WorldWideScience

Sample records for ccch-type zinc finger

  1. Comprehensive analysis of CCCH-type zinc finger gene family in citrus (Clementine mandarin) by genome-wide characterization.

    Science.gov (United States)

    Liu, Shengrui; Khan, Muhammad Rehman Gul; Li, Yongping; Zhang, Jinzhi; Hu, Chungen

    2014-10-01

    The CCCH-type zinc finger proteins comprise a large gene family of regulatory proteins and are widely distributed in eukaryotic organisms. The CCCH proteins have been implicated in multiple biological processes and environmental responses in plants. Little information is available, however, about CCCH genes in plants, especially in woody plants such as citrus. The release of the whole-genome sequence of citrus allowed us to perform a genome-wide analysis of CCCH genes and to compare the identified proteins with their orthologs in model plants. In this study, 62 CCCH genes and a total of 132 CCCH motifs were identified, and a comprehensive analysis including the chromosomal locations, phylogenetic relationships, functional annotations, gene structures and conserved motifs was performed. Distribution mapping revealed that 54 of the 62 CCCH genes are unevenly dispersed on the nine citrus chromosomes. Based on phylogenetic analysis and gene structural features, we constructed 5 subfamilies of 62 CCCH members and integrative subfamilies from citrus, Arabidopsis, and rice, respectively. Importantly, large numbers of SNPs and InDels in 26 CCCH genes were identified from Poncirus trifoliata and Fortunella japonica using whole-genome deep re-sequencing. Furthermore, citrus CCCH genes showed distinct temporal and spatial expression patterns in different developmental processes and in response to various stress conditions. Our comprehensive analysis of CleC3Hs is a valuable resource that further elucidates the roles of CCCH family members in plant growth and development. In addition, variants and comparative genomics analyses deepen our understanding of the evolution of the CCCH gene family and will contribute to further genetics and genomics studies of citrus and other plant species.

  2. 布氏锥虫未知CCCH-型锌指蛋白TbZC3H8功能特性的初步分析%Characterization of the hypothetical CCCH-type zinc-finger protein TbZC3H8 in Trypanosoma brucei

    Institute of Scientific and Technical Information of China (English)

    雷霁卿; 刘罗根; 郭学敏

    2013-01-01

    Objective To characterize the properties and functions of putative zinc-finger protein TbZC3H8 in Trypanosoma brucei.Methods Sequence analysis and motifs/domains prediction were performed through the protein database searches.An inducible RNAi cell line was generated to study the effect of TbZC3H8 repression on cell viability,and the RNAi knockdown efficiency was estimated by RT-QPCR and Western blot.The cell line ectopically expressing C-terminal mycTAP tagged TbZC3H8 was generated and used to identify the composition of TbZC3H8 protein complexes through the combination of tandem affinity purification and mass spectrometric analysis.Subcellular localization of the TbZC3H8 protein was determined by immunofluorescence microscopy.Results The CCCH zinc-finger motif of TbZC3H8 is highly conserved in kinetoplastid parasites.Repression of TbZC3H8 by RNAi resulted in the growth inhibition.The TbZC3H8 protein complex was found to contain both unknown proteins and RNA-binding proteins.TbZC3H8 was found to localize in the cytoplasm and the expression level was not changed upon the heat shock and serum starvation.Conclusion TbZC3H8 is essential for the growth of T.brucei.The binding of ZC3H8 with RNA-binding proteins suggested that ZC3H8 might play a role in RNA processing and metabolism.%目的 通过RNAi和蛋白复合物鉴定来初步分析布氏锥虫未知锌指蛋白TbZC3H8的特性及功能.方法 运用蛋白数据库和分析软件对TbZC3H8进行序列分析和结构域预测;构建RNAi诱导表达细胞株分析TbZC3H8经RNAi敲低后对锥虫生长的影响,并通过RT-QPCR和Western blot检测RNAi干扰效率;构建异位融合表达myc-TAP标签的TbZC3H8细胞株,采用串联亲和纯化合并质谱鉴定其蛋白复合物组成;采用免疫荧光分析蛋白定位.结果 TbZC3H8的CCCH结构域在动基体原虫中高度保守;RNAi下调TbZC3H8后明显抑制了锥虫复制;TbZC3H8蛋白复合物中包含多种未知蛋白

  3. Tandem CCCH zinc finger proteins in plant growth, development and stress response.

    Science.gov (United States)

    Bogamuwa, Srimathi P; Jang, Jyan-Chyun

    2014-08-01

    Cysteine3Histidine (CCCH)-type zinc finger proteins comprise a large family that is well conserved across eukaryotes. Among them, tandem CCCH zinc finger proteins (TZFs) play critical roles in mRNA metabolism in animals and yeast. While there are only three TZF members in humans, a much higher number of TZFs has been found in many plant species. Notably, plant TZFs are over-represented by a class of proteins containing a unique TZF domain preceded by an arginine (R)-rich (RR) motif, hereafter called RR-TZF. Recently, there have been a large number of reports indicating that RR-TZF proteins can localize to processing bodies (P-bodies) and stress granules (SG), two novel cytoplasmic aggregations of messenger ribonucleoprotein complexes (mRNPs), and play critical roles in plant growth, development and stress response, probably via RNA regulation. This review focuses on the classification and most recent development of molecular, cellular and genetic analyses of plant RR-TZF proteins.

  4. Zinc finger structure-function in Ikaros

    Institute of Scientific and Technical Information of China (English)

    Marvin; A; Payne

    2011-01-01

    The zinc finger motif was used as a vehicle for the initial discovery of Ikaros in the context of T-cell differentiation and has been central to all subsequent analyses of Ikaros function.The Ikaros gene is alternately spliced to produce several isoforms that confer diversity of function and consequently have complicated analysis of the function of Ikaros in vivo.Key features of Ikaros in vivo function are associated with six C2H2 zinc fingers;four of which are alternately incorporated in the production of the various Ikaros isoforms.Although no complete structures are available for the Ikaros protein or any of its family members,considerable evidence has accumulated about the structure of zinc fingers and the role that this structure plays in the functions of the Ikaros family of proteins.This review summarizes the structural aspects of Ikaros zinc fingers,individually,and in tandem to provide a structural context for Ikaros function and to provide a structural basis to inform the design of future experiments with Ikaros and its family members.

  5. Prediction of DNA-binding specificity in zinc finger proteins

    Indian Academy of Sciences (India)

    Sumedha Roy; Shayoni Dutta; Kanika Khanna; Shruti Singla; Durai Sundar

    2012-07-01

    Zinc finger proteins interact via their individual fingers to three base pair subsites on the target DNA. The four key residue positions −1, 2, 3 and 6 on the alpha-helix of the zinc fingers have hydrogen bond interactions with the DNA. Mutating these key residues enables generation of a plethora of combinatorial possibilities that can bind to any DNA stretch of interest. Exploiting the binding specificity and affinity of the interaction between the zinc fingers and the respective DNA can help to generate engineered zinc fingers for therapeutic purposes involving genome targeting. Exploring the structure–function relationships of the existing zinc finger–DNA complexes can aid in predicting the probable zinc fingers that could bind to any target DNA. Computational tools ease the prediction of such engineered zinc fingers by effectively utilizing information from the available experimental data. A study of literature reveals many approaches for predicting DNA-binding specificity in zinc finger proteins. However, an alternative approach that looks into the physico-chemical properties of these complexes would do away with the difficulties of designing unbiased zinc fingers with the desired affinity and specificity. We present a physico-chemical approach that exploits the relative strengths of hydrogen bonding between the target DNA and all combinatorially possible zinc fingers to select the most optimum zinc finger protein candidate.

  6. Assessment of zinc finger orientations by residual dipolar coupling constants

    Energy Technology Data Exchange (ETDEWEB)

    Tsui, Vickie; Zhu Leiming; Huang, T.-H.; Wright, Peter E.; Case, David A. [Scripps Research Institute, Department of Molecular Biology (United States)

    2000-01-15

    Residual dipolar coupling constants measured in anisotropic solution contain information on orientations between internuclear vectors and the magnetic field, providing long-range information that may help determine the relative orientations of distinct domains in biomolecules. Here we describe the measurement and use of residual dipolar coupling restraints in the refinement of the structure of the complex of DNA with three zinc fingers of transcription factor IIIA (TFIIIA), measured in a DMPC/DHPC bicelle solution. These dipolar restraints were applied on a variety of orientations of the zinc finger domains (derived from crystallography, previous NMR studies, and systematic modeling) in order to examine the validity and sensitivity of using residual dipolar splittings to study interdomain orientations. The spread in interdomain angles between zinc fingers is reduced from 24 deg. to 9 deg. upon incorporation of dipolar restraints. However, the results also show that the ability to determine relative orientations is strongly dependent on the structural accuracy of the local domain structures.

  7. Zinc finger recombinases with adaptable DNA sequence specificity.

    Directory of Open Access Journals (Sweden)

    Chris Proudfoot

    Full Text Available Site-specific recombinases have become essential tools in genetics and molecular biology for the precise excision or integration of DNA sequences. However, their utility is currently limited to circumstances where the sites recognized by the recombinase enzyme have been introduced into the DNA being manipulated, or natural 'pseudosites' are already present. Many new applications would become feasible if recombinase activity could be targeted to chosen sequences in natural genomic DNA. Here we demonstrate efficient site-specific recombination at several sequences taken from a 1.9 kilobasepair locus of biotechnological interest (in the bovine β-casein gene, mediated by zinc finger recombinases (ZFRs, chimaeric enzymes with linked zinc finger (DNA recognition and recombinase (catalytic domains. In the "Z-sites" tested here, 22 bp casein gene sequences are flanked by 9 bp motifs recognized by zinc finger domains. Asymmetric Z-sites were recombined by the concomitant action of two ZFRs with different zinc finger DNA-binding specificities, and could be recombined with a heterologous site in the presence of a third recombinase. Our results show that engineered ZFRs may be designed to promote site-specific recombination at many natural DNA sequences.

  8. Enhanced cleavage of double-stranded DNA by artificial zinc-finger nuclease sandwiched between two zinc-finger proteins.

    Science.gov (United States)

    Mineta, Yusuke; Okamoto, Tomoyuki; Takenaka, Kosuke; Doi, Norio; Aoyama, Yasuhiro; Sera, Takashi

    2008-11-25

    To enhance DNA cleavage by zinc-finger nucleases (ZFNs), we sandwiched a DNA cleavage enzyme with two artificial zinc-finger proteins (AZPs). Because the DNA between the two AZP-binding sites is cleaved, the AZP-sandwiched nuclease is expected to bind preferentially to a DNA substrate rather than to cleavage products and thereby cleave it with multiple turnovers. To demonstrate the concept, we sandwiched a staphylococcal nuclease (SNase), which cleaves DNA as a monomer, between two three-finger AZPs. The AZP-sandwiched SNase cleaved large amounts of dsDNA site-specifically. Such multiple-turnover cleavage was not observed with nucleases that possess a single AZP. Thus, AZP-sandwiched nucleases will further refine ZFN technology.

  9. Toward a Code for the Interactions of Zinc Fingers with DNA: Selection of Randomized Fingers Displayed on Phage

    Science.gov (United States)

    Choo, Yen; Klug, Aaron

    1994-11-01

    We have used two selection techniques to study sequence-specific DNA recognition by the zinc finger, a small, modular DNA-binding minidomain. We have chosen zinc fingers because they bind as independent modules and so can be linked together in a peptide designed to bind a predetermined DNA site. In this paper, we describe how a library of zinc fingers displayed on the surface of bacteriophage enables selection of fingers capable of binding to given DNA triplets. The amino acid sequences of selected fingers which bind the same triplet are compared to examine how sequence-specific DNA recognition occurs. Our results can be rationalized in terms of coded interactions between zinc fingers and DNA, involving base contacts from a few α-helical positions. In the paper following this one, we describe a complementary technique which confirms the identity of amino acids capable of DNA sequence discrimination from these positions.

  10. Emerging roles of zinc finger proteins in regulating adipogenesis.

    Science.gov (United States)

    Wei, Shengjuan; Zhang, Lifan; Zhou, Xiang; Du, Min; Jiang, Zhihua; Hausman, Gary J; Bergen, Werner G; Zan, Linsen; Dodson, Michael V

    2013-12-01

    Proteins containing the zinc finger domain(s) are named zinc finger proteins (ZFPs), one of the largest classes of transcription factors in eukaryotic genomes. A large number of ZFPs have been studied and many of them were found to be involved in regulating normal growth and development of cells and tissues through diverse signal transduction pathways. Recent studies revealed that a small but increasing number of ZFPs could function as key transcriptional regulators involved in adipogenesis. Due to the prevalence of obesity and metabolic disorders, the investigation of molecular regulatory mechanisms of adipocyte development must be more completely understood in order to develop novel and long-term impact strategies for ameliorating obesity. In this review, we discuss recent work that has documented that ZFPs are important functional contributors to the regulation of adipogenesis. Taken together, these data lead to the conclusion that ZFPs may become promising targets to combat human obesity.

  11. Generation and functional analysis of zinc finger nucleases.

    Science.gov (United States)

    Cathomen, Toni; Segal, David J; Brondani, Vincent; Müller-Lerch, Felix

    2008-01-01

    The recent development of artificial endonucleases with tailored specificities has opened the door for a wide range of new applications, including the correction of mutated genes directly in the chromosome. This kind of gene therapy is based on homologous recombination, which can be stimulated by the creation of a targeted DNA double-strand break (DSB) near the site of the desired recombination event. Artificial nucleases containing zinc finger DNA-binding domains have provided important proofs of concept, showing that inserting a DSB in the target locus leads to gene correction frequencies of 1-18% in human cells. In this paper, we describe how zinc finger nucleases are assembled by polymerase chain reaction (PCR) and present two methods to assess these custom nucleases quickly in vitro and in a cell-based recombination assay.

  12. Editing the Plasmodium vivax Genome, Using Zinc-Finger Nucleases

    OpenAIRE

    Moraes Barros, Roberto R.; Straimer, Judith; Sa, Juliana M; Salzman, Rebecca E.; Melendez-Muniz, Viviana A.; Mu, Jianbing; David A Fidock; Thomas E. Wellems

    2014-01-01

    Plasmodium vivax is a major cause of malaria morbidity worldwide yet has remained genetically intractable. To stably modify this organism, we used zinc-finger nucleases (ZFNs), which take advantage of homology-directed DNA repair mechanisms at the site of nuclease action. Using ZFNs specific to the gene encoding P. vivax dihydrofolate reductase (pvdhfr), we transfected blood specimens from Saimiri boliviensis monkeys infected with the pyrimethamine (Pyr)–susceptible Chesson strain with a ZFN ...

  13. Phylogenetic Analysis of the Plant-specific Zinc Finger-Homeobox and Mini Zinc Finger Gene Families

    Institute of Scientific and Technical Information of China (English)

    Wei Hu; Claude W.dePamphilis; Hong Ma

    2008-01-01

    Zinc finger-homaodomain proteins (ZHD) are present in many plants;however,the evolutionary history of the ZHD gene family remains largely unknown.We show here that ZHD genes are plant-specific,nearly all intronless,and related to MINI ZINC FINGER (MIF) genes that possess only the zinc finger.Phylogenetic analyses of ZHD genes from representative land plants suggest that non-seed plant ZHD genes occupy basal positions and angiosperm homologs form seven distinct clades.Several clades contain genes from two or more major angiosperm groups,including eudicots,monocots,magnoliids,and other basal angiosperms,indicating that several duplications occurred before the diversification of flowering plants.In addition,specific lineages have experienced more recent duplications.Unlike the ZHD genes,&fiFs are found only from seed plants,possibly derived from ZHDs by loss of the homeodomain before the divergence of seed plants.Moreover,the MIF genes have also undergone relatively recent gene duplications.Finally,genome duplication might have contributed substantially to the expansion of family size in angiosperms and caused a high level of functional redundancy/overlap in these genes.

  14. Raman spectroscopy study of zinc finger ZNF191(243-368)

    Institute of Scientific and Technical Information of China (English)

    HUANG Zhongxian; ZHANG Hailiang; YAO Wenhua; LIU Yuqi; ZHOU Jing; TANG Liming

    2003-01-01

    The structure of ZNF191(243-368), the zinc finger region protein of zinc finger protein ZNF191, and its structural change upon thermal and EDTA-induced denaturation were investigated by the Raman spectroscopy. It was demonstrated that the coordination between Zn2+ and His/Cys in ZNF191(243-368) is the essential factor to the stability of zinc finger, which plays an important role in maintaining the hydrophobic core and the secondary structure in zinc finger, and the Raman spectroscopy is a powerful tool for investigating the structure of ZNF191(243-368).

  15. ZifBASE: a database of zinc finger proteins and associated resources

    Directory of Open Access Journals (Sweden)

    Punetha Ankita

    2009-09-01

    Full Text Available Abstract Background Information on the occurrence of zinc finger protein motifs in genomes is crucial to the developing field of molecular genome engineering. The knowledge of their target DNA-binding sequences is vital to develop chimeric proteins for targeted genome engineering and site-specific gene correction. There is a need to develop a computational resource of zinc finger proteins (ZFP to identify the potential binding sites and its location, which reduce the time of in vivo task, and overcome the difficulties in selecting the specific type of zinc finger protein and the target site in the DNA sequence. Description ZifBASE provides an extensive collection of various natural and engineered ZFP. It uses standard names and a genetic and structural classification scheme to present data retrieved from UniProtKB, GenBank, Protein Data Bank, ModBase, Protein Model Portal and the literature. It also incorporates specialized features of ZFP including finger sequences and positions, number of fingers, physiochemical properties, classes, framework, PubMed citations with links to experimental structures (PDB, if available and modeled structures of natural zinc finger proteins. ZifBASE provides information on zinc finger proteins (both natural and engineered ones, the number of finger units in each of the zinc finger proteins (with multiple fingers, the synergy between the adjacent fingers and their positions. Additionally, it gives the individual finger sequence and their target DNA site to which it binds for better and clear understanding on the interactions of adjacent fingers. The current version of ZifBASE contains 139 entries of which 89 are engineered ZFPs, containing 3-7F totaling to 296 fingers. There are 50 natural zinc finger protein entries ranging from 2-13F, totaling to 307 fingers. It has sequences and structures from literature, Protein Data Bank, ModBase and Protein Model Portal. The interface is cross linked to other public

  16. Molecular Cloning and Expression Analysis of a Cys2/His2 Type Zinc Finger Protein Gene in Upland Cotton

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The zinc finger proteins belong to the largest family of transcription factors.But there is little research of Cys2/His2 type zinc finger proteins in cotton,and there is no submission of correlating ESTs

  17. Sandwiched zinc-finger nucleases demonstrating higher homologous recombination rates than conventional zinc-finger nucleases in mammalian cells.

    Science.gov (United States)

    Mori, Tomoaki; Mori, Koichi; Tobimatsu, Takamasa; Sera, Takashi

    2014-02-01

    We previously reported that our sandwiched zinc-finger nucleases (ZFNs), in which a DNA cleavage domain is inserted between two artificial zinc-finger proteins, cleave their target DNA much more efficiently than conventional ZFNs in vitro. In the present study, we compared DNA cleaving efficiencies of a sandwiched ZFN with those of its corresponding conventional ZFN in mammalian cells. Using a plasmid-based single-strand annealing reporter assay in HEK293 cells, we confirmed that the sandwiched ZFN induced homologous recombination more efficiently than the conventional ZFN; reporter activation by the sandwiched ZFN was more than eight times that of the conventional one. Western blot analysis showed that the sandwiched ZFN was expressed less frequently than the conventional ZFN, indicating that the greater DNA-cleaving activity of the sandwiched ZFN was not due to higher expression of the sandwiched ZFN. Furthermore, an MTT assay demonstrated that the sandwiched ZFN did not have any significant cytotoxicity under the DNA-cleavage conditions. Thus, because our sandwiched ZFN cleaved more efficiently than its corresponding conventional ZFN in HEK293 cells as well as in vitro, sandwiched ZFNs are expected to serve as an effective molecular tool for genome editing in living cells.

  18. Editing the Plasmodium vivax genome, using zinc-finger nucleases.

    Science.gov (United States)

    Moraes Barros, Roberto R; Straimer, Judith; Sa, Juliana M; Salzman, Rebecca E; Melendez-Muniz, Viviana A; Mu, Jianbing; Fidock, David A; Wellems, Thomas E

    2015-01-01

    Plasmodium vivax is a major cause of malaria morbidity worldwide yet has remained genetically intractable. To stably modify this organism, we used zinc-finger nucleases (ZFNs), which take advantage of homology-directed DNA repair mechanisms at the site of nuclease action. Using ZFNs specific to the gene encoding P. vivax dihydrofolate reductase (pvdhfr), we transfected blood specimens from Saimiri boliviensis monkeys infected with the pyrimethamine (Pyr)-susceptible Chesson strain with a ZFN plasmid carrying a Pyr-resistant mutant pvdhfr sequence. We obtained Pyr-resistant parasites in vivo that carried mutant pvdhfr and additional silent mutations designed to confirm editing. These results herald the era of stable P. vivax genetic modifications.

  19. Genome editing in plant cells by zinc finger nucleases.

    Science.gov (United States)

    Weinthal, Dan; Tovkach, Andriy; Zeevi, Vardit; Tzfira, Tzvi

    2010-06-01

    Gene targeting is a powerful tool for functional gene studies. However, only a handful of reports have been published describing the successful targeting of genome sequences in model and crop plants. Gene targeting can be stimulated by induction of double-strand breaks at specific genomic sites. The expression of zinc finger nucleases (ZFNs) can induce genomic double-strand breaks. Indeed, ZFNs have been used to drive the replacement of native DNA sequences with foreign DNA molecules, to mediate the integration of the targeted transgene into native genome sequences, to stimulate the repair of defective transgenes, and as site-specific mutagens in model and crop plant species. This review introduces the principles underlying the use of ZFNs for genome editing, with an emphasis on their recent use for plant research and biotechnology.

  20. Enhanced protein production by engineered zinc finger proteins.

    Science.gov (United States)

    Reik, Andreas; Zhou, Yuanyue; Collingwood, Trevor N; Warfe, Lyndon; Bartsevich, Victor; Kong, Yanhong; Henning, Karla A; Fallentine, Barrett K; Zhang, Lei; Zhong, Xiaohong; Jouvenot, Yann; Jamieson, Andrew C; Rebar, Edward J; Case, Casey C; Korman, Alan; Li, Xiao-Yong; Black, Amelia; King, David J; Gregory, Philip D

    2007-08-01

    Increasing the yield of therapeutic proteins from mammalian production cell lines reduces costs and decreases the time to market. To this end, we engineered a zinc finger protein transcription factor (ZFP TF) that binds a DNA sequence within the promoter driving transgene expression. This ZFP TF enabled >100% increase in protein yield from CHO cells in transient, stable, and fermentor production run settings. Expression vectors engineered to carry up to 10 ZFP binding sites further enhanced ZFP-mediated increases in protein production up to approximately 500%. The multimerized ZFP binding sites function independently of the promoter, and therefore across vector platforms. CHO cell lines stably expressing ZFP TFs demonstrated growth characteristics similar to parental cell lines. ZFP TF expression and gains in protein production were stable over >30 generations in the absence of antibiotic selection. Our results demonstrate that ZFP TFs can rapidly and stably increase protein production in mammalian cells.

  1. Can Co(II) or Cd(II) substitute for Zn(II) in zinc fingers?

    Indian Academy of Sciences (India)

    P Rabindra Reddy; M Radhika

    2001-02-01

    Zinc finger domains consist of sequences of amino acids containing cysteine and histidine residues tetrahedrally coordinated to a zinc ion. The role of zinc in a DNA binding finger was considered purely structural due to the absence of redox chemistry in zinc. However, whether other metals e.g. Co(II) or Cd(II) can substitute Zn(II) is not settled. For an answer the detailed interaction of Co(II) and Cd(II) with cysteine methylester and histidine methylester has been investigated as a model for the zinc core in zinc fingers. The study was extended to different temperatures to evaluate the thermodynamic parameters associated with these interactions. The results suggest that zinc has a unique role.

  2. 15-zinc finger protein Bloody Fingers is required for zebrafish morphogenetic movements during neurulation.

    Science.gov (United States)

    Sumanas, Saulius; Zhang, Bo; Dai, Rujuan; Lin, Shuo

    2005-07-01

    A novel zebrafish gene bloody fingers (blf) encoding a 478 amino acid protein containing fifteen C(2)H(2) type zinc fingers was identified by expression screening. As determined by in situ hybridization, blf RNA displays strong ubiquitous early zygotic expression, while during late gastrulation and early somitogenesis, blf expression becomes transiently restricted to the posterior dorsal and lateral mesoderm. During later somitogenesis, blf expression appears only in hematopoietic cells. It is completely eliminated in cloche, moonshine but not in vlad tepes (gata1) mutant embryos. Morpholino (MO) knockdown of the Blf protein results in the defects of morphogenetic movements. Blf-MO-injected embryos (morphants) display shortened and widened axial tissues due to defective convergent extension. Unlike other convergent extension mutants, blf morphants display a split neural tube, resulting in a phenotype similar to the human open neural tube defect spina bifida. In addition, dorsal ectodermal cells delaminate in blf morphants during late somitogenesis. We propose a model explaining the role of blf in convergent extension and neurulation. We conclude that blf plays an important role in regulating morphogenetic movements during gastrulation and neurulation while its role in hematopoiesis may be redundant.

  3. [Cloning and structure analysis of zinc finger protein gene in Populus euphratica Oliv].

    Science.gov (United States)

    Wang, Jun-Ying; Yin, Wei-Lun; Xia, Xin-Li

    2005-03-01

    Zinc finger proteins belong to a family of nuclear transcription factors which function is to regulate gene expression in both prokaryotic and eukaryotic cells. A pair of primers was designed after analyzing the conservation of salt-tolerant zinc protein Alfin-1 in such diverse plants as alfalfa and Arabidopsis. The zinc finger protein gene is isolated from total RNA with RT-PCR in aquaculture leaves of Populus euphratica . Its full cDNA length is 924bp. Analysis of its amino acid sequence showed it has a typical Cys(2)/His(2) zinc finger structure and a G-rich promoter binding site GTGGGG, starting from position 556. Since transcrptional factors which have the same function show conservation in structure and amino acid sequence of DNA binding region, the structure analysis in this paper indicates the cloned zinc finger protein gene may have functional correlation to Alfin-1.

  4. Zinc finger peptide based optic sensor for detection of zinc ions.

    Science.gov (United States)

    Verma, Neelam; Kaur, Gagandeep

    2016-12-15

    In the present work, polyacrylamide gel has been used as a matrix for the immobilization of zinc finger peptide and fluorescent dye acrydine orange on the micro well plate to fabricate the fluorescence based biosensor for the detection of zinc ions in milk samples. The fluorescent dye moves in the hydrophobic groove formed after folding of the peptide in the presence of zinc ions. Under optimized conditions, linear range was observed between 0.001µg/l to 10µg/l of Zinc ions, with a lowest detection limit of 0.001µg/l and response time of 5min. Presented biosensor has shown 20% decrease in fluorescent intensity values after 5 regenerations and stable for more than one month, stored at 4°C. Interference study with other metal ions like lead, cadmium and copper showed a negligible change in fluorescence intensity in comparison to zinc ions. Developed bio sensing system was found to be novel, quick, reliable, miniaturized, stable, reproducible and repeatable and specific for zinc ion, which has been applied to various milk samples.

  5. NMR structure of the single QALGGH zinc finger domain from the Arabidopsis thaliana SUPERMAN protein.

    Science.gov (United States)

    Isernia, Carla; Bucci, Enrico; Leone, Marilisa; Zaccaro, Laura; Di Lello, Paola; Digilio, Giuseppe; Esposito, Sabrina; Saviano, Michele; Di Blasio, Benedetto; Pedone, Carlo; Pedone, Paolo V; Fattorusso, Roberto

    2003-03-03

    Zinc finger domains of the classical type represent the most abundant DNA binding domains in eukaryotic transcription factors. Plant proteins contain from one to four zinc finger domains, which are characterized by high conservation of the sequence QALGGH, shown to be critical for DNA-binding activity. The Arabidopsis thaliana SUPERMAN protein, which contains a single QALGGH zinc finger, is necessary for proper spatial development of reproductive floral tissues and has been shown to specifically bind to DNA. Here, we report the synthesis and UV and NMR spectroscopic structural characterization of a 37 amino acid SUPERMAN region complexed to a Zn(2+) ion (Zn-SUP37) and present the first high-resolution structure of a classical zinc finger domain from a plant protein. The NMR structure of the SUPERMAN zinc finger domain consists of a very well-defined betabetaalpha motif, typical of all other Cys(2)-His(2) zinc fingers structurally characterized. As a consequence, the highly conserved QALGGH sequence is located at the N terminus of the alpha helix. This region of the domain of animal zinc finger proteins consists of hypervariable residues that are responsible for recognizing the DNA bases. Therefore, we propose a peculiar DNA recognition code for the QALGGH zinc finger domain that includes all or some of the amino acid residues at positions -1, 2, and 3 (numbered relative to the N terminus of the helix) and possibly others at the C-terminal end of the recognition helix. This study further confirms that the zinc finger domain, though very simple, is an extremely versatile DNA binding motif.

  6. Stability and Folding Behavior Analysis of Zinc-Finger Using Simple Models

    Directory of Open Access Journals (Sweden)

    Xu-Hong Tian

    2010-10-01

    Full Text Available Zinc-fingers play crucial roles in regulating gene expression and mediating protein-protein interactions. In this article, two different proteins (Sp1f2 and FSD-1 are investigated using the Gaussian network model and anisotropy elastic network model. By using these simple coarse-grained methods, we analyze the structural stabilization and establish the unfolding pathway of the two different proteins, in good agreement with related experimental and molecular dynamics simulation data. From the analysis, it is also found that the folding process of the zinc-finger motif is predominated by several factors. Both the zinc ion and C-terminal loop affect the folding pathway of the zinc-finger motif. Knowledge about the stability and folding behavior of zinc-fingers may help in understanding the folding mechanisms of the zinc-finger motif and in designing new zinc-fingers. Meanwhile, these simple coarse-grained analyses can be used as a general and quick method for mechanistic studies of metalloproteins.

  7. Evaluation of Novel Design Strategies for Developing Zinc Finger Nucleases Tools for Treating Human Diseases

    Directory of Open Access Journals (Sweden)

    Christian Bach

    2014-01-01

    Full Text Available Zinc finger nucleases (ZFNs are associated with cell death and apoptosis by binding at countless undesired locations. This cytotoxicity is associated with the binding ability of engineered zinc finger domains to bind dissimilar DNA sequences with high affinity. In general, binding preferences of transcription factors are associated with significant degenerated diversity and complexity which convolutes the design and engineering of precise DNA binding domains. Evolutionary success of natural zinc finger proteins, however, evinces that nature created specific evolutionary traits and strategies, such as modularity and rank-specific recognition to cope with binding complexity that are critical for creating clinical viable tools to precisely modify the human genome. Our findings indicate preservation of general modularity and significant alteration of the rank-specific binding preferences of the three-finger binding domain of transcription factor SP1 when exchanging amino acids in the 2nd finger.

  8. Spectroscopic characterization of copper(I) binding to apo and metal-reconstituted zinc finger peptides.

    Science.gov (United States)

    Doku, Reginald T; Park, Grace; Wheeler, Korin E; Splan, Kathryn E

    2013-08-01

    Cu(I) exhibits high affinity for thiolate ligands, suggesting that thiol-rich zinc or iron binding sites may be subject to disruption during copper stress conditions. Zinc fingers constitute a large class of metalloproteins that use a combination of cysteine and histidine residues that bind Zn(II) as a structural element. Despite the shared preference of both copper and zinc for thiolate and amine coordination, the susceptibility of zinc finger domains toward copper substitution is not well studied. We report spectroscopic studies that characterize the Cu(I) binding properties of the zinc finger consensus peptides CP-CCHH, CP-CCHC, and CP-CCCC and the C-terminal zinc finger domain of HIV-1 nucleocapsid protein p7 (NCp7_C). Cu(I) binds to both the apopeptides and the Co(II)-substituted peptides, and the stoichiometry of Cu(I) binding is dependent on the number of cysteine thiols at the metal binding site. Fluorescence studies of the Zn(II)-NCp7_C complex indicate that Cu(I) also effectively competes with Zn(II) at the metal binding site, despite the high affinity of Zn(II) for the CCHC binding motif. Circular dichroism studies on both CP-CCHC and NCp7_C show that the conformations of the Cu(I)-bound complexes differ substantially from those of the Zn(II) species, implying that Cu(I) substitution is likely to impact zinc finger function. These results show that for the peptides studied here, Cu(I) is the thermodynamically favored metal despite the known high Zn(II) affinity of zinc finger domains, suggesting that Cu(I)-substituted zinc finger domains might be relevant in the context of both copper toxicity mechanisms and copper-responsive transcription factors.

  9. A multiscale approach to simulating the conformational properties of unbound multi-C₂H₂ zinc finger proteins.

    Science.gov (United States)

    Liu, Lei; Wade, Rebecca C; Heermann, Dieter W

    2015-09-01

    The conformational properties of unbound multi-Cys2 His2 (mC2H2) zinc finger proteins, in which zinc finger domains are connected by flexible linkers, are studied by a multiscale approach. Three methods on different length scales are utilized. First, atomic detail molecular dynamics simulations of one zinc finger and its adjacent flexible linker confirmed that the zinc finger is more rigid than the flexible linker. Second, the end-to-end distance distributions of mC2H2 zinc finger proteins are computed using an efficient atomistic pivoting algorithm, which only takes excluded volume interactions into consideration. The end-to-end distance distribution gradually changes its profile, from left-tailed to right-tailed, as the number of zinc fingers increases. This is explained by using a worm-like chain model. For proteins of a few zinc fingers, an effective bending constraint favors an extended conformation. Only for proteins containing more than nine zinc fingers, is a somewhat compacted conformation preferred. Third, a mesoscale model is modified to study both the local and the global conformational properties of multi-C2H2 zinc finger proteins. Simulations of the CCCTC-binding factor (CTCF), an important mC2H2 zinc finger protein for genome spatial organization, are presented.

  10. Molecular Cloning and Expression Analysis of a Cys2/His2 Type Zinc Finger Protein Gene in Upland Cotton

    Institute of Scientific and Technical Information of China (English)

    YANG Yu-wen; NI Wan-chao; ZHANG Bao-long; SHEN Xin-lian

    2008-01-01

    @@ The zinc finger proteins belong to the largest family of transcription factors.But there is little research of Cys2/His2 type zinc finger proteins in cotton,and there is no submission of correlating ESTs to GenBank.In this study,a full length of one Cys2/His2 type zinc finger protein (GZFP) and its 5 flanking sequence were obtained by RT-PCR and tail PCR.

  11. Solution structure of an archaeal DNA binding protein with an eukaryotic zinc finger fold.

    Directory of Open Access Journals (Sweden)

    Florence Guillière

    Full Text Available While the basal transcription machinery in archaea is eukaryal-like, transcription factors in archaea and their viruses are usually related to bacterial transcription factors. Nevertheless, some of these organisms show predicted classical zinc fingers motifs of the C2H2 type, which are almost exclusively found in proteins of eukaryotes and most often associated with transcription regulators. In this work, we focused on the protein AFV1p06 from the hyperthermophilic archaeal virus AFV1. The sequence of the protein consists of the classical eukaryotic C2H2 motif with the fourth histidine coordinating zinc missing, as well as of N- and C-terminal extensions. We showed that the protein AFV1p06 binds zinc and solved its solution structure by NMR. AFV1p06 displays a zinc finger fold with a novel structure extension and disordered N- and C-termini. Structure calculations show that a glutamic acid residue that coordinates zinc replaces the fourth histidine of the C2H2 motif. Electromobility gel shift assays indicate that the protein binds to DNA with different affinities depending on the DNA sequence. AFV1p06 is the first experimentally characterised archaeal zinc finger protein with a DNA binding activity. The AFV1p06 protein family has homologues in diverse viruses of hyperthermophilic archaea. A phylogenetic analysis points out a common origin of archaeal and eukaryotic C2H2 zinc fingers.

  12. Conservation, diversification and expansion of C2H2 zinc finger proteins in the Arabidopsis thaliana genome

    Directory of Open Access Journals (Sweden)

    Böhm Siegfried

    2004-07-01

    Full Text Available Background The classical C2H2 zinc finger domain is involved in a wide range of functions and can bind to DNA, RNA and proteins. The comparison of zinc finger proteins in several eukaryotes has shown that there is a lot of lineage specific diversification and expansion. Although the number of characterized plant proteins that carry the classical C2H2 zinc finger motifs is growing, a systematic classification and analysis of a plant genome zinc finger gene set is lacking. Results We found through in silico analysis 176 zinc finger proteins in Arabidopsis thaliana that hence constitute the most abundant family of putative transcriptional regulators in this plant. Only a minority of 33 A. thaliana zinc finger proteins are conserved in other eukaryotes. In contrast, the majority of these proteins (81% are plant specific. They are derived from extensive duplication events and form expanded families. We assigned the proteins to different subgroups and families and focused specifically on the two largest and evolutionarily youngest families (A1 and C1 that are suggested to be primarily involved in transcriptional regulation. The newly defined family A1 (24 members comprises proteins with tandemly arranged zinc finger domains. Family C1 (64 members, earlier described as the EPF-family in Petunia, comprises proteins with one isolated or two to five dispersed fingers and a mostly invariant QALGGH motif in the zinc finger helices. Based on the amino acid pattern in these helices we could describe five different signature sequences prevalent in C1 zinc finger domains. We also found a number of non-finger domains that are conserved in these families. Conclusions Our analysis of the few evolutionarily conserved zinc finger proteins of A. thaliana suggests that most of them could be involved in ancient biological processes like RNA metabolism and chromatin-remodeling. In contrast, the majority of the unique A. thaliana zinc finger proteins are known or

  13. The artificial zinc finger coding gene 'Jazz' binds the utrophin promoter and activates transcription.

    Science.gov (United States)

    Corbi, N; Libri, V; Fanciulli, M; Tinsley, J M; Davies, K E; Passananti, C

    2000-06-01

    Up-regulation of utrophin gene expression is recognized as a plausible therapeutic approach in the treatment of Duchenne muscular dystrophy (DMD). We have designed and engineered new zinc finger-based transcription factors capable of binding and activating transcription from the promoter of the dystrophin-related gene, utrophin. Using the recognition 'code' that proposes specific rules between zinc finger primary structure and potential DNA binding sites, we engineered a new gene named 'Jazz' that encodes for a three-zinc finger peptide. Jazz belongs to the Cys2-His2 zinc finger type and was engineered to target the nine base pair DNA sequence: 5'-GCT-GCT-GCG-3', present in the promoter region of both the human and mouse utrophin gene. The entire zinc finger alpha-helix region, containing the amino acid positions that are crucial for DNA binding, was specifically chosen on the basis of the contacts more frequently represented in the available list of the 'code'. Here we demonstrate that Jazz protein binds specifically to the double-stranded DNA target, with a dissociation constant of about 32 nM. Band shift and super-shift experiments confirmed the high affinity and specificity of Jazz protein for its DNA target. Moreover, we show that chimeric proteins, named Gal4-Jazz and Sp1-Jazz, are able to drive the transcription of a test gene from the human utrophin promoter.

  14. Comparing zinc finger nucleases and transcription activator-like effector nucleases for gene targeting in Drosophila.

    Science.gov (United States)

    Beumer, Kelly J; Trautman, Jonathan K; Christian, Michelle; Dahlem, Timothy J; Lake, Cathleen M; Hawley, R Scott; Grunwald, David J; Voytas, Daniel F; Carroll, Dana

    2013-10-03

    Zinc-finger nucleases have proven to be successful as reagents for targeted genome manipulation in Drosophila melanogaster and many other organisms. Their utility has been limited, however, by the significant failure rate of new designs, reflecting the complexity of DNA recognition by zinc fingers. Transcription activator-like effector (TALE) DNA-binding domains depend on a simple, one-module-to-one-base-pair recognition code, and they have been very productively incorporated into nucleases (TALENs) for genome engineering. In this report we describe the design of TALENs for a number of different genes in Drosophila, and we explore several parameters of TALEN design. The rate of success with TALENs was substantially greater than for zinc-finger nucleases , and the frequency of mutagenesis was comparable. Knockout mutations were isolated in several genes in which such alleles were not previously available. TALENs are an effective tool for targeted genome manipulation in Drosophila.

  15. Identification and localization of a novel zinc finger gene in developing chick skin and feather buds.

    Science.gov (United States)

    Padanilam, B J; Solursh, M

    1996-03-07

    We have cloned and sequenced a cDNA encoding a novel zinc finger protein (Fzf-1) containing two tandem repeats of zinc finger motifs of the C2H2 type. The cDNA is 3.0 Kb long and has an open reading frame which codes for a protein of 789 amino acids. The expression pattern of the zinc finger gene was studied in chick embryonic skin and feathers by in situ hybridization. The expression of the gene is found to be temporally and spatially regulated. In stage 38 chick embryos, the transcripts are localized to the epidermis but in 10-day-old embryos, the signal is localized to the forming dermis. In 12-day-old chick, the transcripts are localized to the mesenchymal region of the elongated feather buds. Reverse transcription followed by Polymerase Chain Reaction (RT-PCR) did not detect the transcripts in any other tissues.

  16. The (unusual) aspartic acid in the metal coordination sphere of the prokaryotic zinc finger domain.

    Science.gov (United States)

    D'Abrosca, Gianluca; Russo, Luigi; Palmieri, Maddalena; Baglivo, Ilaria; Netti, Fortuna; de Paola, Ivan; Zaccaro, Laura; Farina, Biancamaria; Iacovino, Rosa; Pedone, Paolo Vincenzo; Isernia, Carla; Fattorusso, Roberto; Malgieri, Gaetano

    2016-08-01

    The possibility of choices of protein ligands and coordination geometries leads to diverse Zn(II) binding sites in zinc-proteins, allowing a range of important biological roles. The prokaryotic Cys2His2 zinc finger domain (originally found in the Ros protein from Agrobacterium tumefaciens) tetrahedrally coordinates zinc through two cysteine and two histidine residues and it does not adopt a correct fold in the absence of the metal ion. Ros is the first structurally characterized member of a family of bacterial proteins that presents several amino acid changes in the positions occupied in Ros by the zinc coordinating residues. In particular, the second position is very often occupied by an aspartic acid although the coordination of structural zinc by an aspartate in eukaryotic zinc fingers is very unusual. Here, by appropriately mutating the protein Ros, we characterize the aspartate role within the coordination sphere of this family of proteins demonstrating how the presence of this residue only slightly perturbs the functional structure of the prokaryotic zinc finger domain while it greatly influences its thermodynamic properties.

  17. Stress Responsive Zinc-finger Protein Gene of Populus euphratica in Tobacco Enhances Salt Tolerance

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The Populus euphratica stress responsive zinc-finger protein gene PSTZ, which encodes a protein including typical Cys2/His2 zinc finger structure, was isolated by reverse transcription-polymerase chain reaction from P. euphratica.Northern hybridization revealed that its expression was induced under drought and salt stress conditions. To examine its function, cDNA of the PSTZ gene, driven by the cauliflower mosaic virus 35S promoter, was cloned into a plant expression vector pBin438 and introduced into tobacco plants. Transgenic tobacco showed an enhanced salt tolerance, suggesting that PSTZ may play a role in plant responsiveness to salt stress.

  18. Expression and RNA-binding of human zinc-finger antiviral protein

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Mi Suk; Kim, Eun Jung [Department of Molecular Biology, College of Natural Sciences, Pusan National University, Jangjeon-dong, Geumjeong-gu, Busan 609-735 (Korea, Republic of); Jang, Se Bok, E-mail: sbjang@pusan.ac.kr [Department of Molecular Biology, College of Natural Sciences, Pusan National University, Jangjeon-dong, Geumjeong-gu, Busan 609-735 (Korea, Republic of)

    2010-06-04

    Zinc-finger antiviral protein (ZAP) is a recently isolated host antiviral factor that inhibits the replication of many viruses such as Moloney murine leukemia virus (MLV) and Sindbis virus (SIN) by preventing the accumulation of viral mRNA in the cytoplasm. ZAP comprises four CCCH zinc-finger motifs, the second and fourth of which are responsible for protein activity based on their integrity. Thus far, there have been no reports on whether or not ZAP expressed in Escherichia coli is soluble. Therefore, we expressed N-terminal ZAP (NZAP, 254 amino acids) in E. coli as a fusion protein with several different cleavage sites and protein tags. Cleaved ZAP in soluble form strongly bound to RNA through its four CCCH zinc-finger motifs. Here, we provide evidence indicating that ZAP directly interacted with viral RNA. Each conserved zinc-finger motif of ZAP coordinates a zinc ion using three cysteines and one histidine. These findings suggest that ZAP recruits the cellular RNA degradation machinery for the degradation of viral RNA.

  19. Differential binding of monomethylarsonous acid compared to arsenite and arsenic trioxide with zinc finger peptides and proteins.

    Science.gov (United States)

    Zhou, Xixi; Sun, Xi; Mobarak, Charlotte; Gandolfi, A Jay; Burchiel, Scott W; Hudson, Laurie G; Liu, Ke Jian

    2014-04-21

    Arsenic is an environmental toxin that enhances the carcinogenic effect of DNA-damaging agents, such as ultraviolet radiation and benzo[a]pyrene. Interaction with zinc finger proteins has been shown to be an important molecular mechanism for arsenic toxicity and cocarcinogenesis. Arsenicals such as arsenite, arsenic trioxide (ATO), and monomethylarsonous acid (MMA(III)) have been reported to interact with cysteine residues of zinc finger domains, but little is known about potential differences in their selectivity of interaction. Herein we analyzed the interaction of arsenite, MMA(III), and ATO with C2H2, C3H1, and C4 configurations of zinc fingers using UV-vis, cobalt, fluorescence, and mass spectrometry. We observed that arsenite and ATO both selectively bound to C3H1 and C4 zinc fingers, while MMA(III) interacted with all three configurations of zinc finger peptides. Structurally and functionally, arsenite and ATO caused conformational changes and zinc loss on C3H1 and C4 zinc finger peptide and protein, respectively, whereas MMA(III) changed conformation and displaced zinc on all three types of zinc fingers. The differential selectivity was also demonstrated in zinc finger proteins isolated from cells treated with these arsenicals. Our results show that trivalent inorganic arsenic compounds, arsenite and ATO, have the same selectivity and behavior when interacting with zinc finger proteins, while methylation removes the selectivity. These findings provide insights on the molecular mechanisms underlying the differential effects of inorganic versus methylated arsenicals, as well as the role of in vivo arsenic methylation in arsenic toxicity and carcinogenesis.

  20. High-frequency genome editing using ssDNA oligonucleotides with zinc-finger nucleases

    DEFF Research Database (Denmark)

    Chen, Fuqiang; Pruett-Miller, Shondra M; Huang, Yuping

    2011-01-01

    Zinc-finger nucleases (ZFNs) have enabled highly efficient gene targeting in multiple cell types and organisms. Here we describe methods for using simple ssDNA oligonucleotides in tandem with ZFNs to efficiently produce human cell lines with three distinct genetic outcomes: (i) targeted point...

  1. DUF581 is plant specific FCS-like zinc finger involved in protein-protein interaction.

    Directory of Open Access Journals (Sweden)

    Muhammed Jamsheer K

    Full Text Available Zinc fingers are a ubiquitous class of protein domain with considerable variation in structure and function. Zf-FCS is a highly diverged group of C2-C2 zinc finger which is present in animals, prokaryotes and viruses, but not in plants. In this study we identified that a plant specific domain of unknown function, DUF581 is a zf-FCS type zinc finger. Based on HMM-HMM comparison and signature motif similarity we named this domain as FCS-Like Zinc finger (FLZ domain. A genome wide survey identified that FLZ domain containing genes are bryophytic in origin and this gene family is expanded in spermatophytes. Expression analysis of selected FLZ gene family members of A. thaliana identified an overlapping expression pattern suggesting a possible redundancy in their function. Unlike the zf-FCS domain, the FLZ domain found to be highly conserved in sequence and structure. Using a combination of bioinformatic and protein-protein interaction tools, we identified that FLZ domain is involved in protein-protein interaction.

  2. DUF581 is plant specific FCS-like zinc finger involved in protein-protein interaction.

    Science.gov (United States)

    K, Muhammed Jamsheer; Laxmi, Ashverya

    2014-01-01

    Zinc fingers are a ubiquitous class of protein domain with considerable variation in structure and function. Zf-FCS is a highly diverged group of C2-C2 zinc finger which is present in animals, prokaryotes and viruses, but not in plants. In this study we identified that a plant specific domain of unknown function, DUF581 is a zf-FCS type zinc finger. Based on HMM-HMM comparison and signature motif similarity we named this domain as FCS-Like Zinc finger (FLZ) domain. A genome wide survey identified that FLZ domain containing genes are bryophytic in origin and this gene family is expanded in spermatophytes. Expression analysis of selected FLZ gene family members of A. thaliana identified an overlapping expression pattern suggesting a possible redundancy in their function. Unlike the zf-FCS domain, the FLZ domain found to be highly conserved in sequence and structure. Using a combination of bioinformatic and protein-protein interaction tools, we identified that FLZ domain is involved in protein-protein interaction.

  3. Multiple-turnover cleavage of double-stranded DNA by sandwiched zinc-finger nuclease.

    Science.gov (United States)

    Mineta, Yusuke; Okamoto, Tomoyuki; Takenaka, Kosuke; Doi, Norio; Aoyama, Yasuhiro; Sera, Takashi

    2009-01-01

    To refine zinc-finger nuclease (ZFN) technology, we constructed a sandwiched ZFN, in which a DNA cleavage enzyme was sandwiched with two artificial zinc-finger proteins (AZPs). Because the sandwiched ZFN is designed to cleave the DNA between the two AZP-binding sites, the sandwiched ZFN is expected to bind preferentially to a DNA substrate rather than to cleavage products and thereby cleave it with multiple turnovers. To prove the concept, we sandwiched a staphylococcal nuclease (SNase), which cleaves DNA as a monomer, between two 3-finger AZPs. The AZP-sandwiched SNase cleaved large amounts of dsDNA site-specifically. Such multiple-turnover cleavage was not observed with control nucleases that possess a single AZP.

  4. Rapid mutation of endogenous zebrafish genes using zinc finger nucleases made by Oligomerized Pool ENgineering (OPEN.

    Directory of Open Access Journals (Sweden)

    Jonathan E Foley

    Full Text Available BACKGROUND: Customized zinc finger nucleases (ZFNs form the basis of a broadly applicable tool for highly efficient genome modification. ZFNs are artificial restriction endonucleases consisting of a non-specific nuclease domain fused to a zinc finger array which can be engineered to recognize specific DNA sequences of interest. Recent proof-of-principle experiments have shown that targeted knockout mutations can be efficiently generated in endogenous zebrafish genes via non-homologous end-joining-mediated repair of ZFN-induced DNA double-stranded breaks. The Zinc Finger Consortium, a group of academic laboratories committed to the development of engineered zinc finger technology, recently described the first rapid, highly effective, and publicly available method for engineering zinc finger arrays. The Consortium has previously used this new method (known as OPEN for Oligomerized Pool ENgineering to generate high quality ZFN pairs that function in human and plant cells. METHODOLOGY/PRINCIPAL FINDINGS: Here we show that OPEN can also be used to generate ZFNs that function efficiently in zebrafish. Using OPEN, we successfully engineered ZFN pairs for five endogenous zebrafish genes: tfr2, dopamine transporter, telomerase, hif1aa, and gridlock. Each of these ZFN pairs induces targeted insertions and deletions with high efficiency at its endogenous gene target in somatic zebrafish cells. In addition, these mutations are transmitted through the germline with sufficiently high frequency such that only a small number of fish need to be screened to identify founders. Finally, in silico analysis demonstrates that one or more potential OPEN ZFN sites can be found within the first three coding exons of more than 25,000 different endogenous zebrafish gene transcripts. CONCLUSIONS AND SIGNIFICANCE: In summary, our study nearly triples the total number of endogenous zebrafish genes successfully modified using ZFNs (from three to eight and suggests that OPEN

  5. Expression of putative zinc-finger protein lcn61 gene in lymphocystis disease virus China (LCDV-cn) genome

    Institute of Scientific and Technical Information of China (English)

    YAN Xiuying; SUN Xiuqin

    2009-01-01

    An open reading frame (lcn61) of iymphocystis disease virus China (LCDV-cn), probably responsible for encoding putative zinc-finger proteins was amplified and inserted into pET24a (+) vector.Then it expressed in E. coli BL21 (DE3), and His-tag fusion protein of high yield was obtained. It was found that the fusion protein existed in E. coli mainly as inclusion bodies. The bioinformatics analysis indicates that LCN61 is C2H2 type zinc-finger protein containing four C2H2 zinc-finger motifs. This work provides a theory for functional research of lcn61 gene.

  6. Zinc fingers as protein recognition motifs: structural basis for the GATA-1/friend of GATA interaction.

    Science.gov (United States)

    Liew, Chu Kong; Simpson, Raina J Y; Kwan, Ann H Y; Crofts, Linda A; Loughlin, Fionna E; Matthews, Jacqueline M; Crossley, Merlin; Mackay, Joel P

    2005-01-18

    GATA-1 and friend of GATA (FOG) are zinc-finger transcription factors that physically interact to play essential roles in erythroid and megakaryocytic development. Several naturally occurring mutations in the GATA-1 gene that alter the FOG-binding domain have been reported. The mutations are associated with familial anemias and thrombocytopenias of differing severity. To elucidate the molecular basis for the GATA-1/FOG interaction, we have determined the three-dimensional structure of a complex comprising the interaction domains of these proteins. The structure reveals how zinc fingers can act as protein recognition motifs. Details of the architecture of the contact domains and their physical properties provide a molecular explanation for how the GATA-1 mutations contribute to distinct but related genetic diseases.

  7. Unidirectional cloning by cleaving heterogeneous sites with a single sandwiched zinc finger nuclease.

    Science.gov (United States)

    Shinomiya, Kazuki; Mori, Tomoaki; Aoyama, Yasuhiro; Sera, Takashi

    2011-11-04

    We previously developed a novel type of zinc finger nucleases (ZFNs), sandwiched ZFNs that can discriminate DNA substrates from cleavage products and thus cleave DNA much more efficiently than conventional ZFNs as well as perform with multiple turnovers like restriction endonucleases. In the present study, we used the sandwiched ZFN to unidirectionally clone exogenous genes into target vectors by cleaving heterogeneous sites that contained heterogeneous spacer DNAs between two zinc-finger protein binding sites with a single sandwiched ZFN. We demonstrated that the sandwiched ZFN cleaved a 40-fold excess of both insert and vector plasmids within 1h and confirmed by sequencing that the resulting recombinants harbored the inserted DNA fragment in the desired orientation. Because sandwiched ZFNs can recognize and cleave a variety of long (≥ 26-bp) target DNAs, they may not only expand the utility of ZFNs for construction of recombinant plasmids, but also serve as useful meganucleases for synthesis of artificial genomes.

  8. Glycoengineering of Human Cell Lines Using Zinc Finger Nuclease Gene Targeting

    DEFF Research Database (Denmark)

    Steentoft, Catharina; Bennett, Eric Paul; Clausen, Henrik

    2013-01-01

    Lectin affinity chromatography is a powerful technique for isolation of glycoproteins carrying a specific glycan structure of interest. However, the enormous diversity of glycans present on the cell surface, as well as on individual proteins, makes it difficult to isolate an entire glycoproteome...... with one or even a series of lectins. Here we present a technique to generate cell lines with homogenous truncated O-glycans using zinc finger nuclease gene targeting. Because of their simplified O-glycoproteome, the cells have been named SimpleCells. Glycoproteins from SimpleCells can be isolated...... in a single purification step by lectin chromatography performed on a long lectin column. This protocol describes Zinc finger nuclease gene targeting of human cells to simplify the glycoproteome, as well as lectin chromatography and isolation of glycopeptides from total cell lysates of SimpleCells....

  9. Design, construction, and analysis of specific zinc finger nucleases for microphthalmia - associate transcription factor

    Directory of Open Access Journals (Sweden)

    Wenwen Wang

    2012-08-01

    Full Text Available This work studied the design, construction, and cleavage analysis of zinc finger nucleases (ZFNs that could cut the specific sequences within microphthalmia - associate transcription factor (mitfa of zebra fish. The target site and ZFPs were selected and designed with zinc finger tools, while the ZFPs were synthesized using DNAWorks and two-step PCR. The ZFNs were constructed, expressed, purified, and analyzed in vitro. As expected, the designed ZFNs could create a double-stand break (DSB at the target site in vitro. The DNAWorks, two-step PCR, and an optimized process of protein expression were firstly induced in the construction of ZFNs successfully, which was an effective and simplified protocol. These results could be useful for further application of ZFNs - mediated gene targeting.

  10. Zinc fingers as protein recognition motifs: Structural basis for the GATA-1/Friend of GATA interaction

    OpenAIRE

    Liew, Chu Kong; Simpson, Raina J. Y.; Kwan, Ann H.Y.; Crofts, Linda A.; Loughlin, Fionna E.; Matthews, Jacqueline M; Crossley, Merlin; Mackay, Joel P.

    2005-01-01

    GATA-1 and friend of GATA (FOG) are zinc-finger transcription factors that physically interact to play essential roles in erythroid and megakaryocytic development. Several naturally occurring mutations in the GATA-1 gene that alter the FOG-binding domain have been reported. The mutations are associated with familial anemias and thrombocytopenias of differing severity. To elucidate the molecular basis for the GATA-1/FOG interaction, we have determined the three-dimensional structure of a compl...

  11. Site-Specific Editing of the Plasmodium falciparum Genome Using Engineered Zinc-Finger Nucleases

    OpenAIRE

    Straimer, Judith; Lee, Marcus CS; Lee, Andrew H.; Zeitler, Bryan; Williams, April E.; Pearl, Jocelynn R.; Zhang, Lei; Rebar, Edward J.; Gregory, Philip D.; Llinás, Manuel; Urnov, Fyodor D; David A Fidock

    2012-01-01

    Malaria afflicts over 200 million people worldwide and its most lethal etiologic agent, Plasmodium falciparum, is evolving to resist even the latest-generation therapeutics. Efficient tools for genome-directed investigations of P. falciparum pathogenesis, including drug resistance mechanisms, are clearly required. Here we report rapid and targeted genetic engineering of this parasite, using zinc-finger nucleases (ZFNs) that produce a double-strand break in a user-defined locus and trigger hom...

  12. Zinc Finger Transcription Factors as Novel Genetic Switches to Modulate Metastatic Progression of Breast Tumors

    Science.gov (United States)

    2008-05-01

    Program, Biochemistry, Université de Montréal with Robert Cedergren 1999-2003 - Research Associate in Dr. Carlos Barbas, III laboratory...PCT/US03/03705 (2001). B. Peer Reviewed Publications (* most significant to proposed work) Blancafort, P., Ferbeyre, G., Sariol, C., and Cedergren , R...and Cedergren , R. The recognition of a non-canonical base pair by a zinc finger protein. Chem Biol. 1999; 6:585-97. Segal, D.J., Beerli R.R, Blancafort

  13. Transcripts from a novel human KRAB zinc finger gene contain spliced Alu and endogenous retroviral segments

    Energy Technology Data Exchange (ETDEWEB)

    Baban, S.; Freeman, J.D.; Mager, D.L. [Univ. of British Columbia, Vancouver, British Columbia (Canada)

    1996-05-01

    During the course of an investigation into the potential effects of endogenous retroviruses on adjacent gene expression, we isolated two cDNA clones containing a small sequence segment belonging to the human endogenous retrovirus family, HERV-H. Characterization of the clones revealed that they represent transcripts from a novel KRAB zinc finger gene termed ZNF177. The two cDNA clones differ at their 5{prime} termini and in the presence of a 559-bp internal exon. The clone containing this internal exon has six imperfect zinc finger motifs followed by seven perfect copies of the C{sub 2}H{sub 2} type but has a frame shift between the KRAB domain and the downstream zinc finger region. The smaller clone lacks the six imperfect motifs and has an intact ORF. The 5{prime} putative untranslated regions of both cDNAs contain an 86-bp HERV-H env segment and a segment of an Alu repeat, both in the antisense orientation, that have been incorporated by splicing. RT-PCR experiments show evidence of alternative splicing but the majority of transcripts appear to contain the Alu and env segments. Genomic PCR and hybridization experiments suggest that a partial HERV-H element is integrated within the ZNF177 locus, which Southern analysis has shown to be a single-copy gene. Northern and RT-PCR analyses suggest that ZNF177 is transcribed at a low level in a variety of cell types. 41 refs., 8 figs.

  14. Molecular Cloning and Expression Analysis of a Zinc Finger Protein Gene in Apple

    Institute of Scientific and Technical Information of China (English)

    CAOQiu-Fen; MasatoWADA; MENGYu-Ping; SUNYi; CUIGui-Mei

    2004-01-01

    A cDNA library was created from stem apex tissue from Jonathan apples (Malus domestica Borkh.), harvested in June to August, during which the plant transitions from vegetative growth to reproductive growth. From this library, we isolated an expressed sequence tag (EST) sequence containing a zinc finger motif, using this sequence, a 779 bp cDNA fragment was obtained by using 5' RACE, and a final full-length cDNA encoding an apple zinc finger protein (named MdZF1; GenBank accession number AB116545) was obtained by further PCR. This zinc finger motif of MdZF1 has high homology with INOETERMINATE1 (ID1) gene from maize which seemed to be involved in the transition to flowering. Northern blot and RT-PCR analyses showed that the MdZF1 expressed in the root, stem, leaves, shoot apex and floral organs of the apple, with expression levels higher in root, stem, leaves and floral shoot apex than that in floral organs (sepals, petals, stamens and pistils). Genomic Southern analysis showed that there was a single copy gene in apple genome.

  15. The Arabidopsis SUPERMAN protein is able to specifically bind DNA through its single Cys2-His2 zinc finger motif.

    Science.gov (United States)

    Dathan, Nina; Zaccaro, Laura; Esposito, Sabrina; Isernia, Carla; Omichinski, James G; Riccio, Andrea; Pedone, Carlo; Di Blasio, Benedetto; Fattorusso, Roberto; Pedone, Paolo V

    2002-11-15

    The Arabidopsis SUPERMAN (SUP) gene has been shown to be important in maintaining the boundary between stamens and carpels, and is presumed to act by regulating cell proliferation. In this work, we show that the SUP protein, which contains a single Cys2-His2 zinc finger domain including the QALGGH sequence, highly conserved in the plant zinc finger proteins, binds DNA. Using a series of deletion mutants, it was determined that the minimal domain required for specific DNA binding (residues 15-78) includes the single zinc finger and two basic regions located on either side of this motif. Furthermore, amino acid substitutions in the zinc finger or in the basic regions, including a mutation that knocks out the function of the SUP protein in vivo (glycine 63 to aspartate), have been found to abolish the activity of the SUP DNA-binding domain. These results strongly suggest that the SUP protein functions in vivo by acting as a DNA-binding protein, likely involved in transcriptional regulation. The association of both an N-terminal and a C-terminal basic region with a single Cys2-His2 zinc finger represents a novel DNA-binding motif suggesting that the mechanism of DNA recognition adopted by the SUP protein is different from that described so far in other zinc finger proteins.

  16. Interaction of Sp1 zinc finger with transport factor in the nuclear localization of transcription factor Sp1

    Energy Technology Data Exchange (ETDEWEB)

    Ito, Tatsuo [Department of Medicinal Biotechnology, Institute for Medicinal Research, Graduate School of Pharmaceutical Sciences, The University of Tokushima, 1-78 Sho-machi, Tokushima 770-8505 (Japan); Kitamura, Haruka; Uwatoko, Chisana; Azumano, Makiko [Department of Molecular Biophysical Chemistry, Faculty of Pharmaceutical Sciences, Doshisha Women' s University, Kodo, Kyotanabe City, Kyoto 610-0395 (Japan); Itoh, Kohji, E-mail: kitoh@ph.tokushima-u.ac.jp [Department of Medicinal Biotechnology, Institute for Medicinal Research, Graduate School of Pharmaceutical Sciences, The University of Tokushima, 1-78 Sho-machi, Tokushima 770-8505 (Japan); Kuwahara, Jun, E-mail: jkuwahar@dwc.doshisha.ac.jp [Department of Molecular Biophysical Chemistry, Faculty of Pharmaceutical Sciences, Doshisha Women' s University, Kodo, Kyotanabe City, Kyoto 610-0395 (Japan)

    2010-12-10

    Research highlights: {yields} Sp1 zinc fingers themselves interact with importin {alpha}. {yields} Sp1 zinc finger domains play an essential role as a nuclear localization signal. {yields} Sp1 can be transported into the nucleus in an importin-dependent manner. -- Abstract: Transcription factor Sp1 is localized in the nucleus and regulates the expression of many cellular genes, but the nuclear transport mechanism of Sp1 is not well understood. In this study, we revealed that GST-fused Sp1 protein bound to endogenous importin {alpha} in HeLa cells via the Sp1 zinc finger domains, which comprise the DNA binding domain of Sp1. It was found that the Sp1 zinc finger domains directly interacted with a wide range of importin {alpha} including the armadillo (arm) repeat domain and the C-terminal acidic domain. Furthermore, it turned out that all three zinc fingers of Sp1 are essential for binding to importin {alpha}. Taken together, these results suggest that the Sp1 zinc finger domains play an essential role as a NLS and Sp1 can be transported into the nucleus in an importin-dependent manner even though it possesses no classical NLSs.

  17. Myocardial ischemic preconditioning upregulated protein 1(Mipu1):zinc finger protein 667 - a multifunctional KRAB/C{sub 2}H{sub 2} zinc finger protein

    Energy Technology Data Exchange (ETDEWEB)

    Han, D.; Zhang, C. [Institute of Cardiovascular Disease, Key Lab for Arteriosclerology of Hunan Province, Post-doctoral Mobile Stations for Basic Medicine, University of South China, Hengyang City, Hunan Province (China); Fan, W.J. [Institute of Cardiovascular Disease, Key Lab for Arteriosclerology of Hunan Province, Post-doctoral Mobile Stations for Basic Medicine, University of South China, Hengyang City, Hunan Province (China); The Second Affiliated Hospital, University of South China, Hengyang City, Hunan Province (China); Pan, W.J.; Feng, D.M.; Qu, S.L.; Jiang, Z.S. [Institute of Cardiovascular Disease, Key Lab for Arteriosclerology of Hunan Province, Post-doctoral Mobile Stations for Basic Medicine, University of South China, Hengyang City, Hunan Province (China)

    2014-10-31

    Myocardial ischemic preconditioning upregulated protein 1 (Mipu1) is a newly discovered upregulated gene produced in rats during the myocardial ischemic preconditioning process. Mipu1 cDNA contains a 1824-base pair open reading frame and encodes a 608 amino acid protein with an N-terminal Krüppel-associated box (KRAB) domain and classical zinc finger C{sub 2}H{sub 2} motifs in the C-terminus. Mipu1 protein is located in the cell nucleus. Recent studies found that Mipu1 has a protective effect on the ischemia-reperfusion injury of heart, brain, and other organs. As a nuclear factor, Mipu1 may perform its protective function through directly transcribing and repressing the expression of proapoptotic genes to repress cell apoptosis. In addition, Mipu1 also plays an important role in regulating the gene expression of downstream inflammatory mediators by inhibiting the activation of activator protein-1 and serum response element.

  18. The zinc finger protein ZNF658 regulates the transcription of genes involved in zinc homeostasis and affects ribosome biogenesis through the zinc transcriptional regulatory element.

    Science.gov (United States)

    Ogo, Ogo A; Tyson, John; Cockell, Simon J; Howard, Alison; Valentine, Ruth A; Ford, Dianne

    2015-03-01

    We previously identified the ZTRE (zinc transcriptional regulatory element) in genes involved in zinc homeostasis and showed that it mediates transcriptional repression in response to zinc. We now report that ZNF658 acts at the ZTRE. ZNF658 was identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry of a band excised after electrophoretic mobility shift assay using a ZTRE probe. The protein contains a KRAB domain and 21 zinc fingers. It has similarity with ZAP1 from Saccharomyces cerevisiae, which regulates the response to zinc restriction, including a conserved DNA binding region we show to be functional also in ZNF658. Small interfering RNA (siRNA) targeted to ZNF658 abrogated the zinc-induced, ZTRE-dependent reduction in SLC30A5 (ZnT5 gene), SLC30A10 (ZnT10 gene), and CBWD transcripts in human Caco-2 cells and the ability of zinc to repress reporter gene expression from corresponding promoter-reporter constructs. Microarray analysis of the effect of reducing ZNF658 expression by siRNA uncovered a large decrease in rRNA. We find that ZTREs are clustered within the 45S rRNA precursor. We also saw effects on expression of multiple ribosomal proteins. ZNF658 thus links zinc homeostasis with ribosome biogenesis, the most active transcriptional, and hence zinc-demanding, process in the cell. ZNF658 is thus a novel transcriptional regulator that plays a fundamental role in the orchestrated cellular response to zinc availability.

  19. Characterization of two novel nuclear BTB/POZ domain zinc finger isoforms. Association with differentiation of hippocampal neurons, cerebellar granule cells, and macroglia

    DEFF Research Database (Denmark)

    Mitchelmore, Cathy; Kjaerulff, Karen M; Pedersen, Hans C

    2002-01-01

    BTB/POZ (broad complex tramtrack bric-a-brac/poxvirus and zinc finger) zinc finger factors are a class of nuclear DNA-binding proteins involved in development, chromatin remodeling, and cancer. However, BTB/POZ domain zinc finger factors linked to development of the mammalian cerebral cortex......, cerebellum, and macroglia have not been described previously. We report here the isolation and characterization of two novel nuclear BTB/POZ domain zinc finger isoforms, designated HOF(L) and HOF(S), that are specifically expressed in early hippocampal neurons, cerebellar granule cells, and gliogenic...

  20. Characterization of How DNA Modifications Affect DNA Binding by C2H2 Zinc Finger Proteins

    Science.gov (United States)

    Patel, A.; Hashimoto, H.; Zhang, X.; Cheng, X.

    2016-01-01

    Much is known about vertebrate DNA methylation and oxidation; however, much less is known about how modified cytosine residues within particular sequences are recognized. Among the known methylated DNA-binding domains, the Cys2-His2 zinc finger (ZnF) protein superfamily is the largest with hundreds of members, each containing tandem ZnFs ranging from 3 to >30 fingers. We have begun to biochemically and structurally characterize these ZnFs not only on their sequence specificity but also on their sensitivity to various DNA modifications. Rather than following published methods of refolding insoluble ZnF arrays, we have expressed and purified soluble forms of ZnFs, ranging in size from a tandem array of two to six ZnFs, from seven different proteins. We also describe a fluorescence polarization assay to measure ZnFs affinity with oligonucleotides containing various modifications and our approaches for cocrystallization of ZnFs with oligonucleotides. PMID:27372763

  1. Characterization of the SUMO-binding activity of the myeloproliferative and mental retardation (MYM-type zinc fingers in ZNF261 and ZNF198.

    Directory of Open Access Journals (Sweden)

    Catherine M Guzzo

    Full Text Available SUMO-binding proteins interact with SUMO modified proteins to mediate a wide range of functional consequences. Here, we report the identification of a new SUMO-binding protein, ZNF261. Four human proteins including ZNF261, ZNF198, ZNF262, and ZNF258 contain a stretch of tandem zinc fingers called myeloproliferative and mental retardation (MYM-type zinc fingers. We demonstrated that MYM-type zinc fingers from ZNF261 and ZNF198 are necessary and sufficient for SUMO-binding and that individual MYM-type zinc fingers function as SUMO-interacting motifs (SIMs. Our binding studies revealed that the MYM-type zinc fingers from ZNF261 and ZNF198 interact with the same surface on SUMO-2 recognized by the archetypal consensus SIM. We also present evidence that MYM-type zinc fingers in ZNF261 contain zinc, but that zinc is not required for SUMO-binding. Immunofluorescence microscopy studies using truncated fragments of ZNF198 revealed that MYM-type zinc fingers of ZNF198 are necessary for localization to PML-nuclear bodies (PML-NBs. In summary, our studies have identified and characterized the SUMO-binding activity of the MYM-type zinc fingers in ZNF261 and ZNF198.

  2. The Electronic Behavior of Zinc-Finger Protein Binding Sites in the Context of the DNA Extended Ladder Model

    Science.gov (United States)

    Oiwa, Nestor; Cordeiro, Claudette; Heermann, Dieter

    2016-05-01

    Instead of ATCG letter alignments, typically used in bioinformatics, we propose a new alignment method using the probability distribution function of the bottom of the occupied molecular orbital (BOMO), highest occupied molecular orbital (HOMO) and lowest unoccupied orbital (LUMO). We apply the technique to transcription factors with Cys2His2 zinc fingers. These transcription factors search for binding sites, probing for the electronic patterns at the minor and major DNA groves. The eukaryotic Cys2His2 zinc finger proteins bind to DNA ubiquitously at highly conserved domains. They are responsible for gene regulation and the spatial organization of DNA. To study and understand these zinc finger DNA-protein interactions, we use the extended ladder in the DNA model proposed by Zhu, Rasmussen, Balatsky & Bishop (2007) te{Zhu-2007}. Considering one single spinless electron in each nucleotide π-orbital along a double DNA chain (dDNA), we find a typical pattern for the bottom of BOMO, HOMO and LUMO along the binding sites. We specifically looked at two members of zinc finger protein family: specificity protein 1 (SP1) and early grown response 1 transcription factors (EGR1). When the valence band is filled, we find electrons in the purines along the nucleotide sequence, compatible with the electric charges of the binding amino acids in SP1 and EGR1 zinc finger.

  3. Zinc finger nuclease technology: A stable tool for high efficiency transformation in bloodstream form T. brucei.

    Science.gov (United States)

    Schumann, Gabriela; Kangussu-Marcolino, Monica M; Doiron, Nicholas; Käser, Sandro; de Assis Burle-Caldas, Gabriela; DaRocha, Wanderson D; Teixeira, Santuza M; Roditi, Isabel

    2017-02-20

    In Trypanosoma brucei, the generation of knockout mutants is relatively easy compared to other organisms as transfection methods are well established. These methods have their limitations, however, when it comes to the generation of genome-wide libraries that require a minimum of several hundred thousand transformants. Double-strand breaks with the meganuclease ISce-I dramatically increase transformation efficiency, but are not widely in use as cell lines need to be generated de novo before each transfection. Here we show that zinc finger nucleases are a robust and stable tool that can enhance transformation in bloodstream forms by more than an order of magnitude.

  4. ZFNGenome: A comprehensive resource for locating zinc finger nuclease target sites in model organisms

    Directory of Open Access Journals (Sweden)

    Voytas Daniel F

    2011-01-01

    Full Text Available Abstract Background Zinc Finger Nucleases (ZFNs have tremendous potential as tools to facilitate genomic modifications, such as precise gene knockouts or gene replacements by homologous recombination. ZFNs can be used to advance both basic research and clinical applications, including gene therapy. Recently, the ability to engineer ZFNs that target any desired genomic DNA sequence with high fidelity has improved significantly with the introduction of rapid, robust, and publicly available techniques for ZFN design such as the Oligomerized Pool ENgineering (OPEN method. The motivation for this study is to make resources for genome modifications using OPEN-generated ZFNs more accessible to researchers by creating a user-friendly interface that identifies and provides quality scores for all potential ZFN target sites in the complete genomes of several model organisms. Description ZFNGenome is a GBrowse-based tool for identifying and visualizing potential target sites for OPEN-generated ZFNs. ZFNGenome currently includes a total of more than 11.6 million potential ZFN target sites, mapped within the fully sequenced genomes of seven model organisms; S. cerevisiae, C. reinhardtii, A. thaliana, D. melanogaster, D. rerio, C. elegans, and H. sapiens and can be visualized within the flexible GBrowse environment. Additional model organisms will be included in future updates. ZFNGenome provides information about each potential ZFN target site, including its chromosomal location and position relative to transcription initiation site(s. Users can query ZFNGenome using several different criteria (e.g., gene ID, transcript ID, target site sequence. Tracks in ZFNGenome also provide "uniqueness" and ZiFOpT (Zinc Finger OPEN Targeter "confidence" scores that estimate the likelihood that a chosen ZFN target site will function in vivo. ZFNGenome is dynamically linked to ZiFDB, allowing users access to all available information about zinc finger reagents, such as the

  5. The N-terminal zinc finger of the erythroid transcription factor GATA-1 binds GATC motifs in DNA.

    Science.gov (United States)

    Newton, A; Mackay, J; Crossley, M

    2001-09-21

    The mammalian transcription factor GATA-1 is required for normal erythroid and megakaryocytic development. GATA-1 contains two zinc fingers, the C-terminal finger, which is known to bind (A/T)GATA(A/G) motifs in DNA and the N-finger, which is important for interacting with co-regulatory proteins such as Friend of GATA (FOG). We now show that, like the C-finger, the N-finger of GATA-1 is also capable of binding DNA but recognizes distinct sequences with the core GATC. We demonstrate that the GATA-1 N-finger can bind these sequences in vitro and that in cellular assays, GATA-1 can activate promoters containing GATC motifs. Experiments with mutant GATA-1 proteins confirm the importance of the N-finger, as the C-finger is not required for transactivation from GATC sites. Recently four naturally occurring mutations in GATA-1 have been shown to be associated with familial blood disorders. These mutations all map to the N-finger domain. We have investigated the effect of these mutations on the recognition of GATC sites by the N-finger and show that one mutation R216Q abolishes DNA binding, whereas the others have only minor effects.

