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Sample records for 3prime untranslated region

  1. The myotonic dystrophy kinase 3{prime}-untranslated region and its effect on gene expression

    SciTech Connect

    Ang, C.W.Y.; Sabourin, L.A.; Narang, M.A.

    1994-09-01

    Myotonic dystrophy (DM) is an autosomal dominant neuromuscular disease involving the expansion of an unstable CTG repeat in the 3{prime}-untranslated (3{prime}-UTR) region of the DM kinase (DMK) gene. Increased levels of mRNA in congenital compared to normal tissue have been shown, suggesting elevated DMK levels may be responsible for the disease phenotype. To study the effect of the DMK 3{prime}UTR on gene expression, a reporter gene system was constructed using the constitutive CMV promoter with the chloramphenicol acetyl transferase (CAT) open reading frame and the DMK 3{prime}UTR containing from 5 repeats up to 90 repeats. Transient transfection into a rhabdomyosarcoma cell line shows a three-fold increase in CAT activity from constructs containing a wildtype 3{prime}UTR (5 and 20 repeats) compared to a control construct containing only a poly(A) signal. Reporter constructs with repeats in the protomutation (50 repeats) and mutation (90 repeats) range show a greater than 10-fold increase over control CAT activity. These results suggest the presence of elements in the DMK 3{prime}UTR capable of conferring increased gene expression. We are currently investigating cell-specific activity of the constructs and conducting deletion mapping to identify regulatory elements in the 3{prime}-UTR.

  2. An AU-rich element in the 3{prime} untranslated region of the spinach chloroplast petD gene participates in sequence-specific RNA-protein complex formation

    SciTech Connect

    Chen, Qiuyun; Adams, C.C.; Usack, L.

    1995-04-01

    In chloroplasts, the 3{prime} untranslated regions of most mRNAs contain a stem-loop-forming inverted repeat (IR) sequence that is required for mRNA stability and correct 3{prime}-end formation. The IR regions of several mRNAs are also known to bind chloroplast proteins, as judged from in vitro gel mobility shift and UV cross-linking assays, and these RNA-protein interactions may be involved in the regulation of chloroplast mRNA processing and/or stability. Here we describe in detail the RNA and protein components that are involved in 3{prime} IR-containing RNA (3{prime} IR-RNA)-protein complex formation for the spinach chloroplast petD gene, which encodes subunit IV of the cytochrome b{sub 6}/f complex. We show that the complex contains 55-, 41-, and 29-kDa RNA-binding proteins (ribonucleoproteins [RNPs]). These proteins together protect a 90-nucleotide segment of RNA from RNase T{sub 1} digestion; this RNA contains the IR and downstream flanking sequences. Competition experiments using 3{prime} IR-RNAs from the psbA or rbcL gene demonstrate that the RNPs have a strong specificity for the petD sequence. Site-directed mutagenesis was carried out to define the RNA sequence elements required for complex formation. These studies identified an 8-nucleotide AU-rich sequence downstream of the IR; mutations within this sequence had moderate to severe effects on RNA-protein complex formation. Although other similar sequences are present in the petD 3{prime} untranslated region, only a single copy, which we have termed box II, appears to be essential for in vivo protein binding. In addition, the IR itself is necessary for optimal complex formation. These two sequence elements together with an RNP complex may direct correct 3{prime}-end processing and/or influence the stability of petD mRNA in chloroplasts. 48 refs., 9 figs., 2 tabs.

  3. Untranslated regions (UTRs) orchestrate translation reprogramming in cellular stress responses.

    PubMed

    Sajjanar, Basavaraj; Deb, Rajib; Raina, Susheel Kumar; Pawar, Sachin; Brahmane, Manoj P; Nirmale, Avinash V; Kurade, Nitin P; Manjunathareddy, Gundallahalli B; Bal, Santanu Kumar; Singh, Narendra Pratap

    2017-04-01

    Stress is the result of an organism's interaction with environmental challenges. Regulations of gene expression including translation modulations are critical for adaptation and survival under stress. Untranslated regions (UTRs) of the transcripts play significant roles in translation regulation and continue to raise many intriguing questions in our understanding of cellular stress physiology. IRES (Internal ribosome entry site) and uORF (upstream open reading frame) mediated alternative translation initiations are emerging as unique mechanisms. Recent studies have revealed novel means of mRNAs stabilization in stress granules and their reversible modifications. Differential regulation of select transcripts is possible by the interplay between the adenine/uridine-rich elements (AREs) in 3'UTR with their binding proteins (AUBP) and by microRNA-mediated effects. Coordination of these various mechanisms control translation and thereby enables appropriate responses to environmental stress. In this review, we focus on the role of sequence signatures both at 5' and 3'UTRs in translation reprogramming during cellular stress responses.

  4. Dom34 rescues ribosomes in 3' untranslated regions.

    PubMed

    Guydosh, Nicholas R; Green, Rachel

    2014-02-27

    Ribosomes that stall before completing peptide synthesis must be recycled and returned to the cytoplasmic pool. The protein Dom34 and cofactors Hbs1 and Rli1 can dissociate stalled ribosomes in vitro, but the identity of targets in the cell is unknown. Here, we extend ribosome profiling methodology to reveal a high-resolution molecular characterization of Dom34 function in vivo. Dom34 removes stalled ribosomes from truncated mRNAs, but, in contrast, does not generally dissociate ribosomes on coding sequences known to trigger stalling, such as polyproline. We also show that Dom34 targets arrested ribosomes near the ends of 3' UTRs. These ribosomes appear to gain access to the 3' UTR via a mechanism that does not require decoding of the mRNA. These results suggest that ribosomes frequently enter downstream noncoding regions and that Dom34 carries out the important task of rescuing them.

  5. Alternative 5' untranslated regions are involved in expression regulation of human heme oxygenase-1.

    PubMed

    Kramer, Marcel; Sponholz, Christoph; Slaba, Monique; Wissuwa, Bianka; Claus, Ralf A; Menzel, Uwe; Huse, Klaus; Platzer, Matthias; Bauer, Michael

    2013-01-01

    The single nucleotide polymorphism rs2071746 and a (GT)n microsatellite within the human gene encoding heme oxygenase-1 (HMOX1) are associated with incidence or outcome in a variety of diseases. Most of these associations involve either release of heme or oxidative stress. Both polymorphisms are localized in the promoter region, but previously reported correlations with heme oxygenase-1 expression remain not coherent. This ambiguity suggests a more complex organization of the 5' gene region which we sought to investigate more fully. We evaluated the 5' end of HMOX1 and found a novel first exon 1a placing the two previously reported polymorphisms in intronic or exonic positions within the 5' untranslated region respectively. Expression of exon 1a can be induced in HepG2 hepatoma cells by hemin and is a repressor of heme oxygenase-1 translation as shown by luciferase reporter assays. Moreover, minigene approaches revealed that the quantitative outcome of alternative splicing within the 5' untranslated region is affected by the (GT)n microsatellite. This data supporting an extended HMOX1 gene model and provide further insights into expression regulation of heme oxygenase-1. Alternative splicing within the HMOX1 5' untranslated region contributes to translational regulation and is a mechanistic feature involved in the interplay between genetic variations, heme oxygenase-1 expression and disease outcome.

