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Sample records for oriented variant identification

  1. Control Oriented System Identification

    DTIC Science & Technology

    1993-08-01

    The research goals for this grant were to obtain algorithms for control oriented system identification is to construct dynamical models of systems...and measured information. Algorithms for this type of nonlinear system identification have been given that produce models suitable for gain scheduled

  2. Identification, mRNA expression, and functional analysis of chitin synthase 1 gene and its two alternative splicing variants in oriental fruit fly, Bactrocera dorsalis.

    PubMed

    Yang, Wen-Jia; Xu, Kang-Kang; Cong, Lin; Wang, Jin-Jun

    2013-01-01

    Two alternative splicing variants of chitin synthase 1 gene (BdCHS1) were cloned and characterized from the oriental fruit fly, Bactrocera dorsalis (Hendel). The cDNA of both variants (BdCHS1a and BdCHS1b) consisted of 5,552 nucleotides (nt), with an open reading frame (ORF) of 4,776 nt, encoding a protein of 1,592 amino acid residues, plus 685- and 88-nt of 5'- and 3'-noncoding regions, respectively. The alternative splicing site was located between positions 3,784-3,960 and formed a pair of mutually exclusive exons (a/b) that were same in size (177 nt), but showed only 65% identity at the nucleotide level. During B. dorsalis growth and development, BdCHS1 and BdCHS1a were both mainly expressed during the larval-pupal and pupal-adult transitions, while BdCHS1b was mainly expressed during pupal-adult metamorphosis and in the middle of the pupal stage. BdCHS1a was predominately expressed in the integument whereas BdCHS1b was mainly expressed in the trachea. The 20-hydroxyecdysone (20E) induced the expression of BdCHS1 and its variants. Injection of dsRNA of BdCHS1, BdCHS1a, and BdCHS1b into third-instar larvae significantly reduced the expression levels of the corresponding variants, generated phenotypic defects, and killed most of the treated larvae. Furthermore, silencing of BdCHS1 and BdCHS1a had a similar result in that the larva was trapped in old cuticle and died without tanning completely, while silencing of BdCHS1b has no effect on insect morphology. These results demonstrated that BdCHS1 plays an important role in the larval-pupal transition and the expression of BdCHS1 in B. dorsalis is regulated by 20E.

  3. The orientation selectivity of face identification

    PubMed Central

    Goffaux, Valerie; Greenwood, John A.

    2016-01-01

    Recent work demonstrates that human face identification is most efficient when based on horizontal, rather than vertical, image structure. Because it is unclear how this specialization for upright (compared to inverted) face processing emerges in the visual system, the present study aimed to systematically characterize the orientation sensitivity profile for face identification. With upright faces, identification performance in a delayed match-to-sample task was highest for horizontally filtered images and declined sharply with oblique and vertically filtered images. Performance was well described by a Gaussian function with a bandwidth around 25°. Face inversion reshaped this sensitivity profile dramatically, with a downward shift of the entire tuning curve as well as a reduction in the amplitude of the horizontal peak and a doubling in bandwidth. The use of naturalistic outer contours (vs. a common outline mask) was also found to reshape this sensitivity profile by increasing sensitivity to oblique information in the near-horizontal range. Altogether, although face identification is sharply tuned to horizontal angles, both inversion and outline masking can profoundly reshape this orientation sensitivity profile. This combination of image- and observer-driven effects provides an insight into the functional relationship between orientation-selective processes within primary and high-level stages of the human brain. PMID:27677359

  4. Re-ranking sequencing variants in the post-GWAS era for accurate causal variant identification.

    PubMed

    Faye, Laura L; Machiela, Mitchell J; Kraft, Peter; Bull, Shelley B; Sun, Lei

    2013-01-01

    Next generation sequencing has dramatically increased our ability to localize disease-causing variants by providing base-pair level information at costs increasingly feasible for the large sample sizes required to detect complex-trait associations. Yet, identification of causal variants within an established region of association remains a challenge. Counter-intuitively, certain factors that increase power to detect an associated region can decrease power to localize the causal variant. First, combining GWAS with imputation or low coverage sequencing to achieve the large sample sizes required for high power can have the unintended effect of producing differential genotyping error among SNPs. This tends to bias the relative evidence for association toward better genotyped SNPs. Second, re-use of GWAS data for fine-mapping exploits previous findings to ensure genome-wide significance in GWAS-associated regions. However, using GWAS findings to inform fine-mapping analysis can bias evidence away from the causal SNP toward the tag SNP and SNPs in high LD with the tag. Together these factors can reduce power to localize the causal SNP by more than half. Other strategies commonly employed to increase power to detect association, namely increasing sample size and using higher density genotyping arrays, can, in certain common scenarios, actually exacerbate these effects and further decrease power to localize causal variants. We develop a re-ranking procedure that accounts for these adverse effects and substantially improves the accuracy of causal SNP identification, often doubling the probability that the causal SNP is top-ranked. Application to the NCI BPC3 aggressive prostate cancer GWAS with imputation meta-analysis identified a new top SNP at 2 of 3 associated loci and several additional possible causal SNPs at these loci that may have otherwise been overlooked. This method is simple to implement using R scripts provided on the author's website.

  5. Orientation to Machine Shop. Safety, Machine Identification, Metal Identification.

    ERIC Educational Resources Information Center

    Engelbrecht, Nancy; And Others

    These instructional materials provide an orientation to the machine shop for use at the postsecondary level. The first of three sections discusses four important areas of safety: (1) personal safety; (2) safety procedures; (3) safe work practices; and (4) fire prevention. The second section identifies and describes the general purposes of 12…

  6. Leveraging ancestry to improve causal variant identification in exome sequencing for monogenic disorders.

    PubMed

    Brown, Robert; Lee, Hane; Eskin, Ascia; Kichaev, Gleb; Lohmueller, Kirk E; Reversade, Bruno; Nelson, Stanley F; Pasaniuc, Bogdan

    2016-01-01

    Recent breakthroughs in exome-sequencing technology have made possible the identification of many causal variants of monogenic disorders. Although extremely powerful when closely related individuals (eg, child and parents) are simultaneously sequenced, sequencing of a single case is often unsuccessful due to the large number of variants that need to be followed up for functional validation. Many approaches filter out common variants above a given frequency threshold (eg, 1%), and then prioritize the remaining variants according to their functional, structural and conservation properties. Here we present methods that leverage the genetic structure across different populations to improve filtering performance while accounting for the finite sample size of the reference panels. We show that leveraging genetic structure reduces the number of variants that need to be followed up by 16% in simulations and by up to 38% in empirical data of 20 exomes from individuals with monogenic disorders for which the causal variants are known.

  7. [Identification and characterization of alternativly splicing variants for murine mater gene].

    PubMed

    Cheng, Hui; Zhang, Xiao-Juan; Liu, Hai-Bo; Zhang, Yi; Huang, Zhao-Feng; Ma, Run-Lin

    2004-08-01

    Mater encoding an oocyte-specific autoantigen,and is associated with premature autoimmune ovarian dysgenesis (AOD) in mouse. Based on RT-PCR, cDNA cloning, screening, sequencing and analysis, we have detected a total of four Mater splice variants, designated as variant B, E, F and G. All these splicing forms are in frame in terms of expected protein products. Among these, B was consistent with the previous report, whereas E, F, G belong to novel splice variants that have not been reported previously. Variant E lacks exon 6, variant F both lacks exon 10 and retains a part of intron 8, variant G lacks part of exon 14, and variant H lacks part of exon 13. The cDNA sequences at all the exon-intron boundaries confirms to the "GT-AG" splicing rule. Variant B, E, F exist in all the four strains. Variant G exists only in SWR/J. According to the cDNA sequences of these four splice variants, amimo acid sequences of the corresponding expected protein isoforms were deduced, and their potential functional effects were predicted in this thesis. Further identification and characterization of these expected protein isoforms would provide valuable information for their functional importance.

  8. The identification of mitochondrial DNA variants in glioblastoma multiforme

    PubMed Central

    2014-01-01

    Background Mitochondrial DNA (mtDNA) encodes key proteins of the electron transfer chain (ETC), which produces ATP through oxidative phosphorylation (OXPHOS) and is essential for cells to perform specialised functions. Tumor-initiating cells use aerobic glycolysis, a combination of glycolysis and low levels of OXPHOS, to promote rapid cell proliferation and tumor growth. Glioblastoma multiforme (GBM) is an aggressively malignant brain tumor and mitochondria have been proposed to play a vital role in GBM tumorigenesis. Results Using next generation sequencing and high resolution melt analysis, we identified a large number of mtDNA variants within coding and non-coding regions of GBM cell lines and predicted their disease-causing potential through in silico modeling. The frequency of variants was greatest in the D-loop and origin of light strand replication in non-coding regions. ND6 was the most susceptible coding gene to mutation whilst ND4 had the highest frequency of mutation. Both genes encode subunits of complex I of the ETC. These variants were not detected in unaffected brain samples and many have not been previously reported. Depletion of HSR-GBM1 cells to varying degrees of their mtDNA followed by transplantation into immunedeficient mice resulted in the repopulation of the same variants during tumorigenesis. Likewise, de novo variants identified in other GBM cell lines were also incorporated. Nevertheless, ND4 and ND6 were still the most affected genes. We confirmed the presence of these variants in high grade gliomas. Conclusions These novel variants contribute to GBM by rendering the ETC. partially dysfunctional. This restricts metabolism to anaerobic glycolysis and promotes cell proliferation. PMID:24383468

  9. Identification and characterization of variant alleles at CODIS STR loci.

    PubMed

    Allor, Catherine; Einum, David D; Scarpetta, Marco

    2005-09-01

    Short tandem repeat (STR) profiles from 32,671 individuals generated by the ABI Profiler Plus and Cofiler systems were screened for variant alleles not represented within manufacturer-provided allelic ladders. A total of 85 distinct variants were identified at 12 of the 13 CODIS loci, most of which involve a truncated tetranucleotide repeat unit. Twelve novel alleles, identified at D3S1358, FGA, D18S51, D5S818, D7S820 and TPOX, were confirmed by nucleotide sequence analysis and include both insertions and deletions involving the repeat units themselves as well as DNA flanking the repeat regions. Population genetic data were collected for all variants and frequencies range from 0.0003 (many single observations) to 0.0042 (D7S820 '10.3' in North American Hispanics). In total, the variant alleles identified in this study are carried by 1.6% of the estimated 1 million individuals tested annually in the U.S. for the purposes of parentage resolution. A paternity case involving a recombination event of paternal origin is presented and demonstrates how variant alleles can significantly strengthen the genetic evidence in troublesome cases. In such instances, increased costs and turnaround time associated with additional testing may be eliminated.

  10. Tight control of light beams in photonic crystals with spatially-variant lattice orientation.

    PubMed

    Digaum, Jennefir L; Pazos, Javier J; Chiles, Jeffrey; D'Archangel, Jeffrey; Padilla, Gabriel; Tatulian, Adrian; Rumpf, Raymond C; Fathpour, Sasan; Boreman, Glenn D; Kuebler, Stephen M

    2014-10-20

    Spatially-variant photonic crystals (SVPCs), in which the orientation of the unit cell changes as a function of position, are shown to be capable of abruptly controlling light beams using just low index materials and can be made to have high polarization selectivity. Multi-photon direct laser writing in the photo-polymer SU-8 was used to fabricate three-dimensional SVPCs that direct the flow of light around a 90 degree bend. The lattice spacing and fill factor were maintained nearly constant throughout the structure. The SVPCs were characterized at a wavelength of 2.94 μm by scanning the faces with optical fibers and the results were compared to electromagnetic simulations. The lattices were shown to direct infrared light of one polarization through sharp bends while the other polarization propagated straight through the SVPC. This work introduces a new scheme for controlling light that should be useful for integrated photonics.

  11. Spatially variant tomographic imaging: Estimation, identification, and optimization

    SciTech Connect

    Baker, J.R.

    1991-11-01

    This thesis is an investigation of methods for processing multidimensional signals acquired using modern tomography systems that have an anisotropic or spatially variant response function. The main result of this research is the discovery of a new method to obtain better estimators of an unknown spatial intensity distribution by incorporating detailed knowledge about the tomograph system response function and statistical properties of the acquired signal into a mathematical model.

  12. Crystallographic Texture and Orientation Variants in Al2O3-Y3Al5O12 Directionally Solidified Eutectic Crystals

    NASA Technical Reports Server (NTRS)

    Frazer, Colleen S.; Dickey, Elizabeth C.; Sayir, Ali; Farmer, Serene (Technical Monitor)

    2001-01-01

    Eutectic rods of Al2O3 and Y3Al5O12 were grown by a laser-heated float zone method, and their microstructure and crystallographic texture were studied by scanning electron microscopy, electron backscattered diffraction and x-ray diffraction. The composites were found to be highly textured with two twin-related crystallographic orientation relationships between the phases. Electron backscattered diffraction was employed to determine the spatial distribution of the orientational variants within the samples and to define the crystallographic orientation of various microstructural features.

  13. Identification of a variant-specific phosphorylation of TH2A during spermiogenesis

    PubMed Central

    Hada, Masashi; Masuda, Koji; Yamaguchi, Kosuke; Shirahige, Katsuhiko; Okada, Yuki

    2017-01-01

    Tissue-specific histone variant incorporation into chromatin plays dynamic and important roles in tissue development. Testis is one such tissue, and a number of testis-specific histone variants are expressed that have unique roles. While it is expected that such variants acquire post-transcriptional modifications to be functional, identification of variant-specific histone modifications is challenging because of the high similarity of amino acid sequences between canonical and variant versions. Here we identified a novel phosphorylation on TH2A, a germ cell-specific histone H2A variant. TH2A-Thr127 is unique to the variant and phosphorylated concomitant with chromatin condensation including spermiogenesis and early embryonic mitosis. In sperm chromatin, phosphorylated TH2A-Thr127 (=pTH2A) is co-localized with H3.3 at transcriptional starting sites of the genome, and subsequently becomes absent from the paternal genome upon fertilization. Notably, pTH2A is recurrent and accumulated in the pericentromeric heterochromatin of both paternal and maternal chromosomes in the first mitosis of embryos, suggesting its unique regulation during spermiogenesis and early embryogenesis. PMID:28387373

  14. Orientation selective neural network for cosmic muon identification

    NASA Astrophysics Data System (ADS)

    Abramowicz, Halina; Horn, David; Naftaly, Ury; Sahar-Pikielny, Carmit

    1997-02-01

    We discuss a novel method for identification of a linear pattern of pixels on a two-dimensional grid. Motivated by principles employed by the visual cortex, we construct orientation selective neurons in a neural network that performs this task. The method is then applied to a sample of data collected with the ZEUS detector at HERA in order to identify cosmic muons that leave a linear pattern of signals in the segmented uranium-scintillator calorimeter. A two dimensional representation of the relevant part of the detector is used. The algorithm performs well in the presence of noise and pixels with limited efficiency. Given its architecture, this system becomes a good candidate for fast pattern recognition in parallel processing devices.

  15. Identification of DLK1 variants in pituitary- and neuroendocrine tumors.

    PubMed

    Altenberger, T; Bilban, M; Auer, M; Knosp, E; Wolfsberger, S; Gartner, W; Mineva, I; Zielinski, C; Wagner, L; Luger, A

    2006-02-17

    In a gene chip analysis of common pituitary tumor types, one of the genes with the most impressive tissue-specific expression regulation was delta-like 1 (DLK1), which was strongly expressed in GH-secreting (GH-S) pituitary tumors. In addition to pituitary adenomas, various endocrine tumors were subjected to real-time-quantitative PCR revealing high expression of DLK1 in normal pituitary tissue, in GH-S-, in one prolactin-secreting pituitary adenoma and in pheochromocytomas. Additionally, three DLK1 gene-derived subvariants were identified. The first, lacking 204 bp--coding for epidermal growth factor-like domain 6 and parts of the juxtamembrane region--was named Secredeltin. In the other two splice variants (named Brevideltin and Brevideltinin), a stop codon is introduced due to a frame-shift, leading to truncated proteins of 204 and 213 aas, respectively.

  16. EBSD imaging of orientation relationships and variant groupings in different martensitic alloys and Widmanstätten iron meteorites

    SciTech Connect

    Cayron, Cyril

    2014-08-15

    An automatic method to colorize and quantify the classical Pitsch, Kurdjumov–Sachs, Greninger–Troiano and Nishiyama–Wasserman orientation relationships in the electron backscatter diffraction maps of martensitic/bainitic steels is detailed. Automatic analysis of variant grouping is also presented. The method was applied to low and high carbon steels, and to iron–nickel Widmanstätten meteorites. Many results of recent literature are confirmed. In low carbon steels the individual laths exhibit continuous orientation gradients between the classical orientation relationships, and the laths tend to be grouped by close-packed plane (morphological) packets. A crystallographic scenario describing the formation of the packets is proposed on the base of the one-step model. When the carbon content increases, the orientation spreading is reduced; and martensite tends to form plate groups and burst configurations. In iron–nickel meteorites, the centimeter long Widmanstätten laths do not exhibit continuous orientation gradients but are constituted of subgrains with uniform orientation relationship; the kamacite grains in the plessite regions are grouped into Bain zones, probably due to a recrystallization during the slow cooling of the meteorites. - Highlights: • Analysis of different low and high carbon steels and Widmanstätten meteorites • Automatic color mapping of the classical orientation relationships in EBSD maps • Quantification of variant pairing and grouping tendencies • Crystallographic scenario for the formation of morphological packets.

  17. Global ridge orientation modeling for partial fingerprint identification.

    PubMed

    Wang, Yi Alice; Hu, Jiankun

    2011-01-01

    Identifying incomplete or partial fingerprints from a large fingerprint database remains a difficult challenge today. Existing studies on partial fingerprints focus on one-to-one matching using local ridge details. In this paper, we investigate the problem of retrieving candidate lists for matching partial fingerprints by exploiting global topological features. Specifically, we propose an analytical approach for reconstructing the global topology representation from a partial fingerprint. First, we present an inverse orientation model for describing the reconstruction problem. Then, we provide a general expression for all valid solutions to the inverse model. This allows us to preserve data fidelity in the existing segments while exploring missing structures in the unknown parts. We have further developed algorithms for estimating the missing orientation structures based on some a priori knowledge of ridge topology features. Our statistical experiments show that our proposed model-based approach can effectively reduce the number of candidates for pair-wised fingerprint matching, and thus significantly improve the system retrieval performance for partial fingerprint identification.

  18. Identification of hemoglobin variants by top-down mass spectrometry using selected diagnostic product ions.

    PubMed

    Coelho Graça, Didia; Hartmer, Ralf; Jabs, Wolfgang; Beris, Photis; Clerici, Lorella; Stoermer, Carsten; Samii, Kaveh; Hochstrasser, Denis; Tsybin, Yury O; Scherl, Alexander; Lescuyer, Pierre

    2015-04-01

    Hemoglobin disorder diagnosis is a complex procedure combining several analytical steps. Due to the lack of specificity of the currently used protein analysis methods, the identification of uncommon hemoglobin variants (proteoforms) can become a hard task to accomplish. The aim of this work was to develop a mass spectrometry-based approach to quickly identify mutated protein sequences within globin chain variants. To reach this goal, a top-down electron transfer dissociation mass spectrometry method was developed for hemoglobin β chain analysis. A diagnostic product ion list was established with a color code strategy allowing to quickly and specifically localize a mutation in the hemoglobin β chain sequence. The method was applied to the analysis of rare hemoglobin β chain variants and an (A)γ-β fusion protein. The results showed that the developed data analysis process allows fast and reliable interpretation of top-down electron transfer dissociation mass spectra by nonexpert users in the clinical area.

  19. Resequencing Pathogen Microarray (RPM) for prospective detection and identification of emergent pathogen strains and variants

    NASA Astrophysics Data System (ADS)

    Tibbetts, Clark; Lichanska, Agnieszka M.; Borsuk, Lisa A.; Weslowski, Brian; Morris, Leah M.; Lorence, Matthew C.; Schafer, Klaus O.; Campos, Joseph; Sene, Mohamadou; Myers, Christopher A.; Faix, Dennis; Blair, Patrick J.; Brown, Jason; Metzgar, David

    2010-04-01

    High-density resequencing microarrays support simultaneous detection and identification of multiple viral and bacterial pathogens. Because detection and identification using RPM is based upon multiple specimen-specific target pathogen gene sequences generated in the individual test, the test results enable both a differential diagnostic analysis and epidemiological tracking of detected pathogen strains and variants from one specimen to the next. The RPM assay enables detection and identification of pathogen sequences that share as little as 80% sequence similarity to prototype target gene sequences represented as detector tiles on the array. This capability enables the RPM to detect and identify previously unknown strains and variants of a detected pathogen, as in sentinel cases associated with an infectious disease outbreak. We illustrate this capability using assay results from testing influenza A virus vaccines configured with strains that were first defined years after the design of the RPM microarray. Results are also presented from RPM-Flu testing of three specimens independently confirmed to the positive for the 2009 Novel H1N1 outbreak strain of influenza virus.

  20. Incomplete data based parameter identification of nonlinear and time-variant oscillators with fractional derivative elements

    NASA Astrophysics Data System (ADS)

    Kougioumtzoglou, Ioannis A.; dos Santos, Ketson R. M.; Comerford, Liam

    2017-09-01

    Various system identification techniques exist in the literature that can handle non-stationary measured time-histories, or cases of incomplete data, or address systems following a fractional calculus modeling. However, there are not many (if any) techniques that can address all three aforementioned challenges simultaneously in a consistent manner. In this paper, a novel multiple-input/single-output (MISO) system identification technique is developed for parameter identification of nonlinear and time-variant oscillators with fractional derivative terms subject to incomplete non-stationary data. The technique utilizes a representation of the nonlinear restoring forces as a set of parallel linear sub-systems. In this regard, the oscillator is transformed into an equivalent MISO system in the wavelet domain. Next, a recently developed L1-norm minimization procedure based on compressive sensing theory is applied for determining the wavelet coefficients of the available incomplete non-stationary input-output (excitation-response) data. Finally, these wavelet coefficients are utilized to determine appropriately defined time- and frequency-dependent wavelet based frequency response functions and related oscillator parameters. Several linear and nonlinear time-variant systems with fractional derivative elements are used as numerical examples to demonstrate the reliability of the technique even in cases of noise corrupted and incomplete data.

  1. Hermetic Narratives and False Analysis: A Unique Variant of the Mechanism of Identification With the Aggressor.

    PubMed

    Amir, Dana

    2016-08-01

    This paper focuses on a unique variant of the mechanism of identification with the aggressor. The term "hermetic narrative" refers to a condition in which the trauma victim creates a hermetically sealed narrative of witnessing, which becomes an addictive and subjugating object in itself, while obstructing natural processes of thinking. This paper examines the ways in which the hermetic narrative reconstructs victim-aggressor relations both within the individual and in the analytic relationship. It further discusses the risk of creating a "false analysis" that is based on a malignant cooperation of the analyst and patient with the internalized traumatic object.

  2. A new system identification approach to identify genetic variants in sequencing studies for a binary phenotype.

    PubMed

    Kang, Guolian; Bi, Wenjian; Zhao, Yanlong; Zhang, Ji-Feng; Yang, Jun J; Xu, Heng; Loh, Mignon L; Hunger, Stephen P; Relling, Mary V; Pounds, Stanley; Cheng, Cheng

    2014-01-01

    We propose in this paper a set-valued (SV) system model, which is a generalized form of logistic (LG) and Probit (Probit) regression, to be considered as a method for discovering genetic variants, especially rare genetic variants in next-generation sequencing studies, for a binary phenotype. We propose a new SV system identification method to estimate all underlying key system parameters for the Probit model and compare it with the LG model in the setting of genetic association studies. Across an extensive series of simulation studies, the Probit method maintained type I error control and had similar or greater power than the LG method, which is robust to different distributions of noise: logistic, normal, or t distributions. Additionally, the Probit association parameter estimate was 2.7-46.8-fold less variable than the LG log-odds ratio association parameter estimate. Less variability in the association parameter estimate translates to greater power and robustness across the spectrum of minor allele frequencies (MAFs), and these advantages are the most pronounced for rare variants. For instance, in a simulation that generated data from an additive logistic model with an odds ratio of 7.4 for a rare single nucleotide polymorphism with a MAF of 0.005 and a sample size of 2,300, the Probit method had 60% power whereas the LG method had 25% power at the α = 10(-6) level. Consistent with these simulation results, the set of variants identified by the LG method was a subset of those identified by the Probit method in two example analyses. Thus, we suggest the Probit method may be a competitive alternative to the LG method in genetic association studies such as candidate gene, genome-wide, or next-generation sequencing studies for a binary phenotype.

  3. Identification of lung cancer histology-specific variants applying Bayesian framework variant prioritization approaches within the TRICL and ILCCO consortia

    PubMed Central

    Brenner, Darren R.; Amos, Christopher I.; Brhane, Yonathan; Timofeeva, Maria N.; Caporaso, Neil; Wang, Yufei; Christiani, David C.; Bickeböller, Heike; Yang, Ping; Albanes, Demetrius; Stevens, Victoria L.; Gapstur, Susan; McKay, James; Boffetta, Paolo; Zaridze, David; Szeszenia-Dabrowska, Neonilia; Lissowska, Jolanta; Rudnai, Peter; Fabianova, Eleonora; Mates, Dana; Bencko, Vladimir; Foretova, Lenka; Janout, Vladimir; Krokan, Hans E.; Skorpen, Frank; Gabrielsen, Maiken E.; Vatten, Lars; Njølstad, Inger; Chen, Chu; Goodman, Gary; Lathrop, Mark; Vooder, Tõnu; Välk, Kristjan; Nelis, Mari; Metspalu, Andres; Broderick, Peter; Eisen, Timothy; Wu, Xifeng; Zhang, Di; Chen, Wei; Spitz, Margaret R.; Wei, Yongyue; Su, Li; Xie, Dong; She, Jun; Matsuo, Keitaro; Matsuda, Fumihiko; Ito, Hidemi; Risch, Angela; Heinrich, Joachim; Rosenberger, Albert; Muley, Thomas; Dienemann, Hendrik; Field, John K.; Raji, Olaide; Chen, Ying; Gosney, John; Liloglou, Triantafillos; Davies, Michael P.A.; Marcus, Michael; McLaughlin, John; Orlow, Irene; Han, Younghun; Li, Yafang; Zong, Xuchen; Johansson, Mattias; Liu, Geoffrey; Tworoger, Shelley S.; Le Marchand, Loic; Henderson, Brian E.; Wilkens, Lynne R.; Dai, Juncheng; Shen, Hongbing; Houlston, Richard S.; Landi, Maria T.; Brennan, Paul; Hung, Rayjean J.

    2015-01-01

    Large-scale genome-wide association studies (GWAS) have likely uncovered all common variants at the GWAS significance level. Additional variants within the suggestive range (0.0001> P > 5×10−8) are, however, still of interest for identifying causal associations. This analysis aimed to apply novel variant prioritization approaches to identify additional lung cancer variants that may not reach the GWAS level. Effects were combined across studies with a total of 33456 controls and 6756 adenocarcinoma (AC; 13 studies), 5061 squamous cell carcinoma (SCC; 12 studies) and 2216 small cell lung cancer cases (9 studies). Based on prior information such as variant physical properties and functional significance, we applied stratified false discovery rates, hierarchical modeling and Bayesian false discovery probabilities for variant prioritization. We conducted a fine mapping analysis as validation of our methods by examining top-ranking novel variants in six independent populations with a total of 3128 cases and 2966 controls. Three novel loci in the suggestive range were identified based on our Bayesian framework analyses: KCNIP4 at 4p15.2 (rs6448050, P = 4.6×10−7) and MTMR2 at 11q21 (rs10501831, P = 3.1×10−6) with SCC, as well as GAREM at 18q12.1 (rs11662168, P = 3.4×10−7) with AC. Use of our prioritization methods validated two of the top three loci associated with SCC (P = 1.05×10−4 for KCNIP4, represented by rs9799795) and AC (P = 2.16×10−4 for GAREM, represented by rs3786309) in the independent fine mapping populations. This study highlights the utility of using prior functional data for sequence variants in prioritization analyses to search for robust signals in the suggestive range. PMID:26363033

  4. Functional classification of DNA variants by hybrid minigenes: Identification of 30 spliceogenic variants of BRCA2 exons 17 and 18

    PubMed Central

    Fraile-Bethencourt, Eugenia; Díez-Gómez, Beatriz; Acedo, Alberto; Sanz, David J.

    2017-01-01

    Mutation screening of the breast cancer genes BRCA1 and BRCA2 identifies a large fraction of variants of uncertain clinical significance (VUS) whose functional and clinical interpretations pose a challenge for genomic medicine. Likewise, an increasing amount of evidence indicates that genetic variants can have deleterious effects on pre-mRNA splicing. Our goal was to investigate the impact on splicing of a set of reported variants of BRCA2 exons 17 and 18 to assess their role in hereditary breast cancer and to identify critical regulatory elements that may constitute hotspots for spliceogenic variants. A splicing reporter minigene with BRCA2 exons 14 to-20 (MGBR2_ex14-20) was constructed in the pSAD vector. Fifty-two candidate variants were selected with splicing prediction programs, introduced in MGBR2_ex14-20 by site-directed mutagenesis and assayed in triplicate in MCF-7 cells. Wild type MGBR2_ex14-20 produced a stable transcript of the expected size (1,806 nucleotides) and structure (V1-[BRCA2_exons_14–20]–V2). Functional mapping by microdeletions revealed essential sequences for exon recognition on the 3’ end of exon 17 (c.7944-7973) and the 5’ end of exon 18 (c.7979-7988, c.7999-8013). Thirty out of the 52 selected variants induced anomalous splicing in minigene assays with >16 different aberrant transcripts, where exon skipping was the most common event. A wide range of splicing motifs were affected including the canonical splice sites (15 variants), novel alternative sites (3 variants), the polypyrimidine tract (3 variants) and enhancers/silencers (9 variants). According to the guidelines of the American College of Medical Genetics and Genomics (ACMG), 20 variants could be classified as pathogenic (c.7806-2A>G, c.7806-1G>A, c.7806-1G>T, c.7806-1_7806-2dup, c.7976+1G>A, c.7977-3_7978del, c.7977-2A>T, c.7977-1G>T, c.7977-1G>C, c.8009C>A, c.8331+1G>T and c.8331+2T>C) or likely pathogenic (c.7806-9T>G, c.7976G>C, c.7976G>A, c.7977-7C>G, c.7985C>G, c

  5. Multi-species Identification of Polymorphic Peptide Variants via Propagation in Spectral Networks

    SciTech Connect

    Na, Seungjin; Payne, Samuel H.; Bandeira, Nuno

    2016-09-08

    The spectral networks approach enables the detection of pairs of spectra from related peptides and thus allows for the propagation of annotations from identified peptides to unidentified spectra. Beyond allowing for unbiased discovery of unexpected post-translational modifications, spectral networks are also applicable to multi-species comparative proteomics or metaproteomics to identify numerous orthologous versions of a protein. We present algorithmic and statistical advances in spectral networks that have made it possible to rigorously assess the statistical significance of spectral pairs and accurately estimate the error rate of identifications via propagation. In the analysis of three related Cyanothece species, a model organism for biohydrogen production, spectral networks identified peptides with highly divergent sequences with up to dozens of variants per peptide, including many novel peptides in species that lack a sequenced genome. Furthermore, spectral networks strongly suggested the presence of novel peptides even in genomically characterized species (i.e. missing from databases) in that a significant portion of unidentified multi-species networks included at least two polymorphic peptide variants.

  6. Identification of a variant in KDR associated with serum VEGFR2 and pharmacodynamics of Pazopanib.

    PubMed

    Maitland, Michael L; Xu, Chun-Fang; Cheng, Yu-Ching; Kistner-Griffin, Emily; Ryan, Kathleen A; Karrison, Theodore G; Das, Soma; Torgerson, Dara; Gamazon, Eric R; Thomeas, Vasiliki; Levine, Matthew R; Wilson, Paul A; Bing, Nan; Liu, Yuan; Cardon, Lon R; Pandite, Lini N; O'Connell, Jeffrey R; Cox, Nancy J; Mitchell, Braxton D; Ratain, Mark J; Shuldiner, Alan R

    2015-01-15

    VEGF receptor (VEGFR) kinases are important drug targets in oncology that affect function of systemic endothelial cells. To discover genetic markers that affect VEGFR inhibitor pharmacodynamics, we performed a genome-wide association study of serum soluble vascular VEGFR2 concentrations [sVEGFR2], a pharmacodynamic biomarker for VEGFR2 inhibitors. We conducted a genome-wide association study (GWAS) of [sVEGFR2] in 736 healthy Old Order Amish volunteers. Gene variants identified from the GWAS were genotyped serially in a cohort of 128 patients with advanced solid tumor with baseline [sVEGFR2] measurements, and in 121 patients with renal carcinoma with [sVEGFR2] measured before and during pazopanib therapy. rs34231037 (C482R) in KDR, the gene encoding sVEGFR2 was found to be highly associated with [sVEGFR2], explaining 23% of the variance (P = 2.7 × 10(-37)). Association of rs34231037 with [sVEGFR2] was replicated in 128 patients with cancer with comparable effect size (P = 0.025). Furthermore, rs34231037 was a significant predictor of changes in [sVEGFR2] in response to pazopanib (P = 0.01). Our findings suggest that genome-wide analysis of phenotypes in healthy populations can expedite identification of candidate pharmacogenetic markers. Genotyping for germline variants in KDR may have clinical utility in identifying patients with cancer with unusual sensitivity to effects of VEGFR2 kinase inhibitors. ©2014 American Association for Cancer Research.

  7. Identification of genetic variants associated with alternative splicing using sQTLseekeR

    PubMed Central

    Monlong, Jean; Calvo, Miquel; Ferreira, Pedro G.; Guigó, Roderic

    2014-01-01

    Identification of genetic variants affecting splicing in RNA sequencing population studies is still in its infancy. Splicing phenotype is more complex than gene expression and ought to be treated as a multivariate phenotype to be recapitulated completely. Here we represent the splicing pattern of a gene as the distribution of the relative abundances of a gene’s alternative transcript isoforms. We develop a statistical framework that uses a distance-based approach to compute the variability of splicing ratios across observations, and a non-parametric analogue to multivariate analysis of variance. We implement this approach in the R package sQTLseekeR and use it to analyze RNA-Seq data from the Geuvadis project in 465 individuals. We identify hundreds of single nucleotide polymorphisms (SNPs) as splicing QTLs (sQTLs), including some falling in genome-wide association study SNPs. By developing the appropriate metrics, we show that sQTLseekeR compares favorably with existing methods that rely on univariate approaches, predicting variants that behave as expected from mutations affecting splicing. PMID:25140736

  8. Panel sequencing for clinically oriented variant screening and copy number detection in 142 untreated multiple myeloma patients

    PubMed Central

    Kortuem, K M; Braggio, E; Bruins, L; Barrio, S; Shi, C S; Zhu, Y X; Tibes, R; Viswanatha, D; Votruba, P; Ahmann, G; Fonseca, R; Jedlowski, P; Schlam, I; Kumar, S; Bergsagel, P L; Stewart, A K

    2016-01-01

    We employed a customized Multiple Myeloma (MM)-specific Mutation Panel (M3P) to screen a homogenous cohort of 142 untreated MM patients for relevant mutations in a selection of disease-specific genes. M3Pv2.0 includes 77 genes selected for being either actionable targets, potentially related to drug–response or part of known key pathways in MM biology. We identified mutations in potentially actionable genes in 49% of patients and provided prognostic evidence of STAT3 mutations. This panel may serve as a practical alternative to more comprehensive sequencing approaches, providing genomic information in a timely and cost-effective manner, thus allowing clinically oriented variant screening in MM. PMID:26918361

  9. Identification and characterization of seven new exon 11-associated splice variants of the rat μ opioid receptor gene, OPRM1.

    PubMed

    Xu, Jin; Xu, Mingming; Rossi, Grace C; Pasternak, Gavril W; Pan, Ying-Xian

    2011-01-21

    The mouse mu opioid receptor (OPRM1) gene undergoes extensive alternative splicing at both the 3'- and 5'-ends of the gene. Previously, several C-terminal variants generated through 3' splicing have been identified in the rat OPRM1 gene. In both mice and humans 5' splicing generates a number of exon 11-containing variants. Studies in an exon 11 knockout mouse suggest the functional importance of these exon 11-associated variants in mediating the analgesic actions of a subset of mu opioids, including morphine-6β-glucuronide (M6G) and heroin, but not others such as morphine and methadone. We now have examined 5' splicing in the rat. The current studies identified in the rat a homologous exon 11 and seven exon 11-associated variants, suggesting conservation of exon 11 and its associated variants among mouse, rat and human. RT-PCR revealed marked differences in the expression of these variants across several brain regions, implying region-specific mRNA processing of the exon 11-associated variants. Of the seven rat exon 11-associated variants, four encoded the identical protein as found in rMOR-1, two predicted 6 TM variants, and one, rMOR-1H2, generated a novel N-terminal variant in which a stretch of an additional 50 amino acids was present at the N-terminus of the previously established rMOR-1 sequence. When expressed in CHO cells, the presence of the additional 50 amino acids in rMOR-1H2 significantly altered agonist-induced G protein activation with little effect on opioid binding. The identification of the rat exon 11 and its associated variants further demonstrated conservation of 5' splicing in OPRM1 genes among rodents and humans. The functional relevance of these exon 11 associated variants was suggested by the region-specific expression of their mRNAs and the influence of the N-terminal sequence on agonist-induced G protein coupling in the novel N-terminal variant, rMOR-1H2. The importance of the exon 11-associated variants in mice in M6G and heroin

  10. Evaluating alignment and variant-calling software for mutation identification in C. elegans by whole-genome sequencing

    PubMed Central

    Yun, Sijung

    2017-01-01

    Whole-genome sequencing is a powerful tool for analyzing genetic variation on a global scale. One particularly useful application is the identification of mutations obtained by classical phenotypic screens in model species. Sequence data from the mutant strain is aligned to the reference genome, and then variants are called to generate a list of candidate alleles. A number of software pipelines for mutation identification have been targeted to C. elegans, with particular emphasis on ease of use, incorporation of mapping strain data, subtraction of background variants, and similar criteria. Although success is predicated upon the sensitive and accurate detection of candidate alleles, relatively little effort has been invested in evaluating the underlying software components that are required for mutation identification. Therefore, we have benchmarked a number of commonly used tools for sequence alignment and variant calling, in all pair-wise combinations, against both simulated and actual datasets. We compared the accuracy of those pipelines for mutation identification in C. elegans, and found that the combination of BBMap for alignment plus FreeBayes for variant calling offers the most robust performance. PMID:28333980

  11. Evaluating alignment and variant-calling software for mutation identification in C. elegans by whole-genome sequencing.

    PubMed

    Smith, Harold E; Yun, Sijung

    2017-01-01

    Whole-genome sequencing is a powerful tool for analyzing genetic variation on a global scale. One particularly useful application is the identification of mutations obtained by classical phenotypic screens in model species. Sequence data from the mutant strain is aligned to the reference genome, and then variants are called to generate a list of candidate alleles. A number of software pipelines for mutation identification have been targeted to C. elegans, with particular emphasis on ease of use, incorporation of mapping strain data, subtraction of background variants, and similar criteria. Although success is predicated upon the sensitive and accurate detection of candidate alleles, relatively little effort has been invested in evaluating the underlying software components that are required for mutation identification. Therefore, we have benchmarked a number of commonly used tools for sequence alignment and variant calling, in all pair-wise combinations, against both simulated and actual datasets. We compared the accuracy of those pipelines for mutation identification in C. elegans, and found that the combination of BBMap for alignment plus FreeBayes for variant calling offers the most robust performance.

  12. Multi-species Identification of Polymorphic Peptide Variants via Propagation in Spectral Networks*

    PubMed Central

    Bandeira, Nuno

    2016-01-01

    Peptide and protein identification remains challenging in organisms with poorly annotated or rapidly evolving genomes, as are commonly encountered in environmental or biofuels research. Such limitations render tandem mass spectrometry (MS/MS) database search algorithms ineffective as they lack corresponding sequences required for peptide-spectrum matching. We address this challenge with the spectral networks approach to (1) match spectra of orthologous peptides across multiple related species and then (2) propagate peptide annotations from identified to unidentified spectra. We here present algorithms to assess the statistical significance of spectral alignments (Align-GF), reduce the impurity in spectral networks, and accurately estimate the error rate in propagated identifications. Analyzing three related Cyanothece species, a model organism for biohydrogen production, spectral networks identified peptides from highly divergent sequences from networks with dozens of variant peptides, including thousands of peptides in species lacking a sequenced genome. Our analysis further detected the presence of many novel putative peptides even in genomically characterized species, thus suggesting the possibility of gaps in our understanding of their proteomic and genomic expression. A web-based pipeline for spectral networks analysis is available at http://proteomics.ucsd.edu/software. PMID:27609420

  13. Identification of six new RHCE variant alleles in individuals of diverse racial origin.

    PubMed

    Goldman, Mindy; Cemborain, Arantxa; Cote, Jacqueline; El Hamss, Rachid; Flower, Robert L; Garaizar, Adirane; Garcia-Sanchez, Felix; Hyland, Catherine A; Kalvelage, Monica; Londero, Donatella; Lopez, Genghis H; Revelli, Nicoletta; Rodriguez-Wilhelmi, Pablo; Villa, Antonietta; Ochoa-Garay, Gorka

    2016-01-01

    The introduction of molecular methods into routine blood typing is prompting the identification of new blood group alleles. Discrepancies between the results of genotyping and serology or chance events uncovered during genotyping prompted additional investigations, which revealed six new RHCE variant alleles. Samples from eight blood donors, two patients (one prenatal), and a patient's relative, all of diverse racial origin, were analyzed by standard serology methods, targeted genotyping arrays, DNA sequencing, and allele-specific polymerase chain reaction. Six new RHCE alleles were identified, namely, RHCE*cE84A, RHCE*ce202G, RHCE*ce307T, RHCE*Ce377G, RHCE*ce697G,712G,733G,744C, and RHCE*Ce733G. While implementation of new assays in commercial genotyping platforms to detect the polymorphisms reported here may not be justified given their apparent rarity, software interpretative algorithms may benefit from the identification of new alleles for a more accurate determination of genotypes and prediction of phenotypes. © 2015 AABB.

  14. Emotional and neutral scenes in competition: orienting, efficiency, and identification.

    PubMed

    Calvo, Manuel G; Nummenmaa, Lauri; Hyönä, Jukka

    2007-12-01

    To investigate preferential processing of emotional scenes competing for limited attentional resources with neutral scenes, prime pictures were presented briefly (450 ms), peripherally (5.2 degrees away from fixation), and simultaneously (one emotional and one neutral scene) versus singly. Primes were followed by a mask and a probe for recognition. Hit rate was higher for emotional than for neutral scenes in the dual- but not in the single-prime condition, and A' sensitivity decreased for neutral but not for emotional scenes in the dual-prime condition. This preferential processing involved both selective orienting and efficient encoding, as revealed, respectively, by a higher probability of first fixation on--and shorter saccade latencies to--emotional scenes and by shorter fixation time needed to accurately identify emotional scenes, in comparison with neutral scenes.

  15. The Socializing Function of New Employee Orientation Programs: A Study of Organizational Identification and Job Satisfaction.

    ERIC Educational Resources Information Center

    Gates, Lisa R.; Hellweg, Susan A.

    This study examined one form of organizational socialization, the new employee orientation program, to determine whether such organizational efforts would increase levels of organizational identification and employee perceptions of job satisfaction when employees underwent such a program. An alternative model was tested to ascertain whether or not…

  16. Skeletal idiopathic osteosclerosis helps to perform personal identification of unknown decedents: A novel contribution from anatomical variants through CT scan.

    PubMed

    De Angelis, D; Gibelli, D; Palazzo, E; Sconfienza, L; Obertova, Z; Cattaneo, C

    2016-07-01

    Personal identification consists of the comparison of ante-mortem information from a missing person with post-mortem data obtained from an unidentified corpse. Such procedure is based on the assessment of individualizing features which may help in providing a conclusive identification between ante-mortem and post-mortem material. Anatomical variants may provide important clues to correctly identify human remains. Areas of idiopathic osteosclerosis (IO), or dense bone islands (DBIs) characterized by radiopaque areas of dense, trabeculated, non-inflamed vital bone represent one of these, potentially individualizing, anatomical features. This study presents a case where the finding of DBI was crucial for a positive identification through CT-scan. A decomposed body was found in an apartment in June 2014 in advanced decomposition and no dental records were available to perform a comparison for positive identification. Genetic tests were not applicable because of the lack of relatives in a direct line. The analysis of the only ante-mortem documentation, a CT-scan to the deceased dating back to August 2009, showed the presence of three DBIs within the trabecular bone of the proximal portion of the right femur. The same bony district was removed from the corpse during the autopsy and analysed by CT-scan, which verified the presence of the same features. Forensic practitioners should therefore be aware of the great importance of anatomical bone variants, such as dense bone islands for identification purposes, and the importance of advanced radiological technique for addressing the individualizing potential of such variants. We propose that anatomical variants of the human skeleton should be considered as being "primary identification characteristics" similar to dental status, fingerprints and DNA.

  17. Identification of interleukin-26 in the dromedary camel (Camelus dromedarius): Evidence of alternative splicing and isolation of novel splice variants.

    PubMed

    Premraj, Avinash; Nautiyal, Binita; Aleyas, Abi G; Rasool, Thaha Jamal

    2015-10-01

    Interleukin-26 (IL-26) is a member of the IL-10 family of cytokines. Though conserved across vertebrates, the IL-26 gene is functionally inactivated in a few mammals like rat, mouse and horse. We report here the identification, isolation and cloning of the cDNA of IL-26 from the dromedary camel. The camel cDNA contains a 516 bp open reading frame encoding a 171 amino acid precursor protein, including a 21 amino acid signal peptide. Sequence analysis revealed high similarity with other mammalian IL-26 homologs and the conservation of IL-10 cytokine family domain structure including key amino acid residues. We also report the identification and cloning of four novel transcript variants produced by alternative splicing at the Exon 3-Exon 4 regions of the gene. Three of the alternative splice variants had premature termination codons and are predicted to code for truncated proteins. The transcript variant 4 (Tv4) having an insertion of an extra 120 bp nucleotides in the ORF was predicted to encode a full length protein product with 40 extra amino acid residues. The mRNA transcripts of all the variants were identified in lymph node, where as fewer variants were observed in other tissues like blood, liver and kidney. The expression of Tv2 and Tv3 were found to be up regulated in mitogen induced camel peripheral blood mononuclear cells. IL-26-Tv2 expression was also induced in camel fibroblast cells infected with Camel pox virus in-vitro. The identification of the transcript variants of IL-26 from the dromedary camel is the first report of alternative splicing for IL-26 in a species in which the gene has not been inactivated. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. Identification of copy number variants in whole-genome data using Reference Coverage Profiles

    PubMed Central

    Glusman, Gustavo; Severson, Alissa; Dhankani, Varsha; Robinson, Max; Farrah, Terry; Mauldin, Denise E.; Stittrich, Anna B.; Ament, Seth A.; Roach, Jared C.; Brunkow, Mary E.; Bodian, Dale L.; Vockley, Joseph G.; Shmulevich, Ilya; Niederhuber, John E.; Hood, Leroy

    2015-01-01

    The identification of DNA copy numbers from short-read sequencing data remains a challenge for both technical and algorithmic reasons. The raw data for these analyses are measured in tens to hundreds of gigabytes per genome; transmitting, storing, and analyzing such large files is cumbersome, particularly for methods that analyze several samples simultaneously. We developed a very efficient representation of depth of coverage (150–1000× compression) that enables such analyses. Current methods for analyzing variants in whole-genome sequencing (WGS) data frequently miss copy number variants (CNVs), particularly hemizygous deletions in the 1–100 kb range. To fill this gap, we developed a method to identify CNVs in individual genomes, based on comparison to joint profiles pre-computed from a large set of genomes. We analyzed depth of coverage in over 6000 high quality (>40×) genomes. The depth of coverage has strong sequence-specific fluctuations only partially explained by global parameters like %GC. To account for these fluctuations, we constructed multi-genome profiles representing the observed or inferred diploid depth of coverage at each position along the genome. These Reference Coverage Profiles (RCPs) take into account the diverse technologies and pipeline versions used. Normalization of the scaled coverage to the RCP followed by hidden Markov model (HMM) segmentation enables efficient detection of CNVs and large deletions in individual genomes. Use of pre-computed multi-genome coverage profiles improves our ability to analyze each individual genome. We make available RCPs and tools for performing these analyses on personal genomes. We expect the increased sensitivity and specificity for individual genome analysis to be critical for achieving clinical-grade genome interpretation. PMID:25741365

  19. Identification of copy number variants in whole-genome data using Reference Coverage Profiles.

    PubMed

    Glusman, Gustavo; Severson, Alissa; Dhankani, Varsha; Robinson, Max; Farrah, Terry; Mauldin, Denise E; Stittrich, Anna B; Ament, Seth A; Roach, Jared C; Brunkow, Mary E; Bodian, Dale L; Vockley, Joseph G; Shmulevich, Ilya; Niederhuber, John E; Hood, Leroy

    2015-01-01

    The identification of DNA copy numbers from short-read sequencing data remains a challenge for both technical and algorithmic reasons. The raw data for these analyses are measured in tens to hundreds of gigabytes per genome; transmitting, storing, and analyzing such large files is cumbersome, particularly for methods that analyze several samples simultaneously. We developed a very efficient representation of depth of coverage (150-1000× compression) that enables such analyses. Current methods for analyzing variants in whole-genome sequencing (WGS) data frequently miss copy number variants (CNVs), particularly hemizygous deletions in the 1-100 kb range. To fill this gap, we developed a method to identify CNVs in individual genomes, based on comparison to joint profiles pre-computed from a large set of genomes. We analyzed depth of coverage in over 6000 high quality (>40×) genomes. The depth of coverage has strong sequence-specific fluctuations only partially explained by global parameters like %GC. To account for these fluctuations, we constructed multi-genome profiles representing the observed or inferred diploid depth of coverage at each position along the genome. These Reference Coverage Profiles (RCPs) take into account the diverse technologies and pipeline versions used. Normalization of the scaled coverage to the RCP followed by hidden Markov model (HMM) segmentation enables efficient detection of CNVs and large deletions in individual genomes. Use of pre-computed multi-genome coverage profiles improves our ability to analyze each individual genome. We make available RCPs and tools for performing these analyses on personal genomes. We expect the increased sensitivity and specificity for individual genome analysis to be critical for achieving clinical-grade genome interpretation.

  20. Identification of a variant in KDR associated with serum VEGFR2 and pharmacodynamics of pazopanib

    PubMed Central

    Maitland, Michael L.; Xu, Chun-Fang; Cheng, Yu-Ching; Kistner-Griffin, Emily; Ryan, Kathleen A.; Karrison, Theodore G.; Das, Soma; Torgerson, Dara; Gamazon, Eric; Thomeas, Vasiliki; Levine, Matthew R.; Wilson, Paul A.; Bing, Nan; Liu, Yuan; Cardon, Lon R.; Pandite, Lini N.; O’Connell, Jeffrey R.; Cox, Nancy J.; Mitchell, Braxton D.; Ratain, Mark J.; Shuldiner, Alan R.

    2014-01-01

    Purpose Vascular endothelial growth factor receptor (VEGFR) kinases are important drug targets in oncology that affect function of systemic endothelial cells. To discover genetic markers that affect VEGFR inhibitor pharmacodynamics we performed a genome-wide association study of serum soluble vascular endothelial growth factor receptor-2 concentrations [sVEGFR2], a pharmacodynamic biomarker for VEGFR2 inhibitors. Experimental Design We conducted a genome-wide association study (GWAS) of [sVEGFR2] in 736 healthy Old Order Amish volunteers. Gene variants identified from the GWAS were genotyped serially in a cohort of 128 advanced solid tumor patients with baseline [sVEGR2] measurements, and in 121 renal carcinoma patients with [sVEGFR2] measured before and during pazopanib therapy. Results rs34231037 (C482R) in KDR, the gene encoding sVEGFR2 was found to be highly associated with [sVEGFR2], explaining 23% of the variance (p=2.7×10−37). Association of rs34231037 with [sVEGFR2] was replicated in 128 patients with cancer with comparable effect size (p = 0.025). Furthermore rs34231037 was a significant predictor of changes in [sVEGFR2] in response to pazopanib (p = 0.01). Conclusion Our findings suggest that genome-wide analysis of phenotypes in healthy populations can expedite identification of candidate pharmacogenetic markers. Genotyping for germ-line variants in KDR may have clinical utility in identifying cancer patients with unusual sensitivity to effects of VEGFR2 kinase inhibitors. PMID:25411163

  1. Brief Report: Whole-Exome Sequencing for Identification of Potential Causal Variants for Diffuse Cutaneous Systemic Sclerosis.

    PubMed

    Mak, Angel C Y; Tang, Paul L F; Cleveland, Clare; Smith, Melanie H; Kari Connolly, M; Katsumoto, Tamiko R; Wolters, Paul J; Kwok, Pui-Yan; Criswell, Lindsey A

    2016-09-01

    Scleroderma is a genetically complex autoimmune disease with substantial phenotypic heterogeneity. Previous genome-wide association studies have identified common genetic variants associated with disease risk, but these studies are not designed to capture rare or potential causal variants. Our goal was to identify rare as well as common genetic variants in patients with diffuse cutaneous systemic sclerosis (dcSSc) through whole-exome sequencing (WES) in order to identify potential causal variants. We generated WES data for 32 dcSSc patients with or without interstitial lung disease (ILD) and for 17 healthy "in-house" controls. Variants were annotated and filtered by quality, minor allele frequency, and deleterious effects on gene function. We applied a gene burden test to identify novel dcSSc and dcSSc-associated ILD candidate genes that were enriched with deleterious variants in cases compared to in-house controls as well as controls from the 1000 Genomes Project (n = 130). We identified 70 genes that were enriched with deleterious variants in dcSSc patients. Two of them (BANK1 and TERT) were in pathways previously implicated in SSc or ILD pathogenesis or known susceptibility loci. Newly identified genes (COL4A3, COL4A4, COL5A2, COL13A1, and COL22A1) were significantly enriched in the extracellular matrix-related pathway, which is relevant to the fibrotic features of dcSSc, and in the DNA repair pathway (XRCC4). This study demonstrates the value of WES for the identification of novel gene variants and pathways that may contribute to scleroderma risk and/or severity. The candidate genes we discovered are potential targets for in-depth functional studies. © 2016, American College of Rheumatology.

  2. Team-oriented leadership: the interactive effects of leader group prototypicality, accountability, and team identification.

    PubMed

    Giessner, Steffen R; van Knippenberg, Daan; van Ginkel, Wendy; Sleebos, Ed

    2013-07-01

    We examined the interactive effects of leader group prototypicality, accountability, and team identification on team-oriented behavior of leaders, thus extending the social identity perspective on leadership to the study of leader behavior. An experimental study (N = 152) supported our hypothesis that leader accountability relates more strongly to team-oriented behavior for group nonprototypical leaders than for group prototypical leaders. A multisource field study with leaders (N = 64) and their followers (N = 209) indicated that this interactive effect is more pronounced for leaders who identify more strongly with their team. We discuss how these findings further develop the social identity analysis of leadership.

  3. The effects of local and global processing orientation on eyewitness identification performance.

    PubMed

    Perfect, Timothy J; Dennis, Ian; Snell, Amelia

    2007-10-01

    Recent work has demonstrated that performance on a simultaneous target-present photographic line-up can be enhanced by prior global processing orientation, and hindered by prior local processing orientation induced by processing Navon letter stimuli. A series of studies explore the generality of this processing bias effect using either videotaped scenarios or live interactions. Five experiments demonstrate that these effects are seen across a range of test stimuli, test formats, and test instructions. These data inform the processes engaged in by witnesses when making line-up identifications and indicate that it may be possible to improve the accuracy of witnesses making such judgements.

  4. Identification of Copy Number Variants Defining Genomic Differences among Major Human Groups

    PubMed Central

    Armengol, Lluís; Villatoro, Sergi; González, Juan R.; Pantano, Lorena; García-Aragonés, Manel; Rabionet, Raquel; Cáceres, Mario; Estivill, Xavier

    2009-01-01

    Background Understanding the genetic contribution to phenotype variation of human groups is necessary to elucidate differences in disease predisposition and response to pharmaceutical treatments in different human populations. Methodology/Principal Findings We have investigated the genome-wide profile of structural variation on pooled samples from the three populations studied in the HapMap project by comparative genome hybridization (CGH) in different array platforms. We have identified and experimentally validated 33 genomic loci that show significant copy number differences from one population to the other. Interestingly, we found an enrichment of genes related to environment adaptation (immune response, lipid metabolism and extracellular space) within these regions and the study of expression data revealed that more than half of the copy number variants (CNVs) translate into gene-expression differences among populations, suggesting that they could have functional consequences. In addition, the identification of single nucleotide polymorphisms (SNPs) that are in linkage disequilibrium with the copy number alleles allowed us to detect evidences of population differentiation and recent selection at the nucleotide variation level. Conclusions Overall, our results provide a comprehensive view of relevant copy number changes that might play a role in phenotypic differences among major human populations, and generate a list of interesting candidates for future studies. PMID:19789632

  5. Overcoming asymmetric goals in teams: the interactive roles of team learning orientation and team identification.

    PubMed

    Pearsall, Matthew J; Venkataramani, Vijaya

    2015-05-01

    Although members of teams share a common, ultimate objective, they often have asymmetric or conflicting individual goals that shape the way they contribute to, and pursue, the shared goal of the team. Compounding this problem, they are frequently unaware of the nature of these goal asymmetries or even the fact that such differences exist. Drawing on, and integrating, social interdependence and representational gaps theories, we identify 2 emergent states that combine interactively to enable teams to overcome asymmetric goals: team identification and team learning orientation. Using data from long-term, real-life teams that engaged in a computer simulation designed to create both asymmetric goals and representational gaps about those goals, we found that teams were most effective when they had a high learning orientation coupled with high team identification and that this effect was mediated by teams' ability to form more accurate team goal mental models and engage in effective planning processes. Implications for theory and practice are discussed.

  6. Resequencing, haplotype construction and identification of novel variants of CYP2D6 in Mexican Mestizos.

    PubMed

    Contreras, Alejandra V; Monge-Cazares, Tulia; Alfaro-Ruiz, Luis; Hernandez-Morales, Salvador; Miranda-Ortiz, Haydee; Carrillo-Sanchez, Karol; Jimenez-Sanchez, Gerardo; Silva-Zolezzi, Irma

    2011-05-01

    The CYP2D6 enzyme participates in the metabolism of commonly prescribed drugs: antidepressants, antipsychotics and antihypertensives. The CYP2D6 gene shows a high degree of interindividual and interethnic variability that influences its expression and function. Mexican Mestizos are a recently admixed population resulting from the combination of Amerindian, European and, to a lesser extent, African populations. This study aimed to comprehensively characterize the CYP2D6 gene in Mexican Mestizos. We performed linkage disequilibrium and network analyses in resequencing data of 96 individuals from two regions within Mexico with a different history of admixture and particular population dynamics, the Northwestern state of Sonora and the Central-Pacific state of Guerrero. We identified 64 polymorphisms, including 14 novel variants: 13 SNPs and a CYP2D7 exon 2 conversion, that was assigned CYP2D6*82 by the Human Cytochrome P450 (CYP) Allele Nomenclature Committee. Three novel SNPs were predicted to have functional effects. For CYP2D6*82 we hypothesize an Amerindian origin that is supported by its identification in three Mexican Amerindian groups (Mayas, Tepehuanos and Mixtecos). Frequencies of CYP2D6*1, *2, *4, *5, *10, *29, *53, *82 and its duplications were 50.0, 25.5, 14.1, 2.0, 2.6, 1.0, 0.5, 2.1 and 3.6%, respectively. We found significant frequency differences in CYP2D6*1 and *2 between Mexican Mestizos and in CYP2D6*1, *2, *4, *5, *10 and *29 between Mexicans and at least one other population. We observed strong linkage disequilibrium and phylogenetic relationships between haplotypes. To our knowledge, this study is the first comprehensive resequencing analysis of CYP2D6 in Mexicans or any other Latin American population, providing information about genetic diversity relevant in the development of pharmacogenomics in this region.

  7. Outperforming whom? A multilevel study of performance-prove goal orientation, performance, and the moderating role of shared team identification.

    PubMed

    Dietz, Bart; van Knippenberg, Daan; Hirst, Giles; Restubog, Simon Lloyd D

    2015-11-01

    Performance-prove goal orientation affects performance because it drives people to try to outperform others. A proper understanding of the performance-motivating potential of performance-prove goal orientation requires, however, that we consider the question of whom people desire to outperform. In a multilevel analysis of this issue, we propose that the shared team identification of a team plays an important moderating role here, directing the performance-motivating influence of performance-prove goal orientation to either the team level or the individual level of performance. A multilevel study of salespeople nested in teams supports this proposition, showing that performance-prove goal orientation motivates team performance more with higher shared team identification, whereas performance-prove goal orientation motivates individual performance more with lower shared team identification. Establishing the robustness of these findings, a second study replicates them with individual and team performance in an educational context. (c) 2015 APA, all rights reserved).

  8. Interpretation of Association Signals and Identification of Causal Variants from Genome-wide Association Studies

    PubMed Central

    Wang, Kai; Dickson, Samuel P.; Stolle, Catherine A.; Krantz, Ian D.; Goldstein, David B.; Hakonarson, Hakon

    2010-01-01

    GWAS have been successful in identifying disease susceptibility loci, but it remains a challenge to pinpoint the causal variants in subsequent fine-mapping studies. A conventional fine-mapping effort starts by sequencing dozens of randomly selected samples at susceptibility loci to discover candidate variants, which are then placed on custom arrays or used in imputation algorithms to find the causal variants. We propose that one or several rare or low-frequency causal variants can hitchhike the same common tag SNP, so causal variants may not be easily unveiled by conventional efforts. Here, we first demonstrate that the true effect size and proportion of variance explained by a collection of rare causal variants can be underestimated by a common tag SNP, thereby accounting for some of the “missing heritability” in GWAS. We then describe a case-selection approach based on phasing long-range haplotypes and sequencing cases predicted to harbor causal variants. We compare this approach with conventional strategies on a simulated data set, and we demonstrate its advantages when multiple causal variants are present. We also evaluate this approach in a GWAS on hearing loss, where the most common causal variant has a minor allele frequency (MAF) of 1.3% in the general population and 8.2% in 329 cases. With our case-selection approach, it is present in 88% of the 32 selected cases (MAF = 66%), so sequencing a subset of these cases can readily reveal the causal allele. Our results suggest that thinking beyond common variants is essential in interpreting GWAS signals and identifying causal variants. PMID:20434130

  9. Object-oriented industrial solid waste identification using HJ satellite imagery: a case study of phosphogypsum

    NASA Astrophysics Data System (ADS)

    Fu, Zhuo; Shen, Wenming; Xiao, Rulin; Xiong, Wencheng; Shi, Yuanli; Chen, Baisong

    2012-10-01

    The increasing volume of industrial solid wastes presents a critical problem for the global environment. In the detection and monitoring of these industrial solid wastes, the traditional field methods are generally expensive and time consuming. With the advantages of quick observations taken at a large area, remote sensing provides an effective means for detecting and monitoring the industrial solid wastes in a large scale. In this paper, we employ an object-oriented method for detecting the industrial solid waste from HJ satellite imagery. We select phosphogypsum which is a typical industrial solid waste as our target. Our study area is located in Fuquan in Guizhou province of China. The object oriented method we adopted consists of the following steps: 1) Multiresolution segmentation method is adopted to segment the remote sensing images for obtaining the object-based images. 2) Build the feature knowledge set of the object types. 3) Detect the industrial solid wastes based on the object-oriented decision tree rule set. We analyze the heterogeneity in features of different objects. According to the feature heterogeneity, an object-oriented decision tree rule set is then built for aiding the identification of industrial solid waste. Then, based on this decision tree rule set, the industrial solid waste can be identified automatically from remote sensing images. Finally, the identified results are validated using ground survey data. Experiments and results indicate that the object-oriented method provides an effective method for detecting industrial solid wastes.

  10. Evaluation of BBL CHROMagar orientation medium for detection and presumptive identification of urinary tract pathogens.

    PubMed Central

    Hengstler, K A; Hammann, R; Fahr, A M

    1997-01-01

    The microbiological performance of BBL CHROMagar Orientation medium and CPS ID2 agar was compared to that of Columbia agar with 5% sheep blood and MacConkey agar without crystal violet for the enumeration and presumptive identification of bacteria responsible for urinary tract infections. Of a total of 658 clinical urine specimens, 118 specimens yielded no growth, 402 specimens yielded growth with cell counts of > or = 10(5) CFU/ml, and 138 specimens yielded growth with cell counts of < 10(5) CFU/ml. Of the specimens with cell counts of > or = 10(5) CFU/ml, 163 were pure cultures and 239 were mixed cultures. A total of 266 Escherichia coli organisms were isolated on both chromogenic media, 260 were isolated on blood agar, and 248 were isolated on MacConkey agar. One strain (0.4%) failed to develop the expected pink color on CHROMagar Orientation medium, and 23 strains (8.7%) failed to develop the expected pink color on CPS ID2 agar. Enterococci (CHROMagar Orientation medium, n = 266; CPS ID2 agar, n = 265) produced small blue-green colonies on both chromogenic media. Fifty of the mixed cultures contained enterococci that were detected only on the chromogenic media. The Klebsiella-Enterobacter-Serratia (KES) and the Proteus-Morganella-Providencia (PMP) groups could be identified on both chromogenic media. Of 66 isolates of the KES group, 63 grew with the expected color on CHROMagar Orientation medium and 58 of 64 isolates grew with the expected color on CPS ID2 agar. Other microorganisms required further identification. The use of chromogenic medium formulations offers a time-saving method for the reliable detection, enumeration, and presumptive identification of urinary tract pathogens. One of the greatest advantages of these media is the easy recognition of mixed cultures. PMID:9350731

  11. Semiconductor Whole Exome Sequencing for the Identification of Genetic Variants in Colombian Patients Clinically Diagnosed with Long QT Syndrome.

    PubMed

    Burgos, Mariana; Arenas, Alvaro; Cabrera, Rodrigo

    2016-08-01

    Inherited long QT syndrome (LQTS) is a cardiac channelopathy characterized by a prolongation of QT interval and the risk of syncope, cardiac arrest, and sudden cardiac death. Genetic diagnosis of LQTS is critical in medical practice as results can guide adequate management of patients and distinguish phenocopies such as catecholaminergic polymorphic ventricular tachycardia (CPVT). However, extensive screening of large genomic regions is required in order to reliably identify genetic causes. Semiconductor whole exome sequencing (WES) is a promising approach for the identification of variants in the coding regions of most human genes. DNA samples from 21 Colombian patients clinically diagnosed with LQTS were enriched for coding regions using multiplex polymerase chain reaction (PCR) and subjected to WES using a semiconductor sequencer. Semiconductor WES showed mean coverage of 93.6 % for all coding regions relevant to LQTS at >10× depth with high intra- and inter-assay depth heterogeneity. Fifteen variants were detected in 12 patients in genes associated with LQTS. Three variants were identified in three patients in genes associated with CPVT. Co-segregation analysis was performed when possible. All variants were analyzed with two pathogenicity prediction algorithms. The overall prevalence of LQTS and CPVT variants in our cohort was 71.4 %. All LQTS variants previously identified through commercial genetic testing were identified. Standardized WES assays can be easily implemented, often at a lower cost than sequencing panels. Our results show that WES can identify LQTS-causing mutations and permits differential diagnosis of related conditions in a real-world clinical setting. However, high heterogeneity in sequencing depth and low coverage in the most relevant genes is expected to be associated with reduced analytical sensitivity.

  12. An Optimal Weighted Aggregated Association Test for Identification of Rare Variants Involved in Common Diseases

    PubMed Central

    Sul, Jae Hoon; Han, Buhm; He, Dan; Eskin, Eleazar

    2011-01-01

    The advent of next generation sequencing technologies allows one to discover nearly all rare variants in a genomic region of interest. This technological development increases the need for an effective statistical method for testing the aggregated effect of rare variants in a gene on disease susceptibility. The idea behind this approach is that if a certain gene is involved in a disease, many rare variants within the gene will disrupt the function of the gene and are associated with the disease. In this article, we present the rare variant weighted aggregate statistic (RWAS), a method that groups rare variants and computes a weighted sum of differences between case and control mutation counts. We show that our method outperforms the groupwise association test of Madsen and Browning in the disease-risk model that assumes that each variant makes an equally small contribution to disease risk. In addition, we can incorporate prior information into our method of which variants are likely causal. By using simulated data and real mutation screening data of the susceptibility gene for ataxia telangiectasia, we demonstrate that prior information has a substantial influence on the statistical power of association studies. Our method is publicly available at http://genetics.cs.ucla.edu/rarevariants. PMID:21368279

  13. Identification of Three Novel Splicing Variants and Expression Analysis of Chicken GPR1 Gene.

    PubMed

    Zhang, Xueyou; Xiao, Qihai; Tian, Kai; Wang, Yan; Zhao, Xiaoling; Yin, Huadong; Li, Diyan; Zhu, Qing

    2017-01-01

    GPR1 is a G protein-coupled receptor that plays critical roles in eukaryotic cells: typically, response to glucose stimulation, lipid accumulation, and transmitting nutrition signals to cAMP pathway. However, the alternative splicing of the GPR1 gene and its expression pattern in chicken tissues and ovarian follicles were unknown. In our current study, we used RACE-PCR to identify three GPR1 variants, including the full-length variant (GPR1-va1) and two alternatively spliced variants (GPR1-va2, GPR1-vb). Quantitative real-time PCR examined the expression pattern of GPR1 mRNA in chicken tissues and ovarian follicles. The result reveals that the coding sequence of the three variants cDNA is 1053, 1053, and 627 bp in length, encoding 350, 350, and 208 amino acids, respectively. The three variants of GPR1 show similar tissue distributions; GPR1 expression was abundant in the abdominal fat, lung, and heart. With the follicular development, the expression of GPR1 gene gradually increased, and GPR1-va1 and GPR1-va2 spliced variants expression in F2 were significantly higher than in F5, F4, and prehierarchical follicles (P < 0.05). Taken together, we found three novel variants of GPR1, and the results of GPR1 expression profiling in adipose tissues and ovarian follicles suggest that GPR1 may play a significant role in the lipid accumulation and progression of follicular development.

  14. Identification of msp1 Gene Variants in Populations of Meloidogyne incognita Using PCR-DGGE

    PubMed Central

    Adam, Mohamed; Hallmann, Johannes; Heuer, Holger

    2014-01-01

    Effectors of root-knot nematodes are essential for parasitism and prone to recognition by adapted variants of the host plants. This selective pressure initiates hypervariability of effector genes. Diversity of the gene variants within nematode populations might correlate with host preferences. In this study we developed a method to compare the distribution of variants of the effector gene msp1 among populations of Meloidogyne incognita. Primers were designed to amplify a 234-bp fragment of msp1. Sequencing of cloned PCR products revealed five msp1 variants from seven populations that were distinguishable in their reproduction on five host plants. A protocol for denaturing gradient gel electrophoresis (DGGE) was developed to separate these msp1 variants. DGGE for replicated pools of juveniles from the seven populations revealed ten variants of msp1. A correlation between the presence of a particular gene variant and the reproductive potential on particular hosts was not evident. Especially race 3 showed substantial variation within the population. DGGE fingerprints of msp1 tended to cluster the populations according to their reproduction rate on pepper. The developed method could be useful for analyzing population heterogeneity and epidemiology of M. incognita. PMID:25276001

  15. Identification of Three Novel Splicing Variants and Expression Analysis of Chicken GPR1 Gene

    PubMed Central

    Zhang, Xueyou; Xiao, Qihai; Tian, Kai; Zhao, Xiaoling; Yin, Huadong; Li, Diyan

    2017-01-01

    GPR1 is a G protein-coupled receptor that plays critical roles in eukaryotic cells: typically, response to glucose stimulation, lipid accumulation, and transmitting nutrition signals to cAMP pathway. However, the alternative splicing of the GPR1 gene and its expression pattern in chicken tissues and ovarian follicles were unknown. In our current study, we used RACE-PCR to identify three GPR1 variants, including the full-length variant (GPR1-va1) and two alternatively spliced variants (GPR1-va2, GPR1-vb). Quantitative real-time PCR examined the expression pattern of GPR1 mRNA in chicken tissues and ovarian follicles. The result reveals that the coding sequence of the three variants cDNA is 1053, 1053, and 627 bp in length, encoding 350, 350, and 208 amino acids, respectively. The three variants of GPR1 show similar tissue distributions; GPR1 expression was abundant in the abdominal fat, lung, and heart. With the follicular development, the expression of GPR1 gene gradually increased, and GPR1-va1 and GPR1-va2 spliced variants expression in F2 were significantly higher than in F5, F4, and prehierarchical follicles (P < 0.05). Taken together, we found three novel variants of GPR1, and the results of GPR1 expression profiling in adipose tissues and ovarian follicles suggest that GPR1 may play a significant role in the lipid accumulation and progression of follicular development. PMID:28203567

  16. Approach/Avoidance Orientations Affect Self-Construal and Identification with In-group

    PubMed Central

    Nussinson, Ravit; Häfner, Michael; Seibt, Beate; Strack, Fritz; Trope, Yaacov

    2011-01-01

    Approach and avoidance are two basic motivational orientations. Their activation influences cognitive and perceptive processes: Previous work suggests that an approach orientation instigates a focus on larger units as compared to avoidance. Study 1 confirms this assumption using a paradigm that more directly taps a person’s tendency to represent objects as belonging to small or large units than prior studies. It was further predicted that the self should also be represented as belonging to larger units, and hence be more interdependent under approach than under avoidance. Study 2 supports this prediction. As a consequence of this focus on belonging to larger units, it was finally predicted that approach results in a stronger identification with one’s in-group than avoidance. Studies 3 and 4 support that prediction. PMID:22844229

  17. Identification and in vivo functional characterization of novel compound heterozygous BMP1 variants in osteogenesis imperfecta.

    PubMed

    Cho, Sung Yoon; Asharani, P V; Kim, Ok-Hwa; Iida, Aritoshi; Miyake, Noriko; Matsumoto, Naomichi; Nishimura, Gen; Ki, Chang-Seok; Hong, Geehay; Kim, Su Jin; Sohn, Young Bae; Park, Sung Won; Lee, Jieun; Kwun, Younghee; Carney, Thomas J; Huh, Rimm; Ikegawa, Shiro; Jin, Dong-Kyu

    2015-02-01

    Osteogenesis imperfecta (OI) comprises a heterogeneous group of disorders that are characterized by susceptibility to bone fractures, and range in severity from a subtle increase in fracture frequency to death in the perinatal period. Most patients have defects in type I collagen biosynthesis with autosomal-dominant inheritance, but many autosomal-recessive genes have been reported. We applied whole-exome sequencing to identify mutations in a Korean OI patient who had an umbilical hernia, frequent fractures, a markedly short stature, delayed motor development, scoliosis, and dislocation of the radial head, with a bowed radius and ulna. We identified two novel variants in the BMP1 gene: c.808A>G and c.1297G>T. The former variant caused a missense change p.(Met270Val) and the latter variant caused the skipping of exon 10. The hypofunctional nature of the two variants was demonstrated in a zebrafish assay.

  18. Large-scale identification of sequence variants influencing human transcription factor occupancy in vivo.

    PubMed

    Maurano, Matthew T; Haugen, Eric; Sandstrom, Richard; Vierstra, Jeff; Shafer, Anthony; Kaul, Rajinder; Stamatoyannopoulos, John A

    2015-12-01

    The function of human regulatory regions depends exquisitely on their local genomic environment and on cellular context, complicating experimental analysis of common disease- and trait-associated variants that localize within regulatory DNA. We use allelically resolved genomic DNase I footprinting data encompassing 166 individuals and 114 cell types to identify >60,000 common variants that directly influence transcription factor occupancy and regulatory DNA accessibility in vivo. The unprecedented scale of these data enables systematic analysis of the impact of sequence variation on transcription factor occupancy in vivo. We leverage this analysis to develop accurate models of variation affecting the recognition sites for diverse transcription factors and apply these models to discriminate nearly 500,000 common regulatory variants likely to affect transcription factor occupancy across the human genome. The approach and results provide a new foundation for the analysis and interpretation of noncoding variation in complete human genomes and for systems-level investigation of disease-associated variants.

  19. An orientation selective neural network and its application to cosmic muon identification

    NASA Astrophysics Data System (ADS)

    Abramowicz, Halina; Horn, David; Naftaly, Ury; Sahar-Pikielny, Carmit

    1996-02-01

    We propose a novel method for identification of a linear pattern of pixels on a two-dimensional grid. Following principles employed by the visual cortex, we employ orientation selective neurons in a neural network which performs this task. The method is then applied to a sample of data collected with the ZEUS detector at HERA in order to identify cosmic muons which leave a linear pattern of signals in the segmented uranium-scintillator calorimeter. A two dimensional representation of the relevant part of the detector is used. The results compared with a visual scan point to a very satisfactory cosmic muon identification. The algorithm performs well in the presence of noise and pixels with limited efficiency. Given its architecture, this system becomes a good candidate for fast pattern recognition in parallel processing devices.

  20. Ultrasensitive Identification of Localization Variants of Modified Peptides Using Ion Mobility Spectrometry

    SciTech Connect

    Ibrahim, Yehia M.; Shvartsburg, Alexandre A.; Smith, Richard D.; Belov, Mikhail E.

    2011-05-28

    Localization of the modification sites on peptides is challenging, particularly when multiple modifications or mixtures of localization isomers (variants) are involved. Such variants commonly coelute in liquid chromatography and may be undistinguishable in tandem mass spectrometry (MS/MS) for lack of unique fragments. Here, we have resolved the variants of singly and doubly phosphorylated peptides employing drift tube ion mobility spectrometry (IMS) coupled to time-of-flight mass spectrometry. Even with a moderate IMS resolving power of ~80, substantial separation was achieved for both 2+ and 3+ ions normally generated by electrospray ionization, including for the variant indistinguishable by MS/MS. Variants often exhibit a distribution of 3-D conformers, which can be adjusted for optimum IMS separation by prior field heating of ions in a funnel trap. The peak assignments were confirmed using MS/MS after IMS separation, but known species could be identified using just the ion mobility "tag". Avoiding the MS/MS step lowers the detection limit of localization variants to <100 attomoles, an order of magnitude better than provided by electron transfer dissociation in an Orbitrap MS.

  1. Identification of an APC Variant in a Patient with Clinical Attenuated Familial Adenomatous Polyposis.

    PubMed

    Schlussel, Andrew T; Donlon, Susan S; Eggerding, Faye A; Gagliano, Ronald A

    2014-01-01

    Introduction. The objective of this case report is to discuss an unclassified germline variant of the adenomatous polyposis coli (APC) gene identified in an older patient with attenuated familial adenomatous polyposis syndrome (AFAP). Methods. We present a case report of a 66-year-old man diagnosed with AFAP. Colonoscopy found multiple polyps and invasive adenocarcinoma arising in the transverse colon. Samples were tested for mutations in the APC gene. Results. DNA sequencing of germline DNA identified a cytosine (C) to thymine (T) transition at nucleotide 1240, heterozygous. The C to T transition at codon 414 is predicted to convert an arginine residue to a cysteine that is possibly pathogenic. Analysis of the patient's colon tumor DNA indicated that the tumor had lost the mutant variant allele and retained only the normal allele, suggesting that the variant may not be significant. Conclusions. The p.R414C variant has been described previously as a germline mutation of probable pathogenicity. This substitution should be considered an unclassified variant and possibly not pathogenic. These findings support the need for further genetic testing of tissue, as well as for developing a mechanism for testing all variants, as this could significantly impact the lives of patients and their family members.

  2. Identification and characterisation of alternative transcriptional variants of PDGFRL in two lines of commercial poultry.

    PubMed

    McDerment, N A; Hocking, P M; Dunn, I C

    2015-10-01

    The platelet-derived growth factor (PDGF) family of genes and their receptors are involved in angiogenesis and steroid hormone production. A putative member of the family, platelet-derived growth factor receptor-like (PDGFRL), has been implicated in steroid-based feedback mechanisms within the chicken reproductive system. Three potential variants of PDGFRL were identified in the chicken, supported by in silico prediction and EST sequencing. The three potential transcripts have been further verified and the 5' terminal regions sequenced in this research. The sum of expression of all three transcripts in broiler breeders (the parents of broiler chickens) has been shown to be consistent with total expression of the gene. However, cumulative expression of the three transcripts in a range of tissues in egg layers was significantly short of total expression, indicating the existence of potential additional variants. Two additional variants were subsequently identified in egg layer cerebellum tissue and the 5' terminal regions sequenced. Sequence analysis of the three initial variants suggests that only one variant, which was the most abundant in broiler breeders and the majority of egg layer tissues, had a functional signal peptide. Although 5' RACE identified two additional transcripts in egg layers, the most likely protein translations indicated that these variants possessed no functional signal peptide, suggesting that, if they have a function, it is not a traditional one. © 2015 Stichting International Foundation for Animal Genetics.

  3. Identification of androgen receptor variants in testis from humans and other vertebrates.

    PubMed

    Laurentino, S S; Pinto, P I S; Tomás, J; Cavaco, J E; Sousa, M; Barros, A; Power, D M; Canário, A V M; Socorro, S

    2013-06-01

    The androgen receptor (AR) is a ligand-activated transcription factor member of the nuclear receptor superfamily. The existence of alternatively spliced variants is well recognised for several members of this superfamily, most of them having functional importance. For example, several testicular oestrogen receptor variants have been suggested to play a role in the regulation of spermatogenesis. However, information on AR variants is mostly related to cancer and androgen insensitivity syndrome (AIS) cases. The objective of this study was to investigate the expression of AR variants in the testis from humans and other vertebrates. Four AR variants [ARΔ2(Stop) , ARΔ2(23Stop) , ARΔ3 and ARΔ4(120)] were identified in human testis. ARΔ2(Stop) and ARΔ3, with exon 2 or 3 deleted, respectively, were also expressed in human liver, lung, kidney and heart. In addition, ARΔ2(Stop) was expressed in rat and gilthead seabream testis, while an ARΔ3 was detected in African clawed frog testis. This is the first report revealing the existence of AR variants in the testis of evolutionarily distant vertebrate species and in nonpathological tissues. These data suggest the functional importance of these novel AR forms and demonstrate a complexity in AR signalling that is not exclusive of pathological conditions.

  4. Identification of polymorphisms and sequence variants in the human homologue of the mouse natural resistance-associated macrophage protein gene

    SciTech Connect

    Liu, Jing; Fujiwara, T.M.; Buu, N.T.; Sanchez, F.O.; Cellier, M.; Paradis, A.J.; Frappier, D.; Skamene, E.; Gros, P.; Morgan, K.

