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Sample records for involving genetic modification

  1. Genetic modification and genetic determinism

    PubMed Central

    Resnik, David B; Vorhaus, Daniel B

    2006-01-01

    In this article we examine four objections to the genetic modification of human beings: the freedom argument, the giftedness argument, the authenticity argument, and the uniqueness argument. We then demonstrate that each of these arguments against genetic modification assumes a strong version of genetic determinism. Since these strong deterministic assumptions are false, the arguments against genetic modification, which assume and depend upon these assumptions, are therefore unsound. Serious discussion of the morality of genetic modification, and the development of sound science policy, should be driven by arguments that address the actual consequences of genetic modification for individuals and society, not by ones propped up by false or misleading biological assumptions. PMID:16800884

  2. [Advances in genetic modification technologies].

    PubMed

    Zhang, Baixue; Sun, Qixin; Li, Haifeng

    2015-08-01

    Genetic modification technology is a new molecular tool for targeted genome modification. It includes zinc finger nucleases (ZFN) technology, transcription activator-like effector nucleases (TALEN) technology and clustered regularly interspaced short palindromic repeat (CRISPR)-associated (Cas) (CRISPR-Cas) nucleases technology. All of these nucleases create DNA double-strand breaks (DSB) at chromosomal targeted sites and induce cell endogenous mechanisms that are primarily repaired by the non-homologous end joining (NHEJ) or homologous recombination (HR) pathway, resulting in targeted endogenous gene knock-out or exogenous gene insertion. In recent years, genetic modification technologies have been successfully applied to bacteria, yeast, human cells, fruit fly, zebra fish, mouse, rat, livestock, cynomolgus monkey, Arabidopsis, rice, tobacco, maize, sorghum, wheat, barley and other organisms, showing its enormous advantage in gene editing field. Especially, the newly developed CRISPR-Cas9 system arose more attention because of its low cost, high effectiveness, simplicity and easiness. We reviewed the principles and the latest research progress of these three technologies, as well as prospect of future research and applications.

  3. Genetic Modification of Preimplantation Embryos: Toward Adequate Human Research Policies

    PubMed Central

    Dresser, Rebecca

    2004-01-01

    Citing advances in transgenic animal research and setbacks in human trials of somatic cell genetic interventions, some scientists and others want to begin planning for research involving the genetic modification of human embryos. Because this form of genetic modification could affect later-born children and their offspring, the protection of human subjects should be a priority in decisions about whether to proceed with such research. Yet because of gaps in existing federal policies, embryo modification proposals might not receive adequate scientific and ethical scrutiny. This article describes current policy shortcomings and recommends policy actions designed to ensure that the investigational genetic modification of embryos meets accepted standards for research on human subjects. PMID:15016248

  4. Genetic Modification of Stem Cells for Transplantation

    PubMed Central

    Phillips, M. Ian; Tang, Yao Liang

    2009-01-01

    Gene modification of cells for prior to their transplantation, especially stem cells, enhances their survival and increases their function in cell therapy. Like the Trojan horse, the gene modified cell has to gain entrance inside the host’s walls and survive and deliver its transgene products Using cellular, molecular and gene manipulation techniques the transplanted cell can be protected in a hostile environment from immune rejection, inflammation, hypoxia and apoptosis. Genetic engineering to modify cells involves constructing modules of functional gene sequences. They can be simple reporter genes or complex cassettes with gene switches, cell specific promoters and multiple transgenes. We discuss methods to deliver and construct gene cassettes with viral and non viral delivery, siRNA, and conditional Cre/Lox P. We review the current uses of gene modified stem cells in cardiovascular disease, diabetes, neurological diseases,( including Parkinson’s, Alzheimer’s and spinal cord injury repair), bone defects, hemophilia, and cancer. PMID:18031863

  5. Reproductive cloning combined with genetic modification.

    PubMed

    Strong, C

    2005-11-01

    Although there is widespread opposition to reproductive cloning, some have argued that its use by infertile couples to have genetically related children would be ethically justifiable. Others have suggested that lesbian or gay couples might wish to use cloning to have genetically related children. Most of the main objections to human reproductive cloning are based on the child's lack of unique nuclear DNA. In the future, it may be possible safely to create children using cloning combined with genetic modifications, so that they have unique nuclear DNA. The genetic modifications could be aimed at giving such children genetic characteristics of both members of the couple concerned. Thus, cloning combined with genetic modification could be appealing to infertile, lesbian, or gay couples who seek genetically related children who have genetic characteristics of both members. In such scenarios, the various objections to human reproductive cloning that are based on the lack of genetic uniqueness would no longer be applicable. The author argues that it would be ethically justifiable for such couples to create children in this manner, assuming these techniques could be used safely.

  6. [Impact of genetic modifications on infectious diseases].

    PubMed

    Houdebine, L-M

    2004-11-01

    Genetic engineering offers the theoretical possibility to transfer any natural or modified gene into any living organism. This generates new and diverse situations which may contribute to the spreading of infectious diseases or on the contrary to control them. Problems may theoretically come from uncontrolled genes providing resistance to antibiotics, from the activation of genomic retroviral sequences, from enhanced sensitivity of the organism to pathogens, as well as from the generation of mutated microorganisms with a higher pathogenecity. On the contrary, various genetic modifications may create organisms resistant to infectious diseases, generate safe and efficient recombinant vaccines, or provide patients with proteins which stimulate their defense mechanisms. The major impacts of genetic modifications in the development of infectious diseases or on the contrary in their eradication are analyzed in this article.

  7. Epigenetics: Beyond Chromatin Modifications and Complex Genetic Regulation1

    PubMed Central

    Eichten, Steven R.; Schmitz, Robert J.; Springer, Nathan M.

    2014-01-01

    Chromatin modifications and epigenetics may play important roles in many plant processes, including developmental regulation, responses to environmental stimuli, and local adaptation. Chromatin modifications describe biochemical changes to chromatin state, such as alterations in the specific type or placement of histones, modifications of DNA or histones, or changes in the specific proteins or RNAs that associate with a genomic region. The term epigenetic is often used to describe a variety of unexpected patterns of gene regulation or inheritance. Here, we specifically define epigenetics to include the key aspects of heritability (stable transmission of gene expression states through mitotic or meiotic cell divisions) and independence from DNA sequence changes. We argue against generically equating chromatin and epigenetics; although many examples of epigenetics involve chromatin changes, those chromatin changes are not always heritable or may be influenced by genetic changes. Careful use of the terms chromatin modifications and epigenetics can help separate the biochemical mechanisms of regulation from the inheritance patterns of altered chromatin states. Here, we also highlight examples in which chromatin modifications and epigenetics affect important plant processes. PMID:24872382

  8. [Biotechnology, especially genetic modification, and legislation].

    PubMed

    de Sitter, H; Peters, P W J

    2002-05-15

    Biotechnology and genetic modification (GM) related legislation is not yet fully developed in the European Union (EU). New legislation has been recently issued ('Introduction of GMO's in the environment') and recently proposals from the European Commission ('GMO's in food and feed' and 'Traceability and labelling of GMO's') entered the decision-making process in the end of 2001. The proposals for the establishment of the European Food Authority play a role in this respect. GMO legislation is complex not in the least because of the demands for the dossiers, to be submitted with an application, while these procedures for admission must become more transparent. In this paper the relevant legislation will be discussed with the exception of that related to human health. Because of dissatisfaction with the present legislation, the European Commission in the past years granted no new approvals for introductions on the market of GMO's and for GM novel foods. New legislation should suspend the present de-facto moratorium. The tasks and position of the Inspectorate for the Health Protection and Veterinary Public Health is discussed. A provision has been made in the legislation with respect to adventitious or technically unavoidable contamination of raw materials with GMO's up to a maximum of 1%, of which the enforcement is not yet watertight. The analytical methods are being still developed.

  9. Genetic disorders involving adrenal development.

    PubMed

    Lin, Lin; Ferraz-de-Souza, Bruno; Achermann, John C

    2007-01-01

    The past decade has seen significant advances in our understanding of the genetic aetiology of several forms of adrenal failure that present in infancy or childhood. Several of these disorders affect adrenal development and are termed 'adrenal hypoplasia'. These conditions can be broadly divided into: (1) secondary forms of adrenal hypoplasia due to panhypopituitarism (e.g. HESX1, LHX4, SOX3) or abnormalities in ACTH synthesis (TPIT) or processing (e.g. POMC or PC1); (2) adrenal hypoplasia as part of an ACTH resistance syndrome [MC2R/ACTH receptor, MRAP, AAAS (triple A syndrome)], and (3) primary defects in the development of the adrenal gland itself (primary adrenal hypoplasia). Primary adrenal hypoplasia most commonly occurs in an X-linked form due to mutations in the nuclear receptor DAX1 (NR0B1) but can occur in a poorly understood recessive form or as part of the IMAGe (intrauterine growth retardation, metaphyseal dysplasia, adrenal hypoplasia, genitourinary anomalies) syndrome. Defining the molecular basis of these conditions can have significant clinical implications for management, counselling and presymptomatic diagnosis, as well as providing fascinating insight into normal and abnormal mechanisms of adrenal development in humans.

  10. Genetic Engineering: The Modification of Man

    ERIC Educational Resources Information Center

    Sinsheimer, Robert L.

    1970-01-01

    Describes somatic and genetic manipulations of individual genotypes, using diabetes control as an example of the first mode that is potentially realizable be derepression or viral transduction of genes. Advocates the use of genetic engineering of the second mode to remove man from his biological limitations, but offers maxims to ensure the…

  11. Biochemical And Genetic Modification Of Polysaccharides

    NASA Technical Reports Server (NTRS)

    Kern, Roger G.; Petersen, Gene R.; Richards, Gil F.

    1993-01-01

    Bacteriophages producing endopolysaccharase-type enzymes used to produce, isolate, and purify high yields of modified polysaccharides from polysaccharides produced by, and incorporated into capsules of, certain bacteria. Bacteriophages used in conversion of native polysaccharide materials into polymers of nearly uniform high molecular weight or, alternatively, into highly pure oligosaccharides. Also used in genetic selection of families of polysaccharides structurally related to native polysaccharide materials, but having altered properties. Resulting new polysaccharides and oligosaccharides prove useful in variety of products, including pharmaceutical chemicals, coating materials, biologically active carbohydrates, and drag-reducing additives for fluids.

  12. Infectious diseases: Surveillance, genetic modification and simulation

    USGS Publications Warehouse

    Koh, H. L.; Teh, S.Y.; De Angelis, D. L.; Jiang, J.

    2011-01-01

    Infectious diseases such as influenza and dengue have the potential of becoming a worldwide pandemic that may exert immense pressures on existing medical infrastructures. Careful surveillance of these diseases, supported by consistent model simulations, provides a means for tracking the disease evolution. The integrated surveillance and simulation program is essential in devising effective early warning systems and in implementing efficient emergency preparedness and control measures. This paper presents a summary of simulation analysis on influenza A (H1N1) 2009 in Malaysia. This simulation analysis provides insightful lessons regarding how disease surveillance and simulation should be performed in the future. This paper briefly discusses the controversy over the experimental field release of genetically modified (GM) Aedes aegypti mosquito in Malaysia. Model simulations indicate that the proposed release of GM mosquitoes is neither a viable nor a sustainable control strategy. ?? 2011 WIT Press.

  13. Enzyme Technology of Peroxidases: Immobilization, Chemical and Genetic Modification

    NASA Astrophysics Data System (ADS)

    Longoria, Adriana; Tinoco, Raunel; Torres, Eduardo

    An overview of enzyme technology applied to peroxidases is made. Immobilization on organic, inorganic, and hybrid supports; chemical modification of amino acids and heme group; and genetic modification by site-directed and random mutagenesis are included. Different strategies that were carried out to improve peroxidase performance in terms of stability, selectivity, and catalytic activity are analyzed. Immobilization of peroxidases on inorganic and organic materials enhances the tolerance of peroxidases toward the conditions normally found in many industrial processes, such as the presence of an organic solvent and high temperature. In addition, it is shown that immobilization helps to increase the Total Turnover Number at levels high enough to justify the use of a peroxidase-based biocatalyst in a synthesis process. Chemical modification of peroxidases produces modified enzymes with higher thermostability and wider substrate variability. Finally, through mutagenesis approaches, it is possible to produce modified peroxidases capable of oxidizing nonnatural substrates with high catalytic activity and affinity.

  14. The past, present and future of crop genetic modification.

    PubMed

    Fedoroff, Nina V

    2010-11-30

    The introduction of science and technology into agriculture over the past two centuries has markedly increased agricultural productivity and decreased its labor-intensiveness. Chemical fertilization, mechanization, plant breeding and molecular genetic modification (GM) have contributed to unparalleled productivity increases. Future increases are far from assured because of underinvestment in agricultural research, growing population pressure, decreasing fresh water availability, increasing temperatures and societal rejection of GM crops in many countries.

  15. Genetic modifications of pigs for medicine and agriculture.

    PubMed

    Whyte, Jeffrey J; Prather, Randall S

    2011-01-01

    Genetically modified swine hold great promise in the fields of agriculture and medicine. Currently, these swine are being used to optimize production of quality meat, to improve our understanding of the biology of disease resistance, and to reduced waste. In the field of biomedicine, swine are anatomically and physiologically analogous to humans. Alterations of key swine genes in disease pathways provide model animals to improve our understanding of the causes and potential treatments of many human genetic disorders. The completed sequencing of the swine genome will significantly enhance the specificity of genetic modifications, and allow for more accurate representations of human disease based on syntenic genes between the two species. Improvements in both methods of gene alteration and efficiency of model animal production are key to enabling routine use of these swine models in medicine and agriculture.

  16. Genetic modification in organ transplantation and in vivo luciferase imaging

    NASA Astrophysics Data System (ADS)

    Murakami, Takashi; Inoue, Sei-ichiro; Sato, Yuki; Ajiki, Takashi; Ohsawa, Ichiro; Kobayashi, Eiji

    2005-04-01

    The genetic modification for organ transplantation is one of the most promising strategies to regulate allogeneic immune response. Organ-selective gene transfer has especially benefit to control local immune responses. Based on the catheter technique, we tested to deliver naked plasmid DNA to target graft organs of rats (liver and limbs) by a rapid injection (hydrodynamics-based transfection). Recent advances in transplantation have been achieved by visualization of cellular process and delivered gene expression during the inflammatory process by using non-invasive in vivo imaging. Herein, we examined the fate of genetically modified grafts using a firefly luciferase expression plasmid. For liver modification before transplantation, 6.25% of body weight PBS containing plasmid DNA was injected into the liver through the inferior vena cava using a catheter, and the liver was subsequently transplanted to the recipient rat. For limb modification, the femoral caudal epigastric vein was used. In the rat liver transplantation model, substantial luciferase expression was visualized and sustained for only a few days in the grafted liver. We also addressed stress responses by this hydrodynamics procedure using reporter plasmids containing cis-acting enhancer binding site such as NF-kappa B, cAMP, or heat shock response element. In contrast to hepatic transduction, this genetic limb targeting achieved long lasting luciferase expression in the muscle for 2 months or more. Thus, our results suggest that this catheter-based in vivo transfection technique provides an effective strategy for organ-selective gene modification in transplantation, and the bioluminescent imaging is broadening its potential for evaluation to various preclinical studies.

  17. Protein modifications involved in neurotransmitter and gasotransmitter signaling

    PubMed Central

    Sen, Nilkantha; Snyder, Solomon H.

    2010-01-01

    Covalent modifications of intracellular proteins, such as phosphorylation, are generally thought to occur as secondary or tertiary responses to neurotransmitters, following the intermediation of membrane receptors and second messengers such as cyclic AMP. By contrast, the gasotransmitter nitric oxide directly S-nitrosylates cysteine residues in diverse intracellular proteins. Recently, hydrogen sulfide has been acknowledged as a gaso-transmitter, which analogously sulfhydrates cysteine residues in proteins. Cysteine residues are also modified by palmitoylation in response to neurotransmitter signaling, possibly in reciprocity with S-nitrosylation. Neurotransmission also elicits sumoylation and acetylation of lysine residues within diverse proteins. This review addresses how these recently appreciated protein modifications impact our thinking about ways in which neurotransmission regulates intracellular protein disposition. PMID:20843563

  18. Interactions of Dnd proteins involved in bacterial DNA phosphorothioate modification

    PubMed Central

    Xiong, Wei; Zhao, Gong; Yu, Hao; He, Xinyi

    2015-01-01

    DNA phosphorothioation (PT) is the first discovered physiological DNA backbone modification, in which a non-bridging oxygen atom of the phosphodiester bond is replaced with a sulfur atom in Rp (rectus for plane) configuration. PT modification is governed by a highly conserved gene cluster dndA/iscS-dndBCDE that is widespread across bacterial and archaeal species. However, little is known about how these proteins coordinately react with each other to perform oxygen–sulfur swap. We here demonstrated that IscS, DndC, DndD and DndE form a protein complex of which the molecular ratio for four proteins in the complex is approximate 1:1:1:1. DndB here displayed little or weak affinity to the complex and the constructs harboring dndACDE can confer the host in vivo PT modification. Using co-purification and pull down strategy, we demonstrated that the four proteins assemble into a pipeline in collinear to its gene organization, namely, IscS binding to DndC, DndC binding to DndD, and DndD binding to DndE. Moreover, weak interactions between DndE and IscS, DndE and DndC were also identified. PMID:26539172

  19. Childhood obesity: are genetic differences involved?

    PubMed

    Bouchard, Claude

    2009-05-01

    This brief review focuses on the genetic contribution to childhood obesity. Evidence for a genetic component to excess body weight during growth is presented from the perspective of genetic epidemiology studies. Parental obesity is a predictor of childhood excess weight. The familial risk ratio for childhood obesity when a parent is obese reaches >2.5. Birth weight is characterized by a genetic heritability component on the order of 30%, with significant maternal and paternal effects in addition to the newborn genes. About 5% of childhood obesity cases are caused by a defect that impairs function in a gene, and >/=5 of these genes have been uncovered. However, the common forms of childhood obesity seem to result from a predisposition that primarily favors obesogenic behaviors in an obesogenic environment. Candidate gene and genomewide association studies reveal that these obesogenic genes have small effect sizes but that the risk alleles for obesity are quite common in populations. The latter may translate into a highly significant population-attributable risk of obesity. Gene-environment interaction studies suggest that the effects of predisposing genes can be enhanced or diminished by exposure to relevant behaviors. It is possible that the prevalence of childhood obesity is increasing across generations as a result of positive assortative mating with obese husbands and wives contributing more obese offspring than normal-weight parents.

  20. Human Handedness: More Evidence for Genetic Involvement.

    ERIC Educational Resources Information Center

    Longstreth, Langdon E.

    1980-01-01

    A series of environmental-genetical analyses of the left-handedness of 1,950 college students indicates that left-handedness is familial: it is more frequent in families in which at least one parent is left-handed. (Author/CM)

  1. Advances in genetic modification of pluripotent stem cells.

    PubMed

    Fontes, Andrew; Lakshmipathy, Uma

    2013-11-15

    Genetically engineered stem cells aid in dissecting basic cell function and are valuable tools for drug discovery, in vivo cell tracking, and gene therapy. Gene transfer into pluripotent stem cells has been a challenge due to their intrinsic feature of growing in clusters and hence not amenable to common gene delivery methods. Several advances have been made in the rapid assembly of DNA elements, optimization of culture conditions, and DNA delivery methods. This has lead to the development of viral and non-viral methods for transient or stable modification of cells, albeit with varying efficiencies. Most methods require selection and clonal expansion that demand prolonged culture and are not suited for cells with limited proliferative potential. Choosing the right platform based on preferred length, strength, and context of transgene expression is a critical step. Random integration of the transgene into the genome can be complicated due to silencing or altered regulation of expression due to genomic effects. An alternative to this are site-specific methods that target transgenes followed by screening to identify the genomic loci that support long-term expression with stem cell proliferation and differentiation. A highly precise and accurate editing of the genome driven by homology can be achieved using traditional methods as well as the newer technologies such as zinc finger nuclease, TAL effector nucleases and CRISPR. In this review, we summarize the different genetic engineering methods that have been successfully used to create modified embryonic and induced pluripotent stem cells.

  2. QTL involved in the modification of cyanidin compounds in black and red raspberry fruit.

    PubMed

    Bushakra, J M; Krieger, C; Deng, D; Stephens, M J; Allan, A C; Storey, R; Symonds, V V; Stevenson, D; McGhie, T; Chagné, D; Buck, E J; Gardiner, S E

    2013-03-01

    Fruit from Rubus species are highly valued for their flavor and nutritive qualities. Anthocyanin content contributes to these qualities, and although many studies have been conducted to identify and quantify the major anthocyanin compounds from various Rubus species, the genetic control of the accumulation of these complex traits in Rubus is not yet well understood. The identification of the regions of the genome involved in the production of anthocyanins is an important first step in identifying the genes underlying their expression. In this study, ultra and high-performance liquid chromatography (UHPLC and HPLC) and two newly developed Rubus linkage maps were used to conduct QTL analyses to explore the presence of associations between concentrations of five anthocyanins in fruit and genotype. In total, 27 QTL were identified on the Rubus linkage maps, four of which are associated with molecular markers designed from transcription factors and three of which are associated with molecular markers designed from anthocyanin biosynthetic pathway candidate genes. The results of this study suggest that, while QTL for anthocyanin accumulation have been identified on six of seven Rubus linkage groups (RLG), the QTL on RLG2 and RLG7 may be very important for genetic control of cyanidin modification in Rubus.

  3. Structural modification of polysaccharides: A biochemical-genetic approach

    NASA Technical Reports Server (NTRS)

    Kern, Roger G.; Petersen, Gene R.

    1991-01-01

    Polysaccharides have a wide range of industrial and biomedical applications. An industry trend is underway towards the increased use of bacteria to produce polysaccharides. Long term goals of this work are the adaptation and enhancement of saccharide properties for electronic and optic applications. In this report we illustrate the application of enzyme-bearing bacteriophage on strains of the enteric bacterium Klebsiella pneumoniae, which produces a polysaccharide with the relatively rare rheological property of drag-reduction. This has resulted in the production of new polysaccharides with enhanced rheological properties. Our laboratory is developing techniques for processing and structurally modifying bacterial polysaccharides and oligosaccharides which comprise their basic polymeric repeat units. Our research has focused on bacteriophage which produce specific polysaccharide degrading enzymes. This has lead to the development of enzymes generated by bacteriophage as tools for polysaccharide modification and purification. These enzymes were used to efficiently convert the native material to uniform-sized high molecular weight polymers, or alternatively into high-purity oligosaccharides. Enzyme-bearing bacteriophage also serve as genetic selection tools for bacteria that produce new families of polysaccharides with modified structures.

  4. [Unintended effects of genetic modifications in plants and methods of their analysis].

    PubMed

    Sorochyns'kyĭ, B V; Burlaka, O M; Naumenko, V D; Sekan, A S

    2011-01-01

    The problem of unintended effects caused by genetic modification of plants is analysed. Factors that can provoke the unintended effects in genetically engineered plants, their consequences and possibility of the avoiding of unintended effects with use of current methods of genetic modification are discussed. Modern methodological approaches applied to analyse the unintended effects during the safety assessment of transgenic plants, in particular methods of molecular profiling with different "-omic"-technologies are described. PMID:22168051

  5. [Unintended effects of genetic modifications in plants and methods of their analysis].

    PubMed

    Sorochyns'kyĭ, B V; Burlaka, O M; Naumenko, V D; Sekan, A S

    2011-01-01

    The problem of unintended effects caused by genetic modification of plants is analysed. Factors that can provoke the unintended effects in genetically engineered plants, their consequences and possibility of the avoiding of unintended effects with use of current methods of genetic modification are discussed. Modern methodological approaches applied to analyse the unintended effects during the safety assessment of transgenic plants, in particular methods of molecular profiling with different "-omic"-technologies are described.

  6. On the genetic modification of psychology, personality, and behavior.

    PubMed

    Neitzke, Alex B

    2012-12-01

    I argue that the use of heritable modifications for psychology, personality, and behavior should be limited to the reversal or prevention of relatively unambiguous instances of pathology or likely harm (e.g. sociopathy). Most of the likely modifications of psychological personality would not be of this nature, however, and parents therefore should not have the freedom to make such modifications to future children. I argue by examining the viewpoints of both the individual and society. For individuals, modifications would interfere with their capacity for self-determination in a way that undermines the very concept of self-determination. I argue that modification of psychology and personality is unlike present parenting in morally significant ways. For society, modification offers a medium for power to manipulate the makeup of persons and populations, possibly causing biological harm to the species and altering our conceptions of social responsibility.

  7. Attitudes Toward Genetic Modification Research: An Analysis of the Views of the Sputnik Generation.

    ERIC Educational Resources Information Center

    Miller, Jon D.

    1982-01-01

    Utilizing data from the 1977 National Assessment of Educational Progress (NAEP) survey of young adults, summarizes attitudes toward genetic modification research and the demographic, educational, and occupational correlates of these attitudes. (Author/SK)

  8. The impact of genetic modification of human foods in the 21st century: a review.

    PubMed

    Uzogara, S G

    2000-05-01

    Genetic engineering of food is the science which involves deliberate modification of the genetic material of plants or animals. It is an old agricultural practice carried on by farmers since early historical times, but recently it has been improved by technology. Many foods consumed today are either genetically modified (GM) whole foods, or contain ingredients derived from gene modification technology. Billions of dollars in U.S. food exports are realized from sales of GM seeds and crops. Despite the potential benefits of genetic engineering of foods, the technology is surrounded by controversy. Critics of GM technology include consumer and health groups, grain importers from European Union (EU) countries, organic farmers, environmentalists, concerned scientists, ethicists, religious rights groups, food advocacy groups, some politicians and trade protectionists. Some of the specific fears expressed by opponents of GM technology include alteration in nutritional quality of foods, potential toxicity, possible antibiotic resistance from GM crops, potential allergenicity and carcinogenicity from consuming GM foods. In addition, some more general concerns include environmental pollution, unintentional gene transfer to wild plants, possible creation of new viruses and toxins, limited access to seeds due to patenting of GM food plants, threat to crop genetic diversity, religious, cultural and ethical concerns, as well as fear of the unknown. Supporters of GM technology include private industries, research scientists, some consumers, U.S. farmers and regulatory agencies. Benefits presented by proponents of GM technology include improvement in fruit and vegetable shelf-life and organoleptic quality, improved nutritional quality and health benefits in foods, improved protein and carbohydrate content of foods, improved fat quality, improved quality and quantity of meat, milk and livestock. Other potential benefits are: the use of GM livestock to grow organs for transplant

  9. Genetic, environmental, and epigenetic factors involved in CAKUT.

    PubMed

    Nicolaou, Nayia; Renkema, Kirsten Y; Bongers, Ernie M H F; Giles, Rachel H; Knoers, Nine V A M

    2015-12-01

    Congenital anomalies of the kidney and urinary tract (CAKUT) refer to a spectrum of structural renal malformations and are the leading cause of end-stage renal disease in children. The genetic diagnosis of CAKUT has proven to be challenging due to genetic and phenotypic heterogeneity and incomplete genetic penetrance. Monogenic causes of CAKUT have been identified using different approaches, including single gene screening, and gene panel and whole exome sequencing. The majority of the identified mutations, however, lack substantial evidence to support a pathogenic role in CAKUT. Copy number variants or single nucleotide variants that are associated with CAKUT have also been identified. Numerous studies support the influence of epigenetic and environmental factors on kidney development and the natural history of CAKUT, suggesting that the pathogenesis of this syndrome is multifactorial. In this Review we describe the current knowledge regarding the genetic susceptibility underlying CAKUT and the approaches used to investigate the genetic basis of CAKUT. We outline the associated environmental risk factors and epigenetic influences on CAKUT and discuss the challenges and strategies used to fully address the involvement and interplay of these factors in the pathogenesis of the disease. PMID:26281895

  10. [Modification changes of the genetic material in Saccharomyces yeasts].

    PubMed

    Repnevskaia, M V; Kashkin, P K; Inge-Vechtomov, S G

    1989-03-01

    The problem of mating-type switches in heterothallic yeast cells was investigated. In selective system for cytoduction in alpha x alpha crosses alpha-cytoductants were predominantly obtained. Thus matings in alpha x alpha crosses can proceed through non-heritable changes (modifications) of the mating type alpha----a. The frequency of alpha-cytoductants after UV-irradiation of the recipient cells exceeded the control value 50-90 times. The extra copy of MAT alpha dramatically decreased the frequency of cytoductants in alpha x alpha crosses, either spontaneously or after UV-irradiation. The rad18 recipient defective in postreplication repair had 70-times increased level of mating-type modifications, as compared with isogenic Rad+ strain. An explanation consistent with these data is that mating-type modifications are due to phenotypic expression of primary lesions of MAT alpha locus. Such lesions might be expressed as transient a-mating type. After the mating event, these lesions can be repaired or turned to true mutations within the MAT locus. In fact, approximately half of non-mating cytoductants from alpha x alpha crosses had the phenotype of mat alpha 2 mutants.

  11. Transgenic crops: the present state and new ways of genetic modification.

    PubMed

    Szabala, Bartosz M; Osipowski, Pawel; Malepszy, Stefan

    2014-08-01

    Transgenic crops were first commercialised almost 20 years ago, which makes it a good opportunity to reflect on this technology. In this review, we compare its status with the predictions included in Vasil's forecast published in 2002. Our analysis shows that science has provided a wide range of possibilities to modify different traits in plants, yet the economy benefits from that range to very different extents. We also point out the most important constituents of the technology development involving methodology improvement and novel traits expressed in varieties introduced into agriculture. Using native genes (or their elements) in transgenes, accumulating previously produced transgenes to cascade resistance and using herbicide resistance as a selectable marker have been considered typical of novel genetically modified (GM) plant varieties. A vast portion of the novelties in stacked varieties is doubtful in terms of EU regulations. Attention has also been directed to completely novel methodology solutions that hold out the prospect of a more comprehensive use of genetic modification in agriculture as a whole, and, particularly, make its use possible in the EU and even in sustainable agriculture.

  12. Genetic modification of the diarrheal pathogen Cryptosporidium parvum

    PubMed Central

    Vinayak, Sumiti; Pawlowic, Mattie C.; Sateriale, Adam; Brooks, Carrie F.; Studstill, Caleb J.; Bar-Peled, Yael; Cipriano, Michael J.; Striepen, Boris

    2015-01-01

    Recent studies into the global causes of severe diarrhea in young children have identified the protozoan parasite Cryptosporidium as the second most important diarrheal pathogen after rotavirus1–3. Diarrheal disease is estimated to be responsible for 10.5% of overall child mortality4. Cryptosporidium is also an opportunistic pathogen in the context of HIV-AIDS and organ transplantation5,6. There is no vaccine and only a single approved drug that provides no benefit for those in gravest danger, malnourished children and immunocompromised patients7,8. Cryptosporidiosis drug and vaccine development is limited by the poor tractability of the parasite, which includes lack of continuous culture, facile animal models, and molecular genetic tools3,9. Here we describe an experimental framework to genetically modify this important human pathogen. We establish and optimize transfection of C. parvum sporozoites in tissue culture. To isolate stable transgenics we develop a mouse model that delivers sporozoites directly into the intestine, a Cryptosporidium CRISPR/Cas9 system, and in vivo selection for aminoglycoside resistance. We derive reporter parasites suitable for in vitro and in vivo drug screening, and we evaluate the basis of drug susceptibility by gene knock out. We anticipate the ability to genetically engineer the parasite will be transformative for Cryptosporidium research. Genetic reporters will provide quantitative correlates for disease, cure and protection and the role of parasite genes in these processes is now open to rigorous investigation. PMID:26176919

  13. Genetic modification of the diarrhoeal pathogen Cryptosporidium parvum.

    PubMed

    Vinayak, Sumiti; Pawlowic, Mattie C; Sateriale, Adam; Brooks, Carrie F; Studstill, Caleb J; Bar-Peled, Yael; Cipriano, Michael J; Striepen, Boris

    2015-07-23

    Recent studies into the global causes of severe diarrhoea in young children have identified the protozoan parasite Cryptosporidium as the second most important diarrhoeal pathogen after rotavirus. Diarrhoeal disease is estimated to be responsible for 10.5% of overall child mortality. Cryptosporidium is also an opportunistic pathogen in the contexts of human immunodeficiency virus (HIV)-caused AIDS and organ transplantation. There is no vaccine and only a single approved drug that provides no benefit for those in gravest danger: malnourished children and immunocompromised patients. Cryptosporidiosis drug and vaccine development is limited by the poor tractability of the parasite, which includes a lack of systems for continuous culture, facile animal models, and molecular genetic tools. Here we describe an experimental framework to genetically modify this important human pathogen. We established and optimized transfection of C. parvum sporozoites in tissue culture. To isolate stable transgenics we developed a mouse model that delivers sporozoites directly into the intestine, a Cryptosporidium clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 system, and in vivo selection for aminoglycoside resistance. We derived reporter parasites suitable for in vitro and in vivo drug screening, and we evaluated the basis of drug susceptibility by gene knockout. We anticipate that the ability to genetically engineer this parasite will be transformative for Cryptosporidium research. Genetic reporters will provide quantitative correlates for disease, cure and protection, and the role of parasite genes in these processes is now open to rigorous investigation. PMID:26176919

  14. Restriction-Modification Systems as a Barrier for Genetic Manipulation of Staphylococcus aureus.

    PubMed

    Sadykov, Marat R

    2016-01-01

    Genetic manipulation is a powerful approach to study fundamental aspects of bacterial physiology, metabolism, and pathogenesis. Most Staphylococcus aureus strains are remarkably difficult to genetically manipulate as they possess strong host defense mechanisms that protect bacteria from cellular invasion by foreign DNA. In S. aureus these bacterial "immunity" mechanisms against invading genomes are mainly associated with restriction-modification systems. To date, prokaryotic restriction-modification systems are classified into four different types (Type I-IV), all of which have been found in the sequenced S. aureus genomes. This chapter describes the roles, classification, mechanisms of action of different types of restriction-modification systems and the recent advances in the biology of restriction and modification in S. aureus.

  15. Impaired methylation modifications of FZD3 alter chromatin accessibility and are involved in congenital hydrocephalus pathogenesis.

    PubMed

    Wang, Li; Shangguan, Shaofang; Chang, Shaoyan; Wang, Zhen; Lu, Xiaolin; Wu, Lihua; Li, Rui; Bao, Yihua; Qiu, Zhiyong; Niu, Bo; Zhang, Ting

    2014-06-20

    Congenital hydrocephalus is heterogeneous in its etiology, and in addition to a genetic component, has been shown to be caused by environmental factors. Until now, however, no methylation alterations of target genes have been connected with congenital hydrocephalus in humans. Frizzled 3(FZD3) is a planar cell polarity (PCP) gene required for PCP signaling. Partial restoration of frizzled 3 activities in FZD3 mutant mice results in hydrocephalus. To analyze the possible roles of epigenetic modifications of the FZD3 gene in congenital hydrocephalus pathogenesis, DNA methylation in the promoter region of FZD3 was assayed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Gene expression and chromatin accessibility were also determined to assess the role of methylation alterations. Our study found methylation levels of the FZD3 gene were increased in congenital hydrocephalus, especially in males (10.57 ± 3.90 vs. 7.08 ± 0.94, p=0.001). Hypermethylation of FZD3 increased congenital hydrocephalus risk, with an odds ratio of 10.125 (p=0.003). Aberrant methylation modification of FZD3 altered both chromatin structure in this region and FZD3 expression levels. Totally, aberrant methylation modification of the FZD3 gene increases the risk of congenital hydrocephalus by altering chromatin structure and disturbing gene expression.

  16. Genetic modification and selection of microorganisms for growth on Mars.

    PubMed

    Hiscox, J A; Thomas, D J

    1995-10-01

    Genetic engineering has often been suggested as a mechanism for improving the survival prospects of terrestrial microoganisms when seeded on Mars. The survival characteristics that these pioneer microorganisms could be endowed with and a variety of mechanisms by which this can be achieved are discussed, together with an overview of some of the potential hurdles that must be overcome. Also, a number of biologically useful properties for these microorganisms are presented that could facilitate the initial human colonisation and ultimately the planetary engineering of Mars.

  17. Genetic modification of plants: significant issues and hurdles to success.

    PubMed

    Day, P R

    1996-04-01

    Transformation and regeneration is routine for many crop plants. A genetically engineered tomato with a longer shelf life at full ripeness was introduced in the United States in 1994, and other soon-to-be-released products, both foods and fibers, incorporate genes for resistance to pests, diseases, and environmentally benign herbicides. Other possibilities are altered plant fats and oils, methionine- and lysine-enhanced grain and legume proteins, plant foods that can deliver immunizing antigens, and other ways of controlling fruit ripening. Food safety concerns include the inadvertent production of toxicants and allergens. Foreign DNA can be introduced into plants by bacterial vectors, direct uptake by protoplasts, and mechanical introduction on metal particles or other materials. Limitations include little or no control of copy number or site of integration of the introduced DNA, dependence on selectable markers for recovery of traits, and inadequate knowledge of how to control key metabolic steps to maximize desirable traits. Directed genetic change still requires conventional crop breeding to deliver benefits to farmers and consumers.

  18. Protein kinase C is involved in the regulation of several calreticulin posttranslational modifications.

    PubMed

    Cristina Castañeda-Patlán, M; Razo-Paredes, Roberto; Carrisoza-Gaytán, Rolando; González-Mariscal, Lorenza; Robles-Flores, Martha

    2010-01-01

    Calreticulin (CRT) is a highly versatile lectin-like chaperone that affects many cellular functions both inside and outside the endoplasmic reticulum lumen. We previously reported that calreticulin interacts with several protein kinase C isozymes both in vitro and in vivo. The aim of this study was to elucidate the molecular determinants involved in the association between these proteins and the biochemical significance of their interaction. Using full-length or CRT-domain constructs expressed as GST-fusion proteins, we found that protein kinase C binds to the CRT N domain in overlay and pull-down assays. Phosphorylation experiments showed that only this CRT domain is phosphorylated by the kinase. Lectin blot analysis demonstrated that CRT is modified by N-glycosylation, but this modification did not affect its interaction with protein kinase C. We also demonstrated that although both domains of protein kinase C theta can bind to CRT, it is the catalytic one that binds with higher affinity to CRT. Immunofluorescence studies showed that CRT and PKC co-localize mainly at the ER (estimated in 35%). Activation of protein kinase C induced caused transient changes in CRT localization, and unexpectedly, also induced changes in posttranslational modifications found in the protein: CRT N-glycosylation is abolished, whereas tyrosine phosphorylation and O-linked beta-N-acetylglucosamine modification are increased. Together, these findings suggest that protein kinase C is involved in the regulation of CRT function. PMID:19800981

  19. Postpartum depression: A systematic review of the genetics involved

    PubMed Central

    Couto, Tiago Castro e; Brancaglion, Mayra Yara Martins; Alvim-Soares, António; Moreira, Lafaiete; Garcia, Frederico Duarte; Nicolato, Rodrigo; Aguiar, Regina Amélia Lopes P; Leite, Henrique Vitor; Corrêa, Humberto

    2015-01-01

    Postpartum depression is one of the most prevalent psychopathologies. Its prevalence is estimated to be between 10% and 15%. Despite its multifactorial etiology, it is known that genetics play an important role in the genesis of this disorder. This paper reviews epidemiological evidence supporting the role of genetics in postpartum depression (PPD). The main objectives of this review are to determine which genes and polymorphisms are associated with PPD and discuss how this association may occur. In addition, this paper explores whether these genes are somehow related to or even the same as those linked to Major Depression (MD). To identify gaps in the current knowledge that require investigation, a systematic review was conducted in the electronic databases PubMed, LILACS and SciELO using the index terms “postpartum depression” and “genetics”. Literature searches for articles in peer-reviewed journals were made until April 2014. PPD was indexed 56 times with genetics. The inclusion criteria were articles in Portuguese, Spanish or English that were available by institutional means or sent by authors upon request; this search resulted in 20 papers. Genes and polymorphisms traditionally related to MD, which are those involved in the serotonin, catecholamine, brain-derived neurotrophic factor and tryptophan metabolism, have been the most studied, and some have been related to PPD. The results are conflicting and some depend on epigenetics, which makes the data incipient. Further studies are required to determine the genes that are involved in PPD and establish the nature of the relationship between these genes and PPD. PMID:25815259

  20. Prospects for reducing fumonisin contamination of maize through genetic modification.

    PubMed Central

    Duvick, J

    2001-01-01

    Fumonisins (FB) are mycotoxins found in (italic)Fusarium verticillioides-infected maize grain worldwide. Attention has focused on FBs because of their widespread occurrence, acute toxicity to certain livestock, and their potential carcinogenicity. FBs are present at low levels in most field-grown maize but may spike to high levels depending on both the environment and genetics of the host plant. Among the strategies for reducing risk of FB contamination in maize supplied to the market, development and deployment of Fusarium ear mold-resistant maize germplasm is a high priority. Breeding for increased ear mold tolerance and reduced mycotoxin levels is being practiced today in both commercial and public programs, but the amount of resistance achievable may be limited due to complicated genetics and/or linkage to undesirable agronomic traits. Molecular markers can be employed to speed up the incorporation of chromosomal regions that have a quantitative effect on resistance (quantitative trait loci). Transgenic approaches to ear mold/mycotoxin resistance are now feasible as well. These potentially include genetically enhanced resistance to insect feeding, increased fungal resistance, and detoxification/prevention of mycotoxins in the grain. An example of the first of these approaches is already on the market, namely transgenic maize expressing Bacillus thuringiensis (Bt) toxin, targeted to the European corn borer. Some Bt maize hybrids have the potential to reduce FB levels in field-harvested grain, presumably through reduced feeding of Bt-susceptible insects in ear tissues. However, improved ear mold resistance per se is still an important goal, as the plant will still be vulnerable to noninsect routes of entry to (italic)Fusarium. A second approach, transgene-mediated control of the ability of Fusarium to infect and colonize the ear, could potentially be achieved through overexpression of specific antifungal proteins and metabolites, or enhancement of the plant's own

  1. Prospects for reducing fumonisin contamination of maize through genetic modification.

    PubMed

    Duvick, J

    2001-05-01

    Fumonisins (FB) are mycotoxins found in (italic)Fusarium verticillioides-infected maize grain worldwide. Attention has focused on FBs because of their widespread occurrence, acute toxicity to certain livestock, and their potential carcinogenicity. FBs are present at low levels in most field-grown maize but may spike to high levels depending on both the environment and genetics of the host plant. Among the strategies for reducing risk of FB contamination in maize supplied to the market, development and deployment of Fusarium ear mold-resistant maize germplasm is a high priority. Breeding for increased ear mold tolerance and reduced mycotoxin levels is being practiced today in both commercial and public programs, but the amount of resistance achievable may be limited due to complicated genetics and/or linkage to undesirable agronomic traits. Molecular markers can be employed to speed up the incorporation of chromosomal regions that have a quantitative effect on resistance (quantitative trait loci). Transgenic approaches to ear mold/mycotoxin resistance are now feasible as well. These potentially include genetically enhanced resistance to insect feeding, increased fungal resistance, and detoxification/prevention of mycotoxins in the grain. An example of the first of these approaches is already on the market, namely transgenic maize expressing Bacillus thuringiensis (Bt) toxin, targeted to the European corn borer. Some Bt maize hybrids have the potential to reduce FB levels in field-harvested grain, presumably through reduced feeding of Bt-susceptible insects in ear tissues. However, improved ear mold resistance per se is still an important goal, as the plant will still be vulnerable to noninsect routes of entry to (italic)Fusarium. A second approach, transgene-mediated control of the ability of Fusarium to infect and colonize the ear, could potentially be achieved through overexpression of specific antifungal proteins and metabolites, or enhancement of the plant's own

  2. Targeted genetic modification of cell lines for recombinant protein production

    PubMed Central

    Piskareva, Olga; Muniyappa, Mohan

    2007-01-01

    Considerable increases in productivity have been achieved in biopharmaceutical production processes over the last two decades. Much of this has been a result of improvements in media formulation and process development. Though advances have been made in cell line development, there remains considerable opportunity for improvement in this area. The wealth of transcriptional and proteomic data being generated currently hold the promise of specific molecular interventions to improve the performance of production cell lines in the bioreactor. Achieving this—particularly for multi-gene modification—will require specific, targeted and controlled genetic manipulation of these cells. This review considers some of the current and potential future techniques that might be employed to realise this goal. PMID:19003191

  3. Are You Ready for [a] Roundup?--What Chemistry Has to Do with Genetic Modifications

    NASA Astrophysics Data System (ADS)

    Pöpping, Bert

    2001-06-01

    Genetically modified crops are grown in most parts of the world nowadays. These transgenic plants have new properties such as herbicide tolerance or insect resistance that often cannot be introduced by conventional breeding. Using examples of very common transgenic varieties, the article explains how the knowledge of metabolic pathways and genetic information is used to design these plants and how the same knowledge is used to detect them. It reviews why detection of genetic modifications in plants has become necessary and describes the most common detection methods, from immunological assays to polymerase chain reaction and real-time detection.

  4. Linking the genetic architecture of cytosine modifications with human complex traits

    PubMed Central

    Zhang, Xu; Moen, Erika L.; Liu, Cong; Mu, Wenbo; Gamazon, Eric R.; Delaney, Shannon M.; Wing, Claudia; Godley, Lucy A.; Dolan, M. Eileen; Zhang, Wei

    2014-01-01

    Interindividual variation in cytosine modifications could contribute to heterogeneity in disease risks and other complex traits. We assessed the genetic architecture of cytosine modifications at 283 540 CpG sites in lymphoblastoid cell lines (LCLs) derived from independent samples of European and African descent. Our study suggests that cytosine modification variation was primarily controlled in local by single major modification quantitative trait locus (mQTL) and additional minor loci. Local genetic epistasis was detectable for a small proportion of CpG sites, which were enriched by more than 9-fold for CpG sites mapped to population-specific mQTL. Genetically dependent CpG sites whose modification levels negatively (repressive sites) or positively (facilitative sites) correlated with gene expression levels significantly co-localized with transcription factor binding, with the repressive sites predominantly associated with active promoters whereas the facilitative sites rarely at active promoters. Genetically independent repressive or facilitative sites preferentially modulated gene expression variation by influencing local chromatin accessibility, with the facilitative sites primarily antagonizing H3K27me3 and H3K9me3 deposition. In comparison with expression quantitative trait loci (eQTL), mQTL detected from LCLs were enriched in associations for a broader range of disease categories including chronic inflammatory, autoimmune and psychiatric disorders, suggesting that cytosine modification variation, while possesses a degree of cell linage specificity, is more stably inherited over development than gene expression variation. About 11% of unique single-nucleotide polymorphisms reported in the Genome-Wide Association Study Catalog were annotated, 78% as mQTL and 31% as eQTL in LCLs, which covered 37% of the investigated diseases/traits and provided insights to the biological mechanisms. PMID:24943591

  5. Air pollution and diabetes association: Modification by type 2 diabetes genetic risk score.

    PubMed

    Eze, Ikenna C; Imboden, Medea; Kumar, Ashish; von Eckardstein, Arnold; Stolz, Daiana; Gerbase, Margaret W; Künzli, Nino; Pons, Marco; Kronenberg, Florian; Schindler, Christian; Probst-Hensch, Nicole

    2016-09-01

    Exposure to ambient air pollution (AP) exposure has been linked to type 2 diabetes (T2D) risk. Evidence on the impact of T2D genetic variants on AP susceptibility is lacking. Compared to single variants, joint genetic variants contribute substantially to disease risk. We investigated the modification of AP and diabetes association by a genetic risk score (GRS) covering 63 T2D genes in 1524 first follow-up participants of the Swiss cohort study on air pollution and lung and heart diseases in adults. Genome-wide data and covariates were available from a nested asthma case-control study design. AP was estimated as 10-year mean residential particulate matter <10μm (PM10). We computed count-GRS and weighted-GRS, and applied PM10 interaction terms in mixed logistic regressions, on odds of diabetes. Analyses were stratified by pathways of diabetes pathology and by asthma status. Diabetes prevalence was 4.6% and mean exposure to PM10 was 22μg/m(3). Odds of diabetes increased by 8% (95% confidence interval: 2, 14%) per T2D risk allele and by 35% (-8, 97%) per 10μg/m(3) exposure to PM10. We observed a positive interaction between PM10 and count-GRS on diabetes [ORinteraction=1.10 (1.01, 1.20)], associations being strongest among participants at the highest quartile of count-GRS [OR: 1.97 (1.00, 3.87)]. Stronger interactions were observed with variants of the GRS involved in insulin resistance [(ORinteraction=1.22 (1.00, 1.50)] than with variants related to beta-cell function. Interactions with count-GRS were stronger among asthma cases. We observed similar results with weighted-GRS. Five single variants near GRB14, UBE2E2, PTPRD, VPS26A and KCNQ1 showed nominally significant interactions with PM10 (P<0.05). Our results suggest that genetic risk for T2D may modify susceptibility to air pollution through alterations in insulin sensitivity. These results need confirmation in diabetes cohort consortia.

  6. Air pollution and diabetes association: Modification by type 2 diabetes genetic risk score.

    PubMed

    Eze, Ikenna C; Imboden, Medea; Kumar, Ashish; von Eckardstein, Arnold; Stolz, Daiana; Gerbase, Margaret W; Künzli, Nino; Pons, Marco; Kronenberg, Florian; Schindler, Christian; Probst-Hensch, Nicole

    2016-09-01

    Exposure to ambient air pollution (AP) exposure has been linked to type 2 diabetes (T2D) risk. Evidence on the impact of T2D genetic variants on AP susceptibility is lacking. Compared to single variants, joint genetic variants contribute substantially to disease risk. We investigated the modification of AP and diabetes association by a genetic risk score (GRS) covering 63 T2D genes in 1524 first follow-up participants of the Swiss cohort study on air pollution and lung and heart diseases in adults. Genome-wide data and covariates were available from a nested asthma case-control study design. AP was estimated as 10-year mean residential particulate matter <10μm (PM10). We computed count-GRS and weighted-GRS, and applied PM10 interaction terms in mixed logistic regressions, on odds of diabetes. Analyses were stratified by pathways of diabetes pathology and by asthma status. Diabetes prevalence was 4.6% and mean exposure to PM10 was 22μg/m(3). Odds of diabetes increased by 8% (95% confidence interval: 2, 14%) per T2D risk allele and by 35% (-8, 97%) per 10μg/m(3) exposure to PM10. We observed a positive interaction between PM10 and count-GRS on diabetes [ORinteraction=1.10 (1.01, 1.20)], associations being strongest among participants at the highest quartile of count-GRS [OR: 1.97 (1.00, 3.87)]. Stronger interactions were observed with variants of the GRS involved in insulin resistance [(ORinteraction=1.22 (1.00, 1.50)] than with variants related to beta-cell function. Interactions with count-GRS were stronger among asthma cases. We observed similar results with weighted-GRS. Five single variants near GRB14, UBE2E2, PTPRD, VPS26A and KCNQ1 showed nominally significant interactions with PM10 (P<0.05). Our results suggest that genetic risk for T2D may modify susceptibility to air pollution through alterations in insulin sensitivity. These results need confirmation in diabetes cohort consortia. PMID:27281273

  7. An Efficient and Versatile System for Visualization and Genetic Modification of Dopaminergic Neurons in Transgenic Mice

    PubMed Central

    Kramer, Edgar R.

    2015-01-01

    Background & Aims The brain dopaminergic (DA) system is involved in fine tuning many behaviors and several human diseases are associated with pathological alterations of the DA system such as Parkinson’s disease (PD) and drug addiction. Because of its complex network integration, detailed analyses of physiological and pathophysiological conditions are only possible in a whole organism with a sophisticated tool box for visualization and functional modification. Methods & Results Here, we have generated transgenic mice expressing the tetracycline-regulated transactivator (tTA) or the reverse tetracycline-regulated transactivator (rtTA) under control of the tyrosine hydroxylase (TH) promoter, TH-tTA (tet-OFF) and TH-rtTA (tet-ON) mice, to visualize and genetically modify DA neurons. We show their tight regulation and efficient use to overexpress proteins under the control of tet-responsive elements or to delete genes of interest with tet-responsive Cre. In combination with mice encoding tet-responsive luciferase, we visualized the DA system in living mice progressively over time. Conclusion These experiments establish TH-tTA and TH-rtTA mice as a powerful tool to generate and monitor mouse models for DA system diseases. PMID:26291828

  8. Menkes' kinky hair syndrome: a genetic disease involving copper.

    PubMed

    Holtzman, N A

    1976-09-01

    The kinky hair syndrome (KHS) is an X-linked defect of copper transport in man. An animal model is available in mutants at the X-linked mottled locus in mice. The defect does not involve the uptake of copper from the intestinal lumen but rather the transport of copper from intestinal cells. The reduced activity of several copper-dependent enzymes and the lower copper content of serum, liver, and probably brain account for the manifestations of the disorder which are evident at, or shortly after, birth. Intrauterine involvement is likely but prenatal diagnosis is not yet possible. Although the delivery of iron to the erythropoietic system, and its utilization, are impaired in nutritionally induced copper deficiency, as is neutrophil production, these processes appear normal in KHS. thus, adequate copper to carry them out is available in KHS. While there may be more than one transport system for copper (only one of which is affected in KHS) it is also possible that the hematopoietic tissue in KHS, like the intestinal cells, has abnormally high afficity for copper. The presence of multiple alleles at the KHS locus (and at other genetic loci) in man, which cause different degrees of reduction in copper transport, could account for variations in the susceptibility to copper deficiency observed in infant populations.

  9. Breaking evolution's chains: the prospect of deliberate genetic modification in humans.

    PubMed

    Powell, Russell; Buchanan, Allen

    2011-02-01

    Many philosophers invoke the "wisdom of nature" in arguing for varying degrees of caution in the development and use of genetic enhancement technologies. Because they view natural selection as akin to a master engineer that creates functionally and morally optimal design, these authors tend to regard genetic intervention with suspicion. In Part II, we examine and ultimately reject the evolutionary assumptions that underlie the master engineer analogy (MEA). By highlighting the constraints on ordinary unassisted evolution, we show how intentional genetic modification can overcome many of the natural impediments to the human good. Our contention is that genetic engineering offers a solution that is more eff icient, reliable, versatile, and morally palatable than the lumbering juggernaut of Darwinian evolution. In Part III, we evaluate a recent attempt to ground precautionary enhancement heuristics in adaptive etiology. Our problem with this approach is two-fold: first, it is based on the same "strong adaptationist" interpretation of evolution that motivates the flawed MEA, and second, the etiological concept of function on which it relies provides indirect and potentially misleading information about the likely consequences of genetic intervention. We offer instead enhancement criteria based on causal relationships in ontogeny. We conclude that rather than grounding a presumption against deliberate genetic modification, the causal structure of the living world gives us good moral reason to pursue it.

  10. Genetic modification and screening in rat using haploid embryonic stem cells.

    PubMed

    Li, Wei; Li, Xin; Li, Tianda; Jiang, Ming-Gui; Wan, Haifeng; Luo, Guan-Zheng; Feng, Chunjing; Cui, Xiaolong; Teng, Fei; Yuan, Yan; Zhou, Quan; Gu, Qi; Shuai, Ling; Sha, Jiahao; Xiao, Yamei; Wang, Liu; Liu, Zhonghua; Wang, Xiu-Jie; Zhao, Xiao-Yang; Zhou, Qi

    2014-03-01

    The rat is an important animal model in biomedical research, but practical limitations to genetic manipulation have restricted the application of genetic analysis. Here we report the derivation of rat androgenetic haploid embryonic stem cells (RahESCs) as a tool to facilitate such studies. Our approach is based on removal of the maternal pronucleus from zygotes to generate androgenetic embryos followed by derivation of ESCs. The resulting RahESCs have 21 chromosomes, express pluripotency markers, differentiate into three germ layer cells, and contribute to the germline. Homozygous mutations can be introduced by both large-scale gene trapping and precise gene targeting via homologous recombination or the CRISPR-Cas system. RahESCs can also produce fertile rats after intracytoplasmic injection into oocytes and are therefore able to transmit genetic modifications to offspring. Overall, RahESCs represent a practical tool for functional genetic studies and production of transgenic lines in rat. PMID:24360884

  11. Advances in the Development of Gene-Targeting Vectors to Increase the Efficiency of Genetic Modification.

    PubMed

    Saito, Shinta; Adachi, Noritaka

    2016-01-01

    Gene targeting via homologous recombination, albeit highly inefficient in human cells, is considered a powerful tool for analyzing gene functions. Despite recent progress in the application of artificial nucleases for genome editing, safety issues remain a concern, particularly when genetic modification is used for therapeutic purposes. Therefore, the development of gene-targeting vectors is necessary for safe and sophisticated genetic modification. In this paper, we describe the effect of vector structure on random integration, which is a major obstacle in efficient gene targeting. In addition, we focus on the features of exon-trapping-type gene-targeting vectors, and discuss a novel strategy for negative selection to enhance gene targeting in human cells.

  12. [Predicting genetic modification targets based on metabolic network analysis--a review].

    PubMed

    Li, Peishun; Ma, Hongwu; Zhao, Xueming; Chen, Tao

    2016-01-01

    Construction of artificial cell factory to produce specific compounds of interest needs wild strain to be genetically engineered. In recent years, with the reconstruction of many genome-scale metabolic networks, a number of methods have been proposed based on metabolic network analysis for predicting genetic modification targets that lead to overproduction of compounds of interest. These approaches use constraints of stoichiometry and reaction reversibility in genome-scale models of metabolism and adopt different mathematical algorithms to predict modification targets, and thus can discover new targets that are difficult to find through traditional intuitive methods. In this review, we introduce the principle, merit, demerit and application of various strain optimization methods in detail. The main problems in existing methods and perspectives on this emerging research field are also discussed, aiming to provide guidance to choose the appropriate methods according to different types of products and the reliability of the predicted results. PMID:27363195

  13. Immune modulation by genetic modification of dendritic cells with lentiviral vectors.

    PubMed

    Liechtenstein, Therese; Perez-Janices, Noemi; Bricogne, Christopher; Lanna, Alessio; Dufait, Inès; Goyvaerts, Cleo; Laranga, Roberta; Padella, Antonella; Arce, Frederick; Baratchian, Mehdi; Ramirez, Natalia; Lopez, Natalia; Kochan, Grazyna; Blanco-Luquin, Idoia; Guerrero-Setas, David; Breckpot, Karine; Escors, David

    2013-09-01

    Our work over the past eight years has focused on the use of HIV-1 lentiviral vectors (lentivectors) for the genetic modification of dendritic cells (DCs) to control their functions in immune modulation. DCs are key professional antigen presenting cells which regulate the activity of most effector immune cells, including T, B and NK cells. Their genetic modification provides the means for the development of targeted therapies towards cancer and autoimmune disease. We have been modulating with lentivectors the activity of intracellular signalling pathways and co-stimulation during antigen presentation to T cells, to fine-tune the type and strength of the immune response. In the course of our research, we have found unexpected results such as the surprising immunosuppressive role of anti-viral signalling pathways, and the close link between negative co-stimulation in the immunological synapse and T cell receptor trafficking. Here we review our major findings and put them into context with other published work.

  14. Rice putative methyltransferase gene OsTSD2 is required for root development involving pectin modification

    PubMed Central

    Qu, Lianghuan; Wu, Chunyan; Zhang, Fei; Wu, Yangyang; Fang, Chuanying; Jin, Cheng; Liu, Xianqing; Luo, Jie

    2016-01-01

    Pectin synthesis and modification are vital for plant development, although the underlying mechanisms are still not well understood. Here, we report the functional characterization of the OsTSD2 gene, which encodes a putative methyltransferase in rice. All three independent T-DNA insertion lines of OsTSD2 displayed dwarf phenotypes and serial alterations in different zones of the root. These alterations included abnormal cellular adhesion and schizogenous aerenchyma formation in the meristematic zone, inhibited root elongation in the elongation zone, and higher lateral root density in the mature zone. Immunofluorescence (with LM19) and Ruthenium Red staining of the roots showed that unesterified homogalacturonan (HG) was increased in Ostsd2 mutants. Biochemical analysis of cell wall pectin polysaccharides revealed that both the monosaccharide composition and the uronic acid content were decreased in Ostsd2 mutants. Increased endogenous ABA content and opposite roles performed by ABA and IAA in regulating cellular adhesion in the Ostsd2 mutants suggested that OsTSD2 is required for root development in rice through a pathway involving pectin synthesis/modification. A hypothesis to explain the relationship among OsTSD2, pectin methylesterification, and root development is proposed, based on pectin’s function in regional cell extension/division in a zone-dependent manner. PMID:27497286

  15. Conformational Modification of Serpins Transforms Leukocyte Elastase Inhibitor into an Endonuclease Involved in Apoptosis▿ †

    PubMed Central

    Padron-Barthe, Laura; Leprêtre, Chloé; Martin, Elisabeth; Counis, Marie-France; Torriglia, Alicia

    2007-01-01

    The best-characterized biochemical feature of apoptosis is degradation of genomic DNA into oligonucleosomes. The endonuclease responsible for DNA degradation in caspase-dependent apoptosis is caspase-activated DNase. In caspase-independent apoptosis, different endonucleases may be activated according to the cell line and the original insult. Among the known effectors of caspase-independent cell death, L-DNase II (LEI [leukocyte elastase inhibitor]-derived DNase II) has been previously characterized by our laboratory. We have thus shown that this endonuclease derives from the serpin superfamily member LEI by posttranslational modification (A. Torriglia, P. Perani, J. Y. Brossas, E. Chaudun, J. Treton, Y. Courtois, and M. F. Counis, Mol. Cell. Biol. 18:3612-3619, 1998). In this work, we assessed the molecular mechanism involved in the change in the enzymatic activity of this molecule from an antiprotease to an endonuclease. We report that the cleavage of LEI by elastase at its reactive center loop abolishes its antiprotease activity and leads to a conformational modification that exposes an endonuclease active site and a nuclear localization signal. This represents a novel molecular mechanism for a complete functional conversion induced by changing the conformation of a serpin. We also show that this molecular transformation affects cellular fate and that both endonuclease activity and nuclear translocation of L-DNase II are needed to induce cell death. PMID:17403905

  16. Genetic Modification of Human Pancreatic Progenitor Cells Through Modified mRNA.

    PubMed

    Lu, Song; Chow, Christie C; Zhou, Junwei; Leung, Po Sing; Tsui, Stephen K; Lui, Kathy O

    2016-01-01

    In this chapter, we describe a highly efficient genetic modification strategy for human pancreatic progenitor cells using modified mRNA-encoding GFP and Neurogenin-3. The properties of modified mRNA offer an invaluable platform to drive protein expression, which has broad applicability in pathway regulation, directed differentiation, and lineage specification. This approach can also be used to regulate expression of other pivotal transcription factors during pancreas development and might have potential therapeutic values in regenerative medicine. PMID:27236809

  17. Genetic Modification of Human Pancreatic Progenitor Cells Through Modified mRNA.

    PubMed

    Lu, Song; Chow, Christie C; Zhou, Junwei; Leung, Po Sing; Tsui, Stephen K; Lui, Kathy O

    2016-01-01

    In this chapter, we describe a highly efficient genetic modification strategy for human pancreatic progenitor cells using modified mRNA-encoding GFP and Neurogenin-3. The properties of modified mRNA offer an invaluable platform to drive protein expression, which has broad applicability in pathway regulation, directed differentiation, and lineage specification. This approach can also be used to regulate expression of other pivotal transcription factors during pancreas development and might have potential therapeutic values in regenerative medicine.

  18. Chapter VIII. Contributions of propagation techniques and genetic modification to breeding - genetic engineering for disease resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genetic engineering offers an opportunity to develop flower bulb crops with resistance to fungal, viral, and bacterial pathogens. Several of the flower bulb crops, Lilium spp., Gladiolus, Zantedeschia, Muscari, Hyacinthus, Narcissus, Ornithogalum, Iris, and Alstroemeria, have been transformed with t...

  19. Genetic modification of plant cell walls to enhance biomass yield and biofuel production in bioenergy crops.

    PubMed

    Wang, Yanting; Fan, Chunfen; Hu, Huizhen; Li, Ying; Sun, Dan; Wang, Youmei; Peng, Liangcai

    2016-01-01

    Plant cell walls represent an enormous biomass resource for the generation of biofuels and chemicals. As lignocellulose property principally determines biomass recalcitrance, the genetic modification of plant cell walls has been posed as a powerful solution. Here, we review recent progress in understanding the effects of distinct cell wall polymers (cellulose, hemicelluloses, lignin, pectin, wall proteins) on the enzymatic digestibility of biomass under various physical and chemical pretreatments in herbaceous grasses, major agronomic crops and fast-growing trees. We also compare the main factors of wall polymer features, including cellulose crystallinity (CrI), hemicellulosic Xyl/Ara ratio, monolignol proportion and uronic acid level. Furthermore, the review presents the main gene candidates, such as CesA, GH9, GH10, GT61, GT43 etc., for potential genetic cell wall modification towards enhancing both biomass yield and enzymatic saccharification in genetic mutants and transgenic plants. Regarding cell wall modification, it proposes a novel groove-like cell wall model that highlights to increase amorphous regions (density and depth) of the native cellulose microfibrils, providing a general strategy for bioenergy crop breeding and biofuel processing technology.

  20. Tailored HIV-1 vectors for genetic modification of primary human dendritic cells and monocytes.

    PubMed

    Durand, Stéphanie; Nguyen, Xuan-Nhi; Turpin, Jocelyn; Cordeil, Stephanie; Nazaret, Nicolas; Croze, Séverine; Mahieux, Renaud; Lachuer, Joël; Legras-Lachuer, Catherine; Cimarelli, Andrea

    2013-01-01

    Monocyte-derived dendritic cells (MDDCs) play a key role in the regulation of the immune system and are the target of numerous gene therapy applications. The genetic modification of MDDCs is possible with human immunodeficiency virus type 1 (HIV-1)-derived lentiviral vectors (LVs) but requires high viral doses to bypass their natural resistance to viral infection, and this in turn affects their physiological properties. To date, a single viral protein is able to counter this restrictive phenotype, Vpx, a protein derived from members of the HIV-2/simian immunodeficiency virus SM lineage that counters at least two restriction factors present in myeloid cells. By tagging Vpx with a short heterologous membrane-targeting domain, we have obtained HIV-1 LVs incorporating high levels of this protein (HIV-1-Src-Vpx). These vectors efficiently transduce differentiated MDDCs and monocytes either as previously purified populations or as populations within unsorted peripheral blood mononuclear cells (PBMCs). In addition, these vectors can be efficiently pseudotyped with receptor-specific envelopes, further restricting their cellular tropism almost uniquely to MDDCs. Compared to conventional HIV-1 LVs, these novel vectors allow for an efficient genetic modification of MDDCs and, more importantly, do not cause their maturation or affect their survival, which are unwanted side effects of the transduction process. This study describes HIV-1-Src-Vpx LVs as a novel potent tool for the genetic modification of differentiated MDDCs and of circulating monocyte precursors with strong potential for a wide range of gene therapy applications.

  1. Genetic modification of plant cell walls to enhance biomass yield and biofuel production in bioenergy crops.

    PubMed

    Wang, Yanting; Fan, Chunfen; Hu, Huizhen; Li, Ying; Sun, Dan; Wang, Youmei; Peng, Liangcai

    2016-01-01

    Plant cell walls represent an enormous biomass resource for the generation of biofuels and chemicals. As lignocellulose property principally determines biomass recalcitrance, the genetic modification of plant cell walls has been posed as a powerful solution. Here, we review recent progress in understanding the effects of distinct cell wall polymers (cellulose, hemicelluloses, lignin, pectin, wall proteins) on the enzymatic digestibility of biomass under various physical and chemical pretreatments in herbaceous grasses, major agronomic crops and fast-growing trees. We also compare the main factors of wall polymer features, including cellulose crystallinity (CrI), hemicellulosic Xyl/Ara ratio, monolignol proportion and uronic acid level. Furthermore, the review presents the main gene candidates, such as CesA, GH9, GH10, GT61, GT43 etc., for potential genetic cell wall modification towards enhancing both biomass yield and enzymatic saccharification in genetic mutants and transgenic plants. Regarding cell wall modification, it proposes a novel groove-like cell wall model that highlights to increase amorphous regions (density and depth) of the native cellulose microfibrils, providing a general strategy for bioenergy crop breeding and biofuel processing technology. PMID:27269671

  2. Regulation of genetic flux between bacteria by restriction–modification systems

    PubMed Central

    Touchon, Marie

    2016-01-01

    Restriction–modification (R-M) systems are often regarded as bacteria's innate immune systems, protecting cells from infection by mobile genetic elements (MGEs). Their diversification has been recently associated with the emergence of particularly virulent lineages. However, we have previously found more R-M systems in genomes carrying more MGEs. Furthermore, it has been suggested that R-M systems might favor genetic transfer by producing recombinogenic double-stranded DNA ends. To test whether R-M systems favor or disfavor genetic exchanges, we analyzed their frequency with respect to the inferred events of homologous recombination and horizontal gene transfer within 79 bacterial species. Genetic exchanges were more frequent in bacteria with larger genomes and in those encoding more R-M systems. We created a recognition target motif predictor for Type II R-M systems that identifies genomes encoding systems with similar restriction sites. We found more genetic exchanges between these genomes, independently of their evolutionary distance. Our results reconcile previous studies by showing that R-M systems are more abundant in promiscuous species, wherein they establish preferential paths of genetic exchange within and between lineages with cognate R-M systems. Because the repertoire and/or specificity of R-M systems in bacterial lineages vary quickly, the preferential fluxes of genetic transfer within species are expected to constantly change, producing time-dependent networks of gene transfer. PMID:27140615

  3. Identification of host genes involved in geminivirus infection using a reverse genetics approach.

    PubMed

    Lozano-Durán, Rosa; Rosas-Díaz, Tábata; Luna, Ana P; Bejarano, Eduardo R

    2011-01-01

    Geminiviruses, like all viruses, rely on the host cell machinery to establish a successful infection, but the identity and function of these required host proteins remain largely unknown. Tomato yellow leaf curl Sardinia virus (TYLCSV), a monopartite geminivirus, is one of the causal agents of the devastating Tomato yellow leaf curl disease (TYLCD). The transgenic 2IRGFP N. benthamiana plants, used in combination with Virus Induced Gene Silencing (VIGS), entail an important potential as a tool in reverse genetics studies to identify host factors involved in TYLCSV infection. Using these transgenic plants, we have made an accurate description of the evolution of TYLCSV replication in the host in both space and time. Moreover, we have determined that TYLCSV and Tobacco rattle virus (TRV) do not dramatically influence each other when co-infected in N. benthamiana, what makes the use of TRV-induced gene silencing in combination with TYLCSV for reverse genetic studies feasible. Finally, we have tested the effect of silencing candidate host genes on TYLCSV infection, identifying eighteen genes potentially involved in this process, fifteen of which had never been implicated in geminiviral infections before. Seven of the analyzed genes have a potential anti-viral effect, whereas the expression of the other eleven is required for a full infection. Interestingly, almost half of the genes altering TYLCSV infection play a role in postranslational modifications. Therefore, our results provide new insights into the molecular mechanisms underlying geminivirus infections, and at the same time reveal the 2IRGFP/VIGS system as a powerful tool for functional reverse genetics studies. PMID:21818318

  4. Identification of Host Genes Involved in Geminivirus Infection Using a Reverse Genetics Approach

    PubMed Central

    Luna, Ana P.; Bejarano, Eduardo R.

    2011-01-01

    Geminiviruses, like all viruses, rely on the host cell machinery to establish a successful infection, but the identity and function of these required host proteins remain largely unknown. Tomato yellow leaf curl Sardinia virus (TYLCSV), a monopartite geminivirus, is one of the causal agents of the devastating Tomato yellow leaf curl disease (TYLCD). The transgenic 2IRGFP N. benthamiana plants, used in combination with Virus Induced Gene Silencing (VIGS), entail an important potential as a tool in reverse genetics studies to identify host factors involved in TYLCSV infection. Using these transgenic plants, we have made an accurate description of the evolution of TYLCSV replication in the host in both space and time. Moreover, we have determined that TYLCSV and Tobacco rattle virus (TRV) do not dramatically influence each other when co-infected in N. benthamiana, what makes the use of TRV-induced gene silencing in combination with TYLCSV for reverse genetic studies feasible. Finally, we have tested the effect of silencing candidate host genes on TYLCSV infection, identifying eighteen genes potentially involved in this process, fifteen of which had never been implicated in geminiviral infections before. Seven of the analyzed genes have a potential anti-viral effect, whereas the expression of the other eleven is required for a full infection. Interestingly, almost half of the genes altering TYLCSV infection play a role in postranslational modifications. Therefore, our results provide new insights into the molecular mechanisms underlying geminivirus infections, and at the same time reveal the 2IRGFP/VIGS system as a powerful tool for functional reverse genetics studies. PMID:21818318

  5. mazF as a counter-selectable marker for unmarked genetic modification of Pichia pastoris.

    PubMed

    Yang, Junjie; Jiang, Weihong; Yang, Sheng

    2009-06-01

    In this study, we demonstrate a novel method for unmarked genetic modification of the methylotrophic yeast Pichia pastoris, in which the Escherichia coli toxin gene mazF was used as a counter-selectable marker. mazF was placed under the tightly controlled AOX1 promoter, and the induced expression of MazF in P. pastoris halted cell growth. A modular plasmid was constructed in which mazF and a Zeocin resistance gene acted as counter-selectable and active-selectable markers, respectively, and the MazF-ZeoR cassette was flanked by two direct repeats for marker recycling. Linearized delivery vectors constructed from the modular plasmid were integrated into the P. pastoris genome via homologous recombination, introducing genetic modifications. Upon counter-selection with methanol medium, which induces the AOX1 promoter, the markers were recycled efficiently via homologous recombination between the direct repeats. We used this method successfully to knock-out the ARG1 and MET2 genes, knock-in a green fluorescent protein expression cassette, and perform site-directed mutagenesis on the ARG1 gene, all without introducing unwanted selection markers. The novel method allows repeated use of the selectable marker gene for multiple modifications and will be a useful tool for P. pastoris studies. PMID:19416369

  6. Safety assessment considerations for food and feed derived from plants with genetic modifications that modulate endogenous gene expression and pathways.

    PubMed

    Kier, Larry D; Petrick, Jay S

    2008-08-01

    The current globally recognized comparative food and feed safety assessment paradigm for biotechnology-derived crops is a robust and comprehensive approach for evaluating the safety of both the inserted gene product and the resulting crop. Incorporating many basic concepts from food safety, toxicology, nutrition, molecular biology, and plant breeding, this approach has been used effectively by scientists and regulatory agencies for 10-15 years. Current and future challenges in agriculture include the need for improved yields, tolerance to biotic and abiotic stresses, and improved nutrition. The next generation of biotechnology-derived crops may utilize regulatory proteins, such as transcription factors that modulate gene expression and/or endogenous plant pathways. In this review, we discuss the applicability of the current safety assessment paradigm to biotechnology-derived crops developed using modifications involving regulatory proteins. The growing literature describing the molecular biology underlying plant domestication and conventional breeding demonstrates the naturally occurring genetic variation found in plants, including significant variation in the classes, expression, and activity of regulatory proteins. Specific examples of plant modifications involving insertion or altered expression of regulatory proteins are discussed as illustrative case studies supporting the conclusion that the current comparative safety assessment process is appropriate for these types of biotechnology-developed crops.

  7. Genetic modification of hematopoietic stem cells as a therapy for HIV/AIDS.

    PubMed

    Younan, Patrick; Kowalski, John; Kiem, Hans-Peter

    2013-12-01

    The combination of genetic modification and hematopoietic stem cell (HSC) transplantation may provide the necessary means to develop an alternative treatment option to conventional antiretroviral therapy. As HSCs give rise to all hematopoietic cell types susceptible to HIV infection, modification of HSCs is an ideal strategy for the development of infection-resistant immune cell populations. Although promising results have been obtained in multiple animal models, additional evidence is needed to convincingly demonstrate the feasibility of this approach as a treatment of HIV-1 infected patients. Here, we review the potential of HSC transplantation and the recently identified limitations of this approach. Using the Berlin Patient as a model for a functional cure, we contrast the confines of autologous versus allogeneic transplantation. Finally, we suggest that although autologous, gene-modified HSC-transplantation may significantly reduce plasma viremia, reaching the lower detection limits currently obtainable through daily HAART will remain a challenging endeavor that will require innovative combinatorial therapies.

  8. Two glycosyltransferases involved in anthocyanin modification delineated by transcriptome independent component analysis in Arabidopsis thaliana

    PubMed Central

    Yonekura-Sakakibara, Keiko; Fukushima, Atsushi; Nakabayashi, Ryo; Hanada, Kousuke; Matsuda, Fumio; Sugawara, Satoko; Inoue, Eri; Kuromori, Takashi; Ito, Takuya; Shinozaki, Kazuo; Wangwattana, Bunyapa; Yamazaki, Mami; Saito, Kazuki

    2012-01-01

    To identify candidate genes involved in Arabidopsis flavonoid biosynthesis, we applied transcriptome coexpression analysis and independent component analyses with 1388 microarray data from publicly available databases. Two glycosyltransferases, UGT79B1 and UGT84A2 were found to cluster with anthocyanin biosynthetic genes. Anthocyanin was drastically reduced in ugt79b1 knockout mutants. Recombinant UGT79B1 protein converted cyanidin 3-O-glucoside to cyanidin 3-O-xylosyl(1→2)glucoside. UGT79B1 recognized 3-O-glucosylated anthocyanidins/flavonols and uridine diphosphate (UDP)-xylose, but not 3,5-O-diglucosylated anthocyanidins, indicating that UGT79B1 encodes anthocyanin 3-O-glucoside: 2′′-O-xylosyltransferase. UGT84A2 is known to encode sinapic acid: UDP-glucosyltransferase. In ugt84a2 knockout mutants, a major sinapoylated anthocyanin was drastically reduced. A comparison of anthocyanin profiles in ugt84a knockout mutants indicated that UGT84A2 plays a major role in sinapoylation of anthocyanin, and that other UGT84As contribute the production of 1-O-sinapoylglucose to a lesser extent. These data suggest major routes from cyanidin 3-O-glucoside to the most highly modified cyanidin in the potential intricate anthocyanin modification pathways in Arabidopsis. PMID:21899608

  9. novPTMenzy: a database for enzymes involved in novel post-translational modifications.

    PubMed

    Khater, Shradha; Mohanty, Debasisa

    2015-01-01

    With the recent discoveries of novel post-translational modifications (PTMs) which play important roles in signaling and biosynthetic pathways, identification of such PTM catalyzing enzymes by genome mining has been an area of major interest. Unlike well-known PTMs like phosphorylation, glycosylation, SUMOylation, no bioinformatics resources are available for enzymes associated with novel and unusual PTMs. Therefore, we have developed the novPTMenzy database which catalogs information on the sequence, structure, active site and genomic neighborhood of experimentally characterized enzymes involved in five novel PTMs, namely AMPylation, Eliminylation, Sulfation, Hydroxylation and Deamidation. Based on a comprehensive analysis of the sequence and structural features of these known PTM catalyzing enzymes, we have created Hidden Markov Model profiles for the identification of similar PTM catalyzing enzymatic domains in genomic sequences. We have also created predictive rules for grouping them into functional subfamilies and deciphering their mechanistic details by structure-based analysis of their active site pockets. These analytical modules have been made available as user friendly search interfaces of novPTMenzy database. It also has a specialized analysis interface for some PTMs like AMPylation and Eliminylation. The novPTMenzy database is a unique resource that can aid in discovery of unusual PTM catalyzing enzymes in newly sequenced genomes. Database URL: http://www.nii.ac.in/novptmenzy.html

  10. Genetic modification of Bluetongue virus by uptake of "synthetic" genome segments.

    PubMed

    van Gennip, René G P; Veldman, Daniel; van de Water, Sandra G P; van Rijn, Piet A

    2010-10-07

    Since 1998, several serotypes of Bluetongue virus (BTV) have invaded several southern European countries. In 2006, the unknown BTV serotype 8 (BTV8/net06) unexpectedly invaded North-West Europe and has resulted in the largest BT-outbreak ever recorded. More recently, in 2008 BTV serotype 6 was reported in The Netherlands and Germany. This virus, BTV6/net08, is closely related to modified-live vaccine virus serotype 6, except for genome segment S10. This genome segment is closer related to that of vaccine virus serotype 2, and therefore BTV6/net08 is considered as a result of reassortment. Research on orbiviruses has been hampered by the lack of a genetic modification method. Recently, reverse genetics has been developed for BTV based on ten in vitro synthesized genomic RNAs. Here, we describe a targeted single-gene modification system for BTV based on the uptake of a single in vitro synthesized viral positive-stranded RNA. cDNAs corresponding to BTV8/net06 genome segments S7 and S10 were obtained by gene synthesis and cloned downstream of the T7 RNA-polymerase promoter and upstream of a unique site for a restriction enzyme at the 3'-terminus for run-off transcription. Monolayers of BSR cells were infected by BTV6/net08, and subsequently transfected with purified in vitro synthesized, capped positive-stranded S7 or S10 RNA from BTV8/net06 origin. "Synthetic" reassortants were rescued by endpoint dilutions, and identified by serotype-specific PCR-assays for segment 2, and serogroup-specific PCRs followed by restriction enzyme analysis or sequencing for S7 and S10 segments. The targeted single-gene modification system can also be used to study functions of viral proteins by uptake of mutated genome segments. This method is also useful to generate mutant orbiviruses for other serogroups of the genus Orbivirus for which reverse genetics has not been developed yet.

  11. Scalable human ES culture for therapeutic use: propagation, differentiation, genetic modification and regulatory issues.