  6. Evaluation of a modular strategy for the construction of novel polydactyl zinc finger DNA-binding proteins.

    Science.gov (United States)

    Segal, David J; Beerli, Roger R; Blancafort, Pilar; Dreier, Birgit; Effertz, Karin; Huber, Adrian; Koksch, Beate; Lund, Caren V; Magnenat, Laurent; Valente, David; Barbas, Carlos F

    2003-02-25

    In previous studies, we have developed a technology for the rapid construction of novel DNA-binding proteins with the potential to recognize any unique site in a given genome. This technology relies on the modular assembly of modified zinc finger DNA-binding domains, each of which recognizes a three bp subsite of DNA. A complete set of 64 domains would provide comprehensive recognition of any desired DNA sequence, and new proteins could be assembled by any laboratory in a matter of hours. However, a critical parameter for this approach is the extent to which each domain functions as an independent, modular unit, without influence or dependence on its neighboring domains. We therefore examined the detailed binding behavior of several modularly assembled polydactyl zinc finger proteins. We first demonstrated that 80 modularly assembled 3-finger proteins can recognize their DNA target with very high specificity using a multitarget ELISA-based specificity assay. A more detailed analysis of DNA binding specificity for eight 3-finger proteins and two 6-finger proteins was performed using a target site selection assay. Results showed that the specificity of these proteins was as good or better than that of zinc finger proteins constructed using methods that allow for interdependency. In some cases, near perfect specificity was achieved. Complications due to target site overlap were found to be restricted to only one particular amino acid interaction (involving an aspartate in position 2 of the alpha-helix) that occurs in a minority of cases. As this is the first report of target site selection for designed, well characterized 6-finger proteins, unique insights are discussed concerning the relationship of protein length and specificity. These results have important implications for the design of proteins that can recognize extended DNA sequences, as well as provide insights into the general rules of recognition for naturally occurring zinc finger proteins.

  7. Zinc Finger Nuclease induced DNA double stranded breaks and rearrangements in MLL

    Energy Technology Data Exchange (ETDEWEB)

    Do, To Uyen [Graduate Group in Immunology, University of California Davis, Davis, CA 95616 (United States); Department of Radiation Oncology, University of California Davis, Sacramento CA 95817 (United States); Ho, Bay; Shih, Shyh-Jen [Department of Radiation Oncology, University of California Davis, Sacramento CA 95817 (United States); Vaughan, Andrew, E-mail: Andrew.vaughan@ucdmc.ucdavis.edu [Graduate Group in Immunology, University of California Davis, Davis, CA 95616 (United States); Department of Radiation Oncology, University of California Davis, Sacramento CA 95817 (United States)

    2012-12-15

    Highlights: ► A Zinc Finger Nuclease (ZFN) targeting a leukemogenic hot spot for rearrangement in MLL is created. ► The novel ZFN efficiently cleaves MLL exon 13. ► Despite MLL cleavage and evidence of mis-repair, no leukemogenic translocations were produced. ► MLL cleavage alone is insufficient to generate leukemogenic translocations. - Abstract: Radiation treatment or chemotherapy has been linked with a higher risk of secondary cancers such as therapy related Acute Myeloid Leukemia (tAML). Several of these cancers have been shown to be correlated to the introduction of double stranded breaks (DSB) and rearrangements within the Mixed Lineage Leukemia (MLL) gene. We used Zinc Finger Nucleases (ZFNs) to introduce precise cuts within MLL to examine how a single DNA DSB might lead to chromosomal rearrangements. A ZFN targeting exon 13 within the Breakpoint Cluster Region of MLL was transiently expressed in a human lymphoblast cell line originating from a CML patient. Although FISH analysis showed ZFN DSB at this region increased the rate of MLL fragmentation, we were unable to detect leukemogenic rearrangements or translocations via inverse PCR. Interestingly, gene fragmentation as well as small interstitial deletions, insertions and base substitutions increased with the inhibition of DNA-PK, suggesting repair of this particular DSB is linked to non-homologous end joining (NHEJ). Although mis-repair of DSBs may be necessary for the initiation of leukemogenic translocations, a MLL targeted DNA break alone is insufficient.

  8. Zinc finger protein 278, a potential oncogene in human colorectal cancer

    Institute of Scientific and Technical Information of China (English)

    Xiaoqing Tian; Danfeng Sun; Yanjie Zhang; Shuliang Zhao; Hua Xiong; Jingyuan Fang

    2008-01-01

    Zinc finger protein 278 (ZNF278) is a novel Krueppel Cys2-His2-type zinc finger protein that is ubiquitously distributed in human tissues. Whether ZNF278 is related to the development of colorectal cancer is still unclear. The transcriptional level of ZNF278 was studied in colorectal cancer by real-time polymerase chain reaction. The results showed that ZNF278 expression was increased in 53% of colorectal cancer tissues compared to corresponding non-cancerous tissues. The transcriptional down-regulation of ZNF278 was detected in only three (6%) human colorectal cancer tissues compared to corresponding non-cancer tissues. No significant difference was detected in 19 (41%) pairs of samples.However, we failed to find a significant association between the up-regulation of ZNF278 transcription and age, sex, the degree of infiltration, or the tumor size of colorectal cancer.To study the function of ZNF278 in colorectal carcinogenesis,the colon cancer cell line SW1116 was stably transfected with a wild-type ZNF278 plasmid to construct an overexpression system, and was transiently transfected with the small interfering RNA of ZNF278 to construct a ZNF278 knockdown system. Cell proliferation was assessed with 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide dye and a cell counter. The results show that ZNF278 promotes cell growth, and its knockdown suppresses cell proliferation. ZNF278 could be a potential proto-oncogene in colorectal cancer.

  9. Simultaneous screening and validation of effective zinc finger nucleases in yeast.

    Directory of Open Access Journals (Sweden)

    Ling Wang

    Full Text Available Zinc finger nucleases (ZFNs have been successfully used for genome modification in various cell types and species. However, construction of an effective ZFN remained challenging. Previous studies all focused on obtaining specific zinc finger proteins (ZFPs first via bacterial 2-hybrid approach, and then fusing selected ZFPs to FokI nuclease domain. These assembled ZFNs have high rate of failing to cleave target sites in vivo. In this study, we developed a simultaneous screening and validation system to obtain effective ZFNs directly in yeast AH109. This system is based on Gal4 reporter system carrying a unique intermediate reporter plasmid with two 30-bp Gal4 homology arms and a ZFN target site. DNA double strand breaks introduced on target sequence by ZFNs were repaired by single strand annealing (SSA mechanism, and the restored Gal4 drove reporter genes expression. Taking the advantage of OPEN (Oligomerized Pool ENgineering selection, we constructed 3 randomized ZFNs libraries and 9 reporter strains for each target gene. We tested this system by taking goat α s1-casein as target gene following three-step selection. Consequently, 3 efficient pairs of ZFNs were obtained from positive colonies on selective medium. The ZFNs achieved a 15.9% disruption frequency in goat mammary epithelial cells. In conclusion, we created a novel system to obtain effective ZFNs directly with simultaneous screening and validation.

  10. Requiem: a novel zinc finger gene essential for apoptosis in myeloid cells.

    Science.gov (United States)

    Gabig, T G; Mantel, P L; Rosli, R; Crean, C D

    1994-11-25

    To identify genes mediating programmed cell death triggered by interleukin 3 (IL-3)-deprivation of myeloid cells, the IL-3-dependent murine myeloid cell line FDCP-1 was used to screen a mammalian cell expression library for cDNAs that would promote survival following withdrawal of IL-3. A unique 892-base pair cDNA was cloned that prevented the programmed cell death response following IL-3 deprivation by causing antisense suppression of an endogenous 2.4-kilobase (kb) mRNA. A 2.3-kb cDNA containing the identical 892-base pair over-lapping sequence was cloned that encoded a deduced 371-amino acid protein containing a single Kruppel-type zinc finger and a cluster of 4 cysteine/histidine-rich repeats resembling atypical zinc fingers. The 2.4-kb mRNA was found to be ubiquitously expressed in murine tissues and its abundance in FDCP-1 cells was not altered in response to IL-3 deprivation. Since expression of this 2.4-kb mRNA was a prerequisite for the apoptosis response following IL-3 deprivation, the gene encoding it was named requiem. Requiem is likely to encode a transcription factor required for the apoptosis response following survival factor withdrawal from myeloid cells.

  11. [Molecular cloning and expression analysis of a SUPERMAN-like zinc finger protein gene in upland cotton].

    Science.gov (United States)

    Yang, Yu-Wen; Ni, Wan-Chao; Zhang, Bao-Long; Shen, Xin-Lian; Zhang, Xiang-Gui; Xu, Ying-Jun; Yao, Shu

    2006-04-01

    The zinc finger proteins belong to the largest family of regulatory transcription factors, which play an important role in growth and development in animal and plant systems. SUPERMAN-like zinc finger protein gene has only one "finger like" motif. A pair of degenerate primers was designed according to the conserved regions, and 3 kinds of EST of this family were isolated from cotton through RT-PCR. The full length of one SUPERMAN-like zinc finger protein also has been acquired. The entire coding region is 744 bp and encodes a polypeptide of 248 amino acids with 40% homology to RBE protein of Arabidopsis deposited in the GenBank. This gene was designated as GZFP. It has the conserved zinc finger domain and the leucine rich region at the carboxyl terminus but no intron in the coding region. GZFP also has the plant nuclear localization signal. GZFP shows a more expression pattern in floral buds, ovaries, petals and roots than in phloem, xylem, fibers, leaves and seeds of cotton by RT-PCR, although it has a very low detection level and there is not any homologous ESTs found in the GenBank. Analysis of the 5' flanking sequence shows there are several regulatory elements responsible for pollen and root expression, four core sites required for binding of Dof proteins and four light-regulated elements.

  12. Structural and functional organization of the HF.10 human zinc finger gene (ZNF35) located on chromosome 3p21-p22

    DEFF Research Database (Denmark)

    Lanfrancone, L; Pengue, G; Pandolfi, P P;

    1992-01-01

    We report the structural and functional characterization of the HF.10 zinc finger gene (ZNF35) in normal human cells, as well as a processed pseudogene. The HF.10 gene spans about 13 kb and it is interrupted by three introns. All 11 zinc finger DNA-binding domains are contiguously encoded within...

  13. Non-transgenic genome modifications in a hemimetabolous insect using zinc-finger and TAL effector nucleases

    OpenAIRE

    Watanabe, T; Ochiai, H.; Sakuma, T.; Horch, HW; Hamaguchi, N.; Nakamura, T.; Bando, T.; Ohuchi, H.; Yamamoto, T.; Noji, S; Mito, T.

    2012-01-01

    Hemimetabolous, or incompletely metamorphosing, insects are phylogenetically relatively basal and comprise many pests. However, the absence of a sophisticated genetic model system, or targeted gene-manipulation system, has limited research on hemimetabolous species. Here we use zinc-finger nuclease and transcription activator-like effector nuclease technologies to produce genetic knockouts in the hemimetabolous insect Gryllus bimaculatus. Following the microinjection of mRNAs encoding zinc-fi...

  14. Sexual dimorphism in diverse metazoans is regulated by a novel class of intertwined zinc fingers.

    Science.gov (United States)

    Zhu, L; Wilken, J; Phillips, N B; Narendra, U; Chan, G; Stratton, S M; Kent, S B; Weiss, M A

    2000-07-15

    Sex determination is regulated by diverse pathways. Although upstream signals vary, a cysteine-rich DNA-binding domain (the DM motif) is conserved within downstream transcription factors of Drosophila melanogaster (Doublesex) and Caenorhabditis elegans (MAB-3). Vertebrate DM genes have likewise been identified and, remarkably, are associated with human sex reversal (46, XY gonadal dysgenesis). Here we demonstrate that the structure of the Doublesex domain contains a novel zinc module and disordered tail. The module consists of intertwined CCHC and HCCC Zn(2+)-binding sites; the tail functions as a nascent recognition alpha-helix. Mutations in either Zn(2+)-binding site or tail can lead to an intersex phenotype. The motif binds in the DNA minor groove without sharp DNA bending. These molecular features, unusual among zinc fingers and zinc modules, underlie the organization of a Drosophila enhancer that integrates sex- and tissue-specific signals. The structure provides a foundation for analysis of DM mutations affecting sexual dimorphism and courtship behavior.

  15. Classification of the treble clef zinc finger: noteworthy lessons for structure and function evolution

    Science.gov (United States)

    Kaur, Gurmeet; Subramanian, Srikrishna

    2016-01-01

    Treble clef (TC) zinc fingers constitute a large fold-group of structural zinc-binding protein domains that mediate numerous cellular functions. We have analysed the sequence, structure, and function relationships among all TCs in the Protein Data Bank. This led to the identification of novel TCs, such as lsr2, YggX and TFIIIC τ 60 kDa subunit, and prediction of a nuclease-like function for the DUF1364 family. The structural malleability of TCs is evident from the many examples with variations to the core structural elements of the fold. We observe domains wherein the structural core of the TC fold is circularly permuted, and also some examples where the overall fold resembles both the TC motif and another unrelated fold. All extant TC families do not share a monophyletic origin, as several TC proteins are known to have been present in the last universal common ancestor and the last eukaryotic common ancestor. We identify several TCs where the zinc-chelating site and residues are not merely responsible for structure stabilization but also perform other functions, such as being redox active in C1B domain of protein kinase C, a nucleophilic acceptor in Ada and catalytic in organomercurial lyase, MerB. PMID:27562564

  16. Predicting success of oligomerized pool engineering (OPEN for zinc finger target site sequences

    Directory of Open Access Journals (Sweden)

    Goodwin Mathew J

    2010-11-01

    Full Text Available Abstract Background Precise and efficient methods for gene targeting are critical for detailed functional analysis of genomes and regulatory networks and for potentially improving the efficacy and safety of gene therapies. Oligomerized Pool ENgineering (OPEN is a recently developed method for engineering C2H2 zinc finger proteins (ZFPs designed to bind specific DNA sequences with high affinity and specificity in vivo. Because generation of ZFPs using OPEN requires considerable effort, a computational method for identifying the sites in any given gene that are most likely to be successfully targeted by this method is desirable. Results Analysis of the base composition of experimentally validated ZFP target sites identified important constraints on the DNA sequence space that can be effectively targeted using OPEN. Using alternate encodings to represent ZFP target sites, we implemented Naïve Bayes and Support Vector Machine classifiers capable of distinguishing "active" targets, i.e., ZFP binding sites that can be targeted with a high rate of success, from those that are "inactive" or poor targets for ZFPs generated using current OPEN technologies. When evaluated using leave-one-out cross-validation on a dataset of 135 experimentally validated ZFP target sites, the best Naïve Bayes classifier, designated ZiFOpT, achieved overall accuracy of 87% and specificity+ of 90%, with an ROC AUC of 0.89. When challenged with a completely independent test set of 140 newly validated ZFP target sites, ZiFOpT performance was comparable in terms of overall accuracy (88% and specificity+ (92%, but with reduced ROC AUC (0.77. Users can rank potentially active ZFP target sites using a confidence score derived from the posterior probability returned by ZiFOpT. Conclusion ZiFOpT, a machine learning classifier trained to identify DNA sequences amenable for targeting by OPEN-generated zinc finger arrays, can guide users to target sites that are most likely to function

  17. Probing isoform-specific functions of polypeptide GalNAc-transferases using zinc finger nuclease glycoengineered SimpleCells

    DEFF Research Database (Denmark)

    Schjoldager, Katrine Ter-Borch Gram; Vakhrushev, Sergey Y; Kong, Yun;

    2012-01-01

    NAc-Ts are largely unavailable. We recently introduced SimpleCells, i.e., human cell lines made deficient in O-glycan extension by zinc finger nuclease targeting of a key gene in O-glycan elongation (Cosmc), which allows for proteome-wide discovery of O-glycoproteins. Here we have extended the SimpleCell concept...

  18. Activation of transcriptional activity of HSE by a novel mouse zinc finger protein ZNFD specifically expressed in testis.

    Science.gov (United States)

    Xu, Fengqin; Wang, Weiping; Lei, Chen; Liu, Qingmei; Qiu, Hao; Muraleedharan, Vinaydhar; Zhou, Bin; Cheng, Hongxia; Huang, Zhongkai; Xu, Weian; Li, Bichun; Wang, Minghua

    2012-04-01

    Zinc finger proteins (ZFPs) that contain multiple cysteine and/or histidine residues perform important roles in various cellular functions, including transcriptional regulation, cell proliferation, differentiation, and apoptosis. The Cys-Cys-His-His (C(2)H(2)) type of ZFPs are the well-defined members of this super family and are the largest and most complex proteins in eukaryotic genomes. In this study, we identified a novel C(2)H(2) type of zinc finger gene ZNFD from mice which has a 1,002 bp open reading frame and encodes a protein with 333 amino acid residues. The predicted 37.4 kDa protein contains a C(2)H(2) zinc finger domain. ZNFD gene is located on chromosome 18qD1. RT-PCR analysis revealed that the ZNFD gene was specifically expressed in mouse testis but not in other tissues. Subcellular localization analysis demonstrated that ZNFD was localized in the nucleus. Reporter gene assays showed that overexpression of ZNFD in the COS7 cells activates the transcriptional activities of heat shock element (HSE). Overall, these results suggest that ZNFD is a member of the zinc finger transcription factor family and it participates in the transcriptional regulation of HSE. Many heat shock proteins regulated by HSE are involved in testicular development. Therefore, our results suggest that ZNFD may probably participate in the development of mouse testis and function as a transcription activator in HSE-mediated gene expression and signaling pathways.

  19. Hippocampus-like corticoneurogenesis induced by two isoforms of the BTB-zinc finger gene Zbtb20 in mice

    DEFF Research Database (Denmark)

    Nielsen, Jakob V; Nielsen, Flemming H; Ismail, Rola

    2007-01-01

    Hippocampus-associated genes that orchestrate the formation of the compact stratum pyramidale are largely unknown. The BTB (broad complex, tramtrack, bric-a-brac)-zinc finger gene Zbtb20 (also known as HOF, Znf288, Zfp288) encodes two protein isoforms, designated Zbtb20(S) and Zbtb20(L), which...

  20. Arabidopsis VARIEGATED 3 encodes a chloroplast-targeted, zinc-finger protein required for chloroplast and palisade cell development

    DEFF Research Database (Denmark)

    Næsted, Henrik; Holm, Agnethe; Jenkins, Tom

    2004-01-01

    protein containing novel repeats and zinc fingers described as protein interaction domains. VAR3 interacts specifically in yeast and in vitro with NCED4, a putative polyene chain or carotenoid dioxygenase, and both VAR3 and NCED4 accumulate in the chloroplast stroma. Metabolic profiling demonstrates...

  1. Arabidopsis VARIEGATED 3 encodes a chloroplasttargeted, zinc-finger protein required for chloroplast and palisade cell development

    DEFF Research Database (Denmark)

    Næsted, Henrik; Holm, A.; Jenkins, T.

    2004-01-01

    protein containing novel repeats and zinc fingers described as protein interaction domains. VAR3 interacts specifically in yeast and in vitro with NCED4, a putative polyene chain or carotenoid dioxygenase, and both VAR3 and NCED4 accumulate in the chloroplast stroma. Metabolic profiling demonstrates...

  2. Site-specific genome editing in Plasmodium falciparum using engineered zinc-finger nucleases.

    Science.gov (United States)

    Straimer, Judith; Lee, Marcus C S; Lee, Andrew H; Zeitler, Bryan; Williams, April E; Pearl, Jocelynn R; Zhang, Lei; Rebar, Edward J; Gregory, Philip D; Llinás, Manuel; Urnov, Fyodor D; Fidock, David A

    2012-10-01

    Malaria afflicts over 200 million people worldwide, and its most lethal etiologic agent, Plasmodium falciparum, is evolving to resist even the latest-generation therapeutics. Efficient tools for genome-directed investigations of P. falciparum-induced pathogenesis, including drug-resistance mechanisms, are clearly required. Here we report rapid and targeted genetic engineering of this parasite using zinc-finger nucleases (ZFNs) that produce a double-strand break in a user-defined locus and trigger homology-directed repair. Targeting an integrated egfp locus, we obtained gene-deletion parasites with unprecedented speed (2 weeks), both with and without direct selection. ZFNs engineered against the parasite gene pfcrt, responsible for escape under chloroquine treatment, rapidly produced parasites that carried either an allelic replacement or a panel of specified point mutations. This method will enable a diverse array of genome-editing approaches to interrogate this human pathogen.

  3. Modular system for the construction of zinc-finger libraries and proteins.

    Science.gov (United States)

    Gonzalez, Beatriz; Schwimmer, Lauren J; Fuller, Roberta P; Ye, Yongjun; Asawapornmongkol, Lily; Barbas, Carlos F

    2010-04-01

    Engineered zinc-finger transcription factors (ZF-TF) are powerful tools to modulate the expression of specific genes. Complex libraries of ZF-TF can be delivered into cells to scan the genome for genes responsible for a particular phenotype or to select the most effective ZF-TF to regulate an individual gene. In both cases, the construction of highly representative and unbiased libraries is critical. In this protocol, we describe a user-friendly ZF technology suitable for the creation of complex libraries and the construction of customized ZF-TFs. The new technology described here simplifies the building of ZF libraries, avoids PCR-introduced bias and ensures equal representation of every module. We also describe the construction of a customized ZF-TF that can be transferred to a number of expression vectors. This protocol can be completed in 9-11 d.

  4. Importance of long-time simulations for rare event sampling in zinc finger proteins.

    Science.gov (United States)

    Godwin, Ryan; Gmeiner, William; Salsbury, Freddie R

    2016-01-01

    Molecular dynamics (MD) simulation methods have seen significant improvement since their inception in the late 1950s. Constraints of simulation size and duration that once impeded the field have lessened with the advent of better algorithms, faster processors, and parallel computing. With newer techniques and hardware available, MD simulations of more biologically relevant timescales can now sample a broader range of conformational and dynamical changes including rare events. One concern in the literature has been under which circumstances it is sufficient to perform many shorter timescale simulations and under which circumstances fewer longer simulations are necessary. Herein, our simulations of the zinc finger NEMO (2JVX) using multiple simulations of length 15, 30, 1000, and 3000 ns are analyzed to provide clarity on this point.

  5. Functional promoter variant in zinc finger protein 202 predicts severe atherosclerosis and ischemic heart disease

    DEFF Research Database (Denmark)

    Frikke-Schmidt, R.; Nordestgaard, Børge; Grande, Peer

    2008-01-01

    Objectives This study was designed to test the hypotheses that single nucleotide polymorphisms ( SNPs), in zinc finger protein 202 ( ZNF202), predict severe atherosclerosis and ischemic heart disease ( IHD). Background ZNF202 is a transcriptional repressor controlling promoter elements in genes......,998 controls. Finally, we determined whether g. -660A>G altered transcriptional activity of the ZNF202 promoter in vitro. Results Cross-sectionally, ZNF202 g. -660 GG versus AA homozygosity predicted an odds ratio for severe atherosclerosis of 2.01 ( 95% confidence interval [CI]: 1.34 to 3.01). Prospectively...... were highly correlated with g. -660A>G, also predicted risk of severe atherosclerosis and IHD. Finally, ZNF202 g. -660G versus g. -660A was associated with a 60% reduction in transcriptional activity in vitro, whereas none of the 2 correlated SNPs were predicted to be functional. Conclusions...

  6. Characterization of the tandem CWCH2 sequence motif: a hallmark of inter-zinc finger interactions

    Directory of Open Access Journals (Sweden)

    Aruga Jun

    2010-02-01

    Full Text Available Abstract Background The C2H2 zinc finger (ZF domain is widely conserved among eukaryotic proteins. In Zic/Gli/Zap1 C2H2 ZF proteins, the two N-terminal ZFs form a single structural unit by sharing a hydrophobic core. This structural unit defines a new motif comprised of two tryptophan side chains at the center of the hydrophobic core. Because each tryptophan residue is located between the two cysteine residues of the C2H2 motif, we have named this structure the tandem CWCH2 (tCWCH2 motif. Results Here, we characterized 587 tCWCH2-containing genes using data derived from public databases. We categorized genes into 11 classes including Zic/Gli/Glis, Arid2/Rsc9, PacC, Mizf, Aebp2, Zap1/ZafA, Fungl, Zfp106, Twincl, Clr1, and Fungl-4ZF, based on sequence similarity, domain organization, and functional similarities. tCWCH2 motifs are mostly found in organisms belonging to the Opisthokonta (metazoa, fungi, and choanoflagellates and Amoebozoa (amoeba, Dictyostelium discoideum. By comparison, the C2H2 ZF motif is distributed widely among the eukaryotes. The structure and organization of the tCWCH2 motif, its phylogenetic distribution, and molecular phylogenetic analysis suggest that prototypical tCWCH2 genes existed in the Opisthokonta ancestor. Within-group or between-group comparisons of the tCWCH2 amino acid sequence identified three additional sequence features (site-specific amino acid frequencies, longer linker sequence between two C2H2 ZFs, and frequent extra-sequences within C2H2 ZF motifs. Conclusion These features suggest that the tCWCH2 motif is a specialized motif involved in inter-zinc finger interactions.

  7. Employing libraries of zinc finger artificial transcription factors to screen for homologous recombination mutants in Arabidopsis.

    Science.gov (United States)

    Lindhout, Beatrice I; Pinas, Johan E; Hooykaas, Paul J J; van der Zaal, Bert J

    2006-11-01

    A library of genes for zinc finger artificial transcription factors (ZF-ATF) was generated by fusion of DNA sequences encoding three-finger Cys(2)His(2) ZF domains to the VP16 activation domain under the control of the promoter of the ribosomal protein gene RPS5A from Arabidopsis thaliana. After introduction of this library into an Arabidopsis homologous recombination (HR) indicator line, we selected primary transformants exhibiting multiple somatic recombination events. After PCR-mediated rescue of ZF sequences, reconstituted ZF-ATFs were re-introduced in the target line. In this manner, a ZF-ATF was identified that led to a 200-1000-fold increase in somatic HR (replicated in an independent second target line). A mutant plant line expressing the HR-inducing ZF-ATF exhibited increased resistance to the DNA-damaging agent bleomycin and was more sensitive to methyl methanesulfonate (MMS), a combination of traits not described previously. Our results demonstrate that the use of ZF-ATF pools is highly rewarding when screening for novel dominant phenotypes in Arabidopsis.

  8. Automatic polymerase chain reaction product detection system for food safety monitoring using zinc finger protein fused to luciferase

    Energy Technology Data Exchange (ETDEWEB)

    Yoshida, Wataru; Kezuka, Aki; Murakami, Yoshiyuki; Lee, Jinhee; Abe, Koichi [Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588 (Japan); Motoki, Hiroaki; Matsuo, Takafumi; Shimura, Nobuaki [System Instruments Co., Ltd., 776-2 Komiya-cho, Hachioji, Tokyo 192-0031 (Japan); Noda, Mamoru; Igimi, Shizunobu [Division of Biomedical Food Research, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501 (Japan); Ikebukuro, Kazunori, E-mail: ikebu@cc.tuat.ac.jp [Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588 (Japan)

    2013-11-01

    Graphical abstract: -- Highlights: •Zif268 fused to luciferase was used for E. coli O157, Salmonella and coliform detection. •Artificial zinc finger protein fused to luciferase was constructed for Norovirus detection. •An analyzer that automatically detects PCR products by zinc finger protein fused to luciferase was developed. •Target pathogens were specifically detected by the automatic analyzer with zinc finger protein fused to luciferase. -- Abstract: An automatic polymerase chain reaction (PCR) product detection system for food safety monitoring using zinc finger (ZF) protein fused to luciferase was developed. ZF protein fused to luciferase specifically binds to target double stranded DNA sequence and has luciferase enzymatic activity. Therefore, PCR products that comprise ZF protein recognition sequence can be detected by measuring the luciferase activity of the fusion protein. We previously reported that PCR products from Legionella pneumophila and Escherichia coli (E. coli) O157 genomic DNA were detected by Zif268, a natural ZF protein, fused to luciferase. In this study, Zif268–luciferase was applied to detect the presence of Salmonella and coliforms. Moreover, an artificial zinc finger protein (B2) fused to luciferase was constructed for a Norovirus detection system. In the luciferase activity detection assay, several bound/free separation process is required. Therefore, an analyzer that automatically performed the bound/free separation process was developed to detect PCR products using the ZF–luciferase fusion protein. By means of the automatic analyzer with ZF–luciferase fusion protein, target pathogenic genomes were specifically detected in the presence of other pathogenic genomes. Moreover, we succeeded in the detection of 10 copies of E. coli BL21 without extraction of genomic DNA by the automatic analyzer and E. coli was detected with a logarithmic dependency in the range of 1.0 × 10 to 1.0 × 10{sup 6} copies.

  9. The second zinc-finger domain of poly(ADP-ribose) polymerase determines specificity for single-stranded breaks in DNA.

    NARCIS (Netherlands)

    G. Gradwohl; J.M. de Murcia; M. Molinete; F. Simonin; M.H.M. Koken (Marcel); J.H.J. Hoeijmakers (Jan); G. de Murcia

    1990-01-01

    textabstractPoly(ADP-ribose) polymerase (EC 2.4.2.30) is a zinc-binding protein that specifically binds to a DNA strand break in a zinc-dependent manner. We describe here the cloning and expression in Escherichia coli of a cDNA fragment encoding the two putative zinc fingers (FI and FII) domain of t

  10. Zinc to cadmium replacement in the A. thaliana SUPERMAN Cys₂ His₂ zinc finger induces structural rearrangements of typical DNA base determinant positions.

    Science.gov (United States)

    Malgieri, Gaetano; Zaccaro, Laura; Leone, Marilisa; Bucci, Enrico; Esposito, Sabrina; Baglivo, Ilaria; Del Gatto, Annarita; Russo, Luigi; Scandurra, Roberto; Pedone, Paolo V; Fattorusso, Roberto; Isernia, Carla

    2011-11-01

    Among heavy metals, whose toxicity cause a steadily increasing of environmental pollution, cadmium is of special concern due to its relatively high mobility in soils and potential toxicity at low concentrations. Given their ubiquitous role, zinc fingers domains have been proposed as mediators for the toxic and carcinogenic effects exerted by xenobiotic metals. To verify the structural effects of zinc replacement by cadmium in zinc fingers, we have determined the high resolution structure of the single Cys₂ His₂ zinc finger of the Arabidopsis thaliana SUPERMAN protein (SUP37) complexed to the cadmium ion by means of UV-vis and NMR techniques. SUP37 is able to bind Cd(II), though with a dissociation constant higher than that measured for Zn(II). Cd-SUP37 retains the ββα fold but experiences a global structural rearrangement affecting both the relative orientation of the secondary structure elements and the position of side chains involved in DNA recognition: among them Ser17 side chain, which we show to be essential for DNA binding, experiences the largest displacement.

  11. Embryonic neural inducing factor churchill is not a DNA-binding zinc finger protein: solution structure reveals a solvent-exposed beta-sheet and zinc binuclear cluster.

    Science.gov (United States)

    Lee, Brian M; Buck-Koehntop, Bethany A; Martinez-Yamout, Maria A; Dyson, H Jane; Wright, Peter E

    2007-08-31

    Churchill is a zinc-containing protein that is involved in neural induction during embryogenesis. At the time of its discovery, it was thought on the basis of sequence alignment to contain two zinc fingers of the C4 type. Further, binding of an N-terminal GST-Churchill fusion protein to a particular DNA sequence was demonstrated by immunoprecipitation selection assay, suggesting that Churchill may function as a transcriptional regulator by sequence-specific DNA binding. We show by NMR solution structure determination that, far from containing canonical C4 zinc fingers, the protein contains three bound zinc ions in novel coordination sites, including an unusual binuclear zinc cluster. The secondary structure of Churchill is also unusual, consisting of a highly solvent-exposed single-layer beta-sheet. Hydrogen-deuterium exchange and backbone relaxation measurements reveal that Churchill is unusually dynamic on a number of time scales, with the exception of regions surrounding the zinc coordinating sites, which serve to stabilize the otherwise unstructured N terminus and the single-layer beta-sheet. No binding of Churchill to the previously identified DNA sequence could be detected, and extensive searches using DNA sequence selection techniques could find no other DNA sequence that was bound by Churchill. Since the N-terminal amino acids of Churchill form part of the zinc-binding motif, the addition of a fusion protein at the N terminus causes loss of zinc and unfolding of Churchill. This observation most likely explains the published DNA-binding results, which would arise due to non-specific interaction of the unfolded protein in the immunoprecipitation selection assay. Since Churchill does not appear to bind DNA, we suggest that it may function in embryogenesis as a protein-interaction factor.

  12. Involvement of C2H2 zinc finger proteins in the regulation of epidermal cell fate determination in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    An Yan; Minjie Wu; Yongqin Zhao; Aidong Zhang; Bohan Liu; John Schiefelbein; Yinbo Gan

    2014-01-01

    Cell fate determination is a basic developmental process during the growth of multicellular organisms. Trichomes and root hairs of Arabidopsis are both readily accessible structures originating from the epidermal cells of the aerial tissues and roots respectively, and they serve as excellent models for understanding the molecular mecha-nisms controlling cell fate determination and cell morphogen-esis. The regulation of trichome and root hair formation is a complex program that consists of the integration of hormonal signals with a large number of transcriptional factors, including MYB and bHLH transcriptional factors. Studies during recent years have uncovered an important role of C2H2 type zinc finger proteins in the regulation of epidermal cell fate determination. Here in this minireview we briefly summarize the involvement of C2H2 zinc finger proteins in the control of trichome and root hair formation in Arabidopsis.

  13. A poxvirus protein with a RING finger motif binds zinc and localizes in virus factories.

    Science.gov (United States)

    Upton, C; Schiff, L; Rice, S A; Dowdeswell, T; Yang, X; McFadden, G

    1994-07-01

    Shope fibroma virus (SFV) is a Leporipoxvirus closely related to the highly virulent myxoma virus. The DNA sequence of the BamHI N fragment of the SFV DNA genome was determined, and the single complete open reading frame (N1R) was characterized. The protein encoded by the N1R gene was found to contain a C3HC4 RING finger motif at the C terminus. This C3HC4 motif is the hallmark of a growing family of proteins, many of which are involved in regulation of gene expression, DNA repair, or DNA recombination. Complete homologs of the SFV N1R gene were also detected in variola virus, myxoma virus, and vaccinia virus strain IHD-W. In contrast, the gene is completely absent from vaccinia virus strain Copenhagen, and in vaccinia virus strain WR, the open reading frame is truncated prior to the zinc binding domain because of an 11-bp deletion, thus producing a frameshift and premature stop codon. Recombinant N1R protein from SFV was expressed in Escherichia coli and shown to bind zinc in a specific manner. Using fluorescence microscopy to visualize a peptide epitope tag (derived from ICP27 of herpes simplex virus) fused to the N terminus of the poxvirus proteins, we observed that the N1R protein of SFV and its homologs in myxoma virus and vaccinia virus IHD-W were localized primarily to the virus factories in the cytoplasm of infected cells and, to a lesser degree, the host cell nucleus. The truncated protein of vaccinia virus strain WR failed to localize in this manner but instead was observed throughout the cytoplasm.

  14. An effective approach for generating a three-Cys2His2 zinc-finger-DNA complex model by docking

    Directory of Open Access Journals (Sweden)

    Rajasekaran M

    2010-06-01

    Full Text Available Abstract Background Determination of protein-DNA complex structures with both NMR and X-ray crystallography remains challenging in many cases. High Ambiguity-Driven DOCKing (HADDOCK is an information-driven docking program that has been used to successfully model many protein-DNA complexes. However, a protein-DNA complex model whereby the protein wraps around DNA has not been reported. Defining the ambiguous interaction restraints for the classical three-Cys2His2 zinc-finger proteins that wrap around DNA is critical because of the complicated binding geometry. In this study, we generated a Zif268-DNA complex model using three different sets of ambiguous interaction restraints (AIRs to study the effect of the geometric distribution on the docking and used this approach to generate a newly reported Sp1-DNA complex model. Results The complex models we generated on the basis of two AIRs with a good geometric distribution in each domain are reasonable in terms of the number of models with wrap-around conformation, interface root mean square deviation, AIR energy and fraction native contacts. We derived the modeling approach for generating a three-Cys2His2 zinc-finger-DNA complex model according to the results of docking studies using the Zif268-DNA and other three crystal complex structures. Furthermore, the Sp1-DNA complex model was calculated with this approach, and the interactions between Sp1 and DNA are in good agreement with those previously reported. Conclusions Our docking data demonstrate that two AIRs with a reasonable geometric distribution in each of the three-Cys2His2 zinc-finger domains are sufficient to generate an accurate complex model with protein wrapping around DNA. This approach is efficient for generating a zinc-finger protein-DNA complex model for unknown complex structures in which the protein wraps around DNA. We provide a flowchart showing the detailed procedures of this approach.

  15. Myeloid zinc finger 1 mediates sulindac sulfide-induced upregulation of death receptor 5 of human colon cancer cells

    OpenAIRE

    Mano Horinaka; Tatsushi Yoshida; Mitsuhiro Tomosugi; Shusuke Yasuda; Yoshihiro Sowa; Toshiyuki Sakai

    2014-01-01

    A combined therapy of sulindac sulfide and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising strategy for the treatment of cancer. Sulindac sulfide had been shown to induce the expression of death receptor 5 (DR5), a receptor for TRAIL, and sensitize cancer cells to TRAIL-induced apoptosis; however, the molecular mechanism underlying the upregulation of DR5 has not yet been elucidated. We demonstrate here that myeloid zinc finger 1 (MZF1) mediates the induction of...

  16. p53 Gene repair with zinc finger nucleases optimised by yeast 1-hybrid and validated by Solexa sequencing.

    Directory of Open Access Journals (Sweden)

    Frank Herrmann

    Full Text Available The tumor suppressor gene p53 is mutated or deleted in over 50% of human tumors. As functional p53 plays a pivotal role in protecting against cancer development, several strategies for restoring wild-type (wt p53 function have been investigated. In this study, we applied an approach using gene repair with zinc finger nucleases (ZFNs. We adapted a commercially-available yeast one-hybrid (Y1H selection kit to allow rapid building and optimization of 4-finger constructs from randomized PCR libraries. We thus generated novel functional zinc finger nucleases against two DNA sites in the human p53 gene, near cancer mutation 'hotspots'. The ZFNs were first validated using in vitro cleavage assays and in vivo episomal gene repair assays in HEK293T cells. Subsequently, the ZFNs were used to restore wt-p53 status in the SF268 human cancer cell line, via ZFN-induced homologous recombination. The frequency of gene repair and mutation by non-homologous end-joining was then ascertained in several cancer cell lines, using a deep sequencing strategy. Our Y1H system facilitates the generation and optimisation of novel, sequence-specific four- to six-finger peptides, and the p53-specific ZFN described here can be used to mutate or repair p53 in genomic loci.

  17. p53 Gene Repair with Zinc Finger Nucleases Optimised by Yeast 1-Hybrid and Validated by Solexa Sequencing

    Science.gov (United States)

    Herrmann, Frank; Garriga-Canut, Mireia; Baumstark, Rebecca; Fajardo-Sanchez, Emmanuel; Cotterell, James; Minoche, André; Himmelbauer, Heinz; Isalan, Mark

    2011-01-01

    The tumor suppressor gene p53 is mutated or deleted in over 50% of human tumors. As functional p53 plays a pivotal role in protecting against cancer development, several strategies for restoring wild-type (wt) p53 function have been investigated. In this study, we applied an approach using gene repair with zinc finger nucleases (ZFNs). We adapted a commercially-available yeast one-hybrid (Y1H) selection kit to allow rapid building and optimization of 4-finger constructs from randomized PCR libraries. We thus generated novel functional zinc finger nucleases against two DNA sites in the human p53 gene, near cancer mutation ‘hotspots’. The ZFNs were first validated using in vitro cleavage assays and in vivo episomal gene repair assays in HEK293T cells. Subsequently, the ZFNs were used to restore wt-p53 status in the SF268 human cancer cell line, via ZFN-induced homologous recombination. The frequency of gene repair and mutation by non-homologous end-joining was then ascertained in several cancer cell lines, using a deep sequencing strategy. Our Y1H system facilitates the generation and optimisation of novel, sequence-specific four- to six-finger peptides, and the p53-specific ZFN described here can be used to mutate or repair p53 in genomic loci. PMID:21695267

  18. Cloning and characterization of SmZF1, a gene encoding a Schistosoma mansoni zinc finger protein

    Directory of Open Access Journals (Sweden)

    Souza Paulo R Eleutério de

    2001-01-01

    Full Text Available The zinc finger motifs (Cys2His2 are found in several proteins playing a role in the regulation of transcripton. SmZF1, a Schistosoma mansoni gene encoding a zinc finger protein was initially isolated from an adult worm cDNA library, as a partial cDNA. The full sequence of the gene was obtained by subcloning and sequencing cDNA and genomic fragments. The collated gene sequence is 2181 nt and the complete cDNA sequence is 705 bp containing the full open reading frame of the gene. Analysis of the genome sequence revealed the presence of three introns interrupting the coding region. The open reading frame theoretically encodes a protein of 164 amino acids, with a calculated molecular mass of 18,667Da. The predicted protein contains three zinc finger motifs, usually present in transcription regulatory proteins. PCR amplification with specific primers for the gene allowed for the detection of the target in egg, cercariae, schistosomulum and adult worm cDNA libraries indicating the expression of the mRNA in these life cycle stages of S. mansoni. This pattern of expression suggests the gene plays a role in vital functions of different life cycle stages of the parasite. Future research will be directed to elucidate the functional role of SmZF1.

  19. Non-transgenic genome modifications in a hemimetabolous insect using zinc-finger and TAL effector nucleases.

    Science.gov (United States)

    Watanabe, Takahito; Ochiai, Hiroshi; Sakuma, Tetsushi; Horch, Hadley W; Hamaguchi, Naoya; Nakamura, Taro; Bando, Tetsuya; Ohuchi, Hideyo; Yamamoto, Takashi; Noji, Sumihare; Mito, Taro

    2012-01-01

    Hemimetabolous, or incompletely metamorphosing, insects are phylogenetically relatively basal and comprise many pests. However, the absence of a sophisticated genetic model system, or targeted gene-manipulation system, has limited research on hemimetabolous species. Here we use zinc-finger nuclease and transcription activator-like effector nuclease technologies to produce genetic knockouts in the hemimetabolous insect Gryllus bimaculatus. Following the microinjection of mRNAs encoding zinc-finger nucleases or transcription activator-like effector nucleases into cricket embryos, targeting of a transgene or endogenous gene results in sequence-specific mutations. Up to 48% of founder animals transmit disrupted gene alleles after zinc-finger nucleases microinjection compared with 17% after microinjection of transcription activator-like effector nucleases. Heterozygous offspring is selected using mutation detection assays that use a Surveyor (Cel-I) nuclease, and subsequent sibling crosses create homozygous knockout crickets. This approach is independent from a mutant phenotype or the genetic tractability of the organism of interest and can potentially be applied to manage insect pests using a non-transgenic strategy.

  20. Zinc finger nucleases and their application%锌指核酶技术的原理及应用

    Institute of Scientific and Technical Information of China (English)

    邓珊珊; 王颖芝; 马端

    2010-01-01

    锌指核酶(zinc finger nucleases,ZFNs)是将锌指蛋白的DNA识别域和非特异性核酸内切酶FokⅠ人工连接而构成的一种核酶.1对ZFNs能在DNA上产生双链断裂(double-strand breaks,DSB),诱导细胞发生同源重组(homology recombination,HR)或非同源末端连接(nonhomologous end joining,NHEJ).最近锌指核酶技术在基因功能的研究中受到重视.作者就ZFNs的作用原理、关键技术及其应用领域进行介绍.%Zinc finger nuclease (ZFN), which is a chimeric fusion structure between a Cys2-His2 zinc-finger protein (ZFP) and the cleavage domain of Fok Ⅰ endonuclease,can be used to introduce targeted double-stranded breaks (DSBs). ZFN-mediated cleavage leads to mutations when double-stranded breaks are repaired by homologous recombination (HR) or nonhomologous end joining (NHEJ). In recent years, ZFNs are widely used in the fields of genetic research. In this review, the methodology and technical advantages of ZFNs were briefly discussed.

  1. Induction of Fetal Hemoglobin In Vivo Mediated by a Synthetic γ-Globin Zinc Finger Activator

    Directory of Open Access Journals (Sweden)

    Flávia C. Costa

    2012-01-01

    Full Text Available Sickle cell disease (SCD and β-thalassemia patients are phenotypically normal if they carry compensatory hereditary persistence of fetal hemoglobin (HPFH mutations that result in increased levels of fetal hemoglobin (HbF, γ-globin chains in adulthood. Thus, research has focused on manipulating the reactivation of γ-globin gene expression during adult definitive erythropoiesis as the most promising therapy to treat these hemoglobinopathies. Artificial transcription factors (ATFs are synthetic proteins designed to bind at a specific DNA sequence and modulate gene expression. The artificial zinc finger gg1-VP64 was designed to target the −117 region of the Aγ-globin gene proximal promoter and activate expression of this gene. Previous studies demonstrated that HbF levels were increased in murine chemical inducer of dimerization (CID-dependent bone marrow cells carrying a human β-globin locus yeast artificial chromosome (β-YAC transgene and in CD34+ erythroid progenitor cells from normal donors and β-thalassemia patients. Herein, we report that gg1-VP64 increased γ-globin gene expression in vivo, in peripheral blood samples from gg1-VP64 β-YAC double-transgenic (bigenic mice. Our results demonstrate that ATFs function in an animal model to increase gene expression. Thus, this class of reagent may be an effective gene therapy for treatment of some inherited diseases.

  2. Zinc finger protein 521 overexpression increased transcript levels of Fndc5 in mouse embryonic stem cells

    Indian Academy of Sciences (India)

    Motahere-Sadat Hashemi; Abbas Kiani Esfahani; Maryam Peymani; Alireza Shoaraye Nejati; Kamran Ghaedi; Mohammad Hossein Nasr-Esfahani; Hossein Baharvand

    2016-03-01

    Zinc finger protein 521 is highly expressed in brain, neural stem cells and early progenitors of the human hematopoietic cells. Zfp521 triggers the cascade of neurogenesis inmouse embryonic stemcells through inducing expression of the early neuroectodermal genes Sox1, Sox3 and Pax6. Fndc5, a precursor of Irisin has inducing effects on the expression level of brain derived neurotrophic factor in hippocampus. Therefore, it is most likely that Fndc5 may play an important role in neural differentiation. To exhibit whether the expression of this protein is under regulation with Zfp521, we overexpressed Zfp521 in a stable transformants of mESCs expressing EGFP under control of Fndc5 promoter. Increased expression of Zfp521 enhanced transcription levels of both EGFP and endogenous Fndc5. This result was confirmed by overexpression the aforementioned vectors in HEK cells and indicated that Zfp521 functions upstream of Fndc5 expression. It is most likely that Zfp521 may act through the binding to its response element on Fndc5 core promoter. Therefore it is concluding that an enhanced expression of Fndc5 in neural progenitor cells is stimulated by Zfp521 overexpression in these cells.

  3. The Drosophila Zinc Finger Transcription Factor Ouija Board Controls Ecdysteroid Biosynthesis through Specific Regulation of spookier.

    Directory of Open Access Journals (Sweden)

    Tatsuya Komura-Kawa

    2015-12-01

    Full Text Available Steroid hormones are crucial for many biological events in multicellular organisms. In insects, the principal steroid hormones are ecdysteroids, which play essential roles in regulating molting and metamorphosis. During larval and pupal development, ecdysteroids are synthesized in the prothoracic gland (PG from dietary cholesterol via a series of hydroxylation and oxidation steps. The expression of all but one of the known ecdysteroid biosynthetic enzymes is restricted to the PG, but the transcriptional regulatory networks responsible for generating such exquisite tissue-specific regulation is only beginning to be elucidated. Here, we report identification and characterization of the C2H2-type zinc finger transcription factor Ouija board (Ouib necessary for ecdysteroid production in the PG in the fruit fly Drosophila melanogaster. Expression of ouib is predominantly limited to the PG, and genetic null mutants of ouib result in larval developmental arrest that can be rescued by administrating an active ecdysteroid. Interestingly, ouib mutant animals exhibit a strong reduction in the expression of one ecdysteroid biosynthetic enzyme, spookier. Using a cell culture-based luciferase reporter assay, Ouib protein stimulates transcription of spok by binding to a specific ~15 bp response element in the spok PG enhancer element. Most remarkable, the developmental arrest phenotype of ouib mutants is rescued by over-expression of a functionally-equivalent paralog of spookier. These observations imply that the main biological function of Ouib is to specifically regulate spookier transcription during Drosophila development.

  4. Identification of SCAN domain zinc-finger gene ZNF449 as a novel factor of chondrogenesis.

    Directory of Open Access Journals (Sweden)

    Keita Okada

    Full Text Available Transcription factors SOX9, SOX5 and SOX6 are indispensable for generation and differentiation of chondrocytes. However, molecular mechanisms to induce the SOX genes are poorly understood. To address this issue, we previously determined the human embryonic enhancer of SOX6 by 5'RACE analysis, and identified the 46-bp core enhancer region (CES6. We initially performed yeast one-hybrid assay for screening other chondrogenic factors using CES6 as bait, and identified a zinc finger protein ZNF449. ZNF449 and Zfp449, a counterpart in mouse, transactivated enhancers or promoters of SOX6, SOX9 and COL2A1. Zfp449 was expressed in mesenchyme-derived tissues including cartilage, calvaria, muscle and tendon, as well as in other tissues including brain, lung and kidney. In limb cartilage of mouse embryo, Zfp449 protein was abundantly located in periarticular chondrocytes, and decreased in accordance with the differentiation. Zfp449 protein was also detected in articular cartilage of an adult mouse. During chondrogenic differentiation of human mesenchymal stem cells, ZNF449 was increased at an early stage, and its overexpression enhanced SOX9 and SOX6 only at the initial stage of the differentiation. We further generated Zfp449 knockout mice to examine the in vivo roles; however, no obvious abnormality was observed in skeletal development or articular cartilage homeostasis. ZNF449 may regulate chondrogenic differentiation from mesenchymal progenitor cells, although the underlying mechanisms are still unknown.

  5. A FYVE zinc finger domain protein specifically links mRNA transport to endosome trafficking.

    Science.gov (United States)

    Pohlmann, Thomas; Baumann, Sebastian; Haag, Carl; Albrecht, Mario; Feldbrügge, Michael

    2015-05-18

    An emerging theme in cellular logistics is the close connection between mRNA and membrane trafficking. A prominent example is the microtubule-dependent transport of mRNAs and associated ribosomes on endosomes. This coordinated process is crucial for correct septin filamentation and efficient growth of polarised cells, such as fungal hyphae. Despite detailed knowledge on the key RNA-binding protein and the molecular motors involved, it is unclear how mRNAs are connected to membranes during transport. Here, we identify a novel factor containing a FYVE zinc finger domain for interaction with endosomal lipids and a new PAM2-like domain required for interaction with the MLLE domain of the key RNA-binding protein. Consistently, loss of this FYVE domain protein leads to specific defects in mRNA, ribosome, and septin transport without affecting general functions of endosomes or their movement. Hence, this is the first endosomal component specific for mRNP trafficking uncovering a new mechanism to couple mRNPs to endosomes.

  6. Identification of Mushroom body miniature, a zinc-finger protein implicated in brain development of Drosophila

    Science.gov (United States)

    Raabe, Thomas; Clemens-Richter, Susanne; Twardzik, Thomas; Ebert, Anselm; Gramlich, Gertrud; Heisenberg, Martin

    2004-01-01

    The mushroom bodies are bilaterally arranged structures in the protocerebrum of Drosophila and most other insect species. Mutants with altered mushroom body structure have been instrumental not only in establishing their role in distinct behavioral functions but also in identifying the molecular pathways that control mushroom body development. The mushroom body miniature1 (mbm1) mutation results in grossly reduced mushroom bodies and odor learning deficits in females. With a survey of genomic rescue constructs, we have pinpointed mbm1 to a single transcription unit and identified a single nucleotide exchange in the 5′ untranslated region of the corresponding transcript resulting in a reduced expression of the protein. The most obvious feature of the Mbm protein is a pair of C2HC zinc fingers, implicating a function of the protein in binding nucleic acids. Immunohistochemical analysis shows that expression of the Mbm protein is not restricted to the mushroom bodies. BrdUrd labeling experiments indicate a function of Mbm in neuronal precursor cell proliferation. PMID:15375215

  7. Zinc finger protein 148 is dispensable for primitive and definitive hematopoiesis in mice.

    Directory of Open Access Journals (Sweden)

    Anna Nilton

    Full Text Available Hematopoiesis is regulated by transcription factors that induce cell fate and differentiation in hematopoietic stem cells into fully differentiated hematopoietic cell types. The transcription factor zinc finger protein 148 (Zfp148 interacts with the hematopoietic transcription factor Gata1 and has been implicated to play an important role in primitive and definitive hematopoiesis in zebra fish and mouse chimeras. We have recently created a gene-trap knockout mouse model deficient for Zfp148, opening up for analyses of hematopoiesis in a conventional loss-of-function model in vivo. Here, we show that Zfp148-deficient neonatal and adult mice have normal or slightly increased levels of hemoglobin, hematocrit, platelets and white blood cells, compared to wild type controls. Hematopoietic lineages in bone marrow, thymus and spleen from Zfp148 (gt/gt mice were further investigated by flow cytometry. There were no differences in T-cells (CD4 and CD8 single positive cells, CD4 and CD8 double negative/positive cells in either organ. However, the fraction of CD69- and B220-positive cells among lymphocytes in spleen was slightly lower at postnatal day 14 in Zfp148 (gt/gt mice compared to wild type mice. Our results demonstrate that Zfp148-deficient mice generate normal mature hematopoietic populations thus challenging earlier studies indicating that Zfp148 plays a critical role during hematopoietic development.

  8. Expression of the zinc-finger antiviral protein inhibits alphavirus replication.

    Science.gov (United States)

    Bick, Matthew J; Carroll, John-William N; Gao, Guangxia; Goff, Stephen P; Rice, Charles M; MacDonald, Margaret R

    2003-11-01

    The rat zinc-finger antiviral protein (ZAP) was recently identified as a host protein conferring resistance to retroviral infection. We analyzed ZAP's ability to inhibit viruses from other families and found that ZAP potently inhibits the replication of multiple members of the Alphavirus genus within the Togaviridae, including Sindbis virus, Semliki Forest virus, Ross River virus, and Venezuelan equine encephalitis virus. However, expression of ZAP did not induce a broad-spectrum antiviral state as some viruses, including vesicular stomatitis virus, poliovirus, yellow fever virus, and herpes simplex virus type 1, replicated to normal levels in ZAP-expressing cells. We determined that ZAP expression inhibits Sindbis virus replication after virus penetration and entry, but before the amplification of newly synthesized plus strand genomic RNA. Using a temperature-sensitive Sindbis virus mutant expressing luciferase, we further showed that translation of incoming viral RNA is blocked by ZAP expression. Elucidation of the antiviral mechanism by which ZAP inhibits Sindbis virus translation may lead to the development of agents with broad activity against alphaviruses.

  9. Abnormal spermatogenesis and male infertility in testicular zinc finger protein Zfp318-knockout mice.

    Science.gov (United States)

    Ishizuka, Masamichi; Ohtsuka, Eri; Inoue, Atsuto; Odaka, Mirei; Ohshima, Hirotaka; Tamura, Norihisa; Yoshida, Kaoru; Sako, Norihisa; Baba, Tadashi; Kashiwabara, Shin-Ichi; Okabe, Masaru; Noguchi, Junko; Hagiwara, Hiromi

    2016-09-01

    Zfp318, a mouse gene with a Cys2/His2 zinc finger motif, is mainly expressed in germ cells in the testis. It encodes two alternative transcripts, which regulate androgen receptor-mediated transcriptional activation or repression by overexpression of them. However, the role of Zfp318 is still obscure in vivo, especially in spermatogenesis. To elucidate the role of Zfp318 during gamete production, we established a knockout mouse line. Zfp318-null male mice exhibited infertility, whereas Zfp318-null female mice displayed normal fertility. ZFP318 was expressed during multiple stages of spermatogenesis, from spermatocytes to round spermatids. The nuclei of secondary spermatocytes showed high levels of expression. Histological analysis and quantitative analysis of DNA content showed decreased numbers of both spermatids in the seminiferous tubules and mature spermatozoa in the epididymides of Zfp318-null mice. These results suggest that Zfp318 is expressed as a functional protein in testicular germ cells and plays an important role in meiosis during spermatogenesis.

  10. An elastic-network based local molecular field analysis of zinc-finger proteins

    CERN Document Server

    Dixit, Purushottam D

    2011-01-01

    We study two designed and one natural zinc-finger peptide each with the Cys2His2 (CCHH) type of metal binding motif. In the approach we have developed, we describe the role of the protein and solvent outside the Zn(II)-CCHH metal-residue cluster by a molecular field represented by generalized harmonic restraints. The strength of the field is adjusted to reproduce the binding energy distribution of the metal with the cluster obtained in a reference all-atom simulation with empirical potentials. The quadratic field allows us to investigate analytically the protein restraints on the binding site in terms of its eigenmodes. Examining these eigenmodes suggests, consistent with experimental observations, the importance of the first histidine (H) in the CCHH cluster in metal binding. Further, the eigenvalues corresponding to these modes also indicate that the designed proteins form a tighter complex with the metal. We find that the bulk protein and solvent response tends to destabilize metal-binding, emphasizing tha...

  11. Expression and prognostic significance of zinc fingers and homeoboxes family members in renal cell carcinoma

    Science.gov (United States)

    Jeong, Dae Cheon; Han, Myoung-Eun; Kim, Ji-Young; Liu, Liangwen; Jung, Jin-Sup; Oh, Sae-Ock

    2017-01-01

    Zinc fingers and homeoboxes (ZHX) is a transcription repressor family that contains three members; ZHX1, ZHX2, and ZHX3. Although ZHX family members have been associated with the progression of cancer, their values as prognostic factors in cancer patients have been poorly examined. Renal cell carcinoma (RCC) is a highly heterogeneous, aggressive cancer that responds variably to treatment. Thus, prognostic molecular markers are required to evaluate disease progression and to improve the survival. In clear cell RCC (ccRCC), ZHX1 and ZHX3 expression were found to be down-regulated but ZHX2 was up-regulated, and the expressions of ZHX1 and ZHX3 were significantly associated with pathological stage. Furthermore, Kaplan-Meier and multivariate regression analysis showed that reduction in the mRNA expression of ZHX1 was associated with poorer survival. Taken together, the present study shows loss of ZHX1 is correlated with ccRCC progression and suggests it is an independent prognostic marker in ccRCC. PMID:28152006

  12. Molecular cloning and expression analysis of a zebrafish novel zinc finger protein gene rnf141

    Directory of Open Access Journals (Sweden)

    Wenqian Deng

    2009-01-01

    Full Text Available ZNF230 is a novel zinc finger gene cloned by our laboratory. In order to understand the potential functions of this gene in vertebrate development, we cloned the zebrafish orthologue of human ZNF230, named rnf141. The cDNA fragment of rnf141 was obtained by rapid amplification of cDNA ends (RACE. The open reading frame (ORF encodes a polypeptide of 222 amino acids which shares 75.65% identity with the human ZNF230. RT-PCR analysis in zebrafish embryo and adult tissues revealed that rnf141 transcripts are maternally derived and that rnf141 mRNA has a broad distribution. Zygotic rnf141 message is strongly localized in the central nervous system, as shown by whole-mount in situ hybridization. Knockdown and over expression of rnf141 can induce abnormal phenotypes, including abnormal development of brain, as well as yolk sac and axis extendsion. Marker gene analysis showed that rnf141 may play a role in normal dorsoventral patterning of zebrafish embryos, suggesting that rnf141 may have a broad function during early development of vertebrates.

  13. Targeted genome editing by lentiviral protein transduction of zinc-finger and TAL-effector nucleases.

    Science.gov (United States)

    Cai, Yujia; Bak, Rasmus O; Mikkelsen, Jacob Giehm

    2014-04-24

    Future therapeutic use of engineered site-directed nucleases, like zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs), relies on safe and effective means of delivering nucleases to cells. In this study, we adapt lentiviral vectors as carriers of designer nuclease proteins, providing efficient targeted gene disruption in vector-treated cell lines and primary cells. By co-packaging pairs of ZFN proteins with donor RNA in 'all-in-one' lentiviral particles, we co-deliver ZFN proteins and the donor template for homology-directed repair leading to targeted DNA insertion and gene correction. Comparative studies of ZFN activity in a predetermined target locus and a known nearby off-target locus demonstrate reduced off-target activity after ZFN protein transduction relative to conventional delivery approaches. Additionally, TALEN proteins are added to the repertoire of custom-designed nucleases that can be delivered by protein transduction. Altogether, our findings generate a new platform for genome engineering based on efficient and potentially safer delivery of programmable nucleases.DOI: http://dx.doi.org/10.7554/eLife.01911.001.

  14. TRIM25 Enhances the Antiviral Action of Zinc-Finger Antiviral Protein (ZAP)

    Science.gov (United States)

    Lau, Zerlina; Cheung, Pamela; Schneider, William M.; Bozzacco, Leonia; Buehler, Eugen; Takaoka, Akinori; Rice, Charles M.; Felsenfeld, Dan P.; MacDonald, Margaret R.

    2017-01-01

    The host factor and interferon (IFN)-stimulated gene (ISG) product, zinc-finger antiviral protein (ZAP), inhibits a number of diverse viruses by usurping and intersecting with multiple cellular pathways. To elucidate its antiviral mechanism, we perform a loss-of-function genome-wide RNAi screen to identify cellular cofactors required for ZAP antiviral activity against the prototype alphavirus, Sindbis virus (SINV). In order to exclude off-target effects, we carry out stringent confirmatory assays to verify the top hits. Important ZAP-liaising partners identified include proteins involved in membrane ion permeability, type I IFN signaling, and post-translational protein modification. The factor contributing most to the antiviral function of ZAP is TRIM25, an E3 ubiquitin and ISG15 ligase. We demonstrate here that TRIM25 interacts with ZAP through the SPRY domain, and TRIM25 mutants lacking the RING or coiled coil domain fail to stimulate ZAP’s antiviral activity, suggesting that both TRIM25 ligase activity and its ability to form oligomers are critical for its cofactor function. TRIM25 increases the modification of both the short and long ZAP isoforms by K48- and K63-linked polyubiquitin, although ubiquitination of ZAP does not directly affect its antiviral activity. However, TRIM25 is critical for ZAP’s ability to inhibit translation of the incoming SINV genome. Taken together, these data uncover TRIM25 as a bona fide ZAP cofactor that leads to increased ZAP modification enhancing its translational inhibition activity. PMID:28060952

  15. Zinc finger antiviral protein inhibits coxsackievirus B3 virus replication and protects against viral myocarditis.

    Science.gov (United States)

    Li, Min; Yan, Kepeng; Wei, Lin; Yang, Jie; Lu, Chenyu; Xiong, Fei; Zheng, Chunfu; Xu, Wei

    2015-11-01

    The host Zinc finger antiviral protein (ZAP) has been reported exhibiting antiviral activity against positive-stranded RNA viruses (Togaviridae), negative-stranded RNA viruses (Filoviridae) and retroviruses (Retroviridae). However, whether ZAP restricts the infection of enterovirus and the development of enterovirus mediated disease remains unknown. Here, we reported the antiviral properties of ZAP against coxsackievirus B3 (CVB3), a single-stranded RNA virus of the Enterovirus genus within the Picornaviridae as a major causative agent of viral myocarditis (VMC). We found that the expression of ZAP was significantly induced after CVB3 infection in heart tissues of VMC mice. ZAP potently inhibited CVB3 replication in cells after infection, while overexpression of ZAP in mice significantly increased the resistance to CVB3 replication and viral myocarditis by significantly reducing cardiac inflammatory cytokine production. The ZAP-responsive elements (ZREs) were mapped to the 3'UTR and 5'UTR of viral RNA. Taken together, ZAP confers resistance to CVB3 infection via directly targeting viral RNA and protects mice from acute myocarditis by suppressing viral replication and cardiac inflammatory cytokine production. Our finding further expands ZAP's range of viral targets, and suggests ZAP as a potential therapeutic target for viral myocarditis caused by CVB3.

  16. Donor plasmid design for codon and single base genome editing using zinc finger nucleases.

    Science.gov (United States)

    Pruett-Miller, Shondra M; Davis, Gregory D

    2015-01-01

    In recent years, CompoZr zinc finger nuclease (ZFN) technology has matured to the point that a user-defined double strand break (DSB) can be placed at virtually any location in the human genome within 50 bp of a desired site. Such high resolution ZFN engineering is well within the conversion tract limitations demarcated by the mammalian DNA repair machinery, resulting in a nearly universal ability to create point mutations throughout the human genome. Additionally, new architectures for targeted nuclease engineering have been rapidly developed, namely transcription activator like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas systems, further expanding options for placement of DSBs. This new capability has created a need to explore the practical limitations of delivering plasmid-based information to the sites of chromosomal double strand breaks so that nuclease-donor methods can be widely deployed in fundamental and therapeutic research. In this chapter, we explore a ZFN-compatible donor design in the context of codon changes at an endogenous locus encoding the human RSK2 kinase.

  17. Characterization of the target DNA sequence for the DNA-binding domain of zinc finger protein 191

    Institute of Scientific and Technical Information of China (English)

    Haoyue Wang; Ruilin Sun; Guoxiang Liu; Minghui Yao; Jian Fei; Hebai Shen

    2008-01-01

    Studies on the DNA-binding properties of transcription factors are important in searching for the downstream genes regulated by these factors. In the present study, we report on the DNA-binding property of a Cys2His2-type transcription factor, zinc finger protein 191 (Zfp191), which has been newly found to play a significant role in mice.By constructing a fusion protein containing the DNA-binding domain of Zfp191,we characterized target DNA by determining the protein's binding specificity and dependence on zinc.The data showed that the DNA-binding domain of Zfp191can specifically bind to the TCAT repeat motif and that there is a cooperative effect among the target DNA's multiple binding sites.Furthermore,the binding reaction is dependent on zinc.This work provides a foundation for further studies on the role of Zfp191 in gene regulation and development.

  18. Generation and genetic engineering of human induced pluripotent stem cells using designed zinc finger nucleases.

    Science.gov (United States)

    Ramalingam, Sivaprakash; London, Viktoriya; Kandavelou, Karthikeyan; Cebotaru, Liudmila; Guggino, William; Civin, Curt; Chandrasegaran, Srinivasan

    2013-02-15

    Zinc finger nucleases (ZFNs) have become powerful tools to deliver a targeted double-strand break at a pre-determined chromosomal locus in order to insert an exogenous transgene by homology-directed repair. ZFN-mediated gene targeting was used to generate both single-allele chemokine (C-C motif) receptor 5 (CCR5)-modified human induced pluripotent stem cells (hiPSCs) and biallele CCR5-modified hiPSCs from human lung fibroblasts (IMR90 cells) and human primary cord blood mononuclear cells (CBMNCs) by site-specific insertion of stem cell transcription factor genes flanked by LoxP sites into the endogenous CCR5 locus. The Oct4 and Sox2 reprogramming factors, in combination with valproic acid, induced reprogramming of human lung fibroblasts to form CCR5-modified hiPSCs, while 5 factors, Oct4/Sox2/Klf4/Lin28/Nanog, induced reprogramming of CBMNCs. Subsequent Cre recombinase treatment of the CCR5-modified IMR90 hiPSCs resulted in the removal of the Oct4 and Sox2 transgenes. Further genetic engineering of the single-allele CCR5-modified IMR90 hiPSCs was achieved by site-specific addition of the large CFTR transcription unit to the remaining CCR5 wild-type allele, using CCR5-specific ZFNs and a donor construct containing tdTomato and CFTR transgenes flanked by CCR5 homology arms. CFTR was expressed efficiently from the endogenous CCR5 locus of the CCR5-modified tdTomato/CFTR hiPSCs. These results suggest that it might be feasible to use ZFN-evoked strategies to (1) generate precisely targeted genetically well-defined patient-specific hiPSCs, and (2) then to reshape their function by targeted addition and expression of therapeutic genes from the CCR5 chromosomal locus for autologous cell-based transgene-correction therapy to treat various recessive monogenic human diseases in the future.

  19. A pair of mouse KRAB zinc finger proteins modulates multiple indicators of female reproduction.

    Science.gov (United States)

    Krebs, Christopher J; Robins, Diane M

    2010-04-01

    Krüppel-associated box-zinc finger proteins (KRAB-ZFPs) are the largest class of transcriptional regulators in mammals, yet few have been assigned biological roles. Cloning the genes underlying the regulator of sex-limitation (rsl) phenotype, in which the normally male-specific sex-limited protein (SLP) is expressed in female mice, identified two KRAB-ZFPs, Rsl1 and Rsl2, as influencing sexually dimorphic liver gene expression. Combined absence of both repressors in rsl mice leads to increased expression in female liver of major urinary proteins (MUPs) and certain enzymes of steroid metabolism, as well as SLP. We hypothesized that this altered gene expression might affect reproductive physiology in rsl females. Urinary MUP (uMUP) concentration varied with the estrous cycle in both wt and rsl females but was consistently higher in rsl urine. A behavioral odor test revealed that wild-type (wt) males preferred rsl to wt females, possibly due to elevated uMUPs providing greater pheromone presentation. To ascribe activity to Rsl1, Rsl2, or both, the genes were individually expressed as liver-specific transgenes. RSL2 overexpression accentuated uMUP fluctuations across the estrous cycle, whereas RSL1 overexpression did not. In addition, puberty onset, as indicated by vaginal opening (VO), occurred 2 days earlier in rsl females, and excess RSL2, but not RSL1, restored VO timing to wt. Hence, transcriptional repression by RSL in liver modifies female mouse reproduction via targets that likely impact both hormonal and pheromonal cues. The large and rapidly diversifying KRAB-ZFP family may modulate biological processes, including reproduction, to confer individual differences that may isolate populations and ultimately lead to speciation.

  20. Zinc finger protein A20 protects rats against chronic liver allograft dysfunction

    Institute of Scientific and Technical Information of China (English)

    Jie Yang; Ming-Qing Xu; Lu-Nan Yan; Xiao-Bo Chen; Jiao Liu

    2012-01-01

    AIM:To investigate the effect of zinc finger protein A20 on chronic liver allograft dysfunction in rats.METHODS:Allogeneic liver transplantation from DA rats to Lewis rats was performed.Chronic liver allograft dysfunction was induced in the rats by administering low-dose tacrolimus at postoperative day (POD) 5.Hepatic overexpression of A20 was achieved by recombinant adenovirus (rAd.)-mediated gene transfer administered intravenously every 10 d starting from POD 10.The recipient rats were injected with physiological saline,rAdEasy-A20 (1 x 109 pfu/30 g weight) or rAdEasy (1 x 109 pfu/30 g weight) every 10 d through the tail vein for 3 mo starting from POD 10.Liver tissue samples were harvested on POD 30 and POD 60.RESULTS:Liver-transplanted rats treated with only tacrolimus showed chronic allograft dysfunction with severe hepatic fibrosis.A20 overexpression ameliorated the effects on liver function,attenuated liver allograft fibrosis and prolonged the survival of the recipient rats.Treatment with A20 suppressed hepatic protein production of tumor growth factor (TGF)-β1,interleukin1β,caspase-8,CD40,CD40L,intercellular adhesion molecule-1,vascular cell adhesion molecule-1 and E-selectin.A20 treatment suppressed liver cell apoptosis and inhibited nuclear factor-κB activation of Kupffer cells (KCs),liver sinusoidal endothelial cells (LSECs)and hepatic stellate cells (HSCs),and it subsequently decreased cytokine mRNA expression in KCs and LSECs and reduced the production of TGF-β1 in HSCs.CONCLUSION:A20 might prevent chronic liver allograft dysfunction by re-establishing functional homeostasis of KCs,LSECs and HSCs.

  1. ZFN-Site searches genomes for zinc finger nuclease target sites and off-target sites

    Directory of Open Access Journals (Sweden)

    Iseli Christian

    2011-05-01

    Full Text Available Abstract Background Zinc Finger Nucleases (ZFNs are man-made restriction enzymes useful for manipulating genomes by cleaving target DNA sequences. ZFNs allow therapeutic gene correction or creation of genetically modified model organisms. ZFN specificity is not absolute; therefore, it is essential to select ZFN target sites without similar genomic off-target sites. It is important to assay for off-target cleavage events at sites similar to the target sequence. Results ZFN-Site is a web interface that searches multiple genomes for ZFN off-target sites. Queries can be based on the target sequence or can be expanded using degenerate specificity to account for known ZFN binding preferences. ZFN off-target sites are outputted with links to genome browsers, facilitating off-target cleavage site screening. We verified ZFN-Site using previously published ZFN half-sites and located their target sites and their previously described off-target sites. While we have tailored this tool to ZFNs, ZFN-Site can also be used to find potential off-target sites for other nucleases, such as TALE nucleases. Conclusions ZFN-Site facilitates genome searches for possible ZFN cleavage sites based on user-defined stringency limits. ZFN-Site is an improvement over other methods because the FetchGWI search engine uses an indexed search of genome sequences for all ZFN target sites and possible off-target sites matching the half-sites and stringency limits. Therefore, ZFN-Site does not miss potential off-target sites.

  2. Zinc-finger nuclease mediated disruption of Rag1 in the LEW/Ztm rat

    Directory of Open Access Journals (Sweden)

    Zschemisch Nils-Holger

    2012-11-01

    Full Text Available Abstract Background Engineered zinc-finger nucleases (ZFN represented an innovative method for the genome manipulation in vertebrates. ZFN introduced targeted DNA double strand breaks (DSB and initiated non-homologous end joining (NHEJ after pronuclear or cytoplasmatic microinjection into zygotes. Resulting frame shift mutations led to functional gene ablations in zebra fish, mice, pigs and also in laboratory rats. Therefore, we targeted the rat Rag1 gene essential for the V(DJ recombination within the immunoglobulin production process and for the differentiation of mature B and T lymphocytes to generate an immunodeficient rat model in the LEW/Ztm strain. Results After microinjection of Rag1 specific ZFN mRNAs in 623 zygotes of inbred LEW/Ztm rats 59 offspring were born from which one carried a 4 bp deletion. This frame shift mutation led to a premature stop codon and a subsequently truncated Rag1 protein confirmed by the loss of the full-length protein in Western Blot analysis. Truncation of the Rag1 protein was characterized by the complete depletion of mature B cells. The remaining T cell population contained mature CD4+/CD3+/TCRαβ+ as well as CD8+/CD3+/TCRαβ+ positive lymphocytes accompanied by a compensatory increase of natural killer cells in the peripheral blood. Reduction of T cell development in Rag1 mutant rats was associated with a hypoplastic thymus that lacked follicular structures. Histological evaluation also revealed the near-complete absence of lymphocytes in spleen and lymph nodes in the immunodeficient Rag1 mutant rat. Conclusion The Rag1 mutant rat will serve as an important model for transplantation studies. Furthermore, it may be used as a model for reconstitution experiments related to the immune system, particularly with respect to different populations of human lymphocytes, natural killer cells and autoimmune phenomena.

  3. Zebrafish foxP2 zinc finger nuclease mutant has normal axon pathfinding.

    Directory of Open Access Journals (Sweden)

    Lingyan Xing

    Full Text Available foxP2, a forkhead-domain transcription factor, is critical for speech and language development in humans, but its role in the establishment of CNS connectivity is unclear. While in vitro studies have identified axon guidance molecules as targets of foxP2 regulation, and cell culture assays suggest a role for foxP2 in neurite outgrowth, in vivo studies have been lacking regarding a role for foxP2 in axon pathfinding. We used a modified zinc finger nuclease methodology to generate mutations in the zebrafish foxP2 gene. Using PCR-based high resolution melt curve analysis (HRMA of G0 founder animals, we screened and identified three mutants carrying nonsense mutations in the 2(nd coding exon: a 17 base-pair (bp deletion, an 8bp deletion, and a 4bp insertion. Sequence analysis of cDNA confirmed that these were frameshift mutations with predicted early protein truncations. Homozygous mutant fish were viable and fertile, with unchanged body morphology, and no apparent differences in CNS apoptosis, proliferation, or patterning at embryonic stages. There was a reduction in expression of the known foxP2 target gene cntnap2 that was rescued by injection of wild-type foxP2 transcript. When we examined axon pathfinding using a pan-axonal marker or transgenic lines, including a foxP2-neuron-specific enhancer, we did not observe any axon guidance errors. Our findings suggest that foxP2 is not necessary for axon pathfinding during development.

  4. The Promyelocytic Leukemia Zinc Finger Transcription Factor Is Critical for Human Endometrial Stromal Cell Decidualization.