    1995-04-01

    The most common mycobacterial disease in humans is tuberculosis, and there is evidence for genetic factors in susceptibility to tuberculosis. In the mouse, the Bcg gene controls macrophage priming for activation and is a major gene for susceptibility to infection with mycobacteria. A candidate gene for Bcg was identified by positional cloning and was designated {open_quotes}natural resistance-associated macrophage protein gene{close_quotes} (Nramp1), and the human homologue (NRAMP1) has recently been cloned. Here we report (1) the physical mapping NRAMP1 close to VIL in chromosome region 2q35 by PCR analysis of somatic cell hybrids and YAC cloning and (2) the identification of nine sequence variants in NRAMP1. Of the four variants in the coding region, there were two missense mutations and two silent substitutions. The missense mutations were a conservative alanine-to-valine substitution at codon 318 in exon9 and an aspartic acid-to-asparagine substitution at codon 543 in the predicted cytoplasmic tail of the NRAMP1 protein. A microsatellite was located in the immediate 5{prime} region of the gene, three variants were in introns, and one variant was located in the 3{prime} UTR. The allele frequencies of each of the nine variants were determined in DNA samples of 60 Caucasians and 20 Asians. In addition, we have physically linked two highly polymorphic microsatellite markers, D2S104 and D2S173, to NRAMP1 on a 1.5-Mb YAC contig. These molecular markers will be useful to assess the role of NRAMP1 in susceptibility to tuberculosis and other macrophage-mediated diseases. 40 refs., 3 figs., 2 tabs.

  5. Identification of BRCA1 missense substitutions that confer partial functional activity: potential moderate risk variants?

    PubMed Central

    Lovelock, Paul K; Spurdle, Amanda B; Mok, Myth TS; Farrugia, Daniel J; Lakhani, Sunil R; Healey, Sue; Arnold, Stephen; Buchanan, Daniel; Investigators, kConFab; Couch, Fergus J; Henderson, Beric R; Goldgar, David E; Tavtigian, Sean V; Chenevix-Trench, Georgia; Brown, Melissa A

    2007-01-01

    Introduction Many of the DNA sequence variants identified in the breast cancer susceptibility gene BRCA1 remain unclassified in terms of their potential pathogenicity. Both multifactorial likelihood analysis and functional approaches have been proposed as a means to elucidate likely clinical significance of such variants, but analysis of the comparative value of these methods for classifying all sequence variants has been limited. Methods We have compared the results from multifactorial likelihood analysis with those from several functional analyses for the four BRCA1 sequence variants A1708E, G1738R, R1699Q, and A1708V. Results Our results show that multifactorial likelihood analysis, which incorporates sequence conservation, co-inheritance, segregation, and tumour immunohistochemical analysis, may improve classification of variants. For A1708E, previously shown to be functionally compromised, analysis of oestrogen receptor, cytokeratin 5/6, and cytokeratin 14 tumour expression data significantly strengthened the prediction of pathogenicity, giving a posterior probability of pathogenicity of 99%. For G1738R, shown to be functionally defective in this study, immunohistochemistry analysis confirmed previous findings of inconsistent 'BRCA1-like' phenotypes for the two tumours studied, and the posterior probability for this variant was 96%. The posterior probabilities of R1699Q and A1708V were 54% and 69%, respectively, only moderately suggestive of increased risk. Interestingly, results from functional analyses suggest that both of these variants have only partial functional activity. R1699Q was defective in foci formation in response to DNA damage and displayed intermediate transcriptional transactivation activity but showed no evidence for centrosome amplification. In contrast, A1708V displayed an intermediate transcriptional transactivation activity and a normal foci formation response in response to DNA damage but induced centrosome amplification. Conclusion

  6. Identification of Gender-Specific Genetic Variants in Patients With Bicuspid Aortic Valve.

    PubMed

    Dargis, Natasha; Lamontagne, Maxime; Gaudreault, Nathalie; Sbarra, Laura; Henry, Cyndi; Pibarot, Philippe; Mathieu, Patrick; Bossé, Yohan

    2016-02-01

    Bicuspid aortic valve (BAV) is the most frequent congenital heart defect and has a male predominance of 3 to 1. A large proportion of patients develop valvular and aortic complications. Despite the high prevalence of BAV, its cause and genetic origins remain elusive. The goal of this study was to identify genetic variants associated with BAV. Nine genes previously associated with BAV (NOTCH1, AXIN1, EGFR, ENG, GATA5, NKX2-5, NOS3, PDIA2, and TGFBR2) were sequenced in 48 patients with BAV using the Ion Torrent Personal Genome Machine. Pathogenicity of genetic variants was evaluated with the Combined Annotation Dependent Depletion framework. A selection of 89 variants identified by sequencing or in previous BAV genetic studies was genotyped, and allele frequencies were compared in 323 patients with BAV confirmed at surgery and 584 controls. Analyses were also performed by gender. Nine novel and 19 potentially pathogenic variants were identified by next-generation sequencing and confirmed by Sanger sequencing, but they were not associated with BAV in the case-control population. A significant association was observed between an in silico-predicted benign EGFR intronic variant (rs17290301) and BAV. Analyses performed by gender revealed different variants associated with BAV in men (EGFR rs533525993 and TEX26 rs12857479) and women (NOTCH1 rs61751489, TGFBR2 rs1155705, and NKX2-5 rs2277923). In conclusion, these results constitute the first association between EGFR genetic variants and BAV in humans and support a possible role of gender-specific polymorphisms in the development of BAV.

  7. New CD20 alternative splice variants: molecular identification and differential expression within hematological B cell malignancies.

    PubMed

    Gamonet, Clémentine; Bole-Richard, Elodie; Delherme, Aurélia; Aubin, François; Toussirot, Eric; Garnache-Ottou, Francine; Godet, Yann; Ysebaert, Loïc; Tournilhac, Olivier; Caroline, Dartigeas; Larosa, Fabrice; Deconinck, Eric; Saas, Philippe; Borg, Christophe; Deschamps, Marina; Ferrand, Christophe

    2015-01-01

    CD20 is a B cell lineage-specific marker expressed by normal and leukemic B cells and targeted by several antibody immunotherapies. We have previously shown that the protein from a CD20 mRNA splice variant (D393-CD20) is expressed at various levels in leukemic B cells or lymphoma B cells but not in resting, sorted B cells from the peripheral blood of healthy donors. Western blot (WB) analysis of B malignancy primary samples showed additional CD20 signals. Deep molecular PCR analysis revealed four new sequences corresponding to in-frame CD20 splice variants (D657-CD20, D618-CD20, D480-CD20, and D177-CD20) matching the length of WB signals. We demonstrated that the cell spliceosome machinery can process ex vivo D480-, D657-, and D618-CD20 transcript variants by involving canonical sites associated with cryptic splice sites. Results of specific and quantitative RT-PCR assays showed that these CD20 splice variants are differentially expressed in B malignancies. Moreover, Epstein-Barr virus (EBV) transformation modified the CD20 splicing profile and mainly increased the D393-CD20 variant transcripts. Finally, investigation of three cohorts of chronic lymphocytic leukemia (CLL) patients showed that the total CD20 splice variant expression was higher in a stage B and C sample collection compared to routinely collected CLL samples or relapsed refractory stage A, B, or C CLL. The involvement of these newly discovered alternative CD20 transcript variants in EBV transformation makes them interesting molecular indicators, as does their association with oncogenesis rather than non-oncogenic B cell diseases, differential expression in B cell malignancies, and correlation with CLL stage and some predictive CLL markers. This potential should be investigated in further studies.

  8. A low density microarray method for the identification of human papillomavirus type 18 variants.

    PubMed

    Meza-Menchaca, Thuluz; Williams, John; Rodríguez-Estrada, Rocío B; García-Bravo, Aracely; Ramos-Ligonio, Ángel; López-Monteon, Aracely; Zepeda, Rossana C

    2013-09-26

    We describe a novel microarray based-method for the screening of oncogenic human papillomavirus 18 (HPV-18) molecular variants. Due to the fact that sequencing methodology may underestimate samples containing more than one variant we designed a specific and sensitive stacking DNA hybridization assay. This technology can be used to discriminate between three possible phylogenetic branches of HPV-18. Probes were attached covalently on glass slides and hybridized with single-stranded DNA targets. Prior to hybridization with the probes, the target strands were pre-annealed with the three auxiliary contiguous oligonucleotides flanking the target sequences. Screening HPV-18 positive cell lines and cervical samples were used to evaluate the performance of this HPV DNA microarray. Our results demonstrate that the HPV-18's variants hybridized specifically to probes, with no detection of unspecific signals. Specific probes successfully reveal detectable point mutations in these variants. The present DNA oligoarray system can be used as a reliable, sensitive and specific method for HPV-18 variant screening. Furthermore, this simple assay allows the use of inexpensive equipment, making it accessible in resource-poor settings.

  9. A Low Density Microarray Method for the Identification of Human Papillomavirus Type 18 Variants

    PubMed Central

    Meza-Menchaca, Thuluz; Williams, John; Rodríguez-Estrada, Rocío B.; García-Bravo, Aracely; Ramos-Ligonio, Ángel; López-Monteon, Aracely; Zepeda, Rossana C.

    2013-01-01

    We describe a novel microarray based-method for the screening of oncogenic human papillomavirus 18 (HPV-18) molecular variants. Due to the fact that sequencing methodology may underestimate samples containing more than one variant we designed a specific and sensitive stacking DNA hybridization assay. This technology can be used to discriminate between three possible phylogenetic branches of HPV-18. Probes were attached covalently on glass slides and hybridized with single-stranded DNA targets. Prior to hybridization with the probes, the target strands were pre-annealed with the three auxiliary contiguous oligonucleotides flanking the target sequences. Screening HPV-18 positive cell lines and cervical samples were used to evaluate the performance of this HPV DNA microarray. Our results demonstrate that the HPV-18's variants hybridized specifically to probes, with no detection of unspecific signals. Specific probes successfully reveal detectable point mutations in these variants. The present DNA oligoarray system can be used as a reliable, sensitive and specific method for HPV-18 variant screening. Furthermore, this simple assay allows the use of inexpensive equipment, making it accessible in resource-poor settings. PMID:24077317

  10. Identification of a rare coding variant in TREM2 in a Chinese individual with Alzheimer's disease.

    PubMed

    Bonham, Luke W; Sirkis, Daniel W; Fan, Jia; Aparicio, Renan E; Tse, Marian; Ramos, Eliana Marisa; Wang, Qing; Coppola, Giovanni; Rosen, Howard J; Miller, Bruce L; Yokoyama, Jennifer S

    2017-02-01

    Rare variation in the TREM2 gene is associated with a broad spectrum of neurodegenerative disorders including Alzheimer's disease (AD). TREM2 encodes a receptor expressed in microglia which is thought to influence neurodegeneration by sensing damage signals and regulating neuroinflammation. Many of the variants reported to be associated with AD, including the rare R47H variant, were discovered in populations of European ancestry and have not replicated in diverse populations from other genetic backgrounds. We utilized a cohort of elderly Chinese individuals diagnosed as cognitively normal, or with mild cognitive impairment or AD to identify a rare variant, A192T, present in a single patient diagnosed with AD. We characterized this variant using biochemical cell surface expression assays and found that it significantly altered cell surface expression of the TREM2 protein. Together these data provide evidence that the A192T variant in TREM2 could contribute risk for AD. This study underscores the increasingly recognized role of immune-related processes in AD and highlights the importance of including diverse populations in research to identify genetic variation that contributes risk for AD and other neurodegenerative disorders.

  11. Identification of carriers of a variant plasma prealbumin (transthyretin) associated with familial amyloidotic polyneuropathy type I.

    PubMed Central

    Benson, M D; Dwulet, F E

    1985-01-01

    A method is described for detecting carriers of a variant plasma prealbumin that is associated with familial amyloidotic polyneuropathy (FAP) type I. It is based on the finding of an extra methionine in the variant prealbumin, at position 30 from the amino terminals. Since normal prealbumin has only one methionine (position 13), treatment with cyanogen bromide (CNBr), which cleaves only at methionines, results in two peptides. CNBr treatment of the variant prealbumin gives three peptides. The extra can then be detected in two ways: by HPLC using a reverse phase C18 column, and by sequential Edman degradation. Each method can detect as little as 1% variant prealbumin in isolated plasma prealbumin, and therefore, can identify carriers of the gene for the variant protein. Since FAP type I usually is not manifest until after the childbearing years, this method to identify carriers of the gene offers a new approach for genetic counseling of families with this disease. To date, kindreds with hereditary amyloidosis that could benefit from these studies include those with FAP type I of Swedish, Japanese, and Portuguese origins. PMID:2981253

  12. Identification of Novel FMR1 Variants by Massively Parallel Sequencing in Developmentally Delayed Males

    PubMed Central

    Collins, Stephen C.; Bray, Steven M.; Suhl, Joshua A.; Cutler, David J.; Coffee, Bradford; Zwick, Michael E.; Warren, Stephen T.

    2010-01-01

    Fragile X syndrome (FXS), the most common inherited form of developmental delay, is typically caused by CGG-repeat expansion in FMR1. However, little attention has been paid to sequence variants in FMR1. Through the use of pooled-template massively parallel sequencing, we identified 130 novel FMR1 sequence variants in a population of 963 developmentally delayed males without CGG-repeat expansion mutations. Among these, we identified a novel missense change, p.R138Q, which alters a conserved residue in the nuclear localization signal of FMRP. We have also identified three promoter mutations in this population, all of which significantly reduce in vitro levels of FMR1 transcription. Additionally, we identified 10 noncoding variants of possible functional significance in the introns and 3’-untranslated region of FMR1, including two predicted splice site mutations. These findings greatly expand the catalogue of known FMR1 sequence variants and suggest that FMR1 sequence variants may represent an important cause of developmental delay. PMID:20799337

  13. Identification of novel FMR1 variants by massively parallel sequencing in developmentally delayed males.

    PubMed

    Collins, Stephen C; Bray, Steven M; Suhl, Joshua A; Cutler, David J; Coffee, Bradford; Zwick, Michael E; Warren, Stephen T

    2010-10-01

    Fragile X syndrome (FXS), the most common inherited form of developmental delay, is typically caused by CGG-repeat expansion in FMR1. However, little attention has been paid to sequence variants in FMR1. Through the use of pooled-template massively parallel sequencing, we identified 130 novel FMR1 sequence variants in a population of 963 developmentally delayed males without CGG-repeat expansion mutations. Among these, we identified a novel missense change, p.R138Q, which alters a conserved residue in the nuclear localization signal of FMRP. We have also identified three promoter mutations in this population, all of which significantly reduce in vitro levels of FMR1 transcription. Additionally, we identified 10 noncoding variants of possible functional significance in the introns and 3'-untranslated region of FMR1, including two predicted splice site mutations. These findings greatly expand the catalog of known FMR1 sequence variants and suggest that FMR1 sequence variants may represent an important cause of developmental delay. Copyright © 2010 Wiley-Liss, Inc.

  14. Identification of a Chicken Anemia Virus Variant-Related Gyrovirus in Stray Cats in China, 2012

    PubMed Central

    Liu, Yuanjia; Ji, Jun; Chen, Feng; Sun, Baoli; Xue, Chunyi; Ma, Jingyun; Bi, Yingzuo; Xie, Qingmei

    2014-01-01

    The chicken anemia virus (CAV), is a known member of the genus Gyrovirus and was first isolated from chickens in Japan in 1979. Some reports have also demonstrated that CAV can be identified in human stool specimens. In this study, a variant of CAV was detected using PCR with CAV-based primers in fecal samples of stray cats. The genome of CAV variant was sequenced and the results suggest that it could be a recombinant viral strain from parental CAV strains JQ690762 and AF311900. Recombination is an important evolutionary mechanism that contributes to genetic diversification. These findings indicate that CAV variant might have originated from CAV-infected chickens. The epidemiology and pathogenesis of this novel virus remains to be elucidated. This study underscores the importance of CAV surveillance and it presents the first evidence suggesting the possibility of CAV homologous recombination in cat. PMID:24689034

  15. Identification of Blastocystis Subtype 1 Variants in the Home for Girls, Bangkok, Thailand

    PubMed Central

    Thathaisong, Umaporn; Siripattanapipong, Suradej; Mungthin, Mathirut; Pipatsatitpong, Duangnate; Tan-ariya, Peerapan; Naaglor, Tawee; Leelayoova, Saovanee

    2013-01-01

    A cross-sectional study of Blastocystis infection was conducted to evaluate the prevalence, risk factors, and subtypes of Blastocystis at the Home for Girls, Bangkok, Thailand in November 2008. Of 370 stool samples, 118 (31.9%) were infected with Blastocystis. Genotypic characterization of Blastocystis was performed by polymerase chain reaction and sequence analysis of the partial small subunit ribosomal RNA (SSU rRNA) gene. Subtype 1 was the most predominant (94.8%), followed by subtype 6 (3.5%) and subtype 2 (1.7%). Sequence analyses revealed nucleotide polymorphisms for Blastocystis subtype 1, which were described as subtype 1/variant 1, subtype 1/variant 2. Blastocystis subtype 1/variant 1 was the most predominant infection occurring in almost every house. The results showed that subtype analysis of Blastocystis was useful for molecular epidemiological study. PMID:23166199

  16. Identification of Blastocystis subtype 1 variants in the Home for Girls, Bangkok, Thailand.

    PubMed

    Thathaisong, Umaporn; Siripattanapipong, Suradej; Mungthin, Mathirut; Pipatsatitpong, Duangnate; Tan-ariya, Peerapan; Naaglor, Tawee; Leelayoova, Saovanee

    2013-02-01

    A cross-sectional study of Blastocystis infection was conducted to evaluate the prevalence, risk factors, and subtypes of Blastocystis at the Home for Girls, Bangkok, Thailand in November 2008. Of 370 stool samples, 118 (31.9%) were infected with Blastocystis. Genotypic characterization of Blastocystis was performed by polymerase chain reaction and sequence analysis of the partial small subunit ribosomal RNA (SSU rRNA) gene. Subtype 1 was the most predominant (94.8%), followed by subtype 6 (3.5%) and subtype 2 (1.7%). Sequence analyses revealed nucleotide polymorphisms for Blastocystis subtype 1, which were described as subtype 1/variant 1, subtype 1/variant 2. Blastocystis subtype 1/variant 1 was the most predominant infection occurring in almost every house. The results showed that subtype analysis of Blastocystis was useful for molecular epidemiological study.

  17. Systematic RH genotyping and variant identification in French donors of African origin

    PubMed Central

    Kappler-Gratias, Sandrine; Auxerre, Carine; Dubeaux, Isabelle; Beolet, Marylise; Ripaux, Maryline; Le Pennec, Pierre-Yves; Pham, Bach-Nga

    2014-01-01

    Background RH molecular analysis has enabled the documentation of numerous variants of RHD and RHCE alleles, especially in individuals of African origin. The aim of the present study was to determine the type and frequency of D and/or RhCE variants among blood donors of African origin in France, by performing a systematic RH molecular analysis, in order to evaluate the implications for blood transfusion of patients of African origin. Materials and methods Samples from 316 African blood donors, whose origin was established by their Fy(a−b−) phenotype, were first analysed using the RHD and RHCE BeadChips Kit (BioArray Solutions, Immucor, Warren, NJ, USA). Sequencing was performed when necessary. Results RHD molecular analysis showed that 26.2% of donors had a variant RHD allele. It allowed the prediction of a partial D in 11% of cases. RHCE molecular analysis showed that 14.2% of donors had a variant RHCE allele or RH [RN or (C)ces] haplotype. A rare Rh phenotype associated with the loss of a high-prevalence antigen or partial RhCE antigens were predicted from RHCE molecular analysis in 1 (0.3%) and 17 (5%) cases, respectively. Discussion Systematic RHD and RHCE molecular analysis performed in blood donors of African origin provides transfusion-relevant information for individuals of African origin because of the frequency of variant RH alleles. RH molecular analysis may improve transfusion therapy of patients by allowing better donor and recipient matching, based not only on phenotypically matched red blood cell units, but also on units that are genetically matched with regards to RhCE variants. PMID:23867180

  18. Systematic RH genotyping and variant identification in French donors of African origin.

    PubMed

    Kappler-Gratias, Sandrine; Auxerre, Carine; Dubeaux, Isabelle; Beolet, Marylise; Ripaux, Maryline; Le Pennec, Pierre-Yves; Pham, Bach-Nga

    2014-01-01

    RH molecular analysis has enabled the documentation of numerous variants of RHD and RHCE alleles, especially in individuals of African origin. The aim of the present study was to determine the type and frequency of D and/or RhCE variants among blood donors of African origin in France, by performing a systematic RH molecular analysis, in order to evaluate the implications for blood transfusion of patients of African origin. Samples from 316 African blood donors, whose origin was established by their Fy(a-b-) phenotype, were first analysed using the RHD and RHCE BeadChips Kit (BioArray Solutions, Immucor, Warren, NJ, USA). Sequencing was performed when necessary. RHD molecular analysis showed that 26.2% of donors had a variant RHD allele. It allowed the prediction of a partial D in 11% of cases. RHCE molecular analysis showed that 14.2% of donors had a variant RHCE allele or RH [RN or (C)ces] haplotype. A rare Rh phenotype associated with the loss of a high-prevalence antigen or partial RhCE antigens were predicted from RHCE molecular analysis in 1 (0.3%) and 17 (5%) cases, respectively. Systematic RHD and RHCE molecular analysis performed in blood donors of African origin provides transfusion-relevant information for individuals of African origin because of the frequency of variant RH alleles. RH molecular analysis may improve transfusion therapy of patients by allowing better donor and recipient matching, based not only on phenotypically matched red blood cell units, but also on units that are genetically matched with regards to RhCE variants.

  19. In silico identification of new putative pathogenic variants in the NEU1 sialidase gene affecting enzyme function and subcellular localization.

    PubMed

    Bonardi, Dario; Ravasio, Viola; Borsani, Giuseppe; d'Azzo, Alessandra; Bresciani, Roberto; Monti, Eugenio; Giacopuzzi, Edoardo

    2014-01-01

    The NEU1 gene is the first identified member of the human sialidases, glycohydrolitic enzymes that remove the terminal sialic acid from oligosaccharide chains. Mutations in NEU1 gene are causative of sialidosis (MIM 256550), a severe lysosomal storage disorder showing autosomal recessive mode of inheritance. Sialidosis has been classified into two subtypes: sialidosis type I, a normomorphic, late-onset form, and sialidosis type II, a more severe neonatal or early-onset form. A total of 50 causative mutations are reported in HGMD database, most of which are missense variants. To further characterize the NEU1 gene and identify new functionally relevant protein isoforms, we decided to study its genetic variability in the human population using the data generated by two large sequencing projects: the 1000 Genomes Project (1000G) and the NHLBI GO Exome Sequencing Project (ESP). Together these two datasets comprise a cohort of 7595 sequenced individuals, making it possible to identify rare variants and dissect population specific ones. By integrating this approach with biochemical and cellular studies, we were able to identify new rare missense and frameshift alleles in NEU1 gene. Among the 9 candidate variants tested, only two resulted in significantly lower levels of sialidase activity (p<0.05), namely c.650T>C and c.700G>A. These two mutations give rise to the amino acid substitutions p.V217A and p.D234N, respectively. NEU1 variants including either of these two amino acid changes have 44% and 25% residual sialidase activity when compared to the wild-type enzyme, reduced protein levels and altered subcellular localization. Thus they may represent new, putative pathological mutations resulting in sialidosis type I. The in silico approach used in this study has enabled the identification of previously unknown NEU1 functional alleles that are widespread in the population and could be tested in future functional studies.

  20. Identification and functional characterization of four TRPA1 variants in Apolygus lucorum (Meyer-Dür)

    USDA-ARS?s Scientific Manuscript database

    As signal integrators that respond to various physical and chemical stimuli, transient receptor potential (TRP) channels fulfil critical functional roles in the sensory systems of both vertebrate and invertebrate organisms. Here, four variants of TRP ankyrin 1 (TRPA1) were identified and cloned from...

  1. Large-scale identification of sequence variants impacting human transcription factor occupancy in vivo

    PubMed Central

    Maurano, Matthew T.; Haugen, Eric; Sandstrom, Richard; Vierstra, Jeff; Shafer, Anthony; Kaul, Rajinder; Stamatoyannopoulos, John A.

    2015-01-01

    The function of human regulatory regions depends exquisitely on their local genomic environment and cellular context, complicating experimental analysis of the expanding pool of common disease- and trait-associated variants that localize within regulatory DNA. We leverage allelically resolved genomic DNaseI footprinting data encompassing 166 individuals and 114 cell types to identify >60,000 common variants that directly impact transcription factor occupancy and regulatory DNA accessibility in vivo. The unprecedented scale of these data enable systematic analysis of the impact of sequence variation on transcription factor occupancy in vivo. We leverage this analysis to develop accurate models of variation affecting the recognition sites for diverse transcription factors, and apply these models to discriminate nearly 500,000 common regulatory variants likely to affect transcription factor occupancy across the human genome. The approach and results provide a novel foundation for analysis and interpretation of noncoding variation in complete human genomes, and for systems-level investigation of disease-associated variants. PMID:26502339

  2. The Identification of Splice Variants as Molecular Markers in Parkinson’s Disease

    DTIC Science & Technology

    2007-09-01

    diagnosed PD patients, who are not yet on dopamine therapy . The overall goal is to use splice variants as biomarkers to identify individuals at risk for PD...FosB-like proteins by electroconvulsive seizure and cocaine treatments. Mol Pharmacol. 48, 880-9. Chen, L., Cagniard, B., Mathews, T., Jones, S., Koh

  3. Identification of genomic variants putatively targeted by selection during dog domestication.

    PubMed

    Cagan, Alex; Blass, Torsten

    2016-01-12

    Dogs [Canis lupus familiaris] were the first animal species to be domesticated and continue to occupy an important place in human societies. Recent studies have begun to reveal when and where dog domestication occurred. While much progress has been made in identifying the genetic basis of phenotypic differences between dog breeds we still know relatively little about the genetic changes underlying the phenotypes that differentiate all dogs from their wild progenitors, wolves [Canis lupus]. In particular, dogs generally show reduced aggression and fear towards humans compared to wolves. Therefore, selection for tameness was likely a necessary prerequisite for dog domestication. With the increasing availability of whole-genome sequence data it is possible to try and directly identify the genetic variants contributing to the phenotypic differences between dogs and wolves. We analyse the largest available database of genome-wide polymorphism data in a global sample of dogs 69 and wolves 7. We perform a scan to identify regions of the genome that are highly differentiated between dogs and wolves. We identify putatively functional genomic variants that are segregating or at high frequency [> = 0.75 Fst] for alternative alleles between dogs and wolves. A biological pathways analysis of the genes containing these variants suggests that there has been selection on the 'adrenaline and noradrenaline biosynthesis pathway', well known for its involvement in the fight-or-flight response. We identify 11 genes with putatively functional variants fixed for alternative alleles between dogs and wolves. The segregating variants in these genes are strong candidates for having been targets of selection during early dog domestication. We present the first genome-wide analysis of the different categories of putatively functional variants that are fixed or segregating at high frequency between a global sampling of dogs and wolves. We find evidence that selection has been strongest

  4. Identification of Variant-Specific Functions of PIK3CA by Rapid Phenotyping of Rare Mutations

    PubMed Central

    Dogruluk, Turgut; Tsang, Yiu Huen; Espitia, Maribel; Chen, Fengju; Chen, Tenghui; Chong, Zechen; Appadurai, Vivek; Dogruluk, Armel; Eterovic, Agna Karina; Bonnen, Penelope E.; Creighton, Chad J.; Chen, Ken; Mills, Gordon B.; Scott, Kenneth L.

    2015-01-01

    Large-scale sequencing efforts are uncovering the complexity of cancer genomes, which are comprised of causal “driver” mutations that promote tumor progression along with many more pathologically-neutral “passenger” events. The majority of mutations, both in known cancer drivers and uncharacterized genes, are generally of low occurrence, highlighting the need to functionally annotate the long tail of infrequent mutations present in heterogeneous cancers. Here we describe a mutation assessment pipeline enabled by high-throughput engineering of molecularly-barcoded gene variant expression clones identified by tumor sequencing. We first used this platform to functionally assess tail mutations observed in PIK3CA, which encodes the catalytic subunit alpha of the phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) frequently mutated in cancer. Orthogonal screening for PIK3CA variant activity using in vitro and in vivo cell growth and transformation assays differentiated driver from passenger mutations, revealing that PIK3CA variant activity correlates imperfectly with its mutation frequency across breast cancer populations. While PIK3CA mutations with frequencies above 5% were significantly more oncogenic than wild-type in all assays, mutations occurring at 0.07 – 5.0% included those with and without oncogenic activities that ranged from weak to strong in at least one assay. Proteomic profiling coupled with therapeutic sensitivity assays on PIK3CA variant-expressing cell models revealed variant-specific activation of PI3K signaling as well as other pathways that include the MEK1/2 module of Mitogen-Activated Protein (MAP) Kinase pathway. Our data indicate that cancer treatments will need to increasingly consider the functional relevance of specific mutations in driver genes rather than considering all mutations in drivers as equivalent. PMID:26627007

  5. Identification and functional characterization of NODAL rare variants in heterotaxy and isolated cardiovascular malformations

    PubMed Central

    Mohapatra, Bhagyalaxmi; Casey, Brett; Li, Hua; Ho-Dawson, Trang; Smith, Liana; Fernbach, Susan D.; Molinari, Laura; Niesh, Stephen R.; Jefferies, John Lynn; Craigen, William J.; Towbin, Jeffrey A.; Belmont, John W.; Ware, Stephanie M.

    2009-01-01

    NODAL and its signaling pathway are known to play a key role in specification and patterning of vertebrate embryos. Mutations in several genes encoding components of the NODAL signaling pathway have previously been implicated in the pathogenesis of human left–right (LR) patterning defects. Therefore, NODAL, a member of TGF-β superfamily of developmental regulators, is a strong candidate to be functionally involved in congenital LR axis patterning defects or heterotaxy. Here we have investigated whether variants in NODAL are present in patients with heterotaxy and/or isolated cardiovascular malformations (CVM) thought to be caused by abnormal heart tube looping. Analysis of a large cohort of cases (n = 269) affected with either classic heterotaxy or looping CVM revealed four different missense variants, one in-frame insertion/deletion and two conserved splice site variants in 14 unrelated subjects (14/269, 5.2%). Although similar with regard to other associated defects, individuals with the NODAL mutations had a significantly higher occurrence of pulmonary valve atresia (P = 0.001) compared with cases without a detectable NODAL mutation. Functional analyses demonstrate that the missense variant forms of NODAL exhibit significant impairment of signaling as measured by decreased Cripto (TDGF-1) co-receptor-mediated activation of artificial reporters. Expression of these NODAL proteins also led to reduced induction of Smad2 phosphorylation and impaired Smad2 nuclear import. Taken together, these results support a role for mutations and rare deleterious variants in NODAL as a cause for sporadic human LR patterning defects. PMID:19064609

  6. Orientation and mobility training for partially-sighted older adults using an identification cane: a systematic review

    PubMed Central

    Ballemans, Judith; Kempen, Gertrudis IJM; Zijlstra, GA Rixt

    2011-01-01

    Objective: This study aimed to provide an overview of the development, content, feasibility, and effectiveness of existing orientation and mobility training programmes in the use of the identification cane. Data sources: A systematic bibliographic database search in PubMed, PsychInfo, ERIC, CINAHL and the Cochrane Library was performed, in combination with the expert consultation (n = 42; orientation and mobility experts), and hand-searching of reference lists. Review methods: Selection criteria included a description of the development, the content, the feasibility, or the effectiveness of orientation and mobility training in the use of the identification cane. Two reviewers independently agreed on eligibility and methodological quality. A narrative/qualitative data analysis method was applied to extract data from obtained documents. Results: The sensitive database search and hand-searching of reference lists revealed 248 potentially relevant abstracts. None met the eligibility criteria. Expert consultation resulted in the inclusion of six documents in which the information presented on the orientation and mobility training in the use of the identification cane was incomplete and of low methodological quality. Conclusion: Our review of the literature showed a lack of well-described protocols and studies on orientation and mobility training in identification cane use. PMID:21795405

  7. Identification and analysis of two splice variants of human G2A generated by alternative splicing.

    PubMed

    Ogawa, Ai; Obinata, Hideru; Hattori, Tomoyasu; Kishi, Mikiko; Tatei, Kazuaki; Ishikawa, Osamu; Izumi, Takashi

    2010-02-01

    G2A is a G protein-coupled receptor that can be induced by various stressors. G2A is reported to have proton-sensing activity that mediates intracellular inositol phosphate (IP) accumulation with decreasing pH. We previously showed that G2A is also activated by some oxidized free fatty acids such as 9-hydroxyoctadecadienoic acid (9-HODE). In this study, we identified a novel alternative splice variant of G2A (G2A-b) that has a partially different N terminus compared with the G2A originally reported (G2A-a). The two splice variants of G2A show similar tissue distributions, but G2A-b is expressed more abundantly. There was no difference between the two variants in 9-HODE-induced cellular responses, such as intracellular calcium mobilization and GDP/GTP exchange of Galpha protein, and in proton-sensitive IP accumulation. However, G2A-b showed a higher basal activity in terms of IP accumulation. Mutagenesis study revealed that the difference in the basal activity is attributable to the K7 residue that exists only in G2A-a. We further demonstrated that an R42A mutation largely impaired both the basal and proton-sensing activities, but did not affect the 9-HODE-induced intracellular calcium increase. Taken together, we found an additional novel G2A variant (G2A-b) that is the major transcript with functional response to ligand stimulation as well as G2A-a, and succeeded in discriminating proton-sensing and oxidized fatty acid-sensing activities of G2A.

  8. Doublet N-Terminal Oriented Proteomics for N-Terminomics and Proteolytic Processing Identification.

    PubMed

    Westermann, Benoit; Jacome, Alvaro Sebastian Vaca; Rompais, Magali; Carapito, Christine; Schaeffer-Reiss, Christine

    2017-01-01

    The study of the N-terminome and the precise identification of proteolytic processing events are key in biology. Dedicated methodologies have been developed as the comprehensive characterization of the N-terminome can hardly be achieved by standard proteomics methods. In this context, we have set up a trimethoxyphenyl phosphonium (TMPP) labeling approach that allows the characterization of both N-terminal and internal digestion peptides in a single experiment. This latter point is a major advantage of our strategy as most N-terminomics methods rely on the enrichment of N-terminal peptides and thus exclude internal peptides.We have implemented a double heavy/light TMPP labeling and an automated data validation workflow that make our doublet N-terminal oriented proteomics (dN-TOP) strategy efficient for high-throughput N-terminome analysis.

  9. Identification of splicing variants of Frabin with partly different functions and tissue distribution.

    PubMed

    Ikeda, W; Nakanishi, H; Takekuni, K; Itoh, S; Takai, Y

    2001-09-07

    Frabin is a GDP/GTP exchange protein for Cdc42 small G protein with actin filament-binding activity. Frabin consists of the actin filament-binding domain, the Dbl homology domain, the first pleckstrin homology domain, the FYVE-finger domain, and the second pleckstrin homology domain in this order from the N-terminus. Frabin forms filopodia through direct activation of Cdc42 and lamellipodia through indirect activation of Rac small G protein. We isolated here two smaller splicing variants of frabin and named the original one, middle-size one, and smallest one frabin-alpha, -beta, and -gamma, respectively. Frabin-beta lacked the second pleckstrin homology domain and frabin-gamma lacked the FYVE-finger domain and the second pleckstrin homology domain. These three variants were expressed in all of the tissues examined but their expression levels are different depending on tissues. In L fibroblasts, all the three variants formed filopodia. As to lamellipodia, frabin-alpha formed them; frabin-beta formed them to a small extent; and frabin-gamma did not. In MDCK epithelial cells, frabin-alpha formed microspikes but frabin-beta or -gamma did not. Copyright 2001 Academic Press.

  10. [Identification of variants in LMF1 gene associated with primary hypertriglyceridemia].

    PubMed

    Lamiquiz-Moneo, Itziar; Bea, Ana M; Mateo-Gallego, Rocío; Baila-Rueda, Lucía; Cenarro, Ana; Pocoví, Miguel; Civeira, Fernando; de Castro-Orós, Isabel

    2015-01-01

    The majority of severe primary hypertriglyceridemia (HTG) are diagnosed in adults, and their molecular bases have not yet been fully defined. The promoter, coding regions and intron-exon boundaries of LMF1 were sequenced in 112 patients with severe primary hipertrigliceridemia (defined as TG above 500mg/dl). Five patients (4.46%) were carriers of four rare variants in the LMF1 gene associated with HTG, which participate in lipoprotein lipase (LpL) function. Also, we have identified two common variants, c.194-28 T>G and c.729+18C>G that were associated with HTG, with a different allelic frequency to that observed in the general population. A bioinformatic analysis of all found variants was conducted, defining the following as potentially harmful: p.Arg364Gln, p.Arg451Trp, p.Pro562Arg and p.Leu85Leu. Our results suggest that LMF1 mutations are involved in a substantial proportion of cases with severe HTG, putting together the moderate-aggressive effect of rare mutations with polymorphisms classically associated with this disease.

  11. Identification of a novel alternative splicing variant of hemocyanin from shrimp Litopenaeus vannamei.

    PubMed

    Zhao, Shan; Lu, Xin; Zhang, Yueling; Zhao, Xianliang; Zhong, Mingqi; Li, Shengkang; Lun, Jingsheng

    2013-01-01

    Recent evidences suggest that invertebrates express families of immune molecules with high levels of sequence diversity. Hemocyanin is an important non-specific immune molecule present in the hemolymph of both mollusks and arthropods. In the present study, we characterized a novel alternative splicing variant of hemocyanin (cHE1) from Litopenaeus vannamei that produced mRNA transcript of 2579 bp in length. The isoform contained two additional sequences of 296 and 267 bp in the 5'- and 3'-terminus respectively, in comparison to that of wild type hemocyanin (cHE). Sequence of cHE1 shows 100% identity to that of hemocyanin genomic DNA (HE, which does not form an open reading frame), suggesting that cHE1 might be an alternative splicing variant due to intron retention. Moreover, cHE1 could be detected by RT-PCR from five tissues (heart, gill, stomach, intestine and brain), and from shrimps at stages from nauplius to mysis larva. Further, cHE1 mRNA transcripts were significantly increased in hearts after 12h of infection with Vibrio parahemolyticus or poly I: C, while no significant difference in the transcript levels of hepatopancreas cHE was detected in the pathogen-treated shrimps during the period. In summary, these studies suggested a novel splicing variant of hemocyanin in shrimp, which might be involved in shrimp resistance to pathogenic infection.

  12. Pathogenesis of coronary artery disease: focus on genetic risk factors and identification of genetic variants

    PubMed Central

    Sayols-Baixeras, Sergi; Lluís-Ganella, Carla; Lucas, Gavin; Elosua, Roberto

    2014-01-01

    Coronary artery disease (CAD) is the leading cause of death and disability worldwide, and its prevalence is expected to increase in the coming years. CAD events are caused by the interplay of genetic and environmental factors, the effects of which are mainly mediated through cardiovascular risk factors. The techniques used to study the genetic basis of these diseases have evolved from linkage studies to candidate gene studies and genome-wide association studies. Linkage studies have been able to identify genetic variants associated with monogenic diseases, whereas genome-wide association studies have been more successful in determining genetic variants associated with complex diseases. Currently, genome-wide association studies have identified approximately 40 loci that explain 6% of the heritability of CAD. The application of this knowledge to clinical practice is challenging, but can be achieved using various strategies, such as genetic variants to identify new therapeutic targets, personal genetic information to improve disease risk prediction, and pharmacogenomics. The main aim of this narrative review is to provide a general overview of our current understanding of the genetics of coronary artery disease and its potential clinical utility. PMID:24520200

  13. Identification of two novel chicken GPR133 variants and their expression in different tissues.

    PubMed

    Tian, Kai; Xiao, Qihai; Zhang, Xueyou; Lan, Xi; Zhao, Xiaoling; Wang, Yan; Li, Diyan; Yin, Huadong; Ye, Lin; Zhu, Qing

    2017-06-14

    GPR133 (G protein-coupled receptor 133) plays significant roles in various physiological processes. Alternatively splicing (AS) variants of GPR133 in many species have been predicted in multiple databases, but there is no available information about the AS events of chicken GPR133 (cGPR133). In the present study, two chicken GPR133 variants, cGPR133-va and cGPR133-vb, were identified by a combination of reverse transcription PCR (RT-PCR) and rapid amplification of cDNA 5'-ends (5' RACE). Sequence analysis shows that cGPR133-va and cGPR133-vb are resulting from different AS modules and their sequences are predicted to encode two distinct putative proteins, respectively. Quantitative real-time PCR (qRT-PCR) analysis revealed that cGPR133-va and cGPR133-vb are widely expressed in different tissues, while exhibiting specific expression profile. Altogether, our results first demonstrate the existence of novel cGPR133 variants and illustrate its transcriptional diversity and their widespread distribution, which provides a foundation for the further research of GPR133.

  14. Identification and characterization of the variants of metastasis-associated protein 1 generated following alternative splicing.

    PubMed

    Yaguchi, Masahiro; Wada, Yoriko; Toh, Yasushi; Iguchi, Haruo; Kono, Akira; Matsusue, Kimihiko; Takiguchi, Soichi

    2005-12-30

    The metastasis-associated gene 1 (mta1) was identified initially in rat highly metastatic cancer cell lines and found to be a component of the nucleosome remodeling and histone deacetylase (NuRD) complex. The gene for mouse mta1 was screened and its genomic structure was determined. It consists of 21 exons spanning 40 kb of genomic DNA. The full-length mouse Mta1 cDNA contained a 2145 nucleotide open reading frame encoding 715 amino acids. In addition to the full-length cDNA, several alternative splicing variants were found. Some differences in the splicing variants found were observed among various mouse organs and cells examined by the semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR). The cDNAs of the splicing variants were inserted into green fluorescent protein (GFP) expression vector and the subcellular localization of the GFP-Mta1 fusion proteins were analyzed. Knowledge of the Mta1 gene expression pattern will be useful in better understanding its functional diversity.

  15. Learning to Be (In)variant: Combining Prior Knowledge and Experience to Infer Orientation Invariance in Object Recognition.

    PubMed

    Austerweil, Joseph L; Griffiths, Thomas L; Palmer, Stephen E

    2016-12-21

    How does the visual system recognize images of a novel object after a single observation despite possible variations in the viewpoint of that object relative to the observer? One possibility is comparing the image with a prototype for invariance over a relevant transformation set (e.g., translations and dilations). However, invariance over rotations (i.e., orientation invariance) has proven difficult to analyze, because it applies to some objects but not others. We propose that the invariant transformations of an object are learned by incorporating prior expectations with real-world evidence. We test this proposal by developing an ideal learner model for learning invariance that predicts better learning of orientation dependence when prior expectations about orientation are weak. This prediction was supported in two behavioral experiments, where participants learned the orientation dependence of novel images using feedback from solving arithmetic problems.

  16. Identification of Rare Causal Variants in Sequence-Based Studies: Methods and Applications to VPS13B, a Gene Involved in Cohen Syndrome and Autism

    PubMed Central

    De Rubeis, Silvia; McCallum, Kenneth; Buxbaum, Joseph D.

    2014-01-01

    Pinpointing the small number of causal variants among the abundant naturally occurring genetic variation is a difficult challenge, but a crucial one for understanding precise molecular mechanisms of disease and follow-up functional studies. We propose and investigate two complementary statistical approaches for identification of rare causal variants in sequencing studies: a backward elimination procedure based on groupwise association tests, and a hierarchical approach that can integrate sequencing data with diverse functional and evolutionary conservation annotations for individual variants. Using simulations, we show that incorporation of multiple bioinformatic predictors of deleteriousness, such as PolyPhen-2, SIFT and GERP++ scores, can improve the power to discover truly causal variants. As proof of principle, we apply the proposed methods to VPS13B, a gene mutated in the rare neurodevelopmental disorder called Cohen syndrome, and recently reported with recessive variants in autism. We identify a small set of promising candidates for causal variants, including two loss-of-function variants and a rare, homozygous probably-damaging variant that could contribute to autism risk. PMID:25502226

  17. Mohs Mapping Fidelity: Optimizing Orientation, Accuracy, and Tissue Identification in Mohs Surgery.

    PubMed

    Li, Janet Y; Silapunt, Sirunya; Migden, Michael R; McGinness, Jamie L; Nguyen, Tri H

    2017-06-23

    Mohs micrographic surgery (MMS) is a highly effective process that requires consistent accuracy in resection, mapping, and histologic interpretation. Although the general sequence in MMS is similar, there are numerous variations among Mohs surgeons as to how this process is performed. This article aims to review the process of MMS, with the intent to identify and mitigate the potential errors at each step. Existing variations will be discussed and protocols offered to minimize error and optimize accuracy. A Pubmed search was performed for publications on methods of tissue mapping, orienting, and processing in MMS. Our literature review highlights various techniques for tissue orientation, transfer, flattening, inking, mapping, and processing of later stages and multiple specimens. We discuss our system, which reduces error during tissue transfer, tissue identification in vivo and ex vivo, and tissue flattening. Furthermore, we discuss adaptations to increase the accuracy during reexcisions of subsequent Mohs layers. Variations in MMS reflects the diverse training and creativity among Mohs surgeons. Unless potential errors are addressed, however, false negatives will occur and undermine the superior cure rate of MMS.

  18. [Cotton identification and extraction using near infrared sensor and object-oriented spectral segmentation technique].

    PubMed

    Deng, Jin-Song; Shi, Yuan-Yuan; Chen, Li-Su; Wang, Ke; Zhu, Jin-Xia

    2009-07-01

    The real-time, effective and reliable method of identifying crop is the foundation of scientific management for crop in the precision agriculture. It is also one of the key techniques for the precision agriculture. However, this expectation cannot be fulfilled by the traditional pixel-based information extraction method with respect to complicated image processing and accurate objective identification. In the present study, visible-near infrared image of cotton was acquired using high-resolution sensor. Object-oriented segmentation technique was performed on the image to produce image objects and spatial/spectral features of cotton. Afterwards, nearest neighbor classifier integrated the spectral, shape and topologic information of image objects to precisely identify cotton according to various features. Finally, 300 random samples and an error matrix were applied to undertake the accuracy assessment of identification. Although errors and confusion exist, this method shows satisfying results with an overall accuracy of 96.33% and a KAPPA coefficient of 0.926 7, which can meet the demand of automatic management and decision-making in precision agriculture.

  19. Antagonistic lactic acid bacteria isolated from goat milk and identification of a novel nisin variant Lactococcus lactis

    PubMed Central

    2014-01-01

    Background The raw goat milk microbiota is considered a good source of novel bacteriocinogenic lactic acid bacteria (LAB) strains that can be exploited as an alternative for use as biopreservatives in foods. The constant demand for such alternative tools justifies studies that investigate the antimicrobial potential of such strains. Results The obtained data identified a predominance of Lactococcus and Enterococcus strains in raw goat milk microbiota with antimicrobial activity against Listeria monocytogenes ATCC 7644. Enzymatic assays confirmed the bacteriocinogenic nature of the antimicrobial substances produced by the isolated strains, and PCR reactions detected a variety of bacteriocin-related genes in their genomes. Rep-PCR identified broad genetic variability among the Enterococcus isolates, and close relations between the Lactococcus strains. The sequencing of PCR products from nis-positive Lactococcus allowed the identification of a predicted nisin variant not previously described and possessing a wide inhibitory spectrum. Conclusions Raw goat milk was confirmed as a good source of novel bacteriocinogenic LAB strains, having identified Lactococcus isolates possessing variations in their genomes that suggest the production of a nisin variant not yet described and with potential for use as biopreservatives in food due to its broad spectrum of action. PMID:24521354

  20. Identification of a capsular variant and characterization of capsular acetylation in Klebsiella pneumoniae PLA-associated type K57

    PubMed Central

    Hsu, Chun-Ru; Liao, Chun-Hsing; Lin, Tzu-Lung; Yang, Han-Ru; Yang, Feng-Ling; Hsieh, Pei-Fang; Wu, Shih-Hsiung; Wang, Jin-Town

    2016-01-01

    Klebsiella pneumoniae can cause community-acquired pyogenic liver abscess (PLA). Capsular polysaccharide (CPS) is important for its virulence. Among 79 capsular (K) types discovered thus far, K57 is often associated with PLA. Here, we report the identification of a K57 variant. Cps gene locus sequencing revealed differences between the K57 reference strain 4425/51 (Ref-K57) and a variant, the PLA isolate A1142. While Ref-K57 cps contained orf13 encoding a putative acetyltransferase, the insertion of a putative transposase-encoding gene at this position was detected in A1142. This variation was detected in other K57 clinical strains. Biochemical analyses indicated that A1142 was deficient in CPS acetylation. Genetic replacement and complementation verified that orf13 was responsible for CPS acetylation. Acetylation increased CPS immunoreactivity to antiserum and enhanced K. pneumoniae induction of pro-inflammatory cytokines through JNK and MAPK signaling. While acetylation diminished the serum resistance of bacteria, it promoted adhesion to intestinal epithelial cells possibly via increasing production of type I fimbriae. In conclusion, acetylation-mediated capsular variation in K57 was observed. Capsular acetylation contributed to the variety and antigenic diversity of CPS, influenced its biological activities, and was involved in K. pneumoniae-host interactions. These findings have implications for vaccine design and pathogenicity of K. pneumoniae. PMID:27550826

  1. Antagonistic lactic acid bacteria isolated from goat milk and identification of a novel nisin variant Lactococcus lactis.

    PubMed

    Perin, Luana Martins; Nero, Luís Augusto

    2014-02-12

    The raw goat milk microbiota is considered a good source of novel bacteriocinogenic lactic acid bacteria (LAB) strains that can be exploited as an alternative for use as biopreservatives in foods. The constant demand for such alternative tools justifies studies that investigate the antimicrobial potential of such strains. The obtained data identified a predominance of Lactococcus and Enterococcus strains in raw goat milk microbiota with antimicrobial activity against Listeria monocytogenes ATCC 7644. Enzymatic assays confirmed the bacteriocinogenic nature of the antimicrobial substances produced by the isolated strains, and PCR reactions detected a variety of bacteriocin-related genes in their genomes. Rep-PCR identified broad genetic variability among the Enterococcus isolates, and close relations between the Lactococcus strains. The sequencing of PCR products from nis-positive Lactococcus allowed the identification of a predicted nisin variant not previously described and possessing a wide inhibitory spectrum. Raw goat milk was confirmed as a good source of novel bacteriocinogenic LAB strains, having identified Lactococcus isolates possessing variations in their genomes that suggest the production of a nisin variant not yet described and with potential for use as biopreservatives in food due to its broad spectrum of action.

  2. Identification of five novel STAR variants in ten Chinese patients with congenital lipoid adrenal hyperplasia.

    PubMed

    Huang, Zhuo; Ye, Jun; Han, Lianshu; Qiu, Wenjuan; Zhang, Huiwen; Yu, Yongguo; Liang, Lili; Gong, Zhuwen; Gu, Xuefan

    2016-04-01

    Congenital lipoid adrenal hyperplasia (CLAH) is a rare autosomal recessive disorder caused by defective synthesis of all steroids. This disorder is characterized by 46,XY sex reversal, skin hyperpigmentation, early-onset adrenal crisis and enlarged adrenal with fatty accumulation. CLAH is caused by mutations in the STAR gene. The clinical features and STAR gene mutation spectrum of a large cohort of Chinese patients with CLAH were not reported previously. We performed clinical retrospective review and genetic analysis of the STAR gene in ten unrelated Chinese phenotypic female patients who were clinically diagnosed with CLAH and followed up in our hospital from 2006 to 2015. All ten patients, including two 46,XY females and eight 46,XX females, presented skin hyperpigmentation and early salt-wasting episode, and showed normal growth and development after steroid replacement treatment. Totally 20 mutant alleles containing 11 different STAR gene mutations were identified in these ten patients, including five novel variants (two missense and three null variants), all predicted to be pathogenic in bioinformatics analysis, and six mutations described in previous literature. Among these 11 mutations, a reported mutation c.772C>T and a novel variant c.707_708delinsCTT were most frequent, accounting for 35% and 15% of the total mutant alleles, respectively. This is the first report of a large Chinese cohort with CLAH, presenting the mutation spectrum of the STAR gene and two possible founder mutations in the Chinese population, which may contribute to better genetic counseling and prenatal diagnosis. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Identification of novel functional sequence variants in the gene for peptidase inhibitor 3

    PubMed Central

    Chowdhury, Mahboob A; Kuivaniemi, Helena; Romero, Roberto; Edwin, Samuel; Chaiworapongsa, Tinnakorn; Tromp, Gerard

    2006-01-01

    Background Peptidase inhibitor 3 (PI3) inhibits neutrophil elastase and proteinase-3, and has a potential role in skin and lung diseases as well as in cancer. Genome-wide expression profiling of chorioamniotic membranes revealed decreased expression of PI3 in women with preterm premature rupture of membranes. To elucidate the molecular mechanisms contributing to the decreased expression in amniotic membranes, the PI3 gene was searched for sequence variations and the functional significance of the identified promoter variants was studied. Methods Single nucleotide polymorphisms (SNPs) were identified by direct sequencing of PCR products spanning a region from 1,173 bp upstream to 1,266 bp downstream of the translation start site. Fourteen SNPs were genotyped from 112 and nine SNPs from 24 unrelated individuals. Putative transcription factor binding sites as detected by in silico search were verified by electrophoretic mobility shift assay (EMSA) using nuclear extract from Hela and amnion cell nuclear extract. Deviation from Hardy-Weinberg equilibrium (HWE) was tested by χ2 goodness-of-fit test. Haplotypes were estimated using expectation maximization (EM) algorithm. Results Twenty-three sequence variations were identified by direct sequencing of polymerase chain reaction (PCR) products covering 2,439 nt of the PI3 gene (-1,173 nt of promoter sequences and all three exons). Analysis of 112 unrelated individuals showed that 20 variants had minor allele frequencies (MAF) ranging from 0.02 to 0.46 representing "true polymorphisms", while three had MAF ≤ 0.01. Eleven variants were in the promoter region; several putative transcription factor binding sites were found at these sites by database searches. Differential binding of transcription factors was demonstrated at two polymorphic sites by electrophoretic mobility shift assays, both in amniotic and HeLa cell nuclear extracts. Differential binding of the transcription factor GATA1 at -689C>G site was confirmed by a

  4. Robust HCV Genotype 3a Infectious Cell Culture System Permits Identification of Escape Variants With Resistance to Sofosbuvir.

    PubMed

    Ramirez, Santseharay; Mikkelsen, Lotte S; Gottwein, Judith M; Bukh, Jens

    2016-11-01

    Direct-acting antivirals (DAAs) effectively eradicate chronic hepatitis C virus (HCV) infection, although HCV genotype 3a is less responsive to these drugs. We aimed to develop genotype 3a infectious cultures and study the effects of inhibitors of NS5A and NS5B and resistance to sofosbuvir-the only nucleotide analog approved for treatment of chronic HCV infection. The developed HCV genotype 3a full-length genome (DBN3a), with a strain-DBN coding sequence, modified NS5B consensus sequence, pS52 untranslated regions, and coding mutations from a culture-efficient JFH1-based core-NS5A (DBN) recombinant, was transfected into Huh7.5 cells. The efficacy of selected DAAs was determined in dose-response assays, in which the number of HCV-infected cells was measured after incubation with different concentrations of the specific DAA. Long-term culture of infected Huh7.5 cells with increasing concentrations of sofosbuvir was used to promote selection of HCV-resistant variants. We engineered a DBN3a variant with 17 substitutions (DBN3acc) that had replication and propagation kinetics in Huh7.5 cells comparable with prototype J6/JFH1. The adaptive mutations also produced culture-efficient DBN-based recombinants with NS5B from HCV genotype 3a strains S52 and DH11. Compared with genotype 1a, genotype 3a was less sensitive to daclatasvir, ledipasvir, and elbasvir, but equally sensitive to ombitasvir, velpatasvir, beclabuvir, dasabuvir, MK-3682, and sofosbuvir. Exposure of Huh7.5 cells infected with DBN3a to sofosbuvir led to identification of an escape variant with substitutions in NS5B, including the resistance-associated substitution S282T. This variant showed increased infectivity of Huh7.5 cells, compared with DBN3a, and was genetically stable in cell cultures without sofosbuvir. Sofosbuvir, MK-3682, dasabuvir, or combinations of sofosbuvir and ledipasvir or sofosbuvir and velpatasvir had decreased efficacy against infection with the DBN3a sofosbuvir escape variant. We

  5. Identification and Characterization of Porcine Kobuvirus Variant Isolated from Suckling Piglet in Gansu Province, China

    PubMed Central

    Fan, Shengtao; Sun, Heting; Ying, Ying; Gao, Xiaolong; Wang, Zheng; Yu, Yicong; Li, Yuanguo; Wang, Tiecheng; Yu, Zhijun; Yang, Songtao; Zhao, Yongkun; Qin, Chuan; Gao, Yuwei; Xia, Xianzhu

    2013-01-01

    Kobuviruses comprise three species, the Aichivirus A, Aichivirus B, and Aichivirus C (porcine kobuvirus). Porcine kobuvirus is endemic to pig farms and is not restricted geographically but, rather, is distributed worldwide. The complete genomic sequences of four porcine kobuvirus strains isolated during a diarrhea outbreak in piglets in the Gansu province of China were determined. Two of these strains exhibited variations relative to the traditional strains. The potential 3C/3D cleavage sites of the variant strains were Q/C, which differed from the Q/S in the traditional porcine kobuvirus genome. A 90-nucleotide deletion in the 2B protein and a single nucleotide insertion in the 3′UTR were found in the variant strains. The VP1 regions of all four porcine kobuviruses in our study were highly variable (81%–86%). Ten common amino acid mutations were found specifically at certain positions within the VP1 region. Significant recombination sites were identified using SimPlot scans of whole genome sequences. Porcine kobuviruses were also detected in pig serum, indicating that the virus can escape the gastrointestinal tract and travel to the circulatory system. These findings suggest that mutations and recombination events may have contributed to the high level of genetic diversity of porcine kobuviruses and serve as a driving force in its evolution. PMID:24145960

  6. Identification of a splice variant of mouse TRPA1 that regulates TRPA1 activity.

    PubMed

    Zhou, Yiming; Suzuki, Yoshiro; Uchida, Kunitoshi; Tominaga, Makoto

    2013-01-01

    Transient receptor potential ankyrin 1 (TRPA1) protein is a nonselective cation channel. Although many studies suggest that TRPA1 is involved in inflammatory and neuropathic pain, its mechanism remains unclear. Here we identify an alternative splice variant of the mouse Trpa1 gene. TRPA1a (full-length) and TRPA1b (splice variant) physically interact with each other and TRPA1b increases the expression of TRPA1a in the plasma membrane. TRPA1a and TRPA1b co-expression significantly increases current density in response to different agonists without affecting their single-channel conductance. Exogenous overexpression of Trpa1b gene in wild-type and TRPA1KO DRG neurons also increases TRPA1a-mediated AITC responses. Moreover, expression levels of Trpa1a and Trpa1b mRNAs change dynamically in two pain models (complete Freund's adjuvant-induced inflammatory pain and partial sciatic nerve ligation-induced neuropathic pain models). These results suggest that TRPA1 may be regulated through alternative splicing under these pathological conditions.

  7. Identification of common variants associated with human hippocampal and intracranial volumes

    PubMed Central

    Stein, Jason L; Medland, Sarah E; Vasquez, Alejandro Arias; Hibar, Derrek P; Senstad, Rudy E; Winkler, Anderson M; Toro, Roberto; Appel, Katja; Bartecek, Richard; Bergmann, Ørjan; Bernard, Manon; Brown, Andrew A; Cannon, Dara M; Chakravarty, M Mallar; Christoforou, Andrea; Domin, Martin; Grimm, Oliver; Hollinshead, Marisa; Holmes, Avram J; Homuth, Georg; Hottenga, Jouke-Jan; Langan, Camilla; Lopez, Lorna M; Hansell, Narelle K; Hwang, Kristy S; Kim, Sungeun; Laje, Gonzalo; Lee, Phil H; Liu, Xinmin; Loth, Eva; Lourdusamy, Anbarasu; Mattingsdal, Morten; Mohnke, Sebastian; Maniega, Susana Muñoz; Nho, Kwangsik; Nugent, Allison C; O’Brien, Carol; Papmeyer, Martina; Pütz, Benno; Ramasamy, Adaikalavan; Rasmussen, Jerod; Rijpkema, Mark; Risacher, Shannon L; Roddey, J Cooper; Rose, Emma J; Ryten, Mina; Shen, Li; Sprooten, Emma; Strengman, Eric; Teumer, Alexander; Trabzuni, Daniah; Turner, Jessica; van Eijk, Kristel; van Erp, Theo G M; van Tol, Marie-Jose; Wittfeld, Katharina; Wolf, Christiane; Woudstra, Saskia; Aleman, Andre; Alhusaini, Saud; Almasy, Laura; Binder, Elisabeth B; Brohawn, David G; Cantor, Rita M; Carless, Melanie A; Corvin, Aiden; Czisch, Michael; Curran, Joanne E; Davies, Gail; de Almeida, Marcio A A; Delanty, Norman; Depondt, Chantal; Duggirala, Ravi; Dyer, Thomas D; Erk, Susanne; Fagerness, Jesen; Fox, Peter T; Freimer, Nelson B; Gill, Michael; Göring, Harald H H; Hagler, Donald J; Hoehn, David; Holsboer, Florian; Hoogman, Martine; Hosten, Norbert; Jahanshad, Neda; Johnson, Matthew P; Kasperaviciute, Dalia; Kent, Jack W; Kochunov, Peter; Lancaster, Jack L; Lawrie, Stephen M; Liewald, David C; Mandl, René; Matarin, Mar; Mattheisen, Manuel; Meisenzahl, Eva; Melle, Ingrid; Moses, Eric K; Mühleisen, Thomas W; Nauck, Matthias; Nöthen, Markus M; Olvera, Rene L; Pandolfo, Massimo; Pike, G Bruce; Puls, Ralf; Reinvang, Ivar; Rentería, Miguel E; Rietschel, Marcella; Roffman, Joshua L; Royle, Natalie A; Rujescu, Dan; Savitz, Jonathan; Schnack, Hugo G; Schnell, Knut; Seiferth, Nina; Smith, Colin; Steen, Vidar M; Valdés Hernández, Maria C; Van den Heuvel, Martijn; van der Wee, Nic J; Van Haren, Neeltje E M; Veltman, Joris A; Völzke, Henry; Walker, Robert; Westlye, Lars T; Whelan, Christopher D; Agartz, Ingrid; Boomsma, Dorret I; Cavalleri, Gianpiero L; Dale, Anders M; Djurovic, Srdjan; Drevets, Wayne C; Hagoort, Peter; Hall, Jeremy; Heinz, Andreas; Jack, Clifford R; Foroud, Tatiana M; Le Hellard, Stephanie; Macciardi, Fabio; Montgomery, Grant W; Poline, Jean Baptiste; Porteous, David J; Sisodiya, Sanjay M; Starr, John M; Sussmann, Jessika; Toga, Arthur W; Veltman, Dick J; Walter, Henrik; Weiner, Michael W; Bis, Joshua C; Ikram, M Arfan; Smith, Albert V; Gudnason, Vilmundur; Tzourio, Christophe; Vernooij, Meike W; Launer, Lenore J; DeCarli, Charles; Seshadri, Sudha; Andreassen, Ole A; Apostolova, Liana G; Bastin, Mark E; Blangero, John; Brunner, Han G; Buckner, Randy L; Cichon, Sven; Coppola, Giovanni; de Zubicaray, Greig I; Deary, Ian J; Donohoe, Gary; de Geus, Eco J C; Espeseth, Thomas; Fernández, Guillén; Glahn, David C; Grabe, Hans J; Hardy, John; Hulshoff Pol, Hilleke E; Jenkinson, Mark; Kahn, René S; McDonald, Colm; McIntosh, Andrew M; McMahon, Francis J; McMahon, Katie L; Meyer-Lindenberg, Andreas; Morris, Derek W; Müller-Myhsok, Bertram; Nichols, Thomas E; Ophoff, Roel A; Paus, Tomas; Pausova, Zdenka; Penninx, Brenda W; Potkin, Steven G; Sämann, Philipp G; Saykin, Andrew J; Schumann, Gunter; Smoller, Jordan W; Wardlaw, Joanna M; Weale, Michael E; Martin, Nicholas G; Franke, Barbara; Wright, Margaret J; Thompson, Paul M

    2013-01-01

    Identifying genetic variants influencing human brain structures may reveal new biological mechanisms underlying cognition and neuropsychiatric illness. The volume of the hippocampus is a biomarker of incipient Alzheimer’s disease1,2 and is reduced in schizophrenia3, major depression4 and mesial temporal lobe epilepsy5. Whereas many brain imaging phenotypes are highly heritable6,7, identifying and replicating genetic influences has been difficult, as small effects and the high costs of magnetic resonance imaging (MRI) have led to underpowered studies. Here we report genome-wide association meta-analyses and replication for mean bilateral hippocampal, total brain and intracranial volumes from a large multinational consortium. The intergenic variant rs7294919 was associated with hippocampal volume (12q24.22; N = 21,151; P = 6.70 × 10−16) and the expression levels of the positional candidate gene TESC in brain tissue. Additionally, rs10784502, located within HMGA2, was associated with intracranial volume (12q14.3; N = 15,782; P = 1.12 × 10−12). We also identified a suggestive association with total brain volume at rs10494373 within DDR2 (1q23.3; N = 6,500; P = 5.81 × 10−7). PMID:22504417

  8. Identification of cystic fibrosis variants by polymerase chain reaction/oligonucleotide ligation assay.

    PubMed

    Schwartz, Karen M; Pike-Buchanan, Lisa L; Muralidharan, Kasinathan; Redman, Joy B; Wilson, Jean Amos; Jarvis, Michael; Cura, M Grace; Pratt, Victoria M

    2009-05-01

    The purpose of this work is to define rare variants of cystic fibrosis (CF) that are potential sources of error and can confound molecular genetic testing methods. We performed routine, clinical CF mutation screening using a laboratory-developed test and the oligonucleotide ligation assay reagents from Abbott/Celera. In this report, we describe 11 unique allele drop outs [3849 + 10kb C>T (NM_000492.2:c.3718-2477C>T), V520F (c.1558G>T), 1078delT (c.948delT), A455E (c.1364C>A), R347P (c.1040G>C), 2184delA (c.2052delA), W1282X (c.3846G>A), R117H (c.350G>A), G85E (c.254G>A), 621 + 1G>T (c.489 + 1G>T), and 2789 + 5G>A (c.2657 + 5G>A)] observed with this platform. The allele drop outs account for less than 0.01% of all results reported in our laboratory. Both the recognition and enumeration of such variants along with clinical information in CF testing is valuable in avoiding false-positive and false-negative results.

  9. Identification of Cystic Fibrosis Variants by Polymerase Chain Reaction/Oligonucleotide Ligation Assay

    PubMed Central

    Schwartz, Karen M.; Pike-Buchanan, Lisa L.; Muralidharan, Kasinathan; Redman, Joy B.; Wilson, Jean Amos; Jarvis, Michael; Cura, M. Grace; Pratt, Victoria M.

    2009-01-01

    The purpose of this work is to define rare variants of cystic fibrosis (CF) that are potential sources of error and can confound molecular genetic testing methods. We performed routine, clinical CF mutation screening using a laboratory-developed test and the oligonucleotide ligation assay reagents from Abbott/Celera. In this report, we describe 11 unique allele drop outs [3849 + 10kb C>T (NM_000492.2:c.3718-2477C>T), V520F (c.1558G>T), 1078delT (c.948delT), A455E (c.1364C>A), R347P (c.1040G>C), 2184delA (c.2052delA), W1282X (c.3846G>A), R117H (c.350G>A), G85E (c.254G>A), 621 + 1G>T (c.489 + 1G>T), and 2789 + 5G>A (c.2657 + 5G>A)] observed with this platform. The allele drop outs account for less than 0.01% of all results reported in our laboratory. Both the recognition and enumeration of such variants along with clinical information in CF testing is valuable in avoiding false-positive and false-negative results. PMID:19324992

  10. Identification and characterization of a human smad3 splicing variant lacking part of the linker region.

    PubMed

    Kjellman, Christian; Honeth, Gabriella; Järnum, Sofia; Lindvall, Magnus; Darabi, Anna; Nilsson, Ingar; Edvardsen, Klaus; Salford, Leif G; Widegren, Bengt

    2004-03-03

    Smad3 is one of the signal transducers that are activated in response to transforming growth factor-beta (TGF-beta). We have identified and characterized a splicing variant of smad3. The splicing variant (smad3-Delta3) lacks exon 3 resulting in a truncated linker region. We could detect mRNA expression of smad3-Delta3 in all investigated human tissues. Real-time PCR analyses demonstrated that the fraction of smad3-Delta3 mRNA compared to normal smad3 varies between tissues. The amount of spliced mRNA was estimated to represent 0.5-5% of the normal smad3 mRNA. When smad3-Delta3 is overexpressed in a fibrosarcoma cell line, the Smad3-Delta3 is translocated to the nucleus upon TGF-beta stimulation and binds the Smad responsive element. Using a CAGA luciferase reporter system, we demonstrate that Smad3-Delta3 has transcriptional activity and we conclude that Smad3-Delta3 possesses functional transactivating properties.

  11. Identification and functional characterisation of genetic variants in OLFM2 in children with developmental eye disorders.

    PubMed

    Holt, R; Ugur Iseri, S A; Wyatt, A W; Bax, D A; Gold Diaz, D; Santos, C; Broadgate, S; Dunn, R; Bruty, J; Wallis, Y; McMullan, D; Ogilvie, C; Gerrelli, D; Zhang, Y; Ragge, Nicola

    2017-01-01

    Anophthalmia, microphthalmia, and coloboma are a genetically heterogeneous spectrum of developmental eye disorders and affect around 30 per 100,000 live births. OLFM2 encodes a secreted glycoprotein belonging to the noelin family of olfactomedin domain-containing proteins that modulate the timing of neuronal differentiation during development. OLFM2 SNPs have been associated with open angle glaucoma in a case-control study, and knockdown of Olfm2 in zebrafish results in reduced eye size. From a cohort of 258 individuals with developmental eye anomalies, we identified two with heterozygous variants in OLFM2: an individual with bilateral microphthalmia carrying a de novo 19p13.2 microdeletion involving OLFM2 and a second individual with unilateral microphthalmia and contralateral coloboma who had a novel single base change in the 5' untranslated region. Dual luciferase assays demonstrated that the latter variant causes a significant decrease in expression of OLFM2. Furthermore, RNA in situ hybridisation experiments using human developmental tissue revealed expression in relevant structures, including the lens vesicle and optic cup. Our study indicates that OLFM2 is likely to be important in mammalian eye development and disease and should be considered as a gene for human ocular anomalies.

  12. Identification of CHRNA5 rare variants in African-American heavy smokers

    PubMed Central

    Doyle, Glenn A.; Chou, Andrew D.; Saung, Wint Thu; Lai, Alison T.; Lohoff, Falk W.; Berrettini, Wade H.

    2014-01-01

    The common CHRNA5 mis-sense coding single nucleotide polymorphism (SNP) rs16969968:G>A (D398N) has been shown repeatedly to confer risk for heavy smoking in individuals who carry the ‘A’ allele (encoding the 398N amino acid). The mis-sense SNP has a minor allele frequency (MAF) of ~40% in European-Americans, but only ~7% in African-Americans (http://www.ncbi.nlm.nih.gov/projects/SNP/). We reasoned that there might be other mis-sense variants among African-Americans that could confer the heavy smoking phenotype (defined here as ≥20 cigarettes per day), perhaps in a similar manner to that of the D398N polymorphism in Europeans. As such, we re-sequenced 250 African-American heavy smokers, most of whom were homozygous ‘G’ at rs16969968:G>A (MAF of 9.6% within the population). Although many novel coding SNPs were not observed, we report an interesting, although rare (perhaps personal) variant in CHRNA5 that could result in nonsense-mediated decay of the aberrant transcript. PMID:24682045

  13. Identification of common variants associated with human hippocampal and intracranial volumes.

    PubMed

    Stein, Jason L; Medland, Sarah E; Vasquez, Alejandro Arias; Hibar, Derrek P; Senstad, Rudy E; Winkler, Anderson M; Toro, Roberto; Appel, Katja; Bartecek, Richard; Bergmann, Ørjan; Bernard, Manon; Brown, Andrew A; Cannon, Dara M; Chakravarty, M Mallar; Christoforou, Andrea; Domin, Martin; Grimm, Oliver; Hollinshead, Marisa; Holmes, Avram J; Homuth, Georg; Hottenga, Jouke-Jan; Langan, Camilla; Lopez, Lorna M; Hansell, Narelle K; Hwang, Kristy S; Kim, Sungeun; Laje, Gonzalo; Lee, Phil H; Liu, Xinmin; Loth, Eva; Lourdusamy, Anbarasu; Mattingsdal, Morten; Mohnke, Sebastian; Maniega, Susana Muñoz; Nho, Kwangsik; Nugent, Allison C; O'Brien, Carol; Papmeyer, Martina; Pütz, Benno; Ramasamy, Adaikalavan; Rasmussen, Jerod; Rijpkema, Mark; Risacher, Shannon L; Roddey, J Cooper; Rose, Emma J; Ryten, Mina; Shen, Li; Sprooten, Emma; Strengman, Eric; Teumer, Alexander; Trabzuni, Daniah; Turner, Jessica; van Eijk, Kristel; van Erp, Theo G M; van Tol, Marie-Jose; Wittfeld, Katharina; Wolf, Christiane; Woudstra, Saskia; Aleman, Andre; Alhusaini, Saud; Almasy, Laura; Binder, Elisabeth B; Brohawn, David G; Cantor, Rita M; Carless, Melanie A; Corvin, Aiden; Czisch, Michael; Curran, Joanne E; Davies, Gail; de Almeida, Marcio A A; Delanty, Norman; Depondt, Chantal; Duggirala, Ravi; Dyer, Thomas D; Erk, Susanne; Fagerness, Jesen; Fox, Peter T; Freimer, Nelson B; Gill, Michael; Göring, Harald H H; Hagler, Donald J; Hoehn, David; Holsboer, Florian; Hoogman, Martine; Hosten, Norbert; Jahanshad, Neda; Johnson, Matthew P; Kasperaviciute, Dalia; Kent, Jack W; Kochunov, Peter; Lancaster, Jack L; Lawrie, Stephen M; Liewald, David C; Mandl, René; Matarin, Mar; Mattheisen, Manuel; Meisenzahl, Eva; Melle, Ingrid; Moses, Eric K; Mühleisen, Thomas W; Nauck, Matthias; Nöthen, Markus M; Olvera, Rene L; Pandolfo, Massimo; Pike, G Bruce; Puls, Ralf; Reinvang, Ivar; Rentería, Miguel E; Rietschel, Marcella; Roffman, Joshua L; Royle, Natalie A; Rujescu, Dan; Savitz, Jonathan; Schnack, Hugo G; Schnell, Knut; Seiferth, Nina; Smith, Colin; Steen, Vidar M; Valdés Hernández, Maria C; Van den Heuvel, Martijn; van der Wee, Nic J; Van Haren, Neeltje E M; Veltman, Joris A; Völzke, Henry; Walker, Robert; Westlye, Lars T; Whelan, Christopher D; Agartz, Ingrid; Boomsma, Dorret I; Cavalleri, Gianpiero L; Dale, Anders M; Djurovic, Srdjan; Drevets, Wayne C; Hagoort, Peter; Hall, Jeremy; Heinz, Andreas; Jack, Clifford R; Foroud, Tatiana M; Le Hellard, Stephanie; Macciardi, Fabio; Montgomery, Grant W; Poline, Jean Baptiste; Porteous, David J; Sisodiya, Sanjay M; Starr, John M; Sussmann, Jessika; Toga, Arthur W; Veltman, Dick J; Walter, Henrik; Weiner, Michael W; Bis, Joshua C; Ikram, M Arfan; Smith, Albert V; Gudnason, Vilmundur; Tzourio, Christophe; Vernooij, Meike W; Launer, Lenore J; DeCarli, Charles; Seshadri, Sudha; Andreassen, Ole A; Apostolova, Liana G; Bastin, Mark E; Blangero, John; Brunner, Han G; Buckner, Randy L; Cichon, Sven; Coppola, Giovanni; de Zubicaray, Greig I; Deary, Ian J; Donohoe, Gary; de Geus, Eco J C; Espeseth, Thomas; Fernández, Guillén; Glahn, David C; Grabe, Hans J; Hardy, John; Hulshoff Pol, Hilleke E; Jenkinson, Mark; Kahn, René S; McDonald, Colm; McIntosh, Andrew M; McMahon, Francis J; McMahon, Katie L; Meyer-Lindenberg, Andreas; Morris, Derek W; Müller-Myhsok, Bertram; Nichols, Thomas E; Ophoff, Roel A; Paus, Tomas; Pausova, Zdenka; Penninx, Brenda W; Potkin, Steven G; Sämann, Philipp G; Saykin, Andrew J; Schumann, Gunter; Smoller, Jordan W; Wardlaw, Joanna M; Weale, Michael E; Martin, Nicholas G; Franke, Barbara; Wright, Margaret J; Thompson, Paul M

    2012-04-15

    Identifying genetic variants influencing human brain structures may reveal new biological mechanisms underlying cognition and neuropsychiatric illness. The volume of the hippocampus is a biomarker of incipient Alzheimer's disease and is reduced in schizophrenia, major depression and mesial temporal lobe epilepsy. Whereas many brain imaging phenotypes are highly heritable, identifying and replicating genetic influences has been difficult, as small effects and the high costs of magnetic resonance imaging (MRI) have led to underpowered studies. Here we report genome-wide association meta-analyses and replication for mean bilateral hippocampal, total brain and intracranial volumes from a large multinational consortium. The intergenic variant rs7294919 was associated with hippocampal volume (12q24.22; N = 21,151; P = 6.70 × 10(-16)) and the expression levels of the positional candidate gene TESC in brain tissue. Additionally, rs10784502, located within HMGA2, was associated with intracranial volume (12q14.3; N = 15,782; P = 1.12 × 10(-12)). We also identified a suggestive association with total brain volume at rs10494373 within DDR2 (1q23.3; N = 6,500; P = 5.81 × 10(-7)).

  14. [Selection and identification of salt-tolerant variants of Taraxacum officinale].

    PubMed

    Zhang, Xinguo; Li, Yinxin; Chen, Hua; Shi, Wuliang

    2008-02-01

    In order to obtain salt-tolerant variant plants of Dandelion (Taraxacum officinale Weber), the leaf discs were excised from 20 to 30-day old seedlings to produce callus, then the induced calli were transferred to selection mediums containing 1.5% NaCl. After regenerating and rooting, these salt-tolerant calli finally developed into 12 variant plantlets. Compared with the wild-type, these regenerated plants produced more trichomes on their leaves, and had larger leaves and shorter petioles. Additionally, the dumpy roots and an approximately 2-cm bract in middle parts of the floricanes were clearly observed in these salt-tolerant plants. By RAPD (Random Amplified Polymorphic DNA) and SDS-PAGE analysis, these salt-tolerant plants showed differences from the control at DNA and protein levels. With 1.5% NaCl treatment, the antioxidant enzyme activity, proline content, and flavonoid concentration were higher in these salt-tolerant plants, whereas maloaldehyde concentration was significantly lower. Salt-tolerant lines of T. officinale showed stronger anti-oxidative activity and higher flavonoid contents.

  15. Expression of GLUT8 in mouse intestine: identification of alternative spliced variants.

    PubMed

    Romero, Amparo; Gomez, Olga; Terrado, Jose; Mesonero, Jose E

    2009-04-15

    GLUT8 is a facilitative glucose transporter composed of 10 exons coding for a 477 amino acids protein. It is mainly expressed in the testis, but it has also been studied in a number of tissues such as brain, adipose tissue, and liver. In this work, we have characterized the expression of GLUT8 in the small and large intestine under normal physiological conditions. Protein assay revealed low GLUT8 protein levels in the intestine compared to the testis, with higher levels in the colon than in the small intestine. Immunohistochemistry studies showed an intracellular localization of GLUT8 in enterocytes and colonocytes with a supranuclear distribution next to the apical membrane. GLUT8 immunoreactivity was also detected in the crypt cells. Interestingly, we have identified three additional transcriptional variants in mouse intestine (mGLUT-SP1, mGLUT8-SP2, and mGLUT8-SP3) produced by the deletion of one, two, and four exons, respectively, whereas only the entire mRNA was detected in the testis. Expression of these alternative variants did not have an effect on glucose consumption in 3T3-L1 cells. Although the specific function of GLUT8 in intestine remains unclear, the alternative splicing of GLUT8 could reflect a mechanism for the regulation of the gene expression in a tissue-specific manner by targeting GLUT8 mRNA for nonsense-mediated decay.

  16. RefCNV: Identification of Gene-Based Copy Number Variants Using Whole Exome Sequencing

    PubMed Central

    Chang, Lun-Ching; Das, Biswajit; Lih, Chih-Jian; Si, Han; Camalier, Corinne E.; McGregor, Paul M.; Polley, Eric

    2016-01-01

    With rapid advances in DNA sequencing technologies, whole exome sequencing (WES) has become a popular approach for detecting somatic mutations in oncology studies. The initial intent of WES was to characterize single nucleotide variants, but it was observed that the number of sequencing reads that mapped to a genomic region correlated with the DNA copy number variants (CNVs). We propose a method RefCNV that uses a reference set to estimate the distribution of the coverage for each exon. The construction of the reference set includes an evaluation of the sources of variability in the coverage distribution. We observed that the processing steps had an impact on the coverage distribution. For each exon, we compared the observed coverage with the expected normal coverage. Thresholds for determining CNVs were selected to control the false-positive error rate. RefCNV prediction correlated significantly (r = 0.96–0.86) with CNV measured by digital polymerase chain reaction for MET (7q31), EGFR (7p12), or ERBB2 (17q12) in 13 tumor cell lines. The genome-wide CNV analysis showed a good overall correlation (Spearman’s coefficient = 0.82) between RefCNV estimation and publicly available CNV data in Cancer Cell Line Encyclopedia. RefCNV also showed better performance than three other CNV estimation methods in genome-wide CNV analysis. PMID:27147817

  17. Near-infrared demonstration of computer-generated holograms implemented by using subwavelength gratings with space-variant orientation.

    PubMed

    Levy, Uriel; Kim, Hyo-Chang; Tsai, Chia-Ho; Fainman, Yeshaiahu

    2005-08-15

    We provide an experimental demonstration of novel form-birefringent computer-generated holograms at wavelengths of 1.55 and 10.6 microm. These novel devices utilize a 2-D array of cells that can be fabricated with a single lithographic step. Each cell contains a subwavelength binary grating whose orientation controls the desired continuous phase profile within the cell.

  18. Identification and characterization of rabbit hemorrhagic disease virus genetic variants isolated in Korea.

    PubMed

    OEM, Jae-Ku; LEE, Kwang-Nyeong; ROH, In Soon; LEE, Kyoung-Ki; KIM, Seong-Hee; KIM, Hye-Ryoung; PARK, Choi-Kyu; JOO, Yi-Seok

    2009-11-01

    Nine isolates of rabbit hemorrhagic disease virus (RHDV) were used for the genetic characterization of RHDV strains collected from rabbits in Korea between 2006 and 2008. A phylogenetic analysis of the complete VP60 region was performed and the sequences were divided mainly into two groups. The one group consisted of original RHDV and the other contained antigenic variant strain known as RHDVa strains. Most of the Korean isolates clustered with Chinese RHDV strains and belonged to the RHDVa subtype. A comparison of the amino acid sequences among RHDVa strains and original RHDV strains revealed significant substitutions of two amino acids in the A region, two in the B region, two in the F region, and nine amino acids in the E region. Taken together, the recent RHDVa strains have gradually replaced the original RHDV and are the predominant strains in Korea.

