Mediator binds to boundaries of chromosomal interaction domains and to proteins involved in DNA looping, RNA metabolism, chromatin remodeling, and actin assembly.

Science.gov (United States)

Chereji, Razvan V; Bharatula, Vasudha; Elfving, Nils; Blomberg, Jeanette; Larsson, Miriam; Morozov, Alexandre V; Broach, James R; Björklund, Stefan

2017-09-06

Mediator is a multi-unit molecular complex that plays a key role in transferring signals from transcriptional regulators to RNA polymerase II in eukaryotes. We have combined biochemical purification of the Saccharomyces cerevisiae Mediator from chromatin with chromatin immunoprecipitation in order to reveal Mediator occupancy on DNA genome-wide, and to identify proteins interacting specifically with Mediator on the chromatin template. Tandem mass spectrometry of proteins in immunoprecipitates of mediator complexes revealed specific interactions between Mediator and the RSC, Arp2/Arp3, CPF, CF 1A and Lsm complexes in chromatin. These factors are primarily involved in chromatin remodeling, actin assembly, mRNA 3'-end processing, gene looping and mRNA decay, but they have also been shown to enter the nucleus and participate in Pol II transcription. Moreover, we have found that Mediator, in addition to binding Pol II promoters, occupies chromosomal interacting domain (CID) boundaries and that Mediator in chromatin associates with proteins that have been shown to interact with CID boundaries, such as Sth1, Ssu72 and histone H4. This suggests that Mediator plays a significant role in higher-order genome organization. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Mid-range adiabatic wireless energy transfer via a mediator coil

International Nuclear Information System (INIS)

Rangelov, A.A.; Vitanov, N.V.

2012-01-01

A technique for efficient mid-range wireless energy transfer between two coils via a mediator coil is proposed. By varying the coil frequencies, three resonances are created: emitter–mediator (EM), mediator–receiver (MR) and emitter–receiver (ER). If the frequency sweeps are adiabatic and such that the EM resonance precedes the MR resonance, the energy flows sequentially along the chain emitter–mediator–receiver. If the MR resonance precedes the EM resonance, then the energy flows directly from the emitter to the receiver via the ER resonance; then the losses from the mediator are suppressed. This technique is robust against noise, resonant constraints and external interferences. - Highlights: ► Efficient and robust mid-range wireless energy transfer via a mediator coil. ► The adiabatic energy transfer is analogous to adiabatic passage in quantum optics. ► Wireless energy transfer is insensitive to any resonant constraints. ► Wireless energy transfer is insensitive to noise in the neighborhood of the coils.

Polyoxometalate active charge-transfer material for mediated redox flow battery

Energy Technology Data Exchange (ETDEWEB)

Anderson, Travis Mark; Hudak, Nicholas; Staiger, Chad; Pratt, Harry

2017-01-17

Redox flow batteries including a half-cell electrode chamber coupled to a current collecting electrode are disclosed herein. In a general embodiment, a separator is coupled to the half-cell electrode chamber. The half-cell electrode chamber comprises a first redox-active mediator and a second redox-active mediator. The first redox-active mediator and the second redox-active mediator are circulated through the half-cell electrode chamber into an external container. The container includes an active charge-transfer material. The active charge-transfer material has a redox potential between a redox potential of the first redox-active mediator and a redox potential of the second redox-active mediator. The active charge-transfer material is a polyoxometalate or derivative thereof. The redox flow battery may be particularly useful in energy storage solutions for renewable energy sources and for providing sustained power to an electrical grid.

Speeding-up exchange-mediated saturation transfer experiments by Fourier transform

Energy Technology Data Exchange (ETDEWEB)

Carneiro, Marta G.; Reddy, Jithender G.; Griesinger, Christian; Lee, Donghan, E-mail: dole@nmr.mpibpc.mpg.de [Max-Planck Institute for Biophysical chemistry, Department of NMR-based Structural Biology (Germany)

2015-11-15

Protein motions over various time scales are crucial for protein function. NMR relaxation dispersion experiments play a key role in explaining these motions. However, the study of slow conformational changes with lowly populated states remained elusive. The recently developed exchange-mediated saturation transfer experiments allow the detection and characterization of such motions, but require extensive measurement time. Here we show that, by making use of Fourier transform, the total acquisition time required to measure an exchange-mediated saturation transfer profile can be reduced by twofold in case that one applies linear prediction. In addition, we demonstrate that the analytical solution for R{sub 1}ρ experiments can be used for fitting the exchange-mediated saturation transfer profile. Furthermore, we show that simultaneous analysis of exchange-mediated saturation transfer profiles with two different radio-frequency field strengths is required for accurate and precise characterization of the exchange process and the exchanging states.

Speeding-up exchange-mediated saturation transfer experiments by Fourier transform

International Nuclear Information System (INIS)

Carneiro, Marta G.; Reddy, Jithender G.; Griesinger, Christian; Lee, Donghan

2015-01-01

Protein motions over various time scales are crucial for protein function. NMR relaxation dispersion experiments play a key role in explaining these motions. However, the study of slow conformational changes with lowly populated states remained elusive. The recently developed exchange-mediated saturation transfer experiments allow the detection and characterization of such motions, but require extensive measurement time. Here we show that, by making use of Fourier transform, the total acquisition time required to measure an exchange-mediated saturation transfer profile can be reduced by twofold in case that one applies linear prediction. In addition, we demonstrate that the analytical solution for R 1 ρ experiments can be used for fitting the exchange-mediated saturation transfer profile. Furthermore, we show that simultaneous analysis of exchange-mediated saturation transfer profiles with two different radio-frequency field strengths is required for accurate and precise characterization of the exchange process and the exchanging states

Deep functional analysis of synII, a 770-kilobase synthetic yeast chromosome.

Science.gov (United States)

Shen, Yue; Wang, Yun; Chen, Tai; Gao, Feng; Gong, Jianhui; Abramczyk, Dariusz; Walker, Roy; Zhao, Hongcui; Chen, Shihong; Liu, Wei; Luo, Yisha; Müller, Carolin A; Paul-Dubois-Taine, Adrien; Alver, Bonnie; Stracquadanio, Giovanni; Mitchell, Leslie A; Luo, Zhouqing; Fan, Yanqun; Zhou, Baojin; Wen, Bo; Tan, Fengji; Wang, Yujia; Zi, Jin; Xie, Zexiong; Li, Bingzhi; Yang, Kun; Richardson, Sarah M; Jiang, Hui; French, Christopher E; Nieduszynski, Conrad A; Koszul, Romain; Marston, Adele L; Yuan, Yingjin; Wang, Jian; Bader, Joel S; Dai, Junbiao; Boeke, Jef D; Xu, Xun; Cai, Yizhi; Yang, Huanming

2017-03-10

Here, we report the successful design, construction, and characterization of a 770-kilobase synthetic yeast chromosome II (synII). Our study incorporates characterization at multiple levels-including phenomics, transcriptomics, proteomics, chromosome segregation, and replication analysis-to provide a thorough and comprehensive analysis of a synthetic chromosome. Our Trans-Omics analyses reveal a modest but potentially relevant pervasive up-regulation of translational machinery observed in synII, mainly caused by the deletion of 13 transfer RNAs. By both complementation assays and SCRaMbLE (synthetic chromosome rearrangement and modification by loxP -mediated evolution), we targeted and debugged the origin of a growth defect at 37°C in glycerol medium, which is related to misregulation of the high-osmolarity glycerol response. Despite the subtle differences, the synII strain shows highly consistent biological processes comparable to the native strain. Copyright © 2017, American Association for the Advancement of Science.

Horizontal Gene Transfers in Mycoplasmas (Mollicutes).

Science.gov (United States)

Citti, C; Dordet-Frisoni, E; Nouvel, L X; Kuo, C H; Baranowski, E

2018-04-12

The class Mollicutes (trivial name "mycoplasma") is composed of wall-less bacteria with reduced genomes whose evolution was long thought to be only driven by gene losses. Recent evidences of massive horizontal gene transfer (HGT) within and across species provided a new frame to understand the successful adaptation of these minimal bacteria to a broad range of hosts. Mobile genetic elements are being identified in a growing number of mycoplasma species, but integrative and conjugative elements (ICEs) are emerging as pivotal in HGT. While sharing common traits with other bacterial ICEs, such as their chromosomal integration and the use of a type IV secretion system to mediate horizontal dissemination, mycoplasma ICEs (MICEs) revealed unique features: their chromosomal integration is totally random and driven by a DDE recombinase related to the Mutator-like superfamily. Mycoplasma conjugation is not restricted to ICE transmission, but also involves the transfer of large chromosomal fragments that generates progenies with mosaic genomes, nearly every position of chromosome being mobile. Mycoplasmas have thus developed efficient ways to gain access to a considerable reservoir of genetic resources distributed among a vast number of species expanding the concept of minimal cell to the broader context of flowing information.

Neurogenin 3 Mediates Sex Chromosome Effects on the Generation of Sex Differences in Hypothalamic Neuronal Development

Directory of Open Access Journals (Sweden)

Maria Julia Scerbo

2014-07-01

Full Text Available The organizational action of testosterone during critical periods of development is the cause of numerous sex differences in the brain. However, sex differences in neuritogenesis have been detected in primary neuronal hypothalamic cultures prepared before the peak of testosterone production by fetal testis. In the present study we assessed the hypothesis of that cell-autonomous action of sex chromosomes can differentially regulate the expression of the neuritogenic gene neurogenin 3 (Ngn3 in male and female hypothalamic neurons, generating sex differences in neuronal development. Neuronal cultures were prepared from male and female E14 mouse hypothalami, before the fetal peak of testosterone. Female neurons showed enhanced neuritogenesis and higher expression of Ngn3 than male neurons. The silencing of Ngn3 abolished sex differences in neuritogenesis, decreasing the differentiation of female neurons. The sex difference in Ngn3 expression was determined by sex chromosomes, as demonstrated using the four core genotypes mouse model, in which a spontaneous deletion of the testis-determining gene Sry from the Y chromosome was combined with the insertion of the Sry gene onto an autosome. In addition, the expression of Ngn3, which is also known to mediate the neuritogenic actions of estradiol, was increased in the cultures treated with the hormone, but only in those from male embryos. Furthermore, the hormone reversed the sex differences in neuritogenesis promoting the differentiation of male neurons. These findings indicate that Ngn3 mediates both cell-autonomous actions of sex chromosomes and hormonal effects on neuritogenesis.

Efficient site-specific integration in Plasmodium falciparum chromosomes mediated by mycobacteriophage Bxb1 integrase.

Science.gov (United States)

Nkrumah, Louis J; Muhle, Rebecca A; Moura, Pedro A; Ghosh, Pallavi; Hatfull, Graham F; Jacobs, William R; Fidock, David A

2006-08-01

Here we report an efficient, site-specific system of genetic integration into Plasmodium falciparum malaria parasite chromosomes. This is mediated by mycobacteriophage Bxb1 integrase, which catalyzes recombination between an incoming attP and a chromosomal attB site. We developed P. falciparum lines with the attB site integrated into the glutaredoxin-like cg6 gene. Transfection of these attB(+) lines with a dual-plasmid system, expressing a transgene on an attP-containing plasmid together with a drug resistance gene and the integrase on a separate plasmid, produced recombinant parasites within 2 to 4 weeks that were genetically uniform for single-copy plasmid integration. Integrase-mediated recombination resulted in proper targeting of parasite proteins to intra-erythrocytic compartments, including the apicoplast, a plastid-like organelle. Recombinant attB x attP parasites were genetically stable in the absence of drug and were phenotypically homogeneous. This system can be exploited for rapid genetic integration and complementation analyses at any stage of the P. falciparum life cycle, and it illustrates the utility of Bxb1-based integrative recombination for genetic studies of intracellular eukaryotic organisms.

Chromosome painting reveals asynaptic full alignment of homologs and HIM-8-dependent remodeling of X chromosome territories during Caenorhabditis elegans meiosis.

Science.gov (United States)

Nabeshima, Kentaro; Mlynarczyk-Evans, Susanna; Villeneuve, Anne M

2011-08-01

During early meiotic prophase, a nucleus-wide reorganization leads to sorting of chromosomes into homologous pairs and to establishing associations between homologous chromosomes along their entire lengths. Here, we investigate global features of chromosome organization during this process, using a chromosome painting method in whole-mount Caenorhabditis elegans gonads that enables visualization of whole chromosomes along their entire lengths in the context of preserved 3D nuclear architecture. First, we show that neither spatial proximity of premeiotic chromosome territories nor chromosome-specific timing is a major factor driving homolog pairing. Second, we show that synaptonemal complex-independent associations can support full lengthwise juxtaposition of homologous chromosomes. Third, we reveal a prominent elongation of chromosome territories during meiotic prophase that initiates prior to homolog association and alignment. Mutant analysis indicates that chromosome movement mediated by association of chromosome pairing centers (PCs) with mobile patches of the nuclear envelope (NE)-spanning SUN-1/ZYG-12 protein complexes is not the primary driver of territory elongation. Moreover, we identify new roles for the X chromosome PC (X-PC) and X-PC binding protein HIM-8 in promoting elongation of X chromosome territories, separable from their role(s) in mediating local stabilization of pairing and association of X chromosomes with mobile SUN-1/ZYG-12 patches. Further, we present evidence that HIM-8 functions both at and outside of PCs to mediate chromosome territory elongation. These and other data support a model in which synapsis-independent elongation of chromosome territories, driven by PC binding proteins, enables lengthwise juxtaposition of chromosomes, thereby facilitating assessment of their suitability as potential pairing partners.

HIM-8 binds to the X chromosome pairing center and mediates chromosome-specific meiotic synapsis.

Science.gov (United States)

Phillips, Carolyn M; Wong, Chihunt; Bhalla, Needhi; Carlton, Peter M; Weiser, Pinky; Meneely, Philip M; Dernburg, Abby F

2005-12-16

The him-8 gene is essential for proper meiotic segregation of the X chromosomes in C. elegans. Here we show that loss of him-8 function causes profound X chromosome-specific defects in homolog pairing and synapsis. him-8 encodes a C2H2 zinc-finger protein that is expressed during meiosis and concentrates at a site on the X chromosome known as the meiotic pairing center (PC). A role for HIM-8 in PC function is supported by genetic interactions between PC lesions and him-8 mutations. HIM-8 bound chromosome sites associate with the nuclear envelope (NE) throughout meiotic prophase. Surprisingly, a point mutation in him-8 that retains both chromosome binding and NE localization fails to stabilize pairing or promote synapsis. These observations indicate that stabilization of homolog pairing is an active process in which the tethering of chromosome sites to the NE may be necessary but is not sufficient.

HIM-8 binds to the X chromosome pairing center and mediates chromosome-specific meiotic synapsis

International Nuclear Information System (INIS)

Phillips, Carolyn M.; Wong, Chihunt; Bhalla, Needhi; Carlton, Peter M.; Weiser, Pinky; Meneely, Philip M.; Dernburg, Abby F.

2005-01-01

The him-8 gene is essential for proper meiotic segregation of the X chromosomes in C. elegans. Here we show that loss of him-8 function causes profound X-chromosome-specific defects in homolog pairing and synapsis.him-8 encodes a C2H2 zinc finger protein that is expressed during meiosis and concentrates at a site on the X chromosome known as themeiotic Pairing Center (PC). A role for HIM-8 in PC function is supported by genetic interactions between PC lesions and him-8 mutations. HIM-8-bound chromosome sites associate with the nuclear envelope (NE)throughout meiotic prophase. Surprisingly, a point mutation in him-8 that retains both chromosome binding and NE localization fails to stabilize pairing or promote synapsis. These observations indicate that stabilization of homolog pairing is an active process in which the tethering of chromosome sites to the NE may be necessary but is not sufficient

Modelling of bubble-mediated gas transfer: Fundamental principles and a laboratory test

NARCIS (Netherlands)

Woolf, D.K.; Leifer, I.S.; Nightingale, P.D.; Rhee, T.S.; Bowyer, P.; Caulliez, G.; Leeuw, G. de; Larsen, S.E.; Liddicoat, M.; Baker, J.; Andreae, M.O.

2007-01-01

The air-water exchange of gases can be substantially enhanced by wave breaking and specifically by bubble-mediated transfer. A feature of bubble-mediated transfer is the additional pressure on bubbles resulting from the hydrostatic forces on a submerged bubble and from surface tension and curvature.

Chromosome Painting Reveals Asynaptic Full Alignment of Homologs and HIM-8–Dependent Remodeling of X Chromosome Territories during Caenorhabditis elegans Meiosis

Science.gov (United States)

Nabeshima, Kentaro; Mlynarczyk-Evans, Susanna; Villeneuve, Anne M.

2011-01-01

During early meiotic prophase, a nucleus-wide reorganization leads to sorting of chromosomes into homologous pairs and to establishing associations between homologous chromosomes along their entire lengths. Here, we investigate global features of chromosome organization during this process, using a chromosome painting method in whole-mount Caenorhabditis elegans gonads that enables visualization of whole chromosomes along their entire lengths in the context of preserved 3D nuclear architecture. First, we show that neither spatial proximity of premeiotic chromosome territories nor chromosome-specific timing is a major factor driving homolog pairing. Second, we show that synaptonemal complex-independent associations can support full lengthwise juxtaposition of homologous chromosomes. Third, we reveal a prominent elongation of chromosome territories during meiotic prophase that initiates prior to homolog association and alignment. Mutant analysis indicates that chromosome movement mediated by association of chromosome pairing centers (PCs) with mobile patches of the nuclear envelope (NE)–spanning SUN-1/ZYG-12 protein complexes is not the primary driver of territory elongation. Moreover, we identify new roles for the X chromosome PC (X-PC) and X-PC binding protein HIM-8 in promoting elongation of X chromosome territories, separable from their role(s) in mediating local stabilization of pairing and association of X chromosomes with mobile SUN-1/ZYG-12 patches. Further, we present evidence that HIM-8 functions both at and outside of PCs to mediate chromosome territory elongation. These and other data support a model in which synapsis-independent elongation of chromosome territories, driven by PC binding proteins, enables lengthwise juxtaposition of chromosomes, thereby facilitating assessment of their suitability as potential pairing partners. PMID:21876678

Agrobacterium-mediated gene transfer in plants and biosafety considerations.

Science.gov (United States)

Mehrotra, Shweta; Goyal, Vinod

2012-12-01

Agrobacterium, the natures' genetic engineer, has been used as a vector to create transgenic plants. Agrobacterium-mediated gene transfer in plants is a highly efficient transformation process which is governed by various factors including genotype of the host plant, explant, vector, plasmid, bacterial strain, composition of culture medium, tissue damage, and temperature of co-cultivation. Agrobacterium has been successfully used to transform various economically and horticulturally important monocot and dicot species by standard tissue culture and in planta transformation techniques like floral or seedling infilteration, apical meristem transformation, and the pistil drip methods. Monocots have been comparatively difficult to transform by Agrobacterium. However, successful transformations have been reported in the last few years based on the adjustment of the parameters that govern the responses of monocots to Agrobacterium. A novel Agrobacterium transferred DNA-derived nanocomplex method has been developed which will be highly valuable for plant biology and biotechnology. Agrobacterium-mediated genetic transformation is known to be the preferred method of creating transgenic plants from a commercial and biosafety perspective. Agrobacterium-mediated gene transfer predominantly results in the integration of foreign genes at a single locus in the host plant, without associated vector backbone and is also known to produce marker free plants, which are the prerequisites for commercialization of transgenic crops. Research in Agrobacterium-mediated transformation can provide new and novel insights into the understanding of the regulatory process controlling molecular, cellular, biochemical, physiological, and developmental processes occurring during Agrobacterium-mediated transformation and also into a wide range of aspects on biological safety of transgenic crops to improve crop production to meet the demands of ever-growing world's population.

Chromosomal instability mediated by non-B DNA: cruciform conformation and not DNA sequence is responsible for recurrent translocation in humans.

Science.gov (United States)

Inagaki, Hidehito; Ohye, Tamae; Kogo, Hiroshi; Kato, Takema; Bolor, Hasbaira; Taniguchi, Mariko; Shaikh, Tamim H; Emanuel, Beverly S; Kurahashi, Hiroki

2009-02-01

Chromosomal aberrations have been thought to be random events. However, recent findings introduce a new paradigm in which certain DNA segments have the potential to adopt unusual conformations that lead to genomic instability and nonrandom chromosomal rearrangement. One of the best-studied examples is the palindromic AT-rich repeat (PATRR), which induces recurrent constitutional translocations in humans. Here, we established a plasmid-based model that promotes frequent intermolecular rearrangements between two PATRRs in HEK293 cells. In this model system, the proportion of PATRR plasmid that extrudes a cruciform structure correlates to the levels of rearrangement. Our data suggest that PATRR-mediated translocations are attributable to unusual DNA conformations that confer a common pathway for chromosomal rearrangements in humans.

Study of male–mediated gene flow across a hybrid zone in the common shrew (Sorex araneus using Y chromosome

Directory of Open Access Journals (Sweden)

Andrei V. Polyakov

2017-06-01

Full Text Available Despite many studies, the impact of chromosome rearrangements on gene flow between chromosome races of the common shrew (Sorex araneus Linnaeus, 1758 remains unclear. Interracial hybrids form meiotic chromosome complexes that are associated with reduced fertility. Nevertheless comprehensive investigations of autosomal and mitochondrial markers revealed weak or no barrier to gene flow between chromosomally divergent populations. In a narrow zone of contact between the Novosibirsk and Tomsk races hybrids are produced with extraordinarily complex configurations at meiosis I. Microsatellite markers have not revealed any barrier to gene flow, but the phenotypic differentiation between races is greater than may be expected if gene flow was unrestricted. To explore this contradiction we analyzed the distribution of the Y chromosome SNP markers within this hybrid zone. The Y chromosome variants in combination with race specific autosome complements allow backcrosses to be distinguished and their proportion among individuals within the hybrid zone to be evaluated. The balanced ratio of the Y variants observed among the pure race individuals as well as backcrosses reveals no male mediated barrier to gene flow. The impact of reproductive unfitness of backcrosses on gene flow is discussed as a possible mechanism of the preservation of race-specific morphology within the hybrid zone.

Environmental factors influencing gene transfer agent (GTA mediated transduction in the subtropical ocean.

Directory of Open Access Journals (Sweden)

Lauren D McDaniel

Full Text Available Microbial genomic sequence analyses have indicated widespread horizontal gene transfer (HGT. However, an adequate mechanism accounting for the ubiquity of HGT has been lacking. Recently, high frequencies of interspecific gene transfer have been documented, catalyzed by Gene Transfer Agents (GTAs of marine α-Proteobacteria. It has been proposed that the presence of bacterial genes in highly purified viral metagenomes may be due to GTAs. However, factors influencing GTA-mediated gene transfer in the environment have not yet been determined. Several genomically sequenced strains containing complete GTA sequences similar to Rhodobacter capsulatus (RcGTA, type strain were screened to ascertain if they produced putative GTAs, and at what abundance. Five of nine marine strains screened to date spontaneously produced virus-like particles (VLP's in stationary phase. Three of these strains have demonstrated gene transfer activity, two of which were documented by this lab. These two strains Roseovarius nubinhibens ISM and Nitratireductor 44B9s, were utilized to produce GTAs designated RnGTA and NrGTA and gene transfer activity was verified in culture. Cell-free preparations of purified RnGTA and NrGTA particles from marked donor strains were incubated with natural microbial assemblages to determine the level of GTA-mediated gene transfer. In conjunction, several ambient environmental parameters were measured including lysogeny indicated by prophage induction. GTA production in culture systems indicated that approximately half of the strains produced GTA-like particles and maximal GTA counts ranged from 10-30% of host abundance. Modeling of GTA-mediated gene transfer frequencies in natural samples, along with other measured environmental variables, indicated a strong relationship between GTA mediated gene transfer and the combined factors of salinity, multiplicity of infection (MOI and ambient bacterial abundance. These results indicate that GTA-mediated

Neocentromeres Provide Chromosome Segregation Accuracy and Centromere Clustering to Multiple Loci along a Candida albicans Chromosome.

Directory of Open Access Journals (Sweden)

Laura S Burrack

2016-09-01

Full Text Available Assembly of kinetochore complexes, involving greater than one hundred proteins, is essential for chromosome segregation and genome stability. Neocentromeres, or new centromeres, occur when kinetochores assemble de novo, at DNA loci not previously associated with kinetochore proteins, and they restore chromosome segregation to chromosomes lacking a functional centromere. Neocentromeres have been observed in a number of diseases and may play an evolutionary role in adaptation or speciation. However, the consequences of neocentromere formation on chromosome missegregation rates, gene expression, and three-dimensional (3D nuclear structure are not well understood. Here, we used Candida albicans, an organism with small, epigenetically-inherited centromeres, as a model system to study the functions of twenty different neocentromere loci along a single chromosome, chromosome 5. Comparison of neocentromere properties relative to native centromere functions revealed that all twenty neocentromeres mediated chromosome segregation, albeit to different degrees. Some neocentromeres also caused reduced levels of transcription from genes found within the neocentromere region. Furthermore, like native centromeres, neocentromeres clustered in 3D with active/functional centromeres, indicating that formation of a new centromere mediates the reorganization of 3D nuclear architecture. This demonstrates that centromere clustering depends on epigenetically defined function and not on the primary DNA sequence, and that neocentromere function is independent of its distance from the native centromere position. Together, the results show that a neocentromere can form at many loci along a chromosome and can support the assembly of a functional kinetochore that exhibits native centromere functions including chromosome segregation accuracy and centromere clustering within the nucleus.

Dysregulation of gene expression in the artificial human trisomy cells of chromosome 8 associated with transformed cell phenotypes.

Directory of Open Access Journals (Sweden)

Hisakatsu Nawata

Full Text Available A change in chromosome number, known as aneuploidy, is a common characteristic of cancer. Aneuploidy disrupts gene expression in human cancer cells and immortalized human epithelial cells, but not in normal human cells. However, the relationship between aneuploidy and cancer remains unclear. To study the effects of aneuploidy in normal human cells, we generated artificial cells of human primary fibroblast having three chromosome 8 (trisomy 8 cells by using microcell-mediated chromosome transfer technique. In addition to decreased proliferation, the trisomy 8 cells lost contact inhibition and reproliferated after exhibiting senescence-like characteristics that are typical of transformed cells. Furthermore, the trisomy 8 cells exhibited chromosome instability, and the overall gene expression profile based on microarray analyses was significantly different from that of diploid human primary fibroblasts. Our data suggest that aneuploidy, even a single chromosome gain, can be introduced into normal human cells and causes, in some cases, a partial cancer phenotype due to a disruption in overall gene expression.

BRCA1-mediated repression of select X chromosome genes

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Ropers H Hilger

2004-09-01

Full Text Available Abstract Recently BRCA1 has been implicated in the regulation of gene expression from the X chromosome. In this study the influence of BRCA1 on expression of X chromosome genes was investigated. Complementary DNA microarrays were used to compare the expression levels of X chromosome genes in 18 BRCA1-associated ovarian cancers to those of the 13 "BRCA1-like" and 14 "BRCA2-like" sporadic tumors (as defined by previously reported expression profiling. Significance was determined using parametric statistics with P

Sustainability of University Technology Transfer: Mediating Effect of Inventor’s Technology Service

Directory of Open Access Journals (Sweden)

Fang Li

2018-06-01

Full Text Available Based on the perspective of knowledge transfer and the technology acceptance model (TAM, this paper constructs a university technology transfer sustainable development model that considers the inventor’s technology service from the perspective of the long-term cooperation of enterprise, and analyzes the mediating effect of the inventor’s technology service on university technology transfer sustainability. By using 270 questionnaires as survey data, it is found that the availability of an inventor’s technology service has a significant positive impact on the attitude tendency and practice tendency of enterprise long-term technological cooperation; enterprise technology absorption capacity and trust between a university and an enterprise also have significant influence on an inventor’s technical service availability. Therefore, the inventor’s technology service acts as a mediator in the relationship between university technology transfer sustainability and influence factors. Universities ought to establish the technology transfer model, which focuses on the inventor’s tacit knowledge transfer service, and promotes the sustainable development of the university.

Genetic variability available through cell fusion

Energy Technology Data Exchange (ETDEWEB)

Smith, H.H.; Mastrangelo-Hough, I.A.

1977-01-01

Results are reported for the following studies: plant hybridization through protoplast fusion using species of Nicotiana and Petunia; chromosome instability studies on culture-induced chromosome changes and chromosome elimination; chloroplast distribution in parasexual hybrids; chromosomal introgression following fusion; plant-animal fusion; and microcell-mediated chromosome transfer and chromosome-mediated gene transfer. (HLW)

Culture, characteristics and chromosome complement of Siberian tiger fibroblasts for nuclear transfer.

Science.gov (United States)

Song, Jimei; Hua, Song; Song, Kai; Zhang, Yong

2007-01-01

Tiger (Panthera tigris Linnaeus, 1758) is a characteristic species of Asia, which is in severe danger. Siberian tiger (Panthera tigris altaica) is the largest one of the five existent tiger subspecies. It is extremely endangered. One new way for tiger protection and rescue is to study interspecies cloning. But there is few research data about Siberian tiger. In this study, we cultured Siberian tiger fibroblasts in vitro, analyzed their biological characteristics, chromosomes, and cell cycles, to provide not only nuclear donors with good morphology, normal biological characteristics, and chromosome quantity for tiger interspecies cloning, but also reliable data for further studying Siberian tiger. The results indicated that Siberian tiger ear fibroblasts can be successfully obtained by tissue culture either with or without overnight cold digestion, the cultured cells were typical fibroblasts with normal morphology, growth curve, and chromosome quantity; G0/G1 percentage increased and S percentage decreased with the confluence of cells. G0/G1 and S stage rate was significantly different between 40-50% and 80-90%, 95-100% confluence; there is no distinct difference between 80-90% and 95-100% confluence. The cells at the same density (80-90% confluence) were treated with or without 0.5% serum starving, GO/G1 rate of the former was higher than the latter, but the difference was not significant. GO/G1 proportion of 95-100% confluence was slightly higher than serum starving (80-90% confluence), but no significant difference. Therefore, the Siberian tiger fibroblasts we cultured in vitro can be used as donor cells, and the donor cells do not need to be treated with normal serum starvation during nuclear transfer; if we will just consider the rate of the G0/G1 stage cells, serum starvation can be replaced by confluence inhibition when cultured cells were more than 80-90% confluence.

Numerical chromosome errors in day 7 somatic nuclear blastocysts

DEFF Research Database (Denmark)

Booth, Paul J; Viuff, Dorthe; Tan, Shijian J

2003-01-01

Day 7 bovine somatic nuclear transfer (NT) embryos reconstructed from granulosa cells were examined for numerical chromosome aberrations as a potential cause of the high embryonic and fetal loss observed in such embryos after transfer. The NT embryos were reconstructed using a zona-free manipulat......Day 7 bovine somatic nuclear transfer (NT) embryos reconstructed from granulosa cells were examined for numerical chromosome aberrations as a potential cause of the high embryonic and fetal loss observed in such embryos after transfer. The NT embryos were reconstructed using a zona...... families, consisting of 112 blastocysts reconstructed from five different primary granulosa cell cultures, were examined. Overall, the mean chromosome complement within embryos was 86.9 +/- 3.7% (mean +/- SEM) diploid, 2.6 +/- 0.5% triploid, 10.0 +/- 3.1% tetraploid, and 0.5 +/- 0.2% pentaploid or greater......; the vast majority (>75%) of the abnormal nuclei were tetraploid. Completely diploid and mixoploid embryos represented 22.1 +/- 4.5% and 73.7 +/- 5.5%, respectively, of all clones. Six totally polyploid blastocysts, containing or=5N chromosome complements, respectively) between two clone families were...

Transfer of Hessian fly resistance from rye to wheat via radiation-induced terminal and intercalary chromosomal translocations

International Nuclear Information System (INIS)

Friebe, B.; Hatchett, J.H.; Gill, B.S.; Mukai, Y.; Sebesta, E.E.

1991-01-01

A new Hessian fly (Mayetiola destructor) resistance gene derived from 'Balbo' rye and its transfer to hexaploid wheat via radiation-induced terminal and intercalary chromosomal translocations are described. Crosses between resistant 'Balbo' rye and susceptible 'Suwon 92' wheat and between the F1 amphidiploids and susceptible 'TAM 106' and 'Amigo' wheats produced resistant BC2F3 lines that were identified by C-banding analysis as being 6RL telocentric addition lines. Comparative chromosomal analyses and resistance tests revealed that the resistance gene is located on the 6RL telocentric chromosome. X-irradiated pollen of 6RL addition plants was used to fertilize plants of susceptible wheats 'TAM 106,' 'TAM 101,' and 'Vona.' After several generations of selection for resistance, new sublines were obtained that were homogeneous for resistance. Thirteen of these lines were analyzed by C-banding, and three different wheat-6RL chromosomal translocations (T) were identified. Wheat chromosomes involved in the translocations were 6B, 4B, and 4A. Almost the complete 6RL arm is present in T6BS · 6BL-6RL. Only the distal half of 6RL is present in T4BS · 4BL-6RL, which locates the resistance gene in the distal half of 6RL. Only a very small segment (ca 1.0 μm) of the distal region of 6RL is present in an intercalary translocation (Ti) Ti4AS · 4AL-6RL-4AL. The 6RL segment is inserted in the intercalary region between the centromere of chromosome 4A and the large proximal C-band of 4AL. The break-points of the translocations are outside the region of the centromere, indicating that they were induced by the X-ray treatment. All three translocations are cytologically stable and can be used directly in wheat breeding programs

Telomerase-mediated life-span extension of human primary fibroblasts by human artificial chromosome (HAC) vector

International Nuclear Information System (INIS)

Shitara, Shingo; Kakeda, Minoru; Nagata, Keiko; Hiratsuka, Masaharu; Sano, Akiko; Osawa, Kanako; Okazaki, Akiyo; Katoh, Motonobu; Kazuki, Yasuhiro; Oshimura, Mitsuo; Tomizuka, Kazuma

2008-01-01

Telomerase-mediated life-span extension enables the expansion of normal cells without malignant transformation, and thus has been thought to be useful in cell therapies. Currently, integrating vectors including the retrovirus are used for human telomerase reverse transcriptase (hTERT)-mediated expansion of normal cells; however, the use of these vectors potentially causes unexpected insertional mutagenesis and/or activation of oncogenes. Here, we established normal human fibroblast (hPF) clones retaining non-integrating human artificial chromosome (HAC) vectors harboring the hTERT expression cassette. In hTERT-HAC/hPF clones, we observed the telomerase activity and the suppression of senescent-associated SA-β-galactosidase activity. Furthermore, the hTERT-HAC/hPF clones continued growing beyond 120 days after cloning, whereas the hPF clones retaining the silent hTERT-HAC senesced within 70 days. Thus, hTERT-HAC-mediated episomal expression of hTERT allows the extension of the life-span of human primary cells, implying that gene delivery by non-integrating HAC vectors can be used to control cellular proliferative capacity of primary cultured cells

SAP-like domain in nucleolar spindle associated protein mediates mitotic chromosome loading as well as interphase chromatin interaction

Energy Technology Data Exchange (ETDEWEB)

Verbakel, Werner, E-mail: werner.verbakel@chem.kuleuven.be [Laboratory of Biomolecular Dynamics, Katholieke Universiteit Leuven, Celestijnenlaan 200G, Bus 2403, 3001 Heverlee (Belgium); Carmeliet, Geert, E-mail: geert.carmeliet@med.kuleuven.be [Laboratory of Experimental Medicine and Endocrinology, Katholieke Universiteit Leuven, Herestraat 49, Bus 902, 3000 Leuven (Belgium); Engelborghs, Yves, E-mail: yves.engelborghs@fys.kuleuven.be [Laboratory of Biomolecular Dynamics, Katholieke Universiteit Leuven, Celestijnenlaan 200G, Bus 2403, 3001 Heverlee (Belgium)

2011-08-12

Highlights: {yields} The SAP-like domain in NuSAP is a functional DNA-binding domain with preference for dsDNA. {yields} This SAP-like domain is essential for chromosome loading during early mitosis. {yields} NuSAP is highly dynamic on mitotic chromatin, as evident from photobleaching experiments. {yields} The SAP-like domain also mediates NuSAP-chromatin interaction in interphase nucleoplasm. -- Abstract: Nucleolar spindle associated protein (NuSAP) is a microtubule-stabilizing protein that localizes to chromosome arms and chromosome-proximal microtubules during mitosis and to the nucleus, with enrichment in the nucleoli, during interphase. The critical function of NuSAP is underscored by the finding that its depletion in HeLa cells results in various mitotic defects. Moreover, NuSAP is found overexpressed in multiple cancers and its expression levels often correlate with the aggressiveness of cancer. Due to its localization on chromosome arms and combination of microtubule-stabilizing and DNA-binding properties, NuSAP takes a special place within the extensive group of spindle assembly factors. In this study, we identify a SAP-like domain that shows DNA binding in vitro with a preference for dsDNA. Deletion of the SAP-like domain abolishes chromosome arm binding of NuSAP during mitosis, but is not sufficient to abrogate its chromosome-proximal localization after anaphase onset. Fluorescence recovery after photobleaching experiments revealed the highly dynamic nature of this NuSAP-chromatin interaction during mitosis. In interphase cells, NuSAP also interacts with chromatin through its SAP-like domain, as evident from its enrichment on dense chromatin regions and intranuclear mobility, measured by fluorescence correlation spectroscopy. The obtained results are in agreement with a model where NuSAP dynamically stabilizes newly formed microtubules on mitotic chromosomes to enhance chromosome positioning without immobilizing these microtubules. Interphase Nu

SAP-like domain in nucleolar spindle associated protein mediates mitotic chromosome loading as well as interphase chromatin interaction

International Nuclear Information System (INIS)

Verbakel, Werner; Carmeliet, Geert; Engelborghs, Yves

2011-01-01

Highlights: → The SAP-like domain in NuSAP is a functional DNA-binding domain with preference for dsDNA. → This SAP-like domain is essential for chromosome loading during early mitosis. → NuSAP is highly dynamic on mitotic chromatin, as evident from photobleaching experiments. → The SAP-like domain also mediates NuSAP-chromatin interaction in interphase nucleoplasm. -- Abstract: Nucleolar spindle associated protein (NuSAP) is a microtubule-stabilizing protein that localizes to chromosome arms and chromosome-proximal microtubules during mitosis and to the nucleus, with enrichment in the nucleoli, during interphase. The critical function of NuSAP is underscored by the finding that its depletion in HeLa cells results in various mitotic defects. Moreover, NuSAP is found overexpressed in multiple cancers and its expression levels often correlate with the aggressiveness of cancer. Due to its localization on chromosome arms and combination of microtubule-stabilizing and DNA-binding properties, NuSAP takes a special place within the extensive group of spindle assembly factors. In this study, we identify a SAP-like domain that shows DNA binding in vitro with a preference for dsDNA. Deletion of the SAP-like domain abolishes chromosome arm binding of NuSAP during mitosis, but is not sufficient to abrogate its chromosome-proximal localization after anaphase onset. Fluorescence recovery after photobleaching experiments revealed the highly dynamic nature of this NuSAP-chromatin interaction during mitosis. In interphase cells, NuSAP also interacts with chromatin through its SAP-like domain, as evident from its enrichment on dense chromatin regions and intranuclear mobility, measured by fluorescence correlation spectroscopy. The obtained results are in agreement with a model where NuSAP dynamically stabilizes newly formed microtubules on mitotic chromosomes to enhance chromosome positioning without immobilizing these microtubules. Interphase NuSAP-chromatin interaction

The mediating role of absorptive capacity in knowledge transfer

DEFF Research Database (Denmark)

Adisa, Femi; Rose, Jeremy

2013-01-01

. These problems become acute in implementations in small and medium sized enterprises (SMEs). SMEs often operate with non-standard business processes, making an effective interchange of process knowledge between consultants and end-users crucial. Using a multiple case study method and content analysis......, the authors investigate the mediating role of absorptive capacity in knowledge transfer in SMEs ERP implementations. They present exploratory case studies from 3 Nigerian companies with varying outcomes, and hypothesize that knowledge transfer is complicated by acute information asymmetry, absence of pre...

Electron-transfer mediator for a NAD-glucose dehydrogenase-based glucose sensor.

Science.gov (United States)

Kim, Dong-Min; Kim, Min-yeong; Reddy, Sanapalli S; Cho, Jaegeol; Cho, Chul-ho; Jung, Suntae; Shim, Yoon-Bo

2013-12-03

A new electron-transfer mediator, 5-[2,5-di (thiophen-2-yl)-1H-pyrrol-1-yl]-1,10-phenanthroline iron(III) chloride (FePhenTPy) oriented to the nicotinamide adenine dinucleotide-dependent-glucose dehydrogenase (NAD-GDH) system was synthesized through a Paal-Knorr condensation reaction. The structure of the mediator was confirmed by Fourier-transform infrared spectroscopy, proton and carbon nucler magnetic resonance spectroscopy, and mass spectroscopy, and its electron-transfer characteristic for a glucose sensor was investigated using voltammetry and impedance spectroscopy. A disposable amperometric glucose sensor with NAD-GDH was constructed with FePhenTPy as an electron-transfer mediator on a screen printed carbon electrode (SPCE) and its performance was evaluated, where the addition of reduces graphene oxide (RGO) to the mediator showed the enhanced sensor performance. The experimental parameters to affect the analytical performance and the stability of the proposed glucose sensor were optimized, and the sensor exhibited a dynamic range between 30 mg/dL and 600 mg/dL with the detection limit of 12.02 ± 0.6 mg/dL. In the real sample experiments, the interference effects by acetaminophen, ascorbic acid, dopamine, uric acid, caffeine, and other monosaccharides (fructose, lactose, mannose, and xylose) were completely avoided through coating the sensor surface with the Nafion film containing lead(IV) acetate. The reliability of proposed glucose sensor was evaluated by the determination of glucose in artificial blood and human whole blood samples.

Chromosomal organization of adrenergic receptor genes

International Nuclear Information System (INIS)

Yang-Feng, T.L.; Xue, Feiyu; Zhong, Wuwei; Cotecchia, S.; Frielle, T.; Caron, M.G.; Lefkowitz, R.J.; Francke, U.

1990-01-01

The adrenergic receptors (ARs) (subtypes α 1 , α 2 , β 1 , and β 2 ) are a prototypic family of guanine nucleotide binding regulatory protein-coupled receptors that mediate the physiological effects of the hormone epinephrine and the neurotransmitter norepinephrine. The authors have previously assigned the genes for β 2 -and α 2 -AR to human chromosomes 5 and 10, respectively. By Southern analysis of somatic cell hybrids and in situ chromosomal hybridization, they have now mapped the α 1 -AR gene to chromosome 5q32→q34, the same position as β 2 -AR, and the β 1 -AR gene to chromosome 10q24→q26, the region where α 2 -AR, is located. In mouse, both α 2 -and β 1 -AR genes were assigned to chromosome 19, and the α 1 -AR locus was localized to chromosome 11. Pulsed field gel electrophoresis has shown that the α 1 -and β 2 -AR genes in humans are within 300 kilobases (kb) and the distance between the α 2 - and β 1 -AR genes is <225 kb. The proximity of these two pairs of AR genes and the sequence similarity that exists among all the ARs strongly suggest that they are evolutionarily related. Moreover, they likely arose from a common ancestral receptor gene and subsequently diverged through gene duplication and chromosomal duplication to perform their distinctive roles in mediation the physiological effects of catecholamines. The AR genes thus provide a paradigm for understanding the evolution of such structurally conserved yet functionally divergent families off receptor molecules

ADA1 and NET1 Genes of Yeast Mediate Both Chromosome Maintenance and Mitochondrial $\\rho^{-}$ Mutagenesis

CERN Document Server

Koltovaya, N A; Tchekhouta, I A; Devin, A B

2002-01-01

An increase in the mitochondrial (mt) rho^- mutagenesis is a well-known respose of yeast cells to mutations in the numerous nuclear genes as well as to various kinds of stress. Notwithstanding the extensive studies during several decades the biological significance of this response is not yet fully understood. The genetic approach to solution of this subject includes the study of genes that are required for the high incidence of spontaneous rho^- mutants. Previously we found that mutations in certain nuclear genes including CDC28, the central cell-cycle regulation gene, may decrease the spontaneous rho^- mutability and simultaneously affect maintenance of the yeast chromosomes and plasmids. The present work provides data on identification of two more genes, resembling CDC28 in this respect. These genes NET1 and ADA1 mediate important regulatory protein-protein interactions in the yeast cell. The effects of net1 and ada1 mutations on the maintenance of yeast mt genome, chromosomes and plasmids as well as on ce...

Proton Transfer in Nucleobases is Mediated by Water

Energy Technology Data Exchange (ETDEWEB)

Khistyaev, Kirill; Golan, Amir; Bravaya, Ksenia B.; Orms, Natalie; Krylov, Anna I.; Ahmed, Musahid

2013-08-08

Water plays a central role in chemistry and biology by mediating the interactions between molecules, altering energy levels of solvated species, modifying potential energy proles along reaction coordinates, and facilitating ecient proton transport through ion channels and interfaces. This study investigates proton transfer in a model system comprising dry and microhydrated clusters of nucleobases. With mass spectrometry and tunable vacuum ultraviolet synchrotron radiation, we show that water shuts down ionization-induced proton transfer between nucleobases, which is very ecient in dry clusters. Instead, a new pathway opens up in which protonated nucleo bases are generated by proton transfer from the ionized water molecule and elimination of a hydroxyl radical. Electronic structure calculations reveal that the shape of the potential energy prole along the proton transfer coordinate depends strongly on the character of the molecular orbital from which the electron is removed, i.e., the proton transfer from water to nucleobases is barrierless when an ionized state localized on water is accessed. The computed energetics of proton transfer is in excellent agreement with the experimental appearance energies. Possible adiabatic passage on the ground electronic state of the ionized system, while energetically accessible at lower energies, is not ecient. Thus, proton transfer is controlled electronically, by the character of the ionized state, rather than statistically, by simple energy considerations.

Social Support and Motivation to Transfer as Predictors of Training Transfer: Testing Full and Partial Mediation Using Meta-Analytic Structural Equation Modelling

Science.gov (United States)

Reinhold, Sarah; Gegenfurtner, Andreas; Lewalter, Doris

2018-01-01

Social support and motivation to transfer are important components in conceptual models on transfer of training. Previous research indicates that both support and motivation influence transfer. To date, however, it is not yet clear if social support influences transfer of training directly, or if this influence is mediated by motivation to…

Reversible assembly of protein-DNA nanostructures triggered by mediated electron transfer

International Nuclear Information System (INIS)

Vogt, Stephan; Wenderhold-Reeb, Sabine; Nöll, Gilbert

2017-01-01

Stable protein-DNA nanostructures have been assembled by reconstitution of the multi-ligand binding flavoprotein dodecin on top of flavin-terminated dsDNA monolayers on gold electrodes. These structures could be disassembled by electrochemical flavin reduction via mediated electron transfer. For this purpose a negative potential was applied at the Au working electrode in the presence of the redox mediator bis-(ammoniumethyl)-4,4′-bipyridinium tetrabromide. The stepwise formation of the flavin-terminated dsDNA monolayers as well as the binding and electrochemically triggered release of apododecin were monitored by surface plasmon resonance (SPR) and quartz crystal microbalance (QCM) measurements. The assembly and disassembly of the protein-DNA nanostructures were fully reversible processes, which could be carried out multiple times at the same flavin-dsDNA modified surface. When a negative potential was applied in the absence of a redox mediator apododecin could not be released, i.e. direct electron transfer was not possible. As alternative redox mediators also methylene blue and phenosafranine were studied, but in the presence of these molecules apododecin was released without applying a potential, probably because the tricyclic aromatic compounds are able to replace the flavins at the binding sites.

Follistatin allows efficient retroviral-mediated gene transfer into rat liver

International Nuclear Information System (INIS)

Borgnon, Josephine; Djamouri, Fatima; Lorand, Isabelle; Rico, Virginie Di; Loux, Nathalie; Pages, Jean-Christophe; Franco, Dominique; Capron, Frederique; Weber, Anne

2005-01-01

Retroviral vectors are widely used tools for gene therapy. However, in vivo gene transfer is only effective in dividing cells, which, in liver, requires a regenerative stimulus. Follistatin is effective in promoting liver regeneration after 90% and 70% hepatectomy in rats. We studied its efficacy on liver regeneration and retroviral-mediated gene delivery in 50% hepatectomized rats. When human recombinant follistatin was infused into the portal vein immediately after 50% hepatectomy, hepatocyte proliferation was significantly higher than in control 50% hepatectomized rats. A single injection of virus particles administered 23 h after follistatin infusion resulted in more than 20% gene transduction efficiency in hepatocytes compared to 3% in control rats. It is concluded that a single injection of follistatin induces onset of proliferation in 50% hepatectomized rats and allows efficient retroviral-mediated gene transfer to the liver

Electron Transfer and Solvent-Mediated Electronic Localization in Molecular Photocatalysis

DEFF Research Database (Denmark)

Dohn, Asmus Ougaard; Kjær, Kasper Skov; Harlang, Tobias B.

2016-01-01

This work provides a detailed mechanism for electron transfer in a heterodinuclear complex designed as a model system in which to study homogeneous molecular photocatalysis. With efficient Born–Oppenheimer molecular dynamics simulations, we show how intermediate, charge-separated states can mediate...

Human chromosome 9 can complement UV sensitivity of xeroderma pigmentosum group A cells

International Nuclear Information System (INIS)

Ishizaki, Kanji; Sasaki, Masao S.; Ikenaga, Mituo; Nakamura, Yusuke

1990-01-01

A single human chromosome derived from normal human fibroblasts and tagged with the G418 resistance gene was transferred into SV40-transformed xeroderma pigmentosum group A (XP-A) cells via microcell fusion. When chromosome 1 or 12 was transferred, UV sensitivity of microcell hybrid cells was not changed. By contrast, after transferring chromosome 9,7 of 11 reipient clones were as UV-resistant as normal human cells. Four other clones were still as UV-sensitive as the parental XP-A cells. Southern hybridization analysis using a polymorphic probe, pEKZ19.3, which is homologous to a sequence of the D9S17 locus on chromosome 9, has confirmed that at least a part of normal human chromosome 9 was transferred into the recipient clones. However, amounts iof UV-induced unscheduled DNA synthesis in the UV-resistant clones were only one-third of those in normal human cells. These results indicate that a gene on chromosome 9 can confer complementation of high UV sensitivity of XP-A cells although it is still possible that 2 or more genes might be involved in the defective-repair phenotypes of XP-A. (author). 20 refs.; 3 figs.; 1 tab

Tyrosine kinase chromosomal translocations mediate distinct and overlapping gene regulation events

International Nuclear Information System (INIS)

Kim, Hani; Gillis, Lisa C; Jarvis, Jordan D; Yang, Stuart; Huang, Kai; Der, Sandy; Barber, Dwayne L

2011-01-01

Leukemia is a heterogeneous disease commonly associated with recurrent chromosomal translocations that involve tyrosine kinases including BCR-ABL, TEL-PDGFRB and TEL-JAK2. Most studies on the activated tyrosine kinases have focused on proximal signaling events, but little is known about gene transcription regulated by these fusions. Oligonucleotide microarray was performed to compare mRNA changes attributable to BCR-ABL, TEL-PDGFRB and TEL-JAK2 after 1 week of activation of each fusion in Ba/F3 cell lines. Imatinib was used to control the activation of BCR-ABL and TEL-PDGFRB, and TEL-JAK2-mediated gene expression was examined 1 week after Ba/F3-TEL-JAK2 cells were switched to factor-independent conditions. Microarray analysis revealed between 800 to 2000 genes induced or suppressed by two-fold or greater by each tyrosine kinase, with a subset of these genes commonly induced or suppressed among the three fusions. Validation by Quantitative PCR confirmed that eight genes (Dok2, Mrvi1, Isg20, Id1, gp49b, Cxcl10, Scinderin, and collagen Vα1(Col5a1)) displayed an overlapping regulation among the three tested fusion proteins. Stat1 and Gbp1 were induced uniquely by TEL-PDGFRB. Our results suggest that BCR-ABL, TEL-PDGFRB and TEL-JAK2 regulate distinct and overlapping gene transcription profiles. Many of the genes identified are known to be involved in processes associated with leukemogenesis, including cell migration, proliferation and differentiation. This study offers the basis for further work that could lead to an understanding of the specificity of diseases caused by these three chromosomal translocations

Extra-chromosomal DNA maintenance in Bacillus subtilis, dependence on flagellation factor FliF and moonlighting mediator EdmS.

Science.gov (United States)

Hakumai, Yuichi; Shimomoto, Kouko; Ashiuchi, Makoto

2015-05-15

Extra-chromosomal DNA maintenance (EDM) as an important process in the propagation and genetic engineering of microbes. Bacillus subtilis EdmS (formerly PgsE), a protein comprising 55 amino acids, is a mediator of the EDM process. In this study, the effect of mutation of global regulators on B. subtilis EDM was examined. Mutation of the swrA gene abolished EdmS-mediated EDM. It is known that swrA predominantly regulates expression of the fla/che operon in B. subtilis. We therefore performed EDM analysis using fla/che-deletion mutants and identified an EDM-mediated EDM cooperator in the flgB-fliL region. Further genetic investigation identified the flagellation factor FliF is a crucial EDM cooperator. To our knowledge, this is the first observation of the moonlighting function of FliF in DNA maintenance. Copyright © 2015 Elsevier Inc. All rights reserved.

High rate of translocation-based gene birth on the Drosophila Y chromosome.

Science.gov (United States)

Tobler, Ray; Nolte, Viola; Schlötterer, Christian

2017-10-31

The Y chromosome is a unique genetic environment defined by a lack of recombination and male-limited inheritance. The Drosophila Y chromosome has been gradually acquiring genes from the rest of the genome, with only seven Y-linked genes being gained over the past 63 million years (0.12 gene gains per million years). Using a next-generation sequencing (NGS)-powered genomic scan, we show that gene transfers to the Y chromosome are much more common than previously suspected: at least 25 have arisen across three Drosophila species over the past 5.4 million years (1.67 per million years for each lineage). The gene transfer rate is significantly lower in Drosophila melanogaster than in the Drosophila simulans clade, primarily due to Y-linked retrotranspositions being significantly more common in the latter. Despite all Y-linked gene transfers being evolutionarily recent (Drosophila Y chromosome to be more dynamic than previously appreciated. Our analytical method provides a powerful means to identify Y-linked gene transfers and will help illuminate the evolutionary dynamics of the Y chromosome in Drosophila and other species. Copyright © 2017 the Author(s). Published by PNAS.

Conceptualizing knowledge transfer between expatriates and host country nationals: The mediating effect of social capital

Directory of Open Access Journals (Sweden)

Maimunah Ismail

2015-12-01

Full Text Available This paper aims to propose a conceptual model of knowledge transfer by relating two specific personal factors of expatriate and host country national (HCN dyads as antecedents of knowledge transfer, and mediated by social capital factors. An intensive literature review method was employed to identify and analyse relevant literatures. The paper used a dyadic bi-directional approach in theorizing knowledge transfer by integrating the social capital theory, and the anxiety and uncertainty management theory. The paper considers two personal factors (cultural intelligence and knowledge-seeking behaviour and two social capital variables (trust and shared vision as mediators of knowledge transfer. Upon model validation, the paper could offer practical interventions for human resource practitioners and managers to assist multinational corporations towards managing knowledge transfer involving expatriates and HCNs.

ADA1 and NET1 genes of yeast mediate both chromosome maintenance and mitochondrial rho- mutagenesis

International Nuclear Information System (INIS)

Koltovaya, N.A.; Gerasimova, A.S.; Chekhuta, I.A.; Devin, A.B.

2002-01-01

An increase in the mitochondrial (mt) rho - mutagenesis is a well-known response of yeast cells to mutations in the numerous nuclear genes as well as to various kinds of stress. Notwithstanding the extensive studies during several decades the biological significance of this response is not yet fully understood. The genetic approach to solution of this subject includes the study of genes that are required for the high incidence of spontaneous rho - mutants. Previously we found that mutations in certain nuclear genes including CDC28, the central cell-cycle regulation gene, may decrease the spontaneous rho - mutability and simultaneously affect maintenance of the yeast chromosomes and plasmids. The present work provides data on identification of two more genes, resembling CDC28 in this respect. These genes NET1 and ADA1 mediate important regulatory protein-protein interactions in the yeast cell. The effects of net1 and ada1 mutations on the maintenance of yeast mt genome, chromosomes and plasmids as well on cell sensitivity to ionizing radiation are also described. (author)

DNA repair genes RAD52 and SRS2, a cell wall synthesis regulator gene SMI1, and the membrane sterol synthesis scaffold gene ERG28 are important in efficient Agrobacterium-mediated yeast transformation with chromosomal T-DNA.

Science.gov (United States)

Ohmine, Yuta; Satoh, Yukari; Kiyokawa, Kazuya; Yamamoto, Shinji; Moriguchi, Kazuki; Suzuki, Katsunori

2016-04-02

Plant pathogenic Agrobacterium strains can transfer T-DNA regions of their Ti plasmids to a broad range of eukaryotic hosts, including fungi, in vitro. In the recent decade, the yeast Saccharomyces cerevisiae is used as a model host to reveal important host proteins for the Agrobacterium-mediated transformation (AMT). Further investigation is required to understand the fundamental mechanism of AMT, including interaction at the cell surface, to expand the host range, and to develop new tools. In this study, we screened a yeast mutant library for low AMT mutant strains by advantage of a chromosome type T-DNA, which transfer is efficient and independent on integration into host chromosome. By the mutant screening, we identified four mutant strains (srs2Δ, rad52Δ, smi1Δ and erg28Δ), which showed considerably low AMT efficiency. Structural analysis of T-DNA product replicons in AMT colonies of mutants lacking each of the two DNA repair genes, SRS2 and RAD52, suggested that the genes act soon after T-DNA entry for modification of the chromosomal T-DNA to stably maintain them as linear replicons and to circularize certain T-DNA simultaneously. The cell wall synthesis regulator SMI1 might have a role in the cell surface interaction between the donor and recipient cells, but the smi1Δ mutant exhibited pleiotropic effect, i.e. low effector protein transport as well as low AMT for the chromosomal T-DNA, but relatively high AMT for integrative T-DNAs. The ergosterol synthesis regulator/enzyme-scaffold gene ERG28 probably contributes by sensing a congested environment, because growth of erg28Δ strain was unaffected by the presence of donor bacterial cells, while the growth of the wild-type and other mutant yeast strains was suppressed by their presence. RAD52 and the DNA helicase/anti-recombinase gene SRS2 are necessary to form and maintain artificial chromosomes through the AMT of chromosomal T-DNA. A sterol synthesis scaffold gene ERG28 is important in the high

Drug-induced premature chromosome condensation (PCC) protocols: cytogenetic approaches in mitotic chromosome and interphase chromatin.

Science.gov (United States)

Gotoh, Eisuke

2015-01-01

Chromosome analysis is a fundamental technique which is used in wide areas of cytogenetic study including karyotyping species, hereditary diseases diagnosis, or chromosome biology study. Chromosomes are usually prepared from mitotic cells arrested by colcemid block protocol. However, obtaining mitotic chromosomes is often hampered under several circumstances. As a result, cytogenetic analysis will be sometimes difficult or even impossible in such cases. Premature chromosome condensation (PCC) (see Note 1) is an alternative method that has proved to be a unique and useful way in chromosome analysis. Former, PCC has been achieved following cell fusion method (cell-fusion PCC) mediated either by fusogenic viruses (e.g., Sendai virus) or cell fusion chemicals (e.g., polyethylene glycol), but the cell fusion PCC has several drawbacks. The novel drug-induced PCC using protein phosphatase inhibitors was introduced about 20 years ago. This method is much simpler and easier even than the conventional mitotic chromosome preparation protocol use with colcemid block and furthermore obtained PCC index (equivalent to mitotic index for metaphase chromosome) is usually much higher than colcemid block method. Moreover, this method allows the interphase chromatin to be condensed to visualize like mitotic chromosomes. Therefore drug-induced PCC has opened the way for chromosome analysis not only in metaphase chromosomes but also in interphase chromatin. The drug-induced PCC has thus proven the usefulness in cytogenetics and other cell biology fields. For this second edition version, updated modifications/changes are supplemented in Subheadings 2, 3, and 4, and a new section describing the application of PCC in chromosome science fields is added with citation of updated references.

Exosome-mediated microRNA transfer plays a role in radiation-induced bystander effect.

Science.gov (United States)

Xu, Shuai; Wang, Jufang; Ding, Nan; Hu, Wentao; Zhang, Xurui; Wang, Bing; Hua, Junrui; Wei, Wenjun; Zhu, Qiyun

2015-01-01

Bystander effects can be induced through cellular communication between irradiated cells and non-irradiated cells. The signals that mediate this cellular communication, such as cytokines, reactive oxygen species, nitric oxide and even microRNAs, can be transferred between cells via gap junctions or extracellular medium. We have previously reported that miR-21, a well described DDR (DNA damage response) microRNA, is involved in radiation-induced bystander effects through a medium-mediated way. However, the mechanisms of the microRNA transfer have not been elucidated in details. In the present study, it was found that exosomes isolated from irradiated conditioned medium could induce bystander effects. Furthermore, we demonstrated plenty of evidences that miR-21, which is up-regulated as a result of mimic transfection or irradiation, can be transferred from donor or irradiated cells into extracellular medium and subsequently get access to the recipient or bystander cells through exosomes to induce bystander effects. Inhibiting the miR-21 expression in advance can offset the bystander effects to some extent. From all of these results, it can be concluded that the exosome-mediated microRNA transfer plays an important role in the radiation-induced bystander effects. These findings provide new insights into the functions of microRNAs and the cellular communication between the directly irradiated cells and the non-irradiated cells.

Arabidopsis PCH2 Mediates Meiotic Chromosome Remodeling and Maturation of Crossovers.

Directory of Open Access Journals (Sweden)

Christophe Lambing

2015-07-01

Full Text Available Meiotic chromosomes are organized into linear looped chromatin arrays by a protein axis localized along the loop-bases. Programmed remodelling of the axis occurs during prophase I of meiosis. Structured illumination microscopy (SIM has revealed dynamic changes in the chromosome axis in Arabidopsis thaliana and Brassica oleracea. We show that the axis associated protein ASY1 is depleted during zygotene concomitant with synaptonemal complex (SC formation. Study of an Atpch2 mutant demonstrates this requires the conserved AAA+ ATPase, PCH2, which localizes to the sites of axis remodelling. Loss of PCH2 leads to a failure to deplete ASY1 from the axes and compromizes SC polymerisation. Immunolocalization of recombination proteins in Atpch2 indicates that recombination initiation and CO designation during early prophase I occur normally. Evidence suggests that CO interference is initially functional in the mutant but there is a defect in CO maturation following designation. This leads to a reduction in COs and a failure to form COs between some homologous chromosome pairs leading to univalent chromosomes at metaphase I. Genetic analysis reveals that CO distribution is also affected in some chromosome regions. Together these data indicate that the axis remodelling defect in Atpch2 disrupts normal patterned formation of COs.

Exosome-Mediated Genetic Information Transfer, a Missing Piece of Osteoblast-Osteoclast Communication Puzzle.

Science.gov (United States)

Yin, Pengbin; Lv, Houchen; Li, Yi; Deng, Yuan; Zhang, Licheng; Tang, Peifu

2017-01-01

The skeletal system functions and maintains itself based on communication between cells of diverse origins, especially between osteoblasts (OBs) and osteoclasts (OCs), accounting for bone formation and resorption, respectively. Previously, protein-level information exchange has been the research focus, and this has been discussed in detail. The regulative effects of microRNAs (miRNAs) on OB and OC ignite the question as to whether genetic information could be transferred between bone cells. Exosomes, extracellular membrane vesicles 30-100 nm in diameter, have recently been demonstrated to transfer functional proteins, mRNAs, and miRNAs, and serve as mediators of intercellular communication. By reviewing the distinguishing features of exosomes, a hypothesis was formulated and evaluated in this article that exosome-mediated genetic information transfer may represent a novel strategy for OB-OC communication. The exosomes may coordinately regulate these two cells under certain physiological conditions by transferring genetic information. Further research in exosome-shuttered miRNAs in OB-OC communication may add a missing piece to the bone cells communication "puzzle."