    Directory of Open Access Journals (Sweden)

    Ramakrishna Kommagani

    2016-04-01

    Full Text Available Progesterone, via the progesterone receptor (PGR, is essential for endometrial stromal cell decidualization, a cellular transformation event in which stromal fibroblasts differentiate into decidual cells. Uterine decidualization supports embryo implantation and placentation as well as subsequent events, which together ensure a successful pregnancy. Accordingly, impaired decidualization results not only in implantation failure or early fetal miscarriage, but also may lead to potential adverse outcomes in all three pregnancy trimesters. Transcriptional reprogramming on a genome-wide scale underlies progesterone dependent decidualization of the human endometrial stromal cell (hESC. However, identification of the functionally essential signals encoded by these global transcriptional changes remains incomplete. Importantly, this knowledge-gap undercuts future efforts to improve diagnosis and treatment of implantation failure based on a dysfunctional endometrium. By integrating genome-wide datasets derived from decidualization of hESCs in culture, we reveal that the promyelocytic leukemia zinc finger (PLZF transcription factor is rapidly induced by progesterone and that this induction is indispensable for progesterone-dependent decidualization. Chromatin immunoprecipitation followed by next generation sequencing (ChIP-Seq identified at least ten progesterone response elements within the PLZF gene, indicating that PLZF may act as a direct target of PGR signaling. The spatiotemporal expression profile for PLZF in both the human and mouse endometrium offers further support for stromal PLZF as a mediator of the progesterone decidual signal. To identify functional targets of PLZF, integration of PLZF ChIP-Seq and RNA Pol II RNA-Seq datasets revealed that the early growth response 1 (EGR1 transcription factor is a PLZF target for which its level of expression must be reduced to enable progesterone dependent hESC decidualization. Apart from furnishing

  5. Efficient immunoglobulin gene disruption and targeted replacement in rabbit using zinc finger nucleases.

    Directory of Open Access Journals (Sweden)

    Tatiana Flisikowska

    Full Text Available Rabbits are widely used in biomedical research, yet techniques for their precise genetic modification are lacking. We demonstrate that zinc finger nucleases (ZFNs introduced into fertilized oocytes can inactivate a chosen gene by mutagenesis and also mediate precise homologous recombination with a DNA gene-targeting vector to achieve the first gene knockout and targeted sequence replacement in rabbits. Two ZFN pairs were designed that target the rabbit immunoglobulin M (IgM locus within exons 1 and 2. ZFN mRNAs were microinjected into pronuclear stage fertilized oocytes. Founder animals carrying distinct mutated IgM alleles were identified and bred to produce offspring. Functional knockout of the immunoglobulin heavy chain locus was confirmed by serum IgM and IgG deficiency and lack of IgM(+ and IgG(+ B lymphocytes. We then tested whether ZFN expression would enable efficient targeted sequence replacement in rabbit oocytes. ZFN mRNA was co-injected with a linear DNA vector designed to replace exon 1 of the IgM locus with ∼1.9 kb of novel sequence. Double strand break induced targeted replacement occurred in up to 17% of embryos and in 18% of fetuses analyzed. Two major goals have been achieved. First, inactivation of the endogenous IgM locus, which is an essential step for the production of therapeutic human polyclonal antibodies in the rabbit. Second, establishing efficient targeted gene manipulation and homologous recombination in a refractory animal species. ZFN mediated genetic engineering in the rabbit and other mammals opens new avenues of experimentation in immunology and many other research fields.

  6. Knockout of exogenous EGFP gene in porcine somatic cells using zinc-finger nucleases

    Energy Technology Data Exchange (ETDEWEB)

    Watanabe, Masahito [Japan Science and Technology Agency (JST), ERATO, Nakauchi Stem Cell and Organ Regeneration Project, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639 (Japan); Department of Life Sciences, School of Agriculture, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki, Kanagawa 214-8571 (Japan); Umeyama, Kazuhiro [Department of Life Sciences, School of Agriculture, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki, Kanagawa 214-8571 (Japan); International Cluster for Bio-Resource Research, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki, Kanagawa 214-8571 (Japan); Matsunari, Hitomi [Department of Life Sciences, School of Agriculture, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki, Kanagawa 214-8571 (Japan); Takayanagi, Shuko [Japan Science and Technology Agency (JST), ERATO, Nakauchi Stem Cell and Organ Regeneration Project, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639 (Japan); Department of Life Sciences, School of Agriculture, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki, Kanagawa 214-8571 (Japan); Haruyama, Erika; Nakano, Kazuaki; Fujiwara, Tsukasa; Ikezawa, Yuka [Department of Life Sciences, School of Agriculture, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki, Kanagawa 214-8571 (Japan); Nakauchi, Hiromitsu [Japan Science and Technology Agency (JST), ERATO, Nakauchi Stem Cell and Organ Regeneration Project, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639 (Japan); Center for Stem Cell Biology and Regenerative Medicine, Institute of Medical Science, Tokyo University, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639 (Japan); and others

    2010-11-05

    Research highlights: {yields} EGFP gene integrated in porcine somatic cells could be knocked out using the ZFN-KO system. {yields} ZFNs induced targeted mutations in porcine primary cultured cells. {yields} Complete absence of EGFP fluorescence was confirmed in ZFN-treated cells. -- Abstract: Zinc-finger nucleases (ZFNs) are expected as a powerful tool for generating gene knockouts in laboratory and domestic animals. Currently, it is unclear whether this technology can be utilized for knocking-out genes in pigs. Here, we investigated whether knockout (KO) events in which ZFNs recognize and cleave a target sequence occur in porcine primary cultured somatic cells that harbor the exogenous enhanced green fluorescent protein (EGFP) gene. ZFN-encoding mRNA designed to target the EGFP gene was introduced by electroporation into the cell. Using the Surveyor nuclease assay and flow cytometric analysis, we confirmed ZFN-induced cleavage of the target sequence and the disappearance of EGFP fluorescence expression in ZFN-treated cells. In addition, sequence analysis revealed that ZFN-induced mutations such as base substitution, deletion, or insertion were generated in the ZFN cleavage site of EGFP-expression negative cells that were cloned from ZFN-treated cells, thereby showing it was possible to disrupt (i.e., knock out) the function of the EGFP gene in porcine somatic cells. To our knowledge, this study provides the first evidence that the ZFN-KO system can be applied to pigs. These findings may open a new avenue to the creation of gene KO pigs using ZFN-treated cells and somatic cell nuclear transfer.

  7. Selection-independent generation of gene knockout mouse embryonic stem cells using zinc-finger nucleases.

    Directory of Open Access Journals (Sweden)

    Anna Osiak

    Full Text Available Gene knockout in murine embryonic stem cells (ESCs has been an invaluable tool to study gene function in vitro or to generate animal models with altered phenotypes. Gene targeting using standard techniques, however, is rather inefficient and typically does not exceed frequencies of 10(-6. In consequence, the usage of complex positive/negative selection strategies to isolate targeted clones has been necessary. Here, we present a rapid single-step approach to generate a gene knockout in mouse ESCs using engineered zinc-finger nucleases (ZFNs. Upon transient expression of ZFNs, the target gene is cleaved by the designer nucleases and then repaired by non-homologous end-joining, an error-prone DNA repair process that introduces insertions/deletions at the break site and therefore leads to functional null mutations. To explore and quantify the potential of ZFNs to generate a gene knockout in pluripotent stem cells, we generated a mouse ESC line containing an X-chromosomally integrated EGFP marker gene. Applying optimized conditions, the EGFP locus was disrupted in up to 8% of ESCs after transfection of the ZFN expression vectors, thus obviating the need of selection markers to identify targeted cells, which may impede or complicate downstream applications. Both activity and ZFN-associated cytotoxicity was dependent on vector dose and the architecture of the nuclease domain. Importantly, teratoma formation assays of selected ESC clones confirmed that ZFN-treated ESCs maintained pluripotency. In conclusion, the described ZFN-based approach represents a fast strategy for generating gene knockouts in ESCs in a selection-independent fashion that should be easily transferrable to other pluripotent stem cells.

  8. Selection-independent generation of gene knockout mouse embryonic stem cells using zinc-finger nucleases.

    Science.gov (United States)

    Osiak, Anna; Radecke, Frank; Guhl, Eva; Radecke, Sarah; Dannemann, Nadine; Lütge, Fabienne; Glage, Silke; Rudolph, Cornelia; Cantz, Tobias; Schwarz, Klaus; Heilbronn, Regine; Cathomen, Toni

    2011-01-01

    Gene knockout in murine embryonic stem cells (ESCs) has been an invaluable tool to study gene function in vitro or to generate animal models with altered phenotypes. Gene targeting using standard techniques, however, is rather inefficient and typically does not exceed frequencies of 10(-6). In consequence, the usage of complex positive/negative selection strategies to isolate targeted clones has been necessary. Here, we present a rapid single-step approach to generate a gene knockout in mouse ESCs using engineered zinc-finger nucleases (ZFNs). Upon transient expression of ZFNs, the target gene is cleaved by the designer nucleases and then repaired by non-homologous end-joining, an error-prone DNA repair process that introduces insertions/deletions at the break site and therefore leads to functional null mutations. To explore and quantify the potential of ZFNs to generate a gene knockout in pluripotent stem cells, we generated a mouse ESC line containing an X-chromosomally integrated EGFP marker gene. Applying optimized conditions, the EGFP locus was disrupted in up to 8% of ESCs after transfection of the ZFN expression vectors, thus obviating the need of selection markers to identify targeted cells, which may impede or complicate downstream applications. Both activity and ZFN-associated cytotoxicity was dependent on vector dose and the architecture of the nuclease domain. Importantly, teratoma formation assays of selected ESC clones confirmed that ZFN-treated ESCs maintained pluripotency. In conclusion, the described ZFN-based approach represents a fast strategy for generating gene knockouts in ESCs in a selection-independent fashion that should be easily transferrable to other pluripotent stem cells.

  9. Hydrogen sulfide represses androgen receptor transactivation by targeting at the second zinc finger module.

    Science.gov (United States)

    Zhao, Kexin; Li, Shuangshuang; Wu, Lingyun; Lai, Christopher; Yang, Guangdong

    2014-07-25

    Androgen receptor (AR) signaling is indispensable for the development of prostate cancer from the initial androgen-dependent state to a later aggressive androgen-resistant state. This study examined the role of hydrogen sulfide (H(2)S), a novel gasotransmitter, in the regulation of AR signaling as well as its mediation in androgen-independent cell growth in prostate cancer cells. Here we found that H(2)S inhibits cell proliferation of both androgen-dependent (LNCaP) and antiandrogen-resistant prostate cancer cells (LNCaP-B), with more significance on the latter, which was established by long term treatment of parental LNCaP cells with bicalutamide. The expression of cystathionine γ-lyase (CSE), a major H(2)S producing enzyme in prostate tissue, was reduced in both human prostate cancer tissues and LNCaP-B cells. LNCaP-B cells were resistant to bicalutamide-induced cell growth inhibition, and CSE overexpression could rebuild the sensitivity of LNCaP-B cells to bicalutamide. H(2)S significantly repressed the expression of prostate-specific antigen (PSA) and TMPRSS2, two AR-targeted genes. In addition, H(2)S inhibited AR binding with PSA promoter and androgen-responsive element (ARE) luciferase activity. We further found that AR is post-translationally modified by H(2)S through S-sulfhydration. Mutation of cysteine 611 and cysteine 614 in the second zinc finger module of AR-DNA binding domain diminished the effects of H(2)S on AR S-sulfhydration and AR dimerization. These data suggest that reduced CSE/H2S signaling contributes to antiandrogen-resistant status, and sufficient level of H(2)S is able to inhibit AR transactivation and treat castration-resistant prostate cancer.

  10. Stability of zinc finger nuclease protein is enhanced by the proteasome inhibitor MG132.

    Directory of Open Access Journals (Sweden)

    Suresh Ramakrishna

    Full Text Available BACKGROUND: Zinc finger nucleases (ZFNs are powerful tools for gene therapy and genetic engineering. The characterization of ZFN protein stability and the development of simple methods to improve ZFN function would facilitate the application of this promising technology. However, the factors that affect ZFN protein stability and function are not yet clear. Here, we determined the stability and half-life of two ZFN proteins and examined the effect of MG132 (carbobenzoxyl-leucinyl-leucinyl-leucinal-Hl, a proteasome inhibitor, on ZFN-mediated gene modifications. METHODOLOGY/PRINCIPAL FINDINGS: ZFN proteins were expressed in 293T cells after transfection of ZFN-encoding plasmids. We studied two ZFN pairs: Z-224, which targets the CCR5 gene, and K-230, which targets a region 230 kbp upstream of CCR5. Western blotting after treatment with cycloheximide showed that the half-life of these ZFN proteins was around two hours. An immunoprecipitation assay revealed that the ZFN interacts with ubiquitin molecules and undergoes polyubiquitination in vivo. Western blotting showed that the addition of MG132, a proteasomal inhibitor, increased ZFN protein levels. Finally, a surrogate reporter assay and a T7E1 assay revealed that MG132 treatment enhanced ZFN-directed gene editing. CONCLUSIONS: To our knowledge, this is the first study to investigate ZFN protein stability and to show that a small molecule can increase ZFN activity. Our protein stability study should lay the foundation for further improvement of ZFN technology; as a first step, the use of the small molecule MG132 can enhance the efficiency of ZFN-mediated gene editing.

  11. Molecular cloning and characterization of a gene encoding RING zinc finger ankyrin protein from drought-tolerant Artemisia desertorum

    Indian Academy of Sciences (India)

    Xiuhong Yang; Chao Sun; Yuanlei Hun; Zhongping Lin

    2008-03-01

    A RING zinc finger ankyrin protein gene, designated AdZFP1, was isolated from drought-tolerant Artemisia desertorum Spreng by mRNA differential display and RACE. Its cDNA was 1723 bp and encoded a putative protein of 445 amino acids with a predicted molecular mass of 47.9 kDa and an isoelectric point (pI) of 7.49. A typical C3HC4-type RING finger domain was found at the C-terminal region of the AdZFP1 protein, and several groups of ankyrin repeats were found at the N-terminal region. Alignments of amino acid sequence showed that AdZFP1 was 66% identical to the Arabidopsis thaliana putative RING zinc finger ankyrin protein AAN31869. Transcriptional analysis showed that AdZFP1 was inducible under drought stress in root, stem and leaf of the plant. Semi-quantitative reverse-transcriptase-polymerase chain reaction (RT-PCR) analysis showed that the transcript of AdZFP1 was strongly induced by exogenous abscisic acid (ABA) and also by salinity, cold and heat to some extent. Overexpression of the AdZFP1 gene in transgenic tobacco enhanced their tolerance to drought stress.

  12. Expression and purification of a novel ZNF191 zinc finger protein——ZNF191 protein and its truncated zinc finger region ZNF191 (243—368)

    Institute of Scientific and Technical Information of China (English)

    刘玉奇; 余龙; 余文浩; 施少林; 孙炳耘; 吴国俊; 黄仲贤

    1999-01-01

    ZNF191 is a new zine finger gene whieh has a 1107 bp open reading frame (ORF) and eneodes a 368 amino avid protein including a putative DNA-binding domain of four Cys2 His2 zine finger motifs at its C-terminal region. The ZNF191 cDNA is loeated in the chromosome 18q12.1 region where it is known that a variety of hereditary diseases are related to. Probably, this protein has potential function of stimulating the gene franscription. In order to examine the function and structure of ZNF191 protein, the ORF of ZNF191 and its zine finger region ZNF191(243--368) genes were inserted into PTSA-18 expression vector by PCR amplification, then the constructed genes were expressed in the PTSA-18/B121 (DE3) system. The two proteins were purified by DEAE-52, CM-23 and Heparin-Sepharose 4B columns. The pooled proteins showed a single band as assayed by Coomasie Brillian Blue Staining of an SDS/polyacryamide gel.

  13. Transcriptome wide identification, phylogenetic analysis, and expression profiling of zinc-finger transcription factors from Crocus sativus L.

    Science.gov (United States)

    Malik, Aubid Hussain; Ashraf, Nasheeman

    2017-02-28

    Crocus sativus belongs to Iridaceae family and is the only plant species which produces apocarotenoids like crocin, picrocrocin, and safranal in significant quantities. Besides their organoleptic properties, Crocus apocarotenoids have been found to possess remarkable pharmacological potential. Although apocarotenoid biosynthetic pathway has been worked out to a great degree, but the mechanism that regulates the tissue and developmental stage-specific production of Crocus apocarotenoids is not known. To identify the genes regulating apocarotenoid biosynthesis in Crocus, transcriptome wide identification of zinc-finger transcription factors was undertaken. 81 zinc-finger transcription factors were identified which grouped into eight subfamilies. C2H2, C3H, and AN20/AN1 were the major subfamilies with 29, 20, and 14 members, respectively. Expression profiling revealed CsSAP09 as a potential candidate for regulation of apocarotenoid biosynthesis. CsSAP09 was found to be highly expressed in stigma at anthesis stage corroborating with the accumulation pattern of apocarotenoids. CsSAP09 was nuclear localized and activated reporter gene transcription in yeast. It was highly induced in response to oxidative, salt and dehydration stresses, ABA and methyl jasmonate. Furthermore, upstream region of CsSAP09 was found to contain stress and light responsive elements. To our knowledge, this is the first report on the study of a gene family in C. sativus and may provide basic insights into the putative role of zinc finger genes. It may also serve as a valuable resource for functional characterization of these genes aimed towards unraveling their role in regulation of apocarotenoid biosynthesis.

  14. Direct binding of specific AUF1 isoforms to tandem zinc finger domains of tristetraprolin (TTP) family proteins.

    Science.gov (United States)

    Kedar, Vishram P; Zucconi, Beth E; Wilson, Gerald M; Blackshear, Perry J

    2012-02-17

    Tristetraprolin (TTP) is the prototype of a family of CCCH tandem zinc finger proteins that can bind to AU-rich elements in mRNAs and promote their decay. TTP binds to mRNA through its central tandem zinc finger domain; it then promotes mRNA deadenylation, considered to be the rate-limiting step in eukaryotic mRNA decay. We found that TTP and its related family members could bind to certain isoforms of another AU-rich element-binding protein, HNRNPD/AUF1, as well as a related protein, laAUF1. The interaction domain within AUF1p45 appeared to be a C-terminal "GY" region, and the interaction domain within TTP was the tandem zinc finger domain. Surprisingly, binding of AUF1p45 to TTP occurred even with TTP mutants that lacked RNA binding activity. In cell extracts, binding of AUF1p45 to TTP potentiated TTP binding to ARE-containing RNA probes, as determined by RNA gel shift assays; AUF1p45 did not bind to the RNA probes under these conditions. Using purified, recombinant proteins and a synthetic RNA target in FRET assays, we demonstrated that AUF1p45, but not AUF1p37, increased TTP binding affinity for RNA ∼5-fold. These data suggest that certain isoforms of AUF1 can serve as "co-activators" of TTP family protein binding to RNA. The results raise interesting questions about the ability of AUF1 isoforms to regulate the mRNA binding and decay-promoting activities of TTP and its family members as well as the ability of AUF1 proteins to serve as possible physical links between TTP and other mRNA decay proteins and structures.

  15. Molecular cloning and characterization of a novel RING zinc-finger protein gene up-regulated under in vitro salt stress in cassava.

    Science.gov (United States)

    dos Reis, Sávio Pinho; Tavares, Liliane de Souza Conceição; Costa, Carinne de Nazaré Monteiro; Brígida, Aílton Borges Santa; de Souza, Cláudia Regina Batista

    2012-06-01

    Cassava (Manihot esculenta Crantz) is one of the world's most important food crops. It is cultivated mainly in developing countries of tropics, since its root is a major source of calories for low-income people due to its high productivity and resistance to many abiotic and biotic factors. A previous study has identified a partial cDNA sequence coding for a putative RING zinc finger in cassava storage root. The RING zinc finger protein is a specialized type of zinc finger protein found in many organisms. Here, we isolated the full-length cDNA sequence coding for M. esculenta RZF (MeRZF) protein by a combination of 5' and 3' RACE assays. BLAST analysis showed that its deduced amino acid sequence has a high level of similarity to plant proteins of RZF family. MeRZF protein contains a signature sequence motif for a RING zinc finger at its C-terminal region. In addition, this protein showed a histidine residue at the fifth coordination site, likely belonging to the RING-H2 subgroup, as confirmed by our phylogenetic analysis. There is also a transmembrane domain in its N-terminal region. Finally, semi-quantitative RT-PCR assays showed that MeRZF expression is increased in detached leaves treated with sodium chloride. Here, we report the first evidence of a RING zinc finger gene of cassava showing potential role in response to salt stress.

  16. The maize INDETERMINATE1 flowering time regulator defines a highly conserved zinc finger protein family in higher plants

    Directory of Open Access Journals (Sweden)

    Colasanti Joseph

    2006-06-01

    Full Text Available Abstract Background The maize INDETERMINATE1 gene, ID1, is a key regulator of the transition to flowering and the founding member of a transcription factor gene family that encodes a protein with a distinct arrangement of zinc finger motifs. The zinc fingers and surrounding sequence make up the signature ID domain (IDD, which appears to be found in all higher plant genomes. The presence of zinc finger domains and previous biochemical studies showing that ID1 binds to DNA suggests that members of this gene family are involved in transcriptional regulation. Results Comparison of IDD genes identified in Arabidopsis and rice genomes, and all IDD genes discovered in maize EST and genomic databases, suggest that ID1 is a unique member of this gene family. High levels of sequence similarity amongst all IDD genes from maize, rice and Arabidopsis suggest that they are derived from a common ancestor. Several unique features of ID1 suggest that it is a divergent member of the maize IDD family. Although no clear ID1 ortholog was identified in the Arabidopsis genome, highly similar genes that encode proteins with identity extending beyond the ID domain were isolated from rice and sorghum. Phylogenetic comparisons show that these putative orthologs, along with maize ID1, form a group separate from other IDD genes. In contrast to ID1 mRNA, which is detected exclusively in immature leaves, several maize IDD genes showed a broad range of expression in various tissues. Further, Western analysis with an antibody that cross-reacts with ID1 protein and potential orthologs from rice and sorghum shows that all three proteins are detected in immature leaves only. Conclusion Comparative genomic analysis shows that the IDD zinc finger family is highly conserved among both monocots and dicots. The leaf-specific ID1 expression pattern distinguishes it from other maize IDD genes examined. A similar leaf-specific localization pattern was observed for the putative ID1 protein

  17. P16-specific DNA methylation by engineered zinc finger methyltransferase inactivates gene transcription and promotes cancer metastasis

    OpenAIRE

    Cui, Chenghua; Gan, Ying; Gu, Liankun; Wilson, James; Liu, Zhaojun; Zhang, Baozhen; Deng, Dajun

    2015-01-01

    Background P16 DNA methylation is well known to be the most frequent event in cancer development. It has been reported that genetic inactivation of P16 drives cancer growth and metastasis, however, whether P16 DNA methylation is truly a driver in cancer metastasis remains unknown. Results A P16-specific DNA methyltransferase (P16-dnmt) expression vector is designed using a P16 promoter-specific engineered zinc finger protein fused with the catalytic domain of dnmt3a. P16-dnmt transfection sig...

  18. Identification of off-target cleavage sites of zinc finger nucleases and TAL effector nucleases using predictive models.

    Science.gov (United States)

    Fine, Eli J; Cradick, Thomas J; Bao, Gang

    2014-01-01

    Using engineered nucleases, such as Zinc Finger Nucleases (ZFNs) or Transcription Activator-Like Effector Nucleases (TALENs), to make targeted genomic modifications has become a common technique to create new model organisms and custom cell lines, and has shown great promise for disease treatment. However, these nucleases have the potential for off-target cleavage that could confound interpretation of experimental results and be detrimental for therapeutic use. Here, we describe a method to test for nuclease cleavage at potential off-target sites predicted by bioinformatics models.

  19. Chemical shift assignments of zinc finger domain of methionine aminopeptidase 1 (MetAP1) from Homo sapiens.

    Science.gov (United States)

    Rachineni, Kavitha; Arya, Tarun; Singarapu, Kiran Kumar; Addlagatta, Anthony; Bharatam, Jagadeesh

    2015-10-01

    Methionine aminopeptidase Type I (MetAP1) cleaves the initiator methionine from about 70 % of all newly synthesized proteins in almost every living cell. Human MetAP1 is a two domain protein with a zinc finger on the N-terminus and a catalytic domain on the C-terminus. Here, we report the chemical shift assignments of the amino terminal zinc binding domain (ZBD) (1-83 residues) of the human MetAP1 derived by using advanced NMR spectroscopic methods. We were able to assign the chemical shifts of ZBD of MetAP1 nearly complete, which reveal two helical fragments involving residues P44-L49 (α1) and Q59-K82 (α2). The protein structure unfolds upon complex formation with the addition of 2 M excess EDTA, indicated by the appearance of amide resonances in the random coil chemical shift region of (15)NHSQC spectrum.

  20. BTB-Zinc Finger Oncogenes Are Required for Ras and Notch-Driven Tumorigenesis in Drosophila.

    Directory of Open Access Journals (Sweden)

    Karen Doggett

    Full Text Available During tumorigenesis, pathways that promote the epithelial-to-mesenchymal transition (EMT can both facilitate metastasis and endow tumor cells with cancer stem cell properties. To gain a greater understanding of how these properties are interlinked in cancers we used Drosophila epithelial tumor models, which are driven by orthologues of human oncogenes (activated alleles of Ras and Notch in cooperation with the loss of the cell polarity regulator, scribbled (scrib. Within these tumors, both invasive, mesenchymal-like cell morphology and continual tumor overgrowth, are dependent upon Jun N-terminal kinase (JNK activity. To identify JNK-dependent changes within the tumors we used a comparative microarray analysis to define a JNK gene signature common to both Ras and Notch-driven tumors. Amongst the JNK-dependent changes was a significant enrichment for BTB-Zinc Finger (ZF domain genes, including chronologically inappropriate morphogenesis (chinmo. chinmo was upregulated by JNK within the tumors, and overexpression of chinmo with either RasV12 or Nintra was sufficient to promote JNK-independent epithelial tumor formation in the eye/antennal disc, and, in cooperation with RasV12, promote tumor formation in the adult midgut epithelium. Chinmo primes cells for oncogene-mediated transformation through blocking differentiation in the eye disc, and promoting an escargot-expressing stem or enteroblast cell state in the adult midgut. BTB-ZF genes are also required for Ras and Notch-driven overgrowth of scrib mutant tissue, since, although loss of chinmo alone did not significantly impede tumor development, when loss of chinmo was combined with loss of a functionally related BTB-ZF gene, abrupt, tumor overgrowth was significantly reduced. abrupt is not a JNK-induced gene, however, Abrupt is present in JNK-positive tumor cells, consistent with a JNK-associated oncogenic role. As some mammalian BTB-ZF proteins are also highly oncogenic, our work suggests that

  1. Identification and characterization of a salt stress-inducible zinc finger protein from Festuca arundinacea

    Directory of Open Access Journals (Sweden)

    Martin Ruth C

    2012-01-01

    Full Text Available Abstract Background Increased biotic and abiotic plant stresses due to climate change together with an expected global human population of over 9 billion by 2050 intensifies the demand for agricultural production on marginal lands. Soil salinity is one of the major abiotic stresses responsible for reduced crop productivity worldwide and the salinization of arable land has dramatically increased over the last few decades. Consequently, as land becomes less amenable for conventional agriculture, plants grown on marginal soils will be exposed to higher levels of soil salinity. Forage grasses are a critical component of feed used in livestock production worldwide, with many of these same species of grasses being utilized for lawns, erosion prevention, and recreation. Consequently, it is important to develop a better understanding of salt tolerance in forage and related grass species. Findings A gene encoding a ZnF protein was identified during the analysis of a salt-stress suppression subtractive hybridization (SSH expression library from the forage grass species Festuca arundinacea. The expression pattern of FaZnF was compared to that of the well characterized gene for delta 1-pyrroline-5-carboxylate synthetase (P5CS, a key enzyme in proline biosynthesis, which was also identified in the salt-stress SSH library. The FaZnF and P5CS genes were both up-regulated in response to salt and drought stresses suggesting a role in dehydration stress. FaZnF was also up-regulated in response to heat and wounding, suggesting that it might have a more general function in multiple abiotic stress responses. Additionally, potential downstream targets of FaZnF (a MAPK [Mitogen-Activated Protein Kinase], GST [Glutathione-S-Transferase] and lipoxygenase L2 were found to be up-regulated in calli overexpressing FaZnF when compared to control cell lines. Conclusions This work provides evidence that FaZnF is an AN1/A20 zinc finger protein that is involved in the regulation

  2. The DnaJ-like zinc finger domain protein PSA2 affects light acclimation and chloroplast development in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Yan-Wen eWang

    2016-03-01

    Full Text Available The biosynthesis of chlorophylls and carotenoids and the assembly of thylakoid membranes are critical for the photoautotrophic growth of plants. Different factors are involved in these two processes. In recent years, members of the DnaJ-like zinc finger domain proteins have been found to take part in the biogenesis and/or the maintenance of plastids. One member of this family of proteins, PSA2, was recently found to localize to the thylakoid lumen and regulate the accumulation of photosystem I. In this study, we report that the silencing of PSA2 in Arabidopsis thaliana resulted in variegated leaves and retarded growth. Although both chlorophylls and total carotenoids decreased in the psa2 mutant, violaxanthin and zeaxanthin accumulated in the mutant seedlings grown under growth condition. Lower levels of non-photochemical quenching and electron transport rate were also found in the psa2 mutant seedlings under growth condition compared with those of the wild-type plants, indicating an impaired capability to acclimate to normal light irradiance when PSA2 was silenced. Moreover, we also observed an abnormal assembly of grana thylakoids and poorly developed stroma thylakoids in psa2 chloroplasts. Taken together, our results demonstrate that PSA2 is a member of the DnaJ-like zinc finger domain protein family that affects light acclimation and chloroplast development.

  3. A novel genetic system based on zinc finger nucleases for the identification of interactions between proteins in vivo.

    Directory of Open Access Journals (Sweden)

    Ling Wang

    Full Text Available Yeast two-hybrid (Y2H methods are powerful tools for detecting protein-protein interactions. The traditional Y2H method has been widely applied to screen novel protein interactions since it was established two decades ago. The high false-positive rate of the traditional method drove the development of modified Y2H systems. Here, we describe a novel Y2H system using zinc-finger nucleases (ZFNs. ZFNs contain two functional domains, a zinc-finger DNA-binding domain (ZFP and a non-specific nuclease domain (FokI. In this system, the bait is expressed as a fusion protein with a specific ZFP, and the prey is fused to the FokI. A reporter vector is designed such that the ZFN target site disrupts the Gal4 open reading frame. By transforming the three plasmids into a yeast strain (AH109, the interaction between the bait and prey proteins reconstitutes ZFN function and generates the double-strand break (DSB on its target site. The DNA DSB repair restores Gal4 function, which activates the expression of the four reporter genes. We used p53-SV40LT interacting proteins to prove the concept. In addition, 80% positive rate was observed in a cDNA screening test against WDSV orfA protein. Our results strongly suggested that this Y2H system could increase screening reliability and reproducibility, and provide a novel approach for interactomics research.

  4. Automatic polymerase chain reaction product detection system for food safety monitoring using zinc finger protein fused to luciferase.

    Science.gov (United States)

    Yoshida, Wataru; Kezuka, Aki; Murakami, Yoshiyuki; Lee, Jinhee; Abe, Koichi; Motoki, Hiroaki; Matsuo, Takafumi; Shimura, Nobuaki; Noda, Mamoru; Igimi, Shizunobu; Ikebukuro, Kazunori

    2013-11-01

    An automatic polymerase chain reaction (PCR) product detection system for food safety monitoring using zinc finger (ZF) protein fused to luciferase was developed. ZF protein fused to luciferase specifically binds to target double stranded DNA sequence and has luciferase enzymatic activity. Therefore, PCR products that comprise ZF protein recognition sequence can be detected by measuring the luciferase activity of the fusion protein. We previously reported that PCR products from Legionella pneumophila and Escherichia coli (E. coli) O157 genomic DNA were detected by Zif268, a natural ZF protein, fused to luciferase. In this study, Zif268-luciferase was applied to detect the presence of Salmonella and coliforms. Moreover, an artificial zinc finger protein (B2) fused to luciferase was constructed for a Norovirus detection system. In the luciferase activity detection assay, several bound/free separation process is required. Therefore, an analyzer that automatically performed the bound/free separation process was developed to detect PCR products using the ZF-luciferase fusion protein. By means of the automatic analyzer with ZF-luciferase fusion protein, target pathogenic genomes were specifically detected in the presence of other pathogenic genomes. Moreover, we succeeded in the detection of 10 copies of E. coli BL21 without extraction of genomic DNA by the automatic analyzer and E. coli was detected with a logarithmic dependency in the range of 1.0×10 to 1.0×10(6) copies.

  5. Engineering HIV-resistant human CD4+ T cells with CXCR4-specific zinc-finger nucleases.

    Directory of Open Access Journals (Sweden)

    Craig B Wilen

    2011-04-01

    Full Text Available HIV-1 entry requires the cell surface expression of CD4 and either the CCR5 or CXCR4 coreceptors on host cells. Individuals homozygous for the ccr5Δ32 polymorphism do not express CCR5 and are protected from infection by CCR5-tropic (R5 virus strains. As an approach to inactivating CCR5, we introduced CCR5-specific zinc-finger nucleases into human CD4+ T cells prior to adoptive transfer, but the need to protect cells from virus strains that use CXCR4 (X4 in place of or in addition to CCR5 (R5X4 remains. Here we describe engineering a pair of zinc finger nucleases that, when introduced into human T cells, efficiently disrupt cxcr4 by cleavage and error-prone non-homologous DNA end-joining. The resulting cells proliferated normally and were resistant to infection by X4-tropic HIV-1 strains. CXCR4 could also be inactivated in ccr5Δ32 CD4+ T cells, and we show that such cells were resistant to all strains of HIV-1 tested. Loss of CXCR4 also provided protection from X4 HIV-1 in a humanized mouse model, though this protection was lost over time due to the emergence of R5-tropic viral mutants. These data suggest that CXCR4-specific ZFNs may prove useful in establishing resistance to CXCR4-tropic HIV for autologous transplant in HIV-infected individuals.

  6. ExpandplusCrystal Structures of Poly(ADP-ribose) Polymerase-1 (PARP-1) Zinc Fingers Bound to DNA

    Energy Technology Data Exchange (ETDEWEB)

    M Langelier; J Planck; S Roy; J Pascal

    2011-12-31

    Poly(ADP-ribose) polymerase-1 (PARP-1) has two homologous zinc finger domains, Zn1 and Zn2, that bind to a variety of DNA structures to stimulate poly(ADP-ribose) synthesis activity and to mediate PARP-1 interaction with chromatin. The structural basis for interaction with DNA is unknown, which limits our understanding of PARP-1 regulation and involvement in DNA repair and transcription. Here, we have determined crystal structures for the individual Zn1 and Zn2 domains in complex with a DNA double strand break, providing the first views of PARP-1 zinc fingers bound to DNA. The Zn1-DNA and Zn2-DNA structures establish a novel, bipartite mode of sequence-independent DNA interaction that engages a continuous region of the phosphodiester backbone and the hydrophobic faces of exposed nucleotide bases. Biochemical and cell biological analysis indicate that the Zn1 and Zn2 domains perform distinct functions. The Zn2 domain exhibits high binding affinity to DNA compared with the Zn1 domain. However, the Zn1 domain is essential for DNA-dependent PARP-1 activity in vitro and in vivo, whereas the Zn2 domain is not strictly required. Structural differences between the Zn1-DNA and Zn2-DNA complexes, combined with mutational and structural analysis, indicate that a specialized region of the Zn1 domain is re-configured through the hydrophobic interaction with exposed nucleotide bases to initiate PARP-1 activation.

  7. Zinc-finger directed double-strand breaks within CAG repeat tracts promote repeat instability in human cells.

    Science.gov (United States)

    Mittelman, David; Moye, Christopher; Morton, Jason; Sykoudis, Kristen; Lin, Yunfu; Carroll, Dana; Wilson, John H

    2009-06-16

    Expanded triplet repeats have been identified as the genetic basis for a growing number of neurological and skeletal disorders. To examine the contribution of double-strand break repair to CAG x CTG repeat instability in mammalian systems, we developed zinc finger nucleases (ZFNs) that recognize and cleave CAG repeat sequences. Engineered ZFNs use a tandem array of zinc fingers, fused to the FokI DNA cleavage domain, to direct double-strand breaks (DSBs) in a site-specific manner. We first determined that the ZFNs cleave CAG repeats in vitro. Then, using our previously described tissue culture assay for identifying modifiers of CAG repeat instability, we found that transfection of ZFN-expression vectors induced up to a 15-fold increase in changes to the CAG repeat in human and rodent cell lines, and that longer repeats were much more sensitive to cleavage than shorter ones. Analysis of individual colonies arising after treatment revealed a spectrum of events consistent with ZFN-induced DSBs and dominated by repeat contractions. We also found that expressing a dominant-negative form of RAD51 in combination with a ZFN, dramatically reduced the effect of the nuclease, suggesting that DSB-induced repeat instability is mediated, in part, through homology directed repair. These studies identify a ZFN as a useful reagent for characterizing the effects of DSBs on CAG repeats in cells.

  8. Molecular phylogeny of OVOL genes illustrates a conserved C2H2 zinc finger domain coupled by hypervariable unstructured regions.

    Directory of Open Access Journals (Sweden)

    Abhishek Kumar

    Full Text Available OVO-like proteins (OVOL are members of the zinc finger protein family and serve as transcription factors to regulate gene expression in various differentiation processes. Recent studies have shown that OVOL genes are involved in epithelial development and differentiation in a wide variety of organisms; yet there is a lack of comprehensive studies that describe OVOL proteins from an evolutionary perspective. Using comparative genomic analysis, we traced three different OVOL genes (OVOL1-3 in vertebrates. One gene, OVOL3, was duplicated during a whole-genome-duplication event in fish, but only the copy (OVOL3b was retained. From early-branching metazoa to humans, we found that a core domain, comprising a tetrad of C2H2 zinc fingers, is conserved. By domain comparison of the OVOL proteins, we found that they evolved in different metazoan lineages by attaching intrinsically-disordered (ID segments of N/C-terminal extensions of 100 to 1000 amino acids to this conserved core. These ID regions originated independently across different animal lineages giving rise to different types of OVOL genes over the course of metazoan evolution. We illustrated the molecular evolution of metazoan OVOL genes over a period of 700 million years (MY. This study both extends our current understanding of the structure/function relationship of metazoan OVOL genes, and assembles a good platform for further characterization of OVOL genes from diverged organisms.

  9. The conserved Bud20 zinc finger protein is a new component of the ribosomal 60S subunit export machinery.

    Science.gov (United States)

    Bassler, Jochen; Klein, Isabella; Schmidt, Claudia; Kallas, Martina; Thomson, Emma; Wagner, Maria Anna; Bradatsch, Bettina; Rechberger, Gerald; Strohmaier, Heimo; Hurt, Ed; Bergler, Helmut

    2012-12-01

    The nuclear export of the preribosomal 60S (pre-60S) subunit is coordinated with late steps in ribosome assembly. Here, we show that Bud20, a conserved C(2)H(2)-type zinc finger protein, is an unrecognized shuttling factor required for the efficient export of pre-60S subunits. Bud20 associates with late pre-60S particles in the nucleoplasm and accompanies them into the cytoplasm, where it is released through the action of the Drg1 AAA-ATPase. Cytoplasmic Bud20 is then reimported via a Kap123-dependent pathway. The deletion of Bud20 induces a strong pre-60S export defect and causes synthetic lethality when combined with mutant alleles of known pre-60S subunit export factors. The function of Bud20 in ribosome export depends on a short conserved N-terminal sequence, as we observed that mutations or the deletion of this motif impaired 60S subunit export and generated the genetic link to other pre-60S export factors. We suggest that the shuttling Bud20 is recruited to the nascent 60S subunit via its central zinc finger rRNA binding domain to facilitate the subsequent nuclear export of the preribosome employing its N-terminal extension.

  10. Promiscuous and specific phospholipid binding by domains in ZAC, a membrane-associated Arabidopsis protein with an ARF GAP zinc finger and a C2 domain

    DEFF Research Database (Denmark)

    Jensen, R B; Lykke-Andersen, K; Frandsen, G I

    2000-01-01

    containing the ZAC-C2 domain bind anionic phospholipids non-specifically, with some variance in Ca2+ and salt dependence. Similar assays demonstrated specific affinity of the ZAC N-terminal region (residues 1-174) for phosphatidylinositol 3-monophosphate (PI-3-P). Binding was dependent in part on an intact...... zinc finger motif, but proteins containing only the zinc finger domain (residues 1-105) did not bind PI-3-P. Recombinant ZAC possessed GTPase-activating activity on Arabidopsis ARF proteins. These data identify a novel PI-3-P-binding protein region and thereby provide evidence...

  11. Tailor-made zinc-finger transcription factors activate FLO11 gene expression with phenotypic consequences in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Shieh, Jia-Ching; Cheng, Yu-Che; Su, Mao-Chang; Moore, Michael; Choo, Yen; Klug, Aaron

    2007-08-15

    Cys2His2 zinc fingers are eukaryotic DNA-binding motifs, capable of distinguishing different DNA sequences, and are suitable for engineering artificial transcription factors. In this work, we used the budding yeast Saccharomyces cerevisiae to study the ability of tailor-made zinc finger proteins to activate the expression of the FLO11 gene, with phenotypic consequences. Two three-finger peptides were identified, recognizing sites from the 5' UTR of the FLO11 gene with nanomolar DNA-binding affinity. The three-finger domains and their combined six-finger motif, recognizing an 18-bp site, were fused to the activation domain of VP16 or VP64. These transcription factor constructs retained their DNA-binding ability, with the six-finger ones being the highest in affinity. However, when expressed in haploid yeast cells, only one three-finger recombinant transcription factor was able to activate the expression of FLO11 efficiently. Unlike in the wild-type, cells with such transcriptional activation displayed invasive growth and biofilm formation, without any requirement for glucose depletion. The VP16 and VP64 domains appeared to act equally well in the activation of FLO11 expression, with comparable effects in phenotypic alteration. We conclude that the functional activity of tailor-made transcription factors in cells is not easily predicted by the in vitro DNA-binding activity.

  12. Structural and dynamical characterization of the Miz-1 zinc fingers 5-8 by solution-state NMR

    Energy Technology Data Exchange (ETDEWEB)

    Bernard, David; Bedard, Mikaeel; Bilodeau, Josee; Lavigne, Pierre, E-mail: pierre.lavigne@usherbrooke.ca [Universite de Sherbrooke, Departement de Biochimie, Faculte de Medecine et des Sciences de la Sante, Institut de Pharmacologie de Sherbrooke (Canada)

    2013-10-15

    Myc-interacting zinc finger protein-1 (Miz-1) is a BTB/POZ transcription factor that activates the transcription of cytostatic genes, such as p15{sup INK4B} or p21{sup CIP1}. The C-terminus of Miz-1 contains 13 consensus C{sub 2}H{sub 2} zinc finger domains (ZF). ZFs 1-4 have been shown to interact with SMAD3/4, while the remaining ZFs are expected to bind the promoters of target genes. We have noted unusual features in ZF 5 and the linker between ZFs 5 and 6. Indeed, a glutamate is found instead of the conserved basic residue two positions before the second zinc-coordinating histidine on the ZF 5 helix, and the linker sequence is DTDKE in place of the classical TGEKP sequence. In a canonical {beta}{beta}{alpha} fold, such unusual primary structure elements should cause severe electrostatic repulsions. In this context, we have characterized the structure and the dynamics of a Miz-1 construct comprising ZFs 5-8 (Miz 5-8) by solution-state NMR. Whilst ZFs 5, 7 and 8 were shown to adopt the classical {beta}{beta}{alpha} fold for C{sub 2}H{sub 2} ZFs, the number of long-range NOEs was insufficient to define a classical fold for ZF 6. We show by using {sup 15}N-relaxation dispersion experiments that this lack of NOEs is due to the presence of extensive motions on the {mu}s-ms timescale. Since this negatively charged region would have to be located near the phosphodiester backbone in a DNA complex, we propose that in addition to promoting conformational searches, it could serve as a hinge region to keep ZFs 1-4 away from DNA.

  13. Concise review: putting a finger on stem cell biology: zinc finger nuclease-driven targeted genetic editing in human pluripotent stem cells.

    Science.gov (United States)

    Collin, Joseph; Lako, Majlinda

    2011-07-01

    Human pluripotent stem cells (hPSCs) encompassing human embryonic stem cells and human induced pluripotent stem cells (hiPSCs) have a wide appeal for numerous basic biology studies and for therapeutic applications because of their potential to give rise to almost any cell type in the human body and immense ability to self-renew. Much attention in the stem cell field is focused toward the study of gene-based anomalies relating to the causative affects of human disease and their correction with the potential for patient-specific therapies using gene corrected hiPSCs. Therefore, the genetic manipulation of stem cells is clearly important for the development of future medicine. Although successful targeted genetic engineering in hPSCs has been reported, these cases are surprisingly few because of inherent technical limitations with the methods used. The development of more robust and efficient means by which to achieve specific genomic modifications in hPSCs has far reaching implications for stem cell research and its applications. Recent proof-of-principle reports have shown that genetic alterations with minimal toxicity are now possible through the use of zinc finger nucleases (ZFNs) and the inherent DNA repair mechanisms within the cell. In light of recent comprehensive reviews that highlight the applications, methodologies, and prospects of ZFNs, this article focuses on the application of ZFNs to stem cell biology, discussing the published work to date, potential problems, and future uses for this technology both experimentally and therapeutically.

  14. Identification of genes encoding zinc finger motifs in the cardiovascular system.

    Science.gov (United States)

    Wang, R; Hwang, D M; Cukerman, E; Liew, C C

    1997-01-01

    The Zn2+-finger DNA-binding domain has been identified in several developmental control proteins, transcription factors and gene products associated with diseases, as well as in several RNA-binding proteins. We applied library screening, expressed sequence tagging (EST sequencing), Zn2+-binding assays and Northern blot hybridization, in order to characterize novel cDNA clones of the human cardiovascular system which contain Zn2+-finger motifs. An embryonic (8-10 weeks gestation) heart lambda ZAP Express cDNA library was screened with an oligonucleotide probe deduced from a consensus amino acid sequence which is highly conserved for Zn2+-finger proteins, and approximately 350 positive clones were isolated from 1 x 10(4) plaque-forming units (pfu) initially plated. The isolated clones were classified as known and novel following single pass automated DNA sequencing. Analysis of Northern blot hybridization delineated the tissue specificity of these clones, as well as their association with cardiac growth and development. Existence of Zn2+-finger motifs in the novel clones was confirmed by Zn2+-binding assay. In this report, we present the characterization of eight novel clones, including the complete cDNA sequences of one of these clones (HHZ-123).

  15. Zinc finger artificial transcription factor-based nearest inactive analogue/nearest active analogue strategy used for the identification of plant genes controlling homologous recombination

    NARCIS (Netherlands)

    Jia, Qi; van Verk, Marcel C.; Pinas, Johan E.; Lindhout, Beatrice I.; Hooykaas, Paul J.J.; Van der Zaal, Bert J.

    2013-01-01

    In previous work, we selected a particular transcription factor, designated VP16-HRU, from a pool of zinc finger artificial transcription factors (ZF-ATFs) used for genome interrogation. When expressed in Arabidopsis thaliana under control of the ribosomal protein S5A promoter, the RPS5A::VP16-HRU c

  16. The Role of Cdkn1A-Interacting Zinc Finger Protein 1 (CIZ1 in DNA Replication and Pathophysiology

    Directory of Open Access Journals (Sweden)

    Qiang Liu

    2016-02-01

    Full Text Available Cdkn1A-interacting zinc finger protein 1 (CIZ1 was first identified in a yeast-2-hybrid system searching for interacting proteins of CDK2 inhibitor p21Cip1/Waf1. Ciz1 also binds to CDK2, cyclin A, cyclin E, CDC6, PCNA, TCF4 and estrogen receptor-α. Recent studies reveal numerous biological functions of CIZ1 in DNA replication, cell proliferation, and differentiation. In addition, splicing variants of CIZ1 mRNA is associated with a variety of cancers and Alzheimer’s disease, and mutations of the CIZ1 gene lead to cervical dystonia. CIZ1 expression is increased in cancers and rheumatoid arthritis. In this review, we will summarize the biological functions and molecular mechanisms of CIZ1 in these physiological and pathological processes.

  17. DNA damage-inducible SUMOylation of HERC2 promotes RNF8 binding via a novel SUMO-binding Zinc finger

    DEFF Research Database (Denmark)

    Danielsen, Jannie Michaela Rendtlew; Povlsen, Lou Klitgaard; Villumsen, Bine Hare;

    2012-01-01

    Nonproteolytic ubiquitylation of chromatin surrounding deoxyribonucleic acid (DNA) double-strand breaks (DSBs) by the RNF8/RNF168/HERC2 ubiquitin ligases facilitates restoration of genome integrity by licensing chromatin to concentrate genome caretaker proteins near the lesions. In parallel......, SUMOylation of so-far elusive upstream DSB regulators is also required for execution of this ubiquitin-dependent chromatin response. We show that HERC2 and RNF168 are novel DNA damage-dependent SUMOylation targets in human cells. In response to DSBs, both HERC2 and RNF168 were specifically modified with SUMO1...... at DSB sites in a manner dependent on the SUMO E3 ligase PIAS4. SUMOylation of HERC2 was required for its DSB-induced association with RNF8 and for stabilizing the RNF8-Ubc13 complex. We also demonstrate that the ZZ Zinc finger in HERC2 defined a novel SUMO-specific binding module, which together...

  18. Evolutionary expansion and divergence in a large family of primate-specific zinc finger transcription factor genes

    Energy Technology Data Exchange (ETDEWEB)

    Hamilton, A T; Huntley, S; Tran-Gyamfi, M; Baggott, D; Gordon, L; Stubbs, L

    2005-09-28

    Although most genes are conserved as one-to-one orthologs in different mammalian orders, certain gene families have evolved to comprise different numbers and types of protein-coding genes through independent series of gene duplications, divergence and gene loss in each evolutionary lineage. One such family encodes KRAB-zinc finger (KRAB-ZNF) genes, which are likely to function as transcriptional repressors. One KRAB-ZNF subfamily, the ZNF91 clade, has expanded specifically in primates to comprise more than 110 loci in the human genome, yielding large gene clusters in human chromosomes 19 and 7 and smaller clusters or isolated copies at other chromosomal locations. Although phylogenetic analysis indicates that many of these genes arose before the split between old world monkeys and new world monkeys, the ZNF91 subfamily has continued to expand and diversify throughout the evolution of apes and humans. The paralogous loci are distinguished by sequence divergence within their zinc finger arrays indicating a selection for proteins with different DNA binding specificities. RT-PCR and in situ hybridization data show that some of these ZNF genes can have tissue-specific expression patterns, however many KRAB-ZNFs that are near-ubiquitous could also be playing very specific roles in halting target pathways in all tissues except for a few, where the target is released by the absence of its repressor. The number of variant KRAB-ZNF proteins is increased not only because of the large number of loci, but also because many loci can produce multiple splice variants, which because of the modular structure of these genes may have separate and perhaps even conflicting regulatory roles. The lineage-specific duplication and rapid divergence of this family of transcription factor genes suggests a role in determining species-specific biological differences and the evolution of novel primate traits.

  19. Engineering zinc finger protein transcription factors to downregulate the epithelial glycoprotein-2 promoter as a novel anti-cancer treatment.

    Science.gov (United States)

    Gommans, Willemijn M; McLaughlin, Pamela M J; Lindhout, Beatrice I; Segal, David J; Wiegman, D J; Haisma, Hidde J; van der Zaal, Bert J; Rots, Marianne G

    2007-05-01

    Zinc finger protein transcription factors (ZFP-TFs) are emerging as powerful novel tools for the treatment of many different diseases. ZFPs are DNA-binding motifs and consist of modular zinc finger domains. Each domain can be engineered to recognize a specific DNA triplet, and stitching six domains together results in the recognition of a gene-specific sequence. Inhibition of gene expression can be achieved by fusing a repressor domain to these DNA-binding motifs. In this study, we engineered ZFP-TFs to downregulate the activity of the epithelial glycoprotein-2 (EGP-2) promoter. The protein EGP-2 is overexpressed in a wide variety of cancer types and EGP-2 downregulation has been shown to result in a decreased oncogenic potential of tumor cells. Therefore, downregulation of EGP-2 expression by ZFP-TFs provides a novel anti-cancer therapeutic. Using a straightforward strategy, we engineered a 3-ZFP that could bind a 9 bp sequence within the EGP-2 promoter. After the addition of a repressor domain, this 3-ZFP-TF could efficiently downregulate EGP-2 promoter activity by 60%. To demonstrate the flexibility of this technology, we coupled an activation domain to the engineered ZFP, resulting in a nearly 200% increase in EGP-2 promoter activity. To inhibit the endogenous EGP-2 promoter, we engineered 6-ZFP-TFs. Although none of the constructed ZFP-TFs could convincingly modulate the endogenous promoter, efficient and specific inhibition of the exogenous promoter was observed. Overall, ZFP-TFs are versatile bi-directional modulators of gene expression and downregulation of EGP-2 promoter activity using ZFP-TFs can ultimately result in a novel anti-cancer treatment.

  20. C. elegans PAT-9 is a nuclear zinc finger protein critical for the assembly of muscle attachments

    Directory of Open Access Journals (Sweden)

    Liu Qian

    2012-05-01

    Full Text Available Abstract Background Caenorhabditis elegans sarcomeres have been studied extensively utilizing both forward and reverse genetic techniques to provide insight into muscle development and the mechanisms behind muscle contraction. A previous genetic screen investigating early muscle development produced 13 independent mutant genes exhibiting a Pat (paralyzed and arrested elongation at the two-fold length of embryonic development muscle phenotype. This study reports the identification and characterization of one of those genes, pat-9. Results Positional cloning, reverse genetics, and plasmid rescue experiments were used to identify the predicted C. elegans gene T27B1.2 (recently named ztf-19 as the pat-9 gene. Analysis of pat-9 showed it is expressed early in development and within body wall muscle lineages, consistent with a role in muscle development and producing a Pat phenotype. However, unlike most of the other known Pat gene family members, which encode structural components of muscle attachment sites, PAT-9 is an exclusively nuclear protein. Analysis of the predicted PAT-9 amino acid sequence identified one putative nuclear localization domain and three C2H2 zinc finger domains. Both immunocytochemistry and PAT-9::GFP fusion expression confirm that PAT-9 is primarily a nuclear protein and chromatin immunoprecipitation (ChIP experiments showed that PAT-9 is present on certain gene promoters. Conclusions We have shown that the T27B1.2 gene is pat-9. Considering the Pat-9 mutant phenotype shows severely disrupted muscle attachment sites despite PAT-9 being a nuclear zinc finger protein and not a structural component of muscle attachment sites, we propose that PAT-9 likely functions in the regulation of gene expression for some necessary structural or regulatory component(s of the muscle attachment sites.

  1. Sequence specificity is obtained from the majority of modular C2H2 zinc-finger arrays.

    Science.gov (United States)

    Lam, Kathy N; van Bakel, Harm; Cote, Atina G; van der Ven, Anton; Hughes, Timothy R

    2011-06-01

    C2H2 zinc fingers (C2H2-ZFs) are the most prevalent type of vertebrate DNA-binding domain, and typically appear in tandem arrays (ZFAs), with sequential C2H2-ZFs each contacting three (or more) sequential bases. C2H2-ZFs can be assembled in a modular fashion, providing one explanation for their remarkable evolutionary success. Given a set of modules with defined three-base specificities, modular assembly also presents a way to construct artificial proteins with specific DNA-binding preferences. However, a recent survey of a large number of three-finger ZFAs engineered by modular assembly reported high failure rates (∼70%), casting doubt on the generality of modular assembly. Here, we used protein-binding microarrays to analyze 28 ZFAs that failed in the aforementioned study. Most (17) preferred specific sequences, which in all but one case resembled the intended target sequence. Like natural ZFAs, the engineered ZFAs typically yielded degenerate motifs, binding dozens to hundreds of related individual sequences. Thus, the failure of these proteins in previous assays is not due to lack of sequence-specific DNA-binding activity. Our findings underscore the relevance of individual C2H2-ZF sequence specificities within tandem arrays, and support the general ability of modular assembly to produce ZFAs with sequence-specific DNA-binding activity.

  2. Zinc finger nucleases for targeted mutagenesis and repair of the sickle-cell disease mutation: An in-silico study

    Directory of Open Access Journals (Sweden)

    Wayengera Misaki

    2012-05-01

    Full Text Available Abstract Background Sickle cell disease (or simply, SCD is an inherited hemoglobinopathy which is mostly prevalent among persons of African descent. SCD results from a monogenic (Hemoglobin, beta point-mutation (substitution of the base Adenine with Thymine at position six that leads to replacement of the amino acid glutamic acid (E with valine (V. Management of SCD within resource-poor settings is largely syndromic, since the option of cure offered by bone-marrow transplantation (BMT is risky and unaffordable by most affected individuals. Despite previous reports of repair and inhibition of the sickle beta-globin gene and messenger ribonucleic acids (mRNAs, respectively in erythrocyte precursor cells via gene-targeting using an oligomer-restriction enzyme construct and either ribozyme- or RNA-DNA chimeric oligonucleotides (or simply third strand binding, gene-therapy to treat SCD still remains largely preclinical. In the wake of the advances in target- gene- mutagenesis and repair wrought by zinc finger nuclease (ZFN technology, it was hypothesized that SCD may be cured by the same. The goal of this study thus, was constructing a database of zinc finger arrays (ZFAs and engineering ZFNs, that respectively bind and cleave within or around specific sequences in the sickle hemoglobin, beta (−βS gene. Methods and results First, using the complete 1606 genomic DNA base pair (bp sequences of the normal hemoglobin-beta (βA chain gene, and the ZiFiT-CoDA-ZFA software preset at default, 57 three-finger arrays (ZFAs that specifically bind 9 base-pair sequences within the normal hemoglobin-beta chain, were computationally assembled. Second, by serial linkage of these ZFAs to the Flavobacterium okeanokoites endonuclease Fok I― four ZFNs with unique specificity to >24 bp target-sequences at the genomic contextual positions 82, 1333, 1334, and 1413 of the βA chain-gene were constructed in-silico. Third, localizing the point-mutation of SCD at

  3. Molecular Characterization of the Schistosoma mansoni Zinc Finger Protein SmZF1 as a Transcription Factor

    Science.gov (United States)

    D'Astolfo, Diego S.; Cardoso, Fernanda C.; Rajão, Matheus A.; Mourão, Marina M.; Gava, Elisandra; Oliveira, Sérgio C.; Macedo, Andréa M.; Machado, Carlos R.; Pena, Sérgio D. J.; Kitten, Gregory T.; Franco, Glória R.

    2009-01-01

    Background During its development, the parasite Schistosoma mansoni is exposed to different environments and undergoes many morphological and physiological transformations as a result of profound changes in gene expression. Characterization of proteins involved in the regulation of these processes is of importance for the understanding of schistosome biology. Proteins containing zinc finger motifs usually participate in regulatory processes and are considered the major class of transcription factors in eukaryotes. It has already been shown, by EMSA (Eletrophoretic Mobility Shift Assay), that SmZF1, a S. mansoni zinc finger (ZF) protein, specifically binds both DNA and RNA oligonucleotides. This suggests that this protein might act as a transcription factor in the parasite. Methodology/Principal Findings In this study we extended the characterization of SmZF1 by determining its subcellular localization and by verifying its ability to regulate gene transcription. We performed immunohistochemistry assays using adult male and female worms, cercariae and schistosomula to analyze the distribution pattern of SmZF1 and verified that the protein is mainly detected in the cells nuclei of all tested life cycle stages except for adult female worms. Also, SmZF1 was heterologously expressed in mammalian COS-7 cells to produce the recombinant protein YFP-SmZF1, which was mainly detected in the nucleus of the cells by confocal microscopy and Western blot assays. To evaluate the ability of this protein to regulate gene transcription, cells expressing YFP-SmZF1 were tested in a luciferase reporter system. In this system, the luciferase gene is downstream of a minimal promoter, upstream of which a DNA region containing four copies of the SmZF1 putative best binding site (D1-3DNA) was inserted. SmZF1 increased the reporter gene transcription by two fold (p≤0.003) only when its specific binding site was present. Conclusion Taken together, these results strongly support the hypothesis

  4. ErbB2-Driven Breast Cancer Cell Invasion Depends on a Complex Signaling Network Activating Myeloid Zinc Finger-1-Dependent Cathepsin B Expression

    DEFF Research Database (Denmark)

    Rafn, Bo; Nielsen, Christian Thomas Friberg; Andersen, Sofie Hagel;

    2012-01-01

    signaling network activates the transcription of cathepsin B gene (CTSB) via myeloid zinc finger-1 transcription factor that binds to an ErbB2-responsive enhancer element in the first intron of CTSB. This work provides a model system for ErbB2-induced breast cancer cell invasiveness, reveals a signaling...... network that is crucial for invasion in vitro, and defines a specific role and targets for the identified serine-threonine kinases....

  5. Gene targeting technologies in rats: zinc finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats.

    OpenAIRE

    Mashimo, Tomoji

    2013-01-01

    The laboratory rat has been widely used as an animal model in biomedical science for more than 150 years. Applying zinc-finger nucleases or transcription activator-like effector nucleases to rat embryos via microinjection is an efficient genome editing tool for generating targeted knockout rats. Recently, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated endonucleases have been used as an effective tool for precise and multiplex genome editing in mice and ra...

  6. Generation of mastitis resistance in cows by targeting human lysozyme gene to β-casein locus using zinc-finger nucleases

    OpenAIRE

    Liu, Xu; Wang, Yongsheng; Tian, Yuchen; Yu, Yuan; Gao, Mingqing; Hu, Guangdong; Su, Feng; Pan, Shaohui; Luo, Yan; Guo, Zekun; Quan, Fusheng; Zhang, Yong

    2014-01-01

    Mastitis costs the dairy industry billions of dollars annually and is the most consequential disease of dairy cattle. Transgenic cows secreting an antimicrobial peptide demonstrated resistance to mastitis. The combination of somatic cell gene targeting and nuclear transfer provides a powerful method to produce transgenic animals. Recent studies found that a precisely placed double-strand break induced by engineered zinc-finger nucleases (ZFNs) stimulated the integration of exogenous DNA stret...

  7. The Solanum lycopersicum Zinc Finger2 cysteine-2/histidine-2 repressor-like transcription factor regulates development and tolerance to salinity in tomato and Arabidopsis.

    Science.gov (United States)

    Hichri, Imène; Muhovski, Yordan; Žižkova, Eva; Dobrev, Petre I; Franco-Zorrilla, Jose Manuel; Solano, Roberto; Lopez-Vidriero, Irene; Motyka, Vaclav; Lutts, Stanley

    2014-04-01

    The zinc finger superfamily includes transcription factors that regulate multiple aspects of plant development and were recently shown to regulate abiotic stress tolerance. Cultivated tomato (Solanum lycopersicum Zinc Finger2 [SIZF2]) is a cysteine-2/histidine-2-type zinc finger transcription factor bearing an ERF-associated amphiphilic repression domain and binding to the ACGTCAGTG sequence containing two AGT core motifs. SlZF2 is ubiquitously expressed during plant development, and is rapidly induced by sodium chloride, drought, and potassium chloride treatments. Its ectopic expression in Arabidopsis (Arabidopsis thaliana) and tomato impaired development and influenced leaf and flower shape, while causing a general stress visible by anthocyanin and malonyldialdehyde accumulation. SlZF2 enhanced salt sensitivity in Arabidopsis, whereas SlZF2 delayed senescence and improved tomato salt tolerance, particularly by maintaining photosynthesis and increasing polyamine biosynthesis, in salt-treated hydroponic cultures (125 mm sodium chloride, 20 d). SlZF2 may be involved in abscisic acid (ABA) biosynthesis/signaling, because SlZF2 is rapidly induced by ABA treatment and 35S::SlZF2 tomatoes accumulate more ABA than wild-type plants. Transcriptome analysis of 35S::SlZF2 revealed that SlZF2 both increased and reduced expression of a comparable number of genes involved in various physiological processes such as photosynthesis, polyamine biosynthesis, and hormone (notably ABA) biosynthesis/signaling. Involvement of these different metabolic pathways in salt stress tolerance is discussed.

  8. Rice zinc finger protein DST enhances grain production through controlling Gn1a/OsCKX2 expression.

    Science.gov (United States)

    Li, Shuyu; Zhao, Bingran; Yuan, Dingyang; Duan, Meijuan; Qian, Qian; Tang, Li; Wang, Bao; Liu, Xiaoqiang; Zhang, Jie; Wang, Jun; Sun, Jiaqiang; Liu, Zhao; Feng, Yu-Qi; Yuan, Longping; Li, Chuanyou

    2013-02-19

    The phytohormone cytokinin (CK) positively regulates the activity and function of the shoot apical meristem (SAM), which is a major parameter determining seed production. The rice (Oryza sativa L.) Gn1a/OsCKX2 (Grain number 1a/Cytokinin oxidase 2) gene, which encodes a cytokinin oxidase, has been identified as a major quantitative trait locus contributing to grain number improvement in rice breeding practice. However, the molecular mechanism of how the expression of OsCKX2 is regulated in planta remains elusive. Here, we report that the zinc finger transcription factor DROUGHT AND SALT TOLERANCE (DST) directly regulates OsCKX2 expression in the reproductive meristem. DST-directed expression of OsCKX2 regulates CK accumulation in the SAM and, therefore, controls the number of the reproductive organs. We identify that DST(reg1), a semidominant allele of the DST gene, perturbs DST-directed regulation of OsCKX2 expression and elevates CK levels in the reproductive SAM, leading to increased meristem activity, enhanced panicle branching, and a consequent increase of grain number. Importantly, the DST(reg1) allele provides an approach to pyramid the Gn1a-dependent and Gn1a-independent effects on grain production. Our study reveals that, as a unique regulator of reproductive meristem activity, DST may be explored to facilitate the genetic enhancement of grain production in rice and other small grain cereals.

  9. A previously unknown zinc finger protein, DST, regulates drought and salt tolerance in rice via stomatal aperture control.

    Science.gov (United States)

    Huang, Xin-Yuan; Chao, Dai-Yin; Gao, Ji-Ping; Zhu, Mei-Zhen; Shi, Min; Lin, Hong-Xuan

    2009-08-01

    Abiotic stresses, such as drought and salinity, lead to crop growth damage and a decrease in crop yields. Stomata control CO(2) uptake and optimize water use efficiency, thereby playing crucial roles in abiotic stress tolerance. Hydrogen peroxide (H(2)O(2)) is an important signal molecule that induces stomatal closure. However, the molecular pathway that regulates the H(2)O(2) level in guard cells remains largely unknown. Here, we clone and characterize DST (drought and salt tolerance)-a previously unknown zinc finger transcription factor that negatively regulates stomatal closure by direct modulation of genes related to H(2)O(2) homeostasis-and identify a novel pathway for the signal transduction of DST-mediated H(2)O(2)-induced stomatal closure. Loss of DST function increases stomatal closure and reduces stomatal density, consequently resulting in enhanced drought and salt tolerance in rice. These findings provide an interesting insight into the mechanism of stomata-regulated abiotic stress tolerance, and an important genetic engineering approach for improving abiotic stress tolerance in crops.

  10. A novel cold-inducible zinc finger protein from soybean, SCOF-1, enhances cold tolerance in transgenic plants.

    Science.gov (United States)

    Kim, J C; Lee, S H; Cheong, Y H; Yoo, C M; Lee, S I; Chun, H J; Yun, D J; Hong, J C; Lee, S Y; Lim, C O; Cho, M J

    2001-02-01

    Cold stress on plants induces changes in the transcription of cold response genes. A cDNA clone encoding C2H2-type zinc finger protein, SCOF-1, was isolated from soybean. The transcription of SCOF-1 is specifically induced by low temperature and abscisic acid (ABA) but not by dehydration or high salinity. Constitutive overexpression of SCOF-1 induced cold-regulated (COR) gene expression and enhanced cold tolerance of non-acclimated transgenic Arabidopsis and tobacco plants. SCOF-1 localized to the nucleus but did not bind directly to either C-repeat/dehydration (CRT/DRE) or ABA responsive element (ABRE), cis-acting DNA regulatory elements present in COR gene promoters. However, SCOF-1 greatly enhanced the DNA binding activity of SGBF-1, a soybean G-box binding bZIP transcription factor, to ABRE in vitro. SCOF-1 also interacted with SGBF-1 in a yeast two-hybrid system. The SGBF-1 transactivated the beta-glucuronidase reporter gene driven by the ABRE element in Arabidopsis leaf protoplasts. Furthermore, the SCOF-1 enhanced ABRE-dependent gene expression mediated by SGBF-1. These results suggest that SCOF-1 may function as a positive regulator of COR gene expression mediated by ABRE via protein-protein interaction, which in turn enhances cold tolerance of plants.

  11. Dissection of splicing regulation at an endogenous locus by zinc-finger nuclease-mediated gene editing.

    Directory of Open Access Journals (Sweden)

    Sandra Cristea

    Full Text Available Sequences governing RNA splicing are difficult to study in situ due to the great difficulty of traditional targeted mutagenesis. Zinc-finger nuclease (ZFN technology allows for the rapid and efficient introduction of site-specific mutations into mammalian chromosomes. Using a ZFN pair along with a donor plasmid to manipulate the outcomes of DNA repair, we introduced several discrete, targeted mutations into the fourth intron of the endogenous BAX gene in Chinese hamster ovary cells. Putative lariat branch points, the polypyrimidine tract, and the splice acceptor site were targeted. We recovered numerous otherwise isogenic clones carrying the intended mutations and analyzed the effect of each on BAX pre-mRNA splicing. Mutation of one of three possible branch points, the polypyrimidine tract, and the splice acceptor site all caused exclusion of exon five from BAX mRNA. Interestingly, these exon-skipping mutations allowed usage of cryptic splice acceptor sites within intron four. These data demonstrate that ZFN-mediated gene editing is a highly effective tool for dissection of pre-mRNA splicing regulatory sequences in their endogenous context.

  12. Multiple interferon stimulated genes synergize with the zinc finger antiviral protein to mediate anti-alphavirus activity.

    Directory of Open Access Journals (Sweden)

    Sophiya Karki

    Full Text Available The zinc finger antiviral protein (ZAP is a host factor that mediates inhibition of viruses in the Filoviridae, Retroviridae and Togaviridae families. We previously demonstrated that ZAP blocks replication of Sindbis virus (SINV, the prototype Alphavirus in the Togaviridae family at an early step prior to translation of the incoming genome and that synergy between ZAP and one or more interferon stimulated genes (ISGs resulted in maximal inhibitory activity. The present study aimed to identify those ISGs that synergize with ZAP to mediate Alphavirus inhibition. Using a library of lentiviruses individually expressing more than 350 ISGs, we screened for inhibitory activity in interferon defective cells with or without ZAP overexpression. Confirmatory tests of the 23 ISGs demonstrating the largest infection reduction in combination with ZAP revealed that 16 were synergistic. Confirmatory tests of all potentially synergistic ISGs revealed 15 additional ISGs with a statistically significant synergistic effect in combination with ZAP. These 31 ISGs are candidates for further mechanistic studies. The number and diversity of the identified ZAP-synergistic ISGs lead us to speculate that ZAP may play an important role in priming the cell for optimal ISG function.

  13. Multiple interferon stimulated genes synergize with the zinc finger antiviral protein to mediate anti-alphavirus activity.

    Science.gov (United States)

    Karki, Sophiya; Li, Melody M H; Schoggins, John W; Tian, Suyan; Rice, Charles M; MacDonald, Margaret R

    2012-01-01

    The zinc finger antiviral protein (ZAP) is a host factor that mediates inhibition of viruses in the Filoviridae, Retroviridae and Togaviridae families. We previously demonstrated that ZAP blocks replication of Sindbis virus (SINV), the prototype Alphavirus in the Togaviridae family at an early step prior to translation of the incoming genome and that synergy between ZAP and one or more interferon stimulated genes (ISGs) resulted in maximal inhibitory activity. The present study aimed to identify those ISGs that synergize with ZAP to mediate Alphavirus inhibition. Using a library of lentiviruses individually expressing more than 350 ISGs, we screened for inhibitory activity in interferon defective cells with or without ZAP overexpression. Confirmatory tests of the 23 ISGs demonstrating the largest infection reduction in combination with ZAP revealed that 16 were synergistic. Confirmatory tests of all potentially synergistic ISGs revealed 15 additional ISGs with a statistically significant synergistic effect in combination with ZAP. These 31 ISGs are candidates for further mechanistic studies. The number and diversity of the identified ZAP-synergistic ISGs lead us to speculate that ZAP may play an important role in priming the cell for optimal ISG function.

  14. Generation of Knockout Rats with X-Linked Severe Combined Immunodeficiency (X-SCID) Using Zinc-Finger Nucleases

    Science.gov (United States)

    Mashimo, Tomoji; Takizawa, Akiko; Voigt, Birger; Yoshimi, Kazuto; Hiai, Hiroshi; Kuramoto, Takashi; Serikawa, Tadao

    2010-01-01

    Background Although the rat is extensively used as a laboratory model, the inability to utilize germ line-competent rat embryonic stem (ES) cells has been a major drawback for studies that aim to elucidate gene functions. Recently, zinc-finger nucleases (ZFNs) were successfully used to create genome-specific double-stranded breaks and thereby induce targeted gene mutations in a wide variety of organisms including plants, drosophila, zebrafish, etc. Methodology/Principal Findings We report here on ZFN-induced gene targeting of the rat interleukin 2 receptor gamma (Il2rg) locus, where orthologous human and mouse mutations cause X-linked severe combined immune deficiency (X-SCID). Co-injection of mRNAs encoding custom-designed ZFNs into the pronucleus of fertilized oocytes yielded genetically modified offspring at rates greater than 20%, which possessed a wide variety of deletion/insertion mutations. ZFN-modified founders faithfully transmitted their genetic changes to the next generation along with the severe combined immune deficiency phenotype. Conclusions and Significance The efficient and rapid generation of gene knockout rats shows that using ZFN technology is a new strategy for creating gene-targeted rat models of human diseases. In addition, the X-SCID rats that were established in this study will be valuable in vivo tools for evaluating drug treatment or gene therapy as well as model systems for examining the treatment of xenotransplanted malignancies. PMID:20111598

  15. Myeloid zinc finger 1 mediates sulindac sulfide-induced upregulation of death receptor 5 of human colon cancer cells.

    Science.gov (United States)

    Horinaka, Mano; Yoshida, Tatsushi; Tomosugi, Mitsuhiro; Yasuda, Shusuke; Sowa, Yoshihiro; Sakai, Toshiyuki

    2014-08-08

    A combined therapy of sulindac sulfide and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising strategy for the treatment of cancer. Sulindac sulfide had been shown to induce the expression of death receptor 5 (DR5), a receptor for TRAIL, and sensitize cancer cells to TRAIL-induced apoptosis; however, the molecular mechanism underlying the upregulation of DR5 has not yet been elucidated. We demonstrate here that myeloid zinc finger 1 (MZF1) mediates the induction of DR5 by sulindac sulfide. Sulindac sulfide induced the expression of DR5 at the protein and mRNA levels in colon cancer SW480 cells. Furthermore, sulindac sulfide increased DR5 promoter activity. We showed that sulindac sulfide stimulated DR5 promoter activity via the -301 to -253 region. This region contained a putative MZF1-binding site. Site-directed mutations in the site abrogated the enhancement in DR5 promoter activity by sulindac sulfide. MZF1 directly bound to the putative MZF1-binding site of the DR5 promoter and the binding was increased by sulindac sulfide. The expression of MZF1 was also increased by sulindac sulfide, and MZF1 siRNA attenuated the upregulation of DR5 by sulindac sulfide. These results indicate that sulindac sulfide induces the expression of DR5 by up-regulating MZF1.

  16. Zinc Finger Homeodomain Factor Zfhx3 Is Essential for Mammary Lactogenic Differentiation by Maintaining Prolactin Signaling Activity.

    Science.gov (United States)

    Zhao, Dan; Ma, Gui; Zhang, Xiaolin; He, Yuan; Li, Mei; Han, Xueying; Fu, Liya; Dong, Xue-Yuan; Nagy, Tamas; Zhao, Qiang; Fu, Li; Dong, Jin-Tang

    2016-06-10

    The zinc finger homeobox 3 (ZFHX3, also named ATBF1 for AT motif binding factor 1) is a transcription factor that suppresses prostatic carcinogenesis and induces neuronal differentiation. It also interacts with estrogen receptor α to inhibit cell proliferation and regulate pubertal mammary gland development in mice. In the present study, we examined whether and how Zfhx3 regulates lactogenic differentiation in mouse mammary glands. At different stages of mammary gland development, Zfhx3 protein was expressed at varying levels, with the highest level at lactation. In the HC11 mouse mammary epithelial cell line, an in vitro model of lactogenesis, knockdown of Zfhx3 attenuated prolactin-induced β-casein expression and morphological changes, indicators of lactogenic differentiation. In mouse mammary tissue, knock-out of Zfhx3 interrupted lactogenesis, resulting in underdeveloped glands with much smaller and fewer alveoli, reduced β-casein expression, accumulation of large cytoplasmic lipid droplets in luminal cells after parturition, and failure in lactation. Mechanistically, Zfhx3 maintained the expression of Prlr (prolactin receptor) and Prlr-Jak2-Stat5 signaling activity, whereas knockdown and knock-out of Zfhx3 in HC11 cells and mammary tissues, respectively, decreased Prlr expression, Stat5 phosphorylation, and the expression of Prlr-Jak2-Stat5 target genes. These findings indicate that Zfhx3 plays an essential role in proper lactogenic development in mammary glands, at least in part by maintaining Prlr expression and Prlr-Jak2-Stat5 signaling activity.