  19. Identification of functional DNA variants in the constitutive promoter region of MDM2.

    PubMed

    Lalonde, Marie-Eve; Ouimet, Manon; Larivière, Mathieu; Kritikou, Ekaterini A; Sinnett, Daniel

    2012-09-01

    Although mutations in the oncoprotein murine double minute 2 (MDM2) are rare, MDM2 gene overexpression has been observed in several human tumors. Given that even modest changes in MDM2 levels might influence the p53 tumor suppressor signaling pathway, we postulated that sequence variation in the promoter region of MDM2 could lead to disregulated expression and variation in gene dosage. Two promoters have been reported for MDM2; an internal promoter (P2), which is located near the end of intron 1 and is p53-responsive, and an upstream constitutive promoter (P1), which is p53-independent. Both promoter regions contain DNA variants that could influence the expression levels of MDM2, including the well-studied single nucleotide polymorphism (SNP) SNP309, which is located in the promoter P2; i.e., upstream of exon 2. In this report, we screened the promoter P1 for DNA variants and assessed the functional impact of the corresponding SNPs. Using the dbSNP database and genotyping validation in individuals of European descent, we identified three common SNPs (-1494 G > A; indel 40 bp; and -182 C > G). Three major promoter haplotypes were inferred by using these three promoter SNPs together with rs2279744 (SNP309). Following subcloning into a gene reporter system, we found that two of the haplotypes significantly influenced MDM2 promoter activity in a haplotype-specific manner. Site-directed mutagenesis experiments indicated that the 40 bp insertion/deletion variation is causing the observed allelic promoter activity. This study suggests that part of the variability in the MDM2 expression levels could be explained by allelic p53-independent P1 promoter activity.

  20. A new class of purple membrane variants for the construction of highly oriented membrane assemblies on the basis of noncovalent interactions.

    PubMed

    Baumann, Roelf-Peter; Busch, Annegret P; Heidel, Björn; Hampp, Norbert

    2012-04-12

    Purple membranes (PM) from Halobacterium salinarum have been discussed for several technical applications. These ideas started just several years after its discovery. The biological function of bacteriorhodopsin (BR), the only protein in PM, is the light-driven proton translocation across the membrane thereby converting light energy into chemical energy. The astonishing physicochemical robustness of this molecular assembly and the ease of its isolation triggered ideas for technical uses. All basic molecular functions of BR, that is, photochromism, photoelectrism, and proton pumping, are key elements for technical applications like optical data processing and data storage, ultrafast light detection and processing, and direct utilization of sunlight in adenosine 5'-triphospate (ATP) generation or seawater desalination. In spite of the efforts of several research groups worldwide, which confirmed the proof-of-principle for all these potential applications, only the photochromism-based applications have reached a technical level. The physical reason for this is that no fixation or orientation of the PMs is required. The situation is quite different for photoelectrism and proton pumping where the macroscopic orientation of PMs is a prerequisite. For proton pumping, in addition, the formation of artificial membranes which prevent passive proton leakage is necessary. In this manuscript, we describe a new class of PM variants with oppositely charged membrane sides which enable an almost 100% orientation on a surface, which is the key element for photoelectric applications of BR. As an example, the mutated BR, BR-E234R7, was prepared and analyzed. A nearly 100% self-orientation on mica was obtained.

  1. Accuracy of next-generation sequencing for the identification of clinically relevant variants in cytology smears in lung adenocarcinoma.

    PubMed

    Baum, Jordan E; Zhang, Pan; Hoda, Rana S; Geraghty, Brian; Rennert, Hanna; Narula, Navneet; Fernandes, Helen D

    2017-06-01

    Minimally invasive diagnostic procedures such as needle-core biopsy and fine-needle aspiration provide adequate material for molecular analyses. Advances in precision oncology are trending toward the interrogation of limited amounts of genomic material to guide clinical and therapeutic decisions. The aim of this study was to investigate the minimum cellularity needed on cytologic smears for the identification of clinically relevant variants with next-generation sequencing (NGS). Thirty cases of cytologically diagnosed, resection-proven primary lung adenocarcinoma were identified. Nineteen of the 30 cases were known to harbor actionable variants. One Diff-Quik (DQ)-stained slide and 1 Papanicolaou (Pap)-stained slide were selected from each case. Cases were categorized as containing fewer than 100 tumor cells, 100 to 500 tumor cells, or more than 500 tumor cells. NGS was performed on the Ion Torrent platform. NGS was successfully performed on all cell blocks and on 90% of the smears. Paired DQ and Pap smears showed similar cellularity, and cases that differed in cellularity were within 1 category of each other. The cases with more than 100 tumor cells had a 93% success rate; this was significantly different from the situation for cases with fewer than 100 tumor cells, which were successfully sequenced only 67% of the time. Overall, NGS was able to provide clinically relevant information for 83% of DQ smears and for 90% of Pap smears tested. The data show a significantly higher likelihood of successful NGS with cytologic smears with more than 100 tumor cells. There was a trend for a higher NGS success rate with Pap smears versus DQ smears. Cancer Cytopathol 2017;125:398-406. © 2017 American Cancer Society. © 2017 American Cancer Society.

  2. Identification of common variants influencing risk of the tauopathy Progressive Supranuclear Palsy

    PubMed Central

    Höglinger, Günter U.; Melhem, Nadine M.; Dickson, Dennis W.; Sleiman, Patrick M.A.; Wang, Li-San; Klei, Lambertus; Rademakers, Rosa; de Silva, Rohan; Litvan, Irene; Riley, David E.; van Swieten, John C.; Heutink, Peter; Wszolek, Zbigniew K.; Uitti, Ryan J.; Vandrovcova, Jana; Hurtig, Howard I.; Gross, Rachel G.; Maetzler, Walter; Goldwurm, Stefano; Tolosa, Eduardo; Borroni, Barbara; Pastor, Pau; Cantwell, Laura B.; Han, Mi Ryung; Dillman, Allissa; van der Brug, Marcel P.; Gibbs, J Raphael; Cookson, Mark R.; Hernandez, Dena G.; Singleton, Andrew B.; Farrer, Matthew J.; Yu, Chang-En; Golbe, Lawrence I.; Revesz, Tamas; Hardy, John; Lees, Andrew J.; Devlin, Bernie; Hakonarson, Hakon; Müller, Ulrich; Schellenberg, Gerard D.

    2011-01-01

    Progressive supranuclear palsy (PSP) is a movement disorder with prominent tau neuropathology. Brain diseases with abnormal tau deposits are called tauopathies, the most common being Alzheimer’s disease. Environmental causes of tauopathies include repetitive head trauma associated with some sports. To identify common genetic variation contributing to risk for tauopathies, we carried out a genome-wide association study of 1,114 PSP cases and 3,247 controls (Stage 1) followed up by a second stage where 1,051 cases and 3,560 controls were genotyped for Stage 1 SNPs that yielded P ≤ 10−3. We found significant novel signals (P < 5 × 10−8) associated with PSP risk at STX6, EIF2AK3, and MOBP. We confirmed two independent variants in MAPT affecting risk for PSP, one of which influences MAPT brain expression. The genes implicated encode proteins for vesicle-membrane fusion at the Golgi-endosomal interface, for the endoplasmic reticulum unfolded protein response, and for a myelin structural component. PMID:21685912

  3. Identification of a splice-site mutation in the human growth hormone-variant gene.

    PubMed Central

    MacLeod, J N; Liebhaber, S A; MacGillivray, M H; Cooke, N E

    1991-01-01

    The human growth-hormone-variant (hGH-V) gene normally expresses two alternatively spliced forms of mRNA--hGH-V and hGH-V2--in the placenta. hGH-V2 mRNA differs from hGH-V rDNA by the retention of intron 4 and represents approximately 15% of transcripts at term. In a survey of hGH-V gene expression in 20 placentas of gestational age 8-40 wk, we detected a single placenta that contained, in addition to the two normal hGH-V mRNA species, a set of two slightly larger hGH-V mRNAs. Sequence analysis of the elongated hGH-V mRNA demonstrated retention of the first 12 bases of intron 2, resulting from both a base substitution at the intron 2 splice-donor dinucleotide (GT----AT) and activation of a cryptic splice-donor site 12 bases downstream. Survey of a total of 60 additional chromosomes failed to reveal additional incidence of this mutation. The mutation, which we have designated hGH-Vintron 2, pos 1 (G----A), represents both an initial example of a nondeletional mutation within the hGH-V gene and corresponding structural alteration in the encoded hGH-V hormone. Images Figure 1 Figure 2 Figure 4 PMID:2035535

  4. Identification of copy number variants associated with BPES-like phenotypes.

    PubMed

    Gijsbers, Antoinet C J; D'haene, Barbara; Hilhorst-Hofstee, Yvonne; Mannens, Marcel; Albrecht, Beate; Seidel, Joerg; Witt, David R; Maisenbacher, Melissa K; Loeys, Bart; van Essen, Ton; Bakker, Egbert; Hennekam, Raoul; Breuning, Martijn H; De Baere, Elfride; Ruivenkamp, Claudia A L

    2008-12-01

    Blepharophimosis-Ptosis-Epicanthus inversus syndrome (BPES) is a well-characterized rare syndrome that includes an eyelid malformation associated with (type I) or without premature ovarian failure (type II). Patients with typical BPES have four major characteristics: blepharophimosis, ptosis, epicanthus inversus and telecanthus. Mutations in the FOXL2 gene, encoding a forkhead transcription factor, are responsible for the majority of both types of BPES. However, many patients with BPES-like features, i.e., having at least two major characteristics of BPES, have an unidentified cause. Here, we report on a group of 27 patients with BPES-like features, but without an identified genetic defect in the FOXL2 gene or flanking region. These patients were analyzed with whole-genome high-density arrays in order to identify copy number variants (CNVs) that might explain the BPES-like phenotype. In nine out of 27 patients (33%) CNVs not previously described as polymorphisms were detected. Four of these patients displayed psychomotor retardation as an additional clinical characteristic. In conclusion, we demonstrate that BPES-like phenotypes are frequently caused by CNVs, and we emphasize the importance of whole-genome copy number screening to identify the underlying genetic causes of these phenotypes.

  5. Identification of a novel splice variant isoform of TREM-1 in human neutrophil granules1

    PubMed Central

    Baruah, Sankar; Keck, Kathy; Vrenios, Michelle; Pope, Marshall; Pearl, Merideth; Doerschug, Kevin; Klesney-Tait, Julia

    2015-01-01

    Triggering receptor expressed on myeloid cells-1 (TREM-1) is critical for inflammatory signal amplification. Humans have two forms of TREM-1: a membrane receptor (mbTREM-1), associated with the adaptor DAP12, and a soluble receptor detected at times of infection. The membrane receptor isoform acts synergistically with the TLR pathway to promote cytokine secretion and neutrophil migration while the soluble receptor functions as a counter regulatory molecule. In multiple models of sepsis, exogenous administration of soluble forms of TREM-1 attenuates inflammation and markedly improves survival. Despite intense interest in soluble TREM-1 both as a clinical predictor of survival and as a therapeutic tool, the origin of native soluble TREM-1 remains controversial. Utilizing human neutrophils, we identified a 15 kDa TREM-1 isoform in primary (azurophilic) and secondary (specific) granules. Mass spectrometric analysis, ELISA, and immunoblot confirm that the 15 kD protein is a novel splice variant of TREM-1 (TREM-1sv). Neutrophil stimulation with P. aeruginosa, LPS, or PAM(3)Cys4 resulted in degranulation and release of TREM-1sv. The addition of exogenous TREM-1sv inhibited TREM-1 receptor mediated proinflammatory cytokine production. Thus these data reveal that TREM-1 isoforms simultaneously activate and inhibit inflammation via the canonical membrane TREM-1 molecule and this newly discovered granular isoform, TREM-1sv. PMID:26561551

  6. Identification and characterization of a novel splice variant of rhesus macaque MHC IA.

    PubMed

    Dai, Zheng-Xi; Zhang, Gao-Hong; Zhang, Xi-He; Zheng, Yong-Tang

    2013-03-01

    Major histocompatibility complex class I (MHC I) molecules play a pivotal role in the immune recognition to intracellular pathogens. A number of important splice variants have already been characterized for these molecules in different species, suggesting their important roles in modulation of immune responses. In this study, we have identified and characterized a novel alternatively spliced form of rhesus macaque MHC IA (designated MHC IA-sv2) that lacks exons coding for the α2 and α3 domains. Despite lacking the α2 and α3 domains, MHC IA-sv2 is targeted to the cell surface, as a 23-kDa glycoprotein that is totally susceptible to endoglycosidase-H digestion and is reduced to 18kDa after deglycosylation with PNGase F. In contrast, the full-length MHC IA reaches the cell surface as a 43-kDa protein of form with complex-type N-glycosylation (endoglycosidase-H resistant). Moreover, we provide evidence here that MHC IA-sv2 can self-associate, forming homodimers, or associate with the fully mature MHC IA molecule, forming a heterodimeric structure in mammalian cells. These data demonstrate that the formation of heterodimers may have some functional implications in the fine tuning of MHC IA-mediated innate and adaptive immune responses.

  7. Identification of the topological defects of optical indicatrix orientation in CaB4O7 glasses

    NASA Astrophysics Data System (ADS)

    Vasylkiv, Yu; Skab, I.; Vlokh, R.

    2016-08-01

    We develop an experimental method for the identification of the topological defects of optical indicatrix orientation, which appear in glass samples with residual mechanical stresses. The criteria needed to determine the sign of the strength of topological defects are formulated. The method is verified on the example of a non-annealed CaB4O7 glass sample that manifests spatially inhomogeneous residual stresses. We have found that at least seven topological defects of optical indicatrix orientation exist under the condition when a light beam propagates through a parallelepiped-shaped glass sample. The strengths of all the defects detected by us are equal to +1/2.

  8. Identification of Genetic and Epigenetic Variants Associated with Breast Cancer Prognosis by Integrative Bioinformatics Analysis

    PubMed Central

    Shilpi, Arunima; Bi, Yingtao; Jung, Segun; Patra, Samir K.; Davuluri, Ramana V.

    2017-01-01

    INTRODUCTION Breast cancer being a multifaceted disease constitutes a wide spectrum of histological and molecular variability in tumors. However, the task for the identification of these variances is complicated by the interplay between inherited genetic and epigenetic aberrations. Therefore, this study provides an extrapolate outlook to the sinister partnership between DNA methylation and single-nucleotide polymorphisms (SNPs) in relevance to the identification of prognostic markers in breast cancer. The effect of these SNPs on methylation is defined as methylation quantitative trait loci (meQTL). MATERIALS AND METHODS We developed a novel method to identify prognostic gene signatures for breast cancer by integrating genomic and epigenomic data. This is based on the hypothesis that multiple sources of evidence pointing to the same gene or pathway are likely to lead to reduced false positives. We also apply random resampling to reduce overfitting noise by dividing samples into training and testing data sets. Specifically, the common samples between Illumina 450 DNA methylation, Affymetrix SNP array, and clinical data sets obtained from the Cancer Genome Atlas (TCGA) for breast invasive carcinoma (BRCA) were randomly divided into training and test models. An intensive statistical analysis based on log-rank test and Cox proportional hazard model has established a significant association between differential methylation and the stratification of breast cancer patients into high- and low-risk groups, respectively. RESULTS The comprehensive assessment based on the conjoint effect of CpG–SNP pair has guided in delaminating the breast cancer patients into the high- and low-risk groups. In particular, the most significant association was found with respect to cg05370838–rs2230576, cg00956490–rs940453, and cg11340537–rs2640785 CpG–SNP pairs. These CpG–SNP pairs were strongly associated with differential expression of ADAM8, CREB5, and EXPH5 genes, respectively

  9. Identification of putative second genetic hits in schizophrenia carriers of high-risk copy number variants and resequencing in additional samples.

    PubMed

    Rodríguez-López, Julio; Sobrino, Beatriz; Amigo, Jorge; Carrera, Noa; Brenlla, Julio; Agra, Santiago; Paz, Eduardo; Carracedo, Ángel; Páramo, Mario; Arrojo, Manuel; Costas, Javier

    2017-04-18

    Copy number variants (CNVs) conferring risk of schizophrenia present incomplete penetrance, suggesting the existence of second genetic hits. Identification of second hits may help to find genes with rare variants of susceptibility to schizophrenia. The aim of this work was to search for second hits of moderate/high risk in schizophrenia carriers of risk CNVs and resequencing of the relevant genes in additional samples. To this end, ten patients with risk CNVs at cytobands 15q11.2, 15q11.2-13.1, 16p11.2, or 16p13.11, were subjected to whole-exome sequencing. Rare single nucleotide variants, defined as those absent from main public databases, were classified according to bioinformatic prediction of pathogenicity by CADD scores. The average number of rare predicted pathogenic variants per sample was 13.6 (SD 2.01). Two genes, BFAR and SYNJ1, presented rare predicted pathogenic variants in more than one sample. Follow-up resequencing of these genes in 432 additional cases and 432 controls identified a significant excess of rare predicted pathogenic variants in case samples at SYNJ1. Taking into account its function in clathrin-mediated synaptic vesicle endocytosis at presynaptic terminals, our results suggest an impairment of this process in schizophrenia.

  10. Identification of circadian gene variants in bipolar disorder in Latino populations.

    PubMed

    Gonzalez, Robert; Gonzalez, Suzanne; Villa, Erika; Ramirez, Mercedes; Zavala, Juan; Armas, Regina; Contreras, Javier; Dassori, Albana; Leach, Robin J; Flores, Deborah; Jerez, Alvaro; Raventós, Henriette; Ontiveros, Alfonso; Nicolini, Humberto; Escamilla, Michael

    2015-11-01

    Variations in circadian genes can impact biological rhythms. Given the rhythm disturbances that characterize bipolar disorder (BD), genes encoding components of molecular clocks are good candidate genes for the illness. A family based association analysis of circadian gene single nucleotide polymorphisms (SNPs) and BD was conducted in Latino pedigrees. 884 individuals from 207 pedigrees (473BP phenotype and 411 unaffected family members) were genotyped. Family based single marker association testing was performed. Ancestral haplotypes (SNPs found to be in strong LD defined using confidence intervals) were also tested for association with BD. Multiple suggestive associations between circadian gene SNPs and BD were noted. These included CSNK1E (rs1534891, p=0.00689), ARNTL (rs3789327, p=0.021172), CSNK1D (rs4510078, p=0.022801), CLOCK (rs17777927, p=0.031664). Individually, none of the SNPs were significantly associated with BD after correction for multiple testing. However, a 4-locus CSNK1E haplotype encompassing the rs1534891 SNP (Z-score=2.685, permuted p=0.0076) and a 3-locus haplotype in ARNTL (Z-score=3.269, permuted p=0.0011) showed a significant association with BD. Larger samples are required to confirm these findings and assess the relationship between circadian gene SNPs and BD in Latinos. The results suggest that ARNTL and CSKN1E variants may be associated with BD. Further studies are warranted to assess the relationships between these genes and BD in Latino populations. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

  11. Identification of Common Genetic Variants Influencing Spontaneous Dizygotic Twinning and Female Fertility

    PubMed Central

    Mbarek, Hamdi; Steinberg, Stacy; Nyholt, Dale R.; Gordon, Scott D.; Miller, Michael B.; McRae, Allan F.; Hottenga, Jouke Jan; Day, Felix R.; Willemsen, Gonneke; de Geus, Eco J.; Davies, Gareth E.; Martin, Hilary C.; Penninx, Brenda W.; Jansen, Rick; McAloney, Kerrie; Vink, Jacqueline M.; Kaprio, Jaakko; Plomin, Robert; Spector, Tim D.; Magnusson, Patrik K.; Reversade, Bruno; Harris, R. Alan; Aagaard, Kjersti; Kristjansson, Ragnar P.; Olafsson, Isleifur; Eyjolfsson, Gudmundur Ingi; Sigurdardottir, Olof; Iacono, William G.; Lambalk, Cornelis B.; Montgomery, Grant W.; McGue, Matt; Ong, Ken K.; Perry, John R.B.; Martin, Nicholas G.; Stefánsson, Hreinn; Stefánsson, Kari; Boomsma, Dorret I.

    2016-01-01

    Spontaneous dizygotic (DZ) twinning occurs in 1%–4% of women, with familial clustering and unknown physiological pathways and genetic origin. DZ twinning might index increased fertility and has distinct health implications for mother and child. We performed a GWAS in 1,980 mothers of spontaneous DZ twins and 12,953 control subjects. Findings were replicated in a large Icelandic cohort and tested for association across a broad range of fertility traits in women. Two SNPs were identified (rs11031006 near FSHB, p = 1.54 × 10−9, and rs17293443 in SMAD3, p = 1.57 × 10−8) and replicated (p = 3 × 10−3 and p = 1.44 × 10−4, respectively). Based on ∼90,000 births in Iceland, the risk of a mother delivering twins increased by 18% for each copy of allele rs11031006-G and 9% for rs17293443-C. A higher polygenic risk score (PRS) for DZ twinning, calculated based on the results of the DZ twinning GWAS, was significantly associated with DZ twinning in Iceland (p = 0.001). A higher PRS was also associated with having children (p = 0.01), greater lifetime parity (p = 0.03), and earlier age at first child (p = 0.02). Allele rs11031006-G was associated with higher serum FSH levels, earlier age at menarche, earlier age at first child, higher lifetime parity, lower PCOS risk, and earlier age at menopause. Conversely, rs17293443-C was associated with later age at last child. We identified robust genetic risk variants for DZ twinning: one near FSHB and a second within SMAD3, the product of which plays an important role in gonadal responsiveness to FSH. These loci contribute to crucial aspects of reproductive capacity and health. PMID:27132594

  12. Identification and characterization of alternatively spliced variants of DNA methyltransferase 3a in mammalian cells.

    PubMed

    Weisenberger, Daniel J; Velicescu, Mihaela; Preciado-Lopez, Miguel A; Gonzales, Felicidad A; Tsai, Yvonne C; Liang, Gangning; Jones, Peter A

    2002-09-18

    CpG methylation is mediated by the functions of at least three active DNA methyltransferases (DNMTs). While DNMT1 is thought to perform maintenance methylation, the more recently discovered DNMT3a and DNMT3b enzymes are thought to facilitate de novo methylation. Murine Dnmt3a and 3b are developmentally regulated and a new Dnmt3a isoform, Dnmt3a2, has been recently shown to be expressed preferentially in mouse embryonic stem (ES) cells. Here we have characterized four alternatively spliced variants of human and mouse DNMT3a. These transcripts included a novel exon 1 (1beta) that was spliced into the same exon 2 acceptor splice site used by the original exon 1 (1alpha). Cloning and sequencing of the 5' region of the human DNMT3a gene revealed that exon 1beta was situated upstream of exon 1alpha and that the entire region was contained within a CpG island. We also identified other alternatively spliced species containing intron 4 inclusions that were associated with either exon 1alpha or 1beta. These were expressed at low levels in mouse and human cells. All transcripts were highly conserved between human and mouse. The levels of Dnmt3a mRNA containing exon 1beta were 3-25-fold greater in mouse ES cells than in various somatic cells as determined by semiquantitative reverse transcription-polymerase chain reaction analysis, while the levels of exon 1alpha-containing transcripts were slightly higher in human and mouse somatic cells. The preferential expression of the beta transcript in ES cells suggests that this transcript, in addition to Dnmt3a2, may also be important for de novo methylation during development.

  13. Identification of a genetic variant associated with rotator cuff repair healing.

    PubMed

    Tashjian, Robert Z; Granger, Erin K; Zhang, Yue; Teerlink, Craig C; Cannon-Albright, Lisa A

    2016-06-01

    A familial and genetic predisposition for the development of rotator cuff tearing has been identified. The purpose of this study was to determine if a familial predisposition exists for healing after rotator cuff repair and if the reported significant association with a single-nucleotide polymorphism (SNP) in the ESRRB gene is present in patients who fail to heal. The study recruited 72 patients undergoing arthroscopic rotator cuff repair for a full-thickness posterosuperior tear. Magnetic resonance imaging studies were performed at a minimum of 1 year postoperatively (average, 2.6 years). Healing failures were classified as lateral or medial. Self-reported family history of rotator cuff tearing data and genome-wide genotypes were available. Characteristics of cases with and without a family history of rotator cuff tearing were compared, and a comparison of the frequency of SNP 1758384 (in ESRRB) was performed between patients who healed and those who failed to heal. Of the rotator cuff repairs, 42% failed to heal; 42% of patients reported a family history of rotator cuff tear. Multivariate regression analysis showed a significant association between familiality and overall healing failure (medial and lateral failures) (P = .036) and lateral failures independently (P = .006). An increased risk for the presence of a rare allele for SNP rs17583842 was present in lateral failures compared with those that healed (P = .005). Individuals with a family history of rotator cuff tearing were more likely to have repair failures. Significant association of a SNP variant in the ESRRB gene was also observed with lateral failure. Copyright © 2016 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.

  14. Identification of a functional variant in SPLUNC1 associated with nasopharyngeal carcinoma susceptibility among Malaysian Chinese.

    PubMed

    Yew, Poh-Yin; Mushiroda, Taisei; Kiyotani, Kazuma; Govindasamy, Gopala Krishnan; Yap, Lee-Fah; Teo, Soo-Hwang; Lim, Paul Vey-Hong; Govindaraju, Selvaratnam; Ratnavelu, Kananathan; Sam, Choon-Kook; Yap, Yoke-Yeow; Khoo, Alan Soo-Beng; Pua, Kin-Choo; Nakamura, Yusuke; Ng, Ching-Ching

    2012-10-01

    Nasopharyngeal carcinoma (NPC) is a multifactorial and polygenic disease with high incidence in Asian countries. Epstein-Barr virus infection, environmental and genetic factors are believed to be involved in the tumorigenesis of NPC. The association of single nucleotide polymorphisms (SNPs) in LPLUNC1 and SPLUNC1 genes with NPC was investigated by performing a two-stage case control association study in a Malaysian Chinese population. The initial screening consisted of 81 NPC patients and 147 healthy controls while the replication study consisted of 366 NPC patients and 340 healthy controls. The combined analysis showed that a SNP (rs2752903) of SPLUNC1 was significantly associated with the risk of NPC (combined P = 0.00032, odds ratio = 1.62, 95% confidence interval = 1.25-2.11). In the subsequent dense fine mapping of SPLUNC1 locus, 36 SNPs in strong linkage disequilibrium with rs2752903 (r(2) ≥ 0.85) were associated with NPC susceptibility. Screening of these variants by electrophoretic mobility shift and luciferase reporter assays showed that rs1407019 located in intron 3 (r(2)  = 0.994 with rs2752903) caused allelic difference in the binding of specificity protein 1 (Sp1) transcription factor and affected luciferase activity. This SNP may consequently alter the expression of SPLUNC1 in the epithelial cells. In summary, our study suggested that rs1407019 in intronic enhancer of SPLUNC1 is associated with NPC susceptibility in which its A allele confers an increased risk of NPC in the Malaysian Chinese population. Copyright © 2011 Wiley Periodicals, Inc.

  15. Identification of time-variant river bed properties with the ensemble Kalman filter

    NASA Astrophysics Data System (ADS)

    Kurtz, Wolfgang; Hendricks Franssen, Harrie-Jan; Vereecken, Harry

    2012-10-01

    An adequate characterization of river bed hydraulic conductivities (L) is crucial for a proper assessment of river-aquifer interactions. However, river bed characteristics may change over time due to dynamic morphological processes like scouring or sedimentation what can lead to erroneous model predictions when static leakage parameters are assumed. Sequential data assimilation with the ensemble Kalman filter (EnKF) allows for an update of model parameters in real-time and may thus be capable of assessing the transient behavior of L. Synthetic experiments with a three-dimensional finite element model of the Limmat aquifer in Zurich were used to assess the performance of data assimilation in capturing time-variant river bed properties. Reference runs were generated where L followed different temporal and/or spatial patterns which should mimic real-world sediment dynamics. Hydraulic head (h) data from these reference runs were then used as input data for EnKF which jointly updated h and L. Results showed that EnKF is able to capture the different spatio-temporal patterns of L in the reference runs well. However, the adaptation time was relatively long which was attributed to the fast decrease of ensemble variance. To improve the performance of EnKF also an adaptive filtering approach with covariance inflation was applied that allowed a faster and more accurate adaptation of model parameters. A sensitivity analysis indicated that even for a low amount of observations a reasonable adaptation of L towards the reference values can be achieved and that EnKF is also able to correct for a biased initial ensemble of L.

  16. Identification of Alzheimer disease-associated variants in genes that regulate retromer function.

    PubMed

    Vardarajan, Badri N; Bruesegem, Sophia Y; Harbour, Michael E; Inzelberg, Rivka; Friedland, Robert; St George-Hyslop, Peter; Seaman, Matthew N J; Farrer, Lindsay A

    2012-09-01

    The proteolytic processing of amyloid precursor protein (APP) to generate the neurotoxic amyloid β (Aβ) peptide is central to the pathogenesis of Alzheimer disease (AD). The endocytic system mediates the processing of APP by controlling its access to secretases that cleave APP. A key mediator of APP localization is SorL1-a membrane protein that has been genetically linked to AD. The retromer complex is a conserved protein complex required for endosome-to-Golgi retrieval of a number of physiologically important membrane proteins including SorL1. Based on the prior suggestion that endocytosis and retromer sorting pathways might be involved, we hypothesized that variants in other genes in this pathway might also modulate AD risk. Genetic association of AD with 451 polymorphisms in 15 genes encoding retromer or retromer-associated proteins was tested in a Caucasian sample of 8309 AD cases and 7366 cognitively normal elders using individual single nucleotide polymorphism (SNP)- and gene-based tests. We obtained significant evidence of association with KIAA1033 (VEGAS p = 0.025), SNX1 (VEGAS p = 0.035), SNX3 (p = 0.0057), and RAB7A (VEGAS p = 0.018). Ten KIAA1033 SNPs were also significantly associated with AD in a group of African Americans (513 AD cases, 504 control subjects). Findings with four significant SNX3 SNPs in the discovery sample were replicated in a community-based sample of Israeli-Arabs (124 AD cases, 142 control subjects). We show that Snx3 and Rab7A proteins interact with the cargo-selective retromer complex through independent mechanisms to regulate the membrane association of retromer and thereby are key mediators of retromer function. These data implicate additional AD risk genes in the retromer pathway and formally demonstrate a direct link between the activity of the retromer complex and the pathogenesis of AD. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. Identification of two novel splicing variants of murine NTE-related esterase.

    PubMed

    Chang, Ping-An; Wang, Zhan-Xiang; Long, Ding-Xin; Qin, Wen-Zhen; Wei, Chen-Ying; Wu, Yi-Jun

    2012-04-15

    NTE-related esterase (NRE) is an insulin-regulated lysophospholipase with homology to neuropathy target esterase (NTE), which plays a role in energy metabolism. Here, we reported two alternative splicing variants of the murine NRE (mNRE) gene, termed mNREV1 and mNREV2. Genomic organization analysis indicated that 5' splice site of mNRE intron 33 was changed in both mNREV1 and mNREV2, and mNRE exon 21 was deleted in mNREV2. mNREV1 had the same protein domains with mNRE, while mNREV2 lacked the patatin domain in the C-terminal catalytic region. Green fluorescent protein-mNREV1 or mNREV2 fusion proteins localized to the endoplasmic reticulum. mNREV1 and mNRE exhibited equal hydrolytic activity to the substrate phenyl valerate, whereas mNREV2 did not have any catalytic activity. The expression profiles of mNRE and its splicing isoforms in white adipose tissue, cardiac muscle, skeletal muscle, and testis tissues were further analyzed by real time quantitative-PCR in fed and fasted states, which indicated that the major isoform of mNRE mRNA generated switched from mNREV2 to mNREV1 during fasting. Thus there was a nutritional regulation of mNRE expression at the mRNA levels via alternative splicing. Copyright © 2012 Elsevier B.V. All rights reserved.

  18. Enhanced Identification of Potential Pleiotropic Genetic Variants for Bone Mineral Density and Breast Cancer.

    PubMed

    Peng, Cheng; Lou, Hui-Ling; Liu, Feng; Shen, Jie; Lin, Xu; Zeng, Chun-Ping; Long, Ji-Rong; Su, Kuan-Jui; Zhang, Lan; Greenbaum, Jonathan; Deng, Wei-Feng; Li, Yu-Mei; Deng, Hong-Wen

    2017-07-31

    Epidemiological and clinical evidences have shown that bone mineral density (BMD) has a close relationship with breast cancer (BC). They might potentially have a shared genetic basis. By incorporating information about these pleiotropic effects, we may be able to explore more of the traits' total heritability. We applied a recently developed conditional false discovery rate (cFDR) method to the summary statistics from two independent GWASs to identify the potential pleiotropic genetic variants for BMD and BC. By jointly analyzing two large independent GWASs of BMD and BC, we found strong pleiotropic enrichment between them and identified 102 single-nucleotide polymorphisms (SNPs) in BMD and 192 SNPs in BC with cFDR < 0.05, including 230 SNPs that might have been overlooked by the standard GWAS analysis. cFDR-significant genes were enriched in GO terms and KEGG pathways which were crucial to bone metabolism and/or BC pathology (adjP < 0.05). Some cFDR-significant genes were partially validated in the gene expressional validation assay. Strong interactions were found between proteins produced by cFDR-significant genes in the context of biological mechanism of bone metabolism and/or BC etiology. Totally, we identified 7 pleiotropic SNPs that were associated with both BMD and BC (conjunction cFDR < 0.05); CCDC170, ESR1, RANKL, CPED1, and MEOX1 might play important roles in the pleiotropy of BMD and BC. Our study highlighted the significant pleiotropy between BMD and BC and shed novel insight into trait-specific as well as the potentially shared genetic architecture for both BMD and BC.

  19. IDENTIFICATION AND HORMONE INDUCTION OF PUTATIVE CHITIN SYNTHASE GENES AND SPLICE VARIANTS IN Leptinotarsa decemlineata (SAY).

    PubMed

    Shi, Ji-Feng; Mu, Li-Li; Guo, Wen-Chao; Li, Guo-Qing

    2016-08-01

    Chitin synthase (ChS) plays a critical role in chitin synthesis and excretion. In this study, two ChS genes (LdChSA and LdChSB) were identified in Leptinotarsa decemlineata. LdChSA contains two splicing variants, LdChSAa and LdChSAb. Within the first, second, and third larval instars, the mRNA levels of LdChSAa, LdChSAb, and LdChSB coincide with the peaks of circulating 20-hydroxyecdysone (20E) and juvenile hormone (JH). In vitro culture of midguts and an in vivo bioassay revealed that 20E and an ecdysteroid agonist halofenozide stimulated the expression of the three LdChSs. Conversely, a reduction of 20E by RNA interference (RNAi) of an ecdysteroidogenesis gene LdSHD repressed the expression of these LdChSs, and ingestion of halofenozide by LdSHD RNAi larvae rescued the repression. Moreover, disruption of 20E signaling by RNAi of LdEcR, LdE75, LdHR3, and LdFTZ-F1 reduced the expression levels of these genes. Similarly, in vitro culture and an in vivo bioassay showed that exogenous JH and a JH analog methoprene activated the expression of the three LdChSs, whereas a decrease in JH by RNAi of a JH biosynthesis gene LdJHAMT downregulated these LdChSs. It seems that JH upregulates LdChSs at the early stage of each instar, whereas a 20E pulse triggers the transcription of LdChSs during molting in L. decemlineata.

  20. Identification of homologous, homoeologous and paralogous sequence variants in an outbreeding allopolyploid species based on comparison with progenitor taxa.

    PubMed

    Hand, Melanie L; Ponting, Rebecca C; Drayton, Michelle C; Lawless, Kahlil A; Cogan, Noel O I; Charles Brummer, E; Sawbridge, Timothy I; Spangenberg, German C; Smith, Kevin F; Forster, John W

    2008-10-01

    The combination of homologous, homoeologous and paralogous classes of sequence variation presents major challenges for SNP discovery in outbreeding allopolyploid species. Previous in vitro gene-associated SNP discovery studies in the allotetraploid forage legume white clover (Trifolium repens L.) were vulnerable to such effects, leading to prohibitive levels of attrition during SNP validation. Identification of T. occidentale and T. pallescens as the putative diploid progenitors of white clover has permitted discrimination of the different sequence variant categories. Amplicons from selected abiotic stress tolerance-related genes were obtained using mapping family parents and individuals from each diploid species. Following cloning, progenitor comparison allowed tentative assignment of individual haplotypes to one or other sub-genome, as well as to gene copies within sub-genomes. A high degree of coincidence and identity between SNPs and HSVs was observed. Close similarity was observed between the genome of T. occidentale and one white clover sub-genome, but the affinity between T. pallescens and the other sub-genome was weaker, suggesting that a currently uncharacterised taxon may be the true second progenitor. Selected validated SNPs were attributed to individual sub-genomes by assignment to and naming of homoeologous linkage groups, providing the basis for improved genetic trait-dissection studies. The approach described in this study is broadly applicable to a range of allopolyploid taxa of equivocal ancestry.

  1. Functional effects of different medium-chain acyl-CoA dehydrogenase genotypes and identification of asymptomatic variants.

    PubMed

    Sturm, Marga; Herebian, Diran; Mueller, Martina; Laryea, Maurice D; Spiekerkoetter, Ute

    2012-01-01

    Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency (OMIM 201450) is the most common inherited disorder of fatty acid metabolism presenting with hypoglycaemia, hepatopathy and Reye-like symptoms during catabolism. In the past, the majority of patients carried the prevalent c.985A>G mutation in the ACADM gene. Since the introduction of newborn screening many other mutations with unknown clinical relevance have been identified in asymptomatic newborns. In order to identify functional effects of these mutant genotypes we correlated residual MCAD (OMIM 607008) activities as measured by octanoyl-CoA oxidation in lymphocytes with both genotype and relevant medical reports in 65 newborns harbouring mutant alleles. We identified true disease-causing mutations with residual activities of 0 to 20%. In individuals carrying the c.199T>C or c.127G>A mutation on one allele, residual activities were much higher and in the range of heterozygotes (31%-60%). Therefore, both mutations cannot clearly be associated with a clinical phenotype. This demonstrates a correlation between the octanoyl-CoA oxidation rate in lymphocytes and the clinical outcome. With newborn screening, the natural course of disease is difficult to assess. The octanoyl-CoA oxidation rate, therefore, allows a risk assessment at birth and the identification of new ACADM genotypes associated with asymptomatic disease variants.

  2. Functional identification of a novel transcript variant of INPP4B in human colon and breast cancer cells.

    PubMed

    Croft, Amanda; Guo, Su Tang; Sherwin, Simonne; Farrelly, Margaret; Yan, Xu Guang; Zhang, Xu Dong; Jiang, Chen Chen

    2017-03-25

    The 4-phosphatase Inositol polyphosphate 4-phosphatase II (INPP4B) is a regulator of the PI3K signalling pathway and functions to suppress or promote activation of downstream kinases depending on cell type and context. Here we report the identification of a novel small transcript variant of INPP4B (INPP4B-S) that has a role in promoting proliferation of colon and breast cancer cells. INPP4B-S differed from full length INPP4B (INPP4B-FL) by the insertion of a small exon between exons 15 and 16 and the deletion of exons 20-24. Nevertheless, INPP4B-S retained all the functional domains of INPP4B-FL and was similarly located to the cytoplasm. Overexpression of INPP4B-S increased, whereas selective knockdown of INPP4B-S reduced the rate of proliferation in HCT116 and MCF-7 cells. These results warrant further investigation of the role INPP4B-S in activation of downstream kinases and in regulation of cancer pathogenesis. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Identification and functional characterization of a voltage-gated chloride channel and its novel splice variant in taste bud cells.

    PubMed

    Huang, Liquan; Cao, Jie; Wang, Hong; Vo, Lynn A; Brand, Joseph G

    2005-10-28

    Taste bud cells are epithelial cells with neuronal properties. Voltage-dependent ion channels have been physiologically described in these cells. Here, we report the molecular identification and functional characterization of a voltage-gated chloride channel (ClC-4) and its novel splice variant (ClC-4A) from taste bud cells. ClC-4A skipped an exon near its 5'-end, incurring the loss of 60 amino acids at the N terminus. In situ hybridization and immunohistochemistry localized these two channels' transcripts and proteins to a subset of taste bud cells. Electrophysiological recordings of the heterologously expressed channels in Xenopus oocytes showed that ClC-4 and ClC-4A have opposite sensitivity to pH and unique ion selectivity. The chloride channel blockers niflumic acid and 5-nitro-2-(3-phenylpropylamino)benzoic acid had a slight or no inhibitory effect on the conductance of ClC-4, but both blockers inhibited ClC-4A, suggesting that ClC-4A is a candidate channel for an acid-induced 5-nitro-2-(3-phenylpropylamino)benzoic acid-sensitive current. Furthermore, these two channels may play a role in bitter-, sweet-, and umami-mediated taste transmission by regulating transmitter uptake into synaptic vesicles.

  4. Identification of distinct physiochemical properties of toxic prefibrillar species formed by A{beta} peptide variants

    SciTech Connect

    Goeransson, Anna-Lena; Nilsson, K. Peter R.; Kagedal, Katarina; Brorsson, Ann-Christin

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer Identification of toxic prefibrillar A{beta} species. Black-Right-Pointing-Pointer Fluorescence measurements using a combined set of fluorophores. Black-Right-Pointing-Pointer Morphology studies using transmission electron microscopy. -- Abstract: The formation of amyloid-{beta} peptide (A{beta}) aggregates at an early stage during the self-assembly process is an important factor in the development of Alzheimer's disease. The toxic effect is believed to be exerted by prefibrillar species of A{beta}. It is therefore important to identify which prefibrillar species are toxic and characterize their distinct properties. In the present study, we investigated the in vitro aggregation behavior of A{beta}-derived peptides possessing different levels of neurotoxic activity, using fluorescence spectroscopy in combination with transmission electron microscopy. The toxicity of various A{beta} aggregates was assessed by using cultures of human neuroblastoma cells. Through combined use of the fluorescence probe 8-anilino-1-napthalenesulfonate (ANS) and the novel luminescent probe pentamer formyl thiophene acetic acid (p-FTAA), we were able to identify those A{beta} peptide-derived prefibrillar species which exhibited cellular toxicity. In particular, species, which formed early during the aggregation process and showed strong p-FTAA and ANS fluorescence, were the species that possessed toxic activities. Moreover, by manipulating the aggregation conditions, it was possible to change the capacity of the A{beta} peptide to form nontoxic versus toxic species.

  5. Identification of genetic variants associated with Huntington's disease progression: a genome-wide association study.

    PubMed

    Moss, Davina J Hensman; Pardiñas, Antonio F; Langbehn, Douglas; Lo, Kitty; Leavitt, Blair R; Roos, Raymund; Durr, Alexandra; Mead, Simon; Holmans, Peter; Jones, Lesley; Tabrizi, Sarah J

    2017-09-01

    -wide significant in the meta-analysis (p=1·58 × 10(-8)), and encodes an aminoacid change (Pro67Ala) in MSH3. In TRACK-HD, each copy of the minor allele at this SNP was associated with a 0·4 units per year (95% CI 0·16-0·66) reduction in the rate of change of the Unified Huntington's Disease Rating Scale (UHDRS) Total Motor Score, and a reduction of 0·12 units per year (95% CI 0·06-0·18) in the rate of change of UHDRS Total Functional Capacity score. These associations remained significant after adjusting for age of onset. The multidomain progression measure in TRACK-HD was associated with a functional variant that was genome-wide significant in our meta-analysis. The association in only 216 participants implies that the progression measure is a sensitive reflection of disease burden, that the effect size at this locus is large, or both. Knockout of Msh3 reduces somatic expansion in Huntington's disease mouse models, suggesting this mechanism as an area for future therapeutic investigation. The European Commission FP7 NeurOmics project; CHDI Foundation; the Medical Research Council UK; the Brain Research Trust; and the Guarantors of Brain. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Identification of Genome-Wide Variants and Discovery of Variants Associated with Brassica rapa Clubroot Resistance Gene Rcr1 through Bulked Segregant RNA Sequencing.

    PubMed

    Yu, Fengqun; Zhang, Xingguo; Huang, Zhen; Chu, Mingguang; Song, Tao; Falk, Kevin C; Deora, Abhinandan; Chen, Qilin; Zhang, Yan; McGregor, Linda; Gossen, Bruce D; McDonald, Mary Ruth; Peng, Gary

    2016-01-01

    Clubroot, caused by Plasmodiophora brassicae, is an important disease on Brassica species worldwide. A clubroot resistance gene, Rcr1, with efficacy against pathotype 3 of P. brassicae, was previously mapped to chromosome A03 of B. rapa in pak choy cultivar "Flower Nabana". In the current study, resistance to pathotypes 2, 5 and 6 was shown to be associated with Rcr1 region on chromosome A03. Bulked segregant RNA sequencing was performed and short read sequences were assembled into 10 chromosomes of the B. rapa reference genome v1.5. For the resistant (R) bulks, a total of 351.8 million (M) sequences, 30,836.5 million bases (Mb) in length, produced 120-fold coverage of the reference genome. For the susceptible (S) bulks, 322.9 M sequences, 28,216.6 Mb in length, produced 109-fold coverage. In total, 776.2 K single nucleotide polymorphisms (SNPs) and 122.2 K insertion / deletion (InDels) in R bulks and 762.8 K SNPs and 118.7 K InDels in S bulks were identified; each chromosome had about 87% SNPs and 13% InDels, with 78% monomorphic and 22% polymorphic variants between the R and S bulks. Polymorphic variants on each chromosome were usually below 23%, but made up 34% of the variants on chromosome A03. There were 35 genes annotated in the Rcr1 target region and variants were identified in 21 genes. The numbers of poly variants differed significantly among the genes. Four out of them encode Toll-Interleukin-1 receptor / nucleotide-binding site / leucine-rich-repeat proteins; Bra019409 and Bra019410 harbored the higher numbers of polymorphic variants, which indicates that they are more likely candidates of Rcr1. Fourteen SNP markers in the target region were genotyped using the Kompetitive Allele Specific PCR method and were confirmed to associate with Rcr1. Selected SNP markers were analyzed with 26 recombinants obtained from a segregating population consisting of 1587 plants, indicating that they were completely linked to Rcr1. Nine SNP markers were used for marker

  7. Identification of a functional splice variant of 14-3-3E1 in rainbow trout.

    PubMed

    Wanna, Warapond; Rexroad, Caird E; Yao, Jianbo

    2010-02-01

    The 14-3-3 protein family is a family of regulatory proteins involved in diverse cellular processes. The presence of 14-3-3 isoforms and the diversity of cellular processes regulated by 14-3-3 isoforms suggest functional specificity of the isoforms. In this study, we report the identification and characterization of a new isoform of the rainbow trout 14-3-3E1 gene generated by alternative splicing. The new isoform contains an insertion of 48 nucleotides (from intron 5) in the coding region of 14-3-3E1 which results in the introduction of a premature stop codon between exon 5 and exon 6. Thus, the alternatively spliced form of 14-3-3E1 (14-3-3E1DeltaC17) lacks 17 amino acid residues at the C terminus encoded by the last exon (exon 6). Reverse-transcription polymerase chain reaction analysis revealed that the wild-type 14-3-3E1 (14-3-3E1wt) is ubiquitously expressed, while 14-3-3E1DeltaC17 shows tissue-specific as well as stage-specific expression during ovarian development and early embryogenesis. Analysis by yeast two-hybrid system demonstrated that 14-3-3E1Delta17 interacts with a number of proteins including ATP synthase, ankyrin repeat domain 13b, cytochrome c subunit VIa, cytochrome c subunit VIb, 60S ribosomal protein L34, solute carrier family 17 member 6 (SLC17A6), troponin I, and an unknown protein. Although all of these proteins except for SLC17A6 also interact with 14-3-3E1wt, 14-3-3E1Delta17 appears to have higher binding affinity with these proteins than 14-3-3E1wt. These findings suggest that alternative splicing affects the function and tissue-specific expression of 14-3-3E1.

  8. Identification and functional characterization of de novo FOXP1 variants provides novel insights into the etiology of neurodevelopmental disorder.

    PubMed

    Sollis, Elliot; Graham, Sarah A; Vino, Arianna; Froehlich, Henning; Vreeburg, Maaike; Dimitropoulou, Danai; Gilissen, Christian; Pfundt, Rolph; Rappold, Gudrun A; Brunner, Han G; Deriziotis, Pelagia; Fisher, Simon E

    2016-02-01

    De novo disruptions of the neural transcription factor FOXP1 are a recently discovered, rare cause of sporadic intellectual disability (ID). We report three new cases of FOXP1-related disorder identified through clinical whole-exome sequencing. Detailed phenotypic assessment confirmed that global developmental delay, autistic features, speech/language deficits, hypotonia and mild dysmorphic features are core features of the disorder. We expand the phenotypic spectrum to include sensory integration disorder and hypertelorism. Notably, the etiological variants in these cases include two missense variants within the DNA-binding domain of FOXP1. Only one such variant has been reported previously. The third patient carries a stop-gain variant. We performed functional characterization of the three missense variants alongside our stop-gain and two previously described truncating/frameshift variants. All variants severely disrupted multiple aspects of protein function. Strikingly, the missense variants had similarly severe effects on protein function as the truncating/frameshift variants. Our findings indicate that a loss of transcriptional repression activity of FOXP1 underlies the neurodevelopmental phenotype in FOXP1-related disorder. Interestingly, the three novel variants retained the ability to interact with wild-type FOXP1, suggesting these variants could exert a dominant-negative effect by interfering with the normal FOXP1 protein. These variants also retained the ability to interact with FOXP2, a paralogous transcription factor disrupted in rare cases of speech and language disorder. Thus, speech/language deficits in these individuals might be worsened through deleterious effects on FOXP2 function. Our findings highlight that de novo FOXP1 variants are a cause of sporadic ID and emphasize the importance of this transcription factor in neurodevelopment. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email

  9. Identification and analysis of function of a novel splicing variant of mouse receptor activator of NF-κB.

    PubMed

    Mukai, Satomi; Kitazawa, Riko; Ishii, Junko; Kondo, Takeshi; Hakozaki, Akihiro; Horiuchi, Keisuke; Haraguch, Ryuma; Mori, Kiyoshi; Kitazawa, Sohei

    2011-04-01

    Receptor activator of NF-κB (RANK) is a member of the tumor necrosis factor receptor (TNFR) family expressed in osteoclast precursors, and RANK-RANK ligand (RANKL) signaling is a key system for differentiation, activation and survival of osteoclasts. Here, we report the identification of a novel alternative splicing variant of mouse RANK gene (vRANK) that contains a new intervening exon between exon 1 and exon 2 of mouse full-length RANK (fRANK) mRNA. Since this novel exon contains the stop codon, vRANK encodes truncated amino acids that have a portion of the signal peptide of fRANK and an additional 19 amino acids that show no homology to previously reported domains. By transient transfection studies with vRANK-GFP and -Flag expressing constructs, vRANK was found localized mostly in the cytoplasm and partly in the cell membrane, but was not secreted into the culture supernatant. Under the stimulation of various factors, the expression of vRANK mRNA was almost parallel to that of fRANK in RAW264.7 cells not treated with M-CSF. Overexpression of vRANK, on the other hand, decreased TRACP (a marker of osteoclasts) mRNA expression as well as the number of TRACP-positive multinucleated giant cells. While the mRNA expression levels of NFATc1 (a master transcriptional factor of the osteoclast differentiation program) were not affected, apoptotic cells increased significantly in vRAN K-transfected cells treated with sRANKL. Taken together, these results suggest that vRANK is a novel osteoclast suppressor that reduces the number of RANKL-induced mature osteoclasts mainly by negating the anti-apoptotic effect of RANKL.

  10. Sex, Sexual Orientation, and Identification of Positive and Negative Facial Affect

    ERIC Educational Resources Information Center

    Rahman, Qazi; Wilson, Glenn D.; Abrahams, Sharon

    2004-01-01

    Sex and sexual orientation related differences in processing of happy and sad facial emotions were examined using an experimental facial emotion recognition paradigm with a large sample (N=240). Analysis of covariance (controlling for age and IQ) revealed that women (irrespective of sexual orientation) had faster reaction times than men for…

  11. Sex, Sexual Orientation, and Identification of Positive and Negative Facial Affect

    ERIC Educational Resources Information Center

    Rahman, Qazi; Wilson, Glenn D.; Abrahams, Sharon

    2004-01-01

    Sex and sexual orientation related differences in processing of happy and sad facial emotions were examined using an experimental facial emotion recognition paradigm with a large sample (N=240). Analysis of covariance (controlling for age and IQ) revealed that women (irrespective of sexual orientation) had faster reaction times than men for…

  12. Identification of Low-Level Product-Related Variants in Filgrastim Products Presently Available in Highly Regulated Markets.

    PubMed

    Hausberger, Anna; Lamanna, William C; Hartinger, Martin; Seidl, Andreas; Toll, Hansjoerg; Holzmann, Johann

    2016-06-01

    Filgrastim is a recombinant, non-glycosylated form of human granulocyte colony-stimulating factor, used to stimulate leukocyte proliferation in patients suffering from neutropenia. Since the expiration of patents associated with Amgen's filgrastim biopharmaceutical, Neupogen(®), in 2006, a number of filgrastim products have been marketed; however, a detailed characterization and comparison of variants associated with these products have not been publically reported. The objective of this study was to identify and quantify product-related variants in filgrastim reference products and biosimilars thereof that are presently available in highly regulated markets. In this study, we used intact and top-down mass spectrometry to identify and quantify product-related variants in filgrastim products. Mass spectrometry has become the method of choice for physicochemical characterization of biopharmaceuticals, allowing accurate and sensitive characterization of product-related variants. In addition to modifications ubiquitously present in biopharmaceuticals, such as methionine oxidation and asparagine/glutamine deamidation, we identified six different low-level, product-related variants present in some, but not all, of the tested products. Two variants, an acetylated filgrastim variant and a filgrastim variant containing an additional C-terminal tryptophan extension, are newly identified variants. This study demonstrates that filgrastim products already in widespread clinical use in highly regulated markets differ in low-level, product-related variants present at levels mostly below 1 % relative abundance. This study provides a comprehensive catalog of minor differences between filgrastim products and suggests that the filgrastim product-related variants described here are not clinically relevant when present at low abundance.

  13. Development of a Targeted Multi-Disorder High-Throughput Sequencing Assay for the Effective Identification of Disease-Causing Variants

    PubMed Central

    Delio, Maria; Patel, Kunjan; Maslov, Alex; Marion, Robert W.; McDonald, Thomas V.; Cadoff, Evan M.; Golden, Aaron; Greally, John M.; Vijg, Jan; Morrow, Bernice; Montagna, Cristina

    2015-01-01

    Background While next generation sequencing (NGS) is a useful tool for the identification of genetic variants to aid diagnosis and support therapy decision, high sequencing costs have limited its application within routine clinical care, especially in economically depressed areas. To investigate the utility of a multi-disease NGS based genetic test, we designed a custom sequencing assay targeting over thirty disease-associated areas including cardiac disorders, intellectual disabilities, hearing loss, collagenopathies, muscular dystrophy, Ashkenazi Jewish genetic disorders, and complex Mendelian disorders. We focused on these specific areas based on the interest of our collaborative clinical team, suggesting these diseases being the ones in need for the development of a sequencing-screening assay. Results We targeted all coding, untranslated regions (UTR) and flanking intronic regions of 650 known disease-associated genes using the Roche-NimbleGen EZ SeqCapV3 capture system and sequenced on the Illumina HiSeq 2500 Rapid Run platform. Eight controls with known variants and one HapMap sample were first sequenced to assess the performance of the panel. Subsequently, as a proof of principle and to explore the possible utility of our test, we analyzed test disease subjects (n = 16). Eight had known Mendelian disorders and eight had complex pediatric diseases. In addition to assess whether copy number variation may be of utility as a companion assay relative to these specific disease areas, we used the Affymetrix Genome-Wide SNP Array 6.0 to analyze the same samples. Conclusion We identified potentially disease-associated variants: 22 missense, 4 nonsense, 1 frameshift, and 1 splice variants (16 previously identified, 12 novel among dbSNP and 15 novel among NHLBI Exome Variant Server). We found multi-disease targeted high-throughput sequencing to be a cost efficient approach in detecting disease-associated variants to aid diagnosis. PMID:26214305

  14. Identification of a new hereditary amyloidosis prealbumin variant, Tyr-77, and detection of the gene by DNA analysis.

    PubMed Central

    Wallace, M R; Dwulet, F E; Williams, E C; Conneally, P M; Benson, M D

    1988-01-01

    In the last several years, five human plasma prealbumin (transthyretin) variants have been discovered in association with hereditary amyloidosis, a late-onset fatal disorder. We recently studied a patient of German descent with peripheral neuropathy and bowel dysfunction. Biopsied rectal tissue contained amyloid that stained with anti-human prealbumin. Amino acid sequence analysis of the patient's plasma prealbumin revealed both normal and variant prealbumin molecules, with the variant containing a tyrosine at position 77 instead of serine. We predicted a single nucleotide change in codon 77 of the variant prealbumin gene, which we then detected in the patient's DNA using the restriction enzyme SspI and a specifically tailored genomic prealbumin probe. DNA tests of other family members identified several gene carriers. This is the sixth prealbumin variant implicated in amyloidosis, and adds to the accumulating evidence that the prealbumin amyloidoses are more varied and prevalent than previously thought. Images PMID:2891727

  15. Identification and DNA sequence analysis of 15 new {alpha}{sub 1}-antitrypsin variants, including two PI*QO alleles and one deficient PI*M allele

    SciTech Connect

    Faber, J.P.; Kirchgesser, M.; Schwaab, R.; Bidlingmaier, F.; Poller, W.; Weidinger, S.; Olek, K. |

    1994-12-01

    The authors have investigated the molecular basis of 15 new {alpha}{sub 1}-antitrypsin ({alpha}1AT) variants. Phenotyping by isoelectric focusing (IEF) was used as a screening method to detect {alpha}1AT variants at the protein level. Genotyping was then performed by sequence analysis of all coding exons, exon-intron junctions, and the hepatocyte-specific promotor region including exon Ic. Three of these rare variants are alleles of clinical relevance, associated with undetectable or very low serum levels of {alpha}1AT: the PI*Q0saarbruecken allele generated by a 1-bp C-nucleotide insertion within a stretch of seven cytosines spanning residues 360-362, resulting in a 3{prime} frameshift and the acquisition of a stop codon at residue 376; a point mutation in the PI*Q0lisbon allele, resulting in a single amino acid substitution Thr{sup 68}(ACC){yields}Ile(ATC); and an in-frame trinucleotide deletion {Delta}Phe{sup 51} (TTC) in the highly deficient PI*Mpalermo allele. The remaining 12 alleles are associated with normal {alpha}1AT serum levels and are characterized by point mutations causing single amino acid substitutions in all but one case. This exception is a silent mutation, which does not affect the amino acid sequence. The limitation of IEF compared with DNA sequence analysis, for identification of new variants, their generation by mutagenesis, and the clinical relevance of the three deficiency alleles are discussed.

  16. Identification of novel host-oriented targets for Human Immunodeficiency Virus type 1 using Random Homozygous Gene Perturbation

    PubMed Central

    Mao, Hanwen; Chen, Hanson; Fesseha, Zena; Chang, Shaojing; Ung-Medoff, Huong; Van Dyke, Jessica; Kohli, Manu; Li, Wu-Bo; Goldblatt, Michael; Kinch, Michael S

    2009-01-01

    Background Human Immunodeficiency Virus (HIV) is a global threat to public health. Current therapies that directly target the virus often are rendered ineffective due to the emergence of drug-resistant viral variants. An emerging concept to combat drug resistance is the idea of targeting host mechanisms that are essential for the propagation of the virus, but have a minimal cellular effect. Results Herein, using Random Homozygous Gene Perturbation (RHGP), we have identified cellular targets that allow human MT4 cells to survive otherwise lethal infection by a wild type HIV-1NL4-3. These gene targets were validated by the reversibility of the RHGP technology, which confirmed that the RHGP itself was responsible for the resistance to HIV-1 infection. We further confirmed by siRNA knockdowns that the RHGP-identified cellular pathways are responsible for resistance to infection by either CXCR4 or CCR5 tropic HIV-1 variants. We also demonstrated that cell clones with these gene targets disrupted by RHGP were not permissible to the replication of a drug resistant HIV-1 mutant. Conclusion These studies demonstrate the power of RHGP to identify novel host targets that are essential for the viral life cycle but which can be safely perturbed without overt cytotoxicity. These findings suggest opportunities for the future development of host-oriented therapeutics with the broad spectrum potential for safe and effective inhibition of HIV infection. PMID:19788744

  17. Identification of Putative Olfactory Genes from the Oriental Fruit Moth Grapholita molesta via an Antennal Transcriptome Analysis

    PubMed Central

    Li, Yiping; Wu, Junxiang

    2015-01-01

    Background The oriental fruit moth, Grapholita molesta, is an extremely important oligophagous pest species of stone and pome fruits throughout the world. As a host-switching species, adult moths, especially females, depend on olfactory cues to a large extent in locating host plants, finding mates, and selecting oviposition sites. The identification of olfactory genes can facilitate investigation on mechanisms for chemical communications. Methodology/Principal Finding We generated transcriptome of female antennae of G.molesta using the next-generation sequencing technique, and assembled transcripts from RNA-seq reads using Trinity, SOAPdenovo-trans and Abyss-trans assemblers. We identified 124 putative olfactory genes. Among the identified olfactory genes, 118 were novel to this species, including 28 transcripts encoding for odorant binding proteins, 17 chemosensory proteins, 48 odorant receptors, four gustatory receptors, 24 ionotropic receptors, two sensory neuron membrane proteins, and one odor degrading enzyme. The identified genes were further confirmed through semi-quantitative reverse transcription PCR for transcripts coding for 26 OBPs and 17 CSPs. OBP transcripts showed an obvious antenna bias, whereas CSP transcripts were detected in different tissues. Conclusion Antennal transcriptome data derived from the oriental fruit moth constituted an abundant molecular resource for the identification of genes potentially involved in the olfaction process of the species. This study provides a foundation for future research on the molecules involved in olfactory recognition of this insect pest, and in particular, the feasibility of using semiochemicals to control this pest. PMID:26540284

  18. Identification of 83 novel alpha-mannosidosis-associated sequence variants: functional analysis of MAN2B1 missense mutations.

    PubMed

    Riise Stensland, Hilde Monica Frostad; Klenow, Helle Bagterp; Van Nguyen, Lam; Hansen, Gaute Martin; Malm, Dag; Nilssen, Øivind

    2012-03-01

    The lysosomal storage disorder alpha-mannosidosis is caused by deficiency of the enzyme lysosomal alpha-mannosidase (MAN2B1). In this study, 96 disease-associated sequence variants were identified in 130 unrelated alpha-mannosidosis patients from 30 countries. Eighty-three novel variants were detected, extending the mutation spectrum from 42 to 125. In total, 256 of the 260 mutant alleles (98.5%) were identified. Most of the variants were unique to each family, however, c.2248C>T (p.Arg750Trp) was detected in 50 patients from 16 countries, and accounted for 27.3% of the disease alleles. Haplotype analysis revealed that the c.2248T variant was present on four MAN2B1 haplotype backgrounds, where one major haplotype accounted for 95% of the alleles. The distribution of the c.2248T-associated haplotypes differed remarkably from those of the control populations, suggesting that c.2248C>T has occurred on a few ancestral haplotypes where the major haplotype subsequently has spread by founder effects. The disease-associated missense mutations were introduced into the human MAN2B1 cDNA, expressed in cell culture and assayed for MAN2B1 activity. The majority of the variants were inactive, however, ten showed MAN2B1 activity above background, and more detailed studies are necessary to further evaluate the pathogenicity of these variants. © 2011 Wiley Periodicals, Inc.

  19. Population genetic studies of the Philippine Negritos. III. Identification of the carbonic anhydrase-1 variant with CA1 Guam.

    PubMed Central

    Omoto, K; Ueda, S; Goriki, K; Takahashi, N; Misawa, S; Pagaran, I G

    1981-01-01

    Investigation of blood samples from 277 Mamanwas of northeastern Mindanao, Philippines, confirmed the concentration of the variant carbonic anhydrase-1 (CA1 3N) in this group. The frequency for the variant allele was estimated at .217 +/- .017. It occurs also in the Manobos, the Mongoloid indigenous inhabitants of the same district, although the frequency is low (.019 +/- .008). Survey of samples from other Philippine populations, including the Aeta and the Ifugao of Luzon, failed to find variants. This findings suggests different origins of the Aeta and the Mamanwa, although both are usually referred to as Negritos. The Ca1 3N protein was purified by affinity chromatography using azosulfonamide and rechromatography on a DEAE-Sephadex column. The tryptic peptide pattern of CA1 3N was similar to that of CA1 Guam already reported. Furthermore, amino acid analyses of the tryptic peptides indicated that CA1 3N is characterized by the substitution 253 Gly leads to Arg, confirming the identity of this variant with CA1 Guam. The widespread occurrence of CA1 3 variants in the Western Pacific suggests that this variant was once common in an aboriginal population of this region, from which it was scattered by gene flow. Images Fig. 1 Fig. 2 PMID:6781336

  20. Effect of selective and distributed training on visual identification of orientation.

    PubMed

    Tschopp-Junker, Chantal; Gentaz, Edouard; Viviani, Paolo

    2010-05-01

    An experiment contrasted the effect of four training schedules in a visual orientation reproduction task. Two selective schedules involved repeated presentation of a single target orientation. Two non-selective schedules involved targets covering the first quadrant either at fixed, equispaced orientations, or distributed randomly. In pre-training sessions, we observed the classical oblique effect (precision for vertical and horizontal stimuli higher than for oblique ones). Practice improved precision with both distributed schedules, but was ineffectual for non-selective schedules. However, a significant oblique effect persisted under all conditions. We argue that the pattern of results is compatible with the hypothesis that the oblique effect reflects both the intrinsic neuronal properties of the primary visual system, and the structure of the visual space imposed by higher, more cognitive processes. The results challenge the thesis that only attentional and post-perceptual factors are able to affect the working of the early visual system.

  1. Lepton identification at particle flow oriented detector for the future e+e- Higgs factories

    NASA Astrophysics Data System (ADS)

    Yu, Dan; Ruan, Manqi; Boudry, Vincent; Videau, Henri

    2017-09-01

    The lepton identification is essential for the physics programs at high-energy frontier, especially for the precise measurement of the Higgs boson. For this purpose, a toolkit for multivariate data analysis (TMVA) based lepton identification (LICH) has been developed for detectors using high granularity calorimeters. Using the conceptual detector geometry for the Circular Electron-Positron Collider (CEPC) and single charged particle samples with energy larger than 2 GeV, LICH identifies electrons/muons with efficiencies higher than 99.5% and controls the mis-identification rate of hadron to muons/electrons to better than 1/0.5%. Reducing the calorimeter granularity by 1-2 orders of magnitude, the lepton identification performance is stable for particles with E > 2 GeV. Applied to fully simulated eeH/μ μ H events, the lepton identification performance is consistent with the single particle case: the efficiency of identifying all the high energy leptons in an event, is 95.5-98.5%.

  2. Silencing of two alternative splicing-derived mRNA variants of chitin synthase 1 gene by RNAi is lethal to the oriental migratory locust, Locusta migratoria manilensis (Meyen).

    PubMed

    Zhang, Jianzhen; Liu, Xiaojian; Zhang, Jianqin; Li, Daqi; Sun, Yi; Guo, Yaping; Ma, Enbo; Zhu, Kun Yan

    2010-11-01

    Chitin synthases are crucial enzymes responsible for chitin biosynthesis in fungi, nematodes and arthropods. We characterized two alternative splicing-derived variants of chitin synthase 1 gene (LmCHS1) from the oriental migratory locust, Locusta migratoria manilensis (Meyen). Each cDNA of the two variants (LmCHS1A and LmCHS1B) consists of 5116 nucleotides that include a 4728-nucleotide open reading frame (ORF) encoding 1576 amino acid residues, and 67- and 321-bp non-coding regions at the 5'- and 3'-ends of the cDNA, respectively. The two variants differ only in one exon consisting of 177 nucleotides that encode 59 amino acid residues. The amino acid sequences within this alternative splicing region are 75% identical between the two variants. Both variants were expressed in all the developmental stages. However, LmCHS1A was predominately expressed in the integument whereas LmCHS1B was mainly expressed in the trachea. Our RNAi-based gene silencing study resulted in a dramatic reduction in the levels of the corresponding mRNA in the locust nymphs injected with dsRNA of LmCHS1, or either of its two variants, LmCHS1A and LmCHS1B. Consequentially, 95, 88 and 51% of mortalities were observed in the locusts injected with the LmCHS1, LmCHS1A and LmCHS1B dsRNA, respectively. The phenotypes resulted from the injection of LmCHS1A dsRNA were similar to those from the injection of LmCHS1 dsRNA, whereas the locusts injected with LmCHS1B dsRNA exhibited crimpled cuticle phenotype. Our results suggest that both variants of chitin synthase 1 are essential for insect growth and development.

  3. Identification of a variant antigenic neutralizing epitope in hypervariable region 1 of avian leukosis virus subgroup J.

    PubMed

    Hou, Minbo; Zhou, Defang; Li, Gen; Guo, Huijun; Liu, Jianzhu; Wang, Guihua; Zheng, Qiankun; Cheng, Ziqiang

    2016-03-08

    Avian leukosis virus subgroup J (ALV-J) is a hypervariable oncogenic retrovirus that causes great economic loss in poultry. Antigenic variations in the variable regions make the development of an effective vaccine a challenging task. In the present study, we identified a variant antigenic neutralizing epitope using reverse vaccinology methods. First, we predicted the B-cell epitopes in gp85 gene of ALV-J strains by DNAman and bioinformatics. Fourteen candidate epitopes were selected and linked in tandem with glycines or serines as a multi-epitope gene. The expressed protein of multi-epitope gene can induce high-titer antibody that can recognize nature ALV-J and neutralize the infectivity of ALV-J strains. Next, we identified a high effective epitope using eight overlapping fragments of gp85 gene reacting with mAb 2D5 and anti-multi-epitope sera. The identified epitope contained one of the predicted epitopes and localized in hyervariable region 1 (hr1), indicating a variant epitope. To better understand if the variants of the epitope have a good antigenicity, we synthesized four variants to react with mAb 2D5 and anti-ALV-J sera. The result showed that all variants could react with the two kinds of antibodies though they showed different antigenicity, while could not react with ALV-J negative sera. Thus, the variant antigenic neutralizing epitope was determined as 137-LRDFIA/E/TKWKS/GDDL/HLIRPYVNQS-158. The result shows a potential use of this variant epitopes as a novel multi-epitope vaccine against ALV-J in poultry.

  4. Identification of Genetic Variants Linking Protein C and Lipoprotein Metabolism: The ARIC Study (Atherosclerosis Risk in Communities).

    PubMed

    Pankow, James S; Tang, Weihong; Pankratz, Nathan; Guan, Weihua; Weng, Lu-Chen; Cushman, Mary; Boerwinkle, Eric; Folsom, Aaron R

    2017-03-01

    Previous studies have identified common genetic variants in 4 chromosomal regions that together account for 14% to 15% of the variance in circulating levels of protein C. To further characterize the genetic architecture of protein C, we obtained denser coverage at some loci, extended investigation of protein C to low-frequency and rare variants, and searched for new associations in genes known to influence protein C. Genetic associations with protein C antigen level were evaluated in ≤10 778 European and 3190 black participants aged 45 to 64 years. Analyses included >26 million autosomal variants available after imputation to the 1000 Genomes reference panel along with additional low-frequency and rare variants directly genotyped using the Illumina ITMAT-Broad-CARe chip and Illumina HumanExome BeadChip. Genome-wide significant associations (P<5×10(-8)) were found for common variants in the GCKR, PROC, BAZ1B, and PROCR-EDEM2 regions in whites and PROC and PROCR-EDEM2 regions in blacks, confirming earlier findings. In a novel finding, the low-density lipoprotein cholesterol-lowering allele of rs12740374, located in the CELSR2-PSRC1-SORT1 region, was associated with lower protein C level in both whites and blacks, reaching genome-wide significance in a meta-analysis combining results from both groups (P=1.4×10(-9)). To further investigate a possible link between lipid metabolism and protein C level, we conducted Mendelian randomization analyses using 185 lipid-related genetic variants as instrumental variables. The results indicated that triglycerides, and possibly low-density lipoprotein cholesterol, influence protein C levels. Discovery of variants influencing circulating protein C levels in the CELSR2-PSRC1-SORT1 region may indicate a novel genetic link between lipoprotein metabolism and hemostasis. © 2017 American Heart Association, Inc.

  5. Postmortem genetic screening for the identification, verification, and reporting of genetic variants contributing to the sudden death of the young.

    PubMed

    Methner, D Nicole R; Scherer, Steven E; Welch, Katherine; Walkiewicz, Magdalena; Eng, Christine M; Belmont, John W; Powell, Mark C; Korchina, Viktoriya; Doddapaneni, Harsha Vardhan; Muzny, Donna M; Gibbs, Richard A; Wolf, Dwayne A; Sanchez, Luis A; Kahn, Roger

    2016-09-01

    Each year in the United States, thousands of cases of sudden and unexpected deaths of infants, children, and young adults are assigned an undetermined cause of death after postmortem investigation and autopsy. Heritable genetic variants have been suggested as the cause of up to a third of sudden death (SD) cases. Elucidation of the genetic variants involved in SD cases is important to not only help establish cause and manner of death of these individuals, but to also aid in determining whether familial genetic testing should be considered. Previously, these types of postmortem screenings have not been a feasible option for most county medical examiners' and coroners' offices. We sequenced full exons of 64 genes associated with SD in the largest known cohort (351) of infant and young SD decedents using massively parallel sequencing at <$600 per sample. Genetic variants were assessed through literature review and clinical evaluation by a multidisciplinary consortium of experts. Thirteen individuals (3.7%), eight infants (2.8% of those <1 yr of age) and five children/young adults (7.0% of those >1 yr of age), were found to have a reportable genetic variant contributing to SD. These percentages represent an estimate lower than those previously reported. Overall yields and results likely vary between studies due to differences in evaluation techniques and reporting. Additionally, we recommend ongoing assessment of data, including nonreported novel variants, as technology and literature continually advance. This study demonstrates a strategy to implement molecular autopsies in medicolegal investigations of young SD decedents.

  6. Expanding the mutation spectrum in 182 Spanish probands with craniosynostosis: identification and characterization of novel TCF12 variants.

    PubMed

    Paumard-Hernández, Beatriz; Berges-Soria, Julia; Barroso, Eva; Rivera-Pedroza, Carlos I; Pérez-Carrizosa, Virginia; Benito-Sanz, Sara; López-Messa, Eva; Santos, Fernando; García-Recuero, Ignacio I; Romance, Ana; Ballesta-Martínez, Juliana María; López-González, Vanesa; Campos-Barros, Ángel; Cruz, Jaime; Guillén-Navarro, Encarna; Sánchez Del Pozo, Jaime; Lapunzina, Pablo; García-Miñaur, Sixto; Heath, Karen E

    2015-07-01

    Craniosynostosis, caused by the premature fusion of one or more of the cranial sutures, can be classified into non-syndromic or syndromic and by which sutures are affected. Clinical assignment is a difficult challenge due to the high phenotypic variability observed between syndromes. During routine diagnostics, we screened 182 Spanish craniosynostosis probands, implementing a four-tiered cascade screening of FGFR2, FGFR3, FGFR1, TWIST1 and EFNB1. A total of 43 variants, eight novel, were identified in 113 (62%) patients: 104 (92%) detected in level 1; eight (7%) in level 2 and one (1%) in level 3. We subsequently screened additional genes in the probands with no detected mutation: one duplication of the IHH regulatory region was identified in a patient with craniosynostosis Philadelphia type and five variants, four novel, were identified in the recently described TCF12, in probands with coronal or multisuture affectation. In the 19 Saethre-Chotzen syndrome (SCS) individuals in whom a variant was detected, 15 (79%) carried a TWIST1 variant, whereas four (21%) had a TCF12 variant. Thus, we propose that TCF12 screening should be included for TWIST1 negative SCS patients and in patients where the coronal suture is affected. In summary, a molecular diagnosis was obtained in a total of 119/182 patients (65%), allowing the correct craniosynostosis syndrome classification, aiding genetic counselling and in some cases provided a better planning on how and when surgical intervention should take place and, subsequently the appropriate clinical follow up.

  7. Expanding the mutation spectrum in 182 Spanish probands with craniosynostosis: identification and characterization of novel TCF12 variants

    PubMed Central

    Paumard-Hernández, Beatriz; Berges-Soria, Julia; Barroso, Eva; Rivera-Pedroza, Carlos I; Pérez-Carrizosa, Virginia; Benito-Sanz, Sara; López-Messa, Eva; Santos, Fernando; García-Recuero, Ignacio I; Romance, Ana; Ballesta-Martínez, Juliana María; López-González, Vanesa; Campos-Barros, Ángel; Cruz, Jaime; Guillén-Navarro, Encarna; Sánchez del Pozo, Jaime; Lapunzina, Pablo; García-Miñaur, Sixto; Heath, Karen E

    2015-01-01

    Craniosynostosis, caused by the premature fusion of one or more of the cranial sutures, can be classified into non-syndromic or syndromic and by which sutures are affected. Clinical assignment is a difficult challenge due to the high phenotypic variability observed between syndromes. During routine diagnostics, we screened 182 Spanish craniosynostosis probands, implementing a four-tiered cascade screening of FGFR2, FGFR3, FGFR1, TWIST1 and EFNB1. A total of 43 variants, eight novel, were identified in 113 (62%) patients: 104 (92%) detected in level 1; eight (7%) in level 2 and one (1%) in level 3. We subsequently screened additional genes in the probands with no detected mutation: one duplication of the IHH regulatory region was identified in a patient with craniosynostosis Philadelphia type and five variants, four novel, were identified in the recently described TCF12, in probands with coronal or multisuture affectation. In the 19 Saethre–Chotzen syndrome (SCS) individuals in whom a variant was detected, 15 (79%) carried a TWIST1 variant, whereas four (21%) had a TCF12 variant. Thus, we propose that TCF12 screening should be included for TWIST1 negative SCS patients and in patients where the coronal suture is affected. In summary, a molecular diagnosis was obtained in a total of 119/182 patients (65%), allowing the correct craniosynostosis syndrome classification, aiding genetic counselling and in some cases provided a better planning on how and when surgical intervention should take place and, subsequently the appropriate clinical follow up. PMID:25271085

  8. Study designs for identification of rare disease variants in complex diseases: the utility of family-based designs.

    PubMed

    Ionita-Laza, Iuliana; Ottman, Ruth

    2011-11-01

    The recent progress in sequencing technologies makes possible large-scale medical sequencing efforts to assess the importance of rare variants in complex diseases. The results of such efforts depend heavily on the use of efficient study designs and analytical methods. We introduce here a unified framework for association testing of rare variants in family-based designs or designs based on unselected affected individuals. This framework allows us to quantify the enrichment in rare disease variants in families containing multiple affected individuals and to investigate the optimal design of studies aiming to identify rare disease variants in complex traits. We show that for many complex diseases with small values for the overall sibling recurrence risk ratio, such as Alzheimer's disease and most cancers, sequencing affected individuals with a positive family history of the disease can be extremely advantageous for identifying rare disease variants. In contrast, for complex diseases with large values of the sibling recurrence risk ratio, sequencing unselected affected individuals may be preferable.

  9. Identification as Incentive to Care: Pre-Service Teachers' Orientation towards Inclusion in Nature

    ERIC Educational Resources Information Center

    Gedžune, Inga

    2015-01-01

    This study proposes identification with nature as a pedagogical technique in teacher education for sustainability to facilitate commitment to care about nonhuman nature, such care being an important underpinning of an inclusive and hence sustainable human-nature relationship. 29 pre-service teachers participated in a series of learning activities…

  10. Identification as Incentive to Care: Pre-Service Teachers' Orientation towards Inclusion in Nature

    ERIC Educational Resources Information Center

    Gedžune, Inga

    2015-01-01

    This study proposes identification with nature as a pedagogical technique in teacher education for sustainability to facilitate commitment to care about nonhuman nature, such care being an important underpinning of an inclusive and hence sustainable human-nature relationship. 29 pre-service teachers participated in a series of learning activities…

  11. Identification of a Novel De Novo Variant in the PAX3 Gene in Waardenburg Syndrome by Diagnostic Exome Sequencing: The First Molecular Diagnosis in Korea

    PubMed Central

    Jang, Mi-Ae; Lee, Taeheon; Lee, Junnam

    2015-01-01

    Waardenburg syndrome (WS) is a clinically and genetically heterogeneous hereditary auditory pigmentary disorder characterized by congenital sensorineural hearing loss and iris discoloration. Many genes have been linked to WS, including PAX3, MITF, SNAI2, EDNRB, EDN3, and SOX10, and many additional genes have been associated with disorders with phenotypic overlap with WS. To screen all possible genes associated with WS and congenital deafness simultaneously, we performed diagnostic exome sequencing (DES) in a male patient with clinical features consistent with WS. Using DES, we identified a novel missense variant (c.220C>G; p.Arg74Gly) in exon 2 of the PAX3 gene in the patient. Further analysis by Sanger sequencing of the patient and his parents revealed a de novo occurrence of the variant. Our findings show that DES can be a useful tool for the identification of pathogenic gene variants in WS patients and for differentiation between WS and similar disorders. To the best of our knowledge, this is the first report of genetically confirmed WS in Korea. PMID:25932447

  12. DIVAN: accurate identification of non-coding disease-specific risk variants using multi-omics profiles.

    PubMed

    Chen, Li; Jin, Peng; Qin, Zhaohui S

    2016-12-06

    Understanding the link between non-coding sequence variants, identified in genome-wide association studies, and the pathophysiology of complex diseases remains challenging due to a lack of annotations in non-coding regions. To overcome this, we developed DIVAN, a novel feature selection and ensemble learning framework, which identifies disease-specific risk variants by leveraging a comprehensive collection of genome-wide epigenomic profiles across cell types and factors, along with other static genomic features. DIVAN accurately and robustly recognizes non-coding disease-specific risk variants under multiple testing scenarios; among all the features, histone marks, especially those marks associated with repressed chromatin, are often more informative than others.

  13. Identification of Novel Crk-associated Substrate (p130Cas) Variants with Functionally Distinct Focal Adhesion Kinase Binding Activities*

    PubMed Central

    Kumbrink, Joerg; Soni, Shefali; Laumbacher, Barbara; Loesch, Barbara; Kirsch, Kathrin H.