    PubMed

    Rao, M

    2008-01-01

    Embryonic stem cells unlike most adult stem cell populations can replicate indefinitely while preserving genetic, epigenetic, mitochondrial and functional profiles. ESCs are therefore an excellent candidate cell type for providing a bank of cells for allogenic therapy and for introducing targeted genetic modifications for therapeutic intervention. This ability of prolonged self-renewal of stem cells and the unique advantages that this offers for gene therapy, discovery efforts, cell replacement, personalized medicine and other more direct applications requires the resolution of several important manufacturing, gene targeting and regulatory issues. In this review, we assess some of the advance made in developing scalable culture systems, improvement in vector design and gene insertion technology and the changing regulatory landscape.

  12. The evolution of adenoviral vectors through genetic and chemical surface modifications.

    PubMed

    Capasso, Cristian; Garofalo, Mariangela; Hirvinen, Mari; Cerullo, Vincenzo

    2014-02-17

    A long time has passed since the first clinical trial with adenoviral (Ad) vectors. Despite being very promising, Ad vectors soon revealed their limitations in human clinical trials. The pre-existing immunity, the marked liver tropism and the high toxicity of first generation Ad (FG-Ad) vectors have been the main challenges for the development of new approaches. Significant effort toward the development of genetically and chemically modified adenoviral vectors has enabled researchers to create more sophisticated vectors for gene therapy, with an improved safety profile and a higher transduction ability of different tissues. In this review, we will describe the latest findings in the high-speed, evolving field of genetic and chemical modifications of adenoviral vectors, a field in which different disciplines, such as biomaterial research, virology and immunology, co-operate synergistically to create better gene therapy tools for modern challenges.

  13. Genetic Mechanisms Involved in the Phenotype of Down Syndrome

    ERIC Educational Resources Information Center

    Patterson, David

    2007-01-01

    Down syndrome (DS) is the most common genetic cause of significant intellectual disability in the human population, occurring in roughly 1 in 700 live births. The ultimate cause of DS is trisomy of all or part of the set of genes located on chromosome 21. How this trisomy leads to the phenotype of DS is unclear. The completion of the DNA…

  14. Genetic Modification in Human Pluripotent Stem Cells by Homologous Recombination and CRISPR/Cas9 System.

    PubMed

    Xue, Haipeng; Wu, Jianbo; Li, Shenglan; Rao, Mahendra S; Liu, Ying

    2016-01-01

    Genetic modification is an indispensable tool to study gene function in normal development and disease. The recent breakthrough of creating human induced pluripotent stem cells (iPSCs) by defined factors (Takahashi et al., Cell 131:861-872, 2007) provides a renewable source of patient autologous cells that not only retain identical genetic information but also give rise to many cell types of the body including neurons and glia. Meanwhile, the rapid advancement of genome modification tools such as gene targeting by homologous recombination (Capecchi, Nat Rev Genet 6:507-512, 2005) and genome editing tools such as CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas (CRISPR-associated) system, TALENs (Transcription activator-like effector nucleases), and ZFNs (Zinc finger nucleases) (Wang et al., Cell 153:910-918, 2013; Mali et al., Science 339:823-826, 2013; Hwang et al., Nat Biotechnol 31:227-229, 2013; Friedland et al., Nat Methods 10(8):741-743, 2013; DiCarlo et al., Nucleic Acids Res 41:4336-4343, 2013; Cong et al., Science 339:819-823, 2013) has greatly accelerated the development of human genome manipulation at the molecular level. This chapter describes the protocols for making neural lineage reporter lines using homologous recombination and the CRISPR/Cas system-mediated genome editing, including construction of targeting vectors, guide RNAs, transfection into hPSCs, and selection and verification of successfully targeted clones. This method can be applied to various needs of hPSC genetic engineering at high efficiency and high reliability.

  15. Genetic modification of stem cells for improved therapy of the infarcted myocardium.

    PubMed

    Haider, Husnain Kh; Mustafa, Anique; Feng, Yuliang; Ashraf, Muhammad

    2011-10-01

    The conventional treatment modalities for ischemic heart disease only provide symptomatic relief to the patient without repairing and regenerating the damaged myocardium. Stem cell transplantation has emerged as a promising alternative therapeutic approach for cardiovascular diseases. Stem cells possess the potential of differentiation to adopt morphofunctional cardiac and vasculogenic phenotypes to repopulate the scar tissue and restore regional blood flow in the ischemic myocardium. These beneficial therapeutic effects make stem cell transplantation the method of choice for the treatment of ischemic heart disease. The efficacy of stem cell transplantation may be augmented by genetic manipulation of the cells prior to transplantation. Not only will insertion of therapeutic transgene(s) into the stem cells support the survival and differentiation of cells in the unfavorable microenvironment of the ischemic myocardium, but also the genetically manipulated stem cells will serve as a source of the transgene expression product in the heart for therapeutic benefits. We provide an overview of the extensively studied stem cell types for cardiac regeneration, the various methods in which these cells have been genetically manipulated and rationale of genetic modification of stem cells for use in regenerative cardiovascular therapeutics.

  16. Genetic code expansion in stable cell lines enables encoded chromatin modification.

    PubMed

    Elsässer, Simon J; Ernst, Russell J; Walker, Olivia S; Chin, Jason W

    2016-02-01

    Genetically encoded unnatural amino acids provide powerful strategies for modulating the molecular functions of proteins in mammalian cells. However, this approach has not been coupled to genome-wide measurements, because efficient incorporation of unnatural amino acids is limited to transient expression settings that lead to very heterogeneous expression. We demonstrate that stable integration of the Methanosarcina mazei pyrrolysyl-tRNA synthetase (PylRS)/tRNA(Pyl)CUA pair (and its derivatives) into the mammalian genome enables efficient, homogeneous incorporation of unnatural amino acids into target proteins in diverse mammalian cells, and we reveal the distinct transcriptional responses of embryonic stem cells and mouse embryonic fibroblasts to amber codon suppression. Genetically encoding N-ɛ-acetyl-lysine in place of six lysine residues in histone H3 enables deposition of pre-acetylated histones into cellular chromatin, via a pathway that is orthogonal to enzymatic modification. After synthetically encoding lysine-acetylation at natural modification sites, we determined the consequences of acetylation at specific amino acids in histones for gene expression. PMID:26727110

  17. Genetic code expansion in stable cell lines enables encoded chromatin modification

    PubMed Central

    Elsässer, Simon J.; Ernst, Russell J; Walker, Olivia S.; Chin, Jason W.

    2016-01-01

    Genetically encoded unnatural amino acids provide powerful strategies for modulating the molecular functions of proteins in mammalian cells. However this approach has not been coupled to genome-wide measurements, because efficient unnatural amino acid incorporation is limited to transient expression settings that lead to very heterogeneous expression. We demonstrate that stable \\ integration of the Methanosarcina mazei pyrrolysyl-tRNA synthetase (PylS)/tRNAPylCUA pair (and its derivatives) into the mammalian genome enables efficient, homogeneous unnatural amino acid incorporation into target proteins in diverse mammalian cells, and we reveal the distinct transcriptional responses of ES cells and MEFs to amber suppression. Genetically encoding Nε-acetyl-lysine in place of six lysine residues in histone H3 enables deposition of pre-acetylated histones into cellular chromatin, via a pathway that is orthogonal to enzymatic modification. Upon synthetically encoding lysine-acetylation at natural modification sites we determine the consequences of acetylation at specific amino acids in histones on gene expression. PMID:26727110

  18. Recent patents on genetic modification of plants and microbes for biomass conversion to biofuels.

    PubMed

    Lubieniechi, Simona; Peranantham, Thinesh; Levin, David B

    2013-04-01

    Development of sustainable energy systems based on renewable biomass feedstocks is now a global effort. Lignocellulosic biomass contains polymers of cellulose, hemicellulose, and lignin, bound together in a complex structure. Liquid biofuels, such as ethanol, can be made from biomass via fermentation of sugars derived from the cellulose and hemicellulose within lignocellulosic materials, but pre-treatment of the biomass to release sugars for microbial conversion is a significant barrier to commercial success of lignocellulosic biofuel production. Strategies to reduce the energy and cost inputs required for biomass pre-treatment include genetic modification of plant materials to reduce lignin content. Significant efforts are also underway to create recombinant microorganisms capable of converting sugars derived from lignocellulosic biomass to a variety of biofuels. An alternative strategy to reduce the costs of cellulosic biofuel production is the use of cellulolytic microorganisms capable of direct microbial conversion of ligno-cellulosic biomass to fuels. This paper reviews recent patents on genetic modification of plants and microbes for biomass conversion to biofuels.

  19. Genetically engineered charge modifications to enhance protein separation in aqueous two-phase systems: Electrochemical partitioning

    SciTech Connect

    Luther, J.R.; Glatz, C.E. . Dept. of Chemical Engineering)

    1994-06-20

    The authors examined the effect of genetically engineered charge modifications on the partitioning behavior of proteins in dextran/polyethylene glycol two-phase systems containing potassium phosphate. By genetically altering a protein's charge, the role of charge on partitioning can be assessed directly without the need to modify the phase system. The charge modifications used are of two types: charged tails of polyaspartic acid fused to [beta]-galactosidase and charge-change point mutations of T4 lysozyme which replace positive lysine residues with negative glutamic acids. The partition coefficient K[sub p] for these proteins was related to measured interfacial potential differences [Delta][phi] using the simple thermodynamic model, In K[sub p] = In K, + (FIRT)Z[sub p] [Delta][phi]. The protein net charge Z[sub p] was determined using the Henderson-Hasselbalch relationship with modifications based on experimentally determined titration and isoelectric point data. It was found that when the electropartitioning term Z[sub p] [Delta][phi] was varied by changing the pH, the partitioning of T4 lysozyme was quantitatively described by the thermodynamic model. The [beta]-galactosidase fusions displayed qualitative agreement, and although less than predicted, the partitioning increased more than two orders of magnitude for the pH range examined. Changes in the partitioning of lysozyme due to the various mutations agreed qualitatively with the thermodynamic model, but with a smaller than expected dependence on the estimated charge differences. The [beta]-galactosidase fusions, on the other hand, did not display a consistent charge based trend, which is likely due either to the enzyme's large size and complexity or to nonelectrostatic contributions from the tails. The lack of quantitative fit with the model described above suggests that the assumptions made in developing this model are oversimplified.

  20. GENETIC MODIFICATION OF GIBBERELLIC ACID SIGNALING TO PROMOTE CARBON SEQUESTRATION IN TREE ROOTS AND STEMS

    SciTech Connect

    Busov, Victor

    2013-03-05

    encode proteins involved in gibberellin metabolism or signalling. Intact genomic copies of PtGA20ox7, PtGA2ox2,Pt RGL1_1, PtRGL1_2 and PtGAI1 genes from the genome-sequenced Populus trichocarpa clone Nisqually-1 were transformed into Populus tremula - alba (clone INRA 717-1B4), and growth, morphology and xylem cell size characterized in the greenhouse. Each cisgene encompassed 1-2?kb of 5' and 1?kb of 3' flanking DNA, as well as all native exons and introns. Large numbers of independent insertion events per cisgene (19-38), including empty vector controls, were studied. Three of the cisgenic modifications had significant effects on plant growth rate, morphology or wood properties. The PtGA20ox7 cisgene increased rate of shoot regeneration in vitro, accelerated early growth, and variation in growth rate was correlated with PtGA20ox7 gene expression. PtRGL1_1 and PtGA2ox2 caused reduced growth, while PtRGL1_2 gave rise to plants that grew normally but had significantly longer xylem fibres. RT-PCR studies suggested that the lack of growth inhibition observed in PtRGL1_2 cisgenic plants was a result of co-suppression. PtGAI1 slowed regeneration rate and both PtGAI1 and PtGA20ox7 gave rise to increased variance among events for early diameter and volume index, respectively. Our work suggests that cisgenic insertion of additional copies of native genes involved in growth regulation may provide tools to help modify plant architecture, expand the genetic variance in plant architecture available to breeders and accelerate transfer of alleles between difficult-to-cross species. The role of gibberellins (GAs) in regulation of lateral root development is poorly understood. We show that GA-deficient (35S:PcGA2ox1) and GA-insensitive (35S:rgl1) transgenic Populus exhibited increased lateral root proliferation and elongation under in vitro and greenhouse conditions, and these effects were reversed by exogenous GA treatment. In addition, RNA interference suppression of two poplar GA 2

  1. Detection of thermogenesis in rodents in response to anti-obesity drugs and genetic modification

    PubMed Central

    Arch, Jonathan R. S.; Trayhurn, Paul

    2013-01-01

    Many compounds and genetic manipulations are claimed to confer resistance to obesity in rodents by raising energy expenditure. Examples taken from recent and older literature, demonstrate that such claims are often based on measurements of energy expenditure after body composition has changed, and depend on comparisons of energy expenditure divided by body weight. This is misleading because white adipose tissue has less influence than lean tissue on energy expenditure. Application of this approach to human data would suggest that human obesity is usually due to a low metabolic rate, which is not an accepted view. Increased energy expenditure per animal is a surer way of demonstrating thermogenesis, but even then it is important to know whether this is due to altered body composition (repartitioning), or increased locomotor activity rather than thermogenesis per se. Regression analysis offers other approaches. The thermogenic response to some compounds has a rapid onset and so cannot be due to altered body composition. These compounds usually mimic or activate the sympathetic nervous system. Thermogenesis occurs in, but may not be confined to, brown adipose tissue. It should not be assumed that weight loss in response to these treatments is due to thermogenesis unless there is a sustained increase in 24-h energy expenditure. Thyroid hormones and fibroblast growth factor 21 also raise energy expenditure before they affect body composition. Some treatments and genetic modifications alter the diurnal rhythm of energy expenditure. It is important to establish whether this is due to altered locomotor activity or efficiency of locomotion. There are no good examples of compounds that do not affect short-term energy expenditure but have a delayed effect. How and under what conditions a genetic modification or compound increases energy expenditure influences the decision on whether to seek drugs for the target or take a candidate drug into clinical studies. PMID:23580228

  2. Molecular/genetic determinants of repolarization and their modification by environmental stress.

    PubMed

    Rosen, M R; Cohen, I S

    2006-01-01

    Although a variety of factors, inherited or environmental, can influence expression of ion channel proteins to impact on repolarization, that environment can affect genetic determinants of repolarization for intervals of varying duration is a concept that is not as generally appreciated as it should be. In the following pages we review the molecular/genetic determinants of cardiac repolarization and summarize how pathologic events and environmental intrusions can affect these determinants. Understanding the chains of events involved should yield insights into both the causes and potential avenues of treatment for abnormalities of repolarization.

  3. Genetic disorders involving molecular-chaperone genes: a perspective.

    PubMed

    Macario, Alberto J L; Grippo, Tomas M; Conway de Macario, Everly

    2005-01-01

    Molecular chaperones are important for maintaining a functional set of proteins in all cellular compartments. Together with protein degradation machineries (e.g., the ubiquitin-proteasome system), chaperones form the core of the cellular protein-quality control mechanism. Chaperones are proteins, and as such, they can be affected by mutations. At least 15 disorders have been identified that are associated with mutations in genes encoding chaperones, or molecules with features suggesting that they function as chaperones. These chaperonopathies and a few other candidates are presented in this article. In most cases, the mechanisms by which the defective genes contribute to the observed phenotypes are still uncharacterized. However, the reported observations definitely point to the possibility that abnormal chaperones participate in pathogenesis. The available data open novel perspectives and should encourage searches for new genetic chaperonopathies, as well as further analyses of the disorders discussed in this article, including detection of new cases.

  4. Anticodon Modifications in the tRNA Set of LUCA and the Fundamental Regularity in the Standard Genetic Code.

    PubMed

    van der Gulik, Peter T S; Hoff, Wouter D

    2016-01-01

    Based on (i) an analysis of the regularities in the standard genetic code and (ii) comparative genomics of the anticodon modification machinery in the three branches of life, we derive the tRNA set and its anticodon modifications as it was present in LUCA. Previously we proposed that an early ancestor of LUCA contained a set of 23 tRNAs with unmodified anticodons that was capable of translating all 20 amino acids while reading 55 of the 61 sense codons of the standard genetic code (SGC). Here we use biochemical and genomic evidence to derive that LUCA contained a set of 44 or 45 tRNAs containing 2 or 3 modifications while reading 59 or 60 of the 61 sense codons. Subsequent tRNA modifications occurred independently in the Bacteria and Eucarya, while the Archaea have remained quite close to the tRNA set as it was present in LUCA. PMID:27454314

  5. Anticodon Modifications in the tRNA Set of LUCA and the Fundamental Regularity in the Standard Genetic Code

    PubMed Central

    van der Gulik, Peter T. S.; Hoff, Wouter D.

    2016-01-01

    Based on (i) an analysis of the regularities in the standard genetic code and (ii) comparative genomics of the anticodon modification machinery in the three branches of life, we derive the tRNA set and its anticodon modifications as it was present in LUCA. Previously we proposed that an early ancestor of LUCA contained a set of 23 tRNAs with unmodified anticodons that was capable of translating all 20 amino acids while reading 55 of the 61 sense codons of the standard genetic code (SGC). Here we use biochemical and genomic evidence to derive that LUCA contained a set of 44 or 45 tRNAs containing 2 or 3 modifications while reading 59 or 60 of the 61 sense codons. Subsequent tRNA modifications occurred independently in the Bacteria and Eucarya, while the Archaea have remained quite close to the tRNA set as it was present in LUCA. PMID:27454314

  6. Genetic modification of the human germ line: The reasons why this project has no future.

    PubMed

    Morange, Michel

    2015-01-01

    Modification of the human germ line has remained a distant but valuable objective for most biologists since the emergence of genetics (and even before). To study the historical transformations of this project, I have selected three periods - the 1930s, at the pinnacle of eugenics, around 1974 when molecular biology triumphed, and today - and have adopted three criteria to estimate the feasibility of this project: the state of scientific knowledge, the existence of suitable tools, and societal demands. Although the long-awaited techniques to modify the germ line are now available, I will show that most of the expectations behind this project have disappeared, or are considered as being reachable by highly different strategies.

  7. Restriction-Modification Systems as Mobile Genetic Elements in the Evolution of an Intracellular Symbiont

    PubMed Central

    Zheng, Hao; Dietrich, Carsten; Hongoh, Yuichi; Brune, Andreas

    2016-01-01

    Long-term vertical transmission of intracellular bacteria causes massive genomic erosion and results in extremely small genomes, particularly in ancient symbionts. Genome reduction is typically preceded by the accumulation of pseudogenes and proliferation of mobile genetic elements, which are responsible for chromosome rearrangements during the initial stage of endosymbiosis. We compared the genomes of an endosymbiont of termite gut flagellates, “Candidatus Endomicrobium trichonymphae,” and its free-living relative Endomicrobium proavitum and discovered many remnants of restriction-modification (R-M) systems that are consistently associated with genome rearrangements in the endosymbiont genome. The rearrangements include apparent insertions, transpositions, and the duplication of a genomic region; there was no evidence of transposon structures or other mobile elements. Our study reveals a so far unrecognized mechanism for genome rearrangements in intracellular symbionts and sheds new light on the general role of R-M systems in genome evolution. PMID:26568615

  8. Genetic modification of hematopoietic stem cells with nonviral systems: past progress and future prospects.

    PubMed

    Papapetrou, E P; Zoumbos, N C; Athanassiadou, A

    2005-10-01

    Serious unwanted complications provoked by retroviral gene transfer into hematopoietic stem cells (HSCs) have recently raised the need for the development and assessment of alternative gene transfer vectors. Within this context, nonviral gene transfer systems are attracting increasing interest. Their main advantages include low cost, ease of handling and large-scale production, large packaging capacity and, most importantly, biosafety. While nonviral gene transfer into HSCs has been restricted in the past by poor transfection efficiency and transient maintenance, in recent years, biotechnological developments are converting nonviral transfer into a realistic approach for genetic modification of cells of hematopoietic origin. Herein we provide an overview of past accomplishments in the field of nonviral gene transfer into hematopoietic progenitor/stem cells and we point at future challenges. We argue that episomally maintained self-replicating vectors combined with physical methods of delivery show the greatest promise among nonviral gene transfer strategies for the treatment of disorders of the hematopoietic system.

  9. Genetic modification of the human germ line: The reasons why this project has no future.

    PubMed

    Morange, Michel

    2015-01-01

    Modification of the human germ line has remained a distant but valuable objective for most biologists since the emergence of genetics (and even before). To study the historical transformations of this project, I have selected three periods - the 1930s, at the pinnacle of eugenics, around 1974 when molecular biology triumphed, and today - and have adopted three criteria to estimate the feasibility of this project: the state of scientific knowledge, the existence of suitable tools, and societal demands. Although the long-awaited techniques to modify the germ line are now available, I will show that most of the expectations behind this project have disappeared, or are considered as being reachable by highly different strategies. PMID:26231145

  10. [Detection of genetic modification in maize and maize products by ELISA-test].

    PubMed

    Urbanek-Karłowska, Bogumiła; Sawilska-Rautenstrauch, Dorota; Jedra, Małgorzata; Badowski, Paweł

    2003-01-01

    Enzyme immunoassay methods--TRAIT Test--was applied for detection of genetic modification in maize seeds and foodstuffs, which have been produced from this crop. TRAIT Test is based on the identification GMO protein Cry 1Ab produced by a gene derived from Bacillus thuringiensis (Bt) incorporated into insect resistant corn grain. The experiment was carried out on maize standards and foodstuffs from Warsaw market. The positive result was obtained for one maize product, which was not labelled as GMO. The presence of GMO material was approximately equal to 1%. In conclusion, this test is proper for fast routine qualitative (yes/no) determination GMO material in maize seeds and unprocessed food products.

  11. Genetic mapping of tumor susceptibility genes involved in mouse plasmacytomagenesis

    SciTech Connect

    Mock, B.A.; Krall, M.M.; Dosik, J.K. )

    1993-10-15

    Plasmacytomas (PCTs) were induced in 47% of BALB/cAnPt mice by the intraperitoneal injection of pristane, in 2% of (BALB/c [times] DBA/2N)F[sub 1], and in 11% of 773 BALB/cAnPt [times] (BALB/cAnPt [times] DBA/2N)F[sub 1]N[sub 2] backcross mice. This result indicates a multigenic mode of inheritance for PCT susceptibility. To locate genes controlling this complex genetic trait, tumor susceptibility in backcross progeny generated from BALB/c and DBA/2N (resistant) mice was correlated with alleles of 83 marker loci. The genotypes of the PCT-susceptible progeny displayed an excess homozygosity for BALB/c alleles with a 32-centimorgan stretch of mouse chromosome 4 (>95% probability of linkage) with minimal recombination (12%) near Gt10. Another susceptibility gene on mouse chromosome 1 may be linked to Fcgr2 (90% probability of linkage); there were excess heterozygotes for Fcgr2 among the susceptible progeny and excess homozygotes among the resistant progeny. Regions of mouse chromosomes 4 and 1 that are correlated with PCT susceptibility share extensive linkage homology with regions of human chromosome 1 that have been associated with cytogenetic abnormalities in multiple myeloma and lymphoid, breast, and endocrine tumors. 68 refs., 2 figs., 1 tab.

  12. Two-subunit enzymes involved in eukaryotic post-transcriptional tRNA modification

    PubMed Central

    Guy, Michael P; Phizicky, Eric M

    2014-01-01

    tRNA modifications are crucial for efficient and accurate protein translation, with defects often linked to disease. There are 7 cytoplasmic tRNA modifications in the yeast Saccharomyces cerevisiae that are formed by an enzyme consisting of a catalytic subunit and an auxiliary protein, 5 of which require only a single subunit in bacteria, and 2 of which are not found in bacteria. These enzymes include the deaminase Tad2-Tad3, and the methyltransferases Trm6-Trm61, Trm8-Trm82, Trm7-Trm732, and Trm7-Trm734, Trm9-Trm112, and Trm11-Trm112. We describe the occurrence and biological role of each modification, evidence for a required partner protein in S. cerevisiae and other eukaryotes, evidence for a single subunit in bacteria, and evidence for the role of the non-catalytic binding partner. Although it is unclear why these eukaryotic enzymes require partner proteins, studies of some 2-subunit modification enzymes suggest that the partner proteins help expand substrate range or allow integration of cellular activities. PMID:25625329

  13. Genetic recombination pathways and their application for genome modification of human embryonic stem cells.

    PubMed

    Nieminen, Mikko; Tuuri, Timo; Savilahti, Harri

    2010-10-01

    Human embryonic stem cells are pluripotent cells derived from early human embryo and retain a potential to differentiate into all adult cell types. They provide vast opportunities in cell replacement therapies and are expected to become significant tools in drug discovery as well as in the studies of cellular and developmental functions of human genes. The progress in applying different types of DNA recombination reactions for genome modification in a variety of eukaryotic cell types has provided means to utilize recombination-based strategies also in human embryonic stem cells. Homologous recombination-based methods, particularly those utilizing extended homologous regions and those employing zinc finger nucleases to boost genomic integration, have shown their usefulness in efficient genome modification. Site-specific recombination systems are potent genome modifiers, and they can be used to integrate DNA into loci that contain an appropriate recombination signal sequence, either naturally occurring or suitably pre-engineered. Non-homologous recombination can be used to generate random integrations in genomes relatively effortlessly, albeit with a moderate efficiency and precision. DNA transposition-based strategies offer substantially more efficient random strategies and provide means to generate single-copy insertions, thus potentiating the generation of genome-wide insertion libraries applicable in genetic screens.

  14. Effects of genetic modifications to flax (Linum usitatissimum) on arbuscular mycorrhiza and plant performance.

    PubMed

    Wróbel-Kwiatkowska, Magdalena; Turnau, Katarzyna; Góralska, Katarzyna; Anielska, Teresa; Szopa, Jan

    2012-10-01

    Although arbuscular mycorrhizal fungi (AMF) are known for their positive effect on flax growth, the impact of genetic manipulation in this crop on arbuscular mycorrhiza and plant performance was assessed for the first time. Five types of transgenic flax that were generated to improve fiber quality and resistance to pathogens, through increased levels of either phenylpropanoids (W92.40), glycosyltransferase (GT4, GT5), or PR2 beta-1,3-glucanase (B14) or produce polyhydroxybutyrate (M50), were used. Introduced genetic modifications did not change the degree of mycorrhizal colonization as compared to parent cultivars Linola and Nike. Arbuscules were well developed in each tested transgenic type (except M50). In two lines (W92.40 and B14), a higher abundance of arbuscules was observed when compared to control, untransformed flax plants. However, in some cases (W92.40, GT4, GT5, and B14 Md), the mycorrhizal dependency for biomass production of transgenic plants was slightly lower when compared to the original cultivars. No significant influence of mycorrhiza on the photosynthetic activity of transformed lines was found, but in most cases P concentration in mycorrhizal plants remained higher than in nonmycorrhizal ones. The transformed flax lines meet the demands for better quality of fiber and higher resistance to pathogens, without significantly influencing the interaction with AMF.

  15. Immunotherapy for Osteosarcoma: Genetic Modification of T cells Overcomes Low Levels of Tumor Antigen Expression

    PubMed Central

    Ahmed, Nabil; Salsman, Vita S; Yvon, Eric; Louis, Chrystal U; Perlaky, Laszlo; Wels, Winfried S; Dishop, Meghan K; Kleinerman, Eugenie E; Pule, Martin; Rooney, Cliona M; Heslop, Helen E; Gottschalk, Stephen

    2009-01-01

    Human epidermal growth factor receptor 2 (HER2) is expressed by the majority of human osteosarcomas and is a risk factor for poor outcome. Unlike breast cancer, osteosarcoma cells express HER2 at too low, a level for patients to benefit from HER2 monoclonal antibodies. We reasoned that this limitation might be overcome by genetically modifying T cells with HER2-specific chimeric antigen receptors (CARs), because even a low frequency of receptor engagement could be sufficient to induce effector cell killing of the tumor. HER2-specific T cells were generated by retroviral transduction with a HER2-specific CAR containing a CD28.ζ signaling domain. HER2-specific T cells recognized HER2-positive osteosarcoma cells as judged by their ability to proliferate, produce immunostimulatory T helper 1 cytokines, and kill HER2-positive osteosarcoma cell lines in vitro. The adoptive transfer of HER2-specific T cells caused regression of established osteosarcoma xenografts in locoregional as well as metastatic mouse models. In contrast, delivery of nontransduced (NT) T cells did not change the tumor growth pattern. Genetic modification of T cells with CARs specific for target antigens, expressed at too low a level to be effectively recognized by monoclonal antibodies, may allow immunotherapy to be more broadly applicable for human cancer therapy. PMID:19532139

  16. An injectable spheroid system with genetic modification for cell transplantation therapy.

    PubMed

    Uchida, Satoshi; Itaka, Keiji; Nomoto, Takahiro; Endo, Taisuke; Matsumoto, Yu; Ishii, Takehiko; Kataoka, Kazunori

    2014-03-01

    The new methodology to increase a therapeutic potential of cell transplantation was developed here by the use of three-dimensional spheroids of transplanting cells subsequent to the genetic modification with non-viral DNA vectors, polyplex nanomicelles. Particularly, spheroids in regulated size of 100-μm of primary hepatocytes transfected with luciferase gene were formed on the micropatterned culture plates coated with thermosensitive polymer, and were recovered in the form of injectable liquid suspension simply by cooling the plates. After subcutaneously transplanting these hepatocyte spheroids, efficient transgene expression was observed in host tissue for more than a month, whereas transplantation of a single-cell suspension from a monolayer culture resulted in an only transient expression. The spheroid system contributed to the preservation of innate functions of transplanted hepatocytes in the host tissue, such as albumin expression, thereby possessing high potential for expressing transgene. Intravital observation of transplanted cells showed that those from spheroid cultures had a tendency to localize in the vicinity of blood vessels, making a favorable microenvironment for preserving cell functionality. Furthermore, spheroids transfected with erythropoietin-expressing DNA showed a significantly higher hematopoietic effect than that of cell suspensions from monolayer cultures, demonstrating high potential of this genetically-modified spheroid transplantation system for therapeutic applications. PMID:24388386

  17. N-terminal modifications of cellular proteins: The enzymes involved, their substrate specificities and biological effects

    PubMed Central

    Varland, Sylvia; Osberg, Camilla; Arnesen, Thomas

    2015-01-01

    The vast majority of eukaryotic proteins are N-terminally modified by one or more processing enzymes. Enzymes acting on the very first amino acid of a polypeptide include different peptidases, transferases, and ligases. Methionine aminopeptidases excise the initiator methionine leaving the nascent polypeptide with a newly exposed amino acid that may be further modified. N-terminal acetyl-, methyl-, myristoyl-, and palmitoyltransferases may attach an acetyl, methyl, myristoyl, or palmitoyl group, respectively, to the α-amino group of the target protein N-terminus. With the action of ubiquitin ligases, one or several ubiquitin molecules are transferred, and hence, constitute the N-terminal modification. Modifications at protein N-termini represent an important contribution to proteomic diversity and complexity, and are essential for protein regulation and cellular signaling. Consequently, dysregulation of the N-terminal modifying enzymes is implicated in human diseases. We here review the different protein N-terminal modifications occurring co- or post-translationally with emphasis on the responsible enzymes and their substrate specificities. PMID:25914051

  18. Molecular genetic analysis of activation-tagged transcription factors thought to be involved in photomorphogenesis

    SciTech Connect

    Neff, Michael M.

    2011-06-23

    This is a final report for Department of Energy Grant No. DE-FG02-08ER15927 entitled “Molecular Genetic Analysis of Activation-Tagged Transcription Factors Thought to be Involved in Photomorphogenesis”. Based on our preliminary photobiological and genetic analysis of the sob1-D mutant, we hypothesized that OBP3 is a transcription factor involved in both phytochrome and cryptochrome-mediated signal transduction. In addition, we hypothesized that OBP3 is involved in auxin signaling and root development. Based on our preliminary photobiological and genetic analysis of the sob2-D mutant, we also hypothesized that a related gene, LEP, is involved in hormone signaling and seedling development.

  19. Phycobiliprotein biosynthesis in cyanobacteria: structure and function of enzymes involved in post-translational modification.

    PubMed

    Schluchter, Wendy M; Shen, Gaozhong; Alvey, Richard M; Biswas, Avijit; Saunée, Nicolle A; Williams, Shervonda R; Mille, Crystal A; Bryant, Donald A

    2010-01-01

    Cyanobacterial phycobiliproteins are brilliantly colored due to the presence of covalently attached chromophores called bilins, linear tetrapyrroles derived from heme. For most phycobiliproteins, these post-translational modifications are catalyzed by enzymes called bilin lyases; these enzymes ensure that the appropriate bilins are attached to the correct cysteine residues with the proper stereochemistry on each phycobiliprotein subunit. Phycobiliproteins also contain a unique, post-translational modification, the methylation of a conserved asparagine (Asn) present at beta-72, which occurs on the beta-subunits of all phycobiliproteins. We have identified and characterized several new families of bilin lyases, which are responsible for attaching PCB to phycobiliproteins as well as the Asn methyl transferase for beta-subunits in Synechococcus sp. PCC 7002 and Synechocystis sp. PCC 6803. All of the enzymes responsible for synthesis of holo-phycobiliproteins are now known for this cyanobacterium, and a brief discussion of each enzyme family and its role in the biosynthesis of phycobiliproteins is presented here. In addition, the first structure of a bilin lyase has recently been solved (PDB ID: 3BDR). This structure shows that the bilin lyases are most similar to the lipocalin protein structural family, which also includes the bilin-binding protein found in some butterflies.

  20. Computational identification of novel biochemical systems involved in oxidation, glycosylation and other complex modifications of bases in DNA.

    PubMed

    Iyer, Lakshminarayan M; Zhang, Dapeng; Burroughs, A Maxwell; Aravind, L

    2013-09-01

    Discovery of the TET/JBP family of dioxygenases that modify bases in DNA has sparked considerable interest in novel DNA base modifications and their biological roles. Using sensitive sequence and structure analyses combined with contextual information from comparative genomics, we computationally characterize over 12 novel biochemical systems for DNA modifications. We predict previously unidentified enzymes, such as the kinetoplastid J-base generating glycosyltransferase (and its homolog GREB1), the catalytic specificity of bacteriophage TET/JBP proteins and their role in complex DNA base modifications. We also predict the enzymes involved in synthesis of hypermodified bases such as alpha-glutamylthymine and alpha-putrescinylthymine that have remained enigmatic for several decades. Moreover, the current analysis suggests that bacteriophages and certain nucleo-cytoplasmic large DNA viruses contain an unexpectedly diverse range of DNA modification systems, in addition to those using previously characterized enzymes such as Dam, Dcm, TET/JBP, pyrimidine hydroxymethylases, Mom and glycosyltransferases. These include enzymes generating modified bases such as deazaguanines related to queuine and archaeosine, pyrimidines comparable with lysidine, those derived using modified S-adenosyl methionine derivatives and those using TET/JBP-generated hydroxymethyl pyrimidines as biosynthetic starting points. We present evidence that some of these modification systems are also widely dispersed across prokaryotes and certain eukaryotes such as basidiomycetes, chlorophyte and stramenopile alga, where they could serve as novel epigenetic marks for regulation or discrimination of self from non-self DNA. Our study extends the role of the PUA-like fold domains in recognition of modified nucleic acids and predicts versions of the ASCH and EVE domains to be novel 'readers' of modified bases in DNA. These results open opportunities for the investigation of the biology of these systems and

  1. Computational identification of novel biochemical systems involved in oxidation, glycosylation and other complex modifications of bases in DNA

    PubMed Central

    Iyer, Lakshminarayan M.; Zhang, Dapeng; Maxwell Burroughs, A.; Aravind, L.

    2013-01-01

    Discovery of the TET/JBP family of dioxygenases that modify bases in DNA has sparked considerable interest in novel DNA base modifications and their biological roles. Using sensitive sequence and structure analyses combined with contextual information from comparative genomics, we computationally characterize over 12 novel biochemical systems for DNA modifications. We predict previously unidentified enzymes, such as the kinetoplastid J-base generating glycosyltransferase (and its homolog GREB1), the catalytic specificity of bacteriophage TET/JBP proteins and their role in complex DNA base modifications. We also predict the enzymes involved in synthesis of hypermodified bases such as alpha-glutamylthymine and alpha-putrescinylthymine that have remained enigmatic for several decades. Moreover, the current analysis suggests that bacteriophages and certain nucleo-cytoplasmic large DNA viruses contain an unexpectedly diverse range of DNA modification systems, in addition to those using previously characterized enzymes such as Dam, Dcm, TET/JBP, pyrimidine hydroxymethylases, Mom and glycosyltransferases. These include enzymes generating modified bases such as deazaguanines related to queuine and archaeosine, pyrimidines comparable with lysidine, those derived using modified S-adenosyl methionine derivatives and those using TET/JBP-generated hydroxymethyl pyrimidines as biosynthetic starting points. We present evidence that some of these modification systems are also widely dispersed across prokaryotes and certain eukaryotes such as basidiomycetes, chlorophyte and stramenopile alga, where they could serve as novel epigenetic marks for regulation or discrimination of self from non-self DNA. Our study extends the role of the PUA-like fold domains in recognition of modified nucleic acids and predicts versions of the ASCH and EVE domains to be novel ‘readers’ of modified bases in DNA. These results open opportunities for the investigation of the biology of these systems

  2. Nuclear-encoded factors involved in post-transcriptional processing and modification of mitochondrial tRNAs in human disease

    PubMed Central

    Powell, Christopher A.; Nicholls, Thomas J.; Minczuk, Michal

    2015-01-01

    The human mitochondrial genome (mtDNA) encodes 22 tRNAs (mt-tRNAs) that are necessary for the intraorganellar translation of the 13 mtDNA-encoded subunits of the mitochondrial respiratory chain complexes. Maturation of mt-tRNAs involves 5′ and 3′ nucleolytic excision from precursor RNAs, as well as extensive post-transcriptional modifications. Recent data suggest that over 7% of all mt-tRNA residues in mammals undergo post-transcriptional modification, with over 30 different modified mt-tRNA positions so far described. These processing and modification steps are necessary for proper mt-tRNA function, and are performed by dedicated, nuclear-encoded enzymes. Recent growing evidence suggests that mutations in these nuclear genes (nDNA), leading to incorrect maturation of mt-tRNAs, are a cause of human mitochondrial disease. Furthermore, mtDNA mutations in mt-tRNA genes, which may also affect mt-tRNA function, processing, and modification, are also frequently associated with human disease. In theory, all pathogenic mt-tRNA variants should be expected to affect only a single process, which is mitochondrial translation, albeit to various extents. However, the clinical manifestations of mitochondrial disorders linked to mutations in mt-tRNAs are extremely heterogeneous, ranging from defects of a single tissue to complex multisystem disorders. This review focuses on the current knowledge of nDNA coding for proteins involved in mt-tRNA maturation that have been linked to human mitochondrial pathologies. We further discuss the possibility that tissue specific regulation of mt-tRNA modifying enzymes could play an important role in the clinical heterogeneity observed for mitochondrial diseases caused by mutations in mt-tRNA genes. PMID:25806043

  3. Quantitative Genetics Identifies Cryptic Genetic Variation Involved in the Paternal Regulation of Seed Development.

    PubMed

    Pires, Nuno D; Bemer, Marian; Müller, Lena M; Baroux, Célia; Spillane, Charles; Grossniklaus, Ueli

    2016-01-01

    Embryonic development requires a correct balancing of maternal and paternal genetic information. This balance is mediated by genomic imprinting, an epigenetic mechanism that leads to parent-of-origin-dependent gene expression. The parental conflict (or kinship) theory proposes that imprinting can evolve due to a conflict between maternal and paternal alleles over resource allocation during seed development. One assumption of this theory is that paternal alleles can regulate seed growth; however, paternal effects on seed size are often very low or non-existent. We demonstrate that there is a pool of cryptic genetic variation in the paternal control of Arabidopsis thaliana seed development. Such cryptic variation can be exposed in seeds that maternally inherit a medea mutation, suggesting that MEA acts as a maternal buffer of paternal effects. Genetic mapping using recombinant inbred lines, and a novel method for the mapping of parent-of-origin effects using whole-genome sequencing of segregant bulks, indicate that there are at least six loci with small, paternal effects on seed development. Together, our analyses reveal the existence of a pool of hidden genetic variation on the paternal control of seed development that is likely shaped by parental conflict.

  4. Quantitative Genetics Identifies Cryptic Genetic Variation Involved in the Paternal Regulation of Seed Development

    PubMed Central

    Pires, Nuno D.; Bemer, Marian; Müller, Lena M.; Baroux, Célia; Spillane, Charles; Grossniklaus, Ueli

    2016-01-01

    Embryonic development requires a correct balancing of maternal and paternal genetic information. This balance is mediated by genomic imprinting, an epigenetic mechanism that leads to parent-of-origin-dependent gene expression. The parental conflict (or kinship) theory proposes that imprinting can evolve due to a conflict between maternal and paternal alleles over resource allocation during seed development. One assumption of this theory is that paternal alleles can regulate seed growth; however, paternal effects on seed size are often very low or non-existent. We demonstrate that there is a pool of cryptic genetic variation in the paternal control of Arabidopsis thaliana seed development. Such cryptic variation can be exposed in seeds that maternally inherit a medea mutation, suggesting that MEA acts as a maternal buffer of paternal effects. Genetic mapping using recombinant inbred lines, and a novel method for the mapping of parent-of-origin effects using whole-genome sequencing of segregant bulks, indicate that there are at least six loci with small, paternal effects on seed development. Together, our analyses reveal the existence of a pool of hidden genetic variation on the paternal control of seed development that is likely shaped by parental conflict. PMID:26811909

  5. Quantitative Genetics Identifies Cryptic Genetic Variation Involved in the Paternal Regulation of Seed Development.

    PubMed

    Pires, Nuno D; Bemer, Marian; Müller, Lena M; Baroux, Célia; Spillane, Charles; Grossniklaus, Ueli

    2016-01-01

    Embryonic development requires a correct balancing of maternal and paternal genetic information. This balance is mediated by genomic imprinting, an epigenetic mechanism that leads to parent-of-origin-dependent gene expression. The parental conflict (or kinship) theory proposes that imprinting can evolve due to a conflict between maternal and paternal alleles over resource allocation during seed development. One assumption of this theory is that paternal alleles can regulate seed growth; however, paternal effects on seed size are often very low or non-existent. We demonstrate that there is a pool of cryptic genetic variation in the paternal control of Arabidopsis thaliana seed development. Such cryptic variation can be exposed in seeds that maternally inherit a medea mutation, suggesting that MEA acts as a maternal buffer of paternal effects. Genetic mapping using recombinant inbred lines, and a novel method for the mapping of parent-of-origin effects using whole-genome sequencing of segregant bulks, indicate that there are at least six loci with small, paternal effects on seed development. Together, our analyses reveal the existence of a pool of hidden genetic variation on the paternal control of seed development that is likely shaped by parental conflict. PMID:26811909

  6. Genetic engineering of novel flower colour by suppression of anthocyanin modification genes in gentian.

    PubMed

    Nakatsuka, Takashi; Mishiba, Kei-ichiro; Kubota, Akiko; Abe, Yoshiko; Yamamura, Saburo; Nakamura, Noriko; Tanaka, Yoshikazu; Nishihara, Masahiro

    2010-02-15

    Ornamental gentian plants have vivid-blue flowers. The main factor contributing to the flower colour is the accumulation of a polyacylated delphinidin 'gentiodelphin' in their petals. Although in vitro studies proposed that acylation plays an important role in the stability and development of gentian blue colour, the in vivo stability of the polyacylated anthocyanin was not clearly demonstrated. Thus, to reveal the importance of anthocyanin modification, especially acylation, and to engineer new colours of gentian flowers, we used chimeric RNAi technology to produce transgenic gentian plants with downregulated anthocyanin 5,3'-aromatic acyltransferase (5/3'AT) and flavonoid 3',5'-hydroxylase (F3'5'H) activities, which are both essential enzymes for gentiodelphin biosynthesis. Two lines of flower colour-modified plants were obtained from fifteen transgenic gentian plants. Clone no. 1 exhibited a lilac flower colour and clone no. 15 exhibited pale-blue flowers. RNA gel blot analysis confirmed that both transgenic lines had markedly suppressed 5/3'AT transcripts, whereas clone no. 15 had fewer F3'5'H transcripts than clone no. 1 and untransformed control plants. HPLC analysis of anthocyanin compositions showed that downregulation of the 5/3'AT gene led to increased accumulation of non-acylated anthocyanins, as expected. These results demonstrated that genetic engineering to reduce the accumulation of polyacylated anthocyanins could cause modulations of flower colour.

  7. Factors influencing U.S. consumer support for genetic modification to prevent crop disease.

    PubMed

    McComas, Katherine A; Besley, John C; Steinhardt, Joseph

    2014-07-01

    This study examines support for the genetic modification (GM) of crops in the context of preventing "late blight," a devastating potato and tomato disease that caused the Irish Potato Famine in the 1850s and results in substantial crop loss today. We surveyed U.S. adults who do the primary grocery shopping in their household (n = 859). Half of the respondents were randomly assigned to read a vignette describing late blight before responding to questions about GM, whereas the other half read a vignette about generic crop disease before responding to questions. We also examine how the perceived fairness of decision makers relates to GM support and the perceived legitimacy of GM decision making. We found that disease specificity mattered less to support and legitimacy than the perceived fairness of decision makers. The perceived risks of GM to human and environmental health negatively related to GM support and legitimacy, whereas the perceived benefits (e.g. reduced threats to crops and a more secure food supply) positively related to support and legitimacy. Objective knowledge about GM had a small, negative relationship with legitimacy whereas self-assessed familiarity with GM had a positive relationship. Overall, the results offer additional confirmation of past findings from more localized settings that perceived fairness of decision makers matters to support for GM and underscore the importance of considering how risk managers' behaviors and actions are perceived alongside individuals' perceptions about the risks and benefits. PMID:24630937

  8. Factors influencing U.S. consumer support for genetic modification to prevent crop disease.

    PubMed

    McComas, Katherine A; Besley, John C; Steinhardt, Joseph

    2014-07-01

    This study examines support for the genetic modification (GM) of crops in the context of preventing "late blight," a devastating potato and tomato disease that caused the Irish Potato Famine in the 1850s and results in substantial crop loss today. We surveyed U.S. adults who do the primary grocery shopping in their household (n = 859). Half of the respondents were randomly assigned to read a vignette describing late blight before responding to questions about GM, whereas the other half read a vignette about generic crop disease before responding to questions. We also examine how the perceived fairness of decision makers relates to GM support and the perceived legitimacy of GM decision making. We found that disease specificity mattered less to support and legitimacy than the perceived fairness of decision makers. The perceived risks of GM to human and environmental health negatively related to GM support and legitimacy, whereas the perceived benefits (e.g. reduced threats to crops and a more secure food supply) positively related to support and legitimacy. Objective knowledge about GM had a small, negative relationship with legitimacy whereas self-assessed familiarity with GM had a positive relationship. Overall, the results offer additional confirmation of past findings from more localized settings that perceived fairness of decision makers matters to support for GM and underscore the importance of considering how risk managers' behaviors and actions are perceived alongside individuals' perceptions about the risks and benefits.

  9. Clinical lung xenotransplantation – what donor genetic modifications may be necessary?

    PubMed Central

    Cooper, David K. C.; Ekser, Burcin; Burlak, Christopher; Ezzelarab, Mohamed; Hara, Hidetaka; Paris, Leela; Tector, A. Joseph; Phelps, Carol; Azimzadeh, Agnes M.; Ayares, David; Robson, Simon C.; Pierson, Richard N.

    2013-01-01

    Barriers to successful lung xenotransplantation appear to be even greater than for other organs. This difficulty may be related to several macro anatomic factors, such as the uniquely fragile lung parenchyma and associated blood supply that results in heightened vulnerability of graft function to segmental or lobar airway flooding caused by loss of vascular integrity (also applicable to allotransplants). There are also micro-anatomic considerations, such as the presence of large numbers of resident inflammatory cells, such as pulmonary intravascular macrophages and natural killer (NK) T cells, and the high levels of von Willebrand factor (vWF) associated with the microvasculature. We have considered what developments would be necessary to allow successful clinical lung xenotransplantation. We suggest this will only be achieved by multiple genetic modifications of the organ-source pig, in particular to render the vasculature resistant to thrombosis. The major problems that require to be overcome are multiple and include (i) the innate immune response (antibody, complement, donor pulmonary and recipient macrophages, monocytes, neutrophils, and NK cells), (ii) the adaptive immune response (T and B cells), (iii) coagulation dysregulation, and (iv) an inflammatory response (e.g., TNF-α, IL-6, HMGB1, C-reactive protein). We propose that the genetic manipulation required to provide normal thromboregulation alone may include the introduction of genes for human thrombomodulin/endothelial protein C-receptor, and/or tissue factor pathway inhibitor, and/or CD39/CD73; the problem of pig vWF may also need to be addressed. It would appear that exploration of every available therapeutic path will be required if lung xenotransplantation is to be successful. To initiate a clinical trial of lung xenotransplantation, even as a bridge to allotransplantation (with a realistic possibility of survival long enough for a human lung allograft to be obtained), significant advances and much

  10. Both common and specific genetic factors are involved in polygenic resistance of pepper to several potyviruses.

    PubMed

    Caranta, C; Palloix, A

    1996-01-01

    Absolute resistance to potato virus Y pathotype 0 (PVY 0), potyvirus E and chili veinal mottle virus (CVMV) and a partial resistance to potato virus Y pathotype 1,2 (PVY 1,2) were found in an Indian pepper line, 'Perennial'. In the doubled haploid (DH) progeny from the F1 of a cross 'Perennial' by 'Yolo Wonder', resistance to CVMV was confered by two independent genes, one with a clear dominant effect. Resistance to PVY and potyvirus E was quantitatively expressed and controlled by several recessive genetic factors. Genetic analysis showed that fewer resistance factors were necessary to explain resistance to PVY (0) and potyvirus E than resistance to PVY(1,2). Genetic correlations between resistances to the different potyviruses in the DH progeny showed that most of genetic factors involved in PVY(0) resistance appear to be also involved in potyvirus E resistance, and some of these polyvalent factors may be also involved in PVY(1,2) resistance but, in this case, additional specific genes were necessary. One of the two CVMV resistance genes seems to be implicated in potyvirus E resistance. Thus, the polygenic resistance of 'Perennial' to these potyviruses was due both to polyvalent genetic factors, i.e. factors that apparently interact with several viruses, and strain-specific genetic factors. PMID:24166111

  11. CRISPR/Cas9: a molecular Swiss army knife for simultaneous introduction of multiple genetic modifications in Saccharomyces cerevisiae.

    PubMed

    Mans, Robert; van Rossum, Harmen M; Wijsman, Melanie; Backx, Antoon; Kuijpers, Niels G A; van den Broek, Marcel; Daran-Lapujade, Pascale; Pronk, Jack T; van Maris, Antonius J A; Daran, Jean-Marc G

    2015-03-01

    A variety of techniques for strain engineering in Saccharomyces cerevisiae have recently been developed. However, especially when multiple genetic manipulations are required, strain construction is still a time-consuming process. This study describes new CRISPR/Cas9-based approaches for easy, fast strain construction in yeast and explores their potential for simultaneous introduction of multiple genetic modifications. An open-source tool (http://yeastriction.tnw.tudelft.nl) is presented for identification of suitable Cas9 target sites in S. cerevisiae strains. A transformation strategy, using in vivo assembly of a guideRNA plasmid and subsequent genetic modification, was successfully implemented with high accuracies. An alternative strategy, using in vitro assembled plasmids containing two gRNAs, was used to simultaneously introduce up to six genetic modifications in a single transformation step with high efficiencies. Where previous studies mainly focused on the use of CRISPR/Cas9 for gene inactivation, we demonstrate the versatility of CRISPR/Cas9-based engineering of yeast by achieving simultaneous integration of a multigene construct combined with gene deletion and the simultaneous introduction of two single-nucleotide mutations at different loci. Sets of standardized plasmids, as well as the web-based Yeastriction target-sequence identifier and primer-design tool, are made available to the yeast research community to facilitate fast, standardized and efficient application of the CRISPR/Cas9 system.

  12. CRISPR/Cas9: a molecular Swiss army knife for simultaneous introduction of multiple genetic modifications in Saccharomyces cerevisiae

    PubMed Central

    Mans, Robert; van Rossum, Harmen M.; Wijsman, Melanie; Backx, Antoon; Kuijpers, Niels G.A.; van den Broek, Marcel; Daran-Lapujade, Pascale; Pronk, Jack T.; van Maris, Antonius J.A.; Daran, Jean-Marc G.

    2015-01-01

    A variety of techniques for strain engineering in Saccharomyces cerevisiae have recently been developed. However, especially when multiple genetic manipulations are required, strain construction is still a time-consuming process. This study describes new CRISPR/Cas9-based approaches for easy, fast strain construction in yeast and explores their potential for simultaneous introduction of multiple genetic modifications. An open-source tool (http://yeastriction.tnw.tudelft.nl) is presented for identification of suitable Cas9 target sites in S. cerevisiae strains. A transformation strategy, using in vivo assembly of a guideRNA plasmid and subsequent genetic modification, was successfully implemented with high accuracies. An alternative strategy, using in vitro assembled plasmids containing two gRNAs, was used to simultaneously introduce up to six genetic modifications in a single transformation step with high efficiencies. Where previous studies mainly focused on the use of CRISPR/Cas9 for gene inactivation, we demonstrate the versatility of CRISPR/Cas9-based engineering of yeast by achieving simultaneous integration of a multigene construct combined with gene deletion and the simultaneous introduction of two single-nucleotide mutations at different loci. Sets of standardized plasmids, as well as the web-based Yeastriction target-sequence identifier and primer-design tool, are made available to the yeast research community to facilitate fast, standardized and efficient application of the CRISPR/Cas9 system. PMID:25743786

  13. Alteration of mitochondrial oxidative phosphorylation in aged skeletal muscle involves modification of adenine nucleotide translocator.

    PubMed

    Gouspillou, Gilles; Bourdel-Marchasson, Isabelle; Rouland, Richard; Calmettes, Guillaume; Franconi, Jean-Michel; Deschodt-Arsac, Véronique; Diolez, Philippe

    2010-02-01

    The process of skeletal muscle aging is characterized by a progressive loss of muscle mass and functionality. The underlying mechanisms are highly complex and remain unclear. This study was designed to further investigate the consequences of aging on mitochondrial oxidative phosphorylation in rat gastrocnemius muscle, by comparing young (6 months) and aged (21 months) rats. Maximal oxidative phosphorylation capacity was clearly reduced in older rats, while mitochondrial efficiency was unaffected. Inner membrane properties were unaffected in aged rats since proton leak kinetics were identical to young rats. Application of top-down control analysis revealed a dysfunction of the phosphorylation module in older rats, responsible for a dysregulation of oxidative phosphorylation under low activities close to in vivo ATP turnover. This dysregulation is responsible for an impaired mitochondrial response toward changes in cellular ATP demand, leading to a decreased membrane potential which may in turn affect ROS production and ion homeostasis. Based on our data, we propose that modification of ANT properties with aging could partly explain these mitochondrial dysfunctions.

  14. Histone Modification Is Involved in Okadaic Acid (OA) Induced DNA Damage Response and G2-M Transition Arrest in Maize

    PubMed Central

    Zhang, Hao; Wang, Pu; Hou, Haoli; Wen, Huan; Zhou, Hong; Gao, Fei; Wu, Jinping; Qiu, Zhengming; Li, Lijia

    2016-01-01

    Histone modifications are involved in regulation of chromatin structure. To investigate the relationship between chromatin modification and cell cycle regulation during plant cell proliferation, Okadaic acid (OA), a specific inhibitor of serine/threonine protein phosphatase, was applied in this study. The results showed that OA caused the cell cycle arrest at preprophase, leading to seedling growth inhibition. Western blotting assay revealed that the spatial distribution of phosphorylation of Ser10 histone H3 tails (H3S10ph) signals was altered under OA treatment. Reactive oxygen species (ROS) was found to be at higher levels and TdT-mediated dUTP nick end labeling (TUNEL) assay displayed DNA breaks happened at the chromatin after treatment with OA, companied with an increase in the acetylation of histone H4 at lysine 5 (H4K5ac) level. From these observations, we speculated that the alteration of the spatial distribution of H3S10ph and the level of H4K5ac was involved in the procedure that OA induced DNA breaks and G2-M arrested by the accumulation of ROS, and that the histone H3S10ph and H4K5ac might facilitate DNA repair by their association with the chromatin decondensation. PMID:27196101

  15. Prediction of novel families of enzymes involved in oxidative and other complex modifications of bases in nucleic acids.

    PubMed

    Iyer, Lakshminarayan M; Tahiliani, Mamta; Rao, Anjana; Aravind, L

    2009-06-01

    Modified bases in nucleic acids present a layer of information that directs biological function over and beyond the coding capacity of the conventional bases. While a large number of modified bases have been identified, many of the enzymes generating them still remain to be discovered. Recently, members of the 2-oxoglutarate- and iron(II)-dependent dioxygenase super-family, which modify diverse substrates from small molecules to biopolymers, were predicted and subsequently confirmed to catalyze oxidative modification of bases in nucleic acids. Of these, two distinct families, namely the AlkB and the kinetoplastid base J binding proteins (JBP) catalyze in situ hydroxylation of bases in nucleic acids. Using sensitive computational analysis of sequences, structures and contextual information from genomic structure and protein domain architectures, we report five distinct families of 2-oxoglutarate- and iron(II)-dependent dioxygenase that we predict to be involved in nucleic acid modifications. Among the DNA-modifying families, we show that the dioxygenase domains of the kinetoplastid base J-binding proteins belong to a larger family that includes the Tet proteins, prototyped by the human oncogene Tet1, and proteins from basidiomycete fungi, chlorophyte algae, heterolobosean amoeboflagellates and bacteriophages. We present evidence that some of these proteins are likely to be involved in oxidative modification of the 5-methyl group of cytosine leading to the formation of 5-hydroxymethylcytosine. The Tet/JBP homologs from basidiomycete fungi such as Laccaria and Coprinopsis show large lineage-specific expansions and a tight linkage with genes encoding a novel and distinct family of predicted transposases, and a member of the Maelstrom-like HMG family. We propose that these fungal members are part of a mobile transposon. To the best of our knowledge, this is the first report of a eukaryotic transposable element that encodes its own DNA-modification enzyme with a

  16. Involvement of Genetic and Environmental Factors in the Onset of Depression

    PubMed Central

    Kim, Hyoung-Chun

    2013-01-01

    First, this article provides a brief overview of the previous hypotheses regarding depression and then focuses on involvement of genetic and environmental factors in development of depression. According to epidemiological research, 30~40% of occurrences of bipolar disorder involve a genetic factor. Therefore, environmental factors play a more important role in development of depression. Resilience and resistance to stress are common; therefore, although a certain extent of stress might be received during the embryonic or perinatal period, having a genetic predisposition to mental disorders does not imply that a mental disorder will develop. However, having a genetic predisposition to disorders does weaken resistance to stresses received during puberty, and without the ability to recover, a mental disorder is triggered. The importance of epigenetics in maintaining normal development and biology is reflected by the observation that development of many diseases occurs when the wrong type of epigenetic marks are introduced or are added at the wrong time or in the wrong place. Involvement of genetic and environmental factors in the onset of depression was investigated in relation to epigenetics. When mice with the disrupted in schizophrenia 1 (DISC1) abnormal gene received isolated rearing stress, depression-like abnormal behaviors and decreased gene expression of tyrosine hydroxylase in the frontal cortex by epigenetical suppression via DNA methylation were observed. Decrease of dopamine in the frontal cortex triggers behavioral disorders. Administration of a glucocorticoid receptor antagonist resulted in full recovery from neurological and behavioral disorders. These results suggest a new therapeutic approach to depression. PMID:24465138

  17. Does Religious Involvement Protect against Early Drinking? A Behavior Genetic Approach

    ERIC Educational Resources Information Center

    Harden, K. Paige

    2010-01-01

    Background: Adolescent involvement in religious organizations has been hypothesized to protect against early age at first drink. However, the correlation between adolescent religiosity and later age at first drink may be confounded by environmental or genetic differences between families. This study tests whether, after controlling for shared…

  18. [Genetic and biochemical mechanisms of involvement of antioxidant defense enzymes in the development of bronchial asthma].

    PubMed

    Polonikov, A V; Ivanov, V P; Bogomazov, A D; Solodilova, M A

    2015-01-01

    In the present review we have analyzed and summarized recent literature data on genetic and biochemical mechanisms responsible for involvement of antioxidant defense enzymes in the etiology and pathogenesis of bronchial asthma. It has been shown that the mechanisms of asthma development are linked with genetically determined abnormalities in the functioning of antioxidant defense enzymes. These alterations are accompanied by a systemic imbalance between oxidative and anti-oxidative reactions with the shift of the redox state toward increased free radical production and oxidative stress, a key element in the pathogenesis of bronchial asthma. PMID:26350733

  19. [Genetic and biochemical mechanisms of involvement of antioxidant defense enzymes in the development of bronchial asthma].

    PubMed

    Polonikov, A V; Ivanov, V P; Bogomazov, A D; Solodilova, M A

    2015-01-01

    In the present review we have analyzed and summarized recent literature data on genetic and biochemical mechanisms responsible for involvement of antioxidant defense enzymes in the etiology and pathogenesis of bronchial asthma. It has been shown that the mechanisms of asthma development are linked with genetically determined abnormalities in the functioning of antioxidant defense enzymes. These alterations are accompanied by a systemic imbalance between oxidative and anti-oxidative reactions with the shift of the redox state toward increased free radical production and oxidative stress, a key element in the pathogenesis of bronchial asthma.

  20. Parent involvement, sibling companionship, and adolescent substance use: A longitudinal, genetically informed design.

    PubMed

    Samek, Diana R; Rueter, Martha A; Keyes, Margaret A; McGue, Matt; Iacono, William G

    2015-08-01

    A large literature shows that parent and sibling relationship factors are associated with an increased likelihood of adolescent substance use. Less is known about the etiology of these associations. Using a genetically informed sibling design, we examined the prospective associations between parent involvement, sibling companionship, and adolescent substance use at 2 points in mid- and late-adolescence. Adolescents were adopted (n = 568) or the biological offspring of both parents (n = 412). Cross-lagged panel results showed that higher levels of parent involvement in early adolescence were associated with lower levels of substance use later in adolescence. Results did not significantly differ across adoption status, suggesting this association cannot be due to passive gene-environment correlation. Adolescent substance use at Time 1 was not significantly associated with parent involvement at Time 2, suggesting this association does not appear to be solely due to evocative (i.e., "child-driven") effects either. Together, results support a protective influence of parent involvement on subsequent adolescent substance use that is environmental in nature. The cross-paths between sibling companionship and adolescent substance use were significant and negative in direction (i.e., protective) for sisters, but positive for brothers (in line with a social contagion hypothesis). These effects were consistent across genetically related and unrelated pairs, and thus appear to be environmentally mediated. For mixed gender siblings, results were consistent with environmentally driven, protective influence hypothesis for genetically unrelated pairs, but in line with a genetically influenced, social contagion hypothesis for genetically related pairs. Implications are discussed. PMID:26030026

  1. 3D cultures of human neural progenitor cells: dopaminergic differentiation and genetic modification. [corrected].

    PubMed

    Brito, Catarina; Simão, Daniel; Costa, Inês; Malpique, Rita; Pereira, Cristina I; Fernandes, Paulo; Serra, Margarida; Schwarz, Sigrid C; Schwarz, Johannes; Kremer, Eric J; Alves, Paula M

    2012-03-01

    Central nervous system (CNS) disorders remain a formidable challenge for the development of efficient therapies. Cell and gene therapy approaches are promising alternatives that can have a tremendous impact by treating the causes of the disease rather than the symptoms, providing specific targeting and prolonged duration of action. Hampering translation of gene-based therapeutic treatments of neurodegenerative diseases from experimental to clinical gene therapy is the lack of valid and reliable pre-clinical models that can contribute to evaluate feasibility and safety. Herein we describe a robust and reproducible methodology for the generation of 3D in vitro models of the human CNS following a systematic technological approach based on stirred culture systems. We took advantage of human midbrain-derived neural progenitor cells (hmNPCs) capability to differentiate into the various neural phenotypes and of their commitment to the dopaminergic lineage to generate differentiated neurospheres enriched in dopaminergic neurons. Furthermore, we describe a protocol for efficient gene transfer into differentiated neurospheres using CAV-2 viral vectors and stable expression of the transgene for at least 10 days. CAV-2 vectors, derived from canine adenovirus type 2, are promising tools to understand and treat neurodegenerative diseases, in particular Parkinson's disease. CAV-2 vectors preferentially transduce neurons and have an impressive level of axonal retrograde transport in vivo. Our model provides a practical and versatile in vitro approach to study the CNS in a 3D cellular context. With the successful differentiation and subsequent genetic modification of neurospheres we are increasing the collection of tools available for neuroscience research and contributing for the implementation and widespread utilization of 3D cellular CNS models. These can be applied to study neurodegenerative diseases such as Parkinson's disease; to study the interaction of viral vectors of

  2. 3D cultures of human neural progenitor cells: dopaminergic differentiation and genetic modification. [corrected].

    PubMed

    Brito, Catarina; Simão, Daniel; Costa, Inês; Malpique, Rita; Pereira, Cristina I; Fernandes, Paulo; Serra, Margarida; Schwarz, Sigrid C; Schwarz, Johannes; Kremer, Eric J; Alves, Paula M

    2012-03-01

    Central nervous system (CNS) disorders remain a formidable challenge for the development of efficient therapies. Cell and gene therapy approaches are promising alternatives that can have a tremendous impact by treating the causes of the disease rather than the symptoms, providing specific targeting and prolonged duration of action. Hampering translation of gene-based therapeutic treatments of neurodegenerative diseases from experimental to clinical gene therapy is the lack of valid and reliable pre-clinical models that can contribute to evaluate feasibility and safety. Herein we describe a robust and reproducible methodology for the generation of 3D in vitro models of the human CNS following a systematic technological approach based on stirred culture systems. We took advantage of human midbrain-derived neural progenitor cells (hmNPCs) capability to differentiate into the various neural phenotypes and of their commitment to the dopaminergic lineage to generate differentiated neurospheres enriched in dopaminergic neurons. Furthermore, we describe a protocol for efficient gene transfer into differentiated neurospheres using CAV-2 viral vectors and stable expression of the transgene for at least 10 days. CAV-2 vectors, derived from canine adenovirus type 2, are promising tools to understand and treat neurodegenerative diseases, in particular Parkinson's disease. CAV-2 vectors preferentially transduce neurons and have an impressive level of axonal retrograde transport in vivo. Our model provides a practical and versatile in vitro approach to study the CNS in a 3D cellular context. With the successful differentiation and subsequent genetic modification of neurospheres we are increasing the collection of tools available for neuroscience research and contributing for the implementation and widespread utilization of 3D cellular CNS models. These can be applied to study neurodegenerative diseases such as Parkinson's disease; to study the interaction of viral vectors of

  3. Modifications of a conserved regulatory network involving INDEHISCENT controls multiple aspects of reproductive tissue development in Arabidopsis.

    PubMed

    Kay, P; Groszmann, M; Ross, J J; Parish, R W; Swain, S M

    2013-01-01

    Disrupting pollen tube growth and fertilization in Arabidopsis plants leads to reduced seed set and silique size, providing a powerful genetic system with which to identify genes with important roles in plant fertility. A transgenic Arabidopsis line with reduced pollen tube growth, seed set and silique growth was used as the progenitor in a genetic screen to isolate suppressors with increased seed set and silique size. This screen generated a new allele of INDEHISCENT (IND), a gene originally identified by its role in valve margin development and silique dehiscence (pod shatter). IND forms part of a regulatory network that involves several other transcriptional regulators and involves the plant hormones GA and auxin. Using GA and auxin mutants that alter various aspects of reproductive development, we have identified novel roles for IND, its paralogue HECATE3, and the MADS box proteins SHATTERPROOF1/2 in flower and fruit development. These results suggest that modified forms of the regulatory network originally described for the Arabidopsis valve margin, which include these genes and/or their recently evolved paralogs, function in multiple components of GA/auxin-regulated reproductive development. PMID:23126654

  4. Genetic polymorphism of serotonin transporter 5-HTTLPR: involvement in smoking behaviour.

    PubMed

    Watanabe, Maria Angelica Ehara; Nunes, Sandra Odebrecht Vargas; Nunes, Sandra Odebrechet Vargas; Amarante, Marla Karine; Guembarovski, Roberta Losi; Oda, Julie Massayo Maeda; Lima, Kalil William Alves De; Fungaro, Maria Helena Pelegrinelli

    2011-04-01

    Data suggest that the serotonin (5-hydroxytryptamine, 5-HT) system is implicated in the pathogenesis of multiple neuropsychiatric disorders and may also be involved in smoking behaviour since nicotine increases brain serotonin secretion. It is known that smoking behaviour is influenced by both genetic and environmental factors. The present review examines the role of the serotonin transporter gene (5-HTT) in smoking behaviour and investigating studies that showed association of 5-HTT gene with smoking. This study discusses a polymorphism which has been investigated by many researchers, as the bi-allelic insertion/deletion polymorphism in the 5'- flanking promoter region (5-HTTLPR). This gene has received considerable attention in attempts to understand the molecular determinants of smoking. Therefore, in the present study, the relationship between genetic polymorphism of serotonin transporter in smoking behaviour is reviewed considering the interactive effect of genetic factors.

  5. Metabolic flux responses to genetic modification for shikimic acid production by Bacillus subtilis strains

    PubMed Central

    2014-01-01

    Background Shikimic acid (SA) is a key chiral starting molecule for the synthesis of the neuramidase inhibitor GS4104 against viral influenza. Microbial production of SA has been extensively investigated in Escherichia coli, and to a less extent in Bacillus subtilis. However, metabolic flux of the high SA-producing strains has not been explored. In this study, we constructed with genetic manipulation and further determined metabolic flux with 13C-labeling test of high SA-producing B. subtilis strains. Results B. subtilis 1A474 had a mutation in SA kinase gene (aroI) and accumulated 1.5 g/L of SA. Overexpression of plasmid-encoded aroA, aroB, aroC or aroD in B. subtilis revealed that aroD had the most significantly positive effects on SA production. Simultaneous overexpression of genes for 3-deoxy-D-arabinoheptulosonate-7-phosphate synthase (aroA) and SA dehydrogenase (aroD) in B. subtilis BSSA/pSAAroA/pDGSAAroD resulted in SA production of 3.2 g/L. 13C-Metabolic flux assay (MFA) on the two strains BSSA/pHCMC04/pDG148-stu and BSSA/pSAAroA/pDGSAAroD indicated the carbon flux from glucose to SA increased to 4.6% in BSSA/pSAAroA/pDGSAAroD from 1.9% in strain BSSA/pHCMC04/pDG148-stu. The carbon flux through tricarboxylic acid cycle significantly reduced, while responses of the pentose phosphate pathway and the glycolysis to high SA production were rather weak, in the strain BSSA/pSAAroA/pDGSAAroD. Based on the results from MFA, two potential targets for further optimization of SA production were identified. Experiments on genetic deletion of phosphoenoylpyruvate kinase gene confirmed its positive influence on SA production, while the overexpression of the transketolase gene did not lead to increase in SA production. Conclusion Of the genes involved in shikimate pathway in B. subtilis, aroD exerted most significant influence on SA accumulation. Overexpression of plasmid-encoded aroA and aroD doubled SA production than its parent strain. MFA revealed metabolic flux

  6. A strategy for genetic modification of the spike-encoding segment of human reovirus T3D for reovirus targeting.

    PubMed

    van den Wollenberg, D J M; van den Hengel, S K; Dautzenberg, I J C; Cramer, S J; Kranenburg, O; Hoeben, R C

    2008-12-01

    Human Orthoreovirus Type 3 Dearing is not pathogenic to humans and has been evaluated clinically as an oncolytic agent. Its transduction efficiency and the tumor cell selectivity may be enhanced by incorporating ligands for alternative receptors. However, the genetic modification of reoviruses has been difficult, and genetic targeting of reoviruses has not been reported so far. Here we describe a technique for generating genetically targeted reoviruses. The propagation of wild-type reoviruses on cells expressing a modified sigma 1-encoding segment embedded in a conventional RNA polymerase II transcript leads to substitution of the wild-type genome segment by the modified version. This technique was used for generating reoviruses that are genetically targeted to an artificial receptor expressed on U118MG cells. These cells lack the junction adhesion molecule-1 and therefore resist infection by wild-type reoviruses. The targeted reoviruses were engineered to carry the ligand for this receptor at the C terminus of the sigma 1 spike protein. This demonstrates that the C terminus of the sigma 1 protein is a suitable locale for the insertion of oligopeptide ligands and that targeting of reoviruses is feasible. The genetically targeted viruses can be propagated using the modified U118MG cells as helper cells. This technique may be applicable for the improvement of human reoviruses as oncolytic agents.

  7. Genetic modification of hypoxia signaling in animal models and its effect on cancer.

    PubMed

    García-Heredia, J M; Felipe-Abrio, B; Cano, D A; Carnero, A

    2015-02-01

    Conditions that cause hypoxemia or generalized tissue hypoxia, which can last for days, months, or even years, are very common in the human population and are among the leading causes of morbidity, disability, and mortality. Therefore, the molecular pathophysiology of hypoxia and its potential deleterious effects on human health are important issues at the forefront of biomedical research. Generalized hypoxia is a consequence of highly prevalent medical disorders that can severely reduce the capacity for O2 exchange between the air and pulmonary capillaries. In recent years, some of the key O2-dependent signaling pathways have been characterized at the molecular level. In particular, the prolyl hydroxylase (PHD)-hypoxia-inducible factor (HIF) cascade has emerged as the master regulator of a general gene expression program involved in cell/tissue/organ adaptation to hypoxia. Hypoxia has emerged as a critical factor in cancer because it can promote tumor initiation, progression, and resistance to therapy. Beyond its role in neovascularization as a mechanism of tumor adaptation to nutrient and O2 deprivation, hypoxia has been linked to prolonged cellular lifespan and immortalization, the generation of "oncometabolites", deregulation of stem cell proliferation, and inflammation, among other tumor hallmarks. Hypoxia may contribute to cancer through several independent pathways, the inter-connections of which have yet to be elucidated. Furthermore, the relevance of chronic hypoxemia in the initiation and progression of cancer has not been studied in depth in the whole organism. Therefore, we explore here the contributions of hypoxia to the whole organism by reviewing studies on genetically modified mice with alterations in the key molecular factors regulating hypoxia.

  8. Molecular Genetic Analysis of Activation-tagged Transcription Factors Thought to be Involved in Photomorphogenesis

    SciTech Connect

    Neff, Michael

    2011-06-23

    Plants utilize light as a source of information via families of photoreceptors such as the red/far-red absorbing phytochromes (PHY) and the blue/UVA absorbing cryptochromes (CRY). The main goal of the Neff lab is to use molecular-genetic mutant screens to elucidate signaling components downstream of these photoreceptors. Activation-tagging mutagenesis led to the identification of two putative transcription factors that may be involved in both photomorphogenesis and hormone signaling pathways. sob1-D (suppressor of phyB-dominant) mutant phenotypes are caused by the over-expression of a Dof transcription factor previously named OBP3. Our previous studies indicate that OBP3 is a negative regulator of light-mediated cotyledon expansion and may be involved in modulating responsiveness to the growth-regulating hormone auxin. The sob2-D mutant uncovers a role for LEP, a putative AP2/EREBP-like transcription factor, in seed germination, hypocotyl elongation and responsiveness to the hormone abscisic acid. Based on photobiological and genetic analysis of OBP3-knockdown and LEP-null mutations, we hypothesize that these transcription factors are involved in both light-mediated seedling development and hormone signaling. To examine the role that these genes play in photomorphogenesis we will: 1) Further explore the genetic role of OBP3 in cotyledon/leaf expansion and other photomorphogenic processes as well as examine potential physical interactions between OBP3 and CRY1 or other signaling components that genetically interact with this transcription factor 2) Test the hypothesis that OBP3 is genetically involved in auxin signaling and root development as well as examine the affects of this hormone and light on OBP3 protein accumulation. 3) Test the hypothesis that LEP is involved in seed germination, seedling photomorphogenesis and hormone signaling. Together these experiments will lead to a greater understanding of the complexity of interactions between photoreceptors and DNA

  9. A genetic system to assess in vivo the functions of histones and histone modifications in higher eukaryotes.

    PubMed

    Günesdogan, Ufuk; Jäckle, Herbert; Herzig, Alf

    2010-10-01

    Despite the fundamental role of canonical histones in nucleosome structure, there is no experimental system for higher eukaryotes in which basic questions about histone function can be directly addressed. We developed a new genetic tool for Drosophila melanogaster in which the canonical histone complement can be replaced with multiple copies of experimentally modified histone transgenes. This new histone-replacement system provides a well-defined and direct cellular assay system for histone function with which to critically test models in chromatin biology dealing with chromatin assembly, variant histone functions and the biological significance of distinct histone modifications in a multicellular organism.

  10. Increasing public involvement in enriching our fish stocks through genetic enhancement.

    PubMed

    Halvorson, H O; Quezada, F

    1999-11-01

    A total of 70%, of the world's conventional commercial fish species are now fully exploited, overexploited, depleted or recovering from depletion. This dramatic crash in the capture world fisheries production has led to problems in foods distribution, balance of payments, employment, and ecological depletion. Public support for breeding programs with terrestrial farm animals and plants in agriculture have revolutionized this industry over the past few hundred years. However, new genetic rearing technologies to improve marine animal production through aquaculture that utilize modern biology to obtain sustainable aquaculture and preserve biodiversity provide a promise to address these problems. However aquaculture has not been subject to public discussion and approval. Public involvement, not necessarily acquiescence, provide value added in the decision making process. Public understanding and involvement involves three stages. (i) Public concern over the pool of genetic information; (ii) if aquaculture is to respond to the fisheries crises with innovation, the knowledge gap between public understanding and scientific information must be bridged; and (iii) strategies must be developed for achieving this. Release of recombinant DNA to the environment, and handling exotic species, are useful case studies. Illustrations will be given of communication bridges to the public and ways to involve the public in making policy decisions.

  11. Possible modification of Alzheimer's disease by statins in midlife: interactions with genetic and non-genetic risk factors.

    PubMed

    Shinohara, Mitsuru; Sato, Naoyuki; Shimamura, Munehisa; Kurinami, Hitomi; Hamasaki, Toshimitsu; Chatterjee, Amarnath; Rakugi, Hiromi; Morishita, Ryuichi

    2014-01-01

    The benefits of statins, commonly prescribed for hypercholesterolemia, in treating Alzheimer's disease (AD) have not yet been fully established. A recent randomized clinical trial did not show any therapeutic effects of two statins on cognitive function in AD. Interestingly, however, the results of the Rotterdam study, one of the largest prospective cohort studies, showed reduced risk of AD in statin users. Based on the current understanding of statin actions and AD pathogenesis, it is still worth exploring whether statins can prevent AD when administered decades before the onset of AD or from midlife. This review discusses the possible beneficial effects of statins, drawn from previous clinical observations, pathogenic mechanisms, which include β-amyloid (Aβ) and tau metabolism, genetic and non-genetic risk factors (apolipoprotein E, cholesterol, sex, hypertension, and diabetes), and other clinical features (vascular dysfunction and oxidative and inflammatory stress) of AD. These findings suggest that administration of statins in midlife might prevent AD in late life by modifying genetic and non-genetic risk factors for AD. It should be clarified whether statins inhibit Aβ accumulation, tau pathological features, and brain atrophy in humans. To answer this question, a randomized controlled study using amyloid positron emission tomography (PET), tau-PET, and magnetic resonance imaging would be useful. This clinical evaluation could help us to overcome this devastating disease.

  12. The Functional Significance of Posttranslational Modifications on Polo-Like Kinase 1 Revealed by Chemical Genetic Complementation.

    PubMed

    Lasek, Amber L; McPherson, Brittany M; Trueman, Natalie G; Burkard, Mark E

    2016-01-01

    Mitosis is coordinated by carefully controlled phosphorylation and ubiquitin-mediated proteolysis. Polo-like kinase 1 (Plk1) plays a central role in regulating mitosis and cytokinesis by phosphorylating target proteins. Yet, Plk1 is itself a target for posttranslational modification by phosphorylation and ubiquitination. We developed a chemical-genetic complementation assay to evaluate the functional significance of 34 posttranslational modifications (PTMs) on human Plk1. To do this, we used human cells that solely express a modified analog-sensitive Plk1 (Plk1AS) and complemented with wildtype Plk1. The wildtype Plk1 provides cells with a functional Plk1 allele in the presence of 3-MB-PP1, a bulky ATP-analog inhibitor that specifically inhibits Plk1AS. Using this approach, we evaluated the ability of 34 singly non-modifiable Plk1 mutants to complement Plk1AS in the presence of 3-MB-PP1. Mutation of the T-loop activating residue T210 and adjacent T214 are lethal, but surprisingly individual mutation of the remaining 32 posttranslational modification sites did not disrupt the essential functions of Plk1. To evaluate redundancy, we simultaneously mutated all phosphorylation sites in the kinase domain except for T210 and T214 or all sites in the C-terminal polo-box domain (PBD). We discovered that redundant phosphorylation events within the kinase domain are required for accurate chromosome segregation in anaphase but those in the PBD are dispensable. We conclude that PTMs within the T-loop of Plk1 are essential and nonredundant, additional modifications in the kinase domain provide redundant control of Plk1 function, and those in the PBD are dispensable for essential mitotic functions of Plk1. This comprehensive evaluation of Plk1 modifications demonstrates that although phosphorylation and ubiquitination are important for mitotic progression, many individual PTMs detected in human tissue may have redundant, subtle, or dispensable roles in gene function. PMID

  13. The Functional Significance of Posttranslational Modifications on Polo-Like Kinase 1 Revealed by Chemical Genetic Complementation

    PubMed Central

    Lasek, Amber L.; McPherson, Brittany M.; Trueman, Natalie G.; Burkard, Mark E.