Coordination of KSHV Latent and Lytic Gene Control by CTCF-Cohesin Mediated Chromosome Conformation

Science.gov (United States)

Kang, Hyojeung; Wiedmer, Andreas; Yuan, Yan; Robertson, Erle; Lieberman, Paul M.

2011-01-01

Herpesvirus persistence requires a dynamic balance between latent and lytic cycle gene expression, but how this balance is maintained remains enigmatic. We have previously shown that the Kaposi's Sarcoma-Associated Herpesvirus (KSHV) major latency transcripts encoding LANA, vCyclin, vFLIP, v-miRNAs, and Kaposin are regulated, in part, by a chromatin organizing element that binds CTCF and cohesins. Using viral genome-wide chromatin conformation capture (3C) methods, we now show that KSHV latency control region is physically linked to the promoter regulatory region for ORF50, which encodes the KSHV immediate early protein RTA. Other linkages were also observed, including an interaction between the 5′ and 3′ end of the latency transcription cluster. Mutation of the CTCF-cohesin binding site reduced or eliminated the chromatin conformation linkages, and deregulated viral transcription and genome copy number control. siRNA depletion of CTCF or cohesin subunits also disrupted chromosomal linkages and deregulated viral latent and lytic gene transcription. Furthermore, the linkage between the latent and lytic control region was subject to cell cycle fluctuation and disrupted during lytic cycle reactivation, suggesting that these interactions are dynamic and regulatory. Our findings indicate that KSHV genomes are organized into chromatin loops mediated by CTCF and cohesin interactions, and that these inter-chromosomal linkages coordinate latent and lytic gene control. PMID:21876668

Intravascular local gene transfer mediated by protein-coated metallic stent.

Science.gov (United States)

Yuan, J; Gao, R; Shi, R; Song, L; Tang, J; Li, Y; Tang, C; Meng, L; Yuan, W; Chen, Z

2001-10-01

To assess the feasibility, efficiency and selectivity of adenovirus-mediated gene transfer to local arterial wall by protein-coated metallic stent. A replication-defective recombinant adenovirus carrying the Lac Z reporter gene for nuclear-specific beta-galactosidase (Ad-beta gal) was used in this study. The coating for metallic stent was made by immersing it in a gelatin solution containing crosslinker. The coated stents were mounted on a 4.0 or 3.0 mm percutaneous transluminal coronary angioplasty (PTCA) balloon and submersed into a high-titer Ad-beta gal viral stock (2 x 10(10) pfu/ml) for 3 min, and then implanted into the carotid arteries in 4 mini-swines and into the left anterior descending branch of the coronary artery in 2 mini-swines via 8F large lumen guiding catheters. The animals were sacrificed 7 (n = 4), 14 (n = 1) and 21 (n = 1) days after implantation, respectively. The beta-galactosidase expression was assessed by X-gal staining. The results showed that the expression of transgene was detected in all animal. In 1 of carotid artery with an intact intima, the beta-gal expression was limited to endothelial cells. In vessels with denuded endothelium, gene expression was found in the sub-intima, media and adventitia. The transfection efficiency of medial smooth muscle cells was 38.6%. In 2 animals sacrificed 7 days after transfection, a microscopic examination of X-gal-stained samples did not show evidence of transfection in remote organs and arterial segments adjacent to the treated arterial site. Adenovirus-mediated arterial gene transfer to endothelial, smooth muscle cells and adventitia by protein-coated metallic stent is feasible. The transfection efficiency is higher. The coated stent may act as a good carrier of adenovirus-mediated gene transfer and have a potential to prevent restenosis following PTCA.

Transfer of genetic information via isolated mammalian chromosomes

NARCIS (Netherlands)

G.J. Wullems

1976-01-01

textabstractRecombination of genetic information from different origin has provided insight in many aspects of the genetic mechanisms of the living cell. These aspects concern the location of genes on chromosomes, the regulation of gene expression and the interaction of different genes in the

Microdissection and molecular manipulation of single chromosomes in woody fruit trees with small chromosomes using pomelo (Citrus grandis) as a model. I. Construction of single chromosomal DNA libraries.

Science.gov (United States)

Huang, D; Wu, W; Zhou, Y; Hu, Z; Lu, L

2004-05-01

Construction of single chromosomal DNA libraries by means of chromosome microdissection and microcloning will be useful for genomic research, especially for those species that have not been extensively studied genetically. Application of the technology of microdissection and microcloning to woody fruit plants has not been reported hitherto, largely due to the generally small sizes of metaphase chromosomes and the difficulty of chromosome preparation. The present study was performed to establish a method for single chromosome microdissection and microcloning in woody fruit species using pomelo as a model. The standard karyotype of a pomelo cultivar ( Citrus grandis cv. Guanxi) was established based on 20 prometaphase photomicrographs. According to the standard karyotype, chromosome 1 was identified and isolated with fine glass microneedles controlled by a micromanipulator. DNA fragments ranging from 0.3 kb to 2 kb were acquired from the isolated single chromosome 1 via two rounds of PCR mediated by Sau3A linker adaptors and then cloned into T-easy vectors to generate a DNA library of chromosome 1. Approximately 30,000 recombinant clones were obtained. Evaluation based on 108 randomly selected clones showed that the sizes of the cloned inserts varied from 0.5 kb to 1.5 kb with an average of 860 bp. Our research suggests that microdissection and microcloning of single small chromosomes in woody plants is feasible.

Microdissection and molecular manipulation of single chromosomes in woody fruit trees with small chromosomes using pomelo (Citrus grandis) as a model. II. Cloning of resistance gene analogs from single chromosomes.

Science.gov (United States)

Huang, D; Wu, W; Lu, L

2004-05-01

Amplification of resistance gene analogs (RGAs) is both a useful method for acquiring DNA markers closely linked to disease resistance (R) genes and a potential approach for the rapid cloning of R genes in plants. However, the screening of target sequences from among the numerous amplified RGAs can be very laborious. The amplification of RGAs from specific chromosomes could greatly reduce the number of RGAs to be screened and, consequently, speed up the identification of target RGAs. We have developed two methods for amplifying RGAs from single chromosomes. Method 1 uses products of Sau3A linker adaptor-mediated PCR (LAM-PCR) from a single chromosome as the templates for RGA amplification, while Method 2 directly uses a single chromosomal DNA molecule as the template. Using a pair of degenerate primers designed on the basis of the conserved nucleotide-binding-site motifs in many R genes, RGAs were successfully amplified from single chromosomes of pomelo using both these methods. Sequencing and cluster analysis of RGA clones obtained from single chromosomes revealed the number, type and organization of R-gene clusters on the chromosomes. We suggest that Method 1 is suitable for analyzing chromosomes that are unidentifiable under a microscope, while Method 2 is more appropriate when chromosomes can be clearly identified.

On the length dependence of bridge-mediated electron transfer reactions

International Nuclear Information System (INIS)

Petrov, E.G.; Shevchenko, Ye.V.; May, V.

2003-01-01

Bridge-mediated nonadiabatic donor-acceptor (D-A) electron transfer (ET) is studied for the case of a regular molecular bridge of N identical units. It is shown that the multi-exponential ET kinetics reduces to a single-exponential transfer if, and only if, the integral population of the bridge remains small (less than 10 -2 ). An analytical expression for the overall D-A ET rate is derived and the necessary and sufficient conditions are formulated at which the rate is given as a sum of a superexchange and a sequential contribution. To describe experimental data on the N-dependence of ET reactions an approximate form of the overall transfer rate is derived. This expression is used to reproduce experimental data on distant ET through polyproline chains. Finally it is noted that the obtained analytical results can also be used for the description of more complex two-electron transfer reactions if the latter comprises separate single-electron pathways

Calcium ions function as a booster of chromosome condensation.

Science.gov (United States)

Phengchat, Rinyaporn; Takata, Hideaki; Morii, Kenichi; Inada, Noriko; Murakoshi, Hideji; Uchiyama, Susumu; Fukui, Kiichi

2016-12-02

Chromosome condensation is essential for the faithful transmission of genetic information to daughter cells during cell division. The depletion of chromosome scaffold proteins does not prevent chromosome condensation despite structural defects. This suggests that other factors contribute to condensation. Here we investigated the contribution of divalent cations, particularly Ca 2+ , to chromosome condensation in vitro and in vivo. Ca 2+ depletion caused defects in proper mitotic progression, particularly in chromosome condensation after the breakdown of the nuclear envelope. Fluorescence lifetime imaging microscopy-Förster resonance energy transfer and electron microscopy demonstrated that chromosome condensation is influenced by Ca 2+ . Chromosomes had compact globular structures when exposed to Ca 2+ and expanded fibrous structures without Ca 2+ . Therefore, we have clearly demonstrated a role for Ca 2+ in the compaction of chromatin fibres.

The colocalization transition of homologous chromosomes at meiosis

Science.gov (United States)

Nicodemi, Mario; Panning, Barbara; Prisco, Antonella

2008-06-01

Meiosis is the specialized cell division required in sexual reproduction. During its early stages, in the mother cell nucleus, homologous chromosomes recognize each other and colocalize in a crucial step that remains one of the most mysterious of meiosis. Starting from recent discoveries on the system molecular components and interactions, we discuss a statistical mechanics model of chromosome early pairing. Binding molecules mediate long-distance interaction of special DNA recognition sequences and, if their concentration exceeds a critical threshold, they induce a spontaneous colocalization transition of chromosomes, otherwise independently diffusing.

SMC Progressively Aligns Chromosomal Arms in Caulobacter crescentus but Is Antagonized by Convergent Transcription

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Ngat T. Tran

2017-08-01

Full Text Available The structural maintenance of chromosomes (SMC complex plays an important role in chromosome organization and segregation in most living organisms. In Caulobacter crescentus, SMC is required to align the left and the right arms of the chromosome that run in parallel down the long axis of the cell. However, the mechanism of SMC-mediated alignment of chromosomal arms remains elusive. Here, using genome-wide methods and microscopy of single cells, we show that Caulobacter SMC is recruited to the centromeric parS site and that SMC-mediated arm alignment depends on the chromosome-partitioning protein ParB. We provide evidence that SMC likely tethers the parS-proximal regions of the chromosomal arms together, promoting arm alignment. Furthermore, we show that highly transcribed genes near parS that are oriented against SMC translocation disrupt arm alignment, suggesting that head-on transcription interferes with SMC translocation. Our results demonstrate a tight interdependence of bacterial chromosome organization and global patterns of transcription.

Trust, Knowledge Creation and Mediating Effects of Knowledge Transfer Processes

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Anna Sankowska

2016-01-01

Full Text Available This paper investigates how organizational trust contributes to knowledge transfer processes and knowledge creation both directly and indirectly. The mediation analyses were used. The findings using cross-sectional data from Polish companies suggest a new way of thinking in respect of how crucial is trust to creation of knowledge unconnected to organizational memory.

H-NS Facilitates Sequence Diversification of Horizontally Transferred DNAs during Their Integration in Host Chromosomes.

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Koichi Higashi

2016-01-01

Full Text Available Bacteria can acquire new traits through horizontal gene transfer. Inappropriate expression of transferred genes, however, can disrupt the physiology of the host bacteria. To reduce this risk, Escherichia coli expresses the nucleoid-associated protein, H-NS, which preferentially binds to horizontally transferred genes to control their expression. Once expression is optimized, the horizontally transferred genes may actually contribute to E. coli survival in new habitats. Therefore, we investigated whether and how H-NS contributes to this optimization process. A comparison of H-NS binding profiles on common chromosomal segments of three E. coli strains belonging to different phylogenetic groups indicated that the positions of H-NS-bound regions have been conserved in E. coli strains. The sequences of the H-NS-bound regions appear to have diverged more so than H-NS-unbound regions only when H-NS-bound regions are located upstream or in coding regions of genes. Because these regions generally contain regulatory elements for gene expression, sequence divergence in these regions may be associated with alteration of gene expression. Indeed, nucleotide substitutions in H-NS-bound regions of the ybdO promoter and coding regions have diversified the potential for H-NS-independent negative regulation among E. coli strains. The ybdO expression in these strains was still negatively regulated by H-NS, which reduced the effect of H-NS-independent regulation under normal growth conditions. Hence, we propose that, during E. coli evolution, the conservation of H-NS binding sites resulted in the diversification of the regulation of horizontally transferred genes, which may have facilitated E. coli adaptation to new ecological niches.

H-NS Facilitates Sequence Diversification of Horizontally Transferred DNAs during Their Integration in Host Chromosomes.

Science.gov (United States)

Higashi, Koichi; Tobe, Toru; Kanai, Akinori; Uyar, Ebru; Ishikawa, Shu; Suzuki, Yutaka; Ogasawara, Naotake; Kurokawa, Ken; Oshima, Taku

2016-01-01

Bacteria can acquire new traits through horizontal gene transfer. Inappropriate expression of transferred genes, however, can disrupt the physiology of the host bacteria. To reduce this risk, Escherichia coli expresses the nucleoid-associated protein, H-NS, which preferentially binds to horizontally transferred genes to control their expression. Once expression is optimized, the horizontally transferred genes may actually contribute to E. coli survival in new habitats. Therefore, we investigated whether and how H-NS contributes to this optimization process. A comparison of H-NS binding profiles on common chromosomal segments of three E. coli strains belonging to different phylogenetic groups indicated that the positions of H-NS-bound regions have been conserved in E. coli strains. The sequences of the H-NS-bound regions appear to have diverged more so than H-NS-unbound regions only when H-NS-bound regions are located upstream or in coding regions of genes. Because these regions generally contain regulatory elements for gene expression, sequence divergence in these regions may be associated with alteration of gene expression. Indeed, nucleotide substitutions in H-NS-bound regions of the ybdO promoter and coding regions have diversified the potential for H-NS-independent negative regulation among E. coli strains. The ybdO expression in these strains was still negatively regulated by H-NS, which reduced the effect of H-NS-independent regulation under normal growth conditions. Hence, we propose that, during E. coli evolution, the conservation of H-NS binding sites resulted in the diversification of the regulation of horizontally transferred genes, which may have facilitated E. coli adaptation to new ecological niches.

A bipartite signal mediates the transfer of type IV secretion substrates of Bartonella henselae into human cells.

Science.gov (United States)

Schulein, Ralf; Guye, Patrick; Rhomberg, Thomas A; Schmid, Michael C; Schröder, Gunnar; Vergunst, Annette C; Carena, Ilaria; Dehio, Christoph

2005-01-18

Bacterial type IV secretion (T4S) systems mediate the transfer of macromolecular substrates into various target cells, e.g., the conjugative transfer of DNA into bacteria or the transfer of virulence proteins into eukaryotic host cells. The T4S apparatus VirB of the vascular tumor-inducing pathogen Bartonella henselae causes subversion of human endothelial cell (HEC) function. Here we report the identification of multiple protein substrates of VirB, which, upon translocation into HEC, mediate all known VirB-dependent cellular changes. These Bartonella-translocated effector proteins (Beps) A-G are encoded together with the VirB system and the T4S coupling protein VirD4 on a Bartonella-specific pathogenicity island. The Beps display a modular architecture, suggesting an evolution by extensive domain duplication and reshuffling. The C terminus of each Bep harbors at least one copy of the Bep-intracellular delivery domain and a short positively charged tail sequence. This biparte C terminus constitutes a transfer signal that is sufficient to mediate VirB/VirD4-dependent intracellular delivery of reporter protein fusions. The Bep-intracellular delivery domain is also present in conjugative relaxases of bacterial conjugation systems. We exemplarily show that the C terminus of such a conjugative relaxase mediates protein transfer through the Bartonella henselae VirB/VirD4 system into HEC. Conjugative relaxases may thus represent the evolutionary origin of the here defined T4S signal for protein transfer into human cells.

Alliance entrepreneurship and entrepreneurial orientation: the mediating effect of knowledge transfer

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Arash Rezazadeh

2016-12-01

Full Text Available Today’s rapidly changing business environment has impelled companies to cooperate with their competitors gaining more competitive advantages by achieving win-win situation. Thereby, building alliances as one of the cooperative strategies has been adopted by many enterprises, consequently attracting great attention of numerous scholars. Nevertheless, the alliance literature seems to lack studies in the domain of entrepreneurship. Accordingly, this paper aims to extend entrepreneurship into the field of alliances highlighting two phenomenal concepts: alliance entrepreneurship and entrepreneurial orientation. Hence, the relationship between these two constructs together with the mediating role of knowledge transfer between alliance partners are investigated. We used Structural Equation Modeling with Partial Least Squares (PLS-SEM technique under two sections of measurement model and structural model assessment in order to analyze data. The results gathered from Iran’s automotive industry confirmed the positive significant impact of alliance entrepreneurship on partners’ entrepreneurial orientation and the mediating effect of knowledge transfer

Balanced Chromosomal Translocation of Chromosomes 6 and 7: A Rare Male Factor of Spontaneous Abortions

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Sefa Resim

2013-06-01

Full Text Available Background: Carriers of structural chromosomal rearrangements such as Robertsonian or reciprocal translocations have an increased risk of spontaneous abortion and producing offspring with genetic abnormalities. Case Report: We report a man with balanced chromosomal translocations located at 6p22, and 7q22. His wife has a history of four spontaneous abortions. Conclusion: Couples with a history of abortions should be investigated cytogenetically, after other causes of miscarriages are excluded. The possibility of spontaneous abortions can be reduced with preimplantation genetic diagnosis (PGD before embryo transfer.

Recombinant adenovirus-mediated gene transfer suppresses experimental arthritis

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E. Quattrocchi

2011-09-01

Full Text Available Collagen Induced Arthritis (CIA is a widely studied animal model to develop and test novel therapeutic approaches for treating Rheumatoid Arthritis (RA in humans. Soluble Cytotoxic T-Lymphocyte Antigen 4 (CTLA4-Ig, which binds B7 molecule on antigen presenting cells and blocks CD28 mediated T-lymphocyte activation, has been shown to ameliorate experimental autoimmune diseases such as lupus, diabetes and CIA. Objective of our research was to investigate in vivo the effectiveness of blocking the B7/CD28 T-lymphocyte co-stimulatory pathway, utilizing a gene transfer technology, as a therapeutic strategy against CIA. Replication-deficient adenoviruses encoding a chimeric CTLA4-Ig fusion protein, or β-galactosidase as control, have been injected intravenously once at arthritis onset. Disease activity has been monitored by the assessment of clinical score, paw thickness and type II collagen (CII specific cellular and humoral immune responses for 21 days. The adenovirally delivered CTLA4-Ig fusion protein at a dose of 2×108 pfu suppressed established CIA, whereas the control β-galactosidase did not significantly affect the disease course. CII-specific lymphocyte proliferation, IFNg production and anti-CII antibodies were significantly reduced by CTLA4-Ig treatment. Our results demonstrate that blockade of the B7/CD28 co-stimulatory pathway by adenovirus-mediated CTLA4-Ig gene transfer is effective in treating established CIA suggesting its potential in treating RA.

Isolation of Chromatin from Dysfunctional Telomeres Reveals an Important Role for Ring1b in NHEJ-Mediated Chromosome Fusions

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Cristina Bartocci

2014-05-01

Full Text Available When telomeres become critically short, DNA damage response factors are recruited at chromosome ends, initiating a cellular response to DNA damage. We performed proteomic isolation of chromatin fragments (PICh in order to define changes in chromatin composition that occur upon onset of acute telomere dysfunction triggered by depletion of the telomere-associated factor TRF2. This unbiased purification of telomere-associated proteins in functional or dysfunctional conditions revealed the dynamic changes in chromatin composition that take place at telomeres upon DNA damage induction. On the basis of our results, we describe a critical role for the polycomb group protein Ring1b in nonhomologous end-joining (NHEJ-mediated end-to-end chromosome fusions. We show that cells with reduced levels of Ring1b have a reduced ability to repair uncapped telomeric chromatin. Our data represent an unbiased isolation of chromatin undergoing DNA damage and are a valuable resource to map the changes in chromatin composition in response to DNA damage activation.

X-Inactivation: Xist RNA Uses Chromosome Contacts to Coat the X

OpenAIRE

Leung, Karen N.; Panning, Barbara

2014-01-01

The mechanisms by which Xist RNA associates with the X chromosome to mediate alterations in chromatin structure remain mysterious. Recent genome-wide Xist RNA distribution studies suggest that this long noncoding RNA uses 3-dimensional chromosome contacts to move to its sites of action.

Current Status of Comprehensive Chromosome Screening for Elective Single-Embryo Transfer

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Ming-Yih Wu

2014-01-01

Full Text Available Most in vitro fertilization (IVF experts and infertility patients agree that the most ideal assisted reproductive technology (ART outcome is to have a healthy, full-term singleton born. To this end, the most reliable policy is the single-embryo transfer (SET. However, unsatisfactory results in IVF may result from plenty of factors, in which aneuploidy associated with advanced maternal age is a major hurdle. Throughout the past few years, we have got a big leap in advancement of the genetic screening of embryos on aneuploidy, translocation, or mutations. This facilitates a higher success rate in IVF accompanied by the policy of elective SET (eSET. As the cost is lowering while the scale of genome characterization continues to be up over the recent years, the contemporary technologies on trophectoderm biopsy and freezing-thaw, comprehensive chromosome screening (CCS with eSET appear to be getting more and more popular for modern IVF centers. Furthermore, evidence has showen that, by these avant-garde techniques (trophectoderm biopsy, vitrification, and CCS, older infertile women with the help of eSET may have an opportunity to increase the success of their live birth rates approaching those reported in younger infertility patients.

Electron Transfer Mediators for Photoelectrochemical Cells Based on Cu(I Metal Complexes

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Michele Brugnati

2007-01-01

Full Text Available The preparation and the photoelectrochemical characterization of a series of bipyridine and pyridyl-quinoline Cu(I complexes, used as electron transfer mediators in regenerative photoelectrochemical cells, are reported. The best performing mediators produced maximum IPCEs of the order of 35–40%. The J-V curves recorded under monochromatic light showed that the selected Cu(I/(II couples generated higher Vocs and fill factors compared to an equivalent I-/I3- cell, due to a decreased dark current.

Growth Conditions Regulate the Requirements for Caulobacter Chromosome Segregation

DEFF Research Database (Denmark)

Shebelut, Conrad W.; Jensen, Rasmus Bugge; Gitai, Zemer

2009-01-01

Growth environments are important metabolic and developmental regulators. Here we demonstrate a growth environment-dependent effect on Caulobacter chromosome segregation of a small-molecule inhibitor of the MreB bacterial actin cytoskeleton. Our results also implicate ParAB as important segregation...... determinants, suggesting that multiple distinct mechanisms can mediate Caulobacter chromosome segregation and that their relative contributions can be environmentally regulated....

Plant–Agrobacterium interaction mediated by ethylene and super-Agrobacterium conferring efficient gene transfer

OpenAIRE

Nonaka, Satoko; Ezura, Hiroshi

2014-01-01

Agrobacterium tumefaciens has a unique ability to transfer genes into plant genomes. This ability has been utilized for plant genetic engineering. However, the efficiency is not sufficient for all plant species. Several studies have shown that ethylene decreased the Agrobacterium-mediated transformation frequency. Thus, A. tumefaciens with an ability to suppress ethylene evolution would increase the efficiency of Agrobacterium-mediated transformation. Some studies showed that plant growth-pro...

Satellite DNA-based artificial chromosomes for use in gene therapy.

Science.gov (United States)

Hadlaczky, G

2001-04-01

Satellite DNA-based artificial chromosomes (SATACs) can be made by induced de novo chromosome formation in cells of different mammalian species. These artificially generated accessory chromosomes are composed of predictable DNA sequences and they contain defined genetic information. Prototype human SATACs have been successfully constructed in different cell types from 'neutral' endogenous DNA sequences from the short arm of the human chromosome 15. SATACs have already passed a number of hurdles crucial to their further development as gene therapy vectors, including: large-scale purification; transfer of purified artificial chromosomes into different cells and embryos; generation of transgenic animals and germline transmission with purified SATACs; and the tissue-specific expression of a therapeutic gene from an artificial chromosome in the milk of transgenic animals.

An Examination of Mediators of the Transfer of Cognitive Speed of Processing Training to Everyday Functional Performance

OpenAIRE

Edwards, Jerri D.; Ruva, Christine L.; O’Brien, Jennifer L.; Haley, Christine B.; Lister, Jennifer J.

2012-01-01

The purpose of these analyses was to examine mediators of the transfer of cognitive speed of processing training to improved everyday functional performance (Edwards, Wadley, Vance, Roenker, & Ball, 2005). Cognitive speed of processing and visual attention (as measured by the Useful Field of View Test; UFOV) were examined as mediators of training transfer. Secondary data analyses were conducted from the Staying Keen in Later Life (SKILL) study, a randomized cohort study including 126 communit...

Sensitivity of Electron Transfer Mediated Decay to Ion Pairing.

Science.gov (United States)

Pohl, Marvin N; Richter, Clemens; Lugovoy, Evgeny; Seidel, Robert; Slavíček, Petr; Aziz, Emad F; Abel, Bernd; Winter, Bernd; Hergenhahn, Uwe

2017-08-17

Ion pairing in electrolyte solutions remains a topic of discussion despite a long history of research. Very recently, nearest-neighbor mediated electronic de-excitation processes of core hole vacancies (electron transfer mediated decay, ETMD) were proposed to carry a spectral fingerprint of local solvation structure and in particular of contact ion pairs. Here, for the first time, we apply electron-electron coincidence detection to a liquid microjet, and record ETMD spectra of Li 1s vacancies in aqueous solutions of lithium chloride (LiCl) in direct comparison to lithium acetate (LiOAc). A change in the ETMD spectrum dependent on the electrolyte anion identity is observed for 4.5 M salt concentration. We discuss these findings within the framework of the formation and presence of contact ion pairs and the unique sensitivity of ETMD spectroscopy to ion pairing.

Optimal design of implants for magnetically mediated hyperthermia: A wireless power transfer approach

Science.gov (United States)

Lang, Hans-Dieter; Sarris, Costas D.

2017-09-01

In magnetically mediated hyperthermia (MMH), an externally applied alternating magnetic field interacts with a mediator (such as a magnetic nanoparticle or an implant) inside the body to heat up the tissue in its proximity. Producing heat via induced currents in this manner is strikingly similar to wireless power transfer (WPT) for implants, where power is transferred from a transmitter outside of the body to an implanted receiver, in most cases via magnetic fields as well. Leveraging this analogy, a systematic method to design MMH implants for optimal heating efficiency is introduced, akin to the design of WPT systems for optimal power transfer efficiency. This paper provides analytical formulas for the achievable heating efficiency bounds as well as the optimal operating frequency and the implant material. Multiphysics simulations validate the approach and further demonstrate that optimization with respect to maximum heating efficiency is accompanied by minimizing heat delivery to healthy tissue. This is a property that is highly desirable when considering MMH as a key component or complementary method of cancer treatment and other applications.

Area-Specific Cell Stimulation via Surface-Mediated Gene Transfer Using Apatite-Based Composite Layers

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Yushin Yazaki

2015-04-01

Full Text Available Surface-mediated gene transfer systems using biocompatible calcium phosphate (CaP-based composite layers have attracted attention as a tool for controlling cell behaviors. In the present study we aimed to demonstrate the potential of CaP-based composite layers to mediate area-specific dual gene transfer and to stimulate cells on an area-by-area basis in the same well. For this purpose we prepared two pairs of DNA–fibronectin–apatite composite (DF-Ap layers using a pair of reporter genes and pair of differentiation factor genes. The results of the area-specific dual gene transfer successfully demonstrated that the cells cultured on a pair of DF-Ap layers that were adjacently placed in the same well showed specific gene expression patterns depending on the gene that was immobilized in theunderlying layer. Moreover, preliminary real-time PCR results indicated that multipotential C3H10T1/2 cells may have a potential to change into different types of cells depending on the differentiation factor gene that was immobilized in the underlying layer, even in the same well. Because DF-Ap layers have a potential to mediate area-specific cell stimulation on their surfaces, they could be useful in tissue engineering applications.

Beyond the chromosome: the prevalence of unique extra-chromosomal bacteriophages with integrated virulence genes in pathogenic Staphylococcus aureus.

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Bryan Utter

Full Text Available In Staphylococcus aureus, the disease impact of chromosomally integrated prophages on virulence is well described. However, the existence of extra-chromosomal prophages, both plasmidial and episomal, remains obscure. Despite the recent explosion in bacterial and bacteriophage genomic sequencing, studies have failed to specifically focus on extra-chromosomal elements. We selectively enriched and sequenced extra-chromosomal DNA from S. aureus isolates using Roche-454 technology and uncovered evidence for the widespread distribution of multiple extra-chromosomal prophages (ExPΦs throughout both antibiotic-sensitive and -resistant strains. We completely sequenced one such element comprised of a 43.8 kbp, circular ExPΦ (designated ФBU01 from a vancomycin-intermediate S. aureus (VISA strain. Assembly and annotation of ФBU01 revealed a number of putative virulence determinants encoded within a bacteriophage immune evasion cluster (IEC. Our identification of several potential ExPΦs and mobile genetic elements (MGEs also revealed numerous putative virulence factors and antibiotic resistance genes. We describe here a previously unidentified level of genetic diversity of stealth extra-chromosomal elements in S. aureus, including phages with a larger presence outside the chromosome that likely play a prominent role in pathogenesis and strain diversity driven by horizontal gene transfer (HGT.

Role of chromosome stability and telomere length in the production of viable cell lines for somatic cell nuclear transfer

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Betts Dean H

2006-08-01

Full Text Available Abstract Background Somatic cell nuclear transfer (SCNT provides an appealing alternative for the preservation of genetic material in non-domestic and endangered species. An important prerequisite for successful SCNT is the availability of good quality donor cells, as normal embryo development is dependent upon proper reprogramming of the donor genome so that embryonic genes can be appropriately expressed. The characteristics of donor cell lines and their ability to produce embryos by SCNT were evaluated by testing the effects of tissue sample collection (DART biopsy, PUNCH biopsy, post-mortem EAR sample and culture initiation (explant, collagenase digestion techniques. Results Differences in initial sample size based on sample collection technique had an effect on the amount of time necessary for achieving primary confluence and the number of population doublings (PDL produced. Thus, DART and PUNCH biopsies resulted in cultures with decreased lifespans (50 PDL and chromosomally stable (>70% normal cells at 20 PDL cultures produced by post-mortem EAR samples. Chromosome stability was influenced by sample collection technique and was dependent upon the culture's initial telomere length and its rate of shortening over cell passages. Following SCNT, short-lived cultures resulted in significantly lower blastocyst development (≤ 0.9% compared to highly proliferative cultures (11.8%. Chromosome stability and sample collection technique were significant factors in determining blastocyst development outcome. Conclusion These data demonstrate the influence of culture establishment techniques on cell culture characteristics, including the viability, longevity and normality of cells. The identification of a quantifiable marker associated with SCNT embryo developmental potential, chromosome stability, provides a means by which cell culture conditions can be monitored and improved.

Taxane-mediated radiosensitization derives from chromosomal missegregation on tripolar mitotic spindles orchestrated by AURKA and TPX2.

Science.gov (United States)

Orth, M; Unger, K; Schoetz, U; Belka, C; Lauber, K

2018-01-04

Taxane-based radiochemotherapy is a central treatment option for various cancer entities in locally advanced stages. The therapeutic synergism of this combined modality approach due to taxane-mediated radiosensitization of cancer cells is well-known. However, the underlying molecular mechanisms remain largely elusive, and mechanism-derived predictive markers of taxane-based radiochemotherapy are currently not available. Here, we show that clinically relevant doses of Paclitaxel, the prototype taxane, stimulate a tripolar mode of mitosis leading to chromosomal missegregation and aneuploidization rather than interfering with cell cycle progression. This distinct mitotic phenotype was interlinked with Paclitaxel-mediated radiosensitization via overexpression of mitotic Aurora kinase A (AURKA) and its cofactor TPX2 whose knockdown rescued the bipolar mode of cell division and largely attenuated the radiosensitizing effects of Paclitaxel. In the cancer genome atlas (TCGA) lung adenocarcinoma cohort, high expression levels of AURKA and TPX2 were associated with specifically improved overall survival upon taxane-based radiochemotherapy, but not in case of non-taxane-based radiochemotherapy, chemo- or radiotherapy only. Thus, our data provide insights into Paclitaxel-mediated radiosensitization on a mechanistic and molecular level and identify AURKA and TPX2 as the first potential mechanism-based, predictive markers of taxane-based radiochemotherapy.

General theory of excitation energy transfer in donor-mediator-acceptor systems.

Science.gov (United States)

Kimura, Akihiro

2009-04-21

General theory of the excitation energy transfer (EET) in the case of donor-mediator-acceptor system was constructed by using generalized master equation (GME). In this theory, we consider the direct and indirect transitions in the EET consistently. Hence, our theory includes the quantum mechanical interference between the direct and indirect transitions automatically. Memory functions in the GME were expressed by the overlap integrals among the time-dependent emission spectrum of the donor, the absorption spectrum of the mediator, the time-dependent emission spectrum of the mediator, and the absorption spectrum of the acceptor. In the Markov limit of the memory functions, we obtained the rate of EET which consists of three terms due to the direct transition, the indirect transition, and the interference between them. We found that the interference works effectively in the limit of slow thermalization at the intermediate state. The formula of EET rate in this limit was expressed by the convolution of the EET interaction and optical spectra. The interference effect strongly depends on the width of the absorption spectrum of mediator molecule and the energy gap between the donor and the mediator molecules.

X-inactivation: Xist RNA uses chromosome contacts to coat the X.

Science.gov (United States)

Leung, Karen N; Panning, Barbara

2014-01-20

The mechanisms by which Xist RNA associates with the X chromosome to mediate alterations in chromatin structure remain mysterious. Recent genome-wide Xist RNA distribution studies suggest that this long noncoding RNA uses 3-dimensional chromosome contacts to move to its sites of action. Copyright © 2014 Elsevier Ltd. All rights reserved.

Assignment of electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO) to human chromosome 4q33 by fluorescence in situ hybridization and somatic cell hybridization.

Science.gov (United States)

Spector, E B; Seltzer, W K; Goodman, S I

1999-08-01

Electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO) is a nuclear-encoded protein located in the inner mitochondrial membrane. Inherited defects of ETF-QO cause glutaric acidemia type II. We here describe the localization of the ETF-QO gene to human chromosome 4q33 by somatic cell hybridization and fluorescence in situ hybridization. Copyright 1999 Academic Press.

Functional Identification of the Plasmodium Centromere and Generation of a Plasmodium Artificial Chromosome

OpenAIRE

Iwanaga, Shiroh; Khan, Shahid M.; Kaneko, Izumi; Christodoulou, Zoe; Newbold, Chris; Yuda, Masao; Janse, Chris J.; Waters, Andrew P.

2010-01-01

Summary The artificial chromosome represents a useful tool for gene transfer, both as cloning vectors and in chromosome biology research. To generate a Plasmodium artificial chromosome (PAC), we had to first functionally identify and characterize the parasite's centromere. A putative centromere (pbcen5) was cloned from chromosome 5 of the rodent parasite P. berghei based on a Plasmodium gene-synteny map. Plasmids containing pbcen5 were stably maintained in parasites during a blood-stage infec...

Extensive duplication of the Wolbachia DNA in chromosome four of Drosophila ananassae.

Science.gov (United States)

Klasson, Lisa; Kumar, Nikhil; Bromley, Robin; Sieber, Karsten; Flowers, Melissa; Ott, Sandra H; Tallon, Luke J; Andersson, Siv G E; Dunning Hotopp, Julie C

2014-12-12

Lateral gene transfer (LGT) from bacterial Wolbachia endosymbionts has been detected in ~20% of arthropod and nematode genome sequencing projects. Many of these transfers are large and contain a substantial part of the Wolbachia genome. Here, we re-sequenced three D. ananassae genomes from Asia and the Pacific that contain large LGTs from Wolbachia. We find that multiple copies of the Wolbachia genome are transferred to the Drosophila nuclear genome in all three lines. In the D. ananassae line from Indonesia, the copies of Wolbachia DNA in the nuclear genome are nearly identical in size and sequence yielding an even coverage of mapped reads over the Wolbachia genome. In contrast, the D. ananassae lines from Hawaii and India show an uneven coverage of mapped reads over the Wolbachia genome suggesting that different parts of these LGTs are present in different copy numbers. In the Hawaii line, we find that this LGT is underrepresented in third instar larvae indicative of being heterochromatic. Fluorescence in situ hybridization of mitotic chromosomes confirms that the LGT in the Hawaii line is heterochromatic and represents ~20% of the sequence on chromosome 4 (dot chromosome, Muller element F). This collection of related lines contain large lateral gene transfers composed of multiple Wolbachia genomes that constitute >2% of the D. ananassae genome (~5 Mbp) and partially explain the abnormally large size of chromosome 4 in D. ananassae.

The Emerging Role of the Cytoskeleton in Chromosome Dynamics

Directory of Open Access Journals (Sweden)

Maya Spichal

2017-05-01

Full Text Available Chromosomes underlie a dynamic organization that fulfills functional roles in processes like transcription, DNA repair, nuclear envelope stability, and cell division. Chromosome dynamics depend on chromosome structure and cannot freely diffuse. Furthermore, chromosomes interact closely with their surrounding nuclear environment, which further constrains chromosome dynamics. Recently, several studies enlighten that cytoskeletal proteins regulate dynamic chromosome organization. Cytoskeletal polymers that include actin filaments, microtubules and intermediate filaments can connect to the nuclear envelope via Linker of the Nucleoskeleton and Cytoskeleton (LINC complexes and transfer forces onto chromosomes inside the nucleus. Monomers of these cytoplasmic polymers and related proteins can also enter the nucleus and play different roles in the interior of the nucleus than they do in the cytoplasm. Nuclear cytoskeletal proteins can act as chromatin remodelers alone or in complexes with other nuclear proteins. They can also act as transcription factors. Many of these mechanisms have been conserved during evolution, indicating that the cytoskeletal regulation of chromosome dynamics is an essential process. In this review, we discuss the different influences of cytoskeletal proteins on chromosome dynamics by focusing on the well-studied model organism budding yeast.

DNA-mediated gene transfer into human diploid fibroblasts derived from normal and ataxia-telangiectasia donors: parameters for DNA transfer and properties of DNA transformants

International Nuclear Information System (INIS)

Debenham, P.G.; Webb, M.B.T.; Masson, W.K.; Cox, R.

1984-01-01

An investigation was made of the feasibility of DNA-mediated gene transfer into human diploid fibroblasts derived from patients with the radiation sensitive syndrome ataxia-telangiectasia (A-T) and from a normal donor. Although they are markedly different in their growth characteristics, both normal and A-T strains give similar frequencies for DNA transfer in a model system using the recombinant plasmid pSV2-gpt. pSV2-gpt DNA transformants arise with a frequency between 10 -5 and 10 -4 per viable cell. Analysis of such transformants, although possible, is severely handicapped by the limited clonal life span of diploid human cells. Despite these problems it may be concluded that diploid human fibroblasts are competent recipients for DNA-mediated gene transfer and the putative repair deficiency of A-T does not markedly effect the efficiency of this process. (author)

Efficient generation of recombinant RNA viruses using targeted recombination-mediated mutagenesis of bacterial artificial chromosomes containing full-length cDNA

DEFF Research Database (Denmark)

Rasmussen, Thomas Bruun; Risager, Peter Christian; Fahnøe, Ulrik

2013-01-01

Background Infectious cDNA clones are a prerequisite for directed genetic manipulation of RNA viruses. Here, a strategy to facilitate manipulation and rescue of classical swine fever viruses (CSFVs) from full-length cDNAs present within bacterial artificial chromosomes (BACs) is described....... This strategy allows manipulation of viral cDNA by targeted recombination-mediated mutagenesis within bacteria. Results A new CSFV-BAC (pBeloR26) derived from the Riems vaccine strain has been constructed and subsequently modified in the E2 coding sequence, using the targeted recombination strategy to enable...

Neurotransmitter-Triggered Transfer of Exosomes Mediates Oligodendrocyte–Neuron Communication

Science.gov (United States)

Kuo, Wen Ping; Amphornrat, Jesa; Thilemann, Sebastian; Saab, Aiman S.; Kirchhoff, Frank; Möbius, Wiebke; Goebbels, Sandra; Nave, Klaus-Armin; Schneider, Anja; Simons, Mikael; Klugmann, Matthias; Trotter, Jacqueline; Krämer-Albers, Eva-Maria

2013-01-01

Reciprocal interactions between neurons and oligodendrocytes are not only crucial for myelination, but also for long-term survival of axons. Degeneration of axons occurs in several human myelin diseases, however the molecular mechanisms of axon-glia communication maintaining axon integrity are poorly understood. Here, we describe the signal-mediated transfer of exosomes from oligodendrocytes to neurons. These endosome-derived vesicles are secreted by oligodendrocytes and carry specific protein and RNA cargo. We show that activity-dependent release of the neurotransmitter glutamate triggers oligodendroglial exosome secretion mediated by Ca2+ entry through oligodendroglial NMDA and AMPA receptors. In turn, neurons internalize the released exosomes by endocytosis. Injection of oligodendroglia-derived exosomes into the mouse brain results in functional retrieval of exosome cargo in neurons. Supply of cultured neurons with oligodendroglial exosomes improves neuronal viability under conditions of cell stress. These findings indicate that oligodendroglial exosomes participate in a novel mode of bidirectional neuron-glia communication contributing to neuronal integrity. PMID:23874151

Neurotransmitter-triggered transfer of exosomes mediates oligodendrocyte-neuron communication.

Science.gov (United States)

Frühbeis, Carsten; Fröhlich, Dominik; Kuo, Wen Ping; Amphornrat, Jesa; Thilemann, Sebastian; Saab, Aiman S; Kirchhoff, Frank; Möbius, Wiebke; Goebbels, Sandra; Nave, Klaus-Armin; Schneider, Anja; Simons, Mikael; Klugmann, Matthias; Trotter, Jacqueline; Krämer-Albers, Eva-Maria

2013-07-01

Reciprocal interactions between neurons and oligodendrocytes are not only crucial for myelination, but also for long-term survival of axons. Degeneration of axons occurs in several human myelin diseases, however the molecular mechanisms of axon-glia communication maintaining axon integrity are poorly understood. Here, we describe the signal-mediated transfer of exosomes from oligodendrocytes to neurons. These endosome-derived vesicles are secreted by oligodendrocytes and carry specific protein and RNA cargo. We show that activity-dependent release of the neurotransmitter glutamate triggers oligodendroglial exosome secretion mediated by Ca²⁺ entry through oligodendroglial NMDA and AMPA receptors. In turn, neurons internalize the released exosomes by endocytosis. Injection of oligodendroglia-derived exosomes into the mouse brain results in functional retrieval of exosome cargo in neurons. Supply of cultured neurons with oligodendroglial exosomes improves neuronal viability under conditions of cell stress. These findings indicate that oligodendroglial exosomes participate in a novel mode of bidirectional neuron-glia communication contributing to neuronal integrity.

Homologous subfamilies of human alphoid repetitive DNA on different nucleolus organizing chromosomes

International Nuclear Information System (INIS)

Joergensen, A.L.; Bostock, C.J.; Bak, A.L.

1987-01-01

The organization of alphoid repeated sequences on human nucleolus-organizing (NOR) chromosomes 13, 21, and 22 has been investigated. Analysis of hybridization of alphoid DNA probes to Southern transfers of restriction enzyme-digested DNA fragments from hybrid cells containing single human chromosomes shows that chromosomes 13 and 21 share one subfamily of alphoid repeats, whereas a different subfamily may be held in common by chromosomes 13 and 22. The sequences of cloned 680-base-pair EcoRI fragments of the alphoid DNA from chromosomes 13 and 21 show that the basic unit of this subfamily is indistinguishable on each chromosome. The sequence of cloned 1020-base-pair Xba I fragments from chromosome 22 is related to, but distinguishable from, that of the 680-base-pair EcoRI alphoid subfamily of chromosomes 13 and 21. These results suggest that, at some point after they originated and were homogenized, different subfamilies of alphoid sequences must have exchanged between chromosomes 13 and 21 and separately between chromosomes 13 and 22

Effective generation of transgenic pigs and mice by linker based sperm-mediated gene transfer.

OpenAIRE

Chang, Keejong; Qian, Jin; Jiang, MeiSheng; Liu, Yi-Hsin; Wu, Ming-Che; Chen, Chi-Dar; Lai, Chao-Kuen; Lo, Hsin-Lung; Hsiao, Chin-Ton; Brown, Lucy; Bolen, James; Huang, Hsiao-I; Ho, Pei-Yu; Shih, Ping Yao; Yao, Chen-Wen

2002-01-01

Abstract Background Transgenic animals have become valuable tools for both research and applied purposes. The current method of gene transfer, microinjection, which is widely used in transgenic mouse production, has only had limited success in producing transgenic animals of larger or higher species. Here, we report a linker based sperm-mediated gene transfer method (LB-SMGT) that greatly improves the production efficiency of large transgenic animals. Results The linker protein, a monoclonal ...

Optimization of cationic lipid mediated gene transfer: structure-function, physico-chemical, and cellular studies.

Science.gov (United States)

Carrière, Marie; Tranchant, Isabelle; Niore, Pierre-Antoine; Byk, Gerardo; Mignet, Nathalie; Escriou, Virginie; Scherman, Daniel; Herscovici, Jean

2002-01-01

The rationale design aimed at the enhancement of cationic lipid mediated gene transfer is discussed. These improvements are based on the straight evaluation of the structure-activity relationship and on the introduction of new structures. Much attention have been given to the supramolecular structures of the lipid/DNA complexes, to the effect of serum on gene transfer and to the intracellular trafficking of the lipoplexes. Finally new avenue using reducible cationic lipids has been discussed.

Integration of replication-defective R68.45-like plasmids into the Pseudomonas aeruginosa chromosome.

Science.gov (United States)

Reimmann, C; Rella, M; Haas, D

1988-06-01

R68.45 and other similar broad-host-range (IncP) plasmids carrying a tandem repeat of the 2.1 kb insertion element IS21 mobilize the chromosome of many different Gram-negative bacteria. To analyse the structure of R68.45-chromosome cointegrates, whose involvement in the mobilization process had been postulated previously, we selected for the stable integration of R68.45-like plasmids into the Pseudomonas aeruginosa chromosome. Two plasmids were chosen: pME28, a transfer-deficient, mobilizable RP1 derivative with an inactive replication control (trfA) gene, and pME487, an R68.45 derivative with a trfA(ts) mutation causing temperature-sensitive replication. Chromosomally integrated pME28 and pME487 were found to be flanked by single IS21 elements. This structure is in agreement with a 'cut-and-paste' mode of R68.45 transposition. pME28 and pME487 showed a low specificity of insertion but rarely (less than 0.1%) induced auxotrophic mutations. Hfr (high-frequency-of-recombination) donors of P. aeruginosa could be obtained by chromosomal integration of pME487 or pME28; in the latter case, the transfer functions lacking from pME28 had to be provided in trans on an autonomous plasmid. Hfr donors gave higher conjugational linkage and transferred longer stretches of the P. aeruginosa chromosome than did R68.45 donors. This suggests that the integration of R68.45 into the donor chromosome is short-lived in P. aeruginosa.

Chromosome break points of T-lymphocytes from atomic bomb survivors

International Nuclear Information System (INIS)

Tanaka, Kimio; Kamada, Nanao; Kuramoto, Atsushi; Ohkita, Takeshi

1980-01-01

Chromosome break points of T-lymphocytes were investigated for 9 atomic bomb survivors estimated to be irradiated with 100 - 630 red. Chromosome aberration was found in 199 cells out of 678 cells investigated, with non-random distribution. The types of the chromosome aberration were, transfer: 56%, deficit: 38%, additional abnormality 3%, and reverse: 2%. High and low incidence of chromosome aberrations were observed at the chromosome numbers of 22, 21, and 13, and 11, 12, and 4, respectively. The aberration numbers per arm were high in 22q, 21q, and 18p and low in 11q, 5p, and 12q. For the distribution of aberration number within a chromosome, 50.7% was observed at the terminal portion and 73% was at the pale band appeared by Q-partial-stain method, suggesting the non-random distribution. The incidence of aberration number in 22q was statistically significant (P 1 chromosome in chronic myelocytic leukemia. The non-random distribution of chromosome break points seemed to reflect the selection effect since irradiation. (Nakanishi, T.)

Genome Organization Drives Chromosome Fragility.

Science.gov (United States)

Canela, Andres; Maman, Yaakov; Jung, Seolkyoung; Wong, Nancy; Callen, Elsa; Day, Amanda; Kieffer-Kwon, Kyong-Rim; Pekowska, Aleksandra; Zhang, Hongliang; Rao, Suhas S P; Huang, Su-Chen; Mckinnon, Peter J; Aplan, Peter D; Pommier, Yves; Aiden, Erez Lieberman; Casellas, Rafael; Nussenzweig, André

2017-07-27

In this study, we show that evolutionarily conserved chromosome loop anchors bound by CCCTC-binding factor (CTCF) and cohesin are vulnerable to DNA double strand breaks (DSBs) mediated by topoisomerase 2B (TOP2B). Polymorphisms in the genome that redistribute CTCF/cohesin occupancy rewire DNA cleavage sites to novel loop anchors. While transcription- and replication-coupled genomic rearrangements have been well documented, we demonstrate that DSBs formed at loop anchors are largely transcription-, replication-, and cell-type-independent. DSBs are continuously formed throughout interphase, are enriched on both sides of strong topological domain borders, and frequently occur at breakpoint clusters commonly translocated in cancer. Thus, loop anchors serve as fragile sites that generate DSBs and chromosomal rearrangements. VIDEO ABSTRACT. Published by Elsevier Inc.

Ionizing and ultraviolet radiation enhances the efficiency of DNA mediated gene transfer in vitro

International Nuclear Information System (INIS)

Perez, C.F.

1984-08-01

The enhancement effects of ionizing and non-ionizing radiation on the efficiency of DNA mediated gene transfer were studied. Confluent Rat-2 cells were transfected with purified SV40 viral DNA, irradiated with either X-rays or ultraviolet, trypsinized, plated, and assayed for the formation of foci on Rat-2 monolayers. Both ionizing and ultraviolet radiation enhanced the frequency of A-gene transformants/survivor compared to unirradiated transfected cells. These enhancements were non-linear and dose dependent. A recombinant plasmid, pOT-TK5, was constructed that contained the SV40 virus A-gene and the Herpes Simplex virus (HSV) thymidine kinase (TK) gene. Confluent Rat-2 cells transfected with pOT-TK5 DNA and then immediately irradiated with either X-rays or 330 MeV/amu argon particles at the Berkeley Bevalac showed a higher frequency of HAT + colonies/survivor than unirradiated transfected cells. Rat-2 cells transfected with the plasmid, pTK2, containing only the HSV TK-gene were enhanced for TK-transformation by both X-rays and ultraviolet radiation. The results demonstrate that radiation enhancement of the efficiency of DNA mediated gene transfer is not explained by increased nuclear uptake of the transfected DNA. Radiation increases the competence of the transfected cell population for genetic transformation. Three models for this increased competence are presented. The targeted integration model, the inducible recombination model, the partition model, and the utilization of DNA mediated gene transfer for DNA repair studies are discussed. 465 references

Ionizing and ultraviolet radiation enhances the efficiency of DNA mediated gene transfer in vitro

Energy Technology Data Exchange (ETDEWEB)

Perez, C.F.

1984-08-01

The enhancement effects of ionizing and non-ionizing radiation on the efficiency of DNA mediated gene transfer were studied. Confluent Rat-2 cells were transfected with purified SV40 viral DNA, irradiated with either X-rays or ultraviolet, trypsinized, plated, and assayed for the formation of foci on Rat-2 monolayers. Both ionizing and ultraviolet radiation enhanced the frequency of A-gene transformants/survivor compared to unirradiated transfected cells. These enhancements were non-linear and dose dependent. A recombinant plasmid, pOT-TK5, was constructed that contained the SV40 virus A-gene and the Herpes Simplex virus (HSV) thymidine kinase (TK) gene. Confluent Rat-2 cells transfected with pOT-TK5 DNA and then immediately irradiated with either X-rays or 330 MeV/amu argon particles at the Berkeley Bevalac showed a higher frequency of HAT/sup +/ colonies/survivor than unirradiated transfected cells. Rat-2 cells transfected with the plasmid, pTK2, containing only the HSV TK-gene were enhanced for TK-transformation by both X-rays and ultraviolet radiation. The results demonstrate that radiation enhancement of the efficiency of DNA mediated gene transfer is not explained by increased nuclear uptake of the transfected DNA. Radiation increases the competence of the transfected cell population for genetic transformation. Three models for this increased competence are presented. The targeted integration model, the inducible recombination model, the partition model, and the utilization of DNA mediated gene transfer for DNA repair studies are discussed. 465 references.

Chromosomal organization and segregation in Pseudomonas aeruginosa.

Directory of Open Access Journals (Sweden)

Isabelle Vallet-Gely

2013-05-01

Full Text Available The study of chromosomal organization and segregation in a handful of bacteria has revealed surprising variety in the mechanisms mediating such fundamental processes. In this study, we further emphasized this diversity by revealing an original organization of the Pseudomonas aeruginosa chromosome. We analyzed the localization of 20 chromosomal markers and several components of the replication machinery in this important opportunistic γ-proteobacteria pathogen. This technique allowed us to show that the 6.3 Mb unique circular chromosome of P. aeruginosa is globally oriented from the old pole of the cell to the division plane/new pole along the oriC-dif axis. The replication machinery is positioned at mid-cell, and the chromosomal loci from oriC to dif are moved sequentially to mid-cell prior to replication. The two chromosomal copies are subsequently segregated at their final subcellular destination in the two halves of the cell. We identified two regions in which markers localize at similar positions, suggesting a bias in the distribution of chromosomal regions in the cell. The first region encompasses 1.4 Mb surrounding oriC, where loci are positioned around the 0.2/0.8 relative cell length upon segregation. The second region contains at least 800 kb surrounding dif, where loci show an extensive colocalization step following replication. We also showed that disrupting the ParABS system is very detrimental in P. aeruginosa. Possible mechanisms responsible for the coordinated chromosomal segregation process and for the presence of large distinctive regions are discussed.

Electron transfer mediators accelerated the microbiologically influence corrosion against carbon steel by nitrate reducing Pseudomonas aeruginosa biofilm.

Science.gov (United States)

Jia, Ru; Yang, Dongqing; Xu, Dake; Gu, Tingyue

2017-12-01

Electron transfer is a rate-limiting step in microbiologically influenced corrosion (MIC) caused by microbes that utilize extracellular electrons. Cross-cell wall electron transfer is necessary to transport the electrons released from extracellular iron oxidation into the cytoplasm of cells. Electron transfer mediators were found to accelerate the MIC caused by sulfate reducing bacteria. However, there is no publication in the literature showing the effect of electron transfer mediators on MIC caused by nitrate reducing bacteria (NRB). This work demonstrated that the corrosion of anaerobic Pseudomonas aeruginosa (PAO1) grown as a nitrate reducing bacterium biofilm on C1018 carbon steel was enhanced by two electron transfer mediators, riboflavin and flavin adenine dinucleotide (FAD) separately during a 7-day incubation period. The addition of either 10ppm (w/w) (26.6μM) riboflavin or 10ppm (12.7μM) FAD did not increase planktonic cell counts, but they increased the maximum pit depth on carbon steel coupons considerably from 17.5μm to 24.4μm and 25.0μm, respectively. Riboflavin and FAD also increased the specific weight loss of carbon steel from 2.06mg/cm 2 to 2.34mg/cm 2 and 2.61mg/cm 2 , respectively. Linear polarization resistance, electrochemical impedance spectroscopy and potentiodynamic polarization curves all corroborated the pitting and weight loss data. Copyright © 2017 Elsevier B.V. All rights reserved.

Direct kinetochore-spindle pole connections are not required for chromosome segregation.

Science.gov (United States)

Sikirzhytski, Vitali; Magidson, Valentin; Steinman, Jonathan B; He, Jie; Le Berre, Maël; Tikhonenko, Irina; Ault, Jeffrey G; McEwen, Bruce F; Chen, James K; Sui, Haixin; Piel, Matthieu; Kapoor, Tarun M; Khodjakov, Alexey

2014-07-21

Segregation of genetic material occurs when chromosomes move to opposite spindle poles during mitosis. This movement depends on K-fibers, specialized microtubule (MT) bundles attached to the chromosomes' kinetochores. A long-standing assumption is that continuous K-fibers connect every kinetochore to a spindle pole and the force for chromosome movement is produced at the kinetochore and coupled with MT depolymerization. However, we found that chromosomes still maintained their position at the spindle equator during metaphase and segregated properly during anaphase when one of their K-fibers was severed near the kinetochore with a laser microbeam. We also found that, in normal fully assembled spindles, K-fibers of some chromosomes did not extend to the spindle pole. These K-fibers connected to adjacent K-fibers and/or nonkinetochore MTs. Poleward movement of chromosomes with short K-fibers was uncoupled from MT depolymerization at the kinetochore. Instead, these chromosomes moved by dynein-mediated transport of the entire K-fiber/kinetochore assembly. Thus, at least two distinct parallel mechanisms drive chromosome segregation in mammalian cells.

Mandatory chromosomal segment balance in aneuploid tumor cells

International Nuclear Information System (INIS)

Kost-Alimova, Maria; Stanbridge, Eric; Klein, George; Imreh, Stefan; Darai-Ramqvist, Eva; Yau, Wing Lung; Sandlund, Agneta; Fedorova, Ludmila; Yang, Ying; Kholodnyuk, Irina; Cheng, Yue; Li Lung, Maria

2007-01-01

Euploid chromosome balance is vitally important for normal development, but is profoundly changed in many tumors. Is each tumor dependent on its own structurally and numerically changed chromosome complement that has evolved during its development and progression? We have previously shown that normal chromosome 3 transfer into the KH39 renal cell carcinoma line and into the Hone1 nasopharyngeal carcinoma line inhibited their tumorigenicity. The aim of the present study was to distinguish between a qualitative and a quantitative model of this suppression. According to the former, a damaged or deleted tumor suppressor gene would be restored by the transfer of a normal chromosome. If so, suppression would be released only when the corresponding sequences of the exogenous normal chromosome are lost or inactivated. According to the alternative quantitative model, the tumor cell would not tolerate an increased dosage of the relevant gene or segment. If so, either a normal cell derived, or, a tumor derived endogenous segment could be lost. Fluorescence in Situ Hybridization based methods, as well as analysis of polymorphic microsatellite markers were used to follow chromosome 3 constitution changes in monochromosomal hybrids. In both tumor lines with introduced supernumerary chromosomes 3, the copy number of 3p21 or the entire 3p tended to fall back to the original level during both in vitro and in vivo growth. An exogenous, normal cell derived, or an endogenous, tumor derived, chromosome segment was lost with similar probability. Identification of the lost versus retained segments showed that the intolerance for increased copy number was particularly strong for 3p14-p21, and weaker for other 3p regions. Gains in copy number were, on the other hand, well tolerated in the long arm and particularly the 3q26-q27 region. The inability of the cell to tolerate an experimentally imposed gain in 3p14-p21 in contrast to the well tolerated gain in 3q26-q27 is consistent with the

Comparative genomics reveals mobile pathogenicity chromosomes in Fusarium

Energy Technology Data Exchange (ETDEWEB)

Ma, Li Jun; van der Does, H. C.; Borkovich, Katherine A.; Coleman, Jeffrey J.; Daboussi, Marie-Jose; Di Pietro, Antonio; Dufresne, Marie; Freitag, Michael; Grabherr, Manfred; Henrissat, Bernard; Houterman, Petra M.; Kang, Seogchan; Shim, Won-Bo; Wolochuk, Charles; Xie, Xiaohui; Xu, Jin Rong; Antoniw, John; Baker, Scott E.; Bluhm, Burton H.; Breakspear, Andrew; Brown, Daren W.; Butchko, Robert A.; Chapman, Sinead; Coulson, Richard; Coutinho, Pedro M.; Danchin, Etienne G.; Diener, Andrew; Gale, Liane R.; Gardiner, Donald; Goff, Steven; Hammond-Kossack, Kim; Hilburn, Karen; Hua-Van, Aurelie; Jonkers, Wilfried; Kazan, Kemal; Kodira, Chinnappa D.; Koehrsen, Michael; Kumar, Lokesh; Lee, Yong Hwan; Li, Liande; Manners, John M.; Miranda-Saavedra, Diego; Mukherjee, Mala; Park, Gyungsoon; Park, Jongsun; Park, Sook Young; Proctor, Robert H.; Regev, Aviv; Ruiz-Roldan, M. C.; Sain, Divya; Sakthikumar, Sharadha; Sykes, Sean; Schwartz, David C.; Turgeon, Barbara G.; Wapinski, Ilan; Yoder, Olen; Young, Sarah; Zeng, Qiandong; Zhou, Shiguo; Galagan, James; Cuomo, Christina A.; Kistler, H. Corby; Rep, Martijn

2010-03-18

Fusarium species are among the most important phytopathogenic and toxigenic fungi, having significant impact on crop production and animal health. Distinctively, members of the F. oxysporum species complex exhibit wide host range but discontinuously distributed host specificity, reflecting remarkable genetic adaptability. To understand the molecular underpinnings of diverse phenotypic traits and their evolution in Fusarium, we compared the genomes of three economically important and phylogenetically related, yet phenotypically diverse plant-pathogenic species, F. graminearum, F. verticillioides and F. oxysporum f. sp. lycopersici. Our analysis revealed greatly expanded lineage-specific (LS) genomic regions in F. oxysporum that include four entire chromosomes, accounting for more than one-quarter of the genome. LS regions are rich in transposons and genes with distinct evolutionary profiles but related to pathogenicity. Experimentally, we demonstrate for the first time the transfer of two LS chromosomes between strains of F. oxysporum, resulting in the conversion of a non-pathogenic strain into a pathogen. Transfer of LS chromosomes between otherwise genetically isolated strains explains the polyphyletic origin of host specificity and the emergence of new pathogenic lineages in the F. oxysporum species complex, putting the evolution of fungal pathogenicity into a new perspective.

Sterol transfer between cyclodextrin and membranes: similar but not identical mechanism to NPC2-mediated cholesterol transfer.

Science.gov (United States)

McCauliff, Leslie A; Xu, Zhi; Storch, Judith

2011-08-30

Niemann--Pick C disease is an inherited disorder in which cholesterol and other lipids accumulate in the late endosomal/lysosomal compartment. Recently, cyclodextrins (CD) have been shown to reduce symptoms and extend lifespan in animal models of the disease. In the present studies we examined the mechanism of sterol transport by CD using in vitro model systems and fluorescence spectroscopy and NPC2-deficient fibroblasts. We demonstrate that cholesterol transport from the lysosomal cholesterol-binding protein NPC2 to CD occurs via aqueous diffusional transfer and is very slow; the rate-limiting step appears to be dissociation of cholesterol from NPC2, suggesting that specific interactions between NPC2 and CD do not occur. In contrast, the transfer rate of the fluorescent cholesterol analogue dehydroergosterol (DHE) from CD to phospholipid membranes is very rapid and is directly proportional to the acceptor membrane concentration, as is DHE transfer from membranes to CD. Moreover, CD dramatically increases the rate of sterol transfer between membranes, with rates that can approach those mediated by NPC2. The results suggest that sterol transfer from CD to membranes occurs by a collisional transfer mechanism involving direct interaction of CD with membranes, similar to that shown previously for NPC2. For CD, however, absolute rates are slower compared to NPC2 for a given concentration, and the lysosomal phospholipid lysobisphosphatidic acid (LBPA) does not stimulate rates of sterol transfer between membranes and CD. As expected from the apparent absence of interaction between CD and NPC2, the addition of CD to NPC2-deficient fibroblasts rapidly rescued the cholesterol accumulation phenotype. Thus, the recent observations of CD efficacy in mouse models of NPC disease are likely the result of CD enhancement of cholesterol transport between membranes, with rapid sterol transfer occurring during CD--membrane interactions.