  6. HLA-G coding region and 3'untranslated region (3'UTR) in two Chinese Han populations.

    PubMed

    Wang, Wen Yi; Tian, Wei; Liu, Xue Xiang; Li, Li Xin

    2016-08-01

    In this study, exons 2-4 and 3'untranslated region (3'UTR) of human leukocyte antigen (HLA)-G gene were investigated for 201 and 104 healthy unrelated Han samples recruited from Hunan Province, southern China and central Inner Mongolia Autonomous Region, northern China, respectively, using sequence-based typing and cloning methods. Totally 12 HLA-G alleles in the coding region, 9 variable sites in 3'UTR, 8 3'UTR haplotypes and 15 HLA-G extended haplotypes (EHs) incorporating the coding region and 3'UTR were observed. Very strong linkage disequilibrium (LD) was observed between HLA-A and HLA-G, and between HLA-G coding region and 3'UTR in each population (all global P=0.0000). Seven HLA-A-G haplotypes showed significant LD in both populations. Three HLA-G alleles in the coding region, 4 polymorphic sites in the 3'UTR, 3 3'UTR haplotypes and 4 HLA-G EHs differed significantly in their distributions between the 2 Chinese Han populations (all P≤0.0001). There was evidence for balancing selection acting on HLA-G 3'UTR positions +3010, +3142 and +3187 in the two populations. The NJ dendrograms demonstrated the existence of two basic HLA-G lineages and indicated that, HLA-G*01:01:01, the most common HLA-G allele, formed a separate lineage from other alleles. Our results shed new lights into HLA-G genetics among Chinese Han populations. The findings reported here are of importance for future studies related to post-transcriptional regulation of HLA-G allelic expression and the potential role of HLA-G in disease association in populations of Chinese ancestry.

  7. Two lck transcripts containing different 5' untranslated regions are present in T cells.

    PubMed Central

    Voronova, A F; Adler, H T; Sefton, B M

    1987-01-01

    p56lck is a new member of the src family of cellular tyrosine protein kinases. It is expressed constitutively at a low level in normal T cells and at an elevated level in the LSTRA and Thy19 Moloney murine leukemia virus-induced thymoma cell lines. It is possible that the expression of p56lck at an elevated level contributes to the transformation of these thymoma cells. The structure of the mRNAs encoding p56lck was examined by using an RNase protection assay. Both a chimeric lck mRNA containing the 5' untranslated region of Moloney virus mRNA and a normal lck mRNA were found in Thy19 and LSTRA cells. The chimeric lck transcript was 4- to 10-fold more abundant than the normal transcript. Transcription arising from a viral promoter is therefore responsible for the elevated levels of lck mRNA in these two cell lines. Surprisingly, uninfected murine T cells were also found to contain lck transcripts with differing 5' untranslated regions. One species of mRNA was colinear with the region of the chromosome just upstream of the initiation codon for p56lck. The other appeared to arise through splicing of an unidentified 5' untranslated exon to a sometimes cryptic splice acceptor just upstream of the region encoding p56lck. These data suggest that lck is expressed through the use of at least two different promoters. The promoters could be subject to different forms of regulation. Images PMID:3501824

  8. Sdt97: A Point Mutation in the 5′ Untranslated Region Confers Semidwarfism in Rice

    PubMed Central

    Tong, Jiping; Han, Zhengshu; Han, Aonan; Liu, Xuejun; Zhang, Shiyong; Fu, Binying; Hu, Jun; Su, Jingping; Li, Shaoqing; Wang, Shengjun; Zhu, Yingguo

    2016-01-01

    Semidwarfism is an important agronomic trait in rice breeding programs. The semidwarf mutant gene Sdt97 was previously described. However, the molecular mechanism underlying the mutant is yet to be elucidated. In this study, we identified the mutant gene by a map-based cloning method. Using a residual heterozygous line (RHL) population, Sdt97 was mapped to the long arm of chromosome 6 in the interval of nearly 60 kb between STS marker N6 and SNP marker N16 within the PAC clone P0453H04. Sequencing of the candidate genes in the target region revealed that a base transversion from G to C occurred in the 5′ untranslated region of Sdt97. qRT-PCR results confirmed that the transversion induced an obvious change in the expression pattern of Sdt97 at different growth and developmental stages. Plants transgenic for Sdt97 resulted in the restoration of semidwarfism of the mutant phenotype, or displayed a greater dwarf phenotype than the mutant. Our results indicate that a point mutation in the 5′ untranslated region of Sdt97 confers semidwarfism in rice. Functional analysis of Sdt97 will open a new field of study for rice semidwarfism, and also expand our knowledge of the molecular mechanism of semidwarfism in rice. PMID:27172200

  9. Tissue-specific expression of insulin-like growth factor II mRNAs with distinct 5' untranslated regions

    SciTech Connect

    Irminger, J.C.; Rosen, K.M.; Humble, R.E.; Villa-Komaroff, L.

    1987-09-01

    The authors have used RNA from human hypothalamus as template for the production of cDNAs encoding insulin-like growth factor II (IGF-II). The prohormone coding sequence of brain IGF-II RNA is identical to that found in liver; however, the 5' untranslated sequence of the brain cDNA has no homology to the 5' untranslated sequence of the previously reported liver cDNAs. By using hybridization to specific probes as well as a method based on the properties of RNase H, they found that the human IGF-II gene has at least three exons that encode alternative 5' untranslated regions and that are expressed in a tissue-specific manner. A probe specific to the brain cDNA 5' untranslated region hybridizes to a 6.0-kilobase transcript present in placenta, hypothalamus, adrenal gland, kidney, Wilms tumor, and a pheochromocytoma. The 5' untranslated sequence of the brain cDNA does not hybridize to a 5.3-kilobase transcript found in liver or to a 5.0-kb transcript found in pheochromocytoma. By using RNase H to specifically fragment the IGF-II transcripts into 3' and 5' fragments, they found that the RNAs vary in size due to differences in the 5' end but not the 3' end.