    2016-01-01

    Mitosis is coordinated by carefully controlled phosphorylation and ubiquitin-mediated proteolysis. Polo-like kinase 1 (Plk1) plays a central role in regulating mitosis and cytokinesis by phosphorylating target proteins. Yet, Plk1 is itself a target for posttranslational modification by phosphorylation and ubiquitination. We developed a chemical-genetic complementation assay to evaluate the functional significance of 34 posttranslational modifications (PTMs) on human Plk1. To do this, we used human cells that solely express a modified analog-sensitive Plk1 (Plk1AS) and complemented with wildtype Plk1. The wildtype Plk1 provides cells with a functional Plk1 allele in the presence of 3-MB-PP1, a bulky ATP-analog inhibitor that specifically inhibits Plk1AS. Using this approach, we evaluated the ability of 34 singly non-modifiable Plk1 mutants to complement Plk1AS in the presence of 3-MB-PP1. Mutation of the T-loop activating residue T210 and adjacent T214 are lethal, but surprisingly individual mutation of the remaining 32 posttranslational modification sites did not disrupt the essential functions of Plk1. To evaluate redundancy, we simultaneously mutated all phosphorylation sites in the kinase domain except for T210 and T214 or all sites in the C-terminal polo-box domain (PBD). We discovered that redundant phosphorylation events within the kinase domain are required for accurate chromosome segregation in anaphase but those in the PBD are dispensable. We conclude that PTMs within the T-loop of Plk1 are essential and nonredundant, additional modifications in the kinase domain provide redundant control of Plk1 function, and those in the PBD are dispensable for essential mitotic functions of Plk1. This comprehensive evaluation of Plk1 modifications demonstrates that although phosphorylation and ubiquitination are important for mitotic progression, many individual PTMs detected in human tissue may have redundant, subtle, or dispensable roles in gene function. PMID

  14. Efficient marker-free recovery of custom genetic modifications with CRISPR/Cas9 in Caenorhabditis elegans.

    PubMed

    Arribere, Joshua A; Bell, Ryan T; Fu, Becky X H; Artiles, Karen L; Hartman, Phil S; Fire, Andrew Z

    2014-11-01

    Facilitated by recent advances using CRISPR/Cas9, genome editing technologies now permit custom genetic modifications in a wide variety of organisms. Ideally, modified animals could be both efficiently made and easily identified with minimal initial screening and without introducing exogenous sequence at the locus of interest or marker mutations elsewhere. To this end, we describe a coconversion strategy, using CRISPR/Cas9 in which screening for a dominant phenotypic oligonucleotide-templated conversion event at one locus can be used to enrich for custom modifications at another unlinked locus. After the desired mutation is identified among the F1 progeny heterozygous for the dominant marker mutation, F2 animals that have lost the marker mutation are picked to obtain the desired mutation in an unmarked genetic background. We have developed such a coconversion strategy for Caenorhabditis elegans, using a number of dominant phenotypic markers. Examining the coconversion at a second (unselected) locus of interest in the marked F1 animals, we observed that 14-84% of screened animals showed homologous recombination. By reconstituting the unmarked background through segregation of the dominant marker mutation at each step, we show that custom modification events can be carried out recursively, enabling multiple mutant animals to be made. While our initial choice of a coconversion marker [rol-6(su1006)] was readily applicable in a single round of coconversion, the genetic properties of this locus were not optimal in that CRISPR-mediated deletion mutations at the unselected rol-6 locus can render a fraction of coconverted strains recalcitrant to further rounds of similar mutagenesis. An optimal marker in this sense would provide phenotypic distinctions between the desired mutant/+ class and alternative +/+, mutant/null, null/null, and null/+ genotypes. Reviewing dominant alleles from classical C. elegans genetics, we identified one mutation in dpy-10 and one mutation in

  15. Efficient marker-free recovery of custom genetic modifications with CRISPR/Cas9 in Caenorhabditis elegans.

    PubMed

    Arribere, Joshua A; Bell, Ryan T; Fu, Becky X H; Artiles, Karen L; Hartman, Phil S; Fire, Andrew Z

    2014-11-01

    Facilitated by recent advances using CRISPR/Cas9, genome editing technologies now permit custom genetic modifications in a wide variety of organisms. Ideally, modified animals could be both efficiently made and easily identified with minimal initial screening and without introducing exogenous sequence at the locus of interest or marker mutations elsewhere. To this end, we describe a coconversion strategy, using CRISPR/Cas9 in which screening for a dominant phenotypic oligonucleotide-templated conversion event at one locus can be used to enrich for custom modifications at another unlinked locus. After the desired mutation is identified among the F1 progeny heterozygous for the dominant marker mutation, F2 animals that have lost the marker mutation are picked to obtain the desired mutation in an unmarked genetic background. We have developed such a coconversion strategy for Caenorhabditis elegans, using a number of dominant phenotypic markers. Examining the coconversion at a second (unselected) locus of interest in the marked F1 animals, we observed that 14-84% of screened animals showed homologous recombination. By reconstituting the unmarked background through segregation of the dominant marker mutation at each step, we show that custom modification events can be carried out recursively, enabling multiple mutant animals to be made. While our initial choice of a coconversion marker [rol-6(su1006)] was readily applicable in a single round of coconversion, the genetic properties of this locus were not optimal in that CRISPR-mediated deletion mutations at the unselected rol-6 locus can render a fraction of coconverted strains recalcitrant to further rounds of similar mutagenesis. An optimal marker in this sense would provide phenotypic distinctions between the desired mutant/+ class and alternative +/+, mutant/null, null/null, and null/+ genotypes. Reviewing dominant alleles from classical C. elegans genetics, we identified one mutation in dpy-10 and one mutation in

  16. Systematic review of genetic association studies involving histologically confirmed non-alcoholic fatty liver disease

    PubMed Central

    Wood, Kayleigh L; Miller, Michael H; Dillon, John F

    2015-01-01

    Non-alcoholic fatty liver disease has an increasing prevalence in Western countries, affecting up to 20% of the population. Objective The aim of this project was to systematically review and summarise the genetic association studies that investigate possible genetic influences that confer susceptibility to non-alcoholic fatty liver disease and non-alcoholic steatohepatitis. Design The MEDLINE and SCOPUS databases were searched to identify candidate gene studies on histologically diagnosed non-alcoholic fatty liver disease. Results A total of 85 articles have been summarised and categorised on the basis of the general pathway each candidate gene is involved in, including lipid metabolism, lipoprotein processing, cholesterol synthesis, glucose homoeostasis, inflammatory response, protection against oxidative stress and whole body metabolism. Conclusions The main findings demonstrate a small but consistent association of PNPLA3 with non-alcoholic fatty liver disease and non-alcoholic steatohepatitis. Genetic association studies have investigated general disease susceptibility, histological characteristics, severity and progression. However, further study is required to better elucidate the genetic factors influencing fatty liver disease. PMID:26462272

  17. Genes and quantitative genetic variation involved with senescence in cells, organs, and the whole plant

    PubMed Central

    Pujol, Benoit

    2015-01-01

    Senescence, the deterioration of morphological, physiological, and reproductive functions with age that ends with the death of the organism, was widely studied in plants. Genes were identified that are linked to the deterioration of cells, organs and the whole plant. It is, however, unclear whether those genes are the source of age dependent deterioration or get activated to regulate such deterioration. Furthermore, it is also unclear whether such genes are active as a direct consequence of age or because they are specifically involved in some developmental stages. At the individual level, it is the relationship between quantitative genetic variation, and age that can be used to detect the genetic signature of senescence. Surprisingly, the latter approach was only scarcely applied to plants. This may be the consequence of the demanding requirements for such approaches and/or the fact that most research interest was directed toward plants that avoid senescence. Here, I review those aspects in turn and call for an integrative genetic theory of senescence in plants. Such conceptual development would have implications for the management of plant genetic resources and generate progress on fundamental questions raised by aging research. PMID:25755664

  18. Phenotypic and genotypic variation in methylases involved in type II restriction-modification systems in Helicobacter pylori

    PubMed Central

    Takata, Tohru; Aras, Rahul; Tavakoli, Donald; Ando, Takafumi; Olivares, Asalia Z.; Blaser, Martin J.

    2002-01-01

    To determine relationships between Helicobacter pylori geographical origin and type II methylase activity, we examined 122 strains from various locations around the world for methylase expression. Most geographic regions possessed at least one strain resistant to digestion by each of 14 restriction endonucleases studied. Across all of the strains studied, the average number of active methylases was 8.2 ± 1.9 with no significant variation between the major geographic regions. Although seven pairs of isolates showed the same susceptibility patterns, their cagA/vacA status differed, and the remaining 108 strains each possessed unique patterns of susceptibility. From a single clonal group, 15 of 18 strains showed identical patterns of resistance, but diverged with respect to M.MboII activity. All of the methylases studied were present in all major human population groupings, suggesting that their horizontal acquisition pre-dated the separation of these populations. For the hpyV and hpyAIV restriction-modification systems, an in-depth analysis of genotype, indicating extensive diversity of cassette size and chromosomal locations regardless of the susceptibility phenotype, points toward substantial strain-specific selection involving these loci. PMID:12034832

  19. Gene flow in genetically engineered perennial grasses: Lessons for modification of dedicated bioenergy crops

    EPA Science Inventory

    The potential ecological consequences of the commercialization of genetically engineered (GD) crops have been the subject of intense debate, particularly when the GE crops are perennial and capable of outcrossing to wild relatives. The essential ecological impact issues for engi...

  20. Genetically engineered charge modifications to enhance protein separation in aqueous two-phase systems: Charge directed partitioning

    SciTech Connect

    Luther, J.R.; Glatz, C.E.

    1995-04-05

    This report continues the authors` examination of the effect of genetically engineered charge modifications on the partitioning behavior of proteins in aqueous two-phase extraction. The genetic modifications consisted of the fusion of charged peptide tails to {beta}-galactosidase and charge-change point mutations to T4 lysozyme. In this study, they examined charge directed partitioning behavior in PEG/dextran systems containing small amounts of the charged polymers diethylaminoethyl-dextran (DEAE-dextran) or dextran sulfate. The best results were obtained when attractive forces between the protein and polymer were present. Nearly 100% of the {beta}-galactosidase, which carries a net negative charge, partitioned to the DEAE-dextran-rich phase regardless of whether the phase was dextran or PEG. In these cases, cloudiness of the protein-rich phases suggest that strong charge interactions resulted in protein/polymer aggregation, which may have contribution to the extreme partitioning. Unlike the potential-driven partitioning reported previously, consistent partitioning trends were observed as a result of the fusion tails, with observed shifts in partition coefficient (K{sub p}) of up to 37-fold. However, these changes could not be solely attributed to charge-based interactions.

  1. Identification of plant genetic loci involved in a posttranscriptional mechanism for meiotically reversible transgene silencing.

    PubMed Central

    Dehio, C; Schell, J

    1994-01-01

    Numerous reports describe phenomena of transgene silencing in plants, yet the underlying genetic and molecular mechanisms are poorly understood. We observed that regeneration of Arabidopsis thaliana plants transgenic for the rolB gene of Agrobacterium rhizogenes results in a selection for transgene silencing. Transgene silencing could be monitored in this system by reversion of the visible RolB phenotype. We report a phenotypic, molecular, and genetic characterization of a meiotically reversible transgene silencing phenomenon observed in a rolB transgenic line. In this line, the rolB gene is expressed strongly and uniformly in seedlings, but in the course of further development, the rolB gene is silenced erratically at a frequency that depends on the dosage of rolB. The silenced state is mitotically stable, while complete resetting of rolB gene expression occurs in seedlings of the following generation. The silencing of rolB correlates with a dramatic reduction of steady-state rolB transcripts, while rolB nuclear run-off transcripts are only moderately reduced. Therefore, rolB gene silencing seems to act predominantly at the posttranscriptional level. The process of rolB gene silencing was found to be affected by two extragenic modifier loci that influence both the frequency and the timing of rolB gene silencing during plant development. These genetic data demonstrate a direct involvement of defined plant genes in this form of gene silencing. Images PMID:8202523

  2. Genetic, metabolic and environmental factors involved in the development of liver cirrhosis in Mexico

    PubMed Central

    Ramos-Lopez, Omar; Martinez-Lopez, Erika; Roman, Sonia; Fierro, Nora A; Panduro, Arturo

    2015-01-01

    Liver cirrhosis (LC) is a chronic illness caused by inflammatory responses and progressive fibrosis. Globally, the most common causes of chronic liver disease include persistent alcohol abuse, followed by viral hepatitis infections and nonalcoholic fatty liver disease. However, regardless of the etiological factors, the susceptibility and degree of liver damage may be influenced by genetic polymorphisms that are associated with distinct ethnic and cultural backgrounds. Consequently, metabolic genes are influenced by variable environmental lifestyle factors, such as diet, physical inactivity, and emotional stress, which are associated with regional differences among populations. This Topic Highlight will focus on the genetic and environmental factors that may influence the metabolism of alcohol and nutrients in the setting of distinct etiologies of liver disease. The interaction between genes and environment in the current-day admixed population, Mestizo and Native Mexican, will be described. Additionally, genes involved in immune regulation, insulin sensitivity, oxidative stress and extracellular matrix deposition may modulate the degree of severity. In conclusion, LC is a complex disease. The onset, progression, and clinical outcome of LC among the Mexican population are influenced by specific genetic and environmental factors. Among these are an admixed genome with a heterogenic distribution of European, Amerindian and African ancestry; a high score of alcohol consumption; viral infections; a hepatopathogenic diet; and a high prevalence of obesity. The variance in risk factors among populations suggests that intervention strategies directed towards the prevention and management of LC should be tailored according to such population-based features. PMID:26556986

  3. Gene discovery for enzymes involved in limonene modification or utilization by the mountain pine beetle-associated pathogen Grosmannia clavigera.

    PubMed

    Wang, Ye; Lim, Lynette; Madilao, Lina; Lah, Ljerka; Bohlmann, Joerg; Breuil, Colette

    2014-08-01

    To successfully colonize and eventually kill pine trees, Grosmannia clavigera (Gs cryptic species), the main fungal pathogen associated with the mountain pine beetle (Dendroctonus ponderosae), has developed multiple mechanisms to overcome host tree chemical defenses, of which terpenoids are a major component. In addition to a monoterpene efflux system mediated by a recently discovered ABC transporter, Gs has genes that are highly induced by monoterpenes and that encode enzymes that modify or utilize monoterpenes [especially (+)-limonene]. We showed that pine-inhabiting Ophiostomale fungi are tolerant to monoterpenes, but only a few, including Gs, are known to utilize monoterpenes as a carbon source. Gas chromatography-mass spectrometry (GC-MS) revealed that Gs can modify (+)-limonene through various oxygenation pathways, producing carvone, p-mentha-2,8-dienol, perillyl alcohol, and isopiperitenol. It can also degrade (+)-limonene through the C-1-oxygenated pathway, producing limonene-1,2-diol as the most abundant intermediate. Transcriptome sequencing (RNA-seq) data indicated that Gs may utilize limonene 1,2-diol through beta-oxidation and then valine and tricarboxylic acid (TCA) metabolic pathways. The data also suggested that at least two gene clusters, located in genome contigs 108 and 161, were highly induced by monoterpenes and may be involved in monoterpene degradation processes. Further, gene knockouts indicated that limonene degradation required two distinct Baeyer-Villiger monooxygenases (BVMOs), an epoxide hydrolase and an enoyl coenzyme A (enoyl-CoA) hydratase. Our work provides information on enzyme-mediated limonene utilization or modification and a more comprehensive understanding of the interaction between an economically important fungal pathogen and its host's defense chemicals.

  4. Gene Discovery for Enzymes Involved in Limonene Modification or Utilization by the Mountain Pine Beetle-Associated Pathogen Grosmannia clavigera

    PubMed Central

    Wang, Ye; Lim, Lynette; Madilao, Lina; Lah, Ljerka; Bohlmann, Joerg

    2014-01-01

    To successfully colonize and eventually kill pine trees, Grosmannia clavigera (Gs cryptic species), the main fungal pathogen associated with the mountain pine beetle (Dendroctonus ponderosae), has developed multiple mechanisms to overcome host tree chemical defenses, of which terpenoids are a major component. In addition to a monoterpene efflux system mediated by a recently discovered ABC transporter, Gs has genes that are highly induced by monoterpenes and that encode enzymes that modify or utilize monoterpenes [especially (+)-limonene]. We showed that pine-inhabiting Ophiostomale fungi are tolerant to monoterpenes, but only a few, including Gs, are known to utilize monoterpenes as a carbon source. Gas chromatography-mass spectrometry (GC-MS) revealed that Gs can modify (+)-limonene through various oxygenation pathways, producing carvone, p-mentha-2,8-dienol, perillyl alcohol, and isopiperitenol. It can also degrade (+)-limonene through the C-1-oxygenated pathway, producing limonene-1,2-diol as the most abundant intermediate. Transcriptome sequencing (RNA-seq) data indicated that Gs may utilize limonene 1,2-diol through beta-oxidation and then valine and tricarboxylic acid (TCA) metabolic pathways. The data also suggested that at least two gene clusters, located in genome contigs 108 and 161, were highly induced by monoterpenes and may be involved in monoterpene degradation processes. Further, gene knockouts indicated that limonene degradation required two distinct Baeyer-Villiger monooxygenases (BVMOs), an epoxide hydrolase and an enoyl coenzyme A (enoyl-CoA) hydratase. Our work provides information on enzyme-mediated limonene utilization or modification and a more comprehensive understanding of the interaction between an economically important fungal pathogen and its host's defense chemicals. PMID:24837377

  5. Guided implant surgery with modification of the technique involving the raising of a semicircular miniflap: A preliminary study

    PubMed Central

    Viña, José; Maestre, Laura; Peñarrocha, David; Balaguer, José

    2012-01-01

    Objective: An evaluation is made of pain, swelling and peri-implant attached mucosal width after implant-based rehabilitation involving guided surgery and a modification of the technique with the raising of a semicircular miniflap, in single and partial replacements. Study design: A case-control study was carried out. The study group consisted of 12 patients with the placement of 19 implants using a guided surgery and miniflap technique. The control group consisted of 12 patients with the placement of 22 implants using the conventional technique. Each patient scored postoperative swelling and pain by means of a visual analog scale (VAS). Attached vestibular mucosa width was evaluated 12 weeks after implant placement. Results: Twelve operations were carried out in each group. Immediate aesthetics were established for all implants of the study group. One implant failed in each group. Maximum pain was recorded after 6 hours in both groups (mean VAS score 4 and 4.9 in the study and control group, respectively). Maximum swelling was recorded after 24 hours (mean VAS score 2.5) in the study group and on the second day (mean VAS score 3.4) in the control group. The mean attached vestibular mucosa width was 2.9 mm in the study group and 3.2 mm in the control group. Conclusion: In this preliminary study, guided implant surgery with a semicircular miniflap in single and partial replacements resulted in slightly less postoperative pain and swelling than with the conventional implant technique. The attached vestibular mucosa width was greater in the control group, though the differences were very small. Key words:Guided surgery, flapless surgery, miniflap, peri-implant mucosa. PMID:22549666

  6. DNA Mapping Made Simple: An Intellectual Activity about the Genetic Modification of Organisms

    ERIC Educational Resources Information Center

    Marques, Miguel; Arrabaca, Joao; Chagas, Isabel

    2004-01-01

    Since the discovery of the DNA double helix (in 1953 by Watson and Crick), technologies have been developed that allow scientists to manipulate the genome of bacteria to produce human hormones, as well as the genome of crop plants to achieve high yield and enhanced flavor. The universality of the genetic code has allowed DNA isolated from a…

  7. Design of a novel molecular beacon: modification of the stem with artificially genetic alphabet.

    PubMed

    Sheng, Pinpin; Yang, Zunyi; Kim, Youngmi; Wu, Yanrong; Tan, Weihong; Benner, Steven A

    2008-11-01

    A molecular beacon that incorporates components of an artificially expanded genetic information system (Aegis) in its stem is shown not to be opened by unwanted stem invasion by adventitious standard DNA; this should improve the "darkness" of the beacon in real-world applications.

  8. Progress and Prospects for Genetic Modification of Nonhuman Primate Models in Biomedical Research

    PubMed Central

    Chan, Anthony W. S.

    2013-01-01

    The growing interest of modeling human diseases using genetically modified (transgenic) nonhuman primates (NHPs) is a direct result of NHPs (rhesus macaque, etc.) close relation to humans. NHPs share similar developmental paths with humans in their anatomy, physiology, genetics, and neural functions; and in their cognition, emotion, and social behavior. The NHP model within biomedical research has played an important role in the development of vaccines, assisted reproductive technologies, and new therapies for many diseases. Biomedical research has not been the primary role of NHPs. They have mainly been used for safety evaluation and pharmacokinetics studies, rather than determining therapeutic efficacy. The development of the first transgenic rhesus macaque (2001) revolutionized the role of NHP models in biomedicine. Development of the transgenic NHP model of Huntington's disease (2008), with distinctive clinical features, further suggested the uniqueness of the model system; and the potential role of the NHP model for human genetic disorders. Modeling human genetic diseases using NHPs will continue to thrive because of the latest advances in molecular, genetic, and embryo technologies. NHPs rising role in biomedical research, specifically pre-clinical studies, is foreseeable. The path toward the development of transgenic NHPs and the prospect of transgenic NHPs in their new role in future biomedicine needs to be reviewed. This article will focus on the advancement of transgenic NHPs in the past decade, including transgenic technologies and disease modeling. It will outline new technologies that may have significant impact in future NHP modeling and will conclude with a discussion of the future prospects of the transgenic NHP model. PMID:24174443

  9. Complex molecular genetic abnormalities involving three or more genetic mutations are important prognostic factors for acute myeloid leukemia.

    PubMed

    Wakita, S; Yamaguchi, H; Ueki, T; Usuki, K; Kurosawa, S; Kobayashi, Y; Kawata, E; Tajika, K; Gomi, S; Koizumi, M; Fujiwara, Y; Yui, S; Fukunaga, K; Ryotokuji, T; Hirakawa, T; Arai, K; Kitano, T; Kosaka, F; Tamai, H; Nakayama, K; Fukuda, T; Inokuchi, K

    2016-03-01

    We conducted a comprehensive analysis of 28 recurrently mutated genes in acute myeloid leukemia (AML) in 271 patients with de novo AML. Co-mutations were frequently detected in the intermediate cytogenetic risk group, at an average of 2.76 co-mutations per patient. When assessing the prognostic impact of these co-mutations in the intermediate cytogenetic risk group, overall survival (OS) was found to be significantly shorter (P=0.0006) and cumulative incidence of relapse (CIR) significantly higher (P=0.0052) in patients with complex molecular genetic abnormalities (CMGAs) involving three or more mutations. This trend was marked even among patients aged ⩽65 years who were also FLT3-ITD (FMS-like tyrosine kinase 3 internal tandem duplications)-negative (OS: P=0.0010; CIR: P=0.1800). Moreover, the multivariate analysis revealed that CMGA positivity was an independent prognostic factor associated with OS (P=0.0007). In stratification based on FLT3-ITD and CEBPA status and 'simplified analysis of co-mutations' using seven genes that featured frequently in CMGAs, CMGA positivity retained its prognostic value in transplantation-aged patients of the intermediate cytogenetic risk group (OS: P=0.0002. CIR: P<0.0001). In conclusion, CMGAs in AML were found to be strong independent adverse prognostic factors and simplified co-mutation analysis to have clinical usefulness and applicability.

  10. Advances in genetic modification of farm animals using zinc-finger nucleases (ZFN).

    PubMed

    Petersen, Bjoern; Niemann, Heiner

    2015-02-01

    Genome editing tools (GET), including zinc-finger nucleases (ZFN), transcription activator-like endonucleases (TALENS), and meganucleases possess long recognition sites and are thus capable of cutting DNA in a very specific manner. These genome editing tools mediate targeted genetic alterations by enhancing DNA mutation frequency via induction of double-strand breaks at a predetermined genomic site. Compared to conventional homologous recombination based gene targeting, GETs can increase gene targeting and gene disruption via mutagenic DNA repair more than 10,000-fold. Recently, a novel class of genome editing tools was described that uses RNAs to target a specific genomic site. The CRISPR/Cas9 system is capable of targeting even multiple genomic sites in one shot and thus could be superior to ZFNs or TALEN. Current results indicate that these tools can be successfully employed in a broad range of organisms which renders them useful for improving the understanding of complex physiological systems, producing transgenic animals, including creating large animal models for human diseases, creating specific cell lines, and plants, and even for treating human genetic diseases. This review provides an update on the use of ZFNs to modify the genome of farm animals, summarizes current knowledge on the underlying mechanism, and discusses new opportunities for generating genetically modified farm animals.

  11. Behavioral and environmental modification of the genetic influence on body mass index: A twin study

    PubMed Central

    Horn, Erin E.; Turkheimer, Eric; Strachan, Eric; Duncan, Glen E.

    2015-01-01

    Body mass index (BMI) has a strong genetic basis, with a heritability around 0.75, but is also influenced by numerous behavioral and environmental factors. Aspects of the built environment (e.g., environmental walkability) are hypothesized to influence obesity by directly affecting BMI, by facilitating or inhibiting behaviors such as physical activity that are related to BMI, or by suppressing genetic tendencies toward higher BMI. The present study investigated relative influences of physical activity and walkability on variance in BMI using 5,079 same-sex adult twin pairs (70% monozygotic, 65% female). High activity and walkability levels independently suppressed genetic variance in BMI. Estimating their effects simultaneously, however, suggested that the walkability effect was mediated by activity. The suppressive effect of activity on variance in BMI was present even with a tendency for low-BMI individuals to select into environments that require higher activity levels. Overall, our results point to community- or macro-level interventions that facilitate individual-level behaviors as a plausible approach to addressing the obesity epidemic among U.S. adults. PMID:25894925

  12. A survey of the use of soy in processed Turkish meat products and detection of genetic modification.

    PubMed

    Ulca, Pelin; Balta, Handan; Senyuva, Hamide Z

    2014-01-01

    To screen for possible illegal use of soybeans in meat products, the performance characteristics of a commercial polymer chain reaction (PCR) kit for detection of soybean DNA in raw and cooked meat products were established. Minced chicken and beef products containing soybean at levels from 0.1% to 10.0% were analysed by real-time PCR to amplify the soybean lectin gene. The PCR method could reliably detect the addition of soybean at a level of 0.1%. A survey of 38 Turkish processed meat products found only six samples to be negative for the presence of soybean. In 32 (84%) positive samples, 13 (34%) contained levels of soy above 0.1%. Of soybean positive samples, further DNA analysis was conducted by real-time PCR to detect whether genetically modified (GM) soybean had been used. Of 32 meat samples containing soybean, two samples were positive for GM modification.

  13. Genetic Modification of Short Rotation Poplar Biomass Feedstock for Efficient Conversion to Ethanol

    SciTech Connect

    Dinus, R.J.

    2000-08-30

    The Bioenergy Feedstock Development Program, Environmental Sciences Division, Oak Ridge National Laboratory is developing poplars (Populus species and hybrids) as sources of renewable energy, i.e., ethanol. Notable increases in adaptability, volume productivity, and pest/stress resistance have been achieved via classical selection and breeding and intensified cultural practices. Significant advances have also been made in the efficiencies of harvesting and handling systems. Given these and anticipated accomplishments, program leaders are considering shifting some attention to genetically modifying feedstock physical and chemical properties, so as to improve the efficiency with which feedstocks can be converted to ethanol. This report provides an in-depth review and synthesis of opportunities for and feasibilities of genetically modifying feedstock qualities via classical selection and breeding, marker-aided selection and breeding, and genetic transformation. Information was collected by analysis of the literature, with emphasis on that published since 1995, and interviews with prominent scientists, breeders, and growers. Poplar research is well advanced, and literature is abundant. The report therefore primarily reflects advances in poplars, but data from other species, particularly other shortrotation hardwoods, are incorporated to fill gaps. An executive summary and recommendations for research, development, and technology transfer are provided immediately after the table of contents. The first major section of the report describes processes most likely to be used for conversion of poplar biomass to ethanol, the various physical and chemical properties of poplar feedstocks, and how such properties are expected to affect process efficiency. The need is stressed for improved understanding of the impact of change on both overall process and individual process step efficiencies. The second part documents advances in trait measurement instrumentation and methodology

  14. Modification of neurobehavioral effects of mercury by genetic polymorphisms of metallothionein in children.

    PubMed

    Woods, James S; Heyer, Nicholas J; Russo, Joan E; Martin, Michael D; Pillai, Pradeep B; Farin, Federico M

    2013-01-01

    Mercury (Hg) is neurotoxic, and children may be particularly susceptible to this effect. A current major challenge is the identification of children who may be uniquely susceptible to Hg toxicity because of genetic disposition. We examined the hypothesis that genetic variants of metallothionein (MT) that are reported to affect Hg toxicokinetics in adults would modify the neurotoxic effects of Hg in children. Five hundred seven children, 8-12 years of age at baseline, participated in a clinical trial to evaluate the neurobehavioral effects of Hg from dental amalgam tooth fillings. Subjects were evaluated at baseline and at 7 subsequent annual intervals for neurobehavioral performance and urinary Hg levels. Following the completion of the clinical trial, we performed genotyping assays for variants of MT isoforms MT1M (rs2270837) and MT2A (rs10636) on biological samples provided by 330 of the trial participants. Regression modeling strategies were employed to evaluate associations between allelic status, Hg exposure, and neurobehavioral test outcomes. Among girls, few significant interactions or independent main effects for Hg exposure and either of the MT gene variants were observed. In contrast, among boys, numerous significant interaction effects between variants of MT1M and MT2A, alone and combined, with Hg exposure were observed spanning multiple domains of neurobehavioral function. All dose-response associations between Hg exposure and test performance were restricted to boys and were in the direction of impaired performance. These findings suggest increased susceptibility to the adverse neurobehavioral effects of Hg among children with relatively common genetic variants of MT, and may have important public health implications for future strategies aimed at protecting children and adolescents from the potential health risks associated with Hg exposure. We note that because urinary Hg reflects a composite exposure index that cannot be attributed to a specific

  15. Association genetics and transcriptome analysis reveal a gibberellin-responsive pathway involved in regulating photosynthesis.

    PubMed

    Xie, Jianbo; Tian, Jiaxing; Du, Qingzhang; Chen, Jinhui; Li, Ying; Yang, Xiaohui; Li, Bailian; Zhang, Deqiang

    2016-05-01

    Gibberellins (GAs) regulate a wide range of important processes in plant growth and development, including photosynthesis. However, the mechanism by which GAs regulate photosynthesis remains to be understood. Here, we used multi-gene association to investigate the effect of genes in the GA-responsive pathway, as constructed by RNA sequencing, on photosynthesis, growth, and wood property traits, in a population of 435 Populus tomentosa By analyzing changes in the transcriptome following GA treatment, we identified many key photosynthetic genes, in agreement with the observed increase in measurements of photosynthesis. Regulatory motif enrichment analysis revealed that 37 differentially expressed genes related to photosynthesis shared two essential GA-related cis-regulatory elements, the GA response element and the pyrimidine box. Thus, we constructed a GA-responsive pathway consisting of 47 genes involved in regulating photosynthesis, including GID1, RGA, GID2, MYBGa, and 37 photosynthetic differentially expressed genes. Single nucleotide polymorphism (SNP)-based association analysis showed that 142 SNPs, representing 40 candidate genes in this pathway, were significantly associated with photosynthesis, growth, and wood property traits. Epistasis analysis uncovered interactions between 310 SNP-SNP pairs from 37 genes in this pathway, revealing possible genetic interactions. Moreover, a structural gene-gene matrix based on a time-course of transcript abundances provided a better understanding of the multi-gene pathway affecting photosynthesis. The results imply a functional role for these genes in mediating photosynthesis, growth, and wood properties, demonstrating the potential of combining transcriptome-based regulatory pathway construction and genetic association approaches to detect the complex genetic networks underlying quantitative traits.

  16. Association genetics and transcriptome analysis reveal a gibberellin-responsive pathway involved in regulating photosynthesis.

    PubMed

    Xie, Jianbo; Tian, Jiaxing; Du, Qingzhang; Chen, Jinhui; Li, Ying; Yang, Xiaohui; Li, Bailian; Zhang, Deqiang

    2016-05-01

    Gibberellins (GAs) regulate a wide range of important processes in plant growth and development, including photosynthesis. However, the mechanism by which GAs regulate photosynthesis remains to be understood. Here, we used multi-gene association to investigate the effect of genes in the GA-responsive pathway, as constructed by RNA sequencing, on photosynthesis, growth, and wood property traits, in a population of 435 Populus tomentosa By analyzing changes in the transcriptome following GA treatment, we identified many key photosynthetic genes, in agreement with the observed increase in measurements of photosynthesis. Regulatory motif enrichment analysis revealed that 37 differentially expressed genes related to photosynthesis shared two essential GA-related cis-regulatory elements, the GA response element and the pyrimidine box. Thus, we constructed a GA-responsive pathway consisting of 47 genes involved in regulating photosynthesis, including GID1, RGA, GID2, MYBGa, and 37 photosynthetic differentially expressed genes. Single nucleotide polymorphism (SNP)-based association analysis showed that 142 SNPs, representing 40 candidate genes in this pathway, were significantly associated with photosynthesis, growth, and wood property traits. Epistasis analysis uncovered interactions between 310 SNP-SNP pairs from 37 genes in this pathway, revealing possible genetic interactions. Moreover, a structural gene-gene matrix based on a time-course of transcript abundances provided a better understanding of the multi-gene pathway affecting photosynthesis. The results imply a functional role for these genes in mediating photosynthesis, growth, and wood properties, demonstrating the potential of combining transcriptome-based regulatory pathway construction and genetic association approaches to detect the complex genetic networks underlying quantitative traits. PMID:27091876

  17. Specific genetic modifications of domestic animals by gene targeting and animal cloning.

    PubMed

    Wang, Bin; Zhou, Jiangfeng

    2003-11-13

    The technology of gene targeting through homologous recombination has been extremely useful for elucidating gene functions in mice. The application of this technology was thought impossible in the large livestock species until the successful creation of the first mammalian clone "Dolly" the sheep. The combination of the technologies for gene targeting of somatic cells with those of animal cloning made it possible to introduce specific genetic mutations into domestic animals. In this review, the principles of gene targeting in somatic cells and the challenges of nuclear transfer using gene-targeted cells are discussed. The relevance of gene targeting in domestic animals for applications in bio-medicine and agriculture are also examined.

  18. Genetic Modulation of Lipid Profiles following Lifestyle Modification or Metformin Treatment: The Diabetes Prevention Program

    PubMed Central

    Jablonski, Kathleen A.; de Bakker, Paul I. W.; Taylor, Andrew; McAteer, Jarred; Pan, Qing; Horton, Edward S.; Delahanty, Linda M.; Altshuler, David; Shuldiner, Alan R.; Goldberg, Ronald B.; Florez, Jose C.; Bray, George A.; Culbert, Iris W.; Champagne, Catherine M.; Eberhardt, Barbara; Greenway, Frank; Guillory, Fonda G.; Herbert, April A.; Jeffirs, Michael L.; Kennedy, Betty M.; Lovejoy, Jennifer C.; Morris, Laura H.; Melancon, Lee E.; Ryan, Donna; Sanford, Deborah A.; Smith, Kenneth G.; Smith, Lisa L.; Amant, Julia A. St.; Tulley, Richard T.; Vicknair, Paula C.; Williamson, Donald; Zachwieja, Jeffery J.; Polonsky, Kenneth S.; Tobian, Janet; Ehrmann, David; Matulik, Margaret J.; Clark, Bart; Czech, Kirsten; DeSandre, Catherine; Hilbrich, Ruthanne; McNabb, Wylie; Semenske, Ann R.; Caro, Jose F.; Watson, Pamela G.; Goldstein, Barry J.; Smith, Kellie A.; Mendoza, Jewel; Liberoni, Renee; Pepe, Constance; Spandorfer, John; Donahue, Richard P.; Goldberg, Ronald B.; Prineas, Ronald; Rowe, Patricia; Calles, Jeanette; Cassanova-Romero, Paul; Florez, Hermes J.; Giannella, Anna; Kirby, Lascelles; Larreal, Carmen; McLymont, Valerie; Mendez, Jadell; Ojito, Juliet; Perry, Arlette; Saab, Patrice; Haffner, Steven M.; Montez, Maria G.; Lorenzo, Carlos; Martinez, Arlene; Hamman, Richard F.; Nash, Patricia V.; Testaverde, Lisa; Anderson, Denise R.; Ballonoff, Larry B.; Bouffard, Alexis; Calonge, B. Ned; Delve, Lynne; Farago, Martha; Hill, James O.; Hoyer, Shelley R.; Jortberg, Bonnie T.; Lenz, Dione; Miller, Marsha; Price, David W.; Regensteiner, Judith G.; Seagle, Helen; Smith, Carissa M.; Steinke, Sheila C.; VanDorsten, Brent; Horton, Edward S.; Lawton, Kathleen E.; Arky, Ronald A.; Bryant, Marybeth; Burke, Jacqueline P.; Caballero, Enrique; Callaphan, Karen M.; Ganda, Om P.; Franklin, Therese; Jackson, Sharon D.; Jacobsen, Alan M.; Jacobsen, Alan M.; Kula, Lyn M.; Kocal, Margaret; Malloy, Maureen A.; Nicosia, Maryanne; Oldmixon, Cathryn F.; Pan, Jocelyn; Quitingon, Marizel; Rubtchinsky, Stacy; Seely, Ellen W.; Schweizer, Dana; Simonson, Donald; Smith, Fannie; Solomon, Caren G.; Warram, James; Kahn, Steven E.; Montgomery, Brenda K.; Fujimoto, Wilfred; Knopp, Robert H.; Lipkin, Edward W.; Marr, Michelle; Trence, Dace; Kitabchi, Abbas E.; Murphy, Mary E.; Applegate, William B.; Bryer-Ash, Michael; Frieson, Sandra L.; Imseis, Raed; Lambeth, Helen; Lichtermann, Lynne C.; Oktaei, Hooman; Rutledge, Lily M.K.; Sherman, Amy R.; Smith, Clara M.; Soberman, Judith E.; Williams-Cleaves, Beverly; Metzger, Boyd E.; Johnson, Mariana K.; Behrends, Catherine; Cook, Michelle; Fitzgibbon, Marian; Giles, Mimi M.; Heard, Deloris; Johnson, Cheryl K.H.; Larsen, Diane; Lowe, Anne; Lyman, Megan; McPherson, David; Molitch, Mark E.; Pitts, Thomas; Reinhart, Renee; Roston, Susan; Schinleber, Pamela A.; Nathan, David M.; McKitrick, Charles; Turgeon, Heather; Abbott, Kathy; Anderson, Ellen; Bissett, Laurie; Cagliero, Enrico; Florez, Jose C.; Delahanty, Linda; Goldman, Valerie; Poulos, Alexandra; Olefsky, Jerrold M.; Carrion-Petersen, Mary Lou; Barrett-Connor, Elizabeth; Edelman, Steven V.; Henry, Robert R.; Horne, Javiva; Janesch, Simona Szerdi; Leos, Diana; Mudaliar, Sundar; Polonsky, William; Smith, Jean; Vejvoda, Karen; Pi-Sunyer, F. Xavier; Lee, Jane E.; Allison, David B.; Aronoff, Nancy J.; Crandall, Jill P.; Foo, Sandra T.; Pal, Carmen; Parkes, Kathy; Pena, Mary Beth; Rooney, Ellen S.; Wye, Gretchen E.H. Van; Viscovich, Kristine A.; Marrero, David G.; Prince, Melvin J.; Kelly, Susie M.; Dotson, Yolanda F.; Fineberg, Edwin S.; Guare, John C; Hadden, Angela M.; Ignaut, James M.; Jackson, Marcia L.; Kirkman, Marion S.; Mather, Kieren J.; Porter, Beverly D.; Roach, Paris J.; Rowland, Nancy D.; Wheeler, Madelyn L.; Ratner, Robert E.; Youssef, Gretchen; Shapiro, Sue; Bavido-Arrage, Catherine; Boggs, Geraldine; Bronsord, Marjorie; Brown, Ernestine; Cheatham, Wayman W.; Cola, Susan; Evans, Cindy; Gibbs, Peggy; Kellum, Tracy; Levatan, Claresa; Nair, Asha K.; Passaro, Maureen; Uwaifo, Gabriel; Saad, Mohammed F.; Budget, Maria; Jinagouda, Sujata; Akbar, Khan; Conzues, Claudia; Magpuri, Perpetua; Ngo, Kathy; Rassam, Amer; Waters, Debra; Xapthalamous, Kathy; Santiago, Julio V.; Dagogo-Jack, Samuel; White, Neil H.; Das, Samia; Santiago, Ana; Brown, Angela; Fisher, Edwin; Hurt, Emma; Jones, Tracy; Kerr, Michelle; Ryder, Lucy; Wernimont, Cormarie; Saudek, Christopher D.; Bradley, Vanessa; Sullivan, Emily; Whittington, Tracy; Abbas, Caroline; Brancati, Frederick L.; Clark, Jeanne M.; Charleston, Jeanne B.; Freel, Janice; Horak, Katherine; Jiggetts, Dawn; Johnson, Deloris; Joseph, Hope; Loman, Kimberly; Mosley, Henry; Rubin, Richard R.; Samuels, Alafia; Stewart, Kerry J.; Williamson, Paula; Schade, David S.; Adams, Karwyn S.; Johannes, Carolyn; Atler, Leslie F.; Boyle, Patrick J.; Burge, Mark R.; Canady, Janene L.; Chai, Lisa; Gonzales, Ysela; Hernandez-McGinnis, Doris A.; Katz, Patricia; King, Carolyn; Rassam, Amer; Rubinchik, Sofya; Senter, Willette; Waters, Debra; Shamoon, Harry; Brown, Janet O.; Adorno, Elsie; Cox, Liane; Crandall, Jill; Duffy, Helena; Engel, Samuel; Friedler, Allison; Howard-Century, Crystal J.; Kloiber, Stacey; Longchamp, Nadege; Martinez, Helen; Pompi, Dorothy; Scheindlin, Jonathan; Violino, Elissa; Walker, Elizabeth; Wylie-Rosett, Judith; Zimmerman, Elise; Zonszein, Joel; Orchard, Trevor; Wing, Rena R.; Koenning, Gaye; Kramer, M. Kaye; Barr, Susan; Boraz, Miriam; Clifford, Lisa; Culyba, Rebecca; Frazier, Marlene; Gilligan, Ryan; Harrier, Susan; Harris, Louann; Jeffries, Susan; Kriska, Andrea; Manjoo, Qurashia; Mullen, Monica; Noel, Alicia; Otto, Amy; Semler, Linda; Smith, Cheryl F.; Smith, Marie; Venditti, Elizabeth; Weinzierl, Valarie; Williams, Katherine V.; Wilson, Tara; Arakaki, Richard F.; Latimer, Renee W.; Baker-Ladao, Narleen K.; Beddow, Ralph; Dias, Lorna; Inouye, Jillian; Mau, Marjorie K.; Mikami, Kathy; Mohideen, Pharis; Odom, Sharon K.; Perry, Raynette U.; Knowler, William C.; Cooeyate, Norman; Hoskin, Mary A.; Percy, Carol A.; Acton, Kelly J.; Andre, Vickie L.; Barber, Rosalyn; Begay, Shandiin; Bennett, Peter H.; Benson, Mary Beth; Bird, Evelyn C.; Broussard, Brenda A.; Chavez, Marcella; Dacawyma, Tara; Doughty, Matthew S.; Duncan, Roberta; Edgerton, Cyndy; Ghahate, Jacqueline M.; Glass, Justin; Glass, Martia; Gohdes, Dorothy; Grant, Wendy; Hanson, Robert L.; Horse, Ellie; Ingraham, Louise E.; Jackson, Merry; Jay, Priscilla; Kaskalla, Roylen S.; Kessler, David; Kobus, Kathleen M.; Krakoff, Jonathan; Manus, Catherine; Michaels, Sara; Morgan, Tina; Nashboo, Yolanda; Nelson, Julie A.; Poirier, Steven; Polczynski, Evette; Reidy, Mike; Roumain, Jeanine; Rowse, Debra; Sangster, Sandra; Sewenemewa, Janet; Tonemah, Darryl; Wilson, Charlton; Yazzie, Michelle; Bain, Raymond; Fowler, Sarah; Brenneman, Tina; Abebe, Solome; Bamdad, Julie; Callaghan, Jackie; Edelstein, Sharon L.; Gao, Yuping; Grimes, Kristina L.; Grover, Nisha; Haffner, Lori; Jones, Steve; Jones, Tara L.; Katz, Richard; Lachin, John M.; Mucik, Pamela; Orlosky, Robert; Rochon, James; Sapozhnikova, Alla; Sherif, Hanna; Stimpson, Charlotte; Temprosa, Marinella; Walker-Murray, Fredricka; Marcovina, Santica; Strylewicz, Greg; Aldrich, F. Alan; O'Leary, Dan; Stamm, Elizabeth; Rautaharju, Pentti; Prineas, Ronald J.; Alexander, Teresa; Campbell, Charles; Hall, Sharon; Li, Yabing; Mills, Margaret; Pemberton, Nancy; Rautaharju, Farida; Zhang, Zhuming; Mayer-Davis, Elizabeth; Moran, Robert R.; Ganiats, Ted; David, Kristin; Sarkin, Andrew J.; Eastman, R.; Fradkin, Judith; Garfield, Sanford; Gregg, Edward; Zhang, Ping; Herman, William; Florez, Jose C.; Altshuler, David; de Bakker, Paul I.W.; Franks, Paul W.; Hanson, Robert L.; Jablonski, Kathleen; Knowler, William C.; McAteer, Jarred B.; Pollin, Toni I.; Shuldiner, Alan R.

    2012-01-01

    Weight-loss interventions generally improve lipid profiles and reduce cardiovascular disease risk, but effects are variable and may depend on genetic factors. We performed a genetic association analysis of data from 2,993 participants in the Diabetes Prevention Program to test the hypotheses that a genetic risk score (GRS) based on deleterious alleles at 32 lipid-associated single-nucleotide polymorphisms modifies the effects of lifestyle and/or metformin interventions on lipid levels and nuclear magnetic resonance (NMR) lipoprotein subfraction size and number. Twenty-three loci previously associated with fasting LDL-C, HDL-C, or triglycerides replicated (P = 0.04–1×10−17). Except for total HDL particles (r = −0.03, P = 0.26), all components of the lipid profile correlated with the GRS (partial |r| = 0.07–0.17, P = 5×10−5–1×10−19). The GRS was associated with higher baseline-adjusted 1-year LDL cholesterol levels (β = +0.87, SEE±0.22 mg/dl/allele, P = 8×10−5, Pinteraction = 0.02) in the lifestyle intervention group, but not in the placebo (β = +0.20, SEE±0.22 mg/dl/allele, P = 0.35) or metformin (β = −0.03, SEE±0.22 mg/dl/allele, P = 0.90; Pinteraction = 0.64) groups. Similarly, a higher GRS predicted a greater number of baseline-adjusted small LDL particles at 1 year in the lifestyle intervention arm (β = +0.30, SEE±0.012 ln nmol/L/allele, P = 0.01, Pinteraction = 0.01) but not in the placebo (β = −0.002, SEE±0.008 ln nmol/L/allele, P = 0.74) or metformin (β = +0.013, SEE±0.008 nmol/L/allele, P = 0.12; Pinteraction = 0.24) groups. Our findings suggest that a high genetic burden confers an adverse lipid profile and predicts attenuated response in LDL-C levels and small LDL particle number to dietary and physical activity interventions aimed at weight loss. PMID:22951888

  19. Genetic modulation of lipid profiles following lifestyle modification or metformin treatment: the Diabetes Prevention Program.

    PubMed

    Pollin, Toni I; Isakova, Tamara; Jablonski, Kathleen A; de Bakker, Paul I W; Taylor, Andrew; McAteer, Jarred; Pan, Qing; Horton, Edward S; Delahanty, Linda M; Altshuler, David; Shuldiner, Alan R; Goldberg, Ronald B; Florez, Jose C; Franks, Paul W

    2012-01-01

    Weight-loss interventions generally improve lipid profiles and reduce cardiovascular disease risk, but effects are variable and may depend on genetic factors. We performed a genetic association analysis of data from 2,993 participants in the Diabetes Prevention Program to test the hypotheses that a genetic risk score (GRS) based on deleterious alleles at 32 lipid-associated single-nucleotide polymorphisms modifies the effects of lifestyle and/or metformin interventions on lipid levels and nuclear magnetic resonance (NMR) lipoprotein subfraction size and number. Twenty-three loci previously associated with fasting LDL-C, HDL-C, or triglycerides replicated (P = 0.04-1 × 10(-17)). Except for total HDL particles (r = -0.03, P = 0.26), all components of the lipid profile correlated with the GRS (partial |r| = 0.07-0.17, P = 5 × 10(-5)-1 10(-19)). The GRS was associated with higher baseline-adjusted 1-year LDL cholesterol levels (β = +0.87, SEE ± 0.22 mg/dl/allele, P = 8 × 10(-5), P(interaction) = 0.02) in the lifestyle intervention group, but not in the placebo (β = +0.20, SEE ± 0.22 mg/dl/allele, P = 0.35) or metformin (β = -0.03, SEE ± 0.22 mg/dl/allele, P = 0.90; P(interaction) = 0.64) groups. Similarly, a higher GRS predicted a greater number of baseline-adjusted small LDL particles at 1 year in the lifestyle intervention arm (β = +0.30, SEE ± 0.012 ln nmol/L/allele, P = 0.01, P(interaction) = 0.01) but not in the placebo (β = -0.002, SEE ± 0.008 ln nmol/L/allele, P = 0.74) or metformin (β = +0.013, SEE ± 0.008 nmol/L/allele, P = 0.12; P(interaction) = 0.24) groups. Our findings suggest that a high genetic burden confers an adverse lipid profile and predicts attenuated response in LDL-C levels and small LDL particle number to dietary and physical activity interventions aimed at weight loss.

  20. Idiopathic Pulmonary Fibrosis: A Genetic Disease That Involves Mucociliary Dysfunction of the Peripheral Airways.

    PubMed

    Evans, Christopher M; Fingerlin, Tasha E; Schwarz, Marvin I; Lynch, David; Kurche, Jonathan; Warg, Laura; Yang, Ivana V; Schwartz, David A

    2016-10-01

    Idiopathic pulmonary fibrosis (IPF) is an incurable complex genetic disorder that is associated with sequence changes in 7 genes (MUC5B, TERT, TERC, RTEL1, PARN, SFTPC, and SFTPA2) and with variants in at least 11 novel loci. We have previously found that 1) a common gain-of-function promoter variant in MUC5B rs35705950 is the strongest risk factor (genetic and otherwise), accounting for 30-35% of the risk of developing IPF, a disease that was previously considered idiopathic; 2) the MUC5B promoter variant can potentially be used to identify individuals with preclinical pulmonary fibrosis and is predictive of radiologic progression of preclinical pulmonary fibrosis; and 3) MUC5B may be involved in the pathogenesis of pulmonary fibrosis with MUC5B message and protein expressed in bronchiolo-alveolar epithelia of IPF and the characteristic IPF honeycomb cysts. Based on these considerations, we hypothesize that excessive production of MUC5B either enhances injury due to reduced mucociliary clearance or impedes repair consequent to disruption of normal regenerative mechanisms in the distal lung. In aggregate, these novel considerations should have broad impact, resulting in specific etiologic targets, early detection of disease, and novel biologic pathways for use in the design of future intervention, prevention, and mechanistic studies of IPF. PMID:27630174

  1. Systematic Analysis of the Genetic Variability That Impacts SUMO Conjugation and Their Involvement in Human Diseases

    NASA Astrophysics Data System (ADS)

    Xu, Hao-Dong; Shi, Shao-Ping; Chen, Xiang; Qiu, Jian-Ding

    2015-07-01

    Protein function has been observed to rely on select essential sites instead of requiring all sites to be indispensable. Small ubiquitin-related modifier (SUMO) conjugation or sumoylation, which is a highly dynamic reversible process and its outcomes are extremely diverse, ranging from changes in localization to altered activity and, in some cases, stability of the modified, has shown to be especially valuable in cellular biology. Motivated by the significance of SUMO conjugation in biological processes, we report here on the first exploratory assessment whether sumoylation related genetic variability impacts protein functions as well as the occurrence of diseases related to SUMO. Here, we defined the SUMOAMVR as sumoylation related amino acid variations that affect sumoylation sites or enzymes involved in the process of connectivity, and categorized four types of potential SUMOAMVRs. We detected that 17.13% of amino acid variations are potential SUMOAMVRs and 4.83% of disease mutations could lead to SUMOAMVR with our system. More interestingly, the statistical analysis demonstrates that the amino acid variations that directly create new potential lysine sumoylation sites are more likely to cause diseases. It can be anticipated that our method can provide more instructive guidance to identify the mechanisms of genetic diseases.

  2. Genetic modification of T cells improves the effectiveness of adoptive tumor immunotherapy.

    PubMed

    Jakóbisiak, Marek; Gołab, Jakub

    2010-10-01

    Appropriate combinations of immunotherapy and gene therapy promise to be more effective in the treatment of cancer patients than either of these therapeutic approaches alone. One such treatment is based on the application of patients' cytotoxic T cells, which can be activated, expanded, and genetically engineered to recognize particular tumor-associated antigens (TAAs). Because T cells recognizing TAAs might become unresponsive in the process of tumor development as a result of tumor evasion strategies, immunogenic viral antigens or alloantigens could be used for the expansion of cytotoxic T cells and then redirected through genetic engineering. This therapeutic approach has already demonstrated promising results in melanoma patients and could be used in the treatment of many other tumors. The graft-versus-leukemia, or more generally graft-versus-tumor, reaction based on the application of a donor lymphocyte infusion can also be ameliorated through the incorporation of suicide genes into donor lymphocytes. Such lymphocytes could be safely and more extensively used in tumor patients because they could be eliminated should a severe graft-versus-host reaction develop.

  3. A BioBrick compatible strategy for genetic modification of plants

    PubMed Central

    2012-01-01

    Background Plant biotechnology can be leveraged to produce food, fuel, medicine, and materials. Standardized methods advocated by the synthetic biology community can accelerate the plant design cycle, ultimately making plant engineering more widely accessible to bioengineers who can contribute diverse creative input to the design process. Results This paper presents work done largely by undergraduate students participating in the 2010 International Genetically Engineered Machines (iGEM) competition. Described here is a framework for engineering the model plant Arabidopsis thaliana with standardized, BioBrick compatible vectors and parts available through the Registry of Standard Biological Parts (http://www.partsregistry.org). This system was used to engineer a proof-of-concept plant that exogenously expresses the taste-inverting protein miraculin. Conclusions Our work is intended to encourage future iGEM teams and other synthetic biologists to use plants as a genetic chassis. Our workflow simplifies the use of standardized parts in plant systems, allowing the construction and expression of heterologous genes in plants within the timeframe allotted for typical iGEM projects. PMID:22716313

  4. Meta-analysis of the independent and cumulative effects of multiple genetic modifications on pig lung xenograft performance during ex vivo perfusion with human blood

    PubMed Central

    Harris, Donald G.; Quinn, Kevin J.; French, Beth M.; Schwartz, Evan; Kang, Elizabeth; Dahi, Siamak; Phelps, Carol J.; Ayares, David L.; Burdorf, Lars; Azimzadeh, Agnes M.; Pierson, Richard N.

    2014-01-01

    Background Genetically modified pigs are a promising potential source of lung xenografts. Ex-vivo xenoperfusion is an effective platform for testing the effect of new modifications, but typical experiments are limited by testing of a single genetic intervention and small sample sizes. The purpose of this study was to analyze the individual and aggregate effects of donor genetic modifications on porcine lung xenograft survival and injury in an extensive pig lung xenoperfusion series. Methods Data from 157 porcine lung xenoperfusion experiments using otherwise unmodified heparinized human blood were aggregated as either continuous or dichotomous variables. Lungs were wild type in 17 perfusions (11% of the study group), while 31 lungs (20% of the study group) had 1 genetic modification, 40 lungs (39%) had 2, and 47 lungs (30%) had 3 or more modifications. The primary endpoint was functional lung survival to 4 hours of perfusion. Secondary analyses evaluated previously identified markers associated with known lung xenograft injury mechanisms. In addition to comparison among all xenografts grouped by survival status, a subgroup analysis was performed of lungs incorporating the GalTKO.hCD46 genotype. Results Each increase in the number of genetic modifications was associated with additional prolongation of lung xenograft survival. Lungs that exhibited survival to 4 hours generally had reduced platelet activation and thrombin generation. GalTKO and the expression of hCD46, HO-1, hCD55 or hEPCR were associated with improved survival. hTBM, HLA-E, and hCD39 were associated with no significant effect on the primary outcome. Conclusion This meta-analysis of an extensive lung xenotransplantation series demonstrates that increasing the number of genetic modifications targeting known xenogeneic lung injury mechanisms is associated with incremental improvements in lung survival. While more detailed mechanistic studies are needed to explore the relationship between gene expression

  5. Enhanced genetic modification of adult growth factor mobilized peripheral blood hematopoietic stem and progenitor cells with rapamycin.

    PubMed

    Li, Lijing; Torres-Coronado, Mónica; Gu, Angel; Rao, Anitha; Gardner, Agnes M; Epps, Elizabeth W; Gonzalez, Nancy; Tran, Chy-Anh; Wu, Xiwei; Wang, Jin-Hui; DiGiusto, David L

    2014-10-01

    Genetic modification of adult human hematopoietic stem and progenitor cells (HSPCs) with lentiviral vectors leads to long-term gene expression in the progeny of the HSPCs and has been used to successfully treat several monogenic diseases. In some cases, the gene-modified cells have a selective growth advantage over nonmodified cells and eventually are the dominant engrafted population. However, in disease indications for which the gene-modified cells do not have a selective advantage, optimizing transduction of HSPC is paramount to successful stem cell-based gene therapy. We demonstrate here that transduction of adult CD34+ HSPCs with lentiviral vectors in the presence of rapamycin, a widely used mTORC1 inhibitor, results in an approximately threefold increase in stable gene marking with minimal effects on HSPC growth and differentiation. Using this approach, we have demonstrated that we can enhance the frequency of gene-modified HSPCs that give rise to clonogenic progeny in vitro without excessive increases in the number of vector copies per cell or changes in integration pattern. The genetic marking of HSPCs and expression of transgenes is durable, and transplantation of gene-modified HSPCs into immunodeficient mice results in high levels of gene marking of the lymphoid and myeloid progeny in vivo. The prior safe clinical history of rapamycin in other applications supports the use of this compound to generate gene-modified autologous HSPCs for our HIV gene therapy clinical trials.

  6. Genetically Determined Susceptibility to Tuberculosis in Mice Causally Involves Accelerated and Enhanced Recruitment of Granulocytes

    PubMed Central

    Keller, Christine; Hoffmann, Reinhard; Lang, Roland; Brandau, Sven; Hermann, Corinna; Ehlers, Stefan

    2006-01-01

    Classical twin studies and recent linkage analyses of African populations have revealed a potential involvement of host genetic factors in susceptibility or resistance to Mycobacterium tuberculosis infection. In order to identify the candidate genes involved and test their causal implication, we capitalized on the mouse model of tuberculosis, since inbred mouse strains also differ substantially in their susceptibility to infection. Two susceptible and two resistant mouse strains were aerogenically infected with 1,000 CFU of M. tuberculosis, and the regulation of gene expression was examined by Affymetrix GeneChip U74A array with total lung RNA 2 and 4 weeks postinfection. Four weeks after infection, 96 genes, many of which are involved in inflammatory cell recruitment and activation, were regulated in common. One hundred seven genes were differentially regulated in susceptible mouse strains, whereas 43 genes were differentially expressed only in resistant mice. Data mining revealed a bias towards the expression of genes involved in granulocyte pathophysiology in susceptible mice, such as an upregulation of those for the neutrophil chemoattractant LIX (CXCL5), interleukin 17 receptor, phosphoinositide kinase 3 delta, or gamma interferon-inducible protein 10. Following M. tuberculosis challenge in both airways or peritoneum, granulocytes were recruited significantly faster and at higher numbers in susceptible than in resistant mice. When granulocytes were efficiently depleted by either of two regimens at the onset of infection, only susceptible mice survived aerosol challenge with M. tuberculosis significantly longer than control mice. We conclude that initially enhanced recruitment of granulocytes contributes to susceptibility to tuberculosis. PMID:16790804

  7. Large-Scale Culture and Genetic Modification of Human Natural Killer Cells for Cellular Therapy.

    PubMed

    Lapteva, Natalia; Parihar, Robin; Rollins, Lisa A; Gee, Adrian P; Rooney, Cliona M

    2016-01-01

    Recent advances in methods for the ex vivo expansion of human natural killer (NK) cells have facilitated the use of these powerful immune cells in clinical protocols. Further, the ability to genetically modify primary human NK cells following rapid expansion allows targeting and enhancement of their immune function. We have successfully adapted an expansion method for primary NK cells from peripheral blood mononuclear cells or from apheresis products in gas permeable rapid expansion devices (G-Rexes). Here, we describe an optimized protocol for rapid and robust NK cell expansion as well as a method for highly efficient retroviral transduction of these ex vivo expanded cells. These methodologies are good manufacturing practice (GMP) compliant and could be used for clinical-grade product manufacturing. PMID:27177667

  8. Large-Scale Culture and Genetic Modification of Human Natural Killer Cells for Cellular Therapy.

    PubMed

    Lapteva, Natalia; Parihar, Robin; Rollins, Lisa A; Gee, Adrian P; Rooney, Cliona M

    2016-01-01

    Recent advances in methods for the ex vivo expansion of human natural killer (NK) cells have facilitated the use of these powerful immune cells in clinical protocols. Further, the ability to genetically modify primary human NK cells following rapid expansion allows targeting and enhancement of their immune function. We have successfully adapted an expansion method for primary NK cells from peripheral blood mononuclear cells or from apheresis products in gas permeable rapid expansion devices (G-Rexes). Here, we describe an optimized protocol for rapid and robust NK cell expansion as well as a method for highly efficient retroviral transduction of these ex vivo expanded cells. These methodologies are good manufacturing practice (GMP) compliant and could be used for clinical-grade product manufacturing.

  9. Genetic modification of primary chronic lymphocytic leukemia cells with a lentivirus expressing CD38

    PubMed Central

    Pearce, Laurence; Morgan, Liam; Lin, Thet Thet; Hewamana, Saman; Matthews, R. James; Deaglio, Silvia; Rowntree, Clare; Fegan, Christopher; Pepper, Christopher; Brennan, Paul

    2010-01-01

    Studies of the role of individual genes in chronic lymphocytic leukemia (CLL) have been hampered by the inability to consistently transfect primary tumor cells. Here, we describe a highly efficient method of genetically modifying primary CLL cells using a VSVG pseudotyped lentiviral vector. We transduced CD38 negative CLL cells with a lentiviral vector encoding CD38 which caused increased surface CD38 expression in all the samples tested (n=17) with no evidence of plasmacytoid differentiation. The mean percentage of positive cells expressing CD38 was 87%±8.5% and the mean cell viability 74%±17%. This high level of transduction of all the CLL cell samples tested demonstrates the utility of this technique which should prove applicable for the introduction and analysis of other genes in these non-dividing cells. PMID:20207849

  10. Enhancing cellulose utilization for fuels and chemicals by genetic modification of plant cell wall architecture.

    PubMed

    Vermerris, Wilfred; Abril, Alejandra

    2015-04-01

    Cellulose from plant biomass can serve as a sustainable feedstock for fuels, chemicals and polymers that are currently produced from petroleum. In order to enhance economic feasibility, the efficiency of cell wall deconstruction needs to be enhanced. With the use of genetic and biotechnological approaches cell wall composition can be modified in such a way that interactions between the major cell wall polymers—cellulose, hemicellulosic polysaccharides and lignin—are altered. Some of the resulting plants are compromised in their growth and development, but this may be caused in part by the plant's overcompensation for metabolic perturbances. In other cases novel structures have been introduced in the cell wall without negative effects. The first field studies with engineered bioenergy crops look promising, while detailed structural analyses of cellulose synthase offer new opportunities to modify cellulose itself.

  11. Specific genetic modifications of domestic animals by gene targeting and animal cloning.

    PubMed

    Wang, Bin; Zhou, Jiangfeng

    2003-11-13

    The technology of gene targeting through homologous recombination has been extremely useful for elucidating gene functions in mice. The application of this technology was thought impossible in the large livestock species until the successful creation of the first mammalian clone "Dolly" the sheep. The combination of the technologies for gene targeting of somatic cells with those of animal cloning made it possible to introduce specific genetic mutations into domestic animals. In this review, the principles of gene targeting in somatic cells and the challenges of nuclear transfer using gene-targeted cells are discussed. The relevance of gene targeting in domestic animals for applications in bio-medicine and agriculture are also examined. PMID:14614774

  12. Conservation and modification of genetic and physiological toolkits underpinning diapause in bumble bee queens.

    PubMed

    Amsalem, Etya; Galbraith, David A; Cnaani, Jonathan; Teal, Peter E A; Grozinger, Christina M

    2015-11-01

    Diapause is the key adaptation allowing insects to survive unfavourable conditions and inhabit an array of environments. Physiological changes during diapause are largely conserved across species and are hypothesized to be regulated by a conserved suite of genes (a 'toolkit'). Furthermore, it is hypothesized that in social insects, this toolkit was co-opted to mediate caste differentiation between long-lived, reproductive, diapause-capable queens and short-lived, sterile workers. Using Bombus terrestris queens, we examined the physiological and transcriptomic changes associated with diapause and CO2 treatment, which causes queens to bypass diapause. We performed comparative analyses with genes previously identified to be associated with diapause in the Dipteran Sarcophaga crassipalpis and with caste differentiation in bumble bees. As in Diptera, diapause in bumble bees is associated with physiological and transcriptional changes related to nutrient storage, stress resistance and core metabolic pathways. There is a significant overlap, both at the level of transcript and gene ontology, between the genetic mechanisms mediating diapause in B. terrestris and S. crassipalpis, reaffirming the existence of a conserved insect diapause genetic toolkit. However, a substantial proportion (10%) of the differentially regulated transcripts in diapausing queens have no clear orthologs in other species, and key players regulating diapause in Diptera (juvenile hormone and vitellogenin) appear to have distinct functions in bumble bees. We also found a substantial overlap between genes related to caste determination and diapause in bumble bees. Thus, our studies demonstrate an intriguing interplay between pathways underpinning adaptation to environmental extremes and the evolution of sociality in insects.

  13. Non-coding RNAs in crop genetic modification: considerations and predictable environmental risk assessments (ERA).

    PubMed

    Ramesh, S V

    2013-09-01

    Of late non-coding RNAs (ncRNAs)-mediated gene silencing is an influential tool deliberately deployed to negatively regulate the expression of targeted genes. In addition to the widely employed small interfering RNA (siRNA)-mediated gene silencing approach, other variants like artificial miRNA (amiRNA), miRNA mimics, and artificial transacting siRNAs (tasiRNAs) are being explored and successfully deployed in developing non-coding RNA-based genetically modified plants. The ncRNA-based gene manipulations are typified with mobile nature of silencing signals, interference from viral genome-derived suppressor proteins, and an obligation for meticulous computational analysis to prevaricate any inadvertent effects. In a broad sense, risk assessment inquiries for genetically modified plants based on the expression of ncRNAs are competently addressed by the environmental risk assessment (ERA) models, currently in vogue, designed for the first generation transgenic plants which are based on the expression of heterologous proteins. Nevertheless, transgenic plants functioning on the foundation of ncRNAs warrant due attention with respect to their unique attributes like off-target or non-target gene silencing effects, small RNAs (sRNAs) persistence, food and feed safety assessments, problems in detection and tracking of sRNAs in food, impact of ncRNAs in plant protection measures, effect of mutations etc. The role of recent developments in sequencing techniques like next generation sequencing (NGS) and the ERA paradigm of the different countries in vogue are also discussed in the context of ncRNA-based gene manipulations.

  14. Kernel-Based Aggregation of Marker-Level Genetic Association Tests Involving Copy-Number Variation

    PubMed Central

    Li, Yinglei; Breheny, Patrick

    2013-01-01

    Genetic association tests involving copy-number variants (CNVs) are complicated by the fact that CNVs span multiple markers at which measurements are taken. The power of an association test at a single marker is typically low, and it is desirable to pool information across the markers spanned by the CNV. However, CNV boundaries are not known in advance, and the best way to proceed with this pooling is unclear. In this article, we propose a kernel-based method for aggregation of marker-level tests and explore several aspects of its implementation. In addition, we explore some of the theoretical aspects of marker-level test aggregation, proposing a permutation-based approach that preserves the family-wise error rate of the testing procedure, while demonstrating that several simpler alternatives fail to do so. The empirical power of the approach is studied in a number of simulations constructed from real data involving a pharmacogenomic study of gemcitabine and compares favorably with several competing approaches.

  15. Improving the immunogenicity of a trivalent Neisseria meningitidis native outer membrane vesicle vaccine by genetic modification.

    PubMed

    Zhang, Lan; Wen, Zhiyun; Lin, Jing; Xu, Hui; Herbert, Paul; Wang, Xin-Min; Mehl, John T; Ahl, Patrick L; Dieter, Lance; Russell, Ryann; Kosinski, Mike J; Przysiecki, Craig T

    2016-07-29

    Trivalent native outer membrane vesicles (nOMVs) derived from three genetically modified Neisseria meningitidis serogroup B strains have been previously evaluated immunologically in mice and rabbits. This nOMV vaccine elicited serum bactericidal activity (SBA) against multiple N. meningitidis serogroup B strains as well as strains from serogroups C, Y, W, and X. In this study, we used trivalent nOMVs isolated from the same vaccine strains and evaluated their immunogenicity in an infant Rhesus macaque (IRM) model whose immune responses to the vaccine are likely to be more predictive of the responses in human infants. IRMs were immunized with trivalent nOMV vaccines and sera were evaluated for exogenous human serum complement-dependent SBA (hSBA). Antibody responses to selected hSBA generating antigens contained within the trivalent nOMVs were also measured and we found that antibody titers against factor H binding protein variant 2 (fHbpv2) were very low in the sera from animals immunized with these original nOMV vaccines. To increase the fHbp content in the nOMVs, the vaccine strains were further genetically altered by addition of another fHbp gene copy into the porB locus. Trivalent nOMVs from the three new vaccine strains had higher fHbp antigen levels and generated higher anti-fHbp antibody responses in immunized mice and IRMs. As expected, fHbp insertion into the porB locus resulted in no PorB expression. Interestingly, higher expression of PorA, an hSBA generating antigen, was observed for all three modified vaccine strains. Compared to the trivalent nOMVs from the original strains, higher PorA levels in the improved nOMVs resulted in higher anti-PorA antibody responses in mice and IRMs. In addition, hSBA titers against other strains with PorA as the only hSBA antigen in common with the vaccine strains also increased. PMID:27269057

  16. Improving the immunogenicity of a trivalent Neisseria meningitidis native outer membrane vesicle vaccine by genetic modification.

    PubMed

    Zhang, Lan; Wen, Zhiyun; Lin, Jing; Xu, Hui; Herbert, Paul; Wang, Xin-Min; Mehl, John T; Ahl, Patrick L; Dieter, Lance; Russell, Ryann; Kosinski, Mike J; Przysiecki, Craig T

    2016-07-29

    Trivalent native outer membrane vesicles (nOMVs) derived from three genetically modified Neisseria meningitidis serogroup B strains have been previously evaluated immunologically in mice and rabbits. This nOMV vaccine elicited serum bactericidal activity (SBA) against multiple N. meningitidis serogroup B strains as well as strains from serogroups C, Y, W, and X. In this study, we used trivalent nOMVs isolated from the same vaccine strains and evaluated their immunogenicity in an infant Rhesus macaque (IRM) model whose immune responses to the vaccine are likely to be more predictive of the responses in human infants. IRMs were immunized with trivalent nOMV vaccines and sera were evaluated for exogenous human serum complement-dependent SBA (hSBA). Antibody responses to selected hSBA generating antigens contained within the trivalent nOMVs were also measured and we found that antibody titers against factor H binding protein variant 2 (fHbpv2) were very low in the sera from animals immunized with these original nOMV vaccines. To increase the fHbp content in the nOMVs, the vaccine strains were further genetically altered by addition of another fHbp gene copy into the porB locus. Trivalent nOMVs from the three new vaccine strains had higher fHbp antigen levels and generated higher anti-fHbp antibody responses in immunized mice and IRMs. As expected, fHbp insertion into the porB locus resulted in no PorB expression. Interestingly, higher expression of PorA, an hSBA generating antigen, was observed for all three modified vaccine strains. Compared to the trivalent nOMVs from the original strains, higher PorA levels in the improved nOMVs resulted in higher anti-PorA antibody responses in mice and IRMs. In addition, hSBA titers against other strains with PorA as the only hSBA antigen in common with the vaccine strains also increased.

  17. Modification of Occupational Exposures on Bladder Cancer Risk by Common Genetic Polymorphisms.

    PubMed

    Figueroa, Jonine D; Koutros, Stella; Colt, Joanne S; Kogevinas, Manolis; Garcia-Closas, Montserrat; Real, Francisco X; Friesen, Melissa C; Baris, Dalsu; Stewart, Patricia; Schwenn, Molly; Johnson, Alison; Karagas, Margaret R; Armenti, Karla R; Moore, Lee E; Schned, Alan; Lenz, Petra; Prokunina-Olsson, Ludmila; Banday, A Rouf; Paquin, Ashley; Ylaya, Kris; Chung, Joon-Yong; Hewitt, Stephen M; Nickerson, Michael L; Tardón, Adonina; Serra, Consol; Carrato, Alfredo; García-Closas, Reina; Lloreta, Josep; Malats, Núria; Fraumeni, Joseph F; Chanock, Stephen J; Chatterjee, Nilanjan; Rothman, Nathaniel; Silverman, Debra T

    2015-11-01

    Few studies have demonstrated gene/environment interactions in cancer research. Using data on high-risk occupations for 2258 case patients and 2410 control patients from two bladder cancer studies, we observed that three of 16 known or candidate bladder cancer susceptibility variants displayed statistically significant and consistent evidence of additive interactions; specifically, the GSTM1 deletion polymorphism (P interaction ≤ .001), rs11892031 (UGT1A, P interaction = .01), and rs798766 (TMEM129-TACC3-FGFR3, P interaction = .03). There was limited evidence for multiplicative interactions. When we examined detailed data on a prevalent occupational exposure associated with increased bladder cancer risk, straight metalworking fluids, we also observed statistically significant additive interaction for rs798766 (TMEM129-TACC3-FGFR3, P interaction = .02), with the interaction more apparent in patients with tumors positive for FGFR3 expression. All statistical tests were two-sided. The interaction we observed for rs798766 (TMEM129-TACC3-FGFR3) with specific exposure to straight metalworking fluids illustrates the value of integrating germline genetic variation, environmental exposures, and tumor marker data to provide insight into the mechanisms of bladder carcinogenesis. PMID:26374428

  18. Elimination of ergovaline from a grass–Neotyphodium endophyte symbiosis by genetic modification of the endophyte

    PubMed Central

    Panaccione, Daniel G.; Johnson, Richard D.; Wang, Jinghong; Young, Carolyn A.; Damrongkool, Prapassorn; Scott, Barry; Schardl, Christopher L.

    2001-01-01

    The fungal endophytes Neotyphodium lolii and Neotyphodium sp. Lp1 from perennial ryegrass (Lolium perenne), and related endophytes in other grasses, produce the ergopeptine toxin ergovaline, among other alkaloids, while also increasing plant fitness and resistance to biotic and abiotic stress. In the related fungus, Claviceps purpurea, the biosynthesis of ergopeptines requires the activities of two peptide synthetases, LPS1 and LPS2. A peptide synthetase gene hypothesized to be important for ergopeptine biosynthesis was identified in C. purpurea by its clustering with another ergot alkaloid biosynthetic gene, dmaW. Sequence analysis conducted independently of the research presented here indicates that this gene encodes LPS1 [Tudzynski, P., Holter, K., Correia, T., Arntz, C., Grammel, N. & Keller, U. (1999) Mol. Gen. Genet. 261, 133–141]. We have cloned a similar peptide synthetase gene from Neotyphodium lolii and inactivated it by gene knockout in Neotyphodium sp. Lp1. The resulting strain retained full compatibility with its perennial ryegrass host plant as assessed by immunoblotting of tillers and quantitative PCR. However, grass–endophyte associations containing the knockout strain did not produce detectable quantities of ergovaline as analyzed by HPLC with fluorescence detection. Disruption of this gene provides a means to manipulate the accumulation of ergovaline in endophyte-infected grasses for the purpose of determining the roles of ergovaline in endophyte-associated traits and, potentially, for ameliorating toxicoses in livestock. PMID:11592979

  19. Brain Tumor Genetic Modification Yields Increased Resistance to Paclitaxel in Physical Confinement.

    PubMed

    Bui, Loan; Hendricks, Alissa; Wright, Jamie; Chuong, Cheng-Jen; Davé, Digant; Bachoo, Robert; Kim, Young-Tae

    2016-01-01

    Brain tumor cells remain highly resistant to radiation and chemotherapy, particularly malignant and secondary cancers. In this study, we utilized microchannel devices to examine the effect of a confined environment on the viability and drug resistance of the following brain cancer cell lines: primary cancers (glioblastoma multiforme and neuroblastoma), human brain cancer cell lines (D54 and D54-EGFRvIII), and genetically modified mouse astrocytes (wild type, p53-/-, p53-/- PTEN-/-, p53-/- Braf, and p53-/- PTEN-/- Braf). We found that loss of PTEN combined with Braf activation resulted in higher viability in narrow microchannels. In addition, Braf conferred increased resistance to the microtubule-stabilizing drug Taxol in narrow confinement. Similarly, survival of D54-EGFRvIII cells was unaffected following treatment with Taxol, whereas the viability of D54 cells was reduced by 75% under these conditions. Taken together, our data suggests key targets for anticancer drugs based on cellular genotypes and their specific survival phenotypes during confined migration. PMID:27184621

  20. Genetic modification in Bacillus subtilis for production of C30 carotenoids.

    PubMed

    Maeda, Isamu

    2012-01-01

    C30 carotenoids, which have shorter backbones than C40 carotenoids, are known to be produced in the pathogenic bacterium Staphylococcus aureus that causes opportunistic infection. The first committed enzyme in the C30 carotenoid synthetic pathway is dehydrosqualene synthase CrtM. CrtM converts farnesyl pyrophosphate to dehydrosqualene. Dehydrosqualene desaturase CrtN then converts dehydrosqualene to the yellow C30 carotenoid, 4,4'-diaponeurosporene. This chapter describes a method to synthesize C30 carotenoids in Bacillus subtilis, which is generally recognized as a safe (GRAS) organism. Introduction of S. aureus crtM and crtN genes into B. subtilis results in yellow pigmentation. The B. subtilis transformant accumulates two C30 carotenoids, 4,4'-diapolycopene and 4,4'-diaponeurosporene. Furthermore, together with crtMN, introduction of S. aureus crtP and crtQ genes, which encode mixed function oxidase and glycosyltransferase, respectively, donates the ability to produce glycosylated C30 carotenoic acid. Thus, carotenoid biosynthesis genes of S. aureus is applicable to genetically modify B. subtilis in order to construct a safe organism producing C30 carotenoids.

  1. Genetic modification of neurons to express bevacizumab for local anti-angiogenesis treatment of glioblastoma.

    PubMed

    Hicks, M J; Funato, K; Wang, L; Aronowitz, E; Dyke, J P; Ballon, D J; Havlicek, D F; Frenk, E Z; De, B P; Chiuchiolo, M J; Sondhi, D; Hackett, N R; Kaminsky, S M; Tabar, V; Crystal, R G

    2015-01-01

    The median survival of glioblastoma multiforme (GBM) is approximately 1 year. Following surgical removal, systemic therapies are limited by the blood-brain barrier. To circumvent this, we developed a method to modify neurons with the genetic sequence for therapeutic monoclonal antibodies using adeno-associated virus (AAV) gene transfer vectors, directing persistent, local expression in the tumor milieu. The human U87MG GBM cell line or patient-derived early passage GBM cells were administered to the striatum of NOD/SCID immunodeficient mice. AAVrh.10BevMab, an AAVrh.10-based vector coding for bevacizumab (Avastin), an anti-human vascular endothelial growth factor (VEGF) monoclonal antibody, was delivered to the area of the GBM xenograft. Localized expression of bevacizumab was demonstrated by quantitative PCR, ELISA and western blotting. Immunohistochemistry showed that bevacizumab was expressed in neurons. Concurrent administration of AAVrh.10BevMab with the U87MG tumor reduced tumor blood vessel density and tumor volume, and increased survival. Administration of AAVrh.10BevMab 1 week after U87MG xenograft reduced growth and increased survival. Studies with patient-derived early passage GBM primary cells showed a reduction in primary tumor burden with an increased survival. These data support the strategy of AAV-mediated central nervous system gene therapy to treat GBM, overcoming the blood-brain barrier through local, persistent delivery of an anti-angiogenesis monoclonal antibody.