Transfer of Chinese hamster DNA repair gene(s) into repair-deficient human cells (Xeroderma pigmentosum)

International Nuclear Information System (INIS)

Karentz, D.; Cleaver, J.E.

1985-01-01

Transfer of repair genes by DNA transfection into repair-deficient Xeroderma pigmentosum (XP) cells has thus far been unsuccessful, presenting an obstacle to cloning XP genes. The authors chose an indirect route to transfer repair genes in chromosome fragments. DNA repair-competent (UV resistant) hybrid cell lines were established by PEG-mediated fusions of DNA repair-deficient (UV sensitive) human fibroblasts (XP12RO) with wild type Chinese hamster (CHO) cells (AA8). CHO cells were exposed to 5 Krad X-rays prior to fusions, predisposing hybrid cells to lose CHO chromosome fragments preferentially. Repair-competent hybrids were selected by periodic exposures to UV light. Secondary and tertiary hybrid cell lines were developed by fusion of X-irradiated hybrids to XP12RO. The hybrid cell lines exhibit resistance to UV that is comparable to that of CHO cells and they are proficient at repair replication after UV exposure. Whole cell DNA-DNA hybridizations indicate that the hybrids have greater homology to CHO DNA than is evident between XP12RO and CHO. These observations indicate that CHO DNA sequences which can function in repair of UV-damaged DNA in human cells have been transferred into the genome of the repair-deficient XP12RO cells

Ploidy of Bovine Nuclear Transfer Blastocysts Blastomere Donors

DEFF Research Database (Denmark)

Booth, P J; VIUFF, D; THOMSEN, P D

2000-01-01

The higher rate of embryonic loss in nuclear transfer compared to in vitro produced embryos may be due to chromosome abnormalities that occur during preimplantation in vitro devel- opment. Because little is known about ploidy errors in nuclear transfer embryos, this was ex- amined using embryos...... cultured until day 7 at which time blastocyst nuclei were extracted and chromosome abnormalities were evaluated by fluorescent in situ hybridization using two probes that bind to the subcentromeric regions on chromosomes 6 and 7. In 16 nuclear transfer blastocysts generated from 5 donor embryos, 53.8 6 20...... comprised mainly triploid (8.2 6 10.3 [0–26.3]: SD [range]) and tetraploid (10.6 6 19.9 [0–54.9]) nuclei with other ploidy com- binations accounting for only 0.9 6 2.1 [0–2.1]% of deviant nuclei. The proportion of com- pletely normal nuclear transfer embryos was no less than those produced by in vitro...

Direct electron transfer of glucose oxidase promoted by carbon nanotubes is without value in certain mediator-free applications

International Nuclear Information System (INIS)

Wang, Y.; Yao, Y.

2012-01-01

We have investigated the direct electron transfer (DET) promoted by carbon nanotubes (CNTs) on an electrode containing immobilized glucose oxidase (GOx) with the aim to develop a third-generation glucose biosensor and a mediator-free glucose biofuel cell anode. GOx was immobilized via chitosan (CS) on a glassy carbon electrode (GCE) modified with multi-walled carbon nanotubes (MWCNTs). Cyclic voltammetric revealed that the GOx on the surface of such an electrode is unable to simultaneously demonstrate DET with the electrode and to retain its catalytic activity towards glucose, although the MWCNTs alone can promote electron transfer between GOx and electrode. This is interpreted in terms of two types of GOx on the surface, the distribution and properties of which are quite different. The first type exhibits DET capability that results from the collaboration of MWCNTs and metal impurities, but is unable to catalyze the oxidation of glucose. The second type maintains its glucose-specific catalytic capability in the presence of a mediator, which can be enhanced by MWCNTs, but cannot undergo DET with the electrode. As a result, the MWCNTs are capable of promoting the electron transfer, but this is without value in some mediator-free applications such as in third-generation glucose biosensors and in mediator-free anodes for glucose biofuel cells. (author)

Effect of Phosphatase and Tensin Homologue on Chromosome 10 on Angiotensin II-Mediated Proliferation, Collagen Synthesis, and Akt/P27 Signaling in Neonatal Rat Cardiac Fibroblasts

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Ling Nie

2016-01-01

Full Text Available Cardiac fibroblasts (CFs play a key role in cardiac fibrosis by regulating the balance between extracellular matrix synthesis and breakdown. Although phosphatase and tensin homologue on chromosome 10 (PTEN has been found to play an important role in cardiovascular disease, it is not clear whether PTEN is involved in functional regulation of CFs. In the present study, PTEN was overexpressed in neonatal rat CFs via recombinant adenovirus-mediated gene transfer. The effects of PTEN overexpression on cell-cycle progression and angiotensin II- (Ang II- mediated regulation of collagen metabolism, synthesis of matrix metalloproteinases, and Akt/P27 signaling were investigated. Compared with uninfected cells and cells infected with green fluorescent protein-expressing adenovirus (Ad-GFP, cells infected with PTEN-expressing adenovirus (Ad-PTEN significantly increased PTEN protein and mRNA levels in CFs (P<0.05. The proportion of CFs in the G1/S cell-cycle phase was significantly higher for PTEN-overexpressing cells. In addition, Ad-PTEN decreased mRNA expression and the protein synthesis rate of collagen types I and III and antagonized Ang II-induced collagen synthesis. Overexpression of PTEN also decreased Ang II-induced matrix metalloproteinase-2 (MMP-2 and tissue inhibitor of metalloproteinase-1 (TIMP-1 production as well as gelatinase activity. Moreover, Ad-PTEN decreased Akt expression and increased P27 expression independent of Ang II stimulation. These results suggest that PTEN could regulate its functional effects in neonatal rat CFs partially via the Akt/P27 signaling pathway.

Direct kinetochore–spindle pole connections are not required for chromosome segregation

Science.gov (United States)

Sikirzhytski, Vitali; Magidson, Valentin; Steinman, Jonathan B.; He, Jie; Le Berre, Maël; Tikhonenko, Irina; Ault, Jeffrey G.; McEwen, Bruce F.; Chen, James K.; Sui, Haixin; Piel, Matthieu; Kapoor, Tarun M.

2014-01-01

Segregation of genetic material occurs when chromosomes move to opposite spindle poles during mitosis. This movement depends on K-fibers, specialized microtubule (MT) bundles attached to the chromosomes′ kinetochores. A long-standing assumption is that continuous K-fibers connect every kinetochore to a spindle pole and the force for chromosome movement is produced at the kinetochore and coupled with MT depolymerization. However, we found that chromosomes still maintained their position at the spindle equator during metaphase and segregated properly during anaphase when one of their K-fibers was severed near the kinetochore with a laser microbeam. We also found that, in normal fully assembled spindles, K-fibers of some chromosomes did not extend to the spindle pole. These K-fibers connected to adjacent K-fibers and/or nonkinetochore MTs. Poleward movement of chromosomes with short K-fibers was uncoupled from MT depolymerization at the kinetochore. Instead, these chromosomes moved by dynein-mediated transport of the entire K-fiber/kinetochore assembly. Thus, at least two distinct parallel mechanisms drive chromosome segregation in mammalian cells. PMID:25023516

Kinetics of water-mediated proton transfer in our atmosphere

International Nuclear Information System (INIS)

Loerting, T.

2000-07-01

Variational transition state theory and multidimensional tunneling methods on hybrid density functional theory generated hypersurfaces have been used to investigate the temperature dependence of the reaction rate constants of water-mediated proton transfer reactions relevant to the chemistry of our atmosphere, namely the hydration of sulfur dioxide and sulfur trioxide and the decomposition of chlorine nitrate. Highly accurate reaction barriers were calculated using ab initio methods taking into account most of the electron correlation, namely CCSD(T)/aug-cc-pVDZ//MP2/aug-cc-pVDZ and G2(MP2). On comparing the determined rate constants with laboratory and atmospheric data, the following points could be established: All of the investigated reactions are highly sensitive to changes in humidity, as water acts as efficient catalyst, i.e., the barrier to the reaction is reduced drastically. Present-day atmospheric chemistry can only be explained when a limited number of water molecules is available for the formation of molecular clusters. Both in the troposphere and in the stratosphere SO 3 is hydrated rather than SO 2 . SO 2 emissions have to be oxidized, therefore, before being subject to hydration. A mechanism involving two or three water molecules is relevant for the production of sulfate aerosols, which play a decisive role in the context of global climate change and acid rain. A third water molecule has the function of assisting double-proton transfer rather than acting as active participant in triple-proton transfer in the case of the hydration of sulfur oxides. The observed ozone depletion above Arctica and Antarctica can be explained either by decomposition of chlorine nitrate in the presence of three water molecules (triple proton transfer) or by decomposition of chlorine nitrate in the presence of one molecule of HCl and one molecule of water (double proton transfer). The preassociation reaction required for homogeneous gas-phase conversion of chlorine

Nuclear overhauser enhancement mediated chemical exchange saturation transfer imaging at 7 Tesla in glioblastoma patients.

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Daniel Paech

Full Text Available BACKGROUND AND PURPOSE: Nuclear Overhauser Enhancement (NOE mediated chemical exchange saturation transfer (CEST is a novel magnetic resonance imaging (MRI technique on the basis of saturation transfer between exchanging protons of tissue proteins and bulk water. The purpose of this study was to evaluate and compare the information provided by three dimensional NOE mediated CEST at 7 Tesla (7T and standard MRI in glioblastoma patients. PATIENTS AND METHODS: Twelve patients with newly diagnosed histologically proven glioblastoma were enrolled in this prospective ethics committee-approved study. NOE mediated CEST contrast was acquired with a modified three-dimensional gradient-echo sequence and asymmetry analysis was conducted at 3.3 ppm (B1 = 0.7 µT to calculate the magnetization transfer ratio asymmetry (MTR(asym. Contrast enhanced T1 (CE-T1 and T2-weighted images were acquired at 3T and used for data co-registration and comparison. RESULTS: Mean NOE mediated CEST signal based on MTR(asym values over all patients was significantly increased (p<0.001 in CE-T1 tumor (-1.99 ± 1.22%, tumor necrosis (-1.36 ± 1.30% and peritumoral CEST hyperintensities (PTCH within T2 edema margins (-3.56 ± 1.24% compared to contralateral normal appearing white matter (-8.38 ± 1.19%. In CE-T1 tumor (p = 0.015 and tumor necrosis (p<0.001 mean MTR(asym values were significantly higher than in PTCH. Extent of the surrounding tumor hyperintensity was smaller in eight out of 12 patients on CEST than on T2-weighted images, while four displayed at equal size. In all patients, isolated high intensity regions (0.40 ± 2.21% displayed on CEST within the CE-T1 tumor that were not discernible on CE-T1 or T2-weighted images. CONCLUSION: NOE mediated CEST Imaging at 7 T provides additional information on the structure of peritumoral hyperintensities in glioblastoma and displays isolated high intensity regions within the CE-T1 tumor that cannot be acquired on CE-T1 or T2

Development of a multiple-gene-loading method by combining multi-integration system-equipped mouse artificial chromosome vector and CRISPR-Cas9.

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Kazuhisa Honma

Full Text Available Mouse artificial chromosome (MAC vectors have several advantages as gene delivery vectors, such as stable and independent maintenance in host cells without integration, transferability from donor cells to recipient cells via microcell-mediated chromosome transfer (MMCT, and the potential for loading a megabase-sized DNA fragment. Previously, a MAC containing a multi-integrase platform (MI-MAC was developed to facilitate the transfer of multiple genes into desired cells. Although the MI system can theoretically hold five gene-loading vectors (GLVs, there are a limited number of drugs available for the selection of multiple-GLV integration. To overcome this issue, we attempted to knock out and reuse drug resistance genes (DRGs using the CRISPR-Cas9 system. In this study, we developed new methods for multiple-GLV integration. As a proof of concept, we introduced five GLVs in the MI-MAC by these methods, in which each GLV contained a gene encoding a fluorescent or luminescent protein (EGFP, mCherry, BFP, Eluc, and Cluc. Genes of interest (GOI on the MI-MAC were expressed stably and functionally without silencing in the host cells. Furthermore, the MI-MAC carrying five GLVs was transferred to other cells by MMCT, and the resultant recipient cells exhibited all five fluorescence/luminescence signals. Thus, the MI-MAC was successfully used as a multiple-GLV integration vector using the CRISPR-Cas9 system. The MI-MAC employing these methods may resolve bottlenecks in developing multiple-gene humanized models, multiple-gene monitoring models, disease models, reprogramming, and inducible gene expression systems.

Comparison of lentiviral and sleeping beauty mediated αβ T cell receptor gene transfer.

Directory of Open Access Journals (Sweden)

Anne-Christine Field

Full Text Available Transfer of tumour antigen-specific receptors to T cells requires efficient delivery and integration of transgenes, and currently most clinical studies are using gamma retroviral or lentiviral systems. Whilst important proof-of-principle data has been generated for both chimeric antigen receptors and αβ T cell receptors, the current platforms are costly, time-consuming and relatively inflexible. Alternative, more cost-effective, Sleeping Beauty transposon-based plasmid systems could offer a pathway to accelerated clinical testing of a more diverse repertoire of recombinant high affinity T cell receptors. Nucleofection of hyperactive SB100X transposase-mediated stable transposition of an optimised murine-human chimeric T cell receptor specific for Wilm's tumour antigen from a Sleeping Beauty transposon plasmid. Whilst transfer efficiency was lower than that mediated by lentiviral transduction, cells could be readily enriched and expanded, and mediated effective target cells lysis in vitro and in vivo. Integration sites of transposed TCR genes in primary T cells were almost randomly distributed, contrasting the predilection of lentiviral vectors for transcriptionally active sites. The results support exploitation of the Sleeping Beauty plasmid based system as a flexible and adaptable platform for accelerated, early-phase assessment of T cell receptor gene therapies.

A- or C-chromosomes, does it matter for the transfer of transgenes from ¤Brassica napus¤

DEFF Research Database (Denmark)

Tomiuk, J.; Hauser, T.P.; Bagger Jørgensen, Rikke

2000-01-01

of herbicide-tolerant plants was explained by selection against the C-chromosomes of B. napus in favor of the homeologous ii-chromosomes. Obviously, such C-chromosomes could be potential candidates as safe integration sites for transgenes. We considered these safety aspects using a simple population genetic...... model. Theory and experiments, however, do not favor the chromosomes of B. napus as safe candidates with respect to the introgression of transgenes into wild populations of B. rapa....

Exosome-mediated transfer of miR-10b promotes cell invasion in breast cancer.

Science.gov (United States)

Singh, Ramesh; Pochampally, Radhika; Watabe, Kounosuke; Lu, Zhaohui; Mo, Yin-Yuan

2014-11-26

Exosomes are 30-100 nm membrane vesicles of endocytic origin, mediating diverse biological functions including tumor cell invasion, cell-cell communication and antigen presentation through transfer of proteins, mRNAs and microRNAs. Recent evidence suggests that microRNAs can be released through ceramide-dependent secretory machinery regulated by neutral sphingomyelinase 2 (nSMase2) enzyme encoded by the smpd3 gene that triggers exosome secretion. However, whether exosome-mediated microRNA transfer plays any role in cell invasion remains poorly understood. Thus, the aim of this study was to identify the exosomal microRNAs involved in breast cancer invasion. The expression level of endogenous and exosomal miRNAs were examined by real time PCR and the expression level of target proteins were detected by western blot. Scanning electron and confocal microscopy were used to characterize exosomes and to study its uptake and transfer. Luciferase reporter plasmids and its mutant were used to confirm direct targeting. Furthermore, the functional significance of exosomal miR-10b was estimated by invasion assay. In this study, we demonstrate that microRNA carrying exosomes can be transferred among different cell lines through direct uptake. miR-10b is highly expressed in metastatic breast cancer MDA-MB-231 cells as compared to non-metastatic breast cancer cells or non-malignant breast cells; it is actively secreted into medium via exosomes. In particular, nSMase2 or ceramide promotes the exosome-mediated miR-10b secretion whereas ceramide inhibitor suppresses this secretion. Moreover, upon uptake, miR-10b can suppress the protein level of its target genes such as HOXD10 and KLF4, indicating its functional significance. Finally, treatment with exosomes derived from MDA-MB-231 cells could induce the invasion ability of non-malignant HMLE cells. Together, our results suggest that a set of specific microRNAs may play an important role in modulating tumor microenvironment through

Characterization of chromosome instability in interspecific somatic hybrids obtained by X-ray fusion between potato (Solanum tuberosum L.) and S. brevidens Phil

International Nuclear Information System (INIS)

Fehér, A.; Preiszner, J.; Litkey, Z.; Csanádi, G; Dudits, D.

1992-01-01

Asymmetric somatic hybrids between Solanum tuberosum L. and S. brevidens Phil. have been obtained via the fusion of protoplasts from potato leaves and from cell suspension culture of S. brevidens. The wild Solanum species served as donor after irradiation of its protoplasts with a lethal X-ray dose (200 Gy). Selection of the putative hybrids was based on the kanamycin-resistance marker gene previously introduced into the genome of Solanum brevidens by Agrobacterium-mediated gene transfer. Thirteen out of the 45 selected clones exhibited reduced morphogenic potential. The morphological abnormalities of the regenerated plantlets were gradually eliminated during the extended in vitro culture period. Cytological investigations revealed that the number of chromosomes in the cultured S. brevidens cells used as protoplast source ranged between 28-40 instead of the basic 2n=24 value. There was a high degree of aneuploidy in all of the investigated hybrid clones, and at least 12 extra chromosomes were observed in addition to the potato chromosomes (2n=48). Interand intraclonal variation and segregation during vegetative propagation indicated the genetic instability of the hybrids, which can be ascribed to the pre-existing and X-ray irradiation-induced chromosomal abnormalities in the donor S. brevidens cells. The detection of centromeric chromosome fragments and long, poly-constrictional chromosomes in cytological preparations as well as non-parental bands in Southern hybridizations with restriction fragment length polymorphism (RFLP) markers revealed extensive chromosome rearrangements in most of the regenerated clones. On the basis of the limited number of RFLP probes used, preferential loss of S. brevidens specific markers with a non-random elimination pattern could be detected in hybrid regenerants

Identification of embryonic chromosomal abnormality using FISH-based preimplantaion genetic diagnosis

Institute of Scientific and Technical Information of China (English)

叶英辉; 徐晨明; 金帆; 钱羽力

2004-01-01

Objective: Embryonic chromosomal abnormality is one of the main reasons for in vitro fertilization (IVF) failure. This study aimed at evaluating the value of Fluorescence in-situ Hybridization (FISH)-based Preimplantation Genetic Diagnosis (PGD) in screening for embryonic chromosomal abnormality to increase the successful rate of IVF. Method: Ten couples, four with high risk of chromosomal abnormality and six infertile couples, underwent FISH-based PGD during IVF procedure. At day 3, one or two blastomeres were aspirated from each embryo. Biopsied blastomeres were examined using FISH analysis to screen out embryos with chromosomal abnormalities. At day 4, embryos without detectable chromosomal abnormality were transferred to the mother bodies as in regular IVF. Results: Among 54 embryos screened using FISH-based PGD, 30 embryos were detected to have chromosomal abnormalities. The 24 healthy embryos were implanted, resulting in four clinical pregnancies, two of which led to successful normal birth of two healthy babies; one to ongoing pregnancy during the writing of this article; and one to ectopic pregnancy. Conclusion: FISH-based PGD is an effective method for detecting embryonic chromosomal abnormality, which is one of the common causes of spontaneous miscarriages and chromosomally unbalanced offsprings.

Identification of embryonic chromosomal abnormality using FISH-based preimplantaion genetic diagnosis

Institute of Scientific and Technical Information of China (English)

叶英辉; 徐晨明; 金帆; 钱羽力

2004-01-01

Objective: Embryonic chromosomal abnormality is one of the main reasons for in vitro fertilization (IVF)failure. This study aimed at evaluating the value of Fluorescence in-situ Hybridization (FISH)-based Preimplantation Genetic Diagnosis (PGD) in screening for embryonic chromosomal abnormality to increase the successful rate of IVF. Method:Ten couples, four with high risk of chromosomal abnormality and six infertile couples, underwent FISH-based PGD during IVF procedure. At day 3, one or two blastomeres were aspirated from each embryo. Biopsied blastomeres were examined using FISH analysis to screen out embryos with chromosomal abnormalities. At day 4, embryos without detectable chromosomal abnormality were transferred to the mother bodies as in regular IVF. Results: Among 54 embryos screened using FISH-based PGD, 30 embryos were detected to have chromosomal abnormalities. The 24 healthy embryos were implanted,resulting in four clinical pregnancies, two of which led to successful normal birth of two healthy babies; one to ongoing pregnancy during the writing of this article; and one to ectopic pregnancy. Conclusion: FISH-based PGD is an effective method for detecting embryonic chromosomal abnormality, which is one of the common causes of spontaneous miscarriages and chromosomally unbalanced offsprings.

Bridge mediated two-electron transfer reactions: Analysis of stepwise and concerted pathways

International Nuclear Information System (INIS)

Petrov, E.G.; May, V.

2004-01-01

A theory of nonadiabatic donor (D)-acceptor (A) two-electron transfer (TET) mediated by a single regular bridge (B) is developed. The presence of different intermediate two-electron states connecting the reactant state D -- BA with the product state DBA -- results in complex multiexponential kinetics. The conditions are discussed at which a reduction to two-exponential as well as single-exponential kinetics becomes possible. For the latter case the rate K TET is calculated, which describes the bridge-mediated reaction as an effective two-electron D-A transfer. In the limit of small populations of the intermediate TET states D - B - A, DB -- A, D - BA - , and DB - A - , K TET is obtained as a sum of the rates K TET (step) and K TET (sup) . The first rate describes stepwise TET originated by transitions of a single electron. It starts at D -- BA and reaches DBA -- via the intermediate state D - BA - . These transitions cover contributions from sequential as well as superexchange reactions all including reduced bridge states. In contrast, a specific two-electron superexchange mechanism from D -- BA to DBA -- defines K TET (sup) . An analytic dependence of K TET (step) and K TET (sup) on the number of bridging units is presented and different regimes of D-A TET are studied

Sequential actin-based pushing forces drive meiosis I chromosome migration and symmetry breaking in oocytes

Science.gov (United States)

Yi, Kexi; Rubinstein, Boris; Unruh, Jay R.; Guo, Fengli; Slaughter, Brian D.

2013-01-01

Polar body extrusion during oocyte maturation is critically dependent on asymmetric positioning of the meiotic spindle, which is established through migration of the meiosis I (MI) spindle/chromosomes from the oocyte interior to a subcortical location. In this study, we show that MI chromosome migration is biphasic and driven by consecutive actin-based pushing forces regulated by two actin nucleators, Fmn2, a formin family protein, and the Arp2/3 complex. Fmn2 was recruited to endoplasmic reticulum structures surrounding the MI spindle, where it nucleated actin filaments to initiate an initially slow and poorly directed motion of the spindle away from the cell center. A fast and highly directed second migration phase was driven by actin-mediated cytoplasmic streaming and occurred as the chromosomes reach a sufficient proximity to the cortex to activate the Arp2/3 complex. We propose that decisive symmetry breaking in mouse oocytes results from Fmn2-mediated perturbation of spindle position and the positive feedback loop between chromosome signal-induced Arp2/3 activation and Arp2/3-orchestrated cytoplasmic streaming that transports the chromosomes. PMID:23439682

Mapping of the human NMDA receptor subunit (NMDAR1) and the proposed NMDA receptor glutamate-binding subunit (NMDARA1) to chromosomes 9q34.3 and chromosome 8, respectively

DEFF Research Database (Denmark)

Collins, C; Duff, C; Duncan, A M

1993-01-01

to human chromosome 8 using a somatic cell hybrid panel. Because the gene causing HD has been localized to chromosome 4p16.3, the chromosome assignments reported here are inconsistent with either of these genes playing a causative role in the molecular pathology of HD. However, it is noteworthy......A role for the N-methyl-D-aspartate (NMDA) receptor in the molecular pathology underlying Huntington disease (HD) has been proposed on the basis of neurochemical studies in HD and the ability of the NMDA receptor to mediate neuronal cell death. The molecular cloning of the human NMDA receptor...

Dual effects of adenovirus-mediated thrombopoietin gene transfer on hepatic oval cell proliferation and platelet counts

International Nuclear Information System (INIS)

Ichiba, Miho; Shimomura, Takashi; Murai, Rie; Hashiguchi, Koichi; Saeki, Toshiya; Yoshida, Yoko; Kanbe, Takamasa; Tanabe, Naotada; Tsuchiya, Hiroyuki; Miura, Norimasa; Tajima, Fumihito; Kurimasa, Akihiro; Hamada, Hirofumi; Shiota, Goshi

2005-01-01

Thrombopoietin (TPO) is the growth factor for megakaryocytes and platelets, however, it also acts as a potent regulator of stem cell proliferation. To examine the significance of TPO expression in proliferation of hepatic oval cells, the effect of adenovirus-mediated TPO gene transfer into livers of the Solt-Farber model, which mimics the condition where liver regeneration is impaired, was examined. Hepatic TPO mRNA peaked its expression at 2 days after gene transduction and then gradually decreased. The peripheral platelet number began to increase at 4 days (P < 0.05) and reached its plateau at 9 days (P < 0.01). Oval cells expressed c-Mpl, a receptor for TPO as well as immature hematopoietic and hepatocytic surface markers such as CD34 and AFP. The proliferating cell nuclear antigen-positive oval cells in rats into which adenovirus-TPO gene was transferred at 7 and 9 days were significantly greater than those in adenovirus-LacZ gene transferred (P < 0.05, each), and the total numbers of oval cells in the adenovirus-TPO gene transferred at 9 and 13 days were also significantly greater than those in adenovirus-LacZ gene transferred (P < 0.05, each). Expression of SCF protein was increased at 4, 7, and 9 days by TPO gene administration and that of c-Kit was increased at 4 and 7 days. These data suggest that adenovirus-mediated TPO gene transfer stimulated oval cell proliferation in liver as well as increasing peripheral platelet counts, emphasizing the significance of the TPO/c-Mpl system in proliferation of hepatic oval cells

DNA-mediated gene transfer into ataxia-telangiectasia cells

International Nuclear Information System (INIS)

Crescenzi, M.; Pulciani, S.; Carbonari, M.; Tedesco, L.; Russo, G.; Gaetano, C.; Fiorilli, M.

1986-01-01

The complete description of the genetic lesion(s) underlying the AT mutation might, therefore, highlight not only a DNA-repair pathwa, but also an important aspect of the physiology of lymphocytes. DNA-mediated gene transfer into eukaryotic cells has proved a powerful tool for the molecular cloning of certain mammalian genes. The possibility to clone a given gene using this technology depends, basically, on the availability of a selectable marker associated with the expression of the transfected gene in the recipient cell. Recently, a human DNA repair gene has been cloned in CHO mutant cells by taking advantage of the increased resistance to ultraviolet radiation of the transformants. As a preliminary step toward the molecular cloning of the AT gene(s), the authors have attempted to confer radioresistance to AT cells by transfection with normal human DNA

Tolerance of DNA Mismatches in Dmc1 Recombinase-mediated DNA Strand Exchange*

Science.gov (United States)

Borgogno, María V.; Monti, Mariela R.; Zhao, Weixing; Sung, Patrick; Argaraña, Carlos E.; Pezza, Roberto J.

2016-01-01

Recombination between homologous chromosomes is required for the faithful meiotic segregation of chromosomes and leads to the generation of genetic diversity. The conserved meiosis-specific Dmc1 recombinase catalyzes homologous recombination triggered by DNA double strand breaks through the exchange of parental DNA sequences. Although providing an efficient rate of DNA strand exchange between polymorphic alleles, Dmc1 must also guard against recombination between divergent sequences. How DNA mismatches affect Dmc1-mediated DNA strand exchange is not understood. We have used fluorescence resonance energy transfer to study the mechanism of Dmc1-mediated strand exchange between DNA oligonucleotides with different degrees of heterology. The efficiency of strand exchange is highly sensitive to the location, type, and distribution of mismatches. Mismatches near the 3′ end of the initiating DNA strand have a small effect, whereas most mismatches near the 5′ end impede strand exchange dramatically. The Hop2-Mnd1 protein complex stimulates Dmc1-catalyzed strand exchange on homologous DNA or containing a single mismatch. We observed that Dmc1 can reject divergent DNA sequences while bypassing a few mismatches in the DNA sequence. Our findings have important implications in understanding meiotic recombination. First, Dmc1 acts as an initial barrier for heterologous recombination, with the mismatch repair system providing a second level of proofreading, to ensure that ectopic sequences are not recombined. Second, Dmc1 stepping over infrequent mismatches is likely critical for allowing recombination between the polymorphic sequences of homologous chromosomes, thus contributing to gene conversion and genetic diversity. PMID:26709229

Chromosome

Science.gov (United States)

... St Louis, MO: Elsevier; 2017:chap 69. Taber's Medical Dictionary Online. Chromosome. www.tabers.com/tabersonline/view/Tabers-Dictionary/753321/all/chromosome?q=Chromosome&ti=0 . Accessed June 11, 2017.

Bacteriophage Mediates Efficient Gene Transfer in Combination with Conventional Transfection Reagents.

Science.gov (United States)

Donnelly, Amanda; Yata, Teerapong; Bentayebi, Kaoutar; Suwan, Keittisak; Hajitou, Amin

2015-12-08

The development of commercially available transfection reagents for gene transfer applications has revolutionized the field of molecular biology and scientific research. However, the challenge remains in ensuring that they are efficient, safe, reproducible and cost effective. Bacteriophage (phage)-based viral vectors have the potential to be utilized for general gene transfer applications within research and industry. Yet, they require adaptations in order to enable them to efficiently enter cells and overcome mammalian cellular barriers, as they infect bacteria only; furthermore, limited progress has been made at increasing their efficiency. The production of a novel hybrid nanocomplex system consisting of two different nanomaterial systems, phage vectors and conventional transfection reagents, could overcome these limitations. Here we demonstrate that the combination of cationic lipids, cationic polymers or calcium phosphate with M13 bacteriophage-derived vectors, engineered to carry a mammalian transgene cassette, resulted in increased cellular attachment, entry and improved transgene expression in human cells. Moreover, addition of a targeting ligand into the nanocomplex system, through genetic engineering of the phage capsid further increased gene expression and was effective in a stable cell line generation application. Overall, this new hybrid nanocomplex system (i) provides enhanced phage-mediated gene transfer; (ii) is applicable for laboratory transfection processes and (iii) shows promise within industry for large-scale gene transfer applications.

The functional role for condensin in the regulation of chromosomal organization during the cell cycle.

Science.gov (United States)

Kagami, Yuya; Yoshida, Kiyotsugu

2016-12-01

In all organisms, the control of cell cycle progression is a fundamental process that is essential for cell growth, development, and survival. Through each cell cycle phase, the regulation of chromatin organization is essential for natural cell proliferation and maintaining cellular homeostasis. During mitosis, the chromatin morphology is dramatically changed to have a "thread-like" shape and the condensed chromosomes are segregated equally into two daughter cells. Disruption of the mitotic chromosome architecture physically impedes chromosomal behaviors, such as chromosome alignment and chromosome segregation; therefore, the proper mitotic chromosome structure is required to maintain chromosomal stability. Accumulating evidence has demonstrated that mitotic chromosome condensation is induced by condensin complexes. Moreover, recent studies have shown that condensin also modulates interphase chromatin and regulates gene expression. This review mainly focuses on the molecular mechanisms that condensin uses to exert its functions during the cell cycle progression. Moreover, we discuss the condensin-mediated chromosomal organization in cancer cells.

Bloom syndrome and maternal uniparental disomy for chromosome 15

Energy Technology Data Exchange (ETDEWEB)

Woodage, T.; Prasad, M.; Trent, R.J.; Smith, A. (Children' s Hospital, Camperdown, New South Wales (New Zealand)); Dixon, J.W.; Romain, D.R.; Columbano-Green, L.M.; Selby, R.E. (Wellington Hospital (New Zealand)); Graham, D. (Waikato Hospital, Hamilton (New Zealand)); Rogan, P.K. (Pennsylvania State Univ., Hershey, PA (United States)) (and others)

1994-07-01

Bloom syndrome (BS) is an autosomal recessive disorder characterized by increases in the frequency of sister-chromatid exchange and in the incidence of malignancy. Chromosome-transfer studies have shown the BS locus to map to chromosome 15q. This report describes a subject with features of both BS and Prader-Willi syndrome (PWS). Molecular analysis showed maternal uniparental disomy for chromosome 15. Meiotic recombination between the two disomic chromosomes 15 has resulted in heterodisomy for proximal 15q and isodisomy for distal 15q. In this individual BS is probably due to homozygosity for a gene that is telomeric to D15S95 (15q25), rather than to genetic imprinting, the mechanism responsible for the development of PWS. This report represents the first application of disomy analysis to the regional localization of a disease gene. This strategy promises to be useful in the genetic mapping of other uncommon autosomal recessive conditions. 37 refs., 3 figs., 2 tabs.

Charge and energy transfer interplay in hybrid sensitized solar cells mediated by graphene quantum dots

International Nuclear Information System (INIS)

Mihalache, Iuliana; Radoi, Antonio; Mihaila, Mihai; Munteanu, Cornel; Marin, Alexandru; Danila, Mihai; Kusko, Mihaela; Kusko, Cristian

2015-01-01

Highlights: • We report a one pot synthesis metod of GQD with controlled size and optoelectronic properties. • An improvement of common N3-DSSC characteristics is achieved when GQDs are used as co-sensitiser. • The role of GQD as cosensitisers in hybrid DSSC was investigated and the interplay between charge and energy transfer phenomena mediated by GQDs was demonstrated. • The GQDs presence determines an inhibition of the recombination processes at the TiO 2 /electrolyte interface. - Abstract: We explored the role of graphene quantum dots (GQDs) as co-sensitizers in hybrid dye sensitized solar cell (DSSC) architectures, focusing on various concurring mechanisms, such as: charge transfer, energy transfer and recombination rate, towards light harvesting improvement. GQDs were prepared by the hydrothermal method that allows the tuning of electronic levels and optical properties by employing appropriate precursors and synthesis conditions. The aim was to realize a type II alignment for TiO 2 /GQD/dye hybrid configuration, using standard N3 Ru-dye in order to improve charge transfer. When GQDs were used as co-sensitizers together with N3 Ru-dye, an improvement in power conversion efficiency was achieved, as shown by electrical measurements. The experimental analysis indicates that this improvement arises from the interplay of various mechanisms mediated by GQDs: (i) enhancement of charge separation and collection due to the cascaded alignment of the energy levels; (ii) energy transfer from GQDs to N3 Ru-dye due to the overlap between GQD photoluminescence and N3 Ru-dye absorption spectra; and (iii) reduction of the electron recombination to the redox couple due to the inhibition of the back electron transfer to the electrolyte by the GQDs

Type 3 Fimbriae Encoded on Plasmids Are Expressed from a Unique Promoter without Affecting Host Motility, Facilitating an Exceptional Phenotype That Enhances Conjugal Plasmid Transfer

DEFF Research Database (Denmark)

Madsen, Jonas Stenlokke; Riber, Leise; Kot, Witold

2016-01-01

Horizontal gene transfer (HGT), the transmission of genetic material to a recipient that is not the progeny of the donor, is fundamental in bacterial evolution. HGT is often mediated by mobile genetic elements such as conjugative plasmids, which may be in conflict with the chromosomal elements...... of the genome because they are independent replicons that may petition their own evolutionary strategy. Here we study differences between type 3 fimbriae encoded on wild type plasmids and in chromosomes. Using known and newly characterized plasmids we show that the expression of type 3 fimbriae encoded...... on plasmids is systematically different, as MrkH, a c-di-GMP dependent transcriptional activator is not needed for strong expression of the fimbriae. MrkH is required for expression of type 3 fimbriae of the Klebsiella pneumoniae chromosome, wherefrom the fimbriae operon (mrkABCDF) of plasmids is believed...

Syntrophic Growth via Quinone-Mediated Interspecies Electron Transfer

Directory of Open Access Journals (Sweden)

Jessica A Smith

2015-02-01

Full Text Available The mechanisms by which microbial species exchange electrons are of interest because interspecies electron transfer can expand the metabolic capabilities of microbial communities. Previous studies with the humic substance analog anthraquinone-2,6-disulfonate (AQDS suggested that quinone-mediated interspecies electron transfer (QUIET is feasible, but it was not determined if sufficient energy is available from QUIET to support the growth of both species. Furthermore, there have been no previous studies on the mechanisms for the oxidation of anthrahydroquinone-2,6-disulfonate (AHQDS. A co-culture of Geobacter metallireducens and Geobacter sulfurreducens metabolized ethanol with the reduction of fumarate much faster in the presence of AQDS, and there was an increase in cell protein. G. sulfurreducens was more abundant, consistent with G. sulfurreducens obtaining electrons from acetate that G. metallireducens produced from ethanol, as well as from AHQDS. Cocultures initiated with a citrate synthase-deficient strain of G. sulfurreducens that was unable to use acetate as an electron donor also metabolized ethanol with the reduction of fumarate and cell growth, but acetate accumulated over time. G. sulfurreducens and G. metallireducens were equally abundant in these co-cultures reflecting the inability of the citrate synthase-deficient strain of G. sulfurreducens to metabolize acetate. Evaluation of the mechanisms by which G. sulfurreducens accepts electrons from AHQDS demonstrated that a strain deficient in outer-surface c-type cytochromes that are required for AQDS reduction was as effective at QUIET as the wild-type strain. Deletion of additional genes previously implicated in extracellular electron transfer also had no impact on QUIET. These results demonstrate that QUIET can yield sufficient energy to support the growth of both syntrophic partners, but that the mechanisms by which electrons are derived from extracellular hydroquinones require

RecET driven chromosomal gene targeting to generate a RecA deficient Escherichia coli strain for Cre mediated production of minicircle DNA

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Coutelle Charles

2006-03-01

Full Text Available Abstract Background Minicircle DNA is the non-replicating product of intramolecular site-specific recombination within a bacterial minicircle producer plasmid. Minicircle DNA can be engineered to contain predominantly human sequences which have a low content of CpG dinucleotides and thus reduced immunotoxicity for humans, whilst the immunogenic bacterial origin and antibiotic resistance marker gene sequences are entirely removed by site-specific recombination. This property makes minicircle DNA an excellent vector for non-viral gene therapy. Large-scale production of minicircle DNA requires a bacterial strain expressing tightly controlled site-specific recombinase, such as Cre recombinase. As recombinant plasmids tend to be more stable in RecA-deficient strains, we aimed to construct a recA- bacterial strain for generation of minicircle vector DNA with less chance of unwanted deletions. Results We describe here the construction of the RecA-deficient minicircle DNA producer Escherichia coli HB101Cre with a chromosomally located Cre recombinase gene under the tight control of the araC regulon. The Cre gene expression cassette was inserted into the chromosomal lacZ gene by creating transient homologous recombination proficiency in the recA- strain HB101 using plasmid-born recET genes and homology-mediated chromosomal "pop-in, pop-out" of the plasmid pBAD75Cre containing the Cre gene and a temperature sensitive replication origin. Favourably for the Cre gene placement, at the "pop-out" step, the observed frequency of RecET-led recombination between the proximal regions of homology was 10 times higher than between the distal regions. Using the minicircle producing plasmid pFIXluc containing mutant loxP66 and loxP71 sites, we isolated pure minicircle DNA from the obtained recA- producer strain HB101Cre. The minicircle DNA preparation consisted of monomeric and, unexpectedly, also multimeric minicircle DNA forms, all containing the hybrid loxP66

Dexter energy transfer pathways.

Science.gov (United States)

Skourtis, Spiros S; Liu, Chaoren; Antoniou, Panayiotis; Virshup, Aaron M; Beratan, David N

2016-07-19

Energy transfer with an associated spin change of the donor and acceptor, Dexter energy transfer, is critically important in solar energy harvesting assemblies, damage protection schemes of photobiology, and organometallic opto-electronic materials. Dexter transfer between chemically linked donors and acceptors is bridge mediated, presenting an enticing analogy with bridge-mediated electron and hole transfer. However, Dexter coupling pathways must convey both an electron and a hole from donor to acceptor, and this adds considerable richness to the mediation process. We dissect the bridge-mediated Dexter coupling mechanisms and formulate a theory for triplet energy transfer coupling pathways. Virtual donor-acceptor charge-transfer exciton intermediates dominate at shorter distances or higher tunneling energy gaps, whereas virtual intermediates with an electron and a hole both on the bridge (virtual bridge excitons) dominate for longer distances or lower energy gaps. The effects of virtual bridge excitons were neglected in earlier treatments. The two-particle pathway framework developed here shows how Dexter energy-transfer rates depend on donor, bridge, and acceptor energetics, as well as on orbital symmetry and quantum interference among pathways.

Exploration of new perspectives and limitations in Agrobacterium mediated gene transfer technology. Progress report, [June 1, 1992-- May 31, 1994

Energy Technology Data Exchange (ETDEWEB)

Marton, L.

1994-12-31

This report describes progress aimed at constructing gene-transfer technology for Nicotiana plumbaginifolia. Most actual effort as described herein has so far been directed at exploring new perspectives and limitations in Agrobacterium mediated gene transfer. Accomplishments are described using a core homologous gene targeting vector.

Construction of an Unstable Ring-X Chromosome Bearing the Autosomal Dopa Decarboxylase Gene in Drosophila melanogaster and Analysis of Ddc Mosaics

OpenAIRE

Gailey, Donald A.; Bordne, Deborah L.; Vallés, Ana Maria; Hall, Jeffrey C.; White, Kalpana

1987-01-01

An unstable Ring-X chromosome, Ddc+- Ring-X carrying a cloned Dopa decarboxylase (Ddc) encoding segment was constructed. The construction involved a double recombination event between the unstable Ring-X, R(1)wvC and a Rod-X chromosome which contained a P-element mediated Ddc + insert. The resulting Ddc+-Ring-X chromosome behaves similarly to the parent chromosome with respect to somatic instability. The Ddc+-Ring-X chromosome was used to generate Ddc mosaics. Analyses of Ddc mosaics reveal...

Electron Transfer Mediator Effects in Water Oxidation Catalysis by Solution and Surface-Bound Ruthenium Bpy-Dicarboxylate Complexes

Energy Technology Data Exchange (ETDEWEB)

Sheridan, Matthew V.; Sherman, Benjamin D.; Marquard, Seth L.; Fang, Zhen; Ashford, Dennis L.; Wee, Kyung-Ryang; Gold, Alexander S.; Alibabaei, Leila; Rudd, Jennifer A.; Coggins, Michael K.; Meyer, Thomas J.

2015-11-12

Electrocatalytic water oxidation by the catalyst, ruthenium 2,2'-bipyridine-6,6'-dicarboxylate (bda) bis-isoquinoline (isoq), [Ru(bda)(isoq)₂], 1, was investigated at metal oxide electrodes surface-derivatized with electron transfer (ET) mediators. At indium-doped tin oxide (ITO) in pH 7.2 in H₂PO₄–/HPO₄²– buffers in 0.5 M NaClO₄ with added acetonitrile (MeCN), the catalytic activity of 1 is enhanced by the surface-bound redox mediators [Ru (4,4'-PO₃H₂-bpy)(4,4'-R-bpy)₂]²⁺ (RuPbpyR₂²⁺, R = Br, H, Me, or OMe, bpy = 2,2'-bipyridine). Rate-limiting ET between the Ru³⁺ form of the mediator and the Ru^IV(O) form in the [Ru^V/IV(O)]^+/0 couple of 1 is observed at relatively high concentrations of HPO₄^2– buffer base under conditions where O···O bond formation is facilitated by atom-proton transfer (APT). For the solution [Ru(bpy)₃]^3+/2+ mediator couple and 1 as the catalyst, catalytic currents vary systematically with the concentration of mediator and the HPO₄^2– buffer base concentration. Electron transfer mediation of water oxidation catalysis was also investigated on nanoparticle TiO₂ electrodes co-loaded with catalyst [Ru(bda)(py-4-O(CH₂)₃-PO₃H₂)₂], 2, (py = pyridine) and RuPbpyR₂²⁺ (R = H, Me, or OMe) with an interplay between rate-limiting catalyst oxidation and rate-limiting O···O bond formation by APT. Lastly, the co-loaded assembly RuPbpyR₂²⁺ + 2 has been investigated in a dye-sensitized photoelectrosynthesis cell for water splitting.

Tolerance of DNA Mismatches in Dmc1 Recombinase-mediated DNA Strand Exchange.

Science.gov (United States)

Borgogno, María V; Monti, Mariela R; Zhao, Weixing; Sung, Patrick; Argaraña, Carlos E; Pezza, Roberto J

2016-03-04

Recombination between homologous chromosomes is required for the faithful meiotic segregation of chromosomes and leads to the generation of genetic diversity. The conserved meiosis-specific Dmc1 recombinase catalyzes homologous recombination triggered by DNA double strand breaks through the exchange of parental DNA sequences. Although providing an efficient rate of DNA strand exchange between polymorphic alleles, Dmc1 must also guard against recombination between divergent sequences. How DNA mismatches affect Dmc1-mediated DNA strand exchange is not understood. We have used fluorescence resonance energy transfer to study the mechanism of Dmc1-mediated strand exchange between DNA oligonucleotides with different degrees of heterology. The efficiency of strand exchange is highly sensitive to the location, type, and distribution of mismatches. Mismatches near the 3' end of the initiating DNA strand have a small effect, whereas most mismatches near the 5' end impede strand exchange dramatically. The Hop2-Mnd1 protein complex stimulates Dmc1-catalyzed strand exchange on homologous DNA or containing a single mismatch. We observed that Dmc1 can reject divergent DNA sequences while bypassing a few mismatches in the DNA sequence. Our findings have important implications in understanding meiotic recombination. First, Dmc1 acts as an initial barrier for heterologous recombination, with the mismatch repair system providing a second level of proofreading, to ensure that ectopic sequences are not recombined. Second, Dmc1 stepping over infrequent mismatches is likely critical for allowing recombination between the polymorphic sequences of homologous chromosomes, thus contributing to gene conversion and genetic diversity. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Loading of PAX3 to Mitotic Chromosomes Is Mediated by Arginine Methylation and Associated with Waardenburg Syndrome*

Science.gov (United States)

Wu, Tsu-Fang; Yao, Ya-Li; Lai, I-Lu; Lai, Chien-Chen; Lin, Pei-Lun; Yang, Wen-Ming

2015-01-01

PAX3 is a transcription factor critical to gene regulation in mammalian development. Mutations in PAX3 are associated with Waardenburg syndrome (WS), but the mechanism of how mutant PAX3 proteins cause WS remains unclear. Here, we found that PAX3 loads on mitotic chromosomes using its homeodomain. PAX3 WS mutants with mutations in homeodomain lose the ability to bind mitotic chromosomes. Moreover, loading of PAX3 on mitotic chromosomes requires arginine methylation, which is regulated by methyltransferase PRMT5 and demethylase JMJD6. Mutant PAX3 proteins that lose mitotic chromosome localization block cell proliferation and normal development of zebrafish. These results reveal the molecular mechanism of PAX3s loading on mitotic chromosomes and the importance of this localization pattern in normal development. Our findings suggest that PAX3 WS mutants interfere with the normal functions of PAX3 in a dominant negative manner, which is important to the understanding of the pathogenesis of Waardenburg syndrome. PMID:26149688

Computational model of dose response for low-LET-induced complex chromosomal aberrations

International Nuclear Information System (INIS)

Eidelman, Y.A.; Andreev, S.G.

2015-01-01

Experiments with full-colour mFISH chromosome painting have revealed high yield of radiation-induced complex chromosomal aberrations (CAs). The ratio of complex to simple aberrations is dependent on cell type and linear energy transfer. Theoretical analysis has demonstrated that the mechanism of CA formation as a result of interaction between lesions at a surface of chromosome territories does not explain high complexes-to-simples ratio in human lymphocytes. The possible origin of high yields of γ-induced complex CAs was investigated in the present work by computer simulation. CAs were studied on the basis of chromosome structure and dynamics modelling and the hypothesis of CA formation on nuclear centres. The spatial organisation of all chromosomes in a human interphase nucleus was predicted by simulation of mitosis-to-interphase chromosome structure transition. Two scenarios of CA formation were analysed, 'static' (existing in a nucleus prior to irradiation) centres and 'dynamic' (formed in response to irradiation) centres. The modelling results reveal that under certain conditions, both scenarios explain quantitatively the dose-response relationships for both simple and complex γ-induced inter-chromosomal exchanges observed by mFISH chromosome painting in the first post-irradiation mitosis in human lymphocytes. (authors)

Construction of a complete set of alien chromosome addition lines from Gossypium australe in Gossypium hirsutum: morphological, cytological, and genotypic characterization.

Science.gov (United States)

Chen, Yu; Wang, Yingying; Wang, Kai; Zhu, Xiefei; Guo, Wangzhen; Zhang, Tianzhen; Zhou, Baoliang

2014-05-01

We report the first complete set of alien addition lines of G. hirsutum . The characterized lines can be used to introduce valuable traits from G. australe into cultivated cotton. Gossypium australe is a diploid wild cotton species (2n = 26, GG) native to Australia that possesses valuable characteristics unavailable in the cultivated cotton gene pool, such as delayed pigment gland morphogenesis in the seed and resistances to pests and diseases. However, it is very difficult to directly transfer favorable traits into cultivated cotton through conventional gene recombination due to the absence of pairing and crossover between chromosomes of G. australe and Gossypium hirsutum (2n = 52, AADD). To enhance the transfer of favorable genes from wild species into cultivated cotton, we developed a set of hirsutum-australe monosomic alien chromosome addition lines (MAAL) using a combination of morphological survey, microsatellite marker-assisted selection, and molecular cytogenetic analysis. The amphidiploid (2n = 78, AADDGG) of G. australe and G. hirsutum was consecutively backcrossed with upland cotton to develop alien addition lines of individual G. australe chromosomes in G. hirsutum. From these backcross progeny, we generated the first complete set of chromosome addition lines in cotton; 11 of 13 lines are monosomic additions, and chromosomes 7G(a) and 13G(a) are multiple additions. MAALs of 1G(a) and 11G(a) were the first to be isolated. The chromosome addition lines can be employed as bridges for the transfer of desired genes from G. australe into G. hirsutum, as well as for gene assignment, isolation of chromosome-specific probes, flow sorting and microdissection of chromosome, development of chromosome-specific ''paints'' for fluorochrome-labeled DNA fragments, physical mapping, and selective isolation and mapping of cDNAs for a particular G. australe chromosome.

Bacterial Artificial Chromosome Mutagenesis Using Recombineering

Directory of Open Access Journals (Sweden)

Kumaran Narayanan

2011-01-01

Full Text Available Gene expression from bacterial artificial chromosome (BAC clones has been demonstrated to facilitate physiologically relevant levels compared to viral and nonviral cDNA vectors. BACs are large enough to transfer intact genes in their native chromosomal setting together with flanking regulatory elements to provide all the signals for correct spatiotemporal gene expression. Until recently, the use of BACs for functional studies has been limited because their large size has inherently presented a major obstacle for introducing modifications using conventional genetic engineering strategies. The development of in vivo homologous recombination strategies based on recombineering in E. coli has helped resolve this problem by enabling facile engineering of high molecular weight BAC DNA without dependence on suitably placed restriction enzymes or cloning steps. These techniques have considerably expanded the possibilities for studying functional genetics using BACs in vitro and in vivo.

Csm4, in collaboration with Ndj1, mediates telomere-led chromosome dynamics and recombination during yeast meiosis.

Directory of Open Access Journals (Sweden)

Jennifer J Wanat

2008-09-01

Full Text Available Chromosome movements are a general feature of mid-prophase of meiosis. In budding yeast, meiotic chromosomes exhibit dynamic movements, led by nuclear envelope (NE-associated telomeres, throughout the zygotene and pachytene stages. Zygotene motion underlies the global tendency for colocalization of NE-associated chromosome ends in a "bouquet." In this study, we identify Csm4 as a new molecular participant in these processes and show that, unlike the two previously identified components, Ndj1 and Mps3, Csm4 is not required for meiosis-specific telomere/NE association. Instead, it acts to couple telomere/NE ensembles to a force generation mechanism. Mutants lacking Csm4 and/or Ndj1 display the following closely related phenotypes: (i elevated crossover (CO frequencies and decreased CO interference without abrogation of normal pathways; (ii delayed progression of recombination, and recombination-coupled chromosome morphogenesis, with resulting delays in the MI division; and (iii nondisjunction of homologs at the MI division for some reason other than absence of (the obligatory CO(s. The recombination effects are discussed in the context of a model where the underlying defect is chromosome movement, the absence of which results in persistence of inappropriate chromosome relationships that, in turn, results in the observed mutant phenotypes.

Production of alien chromosome additions and their utility in plant genetics

NARCIS (Netherlands)

Chang, S.B.; Jong, de J.H.S.G.M.

2005-01-01

Breeding programs aiming at transferring desirable genes from one species to another through interspecific hybridization and backcrossings often produce monosomic and disomic additions as intermediate crossing products. Such aneuploids contain alien chromosomes added to the complements of the

Gametocidal chromosomes enhancing chromosome aberration in common wheat induced by 5-azacytidine.

Science.gov (United States)

Su, W-Y; Cong, W-W; Shu, Y-J; Wang, D; Xu, G-H; Guo, C-H

2013-07-08

The gametocidal (Gc) chromosome from Aegilops spp induces chromosome mutation, which is introduced into common wheat as a tool of chromosome manipulation for genetic improvement. The Gc chromosome functions similar to a restriction-modification system in bacteria, in which DNA methylation is an important regulator. We treated root tips of wheat carrying Gc chromosomes with the hypomethylation agent 5-azacytidine; chromosome breakage and micronuclei were observed in these root tips. The frequency of aberrations differed in wheat containing different Gc chromosomes, suggesting different functions inducing chromosome breakage. Gc chromosome 3C caused the greatest degree of chromosome aberration, while Gc chromosome 3C(SAT) and 2C caused only slight chromosome aberration. Gc chromosome 3C induced different degrees of chromosome aberration in wheat varieties Triticum aestivum var. Chinese Spring and Norin 26, demonstrating an inhibition function in common wheat.

Rf8-Mediated T-urf13 Transcript Accumulation Coincides with a Pentatricopeptide Repeat Cluster on Maize Chromosome 2L

Directory of Open Access Journals (Sweden)

Julie Meyer

2011-11-01

Full Text Available Cytoplasmic male sterility (CMS is a maternally inherited inability to produce functional pollen. In Texas (T-cytoplasm maize ( L., CMS results from the action of the URF13 mitochondrial pore-forming protein encoded by the unique T- mitochondrial gene. Full or partial restoration of fertility to T-cytoplasm maize is mediated by the nuclear gene in combination with one of three other genes: , , or *. encodes a mitochondrial aldehyde dehydrogenase whereas , , and * are associated with the accumulation of distinctive T- mitochondrial transcripts. -associated RNA processing activity was mapped to a 4.55-Mbp region on chromosome 2L that contains 10 pentatricopeptide repeat (PPR encoding genes in the B73 5b.60 genome assembly. Genetic linkage analysis also indicated that * is positioned within this PPR cluster as well as , which restores USDA (S-cytoplasm maize. Partially male-fertile plants segregated for the presence or absence of the -associated T- 1.42- and 0.42-kbp transcripts, indicating that the RNA processing event associated with these transcripts is not necessary for anther exsertion. In addition, a statistically significant delay in flowering was observed between partially male-fertile and mostly male-fertile plants. Taken together, these new results indicate that -mediated male fertility is under the control of more than one nuclear locus.

Fetal chromosome analysis: screening for chromosome disease?

DEFF Research Database (Denmark)

Philip, J; Tabor, Ann; Bang, J

1983-01-01

The aim of the study was to investigate the rationale of the current indications for fetal chromosome analysis. 5372 women had 5423 amniocentesis performed, this group constituting a consecutive sample at the chromosome laboratory, Rigshospitalet, Copenhagen from March 1973 to September 1980 (Group...... A + B). Pregnant women 35 years of age, women who previously had a chromosomally abnormal child, families with translocation carriers or other heritable chromosomal disease, families where the father was 50 years or more and women in families with a history of Down's syndrome (group A), were compared...... to women having amniocentesis, although considered not to have any increased risk of fetal chromosome abnormality (1390 pregnancies, group B). They were also compared with 750 consecutive pregnancies in women 25-34 years of age, in whom all heritable diseases were excluded (group C). The risk of unbalanced...

Loading of PAX3 to Mitotic Chromosomes Is Mediated by Arginine Methylation and Associated with Waardenburg Syndrome.

Science.gov (United States)

Wu, Tsu-Fang; Yao, Ya-Li; Lai, I-Lu; Lai, Chien-Chen; Lin, Pei-Lun; Yang, Wen-Ming

2015-08-14

PAX3 is a transcription factor critical to gene regulation in mammalian development. Mutations in PAX3 are associated with Waardenburg syndrome (WS), but the mechanism of how mutant PAX3 proteins cause WS remains unclear. Here, we found that PAX3 loads on mitotic chromosomes using its homeodomain. PAX3 WS mutants with mutations in homeodomain lose the ability to bind mitotic chromosomes. Moreover, loading of PAX3 on mitotic chromosomes requires arginine methylation, which is regulated by methyltransferase PRMT5 and demethylase JMJD6. Mutant PAX3 proteins that lose mitotic chromosome localization block cell proliferation and normal development of zebrafish. These results reveal the molecular mechanism of PAX3s loading on mitotic chromosomes and the importance of this localization pattern in normal development. Our findings suggest that PAX3 WS mutants interfere with the normal functions of PAX3 in a dominant negative manner, which is important to the understanding of the pathogenesis of Waardenburg syndrome. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

The Human Proteome Organization Chromosome 6 Consortium: integrating chromosome-centric and biology/disease driven strategies.

Science.gov (United States)

Borchers, C H; Kast, J; Foster, L J; Siu, K W M; Overall, C M; Binkowski, T A; Hildebrand, W H; Scherer, A; Mansoor, M; Keown, P A

2014-04-04

The Human Proteome Project (HPP) is designed to generate a comprehensive map of the protein-based molecular architecture of the human body, to provide a resource to help elucidate biological and molecular function, and to advance diagnosis and treatment of diseases. Within this framework, the chromosome-based HPP (C-HPP) has allocated responsibility for mapping individual chromosomes by country or region, while the biology/disease HPP (B/D-HPP) coordinates these teams in cross-functional disease-based groups. Chromosome 6 (Ch6) provides an excellent model for integration of these two tasks. This metacentric chromosome has a complement of 1002-1034 genes that code for known, novel or putative proteins. Ch6 is functionally associated with more than 120 major human diseases, many with high population prevalence, devastating clinical impact and profound societal consequences. The unique combination of genomic, proteomic, metabolomic, phenomic and health services data being drawn together within the Ch6 program has enormous potential to advance personalized medicine by promoting robust biomarkers, subunit vaccines and new drug targets. The strong liaison between the clinical and laboratory teams, and the structured framework for technology transfer and health policy decisions within Canada will increase the speed and efficacy of this transition, and the value of this translational research. Canada has been selected to play a leading role in the international Human Proteome Project, the global counterpart of the Human Genome Project designed to understand the structure and function of the human proteome in health and disease. Canada will lead an international team focusing on chromosome 6, which is functionally associated with more than 120 major human diseases, including immune and inflammatory disorders affecting the brain, skeletal system, heart and blood vessels, lungs, kidney, liver, gastrointestinal tract and endocrine system. Many of these chronic and persistent

Substrate analog interaction with MCR-1 offers insight into the rising threat of the plasmid-mediated transferable colistin resistance.

Science.gov (United States)

Wei, Pengcheng; Song, Guangji; Shi, Mengyang; Zhou, Yafei; Liu, Yang; Lei, Jun; Chen, Peng; Yin, Lei

2018-02-01

Colistin is considered a last-resort antibiotic against most gram-negative bacteria. Recent discoveries of a plasmid-mediated, transferable mobilized colistin-resistance gene ( mcr-1) on all continents have heralded the imminent emergence of pan-drug-resistant superbacteria. The inner-membrane protein MCR-1 can catalyze the transfer of phosphoethanolamine (PEA) to lipid A, resulting in colistin resistance. However, little is known about the mechanism, and few drugs exist to address this issue. We present crystal structures revealing the MCR-1 catalytic domain (cMCR-1) as a monozinc metalloprotein with ethanolamine (ETA) and d-glucose, respectively, thus highlighting 2 possible substrate-binding pockets in the MCR-1-catalyzed PEA transfer reaction. Mutation of the residues involved in ETA and d-glucose binding impairs colistin resistance in recombinant Escherichia coli containing full-length MCR-1. Partial analogs of the substrate are used for cocrystallization with cMCR-1, providing valuable information about the family of PEA transferases. One of the analogs, ETA, causes clear inhibition of polymyxin B resistance, highlighting its potential for drug development. These data demonstrate the crucial role of the PEA- and lipid A-binding pockets and provide novel insights into the structure-based mechanisms, important drug-target hot spots, and a drug template for further drug development to combat the urgent, rising threat of MCR-1-mediated antibiotic resistance.-Wei, P., Song, G., Shi, M., Zhou, Y., Liu, Y., Lei, J., Chen, P., Yin, L. Substrate analog interaction with MCR-1 offers insight into the rising threat of the plasmid-mediated transferable colistin resistance.

Membrane microparticles mediate transfer of P-glycoprotein to drug sensitive cancer cells.

Science.gov (United States)

Bebawy, M; Combes, V; Lee, E; Jaiswal, R; Gong, J; Bonhoure, A; Grau, G E R

2009-09-01

Multidrug resistance (MDR), a significant impediment to the successful treatment of cancer clinically, has been attributed to the overexpression of P-glycoprotein (P-gp), a plasma membrane multidrug efflux transporter. P-gp maintains sublethal intracellular drug concentrations by virtue of its drug efflux capacity. The cellular regulation of P-gp expression is currently known to occur at either pre- or post-transcriptional levels. In this study, we identify a 'non-genetic' mechanism whereby microparticles (MPs) serve as vectors in the acquisition and spread of MDR. MPs isolated from drug-resistant cancer cells (VLB(100)) were co-cultured with drug sensitive cells (CCRF-CEM) over a 4 h period to allow for MP binding and P-gp transfer. Presence of P-gp on MPs was established using flow cytometry (FCM) and western blotting. Whole-cell drug accumulation assays using rhodamine 123 and daunorubicin (DNR) were carried out to validate the transfer of functional P-gp after co-culture. We establish that MPs shed in vitro from drug-resistant cancer cells incorporate cell surface P-gp from their donor cells, effectively bind to drug-sensitive recipient cells and transfer functional P-gp to the latter. These findings serve to substantially advance our understanding of the molecular basis for the emergence of MDR in cancer clinically and lead to new treatment strategies which target and inhibit MP mediated transfer of P-gp during the course of treatment.

CTCF-mediated transcriptional regulation through cell type-specific chromosome organization in the β-globin locus

OpenAIRE

Junier, Ivan; Dale, Ryan K.; Hou, Chunhui; Képès, François; Dean, Ann

2012-01-01

International audience; The principles underlying the architectural landscape of chromatin beyond the nucleosome level in living cells remains largely unknown despite its potential to play a role in mammalian gene regulation. We investigated the three-dimensional folding of a 1 Mbp region of human chromosome 11 containing the β-globin genes by integrating looping interactions of the CCCTC-binding insulator protein CTCF determined comprehensively by chromosome conformation capture (3C) into a ...

Inter-chromosomal heterogeneity in the formation of radiation induced chromosomal aberrations

International Nuclear Information System (INIS)

Natarajan, A.T.; Vermeulen, S.; Boei, J.J.W.A.