  10. Gene Expression in Archaea: Studies of Transcriptional Promoters, Messenger RNA Processing, and Five Prime Untranslated Regions in "Methanocaldococcus Jannashchii"

    ERIC Educational Resources Information Center

    Zhang, Jian

    2009-01-01

    Gene expression in Archaea is less understood than those in Bacteria and Eucarya. In general, three steps are involved in gene expression--transcription, RNA processing, and translation. To expand our knowledge of these processes in Archaea, I have studied transcriptional promoters, messenger RNA processing, and 5'-untranslated regions in…

  11. Functions of the 5'- and 3'-untranslated regions of tobamovirus RNA.

    PubMed

    Chujo, Tetsuya; Ishibashi, Kazuhiro; Miyashita, Shuhei; Ishikawa, Masayuki

    2015-08-03

    The tobamovirus genome is a 5'-m(7)G-capped RNA that carries a tRNA-like structure at its 3'-terminus. The genomic RNA serves as the template for both translation and negative-strand RNA synthesis. The 5'- and 3'-untranslated regions (UTRs) of the genomic RNA contain elements that enhance translation, and the 3'-UTR also contains the elements necessary for the initiation of negative-strand RNA synthesis. Recent studies using a cell-free viral RNA translation-replication system revealed that a 70-nucleotide region containing a part of the 5'-UTR is bound cotranslationally by tobacco mosaic virus (TMV) replication proteins translated from the genomic RNA and that the binding leads the genomic RNA to RNA replication pathway. This mechanism explains the cis-preferential replication of TMV by the replication proteins. The binding also inhibits further translation to avoid a fatal ribosome-RNA polymerase collision, which might arise if translation and negative-strand synthesis occur simultaneously on a single genomic RNA molecule. Therefore, the 5'- and 3'-UTRs play multiple important roles in the life cycle of tobamovirus.

  12. Cellular DDX3 regulates Japanese encephalitis virus replication by interacting with viral un-translated regions.

    PubMed

    Li, Chen; Ge, Ling-ling; Li, Peng-peng; Wang, Yue; Dai, Juan-juan; Sun, Ming-xia; Huang, Li; Shen, Zhi-qiang; Hu, Xiao-chun; Ishag, Hassan; Mao, Xiang

    2014-01-20

    Japanese encephalitis virus is one of the most common causes for epidemic viral encephalitis in humans and animals. Herein we demonstrated that cellular helicase DDX3 is involved in JEV replication. DDX3 knockdown inhibits JEV replication. The helicase activity of DDX3 is crucial for JEV replication. GST-pulldown and co-immunoprecipitation experiments demonstrated that DDX3 could interact with JEV non-structural proteins 3 and 5. Co-immunoprecipitation and confocal microscopy analysis confirmed that DDX3 interacts and colocalizes with these viral proteins and viral RNA during the infection. We determined that DDX3 binds to JEV 5' and 3' un-translated regions. We used a JEV-replicon system to demonstrate that DDX3 positively regulates viral RNA translation, which might affect viral RNA replication at the late stage of virus infection. Collectively, we identified that DDX3 is necessary for JEV infection, suggesting that DDX3 might be a novel target to design new antiviral agents against JEV or other flavivirus infections.

  13. The role of RNA structure at 5' untranslated region in microRNA-mediated gene regulation.

    PubMed

    Gu, Wanjun; Xu, Yuming; Xie, Xueying; Wang, Ting; Ko, Jae-Hong; Zhou, Tong

    2014-09-01

    Recent studies have suggested that the secondary structure of the 5' untranslated region (5' UTR) of messenger RNA (mRNA) is important for microRNA (miRNA)-mediated gene regulation in humans. mRNAs that are targeted by miRNA tend to have a higher degree of local secondary structure in their 5' UTR; however, the general role of the 5' UTR in miRNA-mediated gene regulation remains unknown. We systematically surveyed the secondary structure of 5' UTRs in both plant and animal species and found a universal trend of increased mRNA stability near the 5' cap in mRNAs that are regulated by miRNA in animals, but not in plants. Intra-genome comparison showed that gene expression level, GC content of the 5' UTR, number of miRNA target sites, and 5' UTR length may influence mRNA structure near the 5' cap. Our results suggest that the 5' UTR secondary structure performs multiple functions in regulating post-transcriptional processes. Although the local structure immediately upstream of the start codon is involved in translation initiation, RNA structure near the 5' cap site, rather than the structure of the full-length 5' UTR sequences, plays an important role in miRNA-mediated gene regulation.

  14. Global changes in processing of mRNA 3' untranslated regions characterize clinically distinct cancer subtypes.

    PubMed

    Singh, Priyam; Alley, Travis L; Wright, Sarah M; Kamdar, Sonya; Schott, William; Wilpan, Robert Y; Mills, Kevin D; Graber, Joel H

    2009-12-15

    Molecular cancer diagnostics are an important clinical advance in cancer management, but new methods are still needed. In this context, gene expression signatures obtained by microarray represent a useful molecular diagnostic. Here, we describe novel probe-level microarray analyses that reveal connections between mRNA processing and neoplasia in multiple tumor types, with diagnostic potential. We now show that characteristic differences in mRNA processing, primarily in the 3'-untranslated region, define molecular signatures that can distinguish similar tumor subtypes with different survival characteristics, with at least 74% accuracy. Using a mouse model of B-cell leukemia/lymphoma, we find that differences in transcript isoform abundance are likely due to both alternative polyadenylation (APA) and differential degradation. While truncation of the 3'-UTR is the most common observed pattern, genes with elongated transcripts were also observed, and distinct groups of affected genes are found in related but distinct tumor types. Genes with elongated transcripts are overrepresented in ontology categories related to cell-cell adhesion and morphology. Analysis of microarray data from human primary tumor samples revealed similar phenomena. Western blot analysis of selected proteins confirms that changes in the 3'-UTR can correlate with changes in protein expression. Our work suggests that alternative mRNA processing, particularly APA, can be a powerful molecular biomarker with prognostic potential. Finally, these findings provide insights into the molecular mechanisms of gene deregulation in tumorigenesis.

  15. Identification of short untranslated regions that sufficiently enhance translation in high-quality wheat germ extract.

    PubMed

    Ogawa, Atsushi; Tabuchi, Junichiro; Doi, Yasunori

    2014-08-15

    High-quality wheat germ extract (hqWGE) is very useful for the high-yield production of various types of protein. The most important key to high productivity is the design of mRNA templates. Although the design has been refined for straightforward and efficient translation in hqWGE, there is still room for improvement in untranslated regions (UTRs), especially the 3' UTR length, because a long, cumbersome 3' UTR is commonly used for translation enhancement. Here we examined some short viral 3' cap-independent translation enhancers (3' CITEs) to identify effective ones for efficient translation in hqWGE. We then combined the most effective 3' CITE and a 5' enhancer to further increase the translation efficiency. mRNA with the optimal short 3' and 5' UTRs, both of whose length was less than 150 nt, exhibited a productivity of 1.4 mg/mL in prolonged large-scale protein synthesis in hqWGE, which was comparable to that of control mRNA with a commonly-used long 3' UTR (∼1200 nt).