  2. Brain Tumor Genetic Modification Yields Increased Resistance to Paclitaxel in Physical Confinement

    PubMed Central

    Bui, Loan; Hendricks, Alissa; Wright, Jamie; Chuong, Cheng-Jen; Davé, Digant; Bachoo, Robert; Kim, Young-tae

    2016-01-01

    Brain tumor cells remain highly resistant to radiation and chemotherapy, particularly malignant and secondary cancers. In this study, we utilized microchannel devices to examine the effect of a confined environment on the viability and drug resistance of the following brain cancer cell lines: primary cancers (glioblastoma multiforme and neuroblastoma), human brain cancer cell lines (D54 and D54-EGFRvIII), and genetically modified mouse astrocytes (wild type, p53−/−, p53−/− PTEN−/−, p53−/− Braf, and p53−/− PTEN−/− Braf). We found that loss of PTEN combined with Braf activation resulted in higher viability in narrow microchannels. In addition, Braf conferred increased resistance to the microtubule-stabilizing drug Taxol in narrow confinement. Similarly, survival of D54-EGFRvIII cells was unaffected following treatment with Taxol, whereas the viability of D54 cells was reduced by 75% under these conditions. Taken together, our data suggests key targets for anticancer drugs based on cellular genotypes and their specific survival phenotypes during confined migration. PMID:27184621

  3. Expression of genes encoding for proteins involved in heparan sulphate and chondroitin sulphate chain synthesis and modification in normal and malignant plasma cells.

    PubMed

    Bret, Caroline; Hose, Dirk; Reme, Thierry; Sprynski, Anne-Catherine; Mahtouk, Karène; Schved, Jean-François; Quittet, Philippe; Rossi, Jean-François; Goldschmidt, Hartmut; Klein, Bernard

    2009-05-01

    Syndecan-1 is a proteoglycan that concentrates heparin-binding factors on the surface of multiple myeloma cells, and probably plays a major role in multiple myeloma biology. As heparan sulphate and chondroitin sulphate are the bioactive components of syndecan-1, we analysed the signature of genes encoding 100 proteins involved in synthesis of these chains, i.e. from precursor uptake to post-translational modifications, using Affymetrix microarrays. The expression of enzymes required for heparan sulphate and chondroitin sulphate biosynthesis was shown to increase in parallel with syndecan-1 expression, throughout the differentiation of memory B cells into plasmablasts and normal bone marrow plasma cells. Sixteen genes were significantly different between normal and malignant plasma cells, nine of these genes -EXT2, CHSY3, CSGALNACT1, HS3ST2, HS2ST1, CHST11, CSGALNACT2, HPSE, SULF2 - encode proteins involved in glycosaminoglycan chain synthesis or modifications. Kaplan-Meier analysis was performed in two independent series of patients: B4GALT7, CSGALNACT1, HS2ST1 were associated with a good prognosis whereas EXT1 was linked to a bad prognosis. This study provides an overall picture of the major genes encoding for proteins involved in heparan sulphate and chondroitin sulphate synthesis and modifications that can be implicated in normal and malignant plasma cells. PMID:19298595

  4. Common Genetic Variants and Modification of Penetrance of BRCA2-Associated Breast Cancer

    PubMed Central

    Guiducci, Candace; Segrè, Ayellet V.; McGee, Kate; McGuffog, Lesley; Kartsonaki, Christiana; Morrison, Jonathan; Healey, Sue; Sinilnikova, Olga M.; Stoppa-Lyonnet, Dominique; Mazoyer, Sylvie; Gauthier-Villars, Marion; Sobol, Hagay; Longy, Michel; Frenay, Marc; GEMO Study Collaborators; Hogervorst, Frans B. L.; Rookus, Matti A.; Collée, J. Margriet; Hoogerbrugge, Nicoline; van Roozendaal, Kees E. P.; Piedmonte, Marion; Rubinstein, Wendy; Nerenstone, Stacy; Van Le, Linda; Blank, Stephanie V.; Caldés, Trinidad; de la Hoya, Miguel; Nevanlinna, Heli; Aittomäki, Kristiina; Lazaro, Conxi; Blanco, Ignacio; Arason, Adalgeir; Johannsson, Oskar T.; Barkardottir, Rosa B.; Devilee, Peter; Olopade, Olofunmilayo I.; Neuhausen, Susan L.; Wang, Xianshu; Fredericksen, Zachary S.; Peterlongo, Paolo; Manoukian, Siranoush; Barile, Monica; Viel, Alessandra; Radice, Paolo; Phelan, Catherine M.; Narod, Steven; Rennert, Gad; Lejbkowicz, Flavio; Flugelman, Anath; Andrulis, Irene L.; Glendon, Gord; Ozcelik, Hilmi; Toland, Amanda E.; Montagna, Marco; D'Andrea, Emma; Friedman, Eitan; Laitman, Yael; Borg, Ake; Beattie, Mary; Ramus, Susan J.; Domchek, Susan M.; Nathanson, Katherine L.; Rebbeck, Tim; Spurdle, Amanda B.; Chen, Xiaoqing; Holland, Helene; John, Esther M.; Hopper, John L.; Buys, Saundra S.; Daly, Mary B.; Southey, Melissa C.; Terry, Mary Beth; Tung, Nadine; Overeem Hansen, Thomas V.; Nielsen, Finn C.; Greene, Mark I.; Mai, Phuong L.; Osorio, Ana; Durán, Mercedes; Andres, Raquel; Benítez, Javier; Weitzel, Jeffrey N.; Garber, Judy; Hamann, Ute; Peock, Susan; Cook, Margaret; Oliver, Clare; Frost, Debra; Platte, Radka; Evans, D. Gareth; Lalloo, Fiona; Eeles, Ros; Izatt, Louise; Walker, Lisa; Eason, Jacqueline; Barwell, Julian; Godwin, Andrew K.; Schmutzler, Rita K.; Wappenschmidt, Barbara; Engert, Stefanie; Arnold, Norbert; Gadzicki, Dorothea; Dean, Michael; Gold, Bert; Klein, Robert J.; Couch, Fergus J.; Chenevix-Trench, Georgia; Easton, Douglas F.; Daly, Mark J.; Antoniou, Antonis C.; Altshuler, David M.; Offit, Kenneth

    2010-01-01

    The considerable uncertainty regarding cancer risks associated with inherited mutations of BRCA2 is due to unknown factors. To investigate whether common genetic variants modify penetrance for BRCA2 mutation carriers, we undertook a two-staged genome-wide association study in BRCA2 mutation carriers. In stage 1 using the Affymetrix 6.0 platform, 592,163 filtered SNPs genotyped were available on 899 young (<40 years) affected and 804 unaffected carriers of European ancestry. Associations were evaluated using a survival-based score test adjusted for familial correlations and stratified by country of the study and BRCA2*6174delT mutation status. The genomic inflation factor (λ) was 1.011. The stage 1 association analysis revealed multiple variants associated with breast cancer risk: 3 SNPs had p-values<10−5 and 39 SNPs had p-values<10−4. These variants included several previously associated with sporadic breast cancer risk and two novel loci on chromosome 20 (rs311499) and chromosome 10 (rs16917302). The chromosome 10 locus was in ZNF365, which contains another variant that has recently been associated with breast cancer in an independent study of unselected cases. In stage 2, the top 85 loci from stage 1 were genotyped in 1,264 cases and 1,222 controls. Hazard ratios (HR) and 95% confidence intervals (CI) for stage 1 and 2 were combined and estimated using a retrospective likelihood approach, stratified by country of residence and the most common mutation, BRCA2*6174delT. The combined per allele HR of the minor allele for the novel loci rs16917302 was 0.75 (95% CI 0.66–0.86, ) and for rs311499 was 0.72 (95% CI 0.61–0.85, ). FGFR2 rs2981575 had the strongest association with breast cancer risk (per allele HR = 1.28, 95% CI 1.18–1.39, ). These results indicate that SNPs that modify BRCA2 penetrance identified by an agnostic approach thus far are limited to variants that also modify risk of sporadic BRCA2 wild-type breast cancer. PMID:21060860

  5. Optimization of methods for the genetic modification of human T cells.

    PubMed

    Bilal, Mahmood Y; Vacaflores, Aldo; Houtman, Jon Cd

    2015-11-01

    CD4(+) T cells are not only critical in the fight against parasitic, bacterial and viral infections, but are also involved in many autoimmune and pathological disorders. Studies of protein function in human T cells are confined to techniques such as RNA interference (RNAi) owing to ethical reasons and relative simplicity of these methods. However, introduction of RNAi or genes into primary human T cells is often hampered by toxic effects from transfection or transduction methods that yield cell numbers inadequate for downstream assays. Additionally, the efficiency of recombinant DNA expression is frequently low because of multiple factors including efficacy of the method and strength of the targeting RNAs. Here, we describe detailed protocols that will aid in the study of primary human CD4(+) T cells. First, we describe a method for development of effective microRNA/shRNAs using available online algorithms. Second, we illustrate an optimized protocol for high efficacy retroviral or lentiviral transduction of human T-cell lines. Importantly, we demonstrate that activated primary human CD4(+) T cells can be transduced efficiently with lentiviruses, with a highly activated population of T cells receiving the largest number of copies of integrated DNA. We also illustrate a method for efficient lentiviral transduction of hard-to-transduce un-activated primary human CD4(+) T cells. These protocols will significantly assist in understanding the activation and function of human T cells and will ultimately aid in the development or improvement of current drugs that target human CD4(+) T cells.

  6. Optimization of Methods for the Genetic Modification of Human T Cells

    PubMed Central

    Bilal, Mahmood Y.; Vacaflores, Aldo; Houtman, Jon C.D.

    2015-01-01

    CD4+ T cells are critical in the fight against parasitic, bacterial, and viral infections, but are also involved in many autoimmune and pathological disorders. Studies of protein function in human T cells are confined to techniques such as RNAi due to ethical reasons and relative simplicity of these methods. However, introduction of RNAi or genes into primary human T cells is often hampered by toxic effects from transfection or transduction methods that yield cell numbers inadequate for downstream assays. Additionally, the efficiency of recombinant DNA expression is frequently low due to multiple factors including efficacy of the method and strength of the targeting RNAs. Here, we describe detailed protocols that will aid in the study of primary human CD4+ T cells. First, we describe a method for development of effective microRNA/shRNAs using available online algorithms. Second, we illustrate an optimized protocol for high efficacy retroviral or lentiviral transduction of human T cell lines. Importantly, we demonstrate that activated primary human CD4+ T cells can be transduced efficiently with lentiviruses, with a highly activated population of T cells receiving the largest number of copies of integrated DNA. We also illustrate a method for efficient lentiviral transduction of hard-to-transduce un-activated primary human CD4+ T cells. These protocols will significantly assist in understanding the activation and function of human T cells and will ultimately aid in the development or improvement of current drugs that target human CD4+ T cells. PMID:26027856

  7. Genetic Modification of the Salmonella Membrane Physical State Alters the Pattern of Heat Shock Response ▿

    PubMed Central

    Porta, Amalia; Török, Zsolt; Horvath, Ibolya; Franceschelli, Silvia; Vígh, László; Maresca, Bruno

    2010-01-01

    It is now recognized that membranes are not simple physical barriers but represent a complex and dynamic environment that affects membrane protein structures and their functions. Recent data emphasize the role of membranes in sensing temperature changes, and it has been shown that the physical state of the plasma membrane influences the expression of a variety of genes such as heat shock genes. It has been widely shown that minor alterations in lipid membranes are critically involved in the conversion of signals from the environment to the transcriptional activation of heat shock genes. Previously, we have proposed that the composition, molecular arrangement, and physical state of lipid membranes and their organization have crucial roles in cellular responses during stress caused by physical and chemical factors as well as in pathological states. Here, we show that transformation of Salmonella enterica serovar Typhimurium LT2 (Salmonella Typhimurium) with a heterologous Δ12-desaturase (or with its trans-membrane regions) causes major changes in the pathogen's membrane dynamic. In addition, this pathogen is strongly impaired in the synthesis of major stress proteins (heat shock proteins) under heat shock. These data support the hypothesis that the perception of temperature in Salmonella is strictly controlled by membrane order and by a specific membrane lipid/protein ratio that ultimately causes transcriptional activation of heat shock genes. These results represent a previously unrecognized mode of sensing temperature variation used by this pathogen at the onset of infection. PMID:20139186

  8. Recent advances in the molecular genetics of hereditary retinal dystrophies with primary involvement of the macula.

    PubMed

    Weber, B H

    1998-01-01

    Hereditary dystrophies of the central retina and choroid are a heterogeneous group of disorders characterized by preferential loss of macular function and consequently loss of central and color vision. The primary causes leading to the degenerative processes are largely unknown although recent progress in human molecular genetics is most promising in providing novel insights into the basic biochemical mechanisms of these dystrophies. To date, the disease loci of more than 20 maculopathies including cone and cone-rod dystrophies have been mapped to specific chromosomal regions of which seven disease genes have already been identified. As the goals of the Human Genome Initiative approach completion, the cloning of the genes involved in the etiology of human retinopathies will be greatly simplified providing the basis for a more comprehensive understanding of retinal function and dysfunction. In addition, these advances will facilitate the identification of individuals at risk at a presymptomatic or initial stage of disease, thus creating a unique opportunity to devise novel therapeutic strategies that will primarily be aimed at an early intervention with the potential to either delay or even prevent the development of disease pathology.

  9. A Forward Genetic Screen for Molecules Involved in Pheromone-Induced Dauer Formation in Caenorhabditis elegans

    PubMed Central

    Neal, Scott J.; Park, JiSoo; DiTirro, Danielle; Yoon, Jason; Shibuya, Mayumi; Choi, Woochan; Schroeder, Frank C.; Butcher, Rebecca A.; Kim, Kyuhyung; Sengupta, Piali

    2016-01-01

    Animals must constantly assess their surroundings and integrate sensory cues to make appropriate behavioral and developmental decisions. Pheromones produced by conspecific individuals provide critical information regarding environmental conditions. Ascaroside pheromone concentration and composition are instructive in the decision of Caenorhabditis elegans to either develop into a reproductive adult or enter into the stress-resistant alternate dauer developmental stage. Pheromones are sensed by a small set of sensory neurons, and integrated with additional environmental cues, to regulate neuroendocrine signaling and dauer formation. To identify molecules required for pheromone-induced dauer formation, we performed an unbiased forward genetic screen and identified phd (pheromone response-defective dauer) mutants. Here, we describe new roles in dauer formation for previously identified neuronal molecules such as the WD40 domain protein QUI-1 and MACO-1 Macoilin, report new roles for nociceptive neurons in modulating pheromone-induced dauer formation, and identify tau tubulin kinases as new genes involved in dauer formation. Thus, phd mutants define loci required for the detection, transmission, or integration of pheromone signals in the regulation of dauer formation. PMID:26976437

  10. Chemical genetic screen for AMPKα2 substrates uncovers a network of proteins involved in mitosis

    PubMed Central

    Banko, Max R.; Allen, Jasmina J.; Schaffer, Bethany E.; Wilker, Erik W.; Tsou, Peiling; White, Jamie L.; Villén, Judit; Wang, Beatrice; Kim, Sara R.; Sakamoto, Kei; Gygi, Steven P.; Cantley, Lewis C.; Yaffe, Michael B.; Shokat, Kevan M.; Brunet, Anne

    2011-01-01

    SUMMARY The energy-sensing AMP-activated protein kinase (AMPK) is activated by low nutrient levels. Functions of AMPK, other than its role in cellular metabolism, are just beginning to emerge. Here we use a chemical genetics screen to identify direct substrates of AMPK in human cells. We find that AMPK phosphorylates 28 previously unidentified substrates, several of which are involved in mitosis and cytokinesis. We identify the residues phosphorylated by AMPK in vivo in several substrates, including protein phosphatase 1 regulatory subunit 12C (PPP1R12C) and p21 -activated protein kinase (PAK2). AMPK-induced phosphorylation is necessary for PPP1R12C interaction with 14-3-3 and phosphorylation of myosin regulatory light chain. Both AMPK activity and PPP1R12C phosphorylation are increased in mitotic cells and are important for mitosis completion. These findings suggest that AMPK coordinates nutrient status with mitosis completion, which may be critical for the organism’s response to low nutrients during development, or in adult stem and cancer cells. PMID:22137581

  11. Genetic Polymorphisms Involved in Folate Metabolism and Maternal Risk for Down Syndrome: A Meta-Analysis

    PubMed Central

    Balduino Victorino, Daniella; de Godoy, Moacir Fernandes; Goloni-Bertollo, Eny Maria; Pavarino, Érika Cristina

    2014-01-01

    Inconclusive results of the association between genetic polymorphisms involved in folate metabolism and maternal risk for Down syndrome (DS) have been reported. Therefore, this meta-analysis was conducted. We searched electronic databases through May, 2014, for eligible studies. Pooled odds ratios with 95% confidence intervals were used to assess the strength of the association, which was estimated by fixed or random effects models. Heterogeneity among studies was evaluated using Q-test and I2 statistic. Subgroup and sensitivity analyses were also conducted. Publication bias was estimated using Begg's and Egger's tests. A total of 17 case-controls studies were included. There was evidence for an association between the MTRR c.66A>G (rs1801394) polymorphism and maternal risk for DS. In the subgroup analysis, increased maternal risk for DS was found in Caucasians. Additionally, the polymorphic heterozygote MTHFD1 1958GA genotype was associated significantly with maternal risk for DS, when we limit the analysis by studies conformed to Hardy-Weinberg equilibrium. Finally, considering MTR c.2756A>G (rs1805087), TC2 c.776C>G (rs1801198), and CBS c.844ins68, no significant associations have been found, neither in the overall analyses nor in the stratified analyses by ethnicity. In conclusion, our meta-analysis suggested that the MTRR c.66A>G (rs1801394) polymorphism and MTHFD1 c.1958G>A (rs2236225) were associated with increased maternal risk for DS. PMID:25544792

  12. A Systems Biology Overview on Human Diabetic Nephropathy: From Genetic Susceptibility to Post-Transcriptional and Post-Translational Modifications

    PubMed Central

    Conserva, Francesca; Gesualdo, Loreto; Papale, Massimo

    2016-01-01

    Diabetic nephropathy (DN), a microvascular complication occurring in approximately 20–40% of patients with type 2 diabetes mellitus (T2DM), is characterized by the progressive impairment of glomerular filtration and the development of Kimmelstiel-Wilson lesions leading to end-stage renal failure (ESRD). The causes and molecular mechanisms mediating the onset of T2DM chronic complications are yet sketchy and it is not clear why disease progression occurs only in some patients. We performed a systematic analysis of the most relevant studies investigating genetic susceptibility and specific transcriptomic, epigenetic, proteomic, and metabolomic patterns in order to summarize the most significant traits associated with the disease onset and progression. The picture that emerges is complex and fascinating as it includes the regulation/dysregulation of numerous biological processes, converging toward the activation of inflammatory processes, oxidative stress, remodeling of cellular function and morphology, and disturbance of metabolic pathways. The growing interest in the characterization of protein post-translational modifications and the importance of handling large datasets using a systems biology approach are also discussed. PMID:26798653

  13. Establishment of a tamoxifen-inducible Cre-driver mouse strain for widespread and temporal genetic modification in adult mice.

    PubMed

    Ichise, Hirotake; Hori, Akiko; Shiozawa, Seiji; Kondo, Saki; Kanegae, Yumi; Saito, Izumu; Ichise, Taeko; Yoshida, Nobuaki

    2016-07-29

    Temporal genetic modification of mice using the ligand-inducible Cre/loxP system is an important technique that allows the bypass of embryonic lethal phenotypes and access to adult phenotypes. In this study, we generated a tamoxifen-inducible Cre-driver mouse strain for the purpose of widespread and temporal Cre recombination. The new line, named CM32, expresses the GFPneo-fusion gene in a wide variety of tissues before FLP recombination and tamoxifen-inducible Cre after FLP recombination. Using FLP-recombined CM32 mice (CM32Δ mice) and Cre reporter mouse lines, we evaluated the efficiency of Cre recombination with and without tamoxifen administration to adult mice, and found tamoxifen-dependent induction of Cre recombination in a variety of adult tissues. In addition, we demonstrated that conditional activation of an oncogene could be achieved in adults using CM32Δ mice. CM32Δ;T26 mice, which harbored a Cre recombination-driven, SV40 large T antigen-expressing transgene, were viable and fertile. No overt phenotype was found in the mice up to 3 months after birth. Although they displayed pineoblastomas (pinealoblastomas) and/or thymic enlargement due to background Cre recombination by 6 months after birth, they developed epidermal hyperplasia when administered tamoxifen. Collectively, our results suggest that the CM32Δ transgenic mouse line can be applied to the assessment of adult phenotypes in mice with loxP-flanked transgenes.

  14. A simple method to introduce marker-free genetic modifications into the chromosome of naturally nontransformable Bacillus amyloliquefaciens strains.

    PubMed

    Zakataeva, Natalia P; Nikitina, Oksana V; Gronskiy, Sergey V; Romanenkov, Dmitriy V; Livshits, Vitaliy A

    2010-01-01

    A simple method to introduce marker-free deletions, insertions, and point mutations into the chromosomes of naturally nontransformable Bacillus amyloliquefaciens strains has been developed. The method is efficient and fast, and it allows for the generation of genetic modifications without the use of a counter-selectable marker or a special prerequisite strain. This method uses the combination of the following: the effective introduction of a delivery plasmid into cells for gene replacement; a two-step replacement recombination procedure, which occurs at a very high frequency due to the use of a thermosensitive rolling-circle replication plasmid; and colony polymerase chain reaction (PCR) analysis for screening. Using PCR primers with mismatches at the 3' end enables the selection of strains that contain a single nucleotide substitution in the target gene. This approach can be used as a routine method for the investigation of complex physiological pathways and for the metabolic engineering of food-grade industrial B. amyloliquefaciens and other Bacillus strains.

  15. Establishment of a tamoxifen-inducible Cre-driver mouse strain for widespread and temporal genetic modification in adult mice

    PubMed Central

    Ichise, Hirotake; Hori, Akiko; Shiozawa, Seiji; Kondo, Saki; Kanegae, Yumi; Saito, Izumu; Ichise, Taeko; Yoshida, Nobuaki

    2016-01-01

    Temporal genetic modification of mice using the ligand-inducible Cre/loxP system is an important technique that allows the bypass of embryonic lethal phenotypes and access to adult phenotypes. In this study, we generated a tamoxifen-inducible Cre-driver mouse strain for the purpose of widespread and temporal Cre recombination. The new line, named CM32, expresses the GFPneo-fusion gene in a wide variety of tissues before FLP recombination and tamoxifen-inducible Cre after FLP recombination. Using FLP-recombined CM32 mice (CM32Δ mice) and Cre reporter mouse lines, we evaluated the efficiency of Cre recombination with and without tamoxifen administration to adult mice, and found tamoxifen-dependent induction of Cre recombination in a variety of adult tissues. In addition, we demonstrated that conditional activation of an oncogene could be achieved in adults using CM32Δ mice. CM32Δ;T26 mice, which harbored a Cre recombination-driven, SV40 large T antigen-expressing transgene, were viable and fertile. No overt phenotype was found in the mice up to 3 months after birth. Although they displayed pineoblastomas (pinealoblastomas) and/or thymic enlargement due to background Cre recombination by 6 months after birth, they developed epidermal hyperplasia when administered tamoxifen. Collectively, our results suggest that the CM32Δ transgenic mouse line can be applied to the assessment of adult phenotypes in mice with loxP-flanked transgenes. PMID:26923756

  16. Informed consent, participation in, and withdrawal from a population based cohort study involving genetic analysis

    PubMed Central

    Matsui, K; Kita, Y; Ueshima, H

    2005-01-01

    Design: Descriptive analyses. Setting and participants: The study evaluated two non-genetic subcohorts comprising 3166 people attending for a health checkup during 2002, and two genetic subcohorts comprising 2195 people who underwent a checkup during 2003. Main outcome measurements: Analysis endpoints were differences in participation rates between the non-genetic and genetic subcohorts, differences between providing non-extensive and extensive preliminary information, and changes in participation status between baseline and at 6 months. Results: Participation rates in the genetic subcohorts were 4·7–9·3% lower than those in the non-genetic subcohorts. The odds ratios (OR) of participation in genetic research were between 0·60 and 0·77, and the OR for withdrawal from the research was over 7·70; providing preliminary extensive information about genetic research reduced the withdrawal risks (OR 0·15 for all dependent variables) but worsened participation rates (OR 0·63–0·74). Conclusions: The general population responded sceptically towards genetic research. It is crucial that genetic researchers utilise an informative and educational consent process worthy of public trust. PMID:15994356

  17. Genetic and Epigenetic Modifications of Sox2 Contribute to the Invasive Phenotype of Malignant Gliomas

    PubMed Central

    Alonso, Marta M.; Diez-Valle, Ricardo; Manterola, Lorea; Rubio, Angel; Liu, Dan; Cortes-Santiago, Nahir; Urquiza, Leire; Jauregi, Patricia; de Munain, Adolfo Lopez; Sampron, Nicolás; Aramburu, Ander; Tejada-Solís, Sonia; Vicente, Carmen; Odero, María D.; Bandrés, Eva; García-Foncillas, Jesús; Idoate, Miguel A.; Lang, Frederick F.; Fueyo, Juan; Gomez-Manzano, Candelaria

    2011-01-01

    We undertook this study to understand how the transcription factor Sox2 contributes to the malignant phenotype of glioblastoma multiforme (GBM), the most aggressive primary brain tumor. We initially looked for unbalanced genomic rearrangements in the Sox2 locus in 42 GBM samples and found that Sox2 was amplified in 11.5% and overexpressed in all the samples. These results prompted us to further investigate the mechanisms involved in Sox2 overexpression in GBM. We analyzed the methylation status of the Sox2 promoter because high CpG density promoters are associated with key developmental genes. The Sox2 promoter presented a CpG island that was hypomethylated in all the patient samples when compared to normal cell lines. Treatment of Sox2-negative glioma cell lines with 5-azacitidine resulted in the re-expression of Sox2 and in a change in the methylation status of the Sox2 promoter. We further confirmed these results by analyzing data from GBM cases generated by The Cancer Genome Atlas project. We observed Sox2 overexpression (86%; N = 414), Sox2 gene amplification (8.5%; N = 492), and Sox 2 promoter hypomethylation (100%; N = 258), suggesting the relevance of this factor in the malignant phenotype of GBMs. To further explore the role of Sox2, we performed in vitro analysis with brain tumor stem cells (BTSCs) and established glioma cell lines. Downmodulation of Sox2 in BTSCs resulted in the loss of their self-renewal properties. Surprisingly, ectopic expression of Sox2 in established glioma cells was not sufficient to support self-renewal, suggesting that additional factors are required. Furthermore, we observed that ectopic Sox2 expression was sufficient to induce invasion and migration of glioma cells, and knockdown experiments demonstrated that Sox2 was essential for maintaining these properties. Altogether, our data underscore the importance of a pleiotropic role of Sox2 and suggest that it could be used as a therapeutic target in GBM. PMID:22069467

  18. Transcriptome-wide N⁶-methyladenosine profiling of rice callus and leaf reveals the presence of tissue-specific competitors involved in selective mRNA modification.

    PubMed

    Li, Yuli; Wang, Xiliang; Li, Cuiping; Hu, Songnian; Yu, Jun; Song, Shuhui

    2014-01-01

    N(6)-methyladenosine (m(6)A) is the most prevalent internal modification present in mRNAs of all higher eukaryotes. With the development of MeRIP-seq technique, in-depth identification of mRNAs with m(6)A modification becomes feasible. Here we present a transcriptome-wide m(6)A modification profiling effort for rice transcriptomes of differentiated callus and leaf, which yields 8,138 and 14,253 m(6)A-modified genes, respectively. The m(6)A peak (m(6)A-modified nucleotide position on mRNAs) distribution exhibits preference toward both translation termination and initiation sites. The m(6)A peak enrichment is negatively correlated with gene expression and weakly positively correlated with certain gene features, such as exon length and number. By comparing m(6)A-modified genes between the 2 samples, we define 1,792 and 6,508 tissue-specific m(6)A-modified genes (TSMGs) in callus and leaf, respectively. Among which, 626 and 5,509 TSMGs are actively expressed in both tissues but are selectively m(6)A-modified (SMGs) only in one of the 2 tissues. Further analyses reveal characteristics of SMGs: (1) Most SMGs are differentially expressed between callus and leaf. (2) Two conserved RNA-binding motifs, predicted to be recognized by PUM and RNP4F, are significantly over-represented in SMGs. (3) GO enrichment analysis shows that SMGs in callus mainly participate in transcription regulator/factor activity whereas SMGs in leaf are mainly involved in plastid and thylakoid. Our results suggest the presence of tissue-specific competitors involved in SMGs. These findings provide a resource for plant RNA epitranscriptomic studies and further enlarge our knowledge on the function of RNA m(6)A modification. PMID:25483034

  19. Transcriptome-wide N⁶-methyladenosine profiling of rice callus and leaf reveals the presence of tissue-specific competitors involved in selective mRNA modification.

    PubMed

    Li, Yuli; Wang, Xiliang; Li, Cuiping; Hu, Songnian; Yu, Jun; Song, Shuhui

    2014-01-01

    N(6)-methyladenosine (m(6)A) is the most prevalent internal modification present in mRNAs of all higher eukaryotes. With the development of MeRIP-seq technique, in-depth identification of mRNAs with m(6)A modification becomes feasible. Here we present a transcriptome-wide m(6)A modification profiling effort for rice transcriptomes of differentiated callus and leaf, which yields 8,138 and 14,253 m(6)A-modified genes, respectively. The m(6)A peak (m(6)A-modified nucleotide position on mRNAs) distribution exhibits preference toward both translation termination and initiation sites. The m(6)A peak enrichment is negatively correlated with gene expression and weakly positively correlated with certain gene features, such as exon length and number. By comparing m(6)A-modified genes between the 2 samples, we define 1,792 and 6,508 tissue-specific m(6)A-modified genes (TSMGs) in callus and leaf, respectively. Among which, 626 and 5,509 TSMGs are actively expressed in both tissues but are selectively m(6)A-modified (SMGs) only in one of the 2 tissues. Further analyses reveal characteristics of SMGs: (1) Most SMGs are differentially expressed between callus and leaf. (2) Two conserved RNA-binding motifs, predicted to be recognized by PUM and RNP4F, are significantly over-represented in SMGs. (3) GO enrichment analysis shows that SMGs in callus mainly participate in transcription regulator/factor activity whereas SMGs in leaf are mainly involved in plastid and thylakoid. Our results suggest the presence of tissue-specific competitors involved in SMGs. These findings provide a resource for plant RNA epitranscriptomic studies and further enlarge our knowledge on the function of RNA m(6)A modification.

  20. Detection of modifications in the glucose metabolism induced by genetic mutations in Saccharomyces cerevisiae by 13C- and H-NMR spectroscopy.

    PubMed

    Herve, M; Buffin-Meyer, B; Bouet, F; Son, T D

    2000-06-01

    NMR spectroscopy may offer a suitable technique to characterize the glucose metabolism in response to genetic mutations in cells. The effects of various genetic modifications in Saccharomyces cerevisiae yeast were investigated using 13C- and 1H-NMR spectroscopy associated with biochemical techniques. Cells were incubated with [1-13C]glucose in order to study glucose consumption and the formation of various end-products (ethanol, trehalose, glycerol, glutamate and amino acids) as a function of time. Two types of genetic modifications were studied in S. cerevisiae. A genetic modification deleted the N-terminal part of the TFC7 protein which is the smallest subunit (tau55) of the TFIIIC transcription factor. One secondary effect of this mutation was a large deletion of mitochondrial DNA giving the rho-phenotype. The other genetic modification corresponded to the disruption of the HUF gene; the mutated cells were rho+ like the reference strain. Both mutations increase the glycolysis rate and glycerol synthesis and decrease trehalose production. The most modified cells, which contain both TFC7 deletion and HUF gene disruption, utilize glucose in the most extreme manner as in these cells the largest production of the two glycolytic products (ethanol and glycerol) and the smallest trehalose formation occur. The HUF gene disruption serves as a positive modulator of glycolysis and respiration. However, the TFC7 deletion, associated with the phenotype rho-, induces the most damage in the cellular function, dramatically altering the behaviour of the Krebs cycle. The cycle becomes blocked at the level of 2-oxoglutarate, detected by a characteristic pattern of the 13C-NMR glutamate spectra. These NMR spectra corroborate the phenotypic data, the rho-phenotype corresponding to deletions of mitochondria DNA which block all mitochondria protein synthesis and render the cells unable to derive energy from respiration. Moreover, as a consequence of the Krebs cycle blocking, alanine

  1. Epidemiological and genetic clues for molecular mechanisms involved in uterine leiomyoma development and growth

    PubMed Central

    Commandeur, Arno E.; Styer, Aaron K.; Teixeira, Jose M.

    2015-01-01

    BACKGROUND Uterine leiomyomas (fibroids) are highly prevalent benign smooth muscle tumors of the uterus. In the USA, the lifetime risk for women developing uterine leiomyomas is estimated as up to 75%. Except for hysterectomy, most therapies or treatments often provide only partial or temporary relief and are not successful in every patient. There is a clear racial disparity in the disease; African-American women are estimated to be three times more likely to develop uterine leiomyomas and generally develop more severe symptoms. There is also familial clustering between first-degree relatives and twins, and multiple inherited syndromes in which fibroid development occurs. Leiomyomas have been described as clonal and hormonally regulated, but despite the healthcare burden imposed by the disease, the etiology of uterine leiomyomas remains largely unknown. The mechanisms involved in their growth are also essentially unknown, which has contributed to the slow progress in development of effective treatment options. METHODS A comprehensive PubMed search for and critical assessment of articles related to the epidemiological, biological and genetic clues for uterine leiomyoma development was performed. The individual functions of some of the best candidate genes are explained to provide more insight into their biological function and to interconnect and organize genes and pathways in one overarching figure that represents the current state of knowledge about uterine leiomyoma development and growth. RESULTS In this review, the widely recognized roles of estrogen and progesterone in uterine leiomyoma pathobiology on the basis of clinical and experimental data are presented. This is followed by fundamental aspects and concepts including the possible cellular origin of uterine fibroids. The central themes in the subsequent parts are cytogenetic aberrations in leiomyomas and the racial/ethnic disparities in uterine fibroid biology. Then, the attributes of various in vitro and

  2. Environmental Moderators of Genetic Influences on Adolescent Delinquent Involvement and Victimization

    ERIC Educational Resources Information Center

    Beaver, Kevin M.

    2011-01-01

    A growing body of empirical research reveals that genetic factors account for a substantial amount of variance in measures of antisocial behaviors. At the same time, evidence is also emerging indicating that certain environmental factors moderate the effects that genetic factors have on antisocial outcomes. Despite this line of research, much…

  3. Huntingtin's WW domain partners in Huntington's disease post-mortem brain fulfill genetic criteria for direct involvement in Huntington's disease pathogenesis.

    PubMed

    Passani, L A; Bedford, M T; Faber, P W; McGinnis, K M; Sharp, A H; Gusella, J F; Vonsattel, J P; MacDonald, M E

    2000-09-01

    An elongated glutamine tract in mutant huntingtin initiates Huntington's disease (HD) pathogenesis via a novel structural property that displays neuronal selectivity, glutamine progressivity and dominance over the normal protein based on genetic criteria. As this mechanism is likely to involve a deleterious protein interaction, we have assessed the major class of huntingtin interactors comprising three WW domain proteins. These are revealed to be related spliceosome proteins (HYPA/FBP-11 and HYPC) and a transcription factor (HYPB) that implicate huntingtin in mRNA biogenesis. In HD post-mortem brain, specific antibody reagents detect each partner in HD target neurons, in association with disease-related N-terminal morphologic deposits but not with filter trapped insoluble-aggregate. Glutathione S:-transferase partner 'pull-down' assays reveal soluble, aberrantly migrating, forms of full-length mutant huntingtin specific to HD target tissue. Importantly, these novel mutant species exhibit exaggerated WW domain binding that abrogates partner association with other huntingtin isoforms. Thus, each WW domain partner's association with huntingtin fulfills HD genetic criteria, supporting a direct role in pathogenesis. Our findings indicate that modification of mutant huntingtin in target neurons may promote an abnormal interaction with one, or all, of huntingtin's WW domain partners, perhaps altering ribonucleoprotein function with toxic consequences.

  4. Lysine pyrrolation is a naturally-occurring covalent modification involved in the production of DNA mimic proteins

    PubMed Central

    Miyashita, Hiroaki; Chikazawa, Miho; Otaki, Natsuki; Hioki, Yusuke; Shimozu, Yuki; Nakashima, Fumie; Shibata, Takahiro; Hagihara, Yoshihisa; Maruyama, Shoichi; Matsumi, Noriyoshi; Uchida, Koji

    2014-01-01

    Covalent modification of proteins exerts significant effects on their chemical properties and has important functional and regulatory consequences. We now report the identification and verification of an electrically-active form of modified proteins recognized by a group of small molecules commonly used to interact with DNA. This previously unreported property of proteins was initially discovered when the γ-ketoaldehydes were identified as a source of the proteins stained by the DNA intercalators. Using 1,4-butanedial, the simplest γ-ketoaldehyde, we characterized the structural and chemical criteria governing the recognition of the modified proteins by the DNA intercalators and identified Nε-pyrrolelysine as a key adduct. Unexpectedly, the pyrrolation conferred an electronegativity and electronic properties on the proteins that potentially constitute an electrical mimic to the DNA. In addition, we found that the pyrrolated proteins indeed triggered an autoimmune response and that the production of specific antibodies against the pyrrolated proteins was accelerated in human systemic lupus erythematosus. These findings and the apparent high abundance of Nε-pyrrolelysine in vivo suggest that protein pyrrolation could be an endogenous source of DNA mimic proteins, providing a possible link connecting protein turnover and immune disorders. PMID:24938734

  5. Molecular evolution and population genetics of two Drosophila mettleri cytochrome P450 genes involved in host plant utilization.

    PubMed

    Bono, Jeremy M; Matzkin, Luciano M; Castrezana, Sergio; Markow, Therese A

    2008-07-01

    Understanding the genetic basis of adaptation is one of the primary goals of evolutionary biology. The evolution of xenobiotic resistance in insects has proven to be an especially suitable arena for studying the genetics of adaptation, and resistant phenotypes are known to result from both coding and regulatory changes. In this study, we examine the evolutionary history and population genetics of two Drosophila mettleri cytochrome P450 genes that are putatively involved in the detoxification of alkaloids present in two of its cactus hosts: saguaro (Carnegiea gigantea) and senita (Lophocereus schottii). Previous studies demonstrated that Cyp28A1 was highly up-regulated following exposure to rotting senita tissue while Cyp4D10 was highly up-regulated following exposure to rotting saguaro tissue. Here, we show that a subset of sites in Cyp28A1 experienced adaptive evolution specifically in the D. mettleri lineage. Moreover, neutrality tests in several populations were also consistent with a history of selection on Cyp28A1. In contrast, we did not find evidence for positive selection on Cyp4D10, although this certainly does not preclude its involvement in host plant use. A surprising result that emerged from our population genetic analyses was the presence of significant genetic differentiation between flies collected from different host plant species (saguaro and senita) at Organ Pipe National Monument, Arizona, USA. This preliminary evidence suggests that D. mettleri may have evolved into distinctive host races that specialize on different hosts, a possibility that warrants further investigation. PMID:18510584

  6. Searching Online Mendelian Inheritance in Man (OMIM) for information on genetic loci involved in human disease.

    PubMed

    Baxevanis, Andreas D

    2012-04-01

    Online Mendelian Inheritance in Man (OMIM) is a comprehensive compendium of information on human genes and genetic disorders, with a particular emphasis on the interplay between observed phenotypes and underlying genotypes. This unit focuses on the basic methodology for formulating OMIM searches and illustrates the types of information that can be retrieved from OMIM, including descriptions of clinical manifestations resulting from genetic abnormalities. This unit also provides information on additional relevant medical and molecular biology databases. A basic knowledge of OMIM should be part of the armamentarium of physicians and scientists with an interest in research on the clinical aspects of genetic disorders.

  7. Genetic alterations of JAK/STAT cascade and histone modification in extranodal NK/T-cell lymphoma nasal type.

    PubMed

    Lee, Seungbok; Park, Ha Young; Kang, So Young; Kim, Seok Jin; Hwang, Jinha; Lee, Seungho; Kwak, Soo Heon; Park, Kyong Soo; Yoo, Hae Yong; Kim, Won Seog; Kim, Jong-Il; Ko, Young Hyeh

    2015-07-10

    Extranodal NK/T-cell lymphoma nasal type (ENKL) is a rare type of non-Hodgkin lymphoma that more frequently occurs in East Asia and Latin America. Even though its molecular background has been discussed in the last few years, the current knowledge does not explain the disease pathogenesis in most cases of ENKL. Here, we performed multiple types of next-generation sequencing on 34 ENKL samples, including whole-exome sequencing (9 cancer tissues and 4 cancer cell lines), targeted sequencing (21 cancer tissues), and RNA sequencing (3 cancer tissues and 4 cancer cell lines). Mutations were found most frequently in 3 genes, STAT3, BCOR, and MLL2 (which were present in 9, 7, and 6 cancer samples, respectively), whereas there were only 2 cases of JAK3 mutation. In total, JAK/STAT pathway- and histone modification-related genes accounted for 55.9% and 38.2% of cancer samples, respectively, and their involvement in ENKL pathogenesis was also supported by gene expression analysis. In addition, we provided 177 genes upregulated only in cancer tissues, which appear to be linked with angiocentric and angiodestructive growth of ENKL. In this study, we propose several novel driver genes of ENKL, and show that these genes and their functional groups may be future therapeutic targets of this disease.

  8. Genetic alterations of JAK/STAT cascade and histone modification in extranodal NK/T-cell lymphoma nasal type

    PubMed Central

    Kang, So Young; Kim, Seok Jin; Hwang, Jinha; Lee, Seungho; Kwak, Soo Heon; Park, Kyong Soo; Yoo, Hae Yong

    2015-01-01

    Extranodal NK/T-cell lymphoma nasal type (ENKL) is a rare type of non-Hodgkin lymphoma that more frequently occurs in East Asia and Latin America. Even though its molecular background has been discussed in the last few years, the current knowledge does not explain the disease pathogenesis in most cases of ENKL. Here, we performed multiple types of next-generation sequencing on 34 ENKL samples, including whole-exome sequencing (9 cancer tissues and 4 cancer cell lines), targeted sequencing (21 cancer tissues), and RNA sequencing (3 cancer tissues and 4 cancer cell lines). Mutations were found most frequently in 3 genes, STAT3, BCOR, and MLL2 (which were present in 9, 7, and 6 cancer samples, respectively), whereas there were only 2 cases of JAK3 mutation. In total, JAK/STAT pathway- and histone modification-related genes accounted for 55.9% and 38.2% of cancer samples, respectively, and their involvement in ENKL pathogenesis was also supported by gene expression analysis. In addition, we provided 177 genes upregulated only in cancer tissues, which appear to be linked with angiocentric and angiodestructive growth of ENKL. In this study, we propose several novel driver genes of ENKL, and show that these genes and their functional groups may be future therapeutic targets of this disease. PMID:25980440

  9. Genetic alterations of JAK/STAT cascade and histone modification in extranodal NK/T-cell lymphoma nasal type.

    PubMed

    Lee, Seungbok; Park, Ha Young; Kang, So Young; Kim, Seok Jin; Hwang, Jinha; Lee, Seungho; Kwak, Soo Heon; Park, Kyong Soo; Yoo, Hae Yong; Kim, Won Seog; Kim, Jong-Il; Ko, Young Hyeh

    2015-07-10

    Extranodal NK/T-cell lymphoma nasal type (ENKL) is a rare type of non-Hodgkin lymphoma that more frequently occurs in East Asia and Latin America. Even though its molecular background has been discussed in the last few years, the current knowledge does not explain the disease pathogenesis in most cases of ENKL. Here, we performed multiple types of next-generation sequencing on 34 ENKL samples, including whole-exome sequencing (9 cancer tissues and 4 cancer cell lines), targeted sequencing (21 cancer tissues), and RNA sequencing (3 cancer tissues and 4 cancer cell lines). Mutations were found most frequently in 3 genes, STAT3, BCOR, and MLL2 (which were present in 9, 7, and 6 cancer samples, respectively), whereas there were only 2 cases of JAK3 mutation. In total, JAK/STAT pathway- and histone modification-related genes accounted for 55.9% and 38.2% of cancer samples, respectively, and their involvement in ENKL pathogenesis was also supported by gene expression analysis. In addition, we provided 177 genes upregulated only in cancer tissues, which appear to be linked with angiocentric and angiodestructive growth of ENKL. In this study, we propose several novel driver genes of ENKL, and show that these genes and their functional groups may be future therapeutic targets of this disease. PMID:25980440

  10. Nucleotide modifications within bacterial messenger RNAs regulate their translation and are able to rewire the genetic code

    PubMed Central

    Hoernes, Thomas Philipp; Clementi, Nina; Faserl, Klaus; Glasner, Heidelinde; Breuker, Kathrin; Lindner, Herbert; Hüttenhofer, Alexander; Erlacher, Matthias David

    2016-01-01

    Nucleotide modifications within RNA transcripts are found in every organism in all three domains of life. 6-methyladeonsine (m6A), 5-methylcytosine (m5C) and pseudouridine (Ψ) are highly abundant nucleotide modifications in coding sequences of eukaryal mRNAs, while m5C and m6A modifications have also been discovered in archaeal and bacterial mRNAs. Employing in vitro translation assays, we systematically investigated the influence of nucleotide modifications on translation. We introduced m5C, m6A, Ψ or 2′-O-methylated nucleotides at each of the three positions within a codon of the bacterial ErmCL mRNA and analyzed their influence on translation. Depending on the respective nucleotide modification, as well as its position within a codon, protein synthesis remained either unaffected or was prematurely terminated at the modification site, resulting in reduced amounts of the full-length peptide. In the latter case, toeprint analysis of ribosomal complexes was consistent with stalling of translation at the modified codon. When multiple nucleotide modifications were introduced within one codon, an additive inhibitory effect on translation was observed. We also identified the m5C modification to alter the amino acid identity of the corresponding codon, when positioned at the second codon position. Our results suggest a novel mode of gene regulation by nucleotide modifications in bacterial mRNAs. PMID:26578598

  11. Indole-3-acetic acid UDP-glucosyltransferase from immature seeds of pea is involved in modification of glycoproteins.

    PubMed

    Ostrowski, Maciej; Hetmann, Anna; Jakubowska, Anna

    2015-09-01

    The glycosylation of auxin is one of mechanisms contributing to hormonal homeostasis. The enzyme UDPG: indole-3-ylacetyl-β-D-glucosyltransferase (IAA glucosyltransferase, IAGlc synthase) catalyzes the reversible reaction: IAA+UDPG↔1-O-IA-glucose+UDP, which is the first step in the biosynthesis of IAA-ester conjugates in monocotyledonous plants. In this study, we report IAA-glucosyltransferase isolated using a biochemical approach from immature seed of pea (Pisum sativum). The enzyme was purified by PEG fractionation, DEAE-Sephacel anion-exchange chromatography and preparative PAGE. LC-MS/MS analysis of tryptic peptides of the enzyme revealed the high identity with maize IAGlc synthase, but lack of homology with other IAA-glucosyltransferases from dicots. Biochemical characterization showed that of several acyl acceptors tested, the enzyme had the highest activity on IAA as the glucosyl acceptor (Km=0.52 mM, Vmax=161 nmol min(-1), kcat/Km=4.36 mM s(-1)) and lower activity on indole-3-propionic acid and 1-naphthalene acetic acid. Whereas indole-3-butyric acid and indole-3-propionic acid were competitive inhibitors of IAGlc synthase, D-gluconic acid lactone, an inhibitor of β-glucosidase activity, potentiated the enzyme activity at the optimal concentration of 0.3mM. Moreover, we demonstrated that the 1-O-IA-glucose synthesized by IAGlc synthase is the substrate for IAA labeling of glycoproteins from pea seeds indicating a possible role of this enzyme in the covalent modification of a class of proteins by a plant hormone.

  12. Indole-3-acetic acid UDP-glucosyltransferase from immature seeds of pea is involved in modification of glycoproteins.

    PubMed

    Ostrowski, Maciej; Hetmann, Anna; Jakubowska, Anna

    2015-09-01

    The glycosylation of auxin is one of mechanisms contributing to hormonal homeostasis. The enzyme UDPG: indole-3-ylacetyl-β-D-glucosyltransferase (IAA glucosyltransferase, IAGlc synthase) catalyzes the reversible reaction: IAA+UDPG↔1-O-IA-glucose+UDP, which is the first step in the biosynthesis of IAA-ester conjugates in monocotyledonous plants. In this study, we report IAA-glucosyltransferase isolated using a biochemical approach from immature seed of pea (Pisum sativum). The enzyme was purified by PEG fractionation, DEAE-Sephacel anion-exchange chromatography and preparative PAGE. LC-MS/MS analysis of tryptic peptides of the enzyme revealed the high identity with maize IAGlc synthase, but lack of homology with other IAA-glucosyltransferases from dicots. Biochemical characterization showed that of several acyl acceptors tested, the enzyme had the highest activity on IAA as the glucosyl acceptor (Km=0.52 mM, Vmax=161 nmol min(-1), kcat/Km=4.36 mM s(-1)) and lower activity on indole-3-propionic acid and 1-naphthalene acetic acid. Whereas indole-3-butyric acid and indole-3-propionic acid were competitive inhibitors of IAGlc synthase, D-gluconic acid lactone, an inhibitor of β-glucosidase activity, potentiated the enzyme activity at the optimal concentration of 0.3mM. Moreover, we demonstrated that the 1-O-IA-glucose synthesized by IAGlc synthase is the substrate for IAA labeling of glycoproteins from pea seeds indicating a possible role of this enzyme in the covalent modification of a class of proteins by a plant hormone. PMID:26057226

  13. MeCP2 post-translational modifications: a mechanism to control its involvement in synaptic plasticity and homeostasis?

    PubMed Central

    Bellini, Elisa; Pavesi, Giulio; Barbiero, Isabella; Bergo, Anna; Chandola, Chetan; Nawaz, Mohammad S.; Rusconi, Laura; Stefanelli, Gilda; Strollo, Marta; Valente, Maria M.; Kilstrup-Nielsen, Charlotte; Landsberger, Nicoletta

    2014-01-01

    Although Rett syndrome (RTT) represents one of the most frequent forms of severe intellectual disability in females worldwide, we still have an inadequate knowledge of the many roles played by MeCP2 (whose mutations are responsible for most cases of RTT) and their relevance for RTT pathobiology. Several studies support a role of MeCP2 in the regulation of synaptic plasticity and homeostasis. At the molecular level, MeCP2 is described as a repressor capable of inhibiting gene transcription through chromatin compaction. Indeed, it interacts with several chromatin remodeling factors, such as HDAC-containing complexes and ATRX. Other studies have inferred that MeCP2 functions also as an activator; a role in regulating mRNA splicing and in modulating protein synthesis has also been proposed. Further, MeCP2 avidly binds both 5-methyl- and 5-hydroxymethyl-cytosine. Recent evidence suggests that it is the highly disorganized structure of MeCP2, together with its post-translational modifications (PTMs) that generate and regulate this functional versatility. Indeed, several reports have demonstrated that differential phosphorylation of MeCP2 is a key mechanism by which the methyl binding protein modulates its affinity for its partners, gene expression and cellular adaptations to stimuli and neuronal plasticity. As logic consequence, generation of phospho-defective Mecp2 knock-in mice has permitted associating alterations in neuronal morphology, circuit formation, and mouse behavioral phenotypes with specific phosphorylation events. MeCP2 undergoes various other PTMs, including acetylation, ubiquitination and sumoylation, whose functional roles remain largely unexplored. These results, together with the genome-wide distribution of MeCP2 and its capability to substitute histone H1, recall the complex regulation of histones and suggest the relevance of quickly gaining a deeper comprehension of MeCP2 PTMs, the respective writers and readers and the consequent functional outcomes

  14. Modifications to the HIPAA Privacy, Security, Enforcement, and Breach Notification rules under the Health Information Technology for Economic and Clinical Health Act and the Genetic Information Nondiscrimination Act; other modifications to the HIPAA rules.

    PubMed

    2013-01-25

    The Department of Health and Human Services (HHS or ``the Department'') is issuing this final rule to: Modify the Health Insurance Portability and Accountability Act (HIPAA) Privacy, Security, and Enforcement Rules to implement statutory amendments under the Health Information Technology for Economic and Clinical Health Act (``the HITECH Act'' or ``the Act'') to strengthen the privacy and security protection for individuals' health information; modify the rule for Breach Notification for Unsecured Protected Health Information (Breach Notification Rule) under the HITECH Act to address public comment received on the interim final rule; modify the HIPAA Privacy Rule to strengthen the privacy protections for genetic information by implementing section 105 of Title I of the Genetic Information Nondiscrimination Act of 2008 (GINA); and make certain other modifications to the HIPAA Privacy, Security, Breach Notification, and Enforcement Rules (the HIPAA Rules) to improve their workability and effectiveness and to increase flexibility for and decrease burden on the regulated entities.

  15. Genetic variation in genes involved in hormones, inflammation and energetic factors and breast cancer risk in an admixed population.

    PubMed

    Slattery, Martha L; John, Esther M; Torres-Mejia, Gabriela; Lundgreen, Abbie; Herrick, Jennifer S; Baumgartner, Kathy B; Hines, Lisa M; Stern, Mariana C; Wolff, Roger K

    2012-08-01

    Breast cancer incidence rates are characterized by unique racial and ethnic differences. Native American ancestry has been associated with reduced breast cancer risk. We explore the biological basis of disparities in breast cancer risk in Hispanic and non-Hispanic white women by evaluating genetic variation in genes involved in inflammation, insulin and energy homeostasis in conjunction with genetic ancestry. Hispanic (2111 cases, 2597 controls) and non-Hispanic white (1481 cases, 1586 controls) women enrolled in the 4-Corner's Breast Cancer Study, the Mexico Breast Cancer Study and the San Francisco Bay Area Breast Cancer Study were included. Genetic admixture was determined from 104 ancestral informative markers that discriminate between European and Native American ancestry. Twenty-one genes in the CHIEF candidate pathway were evaluated. Higher Native American ancestry was associated with reduced risk of breast cancer (odds ratio = 0.79, 95% confidence interval 0.65, 0.95) but was limited to postmenopausal women (odds ratio = 0.66, 95% confidence interval 0.52, 0.85). After adjusting for genetic ancestry and multiple comparisons, four genes were significantly associated with breast cancer risk, NFκB1, NFκB1A, PTEN and STK11. Within admixture strata, breast cancer risk among women with low Native American ancestry was associated with IkBKB, NFκB1, PTEN and RPS6KA2, whereas among women with high Native American ancestry, breast cancer risk was associated with IkBKB, mTOR, PDK2, PRKAA1, RPS6KA2 and TSC1. Higher Native American ancestry was associated with reduced breast cancer risk. Breast cancer risk differed by genetic ancestry along with genetic variation in genes involved in inflammation, insulin, and energy homeostasis. PMID:22562547

  16. Genetic Adaptation to Climate in White Spruce Involves Small to Moderate Allele Frequency Shifts in Functionally Diverse Genes

    PubMed Central

    Hornoy, Benjamin; Pavy, Nathalie; Gérardi, Sébastien; Beaulieu, Jean; Bousquet, Jean

    2015-01-01

    Understanding the genetic basis of adaptation to climate is of paramount importance for preserving and managing genetic diversity in plants in a context of climate change. Yet, this objective has been addressed mainly in short-lived model species. Thus, expanding knowledge to nonmodel species with contrasting life histories, such as forest trees, appears necessary. To uncover the genetic basis of adaptation to climate in the widely distributed boreal conifer white spruce (Picea glauca), an environmental association study was conducted using 11,085 single nucleotide polymorphisms representing 7,819 genes, that is, approximately a quarter of the transcriptome. Linear and quadratic regressions controlling for isolation-by-distance, and the Random Forest algorithm, identified several dozen genes putatively under selection, among which 43 showed strongest signals along temperature and precipitation gradients. Most of them were related to temperature. Small to moderate shifts in allele frequencies were observed. Genes involved encompassed a wide variety of functions and processes, some of them being likely important for plant survival under biotic and abiotic environmental stresses according to expression data. Literature mining and sequence comparison also highlighted conserved sequences and functions with angiosperm homologs. Our results are consistent with theoretical predictions that local adaptation involves genes with small frequency shifts when selection is recent and gene flow among populations is high. Accordingly, genetic adaptation to climate in P. glauca appears to be complex, involving many independent and interacting gene functions, biochemical pathways, and processes. From an applied perspective, these results shall lead to specific functional/association studies in conifers and to the development of markers useful for the conservation of genetic resources. PMID:26560341

  17. The genetic diversity of triticale genotypes involved in Polish breeding programs.

    PubMed

    Niedziela, Agnieszka; Orłowska, Renata; Machczyńska, Joanna; Bednarek, Piotr T

    2016-01-01

    Genetic diversity analysis of triticale populations is useful for breeding programs, as it helps to select appropriate genetic material for classifying the parental lines, heterotic groups and predicting hybrid performance. In our study 232 breeding forms were analyzed using diversity arrays technology markers. Principal coordinate analysis followed by model-based Bayesian analysis of population structure revealed the presence of weak data structuring with three groups of data. In the first group, 17 spring and 17 winter forms were clustered. The second and the third groups were represented by 101 and 26 winter forms, respectively. Polymorphic information content values, as well as Shannon's Information Index, were higher for the first (0.319) and second (0.309) than for third (0.234) group. AMOVA analysis demonstrated a higher level of within variation (86 %) than among populations (14 %). This study provides the basic information on the presence of structure within a genetic pool of triticale breeding forms. PMID:27066368

  18. Modification of hydrological properties in a fine textured soil following field application of pelletized biochar: investigation of the mechanism involved.

    NASA Astrophysics Data System (ADS)

    Costanza Andrenelli, Maria; Mocali, Stefano; Pellegrini, Sergio; Vignozzi, Nadia

    2016-04-01

    The application of pelletized biochar is seldom employed in field, and its effect on soil hydrological behaviour scarcely investigated. Biochar is usually added in powdered or granular form to improve the homogeneity of distribution, meanwhile favouring its interaction with soil matrix. In this study we evaluated the possibility of applying pelletized biochar as soil conditioner to enhance, during a single cropping season, the hydrological behaviour of a silty clay loam soil prone to structure degradation. For that purpose, the water retention curves (WRCs) were determined on undisturbed soil samples (0-15 cm) three months after the addition, at the rate of 14 Mg ha-1, of two differently pyrolyzed biochars (B1 and B2). Starting from the WRCs the pore size distribution was determined. The gravimetric water content at both field capacity (-10 kPa) and wilting point (-1,500 kPa) was also measured on biochar samples to assess their available water capacity (AWC). In both the treatments, soil bulk density (BD) was significantly lower compared to control, apparently as direct consequence of the addition of low density pellets. Actually, excluding the intrinsic biochar porosity from soil bulk density calculation, BD values of the treated soils remain lower of around 10% over control. Such findings suggest that a modification of soil structural characteristics might have been induced by pellet addition. Data of the WRCs indicate a significant increase of transmission (500-50 micron), storage (50-0.5 micron) and AWC pores (30-0.2 micron) in the amended soils. The two biochars affected the AWC by direct pore contribution, but the extent of such effect was related to the biochar type: the tested pelletized biomass seems to have positive effects provided that the pyrolysis temperature does not exceed 800°C, as in the case of B1. The overall hydrological improvement might be correlated to both the inherent biochar retention capacity and a merely mechanical process of

  19. Modification of hydrological properties in a fine textured soil following field application of pelletized biochar: investigation of the mechanism involved.

    NASA Astrophysics Data System (ADS)

    Costanza Andrenelli, Maria; Mocali, Stefano; Pellegrini, Sergio; Vignozzi, Nadia

    2016-04-01

    The application of pelletized biochar is seldom employed in field, and its effect on soil hydrological behaviour scarcely investigated. Biochar is usually added in powdered or granular form to improve the homogeneity of distribution, meanwhile favouring its interaction with soil matrix. In this study we evaluated the possibility of applying pelletized biochar as soil conditioner to enhance, during a single cropping season, the hydrological behaviour of a silty clay loam soil prone to structure degradation. For that purpose, the water retention curves (WRCs) were determined on undisturbed soil samples (0-15 cm) three months after the addition, at the rate of 14 Mg ha‑1, of two differently pyrolyzed biochars (B1 and B2). Starting from the WRCs the pore size distribution was determined. The gravimetric water content at both field capacity (-10 kPa) and wilting point (-1,500 kPa) was also measured on biochar samples to assess their available water capacity (AWC). In both the treatments, soil bulk density (BD) was significantly lower compared to control, apparently as direct consequence of the addition of low density pellets. Actually, excluding the intrinsic biochar porosity from soil bulk density calculation, BD values of the treated soils remain lower of around 10% over control. Such findings suggest that a modification of soil structural characteristics might have been induced by pellet addition. Data of the WRCs indicate a significant increase of transmission (500-50 micron), storage (50-0.5 micron) and AWC pores (30-0.2 micron) in the amended soils. The two biochars affected the AWC by direct pore contribution, but the extent of such effect was related to the biochar type: the tested pelletized biomass seems to have positive effects provided that the pyrolysis temperature does not exceed 800°C, as in the case of B1. The overall hydrological improvement might be correlated to both the inherent biochar retention capacity and a merely mechanical process of

  20. Conserved genetic basis of a quantitative plumage trait involved in mate choice.

    PubMed

    Mundy, Nicholas I; Badcock, Nichola S; Hart, Tom; Scribner, Kim; Janssen, Kirstin; Nadeau, Nicola J

    2004-03-19

    A key question in evolutionary genetics is whether shared genetic mechanisms underlie the independent evolution of similar phenotypes across phylogenetically divergent lineages. Here we show that in two classic examples of melanic plumage polymorphisms in birds, lesser snow geese (Anser c. caerulescens) and arctic skuas (Stercorarius parasiticus), melanism is perfectly associated with variation in the melanocortin-1 receptor (MC1R) gene. In both species, the degree of melanism correlates with the number of copies of variant MC1R alleles. Phylogenetic reconstructions of variant MC1R alleles in geese and skuas show that melanism is a derived trait that evolved in the Pleistocene. PMID:15031505

  1. Shared Genetic Factors Involved in Celiac Disease, Type 2 Diabetes and Anorexia Nervosa Suggest Common Molecular Pathways for Chronic Diseases

    PubMed Central

    Mostowy, Joanna; Montén, Caroline; Gudjonsdottir, Audur H.; Arnell, Henrik; Browaldh, Lars; Nilsson, Staffan; Agardh, Daniel

    2016-01-01

    Background and Objectives Genome-wide association studies (GWAS) have identified several genetic regions involved in immune-regulatory mechanisms to be associated with celiac disease. Previous GWAS also revealed an over-representation of genes involved in type 2 diabetes and anorexia nervosa associated with celiac disease, suggesting involvement of common metabolic pathways for development of these chronic diseases. The aim of this study was to extend these previous analyses to study the gene expression in the gut from children with active celiac disease. Material and Methods Thirty six target genes involved in type 2 diabetes and four genes associated with anorexia nervosa were investigated for gene expression in small intestinal biopsies from 144 children with celiac disease at median (range) age of 7.4 years (1.6–17.8) and from 154 disease controls at a median (range) age 11.4.years (1.4–18.3). Results A total of eleven of genes were differently expressed in celiac patients compared with disease controls of which CD36, CD38, FOXP1, SELL, PPARA, PPARG, AGT previously associated with type 2 diabetes and AKAP6, NTNG1 with anorexia nervosa remained significant after correction for multiple testing. Conclusion Shared genetic factors involved in celiac disease, type 2 diabetes and anorexia nervosa suggest common underlying molecular pathways for these diseases. PMID:27483138

  2. High acceptance of an early dyslexia screening test involving genetic analyses in Germany.

    PubMed

    Wilcke, Arndt; Müller, Bent; Schaadt, Gesa; Kirsten, Holger; Boltze, Johannes

    2016-02-01

    Dyslexia is a developmental disorder characterized by severe problems in the acquisition of reading and writing skills. It has a strong neurobiological basis. Genetic influence is estimated at 50-70%. One of the central problems with dyslexia is its late diagnosis, normally not before the end of the 2nd grade, resulting in the loss of several years for early therapy. Currently, research is focusing on the development of early tests for dyslexia, which may be based on EEG and genetics. Our aim was to determine the acceptance of such a future test among parents. We conducted a representative survey in Germany with 1000 parents of children aged 3-7 years, with and without experience of dyslexia. 88.7% of the parents supported the introduction of an early test for dyslexia based on EEG and genetics; 82.8% would have their own children tested, and 57.9% were willing to pay for the test if health insurance did not cover the costs. Test acceptance was significantly higher if parents had prior experience with dyslexia. The perceived benefits of such a test were early recognition and remediation and, preventing deficits. Concerns regarded the precision of the test, its potentially stigmatizing effect and its costs. The high overall support for the test leads to the conclusion that parents would accept a test for dyslexia based on EEG and genetics.

  3. Identification of Candida glabrata genes involved in pH modulation and modification of the phagosomal environment in macrophages.

    PubMed

    Kasper, Lydia; Seider, Katja; Gerwien, Franziska; Allert, Stefanie; Brunke, Sascha; Schwarzmüller, Tobias; Ames, Lauren; Zubiria-Barrera, Cristina; Mansour, Michael K; Becken, Ulrike; Barz, Dagmar; Vyas, Jatin M; Reiling, Norbert; Haas, Albert; Haynes, Ken; Kuchler, Karl; Hube, Bernhard

    2014-01-01

    Candida glabrata currently ranks as the second most frequent cause of invasive candidiasis. Our previous work has shown that C. glabrata is adapted to intracellular survival in macrophages and replicates within non-acidified late endosomal-stage phagosomes. In contrast, heat killed yeasts are found in acidified matured phagosomes. In the present study, we aimed at elucidating the processes leading to inhibition of phagosome acidification and maturation. We show that phagosomes containing viable C. glabrata cells do not fuse with pre-labeled lysosomes and possess low phagosomal hydrolase activity. Inhibition of acidification occurs independent of macrophage type (human/murine), differentiation (M1-/M2-type) or activation status (vitamin D3 stimulation). We observed no differential activation of macrophage MAPK or NFκB signaling cascades downstream of pattern recognition receptors after internalization of viable compared to heat killed yeasts, but Syk activation decayed faster in macrophages containing viable yeasts. Thus, delivery of viable yeasts to non-matured phagosomes is likely not triggered by initial recognition events via MAPK or NFκB signaling, but Syk activation may be involved. Although V-ATPase is abundant in C. glabrata phagosomes, the influence of this proton pump on intracellular survival is low since blocking V-ATPase activity with bafilomycin A1 has no influence on fungal viability. Active pH modulation is one possible fungal strategy to change phagosome pH. In fact, C. glabrata is able to alkalinize its extracellular environment, when growing on amino acids as the sole carbon source in vitro. By screening a C. glabrata mutant library we identified genes important for environmental alkalinization that were further tested for their impact on phagosome pH. We found that the lack of fungal mannosyltransferases resulted in severely reduced alkalinization in vitro and in the delivery of C. glabrata to acidified phagosomes. Therefore, protein

  4. Rice (Oryza sativa) Laccases Involved in Modification and Detoxification of Herbicides Atrazine and Isoproturon Residues in Plants.

    PubMed

    Huang, Meng Tian; Lu, Yi Chen; Zhang, Shuang; Luo, Fang; Yang, Hong

    2016-08-24

    Atrazine (ATR) and isoproturon (IPU) as herbicides have become serious environmental contaminants due to their overuse in crop production. Although ATR and IPU in soils are easily absorbed by many crops, the mechanisms for their degradation or detoxification in plants are poorly understood. This study identified a group of novel genes encoding laccases (EC 1.10.3.2) that are possibly involved in catabolism or detoxification of ATR and IPU residues in rice. Transcriptome profiling shows at least 22 differentially expressed laccase genes in ATR/IPU-exposed rice. Some of the laccase genes were validated by RT-PCR analysis. The biochemical properties of the laccases were analyzed, and their activities in rice were induced under ATR/IPU exposure. To investigate the roles of laccases in degrading or detoxifying ATR/IPU in rice, transgenic yeast cells (Pichia pastoris X-33) expressing two rice laccase genes (LOC_Os01g63180 and LOC_Os12g15680) were generated. Both transformants were found to accumulate less ATR/IPU compared to the control. The ATR/IPU-degraded products in the transformed yeast cells using UPLC-TOF-MS/MS were further characterized. Two metabolites, hydroxy-dehydrogenated atrazine (HDHA) and 2-OH-isopropyl-IPU, catalyzed by laccases were detected in the eukaryotic cells. These results indicate that the laccase-coding genes identified here could confer degradation or detoxification of the herbicides and suggest that the laccases could be one of the important enzymatic pathways responsible for ATR/IPU degradation/detoxification in rice. PMID:27499219

  5. Ubiquitin modifications

    PubMed Central

    Swatek, Kirby N; Komander, David

    2016-01-01

    Protein ubiquitination is a dynamic multifaceted post-translational modification involved in nearly all aspects of eukaryotic biology. Once attached to a substrate, the 76-amino acid protein ubiquitin is subjected to further modifications, creating a multitude of distinct signals with distinct cellular outcomes, referred to as the 'ubiquitin code'. Ubiquitin can be ubiquitinated on seven lysine (Lys) residues or on the N-terminus, leading to polyubiquitin chains that can encompass complex topologies. Alternatively or in addition, ubiquitin Lys residues can be modified by ubiquitin-like molecules (such as SUMO or NEDD8). Finally, ubiquitin can also be acetylated on Lys, or phosphorylated on Ser, Thr or Tyr residues, and each modification has the potential to dramatically alter the signaling outcome. While the number of distinctly modified ubiquitin species in cells is mind-boggling, much progress has been made to characterize the roles of distinct ubiquitin modifications, and many enzymes and receptors have been identified that create, recognize or remove these ubiquitin modifications. We here provide an overview of the various ubiquitin modifications present in cells, and highlight recent progress on ubiquitin chain biology. We then discuss the recent findings in the field of ubiquitin acetylation and phosphorylation, with a focus on Ser65-phosphorylation and its role in mitophagy and Parkin activation. PMID:27012465

  6. Multiple Genetic Modifiers of Bilirubin Metabolism Involvement in Significant Neonatal Hyperbilirubinemia in Patients of Chinese Descent

    PubMed Central

    Yang, Hui; Zheng, Lei; Lin, Min; Zheng, Xiang-bin; Lin, Fen

    2015-01-01

    The potential for genetic variation to modulate neonatal hyperbilirubinemia risk is increasingly being recognized. A case-control study was designed to assess comprehensive contributions of the multiple genetic modifiers of bilirubin metabolism on significant neonatal hyperbilirubinemia in Chinese descendents. Eleven common mutations and polymorphisms across five bilirubin metabolism genes, namely those encoding UGT1A1, HMOX1, BLVRA, SLCO1B1 and SLCO1B3, were determined using the high resolution melt (HRM) assay or PCR-capillary electrophoresis analysis. A total of 129 hyperbilirubinemic infants and 108 control subjects were evaluated. Breastfeeding and the presence of the minor A allele of rs4148323 (UGTA*6) were correlated with an increased risk of hyperbilirubinemia (OR=2.17, P=0.02 for breastfeeding; OR=9.776, P=0.000 for UGTA*6 homozygote; OR=3.151, P=0.000 for UGTA*6 heterozygote); whereas, increasing gestational age and the presence of –TA7 repeat variant of UGT1A1 decreased the risk (OR=0.721, P=0.003 for gestational age; OR=0.313, P=0.002 for heterozygote TA6/TA7). In addition, the SLCO1B1 and SLCO1B3 polymorphisms also contributed to an increased risk of hyperbilirubinemia. This detailed analysis revealed the impact of multiple genetic modifiers on neonatal hyperbilirubinemia. This may support the use of genetic tests for clinical risk assessment. Furthermore, the established HRM assay can serve as an effective method for large-scale investigation. PMID:26146841

  7. Evolutionary genetics as a tool to target genes involved in phenotypes of medical relevance

    PubMed Central

    Heyer, Evelyne; Quintana-Murci, Lluis

    2009-01-01

    There is an increasing interest in detecting genes, or genomic regions, that have been targeted by natural selection. Indeed, the evolutionary approach for inferring the action of natural selection in the human genome represents a powerful tool for predicting regions of the genome potentially associated with disease and of interest in epidemiological genetic studies. Here, we review several examples going from candidate gene studies associated with specific phenotypes, including nutrition, infectious disease and climate adaptation, to whole genome scans for natural selection. All these studies illustrate the power of the evolutionary approach in identifying regions of the genome having played a major role in human survival and adaptation. PMID:25567848

  8. Genetic Factors Involved in Fumonisin Accumulation in Maize Kernels and Their Implications in Maize Agronomic Management and Breeding.

    PubMed

    Santiago, Rogelio; Cao, Ana; Butrón, Ana

    2015-08-20

    Contamination of maize with fumonisins depends on the environmental conditions; the maize resistance to contamination and the interaction between both factors. Although the effect of environmental factors is a determinant for establishing the risk of kernel contamination in a region, there is sufficient genetic variability among maize to develop resistance to fumonisin contamination and to breed varieties with contamination at safe levels. In addition, ascertaining which environmental factors are the most important in a region will allow the implementation of risk monitoring programs and suitable cultural practices to reduce the impact of such environmental variables. The current paper reviews all works done to address the influence of environmental variables on fumonisin accumulation, the genetics of maize resistance to fumonisin accumulation, and the search for the biochemical and/or structural mechanisms of the maize plant that could be involved in resistance to fumonisin contamination. We also explore the outcomes of breeding programs and risk monitoring of undertaken projects.

  9. Genetic Factors Involved in Fumonisin Accumulation in Maize Kernels and Their Implications in Maize Agronomic Management and Breeding.

    PubMed

    Santiago, Rogelio; Cao, Ana; Butrón, Ana

    2015-08-01

    Contamination of maize with fumonisins depends on the environmental conditions; the maize resistance to contamination and the interaction between both factors. Although the effect of environmental factors is a determinant for establishing the risk of kernel contamination in a region, there is sufficient genetic variability among maize to develop resistance to fumonisin contamination and to breed varieties with contamination at safe levels. In addition, ascertaining which environmental factors are the most important in a region will allow the implementation of risk monitoring programs and suitable cultural practices to reduce the impact of such environmental variables. The current paper reviews all works done to address the influence of environmental variables on fumonisin accumulation, the genetics of maize resistance to fumonisin accumulation, and the search for the biochemical and/or structural mechanisms of the maize plant that could be involved in resistance to fumonisin contamination. We also explore the outcomes of breeding programs and risk monitoring of undertaken projects. PMID:26308050

  10. Genetic Factors Involved in Fumonisin Accumulation in Maize Kernels and Their Implications in Maize Agronomic Management and Breeding

    PubMed Central

    Santiago, Rogelio; Cao, Ana; Butrón, Ana

    2015-01-01

    Contamination of maize with fumonisins depends on the environmental conditions; the maize resistance to contamination and the interaction between both factors. Although the effect of environmental factors is a determinant for establishing the risk of kernel contamination in a region, there is sufficient genetic variability among maize to develop resistance to fumonisin contamination and to breed varieties with contamination at safe levels. In addition, ascertaining which environmental factors are the most important in a region will allow the implementation of risk monitoring programs and suitable cultural practices to reduce the impact of such environmental variables. The current paper reviews all works done to address the influence of environmental variables on fumonisin accumulation, the genetics of maize resistance to fumonisin accumulation, and the search for the biochemical and/or structural mechanisms of the maize plant that could be involved in resistance to fumonisin contamination. We also explore the outcomes of breeding programs and risk monitoring of undertaken projects. PMID:26308050

  11. Chromatin remodeling gene EZH2 involved in the genetic etiology of autism in Chinese Han population.

    PubMed

    Li, Jun; You, Yang; Yue, Weihua; Yu, Hao; Lu, Tianlan; Wu, Zhiliu; Jia, Meixiang; Ruan, Yanyan; Liu, Jing; Zhang, Dai; Wang, Lifang

    2016-01-01

    Autism spectrum disorder (ASD) is a group of severe neurodevelopmental disorders. Epigenetic factors play a critical role in the etiology of ASD. Enhancer of zest homolog 2 (EZH2), which encodes a histone methyltransferase, plays an important role in the process of chromatin remodeling during neurodevelopment. Further, EZH2 is located in chromosome 7q35-36, which is one of the linkage regions for autism. However, the genetic relationship between autism and EZH2 remains unclear. To investigate the association between EZH2 and autism in Chinese Han population, we performed a family-based association study between autism and three tagged single nucleotide polymorphisms (SNPs) that covered 95.4% of the whole region of EZH2. In the discovery cohort of 239 trios, two SNPs (rs740949 and rs6464926) showed a significant association with autism. To decrease false positive results, we expanded the sample size to 427 trios. A SNP (rs6464926) was significantly associated with autism even after Bonferroni correction (p=0.008). Haplotype G-T (rs740949 and rs6464926) was a risk factor for autism (Z=2.655, p=0.008, Global p=0.024). In silico function prediction for SNPs indicated that these two SNPs might be regulatory SNPs. Expression pattern of EZH2 showed that it is highly expressed in human embryonic brains. In conclusion, our findings demonstrate that EZH2 might contribute to the genetic etiology of autism in Chinese Han population.

  12. A Mosaic Genetic Screen for Genes Involved in the Early Steps of Drosophila Oogenesis

    PubMed Central

    Jagut, Marlène; Mihaila-Bodart, Ludivine; Molla-Herman, Anahi; Alin, Marie-Françoise; Lepesant, Jean-Antoine; Huynh, Jean-René

    2013-01-01

    The first hours of Drosophila embryogenesis rely exclusively on maternal information stored within the egg during oogenesis. The formation of the egg chamber is thus a crucial step for the development of the future adult. It has emerged that many key developmental decisions are made during the very first stages of oogenesis. We performed a clonal genetic screen on the left arm of chromosome 2 for mutations affecting early oogenesis. During the first round of screening, we scored for defects in egg chambers morphology as an easy read-out of early abnormalities. In a second round of screening, we analyzed the localization of centrosomes and Orb protein within the oocyte, the position of the oocyte within the egg chamber, and the progression through meiosis. We have generated a collection of 71 EMS-induced mutants that affect oocyte determination, polarization, or localization. We also recovered mutants affecting the number of germline cyst divisions or the differentiation of follicle cells. Here, we describe the analysis of nine complementation groups and eight single alleles. We mapped several mutations and identified alleles of Bicaudal-D, lethal(2) giant larvae, kuzbanian, GDP-mannose 4,6-dehydratase, tho2, and eiF4A. We further report the molecular identification of two alleles of the Drosophila homolog of Che-1/AATF and demonstrate its antiapoptotic activity in vivo. This collection of mutants will be useful to investigate further the early steps of Drosophila oogenesis at a genetic level. PMID:23450845

  13. Identification of Genetic Loci Involved in Diabetes using a Rat Model of Depression

    PubMed Central

    Woods, Leah C Solberg; Ahmadiyeh, Nasim; Baum, Amber; Shimomura, Kazuhiro; Li, Qian; Steiner, Donald F; Turek, Fred W; Takahashi, Joseph S; Churchill, Gary A; Redei, Eva E

    2009-01-01

    While diabetic patients often present with comorbid depression, the underlying mechanisms linking diabetes and depression are unknown. The Wistar Kyoto (WKY) rat is a well-known animal model of depression and stress hyper-reactivity. In addition, the WKY rat is glucose intolerant and likely harbors diabetes susceptibility alleles. We conducted a quantitative trait loci (QTL) analysis in the segregating F2 population of a WKY × Fischer 344 (F344) inter-cross. We have previously published QTL analyses for depressive behavior and hypothalamic-pituitary-adrenal (HPA) activity in this cross. In the current study, we report results from the QTL analysis for multiple metabolic phenotypes, including fasting glucose, post-restraint stress glucose, post-prandial glucose and insulin, and body weight. We identified multiple QTLs for each trait and many of the QTLs overlap with those previously identified using inbred models of type 2 diabetes (T2D). Significant correlations were found between metabolic traits and HPA axis measures and several metabolic loci overlap with loci previously identified for HPA activity in this F2 intercross, suggesting the genetic mechanisms underlying these traits may be similar. These results indicate that WKY rats harbor diabetes susceptibility alleles and suggest that this strain may be useful for dissecting the underlying genetic mechanisms linking diabetes, HPA activity and depression. PMID:19697080

  14. Genetic susceptibility to Chagas disease cardiomyopathy: involvement of several genes of the innate immunity and chemokine-dependent migration pathways

    PubMed Central

    2013-01-01

    Background Chagas disease, caused by the protozoan Trypanosoma cruzi is endemic in Latin America. Thirty percent of infected individuals develop chronic Chagas cardiomyopathy (CCC), an inflammatory dilated cardiomyopathy that is, by far, the most important clinical consequence of T. cruzi infection. The others remain asymptomatic (ASY). A possible genetic component to disease progression was suggested by familial aggregation of cases and the association of markers of innate and adaptive immunity genes with CCC development. Migration of Th1-type T cells play a major role in myocardial damage. Methods Our genetic analysis focused on CCR5, CCL2 and MAL/TIRAP genes. We used the Tag SNPs based approach, defined to catch all the genetic information from each gene. The study was conducted on a large Brazilian population including 315 CCC cases and 118 ASY subjects. Results The CCL2rs2530797A/A and TIRAPrs8177376A/A were associated to an increase susceptibility whereas the CCR5rs3176763C/C genotype is associated to protection to CCC. These associations were confirmed when we restricted the analysis to severe CCC, characterized by a left ventricular ejection fraction under 40%. Conclusions Our data show that polymorphisms affecting key molecules involved in several immune parameters (innate immunity signal transduction and T cell/monocyte migration) play a role in genetic susceptibility to CCC development. This also points out to the multigenic character of CCC, each polymorphism imparting a small contribution. The identification of genetic markers for CCC will provide information for pathogenesis as well as therapeutic targets. PMID:24330528

  15. Human amniotic fluid stem cells as a model for functional studies of genes involved in human genetic diseases or oncogenesis.