    2015-01-01

    Elevated levels of p130Cas (Crk-associated substrate)/BCAR1 (breast cancer antiestrogen resistance 1 gene) are associated with aggressiveness of breast tumors. Following phosphorylation of its substrate domain, p130Cas promotes the integration of protein complexes involved in multiple signaling pathways and mediates cell proliferation, adhesion, and migration. In addition to the known BCAR1-1A (wild-type) and 1C variants, we identified four novel BCAR1 mRNA variants, generated by alternative first exon usage (1B, 1B1, 1D, and 1E). Exons 1A and 1C encode for four amino acids (aa), whereas 1D and 1E encode for 22 aa and 1B1 encodes for 50 aa. Exon 1B is non-coding, resulting in a truncated p130Cas protein (Cas1B). BCAR1-1A, 1B1, and variant 1C mRNAs were ubiquitously expressed in cell lines and a survey of human tissues, whereas 1B, 1D, and 1E expression was more restricted. Reconstitution of all isoforms except for 1B in p130Cas-deficient murine fibroblasts induced lamellipodia formation and membrane ruffling, which was unrelated to the substrate domain phosphorylation status. The longer isoforms exhibited increased binding to focal adhesion kinase (FAK), a molecule important for migration and adhesion. The shorter 1B isoform exhibited diminished FAK binding activity and significantly reduced migration and invasion. In contrast, the longest variant 1B1 established the most efficient FAK binding and greatly enhanced migration. Our results indicate that the p130Cas exon 1 variants display altered functional properties. The truncated variant 1B and the longer isoform 1B1 may contribute to the diverse effects of p130Cas on cell biology and therefore will be the target of future studies. PMID:25805500

  14. Identification of Novel Crk-associated Substrate (p130Cas) Variants with Functionally Distinct Focal Adhesion Kinase Binding Activities.

    PubMed

    Kumbrink, Joerg; Soni, Shefali; Laumbacher, Barbara; Loesch, Barbara; Kirsch, Kathrin H

    2015-05-08

    Elevated levels of p130(Cas) (Crk-associated substrate)/BCAR1 (breast cancer antiestrogen resistance 1 gene) are associated with aggressiveness of breast tumors. Following phosphorylation of its substrate domain, p130(Cas) promotes the integration of protein complexes involved in multiple signaling pathways and mediates cell proliferation, adhesion, and migration. In addition to the known BCAR1-1A (wild-type) and 1C variants, we identified four novel BCAR1 mRNA variants, generated by alternative first exon usage (1B, 1B1, 1D, and 1E). Exons 1A and 1C encode for four amino acids (aa), whereas 1D and 1E encode for 22 aa and 1B1 encodes for 50 aa. Exon 1B is non-coding, resulting in a truncated p130(Cas) protein (Cas1B). BCAR1-1A, 1B1, and variant 1C mRNAs were ubiquitously expressed in cell lines and a survey of human tissues, whereas 1B, 1D, and 1E expression was more restricted. Reconstitution of all isoforms except for 1B in p130(Cas)-deficient murine fibroblasts induced lamellipodia formation and membrane ruffling, which was unrelated to the substrate domain phosphorylation status. The longer isoforms exhibited increased binding to focal adhesion kinase (FAK), a molecule important for migration and adhesion. The shorter 1B isoform exhibited diminished FAK binding activity and significantly reduced migration and invasion. In contrast, the longest variant 1B1 established the most efficient FAK binding and greatly enhanced migration. Our results indicate that the p130(Cas) exon 1 variants display altered functional properties. The truncated variant 1B and the longer isoform 1B1 may contribute to the diverse effects of p130(Cas) on cell biology and therefore will be the target of future studies. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. Identification of a functional variant in the KIF5A-CYP27B1-METTL1-FAM119B locus associated with multiple sclerosis

    PubMed Central

    Alcina, Antonio; Fedetz, Maria; Fernández, Óscar; Saiz, Albert; Izquierdo, Guillermo; Lucas, Miguel; Leyva, Laura; García-León, Juan-Antonio; Abad-Grau, María del Mar; Alloza, Iraide; Antigüedad, Alfredo; Garcia-Barcina, María J; Vandenbroeck, Koen; Varadé, Jezabel; de la Hera, Belén; Arroyo, Rafael; Comabella, Manuel; Montalban, Xavier; Petit-Marty, Natalia; Navarro, Arcadi; Otaegui, David; Olascoaga, Javier; Blanco, Yolanda; Urcelay, Elena; Matesanz, Fuencisla

    2013-01-01

    Background and aim Several studies have highlighted the association of the 12q13.3–12q14.1 region with coeliac disease, type 1 diabetes, rheumatoid arthritis and multiple sclerosis (MS); however, the causal variants underlying diseases are still unclear. The authors sought to identify the functional variant of this region associated with MS. Methods Tag-single nucleotide polymorphism (SNP) analysis of the associated region encoding 15 genes was performed in 2876 MS patients and 2910 healthy Caucasian controls together with expression regulation analyses. Results rs6581155, which tagged 18 variants within a region where 9 genes map, was sufficient to model the association. This SNP was in total linkage disequilibrium (LD) with other polymorphisms that associated with the expression levels of FAM119B, AVIL, TSFM, TSPAN31 and CYP27B1 genes in different expression quantitative trait loci studies. Functional annotations from Encyclopedia of DNA Elements (ENCODE) showed that six out of these rs6581155-tagged-SNPs were located in regions with regulatory potential and only one of them, rs10877013, exhibited allele-dependent (ratio A/G=9.5-fold) and orientation-dependent (forward/reverse=2.7-fold) enhancer activity as determined by luciferase reporter assays. This enhancer is located in a region where a long-range chromatin interaction among the promoters and promoter-enhancer of several genes has been described, possibly affecting their expression simultaneously. Conclusions This study determines a functional variant which alters the enhancer activity of a regulatory element in the locus affecting the expression of several genes and explains the association of the 12q13.3–12q14.1 region with MS. PMID:23160276

  16. Identification of a novel nonsense mutation in POLH in a Chinese pedigree with xeroderma pigmentosum, variant type.

    PubMed

    Liu, Xiaoyan; Zhang, Xianning; Qiao, Jianjun; Fang, Hong

    2013-01-01

    Xeroderma pigmentosum-variant (XPV) is one type of XP, a rare autosomal recessive disorder, and caused by defects in the post replication repair machinery while nucleotide-excision repair (NER) is not impaired. In the present study, we reported a Chinese family with XPV phenotype, which was confirmed by histopathological results. Genetic variants were detected by polymerase chain reaction and exon sequencing. Furthermore, the reported molecular defects in XPV patients from previous literatures were reviewed. A homozygous c.67C>T mutation in the exon 2 of DNA polymerase eta (POLH), a novel non-sense mutation in POLH, was discovered.

  17. Identification of two novel Mycobacterium avium allelic variants in pig and human isolates from Brazil by PCR-restriction enzyme analysis.

    PubMed

    Leão, S C; Briones, M R; Sircili, M P; Balian, S C; Mores, N; Ferreira-Neto, J S

    1999-08-01

    Mycobacterium avium complex (MAC) is composed of environmental mycobacteria found widely in soil, water, and aerosols that can cause disease in animals and humans, especially disseminated infections in AIDS patients. MAC consists of two closely related species, M. avium and M. intracellulare, and may also include other, less-defined groups. The precise differentiation of MAC species is a fundamental step in epidemiological studies and for the evaluation of possible reservoirs for MAC infection in humans and animals. In this study, which included 111 pig and 26 clinical MAC isolates, two novel allelic M. avium PCR-restriction enzyme analysis (PRA) variants were identified, differing from the M. avium PRA prototype in the HaeIII digestion pattern. Mutations in HaeIII sites were confirmed by DNA sequencing. Identification of these isolates as M. avium was confirmed by PCR with DT1-DT6 and IS1245 primers, nucleic acid hybridization with the AccuProbe system, 16S ribosomal DNA sequencing, and biochemical tests. The characterization of M. avium PRA variants can be useful in the elucidation of factors involved in mycobacterial virulence and routes of infection and also has diagnostic significance, since they can be misidentified as M. simiae II and M. kansasii I if the PRA method is used in the clinical laboratory for identification of mycobacteria.

  18. Computerized image analysis: Texture-field orientation method for pectoral muscle identification on MLO-view mammograms

    SciTech Connect

    Zhou Chuan; Wei Jun; Chan, Heang-Ping; Paramagul, Chintana; Hadjiiski, Lubomir M.; Sahiner, Berkman; Douglas, Julie A.

    2010-05-15

    Purpose: To develop a new texture-field orientation (TFO) method that combines a priori knowledge, local and global information for the automated identification of pectoral muscle on mammograms. Methods: The authors designed a gradient-based directional kernel (GDK) filter to enhance the linear texture structures, and a gradient-based texture analysis to extract a texture orientation image that represented the dominant texture orientation at each pixel. The texture orientation image was enhanced by a second GDK filter for ridge point extraction. The extracted ridge points were validated and the ridges that were less likely to lie on the pectoral boundary were removed automatically. A shortest-path finding method was used to generate a probability image that represented the likelihood that each remaining ridge point lay on the true pectoral boundary. Finally, the pectoral boundary was tracked by searching for the ridge points with the highest probability lying on the pectoral boundary. A data set of 130 MLO-view digitized film mammograms (DFMs) from 65 patients was used to train the TFO algorithm. An independent data set of 637 MLO-view DFMs from 562 patients was used to evaluate its performance. Another independent data set of 92 MLO-view full field digital mammograms (FFDMs) from 92 patients was used to assess the adaptability of the TFO algorithm to FFDMs. The pectoral boundary detection accuracy of the TFO method was quantified by comparison with an experienced radiologist's manually drawn pectoral boundary using three performance metrics: The percent overlap area (POA), the Hausdorff distance (Hdist), and the average distance (AvgDist). Results: The mean and standard deviation of POA, Hdist, and AvgDist were 95.0{+-}3.6%, 3.45{+-}2.16 mm, and 1.12{+-}0.82 mm, respectively. For the POA measure, 91.5%, 97.3%, and 98.9% of the computer detected pectoral muscles had POA larger than 90%, 85%, and 80%, respectively. For the distance measures, 85.4% and 98.0% of the

  19. Identification of a novel alternative splicing transcript variant of the suppressor of fused: Relationship with lymph node metastasis in pancreatic ductal adenocarcinoma.

    PubMed

    Xu, Qing; Gao, Jun; Li, Zhaoshen

    2016-12-01

    Pancreatic ductal adenocarcinoma (PDAC) is one of the most fatal diseases, and the median survival time is very short. Upregulation of hedgehog signaling pathway activity is a vital factor in pathogenesis of PDAC. However, as a negative regulator of hedgehog signaling, it is not very clear what role the suppressor of fused (SUFU) plays in PDAC tissue. In our study for the identification of alternative splicing transcripts of SUFU gene in human PDAC cells, a novel transcript variant of SUFU (SUFUvN) was discovered by 3' rapid amplification of cDNA ends (3'RACE) and cDNA clone. SUFUvN contained an additional new protein-coding exon compared with the transcript variant 1 of SUFU (SUFUv1, NM_016169) published in NCBI website. The sequence of the new protein-coding exon was the same as a fragment of intron between exon 10 and 11 of SUFUv1. Thus, an exon skipping occurred in transcription of SUFUv1. Compared with the expression vector of SUFUvN transfected PDAC cells, the corresponding protein expression encoded by SUFUvN (SUFU isoform N) was detected in PDAC tissue. Furthermore, it was observed that elevated SUFUvN transcription level was related with lymph node metastasis in PDAC tissues, while neither SUFUv1 nor transcript variant 2 of SUFU (SUFUv2, NM_001178133) did. Our data indicate that there exists a novel transcript variant of SUFU which can be transcribed and translated into corresponding protein and its transcription is related with metastasis of lymph nodes in PDAC.

  20. Whole-genome re-sequencing for the identification of high contribution susceptibility gene variants in patients with type 2 diabetes

    PubMed Central

    SUN, XIAOJUAN; SUI, WEIGUO; WANG, XIAOBING; HOU, XIANLIANG; OU, MINGLIN; DAI, YONG; XIANG, YUEYING

    2016-01-01

    There is increasing evidence that several genes are associated with an increased risk of type 2 diabetes (T2D); genome-wide association investigations and whole-genome re-sequencing investigations offer a useful approach for the identification of genes involved in common human diseases. To further investigate which polymorphisms confer susceptibility to T2D, the present study screened for high-contribution susceptibility gene variants Chinese patients with T2D using whole-genome re-sequencing with DNA pooling. In total, 100 Chinese individuals with T2D and 100 healthy Chinese individuals were analyzed using whole-genome re-sequencing using DNA pooling. To minimize the likelihood of systematic bias in sampling, paired-end libraries with an insert size of 500 bp were prepared for in T2D in all samples, which were then subjected to whole-genome sequencing. Each library contained four lanes. The average sequencing depth was 35.70. In the present study, 1.36 GB of clean sequence data were generated, and the resulting calculated T2D genome consensus sequence covered 99.88% of the hg19 sequence. A total of 3,974,307 single nucleotide polymorphisms were identified, of which 99.88% were in the dbSNP database. The present study also found 642,189 insertions and deletions, 5,590 structure variants (SVs), 4,713 copy number variants (CNVs) and 13,049 single nucleotide variants. A total of 1,884 somatic CNVs and 74 somatic SVs were significantly different between the cases and controls. Therefore, the present study provided validation of whole-genome re-sequencing using the DNA pooling approach. It also generated a whole-genome re-sequencing genotype database for future investigations of T2D. PMID:27035118

  1. Identification of a novel pathogenic OTOF variant causative of nonsyndromic hearing loss with high frequency in the Ashkenazi Jewish population

    PubMed Central

    Fedick, Anastasia M; Jalas, Chaim; Swaroop, Ananya; Smouha, Eric E; Webb, Bryn D

    2016-01-01

    Mutations in the OTOF gene have previously been shown to cause nonsyndromic prelingual deafness (DFNB9, OMIM 601071) as well as auditory neuropathy/dys-synchrony. In this study, the OTOF NM_194248.2 c.5332G>T, p.Val1778Phe variant was identified in a large Ashkenazi Jewish family as the causative variant in four siblings with hearing loss. Our analysis reveals a carrier frequency of the OTOF c.5332G>T, p.Val1778Phe variant of 1.27% in the Ashkenazi Jewish population, suggesting that this variant may be a significant contributor to nonsyndromic sensorineural hearing loss and should be considered for inclusion in targeted hearing loss panels for this population. Of note, the degree of hearing loss associated with this phenotype ranged from mild to moderately severe, with two of the four siblings not known to have hearing loss until they were genotyped and underwent pure tone audiometry and auditory brainstem response testing. The phenotypic variability along with the auditory neuropathy/dys-synchrony, which allows for the production of otoacoustic emissions, supports that nonsyndromic hearing loss caused by OTOF mutations may be much more common in the Ashkenazi Jewish population than currently appreciated due to a lack of diagnosis. PMID:27621663

  2. Identification of genetic variants associated with maize flowering time using an extremely large multi-genetic background population

    USDA-ARS?s Scientific Manuscript database

    Flowering time is one of the major adaptive traits in domestication of maize and an important selection criterion in breeding. To detect more maize flowering time variants we evaluated flowering time traits using an extremely large multi- genetic background population that contained more than 8000 l...

  3. Identification of rare variants of DSP gene in Sudden Unexplained Nocturnal Death Syndrome in the southern Chinese Han population

    PubMed Central

    Zhao, Qianhao; Chen, Yili; Peng, Longlun; Gao, Rui; Liu, Nian; Jiang, Pingping; Liu, Chao; Tang, Shuangbo

    2016-01-01

    Sudden unexplained nocturnal death syndrome (SUNDS) is a perplexing disorder to both forensic pathologists and clinic physicians. Desmoplakin (DSP) gene was the first desmosomal gene linked to arrhythmogenic right ventricular cardiomyopathy (ARVC) which was associated with sudden death. To identify the genetic variants of the DSP gene in SUNDS in the southern Chinese Han population, we genetically screened the DSP gene in 40 sporadic SUNDS victims, 16 Brugada syndrome (BrS) patients and 2 Early Repolarization syndrome (ERS) patients using Next Generation Sequencing (NSG) and direct Sanger sequencing. A total of 10 genetic variants of the DSP gene were detected in 11 cases, comprised of two novel missense mutations (p.I125F and p.D521A) and eight previously reported rare variants. Of eight reported variants, two were previously considered pathogenic (p.Q90R and p.R2639Q), three were predicted in silico to bepathogenic (p.R315C, p.E1357D and p.D2579H), and the rest three were predicted to be benign (p.N1234S, p.R1308Q and p.T2267S). This is the first report of DSP genetic screening in Chinese SUNDS and Brugada syndrome. Our results implies that DSP mutations contribute to the genetic cause of some SUNDS victims and maybe a new susceptible gene for Brugada syndrome. PMID:26585738

  4. Identification of rare variants of DSP gene in sudden unexplained nocturnal death syndrome in the southern Chinese Han population.

    PubMed

    Zhao, Qianhao; Chen, Yili; Peng, Longlun; Gao, Rui; Liu, Nian; Jiang, Pingping; Liu, Chao; Tang, Shuangbo; Quan, Li; Makielski, Jonathan C; Cheng, Jianding

    2016-03-01

    Sudden unexplained nocturnal death syndrome (SUNDS) is a perplexing disorder to both forensic pathologists and clinic physicians. Desmoplakin (DSP) gene was the first desmosomal gene linked to arrhythmogenic right ventricular cardiomyopathy (ARVC) which was associated with sudden death. To identify the genetic variants of the DSP gene in SUNDS in the southern Chinese Han population, we genetically screened the DSP gene in 40 sporadic SUNDS victims, 16 Brugada syndrome (BrS) patients, and 2 early repolarization syndrome (ERS) patients using next generation sequencing (NSG) and direct Sanger sequencing. A total of 10 genetic variants of the DSP gene were detected in 11 cases, comprised of two novel missense mutations (p.I125F and p.D521A) and eight previously reported rare variants. Of eight reported variants, two were previously considered pathogenic (p.Q90R and p.R2639Q), three were predicted in silico to be pathogenic (p.R315C, p.E1357D and p.D2579H), and the rest three were predicted to be benign (p.N1234S, p.R1308Q, and p.T2267S). This is the first report of DSP genetic screening in Chinese SUNDS and Brugada syndrome. Our results imply that DSP mutations contribute to the genetic cause of some SUNDS victims and maybe a new susceptible gene for Brugada syndrome.

  5. Rapid Identification of Pathogenic Variants in Two Cases of Charcot-Marie-Tooth Disease by Gene-Panel Sequencing.

    PubMed

    Ho, Chi-Chun; Tai, Shuk-Mui; Lee, Edmond Chi-Nam; Mak, Timothy Shin-Heng; Liu, Timothy Kam-Tim; Tang, Victor Wai-Lun; Poon, Wing-Tat

    2017-04-05

    Charcot-Marie-Tooth disease (CMT) is a common inherited peripheral neuropathy affecting up to 1 in 1214 of the general population with more than 60 nuclear genes implicated in its pathogenesis. Traditional molecular diagnostic pathways based on relative prevalence and clinical phenotyping are limited by long turnaround time, population-specific prevalence of causative variants and inability to assess multiple co-existing variants. In this study, a CMT gene panel comprising 27 genes was used to uncover the pathogenic mutations in two index patients. The first patient is a 15-year-old boy, born of consanguineous parents, who has had frequent trips and falls since infancy, and was later found to have inverted champagne bottle appearance of bilateral legs and foot drop. His elder sister is similarly affected. The second patient is a 37-year-old woman referred for pre-pregnancy genetic diagnosis. During early adulthood, she developed progressive lower limb weakness, difficulties in tip-toe walking and thinning of calf muscles. Both patients are clinically compatible with CMT, have undergone multiple genetic testings and have not previously received a definitive genetic diagnosis. Patients 1 and 2 were found to have pathogenic homozygous HSPB1:NM_001540:c.250G>A (p.G84R) variant and heterozygous GDAP1:NM_018972:c.358C>T (p.R120W) variant, respectively. Advantages and limitations of the current approach are discussed.

  6. Satellite identification: object oriented tools for accurate maintenance of the catalog

    NASA Astrophysics Data System (ADS)

    Kamensky, S.

    2001-10-01

    Satellite identification procedures are based on joint analysis of orbital and non-coordinate data available in the Data Processing Center. The work presents the system of software tools used for this analysis. The first section of the paper presents general composition of the model of the space situation used by the Data Processing Center, indicating and describing the basic entities, objects, classes, attributes and techniques (with the required databases and archives) used for statistical and logical inference. The place of satellite identification tasks in this system is outlined. Then a set of classifiers, used for statistical inference on satellite type (spacecraft, rocket-body, fragment; specific type (series) of a spacecraft or rocket-body) is described. These classifiers use orbital and non-orbital (estimations of sizes, rotation, ballistic characteristics) data and evaluation of the evolution of these parameters if needed. However, the actual satellite identification techniques normally involve the data on all the objects of the launch (for analysis of new satellites) and consider the structure of satellite groups and constellations - regarding the place of the analyzed satellites in them and their evolution. The examples of enhanced efficiency of the system compared to the simple cluster-based analysis are presented as well as the illustrations of the structural advantages and "richer" inference capabilities.

  7. Identification of a novel variant of LMP-1 of EBV in patients with endemic Burkitt lymphoma in western Kenya.

    PubMed

    Wohlford, Eric M; Asito, Amolo S; Chelimo, Kiprotich; Sumba, Peter O; Baresel, Paul C; Oot, Rebecca A; Moormann, Ann M; Rochford, Rosemary

    2013-09-09

    Epstein Barr virus (EBV) is a gammaherpesvirus that is associated with nasopharyngeal carcinoma (NPC) and endemic Burkitt lymphoma (eBL). EBV carries several latent genes that contribute to oncogenesis including the latent membrane protein 1 (LMP-1), a known oncogene and constitutively active CD40 homolog. Variation in the C terminal region of LMP-1 has been linked to NPC pathogenesis, but little is known regarding LMP-1 variation and eBL. In the present study, peripheral blood samples were obtained from 38 eBL patients and 22 healthy controls in western Kenya, where the disease is endemic. The LMP-1 C-terminal region from these samples was sequenced and analyzed. The frequency of a 30 base pair deletion of LMP-1 previously linked to NPC was not associated with eBL compared to healthy controls. However a novel LMP-1 variant was identified, called K for Kenya and for the G318K mutation that characterizes it. The K variant LMP-1 was found in 40.5% of eBL sequences and 25.0% of healthy controls. All K variant sequences contained mutations in both of the previously described minimal T cell epitopes in the C terminal end of LMP-1. These mutations occurred in the anchor residue at the C-terminal binding groove of both epitopes, a pocket necessary for MHC loading. Overall, our results suggest that there is a novel K variant of LMP-1 in Kenya that may be associated with eBL. Further studies are necessary to determine the functional implications of the LMP-1 variant on early events in eBL genesis.

  8. Identification of a novel variant of LMP-1 of EBV in patients with endemic Burkitt lymphoma in western Kenya

    PubMed Central

    2013-01-01

    Background Epstein Barr virus (EBV) is a gammaherpesvirus that is associated with nasopharyngeal carcinoma (NPC) and endemic Burkitt lymphoma (eBL). EBV carries several latent genes that contribute to oncogenesis including the latent membrane protein 1 (LMP-1), a known oncogene and constitutively active CD40 homolog. Variation in the C terminal region of LMP-1 has been linked to NPC pathogenesis, but little is known regarding LMP-1 variation and eBL. Results In the present study, peripheral blood samples were obtained from 38 eBL patients and 22 healthy controls in western Kenya, where the disease is endemic. The LMP-1 C-terminal region from these samples was sequenced and analyzed. The frequency of a 30 base pair deletion of LMP-1 previously linked to NPC was not associated with eBL compared to healthy controls. However a novel LMP-1 variant was identified, called K for Kenya and for the G318K mutation that characterizes it. The K variant LMP-1 was found in 40.5% of eBL sequences and 25.0% of healthy controls. All K variant sequences contained mutations in both of the previously described minimal T cell epitopes in the C terminal end of LMP-1. These mutations occurred in the anchor residue at the C-terminal binding groove of both epitopes, a pocket necessary for MHC loading. Conclusions Overall, our results suggest that there is a novel K variant of LMP-1 in Kenya that may be associated with eBL. Further studies are necessary to determine the functional implications of the LMP-1 variant on early events in eBL genesis. PMID:24016332

  9. Postmortem genetic screening for the identification, verification, and reporting of genetic variants contributing to the sudden death of the young

    PubMed Central

    Methner, D. Nicole R.; Scherer, Steven E.; Welch, Katherine; Walkiewicz, Magdalena; Eng, Christine M.; Belmont, John W.; Powell, Mark C.; Korchina, Viktoriya; Doddapaneni, Harsha Vardhan; Muzny, Donna M.; Gibbs, Richard A.; Wolf, Dwayne A.; Sanchez, Luis A.; Kahn, Roger

    2016-01-01

    Each year in the United States, thousands of cases of sudden and unexpected deaths of infants, children, and young adults are assigned an undetermined cause of death after postmortem investigation and autopsy. Heritable genetic variants have been suggested as the cause of up to a third of sudden death (SD) cases. Elucidation of the genetic variants involved in SD cases is important to not only help establish cause and manner of death of these individuals, but to also aid in determining whether familial genetic testing should be considered. Previously, these types of postmortem screenings have not been a feasible option for most county medical examiners’ and coroners’ offices. We sequenced full exons of 64 genes associated with SD in the largest known cohort (351) of infant and young SD decedents using massively parallel sequencing at <$600 per sample. Genetic variants were assessed through literature review and clinical evaluation by a multidisciplinary consortium of experts. Thirteen individuals (3.7%), eight infants (2.8% of those <1 yr of age) and five children/young adults (7.0% of those >1 yr of age), were found to have a reportable genetic variant contributing to SD. These percentages represent an estimate lower than those previously reported. Overall yields and results likely vary between studies due to differences in evaluation techniques and reporting. Additionally, we recommend ongoing assessment of data, including nonreported novel variants, as technology and literature continually advance. This study demonstrates a strategy to implement molecular autopsies in medicolegal investigations of young SD decedents. PMID:27435932

  10. Identification of molecular variants of the nonrestoring restorer-of-fertility 1 allele in sugar beet (Beta vulgaris L.).

    PubMed

    Ohgami, Takashi; Uchiyama, Daisuke; Ue, Sachiyo; Yui-Kurino, Rika; Yoshida, Yu; Kamei, Yoko; Kuroda, Yosuke; Taguchi, Kazunori; Kubo, Tomohiko

    2016-04-01

    Only three variants of nonrestoring alleles for sugar beet Rf1 were found from the US maintainer lines which were the selections from a broad range of genetic resources. Cytoplasmic male sterility is widely used for hybrid breeding of sugar beets. Specific genotypes with a nonsterility-inducing cytoplasm and a nonrestoring allele of restorer-of-fertility gene (rf) are called maintainers. The infrequent occurrence of the maintainer genotype evokes the need to diagnose rf alleles. Molecular analysis of Rf1, one of the sugar beet Rfs, revealed a high level of nucleotide sequence diversity, but three variants were tightly associated with maintainer selection in Japan. The question was raised whether this small number of variants would be seen in cases where a wider range of genetic resources was used for maintainer selection. Fifty-seven accessions registered as maintainers in the USDA germplasm collection were characterized in this study. Mitochondrial DNA types (mitotypes) of 551 plants were diagnosed based on minisatellite polymorphism. A mitotype associated with sterility-inducing (S) cytoplasm was identified in 58 plants, indicating S-cytoplasm contamination. The organization of rf1 was investigated by two PCR markers and DNA gel blot analysis. Eight haplotypes were found among the US maintainers, but subsequently two haplotypes were judged as restoring alleles after a test cross and another haplotype was not inherited by the progeny. Nucleotide sequences of rf1 regions in the remaining five haplotypes were compared, and despite the sequence diversity of the gene-flanking regions, the gene-coding regions were identified to be three types. Therefore, there are three rf1 variants in US maintainers, the same number as in the Japanese sugar beet germplasm collection. The implications of having a small repertoire of rf1 variants are discussed.

  11. Identification of cultivated land using remote sensing images based on object-oriented artificial bee colony algorithm

    NASA Astrophysics Data System (ADS)

    Li, Nan; Zhu, Xiufang

    2017-04-01

    Cultivated land resources is the key to ensure food security. Timely and accurate access to cultivated land information is conducive to a scientific planning of food production and management policies. The GaoFen 1 (GF-1) images have high spatial resolution and abundant texture information and thus can be used to identify fragmentized cultivated land. In this paper, an object-oriented artificial bee colony algorithm was proposed for extracting cultivated land from GF-1 images. Firstly, the GF-1 image was segmented by eCognition software and some samples from the segments were manually identified into 2 types (cultivated land and non-cultivated land). Secondly, the artificial bee colony (ABC) algorithm was used to search for classification rules based on the spectral and texture information extracted from the image objects. Finally, the extracted classification rules were used to identify the cultivated land area on the image. The experiment was carried out in Hongze area, Jiangsu Province using wide field-of-view sensor on the GF-1 satellite image. The total precision of classification result was 94.95%, and the precision of cultivated land was 92.85%. The results show that the object-oriented ABC algorithm can overcome the defect of insufficient spectral information in GF-1 images and obtain high precision in cultivated identification.

  12. Single assay for simultaneous detection and differential identification of human and avian influenza virus types, subtypes, and emergent variants

    USDA-ARS?s Scientific Manuscript database

    Rapid and accurate detection, identification and genetic characterization are essential for effective surveillance and epidemiological tracking of influenza viruses. This report describes applications of a resequencing pathogen microarray (RPM) assay that is capable of simultaneous sequencing of su...

  13. Identification of novel OCT4 genetic variant associated with the risk of chronic hepatitis B in a Korean population.

    PubMed

    Shin, Joong-Gon; Cheong, Hyun Sub; Lee, Kwanghyun; Ju, Bong-Gun; Lee, Jeong-Hoon; Yu, Su Jong; Yoon, Jung-Hwan; Cheong, Jae Youn; Cho, Sung Won; Park, Neung Hwa; Namgoong, Suhg; Kim, Lyoung Hyo; Kim, Yoon Jun; Shin, Hyoung Doo

    2017-03-01

    Hepatitis B viral infection is a serious risk factor for chronic hepatitis B (CHB), cirrhosis and hepatocellular carcinoma. Recently, several genome-wide association studies (GWASs) have been conducted to identify important genetic variant associated with the risk of CHB. In our previous GWAS, TCF19 was identified as one of the susceptibility genes for CHB risk (P=4.2×10(-9) at rs1419881). In order to discover possible additional causal variants around TCF19, we performed an association study by genotyping single nucleotide polymorphisms (SNPs) in OCT4, a nearby gene to TCF19. Nineteen OCT4 genetic variants were selected and genotyped in 3902 subjects (1046 CHB patients and 2856 population controls). Logistic regression analysis revealed that OCT4 rs1265163 showed the most significant association signal for the risk of CHB (OR=1.46, P=4.78×10(-12) ). Linkage disequilibrium and conditional analysis confirmed rs1265163 in OCT4 as a novel genetic marker for CHB susceptibility. The genetic risk scores (GRSs) were calculated to visualize the combined genetic effects of all known CHB-associated loci, including OCT4 rs1265163, which had been identified in this study. Individuals with higher cumulative GRSs showed significantly increased ORs. The luciferase activity of rs885952, a tagging SNP of rs1265163, showed that OCT4 promoter activity was significantly different between the wild-type and SNP mutant form (P<.05). This follow-up study to our previous GWAS identified a possible causal genetic variant associated with the risk of CHB, and findings from this study may prove useful in the understanding of genetic susceptibility to CHB. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  14. Identification of the PS1 Thr147Ile Variant in a Family with Very Early Onset Dementia and Expressive Aphasia

    PubMed Central

    Denvir, James; Neitch, Shirley; Fan, Jun; Niles, Richard M.; Boskovic, Goran; Schreurs, Bernard G.; Primerano, Donald A.; Alkon, Daniel L.

    2015-01-01

    Abstract Background: Early onset dementias have variable clinical presentations and are often difficult to diagnose. We established a family pedigree that demonstrated consistent recurrence of very early onset dementia in successive generations. Objective and Method: In order to refine the diagnosis in this family, we sequenced the exomes of two affected family members and relied on discrete filtering to identify disease genes and the corresponding causal variants. Results: Among the 720 nonsynonymous single nucleotide polymorphisms (SNPs) shared by two affected members, we found a C to T transition that gives rise to a Thr147Ile missense substitution in the presenilin 1 (PS1) protein. The presence of this same mutation in a French early-onset Alzheimer’s disease family, other affected members of the family, and the predicted high pathogenicity of the substitution strongly suggest that it is the causal variant. In addition to exceptionally young age of onset, we also observed significant limb spasticity and early loss of speech, concurrent with progression of dementia in affected family members. These findings extend the clinical presentation associated with the Thr147Ile variant. Lastly, one member with the Thr147Ile variant was treated with the PKC epsilon activator, bryostatin, in a compassionate use trial after successful FDA review. Initial improvements with this treatment were unexpectedly clear, including return of some speech, increased attentional focus, ability to swallow, and some apparent decrease in limb spasticity. Conclusions: Our findings confirm the role of the PS1 Thr147Ile substitution in Alzheimer’s disease and expand the clinical phenotype to include expressive aphasia and very early onset of dementia. PMID:25812849

  15. Identifications of polyphyletic variants in acute hepatitis suggest an underdiagnosed circulation of hepatitis E virus in Argentina.

    PubMed

    Munné, María Silvina; Altabert, Nancy Rosana; Vladimirsky, Sara Noemí; Moreiro, Rita; Mares, Lucio Oscar Otegui; Soto, Sonia Soledad; Brajterman, Leonardo Sergio; Castro, Raúl Enrique; González, Jorge Enrique

    2011-10-01

    In recent years, an increasing number of infections with genotype 3 hepatitis E virus (HEV) have been reported in western countries. Data in South America, however, are still scarce. Swine and human variants previously described in Argentina are closely related to a human Austrian one. To identify whether HEV is still circulating in Argentina. Sera and stool samples from adults and children with unexplained acute liver disease referred to our center during the last six years were prospectively studied. Dual infection with hepatitis A was retrospectively studied in a group of children with fulminant hepatic failure. Fifteen new cases (13 adults and 2 children), seven of whom required hospitalization, were diagnosed. Nine had detectable HEV RNA, and one had imported genotype 1. Subgenotype 3i HEV-related variants are still circulating. Five autochthonous sequences, related to European, American and Japanese ones, grouped in subgenotype 3a. One case had a subgenotype 3b variant. The polyphyletic variants widespread in Argentina suggest multiple sources of infection. Whether or not their reservoir is swine merits further investigation. Since hepatitis E is still considered rare, differential laboratory testing in unexplained acute liver disease is not routinely performed in Argentina. Broadening awareness of this disease is important in light of the decrease in hepatitis A incidence since universal vaccination was implemented in 2005. The diagnosis of hepatitis E with a combination of serological and molecular tools is needed to better understand its epidemiology and impact on the clinical management of patients with unexplained increased transaminases. Copyright © 2011 Elsevier B.V. All rights reserved.

  16. Identification of a novel pituitary-specific chicken gonadotropin-releasing hormone receptor and its splice variants.

    PubMed

    Shimizu, Mamiko; Bédécarrats, Grégoy Y

    2006-11-01

    In all vertebrates, GnRH regulates gonadotropin secretion through binding to a specific receptor on the surface of pituitary gonadotropes. At least two forms of GnRH exist within a single species, and several corresponding GnRH receptors (GNRHRs) have been isolated with one form being pituitary specific. In chickens, only one type of widely expressed GNRHR has previously been identified. The objectives of this study were to isolate a chicken pituitary-specific GNRHR and to determine its expression pattern during a reproductive cycle. Using a combined strategy of PCR and rapid amplification of cDNA ends (RACE), a new GNRHR (chicken GNRHR2) and two splice variants were isolated in domestic fowl (Gallus gallus domesticus). Full-length GNRHR2 and one of its splice variant mRNAs were expressed exclusively in the pituitary, whereas mRNA of the other splice variant was expressed in most brain tissues examined. The deduced amino acid sequence of full-length chicken GNRHR2 reveals a seven transmembrane domain protein with 57%-65% homology to nonmammalian GNRHRs. Semiquantitative real-time PCR revealed that mRNA levels of full-length chicken GNRHR2 in the pituitary correlate with the reproductive status of birds, with maximum levels observed during the peak of lay and 4 wk postphotostimulation in females and males, respectively. Furthermore, GnRH stimulation of GH3 cells that were transiently transfected with cDNA that encodes chicken GNRHR2 resulted in a significant increase in inositol phosphate accumulation. In conclusion, we isolated a novel GNRHR and its splice variants in chickens, and spatial and temporal gene expression patterns suggest that this receptor plays an important role in the regulation of reproduction.

  17. Identification and functional characterization of four transient receptor potential ankyrin 1 variants in Apolygus lucorum (Meyer-Dür).

    PubMed

    Fu, T; Hull, J J; Yang, T; Wang, G

    2016-08-01

    As signal integrators that respond to various physical and chemical stimuli, transient receptor potential (TRP) channels fulfil critical functional roles in the sensory systems of both vertebrate and invertebrate organisms. Here, four variants of TRP ankyrin 1 (TRPA1) were identified and cloned from the green plant bug, Apolygus lucorum. Spatiotemporal expression profiling across development and in different adult tissues revealed that the highest relative-transcript levels occurred in first-instar nymphs and antennae, respectively. In Xenopus laevis-based functional assays, Apo. lucorum TRPA1-A (AlucTRPA1-A), AlucTRPA1-B and AlucTRPA1-C were activated by increasing the temperature from 20 to 40 °C with no significant desensitization observed after repeated temperature stimuli. The activation temperature of AlucTRPA1-A and AlucTRPA1-B was < 25 °C, whereas the activation temperature of AlucTRPA1-C was between 25 and 30 °C. Amongst the variants, only AlucTRPA1-A and AlucTRPA1-C were directly activated by high concentrations of allyl isothiocyanate, cinnamaldehyde and citronellal. Taken together, these results suggest that AlucTRPA1 variants may function in vivo as both thermal and chemical sensors, with the four variants potentially mediating different physiological functions. This study not only enriches our understanding of TRPA1 function in Hemiptera (Miridae), but also offers a foundation for developing new pest control strategies. © 2016 The Royal Entomological Society.

  18. Trypanosoma evansi: identification and characterization of a variant surface glycoprotein lacking cysteine residues in its C-terminal domain.

    PubMed

    Jia, Yonggen; Zhao, Xinxin; Zou, Jingru; Suo, Xun

    2011-01-01

    African trypanosomes are flagellated unicellular parasites which proliferate extracellularly in the mammalian host blood-stream and tissue spaces. They evade the hosts' antibody-mediated lyses by sequentially changing their variant surface glycoprotein (VSG). VSG tightly coats the entire parasite body, serving as a physical barrier. In Trypanosoma brucei and the closely related species Trypanosoma evansi, Trypanosoma equiperdum, each VSG polypeptide can be divided into N- and C-terminal domains, based on cysteine distribution and sequence homology. N-terminal domain, the basis of antigenic variation, is hypervariable and contains all the exposed epitopes; C-terminal domain is relatively conserved and a full set of four or eight cysteines were generally observed. We cloned two genes from two distinct variants of T. evansi, utilizing RT-PCR with VSG-specific primers. One contained a VSG type A N-terminal domain followed a C-terminal domain lacking cysteine residues. To confirm that this gene is expressed as a functional VSG, the expression and localization of the corresponding gene product were characterized using Western blotting and immunofluorescent staining of living trypanosomes. Expression analysis showed that this protein was highly expressed, variant-specific, and had a ubiquitous cellular surface localization. All these results indicated that it was expressed as a functional VSG. Our finding showed that cysteine residues in VSG C-terminal domain were not essential; the conserved C-terminal domain generally in T. brucei like VSGs would possibly evolve for regulating the VSG expression.

  19. Identification of Nitrogen Consumption Genetic Variants in Yeast Through QTL Mapping and Bulk Segregant RNA-Seq Analyses

    PubMed Central

    Cubillos, Francisco A.; Brice, Claire; Molinet, Jennifer; Tisné, Sebastién; Abarca, Valentina; Tapia, Sebastián M.; Oporto, Christian; García, Verónica; Liti, Gianni; Martínez, Claudio

    2017-01-01

    Saccharomyces cerevisiae is responsible for wine must fermentation. In this process, nitrogen represents a limiting nutrient and its scarcity results in important economic losses for the wine industry. Yeast isolates use different strategies to grow in poor nitrogen environments and their genomic plasticity enables adaptation to multiple habitats through improvements in nitrogen consumption. Here, we used a highly recombinant S. cerevisiae multi-parent population (SGRP-4X) derived from the intercross of four parental strains of different origins to identify new genetic variants responsible for nitrogen consumption differences during wine fermentation. Analysis of 165 fully sequenced F12 segregants allowed us to map 26 QTL in narrow intervals for 14 amino acid sources and ammonium, the majority of which represent genomic regions previously unmapped for these traits. To complement this strategy, we performed Bulk segregant RNA-seq (BSR-seq) analysis in segregants exhibiting extremely high and low ammonium consumption levels. This identified several QTL overlapping differentially expressed genes and refined the gene candidate search. Based on these approaches, we were able to validate ARO1, PDC1, CPS1, ASI2, LYP1, and ALP1 allelic variants underlying nitrogen consumption differences between strains, providing evidence of many genes with small phenotypic effects. Altogether, these variants significantly shape yeast nitrogen consumption with important implications for evolution, ecological, and quantitative genomics. PMID:28592651

  20. Identification and characterization of a truncated variant of the 5-hydroxytryptamine(2A) receptor produced by alternative splicing.

    PubMed

    Guest, P C; Salim, K; Skynner, H A; George, S E; Bresnick, J N; McAllister, G

    2000-09-08

    We have identified an alternatively spliced 5-hydroxytryptamine 2A receptor (5-HT(2A)-R) transcript by PCR of human brain cDNA using degenerate oligonucleotide primers to transmembrane (TM) domains 3 and 7 of the 5-HT(2)-R subfamily. The variant contains a 118-bp insertion at the exon II/III boundary of the 5-HT(2A)-R, which produces a frame shift in the coding sequence and a premature stop codon. PCR analysis showed that the truncated receptor (5-HT(2A-tr)) and native 5-HT(2A)-R were co-expressed in most brain tissues, with the highest levels being found in hippocampus, corpus collosum, amygdala and caudate nucleus. Western blot analysis of HEK-293 cells transfected transiently with a 5-HT(2A-tr) construct showed that a 30-kDa protein was expressed on cell membranes. Co-transfection studies showed no effect of the 5-HT(2A-tr) variant on 3H-ketanserin binding to the native 5-HT(2A)-R or on functional coupling of the 5-HT(2A)-R to 5-HT-stimulated Ca(2+) mobilization. The functional significance of the 5-HT(2A-tr) variant and other truncated receptors remains to be established.

  1. User oriented ERTS-1 images. [vegetation identification in Canada through image enhancement

    NASA Technical Reports Server (NTRS)

    Shlien, S.; Goodenough, D.

    1974-01-01

    Photographic reproduction of ERTS-1 images are capable of displaying only a portion of the total information available from the multispectral scanner. Methods are being developed to generate ERTS-1 images oriented towards special users such as agriculturists, foresters, and hydrologists by applying image enhancement techniques and interactive statistical classification schemes. Spatial boundaries and linear features can be emphasized and delineated using simple filters. Linear and nonlinear transformations can be applied to the spectral data to emphasize certain ground information. An automatic classification scheme was developed to identify particular ground cover classes such as fallow, grain, rape seed or various vegetation covers. The scheme applies the maximum likelihood decision rule to the spectral information and classifies the ERTS-1 image on a pixel by pixel basis. Preliminary results indicate that the classifier has limited success in distinguishing crops, but is well adapted for identifying different types of vegetation.

  2. Identification of a Pyridopyrimidinone Inhibitor of Orthopoxviruses from a Diversity-Oriented Synthesis Library

    PubMed Central

    Dower, Ken; Filone, Claire Marie; Hodges, Erin N.; Bjornson, Zach B.; Rubins, Kathleen H.; Brown, Lauren E.; Schaus, Scott; Hensley, Lisa E.

    2012-01-01

    Orthopoxviruses include the prototypical vaccinia virus, the emerging infectious agent monkeypox virus, and the potential biothreat variola virus (the causative agent of smallpox). There is currently no FDA-approved drug for humans infected with orthopoxviruses. We screened a diversity-oriented synthesis library for new scaffolds with activity against vaccinia virus. This screen identified a nonnucleoside analog that blocked postreplicative intermediate and late gene expression. Viral genome replication was unaffected, and inhibition could be elicited late in infection and persisted upon drug removal. Sequencing of drug-resistant viruses revealed mutations predicted to be on the periphery of the highly conserved viral RNA polymerase large subunit. Consistent with this, the compound had broad-spectrum activity against orthopoxviruses in vitro. These findings indicate that novel chemical synthesis approaches are a potential source for new infectious disease therapeutics and identify a potentially promising candidate for development to treat orthopoxvirus-infected individuals. PMID:22205744

  3. Identification and synthesis of female sex pheromone of Oriental beetle,Anomala orientalis (Coleoptera: Scarabaeidae).

    PubMed

    Zhang, A; Facundo, H T; Robbins, P S; Linn, C E; Hanula, J L; Villani, M G; Roelofs, W L

    1994-09-01

    Females of the Oriental beetle,Anomala orientalis (Waterhouse), release a sex pheromone composed of a 9:1 blend of (Z)- and (E)-7-tetradecen-2-one. The double-bond position of the pheromone was determined by DMDS derivatization and interpretation of the fragmentation patterns produced by monounsaturated ketones. In a sustained-flight tunnel, males responded by flying toward female beetles and attempting to copulate with them. Both effluvium and whole-body extracts of OB females were analyzed, and the activity was found only in the airborne extracts. Flight-tunnel bioassays also showed that a synthetic 90:10Z/E blend on a rubber septum was attractive and that the responses of males to this blend were equivalent toZ isomer alone, but much better than to the singleE isomer.

  4. Identification and characterization of G beta 3s2, a novel splice variant of the G-protein beta 3 subunit.

    PubMed Central

    Rosskopf, Dieter; Manthey, Iris; Habich, Christiane; Kielbik, Marzena; Eisenhardt, Andreas; Nikula, Christiane; Urban, Melanie; Kohnen, Stefanie; Graf, Eva; Ravens, Ursula; Siffert, Winfried

    2003-01-01

    The T-allele of a polymorphism (C825T) in the gene for the G-protein beta 3 subunit (GNB3) is associated with cardiovascular and metabolic disorders, distinct cellular features and altered drug responses. The molecular mechanisms that give rise to this complex phenotype have been linked to the occurrence of G beta 3s, a splice variant of GNB3. G beta 3s is predominantly expressed in cells with the 825T-allele. In the present study we describe the identification and characterization of an additional G beta 3 splice variant referred to as G beta 3s2. Its mRNA is expressed in heart, blood cells and tumour tissue, and its expression is also tightly associated with the GNB3 825T-allele. G beta 3s2 is generated by alternative splicing using non-canonical splice sites. G beta subunits belong to the family of propeller proteins and consist of seven regular propeller blades. Transcripts for G beta 3s2 are lacking 129 bp of the coding sequence of the wild-type G beta 3 protein. Thus the predicted structure consists of only six propeller blades, which resembles the structure of G beta 3s. Co-immunoprecipitation analyses indicated that G beta 3s2 dimerizes with different G gamma subunits, e.g. G gamma 5, G gamma 8(C) and G gamma 12. In Sf9 insect cells, expression of G beta 3s2 together with G gamma 12 enhances receptor-stimulated activation of G alpha(i2). Expression of G beta 3s2 in mammalian cells activated the mitogen-activated protein kinase cascade. Together, these results suggest that G beta 3s2 is a biologically active G beta variant which may play a role in the manifestation of the complex phenotype associated with the 825T-allele. PMID:12431187

  5. Emotional scenes in peripheral vision: selective orienting and gist processing, but not content identification.

    PubMed

    Calvo, Manuel G; Nummenmaa, Lauri; Hyönä, Jukka

    2008-02-01

    Emotional-neutral pairs of visual scenes were presented peripherally (with their inner edges 5.2 degrees away from fixation) as primes for 150 to 900 ms, followed by a centrally presented recognition probe scene, which was either identical in specific content to one of the primes or related in general content and affective valence. Results indicated that (a) if no foveal fixations on the primes were allowed, the false alarm rate for emotional probes was increased; (b) hit rate and sensitivity (A') were higher for emotional than for neutral probes only when a fixation was possible on only one prime; and (c) emotional scenes were more likely to attract the first fixation than neutral scenes. It is concluded that the specific content of emotional or neutral scenes is not processed in peripheral vision. Nevertheless, a coarse impression of emotional scenes may be extracted, which then leads to selective attentional orienting or--in the absence of overt attention--causes false alarms for related probes.

  6. Large-scale identification and characterization of alternative splicing variants of human gene transcripts using 56 419 completely sequenced and manually annotated full-length cDNAs

    PubMed Central

    Takeda, Jun-ichi; Suzuki, Yutaka; Nakao, Mitsuteru; Barrero, Roberto A.; Koyanagi, Kanako O.; Jin, Lihua; Motono, Chie; Hata, Hiroko; Isogai, Takao; Nagai, Keiichi; Otsuki, Tetsuji; Kuryshev, Vladimir; Shionyu, Masafumi; Yura, Kei; Go, Mitiko; Thierry-Mieg, Jean; Thierry-Mieg, Danielle; Wiemann, Stefan; Nomura, Nobuo; Sugano, Sumio; Gojobori, Takashi; Imanishi, Tadashi

    2006-01-01

    We report the first genome-wide identification and characterization of alternative splicing in human gene transcripts based on analysis of the full-length cDNAs. Applying both manual and computational analyses for 56 419 completely sequenced and precisely annotated full-length cDNAs selected for the H-Invitational human transcriptome annotation meetings, we identified 6877 alternative splicing genes with 18 297 different alternative splicing variants. A total of 37 670 exons were involved in these alternative splicing events. The encoded protein sequences were affected in 6005 of the 6877 genes. Notably, alternative splicing affected protein motifs in 3015 genes, subcellular localizations in 2982 genes and transmembrane domains in 1348 genes. We also identified interesting patterns of alternative splicing, in which two distinct genes seemed to be bridged, nested or having overlapping protein coding sequences (CDSs) of different reading frames (multiple CDS). In these cases, completely unrelated proteins are encoded by a single locus. Genome-wide annotations of alternative splicing, relying on full-length cDNAs, should lay firm groundwork for exploring in detail the diversification of protein function, which is mediated by the fast expanding universe of alternative splicing variants. PMID:16914452

  7. SVSI: fast and powerful set-valued system identification approach to identifying rare variants in sequencing studies for ordered categorical traits.

    PubMed

    Bi, Wenjian; Kang, Guolian; Zhao, Yanlong; Cui, Yuehua; Yan, Song; Li, Yun; Cheng, Cheng; Pounds, Stanley B; Borowitz, Michael J; Relling, Mary V; Yang, Jun J; Liu, Zhifa; Pui, Ching-Hon; Hunger, Stephen P; Hartford, Christine M; Leung, Wing; Zhang, Ji-Feng

    2015-07-01

    In genetic association studies of an ordered categorical phenotype, it is usual to either regroup multiple categories of the phenotype into two categories and then apply the logistic regression (LG), or apply ordered logistic (oLG), or ordered probit (oPRB) regression, which accounts for the ordinal nature of the phenotype. However, they may lose statistical power or may not control type I error due to their model assumption and/or instable parameter estimation algorithm when the genetic variant is rare or sample size is limited. To solve this problem, we propose a set-valued (SV) system model to identify genetic variants associated with an ordinal categorical phenotype. We couple this model with a SV system identification algorithm to identify all the key system parameters. Simulations and two real data analyses show that SV and LG accurately controlled the Type I error rate even at a significance level of 10(-6) but not oLG and oPRB in some cases. LG had significantly less power than the other three methods due to disregarding of the ordinal nature of the phenotype, and SV had similar or greater power than oLG and oPRB. We argue that SV should be employed in genetic association studies for ordered categorical phenotype.

  8. Systemic senile amyloidosis. Identification of a new prealbumin (transthyretin) variant in cardiac tissue: immunologic and biochemical similarity to one form of familial amyloidotic polyneuropathy.

    PubMed Central

    Gorevic, P D; Prelli, F C; Wright, J; Pras, M; Frangione, B

    1989-01-01

    Isolated amyloid fibrils from three cases of systemic senile amyloidosis (SSA) contained subunit proteins with molecular masses of 14 (10-20%), 10-12 (60-80%), and 5-6 kD (5-10%) when fractionated under reducing and dissociating conditions. This grouping was identical to that seen in SKO, a case of familial amyloidotic polyneuropathy (FAP) studied earlier. Amino acid sequencing confirmed that SSA subunit proteins were in fact prealbumin (transthyretin). Complete sequence analysis of one SSA preparation revealed the presence of a new variant Pa (TTr) molecule with a single amino acid substitution of isoleucine for valine at position 122. Further studies used an antiserum specific for SKO IV, a subunit protein of SKO previously shown to correspond to carboxy-terminal 78 residues (positions 49-127) of (TTr). Anti-SKO IV reacted with SSA in tissue at equivalent dilutions to anti-Pa (TTr) and with the 10-12-kD fraction of SSA on Western blots; reactivity was blocked by SKO IV, but not by Pa (TTr). SSA is a form of systemic amyloidosis caused by tissue deposition of Pa (TTr) and its fragments, with shared conformational or subunit antigenicity to at least one form of FAP. Identification of a new variant Pa (TTr) molecule in one case suggests further that SSA may be a genetically determined disease expressed late in life. Images PMID:2646319

  9. Multiplex PCR strategy for rapid identification of structural types and variants of the mec element in methicillin-resistant Staphylococcus aureus.

    PubMed

    Oliveira, Duarte C; de Lencastre, Hermínia

    2002-07-01

    Full characterization of methicillin-resistant Staphylococcus aureus (MRSA) requires definition of not only the bacterial genetic background but also the structure of the complex and heterologous mec element these bacteria carry, which is associated with drug resistance determinant mecA. We report the development, validation, and application of a multiplex PCR strategy that allows quick presumptive characterization of the mec element types based on the structural features that were shown to be typical of mec elements carried by several MRSA clones. The strategy was validated by using a representative collection of pandemic MRSA clones in which the full structure of the associated mec elements was previously determined by hybridization and PCR screenings and also by DNA sequencing. The method was tested together with multilocus sequence typing and other typing methods for the characterization of 18 isolates representative of the MRSA clones recovered during a hospital outbreak in Barcelona, Spain. The multiplex PCR was shown to be rapid, robust, and capable in a single assay of identifying five structural types of the mec element among these strains, three major and two minor variants, each one of which has been already been seen among MRSA characterized earlier. This technique should be a useful addition to the armamentarium of molecular typing tools for the characterization of MRSA clonal types and for the rapid tentative identification of structural variants of the mec element.

  10. Identification and expression pattern of two novel alternative splicing variants of EEF1D gene of dairy cattle.

    PubMed

    Xie, Yan; Yang, Shaohua; Cui, Xiaogang; Jiang, Li; Zhang, Shengli; Zhang, Qin; Zhang, Yuan; Sun, Dongxiao

    2014-01-25

    Our recent genome-wide association study (GWAS) has identified 105 genome-wide significant SNPs for milk production traits. Of these, one SNP (rs109661298) for milk fat percentage is located within the first intron of the bovine EEF1D gene on BTA14, thus that the EEF1D gene was considered as a novel candidate in dairy cattle. Until now, however, its genomic organization remains undetermined yet and no studies of EEF1D in relation to milk production traits have been reported. To layout the groundwork for the validation of gene function in dairy cattle, we herein investigated its expression pattern in lactating dairy cows. With rapid amplification of 5' cDNA end (5' RACE), two novel alternatively spliced transcript variants of 1202bp and 2195bp were isolated in bovine mammary in lactation, named EEF1Da and EEF1Db (GenBank: KC190039 and KC190038KC190039KC190038). Such two variants contain the different first exon from each other (exon1a vs exon1b: 294bp vs 1287bp) with no overlap and the same remaining 7 exons. Coding sequence similarity between EEF1Da and EEF1Db and three of human EEF1D transcript variants were 85-88%. With semi-quantitative and quantitative real-time RT-PCR, we found that the mRNA level of EEF1Da was similar to the overall EEF1D mRNA and much higher than EEF1Db in the mammary of lactating cows, indicating EEF1Da functions as the dominant transcript variant to encode the EEF1D protein. Tissue expression pattern showed that the mRNA expression of EEF1D and EEF1Da in mammary gland was significantly higher compared with other 7 tissues (P<0.05, P<0.01) with the exception of EEF1D between mammary and lung. Together, our findings present the first report on the alternative splicing of the bovine EEF1D gene and provided basis for further investigation on function validation of EEF1D in dairy cattle. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. Identification and expression profile analysis of odorant binding proteins in the oriental fruit fly Bactrocera dorsalis.

    PubMed

    Zheng, Weiwei; Peng, Wei; Zhu, Chipan; Zhang, Qun; Saccone, Giuseppe; Zhang, Hongyu

    2013-07-17

    Olfaction is crucial in many insects for critical behaviors, including those regulating survival and reproduction. Insect odorant-binding proteins (OBPs) function in the first step of the olfactory system and play an essential role in the perception of odorants, such as pheromones and host chemicals. The oriental fruit fly, Bactrocera dorsalis, is a destructive fruit-eating pest, due to its wide host range of up to 250 different types of fruits and vegetables, and this fly causes severe economic damage to the fruit and vegetable industry. However, OBP genes have not been largely identified in B. dorsalis. Based on our previously constructed B. dorsalis cDNA library, ten OBP genes were identified in B. dorsalis for the first time. A phylogenetic tree was generated to show the relationships among the 10 OBPs of B. dorsalis to OBP sequences of two other Dipteran species, including Drosophila melanogaster and the mosquito Anopheles gambiae. The expression profiles of the ten OBPs in different tissues (heads, thoraxes, abdomens, legs, wings, male antennae and female antenna) of the mated adults were analyzed by real-time PCR. The results showed that nine of them are highly expressed in the antenna of both sexes, except BdorOBP7. Four OBPs (BdorOBP1, BdorOBP4, BdorOBP8, and BdorOBP10) are also enriched in the abdomen, and BdorOBP7 is specifically expressed in leg, indicating that it may function in other biological processes. This work will provide insight into the roles of OBPs in chemoreception and help develop new pest-control strategies.

  12. Observation of 'hidden' planar defects in boron carbide nanowires and identification of their orientations.

    PubMed

    Guan, Zhe; Cao, Baobao; Yang, Yang; Jiang, Youfei; Li, Deyu; Xu, Terry T

    2014-01-15

    The physical properties of nanostructures strongly depend on their structures, and planar defects in particular could significantly affect the behavior of the nanowires. In this work, planar defects (twins or stacking faults) in boron carbide nanowires are extensively studied by transmission electron microscopy (TEM). Results show that these defects can easily be invisible, i.e., no presence of characteristic defect features like modulated contrast in high-resolution TEM images and streaks in diffraction patterns. The simplified reason of this invisibility is that the viewing direction during TEM examination is not parallel to the (001)-type planar defects. Due to the unique rhombohedral structure of boron carbide, planar defects are only distinctive when the viewing direction is along the axial or short diagonal directions ([100], [010], or 1¯10) within the (001) plane (in-zone condition). However, in most cases, these three characteristic directions are not parallel to the viewing direction when boron carbide nanowires are randomly dispersed on TEM grids. To identify fault orientations (transverse faults or axial faults) of those nanowires whose planar defects are not revealed by TEM, a new approach is developed based on the geometrical analysis between the projected preferred growth direction of a nanowire and specific diffraction spots from diffraction patterns recorded along the axial or short diagonal directions out of the (001) plane (off-zone condition). The approach greatly alleviates tedious TEM examination of the nanowire and helps to establish the reliable structure-property relations. Our study calls attention to researchers to be extremely careful when studying nanowires with potential planar defects by TEM. Understanding the true nature of planar defects is essential in tuning the properties of these nanostructures through manipulating their structures.

  13. Identification and Expression Profile Analysis of Odorant Binding Proteins in the Oriental Fruit Fly Bactrocera dorsalis

    PubMed Central

    Zheng, Weiwei; Peng, Wei; Zhu, Chipan; Zhang, Qun; Saccone, Giuseppe; Zhang, Hongyu

    2013-01-01

    Olfaction is crucial in many insects for critical behaviors, including those regulating survival and reproduction. Insect odorant-binding proteins (OBPs) function in the first step of the olfactory system and play an essential role in the perception of odorants, such as pheromones and host chemicals. The oriental fruit fly, Bactrocera dorsalis, is a destructive fruit-eating pest, due to its wide host range of up to 250 different types of fruits and vegetables, and this fly causes severe economic damage to the fruit and vegetable industry. However, OBP genes have not been largely identified in B. dorsalis. Based on our previously constructed B. dorsalis cDNA library, ten OBP genes were identified in B. dorsalis for the first time. A phylogenetic tree was generated to show the relationships among the 10 OBPs of B. dorsalis to OBP sequences of two other Dipteran species, including Drosophila melanogaster and the mosquito Anopheles gambiae. The expression profiles of the ten OBPs in different tissues (heads, thoraxes, abdomens, legs, wings, male antennae and female antenna) of the mated adults were analyzed by real-time PCR. The results showed that nine of them are highly expressed in the antenna of both sexes, except BdorOBP7. Four OBPs (BdorOBP1, BdorOBP4, BdorOBP8, and BdorOBP10) are also enriched in the abdomen, and BdorOBP7 is specifically expressed in leg, indicating that it may function in other biological processes. This work will provide insight into the roles of OBPs in chemoreception and help develop new pest-control strategies. PMID:23867609

  14. Identification in Saccharomyces cerevisiae of a new stable variant of alkyl hydroperoxide reductase 1 (Ahp1) induced by oxidative stress.

    PubMed

    Prouzet-Mauléon, Valérie; Monribot-Espagne, Christelle; Boucherie, Hélian; Lagniel, Gilles; Lopez, Sébastien; Labarre, Jean; Garin, Jérome; Lauquin, Guy J-M

    2002-02-15

    Yeasts lacking cytoplasmic superoxide dismutase (Cu,Zn-SOD) activity are permanently subjected to oxidative stress. We used two-dimensional PAGE to examine the proteome pattern of Saccharomyces cerevisiae strains lacking Cu,Zn-SOD. We found a new stable form of alkyl hydroperoxide reductase 1 (Ahp1) with a lower isoelectric point. This form was also present in wild type strains after treatment with tert-butyl hydroperoxide. In vitro enzyme assays showed that Ahp1p had lower specific activity in strains lacking Cu,Zn-SOD. We studied three mutants presenting a reduced production of the low pI variant under oxidative stress conditions. Two of the mutants (C62S and S59D) were totally inactive, thus suggesting that the acidic form of Ahp1p may only appear when the enzyme is functional. The other mutant (S59A) was active in vitro and was more resistant to inactivation by tert-butyl hydroperoxide than the wild type enzyme. Furthermore, the inactivation of Ahp1p in vitro is correlated with its conversion to the low pI form. These results suggest that in vivo during some particular oxidative stress (alkyl hydroperoxide treatment or lack of Cu,Zn-SOD activity but not hydrogen peroxide treatment), the catalytic cysteine of Ahp1p is more oxidized than cysteine-sulfenic acid (a natural occurring intermediate of the enzymatic reaction) and that cysteine-sulfinic acid or cysteine-sulfonic acid variant may be inactive.

  15. Identification and characterization of the two isoforms of the vertebrate H2A.Z histone variant

    PubMed Central

    Matsuda, Ryo; Hori, Tetsuya; Kitamura, Hiroshi; Takeuchi, Kozo; Fukagawa, Tatsuo; Harata, Masahiko

    2010-01-01

    Histone variants play important roles in the epigenetic regulation of genome function. The histone variant H2A.Z is evolutionarily conserved from yeast to vertebrates, and it has been reported to have multiple effects upon gene expression and insulation, and chromosome segregation. Recently two genes encoding H2A.Z were identified in the vertebrate genome. However, it is not yet clear whether the proteins transcribed from these genes are functionally distinct. To address this issue, we knocked out each gene individually in chicken DT40 cells. We found that two distinct proteins, H2A.Z-1 and H2A.Z-2, were produced from these genes, and that these proteins could be separated on a long SDS–PAGE gel. The two isoforms were deposited to a similar extent by the SRCAP chromatin-remodeling complex, suggesting redundancy to their function. However, cells lacking either one of the two isoforms exhibited distinct alterations in cell growth and gene expression, suggesting that the two isoforms have differential effects upon nucleosome stability and chromatin structure. These findings provide insight into the molecular basis of the multiple functions of the H2A.Z gene products. PMID:20299344

  16. Mass Spectrometry-Based Methodology for Identification of Native Histone Variant Modifications From Mammalian Tissues and Solid Tumors.

    PubMed

    Nuccio, A G; Bui, M; Dalal, Y; Nita-Lazar, A

    2017-01-01

    Histone posttranslational modifications (PTMs) are key epigenetic marks involved in gene silencing or activation. Histone modifications impact chromatin organization and transcriptional processes through the changes in charge density between histones and DNA. They also serve as recognition and binding sites for specific binding proteins. Histone tails and globular cores contain many basic amino acid residues, which are subject to various dynamic modifications, making the modification repertoire extremely diverse. Consequently, determination of histone PTM identity and quantity has been a challenging task. In recent years, mass spectrometry-based methods have proven useful in histone PTM characterization. This chapter provides a brief overview of these methods and describes the approach to analyze the PTMs of the histone variant CENP-A, essential for the cell cycle progression, when present in minute amounts from tumor and mammalian tissues. Because this method does not rely on antibody-based immunopurification, we anticipate that these tools could be readily adaptable to the investigation to other histone variants in a range of mammalian tissues and solid tumors. Published by Elsevier Inc.

  17. Identification of a truncated alternative splicing variant of human PPAR{gamma}1 that exhibits dominant negative activity

    SciTech Connect

    Kim, Hyo Jung; Woo, Im Sun; Kang, Eun Sil; Eun, So Young; Kim, Hye Jung; Lee, Jae Heun; Chang, Ki Churl; Kim, Jin-Hoi; Seo, Han Geuk . E-mail: hgseo@gnu.ac.kr

    2006-09-01

    We have identified a novel variant of human peroxisome proliferator-activated receptor gamma (hPPAR{gamma}), derived from insertion of a novel exon 3'. Insertion leads to the introduction of a premature stop codon, resulting in the formation of a truncated splice variant of PPAR{gamma}1 (PPAR{gamma}1{sub tr}). Western blot analysis confirmed the presence of PPAR{gamma}1{sub tr} in tumor-derived cell lines. Although PPAR{gamma}1{sub tr} interfered with transcriptional activity of wild-type PPAR{gamma}1 (PPAR{gamma}1{sub wt}), activity could be rescued by cotransfection with a vector expressing p300. Overexpression of PPAR{gamma}1{sub tr} protein in CHO cells greatly enhanced their proliferation and anchorage-independent colony growth on soft agar. These data demonstrate that PPAR{gamma}1{sub tr} is an important physiologic isoform of PPAR{gamma} that modulates cellular functions of PPAR{gamma}1{sub wt}.

  18. Identification of a Functionally Distinct Truncated BDNF mRNA Splice Variant and Protein in Trachemys scripta elegans

    PubMed Central

    Ambigapathy, Ganesh; Zheng, Zhaoqing; Li, Wei; Keifer, Joyce

    2013-01-01

    Brain-derived neurotrophic factor (BDNF) has a diverse functional role and complex pattern of gene expression. Alternative splicing of mRNA transcripts leads to further diversity of mRNAs and protein isoforms. Here, we describe the regulation of BDNF mRNA transcripts in an in vitro model of eyeblink classical conditioning and a unique transcript that forms a functionally distinct truncated BDNF protein isoform. Nine different mRNA transcripts from the BDNF gene of the pond turtle Trachemys scripta elegans (tBDNF) are selectively regulated during classical conditioning: exon I mRNA transcripts show no change, exon II transcripts are downregulated, while exon III transcripts are upregulated. One unique transcript that codes from exon II, tBDNF2a, contains a 40 base pair deletion in the protein coding exon that generates a truncated tBDNF protein. The truncated transcript and protein are expressed in the naïve untrained state and are fully repressed during conditioning when full-length mature tBDNF is expressed, thereby having an alternate pattern of expression in conditioning. Truncated BDNF is not restricted to turtles as a truncated mRNA splice variant has been described for the human BDNF gene. Further studies are required to determine the ubiquity of truncated BDNF alternative splice variants across species and the mechanisms of regulation and function of this newly recognized BDNF protein. PMID:23825634

  19. Re-fraction: a machine learning approach for deterministic identification of protein homologues and splice variants in large-scale MS-based proteomics.

    PubMed

    Yang, Pengyi; Humphrey, Sean J; Fazakerley, Daniel J; Prior, Matthew J; Yang, Guang; James, David E; Yang, Jean Yee-Hwa

    2012-05-04

    A key step in the analysis of mass spectrometry (MS)-based proteomics data is the inference of proteins from identified peptide sequences. Here we describe Re-Fraction, a novel machine learning algorithm that enhances deterministic protein identification. Re-Fraction utilizes several protein physical properties to assign proteins to expected protein fractions that comprise large-scale MS-based proteomics data. This information is then used to appropriately assign peptides to specific proteins. This approach is sensitive, highly specific, and computationally efficient. We provide algorithms and source code for the current version of Re-Fraction, which accepts output tables from the MaxQuant environment. Nevertheless, the principles behind Re-Fraction can be applied to other protein identification pipelines where data are generated from samples fractionated at the protein level. We demonstrate the utility of this approach through reanalysis of data from a previously published study and generate lists of proteins deterministically identified by Re-Fraction that were previously only identified as members of a protein group. We find that this approach is particularly useful in resolving protein groups composed of splice variants and homologues, which are frequently expressed in a cell- or tissue-specific manner and may have important biological consequences.

  20. Identification of an erythrocyte pyruvate kinase variant in a family from Latium with non-spherocytic congenital haemolytic anaemia.

    PubMed

    Papa, G; De Laurenzi, A; Isacchi, G C; Bonifazi, G; Parziale, L; Salvati, A M

    1979-01-01

    Erythrocyte PK deficiency was detected in a family from Latium in Italy. This PK variant is characterized by normal or increased activity immediately after blood collection, instability to storage, to heat and to urea. Only in the propositus the mutant enzyme exhibited an increased Michaelis constant for PEP, slightly increased inhibition by ATP and an altered optimum pH value. The kinetic anomaly was only partially corrected by activation with F-1, 6-DP and by addition of 2-ME. From these results it can be concluded that in the family observed two distinct erythrocyte PK alterations were demonstrable: instability in the propositus and his father; low affinity for PEP and altered optimum pH value only in the propositus.

  1. Versatile O-GlcNAc transferase assay for high-throughput identification of enzyme variants, substrates, and inhibitors.

    PubMed

    Kim, Eun J; Abramowitz, Lara K; Bond, Michelle R; Love, Dona C; Kang, Dong W; Leucke, Hans F; Kang, Dae W; Ahn, Jong-Seog; Hanover, John A

    2014-06-18

    The dynamic glycosylation of serine/threonine residues on nucleocytoplasmic proteins with a single N-acetylglucosamine (O-GlcNAcylation) is critical for many important cellular processes. Cellular O-GlcNAc levels are highly regulated by two enzymes: O-GlcNAc transferase (OGT) is responsible for GlcNAc addition and O-GlcNAcase (OGA) is responsible for removal of the sugar. The lack of a rapid and simple method for monitoring OGT activity has impeded the efficient discovery of potent OGT inhibitors. In this study we describe a novel, single-well OGT enzyme assay that utilizes 6 × His-tagged substrates, a chemoselective chemical reaction, and unpurified OGT. The high-throughput Ni-NTA Plate OGT Assay will facilitate discovery of potent OGT-specific inhibitors on versatile substrates and the characterization of new enzyme variants.

  2. Identification of multiple risk variants for ankylosing spondylitis through high-density genotyping of immune-related loci.

    PubMed

    Cortes, Adrian; Hadler, Johanna; Pointon, Jenny P; Robinson, Philip C; Karaderi, Tugce; Leo, Paul; Cremin, Katie; Pryce, Karena; Harris, Jessica; Lee, Seunghun; Joo, Kyung Bin; Shim, Seung-Cheol; Weisman, Michael; Ward, Michael; Zhou, Xiaodong; Garchon, Henri-Jean; Chiocchia, Gilles; Nossent, Johannes; Lie, Benedicte A; Førre, Øystein; Tuomilehto, Jaakko; Laiho, Kari; Jiang, Lei; Liu, Yu; Wu, Xin; Bradbury, Linda A; Elewaut, Dirk; Burgos-Vargas, Ruben; Stebbings, Simon; Appleton, Louise; Farrah, Claire; Lau, Jonathan; Kenna, Tony J; Haroon, Nigil; Ferreira, Manuel A; Yang, Jian; Mulero, Juan; Fernandez-Sueiro, Jose Luis; Gonzalez-Gay, Miguel A; Lopez-Larrea, Carlos; Deloukas, Panos; Donnelly, Peter; Bowness, Paul; Gafney, Karl; Gaston, Hill; Gladman, Dafna D; Rahman, Proton; Maksymowych, Walter P; Xu, Huji; Crusius, J Bart A; van der Horst-Bruinsma, Irene E; Chou, Chung-Tei; Valle-Oñate, Raphael; Romero-Sánchez, Consuelo; Hansen, Inger Myrnes; Pimentel-Santos, Fernando M; Inman, Robert D; Videm, Vibeke; Martin, Javier; Breban, Maxime; Reveille, John D; Evans, David M; Kim, Tae-Hwan; Wordsworth, Bryan Paul; Brown, Matthew A

    2013-07-01

    Ankylosing spondylitis is a common, highly heritable inflammatory arthritis affecting primarily the spine and pelvis. In addition to HLA-B*27 alleles, 12 loci have previously been identified that are associated with ankylosing spondylitis in populations of European ancestry, and 2 associated loci have been identified in Asians. In this study, we used the Illumina Immunochip microarray to perform a case-control association study involving 10,619 individuals with ankylosing spondylitis (cases) and 15,145 controls. We identified 13 new risk loci and 12 additional ankylosing spondylitis-associated haplotypes at 11 loci. Two ankylosing spondylitis-associated regions have now been identified encoding four aminopeptidases that are involved in peptide processing before major histocompatibility complex (MHC) class I presentation. Protective variants at two of these loci are associated both with reduced aminopeptidase function and with MHC class I cell surface expression.

  3. Identification of multiple risk variants for ankylosing spondylitis through high-density genotyping of immune-related loci

    PubMed Central

    Cortes, Adrian; Hadler, Johanna; Pointon, Jenny P; Robinson, Philip C; Karaderi, Tugce; Leo, Paul; Cremin, Katie; Pryce, Karena; Harris, Jessica; lee, Seunghun; Joo, Kyung Bin; Shim, Seung-Cheol; Weisman, Michael; Ward, Michael; Zhou, Xiaodong; Garchon, Henri-Jean; Chiocchia, Gilles; Nossent, Johannes; Lie, Benedicte A; Førre, Øystein; Tuomilehto, Jaakko; Laiho, Kari; Jiang, Lei; Liu, Yu; Wu, Xin; Bradbury, Linda A; Elewaut, Dirk; Burgos-Vargas, Ruben; Stebbings, Simon; Appleton, Louise; Farrah, Claire; Lau, Jonathan; Kenna, Tony J; Haroon, Nigil; Ferreira, Manuel A; Yang, Jian; Mulero, Juan; Fernandez-Sueiro, Jose Luis; Gonzalez-Gay, Miguel A; lopez-Larrea, Carlos; Deloukas, Panos; Donnelly, Peter; Bowness, Paul; Gafney, Karl; Gaston, Hill; Gladman, Dafna D; Rahman, Proton; Maksymowych, Walter P; Xu, Huji; Crusius, J Bart A; van der Horst-Bruinsma, Irene E; Chou, Chung-Tei; Valle-Oñate, Raphael; Romero-Sánchez, Consuelo; Hansen, Inger Myrnes; Pimentel-Santos, Fernando M; Inman, Robert D; Videm, Vibeke; Martin, Javier; Breban, Maxime; Reveille, John D; Evans, David M; Kim, Tae-Hwan; Wordsworth, Bryan Paul; Brown, Matthew A

    2013-01-01

    Ankylosing spondylitis is a common, highly heritable inflammatory arthritis affecting primarily the spine and pelvis. In addition to HLA-B*27 alleles, 12 loci have previously been identified that are associated with ankylosing spondylitis in populations of European ancestry, and 2 associated loci have been identified in Asians. In this study, we used the Illumina Immunochip microarray to perform a case-control association study involving 10,619 individuals with ankylosing spondylitis (cases) and 15,145 controls. We identified 13 new risk loci and 12 additional ankylosing spondylitis–associated haplotypes at 11 loci. Two ankylosing spondylitis–associated regions have now been identified encoding four aminopeptidases that are involved in peptide processing before major histocompatibility complex (MHC) class I presentation. Protective variants at two of these loci are associated both with reduced aminopeptidase function and with MHC class I cell surface expression. PMID:23749187

  4. A Markov chain Monte Carlo technique for identification of combinations of allelic variants underlying complex diseases in humans.

    PubMed

    Favorov, Alexander V; Andreewski, Timophey V; Sudomoina, Marina A; Favorova, Olga O; Parmigiani, Giovanni; Ochs, Michael F

    2005-12-01

    In recent years, the number of studies focusing on the genetic basis of common disorders with a complex mode of inheritance, in which multiple genes of small effect are involved, has been steadily increasing. An improved methodology to identify the cumulative contribution of several polymorphous genes would accelerate our understanding of their importance in disease susceptibility and our ability to develop new treatments. A critical bottleneck is the inability of standard statistical approaches, developed for relatively modest predictor sets, to achieve power in the face of the enormous growth in our knowledge of genomics. The inability is due to the combinatorial complexity arising in searches for multiple interacting genes. Similar "curse of dimensionality" problems have arisen in other fields, and Bayesian statistical approaches coupled to Markov chain Monte Carlo (MCMC) techniques have led to significant improvements in understanding. We present here an algorithm, APSampler, for the exploration of potential combinations of allelic variations positively or negatively associated with a disease or with a phenotype. The algorithm relies on the rank comparison of phenotype for individuals with and without specific patterns (i.e., combinations of allelic variants) isolated in genetic backgrounds matched for the remaining significant patterns. It constructs a Markov chain to sample only potentially significant variants, minimizing the potential of large data sets to overwhelm the search. We tested APSampler on a simulated data set and on a case-control MS (multiple sclerosis) study for ethnic Russians. For the simulated data, the algorithm identified all the phenotype-associated allele combinations coded into the data and, for the MS data, it replicated the previously known findings.

  5. Single Assay for Simultaneous Detection and Differential Identification of Human and Avian Influenza Virus Types, Subtypes, and Emergent Variants

    PubMed Central

    Metzgar, David; Myers, Christopher A.; Russell, Kevin L.; Faix, Dennis; Blair, Patrick J.; Brown, Jason; Vo, Scott; Swayne, David E.; Thomas, Colleen; Stenger, David A.; Lin, Baochuan; Malanoski, Anthony P.; Wang, Zheng; Blaney, Kate M.; Long, Nina C.; Schnur, Joel M.; Saad, Magdi D.; Borsuk, Lisa A.; Lichanska, Agnieszka M.; Lorence, Matthew C.; Weslowski, Brian; Schafer, Klaus O.; Tibbetts, Clark

    2010-01-01

    For more than four decades the cause of most type A influenza virus infections of humans has been attributed to only two viral subtypes, A/H1N1 or A/H3N2. In contrast, avian and other vertebrate species are a reservoir of type A influenza virus genome diversity, hosting strains representing at least 120 of 144 combinations of 16 viral hemagglutinin and 9 viral neuraminidase subtypes. Viral genome segment reassortments and mutations emerging within this reservoir may spawn new influenza virus strains as imminent epidemic or pandemic threats to human health and poultry production. Traditional methods to detect and differentiate influenza virus subtypes are either time-consuming and labor-intensive (culture-based) or remarkably insensitive (antibody-based). Molecular diagnostic assays based upon reverse transcriptase-polymerase chain reaction (RT-PCR) have short assay cycle time, and high analytical sensitivity and specificity. However, none of these diagnostic tests determine viral gene nucleotide sequences to distinguish strains and variants of a detected pathogen from one specimen to the next. Decision-quality, strain- and variant-specific pathogen gene sequence information may be critical for public health, infection control, surveillance, epidemiology, or medical/veterinary treatment planning. The Resequencing Pathogen Microarray (RPM-Flu) is a robust, highly multiplexed and target gene sequencing-based alternative to both traditional culture- or biomarker-based diagnostic tests. RPM-Flu is a single, simultaneous differential diagnostic assay for all subtype combinations of type A influenza viruses and for 30 other viral and bacterial pathogens that may cause influenza-like illness. These other pathogen targets of RPM-Flu may co-infect and compound the morbidity and/or mortality of patients with influenza. The informative specificity of a single RPM-Flu test represents specimen-specific viral gene sequences as determinants of virus type, A/HN subtype, virulence

  6. Clinical features and MUT gene mutation spectrum in Chinese patients with isolated methylmalonic acidemia: identification of ten novel allelic variants.

    PubMed

    Han, Lian-Shu; Huang, Zhuo; Han, Feng; Ye, Jun; Qiu, Wen-Juan; Zhang, Hui-Wen; Wang, Yu; Gong, Zhu-Wen; Gu, Xue-Fan

    2015-11-01

    This study aims to study MUT gene mutation spectrum in Chinese patients with isolated methylmalonic academia (MMA) and their clinical features for the potential genotype-phenotype correlation. Forty-three patients were diagnosed with isolated MMA by elevated blood propionylcarnitine, propionylcarnitine to acetylcarnitine ratio, and urine methylmalonate without hyperhomocysteinemia. The MUT gene was amplified by polymerase chain reaction and directly sequenced. Those patients with at least one variant allele were included. The novel missense mutations were assessed by bioinformatic analysis and screened against alleles sequenced from 50 control participants. Among the 43 patients, 38 had typical clinical presentations, and the majority (30/38) experienced earlyonset MMA. Eight patients died and seven were lost to follow-up. Twenty patients had poor outcomes and eight showed normal development. The 43 identified MUT gene mutations had at least one variant allele, whereas 35 had two mutant alleles. Of the 33 mutations reported before, eight recurrent mutations were identified in 32 patients, and c.729_730insTT (p.D244Lfs*39) was the most common (12/78) in the mutant alleles. Of the 10 novel mutations, six were missense mutations and four were premature termination codon mutations. The six novel missense mutations seemed to be pathogenic. A total of 10 novel MUT mutations were detected in the Chinese population. c.729_730insTT (p.D244Lfs*39) was the most frequent mutation. A genotype-phenotype correlation could not be found, but the genotypic characterization indicated the need of genetic counseling for MMA patients and early prenatal diagnoses for high-risk families.

  7. Identification of Novel Variants in LTBP2 and PXDN Using Whole-Exome Sequencing in Developmental and Congenital Glaucoma

    PubMed Central

    Micheal, Shazia; Siddiqui, Sorath Noorani; Zafar, Saemah Nuzhat; Iqbal, Aftab; Khan, Muhammad Imran; den Hollander, Anneke I.