  16. Translational repression by a miniature inverted-repeat transposable element in the 3′ untranslated region

    PubMed Central

    Shen, Jianqiang; Liu, Juhong; Xie, Kabin; Xing, Feng; Xiong, Fang; Xiao, Jinghua; Li, Xianghua; Xiong, Lizhong

    2017-01-01

    Transposable elements constitute a substantial portion of eukaryotic genomes and contribute to genomic variation, function, and evolution. Miniature inverted-repeat transposable elements (MITEs), as DNA transposons, are widely distributed in plant and animal genomes. Previous studies have suggested that retrotransposons act as translational regulators; however, it remains unknown how host mRNAs are influenced by DNA transposons. Here we report a translational repression mechanism mediated by a stowaway-like MITE (sMITE) embedded in the 3′-untranslated region (3′-UTR) of Ghd2, a member of the CCT (CONSTANS [CO], CO-LIKE and TIMING OF CAB1) gene family in rice. Ghd2 regulates important agronomic traits, including grain number, plant height and heading date. Interestingly, the translational repression of Ghd2 by the sMITE mainly relies on Dicer-like 3a (OsDCL3a). Furthermore, other MITEs in the 3′-UTRs of different rice genes exhibit a similar effect on translational repression, thus suggesting that MITEs may exert a general regulatory function at the translational level. PMID:28256530

  17. Differential regulation of oestrogen receptor β isoforms by 5' untranslated regions in cancer.

    PubMed

    Smith, Laura; Brannan, Rebecca A; Hanby, Andrew M; Shaaban, Abeer M; Verghese, Eldo T; Peter, Mark B; Pollock, Steven; Satheesha, Sampoorna; Szynkiewicz, Marcin; Speirs, Valerie; Hughes, Thomas A

    2010-08-01

    Oestrogen receptors (ERs) are critical regulators of the behaviour of many cancers. Despite this, the roles and regulation of one of the two known ERs - ERβ- are poorly understood. This is partly because analyses have been confused by discrepancies between ERβ expression at mRNA and proteins levels, and because ERβ is expressed as several functionally distinct isoforms. We investigated human ERβ 5' untranslated regions (UTRs) and their influences on ERβ expression and function. We demonstrate that two alternative ERβ 5'UTRs have potent and differential influences on expression acting at the level of translation. We show that their influences are modulated by cellular context and in carcinogenesis, and demonstrate the contributions of both upstream open reading frames and RNA secondary structure. These regulatory mechanisms offer explanations for the non-concordance of ERβ mRNA and protein. Importantly, we also demonstrate that 5'UTRs allow the first reported mechanisms for differential regulation of the expression of the ERβ isoforms 1, 2 and 5, and thereby have critical influences on ERβ function.

  18. Skipping of an alternative intron in the srsf1 3' untranslated region increases transcript stability.

    PubMed

    Akaike, Yoko; Kurokawa, Ken; Kajita, Keisuke; Kuwano, Yuki; Masuda, Kiyoshi; Nishida, Kensei; Kang, Seung Wan; Tanahashi, Toshihito; Rokutan, Kazuhito

    2011-08-01

    The srsf1 gene encodes serine/arginine-rich splicing factor 1 (SRSF1) that participates in both constitutive and alternative splicing reactions. This gene possesses two ultraconserved elements in the 3' untranslated region (UTR). Skipping of an alternative intron between the two elements has no effect on the protein-coding sequence, but it generates a premature stop codon (PTC)-containing mRNA isoform, whose degradation is considered to depend on nonsense-mediated mRNA decay (NMD). However, several cell lines (HCT116, RKO, HeLa, and WI38 cells) constitutively expressed significant amounts of the srsf1 PTC variant. HCT116 cells expressed the PTC variant nearly equivalent to the major isoform that includes the alternative intron in the 3' UTR. Inhibition of NMD by silencing a key effecter UPF1 or by treatment with cycloheximide failed to increase amounts of the PTC variant in HCT116 cells, and the PTC variant was rather more stable than the major isoform in the presence of actinomycin D. Our results suggest that the original stop codon may escape from the NMD surveillance even in skipping of the alternative intron. The srsf1 gene may produce an alternative splice variant having truncated 3' UTR to relief the microRNA- and/or RNA-binding protein-mediated control of translation or degradation.

  19. Constitutive translation of human α-synuclein is mediated by the 5′-untranslated region

    PubMed Central

    Koukouraki, Pelagia; Doxakis, Epaminondas

    2016-01-01

    Genetic and biochemical studies have established a central role for α-synuclein (SNCA) accumulation in the pathogenesis of Parkinson's disease. Uncovering and subsequently interfering with physiological mechanisms that control SNCA expression is one approach to limit disease progression. To this end, the long and GC-rich 5′-untranslated region (UTR) of SNCA, which is predicted to fold into stable hairpin and G-quadruplex RNA motifs, was investigated for its role in mRNA translation. Inclusion of SNCA 5′-UTR significantly induced expression of both SNCA and luciferase ORF constructs. This effect was not associated with a change in mRNA levels or differential nucleocytoplasmic shuttling. Further, the presence of the 5′-UTR enhanced SNCA synthesis when cap-dependent translation was attenuated with rapamycin treatment. Analysis using multiple methodologies revealed that the 5′-UTR harbours an internal ribosome entry site (IRES) element that spans most of its nucleotide sequence. Signals such as plasma-membrane depolarization, serum starvation and oxidative stress stimulated SNCA protein translation via its 5′-UTR as well as enhanced its IRES activity. Taken together, these data support the idea that the 5′-UTR is an important positive regulator of SNCA synthesis under diverse physiological and pathological conditions, explaining in part the abundance of SNCA in healthy neurons and its accumulation in degenerative cells. PMID:27248657

  20. RNA-protein interactions of the 3' untranslated regions of myosin heavy chain transcripts.

    PubMed

    Kiri, Arpna; Goldspink, Geoffrey

    2002-01-01

    The RNA-protein interactions of the myosin heavy chain (MyHC) 3' untranslated regions (3'UTRs) were investigated using gel mobility shift assays. Marine skeletal myosin heavy chain mRNAs were amplified using reverse transcription coupled with the polymerase chain reaction (RT-PCR). Four cloned MyHC sequences were identified as slow type 1, fast 2a, fast 2b and fast 2x. The 3'UTRs of the four MyHC mRNAs were shown to interact with muscle protein in a tissue-specific manner as illustrated by gel retardation assays with protein extracts from various tissues. Competition assays indicate that this interaction is specific to the MyHC 3'UTR sequence. UV cross-linking suggests that several small proteins bind to the 3'UTR's. Peptide sequencing identified aldolase A and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as MyHC 3'UTR RNA-binding proteins. The implications of these interactions and post-transcriptional regulation of the MyHC genes are discussed.

  1. Irregular G-quadruplexes Found in the Untranslated Regions of Human mRNAs Influence Translation.

    PubMed

    Bolduc, François; Garant, Jean-Michel; Allard, Félix; Perreault, Jean-Pierre

    2016-10-07

    G-quadruplex structures are composed of coplanar guanines and are found in both DNA and RNA. They are formed by the stacking of two or more G-quartets that are linked together by three loops. The current belief is that RNA G-quadruplexes include loops of l to 7 nucleotides in length, although recent evidence indicates that the central loop (loop 2) can be longer if loops 1 and 3 are limited to a single nucleotide each. With the objective of broadening the definition of irregular RNA G-quadruplexes, a bioinformatic search was performed to find potential G-quadruplexes located in the untranslated regions of human mRNAs (i.e. in the 5' and 3'-UTRs) that contain either a long loop 1 or 3 of up to 40 nucleotides in length. RNA molecules including the potential sequences were then synthesized and examined in vitro by in-line probing for the formation of G-quadruplex structures. The sequences that adopted a G-quadruplex structure were cloned into a luciferase dual vector and examined for their ability to modulate translation in cellulo Some irregular G-quadruplexes were observed to either promote or repress translation regardless of the position or the size of the long loop they possessed. Even if the composition of a RNA G-quadruplex is not quite completely understood, the results presented in this report clearly demonstrate that what defines a RNA G-quadruplex is much broader than what we previously believed.