    PubMed

    Rosner, Margit; Dolznig, Helmut; Schipany, Katharina; Mikula, Mario; Brandau, Oliver; Hengstschläger, Markus

    2011-09-01

    Besides their putative usage for therapies, stem cells are a promising tool for functional studies of genes involved in human genetic diseases or oncogenesis. For this purpose induced pluripotent stem (iPS) cells can be derived from patients harbouring specific mutations. In contrast to adult stem cells, iPS cells are pluripotent and can efficiently be grown in culture. However, iPS cells are modulated due to the ectopic induction of pluripotency, harbour other somatic mutations accumulated during the life span of the source cells, exhibit only imperfectly cleared epigenetic memory of the source cell, and are often genomically instable. In addition, iPS cells from patients only allow the investigation of mutations, which are not prenatally lethal. Embryonic stem (ES) cells have a high proliferation and differentiation potential, but raise ethical issues. Human embryos, which are not transferred in the course of in vitro fertilization, because of preimplantation genetic diagnosis of a genetic defect, are still rarely donated for the establishment of ES cell lines. In addition, their usage for studies on gene functions for oncogenesis is hampered by the fact the ES cells are already tumorigenic per se. In 2003 amniotic fluid stem (AFS) cells have been discovered, which meanwhile have been demonstrated to harbour the potential to differentiate into cells of all three germ layers. Monoclonal human AFS cell lines derived from amniocenteses have a high proliferative potential, are genomically stable and are not associated with ethical controversies. Worldwide amniocenteses are performed for routine human genetic diagnosis. We here discuss how generation and banking of monoclonal human AFS cell lines with specific chromosomal aberrations or monogenic disease mutations would allow to study the functional consequences of disease causing mutations. In addition, recently a protocol for efficient and highly reproducible siRNA-mediated long-term knockdown of endogenous gene

  16. Rapid sensitive analysis of IDH1 mutation in lower-grade gliomas by automated genetic typing involving a quenching probe.

    PubMed

    Kurimoto, Michihiro; Suzuki, Hiromichi; Aoki, Kosuke; Ohka, Fumiharu; Kondo, Goro; Motomura, Kazuya; Iijima, Kentaro; Yamamichi, Akane; Ranjit, Melissa; Wakabayashi, Toshihiko; Kimura, Shinya; Natsume, Atsushi

    2016-01-01

    The authors recently found that 80% of lower-grade gliomas (LGGs) harbored a mutation in IDH1. Intraoperative detection of the mutated IDH1 helps not only differentiate LGGs from other type of brain tumors, but determine the resection border. In the current study, the authors have applied an automated genetic typing involving a quenching probe to detect the mutated IDH1. If tumor cells with the mutated IDH1 contained 10% or more in the mixture of normal and tumor cells, the device could detect it sensitively. The intraoperative assessment of IDH1 mutation is useful in brain tumor surgeries.

  17. Genetic diversity, dynamics, and activity of Lactobacillus community involved in traditional processing of artisanal Manchego cheese.

    PubMed

    Sánchez, Isabel; Seseña, Susana; Poveda, Justa M; Cabezas, Lourdes; Palop, Llanos

    2006-04-01

    A total of 248 strains of predominant lactobacilli isolated during the manufacture and ripening of artisanal Manchego cheeses obtained from two dairies were obtained and the genetic diversity of 197 investigated using random amplified polymorphic DNA (RAPD-PCR). 51 isolates could not be lysed and were therefore not genotyped. Forty-two distinct RAPD patterns, grouped in six major clusters at a similarity level of 54%, were obtained. Phenotypic characterization of isolates enabled their assignment to the species L. plantarum, L. brevis, L. paracasei subsp. paracasei, L. fermentum, L. pentosus, L. acidophilus and L. curvatus. In samples from both dairies, the species L. plantarum, L. brevis and L. paracasei subsp. paracasei dominated during ripening. Three genotypes showed excellent physiological characteristics and were therefore proposed as adjunct cultures for Manchego cheese manufacture. PMID:16481060

  18. An integrated approach to defining genetic and environmental determinants for major clinical outcomes involving vitamin D.

    PubMed

    Berlanga-Taylor, Antonio J; Knight, Julian C

    2014-06-01

    There is substantial genetic and epidemiological evidence implicating vitamin D in the pathogenesis of many common diseases. A number of studies have sought to define an association for disease with sequence variation in the VDR gene, encoding the ligand-activated nuclear hormone receptor for vitamin D. The results of such studies have been difficult to replicate and are likely to need to account for specific environmental exposures. Here, we review recent work that has begun to study the interactions between VDR gene polymorphisms, vitamin D blood levels, and complex disease susceptibility, notably in the context of major clinical outcomes. We highlight the challenges moving forward in this area and its importance for effective clinical translation of current research.

  19. Integument pattern formation involves genetic and epigenetic controls: feather arrays simulated by digital hormone models

    PubMed Central

    Jiang, Ting-Xin; Widelitz, Randall B.; Shen, Wei-Min; Will, Peter; Wu, Da-Yu; Lin, Chih-Min; Jung, Han-Sung; Chuong, Cheng-Ming

    2015-01-01

    Pattern formation is a fundamental morphogenetic process. Models based on genetic and epigenetic control have been proposed but remain controversial. Here we use feather morphogenesis for further evaluation. Adhesion molecules and/or signaling molecules were first expressed homogenously in feather tracts (restrictive mode, appear earlier) or directly in bud or inter-bud regions (de novo mode, appear later). They either activate or inhibit bud formation, but paradoxically co-localize in the bud. Using feather bud reconstitution, we showed that completely dissociated cells can reform periodic patterns without reference to previous positional codes. The patterning process has the characteristics of being self-organizing, dynamic and plastic. The final pattern is an equilibrium state reached by competition, and the number and size of buds can be altered based on cell number and activator/inhibitor ratio, respectively. We developed a Digital Hormone Model which consists of (1) competent cells without identity that move randomly in a space, (2) extracellular signaling hormones which diffuse by a reaction-diffusion mechanism and activate or inhibit cell adhesion, and (3) cells which respond with topological stochastic actions manifested as changes in cell adhesion. Based on probability, the results are cell clusters arranged in dots or stripes. Thus genetic control provides combinational molecular information which defines the properties of the cells but not the final pattern. Epigenetic control governs interactions among cells and their environment based on physical-chemical rules (such as those described in the Digital Hormone Model). Complex integument patterning is the sum of these two components of control and that is why integument patterns are usually similar but non-identical. These principles may be shared by other pattern formation processes such as barb ridge formation, fingerprints, pigmentation patterning, etc. The Digital Hormone Model can also be applied to

  20. Developmental and genetic analysis of a short leaf mutant, a key resource for plant architecture modification in sorghum

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Modification in plant architecture have been demonstrated as one of the major contributing factors that ushered in the Green Revolution resulting in achieving dramatic increases in grain yield for wheat and rice. For sorghum (Sorghum bicolor L. Moench.), possible alteration in plant architecture is ...

  1. Genetics

    MedlinePlus

    ... Inheritance; Heterozygous; Inheritance patterns; Heredity and disease; Heritable; Genetic markers ... The chromosomes are made up of strands of genetic information called DNA. Each chromosome contains sections of ...

  2. Genetic Characterization of the Klebsiella pneumoniae waa Gene Cluster, Involved in Core Lipopolysaccharide Biosynthesis

    PubMed Central

    Regué, Miguel; Climent, Núria; Abitiu, Nihal; Coderch, Núria; Merino, Susana; Izquierdo, Luis; Altarriba, Maria; Tomás, Juan M.

    2001-01-01

    A recombinant cosmid containing genes involved in Klebsiella pneumoniae C3 core lipopolysaccharide biosynthesis was identified by its ability to confer bacteriocin 28b resistance to Escherichia coli K-12. The recombinant cosmid contains 12 genes, the whole waa gene cluster, flanked by kbl and coaD genes, as was found in E. coli K-12. PCR amplification analysis showed that this cluster is conserved in representative K. pneumoniae strains. Partial nucleotide sequence determination showed that the same genes and gene order are found in K. pneumoniae subsp. ozaenae, for which the core chemical structure is known. Complementation analysis of known waa mutants from E. coli K-12 and/or Salmonella enterica led to the identification of genes involved in biosynthesis of the inner core backbone that are shared by these three members of the Enterobacteriaceae. K. pneumoniae orf10 mutants showed a two-log-fold reduction in a mice virulence assay and a strong decrease in capsule amount. Analysis of a constructed K. pneumoniae waaE deletion mutant suggests that the WaaE protein is involved in the transfer of the branch β-d-Glc to the O-4 position of l-glycero-d-manno-heptose I, a feature shared by K. pneumoniae, Proteus mirabilis, and Yersinia enterocolitica. PMID:11371519

  3. Bcl-xL Genetic Modification Enhanced the Therapeutic Efficacy of Mesenchymal Stem Cell Transplantation in the Treatment of Heart Infarction.

    PubMed

    Xue, Xiaodong; Liu, Yu; Zhang, Jian; Liu, Tao; Yang, Zhonglu; Wang, Huishan

    2015-01-01

    Objectives. Low survival rate of mesenchymal stem cells (MSCs) severely limited the therapeutic efficacy of cell therapy in the treatment of myocardial infarction (MI). Bcl-xL genetic modification might enhance MSC survival after transplantation. Methods. Adult rat bone marrow MSCs were modified with human Bcl-xL gene (hBcl-xL-MSCs) or empty vector (vector-MSCs). MSC apoptosis and paracrine secretions were characterized using flow cytometry, TUNEL, and ELISA in vitro. In vivo, randomized adult rats with MI received myocardial injections of one of the three reagents: hBcl-xL-MSCs, vector-MSCs, or culture medium. Histochemistry, TUNEL, and echocardiography were carried out to evaluate cell engraftment, apoptosis, angiogenesis, scar formation, and cardiac functional recovery. Results. In vitro, cell apoptosis decreased 43%, and vascular endothelial growth factor (VEGF), insulin-like growth factor-1 (IGF-1), and plate-derived growth factor (PDGF) increased 1.5-, 0.7-, and 1.2-fold, respectively, in hBcl-xL-MSCs versus wild type and vector-MSCs. In vivo, cell apoptosis decreased 40% and 26% in hBcl-xL-MSC group versus medium and vector-MSC group, respectively. Similar results were observed in cell engraftment, angiogenesis, scar formation, and cardiac functional recovery. Conclusions. Genetic modification of MSCs with hBcl-xL gene could be an intriguing strategy to improve the therapeutic efficacy of cell therapy in the treatment of heart infarction.

  4. Genetic Variation and Divergence of Genes Involved in Leaf Adaxial-Abaxial Polarity Establishment in Brassica rapa

    PubMed Central

    Liang, Jianli; Liu, Bo; Wu, Jian; Cheng, Feng; Wang, Xiaowu

    2016-01-01

    Alterations in leaf adaxial-abaxial (ad-ab) polarity are one of the main factors that influence leaf curvature. In Chinese cabbage, leaf incurvature is an essential prerequisite to the formation of a leafy head. Identifying ad-ab patterning genes and investigating their genetic variation may facilitate elucidation of the mechanisms underlying leaf incurvature during head formation. Comparative genomic analysis of 45 leaf ad-ab patterning genes in Brassica rapa based on 26 homologs of Arabidopsis thaliana indicated that these genes underwent expansion and were retained after whole genome triplication (WGT). We also assessed the nucleotide diversity and selection footprints of these 45 genes in a collection of 94 Brassica rapa accessions that were composed of heading and non-heading morphotypes. Six of the 45 genes showed significant negative Tajima's D indices and nucleotide diversity reduction in heading accessions compared to those in non-heading accessions, indicating that they underwent purifying selection. Further testing of the BrARF3.1 gene, which was one of the selection signals from a larger collection, confirmed that purifying selection did occur. Our results provide genetic evidence that ad-ab patterning genes are involved in leaf incurvature, which is associated with formation of a leafy head, as well as promote an understanding of the genetic mechanism underlying leafy head formation in Chinese cabbage. PMID:26904064

  5. Genetic Variation and Divergence of Genes Involved in Leaf Adaxial-Abaxial Polarity Establishment in Brassica rapa.

    PubMed

    Liang, Jianli; Liu, Bo; Wu, Jian; Cheng, Feng; Wang, Xiaowu

    2016-01-01

    Alterations in leaf adaxial-abaxial (ad-ab) polarity are one of the main factors that influence leaf curvature. In Chinese cabbage, leaf incurvature is an essential prerequisite to the formation of a leafy head. Identifying ad-ab patterning genes and investigating their genetic variation may facilitate elucidation of the mechanisms underlying leaf incurvature during head formation. Comparative genomic analysis of 45 leaf ad-ab patterning genes in Brassica rapa based on 26 homologs of Arabidopsis thaliana indicated that these genes underwent expansion and were retained after whole genome triplication (WGT). We also assessed the nucleotide diversity and selection footprints of these 45 genes in a collection of 94 Brassica rapa accessions that were composed of heading and non-heading morphotypes. Six of the 45 genes showed significant negative Tajima's D indices and nucleotide diversity reduction in heading accessions compared to those in non-heading accessions, indicating that they underwent purifying selection. Further testing of the BrARF3.1 gene, which was one of the selection signals from a larger collection, confirmed that purifying selection did occur. Our results provide genetic evidence that ad-ab patterning genes are involved in leaf incurvature, which is associated with formation of a leafy head, as well as promote an understanding of the genetic mechanism underlying leafy head formation in Chinese cabbage. PMID:26904064

  6. Genetic Variation and Divergence of Genes Involved in Leaf Adaxial-Abaxial Polarity Establishment in Brassica rapa.

    PubMed

    Liang, Jianli; Liu, Bo; Wu, Jian; Cheng, Feng; Wang, Xiaowu

    2016-01-01

    Alterations in leaf adaxial-abaxial (ad-ab) polarity are one of the main factors that influence leaf curvature. In Chinese cabbage, leaf incurvature is an essential prerequisite to the formation of a leafy head. Identifying ad-ab patterning genes and investigating their genetic variation may facilitate elucidation of the mechanisms underlying leaf incurvature during head formation. Comparative genomic analysis of 45 leaf ad-ab patterning genes in Brassica rapa based on 26 homologs of Arabidopsis thaliana indicated that these genes underwent expansion and were retained after whole genome triplication (WGT). We also assessed the nucleotide diversity and selection footprints of these 45 genes in a collection of 94 Brassica rapa accessions that were composed of heading and non-heading morphotypes. Six of the 45 genes showed significant negative Tajima's D indices and nucleotide diversity reduction in heading accessions compared to those in non-heading accessions, indicating that they underwent purifying selection. Further testing of the BrARF3.1 gene, which was one of the selection signals from a larger collection, confirmed that purifying selection did occur. Our results provide genetic evidence that ad-ab patterning genes are involved in leaf incurvature, which is associated with formation of a leafy head, as well as promote an understanding of the genetic mechanism underlying leafy head formation in Chinese cabbage.

  7. Genetic variants involved in gallstone formation and capsaicin metabolism, and the risk of gallbladder cancer in Chilean women

    PubMed Central

    Báez, Sergio; Tsuchiya, Yasuo; Calvo, Alfonso; Pruyas, Martha; Nakamura, Kazutoshi; Kiyohara, Chikako; Oyama, Mari; Yamamoto, Masaharu

    2010-01-01

    AIM: To determine the effects of genetic variants associated with gallstone formation and capsaicin (a pungent component of chili pepper) metabolism on the risk of gallbladder cancer (GBC). METHODS: A total of 57 patients with GBC, 119 patients with gallstones, and 70 controls were enrolled in this study. DNA was extracted from their blood or paraffin block sample using standard commercial kits. The statuses of the genetic variants were assayed using Taqman® SNP Genotyping Assays or Custom Taqman® SNP Genotyping Assays. RESULTS: The non-ancestral T/T genotype of apolipoprotein B rs693 polymorphism was associated with a decreased risk of GBC (OR: 0.14, 95% CI: 0.03-0.63). The T/T genotype of cholesteryl ester transfer protein (CETP) rs708272 polymorphism was associated with an increased risk of GBC (OR: 5.04, 95% CI: 1.43-17.8). CONCLUSION: Genetic variants involved in gallstone formation such as the apolipoprotein B rs693 and CETP rs708272 polymorphisms may be related to the risk of developing GBC in Chilean women. PMID:20082485

  8. Genetic risk for schizophrenia: convergence on synaptic pathways involved in plasticity.

    PubMed

    Hall, Jeremy; Trent, Simon; Thomas, Kerrie L; O'Donovan, Michael C; Owen, Michael J

    2015-01-01

    Recent large-scale genomic studies have revealed two broad classes of risk alleles for schizophrenia: a polygenic component of risk mediated through multiple common risk variants and rarer more highly penetrant submicroscopic chromosomal deletions and duplications, known as copy number variants. The focus of this review is on the emerging findings from the latter and subsequent exome sequencing data of smaller, deleterious single nucleotide variants and indels. In these studies, schizophrenia patients were found to have enriched de novo mutations in genes belonging to the postsynaptic density at glutamatergic synapses, particularly components of the N-methyl-D-aspartate receptor signaling complex, including the PSD-95 complex, activity-regulated cytoskeleton-associated protein interactors, the fragile X mental retardation protein complex, voltage-gated calcium channels, and genes implicated in actin cytoskeletal dynamics. The convergence of these implicated genes onto a coherent biological pathway at the synapse, with a specific role in plasticity, provides a significant advance in understanding pathogenesis and points to new targets for biological investigation. We consider the implications of these studies in the context of existing genetic data and the potential need to reassess diagnostic boundaries of neuropsychiatric disorders before discussing ways forward for more directed mechanistic studies to develop stratified, novel therapeutic approaches in the future. PMID:25152434

  9. Genetic risk for schizophrenia: convergence on synaptic pathways involved in plasticity.

    PubMed

    Hall, Jeremy; Trent, Simon; Thomas, Kerrie L; O'Donovan, Michael C; Owen, Michael J

    2015-01-01

    Recent large-scale genomic studies have revealed two broad classes of risk alleles for schizophrenia: a polygenic component of risk mediated through multiple common risk variants and rarer more highly penetrant submicroscopic chromosomal deletions and duplications, known as copy number variants. The focus of this review is on the emerging findings from the latter and subsequent exome sequencing data of smaller, deleterious single nucleotide variants and indels. In these studies, schizophrenia patients were found to have enriched de novo mutations in genes belonging to the postsynaptic density at glutamatergic synapses, particularly components of the N-methyl-D-aspartate receptor signaling complex, including the PSD-95 complex, activity-regulated cytoskeleton-associated protein interactors, the fragile X mental retardation protein complex, voltage-gated calcium channels, and genes implicated in actin cytoskeletal dynamics. The convergence of these implicated genes onto a coherent biological pathway at the synapse, with a specific role in plasticity, provides a significant advance in understanding pathogenesis and points to new targets for biological investigation. We consider the implications of these studies in the context of existing genetic data and the potential need to reassess diagnostic boundaries of neuropsychiatric disorders before discussing ways forward for more directed mechanistic studies to develop stratified, novel therapeutic approaches in the future.

  10. Combining gene expression and genetic analyses to identify candidate genes involved in cold responses in pea.

    PubMed

    Legrand, Sylvain; Marque, Gilles; Blassiau, Christelle; Bluteau, Aurélie; Canoy, Anne-Sophie; Fontaine, Véronique; Jaminon, Odile; Bahrman, Nasser; Mautord, Julie; Morin, Julie; Petit, Aurélie; Baranger, Alain; Rivière, Nathalie; Wilmer, Jeroen; Delbreil, Bruno; Lejeune-Hénaut, Isabelle

    2013-09-01

    Cold stress affects plant growth and development. In order to better understand the responses to cold (chilling or freezing tolerance), we used two contrasted pea lines. Following a chilling period, the Champagne line becomes tolerant to frost whereas the Terese line remains sensitive. Four suppression subtractive hybridisation libraries were obtained using mRNAs isolated from pea genotypes Champagne and Terese. Using quantitative polymerase chain reaction (qPCR) performed on 159 genes, 43 and 54 genes were identified as differentially expressed at the initial time point and during the time course study, respectively. Molecular markers were developed from the differentially expressed genes and were genotyped on a population of 164 RILs derived from a cross between Champagne and Terese. We identified 5 candidate genes colocalizing with 3 different frost damage quantitative trait loci (QTL) intervals and a protein quantity locus (PQL) rich region previously reported. This investigation revealed the role of constitutive differences between both genotypes in the cold responses, in particular with genes related to glycine degradation pathway that could confer to Champagne a better frost tolerance. We showed that freezing tolerance involves a decrease of expression of genes related to photosynthesis and the expression of a gene involved in the production of cysteine and methionine that could act as cryoprotectant molecules. Although it remains to be confirmed, this study could also reveal the involvement of the jasmonate pathway in the cold responses, since we observed that two genes related to this pathway were mapped in a frost damage QTL interval and in a PQL rich region interval, respectively.

  11. Genetic diversity of variants involved in drug response and metabolism in Sri Lankan populations: implications for clinical implementation of pharmacogenomics

    PubMed Central

    Chan, Sze Ling; Samaranayake, Nilakshi; Ross, Colin J.D.; Toh, Meng Tiak; Carleton, Bruce; Hayden, Michael R.; Teo, Yik Ying; Dissanayake, Vajira H.W.

    2016-01-01

    Background Interpopulation differences in drug responses are well documented, and in some cases they correspond to differences in the frequency of associated genetic markers. Understanding the diversity of genetic markers associated with drug response across different global populations is essential to infer population rates of drug response or risk for adverse drug reactions, and to guide implementation of pharmacogenomic testing. Sri Lanka is a culturally and linguistically diverse nation, but little is known about the population genetics of the major Sri Lankan ethnic groups. The objective of this study was to investigate the diversity of pharmacogenomic variants in the major Sri Lankan ethnic groups. Methods We examined the allelic diversity of more than 7000 variants in genes involved in drug biotransformation and response in the three major ethnic populations of Sri Lanka (Sinhalese, Sri Lankan Tamils, and Moors), and compared them with other South Asian, South East Asian, and European populations using Wright’s Fixation Index, principal component analysis, and STRUCTURE analysis. Results We observed overall high levels of similarity within the Sri Lankan populations (median FST=0.0034), and between Sri Lankan and other South Asian populations (median FST=0.0064). Notably, we observed substantial differentiation between Sri Lankan and European populations for important pharmacogenomic variants related to warfarin (VKORC1 rs9923231) and clopidogrel (CYP2C19 rs4986893) response. Conclusion These data expand our understanding of the population structure of Sri Lanka, provide a resource for pharmacogenomic research, and have implications for the clinical use of genetic testing of pharmacogenomic variants in these populations. PMID:26444257

  12. Molecular genetics of Erwinia amylovora involved in the development of fire blight.

    PubMed

    Oh, Chang-Sik; Beer, Steven V

    2005-12-15

    The bacterial plant pathogen, Erwinia amylovora, causes the devastating disease known as fire blight in some Rosaceous plants like apple, pear, quince, raspberry and several ornamentals. Knowledge of the factors affecting the development of fire blight has mushroomed in the last quarter century. On the molecular level, genes encoding a Hrp type III secretion system, genes encoding enzymes involved in synthesis of extracellular polysaccharides and genes facilitating the growth of E. amylovora in its host plants have been characterized. The Hrp pathogenicity island, delimited by genes suggesting horizontal gene transfer, is composed of four distinct regions, the hrp/hrc region, the HEE (Hrp effectors and elicitors) region, the HAE (Hrp-associated enzymes) region, and the IT (Island transfer) region. The Hrp pathogenicity island encodes a Hrp type III secretion system (TTSS), which delivers several proteins from bacteria to plant apoplasts or cytoplasm. E. amylovora produces two exopolysaccharides, amylovoran and levan, which cause the characteristic fire blight wilting symptom in host plants. In addition, other genes, and their encoded proteins, have been characterized as virulence factors of E. amylovora that encode enzymes facilitating sorbitol metabolism, proteolytic activity and iron harvesting. This review summarizes our understanding of the genes and gene products of E. amylovora that are involved in the development of the fire blight disease. PMID:16253442

  13. Yeast Genetic Analysis Reveals the Involvement of Chromatin Reassembly Factors in Repressing HIV-1 Basal Transcription

    PubMed Central

    Respaldiza, Iñaki; Rodríguez-Gil, Alfonso; Gómez-Herreros, Fernando; Jimeno-González, Silvia; Jordan, Albert; Chávez, Sebastián

    2009-01-01

    Rebound of HIV viremia after interruption of anti-retroviral therapy is due to the small population of CD4+ T cells that remain latently infected. HIV-1 transcription is the main process controlling post-integration latency. Regulation of HIV-1 transcription takes place at both initiation and elongation levels. Pausing of RNA polymerase II at the 5′ end of HIV-1 transcribed region (5′HIV-TR), which is immediately downstream of the transcription start site, plays an important role in the regulation of viral expression. The activation of HIV-1 transcription correlates with the rearrangement of a positioned nucleosome located at this region. These two facts suggest that the 5′HIV-TR contributes to inhibit basal transcription of those HIV-1 proviruses that remain latently inactive. However, little is known about the cell elements mediating the repressive role of the 5′HIV-TR. We performed a genetic analysis of this phenomenon in Saccharomyces cerevisiae after reconstructing a minimal HIV-1 transcriptional system in this yeast. Unexpectedly, we found that the critical role played by the 5′HIV-TR in maintaining low levels of basal transcription in yeast is mediated by FACT, Spt6, and Chd1, proteins so far associated with chromatin assembly and disassembly during ongoing transcription. We confirmed that this group of factors plays a role in HIV-1 postintegration latency in human cells by depleting the corresponding human orthologs with shRNAs, both in HIV latently infected cell populations and in particular single-integration clones, including a latent clone with a provirus integrated in a highly transcribed gene. Our results indicate that chromatin reassembly factors participate in the establishment of the equilibrium between activation and repression of HIV-1 when it integrates into the human genome, and they open the possibility of considering these factors as therapeutic targets of HIV-1 latency. PMID:19148280

  14. Overexpression, crystallization and preliminary X-ray crystallographic analysis of Pseudomonas aeruginosa MnmE, a GTPase involved in tRNA modification.

    PubMed

    Lee, Hyung Ho; Suh, Se Won

    2010-08-01

    MnmE, an evolutionarily conserved GTPase, is involved in modification of the uridine base (U34) at the wobble position of certain tRNAs. Previous crystal structures of MnmE suggest that it is a dimer with considerable conformational flexibility. To facilitate structural comparisons among MnmE proteins, structural analysis of MnmE from Pseudomonas aeruginosa encoded by the PA5567 gene was initiated. It was overexpressed in Escherichia coli and crystallized at 297 K using a reservoir solution consisting of 100 mM sodium ADA pH 6.5, 12%(w/v) polyethylene glycol 4000, 100 mM lithium sulfate, 2%(v/v) 2-propanol and 2.5 mM dithiothreitol. X-ray diffraction data were collected to 2.69 A resolution. The crystals belonged to the orthorhombic space group C222(1), with unit-cell parameters a=96.74, b=204.66, c=120.90 A. Two monomers were present in the asymmetric unit, resulting in a crystal volume per protein mass (VM) of 2.99 A3 Da(-1) and a solvent content of 58.8%.

  15. Brucella melitensis MucR, an Orthologue of Sinorhizobium meliloti MucR, Is Involved in Resistance to Oxidative, Detergent, and Saline Stresses and Cell Envelope Modifications

    PubMed Central

    Mirabella, A.; Terwagne, M.; Zygmunt, M. S.; Cloeckaert, A.; De Bolle, X.

    2013-01-01

    Brucella spp. and Sinorhizobium meliloti are alphaproteobacteria that share not only an intracellular lifestyle in their respective hosts, but also a crucial requirement for cell envelope components and their timely regulation for a successful infectious cycle. Here, we report the characterization of Brucella melitensis mucR, which encodes a zinc finger transcriptional regulator that has previously been shown to be involved in cellular and mouse infections at early time points. MucR modulates the surface properties of the bacteria and their resistance to environmental stresses (i.e., oxidative stress, cationic peptide, and detergents). We show that B. melitensis mucR is a functional orthologue of S. meliloti mucR, because it was able to restore the production of succinoglycan in an S. meliloti mucR mutant, as detected by calcofluor staining. Similar to S. meliloti MucR, B. melitensis MucR also represses its own transcription and flagellar gene expression via the flagellar master regulator ftcR. More surprisingly, we demonstrate that MucR regulates a lipid A core modification in B. melitensis. These changes could account for the attenuated virulence of a mucR mutant. These data reinforce the idea that there is a common conserved circuitry between plant symbionts and animal pathogens that regulates the relationship they have with their hosts. PMID:23161025

  16. A collection of cytochrome P450 monooxygenase genes involved in modification and detoxification of herbicide atrazine in rice (Oryza sativa) plants.

    PubMed

    Rong Tan, Li; Chen Lu, Yi; Jing Zhang, Jing; Luo, Fang; Yang, Hong

    2015-09-01

    Plant cytochrome P450 monooxygenases constitute one of the largest families of protein genes involved in plant growth, development and acclimation to biotic and abiotic stresses. However, whether these genes respond to organic toxic compounds and their biological functions for detoxifying toxic compounds such as herbicides in rice are poorly understood. The present study identified 201 genes encoding cytochrome P450s from an atrazine-exposed rice transcriptome through high-throughput sequencing. Of these, 69 cytochrome P450 genes were validated by microarray and some of them were confirmed by real time PCR. Activities of NADPH-cytochrome P450 reductase (CPR) and p-nitroanisole O-demethylase (PNOD) related to toxicity were determined and significantly induced by atrazine exposure. To dissect the mechanism underlying atrazine modification and detoxification by P450, metabolites (or derivatives) of atrazine in plants were analyzed by ultra performance liquid chromatography mass spectrometry (UPLC/MS). Major metabolites comprised desmethylatrazine (DMA), desethylatrazine (DEA), desisopropylatrazine (DIA), hydroxyatrazine (HA), hydroxyethylatrazine (HEA) and hydroxyisopropylatrazine (HIA). All of them were chemically modified by P450s. Furthermore, two specific inhibitors of piperonyl butoxide (PBO) and malathion (MAL) were used to assess the correlation between the P450s activity and rice responses including accumulation of atrazine in tissues, shoot and root growth and detoxification.

  17. Analysis and interpretation of RNA splicing alterations in genes involved in genetic disorders.

    PubMed

    Vreeswijk, Maaike P G; van der Klift, Heleen M

    2012-01-01

    Germ line mutations in genes involved in hereditary cancer syndromes, such as BRCA1 and BRCA2 in breast cancer and MSH2, MSH6, MLH1, and PSM2 in hereditary nonpolyposis colorectal cancer (HNPCC, more recently indicated as Lynch syndrome), confer a high risk to develop cancer. Mutation analysis in these genes has resulted in the identification of a large number of sequence variants, of which mutations causing frame shifts and nonsense codons are considered undoubtedly to be pathogenic. Many variants, however, cannot be classified as either disease-causing mutations or neutral variants and are therefore called unclassified variants (UVs). A subset of these variants may have an effect on RNA splicing. Appropriate RNA analysis will enable the characterization of the exact molecular nature of this effect and hence, is essential to determine the clinical relevance of the genomic variant. This chapter describes the design and implementation of RNA analysis as an indispensible tool in today's clinical diagnostic setting.

  18. The genetic basis of inherited anomalies of the teeth. Part 2: syndromes with significant dental involvement.

    PubMed

    Bailleul-Forestier, Isabelle; Berdal, Ariane; Vinckier, Frans; de Ravel, Thomy; Fryns, Jean Pierre; Verloes, Alain

    2008-01-01

    Teeth are specialized structural components of the craniofacial skeleton. Developmental defects occur either alone or in combination with other birth defects. In this paper, we review the dental anomalies in several multiple congenital anomaly (MCA) syndromes, in which the dental component is pivotal in the recognition of the phenotype and/or the molecular basis of the disorder is known. We will consider successively syndromic forms of amelogenesis imperfecta or enamel defects, dentinogenesis imperfecta (i.e. osteogenesis imperfecta) and other dentine anomalies. Focusing on dental aspects, we will review a selection of MCA syndromes associated with teeth number and/or shape anomalies. A better knowledge of the dental phenotype may contribute to an earlier diagnosis of some MCA syndromes involving teeth anomalies. They may serve as a diagnostic indicator or help confirm a syndrome diagnosis. PMID:18599376

  19. Plasmid pEC156, a Naturally Occurring Escherichia coli Genetic Element That Carries Genes of the EcoVIII Restriction-Modification System, Is Mobilizable among Enterobacteria.

    PubMed

    Werbowy, Olesia; Kaczorowski, Tadeusz

    2016-01-01

    Type II restriction-modification systems are ubiquitous in prokaryotes. Some of them are present in naturally occurring plasmids, which may facilitate the spread of these systems in bacterial populations by horizontal gene transfer. However, little is known about the routes of their dissemination. As a model to study this, we have chosen an Escherichia coli natural plasmid pEC156 that carries the EcoVIII restriction modification system. The presence of this system as well as the cis-acting cer site involved in resolution of plasmid multimers determines the stable maintenance of pEC156 not only in Escherichia coli but also in other enterobacteria. We have shown that due to the presence of oriT-type F and oriT-type R64 loci it is possible to mobilize pEC156 by conjugative plasmids (F and R64, respectively). The highest mobilization frequency was observed when pEC156-derivatives were transferred between Escherichia coli strains, Enterobacter cloacae and Citrobacter freundii representing coliform bacteria. We found that a pEC156-derivative with a functional EcoVIII restriction-modification system was mobilized in enterobacteria at a frequency lower than a plasmid lacking this system. In addition, we found that bacteria that possess the EcoVIII restriction-modification system can efficiently release plasmid content to the environment. We have shown that E. coli cells can be naturally transformed with pEC156-derivatives, however, with low efficiency. The transformation protocol employed neither involved chemical agents (e.g. CaCl2) nor temperature shift which could induce plasmid DNA uptake.

  20. Plasmid pEC156, a Naturally Occurring Escherichia coli Genetic Element That Carries Genes of the EcoVIII Restriction-Modification System, Is Mobilizable among Enterobacteria

    PubMed Central

    Werbowy, Olesia; Kaczorowski, Tadeusz

    2016-01-01

    Type II restriction-modification systems are ubiquitous in prokaryotes. Some of them are present in naturally occurring plasmids, which may facilitate the spread of these systems in bacterial populations by horizontal gene transfer. However, little is known about the routes of their dissemination. As a model to study this, we have chosen an Escherichia coli natural plasmid pEC156 that carries the EcoVIII restriction modification system. The presence of this system as well as the cis-acting cer site involved in resolution of plasmid multimers determines the stable maintenance of pEC156 not only in Escherichia coli but also in other enterobacteria. We have shown that due to the presence of oriT-type F and oriT-type R64 loci it is possible to mobilize pEC156 by conjugative plasmids (F and R64, respectively). The highest mobilization frequency was observed when pEC156-derivatives were transferred between Escherichia coli strains, Enterobacter cloacae and Citrobacter freundii representing coliform bacteria. We found that a pEC156-derivative with a functional EcoVIII restriction-modification system was mobilized in enterobacteria at a frequency lower than a plasmid lacking this system. In addition, we found that bacteria that possess the EcoVIII restriction-modification system can efficiently release plasmid content to the environment. We have shown that E. coli cells can be naturally transformed with pEC156-derivatives, however, with low efficiency. The transformation protocol employed neither involved chemical agents (e.g. CaCl2) nor temperature shift which could induce plasmid DNA uptake. PMID:26848973

  1. Trends in lignin modification: a comprehensive analysis of the effects of genetic manipulations/mutations on lignification and vascular integrity

    NASA Technical Reports Server (NTRS)

    Anterola, Aldwin M.; Lewis, Norman G.

    2002-01-01

    A comprehensive assessment of lignin configuration in transgenic and mutant plants is long overdue. This review thus undertook the systematic analysis of trends manifested through genetic and mutational manipulations of the various steps associated with monolignol biosynthesis; this included consideration of the downstream effects on organized lignin assembly in the various cell types, on vascular function/integrity, and on plant growth and development. As previously noted for dirigent protein (homologs), distinct and sophisticated monolignol forming metabolic networks were operative in various cell types, tissues and organs, and form the cell-specific guaiacyl (G) and guaiacyl-syringyl (G-S) enriched lignin biopolymers, respectively. Regardless of cell type undergoing lignification, carbon allocation to the different monolignol pools is apparently determined by a combination of phenylalanine availability and cinnamate-4-hydroxylase/"p-coumarate-3-hydroxylase" (C4H/C3H) activities, as revealed by transcriptional and metabolic profiling. Downregulation of either phenylalanine ammonia lyase or cinnamate-4-hydroxylase thus predictably results in reduced lignin levels and impaired vascular integrity, as well as affecting related (phenylpropanoid-dependent) metabolism. Depletion of C3H activity also results in reduced lignin deposition, albeit with the latter being derived only from hydroxyphenyl (H) units, due to both the guaiacyl (G) and syringyl (S) pathways being blocked. Apparently the cells affected are unable to compensate for reduced G/S levels by increasing the amounts of H-components. The downstream metabolic networks for G-lignin enriched formation in both angiosperms and gymnosperms utilize specific cinnamoyl CoA O-methyltransferase (CCOMT), 4-coumarate:CoA ligase (4CL), cinnamoyl CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD) isoforms: however, these steps neither affect carbon allocation nor H/G designations, this being determined by C4H/C3H

  2. Plasmid-based genetic modification of human bone marrow-derived stromal cells: analysis of cell survival and transgene expression after transplantation in rat spinal cord

    PubMed Central

    Ronsyn, Mark W; Daans, Jasmijn; Spaepen, Gie; Chatterjee, Shyama; Vermeulen, Katrien; D'Haese, Patrick; Van Tendeloo, Viggo FI; Van Marck, Eric; Ysebaert, Dirk; Berneman, Zwi N; Jorens, Philippe G; Ponsaerts, Peter

    2007-01-01

    Background Bone marrow-derived stromal cells (MSC) are attractive targets for ex vivo cell and gene therapy. In this context, we investigated the feasibility of a plasmid-based strategy for genetic modification of human (h)MSC with enhanced green fluorescent protein (EGFP) and neurotrophin (NT)3. Three genetically modified hMSC lines (EGFP, NT3, NT3-EGFP) were established and used to study cell survival and transgene expression following transplantation in rat spinal cord. Results First, we demonstrate long-term survival of transplanted hMSC-EGFP cells in rat spinal cord under, but not without, appropriate immune suppression. Next, we examined the stability of EGFP or NT3 transgene expression following transplantation of hMSC-EGFP, hMSC-NT3 and hMSC-NT3-EGFP in rat spinal cord. While in vivo EGFP mRNA and protein expression by transplanted hMSC-EGFP cells was readily detectable at different time points post-transplantation, in vivo NT3 mRNA expression by hMSC-NT3 cells and in vivo EGFP protein expression by hMSC-NT3-EGFP cells was, respectively, undetectable or declined rapidly between day 1 and 7 post-transplantation. Further investigation revealed that the observed in vivo decline of EGFP protein expression by hMSC-NT3-EGFP cells: (i) was associated with a decrease in transgenic NT3-EGFP mRNA expression as suggested following laser capture micro-dissection analysis of hMSC-NT3-EGFP cell transplants at day 1 and day 7 post-transplantation, (ii) did not occur when hMSC-NT3-EGFP cells were transplanted subcutaneously, and (iii) was reversed upon re-establishment of hMSC-NT3-EGFP cell cultures at 2 weeks post-transplantation. Finally, because we observed a slowly progressing tumour growth following transplantation of all our hMSC cell transplants, we here demonstrate that omitting immune suppressive therapy is sufficient to prevent further tumour growth and to eradicate malignant xenogeneic cell transplants. Conclusion In this study, we demonstrate that genetically

  3. Genetic analysis of functions involved in adhesion of Pseudomonas putida to seeds.

    PubMed

    Espinosa-Urgel, M; Salido, A; Ramos, J L

    2000-05-01

    Many agricultural uses of bacteria require the establishment of efficient bacterial populations in the rhizosphere, for which colonization of plant seeds often constitutes a critical first step. Pseudomonas putida KT2440 is a strain that colonizes the rhizosphere of a number of agronomically important plants at high population densities. To identify the functions involved in initial seed colonization by P. putida KT2440, we subjected this strain to transposon mutagenesis and screened for mutants defective in attachment to corn seeds. Eight different mutants were isolated and characterized. While all of them showed reduced attachment to seeds, only two had strong defects in their adhesion to abiotic surfaces (glass and different plastics). Sequences of the loci affected in all eight mutants were obtained. None of the isolated genes had previously been described in P. putida, although four of them showed clear similarities with genes of known functions in other organisms. They corresponded to putative surface and membrane proteins, including a calcium-binding protein, a hemolysin, a peptide transporter, and a potential multidrug efflux pump. One other showed limited similarities with surface proteins, while the remaining three presented no obvious similarities with known genes, indicating that this study has disclosed novel functions.

  4. A Protein-Based Genetic Screening Uncovers Mutants Involved in Phytochrome Signaling in Arabidopsis

    PubMed Central

    Zhu, Ling; Xin, Ruijiao; Huq, Enamul

    2016-01-01

    Plants perceive red and far-red region of the light spectrum to regulate photomorphogenesis through a family of photoreceptors called phytochromes. Phytochromes transduce the light signals to trigger a cascade of downstream gene regulation in part via a subfamily of bHLH transcription factors called Phytochrome Interacting Factors (PIFs). As the repressors of light signaling pathways, most PIFs are phosphorylated and degraded through the ubiquitin/26S proteasome pathway in response to light. The mechanisms involved in the phosphorylation and degradation of PIFs have not been fully understood yet. Here we used an EMS mutagenesis and luminescent imaging system to identify mutants defective in the degradation of one of the PIFs, called PIF1. We identified five mutants named stable PIF (spf) that showed reduced degradation of PIF1 under light treatment in both luminescent imaging and immunoblot assays. The amounts of PIF1 in spf3, spf4, and spf5 were similar to a PIF1 missense mutant (PIF1–3M) that lacks interactions between PIF1 and phyA/phyB under light. The hypocotyl lengths of spf1 and spf2 were slightly longer under red light compared to the LUC-PIF1 control, while only spf1 displayed weak phenotype under far-red light conditions. Interestingly, the spf3, spf4, and spf5 displayed high abundance of PIF1, yet the hypocotyl lengths were similar to the wild type under these conditions. Cloning and characterization of these mutants will help identify key players in the light signaling pathways including, the light-regulated kinase(s) and the E3 ligase(s) necessary for the light-induced degradation of PIFs. PMID:27499759

  5. A Protein-Based Genetic Screening Uncovers Mutants Involved in Phytochrome Signaling in Arabidopsis.

    PubMed

    Zhu, Ling; Xin, Ruijiao; Huq, Enamul

    2016-01-01

    Plants perceive red and far-red region of the light spectrum to regulate photomorphogenesis through a family of photoreceptors called phytochromes. Phytochromes transduce the light signals to trigger a cascade of downstream gene regulation in part via a subfamily of bHLH transcription factors called Phytochrome Interacting Factors (PIFs). As the repressors of light signaling pathways, most PIFs are phosphorylated and degraded through the ubiquitin/26S proteasome pathway in response to light. The mechanisms involved in the phosphorylation and degradation of PIFs have not been fully understood yet. Here we used an EMS mutagenesis and luminescent imaging system to identify mutants defective in the degradation of one of the PIFs, called PIF1. We identified five mutants named stable PIF (spf) that showed reduced degradation of PIF1 under light treatment in both luminescent imaging and immunoblot assays. The amounts of PIF1 in spf3, spf4, and spf5 were similar to a PIF1 missense mutant (PIF1-3M) that lacks interactions between PIF1 and phyA/phyB under light. The hypocotyl lengths of spf1 and spf2 were slightly longer under red light compared to the LUC-PIF1 control, while only spf1 displayed weak phenotype under far-red light conditions. Interestingly, the spf3, spf4, and spf5 displayed high abundance of PIF1, yet the hypocotyl lengths were similar to the wild type under these conditions. Cloning and characterization of these mutants will help identify key players in the light signaling pathways including, the light-regulated kinase(s) and the E3 ligase(s) necessary for the light-induced degradation of PIFs. PMID:27499759

  6. Genetic Manipulation of Leishmania donovani to Explore the Involvement of Argininosuccinate Synthase in Oxidative Stress Management.

    PubMed

    Sardar, Abul Hasan; Jardim, Armando; Ghosh, Ayan Kumar; Mandal, Abhishek; Das, Sushmita; Saini, Savita; Abhishek, Kumar; Singh, Ruby; Verma, Sudha; Kumar, Ajay; Das, Pradeep

    2016-03-01

    Reactive oxygen and nitrogen species (ROS and RNS) produced by the phagocytic cells are the most common arsenals used to kill the intracellular pathogens. However, Leishmania, an intracellular pathogen, has evolved mechanisms to survive by counterbalancing the toxic oxygen metabolites produced during infection. Polyamines, the major contributor in this anti-oxidant machinery, are largely dependent on the availability of L-arginine in the intracellular milieu. Argininosuccinate synthase (ASS) plays an important role as the rate-limiting step required for converting L-citrulline to argininosuccinate to provide arginine for an assortment of metabolic processes. Leishmania produce an active ASS enzyme, yet it has an incomplete urea cycle as it lacks an argininosuccinate lyase (ASL). There is no evidence for endogenous synthesis of L-arginine in Leishmania, which suggests that these parasites salvage L-arginine from extracellular milieu and makes the biological function of ASS and the production of argininosuccinate in Leishmania unclear. Our previous quantitative proteomic analysis of Leishmania promastigotes treated with sub-lethal doses of ROS, RNS, or a combination of both, led to the identification of several differentially expressed proteins which included ASS. To assess the involvement of ASS in stress management, a mutant cell line with greatly reduced ASS activity was created by a double-targeted gene replacement strategy in L. donovani promastigote. Interestingly, LdASS is encoded by three copies of allele, but Western blot analysis showed the third allele did not appear to express ASS. The free thiol levels in the mutant LdASS-/-/+ cell line were decreased. Furthermore, the cell viability in L-arginine depleted medium was greatly attenuated on exposure to different stress environments and was adversely impacted in its ability to infect mice. These findings suggest that ASS is important for Leishmania donovani to counterbalance the stressed environments

  7. Genetic Manipulation of Leishmania donovani to Explore the Involvement of Argininosuccinate Synthase in Oxidative Stress Management.

    PubMed

    Sardar, Abul Hasan; Jardim, Armando; Ghosh, Ayan Kumar; Mandal, Abhishek; Das, Sushmita; Saini, Savita; Abhishek, Kumar; Singh, Ruby; Verma, Sudha; Kumar, Ajay; Das, Pradeep

    2016-03-01

    Reactive oxygen and nitrogen species (ROS and RNS) produced by the phagocytic cells are the most common arsenals used to kill the intracellular pathogens. However, Leishmania, an intracellular pathogen, has evolved mechanisms to survive by counterbalancing the toxic oxygen metabolites produced during infection. Polyamines, the major contributor in this anti-oxidant machinery, are largely dependent on the availability of L-arginine in the intracellular milieu. Argininosuccinate synthase (ASS) plays an important role as the rate-limiting step required for converting L-citrulline to argininosuccinate to provide arginine for an assortment of metabolic processes. Leishmania produce an active ASS enzyme, yet it has an incomplete urea cycle as it lacks an argininosuccinate lyase (ASL). There is no evidence for endogenous synthesis of L-arginine in Leishmania, which suggests that these parasites salvage L-arginine from extracellular milieu and makes the biological function of ASS and the production of argininosuccinate in Leishmania unclear. Our previous quantitative proteomic analysis of Leishmania promastigotes treated with sub-lethal doses of ROS, RNS, or a combination of both, led to the identification of several differentially expressed proteins which included ASS. To assess the involvement of ASS in stress management, a mutant cell line with greatly reduced ASS activity was created by a double-targeted gene replacement strategy in L. donovani promastigote. Interestingly, LdASS is encoded by three copies of allele, but Western blot analysis showed the third allele did not appear to express ASS. The free thiol levels in the mutant LdASS-/-/+ cell line were decreased. Furthermore, the cell viability in L-arginine depleted medium was greatly attenuated on exposure to different stress environments and was adversely impacted in its ability to infect mice. These findings suggest that ASS is important for Leishmania donovani to counterbalance the stressed environments

  8. Genetic Manipulation of Leishmania donovani to Explore the Involvement of Argininosuccinate Synthase in Oxidative Stress Management

    PubMed Central

    Sardar, Abul Hasan; Jardim, Armando; Ghosh, Ayan Kumar; Mandal, Abhishek; Das, Sushmita; Saini, Savita; Abhishek, Kumar; Singh, Ruby; Verma, Sudha; Kumar, Ajay; Das, Pradeep

    2016-01-01

    Reactive oxygen and nitrogen species (ROS and RNS) produced by the phagocytic cells are the most common arsenals used to kill the intracellular pathogens. However, Leishmania, an intracellular pathogen, has evolved mechanisms to survive by counterbalancing the toxic oxygen metabolites produced during infection. Polyamines, the major contributor in this anti-oxidant machinery, are largely dependent on the availability of L-arginine in the intracellular milieu. Argininosuccinate synthase (ASS) plays an important role as the rate-limiting step required for converting L-citrulline to argininosuccinate to provide arginine for an assortment of metabolic processes. Leishmania produce an active ASS enzyme, yet it has an incomplete urea cycle as it lacks an argininosuccinate lyase (ASL). There is no evidence for endogenous synthesis of L-arginine in Leishmania, which suggests that these parasites salvage L-arginine from extracellular milieu and makes the biological function of ASS and the production of argininosuccinate in Leishmania unclear. Our previous quantitative proteomic analysis of Leishmania promastigotes treated with sub-lethal doses of ROS, RNS, or a combination of both, led to the identification of several differentially expressed proteins which included ASS. To assess the involvement of ASS in stress management, a mutant cell line with greatly reduced ASS activity was created by a double-targeted gene replacement strategy in L. donovani promastigote. Interestingly, LdASS is encoded by three copies of allele, but Western blot analysis showed the third allele did not appear to express ASS. The free thiol levels in the mutant LdASS-/-/+ cell line were decreased. Furthermore, the cell viability in L-arginine depleted medium was greatly attenuated on exposure to different stress environments and was adversely impacted in its ability to infect mice. These findings suggest that ASS is important for Leishmania donovani to counterbalance the stressed environments

  9. Mitochondrial 12S rRNA A827G mutation is involved in the genetic susceptibility to aminoglycoside ototoxicity

    SciTech Connect

    Xing Guangqian; Chen Zhibin; Wei Qinjun; Tian Huiqin; Li Xiaolu; Zhou Aidong; Bu Xingkuan; Cao Xin . E-mail: caoxin@njmu.edu.cn

    2006-08-11

    We have analyzed the clinical and molecular characterization of a Chinese family with aminoglycoside-induced and non-syndromic hearing impairment. Clinical evaluations revealed that only those family members who had a history of exposure to aminoglycoside antibiotics subsequently developed hearing loss, suggesting mitochondrial genome involvement. Sequence analysis of the mitochondrial 12S rRNA and tRNA{sup Ser(UCN)} genes led to the identification of a homoplasmic A827G mutation in all maternal relatives, a mutation that was identified previously in a few sporadic patients and in another Chinese family with non-syndromic deafness. The pathogenicity of the A827G mutation is strongly supported by the occurrence of the same mutation in two independent families and several genetically unrelated subjects. The A827G mutation is located at the A-site of the mitochondrial 12S rRNA gene which is highly conserved in mammals. It is possible that the alteration of the tertiary or quaternary structure of this rRNA by the A827G mutation may lead to mitochondrial dysfunction, thereby playing a role in the pathogenesis of hearing loss and aminoglycoside hypersensitivity. However, incomplete penetrance of hearing impairment indicates that the A827G mutation itself is not sufficient to produce clinical phenotype but requires the involvement of modifier factors for the phenotypic expression. Indeed, aminoglycosides may contribute to the phenotypic manifestation of the A827G mutation in this family. In contrast with the congenital or early-onset hearing impairment in another Chinese family carrying the A827G mutation, three patients in this pedigree developed hearing loss only after use of aminoglycosides. This discrepancy likely reflects the difference of genetic backgrounds, either mitochondrial haplotypes or nuclear modifier genes, between two families.

  10. Trends in lignin modification: a comprehensive analysis of the effects of genetic manipulations/mutations on lignification and vascular integrity

    NASA Technical Reports Server (NTRS)

    Anterola, Aldwin M.; Lewis, Norman G.

    2002-01-01

    A comprehensive assessment of lignin configuration in transgenic and mutant plants is long overdue. This review thus undertook the systematic analysis of trends manifested through genetic and mutational manipulations of the various steps associated with monolignol biosynthesis; this included consideration of the downstream effects on organized lignin assembly in the various cell types, on vascular function/integrity, and on plant growth and development. As previously noted for dirigent protein (homologs), distinct and sophisticated monolignol forming metabolic networks were operative in various cell types, tissues and organs, and form the cell-specific guaiacyl (G) and guaiacyl-syringyl (G-S) enriched lignin biopolymers, respectively. Regardless of cell type undergoing lignification, carbon allocation to the different monolignol pools is apparently determined by a combination of phenylalanine availability and cinnamate-4-hydroxylase/"p-coumarate-3-hydroxylase" (C4H/C3H) activities, as revealed by transcriptional and metabolic profiling. Downregulation of either phenylalanine ammonia lyase or cinnamate-4-hydroxylase thus predictably results in reduced lignin levels and impaired vascular integrity, as well as affecting related (phenylpropanoid-dependent) metabolism. Depletion of C3H activity also results in reduced lignin deposition, albeit with the latter being derived only from hydroxyphenyl (H) units, due to both the guaiacyl (G) and syringyl (S) pathways being blocked. Apparently the cells affected are unable to compensate for reduced G/S levels by increasing the amounts of H-components. The downstream metabolic networks for G-lignin enriched formation in both angiosperms and gymnosperms utilize specific cinnamoyl CoA O-methyltransferase (CCOMT), 4-coumarate:CoA ligase (4CL), cinnamoyl CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD) isoforms: however, these steps neither affect carbon allocation nor H/G designations, this being determined by C4H/C3H

  11. Integration-free reprogramming of human somatic cells to induced pluripotent stem cells (iPSCs) without viral vectors, recombinant DNA, and genetic modification.

    PubMed

    Heng, Boon Chin; Fussenegger, Martin

    2014-01-01

    Stem cells are envisaged to be integral components of multicellular systems engineered for therapeutic applications. The reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) via recombinant expression of a limited number of transcription factors, which was first achieved by Yamanaka and colleagues in 2007, heralded a major breakthrough in the stem cell field. Since then, there has been rapid progress in the field of iPSC generation, including the identification of various small molecules that can enhance reprogramming efficiency and reduce the number of different transcription factors required for reprogramming. Nevertheless, the major obstacles facing clinical applications of iPSCs are safety concerns associated with the use of viral vectors and recombinant DNA for expressing the appropriate transcription factors during reprogramming. In particular, permanent genetic modifications to newly reprogrammed iPSCs have to be avoided in order to meet stringent safety requirements for clinical therapy. These safety challenges can be overcome by new technology platforms that enable cellular reprogramming to iPSCs without the need to utilize either recombinant DNA or viral vectors. The use of recombinant cell-penetrating peptides and direct transfection of synthetic mRNA encoding appropriate transcription factors have both been shown to successfully reprogram somatic cells to iPSCs. It has also been shown more recently that the direct transfection of certain miRNA species can reprogram somatic cells to pluripotency without the need for any of the transcription factors commonly utilized for iPSC generation. This chapter describes protocols for iPSC generation with these new techniques, which would obviate the use of recombinant DNA and viral vectors in cellular reprogramming, thus avoiding permanent genetic modification to the reprogrammed cells.

  12. Reverse genetic screening identifies five E-class PPR proteins involved in RNA editing in mitochondria of Arabidopsis thaliana.

    PubMed

    Takenaka, Mizuki; Verbitskiy, Daniil; Zehrmann, Anja; Brennicke, Axel

    2010-08-27

    RNA editing in flowering plant mitochondria post-transcriptionally alters several hundred nucleotides from C to U, mostly in mRNAs. Several factors required for specific RNA-editing events in plant mitochondria and plastids have been identified, all of them PPR proteins of the PLS subclass with a C-terminal E-domain and about half also with an additional DYW domain. Based on this information, we here probe the connection between E-PPR proteins and RNA editing in plant mitochondria. We initiated a reverse genetics screen of T-DNA insertion lines in Arabidopsis thaliana and investigated 58 of the 150 E-PPR-coding genes for a function in RNA editing. Six genes were identified to be involved in mitochondrial RNA editing at specific sites. Homozygous mutants of the five genes MEF18-MEF22 display no gross disturbance in their growth or development patterns, suggesting that the editing sites affected are not crucial at least in the greenhouse. These results show that a considerable percentage of the E-PPR proteins are involved in the functional processing of site-specific RNA editing in plant mitochondria.

  13. Application of capillary electrophoretic chips in protein profiling of plant extracts for identification of genetic modifications of maize.

    PubMed

    Poboży, Ewa; Filaber, Monika; Koc, Anna; Garcia-Reyes, Juan F

    2013-09-01

    In this study, the chip gel electrophoresis with LIF detection was applied in protein profiling of fractionated and total extracts of maize standards. The sensitivity of such determinations can be enhanced by lyophilization of extracts or employing filtering and preconcentration with cutoff filters. Combinatorial peptide ligand library applied for sample processing prior to the electrophoretic analysis was, especially, an effective pretreatment step in the determination of low-abundance proteins. Several repeatable differences were observed for protein profiles between maize standards not containing the genetically modified organisms (GMOs) and those containing GMO, which can be potentially employed for identification of GMO in maize samples and foods of maize origin. PMID:23856913

  14. Application of capillary electrophoretic chips in protein profiling of plant extracts for identification of genetic modifications of maize.

    PubMed

    Poboży, Ewa; Filaber, Monika; Koc, Anna; Garcia-Reyes, Juan F

    2013-09-01

    In this study, the chip gel electrophoresis with LIF detection was applied in protein profiling of fractionated and total extracts of maize standards. The sensitivity of such determinations can be enhanced by lyophilization of extracts or employing filtering and preconcentration with cutoff filters. Combinatorial peptide ligand library applied for sample processing prior to the electrophoretic analysis was, especially, an effective pretreatment step in the determination of low-abundance proteins. Several repeatable differences were observed for protein profiles between maize standards not containing the genetically modified organisms (GMOs) and those containing GMO, which can be potentially employed for identification of GMO in maize samples and foods of maize origin.

  15. Pathway analysis of whole exome sequence data provides further support for the involvement of histone modification in the aetiology of schizophrenia.

    PubMed

    Curtis, David

    2016-10-01

    Weighted burden pathway analysis was applied to whole exome sequence data for 2045 schizophrenic patients and 2045 controls. Overall, there was a statistically significant excess of pathways with more rare, functional variants in cases than in controls. Among the highest ranked were pathways relating to histone modification, as well as neuron differentiation and membrane and vesicle function. This bolsters the evidence from previous studies that histone modification pathways may be important in the aetiology of schizophrenia.

  16. Pathway analysis of whole exome sequence data provides further support for the involvement of histone modification in the aetiology of schizophrenia.

    PubMed

    Curtis, David

    2016-10-01

    Weighted burden pathway analysis was applied to whole exome sequence data for 2045 schizophrenic patients and 2045 controls. Overall, there was a statistically significant excess of pathways with more rare, functional variants in cases than in controls. Among the highest ranked were pathways relating to histone modification, as well as neuron differentiation and membrane and vesicle function. This bolsters the evidence from previous studies that histone modification pathways may be important in the aetiology of schizophrenia. PMID:26981879

  17. The potential for modification in cloning and vitrification technology to enhance genetic progress in beef cattle in Northern Australia.

    PubMed

    Taylor-Robinson, Andrew W; Walton, Simon; Swain, David L; Walsh, Kerry B; Vajta, Gábor

    2014-08-01

    Recent advances in embryology and related research offer considerable possibilities to accelerate genetic improvement in cattle breeding. Such progress includes optimization and standardization of laboratory embryo production (in vitro fertilization - IVF), introduction of a highly efficient method for cryopreservation (vitrification), and dramatic improvement in the efficiency of somatic cell nuclear transfer (cloning) in terms of required effort, cost, and overall outcome. Handmade cloning (HMC), a simplified version of somatic cell nuclear transfer, offers the potential for relatively easy and low-cost production of clones. A potentially modified method of vitrification used at a centrally located laboratory facility could result in cloned offspring that are economically competitive with elite animals produced by more traditional means. Apart from routine legal and intellectual property issues, the main obstacle that hampers rapid uptake of these technologies by the beef cattle industry is a lack of confidence from scientific and commercial sources. Once stakeholder support is increased, the combined application of these methods makes a rapid advance toward desirable traits (rapid growth, high-quality beef, optimized reproductive performance) a realistic goal. The potential impact of these technologies on genetic advancement in beef cattle herds in which improvement of stock is sought, such as in northern Australia, is hard to overestimate.

  18. Lentiviral vector-mediated genetic modification of cell substrates for the manufacture of proteins and other biologics.

    PubMed

    Baranyi, Lajos; Roy, Andre; Embree, Heather D; Dropulic, Boro

    2010-01-01

    Transduction with Lentiviral vectors has been shown to be the most efficient method for the stable delivery of nucleic acid sequences into mammalian cells. Lentiviral vectors have been widely used in research and have recently shown success in clinical trials for human gene therapy. In this paper, we describe the use of lentiviral vectors to generate genetically modified cell substrates for the manufacture of proteins and other complex biologics. The use of lentiviral vectors for the generation of genetically modified cell substrates for the production of biologic material has several advantages over other systems: (1) highly productive mammalian cell lines can be rapidly generated without selection or gene amplification; (2) the high number of vector copies are distributed throughout the open chromatin of the genome, resulting in cell lines that are extremely stable for high levels of gene expression and, consequently, protein production; and (3) high levels of protein glycosylation are maintained despite very high levels of protein production. These advantages offer the potential to significantly improve the quality, time-to-market, and manufacturing cost of biologics for human use.

  19. Spatial soil heterogeneity has a greater effect on symbiotic arbuscular mycorrhizal fungal communities and plant growth than genetic modification with Bacillus thuringiensis toxin genes.

    PubMed

    Cheeke, Tanya E; Schütte, Ursel M; Hemmerich, Chris M; Cruzan, Mitchell B; Rosenstiel, Todd N; Bever, James D

    2015-05-01

    Maize, genetically modified with the insect toxin genes of Bacillus thuringiensis (Bt), is widely cultivated, yet its impacts on soil organisms are poorly understood. Arbuscular mycorrhizal fungi (AMF) form symbiotic associations with plant roots and may be uniquely sensitive to genetic changes within a plant host. In this field study, the effects of nine different lines of Bt maize and their corresponding non-Bt parental isolines were evaluated on AMF colonization and community diversity in plant roots. Plants were harvested 60 days after sowing, and data were collected on plant growth and per cent AMF colonization of roots. AMF community composition in roots was assessed using 454 pyrosequencing of the 28S rRNA genes, and spatial variation in mycorrhizal communities within replicated experimental field plots was examined. Growth responses, per cent AMF colonization of roots and AMF community diversity in roots did not differ between Bt and non-Bt maize, but root and shoot biomass and per cent colonization by arbuscules varied by maize cultivar. Plot identity had the most significant effect on plant growth, AMF colonization and AMF community composition in roots, indicating spatial heterogeneity in the field. Mycorrhizal fungal communities in maize roots were autocorrelated within approximately 1 m, but at greater distances, AMF community composition of roots differed between plants. Our findings indicate that spatial variation and heterogeneity in the field has a greater effect on the structure of AMF communities than host plant cultivar or modification by Bt toxin genes.

  20. Epigenetic Modifications in Essential Hypertension

    PubMed Central

    Wise, Ingrid A.; Charchar, Fadi J.

    2016-01-01

    Essential hypertension (EH) is a complex, polygenic condition with no single causative agent. Despite advances in our understanding of the pathophysiology of EH, hypertension remains one of the world’s leading public health problems. Furthermore, there is increasing evidence that epigenetic modifications are as important as genetic predisposition in the development of EH. Indeed, a complex and interactive genetic and environmental system exists to determine an individual’s risk of EH. Epigenetics refers to all heritable changes to the regulation of gene expression as well as chromatin remodelling, without involvement of nucleotide sequence changes. Epigenetic modification is recognized as an essential process in biology, but is now being investigated for its role in the development of specific pathologic conditions, including EH. Epigenetic research will provide insights into the pathogenesis of blood pressure regulation that cannot be explained by classic Mendelian inheritance. This review concentrates on epigenetic modifications to DNA structure, including the influence of non-coding RNAs on hypertension development. PMID:27023534

  1. Genetic modification of the effect of maternal household air pollution exposure on birth weight in Guatemalan newborns

    PubMed Central

    Thompson, Lisa M.; Yousefi, Paul; Penaloza, Renee; Balmes, John; Holland, Nina

    2014-01-01

    Low birth weight is associated with exposure to air pollution during pregnancy. The purpose of this study was to evaluate whether null polymorphisms of Glutathione S-transferases (GSTs), specifically GSTM1 and GSTT1 genes in infants or mothers, modifies the association between high exposures to household air pollution (HAP) from cooking fires and birth weight. Pregnant women in rural Guatemala were randomized to receive a chimney stove or continue to use open fires for cooking. Newborns were measured within 48 hours of birth. 132 mother-infant pairs provided infant genotypes (n=130) and/or maternal genotypes (n=116). Maternal null GSTM1 was associated with a 144 gram (95% CI: -291, 1) and combined maternal/infant null GSTT1 was associated with a 155 gram (95% CI -303, -8) decrease in birth weight. Although there was a trend toward higher birth weights with increasing number of expressed GST genes, the effect modification by chimney stove use was not demonstrated. PMID:25305053

  2. Further evidence for genetic heterogeneity of distal HMN type V, CMT2 with predominant hand involvement and Silver syndrome

    PubMed Central

    Rohkamm, Barbara; Reilly, Mary M.; Lochmüller, Hanns; Schlotter-Weigel, Beate; Barisic, Nina; Schöls, Ludger; Nicholson, Garth; Pareyson, Davide; Laurà, Matilde; Janecke, Andreas R.; Miltenberger-Miltenyi, Gabriel; John, Elisabeth; Fischer, Carina; Grill, Franz; Wakeling, William; Davis, Mary; Pieber, Thomas R.; Auer-Grumbach, Michaela

    2011-01-01

    Objective Distal hereditary motor neuropathy type V (dHMN-V) and Charcot–Marie–Tooth syndrome (CMT) type 2 presenting with predominant hand involvement, also known as CMT2D and Silver syndrome (SS) are rare phenotypically overlapping diseases which can be caused by mutations in the Berardinelli–Seip Congenital Lipodystrophy 2 (BSCL2) and in the glycyl-tRNA synthetase encoding (GARS) genes. Mutations in the heat-shock proteins HSPB1 and HSPB8 can cause related distal hereditary motor neuropathies (dHMN) and are considered candidates for dHMN-V, CMT2, and SS. Design To define the frequency and distribution of mutations in the GARS, BSCL2, HSPB1 and HSPB8 genes we screened 33 unrelated sporadic and familial patients diagnosed as either dHMN-V, CMT2D or SS. Exon 3 of the BSCL2 gene was screened in further 69 individuals with an unclassified dHMN phenotype or diagnosed as hereditary spastic paraplegia (HSP) complicated by pure motor neuropathy. Results Four patients diagnosed with dHMN-Vor SS carried known heterozygous BSCL2 mutations (N88S and S90L). In one dHMN-V patient we detected a putative GARS mutation (A57V). No mutations were detected in HSPB1 and HSPB8. The diagnostic yield gained in the series of 33 probands was 12% for BSCL2 mutations and 3% for GARS mutations. In the series of unclassified dHMN and complicated HSP cases no mutations were found. Conclusions Our data confirm that most likely only two mutations (N88S, S90L) in exon 3 of BSCL2 may lead to dHMN-V or SS phenotypes. Mutations in GARS, HSPB1 and HSPB8. are not a common cause of dHMN-V, SS and CMT2D. We would therefore suggest that a genetic testing of dHMN-V and SS patients should begin with screening of exon 3 of the BSCL2 gene. Screening of the GARS gene is useful in patients with CMT2 with predominant hand involvement and dHMN-V. The rather low frequencies of BSCL2, GARS, HSPB1 and HSPB8 mutations in dHMN-V, CMT2D and SS patients strongly point to further genetic heterogeneity of these

  3. Attention Deficit Hyperactivity Disorder with Reading Disabilities: Preliminary Genetic Findings on the Involvement of the ADRA2A Gene

    ERIC Educational Resources Information Center

    Stevenson, J.; Langley, K.; Pay, H.; Payton, A.; Worthington, J.; Ollier, W.; Thapar, A.

    2005-01-01

    Background: Attention deficit/hyperactivity disorder (ADHD) and reading disability (RD) tend to co-occur and quantitative genetic studies have shown this to arise primarily through shared genetic influences. However, molecular genetic studies have shown different genes to be associated with each of these conditions. Neurobiological studies have…

  4. Proteomics insight into the biological safety of transgenic modification of rice as compared with conventional genetic breeding and spontaneous genotypic variation.

    PubMed

    Gong, Chun Yan; Li, Qi; Yu, Hua Tao; Wang, Zizhang; Wang, Tai

    2012-05-01

    The potential of unintended effects caused by transgenic events is a key issue in the commercialization of genetically modified (GM) crops. To investigate whether transgenic events cause unintended effects, we used comparative proteomics approaches to evaluate proteome differences in seeds from 2 sets of GM indica rice, herbicide-resistant Bar68-1 carrying bar and insect-resistant 2036-1a carrying cry1Ac/sck, and their respective controls D68 and MH86, as well as indica variety MH63, a parental line for breeding MH86, and japonica variety ZH10. This experimental design allowed for comparing proteome difference caused by transgenes, conventional genetic breeding, and natural genetic variation. Proteomics analysis revealed the maximum numbers of differentially expressed proteins between indica and japonica cultivars, second among indica varieties with relative small difference between MH86 and MH63, and the minimum between GM rice and respective control, thus indicating GM events do not substantially alter proteome profiles as compared with conventional genetic breeding and natural genetic variation. Mass spectrometry analysis revealed 234 proteins differentially expressed in the 6 materials, and these proteins were involved in different cellular and metabolic processes with a prominent skew toward metabolism (31.2%), protein synthesis and destination (25.2%), and defense response (22.4%). In these seed proteomes, proteins implicated in the 3 prominent biological processes showed significantly different composite expression patterns and were major factors differentiating japonica and indica cultivars, as well as indica varieties. Thus, metabolism, protein synthesis and destination, and defense response in seeds are important in differentiating rice cultivars and varieties.

  5. Genetic Modification of the Marine-Isolated Yeast Aureobasidium melanogenum P16 for Efficient Pullulan Production from Inulin.

    PubMed

    Ma, Zai-Chao; Liu, Nan-Nan; Chi, Zhe; Liu, Guang-Lei; Chi, Zhen-Ming

    2015-08-01

    In this study, in order to directly and efficiently convert inulin into pullulan, the INU1 gene from Kluyveromyces maximum KM was integrated into the genomic DNA and actively expressed in the high pullulan producer Aureobasidium melanogenum P16 isolated from the mangrove ecosystem. After the ability to produce pullulan from inulin by different transformants was examined, it was found that the recombinant strain EI36, one of the transformants, produced 40.92 U/ml of inulinase activity while its wild-type strain P16 only yielded 7.57 U/ml of inulinase activity. Most (99.27 %) of the inulinase produced by the recombinant strain EI36 was secreted into the culture. During the 10-l fermentation, 70.57 ± 1.3 g/l of pullulan in the fermented medium was attained from inulin (138.0 g/l) within 108 h, high inulinase activity (42.03 U/ml) was produced within 60 h, the added inulin was actively hydrolyzed by the secreted inulinase, and most of the reducing sugars were used by the recombinant strain EI36. This confirmed that the genetically engineered yeast of A. melanogenum strain P16 was suitable for direct pullulan production from inulin.

  6. Genetic Modification of the Marine-Isolated Yeast Aureobasidium melanogenum P16 for Efficient Pullulan Production from Inulin.

    PubMed

    Ma, Zai-Chao; Liu, Nan-Nan; Chi, Zhe; Liu, Guang-Lei; Chi, Zhen-Ming

    2015-08-01

    In this study, in order to directly and efficiently convert inulin into pullulan, the INU1 gene from Kluyveromyces maximum KM was integrated into the genomic DNA and actively expressed in the high pullulan producer Aureobasidium melanogenum P16 isolated from the mangrove ecosystem. After the ability to produce pullulan from inulin by different transformants was examined, it was found that the recombinant strain EI36, one of the transformants, produced 40.92 U/ml of inulinase activity while its wild-type strain P16 only yielded 7.57 U/ml of inulinase activity. Most (99.27 %) of the inulinase produced by the recombinant strain EI36 was secreted into the culture. During the 10-l fermentation, 70.57 ± 1.3 g/l of pullulan in the fermented medium was attained from inulin (138.0 g/l) within 108 h, high inulinase activity (42.03 U/ml) was produced within 60 h, the added inulin was actively hydrolyzed by the secreted inulinase, and most of the reducing sugars were used by the recombinant strain EI36. This confirmed that the genetically engineered yeast of A. melanogenum strain P16 was suitable for direct pullulan production from inulin. PMID:25985744

  7. The cox1 gene from Euglena gracilis: a protist mitochondrial gene without introns and genetic code modifications.

    PubMed

    Tessier, L H; van der Speck, H; Gualberto, J M; Grienenberger, J M

    1997-03-01

    We present the nucleotide sequence of the cox 1 gene encoding subunit 1 of cytochrome c oxidase in Euglena gracilis, the first report on a mitochondrial gene from this protist. Its study reveals that the Euglena mitochondrial genome does not appear as a compact and homogeneous structure and that its A+T content is high (about 76%) whereas this value is less than 50% in nuclear DNA. The Euglena cox1 gene does not exhibit any intron, and an amino-acid alignment of Euglena COX1 with homologous proteins shows that the universal genetic code is used. Comparisons of the genomic and cDNA sequences of Euglena cox1 indicate that the transcript does not undergo RNA editing as found in trypanosomes and in higher plants. The phylogeny obtained with COX1 protein sequences is in agreement with that obtained with nuclear rRNA sequences and places Euglena and Trypanosoma far apart from other eukaryotes. This result strengthens the hypothesis that these protists represent the earliest mitochondrion-containing organisms.

  8. Testing directed evolution strategies for space exploration: genetic modification of photosystem II to increase stress tolerance under space conditions

    NASA Astrophysics Data System (ADS)

    Bertalan, I.; Giardi, M. T.; Johanningmeier, U.

    Plants and many microorganisms are able to convert and store solar energy in chemical bonds by a process called photosynthesis They remove CO 2 from the atmosphere fix it as carbohydrate and simultaneously evolve oxygen Oxygen evolution is of supreme relevance for all higher life forms and results from the splitting of water molecules This process is catalyzed by the so called photosystem II PSII complex and represents the very beginning of biomass production PS II is also a central point of regulation being responsive to various physical and physiological parameters Complex space radiation is damaging PS II and reduces photosynthetic efficiency Thus bioregenerative life-support systems are severely disturbed at this point Genetic manipulation of photosynthesis checkpoints offer the possibility to adjust biomass and oxygen production to changing environmental conditions As the photosynthetic apparatus has adapted to terrestrial and not to space conditions we are trying to adapt a central and particularly stress-susceptible element of the photosynthesis apparatus - the D1 subunit of PS II - to space radiation by a strategy of directed evolution The D1 subunit together with its sister subunit D2 form the reaction centre of PS II D1 presents a central weak point for radiation energy that hits the chloroplast We have constructed a mutant of the green alga Chlamydomonas reinhardtii with a defect D1 protein This mutant is easily transformable with D1-encoding PCR fragments without purification and cloning steps 1 When

  9. Cellular and genetic mechanisms involved in the generation of protective and pathogenic immune responses in human Chagas disease

    PubMed Central

    Dutra, Walderez Ornelas; Menezes, Cristiane Alves Silva; Villani, Fernanda Nobre Amaral; da Costa, Germano Carneiro; da Silveira, Alexandre Barcelos Morais; d’Ávila Reis, Débora; Gollob, Kenneth J

    2012-01-01

    Perhaps one of the most intriguing aspects of human Chagas disease is the complex network of events that underlie the generation of protective versus pathogenic immune responses during the chronic phase of the disease. While most individuals do not develop patent disease, a large percentage may develop severe forms that eventually lead to death. Although many efforts have been devoted to deciphering these mechanisms, there is still much to be learned before we can fully understand the pathogenesis of Chagas disease. It is clear that the host’s immune response is decisive in this process. While characteristics of the parasite influence the immune response, it is becoming evident that the host genetic background plays a fundamental role in the establishment of pathogenic versus protective responses. The involvement of three complex organisms, host, parasite and vector, is certainly one of the key aspects that calls for multidisciplinary approaches towards the understanding of Chagas disease. We believe that now, one hundred years after the discovery of Chagas disease, it is imperative to continue with highly interactive research in order to elucidate the immune response associated with disease evolution, which will be essential in designing prophylactic or therapeutic interventions. PMID:19753476

  10. Genetics and Gene Expression Involving Stress and Distress Pathways in Fibromyalgia with and without Comorbid Chronic Fatigue Syndrome

    PubMed Central

    Light, Kathleen C.; White, Andrea T.; Tadler, Scott; Iacob, Eli; Light, Alan R.

    2012-01-01

    In complex multisymptom disorders like fibromyalgia syndrome (FMS) and chronic fatigue syndrome (CFS) that are defined primarily by subjective symptoms, genetic and gene expression profiles can provide very useful objective information. This paper summarizes research on genes that may be linked to increased susceptibility in developing and maintaining these disorders, and research on resting and stressor-evoked changes in leukocyte gene expression, highlighting physiological pathways linked to stress and distress. These include the adrenergic nervous system, the hypothalamic-pituitary-adrenal axis and serotonergic pathways, and exercise responsive metabolite-detecting ion channels. The findings to date provide some support for both inherited susceptibility and/or physiological dysregulation in all three systems, particularly for catechol-O-methyl transferase (COMT) genes, the glucocorticoid and the related mineralocorticoid receptors (NR3C1, NR3C2), and the purinergic 2X4 (P2X4) ion channel involved as a sensory receptor for muscle pain and fatigue and also in upregulation of spinal microglia in chronic pain models. Methodological concerns for future research, including potential influences of comorbid clinical depression and antidepressants and other medications, on gene expression are also addressed. PMID:22110941

  11. Effect modification by population dietary folate on the association between MTHFR genotype, homocysteine, and stroke risk: a meta-analysis of genetic studies and randomised trials

    PubMed Central

    Holmes, Michael V; Newcombe, Paul; Hubacek, Jaroslav A; Sofat, Reecha; Ricketts, Sally L; Cooper, Jackie; Breteler, Monique MB; Bautista, Leonelo E; Sharma, Pankaj; Whittaker, John C; Smeeth, Liam; Fowkes, F Gerald R; Algra, Ale; Shmeleva, Veronika; Szolnoki, Zoltan; Roest, Mark; Linnebank, Michael; Zacho, Jeppe; Nalls, Michael A; Singleton, Andrew B; Ferrucci, Luigi; Hardy, John; Worrall, Bradford B; Rich, Stephen S; Matarin, Mar; Norman, Paul E; Flicker, Leon; Almeida, Osvaldo P; van Bockxmeer, Frank M; Shimokata, Hiroshi; Khaw, Kay-Tee; Wareham, Nicholas J; Bobak, Martin; Sterne, Jonathan AC; Smith, George Davey; Talmud, Philippa J; van Duijn, Cornelia; Humphries, Steve E; Price, Jackie F; Ebrahim, Shah; Lawlor, Debbie A; Hankey, Graeme J; Meschia, James F; Sandhu, Manjinder S; Hingorani, Aroon D; Casas, Juan P

    2011-01-01

    folate status (predicted RR 1·00, 95% CI 0·90 to 1·11). Although the predicted effect of homocysteine reduction from large genetic studies in low folate regions (Asia) was larger (RR 0·78, 95% CI 0·68 to 0·90), no trial has evaluated the effect of lowering of homocysteine on stroke risk exclusively in a low folate region. Interpretation In regions with increasing levels or established policies of population folate supplementation, evidence from genetic studies and randomised trials is concordant in suggesting an absence of benefit from lowering of homocysteine for prevention of stroke. Further large-scale genetic studies of the association between MTHFR 677C→T and stroke in low folate settings are needed to distinguish effect modification by folate from small-study bias. If future randomised trials of homocysteine-lowering interventions for stroke prevention are undertaken, they should take place in regions with low folate consumption. Funding Full funding sources listed at end of paper (see Acknowledgments). PMID:21803414

  12. Genome-wide analysis of small nucleolar RNAs of Leishmania major reveals a rich repertoire of RNAs involved in modification and processing of rRNA

    PubMed Central

    Eliaz, Dror; Doniger, Tirza; Tkacz, Itai Dov; Biswas, Viplov Kumar; Gupta, Sachin Kumar; Kolev, Nikolay G; Unger, Ron; Ullu, Elisabetta; Tschudi, Christian; Michaeli, Shulamit

    2015-01-01

    Trypanosomatids are protozoan parasites and the causative agent of infamous infectious diseases. These organisms regulate their gene expression mainly at the post-transcriptional level and possess characteristic RNA processing mechanisms. In this study, we analyzed the complete repertoire of Leishmania major small nucleolar (snoRNA) RNAs by performing RNA-seq analysis on RNAs that were affinity-purified using the C/D snoRNA core protein, SNU13, and the H/ACA core protein, NHP2. This study revealed a large collection of C/D and H/ACA snoRNAs, organized in gene clusters generally containing both snoRNA types. Abundant snoRNAs were identified and predicted to guide trypanosome-specific rRNA cleavages. The repertoire of snoRNAs was compared to that of the closely related Trypanosoma brucei, and 80% of both C/D and H/ACA molecules were found to have functional homologues. The comparative analyses elucidated how snoRNAs evolved to generate molecules with analogous functions in both species. Interestingly, H/ACA RNAs have great flexibility in their ability to guide modifications, and several of the RNA species can guide more than one modification, compensating for the presence of single hairpin H/ACA snoRNA in these organisms. Placing the predicted modifications on the rRNA secondary structure revealed hypermodification regions mostly in domains which are modified in other eukaryotes, in addition to trypanosome-specific modifications. PMID:25970223

  13. Genetic modifications to temperate Enterococcus faecalis phage Ef11 that abolish the establishment of lysogeny and sensitivity to repressor, and increase host range and productivity of lytic infection.