1997-01-01

It is generally assumed that radiation induced chromosomal lesions are distributed randomly and repaired randomly among the genome. Recent studies using fluorescent in situ hybridization (FISH) and chromosome specific DNA libraries indicate that some chromosomes are more sensitive for radiation induced aberration formation than others. Chromosome No. 4 in human and chromosome No. 8 in Chinese hamster have been found to involve more in exchange aberrations than others, when calculated on the basis of their DNA content. Painting with arm specific chromosome libraries indicate that the frequencies of radiation induced intra-chromosome exchanges (i.e., between the arms of a chromosome, such as centric rings and inversions) are far in excess than one would expect on the basis of the frequencies of observed inter-chromosomal exchanges. The possible factors leading to the observed heterogeneity will be discussed

Bayesian analysis of overdispersed chromosome aberration data with the negative binomial model

International Nuclear Information System (INIS)

Brame, R.S.; Groer, P.G.

2002-01-01

The usual assumption of a Poisson model for the number of chromosome aberrations in controlled calibration experiments implies variance equal to the mean. However, it is known that chromosome aberration data from experiments involving high linear energy transfer radiations can be overdispersed, i.e. the variance is greater than the mean. Present methods for dealing with overdispersed chromosome data rely on frequentist statistical techniques. In this paper, the problem of overdispersion is considered from a Bayesian standpoint. The Bayes Factor is used to compare Poisson and negative binomial models for two previously published calibration data sets describing the induction of dicentric chromosome aberrations by high doses of neutrons. Posterior densities for the model parameters, which characterise dose response and overdispersion are calculated and graphed. Calibrative densities are derived for unknown neutron doses from hypothetical radiation accident data to determine the impact of different model assumptions on dose estimates. The main conclusion is that an initial assumption of a negative binomial model is the conservative approach to chromosome dosimetry for high LET radiations. (author)

Mitotic chromosome structure

International Nuclear Information System (INIS)

Heermann, Dieter W.

2012-01-01

Mounting evidence is compiling linking the physical organizational structure of chromosomes and the nuclear structure to biological function. At the base of the physical organizational structure of both is the concept of loop formation. This implies that physical proximity within chromosomes is provided for otherwise distal genomic regions and thus hierarchically organizing the chromosomes. Together with entropy many experimental observations can be explained with these two concepts. Among the observations that can be explained are the measured physical extent of the chromosomes, their shape, mechanical behavior, the segregation into territories (chromosomal and territories within chromosomes), the results from chromosome conformation capture experiments, as well as linking gene expression to structural organization.

Mitotic chromosome structure

Energy Technology Data Exchange (ETDEWEB)

Heermann, Dieter W., E-mail: heermann@tphys.uni-heidelberg.de

2012-07-15

Mounting evidence is compiling linking the physical organizational structure of chromosomes and the nuclear structure to biological function. At the base of the physical organizational structure of both is the concept of loop formation. This implies that physical proximity within chromosomes is provided for otherwise distal genomic regions and thus hierarchically organizing the chromosomes. Together with entropy many experimental observations can be explained with these two concepts. Among the observations that can be explained are the measured physical extent of the chromosomes, their shape, mechanical behavior, the segregation into territories (chromosomal and territories within chromosomes), the results from chromosome conformation capture experiments, as well as linking gene expression to structural organization.

Nonadiabatic two-electron transfer mediated by an irregular bridge

International Nuclear Information System (INIS)

Petrov, E.G.; Shevchenko, Ye.V.; May, V.

2004-01-01

Nonadiabatic two-electron transfer (TET) mediated by a linear molecular bridge is studied theoretically. Special attention is put on the case of a irregular distribution of bridge site energies as well as on the inter-site Coulomb interaction. Based on the unified description of electron transfer reactions [J. Chem. Phys. 115 (2001) 7107] a closed set of kinetic equations describing the TET process is derived. A reduction of this set to a single exponential donor-acceptor (D-A) TET is performed together with a derivation of an overall D-A TET rate. The latter contains a contribution of the stepwise as well as of the concerted route of D-A TET. The stepwise contribution is determined by two single-electron steps each of them associated with a sequential and a superexchange pathway. A two-electron unistep superexchange transition between the D and A forms the concerted contribution to the overall rate. Both contributions are analyzed in their dependency on the bridge length. The irregular distribution of the bridge site energies as well as the influence of the Coulomb interaction facilitates the D-A TET via a modification of the stepwise and the concerted part of the overall rate. At low temperatures and for short bridges with a single or two units the concerted contribution exceeds the stepwise contribution. If the bridge contains more than two units, the stepwise contribution dominates the overall rate

Repetitive sequences and epigenetic modification: inseparable partners play important roles in the evolution of plant sex chromosomes.

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Li, Shu-Fen; Zhang, Guo-Jun; Yuan, Jin-Hong; Deng, Chuan-Liang; Gao, Wu-Jun

2016-05-01

The present review discusses the roles of repetitive sequences played in plant sex chromosome evolution, and highlights epigenetic modification as potential mechanism of repetitive sequences involved in sex chromosome evolution. Sex determination in plants is mostly based on sex chromosomes. Classic theory proposes that sex chromosomes evolve from a specific pair of autosomes with emergence of a sex-determining gene(s). Subsequently, the newly formed sex chromosomes stop recombination in a small region around the sex-determining locus, and over time, the non-recombining region expands to almost all parts of the sex chromosomes. Accumulation of repetitive sequences, mostly transposable elements and tandem repeats, is a conspicuous feature of the non-recombining region of the Y chromosome, even in primitive one. Repetitive sequences may play multiple roles in sex chromosome evolution, such as triggering heterochromatization and causing recombination suppression, leading to structural and morphological differentiation of sex chromosomes, and promoting Y chromosome degeneration and X chromosome dosage compensation. In this article, we review the current status of this field, and based on preliminary evidence, we posit that repetitive sequences are involved in sex chromosome evolution probably via epigenetic modification, such as DNA and histone methylation, with small interfering RNAs as the mediator.

Chromosome-wise dissection of the genome of the extremely big mouse line DU6i.

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Bevova, Marianna R; Aulchenko, Yurii S; Aksu, Soner; Renne, Ulla; Brockmann, Gudrun A

2006-01-01

The extreme high-body-weight-selected mouse line DU6i is a polygenic model for growth research, harboring many small-effect QTL. We dissected the genome of this line into 19 autosomes and the Y chromosome by the construction of a new panel of chromosome substitution strains (CSS). The DU6i chromosomes were transferred to a DBA/2 mice genetic background by marker-assisted recurrent backcrossing. Mitochondria and the X chromosome were of DBA/2 origin in the backcross. During the construction of these novel strains, >4000 animals were generated, phenotyped, and genotyped. Using these data, we studied the genetic control of variation in body weight and weight gain at 21, 42, and 63 days. The unique data set facilitated the analysis of chromosomal interaction with sex and parent-of-origin effects. All analyzed chromosomes affected body weight and weight gain either directly or in interaction with sex or parent of origin. The effects were age specific, with some chromosomes showing opposite effects at different stages of development.

Wrestling with Chromosomes: The Roles of SUMO During Meiosis.

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Nottke, Amanda C; Kim, Hyun-Min; Colaiácovo, Monica P

2017-01-01

Meiosis is a specialized form of cell division required for the formation of haploid gametes and therefore is essential for successful sexual reproduction. Various steps are exquisitely coordinated to ensure accurate chromosome segregation during meiosis, thereby promoting the formation of haploid gametes from diploid cells. Recent studies are demonstrating that an important form of regulation during meiosis is exerted by the post-translational protein modification known as sumoylation. Here, we review and discuss the various critical steps of meiosis in which SUMO-mediated regulation has been implicated thus far. These include the maintenance of meiotic centromeric heterochromatin , meiotic DNA double-strand break repair and homologous recombination, centromeric coupling, and the assembly of a proteinaceous scaffold between homologous chromosomes known as the synaptonemal complex.

Chromosomal aberration

International Nuclear Information System (INIS)

Ishii, Yutaka

1988-01-01

Chromosomal aberrations are classified into two types, chromosome-type and chromatid-type. Chromosom-type aberrations include terminal deletion, dicentric, ring and interstitial deletion, and chromatid-type aberrations include achromatic lesion, chromatid deletion, isochromatid deletion and chromatid exchange. Clastogens which induce chromosomal aberration are divided into ''S-dependent'' agents and ''S-independent''. It might mean whether they can induce double strand breaks independent of the S phase or not. Double strand breaks may be the ultimate lesions to induce chromosomal aberrations. Caffeine added even in the G 2 phase appeared to modify the frequency of chromatid aberrations induced by X-rays and mitomycin C. Those might suggest that the G 2 phase involves in the chromatid aberration formation. The double strand breaks might be repaired by ''G 2 repair system'', the error of which might yield breakage types of chromatid aberrations and the by-pass of which might yield chromatid exchanges. Chromosome-type aberrations might be formed in the G 1 phase. (author)

TMAP/CKAP2 is essential for proper chromosome segregation.

Science.gov (United States)

Hong, Kyung Uk; Kim, Eunhee; Bae, Chang-Dae; Park, Joobae

2009-01-15

Tumor-associated microtubule-associated protein (TMAP), also known as cytoskeleton associated protein 2 (CKAP2), is a novel mitotic spindle-associated protein which is frequently up-regulated in various malignances. However, its cellular functions remain unknown. Previous reports suggested that the cellular functions of TMAP/CKAP2 pertain to regulation of the dynamics and assembly of the mitotic spindle. To investigate its role in mitosis, we studied the effects of siRNA-mediated depletion of TMAP/CKAP2 in cultured mammalian cells. Unexpectedly, TMAP/CKAP2 knockdown did not result in significant alterations of the spindle apparatus. However, TMAP/CKAP2-depleted cells often exhibited abnormal nuclear morphologies, which were accompanied by abnormal organization of the nuclear lamina, and chromatin bridge formation between two daughter cell nuclei. Time lapse video microscopy revealed that the changes in nuclear morphology and chromatin bridge formations observed in TMAP/CKAP2-depleted cells are the result of defects in chromosome segregation. Consistent with this, the spindle checkpoint activity was significantly reduced in TMAP/CKAP2-depleted cells. Moreover, chromosome missegregation induced by depletion of TMAP/CKAP2 ultimately resulted in reduced cell viability and increased chromosomal instability. Our present findings demonstrate that TMAP/CKAP2 is essential for proper chromosome segregation and for maintaining genomic stability.

Y-chromosome evolution: emerging insights into processes of Y-chromosome degeneration.

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Bachtrog, Doris

2013-02-01

The human Y chromosome is intriguing not only because it harbours the master-switch gene that determines gender but also because of its unusual evolutionary history. The Y chromosome evolved from an autosome, and its evolution has been characterized by massive gene decay. Recent whole-genome and transcriptome analyses of Y chromosomes in humans and other primates, in Drosophila species and in plants have shed light on the current gene content of the Y chromosome, its origins and its long-term fate. Furthermore, comparative analysis of young and old Y chromosomes has given further insights into the evolutionary and molecular forces triggering Y-chromosome degeneration and into the evolutionary destiny of the Y chromosome.

Loss of heterozygosity of chromosome 15 in human lung carcinomas

International Nuclear Information System (INIS)

Mitchell, C.E.; Palmisano, W.A.; Lechner, J.F.

1994-01-01

Loss of heterozygosity (LOH) in tumors may be associated with the inactivation of tumor suppressor genes. A tumor suppressor gene for lung cancer may reside on chromosome 15, because deletions in this chromosome are frequently observed. Recently, it was reported that a newly discovered gene, GTPase-activating protein-3 (GAP3) maps to chromosome 15. GAP3 is a member of a family of GAP-related genes. Although the precise function of GAP3 is not known, it is thought that GAP3 is involved in the regulation of ras-like GTPase activities. Ras proteins have a low intrinsic activity, and their inactivation is dependent on GAPS in vivo. Oncogenic mutants of ras proteins, for example, at codons 12, 13, or 61, are resistant to GAP-mediated GTPase stimulation and are constituitively locked in their active, GTP-bound states. The purpose of this investigation was to determine the frequency and extent of LOH of GAP3 in a group of patients with lung cancer

Cooperative electrocatalytic alcohol oxidation with electron-proton-transfer mediators

Science.gov (United States)

Badalyan, Artavazd; Stahl, Shannon S.

2016-07-01

electron-proton-transfer mediators, such as TEMPO, may be used in combination with first-row transition metals, such as copper, to achieve efficient two-electron electrochemical processes, thereby introducing a new concept for the development of non-precious-metal electrocatalysts.

Chromosomal abnormalities in human glioblastomas: gain in chromosome 7p correlating with loss in chromosome 10q.

Science.gov (United States)

Inda, María del Mar; Fan, Xing; Muñoz, Jorge; Perot, Christine; Fauvet, Didier; Danglot, Giselle; Palacio, Ana; Madero, Pilar; Zazpe, Idoya; Portillo, Eduardo; Tuñón, Teresa; Martínez-Peñuela, José María; Alfaro, Jorge; Eiras, José; Bernheim, Alain; Castresana, Javier S

2003-01-01

Various genomic alterations have been detected in glioblastoma. Chromosome 7p, with the epidermal growth factor receptor locus, together with chromosome 10q, with the phosphatase and tensin homologue deleted in chromosome 10 and deleted in malignant brain tumors-1 loci, and chromosome 9p, with the cyclin-dependent kinase inhibitor 2A locus, are among the most frequently damaged chromosomal regions in glioblastoma. In this study, we evaluated the genetic status of 32 glioblastomas by comparative genomic hybridization; the sensitivity of comparative genomic hybridization versus differential polymerase chain reaction to detect deletions at the phosphatase and tensin homologue deleted in chromosome 10, deleted in malignant brain tumors-1, and cyclin-dependent kinase inhibitor 2A loci and amplifications at the cyclin-dependent kinase 4 locus; the frequency of genetic lesions (gain or loss) at 16 different selected loci (including oncogenes, tumor-suppressor genes, and proliferation markers) mapping on 13 different chromosomes; and the possible existence of a statistical association between any pair of molecular markers studied, to subdivide the glioblastoma entity molecularly. Comparative genomic hybridization showed that the most frequent region of gain was chromosome 7p, whereas the most frequent losses occurred on chromosomes 10q and 13q. The only statistically significant association was found for 7p gain and 10q loss. Copyright 2002 Wiley-Liss, Inc.

Observation of electron-transfer-mediated decay in aqueous solution

Science.gov (United States)

Unger, Isaak; Seidel, Robert; Thürmer, Stephan; Pohl, Marvin N.; Aziz, Emad F.; Cederbaum, Lorenz S.; Muchová, Eva; Slavíček, Petr; Winter, Bernd; Kryzhevoi, Nikolai V.

2017-07-01

Photoionization is at the heart of X-ray photoelectron spectroscopy (XPS), which gives access to important information on a sample's local chemical environment. Local and non-local electronic decay after photoionization—in which the refilling of core holes results in electron emission from either the initially ionized species or a neighbour, respectively—have been well studied. However, electron-transfer-mediated decay (ETMD), which involves the refilling of a core hole by an electron from a neighbouring species, has not yet been observed in condensed phase. Here we report the experimental observation of ETMD in an aqueous LiCl solution by detecting characteristic secondary low-energy electrons using liquid-microjet soft XPS. Experimental results are interpreted using molecular dynamics and high-level ab initio calculations. We show that both solvent molecules and counterions participate in the ETMD processes, and different ion associations have distinctive spectral fingerprints. Furthermore, ETMD spectra are sensitive to coordination numbers, ion-solvent distances and solvent arrangement.

Chromosome engineering for alien gene introgression in wheat: Progress and prospective

Science.gov (United States)

Chromosome engineering is a useful strategy for introgression of desirable genes from wild relatives into cultivated wheat. However, it has been a challenge to transfer a small amount of alien chromatin containing the gene of interest from one genome to another non-homologous genome through classic...

De novo formed satellite DNA-based mammalian artificial chromosomes and their possible applications.

Science.gov (United States)

Katona, Robert L

2015-02-01

Mammalian artificial chromosomes (MACs) are non-integrating, autonomously replicating natural chromosome-based vectors that may carry a vast amount of genetic material, which in turn enable potentially prolonged, safe, and regulated therapeutic transgene expression and render MACs as attractive genetic vectors for "gene replacement" or for controlling differentiation pathways in target cells. Satellite-DNA-based artificial chromosomes (SATACs) can be made by induced de novo chromosome formation in cells of different mammalian and plant species. These artificially generated accessory chromosomes are composed of predictable DNA sequences, and they contain defined genetic information. SATACs have already passed a number of obstacles crucial to their further development as gene therapy vectors, including large-scale purification, transfer of purified artificial chromosomes into different cells and embryos, generation of transgenic animals and germline transmission with purified SATACs, and the tissue-specific expression of a therapeutic gene from an artificial chromosome in the milk of transgenic animals. SATACs could be used in cell therapy protocols. For these methods, the most versatile target cell would be one that was pluripotent and self-renewing to address multiple disease target cell types, thus making multilineage stem cells, such as adult derived early progenitor cells and embryonic stem cells, as attractive universal host cells.

A dominant negative mutant of TLK1 causes chromosome missegregation and aneuploidy in normal breast epithelial cells

Directory of Open Access Journals (Sweden)

Williams Briana

2003-10-01

Full Text Available Abstract Background In Arabidopsis thaliana, the gene Tousled encodes a protein kinase of unknown function, but mutations in the gene lead to flowering and leaf morphology defects. We have recently cloned a mammalian Tousled-Like Kinase (TLK1B and found that it phosphorylates specifically histone H3, in vitro and in vivo. We now report the effects that overexpression of a kinase-dead mutant of TLK1B mediates in a normal diploid cell line. Results Expression of a kinase-dead mutant resulted in reduction of phosphorylated histone H3, which could have consequences in mitotic segregation of chromosomes. When analyzed by FACS and microscopy, these cells displayed high chromosome number instability and aneuploidy. This phenomenon was accompanied by less condensed chromosomes at mitosis; failure of a number of chromosomes to align properly on the metaphase plate; failure of some chromosomes to attach to microtubules; and the occasional presentation of two bipolar spindles. We also used a different method (siRNA to reduce the level of endogenous TLK1, but in this case, the main result was a strong block of cell cycle progression suggesting that TLK1 may also play a role in progression from G1. This block in S phase progression could also offer a different explanation of some of the later mitotic defects. Conclusions TLK1 has a function important for proper chromosome segregation and maintenance of diploid cells at mitosis in mammalian cells that could be mediated by reduced phosphorylation of histone H3 and condensation of chromosomes, although other explanations to the phenotype are possible.

A Family of Zinc Finger Proteins Is Required forChromosome-specific Pairing and Synapsis during Meiosis in C.elegans

Energy Technology Data Exchange (ETDEWEB)

Phillips, Carolyn M.; Dernburg, Abby F.

2006-06-07

Homologous chromosome pairing and synapsis are prerequisitefor accurate chromosome segregation during meiosis. Here, we show that afamily of four related C2H2 zinc-finger proteins plays a central role inthese events in C. elegans. These proteins are encoded within a tandemgene cluster. In addition to the X-specific HIM-8 protein, threeadditional paralogs collectively mediate the behavior of the fiveautosomes. Each chromosome relies on a specific member of the family topair and synapse with its homolog. These "ZIM" proteins concentrate atspecial regions called meiotic pairing centers on the correspondingchromosomes. These sites are dispersed along the nuclear envelope duringearly meiotic prophase, suggesting a role analogous to thetelomere-mediated meiotic bouquet in other organisms. To gain insightinto the evolution of these components, wecharacterized homologs in C.briggsae and C. remanei, which revealed changes in copy number of thisgene family within the nematode lineage.

Slit scan flow cytometry of isolated chromosomes following fluorescence hybridization: an approach of online screening for specific chromosomes and chromosome translocations

NARCIS (Netherlands)

Hausmann, M.; Dudin, G.; Aten, J. A.; Heilig, R.; Diaz, E.; Cremer, C.

1991-01-01

The recently developed methods of non radioactive in situ hybridization of chromosomes offer new aspects for chromosome analysis. Fluorescent labelling of hybridized chromosomes or chromosomal subregions allows to facilitate considerably the detection of specific chromosomal abnormalities. For many

An examination of mediators of the transfer of cognitive speed of processing training to everyday functional performance.

Science.gov (United States)

Edwards, Jerri D; Ruva, Christine L; O'Brien, Jennifer L; Haley, Christine B; Lister, Jennifer J

2013-06-01

The purpose of these analyses was to examine mediators of the transfer of cognitive speed of processing training to improved everyday functional performance (J. D. Edwards, V. G. Wadley,, D. E. Vance, D. L. Roenker, & K. K. Ball, 2005, The impact of speed of processing training on cognitive and everyday performance. Aging & Mental Health, 9, 262-271). Cognitive speed of processing and visual attention (as measured by the Useful Field of View Test; UFOV) were examined as mediators of training transfer. Secondary data analyses were conducted from the Staying Keen in Later Life (SKILL) study, a randomized cohort study including 126 community dwelling adults 63 to 87 years of age. In the SKILL study, participants were randomized to an active control group or cognitive speed of processing training (SOPT), a nonverbal, computerized intervention involving perceptual practice of visual tasks. Prior analyses found significant effects of training as measured by the UFOV and Timed Instrumental Activities of Daily Living (TIADL) Tests. Results from the present analyses indicate that speed of processing for a divided attention task significantly mediated the effect of SOPT on everyday performance (e.g., TIADL) in a multiple mediation model accounting for 91% of the variance. These findings suggest that everyday functional improvements found from SOPT are directly attributable to improved UFOV performance, speed of processing for divided attention in particular. Targeting divided attention in cognitive interventions may be important to positively affect everyday functioning among older adults. PsycINFO Database Record (c) 2013 APA, all rights reserved.

Adenoviral vector-mediated gene transfer and neurotransplantation : possibilities and limitations in grafting of the fetal rat suprachiasmatic nucleus

NARCIS (Netherlands)

van Esseveldt, K E; Liu, R.; Hermens, W.T.J.M.C.; Verhaagen, J; Boer, G J

Several studies have reported on the use of primary neural cells transduced by adenoviral vectors as donor cells in neurotransplantation. In the present investigation, we examined whether adenoviral vector-mediated gene transfer could be used to introduce and express a foreign gene in solid neural

Rare recombination events generate sequence diversity among balancer chromosomes in Drosophila melanogaster.

Science.gov (United States)

Miller, Danny E; Cook, Kevin R; Yeganeh Kazemi, Nazanin; Smith, Clarissa B; Cockrell, Alexandria J; Hawley, R Scott; Bergman, Casey M

2016-03-08

Multiply inverted balancer chromosomes that suppress exchange with their homologs are an essential part of the Drosophila melanogaster genetic toolkit. Despite their widespread use, the organization of balancer chromosomes has not been characterized at the molecular level, and the degree of sequence variation among copies of balancer chromosomes is unknown. To map inversion breakpoints and study potential diversity in descendants of a structurally identical balancer chromosome, we sequenced a panel of laboratory stocks containing the most widely used X chromosome balancer, First Multiple 7 (FM7). We mapped the locations of FM7 breakpoints to precise euchromatic coordinates and identified the flanking sequence of breakpoints in heterochromatic regions. Analysis of SNP variation revealed megabase-scale blocks of sequence divergence among currently used FM7 stocks. We present evidence that this divergence arose through rare double-crossover events that replaced a female-sterile allele of the singed gene (sn(X2)) on FM7c with a sequence from balanced chromosomes. We propose that although double-crossover events are rare in individual crosses, many FM7c chromosomes in the Bloomington Drosophila Stock Center have lost sn(X2) by this mechanism on a historical timescale. Finally, we characterize the original allele of the Bar gene (B(1)) that is carried on FM7, and validate the hypothesis that the origin and subsequent reversion of the B(1) duplication are mediated by unequal exchange. Our results reject a simple nonrecombining, clonal mode for the laboratory evolution of balancer chromosomes and have implications for how balancer chromosomes should be used in the design and interpretation of genetic experiments in Drosophila.

CDE-1 affects chromosome segregation through uridylation of CSR-1-bound siRNAs.

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van Wolfswinkel, Josien C; Claycomb, Julie M; Batista, Pedro J; Mello, Craig C; Berezikov, Eugene; Ketting, René F

2009-10-02

We have studied the function of a conserved germline-specific nucleotidyltransferase protein, CDE-1, in RNAi and chromosome segregation in C. elegans. CDE-1 localizes specifically to mitotic chromosomes in embryos. This localization requires the RdRP EGO-1, which physically interacts with CDE-1, and the Argonaute protein CSR-1. We found that CDE-1 is required for the uridylation of CSR-1 bound siRNAs, and that in the absence of CDE-1 these siRNAs accumulate to inappropriate levels, accompanied by defects in both meiotic and mitotic chromosome segregation. Elevated siRNA levels are associated with erroneous gene silencing, most likely through the inappropriate loading of CSR-1 siRNAs into other Argonaute proteins. We propose a model in which CDE-1 restricts specific EGO-1-generated siRNAs to the CSR-1 mediated, chromosome associated RNAi pathway, thus separating it from other endogenous RNAi pathways. The conserved nature of CDE-1 suggests that similar sorting mechanisms may operate in other animals, including mammals.

Speaker transfer in children's peer conversation: completing communication-aid-mediated contributions.

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Clarke, Michael; Bloch, Steven; Wilkinson, Ray

2013-03-01

Managing the exchange of speakers from one person to another effectively is a key issue for participants in everyday conversational interaction. Speakers use a range of resources to indicate, in advance, when their turn will come to an end, and listeners attend to such signals in order to know when they might legitimately speak. Using the principles and findings from conversation analysis, this paper examines features of speaker transfer in a conversation between a boy with cerebral palsy who has been provided with a voice-output communication aid (VOCA), and a peer without physical or communication difficulties. Specifically, the analysis focuses on turn exchange, where a VOCA-mediated contribution approach completion, and the child without communication needs is due to speak next.

Exogenous electron shuttle-mediated extracellular electron transfer of Shewanella putrefaciens 200: electrochemical parameters and thermodynamics.

Science.gov (United States)

Wu, Yundang; Liu, Tongxu; Li, Xiaomin; Li, Fangbai

2014-08-19

Despite the importance of exogenous electron shuttles (ESs) in extracellular electron transfer (EET), a lack of understanding of the key properties of ESs is a concern given their different influences on EET processes. Here, the ES-mediated EET capacity of Shewanella putrefaciens 200 (SP200) was evaluated by examining the electricity generated in a microbial fuel cell. The results indicated that all the ESs substantially accelerated the current generation compared to only SP200. The current and polarization parameters were linearly correlated with both the standard redox potential (E(ES)(0)) and the electron accepting capacity (EAC) of the ESs. A thermodynamic analysis of the electron transfer from the electron donor to the electrode suggested that the EET from c-type cytochromes (c-Cyts) to ESs is a crucial step causing the differences in EET capacities among various ESs. Based on the derived equations, both E(ES)(0) and EAC can quantitatively determine potential losses (ΔE) that reflect the potential loss of the ES-mediated EET. In situ spectral kinetic analysis of ES reduction by c-Cyts in a living SP200 suspension was first investigated with the E(ES), E(c-Cyt), and ΔE values being calculated. This study can provide a comprehensive understanding of the role of ESs in EET.

APC/C Dysfunction Limits Excessive Cancer Chromosomal Instability.

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Sansregret, Laurent; Patterson, James O; Dewhurst, Sally; López-García, Carlos; Koch, André; McGranahan, Nicholas; Chao, William Chong Hang; Barry, David J; Rowan, Andrew; Instrell, Rachael; Horswell, Stuart; Way, Michael; Howell, Michael; Singleton, Martin R; Medema, René H; Nurse, Paul; Petronczki, Mark; Swanton, Charles

2017-02-01

Intercellular heterogeneity, exacerbated by chromosomal instability (CIN), fosters tumor heterogeneity and drug resistance. However, extreme CIN correlates with improved cancer outcome, suggesting that karyotypic diversity required to adapt to selection pressures might be balanced in tumors against the risk of excessive instability. Here, we used a functional genomics screen, genome editing, and pharmacologic approaches to identify CIN-survival factors in diploid cells. We find partial anaphase-promoting complex/cyclosome (APC/C) dysfunction lengthens mitosis, suppresses pharmacologically induced chromosome segregation errors, and reduces naturally occurring lagging chromosomes in cancer cell lines or following tetraploidization. APC/C impairment caused adaptation to MPS1 inhibitors, revealing a likely resistance mechanism to therapies targeting the spindle assembly checkpoint. Finally, CRISPR-mediated introduction of cancer somatic mutations in the APC/C subunit cancer driver gene CDC27 reduces chromosome segregation errors, whereas reversal of an APC/C subunit nonsense mutation increases CIN. Subtle variations in mitotic duration, determined by APC/C activity, influence the extent of CIN, allowing cancer cells to dynamically optimize fitness during tumor evolution. We report a mechanism whereby cancers balance the evolutionary advantages associated with CIN against the fitness costs caused by excessive genome instability, providing insight into the consequence of CDC27 APC/C subunit driver mutations in cancer. Lengthening of mitosis through APC/C modulation may be a common mechanism of resistance to cancer therapeutics that increase chromosome segregation errors. Cancer Discov; 7(2); 218-33. ©2017 AACR.See related commentary by Burkard and Weaver, p. 134This article is highlighted in the In This Issue feature, p. 115. ©2017 American Association for Cancer Research.

DNA damage and chromosome aberration induced by heavy-ion beams

International Nuclear Information System (INIS)

Takakura, Kahoru; Funada, Aya; Aoki, Mizuho; Furusawa, Yoshiya

2003-01-01

The aim of this study is to clarify the relation between cell death and chromosomal aberration in cultured human cells (human salivary gland (HSG) tumor cells and GM05389 human normal fibroblasts) irradiated with heavy ion beams on the basis of linear energy transfer (LET) values. The LET dependences of cell death were observed for the both cells by the method of colony assay. The LET dependences of the chromosomal aberrations, breaks and gaps, isochromatid breaks and exchanges were also observed for the both cells using the premature chromosome condensation (PCC) method. From these results it is suggested that exchange formation is essential for the cell death caused by heavy ion beam irradiation. It is suspected that the densely ionizing track structure of hight LET heavy ions inhibits the effective repair in the chromatid breaks and isochromatid breaks and finally induce much exchange in the cells, which should be essential cause of cell death. (author)

Rapid and efficient introduction of a foreign gene into bacterial artificial chromosome-cloned varicella vaccine by Tn7-mediated site-specific transposition

International Nuclear Information System (INIS)

Somboonthum, Pranee; Koshizuka, Tetsuo; Okamoto, Shigefumi; Matsuura, Masaaki; Gomi, Yasuyuki; Takahashi, Michiaki; Yamanishi, Koichi; Mori, Yasuko

2010-01-01

Using a rapid and reliable system based on Tn7-mediated site-specific transposition, we have successfully constructed a recombinant Oka varicella vaccine (vOka) expressing the mumps virus (MuV) fusion protein (F). The backbone of the vector was our previously reported vOka-BAC (bacterial artificial chromosome) genome. We inserted the transposon Tn7 attachment sequence, LacZα-mini-attTn7, into the region between ORF12 and ORF13 to generate a vOka-BAC-Tn genome. The MuV-F expressing cassette was transposed into the vOka-BAC genome at the mini-attTn7 transposition site. MuV-F protein was expressed in recombinant virus, rvOka-F infected cells. In addition, the MuV-F protein was cleaved in the rvOka-F infected cells as in MuV-infected cells. The growth of rvOka-F was similar to that of the original recombinant vOka without the F gene. Thus, we show that Tn7-mediated transposition is an efficient method for introducing a foreign gene expression cassette into the vOka-BAC genome as a live virus vector.

Enrichment of dynamic chromosomal crosslinks drive phase separation of the nucleolus.

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Hult, Caitlin; Adalsteinsson, David; Vasquez, Paula A; Lawrimore, Josh; Bennett, Maggie; York, Alyssa; Cook, Diana; Yeh, Elaine; Forest, Mark Gregory; Bloom, Kerry

2017-11-02

Regions of highly repetitive DNA, such as those found in the nucleolus, show a self-organization that is marked by spatial segregation and frequent self-interaction. The mechanisms that underlie the sequestration of these sub-domains are largely unknown. Using a stochastic, bead-spring representation of chromatin in budding yeast, we find enrichment of protein-mediated, dynamic chromosomal cross-links recapitulates the segregation, morphology and self-interaction of the nucleolus. Rates and enrichment of dynamic crosslinking have profound consequences on domain morphology. Our model demonstrates the nucleolus is phase separated from other chromatin in the nucleus and predicts that multiple rDNA loci will form a single nucleolus independent of their location within the genome. Fluorescent labeling of budding yeast nucleoli with CDC14-GFP revealed that a split rDNA locus indeed forms a single nucleolus. We propose that nuclear sub-domains, such as the nucleolus, result from phase separations within the nucleus, which are driven by the enrichment of protein-mediated, dynamic chromosomal crosslinks. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Recombinase, chromosomal translocations and lymphoid neoplasia: targeting mistakes and repair failures.

Science.gov (United States)

Marculescu, Rodrig; Vanura, Katrina; Montpellier, Bertrand; Roulland, Sandrine; Le, Trang; Navarro, Jean-Marc; Jäger, Ulrich; McBlane, Fraser; Nadel, Bertrand

2006-09-08

A large number of lymphoid malignancies is characterized by specific chromosomal translocations, which are closely linked to the initial steps of pathogenesis. The hallmark of these translocations is the ectopic activation of a silent proto-oncogene through its relocation at the vicinity of an active regulatory element. Due to the unique feature of lymphoid cells to somatically rearrange and mutate receptor genes, and to the corresponding strong activity of the immune enhancers/promoters at that stage of cell development, B- and T-cell differentiation pathways represent propitious targets for chromosomal translocations and oncogene activation. Recent progress in the understanding of the V(D)J recombination process has allowed a more accurate definition of the translocation mechanisms involved, and has revealed that V(D)J-mediated translocations result both from targeting mistakes of the recombinase, and from illegitimate repair of the V(D)J recombination intermediates. Surprisingly, V(D)J-mediated translocations turn out to be restricted to two specific sub-types of lymphoid malignancies, T-cell acute lymphoblastic leukemias, and a restricted set of mature B-cell Non-Hodgkin's lymphomas.

Rab11b mediates melanin transfer between donor melanocytes and acceptor keratinocytes via coupled exo/endocytosis.

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Tarafder, Abul K; Bolasco, Giulia; Correia, Maria S; Pereira, Francisco J C; Iannone, Lucio; Hume, Alistair N; Kirkpatrick, Niall; Picardo, Mauro; Torrisi, Maria R; Rodrigues, Inês P; Ramalho, José S; Futter, Clare E; Barral, Duarte C; Seabra, Miguel C

2014-04-01

The transfer of melanin from melanocytes to keratinocytes is a crucial process underlying maintenance of skin pigmentation and photoprotection against UV damage. Here, we present evidence supporting coupled exocytosis of the melanin core, or melanocore, by melanocytes and subsequent endocytosis by keratinocytes as a predominant mechanism of melanin transfer. Electron microscopy analysis of human skin samples revealed three lines of evidence supporting this: (1) the presence of melanocores in the extracellular space; (2) within keratinocytes, melanin was surrounded by a single membrane; and (3) this membrane lacked the melanosomal membrane protein tyrosinase-related protein 1 (TYRP1). Moreover, co-culture of melanocytes and keratinocytes suggests that melanin exocytosis is specifically induced by keratinocytes. Furthermore, depletion of Rab11b, but not Rab27a, caused a marked decrease in both keratinocyte-stimulated melanin exocytosis and transfer to keratinocytes. Thus, we propose that the predominant mechanism of melanin transfer is keratinocyte-induced exocytosis, mediated by Rab11b through remodeling of the melanosome membrane, followed by subsequent endocytosis by keratinocytes.

Mode of ATM-dependent suppression of chromosome translocation

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Yamauchi, Motohiro, E-mail: motoyama@nagasaki-u.ac.jp [Graduate School of Biomedical Sciences, Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523 (Japan); Suzuki, Keiji; Oka, Yasuyoshi; Suzuki, Masatoshi; Kondo, Hisayoshi; Yamashita, Shunichi [Graduate School of Biomedical Sciences, Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523 (Japan)

2011-12-09

Highlights: Black-Right-Pointing-Pointer We addressed how ATM suppresses frequency of chromosome translocation. Black-Right-Pointing-Pointer We found ATM/p53-dependent G1 checkpoint suppresses translocation frequency. Black-Right-Pointing-Pointer We found ATM and DNA-PKcs function in a common pathway to suppress translocation. -- Abstract: It is well documented that deficiency in ataxia telangiectasia mutated (ATM) protein leads to elevated frequency of chromosome translocation, however, it remains poorly understood how ATM suppresses translocation frequency. In the present study, we addressed the mechanism of ATM-dependent suppression of translocation frequency. To know frequency of translocation events in a whole genome at once, we performed centromere/telomere FISH and scored dicentric chromosomes, because dicentric and translocation occur with equal frequency and by identical mechanism. By centromere/telomere FISH analysis, we confirmed that chemical inhibition or RNAi-mediated knockdown of ATM causes 2 to 2.5-fold increase in dicentric frequency at first mitosis after 2 Gy of gamma-irradiation in G0/G1. The FISH analysis revealed that ATM/p53-dependent G1 checkpoint suppresses dicentric frequency, since RNAi-mediated knockdown of p53 elevated dicentric frequency by 1.5-fold. We found ATM also suppresses dicentric occurrence independently of its checkpoint role, as ATM inhibitor showed additional effect on dicentric frequency in the context of p53 depletion and Chk1/2 inactivation. Epistasis analysis using chemical inhibitors revealed that ATM kinase functions in the same pathway that requires kinase activity of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) to suppress dicentric frequency. From the results in the present study, we conclude that ATM minimizes translocation frequency through its commitment to G1 checkpoint and DNA double-strand break repair pathway that requires kinase activity of DNA-PKcs.

Enhancement and Tunability of Near-Field Radiative Heat Transfer Mediated by Surface Plasmon Polaritons in Thin Plasmonic Films

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Svetlana V. Boriskina

2015-06-01

Full Text Available The properties of thermal radiation exchange between hot and cold objects can be strongly modified if they interact in the near field where electromagnetic coupling occurs across gaps narrower than the dominant wavelength of thermal radiation. Using a rigorous fluctuational electrodynamics approach, we predict that ultra-thin films of plasmonic materials can be used to dramatically enhance near-field heat transfer. The total spectrally integrated film-to-film heat transfer is over an order of magnitude larger than between the same materials in bulk form and also exceeds the levels achievable with polar dielectrics such as SiC. We attribute this enhancement to the significant spectral broadening of radiative heat transfer due to coupling between surface plasmon polaritons (SPPs on both sides of each thin film. We show that the radiative heat flux spectrum can be further shaped by the choice of the substrate onto which the thin film is deposited. In particular, substrates supporting surface phonon polaritons (SPhP strongly modify the heat flux spectrum owing to the interactions between SPPs on thin films and SPhPs of the substrate. The use of thin film phase change materials on polar dielectric substrates allows for dynamic switching of the heat flux spectrum between SPP-mediated and SPhP-mediated peaks.

Initial damage in human interphase chromosomes from alpha particles with linear energy transfers relevant to radon exposure

International Nuclear Information System (INIS)

Loucas, B.D.; Geard, C.R.

1994-01-01

To determine the efficiency at which α particles at LETs chosen to simulate exposure to radon progeny break chromosomes, the premature chromosome condensation technique was used to measure breaks soon after irradiation. Noncycling human fibroblasts were irradiated with graded doses of monoenergetic α particles accelerated to produce LETs of 90, 120, 150, 180 and 200 keV/pm at the midpoint of the cell nuclei. Premature chromosome condensation was initiated immediately after irradiation and cells were scored for the total number of prematurely condensed chromosomes and fragments per cell. Similar experiments were conducted with 250 kVp X rays for comparison. Irradiation with α particles produced 8.6 to 13.1 excess fragments per gray, while X rays produced 5.8 excess fragments, resulting in RBEs around 2. Calculations of the number of breaks produced on average by a single particle traversal of a cell nucleus indicated that at the LETs tested more than one break was produced by each traversal, the maximum being that produced by 180 keV/μm α particles. When chromosome aberrations are scored at metaphase after high-LET irradiation, RBEs considerably greater than those recorded here have been reported. These results showing relatively small differences in initial break levels for α particles in the LET range of the radon progeny relative to X rays indicate that the great aberration frequencies are not due principally to an increase in breakage efficiency, but interactions between breaks along the same particle track are important. 16 refs., 4 figs

Assay for Human Rad51-Mediated DNA Displacement Loop Formation

OpenAIRE

sprotocols

2014-01-01

Authors: Steven Raynard and Patrick Sung Corresponding author ([]()) ### INTRODUCTION Homologous recombination is an important mechanism for the repair of damaged chromosomes, for preventing the demise of damaged replication forks, and for several other aspects of chromosome metabolism and maintenance. The homologous recombination reaction is mediated by the Rad51 recombinase. In the presence of ATP, Rad51 polymerizes on single-stranded D...

Mapping EBNA-1 Domains Involved in Binding to Metaphase Chromosomes

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Marechal, Vincent; Dehee, Axelle; Chikhi-Brachet, Roxane; Piolot, Tristan; Coppey-Moisan, Maité; Nicolas, Jean-Claude

1999-01-01

The Epstein-Barr virus (EBV) genome can persist in dividing human B cells as multicopy circular episomes. Viral episomes replicate in synchrony with host cell DNA and are maintained at a relatively constant copy number for a long time. Only two viral elements, the replication origin OriP and the EBNA-1 protein, are required for the persistence of viral genomes during latency. EBNA-1 activates OriP during the S phase and may also contribute to the partition and/or retention of viral genomes during mitosis. Indeed, EBNA-1 has been shown to interact with mitotic chromatin. Moreover, viral genomes are noncovalently associated with metaphase chromosomes. This suggests that EBNA-1 may facilitate the anchorage of viral genomes on cellular chromosomes, thus ensuring proper partition and retention. In the present paper, we have investigated the chromosome-binding activity of EBV EBNA-1, herpesvirus papio (HVP) EBNA-1, and various derivatives of EBV EBNA-1, fused to a variant of the green fluorescent protein. The results show that binding to metaphase chromosomes is a common property of EBV and HVP EBNA-1. Further studies indicated that at least three independent domains (CBS-1, -2, and -3) mediate EBNA-1 binding to metaphase chromosomes. In agreement with the anchorage model, two of these domains mapped to a region that has been previously demonstrated to be required for the long-term persistence of OriP-containing plasmids. PMID:10196336

Vibrio chromosome-specific families

DEFF Research Database (Denmark)

Lukjancenko, Oksana; Ussery, David

2014-01-01

We have compared chromosome-specific genes in a set of 18 finished Vibrio genomes, and, in addition, also calculated the pan- and core-genomes from a data set of more than 250 draft Vibrio genome sequences. These genomes come from 9 known species and 2 unknown species. Within the finished...... chromosomes, we find a core set of 1269 encoded protein families for chromosome 1, and a core of 252 encoded protein families for chromosome 2. Many of these core proteins are also found in the draft genomes (although which chromosome they are located on is unknown.) Of the chromosome specific core protein...... families, 1169 and 153 are uniquely found in chromosomes 1 and 2, respectively. Gene ontology (GO) terms for each of the protein families were determined, and the different sets for each chromosome were compared. A total of 363 different "Molecular Function" GO categories were found for chromosome 1...

Giemsa C-banding of Barley Chromosomes. IV. Chromosomal Constitution of Autotetraploid Barley

DEFF Research Database (Denmark)

Linde-Laursen, Ib

1984-01-01

The progeny of an autotetraploid barley plant (C1) consisted of 45 tetraploids and 33 aneuploids. Giemsa C-banding was used to identify each of the chromosomes in 20 euploid and 31 aneuploid C2--seedlings, and in 11 C3--offspring of aneuploid C2--plants. The euploid C2--seedlings all had four...... homologues of each of the chromosomes. The aneuploid C2--seedlings were fairly equally distributed on hypo-and hyperploids, and on the seven chromosome groups. This suggests that a particular chromosome is lost or gained at random in gametes and embryos. The 11 C3--seedlings comprised seven true euploids......, one seedling with 2n=28 having an extra chromosome 6 and missing one chromosome 3, and three seedlings with 2n=29. The chromosomal composition of aneuploid C3--seedlings did not reflect that of their aneuploid C2--parents with respect to missing or extra chromosomes. Two hypohexaploid C2--seedlings...

Telomere healing following DNA polymerase arrest-induced breakages is likely the main mechanism generating chromosome 4p terminal deletions.

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Hannes, Femke; Van Houdt, Jeroen; Quarrell, Oliver W; Poot, Martin; Hochstenbach, Ron; Fryns, Jean-Pierre; Vermeesch, Joris R

2010-12-01

Constitutional developmental disorders are frequently caused by terminal chromosomal deletions. The mechanisms and/or architectural features that might underlie those chromosome breakages remain largely unexplored. Because telomeres are the vital DNA protein complexes stabilizing linear chromosomes against chromosome degradation, fusion, and incomplete replication, those terminal-deleted chromosomes acquired new telomeres either by telomere healing or by telomere capture. To unravel the mechanisms leading to chromosomal breakage and healing, we sequenced nine chromosome 4p terminal deletion boundaries. A computational analysis of the breakpoint flanking region, including 12 previously published pure terminal breakage sites, was performed in order to identify architectural features that might be involved in this process. All terminal 4p truncations were likely stabilized by telomerase-mediated telomere healing. In the majority of breakpoints multiple genetic elements have a potential to induce secondary structures and an enrichment in replication stalling site motifs were identified. These findings suggest DNA replication stalling-induced chromosome breakage during early development is the first mechanistic step leading toward terminal deletion syndromes. © 2010 Wiley-Liss, Inc.

Allocation of the S-genome chromosomes of Aegilops variabilis Eig. carrying powdery mildew resistance in triticale (× Triticosecale Wittmack).

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Kwiatek, M; Belter, J; Majka, M; Wiśniewska, H

2016-03-01

It has been hypothesized that the powdery mildew adult plant resistance (APR) controlled by the Pm13 gene in Aegilops longissima Schweinf. & Muschl. (S(l)S(l)) has been evolutionary transferred to Aegilops variabilis Eig. (UUSS). The molecular marker analysis and the visual evaluation of powdery mildew symptoms in Ae. variabilis and the Ae. variabilis × Secale cereale amphiploid forms (2n = 6x = 42, UUSSRR) showed the presence of product that corresponded to Pm13 marker and the lower infection level compared to susceptible model, respectively. This study also describes the transfer of Ae. variabilis Eig. (2n = 4x = 28, U(v)U(v)S(v)S(v)) chromosomes, carrying powdery mildew resistance, into triticale (× Triticosecale Wittm., 2n = 6x = 42, AABBRR) using Ae. variabilis × S. cereale amphiploid forms. The individual chromosomes of Ae. variabilis, triticale 'Lamberto' and hybrids were characterized by genomic and fluorescence in situ hybridization (GISH/FISH). The chromosome configurations of obtained hybrid forms were studied at first metaphase of meiosis of pollen mother cells (PMCs) using GISH. The statistical analysis showed that the way of S-genome chromosome pairing and transmission to subsequent hybrid generations was diploid-like and had no influence on chromosome pairing of triticale chromosomes. The cytogenetic study of hybrid forms were supported by the marker-assisted selection using Pm13 marker and visual evaluation of natural infection by Blumeria graminis, that allowed to select the addition or substitution lines of hybrids carrying chromosome 3S(v) which were tolerant to the powdery mildew infection.

Involvement of β-carbonic anhydrase (β-CA) genes in bacterial genomic islands and horizontal transfer to protists.

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Zolfaghari Emameh, Reza; Barker, Harlan R; Hytönen, Vesa P; Parkkila, Seppo

2018-05-25

Genomic islands (GIs) are a type of mobile genetic element (MGE) that are present in bacterial chromosomes. They consist of a cluster of genes which produce proteins that contribute to a variety of functions, including, but not limited to, regulation of cell metabolism, anti-microbial resistance, pathogenicity, virulence, and resistance to heavy metals. The genes carried in MGEs can be used as a trait reservoir in times of adversity. Transfer of genes using MGEs, occurring outside of reproduction, is called horizontal gene transfer (HGT). Previous literature has shown that numerous HGT events have occurred through endosymbiosis between prokaryotes and eukaryotes.Beta carbonic anhydrase (β-CA) enzymes play a critical role in the biochemical pathways of many prokaryotes and eukaryotes. We have previously suggested horizontal transfer of β-CA genes from plasmids of some prokaryotic endosymbionts to their protozoan hosts. In this study, we set out to identify β-CA genes that might have transferred between prokaryotic and protist species through HGT in GIs. Therefore, we investigated prokaryotic chromosomes containing β-CA-encoding GIs and utilized multiple bioinformatics tools to reveal the distinct movements of β-CA genes among a wide variety of organisms. Our results identify the presence of β-CA genes in GIs of several medically and industrially relevant bacterial species, and phylogenetic analyses reveal multiple cases of likely horizontal transfer of β-CA genes from GIs of ancestral prokaryotes to protists. IMPORTANCE The evolutionary process is mediated by mobile genetic elements (MGEs), such as genomic islands (GIs). A gene or set of genes in the GIs are exchanged between and within various species through horizontal gene transfer (HGT). Based on the crucial role that GIs can play in bacterial survival and proliferation, they were introduced as the environmental- and pathogen-associated factors. Carbonic anhydrases (CAs) are involved in many critical

Electrochemical ion transfer mediated by a lipophilic Os(ii)/Os(iii) dinonyl bipyridyl probe incorporated in thin film membranes.

Science.gov (United States)

Jansod, Sutida; Wang, Lu; Cuartero, Maria; Bakker, Eric

2017-09-28

A new lipophilic dinonyl bipyridyl Os(ii)/Os(iii) complex successfully mediates ion transfer processes across voltammetric thin membranes. An added lipophilic cation-exchanger may impose voltammetric anion or cation transfer waves of Gaussian shape that are reversible and repeatable. The peak potential is found to shift with the ion concentration in agreement with the Nernst equation. The addition of tridodecylmethylammonium nitrate to the polymeric film dramatically reduces the peak separation from 240 mV to 65 mV, and the peak width to a near-theoretical value of 85 mV, which agrees with a surface confined process. It is suggested that the cationic additive serves as a phase transfer catalyst.

Discrimination of chromosome by autoradiography

International Nuclear Information System (INIS)

Masubuchi, Masanori

1975-01-01

This paper describes discrimination of chromosome by autoradiography. In this method, the difference in DNA synthetic phase between each chromosome was used as a standard, and the used chromosome was in metaphase, as morphological characteristics were markedly in this phase. Cell cycle and autoradiography with 3 H-thymidine were also examined. In order to discriminate chromosome by autoradiography, it was effective to utilize the labelled pattern in late DNA synthetic phase, where asynchronous replication of chromosome appeared most obviously. DNA synthesis in chromosome was examined in each DNA synthetic phase by culturing the chromosome after the treatment with 3 H-thymidine and altering the time to prepare chromosome specimen. Discrimination of chromosome in plants and animals by autoradiography was also mentioned. It was noticed as a structural and functional discrimination of chromosome to observe amino acid uptake into chromosome protein and to utilize the difference in labelled pattern between the sites of chromosome. (K. Serizawa)

The Y chromosome of the Atelidae family (Platyrrhini): study by chromosome microdissection.

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Gifalli-Iughetti, C; Koiffmann, C P

2009-01-01

In order to study the intergeneric variability of the Y chromosome, we describe the hybridization of the Y chromosome of Brachytelesarachnoides, obtained by microdissection, to metaphases of Atelesbelzebuthmarginatus, Lagothrixlagothricha, and Alouatta male specimens. Brachytelesarachnoides (Atelinae) has 62 chromosomes and a very small Y chromosome. Our results showed that the Brachytelesarachnoides Y chromosome probe hybridized to Lagothrixlagothricha metaphases yielding one hybridization signal on only the tiny Y chromosome, and when hybridized with Atelesbelzebuthmarginatus metaphases it yielded one hybridization signal on two thirds of the small acrocentric Y chromosome. However, no hybridization signal was observed in Alouatta metaphases (subfamily Alouattinae), a closely related genus in the Atelidae family. Furthermore, our data support a close phylogenetic relationship among Brachyteles, Ateles, and Lagothrix and their placement in the Atelinae subfamily, but exclude Alouatta from this group indicating its placement as basal to this group. Copyright 2009 S. Karger AG, Basel.

Cobamide-mediated enzymatic reductive dehalogenation via long-range electron transfer.

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Kunze, Cindy; Bommer, Martin; Hagen, Wilfred R; Uksa, Marie; Dobbek, Holger; Schubert, Torsten; Diekert, Gabriele

2017-07-03

The capacity of metal-containing porphyrinoids to mediate reductive dehalogenation is implemented in cobamide-containing reductive dehalogenases (RDases), which serve as terminal reductases in organohalide-respiring microbes. RDases allow for the exploitation of halogenated compounds as electron acceptors. Their reaction mechanism is under debate. Here we report on substrate-enzyme interactions in a tetrachloroethene RDase (PceA) that also converts aryl halides. The shape of PceA's highly apolar active site directs binding of bromophenols at some distance from the cobalt and with the hydroxyl substituent towards the metal. A close cobalt-substrate interaction is not observed by electron paramagnetic resonance spectroscopy. Nonetheless, a halogen substituent para to the hydroxyl group is reductively eliminated and the path of the leaving halide is traced in the structure. Based on these findings, an enzymatic mechanism relying on a long-range electron transfer is concluded, which is without parallel in vitamin B 12 -dependent biochemistry and represents an effective mode of RDase catalysis.

Features of genomic organization in a nucleotide-resolution molecular model of the Escherichia coli chromosome.

Science.gov (United States)

Hacker, William C; Li, Shuxiang; Elcock, Adrian H

2017-07-27

We describe structural models of the Escherichia coli chromosome in which the positions of all 4.6 million nucleotides of each DNA strand are resolved. Models consistent with two basic chromosomal orientations, differing in their positioning of the origin of replication, have been constructed. In both types of model, the chromosome is partitioned into plectoneme-abundant and plectoneme-free regions, with plectoneme lengths and branching patterns matching experimental distributions, and with spatial distributions of highly-transcribed chromosomal regions matching recent experimental measurements of the distribution of RNA polymerases. Physical analysis of the models indicates that the effective persistence length of the DNA and relative contributions of twist and writhe to the chromosome's negative supercoiling are in good correspondence with experimental estimates. The models exhibit characteristics similar to those of 'fractal globules,' and even the most genomically-distant parts of the chromosome can be physically connected, through paths combining linear diffusion and inter-segmental transfer, by an average of only ∼10 000 bp. Finally, macrodomain structures and the spatial distributions of co-expressed genes are analyzed: the latter are shown to depend strongly on the overall orientation of the chromosome. We anticipate that the models will prove useful in exploring other static and dynamic features of the bacterial chromosome. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Chromosomal instability can be induced by the formation of breakage-prone chromosome rearrangement junctions

International Nuclear Information System (INIS)

Allen, R.N.; Ritter, L.; Moore, S.R.; Grosovsky, A.J.

2003-01-01

Full text: Studies in our lab have led to the hypothesis that chromosomal rearrangements can generate novel breakage-prone sites, resulting in chromosomal instability acting predominantly in cis. For example, specific breakage of large blocks of centromeric region heterochromatin on chromosome 16q by treatment with 2,6-diaminopurine (DAP) is associated with repeated rearrangement of chromosome 16q during outgrowth of DAP-treated clones, thereby establishing a link between the initial site of damage and the occurrence of persistent chromosomal instability. Similarly, karyotypic analysis of gamma ray induced instability demonstrated that chromosomal rearrangements in sub-clones were significantly clustered near the site of previously identified chromosomal rearrangement junctions in unstable parental clones. This study investigates the hypothesis that integration of transfected sequences into host chromosomes could create breakage-prone junction regions and persistent genomic instability without exposure to DNA-damage agents. These junctions may mimic the unstable chromosomal rearrangements induced by DAP or radiation, and thus provide a test of the broader hypothesis that instability can to some extent be attributed to the formation of novel chromosomal breakage hot spots. These experiments were performed using human-hamster hybrid AL cells containing a single human chromosome 11, which was used to monitor instability in a chromosomal painting assay. AL cells were transfected with a 2.5 Kb fragment containing multiple copies of the 180 bp human alpha heterochromatic repeat, which resulted in chromosomal instability in 41% of the transfected clones. Parallel exposure to gamma-radiation resulted in a similar level of chromosomal instability, although control transfections with plasmid alone did not lead to karyotypic instability. Chromosomal instability induced by integration of alpha heterochromatic repeats was also frequently associated with delayed reproductive

Micromechanics of human mitotic chromosomes

International Nuclear Information System (INIS)

Sun, Mingxuan; Kawamura, Ryo; Marko, John F

2011-01-01

Eukaryote cells dramatically reorganize their long chromosomal DNAs to facilitate their physical segregation during mitosis. The internal organization of folded mitotic chromosomes remains a basic mystery of cell biology; its understanding would likely shed light on how chromosomes are separated from one another as well as into chromosome structure between cell divisions. We report biophysical experiments on single mitotic chromosomes from human cells, where we combine micromanipulation, nano-Newton-scale force measurement and biochemical treatments to study chromosome connectivity and topology. Results are in accord with previous experiments on amphibian chromosomes and support the 'chromatin network' model of mitotic chromosome structure. Prospects for studies of chromosome-organizing proteins using siRNA expression knockdowns, as well as for differential studies of chromosomes with and without mutations associated with genetic diseases, are also discussed

Effect of met-enkephalin on chromosomal aberrations in the lymphocytes of the peripheral blood of patients with multiple sclerosis.

Science.gov (United States)

Rakanović-Todić, Maida; Burnazović-Ristić, Lejla; Ibrulj, Slavka; Mulbegović, Nedžad

2014-05-01

Endogenious opiod met-enkephalin throughout previous research manifested cytoprotective and anti-inflammatory effects. Previous research suggests that met-enkephalin has cytogenetic effects. Reducement in the frequency of structural chromosome aberrations as well as a suppressive effect on lymphocyte cell cycle is found. It also reduces apoptosis in the blood samples of the patients with immune-mediated diseases. Met-enkephalin exerts immunomodulatory properties and induces stabilization of the clinical condition in patients with multiple Sclerosis (MS). The goal of the present research was to evaluate met-enkephalin in vitro effects on the number and type of chromosome aberrations in the peripheral blood lymphocytes of patients with MS. Our research detected disappearance of ring chromosomes and chromosome fragmentations in the cultures of the peripheral blood lymphocytes treated with met-enkephalin (1.2 μg/mL). However, this research did not detect any significant effects of met-enkephalin on the reduction of structural chromosome aberrations and disappearance of dicentric chromosomes. Chromosomes with the greatest percent of inclusion in chromosome aberrations were noted as: chromosome 1, chromosome 2 and chromosome 9. Additionally, we confirmed chromosome 14 as the most frequently included in translocations. Furthermore, met-enkephalin effects on the increase of the numerical aberrations in both concentrations applied were detected. Those findings should be interpreted cautiously and more research in this field should be conducted.

Effect of met-enkephalin on chromosomal aberrations in the lymphocytes of the peripheral blood of patients with multiple sclerosis

Directory of Open Access Journals (Sweden)

Maida Rakanović-Todić

2014-05-01

Full Text Available Endogenious opiod met-enkephalin throughout previous research manifested cytoprotective and anti-inflammatory effects. Previous research suggests that met-enkephalin has cytogenetic effects. Reducement in the frequency of structural chromosome aberrations as well as a suppressive effect on lymphocyte cell cycle is found. It also reduces apoptosis in the blood samples of the patients with immune-mediated diseases. Met-enkephalin exerts immunomodulatory properties and induces stabilization of the clinical condition in patients with multiple Sclerosis (MS. The goal of the present research was to evaluate met-enkephalin in vitro effects on the number and type of chromosome aberrations in the peripheral blood lymphocytes of patients with MS. Our research detected disappearance of ring chromosomes and chromosome fragmentations in the cultures of the peripheral blood lymphocytes treated with met-enkephalin (1.2 μg/mL. However, this research did not detect any significant effects of met-enkephalin on the reduction of structural chromosome aberrations and disappearance of dicentric chromosomes. Chromosomes with the greatest percent of inclusion in chromosome aberrations were noted as: chromosome 1, chromosome 2 and chromosome 9. Additionally, we confirmed chromosome 14 as the most frequently included in translocations. Furthermore, met-enkephalin effects on the increase of the numerical aberrations in both concentrations applied were detected. Those findings should be interpreted cautiously and more research in this field should be conducted. 

Transfer of unstable chromosomal aberrations in human peripheral lymphocytes at cell division and their significance for the aberration frequency

International Nuclear Information System (INIS)

Stephan, G.; Chang Tsangpi.

1986-04-01

In 48 h cultures, the fraction of human lymphocytes in 2nd mitosis was found to be between 0 and 42.5% (mean value 8.7%). The X-ray exposure from irradiating with 2 Gy resulted in a cell cycle delay which varied from donor to donor. A loss of nearly 50% of dicentric chromosomes and acentric fragments from unstable chromosomes occurred at cell division, while centric rings were not impeded. When dicentric chromosomes, or acentric fragments are found in 2nd mitosis, they show a characteristic differential staining, which means that chromatides at cell division fall free and are replicated in daughter cells. When plotting dose effect curves of dicentric chromosomes, up to 20% of 2nd mitosis fractions have little influence on the aberration rate. This may be additionally verified as part of the 'biological dosimetry' in a person with 24% of 2nd mitosis. When the rates of dicentric chromosomes exclusively evaluated from 1st mitosis after irradiation with 2.0 Gy were related to the donors age, no age-dependent sensitivity to radiation could be observed. Aberration rates which deviate from person to person are comparable to the results achieved by conventional staining methods. (orig./MG) [de

Shelterin Protects Chromosome Ends by Compacting Telomeric Chromatin

Science.gov (United States)

Bandaria, Jigar N.; Qin, Peiwu; Berk, Veysel; Chu, Steven; Yildiz, Ahmet

2016-01-01

SUMMARY Telomeres, repetitive DNA sequences at chromosome ends, are shielded against the DNA damage response (DDR) by the shelterin complex. To understand how shelterin protects telomere ends, we investigated the structural organization of telomeric chromatin in human cells using super-resolution microscopy. We found that telomeres form compact globular structures through a complex network of interactions between shelterin subunits and telomeric DNA, and not by DNA methylation, histone deacetylation or histone trimethylation at telomeres and subtelomeric regions. Mutations that abrogate shelterin assembly or removal of individual subunits from telomeres cause up to a 10-fold increase in telomere volume. Decompacted telomeres become more accessible to telomere-associated proteins and accumulate DDR signals. Recompaction of telomeric chromatin using an orthogonal method displaces DDR signals from telomeres. These results reveal the chromatin remodeling activity of shelterin and demonstrate that shelterin-mediated compaction of telomeric chromatin provides robust protection of chromosome ends against the DDR machinery. PMID:26871633

Ten alien chromosome additions of Gossypium hirsutum-Gossypium bickii developed by integrative uses of GISH and species-specific SSR markers.

Science.gov (United States)

Tang, Dong; Feng, Shouli; Li, Sai; Chen, Yu; Zhou, Baoliang

2018-03-27

Gossypium bickii: (2n = 26, G 1 G 1 ), a wild diploid cotton, carries many favourable traits. However, these favourable traits cannot be directly transferred into G. hirsutum (2n = 52, AADD) cultivars due to the differences in genomes. Monosomic alien addition lines (MAALs) are considered an invaluable tool for the introgression of genes of interest from wild relatives into cultivated crops. In this study, the G. hirsutum-G. bickii amphidiploid (2n = 78, AADDG 1 G 1 ) was backcrossed with G. hirsutum to develop alien additions containing individual G. bickii chromosomes in a G. hirsutum background. Genomic in situ hybridization was employed to detect the number of alien chromosomes added to the backcross progenies. A total of 183 G. bickii-specific DNA markers were developed to discriminate the identities of the G. bickii chromosomes added to G. hirsutum and assess the alien chromosome transmissibility. Chromosomes 4G b and 13G b showed the highest transmissibility, while chromosomes 1G b , 7G b and 11G b showed the lowest. Ten of the 13 possible G. hirsutum-G. bickii MAALs were isolated and characterized, which will lay the foundation for transferring resistance genes of G. bickii into G. hirsutum, as well as for gene assignment, physical mapping, and selective isolation and mapping of cDNAs for particular G. bickii chromosomes. The strategies of how to use MAALs to develop varieties with the trait of interest from wild species (such as glanded plant-glandless seed) were proposed and discussed.