  2. Constitutive translation of human α-synuclein is mediated by the 5'-untranslated region.

    PubMed

    Koukouraki, Pelagia; Doxakis, Epaminondas

    2016-04-01

    Genetic and biochemical studies have established a central role for α-synuclein (SNCA) accumulation in the pathogenesis of Parkinson's disease. Uncovering and subsequently interfering with physiological mechanisms that control SNCA expression is one approach to limit disease progression. To this end, the long and GC-rich 5'-untranslated region (UTR) of SNCA, which is predicted to fold into stable hairpin and G-quadruplex RNA motifs, was investigated for its role in mRNA translation. Inclusion of SNCA 5'-UTR significantly induced expression of both SNCA and luciferase ORF constructs. This effect was not associated with a change in mRNA levels or differential nucleocytoplasmic shuttling. Further, the presence of the 5'-UTR enhanced SNCA synthesis when cap-dependent translation was attenuated with rapamycin treatment. Analysis using multiple methodologies revealed that the 5'-UTR harbours an internal ribosome entry site (IRES) element that spans most of its nucleotide sequence. Signals such as plasma-membrane depolarization, serum starvation and oxidative stress stimulated SNCA protein translation via its 5'-UTR as well as enhanced its IRES activity. Taken together, these data support the idea that the 5'-UTR is an important positive regulator of SNCA synthesis under diverse physiological and pathological conditions, explaining in part the abundance of SNCA in healthy neurons and its accumulation in degenerative cells.

  3. Numerical taxonomy of the genus Pestivirus based on palindromic nucleotide substitutions in the 5' untranslated region.

    PubMed

    Giangaspero, Massimo; Harasawa, Ryô

    2007-12-01

    The palindromic nucleotide substitutions (PNS) at the three variable loci (V1, V2 and V3) in the 5' untranslated region (UTR) of Pestivirus RNA have been considered for taxonomical segregation of species, through the evaluation of 430 genomic sequences. On the basis of qualitative and quantitative secondary structure characteristics, six species have been identified: Bovine viral diarrhea virus 1 (BVDV-1), Bovine viral diarrhea virus 2 (BVDV-2), Classical swine fever virus (CSFV), Border disease virus (BDV), the tentative species Giraffe and a new proposed taxon named Pronghorn. The first step was qualitative and consisted in the characterization of the different positions of the three stems and loops in the 5' UTR sequences of all the strains under consideration belonging to the genus. Secondary structure sequences showing divergent base-pair combinations have been aligned for comparison. Palindromic positions have been characterized according to changes in nucleotide base-pairs identifying low-variable positions (LVP) including base-pairs present in less than 80% of the genus. The second step was quantitative, allowing the identification of genomic groups by clustering the base-pair combinations according to LVP. Relatedness among types was evaluated to identify homogeneous groups. Cross comparisons between types within the genus have been evaluated by computing the divergence percentage thus clarifying borderline and multirelated sequences.

  4. Dynamic Regulation of Tandem 3' Untranslated Regions in Zebrafish Spleen Cells during Immune Response.

    PubMed

    Huang, Guangrui; Huang, Shengfeng; Wang, Ruihua; Yan, Xinyu; Li, Yuxin; Feng, Yuchao; Wang, Shaozhou; Yang, Xia; Chen, Liutao; Li, Jun; You, Leiming; Chen, Shangwu; Luo, Guangbin; Xu, Anlong

    2016-01-15

    Alternative polyadenylation (APA) has been found to be involved in tumorigenesis, development, and cell differentiation, as well as in the activation of several subsets of immune cells in vitro. Whether APA takes place in immune responses in vivo is largely unknown. We profiled the variation in tandem 3' untranslated regions (UTRs) in pathogen-challenged zebrafish and identified hundreds of APA genes with ∼ 10% being immune response genes. The detected immune response APA genes were enriched in TLR signaling, apoptosis, and JAK-STAT signaling pathways. A greater number of microRNA target sites and AU-rich elements were found in the extended 3' UTRs than in the common 3' UTRs of these APA genes. Further analysis suggested that microRNA and AU-rich element-mediated posttranscriptional regulation plays an important role in modulating the expression of APA genes. These results indicate that APA is extensively involved in immune responses in vivo, and it may be a potential new paradigm for immune regulation.

  5. Genome-wide profiling of untranslated regions by paired-end ditag sequencing reveals unexpected transcriptome complexity in yeast.

    PubMed

    Kang, Ya-Ni; Lai, Deng-Pan; Ooi, Hong Sain; Shen, Ting-Ting; Kou, Yao; Tian, Jing; Czajkowsky, Daniel M; Shao, Zhifeng; Zhao, Xiaodong

    2015-02-01

    The identification of structural and functional elements encoded in a genome is a challenging task. Although the transcriptome of budding yeast has been extensively analyzed, the boundaries and untranslated regions of yeast genes remain elusive. To address this least-explored field of yeast genomics, we performed a transcript profiling analysis through paired-end ditag (PET) approach coupled with deep sequencing. With 562,133 PET sequences we accurately defined the boundaries and untranslated regions of 3,409 ORFs, suggesting many yeast genes have multiple transcription start sites (TSSs). We also identified 85 previously uncharacterized transcripts either in intergenic regions or from the opposite strand of reported genomic features. Furthermore, our data revealed the extensive 3' end heterogeneity of yeast genes and identified a novel putative motif for polyadenylation. Our results indicate the yeast transcriptome is more complex than expected. This study would serve as an invaluable resource for elucidating the regulation and evolution of yeast genes.

  6. Association of HLA-G 3' untranslated region variants with type 1 diabetes mellitus.

    PubMed

    de Albuquerque, Rafael S; Mendes-Junior, Celso Teixeira; Lucena-Silva, Norma; da Silva, Camila Leal Lopes; Rassi, Diane Meire; Veiga-Castelli, Luciana C; Foss-Freitas, Maria Cristina; Foss, Milton César; Deghaide, Neifi Hassan Saloum; Moreau, Philippe; Gregori, Silvia; Castelli, Erick C; Donadi, Eduardo Antônio

    2016-04-01

    Besides the well recognized association of HLA-DRB1 and DQB1 alleles with type 1 diabetes mellitus (T1D), linkage studies have identified a gene region close to the non-classical class I HLA-G gene as an independent susceptibility marker. HLA-G is constitutively expressed in the endocrine compartment of the human pancreas and may play a role in controlling autoimmune responses. We evaluated the genetic diversity of the 3' untranslated region (3'UTR) of HLA-G, which have been associated with HLA-G mRNA post-transcriptional regulation, in 120 Brazilian T1D patients and in 120 healthy controls. We found the +3001 T allele was observed only in T1D patients. Notably, the +3001 T allele was in linkage disequilibrium with polymorphic sites associated with low production of HLA-G mRNA or soluble HLA-G levels. Moreover, T1D patients showed a low frequency of the HLA-G 3'UTR-17 (14bpINS/+3001T/+3003T/+3010C/+3027C/+3035T/+3142G/+3187A/+3196C). The +3010 CC genotype and the UTR-3 haplotype (14bpDEL/+3001C/+3003T/+3010C/+3027C/+3035C/+3142G/+3187A/+3196C), associated with low and moderate soluble HLA-G expression, respectively, were underrepresented in patients. The decreased expression of HLA-G at the pancreas level should be detrimental in individuals genetically prone to produce less HLA-G.