  17. Generation of knockout rats with X-linked severe combined immunodeficiency (X-SCID using zinc-finger nucleases.

    Directory of Open Access Journals (Sweden)

    Tomoji Mashimo

    Full Text Available BACKGROUND: Although the rat is extensively used as a laboratory model, the inability to utilize germ line-competent rat embryonic stem (ES cells has been a major drawback for studies that aim to elucidate gene functions. Recently, zinc-finger nucleases (ZFNs were successfully used to create genome-specific double-stranded breaks and thereby induce targeted gene mutations in a wide variety of organisms including plants, drosophila, zebrafish, etc. METHODOLOGY/PRINCIPAL FINDINGS: We report here on ZFN-induced gene targeting of the rat interleukin 2 receptor gamma (Il2rg locus, where orthologous human and mouse mutations cause X-linked severe combined immune deficiency (X-SCID. Co-injection of mRNAs encoding custom-designed ZFNs into the pronucleus of fertilized oocytes yielded genetically modified offspring at rates greater than 20%, which possessed a wide variety of deletion/insertion mutations. ZFN-modified founders faithfully transmitted their genetic changes to the next generation along with the severe combined immune deficiency phenotype. CONCLUSIONS AND SIGNIFICANCE: The efficient and rapid generation of gene knockout rats shows that using ZFN technology is a new strategy for creating gene-targeted rat models of human diseases. In addition, the X-SCID rats that were established in this study will be valuable in vivo tools for evaluating drug treatment or gene therapy as well as model systems for examining the treatment of xenotransplanted malignancies.

  18. The Fruitless gene in Nasonia displays complex sex-specific splicing and contains new zinc finger domains.

    Science.gov (United States)

    Bertossa, Rinaldo C; van de Zande, Louis; Beukeboom, Leo W

    2009-07-01

    The transcription factor Fruitless exerts a broad range of functions during Drosophila development, the most apparent of which is the determination of sexual behavior in males. Although fruitless sequences are found in other insect orders, little is known about fruitless structure and function outside Diptera. We have performed a thorough analysis of fruitless transcripts in the haplo-diploid wasp Nasonia vitripennis and found both sex-specific and non-sex-specific transcripts similar to those found in Drosophila. In Nasonia, however, a novel, large fruitless transcript is present in females only. Putative binding sites for sex-specific splicing factors found in Nasonia fruitless and doublesex as well as Apis mellifera doublesex transcripts were sufficient to identify a corresponding female-specific fruitless exon in A. mellifera, suggesting that similar factors in both hymenopteran species could be responsible for sex-specific splicing of both genes. Furthermore, new C(2)H(2) zinc finger domains found in Nasonia fruitless transcripts were also identified in the fruitless locus of major holometabolous insect species but not in drosophilids. Conservation of important domains and sex-specific splicing in Diptera and Hymenoptera support the hypothesis that fruitless is an ancient gene and has conserved functions in insects. Considerable divergences in other parts of the gene are expected to underlie species-specific differences and may help to explain diversity observed in insect sexual behaviors.

  19. Evaluation of OPEN zinc finger nucleases for direct gene targeting of the ROSA26 locus in mouse embryos.

    Directory of Open Access Journals (Sweden)

    Mario Hermann

    Full Text Available Zinc finger nucleases (ZFNs enable precise genome modification in a variety of organisms and cell types. Commercial ZFNs were reported to enhance gene targeting directly in mouse zygotes, whereas similar approaches using publicly available resources have not yet been described. Here we report precise targeted mutagenesis of the mouse genome using Oligomerized Pool Engineering (OPEN ZFNs. OPEN ZFN can be constructed using publicly available resources and therefore provide an attractive alternative for academic researchers. Two ZFN pairs specific to the mouse genomic locus gt(ROSA26Sor were generated by OPEN selections and used for gene disruption and homology-mediated gene replacement in single cell mouse embryos. One specific ZFN pair facilitated non-homologous end joining (NHEJ-mediated gene disruption when expressed in mouse zygotes. We also observed a single homologous recombination (HR-driven gene replacement event when this ZFN pair was co-injected with a targeting vector. Our experiments demonstrate the feasibility of achieving both gene ablation through NHEJ and gene replacement by HR by using the OPEN ZFN technology directly in mouse zygotes.

  20. Genome-Wide Identification, Evolution and Expression Analysis of the Grape (Vitis vinifera L. Zinc Finger-Homeodomain Gene Family

    Directory of Open Access Journals (Sweden)

    Hao Wang

    2014-04-01

    Full Text Available Plant zinc finger-homeodomain (ZHD genes encode a family of transcription factors that have been demonstrated to play an important role in the regulation of plant growth and development. In this study, we identified a total of 13 ZHD genes (VvZHD in the grape genome that were further classified into at least seven groups. Genome synteny analysis revealed that a number of VvZHD genes were present in the corresponding syntenic blocks of Arabidopsis, indicating that they arose before the divergence of these two species. Gene expression analysis showed that the identified VvZHD genes displayed distinct spatiotemporal expression patterns, and were differentially regulated under various stress conditions and hormone treatments, suggesting that the grape VvZHDs might be also involved in plant response to a variety of biotic and abiotic insults. Our work provides insightful information and knowledge about the ZHD genes in grape, which provides a framework for further characterization of their roles in regulation of stress tolerance as well as other aspects of grape productivity.

  1. Genomic editing of the HIV-1 coreceptor CCR5 in adult hematopoietic stem and progenitor cells using zinc finger nucleases.

    Science.gov (United States)

    Li, Lijing; Krymskaya, Ludmila; Wang, Jianbin; Henley, Jill; Rao, Anitha; Cao, Lan-Feng; Tran, Chy-Anh; Torres-Coronado, Monica; Gardner, Agnes; Gonzalez, Nancy; Kim, Kenneth; Liu, Pei-Qi; Hofer, Ursula; Lopez, Evan; Gregory, Philip D; Liu, Qing; Holmes, Michael C; Cannon, Paula M; Zaia, John A; DiGiusto, David L

    2013-06-01

    The HIV-1 coreceptor CCR5 is a validated target for HIV/AIDS therapy. The apparent elimination of HIV-1 in a patient treated with an allogeneic stem cell transplant homozygous for a naturally occurring CCR5 deletion mutation (CCR5(Δ32/Δ32)) supports the concept that a single dose of HIV-resistant hematopoietic stem cells can provide disease protection. Given the low frequency of naturally occurring CCR5(Δ32/Δ32) donors, we reasoned that engineered autologous CD34(+) hematopoietic stem/progenitor cells (HSPCs) could be used for AIDS therapy. We evaluated disruption of CCR5 gene expression in HSPCs isolated from granulocyte colony-stimulating factor (CSF)-mobilized adult blood using a recombinant adenoviral vector encoding a CCR5-specific pair of zinc finger nucleases (CCR5-ZFN). Our results demonstrate that CCR5-ZFN RNA and protein expression from the adenoviral vector is enhanced by pretreatment of HSPC with protein kinase C (PKC) activators resulting in >25% CCR5 gene disruption and that activation of the mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) signaling pathway is responsible for this activity. Importantly, using an optimized dose of PKC activator and adenoviral vector we could generate CCR5-modified HSPCs which engraft in a humanized mouse model (albeit at a reduced level) and support multilineage differentiation in vitro and in vivo. Together, these data establish the basis for improved approaches exploiting adenoviral vector delivery in the modification of HSPCs.

  2. Conversion of Human Fibroblasts to Stably Self-Renewing Neural Stem Cells with a Single Zinc-Finger Transcription Factor

    Directory of Open Access Journals (Sweden)

    Ebrahim Shahbazi

    2016-04-01

    Full Text Available Direct conversion of somatic cells into neural stem cells (NSCs by defined factors holds great promise for mechanistic studies, drug screening, and potential cell therapies for different neurodegenerative diseases. Here, we report that a single zinc-finger transcription factor, Zfp521, is sufficient for direct conversion of human fibroblasts into long-term self-renewable and multipotent NSCs. In vitro, Zfp521-induced NSCs maintained their characteristics in the absence of exogenous factor expression and exhibited morphological, molecular, developmental, and functional properties that were similar to control NSCs. In addition, the single-seeded induced NSCs were able to form NSC colonies with efficiency comparable with control NSCs and expressed NSC markers. The converted cells were capable of surviving, migrating, and attaining neural phenotypes after transplantation into neonatal mouse and adult rat brains, without forming tumors. Moreover, the Zfp521-induced NSCs predominantly expressed rostral genes. Our results suggest a facilitated approach for establishing human NSCs through Zfp521-driven conversion of fibroblasts.

  3. The Transcriptional Repressive Activity of KRAB Zinc Finger Proteins Does Not Correlate with Their Ability to Recruit TRIM28

    Science.gov (United States)

    Murphy, Kristin E.; Shylo, Natalia A.; Alexander, Katherine A.; Churchill, Angela J.; Copperman, Cecilia; García-García, María J.

    2016-01-01

    KRAB domain Zinc finger proteins are one of the most abundant families of transcriptional regulators in higher vertebrates. The prevailing view is that KRAB domain proteins function as potent transcriptional repressors by recruiting TRIM28 and promoting heterochromatin spreading. However, the extent to which all KRAB domain proteins are TRIM28-dependent transcriptional repressors is currently unclear. Our studies on mouse ZFP568 revealed that TRIM28 recruitment by KRAB domain proteins is not sufficient to warrant transcriptional repressive activity. By using luciferase reporter assays and yeast two-hybrid experiments, we tested the ability of ZFP568 and other mouse KRAB domain proteins to repress transcription and bind TRIM28. We found that some mouse KRAB domain proteins are poor transcriptional repressors despite their ability to recruit TRIM28, while others showed strong KRAB-dependent transcriptional repression, but no TRIM28 binding. Together, our results show that the transcriptional repressive activity of KRAB-ZNF proteins does not correlate with their ability to recruit TRIM28, and provide evidence that KRAB domains can regulate transcription in a TRIM28-independent fashion. Our findings challenge the current understanding of the molecular mechanisms used by KRAB domain proteins to control gene expression and highlight that a high percentage of KRAB domain proteins in the mouse genome differ from the consensus KRAB sequence at amino acid residues that are critical for TRIM28 binding and/or repressive activity. PMID:27658112

  4. Generation of PPARγ mono-allelic knockout pigs via zinc-finger nucleases and nuclear transfer cloning

    Institute of Scientific and Technical Information of China (English)

    Dongshan Yang; Jiangtian Tian; Feng Li; Jifeng Zhang; Lin Chang; Duanqing Pei; Y Eugene Chen; Liangxue Lai; Huaqiang Yang; Wei Li; Bentian Zhao; Zhen Ouyang; Zhaoming Liu; Yu Zhao; Nana Fan; Jun Song

    2011-01-01

    @@ Dear Editor, Gene targeting in mouse embryonic stem (ES) cells has revolutionized the field of mouse genetics and allowed for the analysis of diverse aspects of gene function in vivo.For more than two decades,researchers have been actively searching for ES cells from large animals such as pigs and cattle.Unfortunately,to date,no ES cell lines from large animals have passed the crucial test of germ line contribution.The sole method of gene targeting to date in these species remains somatic cell nuclear transfer (SCNT) combined with DNA homologous recombination (HR).Due to the limited proliferation competency and extremely low frequency of HR in somatic cells (less than 10-6),this process is highly inefficient and only a few successful examples have been achieved,even though enrichment strategies such as positivenegative marker selection,promoter-trap and adenoassociated viral delivery were previously used [1-3].The low efficiency of gene targeting in cultured somatic cells is the main barrier for gene targeting in large animals.Recently,zinc-finger nuclease (ZFN) technology has emerged as a powerful tool for genome editing.The success of ZFN technology for gene targeting in fruit flies,zebra fish,rodents as well as human cell lines encouraged us to establish a high-efficiency gene-targeting platform in large animals such as pigs [4-8].

  5. The multi zinc-finger protein Trps1 acts as a regulator of histone deacetylation during mitosis.

    Science.gov (United States)

    Wuelling, Manuela; Pasdziernik, Markus; Moll, Carina N; Thiesen, Andrea M; Schneider, Sabine; Johannes, Christian; Vortkamp, Andrea

    2013-07-15

    TRPS1, the gene mutated in human "Tricho-Rhino-Phalangeal syndrome," encodes a multi zinc-finger nuclear regulator of chondrocyte proliferation and differentiation. Here, we have identified a new function of Trps1 in controlling mitotic progression in chondrocytes. Loss of Trps1 in mice leads to an increased proportion of cells arrested in mitosis and, subsequently, to chromosome segregation defects. Searching for the molecular basis of the defect, we found that Trps1 acts as regulator of histone deacetylation. Trps1 interacts with two histone deacetylases, Hdac1 and Hdac4, thereby increasing their activity. Loss of Trps1 results in histone H3 hyperacetylation, which is maintained during mitosis. Consequently, chromatin condensation and binding of HP1 is impaired, and Trps1-deficient chondrocytes accumulate in prometaphase. Overexpression of Hdac4 rescues the mitotic defect of Trps1-deficient chondrocytes, identifying Trps1 as an important regulator of chromatin deacetylation during mitosis in chondrocytes. Our data provide the first evidence that the control of mitosis can be linked to the regulation of chondrocyte differentiation by epigenetic consequences of altered Hdac activity.

  6. Suppression of zinc finger protein 467 alleviates osteoporosis through promoting differentiation of adipose derived stem cells to osteoblasts

    Directory of Open Access Journals (Sweden)

    You Li

    2012-01-01

    Full Text Available Abstract Osteoblast and adipocyte are derived from common mesenchymal progenitor cells. The bone loss of osteoporosis is associated with altered progenitor differentiation from an osteoblastic to an adipocytic lineage. In this study, a comparative analysis of gene expression profiling using cDNA microarray and realtime-PCR indicated that Zinc finger protein 467 (Zfp467 involved in adipocyte and osteoblast differentiation of cultured adipose derived stem cells (ADSCs. Our results showed that RNA interference for Zfp467 in ADSCs inhibited adipocyte formation and stimulated osteoblast commitment. The mRNA levels of osteogenic and adipogenic markers in ADSCs were regulated by si-Zfp467. Zfp467 RNAi in ADSCs could restore bone function and structure in an ovariectomized (OVX-induced osteoporotic mouse model. Thus Zfp467 play an important role in ADSCs differentiation to adipocyte and osteoblast. This has relevance to therapeutic interventions in osteoporosis, including si-Zfp467-based therapies currently available, and may be of relevance for the use of adipose-derived stem cells for tissue engineering.

  7. Waking up dormant tumor suppressor genes with zinc fingers, TALEs and the CRISPR/dCas9 system

    Science.gov (United States)

    Garcia-Bloj, Benjamin; Moses, Colette; Sgro, Agustin; Plani-Lam, Janice; Arooj, Mahira; Duffy, Ciara; Thiruvengadam, Shreyas; Sorolla, Anabel; Rashwan, Rabab; Mancera, Ricardo L.; Leisewitz, Andrea; Swift-Scanlan, Theresa; Corvalan, Alejandro H.; Blancafort, Pilar

    2016-01-01

    The aberrant epigenetic silencing of tumor suppressor genes (TSGs) plays a major role during carcinogenesis and regaining these dormant functions by engineering of sequence-specific epigenome editing tools offers a unique opportunity for targeted therapies. However, effectively normalizing the expression and regaining tumor suppressive functions of silenced TSGs by artificial transcription factors (ATFs) still remains a major challenge. Herein we describe novel combinatorial strategies for the potent reactivation of two class II TSGs, MASPIN and REPRIMO, in cell lines with varying epigenetic states, using the CRISPR/dCas9 associated system linked to a panel of effector domains (VP64, p300, VPR and SAM complex), as well as with protein-based ATFs, Zinc Fingers and TALEs. We found that co-delivery of multiple effector domains using a combination of CRISPR/dCas9 and TALEs or SAM complex maximized activation in highly methylated promoters. In particular, CRISPR/dCas9 VPR with SAM upregulated MASPIN mRNA (22,145-fold change) in H157 lung cancer cells, with accompanying re-expression of MASPIN protein, which led to a concomitant inhibition of cell proliferation and induction of apoptotic cell death. Consistently, CRISPR/dCas9 VP64 with SAM upregulated REPRIMO (680-fold change), which led to phenotypic reprogramming in AGS gastric cancer cells. Altogether, our results outlined novel sequence-specific, combinatorial epigenome editing approaches to reactivate highly methylated TSGs as a promising therapy for cancer and other diseases. PMID:27528034

  8. A homozygous mutation in a novel zinc-finger protein, ERIS, is responsible for Wolfram syndrome 2.

    Science.gov (United States)

    Amr, Sami; Heisey, Cindy; Zhang, Min; Xia, Xia-Juan; Shows, Kathryn H; Ajlouni, Kamel; Pandya, Arti; Satin, Leslie S; El-Shanti, Hatem; Shiang, Rita

    2007-10-01

    A single missense mutation was identified in a novel, highly conserved zinc-finger gene, ZCD2, in three consanguineous families of Jordanian descent with Wolfram syndrome (WFS). It had been shown that these families did not have mutations in the WFS1 gene (WFS1) but were mapped to the WFS2 locus at 4q22-25. A G-->C transversion at nucleotide 109 predicts an amino acid change from glutamic acid to glutamine (E37Q). Although the amino acid is conserved and the mutation is nonsynonymous, the pathogenesis for the disorder is because the mutation also causes aberrant splicing. The mutation was found to disrupt messenger RNA splicing by eliminating exon 2, and it results in the introduction of a premature stop codon. Mutations in WFS1 have also been found to cause low-frequency nonsyndromic hearing loss, progressive hearing loss, and isolated optic atrophy associated with hearing loss. Screening of 377 probands with hearing loss did not identify mutations in the WFS2 gene. The WFS1-encoded protein, Wolframin, is known to localize to the endoplasmic reticulum and plays a role in calcium homeostasis. The ZCD2-encoded protein, ERIS (endoplasmic reticulum intermembrane small protein), is also shown to localize to the endoplasmic reticulum but does not interact directly with Wolframin. Lymphoblastoid cells from affected individuals show a significantly greater rise in intracellular calcium when stimulated with thapsigargin, compared with controls, although no difference was observed in resting concentrations of intracellular calcium.

  9. Expression Pattern Analysis of Zinc Finger Protein Genes in Wheat (Triticum aestivum L.) Under Phosphorus Deprivation

    Institute of Scientific and Technical Information of China (English)

    LI Xiao-juan; GUO Cheng-jin; LU Wen-jing; DUAN Wei-wei; ZHAO Miao; MA Chun-ying; GU Jun-tao; XIAO Kai

    2014-01-01

    Zinc ifnger protein (ZFP) genes comprise a large and diverse gene family, and are involved in biotic and abiotic stress responses in plants. In this study, a total of 126 ZFP genes classiifed into various types in wheat were characterized and subjected to expression pattern analysis under inorganic phosphate (Pi) deprivation. The wheat ZFP genes and their corresponding GenBank numbers were obtained from the information of a 4×44K wheat gene expression microarray chip. They were conifrmed by sequence similarity analysis and named based on their homologs in Brachypodium distachyon or Oriza sativa. Expression analysis based on the microarray chip revealed that these ZFP genes are categorized into 11 classes according to their gene expression patterns in a 24-h of Pi deprivation regime. Among them, ten genes were differentially up-regulated, ten genes differentially down-regulated, and two genes both differentially up-and down-regulated by Pi deprivation. The differentially up-or down-regulated genes exhibited signiifcantly more or less transcripts at one, two, or all of the checking time points (1, 6, and 24 h) of Pi stress in comparison with those of normal growth, respectively. The both differentially up-and down-regulated genes exhibited contrasting expression patterns, of these, TaWRKY70;5 showed significantly up-regulated at 1 and 6 h and down-regulated at 24 h whereas TaAN1AN20-8;2 displayed signiifcantly upregulated at 1 h and downregulated at 6 h under deprivation Pi condition. Real time PCR analysis conifrmed the expression patterns of the differentially expressed genes obtained by the microarray chip. Our results indicate that numerous ZFP genes in wheat respond to Pi deprivation and have provided further insight into the molecular basis that plants respond to Pi deprivation mediated by the ZFP gene family.

  10. The rice B-box zinc finger gene family: genomic identification, characterization, expression profiling and diurnal analysis.

    Directory of Open Access Journals (Sweden)

    Jianyan Huang

    Full Text Available BACKGROUND: The B-box (BBX -containing proteins are a class of zinc finger proteins that contain one or two B-box domains and play important roles in plant growth and development. The Arabidopsis BBX gene family has recently been re-identified and renamed. However, there has not been a genome-wide survey of the rice BBX (OsBBX gene family until now. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we identified 30 rice BBX genes through a comprehensive bioinformatics analysis. Each gene was assigned a uniform nomenclature. We described the chromosome localizations, gene structures, protein domains, phylogenetic relationship, whole life-cycle expression profile and diurnal expression patterns of the OsBBX family members. Based on the phylogeny and domain constitution, the OsBBX gene family was classified into five subfamilies. The gene duplication analysis revealed that only chromosomal segmental duplication contributed to the expansion of the OsBBX gene family. The expression profile of the OsBBX genes was analyzed by Affymetrix GeneChip microarrays throughout the entire life-cycle of rice cultivar Zhenshan 97 (ZS97. In addition, microarray analysis was performed to obtain the expression patterns of these genes under light/dark conditions and after three phytohormone treatments. This analysis revealed that the expression patterns of the OsBBX genes could be classified into eight groups. Eight genes were regulated under the light/dark treatments, and eleven genes showed differential expression under at least one phytohormone treatment. Moreover, we verified the diurnal expression of the OsBBX genes using the data obtained from the Diurnal Project and qPCR analysis, and the results indicated that many of these genes had a diurnal expression pattern. CONCLUSIONS/SIGNIFICANCE: The combination of the genome-wide identification and the expression and diurnal analysis of the OsBBX gene family should facilitate additional functional studies of the Os

  11. Association of a genetic variant of the ZPR1 zinc finger gene with type 2 diabetes mellitus.

    Science.gov (United States)

    Tokoro, Fumitaka; Matsuoka, Reiko; Abe, Shintaro; Arai, Masazumi; Noda, Toshiyuki; Watanabe, Sachiro; Horibe, Hideki; Fujimaki, Tetsuo; Oguri, Mitsutoshi; Kato, Kimihiko; Minatoguchi, Shinya; Yamada, Yoshiji

    2015-01-01

    Various loci and genes that confer susceptibility to coronary heart disease (CHD) have been identified in Caucasian populations by genome-wide association studies (GWASs). As type 2 diabetes mellitus (DM) is an important risk factor for CHD, we hypothesized that certain polymorphisms may contribute to the genetic susceptibility to CHD through affecting the susceptibility to type 2 DM. The purpose of the present study was to examine a possible association of type 2 DM in Japanese individuals with 29 polymorphisms identified as susceptibility loci for CHD by meta-analyses of the GWASs. The study subjects comprised of 3,757 individuals (1,444 subjects with type 2 DM and 2,313 controls). The polymorphism genotypes were determined by the multiplex bead-based Luminex assay, which combines the polymerase chain reaction and sequence-specific oligonucleotide probes with suspension array technology. To compensate for multiple comparisons of genotypes, the criterion of a false discovery rate (FDR) ≤0.05 was adopted for testing the statistical significance of the association. The comparisons of allele frequencies by the χ(2) test revealed that the rs964184 (C→G) of the ZPR1 zinc finger gene (ZPR1) was significantly associated (P=0.0017; FDR=0.050) with type 2 DM. Multivariable logistic regression analysis with adjustment for age, gender and body mass index revealed that rs964184 of ZPR1 was significantly associated (P=0.0012; odds ratio, 1.25; dominant model) with type 2 DM with the minor G allele representing a risk factor for this condition. Fasting plasma glucose levels (P=0.0076) and blood glycosylated hemoglobin contents (P=0.0132) significantly differed among ZPR1 genotypes with the G allele associated with increases in these parameters. ZPR1 may thus be a susceptibility locus for type 2 DM in Japanese individuals.

  12. The ZNF75 zinc finger gene subfamily: Isolation and mapping of the four members in humans and great apes

    Energy Technology Data Exchange (ETDEWEB)

    Villa, A.; Strina, D.; Frattini, A. [Consiglio Nazionale delle Ricerche, Milan (Italy)] [and others

    1996-07-15

    We have previously reported the characterization of the human ZNF75 gene located on Xq26, which has only limited homology (less than 65%) to other ZF genes in the databases. Here, we describe three human zinc finger genes with 86 to 95% homology to ZNF75 at the nucleotide level, which represent all the members of the human ZNF75 subfamily. One of these, ZNF75B, is a pseudogene mapped to chromosome 12q13. The other two, ZNF75A and ZNF75C, maintain on ORF in the sequenced region, and at least the latter is expressed in the U937 cell line. They were mapped to chromosomes 16 and 11, respectively. All these genes are conserved in chimpanzees, gorillas, and orangutans. The ZNF75B homologue is a pseudogene in all three great apes, and in chimpanzee it is located on chromosome 10 (phylogenetic XII), at p13 (corresponding to the human 12q13). The chimpanzee homologue of ZNF75 is also located on the Xq26 chromosome, in the same region, as detected by in situ hybridization. As expected, nucleotide changes were clearly more abundant between human and organutan than between human and chimpanzee or gorilla homologues. Members of the same class were more similar to each other than to the other homologues within the same species. This suggests that the duplication and/or retrotranscription events occurred in a common ancestor long before great ape speciation. This, together with the existance of at least two genes in cows and horses, suggests a relatively high conservation of this gene family. 20 refs., 5 figs., 1 tab.

  13. A C2H2 zinc finger protein FEMU2 is required for fox1 expression in Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Xiaodong Deng

    Full Text Available Chlamydomonas reinhardtii fox1 gene encodes a ferroxidase that is involved in cellular Fe uptake and highly induced during Fe deficient conditions. In an effort to identify fox1 promoter regulatory elements, an insertional library was generated in a transgenic Chlamydomonas strain (2A38 harboring an arylsulfatase (ARS reporter gene driven by the fox1 promoter. Mutants with a defective response to low iron conditions were selected for further study. Among these, a strain containing a disrupted femu2 gene was identified. Activation of the fox1 promoter by the femu2 gene product was confirmed by silencing the femu2 gene using RNA interference. In three femu2 RNAi transgenic lines (IR3, IR6, and IR7, ARS reporter gene activities declined by 84.3%, 86.4%, and 88.8%, respectively under Fe deficient conditions. Furthermore, RT-PCR analysis of both the femu2 mutant and the RNAi transgenic lines showed significantly decreased transcript abundance of the endogenous fox1 gene under Fe deficient conditions. Amino acid sequence analysis of the femu2 gene product identified three potential C2H2 zinc finger (ZF motifs and a nuclear localization study suggests that FEMU2 is localized to the nucleus. In addition, a potential FEMU2 binding site ((G/TTTGG(G/T(G/TT was identified using PCR-mediated random binding site selection. Taken together, this evidence suggests that FEMU2 is involved in up-regulation of the fox1 gene in Fe deficient cells.

  14. Transient Expression of Fez Family Zinc Finger 2 Protein Regulates the Brn3b Gene in Developing Retinal Ganglion Cells.

    Science.gov (United States)

    Qu, Chunsheng; Bian, Dandan; Li, Xue; Xiao, Jian; Wu, Chunping; Li, Yue; Jiang, Tian; Zhou, Xiangtian; Qu, Jia; Chen, Jie-Guang

    2016-04-01

    Retinal ganglion cells (RGCs) are projection neurons in the neural retina that relay visual information from the environment to the central nervous system. The early expression of MATH5 endows the post-mitotic precursors with RGC competence and leads to the activation ofBrn3bthat marks committed RGCs. Nevertheless, this fate commitment process and, specifically, regulation ofBrn3bremain elusive. To explore the molecular mechanisms underlying RGC generation in the mouse retina, we analyzed the expression and function of Fez family zinc finger 2 (FEZF2), a transcription factor critical for the development of projection neurons in the cerebral cortex.Fezf2mRNA and protein were transiently expressed at embryonic day 16.5 in the inner neuroblast layer and the prospective ganglion cell layer of the retina, respectively. Knockout ofFezf2in the developing retina reduced BRN3B+ cells and increased apoptotic cell markers.Fezf2knockdown by retinalin uteroelectroporation diminished BRN3B but not the coexpressed ISLET1 and BRN3A, indicating that the BRN3B decrease was the cause, not the result, of the overall reduction of BRN3B+ RGCs in theFezf2knockout retina. Moreover, the mRNA and promoter activity ofBrn3bwere increasedin vitroby FEZF2, which bound to a 5' regulatory fragment in theBrn3bgenomic locus. These results indicate that transient expression ofFezf2in the retina modulates the transcription ofBrn3band the survival of RGCs. This study improves our understanding of the transcriptional cascade required for the specification of RGCs and provides novel insights into the molecular basis of retinal development.

  15. Induction of stable drug resistance in human breast cancer cells using a combinatorial zinc finger transcription factor library.

    Directory of Open Access Journals (Sweden)

    Jeongeun Lee

    Full Text Available Combinatorial libraries of artificial zinc-finger transcription factors (ZF-TFs provide a robust tool for inducing and understanding various functional components of the cancer phenotype. Herein, we utilized combinatorial ZF-TF library technology to better understand how breast cancer cells acquire resistance to fulvestrant, a clinically important anti-endocrine therapeutic agent. From a diverse collection of nearly 400,000 different ZF-TFs, we isolated six ZF-TF library members capable of inducing stable, long-term anti-endocrine drug-resistance in two independent estrogen receptor-positive breast cancer cell lines. Comparative gene expression profile analysis of the six different ZF-TF-transduced breast cancer cell lines revealed five distinct clusters of differentially expressed genes. One cluster was shared among all 6 ZF-TF-transduced cell lines and therefore constituted a common fulvestrant-resistant gene expression signature. Pathway enrichment-analysis of this common fulvestrant resistant signature also revealed significant overlap with gene sets associated with an estrogen receptor-negative-like state and with gene sets associated with drug resistance to different classes of breast cancer anti-endocrine therapeutic agents. Enrichment-analysis of the four remaining unique gene clusters revealed overlap with myb-regulated genes. Finally, we also demonstrated that the common fulvestrant-resistant signature is associated with poor prognosis by interrogating five independent, publicly available human breast cancer gene expression datasets. Our results demonstrate that artificial ZF-TF libraries can be used successfully to induce stable drug-resistance in human cancer cell lines and to identify a gene expression signature that is associated with a clinically relevant drug-resistance phenotype.

  16. Integrative analysis of the zinc finger transcription factor Lame duck in the Drosophila myogenic gene regulatory network.

    Science.gov (United States)

    Busser, Brian W; Huang, Di; Rogacki, Kevin R; Lane, Elizabeth A; Shokri, Leila; Ni, Ting; Gamble, Caitlin E; Gisselbrecht, Stephen S; Zhu, Jun; Bulyk, Martha L; Ovcharenko, Ivan; Michelson, Alan M

    2012-12-11

    Contemporary high-throughput technologies permit the rapid identification of transcription factor (TF) target genes on a genome-wide scale, yet the functional significance of TFs requires knowledge of target gene expression patterns, cooperating TFs, and cis-regulatory element (CRE) structures. Here we investigated the myogenic regulatory network downstream of the Drosophila zinc finger TF Lame duck (Lmd) by combining both previously published and newly performed genomic data sets, including ChIP sequencing (ChIP-seq), genome-wide mRNA profiling, cell-specific expression patterns of putative transcriptional targets, analysis of histone mark signatures, studies of TF cooccupancy by additional mesodermal regulators, TF binding site determination using protein binding microarrays (PBMs), and machine learning of candidate CRE motif compositions. Our findings suggest that Lmd orchestrates an extensive myogenic regulatory network, a conclusion supported by the identification of Lmd-dependent genes, histone signatures of Lmd-bound genomic regions, and the relationship of these features to cell-specific gene expression patterns. The heterogeneous cooccupancy of Lmd-bound regions with additional mesodermal regulators revealed that different transcriptional inputs are used to mediate similar myogenic gene expression patterns. Machine learning further demonstrated diverse combinatorial motif patterns within tissue-specific Lmd-bound regions. PBM analysis established the complete spectrum of Lmd DNA binding specificities, and site-directed mutagenesis of Lmd and additional newly discovered motifs in known enhancers demonstrated the critical role of these TF binding sites in supporting full enhancer activity. Collectively, these findings provide insights into the transcriptional codes regulating muscle gene expression and offer a generalizable approach for similar studies in other systems.

  17. The zinc finger transcription factor SlZFP2 negatively regulates abscisic acid biosynthesis and fruit ripening in tomato.

    Science.gov (United States)

    Weng, Lin; Zhao, Fangfang; Li, Rong; Xu, Changjie; Chen, Kunsong; Xiao, Han

    2015-03-01

    Abscisic acid (ABA) regulates plant development and adaptation to environmental conditions. Although the ABA biosynthesis pathway in plants has been thoroughly elucidated, how ABA biosynthetic genes are regulated at the molecular level during plant development is less well understood. Here, we show that the tomato (Solanum lycopersicum) zinc finger transcription factor SlZFP2 is involved in the regulation of ABA biosynthesis during fruit development. Overexpression of SlZFP2 resulted in multiple phenotypic changes, including more branches, early flowering, delayed fruit ripening, lighter seeds, and faster seed germination, whereas down-regulation of its expression caused problematic fruit set, accelerated ripening, and inhibited seed germination. SlZFP2 represses ABA biosynthesis during fruit development through direct suppression of the ABA biosynthetic genes NOTABILIS, SITIENS, and FLACCA and the aldehyde oxidase SlAO1. We also show that SlZFP2 regulates fruit ripening through transcriptional suppression of the ripening regulator COLORLESS NON-RIPENING. Using bacterial one-hybrid screening and a selected amplification and binding assay, we identified the (A/T)(G/C)TT motif as the core binding sequence of SlZFP2. Furthermore, by RNA sequencing profiling, we found that 193 genes containing the SlZFP2-binding motifs in their promoters were differentially expressed in 2 d post anthesis fruits between the SlZFP2 RNA interference line and its nontransgenic sibling. We propose that SlZFP2 functions as a repressor to fine-tune ABA biosynthesis during fruit development and provides a potentially valuable tool for dissecting the role of ABA in fruit ripening.

  18. The role of Zic family zinc finger transcription factors in the proliferation and differentiation of retinal progenitor cells

    Energy Technology Data Exchange (ETDEWEB)

    Watabe, Yui [Division of Molecular and Developmental Biology, Institute of Medical Science, University of Tokyo (Japan); Department of Ophthalmology, Teikyo University School of Medicine, Tokyo (Japan); Division of Orthoptics, Teikyo University School of Medical Care and Technology, Tokyo (Japan); Baba, Yukihiro [Division of Molecular and Developmental Biology, Institute of Medical Science, University of Tokyo (Japan); Nakauchi, Hiromitsu [Division of Stem Cell Therapy, Center for Stem Cell Biology and Regenerative Medicine, Institute of Medical Science, University of Tokyo (Japan); Mizota, Atsushi [Department of Ophthalmology, Teikyo University School of Medicine, Tokyo (Japan); Watanabe, Sumiko, E-mail: sumiko@ims.u-tokyo.ac.jp [Division of Molecular and Developmental Biology, Institute of Medical Science, University of Tokyo (Japan)

    2011-11-11

    Highlights: Black-Right-Pointing-Pointer Zic transcription factors expressed early retinal progenitor cells. Black-Right-Pointing-Pointer Zics sustain proliferation activity of retinal progenitor cells. Black-Right-Pointing-Pointer Overexpression of Zic in retinal progenitors perturbed rod differentiation. Black-Right-Pointing-Pointer Fate determination to rod photoreceptor was not affected. -- Abstract: Members of the Zic family of zinc finger transcription factors play critical roles in a variety of developmental processes. Using DNA microarray analysis, we found that Zics are strongly expressed in SSEA-1-positive early retinal progenitors in the peripheral region of the mouse retina. Reverse-transcription polymerase chain reaction using mRNA from the retina at various developmental stages showed that Zic1 and Zic2 are expressed in the embryonic retina and then gradually disappear during retinal development. Zic3 is also expressed in the embryonic retina; its expression level slightly decreases but it is expressed until adulthood. We overexpressed Zic1, Zic2, or Zic3 in retinal progenitors at embryonic day 17.5 and cultured the retina as explants for 2 weeks. The number of rod photoreceptors was fewer than in the control, but no other cell types showed significant differences between control and Zic overexpressing cells. The proliferation activity of normal retinal progenitors decreased after 5 days in culture, as observed in normal in vivo developmental processes. However, Zic expressing retinal cells continued to proliferate at days 5 and 7, suggesting that Zics sustain the proliferation activities of retinal progenitor cells. Since the effects of Zic1, 2, and 3 are indistinguishable in terms of differentiation and proliferation of retinal progenitors, the redundant function of Zics in retinal development is suggested.

  19. Probing isoform-specific functions of polypeptide GalNAc-transferases using zinc finger nuclease glycoengineered SimpleCells.

    Science.gov (United States)

    Schjoldager, Katrine T-B G; Vakhrushev, Sergey Y; Kong, Yun; Steentoft, Catharina; Nudelman, Aaron S; Pedersen, Nis B; Wandall, Hans H; Mandel, Ulla; Bennett, Eric P; Levery, Steven B; Clausen, Henrik

    2012-06-19

    Our knowledge of the O-glycoproteome [N-acetylgalactosamine (GalNAc) type] is highly limited. The O-glycoproteome is differentially regulated in cells by dynamic expression of a subset of 20 polypeptide GalNAc-transferases (GalNAc-Ts), and methods to identify important functions of individual GalNAc-Ts are largely unavailable. We recently introduced SimpleCells, i.e., human cell lines made deficient in O-glycan extension by zinc finger nuclease targeting of a key gene in O-glycan elongation (Cosmc), which allows for proteome-wide discovery of O-glycoproteins. Here we have extended the SimpleCell concept to include proteome-wide discovery of unique functions of individual GalNAc-Ts. We used the GalNAc-T2 isoform implicated in dyslipidemia and the human HepG2 liver cell line to demonstrate unique functions of this isoform. We confirm that GalNAc-T2-directed site-specific O-glycosylation inhibits proprotein activation of the lipase inhibitor ANGPTL3 in HepG2 cells and further identify eight O-glycoproteins exclusively glycosylated by T2 of which one, ApoC-III, is implicated in dyslipidemia. Our study supports an essential role for GalNAc-T2 in lipid metabolism, provides serum biomarkers for GalNAc-T2 enzyme function, and validates the use of GALNT gene targeting with SimpleCells for broad discovery of disease-causing deficiencies in O-glycosylation. The presented glycoengineering strategy opens the way for proteome-wide discovery of functions of GalNAc-T isoforms and their role in congenital diseases and disorders.

  20. A classic zinc finger from friend of GATA mediates an interaction with the coiled-coil of transforming acidic coiled-coil 3.

    Science.gov (United States)

    Simpson, Raina J Y; Yi Lee, Stella Hoi; Bartle, Natalie; Sum, Eleanor Y; Visvader, Jane E; Matthews, Jacqueline M; Mackay, Joel P; Crossley, Merlin

    2004-09-17

    Classic zinc finger domains (cZFs) consist of a beta-hairpin followed by an alpha-helix. They are among the most abundant of all protein domains and are often found in tandem arrays in DNA-binding proteins, with each finger contributing an alpha-helix to effect sequence-specific DNA recognition. Lone cZFs, not found in tandem arrays, have been postulated to function in protein interactions. We have studied the transcriptional co-regulator Friend of GATA (FOG), which contains nine zinc fingers. We have discovered that the third cZF of FOG contacts a coiled-coil domain in the centrosomal protein transforming acidic coiled-coil 3 (TACC3). Although FOG-ZF3 exhibited low solubility, we have used a combination of mutational mapping and protein engineering to generate a derivative that was suitable for in vitro and structural analysis. We report that the alpha-helix of FOG-ZF3 recognizes a C-terminal portion of the TACC3 coiled-coil. Remarkably, the alpha-helical surface utilized by FOG-ZF3 is the same surface responsible for the well established sequence-specific DNA-binding properties of many other cZFs. Our data demonstrate the versatility of cZFs and have implications for the analysis of many as yet uncharacterized cZF proteins.

  1. UtroUp is a novel six zinc finger artificial transcription factor that recognises 18 base pairs of the utrophin promoter and efficiently drives utrophin upregulation

    Directory of Open Access Journals (Sweden)

    Onori Annalisa

    2013-01-01

    Full Text Available Abstract Background Duchenne muscular dystrophy (DMD is the most common X-linked muscle degenerative disease and it is due to the absence of the cytoskeletal protein dystrophin. Currently there is no effective treatment for DMD. Among the different strategies for achieving a functional recovery of the dystrophic muscle, the upregulation of the dystrophin-related gene utrophin is becoming more and more feasible. Results We have previously shown that the zinc finger-based artificial transcriptional factor “Jazz” corrects the dystrophic pathology in mdx mice by upregulating utrophin gene expression. Here we describe a novel artificial transcription factor, named “UtroUp”, engineered to further improve the DNA-binding specificity. UtroUp has been designed to recognise an extended DNA target sequence on both the human and mouse utrophin gene promoters. The UtroUp DNA-binding domain contains six zinc finger motifs in tandem, which is able to recognise an 18-base-pair DNA target sequence that statistically is present only once in the human genome. To achieve a higher transcriptional activation, we coupled the UtroUp DNA-binding domain with the innovative transcriptional activation domain, which was derived from the multivalent adaptor protein Che-1/AATF. We show that the artificial transcription factor UtroUp, due to its six zinc finger tandem motif, possesses a low dissociation constant that is consistent with a strong affinity/specificity toward its DNA-binding site. When expressed in mammalian cell lines, UtroUp promotes utrophin transcription and efficiently accesses active chromatin promoting accumulation of the acetylated form of histone H3 in the utrophin promoter locus. Conclusions This novel artificial molecule may represent an improved platform for the development of future applications in DMD treatment.

  2. 锌指核酸酶在基因组编辑中的应用%Application of Zinc Finger Nuclease in Genome Editing

    Institute of Scientific and Technical Information of China (English)

    沈忠福

    2012-01-01

    基因组编辑是建立在基因靶向修饰的基础上,对生物基因组进行改造的一项新技术.锌指核酸酶(zinc finger nucleases,ZFN)是一种特异性强、应用范围广的人工合成酶,可在靶位点制造DNA双链切口(double-strand break,DSB)进而诱导细胞内源性修复机制,通过同源重组修复(homology-directed repair,HDR)或非同源末端连接(non-homologous end joining,NHEJ)途径实现基因替换和纠正,因而成为基因组编辑的良好工具.本文将就最近文献中提到的锌指核酸酶在基因组编辑中的应用做一综述.%Genome editing is a new technology for biological genome transformation based on gene targeting modification. Zinc finger nucleases can induce a double-strand break (DSB) in a specific genomic target sequence, followed by the generation of desired modifications during subsequent DNA break repair, such as replacing or correcting a gene through homology-directed repair (HDR) or non-homologous end joining (NHEJ) pathway, which makes it the excellent tool in genome editing. Thus, I will make a review of application of zinc finger nuclease in genome editing in this article.

  3. High-efficiency genome editing via 2A-coupled co-expression of fluorescent proteins and zinc finger nucleases or CRISPR/Cas9 nickase pairs

    DEFF Research Database (Denmark)

    Duda, Katarzyna; Lonowski, Lindsey A; Kofoed-Nielsen, Michael

    2014-01-01

    Targeted endonucleases including zinc finger nucleases (ZFNs) and clustered regularly interspaced short palindromic repeats (CRISPRs)/Cas9 are increasingly being used for genome editing in higher species. We therefore devised a broadly applicable and versatile method for increasing editing...... were minimal, and when occurring, our data suggest that they may be counteracted by selecting intermediate nuclease levels where off-target mutagenesis is low, but on-target mutagenesis remains relatively high. The method was also applicable to the CRISPR/Cas9 system, including CRISPR/Cas9 mutant...

  4. Isolation and gene expression analysis of Arabidopsis thaliana mutants with constitutive expression of ATL2, an early elicitor-response RING-H2 zinc-finger gene.

    OpenAIRE

    2004-01-01

    Genes with unstable transcripts often encode proteins that play important regulatory roles. ATL2 is a member of a multigene family coding highly related RING-H2 zinc-finger proteins that may function as E3 ubiquitin ligases. ATL2 mRNA accumulation occurs rapidly and transiently after incubation with elicitors of pathogen response. We screened 50,000 M(2) families from a line that carries a fusion of pATL2 to the GUS reporter gene and isolated five mutants, which we named eca (expresión consti...

  5. The LSD1-Type Zinc Finger Motifs of Pisum sativa LSD1 Are a Novel Nuclear Localization Signal and Interact with Importin Alpha

    OpenAIRE

    Shanping He; Kuowei Huang; Xu Zhang; Xiangchun Yu; Ping Huang; Chengcai An

    2011-01-01

    BACKGROUND: Genetic studies of the Arabidopsis mutant lsd1 highlight the important role of LSD1 in the negative regulation of plant programmed cell death (PCD). Arabidopsis thaliana LSD1 (AtLSD1) contains three LSD1-type zinc finger motifs, which are involved in the protein-protein interaction. METHODOLOGY/PRINCIPAL FINDINGS: To further understand the function of LSD1, we have analyzed cellular localization and functional localization domains of Pisum sativa LSD1 (PsLSD1), which is a homolog ...

  6. In Vivo Zinc Finger Nuclease-mediated Targeted Integration of a Glucose-6-phosphatase Transgene Promotes Survival in Mice With Glycogen Storage Disease Type IA.

    Science.gov (United States)

    Landau, Dustin J; Brooks, Elizabeth Drake; Perez-Pinera, Pablo; Amarasekara, Hiruni; Mefferd, Adam; Li, Songtao; Bird, Andrew; Gersbach, Charles A; Koeberl, Dwight D

    2016-04-01

    Glycogen storage disease type Ia (GSD Ia) is caused by glucose-6-phosphatase (G6Pase) deficiency in association with severe, life-threatening hypoglycemia that necessitates lifelong dietary therapy. Here we show that use of a zinc-finger nuclease (ZFN) targeted to the ROSA26 safe harbor locus and a ROSA26-targeting vector containing a G6PC donor transgene, both delivered with adeno-associated virus (AAV) vectors, markedly improved survival of G6Pase knockout (G6Pase-KO) mice compared with mice receiving the donor vector alone (P Ia, as compared with normal littermates, at 8 months following vector administration (P Ia.

  7. A conserved function of the zinc finger transcription factor Sp8/9 in allometric appendage growth in the milkweed bug Oncopeltus fasciatus

    OpenAIRE

    Schaeper, Nina; Prpic, Nikola-Michael; Wimmer, Ernst

    2009-01-01

    The genes encoding the closely related zinc finger transcription factors Buttonhead (Btd) and D-Sp1 are expressed in the developing limb primordia of Drosophila melanogaster and are required for normal growth of the legs. The D-Sp1 homolog of the red flour beetle Tribolium castaneum, Sp8 (appropriately termed Sp8/9), is also required for the proper growth of the leg segments. Here we report on the isolation and functional study of the Sp8/9 gene from the milkweed bug Oncopeltus fasciatus. We ...

  8. NF-κB activation and zinc finger protein A20 expression in mature dendritic cells derived from liver allografts undergoing acute rejection

    Institute of Scientific and Technical Information of China (English)

    Ming-Qing Xu; Wei Wang; Lan Xue; Lv-Nan Yan

    2003-01-01

    AIM: To investigate the role of NF-κB activation and zinc finger protein A20 expression in the regulation of maturation of dendritic cells (DCs) derived from liver allografts undergoing acute rejection. METHODS: Sixty donor male SD rats and sixty recipient male LEW rats weighing 220-300 g were randomly divided into whole liver transplantation group and partial liver transplantation group. Allogeneic (SD rat to LEW rat) whole and 50 % partial liver transplantation were performed. DCs from liver grafts 0 hour and 4 days after transplantation were isolated and propagated in the presence of GM-CSF in vitro. Morphological characteristics and phenotypical features of DCs propagated for 10 days were analyzed by electron microscopy and flow cytometry, respectively. NF-κB binding activity, IL-12p70 protein and zinc finger protein A20expression in these DCs were measured by EMSA and Western blotting, respectively. Histological grading of rejection was determined. RESULTS: Allogeneic whole liver grafts showed no signs of rejection on day 4 after the transplantation. In contrast,allogeneic partial liver grafts demonstrated moderate to severe rejection on day 4 after the transplantation. After propagation for 10 days in the presence of GM-CSF in vitro,DCs from allogeneic whole liver grafts exhibited features of immature DC with absence of CD40 surface expression,these DCs were found to exhibit detectable but very low level of NF-κB activity, IL-12 p70 protein and zinc finger protein A20 expression. Whereas, DCs from allogeneic partial liver graft 4 days after transplantation displayed features of mature DC, with high level of CD40 surface expression, and as a consequence, higher expression of IL-12p70 protein, higher activities of NF-κB and higher expression of zinc finger protein A20 compared with those of DCs from whole liver grafts (P<0.001). CONCLUSION: These results suggest that A20expression is up-regulated in response to NF-κB activation in mature DCs derived from

  9. Gene targeting technologies in rats: zinc finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats.

    Science.gov (United States)

    Mashimo, Tomoji

    2014-01-01

    The laboratory rat has been widely used as an animal model in biomedical science for more than 150 years. Applying zinc-finger nucleases or transcription activator-like effector nucleases to rat embryos via microinjection is an efficient genome editing tool for generating targeted knockout rats. Recently, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated endonucleases have been used as an effective tool for precise and multiplex genome editing in mice and rats. In this review, the advantages and disadvantages of these site-specific nuclease technologies for genetic analysis and manipulation in rats are discussed.

  10. The zinc finger protein Zfr1p is localized specifically to conjugation junction and required for sexual development in Tetrahymena thermophila.

    Directory of Open Access Journals (Sweden)

    Jing Xu

    Full Text Available Conjugation in Tetrahymena thermophila involves a developmental program consisting of three prezygotic nuclear divisions, pronuclear exchange and fusion, and postzygotic and exconjugant stages. The conjugation junction structure appears during the initiation of conjugation development, and disappears during the exconjugant stage. Many structural and functional proteins are involved in the establishment and maintenance of the junction structure in T. thermophila. In the present study, a zinc finger protein-encoding gene ZFR1 was found to be expressed specifically during conjugation and to localize specifically to the conjugation junction region. Truncated Zfr1p localized at the plasma membrane in ordered arrays and decorated Golgi apparatus located adjacent to basal body. The N-terminal zinc finger and C-terminal hydrophobic domains of Zfr1p were found to be required for its specific conjugation junction localization. Conjugation development of ZFR1 somatic knockout cells was aborted at the pronuclear exchange and fusion conjugation stages. Furthermore, Zfr1p was found to be important for conjugation junction stability during the prezygotic nuclear division stage. Taken together, our data reveal that Zfr1p is required for the stability and integrity of the conjugation junction structure and essential for the sexual life cycle of the Tetrahymena cell.

  11. Effect of Zinc Finger Antiviral Protein on Virus%锌指抗病毒蛋白ZAP对病毒的作用

    Institute of Scientific and Technical Information of China (English)

    2015-01-01

    Zinc finger antiviral protein ( ZAP) , a kind of Restriction Factor, can mediate deg⁃radation of viral mRNA, inhibit viral translation and finally suppress viral replication�MMLV、HIV、 EBOV、 MARV、 SINV、 HBV、 HCV could be inhibited by ZAP�This paper mainly re⁃viewed the antiviral ability of ZAP.%锌指抗病毒蛋白( zinc finger antiviral protein, ZAP)是一种具有抗病毒活性的宿主细胞限制因子, ZAP能通过介导病毒mRNA的降解以及抑制翻译,从而抑制病毒的复制。 ZAP能够抑制鼠白血病病毒、人类免疫缺陷病毒、埃博拉病毒、马尔堡病毒、辛德比斯病毒、乙型肝炎病毒、丙型肝炎病毒的复制。本文主要就ZAP的抗病毒作用作一综述。

  12. Overexpression of myeloid zinc finger 1 suppresses matrix metalloproteinase-2 expression and reduces invasiveness of SiHa human cervical cancer cells.

    Science.gov (United States)

    Tsai, Su-Ju; Hwang, Jin-Ming; Hsieh, Shu-Ching; Ying, Tsung-Ho; Hsieh, Yi-Hsien

    2012-08-24

    Myeloid zinc finger 1 (MZF1) gene belongs to the Kruppel family of zinc finger transcription factors. MZF1 has been suggested to play an important role in the tumorigenesis, invasion, and apoptosis of various tumor cells. However, the role of MZF1 in human cervical cancer remains unclear. To investigate the molecular mechanisms of MZF1 and its functional role in human cervical cancer cell migration and invasion, we experimented on stable SiHa cells overexpressing MZF1. We found that MZF1 overexpression inhibits the migratory and invasive abilities of SiHa cervical cancer cells. In addition, the overexpression of MZF1 significantly reduces MMP-2 protein and mRNA levels. Luciferase and ChIP assays suggested that MZF1 directly binds to MMP-2 gene regulatory sequences in vivo and suppresses MMP-2 promoter activity in vitro. This study shows that MZF-1 represses MMP-2 transcription and suggests that this repression may be linked to inhibition of human cervical cancer cell migration and metastasis.

  13. An A20/AN1-type zinc finger protein modulates gibberellins and abscisic acid contents and increases sensitivity to abiotic stress in rice (Oryza sativa).

    Science.gov (United States)

    Zhang, Ye; Lan, Hongxia; Shao, Qiaolin; Wang, Ruqin; Chen, Hui; Tang, Haijuan; Zhang, Hongsheng; Huang, Ji

    2016-01-01

    The plant hormones gibberellins (GA) and abscisic acid (ABA) play important roles in plant development and stress responses. Here we report a novel A20/AN1-type zinc finger protein ZFP185 involved in GA and ABA signaling in the regulation of growth and stress response. ZFP185 was constitutively expressed in various rice tissues. Overexpression of ZFP185 in rice results in a semi-dwarfism phenotype, reduced cell size, and the decrease of endogenous GA3 content. By contrast, higher GA3 content was observed in RNAi plants. The application of exogenous GA3 can fully rescue the semi-dwarfism phenotype of ZFP185 overexpressing plants, suggesting the negative role of ZFP185 in GA biosynthesis. Besides GA, overexpression of ZFP185 decreased ABA content and expression of several ABA biosynthesis-related genes. Moreover, it was found that ZFP185, unlike previously known A20/AN1-type zinc finger genes, increases sensitivity to drought, cold, and salt stresses, implying the negative role of ZFP185 in stress tolerance. ZFP185 was localized in the cytoplasm and lacked transcriptional activation potential. Our study suggests that ZFP185 regulates plant growth and stress responses by affecting GA and ABA biosynthesis in rice.

  14. A novel zinc finger protein Zfp277 mediates transcriptional repression of the Ink4a/arf locus through polycomb repressive complex 1

    DEFF Research Database (Denmark)

    Negishi, Masamitsu; Saraya, Atsunori; Mochizuki, Shinobu

    2010-01-01

    . METHODOLOGY/PRINCIPAL FINDINGS: We examined the function of Zinc finger domain-containing protein 277 (Zfp277), a novel zinc finger protein that interacts with the PcG protein Bmi1. Zfp277 binds to the Ink4a/Arf locus in a Bmi1-independent manner and interacts with polycomb repressor complex (PRC) 1 through......BACKGROUND: Polycomb group (PcG) proteins play a crucial role in cellular senescence as key transcriptional regulators of the Ink4a/Arf tumor suppressor gene locus. However, how PcG complexes target and contribute to stable gene silencing of the Ink4a/Arf locus remains little understood...... direct interaction with Bmi1. Loss of Zfp277 in mouse embryonic fibroblasts (MEFs) caused dissociation of PcG proteins from the Ink4a/Arf locus, resulting in premature senescence associated with derepressed p16(Ink4a) and p19(Arf) expression. Levels of both Zfp277 and PcG proteins inversely correlated...

  15. Zinc finger-inspired nanohydrogels with glutathione/pH triggered degradation based on coordination substitution for highly efficient delivery of anti-cancer drugs.

    Science.gov (United States)

    Zhang, Zihao; Wan, Jiaxun; Sun, Luyan; Li, Yongjing; Guo, Jia; Wang, Changchun

    2016-03-10

    Biodegradable materials used for drug delivery are of great demand due to their ability to degrade into low molecular weight species and further excrete from the body by metabolism. Herein, we report a new kind of zinc(II) crosslinked poly(methacrylic acid) nanohydrogels (ZCLNs) inspired by zinc finger proteins with dual stimuli-triggered degradation (glutathione and pH) for the first time. Compared with the disulfide bond crosslinked nanohydrogels, this new kind of ZCLNs is beneficial to the degradation of a wide range of cells, including normal cells. Ex vivo fluorescence images showed that the DOX-loaded folate-PEG conjugated zinc(II)-crosslinked polymeric nanohydrogels (FPZCLNs-15) preferentially accumulated in tumor tissue and the accumulation in normal tissues was much less compared with DOX-loaded PZCLNs-15 (non-targeted nanohydrogels) and free DOX, the FPZCLNs-15 (targeting system) delivered DOX to the tumor site with approximately 3.6- and 1.6-fold higher than free DOX and PZCLNs-15, respectively. Meanwhile, the PZCLNs-15 and FPZCLNs-15 reduced the concentration of DOX in the heart by 3.2- and 5.0-fold respectively, as compared to the free DOX. Moreover, a superior tumor growth inhibition and negligible damage to normal organs like the heart and kidney, which is reported to be vulnerable to DOX-associated side effects, are further demonstrated.

  16. Genetic analysis of Kruppel-like zinc finger 11 variants in 5864 Danish individuals: potential effect on insulin resistance and modified signal transducer and activator of transcription-3 binding by promoter variant -1659G>C

    DEFF Research Database (Denmark)

    Gutiérrez-Aguilar, Ruth; Froguel, Philippe; Hamid, Yasmin H

    2008-01-01

    CONTEXT: The transcription factor Krüppel-like zinc finger 11 (KLF11) has been suggested to contribute to genetic risk of type 2 diabetes (T2D). Our previous results showed that four KLF11 variants, in strong linkage disequilibrium (LD block including +185 A>G/Gln62Arg and -1659 G>C) were...

  17. The human zinc-finger protein-7 gene is located 90 kb 3' of MYC and is not expressed in Burkitt lymphoma cell lines.

    Science.gov (United States)

    Feduchi, E; Gallego, M I; Lazo, P A

    1994-09-15

    The zinc-finger gene-7 (ZNF7) was located 90 kb 3' of MYC on human chromosome 8 band q24 by pulsed-field gel electrophoresis (PFGE). This position lies between the MLV14 and BVR1 loci, 2 variant translocation breakpoints in Burkitt lymphomas. The structure of the ZNF7 gene was not altered by translocations in Burkitt-lymphoma cell lines as shown by its germline-restriction map configuration. The chromosomal region surrounding the ZNF7 gene was extensively methylated. The ZNF7 gene was not expressed in 19 BL cell lines. Expression was detected only in the BL41 and BL47 cell lines and in the SW756 cervical-carcinoma cell line. The RNA in each was of a different size. We postulate that the lack of ZNF7 expression in Burkitt lymphomas might contribute to the tumor phenotype.

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

    Energy Technology Data Exchange (ETDEWEB)

    Phillips, Carolyn M.; Dernburg, Abby F.

    2006-06-07

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

  19. Activating human genes with zinc finger proteins, transcription activator-like effectors and CRISPR/Cas9 for gene therapy and regenerative medicine.

    Science.gov (United States)

    Gersbach, Charles A; Perez-Pinera, Pablo

    2014-08-01

    New technologies have recently been developed to control the expression of human genes in their native genomic context by engineering synthetic transcription factors that can be targeted to any DNA sequence. The ability to precisely regulate any gene as it occurs naturally in the genome provides a means to address a variety of diseases and disorders. This approach also circumvents some of the traditional challenges of gene therapy. In this editorial, we review the technologies that have enabled targeted human gene activation, including the engineering of transcription factors based on zinc finger proteins, transcription activator-like effectors and the CRISPR/Cas9 system. Additionally, we highlight examples in which these methods have been developed for therapeutic applications and discuss challenges and opportunities.

  20. A Comprehensive Catalog of Human KRAB-associated Zinc Finger Genes: Insights into the Evolutionary History of a Large Family of Transcriptional Repressors

    Energy Technology Data Exchange (ETDEWEB)

    Huntley, S; Baggott, D M; Hamilton, A T; Tran-Gyamfi, M; Yang, S; Kim, J; Gordon, L; Branscomb, E; Stubbs, L

    2005-09-30

    Krueppel-type zinc finger (ZNF) motifs are prevalent components of transcription factor proteins in all eukaryotic species. In mammals, most ZNF proteins comprise a single class of transcriptional repressors in which a chromatin interaction domain, called the Krueppel-associated box (KRAB) is attached to a tandem array of DNA-binding zinc-finger motifs. KRAB-ZNF loci are specific to tetrapod vertebrates, but have expanded dramatically in numbers through repeated rounds of segmental duplication to create a gene family with hundreds of members in mammals. To define the full repertoire of human KRAB-ZNF proteins, we searched the human genome for key motifs and used them to construct and manually curate gene models. The resulting KRAB-ZNF gene catalog includes 326 known genes, 243 of which were structurally corrected by manual annotation, and 97 novel KRAB-ZNF genes; this single family therefore comprises 20% of all predicted human transcription factor genes. Many of the genes are alternatively spliced, yielding a total of 743 distinct predicted proteins. Although many human KRAB-ZNF genes are conserved in mammals, at least 136 and potentially more than 200 genes of this type are primate-specific including many recent segmental duplicates. KRAB-ZNF genes are active in a wide variety of human tissues suggesting roles in many key biological processes, but most member genes remain completely uncharacterized. Because of their sheer numbers, wide-ranging tissue-specific expression patterns, and remarkable evolutionary divergence we predict that KRAB-ZNF transcription factors have played critical roles in crafting many aspects of human biology, including both deeply conserved and primate-specific traits.

  1. ATRX binds to atypical chromatin domains at the 3' exons of zinc finger genes to preserve H3K9me3 enrichment.

    Science.gov (United States)

    Valle-García, David; Qadeer, Zulekha A; McHugh, Domhnall S; Ghiraldini, Flávia G; Chowdhury, Asif H; Hasson, Dan; Dyer, Michael A; Recillas-Targa, Félix; Bernstein, Emily

    2016-06-02

    ATRX is a SWI/SNF chromatin remodeler proposed to govern genomic stability through the regulation of repetitive sequences, such as rDNA, retrotransposons, and pericentromeric and telomeric repeats. However, few direct ATRX target genes have been identified and high-throughput genomic approaches are currently lacking for ATRX. Here we present a comprehensive ChIP-sequencing study of ATRX in multiple human cell lines, in which we identify the 3' exons of zinc finger genes (ZNFs) as a new class of ATRX targets. These 3' exonic regions encode the zinc finger motifs, which can range from 1-40 copies per ZNF gene and share large stretches of sequence similarity. These regions often contain an atypical chromatin signature: they are transcriptionally active, contain high levels of H3K36me3, and are paradoxically enriched in H3K9me3. We find that these ZNF 3' exons are co-occupied by SETDB1, TRIM28, and ZNF274, which form a complex with ATRX. CRISPR/Cas9-mediated loss-of-function studies demonstrate (i) a reduction of H3K9me3 at the ZNF 3' exons in the absence of ATRX and ZNF274 and, (ii) H3K9me3 levels at atypical chromatin regions are particularly sensitive to ATRX loss compared to other H3K9me3-occupied regions. As a consequence of ATRX or ZNF274 depletion, cells with reduced levels of H3K9me3 show increased levels of DNA damage, suggesting that ATRX binds to the 3' exons of ZNFs to maintain their genomic stability through preservation of H3K9me3.

  2. Zinc

    Science.gov (United States)

    ... increase zinc absorption, although a protein in cow's milk slows absorption down. Soy proteins also reduce zinc absorption, possibly due to their ... cow's milk, and even less from soy-based milk. It isn't known whether high-protein diets influence zinc balance in adults.VegetarianismVegetarian diets ...

  3. Mycobacterial and HIV infections up-regulated human zinc finger protein 134, a novel positive regulator of HIV-1 LTR activity and viral propagation.

    Directory of Open Access Journals (Sweden)

    Ronald Benjamin

    Full Text Available BACKGROUND: Concurrent occurrence of HIV and Tuberculosis (TB infections influence the cellular environment of the host for synergistic existence. An elementary approach to understand such coalition at the molecular level is to understand the interactions of the host and the viral factors that subsequently effect viral replication. Long terminal repeats (LTR of HIV genome serve as a template for binding trans-acting viral and cellular factors that regulate its transcriptional activity, thereby, deciding the fate of HIV pathogenesis, making it an ideal system to explore the interplay between HIV and the host. METHODOLOGY/PRINCIPAL FINDINGS: In this study, using biotinylated full length HIV-1 LTR sequence as bait followed by MALDI analyses, we identified and further characterized human-Zinc-finger-protein-134 (hZNF-134 as a novel positive regulator of HIV-1 that promoted LTR-driven transcription and viral production. Over-expression of hZNF-134 promoted LTR driven luciferase activity and viral transcripts, resulting in increased virus production while siRNA mediated knockdown reduced both the viral transcripts and the viral titers, establishing hZNF-134 as a positive effector of HIV-1. HIV, Mycobacteria and HIV-TB co-infections increased hZNF-134 expressions in PBMCs, the impact being highest by mycobacteria. Corroborating these observations, primary TB patients (n = 22 recorded extraordinarily high transcript levels of hZNF-134 as compared to healthy controls (n = 16. CONCLUSIONS/SIGNIFICANCE: With these observations, it was concluded that hZNF-134, which promoted HIV-1 LTR activity acted as a positive regulator of HIV propagation in human host. High titers of hZNF-134 transcripts in TB patients suggest that up-regulation of such positive effectors of HIV-1 upon mycobacterial infection can be yet another mechanism by which mycobacteria assists HIV-1 propagation during HIV-TB co-infections. hZNF-134, an uncharacterized host protein, thus

  4. Evolution of C2H2-zinc finger genes and subfamilies in mammals: Species-specific duplication and loss of clusters, genes and effector domains

    Directory of Open Access Journals (Sweden)

    Aubry Muriel

    2008-06-01

    Full Text Available Abstract Background C2H2 zinc finger genes (C2H2-ZNF constitute the largest class of transcription factors in humans and one of the largest gene families in mammals. Often arranged in clusters in the genome, these genes are thought to have undergone a massive expansion in vertebrates, primarily by tandem duplication. However, this view is based on limited datasets restricted to a single chromosome or a specific subset of genes belonging to the large KRAB domain-containing C2H2-ZNF subfamily. Results Here, we present the first comprehensive study of the evolution of the C2H2-ZNF family in mammals. We assembled the complete repertoire of human C2H2-ZNF genes (718 in total, about 70% of which are organized into 81 clusters across all chromosomes. Based on an analysis of their N-terminal effector domains, we identified two new C2H2-ZNF subfamilies encoding genes with a SET or a HOMEO domain. We searched for the syntenic counterparts of the human clusters in other mammals for which complete gene data are available: chimpanzee, mouse, rat and dog. Cross-species comparisons show a large variation in the numbers of C2H2-ZNF genes within homologous mammalian clusters, suggesting differential patterns of evolution. Phylogenetic analysis of selected clusters reveals that the disparity in C2H2-ZNF gene repertoires across mammals not only originates from differential gene duplication but also from gene loss. Further, we discovered variations among orthologs in the number of zinc finger motifs and association of the effector domains, the latter often undergoing sequence degeneration. Combined with phylogenetic studies, physical maps and an analysis of the exon-intron organization of genes from the SCAN and KRAB domains-containing subfamilies, this result suggests that the SCAN subfamily emerged first, followed by the SCAN-KRAB and finally by the KRAB subfamily. Conclusion Our results are in agreement with the "birth and death hypothesis" for the evolution of

  5. Transcriptional activation is a conserved feature of the early embryonic factor Zelda that requires a cluster of four zinc fingers for DNA binding and a low-complexity activation domain.

    Science.gov (United States)

    Hamm, Danielle C; Bondra, Eliana R; Harrison, Melissa M

    2015-02-01

    Delayed transcriptional activation of the zygotic genome is a nearly universal phenomenon in metazoans. Immediately following fertilization, development is controlled by maternally deposited products, and it is not until later stages that widespread activation of the zygotic genome occurs. Although the mechanisms driving this genome activation are currently unknown, the transcriptional activator Zelda (ZLD) has been shown to be instrumental in driving this process in Drosophila melanogaster. Here we define functional domains of ZLD required for both DNA binding and transcriptional activation. We show that the C-terminal cluster of four zinc fingers mediates binding to TAGteam DNA elements in the promoters of early expressed genes. All four zinc fingers are required for this activity, and splice isoforms lacking three of the four zinc fingers fail to activate transcription. These truncated splice isoforms dominantly suppress activation by the full-length, embryonically expressed isoform. We map the transcriptional activation domain of ZLD to a central region characterized by low complexity. Despite relatively little sequence conservation within this domain, ZLD orthologs from Drosophila virilis, Anopheles gambiae, and Nasonia vitripennis activate transcription in D. melanogaster cells. Transcriptional activation by these ZLD orthologs suggests that ZLD functions through conserved interactions with a protein cofactor(s). We have identified distinct DNA-binding and activation domains within the critical transcription factor ZLD that controls the initial activation of the zygotic genome.

  6. A20 zinc finger protein inhibits TNF-induced apoptosis and stress response early in the signaling cascades and independently of binding to TRAF2 or 14-3-3 proteins.

    Science.gov (United States)

    Lademann, U; Kallunki, T; Jäättelä, M

    2001-03-01

    A20 zinc finger protein is a negative regulator of tumor necrosis factor (TNF)-induced signaling pathways leading to apoptosis, stress response and inflammation. A20 has been shown to bind to TNF-receptor-associated factor 2 (TRAF2) and 14-3-3 chaperone proteins. Our data indicate that the zinc finger domain of A20 is sufficient and that neither TRAF2 nor 14-3-3 binding is necessary for the inhibitory effects of A20. Mutations in the 14-3-3 binding site of A20 did, however, result in a partial cleavage of A20 protein suggesting that 14-3-3 chaperone proteins may stabilize A20. Furthermore, we show that A20 acts early in TNF-induced signaling cascades blocking both TNF-induced rapid activation of c-Jun N-terminal kinase and processing of the receptor-associated caspase-8. Taken together our data indicate that the zinc finger domain of A20 contains all necessary functional domains required for the inhibition of TNF signaling and that A20 may function at the level of the receptor signaling complex.

  7. The Arabidopsis a zinc finger domain protein ARS1 is essential for seed germination and ROS homeostasis in response to ABA and oxidative stress

    Directory of Open Access Journals (Sweden)

    Dongwon eBaek

    2015-11-01

    Full Text Available The phytohormone abscisic acid (ABA induces accumulation of reactive oxygen species (ROS, which can disrupt seed dormancy and plant development. Here, we report the isolation and characterization of an Arabidopsis thaliana mutant called ars1 (aba and ros sensitive 1 that showed hypersensitivity to ABA during seed germination and to methyl viologen (MV at the seedling stage. ARS1 encodes a nuclear protein with one zinc finger domain, two nuclear localization signal (NLS domains, and one nuclear export signal (NES. The ars1 mutants showed reduced expression of a gene for superoxide dismutase (CSD3 and enhanced accumulation of ROS after ABA treatment. Transient expression of ARS1 in Arabidopsis protoplasts strongly suppressed ABA-mediated ROS production. Interestingly, nuclear-localized ARS1 translocated to the cytoplasm in response to treatment with ABA, H2O2, or MV. Taken together, these results suggest that ARS1 modulates seed germination and ROS homeostasis in response to ABA and oxidative stress in plants.

  8. Human and mouse ZFY genes produce a conserved testis-specific transcript encoding a zinc finger protein with a short acidic domain and modified transactivation potential.

    Science.gov (United States)

    Decarpentrie, Fanny; Vernet, Nadège; Mahadevaiah, Shantha K; Longepied, Guy; Streichemberger, Eric; Aknin-Seifer, Isabelle; Ojarikre, Obah A; Burgoyne, Paul S; Metzler-Guillemain, Catherine; Mitchell, Michael J

    2012-06-15

    Mammalian ZFY genes are located on the Y chromosome, and code putative transcription factors with 12-13 zinc fingers preceded by a large acidic (activating) domain. In mice, there are two genes, Zfy1 and Zfy2, which are expressed mainly in the testis. Their transcription increases in germ cells as they enter meiosis, both are silenced by meiotic sex chromosome inactivation (MSCI) during pachytene, and Zfy2 is strongly reactivated later in spermatids. Recently, we have shown that mouse Zfy2, but not Zfy1, is involved in triggering the apoptotic elimination of specific types of sex chromosomally aberrant spermatocytes. In humans, there is a single widely transcribed ZFY gene, and there is no evidence for a specific role in the testis. Here, we characterize ZFY transcription during spermatogenesis in mice and humans. In mice, we define a variety of Zfy transcripts, among which is a Zfy2 transcript that predominates in spermatids, and a Zfy1 transcript, lacking an exon encoding approximately half of the acidic domain, which predominates prior to MSCI. In humans, we have identified a major testis-specific ZFY transcript that encodes a protein with the same short acidic domain. This represents the first evidence that ZFY has a conserved function during human spermatogenesis. We further show that, in contrast to the full acidic domain, the short domain does not activate transcription in yeast, and we hypothesize that this explains the functional difference observed between Zfy1 and Zfy2 during mouse meiosis.

  9. Generation of Interleukin-2 Receptor Gamma Gene Knockout Pigs from Somatic Cells Genetically Modified by Zinc Finger Nuclease-Encoding mRNA

    Science.gov (United States)

    Watanabe, Masahito; Nakano, Kazuaki; Matsunari, Hitomi; Matsuda, Taisuke; Maehara, Miki; Kanai, Takahiro; Kobayashi, Mirina; Matsumura, Yukina; Sakai, Rieko; Kuramoto, Momoko; Hayashida, Gota; Asano, Yoshinori; Takayanagi, Shuko; Arai, Yoshikazu; Umeyama, Kazuhiro; Nagaya, Masaki; Hanazono, Yutaka; Nagashima, Hiroshi

    2013-01-01

    Zinc finger nuclease (ZFN) is a powerful tool for genome editing. ZFN-encoding plasmid DNA expression systems have been recently employed for the generation of gene knockout (KO) pigs, although one major limitation of this technology is the use of potentially harmful genome-integrating plasmid DNAs. Here we describe a simple, non-integrating strategy for generating KO pigs using ZFN-encoding mRNA. The interleukin-2 receptor gamma (IL2RG) gene was knocked out in porcine fetal fibroblasts using ZFN-encoding mRNAs, and IL2RG KO pigs were subsequently generated using these KO cells through somatic cell nuclear transfer (SCNT). The resulting IL2RG KO pigs completely lacked a thymus and were deficient in T and NK cells, similar to human X-linked SCID patients. Our findings demonstrate that the combination of ZFN-encoding mRNAs and SCNT provides a simple robust method for producing KO pigs without genomic integration. PMID:24130776

  10. Roles of Candida albicans Gat2, a GATA-type zinc finger transcription factor, in biofilm formation, filamentous growth and virulence.

    Science.gov (United States)

    Du, Han; Guan, Guobo; Xie, Jing; Sun, Yuan; Tong, Yaojun; Zhang, Lixin; Huang, Guanghua

    2012-01-01

    Candida albicans is the most common human fungal pathogen, causing not only superficial infections, but also life-threatening systemic disease. C. albicans can grow in several morphological forms including unicellular yeast-form, elongated hyphae and pseudohyphae. In certain natural environments, C. albicans also exists as biofilms, which are structured and surface-attached microbial communities. Transcription factors play a critical role in morphogenesis and biofilm development. In this study, we identified four adhesion-promoting transcription factors (Tec1, Cph1, Ume6 and Gat2) by screening a transcription factor overexpression library. Sequence analysis indicates that Gat2 is a GATA-type zinc finger transcription factor. Here we showed that the gat2/gat2 mutant failed to form biofilms on the plastic and silicone surfaces. Overexpression of GAT2 gene promoted filamentous and invasive growth on agar containing Lee's medium, while deletion of this gene had an opposite effect. However, inactivation of Gat2 had no obvious effect on N-acetyl-glucosamine (GlcNAc) induced hyphal development. In a mouse model of systemic infection, the gat2/gat2 mutant showed strongly attenuated virulence. Our results suggest that Gat2 plays a critical role in C. albicans biofilm formation, filamentous growth and virulence.

  11. Regions outside the DNA-binding domain are critical for proper in vivo specificity of an archetypal zinc finger transcription factor.

    Science.gov (United States)

    Burdach, Jon; Funnell, Alister P W; Mak, Ka Sin; Artuz, Crisbel M; Wienert, Beeke; Lim, Wooi F; Tan, Lit Yeen; Pearson, Richard C M; Crossley, Merlin

    2014-01-01

    Transcription factors (TFs) are often regarded as being composed of a DNA-binding domain (DBD) and a functional domain. The two domains are considered separable and autonomous, with the DBD directing the factor to its target genes and the functional domain imparting transcriptional regulation. We examined an archetypal zinc finger (ZF) TF, Krüppel-like factor 3 with an N-terminal domain that binds the corepressor CtBP and a DBD composed of three ZFs at its C-terminus. We established a system to compare the genomic occupancy profile of wild-type Krüppel-like factor 3 with two mutants affecting the N-terminal functional domain: a mutant unable to contact the cofactor CtBP and a mutant lacking the entire N-terminal domain, but retaining the ZFs intact. Chromatin immunoprecipitation followed by sequencing was used to assess binding across the genome in murine embryonic fibroblasts. Unexpectedly, we observe that mutations in the N-terminal domain generally reduced binding, but there were also instances where binding was retained or even increased. These results provide a clear demonstration that the correct localization of TFs to their target genes is not solely dependent on their DNA-contact domains. This informs our understanding of how TFs operate and is of relevance to the design of artificial ZF proteins.