Complementation of a threonine dehydratase-deficient Nicotiana plumbaginifolia mutant after Agrobacterium tumefaciens-mediated transfer of the Saccharomyces cerevisiae ILV1 gene.

OpenAIRE

Colau, D; Negrutiu, I; Van Montagu, M; Hernalsteens, J P

1987-01-01

The Saccharomyces cerevisiae ILV1 gene, encoding threonine dehydratase (EC 4.2.1.16) was fused to the transferred DNA nopaline synthase promoter and the 3' noncoding region of the octopine synthase gene. It was introduced, by Agrobacterium tumefaciens-mediated gene transfer, into an isoleucine-requiring Nicotiana plumbaginifolia auxotroph deficient in threonine dehydratase. Functional complementation by the ILV1 gene product was demonstrated by the selection of several transformed lines on a ...

Comprehensive preimplantation genetic screening and sperm deoxyribonucleic acid fragmentation from three males carrying balanced chromosome rearrangements.

Science.gov (United States)

Ramos, Laia; Daina, Gemma; Del Rey, Javier; Ribas-Maynou, Jordi; Fernández-Encinas, Alba; Martinez-Passarell, Olga; Boada, Montserrat; Benet, Jordi; Navarro, Joaquima

2015-09-01

To assess whether preimplantation genetic screening can successfully identify cytogenetically normal embryos in couples carrying balanced chromosome rearrangements in addition to increased sperm DNA fragmentation. Comprehensive preimplantation genetic screening was performed on three couples carrying chromosome rearrangements. Sperm DNA fragmentation was assessed for each patient. Academic center. One couple with the male partner carrying a chromosome 2 pericentric inversion and two couples with the male partners carrying a Robertsonian translocation (13:14 and 14:21, respectively). A single blastomere from each of the 18 cleavage-stage embryos obtained was analysed by metaphase comparative genomic hybridization. Single- and double-strand sperm DNA fragmentation was determined by the alkaline and neutral Comet assays. Single- and double-strand sperm DNA fragmentation values and incidence of chromosome imbalances in the blastomeres were analyzed. The obtained values of single-strand sperm DNA fragmentation were between 47% and 59%, and the double-strand sperm DNA fragmentation values were between 43% and 54%. No euploid embryos were observed in the couple showing the highest single-strand sperm DNA fragmentation. However, euploid embryos were observed in the other two couples: embryo transfer was performed, and pregnancy was achieved by the couple showing the lowest sperm DNA fragmentation values. Preimplantation genetic screening enables the detection of euploid embryos in couples affected by balanced chromosome rearrangements and increased sperm DNA fragmentation. Even though sperm DNA fragmentation may potentially have clinical consequences on fertility, comprehensive preimplantation genetic screening allows for the identification and transfer of euploid embryos. Copyright © 2015. Published by Elsevier Inc.

Regulation of CD95 expression and CD95-mediated cell death by interferon-gamma in acute lymphoblastic leukemia with chromosomal translocation t(4;11).

Science.gov (United States)

Dörrie, J; Schuh, W; Keil, A; Bongards, E; Greil, J; Fey, G H; Zunino, S J

1999-10-01

The regulatory effects of IFNgamma on CD95 expression and CD95-mediated cell death were investigated in three high-risk pro-B acute lymphoblastic leukemia (ALL) lines that carry the chromosomal translocation t(4;11)(q21;q23). These leukemias are characteristically refractory to conventional chemotherapeutic treatments operating through the induction of apoptosis. However, the mechanisms leading to increased cell survival and resistance to cell death in these leukemias are largely unknown. Interferon-gamma (IFNgamma), a potent inhibitor of hematopoiesis, acts in part by upregulating CD95 and sensitizing cells to CD95-induced apoptosis. The t(4;11) lines SEM, RS4;11, and MV4;11 expressed low levels of CD95, but were completely resistant to CD95-mediated death. Addition of IFNgamma markedly upregulated CD95 expression in SEM (8-9-fold), RS4;11 (2-3-fold), and MV4;11 (2-3-fold) lines. However, after treatment with IFNgamma, only an 11% increase in sensitivity to CD95-mediated cell death was observed in SEM cells, whereas RS4;11 and MV4;11 cells remained resistant. Cycloheximide, but not actinomycin D or brefeldin A, increased CD95-specific cell death only in IFNgamma-treated RS4;11 cells by approximately 12%. Abundant levels of Bcl-2 and Bcl-XL, known to inhibit CD95-signaling in some cells, were present suggesting a possible role for both molecules in the resistance to CD95-mediated cell death. Resistance of the leukemic blasts to CD95-mediated cell death and the failure of IFNgamma to substantially sensitize the CD95-signaling pathway may contribute to the highly malignant phenotype of pro-B ALL with translocation t(4;11).

Hominoid chromosomal rearrangements on 17q map to complex regions of segmental duplication.

Science.gov (United States)

Cardone, Maria Francesca; Jiang, Zhaoshi; D'Addabbo, Pietro; Archidiacono, Nicoletta; Rocchi, Mariano; Eichler, Evan E; Ventura, Mario

2008-01-01

Chromosomal rearrangements, such as translocations and inversions, are recurrent phenomena during evolution, and both of them are involved in reproductive isolation and speciation. To better understand the molecular basis of chromosome rearrangements and their part in karyotype evolution, we have investigated the history of human chromosome 17 by comparative fluorescence in situ hybridization (FISH) and sequence analysis. Human bacterial artificial chromosome/p1 artificial chromosome probes spanning the length of chromosome 17 were used in FISH experiments on great apes, Old World monkeys and New World monkeys to study the evolutionary history of this chromosome. We observed that the macaque marker order represents the ancestral organization. Human, chimpanzee and gorilla homologous chromosomes differ by a paracentric inversion that occurred specifically in the Homo sapiens/Pan troglodytes/Gorilla gorilla ancestor. Detailed analyses of the paracentric inversion revealed that the breakpoints mapped to two regions syntenic to human 17q12/21 and 17q23, both rich in segmental duplications. Sequence analyses of the human and macaque organization suggest that the duplication events occurred in the catarrhine ancestor with the duplication blocks continuing to duplicate or undergo gene conversion during evolution of the hominoid lineage. We propose that the presence of these duplicons has mediated the inversion in the H. sapiens/P. troglodytes/G. gorilla ancestor. Recently, the same duplication blocks have been shown to be polymorphic in the human population and to be involved in triggering microdeletion and duplication in human. These results further support a model where genomic architecture has a direct role in both rearrangement involved in karyotype evolution and genomic instability in human.

Efficient anchoring of alien chromosome segments introgressed into bread wheat by new Leymus racemosus genome-based markers.

Science.gov (United States)

Edet, Offiong Ukpong; Kim, June-Sik; Okamoto, Masanori; Hanada, Kousuke; Takeda, Tomoyuki; Kishii, Masahiro; Gorafi, Yasir Serag Alnor; Tsujimoto, Hisashi

2018-03-27

The tertiary gene pool of bread wheat, to which Leymus racemosus belongs, has remained underutilized due to the current limited genomic resources of the species that constitute it. Continuous enrichment of public databases with useful information regarding these species is, therefore, needed to provide insights on their genome structures and aid successful utilization of their genes to develop improved wheat cultivars for effective management of environmental stresses. We generated de novo DNA and mRNA sequence information of L. racemosus and developed 110 polymorphic PCR-based markers from the data, and to complement the PCR markers, DArT-seq genotyping was applied to develop additional 9990 SNP markers. Approximately 52% of all the markers enabled us to clearly genotype 22 wheat-L. racemosus chromosome introgression lines, and L. racemosus chromosome-specific markers were highly efficient in detailed characterization of the translocation and recombination lines analyzed. A further analysis revealed remarkable transferability of the PCR markers to three other important Triticeae perennial species: L. mollis, Psathyrostachys huashanica and Elymus ciliaris, indicating their suitability for characterizing wheat-alien chromosome introgressions carrying chromosomes of these genomes. The efficiency of the markers in characterizing wheat-L. racemosus chromosome introgression lines proves their reliability, and their high transferability further broadens their scope of application. This is the first report on sequencing and development of markers from L. racemosus genome and the application of DArT-seq to develop markers from a perennial wild relative of wheat, marking a paradigm shift from the seeming concentration of the technology on cultivated species. Integration of these markers with appropriate cytogenetic methods would accelerate development and characterization of wheat-alien chromosome introgression lines.

Interphase Chromosome Profiling: A Method for Conventional Banded Chromosome Analysis Using Interphase Nuclei.

Science.gov (United States)

Babu, Ramesh; Van Dyke, Daniel L; Dev, Vaithilingam G; Koduru, Prasad; Rao, Nagesh; Mitter, Navnit S; Liu, Mingya; Fuentes, Ernesto; Fuentes, Sarah; Papa, Stephen

2018-02-01

- Chromosome analysis on bone marrow or peripheral blood samples fails in a small proportion of attempts. A method that is more reliable, with similar or better resolution, would be a welcome addition to the armamentarium of the cytogenetics laboratory. - To develop a method similar to banded metaphase chromosome analysis that relies only on interphase nuclei. - To label multiple targets in an equidistant fashion along the entire length of each chromosome, including landmark subtelomere and centromere regions. Each label so generated by using cloned bacterial artificial chromosome probes is molecularly distinct with unique spectral characteristics, so the number and position of the labels can be tracked to identify chromosome abnormalities. - Interphase chromosome profiling (ICP) demonstrated results similar to conventional chromosome analysis and fluorescence in situ hybridization in 55 previously studied cases and obtained useful ICP chromosome analysis results on another 29 cases in which conventional methods failed. - ICP is a new and powerful method to karyotype peripheral blood and bone marrow aspirate preparations without reliance on metaphase chromosome preparations. It will be of particular value for cases with a failed conventional analysis or when a fast turnaround time is required.

Exploring the Critical Role of Motivation to Transfer in the Training Transfer Process

Science.gov (United States)

Grohmann, Anna; Beller, Johannes; Kauffeld, Simone

2014-01-01

The present study aims at exploring the critical role of motivation to transfer within the training transfer process. In a sample of N?=?252 employees of one industrial company, one peer rating and several self-ratings of transfer were used to investigate the mediating role of motivation to transfer in the relationship between training…

Lipid-mediated glial cell line-derived neurotrophic factor gene transfer to cultured porcine ventral mesencephalic tissue

DEFF Research Database (Denmark)

Bauer, Matthias; Meyer, Morten; Brevig, Thomas

2002-01-01

Transplantation of dopaminergic ventral mesencephalic (VM) tissue into the basal ganglia of patients with Parkinson's disease (PD) shows at best moderate symptomatic relief in some of the treated cases. Experimental animal studies and clinical trials with allogenic and xenogenic pig-derived VM...... tissue grafts to PD patients indicate that one reason for the poor outcome of neural transplantation is the low survival and differentiation of grafted dopaminergic neurons. To improve dopaminergic cell survival through a gene-therapeutic approach we have established and report here results of lipid-mediated...... numbers of tyrosine hydroxylase-positive neurons in the cultured VM tissue. We conclude that lipid-mediated gene transfer employed on embryonic pig VM explant cultures is a safe and effective method to improve survival of dopaminergic neurons and may become a valuable tool to improve allo...

Extracellular Electron Transfer Mediated by Flavins in Gram-positive Bacillus sp. WS-XY1 and Yeast Pichia stipitis

International Nuclear Information System (INIS)

Wu, Song; Xiao, Yong; Wang, Lu; Zheng, Yue; Chang, Kenlin; Zheng, Zhiyong; Yang, Zhaohui; Varcoe, John R.; Zhao, Feng

2014-01-01

Extracellular electron transfer (EET) of microorganisms represents a communicative bridge between the interior and exterior of the cells. Most prior EET studies have focused on Gram-negative bacteria. However, fungi and Gram-positive bacteria, that contain dense cellular walls, have rarely been reported. Herein, two model dense cell wall microorganisms (Bacillus sp. WS-XY1 and the yeast Pichia stipitis) were identified to be electrochemically active. Further analysis indicated that the two microorganisms were able to secrete flavins to mediate their EET. The discovery, that dense cell wall containing microorganisms can undertake mediated EET, adds to the body of knowledge towards building a comprehensive understanding of biogeochemical and bioelectrical processes

Homologous recombination mediates functional recovery of dysferlin deficiency following AAV5 gene transfer.

Directory of Open Access Journals (Sweden)

William E Grose

Full Text Available The dysferlinopathies comprise a group of untreatable muscle disorders including limb girdle muscular dystrophy type 2B, Miyoshi myopathy, distal anterior compartment syndrome, and rigid spine syndrome. As with other forms of muscular dystrophy, adeno-associated virus (AAV gene transfer is a particularly auspicious treatment strategy, however the size of the DYSF cDNA (6.5 kb negates packaging into traditional AAV serotypes known to express well in muscle (i.e. rAAV1, 2, 6, 8, 9. Potential advantages of a full cDNA versus a mini-gene include: maintaining structural-functional protein domains, evading protein misfolding, and avoiding novel epitopes that could be immunogenic. AAV5 has demonstrated unique plasticity with regards to packaging capacity and recombination of virions containing homologous regions of cDNA inserts has been implicated in the generation of full-length transcripts. Herein we show for the first time in vivo that homologous recombination following AAV5.DYSF gene transfer leads to the production of full length transcript and protein. Moreover, gene transfer of full-length dysferlin protein in dysferlin deficient mice resulted in expression levels sufficient to correct functional deficits in the diaphragm and importantly in skeletal muscle membrane repair. Intravascular regional gene transfer through the femoral artery produced high levels of transduction and enabled targeting of specific muscle groups affected by the dysferlinopathies setting the stage for potential translation to clinical trials. We provide proof of principle that AAV5 mediated delivery of dysferlin is a highly promising strategy for treatment of dysferlinopathies and has far-reaching implications for the therapeutic delivery of other large genes.

Sex reversal in the mouse (Mus musculus) is caused by a recurrent nonreciprocal crossover involving the x and an aberrant y chromosome.

Science.gov (United States)

Singh, L; Jones, K W

1982-02-01

Satellite DNA (Bkm) from the W sex-determining chromosome of snakes, which is related to sequences on the mouse Y chromosome, has been used to analyze the DNA and chromosomes of sex-reversed (Sxr) XXSxr male mice. Such mice exhibit a male-specific Southern blot Bkm hybridization pattern, consistent with the presence of Y-chromosome DNA. In situ hybridization of Bkm to chromosomes of XXSxr mice shows an aberrant concentration of related sequences on the distal terminus of a large mouse chromosome. The XYSxr carrier male, however, shows a pair of small chromosomes, which are presumed to be aberrant Y derivatives. Meiosis in the XYSxr mouse involves transfer of chromatin rich in Bkm-related DNA from the Y-Y1 complex to the X distal terminus. We suggest that this event is responsible for the transmission of the Sxr trait.

Chromosomal divergence and evolutionary inferences in Rhodniini based on the chromosomal location of ribosomal genes

Directory of Open Access Journals (Sweden)

Sebastian Pita

2013-05-01

Full Text Available In this study, we used fluorescence in situ hybridisation to determine the chromosomal location of 45S rDNA clusters in 10 species of the tribe Rhodniini (Hemiptera: Reduviidae: Triatominae. The results showed striking inter and intraspecific variability, with the location of the rDNA clusters restricted to sex chromosomes with two patterns: either on one (X chromosome or both sex chromosomes (X and Y chromosomes. This variation occurs within a genus that has an unchanging diploid chromosome number (2n = 22, including 20 autosomes and 2 sex chromosomes and a similar chromosome size and genomic DNA content, reflecting a genome dynamic not revealed by these chromosome traits. The rDNA variation in closely related species and the intraspecific polymorphism in Rhodnius ecuadoriensis suggested that the chromosomal position of rDNA clusters might be a useful marker to identify recently diverged species or populations. We discuss the ancestral position of ribosomal genes in the tribe Rhodniini and the possible mechanisms involved in the variation of the rDNA clusters, including the loss of rDNA loci on the Y chromosome, transposition and ectopic pairing. The last two processes involve chromosomal exchanges between both sex chromosomes, in contrast to the widely accepted idea that the achiasmatic sex chromosomes of Heteroptera do not interchange sequences.

Effects of hormone treatment on chromosomal radiosensitivity of somatic and germ cells of Snell's dwarf mice

International Nuclear Information System (INIS)

Buul, P.P.W. van; Buul-Offers, S.C. van

1988-01-01

The X-ray induction of micronuclei and structural chromosomal aberrations was studied in bone-marrow cells of normal and dwarf mice in combination with thyroxin and/or prolactin treatment or otherwise. Hormone treatment clearly increased micronuclei induction but not chromosome breakage, suggesting that indirect effects were involved. Since no clear differences in the timing of the final stage of erythropoiesis could be found, it is likely that the indirect effects are mediated via the formation-differentiation kinetics of erythroblasts. The induction of reciprocal translocations by X-rays in stem cell spermatogonia of dwarf mice was lower than in normals and treatment with prolactin, growth hormone and/or thyroxin, did not influence the chromosomal radiosensitivity of spermatogonial cells. 19 refs.; 1 figure; 4 tabs

Fetal chromosome analysis

DEFF Research Database (Denmark)

Philip, J; Tabor, A; Bang, J

1983-01-01

The aim of the study was to investigate the rationale of the current indications for fetal chromosome analysis. 5372 women had 5423 amniocentesis performed, this group constituting a consecutive sample at the chromosome laboratory, Rigshospitalet, Copenhagen from March 1973 to September 1980 (Group...... A + B). Pregnant women 35 years of age, women who previously had a chromosomally abnormal child, families with translocation carriers or other heritable chromosomal disease, families where the father was 50 years or more and women in families with a history of Down's syndrome (group A), were compared...... to women having amniocentesis, although considered not to have any increased risk of fetal chromosome abnormality (1390 pregnancies, group B). They were also compared with 750 consecutive pregnancies in women 25-34 years of age, in whom all heritable diseases were excluded (group C). The risk of unbalanced...

Microdissection and chromosome painting of the alien chromosome in an addition line of wheat-Thinopyrum intermedium

Science.gov (United States)

The chromosome painting is an efficient tool for chromosome research. However, plant chromosome painting is relatively underdeveloped. In this study, chromosome painting was developed and used to identify alien chromosomes in TAi-27, a wheat-Thinopyrum intermedium addition line, and chromosomes of...

Microdissection and chromosome painting of the alien chromosome in an addition line of wheat--Thinopyrum intermedium.

Science.gov (United States)

Deng, Chuanliang; Bai, Lili; Fu, Shulan; Yin, Weibo; Zhang, Yingxin; Chen, Yuhong; Wang, Richard R-C; Zhang, Xiangqi; Han, Fangpu; Hu, Zanmin

2013-01-01

In this study, chromosome painting was developed and used to identify alien chromosomes in TAi-27, a wheat--Thinopyrum intermedium addition line, and the chromosomes of the three different genomes of Th. Intermedium. The smallest alien chromosome of TAi-27 was microdissected and its DNA amplified by DOP-PCR was used as a probe to hybridize with metaphase chromosomes of TAi-27 and Th. intermedium. Results showed that hybridization signals were observed in all regions of a pair of the smallest alien chromosomes and the pericentromeric area of another pair of alien chromosomes in TAi-27, indicating that the probe from microdissected chromosome is species specific. In Th. intermedium, 14 chromosomes had wide and strong hybridization signals distributed mainly on the pericentromere area and 9 chromosomes with narrow and weak signals on the pericentromere area. The remaining chromosomes displayed a very weak or no signal. Sequential FISH/GISH on Th. intermedium chromosomes using the DNAs of microdissected chromosome, Pseudoroegneria spicata (St genome) and pDbH12 (a J(s) genome specific probe) as the probes indicated that the microdissected chromosome belonged to the St genome, three genomes (J(s) , J and St) in Th. intermedium could be distinguished, in which there is no hybridization signal on J genome that is similar to the genome of Th. bessarabicum. Our results showed that the smallest alien chromosomes may represent a truncated chromosome and the repetitive sequence distribution might be similar in different chromosomes within the St genome. However, the repetitive sequence distributions are different within the J(s) genome, within a single chromosome, and among different genomes in Th. intermedium. Our results suggested that chromosome painting could be feasible in some plants and useful in detecting chromosome variation and repetitive sequence distribution in different genomes of polyploidy plants, which is helpful for understanding the evolution of different

Microdissection and Chromosome Painting of the Alien Chromosome in an Addition Line of Wheat - Thinopyrum intermedium

Science.gov (United States)

Yin, Weibo; Zhang, Yingxin; Chen, Yuhong; Wang, Richard R.-C.; Zhang, Xiangqi; Han, Fangpu; Hu, Zanmin

2013-01-01

In this study, chromosome painting was developed and used to identify alien chromosomes in TAi-27, a wheat - Thinopyrum intermedium addition line, and the chromosomes of the three different genomes of Th. Intermedium. The smallest alien chromosome of TAi-27 was microdissected and its DNA amplified by DOP-PCR was used as a probe to hybridize with metaphase chromosomes of TAi-27 and Th . intermedium . Results showed that hybridization signals were observed in all regions of a pair of the smallest alien chromosomes and the pericentromeric area of another pair of alien chromosomes in TAi-27, indicating that the probe from microdissected chromosome is species specific. In Th . intermedium , 14 chromosomes had wide and strong hybridization signals distributed mainly on the pericentromere area and 9 chromosomes with narrow and weak signals on the pericentromere area. The remaining chromosomes displayed a very weak or no signal. Sequential FISH/GISH on Th . intermedium chromosomes using the DNAs of microdissected chromosome, Pseudoroegneria spicata (St genome) and pDbH12 (a Js genome specific probe) as the probes indicated that the microdissected chromosome belonged to the St genome, three genomes (Js, J and St) in Th . intermedium could be distinguished, in which there is no hybridization signal on J genome that is similar to the genome of Th . bessarabicum . Our results showed that the smallest alien chromosomes may represent a truncated chromosome and the repetitive sequence distribution might be similar in different chromosomes within the St genome. However, the repetitive sequence distributions are different within the Js genome, within a single chromosome, and among different genomes in Th . intermedium . Our results suggested that chromosome painting could be feasible in some plants and useful in detecting chromosome variation and repetitive sequence distribution in different genomes of polyploidy plants, which is helpful for understanding the evolution of different

Retroviral-mediated transfer and expression of human β-globin genes in cultured murine and human erythroid cells

International Nuclear Information System (INIS)

Weber-Benarous, A.; Cone, R.D.; London, I.M.; Mulligan, R.C.

1988-01-01

The authors cloned human β-globin DNA sequences from a genomic library prepared from DNA isolated from the human leukemia cell line K562 and have used the retroviral vector pZip-NeoSV(X)1 to introduce a 3.0-kilobase segment encompassing the globin gene into mouse erythroleukemia cells. Whereas the endogenous K562 β-globin gene is repressed in K562 cells, when introduced into mouse erythroleukemia cells by retroviral-mediated gene transfer, the β-globin gene from K562 cells was transcribed and induced 5-20-fold after treatment of the cells with dimethyl sulfoxide. The transcripts were correctly initiated, and expression and regulation of the K562 gene were identical to the expression of a normal human β-globin gene transferred into mouse erythroleukemia cells in the same way. They have also introduced the normal human β-globin gene into K562 cells using the same retrovirus vector. SP6 analysis of the RNA isolated from the transduced cells showed that the normal β-globin gene was transcribed at a moderately high level, before or after treatment with hemin. Based on these data, they suggest that the lack of expression of the endogenous β-globin gene in K562 cells does not result from an alteration in the gene itself and may not result from a lack of factor(s) necessary for β-lobin gene transcription. Retroviral-mediated transfer of the human β-globin gene may, however, uniquely influence expression of the gene K562 cells

Designing of plant artificial chromosome (PAC) by using the Chlorella smallest chromosome as a model system.

Science.gov (United States)

Noutoshi, Y; Arai, R; Fujie, M; Yamada, T

1997-01-01

As a model for plant-type chromosomes, we have been characterizing molecular organization of the Chlorella vulgaris C-169 chromosome I. To identify chromosome structural elements including the centromeric region and replication origins, we constructed a chromosome I specific cosmid library and aligned each cosmid clones to generate contigs. So far, more than 80% of the entire chromosome I has been covered. A complete clonal physical reconstitution of chromosome I provides information on the structure and genomic organization of plant genome. We propose our strategy to construct an artificial chromosome by assembling the functional chromosome structural elements identified on Chrorella chromosome I.

Different regions of the immunoglobulin heavy-chain locus are involved in chromosomal translocations in distinct pathogenic forms of Burkitt lymphoma

Energy Technology Data Exchange (ETDEWEB)

Neri, A.; Barriga, F.; Knowles, D.M.; Magrath, I.T.; Dalla-Favera, R.

1988-04-01

The authors show that endemic (eBL), sporadic (sBL), and acquired immunodeficiency syndrome-associated (AIDS-BL) forms of Burkitt lymphoma (BL) carrying t(8; 14) chromosomal translocations display different breakpoints within the immunoglobulin heavy-chain locus (IGH) on chromosome 14. In sBL (7 out of 11) and AIDS-BL (5 out of 6), the breakpoints occurred within or near the IGH ..mu.. switch (S/sub mu/) region on chromosome 14 and within the c-myc locus (MYC) on chromosome 8. In most eBL (13 out of 16) the breakpoints were mapped within or 5' to the IGH joining J/sub H/ region on chromosome 14 and outside the MYC locus on chromosome 8. Cloning and sequencing of the (8; 14) chromosomal junctions from two eBL cell lines and one eBL biopsy sample show that the recombination do not involve IGH-specific recombination signals on chromosome 14 or homologous sequences on chromosome 8, suggesting that these events are not likely to be mediated by the same mechanisms or enzymes as in IGH rearrangements. In general, these data have implications for the timing of occurrence of chromosomal translocations during B-cell differentiation in different BL types.

Sex-chromosome anaphase movements in crane-fly spermatocytes are coordinated: ultraviolet microbeam irradiation of one kinetochore of one sex chromosome blocks the movements of both sex chromosomes

International Nuclear Information System (INIS)

Swedak, J.A.M.; Forer, A.

1987-01-01

Sex chromosomes in crane-fly spermatocytes move polewards at anaphase after the autosomes have reached the poles. We irradiated one kinetochore of one sex chromosome using an ultraviolet microbeam. When both sex chromosomes were normally oriented, irradiation of a single kinetochore permanently blocked movement of both sex chromosomes. Irradiation of non-kinetochore chromosomal regions or of spindle fibres did not block movement, or blocked movement only temporarily. We argue that ultraviolet irradiation of one kinetochore blocks movement of both sex chromosomes because of effects on a 'signal' system. Irradiation of one kinetochore of a maloriented sex chromosome did not block motion of either sex chromosome. However, irradiation of one kinetochore of a normally oriented sex chromosome permanently blocked motion of both that sex chromosome and the maloriented sex chromosome. Thus for the signal system to allow the sex chromosomes to move to the pole each sex chromosome must have one spindle fibre to each pole. (author)

Modeling Chromosomes

Science.gov (United States)

Robertson, Carol

2016-01-01

Learning about chromosomes is standard fare in biology classrooms today. However, students may find it difficult to understand the relationships among the "genome", "chromosomes", "genes", a "gene locus", and "alleles". In the simple activity described in this article, which follows the 5E approach…

Chromosomal Conditions

Science.gov (United States)

... and more. Stony Point, NY 10980 Close X Home > Complications & Loss > Birth defects & other health conditions > Chromosomal conditions Chromosomal conditions ... Disorders See also: Genetic counseling , Your family health history Last reviewed: February, 2013 ... labor & premature birth The newborn intensive care unit (NICU) Birth defects & ...

Electochemical detection of chromosome translocation

DEFF Research Database (Denmark)

Kwasny, Dorota; Dimaki, Maria; Silahtaroglu, Asli

2014-01-01

Cytogenetics is a study of the cell structure with a main focus on chromosomes content and their structure. Chromosome abnormalities, such as translocations may cause various genetic disorders and heametological malignancies. Chromosome translocations are structural rearrangements of two...... chromosomes that results in formation of derivative chromosomes with a mixed DNA sequence. The method currently used for their detection is Fluorescent In Situ Hybridization, which requires a use of expensive, fluorescently labeled probes that target the derivative chromosomes. We present here a double...... hybridization approach developed for label-free detection of the chromosome translocations. For specific translocation detection it is necessary to determine that the two DNA sequences forming a derivative chromosome are connected, which is achieved by two subsequent hybridization steps. The electrochemical...

Pathogenicity Island Cross Talk Mediated by Recombination Directionality Factors Facilitates Excision from the Chromosome.

Science.gov (United States)

Carpenter, Megan R; Rozovsky, Sharon; Boyd, E Fidelma

2015-12-14

Pathogenicity islands (PAIs) are mobile integrated genetic elements (MIGEs) that contain a diverse range of virulence factors and are essential in the evolution of pathogenic bacteria. PAIs are widespread among bacteria and integrate into the host genome, commonly at a tRNA locus, via integrase-mediated site-specific recombination. The excision of PAIs is the first step in the horizontal transfer of these elements and is not well understood. In this study, we examined the role of recombination directionality factors (RDFs) and their relationship with integrases in the excision of two PAIs essential for Vibrio cholerae host colonization: Vibrio pathogenicity island 1 (VPI-1) and VPI-2. VPI-1 does not contain an RDF, which allowed us to answer the question of whether RDFs are an absolute requirement for excision. We found that an RDF was required for efficient excision of VPI-2 but not VPI-1 and that RDFs can induce excision of both islands. Expression data revealed that the RDFs act as transcriptional repressors to both VPI-1- and VPI-2-encoded integrases. We demonstrated that the RDFs Vibrio excision factor A (VefA) and VefB bind at the attachment sites (overlapping the int promoter region) of VPI-1 and VPI-2, thus supporting this mode of integrase repression. In addition, V. cholerae RDFs are promiscuous due to their dual functions of promoting excision of both VPI-1 and VPI-2 and acting as negative transcriptional regulators of the integrases. This is the first demonstration of cross talk between PAIs mediated via RDFs which reveals the complex interactions that occur between separately acquired MIGEs. Deciphering the mechanisms of pathogenicity island excision is necessary for understanding the evolution and spread of these elements to their nonpathogenic counterparts. Such mechanistic insight would assist in predicting the mobility of uncharacterized genetic elements. This study identified extensive RDF-mediated cross talk between two nonhomologous VPIs and

Birth and death of genes linked to chromosomal inversion

Science.gov (United States)

Furuta, Yoshikazu; Kawai, Mikihiko; Yahara, Koji; Takahashi, Noriko; Handa, Naofumi; Tsuru, Takeshi; Oshima, Kenshiro; Yoshida, Masaru; Azuma, Takeshi; Hattori, Masahira; Uchiyama, Ikuo; Kobayashi, Ichizo

2011-01-01

The birth and death of genes is central to adaptive evolution, yet the underlying genome dynamics remain elusive. The availability of closely related complete genome sequences helps to follow changes in gene contents and clarify their relationship to overall genome organization. Helicobacter pylori, bacteria in our stomach, are known for their extreme genome plasticity through mutation and recombination and will make a good target for such an analysis. In comparing their complete genome sequences, we found that gain and loss of genes (loci) for outer membrane proteins, which mediate host interaction, occurred at breakpoints of chromosomal inversions. Sequence comparison there revealed a unique mechanism of DNA duplication: DNA duplication associated with inversion. In this process, a DNA segment at one chromosomal locus is copied and inserted, in an inverted orientation, into a distant locus on the same chromosome, while the entire region between these two loci is also inverted. Recognition of this and three more inversion modes, which occur through reciprocal recombination between long or short sequence similarity or adjacent to a mobile element, allowed reconstruction of synteny evolution through inversion events in this species. These results will guide the interpretation of extensive DNA sequencing results for understanding long- and short-term genome evolution in various organisms and in cancer cells. PMID:21212362

Emotional contagion of dental fear to children: the fathers' mediating role in parental transfer of fear.

Science.gov (United States)

Lara, America; Crego, Antonio; Romero-Maroto, Martin

2012-09-01

Dental fear is considered to be one of the most frequent problems in paediatric dentistry. According to literature, parents' levels of dental fear play a key role in the development of child's dental anxiety. HYPOTHESIS OR AIM: We have tried to identify the presence of emotional transmission of dental fear among family members and to analyse the different roles that mothers and fathers might play concerning the contagion of dental fear to children. We have hypothesized a key role of the father in the transfer of dental fear from mother to child. A questionnaire-based survey (Children's Fear Survey Schedule-Dental Subscale) has been distributed among 183 schoolchildren and their parents in Madrid (Spain). Inferential statistical analyses, i.e. correlation and hierarchical multiple regression, were carried out and possible mediating effects between variables have been tested. Our results support the hypothesis that family members' levels of dental fear are significantly correlated, and they also allow us to affirm that fathers' dental fear is a mediating variable in the relationship between mothers and children's fear scores. Together with the presence of emotional transmission of dental fear among family members, we identified the relevant role that fathers play as regards the transfer of dental fear from parents to children. © 2011 The Authors. International Journal of Paediatric Dentistry © 2011 BSPD, IAPD and Blackwell Publishing Ltd.

The genome of Nectria haematococca: contribution of supernumerary chromosomes to gene expansion.

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Jeffrey J Coleman

2009-08-01

Full Text Available The ascomycetous fungus Nectria haematococca, (asexual name Fusarium solani, is a member of a group of >50 species known as the "Fusarium solani species complex". Members of this complex have diverse biological properties including the ability to cause disease on >100 genera of plants and opportunistic infections in humans. The current research analyzed the most extensively studied member of this complex, N. haematococca mating population VI (MPVI. Several genes controlling the ability of individual isolates of this species to colonize specific habitats are located on supernumerary chromosomes. Optical mapping revealed that the sequenced isolate has 17 chromosomes ranging from 530 kb to 6.52 Mb and that the physical size of the genome, 54.43 Mb, and the number of predicted genes, 15,707, are among the largest reported for ascomycetes. Two classes of genes have contributed to gene expansion: specific genes that are not found in other fungi including its closest sequenced relative, Fusarium graminearum; and genes that commonly occur as single copies in other fungi but are present as multiple copies in N. haematococca MPVI. Some of these additional genes appear to have resulted from gene duplication events, while others may have been acquired through horizontal gene transfer. The supernumerary nature of three chromosomes, 14, 15, and 17, was confirmed by their absence in pulsed field gel electrophoresis experiments of some isolates and by demonstrating that these isolates lacked chromosome-specific sequences found on the ends of these chromosomes. These supernumerary chromosomes contain more repeat sequences, are enriched in unique and duplicated genes, and have a lower G+C content in comparison to the other chromosomes. Although the origin(s of the extra genes and the supernumerary chromosomes is not known, the gene expansion and its large genome size are consistent with this species' diverse range of habitats. Furthermore, the presence of unique

The genome of Nectria haematococca: contribution of supernumerary chromosomes to gene expansion

Energy Technology Data Exchange (ETDEWEB)

Coleman, J.J.; Rounsley, S.D.; Rodriguez-Carres, M.; Kuo, A.; Wasmann, C.c.; Grimwood, J.; Schmutz, J.; Taga, M.; White, G.J.; Zhuo, S.; Schwartz, D.C.; Freitag, M.; Ma, L.-J.; Danchin, E.G.J.; Henrissat, B.; Cutinho, P.M.; Nelson, D.R.; Straney, D.; Napoli, C.A.; Baker, B.M.; Gribskov, M.; Rep, M.; Kroken, S.; Molnar, I.; Rensing, C.; Kennell, J.C.; Zamora, J.; Farman, M.L.; Selker, E.U.; Salamov, A.; Shapiro, H.; Pangilinan, J.; Lindquist, E.; Lamers, C.; Grigoriev, I.V.; Geiser, D.M.; Covert, S.F.; Temporini, S.; VanEtten, H.D.

2009-04-20

The ascomycetous fungus Nectria haematococca, (asexual name Fusarium solani), is a member of a group of .50 species known as the"Fusarium solani species complex". Members of this complex have diverse biological properties including the ability to cause disease on .100 genera of plants and opportunistic infections in humans. The current research analyzed the most extensively studied member of this complex, N. haematococca mating population VI (MPVI). Several genes controlling the ability of individual isolates of this species to colonize specific habitats are located on supernumerary chromosomes. Optical mapping revealed that the sequenced isolate has 17 chromosomes ranging from 530 kb to 6.52 Mb and that the physical size of the genome, 54.43 Mb, and the number of predicted genes, 15,707, are among the largest reported for ascomycetes. Two classes of genes have contributed to gene expansion: specific genes that are not found in other fungi including its closest sequenced relative, Fusarium graminearum; and genes that commonly occur as single copies in other fungi but are present as multiple copies in N. haematococca MPVI. Some of these additional genes appear to have resulted from gene duplication events, while others may have been acquired through horizontal gene transfer. The supernumerary nature of three chromosomes, 14, 15, and 17, was confirmed by their absence in pulsed field gel electrophoresis experiments of some isolates and by demonstrating that these isolates lacked chromosome-specific sequences found on the ends of these chromosomes. These supernumerary chromosomes contain more repeat sequences, are enriched in unique and duplicated genes, and have a lower G+C content in comparison to the other chromosomes. Although the origin(s) of the extra genes and the supernumerary chromosomes is not known, the gene expansion and its large genome size are consistent with this species' diverse range of habitats. Furthermore, the presence of unique genes on

Chromosome aberration analysis in peripheral lymphocytes of Gulf war and Balkans war veterans

Energy Technology Data Exchange (ETDEWEB)

Schroeder, H.; Heimers, A.; Frentzel-Beyme, R.; Schott, A.; Hoffmann, W

2003-07-01

Chromosome aberrations and sister chromatid exchanges (SCEs) were determined in standard peripheral lymphocyte metaphase preparations of 13 British Gulf War veterans, two veterans of the recent war in the Balkans, and one veteran of both wars. All 16 volunteers suspect exposures to depleted uranium while deployed at the two different theatres of war in 1990 and later on. The Bremen laboratory control served as a reference in this study. Compared with this control there was a statistically significant increase in the frequency of dicentric chromosomes (dic) and centric ring chromosomes (cR) in the veterans' group, indicating a previous exposure to ionising radiation. The statistically significant overdispersion of dic and cR indicates non-uniform irradiation as would be expected after non-uniform exposure and/or exposure to radiation with a high linear energy transfer. The frequency of SCEs was decreased when compared with the laboratory control. (author)

Chromosome aberration analysis in peripheral lymphocytes of Gulf war and Balkans war veterans

International Nuclear Information System (INIS)

Schroeder, H.; Heimers, A.; Frentzel-Beyme, R.; Schott, A.; Hoffmann, W.

2003-01-01

Chromosome aberrations and sister chromatid exchanges (SCEs) were determined in standard peripheral lymphocyte metaphase preparations of 13 British Gulf War veterans, two veterans of the recent war in the Balkans, and one veteran of both wars. All 16 volunteers suspect exposures to depleted uranium while deployed at the two different theatres of war in 1990 and later on. The Bremen laboratory control served as a reference in this study. Compared with this control there was a statistically significant increase in the frequency of dicentric chromosomes (dic) and centric ring chromosomes (cR) in the veterans' group, indicating a previous exposure to ionising radiation. The statistically significant overdispersion of dic and cR indicates non-uniform irradiation as would be expected after non-uniform exposure and/or exposure to radiation with a high linear energy transfer. The frequency of SCEs was decreased when compared with the laboratory control. (author)

Microdissection and chromosome painting of the alien chromosome in an addition line of wheat--Thinopyrum intermedium.

Directory of Open Access Journals (Sweden)

Chuanliang Deng

Full Text Available In this study, chromosome painting was developed and used to identify alien chromosomes in TAi-27, a wheat--Thinopyrum intermedium addition line, and the chromosomes of the three different genomes of Th. Intermedium. The smallest alien chromosome of TAi-27 was microdissected and its DNA amplified by DOP-PCR was used as a probe to hybridize with metaphase chromosomes of TAi-27 and Th. intermedium. Results showed that hybridization signals were observed in all regions of a pair of the smallest alien chromosomes and the pericentromeric area of another pair of alien chromosomes in TAi-27, indicating that the probe from microdissected chromosome is species specific. In Th. intermedium, 14 chromosomes had wide and strong hybridization signals distributed mainly on the pericentromere area and 9 chromosomes with narrow and weak signals on the pericentromere area. The remaining chromosomes displayed a very weak or no signal. Sequential FISH/GISH on Th. intermedium chromosomes using the DNAs of microdissected chromosome, Pseudoroegneria spicata (St genome and pDbH12 (a J(s genome specific probe as the probes indicated that the microdissected chromosome belonged to the St genome, three genomes (J(s , J and St in Th. intermedium could be distinguished, in which there is no hybridization signal on J genome that is similar to the genome of Th. bessarabicum. Our results showed that the smallest alien chromosomes may represent a truncated chromosome and the repetitive sequence distribution might be similar in different chromosomes within the St genome. However, the repetitive sequence distributions are different within the J(s genome, within a single chromosome, and among different genomes in Th. intermedium. Our results suggested that chromosome painting could be feasible in some plants and useful in detecting chromosome variation and repetitive sequence distribution in different genomes of polyploidy plants, which is helpful for understanding the evolution of

Pure chromosome-specific PCR libraries from single sorted chromosomes

NARCIS (Netherlands)

VanDevanter, D. R.; Choongkittaworn, N. M.; Dyer, K. A.; Aten, J. A.; Otto, P.; Behler, C.; Bryant, E. M.; Rabinovitch, P. S.

1994-01-01

Chromosome-specific DNA libraries can be very useful in molecular and cytogenetic genome mapping studies. We have developed a rapid and simple method for the generation of chromosome-specific DNA sequences that relies on polymerase chain reaction (PCR) amplification of a single flow-sorted

Binding of Multiple Rap1 Proteins Stimulates Chromosome Breakage Induction during DNA Replication.

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Greicy H Goto

2015-08-01

Full Text Available Telomeres, the ends of linear eukaryotic chromosomes, have a specialized chromatin structure that provides a stable chromosomal terminus. In budding yeast Rap1 protein binds to telomeric TG repeat and negatively regulates telomere length. Here we show that binding of multiple Rap1 proteins stimulates DNA double-stranded break (DSB induction at both telomeric and non-telomeric regions. Consistent with the role of DSB induction, Rap1 stimulates nearby recombination events in a dosage-dependent manner. Rap1 recruits Rif1 and Rif2 to telomeres, but neither Rif1 nor Rif2 is required for DSB induction. Rap1-mediated DSB induction involves replication fork progression but inactivation of checkpoint kinase Mec1 does not affect DSB induction. Rap1 tethering shortens artificially elongated telomeres in parallel with telomerase inhibition, and this telomere shortening does not require homologous recombination. These results suggest that Rap1 contributes to telomere homeostasis by promoting chromosome breakage.

Chromosomal homologies among vampire bats revealed by chromosome painting (phyllostomidae, chiroptera).

Science.gov (United States)

Sotero-Caio, C G; Pieczarka, J C; Nagamachi, C Y; Gomes, A J B; Lira, T C; O'Brien, P C M; Ferguson-Smith, M A; Souza, M J; Santos, N

2011-01-01

Substantial effort has been made to elucidate karyotypic evolution of phyllostomid bats, mostly through comparisons of G-banding patterns. However, due to the limited number of G-bands in respective karyotypes and to the similarity of non-homologous bands, an accurate evolutionary history of chromosome segments remains questionable. This is the case for vampire bats (Desmodontinae). Despite several proposed homologies, banding data have not yet provided a detailed understanding of the chromosomal changes within vampire genera. We examined karyotype differentiation of the 3 species within this subfamily using whole chromosomal probes from Phyllostomus hastatus (Phyllostominae) and Carollia brevicauda (Carolliinae). Painting probes of P. hastatus respectively detected 22, 21 and 23 conserved segments in Diphylla ecaudata, Diaemus youngi, and Desmodus rotundus karyotypes, whereas 27, 27 and 28 were respectively detectedwith C. brevicauda paints. Based on the evolutionary relationships proposed by morphological and molecular data, we present probable chromosomal synapomorphies for vampire bats and propose chromosomes that were present in the common ancestor of the 5 genera analyzed. Karyotype comparisons allowed us to relate a number of conserved chromosomal segments among the 5 species, providing a broader database for understanding karyotype evolution in the family. 2010 S. Karger AG, Basel.

Y-chromosome DNA is present in the blood of female dogs suggesting the presence of fetal microchimerism.

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Sandra M Axiak-Bechtel

Full Text Available Fetal microchimerism has been suggested to play contradictory roles in women's health, with factors including age of the recipient, time elapsed since microchimerism occurred, and microchimeric cell type modulating disease. Both beneficial and harmful effects have been identified in wound healing and tissue regeneration, immune mediated disease, and cancer. This area of research is relatively new, and hindered by the time course from occurrence of fetal microchimerism to the multi-factorial development of disease. Dogs represent an excellent model for study of fetal microchimerism, as they share our environment, have a naturally condensed lifespan, and spontaneously develop immune-mediated diseases and cancers similar to their human counterparts. However, fetal microchimerism has not been described in dogs. These experiments sought preliminary evidence that dogs develop fetal microchimerism following pregnancy. We hypothesized that Y chromosomal DNA would be detected in the peripheral blood mononuclear cells of female dogs collected within two months of parturition. We further hypothesized that Y chromosomal DNA would be detected in banked whole blood DNA samples from parous female Golden Retrievers with at least one male puppy in a prior litter. Amplification of DNA extracted from five female Golden Retrievers that had whelped within the two months prior to collection revealed strong positive bands for the Y chromosome. Of banked, parous samples, 36% yielded positive bands for the Y chromosome. This is the first report of persistent Y chromosomal DNA in post-partum female dogs and these results suggest that fetal microchimerism occurs in the canine species. Evaluation of the contributions of fetal microchimeric cells to disease processes in dogs as a model for human disease is warranted.

Y-chromosome DNA is present in the blood of female dogs suggesting the presence of fetal microchimerism.

Science.gov (United States)

Axiak-Bechtel, Sandra M; Kumar, Senthil R; Hansen, Sarah A; Bryan, Jeffrey N

2013-01-01

Fetal microchimerism has been suggested to play contradictory roles in women's health, with factors including age of the recipient, time elapsed since microchimerism occurred, and microchimeric cell type modulating disease. Both beneficial and harmful effects have been identified in wound healing and tissue regeneration, immune mediated disease, and cancer. This area of research is relatively new, and hindered by the time course from occurrence of fetal microchimerism to the multi-factorial development of disease. Dogs represent an excellent model for study of fetal microchimerism, as they share our environment, have a naturally condensed lifespan, and spontaneously develop immune-mediated diseases and cancers similar to their human counterparts. However, fetal microchimerism has not been described in dogs. These experiments sought preliminary evidence that dogs develop fetal microchimerism following pregnancy. We hypothesized that Y chromosomal DNA would be detected in the peripheral blood mononuclear cells of female dogs collected within two months of parturition. We further hypothesized that Y chromosomal DNA would be detected in banked whole blood DNA samples from parous female Golden Retrievers with at least one male puppy in a prior litter. Amplification of DNA extracted from five female Golden Retrievers that had whelped within the two months prior to collection revealed strong positive bands for the Y chromosome. Of banked, parous samples, 36% yielded positive bands for the Y chromosome. This is the first report of persistent Y chromosomal DNA in post-partum female dogs and these results suggest that fetal microchimerism occurs in the canine species. Evaluation of the contributions of fetal microchimeric cells to disease processes in dogs as a model for human disease is warranted.

Colocalization of coregulated genes: a steered molecular dynamics study of human chromosome 19.

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Marco Di Stefano

Full Text Available The connection between chromatin nuclear organization and gene activity is vividly illustrated by the observation that transcriptional coregulation of certain genes appears to be directly influenced by their spatial proximity. This fact poses the more general question of whether it is at all feasible that the numerous genes that are coregulated on a given chromosome, especially those at large genomic distances, might become proximate inside the nucleus. This problem is studied here using steered molecular dynamics simulations in order to enforce the colocalization of thousands of knowledge-based gene sequences on a model for the gene-rich human chromosome 19. Remarkably, it is found that most (≈ 88% gene pairs can be brought simultaneously into contact. This is made possible by the low degree of intra-chromosome entanglement and the large number of cliques in the gene coregulatory network. A clique is a set of genes coregulated all together as a group. The constrained conformations for the model chromosome 19 are further shown to be organized in spatial macrodomains that are similar to those inferred from recent HiC measurements. The findings indicate that gene coregulation and colocalization are largely compatible and that this relationship can be exploited to draft the overall spatial organization of the chromosome in vivo. The more general validity and implications of these findings could be investigated by applying to other eukaryotic chromosomes the general and transferable computational strategy introduced here.

Plasmid Transfer in the Ocean – A Case Study from the Roseobacter Group

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Jörn Petersen

2017-07-01

Full Text Available Plasmid mediated horizontal gene transfer (HGT has been speculated to be one of the prime mechanisms for the adaptation of roseobacters (Rhodobacteraceae to their ecological niches in the marine habitat. Their plasmids contain ecologically crucial functional modules of up to ∼40-kb in size, e.g., for aerobic anoxygenic photosynthesis, flagellar formation and the biosynthesis of the antibiotic tropodithietic acid. Furthermore, the widely present type four secretion system (T4SS of roseobacters has been shown to mediate conjugation across genus barriers, albeit in the laboratory. Here we discovered that Confluentimicrobium naphthalenivorans NS6T, a tidal flat bacterium isolated in Korea, carries a 185-kb plasmid, which exhibits a long-range synteny with the conjugative 126-kb plasmid of Dinoroseobacter shibae DFL12T. Both replicons are stably maintained by RepABC operons of the same compatibility group (-2 and they harbor a homologous T4SS. Principal component analysis of the codon usage shows a large similarity between the two plasmids, while the chromosomes are very distinct, showing that neither of the two bacterial species represents the original host of those RepABC-2 type plasmids. The two species do not share a common habitat today and they are phylogenetically only distantly related. Our finding demonstrates the first clear-cut evidence for conjugational plasmid transfer across biogeographical and phylogenetic barriers in Rhodobacteraceae and documents the importance of conjugative HGT in the ocean.

The X chromosome in space.

Science.gov (United States)

Jégu, Teddy; Aeby, Eric; Lee, Jeannie T

2017-06-01

Extensive 3D folding is required to package a genome into the tiny nuclear space, and this packaging must be compatible with proper gene expression. Thus, in the well-hierarchized nucleus, chromosomes occupy discrete territories and adopt specific 3D organizational structures that facilitate interactions between regulatory elements for gene expression. The mammalian X chromosome exemplifies this structure-function relationship. Recent studies have shown that, upon X-chromosome inactivation, active and inactive X chromosomes localize to different subnuclear positions and adopt distinct chromosomal architectures that reflect their activity states. Here, we review the roles of long non-coding RNAs, chromosomal organizational structures and the subnuclear localization of chromosomes as they relate to X-linked gene expression.

NONO ubiquitination is mediated by FBW7 and GSK3 β via a degron lost upon chromosomal rearrangement in cancer.

Science.gov (United States)

Alfano, Luigi; Caporaso, Antonella; Altieri, Angela; Costa, Caterina; Forte, Iris M; Iannuzzi, Carmelina A; Barone, Daniela; Esposito, Luca; Giordano, Antonio; Pentimalli, Francesca

2018-05-01

NONO is an RNA-binding protein involved in transcription, mRNA splicing, DNA repair, and checkpoint activation in response to UV radiation. NONO expression has been found altered in several tumor types, including prostate, colon, breast, melanoma, and in papillary renal carcinoma, in which an X chromosome inversion generates a NONO-TFE3 fusion protein. Upon such rearrangement, NONO loses its C-terminal domain. Through bioinformatics analysis, we identified a putative degron motif, known to be recognized by the Skp1-Cul1-F-box-protein (SCF) complex. Here, we evaluated how this domain could affect NONO protein biology. We showed that NONO interacts with the nuclear FBW7α isoform and its ubiquitination is regulated following modulation of the GSK3β kinase. Mutation of T428A/T432A within the degron impaired polyubiquitination upon FBW7α and GSK3β overexpression. Overall, our data suggest that NONO is likely subjected to proteasome-mediated degradation and add NONO to the list of proteins targeted by FBW7, which is itself often deregulated in cancer. © 2017 Wiley Periodicals, Inc.

Chromosome isolation by flow sorting in Aegilops umbellulata and Ae. comosa and their allotetraploid hybrids Ae. biuncialis and Ae. geniculata.

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István Molnár

Full Text Available This study evaluates the potential of flow cytometry for chromosome sorting in two wild diploid wheats Aegilops umbellulata and Ae. comosa and their natural allotetraploid hybrids Ae. biuncialis and Ae. geniculata. Flow karyotypes obtained after the analysis of DAPI-stained chromosomes were characterized and content of chromosome peaks was determined. Peaks of chromosome 1U could be discriminated in flow karyotypes of Ae. umbellulata and Ae. biuncialis and the chromosome could be sorted with purities exceeding 95%. The remaining chromosomes formed composite peaks and could be sorted in groups of two to four. Twenty four wheat SSR markers were tested for their position on chromosomes of Ae. umbellulata and Ae. comosa using PCR on DNA amplified from flow-sorted chromosomes and genomic DNA of wheat-Ae. geniculata addition lines, respectively. Six SSR markers were located on particular Aegilops chromosomes using sorted chromosomes, thus confirming the usefulness of this approach for physical mapping. The SSR markers are suitable for marker assisted selection of wheat-Aegilops introgression lines. The results obtained in this work provide new opportunities for dissecting genomes of wild relatives of wheat with the aim to assist in alien gene transfer and discovery of novel genes for wheat improvement.

Effective generation of transgenic pigs and mice by linker based sperm-mediated gene transfer.

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Shih Ping Yao

2002-04-01

Full Text Available Abstract Background Transgenic animals have become valuable tools for both research and applied purposes. The current method of gene transfer, microinjection, which is widely used in transgenic mouse production, has only had limited success in producing transgenic animals of larger or higher species. Here, we report a linker based sperm-mediated gene transfer method (LB-SMGT that greatly improves the production efficiency of large transgenic animals. Results The linker protein, a monoclonal antibody (mAb C, is reactive to a surface antigen on sperm of all tested species including pig, mouse, chicken, cow, goat, sheep, and human. mAb C is a basic protein that binds to DNA through ionic interaction allowing exogenous DNA to be linked specifically to sperm. After fertilization of the egg, the DNA is shown to be successfully integrated into the genome of viable pig and mouse offspring with germ-line transfer to the F1 generation at a highly efficient rate: 37.5% of pigs and 33% of mice. The integration is demonstrated again by FISH analysis and F2 transmission in pigs. Furthermore, expression of the transgene is demonstrated in 61% (35/57 of transgenic pigs (F0 generation. Conclusions Our data suggests that LB-SMGT could be used to generate transgenic animals efficiently in many different species.

Social Support at the Workplace, Motivation to Transfer and Training Transfer: A Multilevel Indirect Effects Model

Science.gov (United States)

Massenberg, Ann-Christine; Spurk, Daniel; Kauffeld, Simone

2015-01-01

Supervisor support, peer support and transfer motivation have been identified as important predictors of training transfer. Transfer motivation is thought to mediate the support-training transfer relationship. Especially after team training interventions that include all team members (i.e. whole-team training), individual perception of these…

Liposome-mediated transfer of IL-1 receptor antagonist gene to dispersed islet cells does not prevent recurrence of disease in syngeneically transplanted NOD mice

DEFF Research Database (Denmark)

Saldeen, J; Sandler, S; Bendtzen, K

2000-01-01

transplanted non-obese diabetic (NOD) mice. NOD mouse islet cells were transfected using liposome-mediated gene transfer with a human IL-1ra cDNA construct and transplanted two days later to prediabetic NOD mice. Graft infiltration and destruction were monitored three, five and eight days posttransplantation...... by histology and determination of insulin and cytokine content. IL-1ra gene transfer resulted in transient expression of IL-1ra protein in islet cells in vitro as assessed by ELISA and of IL-1ra mRNA in transplanted islets as revealed by RT-PCR. However, both control and IL-1ra transfected NOD grafts exhibited......IL-1beta is cytotoxic to pancreatic beta-cells in vitro but its role in the vicinity of beta-cells in vivo is unknown. We explored whether liposome-mediated transfer of the interleukin 1 receptor antagonist (IL-1ra) gene to islet cells might prevent recurrence of disease in syngeneically...

Photoinduced electron transfer from organic semiconductors onto redox mediators for CO2

International Nuclear Information System (INIS)

Portenkirchner, E.

2014-01-01

In this work the photoinduced electron transfer from organic semiconductors onto redox mediator catalysts for CO 2 reduction has been investigated. In the beginning, the work focuses on the identication, characterization and test of suitable catalyst materials. For this purpose, rhenium compounds with 2,2'-bipyridine bis(arylimino) acenaphthene ligands and pyridinium were tested for molecular homogenous catalysis. Infrared, ultraviolet-visible (UV-Vis) and nuclear magnetic resonance (NMR) spectroscopy were used for initial characterization of the catalyst substances. Since the interpretation of infrared spectra was difficult for large molecules based on measured data only, additionally infrared absorption spectra obtained by quantum mechanical density functional theory(DFT) calculations were successfully used to correlate characteristic features in the measured spectra to their molecular origin. It was found that experimentally observed data and quantum chemical predictions for the infrared spectra of the novel compounds are in good agreement. Additionally, quantum mechanical calculations were carried out for the determination of molecular orbital frontier energy levels and correlated to UV-Vis absorption and cyclic voltammetry measurements. Extensive cyclic voltammetry measurements and bulk controlled-potential electrolysis experiments were performed using a N 2 - and CO 2 -saturated electrolyte solution. Together with a detailed product analysis via infrared spectroscopy, gas and ion chromatography the results allowed electrochemical characterizations of the novel catalysts regarding their suitability for electrochemical CO 2 reduction. Once suitable catalysts were identied, the materials were immobilized on the electrode surface by electro-polymerization of the catalyst (5,5'bisphenylethynyl-2,2'-bipyridyl)Re(CO) 3 Cl itself or by incorporation of (2,2'-bipyridyl)Re(CO) 3 Cl into a polypyrrole matrix, thereby changing from homogeneous to

New Y chromosomes and early stages of sex chromosome ...

Indian Academy of Sciences (India)

2010-09-06

Sep 6, 2010 ... chromosomes are evolutionary consequences of that func- tion. Given sufficient ... (for a review, see Charlesworth et al. 2005). ... In the present paper, I review sex deter- mination .... part had apparently been exchanged against the homologous ... age group III-Y chromosomes were successful while in well-.

Using 3-color chromosome painting to decide between chromosome aberration models

International Nuclear Information System (INIS)

Lucas, J.N.; Sachs, R.K.

1993-01-01

Ionizing radiation produces chromosome aberrations when DNA double strand breaks (DSB) interact pairwise. For more than 30 years there have been two main, competing theories of such binary DSB interactions. The classical theory asserts that an unrepaired DSB makes two ends which separate, with each end subsequently able to join any similar (non-telomeric) end. The exchange theory asserts that the two DSB ends remain associated until repair or a reciprocal chromosome exchange involving a second DSB occurs. The authors conducted an experiment to test these models, using 3-color chromosome painting. After in vitro irradiation of resting human lymphocytes, they observed cells with three-color triplets at first metaphase: three derivative chromosomes having permuted colors, as if three broken chromosomes had played musical chairs. On the exchange model in its standard form such 3-color triplets cannot occur. On the classical model the expected frequency can be calculated. They report data and computer calculations which exclude the exchange model and favor the classical model

Mediated Electron Transfer at Vertically Aligned Single-Walled Carbon Nanotube Electrodes During Detection of DNA Hybridization

Science.gov (United States)

Wallen, Rachel; Gokarn, Nirmal; Bercea, Priscila; Grzincic, Elissa; Bandyopadhyay, Krisanu

2015-06-01

Vertically aligned single-walled carbon nanotube (VASWCNT) assemblies are generated on cysteamine and 2-mercaptoethanol (2-ME)-functionalized gold surfaces through amide bond formation between carboxylic groups generated at the end of acid-shortened single-walled carbon nanotubes (SWCNTs) and amine groups present on the gold surfaces. Atomic force microscopy (AFM) imaging confirms the vertical alignment mode of SWCNT attachment through significant changes in surface roughness compared to bare gold surfaces and the lack of any horizontally aligned SWCNTs present. These SWCNT assemblies are further modified with an amine-terminated single-stranded probe-DNA. Subsequent hybridization of the surface-bound probe-DNA in the presence of complementary strands in solution is followed using impedance measurements in the presence of Fe(CN)6 3-/4- as the redox probe in solution, which show changes in the interfacial electrochemical properties, specifically the charge-transfer resistance, due to hybridization. In addition, hybridization of the probe-DNA is also compared when it is attached directly to the gold surfaces without any intermediary SWCNTs. Contrary to our expectations, impedance measurements show a decrease in charge-transfer resistance with time due to hybridization with 300 nM complementary DNA in solution with the probe-DNA attached to SWCNTs. In contrast, an increase in charge-transfer resistance is observed with time during hybridization when the probe-DNA is attached directly to the gold surfaces. The decrease in charge-transfer resistance during hybridization in the presence of VASWCNTs indicates an enhancement in the electron transfer process of the redox probe at the VASWCNT-modified electrode. The results suggest that VASWCNTs are acting as mediators of electron transfer, which facilitate the charge transfer of the redox probe at the electrode-solution interface.

Somatically Acquired Isodicentric Y and Mosaic Loss of Chromosome Y in a Boy with Hypospadias.

Science.gov (United States)

Miyado, Mami; Muroya, Koji; Katsumi, Momori; Saito, Kazuki; Kon, Masafumi; Fukami, Maki

2018-04-07

Isodicentric Y chromosome [idic(Y)] represents a relatively common subtype of Y chromosomal rearrangements in the germline; however, limited evidence supports the postzygotic occurrence of idic(Y). Here, we report a boy with hypospadias and somatically acquired idic(Y). The 3.5-year-old boy has been identified in our previous study for patients with hypospadias. In the present study, cytogenetic analysis including FISH revealed a 45,X[5]/46,X,idic(Y)[7]/46,XY[8] karyotype. MLPA showed a mosaic deletion involving PPP1R12BP1 and RBMY2DP. The idic(Y) was likely to have been formed through aberrant recombination between P1 palindromes and subsequently underwent mosaic loss. The patient's phenotype was attributable to deletion of some Y chromosomal genes and/or mosaic loss of chromosome Y (mLOY). The results suggest that idic(Y) can originate in postzygotic cells via palindrome-mediated crossovers. Moreover, our data indicate that somatically acquired idic(Y) can trigger mLOY, which usually appears as an aging-related phenomenon in elderly men. © 2018 S. Karger AG, Basel.

Cyclopentadiene-mediated hydride transfer from rhodium complexes.

Science.gov (United States)

Pitman, C L; Finster, O N L; Miller, A J M

2016-07-12

Attempts to generate a proposed rhodium hydride catalytic intermediate instead resulted in isolation of (Cp*H)Rh(bpy)Cl (1), a pentamethylcyclopentadiene complex, formed by C-H bond-forming reductive elimination from the fleeting rhodium hydride. The hydride transfer ability of diene 1 was explored through thermochemistry and hydride transfer reactions, including the reduction of NAD(+).

Persistence of chromosomal abnormalities additional to the Philadelphia chromosome after Philadelphia chromosome disappearance during imatinib therapy for chronic myeloid leukemia.