  7. Feeding and insulin increase leptin translation. Importance of the leptin mRNA untranslated regions.

    PubMed

    Lee, Mi-Jeong; Yang, Rong-Ze; Gong, Da-Wei; Fried, Susan K

    2007-01-05

    The post-transcriptional mechanisms by which feeding and insulin increase leptin production are poorly understood. Starvation of 6-7-week-old rats for 14 h decreased leptin mRNA level by only 22% but decreased plasma levels, adipose tissue leptin content, and release by over 75%. The decreased leptin with starvation was explained by >85% decrease in relative rates of leptin biosynthesis measured by metabolic labeling and immunoprecipitation. In vitro insulin treatment of adipose tissue from fed or starved rats for 2 h increased relative rates of leptin biosynthesis by 2-3-fold, and the effect was blocked by inhibition of phosphatidylinositol 3-kinase or mammalian target of rapamycin. Consistent with the hypothesis that feeding/insulin increases leptin translation, more leptin mRNA was associated with polysomes in adipose tissue of fed than starved rats, and in vitro incubation of adipose tissue of starved rats with insulin shifted leptin mRNA into polysomes. To assess the mechanisms regulating leptin translation, chimeric human leptin untranslated region (UTR) reporter constructs were transiently transfected into differentiated 3T3-L1 adipocytes. The 5'-UTR of leptin mRNA increased luciferase reporter activity 2-3-fold, whereas the full-length 3'-UTR (nucleotides 1-2804) was inhibitory (-65%). Sequences between nucleotides 462 and 1130 of the leptin 3'-UTR conferred most of the inhibitory effect. Insulin stimulated the expression of constructs that included both the full-length 5'-UTR and the inhibitory 3'-UTR, and the effect was blocked by inhibition of phosphatidylinositol 3-kinase or mammalian target of rapamycin. Our data suggest that insulin derepresses leptin translation by a mechanism that requires both the 5'-UTR and the 3'-UTR and may contribute to the increase in leptin production with feeding.

  8. Irregular G-quadruplexes Found in the Untranslated Regions of Human mRNAs Influence Translation*

    PubMed Central

    Bolduc, François; Garant, Jean-Michel; Allard, Félix; Perreault, Jean-Pierre

    2016-01-01

    G-quadruplex structures are composed of coplanar guanines and are found in both DNA and RNA. They are formed by the stacking of two or more G-quartets that are linked together by three loops. The current belief is that RNA G-quadruplexes include loops of l to 7 nucleotides in length, although recent evidence indicates that the central loop (loop 2) can be longer if loops 1 and 3 are limited to a single nucleotide each. With the objective of broadening the definition of irregular RNA G-quadruplexes, a bioinformatic search was performed to find potential G-quadruplexes located in the untranslated regions of human mRNAs (i.e. in the 5′ and 3′-UTRs) that contain either a long loop 1 or 3 of up to 40 nucleotides in length. RNA molecules including the potential sequences were then synthesized and examined in vitro by in-line probing for the formation of G-quadruplex structures. The sequences that adopted a G-quadruplex structure were cloned into a luciferase dual vector and examined for their ability to modulate translation in cellulo. Some irregular G-quadruplexes were observed to either promote or repress translation regardless of the position or the size of the long loop they possessed. Even if the composition of a RNA G-quadruplex is not quite completely understood, the results presented in this report clearly demonstrate that what defines a RNA G-quadruplex is much broader than what we previously believed. PMID:27557661

  9. Upstream open reading frame in 5'-untranslated region reduces titin mRNA translational efficiency.

    PubMed

    Cadar, Adrian G; Zhong, Lin; Lin, Angel; Valenzuela, Mauricio O; Lim, Chee C

    2014-10-10

    Titin is the largest known protein and a critical determinant of myofibril elasticity and sarcomere structure in striated muscle. Accumulating evidence that mRNA transcripts are post-transcriptionally regulated by specific motifs located in the flanking untranslated regions (UTRs) led us to consider the role of titin 5'-UTR in regulating its translational efficiency. Titin 5'-UTR is highly homologous between human, mouse, and rat, and sequence analysis revealed the presence of a stem-loop and two upstream AUG codons (uAUGs) converging on a shared in frame stop codon. We generated a mouse titin 5'-UTR luciferase reporter construct and targeted the stem-loop and each uAUG for mutation. The wild-type and mutated constructs were transfected into the cardiac HL-1 cell line and primary neonatal rat ventricular myocytes (NRVM). SV40 driven 5'-UTR luciferase activity was significantly suppressed by wild-type titin 5'-UTR (∼ 70% in HL-1 cells and ∼ 60% in NRVM). Mutating both uAUGs was found to alleviate titin 5'-UTR suppression, while eliminating the stem-loop had no effect. Treatment with various growth stimuli: pacing, PMA or neuregulin had no effect on titin 5'-UTR luciferase activity. Doxorubicin stress stimuli reduced titin 5'-UTR suppression, while H2O2 had no effect. A reported single nucleotide polymorphism (SNP) rs13422986 at position -4 of the uAUG2 was introduced and found to further repress titin 5'-UTR luciferase activity. We conclude that the uAUG motifs in titin 5'-UTR serve as translational repressors in the control of titin gene expression, and that mutations/SNPs of the uAUGs or doxorubicin stress could alter titin translational efficiency.

  10. Does HIV-1 mRNA 5'-untranslated region bear an internal ribosome entry site?

    PubMed

    Smirnova, Victoria V; Terenin, Ilya M; Khutornenko, Anastasia A; Andreev, Dmitri E; Dmitriev, Sergey E; Shatsky, Ivan N

    2016-02-01

    Unspliced human immunodeficiency virus-1 (HIV-1) mRNA is capped and therefore can be translated via conventional scanning mechanism. In addition, its 5' untranslated region (5'UTR) is thought to function as an internal ribosome entry site (IRES) during G2/M-phase of cell cycle or when cap-dependent translation is inhibited. Recently, customary methods of internal initiation demonstrating have been challenged, and consequently existence of certain IRESs of cellular origin has been put under question. Since a precise knowledge of translation initiation mechanism used by HIV may be important for cure development, presence of the IRES in HIV-1 mRNA demands a careful reexamination using contemporary stringent criteria. The key point of our strategy is to compare translation efficiency of bicistronic mRNA bearing HIV-1 unspliced mRNA 5' UTR in the intercistronic position to that of the corresponding capped monocistronic mRNA. This approach allows determination of internal initiation contribution into the overall level of particular mRNA translation. We found that both in cell-free systems and in cultured cells monocistronic mRNA with HIV-1 unspliced mRNA 5'UTR is translated significantly better than bicistronic one. Importantly, it is also true for G2/M-phase stalled cells or for cells under conditions of inhibited cap-dependent translation. Thus, in our hands contribution of internal ribosome entry into the overall level of translation driven by HIV-1 unspliced mRNA 5'UTR is negligible, and 5'-dependent scanning is a primary mechanism of its translation initiation.