    2016-01-01

    Background Primary congenital glaucoma (PCG) is the most common form of glaucoma in children. PCG occurs due to the developmental defects in the trabecular meshwork and anterior chamber of the eye. The purpose of this study is to identify the causative genetic variants in three families with developmental and primary congenital glaucoma (PCG) with a recessive inheritance pattern. Methods DNA samples were obtained from consanguineous families of Pakistani ancestry. The CYP1B1 gene was sequenced in the affected probands by conventional Sanger DNA sequencing. Whole exome sequencing (WES) was performed in DNA samples of four individuals belonging to three different CYP1B1-negative families. Variants identified by WES were validated by Sanger sequencing. Results WES identified potentially causative novel mutations in the latent transforming growth factor beta binding protein 2 (LTBP2) gene in two PCG families. In the first family a novel missense mutation (c.4934G>A; p.Arg1645Glu) co-segregates with the disease phenotype, and in the second family a novel frameshift mutation (c.4031_4032insA; p.Asp1345Glyfs*6) was identified. In a third family with developmental glaucoma a novel mutation (c.3496G>A; p.Gly1166Arg) was identified in the PXDN gene, which segregates with the disease. Conclusions We identified three novel mutations in glaucoma families using WES; two in the LTBP2 gene and one in the PXDN gene. The results will not only enhance our current understanding of the genetic basis of glaucoma, but may also contribute to a better understanding of the diverse phenotypic consequences caused by mutations in these genes. PMID:27409795

  8. Identification of a common variant with potential pleiotropic effect on risk of inflammatory bowel disease and colorectal cancer

    PubMed Central

    Khalili, Hamed; Gong, Jian; Brenner, Hermann; Austin, Thomas R.; Hutter, Carolyn M.; Baba, Yoshifumi; Baron, John A.; Berndt, Sonja I.; Bézieau, Stéphane; Caan, Bette; Campbell, Peter T.; Chang-Claude, Jenny; Chanock, Stephen J.; Chen, Constance; Hsu, Li; Jiao, Shuo; Conti, David V.; Duggan, David; Fuchs, Charles S.; Gala, Manish; Gallinger, Steven; Haile, Robert W.; Harrison, Tabitha A.; Hayes, Richard; Hazra, Aditi; Henderson, Brian; Haiman, Chris; Hoffmeister, Michael; Hopper, John L.; Jenkins, Mark A.; Kolonel, Laurence N.; Küry, Sébastien; LaCroix, Andrea; Marchand, Loic Le; Lemire, Mathieu; Lindor, Noralane M.; Ma, Jing; Manson, JoAnn E.; Morikawa, Teppei; Nan, Hongmei; Ng, Kimmie; Newcomb, Polly A.; Nishihara, Reiko; Potter, John D.; Qu, Conghui; Schoen, Robert E.; Schumacher, Fredrick R.; Seminara, Daniela; Taverna, Darin; Thibodeau, Stephen; Wactawski-Wende, Jean; White, Emily; Wu, Kana; Zanke, Brent W.; Casey, Graham; Hudson, Thomas J.; Kraft, Peter; Peters, Ulrike; Slattery, Martha L.; Ogino, Shuji; Chan, Andrew T.

    2015-01-01

    Although genome-wide association studies (GWAS) have separately identified many genetic susceptibility loci for ulcerative colitis (UC), Crohn’s disease (CD) and colorectal cancer (CRC), there has been no large-scale examination for pleiotropy, or shared genetic susceptibility, for these conditions. We used logistic regression modeling to examine the associations of 181 UC and CD susceptibility variants previously identified by GWAS with risk of CRC using data from the Genetics and Epidemiology of Colorectal Cancer Consortium and the Colon Cancer Family Registry. We also examined associations of significant variants with clinical and molecular characteristics in a subset of the studies. Among 11794 CRC cases and 14190 controls, rs11676348, the susceptibility single nucleotide polymorphism (SNP) for UC, was significantly associated with reduced risk of CRC (P = 7E−05). The multivariate-adjusted odds ratio of CRC with each copy of the T allele was 0.93 (95% CI 0.89–0.96). The association of the SNP with risk of CRC differed according to mucinous histological features (P heterogeneity = 0.008). In addition, the (T) allele was associated with lower risk of tumors with Crohn’s-like reaction but not tumors without such immune infiltrate (P heterogeneity = 0.02) and microsatellite instability-high (MSI-high) but not microsatellite stable or MSI-low tumors (P heterogeneity = 0.03). The minor allele (T) in SNP rs11676348, located downstream from CXCR2 that has been implicated in CRC progression, is associated with a lower risk of CRC, particularly tumors with a mucinous component, Crohn’s-like reaction and MSI-high. Our findings offer the promise of risk stratification of inflammatory bowel disease patients for complications such as CRC. PMID:26071399

  9. Identification of beta1C-2, a novel variant of the integrin beta1 subunit generated by utilization of an alternative splice acceptor site in exon C.

    PubMed Central

    Svineng, G; Fässler, R; Johansson, S

    1998-01-01

    A new splice variant of the human integrin subunit beta1 has been identified and designated beta1C-2. It differs from the previously reported beta1C (in this report designated beta1C-1) by 18 nucleotides, and is generated by splicing from exon 6 to an alternative splice acceptor site within exon C, causing an in-frame deletion of six amino acids of the cytoplasmic region of beta1C-1. The beta1C-2 mRNA is present in several human cell lines and tissues at low levels, similarly to beta1C-1. In peripheral T-lymphocytes, beta1C-2 is the selectively expressed isoform. Neither beta1C-1 nor beta1C-2 mRNA could be detected in mouse tissues, and Southern hybridization of a mouse genomic beta1 clone with a human exon-C-specific probe failed to identify a corresponding mouse exon. The antisense orientation of exon C is highly homologous to an Alu element. Since Alu elements are restricted to primates, the beta1C-1 and beta1C-2 variants of the integrin subunit beta1 are specific for these species. The protein coded for by the beta1C-2 cDNA can be expressed and localized to the surface of beta1 deficient mouse cells. However, while stable transformed clones expressing high levels of the beta1A were commonly found, the beta1C-1 and beta1C-2 expressing clones expressed barely detectable amounts of the beta1 protein. Hence, high levels of beta1C-2 may be incompatible with cell proliferation, as previously suggested for beta1C-1. PMID:9494094

  10. Molecular identification of transcription factor Runx1 variants in grass carp (Ctenopharyngodon idella) and their responses to immune stimuli.

    PubMed

    Yao, Fuli; Liu, Yazhen; Du, Linyong; Wang, Xinyan; Zhang, Anying; Wei, He; Zhou, Hong

    2014-08-15

    The Runt-related transcription factor (Runx) family consists of three members, Runx1, Runx2 and Runx3 in mammals, which are involved in various biological processes. Recent studies have demonstrated that Runx1 plays critical roles in the immunity of higher vertebrates. In fish, zebrafish and fugu Runx family members have been identified, and their chromosome location, promoter usage and expression patterns have been elucidated. However, their expression profiles in immune responses are still unknown. In this study, we identified grass carp five Runx1 (gcRunx1) variants (v1-5) possibly generated through alternative promoter usage and alternative splicing. The gcRunx1 v1-3 encodes the proteins possessing intact structural characteristics of Runx family, but the putative proteins of gcRunx1 v4-5 lack a transactivation domain, an inhibitory domain and a C-terminal pentapeptide motif (VWRPY). Tissue distribution assays revealed that gcRunx1 was preferentially expressed in some immune-related tissues including thymus and spleen, indicating its potential roles in teleost immunity. The changes of gcRunx1 expression to various immune stimuli was examined in periphery blood lymphocytes, showing that gcRunx1 v1-3 mRNA levels were increased after LPS, poly I:C and PHA treatment, whereas gcRunx1 v4-5 mRNA expression were stimulated only by LPS and PHA. Furthermore, in vivo studies confirmed that bacterial challenge enhanced gcRunx1 mRNA levels. In particular, in vitro and in vivo studies revealed that gcRunx1 v4-5 mRNA expression was induced with a delayed kinetics compared with that of gcRunx1 v1-3. These findings not only provide the evidence for the involvement of gcRunx1 in immune response, but also reveal the inducible expression diversity of fish Runx1 splicing variants, thereby facilitating further elucidating the role of Runx1 in piscine immunity. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. EcoTILLING for the identification of allelic variants of melon eIF4E, a factor that controls virus susceptibility

    PubMed Central

    Nieto, Cristina; Piron, Florence; Dalmais, Marion; Marco, Cristina F; Moriones, Enrique; Gómez-Guillamón, Ma Luisa; Truniger, Verónica; Gómez, Pedro; Garcia-Mas, Jordi; Aranda, Miguel A; Bendahmane, Abdelhafid

    2007-01-01

    Background Translation initiation factors of the 4E and 4G protein families mediate resistance to several RNA plant viruses in the natural diversity of crops. Particularly, a single point mutation in melon eukaryotic translation initiation factor 4E (eIF4E) controls resistance to Melon necrotic spot virus (MNSV) in melon. Identification of allelic variants within natural populations by EcoTILLING has become a rapid genotype discovery method. Results A collection of Cucumis spp. was characterised for susceptibility to MNSV and Cucumber vein yellowing virus (CVYV) and used for the implementation of EcoTILLING to identify new allelic variants of eIF4E. A high conservation of eIF4E exonic regions was found, with six polymorphic sites identified out of EcoTILLING 113 accessions. Sequencing of regions surrounding polymorphisms revealed that all of them corresponded to silent nucleotide changes and just one to a non-silent change correlating with MNSV resistance. Except for the MNSV case, no correlation was found between variation of eIF4E and virus resistance, suggesting the implication of different and/or additional genes in previously identified resistance phenotypes. We have also characterized a new allele of eIF4E from Cucumis zeyheri, a wild relative of melon. Functional analyses suggested that this new eIF4E allele might be responsible for resistance to MNSV. Conclusion This study shows the applicability of EcoTILLING in Cucumis spp., but given the conservation of eIF4E, new candidate genes should probably be considered to identify new sources of resistance to plant viruses. Part of the methodology described here could alternatively be used in TILLING experiments that serve to generate new eIF4E alleles. PMID:17584936

  12. Identification of a conserved aggregation-prone intermediate state in the folding pathways of Spc-SH3 amyloidogenic variants.

    PubMed

    Krobath, H; Estácio, S G; Faísca, P F N; Shakhnovich, E I

    2012-10-05

    We compared the folding pathways of selected mutational variants of the α-spectrin SH3 domain (Spc-SH3) by using a continuum model that combines a full atomistic protein representation with the Gō potential. Experimental data show that the N47G mutant shows very little tendency to aggregate while the N47A and triple mutant D48G(2Y) are both amyloidogenic, with the latter being clearly more aggregation prone. We identified a strikingly similar native-like folding intermediate across the three mutants, in which strand β(1) is totally unstructured and more than half of the major hydrophobic core residues are highly solvent exposed. Results from extensive docking simulations show that the ability of the intermediates to dimerize is largely driven by strand β(1) and is consistent with the in vitro aggregation behavior reported for the corresponding mutants. They further suggest that residues 44 and 53, which are key players in the nucleation-condensation mechanism of folding, are also important triggers of the aggregation process. Copyright © 2012 Elsevier Ltd. All rights reserved.

  13. Identification of rare variants in KCTD13 at the schizophrenia risk locus 16p11.2

    PubMed Central

    Heinemann, Barbara; Strohmaier, Jana; Pfohl, Marvin A.; Giegling, Ina; Hofmann, Andrea; Ludwig, Kerstin U.; Witt, Stephanie H.; Ludwig, Michael; Forstner, Andreas J.; Albus, Margot; Schwab, Sibylle G.; Borrmann-Hassenbach, Margitta; Lennertz, Leonard; Wagner, Michael; Hoffmann, Per; Rujescu, Dan; Maier, Wolfgang; Cichon, Sven; Rietschel, Marcella; Nöthen, Markus M.

    2016-01-01

    Duplications in 16p11.2 are a risk factor for schizophrenia (SCZ). Using genetically modified zebrafish, Golzio and colleagues identified KCTD13 within 16p11.2 as a major driver of the neuropsychiatric phenotype observed in humans. The aims of the present study were to explore the role of KCTD13 in the development of SCZ and to provide a more complete picture of the allelic architecture at this risk locus. The exons of KCTD13 were sequenced in 576 patients. The mutations c.6G>T and c.598G>A were identified in one patient each. Both mutations were predicted to be functionally relevant and were absent from the 1000 Genomes Project data and the Exome Variant Server. The mutation c.6G>T was predicted to abolish a potential transcription factor-binding site for specifity protein 1. Altered specifity protein 1 expression has been reported in SCZ patients compared with controls. Further studies in large cohorts are warranted to determine the relevance of the two identified mutations. PMID:27668412

  14. Detection and variant identification of HHV-6 by a non-radioactive hybridization microplate assay for amplimers detection.

    PubMed

    Gautheret, A; Aubin, J T; Poirel, L; Chopineau, S; Agut, H; Dussaix, E

    1996-04-26

    A non-radioactive hybridization microtiter plate assay was developed and evaluated for detection of the HHV-6 genome and to identify HHV-6 variants A and B. The viral DNA is amplified by the polymerase chain reaction using a 5'-end-biotinylated primer. The biotinylated amplimers are captured on avidin-coated microtiter plates, denaturated with sodium hydroxide and hybridized to a 3'-end-digoxigenin-labelled probe. Subsequently, anti-digoxigenin Fab fragments conjugated with alkaline phosphatase are used for the revelation of the hybridized probe. The result is obtained by measuring the intensity of light emitted with a spectrophotometer. This new assay was compared to the standard analysis of amplified products by Southern hybridization consisting of gel electrophoresis of the amplimers, transfer onto a nylon membrane, and hybridization with a 32P-labelled oligomeric probe. Both methods exhibited the same sensitivity and specificity. Thus, a non-radioactive hybridization microtiter plate assay may be a suitable alternative to isotopic techniques.

  15. Evaluation of CHROMagar Orientation for differentiation and presumptive identification of gram-negative bacilli and Enterococcus species.

    PubMed Central

    Merlino, J; Siarakas, S; Robertson, G J; Funnell, G R; Gottlieb, T; Bradbury, R

    1996-01-01

    A new chromogenic plate medium, CHROMagar Orientation, was evaluated for use in the differentiation and presumptive identification of gram-negative bacilli and Enterococcus species by a multipoint inoculation (replicator) technique. In this study, 1,404 gram-negative bacilli and 74 enterococcal isolates were tested on CHROMagar Orientation. Six control American Type Culture Collection strains were also included with the testing to ensure quality control of the media. Of the Escherichia coli isolates (n = 588) tested, 99.3% produced a pink-to-red color. Only in four isolates that were O-nitrophenyl-beta-D-galactopyranoside (ONPG) negative did this result differ. Proteus mirabilis and P. vulgaris were well differentiated on this medium. P. mirabilis (n = 184) produced a clear colony with diffusible brown pigment around the periphery. By contrast, 15 of 16 P. vulgaris isolates produced bluish-green colonies with a slight brown background. All Aeromonas hydrophila isolates (n = 26) tested produced clear to pink colonies at 35 to 37 degrees C. This colony color changed to blue after 2 to 3 h of incubation at room temperature. A. hydrophila exhibited stronger color and better growth at 30 degrees C. Serratia marcescens (n = 29) demonstrated an aqua blue color that deepened to a darker blue when exposed to room temperature. All enterococcal isolates (n = 74) resulted in a blue color and gave pinpoint colonies on purity subcultures at 35 to 37 degrees C after 18 h of incubation. Similarity in color resulted in failure to discriminate accurately between Klebsiella, Enterobacter, and Citrobacter species. However, these species could be readily differentiated from other members of the family Enterobacteriaceae. Pseudomonas aeruginosa (n = 151) was easily differentiated from members of the Enterobacteriaceae but was less easily distinguishable from other gram-negative nonmembers of the Enterobacteriaceae. The medium was found to facilitate easy visual detection of mixed

  16. Molecular cloning, genomic organization, chromosome mapping, tissues expression pattern and identification of a novel splicing variant of porcine CIDEb gene

    SciTech Connect

    Li, YanHua; Li, AiHua; Yang, Z.Q.

    2016-09-09

    Cell death-inducing DNA fragmentation factor-α-like effector b (CIDEb) is a member of the CIDE family of apoptosis-inducing factors, CIDEa and CIDEc have been reported to be Lipid droplets (LDs)-associated proteins that promote atypical LD fusion in adipocytes, and responsible for liver steatosis under fasting and obese conditions, whereas CIDEb promotes lipid storage under normal diet conditions [1], and promotes the formation of triacylglyceride-enriched VLDL particles in hepatocytes [2]. Here, we report the gene cloning, chromosome mapping, tissue distribution, genetic expression analysis, and identification of a novel splicing variant of the porcine CIDEb gene. Sequence analysis shows that the open reading frame of the normal porcine CIDEb isoform covers 660bp and encodes a 219-amino acid polypeptide, whereas its alternative splicing variant encodes a 142-amino acid polypeptide truncated at the fourth exon and comprised of the CIDE-N domain and part of the CIDE-C domain. The deduced amino acid sequence of normal porcine CIDEb shows an 85.8% similarity to the human protein and 80.0% to the mouse protein. The CIDEb genomic sequence spans approximately 6KB comprised of five exons and four introns. Radiation hybrid mapping demonstrated that porcine CIDEb is located at chromosome 7q21 and at a distance of 57cR from the most significantly linked marker, S0334, regions that are syntenic with the corresponding region in the human genome. Tissue expression analysis indicated that normal CIDEb mRNA is ubiquitously expressed in many porcine tissues. It was highly expressed in white adipose tissue and was observed at relatively high levels in the liver, lung, small intestine, lymphatic tissue and brain. The normal version of CIDEb was the predominant form in all tested tissues, whereas the splicing variant was expressed at low levels in all examined tissues except the lymphatic tissue. Furthermore, genetic expression analysis indicated that CIDEb mRNA levels were

  17. Linking parental socialization about discrimination to intergroup attitudes: The role of social dominance orientation and cultural identification.

    PubMed

    Su, Jenny C; Gries, Peter H; Lee, I-Ching; Tran, Alisia G T T

    2017-07-01

    This study investigated the interaction of parental socialization about discrimination and social dominance orientation (SDO) in predicting the cultural identity and intergroup attitudes of the Minnanese, an ethnic group in Taiwan that faced systematic discrimination during the early decades of Chinese Nationalist rule. Because high SDO individuals tend to support group-based dominance, we hypothesized that under high preparation for bias, which may reinforce narratives that place the historically disadvantaged Taiwanese in a subordinate position, Minnanese high in SDO would identify less with Taiwanese and more with Chinese (the historically high-status outgroup) compared with their low SDO counterparts. We examined our hypotheses using a sample of Minnanese (N = 365; 183 women, 182 men; average age = 44.35) who participated in a nationally representative survey of Taiwanese adults. As predicted, among Minnanese exposed to high levels of preparation for bias, those with high SDO expressed greater levels of Chinese identification and more favorable attitudes toward Chinese than their low SDO counterparts (no difference was found in attitudes toward Taiwanese). Among Minnanese exposed to low levels of preparation for bias, SDO predicted neither Chinese nor Taiwanese identity. Moreover, the interaction effect of preparation for bias and SDO on attitudes toward Chinese was mediated by Chinese identity. Using a unique, non-Western sample, this study demonstrated the role that parental socialization about past discrimination, in combination with belief in group-based dominance, plays in the construction of group identity and intergroup attitudes among members of historically disadvantaged ethnic groups. (PsycINFO Database Record (c) 2017 APA, all rights reserved).

  18. Identification, characterization and target gene analysis of testicular microRNAs in the oriental fruit fly Bactrocera dorsalis.

    PubMed

    Tariq, K; Peng, W; Saccone, G; Zhang, H

    2016-02-01

    MicroRNAs (miRNAs) are small noncoding RNAs that regulate various diverse biological processes including insect spermatogenesis. The oriental fruit fly, Bactrocera dorsalis, is one of the most destructive horticultural pests in East Asia and the Pacific region. Although developmental miRNA profiles of B. dorsalis have enriched our knowledge, specific testicular miRNAs in this dipteran species are unexplored. In this study, we identified miRNAs from B. dorsalis testes by deep sequencing, which provided an overview of miRNA expression during spermatogenesis. Small RNA libraries were constructed from the testes of fully mature (FM), immature (IM) and middle-aged (MA) adult flies of B. dorsalis. Small RNA sequencing and data analysis revealed 172 known and 78 novel miRNAs amongst these libraries. Pairwise comparisons of libraries led to the identification of 24, 15 and 14 differentially expressed miRNAs in FM vs. IM, FM vs. MA and IM vs. MA insects, respectively. Using a bioinformatic approach, we predicted 124 target genes against the 13 most differentially expressed miRNAs. Furthermore, the expression patterns of six randomly selected miRNAs (from the 13 most differentially expressed miRNAs) and their putative target genes (from the 124 predicted target genes) were analysed in the testis of B. dorsalis by quantitative real-time PCR, which showed that out of six, four tested miRNAs-mRNAs had an inverse expression pattern and are probably co-regulated. This study is the first comparative profile of the miRNA transcriptome in three developmental stages of the testis, and provides a useful resource for further studies on the role of miRNAs in spermatogenesis in B. dorsalis. © 2015 The Royal Entomological Society.

  19. Identification of Bari Transposons in 23 Sequenced Drosophila Genomes Reveals Novel Structural Variants, MITEs and Horizontal Transfer

    PubMed Central

    D’Addabbo, Pietro; Caizzi, Ruggiero

    2016-01-01

    Bari elements are members of the Tc1-mariner superfamily of DNA transposons, originally discovered in Drosophila melanogaster, and subsequently identified in silico in 11 sequenced Drosophila genomes and as experimentally isolated in four non-sequenced Drosophila species. Bari-like elements have been also studied for their mobility both in vivo and in vitro. We analyzed 23 Drosophila genomes and carried out a detailed characterization of the Bari elements identified, including those from the heterochromatic Bari1 cluster in D. melanogaster. We have annotated 401 copies of Bari elements classified either as putatively autonomous or inactive according to the structure of the terminal sequences and the presence of a complete transposase-coding region. Analyses of the integration sites revealed that Bari transposase prefers AT-rich sequences in which the TA target is cleaved and duplicated. Furthermore evaluation of transposon’s co-occurrence near the integration sites of Bari elements showed a non-random distribution of other transposable elements. We also unveil the existence of a putatively autonomous Bari1 variant characterized by two identical long Terminal Inverted Repeats, in D. rhopaloa. In addition, we detected MITEs related to Bari transposons in 9 species. Phylogenetic analyses based on transposase gene and the terminal sequences confirmed that Bari-like elements are distributed into three subfamilies. A few inconsistencies in Bari phylogenetic tree with respect to the Drosophila species tree could be explained by the occurrence of horizontal transfer events as also suggested by the results of dS analyses. This study further clarifies the Bari transposon’s evolutionary dynamics and increases our understanding on the Tc1-mariner elements’ biology. PMID:27213270

  20. Identification of a Putative Crf Splice Variant and Generation of Recombinant Antibodies for the Specific Detection of Aspergillus fumigatus

    PubMed Central

    Schütte, Mark; Thullier, Philippe; Pelat, Thibaut; Wezler, Xenia; Rosenstock, Philip; Hinz, Dominik; Kirsch, Martina Inga; Hasenberg, Mike; Frank, Ronald; Schirrmann, Thomas; Gunzer, Matthias

    2009-01-01

    Background Aspergillus fumigatus is a common airborne fungal pathogen for humans. It frequently causes an invasive aspergillosis (IA) in immunocompromised patients with poor prognosis. Potent antifungal drugs are very expensive and cause serious adverse effects. Their correct application requires an early and specific diagnosis of IA, which is still not properly achievable. This work aims to a specific detection of A. fumigatus by immunofluorescence and the generation of recombinant antibodies for the detection of A. fumigatus by ELISA. Results The A. fumigatus antigen Crf2 was isolated from a human patient with proven IA. It is a novel variant of a group of surface proteins (Crf1, Asp f9, Asp f16) which belong to the glycosylhydrolase family. Single chain fragment variables (scFvs) were obtained by phage display from a human naive antibody gene library and an immune antibody gene library generated from a macaque immunized with recombinant Crf2. Two different selection strategies were performed and shown to influence the selection of scFvs recognizing the Crf2 antigen in its native conformation. Using these antibodies, Crf2 was localized in growing hyphae of A. fumigatus but not in spores. In addition, the antibodies allowed differentiation between A. fumigatus and related Aspergillus species or Candida albicans by immunofluorescence microscopy. The scFv antibody clones were further characterized for their affinity, the nature of their epitope, their serum stability and their detection limit of Crf2 in human serum. Conclusion Crf2 and the corresponding recombinant antibodies offer a novel approach for the early diagnostics of IA caused by A. fumigatus. PMID:19675673

  1. Identification of genes expressed by immune cells of the colon that are regulated by colorectal cancer-associated variants.

    PubMed

    Peltekova, Vanya D; Lemire, Mathieu; Qazi, Aamer M; Zaidi, Syed H E; Trinh, Quang M; Bielecki, Ryszard; Rogers, Marianne; Hodgson, Lyndsey; Wang, Mike; D'Souza, David J A; Zandi, Sasan; Chong, Taryne; Kwan, Jennifer Y Y; Kozak, Krystian; De Borja, Richard; Timms, Lee; Rangrej, Jagadish; Volar, Milica; Chan-Seng-Yue, Michelle; Beck, Timothy; Ash, Colleen; Lee, Shawna; Wang, Jianxin; Boutros, Paul C; Stein, Lincoln D; Dick, John E; Gryfe, Robert; McPherson, John D; Zanke, Brent W; Pollett, Aaron; Gallinger, Steven; Hudson, Thomas J

    2014-05-15

    A locus on human chromosome 11q23 tagged by marker rs3802842 was associated with colorectal cancer (CRC) in a genome-wide association study; this finding has been replicated in case-control studies worldwide. In order to identify biologic factors at this locus that are related to the etiopathology of CRC, we used microarray-based target selection methods, coupled to next-generation sequencing, to study 103 kb at the 11q23 locus. We genotyped 369 putative variants from 1,030 patients with CRC (cases) and 1,061 individuals without CRC (controls) from the Ontario Familial Colorectal Cancer Registry. Two previously uncharacterized genes, COLCA1 and COLCA2, were found to be co-regulated genes that are transcribed from opposite strands. Expression levels of COLCA1 and COLCA2 transcripts correlate with rs3802842 genotypes. In colon tissues, COLCA1 co-localizes with crystalloid granules of eosinophils and granular organelles of mast cells, neutrophils, macrophages, dendritic cells and differentiated myeloid-derived cell lines. COLCA2 is present in the cytoplasm of normal epithelial, immune and other cell lineages, as well as tumor cells. Tissue microarray analysis demonstrates the association of rs3802842 with lymphocyte density in the lamina propria (p = 0.014) and levels of COLCA1 in the lamina propria (p = 0.00016) and COLCA2 (tumor cells, p = 0.0041 and lamina propria, p = 6 × 10(-5)). In conclusion, genetic, expression and immunohistochemical data implicate COLCA1 and COLCA2 in the pathogenesis of colon cancer. Histologic analyses indicate the involvement of immune pathways.

  2. Mutation analysis of methylmalonyl CoA mutase gene exon 2 in Egyptian families: Identification of 25 novel allelic variants.

    PubMed

    Ghoraba, Dina A; Mohammed, Magdy M; Zaki, Osama K

    2015-02-01

    Methylmalonic aciduria (MMA) is an autosomal recessive disorder of methylmalonate and cobalamin (cbl; vitamin B12) metabolism. It is an inborn error of organic acid metabolism which commonly results from a defect in the gene encoding the methylmalonyl-CoA mutase (MCM) apoenzyme. Here we report the results of mutation study of exon 2 of the methylmalonyl CoA mutase (MUT) gene, coding MCM residues from 1 to 128, in ten unrelated Egyptian families affected with methylmalonic aciduria. Patients were presented with a wide-anion gap metabolic acidosis. The diagnosis has established by the measurement of C3 (propionylcarnitine) and C3:C2 (propionylcarnitine/acetylcarnitine) in blood by using liquid chromatography-tandem mass spectrometry (LC/MS-MS) and was confirmed by the detection of an abnormally elevated level of methylmalonic acid in urine by using gas chromatography-mass spectrometry (GC/MS) and isocratic cation exchange high-performance liquid-chromatography (HPLC). Direct sequencing of gDNA of the MUT gene exon 2 has revealed a total of 26 allelic variants: ten of which were intronic, eight were located upstream to the exon 2 coding region, four were novel modifications predicted to affect the splicing region, three were novel mutations within the coding region: c.15G > A (p.K5K), c.165C > A (p.N55K) and c.7del (p.R3EfsX14), as well as the previously reported mutation c.323G > A (p.R108H).

  3. Identification of genetic variants in the TNF promoter associated with COPD secondary to tobacco smoking and its severity.

    PubMed

    Reséndiz-Hernández, Juan Manuel; Sansores, Raúl H; Hernández-Zenteno, Rafael de Jesús; Vargas-Alarcón, Gilber; Colín-Barenque, Laura; Velázquez-Uncal, Mónica; Camarena, Angel; Ramírez-Venegas, Alejandra; Falfán-Valencia, Ramcés

    2015-01-01

    Chronic obstructive pulmonary disease (COPD) is an inflammatory disease that arises in response to noxious particles or gases. Associations of genetic polymorphisms in TNF have been reported in Asians and Caucasians, but not in Mestizo populations. A case-control study was conducted in two stages: in the first stage, patients with COPD (COPD group, n=165) and smokers without disease (SNC group, n=165) were included and the TNF promoter sequence was determined using direct sequencing. In the second stage, the identified polymorphisms were validated by real-time polymerase chain reaction (PCR) in COPD (n=260) and SNC (n=506). In the first stage, 11 different sets of "contig" alignments were determined, of which contig 10 was found to be associated with susceptibility (P=5.0E-04, OR [odds ratio] =3.64) and contig 1 with Global Initiative for COPD (GOLD) greater grade (P=1.0E-02, OR =3.82). The single nucleotide polymorphisms found in this region were individually identified; the GA genotypes of rs1800629 (P=0.038, OR =2.07), rs56036015 (P=0.0082, OR =3.18), and rs361525 (P=1.0E-02, OR =4.220) were higher in the COPD group vs the SNC group; after second-stage validation, rs1800629 (P=6.00E-03, OR =2.26) and rs56036015 (P=1.10E-03, OR =2.54) are maintained. There are genetic variants in the TNF promoter associated with increased risk of COPD secondary to smoking and with a higher GOLD grade in the Mexican Mestizo population.

  4. Identification of genetic variants predictive of early onset pancreatic cancer through a population science analysis of functional genomic datasets.

    PubMed

    Chen, Jinyun; Wu, Xifeng; Huang, Yujing; Chen, Wei; Brand, Randall E; Killary, Ann M; Sen, Subrata; Frazier, Marsha L

    2016-08-30

    Biomarkers are critically needed for the early detection of pancreatic cancer (PC) are urgently needed. Our purpose was to identify a panel of genetic variants that, combined, can predict increased risk for early-onset PC and thereby identify individuals who should begin screening at an early age. Previously, we identified genes using a functional genomic approach that were aberrantly expressed in early pathways to PC tumorigenesis. We now report the discovery of single nucleotide polymorphisms (SNPs) in these genes associated with early age at diagnosis of PC using a two-phase study design. In silico and bioinformatics tools were used to examine functional relevance of the identified SNPs. Eight SNPs were consistently associated with age at diagnosis in the discovery phase, validation phase and pooled analysis. Further analysis of the joint effects of these 8 SNPs showed that, compared to participants carrying none of these unfavorable genotypes (median age at PC diagnosis 70 years), those carrying 1-2, 3-4, or 5 or more unfavorable genotypes had median ages at diagnosis of 64, 63, and 62 years, respectively (P = 3.0E-04). A gene-dosage effect was observed, with age at diagnosis inversely related to number of unfavorable genotypes (Ptrend = 1.0E-04). Using bioinformatics tools, we found that all of the 8 SNPs were predicted to play functional roles in the disruption of transcription factor and/or enhancer binding sites and most of them were expression quantitative trait loci (eQTL) of the target genes. The panel of genetic markers identified may serve as susceptibility markers for earlier PC diagnosis.

  5. Identification of Bari Transposons in 23 Sequenced Drosophila Genomes Reveals Novel Structural Variants, MITEs and Horizontal Transfer.

    PubMed

    Palazzo, Antonio; Lovero, Domenica; D'Addabbo, Pietro; Caizzi, Ruggiero; Marsano, René Massimiliano

    2016-01-01

    Bari elements are members of the Tc1-mariner superfamily of DNA transposons, originally discovered in Drosophila melanogaster, and subsequently identified in silico in 11 sequenced Drosophila genomes and as experimentally isolated in four non-sequenced Drosophila species. Bari-like elements have been also studied for their mobility both in vivo and in vitro. We analyzed 23 Drosophila genomes and carried out a detailed characterization of the Bari elements identified, including those from the heterochromatic Bari1 cluster in D. melanogaster. We have annotated 401 copies of Bari elements classified either as putatively autonomous or inactive according to the structure of the terminal sequences and the presence of a complete transposase-coding region. Analyses of the integration sites revealed that Bari transposase prefers AT-rich sequences in which the TA target is cleaved and duplicated. Furthermore evaluation of transposon's co-occurrence near the integration sites of Bari elements showed a non-random distribution of other transposable elements. We also unveil the existence of a putatively autonomous Bari1 variant characterized by two identical long Terminal Inverted Repeats, in D. rhopaloa. In addition, we detected MITEs related to Bari transposons in 9 species. Phylogenetic analyses based on transposase gene and the terminal sequences confirmed that Bari-like elements are distributed into three subfamilies. A few inconsistencies in Bari phylogenetic tree with respect to the Drosophila species tree could be explained by the occurrence of horizontal transfer events as also suggested by the results of dS analyses. This study further clarifies the Bari transposon's evolutionary dynamics and increases our understanding on the Tc1-mariner elements' biology.

  6. A Statistical Approach to Fine Mapping for the Identification of Potential Causal Variants Related to Bone Mineral Density.

    PubMed

    Greenbaum, Jonathan; Deng, Hong-Wen

    2017-08-01

    Although genomewide association studies (GWASs) have been able to successfully identify dozens of genetic loci associated with bone mineral density (BMD) and osteoporosis-related traits, very few of these loci have been confirmed to be causal. This is because in a given genetic region there may exist many trait-associated SNPs that are highly correlated. Although this correlation is useful for discovering novel associations, the high degree of linkage disequilibrium that persists throughout the genome presents a major challenge to discern which among these correlated variants has a direct effect on the trait. In this study we apply a recently developed Bayesian fine-mapping method, PAINTOR, to determine the SNPs that have the highest probability of causality for femoral neck (FNK) BMD and lumbar spine (LS) BMD. The advantage of this method is that it allows for the incorporation of information about GWAS summary statistics, linkage disequilibrium, and functional annotations to calculate a posterior probability of causality for SNPs across all loci of interest. We present a list of the top 10 candidate SNPs for each BMD trait to be followed up in future functional validation experiments. The SNPs rs2566752 (WLS) and rs436792 (ZNF621 and CTNNB1) are particularly noteworthy because they have more than 90% probability to be causal for both FNK and LS BMD. Using this statistical fine-mapping approach we expect to gain a better understanding of the genetic determinants contributing to BMD at multiple skeletal sites. © 2017 American Society for Bone and Mineral Research. © 2017 American Society for Bone and Mineral Research.

  7. Identification of a Novel Alternative Splicing Variant of VvPMA1 in Grape Root under Salinity.

    PubMed

    Han, Ning; Ji, Xing-Long; Du, Yuan-Peng; He, Xi; Zhao, Xin-Jie; Zhai, Heng

    2017-01-01

    It has been well-demonstrated that the control of plasma membrane H(+)-ATPase (PM H(+)-ATPase) activity is important to plant salt tolerance. This study found a significant increase in PM H(+)-ATPase (PMA) activity in grape root exposed to NaCl. Furthermore, 7 Vitis vinifera PM H(+)-ATPase genes (VvPMAs) were identified within the grape genome and the expression response of these VvPMAs in grape root under salinity was analyzed. Two VvPMAs (VvPMA1 and VvPMA3) were expressed more strongly in roots than the other five VvPMAs. Moreover, roots exhibited diverse patterns of gene expression of VvPMA1 and VvPMA3 responses to salt stress. Interestingly, two transcripts of VvPMA1, which were created through alternative splicing (AS), were discovered and isolated from salt stressed root. Comparing the two VvPMA1 cDNA sequences (designated VvPMA1α and VvPMA1β) with the genomic sequence revealed that the second intron was retained in the VvPMA1β cDNA. This intron retention was predicted to generate a novel VvPMA1 through N-terminal truncation because of a 5'- terminal frame shift. Yeast complementation assays of the two splice variants showed that VvPMA1β could enhance the ability to complement Saccharomyces cerevisiae deficient in PM H(+)-ATPase activity. In addition, the expression profiles of VvPMA1α and VvPMA1β differed under salinity. Our data suggests that through AS, the N-terminal length of VvPMA1 may be regulated to accurately modulate PM H(+)-ATPase activity of grape root in salt stress.

  8. Mutational spectrum in congenital dyserythropoietic anemia type II: Identification of 19 novel variants in SEC23B gene

    PubMed Central

    Russo, Roberta; Esposito, Maria Rosaria; Asci, Roberta; Gambale, Antonella; Perrotta, Silverio; Ramenghi, Ugo; Forni, Gian Luca; Uygun, Vedat; Delaunay, Jean; Iolascon, Achille

    2010-01-01

    SEC23B gene encodes an essential component of the coat protein complex II (COPII)-coated vesicles. Mutations in this gene cause the vast majority the congenital dyserythropoietic anemia Type II (CDA II), a rare disorder resulting from impaired erythropoiesis. Here, we investigated 28 CDA II patients from 21 unrelated families enrolled in the CDA II International Registry. Overall, we found 19 novel variants [c.2270 A>C p.H757P; c.2149−2 A>G; c.1109+1 G>A; c.387(delG) p.L129LfsX26; c.1858 A>G p.M620V; c.1832 G>C p.R611P; c.1735 T>A p.Y579N; c.1254 T>G p.I418M; c.1015 C>T p.R339X; c.1603 C>T p.R535X; c.1654 C>T p.L552F; c.1307 C>T p.S436L; c.279+3 A>G; c. 2150(delC) p.A717VfsX7; c.1733 T>C p.L578P; c.1109+5 G>A; c.221+31 A>G; c.367 C>T p.R123X; c.1857_1859delCAT; p.I619del] in the homozygous or the compound heterozygous state. Homozygosity or compound heterozygosity for two nonsense mutations was never found. In four cases the sequencing analysis has failed to find two mutations. To discuss the putative functional consequences of missense mutations, computational analysis and sequence alignment were performed. Our data underscore the high allelic heterogeneity of CDA II, as the most of SEC23B variations are inherited as private mutations. In this mutation update, we also provided a tool to improve and facilitate the molecular diagnosis of CDA II by defining the frequency of mutations in each exon. Am. J. Hematol., 2010. © 2010 Wiley-Liss, Inc. PMID:20941788

  9. Sensitive and Specific Identification of Wild Type and Variant Proteins from 8 to 669 kDa Using Top-down Mass Spectrometry*S⃞

    PubMed Central

    Karabacak, N. Murat; Li, Long; Tiwari, Ashutosh; Hayward, Lawrence J.; Hong, Pengyu; Easterling, Michael L.; Agar, Jeffrey N.

    2009-01-01

    Top-down and bottom-up mass spectrometry methods can generate gas phase fragments and use these to identify proteins. Top-down methods, in addition, can provide the mass of the protein itself and therefore additional structural information. Despite the conceptual advantage of top-down methods, the market share advantage belongs to bottom-up methods as a result of their more robust sample preparation, fragmentation, and data processing methods. Here we report improved fragmentation and data processing methods for top-down mass spectrometry. Specifically we report the use of funnel-skimmer dissociation, a variation of nozzle-skimmer dissociation, and compare its performance with electron capture dissociation. We also debut BIG Mascot, an extended version of Mascot with incorporated top-down MS2 search ability and the first search engine that can perform both bottom-up and top-down searches. Using BIG Mascot, we demonstrated the ability to identify proteins 1) using only intact protein MS1, 2) using only MS2, and 3) using the combination of MS1 and MS2. We correctly identified proteins with a wide range of masses, including 13 amyotrophic lateral sclerosis-associated variants of the protein Cu/Zn-superoxide dismutase, and extended the upper mass limit of top-down protein identification to 669 kDa by identifying thyroglobulin. PMID:19074999

  10. Genome-wide identification of histone H2A and histone variant H2A.Z-interacting proteins by bPPI-seq.

    PubMed

    Zhang, Yi; Ku, Wai Lim; Liu, Shuai; Cui, Kairong; Jin, Wenfei; Tang, Qingsong; Lu, William; Ni, Bing; Zhao, Keji

    2017-09-01

    H2A is a nucleosome core subunit involved in organizing DNA into a chromatin structure that is often inaccessible to regulatory enzymes. Replacement of H2A by its variant H2A.Z renders chromatin accessible at enhancers and promoters. However, it remains unclear how H2A.Z functions so differently from canonical H2A. Here we report the genome-wide identification of proteins that directly interact with H2A and H2A.Z in vivo using a novel strategy, bPPI-seq. We show that bPPI-seq is a sensitive and robust technique to identify protein-protein interactions in vivo. Our data indicate that H2A.Z-interacting proteins and H2A-interacting proteins participate in distinct biological processes. In contrast to H2A-interacting proteins, the H2A.Z-interacting proteins are involved in transcriptional regulation. We found that the transcription factor Osr1 interacts with H2A.Z both in vitro and in vivo. It also mediates H2A.Z incorporation to a large number of target sites and regulates gene expression. Our data indicate that bPPI-seq can be widely applied to identify genome-wide interacting proteins under physiological conditions.Cell Research advance online publication 1 September 2017; doi:10.1038/cr.2017.112.

  11. Functional identification of a novel 14-3-3 epsilon splicing variant suggests dimerization is not necessary for 14-3-3 epsilon to inhibit UV-induced apoptosis

    SciTech Connect

    Han, Dingding; Ye, Guangming; Liu, Tingting; Chen, Cong; Yang, Xianmei; Wan, Bo; Pan, Yuanwang; Yu, Long

    2010-05-28

    14-3-3 proteins function as a dimer and have been identified to involve in diverse signaling pathways. Here we reported the identification of a novel splicing variant of human 14-3-3 epsilon (14-3-3 epsilon sv), which is derived from a novel exon 1' insertion. The insertion contains a stop codon and leads to a truncated splicing variant of 14-3-3 epsilon. The splicing variant is translated from the exon 2 and results in the deletion of an N-terminal {alpha}-helix which is crucial for the dimerization. Therefore, the 14-3-3 epsilon sv could not form a dimer with 14-3-3 zeta. However, after UV irradiation 14-3-3 epsilon sv could also support cell survival, suggesting monomer of 14-3-3 epsilon is sufficient to protect cell from apoptosis.

  12. Identification of rare DNA sequence variants in high-risk autism families and their prevalence in a large case/control population

    PubMed Central

    2014-01-01

    Background Genetics clearly plays a major role in the etiology of autism spectrum disorders (ASDs), but studies to date are only beginning to characterize the causal genetic variants responsible. Until recently, studies using multiple extended multi-generation families to identify ASD risk genes had not been undertaken. Methods We identified haplotypes shared among individuals with ASDs in large multiplex families, followed by targeted DNA capture and sequencing to identify potential causal variants. We also assayed the prevalence of the identified variants in a large ASD case/control population. Results We identified 584 non-conservative missense, nonsense, frameshift and splice site variants that might predispose to autism in our high-risk families. Eleven of these variants were observed to have odds ratios greater than 1.5 in a set of 1,541 unrelated children with autism and 5,785 controls. Three variants, in the RAB11FIP5, ABP1, and JMJD7-PLA2G4B genes, each were observed in a single case and not in any controls. These variants also were not seen in public sequence databases, suggesting that they may be rare causal ASD variants. Twenty-eight additional rare variants were observed only in high-risk ASD families. Collectively, these 39 variants identify 36 genes as ASD risk genes. Segregation of sequence variants and of copy number variants previously detected in these families reveals a complex pattern, with only a RAB11FIP5 variant segregating to all affected individuals in one two-generation pedigree. Some affected individuals were found to have multiple potential risk alleles, including sequence variants and copy number variants (CNVs), suggesting that the high incidence of autism in these families could be best explained by variants at multiple loci. Conclusions Our study is the first to use haplotype sharing to identify familial ASD risk loci. In total, we identified 39 variants in 36 genes that may confer a genetic risk of developing autism. The

  13. Cellulase variants

    DOEpatents

    Blazej, Robert; Toriello, Nicholas; Emrich, Charles; Cohen, Richard N.; Koppel, Nitzan

    2015-07-14

    This invention provides novel variant cellulolytic enzymes having improved activity and/or stability. In certain embodiments the variant cellulotyic enzymes comprise a glycoside hydrolase with or comprising a substitution at one or more positions corresponding to one or more of residues F64, A226, and/or E246 in Thermobifida fusca Cel9A enzyme. In certain embodiments the glycoside hydrolase is a variant of a family 9 glycoside hydrolase. In certain embodiments the glycoside hydrolase is a variant of a theme B family 9 glycoside hydrolase.

  14. Usefulness and acceptability of a standardised orientation and mobility training for partially-sighted older adults using an identification cane

    PubMed Central

    2012-01-01

    Background Orientation and mobility (O&M) training in using an identification (ID) cane is provided to partially-sighted older adults to facilitate independent functioning and participation in the community. Recently, a protocolised standardised O&M-training in the use of the ID cane was developed in The Netherlands. The purpose of this study is to assess the usefulness and acceptability of both the standardised training and the regular training for participants and O&M-trainers in a randomised controlled trial (NCT00946062). Methods The standardised O&M-training consists of two structured face-to-face sessions and one telephone follow-up, in which, in addition to the regular training, self-management and behavioural change techniques are applied. Questionnaires and interviews were used to collect data on the training’s usefulness, e.g. the population reached, self-reported benefits or achievements, and acceptability, e.g. the performance of the intervention according to protocol and participants’ exposure to and engagement in the training. Results Data was collected from 29 O&M-trainers and 68 participants. Regarding the self-reported benefits, outcomes were comparable for the standardised training and the regular training according the trainers and participants e.g., about 85% of the participants in both groups experienced benefits of the cane and about 70% gained confidence in their capabilities. Participants were actively involved in the standardised training. Nearly 40% of the participants in the standardised training group was not exposed to the training according to protocol regarding the number of sessions scheduled and several intervention elements, such as action planning and contracting. Conclusions The standardised and regular O&M-training showed to be useful and mostly acceptable for the partially-sighted older adults and trainers. Yet, a concern is the deviation from the protocol of the standardised O&M-training by the O&M-trainers regarding

  15. The impact of professional and organizational identification on the relationship between hospital-physician exchange and customer-oriented behaviour of physicians.

    PubMed

    Trybou, Jeroen; De Caluwé, Gaelle; Verleye, Katrien; Gemmel, Paul; Annemans, Lieven

    2015-02-17

    Hospitals face increasingly competitive market conditions. In this challenging environment, hospitals have been struggling to build high-quality hospital-physician relationships. In the literature, two types of managerial strategies for optimizing relationships have been identified. The first focuses on optimizing the economic relationship; the second focuses on the noneconomic dimension and emphasizes the cooperative structure and collaborative nature of the hospital-physician relationship. We investigate potential spillover effects between the perceptions of physicians of organizational exchange and their customer-oriented behaviors. A cross-sectional study was conducted on 130 self-employed physicians practicing at six Belgian hospitals. Economic exchange was measured using the concept of distributive justice (DJ); noneconomic exchange was measured by the concept of perceived organizational support (POS). Our outcomes consist of three types of customer-oriented behaviours: internal influence (II), external representation (ER), and service delivery (SD). Our results show a positive relationship between DJ and II (adjusted R(2) = 0.038, t = 2.35; p = 0.028) and ER (adjusted R(2) = 0.15, t = 4.59; p < 0.001) and a positive relationship between POS and II (adjusted R(2) = 0.032, t = 2.26; p = 0.026) and ER (adjusted R(2) = 0.22, t = 5.81; p < 0.001). No relationship was present between DJ (p = 0.54) or POS (p = 0.57) and SD. Organizational identification positively moderates the relationship between POS and ER (p = 0.045) and between DJ and ER (p = 0.056). The relationships between POS and II (p = 0.54) and between DJ and II (p = 0.99) were not moderated by OI. Professional identification did not moderate the studied relationships. Our results demonstrate that both perceptions of economic and noneconomic exchange are important to self-employed physicians' customer-oriented behaviours. Fostering organizational identification could enhance this reciprocity

  16. Differential expression of P-type ATPases in intestinal epithelial cells: Identification of putative new atp1a1 splice-variant

    SciTech Connect

    Rocafull, Miguel A.; Thomas, Luz E.; Barrera, Girolamo J.; Castillo, Jesus R. del

    2010-01-01

    P-type ATPases are membrane proteins that couple ATP hydrolysis with cation transport across the membrane. Ten different subtypes have been described. In mammalia, 15 genes of P-type ATPases from subtypes II-A, II-B and II-C, that transport low-atomic-weight cations (Ca{sup 2+}, Na{sup +}, K{sup +} and H{sup +}), have been reported. They include reticulum and plasma-membrane Ca{sup 2+}-ATPases, Na{sup +}/K{sup +}-ATPase and H{sup +}/K{sup +}-ATPases. Enterocytes and colonocytes show functional differences, which seem to be partially due to the differential expression of P-type ATPases. These enzymes have 9 structural motifs, being the phosphorylation (E) and the Mg{sup 2+}ATP-binding (H) motifs the most preserved. These structural characteristics permitted developing a Multiplex-Nested-PCR (MN-PCR) for the simultaneous identification of different P-type ATPases. Thus, using MN-PCR, seven different cDNAs were cloned from enterocytes and colonocytes, including SERCA3, SERCA2, Na{sup +}/K{sup +}-ATPase {alpha}1-isoform, H{sup +}/K{sup +}-ATPase {alpha}2-isoform, PMCA1, PMCA4 and a cDNA-fragment that seems to be a new cassette-type splice-variant of the atp1a1 gen. PMCA4 in enterocytes and H{sup +}/K{sup +}-ATPase {alpha}2-isoform in colonocytes were differentially expressed. This cell-specific expression pattern is related with the distinctive enterocyte and colonocyte functions.

  17. Identification with monoclonal antibody 140.240 of a structural variant of melanotransferrin shed by human melanoma cell lines in vitro.

    PubMed

    Liao, S K

    1996-01-01

    Shedding by cultured human melanoma cells of a well-characterised cell- surface glycoprotein antigen known as "melanotransferrin" was studied with two monoclonal antibodies, 140.240 and 96.5. By means of [35S]-cysteine metabolically-labelled melanoma cells and immunoprecipitation studies, identification was made, by 140.240 in the spent media of two of six melanoma cell lines, of a new molecule of 100-kDa, aside from the 88-kDa molecule. Only the 88-kDa shed molecule was detected in the remaining four melanoma cell lines with both antibodies. None of nine clonal sublines derived from the two melanoma cell lines were found to shed the 100-kDa or 88-kDa molecule exclusively. Both shed antigens were released spontaneously to the medium from the live melanoma cells rather than as a result of cell death and lysis, since there was no obvious cell death or debris in the spent medium nor in the monolayer cells detected at the time of spent medium collection. Digestion of the isolated 100-kDa and 88-kDa shed molecules with N-glycanase followed by SDS-polyacrylamide gel electrophoresis resulted in the appearance of a single band of the 77-kDa molecule, which is deduced to be the polypeptide precursor of the cell-associated 87-kDa antigen. It is concluded that some melanoma lines shed the variant 100-kDa molecule, in addition to the 88-kDa molecule, and that both shed molecules and their cellular counterpart 87-kDa differ in their degrees of glycosylation.

  18. Identification and characterization of the tyrosinase gene (TYR) and its transcript variants (TYR_1 and TYR_2) in the crab-eating macaque (Macaca fascicularis).

    PubMed

    Kim, Young-Hyun; Park, Sang-Je; Choe, Se-Hee; Lee, Ja-Rang; Cho, Hyeon-Mu; Kim, Sun-Uk; Kim, Ji-Su; Sim, Bo-Woong; Song, Bong-Seok; Lee, Youngjeon; Jin, Yeung Bae; Hong, Jung-Joo; Jeong, Kang-Jin; Kang, Philyong; Baek, Seung-Ho; Lee, Sang-Rae; Huh, Jae-Won; Chang, Kyu-Tae

    2017-09-30

    Tyrosinase is a copper-containing enzyme that regulates melanin biosynthesis and is encoded by the tyrosinase (TYR) gene. Previous studies demonstrated that mutations in TYR could lead to oculocutaneous albinism type 1 (OCA1) owing to the failure of melanin formation. Although a previous study found that albinism in the rhesus monkey was derived from a mutation in TYR, the identification and characterization of this gene in non-human primates has not been achieved thus far. Thus, using the rapid amplification of cDNA ends (RACE) and internal reverse transcription PCR (RT-PCR) we identified the full-length sequence of TYR in the crab-eating macaque, and two different transcript variants (TYR_1 and TYR_2). While TYR_1 comprised five exons and its coding sequence was highly similar to that of humans, TYR_2 comprised four exons and was generated by a third-exon-skipping event. Interestingly, these two transcripts were also present in the African green monkey (Old World monkey) and the common marmoset (New World monkey). Deduced amino acid sequence analyses revealed that TYR_2 had a shorter C-terminal region than TYR_1 owing to the exon-skipping event. Thus, the present study is the first to identify and characterize a full-length TYR gene in a non-human primate, while the further validation of the third-exon-skipping in TYR indicates that this event is well conserved in the primate lineage. Therefore, this study provides useful and important information for the study of albinism using non-human primate models. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  19. Characterization of glucagon-like peptide 1 receptor (GLP1R) gene in chickens: functional analysis, tissue distribution, and identification of its transcript variants.

    PubMed

    Huang, G; Li, J; Fu, H; Yan, Z; Bu, G; He, X; Wang, Y

    2012-07-01

    Glucagon-like peptide 1 (GLP1) receptor plays a critical role in mediating the biological actions of GLP1 in mammals and fish; however, the gene structure, expression, and functionality of GLP1 receptor (GLP1R) remain largely unknown in birds. In this study, the full-length cDNA of chicken GLP1R (cGLP1R) was first cloned from brain tissue by reverse transcription PCR. The putative cGLP1R is 459 amino acids in length and shares high amino acid sequence identity with that of human (79%), rat (80%), and Xenopus (75%). Using a pGL3-CRE luciferase reporter system, we found that cGLP1R expressed in Chinese hamster ovary cells could be potently activated by cGLP1 (EC(50), 0.11 nM) but not by other structurally related peptides, indicating that cGLP1R is a functional receptor specific to cGLP1. Interestingly, in addition to identification of the transcript encoding cGLP1R of 459 amino acids, eight transcript variants, which were generated by alternative mRNA splicing and predicted to encode either C-terminally or N-terminally truncated cGLP1Rs, were also identified from chicken brain or testis. In line with this finding, multiple cGLP1R transcripts were detected to be expressed in most chicken tissues examined, including pancreas, gastrointestinal tract, and various brain regions by reverse transcription PCR. Using the dual-luciferase reporter assay system, we further found that the 5'-flanking region of cGLP1R gene displays promoter activities in cultured HepG2 and HEK293 cells, suggesting that it may control cGLP1R gene transcription in chicken tissues, including nonpancreatic tissues. Taken together, the results from the present study establish a molecular basis to investigate the roles of GLP1 in chickens.

  20. Identification of the chemical constituents of Chinese medicine Yi-Xin-Shu capsule by molecular feature orientated precursor ion selection and tandem mass spectrometry structure elucidation.

    PubMed

    Wang, Hong-ping; Chen, Chang; Liu, Yan; Yang, Hong-Jun; Wu, Hong-Wei; Xiao, Hong-Bin

    2015-11-01

    The incomplete identification of the chemical components of traditional Chinese medicinal formula has been one of the bottlenecks in the modernization of traditional Chinese medicine. Tandem mass spectrometry has been widely used for the identification of chemical substances. Current automatic tandem mass spectrometry acquisition, where precursor ions were selected according to their signal intensity, encounters a drawback in chemical substances identification when samples contain many overlapping signals. Compounds in minor or trace amounts could not be identified because most tandem mass spectrometry information was lost. Herein, a molecular feature orientated precursor ion selection and tandem mass spectrometry structure elucidation method for complex Chinese medicine chemical constituent analysis was developed. The precursor ions were selected according to their two-dimensional characteristics of retention times and mass-to-charge ratio ranges from herbal compounds, so that all precursor ions from herbal compounds were included and more minor chemical constituents in Chinese medicine were identified. Compared to the conventional automatic tandem mass spectrometry setups, the approach is novel and can overcome the drawback for chemical substances identification. As an example, 276 compounds from the Chinese Medicine of Yi-Xin-Shu capsule were identified. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. CADD score has limited clinical validity for the identification of pathogenic variants in non-coding regions in a hereditary cancer panel

    PubMed Central

    Mather, Cheryl A; Mooney, Sean D; Salipante, Stephen J; Scroggins, Sheena; Wu, David; Pritchard, Colin C; Shirts, Brian H

    2016-01-01

    PURPOSE Several in silico tools have been shown to have reasonable research sensitivity and specificity for classifying sequence variants in coding regions. The recently-developed Combined Annotation Dependent Depletion (CADD) method generates predictive scores for single nucleotide variants (SNVs) in all areas of the genome, including non-coding regions. We sought to determine the clinical validity of non-coding variant CADD scores. METHODS We evaluated 12,391 unique SNVs in 624 patient samples submitted for germline mutation testing in a cancer-related gene panel. We compared the distributions of CADD scores of rare SNVs, common SNVs in our patient population, and the null distribution of all possible SNVs stratifying by genomic region. RESULTS The median CADD scores of intronic and nonsynonymous variants were significantly different between rare and common SNVs (p<0.0001). Despite these different distributions, no individual variants could be identified as plausibly causative among rare intronic variants with the highest scores. The ROC AUC for non-coding variants is modest, and the positive predictive value of CADD for intronic variants in panel testing was found to be 0.088. CONCLUSION Focused in-silico scoring systems with much higher predictive value will be necessary for clinical genomic applications. PMID:27148939

  2. Identification of a rare variant haemoglobin (Hb Sinai-Baltimore) causing spuriously low haemoglobin A(1c) values on ion exchange chromatography.

    PubMed

    Smith, Geoff; Murray, Heather; Brennan, Stephen O

    2013-01-01

    Commonly used methods for assay of haemoglobin A(1c) (HbA(1c)) are susceptible to interference from the presence of haemoglobin variants. In many systems, the common variants can be identified but scientists and pathologists must remain vigilant for more subtle variants that may result in spuriously high or low HbA(1c) values. It is clearly important to recognize these events whether HbA(1c) is being used as a monitoring tool or, as is increasingly the case, for diagnostic purposes. We report a patient with a rare haemoglobin variant (Hb Sinai-Baltimore) that resulted in spuriously low values of HbA(1c) when assayed using ion exchange chromatography, and the steps taken to elucidate the nature of the variant.

  3. Structural Identification of a Non-Glycosylated Variant at Ser126 for O-Glycosylation Site from EPO BRP, Human Recombinant Erythropoietin by LC/MS Analysis.

    PubMed

    Byeon, Jaehee; Lim, Yu-Ri; Kim, Hyong-Ha; Suh, Jung-Keun

    2015-06-01

    A variant peak was detected in the analysis of RP-HPLC of rHu-EPO, which has about 7% relative content. Fractions of the main and the variant peaks were pooled separately and further analyzed to identify the molecular structure of the variant peak. Total mass analysis for each peak fraction using ESI-TOF MS shows differences in molecular mass. The fraction of the main peak tends to result in higher molecular masses than the fraction of the variant. The detected masses for the variant are about 600-1000 Da smaller than those for the main peak. Peptide mapping analysis for each peak fraction using Asp-N and Glu-C shows differences in O-glycopeptide profiles at Ser126. The O-glycopeptides were not detected in the fraction of the variant. It is concluded that the variant peak is non-O-glycosylated rHu-EPO and the main peak is fully O-glycosylated rHu-EPO at Ser126.

  4. Identification and Functional Characterization of G6PC2 Coding Variants Influencing Glycemic Traits Define an Effector Transcript at the G6PC2-ABCB11 Locus

    PubMed Central

    Mahajan, Anubha; Sim, Xueling; Ng, Hui Jin; Manning, Alisa; Rivas, Manuel A.; Highland, Heather M.; Locke, Adam E.; Grarup, Niels; Im, Hae Kyung; Cingolani, Pablo; Flannick, Jason; Fontanillas, Pierre; Fuchsberger, Christian; Gaulton, Kyle J.; Teslovich, Tanya M.; Rayner, N. William; Robertson, Neil R.; Beer, Nicola L.; Rundle, Jana K.; Bork-Jensen, Jette; Ladenvall, Claes; Blancher, Christine; Buck, David; Buck, Gemma; Burtt, Noël P.; Gabriel, Stacey; Gjesing, Anette P.; Groves, Christopher J.; Hollensted, Mette; Huyghe, Jeroen R.; Jackson, Anne U.; Jun, Goo; Justesen, Johanne Marie; Mangino, Massimo; Murphy, Jacquelyn; Neville, Matt; Onofrio, Robert; Small, Kerrin S.; Stringham, Heather M.; Syvänen, Ann-Christine; Trakalo, Joseph; Abecasis, Goncalo; Bell, Graeme I.; Blangero, John; Cox, Nancy J.; Duggirala, Ravindranath; Hanis, Craig L.; Seielstad, Mark; Wilson, James G.; Christensen, Cramer; Brandslund, Ivan; Rauramaa, Rainer; Surdulescu, Gabriela L.; Doney, Alex S. F.; Lannfelt, Lars; Linneberg, Allan; Isomaa, Bo; Tuomi, Tiinamaija; Jørgensen, Marit E.; Jørgensen, Torben; Kuusisto, Johanna; Uusitupa, Matti; Salomaa, Veikko; Spector, Timothy D.; Morris, Andrew D.; Palmer, Colin N. A.; Collins, Francis S.; Mohlke, Karen L.; Bergman, Richard N.; Ingelsson, Erik; Lind, Lars; Tuomilehto, Jaakko; Hansen, Torben; Watanabe, Richard M.; Prokopenko, Inga; Dupuis, Josee; Karpe, Fredrik; Groop, Leif; Laakso, Markku; Pedersen, Oluf; Florez, Jose C.; Morris, Andrew P.; Altshuler, David; Meigs, James B.; Boehnke, Michael; McCarthy, Mark I.; Lindgren, Cecilia M.; Gloyn, Anna L.

    2015-01-01

    Genome wide association studies (GWAS) for fasting glucose (FG) and insulin (FI) have identified common variant signals which explain 4.8% and 1.2% of trait variance, respectively. It is hypothesized that low-frequency and rare variants could contribute substantially to unexplained genetic variance. To test this, we analyzed exome-array data from up to 33,231 non-diabetic individuals of European ancestry. We found exome-wide significant (P<5×10-7) evidence for two loci not previously highlighted by common variant GWAS: GLP1R (p.Ala316Thr, minor allele frequency (MAF)=1.5%) influencing FG levels, and URB2 (p.Glu594Val, MAF = 0.1%) influencing FI levels. Coding variant associations can highlight potential effector genes at (non-coding) GWAS signals. At the G6PC2/ABCB11 locus, we identified multiple coding variants in G6PC2 (p.Val219Leu, p.His177Tyr, and p.Tyr207Ser) influencing FG levels, conditionally independent of each other and the non-coding GWAS signal. In vitro assays demonstrate that these associated coding alleles result in reduced protein abundance via proteasomal degradation, establishing G6PC2 as an effector gene at this locus. Reconciliation of single-variant associations and functional effects was only possible when haplotype phase was considered. In contrast to earlier reports suggesting that, paradoxically, glucose-raising alleles at this locus are protective against type 2 diabetes (T2D), the p.Val219Leu G6PC2 variant displayed a modest but directionally consistent association with T2D risk. Coding variant associations for glycemic traits in GWAS signals highlight PCSK1, RREB1, and ZHX3 as likely effector transcripts. These coding variant association signals do not have a major impact on the trait variance explained, but they do provide valuable biological insights. PMID:25625282

  5. Identification and functional characterization of G6PC2 coding variants influencing glycemic traits define an effector transcript at the G6PC2-ABCB11 locus.

    PubMed

    Mahajan, Anubha; Sim, Xueling; Ng, Hui Jin; Manning, Alisa; Rivas, Manuel A; Highland, Heather M; Locke, Adam E; Grarup, Niels; Im, Hae Kyung; Cingolani, Pablo; Flannick, Jason; Fontanillas, Pierre; Fuchsberger, Christian; Gaulton, Kyle J; Teslovich, Tanya M; Rayner, N William; Robertson, Neil R; Beer, Nicola L; Rundle, Jana K; Bork-Jensen, Jette; Ladenvall, Claes; Blancher, Christine; Buck, David; Buck, Gemma; Burtt, Noël P; Gabriel, Stacey; Gjesing, Anette P; Groves, Christopher J; Hollensted, Mette; Huyghe, Jeroen R; Jackson, Anne U; Jun, Goo; Justesen, Johanne Marie; Mangino, Massimo; Murphy, Jacquelyn; Neville, Matt; Onofrio, Robert; Small, Kerrin S; Stringham, Heather M; Syvänen, Ann-Christine; Trakalo, Joseph; Abecasis, Goncalo; Bell, Graeme I; Blangero, John; Cox, Nancy J; Duggirala, Ravindranath; Hanis, Craig L; Seielstad, Mark; Wilson, James G; Christensen, Cramer; Brandslund, Ivan; Rauramaa, Rainer; Surdulescu, Gabriela L; Doney, Alex S F; Lannfelt, Lars; Linneberg, Allan; Isomaa, Bo; Tuomi, Tiinamaija; Jørgensen, Marit E; Jørgensen, Torben; Kuusisto, Johanna; Uusitupa, Matti; Salomaa, Veikko; Spector, Timothy D; Morris, Andrew D; Palmer, Colin N A; Collins, Francis S; Mohlke, Karen L; Bergman, Richard N; Ingelsson, Erik; Lind, Lars; Tuomilehto, Jaakko; Hansen, Torben; Watanabe, Richard M; Prokopenko, Inga; Dupuis, Josee; Karpe, Fredrik; Groop, Leif; Laakso, Markku; Pedersen, Oluf; Florez, Jose C; Morris, Andrew P; Altshuler, David; Meigs, James B; Boehnke, Michael; McCarthy, Mark I; Lindgren, Cecilia M; Gloyn, Anna L

    2015-01-01

    Genome wide association studies (GWAS) for fasting glucose (FG) and insulin (FI) have identified common variant signals which explain 4.8% and 1.2% of trait variance, respectively. It is hypothesized that low-frequency and rare variants could contribute substantially to unexplained genetic variance. To test this, we analyzed exome-array data from up to 33,231 non-diabetic individuals of European ancestry. We found exome-wide significant (P<5×10-7) evidence for two loci not previously highlighted by common variant GWAS: GLP1R (p.Ala316Thr, minor allele frequency (MAF)=1.5%) influencing FG levels, and URB2 (p.Glu594Val, MAF = 0.1%) influencing FI levels. Coding variant associations can highlight potential effector genes at (non-coding) GWAS signals. At the G6PC2/ABCB11 locus, we identified multiple coding variants in G6PC2 (p.Val219Leu, p.His177Tyr, and p.Tyr207Ser) influencing FG levels, conditionally independent of each other and the non-coding GWAS signal. In vitro assays demonstrate that these associated coding alleles result in reduced protein abundance via proteasomal degradation, establishing G6PC2 as an effector gene at this locus. Reconciliation of single-variant associations and functional effects was only possible when haplotype phase was considered. In contrast to earlier reports suggesting that, paradoxically, glucose-raising alleles at this locus are protective against type 2 diabetes (T2D), the p.Val219Leu G6PC2 variant displayed a modest but directionally consistent association with T2D risk. Coding variant associations for glycemic traits in GWAS signals highlight PCSK1, RREB1, and ZHX3 as likely effector transcripts. These coding variant association signals do not have a major impact on the trait variance explained, but they do provide valuable biological insights.

  6. Identification of strain harboring both aac(6')-Ib and aac(6')-Ib-cr variant simultaneously in Escherichia coli and Klebsiella pneumoniae.

    PubMed

    Kim, Yun-Tae; Jang, Ji-Hyun; Kim, Hyun-Chul; Kim, Hyogyeong; Lee, Kyoung-Ryul; Park, Kyung Sun; Lee, Hee-Joo; Kim, Young-Jin

    2011-04-01

    The aac(6')-Ib gene is the most prevalent gene that encodes aminoglycoside-modifying enzymes and confers resistance to tobramycin, kanamycin, and amikacin. The aac(6')-Ib-cr variant gene can induce resistance against aminoglycoside and fluoroquinolone simultaneously. Two main methods, sequence analysis and the restriction enzyme method, can detect the aac(6')-Ib-cr variant in clinical strains. We collected the 85 strains that were believed to be aac(6')-Ib positive from clinical isolates. Among them, 38 strains were the wild-type; the remaining 47 strains were the aac(6')-Ib-cr variant. Of these 47 strains, 19 simultaneously harbored aac(6')-Ib and aac(6')-Ib-cr. Our study aims to report the characteristics of the 19 strains that simultaneously harbored both genes. This study is the first investigation published in Korea of strains that included both aac(6')-Ib and aac(6')-Ib-cr variant.

  7. Identification and characterization of a spontaneously aggregating amyloid-forming variant of human PrP((90-231)) through phage-display screening of variants randomized between residues 101 and 112.

    PubMed

    Verma, Archana; Sharma, Swati; Ganguly, Nirmal Kumar; Majumdar, Siddharta; Guptasarma, Purnananda; Luthra-Guptasarma, Manni

    2008-01-01

    The N-terminal 'unstructured' region of the human prion protein [PrP((90-231))] is believed to play a role in its aggregation because mutations in this region are associated with seeding-independent deposition disorders like Gerstmann-Straussler-Scheinker disease (GSS). One way of examining the effects of such mutations is to search combinatorially derived libraries for sequence variants showing a propensity to aggregate and/or the ability to interact with prion molecules folded into a beta-sheet-based conformation (i.e., beta-PrP or PrP(Sc)). We created a library of 1.8x10(7) variants randomized between positions 101 and 112, displayed it on filamentous bacteriophage, and 'spiked' it with a approximately 25% population of phages-bearing wild-type prion (wt-PrP). Screening was performed through four rounds of biopanning and amplification against immobilized beta-PrP, and yielded three beta-PrP-binding populations: wt-PrP (26% representation) and two non-wt-PrP variants ( approximately 10% and approximately 64% representation, respectively). The remarkable enrichment of one non-wt-PrP variant (MutPrP) incorporating residues KPSKPKTNMKHM in place of KGVLTWFSPLWQ, despite its initial representation at a 5 million-fold lower level than wt-PrP, caused us to produce it and discover: (i) that it readily aggregates into thioflavin-T-binding amyloids between pH 6.0 and 9.0, (ii) that it adopts a soluble beta-sheet based monomeric structure at pH 10.0, (iii) that it is less thermally stable and more compact than wt-PrP, and (iv) that it displays significantly greater resistance to proteolysis than wt-PrP. Our results suggest that sequence variations in the 101-112 region can indeed predispose the prion for aggregation.

  8. Identification of Functional Variants for Cleft Lip with or without Cleft Palate in or near PAX7, FGFR2, and NOG by Targeted Sequencing of GWAS Loci

    PubMed Central

    Leslie, Elizabeth J.; Taub, Margaret A.; Liu, Huan; Steinberg, Karyn Meltz; Koboldt, Daniel C.; Zhang, Qunyuan; Carlson, Jenna C.; Hetmanski, Jacqueline B.; Wang, Hang; Larson, David E.; Fulton, Robert S.; Kousa, Youssef A.; Fakhouri, Walid D.; Naji, Ali; Ruczinski, Ingo; Begum, Ferdouse; Parker, Margaret M.; Busch, Tamara; Standley, Jennifer; Rigdon, Jennifer; Hecht, Jacqueline T.; Scott, Alan F.; Wehby, George L.; Christensen, Kaare; Czeizel, Andrew E.; Deleyiannis, Frederic W.-B.; Schutte, Brian C.; Wilson, Richard K.; Cornell, Robert A.; Lidral, Andrew C.; Weinstock, George M.; Beaty, Terri H.; Marazita, Mary L.; Murray, Jeffrey C.

    2015-01-01

    Although genome-wide association studies (GWASs) for nonsyndromic orofacial clefts have identified multiple strongly associated regions, the causal variants are unknown. To address this, we selected 13 regions from GWASs and other studies, performed targeted sequencing in 1,409 Asian and European trios, and carried out a series of statistical and functional analyses. Within a cluster of strongly associated common variants near NOG, we found that one, rs227727, disrupts enhancer activity. We furthermore identified significant clusters of non-coding rare variants near NTN1 and NOG and found several rare coding variants likely to affect protein function, including four nonsense variants in ARHGAP29. We confirmed 48 de novo mutations and, based on best biological evidence available, chose two of these for functional assays. One mutation in PAX7 disrupted the DNA binding of the encoded transcription factor in an in vitro assay. The second, a non-coding mutation, disrupted the activity of a neural crest enhancer downstream of FGFR2 both in vitro and in vivo. This targeted sequencing study provides strong functional evidence implicating several specific variants as primary contributory risk alleles for nonsyndromic clefting in humans. PMID:25704602

  9. Identification of functional variants for cleft lip with or without cleft palate in or near PAX7, FGFR2, and NOG by targeted sequencing of GWAS loci.

    PubMed

    Leslie, Elizabeth J; Taub, Margaret A; Liu, Huan; Steinberg, Karyn Meltz; Koboldt, Daniel C; Zhang, Qunyuan; Carlson, Jenna C; Hetmanski, Jacqueline B; Wang, Hang; Larson, David E; Fulton, Robert S; Kousa, Youssef A; Fakhouri, Walid D; Naji, Ali; Ruczinski, Ingo; Begum, Ferdouse; Parker, Margaret M; Busch, Tamara; Standley, Jennifer; Rigdon, Jennifer; Hecht, Jacqueline T; Scott, Alan F; Wehby, George L; Christensen, Kaare; Czeizel, Andrew E; Deleyiannis, Frederic W-B; Schutte, Brian C; Wilson, Richard K; Cornell, Robert A; Lidral, Andrew C; Weinstock, George M; Beaty, Terri H; Marazita, Mary L; Murray, Jeffrey C

    2015-03-05

    Although genome-wide association studies (GWASs) for nonsyndromic orofacial clefts have identified multiple strongly associated regions, the causal variants are unknown. To address this, we selected 13 regions from GWASs and other studies, performed targeted sequencing in 1,409 Asian and European trios, and carried out a series of statistical and functional analyses. Within a cluster of strongly associated common variants near NOG, we found that one, rs227727, disrupts enhancer activity. We furthermore identified significant clusters of non-coding rare variants near NTN1 and NOG and found several rare coding variants likely to affect protein function, including four nonsense variants in ARHGAP29. We confirmed 48 de novo mutations and, based on best biological evidence available, chose two of these for functional assays. One mutation in PAX7 disrupted the DNA binding of the encoded transcription factor in an in vitro assay. The second, a non-coding mutation, disrupted the activity of a neural crest enhancer downstream of FGFR2 both in vitro and in vivo. This targeted sequencing study provides strong functional evidence implicating several specific variants as primary contributory risk alleles for nonsyndromic clefting in humans. Copyright © 2015 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

  10. Identification of human IgG1 variant with enhanced FcRn binding and without increased binding to rheumatoid factor autoantibody

    PubMed Central

    Maeda, Atsuhiko; Iwayanagi, Yuki; Haraya, Kenta; Tachibana, Tatsuhiko; Nakamura, Genki; Nambu, Takeru; Esaki, Keiko; Hattori, Kunihiro; Igawa, Tomoyuki

    2017-01-01

    ABSTRACT Various studies have demonstrated that Fc engineering to enhance neonatal Fc receptor (FcRn) binding is effective for elongating half-life or increasing cellular uptake of IgG. A previous study has shown that a N434H mutation to enhance FcRn binding resulted in increased binding to rheumatoid factor (RF) autoantibody, which is not desirable for therapeutic use in autoimmune disease. In this study, we first showed that all the existing Fc variants with enhanced FcRn binding also show increased RF binding, and then identified specific mutations that could be introduced to those Fc variants to reduce the RF binding. Furthermore, we generated novel Fc variants that do not increase RF binding and show half-lives of 45 d in cynomolgus monkey, which is longer than those of previously reported Fc variants. In addition, we generated novel Fc variants with antigen sweeping activity that do not increase RF binding. We expect that these novel Fc variants will be useful as antibody therapeutics against autoimmune diseases. PMID:28387635

  11. Identification of a novel CHEK2 variant and assessment of its contribution to the risk of breast cancer in French Canadian women

    PubMed Central

    Novak, David J; Chen, Long Qi; Ghadirian, Parviz; Hamel, Nancy; Zhang, Phil; Rossiny, Vanessa; Cardinal, Guy; Robidoux, André; Tonin, Patricia N; Rousseau, Francois; Narod, Steven A; Foulkes, William D

    2008-01-01

    Background BRCA1 and BRCA2 account for the majority of the known familial breast cancer risk, however, the impact of other cancer susceptibility genes largely remains to be elucidated. Checkpoint Kinase 2 (CHEK2) is an important signal transducer of cellular responses to DNA damage, whose defects have been associated with an increase in breast cancer risk. Previous studies have identified low penetrance CHEK2 alleles such as 1100delC and I157T, as well as variants such as S428F in the Ashkenazi Jewish population and IVS2 + 1G>A in the Polish population. No founder allele has been specifically identified in the French Canadian population. Methods The 14 coding exons of CHEK2 were fully sequenced for variant alleles in a panel of 25 affected French Canadian women and 25 healthy controls. Two variants were identified of which one novel variant was further screened for in an additional panel of 667 breast cancer patients and 6548 healthy controls. Additional genotyping was conducted using allele specific PCR and a restriction digest assay. Significance of amino acid substitutions were deduced by employing comparative analysis techniques. Results Two variants were identified: the previously reported silent substitution 252A>G (E84E) and the novel missense variant, 1217G>A (R406H). No significant difference in allele distribution between French Canadian women with breast cancer and healthy controls was observed (3/692, 0.43% vs. 22/6573, 0.33%, respectively, P = 0.73). Conclusion The novel CHEK2 missense variant identified in this study, R406H, is unlikely to contribute to breast cancer risk in French Canadian women. PMID:18706089

  12. Characterization and identification of alanine to serine sequence variants in an IgG4 monoclonal antibody produced in mammalian cell lines.

    PubMed

    Fu, Jinmei; Bongers, Jacob; Tao, Li; Huang, Dan; Ludwig, Richard; Huang, Yunping; Qian, Yueming; Basch, Jonathan; Goldstein, Joel; Krishnan, Ramji; You, Li; Li, Zheng Jian; Russell, Reb J

    2012-11-01

    Low levels of alanine to serine sequence variants were identified in an IgG4 monoclonal antibody by ultra/high performance liquid chromatography and tandem mass spectrometry. The levels of the identified sequence variants A183S and A152S, both in the light chain, have been determined to be 7.8-9.9% and 0.5-0.6%, by extracted ion currents of the tryptic peptides L16 and L14, respectively. The A183S variant was confirmed through tryptic map spiking experiments using synthetic peptide, SDYEK, which incorporated Ser at the position of native Ala in the tryptic peptide L16. Both mutations were also observed by endoproteinase Asp-N peptide mapping. The variant level of A183S was also quantified by LC-UV with detection at 280nm and fluorescence detection of tyrosine residues on the tryptic peptides. The results from LC-MS, UV, and fluorescence detection are in close agreement with each other. The levels of the sequence variants are comparable among the antibody samples manufactured at different scales as well as locations, indicating that the variants' levels are not affected by manufacture scale or locations. DNA sequencing of the master cell bank revealed the presence of mixed bases at position 183 encoding both wild and mutated populations, whereas bases encoding the minor sequence variant at position 152 were not detected. The root cause for A152S mutation is not yet clearly understood at this moment. Copyright © 2012 Elsevier B.V. All rights reserved.

  13. Phonological variant recognition: representations and rules.

    PubMed

    Pinnow, Eleni; Connine, Cynthia M

    2014-03-01

    The current research explores the role of lexical representations and processing in the recognition of phonological variants. Two alternative approaches for variant recognition are considered: a representational approach that posits frequency-graded lexical representations for variant forms and inferential processes that mediate between the spoken variant and the lexical representation. In a lexical decision task (Experiment 1) and in a phoneme identification task (Experiment 2) using real words, low-frequency variants, but not high-frequency variants, show improved recognition rates following additional experience with the variants. This knowledge generalized to novel variant forms. Experiment 3 replicated these results using an artificial lexicon and showed that recognition of low-frequency variants was influenced by similarity to a high-frequency variant form. Similarity to a high-frequency variant alone, however, was insufficient to explain recognition of the infrequent variants (Experiments 4 and 5). The results support a hybrid account of variant recognition that relies on both multiple frequency-graded representations and inference processes.

  14. A Bioinformatics Approach to the Identification of Variants Associated with Type 1 and Type 2 Diabetes Mellitus that Reside in Functionally Validated miRNAs Binding Sites.

    PubMed

    Ghaedi, Hamid; Bastami, Milad; Jahani, Mohammad Mehdi; Alipoor, Behnam; Tabasinezhad, Maryam; Ghaderi, Omar; Nariman-Saleh-Fam, Ziba; Mirfakhraie, Reza; Movafagh, Abolfazl; Omrani, Mir Davood; Masotti, Andrea

    2016-06-01

    The present work is aimed at finding variants associated with Type 1 and Type 2 diabetes mellitus (DM) that reside in functionally validated miRNAs binding sites and that can have a functional role in determining diabetes and related pathologies. Using bioinformatics analyses we obtained a database of validated polymorphic miRNA binding sites which has been intersected with genes related to DM or to variants associated and/or in linkage disequilibrium (LD) with it and is reported in genome-wide association studies (GWAS). The workflow we followed allowed us to find variants associated with DM that also reside in functional miRNA binding sites. These data have been demonstrated to have a functional role by impairing the functions of genes implicated in biological processes linked to DM. In conclusion, our work emphasized the importance of SNPs located in miRNA binding sites. The results discussed in this work may constitute the basis of further works aimed at finding functional candidates and variants affecting protein structure and function, transcription factor binding sites, and non-coding epigenetic variants, contributing to widen the knowledge about the pathogenesis of this important disease.