Science.gov (United States)

Zaccaria, Alfonso; Valenti, Anna Maria; Donti, Emilio; Gozzetti, Alessandro; Ronconi, Sonia; Spedicato, Francesco

2007-04-01

Five Philadelphia chromosome positive (Ph+) chronic myeloid leukemia (CML) patients with additional chromosome abnormalities at diagnosis have been followed during Imatinib therapy. In all, the Ph chromosome disappeared, while the 5 cases, additional abnormalities [dup(1); del(5), +8 (2 patients) and +14] persisted in the subsequent studies, performed over a period of 11 to 49 months, either alone or together with a karyotypically normal cell population. This finding is consistent with a secondary origin of the Ph chromosome in these patients. It is still to early to evaluate the possible prognostic value of these additional abnormalities.

Targeted introgression of a wheat stem rust resistance gene by DNA marker-assisted chromosome engineering.

Science.gov (United States)

Niu, Zhixia; Klindworth, Daryl L; Friesen, Timothy L; Chao, Shiaoman; Jin, Yue; Cai, Xiwen; Xu, Steven S

2011-04-01

Chromosome engineering is a useful strategy for transfer of alien genes from wild relatives into modern crops. However, this strategy has not been extensively used for alien gene introgression in most crops due to low efficiency of conventional cytogenetic techniques. Here, we report an improved scheme of chromosome engineering for efficient elimination of a large amount of goatgrass (Aegilops speltoides) chromatin surrounding Sr39, a gene that provides resistance to multiple stem rust races, including Ug99 (TTKSK) in wheat. The wheat ph1b mutation, which promotes meiotic pairing between homoeologous chromosomes, was employed to induce recombination between wheat chromosome 2B and goatgrass 2S chromatin using a backcross scheme favorable for inducing and detecting the homoeologous recombinants with small goatgrass chromosome segments. Forty recombinants with Sr39 with reduced surrounding goatgrass chromatin were quickly identified from 1048 backcross progenies through disease screening and molecular marker analysis. Four of the recombinants carrying Sr39 with a minimal amount of goatgrass chromatin (2.87-9.15% of the translocated chromosomes) were verified using genomic in situ hybridization. Approximately 97% of the goatgrass chromatin was eliminated in one of the recombinants, in which a tiny goatgrass chromosome segment containing Sr39 was retained in the wheat genome. Localization of the goatgrass chromatin in the recombinants led to rapid development of three molecular markers tightly linked to Sr39. The new wheat lines and markers provide useful resources for the ongoing global effort to combat Ug99. This study has demonstrated great potential of chromosome engineering in genome manipulation for plant improvement.

Biodosimetry of ionizing radiation by selective painting of prematurely condensed chromosomes in human lymphocytes

Science.gov (United States)

Durante, M.; George, K.; Yang, T. C.

1997-01-01

Painting of interphase chromosomes can be useful for biodosimetric purposes in particular cases such as radiation therapy, accidental exposure to very high radiation doses and exposure to densely ionizing radiation, for example during space missions. Biodosimetry of charged-particle radiation is analyzed in the present paper. Target cells were human peripheral blood lymphocytes irradiated in vitro with gamma rays, protons and iron ions. After exposure, lymphocytes were incubated for different times to allow repair of radiation-induced damage and then fused to mitotic hamster cells to promote premature condensation in the interphase chromosomes. Chromosome spreads were then hybridized with whole-chromosome DNA probes labeled with fluorescent stains. Dose-response curves for the induction of chromatin fragments shortly after exposure, as well as the kinetics of rejoining and misrejoining, were not markedly dependent on linear energy transfer. However, after exposure to heavy ions, more aberrations were scored in the interphase cells after incubation for repair than in metaphase samples harvested at the first postirradiation mitosis. On the other hand, no significant differences were observed in the two samples after exposure to sparsely ionizing radiation. These results suggest that interphase chromosome painting can be a useful tool for biodosimetry of particle radiation.

Chromosomal instability and double minute chromosomes in a breast cancer patient

International Nuclear Information System (INIS)

Lalic, H.; Radosevic-Stasic, B.

2004-01-01

Cytogenetic analysis was performed in peripheral blood lymphocytes (PBL) of a woman with ductal breast carcinoma, who as a hospital employee was exposed professionally for 15 years to low doses of ionizing radiation. The most important finding after the chemotherapy in combination with radiotherapy was the presence of double minutes (DM) chromosomes, in combination with other chromosomal abnormalities (on 200 scored metaphases were found 2 chromatid breaks, 10 dicentrics, 11 acentric fragments, 2 gaps, and 3 double min chromosomes). In a repeated analysis (after 6 months), DM chromosomes were still present. To rule out the possibility that the patient was overexposed to ionizing radiation at work, her blood test was compared with a group of coworkers as well as with a group of professionally unexposed people. The data rejected this possibility, but the retroactive analysis showed that the patient even at the time of employment had a moderately increased number of chromosomal aberrations (3.5%) consisting of 3 isochromatids and 4 gaps, suggesting that her initial genomic instability enhanced the later development. The finding of a continuous presence of rare DM chromosomes in her PBL (4 and 10 months after radio-chemotherapy) was considered as an indicator of additional risk, which might have some prognostic significance. (author)

Exploration of new perspectives and limitations in Agrobacterium-mediated gene transfer technology. Final report, June 1, 1992--May 31, 1995

Energy Technology Data Exchange (ETDEWEB)

Marton, L.

1996-02-01

Genetic manipulation of plants often involves the introduction of homologous or partly homologous genes. Ectropic introduction of homologous sequences into plant genomes may trigger epigenetic changes, making expression of the genes unpredictable. The main project objective was to examine the feasibility of using Agrobacterium-mediated gene transfer for homologous gene targeting in plants.

Chromosome painting in plants.

NARCIS (Netherlands)

Schubert, I.; Fransz, P.F.; Fuchs, J.; Jong, de J.H.

2001-01-01

The current 'state-of-art' as to chromosome painting in plants is reviewed. We define different situations described as painting so far: i) Genomic in situ hybridisation (GISH) with total genomic DNA to distinguish alien chromosomes on the basis of divergent dispersed repeats, ii) 'Chromosomal in

Efficient gene transfer into nondividing cells by adeno-associated virus-based vectors.

Science.gov (United States)

Podsakoff, G; Wong, K K; Chatterjee, S

1994-09-01

Gene transfer vectors based on adeno-associated virus (AAV) are emerging as highly promising for use in human gene therapy by virtue of their characteristics of wide host range, high transduction efficiencies, and lack of cytopathogenicity. To better define the biology of AAV-mediated gene transfer, we tested the ability of an AAV vector to efficiently introduce transgenes into nonproliferating cell populations. Cells were induced into a nonproliferative state by treatment with the DNA synthesis inhibitors fluorodeoxyuridine and aphidicolin or by contact inhibition induced by confluence and serum starvation. Cells in logarithmic growth or DNA synthesis arrest were transduced with vCWR:beta gal, an AAV-based vector encoding beta-galactosidase under Rous sarcoma virus long terminal repeat promoter control. Under each condition tested, vCWR:beta Gal expression in nondividing cells was at least equivalent to that in actively proliferating cells, suggesting that mechanisms for virus attachment, nuclear transport, virion uncoating, and perhaps some limited second-strand synthesis of AAV vectors were present in nondividing cells. Southern hybridization analysis of vector sequences from cells transduced while in DNA synthetic arrest and expanded after release of the block confirmed ultimate integration of the vector genome into cellular chromosomal DNA. These findings may provide the basis for the use of AAV-based vectors for gene transfer into quiescent cell populations such as totipotent hematopoietic stem cells.

Frequency of chromosomal aneuploidy in high quality embryos from young couples using preimplantation genetic screening

Directory of Open Access Journals (Sweden)

Farzaneh Fesahat

2017-09-01

Full Text Available Background: Selection of the best embryo for transfer is very important in assisted reproductive technology (ART. Using morphological assessment for this selection demonstrated that the correlation between embryo morphology and implantation potential is relatively weak. On the other hand, aneuploidy is a key genetic factor that can influence human reproductive success in ART. Objective: The aim of this lab trial study was to evaluate the incidence of aneuploidies in five chromosomes in the morphologically high-quality embryos from young patients undergoing ART for sex selection. Materials and Methods: A total of 97 high quality embryos from 23 women at the age of 37or younger years that had previously undergone preimplantation genetic screening for sex selection were included in this study. After washing, the slides of blastomeres from embryos of patients were reanalyzed by fluorescence in-situ hybridization for chromosomes 13, 18 and 21. Results: There was a significant rate of aneuploidy determination in the embryos using preimplantation genetic screening for both sex and three evaluated autosomal chromosomes compared to preimplantation genetic screening for only sex chromosomes (62.9% vs. 24.7%, p=0.000. The most frequent detected chromosomal aneuploidy was trisomy or monosomy of chromosome 13. Conclusion: There is considerable numbers of chromosomal abnormalities in embryos generated in vitro which cause in vitro fertilization failure and it seems that morphological characterization of embryos is not a suitable method for choosing the embryos without these abnormalities

Noninvolvement of the X chromosome in radiation-induced chromosome translocations in the human lymphoblastoid cell line TK6

International Nuclear Information System (INIS)

Jordan, R.; Schwartz, J.L.

1994-01-01

Fluorescence in situ hybridization procedures were used to examine the influence of chromosome locus on the frequency and type of chromosome aberrations induced by 60 Co γ rays in the human lymphoblastoid cell line TK6. Aberrations involving the X chromosome were compared to those involving the similarly sized autosome chromosome 7. When corrected for DNA content, acentric fragments were induced with equal frequency in the X and 7 chromosomes. Dose-dependent increases in chromosomal interchanges involving chromosome 7 were noted, and the frequencies of balanced translocations and dicentrics produced were approximately equal. Chromosome interchanges involving the X chromosome were rare and showed no apparent dose dependence. Thus, while chromosomes 7 and X are equally sensitive to the induction of chromosome breaks, the X chromosome is much less likely to interact with autosomes than chromosome 7. The noninvolvement of the X chromosome in translocations with autosomes may reflect a more peripheral and separate location for the X chromosome in the mammalian nucleus. 20 refs., 2 figs., 1 tab

The convergence of quantum-dot-mediated fluorescence resonance energy transfer and microfluidics for monitoring DNA polyplex self-assembly in real time

International Nuclear Information System (INIS)

Ho Yiping; Wang, T-H; Chen, Hunter H; Leong, Kam W

2009-01-01

We present a novel convergence of quantum-dot-mediated fluorescence resonance energy transfer (QD-FRET) and microfluidics, through which molecular interactions were precisely controlled and monitored using highly sensitive quantum-dot-mediated FRET. We demonstrate its potential in studying the kinetics of self-assembly of DNA polyplexes under laminar flow in real time with millisecond resolution. The integration of nanophotonics and microfluidics offers a powerful tool for elucidating the formation of polyelectrolyte polyplexes, which is expected to provide better control and synthesis of uniform and customizable polyplexes for future nucleic acid-based therapeutics.

Chromosome painting in biological dosimetry: Semi-automatic system to score stable chromosome aberrations

International Nuclear Information System (INIS)

Garcia-Sagredo, J.M.; Vallcorba, I.; Sanchez-Hombre, M.C.; Ferro, M.T.; San Roman Cos-Gayon, C.; Santos, A.; Malpica, N.; Ortiz, C.

1997-01-01

From the beginning of the description of the procedure of chromosome painting by fluorescence in situ hybridization (FISH), it was thought its possible application to score induced chromosomal aberrations in radiation exposition. With chromosome painting it is possible to detect changes between chromosomes that has been validated in radiation exposition. Translocation scoring by FISH, contrarily to the unstable dicentrics, mainly detect stable chromosome aberrations that do not disappear, it allows the capability of quantify delayed acute expositions or chronic cumulative expositions. The large number of cells that have to be analyzed for high accuracy, specially when dealing with low radiation doses, makes it almost imperative to use an automatic analysis system. After validate translocation scoring by FISH in our, we have evaluated the ability and sensitivity to detect chromosomal aberrations by chromosome using different paint probes used, showing that any combination of paint probes can be used to score induced chromosomal aberrations. Our group has developed a FISH analysis that is currently being adapted for translocation scoring analysis. It includes systematic error correction and internal control probes. The performance tests carried out show that 9,000 cells can be analyzed in 10 hr. using a Sparc 4/370. Although with a faster computer, a higher throughput is expected, for large population screening or very low radiation doses, this performance still has to be improved. (author)

A simple strategy for subcloning and amplifying random multimegabase subchromosomal acentric DNA fragments as double minute chromosomes

International Nuclear Information System (INIS)

Hahn, P.J.; Giddings, L.; Lane, M.J.

1989-01-01

Restriction mapping of relatively large genomes (e.g. human) utilizing randomly generated DNA segments requires high mapping redundancy to successfully organize 'contigs' to represent the entire genome. The number of independent DNA segment maps required is dependent on the average size of a mapping segment; the larger the segment, the fewer required. The authors have developed a strategy for subcloning intact multimegabase subchromosomal fragments as double minute chromosomes. Such fragments could serve as primary mapping elements or as adjunct (linking) fragments to rapidly connect already existent contigs generated using yeast artificial chromosomes or cosmids. They present several lines of evidence supporting the viability of this approach. (1) X-ray treated EMT-6 mouse cells (7.5 Gr.) which are selected over several months with increasing levels of methotrexate (MTX) contain highly amplified circular DNA molecules (double minutes) which include the dihydrofolate reductase (DHFR) gene in a size range between 1,000 and 3,500 kilobases as determined by pulsed-field gel electrophoresis and these acentric chromosomal fragments have been stably maintained in culture for at least a year. (2) Preliminary data based on experiments involving fusion of X-irradiated Chinese Hamster Ovary (CH0 DG44) cells containing randomly inserted cotransfected Neomycin resistance and DHFR genes to mouse EMT-6 cells shows that the linked genes can be readily cotransferred as acentric subchromosomal fragment(s) suitable for gene amplification. (3) The studies of CHO cells with cell fusion transferred X-ray induced chromosomal fragments containing the natural CHO DHFR gene suggest that transferred chromosome fragments undergo gene amplification much more readily than nonfragmented endogenous DHFR genes

Speculations on the initiation of chromosome replication in Escherichia coli: the dualism hypothesis.

Science.gov (United States)

Norris, Vic

2011-05-01

The exact nature of the mechanism that triggers initiation of chromosome replication in the best understood of all organisms, Escherichia coli, remains mysterious. Here, I suggest that this mechanism evolved in response to the problems that arise if chromosome replication does not occur. E. coli is now known to be highly structured. This leads me to propose a mechanism for initiation of replication based on the dynamics of large assemblies of molecules and macromolecules termed hyperstructures. In this proposal, hyperstructures and their constituents are put into two classes, non-equilibrium and equilibrium, that spontaneously separate and that are appropriate for life in either good or bad conditions. Maintaining the right ratio(s) of non-equilibrium to equilibrium hyperstructures is therefore a major challenge for cells. I propose that this maintenance entails a major transfer of material from equilibrium to non-equilibrium hyperstructures once per cell and I further propose that this transfer times the cell cycle. More specifically, I speculate that the dialogue between hyperstructures involves the structuring of water and the condensation of cations and that one of the outcomes of ion condensation on ribosomal hyperstructures and decondensation from the origin hyperstructure is the separation of strands at oriC responsible for triggering initiation of replication. The dualism hypothesis that comes out of these speculations may help integrate models for initiation of replication, chromosome segregation and cell division with the 'prebiotic ecology' scenario of the origins of life. Copyright © 2011 Elsevier Ltd. All rights reserved.

Inhibition by TNF-alpha and IL-4 of cationic lipid mediated gene transfer in cystic fibrosis tracheal gland cells.

Science.gov (United States)

Bastonero, Sonia; Gargouri, Myriem; Ortiou, Sandrine; Guéant, Jean-Louis; Merten, Marc D

2005-11-01

In vivo, tracheal gland serous cells highly express the cystic fibrosis transmembrane conductance regulator (cftr) gene. This gene is mutated in the lethal monogenic disease cystic fibrosis (CF). Clinical trials in which the human CFTR cDNA was delivered to the respiratory epithelia of CF patients have resulted in weak and transient gene expression. As CF is characterized by mucus inspissation, airway infection, and severe inflammation, we tested the hypothesis that inflammation and especially two cytokines involved in the Th1/Th2 inflammatory response, interleukin 4 (IL-4) and TNFalpha, could inhibit gene transfer efficiency using a model of human CF tracheal gland cells (CF-KM4) and Lipofectamine reagent as a transfection reagent. The specific secretory defects of CF-KM4 cells were corrected by Lipofectamine-mediated human CFTR gene transfer. However, this was altered when cells were pre-treated with IL-4 and TNFalpha. Inhibition of luciferase reporter gene expression by IL-4 and TNFalpha pre-treated CF-KM4 cells was measured by activity and real-time RT-PCR. Both cytokines induced similar and synergistic inhibition of transgene expression and activity. This cytokine-mediated inhibition could be prevented by anti-inflammatory agents such as glucocorticoids but not by non-steroidal (NSAI) agents. This data suggests that an inflammatory context generated by IL-4 and TNFalpha can inhibit human CFTR gene transfer in CF tracheal gland cells and that glucocorticoids may have a protecting action. Copyright (c) 2005 John Wiley & Sons, Ltd.

A First Generation Comparative Chromosome Map between Guinea Pig (Cavia porcellus) and Humans.

Science.gov (United States)

Romanenko, Svetlana A; Perelman, Polina L; Trifonov, Vladimir A; Serdyukova, Natalia A; Li, Tangliang; Fu, Beiyuan; O'Brien, Patricia C M; Ng, Bee L; Nie, Wenhui; Liehr, Thomas; Stanyon, Roscoe; Graphodatsky, Alexander S; Yang, Fengtang

2015-01-01

The domesticated guinea pig, Cavia porcellus (Hystricomorpha, Rodentia), is an important laboratory species and a model for a number of human diseases. Nevertheless, genomic tools for this species are lacking; even its karyotype is poorly characterized. The guinea pig belongs to Hystricomorpha, a widespread and important group of rodents; so far the chromosomes of guinea pigs have not been compared with that of other hystricomorph species or with any other mammals. We generated full sets of chromosome-specific painting probes for the guinea pig by flow sorting and microdissection, and for the first time, mapped the chromosomal homologies between guinea pig and human by reciprocal chromosome painting. Our data demonstrate that the guinea pig karyotype has undergone extensive rearrangements: 78 synteny-conserved human autosomal segments were delimited in the guinea pig genome. The high rate of genome evolution in the guinea pig may explain why the HSA7/16 and HSA16/19 associations presumed ancestral for eutherians and the three syntenic associations (HSA1/10, 3/19, and 9/11) considered ancestral for rodents were not found in C. porcellus. The comparative chromosome map presented here is a starting point for further development of physical and genetic maps of the guinea pig as well as an aid for genome assembly assignment to specific chromosomes. Furthermore, the comparative mapping will allow a transfer of gene map data from other species. The probes developed here provide a genomic toolkit, which will make the guinea pig a key species to unravel the evolutionary biology of the Hystricomorph rodents.

Comparative genome sequencing of Drosophila pseudoobscura: Chromosomal, gene, and cis-element evolution

DEFF Research Database (Denmark)

Richards, Stephen; Liu, Yue; Bettencourt, Brian R.

2005-01-01

years (Myr) since the pseudoobscura/melanogaster divergence. Genes expressed in the testes had higher amino acid sequence divergence than the genome-wide average, consistent with the rapid evolution of sex-specific proteins. Cis-regulatory sequences are more conserved than random and nearby sequences......We have sequenced the genome of a second Drosophila species, Drosophila pseudoobscura, and compared this to the genome sequence of Drosophila melanogaster, a primary model organism. Throughout evolution the vast majority of Drosophila genes have remained on the same chromosome arm, but within each...... between the species-but the difference is slight, suggesting that the evolution of cis-regulatory elements is flexible. Overall, a pattern of repeat-mediated chromosomal rearrangement, and high coadaptation of both male genes and cis-regulatory sequences emerges as important themes of genome divergence...

Mitotic chromosome condensation in vertebrates

International Nuclear Information System (INIS)

Vagnarelli, Paola

2012-01-01

Work from several laboratories over the past 10–15 years has revealed that, within the interphase nucleus, chromosomes are organized into spatially distinct territories [T. Cremer, C. Cremer, Chromosome territories, nuclear architecture and gene regulation in mammalian cells, Nat. Rev. Genet. 2 (2001) 292–301 and T. Cremer, M. Cremer, S. Dietzel, S. Muller, I. Solovei, S. Fakan, Chromosome territories—a functional nuclear landscape, Curr. Opin. Cell Biol. 18 (2006) 307–316]. The overall compaction level and intranuclear location varies as a function of gene density for both entire chromosomes [J.A. Croft, J.M. Bridger, S. Boyle, P. Perry, P. Teague,W.A. Bickmore, Differences in the localization and morphology of chromosomes in the human nucleus, J. Cell Biol. 145 (1999) 1119–1131] and specific chromosomal regions [N.L. Mahy, P.E. Perry, S. Gilchrist, R.A. Baldock, W.A. Bickmore, Spatial organization of active and inactive genes and noncoding DNA within chromosome territories, J. Cell Biol. 157 (2002) 579–589] (Fig. 1A, A'). In prophase, when cyclin B activity reaches a high threshold, chromosome condensation occurs followed by Nuclear Envelope Breakdown (NEB) [1]. At this point vertebrate chromosomes appear as compact structures harboring an attachment point for the spindle microtubules physically recognizable as a primary constriction where the two sister chromatids are held together. The transition from an unshaped interphase chromosome to the highly structured mitotic chromosome (compare Figs. 1A and B) has fascinated researchers for several decades now; however a definite picture of how this process is achieved and regulated is not yet in our hands and it will require more investigation to comprehend the complete process. From a biochemical point of view a vertebrate mitotic chromosomes is composed of DNA, histone proteins (60%) and non-histone proteins (40%) [6]. I will discuss below what is known to date on the contribution of these two different

Mitotic chromosome condensation in vertebrates

Energy Technology Data Exchange (ETDEWEB)

Vagnarelli, Paola, E-mail: P.Vagnarelli@ed.ac.uk

2012-07-15

Work from several laboratories over the past 10-15 years has revealed that, within the interphase nucleus, chromosomes are organized into spatially distinct territories [T. Cremer, C. Cremer, Chromosome territories, nuclear architecture and gene regulation in mammalian cells, Nat. Rev. Genet. 2 (2001) 292-301 and T. Cremer, M. Cremer, S. Dietzel, S. Muller, I. Solovei, S. Fakan, Chromosome territories-a functional nuclear landscape, Curr. Opin. Cell Biol. 18 (2006) 307-316]. The overall compaction level and intranuclear location varies as a function of gene density for both entire chromosomes [J.A. Croft, J.M. Bridger, S. Boyle, P. Perry, P. Teague,W.A. Bickmore, Differences in the localization and morphology of chromosomes in the human nucleus, J. Cell Biol. 145 (1999) 1119-1131] and specific chromosomal regions [N.L. Mahy, P.E. Perry, S. Gilchrist, R.A. Baldock, W.A. Bickmore, Spatial organization of active and inactive genes and noncoding DNA within chromosome territories, J. Cell Biol. 157 (2002) 579-589] (Fig. 1A, A'). In prophase, when cyclin B activity reaches a high threshold, chromosome condensation occurs followed by Nuclear Envelope Breakdown (NEB) [1]. At this point vertebrate chromosomes appear as compact structures harboring an attachment point for the spindle microtubules physically recognizable as a primary constriction where the two sister chromatids are held together. The transition from an unshaped interphase chromosome to the highly structured mitotic chromosome (compare Figs. 1A and B) has fascinated researchers for several decades now; however a definite picture of how this process is achieved and regulated is not yet in our hands and it will require more investigation to comprehend the complete process. From a biochemical point of view a vertebrate mitotic chromosomes is composed of DNA, histone proteins (60%) and non-histone proteins (40%) [6]. I will discuss below what is known to date on the contribution of these two different classes

Chromosomal rearrangement interferes with meiotic X chromosome inactivation

Czech Academy of Sciences Publication Activity Database

Homolka, David; Ivánek, Robert; Čapková, Jana; Jansa, Petr; Forejt, Jiří

2007-01-01

Roč. 17, č. 10 (2007), s. 1431-1437 ISSN 1088-9051 R&D Projects: GA MŠk(CZ) 1M0520; GA ČR GA301/06/1334; GA ČR GA301/07/1383 Grant - others:Howard Hughes Medical Institute(US) HHMI 55000306 Institutional research plan: CEZ:AV0Z50520514 Keywords : chromosomal translocations * meiotic X chromosome inactivation * spermatogenesis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 11.224, year: 2007

Paternal isodisomy of chromosome 6 in association with a maternal supernumerary marker chromosome (6)

Energy Technology Data Exchange (ETDEWEB)

James, R.S.; Crolla, J.A.; Sitch, F.L. [Salisbury District Hospital, Wiltshire (United Kingdom)] [and others

1994-09-01

Uniparental disomy may arise by a number of different mechanisms of aneuploidy correction. A population that has been identified as being at increased risk of aneuploidy are those individuals bearing supernumerary marker chromosomes (SMCs). There have been a number of cases reported of trisomy 21 in association with bi-satellited marker chromosomes have described two individuals with small inv dup (15) markers. One had paternal isodisomy of chromosome 15 and Angelman syndrome. The other had maternal heterodisomy (15) and Prader-Willi syndrome. At the Wessex Regional Genetics Laboratory we have conducted a search for uniparental disomy of the normal homologues of the chromosomes from which SMCs originated. Our study population consists of 39 probands with SMCs originating from a number of different autosomes, including 17 with SMCs of chromosome 15 origin. Using PCR amplification of microsatellite repeat sequences located distal to the regions included in the SMCs we have determined the parental origin of the two normal homologues in each case. We have identified paternal isodisomy of chromosome 6 in a female child with a supernumerary marker ring chromosome 6 in approximately 70% of peripheral blood lymphocytes. The marker was found to be of maternal origin. This is the second case of paternal isodisomy of chromosome 6 to be reported, and the first in association with a SMC resulting in a partial trisomy for a portion of the short arm of chromosome 6. In spite of this, the patient appears to be functioning appropriately for her age.

Gonadal sex chromosome complement in individuals with sex chromosomal and/or gonadal disorders

Energy Technology Data Exchange (ETDEWEB)

Bridge, J.A.; Sanger, W.G.; Seemayer, T. [Univ. of Nebraska Medical Center, Omaha, NE (United States)] [and others

1994-09-01

Gonadal abnormalities are characteristically seen in patients with sex chromosomal aneuploidy. Morphologically these abnormalities can be variable and are hypothesized to be dependent on the sex chromosomal consititution of the gonad (independent of the chromosomal complement of other tissues, such as peripheral blood lymphocytes). In this study, the gonadal sex chromosome complement was evaluated for potential mosaicism and correlated with the histopathology from 5 patients with known sex chromosomal and/or gonadal disorders. FISH techniques using X and Y chromosome specific probes were performed on nuclei extracted from paraffin embedded tissue. Gonadal tissue obtained from case 1 (a true hemaphroditic newborn) consisted of ovotestes and epididymis (left side) and ovary with fallopian tube (right side). Cytogenetic and FISH studies performed on blood, ovotestes and ovary revealed an XX complement. Cytogenetic analysis of blood from case 2, a 4-year-old with suspected Turner syndrome revealed 45,X/46,X,del(Y)(q11.21). FISH analysis of the resected gonads (histologically = immature testes) confirmed an X/XY mosaic complement. Histologically, the gonadal tissue was testicular. Severe autolysis prohibited successful analysis in the 2 remaining cases. In summary, molecular cytogenetic evaluation of gonadal tissue from individuals with sex chromosomal and/or gonadal disorders did not reveal tissue-specific anomalies which could account for differences observed pathologically.

The distribution of chromosome aberrations among chromosomes of karyotype in exposed human lymphocyte

International Nuclear Information System (INIS)

Que Tran; Tien Hoang Hung

1997-01-01

Induced chromosome aberrations (ch. ab.) in exposed Human peripheral blood lymphocyte have been used to assay radio.bio.doses, because of their characters such as: the maintaining Go phase in cell cycle in body, the distribution of cell in blood system and the distribution of ch. ab. in exposed cells of body and among chromosomes of karyotype. The frequency of ch. ab. reflected the quantity of radiation dose, dose rate and radiation energy. The dependence between radiation dose and frequency of ch. ab. was illustrated by the mathematic equations. The distribution of induced ch. ab. among the cells exposed to uniform radiation fields was Poisson's, but the distribution of ch. ab. among chromosomes in karyotype depended on radiation field and mononucleotid sequence of DNA molecular of each chromosome. The minimum influence of mononucleotid sequence of DNA molecular in inform ch. ab. will be advantageous state for dose-assessments. The location of induced ch. ab. in exposed Human lymphocyte had been determined by karyotype analyses. The data of statistic analyse had improved that the number of ch. ab. depended on the size of chromosomes in karyotype. The equal distribution of ch. ab.among chromosomes in karyotype provided the objectiveness and the accuracy of using the chromosomal aberrant analysis technique on bio-dosimetry. (author)

Detection of Alien Oryza punctata Kotschy Chromosomes in Rice, Oryza sativa L., by Genomic in situ Hybridization

OpenAIRE

Yasui, Hideshi; Nonomura, Ken-ichi; Iwata, Nobuo; 安井, 秀; 野々村, 賢一; 岩田, 伸夫

1997-01-01

Genomic in situ hybridization (GIS H) using total Oryza punctata Kotschy genomic DNA as a probe was applied to detect alien chromosomes transferred from O. punctata (W1514: 2n=2x=24: BB) to O. sativa Japonica cultivar, Nipponbare (2n=2x=24: AA). Only 12 chromosomes in the interspecific hybrids (2n=3x=36: AAB) between autotetraploid of O. sativa cultivar Nipponbare and a diploid strain of O. punctata (W1514) showed intense staining by FITC in mitotic metaphase spreads. Only one homologous pair...

Analysis of the Ceratitis capitata y chromosome using in situ hybridization to mitotic chromosomes

International Nuclear Information System (INIS)

Willhoeft, U.; Franz, G.

1998-01-01

In Ceratitis capitata the Y chromosome is responsible for sex-determination. We used fluorescence in situ hybridization (FISH) for cytogenetic analysis of mitotic chromosomes. FISH with the wild-type strain EgyptII and two repetitive DNA probes enabled us to differentiate between the short and the long arm of the Y chromosome and gives a much better resolution than C-banding of mitotic chromosomes. We identified the Y-chromosomal breakpoints in Y-autosome translocations using FISH. Even more complex rearrangements i.e. deletions and insertions in some translocation strains were detected by this method. A strategy for mapping the primary sex determination factor in Ceratitis capitata by FISH is presented. (author)

Numerically abnormal chromosome constitutions in humans

Energy Technology Data Exchange (ETDEWEB)

NONE

1993-12-31

Chapter 24, discusses numerically abnormal chromosome constitutions in humans. This involves abnormalities of human chromosome number, including polyploidy (when the number of sets of chromosomes increases) and aneuploidy (when the number of individual normal chromosomes changes). Chapter sections discuss the following chromosomal abnormalities: human triploids, imprinting and uniparental disomy, human tetraploids, hydatidiform moles, anomalies caused by chromosomal imbalance, 13 trisomy (D{sub 1} trisomy, Patau syndrome), 21 trisomy (Down syndrome), 18 trisomy syndrome (Edwards syndrome), other autosomal aneuploidy syndromes, and spontaneous abortions. The chapter concludes with remarks on the nonrandom participation of chromosomes in trisomy. 69 refs., 3 figs., 4 tabs.

A recurrent translocation is mediated by homologous recombination between HERV-H elements

Directory of Open Access Journals (Sweden)

Hermetz Karen E

2012-01-01

Full Text Available Abstract Background Chromosome rearrangements are caused by many mutational mechanisms; of these, recurrent rearrangements can be particularly informative for teasing apart DNA sequence-specific factors. Some recurrent translocations are mediated by homologous recombination between large blocks of segmental duplications on different chromosomes. Here we describe a recurrent unbalanced translocation casued by recombination between shorter homologous regions on chromosomes 4 and 18 in two unrelated children with intellectual disability. Results Array CGH resolved the breakpoints of the 6.97-Megabase (Mb loss of 18q and the 7.30-Mb gain of 4q. Sequencing across the translocation breakpoints revealed that both translocations occurred between 92%-identical human endogenous retrovirus (HERV elements in the same orientation on chromosomes 4 and 18. In addition, we find sequence variation in the chromosome 4 HERV that makes one allele more like the chromosome 18 HERV. Conclusions Homologous recombination between HERVs on the same chromosome is known to cause chromosome deletions, but this is the first report of interchromosomal HERV-HERV recombination leading to a translocation. It is possible that normal sequence variation in substrates of non-allelic homologous recombination (NAHR affects the alignment of recombining segments and influences the propensity to chromosome rearrangement.

Combining M-FISH and Quantum Dot technology for fast chromosomal assignment of transgenic insertions

Directory of Open Access Journals (Sweden)

Yusuf Mohammed

2011-12-01

Full Text Available Abstract Background Physical mapping of transgenic insertions by Fluorescence in situ Hybridization (FISH is a reliable and cost-effective technique. Chromosomal assignment is commonly achieved either by concurrent G-banding or by a multi-color FISH approach consisting of iteratively co-hybridizing the transgenic sequence of interest with one or more chromosome-specific probes at a time, until the location of the transgenic insertion is identified. Results Here we report a technical development for fast chromosomal assignment of transgenic insertions at the single cell level in mouse and rat models. This comprises a simplified 'single denaturation mixed hybridization' procedure that combines multi-color karyotyping by Multiplex FISH (M-FISH, for simultaneous and unambiguous identification of all chromosomes at once, and the use of a Quantum Dot (QD conjugate for the transgene detection. Conclusions Although the exploitation of the unique optical properties of QD nanocrystals, such as photo-stability and brightness, to improve FISH performance generally has been previously investigated, to our knowledge this is the first report of a purpose-designed molecular cytogenetic protocol in which the combined use of QDs and standard organic fluorophores is specifically tailored to assist gene transfer technology.

Charge transfer mediator based systems for electrocatalytic oxygen reduction

Science.gov (United States)

Stahl, Shannon S.; Gerken, James B.; Anson, Colin W.

2017-07-18

Disclosed are systems for the electrocatalytic reduction of oxygen, having redox mediator/redox catalyst pairs and an electrolyte solution in contact with an electrode. The redox mediator is included in the electrolyte solution, and the redox catalyst may be included in the electrolyte solution, or alternatively, may be in contact with the electrolyte solution. In one form a cobalt redox catalyst is used with a quinone redox mediator. In another form a nitrogen oxide redox catalyst is used with a nitroxyl type redox mediator. The systems can be used in electrochemical cells wherein neither the anode nor the cathode comprise an expensive metal such as platinum.

Charge transfer mediator based systems for electrocatalytic oxygen reduction

Energy Technology Data Exchange (ETDEWEB)

Stahl, Shannon S.; Gerken, James B.; Anson, Colin W.

2017-11-07

Disclosed are systems for the electrocatalytic reduction of oxygen, having redox mediator/redox catalyst pairs and an electrolyte solution in contact with an electrode. The redox mediator is included in the electrolyte solution, and the redox catalyst may be included in the electrolyte solution, or alternatively, may be in contact with the electrolyte solution. In one form a cobalt redox catalyst is used with a quinone redox mediator. In another form a nitrogen oxide redox catalyst is used with a nitroxyl type redox mediator. The systems can be used in electrochemical cells wherein neither the anode nor the cathode comprise an expensive metal such as platinum.

Origin of amphibian and avian chromosomes by fission, fusion, and retention of ancestral chromosomes

Science.gov (United States)

Voss, Stephen R.; Kump, D. Kevin; Putta, Srikrishna; Pauly, Nathan; Reynolds, Anna; Henry, Rema J.; Basa, Saritha; Walker, John A.; Smith, Jeramiah J.

2011-01-01

Amphibian genomes differ greatly in DNA content and chromosome size, morphology, and number. Investigations of this diversity are needed to identify mechanisms that have shaped the evolution of vertebrate genomes. We used comparative mapping to investigate the organization of genes in the Mexican axolotl (Ambystoma mexicanum), a species that presents relatively few chromosomes (n = 14) and a gigantic genome (>20 pg/N). We show extensive conservation of synteny between Ambystoma, chicken, and human, and a positive correlation between the length of conserved segments and genome size. Ambystoma segments are estimated to be four to 51 times longer than homologous human and chicken segments. Strikingly, genes demarking the structures of 28 chicken chromosomes are ordered among linkage groups defining the Ambystoma genome, and we show that these same chromosomal segments are also conserved in a distantly related anuran amphibian (Xenopus tropicalis). Using linkage relationships from the amphibian maps, we predict that three chicken chromosomes originated by fusion, nine to 14 originated by fission, and 12–17 evolved directly from ancestral tetrapod chromosomes. We further show that some ancestral segments were fused prior to the divergence of salamanders and anurans, while others fused independently and randomly as chromosome numbers were reduced in lineages leading to Ambystoma and Xenopus. The maintenance of gene order relationships between chromosomal segments that have greatly expanded and contracted in salamander and chicken genomes, respectively, suggests selection to maintain synteny relationships and/or extremely low rates of chromosomal rearrangement. Overall, the results demonstrate the value of data from diverse, amphibian genomes in studies of vertebrate genome evolution. PMID:21482624

Molecular fundamentals of chromosomal mutagenesis

International Nuclear Information System (INIS)

Ganassi, E.Eh.; Zaichkina, S.I.; Malakhova, L.V.

1987-01-01

Precise quantitative correlation between the yield of chromosome structure damages and the yield of DNA damages is shown when comparing data on molecular and cytogenetic investigations carried out in cultural Mammalia cells. As the chromosome structure damage is to be connected with the damage of its carcass structure, then it is natural that DNA damage in loop regions is not to affect considerably the structure, while DNA damage lying on the loop base and connected with the chromosome carcass is to play a determining role in chromosomal mutagenesis. This DNA constitutes 1-2% from the total quantity of nuclear DNA. If one accepts that damages of these regions of DNA are ''hot'' points of chromosomal mutagenesis, then it becomes clear why 1-2% of preparation damages in a cell are realized in chromosome structural damages

Chromosomal geometry in the interface from the frequency of the radiation induced chromosome aberrations

International Nuclear Information System (INIS)

Nasazzi, N.; Otero, D.; Di Giorgio, M.

1996-01-01

Ionizing radiation induces DNA double-strand breaks (DSBs) and their interaction and illegitimate recombination produces chromosomal aberrations. Stable chromosomal aberrations comprise inter-chromosomal events (translocations) and intra-chromosomal events (inversions). When DSBs induction and interaction is done at random, and the proximity effects are neglected, the expected relation between translocations and inversions is F=86, based on chromosome arm length. The number of translocations and inversions is analyzed by using G-banding in 16 lymphocytes cultures from blood samples acutely irradiated with γ-rays (dose range: 0,5 Gy - 3 Gy). The result obtained was: F=13,5, significantly smaller than F=86. Literature data show similar small F values, but strongly spread. The excess of inversions could be explained by a 'proximity effect', it means that more proximate DSBs have more interaction probability. Therefore, it is possible to postulate a special chromosome arrangement during irradiation and the subsequent interval. We propose a model where individual chromosomes show spherical confinement with some degree of overlapping and DSBs induction proportional to cross section. A DSBs interaction probability function with cut-off length= 1μ is assumed. According to our results, the confinement volume is ≅ 6.4% of the nuclear volume. Nevertheless, we presume that large spread in F data could be due to temporal variation in overlapping and spatial chromosomal confinement. (authors). 14 refs

Painting of fourth and chromosome-wide regulation of the 4th chromosome in Drosophila melanogaster.

Science.gov (United States)

Johansson, Anna-Mia; Stenberg, Per; Bernhardsson, Carolina; Larsson, Jan

2007-05-02

Drosophila melanogaster exhibits two expression-regulating systems that target whole, specific chromosomes: the dosage compensation system whereby the male-specific lethal complex doubles transcription of genes on the male X-chromosome and the chromosome 4-specific protein Painting of fourth, POF. POF is the first example of an autosome-specific protein and its presence raises the question of the universality of chromosome-specific regulation. Here we show that POF and heterochromatin protein 1 (HP1) are involved in the global regulation of the 4th chromosome. Contrary to previous conclusions, Pof is not essential for survival of diplo-4th karyotype flies. However, Pof is essential for survival of haplo-4th individuals and expression of chromosome 4 genes in diplo-4th individuals is decreased in the absence of Pof. Mapping of POF using chromatin immunoprecipitation suggested that it binds within genes. Furthermore, we show that POF binding is dependent on heterochromatin and that POF and HP1 bind interdependently to the 4th chromosome. We propose a balancing mechanism involving POF and HP1 that provides a feedback system for fine-tuning expression status of genes on the 4th chromosome.

Chromosome Territories

OpenAIRE

Cremer, Thomas; Cremer, Marion

2010-01-01

Chromosome territories (CTs) constitute a major feature of nuclear architecture. In a brief statement, the possible contribution of nuclear architecture studies to the field of epigenomics is considered, followed by a historical account of the CT concept and the final compelling experimental evidence of a territorial organization of chromosomes in all eukaryotes studied to date. Present knowledge of nonrandom CT arrangements, of the internal CT architecture, and of structural interactions wit...

Reversible immortalisation enables genetic correction of human muscle progenitors and engineering of next-generation human artificial chromosomes for Duchenne muscular dystrophy.

Science.gov (United States)

Benedetti, Sara; Uno, Narumi; Hoshiya, Hidetoshi; Ragazzi, Martina; Ferrari, Giulia; Kazuki, Yasuhiro; Moyle, Louise Anne; Tonlorenzi, Rossana; Lombardo, Angelo; Chaouch, Soraya; Mouly, Vincent; Moore, Marc; Popplewell, Linda; Kazuki, Kanako; Katoh, Motonobu; Naldini, Luigi; Dickson, George; Messina, Graziella; Oshimura, Mitsuo; Cossu, Giulio; Tedesco, Francesco Saverio

2018-02-01

Transferring large or multiple genes into primary human stem/progenitor cells is challenged by restrictions in vector capacity, and this hurdle limits the success of gene therapy. A paradigm is Duchenne muscular dystrophy (DMD), an incurable disorder caused by mutations in the largest human gene: dystrophin. The combination of large-capacity vectors, such as human artificial chromosomes (HACs), with stem/progenitor cells may overcome this limitation. We previously reported amelioration of the dystrophic phenotype in mice transplanted with murine muscle progenitors containing a HAC with the entire dystrophin locus (DYS-HAC). However, translation of this strategy to human muscle progenitors requires extension of their proliferative potential to withstand clonal cell expansion after HAC transfer. Here, we show that reversible cell immortalisation mediated by lentivirally delivered excisable hTERT and Bmi1 transgenes extended cell proliferation, enabling transfer of a novel DYS-HAC into DMD satellite cell-derived myoblasts and perivascular cell-derived mesoangioblasts. Genetically corrected cells maintained a stable karyotype, did not undergo tumorigenic transformation and retained their migration ability. Cells remained myogenic in vitro (spontaneously or upon MyoD induction) and engrafted murine skeletal muscle upon transplantation. Finally, we combined the aforementioned functions into a next-generation HAC capable of delivering reversible immortalisation, complete genetic correction, additional dystrophin expression, inducible differentiation and controllable cell death. This work establishes a novel platform for complex gene transfer into clinically relevant human muscle progenitors for DMD gene therapy. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

Chromosomal Evolution in Chiroptera.

Science.gov (United States)

Sotero-Caio, Cibele G; Baker, Robert J; Volleth, Marianne

2017-10-13

Chiroptera is the second largest order among mammals, with over 1300 species in 21 extant families. The group is extremely diverse in several aspects of its natural history, including dietary strategies, ecology, behavior and morphology. Bat genomes show ample chromosome diversity (from 2n = 14 to 62). As with other mammalian orders, Chiroptera is characterized by clades with low, moderate and extreme chromosomal change. In this article, we will discuss trends of karyotypic evolution within distinct bat lineages (especially Phyllostomidae, Hipposideridae and Rhinolophidae), focusing on two perspectives: evolution of genome architecture, modes of chromosomal evolution, and the use of chromosome data to resolve taxonomic problems.

Chromosomal Evolution in Chiroptera

Directory of Open Access Journals (Sweden)

Cibele G. Sotero-Caio

2017-10-01

Full Text Available Chiroptera is the second largest order among mammals, with over 1300 species in 21 extant families. The group is extremely diverse in several aspects of its natural history, including dietary strategies, ecology, behavior and morphology. Bat genomes show ample chromosome diversity (from 2n = 14 to 62. As with other mammalian orders, Chiroptera is characterized by clades with low, moderate and extreme chromosomal change. In this article, we will discuss trends of karyotypic evolution within distinct bat lineages (especially Phyllostomidae, Hipposideridae and Rhinolophidae, focusing on two perspectives: evolution of genome architecture, modes of chromosomal evolution, and the use of chromosome data to resolve taxonomic problems.

GSK-3 inhibitors induce chromosome instability

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Staples Oliver D

2007-08-01

Full Text Available Abstract Background Several mechanisms operate during mitosis to ensure accurate chromosome segregation. However, during tumour evolution these mechanisms go awry resulting in chromosome instability. While several lines of evidence suggest that mutations in adenomatous polyposis coli (APC may promote chromosome instability, at least in colon cancer, the underlying mechanisms remain unclear. Here, we turn our attention to GSK-3 – a protein kinase, which in concert with APC, targets β-catenin for proteolysis – and ask whether GSK-3 is required for accurate chromosome segregation. Results To probe the role of GSK-3 in mitosis, we inhibited GSK-3 kinase activity in cells using a panel of small molecule inhibitors, including SB-415286, AR-A014418, 1-Azakenpaullone and CHIR99021. Analysis of synchronised HeLa cells shows that GSK-3 inhibitors do not prevent G1/S progression or cell division. They do, however, significantly delay mitotic exit, largely because inhibitor-treated cells have difficulty aligning all their chromosomes. Although bipolar spindles form and the majority of chromosomes biorient, one or more chromosomes often remain mono-oriented near the spindle poles. Despite a prolonged mitotic delay, anaphase frequently initiates without the last chromosome aligning, resulting in chromosome non-disjunction. To rule out the possibility of "off-target" effects, we also used RNA interference to selectively repress GSK-3β. Cells deficient for GSK-3β exhibit a similar chromosome alignment defect, with chromosomes clustered near the spindle poles. GSK-3β repression also results in cells accumulating micronuclei, a hallmark of chromosome missegregation. Conclusion Thus, not only do our observations indicate a role for GSK-3 in accurate chromosome segregation, but they also raise the possibility that, if used as therapeutic agents, GSK-3 inhibitors may induce unwanted side effects by inducing chromosome instability.

Dielectrophoretic manipulation of human chromosomes in microfluidic channels: extracting chromosome dielectric properties

DEFF Research Database (Denmark)

Clausen, Casper Hyttel; Dimaki, Maria; Buckley, Sonia

2011-01-01

An investigation of the dielectric properties of polyamine buffer prepared human chromosomes is presented in this paper. Chromosomes prepared in this buffer are only a few micrometers in size and shaped roughly like spherical discs. Dielectrophoresis was therefore chosen as the method...... of manipulation combined with a custom designed microfluidic system containing the required electrodes for dielectrophoresis experiments. Our results show that although this system is presently not able to distinguish between the different chromosomes, it can provide average data for the dielectric properties...... of human chromosomes in polyamine buffer. These can then be used to optimize system designs for further characterization and even sorting. The experimental data from the dielectrophoretic manipulation were combined with theoretical calculations to extract a range of values for the permittivity...

Mobilisation and remobilisation of a large archetypal pathogenicity island of uropathogenic Escherichia coli in vitro support the role of conjugation for horizontal transfer of genomic islands

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Hochhut Bianca

2011-09-01

Full Text Available Abstract Background A substantial amount of data has been accumulated supporting the important role of genomic islands (GEIs - including pathogenicity islands (PAIs - in bacterial genome plasticity and the evolution of bacterial pathogens. Their instability and the high level sequence similarity of different (partial islands suggest an exchange of PAIs between strains of the same or even different bacterial species by horizontal gene transfer (HGT. Transfer events of archetypal large genomic islands of enterobacteria which often lack genes required for mobilisation or transfer have been rarely investigated so far. Results To study mobilisation of such large genomic regions in prototypic uropathogenic E. coli (UPEC strain 536, PAI II536 was supplemented with the mobRP4 region, an origin of replication (oriVR6K, an origin of transfer (oriTRP4 and a chloramphenicol resistance selection marker. In the presence of helper plasmid RP4, conjugative transfer of the 107-kb PAI II536 construct occured from strain 536 into an E. coli K-12 recipient. In transconjugants, PAI II536 existed either as a cytoplasmic circular intermediate (CI or integrated site-specifically into the recipient's chromosome at the leuX tRNA gene. This locus is the chromosomal integration site of PAI II536 in UPEC strain 536. From the E. coli K-12 recipient, the chromosomal PAI II536 construct as well as the CIs could be successfully remobilised and inserted into leuX in a PAI II536 deletion mutant of E. coli 536. Conclusions Our results corroborate that mobilisation and conjugal transfer may contribute to evolution of bacterial pathogens through horizontal transfer of large chromosomal regions such as PAIs. Stabilisation of these mobile genetic elements in the bacterial chromosome result from selective loss of mobilisation and transfer functions of genomic islands.

Macronuclear genome structure of the ciliate Nyctotherus ovalis: Single-gene chromosomes and tiny introns

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Landweber Laura F

2008-12-01

Full Text Available Abstract Background Nyctotherus ovalis is a single-celled eukaryote that has hydrogen-producing mitochondria and lives in the hindgut of cockroaches. Like all members of the ciliate taxon, it has two types of nuclei, a micronucleus and a macronucleus. N. ovalis generates its macronuclear chromosomes by forming polytene chromosomes that subsequently develop into macronuclear chromosomes by DNA elimination and rearrangement. Results We examined the structure of these gene-sized macronuclear chromosomes in N. ovalis. We determined the telomeres, subtelomeric regions, UTRs, coding regions and introns by sequencing a large set of macronuclear DNA sequences (4,242 and cDNAs (5,484 and comparing them with each other. The telomeres consist of repeats CCC(AAAACCCCn, similar to those in spirotrichous ciliates such as Euplotes, Sterkiella (Oxytricha and Stylonychia. Per sequenced chromosome we found evidence for either a single protein-coding gene, a single tRNA, or the complete ribosomal RNAs cluster. Hence the chromosomes appear to encode single transcripts. In the short subtelomeric regions we identified a few overrepresented motifs that could be involved in gene regulation, but there is no consensus polyadenylation site. The introns are short (21–29 nucleotides, and a significant fraction (1/3 of the tiny introns is conserved in the distantly related ciliate Paramecium tetraurelia. As has been observed in P. tetraurelia, the N. ovalis introns tend to contain in-frame stop codons or have a length that is not dividable by three. This pattern causes premature termination of mRNA translation in the event of intron retention, and potentially degradation of unspliced mRNAs by the nonsense-mediated mRNA decay pathway. Conclusion The combination of short leaders, tiny introns and single genes leads to very minimal macronuclear chromosomes. The smallest we identified contained only 150 nucleotides.

Frequent gene conversion events between the X and Y homologous chromosomal regions in primates

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Hirai Hirohisa

2010-07-01

Full Text Available Abstract Background Mammalian sex-chromosomes originated from a pair of autosomes. A step-wise cessation of recombination is necessary for the proper maintenance of sex-determination and, consequently, generates a four strata structure on the X chromosome. Each stratum shows a specific per-site nucleotide sequence difference (p-distance between the X and Y chromosomes, depending on the time of recombination arrest. Stratum 4 covers the distal half of the human X chromosome short arm and the p-distance of the stratum is ~10%, on average. However, a 100-kb region, which includes KALX and VCX, in the middle of stratum 4 shows a significantly lower p-distance (1-5%, suggesting frequent sequence exchanges or gene conversions between the X and Y chromosomes in humans. To examine the evolutionary mechanism for this low p-distance region, sequences of a corresponding region including KALX/Y from seven species of non-human primates were analyzed. Results Phylogenetic analysis of this low p-distance region in humans and non-human primate species revealed that gene conversion like events have taken place at least ten times after the divergence of New World monkeys and Catarrhini (i.e., Old World monkeys and hominoids. A KALY-converted KALX allele in white-handed gibbons also suggests a possible recent gene conversion between the X and Y chromosomes. In these primate sequences, the proximal boundary of this low p-distance region is located in a LINE element shared between the X and Y chromosomes, suggesting the involvement of this element in frequent gene conversions. Together with a palindrome on the Y chromosome, a segmental palindrome structure on the X chromosome at the distal boundary near VCX, in humans and chimpanzees, may mediate frequent sequence exchanges between X and Y chromosomes. Conclusion Gene conversion events between the X and Y homologous regions have been suggested, mainly in humans. Here, we found frequent gene conversions in the

Chromatin Structure and Replication Origins: Determinants Of Chromosome Replication And Nuclear Organization

Science.gov (United States)

Smith, Owen K.; Aladjem, Mirit I.

2014-01-01

The DNA replication program is, in part, determined by the epigenetic landscape that governs local chromosome architecture and directs chromosome duplication. Replication must coordinate with other biochemical processes occurring concomitantly on chromatin, such as transcription and remodeling, to insure accurate duplication of both genetic and epigenetic features and to preserve genomic stability. The importance of genome architecture and chromatin looping in coordinating cellular processes on chromatin is illustrated by two recent sets of discoveries. First, chromatin-associated proteins that are not part of the core replication machinery were shown to affect the timing of DNA replication. These chromatin-associated proteins could be working in concert, or perhaps in competition, with the transcriptional machinery and with chromatin modifiers to determine the spatial and temporal organization of replication initiation events. Second, epigenetic interactions are mediated by DNA sequences that determine chromosomal replication. In this review we summarize recent findings and current models linking spatial and temporal regulation of the replication program with epigenetic signaling. We discuss these issues in the context of the genome’s three-dimensional structure with an emphasis on events occurring during the initiation of DNA replication. PMID:24905010

Mechanisms of ring chromosome formation in 11 cases of human ring chromosome 21

DEFF Research Database (Denmark)

McGinniss, M J; Kazazian, H H; Stetten, G

1992-01-01

We studied the mechanism of ring chromosome 21 (r(21)) formation in 13 patients (11 unique r(21)s), consisting of 7 from five families with familial r(21) and 6 with de novo r(21). The copy number of chromosome 21 sequences in the rings of these patients was determined by quantitative dosage......), resulting in deletion of varying amounts of 21q22.1 to 21qter. The data from one individual who had a Down syndrome phenotype were consistent with asymmetric breakage and reunion of 21q sequences from an intermediate isochromosome or Robertsonian translocation chromosome as reported by Wong et al. Another......). The phenotype of patients correlated well with the extent of deletion or duplication of chromosome 21 sequences. These data demonstrate three mechanisms of r(21) formation and show that the phenotype of r(21) patients varies with the extent of chromosome 21 monosomy or trisomy....

Appearance and evolution of the specific chromosomal rearrangements associated with malignant transformation of mouse m5S cells

International Nuclear Information System (INIS)

Kodama, S.; Okumura, Y.; Komatsu, K.; Sasaki, M.S.

1991-01-01

Chromosomal alterations were studied during the acquisition of malignant phenotypes in two karyotypically distinct cells isolated from transformed foci induced by x-irradiation in mouse m5S cells. Because the transformants, despite foci origin, showed low ability to grow in agar, they were cultured in vitro with serial transfer schedules to allow further cell generations and assayed for anchorage independence (AI) at each passage level. The AI frequency increased with the cell doubling numbers. Chromosome analysis showed that a focus was one cell origin, but the transformants showed karyotypic instability during cell proliferation, giving rise to the rearrangements clustered in the distal region of the specific chromosomes. These rearrangements appeared to be directed toward the acquisition of malignant phenotypes. Analysis of the types and sites of rearrangements indicated that a mechanism exists that induces frequent rearrangements of the specific region of a chromosome during the process of transformation into the malignant state

Sexually antagonistic "zygotic drive" of the sex chromosomes.

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William R Rice

2008-12-01

Full Text Available Genomic conflict is perplexing because it causes the fitness of a species to decline rather than improve. Many diverse forms of genomic conflict have been identified, but this extant tally may be incomplete. Here, we show that the unusual characteristics of the sex chromosomes can, in principle, lead to a previously unappreciated form of sexual genomic conflict. The phenomenon occurs because there is selection in the heterogametic sex for sex-linked mutations that harm the sex of offspring that does not carry them, whenever there is competition among siblings. This harmful phenotype can be expressed as an antagonistic green-beard effect that is mediated by epigenetic parental effects, parental investment, and/or interactions among siblings. We call this form of genomic conflict sexually antagonistic "zygotic drive", because it is functionally equivalent to meiotic drive, except that it operates during the zygotic and postzygotic stages of the life cycle rather than the meiotic and gametic stages. A combination of mathematical modeling and a survey of empirical studies is used to show that sexually antagonistic zygotic drive is feasible, likely to be widespread in nature, and that it can promote a genetic "arms race" between the homo- and heteromorphic sex chromosomes. This new category of genomic conflict has the potential to strongly influence other fundamental evolutionary processes, such as speciation and the degeneration of the Y and W sex chromosomes. It also fosters a new genetic hypothesis for the evolution of enigmatic fitness-reducing traits like the high frequency of spontaneous abortion, sterility, and homosexuality observed in humans.

Electrochemical Water Oxidation and Stereoselective Oxygen Atom Transfer Mediated by a Copper Complex.

Science.gov (United States)

Kafentzi, Maria-Chrysanthi; Papadakis, Raffaello; Gennarini, Federica; Kochem, Amélie; Iranzo, Olga; Le Mest, Yves; Le Poul, Nicolas; Tron, Thierry; Faure, Bruno; Simaan, A Jalila; Réglier, Marius

2018-04-06

Water oxidation by copper-based complexes to form dioxygen has attracted attention in recent years, with the aim of developing efficient and cheap catalysts for chemical energy storage. In addition, high-valent metal-oxo species produced by the oxidation of metal complexes in the presence of water can be used to achieve substrate oxygenation with the use of H 2 O as an oxygen source. To date, this strategy has not been reported for copper complexes. Herein, a copper(II) complex, [(RPY2)Cu(OTf) 2 ] (RPY2=N-substituted bis[2-pyridyl(ethylamine)] ligands; R=indane; OTf=triflate), is used. This complex, which contains an oxidizable substrate moiety (indane), is used as a tool to monitor an intramolecular oxygen atom transfer reaction. Electrochemical properties were investigated and, upon electrolysis at 1.30 V versus a normal hydrogen electrode (NHE), both dioxygen production and oxygenation of the indane moiety were observed. The ligand was oxidized in a highly diastereoselective manner, which indicated that the observed reactivity was mediated by metal-centered reactive species. The pH dependence of the reactivity was monitored and correlated with speciation deduced from different techniques, ranging from potentiometric titrations to spectroscopic studies and DFT calculations. Water oxidation for dioxygen production occurs at neutral pH and is probably mediated by the oxidation of a mononuclear copper(II) precursor. It is achieved with a rather low overpotential (280 mV at pH 7), although with limited efficiency. On the other hand, oxygenation is maximum at pH 8-8.5 and is probably mediated by the electrochemical oxidation of an antiferromagnetically coupled dinuclear bis(μ-hydroxo) copper(II) precursor. This constitutes the first example of copper-centered oxidative water activation for a selective oxygenation reaction. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Chromosome analysis of arsenic affected cattle

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S. Shekhar

2014-10-01

Full Text Available Aim: The aim was to study the chromosome analysis of arsenic affected cattle. Materials and Methods: 27 female cattle (21 arsenic affected and 6 normal were selected for cytogenetical study. The blood samples were collected, incubated, and cultured using appropriate media and specific methods. The samples were analyzed for chromosome number and morphology, relative length of the chromosome, arm ratio, and centromere index of X chromosome and chromosomal abnormalities in arsenic affected cattle to that of normal ones. Results: The diploid number of metaphase chromosomes in arsenic affected cattle as well as in normal cattle were all 2n=60, 58 being autosomes and 2 being sex chromosomes. From the centromeric position, karyotyping studies revealed that all the 29 pair of autosomes was found to be acrocentric or telocentric, and the sex chromosomes (XX were submetacentric in both normal and arsenic affected cattle. The relative length of all the autosome pairs and sex chrosomosome pair was found to be higher in normal than that of arsenic affected cattle. The mean arm ratio of X-chromosome was higher in normal than that of arsenic affected cattle, but it is reverse in case of centromere index value of X-chromosome. There was no significant difference of arm ratio and centromere index of X-chromosomes between arsenic affected and normal cattle. No chromosomal abnormalities were found in arsenic affected cattle. Conclusion: The chromosome analysis of arsenic affected cattle in West Bengal reported for the first time in this present study which may serve as a guideline for future studies in other species. These reference values will also help in comparison of cytological studies of arsenic affected cattle to that of various toxicants.

Effect of genomic location on horizontal transfer of a recombinant gene cassette between Pseudomonas strains in the rhizosphere and spermosphere of barley seedlings

DEFF Research Database (Denmark)

Sengelov, G.; Kristensen, K. J.; Sørensen, Anders Morten Hay

2001-01-01

, horizontal transfer of a recombinant gene cassette inserted into the chromosome of a Pseudomonas strutzeri strain, into a mobilizable plasmid (pAGM42), and into a conjugative plasmid (pKJK5) isolated from barley rhizosphere was investigated. Horizontal transfer efficiencies of the gene cassette inserted...... efficiencies were up to 4.36 x 10(-3) transconjugants/(donors x recipients)(1/2). Transfer of chromosomal encoded genes could not be detected in the microcosms by conjugation or transformation. However, transformation did occur by using the same bacterial strains under laboratory conditions. The rhizosphere...

Enabling fast electron transfer through both bacterial outer-membrane redox centers and endogenous electron mediators by polyaniline hybridized large-mesoporous carbon anode for high-performance microbial fuel cells

International Nuclear Information System (INIS)

Zou, Long; Qiao, Yan; Zhong, Canyu; Li, Chang Ming

2017-01-01

Both physical structure and chemical property of an electrode play critical roles in extracellular electron transfer from microbes to electrodes in microbial fuel cells (MFCs). Herein a novel polyaniline hybridized large mesoporous carbon (PANI-LMC) anode is fabricated from natural biomass by nanostructured CaCO 3 template-assisted carbonization followed by in situ chemical polymerizing PANI to enable fast extracellular electron transfer, in which the LMC with rich disorder-interconnected large mesopores (∼20−50 nm) and large surface area facilitates a fast mediated electron transfer through electron mediators, while the decorated PANI on LMC surface enables the direct electron transfer via bacterial outer-membrane redox centers. Owing to the unique synergistic effect from both excellent electron transfer paths, the PANI-LMC hybrid anode harvests high power electricity with a maximum output power density of 1280 mW m −2 in Shewanella putrefaciens CN32 MFCs, 10-fold higher than that of conventional carbon cloth. The findings from this work suggest a new insight on design of high-efficient anode according to the multiple and flexible electrochemical process for practical MFC applications.

Chromosome heteromorphisms in the Japanese, 3

International Nuclear Information System (INIS)

Sofuni, Toshio; Awa, A.A.

1982-12-01

The type and frequency of chromosome variants detected by the C-staining method were ascertained in 1,857 individuals residing in Hiroshima. The most frequent heteromorphic variant was the total inversion of the C-band in chromosome 9 found in 27 individuals (1.45%). The total inversion of the C-band in chromosome 1 was not seen in this sample, but the partial inversion of the C-band in chromosome 1 was found in 18 persons (0.97%). Partial inversion was also detected in the C-band in chromosome 9 in 22 individuals (1.18%). In chromosome 16, neither total nor partial inversion of the C-band was observed in the present study. The frequencies of chromosomes 1, 9, and 16 with a very large C-band were 0.70%, 0.22%, and 0.54%, respectively. Aside from these (1, 9, and 16) a very large C-band was found occasionally in chromosomes 4, 5, 6, 11, 12, 14, and 15, and an unusual insertion of the Y chromosome was observed. A total of 128 C-band variants (6.89%) was found in the 1,857 Hiroshima residents. (author)

Semi-automatic laser beam microdissection of the Y chromosome and analysis of Y chromosome DNA in a dioecious plant, Silene latifolia

International Nuclear Information System (INIS)

Matsunaga, S.; Kawano, S.; Michimoto, T.; Higashiyama, T.; Nakao, S.; Sakai, A.; Kuroiwa, T.