  11. Phylogenetic study on the 5'-untranslated region of bovine viral diarrhoea virus isolates from Iran.

    PubMed

    Esmaelizad, Majid; Kargar-Moakhar, Rohani

    2014-01-01

    Bovine viral diarrhoea virus is a pathogen of bovids associated with reproduction system, causing in infected animals a range of ailments, from abortion to congenital defects. In this article, the nucleotide structure of the 5'-untranslated region (5-UTR) from 7 Iranian bovine diarrhoea virus (BVDV) isolates was characterized and subjected to comparative analysis against a panel of BVDV isolates from different sources. To this end, a 288 bp-long stretch of the internal ribosome entry site was amplified by RT-PCR. The PCR products subsequently cloned into PTZ57T vector and sequenced using T7 promoter primers. This resulted in detection of 3 new point mutations G → A and G → T in 2 isolates. When these findings were phylogenetically assessed, all the examined Iranian isolates were deemed to belong to the type1 of BVDV. Besides, 2 subtypes were identified among these isolates. In group A, a high level of similarity (99.2%) between Iranian isolates with a cytopathic Australian strain of BVDV-1c was detected; while in group B, the 4 Iranian isolates proved to be very similar to NADL-like BVDV-1a strains. We believe that the surprisingly high level of similarity between group A Iranian isolates and their corresponding Australian strain is likely to be an indication of a shared common ancestor. If correct, the most likely explanation of this observation is the introduction of such strains from Australia to Iran, possibly through exportation of infected live animals or animal productions (e.g. semen and meat) at some points in the past. Nevertheless, this hypothesis remains to be proved as further epidemiological work at genomic level is required to understand population of BVDV in Iran.

  12. 5' and 3' untranslated regions contribute to the differential expression of specific HLA-A alleles.

    PubMed

    René, Céline; Lozano, Claire; Villalba, Martin; Eliaou, Jean-François

    2015-12-01

    In hematopoietic stem cell transplantation (HSCT), when no HLA full-matched donor is available, alternative donors could include one HLA-mismatched donor. Recently, the low expressed HLA-C alleles have been identified as permissive mismatches for the best donor choice. Concerning HLA-A, the degree of variability of expression is poorly understood. Here, we evaluated HLA-A expression in healthy individuals carrying HLA-A*02 allele in different genotypes using flow cytometry and allele-specific quantitative RT-PCR. While an interindividual variability of HLA-A*02 cell surface expression, not due to the allele associated, was observed, no difference of the mRNA expression level was shown, suggesting the involvement of the posttranscriptional regulation. The results of qRT-PCR analyses exhibit a differential expression of HLA-A alleles with HLA-A*02 as the strongest expressed allele independently of the second allele. The associated non-HLA-A*02 alleles were differentially expressed, particularly the HLA-A*31 and HLA-A*33 alleles (strong expression) and the HLA-A*29 (low expression). The presence of specific polymorphisms in the 5' and 3' untranslated regions of the HLA-A*31 and HLA-A*33 alleles could contribute to this high level of expression. As previously described for HLA-C, low-expressed HLA-A alleles, such as HLA-A*29, could be considered as a permissive mismatch, although this needs to be confirmed by clinical studies.

  13. MicroRNA genes and their target 3'-untranslated regions are infrequently somatically mutated in ovarian cancers.

    PubMed

    Ryland, Georgina L; Bearfoot, Jennifer L; Doyle, Maria A; Boyle, Samantha E; Choong, David Y H; Rowley, Simone M; Tothill, Richard W; Gorringe, Kylie L; Campbell, Ian G

    2012-01-01

    MicroRNAs are key regulators of gene expression and have been shown to have altered expression in a variety of cancer types, including epithelial ovarian cancer. MiRNA function is most often achieved through binding to the 3'-untranslated region of the target protein coding gene. Mutation screening using massively-parallel sequencing of 712 miRNA genes in 86 ovarian cancer cases identified only 5 mutated miRNA genes, each in a different case. One mutation was located in the mature miRNA, and three mutations were predicted to alter the secondary structure of the miRNA transcript. Screening of the 3'-untranslated region of 18 candidate cancer genes identified one mutation in each of AKT2, EGFR, ERRB2 and CTNNB1. The functional effect of these mutations is unclear, as expression data available for AKT2 and EGFR showed no increase in gene transcript. Mutations in miRNA genes and 3'-untranslated regions are thus uncommon in ovarian cancer.

  14. MicroRNA Genes and Their Target 3′-Untranslated Regions Are Infrequently Somatically Mutated in Ovarian Cancers

    PubMed Central

    Doyle, Maria A.; Boyle, Samantha E.; Choong, David Y. H.; Rowley, Simone M.; Tothill, Richard W.; Gorringe, Kylie L.; Campbell, Ian G.

    2012-01-01

    MicroRNAs are key regulators of gene expression and have been shown to have altered expression in a variety of cancer types, including epithelial ovarian cancer. MiRNA function is most often achieved through binding to the 3′-untranslated region of the target protein coding gene. Mutation screening using massively-parallel sequencing of 712 miRNA genes in 86 ovarian cancer cases identified only 5 mutated miRNA genes, each in a different case. One mutation was located in the mature miRNA, and three mutations were predicted to alter the secondary structure of the miRNA transcript. Screening of the 3′-untranslated region of 18 candidate cancer genes identified one mutation in each of AKT2, EGFR, ERRB2 and CTNNB1. The functional effect of these mutations is unclear, as expression data available for AKT2 and EGFR showed no increase in gene transcript. Mutations in miRNA genes and 3′-untranslated regions are thus uncommon in ovarian cancer. PMID:22536442

  15. Species characterization in the genus Pestivirus according to palindromic nucleotide substitutions in the 5'-untranslated region.