  12. Sda1, a Cys2-His2 zinc finger transcription factor, is involved in polyol metabolism and fumonisin B1 production in Fusarium verticillioides.

    Science.gov (United States)

    Malapi-Wight, Martha; Smith, Jonathon; Campbell, Jacquelyn; Bluhm, Burton H; Shim, Won-Bo

    2013-01-01

    The ubiquitous ascomycete Fusarium verticillioides causes ear rot and stalk rot of maize, both of which reduce grain quality and yield. Additionally, F. verticillioides produces the mycotoxin fumonisin B1 (FB1) during infection of maize kernels, and thus potentially compromises human and animal health. The current knowledge is fragmentary regarding the regulation of FB1 biosynthesis, particularly when considering interplay with environmental factors such as nutrient availability. In this study, SDA1 of F. verticillioides, predicted to encode a Cys-2 His-2 zinc finger transcription factor, was shown to play a key role in catabolizing select carbon sources. Growth of the SDA1 knock-out mutant (Δsda1) was completely inhibited when sorbitol was the sole carbon source and was severely impaired when exclusively provided mannitol or glycerol. Deletion of SDA1 unexpectedly increased FB1 biosynthesis, but reduced arabitol and mannitol biosynthesis, as compared to the wild-type progenitor. Trichoderma reesei ACE1, a regulator of cellulase and xylanase expression, complemented the F. verticillioides Δsda1 mutant, which indicates that Ace1 and Sda1 are functional orthologs. Taken together, the data indicate that Sda1 is a transcriptional regulator of carbon metabolism and toxin production in F. verticillioides.

  13. Expression Profiling and Functional Implications of a Set of Zinc Finger Proteins, ZNF423, ZNF470, ZNF521, and ZNF780B, in Primary Osteoarthritic Articular Chondrocytes

    Directory of Open Access Journals (Sweden)

    Maria Mesuraca

    2014-01-01

    Full Text Available Articular chondrocytes are responsible for the maintenance of healthy articulations; indeed, dysregulation of their functions, including the production of matrix proteins and matrix-remodeling proteases, may result in fraying of the tissue and development of osteoarthritis (OA. To explore transcriptional mechanisms that contribute to the regulation of chondrocyte homeostasis and may be implicated in OA development, we compared the gene expression profile of a set of zinc finger proteins potentially linked to the control of chondrocyte differentiation and/or functions (ZNF423, ZNF470, ZNF521, and ZNF780B in chondrocytes from patients affected by OA and from subjects not affected by OA. This analysis highlighted a significantly lower expression of the transcript encoding ZNF423 in chondrocytes from OA, particularly in elderly patients. Interestingly, this decrease was mirrored by the similarly reduced expression of PPARγ, a known target of ZNF423 with anti-inflammatory and chondroprotective properties. The ZNF521 mRNA instead was abundant in all primary chondrocytes studied; the RNAi-mediated silencing of this gene significantly altered the COL2A/COL1 expression ratio, associated with the maintenance of the differentiated phenotype, in chondrocytes cultivated in alginate beads. These results suggest a role for ZNF423 and ZNF521 in the regulation of chondrocyte homeostasis and warrant further investigations to elucidate their mechanism of action.

  14. Loss of c-Kit and bone marrow failure upon conditional removal of the GATA-2 C-terminal zinc finger domain in adult mice.

    Science.gov (United States)

    Li, Haiyan S; Jin, Jin; Liang, Xiaoxuan; Matatall, Katie A; Ma, Ying; Zhang, Huiyuan; Ullrich, Stephen E; King, Katherine Y; Sun, Shao-Cong; Watowich, Stephanie S

    2016-09-01

    Heterozygous mutations in the transcriptional regulator GATA-2 associate with multilineage immunodeficiency, myelodysplastic syndrome (MDS), and acute myeloid leukemia (AML). The majority of these mutations localize in the zinc finger (ZnF) domains, which mediate GATA-2 DNA binding. Deregulated hematopoiesis with GATA-2 mutation frequently develops in adulthood, yet GATA-2 function in the bone marrow remains unresolved. To investigate this, we conditionally deleted the GATA-2 C-terminal ZnF (C-ZnF) coding sequences in adult mice. Upon Gata2 C-ZnF deletion, we observed rapid peripheral cytopenia, bone marrow failure, and decreased c-Kit expression on hematopoietic progenitors. Transplant studies indicated GATA-2 has a cell-autonomous role in bone marrow hematopoiesis. Moreover, myeloid lineage populations were particularly sensitive to Gata2 hemizygosity, while molecular assays indicated GATA-2 regulates c-Kit expression in multilineage progenitor cells. Enforced c-Kit expression in Gata2 C-ZnF-deficient hematopoietic progenitors enhanced myeloid colony activity, suggesting GATA-2 sustains myelopoiesis via a cell intrinsic role involving maintenance of c-Kit expression. Our results provide insight into mechanisms regulating hematopoiesis in bone marrow and may contribute to a better understanding of immunodeficiency and bone marrow failure associated with GATA-2 mutation.

  15. Sexual orientation in Drosophila is altered by the satori mutation in the sex-determination gene fruitless that encodes a zinc finger protein with a BTB domain.

    Science.gov (United States)

    Ito, H; Fujitani, K; Usui, K; Shimizu-Nishikawa, K; Tanaka, S; Yamamoto, D

    1996-09-03

    We have isolated a new Drosophila mutant, satori (sat), the males of which do not court or copulate with female flies. The sat mutation comaps with fruitless (fru) at 91B and does not rescue the bisexual phenotype of fru, indicating that sat is allelic to fru (fru(sat)). The fru(sat) adult males lack a male-specific muscle, the muscle of Lawrence, as do adult males with other fru alleles. Molecular cloning and analyses of the genomic and complementary DNAs indicated that transcription of the fru locus yields several different transcripts. The sequence of fru cDNA clones revealed a long open reading frame that potentially encodes a putative transcription regulator with a BTB domain and two zinc finger motifs. In the 5' noncoding region, three putative transformer binding sites were identified in the female transcript but not in male transcripts. The fru gene is expressed in a population of brain cells, including those in the antennal lobe, that have been suggested to be involved in determination of male sexual orientation. We suggest that fru functions downstream of tra in the sex-determination cascade in some neural cells and that inappropriate sexual development of these cells in the fru mutants results in altered sexual orientation of the fly.

  16. Chemical shift as a probe of molecular interfaces: NMR studies of DNA binding by the three amino-terminal zinc finger domains from transcription factor IIIA

    Energy Technology Data Exchange (ETDEWEB)

    Foster, Mark P.; Wuttke, Deborah S.; Clemens, Karen R.; Jahnke, Wolfgang; Radhakrishnan, Ishwar; Tennant, Linda; Reymond, Martine; Chung, John; Wright, Peter E. [Scripps Research Institute, Department of Molecular Biology and Skaggs Institute for Chemical Biology (United States)

    1998-07-15

    We report the NMR resonance assignments for a macromolecular protein/DNA complex containing the three amino-terminal zinc fingers (92 amino acid residues) of Xenopus laevis TFIIIA (termed zf1-3) bound to the physiological DNA target (15 base pairs), and for the free DNA. Comparisons are made of the chemical shifts of protein backbone{sup 1} H{sup N}, {sup 15}N,{sup 13} C{sup {alpha}} and{sup 13} C{sup {beta}} and DNA base and sugar protons of the free and bound species. Chemical shift changes are analyzed in the context of the structures of the zf1-3/DNA complex to assess the utility of chemical shift change as a probe of molecular interfaces. Chemical shift perturbations that occur upon binding in the zf1-3/DNA complex do not correspond directly to the structural interface, but rather arise from a number of direct and indirect structural and dynamic effects.

  17. Engineered zinc-finger proteins can compensate genetic haploinsufficiency by transcriptional activation of the wild-type allele: application to Willams-Beuren syndrome and supravalvular aortic stenosis.

    Science.gov (United States)

    Zhang, Pei; Huang, Angela; Morales-Ruiz, Manuel; Starcher, Barry C; Huang, Yan; Sessa, William C; Niklason, Laura E; Giordano, Frank J

    2012-11-01

    Williams-Beuren syndrome (WBS) and supravalvular aortic stenosis (SVAS) are genetic syndromes marked by the propensity to develop severe vascular stenoses. Vascular lesions in both syndromes are caused by haploinsufficiency of the elastin gene. We used these distinct genetic syndromes as models to evaluate the feasibility of using engineered zinc-finger protein transcription factors (ZFPs) to achieve compensatory expression of haploinsufficient genes by inducing augmented expression from the remaining wild-type allele. For complex genes with multiple splice variants, this approach could have distinct advantages over cDNA-based gene replacement strategies. Targeting the elastin gene, we show that transcriptional activation by engineered ZFPs can induce compensatory expression from the wild-type allele in the setting of classic WBS and SVAS genetic mutations, increase elastin expression in wild-type cells, induce expression of the major elastin splice variants, and recapitulate their natural stoichiometry. Further, we establish that transcriptional activation of the mutant allele in SVAS does not overcome nonsense-mediated decay, and thus ZFP-mediated transcriptional activation is not likely to induce production of a mutant protein, a crucial consideration. Finally, we show in bioengineered blood vessels that ZFP-mediated induction of elastin expression is capable of stimulating functional elastogenesis. Haploinsufficiency is a common mechanism of genetic disease. These findings have significant implications for WBS and SVAS, and establish that haploinsufficiency can be overcome by targeted transcriptional activation without inducing protein expression from the mutant allele.

  18. A conserved function of the zinc finger transcription factor Sp8/9 in allometric appendage growth in the milkweed bug Oncopeltus fasciatus.

    Science.gov (United States)

    Schaeper, Nina D; Prpic, Nikola-Michael; Wimmer, Ernst A

    2009-08-01

    The genes encoding the closely related zinc finger transcription factors Buttonhead (Btd) and D-Sp1 are expressed in the developing limb primordia of Drosophila melanogaster and are required for normal growth of the legs. The D-Sp1 homolog of the red flour beetle Tribolium castaneum, Sp8 (appropriately termed Sp8/9), is also required for the proper growth of the leg segments. Here we report on the isolation and functional study of the Sp8/9 gene from the milkweed bug Oncopeltus fasciatus. We show that Sp8/9 is expressed in the developing appendages throughout development and that the downregulation of Sp8/9 via RNAi leads to antennae, rostrum, and legs with shortened and fused segments. This supports a conserved role of Sp8/9 in allometric leg segment growth. However, all leg segments including the claws are present and the expression of the leg genes Distal-less, dachshund, and homothorax are proportionally normal, thus providing no evidence for a role of Sp8/9 in appendage specification.

  19. Engineering drought tolerant tomato plants over-expressing BcZAT12 gene encoding a C₂H₂ zinc finger transcription factor.

    Science.gov (United States)

    Rai, Avinash Chandra; Singh, Major; Shah, Kavita

    2013-01-01

    Efficient genetic transformation of cotyledonary explants of tomato (Solanum lycopersicum, cv. H-86, Kashi vishesh) was obtained. Disarmed Agrobacterium tumifaciens strain GV 3101 was used in conjugation with binary vector pBinAR containing a construct consisting of the coding sequence of the BcZAT12 gene under the regulatory control of the stress inducible Bclea1a promoter. ZAT12 encodes a C₂H₂ zinc finger protein which confers multiple abiotic stress tolerance to plants. Integration of ZAT12 gene into nuclear genome of individual kanamycin resistant transformed T₀ tomato lines was confirmed by Southern blot hybridization with segregation analysis of T(1) plants showing Mendelian inheritance of the transgene. Expression of ZAT12 in drought-stressed transformed tomato lines was verified in T₂ generation plants using RT-PCR. Of the six transformed tomato lines (ZT1-ZT6) the transformants ZT1 and ZT5 showed maximum expression of BcZAT12 gene transcripts when exposed to 7 days drought stress. Analysis of relative water content (RWC), electrolyte leakage (EL), chlorophyll colour index (CCI), H₂O₂ level and catalase activity suggested that tomato BcZAT12 transformants ZT1 and ZT5 have significantly increased levels of drought tolerance. These results suggest that BcZAT12 transformed tomato cv. H-86 has real potential for molecular breeding programs aimed at augmenting yield of tomato in regions affected with drought stress.

  20. Sda1, a Cys2-His2 zinc finger transcription factor, is involved in polyol metabolism and fumonisin B1 production in Fusarium verticillioides.

    Directory of Open Access Journals (Sweden)

    Martha Malapi-Wight

    Full Text Available The ubiquitous ascomycete Fusarium verticillioides causes ear rot and stalk rot of maize, both of which reduce grain quality and yield. Additionally, F. verticillioides produces the mycotoxin fumonisin B1 (FB1 during infection of maize kernels, and thus potentially compromises human and animal health. The current knowledge is fragmentary regarding the regulation of FB1 biosynthesis, particularly when considering interplay with environmental factors such as nutrient availability. In this study, SDA1 of F. verticillioides, predicted to encode a Cys-2 His-2 zinc finger transcription factor, was shown to play a key role in catabolizing select carbon sources. Growth of the SDA1 knock-out mutant (Δsda1 was completely inhibited when sorbitol was the sole carbon source and was severely impaired when exclusively provided mannitol or glycerol. Deletion of SDA1 unexpectedly increased FB1 biosynthesis, but reduced arabitol and mannitol biosynthesis, as compared to the wild-type progenitor. Trichoderma reesei ACE1, a regulator of cellulase and xylanase expression, complemented the F. verticillioides Δsda1 mutant, which indicates that Ace1 and Sda1 are functional orthologs. Taken together, the data indicate that Sda1 is a transcriptional regulator of carbon metabolism and toxin production in F. verticillioides.

  1. A Novel Role for the Zinc-Finger Transcription Factor EGL-46 in the Differentiation of Gas-Sensing Neurons in Caenorhabditis elegans

    DEFF Research Database (Denmark)

    Rojo Romanos, Teresa; Gramstrup Petersen, Jakob; Redo Riveiro, Alba;

    2014-01-01

    Oxygen (O2) and carbon dioxide (CO2) provoke distinct olfactory behaviors via specialized sensory neurons across metazoa. In the nematode Caenorhabditis elegans, the BAG sensory neurons are specialized to sense changes in both O2 and CO2 levels in the environment. The precise functionality of the......-13. Thereby, three conserved transcription factors collaborate to ensure neuron type-specific identity features of the BAG gas-sensing neurons.......Oxygen (O2) and carbon dioxide (CO2) provoke distinct olfactory behaviors via specialized sensory neurons across metazoa. In the nematode Caenorhabditis elegans, the BAG sensory neurons are specialized to sense changes in both O2 and CO2 levels in the environment. The precise functionality...... is partially, though not completely, controlled by ETS-5, an ETS-domain-containing transcription factor, and EGL-13, a Sox transcription factor. We report here, the identification of EGL-46, a zinc-finger transcription factor, which regulates BAG gas-sensing fate in partially parallel pathways to ETS-5 and EGL...

  2. A Zinc-Finger-Family Transcription Factor, AbVf19, Is Required for the Induction of a Gene Subset Important for Virulence in Alternaria brassicicola

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, Akhil [Univ. of Hawaii, Manoa, HI (United States); Ohm, Robin A. [USDOE Joint Genome Inst., Walnut Creek, CA (United States); Oxiles, Lindsay [Univ. of Hawaii, Manoa, HI (United States); Brooks, Fred [Univ. of Hawaii, Manoa, HI (United States); Lawrence, Christopher B. [Virginia Polytechnic Inst. and State Univ. (Virginia Tech), Blacksburg, VA (United States); Grigoriev, Igor V. [USDOE Joint Genome Inst., Walnut Creek, CA (United States); Cho, Yangrae [Univ. of Hawaii, Manoa, HI (United States)

    2011-10-26

    Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen with a broad host range within the family Brassicaceae. It produces secondary metabolites that marginally affect virulence. Cell wall degrading enzymes (CDWE) have been considered important for pathogenesis but none of them individually have been identified as significant virulence factors in A. brassicicola. In this study, knockout mutants of a gene, AbVf19, were created and produced considerably smaller lesions than the wild type on inoculated host plants. The presence of tandem zinc-finger domains in the predicted amino acid sequence and nuclear localization of AbVf19- reporter protein suggested that it was a transcription factor. Gene expression comparisons using RNA-seq identified 74 genes being downregulated in the mutant during a late stage of infection. Among the 74 downregulated genes, 28 were putative CWDE genes. These were hydrolytic enzyme genes that composed a small fraction of genes within each family of cellulases, pectinases, cutinases, and proteinases. The mutants grew slower than the wild type on an axenic medium with pectin as a major carbon source. This study demonstrated the existence and the importance of a transcription factor that regulates a suite of genes that are important for decomposing and utilizing plant material during the late stage of plant infection.

  3. Zinc finger protein genes from Cucurbita pepo are promising tools for conferring non-Cucurbitaceae plants with ability to accumulate persistent organic pollutants.

    Science.gov (United States)

    Inui, Hideyuki; Hirota, Matashi; Goto, Junya; Yoshihara, Ryouhei; Kodama, Noriko; Matsui, Tomomi; Yamazaki, Kiyoshi; Eun, Heesoo

    2015-03-01

    Some cultivars of cucumbers, melons, pumpkins, and zucchini, which are members of the Cucurbitaceae family, are uniquely subject to contamination by hydrophobic pollutants such as the organohalogen insecticides DDT. However, the molecular mechanisms for the accumulation of these pollutants in cucurbits have not been determined. Here, cDNA subtraction analysis of Cucurbita pepo cultivars that are low and high accumulators of hydrophobic contaminants revealed that a gene for zinc finger proteins (ZFPs) are preferentially expressed in high accumulators. The cloned CpZFP genes were classified into 2 types: (1) the PBG type, which were expressed in C. pepo cultivars Patty Green, Black Beauty, and Gold Rush, and (2) the BG type, which were expressed in Black Beauty and Gold Rush. Expression of these CpZFP genes in transgenic tobacco plants carrying an aryl hydrocarbon receptor-based inducible gene expression system significantly induced β-glucuronidase activity when the plants were treated with a polychlorinated biphenyl (PCB) compound, indicating that highly hydrophobic PCBs accumulated in the plants. In transgenic tobacco plants carrying CpZFPs, accumulation of dioxins and dioxin-like compounds increased in their aerial parts when they were cultivated in the dioxin-contaminated soil. In summary, we propose that addition of CpZFP genes is a promising tool for conferring noncucurbits with the ability to accumulate hydrophobic contaminants.

  4. Identification of ‘safe harbor’ loci in indica rice genome by harnessing the property of zinc-finger nucleases to induce DNA damage and repair.

    Directory of Open Access Journals (Sweden)

    Christian eCantos

    2014-06-01

    Full Text Available Zinc-finger nucleases (ZFNs have proved to be successful tools for targeted genome manipulation in several organisms. Their main property is the induction of double-strand breaks (DSBs at specific sites, which are further repaired through homologous recombination (HR or non-homologous end joining (NHEJ. However, for the appropriate integration of genes at specific chromosomal locations, proper sites for gene integration need to be identified. These regions, hereby named safe harbor loci, must be localized in non-coding regions and possess high gene expression. In the present study, three different ZFN constructs (pZFN1, pZFN2, pZFN3, harboring β-glucuronidase (GUS as a reporter gene, were used to identify safe harbor loci regions on rice chromosomes. The constructs were delivered into IR64 rice by using an improved Agrobacterium-mediated transformation protocol, based on the use of immature embryos. Gene expression was measured by histochemical GUS activity and the flanking regions were determined through thermal-asymmetric interlaced polymerase chain reaction (TAIL PCR. Following sequencing, 28 regions were identified as putative sites for safe integration, but only one was localized in a non-coding region and it also possessed high GUS expression. These findings have significant applicability to create crops with new and valuable traits, since the site can be subsequently used to stably introduce one or more genes in a targeted manner.

  5. Screening and validation of specific zinc finger nucleases targeted pig CFTR gene%猪CFTR基因特异性锌指核酸酶的筛选与鉴定

    Institute of Scientific and Technical Information of China (English)

    王瑞; 王令; 林娟; 张存芳; 张智英

    2012-01-01

    【目的】获得靶向猪囊性纤维化跨膜传导调节因子(Cystic fibrosis transmembrane conductance regulator,CFTR)基因的特异锌指核酸酶(Zinc finger nuclease,ZFN),为建立CFTR基因敲除猪细胞系提供技术支持。【方法】通过开源式(Oligomerized pool engineering,OPEN)方法筛选,首先以已知三锌指蛋白为框架,随机突变单个锌指关键位点氨基酸编码序列,建立人工三锌指蛋白随机库;然后应用细菌双杂交技术,从库中筛选出能够结合CFTR基因靶位点的三锌指蛋白;最后将获得的锌指蛋白与非限制性核酸内切酶FokⅠ组装成特异ZFN,通过酵母验证体系,检测ZFN靶向切割其识别序列的效率。【结果】获得3个人工三锌指蛋白随机库,每个库约含有2×106个单克隆,通过2轮细菌双杂交筛选,分别获得48个针对CFTR基因左右两侧靶位点的三锌指蛋白。ZFN活性验证结果表明,约90%的ZFN能够在酵母细胞内实现靶向切割,获得了高效特异的ZFN。【结论】获得了猪CFTR基因高效特异的ZFN。%【Objective】Specific zinc finger nucleases(ZFN)were screened to target pig cystic fibrosis transmembrane conductance regulator(CFTR)gene for the preparation of generating CFTR gene knockout pig cell line.【Method】Upon the oligomerized pool engineering(OPEN)strategy,firstly,three libraries of 3-zinc finger proteins were constructed.The DNA sequences of recognition amino acid residues of a finger motif were randomized by cassette mutagenesis.Secondly,bacterial twohybrid(B2H)system was used to screen for ZFPs from the three zinc finger protein libraries.Finally,selected ZFPs were fused to FokⅠdomain to generate specific ZFN for subsequent validation of ZFN cleavage activity in yeast.【Result】Three artificial randomly libraries of 3-zinc finger proteins were obtained,and each contained about 2×106 colonies.The three zinc finger random libraries were used

  6. 菠萝锌指蛋白基因AcRCHYI的克隆与表达分析%Cloning and Expression Analysis of a Zinc Finger Protein Gene AcRCHY1 from Pineapple

    Institute of Scientific and Technical Information of China (English)

    杨祥燕; 蔡元保; 吴青松; 孙光明

    2009-01-01

    根据植物C3HCA型兼CHY型锌指蛋白的功能保守区设计简并引物,通过RT-PCR结合RACE方法从菠萝(Ananas comosus L.Merr)幼苗中克隆获得了一个新的菠萝锌指蛋白基因cDNA全长,将其命名为AcRCHY1.该基因cDNA全长1 261 bp,开放阅读框ORF为918 bp,推测其编码一个含有306个氨基酸残基的多肽.AcRCHY1蛋白具有保守的C3HC4(RING finger)和CHY两个锌指结构域,与其它植物锌指蛋白的同源性高达81%-91%.半定量RT-PCR分析表明,AcRCHY1在菠萝中呈组成性表达,在子房、花瓣和小花中的表达量明显高于根和叶;低温、高盐、干旱和ABA等非生物胁迫处理后,AcRCHY1在叶片中的表达明显增强.因此,AcRCHY1蛋白可能与花器官生长发育的调控有关,而且可能作为一个转录调控因子在菠萝响应低温、高盐和渗透胁迫过程中参与了依赖ABA的信号转导途径.%Employing a pair of degenerate primers designed from the conserved domain of plant C3HC4 type and CHY type zinc-finger proteins, a novel zinc-finger protein gene AcRCHY1 was isolated from pineapple (Ananas comosus) by RACE-PCR and RT-PCR techniques. The full length cDNA has 1 261 bp nucleotides with an open reading frame (ORF) of 918 bp encoding a precursor protein of 306 amino acid residues. AcRCHY1 protein sharing 81% -91% homology with zinc-finger proteins from other plants contains two conservative zinc finger motif, C3HC4 zinc finger (RING finger) and CHY zinc finger. Semi-quantitive RT-PCR analysis indicated that AcRCHY1 was constitutively expressed in all plant organs, but stronger in ovaries, petals and florets than in roots and leaves, and the abiotic stresses such as low temperature, high salt , drought (20% PEG6000) and ABA, could trigger a significant induction of AcRCHY1 in leaves. All these results suggest that the AcRCHY1 protein may be involved in the regulation of floral organ growth and development, and play an important role in signal transduction pathway of

  7. Evolution of KRAB-containing zinc finger proteins and their roles in species evolution%KRAB型锌指蛋白的进化及在物种演化中的功能

    Institute of Scientific and Technical Information of China (English)

    王进龙; 王建; 田春艳

    2016-01-01

    C2H2型锌指蛋白家族是目前发现的哺乳动物中最大的转录/转录调控因子家族,由一小群古老的含有真核锌指结构的转录因子经过多次基因复制和功能分化演化而来。KRAB型锌指蛋白(KRAB-containing zinc finger proteins, KRAB-ZFPs)作为C2H2型锌指蛋白家族中最大的亚家族,最早出现在四足脊椎动物,并随物种的进化数量快速增长,在人类中占据C2H2型锌指蛋白的60%左右。在物种演化中,进化压力主要改变KRAB-ZFPs的DNA结合能力,而KRAB-ZFPs介导的转录抑制能力则稳定存在。同时,多种KRAB-ZFPs能够与KRAB相关蛋白1(KRAB-associated protein 1, KAP1)协同作用沉默哺乳动物中反转录元件的活性,并与之协同进化,严格限制反转录原件的跳跃能力。本文综述了 KRAB-ZFPs 的数量倍增、锌指结构的灵活多变、KRAB-ZFPs/KAP1的转录抑制能力和反转录元件的跳跃性在促进哺乳动物调控网络的差异、基因组稳定性的变化和物种进化中的作用,旨在进一步揭示KRAB-ZFPs在推动物种稳定演化中的特点和功能。%The C2H2 zinc finger protein family, one of the largest families of transcription factor/transcriptional regulator in mammal, arose from a small ancestral group of eukaryotic zinc finger transcription factors through many repeated gene duplications accompanied by functional divergence. As the biggest subfamily of C2H2 zinc finger protein family, Kruppel-associated box-containing zinc finger proteins (KRAB-ZFPs) appeared at the period oftetrapod, expand rapidly along with species evolution, and take about 60%of the total C2H2 zinc finger proteins in human. During species evolution, the DNA binding ability of KRAB-ZFPs is changed while the KRAB-ZFPs-mediate transcriptional repression ability maintains stable under the evolution pressure. Moreover, multiple KRAB-ZFPs function synergistically with KAP1 on transcriptional silencing of retroelements, and

  8. Identity of zinc finger nucleases with specificity to herpes simplex virus type II genomic DNA: novel HSV-2 vaccine/therapy precursors

    Directory of Open Access Journals (Sweden)

    Wayengera Misaki

    2011-06-01

    Full Text Available Abstract Background Herpes simplex type II (HSV-2 is a member of the family herpesviridae. Human infection with this double stranded linear DNA virus causes genital ulcerative disease and existing treatment options only serve to resolve the symptomatology (ulcers associated with active HSV-2 infection but do not eliminate latent virus. As a result, infection with HSV-2 follows a life-long relapsing (active versus latent course. On the basis of a primitive bacterium anti-phage DNA defense, the restriction modification (R-M system, we previously identified the Escherichia coli restriction enzyme (REase EcoRII as a novel peptide to excise or irreversibly disrupt latent HSV-2 DNA from infected cells. However, sequences of the site specificity palindrome of EcoRII 5'-CCWGG-3' (W = A or T are equally present within the human genome and are a potential source of host-genome toxicity. This feature has limited previous HSV-2 EcoRII based therapeutic models to microbicides only, and highlights the need to engineer artificial REases (zinc finger nucleases-ZFNs with specificity to HSV-2 genomic-DNA only. Herein, the therapeutic-potential of zinc finger arrays (ZFAs and ZFNs is identified and modeled, with unique specificity to the HSV-2 genome. Methods and results Using the whole genome of HSV-2 strain HG52 (Dolan A et al.,, and with the ZFN-consortium's CoDA-ZiFiT software pre-set at default, more than 28,000 ZFAs with specificity to HSV-2 DNA were identified. Using computational assembly (through in-silico linkage to the Flavobacterium okeanokoites endonuclease Fok I of the type IIS class, 684 ZFNs with specificity to the HSV-2 genome, were constructed. Graphic-analysis of the HSV-2 genome-cleavage pattern using the afore-identified ZFNs revealed that the highest cleavage-incidence occurred within the 30,950 base-pairs (~between the genomic context coordinates 0.80 and 1.00 at the 3' end of the HSV-2 genome. At approximately 3,095 bp before and after the

  9. Cytotoxic Effects during Knock Out of Multiple Porcine Endogenous Retrovirus (PERV Sequences in the Pig Genome by Zinc Finger Nucleases (ZFN.

    Directory of Open Access Journals (Sweden)

    Marwan Semaan

    Full Text Available Xenotransplantation has been proposed as a solution to the shortage of suitable human donors for transplantation and pigs are currently favoured as donor animals. However, xenotransplantation may be associated with the transmission of zoonotic microorganisms. Whereas most porcine microorganisms representing a risk for the human recipient may be eliminated by designated pathogen free breeding, multiple copies of porcine endogenous retroviruses (PERVs are integrated in the genome of all pigs and cannot be eliminated this way. PERVs are released as infectious particles and infect human cells. The zinc finger nuclease (ZFN technology allows knocking out specifically cellular genes, however it was not yet used to eliminate multiple integrated proviral sequences with a strong conservation in the target sequence. To reduce the risk of horizontal PERV transmission and to knock out as many as possible proviruses, for the first time the powerful tool of the ZFN technology was used. ZFN were designed to bind specifically to sequences conserved in all known replication-competent proviruses. Expression and transport of the ZFN into the nucleus was shown by Western blot analysis, co-localisation analysis, PLA and FRET. Survival of transfected cells was analysed using fluorescent ZFN and cell counting. After transfection a strong expression of the ZFN proteins and a co-localisation of the expressed ZFN proteins were shown. However, expression of the ZFN was found to be extremely toxic for the transfected cells. The induced cytotoxicity was likely due to the specific cutting of the high copy number of the PERV proviruses, which is also commonly observed when ZFN with low specificity cleave numerous off-target sites in a genome. This is the first attempt to knock out multiple, nearly identical, genes in a cellular genome using ZFN. The attempt failed, and other strategies should be used to prevent PERV transmission.

  10. In situ genetic correction of the sickle cell anemia mutation in human induced pluripotent stem cells using engineered zinc finger nucleases.

    Science.gov (United States)

    Sebastiano, Vittorio; Maeder, Morgan L; Angstman, James F; Haddad, Bahareh; Khayter, Cyd; Yeo, Dana T; Goodwin, Mathew J; Hawkins, John S; Ramirez, Cherie L; Batista, Luis F Z; Artandi, Steven E; Wernig, Marius; Joung, J Keith

    2011-11-01

    The combination of induced pluripotent stem cell (iPSC) technology and targeted gene modification by homologous recombination (HR) represents a promising new approach to generate genetically corrected, patient-derived cells that could be used for autologous transplantation therapies. This strategy has several potential advantages over conventional gene therapy including eliminating the need for immunosuppression, avoiding the risk of insertional mutagenesis by therapeutic vectors, and maintaining expression of the corrected gene by endogenous control elements rather than a constitutive promoter. However, gene targeting in human pluripotent cells has remained challenging and inefficient. Recently, engineered zinc finger nucleases (ZFNs) have been shown to substantially increase HR frequencies in human iPSCs, raising the prospect of using this technology to correct disease causing mutations. Here, we describe the generation of iPSC lines from sickle cell anemia patients and in situ correction of the disease causing mutation using three ZFN pairs made by the publicly available oligomerized pool engineering method (OPEN). Gene-corrected cells retained full pluripotency and a normal karyotype following removal of reprogramming factor and drug-resistance genes. By testing various conditions, we also demonstrated that HR events in human iPSCs can occur as far as 82 bps from a ZFN-induced break. Our approach delineates a roadmap for using ZFNs made by an open-source method to achieve efficient, transgene-free correction of monogenic disease mutations in patient-derived iPSCs. Our results provide an important proof of principle that ZFNs can be used to produce gene-corrected human iPSCs that could be used for therapeutic applications.

  11. Positional cloning of zinc finger domain transcription factor Zfp69, a candidate gene for obesity-associated diabetes contributed by mouse locus Nidd/SJL.

    Directory of Open Access Journals (Sweden)

    Stephan Scherneck

    2009-07-01

    Full Text Available Polygenic type 2 diabetes in mouse models is associated with obesity and results from a combination of adipogenic and diabetogenic alleles. Here we report the identification of a candidate gene for the diabetogenic effect of a QTL (Nidd/SJL, Nidd1 contributed by the SJL, NON, and NZB strains in outcross populations with New Zealand Obese (NZO mice. A critical interval of distal chromosome 4 (2.1 Mbp conferring the diabetic phenotype was identified by interval-specific congenic introgression of SJL into diabetes-resistant C57BL/6J, and subsequent reporter cross with NZO. Analysis of the 10 genes in the critical interval by sequencing, qRT-PCR, and RACE-PCR revealed a striking allelic variance of Zfp69 encoding zinc finger domain transcription factor 69. In NZO and C57BL/6J, a retrotransposon (IAPLTR1a in intron 3 disrupted the gene by formation of a truncated mRNA that lacked the coding sequence for the KRAB (Krüppel-associated box and Znf-C2H2 domains of Zfp69, whereas the diabetogenic SJL, NON, and NZB alleles generated a normal mRNA. When combined with the B6.V-Lep(ob background, the diabetogenic Zfp69(SJL allele produced hyperglycaemia, reduced gonadal fat, and increased plasma and liver triglycerides. mRNA levels of the human orthologue of Zfp69, ZNF642, were significantly increased in adipose tissue from patients with type 2 diabetes. We conclude that Zfp69 is the most likely candidate for the diabetogenic effect of Nidd/SJL, and that retrotransposon IAPLTR1a contributes substantially to the genetic heterogeneity of mouse strains. Expression of the transcription factor in adipose tissue may play a role in the pathogenesis of type 2 diabetes.

  12. Generation of GFAP::GFP astrocyte reporter lines from human adult fibroblast-derived iPS cells using zinc-finger nuclease technology.

    Science.gov (United States)

    Zhang, Ping-Wu; Haidet-Phillips, Amanda M; Pham, Jacqueline T; Lee, Youngjin; Huo, Yuqing; Tienari, Pentti J; Maragakis, Nicholas J; Sattler, Rita; Rothstein, Jeffrey D

    2016-01-01

    Astrocytes are instrumental to major brain functions, including metabolic support, extracellular ion regulation, the shaping of excitatory signaling events and maintenance of synaptic glutamate homeostasis. Astrocyte dysfunction contributes to numerous developmental, psychiatric and neurodegenerative disorders. The generation of adult human fibroblast-derived induced pluripotent stem cells (iPSCs) has provided novel opportunities to study mechanisms of astrocyte dysfunction in human-derived cells. To overcome the difficulties of cell type heterogeneity during the differentiation process from iPSCs to astroglial cells (iPS astrocytes), we generated homogenous populations of iPS astrocytes using zinc-finger nuclease (ZFN) technology. Enhanced green fluorescent protein (eGFP) driven by the astrocyte-specific glial fibrillary acidic protein (GFAP) promoter was inserted into the safe harbor adeno-associated virus integration site 1 (AAVS1) locus in disease and control-derived iPSCs. Astrocyte populations were enriched using Fluorescence Activated Cell Sorting (FACS) and after enrichment more than 99% of iPS astrocytes expressed mature astrocyte markers including GFAP, S100β, NFIA and ALDH1L1. In addition, mature pure GFP-iPS astrocytes exhibited a well-described functional astrocytic activity in vitro characterized by neuron-dependent regulation of glutamate transporters to regulate extracellular glutamate concentrations. Engraftment of GFP-iPS astrocytes into rat spinal cord grey matter confirmed in vivo cell survival and continued astrocytic maturation. In conclusion, the generation of GFAP::GFP-iPS astrocytes provides a powerful in vitro and in vivo tool for studying astrocyte biology and astrocyte-driven disease pathogenesis and therapy.

  13. Drosophila Zpr1 (Zinc finger protein 1 is required downstream of both EGFR and FGFR signaling in tracheal subcellular lumen formation.

    Directory of Open Access Journals (Sweden)

    Oscar E Ruiz

    Full Text Available The cellular and molecular cues involved in creating branched tubular networks that transport liquids or gases throughout an organism are not well understood. To identify factors required in branching and lumen formation of Drosophila tracheal terminal cells, a model for branched tubular networks, we performed a forward genetic-mosaic screen to isolate mutations affecting these processes. From this screen, we have identified the first Drosophila mutation in the gene Zpr1 (Zinc finger protein 1 by the inability of Zpr1-mutant terminal cells to form functional, gas-filled lumens. We show that Zpr1 defective cells initiate lumen formation, but are blocked from completing the maturation required for gas filling. Zpr1 is an evolutionarily conserved protein first identified in mammalian cells as a factor that binds the intracellular domain of the unactivated epidermal growth factor receptor (EGFR. We show that down-regulation of EGFR in terminal cells phenocopies Zpr1 mutations and that Zpr1 is epistatic to ectopic lumen formation driven by EGFR overexpression. However, while Zpr1 mutants are fully penetrant, defects observed when reducing EGFR activity are only partially penetrant. These results suggest that a distinct pathway operating in parallel to the EGFR pathway contributes to lumen formation, and this pathway is also dependent on Zpr1. We provide evidence that this alternative pathway may involve fibroblast growth factor receptor (FGFR signaling. We suggest a model in which Zpr1 mediates both EGFR and FGFR signal transduction cascades required for lumen formation in terminal cells. To our knowledge, this is the first genetic evidence placing Zpr1 downstream of EGFR signaling, and the first time Zpr1 has been implicated in FGFR signaling. Finally, we show that down-regulation of Smn, a protein known to interact with Zpr1 in mammalian cells, shows defects similar to Zpr1 mutants.

  14. The C2H2-type Zinc Finger Protein ZFP182 is Involved in Abscisic Acid-Induced Antioxidant Defense in Rice

    Institute of Scientific and Technical Information of China (English)

    Hong Zhang; Lan Ni; Yanpei Liu; Yunfei Wang; Aying Zhang; Mingpu Tan; Mingyi Jiang

    2012-01-01

    C2H2-type zinc finger proteins (ZFPs) are thought to play important roles in modulating the responses of plants to drought,salinity and oxidative stress.However,direct evidence is lacking for the involvement of these ZFPs in abscisic acid (ABA)-induced antioxidant defense in plants.In this study,the role of the rice (Oryza sativa L.sub.japonica cv.Nipponbare) C2H2-type ZFP ZFP182 in ABA-induced antioxidant defense and the relationship between ZFP182 and two rice MAPKs,OsMPK1 and OsMPK5 in ABA signaling were investigated.ABA treatment induced the increases in the expression of ZFP182,OsMPK1 and OsMPK5,and the activities of superoxide dismutase (SOD) and ascorbate peroxidase (APX) in rice leaves.The transient gene expression analysis and the transient RNA interference (RNAi) analysis in protoplasts showed that ZFP182,OsMPK1 and OsMPK5 are involved in ABA-induced up-regulation in the activities of SOD and APX.Besides,OsMPK1 and OsMPK5 were shown to be required for the up-regulation in the expression of ZFP182 in ABA signaling,but ZFP182 did not mediate the ABA-induced up-regulation in the expression of OsMPK1 and OsMPK5.These results indicate that ZFP182 is required for ABA-induced antioxidant defense and the expression of ZFP182 is regulated by rice MAPKs in ABA signaling.

  15. Improvement of acetic acid tolerance of Saccharomyces cerevisiae using a zinc-finger-based artificial transcription factor and identification of novel genes involved in acetic acid tolerance.

    Science.gov (United States)

    Ma, Cui; Wei, Xiaowen; Sun, Cuihuan; Zhang, Fei; Xu, Jianren; Zhao, Xinqing; Bai, Fengwu

    2015-03-01

    Acetic acid is present in cellulosic hydrolysate as a potent inhibitor, and the superior acetic acid tolerance of Saccharomyces cerevisiae ensures good cell viability and efficient ethanol production when cellulosic raw materials are used as substrates. In this study, a mutant strain of S. cerevisiae ATCC4126 (Sc4126-M01) with improved acetic acid tolerance was obtained through screening strains transformed with an artificial zinc finger protein transcription factor (ZFP-TF) library. Further analysis indicated that improved acetic acid tolerance was associated with improved catalase (CAT) activity. The ZFP coding sequence associated with the improved phenotype was identified, and real-time RT-PCR analysis revealed that three of the possible genes involved in the enhanced acetic acid tolerance regulated by this ZFP-TF, namely YFL040W, QDR3, and IKS1, showed decreased transcription levels in Sc4126-M01 in the presence of acetic acid, compared to those in the control strain. Sc4126-M01 mutants having QDR3 and IKS1 deletion (ΔQDR3 and ΔIKS1) exhibited higher acetic acid tolerance than the wild-type strain under acetic acid treatment. Glucose consumption rate and ethanol productivity in the presence of 5 g/L acetic acid were improved in the ΔQDR3 mutant compared to the wild-type strain. Our studies demonstrated that the synthetic ZFP-TF library can be used to improve acetic acid tolerance of S. cerevisiae and that the employment of an artificial transcription factor can facilitate the exploration of novel functional genes involved in stress tolerance of S. cerevisiae.

  16. The brain-specific neural zinc finger transcription factor 2b (NZF-2b/7ZFMyt1 suppresses cocaine self-administration in rats

    Directory of Open Access Journals (Sweden)

    Vijay Chandrasekar

    2010-04-01

    Full Text Available Brain-specific neural-zinc-finger transcription factor-2b (NZF2b/7ZFMyt1 is induced in the mesolimbic dopaminergic region after chronic cocaine exposure and lentiviral-mediated expression of NZF2b/7ZFMyt1 in the nucleus accumbens results in decreased locomotor activity (Chandrasekar and Dreyer, 2009. In this study the role of NZF2b/7ZFMyt1 in active cocaine seeking and of its interaction with histone deacetylase on the altered behavior has been observed. Localized expression of NZF2b/7ZFMyt1 in the nucleus accumbens resulted in attenuated cocaine self-administration, whereas silencing this transcription factor with lentiviruses expressing siRNAs increased the animal′s motivation to self-infuse cocaine. Low doses of sodium butyrate, a potent inhibitor of histone deacetylase, were sufficient to reverse the NZF2b/7ZFMyt1-mediated decrease in cocaine self-administration. NZF2b/7ZFMyt1 expression resulted in strong induction of transcription factors REST1 and NAC1 and of the dopamine D2 receptor, with concomitant inhibition of BDNF and its receptor TrkB. We show that NZF2b/7ZFMyt1 colocalizes with histone deacetylase-2 (HDAC2, probably overcoming the suppression of transcriptional activity caused by Lingo1. These findings show that molecular adaptations mediated by NZF2b/7ZFMyt1 expression possibly lead to decreased responsiveness to the reinforcing properties of cocaine and play a prominent role in affecting the behavioral changes induced by the drug.

  17. Tuberculate fruit gene Tu encodes a C2 H2 zinc finger protein that is required for the warty fruit phenotype in cucumber (Cucumis sativus L.).

    Science.gov (United States)

    Yang, Xuqin; Zhang, Weiwei; He, Huanle; Nie, Jingtao; Bie, Beibei; Zhao, Junlong; Ren, Guoliang; Li, Yue; Zhang, Dabing; Pan, Junsong; Cai, Run

    2014-06-01

    Cucumber fruits that have tubercules and spines (trichomes) are known to possess a warty (Wty) phenotype. In this study, the tuberculate fruit gene Tu was identified by map-based cloning, and was found to encode a transcription factor (TF) with a single C2 H2 zinc finger domain. Tu was identified in all 38 Wty lines examined, and was completely absent from all 56 non-warty (nWty) lines. Cucumber plants transgenic for Tu (TCP) revealed that Tu was required for the Wty fruit phenotype. Subcellular localization showed that the fusion protein GFP-Tu was localized mainly to the nucleus. Based on analyses of semi-quantitative and quantitative reverse transcription polymerase chain reaction (RT-PCR), and mRNA in situ hybridization, we found that Tu was expressed specifically in fruit spine cells during development of fruit tubercules. Moreover, cytokinin (CTK) content measurements and cytological observations in Wty and nWty fruits revealed that the Wty fruit phenotype correlated with high endogenous CTK concentrations. As a result of further analyses on the transcriptomic profile of the nWty fruit epidermis and TCP fruit warts, expression of CTK-associated genes, and hormone content in nWty fruit epidermis, Wty fruit warts and epidermis, and TCP fruit warts and epidermis, we found that Tu probably promoted CTK biosynthesis in fruit warts. Here we show that Tu could not be expressed in the glabrous and tubercule-free mutant line gl that contained Tu, this result that futher confirmed the epistatic effect of the trichome (spine) gene Gl over Tu. Taken together, these data led us to propose a genetic pathway for the Wty fruit trait that could guide future mechanistic studies.

  18. Zinc-Finger Transcription Factor ZAT6 Positively Regulates Cadmium Tolerance through the Glutathione-Dependent Pathway in Arabidopsis1[OPEN

    Science.gov (United States)

    Chen, Jian; Yan, Xingxing; Liu, Yunlei; Wang, Ren; Fan, Tingting; Ren, Yongbing; Tang, Xiaofeng; Xiao, Fangming

    2016-01-01

    Cadmium (Cd) is an environmental pollutant with high toxicity to animals and plants. It has been established that the glutathione (GSH)-dependent phytochelatin (PC) synthesis pathway is one of the most important mechanisms contributing to Cd accumulation and tolerance in plants. However, the transcription factors involved in regulating GSH-dependent PC synthesis pathway remain largely unknown. Here, we identified an Arabidopsis (Arabidopsis thaliana) Cd-resistant mutant xcd2-D (XVE system-induced cadmium-tolerance2) using a forward genetics approach. The mutant gene underlying xcd2-D mutation was revealed to encode a known zinc-finger transcription factor, ZAT6. Transgenic plants overexpressing ZAT6 showed significant increase of Cd tolerance, whereas loss of function of ZAT6 led to decreased Cd tolerance. Increased Cd accumulation and tolerance in ZAT6-overexpressing lines was GSH dependent and associated with Cd-activated synthesis of PC, which was correlated with coordinated activation of PC-synthesis related gene expression. By contrast, loss of function of ZAT6 reduced Cd accumulation and tolerance, which was accompanied by abolished PC synthesis and gene expression. Further analysis revealed that ZAT6 positively regulates the transcription of GSH1, GSH2, PCS1, and PCS2, but ZAT6 is capable of specifically binding to GSH1 promoter in vivo. Consistently, overexpression of GSH1 has been shown to restore Cd sensitivity in the zat6-1 mutant, suggesting that GSH1 is a key target of ZAT6. Taken together, our data provide evidence that ZAT6 coordinately activates PC synthesis-related gene expression and directly targets GSH1 to positively regulate Cd accumulation and tolerance in Arabidopsis. PMID:26983992

  19. Positional cloning of zinc finger domain transcription factor Zfp69, a candidate gene for obesity-associated diabetes contributed by mouse locus Nidd/SJL.

    Science.gov (United States)

    Scherneck, Stephan; Nestler, Matthias; Vogel, Heike; Blüher, Matthias; Block, Marcel-Dominique; Berriel Diaz, Mauricio; Herzig, Stephan; Schulz, Nadja; Teichert, Marko; Tischer, Sina; Al-Hasani, Hadi; Kluge, Reinhart; Schürmann, Annette; Joost, Hans-Georg

    2009-07-01

    Polygenic type 2 diabetes in mouse models is associated with obesity and results from a combination of adipogenic and diabetogenic alleles. Here we report the identification of a candidate gene for the diabetogenic effect of a QTL (Nidd/SJL, Nidd1) contributed by the SJL, NON, and NZB strains in outcross populations with New Zealand Obese (NZO) mice. A critical interval of distal chromosome 4 (2.1 Mbp) conferring the diabetic phenotype was identified by interval-specific congenic introgression of SJL into diabetes-resistant C57BL/6J, and subsequent reporter cross with NZO. Analysis of the 10 genes in the critical interval by sequencing, qRT-PCR, and RACE-PCR revealed a striking allelic variance of Zfp69 encoding zinc finger domain transcription factor 69. In NZO and C57BL/6J, a retrotransposon (IAPLTR1a) in intron 3 disrupted the gene by formation of a truncated mRNA that lacked the coding sequence for the KRAB (Krüppel-associated box) and Znf-C2H2 domains of Zfp69, whereas the diabetogenic SJL, NON, and NZB alleles generated a normal mRNA. When combined with the B6.V-Lep(ob) background, the diabetogenic Zfp69(SJL) allele produced hyperglycaemia, reduced gonadal fat, and increased plasma and liver triglycerides. mRNA levels of the human orthologue of Zfp69, ZNF642, were significantly increased in adipose tissue from patients with type 2 diabetes. We conclude that Zfp69 is the most likely candidate for the diabetogenic effect of Nidd/SJL, and that retrotransposon IAPLTR1a contributes substantially to the genetic heterogeneity of mouse strains. Expression of the transcription factor in adipose tissue may play a role in the pathogenesis of type 2 diabetes.

  20. Efficient methods for targeted mutagenesis in zebrafish using zinc-finger nucleases: data from targeting of nine genes using CompoZr or CoDA ZFNs.

    Directory of Open Access Journals (Sweden)

    Raman Sood

    Full Text Available Recently, it has been shown that targeted mutagenesis using zinc-finger nucleases (ZFNs and transcription activator-like effector nucleases (TALENs can be used to generate knockout zebrafish lines for analysis of their function and/or developing disease models. A number of different methods have been developed for the design and assembly of gene-specific ZFNs and TALENs, making them easily available to most zebrafish researchers. Regardless of the choice of targeting nuclease, the process of generating mutant fish is similar. It is a time-consuming and multi-step process that can benefit significantly from development of efficient high throughput methods. In this study, we used ZFNs assembled through either the CompoZr (Sigma-Aldrich or the CoDA (context-dependent assembly platforms to generate mutant zebrafish for nine genes. We report our improved high throughput methods for 1 evaluation of ZFNs activity by somatic lesion analysis using colony PCR, eliminating the need for plasmid DNA extractions from a large number of clones, and 2 a sensitive founder screening strategy using fluorescent PCR with PIG-tailed primers that eliminates the stutter bands and accurately identifies even single nucleotide insertions and deletions. Using these protocols, we have generated multiple mutant alleles for seven genes, five of which were targeted with CompoZr ZFNs and two with CoDA ZFNs. Our data also revealed that at least five-fold higher mRNA dose was required to achieve mutagenesis with CoDA ZFNs than with CompoZr ZFNs, and their somatic lesion frequency was lower (<5% when compared to CopmoZr ZFNs (9-98%. This work provides high throughput protocols for efficient generation of zebrafish mutants using ZFNs and TALENs.

  1. Zinc finger artificial transcription factor-based nearest inactive analogue/nearest active analogue strategy used for the identification of plant genes controlling homologous recombination.

    Science.gov (United States)

    Jia, Qi; van Verk, Marcel C; Pinas, Johan E; Lindhout, Beatrice I; Hooykaas, Paul J J; van der Zaal, Bert J

    2013-12-01

    In previous work, we selected a particular transcription factor, designated VP16-HRU, from a pool of zinc finger artificial transcription factors (ZF-ATFs) used for genome interrogation. When expressed in Arabidopsis thaliana under control of the ribosomal protein S5A promoter, the RPS5A::VP16-HRU construct led to a 200- to 300-fold increase in the frequency of somatic intrachromosomal homologous recombination (iHR). Because the expression of each ZF-ATF leads to a large number of transcriptional changes, we designed a strategy employing a collection of structurally similar ZF-ATFs to filter out the transcriptional changes relevant to the phenotype by deep sequencing. In that manner, 30 transcripts were found to be consistently induced in plants with enhanced homologous recombination (HR). For 25 of the cognate genes, their effect on the HR process was assessed using cDNA/gDNA expression constructs. For three genes, ectopic expression indeed led to enhanced iHR frequencies, albeit much lower than the frequency observed when a HR-inducing ZF-ATF was present. Altogether, our data demonstrate that despite the large number of transcriptional changes brought about by individual ZF-ATFs, causal changes can be identified. In our case, the picture emerged that a natural regulatory switch for iHR does not exist but that ZF-ATFs-like VP16-HRU act as an ectopic master switch, orchestrating the timely expression of a set of plant genes that each by themselves only have modest effects, but when acting together support an extremely high iHR frequency.

  2. Annexin A2 and zinc finger transcription factor Snail expression in glioma tissue and the regulating effect of corresponding siRNA on glioma cells

    Institute of Scientific and Technical Information of China (English)

    Zheng-Hai Deng; Ying-Zhi Hou

    2016-01-01

    Objective:To study the Annexin A2 and zinc finger transcription factor Snail expression in glioma tissue and the regulating effect of corresponding siRNA on glioma cells.Methods:Glioma and peri-tumor tissue were collected to determine AnnexinA2 and Snail expression; glioma cell lines U373-MG were cultured and transfected with AnnexinA2, Snail and NC siRNA, and then the cell viability, number of migrating and invading cells as well as the expression levels of proliferation and epithelial-mesenchymal transition genes were detected. Results:AnnexinA2 and SnailmRNA levels in glioma tissues were significantly higher than those in peri-tumor tissues; cell viability as well asRas, Raf, MEKandERK mRNA levels of AnnexinA2-siRNA group was significantly lower than those of NC-siRNA group, and the migrating cell number and invading cell number as well as E-cadherin, N-cadherin, Vimentin andα-SMA mRNA levels were not significantly different from those of NC-siRNA group; migrating cell number and invading cell number as well as N-cadherin, Vimentin andα-SMA mRNA levels of Snail-siRNA group were significantly lower than those of NC-siRNA group, E-cadherin mRNA level was significantly higher than that of NC-siRNA group, and the cell viability as well asRas, Raf, MEK andERK mRNA levels were not significantly different from those of NC-siRNA group.Conclusions:AnnexinA2 and Snail expression levels significantly increase in glioma tissues, highly expressed AnnexinA2 can promote cell proliferation and highly expressed Snail can promote epithelial-mesenchymal transition.

  3. A unique sequence in the N-terminal regulatory region controls the nuclear localization of KLF8 by cooperating with the C-terminal zinc-fingers

    Institute of Scientific and Technical Information of China (English)

    Tina S Mehta; Heng Lu; Xianhui Wang; Alison M Urvalek; Kim-Hang H Nguyen; Farah Monzur; Jojo D Hammond; Jameson Q Ma; Jihe Zhao

    2009-01-01

    Kruppel-like factor 8 (KLF8) transcription factor plays a critical role in cell cycle progression, oncogenic trans-formation, epithelial to mesenchymal transition and invasion. However, its nuclear localization signal(s) (NLS) has not been identified. KLF8 shares with other KLFs monopartite NLSs (mNLS) and C2H2 zinc fingers (ZFs), both of which have been shown to be the NLSs for some other KLFs. In this report, using PCR-directed mutagenesis and immunofluorescent microscopy, we show that disruption of the mNLSs, deletion of any single ZF, or mutation of the Zn2+-binding or DNA-contacting motifs did not affect the nuclear localization of KLF8. Deletion of>1.5 ZFs from C-terminus, however, caused cytoplasmic accumulation of KLF8. Surprisingly, deletion of amino acid (aa) 151-200 re-gion almost eliminated KLF8 from the nucleus. S165A, K171E or K171R mutation, or treatment with PKC inhibitor led to partial cytoplasmic accumulation. Co-immunoprecipitation demonstrated that KLF8 interacted with importin-β and this interaction required the ZF motif. Deletion of aa 1-150 or 201-261 region alone did not alter the nuclear lo-calization. BrdU incorporation and cyclin D1 promoter luciferase assays showed that the KLF8 mutants defective in nuclear localization could not promote DNA synthesis or cyclin D1 promoter activation as the wild-type KLF8 did. Taken together, these results suggest that KLF8 has two NLSs, one surrounding S165 and K171 and the other being two tandem ZFs, which are critical for the regulation of KLF8 nuclear localization and its cellular functions.

  4. ZINC FINGER OF ARABIDOPSIS THALIANA12 (ZAT12) Interacts with FER-LIKE IRON DEFICIENCY-INDUCED TRANSCRIPTION FACTOR (FIT) Linking Iron Deficiency and Oxidative Stress Responses.

    Science.gov (United States)

    Le, Cham Thi Tuyet; Brumbarova, Tzvetina; Ivanov, Rumen; Stoof, Claudia; Weber, Eva; Mohrbacher, Julia; Fink-Straube, Claudia; Bauer, Petra

    2016-01-01

    Plants grown under iron (Fe)-deficient conditions induce a set of genes that enhance the efficiency of Fe uptake by the roots. In Arabidopsis (Arabidopsis thaliana), the central regulator of this response is the basic helix-loop-helix transcription factor FER-LIKE IRON DEFICIENCY-INDUCED TRANSCRIPTION FACTOR (FIT). FIT activity is regulated by protein-protein interactions, which also serve to integrate external signals that stimulate and possibly inhibit Fe uptake. In the search of signaling components regulating FIT function, we identified ZINC FINGER OF ARABIDOPSIS THALIANA12 (ZAT12), an abiotic stress-induced transcription factor. ZAT12 interacted with FIT, dependent on the presence of the ethylene-responsive element-binding factor-associated amphiphilic repression motif. ZAT12 protein was found expressed in the root early differentiation zone, where its abundance was modulated in a root layer-specific manner. In the absence of ZAT12, FIT expression was upregulated, suggesting a negative effect of ZAT12 on Fe uptake. Consistently, zat12 loss-of-function mutants had higher Fe content than the wild type at sufficient Fe. We found that under Fe deficiency, hydrogen peroxide (H2O2) levels were enhanced in a FIT-dependent manner. FIT protein, in turn, was stabilized by H2O2 but only in the presence of ZAT12, showing that H2O2 serves as a signal for Fe deficiency responses. We propose that oxidative stress-induced ZAT12 functions as a negative regulator of Fe acquisition. A model where H2O2 mediates the negative regulation of plant responses to prolonged stress might be applicable to a variety of stress conditions.

  5. Effects of HAb18G/CD147 knockout on hepatocellular carcinoma cells in vitro using a novel zinc-finger nuclease-targeted gene knockout approach.

    Science.gov (United States)

    Li, Hong-Wei; Yang, Xiang-Min; Tang, Juan; Wang, Shi-Jie; Chen, Zhi-Nan; Jiang, Jian-Li

    2015-03-01

    HAb18G/CD147 belongs to the immunoglobulin superfamily and predominantly functions as an inducer of matrix metalloproteinase secretion for tumor invasion and metastasis. This study was designed to investigate the effects of HAb18G/CD147 knockout on hepatocellular carcinoma cells using zinc-finger nuclease (ZFNs)-targeted gene knockout approach. The HCC cell line SMMC-7721 was used for ZFNs-targeted cleavage of the HAb18G/CD147 gene. RT-PCR and Western blot assays were used to detect HAb18G/CD147 expression. HAb18G phenotypic changes following HAb18G/CD147 knockout in SMMC-K7721 cells were assessed using tumor cell adhesion, invasion, migration and colony formation and flow cytometric assays. These data demonstrated that tumor cell adhesion, invasion, migration, and colony formation capabilities of SMMC-K7721 were significantly reduced compared to parental cells or SMMC-7721 with re-expression of HAb18G/CD147 protein transfected with HAb18G/CD147 cDNA. Moreover, knockout of HAb18G/CD147 expression also induced SMMC-K7721 cells to undergo apoptosis compared to SMMC-7721 and SMMC-R7721 (P CD147 reduced p53 levels in SMMC-R7721 cells, possibly through inhibition of the PI3K-Akt-MDM2 signaling pathway. The findings provide a novel insight into the mechanisms underlying HAb18G/CD147-induced progression of HCC cells.

  6. Diversity of Prdm9 zinc finger array in wild mice unravels new facets of the evolutionary turnover of this coding minisatellite.

    Directory of Open Access Journals (Sweden)

    Jérôme Buard

    Full Text Available In humans and mice, meiotic recombination events cluster into narrow hotspots whose genomic positions are defined by the PRDM9 protein via its DNA binding domain constituted of an array of zinc fingers (ZnFs. High polymorphism and rapid divergence of the Prdm9 gene ZnF domain appear to involve positive selection at DNA-recognition amino-acid positions, but the nature of the underlying evolutionary pressures remains a puzzle. Here we explore the variability of the Prdm9 ZnF array in wild mice, and uncovered a high allelic diversity of both ZnF copy number and identity with the caracterization of 113 alleles. We analyze features of the diversity of ZnF identity which is mostly due to non-synonymous changes at codons -1, 3 and 6 of each ZnF, corresponding to amino-acids involved in DNA binding. Using methods adapted to the minisatellite structure of the ZnF array, we infer a phylogenetic tree of these alleles. We find the sister species Mus spicilegus and M. macedonicus as well as the three house mouse (Mus musculus subspecies to be polyphyletic. However some sublineages have expanded independently in Mus musculus musculus and M. m. domesticus, the latter further showing phylogeographic substructure. Compared to random genomic regions and non-coding minisatellites, none of these patterns appears exceptional. In silico prediction of DNA binding sites for each allele, overlap of their alignments to the genome and relative coverage of the different families of interspersed repeated elements suggest a large diversity between PRDM9 variants with a potential for highly divergent distributions of recombination events in the genome with little correlation to evolutionary distance. By compiling PRDM9 ZnF protein sequences in Primates, Muridae and Equids, we find different diversity patterns among the three amino-acids most critical for the DNA-recognition function, suggesting different diversification timescales.

  7. Scientific opinion addressing the safety assessment of plants developed using Zinc Finger Nuclease 3 and other Site-Directed Nucleases with similar function

    Directory of Open Access Journals (Sweden)

    EFSA Panel on Genetically Modified Organisms (GMO

    2012-10-01

    Full Text Available

    The European Commission requested that the EFSA Panel on Genetically Modified Organisms deliver a scientific opinion related to risk assessment of plants developed using the zinc finger nuclease 3 technique (ZFN-3 which allows the integration of gene(s in a predefined insertion site in the genome of the recipient species. Since other nucleases with a similar function to ZFN are considered in this opinion the term site-directed nuclease 3 (SDN-3 is used to describe the technique rather than ZFN-3 specifically. The EFSA GMO Panel considers that its guidance documents are applicable for the evaluation of food and feed products derived from plants developed using the SDN-3 technique and for performing an environmental risk assessment. However, on a case-by-case basis lesser amounts of event specific data may be needed for the risk assessment of plants developed using the SDN-3 technique. The EFSA GMO Panel compared the hazards associated with plants produced by the SDN-3 technique with those obtained by conventional plant breeding techniques and by currently used transgenesis. With respect to the genes introduced, the SDN-3 technique does not differ from transgenesis or from the other genetic modification techniques currently used, and can be used to introduce transgenes, intragenes or cisgenes. The main difference between the SDN-3 technique and transgenesis is that the insertion of DNA is targeted to a predefined region of the genome. Therefore, the SDN-3 technique can minimise hazards associated with the disruption of genes and/or regulatory elements in the recipient genome. Whilst the SDN-3 technique can induce off-target changes in the genome of the recipient plant these would be fewer than those occurring with most mutagenesis techniques. Furthermore, where such changes occur they would be of the same types as those produced by conventional breeding techniques.

  8. The BTB-zinc finger transcription factor abrupt acts as an epithelial oncogene in Drosophila melanogaster through maintaining a progenitor-like cell state.

    Directory of Open Access Journals (Sweden)

    Nezaket Turkel

    Full Text Available The capacity of tumour cells to maintain continual overgrowth potential has been linked to the commandeering of normal self-renewal pathways. Using an epithelial cancer model in Drosophila melanogaster, we carried out an overexpression screen for oncogenes capable of cooperating with the loss of the epithelial apico-basal cell polarity regulator, scribbled (scrib, and identified the cell fate regulator, Abrupt, a BTB-zinc finger protein. Abrupt overexpression alone is insufficient to transform cells, but in cooperation with scrib loss of function, Abrupt promotes the formation of massive tumours in the eye/antennal disc. The steroid hormone receptor coactivator, Taiman (a homologue of SRC3/AIB1, is known to associate with Abrupt, and Taiman overexpression also drives tumour formation in cooperation with the loss of Scrib. Expression arrays and ChIP-Seq indicates that Abrupt overexpression represses a large number of genes, including steroid hormone-response genes and multiple cell fate regulators, thereby maintaining cells within an epithelial progenitor-like state. The progenitor-like state is characterised by the failure to express the conserved Eyes absent/Dachshund regulatory complex in the eye disc, and in the antennal disc by the failure to express cell fate regulators that define the temporal elaboration of the appendage along the proximo-distal axis downstream of Distalless. Loss of scrib promotes cooperation with Abrupt through impaired Hippo signalling, which is required and sufficient for cooperative overgrowth with Abrupt, and JNK (Jun kinase signalling, which is required for tumour cell migration/invasion but not overgrowth. These results thus identify a novel cooperating oncogene, identify mammalian family members of which are also known oncogenes, and demonstrate that epithelial tumours in Drosophila can be characterised by the maintenance of a progenitor-like state.

  9. The BTB-zinc Finger Transcription Factor Abrupt Acts as an Epithelial Oncogene in Drosophila melanogaster through Maintaining a Progenitor-like Cell State

    Science.gov (United States)

    Turkel, Nezaket; Sahota, Virender K.; Bolden, Jessica E.; Goulding, Karen R.; Doggett, Karen; Willoughby, Lee F.; Blanco, Enrique; Martin-Blanco, Enrique; Corominas, Montserrat; Ellul, Jason; Aigaki, Toshiro; Richardson, Helena E.; Brumby, Anthony M.

    2013-01-01

    The capacity of tumour cells to maintain continual overgrowth potential has been linked to the commandeering of normal self-renewal pathways. Using an epithelial cancer model in Drosophila melanogaster, we carried out an overexpression screen for oncogenes capable of cooperating with the loss of the epithelial apico-basal cell polarity regulator, scribbled (scrib), and identified the cell fate regulator, Abrupt, a BTB-zinc finger protein. Abrupt overexpression alone is insufficient to transform cells, but in cooperation with scrib loss of function, Abrupt promotes the formation of massive tumours in the eye/antennal disc. The steroid hormone receptor coactivator, Taiman (a homologue of SRC3/AIB1), is known to associate with Abrupt, and Taiman overexpression also drives tumour formation in cooperation with the loss of Scrib. Expression arrays and ChIP-Seq indicates that Abrupt overexpression represses a large number of genes, including steroid hormone-response genes and multiple cell fate regulators, thereby maintaining cells within an epithelial progenitor-like state. The progenitor-like state is characterised by the failure to express the conserved Eyes absent/Dachshund regulatory complex in the eye disc, and in the antennal disc by the failure to express cell fate regulators that define the temporal elaboration of the appendage along the proximo-distal axis downstream of Distalless. Loss of scrib promotes cooperation with Abrupt through impaired Hippo signalling, which is required and sufficient for cooperative overgrowth with Abrupt, and JNK (Jun kinase) signalling, which is required for tumour cell migration/invasion but not overgrowth. These results thus identify a novel cooperating oncogene, identify mammalian family members of which are also known oncogenes, and demonstrate that epithelial tumours in Drosophila can be characterised by the maintenance of a progenitor-like state. PMID:23874226

  10. [Zinc].

    Science.gov (United States)

    Couinaud, C

    1984-10-01

    Zinc is indispensable for life from bacteria to man. As a trace element it is included in numerous enzymes or serves as their activator (more than 80 zinc metallo-enzymes). It is necessary for nucleic acid and protein synthesis, the formation of sulphated molecules (insulin, growth hormone, keratin, immunoglobulins), and the functioning of carbonic anhydrase, aldolases, many dehydrogenases (including alcohol-dehydrogenase, retinal reductase indispensable for retinal rod function), alkaline phosphatase, T cells and superoxide dismutase. Its lack provokes distinctive signs: anorexia, diarrhea, taste, smell and vision disorders, skin lesions, delayed healing, growth retardation, delayed appearance of sexual characteristics, diminished resistance to infection, and it may be the cause of congenital malformations. Assay is now simplified by atomic absorption spectrophotometry in blood or hair. There is a latent lack prior to any disease because of the vices of modern eating habits, and this increases during stress, infections or tissue healing processes. Its lack is accentuated during long-term parenteral feeding or chronic gastrointestinal affections. Correction is as simple as it is innocuous, and zinc supplements should be given more routinely during surgical procedures.

  11. Study on a cDNA sequence of cold inducible zinc finger protein in albinism tea cultivar "Xiaoxueya"%茶树品种“小雪芽”冷诱导锌指蛋白基因cDNA研究

    Institute of Scientific and Technical Information of China (English)

    王开荣; 李娜娜; 陆建良; 郑新强; 梁月荣; 吴颖; 李明

    2012-01-01

    A cDNA sequence of cold inducible zinc finger protein in leaf of albinism tea cultivar " Xiaoxueya" was investigated. The results shows that the cDNA sequence had 698 bp in length, with 83% and 82% identity to zinc finger protein mRNA of Glycine max and Ricinus communis respectively. It had an opening reading frame encoding 230 amino acids. Compared to that of common tea cuhivar " Fudingdabai" , there were three loci of nucleotide deletion and one locus of nucleotide substitution. Its deduced amino acid sequence had 99% identity to that of cuhivar " Fudingdabai" , among which there were 3 loci of amino acid substitution. The expression of cold inducible zinc finger protein in " Xiaoxueya" ,sas significantly lower than in " Fudingdabai". It is considered that the mutation in gene sequence resulted in the sensitivity of cultivar "Xiaoxueya" to low temperature through low expression and mutation of the cold inducible zinc finger protein.%分析了低温诱导型新梢白化茶树品种“小雪芽”叶片低温诱导锌指蛋白基因cDNA序列。该序列长度为698bp,与大豆锌指蛋白mRNA同源性为83%,与蓖麻锌指蛋白mRNA同源性为82%;具有可编码230个氨基酸的开放阅读框。与“福鼎大白茶”冷诱导锌指蛋白eDNA序列相比,该eDNA序列在50—51位上核苷酸AT缺失,第143位的A被置换为G,第654位T缺失。其翻译的蛋白质氨基酸序列与“福鼎大白茶”同源性达到99%,但有3个位点的氨基酸变异。该基因表达丰度明显低于“福鼎大白茶”。研究认为,基因结构差异,引起表达强度和蛋白质氨基酸序列的差异,可能是引起“小雪芽”品种对低温敏感的重要因素。

  12. The BTB/POZ zinc finger protein Broad-Z3 promotes dendritic outgrowth during metamorphic remodeling of the peripheral stretch receptor dbd

    Directory of Open Access Journals (Sweden)

    Scott Janet A

    2011-12-01

    Full Text Available Abstract Background Various members of the family of BTB/POZ zinc-finger transcription factors influence patterns of dendritic branching. One such member, Broad, is notable because its BrZ3 isoform is widely expressed in Drosophila in immature neurons around the time of arbor outgrowth. We used the metamorphic remodeling of an identified sensory neuron, the dorsal bipolar dendrite sensory neuron (dbd, to examine the effects of BrZ3 expression on the extent and pattern of dendrite growth during metamorphosis. Results Using live imaging of dbd in Drosophila pupae, we followed its normal development during metamorphosis and the effect of ectopic expression of BrZ3 on this development. After migration of its cell body, dbd extends a growth-cone that grows between two muscle bands followed by branching and turning back on itself to form a compact dendritic bundle. The ectopic expression of the BrZ3 isoform, using the GAL4/UAS system, caused dbd's dendritic tree to transform from its normal, compact, fasciculated form into a comb-like arbor that spread over on the body wall. Time-lapse analysis revealed that the expression of BrZ3 caused the premature extension of the primary dendrite onto immature myoblasts, ectopic growth past the muscle target region, and subsequent elaboration onto the epidermis. To control the timing of expression of BrZ3, we used a temperature-sensitive GAL80 mutant. When BrZ3 expression was delayed until after the extension of the primary dendrite, then a normal arbor was formed. By contrast, when BrZ3 expression was confined to only the early outgrowth phase, then ectopic arbors were subsequently formed and maintained on the epidermis despite the subsequent absence of BrZ3. Conclusions The adult arbor of dbd is a highly branched arbor whose branches self-fasciculate to form a compact dendritic bundle. The ectopic expression of BrZ3 in this cell causes a premature extension of its growth-cone, resulting in dendrites that extend

  13. Zinc finger nuclease mediated knockout of ADP-dependent glucokinase in cancer cell lines: effects on cell survival and mitochondrial oxidative metabolism.

    Directory of Open Access Journals (Sweden)

    Susan Richter

    Full Text Available Zinc finger nucleases (ZFN are powerful tools for editing genes in cells. Here we use ZFNs to interrogate the biological function of ADPGK, which encodes an ADP-dependent glucokinase (ADPGK, in human tumour cell lines. The hypothesis we tested is that ADPGK utilises ADP to phosphorylate glucose under conditions where ATP becomes limiting, such as hypoxia. We characterised two ZFN knockout clones in each of two lines (H460 and HCT116. All four clones had frameshift mutations in all alleles at the target site in exon 1 of ADPGK, and were ADPGK-null by immunoblotting. ADPGK knockout had little or no effect on cell proliferation, but compromised the ability of H460 cells to survive siRNA silencing of hexokinase-2 under oxic conditions, with clonogenic survival falling from 21±3% for the parental line to 6.4±0.8% (p = 0.002 and 4.3±0.8% (p = 0.001 for the two knockouts. A similar increased sensitivity to clonogenic cell killing was observed under anoxia. No such changes were found when ADPGK was knocked out in HCT116 cells, for which the parental line was less sensitive than H460 to anoxia and to hexokinase-2 silencing. While knockout of ADPGK in HCT116 cells caused few changes in global gene expression, knockout of ADPGK in H460 cells caused notable up-regulation of mRNAs encoding cell adhesion proteins. Surprisingly, we could discern no consistent effect on glycolysis as measured by glucose consumption or lactate formation under anoxia, or extracellular acidification rate (Seahorse XF analyser under oxic conditions in a variety of media. However, oxygen consumption rates were generally lower in the ADPGK knockouts, in some cases markedly so. Collectively, the results demonstrate that ADPGK can contribute to tumour cell survival under conditions of high glycolytic dependence, but the phenotype resulting from knockout of ADPGK is cell line dependent and appears to be unrelated to priming of glycolysis in these lines.

  14. Use of zinc-finger nucleases to knock out the WAS gene in K562 cells: a human cellular model for Wiskott-Aldrich syndrome

    Directory of Open Access Journals (Sweden)

    Miguel G. Toscano

    2013-03-01

    Mutations in the WAS gene cause Wiskott-Aldrich syndrome (WAS, which is characterized by eczema, immunodeficiency and microthrombocytopenia. Although the role of WASP in lymphocytes and myeloid cells is well characterized, its role on megakaryocyte (MK development is poorly understood. In order to develop a human cellular model that mimics the megakaryocytic-derived defects observed in WAS patients we used K562 cells, a well-known model for study of megakaryocytic development. We knocked out the WAS gene in K562 cells using a zinc-finger nuclease (ZFN pair targeting the WAS intron 1 and a homologous donor DNA that disrupted WASP expression. Knockout of WASP on K562 cells (K562WASKO cells resulted in several megakaryocytic-related defects such as morphological alterations, lower expression of CD41ɑ, lower increments in F-actin polymerization upon stimulation, reduced CD43 expression and increased phosphatidylserine exposure. All these defects have been previously described either in WAS-knockout mice or in WAS patients, validating K562WASKO as a cell model for WAS. However, K562WASPKO cells showed also increased basal F-actin and adhesion, increased expression of CD61 and reduced expression of TGFβ and Factor VIII, defects that have never been described before for WAS-deficient cells. Interestingly, these phenotypic alterations correlate with different roles for WASP in megakaryocytic differentiation. All phenotypic alterations observed in K562WASKO cells were alleviated upon expression of WAS following lentiviral transduction, confirming the role of WASP in these phenotypes. In summary, in this work we have validated a human cellular model, K562WASPKO, that mimics the megakaryocytic-related defects found in WAS-knockout mice and have found evidences for a role of WASP as regulator of megakaryocytic differentiation. We propose the use of K562WASPKO cells as a tool to study the molecular mechanisms involved in the megakaryocytic-related defects observed in WAS

  15. Genome-Wide Analysis of C2H2 Zinc-Finger Family Transcription Factors and Their Responses to Abiotic Stresses in Poplar (Populus trichocarpa.

    Directory of Open Access Journals (Sweden)

    Quangang Liu

    Full Text Available C2H2 zinc-finger (C2H2-ZF proteins are a large gene family in plants that participate in various aspects of normal plant growth and development, as well as in biotic and abiotic stress responses. To date, no overall analysis incorporating evolutionary history and expression profiling of the C2H2-ZF gene family in model tree species poplar (Populus trichocarpa has been reported.Here, we identified 109 full-length C2H2-ZF genes in P. trichocarpa, and classified them into four groups, based on phylogenetic analysis. The 109 C2H2-ZF genes were distributed unequally on 19 P. trichocarpa linkage groups (LGs, with 39 segmental duplication events, indicating that segmental duplication has been important in the expansion of the C2H2-ZF gene family. Promoter cis-element analysis indicated that most of the C2H2-ZF genes contain phytohormone or abiotic stress-related cis-elements. The expression patterns of C2H2-ZF genes, based on heatmap analysis, suggested that C2H2-ZF genes are involved in tissue and organ development, especially root and floral development. Expression analysis based on quantitative real-time reverse transcription polymerase chain reaction indicated that C2H2-ZF genes are significantly involved in drought, heat and salt response, possibly via different mechanisms.This study provides a thorough overview of the P. trichocarpa C2H2-ZF gene family and presents a new perspective on the evolution of this gene family. In particular, some C2H2-ZF genes may be involved in environmental stress tolerance regulation. PtrZFP2, 19 and 95 showed high expression levels in leaves and/or roots under environmental stresses. Additionally, this study provided a solid foundation for studying the biological roles of C2H2-ZF genes in Populus growth and development. These results form the basis for further investigation of the roles of these candidate genes and for future genetic engineering and gene functional studies in Populus.