1999-01-01

Silene latifolia has heteromorphic sex chromosomes, the X and Y chromosomes. The Y chromosome, which is thought to carry the male determining gene, was isolated by UV laser microdissection and amplified by degenerate oligonucleotide-primed PCR. In situ chromosome suppression of the amplified Y chromosome DNA in the presence of female genomic DNA as a competitor showed that the microdissected Y chromosome DNA did not specifically hybridize to the Y chromosome, but-hybridized to all chromosomes. This result suggests that the Y chromosome does not contain Y chromosome-enriched repetitive sequences. A repetitive sequence in the microdissected Y chromosome, RMY1, was isolated while screening repetitive sequences in the amplified Y chromosome. Part of the nucleotide sequence shared a similarity to that of X-43.1, which was isolated from microdissected X chromosomes. Since fluorescence in situ hybridization analysis with RMY1 demonstrated that RMY1 was localized at the ends of the chromosome, RMY1 may be a subtelomeric repetitive sequence. Regarding the sex chromosomes, RMY1 was detected at both ends of the X chromosome and at one end near the pseudoautosomal region of the Y chromosome. The different localization of RMY1 on the sex chromosomes provides a clue to the problem of how the sex chromosomes arose from autosomes

Problems with multiple use of transfer buffer in protein electrophoretic transfer.

Science.gov (United States)

Dorri, Yaser; Kurien, Biji T; Scofield, R Hal

2010-04-01

Two-dimensional gel electrophoresis (2DE) and SDS-PAGE are the two most useful methods in protein separation. Proteins separated by 2DE or SDS-PAGE are usually transferred to membranes using a variety of methods, such as electrophoretic transfer, heat-mediated transfer, or nonelectrophoretic transfer, for specific protein detection and/or analysis. In a recent study, Pettegrew et al. claim to reuse transfer buffer containing methanol for at least five times for transferring proteins from SDS-PAGE to polyvinylidene difluoride. They add 150-200 ml fresh transfer solution each time for extended use as a result of loss of transfer buffer. Finally, they test efficiency of each protein transfer by chemiluminescence detection. Here, we comment on this report, as we believe this method is not accurate and useful for protein analysis, and it can cause background binding as well as inaccurate protein analysis.

Radioiodine uptake of undifferentiated thyroid cancer cells by adenovirus-mediated Na+/ I- symporter gene transfer

Energy Technology Data Exchange (ETDEWEB)

So, Y.; Lee, Y. J.; Shin, J. H.; Oh, H. J.; Chung, J. K.; Lee, M. C.; Cho, B. Y. [College of Medicine, Univ. of Seoul National, Seoul (Korea, Republic of); Lee, K. H. [Samsung Medical Center, Seoul (Korea, Republic of)

2003-07-01

To increase radioiodine uptake on undifferentiated thyroid cancer cell (ARO cells) by adenovirus-mediated human Na+/I- symporter (hNIS) gene transfer. Recombinant adenovirus Ad-hNIS was manufactured successfully. After transfecting Ad-hNIS on ARO cells, in vitro I-125 uptake and efflux studies were performed. For in vivo studies, 1.510'8 p.f.u. (50 1) of Ad-hNIS was injected into xenograft ARO tumors on the R thigh of BALB/c nu/nu mice (n=12), and same amount of normal saline was injected into xenograft ARO tumors on the L thigh. Two, 3, 4 and 6 days after intratumoral injection of Ad-hNIS, I-131 images (3 mice per day) were taken and xenograft tumors on both thighs were all excised. Total RNA was extracted from each tumor tissue and RT-PCR was performed to confirm the hNIS expression of Ad-hNIS injected xenograft ARO tumors. I-125 uptake of Ad-hNIS transfected ARO cells was increased up to 233 folds at 120 minutes in vitro. I-125 efflux study revealed rapid washout of I-125 from Ad-hNIS transfected ARO cells. On dynamic image, I-131 uptake of Ad-hNIS injected ARO tumor was continuously increased until 60 minutes. Mean count ratios of xenograft ARO tumors (R/L) of 60 minutes I-131 images at 2, 3, 4 and 6 days after Ad-hNIS injection were 2.85, 2.54, 2.31, and 2.18, each. On RT-PCR, hNIS expression of Ad-hNIS transfected ARO xenograft tumors was confirmed. Radioiodine uptake was successfully increased in ARO cells by adenovirus-mediated hNIs gene transfer both in vitro and in vivo.

Electron transfer capacity dependence of quinone-mediated Fe(III) reduction and current generation by Klebsiella pneumoniae L17.

Science.gov (United States)

Li, Xiaomin; Liu, Liang; Liu, Tongxu; Yuan, Tian; Zhang, Wei; Li, Fangbai; Zhou, Shungui; Li, Yongtao

2013-06-01

Quinone groups in exogenous electron shuttles can accelerate extracellular electron transfer (EET) from bacteria to insoluble terminal electron acceptors, such as Fe(III) oxides and electrodes, which are important in biogeochemical redox processes and microbial electricity generation. However, the relationship between quinone-mediated EET performance and electron-shuttling properties of the quinones remains incompletely characterized. This study investigates the effects of a series of synthetic quinones (SQs) on goethite reduction and current generation by a fermenting bacterium Klebsiella pneumoniae L17. In addition, the voltammetric behavior and electron transfer capacities (ETCs) of SQ, including electron accepting (EAC) and donating (EDC) capacities, is also examined using electrochemical methods. The results showed that SQ can significantly increase both the Fe(III) reduction rates and current outputs of L17. Each tested SQ reversibly accepted and donated electrons as indicated by the cyclic voltammograms. The EAC and EDC results showed that Carmine and Alizarin had low relative capacities of electron transfer, whereas 9,10-anthraquinone-2,6-disulfonic acid (AQDS), 2-hydroxy-1,4-naphthoquinone (2-HNQ), and 5-hydroxy-1,4-naphthoquinone (5-HNQ) showed stronger relative ETC, and 9,10-anthraquinone-2-carboxylic acid (AQC) and 9,10-anthraquinone-2-sulfonic acid (AQS) had high relative ETC. Enhancement of microbial goethite reduction kinetics and current outputs by SQ had a good linear relationship with their ETC, indicating that the effectiveness of quinone-mediated EET may be strongly dependent on the ETC of the quinones. Therefore, the presence of quinone compounds and fermenting microorganisms may increase the diversity of microbial populations that contribute to element transformation in natural environments. Moreover, ETC determination of different SQ would help to evaluate their performance for microbial EET under anoxic conditions. Copyright © 2013 Elsevier

Chromosomal instability induced by ionizing radiation

International Nuclear Information System (INIS)

Morgan, W.F.; Marder, B.A.; Day, J.P.

1995-01-01

There is accumulating evidence indicating genomic instability can manifest multiple generations after cellular exposure to DNA damaging agents. For instance, some cells surviving exposure to ionizing radiations show delayed reproductive cell death, delayed mutation and / or delayed chromosomal instability. Such instability, especially chromosome destabilization has been implicated in mutation, gene amplification, cellular transformation, and cell killing. To investigate chromosomal instability following DNA damage, we have used fluorescence in situ hybridization to detect chromosomal rearrangements in a human/hamster somatic hybrid cell line following exposure to ionizing radiation. Delayed chromosomal instability was detected when multiple populations of uniquely arranged metaphases were observed in clonal isolates raised from single cells. The relationship between delayed chromosomal destabilization and other endpoints of genomic instability, namely; delayed mutation and gene amplification will be discussed, as will the potential cytogenetic and molecular mechanisms contributing to delayed chromosomal instability

Unique mosaicism of structural chromosomal rearrangement: is chromosome 18 preferentially involved?

NARCIS (Netherlands)

Pater, J.M. de; Smeets, D.F.C.M.; Scheres, J.M.J.C.

2003-01-01

The mentally normal mother of a 4-year-old boy with del(18)(q21.3) syndrome was tested cytogenetically to study the possibility of an inherited structural rearrangement of chromosome 18. She was found to carry an unusual mosaicism involving chromosomes 18 and 21. Two unbalanced cell lines were seen

B chromosomes are associated with redistribution of genetic recombination towards lower recombination chromosomal regions in perennial ryegrass.

Science.gov (United States)

Harper, John; Phillips, Dylan; Thomas, Ann; Gasior, Dagmara; Evans, Caron; Powell, Wayne; King, Julie; King, Ian; Jenkins, Glyn; Armstead, Ian

2018-04-09

Supernumerary 'B' chromosomes are non-essential components of the genome present in a range of plant and animal species-including many grasses. Within diploid and polyploid ryegrass and fescue species, including the forage grass perennial ryegrass (Lolium perenne L.), the presence of B chromosomes has been reported as influencing both chromosome pairing and chiasma frequencies. In this study, the effects of the presence/absence of B chromosomes on genetic recombination has been investigated through generating DArT (Diversity Arrays Technology) marker genetic maps for six perennial ryegrass diploid populations, the pollen parents of which contained either two B or zero B chromosomes. Through genetic and cytological analyses of these progeny and their parents, we have identified that, while overall cytological estimates of chiasma frequencies were significantly lower in pollen mother cells with two B chromosomes as compared with zero B chromosomes, the recombination frequencies within some marker intervals were actually increased, particularly for marker intervals in lower recombination regions of chromosomes, namely pericentromeric regions. Thus, in perennial ryegrass, the presence of two B chromosomes redistributed patterns of meiotic recombination in pollen mother cells in ways which could increase the range of allelic variation available to plant breeders.

Activating RNAs associate with Mediator to enhance chromatin architecture and transcription.

Science.gov (United States)

Lai, Fan; Orom, Ulf A; Cesaroni, Matteo; Beringer, Malte; Taatjes, Dylan J; Blobel, Gerd A; Shiekhattar, Ramin

2013-02-28

Recent advances in genomic research have revealed the existence of a large number of transcripts devoid of protein-coding potential in multiple organisms. Although the functional role for long non-coding RNAs (lncRNAs) has been best defined in epigenetic phenomena such as X-chromosome inactivation and imprinting, different classes of lncRNAs may have varied biological functions. We and others have identified a class of lncRNAs, termed ncRNA-activating (ncRNA-a), that function to activate their neighbouring genes using a cis-mediated mechanism. To define the precise mode by which such enhancer-like RNAs function, we depleted factors with known roles in transcriptional activation and assessed their role in RNA-dependent activation. Here we report that depletion of the components of the co-activator complex, Mediator, specifically and potently diminished the ncRNA-induced activation of transcription in a heterologous reporter assay using human HEK293 cells. In vivo, Mediator is recruited to ncRNA-a target genes and regulates their expression. We show that ncRNA-a interact with Mediator to regulate its chromatin localization and kinase activity towards histone H3 serine 10. The Mediator complex harbouring disease- displays diminished ability to associate with activating ncRNAs. Chromosome conformation capture confirmed the presence of DNA looping between the ncRNA-a loci and its targets. Importantly, depletion of Mediator subunits or ncRNA-a reduced the chromatin looping between the two loci. Our results identify the human Mediator complex as the transducer of activating ncRNAs and highlight the importance of Mediator and activating ncRNA association in human disease.

Statistics for X-chromosome associations.

Science.gov (United States)

Özbek, Umut; Lin, Hui-Min; Lin, Yan; Weeks, Daniel E; Chen, Wei; Shaffer, John R; Purcell, Shaun M; Feingold, Eleanor

2018-06-13

In a genome-wide association study (GWAS), association between genotype and phenotype at autosomal loci is generally tested by regression models. However, X-chromosome data are often excluded from published analyses of autosomes because of the difference between males and females in number of X chromosomes. Failure to analyze X-chromosome data at all is obviously less than ideal, and can lead to missed discoveries. Even when X-chromosome data are included, they are often analyzed with suboptimal statistics. Several mathematically sensible statistics for X-chromosome association have been proposed. The optimality of these statistics, however, is based on very specific simple genetic models. In addition, while previous simulation studies of these statistics have been informative, they have focused on single-marker tests and have not considered the types of error that occur even under the null hypothesis when the entire X chromosome is scanned. In this study, we comprehensively tested several X-chromosome association statistics using simulation studies that include the entire chromosome. We also considered a wide range of trait models for sex differences and phenotypic effects of X inactivation. We found that models that do not incorporate a sex effect can have large type I error in some cases. We also found that many of the best statistics perform well even when there are modest deviations, such as trait variance differences between the sexes or small sex differences in allele frequencies, from assumptions. © 2018 WILEY PERIODICALS, INC.

Conversion and non-conversion approach to preimplantation diagnosis for chromosomal rearrangements in 475 cycles.

Science.gov (United States)

Kuliev, Anver; Janzen, Jeanine Cieslak; Zlatopolsky, Zev; Kirillova, Irina; Ilkevitch, Yury; Verlinsky, Yury

2010-07-01

Due to the limitations of preimplantation genetic diagnosis (PGD) for chromosomal rearrangements by interphase fluorescent in-situ hybridization (FISH) analysis, a method for obtaining chromosomes from single blastomeres was introduced by their fusion with enucleated or intact mouse zygotes, followed by FISH analysis of the resulting heterokaryons. Although this allowed a significant improvement in the accuracy of testing of both maternally and paternally derived translocations, it is still labour intensive and requires the availability of fertilized mouse oocytes, also creating ethical issues related to the formation of interspecies heterokaryons. This method was modified with a chemical conversion procedure that has now been clinically applied for the first time on 877 embryos from PGD cycles for chromosomal rearrangements and has become the method of choice for performing PGD for structural rearrangements. This is presented within the context of overall experience of 475 PGD cycles for translocations with pre-selection and transfer of balanced or normal embryos in 342 (72%) of these cycles, which resulted in 131 clinical pregnancies (38%), with healthy deliveries of 113 unaffected children. The spontaneous abortion rate in these cycles was as low as 17%, which confirms an almost five-fold reduction of spontaneous abortion rate following PGD for chromosomal rearrangements. 2010 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

A novel system for simultaneous or sequential integration of multiple gene-loading vectors into a defined site of a human artificial chromosome.

Science.gov (United States)

Suzuki, Teruhiko; Kazuki, Yasuhiro; Oshimura, Mitsuo; Hara, Takahiko

2014-01-01

Human artificial chromosomes (HACs) are gene-delivery vectors suitable for introducing large DNA fragments into mammalian cells. Although a HAC theoretically incorporates multiple gene expression cassettes of unlimited DNA size, its application has been limited because the conventional gene-loading system accepts only one gene-loading vector (GLV) into a HAC. We report a novel method for the simultaneous or sequential integration of multiple GLVs into a HAC vector (designated as the SIM system) via combined usage of Cre, FLP, Bxb1, and φC31 recombinase/integrase. As a proof of principle, we first attempted simultaneous integration of three GLVs encoding EGFP, Venus, and TdTomato into a gene-loading site of a HAC in CHO cells. These cells successfully expressed all three fluorescent proteins. Furthermore, microcell-mediated transfer of HACs enabled the expression of those fluorescent proteins in recipient cells. We next demonstrated that GLVs could be introduced into a HAC one-by-one via reciprocal usage of recombinase/integrase. Lastly, we introduced a fourth GLV into a HAC after simultaneous integration of three GLVs by FLP-mediated DNA recombination. The SIM system expands the applicability of HAC vectors and is useful for various biomedical studies, including cell reprogramming.

A novel system for simultaneous or sequential integration of multiple gene-loading vectors into a defined site of a human artificial chromosome.

Directory of Open Access Journals (Sweden)

Teruhiko Suzuki

Full Text Available Human artificial chromosomes (HACs are gene-delivery vectors suitable for introducing large DNA fragments into mammalian cells. Although a HAC theoretically incorporates multiple gene expression cassettes of unlimited DNA size, its application has been limited because the conventional gene-loading system accepts only one gene-loading vector (GLV into a HAC. We report a novel method for the simultaneous or sequential integration of multiple GLVs into a HAC vector (designated as the SIM system via combined usage of Cre, FLP, Bxb1, and φC31 recombinase/integrase. As a proof of principle, we first attempted simultaneous integration of three GLVs encoding EGFP, Venus, and TdTomato into a gene-loading site of a HAC in CHO cells. These cells successfully expressed all three fluorescent proteins. Furthermore, microcell-mediated transfer of HACs enabled the expression of those fluorescent proteins in recipient cells. We next demonstrated that GLVs could be introduced into a HAC one-by-one via reciprocal usage of recombinase/integrase. Lastly, we introduced a fourth GLV into a HAC after simultaneous integration of three GLVs by FLP-mediated DNA recombination. The SIM system expands the applicability of HAC vectors and is useful for various biomedical studies, including cell reprogramming.

Fungal genome and mating system transitions facilitated by chromosomal translocations involving intercentromeric recombination.

Directory of Open Access Journals (Sweden)

Sheng Sun

2017-08-01

rearrangements mediated by intercentromeric recombination have occurred during descent of the 2 lineages from their common ancestor. Taken together, our findings support a model in which the evolution of the bipolar mating system was initiated by an ectopic recombination event mediated by similar repetitive centromeric DNA elements shared between chromosomes. This translocation brought the P/R and HD loci onto the same chromosome, and further chromosomal rearrangements then resulted in the 2 MAT loci becoming physically linked and eventually fusing to form the single contiguous MAT locus that is now extant in the pathogenic Cryptococcus species.

Genetic architecture of autosome-mediated hybrid male sterility in Drosophila.

Science.gov (United States)

Marín, I

1996-04-01

Several estimators have been developed for assessing the number of sterility factors in a chromosome based on the sizes of fertile and sterile introgressed fragments. Assuming that two factors are required for producing sterility, simulations show that one of these, twice the inverse of the relative size of the largest fertile fragment, provides good average approximations when as few as five fertile fragments are analyzed. The estimators have been used for deducing the number of factors from previous data on several pairs of species. A particular result contrasts with the authors' interpretations: instead of the high number of sterility factors suggested, only a few per autosome are estimated in both reciprocal crosses involving Drosophila buzzatii and D. koepferae. It has been possible to map these factors, between three and six per chromosome, in the autosomes 3 and 4 of these species. Out of 203 introgressions of different fragments or combinations of fragments, the outcome of at least 192 is explained by the mapped zones. These results suggest that autosome-mediated sterility in the male hybrids of these species is mediated by a few epistatic factors, similarly to X-mediated sterility in the hybrids of other Drosophila species.

Knock-in/Knock-out (KIKO) vectors for rapid integration of large DNA sequences, including whole metabolic pathways, onto the Escherichia coli chromosome at well-characterised loci.

Science.gov (United States)

Sabri, Suriana; Steen, Jennifer A; Bongers, Mareike; Nielsen, Lars K; Vickers, Claudia E

2013-06-24

Metabolic engineering projects often require integration of multiple genes in order to control the desired phenotype. However, this often requires iterative rounds of engineering because many current insertion approaches are limited by the size of the DNA that can be transferred onto the chromosome. Consequently, construction of highly engineered strains is very time-consuming. A lack of well-characterised insertion loci is also problematic. A series of knock-in/knock-out (KIKO) vectors was constructed for integration of large DNA sequences onto the E. coli chromosome at well-defined loci. The KIKO plasmids target three nonessential genes/operons as insertion sites: arsB (an arsenite transporter); lacZ (β-galactosidase); and rbsA-rbsR (a ribose metabolism operon). Two homologous 'arms' target each insertion locus; insertion is mediated by λ Red recombinase through these arms. Between the arms is a multiple cloning site for the introduction of exogenous sequences and an antibiotic resistance marker (either chloramphenicol or kanamycin) for selection of positive recombinants. The resistance marker can subsequently be removed by flippase-mediated recombination. The insertion cassette is flanked by hairpin loops to isolate it from the effects of external transcription at the integration locus. To characterize each target locus, a xylanase reporter gene (xynA) was integrated onto the chromosomes of E. coli strains W and K-12 using the KIKO vectors. Expression levels varied between loci, with the arsB locus consistently showing the highest level of expression. To demonstrate the simultaneous use of all three loci in one strain, xynA, green fluorescent protein (gfp) and a sucrose catabolic operon (cscAKB) were introduced into lacZ, arsB and rbsAR respectively, and shown to be functional. The KIKO plasmids are a useful tool for efficient integration of large DNA fragments (including multiple genes and pathways) into E. coli. Chromosomal insertion provides stable

Centromere Destiny in Dicentric Chromosomes: New Insights from the Evolution of Human Chromosome 2 Ancestral Centromeric Region.

Science.gov (United States)

Chiatante, Giorgia; Giannuzzi, Giuliana; Calabrese, Francesco Maria; Eichler, Evan E; Ventura, Mario

2017-07-01

Dicentric chromosomes are products of genomic rearrangements that place two centromeres on the same chromosome. Due to the presence of two primary constrictions, they are inherently unstable and overcome their instability by epigenetically inactivating and/or deleting one of the two centromeres, thus resulting in functionally monocentric chromosomes that segregate normally during cell division. Our understanding to date of dicentric chromosome formation, behavior and fate has been largely inferred from observational studies in plants and humans as well as artificially produced de novo dicentrics in yeast and in human cells. We investigate the most recent product of a chromosome fusion event fixed in the human lineage, human chromosome 2, whose stability was acquired by the suppression of one centromere, resulting in a unique difference in chromosome number between humans (46 chromosomes) and our most closely related ape relatives (48 chromosomes). Using molecular cytogenetics, sequencing, and comparative sequence data, we deeply characterize the relicts of the chromosome 2q ancestral centromere and its flanking regions, gaining insight into the ancestral organization that can be easily broadened to all acrocentric chromosome centromeres. Moreover, our analyses offered the opportunity to trace the evolutionary history of rDNA and satellite III sequences among great apes, thus suggesting a new hypothesis for the preferential inactivation of some human centromeres, including IIq. Our results suggest two possible centromere inactivation models to explain the evolutionarily stabilization of human chromosome 2 over the last 5-6 million years. Our results strongly favor centromere excision through a one-step process. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Chromosome Connections: Compelling Clues to Common Ancestry

Science.gov (United States)

Flammer, Larry

2013-01-01

Students compare banding patterns on hominid chromosomes and see striking evidence of their common ancestry. To test this, human chromosome no. 2 is matched with two shorter chimpanzee chromosomes, leading to the hypothesis that human chromosome 2 resulted from the fusion of the two shorter chromosomes. Students test that hypothesis by looking for…

Baculovirus-mediated gene transfer in butterfly wings in vivo: an efficient expression system with an anti-gp64 antibody.

Science.gov (United States)

Dhungel, Bidur; Ohno, Yoshikazu; Matayoshi, Rie; Otaki, Joji M

2013-03-25

Candidate genes for color pattern formation in butterfly wings have been known based on gene expression patterns since the 1990s, but their functions remain elusive due to a lack of a functional assay. Several methods of transferring and expressing a foreign gene in butterfly wings have been reported, but they have suffered from low success rates or low expression levels. Here, we developed a simple, practical method to efficiently deliver and express a foreign gene using baculovirus-mediated gene transfer in butterfly wings in vivo. A recombinant baculovirus containing a gene for green fluorescent protein (GFP) was injected into pupae of the blue pansy butterfly Junonia orithya (Nymphalidae). GFP fluorescence was detected in the pupal wings and other body parts of the injected individuals three to five days post-injection at various degrees of fluorescence. We obtained a high GFP expression rate at relatively high virus titers, but it was associated with pupal death before color pattern formation in wings. To reduce the high mortality rate caused by the baculovirus treatment, we administered an anti-gp64 antibody, which was raised against baculovirus coat protein gp64, to infected pupae after the baculovirus injection. This treatment greatly reduced the mortality rate of the infected pupae. GFP fluorescence was observed in pupal and adult wings and other body parts of the antibody-treated individuals at various degrees of fluorescence. Importantly, we obtained completely developed wings with a normal color pattern, in which fluorescent signals originated directly from scales or the basal membrane after the removal of scales. GFP fluorescence in wing tissues spatially coincided with anti-GFP antibody staining, confirming that the fluorescent signals originated from the expressed GFP molecules. Our baculovirus-mediated gene transfer system with an anti-gp64 antibody is reasonably efficient, and it can be an invaluable tool to transfer, express, and functionally

Effects of alpha-particles on survival and chromosomal aberrations in human mammary epithelial cells

Science.gov (United States)

Durante, M.; Grossi, G. F.; Gialanella, G.; Pugliese, M.; Nappo, M.; Yang, T. C.

1995-01-01

We have studied the radiation responses of a human mammary epithelial cell line, H184B5 F5-1 M/10. This cell line was derived from primary mammary cells after treatment with chemicals and heavy ions. The F5-1 M/10 cells are immortal, density-inhibited in growth, and non-tumorigenic in athymic nude mice and represent an in vitro model of the human epithelium for radiation studies. Because epithelial cells are the target of alpha-particles emitted from radon daughters, we concentrated our studies on the efficiency of alpha-particles. Confluent cultures of M/10 cells were exposed to accelerated alpha-particles [beam energy incident at the cell monolayer = 3.85 MeV, incident linear energy transfer (LET) in cell = 109 keV/microns] and, for comparison, to 80 kVp x-rays. The following endpoints were studied: (1) survival, (2) chromosome aberrations at the first postirradiation mitosis, and (3) chromosome alterations at later passages following irradiation. The survival curve was exponential for alpha-particles (D0 = 0.73 +/- 0.04 Gy), while a shoulder was observed for x-rays (alpha/beta = 2.9 Gy; D0 = 2.5 Gy, extrapolation number 1.6). The relative biological effectiveness (RBE) of high-LET alpha-particles for human epithelial cell killing was 3.3 at 37% survival. Dose-response curves for the induction of chromosome aberrations were linear for alpha-particles and linearquadratic for x-rays. The RBE for the induction of chromosome aberrations varied with the type of aberration scored and was high (about 5) for chromosome breaks and low (about 2) for chromosome exchanges.(ABSTRACT TRUNCATED AT 250 WORDS).

Fishing for radiation quality: chromosome aberrations and the role of radiation track structure

International Nuclear Information System (INIS)

Hill, M.A.

2015-01-01

The yield of chromosome aberrations is not only dependent on dose but also on radiation quality, with high linear energy transfer (LET) typically having a greater biological effectiveness per unit dose than those of low-LET radiation. Differences in radiation track structure and cell morphology can also lead to quantitative differences in the spectra of the resulting chromosomal rearrangements, especially at low doses associated with typical human exposures. The development of combinatorial fluorescent labelling techniques (such as mFISH and mBAND) has helped to reveal the complexity of rearrangements, showing increasing complexity of observed rearrangements with increasing LET but has a resolution limited to ∼10 MBp. High-LET particles have not only been shown to produce clustered sites of DNA damage but also produce multiple correlated breaks along its path resulting in DNA fragments smaller than the resolution of these techniques. Additionally, studies have shown that the vast majority of radiation-induced HPRT mutations were also not detectable using fluorescent in situ hybridisation (FISH) techniques, with correlation of breaks along the track being reflected in the complexity of mutations, with intra- and inter-chromosomal insertions, and inversions occurring at the sites of some of the deletions. Therefore, the analysis of visible chromosomal rearrangements observed using current FISH techniques is likely to represent just the tip of the iceberg, considerably underestimating the extent and complexity of radiation induced rearrangements. (author)

Linkage group-chromosome correlations in Sordaria macrospora: Chromosome identification by three dimensional reconstruction of their synaptonemal complex.

Science.gov (United States)

Zickler, D; Leblon, G; Haedens, V; Collard, A; Thuriaux, P

1984-01-01

Reconstruction of serially sectioned zygotene and pachytene nuclei has allowed, by measuring the lengths of synaptonemal complexes, an assignment of the 7 linkage (LG) groups to the 7 chromosomes in the fungus Sordaria macrospora. The 7 LG have been established using 19 mutants showing low second division segregation frequencies. Eight chromosomal rearrangements mapped on the 7 LG were used to identify the chromosomes involved. The following one to one assignment of the 7 LG to specific chromosomes was obtained: LG a: chromosome (chr) 1, LG b: chr5, LG c: chr6, LG d: chr7, LG e: chr4, LG f: chr3 and LG g: chr2 (the chromosome carrying the nucleolus organizer region).

Characterization of a chromosome-specific chimpanzee alpha satellite subset: Evolutionary relationship to subsets on human chromosomes

Energy Technology Data Exchange (ETDEWEB)

Warburton, P.E.; Gosden, J.; Lawson, D. [Western General Hospital, Edinburgh (United Kingdom)] [and others

1996-04-15

Alpha satellite DNA is a tandemly repeated DNA family found at the centromeres of all primate chromosomes examined. The fundamental repeat units of alpha satellite DNA are diverged 169- to 172-bp monomers, often found to be organized in chromosome-specific higher-order repeat units. The chromosomes of human (Homo sapiens (HSA)), chimpanzee (Pan troglodytes (PTR) and Pan paniscus), and gorilla (Gorilla gorilla) share a remarkable similarity and synteny. It is of interest to ask if alpha satellite arrays at centromeres of homologous chromosomes between these species are closely related (evolving in an orthologous manner) or if the evolutionary processes that homogenize and spread these arrays within and between chromosomes result in nonorthologous evolution of arrays. By using PCR primers specific for human chromosome 17-specific alpha satellite DNA, we have amplified, cloned, and characterized a chromosome-specific subset from the PTR chimpanzee genome. Hybridization both on Southern blots and in situ as well as sequence analysis show that this subset is most closely related, as expected, to sequences on HSA 17. However, in situ hybridization reveals that this subset is not found on the homologous chromosome in chimpanzee (PTR 19), but instead on PTR 12, which is homologous to HSA 2p. 40 refs., 3 figs.

Addition of Aegilops U and M Chromosomes Affects Protein and Dietary Fiber Content of Wholemeal Wheat Flour

Directory of Open Access Journals (Sweden)

Marianna Rakszegi

2017-09-01

Full Text Available Cereal grain fiber is an important health-promoting component in the human diet. One option to improve dietary fiber content and composition in wheat is to introduce genes from its wild relatives Aegilops biuncialis and Aegilops geniculata. This study showed that the addition of chromosomes 2Ug, 4Ug, 5Ug, 7Ug, 2Mg, 5Mg, and 7Mg of Ae. geniculata and 3Ub, 2Mb, 3Mb, and 7Mb of Ae. biuncialis into bread wheat increased the seed protein content. Chromosomes 1Ug and 1Mg increased the proportion of polymeric glutenin proteins, while the addition of chromosomes 1Ub and 6Ub led to its decrease. Both Aegilops species had higher proportions of β-glucan compared to arabinoxylan (AX than wheat lines, and elevated β-glucan content was also observed in wheat chromosome addition lines 5U, 7U, and 7M. The AX content in wheat was increased by the addition of chromosomes 5Ug, 7Ug, and 1Ub while water-soluble AX was increased by the addition of chromosomes 5U, 5M, and 7M, and to a lesser extent by chromosomes 3, 4, 6Ug, and 2Mb. Chromosomes 5Ug and 7Mb also affected the structure of wheat AX, as shown by the pattern of oligosaccharides released by digestion with endoxylanase. These results will help to map genomic regions responsible for edible fiber content in Aegilops and will contribute to the efficient transfer of wild alleles in introgression breeding programs to obtain wheat varieties with improved health benefits.Key Message: Addition of Aegilops U- and M-genome chromosomes 5 and 7 improves seed protein and fiber content and composition in wheat.

Comparative physical mapping between wheat chromosome arm 2BL and rice chromosome 4.

Science.gov (United States)

Lee, Tong Geon; Lee, Yong Jin; Kim, Dae Yeon; Seo, Yong Weon

2010-12-01

Physical maps of chromosomes provide a framework for organizing and integrating diverse genetic information. DNA microarrays are a valuable technique for physical mapping and can also be used to facilitate the discovery of single feature polymorphisms (SFPs). Wheat chromosome arm 2BL was physically mapped using a Wheat Genome Array onto near-isogenic lines (NILs) with the aid of wheat-rice synteny and mapped wheat EST information. Using high variance probe set (HVP) analysis, 314 HVPs constituting genes present on 2BL were identified. The 314 HVPs were grouped into 3 categories: HVPs that match only rice chromosome 4 (298 HVPs), those that match only wheat ESTs mapped on 2BL (1), and those that match both rice chromosome 4 and wheat ESTs mapped on 2BL (15). All HVPs were converted into gene sets, which represented either unique rice gene models or mapped wheat ESTs that matched identified HVPs. Comparative physical maps were constructed for 16 wheat gene sets and 271 rice gene sets. Of the 271 rice gene sets, 257 were mapped to the 18-35 Mb regions on rice chromosome 4. Based on HVP analysis and sequence similarity between the gene models in the rice chromosomes and mapped wheat ESTs, the outermost rice gene model that limits the translocation breakpoint to orthologous regions was identified.

A Rare De novo Complex Chromosomal Rearrangement (CCR) Involving Four Chromosomes in An Oligo-asthenosperm Infertile Man.

Science.gov (United States)

Asia, Saba; Vaziri Nasab, Hamed; Sabbaghian, Marjan; Kalantari, Hamid; Zari Moradi, Shabnam; Gourabi, Hamid; Mohseni Meybodi, Anahita

2014-01-01

Complex chromosomal rearrangements (CCRs) are rare events involving more than two chromosomes and over two breakpoints. They are usually associated with infertility or sub fertility in male carriers. Here we report a novel case of a CCR in a 30-year-old oligoasthenosperm man with a history of varicocelectomy, normal testes size and normal endocrinology profile referred for chromosome analysis to the Genetics unit of Royan Reproductive Biomedicine Research Center. Chromosomal analysis was performed using peripheral blood lymphocyte cultures and analyzed by GTG banding. Additional tests such as C-banding and multicolor fluorescence in situ hybridization (FISH) procedure for each of the involved chromosomes were performed to determine the patterns of the segregations. Y chromosome microdeletions in the azoospermia factor (AZF) region were analyzed with multiplex polymerase chain reaction. To identify the history and origin of this CCR, all the family members were analyzed. No micro deletion in Y chromosome was detected. The same de novo reciprocal exchange was also found in his monozygous twin brother. The other siblings and parents were normal. CCRs are associated with male infertility as a result of spermatogenic disruption due to complex meiotic configurations and the production of chromosomally abnormal sperms. These chromosomal rearrangements might have an influence on decreasing the number of sperms.

Chromosome Banding in Amphibia. XXXVI. Multimorphic Sex Chromosomes and an Enigmatic Sex Determination in Eleutherodactylus johnstonei (Anura, Eleutherodactylidae).

Science.gov (United States)

Schmid, Michael; Steinlein, Claus

2018-01-01

A detailed cytogenetic study on the leaf litter frog Eleutherodactylus johnstonei from 14 different Caribbean islands and the mainlands of Venezuela and Guyana revealed the existence of multimorphic XY♂/XX♀ sex chromosomes 14. Their male sex determination and development depends either on the presence of 2 telocentric chromosomes 14 (XtYt), or on 1 submetacentric chromosome 14 (Xsm) plus 1 telocentric chromosome 14 (Yt), or on the presence of 2 submetacentric chromosomes 14 (XsmYsm). The female sex determination and development requires either the presence of 2 telocentric chromosomes 14 (XtXt) or 2 submetacentric chromosomes 14 (XsmXsm). In all individuals analyzed, the sex chromosomes 14 carry a prominent nucleolus organizer region in their long arms. An explanation is given for the origin of the (XtYt)♂, (XsmYt)♂, (XsmYsm)♂, (XtXt)♀, and (XsmXsm)♀ in the different populations of E. johnstonei. Furthermore, the present study gives detailed data on the chromosome banding patterns, in situ hybridization experiments, and the genome size of E. johnstonei. © 2018 S. Karger AG, Basel.

Family-based multi-SNP X chromosome analysis using parental information

Directory of Open Access Journals (Sweden)

Alison S. Wise

2016-02-01

Full Text Available We propose a method for association analysis of haplotypes on the X chromosome that offers both improved power and robustness to population stratification in studies of affected offspring and their parents if all three have been genotyped. The method makes use of assumed parental haplotype exchangeability, a weaker assumption than Hardy-Weinberg equilibrium. Parental haplotype exchangeability requires that in the source population, of the three X chromosome haplotypes carried by the two parents, each is equally likely to be carried by the father. We propose a pseudo-sibling approach that exploits that exchangeability assumption. Our method extends the single-SNP PIX-LRT method to multiple SNPs in a high linkage block. We describe methods for testing the parental haplotype exchangeability assumption and also for determining how apparent violations can be distinguished from true fetal effects or maternally-mediated effects. We show results of simulations that demonstrate nominal type I error rate and good power. The methods are then applied to dbGaP data on the birth defect oral cleft, using both Asian and Caucasian families with cleft.

Energy transfer in plasmonic systems

International Nuclear Information System (INIS)

Pustovit, Vitaliy N; Urbas, Augustine M; Shahbazyan, Tigran V

2014-01-01

We present our results on energy transfer between donor and acceptor molecules or quantum dots near a plasmonic nanoparticle. In such systems, the Förster resonance energy transfer is strongly modified due to plasmon-mediated coupling between donors and acceptors. The transfer efficiency is determined by a competition between transfer, radiation and dissipation that depends sensitively on system parameters. When donor and accepror spectral bands overlap with dipole surface plasmon resonance, the dominant transfer mechanism is through plasmon-enhanced radiative coupling. When transfer takes place from an ensemble of donors to an acceptor, a cooperative amplification of energy transfer takes place in a wide range of system parameters. (paper)

Introduction of a normal human chromosome 8 corrects abnormal phenotypes of Werner syndrome cells immortalized by expressing an hTERT gene

International Nuclear Information System (INIS)

Ariyoshi, Kentaro; Kodama, Seiji; Suzuki, Keiji; Goto, Makoto; Oshimura, Mitsuo; Ishizaki, Kanji; Watanabe, Masami

2009-01-01

Werner syndrome (WS) is an autosomal recessive disease characterized by premature aging and caused by mutations of the WRN gene mapped at 8p12. To examine functional complementation of WS phenotypes, we introduced a normal human chromosome 8 into a strain of WS fibroblasts (WS3RGB) immortalized by expressing a human telomerase reverse transcriptase subunit (hTERT) gene. Here, we demonstrate that the abnormal WS phenotypes including cellular sensitivities to 4-nitroquinoline-1-oxide (4NQO) and hydroxy urea (HU), and chromosomal radiosensitivity at G 2 phase are corrected by expression of the WRN gene mediated by introducing a chromosome 8. This indicates that those multiple abnormal WS phenotypes are derived from a primary, but not secondary, defect in the WRN gene. (author)

Frequencies of chromosome aberration on radiation workers

International Nuclear Information System (INIS)

Yanti Lusiyanti; Zubaidah Alatas

2016-01-01

Radiation exposure of the body can cause damage to the genetic material in cells (cytogenetic) in the form of changes in the structure or chromosomal aberrations in peripheral blood lymphocytes. Chromosomal aberrations can be unstable as dicentric and ring chromosomes, and is stable as translocation. Dicentric chromosome is the gold standard biomarker due to radiation exposure, and chromosome translocation is a biomarker for retrospective biodosimetry. The aim of this studi is to conduct examination of chromosomal aberrations in the radiation worker to determine the potential damage of cell that may arise due to occupational radiation exposure. The examination have been carried out on blood samples from 55 radiation workers in the range of 5-30 year of service. Chromosome aberration frequency measurement starts with blood sampling, culturing, harvesting, slide preparations, and lymphocyte chromosome staining with Giemsa and painting with Fluorescence In Situ Hybridization (FISH) technique. The results showed that chromosomal translocations are not found in blood samples radiation workers and dicentric chromosomes found only on 2 blood samples of radiation workers with a frequency of 0.001/cell. The frequency of chromosomal aberrations in the blood cells such workers within normal limits and this means that the workers have been implemented a radiation safety aspects very well. (author)

Agrobacterium-mediated transformation: state of the art and future prospect

Institute of Scientific and Technical Information of China (English)

无

2000-01-01

Great progress has been made in recent years in studies on the mechanism of Agrobacterium-mediated transformation and its application. Many details of the key molecular events within the bacterial cells involved in T-DNA transfer have been elucidated, and it is notable that some plant factors which were elusive before are purified and characterized. Vast kinds of species, which were either recalcitrant to or not included in the host range of Agrobacterium, can now be transformed by this bacterium, and they include the very important cereal species, gymnosperms, yeast and many filamentous fungi. The simple in vivo transformation of tissue in intact plants and the "agrolistic" methods to transform recalcitrant plants are the two novel technical achievements. Combined with other powerful techniques such as bacterial artificial chromosome, very large DNA fragment can be transformed into the plant genome by Agrobacterium. Further studies will elucidate more plant-encoded factors involved in T-DNA transformation and there is a need to develop more powerful Agrobacterium-based transformation systems to meet different needs in basic research and crop improvement practice.

Telomere dysfunction and chromosome structure modulate the contribution of individual chromosomes in abnormal nuclear morphologies

Energy Technology Data Exchange (ETDEWEB)

Pampalona, J.; Soler, D.; Genesca, A. [Department of Cell Biology, Physiology and Immunology, Universitat Autonoma de Barcelona, Bellaterra E-08193 (Spain); Tusell, L., E-mail: laura.tusell@uab.es [Department of Cell Biology, Physiology and Immunology, Universitat Autonoma de Barcelona, Bellaterra E-08193 (Spain)

2010-01-05

The cytokinesis-block micronucleus assay has emerged as a biomarker of chromosome damage relevant to cancer. Although it was initially developed to measure micronuclei, it is also useful for measuring nucleoplasmic bridges and nuclear buds. Abnormal nuclear morphologies are frequently observed in malignant tissues and short-term tumour cell cultures. Changes in chromosome structure and number resulting from chromosome instability are important factors in oncogenesis. Telomeres have become key players in the initiation of chromosome instability related to carcinogenesis by means of breakage-fusion-bridge cycles. To better understand the connection between telomere dysfunction and the appearance of abnormal nuclear morphologies, we have characterised the presence of micronuclei, nucleoplasmic bridges and nuclear buds in human mammary primary epithelial cells. These cells can proliferate beyond the Hayflick limit by spontaneously losing expression of the p16{sup INK4a} protein. Progressive telomere shortening leads to the loss of the capping function, and the appearance of end-to-end chromosome fusions that can enter into breakage-fusion-bridge cycles generating massive chromosomal instability. In human mammary epithelial cells, different types of abnormal nuclear morphologies were observed, however only nucleoplasmatic bridges and buds increased significantly with population doublings. Fluorescent in situ hybridisation using centromeric and painting specific probes for chromosomes with eroded telomeres has revealed that these chromosomes are preferentially included in the different types of abnormal nuclear morphologies observed, thus reflecting their common origin. Accordingly, real-time imaging of cell divisions enabled us to determine that anaphase bridge resolution was mainly through chromatin breakage and the formation of symmetric buds in daughter nuclei. Few micronuclei emerged in this cell system thus validating the scoring of nucleoplasmic bridges and

Telomere dysfunction and chromosome structure modulate the contribution of individual chromosomes in abnormal nuclear morphologies

International Nuclear Information System (INIS)

Pampalona, J.; Soler, D.; Genesca, A.; Tusell, L.

2010-01-01

The cytokinesis-block micronucleus assay has emerged as a biomarker of chromosome damage relevant to cancer. Although it was initially developed to measure micronuclei, it is also useful for measuring nucleoplasmic bridges and nuclear buds. Abnormal nuclear morphologies are frequently observed in malignant tissues and short-term tumour cell cultures. Changes in chromosome structure and number resulting from chromosome instability are important factors in oncogenesis. Telomeres have become key players in the initiation of chromosome instability related to carcinogenesis by means of breakage-fusion-bridge cycles. To better understand the connection between telomere dysfunction and the appearance of abnormal nuclear morphologies, we have characterised the presence of micronuclei, nucleoplasmic bridges and nuclear buds in human mammary primary epithelial cells. These cells can proliferate beyond the Hayflick limit by spontaneously losing expression of the p16 INK4a protein. Progressive telomere shortening leads to the loss of the capping function, and the appearance of end-to-end chromosome fusions that can enter into breakage-fusion-bridge cycles generating massive chromosomal instability. In human mammary epithelial cells, different types of abnormal nuclear morphologies were observed, however only nucleoplasmatic bridges and buds increased significantly with population doublings. Fluorescent in situ hybridisation using centromeric and painting specific probes for chromosomes with eroded telomeres has revealed that these chromosomes are preferentially included in the different types of abnormal nuclear morphologies observed, thus reflecting their common origin. Accordingly, real-time imaging of cell divisions enabled us to determine that anaphase bridge resolution was mainly through chromatin breakage and the formation of symmetric buds in daughter nuclei. Few micronuclei emerged in this cell system thus validating the scoring of nucleoplasmic bridges and nuclear

Telomere dysfunction and chromosome structure modulate the contribution of individual chromosomes in abnormal nuclear morphologies.

Science.gov (United States)

Pampalona, J; Soler, D; Genescà, A; Tusell, L

2010-01-05

The cytokinesis-block micronucleus assay has emerged as a biomarker of chromosome damage relevant to cancer. Although it was initially developed to measure micronuclei, it is also useful for measuring nucleoplasmic bridges and nuclear buds. Abnormal nuclear morphologies are frequently observed in malignant tissues and short-term tumour cell cultures. Changes in chromosome structure and number resulting from chromosome instability are important factors in oncogenesis. Telomeres have become key players in the initiation of chromosome instability related to carcinogenesis by means of breakage-fusion-bridge cycles. To better understand the connection between telomere dysfunction and the appearance of abnormal nuclear morphologies, we have characterised the presence of micronuclei, nucleoplasmic bridges and nuclear buds in human mammary primary epithelial cells. These cells can proliferate beyond the Hayflick limit by spontaneously losing expression of the p16(INK4a) protein. Progressive telomere shortening leads to the loss of the capping function, and the appearance of end-to-end chromosome fusions that can enter into breakage-fusion-bridge cycles generating massive chromosomal instability. In human mammary epithelial cells, different types of abnormal nuclear morphologies were observed, however only nucleoplasmatic bridges and buds increased significantly with population doublings. Fluorescent in situ hybridisation using centromeric and painting specific probes for chromosomes with eroded telomeres has revealed that these chromosomes are preferentially included in the different types of abnormal nuclear morphologies observed, thus reflecting their common origin. Accordingly, real-time imaging of cell divisions enabled us to determine that anaphase bridge resolution was mainly through chromatin breakage and the formation of symmetric buds in daughter nuclei. Few micronuclei emerged in this cell system thus validating the scoring of nucleoplasmic bridges and nuclear

Label Free Chromosome Translocation Detection with Silicon nanowires

DEFF Research Database (Denmark)

Kwasny, Dorota; Andersen, Karsten Brandt; Frøhling, Kasper Bayer

HROMOSOME translocation, which is a rearrangement of arms between two chromosomes, is a major group of chromosome abnormalities leading to cancer. As a result, two derivative chromosomes with sequences coming from both chromosomes are formed. The current translocation detection method is a Fluore......HROMOSOME translocation, which is a rearrangement of arms between two chromosomes, is a major group of chromosome abnormalities leading to cancer. As a result, two derivative chromosomes with sequences coming from both chromosomes are formed. The current translocation detection method...

Female chromosome X mosaicism is age-related and preferentially affects the inactivated X chromosome

Science.gov (United States)

Machiela, Mitchell J.; Zhou, Weiyin; Karlins, Eric; Sampson, Joshua N.; Freedman, Neal D.; Yang, Qi; Hicks, Belynda; Dagnall, Casey; Hautman, Christopher; Jacobs, Kevin B.; Abnet, Christian C.; Aldrich, Melinda C.; Amos, Christopher; Amundadottir, Laufey T.; Arslan, Alan A.; Beane-Freeman, Laura E.; Berndt, Sonja I.; Black, Amanda; Blot, William J.; Bock, Cathryn H.; Bracci, Paige M.; Brinton, Louise A.; Bueno-de-Mesquita, H Bas; Burdett, Laurie; Buring, Julie E.; Butler, Mary A.; Canzian, Federico; Carreón, Tania; Chaffee, Kari G.; Chang, I-Shou; Chatterjee, Nilanjan; Chen, Chu; Chen, Constance; Chen, Kexin; Chung, Charles C.; Cook, Linda S.; Crous Bou, Marta; Cullen, Michael; Davis, Faith G.; De Vivo, Immaculata; Ding, Ti; Doherty, Jennifer; Duell, Eric J.; Epstein, Caroline G.; Fan, Jin-Hu; Figueroa, Jonine D.; Fraumeni, Joseph F.; Friedenreich, Christine M.; Fuchs, Charles S.; Gallinger, Steven; Gao, Yu-Tang; Gapstur, Susan M.; Garcia-Closas, Montserrat; Gaudet, Mia M.; Gaziano, J. Michael; Giles, Graham G.; Gillanders, Elizabeth M.; Giovannucci, Edward L.; Goldin, Lynn; Goldstein, Alisa M.; Haiman, Christopher A.; Hallmans, Goran; Hankinson, Susan E.; Harris, Curtis C.; Henriksson, Roger; Holly, Elizabeth A.; Hong, Yun-Chul; Hoover, Robert N.; Hsiung, Chao A.; Hu, Nan; Hu, Wei; Hunter, David J.; Hutchinson, Amy; Jenab, Mazda; Johansen, Christoffer; Khaw, Kay-Tee; Kim, Hee Nam; Kim, Yeul Hong; Kim, Young Tae; Klein, Alison P.; Klein, Robert; Koh, Woon-Puay; Kolonel, Laurence N.; Kooperberg, Charles; Kraft, Peter; Krogh, Vittorio; Kurtz, Robert C.; LaCroix, Andrea; Lan, Qing; Landi, Maria Teresa; Marchand, Loic Le; Li, Donghui; Liang, Xiaolin; Liao, Linda M.; Lin, Dongxin; Liu, Jianjun; Lissowska, Jolanta; Lu, Lingeng; Magliocco, Anthony M.; Malats, Nuria; Matsuo, Keitaro; McNeill, Lorna H.; McWilliams, Robert R.; Melin, Beatrice S.; Mirabello, Lisa; Moore, Lee; Olson, Sara H.; Orlow, Irene; Park, Jae Yong; Patiño-Garcia, Ana; Peplonska, Beata; Peters, Ulrike; Petersen, Gloria M.; Pooler, Loreall; Prescott, Jennifer; Prokunina-Olsson, Ludmila; Purdue, Mark P.; Qiao, You-Lin; Rajaraman, Preetha; Real, Francisco X.; Riboli, Elio; Risch, Harvey A.; Rodriguez-Santiago, Benjamin; Ruder, Avima M.; Savage, Sharon A.; Schumacher, Fredrick; Schwartz, Ann G.; Schwartz, Kendra L.; Seow, Adeline; Wendy Setiawan, Veronica; Severi, Gianluca; Shen, Hongbing; Sheng, Xin; Shin, Min-Ho; Shu, Xiao-Ou; Silverman, Debra T.; Spitz, Margaret R.; Stevens, Victoria L.; Stolzenberg-Solomon, Rachael; Stram, Daniel; Tang, Ze-Zhong; Taylor, Philip R.; Teras, Lauren R.; Tobias, Geoffrey S.; Van Den Berg, David; Visvanathan, Kala; Wacholder, Sholom; Wang, Jiu-Cun; Wang, Zhaoming; Wentzensen, Nicolas; Wheeler, William; White, Emily; Wiencke, John K.; Wolpin, Brian M.; Wong, Maria Pik; Wu, Chen; Wu, Tangchun; Wu, Xifeng; Wu, Yi-Long; Wunder, Jay S.; Xia, Lucy; Yang, Hannah P.; Yang, Pan-Chyr; Yu, Kai; Zanetti, Krista A.; Zeleniuch-Jacquotte, Anne; Zheng, Wei; Zhou, Baosen; Ziegler, Regina G.; Perez-Jurado, Luis A.; Caporaso, Neil E.; Rothman, Nathaniel; Tucker, Margaret; Dean, Michael C.; Yeager, Meredith; Chanock, Stephen J.

2016-01-01

To investigate large structural clonal mosaicism of chromosome X, we analysed the SNP microarray intensity data of 38,303 women from cancer genome-wide association studies (20,878 cases and 17,425 controls) and detected 124 mosaic X events >2 Mb in 97 (0.25%) women. Here we show rates for X-chromosome mosaicism are four times higher than mean autosomal rates; X mosaic events more often include the entire chromosome and participants with X events more likely harbour autosomal mosaic events. X mosaicism frequency increases with age (0.11% in 50-year olds; 0.45% in 75-year olds), as reported for Y and autosomes. Methylation array analyses of 33 women with X mosaicism indicate events preferentially involve the inactive X chromosome. Our results provide further evidence that the sex chromosomes undergo mosaic events more frequently than autosomes, which could have implications for understanding the underlying mechanisms of mosaic events and their possible contribution to risk for chronic diseases. PMID:27291797

Stabilization of chromosomes by DNA intercalators for flow karyotyping and identification by banding of isolated chromosomes

NARCIS (Netherlands)

Aten, J. A.; Buys, C. H.; van der Veen, A. Y.; Mesa, J. R.; Yu, L. C.; Gray, J. W.; Osinga, J.; Stap, J.

1987-01-01

A number of structurally unrelated DNA intercalators have been studied as stabilizers of mitotic chromosomes during isolation from rodent and human metaphase cells. Seven out of the nine intercalators tested were found to be useful as chromosome stabilizing agents. Chromosome suspensions prepared in

Developing de novo human artificial chromosomes in embryonic stem cells using HSV-1 amplicon technology.

Science.gov (United States)

Moralli, Daniela; Monaco, Zoia L

2015-02-01

De novo artificial chromosomes expressing genes have been generated in human embryonic stem cells (hESc) and are maintained following differentiation into other cell types. Human artificial chromosomes (HAC) are small, functional, extrachromosomal elements, which behave as normal chromosomes in human cells. De novo HAC are generated following delivery of alpha satellite DNA into target cells. HAC are characterized by high levels of mitotic stability and are used as models to study centromere formation and chromosome organisation. They are successful and effective as gene expression vectors since they remain autonomous and can accommodate larger genes and regulatory regions for long-term expression studies in cells unlike other viral gene delivery vectors currently used. Transferring the essential DNA sequences for HAC formation intact across the cell membrane has been challenging for a number of years. A highly efficient delivery system based on HSV-1 amplicons has been used to target DNA directly to the ES cell nucleus and HAC stably generated in human embryonic stem cells (hESc) at high frequency. HAC were detected using an improved protocol for hESc chromosome harvesting, which consistently produced high-quality metaphase spreads that could routinely detect HAC in hESc. In tumour cells, the input DNA often integrated in the host chromosomes, but in the host ES genome, it remained intact. The hESc containing the HAC formed embryoid bodies, generated teratoma in mice, and differentiated into neuronal cells where the HAC were maintained. The HAC structure and chromatin composition was similar to the endogenous hESc chromosomes. This review will discuss the technological advances in HAC vector delivery using HSV-1 amplicons and the improvements in the identification of de novo HAC in hESc.

Kidney-specific Sonoporation-mediated Gene Transfer.

Science.gov (United States)

Ishida, Ryo; Kami, Daisuke; Kusaba, Tetsuro; Kirita, Yuhei; Kishida, Tsunao; Mazda, Osam; Adachi, Takaomi; Gojo, Satoshi

2016-02-01

Sonoporation can deliver agents to target local organs by systemic administration, while decreasing the associated risk of adverse effects. Sonoporation has been used for a variety of materials and in a variety of organs. Herein, we demonstrated that local sonoporation to the kidney can offer highly efficient transfer of oligonucleotides, which were systemically administrated to the tubular epithelium with high specificity. Ultrasonic wave irradiation to the kidney collapsed the microbubbles and transiently affected the glomerular filtration barrier and increased glomerular permeability. Oligonucleotides were passed through the barrier all at once and were absorbed throughout the tubular epithelium. Tumor necrosis factor alpha (TNFα), which plays a central role in renal ischemia-reperfusion injury, was targeted using small interfering RNA (siRNA) with renal sonoporation in a murine model. The reduction of TNFα expression after single gene transfer significantly inhibited the expression of kidney injury markers, suggesting that systemic administration of siRNA under temporary and local sonoporation could be applicable in the clinical setting of ischemic acute kidney injury.

Transcription-factor-mediated DNA looping probed by high-resolution, single-molecule imaging in live E. coli cells.

Directory of Open Access Journals (Sweden)

Zach Hensel

Full Text Available DNA looping mediated by transcription factors plays critical roles in prokaryotic gene regulation. The "genetic switch" of bacteriophage λ determines whether a prophage stays incorporated in the E. coli chromosome or enters the lytic cycle of phage propagation and cell lysis. Past studies have shown that long-range DNA interactions between the operator sequences O(R and O(L (separated by 2.3 kb, mediated by the λ repressor CI (accession number P03034, play key roles in regulating the λ switch. In vitro, it was demonstrated that DNA segments harboring the operator sequences formed loops in the presence of CI, but CI-mediated DNA looping has not been directly visualized in vivo, hindering a deep understanding of the corresponding dynamics in realistic cellular environments. We report a high-resolution, single-molecule imaging method to probe CI-mediated DNA looping in live E. coli cells. We labeled two DNA loci with differently colored fluorescent fusion proteins and tracked their separations in real time with ∼40 nm accuracy, enabling the first direct analysis of transcription-factor-mediated DNA looping in live cells. Combining looping measurements with measurements of CI expression levels in different operator mutants, we show quantitatively that DNA looping activates transcription and enhances repression. Further, we estimated the upper bound of the rate of conformational change from the unlooped to the looped state, and discuss how chromosome compaction may impact looping kinetics. Our results provide insights into transcription-factor-mediated DNA looping in a variety of operator and CI mutant backgrounds in vivo, and our methodology can be applied to a broad range of questions regarding chromosome conformations in prokaryotes and higher organisms.

FISH detection of chromosomal aberrations: the state-of-the-art in Argentina

International Nuclear Information System (INIS)

Nasazzi, Nora B.; Maranon, David; Muhlmann, Maria del C.

2004-01-01

The combined application of cytogenetics and molecular biology allows the identification, through fluorescence in situ hybridization technique (FISH), of numerical and structural chromosomal genetic alterations. The application fields are: the basic genetic research in man and other species, medical diagnosis and prognosis related to constitutive and somatic cell genetics, and biological dosimetry. Up to now, in our country as in the rest of Latin America, FISH is performed using commercial DNA probes. In a joint effort of CONICET, CNEA and ARN, Argentina has concluded the project to develop at pilot scale the synthesis of fluorescent probes by Chromosome Microdissection (SMF), suitable for any species and lowering the costs to about one sixth of the equivalent commercial probes. In this work, we present the general protocol, the cost analysis comparing with commercial probes and the minimum requirements for technology transfer and implementation of this technique in Latin American countries. (author)

Chromosomal mutations and chromosome loss measured in a new human-hamster hybrid cell line, ALC: studies with colcemid, ultraviolet irradiation, and 137Cs gamma-rays

Science.gov (United States)

Kraemer, S. M.; Waldren, C. A.; Chatterjee, A. (Principal Investigator)

1997-01-01

Small mutations, megabase deletions, and aneuploidy are involved in carcinogenesis and genetic defects, so it is important to be able to quantify these mutations and understand mechanisms of their creation. We have previously quantified a spectrum of mutations, including megabase deletions, in human chromosome 11, the sole human chromosome in a hamster-human hybrid cell line AL. S1- mutants have lost expression of a human cell surface antigen, S1, which is encoded by the M1C1 gene at 11p13 so that mutants can be detected via a complement-mediated cytotoxicity assay in which S1+ cells are killed and S1- cells survive. But loss of genes located on the tip of the short arm of 11 (11p15.5) is lethal to the AL hybrid, so that mutants that have lost the entire chromosome 11 die and escape detection. To circumvent this, we fused AL with Chinese hamster ovary (CHO) cells to produce a new hybrid, ALC, in which the requirement for maintaining 11p15.5 is relieved, allowing us to detect mutations events involving loss of 11p15.5. We evaluated the usefulness of this hybrid by conducting mutagenesis studies with colcemid, 137Cs gamma-radiation and UV 254 nm light. Colcemid induced 1000 more S1- mutants per unit dose in ALC than in AL; the increase for UV 254 nm light was only two-fold; and the increase for 137Cs gamma-rays was 12-fold. The increase in S1- mutant fraction in ALC cells treated with colcemid and 137Cs gamma-rays were largely due to chromosome loss and 11p deletions often containing a breakpoint within the centromeric region.

Plasmid-Mediated Antimicrobial Resistance in Staphylococci and Other Firmicutes.

Science.gov (United States)

Schwarz, Stefan; Shen, Jianzhong; Wendlandt, Sarah; Fessler, Andrea T; Wang, Yang; Kadlec, Kristina; Wu, Cong-Ming

2014-12-01

In staphylococci and other Firmicutes, resistance to numerous classes of antimicrobial agents, which are commonly used in human and veterinary medicine, is mediated by genes that are associated with mobile genetic elements. The gene products of some of these antimicrobial resistance genes confer resistance to only specific members of a certain class of antimicrobial agents, whereas others confer resistance to the entire class or even to members of different classes of antimicrobial agents. The resistance mechanisms specified by the resistance genes fall into any of three major categories: active efflux, enzymatic inactivation, and modification/replacement/protection of the target sites of the antimicrobial agents. Among the mobile genetic elements that carry such resistance genes, plasmids play an important role as carriers of primarily plasmid-borne resistance genes, but also as vectors for nonconjugative and conjugative transposons that harbor resistance genes. Plasmids can be exchanged by horizontal gene transfer between members of the same species but also between bacteria belonging to different species and genera. Plasmids are highly flexible elements, and various mechanisms exist by which plasmids can recombine, form cointegrates, or become integrated in part or in toto into the chromosomal DNA or into other plasmids. As such, plasmids play a key role in the dissemination of antimicrobial resistance genes within the gene pool to which staphylococci and other Firmicutes have access. This chapter is intended to provide an overview of the current knowledge of plasmid-mediated antimicrobial resistance in staphylococci and other Firmicutes.

Delimiting the origin of a B chromosome by FISH mapping, chromosome painting and DNA sequence analysis in Astyanax paranae (Teleostei, Characiformes.

Directory of Open Access Journals (Sweden)

Duílio M Z de A Silva

Full Text Available Supernumerary (B chromosomes have been shown to contain a wide variety of repetitive sequences. For this reason, fluorescent in situ hybridisation (FISH is a useful tool for ascertaining the origin of these genomic elements, especially when combined with painting from microdissected B chromosomes. In order to investigate the origin of B chromosomes in the fish species Astyanax paranae, these two approaches were used along with PCR amplification of specific DNA sequences obtained from the B chromosomes and its comparison with those residing in the A chromosomes. Remarkably, chromosome painting with the one-arm metacentric B chromosome probe showed hybridization signals on entire B chromosome, while FISH mapping revealed the presence of H1 histone and 18S rDNA genes symmetrically placed in both arms of the B chromosome. These results support the hypothesis that the B chromosome of A. paranae is an isochromosome. Additionally, the chromosome pairs Nos. 2 or 23 are considered the possible B chromosome ancestors since both contain syntenic H1 and 18S rRNA sequences. The analysis of DNA sequence fragments of the histone and rRNA genes obtained from the microdissected B chromosomes showed high similarity with those obtained from 0B individuals, which supports the intraspecific origin of B chromosomes in A. paranae. Finally, the population hereby analysed showed a female-biased B chromosome presence suggesting that B chromosomes in this species could influence sex determinism.

Retrospective dosimetry using chromosome painting

International Nuclear Information System (INIS)

Nasazzi, N.B.; Giorgio, M.D.; Taja, M.R.