    PubMed

    Zhang, H; Fouts, D E; DePew, J; Stevens, R H

    2013-06-01

    Ef11 is a temperate bacteriophage originally isolated by induction from a lysogenic Enterococcus faecalis strain recovered from an infected root canal, and the Ef11 prophage is widely disseminated among strains of E. faecalis. Because E. faecalis has emerged as a significant opportunistic human pathogen, we were interested in examining the genes and regulatory sequences predicted to be critical in the establishment/maintenance of lysogeny by Ef11 as a first step in the construction of the genome of a virulent, highly lytic phage that could be used in treating serious E. faecalis infections. Passage of Ef11 in E. faecalis JH2-2 yielded a variant that produced large, extensively spreading plaques in lawns of indicator cells, and elevated phage titres in broth cultures. Genetic analysis of the cloned virus producing the large plaques revealed that the variant was a recombinant between Ef11 and a defective FL1C-like prophage located in the E. faecalis JH2-2 chromosome. The recombinant possessed five ORFs of the defective FL1C-like prophage in place of six ORFs of the Ef11 genome. Deletion of the putative lysogeny gene module (ORFs 31-36) and replacement of the putative cro promoter from the recombinant phage genome with a nisin-inducible promoter resulted in no loss of virus infectivity. The genetic construct incorporating all the aforementioned Ef11 genomic modifications resulted in the generation of a variant that was incapable of lysogeny and insensitive to repressor, rendering it virulent and highly lytic, with a notably extended host range. PMID:23579685

  14. Modulation of monoamine oxidase (MAO) expression in neuropsychiatric disorders: genetic and environmental factors involved in type A MAO expression.

    PubMed

    Naoi, Makoto; Riederer, Peter; Maruyama, Wakako

    2016-02-01

    -A activity may increase the levels of serotonin and norepinephrine, resulting in disturbed neurotransmitter system development and behavior. This review discusses genetic and environmental factors involved in the regulation of MAO-A expression, in the contexts of neuropsychiatric function and of the regulation of neuronal survival and death.

  15. Enhancement of antitumor activity of gammaretrovirus carrying IL-12 gene through genetic modification of envelope targeting HER2 receptor: a promising strategy for bladder cancer therapy.

    PubMed

    Tsai, Y-S; Shiau, A-L; Chen, Y-F; Tsai, H-T; Tzai, T-S; Wu, C-L

    2010-01-01

    The objective of this study was to develop an HER2-targeted, envelope-modified Moloney murine leukemia virus (MoMLV)-based gammaretroviral vector carrying interleukin (IL)-12 gene for bladder cancer therapy. It displayed a chimeric envelope protein containing a single-chain variable fragment (scFv) antibody to the HER2 receptor and carried the mouse IL-12 gene. The fragment of anti-erbB2scFv was constructed into the proline-rich region of the viral envelope of the packaging vector lacking a transmembrane subunit of the carboxyl terminal region of surface subunit. As compared with envelope-unmodified gammaretroviruses, envelope-modified ones had extended viral tropism to human HER2-expressing bladder cancer cell lines, induced apoptosis, and affected cell cycle progression despite lower viral titers. Moreover, animal studies showed that envelope-modified gammaretroviruses carrying IL-12 gene exerted higher antitumor activity in terms of retarding tumor growth and prolonging the survival of tumor-bearing mice than unmodified ones, which were associated with enhanced tumor cell apoptosis as well as increased intratumoral levels of IL-12, interferon-gamma, IL-1beta, and tumor necrosis factor-alpha proteins. Therefore, the antitumor activity of gammaretroviruses carrying the IL-12 gene was enhanced through genetic modification of the envelope targeting HER2 receptor, which may be a promising strategy for bladder cancer therapy.

  16. Chemical modification of thiol groups of mitochondrial F1-ATPase from the yeast Schizosaccharomyces pombe. Involvement of alpha- and gamma-subunits in the enzyme activity

    SciTech Connect

    Falson, P.; Di Pietro, A.; Gautheron, D.C.

    1986-06-05

    Mitochondrial F1-ATPase from the yeast Schizosaccharomyces pombe has been prepared under a stable form and in relatively high amounts by an improved purification procedure. Specific chemical modification of the enzyme by the thiol reagent N-ethylmaleimide (NEM) at pH 6.8 leads to complete inactivation characterized by complex kinetics and pH dependence, indicating that several thiols are related to the enzyme activity. A complete protection against NEM effect is afforded by low concentrations of nucleotides in the presence of Mg/sup 2 +/, with ADP and ATP being more efficient than GTP. A total binding of 5 mol of (/sup 14/C)NEM/mol of F1-ATPase is obtained when the enzyme is 85% inactivated: 3 mol of the label are located on the alpha-subunits and 2 on the gamma-subunit. Two out of the 3 mol on the alpha-subunits bind very rapidly before any inactivation occurs. Complete protection by ATP against inactivation by NEM prevents the modification of three essential thiols out of the group of five thiols labeled in the absence of ATP: one is located on a alpha-subunit and two on the gamma-subunit. These two essential thiols of the gamma-subunit can be differentiated by modification with 6,6'-dithiodinicotinic acid (CPDS), another specific thiol reagent. A maximal binding of 4 mol of (/sup 14/C)CPDS/mol of enzyme is obtained, concomitant to a 25% inhibition. Sequential modification of the enzyme by CPDS and (/sup 14/C)NEM leads to the same final deep inactivation as that obtained with (/sup 14/C)NEM alone. One out of the two thiols of the gamma-subunit is no longer accessible to (/sup 14/C)NEM after CPDS treatment. When incubated at pH 6.8 with (/sup 3/H)ATP in the presence of Mg/sup 2 +/, F1-ATPase is able to bind 3, largely exchangeable, mol of nucleotide/mol of enzyme. Modification of the three essential thiols by NEM dramatically decreases the binding of /sup 3/H-nucleotide down to about 1 mol/mol of enzyme.

  17. Genetics

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genus Capsicum represents one of several well characterized Solanaceous genera. A wealth of classical and molecular genetics research is available for the genus. Information gleaned from its cultivated relatives, tomato and potato, provide further insight for basic and applied studies. Early ...

  18. Genetics

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Maintaining genetic variation in wild populations of Arctic organisms is fundamental to the long-term persistence of high latitude biodiversity. Variability is important because it provides options for species to respond to changing environmental conditions and novel challenges such as emerging path...

  19. Complete Bordetella avium, Bordetella hinzii and Bordetella trematum lipid A structures and genomic sequence analyses of the loci involved in their modifications.

    PubMed

    Novikov, Alexey; Shah, Nita R; AlBitar-Nehme, Sami; Basheer, Soorej M; Trento, Ilaria; Tirsoaga, Alina; Moksa, Michelle; Hirst, Martin; Perry, Malcolm B; Hamidi, Asmaa El; Fernandez, Rachel C; Caroff, Martine

    2014-08-01

    Endotoxin is recognized as one of the virulence factors of the Bordetella avium bird pathogen, and characterization of its structure and corresponding genomic features are important for an understanding of its role in pathogenicity and for an improved general knowledge of Bordetella spp virulence factors. The structure of the biologically active part of B. avium LPS, lipid A, is described and compared to those of another bird pathogen, opportunistic in humans, Bordetella hinzii, and to that of Bordetella trematum, a human pathogen. Sequence analyses showed that the three strains have homologues of acyl-chain modifying enzymes PagL, PagP and LpxO, of the 1-phosphatase LpxE, in addition to LgmA, LgmB and LgmC, which are required for the glucosamine modification. MALDI mass spectrometry identified a high amount of glucosamine substituting the phosphate groups of B. avium lipid A; this modification was absent from B. hinzii and B. trematum. The acylation patterns of the three lipid As were similar, but they differed from those of Bordetella pertussis and Bordetella parapertussis. They were also found to be close to the lipid A structure of Bordetella bronchiseptica, a mammalian pathogen, only differing from the latter by the degree of hydroxylation of the branched fatty acid.

  20. Genetic modification of T cells.

    PubMed

    Morgan, Richard A; Kakarla, Sunitha

    2014-01-01

    Gene transfer technology has advanced rapidly from simple physical-chemical laboratory methods in the 1970s and 1980s to the sophisticated viral and nonviral methods currently in clinical practice. Herein, we review 4 gene transfer methodologies applied in human gene therapy clinical trials transferring chimeric antigen receptors into T cells for the treatment of B-cell malignancies. The 4 methods include 2 viral vector gene transfer technologies, gamma retroviral vectors and lentiviral vectors, and 2 nonviral methods, transposons and mRNA electroporation.

  1. Genetic background and epigenetic modifications in the core of the nucleus accumbens predict addiction-like behavior in a rat model.

    PubMed

    Flagel, Shelly B; Chaudhury, Sraboni; Waselus, Maria; Kelly, Rebeca; Sewani, Salima; Clinton, Sarah M; Thompson, Robert C; Watson, Stanley J; Akil, Huda

    2016-05-17

    This study provides a demonstration in the rat of a clear genetic difference in the propensity for addiction-related behaviors following prolonged cocaine self-administration. It relies on the use of selectively bred high-responder (bHR) and low-responder (bLR) rat lines that differ in several characteristics associated with "temperament," including novelty-induced locomotion and impulsivity. We show that bHR rats exhibit behaviors reminiscent of human addiction, including persistent cocaine-seeking and increased reinstatement of cocaine seeking. To uncover potential underlying mechanisms of this differential vulnerability, we focused on the core of the nucleus accumbens and examined expression and epigenetic regulation of two transcripts previously implicated in bHR/bLR differences: fibroblast growth factor (FGF2) and the dopamine D2 receptor (D2). Relative to bHRs, bLRs had lower FGF2 mRNA levels and increased association of a repressive mark on histones (H3K9me3) at the FGF2 promoter. These differences were apparent under basal conditions and persisted even following prolonged cocaine self-administration. In contrast, bHRs had lower D2 mRNA under basal conditions, with greater association of H3K9me3 at the D2 promoter and these differences were no longer apparent following prolonged cocaine self-administration. Correlational analyses indicate that the association of H3K9me3 at D2 may be a critical substrate underlying the propensity to relapse. These findings suggest that low D2 mRNA levels in the nucleus accumbens core, likely mediated via epigenetic modifications, may render individuals more susceptible to cocaine addiction. In contrast, low FGF2 levels, which appear immutable even following prolonged cocaine exposure, may serve as a protective factor. PMID:27114539

  2. Genetic Modification of Mesenchymal Stem Cells Overexpressing CCR1 Increases Cell Viability, Migration, Engraftment and Capillary Density in the Injured Myocardium

    PubMed Central

    Huang, Jing; Zhang, Zhiping; Guo, Jian; Ni, Aiguo; Deb, Arjun; Zhang, Lunan; Mirotsou, Maria; Pratt, Richard E; Dzau, Victor J

    2010-01-01

    Rationale Although mesenchymal stem cell (MSC) transplantation has been shown to promote cardiac repair in acute myocardial injury in vivo, its overall restorative capacity appears to be restricted mainly due to poor cell viability and low engraftment in the ischemic myocardium. Specific chemokines are upregulated in the infarcted myocardium. However the expression levels of the corresponding chemokine receptors (e.g. CCR1, CXCR2) in MSCs are very low. We hypothesized that this discordance may account for the poor MSC engraftment and survival. Objective To determine whether overexpression of CCR1 or CXCR2 chemokine receptors in MSCs augments their cell survival, migration and engraftment after injection in the infarcted myocardium. Methods and Results Overexpression of CCR1, but not CXCR2, dramatically increased chemokine-induced murine MSC migration and protected MSC from apoptosis in vitro. Moreover, when MSCs were injected intramyocardially one hour after coronary artery ligation, CCR1-MSCs accumulated in the infarcted myocardium at significantly higher levels than control-MSCs or CXCR2-MSCs three days post-myocardial infarction (MI). CCR1-MSC injected hearts exhibited a significant reduction in infarct size, reduced cardiomyocytes apoptosis and increased capillary density in injured myocardium three days post-MI. Furthermore, intramyocardial injection of CCR1-MSCs prevented cardiac remodeling and restored cardiac function 4 weeks post-MI. Conclusions Our results demonstrate the in vitro and in vivo salutary effects of genetic modification of stem cells. Specifically, overexpression of chemokine receptor enhances the migration, survival and engraftment of MSCs, and may provide a new therapeutic strategy for the injured myocardium. PMID:20378860

  3. Genetic Stabilization of the Drug-Resistant PMEN1 Pneumococcus Lineage by Its Distinctive DpnIII Restriction-Modification System

    PubMed Central

    Eutsey, Rory A.; Powell, Evan; Dordel, Janina; Salter, Susannah J.; Clark, Tyson A.; Korlach, Jonas

    2015-01-01

    ABSTRACT The human pathogen Streptococcus pneumoniae (pneumococcus) exhibits a high degree of genomic diversity and plasticity. Isolates with high genomic similarity are grouped into lineages that undergo homologous recombination at variable rates. PMEN1 is a pandemic, multidrug-resistant lineage. Heterologous gene exchange between PMEN1 and non-PMEN1 isolates is directional, with extensive gene transfer from PMEN1 strains and only modest transfer into PMEN1 strains. Restriction-modification (R-M) systems can restrict horizontal gene transfer, yet most pneumococcal strains code for either the DpnI or DpnII R-M system and neither limits homologous recombination. Our comparative genomic analysis revealed that PMEN1 isolates code for DpnIII, a third R-M system syntenic to the other Dpn systems. Characterization of DpnIII demonstrated that the endonuclease cleaves unmethylated double-stranded DNA at the tetramer sequence 5′ GATC 3′, and the cognate methylase is a C5 cytosine-specific DNA methylase. We show that DpnIII decreases the frequency of recombination under in vitro conditions, such that the number of transformants is lower for strains transformed with unmethylated DNA than in those transformed with cognately methylated DNA. Furthermore, we have identified two PMEN1 isolates where the DpnIII endonuclease is disrupted, and phylogenetic work by Croucher and colleagues suggests that these strains have accumulated genomic differences at a higher rate than other PMEN1 strains. We propose that the R-M locus is a major determinant of genetic acquisition; the resident R-M system governs the extent of genome plasticity. PMID:26081630

  4. Genetic background and epigenetic modifications in the core of the nucleus accumbens predict addiction-like behavior in a rat model

    PubMed Central

    Flagel, Shelly B.; Chaudhury, Sraboni; Waselus, Maria; Kelly, Rebeca; Sewani, Salima; Clinton, Sarah M.; Thompson, Robert C.; Watson, Stanley J.; Akil, Huda

    2016-01-01

    This study provides a demonstration in the rat of a clear genetic difference in the propensity for addiction-related behaviors following prolonged cocaine self-administration. It relies on the use of selectively bred high-responder (bHR) and low-responder (bLR) rat lines that differ in several characteristics associated with “temperament,” including novelty-induced locomotion and impulsivity. We show that bHR rats exhibit behaviors reminiscent of human addiction, including persistent cocaine-seeking and increased reinstatement of cocaine seeking. To uncover potential underlying mechanisms of this differential vulnerability, we focused on the core of the nucleus accumbens and examined expression and epigenetic regulation of two transcripts previously implicated in bHR/bLR differences: fibroblast growth factor (FGF2) and the dopamine D2 receptor (D2). Relative to bHRs, bLRs had lower FGF2 mRNA levels and increased association of a repressive mark on histones (H3K9me3) at the FGF2 promoter. These differences were apparent under basal conditions and persisted even following prolonged cocaine self-administration. In contrast, bHRs had lower D2 mRNA under basal conditions, with greater association of H3K9me3 at the D2 promoter and these differences were no longer apparent following prolonged cocaine self-administration. Correlational analyses indicate that the association of H3K9me3 at D2 may be a critical substrate underlying the propensity to relapse. These findings suggest that low D2 mRNA levels in the nucleus accumbens core, likely mediated via epigenetic modifications, may render individuals more susceptible to cocaine addiction. In contrast, low FGF2 levels, which appear immutable even following prolonged cocaine exposure, may serve as a protective factor. PMID:27114539

  5. Screening and identification of genetic loci involved in producing more/denser inclusion bodies in Escherichia coli

    PubMed Central

    2013-01-01

    Background Many proteins and peptides have been used in therapeutic or industrial applications. They are often produced in microbial production hosts by fermentation. Robust protein production in the hosts and efficient downstream purification are two critical factors that could significantly reduce cost for microbial protein production by fermentation. Producing proteins/peptides as inclusion bodies in the hosts has the potential to achieve both high titers in fermentation and cost-effective downstream purification. Manipulation of the host cells such as overexpression/deletion of certain genes could lead to producing more and/or denser inclusion bodies. However, there are limited screening methods to help to identify beneficial genetic changes rendering more protein production and/or denser inclusion bodies. Results We report development and optimization of a simple density gradient method that can be used for distinguishing and sorting E. coli cells with different buoyant densities. We demonstrate utilization of the method to screen genetic libraries to identify a) expression of glyQS loci on plasmid that increased expression of a peptide of interest as well as the buoyant density of inclusion body producing E. coli cells; and b) deletion of a host gltA gene that increased the buoyant density of the inclusion body produced in the E. coli cells. Conclusion A novel density gradient sorting method was developed to screen genetic libraries. Beneficial host genetic changes could be exploited to improve recombinant protein expression as well as downstream protein purification. PMID:23638724

  6. The genetic regulatory network centered on Pto-Wuschela and its targets involved in wood formation revealed by association studies.

    PubMed

    Yang, Xiaohui; Wei, Zunzheng; Du, Qingzhang; Chen, Jinhui; Wang, Qingshi; Quan, Mingyang; Song, Yuepeng; Xie, Jianbo; Zhang, Deqiang

    2015-11-09

    Transcription factors (TFs) regulate gene expression and can strongly affect phenotypes. However, few studies have examined TF variants and TF interactions with their targets in plants. Here, we used genetic association in 435 unrelated individuals of Populus tomentosa to explore the variants in Pto-Wuschela and its targets to decipher the genetic regulatory network of Pto-Wuschela. Our bioinformatics and co-expression analysis identified 53 genes with the motif TCACGTGA as putative targets of Pto-Wuschela. Single-marker association analysis showed that Pto-Wuschela was associated with wood properties, which is in agreement with the observation that it has higher expression in stem vascular tissues in Populus. Also, SNPs in the 53 targets were associated with growth or wood properties under additive or dominance effects, suggesting these genes and Pto-Wuschela may act in the same genetic pathways that affect variation in these quantitative traits. Epistasis analysis indicated that 75.5% of these genes directly or indirectly interacted Pto-Wuschela, revealing the coordinated genetic regulatory network formed by Pto-Wuschela and its targets. Thus, our study provides an alternative method for dissection of the interactions between a TF and its targets, which will strength our understanding of the regulatory roles of TFs in complex traits in plants.

  7. The genetic regulatory network centered on Pto-Wuschela and its targets involved in wood formation revealed by association studies

    PubMed Central

    Yang, Xiaohui; Wei, Zunzheng; Du, Qingzhang; Chen, Jinhui; Wang, Qingshi; Quan, Mingyang; Song, Yuepeng; Xie, Jianbo; Zhang, Deqiang

    2015-01-01

    Transcription factors (TFs) regulate gene expression and can strongly affect phenotypes. However, few studies have examined TF variants and TF interactions with their targets in plants. Here, we used genetic association in 435 unrelated individuals of Populus tomentosa to explore the variants in Pto-Wuschela and its targets to decipher the genetic regulatory network of Pto-Wuschela. Our bioinformatics and co-expression analysis identified 53 genes with the motif TCACGTGA as putative targets of Pto-Wuschela. Single-marker association analysis showed that Pto-Wuschela was associated with wood properties, which is in agreement with the observation that it has higher expression in stem vascular tissues in Populus. Also, SNPs in the 53 targets were associated with growth or wood properties under additive or dominance effects, suggesting these genes and Pto-Wuschela may act in the same genetic pathways that affect variation in these quantitative traits. Epistasis analysis indicated that 75.5% of these genes directly or indirectly interacted Pto-Wuschela, revealing the coordinated genetic regulatory network formed by Pto-Wuschela and its targets. Thus, our study provides an alternative method for dissection of the interactions between a TF and its targets, which will strength our understanding of the regulatory roles of TFs in complex traits in plants. PMID:26549216

  8. Food Habits, Lifestyle Factors, and Risk of Prostate Cancer in Central Argentina: A Case Control Study Involving Self-Motivated Health Behavior Modifications after Diagnosis

    PubMed Central

    Pacheco, Sandaly O. S.; Pacheco, Fabio J.; Zapata, Gimena M. J.; Garcia, Julieta M. E.; Previale, Carlos A.; Cura, Héctor E.; Craig, Winston J.

    2016-01-01

    Cancer is the second most important non-communicable disease worldwide and disproportionately impacts low- to middle-income countries. Diet in combination with other lifestyle habits seems to modify the risk for some cancers but little is known about South Americans. Food habits of Argentinean men pre- and post-diagnosis of prostate cancer (n = 326) were assessed along with other lifestyle factors. We studied whether any of the behaviors and risk factors for prostate cancer were found in men with other cancers (n = 394), compared with control subjects (n = 629). Before diagnosis, both cases reported a greater mean consumption of meats and fats and lower intakes of fruits, green vegetables, cruciferous vegetables, legumes, nuts, seeds, and whole grains than the controls (all p < 0.001). After diagnosis, cases significantly reduced the intake of meats and fats, and reported other dietary modifications with increased consumption of fish, fruits (including red fruits in prostate cancer), cruciferous vegetables, legumes, nuts, and black tea (all p < 0.001). Additional lifestyle aspects significantly predominant in cases included a reduced quality of sleep, emotional stress, low physical activity, tobacco smoking, alcohol consumption, living in rural areas, and being exposed to environmental contaminants. Argentinian men were predisposed to modify their unhealthy dietary habits and other lifestyle factors after cancer diagnosis. PMID:27409631

  9. Food Habits, Lifestyle Factors, and Risk of Prostate Cancer in Central Argentina: A Case Control Study Involving Self-Motivated Health Behavior Modifications after Diagnosis.

    PubMed

    Pacheco, Sandaly O S; Pacheco, Fabio J; Zapata, Gimena M J; Garcia, Julieta M E; Previale, Carlos A; Cura, Héctor E; Craig, Winston J

    2016-01-01

    Cancer is the second most important non-communicable disease worldwide and disproportionately impacts low- to middle-income countries. Diet in combination with other lifestyle habits seems to modify the risk for some cancers but little is known about South Americans. Food habits of Argentinean men pre- and post-diagnosis of prostate cancer (n = 326) were assessed along with other lifestyle factors. We studied whether any of the behaviors and risk factors for prostate cancer were found in men with other cancers (n = 394), compared with control subjects (n = 629). Before diagnosis, both cases reported a greater mean consumption of meats and fats and lower intakes of fruits, green vegetables, cruciferous vegetables, legumes, nuts, seeds, and whole grains than the controls (all p < 0.001). After diagnosis, cases significantly reduced the intake of meats and fats, and reported other dietary modifications with increased consumption of fish, fruits (including red fruits in prostate cancer), cruciferous vegetables, legumes, nuts, and black tea (all p < 0.001). Additional lifestyle aspects significantly predominant in cases included a reduced quality of sleep, emotional stress, low physical activity, tobacco smoking, alcohol consumption, living in rural areas, and being exposed to environmental contaminants. Argentinian men were predisposed to modify their unhealthy dietary habits and other lifestyle factors after cancer diagnosis. PMID:27409631

  10. Chemical Modification of a Dehydratase Enzyme Involved in Bacterial Virulence by an Ammonium Derivative: Evidence of its Active Site Covalent Adduct.

    PubMed

    González-Bello, Concepción; Tizón, Lorena; Lence, Emilio; Otero, José M; van Raaij, Mark J; Martinez-Guitian, Marta; Beceiro, Alejandro; Thompson, Paul; Hawkins, Alastair R

    2015-07-29

    The first example of an ammonium derivative that causes a specific modification of the active site of type I dehydroquinase (DHQ1), a dehydratase enzyme that is a promising target for antivirulence drug discovery, is described. The resolution at 1.35 Å of the crystal structure of DHQ1 from Salmonella typhi chemically modified by this ammonium derivative revealed that the ligand is covalently attached to the essential Lys170 through the formation of an amine. The detection by mass spectroscopy of the reaction intermediates, in conjunction with the results of molecular dynamics simulations, allowed us to explain the inhibition mechanism and the experimentally observed differences between S. typhi and Staphylococcus aureus enzymes. The results presented here reveal that the replacement of Phe225 in St-DHQ1 by Tyr214 in Sa-DHQ1 and its hydrogen bonding interaction with the conserved water molecule observed in several crystal structures protects the amino adduct against further dehydration/aromatization reactions. In contrast, for the St-DHQ1 enzyme, the carboxylate group of Asp114, with the assistance of this water molecule, would trigger the formation of a Schiff base that can undergo further dehydration reactions until full aromatization of the cyclohexane ring is achieved. Moreover, in vitro antivirulence studies showed that the reported compound is able to reduce the ability of Salmonella Enteritidis to kill A459 respiratory cells. These studies have identified a good scaffold for the design of irreversible inhibitors that can be used as drugs and has opened up new opportunities for the development of novel antivirulence agents by targeting the DHQ1 enzyme.

  11. The activities of lipoprotein lipase and of enzymes involved in triacylglycerol synthesis in rat adipose tissue. Effects of starvation, dietary modification and of corticotropin injection.

    PubMed Central

    Lawson, N; Pollard, A D; Jennings, R J; Gurr, M I; Brindley, D N

    1981-01-01

    1. The effects of dietary modification, including starvation, and of corticotropin injection on the activities of acyl-CoA synthetase, glycerol phosphate acyltransferase, dihydroxyacetone phosphate acyltransferase, phosphatidate phosphohydrolase, diacylglycerol acyltransferase and lipoprotein lipase were measured in adipose tissue. 2. Lipoprotein lipase activities in heart were increased and those in adipose tissue were decreased when rats were fed on diets enriched with corn oil or beef tallow rather than with sucrose or starch. The lipoprotein lipase activity was lower in the adipose tissue of rats fed on the sucrose rather than on the starch diet. 3. Rats fed on the beef tallow diet had slightly higher activities of the total glycerol phosphate acyltransferase in adipose tissue than did rats fed on the sucrose or starch diet. The diacylglycerol acyltransferase and the mitochondrial glycerol phosphate acyltransferase activities were higher for the rats fed on the tallow diet than for those fed on the corn-oil diet. 4. Starvation significantly decreased the activities of lipoprotein lipase (after 24 and 48 h), acyl-CoA synthetase (after 24 h) and of the mitochondrial glycerol phosphate acyltransferase and the N-ethylmaleimide-insensitive dihydroxyacetone phosphate acyltransferase (after 48 h) in adipose tissue. The activities of the microsomal glycerol phosphate acyltransferase, diacylglycerol acyltransferase and the soluble phosphatidate phosphohydrolase were not significantly changed after 24 or 48 h of starvation. 5. The activities of lipoprotein lipase and phosphatidate phosphohydrolase in adipose tissue were decreased 15 min after corticotropin was injected into rats during November to December. No statistically significant differences were found when these experiments were performed during March to September. These differences may be related to the seasonal variation in acute lipolytic responses. 6. These results are discussed in relation to the control of

  12. Exploring Genetic Factors Involved in Huntington Disease Age of Onset: E2F2 as a New Potential Modifier Gene.

    PubMed

    Valcárcel-Ocete, Leire; Alkorta-Aranburu, Gorka; Iriondo, Mikel; Fullaondo, Asier; García-Barcina, María; Fernández-García, José Manuel; Lezcano-García, Elena; Losada-Domingo, José María; Ruiz-Ojeda, Javier; Álvarez de Arcaya, Amaia; Pérez-Ramos, José María; Roos, Raymund A C; Nielsen, Jørgen E; Saft, Carsten; Zubiaga, Ana M; Aguirre, Ana

    2015-01-01

    Age of onset (AO) of Huntington disease (HD) is mainly determined by the length of the CAG repeat expansion (CAGexp) in exon 1 of the HTT gene. Additional genetic variation has been suggested to contribute to AO, although the mechanism by which it could affect AO is presently unknown. The aim of this study is to explore the contribution of candidate genetic factors to HD AO in order to gain insight into the pathogenic mechanisms underlying this disorder. For that purpose, two AO definitions were used: the earliest age with unequivocal signs of HD (earliest AO or eAO), and the first motor symptoms age (motor AO or mAO). Multiple linear regression analyses were performed between genetic variation within 20 candidate genes and eAO or mAO, using DNA and clinical information of 253 HD patients from REGISTRY project. Gene expression analyses were carried out by RT-qPCR with an independent sample of 35 HD patients from Basque Country Hospitals. We found suggestive association signals between HD eAO and/or mAO and genetic variation within the E2F2, ATF7IP, GRIN2A, GRIN2B, LINC01559, HIP1 and GRIK2 genes. Among them, the most significant was the association between eAO and rs2742976, mapping to the promoter region of E2F2 transcription factor. Furthermore, rs2742976 T allele patient carriers exhibited significantly lower lymphocyte E2F2 gene expression, suggesting a possible implication of E2F2-dependent transcriptional activity in HD pathogenesis. Thus, E2F2 emerges as a new potential HD AO modifier factor.

  13. Investigation of genetic variants in ubiquitin enzyme genes involved in the modulation of neurodevelopmental processes: a role in schizophrenia susceptibility?

    PubMed

    Andrews, Jessica L; Fernandez-Enright, Francesca

    2014-11-24

    Despite extensive research during the last few decades, the etiology of schizophrenia remains unclear. Evidence of both genetic and environmental influences in the developmental profile of schizophrenia has grown, and due to the complexity of this disorder, a polygenic aspect has been associated with this neuropsychiatric pathology. Unfortunately, no diagnostic strategies based on biological measurement or genetic testing is currently available for schizophrenia. Gene-expression profiling and recent protein studies have shown a decrease in the expression of ubiquitin pathway proteins in the prefrontal cortex of schizophrenia patients. We have examined single nucleotide polymorphisms (or SNPs) within three genes from the ubiquitin protein system: the ubiquitin conjugating enzyme E2D1 (UBE2D1) gene, the E3 SUMO-protein ligase protein inhibitor of activated STAT 2 (PIAS2) gene, and the E3 ubiquitin ligase F-box and leucine-rich repeat protein 21 (FBXL21) gene, in a Caucasian case-control population for schizophrenia. After Bonferroni correction for multiple testing was applied, no significant associations were reported for any of the tested SNPs. Additional genetic analyses will be necessary to fully explore the role of these three genes in schizophrenia. Regarding the rising interest in ubiquitin-related proteins as a therapeutic target in other pathologies such as cancer, further research into the role of ubiquitin pathways in schizophrenia seems topical and timely.

  14. Genetic Syndromes and Genes Involved in the Development of the Female Reproductive Tract: A Possible Role for Gene Therapy

    PubMed Central

    Connell, MT; Owen, CM; Segars, JH

    2014-01-01

    Müllerian and vaginal anomalies are congenital malformations of the female reproductive tract resulting from alterations in the normal developmental pathway of the uterus, cervix, fallopian tubes, and vagina. The most common of the Müllerian anomalies affect the uterus and may adversely impact reproductive outcomes highlighting the importance of gaining understanding of the genetic mechanisms that govern normal and abnormal development of the female reproductive tract. Modern molecular genetics with study of knock out animal models as well as several genetic syndromes featuring abnormalities of the female reproductive tract have identified candidate genes significant to this developmental pathway. Further emphasizing the importance of understanding female reproductive tract development, recent evidence has demonstrated expression of embryologically significant genes in the endometrium of adult mice and humans. This recent work suggests that these genes not only play a role in the proper structural development of the female reproductive tract but also may persist in adults to regulate proper function of the endometrium of the uterus. As endometrial function is critical for successful implantation and pregnancy maintenance, these recent data suggest a target for gene therapy. Future research will be needed to determine if gene therapy may improve reproductive outcomes for patients with demonstrated deficient endometrial expression related to abnormal gene expression. PMID:25506511

  15. Evaluation of mating systems involving five breeds for integrated beef production systems: IV. Accounting for variability and genetic trends.

    PubMed

    Lamb, M A; Tess, M W; Robison, O W

    1993-03-01

    Computer models were used to simulate integrated cow-calf-feedlot production systems. Angus (A), Charolais (C), Hereford (H), Limousin (L), and Simmental (S) purebreds and two- and three-breed rotational crossbreds were included. Models were deterministic and based on data reported primarily from the 1970s. Variation in carcass weights were determined to predict distributions of carcass weights and values for 272- to 318-kg carcasses. Data were updated to a 1984 base by increasing birth, weaning, yearling, and mature weights to account for genetic trends within breeds. Two slaughter end points were considered: 288-kg carcass weight and low Choice grade. At low Choice grade, accounting for variation in carcass weights around the 272- to 318-kg target weight increased the estimated efficiency of A and AH crosses (input costs/carcass value), whereas at the 288-kg end point, efficiency rankings among other breed combinations were relatively unchanged. Including genetic trends resulted in increased estimated efficiencies among breed combinations with previously underweight carcasses at low Choice (A and AH), measured either as input costs/carcass weight or lean weight values. Within breeds, accounting for genetic trends and variation for weights caused breeds to be ranked differently when evaluated at low Choice.

  16. Modification of tobacco plant development by sense and antisense expression of the tomato viroid-induced AGC VIIIa protein kinase PKV suggests involvement in gibberellin signaling

    PubMed Central

    2009-01-01

    Background The serine-threonine protein kinase gene, designated pkv (protein kinase- viroid induced) was previously found to be transcriptionally activated in tomato plants infected with the plant pathogen Potato spindle tuber viroid (PSTVd). These plants exhibited symptoms of stunting, and abnormal development of leaf, root, and vascular tissues. The encoded protein, PKV, is a novel member of the AGC VIIIa group of signal-transducing protein kinases; however, the role of PKV in plant development is unknown. In this communication, we report the phenotypic results of over expression and silencing of pkv in transgenic tobacco. Results Over expression of pkv in Nicotiana tabacum cv. Xanthi (tobacco) resulted in stunting, reduced root formation, and delay in flowering, phenotypes similar to symptoms of PSTVd infection of tomato. In addition, homozygous T2 tobacco plants over expressing PKV were male sterile. Antisense expression of pkv, on the other hand, resulted in plants that were taller than non-transformed plants, produced an increased number of flowers, and were fertile. Exogenous application of GA3 stimulated stem elongation in the stunted, sense-expressing plants. PKV sense and antisense expression altered transcript levels of GA biosynthetic genes and genes involved in developmental and signaling pathways, but not genes involved in salicylic acid- or jasmonic acid-dependent pathways. Our data provide evidence suggesting that PKV plays an important role in a GA signaling pathway that controls plant height and fertility. Conclusion We have found that the over expression of the tomato protein kinase PKV resulted in stunting, modified vascular tissue development, reduced root formation, and male sterility in tobacco, and we propose that PKV regulates plant development by functioning in critical signaling pathways involved in gibberellic acid metabolism. PMID:19689802

  17. Acclimation of E miliania huxleyi (1516) to nutrient limitation involves precise modification of the proteome to scavenge alternative sources of N and P

    PubMed Central

    Metodieva, Gergana; Raines, Christine A.; Metodiev, Metodi V.; Geider, Richard J.

    2015-01-01

    Summary Limitation of marine primary production by the availability of nitrogen or phosphorus is common. E miliania huxleyi, a ubiquitous phytoplankter that plays key roles in primary production, calcium carbonate precipitation and production of dimethyl sulfide, often blooms in mid‐latitude at the beginning of summer when inorganic nutrient concentrations are low. To understand physiological mechanisms that allow such blooms, we examined how the proteome of E . huxleyi (strain 1516) responds to N and P limitation. We observed modest changes in much of the proteome despite large physiological changes (e.g. cellular biomass, C, N and P) associated with nutrient limitation of growth rate. Acclimation to nutrient limitation did however involve significant increases in the abundance of transporters for ammonium and nitrate under N limitation and for phosphate under P limitation. More notable were large increases in proteins involved in the acquisition of organic forms of N and P, including urea and amino acid/polyamine transporters and numerous C‐N hydrolases under N limitation and a large upregulation of alkaline phosphatase under P limitation. This highly targeted reorganization of the proteome towards scavenging organic forms of macronutrients gives unique insight into the molecular mechanisms that underpin how E . huxleyi has found its niche to bloom in surface waters depleted of inorganic nutrients. PMID:26119724

  18. Acclimation of Emiliania huxleyi (1516) to nutrient limitation involves precise modification of the proteome to scavenge alternative sources of N and P.

    PubMed

    McKew, Boyd A; Metodieva, Gergana; Raines, Christine A; Metodiev, Metodi V; Geider, Richard J

    2015-10-01

    Limitation of marine primary production by the availability of nitrogen or phosphorus is common. Emiliania huxleyi, a ubiquitous phytoplankter that plays key roles in primary production, calcium carbonate precipitation and production of dimethyl sulfide, often blooms in mid-latitude at the beginning of summer when inorganic nutrient concentrations are low. To understand physiological mechanisms that allow such blooms, we examined how the proteome of E. huxleyi (strain 1516) responds to N and P limitation. We observed modest changes in much of the proteome despite large physiological changes (e.g. cellular biomass, C, N and P) associated with nutrient limitation of growth rate. Acclimation to nutrient limitation did however involve significant increases in the abundance of transporters for ammonium and nitrate under N limitation and for phosphate under P limitation. More notable were large increases in proteins involved in the acquisition of organic forms of N and P, including urea and amino acid/polyamine transporters and numerous C-N hydrolases under N limitation and a large upregulation of alkaline phosphatase under P limitation. This highly targeted reorganization of the proteome towards scavenging organic forms of macronutrients gives unique insight into the molecular mechanisms that underpin how E. huxleyi has found its niche to bloom in surface waters depleted of inorganic nutrients. PMID:26119724

  19. Acclimation of Emiliania huxleyi (1516) to nutrient limitation involves precise modification of the proteome to scavenge alternative sources of N and P.

    PubMed

    McKew, Boyd A; Metodieva, Gergana; Raines, Christine A; Metodiev, Metodi V; Geider, Richard J

    2015-10-01

    Limitation of marine primary production by the availability of nitrogen or phosphorus is common. Emiliania huxleyi, a ubiquitous phytoplankter that plays key roles in primary production, calcium carbonate precipitation and production of dimethyl sulfide, often blooms in mid-latitude at the beginning of summer when inorganic nutrient concentrations are low. To understand physiological mechanisms that allow such blooms, we examined how the proteome of E. huxleyi (strain 1516) responds to N and P limitation. We observed modest changes in much of the proteome despite large physiological changes (e.g. cellular biomass, C, N and P) associated with nutrient limitation of growth rate. Acclimation to nutrient limitation did however involve significant increases in the abundance of transporters for ammonium and nitrate under N limitation and for phosphate under P limitation. More notable were large increases in proteins involved in the acquisition of organic forms of N and P, including urea and amino acid/polyamine transporters and numerous C-N hydrolases under N limitation and a large upregulation of alkaline phosphatase under P limitation. This highly targeted reorganization of the proteome towards scavenging organic forms of macronutrients gives unique insight into the molecular mechanisms that underpin how E. huxleyi has found its niche to bloom in surface waters depleted of inorganic nutrients.

  20. Genetic variability of transcript abundance in pig peri-mortem skeletal muscle: eQTL localized genes involved in stress response, cell death, muscle disorders and metabolism

    PubMed Central

    2011-01-01

    Background The genetics of transcript-level variation is an exciting field that has recently given rise to many studies. Genetical genomics studies have mainly focused on cell lines, blood cells or adipose tissues, from human clinical samples or mice inbred lines. Few eQTL studies have focused on animal tissues sampled from outbred populations to reflect natural genetic variation of gene expression levels in animals. In this work, we analyzed gene expression in a whole tissue, pig skeletal muscle sampled from individuals from a half sib F2 family shortly after slaughtering. Results QTL detection on transcriptome measurements was performed on a family structured population. The analysis identified 335 eQTLs affecting the expression of 272 transcripts. The ontologic annotation of these eQTLs revealed an over-representation of genes encoding proteins involved in processes that are expected to be induced during muscle development and metabolism, cell morphology, assembly and organization and also in stress response and apoptosis. A gene functional network approach was used to evidence existing biological relationships between all the genes whose expression levels are influenced by eQTLs. eQTLs localization revealed a significant clustered organization of about half the genes located on segments of chromosome 1, 2, 10, 13, 16, and 18. Finally, the combined expression and genetic approaches pointed to putative cis-drivers of gene expression programs in skeletal muscle as COQ4 (SSC1), LOC100513192 (SSC18) where both the gene transcription unit and the eQTL affecting its expression level were shown to be localized in the same genomic region. This suggests cis-causing genetic polymorphims affecting gene expression levels, with (e.g. COQ4) or without (e.g. LOC100513192) potential pleiotropic effects that affect the expression of other genes (cluster of trans-eQTLs). Conclusion Genetic analysis of transcription levels revealed dependence among molecular phenotypes as being

  1. [HLA-DQA1 genes involved in genetic susceptibility to rheumatic fever and rheumatic heart disease in southern Hans].

    PubMed

    Gu, J; Yu, B; Zhou, J

    1997-05-01

    The incidence of rheumatic fever (RF) or rheumatic heart disease (RHD) is high in southern China. We studied the genetic susceptibility of HLA-DQA1 alleles to RF or RHD with emphasis on the mechanisms 106 unrelated healthy individuals and 54 patients with RF or RHD of Chinese Han nationality in Guangdong were included. DNA extraction from various biological material using phenol/chloroform method and HLA-DQA1 genotyping by PCR-PAGE and then with silver dyeing was used to show the electrophoretic patterns. A total 6 alleles of HLA-DQA1 were found. Increased allele frequencies of DQA1*0101 (31.48%, RR = 2.89, P < 0.005, EF = 0.206) and decreased allele frequencies of DQA1*0102 (1.85%, RR = 0.106, P < 0.005, PF = 0.134) were observed. Two increased genotyping of HLA-DQA1 (DQA1*0101/0301, chi 2 = 8.84, P < 0.005 and DQA1*0101/0401, chi 2 = 6.23, P < 0.0025) and decreased genotyping of DQA1*0102/0301 (chi 2 = 11.98, P < 0.005) were also observed. These findings suggested that DQA1*0101 contribute to genetic susceptibility for RF or RHD in Guangdong hans while DQA1*0102 to its genetic resistance. Digesting the genotyping of HLA-DQA1 may provide scientific basis for finding susceptible individuals to RF or RHD. Using PCR-PAGE and silver dyeing technique, a new genotyping method for HLA-DQA1, which is simple sensitive and precise, was established and applied. PMID:10374271

  2. Development of a genetic system for Marinobacter adhaerens HP15 involved in marine aggregate formation by interacting with diatom cells.

    PubMed

    Sonnenschein, Eva C; Gärdes, Astrid; Seebah, Shalin; Torres-Monroy, Ingrid; Grossart, Hans-Peter; Ullrich, Matthias S

    2011-11-01

    Diatom aggregation is substantial for organic carbon flux from the photic zone to deeper waters. Many heterotrophic bacteria ubiquitously found in diverse marine environments interact with marine algae and thus impact organic matter and energy cycling in the ocean. In particular, Marinobacter adhaerens HP15 induces aggregate formation while interacting with the diatom, Thalassiosira weissflogii. To study this effect at the molecular level, a genetic tool system was developed for strain HP15. The antibiotic susceptibility spectrum of this organism was determined and electroporation and conjugation protocols were established. Among various plasmids of different incompatibility groups, only two were shown to replicate in M. adhaerens. 1.4×10(-3) transconjugants per recipient were obtained for a broad-host-range vector. Electroporation efficiency corresponded to 1.1×10(5)CFU per μg of DNA. Transposon and gene-specific mutageneses were conducted for flagellum biosynthetic genes. Mutant phenotypes were confirmed by swimming assay and microscopy. Successful expression of two reporter genes in strain HP15 revealed useful tools for gene expression analyses, which will allow studying diverse bacteria-algae interactions at the molecular level and hence to gain a mechanistic understanding of micro-scale processes underlying ocean basin-scale processes. This study is the first report for the genetic manipulation of a Marinobacter species which specifically interacts with marine diatoms and serves as model to additionally analyze various previously reported Marinobacter-algae interactions in depth. PMID:21880271

  3. Genetic screens for factors involved in the notum bristle loss of interspecific hybrids between Drosophila melanogaster and D. simulans.

    PubMed

    Takano-Shimizu, T

    2000-09-01

    Interspecific cross is a powerful means to uncover hidden within- and between-species variation in populations. One example is a bristle loss phenotype of hybrids between Drosophila melanogaster and D. simulans, although both the pure species have exactly the same pattern of bristle formation on the notum. There exists a large amount of genetic variability in the simulans populations with respect to the number of missing bristles in hybrids, and the variation is largely attributable to simulans X chromosomes. Using nine molecular markers, I screened the simulans X chromosome for genetic factors that were responsible for the differences between a pair of simulans lines with high (H) and low (L) missing bristle numbers. Together with duplication-rescue experiments, a single major quantitative locus was mapped to a 13F-14F region. Importantly, this region accounted for most of the differences between H and L lines in three other independent pairs, suggesting segregation of H and L alleles at the single locus in different populations. Moreover, a deficiency screening uncovered several regions with factors that potentially cause the hybrid bristle loss due to epistatic interactions with the other factors.

  4. Development of a genetic system for Marinobacter adhaerens HP15 involved in marine aggregate formation by interacting with diatom cells.

    PubMed

    Sonnenschein, Eva C; Gärdes, Astrid; Seebah, Shalin; Torres-Monroy, Ingrid; Grossart, Hans-Peter; Ullrich, Matthias S

    2011-11-01

    Diatom aggregation is substantial for organic carbon flux from the photic zone to deeper waters. Many heterotrophic bacteria ubiquitously found in diverse marine environments interact with marine algae and thus impact organic matter and energy cycling in the ocean. In particular, Marinobacter adhaerens HP15 induces aggregate formation while interacting with the diatom, Thalassiosira weissflogii. To study this effect at the molecular level, a genetic tool system was developed for strain HP15. The antibiotic susceptibility spectrum of this organism was determined and electroporation and conjugation protocols were established. Among various plasmids of different incompatibility groups, only two were shown to replicate in M. adhaerens. 1.4×10(-3) transconjugants per recipient were obtained for a broad-host-range vector. Electroporation efficiency corresponded to 1.1×10(5)CFU per μg of DNA. Transposon and gene-specific mutageneses were conducted for flagellum biosynthetic genes. Mutant phenotypes were confirmed by swimming assay and microscopy. Successful expression of two reporter genes in strain HP15 revealed useful tools for gene expression analyses, which will allow studying diverse bacteria-algae interactions at the molecular level and hence to gain a mechanistic understanding of micro-scale processes underlying ocean basin-scale processes. This study is the first report for the genetic manipulation of a Marinobacter species which specifically interacts with marine diatoms and serves as model to additionally analyze various previously reported Marinobacter-algae interactions in depth.

  5. Genetic evidence that Ras-like GTPases, Gtr1p, and Gtr2p, are involved in epigenetic control of gene expression in Saccharomyces cerevisiae

    SciTech Connect

    Sekiguchi, Takeshi Hayashi, Naoyuki; Wang, Yonggang; Kobayashi, Hideki

    2008-04-11

    Gtr1p and Gtr2p of Saccharomyces cerevisiae are members of the Ras-like GTP binding family and interact genetically with Prp20p (yeast RCC1), which is a guanine nucleotide exchange factor for Gsp1p (yeast homolog of Ran, involved in nuclear export). Recently, Gtr1p and Gtr2p were suggested to be molecular switches in the rapamycin-sensitive TOR signaling pathway. Here, we show that Gtr1p and Gtr2p genetically interact with the chromatin remodeling factor Ino80p. Gtr2p interacted physically with both Rvb1p and Rvb2p. Consistent with these results, Gtr2p localized to chromatin and could activate transcription. Gtr1p and Gtr2p were found to be involved in chromatin silencing in the vicinity of telomeres. Gtr1p and Gtr2p were required to repress nitrogen catabolite-repressed genes, which are repressed by the TOR signaling pathway. We propose that Gtr1p and Gtr2p are involved in epigenetic control of gene expression in the TOR signaling pathway.

  6. Harnessing epigenome modifications for better crops

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Chemical DNA modifications such as methylation influence translation of the DNA code to specific genetic outcomes. While such modifications can be heritable, others are transient, and their overall contribution to plant genetic diversity remains intriguing but uncertain. The focus of this article is...

  7. Reductive activation in periplasmic nitrate reductase involves chemical modifications of the Mo-cofactor beyond the first coordination sphere of the metal ion.

    PubMed

    Jacques, Julien G J; Fourmond, Vincent; Arnoux, Pascal; Sabaty, Monique; Etienne, Emilien; Grosse, Sandrine; Biaso, Frédéric; Bertrand, Patrick; Pignol, David; Léger, Christophe; Guigliarelli, Bruno; Burlat, Bénédicte

    2014-02-01

    In Rhodobacter sphaeroides periplasmic nitrate reductase NapAB, the major Mo(V) form (the "high g" species) in air-purified samples is inactive and requires reduction to irreversibly convert into a catalytically competent form (Fourmond et al., J. Phys. Chem., 2008). In the present work, we study the kinetics of the activation process by combining EPR spectroscopy and direct electrochemistry. Upon reduction, the Mo (V) "high g" resting EPR signal slowly decays while the other redox centers of the protein are rapidly reduced, which we interpret as a slow and gated (or coupled) intramolecular electron transfer between the [4Fe-4S] center and the Mo cofactor in the inactive enzyme. Besides, we detect spin-spin interactions between the Mo(V) ion and the [4Fe-4S](1+) cluster which are modified upon activation of the enzyme, while the EPR signatures associated to the Mo cofactor remain almost unchanged. This shows that the activation process, which modifies the exchange coupling pathway between the Mo and the [4Fe-4S](1+) centers, occurs further away than in the first coordination sphere of the Mo ion. Relying on structural data and studies on Mo-pyranopterin and models, we propose a molecular mechanism of activation which involves the pyranopterin moiety of the molybdenum cofactor that is proximal to the [4Fe-4S] cluster. The mechanism implies both the cyclization of the pyran ring and the reduction of the oxidized pterin to give the competent tricyclic tetrahydropyranopterin form.

  8. A targeted next-generation sequencing assay for the molecular diagnosis of genetic disorders with orodental involvement

    PubMed Central

    Prasad, Megana K; Geoffroy, Véronique; Vicaire, Serge; Jost, Bernard; Dumas, Michael; Le Gras, Stéphanie; Switala, Marzena; Gasse, Barbara; Laugel-Haushalter, Virginie; Paschaki, Marie; Leheup, Bruno; Droz, Dominique; Dalstein, Amelie; Loing, Adeline; Grollemund, Bruno; Muller-Bolla, Michèle; Lopez-Cazaux, Séréna; Minoux, Maryline; Jung, Sophie; Obry, Frédéric; Vogt, Vincent; Davideau, Jean-Luc; Davit-Beal, Tiphaine; Kaiser, Anne-Sophie; Moog, Ute; Richard, Béatrice; Morrier, Jean-Jacques; Duprez, Jean-Pierre; Odent, Sylvie; Bailleul-Forestier, Isabelle; Rousset, Monique Marie; Merametdijan, Laure; Toutain, Annick; Joseph, Clara; Giuliano, Fabienne; Dahlet, Jean-Christophe; Courval, Aymeric; El Alloussi, Mustapha; Laouina, Samir; Soskin, Sylvie; Guffon, Nathalie; Dieux, Anne; Doray, Bérénice; Feierabend, Stephanie; Ginglinger, Emmanuelle; Fournier, Benjamin; de la Dure Molla, Muriel; Alembik, Yves; Tardieu, Corinne; Clauss, François; Berdal, Ariane; Stoetzel, Corinne; Manière, Marie Cécile; Dollfus, Hélène; Bloch-Zupan, Agnès

    2016-01-01

    Background Orodental diseases include several clinically and genetically heterogeneous disorders that can present in isolation or as part of a genetic syndrome. Due to the vast number of genes implicated in these disorders, establishing a molecular diagnosis can be challenging. We aimed to develop a targeted next-generation sequencing (NGS) assay to diagnose mutations and potentially identify novel genes mutated in this group of disorders. Methods We designed an NGS gene panel that targets 585 known and candidate genes in orodental disease. We screened a cohort of 101 unrelated patients without a molecular diagnosis referred to the Reference Centre for Oro-Dental Manifestations of Rare Diseases, Strasbourg, France, for a variety of orodental disorders including isolated and syndromic amelogenesis imperfecta (AI), isolated and syndromic selective tooth agenesis (STHAG), isolated and syndromic dentinogenesis imperfecta, isolated dentin dysplasia, otodental dysplasia and primary failure of tooth eruption. Results We discovered 21 novel pathogenic variants and identified the causative mutation in 39 unrelated patients in known genes (overall diagnostic rate: 39%). Among the largest subcohorts of patients with isolated AI (50 unrelated patients) and isolated STHAG (21 unrelated patients), we had a definitive diagnosis in 14 (27%) and 15 cases (71%), respectively. Surprisingly, COL17A1 mutations accounted for the majority of autosomal-dominant AI cases. Conclusions We have developed a novel targeted NGS assay for the efficient molecular diagnosis of a wide variety of orodental diseases. Furthermore, our panel will contribute to better understanding the contribution of these genes to orodental disease. Trial registration numbers NCT01746121 and NCT02397824. PMID:26502894

  9. Genetic ablation of caspase-7 promotes solar-simulated light-induced mouse skin carcinogenesis: the involvement of keratin-17.

    PubMed

    Lee, Mee-Hyun; Lim, Do Young; Kim, Myoung Ok; Lee, Sung-Young; Shin, Seung Ho; Kim, Jae Young; Kim, Sung-Hyun; Kim, Dong Joon; Jung, Sung Keun; Yao, Ke; Kundu, Joydeb Kumar; Lee, Hye Suk; Lee, Cheol-Jung; Dickinson, Sally E; Alberts, David; Bowden, G Timothy; Stratton, Steven; Curiel, Clara; Einspahr, Janine; Bode, Ann M; Surh, Young-Joon; Cho, Yong-Yeon; Dong, Zigang

    2015-11-01

    Solar ultraviolet irradiation is an environmental carcinogen that causes skin cancer. Caspase-7 is reportedly expressed at reduced levels in many cancers. The present study was designed to examine the role of caspase-7 in solar-simulated light (SSL)-induced skin cancer and to elucidate its underlying molecular mechanisms. Our study revealed that mice with genetic deficiency of caspase-7 are highly susceptible to SSL-induced skin carcinogenesis. Epidermal hyperplasia, tumor volume and the average number of tumors were significantly increased in caspase-7 knockout (KO) mice compared with SKH1 wild-type mice irradiated with SSL. The expression of cell proliferation markers, such as survivin and Ki-67, was elevated in SSL-irradiated skin of caspase-7 KO mice compared with those observed in SSL-exposed wild-type SKH1 mouse skin. Moreover, SSL-induced apoptosis was abolished in skin from caspase-7 KO mice. Two-dimensional gel electrophoresis, followed by matrix-assisted laser desorption/ionization-time-of-flight analysis of skin tissue lysates from SSL-irradiated SKH1 wild-type and caspase-7 KO mice revealed an aberrant induction of keratin-17 in caspase-7 KO mice. Immunohistochemical analysis of skin tumors also showed an increase of keratin-17 expression in caspase-7 KO mice compared with SKH1 wild-type mice. The expression of keratin-17 was also elevated in SSL-irradiated caspase-7 KO keratinocytes as well as in human basal cell carcinomas. The in vitro caspase activity assay showed keratin-17 as a substrate of caspase-7, but not caspase-3. Overall, our study demonstrates that genetic loss of caspase-7 promotes SSL-induced skin carcinogenesis by blocking caspase-7-mediated cleavage of keratin-17. PMID:26271098

  10. STAT3 activation by cytokines utilizing gp130 and related transducers involves a secondary modification requiring an H7-sensitive kinase.

    PubMed Central

    Boulton, T G; Zhong, Z; Wen, Z; Darnell, J E; Stahl, N; Yancopoulos, G D

    1995-01-01

    Ciliary neurotrophic factor, oncostatin M, leukemia-inhibitory factor, and interleukin 6 are related cytokines that initiate signaling by homodimerizing the signal-transducing receptor component gp130 or by heterodimerizing gp130 with a gp130-related receptor component. Receptor dimerization in turn activates receptor-associated kinases of the Jak/Tyk family, resulting in the rapid tyrosine phosphorylation of several intracellular proteins, including those of two members of the signal transducers and activators of transcription (STAT) family--STAT1 and STAT3. Here we show that all cytokines that utilize gp130 sequentially induce two distinct forms of STAT3 in all responding cells examined, with the two forms apparently differing because of a time-dependent secondary serine/threonine phosphorylation involving an H7-sensitive kinase. While both STAT3 forms bind DNA and translocate to the nucleus, the striking time-dependent progression from one form to the other implies other important functional differences between the two forms. Granulocyte colony-stimulating factor, which utilizes a receptor highly related to gp130, also induces these two forms of STAT3. In contrast to a number of other cytokines and growth factors, all cytokines using gp130 and related signal transducers consistently and preferentially induce the two forms of STAT3 as compared with STAT1; this characteristic STAT activation pattern is seen regardless of which Jak/Tyk kinases are used in a particular response, consistent with the notion that the receptor components themselves are the primary determinants of which STATs are activated. Images Fig. 1 Fig. 2 Fig. 3 PMID:7624343

  11. Structural and Functional Characterization of PseC, an Aminotransferase Involved in the Biosynthesis of Pseudaminic Acid, an Essential Flagellar Modification in Helicobacter Pylori

    SciTech Connect

    Schoenhofen,I.; Lunin, V.; Julien, J.; Li, Y.; Ajamian, E.; Matte, A.; Cygler, M.; Brisson, J.; Aubry, A.; et al.

    2006-01-01

    Helicobacter pylori flagellin is heavily glycosylated with the novel sialic acid-like nonulosonate, pseudaminic acid (Pse). The glycosylation process is essential for assembly of functional flagellar filaments and consequent bacterial motility. As motility is a key virulence factor for this and other important pathogens, the Pse biosynthetic pathway offers potential for novel therapeutic targets. From recent NMR analyses, we determined that the conversion of UDP-a-D-GlcNAc to the central intermediate in the pathway, UDP-4-amino-4,6-dideoxy-{beta}-L-AltNAc, proceeds by formation of UDP-2-acetamido-2,6-dideoxy-{beta}-L-arabino-4-hexulose by the dehydratase/epimerase PseB (HP0840) followed with amino transfer by the aminotransferase, PseC (HP0366). The central role of PseC in the H. pylori Pse biosynthetic pathway prompted us to determine crystal structures of the native protein, its complexes with pyridoxal phosphate alone and in combination with the UDP-4-amino-4,6-dideoxy-{beta}-L-AltNAc product, the latter being converted to the external aldimine form in the enzyme's active site. In the binding site, the AltNAc sugar ring adopts a 4C1 chair conformation which is different from the predominant 1C4 form found in solution. The enzyme forms a homodimer where each monomer contributes to the active site, and these structures have permitted the identification of key residues involved in stabilization, and possibly catalysis, of the {beta}-L-arabino intermediate during the amino transfer reaction. The essential role of Lys183 in the catalytic event was confirmed by site-directed mutagenesis. This work presents for the first time a nucleotide-sugar aminotransferase co-crystallized with its natural ligand, and in conjunction with the recent functional characterization of this enzyme, will assist in elucidating the aminotransferase reaction mechanism within the Pse biosynthetic pathway.

  12. The effect of communicating the genetic risk of cardiometabolic disorders on motivation and actual engagement in preventative lifestyle modification and clinical outcome: a systematic review and meta-analysis of randomised controlled trials.

    PubMed

    Li, Sherly X; Ye, Zheng; Whelan, Kevin; Truby, Helen

    2016-09-01

    Genetic risk prediction of chronic conditions including obesity, diabetes and CVD currently has limited predictive power but its potential to engage healthy behaviour change has been of immense research interest. We aimed to understand whether the latter is indeed true by conducting a systematic review and meta-analysis investigating whether genetic risk communication affects motivation and actual behaviour change towards preventative lifestyle modification. We included all randomised controlled trials (RCT) since 2003 investigating the impact of genetic risk communication on health behaviour to prevent cardiometabolic disease, without restrictions on age, duration of intervention or language. We conducted random-effects meta-analyses for perceived motivation for behaviour change and clinical changes (weight loss) and a narrative analysis for other outcomes. Within the thirteen studies reviewed, five were vignette studies (hypothetical RCT) and seven were clinical RCT. There was no consistent effect of genetic risk on actual motivation for weight loss, perceived motivation for dietary change (control v. genetic risk group standardised mean difference (smd) -0·15; 95 % CI -1·03, 0·73, P=0·74) or actual change in dietary behaviour. Similar results were observed for actual weight loss (control v. high genetic risk SMD 0·29 kg; 95 % CI -0·74, 1·31, P=0·58). This review found no clear or consistent evidence that genetic risk communication alone either raises motivation or translates into actual change in dietary intake or physical activity to reduce the risk of cardiometabolic disorders in adults. Of thirteen studies, eight were at high or unclear risk of bias. Additional larger-scale, high-quality clinical RCT are warranted. PMID:27405704

  13. The effect of communicating the genetic risk of cardiometabolic disorders on motivation and actual engagement in preventative lifestyle modification and clinical outcome: a systematic review and meta-analysis of randomised controlled trials.

    PubMed

    Li, Sherly X; Ye, Zheng; Whelan, Kevin; Truby, Helen

    2016-09-01

    Genetic risk prediction of chronic conditions including obesity, diabetes and CVD currently has limited predictive power but its potential to engage healthy behaviour change has been of immense research interest. We aimed to understand whether the latter is indeed true by conducting a systematic review and meta-analysis investigating whether genetic risk communication affects motivation and actual behaviour change towards preventative lifestyle modification. We included all randomised controlled trials (RCT) since 2003 investigating the impact of genetic risk communication on health behaviour to prevent cardiometabolic disease, without restrictions on age, duration of intervention or language. We conducted random-effects meta-analyses for perceived motivation for behaviour change and clinical changes (weight loss) and a narrative analysis for other outcomes. Within the thirteen studies reviewed, five were vignette studies (hypothetical RCT) and seven were clinical RCT. There was no consistent effect of genetic risk on actual motivation for weight loss, perceived motivation for dietary change (control v. genetic risk group standardised mean difference (smd) -0·15; 95 % CI -1·03, 0·73, P=0·74) or actual change in dietary behaviour. Similar results were observed for actual weight loss (control v. high genetic risk SMD 0·29 kg; 95 % CI -0·74, 1·31, P=0·58). This review found no clear or consistent evidence that genetic risk communication alone either raises motivation or translates into actual change in dietary intake or physical activity to reduce the risk of cardiometabolic disorders in adults. Of thirteen studies, eight were at high or unclear risk of bias. Additional larger-scale, high-quality clinical RCT are warranted.

  14. New challenges and promises in solid organ transplantation pharmacogenetics: the genetic variability of proteins involved in the pharmacodynamics of immunosuppressive drugs.

    PubMed

    Pouché, Lucie; Stojanova, Jana; Marquet, Pierre; Picard, Nicolas

    2016-02-01

    Interindividual variability in immunosuppressive drug responses might be partly explained by genetic variants in proteins involved in the immune response or associated with IS pharmacodynamics. On a general basis, the pharmacogenetics of drug target proteins is less known and understood than that of proteins involved in drug disposition pathways. The aim of this review is to facilitate research related to the pharmacodynamics of the main immunosuppressive drugs used in solid organ transplantation. We elaborated a quality of evidence grading system based on a literature review and identified 'highly recommended', 'recommended' or 'potential' candidates for further research. It is likely that a number of additional rare variants might further explain drug response phenotypes in transplantation, and particularly the most severe ones. The advent of next-generation sequencing will help to identify those variants.

  15. Home Modification

    MedlinePlus

    ... it is important to consider certain safety modifications. Adaptations such as those in the following list can ... The importance of a Consumer Perspective in Home Adaptation of Alzheimer’s Households” (Chapter 6 pp 91-112) ...

  16. Linkage analysis of the genetic determinants of high density lipoprotein concentrations and composition: evidence for involvement of the apolipoprotein A-II and cholesteryl ester transfer protein loci.

    PubMed

    Bu, X; Warden, C H; Xia, Y R; De Meester, C; Puppione, D L; Teruya, S; Lokensgard, B; Daneshmand, S; Brown, J; Gray, R J

    1994-06-01

    We have tested for evidence of linkage between the genetic loci determining concentrations and composition of plasma high density lipoproteins (HDL) with the genes for the major apolipoproteins and enzymes participating in lipoprotein metabolism. These genes include those encoding various apolipoproteins (apo), including apoA-I, apoA-II, apoA-IV, apoB, apoC-I, apoC-II, apoC-III, apoE, and apo(a), cholesteryl ester transfer protein (CETP), HDL-binding protein, lipoprotein lipase, and the low density lipoprotein (LDL) receptor. Polymorphisms of these genes, and nearby highly polymorphic simple sequence repeat markers, were examined by quantitative sib-pair linkage analysis in 30 coronary artery disease families consisting of a total of 366 individuals. Evidence for linkage was observed between a marker locus D16S313 linked to the CETP locus and a locus determining plasma HDL-cholesterol concentration (P = 0.002), and the genetic locus for apoA-II and a locus determining the levels of the major apolipoproteins of HDL, apoA-I and apoA-II (P = 0.009 and 0.02, respectively). HDL level was also influenced by the variation at the apo(a) locus on chromosome 6 (P = 0.02). Thus, these data indicate the simultaneous involvement of at least two different genetic loci in the determination of the levels of HDL and its associated lipoproteins.

  17. Epigenetic modification of OXT and human sociability.

    PubMed

    Haas, Brian W; Filkowski, Megan M; Cochran, R Nick; Denison, Lydia; Ishak, Alexandra; Nishitani, Shota; Smith, Alicia K

    2016-07-01

    Across many mammalian species there exist genetic and biological systems that facilitate the tendency to be social. Oxytocin is a neuropeptide involved in social-approach behaviors in humans and others mammals. Although there exists a large, mounting body of evidence showing that oxytocin signaling genes are associated with human sociability, very little is currently known regarding the way the structural gene for oxytocin (OXT) confers individual differences in human sociability. In this study, we undertook a comprehensive approach to investigate the association between epigenetic modification of OXT via DNA methylation, and overt measures of social processing, including self-report, behavior, and brain function and structure. Genetic data were collected via saliva samples and analyzed to target and quantify DNA methylation across the promoter region of OXT We observed a consistent pattern of results across sociability measures. People that exhibit lower OXT DNA methylation (presumably linked to higher OXT expression) display more secure attachment styles, improved ability to recognize emotional facial expressions, greater superior temporal sulcus activity during two social-cognitive functional MRI tasks, and larger fusiform gyrus gray matter volume than people that exhibit higher OXT DNA methylation. These findings provide empirical evidence that epigenetic modification of OXT is linked to several overt measures of sociability in humans and serve to advance progress in translational social neuroscience research toward a better understanding of the evolutionary and genetic basis of normal and abnormal human sociability. PMID:27325757

  18. Epigenetic modification of OXT and human sociability.

    PubMed

    Haas, Brian W; Filkowski, Megan M; Cochran, R Nick; Denison, Lydia; Ishak, Alexandra; Nishitani, Shota; Smith, Alicia K

    2016-07-01

    Across many mammalian species there exist genetic and biological systems that facilitate the tendency to be social. Oxytocin is a neuropeptide involved in social-approach behaviors in humans and others mammals. Although there exists a large, mounting body of evidence showing that oxytocin signaling genes are associated with human sociability, very little is currently known regarding the way the structural gene for oxytocin (OXT) confers individual differences in human sociability. In this study, we undertook a comprehensive approach to investigate the association between epigenetic modification of OXT via DNA methylation, and overt measures of social processing, including self-report, behavior, and brain function and structure. Genetic data were collected via saliva samples and analyzed to target and quantify DNA methylation across the promoter region of OXT We observed a consistent pattern of results across sociability measures. People that exhibit lower OXT DNA methylation (presumably linked to higher OXT expression) display more secure attachment styles, improved ability to recognize emotional facial expressions, greater superior temporal sulcus activity during two social-cognitive functional MRI tasks, and larger fusiform gyrus gray matter volume than people that exhibit higher OXT DNA methylation. These findings provide empirical evidence that epigenetic modification of OXT is linked to several overt measures of sociability in humans and serve to advance progress in translational social neuroscience research toward a better understanding of the evolutionary and genetic basis of normal and abnormal human sociability.

  19. The EDGE Hypothesis: Epigenetically Directed Genetic Errors in Repeat-Containing Proteins (RCPs) Involved in Evolution, Neuroendocrine Signaling, and Cancer

    PubMed Central

    Ruden, Douglas M.; Jamison, D. Curtis; Zeeberg, Barry R.; Garfinkel, Mark D.; Weinstein, John N.; Rasouli, Parsa; Lu, Xiangyi

    2009-01-01

    Trans-generational epigenetic phenomena, such as endocrine-disrupting chemicals (EDCs) that decrease fertility and the global methylation status of DNA in the offspring, are of great concern because they may affect the health of our children. However, of even greater concern is the possibility that trans-generational changes in the methylation status of the DNA might lead to permanent changes in the DNA sequence itself. By contaminating the environment with EDCs, mankind might be permanently affecting the health of future generations. In this chapter, we present evidence from our laboratory and others that trans-generational epigenetic changes in DNA might lead to mutations directed to genes encoding amino acid repeat-containing proteins (RCPs) that are important for adaptive evolution or cancer progression. Such epigenetic changes can be induced “naturally” by hormones or “unnaturally” by EDCs or environmental stress. To illustrate the phenomenon, we present new bioinformatic evidence that the only RCP ontological categories conserved from Drosophila to humans are “regulation of splicing,” “regulation of transcription,” and “regulation of synaptogenesis,” which are precisely the classes of genes that are important for evolutionary processes. Based on that and other evidence, we propose a model for evolution that we call the EDGE (Epigenetically Directed Genetic Errors) hypothesis for the mechanism by which mutations are targeted at epigenetically-modified “contingency genes” encoding RCPs. In the model, “epigenetic assimilation” of metastable epialleles of RCPs over many generations can lead to mutations directed to those genes, thereby permanently stabilizing the adaptive phenotype. PMID:18295320

  20. Genetic regulation analysis reveals involvement of tumor necrosis factor and alpha-induced protein 3 in stress response in mice.

    PubMed

    Xu, Jian; Dai, Aihua; Chen, Qi; Liu, Xiaorong; Zhang, Yu; Wang, Hongmei; Li, Haizhen; Chen, Ying; Cao, Maohong

    2016-01-15

    In order to study whether Tnfaip3 is related to stress response and further to find it's genetic regulation, we use C57BL/6J (B6) and DBA/2 (D2) mice to built the model of chronic unpredictable mild stress. RT-PCR, Western blotting and immunohistochemistry were used for studying the expression variation of Tnfaip3 in hippocampus tissue of B6 and D2 mice after being stressed. We found that the expression of Tnfaip3 was more remarkably increased in chronic unpredictable stress models than that in untreated mice (P<0.05). It is indicated that Tnfaip3 might take part in the process of stress response. The expression of Tnfaip3 is regulated by a cis-acting quantitative trait locus (cis-eQTL). We identified 5 genes are controlled by Tnfaip3 and the expression of 64 genes highly associated with Tnfaip3, 9 of those have formerly been participate in stress related pathways. In order to estimate the relationship between Tnfaip3 and its downstream genes or network members, we transfected SH-SY5Y cells with Tnfaip3 siRNA leading to down-regulation of Tnfaip3 mRNA. We confirmed a significant influence of Tnfaip3 depletion on the expression of Tsc22d3, Pex7, Rap2a, Slc2a3, and Gap43. These validated downstream genes and members of Tnfaip3 gene network provide us new insight into the biological mechanisms of Tnfaip3 in chronic unpredictable stress. PMID:26546835

  1. Assembly of the mitochondrial membrane system. XVIII. Genetic loci on mitochondrial DNA involved in cytochrome b biosynthesis.

    PubMed

    Tzagoloff, A; Foury, F; Akai, A

    1976-11-24

    1. Fourteen cytoplasmic mutants of Saccharomyces cerevisiae with a specific deficiency of cytochrome b have been studied. The mutations have been shown to occur in two separate genetic loci, COB 1 and COB 2. These loci can be distinguished by mit- X mit- crosses. Pairwise crosses of cytochrome b mutants belonging to different loci yield 4-6% wild type recombinants corresponding to recombinational frequencies of 8-12%. In intra-locus crosses, the recombinational frequencies range from 1% to less than 0.01%. The two loci can also be distinguished by mit- X rho- crosses. Twenty rho- testers have been isolated of which ten preferentially restore mutations in COB 1 and ten others in COB 2. 2. The COB 1 and COB 2 loci have been localized on mitochondrial DNA between the two antibiotic resistance loci OLI 1 and OLI 2 in the order OLI 2-COB 2-COB 1-OLI 1. The results of mit- X mit- and mit- X rho- crosses have also been used to map the cytochrome b mutations relative to each other. The maps obtained by the two independent methods are in good agreement. 3. Mutations in COB 1 have been found to be linked to the OLI1 locus in some but not in other strains of S. cervisiae. This evidence suggests that there may be a spacer region between the two loci whose length varies from strain to strain. 4. Two mutations in COB 2 have been found to cause a loss of a mitochondrial translation product corresponding to the cytochrome b apoprotein. Instead of the wild type protein the mutants have a new low-molecular weight product which is probably a fragment of cytochrome b. The fact that the mutations revert suggests that they are nonsense mutations in the structural gene of cytochrome b. PMID:796670

  2. Genetic organization and transcriptional analysis of a major gene cluster involved in siderophore biosynthesis in Pseudomonas putida WCS358.

    PubMed Central

    Marugg, J D; Nielander, H B; Horrevoets, A J; van Megen, I; van Genderen, I; Weisbeek, P J

    1988-01-01

    In iron-limited environments, the plant-growth-stimulating Pseudomonas putida WCS358 produces a yellow-green fluorescent siderophore called pseudobactin 358. The transcriptional organization and the iron-regulated expression of a major gene cluster involved in the biosynthesis and transport of pseudobactin 358 were analyzed in detail. The cluster comprises a region with a minimum length of 33.5 kilobases and contains at least five transcriptional units, of which some are relatively large. The directions of transcription of four transcriptional units were determined by RNA-RNA hybridization and by analysis in Escherichia coli minicells. The latter also demonstrated that large polypeptides were encoded by these transcriptional units. The results allowed us to localize several promoter regions on the DNA. The iron-dependent expression of at least two genes within this cluster appears to be regulated at the transcriptional level. Images PMID:2450869

  3. A complex genetic switch involving overlapping divergent promoters and DNA looping regulates expression of conjugation genes of a gram-positive plasmid.

    PubMed

    Ramachandran, Gayetri; Singh, Praveen K; Luque-Ortega, Juan Roman; Yuste, Luis; Alfonso, Carlos; Rojo, Fernando; Wu, Ling J; Meijer, Wilfried J J

    2014-10-01

    Plasmid conjugation plays a significant role in the dissemination of antibiotic resistance and pathogenicity determinants. Understanding how conjugation is regulated is important to gain insights into these features. Little is known about regulation of conjugation systems present on plasmids from Gram-positive bacteria. pLS20 is a native conjugative plasmid from the Gram-positive bacterium Bacillus subtilis. Recently the key players that repress and activate pLS20 conjugation have been identified. Here we studied in detail the molecular mechanism regulating the pLS20 conjugation genes using both in vivo and in vitro approaches. Our results show that conjugation is subject to the control of a complex genetic switch where at least three levels of regulation are integrated. The first of the three layers involves overlapping divergent promoters of different strengths regulating expression of the conjugation genes and the key transcriptional regulator RcoLS20. The second layer involves a triple function of RcoLS20 being a repressor of the main conjugation promoter and an activator and repressor of its own promoter at low and high concentrations, respectively. The third level of regulation concerns formation of a DNA loop mediated by simultaneous binding of tetrameric RcoLS20 to two operators, one of which overlaps with the divergent promoters. The combination of these three layers of regulation in the same switch allows the main conjugation promoter to be tightly repressed during conditions unfavorable to conjugation while maintaining the sensitivity to accurately switch on the conjugation genes when appropriate conditions occur. The implications of the regulatory switch and comparison with other genetic switches involving DNA looping are discussed. PMID:25340403

  4. [Lifestyle modifications].

    PubMed

    Kawano, Yuhei

    2015-11-01

    Lifestyle modifications are important in the prevention and treatment of hypertension. The Japanese Society of Hypertension Guidelines for the Management of Hypertension (JSH2014) recommend salt reduction (< 6 g/day), increased intake of vegetables/fruit and fish (fish oil), reduced intake of cholesterol and saturated fatty acids, weight loss (body mass index < 25kg/m2), exercise (≥ 30 min/day), reduction of alcohol intake (≤ 20-30 mL/day in men, ≤ 10-20 mL/day in women as ethanol), and quitting smoking. These lifestyle modifications are capable of reducing blood pressure and ameliorating other cardiovascular risk factors. However, the reduction in blood pressure is mild to moderate and the adherence to lifestyle modifications has been still suboptimal. PMID:26619658

  5. Community involvement in the ethical review of genetic research: lessons from American Indian and Alaska Native populations.

    PubMed Central

    Sharp, Richard R; Foster, Morris W

    2002-01-01

    The National Bioethics Advisory Commission has proposed that regulatory oversight for research with human subjects be extended beyond the protection of individual research participants to include the protection of social groups. To accomplish this, the commission recommends that investigators and ethics review boards a) work directly with community representatives to develop study methods that minimize potential group harms, b) discuss group implications as part of the informed consent process, and c) consider group harms in reporting research results. We examine the utility of these recommendations in the context of research with American Indian and Alaska Native communities. Because much attention has been given to the question of how best to consult with members of these communities in the design and conduct of research, we believe it behooves investigators to consider the lessons to be learned from research involving American Indians and Alaska Natives. After describing several difficulties surrounding the application of the commission's approach to these research contexts, we propose a research agenda to develop best practices for working with local communities in the ethical assessment of epidemiologic and environmental health research. PMID:11929722

  6. Genetic analysis of regions of the Lactococcus lactis subsp. lactis plasmid pRS01 involved in conjugative transfer.

    PubMed Central

    Mills, D A; Choi, C K; Dunny, G M; McKay, L L

    1994-01-01

    The genes responsible for conjugative transfer of the 48.4-kb Lactococcus lactis subsp. lactis ML3 plasmid pRS01 were localized by insertional mutagenesis. Integration of the IS946-containing plasmid pTRK28 into pRS01 generated a pool of stable cointegrates, including a number of plasmids altered in conjugative proficiency. Mapping of pTRK28 insertions and phenotypic analysis of cointegrate plasmids identified four distinct regions (Tra1, Tra2, Tra3, and Tra4) involved in pRS01 conjugative transfer. Tra3 corresponds closely to a region previously identified (D. G. Anderson and L. L. McKay, J. Bacteriol. 158:954-962, 1984). Another region (Tra4) was localized within an inversion sequence shown to correlate with a cell aggregation phenotype. Tra1 and Tra2, two previously unidentified regions, were located at a distance of 9 kb from Tra3. When provided in trans, a cloned portion of the Tra3 region complemented Tra3 mutants. PMID:7811081

  7. [Genetics and genetic counseling].

    PubMed

    Izzi, Claudia; Liut, Francesca; Dallera, Nadia; Mazza, Cinzia; Magistroni, Riccardo; Savoldi, Gianfranco; Scolari, Francesco

    2016-01-01

    Autosomal Dominant Polycystic Kidney Disease (ADPKD) is the most frequent genetic disease, characterized by progressive development of bilateral renal cysts. Two causative genes have been identified: PKD1 and PKD2. ADPKD phenotype is highly variable. Typically, ADPKD is an adult onset disease. However, occasionally, ADPKD manifests as very early onset disease. The phenotypic variability of ADPKD can be explained at three genetic levels: genic, allelic and gene modifier effects. Recent advances in molecular screening for PKD gene mutations and the introduction of the new next generation sequencing (NGS)- based genotyping approach have generated considerable improvement regarding the knowledge of genetic basis of ADPKD. The purpose of this article is to provide a comprehensive review of the genetics of ADPKD, focusing on new insights in genotype-phenotype correlation and exploring novel clinical approach to genetic testing. Evaluation of these new genetic information requires a multidisciplinary approach involving a nephrologist and a clinical geneticist. PMID:27067213

  8. Chemical strategies for the covalent modification of filamentous phage

    PubMed Central

    Bernard, Jenna M. L.; Francis, Matthew B.