  16. Zinc-Finger Nuclease Knockout of Dual-Specificity Protein Phosphatase-5 Enhances the Myogenic Response and Autoregulation of Cerebral Blood Flow in FHH.1BN Rats

    Science.gov (United States)

    Fan, Fan; Geurts, Aron M.; Pabbidi, Mallikarjuna R.; Smith, Stanley V.; Harder, David R.; Jacob, Howard; Roman, Richard J.

    2014-01-01

    We recently reported that the myogenic responses of the renal afferent arteriole (Af-Art) and middle cerebral artery (MCA) and autoregulation of renal and cerebral blood flow (RBF and CBF) were impaired in Fawn Hooded hypertensive (FHH) rats and were restored in a FHH.1BN congenic strain in which a small segment of chromosome 1 from the Brown Norway (BN) containing 15 genes including dual-specificity protein phosphatase-5 (Dusp5) were transferred into the FHH genetic background. We identified 4 single nucleotide polymorphisms in the Dusp5 gene in FHH as compared with BN rats, two of which altered CpG sites and another that caused a G155R mutation. To determine whether Dusp5 contributes to the impaired myogenic response in FHH rats, we created a Dusp5 knockout (KO) rat in the FHH.1BN genetic background using a zinc-finger nuclease that introduced an 11 bp frame-shift deletion and a premature stop codon at AA121. The expression of Dusp5 was decreased and the levels of its substrates, phosphorylated ERK1/2 (p-ERK1/2), were enhanced in the KO rats. The diameter of the MCA decreased to a greater extent in Dusp5 KO rats than in FHH.1BN and FHH rats when the perfusion pressure was increased from 40 to 140 mmHg. CBF increased markedly in FHH rats when MAP was increased from 100 to 160 mmHg, and CBF was better autoregulated in the Dusp5 KO and FHH.1BN rats. The expression of Dusp5 was higher at the mRNA level but not at the protein level and the levels of p-ERK1/2 and p-PKC were lower in cerebral microvessels and brain tissue isolated from FHH than in FHH.1BN rats. These results indicate that Dusp5 modulates myogenic reactivity in the cerebral circulation and support the view that a mutation in Dusp5 may enhance Dusp5 activity and contribute to the impaired myogenic response in FHH rats. PMID:25397684

  17. Zinc-finger nuclease knockout of dual-specificity protein phosphatase-5 enhances the myogenic response and autoregulation of cerebral blood flow in FHH.1BN rats.

    Directory of Open Access Journals (Sweden)

    Fan Fan

    Full Text Available We recently reported that the myogenic responses of the renal afferent arteriole (Af-Art and middle cerebral artery (MCA and autoregulation of renal and cerebral blood flow (RBF and CBF were impaired in Fawn Hooded hypertensive (FHH rats and were restored in a FHH.1BN congenic strain in which a small segment of chromosome 1 from the Brown Norway (BN containing 15 genes including dual-specificity protein phosphatase-5 (Dusp5 were transferred into the FHH genetic background. We identified 4 single nucleotide polymorphisms in the Dusp5 gene in FHH as compared with BN rats, two of which altered CpG sites and another that caused a G155R mutation. To determine whether Dusp5 contributes to the impaired myogenic response in FHH rats, we created a Dusp5 knockout (KO rat in the FHH.1BN genetic background using a zinc-finger nuclease that introduced an 11 bp frame-shift deletion and a premature stop codon at AA121. The expression of Dusp5 was decreased and the levels of its substrates, phosphorylated ERK1/2 (p-ERK1/2, were enhanced in the KO rats. The diameter of the MCA decreased to a greater extent in Dusp5 KO rats than in FHH.1BN and FHH rats when the perfusion pressure was increased from 40 to 140 mmHg. CBF increased markedly in FHH rats when MAP was increased from 100 to 160 mmHg, and CBF was better autoregulated in the Dusp5 KO and FHH.1BN rats. The expression of Dusp5 was higher at the mRNA level but not at the protein level and the levels of p-ERK1/2 and p-PKC were lower in cerebral microvessels and brain tissue isolated from FHH than in FHH.1BN rats. These results indicate that Dusp5 modulates myogenic reactivity in the cerebral circulation and support the view that a mutation in Dusp5 may enhance Dusp5 activity and contribute to the impaired myogenic response in FHH rats.

  18. MiRNA-205 modulates cellular invasion and migration via regulating zinc finger E-box binding homeobox 2 expression in esophageal squamous cell carcinoma cells

    Directory of Open Access Journals (Sweden)

    Yamashita Shunichi

    2011-03-01

    Full Text Available Abstract Background Esophageal squamous cell carcinoma (ESCC is often diagnosed at later stages until they are incurable. MicroRNA (miR is a small, non-coding RNA that negatively regulates gene expression mainly via translational repression. Accumulating evidence indicates that deregulation of miR is associated with human malignancies including ESCC. The aim of this study was to identify miR that could be specifically expressed and exert distinct biological actions in ESCC. Methods Total RNA was extracted from ESCC cell lines, OE21 and TE10, and a non-malignant human esophageal squamous cell line, Het-1A, and subjected to microarray analysis. Expression levels of miR that showed significant differences between the 2 ESCC and Het-1A cells based on the comprehensive analysis were analyzed by the quantitative reverse transcriptase (RT-PCR method. Then, functional analyses, including cellular proliferation, apoptosis and Matrigel invasion and the wound healing assay, for the specific miR were conducted. Using ESCC tumor samples and paired surrounding non-cancerous tissue obtained endoscopically, the association with histopathological differentiation was examined with quantitative RT-PCR. Results Based on the miR microarray analysis, there were 14 miRs that showed significant differences (more than 2-fold in expression between the 2 ESCC cells and non-malignant Het-1A. Among the significantly altered miRs, miR-205 expression levels were exclusively higher in 5 ESCC cell lines examined than any other types of malignant cell lines and Het-1A. Thus, miR-205 could be a specific miR in ESCC. Modulation of miR-205 expression by transfection with its precursor or anti-miR-205 inhibitor did not affect ESCC cell proliferation and apoptosis, but miR-205 was found to be involved in cell invasion and migration. Western blot revealed that knockdown of miR-205 expression in ESCC cells substantially enhanced expression of zinc finger E-box binding homeobox 2

  19. DPL-1 DP, LIN-35 Rb and EFL-1 E2F act with the MCD-1 zinc-finger protein to promote programmed cell death in Caenorhabditis elegans.

    Science.gov (United States)

    Reddien, Peter W; Andersen, Erik C; Huang, Michael C; Horvitz, H Robert

    2007-04-01

    The genes egl-1, ced-9, ced-4, and ced-3 play major roles in programmed cell death in Caenorhabditis elegans. To identify genes that have more subtle activities, we sought mutations that confer strong cell-death defects in a genetically sensitized mutant background. Specifically, we screened for mutations that enhance the cell-death defects caused by a partial loss-of-function allele of the ced-3 caspase gene. We identified mutations in two genes not previously known to affect cell death, dpl-1 and mcd-1 (modifier of cell death). dpl-1 encodes the C. elegans homolog of DP, the human E2F-heterodimerization partner. By testing genes known to interact with dpl-1, we identified roles in cell death for four additional genes: efl-1 E2F, lin-35 Rb, lin-37 Mip40, and lin-52 dLin52. mcd-1 encodes a novel protein that contains one zinc finger and that is synthetically required with lin-35 Rb for animal viability. dpl-1 and mcd-1 act with efl-1 E2F and lin-35 Rb to promote programmed cell death and do so by regulating the killing process rather than by affecting the decision between survival and death. We propose that the DPL-1 DP, MCD-1 zinc finger, EFL-1 E2F, LIN-35 Rb, LIN-37 Mip40, and LIN-52 dLin52 proteins act together in transcriptional regulation to promote programmed cell death.

  20. 绿竹锌指蛋白基因BoBZF克隆及功能初步分析%Cloning and Primary Functional Analysis of a Zinc Finger Protein Gene of BoBZF from Bambusa oldhamii

    Institute of Scientific and Technical Information of China (English)

    高志民; 刘青; 牟少华; 李雪平; 胡陶

    2012-01-01

    Zinc finger proteins play important roles in regulating gene expression in both prokaryote and eukaryotice. A zinc finger protein gene was isolated from leaves of Bambusa oldhamii with RT-PCR and RACE methods, and designed as BoBZF ( GenBank No. EU606025 ) . The full length cDNA of BoBZF is 1 076 bp containing an open reading frame which encodes 256 amino acids. The protein encoded by BoBZF has two B-Box domains, which indicates BoBZF belongs to B-Box zinc finger protein. Tissue specific expression showed that BoBZF expressed in leaf, sheath, stem and root, with the highest level in leaf. BoBZF gene was subcloned into the multiple cloning sites of pBI121 vector driven by 35S promoter, and transferred into Arabidopsis thaliana. RT-PCR analysis showed that BoBZF was expressed in the Arabidopsis. The transgenic plant was more drought tolerance than control, indicating that BoBZF was related to the drought tolerance ability of the bamboo.%锌指蛋白在原核生物与真核生物基因转录调控中发挥着重要作用.采用RT-PCR和RACE技术,从绿竹叶片中分离到1个锌指蛋白基因,命名为BoBZF(GenBank登记号:EU606025).BoBZF cDNA全长1 076 bp,编码256个氨基酸.蛋白结构分析表明,其编码的蛋白具有2个B-Box结构域,属于B-Box型锌指蛋白.组织特异性表达显示BoBZF在叶片、叶鞘、幼茎和根中均有表达,其中在叶片的表达丰度较高.将BoBZF置于CaMV 35S启动子控制下,构建到载体pBI121的多克隆位点,并转化拟南芥.RT-PCR分析表明,BoBZF已在拟南芥中表达.转基因植株耐旱性明显提高,意味着BoBZF与竹子的耐旱能力有关.

  1. The stem cell zinc finger 1 (SZF1)/ZNF589 protein has a human-specific evolutionary nucleotide DNA change and acts as a regulator of cell viability in the hematopoietic system.

    Science.gov (United States)

    Venturini, Letizia; Stadler, Michael; Manukjan, Georgi; Scherr, Michaela; Schlegelberger, Brigitte; Steinemann, Doris; Ganser, Arnold

    2016-04-01

    The stem cell zinc finger 1 (SZF1)/ZNF589 protein belongs to the large family of Krüppel-associated box domain-zinc finger (KRAB-ZNF) transcription factors, which are present only in higher vertebrates and epigenetically repress transcription by recruiting chromatin-modifying complexes to the promoter regions of their respective target genes. Although the distinct biological functions of most KRAB-ZNF proteins remain unknown, recent publications indicate their implication in fundamental processes, such as cell proliferation, apoptosis, differentiation, development, and tumorigenesis. SZF1/ZNF589 was first identified as a gene with SZF1-1 isoform specifically expressed in CD34(+) hematopoietic cells, strongly suggesting a role in epigenetic control of gene expression in hematopoietic stem/progenitor cells (HSPCs). However, the function of SZF1/ZNF589 in hematopoiesis has not yet been elucidated. Our study reveals SZF1/ZNF589 as a gene with a human-specific nucleotide DNA-change, conferring potential species-specific functional properties. Through shRNA-mediated loss-of-function experiments, we found that changes in expression of fundamental apoptosis-controlling genes are induced on SZF1/ZNF589 knockdown, resulting in inhibited growth of hematopoietic cell lines and decreased progenitor potential of primary human bone marrow CD34(+) cells. Moreover, we found that the SZF1/ZNF589 gene is differentially regulated during hypoxia in CD34(+) HSPCs in a cytokine-dependent manner, implicating its possible involvement in the maintenance of the hypoxic physiologic status of hematopoietic stem cells. Our results establish the role of SZF1/ZNF589 as a new functional regulator of the hematopoietic system.

  2. Nucleus Accumbens 1, a Pox virus and Zinc finger/Bric-a-brac Tramtrack Broad protein binds to TAR DNA-binding protein 43 and has a potential role in Amyotrophic Lateral Sclerosis.

    Science.gov (United States)

    Scofield, M D; Korutla, L; Jackson, T G; Kalivas, P W; Mackler, S A

    2012-12-27

    Protein degradation is a critical component of cellular maintenance. The intracellular translocation and targeting of the Ubiquitin Proteasome System (UPS) differentially coordinates a protein's half-life and thereby its function. Nucleus Accumbens 1 (NAC1), a member of the Pox virus and Zinc finger/Bric-a-brac Tramtrack Broad complex (POZ/BTB) family of proteins, participates in the coordinated proteolysis of synaptic proteins by mediating recruitment of the UPS to dendritic spines. Here we report a novel interaction between NAC1 and TAR DNA-binding protein 43 (TDP-43), a protein identified as the primary component of ubiquitinated protein aggregates found in patients with Amyotrophic Lateral Sclerosis (ALS). In vitro translated full-length TDP-43 associated with both the POZ/BTB domain and the non-POZ/BTB domain of NAC1 in GST pulldown assays. Other POZ/BTB proteins (including zinc finger POZ/BTB proteins and atypical POZ/BTB proteins) showed weak interactions with TDP-43. In addition, NAC1 and TDP-43 were present in the same immunocomplexes in different regions of mouse brain and spinal cord. In primary spinal cord cultures, TDP-43 expression was mainly nuclear, whereas NAC1 was both nuclear and cytoplasmic. In order to mimic ALS-like toxicity in the spinal cord culture system, we elevated extracellular glutamate levels resulting in the selective loss of motor neurons. Using this model, it was found that glutamate toxicity elicited a dose-dependent translocation of TDP-43 out of the nucleus of cholinergic neurons and increased the co-localization of NAC1 and TDP-43. These findings suggest that NAC1 may function to link TDP-43 to the proteasome; thereby, facilitating the post-translational modifications of TDP-43 that lead to the development of ALS.

  3. Requirement of the C3HC4 zinc RING finger of the Arabidopsis PEX10 for photorespiration and leaf peroxisome contact with chloroplasts.

    Science.gov (United States)

    Schumann, Uwe; Prestele, Jakob; O'Geen, Henriette; Brueggeman, Robert; Wanner, Gerhard; Gietl, Christine

    2007-01-16

    Plant peroxisomes perform multiple vital metabolic processes including lipid mobilization in oil-storing seeds, photorespiration, and hormone biosynthesis. Peroxisome biogenesis requires the function of peroxin (PEX) proteins, including PEX10, a C(3)HC(4) Zn RING finger peroxisomal membrane protein. Loss of function of PEX10 causes embryo lethality at the heart stage. We investigated the function of PEX10 with conditional sublethal mutants. Four T-DNA insertion lines expressing pex10 with a dysfunctional RING finger were created in an Arabidopsis WT background (DeltaZn plants). They could be normalized by growth in an atmosphere of high CO(2) partial pressure, indicating a defect in photorespiration. beta-Oxidation in mutant glyoxysomes was not affected. However, an abnormal accumulation of the photorespiratory metabolite glyoxylate, a lowered content of carotenoids and chlorophyll a and b, and a decreased quantum yield of photosystem II were detected under normal atmosphere, suggesting impaired leaf peroxisomes. Light and transmission electron microscopy demonstrated leaf peroxisomes of the DeltaZn plants to be more numerous, multilobed, clustered, and not appressed to the chloroplast envelope as in WT. We suggest that inactivation of the RING finger domain in PEX10 has eliminated protein interaction required for attachment of peroxisomes to chloroplasts and movement of metabolites between peroxisomes and chloroplasts.

  4. Finger pain

    Science.gov (United States)

    ... hands from the cold. When to Contact a Medical Professional Call your health care provider if: Your finger pain is caused by injury ... to Expect at Your Office Visit The health ... will be asked questions about your medical history and symptoms. You may have an x- ...

  5. Cloning and Molecular Characterization of a RING Zinc-Finger Gene of Hevea brasiliensis%巴西橡胶树环锌指蛋白基因克隆及其分子特性

    Institute of Scientific and Technical Information of China (English)

    朱家红; 李辉亮; 屠发志; 田维敏; 彭世清

    2006-01-01

    Using the information about the sequence from a differentially expressed clone (designated as HbSSH10)encodes a protein specifying a cysteine-rich sequence containing a putative "RING finger" or "C3HC4" consensus motif that was cloned recently by the subtractive hybridization between latex and leaves from rubber tree (Hevea brasiliensis). A full-length cDNA encoding C3HC4 type zinc-finger protein was isolated and characterized from rubber tree. Sequence analysis revealed that the ORFs of HbRZF encode 156 amino acid residues with a total predicted molecular mass of 17.2 kD,HbRZF protein having a putative "RING finger" segment (amino acid residues 100-144). The deduced amino acid sequences of HbRZF showed high identities of 48%,52% and 50% to those of the ring zinc protein from Poncirus trifoliata, Arabidopsis thaliana, Thellungiella halophila. The result of Northern blot analysis indicated that the transcripts of the HbRZF were expressed more in the latex than in the leaves, whereas little expression was detected in roots and flowers. The transcription of HbRZF was induced by jasmonic acid, whereas ethylene had little effect.%在先前的研究中通过抑制缩减杂交获得了一个在巴西橡胶树胶乳中特异表达的片段(HbSSH10),该片段含有"RING finger"或"C3HC4"保守序列.根据HbSSH10的序列信息设计引物并通过3'-RACE和5'-RACE的方法,获得了一个全长的cDNA(HbRZF).该cDNA含有589个核苷酸,含有完整的阅读框架,编码156个氨基酸.从它推导出的氨基酸序列中含有"RING finger"或"C3HC4"保守区(氨基酸100~144).该氨基酸序列与Poncirus trifoliata、Arabidopsis thaliana和Thellungiella halophila的环锌指蛋白的同源性分别为48%、52%和50%.Northern杂交分析表明HbRZF在胶乳中大量表达,在叶片中微量表达,而在根和花中几乎没有表达.茉莉酸处理可以诱导胶乳中HbRZF的表达,而乙烯对胶乳中HbRZF的表达基本上没有影响.

  6. Gclust Server: 3232 [Gclust Server

    Lifescience Database Archive (English)

    Full Text Available 3232 ATH_AT1G68200_145326690 Cluster Sequences Related Sequences(96) 307 zinc finge...r (CCCH-type) family protein 30 1.00e-45 42.86 33.33 0.0 0.0 0.0 75.0 Show 3232 Cluster ID 3232 Sequence ID ATH_AT1G68200_14532

  7. Research Advance on Zinc-finger Nucleases Technology in Mammalian Transgenesis%锌指核酸酶技术在动物转基因中的研究进展

    Institute of Scientific and Technical Information of China (English)

    袁玉国; 彭秋玲

    2014-01-01

    锌指核酸酶(zinc finger nucleases,ZFNs)技术是近年来发展起来的一种基因修饰技术,利用ZFNs可特异识别靶位点DNA序列并使其断裂.与传统基于同源重组的基因打靶相比,ZFNs能使打靶效率提高100000倍,并使基因断裂与突变修复达到同样效果.ZFNs已经应用于多种生物,包括昆虫、两栖类生物、线虫、植物、多种动物和人类细胞.这些应用提高了人们对复杂生理系统的理解,使ZFNs成为一种生产转基因动物、细胞系和植物的有力工具,并在治疗人类疾病中发挥重要作用.文章对ZFNs技术原理及其应用于哺乳类动物转基因的研究进行了回顾和展望.

  8. Herpes simplex virus induces the marked up-regulation of the zinc finger transcriptional factor INSM1, which modulates the expression and localization of the immediate early protein ICP0

    Directory of Open Access Journals (Sweden)

    Kimura Hiroshi

    2011-05-01

    Full Text Available Abstract Background Herpes simplex viruses (HSVs rapidly shut off macromolecular synthesis in host cells. In contrast, global microarray analyses have shown that HSV infection markedly up-regulates a number of host cell genes that may play important roles in HSV-host cell interactions. To understand the regulatory mechanisms involved, we initiated studies focusing on the zinc finger transcription factor insulinoma-associated 1 (INSM1, a host cell protein markedly up-regulated by HSV infection. Results INSM1 gene expression in HSV-1-infected normal human epidermal keratinocytes increased at least 400-fold 9 h after infection; INSM1 promoter activity was also markedly stimulated. Expression and subcellular localization of the immediate early HSV protein ICP0 was affected by INSM1 expression, and chromatin immunoprecipitation (ChIP assays revealed binding of INSM1 to the ICP0 promoter. Moreover, the role of INSM1 in HSV-1 infection was further clarified by inhibition of HSV-1 replication by INSM1-specific siRNA. Conclusions The results suggest that INSM1 up-regulation plays a positive role in HSV-1 replication, probably by binding to the ICP0 promoter.

  9. The C3H-type zinc finger protein GDS1/C3H42 is a nuclear-speckle-localized protein that is essential for normal growth and development in Arabidopsis.

    Science.gov (United States)

    Kim, Dae Won; Jeon, Su Jeong; Hwang, Sung Min; Hong, Jong Chan; Bahk, Jeong Dong

    2016-09-01

    Eukaryotic C3H-type zinc finger proteins (Znfs) comprise a large family of regulatory proteins involved in many aspects of plant stress response, growth and development. However, compared to mammalian, only a few plant Znfs have been functionally characterized. Here, T-DNA inserted gds1 (growth, development and splicing 1) mutant, displayed abnormal growth throughout the lifecycle owing to the reduction of cell size and number. Inverse PCR analysis revealed that the abnormal growth was caused by the disruption of At3g47120, which encodes a C3H42 protein belonging to the C-X7-C-X5-C-X3-H class of the Znf family. GDS1 was ubiquitously transcribed, but shows high levels of expression in young seedling and unexpanded new leaves. In gds1, the transcripts of many growth- and development-related genes were down-regulated, and the auxin response was dramatically reduced. A fluorescence-based assay revealed that the GDS1 protein was localized to the nucleus, prominently in the speckle compartments. Its arginine/serine dipeptide-rich-like (RS-like) domain was essential for nuclear localization. In addition, the SR1, SRm102 and U1-70K components of the U1 spliceosome interacted with GDS1 in the nuclear speckle compartments. Taken together, these suggest that GDS1, a nuclear-speckle-associated Znf, might play a significant role in splicing during plant growth and development.

  10. CLONING AND EVOLUTIONARY ANALYSIS OF HOMOLOGOUS SEQUENCES OF A NOVEL GENE ENCODING C2H2 ZINC FINGER PROTEIN IN CRUCIFERAE%十字花科植物C2H2型锌指蛋白新基因BcMF20同源序列克隆与进化分析

    Institute of Scientific and Technical Information of China (English)

    韩莹琰; 张爱红; 范双喜; 曹家树

    2011-01-01

    为获取C2H2型锌指蛋白基因在物种间的分类和进化关系,分析其在基因结构上的保守性,更好地研究该基因在植物花粉发育中的功能,根据白菜花粉发育相关基因BcMF20 DNA序列设计引物,运用PCR技术分别从十字花科芸薹属和萝卜属18种材料中克隆了C2H2型锌指蛋白基因BcMF20的同源序列。经序列比对分析,BcMF20同源基因在DNA序列上的相似性为86.9%~100%,所推导的氨基酸序列相似性为77.6%~100%,在锌指的保守区域,氨基酸序列完全相同。这些结果表明该基因具有较好的保守性,可能在十字花科植物的花粉发育中行使重要的功能。%In order to elucidate categorization evolvement of C2H2 zinc finger protein,and clarify mechanism on C2H2 zinc finger protein in pollen development among plants at molecular level,the genes encoding C2H2 zinc finger protein analogues from 18 species of genera Brassicaand Raphanusin Cuciferae were obtained by PCR strategy using specific primers designed from the full length of BcMF20,a putative gene encoding C2H2 zinc finger protein which was related to the male sterility.The phylogenetic relationships of these species belonging to the family Cruciferae were investigated through comparison of the sequences.Homologous sequences of BcMF20comparison indicated that the similarities among the genes at nucleotide and amino acid levels were 86.9% ~ 100% and 77.6%~100%,respectively.In the zinc finger regions of homologous sequences,the amino acid sequences were identical.These results showed that the BcMF20was relative conservation in evolution in Cruciferae,and BcMF20may play an important role in pollen development.

  11. Zinc finger protein ZPR9 functions as an activator of AMPK-related serine/threonine kinase MPK38/MELK involved in ASK1/TGF-β/p53 signaling pathways

    Science.gov (United States)

    Seong, Hyun-A; Manoharan, Ravi; Ha, Hyunjung

    2017-01-01

    Murine protein serine-threonine kinase 38 (MPK38), an AMP‐activated protein kinase (AMPK)-related kinase, has been implicated in the induction of apoptosis signal-regulating kinase 1 (ASK1)-, transforming growth factor-β (TGF‐β)-, and p53-mediated activity involved in metabolic homeostasis. Here, zinc finger protein ZPR9 was found to be an activator of MPK38. The association of MPK38 and ZPR9 was mediated by cysteine residues present in each of these two proteins, Cys269 and Cys286 of MPK38 and Cys305 and Cys308 of ZPR9. MPK38 phosphorylated ZPR9 at Thr252. Wild‐type ZPR9, but not the ZPR9 mutant T252A, enhanced ASK1, TGF‐β, and p53 function by stabilizing MPK38. The requirement of ZPR9 Thr252 phosphorylation was validated using CRISPR/Cas9-mediated ZPR9 (T252A) knockin cell lines. The knockdown of endogenous ZPR9 showed an opposite trend, resulting in the inhibition of MPK38‐dependent ASK1, TGF‐β, and p53 function. This effect was also demonstrated in mouse embryonic fibroblast (MEF) cells that were haploinsufficient (+/−) for ZPR9, NIH 3T3 cells with inducible knockdown of ZPR9, and CRISPR/Cas9-mediated ZPR9 knockout cells. Furthermore, high-fat diet (HFD)-fed mice displayed reduced MPK38 kinase activity and ZPR9 expression compared to that in mice on control chow, suggesting that ZPR9 acts as a physiological activator of MPK38 that may participate in obesity. PMID:28195154

  12. The mouse nac1 gene, encoding a cocaine-regulated Bric-a-brac Tramtrac Broad complex/Pox virus and Zinc finger protein, is regulated by AP1.

    Science.gov (United States)

    Mackler, S A; Homan, Y X; Korutla, L; Conti, A C; Blendy, J A

    2003-01-01

    NAC1 cDNA was identified as a novel transcript induced in the nucleus accumbens from rats chronically treated with cocaine. NAC1 is a member of the Bric-a-brac Tramtrac Broad complex/Pox virus and Zinc finger family of transcription factors and has been shown by overexpression studies to prevent the development of behavioral sensitization resulting from repeated cocaine treatment. This paper reports the cloning and characterization of the corresponding gene. The mouse Nac1 gene consist of six exons, with exon 2 containing an alternative splice donor, providing a molecular explanation of the splice variants observed in mouse and rat. Transcripts of Nac1 were ubiquitously detected in different mouse tissues with prominent expression in the brain. The mouse Nac1 gene was localized to chromosome 8, suggesting a highly plausible candidate gene to explain differences in cocaine-induced behaviors between C57BL6/J and DBA/2J mice that had previously been mapped to the area. In addition, a functional AP1 binding site has been identified in an intron 1 enhancer of the Nac1 gene that plays an essential role in the activation of the gene in differentiation of neuroblastoma cells. Co-transfection with c-jun and c-fos expression plasmids, which encode the two subunits of AP1, activated the wild type Nac1 intron 1 enhancer two-fold over basal, nearly at the level of NAC1 enhancer activity seen in differentiated N2A cells. Mutation of the AP1 site completely abrogated all activation of the NAC1 enhancer in differentiated N2A cells. Activation of immediate early genes such as c-fos and c-jun following chronic drug treatments has been well characterized. The present data describe one potential regulatory cascade involving these transcription factors and activation of NAC1. Identification of drug induced alterations in gene expression is key to understanding the types of molecular adaptations underlying addiction.

  13. Association studies and gene expression analyses of the DISC1-interacting molecules, pericentrin 2 (PCNT2) and DISC1-binding zinc finger protein (DBZ), with schizophrenia and with bipolar disorder.

    Science.gov (United States)

    Anitha, Ayyappan; Nakamura, Kazuhiko; Yamada, Kazuo; Iwayama, Yoshimi; Toyota, Tomoko; Takei, Nori; Iwata, Yasuhide; Suzuki, Katsuaki; Sekine, Yoshimoto; Matsuzaki, Hideo; Kawai, Masayoshi; Thanseem, Ismail; Miyoshi, Ko; Katayama, Taiichi; Matsuzaki, Shinsuke; Baba, Kousuke; Honda, Akiko; Hattori, Tsuyoshi; Shimizu, Shoko; Kumamoto, Natsuko; Kikuchi, Mitsuru; Tohyama, Masaya; Yoshikawa, Takeo; Mori, Norio

    2009-10-05

    Disrupted-in-Schizophrenia 1 (DISC1) and its molecular cascade have been implicated in the pathophysiology of major psychoses. Previously, we identified pericentrin 2 (PCNT2) and DISC1-binding zinc finger protein (DBZ) as binding partners of DISC1; further, we observed elevated expression of PCNT2 in the postmortem brains and in the lymphocytes of bipolar disorder patients, compared to controls. Here, we examined the association of PCNT2 with schizophrenia in a case-control study of Japanese cohorts. We also examined the association of DBZ with schizophrenia and with bipolar disorder, and compared the mRNA levels of DBZ in the postmortem brains of schizophrenia, bipolar and control samples. DNA from 180 schizophrenia patients 201 controls were used for the association study of PCNT2 and DBZ with schizophrenia. Association of DBZ with bipolar disorder was examined in DNA from 238 bipolar patients and 240 age- and gender-matched controls. We observed significant allelic and genotypic associations of the PCNT2 SNPs, rs2249057, rs2268524, and rs2073380 (Ser/Arg) with schizophrenia; the association of rs2249057 (P = 0.002) withstand multiple testing correction. Several two SNP- and three SNP-haplotypes showed significant associations; the associations of haplotypes involving rs2249057 withstand multiple testing correction. No associations were observed for DBZ with schizophrenia or with bipolar disorder; further, there was no significant difference between the DBZ mRNA levels of control, schizophrenia and bipolar postmortem brains. We suggest a possible role of PCNT2 in the pathogenesis of schizophrenia. Abnormalities of PCNT2, the centrosomal protein essential for microtubule organization, may be suggested to lead to neurodevelopmental abnormalities.

  14. An A20/AN1-zinc-finger domain containing protein gene in tea is differentially expressed during winter dormancy and in response to abiotic stress and plant growth regulators

    Directory of Open Access Journals (Sweden)

    Asosii Paul

    2015-03-01

    Full Text Available The present manuscript describes cloning and expression characterization of A20/AN1-zinc-finger domain containing protein (CsZfp gene in an evergreen tree tea [Camellia sinensis (L. O. Kuntze] in response to winter dormancy (WD, abiotic stresses (polyethylene glycol, hydrogen peroxide, and sodium chloride and plant growth regulators [abscisic acid (ABA, and gibberellic acid (GA3]. CsZfp encoded a putative protein of 173 amino acids with a calculated molecular weight of 18.44 kDa, an isoelectric point (pI of 6.50 and grand average of hydropathicity (GRAVY value of −0.334. The gene did not have an intron, and belonged to a multi-gene family. During the period of active growth (PAG, CsZfp showed maximum expression in root and fruit as compared to leaf, floral bud and stem. Interaction studies between temperature and plant growth regulators on the expression of CsZfp showed that ABA upregulated CsZfp expression at growth temperature (GT; 25 °C but had no effect at low temperature (LT; 4 °C. In response to GA3, upregulation was observed at LT but not at GT. Further, the expression was not modulated by LT either in the tissue harvested during PAG or during WD. It was interesting to record that the expression of CsZfp was upregulated by hydrogen peroxide and sodium chloride, whereas it was non-responsive to polyethylene glycol. The possible role of CsZfp in playing key but differential roles in tea to various abiotic stresses is discussed.

  15. Aesthetic Finger Prosthesis

    OpenAIRE

    2011-01-01

    Complete or partial fingers are the most commonly encountered forms of partial hand losses. Though finger amputations are commonly due to traumatic injuries, digit loss may also be attributed to congenital malformations and disease. Irrespective of the etiology, the loss of a finger has a considerable functional and psychological impact on an individual. In order to alleviate these problems, partial or complete finger prosthesis may be fabricated. This clinical report portrays a method to fab...

  16. Nuclear receptors from the ctenophore Mnemiopsis leidyi lack a zinc-finger DNA-binding domain: lineage-specific loss or ancestral condition in the emergence of the nuclear receptor superfamily?

    Directory of Open Access Journals (Sweden)

    Reitzel Adam M

    2011-02-01

    sponges have a similarly restricted NR complement supporting the hypothesis that the original NR was HNF4-like and that these lineages are the first two branches from the animal tree. The absence of a zinc-finger DNA-binding domain in the two ctenophore species suggests two hypotheses: this domain may have been secondarily lost within the ctenophore lineage or, if ctenophores are the first branch off the animal tree, the original NR may have lacked the canonical DBD. Phylogenomic analyses and categorization of NRs from all four early diverging animal phyla compared with the complement from bilaterians suggest the rate of NR diversification prior to the cnidarian-bilaterian split was relatively modest, with independent radiations of several NR subfamilies within the cnidarian lineage.

  17. Aesthetic finger prosthesis.

    Science.gov (United States)

    Shanmuganathan, N; Maheswari, M Uma; Anandkumar, V; Padmanabhan, T V; Swarup, Shailee; Jibran, Ahmed Hasan

    2011-12-01

    Complete or partial fingers are the most commonly encountered forms of partial hand losses. Though finger amputations are commonly due to traumatic injuries, digit loss may also be attributed to congenital malformations and disease. Irrespective of the etiology, the loss of a finger has a considerable functional and psychological impact on an individual. In order to alleviate these problems, partial or complete finger prosthesis may be fabricated. This clinical report portrays a method to fabricate silicone rubber prosthesis for a patient who has a partial finger loss caused due to trauma.

  18. 小麦锌指蛋白基因TaLOL2的克隆及特征分析%Cloning and Characterization of a Zinc Finger Protein Gene TaLOL2 in Wheat

    Institute of Scientific and Technical Information of China (English)

    白鹏飞; 杨倩; 康振生; 郭军

    2012-01-01

    通过电子克隆与RT-PCR相结合的方法,在条锈菌诱导的小麦叶片中克隆获得1个新的LSD1型锌指蛋白基因TaLOL2,并用qRT PCR技术分析了其转录表达特征.结果显示:(1)小麦锌指蛋白基因TaLOL2的cDNA全长1 095 bp,编码1 79个氨基酸.(2)TaLOL2含有3个典型的zf-LSD1型(CxxCxRxxLMYxxGASxVxCxxC)保守结构域,与水稻、拟南芥、大麦等植物LSD1型锌指蛋白序列具有高度相似性,其中与水稻OsLOL2相似度达86.0%.(3)进化树分析表明,TaLOL2与水稻、拟南芥和大麦中部分含有3个保守zf-LSD1锌指结构的基因亲缘关系较近,而与其它包含不同数目的zf-LSD1锌指结构的基因亲缘关系较远.(4)qRT-PCR定量分析表明,Ta LOL2在条锈菌侵染前期呈上调表达,在亲和及非亲和反应中差异表达.研究表明,TaLOL2参与了条锈菌诱导的小麦抗病防卫反应,很可能作为正调控因子参与了小麦-条锈菌非亲和互作中对条锈菌的抗性信号途径.%A LSD1-type zinc finger protein gene, TaLOL2, was cloned from the wheat leaves infected by stripe rust fungus (Puccinia striiformis f. sp. tritici,Pst) using in silico cloning and RT-PCR and its expression pattern was analyzed through qRT-PCR. (1) The full cDNA length of TaLOL2 was 1 095 bp, which encodes a 179 amino acid protein. (2)Analysis of the protein domain features indicated that TaLOL2 contains three internally conserved zf-LSD1 domains defined as CxxCxRxxLMYxxGASxVxCxxC. Multi-sequence alignment showed that TaLOL2 shares high similarity with other LSD1-type proteins in rice (Oryza sativa) ,Arabidopsis thaliana and barley (Hordeum vulgare) ,and is 86. 0% identical to the rice OsLOL2. (3)Phylogenetic analysis of the zf-LSD1-containing proteins in plants demonstrated that TaLOL2 clusters with three zf-LSD1 containing proteins and shares highly homology with OsLOL2 and HvLSD1-likel. (4) Transcriptional analysis of TaLOL2 suggested that TaLOL2 was highly induced by Pst during the early

  19. Functional Analysis of the SUPERMAN-like Zinc Finger Protein Gene GZFP and Its Promoter%棉花SUPERMAN类锌指蛋白基因GZFP的启动子及功能分析

    Institute of Scientific and Technical Information of China (English)

    杨郁文; 周建武; 张保龙; 范晓慧; 任永哲; 陈天子

    2011-01-01

    GZFP是来源于陆地棉的一个SUPERMAN类锌指蛋白基因.为了进一步了解该基因功能,本研究通过染色体步移(Genome walking)获得其启动子区域,并构建该片段与GUS连接的重组载体pBI-pGZFP-5::GUS,通过花浸染法转化拟南芥.转基因植株的GUS染色结果表明GUS基因主要在根部以及花器官表达,而叶片中的表达量很低,并且根部的表达量在整个生育期都很强.通过半定量RT-PCR分析不同诱导下GZFP基因的表达变化,发现ABA、干旱、盐胁迫诱导可以提高其表达量.构建GZFP过量表达载体并通过叶盘法转化烟草,烟草转化株的表型无明显变化,但是转化株中NtOPBP1和NtERD10的表达量较未转化株明显增强,而这两个基因都参与了植物的抗逆反应,说明GZFP基因可能与植物抗逆相关.%GZFP, derived from Gossypium hirsutum L, belongs to the SUPERMAN-like zinc finger protein. To further analyze the function of GZFP, the promoter region of GZFP was obtained by genome walking. The recombination vector pBI-pGZFP-5::GUS containing the promoter was constructed and transformed to Aiabidopsis by Agrobacterium-mediated method. GUS staining showed GZFP promoter driven GUS activity was specifically detected in roots and flowers and the activity was very low in leaves. In addition, the GUS activity in roots was very strong throughout the whole growth period. By semi-quantitative RT-PCR, we have found the expression of GZFP was induced by ABA, drought and salt. The over expression vector was constructed by inserting GZFP into the pCAMBIA2301, tobacco plants were transformed by co-cultivating leaves method via A-grobacterium mediation. The object gene was verified to have been integrated into the genome of tobacco by PCR. The pheno-type of transgenic plants was normal compared to the wild type, but the expression of NtOPBP\\ and NtERD10 was relatively higher than the wild type. As the two genes involved in the stress reaction, GZFP may have

  20. Gold finger formation studied by high-resolution mass spectrometry and in silico methods

    NARCIS (Netherlands)

    Laskay, Ü.A.; Garino, C.; Tsybin, Y.O.; Salassa, L.; Casini, A.

    2015-01-01

    High-resolution mass spectrometry and quantum mechanics/molecular mechanics studies were employed for characterizing the formation of two gold finger (GF) domains from the reaction of zinc fingers (ZF) with gold complexes. The influence of both the gold oxidation state and the ZF coordination sphere

  1. Multiple Fingers - One Gestalt.

    Science.gov (United States)

    Lezkan, Alexandra; Manuel, Steven G; Colgate, J Edward; Klatzky, Roberta L; Peshkin, Michael A; Drewing, Knut

    2016-01-01

    The Gestalt theory of perception offered principles by which distributed visual sensations are combined into a structured experience ("Gestalt"). We demonstrate conditions whereby haptic sensations at two fingertips are integrated in the perception of a single object. When virtual bumps were presented simultaneously to the right hand's thumb and index finger during lateral arm movements, participants reported perceiving a single bump. A discrimination task measured the bump's perceived location and perceptual reliability (assessed by differential thresholds) for four finger configurations, which varied in their adherence to the Gestalt principles of proximity (small versus large finger separation) and synchrony (virtual spring to link movements of the two fingers versus no spring). According to models of integration, reliability should increase with the degree to which multi-finger cues integrate into a unified percept. Differential thresholds were smaller in the virtual-spring condition (synchrony) than when fingers were unlinked. Additionally, in the condition with reduced synchrony, greater proximity led to lower differential thresholds. Thus, with greater adherence to Gestalt principles, thresholds approached values predicted for optimal integration. We conclude that the Gestalt principles of synchrony and proximity apply to haptic perception of surface properties and that these principles can interact to promote multi-finger integration.

  2. Pathogen-induced binding of the soybean zinc finger homeodomain proteins GmZF-HD1 and GmZF-HD2 to two repeats of ATTA homeodomain binding site in the calmodulin isoform 4 (GmCaM4) promoter.

    Science.gov (United States)

    Park, Hyeong Cheol; Kim, Man Lyang; Lee, Sang Min; Bahk, Jeong Dong; Yun, Dae-Jin; Lim, Chae Oh; Hong, Jong Chan; Lee, Sang Yeol; Cho, Moo Je; Chung, Woo Sik

    2007-01-01

    Calmodulin (CaM) is involved in defense responses in plants. In soybean (Glycine max), transcription of calmodulin isoform 4 (GmCaM4) is rapidly induced within 30 min after pathogen stimulation, but regulation of the GmCaM4 gene in response to pathogen is poorly understood. Here, we used the yeast one-hybrid system to isolate two cDNA clones encoding proteins that bind to a 30-nt A/T-rich sequence in the GmCaM4 promoter, a region that contains two repeats of a conserved homeodomain binding site, ATTA. The two proteins, GmZF-HD1 and GmZF-HD2, belong to the zinc finger homeodomain (ZF-HD) transcription factor family. Domain deletion analysis showed that a homeodomain motif can bind to the 30-nt GmCaM4 promoter sequence, whereas the two zinc finger domains cannot. Critically, the formation of super-shifted complexes by an anti-GmZF-HD1 antibody incubated with nuclear extracts from pathogen-treated cells suggests that the interaction between GmZF-HD1 and two homeodomain binding site repeats is regulated by pathogen stimulation. Finally, a transient expression assay with Arabidopsis protoplasts confirmed that GmZF-HD1 can activate the expression of GmCaM4 by specifically interacting with the two repeats. These results suggest that the GmZF-HD1 and -2 proteins function as ZF-HD transcription factors to activate GmCaM4 gene expression in response to pathogen.

  3. 锌指蛋白1对大鼠肝纤维化转化生长因子β的影响%Role of zinc finger protein 1 in rat liver fibrosis and as related to TGFβ expression

    Institute of Scientific and Technical Information of China (English)

    沈宇娟; 陈永平; 刘真真; 王晓东; 戴春蕾; 林镯

    2014-01-01

    Objective To determine the role of zinc finger protein 1 (ZEB 1) in liver fibrosis and in regards to expression of the tumor growth factor-beta (TGFβ) signaling factor using a rat model system.Methods Sprague-Dawley rats were randomly divided into a normal (control) group,liver fibrosis (model) group and a liver fibrosis + therapy (ZEB1 intervention) group.The model group and the ZEB1 intervention group were given intraperitoneal injections of dimethylnitrosamine (DMN) for the first 3 days of each week over a 7-week period; starting at week 5,the ZEB 1 intervention group was started on a routine of every other day tail vein injections of recombinant ZEB1.During this 7-week period,the control group was given intraperitoneal injections of 0.9% NaC1 alone on the DMN schedule.Liver tissues were collected for pathological examination (with hematoxylin-eosin and Masson staining) and for detection of TGFβ1 and ZEB 1 expression (by RT-PCR and western blotting).Measurement data were compared between groups using the single-factor analysis of variance test,followed by the least significant difference LSD test.Count data were analyzed by Fisher's exact test.Results The model group's liver tissues showed degeneration and necrosis,as well as obvious fibrous septa accompanied by pseudo lobules.The ZEB 1 intervention group's liver tissues showed a significantly higher degree of fibrosis (x2 =21.63,P =0),with more coarse fiber cords.The expression of ZEB1 and TGFβ1 was significantly higher in the model group than in the control group (both P < 0.05).However,the ZEB 1 intervention group showed the highest levels of ZEB 1 and TGFβ1 expression (vs.model group,P < 0.05).Conclusion ZEB 1 may promote the development of liver fibrosis in rats through a mechanism involving the TGFβ/Smad signaling pathway.%目的 观察锌指蛋白(ZEB)1在大鼠肝纤维化模型中的表达情况及探讨重组ZEB1蛋白干预对纤维化程度的影响.方法 36只健康SD大鼠随机分为

  4. Nickel transfer by fingers.

    Science.gov (United States)

    Isnardo, D; Vidal, J; Panyella, D; Vilaplana, J

    2015-06-01

    We investigated fingers as a potential source of nickel transfer to the face in patients with allergic contact dermatitis to nickel and a history of facial dermatitis. Samples were collected from the fingers and cheeks of volunteers using the stripping method with standard adhesive tape, and nickel levels were quantified using mass spectrometry. Fingers and cheeks of individuals who had handled coins were both positive for nickel, with levels ranging from 14.67 to 58.64 ppm and 1.28 to 8.52 ppm, respectively. The levels in a control group were considerably and significantly lower. Transfer of nickel from a person's fingers to their face after handling a nickel-containing object could explain the presence of facial dermatitis in patients with nickel hypersensitivity.

  5. Cross finger flaps.

    Science.gov (United States)

    Kisner, W H

    1979-01-01

    Proper fingertip reconstruction requires good skin and soft tissue coverage, preservation of function and as normal an appearance as possible. The cross finger flap results in negligible joint stiffness, minimal morbidity and little work-time loss. An important factor is the conservation of finger length permitted by this technique. This method of repair is underutilized. It is indicated in several types of fingertip amputations where bone shortening would be detrimental.

  6. Tendon Driven Finger Actuation System

    Science.gov (United States)

    Ihrke, Chris A. (Inventor); Reich, David M. (Inventor); Bridgwater, Lyndon (Inventor); Linn, Douglas Martin (Inventor); Askew, Scott R. (Inventor); Diftler, Myron A. (Inventor); Platt, Robert (Inventor); Hargrave, Brian (Inventor); Valvo, Michael C. (Inventor); Abdallah, Muhammad E. (Inventor); Permenter, Frank Noble (Inventor); Mehling, Joshua S. (Inventor)

    2013-01-01

    A humanoid robot includes a robotic hand having at least one finger. An actuation system for the robotic finger includes an actuator assembly which is supported by the robot and is spaced apart from the finger. A tendon extends from the actuator assembly to the at least one finger and ends in a tendon terminator. The actuator assembly is operable to actuate the tendon to move the tendon terminator and, thus, the finger.

  7. Arsenite binding-induced zinc loss from PARP-1 is equivalent to zinc deficiency in reducing PARP-1 activity, leading to inhibition of DNA repair

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Xi; Zhou, Xixi [Department of Pharmaceutical Sciences, College of Pharmacy, University of New Mexico Health Sciences Center, Albuquerque, NM 87131 (United States); Du, Libo [Center for Molecular Science, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Liu, Wenlan [Department of Pharmaceutical Sciences, College of Pharmacy, University of New Mexico Health Sciences Center, Albuquerque, NM 87131 (United States); Liu, Yang [Center for Molecular Science, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Hudson, Laurie G. [Department of Pharmaceutical Sciences, College of Pharmacy, University of New Mexico Health Sciences Center, Albuquerque, NM 87131 (United States); Liu, Ke Jian, E-mail: kliu@salud.unm.edu [Department of Pharmaceutical Sciences, College of Pharmacy, University of New Mexico Health Sciences Center, Albuquerque, NM 87131 (United States)

    2014-01-15

    Inhibition of DNA repair is a recognized mechanism for arsenic enhancement of ultraviolet radiation-induced DNA damage and carcinogenesis. Poly(ADP-ribose) polymerase-1 (PARP-1), a zinc finger DNA repair protein, has been identified as a sensitive molecular target for arsenic. The zinc finger domains of PARP-1 protein function as a critical structure in DNA recognition and binding. Since cellular poly(ADP-ribosyl)ation capacity has been positively correlated with zinc status in cells, we hypothesize that arsenite binding-induced zinc loss from PARP-1 is equivalent to zinc deficiency in reducing PARP-1 activity, leading to inhibition of DNA repair. To test this hypothesis, we compared the effects of arsenite exposure with zinc deficiency, created by using the membrane-permeable zinc chelator TPEN, on 8-OHdG formation, PARP-1 activity and zinc binding to PARP-1 in HaCat cells. Our results show that arsenite exposure and zinc deficiency had similar effects on PARP-1 protein, whereas supplemental zinc reversed these effects. To investigate the molecular mechanism of zinc loss induced by arsenite, ICP-AES, near UV spectroscopy, fluorescence, and circular dichroism spectroscopy were utilized to examine arsenite binding and occupation of a peptide representing the first zinc finger of PARP-1. We found that arsenite binding as well as zinc loss altered the conformation of zinc finger structure which functionally leads to PARP-1 inhibition. These findings suggest that arsenite binding to PARP-1 protein created similar adverse biological effects as zinc deficiency, which establishes the molecular mechanism for zinc supplementation as a potentially effective treatment to reverse the detrimental outcomes of arsenic exposure. - Highlights: • Arsenite binding is equivalent to zinc deficiency in reducing PARP-1 function. • Zinc reverses arsenic inhibition of PARP-1 activity and enhancement of DNA damage. • Arsenite binding and zinc loss alter the conformation of zinc finger

  8. 青花菜C3H型锌指蛋白基因 BoCCCH2的克隆与表达%Cloning and expression of a C3H-type zinc finger protein gene BoCCCH2 from Brassica oleracea var . italica

    Institute of Scientific and Technical Information of China (English)

    蒋明; 刘青娥; 章燕如; 祝琦; 龚秀; 俞可可; 周秀倩

    2016-01-01

    以青花菜为材料,在克隆C3 H型锌指蛋白基因 BoCCC H2的基础上,研究该基因在不同器官及霜霉菌和灰葡萄孢菌侵染叶片中的表达模式。测序结果表明,BoCCC H2没有内含子,编码区全长为1740 bp ,编码579个氨基酸,推导的蛋白质具2个 ANK结构域和2种CCCH 锌指结构,锌指结构的类型分别为C—X8—C—X5—C—X3—H和C—X5—C—X4—C—X3—H 。反转录聚合酶链反应表明:BoCCC H2在根、叶、花茎、嫩角果、花蕾和花中均有表达,其中在根中的表达量最高;经霜霉菌和灰葡萄孢菌侵染后,该基因表达量均有不同程度的增加,其中在霜霉菌侵染下,表达量在24 h后开始增加,72 h时下降,而在灰葡萄孢菌侵染下,6 h时的表达量最大,12 h时开始缓慢下降。聚类结果表明,BoCCCH2与其他十字花科植物的同源序列聚为一类,支持率达100%,而与豆科、大戟科和蔷薇科等植物的序列处于不同分支。对 BoCCC H2基因的克隆和表达分析为该基因功能研究奠定了基础。%Summary Brassicaoleraceavar.italicaisanimportantvegetablecropworldwide,andinChina,TaizhouCity of Zhejiang Province is one of the major broccoli production areas . Downy mildew and grey mold rot are two common fungal diseases caused by Hyaloperonospora parasitica and Botrytis cinerea , respectively . In recent years , broccoli production in Taizhou was frequently affected by these two fungal diseases , resulting in yield and quality loss . Broccoli germplasm resources resistance to disease is scarce; therefore , molecular breeding is regarded as an effective solution to solve the problem . This is critically important to isolate genes associated with disease resistance , which will act as potential target genes for broccoli breeding . Zinc finger proteins are kinds of important transcription factors in eukaryotic organisms , which involve in various biological activities , such as replication , transcription

  9. Three-Fingered Robot Hand

    Science.gov (United States)

    Ruoff, C. F.; Salisbury, J. K.

    1984-01-01

    Mechanical joints and tendons resemble human hand. Robot hand has three "human-like" fingers. "Thumb" at top. Rounded tips of fingers covered with resilient material provides high friction for griping. Hand potential as prosthesis for humans.

  10. Mixing methods, tasting fingers

    DEFF Research Database (Denmark)

    Mann, Anna; Mol, Annemarie; Satalkar, Priya;

    2011-01-01

    This article reports on an ethnographic experiment. Four finger eating experts and three novices sat down for a hot meal and ate with their hands. Drawing on the technique of playing with the familiar and the strange, our aim was not to explain our responses, but to articulate them. As we seek wo...

  11. Finger cold induced vasodilation

    NARCIS (Netherlands)

    Daanen, H.A.M.

    2007-01-01

    There are indications that subjects with a reduced finger CIVD response are more prone to get local cold injuries, but more epidemiological research is needed to establish a firm relationship. Although it was observed that an early CIVD onset was associated with initially superior manual performance

  12. X-Ray Exam: Finger

    Science.gov (United States)

    ... Old Feeding Your 1- to 2-Year-Old X-Ray Exam: Finger KidsHealth > For Parents > X-Ray Exam: Finger A A A What's in ... español Radiografía: dedo What It Is A finger X-ray is a safe and painless test that ...

  13. Design of lanthanide fingers: compact lanthanide-binding metalloproteins.

    Science.gov (United States)

    am Ende, Christopher W; Meng, Hai Yun; Ye, Mao; Pandey, Anil K; Zondlo, Neal J

    2010-08-16

    Lanthanides have interesting chemical properties; these include luminescent, magnetic, and catalytic functions. Toward the development of proteins incorporating novel functions, we have designed a new lanthanide-binding motif, lanthanide fingers. These were designed based on the Zif268 zinc finger, which exhibits a beta beta alpha structural motif. Lanthanide fingers utilize an Asp(2)Glu(2) metal-coordination environment to bind lanthanides through a tetracarboxylate peptide ligand. The iterative design of a general lanthanide-binding peptide incorporated the following key elements: 1) residues with high alpha-helix and beta-sheet propensities in the respective secondary structures; 2) an optimized big box alpha-helix N-cap; 3) a Schellman alpha-helix C-cap motif; and 4) an optional D-Pro-Ser type II' beta-turn in the beta-hairpin. The peptides were characterized for lanthanide binding by circular dichroism (CD), NMR, and fluorescence spectroscopy. In all instances, stabilization of the peptide secondary structures resulted in an increase in metal affinity. The optimized protein design was a 25-residue peptide that was a general lanthanide-binding motif; this binds all lanthanides examined in a competitive aqueous environment, with a dissociation constant of 9.3 microM for binding Er(3+). CD spectra of the peptide-lanthanide complexes are similar to those of zinc fingers and other beta beta alpha proteins. Metal binding involves residues from the N-terminal beta-hairpin and the C terminal alpha-helical segments of the peptide. NMR data indicated that metal binding induced a global change in the peptide structure. The D-Pro-Ser type II' beta-turn motif could be replaced by Thr-Ile to generate genetically encodable lanthanide fingers. Replacement of the central Phe with Trp generated genetically encodable lanthanide fingers that exhibited terbium luminescence greater than that of an EF-hand peptide.

  14. Finger vein recognition based on finger crease location

    Science.gov (United States)

    Lu, Zhiying; Ding, Shumeng; Yin, Jing

    2016-07-01

    Finger vein recognition technology has significant advantages over other methods in terms of accuracy, uniqueness, and stability, and it has wide promising applications in the field of biometric recognition. We propose using finger creases to locate and extract an object region. Then we use linear fitting to overcome the problem of finger rotation in the plane. The method of modular adaptive histogram equalization (MAHE) is presented to enhance image contrast and reduce computational cost. To extract the finger vein features, we use a fusion method, which can obtain clear and distinguishable vein patterns under different conditions. We used the Hausdorff average distance algorithm to examine the recognition performance of the system. The experimental results demonstrate that MAHE can better balance the recognition accuracy and the expenditure of time compared with three other methods. Our resulting equal error rate throughout the total procedure was 3.268% in a database of 153 finger vein images.

  15. Emotional Communication in Finger Braille

    Directory of Open Access Journals (Sweden)

    Yasuhiro Matsuda

    2010-01-01

    Full Text Available We describe analyses of the features of emotions (neutral, joy, sadness, and anger expressed by Finger Braille interpreters and subsequently examine the effectiveness of emotional expression and emotional communication between people unskilled in Finger Braille. The goal is to develop a Finger Braille system to teach emotional expression and a system to recognize emotion. The results indicate the following features of emotional expression by interpreters. The durations of the code of joy were significantly shorter than the durations of the other emotions, the durations of the code of sadness were significantly longer, and the finger loads of anger were significantly larger. The features of emotional expression by unskilled subjects were very similar to those of the interpreters, and the coincidence ratio of emotional communication was 75.1%. Therefore, it was confirmed that people unskilled in Finger Braille can express and communicate emotions using this communication medium.

  16. Zinc Deficiency in Humans and its Amelioration

    OpenAIRE

    Yashbir Singh Shivay

    2015-01-01

    Zinc (Zn) deficiency in humans has recently received considerable attention. Global mortality in children under 5 years of age in 2004 due to Zn deficiency was estimated at 4,53,207 as against 6,66,771 for vitamin A deficiency; 20,854 for iron deficiency and 3,619 for iodine deficiency. In humans 2800-3000 proteins contain Zn prosthetic group and Zn is an integral component of zinc finger prints that regulate DNA transcription. Zinc is a Type-2 nutrient, which means that its concentration in ...

  17. Covering the Dorsal Finger Defect with Reverse Cross Finger Flap

    Directory of Open Access Journals (Sweden)

    Kaan Gurbuz

    2014-12-01

    Full Text Available Reconstruction of finger extensor zone defects with or without tendon gaps still remains a challenge for surgeons. Although surgical treatments may differ, and range from the use of local, regional, to free flaps, the outcomes for all cases are not satisfactory. In this case report, we present a case of a 3rd finger extensor side crush injury including a defect of Dd (Digit Dorsal 1, Dd2 and Dd3 defects of extensor zones with tendon gap. Tendon gap was reconstructed using m. palmaris longus tendon graft and the defect was covered with reversed cross-finger flap (random pattern with good cosmetic and excellent functional results.

  18. Steadily translating parabolic dissolution fingers

    CERN Document Server

    Kondratiuk, Paweł

    2015-01-01

    Dissolution fingers (or wormholes) are formed during the dissolution of a porous rock as a result of nonlinear feedbacks between the flow, transport and chemical reactions at pore surfaces. We analyze the shapes and growth velocities of such fingers within the thin-front approximation, in which the reaction is assumed to take place instantaneously with the reactants fully consumed at the dissolution front. We concentrate on the case when the main flow is driven by the constant pressure gradient far from the finger, and the permeability contrast between the inside and the outside of the finger is finite. Using Ivantsov ansatz and conformal transformations we find the family of steadily translating fingers characterized by a parabolic shape. We derive the reactant concentration field and the pressure field inside and outside of the fingers and show that the flow within them is uniform. The advancement velocity of the finger is shown to be inversely proportional to its radius of curvature in the small P\\'{e}clet...

  19. Finger Forces in Clarinet Playing.

    Science.gov (United States)

    Hofmann, Alex; Goebl, Werner

    2016-01-01

    Clarinettists close and open multiple tone holes to alter the pitch of the tones. Their fingering technique must be fast, precise, and coordinated with the tongue articulation. In this empirical study, finger force profiles and tongue techniques of clarinet students (N = 17) and professional clarinettists (N = 6) were investigated under controlled performance conditions. First, in an expressive-performance task, eight selected excerpts from the first Weber Concerto were performed. These excerpts were chosen to fit in a 2 × 2 × 2 design (register: low-high; tempo: slow-fast, dynamics: soft-loud). There was an additional condition controlled by the experimenter, which determined the expression levels (low-high) of the performers. Second, a technical-exercise task, an isochronous 23-tone melody was designed that required different effectors to produce the sequence (finger-only, tongue-only, combined tongue-finger actions). The melody was performed in three tempo conditions (slow, medium, fast) in a synchronization-continuation paradigm. Participants played on a sensor-equipped Viennese clarinet, which tracked finger forces and reed oscillations simultaneously. From the data, average finger force (F mean ) and peak force (F max ) were calculated. The overall finger forces were low (F mean = 1.17 N, F max = 3.05 N) compared to those on other musical instruments (e.g., guitar). Participants applied the largest finger forces during the high expression level performance conditions (F mean = 1.21 N). For the technical exercise task, timing and articulation information were extracted from the reed signal. Here, the timing precision of the fingers deteriorated the timing precision of the tongue for combined tongue-finger actions, especially for faster tempi. Although individual finger force profiles were overlapping, the group of professional players applied less finger force overall (F mean = 0.54 N). Such sensor instruments provide useful insights into player

  20. Finger Forces in Clarinet Playing

    Directory of Open Access Journals (Sweden)

    Alex Hofmann

    2016-08-01

    Full Text Available Clarinettists close and open multiple tone holes to alter the pitch of the tones. Their fingering technique must be fast, precise, and coordinated with the tongue articulation. In this empirical study, finger force profiles and tongue techniques of clarinet students (N = 17 and professional clarinettists (N = 6 were investigated under controlled performance conditions. First, in an expressive-performance task, eight selected excerpts from the first Weber Concerto were performed. These excerpts were chosen to fit in a 2 x 2 x 2 design (register: low--high; tempo: slow--fast, dynamics: soft--loud. There was an additional condition controlled by the experimenter, which determined the expression levels (low--high of the performers. Second, a technical-exercise task, an isochronous 23-tone melody was designed that required different effectors to produce the sequence (finger-only, tongue-only, combined tongue-finger actions. The melody was performed in three tempo conditions (slow, medium, fast in a synchronization-continuation paradigm. Participants played on a sensor-equipped Viennese clarinet, which tracked finger forces and reed oscillations simultaneously. From the data, average finger force (Fmean and peak force (Fmax were calculated. The overall finger forces were low (Fmean = 1.17 N, Fmax = 3.05 N compared to those on other musical instruments (e.g. guitar. Participants applied the largest finger forces during the high expression level performance conditions (Fmean = 1.21 N.For the technical exercise task, timing and articulation information were extracted from the reed signal. Here, the timing precision of the fingers deteriorated the timing precision of the tongue for combined tongue-finger actions, especially for faster tempi. Although individual finger force profiles were overlapping, the group of professional players applied less finger force overall (Fmean = 0.54 N. Such sensor instruments provide useful insights into player

  1. Viscous fingering of miscible slices

    CERN Document Server

    De Wit, A; Martin, M; Wit, Anne De; Bertho, Yann; Martin, Michel

    2005-01-01

    Viscous fingering of a miscible high viscosity slice of fluid displaced by a lower viscosity fluid is studied in porous media by direct numerical simulations of Darcy's law coupled to the evolution equation for the concentration of a solute controlling the viscosity of miscible solutions. In contrast with fingering between two semi-infinite regions, fingering of finite slices is a transient phenomenon due to the decrease in time of the viscosity ratio across the interface induced by fingering and dispersion processes. We show that fingering contributes transiently to the broadening of the peak in time by increasing its variance. A quantitative analysis of the asymptotic contribution of fingering to this variance is conducted as a function of the four relevant parameters of the problem i.e. the log-mobility ratio R, the length of the slice l, the Peclet number Pe and the ratio between transverse and axial dispersion coefficients $\\epsilon$. Relevance of the results is discussed in relation with transport of vi...

  2. Association analysis of zinc finger protein 804A gene with bipolar disorders and its subtypes in Han Chinese%锌指蛋白804A基因与中国汉族双相障碍及其亚型的关联分析

    Institute of Scientific and Technical Information of China (English)

    张晨; 汪作为; 吴志国; 洪武; 陈俊; 彭代辉; 方贻儒

    2015-01-01

    Objective To investigate the potential association between zinc finger protein 804A (ZNF804A) rs1344706 polymorphism and bipolar disorder as well as its subtypes.Methods A total of 304 bipolar Ⅰ disorder,442 bipolar Ⅱ disorder and 762 healthy controls subjects were recruited from Division of Mood Disorders,Shanghai Mental Health Center,Shanghai Jiao Tong University School of Medicine,from November 2006 to August 2014.Rs1344706 was genotyped by SNaPshot genotyping assay.Results In comparison between the patients with bipolar disorder and controls,there were significant differences in the genotypic (TT:210(28.2%) vs.175(23.0%),TG:378(50.7%) vs.399(52.4%),GG:158 (21.2%) vs.188(24.7%)) and allelic (T:798(53.5%) vs.749(49.1%),G:694(46.5%) vs.775(50.9%)) distributions (x2=6.18,P=0.04,x2=5.68,P=0.017,respectively).The frequency of T allele in patients with bipolar disorder was higher than that in controls (OR=1.19,95%CI 1.03-1.37).After stratified by diagnosed subtypes,there were significant differences in the genotypic and allelic distribution between patients with bipolar Ⅰ disorder and controls (x2=7.57,P=0.02,x2=7.22,P=0.007).The frequency of T allele in patients with bipolar Ⅰ disorder was higher than those in controls (OR=1.30,95% CI 1.07-1.56).In comparison between patients with bipolar Ⅱ disorder and controls,there was no significant difference in the distributions of genotype and allele.Conclusion There is positive association between rs1344706 polymorphism and bipolar Ⅰ disorder in a Chinese Han population.ZNF804A gene may confer susceptibility to bipolar Ⅰ disorder in Han Chinese.%目的 探讨位于锌指蛋白804A(zinc finger protein 804A,ZNF804A)内含子2区的多态性位点rs 1344706与双相障碍及其亚型的关系.方法 收集2006年11月至2014年8月在上海交通大学医学院附属精神卫生中心心境障碍科就诊的双相障碍患者746例(患者组),根据DSM-Ⅳ中双相障碍诊断亚型

  3. An Underactuated Multi-finger Grasping Device

    Directory of Open Access Journals (Sweden)

    Cesare Rossi

    2014-02-01

    Full Text Available In this paper, a mechanical model for an underactuated multi-finger grasping device is presented. The device has single-tendon, three-phalanx fingers, all moved by only one actuator. By means of the model, both the kinematic and dynamical behaviour of the finger itself can be studied. The finger is part of a more complex mechanical system that consists of a four-finger grasping device for robots or a five-finger human hand prosthesis. Some results of both the kinematic and dynamical behaviour are also presented.

  4. Mesofluidic controlled robotic or prosthetic finger

    Energy Technology Data Exchange (ETDEWEB)

    Lind, Randall F; Jansen, John F; Love, Lonnie J

    2013-11-19

    A mesofluidic powered robotic and/or prosthetic finger joint includes a first finger section having at least one mesofluidic actuator in fluid communication with a first actuator, a second mesofluidic actuator in fluid communication with a second actuator and a second prosthetic finger section pivotally connected to the first finger section by a joint pivot, wherein the first actuator pivotally cooperates with the second finger to provide a first mechanical advantage relative to the joint point and wherein the second actuator pivotally cooperates with the second finger section to provide a second mechanical advantage relative to the joint point.

  5. Mechanical model of a single tendon finger

    Science.gov (United States)

    Rossi, Cesare; Savino, Sergio

    2013-10-01

    The mechanical model of a single tendon three phalanxes finger is presented. By means of the model both kinematic and dynamical behavior of the finger itself can be studied. This finger is a part of a more complex mechanical system that consists in a four finger grasping device for robots or in a five finger human hand prosthesis. A first prototype has been realized in our department in order to verify the real behavior of the model. Some results of both kinematic and dynamical behavior are presented.

  6. FINGER-VEIN RECOGNITION SYSTEMS

    Directory of Open Access Journals (Sweden)

    A.Haritha Deepthi

    2015-10-01

    Full Text Available As the Person‟s/Organization‟s Private information‟s are becoming very easy to access, the demand for a Simple, Convenient, Efficient, and a highly Securable Authentication System has been increased. In considering these requirements for data Protection, Biometrics, which uses human physiological or behavioral system for personal Identification has been found as a solution for these difficulties. However most of the biometric systems have high complexity in both time and space. So we are going to use a Real time Finger-Vein recognition System for authentication purposes. In this paper we had implemented the Finger Vein Recognition concept using MATLAB R2013a. The features used are Lacunarity Distance, Blanket Dimension distance. This has more accuracy when compared to conventional methods.

  7. Finger Motion Decoding Using EMG Signals Corresponding Various Arm Postures

    OpenAIRE

    You, Kyung-Jin; Rhee, Ki-Won; Shin, Hyun-Chool

    2010-01-01

    We provide a novel method to infer finger flexing motions using a four-channel surface electromyogram (EMG). Surface EMG signals can be recorded from the human body non-invasively and easily. Surface EMG signals in this study were obtained from four channel electrodes placed around the forearm. The motions consist of the flexion of five single fingers (thumb, index finger, middle finger, ring finger, and little finger) and three multi.finger motions. The maximum likelihood estimation was used...

  8. Reconstruction of finger pulp defect with reversed fasciocutaneous island flap from same finger

    Institute of Scientific and Technical Information of China (English)

    LU Yao-jun; HONG Guang-xiang; XU Nan-wei; HU Zhi-yong; SHAO Lei

    2007-01-01

    Objective: To investigate the clinical curative effect of reconstruction of finger pulp defect by anastomosis of reversed fasciocutaneous island flap with dorsal branch of the digital nerve of the same finger. Methods: The restoration of finger pulp defect with fasciocutaneous island flap from the same finger was conducted in 25 cases (30 fingers) from January 2002 to June 2003. Nine patients (11 fingers) whose flaps with dorsal branch of the digital nerve anastomosed with the digital inherent nerve around the surface of the wound were Group A and the others were Group B. The follow-up was carried out at 3 and 9 months after the operation to observe the shape of finger pulp and the sense restoration between two groups.Results: All flaps of 25 cases (30 fingers) survived. Three months after operation, the patients had fully grown finger pulps and recovered the superficial sensation and tactile sense of finger pulps. The two point discrimination on average was 5.00 mm ± 0.23 mm in Group A and 6.00 mm ± 0.30 mm in Group B. The difference between two groups was highly significant. Nine months later, their senses of finger pulps between two groups were recovered basically. Conclusions:The reversed fasciocutaneous island flap from the same finger is the first choice to reconstruct the finger pulp defect, and the anastomosis of dorsal branch of the digital nerve shall be determined according to the specific condition.

  9. Current status of ultrasonography of the finger

    Directory of Open Access Journals (Sweden)

    Seun Ah Lee

    2016-04-01

    Full Text Available The recent development of advanced high-resolution transducers has enabled the fast, easy, and dynamic ultrasonographic evaluation of small, superficial structures such as the finger. In order to best exploit these advances, it is important to understand the normal anatomy and the basic pathologies of the finger, as exemplified by the following conditions involving the dorsal, volar, and lateral sections of the finger: sagittal band injuries, mallet finger, and Boutonnière deformity (dorsal aspect; flexor tendon tears, trigger finger, and volar plate injuries (volar aspect; gamekeeper’s thumb (Stener lesions and other collateral ligament tears (lateral aspect; and other lesions. This review provides a basis for understanding the ultrasonography of the finger and will therefore be useful for radiologists.

  10. Current status of ultrasonography of the finger

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Seun Ah; Kim, Baek Hyun [Dept. of Radiology, Korea University Ansan Hospital, Korea University College of Medicine, Ansan (Korea, Republic of); Kim, Seon Jeong [Dept. of Radiology, Myongji Hospital, Seonam University College of Medicine, Goyang (Korea, Republic of); Kim, Ji Na [Dept. of Radiology, Chungnam National University Hospital, Chungnam National University School of Medicine, Daejeon (Korea, Republic of); Park, Sun Young [Dept. of Radiology, Hallym University Sacred Heart Hospital, Hallym University College of Medicine, Anyang (Korea, Republic of); Choi, Kyung Hee [Incheon Baek Hospital, Incheon (Korea, Republic of)

    2016-03-15

    The recent development of advanced high-resolution transducers has enabled the fast, easy, and dynamic ultrasonographic evaluation of small, superficial structures such as the finger. In order to best exploit these advances, it is important to understand the normal anatomy and the basic pathologies of the finger, as exemplified by the following conditions involving the dorsal, volar, and lateral sections of the finger: sagittal band injuries, mallet finger, and Boutonnière deformity (dorsal aspect); flexor tendon tears, trigger finger, and volar plate injuries (volar aspect); gamekeeper’s thumb (Stener lesions) and other collateral ligament tears (lateral aspect); and other lesions. This review provides a basis for understanding the ultrasonography of the finger and will therefore be useful for radiologists.

  11. Prosthetic Hand With Two Gripping Fingers

    Science.gov (United States)

    Norton, William E.; Belcher, Jewell B.; Vest, Thomas W.; Carden, James R.

    1993-01-01

    Prosthetic hand developed for amputee who retains significant portion of forearm. Outer end of device is end effector including two fingers, one moved by rotating remaining part of forearm about its longitudinal axis. Main body of end effector is end member supporting fingers, roller bearing assembly, and rack-and-pinion mechanism. Advantage of rack-and-pinion mechanism enables user to open or close gap between fingers with precision and force.

  12. Expression of zinc finger protein X-linked in childhood B lineage acute lymphoblastic leukemia%锌指蛋白ZFX在儿童B系急性淋巴细胞性白血病中的表达及意义

    Institute of Scientific and Technical Information of China (English)

    王卓; 梁欢欢; 李本尚; 黄小航; 张婧; 王翔; 丁丽霞; 江华

    2013-01-01

    Objective To study the expression of zinc finger protein X-linked (ZFX) in bone marrow mononuclear cells (BMMCs) of children with B lineage acute lymphoblastic leukemia (B-ALL)and its relationship with prognosis.Methods The expression of ZFX in human leukemia cell lines (REH,HL-60,NB4 and K562) was measured by Western blot.ZFX gene was cloned by PCR from one patient and DNA sequencing technology was used to confirm it.Real-time PCR was used for detecting ZFX mRNA expression in the BMMCs of 82 children with newly-diagnosed B-ALL,24 children with complete remission (CR) after induction therapy and 64 control children (fracture or congenital heart disease patients).According to the presence of bone marrow or central nervous system relapse during a follow-up of 3 years,the patients were identified as having a good or poor prognosis.Their ZFX mRNA levels in BMMCs at diagnosis were compared.Results ZFX protein was expressed in human leukemia cell lines REH,HL-60,NB4 and K562.ZFX mRNA expression was significantly higher in the newly-diagnosed ALL group than in the control group (P < 0.01).ZFX mRNA expression in the ALL CR group was significantly reduced compared with the newly-diagnosed ALL group (P < 0.01).Children with a poor prognosis had significantly higher ZFX mRNA levels at diagnosis than those with a good prognosis (P< 0.05).Conclusions ZFX is over-expressed in children with B-ALL and its levels are higher in those with a poor prognosis than those with a good prognosis,which suggests that ZFX is important in the prognosis evaluation of B-ALL.%目的 研究儿童B系急性淋巴细胞性白血病(B-ALL)患者骨髓单个核细胞中X染色体耦联锌指蛋白(ZFX)的表达及其与白血病预后的关系.方法 通过Western blot方法检测白血病细胞株(REH、HL-60、NB4、K562)中ZFX蛋白的表达;利用PCR方法扩增获得全长ZFX,并测序验证;通过实时定量PCR检测82例初发B-ALL儿童骨髓样本,24例B-ALL缓解期样本,以及64

  13. Zinc coordination spheres in protein structures.

    Science.gov (United States)

    Laitaoja, Mikko; Valjakka, Jarkko; Jänis, Janne

    2013-10-07

    Zinc metalloproteins are one of the most abundant and structurally diverse proteins in nature. In these proteins, the Zn(II) ion possesses a multifunctional role as it stabilizes the fold of small zinc fingers, catalyzes essential reactions in enzymes of all six classes, or assists in the formation of biological oligomers. Previously, a number of database surveys have been conducted on zinc proteins to gain broader insights into their rich coordination chemistry. However, many of these surveys suffer from severe flaws and misinterpretations or are otherwise limited. To provide a more comprehensive, up-to-date picture on zinc coordination environments in proteins, zinc containing protein structures deposited in the Protein Data Bank (PDB) were analyzed in detail. A statistical analysis in terms of zinc coordinating amino acids, metal-to-ligand bond lengths, coordination number, and structural classification was performed, revealing coordination spheres from classical tetrahedral cysteine/histidine binding sites to more complex binuclear sites with carboxylated lysine residues. According to the results, coordination spheres of hundreds of crystal structures in the PDB could be misinterpreted due to symmetry-related molecules or missing electron densities for ligands. The analysis also revealed increasing average metal-to-ligand bond length as a function of crystallographic resolution, which should be taken into account when interrogating metal ion binding sites. Moreover, one-third of the zinc ions present in crystal structures are artifacts, merely aiding crystal formation and packing with no biological significance. Our analysis provides solid evidence that a minimal stable zinc coordination sphere is made up by four ligands and adopts a tetrahedral coordination geometry.