  15. Identification of novel splice variants of the voltage- and Ca²⁺-dependent K⁺-channel SLO-1 of Trichuris muris.

    PubMed

    Yilmaz, Esra; Kulke, Daniel; von Samson-Himmelstjerna, Georg; Krücken, Jürgen

    2015-01-01

    The anthelmintic cyclooctadepsipeptide emodepside is effective against nematodes showing resistance against established drug classes. Emodepside exerts its nematicidal effects mainly through its validated target, the tetrameric voltage- and calcium-activated potassium channel SLO-1. Two slo-1 genes were described in Trichuris muris. Alternative splicing is known to alter SLO-1 properties. Here, 16 T. muris splice variants for slo-1.1 and three variants for slo-1.2 were identified in addition to previously described variants. Splice variants caused by intron retentions and/or exon exclusions encode varyingly truncated subunits. Depending on the subunit composition, channels might have altered physiological and pharmacological properties including different modulation by calcium and/or voltage or reduced emodepside susceptibility which might lead to emodepside resistance as observed in Caenorhabditis elegans expressing only similarly truncated Slo-1. The comprehensive characterisation of splice variants is a prerequisite for functional analysis and confirmed conservation of remarkable differences found between both slo-1 paralogs in Trichuris suis. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Identification of Male- and Female-Specific Olfaction Genes in Antennae of the Oriental Fruit Fly (Bactrocera dorsalis)

    PubMed Central

    Liu, Zhao; Smagghe, Guy; Lei, Zhongren; Wang, Jin-Jun

    2016-01-01

    The oriental fruit fly (Bactrocera dorsalis) is a species of tephritid fruit fly, endemic to Southeast Asia but also introduced to many regions of the US, and it is one of the major pest species with a broad host range of cultivated and wild fruits. Although males of B. dorsalis respond strongly to methyl eugenol and this is used for monitoring and estimating populations, the molecular mechanism of the oriental fruit fly olfaction has not been elucidated yet. Therefore, in this project, using next generation sequencing technologies, we sequenced the transcriptome of the antennae of male and female adults of B. dorsalis. We identified a total of 20 candidate odorant binding proteins (OBPs), 5 candidate chemosensory proteins (CSPs), 35 candidate odorant receptors (ORs), 12 candidate ionotropic receptors (IRs) and 4 candidate sensory neuron membrane proteins (SNMPs). The sex-specific expression of these genes was determined and a subset of 9 OR genes was further characterized by qPCR with male and female antenna, head, thorax, abdomen, leg and wing samples. In the male antennae, 595 genes showed a higher expression, while 128 genes demonstrated a higher expression in the female antennae. Interestingly, 2 ORs (BdorOR13 and BdorOR14) were highly and specifically expressed in the antennae of males, and 4 ORs (BdorOR13, BdorOR16, BdorOR18 and BdorOR35) clustered with DmOR677, suggesting pheromone reception. We believe this study with these antennae-enriched OBPs, CSPs, ORs, IRs and SNMPs can play an important role in the detection of pheromones and general odorants, and so in turn our data improve our current understanding of insect olfaction at the molecular level and provide important information for disrupting the behavior of the oriental fruit fly using chemical communication methods. PMID:26845547

  17. Identification of Male- and Female-Specific Olfaction Genes in Antennae of the Oriental Fruit Fly (Bactrocera dorsalis).

    PubMed

    Liu, Zhao; Smagghe, Guy; Lei, Zhongren; Wang, Jin-Jun

    2016-01-01

    The oriental fruit fly (Bactrocera dorsalis) is a species of tephritid fruit fly, endemic to Southeast Asia but also introduced to many regions of the US, and it is one of the major pest species with a broad host range of cultivated and wild fruits. Although males of B. dorsalis respond strongly to methyl eugenol and this is used for monitoring and estimating populations, the molecular mechanism of the oriental fruit fly olfaction has not been elucidated yet. Therefore, in this project, using next generation sequencing technologies, we sequenced the transcriptome of the antennae of male and female adults of B. dorsalis. We identified a total of 20 candidate odorant binding proteins (OBPs), 5 candidate chemosensory proteins (CSPs), 35 candidate odorant receptors (ORs), 12 candidate ionotropic receptors (IRs) and 4 candidate sensory neuron membrane proteins (SNMPs). The sex-specific expression of these genes was determined and a subset of 9 OR genes was further characterized by qPCR with male and female antenna, head, thorax, abdomen, leg and wing samples. In the male antennae, 595 genes showed a higher expression, while 128 genes demonstrated a higher expression in the female antennae. Interestingly, 2 ORs (BdorOR13 and BdorOR14) were highly and specifically expressed in the antennae of males, and 4 ORs (BdorOR13, BdorOR16, BdorOR18 and BdorOR35) clustered with DmOR677, suggesting pheromone reception. We believe this study with these antennae-enriched OBPs, CSPs, ORs, IRs and SNMPs can play an important role in the detection of pheromones and general odorants, and so in turn our data improve our current understanding of insect olfaction at the molecular level and provide important information for disrupting the behavior of the oriental fruit fly using chemical communication methods.

  18. An evaluation of the SPIFE 3000 semi-automated gel electrophoresis system for the identification of hemoglobin variants and comparison of relative electrophoretic mobilities with manual gel electrophoresis methods.

    PubMed

    Hoyer, J D; Markley, K M; Savedra, M E; Kubik, K S; Scheidt, R M

    2010-06-01

    Laboratory identification of hemoglobin (Hb) variants can involve multiple techniques. The use of semi-automated instruments that perform gel electrophoresis and staining, such as the SPIFE 3000 electrophoresis system, can greatly reduce the labor required for these commonly used techniques. We performed a comparison of the method involved in SPIFE 3000 system with those of manual gel electrophoresis. A total of 22 540 samples were analyzed using the SPIFE 3000, and compared with mobilities on cellulose acetate and citrate agar gels using standard manual methods. The results were compared using relative electrophoretic mobilities (REM). Of the 191 Hb variants identified, only 13 had REM that differed from manual electrophoresis when analyzed using the SPIFE 3000 system. One variant (Hb O-Indonesia) showed different mobility on both acid and alkaline gels, two (Hb E, Hb Sunshine Seth) on alkaline gel only, and 10 (Hbs N-Baltimore, N-Seattle, O-Arab, Shelby, Summer Hill, Tak, Hasharon, M-Iwate, Q-Iran, and Setif) on acid gels only. The SPIFE 3000 semi-automated electrophoresis system produces similar results when compared with those of standard manual electrophoresis methods.

  19. Identification of Rhodospirillum rubrum GlnB Variants That Are Altered in Their Ability To Interact with Different Targets in Response to Nitrogen Status Signals

    PubMed Central

    Zhu, Yu; Conrad, Mary C.; Zhang, Yaoping; Roberts, Gary P.

    2006-01-01

    In Rhodospirillum rubrum, NifA, the transcriptional activator for the nif genes, is posttranslationally activated only by the uridylylated form of GlnB, one of three PII homologs in the organism. We have used the yeast two-hybrid system to detect variants of GlnB that interact better with NifA than does wild-type GlnB. When examined for physiological effects in R. rubrum, these GlnB* variants activated NifA in the presence of NH4+, which normally blocks NifA activation completely, and in the absence of GlnD, whose uridylylation of GlnB is also normally essential for NifA activation. When these variants were tested in the two-hybrid system for their interaction with NtrB, a receptor that should interact with the nonuridylylated form of GlnB, they were uniformly weaker than wild-type GlnB in that interaction. When expressed in R. rubrum either as single-copy integrants or on multiple-copy plasmids, these variants were also dramatically altered in terms of their ability to regulate several other receptors involved in nitrogen metabolism, including GlnE, NtrB/NtrC, and DRAT (dinitrogenase reductase ADP-ribosyl transferase)-DRAG (dinitrogenase reductase-activating glycohydrolase). The consistent pattern throughout is that these GlnB variants partially mimic the uridylylated form of wild-type GlnB, even under nitrogen-excess conditions and in strains lacking GlnD. The results suggest that the role of uridylylation of GlnB is primarily to shift the equilibrium of GlnB from a “nitrogen-sufficient” form to a “nitrogen-deficient” form, each of which interacts with different but overlapping receptor proteins in the cell. These GlnB variants apparently shift that equilibrium through direct structural changes. PMID:16484197

  20. Identification of a novel vga(E) gene variant that confers resistance to pleuromutilins, lincosamides and streptogramin A antibiotics in staphylococci of porcine origin.

    PubMed

    Li, Jun; Li, Beibei; Wendlandt, Sarah; Schwarz, Stefan; Wang, Yang; Wu, Congming; Ma, Zhiyong; Shen, Jianzhong

    2014-04-01

    To investigate the genetic basis of pleuromutilin resistance in coagulase-negative staphylococci of porcine origin that do not carry known pleuromutilin resistance genes and to determine the localization and genetic environment of the identified resistance gene. Plasmid DNA of two pleuromutilin-resistant Staphylococcus cohnii and Staphylococcus simulans isolates was transformed into Staphylococcus aureus RN4220. The identified resistance plasmids were sequenced completely. The candidate gene for pleuromutilin resistance was cloned into shuttle vector pAM401. S. aureus RN4220 transformants carrying this recombinant shuttle vector were tested for their MICs. S. cohnii isolate SA-7 and S. simulans isolate SSI1 carried the same plasmid of 5584 bp, designated pSA-7. A variant of the vga(E) gene was detected, which encodes a 524 amino acid ATP-binding cassette protein. The variant gene shared 85.7% nucleotide sequence identity and the variant protein 85.3% amino acid sequence identity with the original vga(E) gene and Vga(E) protein, respectively. The Vga(E) variant conferred cross-resistance to pleuromutilins, lincosamides and streptogramin A antibiotics. Plasmid pSA-7 showed an organization similar to that of the apmA-carrying plasmid pKKS49 from methicillin-resistant S. aureus and the dfrK-carrying plasmid pKKS966 from Staphylococcus hyicus. Sequence comparisons suggested that recombination events may have played a role in the acquisition of this vga(E) variant. A novel vga(E) gene variant was identified, which was located on a small plasmid and was not associated with the transposon Tn6133 [in contrast to the original vga(E) gene]. The plasmid location may enable its further dissemination to other staphylococci and possibly also to other bacteria.

  1. Mutation screening of IRF6 among families with non-syndromic oral clefts and identification of two novel variants: review of the literature.

    PubMed

    Salahshourifar, Iman; Wan Sulaiman, Wan Azman; Halim, Ahmad Sukari; Zilfalil, Bin Alwi

    2012-06-01

    Non-syndromic oral clefts share the main clinical features of Van der Woude Syndrome (VWS), with the exception of the lower lip pit. Thus, about 15% of VWS cases are indistinguishable from cases with non-syndromic oral clefts. IRF6 mutations are the major cause of VWS; however, variants in this gene show strong association with non-syndromic oral clefts, with a higher increased risk among cases with cleft lip only (CLO). A total of 39 individuals, including 16 patients with CLO and 23 patients with a family history of cleft, were examined for IRF6 mutations in the present study. Seven variants, including five known (c.-75-4 A>; G, c.-73T>; C, c.459G>; T 5, c.820G>; A, and c.1060 + 37C>; T) and two novel (c.-75-23G>; C and c.1380G>; T), were found. Both novel variants were inherited from non-affected parents and we did not find also in the 120 control chromosomes. In silico analysis revealed that both c.1380G>; T and c.-75-23G>; C variants may disrupts a putative exonic splicing enhancer and intronic splicing binding site for SC35, respectively. Taken together, the presence of deleterious IRF6 variants in patients with non-syndromic oral clefts could be most likely an evidence for VWS. While, IRF6 variants could, at best, contribute to clefting as part of a complex inheritance pattern, with both additional genes and environmental factors having a role. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  2. Identification of new TSGA10 transcript variants in human testis with conserved regulatory RNA elements in 5'untranslated region and distinct expression in breast cancer.

    PubMed

    Salehipour, Pouya; Nematzadeh, Mahsa; Mobasheri, Maryam Beigom; Afsharpad, Mandana; Mansouri, Kamran; Modarressi, Mohammad Hossein

    2017-09-01

    Testis specific gene antigen 10 (TSGA10) is a cancer testis antigen involved in the process of spermatogenesis. TSGA10 could also play an important role in the inhibition of angiogenesis by preventing nuclear localization of HIF-1α. Although it has been shown that TSGA10 messenger RNA (mRNA) is mainly expressed in testis and some tumors, the transcription pattern and regulatory mechanisms of this gene remain largely unknown. Here, we report that human TSGA10 comprises at least 22 exons and generates four different transcript variants. It was identified that using two distinct promoters and splicing of exons 4 and 7 produced these transcript variants, which have the same coding sequence, but the sequence of 5'untanslated region (5'UTR) is different between them. This is significant because conserved regulatory RNA elements like upstream open reading frame (uORF) and putative internal ribosome entry site (IRES) were found in this region which have different combinations in each transcript variant and it may influence translational efficiency of them in normal or unusual environmental conditions like hypoxia. To indicate the transcription pattern of TSGA10 in breast cancer, expression of identified transcript variants was analyzed in 62 breast cancer samples. We found that TSGA10 tends to express variants with shorter 5'UTR and fewer uORF elements in breast cancer tissues. Our study demonstrates for the first time the expression of different TSGA10 transcript variants in testis and breast cancer tissues and provides a first clue to a role of TSGA10 5'UTR in regulation of translation in unusual environmental conditions like hypoxia. Copyright © 2017. Published by Elsevier B.V.

  3. Identification of human herpesvirus 6 variants A and B by primer-specific real-time PCR may help to revisit their respective role in pathology.

    PubMed

    Boutolleau, David; Duros, Caroline; Bonnafous, Pascale; Caïola, Delphine; Karras, Alexandre; Castro, Nathalie De; Ouachée, Marie; Narcy, Philippe; Gueudin, Marie; Agut, Henri; Gautheret-Dejean, Agnès

    2006-03-01

    Human herpesvirus 6 (HHV-6) isolates are classified into two variants, termed HHV-6A and HHV-6B, on the basis of distinct genetic, antigenic and biological characteristics, but the specific pathogenicity of each variant remains poorly understood. To design a rapid, sensitive and specific real-time variant-specific PCR (VS-PCR) method to differentiate both variants in biological specimens. The VS-PCR was adapted from a real-time PCR assay, based on TaqMan technology, previously developed for the genome quantitation of both HHV-6 variants [Gautheret-Dejean A, Manichanh C, Thien-Ah-Koon F, Fillet AM, Mangeney N, Vidaud M, et al. Development of a real-time polymerase chain reaction assay for the diagnosis of human herpesvirus-6 infection and application to bone marrow transplant patients. J Virol Meth 2002;100:27-35], a consensual reverse primer (Taq2) being changed into two variant-specific primers named H6A and H6B. This method was applied to a large set of biological specimens obtained in different pathological contexts. The sensitivity threshold was about 10 copies/well for HHV-6A-specific PCR (PCR-A) and 1 copy/well for HHV-6B-specific PCR (PCR-B). Both assays showed a linear dynamic range from 10 to 100,000 copies of HHV-6 DNA. Regarding the specificity and the capacity of discrimination of each assay, one variant could be detected and identified in the presence of more than 1000 times higher concentrations of the other variant in virus mixtures. The comparison of the results obtained with this VS-PCR with those previously obtained with a classic PCR method allowed us to validate our new technique on a wide panel of biological samples, including numerous patients with severe HHV-6-related symptoms. The high prevalence of HHV-6B was confirmed in healthy individuals and immunocompromised patients. HHV-6A was identified in distinct samples from several patients exhibiting neurological disorders. We developed a new VS-PCR assay, able to differentiate HHV-6A and HHV-6

  4. Identification of the Functional Variant(s) that Explain the Low-Density Lipoprotein Receptor (LDLR) GWAS SNP rs6511720 Association with Lower LDL-C and Risk of CHD

    PubMed Central

    Palmen, Jutta; Kalea, Anastasia Z.

    2016-01-01

    Background The Low-Density Lipoprotein Receptor (LDLR) SNP rs6511720 (G>T), located in intron-1 of the gene, has been identified in genome-wide association studies (GWAS) as being associated with lower plasma levels of LDL-C and a lower risk of coronary heart disease (CHD). Whether or not rs6511720 is itself functional or a marker for a functional variant elsewhere in the gene is not known. Methods The association of LDLR SNP rs6511720 with incidence of CHD and levels of LDL-C was determined by reference to CARDIoGRAM, C4D and Global lipids genetics consortium (GLGC) data. SNP annotation databases were used to identify possible SNP function and prioritization. Luciferase reporter assays in the liver cell line Huh7 were used to measure the effect of variant genotype on gene expression. Electrophoretic Mobility Shift Assays (EMSAs) were used to identify the Transcription Factors (TFs) involved in gene expression regulation. Results The phenotype-genotype analysis showed that the rs6511720 minor allele is associated with lower level of LDL-C [beta = -0.2209, p = 3.85 x10-262], and lower risk of CHD [log (OR) = 0.1155, p = 1.04 x10-7]. Rs6511720 is in complete linkage. Rs6511720 is in complete linkage disequilibrium (LD) with three intron-1 SNPs (rs141787760, rs60173709, rs57217136). Luciferase reporter assays in Huh7 cells showed that the rare alleles of both rs6511720 and rs57217136 caused a significant increase in LDLR expression compared to the common alleles (+29% and +24%, respectively). Multiplex Competitor-EMSAs (MC-EMSA) identified that the transcription factor serum response element (SRE) binds to rs6511720, while retinoic acid receptor (RAR) and signal transducer and activator of transcription 1 (STAT1) bind to rs57217136. Conclusion Both LDLR rs6511720 and rs57217136 are functional variants. Both these minor alleles create enhancer-binding protein sites for TFs and may contribute to increased LDLR expression, which is consequently associated with reduced

  5. Identification and characterization of a new human type 9 cGMP-specific phosphodiesterase splice variant (PDE9A5). Differential tissue distribution and subcellular localization of PDE9A variants.

    PubMed

    Wang, Peng; Wu, Ping; Egan, Robert W; Billah, M Motasim

    2003-09-18

    Previously, four splice variants of human cGMP-specific phosphodiesterase (PDE) 9A (PDEs 9A1, 9A2, 9A3 and 9A4) have been identified. In this study, we have cloned a cDNA representing a new human PDE9A variant (PDE9A5). PDE9A5 encodes a protein of 492 amino acids, smaller than PDEs 9A1 and 9A2 but larger than PDEs 9A3 and 9A4. The exon structure of PDE9A5 is different from those of PDEs 9A1, 9A2, 9A3 and 9A4 in that, of the 20 exons of PDE9A gene, it lacks exons 2 and 5. PDE9A5 has been characterized in comparison with PDE9A1, the longest PDE9A variant. PDEs 9A5 and 9A1 have similar enzymatic properties. They both have a high affinity for cGMP with similar Km values (0.39 and 0.25 microM, respectively), although they have slightly different Vmax values (2.55 and 0.96 micromol/min/mg, respectively). They exhibit very similar divalent metal ion dependency and inhibitor sensitivity. Real-time quantitative PCR analysis shows that PDEs 9A5 and 9A1 exhibit differential tissue distribution. They are highly expressed in immune tissues (spleen, lymph node and thymus) and are more abundant in T cells than in B cells, neutrophils and monocytes. When transiently expressed in HEK293 cells, PDEs 9A5 and 9A1 proteins exhibit differential subcellular localization. PDE9A5 localizes exclusively in the cytoplasm, whereas PDE9A1 localizes in the nucleus only. The nuclear localization of PDE9A1 is dependent on a unique pat7 motif. By Western blot analysis, native PDE9A1 is detectable in the nucleus but not in the cytoplasm of T cells. Thus, to our knowledge, PDE9A1 is the only PDE isoform found to localize exclusively in the nucleus. We speculate that the physiological role of the PDE9A diversity may be imparting cGMP-metabolizing ability to specific cellular compartments in appropriate tissues.

  6. Identification of Variants of Hepatitis C Virus (HCV) Entry Factors in Patients Highly Exposed to HCV but Remaining Uninfected: An ANRS Case-Control Study.

    PubMed

    Fouquet, Baptiste; Ghosn, Jade; Quertainmont, Yann; Salmon, Dominique; Rioux, Christophe; Duvivier, Claudine; Delfraissy, Jean-François; Misrahi, Micheline

    2015-01-01

    Hepatitis C virus (HCV) causes persistent infection in 75% of cases and is a major public health problem worldwide. More than 92% of intravenous drug users (IDU) infected by human immunodeficiency virus type 1 (HIV-1) are seropositive for HCV, and it is conceivable that some HIV-1-infected IDU who remain uninfected by HCV may be genetically resistant.Here we conducted a case-control study to identify mutations in HCV entry coreceptors in HIV-infected IDU who remained uninfected by HCV. We recruited 138 patients, comprising 22 HIV+ HCV- case IDU and 116 HIV+ HCV+ control IDU. We focused on coreceptors in which point mutations are known to abolish HCV infectivity in vitro. Our previous study of the Claudin-1 gene revealed no specific variants in the same case population. Here we performed direct genomic sequencing of the Claudin-6, Claudin-9, Occludin and Scavenger receptor-B1 (SCARB1) gene coding regions. Most HIV+ HCV- IDU had no mutations in HCV coreceptors. However, two HIV+ HCV- patients harbored a total of four specific mutations/variants of HCV entry factors that were not found in the HIV+ HCV+ controls. One case patient harbored heterozygous variants of both Claudin-6 and Occludin, and the other case patient harbored two heterozygous variants of SCARB1. This suggests that HCV resistance might involve complex genetic events and factors other than coreceptors, a situation similar to that reported for HIV-1 resistance.

  7. Identification of non-coding genetic variants in samples from hypoxemic respiratory disease patients that affect the transcriptional response to hypoxia

    PubMed Central

    Roche, Olga; Deguiz, María Laura; Tiana, María; Galiana-Ribote, Clara; Martinez-Alcazar, Daniel; Rey-Serra, Carlos; Ranz-Ribeiro, Beatriz; Casitas, Raquel; Galera, Raúl; Fernández-Navarro, Isabel; Sánchez-Cuéllar, Silvia; Bernard, Virginie; Ancochea, Julio; Wasserman, Wyeth W.; García-Rio, Francisco; Jimenez, Benilde; del Peso, Luis

    2016-01-01

    A wide range of diseases course with an unbalance between the consumption of oxygen by tissues and its supply. This situation triggers a transcriptional response, mediated by the hypoxia inducible factors (HIFs), that aims to restore oxygen homeostasis. Little is known about the inter-individual variation in this response and its role in the progression of disease. Herein, we sought to identify common genetic variants mapping to hypoxia response elements (HREs) and characterize their effect on transcription. To this end, we constructed a list of genome-wide HIF-binding regions from publicly available experimental datasets and studied the genetic variability in these regions by targeted re-sequencing of genomic samples from 96 chronic obstructive pulmonary disease and 144 obstructive sleep apnea patients. This study identified 14 frequent variants disrupting potential HREs. The analysis of the genomic regions containing these variants by means of reporter assays revealed that variants rs1009329, rs6593210 and rs150921338 impaired the transcriptional response to hypoxia. Finally, using genome editing we confirmed the functional role of rs6593210 in the transcriptional regulation of EGFR. In summary, we found that inter-individual variability in non-coding regions affect the response to hypoxia and could potentially impact on the progression of pulmonary diseases. PMID:27625398

  8. Identification and functional analysis of variants of a cancer/testis antigen LEMD1 in colorectal cancer stem-like cells.

    PubMed

    Takeda, Rui; Hirohashi, Yoshihiko; Shen, Min; Wang, Liming; Ogawa, Tadashi; Murai, Aiko; Yamamoto, Eri; Kubo, Terufumi; Nakatsugawa, Munehide; Kanaseki, Takayuki; Tsukahara, Tomohide; Nishidate, Toshihiko; Okita, Kenji; Kutomi, Goro; Sato, Noriyuki; Takemasa, Ichiro; Torigoe, Toshihiko

    2017-04-08

    Colorectal cancer (CRC) is one of the most common malignancy, and the prognosis is not still satisfactory due to treatment resistance, recurrence and distant metastasis. Cancer stem cells (CSCs)/cancer-initiating cells (CICs) is endowed with higher tumor-initiating ability, self-renewal ability and differentiation ability, and CSCs/CICs are resistant to treatments. Thus, CSCs/CICs are thought to be responsible for recurrence and distant metastasis, and eradication of CSCs/CICs is essential to cure CRCs. However, the molecular mechanisms of CSCs/CICs are remain unknown, and we aimed to elucidate molecular aspects of CR-CSCs/CICs in this study. We screened the transcriptome data of primary human CR-CSCs/CICs that we previously established, and found that LEM domain containing 1 (LEMD1) is preferentially expressed in CR-CSCs/CICs. LEMD1 belongs to cancer-testis (CT) antigen, and has five transcript variants (variant 1 [V1] - variant 5 [V5]). We found that LEMD1 V1, V2 and V3 is expressed in testis and CR-CSCs/CICs, whereas LEMD1 V4 and V5 is ubiquitously expressed. LEMD1 gene knockdown experiments using siRNAs and gene overexpression experiments revealed that LEMD1 has a role in the maintenance of CR-CSCs/CICs. These observations indicate that CR-CSC/CIC-specific LEMD1 variants are reasonable target of CR-CSC/CIC-targeted therapy. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Identification of STAC3 variants in non-Native American families with overlapping features of Carey-Fineman-Ziter syndrome and Moebius syndrome.

    PubMed

    Telegrafi, Aida; Webb, Bryn D; Robbins, Sarah M; Speck-Martins, Carlos E; FitzPatrick, David; Fleming, Leah; Redett, Richard; Dufke, Andreas; Houge, Gunnar; van Harssel, Jeske J T; Verloes, Alain; Robles, Angela; Manoli, Irini; Engle, Elizabeth C; Jabs, Ethylin W; Valle, David; Carey, John; Hoover-Fong, Julie E; Sobreira, Nara L M

    2017-10-01

    Horstick et al. (2013) previously reported a homozygous p.Trp284Ser variant in STAC3 as the cause of Native American myopathy (NAM) in 5 Lumbee Native American families with congenital hypotonia and weakness, cleft palate, short stature, ptosis, kyphoscoliosis, talipes deformities, and susceptibility to malignant hyperthermia (MH). Here we present two non-Native American families, who were found to have STAC3 pathogenic variants. The first proband and her affected older sister are from a consanguineous Qatari family with a suspected clinical diagnosis of Carey-Fineman-Ziter syndrome (CFZS) based on features of hypotonia, myopathic facies with generalized weakness, ptosis, normal extraocular movements, cleft palate, growth delay, and kyphoscoliosis. We identified the homozygous c.851G>C;p.Trp284Ser variant in STAC3 in both sisters. The second proband and his affected sister are from a non-consanguineous, Puerto Rican family who was evaluated for a possible diagnosis of Moebius syndrome (MBS). His features included facial and generalized weakness, minimal limitation of horizontal gaze, cleft palate, and hypotonia, and he has a history of MH. The siblings were identified to be compound heterozygous for STAC3 variants c.851G>C;p.Trp284Ser and c.763_766delCTCT;p.Leu255IlefsX58. Given the phenotypic overlap of individuals with CFZS, MBS, and NAM, we screened STAC3 in 12 individuals diagnosed with CFZS and in 50 individuals diagnosed with MBS or a congenital facial weakness disorder. We did not identify any rare coding variants in STAC3. NAM should be considered in patients presenting with facial and generalized weakness, normal or mildly abnormal extraocular movement, hypotonia, cleft palate, and scoliosis, particularly if there is a history of MH. © 2017 Wiley Periodicals, Inc.

  10. Volatile organic compounds in the Pearl River Delta: Identification of source regions and recommendations for emission-oriented monitoring strategies

    NASA Astrophysics Data System (ADS)

    Yuan, Zibing; Zhong, Liuju; Lau, Alexis Kai Hon; Yu, Jian Zhen; Louie, Peter K. K.

    2013-09-01

    For the purpose of systematically characterizing the ambient volatile organic compound (VOC) levels and their emission sources in the Pearl River Delta (PRD) of China, a grid study with VOC samples simultaneously taken at 84 sites over the PRD was conducted in summer and winter of 2008 and 2009. Positive Matrix Factorization (PMF) model was applied to identify the major VOC contributing sources and their temporal and spatial variations. Nine sources were identified, with gasoline exhaust, industrial emission and LPG leakage & propellant emission the top three significant sources. They accounted for 23%, 16% and 13% of the ambient VOC levels, respectively. Control measures should be therefore targeted on mitigating the VOC emissions from the traffic-related and industrial-related sources. The total VOC level did not show strong increase from 5 a.m. to 10 a.m. during all the four sampling campaigns, which may result from stronger wind and higher mixing height at 10 a.m. Three hotspot areas with significant VOC contributions were identified by source apportionment analysis: (1) the Pearl River Estuary; (2) an area from Central Dongguan to North Shenzhen; and (3) the Zhuhai-Zhongshan-Jiangmen area. For better characterizing the roles of VOC and NOx in producing the secondary pollutants and to identify specific sources emitting excessive concentrations of precursors, the emission-oriented Photochemical Assessment Monitoring Station (PAMS) network is recommended to be established in the PRD. Three PAMS networks are suggested in correspondence to the three identified hotspot areas.

  11. Identification of differentially expressed genes in hepatopancreas of oriental river prawn, Macrobrachium nipponense exposed to environmental hypoxia.

    PubMed

    Sun, Shengming; Xuan, Fujun; Ge, Xianping; Fu, Hongtuo; Zhu, Jian; Zhang, Shiyong

    2013-10-25

    Hypoxia represents a major physiological challenge for prawn culture, and the hepatopancreas plays an important role in these processes. Here, we applied high-throughput sequencing technology to detect the gene expression profile of the hepatopancreas in M. nipponense in response to hypoxia for 3h and hypoxia for 24h. Gene expression profiling identified 1925 genes that were significantly up- or down-regulated by dissolved oxygen availability. Functional categorization of the differentially expressed genes revealed that oxygen transport, electron transport chain, reactive oxygen species generation/scavenging, and immune response were the differentially regulated processes occurring during environmental hypoxia. Finally, quantitative real-time polymerase chain reaction using six genes independently verified the tag-mapped results. Immunohistochemistry analysis revealed, for the first time, hemocyanin protein expression as significant hypoxia-specific signatures in prawns, which opens the way for in depth molecular studies of hypoxia exposure. The analysis of changes in hepatic gene expression in oriental river prawn provides a preliminary basis for a better understanding of the molecular response to hypoxia exposures. Crown Copyright © 2013. All rights reserved.

  12. An automated high-throughput screening method for the identification of high-yield, soluble protein variants using cell-free expression and systematic truncation.

    PubMed

    Bursey, Evan H; Kim, Chang-Yub; Yu, Minmin; Terwilliger, Thomas C; Hung, Li-Wei

    2006-12-01

    A highly automated method for rapidly identifying soluble protein variants with good expression yields has been developed. This method is based on a commercially available in vitro protein expression system. It consists of two polymerase chain reactions (PCR) followed by in vitro protein expression and protein quantification by dot blot. The PCR protocols have been improved and optimized to allow automation using commercial fluid handling devices. A PCR primer design program has also been implemented to streamline protein variant design. This automated protocol is highly reliable and has tremendously improved the throughput of expression screening as compared to conventional cell-based methods and manual in vitro methods. We have applied this method to 32 problematic targets from the TB Structural Genomics Consortium. Experimental results of these studies are reported.

  13. Identification of a new functional splice variant of the enzyme methionine sulphoxide reductase A (MSRA) expressed in rat vascular smooth muscle cells.

    PubMed

    Haenold, Ronny; Wassef, Ramez; Hansel, Alfred; Heinemann, Stefan H; Hoshi, Toshinori

    2007-11-01

    Reactive oxygen species contribute to ageing of the vascular system and development of cardiovascular disease. Methionine-S-sulphoxide, an oxidized form of methionine, is repaired by the enzyme methionine sulphoxide reductase A (MSRA). The enzyme, targeted to mitochondria or the cytosol by alternative splicing, is vital for oxidative stress resistance. This study was designed to examine the endogenous expression and intracellular localization of MSRA in rat aortic vascular smooth muscle cells (VSMCs). We detected robust MSRA immunoreactivity exclusively in mitochondria. Sequence analysis of msrA transcripts revealed the presence of a novel mitochondrial splice variant, msrA2a, in cultured rat VSMCs as well as in aortic tissue preparations. The enzymatic activity of a recombinant MSRA2a protein was confirmed by the reduction of methionine sulphoxide in a model substrate peptide. We conclude that multiple MSRA variants participate in the repair of oxidized proteins in VSMC mitochondria, but that other protective mechanisms may exist in the cytoplasmic compartment.

  14. Identification of a Latin American-specific BabA adhesin variant through whole genome sequencing of Helicobacter pylori patient isolates from Nicaragua

    SciTech Connect

    Thorell, Kaisa; Hosseini, Shaghayegh; Palacios Gonzales, Reyna Victoria Palacios; Chaotham, Chatchai; Graham, David Y.; Paszat, Lawrence; Rabeneck, Linda; Lundin, Samuel B.; Nookaew, Intawat; Sjoling, Asa

    2016-02-29

    In this study, Helicobacter pylori (H. pylori) is one of the most common bacterial infections in humans and this infection can lead to gastric ulcers and gastric cancer. H. pylori is one of the most genetically variable human pathogens and the ability of the bacterium to bind to the host epithelium as well as the presence of different virulence factors and genetic variants within these genes have been associated with disease severity. Nicaragua has particularly high gastric cancer incidence and we therefore studied Nicaraguan clinical H. pylori isolates for factors that could contribute to cancer risk. The complete genomes of fifty-two Nicaraguan H. pylorii isolates were sequenced and assembled de novo, and phylogenetic and virulence factor analyses were performed. The Nicaraguan isolates showed phylogenetic relationship with West African isolates in whole-genome sequence comparisons and with Western and urban South-and Central American isolates using MLSA (Multi-locus sequence analysis). A majority, 77 % of the isolates carried the cancer-associated virulence gene cagA and also the s1/i1/m1 vacuolating cytotoxin, vacA allele combination, which is linked to increased severity of disease. Specifically, we also found that Nicaraguan isolates have a blood group-binding adhesin (BabA) variant highly similar to previously reported BabA sequences from Latin America, including from isolates belonging to other phylogenetic groups. These BabA sequences were found to be under positive selection at several amino acid positions that differed from the global collection of isolates. In conclusion, the discovery of a Latin American BabA variant, independent of overall phylogenetic background, suggests hitherto unknown host or environmental factors within the Latin American population giving H. pylori isolates carrying this adhesin variant a selective advantage, which could affect pathogenesis and risk for sequelae through specific adherence

  15. Identification of a Latin American-specific BabA adhesin variant through whole genome sequencing of Helicobacter pylori patient isolates from Nicaragua

    DOE PAGES

    Thorell, Kaisa; Hosseini, Shaghayegh; Palacios Gonzales, Reyna Victoria Palacios; ...

    2016-02-29

    In this study, Helicobacter pylori (H. pylori) is one of the most common bacterial infections in humans and this infection can lead to gastric ulcers and gastric cancer. H. pylori is one of the most genetically variable human pathogens and the ability of the bacterium to bind to the host epithelium as well as the presence of different virulence factors and genetic variants within these genes have been associated with disease severity. Nicaragua has particularly high gastric cancer incidence and we therefore studied Nicaraguan clinical H. pylori isolates for factors that could contribute to cancer risk. The complete genomes ofmore » fifty-two Nicaraguan H. pylorii isolates were sequenced and assembled de novo, and phylogenetic and virulence factor analyses were performed. The Nicaraguan isolates showed phylogenetic relationship with West African isolates in whole-genome sequence comparisons and with Western and urban South-and Central American isolates using MLSA (Multi-locus sequence analysis). A majority, 77 % of the isolates carried the cancer-associated virulence gene cagA and also the s1/i1/m1 vacuolating cytotoxin, vacA allele combination, which is linked to increased severity of disease. Specifically, we also found that Nicaraguan isolates have a blood group-binding adhesin (BabA) variant highly similar to previously reported BabA sequences from Latin America, including from isolates belonging to other phylogenetic groups. These BabA sequences were found to be under positive selection at several amino acid positions that differed from the global collection of isolates. In conclusion, the discovery of a Latin American BabA variant, independent of overall phylogenetic background, suggests hitherto unknown host or environmental factors within the Latin American population giving H. pylori isolates carrying this adhesin variant a selective advantage, which could affect pathogenesis and risk for sequelae through specific adherence properties.« less

  16. Identification of Rare Recurrent Copy Number Variants in High-Risk Autism Families and Their Prevalence in a Large ASD Population

    PubMed Central

    Christensen, G. Bryce; Kim, Cecilia; Frackelton, Edward; Thomas, Kelly; da Silva, Renata Pellegrino; Stevens, Jeff; Baird, Lisa; Otterud, Brith; Ho, Karen; Varvil, Tena; Leppert, Tami; Lambert, Christophe G.; Leppert, Mark; Hakonarson, Hakon

    2013-01-01

    Structural variation is thought to play a major etiological role in the development of autism spectrum disorders (ASDs), and numerous studies documenting the relevance of copy number variants (CNVs) in ASD have been published since 2006. To determine if large ASD families harbor high-impact CNVs that may have broader impact in the general ASD population, we used the Affymetrix genome-wide human SNP array 6.0 to identify 153 putative autism-specific CNVs present in 55 individuals with ASD from 9 multiplex ASD pedigrees. To evaluate the actual prevalence of these CNVs as well as 185 CNVs reportedly associated with ASD from published studies many of which are insufficiently powered, we designed a custom Illumina array and used it to interrogate these CNVs in 3,000 ASD cases and 6,000 controls. Additional single nucleotide variants (SNVs) on the array identified 25 CNVs that we did not detect in our family studies at the standard SNP array resolution. After molecular validation, our results demonstrated that 15 CNVs identified in high-risk ASD families also were found in two or more ASD cases with odds ratios greater than 2.0, strengthening their support as ASD risk variants. In addition, of the 25 CNVs identified using SNV probes on our custom array, 9 also had odds ratios greater than 2.0, suggesting that these CNVs also are ASD risk variants. Eighteen of the validated CNVs have not been reported previously in individuals with ASD and three have only been observed once. Finally, we confirmed the association of 31 of 185 published ASD-associated CNVs in our dataset with odds ratios greater than 2.0, suggesting they may be of clinical relevance in the evaluation of children with ASDs. Taken together, these data provide strong support for the existence and application of high-impact CNVs in the clinical genetic evaluation of children with ASD. PMID:23341896

  17. Association of breast cancer risk with genetic variants showing differential allelic expression: Identification of a novel breast cancer susceptibility locus at 4q21.

    PubMed

    Hamdi, Yosr; Soucy, Penny; Adoue, Véronique; Michailidou, Kyriaki; Canisius, Sander; Lemaçon, Audrey; Droit, Arnaud; Andrulis, Irene L; Anton-Culver, Hoda; Arndt, Volker; Baynes, Caroline; Blomqvist, Carl; Bogdanova, Natalia V; Bojesen, Stig E; Bolla, Manjeet K; Bonanni, Bernardo; Borresen-Dale, Anne-Lise; Brand, Judith S; Brauch, Hiltrud; Brenner, Hermann; Broeks, Annegien; Burwinkel, Barbara; Chang-Claude, Jenny; Couch, Fergus J; Cox, Angela; Cross, Simon S; Czene, Kamila; Darabi, Hatef; Dennis, Joe; Devilee, Peter; Dörk, Thilo; Dos-Santos-Silva, Isabel; Eriksson, Mikael; Fasching, Peter A; Figueroa, Jonine; Flyger, Henrik; García-Closas, Montserrat; Giles, Graham G; Goldberg, Mark S; González-Neira, Anna; Grenaker-Alnæs, Grethe; Guénel, Pascal; Haeberle, Lothar; Haiman, Christopher A; Hamann, Ute; Hallberg, Emily; Hooning, Maartje J; Hopper, John L; Jakubowska, Anna; Jones, Michael; Kabisch, Maria; Kataja, Vesa; Lambrechts, Diether; Le Marchand, Loic; Lindblom, Annika; Lubinski, Jan; Mannermaa, Arto; Maranian, Mel; Margolin, Sara; Marme, Frederik; Milne, Roger L; Neuhausen, Susan L; Nevanlinna, Heli; Neven, Patrick; Olswold, Curtis; Peto, Julian; Plaseska-Karanfilska, Dijana; Pylkäs, Katri; Radice, Paolo; Rudolph, Anja; Sawyer, Elinor J; Schmidt, Marjanka K; Shu, Xiao-Ou; Southey, Melissa C; Swerdlow, Anthony; Tollenaar, Rob A E M; Tomlinson, Ian; Torres, Diana; Truong, Thérèse; Vachon, Celine; Van Den Ouweland, Ans M W; Wang, Qin; Winqvist, Robert; Zheng, Wei; Benitez, Javier; Chenevix-Trench, Georgia; Dunning, Alison M; Pharoah, Paul D P; Kristensen, Vessela; Hall, Per; Easton, Douglas F; Pastinen, Tomi; Nord, Silje; Simard, Jacques

    2016-12-06

    There are significant inter-individual differences in the levels of gene expression. Through modulation of gene expression, cis-acting variants represent an important source of phenotypic variation. Consequently, cis-regulatory SNPs associated with differential allelic expression are functional candidates for further investigation as disease-causing variants. To investigate whether common variants associated with differential allelic expression were involved in breast cancer susceptibility, a list of genes was established on the basis of their involvement in cancer related pathways and/or mechanisms. Thereafter, using data from a genome-wide map of allelic expression associated SNPs, 313 genetic variants were selected and their association with breast cancer risk was then evaluated in 46,451 breast cancer cases and 42,599 controls of European ancestry ascertained from 41 studies participating in the Breast Cancer Association Consortium. The associations were evaluated with overall breast cancer risk and with estrogen receptor negative and positive disease. One novel breast cancer susceptibility locus on 4q21 (rs11099601) was identified (OR = 1.05, P = 5.6x10-6). rs11099601 lies in a 135 kb linkage disequilibrium block containing several genes, including, HELQ, encoding the protein HEL308 a DNA dependant ATPase and DNA Helicase involved in DNA repair, MRPS18C encoding the Mitochondrial Ribosomal Protein S18C and FAM175A (ABRAXAS), encoding a BRCA1 BRCT domain-interacting protein involved in DNA damage response and double-strand break (DSB) repair. Expression QTL analysis in breast cancer tissue showed rs11099601 to be associated with HELQ (P = 8.28x10-14), MRPS18C (P = 1.94x10-27) and FAM175A (P = 3.83x10-3), explaining about 20%, 14% and 1%, respectively of the variance inexpression of these genes in breast carcinomas.

  18. Association of breast cancer risk with genetic variants showing differential allelic expression: Identification of a novel breast cancer susceptibility locus at 4q21

    PubMed Central

    Adoue, Véronique; Michailidou, Kyriaki; Canisius, Sander; Lemaçon, Audrey; Droit, Arnaud; Andrulis, Irene L; Anton-Culver, Hoda; Arndt, Volker; Baynes, Caroline; Blomqvist, Carl; Bogdanova, Natalia V.; Bojesen, Stig E.; Bolla, Manjeet K.; Bonanni, Bernardo; Borresen-Dale, Anne-Lise; Brand, Judith S.; Brauch, Hiltrud; Brenner, Hermann; Broeks, Annegien; Burwinkel, Barbara; Chang-Claude, Jenny; Couch, Fergus J.; Cox, Angela; Cross, Simon S.; Czene, Kamila; Darabi, Hatef; Dennis, Joe; Devilee, Peter; Dörk, Thilo; Dos-Santos-Silva, Isabel; Eriksson, Mikael; Fasching, Peter A.; Figueroa, Jonine; Flyger, Henrik; García-Closas, Montserrat; Giles, Graham G.; Goldberg, Mark S.; González-Neira, Anna; Grenaker-Alnæs, Grethe; Guénel, Pascal; Haeberle, Lothar; Haiman, Christopher A.; Hamann, Ute; Hallberg, Emily; Hooning, Maartje J.; Hopper, John L.; Jakubowska, Anna; Jones, Michael; Kabisch, Maria; Kataja, Vesa; Lambrechts, Diether; Marchand, Loic Le; Lindblom, Annika; Lubinski, Jan; Mannermaa, Arto; Maranian, Mel; Margolin, Sara; Marme, Frederik; Milne, Roger L.; Neuhausen, Susan L.; Nevanlinna, Heli; Neven, Patrick; Olswold, Curtis; Peto, Julian; Plaseska-Karanfilska, Dijana; Pylkäs, Katri; Radice, Paolo; Rudolph, Anja; Sawyer, Elinor J.; Schmidt, Marjanka K.; Shu, Xiao-Ou; Southey, Melissa C.; Swerdlow, Anthony; Tollenaar, Rob A.E.M.; Tomlinson, Ian; Torres, Diana; Truong, Thérèse; Vachon, Celine; Van Den Ouweland, Ans M. W.; Wang, Qin; Winqvist, Robert; Investigators, kConFab/AOCS; Zheng, Wei; Benitez, Javier; Chenevix-Trench, Georgia; Dunning, Alison M.; Pharoah, Paul D. P.; Kristensen, Vessela; Hall, Per; Easton, Douglas F.; Pastinen, Tomi; Nord, Silje; Simard, Jacques

    2016-01-01

    There are significant inter-individual differences in the levels of gene expression. Through modulation of gene expression, cis-acting variants represent an important source of phenotypic variation. Consequently, cis-regulatory SNPs associated with differential allelic expression are functional candidates for further investigation as disease-causing variants. To investigate whether common variants associated with differential allelic expression were involved in breast cancer susceptibility, a list of genes was established on the basis of their involvement in cancer related pathways and/or mechanisms. Thereafter, using data from a genome-wide map of allelic expression associated SNPs, 313 genetic variants were selected and their association with breast cancer risk was then evaluated in 46,451 breast cancer cases and 42,599 controls of European ancestry ascertained from 41 studies participating in the Breast Cancer Association Consortium. The associations were evaluated with overall breast cancer risk and with estrogen receptor negative and positive disease. One novel breast cancer susceptibility locus on 4q21 (rs11099601) was identified (OR = 1.05, P = 5.6x10-6). rs11099601 lies in a 135 kb linkage disequilibrium block containing several genes, including, HELQ, encoding the protein HEL308 a DNA dependant ATPase and DNA Helicase involved in DNA repair, MRPS18C encoding the Mitochondrial Ribosomal Protein S18C and FAM175A (ABRAXAS), encoding a BRCA1 BRCT domain-interacting protein involved in DNA damage response and double-strand break (DSB) repair. Expression QTL analysis in breast cancer tissue showed rs11099601 to be associated with HELQ (P = 8.28x10-14), MRPS18C (P = 1.94x10-27) and FAM175A (P = 3.83x10-3), explaining about 20%, 14% and 1%, respectively of the variance inexpression of these genes in breast carcinomas. PMID:27792995

  19. NADPH oxidase complex and IBD candidate gene studies: identification of a rare variant in NCF2 that results in reduced binding to RAC2

    PubMed Central

    Muise, Aleixo M; Xu, Wei; Guo, Cong-Hui; Walters, Thomas D; Wolters, Victorien M; Fattouh, Ramzi; Lam, Grace Y; Hu, Pingzhao; Murchie, Ryan; Sherlock, Mary; Gana, Juan Cristóbal; Russell, Richard K; Glogauer, Michael; Duerr, Richard H; Cho, Judy H; Lees, Charlie W; Satsangi, Jack; Wilson, David C; Paterson, Andrew D; Griffiths, Anne M; Silverberg, Mark S; Brumell, John H

    2013-01-01

    Objective The NOX2 NADPH oxidase complex produces reactive oxygen species and plays a critical role in the killing of microbes by phagocytes. Genetic mutations in genes encoding components of the complex result in both X-linked and autosomal recessive forms of chronic granulomatous disease (CGD). Patients with CGD often develop intestinal inflammation that is histologically similar to Crohn's colitis, suggesting a common aetiology for both diseases. The aim of this study is to determine if polymorphisms in NOX2 NADPH oxidase complex genes that do not cause CGD are associated with the development of inflammatory bowel disease (IBD). Methods Direct sequencing and candidate gene approaches were used to identify susceptibility loci in NADPH oxidase complex genes. Functional studies were carried out on identified variants. Novel findings were replicated in independent cohorts. Results Sequence analysis identified a novel missense variant in the neutrophil cytosolic factor 2 (NCF2) gene that is associated with very early onset IBD (VEO-IBD) and subsequently found in 4% of patients with VEO-IBD compared with 0.2% of controls (p=1.3×10−5, OR 23.8 (95% CI 3.9 to 142.5); Fisher exact test). This variant reduced binding of the NCF2 gene product p67phox to RAC2. This study found a novel genetic association of RAC2 with Crohn's disease (CD) and replicated the previously reported association of NCF4 with ileal CD. Conclusion These studies suggest that the rare novel p67phox variant results in partial inhibition of oxidase function and are associated with CD in a subgroup of patients with VEO-IBD; and suggest that components of the NADPH oxidase complex are associated with CD. PMID:21900546

  20. Identification and validation of immunogenic potential of India specific HPV-16 variant constructs: In-silico & in-vivo insight to vaccine development

    PubMed Central

    Kumar, Anoop; Hussain, Showket; Sharma, Gagan; Mehrotra, Ravi; Gissmann, Lutz; Das, Bhudev C.; Bharadwaj, Mausumi

    2015-01-01

    Cervical cancer is one of the most common gynecological cancers in the world but in India, it is the top most cancer among women. Persistent infection with high-risk human papillomaviruses (HR-HPVs) is the most important risk factor. The sequence variation(s) in the most common HR-HPV i.e. HPV type 16 leads to altered biological functions with possible clinical significance in the different geographical locations. Sixteen major variants (V1-V16) in full length L1 gene of HPV-16 were identified following analysis of 250 prospectively collected cervical cancer tissue biopsies and their effect on immunogenicity was studied. The effect of these major variations on the epitopes were predicted by in silico methods and the immunogenicity of variants and respective reference DNA vaccine constructs were evaluated by administration of prepared DNA vaccine constructs in female BALB/c mice to evaluate antibody titer. In the present study, L500F (V16) variation showed a significant ~2.7 fold (p < 0.002) increase in antibody titer, whereas T379P (V8) showed ~0.4 fold (p < 0.328) decrease after final injection. These results showed a promising roadmap for the development of DNA based vaccine and for the generation of effective response, though there is a need to study more prevalent variants of HPV in the Indian population. PMID:26507515

  1. Identification of the thiamin pyrophosphokinase gene in rainbow trout: Characteristic structure and expression of seven splice variants in tissues and cell lines and during embryo development

    USGS Publications Warehouse

    Yuge, Shinya; Richter, Catherine A.; Wright-Osment, Maureen K.; Nicks, Diane; Saloka, Stephanie K.; Tillitt, Donald E.; Li, Weiming

    2012-01-01

    Thiamin pyrophosphokinase (TPK) converts thiamin to its active form, thiamin diphosphate. In humans, TPK expression is down-regulated in some thiamin deficiency related syndrome, and enhanced during pregnancy. Rainbow trout are also vulnerable to thiamin deficiency in wild life and are useful models for thiamin metabolism research. We identified the tpk gene transcript including seven splice variants in the rainbow trout. Almost all cell lines and tissues examined showed co-expression of several tpk splice variants including a potentially major one at both mRNA and protein levels. However, relative to other tissues, the longest variant mRNA expression was predominant in the ovary and abundant in embryos. During embryogenesis, total tpk transcripts increased abruptly in early development, and decreased to about half of the peak shortly after hatching. In rainbow trout, the tpk transcript complex is ubiquitously expressed for all tissues and cells examined, and its increase in expression could be important in the early-middle embryonic stages. Moreover, decimated tpk expression in a hepatoma cell line relative to hepatic and gonadal cell lines appears to be consistent with previously reported down-regulation of thiamin metabolism in cancer.

  2. Identification of the thiamin pyrophosphokinase gene in rainbow trout: characteristic structure and expression of seven splice variants in tissues and cell lines and during embryo development.

    PubMed

    Yuge, Shinya; Richter, Catherine A; Wright-Osment, Maureen K; Nicks, Diane; Saloka, Stephanie K; Tillitt, Donald E; Li, Weiming

    2012-10-01

    Thiamin pyrophosphokinase (TPK) converts thiamin to its active form, thiamin diphosphate. In humans, TPK expression is down-regulated in some thiamin deficiency related syndrome, and enhanced during pregnancy. Rainbow trout are also vulnerable to thiamin deficiency in wild life and are useful models for thiamin metabolism research. We identified the tpk gene transcript including seven splice variants in the rainbow trout. Almost all cell lines and tissues examined showed co-expression of several tpk splice variants including a potentially major one at both mRNA and protein levels. However, relative to other tissues, the longest variant mRNA expression was predominant in the ovary and abundant in embryos. During embryogenesis, total tpk transcripts increased abruptly in early development, and decreased to about half of the peak shortly after hatching. In rainbow trout, the tpk transcript complex is ubiquitously expressed for all tissues and cells examined, and its increase in expression could be important in the early-middle embryonic stages. Moreover, decimated tpk expression in a hepatoma cell line relative to hepatic and gonadal cell lines appears to be consistent with previously reported down-regulation of thiamin metabolism in cancer. Copyright © 2012 Elsevier Inc. All rights reserved.

  3. Caveolin-1 isoforms are encoded by distinct mRNAs. Identification Of mouse caveolin-1 mRNA variants caused by alternative transcription initiation and splicing.

    PubMed

    Kogo, H; Fujimoto, T

    2000-01-14

    By searching the EST database with the known cDNA sequence encoding alpha-caveolin-1 (full-length: FL), we found a variant having a hitherto unknown sequence in place of the first exon (5'-end variant: 5'V). The expression level of 5'V mRNA was equivalent to that of FL mRNA. The entire sequences of FL and 5'V mRNA were determined by 3'- and 5'-RACE analysis; their sizes were 2484 bp and 2533 bp, respectively, and the sequences were identical except for the region of the first exon. By Northern blotting, FL and 5'V mRNAs showed the same tissue distribution, and were intensely expressed in the lung, heart, and skeletal muscle. Analyzing the protein production from these mRNAs using green fluorescent protein as a tag, we found FL mRNA to produce the alpha-isoform predominantly, but to form little beta-isoform. The production of the beta-isoform from 5'V mRNA was also demonstrated. By sequence analysis of the first intron of the caveolin-1 gene, a TATA box was found at 28 bp upstream of the transcription initiation site for 5'V mRNA. This is the first demonstration of caveolin-1 mRNA variants generated by alternative transcription initiation, and it indicates that the two isoforms of caveolin-1 are produced from two distinct mRNAs.

  4. Isolation and identification of host cues from mango, Mangifera indica, that attract gravid female oriental fruit fly, Bactrocera dorsalis.

    PubMed

    Jayanthi, Pagadala D Kamala; Woodcock, Christine M; Caulfield, John; Birkett, Michael A; Bruce, Toby J A

    2012-04-01

    The oriental fruit fly, Bactrocera dorsalis, is an economically damaging, polyphagous pest of fruit crops in South-East Asia and Hawaii, and a quarantine pest in other parts of the world. The objective of our study was to identify new attractants for B. dorsalis from overripe mango fruits. Headspace samples of volatiles were collected from two cultivars of mango, 'Alphonso' and 'Chausa', and a strong positive behavioral response was observed when female B. dorsalis were exposed to these volatiles in olfactometer bioassays. Coupled GC-EAG with female B. dorsalis revealed 7 compounds from 'Alphonso' headspace and 15 compounds from 'Chausa' headspace that elicited an EAG response. The EAG-active compounds, from 'Alphonso', were identified, using GC-MS, as heptane, myrcene, (Z)-ocimene, (E)-ocimene, allo-ocimene, (Z)-myroxide, and γ-octalactone, with the two ocimene isomers being the dominant compounds. The EAG-active compounds from 'Chausa' were 3-hydroxy-2-butanone, 3-methyl-1-butanol, ethyl butanoate, ethyl methacrylate, ethyl crotonate, ethyl tiglate, 1-octen-3-ol, ethyl hexanoate, 3-carene, p-cymene, ethyl sorbate, α-terpinolene, phenyl ethyl alcohol, ethyl octanoate, and benzothiazole. Individual compounds were significantly attractive when a standard dose (1 μg on filter paper) was tested in the olfactometer. Furthermore, synthetic blends with the same concentration and ratio of compounds as in the natural headspace samples were highly attractive (P < 0.001), and in a choice test, fruit flies did not show any preference for the natural samples over the synthetic blends. Results are discussed in relation to developing a lure for female B. dorsalis to bait traps with.

  5. Identification of independent association signals and putative functional variants for breast cancer risk through fine-scale mapping of the 12p11 locus.

    PubMed

    Zeng, Chenjie; Guo, Xingyi; Long, Jirong; Kuchenbaecker, Karoline B; Droit, Arnaud; Michailidou, Kyriaki; Ghoussaini, Maya; Kar, Siddhartha; Freeman, Adam; Hopper, John L; Milne, Roger L; Bolla, Manjeet K; Wang, Qin; Dennis, Joe; Agata, Simona; Ahmed, Shahana; Aittomäki, Kristiina; Andrulis, Irene L; Anton-Culver, Hoda; Antonenkova, Natalia N; Arason, Adalgeir; Arndt, Volker; Arun, Banu K; Arver, Brita; Bacot, Francois; Barrowdale, Daniel; Baynes, Caroline; Beeghly-Fadiel, Alicia; Benitez, Javier; Bermisheva, Marina; Blomqvist, Carl; Blot, William J; Bogdanova, Natalia V; Bojesen, Stig E; Bonanni, Bernardo; Borresen-Dale, Anne-Lise; Brand, Judith S; Brauch, Hiltrud; Brennan, Paul; Brenner, Hermann; Broeks, Annegien; Brüning, Thomas; Burwinkel, Barbara; Buys, Saundra S; Cai, Qiuyin; Caldes, Trinidad; Campbell, Ian; Carpenter, Jane; Chang-Claude, Jenny; Choi, Ji-Yeob; Claes, Kathleen B M; Clarke, Christine; Cox, Angela; Cross, Simon S; Czene, Kamila; Daly, Mary B; de la Hoya, Miguel; De Leeneer, Kim; Devilee, Peter; Diez, Orland; Domchek, Susan M; Doody, Michele; Dorfling, Cecilia M; Dörk, Thilo; Dos-Santos-Silva, Isabel; Dumont, Martine; Dwek, Miriam; Dworniczak, Bernd; Egan, Kathleen; Eilber, Ursula; Einbeigi, Zakaria; Ejlertsen, Bent; Ellis, Steve; Frost, Debra; Lalloo, Fiona; Fasching, Peter A; Figueroa, Jonine; Flyger, Henrik; Friedlander, Michael; Friedman, Eitan; Gambino, Gaetana; Gao, Yu-Tang; Garber, Judy; García-Closas, Montserrat; Gehrig, Andrea; Damiola, Francesca; Lesueur, Fabienne; Mazoyer, Sylvie; Stoppa-Lyonnet, Dominique; Giles, Graham G; Godwin, Andrew K; Goldgar, David E; González-Neira, Anna; Greene, Mark H; Guénel, Pascal; Haeberle, Lothar; Haiman, Christopher A; Hallberg, Emily; Hamann, Ute; Hansen, Thomas V O; Hart, Steven; Hartikainen, Jaana M; Hartman, Mikael; Hassan, Norhashimah; Healey, Sue; Hogervorst, Frans B L; Verhoef, Senno; Hendricks, Carolyn B; Hillemanns, Peter; Hollestelle, Antoinette; Hulick, Peter J; Hunter, David J; Imyanitov, Evgeny N; Isaacs, Claudine; Ito, Hidemi; Jakubowska, Anna; Janavicius, Ramunas; Jaworska-Bieniek, Katarzyna; Jensen, Uffe Birk; John, Esther M; Joly Beauparlant, Charles; Jones, Michael; Kabisch, Maria; Kang, Daehee; Karlan, Beth Y; Kauppila, Saila; Kerin, Michael J; Khan, Sofia; Khusnutdinova, Elza; Knight, Julia A; Konstantopoulou, Irene; Kraft, Peter; Kwong, Ava; Laitman, Yael; Lambrechts, Diether; Lazaro, Conxi; Le Marchand, Loic; Lee, Chuen Neng; Lee, Min Hyuk; Lester, Jenny; Li, Jingmei; Liljegren, Annelie; Lindblom, Annika; Lophatananon, Artitaya; Lubinski, Jan; Mai, Phuong L; Mannermaa, Arto; Manoukian, Siranoush; Margolin, Sara; Marme, Frederik; Matsuo, Keitaro; McGuffog, Lesley; Meindl, Alfons; Menegaux, Florence; Montagna, Marco; Muir, Kenneth; Mulligan, Anna Marie; Nathanson, Katherine L; Neuhausen, Susan L; Nevanlinna, Heli; Newcomb, Polly A; Nord, Silje; Nussbaum, Robert L; Offit, Kenneth; Olah, Edith; Olopade, Olufunmilayo I; Olswold, Curtis; Osorio, Ana; Papi, Laura; Park-Simon, Tjoung-Won; Paulsson-Karlsson, Ylva; Peeters, Stephanie; Peissel, Bernard; Peterlongo, Paolo; Peto, Julian; Pfeiler, Georg; Phelan, Catherine M; Presneau, Nadege; Radice, Paolo; Rahman, Nazneen; Ramus, Susan J; Rashid, Muhammad Usman; Rennert, Gad; Rhiem, Kerstin; Rudolph, Anja; Salani, Ritu; Sangrajrang, Suleeporn; Sawyer, Elinor J; Schmidt, Marjanka K; Schmutzler, Rita K; Schoemaker, Minouk J; Schürmann, Peter; Seynaeve, Caroline; Shen, Chen-Yang; Shrubsole, Martha J; Shu, Xiao-Ou; Sigurdson, Alice; Singer, Christian F; Slager, Susan; Soucy, Penny; Southey, Melissa; Steinemann, Doris; Swerdlow, Anthony; Szabo, Csilla I; Tchatchou, Sandrine; Teixeira, Manuel R; Teo, Soo H; Terry, Mary Beth; Tessier, Daniel C; Teulé, Alex; Thomassen, Mads; Tihomirova, Laima; Tischkowitz, Marc; Toland, Amanda E; Tung, Nadine; Turnbull, Clare; van den Ouweland, Ans M W; van Rensburg, Elizabeth J; Ven den Berg, David; Vijai, Joseph; Wang-Gohrke, Shan; Weitzel, Jeffrey N; Whittemore, Alice S; Winqvist, Robert; Wong, Tien Y; Wu, Anna H; Yannoukakos, Drakoulis; Yu, Jyh-Cherng; Pharoah, Paul D P; Hall, Per; Chenevix-Trench, Georgia; Dunning, Alison M; Simard, Jacques; Couch, Fergus J; Antoniou, Antonis C; Easton, Douglas F; Zheng, Wei

    2016-06-21

    Multiple recent genome-wide association studies (GWAS) have identified a single nucleotide polymorphism (SNP), rs10771399, at 12p11 that is associated with breast cancer risk. We performed a fine-scale mapping study of a 700 kb region including 441 genotyped and more than 1300 imputed genetic variants in 48,155 cases and 43,612 controls of European descent, 6269 cases and 6624 controls of East Asian descent and 1116 cases and 932 controls of African descent in the Breast Cancer Association Consortium (BCAC; http://bcac.ccge.medschl.cam.ac.uk/ ), and in 15,252 BRCA1 mutation carriers in the Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA). Stepwise regression analyses were performed to identify independent association signals. Data from the Encyclopedia of DNA Elements project (ENCODE) and the Cancer Genome Atlas (TCGA) were used for functional annotation. Analysis of data from European descendants found evidence for four independent association signals at 12p11, represented by rs7297051 (odds ratio (OR) = 1.09, 95 % confidence interval (CI) = 1.06-1.12; P = 3 × 10(-9)), rs805510 (OR = 1.08, 95 % CI = 1.04-1.12, P = 2 × 10(-5)), and rs1871152 (OR = 1.04, 95 % CI = 1.02-1.06; P = 2 × 10(-4)) identified in the general populations, and rs113824616 (P = 7 × 10(-5)) identified in the meta-analysis of BCAC ER-negative cases and BRCA1 mutation carriers. SNPs rs7297051, rs805510 and rs113824616 were also associated with breast cancer risk at P < 0.05 in East Asians, but none of the associations were statistically significant in African descendants. Multiple candidate functional variants are located in putative enhancer sequences. Chromatin interaction data suggested that PTHLH was the likely target gene of these enhancers. Of the six variants with the strongest evidence of potential functionality, rs11049453 was statistically significantly associated with the expression of PTHLH and its nearby

  6. Identification of novel subgroup A variants with enhanced receptor binding and replicative capacity in primary isolates of anaemogenic strains of feline leukaemia virus.

    PubMed

    Stewart, Hazel; Adema, Karen W; McMonagle, Elizabeth L; Hosie, Margaret J; Willett, Brian J

    2012-05-31

    The development of anaemia in feline leukaemia virus (FeLV)-infected cats is associated with the emergence of a novel viral subgroup, FeLV-C. FeLV-C arises from the subgroup that is transmitted, FeLV-A, through alterations in the amino acid sequence of the receptor binding domain (RBD) of the envelope glycoprotein that result in a shift in the receptor usage and the cell tropism of the virus. The factors that influence the transition from subgroup A to subgroup C remain unclear, one possibility is that a selective pressure in the host drives the acquisition of mutations in the RBD, creating A/C intermediates with enhanced abilities to interact with the FeLV-C receptor, FLVCR. In order to understand further the emergence of FeLV-C in the infected cat, we examined primary isolates of FeLV-C for evidence of FeLV-A variants that bore mutations consistent with a gradual evolution from FeLV-A to FeLV-C. Within each isolate of FeLV-C, we identified variants that were ostensibly subgroup A by nucleic acid sequence comparisons, but which bore mutations in the RBD. One such mutation, N91D, was present in multiple isolates and when engineered into a molecular clone of the prototypic FeLV-A (Glasgow-1), enhanced replication was noted in feline cells. Expression of the N91D Env on murine leukaemia virus (MLV) pseudotypes enhanced viral entry mediated by the FeLV-A receptor THTR1 while soluble FeLV-A Env bearing the N91D mutation bound more efficiently to mouse or guinea pig cells bearing the FeLV-A and -C receptors. Long-term in vitro culture of variants bearing the N91D substitution in the presence of anti-FeLV gp70 antibodies did not result in the emergence of FeLV-C variants, suggesting that additional selective pressures in the infected cat may drive the subsequent evolution from subgroup A to subgroup C. Our data support a model in which variants of FeLV-A, bearing subtle differences in the RBD of Env, may be predisposed towards enhanced replication in vivo and subsequent

  7. Characterization of recombinant long-chain rat acyl-CoA synthetase isoforms 3 and 6: identification of a novel variant of isoform 6.

    PubMed

    Van Horn, Cynthia G; Caviglia, Jorge M; Li, Lei O; Wang, Shuli; Granger, Deborah A; Coleman, Rosalind A

    2005-02-08

    The metabolism of long-chain fatty acids in brain and their incorporation into signaling molecules such as diacylglycerol and LPA and into structural components of membranes, including myelin, requires activation by long-chain acyl-CoA synthetase (ACSL). Because ACSL3 and ACSL6 are the predominant ACSL isoforms in brain, we cloned and characterized these isoforms from rat brain and identified a novel ACSL6 clone (ACSL6_v2). ACSL6_v2 and the previously reported ACSL6_v1 represent splice variants that include exon 13 or 14, respectively. Homologue sequences of both of these variants are present in the human and mouse databases. ACSL3, ACSL6_v1, and ACSL6_v2 with Flag-epitopes at the C-termini were expressed in Escherichia coli and purified on Flag-affinity columns. The three recombinant proteins were characterized. Compared to ACSL4, another brain isoform, ACSL3, ACSL6_v1, and ACSL6_v2 showed similarities in kinetic values for CoA, palmitate, and arachidonate, but their apparent Km values for oleate were 4- to 6-fold lower than for ACSL4. In a direct competition assay with palmitate, all the polyunsaturated fatty acids tested were strong competitors only for ACSL4 with IC50 values of 0.5 to 5 microM. DHA was also strongly preferred by ACSL6_v2. The apparent Km value for ATP of ACSL6_v1 was 8-fold higher than that of ACSL6_v2. ACSL3 and the two variants of ACSL6 were more resistant than ACSL4 to heat inactivation. Despite the high amino acid identity between ACSL3 and ACSL4, rosiglitazone inhibited only ACSL4. Triacsin C, an inhibitor of ACSL1 and ACSL4, also inhibited ACSL3, but did not inhibit the ACSL6 variants. These data further document important differences in the closely related ACSL isoforms and show that amino acid changes near the consensus nucleotide binding site alter function in the two splice variants of ACSL6.

  8. Automatic identification of agricultural terraces through object-oriented analysis of very high resolution DSMs and multispectral imagery obtained from an unmanned aerial vehicle.

    PubMed

    Diaz-Varela, R A; Zarco-Tejada, P J; Angileri, V; Loudjani, P

    2014-02-15

    Agricultural terraces are features that provide a number of ecosystem services. As a result, their maintenance is supported by measures established by the European Common Agricultural Policy (CAP). In the framework of CAP implementation and monitoring, there is a current and future need for the development of robust, repeatable and cost-effective methodologies for the automatic identification and monitoring of these features at farm scale. This is a complex task, particularly when terraces are associated to complex vegetation cover patterns, as happens with permanent crops (e.g. olive trees). In this study we present a novel methodology for automatic and cost-efficient identification of terraces using only imagery from commercial off-the-shelf (COTS) cameras on board unmanned aerial vehicles (UAVs). Using state-of-the-art computer vision techniques, we generated orthoimagery and digital surface models (DSMs) at 11 cm spatial resolution with low user intervention. In a second stage, these data were used to identify terraces using a multi-scale object-oriented classification method. Results show the potential of this method even in highly complex agricultural areas, both regarding DSM reconstruction and image classification. The UAV-derived DSM had a root mean square error (RMSE) lower than 0.5 m when the height of the terraces was assessed against field GPS data. The subsequent automated terrace classification yielded an overall accuracy of 90% based exclusively on spectral and elevation data derived from the UAV imagery. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. Plant Gene and Alternatively Spliced Variant Annotator. A plant genome annotation pipeline for rice gene and alternatively spliced variant identification with cross-species expressed sequence tag conservation from seven plant species.

    PubMed

    Chen, Feng-Chi; Wang, Sheng-Shun; Chaw, Shu-Miaw; Huang, Yao-Ting; Chuang, Trees-Juen

    2007-03-01

    The completion of the rice (Oryza sativa) genome draft has brought unprecedented opportunities for genomic studies of the world's most important food crop. Previous rice gene annotations have relied mainly on ab initio methods, which usually yield a high rate of false-positive predictions and give only limited information regarding alternative splicing in rice genes. Comparative approaches based on expressed sequence tags (ESTs) can compensate for the drawbacks of ab initio methods because they can simultaneously identify experimental data-supported genes and alternatively spliced transcripts. Furthermore, cross-species EST information can be used to not only offset the insufficiency of same-species ESTs but also derive evolutionary implications. In this study, we used ESTs from seven plant species, rice, wheat (Triticum aestivum), maize (Zea mays), barley (Hordeum vulgare), sorghum (Sorghum bicolor), soybean (Glycine max), and Arabidopsis (Arabidopsis thaliana), to annotate the rice genome. We developed a plant genome annotation pipeline, Plant Gene and Alternatively Spliced Variant Annotator (PGAA). Using this approach, we identified 852 genes (931 isoforms) not annotated in other widely used databases (i.e. the Institute for Genomic Research, National Center for Biotechnology Information, and Rice Annotation Project) and found 87% of them supported by both rice and nonrice EST evidence. PGAA also identified more than 44,000 alternatively spliced events, of which approximately 20% are not observed in the other three annotations. These novel annotations represent rich opportunities for rice genome research, because the functions of most of our annotated genes are currently unknown. Also, in the PGAA annotation, the isoforms with non-rice-EST-supported exons are significantly enriched in transporter activity but significantly underrepresented in transcription regulator activity. We have also identified potential lineage-specific and conserved isoforms, which are

  10. Reliably Detecting Clinically Important Variants Requires Both Combined Variant Calls and Optimized Filtering Strategies

    PubMed Central

    Field, Matthew A.; Cho, Vicky

    2015-01-01

    A diversity of tools is available for identification of variants from genome sequence data. Given the current complexity of incorporating external software into a genome analysis infrastructure, a tendency exists to rely on the results from a single tool alone. The quality of the output variant calls is highly variable however, depending on factors such as sequence library quality as well as the choice of short-read aligner, variant caller, and variant caller filtering strategy. Here we present a two-part study first using the high quality ‘genome in a bottle’ reference set to demonstrate the significant impact the choice of aligner, variant caller, and variant caller filtering strategy has on overall variant call quality and further how certain variant callers outperform others with increased sample contamination, an important consideration when analyzing sequenced cancer samples. This analysis confirms previous work showing that combining variant calls of multiple tools results in the best quality resultant variant set, for either specificity or sensitivity, depending on whether the intersection or union, of all variant calls is used respectively. Second, we analyze a melanoma cell line derived from a control lymphocyte sample to determine whether software choices affect the detection of clinically important melanoma risk-factor variants finding that only one of the three such variants is unanimously detected under all conditions. Finally, we describe a cogent strategy for implementing a clinical variant detection pipeline; a strategy that requires careful software selection, variant caller filtering optimizing, and combined variant calls in order to effectively minimize false negative variants. While implementing such features represents an increase in complexity and computation the results offer indisputable improvements in data quality. PMID:26600436

  11. Acute intermittent porphyria: identification and expression of exonic mutations in the hydroxymethylbilane synthase gene. An initiation codon missense mutation in the housekeeping transcript causes "variant acute intermittent porphyria" with normal expression of the erythroid-specific enzyme.

    PubMed Central

    Chen, C H; Astrin, K H; Lee, G; Anderson, K E; Desnick, R J

    1994-01-01

    Acute intermittent porphyria (AIP), an autosomal dominant inborn error, results from the half-normal activity of the heme biosynthetic enzyme, hydroxymethylbilane synthase (EC 4.3.1.8). Diagnosis of AIP heterozygotes is essential to prevent acute, life-threatening neurologic attacks by avoiding various precipitating factors. Since biochemical diagnosis is problematic, the identification of hydroxymethylbilane synthase mutations has facilitated the detection of AIP heterozygotes. Molecular analyses of unrelated AIP patients revealed six exonic mutations: an initiating methionine to isoleucine substitution (M1I) in a patient with variant AIP, which precluded translation of the housekeeping, but not the erythroid-specific isozyme; four missense mutations in classical AIP patients, V93F, R116W, R201W, C247F; and a nonsense mutation W283X in a classical AIP patient, which truncated the housekeeping and erythroid-specific isozymes. Each mutation was confirmed in genomic DNA from family members. The W283X lesion was found in another unrelated AIP family. Expression of each mutation in Escherichia coli revealed that R201W, C247F, and W283X had residual activity. In vitro transcription/translation studies indicated that the M1I allele produced only the erythroid-specific enzyme, while the other mutant alleles encoded both isozymes. These mutations provide insight into the molecular pathology of classic and variant AIP and facilitate molecular diagnosis in AIP families. Images PMID:7962538

  12. A genetic polymorphism in coumarin 7-hydroxylation: Sequence of the human CYP2A genes and identification of variant CYP2A6 alleles

    SciTech Connect

    Fernandez-Salguero, P.; Hoffman, S.M.G.; Mohrenweiser, H.

    1995-09-01

    A group of human cytochrome P450 genes encompassing the CYP2A, CYP2B, and CYP2F subfamilies were cloned and assembled into a 350-kb contig localized on the long arm of chromosome 19. Three complete CYP2A genes - CYP2A6, CYP2A7, and CYP2A13 - plus two pseudogenes truncated after exon 5 were identified and sequenced. A variant CYP2A6 allele that differed from the corresponding CYP2A6 and CYP2A7 cDNAs previously sequenced was found and was designated CYP2A6{nu}2. Sequence differences in the CY-P2A6{nu}2 gene are restricted to regions encompassing exons 3, 6, and 8, which bear sequence relatedness with the corresponding exons of the CYP2A7 gene, located downstream and centromeric of CYP2A6{nu}2, suggesting recent gene-conversion events. The sequencing of all the CYP2A genes allowed the design of a PCR diagnostic test for the normal CYP2A6 allele, the CYP2A6{nu}2 allele, and a variant - designated CYP2A6{nu}1 - that encodes an enzyme with a single inactivating amino acid change. These variant alleles were found in individuals who were deficient in their ability to metabolize the CYP2A6 probe drug coumarin. The allelic frequencies of CYP2A6{nu}1 and CYP2A6{nu}2 differed significantly between Caucasian, Asian, and African-American populations. These studies establish the existence of a new cytochrome P450 genetic polymorphism. 30 refs., 4 figs., 2 tabs.

  13. Identification of common variants in the SHBG gene affecting sex hormone binding globulin levels and breast cancer risk in postmenopausal women

    PubMed Central

    Thompson, Deborah J; Healey, Catherine S; Baynes, Caroline; Kalmyrzaev, Bolot; Ahmed, Shahana; Dowsett, Mitch; Folkerd, Elizabeth; Luben, Robert N; Cox, David; Ballinger, Dennis; Pharoah, Paul DP; Ponder, Bruce AJ; Dunning, Alison M; Easton, Douglas F

    2009-01-01

    Background Circulating levels of sex hormone-binding globulin (SHBG) are inversely associated with breast cancer risk in post-menopausal women. Three polymorphisms within the SHBG gene have been reported to affect SHBG levels, but there has been no systematic attempt to identify other such variants. Methods We looked for associations between SHBG levels in 1134 healthy, postmenopausal women and 11 tagging single nucleotide polymorphisms (SNPs) in or around the SHBG gene. Associations between SHBG SNPs and breast cancer were tested in up to 6622 post-menopausal breast cancer cases and 6784 controls. Results Ten SNPs within or close to the SHBG gene were significantly associated with SHBG levels, as was the (TAAAA)n polymorphism. The best-fitting combination of rs6259, rs858521, rs727428 and body mass index, waist, hip, age and smoking status accounted for 24% of the variance in SHBG levels (natural logarithm transformed). Haplotype analysis suggested that rs858518, rs727428 or a variant in linkage disequilibrium with them, acts to decrease SHBG levels, but that this effect is neutralised by rs6259 (D356N). rs1799941 increases SHBG levels, but the previously reported association with (TAAAA)n repeat length appears to be a consequence of linkage disequilibrium with these SNPs. One further SHBG SNP was significantly associated with breast cancer (rs6257, per-allele odds ratio 0.88, 95% CI=0.82-0.95, p=0.002). Conclusion At least three SNPs showed associations with SHBG levels that were highly significant but relatively small in magnitude. rs6257 is a potential breast cancer susceptibility variant, but relationships between the genetic determinants of SHBG levels and breast cancer are complex. PMID:19064566

  14. Identification of common variants in the SHBG gene affecting sex hormone-binding globulin levels and breast cancer risk in postmenopausal women.

    PubMed

    Thompson, Deborah J; Healey, Catherine S; Baynes, Caroline; Kalmyrzaev, Bolot; Ahmed, Shahana; Dowsett, Mitch; Folkerd, Elizabeth; Luben, Robert N; Cox, David; Ballinger, Dennis; Pharoah, Paul D P; Ponder, Bruce A J; Dunning, Alison M; Easton, Douglas F

    2008-12-01

    Circulating levels of sex hormone-binding globulin (SHBG) are inversely associated with breast cancer risk in postmenopausal women. Three polymorphisms within the SHBG gene have been reported to affect SHBG levels, but there has been no systematic attempt to identify other such variants. We looked for associations between SHBG levels in 1,134 healthy, postmenopausal women and 11 tagging single nucleotide polymorphisms (SNP) in or around the SHBG gene. Associations between SHBG SNPs and breast cancer were tested in up to 6,622 postmenopausal breast cancer cases and 6,784 controls. Ten SNPs within or close to the SHBG gene were significantly associated with SHBG levels as was the (TAAAA)(n) polymorphism. The best-fitting combination of rs6259, rs858521, and rs727428 and body mass index, waist, hip, age, and smoking status accounted for 24% of the variance in SHBG levels (natural logarithm transformed). Haplotype analysis suggested that rs858518, rs727428, or a variant in linkage disequilibrium with them acts to decrease SHBG levels but that this effect is neutralized by rs6259 (D356N). rs1799941 increases SHBG levels, but the previously reported association with (TAAAA)(n) repeat length appears to be a consequence of linkage disequilibrium with these SNPs. One further SHBG SNP was significantly associated with breast cancer (rs6257, per-allele odds ratio, 0.88; 95% confidence interval, 0.82-0.95; P = 0.002). At least 3 SNPs showed associations with SHBG levels that were highly significant but relatively small in magnitude. rs6257 is a potential breast cancer susceptibility variant, but relationships between the genetic determinants of SHBG levels and breast cancer are complex.

  15. Identification of genetic risk variants for deep vein thrombosis by multiplexed next-generation sequencing of 186 hemostatic/pro-inflammatory genes

    PubMed Central

    2012-01-01

    Background Next-generation DNA sequencing is opening new avenues for genetic association studies in common diseases that, like deep vein thrombosis (DVT), have a strong genetic predisposition still largely unexplained by currently identified risk variants. In order to develop sequencing and analytical pipelines for the application of next-generation sequencing to complex diseases, we conducted a pilot study sequencing the coding area of 186 hemostatic/proinflammatory genes in 10 Italian cases of idiopathic DVT and 12 healthy controls. Results A molecular-barcoding strategy was used to multiplex DNA target capture and sequencing, while retaining individual sequence information. Genomic libraries with barcode sequence-tags were pooled (in pools of 8 or 16 samples) and enriched for target DNA sequences. Sequencing was performed on ABI SOLiD-4 platforms. We produced > 12 gigabases of raw sequence data to sequence at high coverage (average: 42X) the 700-kilobase target area in 22 individuals. A total of 1876 high-quality genetic variants were identified (1778 single nucleotide substitutions and 98 insertions/deletions). Annotation on databases of genetic variation and human disease mutations revealed several novel, potentially deleterious mutations. We tested 576 common variants in a case-control association analysis, carrying the top-5 associations over to replication in up to 719 DVT cases and 719 controls. We also conducted an analysis of the burden of nonsynonymous variants in coagulation factor and anticoagulant genes. We found an excess of rare missense mutations in anticoagulant genes in DVT cases compared to controls and an association for a missense polymorphism of FGA (rs6050; p = 1.9 × 10-5, OR 1.45; 95% CI, 1.22-1.72; after replication in > 1400 individuals). Conclusions We implemented a barcode-based strategy to efficiently multiplex sequencing of hundreds of candidate genes in several individuals. In the relatively small dataset of our pilot study we were

  16. Identification of novel mutations and sequence variants in the SOX2 and CHX10 genes in patients with anophthalmia/microphthalmia.