    2014-01-01

    Historically filamentous bacteriophage have been known to be the workhorse of phage display due to their ability to link genotype to phenotype. More recently, the filamentous phage scaffold has proven to be powerful outside the realm of phage display technology in fields such as molecular imaging, cancer research and materials, and vaccine development. The ability of the virion to serve as a platform for a variety of applications heavily relies on the functionalization of the phage coat proteins with a wide variety of functionalities. Genetic modification of the coat proteins has been the most widely used strategy for functionalizing the virion; however, complementary chemical modification strategies can help to diversify the range of materials that can be developed. This review emphasizes the recent advances that have been made in the chemical modification of filamentous phage as well as some of the challenges that are involved in functionalizing the virion. PMID:25566240

  9. Isolation of Polymyxin B-Susceptible Mutants of Burkholderia pseudomallei and Molecular Characterization of Genetic Loci Involved in Polymyxin B Resistance

    PubMed Central

    Burtnick, Mary N.; Woods, Donald E.

    1999-01-01

    Burkholderia pseudomallei is a gram-negative bacterium that causes the disease known as melioidosis. This pathogen is endemic to Southeast Asia and northern Australia and is particularly problematic in northeastern Thailand. It has been previously reported that B. pseudomallei is resistant to the killing action of cationic antimicrobial peptides, including human neutrophil peptide, protamine sulfate, poly-l-lysine, magainins, and polymyxins. Recently, we have also found that the virulent clinical isolate B. pseudomallei 1026b is capable of replicating in media containing polymyxin B at concentrations of >100 mg/ml. In order to identify genetic loci that are associated with this particular resistance phenotype, we employed a Tn5-OT182 mutagenesis system in coordination with a replica plating screen to isolate polymyxin B-susceptible mutants. Of the 17,000 Tn5-OT182 mutants screened via this approach, five polymyxin B-susceptible mutants were obtained. Three of these mutants harbored Tn5-OT182 insertions within a genetic locus demonstrating strong homology to the lytB gene present in other gram-negative bacteria. Of the remaining two mutants, one contained a transposon insertion in a locus involved in lipopolysaccharide core biosynthesis (waaF), while the other contained an insertion in an open reading frame homologous to UDP-glucose dehydrogenase genes. Isogenic mutants were also constructed via allelic exchange and used in complementation analysis studies to further characterize the relative importance of each of the various genetic loci with respect to the polymyxin B resistance phenotype exhibited by B. pseudomallei 1026b. PMID:10543742

  10. Defined chromosomal assignment of CLN5 demonstrates that at least four genetic loci are involved in the pathogenesis of human ceroid lipofuscinoses

    SciTech Connect

    Savukoski, M.; Peltonen, L.; Santavuori, P.; Kestilae, M. |; Williams, R.; Jaervelae, I.; Sharp, J.; Harris, J.; Gardiner, M.

    1994-10-01

    We demonstrate here that at least four genetically separate loci are involved in the pathogenesis of human neuronal ceroid lipofuscinoses (NCLs), fatal brain disorders of children. Earlier the assignments of the infantile and juvenile subtypes of NCL to 1p32 and 16p12 had revealed two loci; and here a variant subtype of the late-infantile form of NCL is mapped to a well-defined region on 13q21.1-q32, whereas the clinically similar, classical form of late-infantile NCL was found to represent the fourth, yet-unidentified NCL locus. The linkage disequilibrium was crucial for locus assignment in our highly limited family material, and the data exemplify the significance of this phenomenon in the random mapping of rare human diseases. 22 refs., 4 figs., 3 tabs.

  11. Readers of PCNA modifications.

    PubMed

    Ulrich, Helle D; Takahashi, Tomio

    2013-08-01

    The eukaryotic sliding clamp, proliferating cell nuclear antigen (PCNA), acts as a central coordinator of DNA transactions by providing a multivalent interaction surface for factors involved in DNA replication, repair, chromatin dynamics and cell cycle regulation. Posttranslational modifications (PTMs), such as mono- and polyubiquitylation, sumoylation, phosphorylation and acetylation, further expand the repertoire of PCNA's binding partners. These modifications affect PCNA's activity in the bypass of lesions during DNA replication, the regulation of alternative damage processing pathways such as homologous recombination and DNA interstrand cross-link repair, or impact on the stability of PCNA itself. In this review, we summarise our current knowledge about how the PTMs are "read" by downstream effector proteins that mediate the appropriate action. Given the variety of interaction partners responding to PCNA's modified forms, the ensemble of PCNA modifications serves as an instructive model for the study of biological signalling through PTMs in general. PMID:23580141

  12. Genetic analyses involving microsatellite ETH10 genotypes on bovine chromosome 5 and performance trait measures in Angus- and Brahman-influenced cattle.

    PubMed

    DeAtley, K L; Rincon, G; Farber, C R; Medrano, J F; Luna-Nevarez, P; Enns, R M; VanLeeuwen, D M; Silver, G A; Thomas, M G

    2011-07-01

    ETH10 is a dinucleotide microsatellite within the promoter of signal transducer and activator of transcription 6 (STAT6) gene on bovine chromosome 5. ETH10 is included in the panel of genetic markers used in parentage testing procedures of cattle breed associations. Allelic sizes of ETH10 PCR amplicons range from 199 to 225 bp. Objectives of this study were to use microsatellite data from beef cattle breed associations to investigate genetic distance and population stratification among Angus- and Brahman-influenced cattle and to use ETH10 genotypes and growth and ultrasound carcass data to investigate their statistical relationships. Three series of genotype to phenotype association analyses were conducted with 1) Angus data (n=5,094), 2) Brangus data (3/8 Brahman × 5/8 Angus; n=2,296), and 3) multibreed data (n=4,426) of Angus and Brangus cattle. Thirteen alleles and 38 genotypes were observed, but frequencies varied among breed groups. Tests of genetic identity and distance among 6 breed composition groups increasing in Brahman influence from 0 to 75% revealed that as Brahman-influence increased to ≥50%, genetic distance from Angus ranged from 18.3 to 43.5%. This was accomplished with 10 microsatellite loci. A mixed effects model involving genotype as a fixed effect and sire as a random source of variation suggested that Angus cattle with the 217/219 genotype tended to have 2.1% heavier (P=0.07) 205-d BW than other genotypes. In Brangus cattle, allele combinations were classified as small (≤215 bp) or large (≥217 bp). Brangus cattle with the small/large genotype had 2.0% heavier (P<0.05) birth weight, yet cattle with the large/large genotype had approximately 5.1% greater (P<0.05) percentage of fat within LM and more LM per BW than cattle with small/large or small/small genotypes. Genotype-to-phenotype relationships were not detected in multibreed analyses. The ETH10 locus appears to be associated with growth and carcass traits in Angus and Brangus cattle

  13. Genetic analyses involving microsatellite ETH10 genotypes on bovine chromosome 5 and performance trait measures in Angus- and Brahman-influenced cattle.

    PubMed

    DeAtley, K L; Rincon, G; Farber, C R; Medrano, J F; Luna-Nevarez, P; Enns, R M; VanLeeuwen, D M; Silver, G A; Thomas, M G

    2011-07-01

    ETH10 is a dinucleotide microsatellite within the promoter of signal transducer and activator of transcription 6 (STAT6) gene on bovine chromosome 5. ETH10 is included in the panel of genetic markers used in parentage testing procedures of cattle breed associations. Allelic sizes of ETH10 PCR amplicons range from 199 to 225 bp. Objectives of this study were to use microsatellite data from beef cattle breed associations to investigate genetic distance and population stratification among Angus- and Brahman-influenced cattle and to use ETH10 genotypes and growth and ultrasound carcass data to investigate their statistical relationships. Three series of genotype to phenotype association analyses were conducted with 1) Angus data (n=5,094), 2) Brangus data (3/8 Brahman × 5/8 Angus; n=2,296), and 3) multibreed data (n=4,426) of Angus and Brangus cattle. Thirteen alleles and 38 genotypes were observed, but frequencies varied among breed groups. Tests of genetic identity and distance among 6 breed composition groups increasing in Brahman influence from 0 to 75% revealed that as Brahman-influence increased to ≥50%, genetic distance from Angus ranged from 18.3 to 43.5%. This was accomplished with 10 microsatellite loci. A mixed effects model involving genotype as a fixed effect and sire as a random source of variation suggested that Angus cattle with the 217/219 genotype tended to have 2.1% heavier (P=0.07) 205-d BW than other genotypes. In Brangus cattle, allele combinations were classified as small (≤215 bp) or large (≥217 bp). Brangus cattle with the small/large genotype had 2.0% heavier (P<0.05) birth weight, yet cattle with the large/large genotype had approximately 5.1% greater (P<0.05) percentage of fat within LM and more LM per BW than cattle with small/large or small/small genotypes. Genotype-to-phenotype relationships were not detected in multibreed analyses. The ETH10 locus appears to be associated with growth and carcass traits in Angus and Brangus cattle

  14. Functional validation of Capsicum frutescens aminotransferase gene involved in vanillylamine biosynthesis using Agrobacterium mediated genetic transformation studies in Nicotiana tabacum and Capsicum frutescens calli cultures.

    PubMed

    Gururaj, Harishchandra B; Padma, Mallaya N; Giridhar, Parvatam; Ravishankar, Gokare A

    2012-10-01

    Capsaicinoid biosynthesis involves the participation of two substrates viz. vanillylamine and C(9)-C(11) fatty acid moieties. Vanillylamine which is a derivative of vanillin is synthesized through a transaminase reaction in the phenylpropanoid pathway of capsaicinoid synthesis. Here we report the functional validation of earlier reported putative aminotransferase gene for vanillylamine biosynthesis in heterologous system using Agrobacterium mediated genetic transformation studies in Nicotiana tabacum and Capsicum frutescens calli cultures. Molecular analysis tools comprising PCR and Southern blot analysis have shown the integration of the foreign gene in N. tabacum and C. frutescens calli cultures. The study shows the production of vanillylamine in transformed N. tabacum callus cultures and also the reduction of vanillylamine production when whole gene based antisense binary vector construct was used in transformation of C. frutescens callus cultures. Vanillylamine production, aminotransferase assay with Western blot analysis for crude proteins of transformants established the production of putative aminotransferase (pAMT) protein in alternate plant. The result is a clear evidence of involvement of the reported putative aminotransferase responsible for vanillylamine biosynthesis in capsaicinoid biosynthesis pathway, confirming the gene function through functional validation.

  15. [Combination of Genetic and Humanitarian (Cross-Cultural) Methods for the Identification of Human Genes Involved in the Process of Adaptation to Evolutionary New Environmental Factors].

    PubMed

    Borinskaya, S A; Yankovsky, N K

    2015-04-01

    Human settlement from the African ancestral home was accompanied by cultural and genetic adaptation to new habitat conditions (climate, infections, diet, etc.). We previously suggested for the first time an approach to the identification of human genes presumably involved in adaptation to evolutionary new environmental factors based on a combination of genetic and humanitarian methods of study. In order to search for the genes involved in adaptation and for environmental factors (to which this adaptation occurs), we attempted to find correlations between the population allele frequencies of the studied gene and formalized descriptions of peculiarities of the habitat of ethnic groups given in "Ethnographic Atlas" by G. P. Murdock. In the presented review, we summarized our own data on an experimental determination of the allele frequencies for lactase (LCT*), apolipoprotein E (APOE), and alcohol dehydrogenase (ADH1B) genes in populations of Russia. Based on these data and available materials of other investigators, we developed maps of worldwide allele frequency distribution for these genes. We detected a correlation of allele frequencies of these genes in populations with the presence of certain factors of the environment that these populations inhabit. It was also confirmed that the evolutionarily young LCT*-13910T allele, which determines lactase persistence and the possibility of milk consumption in adults, is distributed in populations for which dairy animal husbandry is typical. During the analysis of 68 populations, we for the first time demonstrated that the frequency of the APOE e4 allele (which is ancestral for humans and influences the lipid metabolism) is higher in groups with a high contribution of hunting and gathering. Our data are in favor of the hypothesis that it was exactly the e4 allele that was a subject for selection, while the e3 allele was less important for adaptation. We also for the first time demonstrated that the evolutionarily young ADH

  16. [Combination of Genetic and Humanitarian (Cross-Cultural) Methods for the Identification of Human Genes Involved in the Process of Adaptation to Evolutionary New Environmental Factors].

    PubMed

    Borinskaya, S A; Yankovsky, N K

    2015-04-01

    Human settlement from the African ancestral home was accompanied by cultural and genetic adaptation to new habitat conditions (climate, infections, diet, etc.). We previously suggested for the first time an approach to the identification of human genes presumably involved in adaptation to evolutionary new environmental factors based on a combination of genetic and humanitarian methods of study. In order to search for the genes involved in adaptation and for environmental factors (to which this adaptation occurs), we attempted to find correlations between the population allele frequencies of the studied gene and formalized descriptions of peculiarities of the habitat of ethnic groups given in "Ethnographic Atlas" by G. P. Murdock. In the presented review, we summarized our own data on an experimental determination of the allele frequencies for lactase (LCT*), apolipoprotein E (APOE), and alcohol dehydrogenase (ADH1B) genes in populations of Russia. Based on these data and available materials of other investigators, we developed maps of worldwide allele frequency distribution for these genes. We detected a correlation of allele frequencies of these genes in populations with the presence of certain factors of the environment that these populations inhabit. It was also confirmed that the evolutionarily young LCT*-13910T allele, which determines lactase persistence and the possibility of milk consumption in adults, is distributed in populations for which dairy animal husbandry is typical. During the analysis of 68 populations, we for the first time demonstrated that the frequency of the APOE e4 allele (which is ancestral for humans and influences the lipid metabolism) is higher in groups with a high contribution of hunting and gathering. Our data are in favor of the hypothesis that it was exactly the e4 allele that was a subject for selection, while the e3 allele was less important for adaptation. We also for the first time demonstrated that the evolutionarily young ADH

  17. Chemical modification of tyrosine-38 in p-hydroxybenzoate hydroxylase from Pseudomonas fluorescens by 5'-p-fluorosulfonylbenzoyladenosine: a probe for the elucidation of the NADPH binding site? Involvement in catalysis, assignment in sequence and fitting to the tertiary structure.

    PubMed

    van Berkel, W J; Müller, F; Jekel, P A; Weijer, W J; Schreuder, H A; Wierenga, R K

    1988-09-15

    p-Hydroxybenzoate hydroxylase from Pseudomonas fluorescens was covalently modified by the nucleotide analog 5'-(p-fluorosulfonylbenzoyl)-adenosine in the presence of 20% dimethylsulfoxide. The inactivation reaction is pH-dependent and does not obey pseudo-first-order kinetics, due to spontaneous hydrolysis of the reagent. The kinetic data further indicate that a weak, reversible enzyme-inhibitor complex is an intermediate in the inactivation reaction and that only one amino acid residue is responsible for the loss of activity. The inactivation is strongly inhibited by NADPH and 2',5'ADP. Steady-state kinetics and 2',5'ADP bioaffinity chromatography of the modified enzyme suggest that the essential residue is not directly involved in NADPH binding. Sequence studies show that Tyr-38 is the main residue protected from modification in the presence of NADPH. From crystallographic studies it is known that the hydroxyl group of Tyr-38 is 1.84 nm away from the active site. Model-building studies using computer graphics show that this distance can be accommodated when FSO2BzAdo binds in an extended conformation with the sulfonylbenzoyl portion in an orientation different from the nicotin-amide ring of NADPH. PMID:3138119

  18. Involvement of waaY, waaQ, and waaP in the modification of Escherichia coli lipopolysaccharide and their role in the formation of a stable outer membrane.

    PubMed

    Yethon, J A; Heinrichs, D E; Monteiro, M A; Perry, M B; Whitfield, C

    1998-10-01

    The waaY, waaQ, and waaP genes are located in the central operon of the waa (formerly rfa) locus on the chromosome of Escherichia coli. This locus contains genes whose products are involved in the assembly of the core region of the lipopolysaccharide molecule. In the R1 core prototype strain, E. coli F470, there are nine genes in this operon, and all but waaY, waaQ, and waaP have been assigned function. In this study, the waaY, waaQ, and waaP genes were independently mutated by insertion of a non-polar antibiotic resistance cassette, and the structures of the resulting mutant core oligosaccharides were determined by chemical analyses and phosphorus-nuclear magnetic resonance spectroscopy. All three of these mutations were shown to affect the modification of the heptose region of the core, a region whose structure is critical to outer membrane stability. Mutation of waaY resulted in a core oligosaccharide devoid of phosphate on HepII. Mutation of waaQ resulted in loss of the branch HepIII residue on HepII and impeded the activity of WaaY. Mutation of waaP resulted in loss of phosphoryl substituents on HepI and obviated WaaQ and WaaY activity. Only mutation of waaP resulted in hypersensitivity to novobiocin and sodium dodecyl sulfate, a characteristic of deep-rough mutations.

  19. A formulation of the foundations of genetics and evolution.

    PubMed

    Bahr, Brian Edward

    2016-05-01

    This paper proposes a formulation of theories of the foundations of genetics and evolution that can be used to mathematically simulate phenotype expression, reproduction, mutation, and natural selection. It will be shown that Mendelian inheritance can be mathematically simulated with expressions involving matrices and that these expressions can also simulate phenomena that are modifications to Mendel's basic principles, like alleles that give rise to quantitative effects and traits that are the expression of multiple alleles and/or multiple genetic loci. PMID:26947775

  20. Characterization of the dapA-nlpB genetic locus involved in regulation of swarming motility, cell envelope architecture, hemolysin production, and cell attachment ability in Serratia marcescens.

    PubMed

    Soo, Po-Chi; Wei, Jun-Rong; Horng, Yu-Tze; Hsieh, Shang-Chen; Ho, Shen-Wu; Lai, Hsin-Chih

    2005-09-01

    Swarming migration of Serratia marcescens requires both flagellar motility and cellular differentiation and is a population-density-dependent behavior. While the flhDC and quorum-sensing systems have been characterized as important factors regulating S. marcescens swarming, the underlying molecular mechanisms are currently far from being understood. Serratia swarming is thermoregulated and is characterized by continuous surface migration on rich swarming agar surfaces at 30 degrees C but not at 37 degrees C. To further elucidate the mechanisms, identification of specific and conserved regulators that govern the initiation of swarming is essential. We performed transposon mutagenesis to screen for S. marcescens strain CH-1 mutants that swarmed at 37 degrees C. Analysis of a "precocious-swarming" mutant revealed that the defect in a conserved dapA(Sm)-nlpB(Sm) genetic locus which is closely related to the synthesis of bacterial cell wall peptidoglycan is responsible for the aberrant swarming phenotype. Further complementation and gene knockout studies showed that nlpB(Sm), which encodes a membrane lipoprotein, NlpB(Sm), but not dapA(Sm), is specifically involved in swarming regulation. On the other hand, dapA(Sm) but not nlpB(Sm) is responsible for the determination of cell envelope architecture, regulation of hemolysin production, and cellular attachment capability. While the nlpB(Sm) mutant showed similar cytotoxicity to its parent strain, the dapA(Sm) mutant significantly increased in cytotoxicity. We present evidence that DapA(Sm) is involved in the determination of cell-envelope-associated phenotypes and that NlpB(Sm) is involved in the regulation of swarming motility.

  1. Involvement of endocannabinoids in alcohol “binge” drinking: studies of mice with human fatty acid amide hydrolase genetic variation and after CB1 receptor antagonists

    PubMed Central

    Zhou, Yan; Huang, Ted; Lee, Francis; Kreek, Mary Jeanne

    2016-01-01

    Background The endocannabinoid system has been found to play an important role in modulating alcohol intake. Inhibition or genetic deletion of fatty acid amide hydrolase (FAAH, a key catabolic enzyme for endocannabinoids) leads to increased alcohol consumption and preference in rodent models. A common human single-nucleotide polymorphism (SNP; C385A, rs324420) in the FAAH gene is associated with decreased enzymatic activity of FAAH, resulting in increased anandamide levels in both humans and FAAH C385A knock-in mice. Methods As this FAAH SNP has been reported to be associated with altered alcohol abuse, the present study used these genetic knock-in mice containing the human SNP C385A to determine the impact of variant FAAH gene on alcohol “binge” drinking in the drinking-in-the-dark (DID) model. Results We found that the FAAHA/A mice had greater alcohol intake and preference than the wild-type FAAHC/C mice, suggesting that increased endocannabinoid signaling in FAAHA/A mice led to increased alcohol “binge” consumption. The specificity on alcohol vulnerability was suggested by the lack of any FAAH genotype difference on sucrose or saccharin intake. Using the “binge” DID model, we confirmed that selective CB1 receptor antagonist AM251 reduced alcohol intake in the wild-type mice. Conclusions These data suggest that there is direct and selective involvement of the human FAAH C385A SNP and CB1 receptors in alcohol “binge” drinking. PMID:26857901

  2. Either the dorsal hippocampus or the dorsolateral striatum is selectively involved in consolidation of forced swim-induced immobility depending on genetic background.

    PubMed

    Colelli, V; Campus, P; Conversi, D; Orsini, C; Cabib, S

    2014-05-01

    Healthy subjects differ in the memory system they engage to learn dual-solution tasks. Both genotype and stress experience could contribute to this phenotypic variability. The present experiments tested whether the hippocampus and the dorsal striatum, the core nodes of two different memory systems, are differently involved in 24 h retention of a stress-associated memory in two genetically unrelated inbred strains of mice. Mice from both the C57BL/6J and the DBA/2J inbred strains showed progressive increase of immobility during 10 min exposure to forced swim (FS) and retrieved the acquired levels of immobility when tested 24h later. The pattern of c-fos immunostaining promoted by FS revealed activation of a large number of brain areas in both strains, including CA1 and CA3 fields of the hippocampus. However, only DBA/2J mice showed activation of the dorsolateral striatum (DLS). In addition, FS induced a positive correlation between c-fos expression in the amygdala and CA1 and CA3 in C57BL/6J mice whereas it induced a positive correlation between c-fos expression in the amygdala and DLS in DBA/2J mice. Finally, temporary post-training inactivation of the dorsal hippocampus, by local infusion of lidocaine, prevented 24h retention of immobility in C57BL/6J mice only, whereas inactivation of the DLS prevented retention in DBA/2J mice only. These findings support the view that genetic factors can determine whether the dorsal hippocampus or the DLS are selectively engaged to consolidate stress-related memory.

  3. To Test or Not to Test? The Role of Attitudes, Knowledge, and Religious Involvement among U.s. Adults on Intent-to-Obtain Adult Genetic Testing

    ERIC Educational Resources Information Center

    Botoseneanu, Anda; Alexander, Jeffrey A.; Banaszak-Holl, Jane

    2011-01-01

    Genetic testing can advance cancer prevention if current screening behaviors improve. Increased prevalence of high-risk genotypes within specific religious groups, use of religious venues for recruiting to genetic screening, and ethical-religious considerations argue for exploring the role of religiosity in forming genetic testing decisions. This…

  4. Oligo/Polynucleotide-Based Gene Modification: Strategies and Therapeutic Potential

    PubMed Central

    Sargent, R. Geoffrey; Kim, Soya

    2011-01-01

    Oligonucleotide- and polynucleotide-based gene modification strategies were developed as an alternative to transgene-based and classical gene targeting-based gene therapy approaches for treatment of genetic disorders. Unlike the transgene-based strategies, oligo/polynucleotide gene targeting approaches maintain gene integrity and the relationship between the protein coding and gene-specific regulatory sequences. Oligo/polynucleotide-based gene modification also has several advantages over classical vector-based homologous recombination approaches. These include essentially complete homology to the target sequence and the potential to rapidly engineer patient-specific oligo/polynucleotide gene modification reagents. Several oligo/polynucleotide-based approaches have been shown to successfully mediate sequence-specific modification of genomic DNA in mammalian cells. The strategies involve the use of polynucleotide small DNA fragments, triplex-forming oligonucleotides, and single-stranded oligodeoxynucleotides to mediate homologous exchange. The primary focus of this review will be on the mechanistic aspects of the small fragment homologous replacement, triplex-forming oligonucleotide-mediated, and single-stranded oligodeoxynucleotide-mediated gene modification strategies as it relates to their therapeutic potential. PMID:21417933

  5. Black Psychologists Discuss Behavior Modification.

    ERIC Educational Resources Information Center

    Bardo, Harold R.; And Others

    The primary purpose of this paper is to discuss reasons why blacks should be concerned and actively involved with practices in behavior modification. The concern is that as these techniques are refined it becomes more important to be sure blacks should be involved at all levels of the application of these procedures when other blacks are subjects…

  6. Uniparental disomy for chromosome 6 results in steroid 21-hydroxylase deficiency: evidence of different genetic mechanisms involved in the production of the disease.

    PubMed Central

    López-Gutiérrez, A U; Riba, L; Ordoñez-Sánchez, M L; Ramírez-Jiménez, S; Cerrillo-Hinojosa, M; Tusié-Luna, M T

    1998-01-01

    Congenital adrenal hyperplasia (CAH) is an inherited recessive disorder of adrenal steroidogenesis caused by mutations in the steroid 21-hydroxylase gene (CYP21) in more than 90% of affected patients. The CYP21 gene is located within the HLA complex locus on chromosome 6 (6p21.3). During a molecular characterisation study of a group of 47 Mexican families with 21-hydroxylase deficiency, we identified nine in which the mutation or mutations found in the patient did not appear to originate from one of the parents. Through DNA fingerprinting, paternity was established in all nine families with a probability of non-paternity in the range of 10(-19) to 10(-23). Among these families, we identified one patient with exclusive paternal inheritance of all eight markers tested on chromosome 6p, despite normal maternal and paternal contributions for eight additional markers on three different chromosomes. We did not identify duplication of paternal information for markers in the 6q region, consistent with lack of expression of transient neonatal diabetes owing to genomic imprinting in this patient. Our results substantiate evidence for the existence of different genetic mechanisms involved in the expression of this recessive condition in a substantial portion (approximately 19%) of affected Mexican families. In addition to the identification of a patient with paternal uniparental disomy, the occurrence of germline mutations may explain the unusual pattern of segregation in the majority of the remaining eight families. PMID:9863599

  7. Analysis of the genetic potential and gene expression of microbial communities involved in the in situ bioremediation of uranium and harvesting electrical energy from organic matter.

    PubMed

    Lovley, Derek R

    2002-01-01

    The proposed research will investigate two microbial communities that are of direct relevance to Department of Energy interests. One is the microbial community associated with the in situ bioremediation of uranium-contaminated groundwater. The second is a microbial community that harvests energy from waste organic matter in the form of electricity. These studies will address DOE needs for (1) remediation of metals and radionuclides at DOE sites and (2) the development of cleaner forms of energy and biomass conversion to energy. Our previous studies have demonstrated that the microbial communities involved in uranium bioremediation and energy harvesting are both dominated by microorganisms in the family Geobacteraceae and that the organisms in this family are responsible for uranium bioremediation and electron transfer to electrodes. The initial objectives of this study are to (1) describe the genetic potential of the Geobacteraceae that predominate in the environments of interest; (2) identify conserved patterns of gene expression within the Geobacteraceae family in response to a range of environmental conditions; (3) begin to identify mechanisms controlling the expression of key genes related to survival, growth, and activity in subsurface environments and on electrodes; and (4) use the results from subobjectives 1-3 to develop a conceptual model for predicting gene expression of Geobacteraceae in the environments of interest. This will serve as the basis for a subsequent simulation model of the growth and activity of Geobacteraceae in the subsurface and on electrodes.

  8. Characterization and genetic mapping of modA. A mutation in the post-translational modification of the glycosidases of Dictyostelium discoideum.

    PubMed

    Free, S J; Schimke, R T; Freeze, H; Loomis, W F

    1978-06-25

    We have isolated a mutant of Dictyostelium discoideum, M31, which produces a reduced number of alpha-mannosidase-1 molecules per cell during the developmental program of the organism. We find that several of the glycosidases, a group of lysosomal proteins produced by D. discoideum, are altered in strain M31 and that this strain produces a reduced level of at least three of these activities. These enzymes do not share a common protein subunit but are known to share a common antigenic determinant which is, in part, carbohydrate in nature. In the wild type parent of M31, alpha-mannosidase-1 is modified by the addition of mannose and glucosamine (probably as N-acetylglucosamine) in the molar ratio of 5:2. alpha-Mannosidase-1 was also found to contain phosphoserine/phosphothreonine residues. alpha-Mannosidase-1 and other glycosidases are electrophoretically less negative when isolated from strain M31 than when isolated from wild type cells. The mutation present in M31, modA, appears to affect posttranslational modification, modA is a recessive mutation which we map onto linkage group I.

  9. Identification of potential genetic components involved in the deviant quorum-sensing signaling pathways of Burkholderia glumae through a functional genomics approach.

    PubMed

    Chen, Ruoxi; Barphagha, Inderjit K; Ham, Jong Hyun

    2015-01-01

    Burkholderia glumae is the chief causal agent for bacterial panicle blight of rice. The acyl-homoserine lactone (AHL)-mediated quorum-sensing (QS) system dependent on a pair of luxI and luxR homologs, tofI and tofR, is the primary cell-to-cell signaling mechanism determining the virulence of this bacterium. Production of toxoflavin, a major virulence factor of B. glumae, is known to be dependent on the tofI/tofR QS system. In our previous study, however, it was observed that B. glumae mutants defective in tofI or tofR produced toxoflavin if they grew on the surface of a solid medium, suggesting that alternative signaling pathways independent of tofI or tofR are activated in that growth condition for the production of toxoflavin. In this study, potential genetic components involved in the tofI- and tofR-independent signaling pathways for toxoflavin production were sought through screening random mini-Tn5 mutants of B. glumae to better understand the intercellular signaling pathways of this pathogen. Fifteen and three genes were initially identified as the potential genetic elements of the tofI- and tofR-independent pathways, respectively. Especially, the ORF (bglu_2g06320) divergently transcribed from toxJ, which encodes an orphan LuxR protein and controls toxoflavin biosynthesis, was newly identified in this study as a gene required for the tofR-independent toxoflavin production and named as toxK. Among those genes, flhD, dgcB, and wzyB were further studied to validate their functions in the tofI-independent toxoflavin production, and similar studies were also conducted with qsmR and toxK for their functions in the tofR-independent toxoflavin production. This work provides a foundation for future comprehensive studies of the intercellular signaling systems of B. glumae and other related pathogenic bacteria.

  10. Genetic variability in key genes in prostaglandin E2 pathway (COX-2, HPGD, ABCC4 and SLCO2A1) and their involvement in colorectal cancer development.

    PubMed

    Pereira, Carina; Queirós, Sara; Galaghar, Ana; Sousa, Hugo; Pimentel-Nunes, Pedro; Brandão, Catarina; Moreira-Dias, Luís; Medeiros, Rui; Dinis-Ribeiro, Mário

    2014-01-01

    The pro-carcinogenic effects of prostaglandin E2 (PGE2) in colonic mucosa are not only regulated by the rates between Cyclooxygenase-2 (COX-2) biosynthesis and 15-Hydroxyprostaglandin Dehydrogenase (15-PGDH)-dependent degradation but also the steady-state levels of PGE2 in extracellular microenvironment, maintained by key specific prostaglandin transporters, the Multidrug Resistance Protein (MRP4) (efflux carrier) and Prostaglandin Transporter (PGT) (influx carrier). To understand the contribution of genetic variability in genes coding for COX-2/15-PGDH/MRP4/PGT proteins in CRC development, we conducted a hospital-based case-control study involving 246 CRC patients and 480 cancer-free controls. A total of 51 tagSNPs were characterized using the Sequenom platform through multiplexed amplification followed by mass-spectrometric product separation or allelic discrimination using real-time PCR. Seven tagSNPs were implicated in CRC development: the rs689466 in COX-2 gene, the rs1346271 and rs1426945 in 15-PGDH, the rs6439448 and rs7616492 in PGT and rs1751051 and rs1751031 in MRP4 coding genes. Upon a stratified analysis a measurable gene-environment interaction was noticed between rs689466 and smoking habits, with individuals ever-smokers carriers of rs689466 GG homozygous genotype having a nearly 6-fold increased susceptibility for CRC onset (95%CI: 1.49-22.42, P = 0.011). Furthermore, the multifactor dimensionality reduction (MDR) analysis identified an overall four-factor best gene-gene interactive model, including the rs1426945, rs6439448, rs1751051 and rs1751031 polymorphisms. This model had the highest cross-validation consistency (10/10, P<0.0001) and an accuracy of 0.6957 and was further associated with a 5-fold increased risk for CRC development (95%CI: 3.89-7.02, P<0.0001). In conclusion, specific low penetrance genes in the pro-carcinogenic PGE2 pathway appear to modulate the genetic susceptibility for CRC development. A clearer understanding on CRC

  11. Modification in penicillin-binding proteins during in vivo development of genetic competence of Haemophilus influenzae is associated with a rapid change in the physiological state of cells.

    PubMed Central

    Dargis, M; Gourde, P; Beauchamp, D; Foiry, B; Jacques, M; Malouin, F

    1992-01-01

    By using whole-cell labeling assay with 125I-penicillin V, we observed a reduction in the binding of the radiolabeled beta-lactam to four or five penicillin-binding proteins (PBPs) in Haemophilus influenzae cells cultivated under specific conditions. PBPs 3A, 3B, 4, and 6 were altered after the growth of bacteria in diffusion chambers implanted in the peritoneal cavity of rats. PBP 2 was also modified when cells were cultivated in human cerebrospinal fluids. Because this observation may have important consequences on the efficacy of beta-lactams during antibiotic therapy, we characterized the physiological state of bacteria cultivated in animals in the hope of explaining how such important changes in cell properties develop in vivo. Since the development of natural genetic competence occurs at the stationary phase of growth in H. influenzae, we used a DNA transformation assay to evaluate the physiological state of bacteria grown in diffusion chambers implanted in rats. Chromosomal DNA isolated from an antibiotic-resistant donor strain was mixed with bacteria in diffusion chambers. At different times during a 5-h incubation period, recipient bacteria were collected from the chambers, CFU were determined by plate counting, and antibiotic-resistant transformants were isolated on selective plates. Genetic competence rapidly developed in cells grown in rats, and the frequency of transformation by test DNA was elevated. Electron microscopy revealed an irregular cell shape and blebs at the surface of bacteria cultivated in animals and in cerebrospinal fluids. In an attempt to induce a similar physiological state in vitro, we supplemented broth cultures with cyclic AMP or synchronized cultures by a nutritional upshift. No changes in PBPs were observed with supplemental cyclic AMP or during a single cell cycle. Finally, a reduction in the affinity of PBPs for 125I-penicillin V identical to that observed in bacteria grown in rats was observed in cells isolated from the

  12. Genetic Testing for ALS

    MedlinePlus

    ... Involved Donate Familial Amyotrophic Lateral Sclerosis (FALS) and Genetic Testing By Deborah Hartzfeld, MS, CGC, Certified Genetic ... guarantee a person will develop symptoms of ALS. Genetic Counseling If there is more than one person ...

  13. Alpharetroviral self-inactivating vectors produced by a superinfection-resistant stable packaging cell line allow genetic modification of primary human T lymphocytes.

    PubMed

    Labenski, Verena; Suerth, Julia D; Barczak, Elke; Heckl, Dirk; Levy, Camille; Bernadin, Ornellie; Charpentier, Emmanuelle; Williams, David A; Fehse, Boris; Verhoeyen, Els; Schambach, Axel

    2016-08-01

    Primary human T lymphocytes represent an important cell population for adoptive immunotherapies, including chimeric-antigen and T-cell receptor applications, as they have the capability to eliminate non-self, virus-infected and tumor cells. Given the increasing numbers of clinical immunotherapy applications, the development of an optimal vector platform for genetic T lymphocyte engineering, which allows cost-effective high-quality vector productions, remains a critical goal. Alpharetroviral self-inactivating vectors (ARV) have several advantages compared to other vector platforms, including a more random genomic integration pattern and reduced likelihood for inducing aberrant splicing of integrated proviruses. We developed an ARV platform for the transduction of primary human T lymphocytes. We demonstrated functional transgene transfer using the clinically relevant herpes-simplex-virus thymidine kinase variant TK.007. Proof-of-concept of alpharetroviral-mediated T-lymphocyte engineering was shown in vitro and in a humanized transplantation model in vivo. Furthermore, we established a stable, human alpharetroviral packaging cell line in which we deleted the entry receptor (SLC1A5) for RD114/TR-pseudotyped ARVs to prevent superinfection and enhance genomic integrity of the packaging cell line and viral particles. We showed that superinfection can be entirely prevented, while maintaining high recombinant virus titers. Taken together, this resulted in an improved production platform representing an economic strategy for translating the promising features of ARVs for therapeutic T-lymphocyte engineering. PMID:27162078

  14. The hyperactive Sleeping Beauty transposase SB100X improves the genetic modification of T cells to express a chimeric antigen receptor.

    PubMed

    Jin, Z; Maiti, S; Huls, H; Singh, H; Olivares, S; Mátés, L; Izsvák, Z; Ivics, Z; Lee, D A; Champlin, R E; Cooper, L J N

    2011-09-01

    Sleeping Beauty (SB3) transposon and transposase constitute a DNA plasmid system used for therapeutic human cell genetic engineering. Here we report a comparison of SB100X, a newly developed hyperactive SB transposase, to a previous generation SB11 transposase to achieve stable expression of a CD19-specific chimeric antigen receptor (CAR3) in primary human T cells. The electro-transfer of SB100X expressed from a DNA plasmid or as an introduced mRNA species had superior transposase activity in T cells based on the measurement of excision circles released after transposition and emergence of CAR expression on T cells selectively propagated upon CD19+ artificial antigen-presenting cells. Given that T cells modified with SB100X and SB11 integrate on average one copy of the CAR transposon in each T-cell genome, the improved transposition mediated by SB100X apparently leads to an augmented founder effect of electroporated T cells with durable integration of CAR. In aggregate, SB100X improves SB transposition in primary human T cells and can be titrated with an SB transposon plasmid to improve the generation of CD19-specific CAR+ T cells. PMID:21451576

  15. Alpharetroviral self-inactivating vectors produced by a superinfection-resistant stable packaging cell line allow genetic modification of primary human T lymphocytes.

    PubMed

    Labenski, Verena; Suerth, Julia D; Barczak, Elke; Heckl, Dirk; Levy, Camille; Bernadin, Ornellie; Charpentier, Emmanuelle; Williams, David A; Fehse, Boris; Verhoeyen, Els; Schambach, Axel

    2016-08-01

    Primary human T lymphocytes represent an important cell population for adoptive immunotherapies, including chimeric-antigen and T-cell receptor applications, as they have the capability to eliminate non-self, virus-infected and tumor cells. Given the increasing numbers of clinical immunotherapy applications, the development of an optimal vector platform for genetic T lymphocyte engineering, which allows cost-effective high-quality vector productions, remains a critical goal. Alpharetroviral self-inactivating vectors (ARV) have several advantages compared to other vector platforms, including a more random genomic integration pattern and reduced likelihood for inducing aberrant splicing of integrated proviruses. We developed an ARV platform for the transduction of primary human T lymphocytes. We demonstrated functional transgene transfer using the clinically relevant herpes-simplex-virus thymidine kinase variant TK.007. Proof-of-concept of alpharetroviral-mediated T-lymphocyte engineering was shown in vitro and in a humanized transplantation model in vivo. Furthermore, we established a stable, human alpharetroviral packaging cell line in which we deleted the entry receptor (SLC1A5) for RD114/TR-pseudotyped ARVs to prevent superinfection and enhance genomic integrity of the packaging cell line and viral particles. We showed that superinfection can be entirely prevented, while maintaining high recombinant virus titers. Taken together, this resulted in an improved production platform representing an economic strategy for translating the promising features of ARVs for therapeutic T-lymphocyte engineering.

  16. RNA polymerase II transcriptional fidelity control and its functional interplay with DNA modifications

    PubMed Central

    Xu, Liang; Wang, Wei; Chong, Jenny; Shin, Ji Hyun; Xu, Jun; Wang, Dong

    2016-01-01

    Accurate genetic information transfer is essential for life. As a key enzyme involved in the first step of gene expression, RNA polymerase II (Pol II) must maintain high transcriptional fidelity while it reads along DNA template and synthesizes RNA transcript in a stepwise manner during transcription elongation. DNA lesions or modifications may lead to significant changes in transcriptional fidelity or transcription elongation dynamics. In this review, we will summarize recent progress towards understanding the molecular basis of RNA Pol II transcriptional fidelity control and impacts of DNA lesions and modifications on Pol II transcription elongation. PMID:26392149

  17. The yeast nucleolar protein Cbf5p is involved in rRNA biosynthesis and interacts genetically with the RNA polymerase I transcription factor RRN3.

    PubMed Central

    Cadwell, C; Yoon, H J; Zebarjadian, Y; Carbon, J

    1997-01-01

    Yeast Cbf5p was originally isolated as a low-affinity centromeric DNA binding protein (W. Jiang, K. Middleton, H.-J. Yoon, C. Fouquet, and J. Carbon, Mol. Cell. Biol. 13:4884-4893, 1993). Cbf5p also binds microtubules in vitro and interacts genetically with two known centromere-related protein genes (NDC10/CBF2 and MCK1). However, Cbf5p was found to be nucleolar and is highly homologous to the rat nucleolar protein NAP57, which coimmunoprecipitates with Nopp140 and which is postulated to be involved in nucleolar-cytoplasmic shuttling (U. T. Meier, and G. Blobel, J. Cell Biol. 127:1505-1514, 1994). The temperature-sensitive cbf5-1 mutant demonstrates a pronounced defect in rRNA biosynthesis at restrictive temperatures, while tRNA transcription and pre-rRNA and pre-tRNA cleavage processing appear normal. The cbf5-1 mutant cells are deficient in cytoplasmic ribosomal subunits at both permissive and restrictive temperatures. A high-copy-number yeast genomic library was screened for genes that suppress the cbf5-1 temperature-sensitive growth phenotype. SYC1 (suppressor of yeast cbf5-1) was identified as a multicopy suppressor of cbf5-1 and subsequently was found to be identical to RRN3, an RNA polymerase I transcription factor. A cbf5delta null mutant is not rescued by plasmid pNOY103 containing a yeast 35S rRNA gene under the control of a Pol II promoter, indicating that Cbf5p has one or more essential functions in addition to its role in rRNA transcription. PMID:9315678

  18. Modulation of endotoxicity of Shigella generalized modules for membrane antigens (GMMA) by genetic lipid A modifications: relative activation of TLR4 and TLR2 pathways in different mutants.

    PubMed

    Rossi, Omar; Pesce, Isabella; Giannelli, Carlo; Aprea, Susanna; Caboni, Mariaelena; Citiulo, Francesco; Valentini, Sara; Ferlenghi, Ilaria; MacLennan, Calman Alexander; D'Oro, Ugo; Saul, Allan; Gerke, Christiane

    2014-09-01

    Outer membrane particles from Gram-negative bacteria are attractive vaccine candidates as they present surface antigens in their natural context. We previously developed a high yield production process for genetically derived particles, called generalized modules for membrane antigens (GMMA), from Shigella. As GMMA are derived from the outer membrane, they contain immunostimulatory components, especially lipopolysaccharide (LPS). We examined ways of reducing their reactogenicity by modifying lipid A, the endotoxic part of LPS, through deletion of late acyltransferase genes, msbB or htrB, in GMMA-producing Shigella sonnei and Shigella flexneri strains. GMMA with resulting penta-acylated lipid A from the msbB mutants showed a 600-fold reduced ability, and GMMA from the S. sonnei ΔhtrB mutant showed a 60,000-fold reduced ability compared with GMMA with wild-type lipid A to stimulate human Toll-like receptor 4 (TLR4) in a reporter cell line. In human peripheral blood mononuclear cells, GMMA with penta-acylated lipid A showed a marked reduction in induction of inflammatory cytokines (S. sonnei ΔhtrB, 800-fold; ΔmsbB mutants, 300-fold). We found that the residual activity of these GMMA is largely due to non-lipid A-related TLR2 activation. In contrast, in the S. flexneri ΔhtrB mutant, a compensatory lipid A palmitoleoylation resulted in GMMA with hexa-acylated lipid A with ∼10-fold higher activity to stimulate peripheral blood mononuclear cells than GMMA with penta-acylated lipid A, mostly due to retained TLR4 activity. Thus, for use as vaccines, GMMA will likely require lipid A penta-acylation. The results identify the relative contributions of TLR4 and TLR2 activation by GMMA, which need to be taken into consideration for GMMA vaccine development.

  19. Liquid chromatography-diode array detection to study the metabolism of glufosinate in Triticum aestivum T-590 and influence of the genetic modification on its resistance.

    PubMed

    Rojano-Delgado, Antonia María; Priego-Capote, Feliciano; Barro, Francisco; de Castro, María Dolores Luque; De Prado, Rafael

    2013-12-01

    The resistance to glufosinate of two lines-genetically modified (GM) and unmodified (T-590 and T-549, respectively)-of Triticum aestivum has been studied. In the GM line, the bar gene was introduced to increase the resistance to glufosinate. Experiments in a controlled growth chamber showed that line T-590 presented a high resistance to glufosinate with an ED50 value of 478.59 g active ingredient per hectare (g ai ha(-1)) versus 32.65 g ai ha(-1) for line T-549. The activity of glutamine synthetase (GS) in leaf extracts from both lines was investigated. The I50 for line T-590 was 694.10 μM glufosinate versus 55.46 μM for line T-549, with a resistance factor of 12.51. Metabolism studies showed a higher and faster penetration of glufosinate in line T-549 than in line T-590. LC-TOF/MS analysis of glufosinate metabolism at 48 h after herbicide treatment (300 g ai ha(-1)) revealed an 83.4% conversion of the herbicide (66.5% in N-acetyl-glufosinate metabolite), while in line T-549 conversion of the herbicide was about 40% (0% to N-acetyl-glufosinate). These results suggest that metabolism of glufosinate by the bar gene is a key mechanism of resistance in line T-590 that explains such high levels of herbicide tolerated by the plant, together with other mechanisms due to unmodified pathway, absorption and loss of glufosinate affinity for its target site.

  20. Chemical modification of chitosan for efficient gene therapy.

    PubMed

    Jiang, Hu-Lin; Cui, Peng-Fei; Xie, Rong-Lin; Cho, Chong-Su

    2014-01-01

    Gene therapy involves the introduction of foreign genetic material into cells in order to exert a therapeutic effect. Successful gene therapy relies on effective vector system. Viral vectors are highly efficient in transfecting cells, but the undesirable complications limit their therapeutic applications. As a natural biopolymer, chitosan has been considered to be a good gene carrier candidate due to its ideal character which combines biocompatibility, low toxicity with high cationic density together. However, the low cell specificity and low transfection efficiency of chitosan as a gene carrier need to be overcome before undertaking clinical trials. This chapter is principally on those endeavors such as chemical modifications using cell-specific ligands and stimuli-response groups as well as penetrating modifications that have been done to increase the performances of chitosan in gene therapy.

  1. Arthropod Genetics.

    ERIC Educational Resources Information Center

    Zumwalde, Sharon

    2000-01-01

    Introduces an activity on arthropod genetics that involves phenotype and genotype identification of the creature and the construction process. Includes a list of required materials and directions to build a model arthropod. (YDS)

  2. Germline modification of domestic animals

    PubMed Central

    Tang, L.; González, R.; Dobrinski, I.

    2016-01-01

    Genetically-modified domestic animal models are of increasing significance in biomedical research and agriculture. As authentic ES cells derived from domestic animals are not yet available, the prevailing approaches for engineering genetic modifications in those animals are pronuclear microinjection and somatic cell nuclear transfer (SCNT, also known as cloning). Both pronuclear microinjection and SCNT are inefficient, costly, and time-consuming. In animals produced by pronuclear microinjection, the exogenous transgene is usually inserted randomly into the genome, which results in highly variable expression patterns and levels in different founders. Therefore, significant efforts are required to generate and screen multiple founders to obtain animals with optimal transgene expression. For SCNT, specific genetic modifications (both gain-of-function and loss-of-function) can be engineered and carefully selected in the somatic cell nucleus before nuclear transfer. SCNT has been used to generate a variety of genetically modified animals such as goats, pigs, sheep and cattle; however, animals resulting from SCNT frequently suffer from developmental abnormalities associated with incomplete nuclear reprogramming. Other strategies to generate genetically-modified animals rely on the use of the spermatozoon as a natural vector to introduce genetic material into the female gamete. This sperm mediated DNA transfer (SMGT) combined with intracytoplasmatic sperm injection (ICSI) has relatively high efficiency and allows the insertion of large DNA fragments, which, in turn, enhance proper gene expression. An approach currently being developed to complement SCNT for producing genetically modified animals is germ cell transplantation using genetically modified male germline stem cells (GSCs). This approach relies on the ability of GSCs that are genetically modified in vitro to colonize the recipient testis and produce donor derived sperm upon transplantation. As the genetic change

  3. Involvement of GABAB receptors in biochemical alterations induced by anxiety-related responses to nicotine in mice: genetic and pharmacological approaches.

    PubMed

    Varani, Andrés P; Pedrón, Valeria T; Bettler, Bernhard; Balerio, Graciela N

    2014-06-01

    Previous studies from our laboratory showed that anxiety-related responses induced by nicotine (NIC), measured by the elevated plus maze, were abolished by 2-OH-saclofen (GABAB receptor antagonist) (1 mg/kg; ip) or the lack of GABAB receptors (GABAB1 knockout mice). Based on these behavioral data, the aims of the present study were: 1) to evaluate the possible neurochemical changes (dopamine, DA, serotonin, 5-HT, 3,4-dihydroxyphenylacetic acid, DOPAC, 5-hydroxyindoleacetic acid, 5-HIAA and noradrenaline, NA) and the c-Fos expression induced by the anxiolytic (0.05 mg/kg) or anxiogenic (0.8 mg/kg) doses of NIC in the dorsal raphe (DRN) and lateral septal (LSN) nucleus; 2) to study the possible involvement of GABAB receptors on the neurochemical alterations and c-Fos expression induced by NIC (0.05 and 0.8 mg/kg), using both pharmacological (2-OH-saclofen) and genetic (mice GABAB1 knockout) approaches. The results revealed that in wild-type mice, NIC (0.05 mg/kg) increased the concentration of 5-HT and 5-HIAA (p < 0.05) in the DRN, and NIC (0.8 mg/kg) increased the levels of 5-HT (p < 0.01) and NA (p < 0.05) in the LSN. Additionally, 2-OH-saclofen pretreatment (1 mg/kg, ip) or the lack of GABAB receptors abolished these neurochemical changes induced by NIC (p < 0.01, p < 0.05, respectively). On the other hand, NIC 0.05 and 0.8 mg/kg increased (p < 0.01) the c-Fos expression in the DRN and LSN respectively, in wild-type mice. In addition, 2-OH-saclofen pretreatment (1 mg/kg, ip) or the lack of GABAB receptors prevented the c-Fos alterations induced by NIC (p < 0.01). In summary, both approaches show that GABAB receptors would participate in the modulation of anxiolytic- and anxiogenic-like responses induced by NIC, suggesting the potential therapeutic target of these receptors for the tobacco addiction treatment.

  4. A combined strategy involving Sanger and 454 pyrosequencing increases genomic resources to aid in the management of reproduction, disease control and genetic selection in the turbot (Scophthalmus maximus)

    PubMed Central

    2013-01-01

    Background Genomic resources for plant and animal species that are under exploitation primarily for human consumption are increasingly important, among other things, for understanding physiological processes and for establishing adequate genetic selection programs. Current available techniques for high-throughput sequencing have been implemented in a number of species, including fish, to obtain a proper description of the transcriptome. The objective of this study was to generate a comprehensive transcriptomic database in turbot, a highly priced farmed fish species in Europe, with potential expansion to other areas of the world, for which there are unsolved production bottlenecks, to understand better reproductive- and immune-related functions. This information is essential to implement marker assisted selection programs useful for the turbot industry. Results Expressed sequence tags were generated by Sanger sequencing of cDNA libraries from different immune-related tissues after several parasitic challenges. The resulting database (“Turbot 2 database”) was enlarged with sequences generated from a 454 sequencing run of brain-hypophysis-gonadal axis-derived RNA obtained from turbot at different development stages. The assembly of Sanger and 454 sequences generated 52,427 consensus sequences (“Turbot 3 database”), of which 23,661 were successfully annotated. A total of 1,410 sequences were confirmed to be related to reproduction and key genes involved in sex differentiation and maturation were identified for the first time in turbot (AR, AMH, SRY-related genes, CYP19A, ZPGs, STAR FSHR, etc.). Similarly, 2,241 sequences were related to the immune system and several novel key immune genes were identified (BCL, TRAF, NCK, CD28 and TOLLIP, among others). The number of genes of many relevant reproduction- and immune-related pathways present in the database was 50–90% of the total gene count of each pathway. In addition, 1,237 microsatellites and 7,362 single

  5. Pnp gene modification for improved xylose utilization in Zymomonas

    DOEpatents

    Caimi, Perry G G; Qi, Min; Tao, Luan; Viitanen, Paul V; Yang, Jianjun

    2014-12-16

    The endogenous pnp gene encoding polynucleotide phosphorylase in the Zymomonas genome was identified as a target for modification to provide improved xylose utilizing cells for ethanol production. The cells are in addition genetically modified to have increased expression of ribose-5-phosphate isomerase (RPI) activity, as compared to cells without this genetic modification, and are not limited in xylose isomerase activity in the absence of the pnp modification.

  6. Changing Attitudes Through Behavior Modification.

    ERIC Educational Resources Information Center

    Whipple, W. Scott

    This article describes the philosophy and methods used by the staff at the Granite Alternative School in changing student attitudes through behavior modification. The students involved all have a failure syndrome or low self-image, and are dropouts from traditional high schools. Among the techniques used are: (1) reinforcing good behavior (praise…

  7. [Lung cancer, smoking and genetic modifications].

    PubMed

    Popescu, Iulian

    2008-01-01

    Lung cancer represents a heterogeneity entity consisting of different histopathologic, clinic and therapeutic subtypes. Smoking is the most well-known risk factor, but the mechanisms of occurrence of lung cancer are not completely understood yet. The effects of smoking are related to the young age of beginning of smoking, the daily dose and the length of smoking, but they also depend of the individual heredity due to the gene polymorphism. Smoking induces several deleterious mechanisms: inactivation of tumor-suppressor genes, activation of proto-oncogenes, increase in telomerase's activity.

  8. Somatic Embryogenesis and Genetic Modification of Vitis.

    PubMed

    Dhekney, Sadanand A; Li, Zhijian T; Grant, Trudi N L; Gray, Dennis J

    2016-01-01

    Grapevine embryogenic cultures are ideal target tissues for inserting desired traits of interest and improving existing cultivars via precision breeding (PB). PB is a new approach that, like conventional breeding, utilizes only DNA fragments obtained from sexually compatible grapevine plants. Embryogenic culture induction occurs by placing leaves or stamens and pistils on induction medium with a dark/light photoperiod cycle for 12-16 weeks. Resulting cultures produce sectors of embryogenic and non-embryogenic callus, which can be identified on the basis of callus morphology and color. Somatic embryo development occurs following transfer of embryogenic callus to development medium and cultures can be maintained for extended periods of time by transfer of the proliferating proembryonic masses to fresh medium at 4-6-week intervals. To demonstrate plant recovery via PB, somatic embryos at the mid-cotyledonary stage are cocultivated with Agrobacterium containing the desired gene of interest along with a, non-PB, enhanced green fluorescent protein/neomycin phosphotransferase II (egfp/nptII) fusion gene. Modified cultures are grown on proliferation and development medium to produce uniformly modified somatic embryos via secondary embryogenesis. Modified embryos identified on the basis of green fluorescence and kanamycin resistance are transferred to germination medium for plant development. The resulting plants are considered to prototype examples of the PB approach, since they contain egfp/nptII, a non-grapevine-derived fusion gene. Uniform green fluorescent protein (GFP) fluorescence can be observed in all tissues of regenerated plants.

  9. Current Perspectives on Epigenetic Modifications by Dietary Chemopreventive and Herbal Phytochemicals

    PubMed Central

    Guo, Yue; Su, Zheng-Yuan; Kong, Ah-Ng Tony

    2015-01-01

    Studies during the last two decades have revealed the involvement of epigenetic modifications in the development of human cancer. It is now recognized that the interplay of DNA methylation, post-translational histone modification, and non-coding RNAs can interact with genetic defects to drive tumorigenesis. The early onset, reversibility, and dynamic nature of such epigenetic modifications enable them to be developed as promising cancer biomarkers and preventive/therapeutic targets. In addition to the recent approval of several epigenetic therapies in the treatment of human cancer, emerging studies have indicated that dietary phytochemicals might exert cancer chemopreventive effects by targeting epigenetic mechanisms. In this review, we will present the current understanding of the epigenetic alterations in carcinogenesis and highlight the potential of targeting these mechanisms to treat/prevent cancer. The latest findings, published in the past three years regarding the effects of dietary phytochemicals in modulating epigenetic mechanisms will also be discussed. PMID:26328267

  10. Improving experimental phases for strong reflections prior to density modification

    SciTech Connect

    Uervirojnangkoorn, Monarin; Hilgenfeld, Rolf; Terwilliger, Thomas C.; Read, Randy J.

    2013-10-01

    A genetic algorithm has been developed to optimize the phases of the strongest reflections in SIR/SAD data. This is shown to facilitate density modification and model building in several test cases. Experimental phasing of diffraction data from macromolecular crystals involves deriving phase probability distributions. These distributions are often bimodal, making their weighted average, the centroid phase, improbable, so that electron-density maps computed using centroid phases are often non-interpretable. Density modification brings in information about the characteristics of electron density in protein crystals. In successful cases, this allows a choice between the modes in the phase probability distributions, and the maps can cross the borderline between non-interpretable and interpretable. Based on the suggestions by Vekhter [Vekhter (2005 ▶), Acta Cryst. D61, 899–902], the impact of identifying optimized phases for a small number of strong reflections prior to the density-modification process was investigated while using the centroid phase as a starting point for the remaining reflections. A genetic algorithm was developed that optimizes the quality of such phases using the skewness of the density map as a target function. Phases optimized in this way are then used in density modification. In most of the tests, the resulting maps were of higher quality than maps generated from the original centroid phases. In one of the test cases, the new method sufficiently improved a marginal set of experimental SAD phases to enable successful map interpretation. A computer program, SISA, has been developed to apply this method for phase improvement in macromolecular crystallography.

  11. [Genetics and epigenetics of schizophrenia].

    PubMed

    Fabi, E; Fusco, A; Valiante, M; Celli, R

    2013-01-01

    Schizophrenia is a severe psychiatric disorder with an estimate prevalence of 0.3-0.7%. Studies on family aggregation showed a higher incidence of disease among family members of affected people. This observation lead to formulate the hypothesis that schizophrenia could be inheritable, but twin studies have shown a concordance of disease between monozygotic twins only of 50%, indicating the concomitant role of environmental factors in the pathogenesis of schizophrenia. Researches in molecular biology field have allowed the identification of genes that confer susceptibility to schizophrenia on chromosomes 1, 2, 3, 5, 6, 8, 10, 11, 13, 14, 20 and 22. Epigenetic modifications of gene expression, that not involve the primary DNA sequence, may also predispose to schizophrenia, in particular the methylation of genes involved in neurotransmission (RELN, GAD1, MARLIN-1, and NR3B GRIA2, VGLUT1 and 2, 5HT2a, COMT and BDNF), the histone modifications and the action of non-coding RNAs. This review deals with the results of a bibliographic retrieval on PubMed, carried out, using the key words: schizophrenia, genetics, epigenetics. From the epitomized results it can be derived that schizophrenia seems to be a multifactorial disease. Environmental factors, that can cause epigenetic modifications, are important in its pathogenesis, acting on a biological inheritable vulnerability. PMID:24045531

  12. Mechanisms of Non-Genetic Inheritance and Psychiatric Disorders

    PubMed Central

    Toth, Miklos

    2015-01-01

    Inheritance is typically associated with the Mendelian transmission of information from parents to offspring by alleles (DNA sequence). However, empirical data clearly suggest that traits can be acquired from ancestors by mechanisms that do not involve genetic alleles, referred to as non-genetic inheritance. Information that is non-genetically transmitted across generations includes parental experience and exposure to certain environments, but also parental mutations and polymorphisms, because they can change the parental ‘intrinsic' environment. Non-genetic inheritance is not limited to the first generation of the progeny, but can involve the grandchildren and even further generations. Non-genetic inheritance has been observed for multiple traits including overall development, cardiovascular risk and metabolic symptoms, but this review will focus on the inheritance of behavioral abnormalities pertinent to psychiatric disorders. Multigenerational non-genetic inheritance is often interpreted as the transmission of epigenetic marks, such as DNA methylation and chromatin modifications, via the gametes (transgenerational epigenetic inheritance). However, information can be carried across generations by a large number of bioactive substances, including hormones, cytokines, and even microorganisms, without the involvement of the gametes. We reason that this broader definition of non-genetic inheritance is more appropriate, especially in the context of psychiatric disorders, because of the well-recognized role of parental and early life environmental factors in later life psychopathology. Here we discuss the various forms of non-genetic inheritance in humans and animals, as well as rodent models of psychiatric conditions to illustrate possible mechanisms. PMID:24889369

  13. Chromatin architecture near a potential 3' end of the igh locus involves modular regulation of histone modifications during B-Cell development and in vivo occupancy at CTCF sites.

    PubMed

    Garrett, Francine E; Emelyanov, Alexander V; Sepulveda, Manuel A; Flanagan, Patrick; Volpi, Sabrina; Li, Fubin; Loukinov, Dmitry; Eckhardt, Laurel A; Lobanenkov, Victor V; Birshtein, Barbara K

    2005-02-01

    The murine Igh locus has a 3' regulatory region (3' RR) containing four enhancers (hs3A, hs1,2, hs3B, and hs4) at DNase I-hypersensitive sites. The 3' RR exerts long-range effects on class switch recombination (CSR) to several isotypes through its control of germ line transcription. By measuring levels of acetylated histones H3 and H4 and of dimethylated H3 (K4) with chromatin immunoprecipitation assays, we found that early in B-cell development, chromatin encompassing the enhancers of the 3' RR began to attain stepwise modifications typical of an open conformation. The hs4 enhancer was associated with active chromatin initially in pro- and pre-B cells and then together with hs3A, hs1,2, and hs3B in B and plasma cells. Histone modifications were similar in resting splenic B cells and in splenic B cells induced by lipopolysaccharide to undergo CSR. From the pro-B-cell stage onward, the approximately 11-kb region immediately downstream of hs4 displayed H3 and H4 modifications indicative of open chromatin. This region contained newly identified DNase I-hypersensitive sites and several CTCF target sites, some of which were occupied in vivo in a developmentally regulated manner. The open chromatin environment of the extended 3' RR in mature B cells was flanked by regions associated with dimethylated K9 of histone H3. Together, these data suggest that 3' RR elements are located within a specific chromatin subdomain that contains CTCF binding sites and developmentally regulated modules.

  14. Glacial vicariance in Eurasia: mitochondrial DNA evidence from Scots pine for a complex heritage involving genetically distinct refugia at mid-northern latitudes and in Asia Minor

    PubMed Central

    Naydenov, Krassimir; Senneville, Sauphie; Beaulieu, Jean; Tremblay, Francine; Bousquet, Jean

    2007-01-01

    Background At the last glacial maximum, Fennoscandia was covered by an ice sheet while the tundra occupied most of the rest of northern Eurasia. More or less disjunct refugial populations of plants were dispersed in southern Europe, often trapped between mountain ranges and seas. Genetic and paleobotanical evidences indicate that these populations have contributed much to Holocene recolonization of more northern latitudes. Less supportive evidence has been found for the existence of glacial populations located closer to the ice margin. Scots pine (Pinus sylvestris L.) is a nordic conifer with a wide natural range covering much of Eurasia. Fractures in its extant genetic structure might be indicative of glacial vicariance and how different refugia contributed to the current distribution at the continental level. The population structure of Scots pine was investigated on much of its Eurasian natural range using maternally inherited mitochondrial DNA polymorphisms. Results A novel polymorphic region of the Scots pine mitochondrial genome has been identified, the intron 1 of nad7, with three variants caused by insertions-deletions. From 986 trees distributed among 54 populations, four distinct multi-locus mitochondrial haplotypes (mitotypes) were detected based on the three nad7 intron 1 haplotypes and two previously reported size variants for nad1 intron B/C. Population differentiation was high (GST = 0.657) and the distribution of the mitotypes was geographically highly structured, suggesting at least four genetically distinct ancestral lineages. A cosmopolitan lineage was widely distributed in much of Europe throughout eastern Asia. A previously reported lineage limited to the Iberian Peninsula was confirmed. A new geographically restricted lineage was found confined to Asia Minor. A new lineage was restricted to more northern latitudes in northeastern Europe and the Baltic region. Conclusion The contribution of the various ancestral lineages to the current

  15. Comparative genetic analysis of quantitative traits in sunflower (Helianthus annuus L.). 2. Characterisation of QTL involved in developmental and agronomic traits.

    PubMed

    Bert, P-F; Jouan, I; Tourvieille de Labrouhe, D; Serre, F; Philippon, J; Nicolas, P; Vear, F

    2003-06-01

    Seed weight and oil content are important properties of cultivated sunflower under complex genetic and environmental control, and associated with morphological and developmental characteristics such as plant height or flowering dates. Using a genetic map with 290 markers for a cross between two inbred sunflower lines and 2 years of observations on F3 families, QTL controlling seed weight, oil content, plant height, plant lodging, flowering dates, maturity dates and delay from flowering to maturity were detected. QTL detected were compared between the F2 and F3 generations and between the 2 years of testing for the F3 families in 1997 and 1999. Some of the QTL controlling seed weight overlapped with those controlling oil content. Several other co-localisations of QTL controlling developmental or morphological characteristics were observed and the relationships between the traits were also shown by correlation analyses. The relationships between all these traits and with resistance to Sclerotinia sclerotiorum and Diaporthe helianthi are discussed.

  16. Untangling the interplay of genetic and metabolic influences on beta-cell function: Examples of potential therapeutic implications involving TCF7L2 and FFAR1☆

    PubMed Central

    Wagner, Robert; Staiger, Harald; Ullrich, Susanne; Stefan, Norbert; Fritsche, Andreas; Häring, Hans-Ulrich

    2014-01-01

    Deteriorating beta-cell function is a common feature of type 2 diabetes. In this review, we briefly address the regulation of beta-cell function, and discuss some of the main determinants of beta-cell failure. We will focus on the role of interactions between the genetic background and metabolic environment (insulin resistance, fuel supply and flux as well as metabolic signaling). We present data on the function of the strongest common diabetes risk variant, the single nucleotide polymorphism (SNP) rs7903146 in TCF7L2. As also mirrored by its interaction with glycemia on insulin secretion, this SNP in large part confers resistance against the incretin effect. Genetic influence on insulin secretion also interacts with free fatty acids, as evidenced by data on rs1573611 in FFAR1. Several medications marketed by now or currently under development for diabetes treatment engage these pathways, and therapeutic implications from these findings are soon to be expected. PMID:24749055

  17. Resistance to Botrytis cinerea in sitiens, an Abscisic Acid-Deficient Tomato Mutant, Involves Timely Production of Hydrogen Peroxide and Cell Wall Modifications in the Epidermis1[C][W][OA

    PubMed Central

    Asselbergh, Bob; Curvers, Katrien; França, Soraya C.; Audenaert, Kris; Vuylsteke, Marnik; Van Breusegem, Frank; Höfte, Monica

    2007-01-01

    Plant defense mechanisms against necrotrophic pathogens, such as Botrytis cinerea, are considered to be complex and to differ from those that are effective against biotrophs. In the abscisic acid-deficient sitiens tomato (Solanum lycopersicum) mutant, which is highly resistant to B. cinerea, accumulation of hydrogen peroxide (H2O2) was earlier and stronger than in the susceptible wild type at the site of infection. In sitiens, H2O2 accumulation was observed from 4 h postinoculation (hpi), specifically in the leaf epidermal cell walls, where it caused modification by protein cross-linking and incorporation of phenolic compounds. In wild-type tomato plants, H2O2 started to accumulate 24 hpi in the mesophyll layer and was associated with spreading cell death. Transcript-profiling analysis using TOM1 microarrays revealed that defense-related transcript accumulation prior to infection was higher in sitiens than in wild type. Moreover, further elevation of sitiens defense gene expression was stronger than in wild type 8 hpi both in number of genes and in their expression levels and confirmed a role for cell wall modification in the resistant reaction. Although, in general, plant defense-related reactive oxygen species formation facilitates necrotrophic colonization, these data indicate that timely hyperinduction of H2O2-dependent defenses in the epidermal cell wall can effectively block early development of B. cinerea. PMID:17573540

  18. Genetic and epigenetic basis of psoriasis pathogenesis.

    PubMed

    Chandra, Aditi; Ray, Aditi; Senapati, Swapan; Chatterjee, Raghunath

    2015-04-01

    Psoriasis is a chronic inflammatory skin disease whose prevalence varies among different populations worldwide. It is a complex multi-factorial disease and the exact etiology is largely unknown. Family based studies have indicated a genetic predisposition; however they cannot fully explain the disease pathogenesis. In addition to genetic susceptibility, environmental as well as gender and age related factors were also been found to be associated. Recently, imbalances in epigenetic networks are indicated to be causative elements in psoriasis. The present knowledge of epigenetic involvement, mainly the DNA methylation, chromatin modifications and miRNA deregulation is surveyed here. An integrated approach considering genetic and epigenetic anomalies in the light of immunological network may explore the pathogenesis of psoriasis.

  19. Histone H3 modifications and Cdx-2 binding to the sucrase-isomaltase (SI) gene is involved in induction of the gene in the transition from the crypt to villus in the small intestine of rats.

    PubMed

    Suzuki, Takuji; Mochizuki, Kazuki; Goda, Toshinao

    2008-05-01

    Expression of the sucrase-isomaltase (SI) gene is induced in cells transitioning from the crypt to the villus of rat jejunum. In the present study, we revealed by ChIP assay using a cryostat sectioning technique that binding of the di-acetylated histone H3 at lysine 9/14 and the transcriptional factor Cdx-2 to the promoter region on the SI gene, as well as mRNA, increased in the transient process. Additionally, di-/tri-methylation of histone H3 at lysine 9/14 on the promoter region of the SI gene rapidly decreased with increasing mRNA. These results suggest that induction of the SI gene during the transition from the crypt to the villi is associated with changes in histone H3 modifications from methylation at lysine 9 to di-acetylation at lysine 9/14, as well as increased binding of Cdx-2 to the SI promoter region.

  20. Use of genetically modified viruses and genetically engineered virus-vector vaccines: environmental effects.

    PubMed

    Chan, Vivian S W

    2006-11-01

    Despite major therapeutic advances, infectious diseases remain highly problematic. Recent advancements in technology in producing DNA-based vaccines, together with the growing knowledge of the immune system, have provided new insights into the identification of the epitopes needed to target the development of highly targeted vaccines. Genetically modified (GM) viruses and genetically engineered virus-vector vaccines possess significant unpredictability and a number of inherent harmful potential hazards. For all these vaccines, safety assessment concerning unintended and unwanted side effects with regard to targeted vaccinees has always been the main focus. Important questions concerning effects on nontargeted individuals within the same species or other species remain unknown. Horizontal transfer of genes, though lacking supportive experimental or epidemiological investigations, is well established. New hybrid virus progenies resulting from genetic recombination between genetically engineered vaccine viruses and their naturally occurring relatives may possess totally unpredictable characteristics with regard to host preferences and disease-causing potentials. Furthermore, when genetically modified or engineered virus particles break down in the environment, their nuclei acids are released. Appropriate risk management is the key to minimizing any potential risks to humans and environment resulting from the use of these GM vaccines. There is inadequate knowledge to define either the probability of unintended events or the consequences of genetic modifications. The objective of this article is to highlight the limitations in environmental risk assessment and raise awareness of the potential risks involving the use of genetically modified viruses and genetically engineered virus-vector vaccines. PMID:16982535

  1. Comparative Transcriptome Analysis of the Pacific Oyster Crassostrea gigas Characterized by Shell Colors: Identification of Genetic Bases Potentially Involved in Pigmentation

    PubMed Central

    Feng, Dandan; Li, Qi; Yu, Hong; Zhao, Xuelin; Kong, Lingfeng

    2015-01-01

    Background Shell color polymorphisms of Mollusca have contributed to development of evolutionary biology and population genetics, while the genetic bases and molecular mechanisms underlying shell pigmentation are poorly understood. The Pacific oyster (Crassostrea gigas) is one of the most important farmed oysters worldwide. Through successive family selection, four shell color variants (white, golden, black and partially pigmented) of C. gigas have been developed. To elucidate the genetic mechanisms of shell coloration in C. gigas and facilitate the selection of elite oyster lines with desired coloration patterns, differentially expressed genes (DEGs) were identified among the four shell color variants by RNA-seq. Results Digital gene expression generated over fifteen million reads per sample, producing expression data for 28,027 genes. A total number of 2,645 DEGs were identified from pair-wise comparisons, of which 432, 91, 43 and 39 genes specially were up-regulated in white, black, golden and partially pigmented shell of C. gigas, respectively. Three genes of Abca1, Abca3 and Abcb1 which belong to the ATP-binding cassette (ABC) transporters super-families were significantly associated with white shell formation. A tyrosinase transcript (CGI_10008737) represented consistent up-regulated pattern with golden coloration. We proposed that white shell variant of C. gigas could employ “endocytosis” to down-regulate notch level and to prevent shell pigmentation. Conclusion This study discovered some potential shell coloration genes and related molecular mechanisms by the RNA-seq, which would provide foundational information to further study on shell coloration and assist in selective breeding in C. gigas. PMID:26693729

  2. Optimized BlaM-transposon shuttle mutagenesis of Helicobacter pylori allows the identification of novel genetic loci involved in bacterial virulence.

    PubMed

    Odenbreit, S; Till, M; Haas, R

    1996-04-01

    Helicobacter pylori is an important etiologic agent of gastroduodenal disease in humans. In this report, we describe a general genetic approach for the identification of genes encoding exported proteins in H. pylori. The novel TnMax9 mini-blaM transposon was used for insertion mutagenesis of a H. pylori gene library established in Escherichia coli. A total of 192 E. coli clones expressing active beta-lactamase fusion proteins (BlaM+) were obtained, indicating that the corresponding target plasmids carry H. pylori genes encoding putative extracytoplasmic proteins. Natural transformation of H. pylori P1 or P12 using the 192 mutant plasmids resulted in 135 distinct H. pylori mutant strains (70%). Screening of the H. pylori collection of mutant strains allowed the identification of mutant strains impaired in motility, in natural transformation competence and in adherence to gastric epithelial cell lines. Motility mutants could be grouped into distinct classes: (i) mutant strains lacking the major flagellin subunit FlaA and intact flagella (class I); (ii) mutant strains with apparently normal flagella, but reduced motility (class II), and (iii) mutant strains with obviously normal flagella, but completely abolished motility (class III). Two independent mutations that exhibited defects in natural competence for genetic transformation mapped to different genetic loci. In addition, two independent mutant strains were isolated by their failure to bind to the human gastric carcinoma cell line KatoIII. Both mutant strains carried a transposon in the same gene, 0.8 kb apart, and showed decreased autoagglutination when compared to the wild-type strain. PMID:8733234

  3. Genetic transformation of fruit trees: current status and remaining challenges.

    PubMed

    Gambino, Giorgio; Gribaudo, Ivana

    2012-12-01

    Genetic transformation has emerged as a powerful tool for genetic improvement of fruit trees hindered by their reproductive biology and their high levels of heterozygosity. For years, genetic engineering of fruit trees has focussed principally on enhancing disease resistance (against viruses, fungi, and bacteria), although there are few examples of field cultivation and commercial application of these transgenic plants. In addition, over the years much work has been performed to enhance abiotic stress tolerance, to induce modifications of plant growth and habit, to produce marker-free transgenic plants and to improve fruit quality by modification of genes that are crucially important in the production of specific plant components. Recently, with the release of several genome sequences, studies of functional genomics are becoming increasingly important: by modification (overexpression or silencing) of genes involved in the production of specific plant components is possible to uncover regulatory mechanisms associated with the biosynthesis and catabolism of metabolites in plants. This review focuses on the main advances, in recent years, in genetic transformation of the most important species of fruit trees, devoting particular attention to functional genomics approaches and possible future challenges of genetic engineering for these species in the post-genomic era.

  4. Genetic Influences on Learning Disabilties I: Clinical Genetics.

    ERIC Educational Resources Information Center

    Smith, Shelley D.; Pennington, Bruce F.

    1983-01-01

    A discussion of basic genetic principles is followed by a review of selected genetic syndromes involving learning disabilites (such as Noonan Syndrome, Neurofibromatosis, Pheuylketonuria, and cleft lip and palate). Guidelines for securing a genetic evaluation are given. (CL)

  5. Genetics of canine diabetes mellitus: are the diabetes susceptibility genes identified in humans involved in breed susceptibility to diabetes mellitus in dogs?

    PubMed

    Catchpole, Brian; Adams, Jamie P; Holder, Angela L; Short, Andrea D; Ollier, William E R; Kennedy, Lorna J

    2013-02-01

    Diabetes mellitus is a common endocrinopathy in companion animals, characterised by hyperglycaemia, glycosuria and weight loss, resulting from an absolute or relative deficiency in the pancreatic hormone insulin. There are breed differences in susceptibility to diabetes mellitus in dogs, with the Samoyed breed being overrepresented, while Boxers are relatively absent in the UK population of diabetic dogs, suggesting that genetic factors play an important role in determining susceptibility to the disease. A number of genes, linked with susceptibility to diabetes mellitus in humans, are associated with an increased risk of diabetes mellitus in dogs, some of which appear to be relatively breed-specific. Diabetes mellitus in dogs has been associated with major histocompatibility complex (MHC) class II genes (dog leucocyte antigen; DLA), with similar haplotypes and genotypes being identified in the most susceptible breeds. A region containing a variable number of tandem repeats (VNTR) and several polymorphisms have been identified in the canine insulin gene, with some alleles associated with susceptibility or resistance to diabetes mellitus in a breed-specific manner. Polymorphisms in the canine CTLA4 promoter and in other immune response genes are associated with susceptibility to diabetes mellitus in a number of pedigree breeds. Genome wide association studies are currently underway that should shed further light on the genetic factors responsible for the breed profile seen in the diabetic dog population.

  6. Genetic variation in the non-coding genome: Involvement of micro-RNAs and long non-coding RNAs in disease.

    PubMed

    Hrdlickova, Barbara; de Almeida, Rodrigo Coutinho; Borek, Zuzanna; Withoff, Sebo

    2014-10-01

    It has been found that the majority of disease-associated genetic variants identified by genome-wide association studies are located outside of protein-coding regions, where they seem to affect regions that control transcription (promoters, enhancers) and non-coding RNAs that also can influence gene expression. In this review, we focus on two classes of non-coding RNAs that are currently a major focus of interest: micro-RNAs and long non-coding RNAs. We describe their biogenesis, suggested mechanism of action, and discuss how these non-coding RNAs might be affected by disease-associated genetic alterations. The discovery of these alterations has already contributed to a better understanding of the etiopathology of human diseases and yielded insight into the function of these non-coding RNAs. We also provide an overview of available databases, bioinformatics tools, and high-throughput techniques that can be used to study the mechanism of action of individual non-coding RNAs. This article is part of a Special Issue entitled: From Genome to Function.

  7. SM2PH-db: an interactive system for the integrated analysis of phenotypic consequences of missense mutations in proteins involved in human genetic diseases.

    PubMed

    Friedrich, Anne; Garnier, Nicolas; Gagnière, Nicolas; Nguyen, Hoan; Albou, Laurent-Philippe; Biancalana, Valérie; Bettler, Emmanuel; Deléage, Gilbert; Lecompte, Odile; Muller, Jean; Moras, Dino; Mandel, Jean-Louis; Toursel, Thierry; Moulinier, Luc; Poch, Olivier

    2010-02-01

    Understanding how genetic alterations affect gene products at the molecular level represents a first step in the elucidation of the complex relationships between genotypic and phenotypic variations, and is thus a major challenge in the postgenomic era. Here, we present SM2PH-db (http://decrypthon.igbmc.fr/sm2ph), a new database designed to investigate structural and functional impacts of missense mutations and their phenotypic effects in the context of human genetic diseases. A wealth of up-to-date interconnected information is provided for each of the 2,249 disease-related entry proteins (August 2009), including data retrieved from biological databases and data generated from a Sequence-Structure-Evolution Inference in Systems-based approach, such as multiple alignments, three-dimensional structural models, and multidimensional (physicochemical, functional, structural, and evolutionary) characterizations of mutations. SM2PH-db provides a robust infrastructure associated with interactive analysis tools supporting in-depth study and interpretation of the molecular consequences of mutations, with the more long-term goal of elucidating the chain of events leading from a molecular defect to its pathology. The entire content of SM2PH-db is regularly and automatically updated thanks to a computational grid data federation facilities provided in the context of the Decrypthon program.

  8. Genetic analysis of the genes involved in synthesis of the lipopolysaccharide core in Escherichia coli K-12: three operons in the rfa locus.