  14. Surgical Treatment of Trigger Finger: Open Release

    Directory of Open Access Journals (Sweden)

    Firat Ozan

    2016-01-01

    Full Text Available In this study, open A1 pulley release results were evaluated in patients with a trigger finger diagnosis. 45 patients (29 females, 16 males, mean age 50.7 ± 11.9; range (24-79, 45 trigger fingers were released via open surgical technique. On the 25 of 45 cases were involved in the right hand and 16 of them were at the thumb, 2 at index, 6 at the middle and 1 at ring finger. Similarly, at the left hand, 15 of 20 cases were at the thumb, 1 at the index finger, 2 at middle finger and 2 at ring finger. Average follow-up time was 10.2 ± 2.7 (range, 6-15 months. Comorbidities in patients were; diabetes mellitus at 6 cases (13.3%, hypertension at 11 cases (24.4%, hyperthyroidism at 2 cases (4.4%, dyslipidemia at 2 cases (4.4% and lastly 2 cases had carpal tunnel syndrome operation. The mean time between the onset of symptoms to surgery was 6.9 ± 4.8 (range, 2-24 months. Patient satisfaction was very good in 34 cases (75.4% and good in 11 (24.6% patients. The distance between the pulpa of the operated finger and the palm was normal in every case postoperatively. We have not encountered any postoperative complications. We can recommend that; A1 pulley release via open incision is an effective and reliable method in trigger finger surgery.

  15. Generating and analyzing synthetic finger vein images

    NARCIS (Netherlands)

    Hillerström, Fieke; Kumar, Ajay; Veldhuis, Raymond

    2014-01-01

    Abstract: The finger-vein biometric offers higher degree of security, personal privacy and strong anti-spoofing capabilities than most other biometric modalities employed today. Emerging privacy concerns with the database acquisition and lack of availability of large scale finger-vein database have

  16. Generic Automated Multi-function Finger Design

    Science.gov (United States)

    Honarpardaz, M.; Tarkian, M.; Sirkett, D.; Ölvander, J.; Feng, X.; Elf, J.; Sjögren, R.

    2016-11-01

    Multi-function fingers that are able to handle multiple workpieces are crucial in improvement of a robot workcell. Design automation of multi-function fingers is highly demanded by robot industries to overcome the current iterative, time consuming and complex manual design process. However, the existing approaches for the multi-function finger design automation are unable to entirely meet the robot industries’ need. This paper proposes a generic approach for design automation of multi-function fingers. The proposed approach completely automates the design process and requires no expert skill. In addition, this approach executes the design process much faster than the current manual process. To validate the approach, multi-function fingers are successfully designed for two case studies. Further, the results are discussed and benchmarked with existing approaches.

  17. Double dislocation of finger interphalangeal joints.

    Science.gov (United States)

    Jahangiri, Saqib Aziz; Mestha, Prabhakar; McNally, Scarlett

    2012-10-22

    A 62-year-old, right-hand-dominant man who had dementia and lived in an Elderly Mentally Infirm (EMI) nursing home was admitted through Accident & Emergency (A&E) department following unwitnessed injury to the left little finger. His examination revealed a swollen and deformed left little finger with a laceration along the middle crease on the volar aspect and head of proximal phalanx visible through this. Distally sensations and capillary refill was normal. X-rays showed a double dislocation of both proximal and distal interphalangeal joints. The finger was reduced under ring block and the laceration was washed with saline in A&E. The patient was taken to the operation theatre next morning for wound exploration and wash-out±stabilistion of the finger under general anaesthesia. The wound was thoroughly washed out and closed with 4/0 interrupted nylon. The finger was immobilised with neighbour strapping and bandaged in flexion.

  18. Use of twin dorsal middle phalangeal finger flaps for thumb or index finger reconstruction.

    Science.gov (United States)

    Qi, W; Chen, K J

    2013-05-01

    Amputation or degloving injuries of the thumb or index finger are highly disabling. We describe the use of twin dorsal middle finger flaps harvested from the dorsal aspects of the middle and ring fingers, and based on one palmar proper digital artery, its venae comitantes, and the dorsal branches of the palmar digital nerves of the middle and ring fingers, respectively. These flaps offer advantages when large soft tissue defects of the thumb or index finger are present. In this study, twin dorsal middle finger flaps were used in nine patients (six thumbs, three index fingers). All flaps completely survived. At the mean follow-up of 20 months, the appearance of the reconstructed thumbs or index fingers was acceptable, the length was maintained, and the mean static 2-point discrimination values were 10 mm in the palmar flap and 13 mm in the dorsal flap of the reconstructed digit. All patients were satisfied with the appearance and mobility of the donor fingers. All but one donor finger showed normal finger pulp sensibility, with a static 2-point discrimination between 3 and 6 mm.

  19. An effective preprocessing method for finger vein recognition

    Science.gov (United States)

    Peng, JiaLiang; Li, Qiong; Wang, Ning; Abd El-Latif, Ahmed A.; Niu, Xiamu

    2013-07-01

    The image preprocessing plays an important role in finger vein recognition system. However, previous preprocessing schemes remind weakness to be resolved for the high finger vein recongtion performance. In this paper, we propose a new finger vein preprocessing that includes finger region localization, alignment, finger vein ROI segmentation and enhancement. The experimental results show that the proposed scheme is capable of enhancing the quality of finger vein image effectively and reliably.

  20. Optical flow based finger stroke detection

    Science.gov (United States)

    Zhu, Zhongdi; Li, Bin; Wang, Kongqiao

    2010-07-01

    Finger stroke detection is an important topic in hand based Human Computer Interaction (HCI) system. Few research studies have carried out effective solutions to this problem. In this paper, we present a novel approach for stroke detection based on mono vision. Via analyzing the optical flow field within the finger area, our method is able to detect finger stroke under various camera position and visual angles. We present a thorough evaluation for each component of the algorithm, and show its efficiency and effectiveness on solving difficult stroke detection problems.

  1. Involvement of XZFP36L1,an RNA-binding protein,in Xenopus neural development%Involvement of XZFP36L1, an RNA-binding protein,in Xenopus neural development

    Institute of Scientific and Technical Information of China (English)

    Yingjie XIA; Shuhua ZHAO; Bingyu MAO

    2012-01-01

    Xenopus ZFP36L1 (zinc finger protein 36,C3H type-like 1) belongs to the ZFP36 family of RNA-binding proteins,which contains two characteristic tandem CCCH-type zinc-finger domains.The ZFP36 proteins can bind AU-rich elements in 3' untranslated regions of target mRNAs and promote their turnover.However,the expression and role of ZFP36 genes during neural development in Xenopus embryos remains largely tmknown.The present study showed that Xenopus ZFP36L1 was expressed at the dorsal part of the forebrain,forebrain-midbrain boundary,and midbrain-hindbrain boundary from late neurula stages to tadpole stages of embryonic development.Overexpression of XZFP36L1 in Xenopus embryos inhibited neural induction and differentiation,leading to severe neural tube defects.The function of XZP36L1 requires both its zinc finger and C terminal domains,which also affect its subcellular localization.These results suggest that XZFP36L1 is likely involved in neural development in Xenopus and might play an important role in post-transcriptional regulation.

  2. Fingering Convection in Red Giants Revisited

    CERN Document Server

    Wachlin, F C; Althaus, L G

    2014-01-01

    Fingering (thermohaline) convection has been invoked for several years as a possible extra-mixing which could occur in Red Giant stars due to the modification of the chemical composition induced by nuclear reactions in the hydrogen burning zone. Recent studies show however that this mixing is not sufficient to account for the needed surface abundances. A new prescription for fingering convection, based on 3D numerical simulations has recently been proposed (BGS). The resulting mixing coefficient is larger than the ones previously given in the literature. We compute models using this new coefficient and compare them to previous studies. We use the LPCODE stellar evolution code with the GNA generalized version of the mixing length theory to compute Red Giant models and we introduce fingering convection using the BGS prescription. The results show that, although the fingering zone now reaches the outer dynamical convective zone, the efficiency of the mixing is not enough to account for the observations. The fing...

  3. Case reports: thumb reconstruction using amputated fingers.

    Science.gov (United States)

    Hoang, Nguyen T; Staudenmaier, R; Hoehnke, C

    2008-08-01

    Reconstruction of an irreparably amputated thumb in multiple digit amputations using amputated fingers can considerably improve hand function and allows creation of a newly transplanted thumb with acceptable cosmetic and functional attributes. However, the surgery is challenging and rarely reported. We report six cases using this procedure in patients with crushed thumbs unsuitable for replantation. In four of the patients, the remnant of the index finger was replanted on the thumb stump and in another two patients, an amputated middle finger and ring finger were used. The patients had a minimum followup of 12 months (mean, 18 months; range, 12-45 months). All newly transplanted thumbs survived resulting in the patients having satisfactory postoperative hand function and appearance.

  4. Aesthetic finger prosthesis with silicone biomaterial

    Science.gov (United States)

    Raghu, K M; Gururaju, C R; Sundaresh, K J; Mallikarjuna, Rachappa

    2013-01-01

    The fabrication of finger prosthesis is as much an art as it is science. The ideally constructed prosthesis must duplicate the missing structures so precisely that patients can appear in public without fear of attracting unwanted attraction. A 65-years-old patient reported with loss of his right index finger up to the second phalanx and wanted to get it replaced. An impression of the amputated finger and donor were made. A wax pattern of the prosthesis was fabricated using the donor impression; a trial was performed and flasked. Medical grade silicone was intrinsically stained to match the skin tone, following which it was packed, processed and finished. This clinical report describes a method of attaining retention by selective scoring of the master cast of partially amputated finger to enhance the vacuum effect at par with the proportional distribution of the positive forces on the tissues exerted by the prosthesis. PMID:23975917

  5. Finger prosthesis: a boon to handicapped.

    Science.gov (United States)

    Gupta, Ridhima; Kumar, Lakshya; Rao, Jitendra; Singh, Kamleshwar

    2013-08-29

    This is a clinical case report of a 52-year-old male patient with four partially missing fingers of the left hand. The article describes the clinical and laboratory procedure of making prosthesis with modern silicone material. A wax pattern was fabricated using the right hand of the patient. A special type of wax was formulated to make the pattern so that it can be easily moulded and carved. Intrinsic and extrinsic staining was also performed to match the adjacent skin colour. The patient was given the finger prosthesis and was asked to use a half glove (sports) to mask the junction between the prosthesis and the normal tissue. It also provides additional retention to the artificial fingers. The patient felt his social acceptance improved after wearing the finger prosthesis.

  6. Layer Formation in Sedimentary Fingering Convection

    CERN Document Server

    Reali, J F; Alsinan, A; Meiburg, E

    2016-01-01

    When particles settle through a stable temperature or salinity gradient they can drive an instability known as sedimentary fingering convection. This phenomenon is thought to occur beneath sediment-rich river plumes in lakes and oceans, in the context of marine snow where decaying organic materials serve as the suspended particles, or in the atmosphere in the presence of aerosols or volcanic ash. Laboratory experiments of Houk and Green (1973) and Green (1987) have shown sedimentary fingering convection to be similar to the more commonly known thermohaline fingering convection in many ways. Here, we study the phenomenon using 3D direct numerical simulations. We find evidence for layer formation in sedimentary fingering convection in regions of parameter space where it does not occur for non-sedimentary systems. This is due to two complementary effects. Sedimentation affects the turbulent fluxes and broadens the region of parameter space unstable to the $\\gamma$-instability (Radko 2003) to include systems at l...

  7. Case Reports: Thumb Reconstruction Using Amputated Fingers

    OpenAIRE

    Hoang, Nguyen T.; Staudenmaier, R.; Hoehnke, C.

    2008-01-01

    Reconstruction of an irreparably amputated thumb in multiple digit amputations using amputated fingers can considerably improve hand function and allows creation of a newly transplanted thumb with acceptable cosmetic and functional attributes. However, the surgery is challenging and rarely reported. We report six cases using this procedure in patients with crushed thumbs unsuitable for replantation. In four of the patients, the remnant of the index finger was replanted on the thumb stump and ...

  8. Finger Search in the Implicit Model

    DEFF Research Database (Denmark)

    Brodal, Gerth Stølting; Nielsen, Jesper Asbjørn Sindahl; Truelsen, Jakob

    2012-01-01

    the finger to another element is Ω(q− 1(logn)), where t is the rank distance between the query element and the finger. We present an optimal implicit static structure matching this lower bound. We furthermore present a near optimal implicit dynamic structure supporting search, change-finger, insert......We address the problem of creating a dictionary with the finger search property in the strict implicit model, where no information is stored between operations, except the array of elements. We show that for any implicit dictionary supporting finger searches in q(t) = Ω(logt) time, the time to move......, and delete in times $\\mathcal{O}(q(t))$, $\\mathcal{O}(q^{-1}(\\log n)\\log n)$, $\\mathcal{O}(\\log n)$, and $\\mathcal{O}(\\log n)$, respectively, for any q(t) = Ω(logt). Finally we show that the search operation must take Ω(logn) time for the special case where the finger is always changed to the element...

  9. Scattering removal for finger-vein image restoration.

    Science.gov (United States)

    Yang, Jinfeng; Zhang, Ben; Shi, Yihua

    2012-01-01

    Finger-vein recognition has received increased attention recently. However, the finger-vein images are always captured in poor quality. This certainly makes finger-vein feature representation unreliable, and further impairs the accuracy of finger-vein recognition. In this paper, we first give an analysis of the intrinsic factors causing finger-vein image degradation, and then propose a simple but effective image restoration method based on scattering removal. To give a proper description of finger-vein image degradation, a biological optical model (BOM) specific to finger-vein imaging is proposed according to the principle of light propagation in biological tissues. Based on BOM, the light scattering component is sensibly estimated and properly removed for finger-vein image restoration. Finally, experimental results demonstrate that the proposed method is powerful in enhancing the finger-vein image contrast and in improving the finger-vein image matching accuracy.

  10. Highly transparent front electrodes with metal fingers for p-i-n thin-film silicon solar cells

    Directory of Open Access Journals (Sweden)

    Moulin Etienne

    2015-01-01

    Full Text Available The optical and electrical properties of transparent conductive oxides (TCOs, traditionally used in thin-film silicon (TF-Si solar cells as front-electrode materials, are interlinked, such that an increase in TCO transparency is generally achieved at the cost of reduced lateral conductance. Combining a highly transparent TCO front electrode of moderate conductance with metal fingers to support charge collection is a well-established technique in wafer-based technologies or for TF-Si solar cells in the substrate (n-i-p configuration. Here, we extend this concept to TF-Si solar cells in the superstrate (p-i-n configuration. The metal fingers are used in conjunction with a millimeter-scale textured foil, attached to the glass superstrate, which provides an antireflective and retroreflective effect; the latter effect mitigates the shadowing losses induced by the metal fingers. As a result, a substantial increase in power conversion efficiency, from 8.7% to 9.1%, is achieved for 1-μm-thick microcrystalline silicon solar cells deposited on a highly transparent thermally treated aluminum-doped zinc oxide layer combined with silver fingers, compared to cells deposited on a state-of-the-art zinc oxide layer.

  11. Scattering Removal for Finger-Vein Image Restoration

    OpenAIRE

    Jinfeng Yang; Ben Zhang; Yihua Shi

    2012-01-01

    Finger-vein recognition has received increased attention recently. However, the finger-vein images are always captured in poor quality. This certainly makes finger-vein feature representation unreliable, and further impairs the accuracy of finger-vein recognition. In this paper, we first give an analysis of the intrinsic factors causing finger-vein image degradation, and then propose a simple but effective image restoration method based on scattering removal. To give a proper description of f...

  12. Zinc: the neglected nutrient.

    Science.gov (United States)

    Shambaugh, G E

    1989-03-01

    Zinc was first recognized as essential for animals at the University of Illinois School of Agriculture in 1916, when it was found that zinc-deficient baby pigs were runty, developed dermatitis on their legs, and were sterile. Zinc deficiency was first recognized in man by Dr. Ananda Prasad of Detroit 26 years ago when he measured serum and hair zinc levels in young male Egyptian dwarfs who had failed to mature and were small in stature. By simply adding zinc to their regular diet, they grew in height and became sexually mature. It is now recognized that dwarfism in males is frequent around the Mediterranean, where wheat is the staple of life and has been grown for 4,000 years on the same soil, thereby resulting in the depletion of zinc. Professor Robert Henkin first suggested that zinc deficiency might cause hearing-nerve impairment. Assay of the soft tissues of the cochlea and vestibule revealed a zinc level higher than that of any other part of the body. Previously, the eye was considered to have the highest level of zinc of any organ. To diagnose zinc deficiency clinically, we use serum zinc assays made at the Mayo Clinic Trace Element Laboratory. With zinc supplementation in patients who are marginally zinc deficient, there has been improvement in tinnitus and sensorineural hearing loss in about one-third of elderly adults. We believe zinc deficiency is one causation of presbycusis; by recognizing and correcting it, a progressive hearing loss can be arrested.

  13. Design of a 2-Finger Hand Exoskeleton for Finger Stiffness Measurements

    Directory of Open Access Journals (Sweden)

    Angelo Emanuele Fiorilla

    2009-01-01

    Full Text Available Recent studies of human arm movements have suggested that the control of stiffness may be important both for maintaining stability and for achieving differences in movement accuracy. Several studies in the robotic field demonstrated that grasp stiffness is useful for modelling and controlling manipulators but, even though it is accredited that having models of the human finger impedance would be very desirable for the control of anthropomorphous robot's hands, relatively few studies have focused on finger and hand stiffness. To allow the measurement of such entities at the finger level, an appropriate device capable of applying fast force transients while at the same time be able to monitor the finger movements is required. The work presented in this paper is a very detailed report about the design of a new hand exoskeleton system that will be used in our future works to investigate the finger stiffness range in different grasping postures and conditions.

  14. Trace elements in human physiology and pathology: zinc and metallothioneins.

    Science.gov (United States)

    Tapiero, Haim; Tew, Kenneth D

    2003-11-01

    Zinc is one of the most abundant nutritionally essential elements in the human body. It is found in all body tissues with 85% of the whole body zinc in muscle and bone, 11% in the skin and the liver and the remaining in all the other tissues. In multicellular organisms, virtually all zinc is intracellular, 30-40% is located in the nucleus, 50% in the cytoplasm, organelles and specialized vesicles (for digestive enzymes or hormone storage) and the remainder in the cell membrane. Zinc intake ranges from 107 to 231 micromol/d depending on the source, and human zinc requirement is estimated at 15 mg/d. Zinc has been shown to be essential to the structure and function of a large number of macromolecules and for over 300 enzymic reactions. It has both catalytic and structural roles in enzymes, while in zinc finger motifs, it provides a scaffold that organizes protein sub-domains for the interaction with either DNA or other proteins. It is critical for the function of a number of metalloproteins, inducing members of oxido-reductase, hydrolase ligase, lyase family and has co-activating functions with copper in superoxide dismutase or phospholipase C. The zinc ion (Zn(++)) does not participate in redox reactions, which makes it a stable ion in a biological medium whose potential is in constant flux. Zinc ions are hydrophilic and do not cross cell membranes by passive diffusion. In general, transport has been described as having both saturable and non-saturable components, depending on the Zn(II) concentrations involved. Zinc ions exist primarily in the form of complexes with proteins and nucleic acids and participate in all aspects of intermediary metabolism, transmission and regulation of the expression of genetic information, storage, synthesis and action of peptide hormones and structural maintenance of chromatin and biomembranes.

  15. Is zinc a neuromodulator?

    Science.gov (United States)

    Kay, Alan R; Tóth, Katalin

    2008-01-01

    The vesicles of certain glutamatergic terminals in the mammalian forebrain are replete with ionic zinc. It is believed that during synaptic transmission zinc is released, binds to receptors on the pre- or postsynaptic membranes, and hence acts as a neuromodulator. Although exogenous zinc modulates a wide variety of channels, whether synaptic zinc transits across the synaptic cleft and alters the response of channels has been difficult to establish. We will review the evidence for zinc as a neuromodulator and propose diagnostic criteria for establishing whether it is indeed one. Moreover, we will delineate alternative ways in which zinc might act at synapses.

  16. Saffman-Taylor fingers with kinetic undercooling

    KAUST Repository

    Gardiner, Bennett P. J.

    2015-02-23

    © 2015 American Physical Society. The mathematical model of a steadily propagating Saffman-Taylor finger in a Hele-Shaw channel has applications to two-dimensional interacting streamer discharges which are aligned in a periodic array. In the streamer context, the relevant regularization on the interface is not provided by surface tension but instead has been postulated to involve a mechanism equivalent to kinetic undercooling, which acts to penalize high velocities and prevent blow-up of the unregularized solution. Previous asymptotic results for the Hele-Shaw finger problem with kinetic undercooling suggest that for a given value of the kinetic undercooling parameter, there is a discrete set of possible finger shapes, each analytic at the nose and occupying a different fraction of the channel width. In the limit in which the kinetic undercooling parameter vanishes, the fraction for each family approaches 1/2, suggesting that this "selection" of 1/2 by kinetic undercooling is qualitatively similar to the well-known analog with surface tension. We treat the numerical problem of computing these Saffman-Taylor fingers with kinetic undercooling, which turns out to be more subtle than the analog with surface tension, since kinetic undercooling permits finger shapes which are corner-free but not analytic. We provide numerical evidence for the selection mechanism by setting up a problem with both kinetic undercooling and surface tension and numerically taking the limit that the surface tension vanishes.

  17. Crustal fingering: solidification on a moving interface

    Science.gov (United States)

    Fu, Xiaojing; Jimenez-Martinez, Joaquin; Porter, Mark; Cueto-Felgueroso, Luis; Juanes, Ruben

    2016-11-01

    Viscous fingering-the hydrodynamic instability that takes place when a less viscous fluid displaces a more viscous fluid-is a well known phenomenon. Motivated by the formation of gas hydrates in seafloor sediments and during the ascent of gas bubbles through ocean water, here we study the interplay of immiscible viscous fingering with solidification of the evolving unstable interface. We present experimental observations of the dynamics of a bubble of Xenon in a water-filled and pressurized Hele-Shaw cell. The evolution is controlled by two processes: (1) the formation of a hydrate "crust" around the bubble, and (2) viscous fingering from bubble expansion. To reproduce the experimental observations, we propose a phase-field model that describes the nucleation and thickening of a porous solid shell on a moving gas-liquid interface. We design the free energy of the three-phase system (gas-liquid-hydrate) to rigorously account for interfacial effects, mutual solubility, and phase transformations (hydrate formation and disappearance). We introduce a pseudo-plasticity model with large variations in viscosity to describe the plate-like rheology of the hydrate shell. We present high-resolution numerical simulations of the model, which illustrate the emergence of complex "crustal fingering" patterns as a result of gas fingering dynamics modulated by hydrate growth at the interface.

  18. Perceiving fingers in single-digit arithmetic problems

    Directory of Open Access Journals (Sweden)

    Ilaria eBerteletti

    2015-03-01

    Full Text Available In this study, we investigate in children the neural underpinnings of finger representation and finger movement involved in single-digit arithmetic problems. Evidence suggests that finger representation and finger-based strategies play an important role in learning and understanding arithmetic. Because different operations rely on different networks, we compared activation for subtraction and multiplication problems in independently localized finger somatosensory and motor areas and tested whether activation was related to skill. Brain activations from children between 8 and 13 years of age revealed that only subtraction problems significantly activated finger motor areas, suggesting reliance on finger-based strategies. In addition, larger subtraction problems yielded greater somatosensory activation than smaller problems, suggesting a greater reliance on finger representation for larger numerical values. Interestingly, better performance in subtraction problems was associated with lower activation in the finger somatosensory area. Our results support the importance of fine-grained finger representation in arithmetical skill and are the first neurological evidence for a functional role of the somatosensory finger area in proficient arithmetical problem solving, in particular for those problems requiring quantity manipulation. From an educational perspective, these results encourage investigating whether different finger-based strategies facilitate arithmetical understanding and encourage educational practices aiming at integrating finger representation and finger-based strategies as a tool for instilling stronger numerical sense.

  19. Finger recognition and gesture imitation in Gerstmann's syndrome.

    Science.gov (United States)

    Moro, V; Pernigo, S; Urgesi, C; Zapparoli, P; Aglioti, S M

    2008-01-01

    We report the association between finger agnosia and gesture imitation deficits in a right-handed, right-hemisphere damaged patient with Gerstmann's syndrome (GS), a neuropsychological syndrome characterized by finger and toe agnosia, left-right disorientation and dyscalculia. No language deficits were found. The patient showed a gestural imitation deficit that specifically involved finger movements and postures. The association between finger recognition and imitation deficits suggests that both static and dynamic aspects of finger representations are impaired in GS. We suggest that GS is a disorder of body representation that involves hands and fingers, that is, the non-facial body parts most involved in social interactions.

  20. Zinc oxide overdose

    Science.gov (United States)

    Zinc oxide is an ingredient in many products. Some of these are certain creams and ointments used ... prevent or treat minor skin burns and irritation. Zinc oxide overdose occurs when someone eats one of ...

  1. Reactivity of Cys4 Zinc Finger Domains with Gold(III) Complexes : Insights into the Formation of "Gold Fingers"

    NARCIS (Netherlands)

    Jacques, Aurélie; Lebrun, Colette; Casini, Angela; Kieffer, Isabelle; Proux, Olivier; Latour, Jean-Marc; Sénèque, Olivier

    2015-01-01

    Gold(I) complexes such as auranofin or aurothiomalate have been used as therapeutic agents for the treatment of rheumatoid arthritis for several decades. Several gold(I) and gold(III) complexes have also shown in vitro anticancer properties against human cancer cell lines, including cell lines resis

  2. Autonomous control of multi-fingered hand

    Institute of Scientific and Technical Information of China (English)

    JIANG Li; LIU Hong

    2006-01-01

    This paper describes a novel autonomous control strategy of multi-fingered hand based on a modular control system of dexterous manipulation. A simple proportional-integral-derivative(PID) position control with friction compensation, which requires few friction parameters, is used to realize accurate and smooth trajectory tracking in pregrasp phase. In grasp and manipulation phases, an event-driven switcher is adopted to determine the switching between unconstrained position control and constrained torque control, and an improved explicit integral force control strategy is implemented to realize simultaneously stable contact transition and accurate force tracking. Experimental results have verified the effectiveness of the proposed autonomous control strategy of multi-fingered hand.

  3. Fingering instability in combustion: an extended view.

    Science.gov (United States)

    Zik, O; Moses, E

    1999-07-01

    We detail the experimental situation concerning the fingering instability that occurs when a solid fuel is forced to burn against a horizontal oxidizing wind. The instability appears when the Rayleigh number for convection is below criticality. The focus is on the developed fingering state. We present direct measurements of the depletion of oxygen by the front as well as new results that connect heat losses to the characteristic scale of the instability. In addition, we detail the experimental system, elaborate (qualitatively and quantitatively) on the results that were previously presented, and discuss new observations. We also show that the same phenomenological model applies to electrochemical deposition.

  4. Admittance Control of a Multi-Finger Arm Based on Manipulability of Fingers

    Directory of Open Access Journals (Sweden)

    Takayuki Hori

    2011-09-01

    Full Text Available In the previous studies, admittance control and impedance control for a finger‐arm robot using the manipulability of the finger were studied and methods of realizing the controls have been proposed. In this study, two 3‐DOF fingers are attached to the end‐effector of a 6‐DOF arm to configure a multi‐finger arm robot. Based on the previous methods, the authors have proposed an admittance control for a multi‐finger arm robot using the manipulability of the fingers in this study. Algorithms of the averaging method and the mini‐max method were introduced to establish a manipulability criterion of the two fingers in order to generate a cooperative movement of the arm. Comparison of the admittance controls combined with the top search method and local optimization method for the multi‐finger arm robot was made and features of the control methods were also discussed. The stiffness control and damping control were experimentally evaluated to demonstrate the effectiveness of the proposed methods.

  5. ANALYSIS WITH MSC ADAMS OF A 5-FINGER AND 3-PHALANX /FINGER UNDER-ACTUATEDMECHANICAL HAND

    Directory of Open Access Journals (Sweden)

    Gheorghe POPESCU

    2013-05-01

    Full Text Available This paper studies the analysis with MSC ADAMS of a 5-fingered and 3-phalanx/finger underactuatedmechanical hand, designed by the author to work on industrial robots. Moreover, in order to increasegrasping safety in the automated handling process, the author has fitted each finger with a locking sequence inthe final phase of grasping. Thus, the mechanism of mechanical hand is considered to be a mechanical systemand is treated like a set of rigid bodies connected by mechanical linkages and elastic elements. To model andsimulate this mechanism with MSC ADAMS programme, the author covered the following stages: constructionof the model, testing-simulation, validation, finishing, parameterization, and optimization

  6. The PHD finger of human UHRF1 reveals a new subgroup of unmethylated histone H3 tail readers.

    Directory of Open Access Journals (Sweden)

    Nada Lallous

    Full Text Available The human UHRF1 protein (ubiquitin-like containing PHD and RING finger domains 1 has emerged as a potential cancer target due to its implication in cell cycle regulation, maintenance of DNA methylation after replication and heterochromatin formation. UHRF1 functions as an adaptor protein that binds to histones and recruits histone modifying enzymes, like HDAC1 or G9a, which exert their action on chromatin. In this work, we show the binding specificity of the PHD finger of human UHRF1 (huUHRF1-PHD towards unmodified histone H3 N-terminal tail using native gel electrophoresis and isothermal titration calorimetry. We report the molecular basis of this interaction by determining the crystal structure of huUHRF1-PHD in complex with the histone H3 N-terminal tail. The structure reveals a new mode of histone recognition involving an extra conserved zinc finger preceding the conventional PHD finger region. This additional zinc finger forms part of a large surface cavity that accommodates the side chain of the histone H3 lysine K4 (H3K4 regardless of its methylation state. Mutation of Q330, which specifically interacts with H3K4, to alanine has no effect on the binding, suggesting a loose interaction between huUHRF1-PHD and H3K4. On the other hand, the recognition appears to rely on histone H3R2, which fits snugly into a groove on the protein and makes tight interactions with the conserved aspartates D334 and D337. Indeed, a mutation of the former aspartate disrupts the formation of the complex, while mutating the latter decreases the binding affinity nine-fold.

  7. Acceleration Workspace of Cooperating Multi-Finger Robot Systems

    Institute of Scientific and Technical Information of China (English)

    Hyungwon Shim; Jihong Lee

    2008-01-01

    We present a mathematical method for acceleration workspace analysis of cooperating multi-finger robot systems using a model of point-contact with friction. A new unified formulation from dynamic equations of cooperating multi-finger robots is derived considering the force and acceleration relationships between the fingers and the object to be handled. From the dynamic equation, maximum translational and rotational acceleration bounds of an object are calculated under given constraints of contact conditions, configurations of fingers, and bounds on the torques of joint actuators for each finger. Here, the rotational acceleration bounds can be applied as an important manipulability index when the multi-finger robot grasps an object. To verify the proposed method, we used a set of case studies with a simple multi-finger mechanism system. The achievable acceleration boundary in task space can be obtained successfully with the proposed method and the acceleration boundary depends on the configurations of fingers.

  8. Effective Length Design of Humanoid Robot Fingers Using Biomimetic Optimization

    Directory of Open Access Journals (Sweden)

    Byoung-Ho Kim

    2015-10-01

    Full Text Available In this study, we propose an effective design method for the phalangeal parameters and the total size of humanoid robot fingers based on a biomimetic optimization. For the optimization, an interphalangeal joint coordination parameter and the length constraints inherent in human fingers are considered from a biomimetic perspective. A reasonable grasp formulation is also taken into account from the viewpoint of power grasping, where the grasp space of a humanoid robot finger is importantly considered to determine the phalangeal length parameters. The usefulness of the devised biomimetic optimization method is shown through the design examples of various humanoid robot fingers. In fact, the optimization-based finger design method enables us to determine effectively the proper phalangeal size of humanoid robot fingers for human-like object handling tasks. In addition, we discuss its contribution to the structural configuration and coordinated motion of a humanoid robot finger, and address its practical availability in terms of effective finger design.

  9. Finger Search in Grammar-Compressed Strings

    DEFF Research Database (Denmark)

    Bille, Philip; Christiansen, Anders Roy; Cording, Patrick Hagge;

    2016-01-01

    random access, that is, given a position in the original uncompressed string report the character at that position. In this paper we study the random access problem with the finger search property, that is, the time for a random access query should depend on the distance between a specified index f...

  10. Viscous fingering with partial miscible fluids

    Science.gov (United States)

    Fu, Xiaojing; Cueto-Felgueroso, Luis; Juanes, Ruben

    2015-11-01

    When a less viscous fluid displaces a more viscous fluid, the contrast in viscosity destabilizes the interface between the two fluids, leading to the formation of fingers. Studies of viscous fingering have focused on fluids that are either fully miscible or perfectly immiscible. In practice, however, the miscibility of two fluids can change appreciably with temperature and pressure, and often falls into the case of partial miscibility, where two fluids have limited solubility in each other. Following our recent work for miscible (Jha et al., PRL 2011, 2013) and immiscible systems (Cueto-Felgueroso and Juanes, PRL 2012, JFM 2014), here we propose a phase-field model for fluid-fluid displacements in a Hele-Shaw cell, when the two fluids have limited (but nonzero) solubility in one another. Partial miscibility is characterized through the design of thermodynamic free energy of the two-fluid system. We elucidate the key dimensionless groups that control the behavior of the system. We present high-resolution numerical simulations of the model applied to the viscous fingering problem. On one hand, we demonstrate the effect of partial miscibility on the hydrodynamic instability. On the other, we elucidate the role of the degree of fingering on the rate of mutual fluid dissolution.

  11. Clubbed fingers: the claws we lost?

    NARCIS (Netherlands)

    Brouwers, A.A.M.; Vermeij-Keers, C.; Zoelen, E.J.J. van; Gooren, L.J.G.

    2004-01-01

    Clubbed digits resemble the human embryonic fingers and toes, which took like the digits of a claw. Clubbed digits, thus, may represent the return of the embryonic claw and may even represent the claws man has lost during evolution, if ontogenesis realty recapitulates phylogenesis. We put forward th

  12. Task specificity of finger dexterity tests

    NARCIS (Netherlands)

    Berger, M.A.M.; Krul, A.; Daanen, H.A.M.

    2009-01-01

    Finger dexterity tests are generally used to assess performance decrease due to gloves, cold and pathology. It is generally assumed that the O’Connor and Purdue Pegboard test yield similar results. In this experiment we compared these two tests for dry conditions without gloves, and for dry and wet

  13. Finger-jointed beams in bending

    DEFF Research Database (Denmark)

    Andreasen, Lotte; Hoffmeyer, Preben

    1997-01-01

    An investigation of the dynamic and static fatique of finger-jointed beams in bending was carried out. Results were obtained for five different frequencies from static loading to a load cycle period of two minutes. A total of seven series were long-term tested and five series were short-term tested...

  14. Coriolis effects on fingering patterns under rotation.

    Science.gov (United States)

    Alvarez-Lacalle, Enrique; Gadêlha, Hermes; Miranda, José A

    2008-08-01

    The development of immiscible viscous fingering patterns in a rotating Hele-Shaw cell is investigated. We focus on understanding how the time evolution and the resulting morphologies are affected by the action of the Coriolis force. The problem is approached analytically and numerically by employing a vortex sheet formalism. The vortex sheet strength and a linear dispersion relation are derived analytically, revealing that the most relevant Coriolis force contribution comes from the normal component of the averaged interfacial velocity. It is shown that this normal velocity, uniquely due to the presence of the Coriolis force, is responsible for the complex-valued nature of the linear dispersion relation making the linear phases vary with time. Fully nonlinear stages are studied through intensive numerical simulations. A suggestive interplay between inertial and viscous effects is found, which modifies the dynamics, leading to different pattern-forming structures. The inertial Coriolis contribution plays a characteristic role: it generates a phase drift by deviating the fingers in the sense opposite to the actual rotation of the cell. However, the direction and intensity of finger bending is predominantly determined by viscous effects, being sensitive to changes in the magnitude and sign of the viscosity contrast. The finger competition behavior at advanced time stages is also discussed.

  15. Fingering phenomena during grain-grain displacement

    Science.gov (United States)

    Mello, Nathália M. P.; Paiva, Humberto A.; Combe, G.; Atman, A. P. F.

    2016-05-01

    Spontaneous formation of fingered patterns during the displacement of dense granular assemblies was experimentally reported few years ago, in a radial Hele-Shaw cell. Here, by means of discrete element simulations, we have recovered the experimental findings and extended the original study to explore the control parameters space. In particular, using assemblies of grains with different geometries (monodisperse, bidisperse, or polydisperse), we measured the macroscopic stress tensor in the samples in order to confirm some conjectures proposed in analogy with Saffman-Taylor viscous fingering phenomena for immiscible fluids. Considering an axial setup which allows to control the discharge of grains and to follow the trajectory and the pressure gradient along the displacing interface, we have applied the Darcy law for laminar flow in fluids in order to measure an "effective viscosity" for each assembly combination, in an attempt to mimic variation of the viscosity ratio between the injected/displaced fluids in the Saffman-Taylor experiment. The results corroborate the analogy with the viscous fluids displacement, with the bidisperse assembly corresponding to the less viscous geometry. But, differently to fluid case, granular fingers only develop for a specific combination of displaced/injected geometries, and we have demonstrated that it is always related with the formation of a force chain network along the finger direction.

  16. Cadmium and zinc relationships

    Energy Technology Data Exchange (ETDEWEB)

    Elinder, C.; Piscator, M.

    1978-08-01

    Higher mammals, such as homo sapiens, accumulate zinc in kidney cortex almost equimolarly with cadmium. A different pattern seems to be present in liverthere is a limited increase of zinc in two species of large farm animals compared with a marked increase in the laboratory. In large farm animals, an equimolar increase of zinc with cadmium in renal cortex seems to indicate that the form of metallothionein that binds equal amounts of cadmium and zinc in present. Differences in cadmium and zinc relationships in large animals and humans compared with laboratory animals must be carefully considered. (4 graphs, 26 references)

  17. A palmitoylated RING finger ubiquitin ligase and its homologue in the brain membranes.

    Science.gov (United States)

    Araki, Kazuaki; Kawamura, Meiko; Suzuki, Toshiaki; Matsuda, Noriyuki; Kanbe, Daiji; Ishii, Kyoko; Ichikawa, Tomio; Kumanishi, Toshiro; Chiba, Tomoki; Tanaka, Keiji; Nawa, Hiroyuki

    2003-08-01

    Ubiquitin (Ub) ligation is implicated in active protein metabolism and subcellular trafficking and its impairment is involved in various neurologic diseases. In rat brain, we identified two novel Ub ligases, Momo and Sakura, carrying double zinc finger motif and RING finger domain. Momo expression is enriched in the brain gray matter and testis, and Sakura expression is more widely detected in the brain white matter as well as in many peripheral organs. Both proteins associate with the cell membranes of neuronal and/or glial cells. We examined their Ub ligase activity in vivo and in vitro using viral expression vectors carrying myc-tagged Momo and Sakura. Overexpression of either Momo or Sakura in mixed cortical cultures increased total polyubiquitination levels. In vitro ubiquitination assay revealed that the combination of Momo and UbcH4 and H5c, or of Sakura and UbcH4, H5c and H6 is required for the reaction. Deletion mutagenesis suggested that the E3 Ub ligase activity of Momo and Sakura depended on their C-terminal domains containing RING finger structure, while their N-terminal domains influenced their membrane association. In agreement, Sakura associating with the membrane was specifically palmitoylated. Although the molecular targets of their Ub ligation remain to be identified, these findings imply a novel function of the palmitoylated E3 Ub ligase(s).

  18. Cadmium and zinc relationships.

    Science.gov (United States)

    Elinder, C G; Piscator, M

    1978-08-01

    Cadmium and zinc concentrations in kidney and liver have been measured under different exposure situations in different species including man. The results show that zinc increases almost equimolarly with cadmium in kidney after long-term low-level exposure to cadmium, e.g., in man, horse, pig, and lamb. In contrast, the increase of zinc follows that of cadmium to only a limited extent, e.g., in guinea pig, rabbit, rat, mouse, and chicks. In liver, the cadmium--zinc relationship seems to be reversed in such a way that zinc increases with cadmium more markedly in laboratory animals than in higher mammals. These differences between cadmium and zinc relationships in humans and large farm animals and those in commonly used laboratory animals must be considered carefully before experimental data on cadmium and zinc relationships in laboratory animals can be extrapolated to humans.

  19. Relative finger position influences whether you can localize tactile stimuli

    NARCIS (Netherlands)

    Overvliet, K.E.; Anema, H.A.; Brenner, E.; Dijkerman, H.C.; Smeets, J.B.J.

    2011-01-01

    To investigate whether the relative positions of the fingers influence tactile localization, participants were asked to localize tactile stimuli applied to their fingertips. We measured the location and rate of errors for three finger configurations: fingers stretched out and together so that they a

  20. Robot-assisted Guitar Hero for finger rehabilitation after stroke.

    Science.gov (United States)

    Taheri, Hossein; Rowe, Justin B; Gardner, David; Chan, Vicky; Reinkensmeyer, David J; Wolbrecht, Eric T

    2012-01-01

    This paper describes the design and testing of a robotic device for finger therapy after stroke: FINGER (Finger Individuating Grasp Exercise Robot). FINGER makes use of stacked single degree-of-freedom mechanisms to assist subjects in moving individual fingers in a naturalistic grasping pattern through much of their full range of motion. The device has a high bandwidth of control (-3dB at approximately 8 Hz) and is backdriveable. These characteristics make it capable of assisting in grasping tasks that require precise timing. We therefore used FINGER to assist individuals with a stroke (n= 8) and without impairment (n= 4) in playing a game similar to Guitar Hero©. The subjects attempted to move their fingers to target positions at times specified by notes that were graphically streamed to popular music. We show here that by automatically adjusting the robot gains, it is possible to use FINGER to modulate the subject's success rate at the game, across a range of impairment levels. Modulating success rates did not alter the stroke subject's effort, although the unimpaired subjects exerted more force when they were made less successful. We also present a novel measure of finger individuation that can be assessed as individuals play Guitar Hero with FINGER. The results demonstrate the ability of FINGER to provide controlled levels of assistance during an engaging computer game, and to quantify finger individuation after stroke.

  1. Left hand finger force in violin playing: tempo, loudness, and finger differences.

    Science.gov (United States)

    Kinoshita, Hiroshi; Obata, Satoshi

    2009-07-01

    A three-dimensional force transducer was installed in the neck of a violin under the A string at the D5 position in order to study the force with which the violinist clamps the string against the fingerboard under normal playing conditions. Violinists performed repetitive sequences of open A- and fingered D-tones using the ring finger at tempi of 1, 2, 4, 8, and 16 notes/s at mezzo-forte. At selected tempi, the effects of dynamic level and the use of different fingers were investigated as well. The force profiles were clearly dependent on tempo and dynamic level. At slow tempi, the force profiles were characterized by an initial pulse followed by a level force to the end of the finger contact period. At tempi higher than 2 Hz, only pulsed profiles were observed. The peak force exceeded 4.5 N at 1 and 2 Hz and decreased to 1.7 N at 16 Hz. All force and impulse values were lower at softer dynamic levels, and when using the ring or little finger compared to the index finger.

  2. [Zinc and type 2 diabetes].

    Science.gov (United States)

    Fukunaka, Ayako; Fujitani, Yoshio

    2016-07-01

    Pancreatic β cells contain the highest amount of zinc among cells within the human body, and hence, the relationship between zinc and diabetes has been a topic of great interest. While many studies demonstrating possible involvement of zinc deficiency in diabetes have been reported, precise mechanisms how zinc regulates glucose metabolism are still far from understood. Recent studies revealed that zinc can transmit signals that are driven by a variety of zinc transporters in a tissue and cell-type specific manner and deficiency in some zinc transporters may cause human diseases. Here, we review the role of zinc in metabolism particularly focusing on the emerging role of zinc transporters in diabetes.

  3. Dynamic analysis of C/C composite finger seal

    Institute of Scientific and Technical Information of China (English)

    Chen Guoding; Wang Li’na; Yu Qiangpeng; Su Hua

    2014-01-01

    A seal device as an important component of aeroengines has decisive influence on per-formance, reliability, and working life of aeroengines. With the development of aeroengines, demands on the performance characteristics of seal devices are made strictly. Finger seal as a novel kind of sealing device, recently attracts more and more attentions in academic circles and engineer-ing fields at home and abroad. Research on finger seals has been extensively developed, especially on leakage and wear performances under dynamic conditions. However, it is a pity that the work on finger seals has been limited with a single approach that is improving the performance by structural optimization;in addition, the technology of dynamic analysis on finger seals is weak. Aiming at the problems mentioned above, a distributed mass equivalent dynamic model of finger seals considering the coupling effect of overlaid laminates is established in the present paper, the dynamic perfor-mance of 2.5 dimension C/C composite finger seal is analyzed with the model, and then the effects of fiber bundle density and fiber bundle preparation direction on finger seal’s dynamic performance are discussed, as well as compared with those of Co-based alloy finger seal. The current work is about dynamic analysis of finger seals and application of C/C composite in this paper may have much academic significance and many engineering values for improving research level of finger seal dynamics and exploring feasibility of C/C composite being used for finger seals.

  4. Dynamic analysis of C/C composite finger seal

    Directory of Open Access Journals (Sweden)

    Chen Guoding

    2014-06-01

    Full Text Available A seal device as an important component of aeroengines has decisive influence on performance, reliability, and working life of aeroengines. With the development of aeroengines, demands on the performance characteristics of seal devices are made strictly. Finger seal as a novel kind of sealing device, recently attracts more and more attentions in academic circles and engineering fields at home and abroad. Research on finger seals has been extensively developed, especially on leakage and wear performances under dynamic conditions. However, it is a pity that the work on finger seals has been limited with a single approach that is improving the performance by structural optimization; in addition, the technology of dynamic analysis on finger seals is weak. Aiming at the problems mentioned above, a distributed mass equivalent dynamic model of finger seals considering the coupling effect of overlaid laminates is established in the present paper, the dynamic performance of 2.5 dimension C/C composite finger seal is analyzed with the model, and then the effects of fiber bundle density and fiber bundle preparation direction on finger seal’s dynamic performance are discussed, as well as compared with those of Co-based alloy finger seal. The current work is about dynamic analysis of finger seals and application of C/C composite in this paper may have much academic significance and many engineering values for improving research level of finger seal dynamics and exploring feasibility of C/C composite being used for finger seals.

  5. A Parametric Modelling Method for Dexterous Finger Reachable Workspaces

    Directory of Open Access Journals (Sweden)

    Wenzhen Yang

    2016-03-01

    Full Text Available The well-known algorithms, such as the graphic method, analytical method or numerical method, have some defects when modelling the dexterous finger workspace, which is a significant kinematical feature of dexterous hands and valuable for grasp planning, motion control and mechanical design. A novel modelling method with convenient and parametric performances is introduced to generate the dexterous-finger reachable workspace. This method constructs the geometric topology of the dexterous-finger reachable workspace, and uses a joint feature recognition algorithm to extract the kinematical parameters of the dexterous finger. Compared with graphic, analytical and numerical methods, this parametric modelling method can automatically and conveniently construct a more vivid workspace’ forms and contours of the dexterous finger. The main contribution of this paper is that a workspace-modelling tool with high interactive efficiency is developed for designers to precisely visualize the dexterous-finger reachable workspace, which is valuable for analysing the flexibility of the dexterous finger.

  6. Improved zinc electrode and rechargeable zinc-air battery

    Science.gov (United States)

    Ross, P.N. Jr.

    1988-06-21

    The invention comprises an improved rechargeable zinc-air cell/battery having recirculating alkaline electrolyte and a zinc electrode comprising a porous foam support material which carries the active zinc electrode material. 5 figs.

  7. Angiolipoma of index finger: A case report

    Directory of Open Access Journals (Sweden)

    Muzaffer Durmus

    2016-04-01

    Full Text Available Angiolipomas are usually found in the upper extremities, shoulder and back. They are seldom found in the hands, face and lower extremities. They usually occur as painful soft tissue masses or they may compress the neighboring structures (e.g. nerves depending on the size and location. In this report we present an angiolipoma case located in the finger and discuss related recent cases described in the literature. [Hand Microsurg 2016; 5(1.000: 22-25

  8. Palm to finger ulnar sensory nerve conduction

    Directory of Open Access Journals (Sweden)

    Eduardo Davidowich

    2015-12-01

    Full Text Available Ulnar neuropathy at the wrist (UNW is rare, and always challenging to localize. To increase the sensitivity and specificity of the diagnosis of UNW many authors advocate the stimulation of the ulnar nerve (UN in the segment of the wrist and palm. The focus of this paper is to present a modified and simplified technique of sensory nerve conduction (SNC of the UN in the wrist and palm segments and demonstrate the validity of this technique in the study of five cases of type III UNW. The SNC of UN was performed antidromically with fifth finger ring recording electrodes. The UN was stimulated 14 cm proximal to the active electrode (the standard way and 7 cm proximal to the active electrode. The normal data from amplitude and conduction velocity (CV ratios between the palm to finger and wrist to finger segments were obtained. Normal amplitude ratio was 1.4 to 0.76. Normal CV ratio was 0.8 to 1.23.We found evidences of abnormal SNAP amplitude ratio or substantial slowing of UN sensory fibers across the wrist in 5 of the 5 patients with electrophysiological-definite type III UNW.

  9. Palm to Finger Ulnar Sensory Nerve Conduction.

    Science.gov (United States)

    Davidowich, Eduardo; Nascimento, Osvaldo J M; Orsini, Marco; Pupe, Camila; Pessoa, Bruno; Bittar, Caroline; Pires, Karina Lebeis; Bruno, Carlos; Coutinho, Bruno Mattos; de Souza, Olivia Gameiro; Ribeiro, Pedro; Velasques, Bruna; Bittencourt, Juliana; Teixeira, Silmar; Bastos, Victor Hugo

    2015-12-29

    Ulnar neuropathy at the wrist (UNW) is rare, and always challenging to localize. To increase the sensitivity and specificity of the diagnosis of UNW many authors advocate the stimulation of the ulnar nerve (UN) in the segment of the wrist and palm. The focus of this paper is to present a modified and simplified technique of sensory nerve conduction (SNC) of the UN in the wrist and palm segments and demonstrate the validity of this technique in the study of five cases of type III UNW. The SNC of UN was performed antidromically with fifth finger ring recording electrodes. The UN was stimulated 14 cm proximal to the active electrode (the standard way) and 7 cm proximal to the active electrode. The normal data from amplitude and conduction velocity (CV) ratios between the palm to finger and wrist to finger segments were obtained. Normal amplitude ratio was 1.4 to 0.76. Normal CV ratio was 0.8 to 1.23.We found evidences of abnormal SNAP amplitude ratio or substantial slowing of UN sensory fibers across the wrist in 5 of the 5 patients with electrophysiological-definite type III UNW.

  10. Visual Foraging With Fingers and Eye Gaze.

    Science.gov (United States)

    Jóhannesson, Ómar I; Thornton, Ian M; Smith, Irene J; Chetverikov, Andrey; Kristjánsson, Árni

    2016-03-01

    A popular model of the function of selective visual attention involves search where a single target is to be found among distractors. For many scenarios, a more realistic model involves search for multiple targets of various types, since natural tasks typically do not involve a single target. Here we present results from a novel multiple-target foraging paradigm. We compare finger foraging where observers cancel a set of predesignated targets by tapping them, to gaze foraging where observers cancel items by fixating them for 100 ms. During finger foraging, for most observers, there was a large difference between foraging based on a single feature, where observers switch easily between target types, and foraging based on a conjunction of features where observers tended to stick to one target type. The pattern was notably different during gaze foraging where these condition differences were smaller. Two conclusions follow: (a) The fact that a sizeable number of observers (in particular during gaze foraging) had little trouble switching between different target types raises challenges for many prominent theoretical accounts of visual attention and working memory. (b) While caveats must be noted for the comparison of gaze and finger foraging, the results suggest that selection mechanisms for gaze and pointing have different operational constraints.

  11. Small-scale behavior of single gravity-driven fingers in an initially dry fracture

    Energy Technology Data Exchange (ETDEWEB)

    Nicholl, M.J.; Glass, R.J.; Nguyen, H.A.

    1992-12-31

    Experiments investigating the behavior of individual, gravity-driven fingers in an initially dry, rough-walled analog fracture are presented. Fingers were initiated from constant flow to a point source. Finger structure is described in detail; specific phenomena observed include: desaturation behind the finger-tip, variation in finger path, intermittent flow structures, finger-tip bifurcation, and formation of dendritic sub-fingers. Measurements were made of finger-tip velocity, finger width, and finger-tip length. Non-dimensional forms of the measured variables are analyzed relative to the independent parameters, flow rate and gravitational gradient.

  12. Viscous fingering with partially miscible fluids

    Science.gov (United States)

    Fu, X.; Cueto-Felgueroso, L.; Juanes, R.

    2015-12-01

    When a less viscous fluid displaces a more viscous fluid, the contrast in viscosity destabilizes the interface between the two fluids, leading to the formation of fingers. Experimental and numerical studies of viscous fingering have focused on fluids that are either fully miscible (e.g. water and glycerol) or perfectly immiscible (e.g. water and oil). In practice, however, the miscibility of two fluids can change appreciably with temperature and pressure, and often falls into the case of partial miscibility, where two fluids have limited solubility in each other (e.g. CO2 and water). Following our recent work for miscible systems (Jha et al., PRL 2011, 2013) and immiscible systems (Cueto-Felgueroso and Juanes, PRL 2012, JFM 2014), here we propose a phase-field model for fluid-fluid displacements in a porous medium, when the two fluids have limited (but nonzero) solubility in one another. In our model, partial miscibility is characterized through the design of the thermodynamic free energy of the two-fluid system. We express the model in dimensionless form and elucidate the key dimensionless groups that control the behavior of the system. We present high-resolution numerical simulations of the model applied to the viscous fingering problem. On one hand, we demonstrate the effect of partial miscibility on the hydrodynamic instability. On the other, we elucidate the role of the degree of fingering on the rate of mutual fluid dissolution. Figure caption: final snapshots in simulations of viscous fingering with a two-fluid system mimicking that of CO2 and water. The colormap corresponds to the concentration of CO2. A band of less viscous gas phase rich in CO2 (red) displaces through the more viscous liquid phase that is undersaturated with CO2 (blue). At the fluid interface, an exchange of CO2 occurs as a result of local chemical potentials that drives the system towards thermodynamic equilibrium. This results in a shrinkage of gas phase as well as a local increase in

  13. Proteomic Analysis of Phosphoproteins in the Rice Nucleus During the Early Stage of Seed Germination.

    Science.gov (United States)

    Li, Ming; Yin, Xiaojian; Sakata, Katsumi; Yang, Pingfang; Komatsu, Setsuko

    2015-07-01

    The early stage of seed germination is the first step in the plant life cycle without visible morphological change. To investigate the mechanism controlling the early stage of rice seed germination, we performed gel-and label-free nuclear phosphoproteomics. A total of 3467 phosphopeptides belonging to 102 nuclear phosphoproteins from rice embryos were identified. Protein-synthesis-related proteins were mainly phosphorylated. During the first 24 h following imbibition, 115 nuclear phosphoproteins were identified, and significant changes in the phosphorylation level over time were observed in 29 phosphoproteins. Cluster analysis indicated that nucleotide-binding proteins and zinc finger CCCH- and BED-type proteins increased in abundance during the first 12 h of imbibition and then decreased. The in silico protein-protein interactions for 29 nuclear phosphoproteins indicated that the Sas10/Utp3 protein, which functions in snoRNA binding and gene silencing, was the center of the phosphoprotein network in nuclei. The germination rate of seeds was significantly slowed with phosphatase inhibitor treatment. The mRNA expression of the zinc finger CCCH-type protein did not change, and the zinc finger BED-type protein was upregulated in rice embryos during the early stage of germination with phosphatase inhibitor treatment. These results suggest that the phosphorylation and dephosphorylation of nuclear proteins are involved in rice seed germination. Furthermore, transcription factors such as zinc finger CCCH- and BED-type proteins might play a key role through nuclear phosphoproteins, and Sas10/Utp3 protein might interact with nuclear phosphoproteins in rice embryos to mediate the early stage of seed germination.

  14. Chelators for investigating zinc metalloneurochemistry.

    Science.gov (United States)

    Radford, Robert J; Lippard, Stephen J

    2013-04-01

    The physiology and pathology of mobile zinc signaling has become an important topic in metalloneurochemistry. To study the action of mobile zinc effectively, specialized tools are required that probe the temporal and positional changes of zinc ions within live tissue and cells. In the present article we describe the design and implementation of selective zinc chelators as antagonists to interrogate the function of mobile zinc, with an emphasis on the pools of vesicular zinc in the terminals of hippocampal mossy fiber buttons.

  15. cDNA cloning, characterization and expression analysis of DTX2, a human WWE and RING-finger gene, in human embryos.

    Science.gov (United States)

    Yi, Zhengfang; Yi, Tingfang; Wu, Zirong

    2006-06-01

    The WWE domain is a conserved globular domain in several proteins and predicted to mediate specificprotein-protein interactions in ubiquitin and ADP ribose conjugation systems. The RING domain is a conserved and specialized zinc-finger motif with 40-60 residues binding to two zinc atoms, which is also probably involved in mediating protein-protein interactions. Here, from human fetal heart cDNA library, we identified DTX2, a human WWE & RING-finger gene, with high similarity with its homologues. Evaluation of full-length cDNA obtained by RACE indicated it encodes a protein composed of two WWE domains and a RING-finger region. The DTX2 gene located in human chromosome 7q11.23 spanning approximately 44.3 kb on the genome and the deduced protein is 622 amino acids. Northern analysis revealed DTX2 was expressed in the 18-week, 22.5-week human embryo hearts and adult hearts, especially with high levels in the 18-week and adult hearts. Taken together, these results indicate that DTX2 is a gene encoding a WWE-RING-finger protein and involved in regulating heart development and heart functions.

  16. C3HC4-type RING finger protein NbZFP1 is involved in growth and fruit development in Nicotiana benthamiana.

    Directory of Open Access Journals (Sweden)

    Wenxian Wu

    Full Text Available C3HC4-type RING finger proteins constitute a large family in the plant kingdom and play important roles in various physiological processes of plant life. In this study, a C3HC4-type zinc finger gene was isolated from Nicotiana benthamiana. Sequence analysis indicated that the gene encodes a 24-kDa protein with 191 amino acids containing one typical C3HC4-type zinc finger domain; this gene was named NbZFP1. Transient expression of pGDG-NbZFP1 demonstrated that NbZFP1 was localized to the chloroplast, especially in the chloroplasts of cells surrounding leaf stomata. Virus-induced gene silencing (VIGS analysis indicated that silencing of NbZFP1 hampered fruit development, although the height of the plants was normal. An overexpression construct was then designed and transferred into Nicotiana benthamiana, and PCR and Southern blot showed that the NbZFP1 gene was successfully integrated into the Nicotiana benthamiana genome. The transgenic lines showed typical compactness, with a short internode length and sturdy stems. This is the first report describing the function of a C3HC4-type RING finger protein in tobacco.

  17. The role of fingers in number processing in young children

    Directory of Open Access Journals (Sweden)

    Anne eLafay

    2013-07-01

    Full Text Available The aim of the present study was to investigate the relationship between finger counting and numerical processing in 4- to 7-year-old children. Children were assessed on a variety of numerical tasks and we examined the correlations between their rates of success and their frequency of finger use in a counting task. We showed that children’s performance on finger pattern comparison and identification tasks did not correlate with the frequency of finger use. However, this last variable correlated with the percentages of correct responses in an enumeration task (i.e., Give-N task, even when the age of children was entered as a covariate in the analysis. Despite this correlation, we showed that some children who never used their fingers in the counting task were able to perform optimally in the enumeration task. Overall, our results support the conclusion that finger counting is useful but not necessary to develop accurate symbolic numerical skills. Moreover, our results suggest that the use of fingers in a counting task is related to the ability of children in a dynamic enumeration task but not to static tasks involving recognition or comparison of finger patterns. Therefore, it could be that the link between fingers and numbers remain circumscribed to counting tasks and do not extent to static finger montring situations.

  18. Individual finger contribution in submaximal voluntary contraction of gripping.

    Science.gov (United States)

    Kong, Yong-Ku; Lee, Kyung-Sun; Kim, Dae-Min; Jung, Myung-Chul

    2011-11-01

    The objective of this study was to evaluate individual finger force and contribution to a gripping force, the difference between actual and expected finger forces and subjective discomfort rating at 10 different submaximal voluntary contraction (%MVC) levels (10-100% in 10 increments). Seventy-two participants randomly exerted gripping force with a multi-finger force measurement system. The individual finger force, gripping force and discomfort increased as %MVC levels increased. The middle and ring fingers exerted more force and contributed to a gripping force more than the index and little fingers due to their larger mass fractions of the digit flexor muscles. It was apparent at MVC; however, the index finger increased its contribution and exerted even more force than expected at more than 50% MVC. Subjective discomfort supported the results of the objective measures. This could explain the conflicting findings between index and ring fingers in previous finger contribution studies. STATEMENT OF RELEVANCE: Hand tool design is of special interest in ergonomics due to its association with musculoskeletal disorders in the hand. This study reveals a different contribution pattern of the fingers in submaximal voluntary contraction of gripping exertion.

  19. Torque Control of Underactuated Tendon-driven Robotic Fingers

    Science.gov (United States)

    Abdallah, Muhammad E. (Inventor); Ihrke, Chris A. (Inventor); Reiland, Matthew J. (Inventor); Wampler, Charles W. (Inventor); Diftler, Myron A. (Inventor); Platt, Robert (Inventor); Bridgwater, Lyndon (Inventor)

    2013-01-01

    A robotic system includes a robot having a total number of degrees of freedom (DOF) equal to at least n, an underactuated tendon-driven finger driven by n tendons and n DOF, the finger having at least two joints, being characterized by an asymmetrical joint radius in one embodiment. A controller is in communication with the robot, and controls actuation of the tendon-driven finger using force control. Operating the finger with force control on the tendons, rather than position control, eliminates the unconstrained slack-space that would have otherwise existed. The controller may utilize the asymmetrical joint radii to independently command joint torques. A method of controlling the finger includes commanding either independent or parameterized joint torques to the controller to actuate the fingers via force control on the tendons.

  20. Experiments of periodic forcing of Saffman-Taylor fingers

    Science.gov (United States)

    Torralba, M.; Ortín, J.; Hernández-Machado, A.; Poiré, E. Corvera

    2008-03-01

    We report on an experimental study of long normal Saffman-Taylor fingers subject to periodic forcing. The sides of the finger develop a low amplitude, long wavelength instability. We discuss the finger response in stationary and nonstationary situations, as well as the dynamics towards the stationary states. The response frequency of the instability increases with forcing frequency at low forcing frequencies, while, remarkably, it becomes independent of forcing frequency at large forcing frequencies. This implies a process of wavelength selection. These observations are in good agreement with previous numerical results reported in [Ledesma-Aguilar , Phys. Rev. E 71, 016312 (2005)]. We also study the average value of the finger width, and its fluctuations, as a function of forcing frequency. The average finger width is always smaller than the width of the steady-state finger. Fluctuations have a nonmonotonic behavior with a maximum at a particular frequency.

  1. A Color Based Touchless Finger Mouse

    Directory of Open Access Journals (Sweden)

    Kah-Meng Kwong

    2012-01-01

    Full Text Available People work with computers almost anytime, everywhere  in the current trend. However, continuously controlling a computer with mouse for a long time might cause much strains to people’s wrist. This work proposes a touchless finger mouse using webcam. A marker with different colours representing different actions is used. The webcam will capture the information on the marker and trigger the associated actions. This prototype is proven to be able to perform most of the actions a normal mouser can perform.

  2. Hybrid-Actuated Finger Prosthesis with Tactile Sensing

    OpenAIRE

    2013-01-01

    Finger prostheses are devices developed to emulate the functionality of natural human fingers. On top of their aesthetic appearance in terms of shape, size and colour, such biomimetic devices require a high level of dexterity. They must be capable of gripping an object, and even manipulating it in the hand. This paper presents a biomimetic robotic finger actuated by a hybrid mechanism and integrated with a tactile sensor. The hybrid actuation mechanism comprises a DC micromotor and a Shape Me...

  3. BILATERAL VOLLEYBALL-RELATED DEFORMITY OF THE LITTLE FINGERS: MALLET FINGER AND CLINODACTYLY MIMIC

    Directory of Open Access Journals (Sweden)

    Mustafa Uslu

    2011-03-01

    Full Text Available A 14-year-old male high school volleyball player was seen to evaluate right- and left-hand little-finger distal interphalangeal joint deformity and pain. His symptoms began during his second season of competitive play. The distal interphalangeal (DIP joints of the little fingers flexed 20-30°, and a 10-15° valgus deformity was seen at the same joints. Pain was relieved with rest but returned immediately after playing volleyball, so plain radiographs were obtained. The flexion and valgus deformity was obvious on plain radiographs and through a clinical examination. Thus, a bilateral little-finger distal phalanx base epiphysis injury was seen. This injury is characterized by a biplanar Salter Harris physeal injury; type 5 on anteroposterior radiographs and type 2 on lateral plain radiographs. The deformity occurred as a result of competitive volleyball play. To our knowledge, this is the first reported case of a bilateral biplanar physial injury of the base of distal phalanges of the little fingers. Flexion and valgus deformities of DIP joints are a result of repeated micro traumas around the physis.

  4. Exploring zinc coordination in novel zinc battery electrolytes.

    Science.gov (United States)

    Kar, Mega; Winther-Jensen, Bjorn; Forsyth, Maria; MacFarlane, Douglas R

    2014-06-14

    The coordination of zinc ions by tetraglyme has been investigated here to support the development of novel electrolytes for rechargeable zinc batteries. Zn(2+) reduction is electrochemically reversible from tetraglyme. The spectroscopic data, molar conductivity and thermal behavior as a function of zinc composition, between mole ratios [80 : 20] and [50 : 50] [tetraglyme : zinc chloride], all suggest that strong interactions take place between chloro-zinc complexes and tetraglyme. Varying the concentration of zinc chloride produces a range of zinc-chloro species (ZnClx)(2-x) in solution, which hinder full interaction between the zinc ion and tetraglyme. Both the [70 : 30] and [50 : 50] mixtures are promising electrolyte candidates for reversible zinc batteries, such as the zinc-air device.

  5. Effect of Finger Joint on Flexural Strength of Teak Wood

    Directory of Open Access Journals (Sweden)

    Bharatesh A. Danawade

    2014-01-01

    Full Text Available This paper presents the flexural properties of rectangular Burma teak wood beam without finger joint and with finger joint. Finger joints enable full utilization of wood. Finger jointing technique is also used to eliminate wood defects which weaken the strength of wood. This paper considers finger joint as defined defect and its effect on the flexural strength is determined. Teakwood is hard and heavy, seasons rapidly and has good durability. The specimens were studied under three point bending test. Both edge wise and flat wise tests were carried out. It is observed that Burma teakwood beam without finger joint is stronger than beams with finger joints. Because of finger jointing the flexural strength reduces. It can be concluded that the strength loss can improved upon by selecting suitable geometry of finger joint and a suitable adhesive. It is recognized that further studies are necessary on jointing techniques of wood and type of adhesive so as to equal the flexural strength properties of clear teak wood beams.

  6. Spatial Circular Granulation Method Based on Multimodal Finger Feature

    Directory of Open Access Journals (Sweden)

    Jinfeng Yang

    2016-01-01

    Full Text Available Finger-based personal identification has become an active research topic in recent years because of its high user acceptance and convenience. How to reliably and effectively fuse the multimodal finger features together, however, has still been a challenging problem in practice. In this paper, viewing the finger trait as the combination of a fingerprint, finger vein, and finger-knuckle-print, a new multimodal finger feature recognition scheme is proposed based on granular computing. First, the ridge texture features of FP, FV, and FKP are extracted using Gabor Ordinal Measures (GOM. Second, combining the three-modal GOM feature maps in a color-based manner, we then constitute the original feature object set of a finger. To represent finger features effectively, they are granulated at three levels of feature granules (FGs in a bottom-up manner based on spatial circular granulation. In order to test the performance of the multilevel FGs, a top-down matching method is proposed. Experimental results show that the proposed method achieves higher accuracy recognition rate in finger feature recognition.

  7. Acute finger injuries: part I. Tendons and ligaments.

    Science.gov (United States)

    Leggit, Jeffrey C; Meko, Christian J

    2006-03-01

    Improper diagnosis and treatment of finger injuries can cause deformity and dysfunction over time. A basic understanding of the complex anatomy of the finger and of common tendon and ligament injury mechanisms can help physicians properly diagnose and treat finger injuries. Evaluation includes a general musculoskeletal examination as well as radiography (oblique, anteroposterior, and true lateral views). Splinting and taping are effective treatments for tendon and ligament injuries. Treatment should restrict the motion of injured structures while allowing uninjured joints to remain mobile. Although family physicians are usually the first to evaluate patients with finger injuries, it is important to recognize when a referral is needed to ensure optimal outcomes.

  8. Simulation results of the grasping analysis of an underactuated finger

    Directory of Open Access Journals (Sweden)

    Niola Vincenzo

    2016-01-01

    Full Text Available The results of a number of simulations concerning the grasping analysis is presented. The grasping device consist in an under-actuated finger driven by un-extendible tendon that is one of the fingers of a mechanical prosthesis that was principally conceived as human prosthesis. The results, however, are useful for any similar finger to be used in grasping devices for industrial and agricultural applications, Aanalysis maps of the grasping were obtained which show the “robustness” of the socket. The method seems to be a suitable tool for the optimum design of such under-actuated fingers for grasping devices.

  9. The design and development of a finger joint simulator.

    Science.gov (United States)

    Joyce, Thomas J

    2016-05-01

    Artificial finger joints lack the long-term clinical success seen with hip and knee prostheses. In part, this can be explained by the challenges of rheumatoid arthritis, a progressive disease which attacks surrounding tissues as well as the joint itself. Therefore, the natural finger joints' biomechanics are adversely affected, and consequently, this imbalance due to subluxing forces further challenges any prosthesis. Many different designs of finger prosthesis have been offered over a period of greater than 50 years. Most of these designs have failed, and it is likely that many of these failures could have been identified had the prostheses been appropriately tested prior to implantation into patients. While finger joint simulators have been designed, arguably only those from a single centre have been able to reproduce clinical-type failures of the finger prostheses tested in them. This article describes the design and development of a finger simulator at Durham University, UK. It explains and justifies the engineering decisions made and thus the evolution of the finger simulator. In vitro results and their linkage to clinical-type failures are outlined to help to show the effectiveness of the simulator. Failures of finger implants in vivo continue to occur, and the need for appropriate in vitro testing of finger prostheses remains strong.

  10. Tetanus following replantation of an amputated finger: a case report

    Directory of Open Access Journals (Sweden)

    Hayashida Kenji

    2012-10-01

    Full Text Available Abstract Introduction Tetanus is an infectious disease caused by tetanus toxin produced by Clostridium tetani and induces severe neurological manifestations. We treated a patient who developed tetanus during hospitalization for replantation of an amputated finger. To the best of our knowledge, this is the first published case report of such an entity. Case presentation A 49-year-old Japanese man had an amputation of his right middle finger at the distal interphalangeal joint region in an accident at work. His middle finger was successfully replanted, but his fingertip was partially necrotized because of crushing and so additional reconstruction with a reverse digital arterial flap was performed 15 days after the injury. Tetanus developed 21 days after replantation of the middle finger, but symptoms remitted via rapid diagnosis and treatment. Conclusions In replantation after finger trauma with exposure of nerve and blood vessel bundles, concern over injuring nerves and blood vessels may prevent irrigation and debridement from being performed sufficiently; these treatments may have been insufficiently performed in this patient. It is likely that the replanted middle finger partially adhered, and Clostridium tetani colonized the partially necrotized region. Even when there is only limited soil contamination, administration of tetanus toxoid and anti-tetanus immunoglobulin is necessary when the fingers are injured outdoors and the finger nerves and blood vessels are exposed. The drugs should be administered just after replantation if the finger has been amputated. However, if clinicians pay attention to the possibility of tetanus development, treatment can be rapidly initiated.

  11. Zinc supplementation alters airway inflammation and airway hyperresponsiveness to a common allergen

    Directory of Open Access Journals (Sweden)

    Morgan Carrie I

    2011-12-01

    Full Text Available Abstract Background Zinc supplementation can modulate immunity through inhibition of NF-κB, a transcription factor that controls many immune response genes. Thus, we sought to examine the mechanism by which zinc supplementation tempers the response to a common allergen and determine its effect on allergic airway inflammation. Methods Mice were injected with zinc gluconate prior to German cockroach (GC feces (frass exposure and airway inflammation was assessed. Primary bone marrow-derived neutrophils and DMSO-differentiated HL-60 cells were used to assess the role of zinc gluconate on tumor necrosis factor (TNFα expression. NF-κB:DNA binding and IKK activity were assessed by EMSA and in vitro kinase assay. Protein levels of A20, RIP1 and TRAF6 were assessed by Western blot analysis. Establishment of allergic airway inflammation with GC frass was followed by administration of zinc gluconate. Airway hyperresponsiveness, serum IgE levels, eosinophilia and Th2 cytokine production were assessed. Results Administration of zinc gluconate prior to allergen exposure resulted in significantly decreased neutrophil infiltration and TNFα cytokine release into the airways. This correlated with decreased NF-κB activity in the whole lung. Treatment with zinc gluconate significantly decreased GC frass-mediated TNFα production from bone-marrow derived neutrophils and HL-60 cells. We confirmed zinc-mediated decreases in NF-κB:DNA binding and IKK activity in HL-60 cells. A20, a natural inhibitor of NF-κB and a zinc-fingered protein, is a potential target of zinc. Zinc treatment did not alter A20 levels in the short term, but resulted in the degradation of RIP1, an important upstream activator of IKK. TRAF6 protein levels were unaffected. To determine the application for zinc as a therapeutic for asthma, we administered zinc following the establishment of allergic airway inflammation in a murine model. Zinc supplementation decreased airway hyperresponsiveness

  12. Estetske proteze po delni amputaciji prstov: Aesthetic prostheses after partial finger amputation: Aesthetic prostheses after partial finger amputation:

    OpenAIRE

    2010-01-01

    Partial finger amputation affects patients from the functional as well as from the aesthetic point of view. In accordance with the number of amputated fingers, abilities and capabilities of the hand are reduced and the patient's self-image is altered. The amputated part of the finger can be replaced by an aesthetic silicone prosthesis, which is individually coloured and fitted. In anaplastology, as the field of aesthetic prosthetics is nowadays called, several different approaches to prosthes...

  13. Lipid Gymnastics: Tethers and Fingers in membrane

    Science.gov (United States)

    Tayebi, Lobat; Miller, Gregory; Parikh, Atul

    2009-03-01

    A significant body of evidence now links local mesoscopic structure (e.g., shape and composition) of the cell membrane with its function; the mechanisms by which cellular membranes adopt the specific shapes remain poorly understood. Among all the different structures adopted by cellular membranes, the tubular shape is one of the most surprising one. While their formation is typically attributed to the reorganization of membrane cytoskeleton, many exceptions exist. We report the instantaneous formation of tubular membrane mesophases following the hydration under specific thermal conditions. The shapes emerge in a bimodal way where we have two distinct diameter ranges for tubes, ˜20μm and ˜1μm, namely fat fingers and narrow tethers. We study the roughening of hydrated drops of 3 lipids in 3 different spontaneous curvatures at various temp. and ionic strength to figure out the dominant effect in selection of tethers and fingers. Dynamics of the tubes are of particular interest where we observe four distinct steps of birth, coiling, uncoiling and retraction with different lifetime on different thermal condition. These dynamics appear to reflect interplay between membrane elasticity, surface adhesion, and thermal or hydrodynamic gradient.

  14. Rehabilitation for bilateral amputation of fingers

    Science.gov (United States)

    Stapanian, Martin A.; Stapanian, Adrienne M.P.; Staley, Keith E.

    2010-01-01

    We describe reconstructive surgeries, therapy, prostheses, and adaptations for a patient who experienced bilateral amputation of all five fingers of both hands through the proximal phalanges in January 1992. The patient made considerable progress in the use of his hands in the 10 mo after amputation, including nearly a 120% increase in the active range of flexion of metacarpophalangeal joints. In late 1992 and early 1993, the patient had "on-top plasty" surgeries, in which the index finger remnants were transferred onto the thumb stumps, performed on both hands. The increased web space and functional pinch resulting from these procedures made many tasks much easier. The patient and occupational therapists set challenging goals at all times. Moreover, the patient was actively involved in the design and fabrication of all prostheses and adaptations or he developed them himself. Although he was discharged from occupational therapy in 1997, the patient continues to actively find new solutions for prehension and grip strength 18 yr after amputation.

  15. Rehabilitation for bilateral amputation of fingers.

    Science.gov (United States)

    Stapanian, Martin A; Stapanian, Adrienne M P; Staley, Keith E

    2010-01-01

    We describe reconstructive surgeries, therapy, prostheses, and adaptations for a patient who experienced bilateral amputation of all five fingers of both hands through the proximal phalanges in January 1992. The patient made considerable progress in the use of his hands in the 10 mo after amputation, including nearly a 120% increase in the active range of flexion of metacarpophalangeal joints. In late 1992 and early 1993, the patient had "on-top plasty" surgeries, in which the index finger remnants were transferred onto the thumb stumps, performed on both hands. The increased web space and functional pinch resulting from these procedures made many tasks much easier. The patient and occupational therapists set challenging goals at all times. Moreover, the patient was actively involved in the design and fabrication of all prostheses and adaptations or he developed them himself. Although he was discharged from occupational therapy in 1997, the patient continues to actively find new solutions for prehension and grip strength 18 yr after amputation.