    PubMed

    Zhou, Jie; Kherani, Femida; Bardakjian, Tanya M; Katowitz, James; Hughes, Nkecha; Schimmenti, Lisa A; Schneider, Adele; Young, Terri L

    2008-03-24

    Mutations in the SOX2 and CHX10 genes have been reported in patients with anophthalmia and/or microphthalmia. In this study, we evaluated 34 anophthalmic/microphthalmic patient DNA samples (two sets of siblings included) for mutations and sequence variants in SOX2 and CHX10. Conformational sensitive gel electrophoresis (CSGE) was used for the initial SOX2 and CHX10 screening of 34 affected individuals (two sets of siblings), five unaffected family members, and 80 healthy controls. Patient samples containing heteroduplexes were selected for sequence analysis. Base pair changes in SOX2 and CHX10 were confirmed by sequencing bidirectionally in patient samples. Two novel heterozygous mutations and two sequence variants (one known) in SOX2 were identified in this cohort. Mutation c.310 G>T (p. Glu104X), found in one patient, was in the region encoding the high mobility group (HMG) DNA-binding domain and resulted in a change from glutamic acid to a stop codon. The second mutation, noted in two affected siblings, was a single nucleotide deletion c.549delC (p. Pro184ArgfsX19) in the region encoding the activation domain, resulting in a frameshift and premature termination of the coding sequence. The shortened protein products may result in the loss of function. In addition, a novel nucleotide substitution c.*557G>A was identified in the 3'-untranslated region in one patient. The relationship between the nucleotide change and the protein function is indeterminate. A known single nucleotide polymorphism (c. *469 C>A, SNP rs11915160) was also detected in 2 of the 34 patients. Screening of CHX10 identified two synonymous sequence variants, c.471 C>T (p.Ser157Ser, rs35435463) and c.579 G>A (p. Gln193Gln, novel SNP), and one non-synonymous sequence variant, c.871 G>A (p. Asp291Asn, novel SNP). The non-synonymous polymorphism was also present in healthy controls, suggesting non-causality. These results support the role of SOX2 in ocular development. Loss of SOX2 function results

  17. Anatomy of a Red Copper Center: Spectroscopic Identification and Reactivity of the Copper Centers of Bacillus Subtilis Sco and its Cys to Ala Variants

    PubMed Central

    Siluvai, Gnana S; Mayfield, Mary; Nilges, Mark J; George, Serena DeBeer; Blackburn, Ninian J

    2010-01-01

    Sco is a mononuclear red copper protein involved in the assembly of cytochrome c oxidase. It is spectroscopically similar to red copper nitrosocyanin, but unlike the latter, which has one copper cysteine thiolate, the former has two. In addition to the two cysteine ligands (C45 and C49), the WT protein from Bacillus subtilis (hereafter named BSco) has a histidine (H135) and an unknown endogenous protein oxygen ligand in a distorted tetragonal array. We have compared the properties of the WT protein to variants in which each of the two coordinating Cys residues has been individually mutated to Ala, using UV/vis, Cu and S K edge XAS, EPR, and resonance Raman spectroscopy. Unlike the Cu(II) form of native Sco, the Cu(II) complexes of the Cys variants are unstable. The copper center of C49A undergoes autoreduction to the Cu(I) form which is shown by EXAFS to be composed of a novel 2-coordinate center with one Cys and one His ligand. C45A rearranges to a new stable Cu(II) species coordinated by C49 H135 and a second His ligand recruited from a previously uncoordinated protein side chain. The different chemistry exhibited by the Cys variants can be rationalized by whether a stable Cu(I) species can be formed by autoredox chemistry. For C49A, the remaining Cys and His residues are trans which facilitate the formation of the highly stable 2-coordinate Cu(I) species, while for C45A such a configuration cannot be attained. Resonance Raman spectroscopy of the WT protein indicates a net weak Cu–S bond strength at ~ 2.24 Å corresponding to the two thiolate copper bonds, whereas the single variant C45A shows a moderately strong Cu–S bond at ~ 2.16 Å. S K-edge data gives a total covalency of 28% for both Cu-S bonds in the WT protein. These data suggest an average covalency per Cu-S bond lower than nitrosocyanin and close to that expected for type-2 Cu(II)-thiolate systems. The data are discussed relative to the unique Cu-S characteristics of cupredoxins, whence it is

  18. Identification of Protective Epitopes by Sequencing of the Major Outer Membrane Protein Gene of a Variant Strain of Chlamydia psittaci Serotype 1 (Chlamydophila abortus)

    PubMed Central

    Vretou, Evangelia; Psarrou, Evgenia; Kaisar, Maria; Vlisidou, Isabella; Salti-Montesanto, Viviane; Longbottom, David

    2001-01-01

    Protective monoclonal antibodies (MAbs) to the major outer membrane protein (MOMP) of species of the family Chlamydiaceae, which is the primary vaccine candidate antigen, recognize nonlinear epitopes conferred by the oligomeric conformation of the molecule. Protective MAbs failed to recognize oligomeric MOMP of the variant strain LLG, which bears amino acid substitutions in variable segments (VSs) 1, 2, and 4, and competed with monomer-specific MAbs mapping to these VSs in reference strain 577. The results suggest that multiple sites located in the three VSs contribute to the epitope of protective MAbs. PMID:11119563

  19. Antisolvent crystallization of pharmaceutical excipients from aqueous solutions and the use of preferred orientation in phase identification by powder X-ray diffraction.

    PubMed

    Crisp, J L; Dann, S E; Blatchford, C G

    2011-04-18

    Crystallization of lactose from 10% (w/v) aqueous solutions was investigated with the use of polar antisolvents. Crystal growth was observed at 50-65% antisolvent content and showed a morphological transition from a polyhedral to needle-like habit with increasing antisolvent content, which coincided with a polymorphic transition from alpha lactose monohydrate (Lα·H(2)O) to beta lactose (Lβ). Where dehydrating antisolvents were employed such as methanol and ethanol, evidence of Lα·H(2)O dehydration to form Lα(S) was also observed at 95% antisolvent content. Powder X-ray diffraction (PXRD) analysis of the crystals highlighted the preferred orientation effects exhibited by large crystals of this kind, indicating the difficulties experienced by the non-specialist when performing phase identification of lactose polymorphs. The same studies were applied to raffinose pentahydrate, trehalose dihydrate and mannitol to assess the effects of crystallization conditions on other pharmaceutical excipients. Copyright © 2011 Elsevier B.V. All rights reserved.

  20. Molecular and biochemical identification of inositol 1,3,4,5,6-pentakisphosphate 2-kinase encoding mRNA variants in castor bean (Ricinus communis L.) seeds.

    PubMed

    Yu, Jaeju; Saiardi, Adolfo; Greenwood, John S; Bewley, J Derek

    2014-05-01

    During seed development, phytic acid (PA) associated with mineral cations is stored as phytin and mobilized following germination in support of seedling growth. Two parallel biosynthetic pathways for PA have been proposed; yet the pathway is still poorly understood in terms of its regulation and the enzymes involved. Here, the castor bean (Ricinus communis L.) gene for inositol 1,3,4,5,6-pentakisphosphate 2-kinase (RcIPK1) has been identified. This encodes the enzyme implicated in catalyzing the final reaction in PA biosynthesis, and its expression is enhanced in isolated germinated embryos by application of phosphate and myo-inositol (Ins). Even though only one copy of the RcIPK1 gene is present in the genome, numerous RNA variants are present, most likely due to alternative splicing. These are translated into six closely related protein isoforms according to in silico analysis. Functional analyses using yeast ipk1Δ revealed that only three of the mRNA variants can rescue a temperature-sensitive growth phenotype of this strain. High-performance liquid chromatography (HPLC) analysis of the synthesized inositol phosphates demonstrated that the ability to complement the missing yeast IPK1 enzyme is associated with the production of enzyme activity. The three active isoforms possess unique conserved motifs important for IPK1 catalytic activity.

  1. Identification of gene variants in NOS3, ET-1 and RAS that confer risk and protection against microangiopathy in type 2 diabetic obese subjects.

    PubMed

    Manea, Simona-Adriana; Robciuc, Alexandra; Guja, Cristian; Heltianu, Constantina

    2011-04-15

    The study aim was to investigate NOS3 VNTR, NOS3 G894T, EDN1 C8002T, ACE I/D, AGT M235T and AGTR1 A1166C in nonobese and obese T2DM patients, and their interaction with the incidence of microangiopathy. T2DM subjects (n=250; 166 nonobese, and 84 obese) were genotyped for the gene variants by PCR/RFLP. The interaction of these polymorphisms with obesity and their contribution to microangiopathy were analyzed by multivariate regression analysis. A higher frequency of NOS3 4a allele was found in obese (P=0.027) vs. nonobese subjects. ACE D (P=0.009) and AGT 235T (P=0.026) alleles were associated with the reduced risk of diabetic nephropathy in nonobese and obese patients, respectively. In obese subjects, NOS3 4a (P=0.011) had a converse effect to NOS3 894T (P=0.043), and EDN1 8002T (P=0.035) on the prevalence of combined microangiopathy (neuropathy/retinopathy/nephropathy) vs. microangiopathy-negative subjects. The study indicates association of RAS variants with obesity and nephropathy, and an opposite effect of NOS3 VNTR and NOS3 G894T on the occurrence of combined microangiopathy. Copyright © 2011 Elsevier Inc. All rights reserved.

  2. Identification and characterization of a macrophage-tropic SIV envelope glycoprotein variant in blood from early infection in SIVmac251-infected macaques

    PubMed Central

    Yen, Po-Jen; Mefford, Megan E; Hoxie, James A.; Williams, Kenneth C.; Desrosiers, Ronald C.; Gabuzda, Dana

    2014-01-01

    Macrophages play an important role in HIV/SIV pathogenesis by serving as a reservoir for viral persistence in brain and other tissues. Infected macrophages have been detected in brain early after infection, but macrophage-tropic viruses are rarely isolated until late-stage infection. Little is known about early variants that establish persistent infection in brain. Here, we characterize a unique macrophage-tropic SIV envelope glycoprotein (Env) variant from two weeks post-infection in blood of an SIVmac251-infected macaque that is closely related to sequences in brain from animals with neurological disease. SIVmac251 clones expressing this Env are highly fusogenic, and replicate efficiently in T cells and macrophages. N173 and N481 were identified as novel determinants of macrophage tropism and neutralization sensitivity. These results imply that macrophage-tropic SIV capable of establishing viral reservoirs in brain can be present in blood during early infection. Furthermore, these SIVmac251 clones will be useful for studies on pathogenesis, eradication, and vaccines. PMID:24928039

  3. Identification and characterization of a macrophage-tropic SIV envelope glycoprotein variant in blood from early infection in SIVmac251-infected macaques.

    PubMed

    Yen, Po-Jen; Mefford, Megan E; Hoxie, James A; Williams, Kenneth C; Desrosiers, Ronald C; Gabuzda, Dana

    2014-06-01

    Macrophages play an important role in HIV/SIV pathogenesis by serving as a reservoir for viral persistence in brain and other tissues. Infected macrophages have been detected in brain early after infection, but macrophage-tropic viruses are rarely isolated until late-stage infection. Little is known about early variants that establish persistent infection in brain. Here, we characterize a unique macrophage-tropic SIV envelope glycoprotein (Env) variant from two weeks post-infection in blood of an SIVmac251-infected macaque that is closely related to sequences in brain from animals with neurological disease. SIVmac251 clones expressing this Env are highly fusogenic, and replicate efficiently in T cells and macrophages. N173 and N481 were identified as novel determinants of macrophage tropism and neutralization sensitivity. These results imply that macrophage-tropic SIV capable of establishing viral reservoirs in brain can be present in blood during early infection. Furthermore, these SIVmac251 clones will be useful for studies on pathogenesis, eradication, and vaccines.

  4. Fc gamma receptor IIIb polymorphism and systemic lupus erythematosus: association with disease susceptibility and identification of a novel FCGR3B*01 variant.

    PubMed

    Santos, V C; Grecco, M; Pereira, K M C; Terzian, C C N; Andrade, L E C; Silva, N P

    2016-10-01

    The objective of this study was to evaluate the association between Fc gamma receptor IIIb polymorphism and susceptibility to systemic lupus erythematosus and clinical traits of the disease. Genomic DNA was obtained from 303 consecutive systemic lupus erythematosus patients and 300 healthy blood donors from the southeastern region of Brazil. The polymorphic region of the FCGR3B gene was sequenced and the alleles FCGR3B*01, FCGR3B*02 and FCGR3B*03 were analyzed. The FCGR3B*01 allele was more frequent in systemic lupus erythematosus patients (43.1%) while the FCGR3B*02 allele prevailed among controls (63.7%) (P = 0.001). The FCGR3B*03 allele was found equally in both groups. The FCGR3B*01/*01 (20.7%) and FCGR3B*01/*02 (41.1%) genotypes were more frequent among systemic lupus erythematosus patients (P = 0.028 and P = 0.012, respectively) while the FCGR3B*02/*02 genotype was more frequent in controls (45.5%) (P < 0.001). One variant of the FCGR3B*01 allele previously described in Germany was found in only one control. A new variant of the FCGR3B*01 allele with two substitutions (A227G/G277A) was found in one control. Three variants of the FCGR3B*02 allele previously described in African-Americans, Brazilians, Chinese and Japanese were found in ten 10 patients and two controls. In addition, several single nucleotide polymorphisms at non-polymorphic positions were identified in both patients and controls. Susceptibility to systemic lupus erythematosus was associated with the FCGR3B*01 allele, as well as with the FCGR3B*01/*01 and FCGR3B*01/*02 genotypes. No association was found between FCGR3B genotypes and clinical manifestations, disease severity or the presence of autoantibodies. © The Author(s) 2016.

  5. Identification and functional evaluation of two STAT3 variants in grass carp: Implication for the existence of specific alternative splicing of STAT3 gene in teleost.

    PubMed

    Du, Linyong; Zhou, Hong; Qin, Lei; Wei, He; Zhang, Anying; Yang, Kun; Wang, Xinyan

    2017-11-01

    A STAT family member, STAT3, becomes activated as a DNA binding protein in response to cytokines and growth factors. In teleost, STAT3 cDNA has been cloned and identified in a few species, but only a single STAT3 transcript is revealed in these studies. In the present study, two variants of STAT3 gene generated by alternative splicing were isolated from grass carp and nominated as STAT3α1 and STAT3α2 based on the homology with their mammalian orthologs. In particular, the homologs of STAT3α1/2 were also found in various fish species, including zebrafish, takifugu, tilapia, medaka and goldfish. Intriguingly, sequence alignment and genomic structure analysis revealed that fish STAT3α1/2 are generated through similar alternative splicing events, implying the potential physiological significance of generating STAT3 variants in fish. Grass carp STAT3α1/2 (gcSTAT3α1/2) were ubiquitously expressed although the transcript levels of STAT3α2 were markedly higher than STAT3α1 in all examined tissues. In vivo and in vitro studies showed that the expression patterns of these two variants were similar under the stimulation of immune stimuli. To reveal the role of gcSTAT3α1/2 in fish immunity, their phosphorylation and involvement in IL-17A/F1 mRNA expression were demonstrated in grass carp peripheral blood lymphocytes upon LPS or PHA challenge, providing evidence for the functional conservation of STAT3 signaling in fish. These findings also raise a question of whether both gcSTAT3α1/2 participate in transcriptional regulation in fish. Actually, our results showed that both of them had the ability to translocate into the nucleus upon activation, and to amplify IL-10 signaling, indicating the existence of STAT3 isoforms with functional redundancy in teleost. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Hue-shifted monomeric variants of Clavularia cyan fluorescent protein: identification of the molecular determinants of color and applications in fluorescence imaging.

    PubMed

    Ai, Hui-wang; Olenych, Scott G; Wong, Peter; Davidson, Michael W; Campbell, Robert E

    2008-03-06

    In the 15 years that have passed since the cloning of Aequorea victoria green fluorescent protein (avGFP), the expanding set of fluorescent protein (FP) variants has become entrenched as an indispensable toolkit for cell biology research. One of the latest additions to the toolkit is monomeric teal FP (mTFP1), a bright and photostable FP derived from Clavularia cyan FP. To gain insight into the molecular basis for the blue-shifted fluorescence emission we undertook a mutagenesis-based study of residues in the immediate environment of the chromophore. We also employed site-directed and random mutagenesis in combination with library screening to create new hues of mTFP1-derived variants with wavelength-shifted excitation and emission spectra. Our results demonstrate that the protein-chromophore interactions responsible for blue-shifting the absorbance and emission maxima of mTFP1 operate independently of the chromophore structure. This conclusion is supported by the observation that the Tyr67Trp and Tyr67His mutants of mTFP1 retain a blue-shifted fluorescence emission relative to their avGFP counterparts (that is, Tyr66Trp and Tyr66His). Based on previous work with close homologs, His197 and His163 are likely to be the residues with the greatest contribution towards blue-shifting the fluorescence emission. Indeed we have identified the substitutions His163Met and Thr73Ala that abolish or disrupt the interactions of these residues with the chromophore. The mTFP1-Thr73Ala/His163Met double mutant has an emission peak that is 23 nm red-shifted from that of mTFP1 itself. Directed evolution of this double mutant resulted in the development of mWasabi, a new green fluorescing protein that offers certain advantages over enhanced avGFP (EGFP). To assess the usefulness of mTFP1 and mWasabi in live cell imaging applications, we constructed and imaged more than 20 different fusion proteins. Based on the results of our mutagenesis study, we conclude that the two histidine

  7. Hue-shifted monomeric variants of Clavularia cyan fluorescent protein: identification of the molecular determinants of color and applications in fluorescence imaging

    PubMed Central

    Ai, Hui-wang; Olenych, Scott G; Wong, Peter; Davidson, Michael W; Campbell, Robert E

    2008-01-01

    Background In the 15 years that have passed since the cloning of Aequorea victoria green fluorescent protein (avGFP), the expanding set of fluorescent protein (FP) variants has become entrenched as an indispensable toolkit for cell biology research. One of the latest additions to the toolkit is monomeric teal FP (mTFP1), a bright and photostable FP derived from Clavularia cyan FP. To gain insight into the molecular basis for the blue-shifted fluorescence emission we undertook a mutagenesis-based study of residues in the immediate environment of the chromophore. We also employed site-directed and random mutagenesis in combination with library screening to create new hues of mTFP1-derived variants with wavelength-shifted excitation and emission spectra. Results Our results demonstrate that the protein-chromophore interactions responsible for blue-shifting the absorbance and emission maxima of mTFP1 operate independently of the chromophore structure. This conclusion is supported by the observation that the Tyr67Trp and Tyr67His mutants of mTFP1 retain a blue-shifted fluorescence emission relative to their avGFP counterparts (that is, Tyr66Trp and Tyr66His). Based on previous work with close homologs, His197 and His163 are likely to be the residues with the greatest contribution towards blue-shifting the fluorescence emission. Indeed we have identified the substitutions His163Met and Thr73Ala that abolish or disrupt the interactions of these residues with the chromophore. The mTFP1-Thr73Ala/His163Met double mutant has an emission peak that is 23 nm red-shifted from that of mTFP1 itself. Directed evolution of this double mutant resulted in the development of mWasabi, a new green fluorescing protein that offers certain advantages over enhanced avGFP (EGFP). To assess the usefulness of mTFP1 and mWasabi in live cell imaging applications, we constructed and imaged more than 20 different fusion proteins. Conclusion Based on the results of our mutagenesis study, we conclude

  8. Molecular epidemiology of Theileria annulata and identification of 18S rRNA gene and ITS regions sequences variants in apparently healthy buffaloes and cattle in Pakistan.

    PubMed

    Khan, Muhammad Kasib; He, Lan; Hussain, Altaf; Azam, Sabita; Zhang, Wen-Jie; Wang, Li-Xia; Zhang, Qing-Li; Hu, Min; Zhou, Yan-Qin; Zhao, Junlong

    2013-01-01

    A molecular epidemiological survey was conducted to determine the prevalence of piroplasms in buffaloes and cattle from Sheikhupura and Okara districts of Punjab, Pakistan using reverse line blot (RLB) hybridization assay. The genetic diversity within 18S rRNA gene and ITS regions sequences of various obtained Theileria species (spp.) was also investigated. Briefly, 102 blood samples from buffaloes and cattle in the study districts were collected on blood collection cards and brought to the laboratory. DNA was extracted; the V4 hypervariable region of 18S rRNA was amplified and analyzed using RLB. Out of total samples analyzed, 61 (59.8%) were hybridized with Babesia/Theileria (B/T) genus-specific probe. Only one species of piroplasm was detected in buffaloes and cattle in study districts, i.e. Theileria (T.) annulata. Six samples only hybridized with B/T genus-specific and Theileria genus-specific probes but not with any species-specific probe indicating the presence of novel species or variants. The sequences of 18S rRNA gene and ITS regions of these six samples revealed the presence of T. annulata variants as confirmed through sequence identity estimation and phylogenetic analyses. Meanwhile, an unexpected sequence variation was observed within the 18S rRNA gene and ITS regions sequences of T. annulata identified in the present study. This is the first report on the simultaneous detection of species of piroplasms infecting buffaloes and cattle in Pakistan and molecular characterization of T. annulata 18S rRNA gene and ITS regions. The present study may address the new insights into the epidemiology of theileriosis which will help researches in designing control strategies and developing various molecular diagnostic tools at national level. Copyright © 2012 Elsevier B.V. All rights reserved.

  9. Functional Genomic mRNA Profiling of Colorectal Adenomas: Identification and in vivo Validation of CD44 and Splice Variant CD44v6 as Molecular Imaging Targets.

    PubMed

    Hartmans, Elmire; Orian-Rousseau, Veronique; Matzke-Ogi, Alexandra; Karrenbeld, Arend; de Groot, Derk Jan A; de Jong, Steven; van Dam, Gooitzen M; Fehrmann, Rudolf S N; Nagengast, Wouter B

    2017-01-01

    Colorectal cancer (CRC) is the third leading cause of cancer-related deaths worldwide. High adenoma miss rates, especially seen in high-risk patients, demand for better endoscopic detection. By fluorescently 'highlighting' specific molecular characteristics, endoscopic molecular imaging has great potential to fulfill this need. To implement this technique effectively, target proteins that distinguish adenomas from normal tissue must be identified. In this study we applied in silico Functional Genomic mRNA (FGmRNA) profiling, which is a recently developed method that results in an enhanced view on the downstream effects of genomic alterations occurring in adenomas on gene expression levels. FGmRNA profiles of sporadic adenomas were compared to normal colon tissue to identify overexpressed genes. We validated the protein expression of the top identified genes, AXIN2, CEMIP, CD44 and JUN, in sporadic adenoma patient samples via immunohistochemistry (IHC). CD44 was identified as the most attractive target protein for imaging purposes and we proved its relevance in high-risk patients by demonstrating CD44 protein overexpression in Lynch lesions. Subsequently, we show that the epithelial splice variant CD44V6 is highly overexpressed in our patient samples and we demonstrated the feasibility of visualizing adenomas in Apc(Min/+) mice in vivo by using a fluorescently labeled CD44v6 targeting peptide. In conclusion, via in silico functional genomics and ex vivo protein validation, this study identified CD44 as an attractive molecular target for both sporadic and high-risk Lynch adenomas, and demonstrates the in vivo applicability of a small peptide drug directed against splice variant CD44v6 for adenoma imaging.

  10. Metallo-β-Lactamases in Clinical Pseudomonas Isolates in Taiwan and Identification of VIM-3, a Novel Variant of the VIM-2 Enzyme

    PubMed Central

    Yan, Jing-Jou; Hsueh, Po-Ren; Ko, Wen-Chien; Luh, Kwen-Tay; Tsai, Shu-Huei; Wu, Hsiu-Mei; Wu, Jiunn-Jong

    2001-01-01

    A total of 209 clinical isolates of Pseudomonas (193 Pseudomonas aeruginosa, 10 P. putida, 4 P. stutzeri, and 2 P. fluorescens isolates) with reduced susceptibilities to imipenem and/or ceftazidime were subjected to PCR assays with primers specific for blaIMP-1, blaIMP-2, blaVIM-1, and blaVIM-2 and sequence analysis to identify the metallo-β-lactamases (MBLs) prevalent among these organisms in Taiwan; and 21 isolates gave positive results. Five isolates including two P. putida and three P. stutzeri isolates were found to carry blaIMP-1, and six isolates including five P. putida and one P. stutzeri isolates harbored blaVIM-2. The remaining 10 isolates were P. aeruginosa, and all were found to carry a novel variant of blaVIM-2, designated blaVIM-3. There are only two nucleotide differences between blaVIM-2 and blaVIM-3, leading to two amino acid alterations. Our findings indicate that VIM-2 and its variant have become the most prevalent metalloenzymes in Pseudomonas in Taiwan. Southern hybridization with the blaVIM-2-, blaVIM-3-, and blaIMP-1 -specific probes revealed that only two VIM-2-producing P. putida isolates appeared to carry the MBL gene on plasmids. Pulsed-field gel electrophoresis showed that six VIM-3-producing P. aeruginosa isolates and two IMP-1-producing P. stutzeri isolates were genetically related, suggesting that the spread of these MBL genes in Taiwan could be due to clonal dissemination as well as genetic exchange between different clones. PMID:11451678

  11. Identification of a truncated splice variant of IL-18 receptor alpha in the human and rat, with evidence of wider evolutionary conservation

    PubMed Central

    Grattan, David R.

    2014-01-01

    Interleukin-18 (IL-18) is a pro-inflammatory cytokine which stimulates activation of the nuclear factor kappa beta (NF-κB) pathway via interaction with the IL-18 receptor. The receptor itself is formed from a dimer of two subunits, with the ligand-binding IL-18Rα subunit being encoded by the IL18R1 gene. A splice variant of murine IL18r1, which has been previously described, is formed by transcription of an unspliced intron (forming a ‘type II’ IL18r1 transcript) and is predicted to encode a receptor with a truncated intracellular domain lacking the capacity to generate downstream signalling. In order to examine the relevance of this finding to human IL-18 function, we assessed the presence of a homologous transcript by reverse transcription-polymerase chain reaction (RT-PCR) in the human and rat as another common laboratory animal. We present evidence for type II IL18R1 transcripts in both species. While the mouse and rat transcripts are predicted to encode a truncated receptor with a novel 5 amino acid C-terminal domain, the human sequence is predicted to encode a truncated protein with a novel 22 amino acid sequence bearing resemblance to the ‘Box 1’ motif of the Toll/interleukin-1 receptor (TIR) domain, in a similar fashion to the inhibitory interleukin-1 receptor 2. Given that transcripts from these three species are all formed by inclusion of homologous unspliced intronic regions, an analysis of homologous introns across a wider array of 33 species with available IL18R1 gene records was performed, which suggests similar transcripts may encode truncated type II IL-18Rα subunits in other species. This splice variant may represent a conserved evolutionary mechanism for regulating IL-18 activity. PMID:25250214

  12. Isolation of a hop-sensitive variant of Lactobacillus lindneri and identification of genetic markers for beer spoilage ability of lactic acid bacteria.

    PubMed

    Suzuki, Koji; Iijima, Kazumaru; Ozaki, Kazutaka; Yamashita, Hiroshi

    2005-09-01

    We have isolated a hop-sensitive variant of the beer spoilage bacterium Lactobacillus lindneri DSM 20692. The variant lost a plasmid carrying two contiguous open reading frames (ORF s) designated horB(L) and horC(L) that encode a putative regulator and multidrug transporter presumably belonging to the resistance-nodulation-cell division superfamily. The loss of hop resistance ability occurred with the loss of resistance to other drugs, including ethidium bromide, novobiocin, and cetyltrimethylammonium bromide. PCR and Southern blot analysis using 51 beer spoilage strains of various species of lactic acid bacteria (LAB) revealed that 49 strains possessed homologs of horB and horC. No false-positive results have been observed for nonspoilage LAB or frequently encountered brewery isolates. These features are superior to those of horA and ORF 5, previously reported genetic markers for determining the beer spoilage ability of LAB. It was further shown that the combined use of horB/horC and horA is able to detect all 51 beer spoilage strains examined in this study. Furthermore sequence comparison of horB and horC homologs identified in four different beer spoilage species indicates these homologs are 96.6 to 99.5% identical, which is not typical of distinct species. The wide and exclusive distribution of horB and horC homologs among beer spoilage LAB and their sequence identities suggest that the hop resistance ability of beer spoilage LAB has been acquired through horizontal gene transfer. These insights provide a foundation for applying trans-species genetic markers to differentiating beer spoilage LAB including previously unencountered species.

  13. Characterization of high-molecular-weight glutenin subunits from Eremopyrum bonaepartis and identification of a novel variant with unusual high molecular weight and altered cysteine residues.

    PubMed

    Jiang, Qian-Tao; Zhang, Xiao-Wei; Ma, Jian; Wei, Long; Zhao, Shan; Zhao, Quan-Zhi; Qi, Peng-Fei; Lu, Zhen-Xiang; Zheng, You-Liang; Wei, Yu-Ming

    2014-04-01

    We characterized two high-molecular-weight glutenin subunit (HMW-GS) variants from Eremopyrum bonaepartis, determined their complete open reading frames, and further expressed them in a bacterial system. The variants have many novel structural features compared with typical subunits encoded by Glu-1 loci: 1Fx3.7 and 1Fy1.5 exhibit hybrid properties of x- and y-type subunits. In addition, unusual molecular mass and altered number and distribution of cysteine residues were unique features of HMW-GSs encoded by Glu-F1 from E. bonaepartis. The mature 1Fx3.7 subunit has a full length of 1,223 amino acid residues, making it the largest subunit found thus far, while 1Fy1.5 is just 496 residues. In addition, the mutated PGQQ repeat motif was found in the repetitive region of 1Fx3.7. Although it has a similar molecular mass to that previously reported for 1Dx2.2, 1Dx2.2* and 1S(sh)x2.9 subunits, 1Fx3.7 appears to have had a different evolutionary history. The N-terminal and repetitive regions have a total of four additional cysteine residues, giving 1Fx3.7 a total of eight cysteines, while 1Fy1.5 has only six cysteines because the GHCPTSPQQ nonapeptide at the end of the repetitive region is deleted. With its extra cysteine residues and the longest repetitive region, features that are relevant to good wheat quality, the 1Fx3.7 subunit gene could be an excellent candidate for applications in wheat quality improvement.

  14. Use of heteroduplex mobility assays (HMA) for pre-sequencing screening and identification of variant strains of swine and avian hepatitis E viruses.

    PubMed

    Sun, Z F; Huang, F F; Halbur, P G; Schommer, S K; Pierson, F W; Toth, T E; Meng, X J

    2003-10-17

    Hepatitis E virus (HEV), the causative agent of human hepatitis E, is an important public health problem in many developing countries and is also endemic in many industrialized countries including the US. The discoveries of avian and swine HEVs by our group from chickens and pigs, respectively, suggest that hepatitis E may be a zoonosis. Current methods for molecular epidemiological studies of HEV require PCR amplification of field strains of HEV followed by DNA sequencing and sequence analyses, which are laborious and expensive. As novel or variant strains of HEV continue to evolve rapidly both in humans and other animals, it is important to develop a rapid pre-sequencing screening method to select field isolates for further molecular characterization. In this study, we developed two heteroduplex mobility assays (HMA) (one for swine HEV based on the ORF2 region, and the other for avian HEV based on the ORF1 region) to genetically differentiate field strains of avian and swine HEVs from known reference strains. The ORF2 regions of 22 swine HEV isolates and the ORF1 regions of 13 avian HEV isolates were amplified by PCR, sequenced and analyzed by HMA against reference prototype swine HEV strain and reference prototype avian HEV strain, respectively. We showed that, in general, the HMA profiles correlate well with nucleotide sequence identities and with phylogenetic clustering between field strains and the reference swine HEV or avian HEV strains. Field isolates with similar HMA patterns generally showed similar sequence identities with the reference strains and clustered together in the phylogenetic trees. Therefore, by using different HEV isolates as references, the HMA developed in this study can be used as a pre-sequencing screening tool to identify variant HEV isolates for further molecular epidemiological studies.

  15. Isolation of a Hop-Sensitive Variant of Lactobacillus lindneri and Identification of Genetic Markers for Beer Spoilage Ability of Lactic Acid Bacteria

    PubMed Central

    Suzuki, Koji; Iijima, Kazumaru; Ozaki, Kazutaka; Yamashita, Hiroshi

    2005-01-01

    We have isolated a hop-sensitive variant of the beer spoilage bacterium Lactobacillus lindneri DSM 20692. The variant lost a plasmid carrying two contiguous open reading frames (ORF s) designated horBL and horCL that encode a putative regulator and multidrug transporter presumably belonging to the resistance-nodulation-cell division superfamily. The loss of hop resistance ability occurred with the loss of resistance to other drugs, including ethidium bromide, novobiocin, and cetyltrimethylammonium bromide. PCR and Southern blot analysis using 51 beer spoilage strains of various species of lactic acid bacteria (LAB) revealed that 49 strains possessed homologs of horB and horC. No false-positive results have been observed for nonspoilage LAB or frequently encountered brewery isolates. These features are superior to those of horA and ORF 5, previously reported genetic markers for determining the beer spoilage ability of LAB. It was further shown that the combined use of horB/horC and horA is able to detect all 51 beer spoilage strains examined in this study. Furthermore sequence comparison of horB and horC homologs identified in four different beer spoilage species indicates these homologs are 96.6 to 99.5% identical, which is not typical of distinct species. The wide and exclusive distribution of horB and horC homologs among beer spoilage LAB and their sequence identities suggest that the hop resistance ability of beer spoilage LAB has been acquired through horizontal gene transfer. These insights provide a foundation for applying trans-species genetic markers to differentiating beer spoilage LAB including previously unencountered species. PMID:16151091

  16. Prevalence and genetic diversity of arginine catabolic mobile element (ACME) in clinical isolates of coagulase-negative staphylococci: identification of ACME type I variants in Staphylococcus epidermidis.

    PubMed

    Onishi, Mayumi; Urushibara, Noriko; Kawaguchiya, Mitsuyo; Ghosh, Souvik; Shinagawa, Masaaki; Watanabe, Naoki; Kobayashi, Nobumichi

    2013-12-01

    Arginine catabolic mobile element (ACME), a genomic island consisting of the arc and/or opp3 gene clusters found in staphylococcal species, is related to increased bacterial adaptability to hosts. Staphylococcus epidermidis is considered a major ACME reservoir; however, prevalence and genetic diversity of ACME in coagulase-negative staphylococci (CNS) have not yet been well characterized for clinical isolates in Japan. A total of 271 clinical isolates of CNS in a Japanese hospital were investigated for the presence and genotype of ACME and SCCmec. The prevalence of ACME-arcA was significantly higher (p<0.001) in S. epidermidis (45.8%) than in other CNS species (3.7%). ACME in S. epidermidis isolates (n=87) were differentiated into type I (n=33), variant forms of type I (ΔI, n=26) newly identified in this study, type II (n=6), and type ΔII (n=19). ACME-type ΔI, which were further classified into three subtypes, lacked some genetic components between the arc and opp3 clusters in archetypal type I, whereas the arc and opp3 clusters were intact. The arc cluster exhibited high sequence identity (95.8-100%) to that of type I ACME; in contrast, the opp3 cluster was highly diverse, and showed relatively lower identities (94.8-98.7%) to the identical regions in type I ACME. Twenty-one isolates of ΔI ACME-carrying S. epidermidis possessed SCCmec IVa and belonged to ST5 (clonal complex 2). Phylogenetic analysis revealed that isolates harboring ACME ΔI in this study clustered with previously reported S. epidermidis strains with other lineges, suggesting that S. epidermidis originally had some genetic variations in the opp3 cluster. In summary, ACME type ΔI, a truncated variant of ACME-I, was first identified in S. epidermidis, and revealed to be prevalent in ST5 MRSE clinical isolates with SCCmec IVa.

  17. Identification of a regulatory variant that binds FOXA1 and FOXA2 at the CDC123/CAMK1D type 2 diabetes GWAS locus.

    PubMed

    Fogarty, Marie P; Cannon, Maren E; Vadlamudi, Swarooparani; Gaulton, Kyle J; Mohlke, Karen L

    2014-09-01

    Many of the type 2 diabetes loci identified through genome-wide association studies localize to non-protein-coding intronic and intergenic regions and likely contain variants that regulate gene transcription. The CDC123/CAMK1D type 2 diabetes association signal on chromosome 10 spans an intergenic region between CDC123 and CAMK1D and also overlaps the CDC123 3'UTR. To gain insight into the molecular mechanisms underlying the association signal, we used open chromatin, histone modifications and transcription factor ChIP-seq data sets from type 2 diabetes-relevant cell types to identify SNPs overlapping predicted regulatory regions. Two regions containing type 2 diabetes-associated variants were tested for enhancer activity using luciferase reporter assays. One SNP, rs11257655, displayed allelic differences in transcriptional enhancer activity in 832/13 and MIN6 insulinoma cells as well as in human HepG2 hepatocellular carcinoma cells. The rs11257655 risk allele T showed greater transcriptional activity than the non-risk allele C in all cell types tested. Using electromobility shift and supershift assays we demonstrated that the rs11257655 risk allele showed allele-specific binding to FOXA1 and FOXA2. We validated FOXA1 and FOXA2 enrichment at the rs11257655 risk allele using allele-specific ChIP in human islets. These results suggest that rs11257655 affects transcriptional activity through altered binding of a protein complex that includes FOXA1 and FOXA2, providing a potential molecular mechanism at this GWAS locus.

  18. Functional Genomic mRNA Profiling of Colorectal Adenomas: Identification and in vivo Validation of CD44 and Splice Variant CD44v6 as Molecular Imaging Targets

    PubMed Central

    Hartmans, Elmire; Orian-Rousseau, Veronique; Matzke-Ogi, Alexandra; Karrenbeld, Arend; de Groot, Derk Jan A.; de Jong, Steven; van Dam, Gooitzen M.; Fehrmann, Rudolf S.N.; Nagengast, Wouter B.

    2017-01-01

    Colorectal cancer (CRC) is the third leading cause of cancer-related deaths worldwide. High adenoma miss rates, especially seen in high-risk patients, demand for better endoscopic detection. By fluorescently 'highlighting' specific molecular characteristics, endoscopic molecular imaging has great potential to fulfill this need. To implement this technique effectively, target proteins that distinguish adenomas from normal tissue must be identified. In this study we applied in silico Functional Genomic mRNA (FGmRNA) profiling, which is a recently developed method that results in an enhanced view on the downstream effects of genomic alterations occurring in adenomas on gene expression levels. FGmRNA profiles of sporadic adenomas were compared to normal colon tissue to identify overexpressed genes. We validated the protein expression of the top identified genes, AXIN2, CEMIP, CD44 and JUN, in sporadic adenoma patient samples via immunohistochemistry (IHC). CD44 was identified as the most attractive target protein for imaging purposes and we proved its relevance in high-risk patients by demonstrating CD44 protein overexpression in Lynch lesions. Subsequently, we show that the epithelial splice variant CD44V6 is highly overexpressed in our patient samples and we demonstrated the feasibility of visualizing adenomas in ApcMin/+ mice in vivo by using a fluorescently labeled CD44v6 targeting peptide. In conclusion, via in silico functional genomics and ex vivo protein validation, this study identified CD44 as an attractive molecular target for both sporadic and high-risk Lynch adenomas, and demonstrates the in vivo applicability of a small peptide drug directed against splice variant CD44v6 for adenoma imaging. PMID:28255344

  19. Ultrasonographic imaging of papillary thyroid carcinoma variants

    PubMed Central

    2017-01-01

    Ultrasonography (US) is routinely used to evaluate thyroid nodules. The US features of papillary thyroid carcinoma (PTC), the most common thyroid malignancy, include hypoechogenicity, spiculated/microlobulated margins, microcalcifications, and a nonparallel orientation. However, many PTC variants have been identified, some of which differ from the classic type of PTC in terms of biological behavior and clinical outcomes. This review describes the US features and clinical implications of the variants of PTC. With the introduction of active surveillance replacing immediate biopsy or surgical treatment of indolent, small PTCs, an understanding of the US characteristics of PTC variants will facilitate the individualized management of patients with PTC. PMID:28222584

  20. Ultrasonographic imaging of papillary thyroid carcinoma variants.

    PubMed

    Shin, Jung Hee

    2017-04-01

    Ultrasonography (US) is routinely used to evaluate thyroid nodules. The US features of papillary thyroid carcinoma (PTC), the most common thyroid malignancy, include hypoechogenicity, spiculated/microlobulated margins, microcalcifications, and a nonparallel orientation. However, many PTC variants have been identified, some of which differ from the classic type of PTC in terms of biological behavior and clinical outcomes. This review describes the US features and clinical implications of the variants of PTC. With the introduction of active surveillance replacing immediate biopsy or surgical treatment of indolent, small PTCs, an understanding of the US characteristics of PTC variants will facilitate the individualized management of patients with PTC.

  1. Transcriptomics and Identification of the Chemoreceptor Superfamily of the Pupal Parasitoid of the Oriental Fruit Fly, Spalangia endius Walker (Hymenoptera: Pteromalidae)

    PubMed Central

    Zhang, Yuping; Zheng, Yuan; Li, Dunsong; Fan, Yilin

    2014-01-01

    Background The oriental fruit fly, Bactrocera dorsalis Hendel, causes serious losses to fruit production and is one of the most economically important pests in many countries, including China, Spalangia endius Walker is a pupal parasitoid of various dipteran hosts, and may be considered a potentially important ectoparasitic pupal parasitoid of B. dorsalis. However, lack of genetic information on this organism is an obstacle to understanding the mechanisms behind its interaction with this host. Analysis of the S. endius transcriptome is essential to extend the resources of genetic information on this species and, to support studies on S. endius on the host B. dorsalis. Methodology/Principal Findings We performed de novo assembly RNA-seq of S. endius. We obtained nearly 10 Gbp of data using a HiSeq platform, and 36319 high-quality transcripts using Trinity software. A total of 22443 (61.79%) unigenes were aligned to homologous sequences in the jewel wasp and honeybee (Apis florae) protein set from public databases. A total of 10037 protein domains were identified in 7892 S. endius transcripts using HMMER3 software. We identified expression of six gustatory receptor and 21 odorant receptor genes in the sample, with only one gene having a high expression level in each family. The other genes had a low expression level, including two genes regulated by splicing. This result may be due to the wasps being kept under laboratory conditions. Additionally, a total of 3727 SSR markers were predicted, which could facilitate the identification of polymorphisms and functional genes within wasp populations. Conclusion/Significance This transcriptome greatly improves our genetic understanding of S. endius and provides a large number of gene sequences for further study. PMID:24505315

  2. Identification and comparative analysis of the oriental river prawn (Macrobrachium nipponense) microRNA expression profile during hypoxia using a deep sequencing approach.

    PubMed

    Sun, Shengming; Fu, Hongtuo; Ge, Xianping; Zhu, Jian; Gu, Zhimin; Xuan, Fujun

    2016-03-01

    Hypoxia refers to a state of oxygen deficiency, which is observed frequently in aquaculture ponds. MicroRNAs (miRNAs) are small non-coding RNAs that are important effectors in regulating gene expression through posttranscriptional mechanisms. They are key elements in the response to hypoxia. The oriental river prawn (Macrobrachium nipponense) is an important commercial aquaculture species, and is sensitive to hypoxia. To date, there are no reports describing M. nipponense miRNAs. In this study, Solexa deep sequencing technology was used for high-throughput analysis of miRNAs in a small RNA library isolated from four M. nipponense tissues (gill, hepatopancreas, muscle and hemocytes). In total, 9,227,356 reads were obtained, 4,293,155 of which were related to 267 unique miRNAs, including 203 conserved and 64 prawn-specific miRNAs. Furthermore, miRNA features including length distribution and end variations were characterized. Annotation of targets revealed a broad range of biological processes and signal transduction pathways regulated by M. nipponense miRNAs. In addition, 880 co-expressed and 39 specific (25 normoxia-specific and 14 hypoxia-specific) miRNAs that may be involved in the response to hypoxia were confirmed using miRNA microarray analysis from the four prawn tissues combined. Real-time quantitative PCR (qPCR) analysis of eight miRNAs in the normoxia and hypoxia groups showed good concordance between the sequencing and qPCR data. This study provides the first large-scale identification and characterization of M. nipponense miRNAs and their potential targets, and represents a foundation for further characterization of their roles in the regulation of the diversity of hypoxia processes.

  3. Identification of Theileria parva and Theileria sp. (buffalo) 18S rRNA gene sequence variants in the African Buffalo (Syncerus caffer) in southern Africa.

    PubMed

    Chaisi, Mamohale E; Sibeko, Kgomotso P; Collins, Nicola E; Potgieter, Fred T; Oosthuizen, Marinda C

    2011-12-15

    distinction between Theileria sp. (buffalo) and T. parva and indicate the existence of a single group of T. parva and two Theileria sp. (buffalo) 18S rRNA gene variants in the African buffalo. Despite the observed variation in the full-length parasite 18S rRNA gene sequences, the area in the V4 hypervariable region where the RLB and real-time PCR hybridization probes were developed was relatively conserved. The T. parva specific real-time PCR assay was able to successfully detect all T. parva variants and, although amplicons were obtained from Theileria sp. (buffalo) DNA, none of the Theileria sp. (buffalo) 18S rRNA sequence variants were detected by the T. parva-specific hybridization probes.

  4. Molecular characterization and tissue distribution of aryl hydrocarbon receptor nuclear translocator isoforms, ARNT1 and ARNT2, and identification of novel splice variants in common cormorant (Phalacrocorax carbo).

    PubMed

    Lee, Jin-Seon; Kim, Eun-Young; Iwata, Hisato; Tanabe, Shinsuke

    2007-04-01

    High levels of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and related planar halogenated aromatic hydrocarbons (PHAHs) are accumulated in fish-eating birds including common cormorant (Phalacrocorax carbo). Most of the biochemical and toxic effects of TCDD are mediated by a basic helix-loop-helix and a conserved region among Per, ARNT, and Sim (bHLH/PAS) proteins, aryl hydrocarbon receptor (AHR) and AHR nuclear translocator (ARNT). To study the molecular mechanism of TCDD toxicity in common cormorant as an avian model species, characterization of the AHR/ARNT signaling pathway in this species is necessary. The present study focuses on molecular characterization of ARNT from common cormorant (ccARNT). The cDNA of the ccARNT isoform, ccARNT1 obtained by the screening of hepatic cDNA library contains a 2424-bp open reading frame that encodes 807 amino acids, exhibiting high identities (92%) with chicken ARNT. This isoform contains a unique 22 amino acid residue in 3' end of PAS A domain as is also recognized in chicken ARNT. The ccARNT2 cDNA isolated from brain tissue has a 2151-bp open reading frame. The deduced amino acid sequence of ccARNT2 protein (716 aa) shows a conservation of bHLH and PAS motif in its N-terminal region with high similarities (96% and 78%, respectively) to that of ccARNT1. Using quantitative RT-PCR methods, the tissue distribution profiles of ccARNT1 and ccARNT2 were unveiled. Both ccARNT1 and ccARNT2 mRNAs were ubiquitously expressed in all examined tissues including liver. The expression profile of ccARNT1 was comparable with that of rodent ARNT1, but ccARNT2 was not with rodent ARNT2, implying different roles of ARNT2 between the two species. There was a significant positive correlation between ARNT1 and ARNT2 mRNA expression levels in the liver of wild cormorant population, indicating that their expressions may be enforced by similar transcriptional regulation mechanism. Novel variants of ccARNT1 and ccARNT2 isoforms that were supposed to

  5. Identification of Emerging Macrophage-Tropic HIV-1 R5 Variants in Brain Tissue of AIDS Patients without Severe Neurological Complications.

    PubMed

    Gonzalez-Perez, Maria Paz; Peters, Paul J; O'Connell, Olivia; Silva, Nilsa; Harbison, Carole; Cummings Macri, Sheila; Kaliyaperumal, Saravanan; Luzuriaga, Katherine; Clapham, Paul R

    2017-10-15

    Untreated HIV-positive (HIV-1(+)) individuals frequently suffer from HIV-associated neurocognitive disorders (HAND), with about 30% of AIDS patients suffering severe HIV-associated dementias (HADs). Antiretroviral therapy has greatly reduced the incidence of HAND and HAD. However, there is a continuing problem of milder neurocognitive impairments in treated HIV(+) patients that may be increasing with long-term therapy. In the present study, we investigated whether envelope (env) genes could be amplified from proviral DNA or RNA derived from brain tissue of 12 individuals with normal neurology or minor neurological conditions (N/MC individuals). The tropism and characteristics of the brain-derived Envs were then investigated and compared to those of Envs derived from immune tissue. We showed that (i) macrophage-tropic R5 Envs could be detected in the brain tissue of 4/12 N/MC individuals, (ii) macrophage-tropic Envs in brain tissue formed compartmentalized clusters distinct from non-macrophage-tropic (non-mac-tropic) Envs recovered from the spleen or brain, (iii) the evidence was consistent with active viral expression by macrophage-tropic variants in the brain tissue of some individuals, and (iv) Envs from immune tissue of the N/MC individuals were nearly all tightly non-mac-tropic, contrasting with previous data for neuro-AIDS patients where immune tissue Envs mediated a range of macrophage infectivities, from background levels to modest infection, with a small number of Envs from some patients mediating high macrophage infection levels. In summary, the data presented here show that compartmentalized and active macrophage-tropic HIV-1 variants are present in the brain tissue of individuals before neurological disease becomes overt or serious.IMPORTANCE The detection of highly compartmentalized macrophage-tropic R5 Envs in the brain tissue of HIV patients without serious neurological disease is consistent with their emergence from a viral population already

  6. Identification and characterization of circulating variants of CXCL12 from human plasma: effects on chemotaxis and mobilization of hematopoietic stem and progenitor cells.

    PubMed

    Richter, Rudolf; Jochheim-Richter, Andrea; Ciuculescu, Felicia; Kollar, Katarina; Seifried, Erhard; Forssmann, Ulf; Verzijl, Dennis; Smit, Martine J; Blanchet, Xavier; von Hundelshausen, Philipp; Weber, Christian; Forssmann, Wolf-Georg; Henschler, Reinhard

    2014-08-15

    Mobilization of hematopoietic stem and progenitor cells (HPCs) is induced by treatment with granulocyte-colony stimulating factor, chemotherapy, or irradiation. We observed that these treatments are accompanied by a release of chemotactic activity into the blood. This plasma activity is derived from the bone marrow, liver, and spleen and acts on HPCs via the chemokine receptor CXCR4. A human blood peptide library was used to characterize CXCR4-activating compounds. We identified CXCL12[22-88] and N-terminally truncated variants CXCL12[24-88], CXCL12[25-88], CXCL12[27-88], and CXCL12[29-88]. Only CXCL12[22-88] could effectively bind to CXCR4 and induce intracellular calcium flux and chemotactic migration of HPCs. CXCL12[25-88] and CXCL12[27-88] revealed neither agonistic nor antagonistic activities in vitro, whereas CXCL12[29-88] inhibited CXCL12[22-88]-induced chemotactic migration. Since binding to glycosaminoglycans (GAG) modulates the function of CXCL12, binding to heparin was analyzed. Surface plasmon resonance kinetic analysis showed that N-terminal truncation of Arg22-Pro23 increased the dissociation constant KD by one log10 stage ([22-88]: KD: 5.4 ± 2.6 μM; [24-88]: KD: 54 ± 22.4 μM). Further truncation of the N-terminus decreased the KD ([25-88] KD: 30 ± 4.8 μM; [27-88] KD: 23 ± 1.6 μM; [29-88] KD: 19 ± 5.4 μM), indicating increasing competition for heparin binding. Systemic in vivo application of CXCL12[22-88] as well as CXCL12[27-88] or CXCL12[29-88] induced a significant mobilization of HPCs in mice. Our findings indicate that plasma-derived CXCL12 variants may contribute to the regulation of HPC mobilization by modulating the binding of CXCL12[22-88] to GAGs rather than blocking the CXCR4 receptor and, therefore, may have a contributing role in HPC mobilization.

  7. Identification and Characterization of Circulating Variants of CXCL12 from Human Plasma: Effects on Chemotaxis and Mobilization of Hematopoietic Stem and Progenitor Cells

    PubMed Central

    Jochheim-Richter, Andrea; Ciuculescu, Felicia; Kollar, Katarina; Seifried, Erhard; Forssmann, Ulf; Verzijl, Dennis; Smit, Martine J.; Blanchet, Xavier; von Hundelshausen, Philipp; Weber, Christian; Forssmann, Wolf-Georg; Henschler, Reinhard

    2014-01-01

    Mobilization of hematopoietic stem and progenitor cells (HPCs) is induced by treatment with granulocyte-colony stimulating factor, chemotherapy, or irradiation. We observed that these treatments are accompanied by a release of chemotactic activity into the blood. This plasma activity is derived from the bone marrow, liver, and spleen and acts on HPCs via the chemokine receptor CXCR4. A human blood peptide library was used to characterize CXCR4-activating compounds. We identified CXCL12[22–88] and N-terminally truncated variants CXCL12[24–88], CXCL12[25–88], CXCL12[27–88], and CXCL12[29–88]. Only CXCL12[22–88] could effectively bind to CXCR4 and induce intracellular calcium flux and chemotactic migration of HPCs. CXCL12[25–88] and CXCL12[27–88] revealed neither agonistic nor antagonistic activities in vitro, whereas CXCL12[29–88] inhibited CXCL12[22–88]-induced chemotactic migration. Since binding to glycosaminoglycans (GAG) modulates the function of CXCL12, binding to heparin was analyzed. Surface plasmon resonance kinetic analysis showed that N-terminal truncation of Arg22-Pro23 increased the dissociation constant KD by one log10 stage ([22–88]: KD: 5.4±2.6 μM; [24–88]: KD: 54±22.4 μM). Further truncation of the N-terminus decreased the KD ([25–88] KD: 30±4.8 μM; [27–88] KD: 23±1.6 μM; [29–88] KD: 19±5.4 μM), indicating increasing competition for heparin binding. Systemic in vivo application of CXCL12[22–88] as well as CXCL12[27–88] or CXCL12[29–88] induced a significant mobilization of HPCs in mice. Our findings indicate that plasma-derived CXCL12 variants may contribute to the regulation of HPC mobilization by modulating the binding of CXCL12[22–88] to GAGs rather than blocking the CXCR4 receptor and, therefore, may have a contributing role in HPC mobilization. PMID:24708339

  8. Identification of a splicing variant that regulates type 2 diabetes risk factor CDKAL1 level by a coding-independent mechanism in human.

    PubMed

    Zhou, Bo; Wei, Fan-Yan; Kanai, Narumi; Fujimura, Atsushi; Kaitsuka, Taku; Tomizawa, Kazuhito

    2014-09-01

    Single-nucleotide polymorphisms (SNPs) in CDKAL1 have been associated with the development of type 2 diabetes (T2D). CDKAL1 catalyzes 2-methylthio modification of adenosine at position 37 of tRNA(Lys)(UUU). A deficit of this modification causes aberrant protein synthesis, and is associated with impairment of insulin secretion in both mouse model and human. However, it is unknown whether the T2D-associated SNPs in CDKAL1 are associated with downregulation of CDKAL1 by regulating the gene expression. Here, we report a specific splicing variant of CDKAL1 termed CDKAL1-v1 that is markedly lower in individuals carrying risk SNPs of CDKAL1. Interestingly, CDKAL1-v1 is a non-coding transcript, which regulates the CDKAL1 level by competitive binding to a CDKAL1-targeting miRNA. By direct editing of the genome, we further show that the nucleotides around the SNP regions are critical for the alternative splicing of CDKAL1-v1. These findings reveal that the T2D-associated SNPs in CDKAL1 reduce CDKAL1-v1 levels by impairing splicing, which in turn increases miRNA-mediated suppression of CDKAL1. Our results suggest that CDKAL1-v1-mediated suppression of CDKAL1 might underlie the pathogenesis of T2D in individuals carrying the risk SNPs. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  9. Identification of a Novel Rat NR2B Subunit Gene Promoter Region Variant and Its Association with Microwave-Induced Neuron Impairment.

    PubMed

    Wang, Li-Feng; Tian, Da-Wei; Li, Hai-Juan; Gao, Ya-Bing; Wang, Chang-Zhen; Zhao, Li; Zuo, Hong-Yan; Dong, Ji; Qiao, Si-Mo; Zou, Yong; Xiong, Lu; Zhou, Hong-Mei; Yang, Yue-Feng; Peng, Rui-Yun; Hu, Xiang-Jun

    2016-05-01

    Microwave radiation has been implicated in cognitive dysfunction and neuronal injury in animal models and in human investigations; however, the mechanism of these effects is unclear. In this study, single nucleotide polymorphism (SNP) sites in the rat GRIN2B promoter region were screened. The associations of these SNPs with microwave-induced rat brain dysfunction and with rat pheochromocytoma-12 (PC12) cell function were investigated. Wistar rats (n = 160) were exposed to microwave radiation (30 mW/cm(2) for 5 min/day, 5 days/week, over a period of 2 months). Screening of the GRIN2B promoter region revealed a stable C-to-T variant at nucleotide position -217 that was not induced by microwave exposure. The learning and memory ability, amino acid contents in the hippocampus and cerebrospinal fluid, and NR2B expression were then investigated in the different genotypes. Following microwave exposure, NR2B protein expression decreased, while the Glu contents in the hippocampus and CSF increased, and memory impairment was observed in the TT genotype but not the CC and CT genotypes. In PC12 cells, the effects of the T allele were more pronounced than those of the C allele on transcription factor binding ability, transcriptional activity, NR2B mRNA, and protein expression. These effects may be related to the detrimental role of the T allele and the protective role of the C allele in rat brain function and PC12 cells exposed to microwave radiation.

  10. Identification of proliferation-induced genes in Arabidopsis thaliana. Characterization of a new member of the highly evolutionarily conserved histone H2A.F/Z variant subfamily.

    PubMed Central

    Callard, D; Mazzolini, L

    1997-01-01

    The changes in gene expression associated with the reinitiation of cell division and subsequent progression through the cell cycle in Arabidopsis thaliana cell-suspension cultures were investigated. Partial synchronization of cells was achieved by a technique combining phosphate starvation and a transient treatment with the DNA replication inhibitor aphidicolin. Six cDNAs corresponding to genes highly induced in proliferating cells and showing cell-cycle-regulated expression were obtained by the mRNA differential display technique. Full-length cDNA clones (cH2BAt and cH2AvAt) corresponding to two of the display products were subsequently isolated. The cH2BAt clone codes for a novel histone H2B protein, whereas the cH2AvAt cDNA corresponds to a gene encoding a new member of the highly conserved histone H2A.F/Z subfamily of chromosomal proteins. Further studies indicated that H2AvAt mRNA expression is tightly correlated with cell proliferation in cell-suspension cultures, and that closely related analogs of the encoded protein exist in Arabidopsis. The implications of the conservation of histone H2A.F/Z variants in plants are discussed. PMID:9414552

  11. Insights Into the Pathogenicity of Rare Missense GCK Variants From the Identification and Functional Characterization of Compound Heterozygous and Double Mutations Inherited in Cis

    PubMed Central

    Beer, Nicola L.; Osbak, Kara K.; van de Bunt, Martijn; Tribble, Nicholas D.; Steele, Anna M.; Wensley, Kirsty J.; Edghill, Emma L.; Colcough, Kevin; Barrett, Amy; Valentínová, Lucia; Rundle, Jana K.; Raimondo, Anne; Grimsby, Joseph; Ellard, Sian; Gloyn, Anna L.

    2012-01-01

    OBJECTIVE To demonstrate the importance of using a combined genetic and functional approach to correctly interpret a genetic test for monogenic diabetes. RESEARCH DESIGN AND METHODS We identified three probands with a phenotype consistent with maturity-onset diabetes of the young (MODY) subtype GCK-MODY, in whom two potential pathogenic mutations were identified: [R43H/G68D], [E248 K/I225M], or [G261R/D217N]. Allele-specific PCR and cosegregation were used to determine phase. Single and double mutations were kinetically characterized. RESULTS The mutations occurred in cis (double mutants) in two probands and in trans in one proband. Functional studies of all double mutants revealed inactivating kinetics. The previously reported GCK-MODY mutations R43H and G68D were inherited from an affected father and unaffected mother, respectively. Both our functional and genetic studies support R43H as the cause of GCK-MODY and G68D as a neutral rare variant. CONCLUSIONS These data highlight the need for family/functional studies, even for previously reported pathogenic mutations. PMID:22611063

  12. Identification of a Nuclear Respiratory Factor 1 Recognition Motif in the Apolipoprotein E Variant APOE4 linked to Alzheimer’s Disease

    PubMed Central

    Urfer-Buchwalder, Anne; Urfer, Roman

    2017-01-01

    Alzheimer’s disease affects tens of millions of people worldwide and its prevalence continues to rise. It is caused by a combination of a subject’s heredity, environment, lifestyle, and medical condition. The most significant genetic risk factor for late onset Alzheimer’s disease is a variant of the apolipoprotein E gene, APOE4. Here we show that the single nucleotide polymorphism rs429358 that defines APOE4 is located in a short sequence motif repeated several times within exon 4 of apolipoprotein E, reminiscent of the structure of transcriptional enhancers. A JASPAR database search predicts that the T to C transition in rs429358 generates a binding motif for nuclear respiratory factor NRF1. This site appears to be part of a binding site cluster for this transcription factor on exon 4 of APOE. This de novo NRF1 binding site has therefore the potential to affect the expression of multiple genes in its genomic vicinity. Our in silico analysis, suggesting a novel function for APOE4 at the DNA level, offers a potential mechanism for the observed tissue specific neurodegeneration and the role of environmental factors in Alzheimer’s disease etiology. PMID:28094792

  13. Cost-Effective and Rapid Presumptive Identification of Gram-Negative Bacilli in Routine Urine, Pus, and Stool Cultures: Evaluation of the Use of CHROMagar Orientation Medium in Conjunction with Simple Biochemical Tests

    PubMed Central

    Ohkusu, Kiyofumi

    2000-01-01

    The algorithm for a new identification system was designed on the basis of colony color and morphology on CHROMagar Orientation medium in conjunction with simple biochemical tests such as indole (IND), lysine decarboxylase (LDC), and ornithine decarboxylase (ODC) utilization tests with gram-negative bacilli isolated from urine samples as well as pus, stool, and other clinical specimens by the following colony characteristics, biochemical reactions, and serological results: pinkish to red, IND positive (IND+), Escherichia coli; metallic blue, IND+, LDC+, and ODC negative (ODC−), Klebsiella oxytoca; IND+, LDC−, and ODC+, Citrobacter diversus; IND+ or IND−, LDC−, and ODC−, Citrobacter freundii; IND−, LDC+, and ODC+, Enterobacter aerogenes; IND−, LDC−, and ODC+, Enterobacter cloacae; IND−, LDC+, and ODC−, Klebsiella pneumoniae; diffuse brown and IND+, Morganella morganii; IND−, Proteus mirabilis; aqua blue, Serratia marcescens; bluish green and IND+, Proteus vulgaris; transparent yellow-green, serology positive, Pseudomonas aeruginosa; clear and serology positive, Salmonella sp.; other colors and reactions, the organism was identified by the full identification methods. The accuracy and cost-effectiveness of this new system were prospectively evaluated. During an 8-month period, a total of 345 specimens yielded one or more gram-negative bacilli. A total of 472 gram-negative bacillus isolates were detected on CHROMagar Orientation medium. For 466 of the isolates (98.7%), no discrepancies in the results were obtained on the basis of the identification algorithm. The cost of identification of gram-negative bacilli during this period was reduced by about 70%. The results of this trial for the differentiation of the most commonly encountered gram-negative pathogens in clinical specimens with the new algorithm were favourable in that it permitted reliable detection and presumptive identification. In addition, this rapid identification system not only

  14. Cost-effective and rapid presumptive identification of gram-negative bacilli in routine urine, pus, and stool cultures: evaluation of the use of CHROMagar orientation medium in conjunction with simple biochemical tests.

    PubMed

    Ohkusu, K

    2000-12-01

    The algorithm for a new identification system was designed on the basis of colony color and morphology on CHROMagar Orientation medium in conjunction with simple biochemical tests such as indole (IND), lysine decarboxylase (LDC), and ornithine decarboxylase (ODC) utilization tests with gram-negative bacilli isolated from urine samples as well as pus, stool, and other clinical specimens by the following colony characteristics, biochemical reactions, and serological results: pinkish to red, IND positive (IND(+)), Escherichia coli; metallic blue, IND(+), LDC(+), and ODC negative (ODC(-)), Klebsiella oxytoca; IND(+), LDC(-), and ODC(+), Citrobacter diversus; IND(+) or IND(-), LDC(-), and ODC(-), Citrobacter freundii; IND(-), LDC(+), and ODC(+), Enterobacter aerogenes; IND(-), LDC(-), and ODC(+), Enterobacter cloacae; IND(-), LDC(+), and ODC(-), Klebsiella pneumoniae; diffuse brown and IND(+), Morganella morganii; IND(-), Proteus mirabilis; aqua blue, Serratia marcescens; bluish green and IND(+), Proteus vulgaris; transparent yellow-green, serology positive, Pseudomonas aeruginosa; clear and serology positive, Salmonella sp.; other colors and reactions, the organism was identified by the full identification methods. The accuracy and cost-effectiveness of this new system were prospectively evaluated. During an 8-month period, a total of 345 specimens yielded one or more gram-negative bacilli. A total of 472 gram-negative bacillus isolates were detected on CHROMagar Orientation medium. For 466 of the isolates (98.7%), no discrepancies in the results were obtained on the basis of the identification algorithm. The cost of identification of gram-negative bacilli during this period was reduced by about 70%. The results of this trial for the differentiation of the most commonly encountered gram-negative pathogens in clinical specimens with the new algorithm were favourable in that it permitted reliable detection and presumptive identification. In addition, this rapid

  15. Identification of a genetic variant at 2q12.1 associated with blood pressure in East Asians by genome-wide scan including gene-environment interactions.

    PubMed

    Kim, Yun Kyoung; Kim, Youngdoe; Hwang, Mi Yeong; Shimokawa, Kazuro; Won, Sungho; Kato, Norihiro; Tabara, Yasuharu; Yokota, Mitsuhiro; Han, Bok-Ghee; Lee, Jong Ho; Kim, Bong-Jo

    2014-06-05

    Genome-wide association studies have identified many genetic loci associated with blood pressure (BP). Genetic effects on BP can be altered by environmental exposures via multiple biological pathways. Especially, obesity is one of important environmental risk factors that can have considerable effect on BP and it may interact with genetic factors. Given that, we aimed to test whether genetic factors and obesity may jointly influence BP. We performed meta-analyses of genome-wide association data for systolic blood pressure (SBP) and diastolic blood pressure (DBP) that included analyses of interaction between single nucleotide polymorphisms (SNPs) and the obesity-related anthropometric measures, body mass index (BMI), height, weight, and waist/hip ratio (WHR) in East-Asians (n = 12,030). We identified that rs13390641 on 2q12.1 demonstrated significant association with SBP when the interaction between SNPs and BMI was considered (P < 5 × 10 -8). The gene located nearest to rs13390641, TMEM182, encodes transmembrane protein 182. In stratified analyses, the effect of rs13390641 on BP was much stronger in obese individuals (BMI ≥ 30) than non-obese individuals and the effect of BMI on BP was strongest in individuals with the homozygous A allele of rs13390641. Our analyses that included interactions between SNPs and environmental factors identified a genetic variant associated with BP that was overlooked in standard analyses in which only genetic factors were included. This result also revealed a potential mechanism that integrates genetic factors and obesity related traits in the development of high BP.

  16. Variant CJD

    PubMed Central

    Diack, Abigail B; Head, Mark W; McCutcheon, Sandra; Boyle, Aileen; Knight, Richard; Ironside, James W; Manson, Jean C; Will, Robert G

    2014-01-01

    It is now 18 years since the first identification of a case of vCJD in the UK. Since that time, there has been much speculation over how vCJD might impact human health. To date there have been 177 case reports in the UK and a further 51 cases worldwide in 11 different countries. Since establishing that BSE and vCJD are of the same strain of agent, we have also shown that there is broad similarity between UK and non-UK vCJD cases on first passage to mice. Transgenic mouse studies have indicated that all codon 129 genotypes are susceptible to vCJD and that genotype may influence whether disease appears in a clinical or asymptomatic form, supported by the appearance of the first case of potential asymptomatic vCJD infection in a PRNP 129MV patient. Following evidence of blood transfusion as a route of transmission, we have ascertained that all blood components and leucoreduced blood in a sheep model of vCJD have the ability to transmit disease. Importantly, we recently established that a PRNP 129MV patient blood recipient with an asymptomatic infection and limited PrPSc deposition in the spleen could readily transmit disease into mice, demonstrating the potential for peripheral infection in the absence of clinical disease. This, along with the recent appendix survey which identified 16 positive appendices in a study of 32 441 cases, underlines the importance of continued CJD surveillance and maintaining control measures already in place to protect human health. PMID:25495404

  17. Identification of splice variants, targeted microRNAs and functional single nucleotide polymorphisms of the BOLA-DQA2 gene in dairy cattle.

    PubMed

    Hou, Qinlei; Huang, Jinming; Ju, Zhihua; Li, Qiuling; Li, Liming; Wang, Changfa; Sun, Tao; Wang, Lingling; Hou, Minghai; Hang, Suqin; Zhong, Jifeng

    2012-05-01

    Major histocompatibility complex, class II, DQ alpha 2, also named BOLA-DQA2, belongs to the Bovine Leukocyte Antigen (BOLA) class II genes which are involved in the immune response. To explore the variability of the BOLA-DQA2 gene and resistance to mastitis in cows, the splice variants (SV), targeted microRNAs (miRNAs), and single nucleotide polymorphisms (SNPs) were identified in this study. A new SV (BOLA-DQA2-SV1) lacking part of exon 3 (195 bp) and two 3'-untranslated regions (UTR) (52 bp+167 bp) of the BOLA-DQA2 gene was found in the healthy and mastitis-infected mammary gland tissues. Four of 13 new SNPs and multiple nucleotide polymorphisms resulted in amino acid changes in the protein and SNP (c. +1283 C>T) may affect the binding to the seed sequence of bta-miR-2318. Further, we detected the relative expressions of two BOLA-DQA2 transcripts and five candidated microRNAs binding to the 3'-UTR of two transcripts in the mammary gland tissues in dairy cattle by using the quantitative real-time polymerase chain reaction. The result showed that expression of the BOLA-DQA2-SV1 mRNA was significantly upregulated 2.67-fold (p<0.05) in mastitis-infected mammary tissues (n = 5) compared with the healthy mammary gland mammary tissues (n = 5). Except for bta-miR-1777a, miRNA expression (bta-miR-296, miR-2430, and miR-671) was upregulated 1.75 to 2.59-fold (p<0.05), whereas miR-2318 was downregulated in the mastitis cows. Our findings reveal that BOLA-DQA2-SV1 may play an important role in the mastitis resistance in dairy cattle. Whether the SNPs affect the structure of the BOLA-DQA2 gene or association with mastitis resistance is unknown and warrants further investigation.

  18. Identification of Splice Variants, Targeted MicroRNAs and Functional Single Nucleotide Polymorphisms of the BOLA-DQA2 Gene in Dairy Cattle

    PubMed Central

    Hou, Qinlei; Huang, Jinming; Ju, Zhihua; Li, Qiuling; Li, Liming; Wang, Changfa; Sun, Tao; Wang, Lingling; Hou, Minghai

    2012-01-01

    Major histocompatibility complex, class II, DQ alpha 2, also named BOLA-DQA2, belongs to the Bovine Leukocyte Antigen (BOLA) class II genes which are involved in the immune response. To explore the variability of the BOLA-DQA2 gene and resistance to mastitis in cows, the splice variants (SV), targeted microRNAs (miRNAs), and single nucleotide polymorphisms (SNPs) were identified in this study. A new SV (BOLA-DQA2-SV1) lacking part of exon 3 (195 bp) and two 3′-untranslated regions (UTR) (52 bp+167 bp) of the BOLA-DQA2 gene was found in the healthy and mastitis-infected mammary gland tissues. Four of 13 new SNPs and multiple nucleotide polymorphisms resulted in amino acid changes in the protein and SNP (c. +1283 C>T) may affect the binding to the seed sequence of bta-miR-2318. Further, we detected the relative expressions of two BOLA-DQA2 transcripts and five candidated microRNAs binding to the 3′-UTR of two transcripts in the mammary gland tissues in dairy cattle by using the quantitative real-time polymerase chain reaction. The result showed that expression of the BOLA-DQA2-SV1 mRNA was significantly upregulated 2.67-fold (p<0.05) in mastitis-infected mammary tissues (n=5) compared with the healthy mammary gland mammary tissues (n=5). Except for bta-miR-1777a, miRNA expression (bta-miR-296, miR-2430, and miR-671) was upregulated 1.75 to 2.59-fold (p<0.05), whereas miR-2318 was downregulated in the mastitis cows. Our findings reveal that BOLA-DQA2-SV1 may play an important role in the mastitis resistance in dairy cattle. Whether the SNPs affect the structure of the BOLA-DQA2 gene or association with mastitis resistance is unknown and warrants further investigation. PMID:22084936

  19. Identification of Alternative Variants and Insertion of the Novel Polymorphic AluYl17 in TSEN54 Gene during Primate Evolution

    PubMed Central

    Kim, Young-Hyun; Choe, Se-Hee; Cho, Hyeon-Mu; Lee, Sang-Rae; Kim, Ji-Su; Song, Bong-Seok; Jeong, Kang-Jin; Jin, Yeung Bae; Kang, Philyong

    2016-01-01

    TSEN54 encodes a subunit of the tRNA-splicing endonuclease complex, which catalyzes the identification and cleavage of introns from precursor tRNAs. Previously, we identified an AluSx-derived alternative transcript in TSEN54 of cynomolgus monkey. Reverse transcription-polymerase chain reaction (RT-PCR) amplification and TSEN54 sequence analysis of primate and human samples identified five novel alternative transcripts, including the AluSx exonized transcript. Additionally, we performed comparative expression analysis via RT-qPCR in various cynomolgus, rhesus monkey, and human tissues. RT-qPCR amplification revealed differential expression patterns. Furthermore, genomic PCR amplification and sequencing of primate and human DNA samples revealed that AluSx elements were integrated in human and all of the primate samples tested. Intriguingly, in langur genomic DNA, an additional AluY element was inserted into AluSx of intron eight of TSEN54. The new AluY element showed polymorphic insertion. Using standardized nomenclature for Alu repeats, the polymorphic AluY of the langur TSEN54 was designated as being of the AluYl17 subfamily. Our results suggest that integration of the AluSx element in TSEN54 contributed to diversity in transcripts and induced lineage- or species-specific evolutionary events such as alternative splicing and polymorphic insertion during primate evolution. PMID:28083540

  20. Identification and functional characterization of NifA variants that are independent of GlnB activation in the photosynthetic bacterium Rhodospirillum rubrum

    PubMed Central

    Zou, Xiaoxiao; Zhu, Yu; Pohlmann, Edward L.; Li, Jilun; Zhang, Yaoping; Roberts, Gary P.

    2012-01-01

    The activity of NifA, the transcriptional activator of the nitrogen fixation (nif) gene, is tightly regulated in response to ammonium and oxygen. However, the mechanisms for the regulation of NifA activity are quite different among various nitrogen-fixing bacteria. Unlike the well-studied NifL–NifA regulatory systems in Klebsiella pneumoniae and Azotobacter vinelandii, in Rhodospirillum rubrum NifA is activated by a direct protein–protein interaction with the uridylylated form of GlnB, which in turn causes a conformational change in NifA. We report the identification of several substitutions in the N-terminal GAF domain of R. rubrum NifA that allow NifA to be activated in the absence of GlnB. Presumably these substitutions cause conformational changes in NifA necessary for activation, without interaction with GlnB. We also found that wild-type NifA can be activated in a GlnB-independent manner under certain growth conditions, suggesting that some other effector(s) can also activate NifA. An attempt to use Tn5 mutagenesis to obtain mutants that altered the pool of these presumptive effector(s) failed, though much rarer spontaneous mutations in nifA were detected. This suggests that the necessary alteration of the pool of effector(s) for NifA activation cannot be obtained by knockout mutations. PMID:18757802

  1. Genetic variants of human parvovirus B19 in South Africa: cocirculation of three genotypes and identification of a novel subtype of genotype 1.

    PubMed

    Corcoran, Craig; Hardie, Diana; Yeats, Jane; Smuts, Heidi

    2010-01-01

    Parvovirus B19 comprises three distinct genotypes (1, 2, and 3). The distribution of B19 genotypes has not before been examined in South Africa. Two hundred thirty-nine laboratory samples submitted to a diagnostic virology laboratory for parvovirus DNA detection were analyzed retrospectively. Of the 53 PCR-positive samples investigated, 40 (75.4%) were identified as genotype 1 by genotype-specific PCR or consensus NS1 PCR and sequencing and 3 (5.7%) as genotype 2 and 10 (18.9%) as genotype 3 by analysis of NS1 sequences. Furthermore, phylogenetic analysis identified two genotype 1 sequences which were distinct from the previously described genotypes 1A and 1B. Interestingly, a genotype 2 virus was detected in the serum of an 11-year-old child, providing evidence for its recent circulation. This is the first study to demonstrate the concurrent circulation of all three genotypes of B19 in South Africa and the provisional identification of a novel subtype of genotype 1. The implications of parvovirus B19 variation are discussed.

  2. Detection of 5α-androst-2-en-17-one and variants: Identification of main urinary metabolites in human urine samples by GC-MS and NMR.

    PubMed

    Ayotte, Christiane; Sylvestre, Alexandre; Charlebois, Alain; Poirier, Donald

    2016-11-01

    Two steroids were identified in a supplement named D-2 following the detection of unknown compounds during the routine testing of an athlete's sample. The main glucuroconjugated metabolites were isolated from this urine by high performance liquid chromatography (HPLC) following enzymatic hydrolysis and identified by gas chromatography-mass spectrometry (GC-MS) and nuclear magnetic resonance (NMR) analyses as being 2α-hydroxy-5α-androst-3-en-17-one (M1) and 2β,3α-dihydroxy-5α-androstan-17-one (M2). A third metabolite, 3α,4β-dihydroxy-5α-androstan-17-one (M3) was also detected, however in lower amounts. The precursor steroids, 5α-androst-2-en-17-one (1) and 5α-androst-3-en-17-one (2) were present in the first D-2 products offered on the Internet. Later, the corresponding 17-hydroxyl compounds were offered as such or as esters (acetate, cypionate) in different relative ratios. Both M2 and M3 were synthesized from the trans-diaxial hydrolysis of the corresponding 2α,3α- and 3α,4α-epoxides (3). These were excreted in the hours following the controlled administration of the commercial product called D-2 R to a male volunteer and were also produced from the incubation of 1 and 2 with S9 liver fractions. Some preparations contain predominantly the alkene in C-2 and, therefore, an efficient detection method must include both primary metabolites M1 and M2. The latter was found equally in the fractions extracted following the enzymatic hydrolysis with β-glucuronidase and the chemical solvolysis, which may ease its identification. Copyright © 2016 John Wiley & Sons, Ltd.

  3. Database for Parkinson Disease Mutations and Rare Variants

    DTIC Science & Technology

    2015-07-01

    due to time or knowledge restrained to sift through scattered information. Current databases do not address actual contribution of variant to PD...variant discovery , the Parkinson Disease project focuses on identification of rare variants and their differential accumulation within a gene region...During her three year fellowship in Miami, she has extended her knowledge and acquired vast experience working with next generation sequencing

  4. RHD variants in Flanders, Belgium.

    PubMed

    Van Sandt, Vicky S T; Gassner, Christoph; Emonds, Marie-Paule; Legler, Tobias J; Mahieu, Sarah; Körmöczi, Günther F

    2015-06-01

    D antigen variants may be grouped into partial D, weak D, and DEL types. Cumulative phenotype frequencies of these D variants may approach 1% in certain European regions. Unambiguous and quick identification of D variants is of immediate clinical relevance, with implications for transfusion strategy. A total of 628 samples with ambiguous serologic results from different immunohematology laboratories throughout the Flanders region, Belgium, were genotyped using a commercially available weak D typing approach. After exclusion of detectable weak D types, molecular RHD exon scanning was performed for the remaining samples, and RHD sequencing was performed in two particular cases. Of all samples investigated, 424 (67.5%) were positive for weak D Type 1, 2, or 3, and 22 cases (3.5%) typed weak D Type 4.0/4.1/4.3, 4.2, 5, 11, 15, or 17. Another 49 (7.8%) samples were partial D variants, with a major proportion being category DVI types (n = 27). One RHD(S103P) sample was identified as high-grade partial D, with DIII-like phenotype and anti-D and anti-C immunization. Additionally, a novel DVI Type 3 (A399T) variant was found. Of the remaining 133 samples mainly tested because of ambiguous serologic D typing results due to recent transfusion, 32 (5.1%) were negative for RHD, and 101 (16.1%) were indistinguishable from wild-type RHD and not investigated further. Despite the enormous diversity of RHD alleles, first-line weak D genotyping was remarkably informative, allowing for rapid classification of most samples with conspicuous RhD phenotype in Flanders. The clinical implications are discussed. © 2014 AABB.

  5. A rare hemoglobin variant, Hb Belliard

    PubMed Central

    Benavides, Raul

    2017-01-01

    There are many documented variants of hemoglobin; however, other than a limited number (such as sickle cell disease), very few are known to have any clinical significance. As advances in detection and identification continue through gel electrophoresis, capillary electrophoresis, and DNA sequencing, more rare variants are identified. Without case reporting, the significance of these variants will remain unknown or continue to be thought of as insignificant. Here we report a rare hemoglobin variant, Hb Belliard, which was detected in a 68-year-old Indian immigrant to the United States. He presented with elevated hemoglobin and was found to have a unique peak on capillary electrophoresis. The specimen was sent for sequencing and was subsequently found to have Hb Belliard. Currently, Hb Belliard is thought to be insignificant.

  6. Search-based model identification of smart-structure damage

    NASA Technical Reports Server (NTRS)

    Glass, B. J.; Macalou, A.

    1991-01-01

    This paper describes the use of a combined model and parameter identification approach, based on modal analysis and artificial intelligence (AI) techniques, for identifying damage or flaws in a rotating truss structure incorporating embedded piezoceramic sensors. This smart structure example is representative of a class of structures commonly found in aerospace systems and next generation space structures. Artificial intelligence techniques of classification, heuristic search, and an object-oriented knowledge base are used in an AI-based model identification approach. A finite model space is classified into a search tree, over which a variant of best-first search is used to identify the model whose stored response most closely matches that of the input. Newly-encountered models can be incorporated into the model space. This adaptativeness demonstrates the potential for learning control. Following this output-error model identification, numerical parameter identification is used to further refine the identified model. Given the rotating truss example in this paper, noisy data corresponding to various damage configurations are input to both this approach and a conventional parameter identification method. The combination of the AI-based model identification with parameter identification is shown to lead to smaller parameter corrections than required by the use of parameter identification alone.

  7. Identification of membrane-bound variant of metalloendopeptidase neurolysin (EC 3.4.24.16) as the non-angiotensin type 1 (non-AT1), non-AT2 angiotensin binding site.