    PubMed Central

    Roncero, C; Casadaban, M J

    1992-01-01

    The region of the Escherichia coli K-12 chromosome encoding the enzymes responsible for the synthesis of responsible for the synthesis of the lipopolysaccharide (LPS) core has been cloned in vivo by using a mini-Mu vector. This region, formerly known as the rfa locus, comprises 18 kb of DNA between the markers tdh and rpmBG. Results of in vitro mutagenesis of this region with MudII1734 indicate the presence of at least 17 open reading frames or genes, a number considerably higher than expected on the basis of genetic and biochemical studies. Specific insertions in different genes have been recombined into the chromosome, and the mutations have been phenotypically characterized. Complementation analysis indicates that these genes are arranged in three different operons transcribed in opposite directions. A detailed physical map of this region has been constructed on the basis of complementation analysis, fusion protein data, and phenotypic characterizations. Additionally, the role of some genes in the synthesis of LPS has been defined by complementation analysis with known Salmonella typhimurium LPS mutants. The genetic organization of this locus seems to be identical in E. coli K-12 and S. typhimurium. Images PMID:1577693

  9. A Genetic Mosaic Analysis With a Repressible Cell Marker Screen to Identify Genes Involved in Tracheal Cell Migration During Drosophila Air Sac Morphogenesis

    PubMed Central

    Chanut-Delalande, Hélène; Jung, Alain C.; Lin, Li; Baer, Magdalena M.; Bilstein, Andreas; Cabernard, Clemens; Leptin, Maria; Affolter, Markus

    2007-01-01

    Branching morphogenesis of the Drosophila tracheal system relies on the fibroblast growth factor receptor (FGFR) signaling pathway. The Drosophila FGF ligand Branchless (Bnl) and the FGFR Breathless (Btl/FGFR) are required for cell migration during the establishment of the interconnected network of tracheal tubes. However, due to an important maternal contribution of members of the FGFR pathway in the oocyte, a thorough genetic dissection of the role of components of the FGFR signaling cascade in tracheal cell migration is impossible in the embryo. To bypass this shortcoming, we studied tracheal cell migration in the dorsal air sac primordium, a structure that forms during late larval development. Using a mosaic analysis with a repressible cell marker (MARCM) clone approach in mosaic animals, combined with an ethyl methanesulfonate (EMS)-mutagenesis screen of the left arm of the second chromosome, we identified novel genes implicated in cell migration. We screened 1123 mutagenized lines and identified 47 lines displaying tracheal cell migration defects in the air sac primordium. Using complementation analyses based on lethality, mutations in 20 of these lines were genetically mapped to specific genomic areas. Three of the mutants were mapped to either the Mhc or the stam complementation groups. Further experiments confirmed that these genes are required for cell migration in the tracheal air sac primordium. PMID:17603108

  10. Identification of genetic networks involved in the cell injury accompanying endoplasmic reticulum stress induced by bisphenol A in testicular Sertoli cells

    SciTech Connect

    Tabuchi, Yoshiaki . E-mail: ytabu@cts.u-toyama.ac.jp; Takasaki, Ichiro; Kondo, Takashi

    2006-07-07

    To identify detailed mechanisms by which bisphenol A (BPA), an endocrine-disrupting chemical, induces cell injury in mouse testicular Sertoli TTE3 cells, we performed genome-wide microarray and computational gene network analyses. BPA (200 {mu}M) significantly decreased cell viability and simultaneously induced an increase in mRNA levels of HSPA5 and DDIT3, endoplasmic reticulum (ER) stress marker genes. Of the 22,690 probe sets analyzed, BPA down-regulated 661 probe sets and up-regulated 604 probe sets by >2.0-fold. Hierarchical cluster analysis demonstrated nine gene clusters. In decreased gene clusters, two significant genetic networks were associated with cell growth and proliferation and the cell cycle. In increased gene clusters, two significant genetic networks including many basic-region leucine zipper transcription factors were associated with cell death and DNA replication, recombination, and repair. The present results will provide additional novel insights into the detailed molecular mechanisms of cell injury accompanying ER stress induced by BPA in Sertoli cells.

  11. Improving experimental phases for strong reflections prior to density modification

    SciTech Connect

    Uervirojnangkoorn, Monarin; Hilgenfeld, Rolf; Terwilliger, Thomas C.; Read, Randy J.

    2013-09-20

    Experimental phasing of diffraction data from macromolecular crystals involves deriving phase probability distributions. These distributions are often bimodal, making their weighted average, the centroid phase, improbable, so that electron-density maps computed using centroid phases are often non-interpretable. Density modification brings in information about the characteristics of electron density in protein crystals. In successful cases, this allows a choice between the modes in the phase probability distributions, and the maps can cross the borderline between non-interpretable and interpretable. Based on the suggestions by Vekhter [Vekhter (2005), Acta Cryst. D61, 899–902], the impact of identifying optimized phases for a small number of strong reflections prior to the density-modification process was investigated while using the centroid phase as a starting point for the remaining reflections. A genetic algorithm was developed that optimizes the quality of such phases using the skewness of the density map as a target function. Phases optimized in this way are then used in density modification. In most of the tests, the resulting maps were of higher quality than maps generated from the original centroid phases. In one of the test cases, the new method sufficiently improved a marginal set of experimental SAD phases to enable successful map interpretation. Lastly, a computer program,SISA, has been developed to apply this method for phase improvement in macromolecular crystallography.

  12. Improving experimental phases for strong reflections prior to density modification

    DOE PAGES

    Uervirojnangkoorn, Monarin; Hilgenfeld, Rolf; Terwilliger, Thomas C.; Read, Randy J.

    2013-09-20

    Experimental phasing of diffraction data from macromolecular crystals involves deriving phase probability distributions. These distributions are often bimodal, making their weighted average, the centroid phase, improbable, so that electron-density maps computed using centroid phases are often non-interpretable. Density modification brings in information about the characteristics of electron density in protein crystals. In successful cases, this allows a choice between the modes in the phase probability distributions, and the maps can cross the borderline between non-interpretable and interpretable. Based on the suggestions by Vekhter [Vekhter (2005), Acta Cryst. D61, 899–902], the impact of identifying optimized phases for a small number ofmore » strong reflections prior to the density-modification process was investigated while using the centroid phase as a starting point for the remaining reflections. A genetic algorithm was developed that optimizes the quality of such phases using the skewness of the density map as a target function. Phases optimized in this way are then used in density modification. In most of the tests, the resulting maps were of higher quality than maps generated from the original centroid phases. In one of the test cases, the new method sufficiently improved a marginal set of experimental SAD phases to enable successful map interpretation. Lastly, a computer program,SISA, has been developed to apply this method for phase improvement in macromolecular crystallography.« less

  13. Review of Rule Modification in Sport

    PubMed Central

    Arias, Jose L.; Argudo, Francisco M.; Alonso, Jose I.

    2011-01-01

    The goal of this qualitative review was to analyze the state of the bibliography about rule modification in sport. In the literature reviewed, there are few studies of rule modification and related aspects. Most studies omit mentioning the purpose of the modifications, but they do refer to the goals of their analysis (improving players' performance, attracting spectators and athletes, attending to commercial pressure, adapting the sport to children’s needs and interests, preventing injuries). Eighty percent of the studies did not report the outcome of the previous modifications they analyzed. More than half of the studies (60%) achieved the proposed goals. Nearly two-thirds (63.83%) analyzed the effect of rule modification on game actions occurring during the game or through a test. Most of the studies (91.5%) did not consult the participants. Three-fourths of the studies (74.46%) examined the effect of rule modification without any knowledge of a previous analysis or without any previous analysis, and 74.47% studied rule modification related to internal logic. Modifications to be introduced in a sport should be analyzed through a reflective process before their final introduction. The following points should be considered: establishing goals, respecting the basic rules without modifying them, becoming familiar with players’ and coaches’ opinions, determining the effect of the modification on a wide spectrum of variables, elaborating useful proposals for the organizations that are responsible for competitions, using more than one type of data, modifying the internal logic and, preferably, the functional rules, and following some basic stages to consolidate rule modification. Key points Rule modification involves processes that seek change in the game conditions with a certain goal in mind. The rules related to internal logic model the game actions that are characteristic of a sport. Functional rules facilitate achieving the goals. There are few valid research

  14. Genetics Home Reference: mevalonate kinase deficiency

    MedlinePlus

    ... cytoskeleton), gene activity (expression), and protein production and modification. Most MVK gene mutations that cause mevalonate kinase ... What are the different ways in which a genetic condition can be inherited? More about Inheriting Genetic ...

  15. Mechanism-Based Inactivation of Human Cytochrome P450 2B6 by Clopidogrel: Involvement of Both Covalent Modification of Cysteinyl Residue 475 and Loss of HemeS⃞

    PubMed Central

    Zhang, Haoming; Amunugama, Hemali; Ney, Sarah; Cooper, Nyemade

    2011-01-01

    We have investigated the mechanisms by which clopidogrel inactivates human cytochrome P450 2B6 (CYP2B6) in a reconstituted system. It was found that clopidogrel and its thiolactone metabolite, 2-oxo-clopidogrel, both inactivate CYP2B6 in a time- and concentration-dependent manner. On the basis of kinact/KI ratios, clopidogrel is approximately 5 times more efficient than 2-oxo-clopidogrel in inactivating CYP2B6. Analysis of the molecular mass of the CYP2B6 wild-type (WT) protein that had been inactivated by either clopidogrel or 2-oxo-clopidogrel showed an increase in the mass of the protein by ∼350 Da. This increase in the protein mass corresponds to the addition of the active metabolite of clopidogrel to CYP2B6. It is noteworthy that this adduct can be cleaved from the protein matrix by incubation with dithiothreitol, confirming that the active metabolite is linked to a cysteinyl residue of CYP2B6 via a disulfide bond. Peptide mapping of tryptic digests of the inactivated CYP2B6 using electrospray ionization liquid chromatography-tandem mass spectrometry identified Cys475 as the site of covalent modification by the active metabolite. This was further confirmed by the observation that mutation of Cys475 to a serine residue eliminates the formation of the protein adduct and prevents the C475S variant from mechanism-based inactivation by 2-oxo-clopidogrel. However, this mutation did not prevent the C475S variant from being inactivated by clopidogrel. Furthermore, inactivation of both CYP2B6 WT and C475S by clopidogrel, but not by 2-oxo-clopidogrel, led to the loss of the heme, which accounts for most of the loss of the catalytic activity. Collectively, these results suggest that clopidogrel inactivates CYP2B6 primarily through destruction of the heme, whereas 2-oxo-clopidogrel inactivates CYP2B6 through covalent modification of Cys475. PMID:21862689

  16. Liver PPAR{alpha} and UCP2 are involved in the regulation of obesity and lipid metabolism by swim training in genetically obese db/db mice

    SciTech Connect

    Oh, Ki Sook; Kim, Mina; Lee, Jinmi; Kim, Min Jeong; Nam, Youn Shin; Ham, Jung Eun; Shin, Soon Shik; Lee, Chung Moo . E-mail: Chung@sookmyung.ac.kr; Yoon, Michung . E-mail: yoon60@mokwon.ac.kr

    2006-07-07

    Swim training for 6 weeks significantly decreased body weight gain, adipose tissue mass, and adipocyte size in both sexes of genetically obese db/db mice compared with their respective sedentary controls. Swim training also caused significant decreases in serum levels of free fatty acids, triglycerides, and total cholesterol in both sexes of obese mice. Concomitantly, hepatic mRNA levels of peroxisome proliferator-activated receptor {alpha} (PPAR{alpha}) target enzymes responsible for mitochondrial and peroxisomal fatty acid {beta}-oxidation were significantly increased by swim training. Moreover, mRNA levels of uncoupling protein 2 (UCP2) in liver were also markedly increased by swim training. In conclusion, these results suggest that swim training-induced transcriptional activation of hepatic PPAR{alpha} target enzymes and UCP2 may effectively prevent body weight gain, adiposity, and lipid disorders caused by leptin receptor deficiency in both sexes of mice.

  17. Matrix metalloproteinases and genetic mouse models in cancer research: a mini-review.

    PubMed

    Wieczorek, Edyta; Jablonska, Ewa; Wasowicz, Wojciech; Reszka, Edyta

    2015-01-01

    Carcinogenesis is a multistep and also a multifactorial process that involves agents like genetic and environmental factors. Matrix metalloproteinases (MMPs) are major proteolytic enzymes which are involved in cancer cell migration, invasion, and metastasis. Genetic variations in genes encoding the MMPs were shown in human studies to influence cancer risk and phenotypic features of a tumor. The complex role of MMPs seems to be important in the mechanism of carcinogenesis, but it is not well recognized. Rodent studies concentrated particularly on the better understanding of the biological functions of the MMPs and their impact on the pathological process, also through the modification of Mmp genes. This review presents current knowledge and the existing evidence on the importance of selected MMPs in genetic mouse models of cancer and human genetic association studies. Further, this work can be useful for scientists studying the role of the genetic impact of MMPs in carcinogenesis. PMID:25352026

  18. Genetics of xanthan production in Xanthomonas campestris: the xanA and xanB genes are involved in UDP-glucose and GDP-mannose biosynthesis.

    PubMed Central

    Köplin, R; Arnold, W; Hötte, B; Simon, R; Wang, G; Pühler, A

    1992-01-01

    The nucleotide sequence of a 3.4-kb EcoRI-PstI DNA fragment of Xanthomonas campestris pv. campestris revealed two open reading frames, which were designated xanA and xanB. The genes xanA and xanB encode proteins of 448 amino acids (molecular weight of 48,919) and 466 amino acids (molecular weight of 50,873), respectively. These genes were identified by analyzing insertion mutants which were known to be involved in xanthan production. Specific tests for the activities of enzymes involved in the biosynthesis of UDP-glucose and GDP-mannose indicated that the xanA gene product was involved in the biosynthesis of both glucose 1-phosphate and mannose 1-phosphate. The deduced amino acid sequence of xanB showed a significant degree of homology (59%) to the phosphomannose isomerase of Pseudomonas aeruginosa, a key enzyme in the biosynthesis of alginate. Moreover, biochemical analysis and complementation experiments with the Escherichia coli manA fragment revealed that xanB encoded a bifunctional enzyme, phosphomannose isomerase-GDP-mannose pyrophosphorylase. Images PMID:1370280

  19. Role of posttranslational modifications in C. elegans and ascaris spermatogenesis and sperm function.

    PubMed

    Miao, Long; L'Hernault, Steven W

    2014-01-01

    Generally, spermatogenesis and sperm function involve widespread posttranslational modification of regulatory proteins in many different species. Nematode spermatogenesis has been studied in detail, mostly by genetic/molecular genetic techniques in the free-living Caenorhabditis elegans and by biochemistry/cell biology in the pig parasite Ascaris suum. Like other nematodes, both of these species produce sperm that use a form of amoeboid motility termed crawling, and many aspects of spermatogenesis are likely to be similar in both species. Consequently, work in these two nematode species has been largely complementary. Work in C. elegans has identified a number of spermatogenesis-defective genes and, so far, 12 encode enzymes that are implicated as catalysts of posttranslational protein modification. Crawling motility involves extension of a single pseudopod and this process is powered by a unique cytoskeleton composed of Major Sperm Protein (MSP) and accessory proteins, instead of the more widely observed actin. In Ascaris, pseudopod extension and crawling motility can be reconstituted in vitro, and biochemical studies have begun to reveal how posttranslational protein modifications, including phosphorylation, dephosphorylation and proteolysis, participate in these processes. PMID:25030766

  20. Involvement of EARLY BUD-BREAK, an AP2/ERF Transcription Factor Gene, in Bud Break in Japanese Pear (Pyrus pyrifolia Nakai) Lateral Flower Buds: Expression, Histone Modifications and Possible Target Genes.

    PubMed

    Anh Tuan, Pham; Bai, Songling; Saito, Takanori; Imai, Tsuyoshi; Ito, Akiko; Moriguchi, Takaya

    2016-05-01

    In the Japanese pear (Pyrus pyrifolia Nakai) 'Kosui', three developmental stages of lateral flower buds have been proposed to occur during ecodormancy to the flowering phase, i.e. rapid enlargement, sprouting and flowering. Here, we report an APETALA2/ethylene-responsive factor (AP2/ERF) transcription factor gene, named pear EARLY BUD-BREAK (PpEBB), which was highly expressed during the rapid enlargement stage occurring prior to the onset of bud break in flower buds. Gene expression analysis revealed that PpEBB expression was dramatically increased during the rapid enlargement stage in three successive growing seasons. PpEBB transcript levels peaked 1 week prior to onset of bud break in 'Kosui' potted plants treated with hydrogen cyanamide or water under forcing conditions. Chromatin immunoprecipitation-quantitative PCR showed that higher levels of active histone modifications (trimethylation of the histone H3 tail at Lys4) in the 5'-upstream and start codon regions of the PpEBB gene were associated with the induced expression level of PpEBB during the rapid enlargement stage. In addition, we provide evidence that PpEBB may interact with and regulate pear four D-type cyclin (PpCYCD3) genes during bud break in 'Kosui' lateral flower buds. PpEBB significantly increased the promoter activities of four PpCYCD3 genes in a dual-luciferase assay using tobacco leaves. Taken together, our findings uncovered aspects of the bud break regulatory mechanism in the Japanese pear and provided further evidence that the EBB family plays an important role in bud break in perennial plants. PMID:26940832

  1. Involvement of EARLY BUD-BREAK, an AP2/ERF Transcription Factor Gene, in Bud Break in Japanese Pear (Pyrus pyrifolia Nakai) Lateral Flower Buds: Expression, Histone Modifications and Possible Target Genes.

    PubMed

    Anh Tuan, Pham; Bai, Songling; Saito, Takanori; Imai, Tsuyoshi; Ito, Akiko; Moriguchi, Takaya

    2016-05-01

    In the Japanese pear (Pyrus pyrifolia Nakai) 'Kosui', three developmental stages of lateral flower buds have been proposed to occur during ecodormancy to the flowering phase, i.e. rapid enlargement, sprouting and flowering. Here, we report an APETALA2/ethylene-responsive factor (AP2/ERF) transcription factor gene, named pear EARLY BUD-BREAK (PpEBB), which was highly expressed during the rapid enlargement stage occurring prior to the onset of bud break in flower buds. Gene expression analysis revealed that PpEBB expression was dramatically increased during the rapid enlargement stage in three successive growing seasons. PpEBB transcript levels peaked 1 week prior to onset of bud break in 'Kosui' potted plants treated with hydrogen cyanamide or water under forcing conditions. Chromatin immunoprecipitation-quantitative PCR showed that higher levels of active histone modifications (trimethylation of the histone H3 tail at Lys4) in the 5'-upstream and start codon regions of the PpEBB gene were associated with the induced expression level of PpEBB during the rapid enlargement stage. In addition, we provide evidence that PpEBB may interact with and regulate pear four D-type cyclin (PpCYCD3) genes during bud break in 'Kosui' lateral flower buds. PpEBB significantly increased the promoter activities of four PpCYCD3 genes in a dual-luciferase assay using tobacco leaves. Taken together, our findings uncovered aspects of the bud break regulatory mechanism in the Japanese pear and provided further evidence that the EBB family plays an important role in bud break in perennial plants.

  2. Germline Modification and Engineering in Avian Species.

    PubMed

    Lee, Hong Jo; Lee, Hyung Chul; Han, Jae Yong

    2015-09-01

    Production of genome-edited animals using germline-competent cells and genetic modification tools has provided opportunities for investigation of biological mechanisms in various organisms. The recently reported programmed genome editing technology that can induce gene modification at a target locus in an efficient and precise manner facilitates establishment of animal models. In this regard, the demand for genome-edited avian species, which are some of the most suitable model animals due to their unique embryonic development, has also increased. Furthermore, germline chimera production through long-term culture of chicken primordial germ cells (PGCs) has facilitated research on production of genome-edited chickens. Thus, use of avian germline modification is promising for development of novel avian models for research of disease control and various biological mechanisms. Here, we discuss recent progress in genome modification technology in avian species and its applications and future strategies. PMID:26333275

  3. Germline Modification and Engineering in Avian Species

    PubMed Central

    Lee, Hong Jo; Lee, Hyung Chul; Han, Jae Yong

    2015-01-01

    Production of genome-edited animals using germline-competent cells and genetic modification tools has provided opportunities for investigation of biological mechanisms in various organisms. The recently reported programmed genome editing technology that can induce gene modification at a target locus in an efficient and precise manner facilitates establishment of animal models. In this regard, the demand for genome-edited avian species, which are some of the most suitable model animals due to their unique embryonic development, has also increased. Furthermore, germline chimera production through long-term culture of chicken primordial germ cells (PGCs) has facilitated research on production of genome-edited chickens. Thus, use of avian germline modification is promising for development of novel avian models for research of disease control and various biological mechanisms. Here, we discuss recent progress in genome modification technology in avian species and its applications and future strategies. PMID:26333275

  4. The Yeast Hrs1 Gene Is Involved in Positive and Negative Regulation of Transcription and Shows Genetic Characteristics Similar to Sin4 and Gal11

    PubMed Central

    Piruat, J. I.; Chavez, S.; Aguilera, A.

    1997-01-01

    We provide genetic evidence that HRS1/PGD1, a yeast gene previously identified as a suppressor of the hyper-recombination phenotype of hpr1, has positive and negative roles in transcriptional regulation. We have analyzed three differently regulated promoters, GAL1, PHO5 and HSP26, by β-galactosidase assays of lacZ-fused promoters and by Northern analysis of the endogenous genes. Transcription of these promoters was derepressed in hrs1δ mutants under conditions in which it is normally repressed in wild type. Under induced conditions it was either strongly reduced or significantly enhanced depending on the promoter system analyzed. Constitutive transcription was not affected, as determined in ADH1 and TEF2. In addition, Hrs1p was required for mating-factor expression, telomere-linked DNA silencing and DNA supercoiling of plasmids. Furthermore, hrs1δ suppressed Ty-insertion mutations and conferred a Gal(-) phenotype. Many of these phenotypes also result from mutations in GAL11, SIN4 or RGR1, which encode proteins of the RNA polII mediator. We also show that gal11δ and sin4δ partially suppress the hyper-rec phenotype of hpr1 mutants, although to a lesser extent than hrs1δ. Our results provide new evidence for the connection between hpr1δ-induced deletions and transcription. We discuss the possibility that Hrs1p might be a component of the RNA polII transcription machinery. PMID:9409823

  5. Investigating the specific core genetic-and-epigenetic networks of cellular mechanisms involved in human aging in peripheral blood mononuclear cells

    PubMed Central

    Li, Cheng-Wei; Wang, Wen-Hsin; Chen, Bor-Sen

    2016-01-01

    Aging is an inevitable part of life for humans, and slowing down the aging process has become a main focus of human endeavor. Here, we applied a systems biology approach to construct protein-protein interaction networks, gene regulatory networks, and epigenetic networks, i.e. genetic and epigenetic networks (GENs), of elderly individuals and young controls. We then compared these GENs to extract aging mechanisms using microarray data in peripheral blood mononuclear cells, microRNA (miRNA) data, and database mining. The core GENs of elderly individuals and young controls were obtained by applying principal network projection to GENs based on Principal Component Analysis. By comparing the core networks, we identified that to overcome the accumulated mutation of genes in the aging process the transcription factor JUN can be activated by stress signals, including the MAPK signaling, T-cell receptor signaling, and neurotrophin signaling pathways through DNA methylation of BTG3, G0S2, and AP2B1 and the regulations of mir-223 let-7d, and mir-130a. We also address the aging mechanisms in old men and women. Furthermore, we proposed that drugs designed to target these DNA methylated genes or miRNAs may delay aging. A multiple drug combination comprising phenylalanine, cholesterol, and palbociclib was finally designed for delaying the aging process. PMID:26895224

  6. Suggested Involvement of PP1/PP2A Activity and De Novo Gene Expression in Anhydrobiotic Survival in a Tardigrade, Hypsibius dujardini, by Chemical Genetic Approach.

    PubMed

    Kondo, Koyuki; Kubo, Takeo; Kunieda, Takekazu

    2015-01-01

    Upon desiccation, some tardigrades enter an ametabolic dehydrated state called anhydrobiosis and can survive a desiccated environment in this state. For successful transition to anhydrobiosis, some anhydrobiotic tardigrades require pre-incubation under high humidity conditions, a process called preconditioning, prior to exposure to severe desiccation. Although tardigrades are thought to prepare for transition to anhydrobiosis during preconditioning, the molecular mechanisms governing such processes remain unknown. In this study, we used chemical genetic approaches to elucidate the regulatory mechanisms of anhydrobiosis in the anhydrobiotic tardigrade, Hypsibius dujardini. We first demonstrated that inhibition of transcription or translation drastically impaired anhydrobiotic survival, suggesting that de novo gene expression is required for successful transition to anhydrobiosis in this tardigrade. We then screened 81 chemicals and identified 5 chemicals that significantly impaired anhydrobiotic survival after severe desiccation, in contrast to little or no effect on survival after high humidity exposure only. In particular, cantharidic acid, a selective inhibitor of protein phosphatase (PP) 1 and PP2A, exhibited the most profound inhibitory effects. Another PP1/PP2A inhibitor, okadaic acid, also significantly and specifically impaired anhydrobiotic survival, suggesting that PP1/PP2A activity plays an important role for anhydrobiosis in this species. This is, to our knowledge, the first report of the required activities of signaling molecules for desiccation tolerance in tardigrades. The identified inhibitory chemicals could provide novel clues to elucidate the regulatory mechanisms underlying anhydrobiosis in tardigrades.

  7. Suggested Involvement of PP1/PP2A Activity and De Novo Gene Expression in Anhydrobiotic Survival in a Tardigrade, Hypsibius dujardini, by Chemical Genetic Approach

    PubMed Central

    Kondo, Koyuki; Kubo, Takeo; Kunieda, Takekazu

    2015-01-01

    Upon desiccation, some tardigrades enter an ametabolic dehydrated state called anhydrobiosis and can survive a desiccated environment in this state. For successful transition to anhydrobiosis, some anhydrobiotic tardigrades require pre-incubation under high humidity conditions, a process called preconditioning, prior to exposure to severe desiccation. Although tardigrades are thought to prepare for transition to anhydrobiosis during preconditioning, the molecular mechanisms governing such processes remain unknown. In this study, we used chemical genetic approaches to elucidate the regulatory mechanisms of anhydrobiosis in the anhydrobiotic tardigrade, Hypsibius dujardini. We first demonstrated that inhibition of transcription or translation drastically impaired anhydrobiotic survival, suggesting that de novo gene expression is required for successful transition to anhydrobiosis in this tardigrade. We then screened 81 chemicals and identified 5 chemicals that significantly impaired anhydrobiotic survival after severe desiccation, in contrast to little or no effect on survival after high humidity exposure only. In particular, cantharidic acid, a selective inhibitor of protein phosphatase (PP) 1 and PP2A, exhibited the most profound inhibitory effects. Another PP1/PP2A inhibitor, okadaic acid, also significantly and specifically impaired anhydrobiotic survival, suggesting that PP1/PP2A activity plays an important role for anhydrobiosis in this species. This is, to our knowledge, the first report of the required activities of signaling molecules for desiccation tolerance in tardigrades. The identified inhibitory chemicals could provide novel clues to elucidate the regulatory mechanisms underlying anhydrobiosis in tardigrades. PMID:26690982

  8. Molecular genetic evidence for the involvement of a specific polygalacturonase, P2c, in the invasion and spread of Aspergillus flavus in cotton bolls.

    PubMed Central

    Shieh, M T; Brown, R L; Whitehead, M P; Cary, J W; Cotty, P J; Cleveland, T E; Dean, R A

    1997-01-01

    Isolates of Aspergillus flavus can be differentiated based on production of the polygalacturonase P2c. One group of isolates produces P2c, whereas the other group does not. In general, the group that produces P2c causes more damage and spreads to a greater extent in cotton bolls than those isolates that do not produce P2c. To determine whether P2c contributes to disease, the expression of pecA, the gene previously determined to encode P2c, was genetically altered. Adding the pecA gene to a strain previously lacking the gene resulted in the ability to cause significantly more damage to the intercarpellary membrane and the ability spread to a greater extent within the adjacent locule compared to the abilities of a control transformant. Conversely, eliminating the expression of pecA by targeted disruption caused a significant reduction in aggressiveness compared to that of a nondisrupted control transformant. These results provide direct evidence that P2c contributes to the invasion and spread of A. flavus during infection of cotton bolls. However, other factors not evaluated in this study also contribute to aggressiveness. PMID:9293005

  9. Genetic and biochemical interactions involving tricarboxylic acid cycle (TCA) function using a collection of mutants defective in all TCA cycle genes.

    PubMed

    Przybyla-Zawislak, B; Gadde, D M; Ducharme, K; McCammon, M T

    1999-05-01

    The eight enzymes of the tricarboxylic acid (TCA) cycle are encoded by at least 15 different nuclear genes in Saccharomyces cerevisiae. We have constructed a set of yeast strains defective in these genes as part of a comprehensive analysis of the interactions among the TCA cycle proteins. The 15 major TCA cycle genes can be sorted into five phenotypic categories on the basis of their growth on nonfermentable carbon sources. We have previously reported a novel phenotype associated with mutants defective in the IDH2 gene encoding the Idh2p subunit of the NAD+-dependent isocitrate dehydrogenase (NAD-IDH). Null and nonsense idh2 mutants grow poorly on glycerol, but growth can be enhanced by extragenic mutations, termed glycerol suppressors, in the CIT1 gene encoding the TCA cycle citrate synthase and in other genes of oxidative metabolism. The TCA cycle mutant collection was utilized to search for other genes that can suppress idh2 mutants and to identify TCA cycle genes that display a similar suppressible growth phenotype on glycerol. Mutations in 7 TCA cycle genes were capable of functioning as suppressors for growth of idh2 mutants on glycerol. The only other TCA cycle gene to display the glycerol-suppressor-accumulation phenotype was IDH1, which encodes the companion Idh1p subunit of NAD-IDH. These results provide genetic evidence that NAD-IDH plays a unique role in TCA cycle function.

  10. Key Genetic and Epigenetic Mechanisms in Chemical Carcinogenesis.

    PubMed

    Ravegnini, Gloria; Sammarini, Gulia; Hrelia, Patrizia; Angelini, Sabrina

    2015-11-01

    DNA sequence and genetic factors alone cannot fully explain the many processes implicated in diseases initiation and development. It is now well understood that additional factors are involved in a final resulting phenotype. Epigenetic modifications, heritable changes not affecting the DNA sequence, are a key phenomenon at the basis of normal growth and differentiation. However, these can be defective leading to diseases, such as cancer. An increasing body of literature reports the environmental and occupational exposure to a mixture of natural and man-produced substances leading to epigenetic alterations. The identification of key genetic and/or epigenetic events involved in chemical carcinogenesis is an important step towards the discovery of biomarkers that can be used to evaluate the exposure, predict biological effects, and prevent adverse health consequences. Here, we focus on epidemiological studies to review the most recent advances in understanding genetic and epigenetic factors in relation to particulate matter, benzene and polycyclic aromatic hydrocarbons exposure. PMID:26500287

  11. Genetic identification of ACC-RESISTANT2 reveals involvement of LYSINE HISTIDINE TRANSPORTER1 in the uptake of 1-aminocyclopropane-1-carboxylic acid in Arabidopsis thaliana.

    PubMed

    Shin, Kihye; Lee, Sumin; Song, Won-Yong; Lee, Rin-A; Lee, Inhye; Ha, Kyungsun; Koo, Ja-Choon; Park, Soon-Ki; Nam, Hong-Gil; Lee, Youngsook; Soh, Moon-Soo

    2015-03-01

    1-Aminocyclopropane-1-carboxylic acid (ACC) is a biosynthetic precursor of ethylene, a gaseous plant hormone which controls a myriad of aspects of development and stress adaptation in higher plants. Here, we identified a mutant in Arabidopsis thaliana, designated as ACC-resistant2 (are2), displaying a dose-dependent resistance to exogenously applied ACC. Physiological analyses revealed that mutation of are2 impaired various aspects of exogenous ACC-induced ethylene responses, while not affecting sensitivity to other plant hormones during seedling development. Interestingly, the are2 mutant was normally sensitive to gaseous ethylene, compared with the wild type. Double mutant analysis showed that the ethylene-overproducing mutations, eto1 or eto3, and the constitutive ethylene signaling mutation, ctr1 were epistatic to the are2 mutation. These results suggest that the are2 mutant is not defective in ethylene biosynthesis or ethylene signaling per se. Map-based cloning of ARE2 demonstrated that LYSINE HISTIDINE TRANSPORTER1 (LHT1), encoding an amino acid transporter, is the gene responsible. An uptake experiment with radiolabeled ACC indicated that mutations of LHT1 reduced, albeit not completely, uptake of ACC. Further, we performed an amino acid competition assay and found that two amino acids, alanine and glycine, known as substrates of LHT1, could suppress the ACC-induced triple response in a LHT1-dependent way. Taken together, these results provide the first molecular genetic evidence supporting that a class of amino acid transporters including LHT1 takes part in transport of ACC, thereby influencing exogenous ACC-induced ethylene responses in A. thaliana. PMID:25520403

  12. Substance P induces inflammatory responses involving NF-κB in genetically diabetic mice skin fibroblasts co-cultured with macrophages

    PubMed Central

    Ni, Tao; Liu, Yushu; Peng, Yinbo; Li, Ming; Fang, Yong; Yao, Min

    2016-01-01

    Purpose: Delayed wound healing is an intractable complex of diabetes and substance P (SP) is proved to benefit wound healing, whose functioning mechanism remains elusive. This study aims at revealing whether the influence of SP on diabetic wound healing is dependent on inflammatory responses, particularly NF-κB. Methods: Skin fibroblasts of genetically diabetic mice were co-cultured with bone marrow-derived macrophages, and treated with SP, SP + L703,606 (a neurokinin-1 receptor antagonist), or SP + MG132 (an inhibitor of NF-κB). For macrophages, their migration ability was assessed by Transwell experiments, and their M2 polarization was analyzed by flow cytometry and markers for M2 phenotype. Pro-inflammatory factors in the supernatant were detected by enzyme-linked immunosorbent assay. In fibroblasts, the transcription levels of the four pro-inflammatory factors and the protein levels of NF-κB regulators like inhibitor of NF-κB alpha (IκBα) and IκB kinases (IKKs) were monitored by real-time quantitative PCR and western blot, respectively. Results: SP could significantly induce migration to fibroblasts (P<0.01), M2 polarization (P<0.001) and pro-inflammatory factor concentration (P<0.01) in the co-culture system. It also promotes the transcription process of pro-inflammatory factors in fibroblasts (P<0.01), and induce activation of IKKα/β and phosphorylation of IκBα, which caused NF-κB activation. All these effects were reversed if NF-κB was inhibited. Conclusion: The promoting effects of SP on diabetic wound healing was dependent on enhanced inflammatory responses, especially the activation of NF-κB. This study provided evidence for the potential usage of SP in accelerating diabetic wound healing. PMID:27347325

  13. Construction and analysis of regulatory genetic networks in cervical cancer based on involved microRNAs, target genes, transcription factors and host genes.

    PubMed

    Wang, Ning; Xu, Zhiwen; Wang, Kunhao; Zhu, Minghui; Li, Yang

    2014-04-01

    Over recent years, genes and microRNA (miRNA/miR) have been considered as key biological factors in human carcinogenesis. During cancer development, genes may act as multiple identities, including target genes of miRNA, transcription factors and host genes. The present study concentrated on the regulatory networks consisting of the biological factors involved in cervical cancer in order to investigate their features and affect on this specific pathology. Numerous raw data was collected and organized into purposeful structures, and adaptive procedures were defined for application to the prepared data. The networks were therefore built with the factors as basic components according to their interacting associations. The networks were constructed at three levels of interdependency, including a differentially-expressed network, a related network and a global network. Comparisons and analyses were made at a systematic level rather than from an isolated gene or miRNA. Critical hubs were extracted in the core networks and notable features were discussed, including self-adaption feedback regulation. The present study expounds the pathogenesis from a novel point of view and is proposed to provide inspiration for further investigation and therapy.

  14. Genetic Susceptibility to Cardiac and Digestive Clinical Forms of Chronic Chagas Disease: Involvement of the CCR5 59029 A/G Polymorphism

    PubMed Central

    de Oliveira, Amanda Priscila; Bernardo, Cássia Rubia; Camargo, Ana Vitória da Silveira; Ronchi, Luiz Sérgio; Borim, Aldenis Albaneze; Brandão de Mattos, Cinara Cássia; de Campos Júnior, Eumildo; Castiglioni, Lílian; Netinho, João Gomes; Cavasini, Carlos Eugênio; Bestetti, Reinaldo Bulgarelli; de Mattos, Luiz Carlos

    2015-01-01

    The clinical manifestations of chronic Chagas disease include the cardiac form of the disease and the digestive form. Not all the factors that act in the variable clinical course of this disease are known. This study investigated whether the CCR5Δ32 (rs333) and CCR5 59029 A/G (promoter region—rs1799987) polymorphisms of the CCR5 gene are associated with different clinical forms of chronic Chagas disease and with the severity of left ventricular systolic dysfunction in patients with chronic Chagas heart disease (CCHD). The antibodies anti-T. cruzi were identified by ELISA. PCR and PCR-RFLP were used to identify the CCR5Δ32 and CCR5 59029 A/G polymorphisms. The chi-square test was used to compare variables between groups. There was a higher frequency of the AA genotype in patients with CCHD compared with patients with the digestive form of the disease and the control group. The results also showed a high frequency of the AG genotype in patients with the digestive form of the disease compared to the other groups. The results of this study show that the CCR5Δ32 polymorphism does not seem to influence the different clinical manifestations of Chagas disease but there is involvement of the CCR5 59029 A/G polymorphism in susceptibility to the different forms of chronic Chagas disease. Besides, these polymorphisms do not influence left ventricular systolic dysfunction in patients with CCHD. PMID:26599761

  15. A reversed genetic approach reveals the coenzyme specificity and other catalytic properties of three enzymes putatively involved in anaerobic oxidation of methane with sulfate.

    PubMed

    Kojima, Hisaya; Moll, Johanna; Kahnt, Jörg; Fukui, Manabu; Shima, Seigo

    2014-11-01

    Consortia of anaerobic methanotrophic (ANME) archaea and delta-proteobacteria anaerobically oxidize methane coupled to sulfate reduction to sulfide. The metagenome of ANME-1 archaea contains genes homologous to genes otherwise only found in methanogenic archaea, and transcription of some of these genes in ANME-1 cells has been shown. We now have heterologously expressed three of these genes in Escherichia coli, namely those homologous to genes for formylmethanofuran : tetrahydromethanopterin formyltransferase, methenyltetrahydromethanopterin cyclohydrolase (Mch) and coenzyme F420 -dependent methylenetetrahydromethanopterin dehydrogenase (Mtd), and have characterized the overproduced enzymes with respect to their coenzyme specificity and other catalytic properties. The three enzymes from ANME-1 were found to catalyse the same reactions and with similar specific activities using identical coenzymes as the respective enzymes in methanogenic archaea, the apparent Km for their substrates being in the same concentration range. The results support the proposal that anaerobic oxidation of methane to CO₂in ANME involves the same enzymes and coenzymes as CO₂reduction to methane in methanogenic archaea. Interestingly, the activity of Mch and the stability of Mtd from ANME-1 were found to be dependent on the presence of 0.5-1.0 M potassium phosphate, which suggested that ANME-1 archaea contain high concentrations of lyotropic salts, presumably as compatible solutes.

  16. Genetic linkage facilitates cloning of Ert-m regulating plant architecture in barley and identified a strong candidate of Ant1 involved in anthocyanin biosynthesis.

    PubMed

    Zakhrabekova, Shakhira; Dockter, Christoph; Ahmann, Katharina; Braumann, Ilka; Gough, Simon P; Wendt, Toni; Lundqvist, Udda; Mascher, Martin; Stein, Nils; Hansson, Mats

    2015-08-01

    The erectoides-m anthocyanin-less 1 (ert-m ant1) double mutants are among the very few examples of induced double mutants in barley. From phenotypic observations of mutant plants it is known that the Ert-m gene product regulates plant architecture whereas the Ant1 gene product is involved in anthocyanin biosynthesis. We used a near-isogenic line of the cultivar Bowman, BW316 (ert-m.34), to create four F2-mapping populations by crosses to the barley cultivars Barke, Morex, Bowman and Quench. We phenotyped and genotyped 460 plants, allowing the ert-m mutation to be mapped to an interval of 4.7 cM on the short arm of barley chromosome 7H. Bioinformatic searches identified 21 candidate gene models in the mapped region. One gene was orthologous to a regulator of Arabidopsis thaliana plant architecture, ERECTA, encoding a leucine-rich repeat receptor-like kinase. Sequencing of HvERECTA in barley ert-m mutant accessions identified severe DNA changes in 15 mutants, including full gene deletions in ert-m.40 and ert-m.64. Both deletions, additionally causing anthocyanin deficiency, were found to stretch over a large region including two putative candidate genes for the anthocyanin biosynthesis locus Ant1. Analyses of ert-m and ant1 single- and double-deletion mutants suggest Ant1 as a closely linked gene encoding a R2R3 myeloblastosis transcription factor.

  17. Genetic Susceptibility to Cardiac and Digestive Clinical Forms of Chronic Chagas Disease: Involvement of the CCR5 59029 A/G Polymorphism.

    PubMed

    de Oliveira, Amanda Priscila; Bernardo, Cássia Rubia; Camargo, Ana Vitória da Silveira; Ronchi, Luiz Sérgio; Borim, Aldenis Albaneze; de Mattos, Cinara Cássia Brandão; de Campos Júnior, Eumildo; Castiglioni, Lílian; Netinho, João Gomes; Cavasini, Carlos Eugênio; Bestetti, Reinaldo Bulgarelli; de Mattos, Luiz Carlos

    2015-01-01

    The clinical manifestations of chronic Chagas disease include the cardiac form of the disease and the digestive form. Not all the factors that act in the variable clinical course of this disease are known. This study investigated whether the CCR5Δ32 (rs333) and CCR5 59029 A/G (promoter region--rs1799987) polymorphisms of the CCR5 gene are associated with different clinical forms of chronic Chagas disease and with the severity of left ventricular systolic dysfunction in patients with chronic Chagas heart disease (CCHD). The antibodies anti-T. cruzi were identified by ELISA. PCR and PCR-RFLP were used to identify the CCR5Δ32 and CCR5 59029 A/G polymorphisms. The chi-square test was used to compare variables between groups. There was a higher frequency of the AA genotype in patients with CCHD compared with patients with the digestive form of the disease and the control group. The results also showed a high frequency of the AG genotype in patients with the digestive form of the disease compared to the other groups. The results of this study show that the CCR5Δ32 polymorphism does not seem to influence the different clinical manifestations of Chagas disease but there is involvement of the CCR5 59029 A/G polymorphism in susceptibility to the different forms of chronic Chagas disease. Besides, these polymorphisms do not influence left ventricular systolic dysfunction in patients with CCHD.

  18. Medium Modification of Vector Mesons

    SciTech Connect

    Chaden Djalali, Michael Paolone, Dennis Weygand, Michael H. Wood, Rakhsha Nasseripour

    2011-03-01

    The theory of the strong interaction, Quantum Chromodynamics (QCD), has been remarkably successful in describing high-energy and short-distance-scale experiments involving quarks and gluons. However, applying QCD to low energy and large-distance scale experiments has been a major challenge. Various QCD-inspired models predict a partial restoration of chiral symmetry in nuclear matter with modifications of the properties of hadrons from their free-space values. Measurable changes such as a shift in mass and/or a change of width are predicted at normal nuclear density. Photoproduction of vector mesons off nuclei have been performed at different laboratories. The properties of the ρ, ω and φ mesons are investigated either directly by measuring their mass spectra or indirectly through transparency ratios. The latest results regarding medium modifications of the vector mesons in the nuclear medium will be discussed.

  19. On Involvement.

    ERIC Educational Resources Information Center

    Greene, Michael B.

    Involvement Ratings In Settings (IRIS), a multi-dimensional non-verbal scale of involvement adaptable to a time-sampling method of data collection, was constructed with the aid of the videotapes of second-grade Follow Through classrooms made by CCEP. Scales were defined through observations of involved and alienated behavior, and the IRIS was…

  20. Modifications of mitosis

    SciTech Connect

    1993-12-31

    Chapter 15, discusses modifications of mitosis, including endoreduplication, polyteny, endomitosis, C-mitosis, restitution, amitosis and multipolar mitoses. Apart from multipolar mitosis, all other mitotic modifications are characterized by an absent or defective spindle, and in most cases these result in the duplication of the chromosome number. 29 refs., 4 figs.

  1. Permit application modifications

    SciTech Connect

    1995-11-01

    This document contains the Permit Application Modifications for the Y-12 Industrial Landfill V site on the Oak Ridge Reservation. These modifications include the assessment of stability of the proposed Landfill V under static and loading conditions. Analyses performed include the general slope stability, veneer stability of the bottom liner and cover system, and a liquefaction potential assessment of the foundation soils.

  2. Lifestyle modifications for GDM.

    PubMed

    Dhingra, Atul; Ahuja, Kamlesh

    2016-09-01

    Prevalence of gestational diabetes mellitus (GDM) is increasing worldwide more so in Southeast Asian countries like India and Pakistan. 1 GDM is associated with various adverse foetal and maternal effects. The management of GDM aims at reducing blood glucose to reduce maternal and foetal morbidity and mortality. Various studies have shown that lifestyle modifications are an important tool for reducing blood glucose levels in patients with GDM. Lifestyle modifications consist of dietary modifications and daily physical activity. Dietary modifications aim to achieve glycaemic control by providing adequate calories to the mother and foetus. Exercise is an obvious adjunct to dietary modifications for management of GDM. Therefore the purpose of this review is to summarize the benefits of lifestyle interventions in patients with GDM. PMID:27582149

  3. Behavioral genetics and taste

    PubMed Central

    Boughter, John D; Bachmanov, Alexander A

    2007-01-01

    This review focuses on behavioral genetic studies of sweet, umami, bitter and salt taste responses in mammals. Studies involving mouse inbred strain comparisons and genetic analyses, and their impact on elucidation of taste receptors and transduction mechanisms are discussed. Finally, the effect of genetic variation in taste responsiveness on complex traits such as drug intake is considered. Recent advances in development of genomic resources make behavioral genetics a powerful approach for understanding mechanisms of taste. PMID:17903279

  4. Regulation of chromatin by histone modifications

    PubMed Central

    Bannister, Andrew J; Kouzarides, Tony

    2011-01-01

    Chromatin is not an inert structure, but rather an instructive DNA scaffold that can respond to external cues to regulate the many uses of DNA. A principle component of chromatin that plays a key role in this regulation is the modification of histones. There is an ever-growing list of these modifications and the complexity of their action is only just beginning to be understood. However, it is clear that histone modifications play fundamental roles in most biological processes that are involved in the manipulation and expression of DNA. Here, we describe the known histone modifications, define where they are found genomically and discuss some of their functional consequences, concentrating mostly on transcription where the majority of characterisation has taken place. PMID:21321607

  5. Genetics Home Reference: multiple mitochondrial dysfunctions syndrome

    MedlinePlus

    ... to function; this protein is involved in the modification of additional proteins that aid in energy production ... also known as the oxoglutarate dehydrogenase complex). This modification is also critical to the function of the ...

  6. Welcome to Neurology: Genetics.

    PubMed

    Pulst, Stefan M

    2015-06-01

    The powers of human genetics and genetic technologies have transformed the complexities of neurology and neuroscience at the basic, translational, and now also the clinical level. We have left an era of black and white views of causative genetic variation and are entering a period of more than 50 shades of grey, fascinated with DNA variants that increase or decrease risk, epigenetic modification, and an unexpectedly large number of variants of unknown or potentially pathogenic significance. Loss-of-function alleles and even complete human gene knockouts for certain genes appear to be compatible with a normal phenotype. PMID:27066541

  7. Jackson Creek Spillway modifications

    SciTech Connect

    Freitas, M.J.; Young, D.J.; McCloud, B.J.

    1995-12-31

    The Jackson Creek Spillway in Amador County, California has been modified in response to issues raised during the Federal Energy Regulatory Commission (FERC) mandated 5-year safety inspections. The calculated factors of safety for the Jackson Creek Spillway, under the probable maximum flood (PMF) and maximum credible earthquake (MCE) loading conditions, were below levels considered acceptable by the FERC and modifications to the structure were required. Woodward-Clyde Consultants, under contract to the East Bay Municipal Utility District (EBMUD), designed the modifications and in the summer and fall of 1994 the modifications to the Jackson Creek Spillway were successfully constructed with both FERC and California Division of Safety of Dams (DSOD) approval. This paper will summarize the design and construction issues, and discuss the lessons learned during modification of this 67-year-old structure.

  8. Transgenerational inheritance or resetting of stress-induced epigenetic modifications: two sides of the same coin

    PubMed Central

    Tricker, Penny J.

    2015-01-01

    The transgenerational inheritance of stress-induced epigenetic modifications is still controversial. Despite several examples of defense “priming” and induced genetic rearrangements, the involvement and persistence of transgenerational epigenetic modifications is not known to be general. Here I argue that non-transmission of epigenetic marks through meiosis may be regarded as an epigenetic modification in itself, and that we should understand the implications for plant evolution in the context of both selection for and selection against transgenerational epigenetic memory. Recent data suggest that both epigenetic inheritance and resetting are mechanistically directed and targeted. Stress-induced epigenetic modifications may buffer against DNA sequence-based evolution to maintain plasticity, or may form part of plasticity’s adaptive potential. To date we have tended to concentrate on the question of whether and for how long epigenetic memory persists. I argue that we should now re-direct our question to investigate the differences between where it persists and where it does not, to understand the higher order evolutionary methods in play and their contribution. PMID:26442015

  9. Asbestos surface provides a niche for oxidative modification.

    PubMed

    Nagai, Hirotaka; Ishihara, Toshikazu; Lee, Wen-Hua; Ohara, Hiroki; Okazaki, Yasumasa; Okawa, Katsuya; Toyokuni, Shinya

    2011-12-01

    Asbestos is a potent carcinogen associated with increased risks of malignant mesothelioma and lung cancer in humans. Although the mechanism of carcinogenesis remains elusive, the physicochemical characteristics of asbestos play a role in the progression of asbestos-induced diseases. Among these characteristics, a high capacity to adsorb and accommodate biomolecules on its abundant surface area has been linked to cellular and genetic toxicity. Several previous studies identified asbestos-interacting proteins. Here, with the use of matrix-assisted laser desorption ionization-time of flight mass spectrometry, we systematically identified proteins from various lysates that adsorbed to the surface of commercially used asbestos and classified them into the following groups: chromatin/nucleotide/RNA-binding proteins, ribosomal proteins, cytoprotective proteins, cytoskeleton-associated proteins, histones and hemoglobin. The surfaces of crocidolite and amosite, two iron-rich types of asbestos, caused more protein scissions and oxidative modifications than that of chrysotile by in situ-generated 4-hydroxy-2-nonenal. In contrast, we confirmed the intense hemolytic activity of chrysotile and found that hemoglobin attached to chrysotile, but not silica, can work as a catalyst to induce oxidative DNA damage. This process generates 8-hydroxy-2'-deoxyguanosine and thus corroborates the involvement of iron in the carcinogenicity of chrysotile. This evidence demonstrates that all three types of asbestos adsorb DNA and specific proteins, providing a niche for oxidative modification via catalytic iron. Therefore, considering the affinity of asbestos for histones/DNA and the internalization of asbestos into mesothelial cells, our results suggest a novel hypothetical mechanism causing genetic alterations during asbestos-induced carcinogenesis.

  10. Human Rights and Behavior Modification

    ERIC Educational Resources Information Center

    Roos, Philip

    1974-01-01

    Criticisms of behavior modification, which charge that it violates ethical and legal principles, are discussed and reasons are presented to explain behavior modification's susceptibility to attack. (GW)

  11. Somatic cell reprogramming-free generation of genetically modified pigs.

    PubMed

    Tanihara, Fuminori; Takemoto, Tatsuya; Kitagawa, Eri; Rao, Shengbin; Do, Lanh Thi Kim; Onishi, Akira; Yamashita, Yukiko; Kosugi, Chisato; Suzuki, Hitomi; Sembon, Shoichiro; Suzuki, Shunichi; Nakai, Michiko; Hashimoto, Masakazu; Yasue, Akihiro; Matsuhisa, Munehide; Noji, Sumihare; Fujimura, Tatsuya; Fuchimoto, Dai-Ichiro; Otoi, Takeshige

    2016-09-01

    Genetically modified pigs for biomedical applications have been mainly generated using the somatic cell nuclear transfer technique; however, this approach requires complex micromanipulation techniques and sometimes increases the risks of both prenatal and postnatal death by faulty epigenetic reprogramming of a donor somatic cell nucleus. As a result, the production of genetically modified pigs has not been widely applied. We provide a simple method for CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 gene editing in pigs that involves the introduction of Cas9 protein and single-guide RNA into in vitro fertilized zygotes by electroporation. The use of gene editing by electroporation of Cas9 protein (GEEP) resulted in highly efficient targeted gene disruption and was validated by the efficient production of Myostatin mutant pigs. Because GEEP does not require the complex methods associated with micromanipulation for somatic reprogramming, it has the potential for facilitating the genetic modification of pigs.

  12. Somatic cell reprogramming-free generation of genetically modified pigs.

    PubMed

    Tanihara, Fuminori; Takemoto, Tatsuya; Kitagawa, Eri; Rao, Shengbin; Do, Lanh Thi Kim; Onishi, Akira; Yamashita, Yukiko; Kosugi, Chisato; Suzuki, Hitomi; Sembon, Shoichiro; Suzuki, Shunichi; Nakai, Michiko; Hashimoto, Masakazu; Yasue, Akihiro; Matsuhisa, Munehide; Noji, Sumihare; Fujimura, Tatsuya; Fuchimoto, Dai-Ichiro; Otoi, Takeshige

    2016-09-01

    Genetically modified pigs for biomedical applications have been mainly generated using the somatic cell nuclear transfer technique; however, this approach requires complex micromanipulation techniques and sometimes increases the risks of both prenatal and postnatal death by faulty epigenetic reprogramming of a donor somatic cell nucleus. As a result, the production of genetically modified pigs has not been widely applied. We provide a simple method for CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 gene editing in pigs that involves the introduction of Cas9 protein and single-guide RNA into in vitro fertilized zygotes by electroporation. The use of gene editing by electroporation of Cas9 protein (GEEP) resulted in highly efficient targeted gene disruption and was validated by the efficient production of Myostatin mutant pigs. Because GEEP does not require the complex methods associated with micromanipulation for somatic reprogramming, it has the potential for facilitating the genetic modification of pigs. PMID:27652340

  13. Somatic cell reprogramming-free generation of genetically modified pigs

    PubMed Central

    Tanihara, Fuminori; Takemoto, Tatsuya; Kitagawa, Eri; Rao, Shengbin; Do, Lanh Thi Kim; Onishi, Akira; Yamashita, Yukiko; Kosugi, Chisato; Suzuki, Hitomi; Sembon, Shoichiro; Suzuki, Shunichi; Nakai, Michiko; Hashimoto, Masakazu; Yasue, Akihiro; Matsuhisa, Munehide; Noji, Sumihare; Fujimura, Tatsuya; Fuchimoto, Dai-ichiro; Otoi, Takeshige

    2016-01-01

    Genetically modified pigs for biomedical applications have been mainly generated using the somatic cell nuclear transfer technique; however, this approach requires complex micromanipulation techniques and sometimes increases the risks of both prenatal and postnatal death by faulty epigenetic reprogramming of a donor somatic cell nucleus. As a result, the production of genetically modified pigs has not been widely applied. We provide a simple method for CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 gene editing in pigs that involves the introduction of Cas9 protein and single-guide RNA into in vitro fertilized zygotes by electroporation. The use of gene editing by electroporation of Cas9 protein (GEEP) resulted in highly efficient targeted gene disruption and was validated by the efficient production of Myostatin mutant pigs. Because GEEP does not require the complex methods associated with micromanipulation for somatic reprogramming, it has the potential for facilitating the genetic modification of pigs.

  14. Somatic cell reprogramming-free generation of genetically modified pigs

    PubMed Central

    Tanihara, Fuminori; Takemoto, Tatsuya; Kitagawa, Eri; Rao, Shengbin; Do, Lanh Thi Kim; Onishi, Akira; Yamashita, Yukiko; Kosugi, Chisato; Suzuki, Hitomi; Sembon, Shoichiro; Suzuki, Shunichi; Nakai, Michiko; Hashimoto, Masakazu; Yasue, Akihiro; Matsuhisa, Munehide; Noji, Sumihare; Fujimura, Tatsuya; Fuchimoto, Dai-ichiro; Otoi, Takeshige

    2016-01-01

    Genetically modified pigs for biomedical applications have been mainly generated using the somatic cell nuclear transfer technique; however, this approach requires complex micromanipulation techniques and sometimes increases the risks of both prenatal and postnatal death by faulty epigenetic reprogramming of a donor somatic cell nucleus. As a result, the production of genetically modified pigs has not been widely applied. We provide a simple method for CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 gene editing in pigs that involves the introduction of Cas9 protein and single-guide RNA into in vitro fertilized zygotes by electroporation. The use of gene editing by electroporation of Cas9 protein (GEEP) resulted in highly efficient targeted gene disruption and was validated by the efficient production of Myostatin mutant pigs. Because GEEP does not require the complex methods associated with micromanipulation for somatic reprogramming, it has the potential for facilitating the genetic modification of pigs. PMID:27652340

  15. Interpreting thelanguage of histone and DNA modifications

    PubMed Central

    Rothbart, Scott B.; Strahl, Brian D.

    2014-01-01

    A major mechanism regulating the accessibility and function of eukaryotic genomes are the covalent modifications to DNA and histone proteins that dependably package our genetic information inside the nucleus of every cell. Formally postulated over a decade ago, it is becoming increasingly clear that post-translational modifications (PTMs) on histones act singly and in combination to form a language or ‘code’ that is read by specialized proteins to facilitate downstream functions in chromatin. Underappreciated at the time was the level of complexity harbored both within histone PTMs and their combinations, as well as within the proteins that read and interpret the language. In addition to histone PTMs, newly-identified DNA modifications that can recruit specific effector proteins has raised further awareness that histone PTMs operate within a broader language of epigenetic modifications to orchestrate the dynamic functions associated with chromatin. Here, we highlight key recent advances in our understanding of the epigenetic language encompassing histone and DNA modifications and foreshadow challenges that lie ahead as we continue our quest to decipher the fundamental mechanisms of chromatin regulation. PMID:24631868

  16. Molecular and genetic analysis of a region of plasmid pCF10 containing positive control genes and structural genes encoding surface proteins involved in pheromone-inducible conjugation in Enterococcus faecalis.

    PubMed Central

    Kao, S M; Olmsted, S B; Viksnins, A S; Gallo, J C; Dunny, G M

    1991-01-01

    Exposure of Enterococcus faecalis cells carrying the tetracycline resistance plasmid pCF10 to the heptapeptide pheromone cCF10 results in an increase in conjugal transfer frequency by as much as 10(6)-fold. Pheromone-induced donor cells also express at least two plasmid-encoded surface proteins, the 130-kDa Sec 10 protein, which is involved in surface exclusion, and the 150-kDa Asc10 protein, which has been associated with the formation of mating aggregates. Previous subcloning and transposon mutagenesis studies indicated that the adjacent EcoRI c (7.5 kb) and e (4.5 kb) fragments of pCF10 encode the structural genes for these proteins and that the EcoRI c fragment also encodes at least two regulatory genes involved in activation of the expression of the genes encoding Asc10 and Sec10. In this paper, the results of physical and genetic analysis of this region of pCF10, along with the complete DNA sequences of the EcoRI c and e fragments, are reported. The results of the genetic studies indicate the location of the structural genes for the surface proteins and reveal important features of their transcription. In addition, we provide evidence here and in the accompanying paper (S. B. Olmsted, S.-M. Kao, L. J. van Putte, J. C. Gallo, and G. M. Dunny, J. Bacteriol. 173:7665-7672, 1991) for a role of Asc10 in mating aggregate formation. The data also reveal a complex positive control system that acts at distances of at least 3 to 6 kb to activate expression of Asc10. DNA sequence analysis presented here reveals the positions of a number of specific genes, termed prg (pheromone-responsive genes) in this region of pCF10. The genes mapped include prgA (encoding Sec10) and prgB (encoding Asc10), as well as four putative regulatory genes, prgX, -R, -S, and -T. Although the predicted amino acid sequences of Sec10 and Asc10 have some structural features in common with a number of surface proteins of gram-positive cocci, and the Asc10 sequence is highly similar to that of a

  17. Nuclear lactate dehydrogenase modulates histone modification in human hepatocytes

    SciTech Connect

    Castonguay, Zachary; Auger, Christopher; Thomas, Sean C.; Chahma, M’hamed; Appanna, Vasu D.

    2014-11-07

    Highlights: • Nuclear LDH is up-regulated under oxidative stress. • SIRT1 is co-immunoprecipitated bound to nuclear LDH. • Nuclear LDH is involved in histone deacetylation and epigenetics. - Abstract: It is becoming increasingly apparent that the nucleus harbors metabolic enzymes that affect genetic transforming events. Here, we describe a nuclear isoform of lactate dehydrogenase (nLDH) and its ability to orchestrate histone deacetylation by controlling the availability of nicotinamide adenine dinucleotide (NAD{sup +}), a key ingredient of the sirtuin-1 (SIRT1) deacetylase system. There was an increase in the expression of nLDH concomitant with the presence of hydrogen peroxide (H{sub 2}O{sub 2}) in the culture medium. Under oxidative stress, the NAD{sup +} generated by nLDH resulted in the enhanced deacetylation of histones compared to the control hepatocytes despite no discernable change in the levels of SIRT1. There appeared to be an intimate association between nLDH and SIRT1 as these two enzymes co-immunoprecipitated. The ability of nLDH to regulate epigenetic modifications by manipulating NAD{sup +} reveals an intricate link between metabolism and the processing of genetic information.

  18. Genetic Screening for Employment Purposes.

    ERIC Educational Resources Information Center

    Olian, Judy D.

    1984-01-01

    Discusses genetic screening in the employment context, which involves identification of individuals hypersusceptible to toxins in the work environment. Examines the status of genetic screening devices against standard testing and legal criteria. (LLL)

  19. Modifications of gravity.

    PubMed

    Skordis, Constantinos

    2011-12-28

    General relativity (GR) is a phenomenologically successful theory that rests on firm foundations, but has not been tested on cosmological scales. The deep mystery of dark energy (and possibly even the requirement of cold dark matter (CDM)) has increased the need for testing modifications to GR, as the inference of such otherwise undetected fluids depends crucially on the theory of gravity. Here, I discuss a general scheme for constructing consistent and covariant modifications to the Einstein equations. This framework is such that there is a clear connection between the modification and the underlying field content that produces it. I argue that this is mandatory for distinguishing modifications of gravity from conventional fluids. I give a non-trivial example, a simple metric-based modification of the fluctuation equations for which the background is exact ΛCDM, but differs from it in the perturbations. I show how this can be generalized and solved in terms of two arbitrary functions. Finally, I discuss future prospects and directions of research.

  20. Laboratory Course on "Streptomyces" Genetics and Secondary Metabolism

    ERIC Educational Resources Information Center

    Siitonen, Vilja; Räty, Kaj; Metsä-Ketelä, Mikko

    2016-01-01

    The "'Streptomyces' genetics and secondary metabolism" laboratory course gives an introduction to the versatile soil dwelling Gram-positive bacteria "Streptomyces" and their secondary metabolism. The course combines genetic modification of "Streptomyces"; growing of the strain and protoplast preparation, plasmid…

  1. Genetics & sport: bioethical concerns.

    PubMed

    Miah, Andy

    2012-12-01

    This paper provides an overview of the ethical issues pertaining to the use of genetic insights and techniques in sport. Initially, it considers a range of scientific findings that have stimulated debate about the ethical issues associated with genetics applied to sport. It also outlines some of the early policy responses to these discoveries from world leading sports organizations, along with knowledge about actual use of gene technologies in sport. Subsequently, it considers the challenges with distinguishing between therapeutic use and human enhancement within genetic science, which is a particularly important issue for the world of sport. Next, particular attention is given to the use of genetic information, which raises questions about the legitimacy and reliability of genetic tests, along with the potential public value of having DNA databanks to economize in health care. Finally, the ethics of gene transfer are considered, inviting questions into the values of sport and humanity. It argues that, while gene modification may seem conceptually similar to other forms of doping, the requirements upon athletes are such that new forms of enhancement become increasingly necessary to discover. Insofar as genetic science is able to create safer, more effective techniques of human modification, then it may be an appealing route through which to modify athletes to safeguard the future of elite sports as enterprises of human excellence. PMID:22830450

  2. Genetics & sport: bioethical concerns.

    PubMed

    Miah, Andy

    2012-12-01

    This paper provides an overview of the ethical issues pertaining to the use of genetic insights and techniques in sport. Initially, it considers a range of scientific findings that have stimulated debate about the ethical issues associated with genetics applied to sport. It also outlines some of the early policy responses to these discoveries from world leading sports organizations, along with knowledge about actual use of gene technologies in sport. Subsequently, it considers the challenges with distinguishing between therapeutic use and human enhancement within genetic science, which is a particularly important issue for the world of sport. Next, particular attention is given to the use of genetic information, which raises questions about the legitimacy and reliability of genetic tests, along with the potential public value of having DNA databanks to economize in health care. Finally, the ethics of gene transfer are considered, inviting questions into the values of sport and humanity. It argues that, while gene modification may seem conceptually similar to other forms of doping, the requirements upon athletes are such that new forms of enhancement become increasingly necessary to discover. Insofar as genetic science is able to create safer, more effective techniques of human modification, then it may be an appealing route through which to modify athletes to safeguard the future of elite sports as enterprises of human excellence.

  3. Parent Involvement.

    ERIC Educational Resources Information Center

    LaCrosse, Ed

    The paper discusses the rationale and guidelines for parent involvement in HCEEP (Handicapped Children's Early Education Program) projects. Ways of assessing parents' needs are reviewed, as are four types of services to meet the identified needs: parent education, direct participation, parent counseling, and parent provided programs. Materials and…

  4. Adding 'epi-' to behaviour genetics: implications for animal domestication.

    PubMed

    Jensen, Per

    2015-01-01

    In this review, it is argued that greatly improved understanding of domestication may be gained from extending the field of behaviour genetics to also include epigenetics. Domestication offers an interesting framework of rapid evolutionary changes caused by well-defined selection pressures. Behaviour is an important phenotype in this context, as it represents the primary means of response to environmental challenges. An overview is provided of the evidence for genetic involvement in behavioural control and the presently used methods for finding so-called behaviour genes. This shows that evolutionary changes in behaviour are to a large extent correlated to changes in patterns of gene expression, which brings epigenetics into the focus. This area is concerned with the mechanisms controlling the timing and extent of gene expression, and a lot of focus has been placed on methylation of cytosine in promoter regions, usually associated with genetic downregulation. The review considers the available evidence that environmental input, for example stress, can modify methylation and other epigenetic marks and subsequently affect behaviour. Furthermore, several studies are reviewed, demonstrating that acquired epigenetic modifications can be inherited and cause trans-generational behaviour changes. In conclusion, epigenetics may signify a new paradigm in this respect, as it shows that genomic modifications can be caused by environmental signals, and random mutations in DNA sequence are therefore not the only sources of heritable genetic variation.

  5. Potential adverse health effects of genetically modified crops.

    PubMed

    Bakshi, Anita

    2003-01-01

    Genetically modified crops have the potential to eliminate hunger and starvation in millions of people, especially in developing countries because the genetic modification can produce large amounts of foods that are more nutritious. Large quantities are produced because genetically modified crops are more resistant to pests and drought. They also contain greater amounts of nutrients, such as proteins and vitamins. However, there are concerns about the safety of genetically modified crops. The concerns are that they may contain allergenic substances due to introduction of new genes into crops. Another concern is that genetic engineering often involves the use of antibiotic-resistance genes as "selectable markers" and this could lead to production of antibiotic-resistant bacterial strains that are resistant to available antibiotics. This would create a serious public health problem. The genetically modified crops might contain other toxic substances (such as enhanced amounts of heavy metals) and the crops might not be "substantially equivalent" in genome, proteome, and metabolome compared with unmodified crops. Another concern is that genetically modified crops may be less nutritious; for example, they might contain lower amounts of phytoestrogens, which protect against heart disease and cancer. The review of available literature indicates that the genetically modified crops available in the market that are intended for human consumption are generally safe; their consumption is not associated with serious health problems. However, because of potential for exposure of a large segment of human population to genetically modified foods, more research is needed to ensure that the genetically modified foods are safe for human consumption.

  6. Genetic Variation and Atherosclerosis

    PubMed Central

    Biros, Erik; Karan, Mirko; Golledge, Jonathan

    2008-01-01

    A family history of atherosclerosis is independently associated with an increased incidence of cardiovascular events. The genetic factors underlying the importance of inheritance in atherosclerosis are starting to be understood. Genetic variation, such as mutations or common polymorphisms has been shown to be involved in modulation of a range of risk factors, such as plasma lipoprotein levels, inflammation and vascular calcification. This review presents examples of present studies of the role of genetic polymorphism in atherosclerosis. PMID:19424482

  7. Ion Beam Modification of Materials

    SciTech Connect

    Averback, B; de la Rubia, T D; Felter, T E; Hamza, A V; Rehn, L E

    2005-10-10

    This volume contains the proceedings of the 14th International Conference on Ion Beam Modification of Materials, IBMM 2004, and is published by Elsevier-Science Publishers as a special issue of Nuclear Instruments and Methods B. The conference series is the major international forum to present and discuss recent research results and future directions in the field of ion beam modification, synthesis and characterization of materials. The first conference in the series was held in Budapest, Hungary, 1978, and subsequent conferences were held every two years at locations around the Globe, most recently in Japan, Brazil, and the Netherlands. The series brings together physicists, materials scientists, and ion beam specialists from all over the world. The official conference language is English. IBMM 2004 was held on September 5-10, 2004. The focus was on materials science involving both basic ion-solid interaction processes and property changes occurring either during or subsequent to ion bombardment and ion beam processing in relation to materials and device applications. Areas of research included Nanostructures, Multiscale Modeling, Patterning of Surfaces, Focused Ion Beams, Defects in Semiconductors, Insulators and Metals, Cluster Beams, Radiation Effects in Materials, Photonic Devices, Ion Implantation, Ion Beams in Biology and Medicine including New Materials, Imaging, and Treatment.

  8. Biblical behavior modification.

    PubMed

    Lasure, L C; Mikulas, W L

    1996-07-01

    Although we may have formalized and systematized the field of behavior modification in the last few decades, people around the world have been using behavioral change strategies throughout history. Premack's (1965) theory of reinforcement is often called "Grandma's rule" because grandmothers have long been using it (e.g. You must finish your vegetables before you may go out and play). Franks (1969, p. 4), in one of the first behavioral texts, gave historical examples from China, Turkey, France, and Italy. Knapp and Shodahl (1974) showed how Benjamin Franklin used behavior modification. And de Silva (1984, 1985) gave examples of behavior modification by the Buddha and other early Buddhists. Conspicuously absent from our literature are examples from the Judeo-Christian tradition. In this paper, we provide a number of behavior modification examples from the Bible (New International Version). Footnotes provide references for many more examples. In the discussion, we explore implications for education and therapy. Examples are grouped by the following categories: operant conditioning, respondent conditioning, modeling, and cognitive interventions. However, the Biblical examples, like contemporary case studies, do not always fall neatly into discrete categories. They often are a combination, particularly operant and respondent conditioning interweaving.

  9. Diet Modification for Hyperlipidemia

    PubMed Central

    Mann, Heather D.; Piotrowski, Pamela

    1992-01-01

    Hyperlipidemia is a major risk factor associated with cardiovascular disease. Dietary modification is effective in achieving and maintaining improved serum lipid levels. Nutritional care provided by a dietitian includes individual dietary and lifestyle assessment, formulating an appropriate dietary regimen, education, and follow-up assessments. PMID:21221406

  10. Behavior Modification with Children

    ERIC Educational Resources Information Center

    Brown, Daniel G.

    1972-01-01

    The author urges wider use of positive reinforcement theories in helping emotionally disturbed and mentally handicapped children. Underlining the influence of environment on behavior, he also notes that behavior modification programs utilize fewer trained personnel more effectively and, like Tennessee's Re-Education Treatment, allow for therapy in…

  11. Behavior Modification for Teachers.

    ERIC Educational Resources Information Center

    Maryland Univ., College Park. Coll. of Education.

    The Active Student Participation Program (ASPP), developed by the Department of Counseling and Personnel Services in the University of Maryland College of Education, was designed to disseminate knowledge of behavior modification principles. The course has been taught since the fall of 1971, and approximately 81 in-service teachers have completed…

  12. Toy Modification Note. Revised.

    ERIC Educational Resources Information Center

    Vanderheiden, Gregg C.; And Others

    Described are toy modifications which enable handicapped individuals to operate battery-powered toys. A battery interrupter is explained as a device which fits between the batteries in a toy and provides the ability to have a separate on-off switch which can be custom designed to fit a handicapped user's needs. Construction and use of three types…

  13. THE MODIFICATION OF STUTTERING.

    ERIC Educational Resources Information Center

    BRUTTEN, EUGENE J.; SHOEMAKER, DONALD J.

    INTENDED FOR BOTH THE COLLEGE STUDENT AND THE PROFESSIONAL SPEECH PATHOLOGIST, THE BOOK PRESENTS CURRENT LEARNING THEORIES CONCERNING STUTTERING, DATA IMPORTANT TO THE THEORIES, AND A 2-PROCESS THEORY OF LEARNING FOR THEORETICAL INTEGRATION OF THE DATA ON STUTTERING AND FOR THERAPEUTIC MODIFICATION. INFORMATION PRESENTED ABOUT BEHAVIORISTIC…

  14. Metabolic regulation of histone post-translational modifications

    PubMed Central

    Fan, Jing; Krautkramer, Kimberly A.; Feldman, Jessica L.; Denu, John M.

    2015-01-01

    Histone post-translational modifications regulate transcription and other DNA-templated functions. This process is dynamically regulated by specific modifying enzymes whose activities require metabolites that either serve as co-substrates or act as activators/inhibitors. Therefore, metabolism can influence histone modification by changing local concentrations of key metabolites. Physiologically, the epigenetic response to metabolism is important for nutrient sensing and environment adaption. In pathologic states, the connection between metabolism and histone modification mediates epigenetic abnormality in complex disease. In this review, we summarize recent studies of the molecular mechanisms involved in metabolic regulation of histone modifications and discuss their biological significance. PMID:25562692

  15. Acute blockade of the Caenorhabditis elegans dopamine transporter DAT-1 by the mammalian norepinephrine transporter inhibitor nisoxetine reveals the influence of genetic modifications of dopamine signaling in vivo.

    PubMed

    Bermingham, Daniel P; Hardaway, J Andrew; Snarrenberg, Chelsea L; Robinson, Sarah B; Folkes, Oakleigh M; Salimando, Greg J; Jinnah, Hussain; Blakely, Randy D

    2016-09-01

    Modulation of neurotransmission by the catecholamine dopamine (DA) is conserved across phylogeny. In the nematode Caenorhabditis elegans, excess DA signaling triggers Swimming-Induced Paralysis (Swip), a phenotype first described in animals with loss of function mutations in the presynaptic DA transporter (dat-1). Swip has proven to be a phenotype suitable for the identification of novel dat-1 mutations as well as the identification of novel genes that impact DA signaling. Pharmacological manipulations can also induce Swip, though the reagents employed to date lack specificity and potency, limiting their use in evaluation of dat-1 expression and function. Our lab previously established the mammalian norepinephrine transporter (NET) inhibitor nisoxetine to be a potent antagonist of DA uptake conferred by DAT-1 following heterologous expression. Here we demonstrate the ability of low (μM) concentrations of nisoxetine to trigger Swip within minutes of incubation, with paralysis dependent on DA release and signaling, and non-additive with Swip triggered by dat-1 deletion. Using nisoxetine in combination with genetic mutations that impact DA release, we further demonstrate the utility of the drug for demonstrating contributions of presynaptic DA receptors and ion channels to Swip. Together, these findings reveal nisoxetine as a powerful reagent for monitoring multiple dimensions of DA signaling in vivo, thus providing a new resource that can be used to evaluate contributions of dat-1 and other genes linked to DA signaling without the potential for compensations that attend constitutive genetic mutations.

  16. Difficult Decisions: Genetic Screening.

    ERIC Educational Resources Information Center

    Slesnick, Irwin L.; Parakh, Jal S.

    1988-01-01

    Discusses the issue of mandatory genetic screening. Poses the question of the benefits, drawbacks, and motives involved. Provides a discussion activity to be used in high school biology classes consisting of a hypothetical situation and several questions. (CW)

  17. Medical genetics

    SciTech Connect

    Nora, J.J.; Fraser, F.C.

    1989-01-01

    This book presents a discussion of medical genetics for the practitioner treating or counseling patients with genetic disease. It includes a discussion of the relationship of heredity and diseases, the chromosomal basis for heredity, gene frequencies, and genetics of development and maldevelopment. The authors also focus on teratology, somatic cell genetics, genetics and cancer, genetics of behavior.

  18. Genetic and molecular alterations associated with oral squamous cell cancer (Review).

    PubMed

    Pérez-Sayáns, Mario; Somoza-Martín, José M; Barros-Angueira, Francisco; Reboiras-López, María D; Gándara Rey, José M; García-García, Abel

    2009-12-01

    The development of oral squamous cell cancer (OSCC) is a multistep process involving the accumulation of multiple genetic alterations modulated by genetic pre-disposition and environmental influences such as tobacco and alcohol use, chronic inflammation, and viral infections. All of these factors can lead to a wide range of genetic and molecular alterations that can be detected using a range of molecular studies. The alterations mostly affect two large groups of genes: oncogenes and tumor suppressor genes, which can be either inactivated or overexpressed through mutations, loss of heterozygosity, deletions, or epigenetic modifications such as methylation. Other molecules that are closely associated with tumor pathogenesis and prognosis also exist and warrant further study. Important advances in molecular biology are helping to shed light on oral cancer and thus aiding in the early diagnosis and development of new personalized treatment approaches. The purpose of the review is to explore the genetic and molecular alterations associated with OSCC.

  19. Effect of phytate reduction of sorghum, through genetic modification, on iron and zinc availability as assessed by an in vitro dialysability bioaccessibility assay, Caco-2 cell uptake assay, and suckling rat pup absorption model.

    PubMed

    Kruger, Johanita; Taylor, John R N; Du, Xiaogu; De Moura, Fabiana F; Lönnerdal, Bo; Oelofse, André

    2013-11-15

    Improved iron and zinc availability from sorghum, a commonly consumed staple, will benefit many malnourished communities in rural Africa burdened with high prevalence of iron and zinc deficiency. This research compared the effect of genetic phytate reduction in sorghum on iron and zinc bioaccessibility and uptake measured by in vitro dialysability and Caco-2 cell uptake assays to that of iron and zinc absorption measured by a suckling rat pup model. The phytate reduction (80-86%) in these sorghums significantly increased zinc availability. The Caco-2 cell method, but not the dialysability assay, proved useful in estimating zinc absorption. The measured increase in iron availability differed between the methods, possibly due to the effect of varying mineral (Ca, Fe, Zn, P) contents of the sorghums. This effect was most prominent in the iron uptake results. More research is needed to determine the effect of naturally occurring variations in mineral contents of sorghum on the iron uptake by Caco-2 cells.

  20. Genetic evidence for the involvement of the S-layer protein gene sap and the sporulation genes spo0A, spo0B, and spo0F in Phage AP50c infection of Bacillus anthracis.

    PubMed

    Plaut, Roger D; Beaber, John W; Zemansky, Jason; Kaur, Ajinder P; George, Matroner; Biswas, Biswajit; Henry, Matthew; Bishop-Lilly, Kimberly A; Mokashi, Vishwesh; Hannah, Ryan M; Pope, Robert K; Read, Timothy D; Stibitz, Scott; Calendar, Richard; Sozhamannan, Shanmuga

    2014-03-01

    In order to better characterize the Bacillus anthracis typing phage AP50c, we designed a genetic screen to identify its bacterial receptor. Insertions of the transposon mariner or targeted deletions of the structural gene for the S-layer protein Sap and the sporulation genes spo0A, spo0B, and spo0F in B. anthracis Sterne resulted in phage resistance with concomitant defects in phage adsorption and infectivity. Electron microscopy of bacteria incubated with AP50c revealed phage particles associated with the surface of bacilli of the Sterne strain but not with the surfaces of Δsap, Δspo0A, Δspo0B, or Δspo0F mutants. The amount of Sap in the S layer of each of the spo0 mutant strains was substantially reduced compared to that of the parent strain, and incubation of AP50c with purified recombinant Sap led to a substantial reduction in phage activity. Phylogenetic analysis based on whole-genome sequences of B. cereus sensu lato strains revealed several closely related B. cereus and B. thuringiensis strains that carry sap genes with very high similarities to the sap gene of B. anthracis. Complementation of the Δsap mutant in trans with the wild-type B. anthracis sap or the sap gene from either of two different B. cereus strains that are sensitive to AP50c infection restored phage sensitivity, and electron microscopy confirmed attachment of phage particles to the surface of each of the complemented strains. Based on these data, we postulate that Sap is involved in AP50c infectivity, most likely acting as the phage receptor, and that the spo0 genes may regulate synthesis of Sap and/or formation of the S layer.

  1. Genetic Evidence for the Involvement of the S-Layer Protein Gene sap and the Sporulation Genes spo0A, spo0B, and spo0F in Phage AP50c Infection of Bacillus anthracis

    PubMed Central

    Beaber, John W.; Zemansky, Jason; Kaur, Ajinder P.; George, Matroner; Biswas, Biswajit; Henry, Matthew; Bishop-Lilly, Kimberly A.; Mokashi, Vishwesh; Hannah, Ryan M.; Pope, Robert K.; Read, Timothy D.; Stibitz, Scott; Calendar, Richard; Sozhamannan, Shanmuga

    2014-01-01

    In order to better characterize the Bacillus anthracis typing phage AP50c, we designed a genetic screen to identify its bacterial receptor. Insertions of the transposon mariner or targeted deletions of the structural gene for the S-layer protein Sap and the sporulation genes spo0A, spo0B, and spo0F in B. anthracis Sterne resulted in phage resistance with concomitant defects in phage ads