  16. Segregation induced fingering instabilities in granular avalanches

    Science.gov (United States)

    Woodhouse, Mark; Thornton, Anthony; Johnson, Chris; Kokelaar, Pete; Gray, Nico

    2013-04-01

    It is important to be able to predict the distance to which a hazardous natural granular flows (e.g. snow slab avalanches, debris-flows and pyroclastic flows) might travel, as this information is vital for accurate assessment of the risks posed by such events. In the high solids fraction regions of these flows the large particles commonly segregate to the surface, where they are transported to the margins to form bouldery flow fronts. In many natural flows these bouldery margins experience a much greater frictional force, leading to frontal instabilities. These instabilities create levees that channelize the flow vastly increasing the run-out distance. A similar effect can be observed in dry granular experiments, which use a combination of small round and large rough particles. When this mixture is poured down an inclined plane, particle size segregation causes the large particles to accumulate near the margins. Being rougher, the large particles experience a greater friction force and this configuration (rougher material in front of smoother) can be unstable. The instability causes the uniform flow front to break up into a series of fingers. A recent model for particle size-segregation has been coupled to existing avalanche models through a particle concentration dependent friction law. In this talk numerical solutions of this coupled system are presented and compared to both large scale experiments carried out at the USGS flume and more controlled small scale laboratory experiments. The coupled depth-averaged model captures the accumulation of large particles at the flow front. We show this large particle accumulation at the head of the flow can lead to the break-up of the initially uniform front into a series of fingers. However, we are unable to obtain a fully grid-resolved numerical solution; the width of the fingers decreases as the grid is refined. By considering the linear stability of a steady, fully-developed, bidisperse granular layer it is shown that

  17. Finger-like voids induced by viscous fingering during phase inversion of alumina/PES/NMP suspensions

    KAUST Repository

    Wang, Bo

    2012-07-01

    The formation mechanism of phase-inversion ceramic hollow fibre membranes has not been well understood. In this paper, we report on the formation of finger-like macrovoids during non-solvent-induced phase inversion of alumina/PES/NMP suspensions. A membrane structure without such finger-like macrovoids was observed when the suspension was slowly immersed into pure ethanol or a mixture of 70. wt% NMP and 30. wt% water, whereas finger-like macrovoids occurred when the suspension was slid into the non-solvents at higher speeds. We found that the formation process of finger-like macrovoids could be fully or partially reversed when nascent membranes were taken out from water shortly after immersion, depending on the duration of the immersion. Splitting of the fingers during the formation of the macrovoids was also observed during the phase inversion of two alumina/PES/NMP suspensions. These experimental observations were not predicted by current theories of finger-like macrovoid formation in polymer membranes, but appear to mimic the well-known viscous fingering phenomenon. We therefore propose that in the phase inversion of ceramic suspensions, the viscous fingering phenomenon is an important mechanism in the formation of finger-like voids. © 2012 Elsevier B.V.

  18. Zinc level and obesity

    Directory of Open Access Journals (Sweden)

    Doaa S.E. Zaky

    2013-01-01

    Conclusion Plasma zinc concentration in obese individuals showed an inverse relationship with the waist circumference and BMI as well as serum low-density lipoprotein-cholesterol and correlated positively with high-density lipoprotein.

  19. Zinc electrode and rechargeable zinc-air battery

    Science.gov (United States)

    Ross, Jr., Philip N.

    1989-01-01

    An improved zinc electrode is disclosed for a rechargeable zinc-air battery comprising an outer frame and a porous foam electrode support within the frame which is treated prior to the deposition of zinc thereon to inhibit the formation of zinc dendrites on the external surface thereof. The outer frame is provided with passageways for circulating an alkaline electrolyte through the treated zinc-coated porous foam. A novel rechargeable zinc-air battery system is also disclosed which utilizes the improved zinc electrode and further includes an alkaline electrolyte within said battery circulating through the passageways in the zinc electrode and an external electrolyte circulation means which has an electrolyte reservoir external to the battery case including filter means to filter solids out of the electrolyte as it circulates to the external reservoir and pump means for recirculating electrolyte from the external reservoir to the zinc electrode.

  20. 21 CFR 888.3230 - Finger joint polymer constrained prosthesis.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Finger joint polymer constrained prosthesis. 888... constrained prosthesis. (a) Identification. A finger joint polymer constrained prosthesis is a device intended... generic type of device includes prostheses that consist of a single flexible across-the-joint...

  1. Compliance Analysis of an Under-Actuated Robotic Finger

    NARCIS (Netherlands)

    Wassink, Martin; Carloni, Raffaella; Stramigioli, Stefano

    2010-01-01

    Under-actuated robotic hands have multiple applications fields, like prosthetics and service robots. They are interesting for their versatility, simple control and minimal component usage. However, when external forces are applied on the finger-tip, the mechanical structure of the finger might not b

  2. A force feedback master finger in exoskeleton type

    Institute of Scientific and Technical Information of China (English)

    Fang Honggen; Liu Hong; Xie Zongwu

    2010-01-01

    In order to eliminate the drawbacks of conventional force feedback gloves,a new type of master finger has been developed.By utilizing three"four-bar mechanism joint"in series and wire coupling mechanism,the master finger transmission ratio is kept exactly 1:1.4:1 in the whole movement range and it can make active motions in both extension and flexion directions.Additionally,to assttre faster data transmission and near zero delay in the master-slave operation,a digital signal processing/field programmable gate array(DSP/FPGA-FPGA)structure with 200μs cycle time is designed.The operating modes of the master finger can be contact or non-contact,which depends on the motion states of a slave finger,free motion or constrained motion.The position control employed in non-contact mode ensures unconstrained motion and the force control adopted in contact mode guarantees natural contact sensation.To evaluate the performances of the master finger,an experiment between the master finger and a DLR/HIT dexterous finger is conducted.The results demonstrate that this new type master finger can augment telepresence.

  3. Evaluation of the finger wrinkling test: a pilot study

    NARCIS (Netherlands)

    Barneveld, van S.; Palen, van der J.; Putten, van M.J.A.M

    2010-01-01

    Purpose: Tilt table testing mainly evaluates the systemic cardiovascular part of the autonomic nervous system, while it is assumed that the finger wrinkling test assesses the peripheral part of the autonomic nervous system. In this study we explored whether the finger wrinkling test could be a usefu

  4. Evaluation of the finger wrinkling test : a pilot study

    NARCIS (Netherlands)

    van Barneveld, S.; van der Palen, J.; van Putten, M. J. A. M.

    2010-01-01

    Tilt table testing mainly evaluates the systemic cardiovascular part of the autonomic nervous system, while it is assumed that the finger wrinkling test assesses the peripheral part of the autonomic nervous system. In this study we explored whether the finger wrinkling test could be a useful test fo

  5. Robust finger vein ROI localization based on flexible segmentation.

    Science.gov (United States)

    Lu, Yu; Xie, Shan Juan; Yoon, Sook; Yang, Jucheng; Park, Dong Sun

    2013-10-24

    Finger veins have been proved to be an effective biometric for personal identification in the recent years. However, finger vein images are easily affected by influences such as image translation, orientation, scale, scattering, finger structure, complicated background, uneven illumination, and collection posture. All these factors may contribute to inaccurate region of interest (ROI) definition, and so degrade the performance of finger vein identification system. To improve this problem, in this paper, we propose a finger vein ROI localization method that has high effectiveness and robustness against the above factors. The proposed method consists of a set of steps to localize ROIs accurately, namely segmentation, orientation correction, and ROI detection. Accurate finger region segmentation and correct calculated orientation can support each other to produce higher accuracy in localizing ROIs. Extensive experiments have been performed on the finger vein image database, MMCBNU_6000, to verify the robustness of the proposed method. The proposed method shows the segmentation accuracy of 100%. Furthermore, the average processing time of the proposed method is 22 ms for an acquired image, which satisfies the criterion of a real-time finger vein identification system.

  6. Robust Finger Vein ROI Localization Based on Flexible Segmentation

    Directory of Open Access Journals (Sweden)

    Dong Sun Park

    2013-10-01

    Full Text Available Finger veins have been proved to be an effective biometric for personal identification in the recent years. However, finger vein images are easily affected by influences such as image translation, orientation, scale, scattering, finger structure, complicated background, uneven illumination, and collection posture. All these factors may contribute to inaccurate region of interest (ROI definition, and so degrade the performance of finger vein identification system. To improve this problem, in this paper, we propose a finger vein ROI localization method that has high effectiveness and robustness against the above factors. The proposed method consists of a set of steps to localize ROIs accurately, namely segmentation, orientation correction, and ROI detection. Accurate finger region segmentation and correct calculated orientation can support each other to produce higher accuracy in localizing ROIs. Extensive experiments have been performed on the finger vein image database, MMCBNU_6000, to verify the robustness of the proposed method. The proposed method shows the segmentation accuracy of 100%. Furthermore, the average processing time of the proposed method is 22 ms for an acquired image, which satisfies the criterion of a real-time finger vein identification system.

  7. Finger-vein image separation algorithms and realization with MATLAB

    Science.gov (United States)

    Gao, Xiaoyan; Ma, Junshan; Wu, Jiajie

    2010-10-01

    According to the characteristics of the finger-vein image, we adopted a series of methods to enhance the contrast of the image in order to separate the finger-vein areas from the background areas, and made prepare for the subsequent research such as feature extraction and recognition processing . The method consists of three steps: denoising, contrast enhancement and image binarization. In denoising, considering the relationship between gray levels in the adjacent areas of the finger-vein image, we adopted the Gradient Inverse Weighted Smoothing method. In contrast enhancement, we improved the conventional High Frequency Stress Filtering method and adopted a method which combined the traditional High Frequency Stress Filtering algorithm together with the Histogram Equalization. With this method, the contrast of the finger-vein area and the background area has been enhanced significantly. During the binarization process, after taking the differences of the gray levels between the different areas of the finger-vein image into consideration, we proposed a method which combined the binarization by dividing the image into several segments and the Morphological Image Processing means. Our experiment results show that after a series of processing mentioned above by using MATLAB, the finger-vein areas can be separated from the background areas obviously. We can get a vivid figure of the finger-vein which provided some references for the following research such as finger-vein image feature extraction, matching and identification.

  8. Predicted and observed finger diameters in field soils

    NARCIS (Netherlands)

    Ritsema, C.J.; Steenhuis, T.S.; Parlange, J.Y.; Dekker, L.W.

    1996-01-01

    Wetting front instability resulting in fingered flow has been found in both wettable and non-wettable soils. Laboratory research has resulted in a number of expressions for finger diameter. The applicability of one of these equations was tested for three soils where detailed soil moisture contents w

  9. The effects of vibration-reducing gloves on finger vibration.

    Science.gov (United States)

    Welcome, Daniel E; Dong, Ren G; Xu, Xueyan S; Warren, Christopher; McDowell, Thomas W

    2014-01-01

    Vibration-reducing (VR) gloves have been used to reduce the hand-transmitted vibration exposures from machines and powered hand tools but their effectiveness remains unclear, especially for finger protection. The objectives of this study are to determine whether VR gloves can attenuate the vibration transmitted to the fingers and to enhance the understanding of the mechanisms of how these gloves work. Seven adult male subjects participated in the experiment. The fixed factors evaluated include hand force (four levels), glove condition (gel-filled, air bladder, no gloves), and location of the finger vibration measurement. A 3-D laser vibrometer was used to measure the vibrations on the fingers with and without wearing a glove on a 3-D hand-arm vibration test system. This study finds that the effect of VR gloves on the finger vibration depends on not only the gloves but also their influence on the distribution of the finger contact stiffness and the grip effort. As a result, the gloves increase the vibration in the fingertip area but marginally reduce the vibration in the proximal area at some frequencies below 100 Hz. On average, the gloves reduce the vibration of the entire fingers by less than 3% at frequencies below 80 Hz but increase at frequencies from 80 to 400 Hz. At higher frequencies, the gel-filled glove is more effective at reducing the finger vibration than the air bladder-filled glove. The implications of these findings are discussed.

  10. Zinc level and obesity

    OpenAIRE

    Doaa S.E Zaky; Eman A Sultan; Mahmoud F Salim; Rana S Dawod

    2013-01-01

    Background Obesity is a chronic condition that is associated with disturbances in the metabolism of zinc. Therefore, the aim of this study was to investigate the relationship between serum zinc level and different clinical and biochemical parameters in obese individuals. Patients and methods Twenty-four individuals with BMI more than 30 kg/m 2 and 14 healthy controls (BMI < 24 kg/m 2 ) were assessed for BMI and waist circumference using anthropometric measurements. Colorimetric tes...

  11. Development of a CPM Machine for Injured Fingers.

    Science.gov (United States)

    Fu, Yili; Zhang, Fuxiang; Ma, Xin; Meng, Qinggang

    2005-01-01

    Human fingers are easy to be injured. A CPM machine is a mechanism based on the rehabilitation theory of continuous passive motion (CPM). To develop a CPM machine for the clinic application in the rehabilitation of injured fingers is a significant task. Therefore, based on the theories of evidence based medicine (EBM) and CPM, we've developed a set of biomimetic mechanism after modeling the motions of fingers and analyzing its kinematics and dynamics analysis. We also design an embedded operating system based on ARM (a kind of 32-bit RISC microprocessor). The equipment can achieve the precise control of moving scope of fingers, finger's force and speed. It can serves as a rational checking method and a way of assessment for functional rehabilitation of human hands. Now, the first prototype has been finished and will start the clinical testing in Harbin Medical University shortly.

  12. Active Finger Recognition from Surface EMG Signal Using Bayesian Filter

    Science.gov (United States)

    Araki, Nozomu; Hoashi, Yuki; Konishi, Yasuo; Mabuchi, Kunihiko; Ishigaki, Hiroyuki

    This paper proposed an active finger recognition method using Bayesian filter in order to control a myoelectric hand. We have previously proposed a finger joint angle estimation method based on measured surface electromyography (EMG) signals and a linear model. However, when we estimate 2 or more finger angles by this estimation method, the estimation angle of the inactive finger is not accurate. This is caused by interference of surface EMG signal. To solve this interference problem, we proposed active finger recognition method from the amplitude spectrum of surface EMG signal using Bayesian filter. To confirm the effectiveness of this recognition method, we developed a myoelectric hand simulator that implements proposed recognition algorithm and carried out real-time recognition experiment.

  13. Finger gnosia: a predictor of numerical abilities in children?

    Science.gov (United States)

    Noël, Marie-Pascale

    2005-10-01

    This paper aimed to test the specificity of predicting power of finger gnosia on later numerical abilities in school-age children and to contribute to the understanding of this effect. Forty-one children were tested in the beginning of Grade 1 on finger gnosia, left-right orientation (another sign of the Gerstmann "syndrome"), and global development. Fifteen months later, numerical and reading abilities were assessed. Analyses of the results indicated that, contrary to the general measures of cognitive development, performance in the finger gnosia test was a good predictor of numerical skills 1 year later but not of reading skills, which proves the specificity of that predictor. The same conclusion was also true for the left-right orientation. However, finger gnosia could equally predict performance in numerical tasks that do or do not rely heavily on finger representation or on magnitude representation. Results are discussed in terms of the localizationist and the functional hypotheses.

  14. The effects of exercise on finger extension of CVA patients.

    Science.gov (United States)

    Trombly, C A; Quintana, L A

    1983-03-01

    The choice of activity to improve finger extension of post-CVA patients is based on untested assumptions and hypotheses. In this study, using electromyography of the extrinsic finger muscles and electrogoniometry of wrist and finger joints, the effects of five types of exercise on the finger extension of post-CVA patients were documented. Results indicated that resisted and rapid exercises recruited high percentages of output of all three muscles. Slow, unresisted extension exercises preferentially recruited the extensor digitorum. No exercise caused significant immediate changes in range of motion (ROM), flexor/extensor balance, time required to open the hand, or level of activity of the extensor digitorum during opening of the hand. Resisted grasp did not limit the patients' ability to extend the fingers. Variability in percent of motor output among the subjects of this study indicates the need to monitor each patient during therapy.

  15. Finger vein image quality evaluation using support vector machines

    Science.gov (United States)

    Yang, Lu; Yang, Gongping; Yin, Yilong; Xiao, Rongyang

    2013-02-01

    In an automatic finger-vein recognition system, finger-vein image quality is significant for segmentation, enhancement, and matching processes. In this paper, we propose a finger-vein image quality evaluation method using support vector machines (SVMs). We extract three features including the gradient, image contrast, and information capacity from the input image. An SVM model is built on the training images with annotated quality labels (i.e., high/low) and then applied to unseen images for quality evaluation. To resolve the class-imbalance problem in the training data, we perform oversampling for the minority class with random-synthetic minority oversampling technique. Cross-validation is also employed to verify the reliability and stability of the learned model. Our experimental results show the effectiveness of our method in evaluating the quality of finger-vein images, and by discarding low-quality images detected by our method, the overall finger-vein recognition performance is considerably improved.

  16. Geometric approach to viscous fingering on a cone

    CERN Document Server

    Miranda, J A

    2003-01-01

    We study fluid flow and the formation of viscous fingering patterns on a two-dimensional conical background space, defined as the conical Hele-Shaw cell. We approach the problem geometrically and study how the nontrivial topological structure of the conical cell affects the evolution of the interface separating two viscous fluids. We perform a perturbative weakly nonlinear analysis of the problem and derive a mode-coupling differential equation which describes fluid-fluid interface behaviour. Our nonlinear study predicts the formation of fingering structures in which fingers of different lengths compete and split at their tips. The shape of the emerging patterns show a significant sensitivity to variations in the cell's topological features, which can be monitored by changing the cone opening angle. We find that for increasingly larger values of the opening angle, finger competition is inhibited while finger tip-splitting is enhanced.

  17. Mechanics of finger-tip electronics.

    Science.gov (United States)

    Su, Yewang; Li, Rui; Cheng, Huanyu; Ying, Ming; Bonifas, Andrew P; Hwang, Keh-Chih; Rogers, John A; Huang, Yonggang

    2013-10-28

    Tactile sensors and electrotactile stimulators can provide important links between humans and virtual environments, through the sensation of touch. Soft materials, such as low modulus silicones, are attractive as platforms and support matrices for arrays sensors and actuators that laminate directly onto the fingertips. Analytic models for the mechanics of three dimensional, form-fitting finger cuffs based on such designs are presented here, along with quantitative validation using the finite element method. The results indicate that the maximum strains in the silicone and the embedded devices are inversely proportional to the square root of radius of curvature of the cuff. These and other findings can be useful in formulating designs for these and related classes of body-worn, three dimensional devices.

  18. Vertical finger displacement is reduced in index finger tapping during repeated bout rate enhancement

    DEFF Research Database (Denmark)

    Mora-Jensen, Mark Holten; Madeleine, Pascal; Hansen, Ernst Albin

    2017-01-01

    , and 2) the hypotheses that the faster tapping was accompanied by changed vertical displacement of the fingertip and by changed peak force during tapping. Right-handed, healthy, and recreationally active individuals (n=24) performed two 3-min index finger tapping bouts at freely chosen tapping frequency......, separated by 10 min rest. The recently reported phenomenon of repeated bout rate enhancement was replicated. The faster tapping (8.8±18.7 taps min-1, corresponding to 6.0±11.0%, p=.033) was accompanied by reduced vertical displacement (1.6±2.9 mm, corresponding to 6.3±14.9%, p=.012) of the fingertip....... Concurrently, peak force was unchanged. The present study points at separate control mechanisms governing kinematics and kinetics during finger tapping....

  19. Treatment of zinc deficiency without zinc fortification

    Institute of Scientific and Technical Information of China (English)

    Donald OBERLEAS; Barbara F. HARLAND

    2008-01-01

    Zinc (Zn) deficiency in animals became of interest until the 1950s. In this paper, progresses in researches on physi-ology of Zn deficiency in animals, phytate effect on bioavailability of Zn, and role of phytase in healing Zn deficiency of animals were reviewed. Several studies demonstrated that Zn is recycled via the pancreas; the problem of Zn deficiency was controlled by Zn homeostasis. The endogenous secretion of Zn is considered as an important factor influencing Zn deficiency, and the critical molar ratio is 10. Phytate (inositol hexaphosphate) constituted up to 90% of the organically bound phosphorus in seeds. Great improvement has been made in recent years on isolating and measuring phytate, and its structure is clear. Phytate is considered to reduce Zn bioavailability in animal. Phytase is the enzyme that hydrolyzes phytate and is present in yeast, rye bran, wheat bran, barley, triticale, and many bacteria and fungi. Zinc nutrition and bioavailability can be enhanced by addition of phytase to animal feeds. Therefore, using phytase as supplements, the most prevalent Zn deficiency in animals may be effectively corrected without the mining and smelting of several tons of zinc daily needed to correct this deficiency by fortification worldwide.

  20. Use of an electronic finger dosimeters in optimisation of finger doses

    Energy Technology Data Exchange (ETDEWEB)

    Martin, C. J.; Whitby, M.; Hilditch, T.; Anstee, D.

    2002-07-01

    Optimisation of radiation doses to the hands is problematic. Doses recorded by passive dosimeters include contributions from many different manipulations, so it is difficult to ascertain which actions make significant contributions to the doses received. A device for electronic monitoring of finger doses called and Advanced Extremity Gamma Instrumentation system (AEGIS), which records instantaneous dose rates at the finger, can assist in providing this information. AEGIS ha been employed in a number of hospital departments where radionuclides are used, including a large hospital radionuclide dispensary and nuclear medicine departments. Data were recorded throughout complete sessions and analysed to identify the actions which made the most significant contributions to doses. Study of the patterns of radiation dose rate has enabled finger doses in the radionuclide dispensary to be optimised through raising staff awareness of doses from each manipulation. Optimisation has been achieved through changes following evaluation of alternative manipulation techniques and use of shielding devices. AEGIS has also been used to determine dose distributions across the hand and establish a relationship between the dose to the most exposed part and that at the monitoring position. For radionuclide dispensary staff, the tip of the index finger on the dominant hand receives the highest dose. Studies carried out in nuclear medicine departments have shown that withdrawals of radiopharmaceutical into a syringe, where a shield is not used routinely, tend to make the largest contribution to the dose. This can vary from 5 to 500 {mu}Gy per manipulation. Doses for injections varied from 1.5 to 300{mu}Gy. (Author)

  1. Intensity Variation Normalization for Finger Vein Recognition Using Guided Filter Based Singe Scale Retinex

    OpenAIRE

    Shan Juan Xie; Yu Lu; Sook Yoon; Jucheng Yang; Dong Sun Park

    2015-01-01

    Finger vein recognition has been considered one of the most promising biometrics for personal authentication. However, the capacities and percentages of finger tissues (e.g., bone, muscle, ligament, water, fat, etc.) vary person by person. This usually causes poor quality of finger vein images, therefore degrading the performance of finger vein recognition systems (FVRSs). In this paper, the intrinsic factors of finger tissue causing poor quality of finger vein images are analyzed, and an int...

  2. The Silent Language of Fingers and Hands-The Role of Finger and Hand Motions in Non-verbal Communication

    Institute of Scientific and Technical Information of China (English)

    周樱

    2013-01-01

    This paper engages in the discussion about nonverbal intercultural communication from the perspective of hand and finger gestures, probing into the functions of non-verbal cues in cross-cultural interaction, talking over two different attitudes to⁃ward gestures, and giving a detailed analysis about the silent language of some specific gestures with fingers and hands in different cultures.

  3. The equine herpesvirus 1 gene 63 RING finger protein partially complements Vmw110, its herpes simplex virus type 1 counterpart.

    Science.gov (United States)

    Everett, R; Orr, A; Elliott, M

    1995-09-01

    All alpha herpesviruses of known DNA sequence have been found to encode a protein with similarities to immediate early protein Vmw110 (ICP0) of herpes simplex virus type 1 (HSV-1). The conserved portion of this family of proteins is a characteristic zinc binding module, known as a RING finger or C3HC4 domain. Examples of RING finger domains occur in many other proteins of diverse evolutionary origin and function. Recently, the solution structure of the equine herpesvirus 1 (EHV-1) RING finger protein, encoded by gene 63, has been solved. To investigate whether this structure could be considered to be a paradigm of herpesvirus RING domains, we have constructed a recombinant HSV-1 which expresses the EHV-1 gene 63 protein (EHVg63) in place of Vmw110. Comparison of the growth properties of the recombinant with those of wild-type and Vmw110-defective viruses indicates that EHVg63 is able to fulfil partially, but not completely, the roles of Vmw110 during virus growth in tissue culture.

  4. The bioavailability of four zinc oxide sources and zinc sulphate in broiler chickens

    OpenAIRE

    Veldkamp, T.; Diepen, van, C.A.; Bikker, P.

    2014-01-01

    Zinc is an essential trace element for all farm animal species. It is commonly included in animal diets as zinc oxide, zinc sulphate or organically bound zinc. Umicore Zinc Chemicals developed zinc oxide products with different mean particle sizes. Umicore Zinc Chemicals requested Wageningen UR Livestock Research to determine the bioavailability of four zinc oxide sources and zinc sulphate in broiler chickens. A precise estimate of the bioavailability of zinc sources is required both for fulf...

  5. TIF1alpha: a possible link between KRAB zinc finger proteins and nuclear receptors

    DEFF Research Database (Denmark)

    Le Douarin, B; You, J; Nielsen, Anders Lade;

    1998-01-01

    -protein interaction sites. Of these, one specifically interacts with NRs bound to their agonistic ligand and not with NR mutants that are defective in the AF-2 activity. Immediately adjacent to this 'NR box', TIF1alpha contains an interaction site for members of the chromatin organization modifier (chromo) family, HP...

  6. Cooperation of the BTB-Zinc finger protein, Abrupt, with cytoskeletal regulators in Drosophila epithelial tumorigenesis

    Directory of Open Access Journals (Sweden)

    Nezaket Turkel

    2015-08-01

    Full Text Available The deregulation of cell polarity or cytoskeletal regulators is a common occurrence in human epithelial cancers. Moreover, there is accumulating evidence in human epithelial cancer that BTB-ZF genes, such as Bcl6 and ZBTB7A, are oncogenic. From our previous studies in the vinegar fly, Drosophila melanogaster, we have identified a cooperative interaction between a mutation in the apico-basal cell polarity regulator Scribble (Scrib and overexpression of the BTB-ZF protein Abrupt (Ab. Herein, we show that co-expression of ab with actin cytoskeletal regulators, RhoGEF2 or Src64B, in the developing eye-antennal epithelial tissue results in the formation of overgrown amorphous tumours, whereas ab and DRac1 co-expression leads to non-cell autonomous overgrowth. Together with ab, these genes affect the expression of differentiation genes, resulting in tumours locked in a progenitor cell fate. Finally, we show that the expression of two mammalian genes related to ab, Bcl6 and ZBTB7A, which are oncogenes in mammalian epithelial cancers, significantly correlate with the upregulation of cytoskeletal genes or downregulation of apico-basal cell polarity neoplastic tumour suppressor genes in colorectal, lung and other human epithelial cancers. Altogether, this analysis has revealed that upregulation of cytoskeletal regulators cooperate with Abrupt in Drosophila epithelial tumorigenesis, and that high expression of human BTB-ZF genes, Bcl6 and ZBTB7A, shows significant correlations with cytoskeletal and cell polarity gene expression in specific epithelial tumour types. This highlights the need for further investigation of the cooperation between these genes in mammalian systems.

  7. Re-engineering the zinc fingers of PRDM9 reverses hybrid sterility in mice.

    Science.gov (United States)

    Davies, Benjamin; Hatton, Edouard; Altemose, Nicolas; Hussin, Julie G; Pratto, Florencia; Zhang, Gang; Hinch, Anjali Gupta; Moralli, Daniela; Biggs, Daniel; Diaz, Rebeca; Preece, Chris; Li, Ran; Bitoun, Emmanuelle; Brick, Kevin; Green, Catherine M; Camerini-Otero, R Daniel; Myers, Simon R; Donnelly, Peter

    2016-02-11

    The DNA-binding protein PRDM9 directs positioning of the double-strand breaks (DSBs) that initiate meiotic recombination in mice and humans. Prdm9 is the only mammalian speciation gene yet identified and is responsible for sterility phenotypes in male hybrids of certain mouse subspecies. To investigate PRDM9 binding and its role in fertility and meiotic recombination, we humanized the DNA-binding domain of PRDM9 in C57BL/6 mice. This change repositions DSB hotspots and completely restores fertility in male hybrids. Here we show that alteration of one Prdm9 allele impacts the behaviour of DSBs controlled by the other allele at chromosome-wide scales. These effects correlate strongly with the degree to which each PRDM9 variant binds both homologues at the DSB sites it controls. Furthermore, higher genome-wide levels of such 'symmetric' PRDM9 binding associate with increasing fertility measures, and comparisons of individual hotspots suggest binding symmetry plays a downstream role in the recombination process. These findings reveal that subspecies-specific degradation of PRDM9 binding sites by meiotic drive, which steadily increases asymmetric PRDM9 binding, has impacts beyond simply changing hotspot positions, and strongly support a direct involvement in hybrid infertility. Because such meiotic drive occurs across mammals, PRDM9 may play a wider, yet transient, role in the early stages of speciation.

  8. Highly efficient modification of beta-lactoglobulin (BLG) gene via zinc-finger nucleases in cattle

    Institute of Scientific and Technical Information of China (English)

    Shengli Yu; Junjie Luo; Zhiyuan Song; Fangrong Ding; Yunping Dai; Ning Li

    2011-01-01

    Dear Editor,Gene targeting is in widespread use as a gold standard for determining the function of genes in mice and human embryonic stem cells [1].However,the poor efficiency of this technology has hindered its application to domestic animals,for which embryonic stem cells are not available.Although gene-targeted large domestic animals have been produced successfully by combination of homologous recombination-based targeting strategy and cloning [2-4],the efficiency is very low and,more importantly,the disruption of the targeted gene is usually mono-allelic.It thus takes a long time to obtain a null mutant.

  9. Brittle Cornea Syndrome Associated with a Missense Mutation in the Zinc-Finger 469 Gene

    DEFF Research Database (Denmark)

    Christensen, Anne Elisabeth; Knappskog, Per Morten; Midtbø, Marit

    2010-01-01

    Purpose: To investigate the diverse clinical manifestations, identify the causative mutation and explain the association with red hair in a family with brittle cornea syndrome (BCS). Methods: Eight family members in three generations underwent ophthalmic, dental, and general medical examination, ...

  10. FYVE zinc-finger proteins in the plant model Arabidopsis thaliana

    DEFF Research Database (Denmark)

    Jensen, R B; La Cour, T; Albrethsen, J

    2001-01-01

    , specificity for PtdIns3P was obtained by mutagenic conversion of the variant into a classic FYVE domain (Asn(4),Tyr(6)-->His(4),Arg(6)). Separate substitutions of the variant residues were not sufficient to impose preferential binding of PtdIns3P, suggesting a co-operative effect of these residues in binding...

  11. Mining the O-glycoproteome using zinc-finger nuclease-glycoengineered SimpleCell lines

    DEFF Research Database (Denmark)

    Steentoft, Catharina; Vakhrushev, Sergey Y; Vester-Christensen, Malene B;

    2011-01-01

    -glycopeptides from total cell lysates using lectin chromatography and nanoflow liquid chromatography-mass spectrometry (nLC-MS/MS) with electron transfer dissociation fragmentation. We identified >100 O-glycoproteins with >350 O-glycan sites (the great majority previously unidentified), including a GalNAc O......-glycan linkage to a tyrosine residue. The SimpleCell method should facilitate analyses of important functions of protein glycosylation. The strategy is also applicable to other O-glycoproteomes....

  12. RABBIT EARS, encoding a SUPERMAN-like zinc finger protein, regulates petal development in Arabidopsis thaliana.

    Science.gov (United States)

    Takeda, Seiji; Matsumoto, Noritaka; Okada, Kiyotaka

    2004-01-01

    Floral organs usually initiate at fixed positions in concentric whorls within a flower. Although it is understood that floral homeotic genes determine the identity of floral organs, the mechanisms of position determination and the development of each organ have not been clearly explained. We isolated a novel mutant, rabbit ears (rbe), with defects in petal development. In rbe, under-developed petals are formed at the correct position in a flower, and the initiation of petal primordia is altered. The rbe mutation affects the second whorl organ shapes independently of the organ identity. RBE encodes a SUPERMAN-like protein and is located in the nucleus, and thus may be a transcription factor. RBE transcripts are expressed in petal primordia and their precursor cells, and disappeared at later stages. When cells that express RBE are ablated genetically, no petal primordia arise. RBE is not expressed in ap1-1 and ptl-1 mutants, indicating that RBE acts downstream of AP1 and PTL genes. These characteristics suggest that RBE is required for the early development of the organ primordia of the second whorl.

  13. The zinc finger protein ZAT11 modulates paraquat-induced programmed cell death in Arabidopsis thaliana

    NARCIS (Netherlands)

    Qureshi, Muhammad Kamran; Sujeeth, Neerakkal; Gechev, Tsanko S.; Hille, Jacques; Liu, J.-H.

    2013-01-01

    Plants use programmed cell death (PCD) as a tool in their growth and development. PCD is also involved in defense against different kinds of stresses including pathogen attack. In both types of PCD, reactive oxygen species (ROS) play an important role. ROS is not only a toxic by-product but also act

  14. Expression of Zinc Finger Protein 804A (ZNF804A) in the brain

    DEFF Research Database (Denmark)

    Benedikz, Eirikur

    to further scrutiny. Recently ZNF804A was the first gene to achieve genome-wide significance for psychosis and several genome-wide association studies have since confirmed an association between schizophrenia and ZNF804A. The function of ZNF804A and its role in the disease are unknown. Interestingly...... the schizophrenia susceptibility genotype of ZNF804A is associated with altered connectivity in the dorsolateral prefrontal cortex, the hippocampus, and the amygdala. Altered connectivity within and between these brain regions has been associated with schizophrenia. In this study we have analyzed the mRNA levels...... of ZNF804A in different brain regions and at different ages in rats using qPCR. Our results show that expression of ZNF804A is developmentally regulated and increases significantly in the brain of embryonic day 18 rats (the developmental equivalent of a 9 week old human fetus). In cortex and cerebellum...

  15. Reduction of arsenite-enhanced ultraviolet radiation-induced DNA damage by supplemental zinc

    Energy Technology Data Exchange (ETDEWEB)

    Cooper, Karen L.; King, Brenee S.; Sandoval, Monica M.; Liu, Ke Jian; Hudson, Laurie G., E-mail: lhudson@salud.unm.edu

    2013-06-01

    Arsenic is a recognized human carcinogen and there is evidence that arsenic augments the carcinogenicity of DNA damaging agents such as ultraviolet radiation (UVR) thereby acting as a co-carcinogen. Inhibition of DNA repair is one proposed mechanism to account for the co-carcinogenic actions of arsenic. We and others find that arsenite interferes with the function of certain zinc finger DNA repair proteins. Furthermore, we reported that zinc reverses the effects of arsenite in cultured cells and a DNA repair target protein, poly (ADP-ribose) polymerase-1. In order to determine whether zinc ameliorates the effects of arsenite on UVR-induced DNA damage in human keratinocytes and in an in vivo model, normal human epidermal keratinocytes and SKH-1 hairless mice were exposed to arsenite, zinc or both before solar-simulated (ss) UVR exposure. Poly (ADP-ribose) polymerase activity, DNA damage and mutation frequencies at the Hprt locus were measured in each treatment group in normal human keratinocytes. DNA damage was assessed in vivo by immunohistochemical staining of skin sections isolated from SKH-1 hairless mice. Cell-based findings demonstrate that ssUVR-induced DNA damage and mutagenesis are enhanced by arsenite, and supplemental zinc partially reverses the arsenite effect. In vivo studies confirm that zinc supplementation decreases arsenite-enhanced DNA damage in response to ssUVR exposure. From these data we can conclude that zinc offsets the impact of arsenic on ssUVR-stimulated DNA damage in cells and in vivo suggesting that zinc supplementation may provide a strategy to improve DNA repair capacity in arsenic exposed human populations. - Highlights: • Low levels of arsenite enhance UV-induced DNA damage in human keratinocytes. • UV-initiated HPRT mutation frequency is enhanced by arsenite. • Zinc supplementation offsets DNA damage and mutation frequency enhanced by arsenite. • Zinc-dependent reduction of arsenite enhanced DNA damage is confirmed in vivo.

  16. Five- to 7-year-olds' finger gnosia and calculation abilities.

    Science.gov (United States)

    Reeve, Robert; Humberstone, Judi

    2011-01-01

    The research examined the relationship between 65 5- to 7-year-olds' finger gnosia, visuo-spatial working memory, and finger-use in solving single-digit addition problems. Their non-verbal IQ and basic reaction time were also assessed. Previous research has found significant changes in children's representational abilities between 5 and 7 years. One aim of the research was to determine whether changes in finger representational abilities (finger gnosia) occur across these ages and whether they are associated with finger-use in computation. A second aim was to determine whether visuo-spatial working memory is associated with finger gnosia and computation abilities. We used latent class profile analysis to identify patterns of similarities and differences in finger gnosia and computation/finger-use abilities. The analysis yielded four finger gnosia subgroups that differed in finger representation ability. It also yielded four finger/computation subgroups that differed in the relationship between finger-use and computation success. Analysis revealed associations between computation finger-use/success subgroups, finger gnosia subgroups, and visuo-spatial working memory. A multinomial logistic regression analysis showed that finger gnosia subgroup membership and visuo-spatial working memory uniquely contribute to a model predicting finger-use in computation group membership. The results show that finger gnosia abilities change in the early school years, and that these changes are associated with the ability to use fingers to aid computation.

  17. Five- to 7-Year-Olds’ Finger Gnosia and Calculation Abilities

    Science.gov (United States)

    Reeve, Robert; Humberstone, Judi

    2011-01-01

    The research examined the relationship between 65 5- to 7-year-olds’ finger gnosia, visuo-spatial working memory, and finger-use in solving single-digit addition problems. Their non-verbal IQ and basic reaction time were also assessed. Previous research has found significant changes in children’s representational abilities between 5 and 7 years. One aim of the research was to determine whether changes in finger representational abilities (finger gnosia) occur across these ages and whether they are associated with finger-use in computation. A second aim was to determine whether visuo-spatial working memory is associated with finger gnosia and computation abilities. We used latent class profile analysis to identify patterns of similarities and differences in finger gnosia and computation/finger-use abilities. The analysis yielded four finger gnosia subgroups that differed in finger representation ability. It also yielded four finger/computation subgroups that differed in the relationship between finger-use and computation success. Analysis revealed associations between computation finger-use/success subgroups, finger gnosia subgroups, and visuo-spatial working memory. A multinomial logistic regression analysis showed that finger gnosia subgroup membership and visuo-spatial working memory uniquely contribute to a model predicting finger-use in computation group membership. The results show that finger gnosia abilities change in the early school years, and that these changes are associated with the ability to use fingers to aid computation. PMID:22171220

  18. Five- to 7-Year-Olds’ Finger Gnosia and Calculation Abilities

    Directory of Open Access Journals (Sweden)

    Robert eReeve

    2011-12-01

    Full Text Available The research examined the relationship between 65 5- to 7-year-olds’ finger gnosia, visuo-spatial working memory and finger-use solving single digit addition problems. Their non-verbal IQ and basic RT were also assessed. Previous research has found significant changes in children’s representational abilities between five and seven years. One aim of the research was to determine whether changes in finger representational abilities (finger gnosia occur across these ages and whether they are associated with finger-use in computation. A second aim was to determine whether visuo-spatial working memory is associated with finger gnosia and computation abilities. We used latent class profile analysis to identify patterns of similarities and differences in finger gnosia and computation/finger-use abilities. The analysis yielded four finger gnosia subgroups that differed in finger representation ability. It also yielded four finger/computation subgroups that differed in the relationship between finger-use and computation success. Analysis revealed associations between computation finger-use/success subgroups, finger gnosia subgroups, and visuo-spatial working memory. A multinomial logistic regression analysis showed that finger gnosia subgroup membership and visuo-spatial working memory uniquely contribute to a model predicting finger-use in computation group membership. The results show that finger gnosia abilities change in the early school years, and that these changes are associated with the ability to use fingers to aid computation.

  19. Design Considerations for an Underactuated Robotic Finger Mechanism

    Institute of Scientific and Technical Information of China (English)

    YAO Shuangji; CECCARELLI Marco; ZHAN Qiang; ARBONE Giuseppe; LU Zhen

    2009-01-01

    A design approach is presented in this paper for underactuation in robotic finger mechanisms. The characters of underactuated finger mechanisms are introduced as based on linkage and spring systems. The feature of self-adaptive enveloping grasp by underactuated finger mechanisms is discussed with feasible in grasping unknown objects. The design problem of robotic fingers is analyzed by looking at many aspects for an optimal functionality. Design problems and requirements for underactuated mechanisms are formulated as related to human-like robotic fingers. In particular, characteristics of finger mechanisms are analyzed and optimality criteria are summarized with the aim to formulate a general design algorithm. A general multi-objective optimization design approach is applied as based on a suitable optimization problem by using suitable expressions of optimality criteria. An example is illustrated as an improvement of finger mechanism in Laboratory of Robotics and Mechatronics (LARM) Hand. Results of design outputs and grasp simulations are reported with the aim to show the practical feasibility of the proposed concepts and computations.

  20. Torque control of underactuated tendon-driven fingers

    Directory of Open Access Journals (Sweden)

    M. E. Abdallah

    2011-02-01

    Full Text Available Given an underactuated tendon-driven finger, the finger posture is underdetermined and can move freely ("flop" in a region of slack tendons. This work shows that such an underactuated finger can be operated in tendon force control (rather than position control with effective performance. The force control eliminates the indeterminate slack while commanding a parameterized space of desired torques. The torque will either push the finger to the joint limits or wrap around an external object with variable torque – behavior that is sufficient for primarily gripping fingers. In addition, introducing asymmetric joint radii to the design allows the finger to command an expanded range of joint torques and to scan an expanded set of external surfaces. This study is motivated by the design and control of the secondary fingers of the NASA-GM R2 humanoid hand.

    This paper was presented at the IFToMM/ASME International Workshop on Underactuated Grasping (UG2010, 19 August 2010, Montréal, Canada.

  1. Tangential finger forces use mechanical advantage during static grasping.

    Science.gov (United States)

    Slota, Gregory P; Latash, Mark L; Zatsiorsky, Vladimir M

    2012-02-01

    When grasping and manipulating objects, the central controller utilizes the mechanical advantage of the normal forces of the fingers for torque production. Whether the same is valid for tangential forces is unknown. The main purpose of this study was to determine the patterns of finger tangential forces and the use of mechanical advantage as a control mechanism when dealing with objects of nonuniform finger positioning. A complementary goal was to explore the interaction of mechanical advantage (moment arm) and the role a finger has as a torque agonist/antagonist with respect to external torques (±0.4 N m). Five 6-df force/torque transducers measured finger forces while subjects held a prism handle (6 cm width × 9 cm height) with and without a single finger displaced 2 cm (handle width). The effect of increasing the tangential moment arm was significant (p forces (in >70% of trials) and hence creating greater moments. Thus, the data provides evidence that the grasping system as a rule utilizes mechanical advantage for generating tangential forces. The increase in tangential force was independent of whether the finger was acting as a torque agonist or antagonist, revealing their effects to be additive.

  2. Finger vein verification system based on sparse representation.

    Science.gov (United States)

    Xin, Yang; Liu, Zhi; Zhang, Haixia; Zhang, Hong

    2012-09-01

    Finger vein verification is a promising biometric pattern for personal identification in terms of security and convenience. The recognition performance of this technology heavily relies on the quality of finger vein images and on the recognition algorithm. To achieve efficient recognition performance, a special finger vein imaging device is developed, and a finger vein recognition method based on sparse representation is proposed. The motivation for the proposed method is that finger vein images exhibit a sparse property. In the proposed system, the regions of interest (ROIs) in the finger vein images are segmented and enhanced. Sparse representation and sparsity preserving projection on ROIs are performed to obtain the features. Finally, the features are measured for recognition. An equal error rate of 0.017% was achieved based on the finger vein image database, which contains images that were captured by using the near-IR imaging device that was developed in this study. The experimental results demonstrate that the proposed method is faster and more robust than previous methods.

  3. The social and economic consequences of finger amputations.

    Science.gov (United States)

    Hovgaard, C; Angermann, P; Hovgaard, D

    1994-06-01

    120 patients with amputation of at least 1 of the 4 ulnar fingers were admitted to hospital. In none was replantation considered to be possible because of serious damage to the soft tissues and bone. 12 (3-18) years after the accident 80 percent of the patients assessed their condition as good or fair, even those with proximal amputation or loss of 2 or 3 fingers. Our observations do not support replantation when only one of the second-to-fifth fingers have been amputated.

  4. Narrow fingers in the Saffman-Taylor instability

    Science.gov (United States)

    Couder, Y.; Gerard, N.; Rabaud, M.

    1986-12-01

    Saffman-Taylor fingers with a relative width much smaller than the classical limit lambda = 0.5 are found when a small isolated bubble is located at their tip. These solutions are members of a family found by Saffman and Taylor (1958) neglecting superficial tension. Recent theories have shown that when capillary forces are taken into account an unphysical cusplike singularity would appear at the tip of all the fingers with lambda less than 0.5. Conversely, here the replacement of the tip by a small bubble makes these solutions possible. At large velocity these fingers show dendritic instability.

  5. Improving esthetics of finger prosthesis by glove silicone

    Directory of Open Access Journals (Sweden)

    Laxman Singh Kaira

    2014-01-01

    Full Text Available Finger and partial finger amputations are some of the most frequently encountered forms of partial hand loss. A high quality esthetic prosthesis with passive function can be helpful to the patient since loss or congenital absence or malformation have both a social and psychological impact on the patient. Prosthetics is an art and science, which provides lifelike appearance to the lost structures of the patient. This case report presents the fabrication of a silicone finger prosthesis, which had good suspension, adequate function, was comfortable to use and esthetically acceptable to the patient.

  6. Dorsal finger texture recognition: Investigating fixed-length SURF

    DEFF Research Database (Denmark)

    Hartung, Daniel; Kückelhahn, Jesper

    2012-01-01

    We seek to create fixed-length features from dorsal finger skin images extracted by the SURF interest point detector to combine it in the privacy enhancing helper data scheme. The source of the biometric samples is the GUC45 database which features finger vein, fingerprint and dorsal finger skin...... images for modality fusion. First, the region of interest (ROI) is extracted, after which SURF features are extracted, and finally two different approaches for creating fixed length feature vectors are applied. SURF performance on the ROI is comparable to the PolyU database reported in the literature...

  7. Fertilization in C. elegans requires an intact C-terminal RING finger in sperm protein SPE-42

    Directory of Open Access Journals (Sweden)

    Rumbley Jon N

    2011-02-01

    Full Text Available Abstract Background The C. elegans sperm protein SPE-42, a membrane protein of unknown structure and molecular function, is required for fertilization. Sperm from worms with spe-42 mutations appear normal but are unable to fertilize eggs. Sequence analysis revealed the presence of 8 conserved cysteine residues in the C-terminal cytoplasmic domain of this protein suggesting these residues form a zinc-coordinating RING finger structure. Results We made an in silico structural model of the SPE-42 RING finger domain based on primary sequence analysis and previously reported RING structures. To test the model, we created spe-42 transgenes coding for mutations in each of the 8 cysteine residues predicted to coordinate Zn++ ions in the RING finger motif. Transgenes were crossed into a spe-42 null background and protein function was measured by counting progeny. We found that all 8 cysteines are required for protein function. We also showed that sequence differences between the C-terminal 29 and 30 amino acids in C. elegans and C. briggsae SPE-42 following the RING finger domain are not responsible for the failure of the C. briggsae SPE-42 homolog to rescue C. elegans spe-42 mutants. Conclusions The results suggest that a bona fide RING domain is present at the C-terminus of the SPE-42 protein and that this motif is required for sperm-egg interactions during C. elegans fertilization. Our structural model of the RING domain provides a starting point for further structure-function analysis of this critical region of the protein. The C-terminal domain swap experiment suggests that the incompatibility between the C. elegans and C. briggsae SPE-42 proteins is caused by small amino acid differences outside the C-terminal domain.

  8. Zinc Determination in Pleural Fluid

    OpenAIRE

    Nazan DEMİR; DEMİR, Yaşar

    2000-01-01

    In this study, an enzymatic zinc determination method was applied to pleural fluid, the basis of which was the regaining of the activity of apo carbonic anhydrase by the zinc present in the sample. The method was used for pleural fluid zinc determination in order to show the application to body fluids other than serum. For this purpose, pleural fluids were obtained from 20 patients and zinc concentrations were determined. Carbonic anhydrase was purified by affinity chromatography from bovine ...

  9. The bioavailability of four zinc oxide sources and zinc sulphate in broiler chickens

    NARCIS (Netherlands)

    Veldkamp, T.; Diepen, van J.T.M.; Bikker, P.

    2014-01-01

    Zinc is an essential trace element for all farm animal species. It is commonly included in animal diets as zinc oxide, zinc sulphate or organically bound zinc. Umicore Zinc Chemicals developed zinc oxide products with different mean particle sizes. Umicore Zinc Chemicals requested Wageningen UR Live

  10. Zinc Phosphide Poisoning

    Directory of Open Access Journals (Sweden)

    Erdal Doğan

    2014-01-01

    Full Text Available Zinc phosphide has been used widely as a rodenticide. Upon ingestion, it gets converted to phosphine gas in the body, which is subsequently absorbed into the bloodstream through the stomach and the intestines and gets captured by the liver and the lungs. Phosphine gas produces various metabolic and nonmetabolic toxic effects. Clinical symptoms are circulatory collapse, hypotension, shock symptoms, myocarditis, pericarditis, acute pulmonary edema, and congestive heart failure. In this case presentation, we aim to present the intensive care process and treatment resistance of a patient who ingested zinc phosphide for suicide purposes.

  11. Environmental risk limits for zinc

    NARCIS (Netherlands)

    Bodar CWM; SEC

    2007-01-01

    Environmental Riks Limits (ERLs) were derived for zinc. ERLs serve as advisory values to set environmental quality standards in the Netherlands. The ERLs for zinc closely follow the outcomes of earlier discussions on zinc within the Water Framework Directive and EC Regulation 793/93. The ERLs refer

  12. Environmental risk limits for zinc

    NARCIS (Netherlands)

    Bodar CWM; SEC

    2007-01-01

    Environmental Riks Limits (ERLs) were derived for zinc. ERLs serve as advisory values to set environmental quality standards in the Netherlands. The ERLs for zinc closely follow the outcomes of earlier discussions on zinc within the Water Framework Directive and EC Regulation 793/93. The ERLs ref

  13. Finger Length Ratios in Serbian Transsexuals

    Science.gov (United States)

    Vujović, Svetlana; Popović, Srdjan; Mrvošević Marojević, Ljiljana; Ivović, Miomira; Tančić-Gajić, Milina; Stojanović, Miloš; Marina, Ljiljana V.; Barać, Marija; Barać, Branko; Kovačević, Milena; Duišin, Dragana; Barišić, Jasmina; Djordjević, Miroslav L.; Micić, Dragan

    2014-01-01

    Atypical prenatal hormone exposure could be a factor in the development of transsexualism. There is evidence that the 2nd and 4th digit ratio (2D : 4D) associates negatively with prenatal testosterone and positively with estrogens. The aim was to assess the difference in 2D : 4D between female to male transsexuals (FMT) and male to female transsexuals (MFT) and controls. We examined 42 MFT, 38 FMT, and 45 control males and 48 control females. Precise measurements were made by X-rays at the ventral surface of both hands from the basal crease of the digit to the tip using vernier calliper. Control male and female patients had larger 2D : 4D of the right hand when compared to the left hand. Control male's left hand ratio was lower than in control female's left hand. There was no difference in 2D : 4D between MFT and control males. MFT showed similar 2D : 4D of the right hand with control women indicating possible influencing factor in embryogenesis and consequently finger length changes. FMT showed the lowest 2D : 4D of the left hand when compared to the control males and females. Results of our study go in favour of the biological aetiology of transsexualism. PMID:24982993

  14. Finger Length Ratios in Serbian Transsexuals

    Directory of Open Access Journals (Sweden)

    Svetlana Vujović

    2014-01-01

    Full Text Available Atypical prenatal hormone exposure could be a factor in the development of transsexualism. There is evidence that the 2nd and 4th digit ratio (2D : 4D associates negatively with prenatal testosterone and positively with estrogens. The aim was to assess the difference in 2D : 4D between female to male transsexuals (FMT and male to female transsexuals (MFT and controls. We examined 42 MFT, 38 FMT, and 45 control males and 48 control females. Precise measurements were made by X-rays at the ventral surface of both hands from the basal crease of the digit to the tip using vernier calliper. Control male and female patients had larger 2D : 4D of the right hand when compared to the left hand. Control male’s left hand ratio was lower than in control female’s left hand. There was no difference in 2D : 4D between MFT and control males. MFT showed similar 2D : 4D of the right hand with control women indicating possible influencing factor in embryogenesis and consequently finger length changes. FMT showed the lowest 2D : 4D of the left hand when compared to the control males and females. Results of our study go in favour of the biological aetiology of transsexualism.

  15. Radial fingering at an active interface

    Science.gov (United States)

    Nagilla, Amarender; Prabhakar, Ranganathan; Jadhav, Sameer

    2016-11-01

    It has been suggested that the shapes of single cells crawling on surfaces and those of the fronts of thin layers of cells collectively expanding to close a wound are the results of fingering instabilities. Motivated by these studies, we investigate the conditions under which an actively forced interface between a pair of immiscible viscous fluids will destabilize under Hele-Shaw confinement. The case of a circular active interface with surface tension and bending resistance is considered. Active forces exerted by the inner fluid at the interfacial region can be either completely internal or due to interactions with the confining substrate. In addition, the effects of cell growth or actin depolymerization or external injection of cell suspensions are modeled by including a distributed source and a point source of arbitrary strengths. Linear stability analysis reveals that at any given mean radius of the interface, its stability is dictated by two key dimensionless parameters. We discuss the different regions in a state space of these parameters.

  16. Examiner's finger-mounted fetal tissue oximetry

    Science.gov (United States)

    Kanayama, Naohiro; Niwayama, Masatsugu

    2014-06-01

    The best way to assess fetal condition is to observe the oxygen status of the fetus (as well as to assess the condition of infants, children, and adults). Previously, several fetal oximeters have been developed; however, no instrument has been utilized in clinical practice because of the low-capturing rate of the fetal oxygen saturation. To overcome the problem, we developed a doctor's finger-mounted fetal tissue oximeter, whose sensor volume is one hundredth of the conventional one. Additionally, we prepared transparent gloves. The calculation algorithm of the hemoglobin concentration was derived from the light propagation analysis based on the transport theory. We measured neonatal and fetal oxygen saturation (StO2) with the new tissue oximeter. Neonatal StO was measured at any position of the head regardless of amount of hair. Neonatal StO was found to be around 77%. Fetal StO was detected in every position of the fetal head during labor regardless of the presence of labor pain. Fetal StO without labor pain was around 70% in the first stage of labor and around 60% in the second stage of labor. We concluded that our new concept of fetal tissue oximetry would be useful for detecting fetal StO in any condition of the fetus.

  17. Numerically determined transport laws for fingering ("thermohaline") convection in astrophysics

    CERN Document Server

    Traxler, Adrienne; Stellmach, Stephan

    2010-01-01

    We present the first three-dimensional simulations of fingering convection performed in a parameter regime close to the one relevant for astrophysics, and reveal the existence of simple asymptotic scaling laws for turbulent heat and compositional transport. These laws can straightforwardly be extrapolated to the true astrophysical regime. Our investigation also indicates that thermocompositional "staircases," a key consequence of fingering convection in the ocean, cannot form spontaneously in stellar interiors. Our proposed empirically-determined transport laws thus provide simple prescriptions for mixing by fingering convection in a variety of astrophysical situations, and should, from here on, be used preferentially over older and less accurate parameterizations. They also establish that fingering convection does not provide sufficient extra mixing to explain observed chemical abundances in RGB stars.

  18. [Raynaud's phenomenon and other circulatory disorders of the fingers].

    Science.gov (United States)

    Mahler, Felix

    2014-02-26

    Raynaud's phenomenon (RP) is defined as attacks of blanking, subsequent cyanosis and rubeosis of fingers due to vasospasms in response to cold or emotional stimuli. Primary RP has no known underlying cause and occurs mainly in young and otherwise healthy women. Secondary RP goes along with various causes such as connective tissue diseases, toxic substances, drugs, physical trauma or organic finger artery occlusions, and occurs at any age and in both genders. Related affections are acrocyanosis and finger artery occlusions either due to arteriosclerosis or vasculitis. Also spontaneous finger hematoma may provoke an episode of RP. Therapeutically strict cold protection and avoidance of possible noxa is recommended besides the treatment of underlying diseases. No standard vasoactive drug has proven ideal for RP due to side effects. In cases with rest pain or ulcerations the same principles are applied as in ischemic diseases with no possibility for revascularization.

  19. Online Credit Card Transaction Using Finger Print Recognition

    Directory of Open Access Journals (Sweden)

    S.Sivasubramanian

    2010-10-01

    Full Text Available Internet shopping, a strong alternative to traditional “go, see, touch and buy” shopping, has been one of the mostly used facilities of the Internet. Security in online payment systems has been a wide research area since the early days of the Internet and several approaches have been devised by various Organizations. But, none of the system overcome the weakness in those system. Several online shopping systems serve internet users all around the world and enable people to get the products they need with a small effort. This paper proposes a new solution that combines finger print recognition with online credit card transactions. Here the proposed system provides more security then existing system with finger print recognitionbecause of finger print is unique. Here no need to remember the more passwords, your finger is your password.

  20. Tension Distribution in a Tendon-Driven Robotic Finger

    Science.gov (United States)

    Abdallah, Muhammad E. (Inventor); Platt, Robert (Inventor); Wampler, II, Charles W. (Inventor)

    2013-01-01

    A method is provided for distributing tension among tendons of a tendon-driven finger in a robotic system, wherein the finger characterized by n degrees of freedom and n+1 tendons. The method includes determining a maximum functional tension and a minimum functional tension of each tendon of the finger, and then using a controller to distribute tension among the tendons, such that each tendon is assigned a tension value less than the maximum functional tension and greater than or equal to the minimum functional tension. The method satisfies the minimum functional tension while minimizing the internal tension in the robotic system, and satisfies the maximum functional tension without introducing a coupled disturbance to the joint torques. A robotic system includes a robot having at least one tendon-driven finger characterized by n degrees of freedom and n+1 tendons, and a controller having an algorithm for controlling the tendons as set forth above.

  1. Repeatability Evaluation of Finger Tapping Device with Magnetic Sensors

    Science.gov (United States)

    Sano, Yuko; Kandori, Akihiko; Shima, Keisuke; Tamura, Yasuhiro; Takagi, Hiroshi; Tsuji, Toshio; Noda, Masafumi; Higashikawa, Fumiko; Yokoe, Masaru; Sakoda, Saburo

    We tested the repeatability of a finger tapping device with magnetic sensors to determine its reliability. This device, which was developed to assist in the diagnosis of movement disorders such as Parkinson's disease (PD) and strokes, measures the distance between the first and index fingers during finger tapping movements (opening and closing the fingers repeatedly). We evaluated three types of repeatability based on ICC (interclass correlation coefficient) and Welch's test (test for equal means in a oneway layout): repeatability when measured at different times, when using different devices, and when using different measurers. We calculated these three types for three finger tapping tasks on both hands for 21 characteristics calculated from finger tapping waveforms. Results demonstrated that the repeatability when using different devices is high regardless of the task or hand. The repeatability when measuring at different times and when using different measurers is high at some tasks, but not all. One of the finger tapping tasks (finger tapping movement with the largest amplitude and highest velocity), which is used in a conventional PD diagnosis method (UPDRS), does not have enough repeatability, while other tasks show high repeatability. Results also showed that five characteristics have the highest repeatability (ICC ≥ 0.5 or significance probability of Welch's test ≥ 5% in all tasks): “total moving distance,” “average of local minimum acceleration in opening motion,” “average of local minimum acceleration in closing motion,” “average of local maximum distance” and “average of local minimum velocity”. These results clearly demonstrate the strong repeatability of this device and lead to more precise diagnosis of movement disorders.

  2. Finger blood content, light transmission, and pulse oximetry errors.

    Science.gov (United States)

    Craft, T M; Lawson, R A; Young, J D

    1992-01-01

    The changes in light emitting diode current necessary to maintain a constant level of light incident upon a photodetector were measured in 20 volunteers at the two wavelengths employed by pulse oximeters. Three states of finger blood content were assessed; exsanguinated, hyperaemic, and normal. The changes in light emitting diode current with changes in finger blood content were small and are not thought to represent a significant source of error in saturation as measured by pulse oximetry.

  3. Structural insight into histone recognition by the ING PHD fingers.

    Science.gov (United States)

    Champagne, Karen S; Kutateladze, Tatiana G

    2009-05-01

    The Inhibitor of Growth (ING) tumor suppressors are implicated in oncogenesis, control of DNA damage repair, cellular senescence and apoptosis. All members of the ING family contain unique amino-terminal regions and a carboxy-terminal plant homeodomain (PHD) finger. While the amino-terminal domains associate with a number of protein effectors including distinct components of histone deacetylase (HDAC) and histone acetyltransferase (HAT) complexes, the PHD finger binds strongly and specifically to histone H3 trimethylated at lysine 4 (H3K4me3). In this review we describe the molecular mechanism of H3K4me3 recognition by the ING1-5 PHD fingers, analyze the determinants of the histone specificity and compare the biological activities and structures within subsets of PHD fingers. The atomic-resolution structures of the ING PHD fingers in complex with a H3K4me3 peptide reveal that the histone tail is bound in a large and deep binding site encompassing nearly one-third of the protein surface. An extensive network of intermolecular hydrogen bonds, hydrophobic and cation-pi contacts, and complementary surface interactions coordinate the first six residues of the H3K4me3 peptide. The trimethylated Lys4 occupies an elongated groove, formed by the highly conserved aromatic and hydrophobic residues of the PHD finger, whereas the adjacent groove accommodates Arg2. The two grooves are connected by a narrow channel, the small size of which defines the PHD finger's specificity, excluding interactions with other modified histone peptides. Binding of the ING PHD fingers to H3K4me3 plays a critical role in regulating chromatin acetylation. The ING proteins function as tethering molecules that physically link the HDAC and HAT enzymatic complexes to chromatin. In this review we also highlight progress recently made in understanding the molecular basis underlying biological and tumorigenic activities of the ING tumor suppressors.

  4. A Parametric Modelling Method for Dexterous Finger Reachable Workspaces

    OpenAIRE

    2016-01-01

    The well-known algorithms, such as the graphic method, analytical method or numerical method, have some defects when modelling the dexterous finger workspace, which is a significant kinematical feature of dexterous hands and valuable for grasp planning, motion control and mechanical design. A novel modelling method with convenient and parametric performances is introduced to generate the dexterous-finger reachable workspace. This method constructs the geometric topology of the dexterous-finge...

  5. Variable and Asymmetric Range of Enslaving: Fingers Can Act Independently over Small Range of Flexion

    NARCIS (Netherlands)

    Noort, J.C. van den; Beek, N. van; Kraan, T. van der; Veeger, D.H.; Stegeman, D.F.; Veltink, P.H.; Maas, H.

    2016-01-01

    The variability in the numerous tasks in which we use our hands is very large. However, independent movement control of individual fingers is limited. To assess the extent of finger independency during full-range finger flexion including all finger joints, we studied enslaving (movement in non-instr

  6. Variable and asymmetric range of enslaving: fingers can act independently over small range of flexion

    NARCIS (Netherlands)

    Noort, van den Josien C.; Beek, van Nathalie; Kraan, van der Thomas; Veeger, DirkJan H.E.J.; Stegeman, Dick F.; Veltink, Peter H.; Maas, Huub; Cymbalyuk, G.

    2016-01-01

    The variability in the numerous tasks in which we use our hands is very large. However, independent movement control of individual fingers is limited. To assess the extent of finger independency during full-range finger flexion including all finger joints, we studied enslaving movement in non-instru

  7. Doped zinc oxide microspheres

    Science.gov (United States)

    Arnold, Jr., Wesley D.; Bond, Walter D.; Lauf, Robert J.

    1993-01-01

    A new composition and method of making same for a doped zinc oxide microsphere and articles made therefrom for use in an electrical surge arrestor which has increased solid content, uniform grain size and is in the form of a gel.

  8. Creep Resistant Zinc Alloy

    Energy Technology Data Exchange (ETDEWEB)

    Frank E. Goodwin

    2002-12-31

    This report covers the development of Hot Chamber Die Castable Zinc Alloys with High Creep Strengths. This project commenced in 2000, with the primary objective of developing a hot chamber zinc die-casting alloy, capable of satisfactory service at 140 C. The core objectives of the development program were to: (1) fill in missing alloy data areas and develop a more complete empirical model of the influence of alloy composition on creep strength and other selected properties, and (2) based on the results from this model, examine promising alloy composition areas, for further development and for meeting the property combination targets, with the view to designing an optimized alloy composition. The target properties identified by ILZRO for an improved creep resistant zinc die-casting alloy were identified as follows: (1) temperature capability of 1470 C; (2) creep stress of 31 MPa (4500 psi); (3) exposure time of 1000 hours; and (4) maximum creep elongation under these conditions of 1%. The project was broadly divided into three tasks: (1) Task 1--General and Modeling, covering Experimental design of a first batch of alloys, alloy preparation and characterization. (2) Task 2--Refinement and Optimization, covering Experimental design of a second batch of alloys. (3) Task 3--Creep Testing and Technology transfer, covering the finalization of testing and the transfer of technology to the Zinc industry should have at least one improved alloy result from this work.

  9. Zinc in Multiple Sclerosis

    DEFF Research Database (Denmark)

    Bredholt, Mikkel; Frederiksen, Jette Lautrup

    2016-01-01

    In the last 35 years, zinc (Zn) has been examined for its potential role in the disease multiple sclerosis (MS). This review gives an overview of the possible role of Zn in the pathogenesis of MS as well as a meta-analysis of studies having measured Zn in serum or plasma in patients with MS...

  10. A Two-Axis Goniometric Sensor for Tracking Finger Motion.

    Science.gov (United States)

    Wang, Lefan; Meydan, Turgut; Williams, Paul Ieuan

    2017-04-05

    The study of finger kinematics has developed into an important research area. Various hand tracking systems are currently available; however, they all have limited functionality. Generally, the most commonly adopted sensors are limited to measurements with one degree of freedom, i.e., flexion/extension of fingers. More advanced measurements including finger abduction, adduction, and circumduction are much more difficult to achieve. To overcome these limitations, we propose a two-axis 3D printed optical sensor with a compact configuration for tracking finger motion. Based on Malus' law, this sensor detects the angular changes by analyzing the attenuation of light transmitted through polarizing film. The sensor consists of two orthogonal axes each containing two pathways. The two readings from each axis are fused using a weighted average approach, enabling a measurement range up to 180 ∘ and an improvement in sensitivity. The sensor demonstrates high accuracy (±0.3 ∘ ), high repeatability, and low hysteresis error. Attaching the sensor to the index finger's metacarpophalangeal joint, real-time movements consisting of flexion/extension, abduction/adduction and circumduction have been successfully recorded. The proposed two-axis sensor has demonstrated its capability for measuring finger movements with two degrees of freedom and can be potentially used to monitor other types of body motion.

  11. Biomechanical Analysis of Force Distribution in Human Finger Extensor Mechanisms

    Directory of Open Access Journals (Sweden)

    Dan Hu

    2014-01-01

    Full Text Available The complexities of the function and structure of human fingers have long been recognised. The in vivo forces in the human finger tendon network during different activities are critical information for clinical diagnosis, surgical treatment, prosthetic finger design, and biomimetic hand development. In this study, we propose a novel method for in vivo force estimation for the finger tendon network by combining a three-dimensional motion analysis technique and a novel biomechanical tendon network model. The extensor mechanism of a human index finger is represented by an interconnected tendinous network moving around the phalanx’s dorsum. A novel analytical approach based on the “Principle of Minimum Total Potential Energy” is used to calculate the forces and deformations throughout the tendon network of the extensor mechanism when subjected to an external load and with the finger posture defined by measurement data. The predicted deformations and forces in the tendon network are in broad agreement with the results obtained by previous experimental in vitro studies. The proposed methodology provides a promising tool for investigating the biomechanical function of complex interconnected tendon networks in vivo.

  12. Elastic fingering in rotating Hele-Shaw flows

    KAUST Repository

    Carvalho, Gabriel D.

    2014-05-21

    The centrifugally driven viscous fingering problem arises when two immiscible fluids of different densities flow in a rotating Hele-Shaw cell. In this conventional setting an interplay between capillary and centrifugal forces makes the fluid-fluid interface unstable, leading to the formation of fingered structures that compete dynamically and reach different lengths. In this context, it is known that finger competition is very sensitive to changes in the viscosity contrast between the fluids. We study a variant of such a rotating flow problem where the fluids react and produce a gellike phase at their separating boundary. This interface is assumed to be elastic, presenting a curvature-dependent bending rigidity. A perturbative weakly nonlinear approach is used to investigate how the elastic nature of the interface affects finger competition events. Our results unveil a very different dynamic scenario, in which finger length variability is not regulated by the viscosity contrast, but rather determined by two controlling quantities: a characteristic radius and a rigidity fraction parameter. By properly tuning these quantities one can describe a whole range of finger competition behaviors even if the viscosity contrast is kept unchanged. © 2014 American Physical Society.

  13. Integrating optical finger motion tracking with surface touch events

    Directory of Open Access Journals (Sweden)

    Jennifer eMacRitchie

    2015-06-01

    Full Text Available This paper presents a method of integrating two contrasting sensor systems for studying human interaction with a mechanical system, using piano performance as the case study. Piano technique requires both precise small-scale motion of fingers on the key surfaces and planned large-scale movement of the hands and arms. Where studies of performance often focus on one of these scales in isolation, this paper investigates the relationship between them. Two sensor systems were installed on an acoustic grand piano: a monocular high-speed camera tracking the position of painted markers on the hands, and capacitive touch sensors attach to the key surfaces which measure the location of finger-key contacts. This paper highlights a method of fusing the data from these systems, including temporal and spatial alignment, segmentation into notes and automatic fingering annotation. Three case studies demonstrate the utility of the multi-sensor data: analysis of finger flexion or extension based on touch and camera marker location, timing analysis of finger-key contact preceding and following key presses, and characterisation of individual finger movements in the transitions between successive key presses. Piano performance is the focus of this paper, but the sensor method could equally apply to other fine motor control scenarios, with applications to human-computer interaction.

  14. Studies on nanocrystalline zinc coating

    Indian Academy of Sciences (India)

    H B Muralidhara; Y Arthoba Naik

    2008-08-01

    Nano zinc coatings were deposited on mild steel by electrodeposition. The effect of additive on the morphology of crystal size on zinc deposit surface and corrosion properties were investigated. Corrosion tests were performed for dull zinc deposits and bright zinc deposits in aqueous NaCl solution (3.5 wt.%) using electrochemical measurements. The results showed that addition of additive in the deposition process of zinc significantly increased the corrosion resistance. The surface morphology of the zinc deposits was studied by scanning electron microscopy (SEM). The preferred orientation and average size of the zinc electrodeposited particles were obtained by X-ray diffraction analysis. The particles size was also characterized by TEM analysis.

  15. Automated FingerPrint Background removal: FPB

    Directory of Open Access Journals (Sweden)

    Morgante Michele

    2009-04-01

    Full Text Available Abstract Background The construction of a whole-genome physical map has been an essential component of numerous genome projects initiated since the inception of the Human Genome Project. Its usefulness has been proved for whole-genome shotgun projects as a post-assembly validation and recently it has also been used in the assembly step to constrain on BACs positions. Fingerprinting is usually the method of choice for construction of physical maps. A clone fingerprint is composed of true peaks representing real fragments and background peaks, mainly composed of E. coli genomic DNA, partial digestions, star activity by-products, and machine background. High-throughput fingerprinting leads to the production of thousands of BAC clone fingerprints per day. That is why background peaks removal has become an important issue and needs to be automatized, especially in capillary electrophoresis based fingerprints. Results At the moment, the only tools available for such a task are GenoProfiler and its descendant FPMiner. The large variation in the quality of fingerprints that is usually present in large fingerprinting projects represents a major difficulty in the correct removal of background peaks that has only been partially addressed by the methods so far adopted that all require a long manual optimization of parameters. Thus, we implemented a new data-independent tool, FPB (FingerPrint Background removal, suitable for large scale projects as well as mapping of few clones. Conclusion FPB is freely available at http://www.appliedgenomics.org/tools.php. FPB was used to remove the background from all fingerprints of three grapevine physical map projects. The first project consists of about 50,000 fingerprints, the second one consists of about 70,000 fingerprints, and the third one consists of about 45,000 fingerprints. In all cases a successful assembly was built.

  16. Child labour. Refuting the "nimble fingers" argument.

    Science.gov (United States)

    1996-01-01

    According to an International Labor Organization (ILO) study, approximately 130,000 children work in India's hand-knotted carpet industry. In one-loom enterprises, children comprise 14% of all weavers; in businesses with five or more looms, this rate increases to 33%. India's Factories Act, which applies costly health, safety, and labor regulations to larger firms, has led to a proliferation of cottage industries. The finding that children are more likely to work on low-quality rather than highest-quality carpets refutes the "nimble fingers" argument used by apologists of child labor. Although child and adult weavers have similar productivity, children earn less while apprentices than trained weavers and serve to depress wages throughout the industry. According to ILO estimates, replacing the 22% of the work force currently occupied by children with adults would cause wages to rise by about 5%. The overall savings in production costs from the use of child labor are very small when compared to the foreign retail price of the carpets, which is often four times the Indian export price. The ILO has urged an international approach to the elimination of child labor, in which all carpet-producing countries simultaneously implement a no-child-labor strategy to avoid placing any one country at a competitive disadvantage. Given the thousands of cottages where one or two carpets are woven per year, strategies such as labelling and regulation are likely to be ineffective. Solutions that address the general problems of poverty, while developing alternative sources of education and employment, are most likely to be effective in reducing child labor in countries such as India.

  17. Thermal stability improvement of a multiple finger power SiGe heterojunction bipolar transistor under different power dissipations using non-uniform finger spacing

    Institute of Scientific and Technical Information of China (English)

    Chen Liang; Zhang Wan-Rong; Jin Dong-Yue; Shen Pei; Xie Hong-Yun; Ding Chun-Bao; Xiaa Ying; Sun Bo-Tao; Wang Ren-Qing

    2011-01-01

    method of non-uniform finger spacing is proposed to enhance thermal stability of a multiple finger power SiGe hererojunction bipolar transistor under different power dissipations. Temperature distribution on the emitter fingers of a multi-finger SiGe heterojunction bipolar transistor is studied using a numerical electro-thermal model. The results show that the SiGe heterojunction bipolar transistor with non-uniform finger spacing has a small temperature difference between fingers compared with a traditional uniform finger spacing heterojunction bipolar transistor at the same power dissipation. What is most important is that the ability to improve temperature non-uniformity is not weakened as power dissipation increases. So the method of non-uniform finger spacing is very effective in enhancing the thermal stability and the power handing capability of power device. Experimental results verify our conclusious.

  18. Numerical simulation of two-dimensional salt fingers

    Science.gov (United States)

    Shen, Colin Y.; Veronis, George

    1997-10-01

    Numerical calculations of unperturbed, regularly spaced fingers in the heat-salt system (with a ratio of salt to heat diffusivities of 1/80) were carried out for a configuration in which a reservoir of uniformly salty, warm fluid lies initially above a reservoir of fresh, cold fluid. Cases were calculated in which the stability ratio, Rρ, was 1.5 and 3.0, and they were calculated for different magnitudes of the destabilizing salt increment, ΔS, expressed in terms of a salt Rayleigh number, Rs. Blobs of fluid with a salt anomaly accumulate at the ends of the evolving fingers. The magnitude and size of the anomaly increase with decreasing Rρ and increasing Rs. The density of those blobs is gravitationally unstable to perturbations. In the range of parameters used in these calculations the ratio of the flux of density due to heat to that due to salt varies from 0.17 to 0.74 for the unperturbed fingers. Essentially, the flux ratio decreases when the vertical velocity in the fingers is small, so that a relatively large amount of heat is diffused laterally from warm, salty descending fingers to cool, fresh ascending ones. A detailed account of the evolution of the perturbed system describes the various stages of the instability, concluding with the formation of larger structures in the reservoirs, which squash the fingers near the interface, so that isotherms and isohaline contours at midlevel are more or less horizontal. There is an indication of three period doublings in the spacing of the unstable blobs as they penetrate into the lower reservoir. The destruction of the regular array of upright, uniformly spaced fingers appears to be the natural evolution of perturbed systems in which Rρ is near unity and Rs is large.

  19. Making fingers and words count in a cognitive robot.

    Science.gov (United States)

    De La Cruz, Vivian M; Di Nuovo, Alessandro; Di Nuovo, Santo; Cangelosi, Angelo

    2014-01-01

    Evidence from developmental as well as neuroscientific studies suggest that finger counting activity plays an important role in the acquisition of numerical skills in children. It has been claimed that this skill helps in building motor-based representations of number that continue to influence number processing well into adulthood, facilitating the emergence of number concepts from sensorimotor experience through a bottom-up process. The act of counting also involves the acquisition and use of a verbal number system of which number words are the basic building blocks. Using a Cognitive Developmental Robotics paradigm we present results of a modeling experiment on whether finger counting and the association of number words (or tags) to fingers, could serve to bootstrap the representation of number in a cognitive robot, enabling it to perform basic numerical operations such as addition. The cognitive architecture of the robot is based on artificial neural networks, which enable the robot to learn both sensorimotor skills (finger counting) and linguistic skills (using number words). The results obtained in our experiments show that learning the number words in sequence along with finger configurations helps the fast building of the initial representation of number in the robot. Number knowledge, is instead, not as efficiently developed when number words are learned out of sequence without finger counting. Furthermore, the internal representations of the finger configurations themselves, developed by the