    PubMed

    Wangler, Naomi J; Santos, Kira L; Schadock, Ines; Hagen, Fred K; Escher, Emanuel; Bader, Michael; Speth, Robert C; Karamyan, Vardan T

    2012-01-02

    Recently, we discovered a novel non-angiotensin type 1 (non-AT1), non-AT2 angiotensin binding site in rodent and human brain membranes, which is distinctly different from angiotensin receptors and key proteases processing angiotensins. It is hypothesized to be a new member of the renin-angiotensin system. This study was designed to isolate and identify this novel angiotensin binding site. An angiotensin analog, photoaffinity probe 125I-SBpa-Ang II, was used to specifically label the non-AT1, non-AT2 angiotensin binding site in mouse forebrain membranes, followed by a two-step purification procedure based on the molecular size and isoelectric point of the photoradiolabeled binding protein. Purified samples were subjected to two-dimensional gel electrophoresis followed by mass spectrometry identification of proteins in the two-dimensional gel sections containing radioactivity. LC-MS/MS analysis revealed eight protein candidates, of which the four most abundant were immunoprecipitated after photoradiolabeling. Immunoprecipitation studies indicated that the angiotensin binding site might be the membrane-bound variant of metalloendopeptidase neurolysin (EC 3.4.24.16). To verify these observations, radioligand binding and photoradiolabeling experiments were conducted in membrane preparations of HEK293 cells overexpressing mouse neurolysin or thimet oligopeptidase (EC 3.4.24.15), a closely related metalloendopeptidase of the same family. These experiments also identified neurolysin as the non-AT1, non-AT2 angiotensin binding site. Finally, brain membranes of mice lacking neurolysin were nearly devoid of the non-AT1, non-AT2 angiotensin binding site, further establishing membrane-bound neurolysin as the binding site. Future studies will focus on the functional significance of this highly specific, high affinity interaction between neurolysin and angiotensins.

  8. Identification of Membrane-bound Variant of Metalloendopeptidase Neurolysin (EC 3.4.24.16) as the Non-angiotensin Type 1 (Non-AT1), Non-AT2 Angiotensin Binding Site*

    PubMed Central

    Wangler, Naomi J.; Santos, Kira L.; Schadock, Ines; Hagen, Fred K.; Escher, Emanuel; Bader, Michael; Speth, Robert C.; Karamyan, Vardan T.

    2012-01-01

    Recently, we discovered a novel non-angiotensin type 1 (non-AT1), non-AT2 angiotensin binding site in rodent and human brain membranes, which is distinctly different from angiotensin receptors and key proteases processing angiotensins. It is hypothesized to be a new member of the renin-angiotensin system. This study was designed to isolate and identify this novel angiotensin binding site. An angiotensin analog, photoaffinity probe 125I-SBpa-Ang II, was used to specifically label the non-AT1, non-AT2 angiotensin binding site in mouse forebrain membranes, followed by a two-step purification procedure based on the molecular size and isoelectric point of the photoradiolabeled binding protein. Purified samples were subjected to two-dimensional gel electrophoresis followed by mass spectrometry identification of proteins in the two-dimensional gel sections containing radioactivity. LC-MS/MS analysis revealed eight protein candidates, of which the four most abundant were immunoprecipitated after photoradiolabeling. Immunoprecipitation studies indicated that the angiotensin binding site might be the membrane-bound variant of metalloendopeptidase neurolysin (EC 3.4.24.16). To verify these observations, radioligand binding and photoradiolabeling experiments were conducted in membrane preparations of HEK293 cells overexpressing mouse neurolysin or thimet oligopeptidase (EC 3.4.24.15), a closely related metalloendopeptidase of the same family. These experiments also identified neurolysin as the non-AT1, non-AT2 angiotensin binding site. Finally, brain membranes of mice lacking neurolysin were nearly devoid of the non-AT1, non-AT2 angiotensin binding site, further establishing membrane-bound neurolysin as the binding site. Future studies will focus on the functional significance of this highly specific, high affinity interaction between neurolysin and angiotensins. PMID:22039052

  9. Identification and Modelling of the In-Plane Reinforcement Orientation Variations in a CFRP Laminate Produced by Manual Lay-Up

    NASA Astrophysics Data System (ADS)

    Davila, Yves; Crouzeix, Laurent; Douchin, Bernard; Collombet, Francis; Grunevald, Yves-Henri

    2017-08-01

    Reinforcement angle orientation has a significant effect on the mechanical properties of composite materials. This work presents a methodology to introduce variable reinforcement angles into finite element (FE) models of composite structures. The study of reinforcement orientation variations uses meta-models to identify and control a continuous variation across the composite ply. First, the reinforcement angle is measured through image analysis techniques of the composite plies during the lay-up phase. Image analysis results show that variations in the mean ply orientations are between -0.5 and 0.5° with standard deviations ranging between 0.34 and 0.41°. An automatic post-treatment of the images determines the global and local angle variations yielding good agreements visually and numerically between the analysed images and the identified parameters. A composite plate analysed at the end of the cooling phase is presented as a case of study. Here, the variation in residual strains induced by the variability in the reinforcement orientation are up to 28% of the strain field of the homogeneous FE model. The proposed methodology has shown its capabilities to introduce material and geometrical variability into FE analysis of layered composite structures.

  10. Evaluation of the Biomic V3 Microbiology System for Identification of Selected Species on BBL CHROMagar Orientation Agar and CHROMagar MRSA Medium ▿

    PubMed Central

    Baron, Ellen Jo; D'Souza, Holly; Qi Wang, Andrew; Gibbs, David L.

    2008-01-01

    The Biomic V3 microbiology system identifies bacteria by reading the color of colonies selected by the user. For CHROMagar orientation, Biomic results agreed with conventional methods for 94% of the strains assayed. For CHROMagar MRSA, Biomic correctly identified 100% of the strains tested and did not misidentify two methicillin-susceptible Staphylococcus aureus strains growing on the plates. PMID:18701661

  11. Orienteering injuries

    PubMed Central

    Folan, Jean M.

    1982-01-01

    At the Irish National Orienteering Championships in 1981 a survey of the injuries occurring over the two days of competition was carried out. Of 285 individual competitors there was a percentage injury rate of 5.26%. The article discusses the injuries and aspects of safety in orienteering. Imagesp236-ap237-ap237-bp238-ap239-ap240-a PMID:7159815

  12. Histone Variants and Epigenetics

    PubMed Central

    Henikoff, Steven; Smith, M. Mitchell

    2015-01-01

    Histones package and compact DNA by assembling into nucleosome core particles. Most histones are synthesized at S phase for rapid deposition behind replication forks. In addition, the replacement of histones deposited during S phase by variants that can be deposited independently of replication provide the most fundamental level of chromatin differentiation. Alternative mechanisms for depositing different variants can potentially establish and maintain epigenetic states. Variants have also evolved crucial roles in chromosome segregation, transcriptional regulation, DNA repair, and other processes. Investigations into the evolution, structure, and metabolism of histone variants provide a foundation for understanding the participation of chromatin in important cellular processes and in epigenetic memory. PMID:25561719

  13. Protein characterization by LC-MS/MS may be required for the DNA identification of a fusion hemoglobin: the example of Hb P-Nilotic.

    PubMed

    Zanella-Cleon, Isabelle; Delolme, Frédéric; Lacan, Philippe; Garcia, Caroline; Vinatier, Isabelle; Francina, Alain; Joly, Philippe

    2012-02-01

    DNA analysis is currently the easiest way to identify a hemoglobin variant in most cases. Nevertheless, in case of complex gene rearrangements, mass spectrometry studies may be required to orientate the DNA diagnosis. The present report shows the use of mass spectrometry techniques prior to DNA analysis for the identification of the rare P-Nilotic fusion hemoglobin. Complete protein analysis is performed by liquid chromatography-tandem mass spectrometry on the abnormal globin chain isolated by reversed-phase liquid chromatography.

  14. Orientation and the Young Orienteer

    NASA Astrophysics Data System (ADS)

    Walsh, S. E.; Martland, J. R.

    Orientation within orienteering is dependent on the use of two basic strategies; that is, either a compass or Magnetic-North-based strategy, which relies on the use of one set of information; or the use of a map and landmark-based strategy which relies on the use of at least two sets of information. Walsh and found that, when given the choice, young children use the compass-based strategy when following complex potentially disorientating routes.The efficacy of these two basic orientation strategies was investigated within three different orienteering environments: (1) a familiar known environment; (2) a familiar unknown environment and (3) an unfamiliar unknown environment.Subjects, age range from 9 to 10think aloud particularly the introduction of basic skills to young performers. They support the argument that is essential to introduce the map and compass simultaneously and that relocation and orientation skills should be coached concurrently.

  15. Twin Research and the Arts: Interconnections / Twin Research: Twin Studies of Sexual Orientation; A Historical Biological Twin Gem; GWAS Approach to Who Has Twins / Newsworthy: Twins on College Campuses; 'Brainprint': Personal Identification by Brain Waves.

    PubMed

    Segal, Nancy L

    2016-08-01

    The interrelatedness between twin research and the arts is explored via a new play about a famous case. In the 1960s, identical twin David Bruce Reimer was accidentally castrated as an infant during circumcision to correct a urinary problem. The decision to raise him as a girl, and the consequences of that decision, are explored in the new theatrical production of Boy. Other examples of the arts mirroring science, and vice versa, are described. Next, brief reviews and summaries of twin research on sexual orientation, 1860s' knowledge of placental arrangements and twinning mechanisms, and genes underlying multiple birth conception and fertility related measures are provided. This article concludes with a look at twins on college campuses and the identification of individuals by their brain waves. A correction and clarification regarding my article on the Brazilian Twin Registry in the last issue of THG (Segal, 2016) is also provided.

  16. Mutation Update: The Spectra of Nebulin Variants and Associated Myopathies

    PubMed Central

    Lehtokari, Vilma-Lotta; Kiiski, Kirsi; Sandaradura, Sarah A.; Laporte, Jocelyn; Repo, Pauliina; Frey, Jennifer A.; Donner, Kati; Marttila, Minttu; Saunders, Carol; Barth, Peter G.; den Dunnen, Johan T.; Beggs, Alan H.; Clarke, Nigel F.; North, Kathryn N.; Laing, Nigel G.; Romero, Norma B.; Winder, Thomas L.; Pelin, Katarina; Wallgren-Pettersson, Carina

    2015-01-01

    A mutation update on the nebulin gene (NEB) is necessary because of recent developments in analysis methodology, the identification of increasing numbers and novel types of variants, and a widening in the spectrum of clinical and histological phenotypes associated with this gigantic, 183 exons containing gene. Recessive pathogenic variants in NEB are the major cause of nemaline myopathy (NM), one of the most common congenital myopathies. Moreover, pathogenic NEB variants have been identified in core-rod myopathy and in distal myopathies. In this update, we present the disease-causing variants in NEB in 159 families, 143 families with NM, and 16 families with NM-related myopathies. Eighty-eight families are presented here for the first time. We summarize 86 previously published and 126 unpublished variants identified in NEB. Furthermore, we have analyzed the NEB variants deposited in the Exome Variant Server (http://evs.gs.washington.edu/EVS/), identifying that pathogenic variants are a minor fraction of all coding variants (~7%). This indicates that nebulin tolerates substantial changes in its amino acid sequence, providing an explanation as to why variants in such a large gene result in relatively rare disorders. Lastly, we discuss the difficulties of drawing reliable genotype–phenotype correlations in NEB-associated disease. PMID:25205138

  17. Analyzing Orientations

    NASA Astrophysics Data System (ADS)

    Ruggles, Clive L. N.

    Archaeoastronomical field survey typically involves the measurement of structural orientations (i.e., orientations along and between built structures) in relation to the visible landscape and particularly the surrounding horizon. This chapter focuses on the process of analyzing the astronomical potential of oriented structures, whether in the field or as a desktop appraisal, with the aim of establishing the archaeoastronomical "facts". It does not address questions of data selection (see instead Chap. 25, "Best Practice for Evaluating the Astronomical Significance of Archaeological Sites", 10.1007/978-1-4614-6141-8_25) or interpretation (see Chap. 24, "Nature and Analysis of Material Evidence Relevant to Archaeoastronomy", 10.1007/978-1-4614-6141-8_22). The main necessity is to determine the azimuth, horizon altitude, and declination in the direction "indicated" by any structural orientation. Normally, there are a range of possibilities, reflecting the various errors and uncertainties in estimating the intended (or, at least, the constructed) orientation, and in more formal approaches an attempt is made to assign a probability distribution extending over a spread of declinations. These probability distributions can then be cumulated in order to visualize and analyze the combined data from several orientations, so as to identify any consistent astronomical associations that can then be correlated with the declinations of particular astronomical objects or phenomena at any era in the past. The whole process raises various procedural and methodological issues and does not proceed in isolation from the consideration of corroborative data, which is essential in order to develop viable cultural interpretations.

  18. Identification of RNO 43 and B335 as two highly collimated bipolar flows oriented nearly in the plane of the sky

    NASA Technical Reports Server (NTRS)

    Cabrit, Sylvie; Goldsmith, Paul F.; Snell, Ronald L.

    1988-01-01

    An analysis of CO and CS line observations shows that RNO 43 and B335 are single highly collimated bipolar flows oriented nearly in the plane of the sky. The angle between the line of sight and the flow axis is estimated to be 85 deg in RNO 43 and 82 deg in B335. The results indicate that each of these flows must have a significant expansion velocity perpendicular to its axis, and that the flow in B335 must be driven by a mostly nonmolecular wind component.

  19. Wham: Identifying Structural Variants of Biological Consequence

    PubMed Central

    Kronenberg, Zev N.; Osborne, Edward J.; Cone, Kelsey R.; Kennedy, Brett J.; Domyan, Eric T.; Shapiro, Michael D.; Elde, Nels C.; Yandell, Mark

    2015-01-01

    Existing methods for identifying structural variants (SVs) from short read datasets are inaccurate. This complicates disease-gene identification and efforts to understand the consequences of genetic variation. In response, we have created Wham (Whole-genome Alignment Metrics) to provide a single, integrated framework for both structural variant calling and association testing, thereby bypassing many of the difficulties that currently frustrate attempts to employ SVs in association testing. Here we describe Wham, benchmark it against three other widely used SV identification tools–Lumpy, Delly and SoftSearch–and demonstrate Wham’s ability to identify and associate SVs with phenotypes using data from humans, domestic pigeons, and vaccinia virus. Wham and all associated software are covered under the MIT License and can be freely downloaded from github (https://github.com/zeeev/wham), with documentation on a wiki (http://zeeev.github.io/wham/). For community support please post questions to https://www.biostars.org/. PMID:26625158

  20. Wham: Identifying Structural Variants of Biological Consequence.

    PubMed

    Kronenberg, Zev N; Osborne, Edward J; Cone, Kelsey R; Kennedy, Brett J; Domyan, Eric T; Shapiro, Michael D; Elde, Nels C; Yandell, Mark

    2015-12-01

    Existing methods for identifying structural variants (SVs) from short read datasets are inaccurate. This complicates disease-gene identification and efforts to understand the consequences of genetic variation. In response, we have created Wham (Whole-genome Alignment Metrics) to provide a single, integrated framework for both structural variant calling and association testing, thereby bypassing many of the difficulties that currently frustrate attempts to employ SVs in association testing. Here we describe Wham, benchmark it against three other widely used SV identification tools-Lumpy, Delly and SoftSearch-and demonstrate Wham's ability to identify and associate SVs with phenotypes using data from humans, domestic pigeons, and vaccinia virus. Wham and all associated software are covered under the MIT License and can be freely downloaded from github (https://github.com/zeeev/wham), with documentation on a wiki (http://zeeev.github.io/wham/). For community support please post questions to https://www.biostars.org/.

  1. Crystallographic and magnetic identification of secondary phase in orientated Bi5Fe0.5Co0.5Ti3O15 ceramics

    NASA Astrophysics Data System (ADS)

    Palizdar, Meghdad; Comyn, Tim P.; Ward, Michael B.; Brown, Andrew P.; Harrington, John P.; Kulkarni, Santosh; Keeney, Lynette; Roy, Saibal; Pemble, Martyn; Whatmore, Roger; Quinn, Christopher; Kilcoyne, Susan H.; Bell, Andrew J.

    2012-10-01

    The fabrication of highly-oriented polycrystalline ceramics of Bi5Fe0.5Co0.5Ti3O15, prepared via molten salt synthesis and uniaxial pressing of high aspect ratio platelets is reported. Electron backscatter images show a secondary phase within the ceramic which is rich in cobalt and iron. The concentration of the secondary phase obtained from scanning electron microscopy is estimated at less than 2% by volume, below the detection limit of x-ray diffraction (XRD). The samples were characterized by x-ray diffraction, polarization-electric field measurements, superconducting quantum interference device as a function of sample orientation and vibrating sample magnetometry as a function of temperature. It is inferred from the data that the observed ferromagnetic response is dominated by the secondary phase. This work highlights the importance of rigorous materials characterisation in the study of multiferroics as small amounts of secondary phase, below the limit of XRD, can lead to false conclusions.

  2. Identification of Key Proteins and Networks Related to Grain Development in Wheat (Triticum aestivum L.) by Comparative Transcription and Proteomic Analysis of Allelic Variants in TaGW2-6A

    PubMed Central

    Du, Dengfeng; Gao, Xin; Geng, Juan; Li, Qingyan; Li, Liqun; Lv, Qian; Li, Xuejun

    2016-01-01

    In wheat, coding region allelic variants of TaGW2-6A are closely associated with grain width and weight, but the genetic mechanisms involved remain unclear. Thus, to obtain insights into the key functions regulated by TaGW2-6A during wheat grain development, we performed transcriptional and proteomic analyses of TaGW2-6A allelic variants. The transcription results showed that the TaGW2-6A allelic variants differed significantly by several orders of magnitude. Each allelic variant of TaGW2-6A reached its first transcription peak at 6 days after anthesis (DAA), but the insertion type TaGW2-6A allelic variant reached its second peak earlier than the normal type, i.e., at 12 DAA rather than 20 DAA. In total, we identified 228 differentially accumulated protein spots representing 138 unique proteins by two-dimensional gel electrophoresis and tandem MALDI-TOF/TOF-MS in these three stages. Based on the results, we found some key proteins that are closely related to wheat grain development. The results of this analysis improve our understanding of the genetic mechanisms related to TaGW2-6A during wheat grain development as well as providing insights into the biological processes involved in seed formation. PMID:27446152

  3. Mucopolysaccharidosis: A New Variant?

    ERIC Educational Resources Information Center

    Primrose, D. A.

    1972-01-01

    Described is a possibly new variant of mucopolysaccharidosis characterized by progressive mental and motor deficiency, bone abnormalities, a generalized skin lesion, and abnormal mucopolysaccharides in the urine as seen in a 20-year-old female. (DB)

  4. Orientation to student placements: needs and benefits.

    PubMed

    Worrall, Katie

    2007-02-01

    A review of evidence on the benefits and challenges of student orientation is used in this article alongside experiences of orientation days on a children's ward to consider ways in which such programmes could be improved. Orientation to clinical placements can enhance learning by helping students to feel they fit in, reduce anxiety and increase motivation to learn through early identification of learning outcomes. However, there are challenges in the practical implementation of orientation including timing of students' starting dates, staff time, consistency and level of information and teaching. Increased involvement of individual mentors could improve orientation and optimise students' learning experiences.

  5. Industrial Orientation.

    ERIC Educational Resources Information Center

    Rasor, Leslie; Brooks, Valerie

    These eight modules for an industrial orientation class were developed by a project to design an interdisciplinary program of basic skills training for disadvantaged students in a Construction Technology Program (see Note). The Drafting module overviews drafting career opportunities, job markets, salaries, educational requirements, and basic…

  6. [Identification of auditory laterality by means of a new dichotic digit test in Spanish, and body laterality and spatial orientation in children with dyslexia and in controls].

    PubMed

    Olivares-García, M R; Peñaloza-López, Y R; García-Pedroza, F; Jesús-Pérez, S; Uribe-Escamilla, R; Jiménez-de la Sancha, S

    In this study, a new dichotic digit test in Spanish (NDDTS) was applied in order to identify auditory laterality. We also evaluated body laterality and spatial location using the Subirana test. Both the dichotic test and the Subirana test for body laterality and spatial location were applied in a group of 40 children with dyslexia and in a control group made up of 40 children who were paired according to age and gender. The results of the three evaluations were analysed using the SPSS 10 software application, with Pearson's chi-squared test. It was seen that 42.5% of the children in the group of dyslexics had mixed auditory laterality, compared to 7.5% in the control group (p < or = 0.05). Body laterality was mixed in 25% of dyslexic children and in 2.5% in the control group (p < or = 0.05) and there was 72.5% spatial disorientation in the group of dyslexics, whereas only 15% (p < or = 0.05) was found in the control group. The NDDTS proved to be a useful tool for demonstrating that mixed auditory laterality and auditory predominance of the left ear are linked to dyslexia. The results of this test exceed those obtained for body laterality. Spatial orientation is indeed altered in children with dyslexia. The importance of this finding makes it necessary to study the central auditory processes in all cases in order to define better rehabilitation strategies in Spanish-speaking children.

  7. Variation in Sexual Identification Among Behaviorally Bisexual Women in the Midwestern United States: Challenging the Established Methods for Collecting Data on Sexual Identity and Orientation.

    PubMed

    Baldwin, Aleta; Schick, Vanessa R; Dodge, Brian; van Der Pol, Barbara; Herbenick, Debby; Sanders, Stephanie A; Fortenberry, J Dennis

    2017-07-01

    Collecting information on sexual identity is critical to ensuring the visibility of minority populations who face stigmatization and discrimination related to sexual identities. However, it is challenging to capture the nuances of sexual identity with traditional survey research methods. Using a mixed-methods approach, we gathered data on the sexual identities of 80 behaviorally bisexual women in the Midwestern United States through an online survey. When provided different types of measures (e.g., open ended and fixed response) and different contexts in which to identify (e.g., private and public), participants varied in how they reported their sexual identities. Qualitative analysis of participant narratives around identity change finds partitioning and ranking of attraction is a key component in understanding behaviorally bisexual women's identities. We further identify a division regarding the desired outcomes of identity development processes. Given the multiple ways in which participants identified depending upon the type of measure and the context specified, and the variation in identification over time, results support reconsidering the capability of typical measures and methods used in survey research to capture sexual identity information. Additionally, findings highlight the utility of including multiple, context-specific measures of sexual identities in future research.

  8. Restaurant-based intervention to facilitate healthy eating choices and the identification of allergenic foods at a family-oriented resort and a campground.

    PubMed

    Tarro, Lucia; Aceves-Martins, Magaly; Tiñena, Yolanda; Parisi, Joan Lluís; Blasi, Xavier; Giralt, Montse; Llauradó, Elisabet; Solà, Rosa

    2017-05-05

    Restaurant-based interventions can be an enjoyable way to encourage healthier eating choices by all members of a family. Thus, the principal aims of this study were a) to promote healthy diets by increasing healthy food offerings and b) to increase the number of foods offered specifically as gluten-free and lactose-free and to inform patrons by including nutritional and allergen information that complies with Regulation 1169/2011 regarding the food served in restaurants, takeaways and snack bars. A restaurant-based intervention was implemented at 16 food establishments at 2 resorts (the Cambrils Park Resort and Camping Sangulí, Spain, from 2014 to 2015) based on the following 4 components: 1) providing nutritional and allergen analyses of the offered dishes, 2) increasing the number of healthy food choices, 3) identifying menu items associated with allergies and intolerance, and 4) training staff on healthy eating and allergens. Customer satisfaction regarding food aspects was assessed using surveys (10-point scale). Both resorts significantly increased their offerings of healthy dishes (28.6% to 44.7%; P = 0.003) and desserts with fruit (20% to 51.3%; P = 0.013), thus obtaining the Spanish Government's Mediterranean Diet certification. Additionally, both resorts obtained Catalan Celiac Association certification. Moreover, both resorts significantly increased their percentages of gluten-free dishes (2.1% to 50.5%; P < 0.001) and lactose-free dishes (5.5% to 37.5%; P < 0.001) after the intervention. Customer satisfaction increased (mean ± standard deviation) from 6.9 ± 1.6 to 8.5 ± 1.5 (P < 0.001). This restaurant-based intervention expanded the number of healthy and allergen-free foods offered in a family-oriented holiday resort environment to encourage healthy food choices, resulting in increased customer satisfaction.

  9. Adipokinetic hormone receptor gene identification and its role in triacylglycerol mobilization and sexual behavior in the oriental fruit fly (Bactrocera dorsalis).

    PubMed

    Hou, Qiu-Li; Chen, Er-Hu; Jiang, Hong-Bo; Wei, Dan-Dan; Gui, Shun-Hua; Wang, Jin-Jun; Smagghe, Guy

    2017-09-15

    Energy homeostasis requires continuous compensation for fluctuations in energy expenditure and availability of food resources. In insects, energy mobilization is under control of the adipokinetic hormone (AKH) where it is regulating the nutritional status by supporting the mobilization of lipids. In this study, we characterized the gene coding for the AKH receptor (AKHR) and investigated its function in the oriental fruit fly (Bactrocera dorsalis) that is economically one of the most important pest insects of tropical and subtropical fruit. Bacdo-AKHR is a typical G protein-coupled receptor (GPCR) and phylogenetic analysis confirmed that Bacdo-AKHR is closely related to insect AKHRs from other species. When expressed in Chinese hamster ovary (CHO) cells, Bacdo-AKHR exhibited a high sensitivity and selectivity for AKH peptide (EC50 = 19.3 nM). Using qPCR, the developmental stage and tissue-specific expression profiles demonstrated that Bacdo-AKHR was highly expressed in both the larval and adult stages, and also specifically in the fat body and midgut of the adult with no difference in sex. To investigate the role of AKHR in B. dorsalis, RNAi assays were performed with dsRNA against Bacdo-AKHR in adult flies of both sexes and under starvation and feeding condition. As major results, the knockdown of this gene resulted in triacylglycerol (TAG) accumulation. With RNAi-males, we observed a severe decrease in their sexual courtship activity when starved, but there was a partial rescue in copulation when refed. Also in RNAi-males, the tethered-flight duration declined compared with the control group when starved, which is confirming the dependency on energy metabolism. In RNAi-females, the sexual behavior was not affected, but their fecundity was decreased. Our findings indicate an interesting role of AKHR in the sexual behavior of males specifically. The effects are associated with TAG accumulation, and we also reported that the conserved role of AKH-mediated system

  10. Chapter 4: Variant descriptions

    Treesearch

    Duncan C. Lutes; Donald C. E. Robinson

    2003-01-01

    The Fire and Fuels Extension (FFE) to the Forest Vegetation Simulator (FVS) simulates fuel dynamics and potential fire behavior over time, in the context of stand development and management. This report documents differences between geographic variants of the FFE. It is a companion document to the FFE "Model Description" and "User's Guide."...

  11. Orienting hypnosis.

    PubMed

    Hope, Anna E; Sugarman, Laurence I

    2015-01-01

    This article presents a new frame for understanding hypnosis and its clinical applications. Despite great potential to transform health and care, hypnosis research and clinical integration is impaired in part by centuries of misrepresentation and ignorance about its demonstrated efficacy. The authors contend that advances in the field are primarily encumbered by the lack of distinct boundaries and definitions. Here, hypnosis, trance, and mind are all redefined and grounded in biological, neurological, and psychological phenomena. Solutions are proposed for boundary and language problems associated with hypnosis. The biological role of novelty stimulating an orienting response that, in turn, potentiates systemic plasticity forms the basis for trance. Hypnosis is merely the skill set that perpetuates and influences trance. This formulation meshes with many aspects of Milton Erickson's legacy and Ernest Rossi's recent theory of mind and health. Implications of this hypothesis for clinical skills, professional training, and research are discussed.

  12. EDITORIAL: Optical orientation Optical orientation

    NASA Astrophysics Data System (ADS)

    SAME ADDRESS *, Yuri; Landwehr, Gottfried

    2008-11-01

    priority of the discovery in the literature, which was partly caused by the existence of the Iron Curtain. I had already enjoyed contact with Boris in the 1980s when the two volumes of Landau Level Spectroscopy were being prepared [2]. He was one of the pioneers of magneto-optics in semiconductors. In the 1950s the band structure of germanium and silicon was investigated by magneto-optical methods, mainly in the United States. No excitonic effects were observed and the band structure parameters were determined without taking account of excitons. However, working with cuprous oxide, which is a direct semiconductor with a relative large energy gap, Zakharchenya and his co-worker Seysan showed that in order to obtain correct band structure parameters, it is necessary to take excitons into account [3]. About 1970 Boris started work on optical orientation. Early work by Hanle in Germany in the 1920s on the depolarization of luminescence in mercury vapour by a transverse magnetic field was not appreciated for a long time. Only in the late 1940s did Kastler and co-workers in Paris begin a systematic study of optical pumping, which led to the award of a Nobel prize. The ideas of optical pumping were first applied by Georges Lampel to solid state physics in 1968. He demonstrated optical orientation of free carriers in silicon. The detection method was nuclear magnetic resonance; optically oriented free electrons dynamically polarized the 29Si nuclei of the host lattice. The first optical detection of spin orientation was demonstrated by with the III-V semiconductor GaSb by Parsons. Due to the various interaction mechanisms of spins with their environment, the effects occurring in semiconductors are naturally more complex than those in atoms. Optical detection is now the preferred method to detect spin alignment in semiconductors. The orientation of spins in crystals pumped with circularly polarized light is deduced from the degree of circular polarization of the recombination

  13. A Bioinformatics Workflow for Variant Peptide Detection in Shotgun Proteomics*

    PubMed Central

    Li, Jing; Su, Zengliu; Ma, Ze-Qiang; Slebos, Robbert J. C.; Halvey, Patrick; Tabb, David L.; Liebler, Daniel C.; Pao, William; Zhang, Bing

    2011-01-01

    Shotgun proteomics data analysis usually relies on database search. However, commonly used protein sequence databases do not contain information on protein variants and thus prevent variant peptides and proteins from been identified. Including known coding variations into protein sequence databases could help alleviate this problem. Based on our recently published human Cancer Proteome Variation Database, we have created a protein sequence database that comprehensively annotates thousands of cancer-related coding variants collected in the Cancer Proteome Variation Database as well as noncancer-specific ones from the Single Nucleotide Polymorphism Database (dbSNP). Using this database, we then developed a data analysis workflow for variant peptide identification in shotgun proteomics. The high risk of false positive variant identifications was addressed by a modified false discovery rate estimation method. Analysis of colorectal cancer cell lines SW480, RKO, and HCT-116 revealed a total of 81 peptides that contain either noncancer-specific or cancer-related variations. Twenty-three out of 26 variants randomly selected from the 81 were confirmed by genomic sequencing. We further applied the workflow on data sets from three individual colorectal tumor specimens. A total of 204 distinct variant peptides were detected, and five carried known cancer-related mutations. Each individual showed a specific pattern of cancer-related mutations, suggesting potential use of this type of information for personalized medicine. Compatibility of the workflow has been tested with four popular database search engines including Sequest, Mascot, X!Tandem, and MyriMatch. In summary, we have developed a workflow that effectively uses existing genomic data to enable variant peptide detection in proteomics. PMID:21389108

  14. Variants of Uncertainty

    DTIC Science & Technology

    1981-05-15

    Variants of Uncertainty Daniel Kahneman University of British Columbia Amos Tversky Stanford University DTI-C &%E-IECTE ~JUNO 1i 19 8 1j May 15, 1981... Dennett , 1979) in which different parts have ac- cess to different data, assign then different weights and hold different views of the situation...2robable and t..h1 provable. Oxford- Claredor Press, 1977. Dennett , D.C. Brainstorms. Hassocks: Harvester, 1979. Donchin, E., Ritter, W. & McCallum, W.C

  15. Efflux-mediated fluoroquinolone resistance in the multidrug-resistant Pseudomonas aeruginosa clinical isolate PA7: identification of a novel MexS variant involved in upregulation of the mexEF-oprN multidrug efflux operon

    PubMed Central

    Morita, Yuji; Tomida, Junko; Kawamura, Yoshiaki

    2014-01-01

    The emergence of multidrug-resistant Pseudomonas aeruginosa has become a serious problem in medical settings. P. aeruginosa clinical isolate PA7 is resistant to fluoroquinolones, aminoglycosides, and most β-lactams but not imipenem. In this study, enhanced efflux-mediated fluoroquinolone resistance of PA7 was shown to reflect increased expression of two resistance nodulation cell division (RND) -type multidrug efflux operons, mexEF-oprN and mexXY-oprA. Such a clinical isolate has rarely been reported because MexEF-OprN-overproducing mutants often increase susceptibility to aminoglycosides apparently owing to impairment of the MexXY system. A mutant of PA7 lacking three RND-type multidrug efflux operons (mexAB-oprM, mexEF-oprN, and mexXY-oprA) was susceptible to all anti-pseudomonas agents we tested, supporting an idea that these RND-type multidrug efflux transporters are molecular targets to overcome multidrug resistance in P. aeruginosa. mexEF-oprN-upregulation in P. aeruginosa PA7 was shown due to a MexS variant harboring the Valine-155 amino acid residue. This is the first genetic evidence shown that a MexS variant causes mexEF-oprN-upregulation in P. aeruginosa clinical isolates. PMID:25653649

  16. Variants of glycoside hydrolases

    DOEpatents

    Teter, Sarah; Ward, Connie; Cherry, Joel; Jones, Aubrey; Harris, Paul; Yi, Jung

    2017-07-11

    The present invention relates to variants of a parent glycoside hydrolase, comprising a substitution at one or more positions corresponding to positions 21, 94, 157, 205, 206, 247, 337, 350, 373, 383, 438, 455, 467, and 486 of amino acids 1 to 513 of SEQ ID NO: 2, and optionally further comprising a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2 a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2, wherein the variants have glycoside hydrolase activity. The present invention also relates to nucleotide sequences encoding the variant glycoside hydrolases and to nucleic acid constructs, vectors, and host cells comprising the nucleotide sequences.

  17. Variants of glycoside hydrolases

    DOEpatents

    Teter, Sarah; Ward, Connie; Cherry, Joel; Jones, Aubrey; Harris, Paul; Yi, Jung

    2013-02-26

    The present invention relates to variants of a parent glycoside hydrolase, comprising a substitution at one or more positions corresponding to positions 21, 94, 157, 205, 206, 247, 337, 350, 373, 383, 438, 455, 467, and 486 of amino acids 1 to 513 of SEQ ID NO: 2, and optionally further comprising a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2 a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2, wherein the variants have glycoside hydrolase activity. The present invention also relates to nucleotide sequences encoding the variant glycoside hydrolases and to nucleic acid constructs, vectors, and host cells comprising the nucleotide sequences.

  18. Variants of glycoside hydrolases

    DOEpatents

    Teter, Sarah [Davis, CA; Ward, Connie [Hamilton, MT; Cherry, Joel [Davis, CA; Jones, Aubrey [Davis, CA; Harris, Paul [Carnation, WA; Yi, Jung [Sacramento, CA

    2011-04-26

    The present invention relates to variants of a parent glycoside hydrolase, comprising a substitution at one or more positions corresponding to positions 21, 94, 157, 205, 206, 247, 337, 350, 373, 383, 438, 455, 467, and 486 of amino acids 1 to 513 of SEQ ID NO: 2, and optionally further comprising a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2 a substitution at one or more positions corresponding to positions 8, 22, 41, 49, 57, 113, 193, 196, 226, 227, 246, 251, 255, 259, 301, 356, 371, 411, and 462 of amino acids 1 to 513 of SEQ ID NO: 2, wherein the variants have glycoside hydrolase activity. The present invention also relates to nucleotide sequences encoding the variant glycoside hydrolases and to nucleic acid constructs, vectors, and host cells comprising the nucleotide sequences.

  19. Enhanced variants of IDE algorithm

    NASA Astrophysics Data System (ADS)

    Bujok, Petr

    2017-07-01

    The performance of a new mechanism applied on a differential evolution algorithm with an individual-dependent mechanism (IDE) is studied experimentally. Three new IDE variants are proposed and compared with the original IDE variant. The performance of all studied algorithms was compared on each problem of CEC 2015 test suite. The newly proposed IDE variants mostly outperformed the original IDE variant significantly. The results show that the best results are for the newly proposed IDE variant with an enhanced mutation scheme and IDE with the control of the population diversity.

  20. Association of genetic variants with diabetic nephropathy.

    PubMed

    Rizvi, Saliha; Raza, Syed Tasleem; Mahdi, Farzana

    2014-12-15

    Diabetic nephropathy accounts for the most serious microvascular complication of diabetes mellitus. It is suggested that the prevalence of diabetic nephropathy will continue to increase in future posing a major challenge to the healthcare system resulting in increased morbidity and mortality. It occurs as a result of interaction between both genetic and environmental factors in individuals with both type 1 and type 2 diabetes. Genetic susceptibility has been proposed as an important factor for the development and progression of diabetic nephropathy, and various research efforts are being executed worldwide to identify the susceptibility gene for diabetic nephropathy. Numerous single nucleotide polymorphisms have been found in various genes giving rise to various gene variants which have been found to play a major role in genetic susceptibility to diabetic nephropathy. The risk of developing diabetic nephropathy is increased several times by inheriting risk alleles at susceptibility loci of various genes like ACE, IL, TNF-α, COL4A1, eNOS, SOD2, APOE, GLUT, etc. The identification of these genetic variants at a biomarker level could thus, allow the detection of those individuals at high risk for diabetic nephropathy which could thus help in the treatment, diagnosis and early prevention of the disease. The present review discusses about the various gene variants found till date to be associated with diabetic nephropathy.

  1. Novel RNA variants in colorectal cancers

    PubMed Central

    Alagaratnam, Sharmini; Zhao, Sen; Nome, Torfinn; Løvf, Marthe; Bakken, Anne C.; Hektoen, Merete; Sveen, Anita; Lothe, Ragnhild A.; Skotheim, Rolf I.

    2015-01-01

    With an annual estimated incidence of 1.4 million, and a five-year survival rate of 60%, colorectal cancer (CRC) is a major clinical burden. To identify novel RNA variants in CRC, we analyzed exon-level microarray expression data from a cohort of 202 CRCs. We nominated 25 genes with increased expression of their 3′ parts in at least one cancer sample each. To efficiently investigate underlying transcript structures, we developed an approach using rapid amplification of cDNA ends followed by high throughput sequencing (RACE-seq). RACE products from the targeted genes in 23 CRC samples were pooled together and sequenced. We identified VWA2-TCF7L2, DHX35-BPIFA2 and CASZ1-MASP2 as private fusion events, and novel transcript structures for 17 of the 23 other candidate genes. The high-throughput approach facilitated identification of CRC specific RNA variants. These include a recurrent read-through fusion transcript between KLK8 and KLK7, and a splice variant of S100A2. Both of these were overrepresented in CRC tissue and cell lines from external RNA-seq datasets. PMID:26474385

  2. Mapping of Epitopes Occurring in Bovine α(s1)-Casein Variants by Peptide Microarray Immunoassay.

    PubMed

    Lisson, Maria; Erhardt, Georg

    2016-01-01

    Immunoglobulin E epitope mapping of milk proteins reveals important information about their immunologic properties. Genetic variants of αS1-casein, one of the major allergens in bovine milk, are until now not considered when discussing the allergenic potential. Here we describe the complete procedure to assess the allergenicity of αS1-casein variants B and C, which are frequent in most breeds, starting from milk with identification and purification of casein variants by isoelectric focusing (IEF) and anion-exchange chromatography, followed by in vitro gastrointestinal digestion of the casein variants, identification of the resulting peptides by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), in silico analysis of the variant-specific peptides as allergenic epitopes, and determination of their IgE-binding properties by microarray immunoassay with cow's milk allergic human sera.

  3. Personal Factors in Organizational Identification

    ERIC Educational Resources Information Center

    Hall, Douglas T.; And Others

    1970-01-01

    Results of a study suggested possibility of a process whereby (1) service-oriented individuals are attracted to and recruited by the Forest Service, (2) service-oriented members are likely to identify strongly with the Service, and (3) this identification results in intrinsic need satisfactions. (Author/KJ)

  4. Personal Factors in Organizational Identification

    ERIC Educational Resources Information Center

    Hall, Douglas T.; And Others

    1970-01-01

    Results of a study suggested possibility of a process whereby (1) service-oriented individuals are attracted to and recruited by the Forest Service, (2) service-oriented members are likely to identify strongly with the Service, and (3) this identification results in intrinsic need satisfactions. (Author/KJ)

  5. Targeted exome sequencing for the identification of a protective variant against Internet gaming disorder at rs2229910 of neurotrophic tyrosine kinase receptor, type 3 (NTRK3): A pilot study

    PubMed Central

    Kim, Jeong-Yu; Jeong, Jo-Eun; Rhee, Je-Keun; Cho, Hyun; Chun, Ji-Won; Kim, Tae-Min; Choi, Sam-Wook; Choi, Jung-Seok; Kim, Dai-Jin

    2016-01-01

    Background and aims Internet gaming disorder (IGD) has gained recognition as a potential new diagnosis in the fifth revision of the Diagnostic and Statistical Manual of Mental Disorders, but genetic evidence supporting this disorder remains scarce. Methods In this study, targeted exome sequencing was conducted in 30 IGD patients and 30 control subjects with a focus on genes linked to various neurotransmitters associated with substance and non-substance addictions, depression, and attention deficit hyperactivity disorder. Results rs2229910 of neurotrophic tyrosine kinase receptor, type 3 (NTRK3) was the only single nucleotide polymorphism (SNP) that exhibited a significantly different minor allele frequency in IGD subjects compared to controls (p = .01932), suggesting that this SNP has a protective effect against IGD (odds ratio = 0.1541). The presence of this potentially protective allele was also associated with less time spent on Internet gaming and lower scores on the Young’s Internet Addiction Test and Korean Internet Addiction Proneness Scale for Adults. Conclusions The results of this first targeted exome sequencing study of IGD subjects indicate that rs2229910 of NTRK3 is a genetic variant that is significantly related to IGD. These findings may have significant implications for future research investigating the genetics of IGD and other behavioral addictions. PMID:27826991

  6. Targeted exome sequencing for the identification of a protective variant against Internet gaming disorder at rs2229910 of neurotrophic tyrosine kinase receptor, type 3 (NTRK3): A pilot study.

    PubMed

    Kim, Jeong-Yu; Jeong, Jo-Eun; Rhee, Je-Keun; Cho, Hyun; Chun, Ji-Won; Kim, Tae-Min; Choi, Sam-Wook; Choi, Jung-Seok; Kim, Dai-Jin

    2016-12-01

    Background and aims Internet gaming disorder (IGD) has gained recognition as a potential new diagnosis in the fifth revision of the Diagnostic and Statistical Manual of Mental Disorders, but genetic evidence supporting this disorder remains scarce. Methods In this study, targeted exome sequencing was conducted in 30 IGD patients and 30 control subjects with a focus on genes linked to various neurotransmitters associated with substance and non-substance addictions, depression, and attention deficit hyperactivity disorder. Results rs2229910 of neurotrophic tyrosine kinase receptor, type 3 (NTRK3) was the only single nucleotide polymorphism (SNP) that exhibited a significantly different minor allele frequency in IGD subjects compared to controls (p = .01932), suggesting that this SNP has a protective effect against IGD (odds ratio = 0.1541). The presence of this potentially protective allele was also associated with less time spent on Internet gaming and lower scores on the Young's Internet Addiction Test and Korean Internet Addiction Proneness Scale for Adults. Conclusions The results of this first targeted exome sequencing study of IGD subjects indicate that rs2229910 of NTRK3 is a genetic variant that is significantly related to IGD. These findings may have significant implications for future research investigating the genetics of IGD and other behavioral addictions.

  7. Identification and characterization of a ross river virus variant that grows persistently in macrophages, shows altered disease kinetics in a mouse model, and exhibits resistance to type I interferon.

    PubMed

    Lidbury, Brett A; Rulli, Nestor E; Musso, Cristina M; Cossetto, Susan B; Zaid, Ali; Suhrbier, Andreas; Rothenfluh, Harald S; Rolph, Michael S; Mahalingam, Suresh

    2011-06-01

    Alphaviruses, such as chikungunya virus, o'nyong-nyong virus, and Ross River virus (RRV), cause outbreaks of human rheumatic disease worldwide. RRV is a positive-sense single-stranded RNA virus endemic to Australia and Papua New Guinea. In this study, we sought to establish an in vitro model of RRV evolution in response to cellular antiviral defense mechanisms. RRV was able to establish persistent infection in activated macrophages, and a small-plaque variant (RRV(PERS)) was isolated after several weeks of culture. Nucleotide sequence analysis of RRV(PERS) found several nucleotide differences in the nonstructural protein (nsP) region of the RRV(PERS) genome. A point mutation was also detected in the E2 gene. Compared to the parent virus (RRV-T48), RRV(PERS) showed significantly enhanced resistance to beta interferon (IFN-β)-stimulated antiviral activity. RRV(PERS) infection of RAW 264.7 macrophages induced lower levels of IFN-β expression and production than infection with RRV-T48. RRV(PERS) was also able to inhibit type I IFN signaling. Mice infected with RRV(PERS) exhibited significantly enhanced disease severity and mortality compared to mice infected with RRV-T48. These results provide strong evidence that the cellular antiviral response can direct selective pressure for viral sequence evolution that impacts on virus fitness and sensitivity to alpha/beta IFN (IFN-α/β).

  8. Identification of two novel HLA-A*02 variants, A*02:319 and A*02:01:64, in two Taiwanese marrow stem cell donors by sequence-based typing.

    PubMed

    Yang, K L; Lee, S K; Yang, S Y; Kao, R H; Lin, C L; Lin, P Y

    2012-06-01

    We report here two novel variants of HLA-A*02 allele, A*02:319 and A*02:01:64, discovered in two Taiwanese unrelated volunteer bone marrow donors by sequence-based typing (SBT) method. The DNA sequence of A*02:319 is identical to A*02:07 in exons 2 and 3 but varies with one nucleotide at codon 9 (TTC->TCC). The variation caused one amino acid substitution at residue 9 (F->S). On the other hand, the DNA sequence of A*02:01:64 is identical to the sequence of A*02:01:01:01 in exons 2 and 3 except a silent mutation at codon 114 (CAC->CAT). The probable HLA-A, HLA-B and HLA-DRB1 haplotypes in association with A*02:319 and A*02:01:64 were deduced as A*02:319-B*46:01-DRB1*04 and A*02:01:64-B*38:02-DRB1*16:02, respectively.

  9. Improving coeliac disease risk prediction by testing non-HLA variants additional to HLA variants

    PubMed Central

    Romanos, Jihane; Rosén, Anna; Kumar, Vinod; Trynka, Gosia; Franke, Lude; Szperl, Agata; Gutierrez-Achury, Javier; van Diemen, Cleo C; Kanninga, Roan; Jankipersadsing, Soesma A; Steck, Andrea; Eisenbarth, Georges; van Heel, David A; Cukrowska, Bozena; Bruno, Valentina; Mazzilli, Maria Cristina; Núñez, Concepcion; Bilbao, Jose Ramon; Mearin, M Luisa; Barisani, Donatella; Rewers, Marian; Norris, Jill M; Ivarsson, Anneli; Boezen, H Marieke; Liu, Edwin; Wijmenga, Cisca

    2014-01-01

    Background The majority of coeliac disease (CD) patients are not being properly diagnosed and therefore remain untreated, leading to a greater risk of developing CD-associated complications. The major genetic risk heterodimer, HLA-DQ2 and DQ8, is already used clinically to help exclude disease. However, approximately 40% of the population carry these alleles and the majority never develop CD. Objective We explored whether CD risk prediction can be improved by adding non-HLA-susceptible variants to common HLA testing. Design We developed an average weighted genetic risk score with 10, 26 and 57 single nucleotide polymorphisms (SNP) in 2675 cases and 2815 controls and assessed the improvement in risk prediction provided by the non-HLA SNP. Moreover, we assessed the transferability of the genetic risk model with 26 non-HLA variants to a nested case–control population (n=1709) and a prospective cohort (n=1245) and then tested how well this model predicted CD outcome for 985 independent individuals. Results Adding 57 non-HLA variants to HLA testing showed a statistically significant improvement compared to scores from models based on HLA only, HLA plus 10 SNP and HLA plus 26 SNP. With 57 non-HLA variants, the area under the receiver operator characteristic curve reached 0.854 compared to 0.823 for HLA only, and 11.1% of individuals were reclassified to a more accurate risk group. We show that the risk model with HLA plus 26 SNP is useful in independent populations. Conclusions Predicting risk with 57 additional non-HLA variants improved the identification of potential CD patients. This demonstrates a possible role for combined HLA and non-HLA genetic testing in diagnostic work for CD. PMID:23704318

  10. Symmetric Euler orientation representations for orientational averaging.

    PubMed

    Mayerhöfer, Thomas G

    2005-09-01

    A new kind of orientation representation called symmetric Euler orientation representation (SEOR) is presented. It is based on a combination of the conventional Euler orientation representations (Euler angles) and Hamilton's quaternions. The properties of the SEORs concerning orientational averaging are explored and compared to those of averaging schemes that are based on conventional Euler orientation representations. To that aim, the reflectance of a hypothetical polycrystalline material with orthorhombic crystal symmetry was calculated. The calculation was carried out according to the average refractive index theory (ARIT [T.G. Mayerhöfer, Appl. Spectrosc. 56 (2002) 1194]). It is shown that the use of averaging schemes based on conventional Euler orientation representations leads to a dependence of the result from the specific Euler orientation representation that was utilized and from the initial position of the crystal. The latter problem can be overcome partly by the introduction of a weighing factor, but only for two-axes-type Euler orientation representations. In case of a numerical evaluation of the average, a residual difference remains also if a two-axes type Euler orientation representation is used despite of the utilization of a weighing factor. In contrast, this problem does not occur if a symmetric Euler orientation representation is used as a matter of principle, while the result of the averaging for both types of orientation representations converges with increasing number of orientations considered in the numerical evaluation. Additionally, the use of a weighing factor and/or non-equally spaced steps in the numerical evaluation of the average is not necessary. The symmetrical Euler orientation representations are therefore ideally suited for the use in orientational averaging procedures.

  11. Group B streptococcal opacity variants.

    PubMed Central

    Pincus, S H; Cole, R L; Wessels, M R; Corwin, M D; Kamanga-Sollo, E; Hayes, S F; Cieplak, W; Swanson, J

    1992-01-01

    Colony opacity variants were detected for type III group B streptococci (GBS). Transparent colonies predominate in the parent GBS, with occasional colonies having opaque portions. Two stable opaque variants (1.1 and 1.5) were compared with three transparent clones (1.2, 1.3, and 1.4). All grew well on blood agar and on GC medium, but variant 1.1 failed to grow on Todd-Hewitt medium. Scanning and transmission electron microscopy demonstrated that colony opacity correlated with bacterial aggregation status, with opaque variants forming longer and more organized chains. Opaque-transparent switches were observed in both directions for most variants, with transparent to opaque noted most frequently, but 1.5 did not switch at all. Switching of the opacity phenotype was observed both in vitro and in neonatal mice. Relationships between colony opacity and several cell surface phenomena were explored. (i) Opaque variant 1.1 had two surface proteins (46 and 75 kDa) that were either unique or greatly overexpressed. (ii) Variant 1.1 was deficient in type III polysaccharide, while 1.5 lacked group B antigen. Diminished capsular polysaccharide of variant 1.1 was reflected in reduced negative electrophoretic mobility and in increased buoyant density. (iii) Transparent variant colonies growing closest to a penicillin disk were opaque, but colonial variants did not differ in their sensitivity to penicillin. These data indicate that GBS can exist in both opaque and transparent forms, with opaque appearance occurring by multiple routes. Opaque variants grow poorly on Todd-Hewitt medium generally used for isolation of GBS, so any possible relationships between opacity variation and pathogenesis of GBS infection are unknown. Images PMID:1592825

  12. Variants of windmill nystagmus.

    PubMed

    Choi, Kwang-Dong; Shin, Hae Kyung; Kim, Ji-Soo; Kim, Sung-Hee; Choi, Jae-Hwan; Kim, Hyo-Jung; Zee, David S

    2016-07-01

    Windmill nystagmus is characterized by a clock-like rotation of the beating direction of a jerk nystagmus suggesting separate horizontal and vertical oscillators, usually 90° out of phase. We report oculographic characteristics in three patients with variants of windmill nystagmus in whom the common denominator was profound visual loss due to retinal diseases. Two patients showed a clock-like pattern, while in the third, the nystagmus was largely diagonal (in phase or 180° out of phase) but also periodically changed direction by 180°. We hypothesize that windmill nystagmus is a unique manifestation of "eye movements of the blind." It emerges when the central structures, including the cerebellum, that normally keep eye movements calibrated and gaze steady can no longer perform their task, because they are deprived of the retinal image motion that signals a need for adaptive recalibration.

  13. Association of breast cancer risk in BRCA1 and BRCA2 mutation carriers with genetic variants showing differential allelic expression: identification of a modifier of breast cancer risk at locus 11q22.3.

    PubMed

    Hamdi, Yosr; Soucy, Penny; Kuchenbaeker, Karoline B; Pastinen, Tomi; Droit, Arnaud; Lemaçon, Audrey; Adlard, Julian; Aittomäki, Kristiina; Andrulis, Irene L; Arason, Adalgeir; Arnold, Norbert; Arun, Banu K; Azzollini, Jacopo; Bane, Anita; Barjhoux, Laure; Barrowdale, Daniel; Benitez, Javier; Berthet, Pascaline; Blok, Marinus J; Bobolis, Kristie; Bonadona, Valérie; Bonanni, Bernardo; Bradbury, Angela R; Brewer, Carole; Buecher, Bruno; Buys, Saundra S; Caligo, Maria A; Chiquette, Jocelyne; Chung, Wendy K; Claes, Kathleen B M; Daly, Mary B; Damiola, Francesca; Davidson, Rosemarie; De la Hoya, Miguel; De Leeneer, Kim; Diez, Orland; Ding, Yuan Chun; Dolcetti, Riccardo; Domchek, Susan M; Dorfling, Cecilia M; Eccles, Diana; Eeles, Ros; Einbeigi, Zakaria; Ejlertsen, Bent; Engel, Christoph; Gareth Evans, D; Feliubadalo, Lidia; Foretova, Lenka; Fostira, Florentia; Foulkes, William D; Fountzilas, George; Friedman, Eitan; Frost, Debra; Ganschow, Pamela; Ganz, Patricia A; Garber, Judy; Gayther, Simon A; Gerdes, Anne-Marie; Glendon, Gord; Godwin, Andrew K; Goldgar, David E; Greene, Mark H; Gronwald, Jacek; Hahnen, Eric; Hamann, Ute; Hansen, Thomas V O; Hart, Steven; Hays, John L; Hogervorst, Frans B L; Hulick, Peter J; Imyanitov, Evgeny N; Isaacs, Claudine; Izatt, Louise; Jakubowska, Anna; James, Paul; Janavicius, Ramunas; Jensen, Uffe Birk; John, Esther M; Joseph, Vijai; Just, Walter; Kaczmarek, Katarzyna; Karlan, Beth Y; Kets, Carolien M; Kirk, Judy; Kriege, Mieke; Laitman, Yael; Laurent, Maïté; Lazaro, Conxi; Leslie, Goska; Lester, Jenny; Lesueur, Fabienne; Liljegren, Annelie; Loman, Niklas; Loud, Jennifer T; Manoukian, Siranoush; Mariani, Milena; Mazoyer, Sylvie; McGuffog, Lesley; Meijers-Heijboer, Hanne E J; Meindl, Alfons; Miller, Austin; Montagna, Marco; Mulligan, Anna Marie; Nathanson, Katherine L; Neuhausen, Susan L; Nevanlinna, Heli; Nussbaum, Robert L; Olah, Edith; Olopade, Olufunmilayo I; Ong, Kai-Ren; Oosterwijk, Jan C; Osorio, Ana; Papi, Laura; Park, Sue Kyung; Pedersen, Inge Sokilde; Peissel, Bernard; Segura, Pedro Perez; Peterlongo, Paolo; Phelan, Catherine M; Radice, Paolo; Rantala, Johanna; Rappaport-Fuerhauser, Christine; Rennert, Gad; Richardson, Andrea; Robson, Mark; Rodriguez, Gustavo C; Rookus, Matti A; Schmutzler, Rita Katharina; Sevenet, Nicolas; Shah, Payal D; Singer, Christian F; Slavin, Thomas P; Snape, Katie; Sokolowska, Johanna; Sønderstrup, Ida Marie Heeholm; Southey, Melissa; Spurdle, Amanda B; Stadler, Zsofia; Stoppa-Lyonnet, Dominique; Sukiennicki, Grzegorz; Sutter, Christian; Tan, Yen; Tea, Muy-Kheng; Teixeira, Manuel R; Teulé, Alex; Teo, Soo-Hwang; Terry, Mary Beth; Thomassen, Mads; Tihomirova, Laima; Tischkowitz, Marc; Tognazzo, Silvia; Toland, Amanda Ewart; Tung, Nadine; van den Ouweland, Ans M W; van der Luijt, Rob B; van Engelen, Klaartje; van Rensburg, Elizabeth J; Varon-Mateeva, Raymonda; Wappenschmidt, Barbara; Wijnen, Juul T; Rebbeck, Timothy; Chenevix-Trench, Georgia; Offit, Kenneth; Couch, Fergus J; Nord, Silje; Easton, Douglas F; Antoniou, Antonis C; Simard, Jacques

    2017-01-01

    Cis-acting regulatory SNPs resulting in differential allelic expression (DAE) may, in part, explain the underlying phenotypic variation associated with many complex diseases. To investigate whether common variants associated with DAE were involved in breast cancer susceptibility among BRCA1 and BRCA2 mutation carriers, a list of 175 genes was developed based of their involvement in cancer-related pathways. Using data from a genome-wide map of SNPs associated with allelic expression, we assessed the association of ~320 SNPs located in the vicinity of these genes with breast and ovarian cancer risks in 15,252 BRCA1 and 8211 BRCA2 mutation carriers ascertained from 54 studies participating in the Consortium of Investigators of Modifiers of BRCA1/2. We identified a region on 11q22.3 that is significantly associated with breast cancer risk in BRCA1 mutation carriers (most significant SNP rs228595 p = 7 × 10(-6)). This association was absent in BRCA2 carriers (p = 0.57). The 11q22.3 region notably encompasses genes such as ACAT1, NPAT, and ATM. Expression quantitative trait loci associations were observed in both normal breast and tumors across this region, namely for ACAT1, ATM, and other genes. In silico analysis revealed some overlap between top risk-associated SNPs and relevant biological features in mammary cell data, which suggests potential functional significance. We identified 11q22.3 as a new modifier locus in BRCA1 carriers. Replication in larger studies using estrogen receptor (ER)-negative or triple-negative (i.e., ER-, progesterone receptor-, and HER2-negative) cases could therefore be helpful to confirm the association of this locus with breast cancer risk.

  14. Identification of bla(IMP-22) in Pseudomonas spp. in urban wastewater and nosocomial environments: biochemical characterization of a new IMP metallo-enzyme variant and its genetic location.

    PubMed

    Pellegrini, Cristina; Mercuri, Paola Sandra; Celenza, Giuseppe; Galleni, Moreno; Segatore, Bernardetta; Sacchetti, Elisa; Volpe, Roberto; Amicosante, Gianfranco; Perilli, Mariagrazia

    2009-05-01

    The aim of the study was the biochemical characterization of a new variant of the metallo-beta-lactamase, IMP-22. Moreover, the genetic environment of the bla(IMP-22) gene was investigated in Pseudomonas fluorescens and Pseudomonas aeruginosa collected from urban wastewater and a teaching hospital in L'Aquila, Italy. Molecular characterization of genetic elements was carried out by PCR and DNA sequencing methods. The new enzyme was purified from recombinant Escherichia coli BL21(DE)Rosetta/pBC-SK/IMP-22. Steady-state kinetic parameters (K(m) and V(max)) were determined for a large pattern of substrates. A new IMP metallo-beta-lactamase gene was found in a class 1 integron and in one case, in a plasmid of Pseudomonas spp. The bla(IMP-22) encodes for a pre-protein of 246 amino acids and the N-terminus of the mature beta-lactamase (NH(2)-PDLK) was also determined. The molecular mass and pI were 24 930 Da and 6.2, respectively. On the basis of the kinetic parameters calculated (K(m) and V(max)), IMP-22 was found to hydrolyse narrow- and extended-spectrum beta-lactams. Enzyme activity was found to be inhibited by metal chelators such as EDTA, 1,10-o-phenathroline and dipicolinic acid with an IC(50) of 800, 750 and 300 microM, respectively. The finding of the bla(IMP-22) gene in P. fluorescens environmental strains and P. aeruginosa clinical isolate suggests the ongoing spread of bla(MBL) genes in several bacterial species and in different environments.

  15. Identification of a new structural variant of human apolipoprotein E, E2(Lys146 leads to Gln), in a type III hyperlipoproteinemic subject with the E3/2 phenotype.

    PubMed Central

    Rall, S C; Weisgraber, K H; Innerarity, T L; Bersot, T P; Mahley, R W; Blum, C B

    1983-01-01

    A type III hyperlipoproteinemic subject having the apolipoprotein E (apo E) phenotype E3/2 was identified. From isoelectric focusing experiments in conjunction with cysteamine treatment (a method that measures cysteine content in apo E), the E2 isoform of this subject was determined to have only one cysteine residue, in contrast to all previously studied E2 apoproteins, which had two cysteines. This single cysteine was shown to be at residue 112, the same site at which it occurs in apo E3. From amino acid and sequence analyses, it was determined that this apo E2 differed from apo E3 by the occurrence of glutamine rather than lysine at residue 146. When phospholipid X protein recombinants of the subject's isolated E3 and E2 isoforms were tested for their ability to bind to the human fibroblast apo-B,E receptor, it was found that the E3 bound normally (compared with an apo E3 control) but that the E2 had defective binding (approximately 40% of normal). Although they contained E3 as well as E2, the beta-very low density lipoproteins (beta-VLDL) from this subject were very similar in character to the beta-VLDL from an E2/2 type III hyperlipoproteinemic subject; similar subfractions could be obtained from each subject and were shown to have a similar ability to stimulate cholesteryl ester accumulation in mouse peritoneal macrophages. The new apo E2 variant has also been detected in a second type III hyperlipoproteinemic subject. Images PMID:6313758

  16. Biometrics, identification and surveillance.

    PubMed

    Lyon, David

    2008-11-01

    Governing by identity describes the emerging regime of a globalizing, mobile world. Governance depends on identification but identification increasingly depends on biometrics. This 'solution' to difficulties of verification is described and some technical weaknesses are discussed. The role of biometrics in classification systems is also considered and is shown to contain possible prejudice in relation to racialized criteria of identity. Lastly, the culture of biometric identification is shown to be limited to abstract data, artificially separated from the lived experience of the body including the orientation to others. It is proposed that creators of national ID systems in particular address these crucial deficiencies in their attempt to provide new modes of verification.

  17. Functional characterization of BRCA1 gene variants by mini-gene splicing assay.

    PubMed

    Steffensen, Ane Y; Dandanell, Mette; Jønson, Lars; Ejlertsen, Bent; Gerdes, Anne-Marie; Nielsen, Finn C; Hansen, Thomas vO

    2014-12-01

    Mutational screening of the breast cancer susceptibility gene BRCA1 leads to the identification of numerous pathogenic variants such as frameshift and nonsense variants, as well as large genomic rearrangements. The screening moreover identifies a large number of variants, for example, missense, silent, and intron variants, which are classified as variants of unknown clinical significance owing to the lack of causal evidence. Variants of unknown clinical significance can potentially have an impact on splicing and therefore functional examinations are warranted to classify whether these variants are pathogenic or benign. Here we validate a mini-gene splicing assay by comparing the results of 24 variants with previously published data from RT-PCR analysis on RNA from blood samples/lymphoblastoid cell lines. The analysis showed an overall concordance of 100%. In addition, we investigated 13 BRCA1 variants of unknown clinical significance or putative variants affecting splicing by in silico analysis and mini-gene splicing assay. Both the in silico analysis and mini-gene splicing assay classified six BRCA1 variants as pathogenic (c.80+1G>A, c.132C>T (p.=), c.213-1G>A, c.670+1delG, c.4185+1G>A, and c.5075-1G>C), whereas six BRCA1 variants were classified as neutral (c.-19-22_-19-21dupAT, c.302-15C>G, c.547+14delG, c.4676-20A>G, c.4987-21G>T, and c.5278-14C>G) and one BRCA1 variant remained unclassified (c.670+16G>A). In conclusion, our study emphasizes that in silico analysis and mini-gene splicing assays are important for the classification of variants, especially if no RNA is available from the patient. This knowledge is crucial for proper genetic counseling of patients and their family members.

  18. Functional characterization of BRCA1 gene variants by mini-gene splicing assay

    PubMed Central

    Steffensen, Ane Y; Dandanell, Mette; Jønson, Lars; Ejlertsen, Bent; Gerdes, Anne-Marie; Nielsen, Finn C; Hansen, Thomas vO

    2014-01-01

    Mutational screening of the breast cancer susceptibility gene BRCA1 leads to the identification of numerous pathogenic variants such as frameshift and nonsense variants, as well as large genomic rearrangements. The screening moreover identifies a large number of variants, for example, missense, silent, and intron variants, which are classified as variants of unknown clinical significance owing to the lack of causal evidence. Variants of unknown clinical significance can potentially have an impact on splicing and therefore functional examinations are warranted to classify whether these variants are pathogenic or benign. Here we validate a mini-gene splicing assay by comparing the results of 24 variants with previously published data from RT-PCR analysis on RNA from blood samples/lymphoblastoid cell lines. The analysis showed an overall concordance of 100%. In addition, we investigated 13 BRCA1 variants of unknown clinical significance or putative variants affecting splicing by in silico analysis and mini-gene splicing assay. Both the in silico analysis and mini-gene splicing assay classified six BRCA1 variants as pathogenic (c.80+1G>A, c.132C>T (p.=), c.213−1G>A, c.670+1delG, c.4185+1G>A, and c.5075−1G>C), whereas six BRCA1 variants were classified as neutral (c.-19-22_-19-21dupAT, c.302−15C>G, c.547+14delG, c.4676−20A>G, c.4987−21G>T, and c.5278−14C>G) and one BRCA1 variant remained unclassified (c.670+16G>A). In conclusion, our study emphasizes that in silico analysis and mini-gene splicing assays are important for the classification of variants, especially if no RNA is available from the patient. This knowledge is crucial for proper genetic counseling of patients and their family members. PMID:24667779

  19. Election 2016: Voting on Variants.

    PubMed

    Cho, Raymond J; Collisson, Eric A

    2016-07-01

    Genome sequencing studies increasingly identify variants of unknown significance in provocative genes. Kim and colleagues present a system with which to functionally annotate such variants in a high-throughput, biologically relevant series of assays. Cancer Discov; 6(7); 694-6. ©2016 AACRSee related article by Kim et al., p. 714.

  20. Human AZU-1 gene, variants thereof and expressed gene products

    DOEpatents

    Chen, Huei-Mei; Bissell, Mina

    2004-06-22

    A human AZU-1 gene, mutants, variants and fragments thereof. Protein products encoded by the AZU-1 gene and homologs encoded by the variants of AZU-1 gene acting as tumor suppressors or markers of malignancy progression and tumorigenicity reversion. Identification, isolation and characterization of AZU-1 and AZU-2 genes localized to a tumor suppressive locus at chromosome 10q26, highly expressed in nonmalignant and premalignant cells derived from a human breast tumor progression model. A recombinant full length protein sequences encoded by the AZU-1 gene and nucleotide sequences of AZU-1 and AZU-2 genes and variant and fragments thereof. Monoclonal or polyclonal antibodies specific to AZU-1, AZU-2 encoded protein and to AZU-1, or AZU-2 encoded protein homologs.

  1. Thinking in Orienteering.

    ERIC Educational Resources Information Center

    Johansen, Bjorn Tore

    1997-01-01

    A think-aloud technique, in which 20 orienteers verbalized their exact thoughts during orienteering, was used to examine the phenomenon of cognition during orienteering. Results indicate that orienteering is experienced as a task to be accomplished, a physical movement, and a dynamic process, and that thinking involves attuning perceptions to…

  2. Family studies to find rare high risk variants in migraine.

    PubMed

    Hansen, Rikke Dyhr; Christensen, Anne Francke; Olesen, Jes

    2017-12-01

    Migraine has long been known as a common complex disease caused by genetic and environmental factors. The pathophysiology and the specific genetic susceptibility are poorly understood. Common variants only explain a small part of the heritability of migraine. It is thought that rare genetic variants with bigger effect size may be involved in the disease. Since migraine has a tendency to cluster in families, a family approach might be the way to find these variants. This is also indicated by identification of migraine-associated loci in classical linkage-analyses in migraine families. A single migraine study using a candidate-gene approach was performed in 2010 identifying a rare mutation in the TRESK potassium channel segregating in a large family with migraine with aura, but this finding has later become questioned. The technologies of next-generation sequencing (NGS) now provides an affordable tool to investigate the genetic variation in the entire exome or genome. The family-based study design using NGS is described in this paper. We also review family studies using NGS that have been successful in finding rare variants in other common complex diseases in order to argue the promising application of a family approach to migraine. PubMed was searched to find studies that looked for rare genetic variants in common complex diseases through a family-based design using NGS, excluding studies looking for de-novo mutations, or using a candidate-gene approach and studies on cancer. All issues from Nature Genetics and PLOS genetics 2014, 2015 and 2016 (UTAI June) were screened for relevant papers. Reference lists from included and other relevant papers were also searched. For the description of the family-based study design using NGS an in-house protocol was used. Thirty-two successful studies, which covered 16 different common complex diseases, were included in this paper. We also found a single migraine study. Twenty-three studies found one or a few family specific

  3. Characterization of a Mycobacterium intracellulare Variant Strain by Molecular Techniques

    PubMed Central

    Menendez, M. C.; Palenque, E.; Navarro, M. C.; Nuñez, M. C.; Rebollo, M. J.; Garcia, M. J.

    2001-01-01

    This paper describes a Mycobacterium intracellulare variant strain causing an unusual infection. Several isolates obtained from an immunocompromised patient were identified as members of the Mycobacterium avium complex (MAC) by the commercial AccuProbe system and biochemical standard identification. Further molecular approaches were undertaken for a more accurate characterization of the bacteria. Up to seven different genomic sequences were analyzed, ranging from conserved mycobacterial genes such as 16S ribosomal DNA to MAC-specific genes such as mig (macrophage-induced gene). The results obtained identify the isolates as a variant of M. intracellulare, an example of the internal variability described for members of the MAC, particularly within that species. The application of other molecular approaches is recommended for more accurate identification of bacteria described as MAC members. PMID:11724827

  4. Characterization of a Mycobacterium intracellulare variant strain by molecular techniques.

    PubMed

    Menendez, M C; Palenque, E; Navarro, M C; Nuñez, M C; Rebollo, M J; Garcia, M J

    2001-12-01

    This paper describes a Mycobacterium intracellulare variant strain causing an unusual infection. Several isolates obtained from an immunocompromised patient were identified as members of the Mycobacterium avium complex (MAC) by the commercial AccuProbe system and biochemical standard identification. Further molecular approaches were undertaken for a more accurate characterization of the bacteria. Up to seven different genomic sequences were analyzed, ranging from conserved mycobacterial genes such as 16S ribosomal DNA to MAC-specific genes such as mig (macrophage-induced gene). The results obtained identify the isolates as a variant of M. intracellulare, an example of the internal variability described for members of the MAC, particularly within that species. The application of other molecular approaches is recommended for more accurate identification of bacteria described as MAC members.

  5. gag, vif, and nef Genes Contribute to the Homologous Viral Interference Induced by a Nonproducer Human Immunodeficiency Virus Type 1 (HIV-1) Variant: Identification of Novel HIV-1-Inhibiting Viral Protein Mutants

    PubMed Central

    D’Aloja, Paola; Olivetta, Eleonora; Bona, Roberta; Nappi, Filomena; Pedacchia, Daniela; Pugliese, Katherina; Ferrari, Giuliana; Verani, Paola; Federico, Maurizio

    1998-01-01

    We previously demonstrated that expression of the nonproducer F12-human immunodeficiency virus type 1 (HIV-1) variant induces a block in the replication of superinfecting HIV that does not depend on the down-regulation of CD4 HIV receptors. In order to individuate the gene(s) involved in F12-HIV-induced interference, vectors expressing each of the nine F12-HIV proteins were transfected in HIV-susceptible HeLa CD4 cells. Pools of cell clones stably producing each viral protein were infected with HIV-1, and virus release was measured in terms of reverse transcriptase activity in supernatants. We hereby demonstrate that HeLa CD4 cells expressing the F12-HIV gag, vif, or nef gene were resistant, to different degrees, to infection with T-cell-line-adapted HIV-1 strains. Conversely, expression of either the tat, rev, or vpu F12-HIV gene increased the rate of HIV release, and no apparent effects on HIV replication were observed in cells expressing either the F12-HIV vpr, pol, or env gene. No variation of CD4 exposure was detected in any of the uninfected HeLa CD4 pools. These data indicate that F12-HIV homologous viral interference is the consequence of the synergistic anti-HIV effects of Gag, Vif, and Nef proteins. Retrovirus vectors expressing F12-HIV vif or nef allowed us to further establish that the expression of each mutated protein (i) inhibits the replication of clinical HIV-1 isolates as well, (ii) impairs the infectivity of the virus released by cells chronically infected with HIV-1, and (iii) limitedly to F12-HIV Vif protein, induces HIV resistance in both vif-permissive and vif-nonpermissive cells. The levels of action of F12-HIV vif and nef anti-HIV effects were also determined. We observed that HIV virions emerging from the first viral cycle on F12-HIV vif-expressing cells, although released in unaltered amounts, had a strongly reduced ability to initiate the retrotranscription process when they reinfected parental HeLa CD4 cells. Differently, we observed

  6. Isotopic variants of light and heavy L-pyroglutamic acid succinimidyl esters as the derivatization reagents for DL-amino acid chiral metabolomics identification by liquid chromatography and electrospray ionization mass spectrometry.

    PubMed

    Mochizuki, Toshiki; Todoroki, Kenichiro; Inoue, Koichi; Min, Jun Zhe; Toyo'oka, Toshimasa

    2014-02-06

    L-Pyroglutamic acid succinimidyl ester (L-PGA-OSu) and its isotopic variant (L-PGA[d5]-OSu) were newly synthesized and evaluated as the chiral labeling reagents for the enantioseparation of amino acids, in terms of separation efficiency by reversed-phase chromatography and detection sensitivity by ESI-MS/MS. The enantiomers of amino acids were easily labeled with the reagents at 60°C within 10 min in an alkaline medium containing triethylamine. Although all the diastereomers derived from 18 proteolytic amino acids could not be satisfactorily separated, the pairs of 9 amino acids were completely separated by reversed-phase chromatography using the small particle (1.7 μm) ODS column (Rs=1.95-8.05). The characteristic daughter ions, i.e., m/z 84.04 and m/z 89.04, were detected from all the derivatives by the collision induced dissociation of the protonated molecular ions. A highly sensitive detection at a low-fmol level (0.5-3.2 fmol) was also obtained from the selected reaction monitoring (SRM) chromatograms. An isotope labeling strategy using light and heavy L-PGA-OSu for the differential analysis of the DL-amino acids in different sample groups is also presented in this paper. The differential analysis of biological sample (i.e., human serum) and food product (i.e., yogurt) were tried to demonstrate the efficiency of the proposed method. The ratios of the DL-amino acids in human serum samples, spiked with the different concentrations of D-amino acids, were determined by the procedures using L-PGA-OSu and L-PGA[d5]-OSu. The D/L ratios in the two sample groups at different concentrations of amino acids were similar to the theoretical values. Furthermore, the ratios of D/L-alanine values in different yogurt products were comparable to the ratios obtained from the d/l values using only light reagent (i.e., L-PGA-OSu). Consequently, the proposed strategy is useful for the differential analysis not only in biological samples but also in food products.

  7. Variant (Swine Origin) Influenza Viruses in Humans

    MedlinePlus

    ... Types Seasonal Avian Swine Variant Other Variant Influenza Viruses: Background and CDC Risk Assessment and Reporting Language: ... Background CDC Assessment Reporting Background On Variant Influenza Viruses Swine flu viruses do not normally infect humans. ...

  8. Cellobiohydrolase variants and polynucleotides encoding same

    DOEpatents

    Wogulis, Mark

    2013-09-24

    The present invention relates to variants of a parent cellobiohydrolase II. The present invention also relates to polynucleotides encoding the variants; nucleic acid constructs, vectors, and host cells comprising the polynucleotides; and methods of using the variants.

  9. Cellobiohydrolase variants and polynucleotides encoding same

    DOEpatents

    Wogulis, Mark

    2014-10-14

    The present invention relates to variants of a parent cellobiohydrolase II. The present invention also relates to polynucleotides encoding the variants; nucleic acid constructs, vectors, and host cells comprising the polynucleotides; and methods of using the variants.

  10. Cellobiohydrolase variants and polynucleotides encoding the same

    DOEpatents

    Wogulis, Mark

    2014-09-09

    The present invention relates to variants of a parent cellobiohydrolase. The present invention also relates to polynucleotides encoding the cellobiohydrolase variants; nucleic acid constructs, vectors, and host cells comprising the polynucleotides; and methods of using the cellobiohydrolase variants.

  11. Occurrence of Fuligo gyrosa Causing Slime Mold of Oriental Melon

    PubMed Central

    Choi, Hyo Won; Hong, Sung Kee; Lee, Young Kee; Lee, Su Heon

    2009-01-01

    Recently, a severe slime mold infestation affected oriental melon plants in fields in Chilgok county, Gyeongbuk province, Korea. Specimens were collected from the fields and examined for identification. A species of Myxomycetes, Fuligo gyrosa, was identified based on its morphological characteristics. This is the first report that F. gyrosa causes slime mold of oriental melon. PMID:23983540

  12. Occurrence of Fuligo gyrosa Causing Slime Mold of Oriental Melon.

    PubMed

    Kim, Wan Gyu; Choi, Hyo Won; Hong, Sung Kee; Lee, Young Kee; Lee, Su Heon

    2009-09-01

    Recently, a severe slime mold infestation affected oriental melon plants in fields in Chilgok county, Gyeongbuk province, Korea. Specimens were collected from the fields and examined for identification. A species of Myxomycetes, Fuligo gyrosa, was identified based on its morphological characteristics. This is the first report that F. gyrosa causes slime mold of oriental melon.

  13. Heteromorphic variants of chromosome 9

    PubMed Central

    2013-01-01

    Background Heterochromatic variants of pericentromere of chromosome 9 are reported and discussed since decades concerning their detailed structure and clinical meaning. However, detailed studies are scarce. Thus, here we provide the largest ever done molecular cytogenetic research based on >300 chromosome 9 heteromorphism carriers. Results In this study, 334 carriers of heterochromatic variants of chromosome 9 were included, being 192 patients from Western Europe and the remainder from Easter-European origin. A 3-color-fluorescence in situ hybridization (FISH) probe-set directed against for 9p12 to 9q13~21.1 (9het-mix) and 8 different locus-specific probes were applied for their characterization. The 9het-mix enables the characterization of 21 of the yet known 24 chromosome 9 heteromorphic patterns. In this study, 17 different variants were detected including five yet unreported; the most frequent were pericentric inversions (49.4%) followed by 9qh-variants (23.9%), variants of 9ph (11.4%), cenh (8.2%), and dicentric- (3.8%) and duplication-variants (3.3%). For reasons of simplicity, a new short nomenclature for the yet reported 24 heteromorphic patterns of chromosome 9 is suggested. Six breakpoints involved in four of the 24 variants could be narrowed down using locus-specific probes. Conclusions Based on this largest study ever done in carriers of chromosome 9 heteromorphisms, three of the 24 detailed variants were more frequently observed in Western than in Eastern Europe. Besides, there is no clear evidence that infertility is linked to any of the 24 chromosome 9 heteromorphic variants. PMID:23547710

  14. Variants of beta-glucosidase

    DOEpatents

    Fidantsef, Ana; Lamsa, Michael; Gorre-Clancy, Brian

    2009-12-29

    The present invention relates to variants of a parent beta-glucosidase, comprising a substitution at one or more positions corresponding to positions 142, 183, 266, and 703 of amino acids 1 to 842 of SEQ ID NO: 2 or corresponding to positions 142, 183, 266, and 705 of amino acids 1 to 844 of SEQ ID NO: 70, wherein the variant has beta-glucosidase activity. The present invention also relates to nucleotide sequences encoding the variant beta-glucosidases and to nucleic acid constructs, vectors, and host cells comprising the nucleotide sequences.

  15. Variants of beta-glucosidase

    SciTech Connect

    Fidantsef, Ana; Lamsa, Michael; Gorre-Clancy, Brian

    2015-07-14

    The present invention relates to variants of a parent beta-glucosidase, comprising a substitution at one or more positions corresponding to positions 142, 183, 266, and 703 of amino acids 1 to 842 of SEQ ID NO: 2 or corresponding to positions 142, 183, 266, and 705 of amino acids 1 to 844 of SEQ ID NO: 70, wherein the variant has beta-glucosidase activity. The present invention also relates to nucleotide sequences encoding the variant beta-glucosidases and to nucleic acid constructs, vectors, and host cells comprising the nucleotide sequences.

  16. Variants of beta-glucosidases

    SciTech Connect

    Fidantsef, Ana; Lamsa, Michael; Gorre-Clancy, Brian

    2014-10-07

    The present invention relates to variants of a parent beta-glucosidase, comprising a substitution at one or more positions corresponding to positions 142, 183, 266, and 703 of amino acids 1 to 842 of SEQ ID NO: 2 or corresponding to positions 142, 183, 266, and 705 of amino acids 1 to 844 of SEQ ID NO: 70, wherein the variant has beta-glucosidase activity. The present invention also relates to nucleotide sequences encoding the variant beta-glucosidases and to nucleic acid constructs, vectors, and host cells comprising the nucleotide sequences.

  17. Variants of beta-glucosidases

    DOEpatents

    Fidantsef, Ana; Lamsa, Michael; Clancy, Brian Gorre

    2008-08-19

    The present invention relates to variants of a parent beta-glucosidase, comprising a substitution at one or more positions corresponding to positions 142, 183, 266, and 703 of amino acids 1 to 842 of SEQ ID NO: 2 or corresponding to positions 142, 183, 266, and 705 of amino acids 1 to 844 of SEQ ID NO: 70, wherein the variant has beta-glucosidase activity. The present invention also relates to nucleotide sequences encoding the variant beta-glucosidases and to nucleic acid constructs, vectors, and host cells comprising the nucleotide sequences.

  18. Identification of an altered peptide ligand based on the endogenously presented, rheumatoid arthritis-associated, human cartilage glycoprotein-39(263–275) epitope: an MHC anchor variant peptide for immune modulation

    PubMed Central

    Boots, Annemieke MH; Hubers, Henk; Kouwijzer, Milou; den Hoed-van Zandbrink, Leontien; Westrek-Esselink, Bernice M; van Doorn, Cindy; Stenger, Rachel; Bos, Ebo S; van Lierop, Marie-jose C; Verheijden, Gijs F; Timmers, Cornelis M; van Staveren, Catharina J

    2007-01-01

    We sought to identify an altered peptide ligand (APL) based on the endogenously expressed synovial auto-epitope of human cartilage glycoprotein-39 (HC gp-39) for modulation of cognate, HLA-DR4-restricted T cells. For this purpose we employed a panel of well-characterized T cell hybridomas generated from HC gp-39-immunized HLA-DR4 transgenic mice. The hybridomas all respond to the HC gp-39(263–275) epitope when bound to HLA-DR4(B1*0401) but differ in their fine specificities. First, the major histocompatibility