2000-01-01

Chromosome aberration frequency measured in peripheral lymphocytes of persons exposed to ionizing radiation has been used since 1960s for dose assessment. Suspected overexposure is usually evaluated by the frequency of dicentrics and centric rings using an appropriate in vitro calibration curve. However, these chromosome aberrations are unstable with time after exposure and dose reconstruction may encounter uncertainties when the time between the exposure and the analysis is considerable or even unknown. It appears that translocations persist with time after exposure and may be used as an indication of acute past overexposures. Moreover, they appear to accumulate the cytogenetical information, which correlates with the dose received under fractionated, chronic or even occupational exposure conditions. Translocations may be detected using G-banding, which allows to score the total amount of radiation induced translocations but it is a time consuming method, or by Chromosome Painting, a method base on the Fluorescence in situ Hybridization (FISH) technique, painting only some chromosome pairs with specific whole chromosome probes and then extrapolating the observed translocation frequencies to the full genome. The latter method allows a faster aberration scoring than G-banding and appears to be the most promissory tool for biodosimetry, particularly when it is necessary to assess low doses and consequently to score a large number of metaphases, e.g. radiation workers exposed within dose limits. As with the unstable chromosome aberration, it is necessary an in vitro calibration curve based on the frequency of stable chromosome aberrations to assess doses. Our laboratory performed calibration curves for Co 60 γ-rays based on the frequencies of unstable (dicentrics and centric rings detected by conventional Giemsa staining) and stable chromosome aberrations (translocations and inversions, detected by G-banding). In order to minimize the interlaboratory variability, we

XY sex chromosome complement, compared with XX, in the CNS confers greater neurodegeneration during experimental autoimmune encephalomyelitis.

Science.gov (United States)

Du, Sienmi; Itoh, Noriko; Askarinam, Sahar; Hill, Haley; Arnold, Arthur P; Voskuhl, Rhonda R

2014-02-18

Women are more susceptible to multiple sclerosis (MS) and have more robust immune responses than men. However, men with MS tend to demonstrate a more progressive disease course than women, suggesting a disconnect between the severity of an immune attack and the CNS response to a given immune attack. We have previously shown in an MS model, experimental autoimmune encephalomyelitis, that autoantigen-sensitized XX lymph node cells, compared with XY, are more encephalitogenic. These studies demonstrated an effect of sex chromosomes in the induction of immune responses, but did not address a potential role of sex chromosomes in the CNS response to immune-mediated injury. Here, we examined this possibility using XX versus XY bone marrow chimeras reconstituted with a common immune system of one sex chromosomal type. We found that experimental autoimmune encephalomyelitis mice with an XY sex chromosome complement in the CNS, compared with XX, demonstrated greater clinical disease severity with more neuropathology in the spinal cord, cerebellum, and cerebral cortex. A candidate gene on the X chromosome, toll-like receptor 7, was then examined. Toll-like receptor 7 expression in cortical neurons was higher in mice with XY compared with mice with XX CNS, consistent with the known neurodegenerative role for toll-like receptor 7 in neurons. These results suggest that sex chromosome effects on neurodegeneration in the CNS run counter to effects on immune responses, and may bear relevance to the clinical enigma of greater MS susceptibility in women but faster disability progression in men. This is a demonstration of a direct effect of sex chromosome complement on neurodegeneration in a neurological disease.

Isolation of probes specific to human chromosomal region 6p21 from immunoselected irradiation-fusion gene transfer hybrids

International Nuclear Information System (INIS)

Ragoussis, J.; Jones, T.A.; Sheer, D.; Shrimpton, A.E.; Goodfellow, P.N.; Trowsdale, J.; Ziegler, A.

1991-01-01

A hybrid cell line (R21/B1) containing a truncated human chromosome 6 (6pter-6q21) and a human Y chromosome on a hamster background was irradiated and fused to A23 (TK-) or W3GH (HPRT-) hamster cells. Clones containing expressed HLA class I genes (4/40) were selected using monoclonal antibodies. These clones were recloned and analyzed with a panel of probes from the HLA region. One hybrid (4G6) contained the entire HLA complex. Two other hybrids (4J4 and 4H2) contained only the HLA class I region, while the fourth hybrid (5P9) contained HLA class I and III genes in addition to other genes located in the 6p21 chromosomal region. In situ hybridization showed that the hybrid cells contained more than one fragment of human DNA. Alu and LINE PCR products were derived from these cells and compared to each other as well as to products from two somatic cell hybrids having the 6p21 region in common. The PCR fragments were then screened on conventional Southern blots of the somatic cell hybrids to select a panel of novel probes encompassing the 6p21 region. In addition, the origin of the human DNA fragments in hybrid 4J4 was determined by regional mapping of PCR products

Chromosomes of South American Bufonidae (Amphibia, Anura Chromosomes of South American Bufonidae (Amphibia, Anura

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Brum Zorrilla N.

1973-09-01

Full Text Available Karyotypes of eight of South American Bufonidae were studied: B.ictericus, B. spinulosus spinulosus, B. arenarum, B. g. fernandezae, B. g. d'orbignyi, B. crucifer, B. paracnemis and B. marinus. In all species 2n = 22 chromosomes were found. Neither heteromorphic pairs of chromosomes nor bivalents with characteristic morphology and behavior of sex chromosomesduring male meiosis were observed in any species.Karyotypes of eight of South American Bufonidae were studied: B.ictericus, B. spinulosus spinulosus, B. arenarum, B. g. fernandezae, B. g. d'orbignyi, B. crucifer, B. paracnemis and B. marinus. In all species 2n = 22 chromosomes were found. Neither heteromorphic pairs of chromosomes nor bivalents with characteristic morphology and behavior of sex chromosomesduring male meiosis were observed in any species.

Transfer of Antibiotic Resistance in Staphylococcus aureus

DEFF Research Database (Denmark)

Haaber, Jakob; Penadés, José R; Ingmer, Hanne

2017-01-01

Staphylococcus aureus is a serious human pathogen with remarkable adaptive powers. Antibiotic-resistant clones rapidly emerge mainly by acquisition of antibiotic-resistance genes from other S. aureus strains or even from other genera. Transfer is mediated by a diverse complement of mobile genetic...... of plasmids that can be transferred by conjugation and the efficiency with which transduction occurs. Here, we review the main routes of antibiotic resistance gene transfer in S. aureus in the context of its biology as a human commensal and a life-threatening pathogen. Staphylococcus aureus cells...... are effective in exchanging mobile genetic elements, including antibiotic-resistance genes.During colonization or infection of host organisms, the exchange appears to be particularly effective.Bacteriophage-mediated transfer involves both transduction and autotransduction, which may enable lysogenic S. aureus...

Rise, fall and resurrection of chromosome territories: a historical perspective. Part I. The rise of chromosome territories

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T Cremer

2009-06-01

Full Text Available It is now generally accepted that chromosomes in the cell nucleus are organized in distinct domains, first called chromosome territories in 1909 by the great cytologist Theodor Boveri. Yet, even today chromosomes have remained enigmatic individuals, whose structures, arrangements and functions in cycling and post-mitotic cells still need to be explored in full detail. Whereas numerous recent reviews describe present evidence for a dynamic architecture of chromosome territories and discuss the potential significance within the functional compartmentalization of the nucleus, a comprehensive historical account of this important concept of nuclear organization was lacking so far. Here, we describe the early rise of chromosome territories within the context of the discovery of chromosomes and their fundamental role in heredity, covering a period from the 1870th to the early 20th century (part I, this volume. In part II (next volume we review the abandonment of the chromosome territory concept during the 1950th to 1980th and the compelling evidence, which led to its resurrection during the 1970th to 1980th.

Sixty-five radiation hybrids for the short arm of human chromosome 6: their value as a mapping panel and as a source for rapid isolation of new probes using repeat element-mediated PCR

International Nuclear Information System (INIS)

Zoghbi, H.Y.; McCall, A.E.; LeBorgne-Demarquoy, F.

1991-01-01

We have used an irradiation and fusion procedure to generate somatic cell hybrids that retain fragments of the short arm of human chromosome 6 (6p). To identify hybrids retaining human material, we performed repeat element-mediated PCR on crude lysates of cells from individual clones. Sixty-five hybrids were shown to contain human material and fifty of those contained one or more 6p-specific probes. Detailed characterization of these hybrids identified a subset that divides 6p into ten mapping intervals. Using repeat element-mediated PCR, we were able to isolate and map 61 new DNA fragments from specific regions of 6p. Fifteen of these fragments were used to screen for restriction fragment length polymorphisms (RFLPs), and nine identified RFLPs with one or more enzymes. The radiation hybrids described in this study provide a valuable resource for high-resolution mapping of 6p and for the rapid isolation of region-specific markers

CHROMOSOMES OF WOODY SPECIES

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Julio R Daviña

2000-01-01

Full Text Available Chromosome numbers of nine subtropical woody species collected in Argentina and Paraguay are reported. The counts tor Coutarea hexandra (2n=52, Inga vera subsp. affinis 2n=26 (Fabaceae and Chorisia speciosa 2n=86 (Bombacaceae are reported for the first time. The chromosome number given for Inga semialata 2n=52 is a new cytotype different from the previously reported. Somatic chromosome numbers of the other taxa studied are: Sesbania punicea 2n=12, S. virgata 2n=12 and Pilocarpus pennatifolius 2n=44 from Argentina

Cells with dysfunctional telomeres are susceptible to reactive oxygen species hydrogen peroxide via generation of multichromosomal fusions and chromosomal fragments bearing telomeres

Energy Technology Data Exchange (ETDEWEB)

Woo, Seon Rang [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Department of Biochemistry, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Park, Jeong-Eun; Juhn, Kyoung-Mi; Ju, Yeun-Jin; Jeong, Jaemin [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Kang, Chang-Mo; Yun, Hyun Jin [Division of Radiation Effect, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Yun, Mi Yong; Shin, Hyun-Jin; Joo, Hyun-Yoo; Park, Eun-Ran; Park, In-Chul; Hong, Sung Hee; Hwang, Sang-Gu [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Kim, Haekwon [Department of Biotechnology, Seoul Woman' s University, Seoul 139-774 (Korea, Republic of); Cho, Myung-Haing [Laboratory of Toxicology, College of Veterinary Medicine, Seoul National University, Seoul 151-742 (Korea, Republic of); Kim, Sang Hoon [Department of Biology, Kyung Hee University, Seoul 130-701 (Korea, Republic of); Park, Gil Hong [Department of Biochemistry, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Lee, Kee-Ho, E-mail: khlee@kirams.re.kr [Division of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of)

2012-01-06

Highlights: Black-Right-Pointing-Pointer Under conditions of telomere erosion, cells become extremely sensitive to H{sub 2}O{sub 2}. Black-Right-Pointing-Pointer Chromosomal regions adjacent to telomeres are cleaved by H{sub 2}O{sub 2} under such conditions. Black-Right-Pointing-Pointer H{sub 2}O{sub 2} thus causes multichromosomal fusions and generation of small chromosomal fragments. Black-Right-Pointing-Pointer N-acetylcysteine prevents H{sub 2}O{sub 2}-induced chromosomal aberrations. -- Abstract: During genotoxic stress, reactive oxygen species hydrogen peroxide (H{sub 2}O{sub 2}) is a prime mediator of the DNA damage response. Telomeres function both to assist in DNA damage repair and to inhibit chromosomal end-to-end fusion. Here, we show that telomere dysfunction renders cells susceptible to H{sub 2}O{sub 2}, via generation of multichromosomal fusion and chromosomal fragments. H{sub 2}O{sub 2} caused formation of multichromosomal end-to-end fusions involving more than three chromosomes, preferentially when telomeres were erosive. Interestingly, extensive chromosomal fragmentation (yielding small-sized fragments) occurred only in cells exhibiting such multichromosomal fusions. Telomeres were absent from fusion points, being rather present in the small fragments, indicating that H{sub 2}O{sub 2} cleaves chromosomal regions adjacent to telomeres. Restoration of telomere function or addition of the antioxidant N-acetylcysteine prevented development of chromosomal aberrations and rescued the observed hypersensitivity to H{sub 2}O{sub 2}. Thus, chromosomal regions adjacent to telomeres become sensitive to reactive oxygen species hydrogen peroxide when telomeres are dysfunctional, and are cleaved to produce multichromosomal fusions and small chromosomal fragments bearing the telomeres.

Rise, fall and resurrection of chromosome territories: a historical perspective. Part I. The rise of chromosome territories

OpenAIRE

T Cremer; C Cremer

2009-01-01

It is now generally accepted that chromosomes in the cell nucleus are organized in distinct domains, first called chromosome territories in 1909 by the great cytologist Theodor Boveri. Yet, even today chromosomes have remained enigmatic individuals, whose structures, arrangements and functions in cycling and post-mitotic cells still need to be explored in full detail. Whereas numerous recent reviews describe present evidence for a dynamic architecture of chromosome territories and discuss the...

Chromosome segregation in plant meiosis

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Linda eZamariola

2014-06-01

Full Text Available Faithful chromosome segregation in meiosis is essential for ploidy stability over sexual life cycles. In plants, defective chromosome segregation caused by gene mutations or other factors leads to the formation of unbalanced or unreduced gametes creating aneuploid or polyploid progeny, respectively. Accurate segregation requires the coordinated execution of conserved processes occurring throughout the two meiotic cell divisions. Synapsis and recombination ensure the establishment of chiasmata that hold homologous chromosomes together allowing their correct segregation in the first meiotic division, which is also tightly regulated by cell-cycle dependent release of cohesin and monopolar attachment of sister kinetochores to microtubules. In meiosis II, bi-orientation of sister kinetochores and proper spindle orientation correctly segregate chromosomes in four haploid cells. Checkpoint mechanisms acting at kinetochores control the accuracy of kinetochore-microtubule attachment, thus ensuring the completion of segregation. Here we review the current knowledge on the processes taking place during chromosome segregation in plant meiosis, focusing on the characterization of the molecular factors involved.

Cellular Immune Response Against Firefly Luciferase After Sleeping Beauty–Mediated Gene Transfer In Vivo

Science.gov (United States)

Podetz-Pedersen, Kelly M.; Vezys, Vaiva; Somia, Nikunj V.; Russell, Stephen J.

2014-01-01

Abstract The Sleeping Beauty (SB) transposon system has been shown to mediate new gene sequence integration resulting in long-term expression. Here the effectiveness of hyperactive SB100X transposase was tested, and we found that hydrodynamic co-delivery of a firefly luciferase transposon (pT2/CaL) along with SB100X transposase (pCMV-SB100X) resulted in remarkably sustained, high levels of luciferase expression. However, after 4 weeks there was a rapid, animal-by-animal loss of luciferase expression that was not observed in immunodeficient mice. We hypothesized that this sustained, high-level luciferase expression achieved using the SB100X transposase elicits an immune response in pT2/CaL co-administered mice, which was supported by the rapid loss of luciferase expression upon challenge of previously treated animals and in naive animals adoptively transferred with splenocytes from previously treated animals. Specificity of the immune response to luciferase was demonstrated by increased cytokine expression in splenocytes after exposure to luciferase peptide in parallel with MHC I–luciferase peptide tetramer binding. This anti-luciferase immune response observed following continuous, high-level luciferase expression in vivo clearly impacts its use as an in vivo reporter. As both an immunogen and an extremely sensitive reporter, luciferase is also a useful model system for the study of immune responses following in vivo gene transfer and expression. PMID:25093708

Isolation and characterization of DNA probes from a flow-sorted human chromosome 8 library that detect restriction fragment length polymorphism (RFLP).

Science.gov (United States)

Wood, S; Starr, T V; Shukin, R J

1986-01-01

We have used a recombinant DNA library constructed from flow-sorted human chromosome 8 as a source of single-copy human probes. These probes have been screened for restriction fragment length polymorphism (RFLP) by hybridization to Southern transfers of genomic DNA from five unrelated individuals. We have detected six RFLPs distributed among four probes after screening 741 base pairs for restriction site variation. These RFLPs all behave as codominant Mendelian alleles. Two of the probes detect rare variants, while the other two detect RFLPs with PIC values of .36 and .16. Informative probes will be useful for the construction of a linkage map for chromosome 8 and for the localization of mutant alleles to this chromosome. Images Fig. 1 PMID:2879441

Recent Male-Mediated Gene Flow over a Linguistic Barrier in Iberia, Suggested by Analysis of a Y-Chromosomal DNA Polymorphism

Science.gov (United States)

Hurles, Matthew E.; Veitia, Reiner; Arroyo, Eduardo; Armenteros, Manuel; Bertranpetit, Jaume; Pérez-Lezaun, Anna; Bosch, Elena; Shlumukova, Maria; Cambon-Thomsen, Anne; McElreavey, Ken; López de Munain, Adolfo; Röhl, Arne; Wilson, Ian J.; Singh, Lalji; Pandya, Arpita; Santos, Fabrício R.; Tyler-Smith, Chris; Jobling, Mark A.

1999-01-01

Summary We have examined the worldwide distribution of a Y-chromosomal base-substitution polymorphism, the T/C transition at SRY-2627, where the T allele defines haplogroup 22; sequencing of primate homologues shows that the ancestral state cannot be determined unambiguously but is probably the C allele. Of 1,191 human Y chromosomes analyzed, 33 belong to haplogroup 22. Twenty-nine come from Iberia, and the highest frequencies are in Basques (11%; n=117) and Catalans (22%; n=32). Microsatellite and minisatellite (MSY1) diversity analysis shows that non-Iberian haplogroup-22 chromosomes are not significantly different from Iberian ones. The simplest interpretation of these data is that haplogroup 22 arose in Iberia and that non-Iberian cases reflect Iberian emigrants. Several different methods were used to date the origin of the polymorphism: microsatellite data gave ages of 1,650, 2,700, 3,100, or 3,450 years, and MSY1 gave ages of 1,000, 2,300, or 2,650 years, although 95% confidence intervals on all of these figures are wide. The age of the split between Basque and Catalan haplogroup-22 chromosomes was calculated as only 20% of the age of the lineage as a whole. This study thus provides evidence for direct or indirect gene flow over the substantial linguistic barrier between the Indo-European and non–Indo-European–speaking populations of the Catalans and the Basques, during the past few thousand years. PMID:10521311

Molecular and cytogenetic characterization of the 5DS-5BS chromosome translocation conditioning soft kernel texture in durum wheat

Science.gov (United States)

Cultivar ‘Soft Svevo’, a new non-GMO soft durum cultivar with soft kernel texture, was developed through a 5DS(5BS) chromosomal translocation from event. cv. Chinese Spring, and subsequently used to create new soft durum germplasm. The development of Soft Svevo featured the Ph1b-mediated homoeologou...

Nonequilibrium Chromosome Looping via Molecular Slip Links

Science.gov (United States)

Brackley, C. A.; Johnson, J.; Michieletto, D.; Morozov, A. N.; Nicodemi, M.; Cook, P. R.; Marenduzzo, D.

2017-09-01

We propose a model for the formation of chromatin loops based on the diffusive sliding of molecular slip links. These mimic the behavior of molecules like cohesin, which, along with the CTCF protein, stabilize loops which contribute to organizing the genome. By combining 3D Brownian dynamics simulations and 1D exactly solvable nonequilibrium models, we show that diffusive sliding is sufficient to account for the strong bias in favor of convergent CTCF-mediated chromosome loops observed experimentally. We also find that the diffusive motion of multiple slip links along chromatin is rectified by an intriguing ratchet effect that arises if slip links bind to the chromatin at a preferred "loading site." This emergent collective behavior favors the extrusion of loops which are much larger than the ones formed by single slip links.

GEMC1 is a TopBP1 interacting protein required for chromosomal DNA replication

Science.gov (United States)

Balestrini, Alessia; Cosentino, Claudia; Errico, Alessia; Garner, Elizabeth; Costanzo, Vincenzo

2010-01-01

Many factors required for chromosomal DNA replication have been identified in unicellular eukaryotes. However, DNA replication in complex multicellular organisms is poorly understood. Here, we report the identification of GEMC1, a novel vertebrate protein required for chromosomal DNA replication. GEMC1 is highly conserved in vertebrates and is preferentially expressed in proliferating cells. Using Xenopus egg extract we show that Xenopus GEMC1 (xGEMC1) binds to checkpoint and replication factor TopBP1, which promotes xGEMC1 binding to chromatin during pre-replication complex (pre-RC) formation. We demonstrate that xGEMC1 directly interacts with replication factors such as Cdc45 and Cdk2-CyclinE by which it is heavily phosphorylated. Phosphorylated xGEMC1 stimulates initiation of DNA replication whereas depletion of xGEMC1 prevents DNA replication onset due to impairment of Cdc45 loading onto chromatin. Likewise, inhibition of GEMC1 expression by morpholino and siRNA oligos prevents DNA replication in embryonic and somatic vertebrate cells. These data suggest that GEMC1 promotes initiation of chromosomal DNA replication in higher eukaryotes by mediating TopBP1 and Cdk2 dependent recruitment of Cdc45 onto replication origins. PMID:20383140

Are There Knots in Chromosomes?

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Jonathan T. Siebert

2017-08-01

Full Text Available Recent developments have for the first time allowed the determination of three-dimensional structures of individual chromosomes and genomes in nuclei of single haploid mouse embryonic stem (ES cells based on Hi–C chromosome conformation contact data. Although these first structures have a relatively low resolution, they provide the first experimental data that can be used to study chromosome and intact genome folding. Here we further analyze these structures and provide the first evidence that G1 phase chromosomes are knotted, consistent with the fact that plots of contact probability vs sequence separation show a power law dependence that is intermediate between that of a fractal globule and an equilibrium structure.

Chromosome condensation and segmentation

International Nuclear Information System (INIS)

Viegas-Pequignot, E.M.

1981-01-01

Some aspects of chromosome condensation in mammalians -humans especially- were studied by means of cytogenetic techniques of chromosome banding. Two further approaches were adopted: a study of normal condensation as early as prophase, and an analysis of chromosome segmentation induced by physical (temperature and γ-rays) or chemical agents (base analogues, antibiotics, ...) in order to show out the factors liable to affect condensation. Here 'segmentation' means an abnormal chromosome condensation appearing systematically and being reproducible. The study of normal condensation was made possible by the development of a technique based on cell synchronization by thymidine and giving prophasic and prometaphasic cells. Besides, the possibility of inducing R-banding segmentations on these cells by BrdU (5-bromodeoxyuridine) allowed a much finer analysis of karyotypes. Another technique was developed using 5-ACR (5-azacytidine), it allowed to induce a segmentation similar to the one obtained using BrdU and identify heterochromatic areas rich in G-C bases pairs [fr

Preimplantation genetic diagnosis for chromosomal rearrangements with the use of array comparative genomic hybridization at the blastocyst stage.

Science.gov (United States)

Christodoulou, Christodoulos; Dheedene, Annelies; Heindryckx, Björn; van Nieuwerburgh, Filip; Deforce, Dieter; De Sutter, Petra; Menten, Björn; Van den Abbeel, Etienne

2017-01-01

To establish the value of array comparative genomic hybridization (CGH) for preimplantation genetic diagnosis (PGD) in embryos of translocation carriers in combination with vitrification and frozen embryo transfer in nonstimulated cycles. Retrospective data analysis study. Academic centers for reproductive medicine and genetics. Thirty-four couples undergoing PGD for chromosomal rearrangements from October 2013 to December 2015. Trophectoderm biopsy at day 5 or day 6 of embryo development and subsequently whole genome amplification and array CGH were performed. This approach revealed a high occurrence of aneuploidies and structural rearrangements unrelated to the parental rearrangement. Nevertheless, we observed a benefit in pregnancy rates of these couples. We detected chromosomal abnormalities in 133/207 embryos (64.2% of successfully amplified), and 74 showed a normal microarray profile (35.7%). In 48 of the 133 abnormal embryos (36.1%), an unbalanced rearrangement originating from the parental translocation was identified. Interestingly, 34.6% of the abnormal embryos (46/133) harbored chromosome rearrangements that were not directly linked to the parental translocation in question. We also detected a combination of unbalanced parental-derived rearrangements and aneuploidies in 27 of the 133 abnormal embryos (20.3%). The use of trophectoderm biopsy at the blastocyst stage is less detrimental to the survival of the embryo and leads to a more reliable estimate of the genomic content of the embryo than cleavage-stage biopsy. In this small cohort PGD study, we describe the successful implementation of array CGH analysis of blastocysts in patients with a chromosomal rearrangement to identify euploid embryos for transfer. Copyright © 2016 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Development of a quantitative pachytene chromosome map and its unification with somatic chromosome and linkage maps of rice (Oryza sativa L.).

Science.gov (United States)

Ohmido, Nobuko; Iwata, Aiko; Kato, Seiji; Wako, Toshiyuki; Fukui, Kiichi

2018-01-01

A quantitative pachytene chromosome map of rice (Oryza sativa L.) was developed using imaging methods. The map depicts not only distribution patterns of chromomeres specific to pachytene chromosomes, but also the higher order information of chromosomal structures, such as heterochromatin (condensed regions), euchromatin (decondensed regions), the primary constrictions (centromeres), and the secondary constriction (nucleolar organizing regions, NOR). These features were image analyzed and quantitatively mapped onto the map by Chromosome Image Analyzing System ver. 4.0 (CHIAS IV). Correlation between H3K9me2, an epigenetic marker and formation and/or maintenance of heterochromatin, thus was, clearly visualized. Then the pachytene chromosome map was unified with the existing somatic chromosome and linkage maps by physically mapping common DNA markers among them, such as a rice A genome specific tandem repeat sequence (TrsA), 5S and 45S ribosomal RNA genes, five bacterial artificial chromosome (BAC) clones, four P1 bacteriophage artificial chromosome (PAC) clones using multicolor fluorescence in situ hybridization (FISH). Detailed comparison between the locations of the DNA probes on the pachytene chromosomes using multicolor FISH, and the linkage map enabled determination of the chromosome number and short/long arms of individual pachytene chromosomes using the chromosome number and arm assignment designated for the linkage map. As a result, the quantitative pachytene chromosome map was unified with two other major rice chromosome maps representing somatic prometaphase chromosomes and genetic linkages. In conclusion, the unification of the three rice maps serves as an indispensable basic information, not only for an in-depth comparison between genetic and chromosomal data, but also for practical breeding programs.

Diagnostic radiation and chromosome aberrations

International Nuclear Information System (INIS)

Patil, S.R.; Hecht, F.; Lubs, H.A.; Kimberling, W.; Brown, J.; Gerald, P.S.; Summitt, R.L.

1977-01-01

Some evidence is presented suggesting that diagnostic X-rays may be important in the origin of a new chromosomal abnormality other than Down syndrome. Chromosome analyses have been carried out on 4342 children, seven or eight years old. Maternal diagnostic irradiation in the year before conception and up to third lunar month of the index pregnancy was recorded, before the chromosome study began, together with a large amount of family and clinical data. Information on X-ray exposure was supplied by the mothers, s o radiation dosage could not be estimated. 21 children (including a pair of twins and a pair of siblings) born to 19 mothers had chromosomal aberrations. The mothers of six children with inherited translocations, rearrangements and XYY karyotypes were excluded, and 3 (23%) of the remaining 13 mothers had received abdominal and pelvic X-ray exposures. In the whole sample, however, only 6% of the mothers had diagnostic irradiation. Two of these mothers, aged sixteen and twenty, gave birth to a child each with de-novo autosomal translocations, and the third mother, aged thirty-two, had a child with a complex mosaicism involving one X chromosome. Although the sample size of the mothers with chromosomally abnormal children is small, the results are significant. (U.K.)

Diagnostic radiation and chromosome aberrations

Energy Technology Data Exchange (ETDEWEB)

Patil, S R; Hecht, F [Dept. of Pediatrics, Child Development and Rehabilitation Center, Univ. of Oregon Health Sciences Center, Portland, Oregon (USA); Lubs, H A; Kimberling, W; Brown, J; Gerald, P S; Summitt, R L

1977-01-15

Some evidence is presented suggesting that diagnostic X-rays may be important in the origin of a new chromosomal abnormality other than Down syndrome. Chromosome analyses have been carried out on 4342 children, seven or eight years old. Maternal diagnostic irradiation in the year before conception and up to third lunar month of the index pregnancy was recorded, before the chromosome study began, together with a large amount of family and clinical data. Information on X-ray exposure was supplied by the mothers, so radiation dosage could not be estimated. 21 children (including a pair of twins and a pair of siblings) born to 19 mothers had chromosomal aberrations. The mothers of six children with inherited translocations, rearrangements and XYY karyotypes were excluded, and 3 (23%) of the remaining 13 mothers had received abdominal and pelvic X-ray exposures. In the whole sample, however, only 6% of the mothers had diagnostic irradiation. Two of these mothers, aged sixteen and twenty, gave birth to a child each with de-novo autosomal translocations, and the third mother, aged thirty-two, had a child with a complex mosaicism involving one X chromosome. Although the sample size of the mothers with chromosomally abnormal children is small, the results are significant.

Updating the maize karyotype by chromosome DNA sizing

Science.gov (United States)

2018-01-01

The karyotype is a basic concept regarding the genome, fundamentally described by the number and morphological features of all chromosomes. Chromosome class, centromeric index, intra- and interchromosomal asymmetry index, and constriction localization are important in clinical, systematic and evolutionary approaches. In spite of the advances in karyotype characterization made over the last years, new data about the chromosomes can be generated from quantitative methods, such as image cytometry. Therefore, using Zea mays L., this study aimed to update the species’ karyotype by supplementing information on chromosome DNA sizing. After adjustment of the procedures, chromosome morphometry and class as well as knob localization enabled describing the Z. mays karyotype. In addition, applying image cytometry, DNA sizing was unprecedentedly measured for the arms and satellite of all chromosomes. This way, unambiguous identification of the chromosome pairs, and hence the assembly of 51 karyograms, were only possible after the DNA sizing of each chromosome, their arms and satellite portions. These accurate, quantitative and reproducible data also enabled determining the distribution and variation of DNA content in each chromosome. From this, a correlation between DNA amount and total chromosome length evidenced that the mean DNA content of chromosome 9 was higher than that of chromosome 8. The chromosomal DNA sizing updated the Z. mays karyotype, providing insights into its dynamic genome with regards to the organization of the ten chromosomes and their respective portions. Considering the results and the relevance of cytogenetics in the current scenario of comparative sequencing and genomics, chromosomal DNA sizing should be incorporated as an additional parameter for karyotype definition. Based on this study, it can be affirmed that cytogenetic approaches go beyond the simple morphological description of chromosomes. PMID:29293613

Continuum-mediated dark matter–baryon scattering

CERN Document Server

Katz, Andrey; Sajjad, Aqil

2016-01-01

Many models of dark matter scattering with baryons may be treated either as a simple contact interaction or as the exchange of a light mediator particle. We study an alternative, in which a continuum of light mediator states may be exchanged. This could arise, for instance, from coupling to a sector which is approximately conformal at the relevant momentum transfer scale. In the non-relativistic effective theory of dark matter-baryon scattering, which is useful for parametrizing direct detection signals, the effect of such continuum mediators is to multiply the amplitude by a function of the momentum transfer q, which in the simplest case is just a power law. We develop the basic framework and study two examples: the case where the mediator is a scalar operator coupling to the Higgs portal (which turns out to be highly constrained) and the case of an antisymmetric tensor operator ${\\cal O}_{\\mu \

Phage-inducible chromosomal islands are ubiquitous within the bacterial universe.

Science.gov (United States)

Fillol-Salom, Alfred; Martínez-Rubio, Roser; Abdulrahman, Rezheen F; Chen, John; Davies, Robert; Penadés, José R

2018-06-06

Phage-inducible chromosomal islands (PICIs) are a recently discovered family of pathogenicity islands that contribute substantively to horizontal gene transfer, host adaptation and virulence in Gram-positive cocci. Here we report that similar elements also occur widely in Gram-negative bacteria. As with the PICIs from Gram-positive cocci, their uniqueness is defined by a constellation of features: unique and specific attachment sites, exclusive PICI genes, a phage-dependent mechanism of induction, conserved replication origin organization, convergent mechanisms of phage interference, and specific packaging of PICI DNA into phage-like infectious particles, resulting in very high transfer frequencies. We suggest that the PICIs represent two or more distinct lineages, have spread widely throughout the bacterial world, and have diverged much more slowly than their host organisms or their prophage cousins. Overall, these findings represent the discovery of a universal class of mobile genetic elements.

H-NS Facilitates Sequence Diversification of Horizontally Transferred DNAs during Their Integration in Host Chromosomes.

OpenAIRE

Koichi Higashi; Toru Tobe; Akinori Kanai; Ebru Uyar; Shu Ishikawa; Yutaka Suzuki; Naotake Ogasawara; Ken Kurokawa; Taku Oshima

2016-01-01

Bacteria can acquire new traits through horizontal gene transfer. Inappropriate expression of transferred genes, however, can disrupt the physiology of the host bacteria. To reduce this risk, Escherichia coli expresses the nucleoid-associated protein, H-NS, which preferentially binds to horizontally transferred genes to control their expression. Once expression is optimized, the horizontally transferred genes may actually contribute to E. coli survival in new habitats. Therefore, we investiga...

Automation of chromosomes analysis. Automatic system for image processing

International Nuclear Information System (INIS)

Le Go, R.; Cosnac, B. de; Spiwack, A.

1975-01-01

The A.S.T.I. is an automatic system relating to the fast conversational processing of all kinds of images (cells, chromosomes) converted to a numerical data set (120000 points, 16 grey levels stored in a MOS memory) through a fast D.O. analyzer. The system performs automatically the isolation of any individual image, the area and weighted area of which are computed. These results are directly displayed on the command panel and can be transferred to a mini-computer for further computations. A bright spot allows parts of an image to be picked out and the results to be displayed. This study is particularly directed towards automatic karyo-typing [fr

Chromosomal Targeting by the Type III-A CRISPR-Cas System Can Reshape Genomes in Staphylococcus aureus.

Science.gov (United States)

Guan, Jing; Wang, Wanying; Sun, Baolin

2017-01-01

CRISPR-Cas (clustered regularly interspaced short palindromic repeat [CRISPR]-CRISPR-associated protein [Cas]) systems can provide protection against invading genetic elements by using CRISPR RNAs (crRNAs) as a guide to locate and degrade the target DNA. CRISPR-Cas systems have been classified into two classes and five types according to the content of cas genes. Previous studies have indicated that CRISPR-Cas systems can avoid viral infection and block plasmid transfer. Here we show that chromosomal targeting by the Staphylococcus aureus type III-A CRISPR-Cas system can drive large-scale genome deletion and alteration within integrated staphylococcal cassette chromosome mec (SCC mec ). The targeting activity of the CRISPR-Cas system is associated with the complementarity between crRNAs and protospacers, and 10- to 13-nucleotide truncations of spacers partially block CRISPR attack and more than 13-nucleotide truncation can fully abolish targeting, suggesting that a minimal length is required to license cleavage. Avoiding base pairings in the upstream region of protospacers is also necessary for CRISPR targeting. Successive trinucleotide complementarity between the 5' tag of crRNAs and protospacers can disrupt targeting. Our findings reveal that type III-A CRISPR-Cas systems can modulate bacterial genome stability and may serve as a high-efficiency tool for deleting resistance or virulence genes in bacteria. IMPORTANCE Staphylococcus aureus is a pathogen that can cause a wide range of infections in humans. Studies have suggested that CRISPR-Cas systems can drive the loss of integrated mobile genetic elements (MGEs) by chromosomal targeting. Here we demonstrate that CRISPR-mediated cleavage contributes to the partial deletion of integrated SCC mec in methicillin-resistant S. aureus (MRSA), which provides a strategy for the treatment of MRSA infections. The spacer within artificial CRISPR arrays should contain more than 25 nucleotides for immunity, and consecutive

CHROMOSOMAL DIFFERENTIATIONS OF THE LAMPBRUSH TYPE FORMED BY THE Y CHROMOSOME IN DROSOPHILA HYDEI AND DROSOPHILA NEOHYDEI

Science.gov (United States)

Hess, Oswald; Meyer, Günther F.

1963-01-01

The nuclei of growing spermatocytes in Drosophila hydei and D. neohydei are characterized by the appearance of phase-specific, paired, loop-shaped structures thought to be similar to the loops in lampbrush chromosomes of amphibian oocytes. In X/O-males of D. hydei spermatogenesis is completely blocked before the first maturation division. No spermatozoa are formed in such testes. In the nuclei of X/O-spermatocytes, paired loop formations are absent. This shows the dependence of these chromosomal functional structures upon the Y chromosome. The basis of this dependence could be shown through an investigation of males with two Y chromosomes. All loop pairs are present in duplicate in XYY males. This proves that the intranuclear formations are structural modifications of the Y chromosome itself. These functional structures are species-specific and characteristically different in Drosophila hydei and D. neohydei. Reciprocal species crosses and a backcross showed that the spermatocyte nuclei of all hybrid males possess the functional structures corresponding to the species which donated the Y chromosome. This shows that the morphological character of the functional structures is also determined by the Y chromosome. PMID:13954225

Comparison of initial DNA (Chromosome) damage/repair in cells exposed to heavy ion particles and X-rays

International Nuclear Information System (INIS)

Okayasu, Ryuichi; Okada, Maki; Noguchi, Mitsuho; Saito, Shiori; Okabe, Atsushi; Takakura, Kahoru

2005-01-01

We have studied cell survival and chromosome damage/repair in normal and non homologous end-joining (NHEJ) deficient human cells exposed to carbon ions (290 MeV/u, ∼70 keV/um), iron ions (500 MeV/u, ∼200 keV/um) and X-rays. In order to examine the effect of heavy ion on double strand break (DSB) repair machinery, the auto-phosphorylation of DNA-PKcs was also investigated. The important discoveries made during this period are: 200 keV/um iron irradiation induced additional molecular damage beyond that 70 keV/um carbon did. Iron irradiation not only caused an inefficient G1 chromosome repair, but also induced non-repairable DSB/chromosome damage. The auto-phosphorylation of DNA-PKcs was significantly affected by high linear energy transfer (LET) irradiation when compared to X-rays. These results indicate NHEJ machinery was markedly disturbed by high LET radiation when compared to low LET radiation. (author)

Chromosomal study of lettuce and its allied species (Lactuca spp., Asteraceae) by means of karyotype analysis and fluorescence in situ hybridization.

Science.gov (United States)

Matoba, Hideyuki; Mizutani, Takayuki; Nagano, Katsuya; Hoshi, Yoshikazu; Uchiyama, Hiroshi

2007-12-01

In this study, in addition to the karyotype analysis, the chromosomal distributions of 5 S and 18 S rDNAs, and the Arabidopsis-type (T3AG3) telomeric sequences were detected by means of fluorescence in situ hybridization (FISH) to promote the information of chromosomal organization and evolution in the cultivated lettuce and its wild relatives, L. sativa, L. serriola, L. saligna and L. virosa. The karyotype analysis revealed the dissimilarity between L. virosa and the remaining species. In all four Lactuca species studied, one 5 S rDNA and two 18 S rDNA loci were detected. The simultaneous FISH of 5 S and 18 S rDNAs revealed that both rDNA loci of L. sativa, L. serriola and L. saligna were identical, however, that of L. virosa was different from the other species. These analyses indicate the closer relationships between L. sativa/L. serriola and L. saligna rather than L. virosa. Arabidopsis-type telomeric sequences were detected at both ends of their chromatids of all chromosomes not in the other regions. This observation suggests the lack of telomere-mediated chromosomal rearrangements among the Lactuca chromosomes.

Genome-wide detection of chromosomal rearrangements, indels, and mutations in circular chromosomes by short read sequencing

DEFF Research Database (Denmark)

Skovgaard, Ole; Bak, Mads; Løbner-Olesen, Anders

2011-01-01

a combination of WGS and genome copy number analysis, for the identification of mutations that suppress the growth deficiency imposed by excessive initiations from the Escherichia coli origin of replication, oriC. The E. coli chromosome, like the majority of bacterial chromosomes, is circular, and DNA...... replication is initiated by assembling two replication complexes at the origin, oriC. These complexes then replicate the chromosome bidirectionally toward the terminus, ter. In a population of growing cells, this results in a copy number gradient, so that origin-proximal sequences are more frequent than...... origin-distal sequences. Major rearrangements in the chromosome are, therefore, readily identified by changes in copy number, i.e., certain sequences become over- or under-represented. Of the eight mutations analyzed in detail here, six were found to affect a single gene only, one was a large chromosomal...

Blastocyst Morphology Holds Clues Concerning The Chromosomal Status of The Embryo

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Rita de Cassia Savio Figueira

2015-07-01

Full Text Available Background: Embryo morphology has been proposed as an alternative marker of chromosomal status. The objective of this retrospective cohort study was to investigate the association between the chromosomal status on day 3 of embryo development and blastocyst morphology. Materials and Methods: A total of 596 embryos obtained from 106 cycles of intracytoplasmic sperm injection (ICSI followed by preimplantation genetic aneuploidy screening (PGS were included in this retrospective study. We evaluated the relationship between blastocyst morphological features and embryonic chromosomal alteration. Results: Of the 564 embryos with fluorescent in situ hybridization (FISH results, 200 reached the blastocyst stage on day 5 of development. There was a significantly higher proportion of euploid embryos in those that achieved the blastocyst stage (59.0% compared to embryos that did not develop to blastocysts (41.2% on day 5 (P<0.001. Regarding blastocyst morphology, we observed that all embryos that had an abnormal inner cell mass (ICM were aneuploid. Embryos with morphologically normal ICM had a significantly higher euploidy rate (62.1%, P<0.001. As regards to the trophectoderm (TE morphology, an increased rate of euploidy was observed in embryos that had normal TE (65.8% compared to embryos with abnormal TE (37.5%, P<0.001. Finally, we observed a two-fold increase in the euploidy rate in high-quality blastocysts with both high-quality ICM and TE (70.4% compared to that found in low-quality blastocysts (31.0%, P<0.001. Conclusion: Chromosomal abnormalities do not impair embryo development as aneuploidy is frequently observed in embryos that reach the blastocyst stage. A high-quality blastocyst does not represent euploidy of chromosomes 13, 14, 15, 16, 18, 21, 22, X and Y. However, aneuploidy is associated with abnormalities in the ICM morphology. Further studies are necessary to confirm whether or not the transfer of blastocysts with low-quality ICM should be

Exceptional Complex Chromosomal Rearrangements in Three Generations

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Hannie Kartapradja

2015-01-01

Full Text Available We report an exceptional complex chromosomal rearrangement (CCR found in three individuals in a family that involves 4 chromosomes with 5 breakpoints. The CCR was ascertained in a phenotypically abnormal newborn with additional chromosomal material on the short arm of chromosome 4. Maternal karyotyping indicated that the mother carried an apparently balanced CCR involving chromosomes 4, 6, 11, and 18. Maternal transmission of the derivative chromosome 4 resulted in partial trisomy for chromosomes 6q and 18q and a partial monosomy of chromosome 4p in the proband. Further family studies found that the maternal grandmother carried the same apparently balanced CCR as the proband’s mother, which was confirmed using the whole chromosome painting (WCP FISH. High resolution whole genome microarray analysis of DNA from the proband’s mother found no evidence for copy number imbalance in the vicinity of the CCR translocation breakpoints, or elsewhere in the genome, providing evidence that the mother’s and grandmother’s CCRs were balanced at a molecular level. This structural rearrangement can be categorized as an exceptional CCR due to its complexity and is a rare example of an exceptional CCR being transmitted in balanced and/or unbalanced form across three generations.

Non-disjunction of chromosome 13

DEFF Research Database (Denmark)

Bugge, Merete; Collins, Andrew; Hertz, Jens Michael

2007-01-01

We performed a molecular study with 21 microsatellites on a sample of 82 trisomy 13 conceptuses, the largest number of cases studied to date. The parental origin was determined in every case and in 89% the extra chromosome 13 was of maternal origin with an almost equal number of maternal MI and MII...... recombination in both maternal MI and MII errors and the former is associated with a significant number of tetrads (33%) that are nullichiasmate, which do not appear to be a feature of normal chromosome 13 meiosis. This study supports the evidence for subtle chromosome-specific influences on the mechanisms...... that determine non-disjunction of human chromosomes, consistent with the diversity of findings for other trisomies. Udgivelsesdato: 2007-Aug-15...

Boundary Associated Long Noncoding RNA Mediates Long-Range Chromosomal Interactions.

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Ifeoma Jane Nwigwe

Full Text Available CCCTC binding factor (CTCF is involved in organizing chromosomes into mega base-sized, topologically associated domains (TADs along with other factors that define sub-TAD organization. CTCF-Cohesin interactions have been shown to be critical for transcription insulation activity as it stabilizes long-range interactions to promote proper gene expression. Previous studies suggest that heterochromatin boundary activity of CTCF may be independent of Cohesin, and there may be additional mechanisms for defining topological domains. Here, we show that a boundary site we previously identified known as CTCF binding site 5 (CBS5 from the homeotic gene cluster A (HOXA locus exhibits robust promoter activity. This promoter activity from the CBS5 boundary element generates a long noncoding RNA that we designate as boundary associated long noncoding RNA-1 (blncRNA1. Functional characterization of this RNA suggests that the transcript stabilizes long-range interactions at the HOXA locus and promotes proper expression of HOXA genes. Additionally, our functional analysis also shows that this RNA is not needed in the stabilization of CTCF-Cohesin interactions however CTCF-Cohesin interactions are critical in the transcription of blncRNA1. Thus, the CTCF-associated boundary element, CBS5, employs both Cohesin and noncoding RNA to establish and maintain topologically associated domains at the HOXA locus.

Chromatin structure and ionizing-radiation-induced chromosome aberrations

International Nuclear Information System (INIS)

Muehlmann-Diaz, M.C.

1993-01-01

The possible influence of chromatic structure or activity on chromosomal radiosensitivity was studied. A cell line was isolated which contained some 10 5 copies of an amplified plasmid in a single large mosquito artificial chromosome (MAC). This chromosome was hypersensitive to DNase I. Its radiosensitivity was some three fold greater than normal mosquito chromosomes in the same cell. In cultured human cells irradiated during G 0 , the initial breakage frequency in chromosome 4, 19 and the euchromatic and heterochromatic portions of the Y chromosome were measured over a wide range of doses by inducing Premature Chromosome Condensation (PCC) immediately after irradiation with Cs-137 gamma rays. No evidence was seen that Y heterochromatin or large fragments of it remained unbroken. The only significant deviation from the expected initial breakage frequency per Gy per unit length of chromosome was that observed for the euchromatic portion of the Y chromosome, with breakage nearly twice that expected. The development of aberrations involving X and Y chromosomes at the first mitosis after irradation was also studied. Normal female cells sustained about twice the frequency of aberrations involving X chromosomes for a dose of 7.3 Gy than the corresponding male cells. Fibroblasts from individuals with supernumerary X chromosomes did not show any further increase in X aberrations for this dos. The frequency of aberrations involving the heterochromatic portion of the long arm of the Y chromosome was about what would be expected for a similar length of autosome, but the euchromatic portion of the Y was about 3 times more radiosensitive per unit length. 5-Azacytidine treatment of cultured human female fibroblasts or fibroblasts from a 49,XXXXY individual, reduced the methylation of cytosine residues in DNA, and resulted in an increased chromosomal radiosensitivity in general, but it did not increase the frequency of aberrations involving the X chromosomes

Plasmid-mediated quinolone resistance; interactions between human, animal and environmental ecologies

Directory of Open Access Journals (Sweden)

Laurent ePOIREL

2012-02-01

Full Text Available Resistance to quinolones and fluoroquinolones is being increasingly reported among human but also veterinary isolates during the last two to three decades, very likely as a consequence of the large clinical usage of those antibiotics. Even if the principle mechanisms of resistance to quinolones are chromosome-encoded, due to modifications of molecular targets (DNA gyrase and topoisomerase IV, decreased outer-membrane permeability (porin defect and overexpression of naturally-occurring efflux, the emergence of plasmid-mediated quinolone resistance (PMQR has been reported since 1998. Although these PMQR determinants confer low-level resistance to quinolones and/or fluoroquinolones, they are a favorable background for selection of additional chromosome-encoded quinolone resistance mechanisms. Different transferable mechanisms have been identified, corresponding to the production of Qnr proteins, of the aminoglycoside acetyltransferase AAC(6’-Ib-cr, or of the QepA-type or OqxAB-type efflux pumps. Qnr proteins protect target enzymes (DNA gyrase and type IV topoisomerase from quinolone inhibition (mostly nalidixic acid. The AAC(6’-Ib-cr determinant acetylates several fluoroquinolones, such as norfloxacin and ciprofloxacin. Finally, the QepA and OqxAB efflux pumps extrude fluoroquinolones from the bacterial cell. A series of studies have identified the environment to be a reservoir of PMQR genes, with farm animals and aquatic habitats being significantly involved. In addition, the origin of the qnr genes has been identified, corresponding to the waterborne species Shewanella sp. Altogether, the recent observations suggest that the aquatic environment might constitute the original source of PMQR genes, that would secondly spread among animal or human isolates.

Catabolite-mediated mutations in alternate toluene degradative pathways in Pseudomonas putida.

Science.gov (United States)

Leddy, M B; Phipps, D W; Ridgway, H F

1995-01-01

Pseudomonas putida 54g grew on mineral salts with toluene and exhibited catechol-2,3-dioxygenase (C23O) activity, indicating a meta pathway. After 10 to 15 days on toluene, nondegrading (Tol-) variants approached nearly 10% of total CFU. Auxotrophs were not detected among variants, suggesting selective loss of catabolic function(s). Variant formation was substrate dependent, since Tol- cells were observed on neither ethylbenzene, glucose, nor peptone-based media nor when toluene catabolism was suppressed by glucose. Unlike wild-type cells, variants did not grow on gasoline, toluene, benzene, ethylbenzene, benzoate, or catechol, suggesting loss of meta pathway function. Catabolic and C23O activities were restored to variants via transfer of a 78-mDa TOL-like plasmid from a wild-type Tol+ donor. Tests for reversion of variants to Tol+ were uniformly negative, suggesting possible delection or excision of catabolic genes. Deletions were confirmed in some variants by failure to hybridize with a DNA probe specific for the xylE gene encoding C23O. Cells grown on benzoate remained Tol+ but were C23O- and contained a plasmid of reduced size or were plasmid free, suggesting an alternate chromosomal catabolic pathway, also defective in variants. Cells exposed to benzyl alcohol, the initial oxidation product of toluene, accumulated > 13% variants in 5 days, even when cell division was repressed by nitrogen deprivation to abrogate selection processes. No variants formed in identical ethylbenzene-exposed controls. The results suggest that benzyl alcohol mediates irreversible defects in both a plasmid-associated meta pathway and an alternate chromosomal pathway. PMID:7642499

The contribution of p53 and Y chromosome long arm genes to regulation of apoptosis in mouse testis.

Science.gov (United States)

Lech, Tomasz; Styrna, Józefa; Kotarska, Katarzyna

2018-03-01

Apoptosis of excessive or defective germ cells is a natural process occurring in mammalian testes. Tumour suppressor protein p53 is involved in this process both in developing and adult male gonads. Its contribution to testicular physiology is known to be modified by genetic background. The aim of this study was to evaluate the combined influence of the p53 and Y chromosome long arm genes on male germ cell apoptosis. Knockout of the transformation related protein 53 (Trp53) gene was introduced into congenic strains: B10.BR (intact Y chromosome) and B10.BR-Ydel (Y chromosome with a deletion in the long arm). The level of apoptosis in the testes of 19-day-old and 3-month-old male mice was determined using the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate in situ nick-end labelling (TUNEL) method. The study revealed that although p53 is involved in germ cell apoptosis in peripubertal testes, this process can also be mediated by p53-independent mechanisms. However, activation of p53-independent apoptotic pathways in the absence of the p53 protein requires engagement of the multicopy Yq genes and was not observed in gonads of B10.BR-Ydel-p53-/- males. The role of Yq genes in the regulation of testicular apoptosis seems to be restricted to the initial wave of spermatogenesis and is not evident in adult gonads. The study confirmed, instead, that p53 does participate in spontaneous apoptosis in mature testes.

Computer aided analysis of additional chromosome aberrations in Philadelphia chromosome positive acute lymphoblastic leukaemia using a simplified computer readable cytogenetic notation

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Mohr Brigitte

2003-01-01

Full Text Available Abstract Background The analysis of complex cytogenetic databases of distinct leukaemia entities may help to detect rare recurring chromosome aberrations, minimal common regions of gains and losses, and also hot spots of genomic rearrangements. The patterns of the karyotype alterations may provide insights into the genetic pathways of disease progression. Results We developed a simplified computer readable cytogenetic notation (SCCN by which chromosome findings are normalised at a resolution of 400 bands. Lost or gained chromosomes or chromosome segments are specified in detail, and ranges of chromosome breakpoint assignments are recorded. Software modules were written to summarise the recorded chromosome changes with regard to the respective chromosome involvement. To assess the degree of karyotype alterations the ploidy levels and numbers of numerical and structural changes were recorded separately, and summarised in a complex karyotype aberration score (CKAS. The SCCN and CKAS were used to analyse the extend and the spectrum of additional chromosome aberrations in 94 patients with Philadelphia chromosome positive (Ph-positive acute lymphoblastic leukemia (ALL and secondary chromosome anomalies. Dosage changes of chromosomal material represented 92.1% of all additional events. Recurring regions of chromosome losses were identified. Structural rearrangements affecting (pericentromeric chromosome regions were recorded in 24.6% of the cases. Conclusions SCCN and CKAS provide unifying elements between karyotypes and computer processable data formats. They proved to be useful in the investigation of additional chromosome aberrations in Ph-positive ALL, and may represent a step towards full automation of the analysis of large and complex karyotype databases.

Chromosome-specific DNA Repeat Probes

Energy Technology Data Exchange (ETDEWEB)

Baumgartner, Adolf; Weier, Jingly Fung; Weier, Heinz-Ulrich G.

2006-03-16

In research as well as in clinical applications, fluorescence in situ hybridization (FISH) has gained increasing popularity as a highly sensitive technique to study cytogenetic changes. Today, hundreds of commercially available DNA probes serve the basic needs of the biomedical research community. Widespread applications, however, are often limited by the lack of appropriately labeled, specific nucleic acid probes. We describe two approaches for an expeditious preparation of chromosome-specific DNAs and the subsequent probe labeling with reporter molecules of choice. The described techniques allow the preparation of highly specific DNA repeat probes suitable for enumeration of chromosomes in interphase cell nuclei or tissue sections. In addition, there is no need for chromosome enrichment by flow cytometry and sorting or molecular cloning. Our PCR-based method uses either bacterial artificial chromosomes or human genomic DNA as templates with {alpha}-satellite-specific primers. Here we demonstrate the production of fluorochrome-labeled DNA repeat probes specific for human chromosomes 17 and 18 in just a few days without the need for highly specialized equipment and without the limitation to only a few fluorochrome labels.

Novel rat Alzheimer's disease models based on AAV-mediated gene transfer to selectively increase hippocampal Aβ levels

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Dicker Bridget L

2007-06-01

Full Text Available Abstract Background Alzheimer's disease (AD is characterized by a decline in cognitive function and accumulation of amyloid-β peptide (Aβ in extracellular plaques. Mutations in amyloid precursor protein (APP and presenilins alter APP metabolism resulting in accumulation of Aβ42, a peptide essential for the formation of amyloid deposits and proposed to initiate the cascade leading to AD. However, the role of Aβ40, the more prevalent Aβ peptide secreted by cells and a major component of cerebral Aβ deposits, is less clear. In this study, virally-mediated gene transfer was used to selectively increase hippocampal levels of human Aβ42 and Aβ40 in adult Wistar rats, allowing examination of the contribution of each to the cognitive deficits and pathology seen in AD. Results Adeno-associated viral (AAV vectors encoding BRI-Aβ cDNAs were generated resulting in high-level hippocampal expression and secretion of the specific encoded Aβ peptide. As a comparison the effect of AAV-mediated overexpression of APPsw was also examined. Animals were tested for development of learning and memory deficits (open field, Morris water maze, passive avoidance, novel object recognition three months after infusion of AAV. A range of impairments was found, with the most pronounced deficits observed in animals co-injected with both AAV-BRI-Aβ40 and AAV-BRI-Aβ42. Brain tissue was analyzed by ELISA and immunohistochemistry to quantify levels of detergent soluble and insoluble Aβ peptides. BRI-Aβ42 and the combination of BRI-Aβ40+42 overexpression resulted in elevated levels of detergent-insoluble Aβ. No significant increase in detergent-insoluble Aβ was seen in the rats expressing APPsw or BRI-Aβ40. No pathological features were noted in any rats, except the AAV-BRI-Aβ42 rats which showed focal, amorphous, Thioflavin-negative Aβ42 deposits. Conclusion The results show that AAV-mediated gene transfer is a valuable tool to model aspects of AD pathology in

Effects of Spirulina platensis on DNA damage and chromosomal aberration against cadmium chloride-induced genotoxicity in rats.

Science.gov (United States)

Aly, Fayza M; Kotb, Ahmed M; Hammad, Seddik

2018-04-01

Todays, bioactive compounds extracted from Spirulina platensis have been intensively studied for their therapeutical values. Therefore, in the present study, we aimed to evaluate the effects of S. platensis extract on DNA damage and chromosomal aberrations induced by cadmium in rats. Four groups of male albino rats (n = 7 rats) were used. The first group served as a control group and received distilled water. The second group was exposed intraperitoneally to cadmium chloride (CdCl 2 ) (3.5 mg/kg body weight dissolved in 2 ml distilled water). The third group included the rats that were orally treated with S. platensis extract (1 g/kg dissolved in 5 ml distilled water, every other day for 30 days). The fourth group included the rats that were intraperitoneally and orally exposed to cadmium chloride and S. platensis, respectively. The experiment in all groups was extended for 60 days. The results of cadmium-mediated toxicity revealed significant genetic effects (DNA fragmentation, deletion or disappearance of some base pairs of DNA, and appearance of few base pairs according to ISSR-PCR analysis). Moreover, chromosomes showed structural aberrations such as reduction of chromosomal number, chromosomal ring, chromatid deletions, chromosomal fragmentations, and dicentric chromosomes. Surprisingly, S. platensis extract plus CdCl 2 -treated group showed less genetic effects compared with CdCl 2 alone. Further, S. platensis extract upon CdCl 2 toxicity was associated with less chromosomal aberration number and nearly normal appearance of DNA fragments as indicated by the bone marrow and ISSR-PCR analysis, respectively. In conclusion, the present novel study showed that co-treatment with S. platensis extract could reduce the genotoxic effects of CdCl 2 in rats.

Adeno-associated virus-mediated gene transfer.

Science.gov (United States)

Srivastava, Arun

2008-09-01

Although the remarkable versatility and efficacy of recombinant adeno-associated virus 2 (AAV2) vectors in transducing a wide variety of cells and tissues in vitro, and in numerous pre-clinical animal models of human diseases in vivo, have been well established, the published literature is replete with controversies with regard to the efficacy of AAV2 vectors in hematopoietic stem cell (HSC) transduction. A number of factors have contributed to these controversies, the molecular bases of which have begun to come to light in recent years. With the availability of several novel serotypes (AAV1 through AAV12), rational design of AAV capsid mutants, and strategies (self-complementary vector genomes, hematopoietic cell-specific promoters), it is indeed becoming feasible to achieve efficient transduction of HSC by AAV vectors. Using a murine serial bone marrow transplantation model in vivo, we have recently documented stable integration of the proviral AAV genome into mouse chromosomes, which does not lead to any overt hematological abnormalities. Thus, a better understanding of the AAV-HSC interactions, and the availability of a vast repertoire of novel serotype and capsid mutant vectors, are likely to have significant implications in the use of AAV vectors in high-efficiency transduction of HSCs as well as in gene therapy applications involving the hematopoietic system. (c) 2008 Wiley-Liss, Inc.

Chromosome and genome size variation in Luzula (Juncaceae), a genus with holocentric chromosomes

Czech Academy of Sciences Publication Activity Database

Bozek, M.; Leitch, A. R.; Leitch, I. J.; Záveská Drábková, Lenka; Kuta, E.

2012-01-01

Roč. 170, č. 4 (2012), s. 529-541 ISSN 0024-4074 R&D Projects: GA ČR GP206/07/P147 Institutional support: RVO:67985939 Keywords : chromosomal evolution * endopolyploidy * holokinetic chromosome * karyotype evolution * tetraploides * centromeres * TRNF intergenic spacer Subject RIV: EF - Botanics Impact factor: 2.589, year: 2012

mBAND analysis for high- and low-LET radiation-induced chromosome aberrations: A review

Energy Technology Data Exchange (ETDEWEB)

Hada, Megumi, E-mail: megumi.hada-1@nasa.gov [NASA Johnson Space Center, Houston, TX 77058 (United States); Universities Space Research Association, Houston, TX 77058 (United States); Wu Honglu; Cucinotta, Francis A. [NASA Johnson Space Center, Houston, TX 77058 (United States)

2011-06-03

During long-term space travel or cancer therapy, humans are exposed to high linear energy transfer (LET) energetic heavy ions. High-LET radiation is much more effective than low-LET radiation in causing various biological effects, including cell inactivation, genetic mutations, cataracts and cancer induction. Most of these biological endpoints are closely related to chromosomal damage, and cytogenetic damage can be utilized as a biomarker for radiation insults. Epidemiological data, mainly from survivors of the atomic bomb detonations in Japan, have enabled risk estimation from low-LET radiation exposures. The identification of a cytogenetic signature that distinguishes high- from low-LET exposure remains a long-term goal in radiobiology. Recently developed fluorescence in situ hybridization (FISH)-painting methodologies have revealed unique endpoints related to radiation quality. Heavy-ions induce a high fraction of complex-type exchanges, and possibly unique chromosome rearrangements. This review will concentrate on recent data obtained with multicolor banding in situ hybridization (mBAND) methods in mammalian cells exposed to low- and high-LET radiations. Chromosome analysis with mBAND technique allows detection of both inter- and intrachromosomal exchanges, and also distribution of the breakpoints of aberrations.

A high-resolution comparative map between pig chromosome 17 and human chromosomes 4, 8, and 20: Identification of synteny breakpoints

DEFF Research Database (Denmark)

Lahbib-Mansais, Yvette; Karlskov-Mortensen, Peter; Mompart, Florence

2005-01-01

We report on the construction of a high-resolution comparative map of porcine chromosome 17 (SSC17) focusing on evolutionary breakpoints with human chromosomes. The comparative map shows high homology with human chromosome 20 but suggests more limited homologies with other human chromosomes. SSC1...

Cholesterol transfer at endosomal-organelle membrane contact sites.

Science.gov (United States)

Ridgway, Neale D; Zhao, Kexin

2018-06-01

Cholesterol is delivered to the limiting membrane of late endosomes by Niemann-Pick Type C1 and C2 proteins. This review summarizes recent evidence that cholesterol transfer from endosomes to the endoplasmic reticulum and other organelles is mediated by lipid-binding proteins that localize to membrane contact sites (MCS). LDL-cholesterol in the late endosomal/lysosomes is exported to the plasma membrane, where most cholesterol resides, and the endoplasmic reticulum, which harbors the regulatory complexes and enzymes that control the synthesis and esterification of cholesterol. A major advance in dissecting these cholesterol transport pathways was identification of frequent and dynamic MCS between endosomes and the endoplasmic reticulum, peroxisomes and plasma membrane. Positioned at these MCS are members of the oxysterol-binding protein (OSBP) and steroidogenic acute regulatory protein-related lipid-transfer family of lipid transfer proteins that bridge the opposing membranes and directly or indirectly mediate cholesterol transfer. OSBP-related protein 1L (ORP1L), ORP5 and ORP6 mediate cholesterol transfer to the endoplasmic reticulum that regulates cholesterol homeostasis. ORP1L and STARD3 also move cholesterol from the endoplasmic reticulum-to-late endosomal/lysosomes under low-cholesterol conditions to facilitate intraluminal vesicle formation. Cholesterol transport also occurs at MCS with peroxisomes and possibly the plasma membrane. Frequent contacts between organelles and the endo-lysosomal vesicles are sites for bidirectional transfer of cholesterol.