    PubMed

    Giangaspero, Massimo; Harasawa, Ryô

    2011-06-01

    The palindromic nucleotide substitutions (PNS) at the three variable loci (V1, V2 and V3) in the 5'-untranslated region (UTR) of the Pestivirus genome have been considered for taxonomical segregation of the species, through the evaluation of 534 strains. On the basis of qualitative and quantitative secondary structure characteristics, species have been identified within the genus, determining genetic distances between species isolates, clarifying borderline and multirelated sequences, and characterizing and clustering the Pestivirus strains showing unexpected genomic sequences. Nine genomic groups have been identified: the species Bovine viral diarrhea virus 1 (BVDV-1), Bovine viral diarrhea virus 2 (BVDV-2), Border disease virus (BDV) and Classical swine fever virus (CSFV) and the tentative species Pronghorn, Giraffe, Bovine viral diarrhea virus 3 (BVDV-3) (HoBi group), Border disease virus 2 (BDV-2) (Italian small ruminant isolates) and Bungowannah. Palindromic positions have been characterized according to changes in nucleotide base-pairs identifying low variable positions (LVP) including base-pairs present in less than 80% of the genus. The determination of divergence between single strain sequences or genetic groups was obtained easily by comparing base-pairing combinations from aligned secondary structures. This provided clear information such as the level of heterogeneity within a species, the relatedness between species, or facilitating the characterization and clustering of specific strains. The BVDV-1 and BDV species resulted heterogeneous, showing isolates located on a borderline in the species. Within the BVDV-2 species, two main genogroups were identified, with strains showing common sequence characteristics to both groups (multirelated strains). They could be allocated correctly by quantitative analysis. Similarly, the relation between CSFV and BDV species appeared very clearly. Also in this case, ambiguous strain sequences could be clustered in the

  16. Molecular characterization of enterovirus 71 and coxsackievirus A16 using the 5' untranslated region and VP1 region.

    PubMed

    Zhou, Fei; Kong, Fanrong; Wang, Bin; McPhie, Kenneth; Gilbert, Gwendolyn L; Dwyer, Dominic E

    2011-03-01

    Enterovirus 71 (EV71) and coxsackievirus A16 (CVA16) are members of the species Human enterovirus A, and are both major and independent aetiological agents of hand-foot-and-mouth disease. The human enterovirus (HEV) 5' untranslated region (UTR) is fundamentally important for efficient virus replication and for virulence, whilst the VP1 region correlates well with antigenic typing by neutralization, and can be used for virus identification and evolutionary studies. A comparison was undertaken of the 5'UTR and VP1 nucleotide sequences of five EV71 clinical isolates and 10 CVA16 clinical isolates from one laboratory with the 5'UTR and VP1 sequences of 104 EV71 strains and 45 CVA16 strains available in GenBank. The genetic relationships were analysed using standard phylogenetic methods. The EV71 phylogenetic analysis showed that the VP1 sequences were clustered into three genogroups, A, B and C, with genogroups B and C further divided into five subgenogroups, B1-B5 and C1-C5, respectively. All EV71 strains were clustered similarly in the 5'UTR and VP1 trees, except for one Taiwanese strain, which demonstrated different clustering in the two trees, suggesting a recombination event in the phylogeny. The CVA16 phylogenetic analysis showed that the VP1 sequences were clustered into two genogroups, A and B, with genogroup B further divided into B1 (B1a and B1b), B2 and a possible B3; and that a similar pattern and grouping of all strains were displayed in the 5'UTR tree. This study demonstrated that comparing the two regions provides evidence of epidemiological linkage of HEV-A strains, and that mutation in the two regions plays a vital role in the evolution of these viruses. The combination of molecular typing and phylogenetic sequence analysis will be beneficial in both individual patient diagnosis and public health measures.

  17. Translational regulation of human neuronal nitric-oxide synthase by an alternatively spliced 5'-untranslated region leader exon.

    PubMed

    Newton, Derek C; Bevan, Sian C; Choi, Stephen; Robb, G Brett; Millar, Adam; Wang, Yang; Marsden, Philip A

    2003-01-03

    Expression of the neuronal nitric-oxide synthase (nNOS) mRNA is subject to complex cell-specific transcriptional regulation, which is mediated by alternative promoters. Unexpectedly, we identified a 89-nucleotide alternatively spliced exon located in the 5'-untranslated region between exon 1 variants and a common exon 2 that contains the translational initiation codon. Alternative splicing events that do not affect the open reading frame are distinctly uncommon in mammals; therefore, we assessed its functional relevance. Transient transfection of reporter RNAs performed in a variety of cell types revealed that this alternatively spliced exon acts as a potent translational repressor. Stably transfected cell lines confirmed that the alternatively spliced exon inhibited translation of the native nNOS open reading frame. Reverse transcription-PCR and RNase protection assays indicated that nNOS mRNAs containing this exon are common and expressed in both a promoter-specific and tissue-restricted fashion. Mutational analysis identified the functional cis-element within this novel exon, and a secondary structure prediction revealed that it forms a putative stem-loop. RNA electrophoretic mobility shift assay techniques revealed that a specific cytoplasmic RNA-binding complex interacts with this motif. Hence, a unique splicing event within a 5'-untranslated region is demonstrated to introduce a translational control element. This represents a newer model for the translational control of a mammalian mRNA.

  18. Incorporation of beta-globin untranslated regions into a Sindbis virus vector for augmentation of heterologous mRNA expression.

    PubMed

    Strong, T V; Hampton, T A; Louro, I; Bilbao, G; Conry, R M; Curiel, D T

    1997-06-01

    Polynucleotide immunization has been employed as a means of inducing immune responses through the introduction of antigen-encoding DNA. While immunization against specific tumor antigens may be achieved through this strategy, various candidate tumor antigens may not be approached via DNA-based vaccines as they represent transforming oncogenes. As an alternative approach, we have explored the utility of mRNA vectors for polynucleotide immunization. The transient expression achieved by mRNA may provide an efficient and safe system for stimulating immune responses to tumor-specific antigens. Our previous work demonstrated that a self-replicating RNA enhances the magnitude and duration of transgene expression for this application. Here we further modify the vector for optimal use in gene therapy through the incorporation of untranslated regions flanking the encoded transgene. The beta-globin 5' and 3' untranslated regions (UTRs) were inserted directly flanking the luciferase gene in both nonreplicative and replicative RNA constructs. In both cases, elevated and prolonged levels of luciferase expression were detected from the beta-globin UTR-flanked luciferase as compared to luciferase without these sequences. These modifications improve the ability of replicative RNA vectors to produce high, yet transient transgene expression for cancer immunotherapy strategies.

  19. A guide to design and optimization of reporter assays for 3' untranslated region mediated regulation of mammalian messenger RNAs.

    PubMed

    Van Etten, Jamie; Schagat, Trista L; Goldstrohm, Aaron C

    2013-09-15

    Post-transcriptional regulatory mechanisms are pervasive in the control of gene expression. Regulatory sequences within transcripts can control RNA processing, localization, translation efficiency, and stability of the RNA. Regulation is mediated by a diverse set of RNA binding regulators, including proteins and RNAs, which interact with specific mRNA sequences that are often found in untranslated regions. The potential for vast post-transcriptional control exists: mammalian mRNAs contain extensive untranslated regions and their genomes encode many hundreds of RNA binding proteins and non-coding RNAs. Facile quantitative methods are necessary to study the activities and mechanisms of regulatory sequences and the RNA binding factors that recognize them. Here we discuss the design and implementation of luciferase-based reporter assays to measure the effect of regulatory RNA sequences on protein and RNA expression. Protocols are described for transfection of the reporter into cells, measurement of protein expression levels with luciferase activity assays, RNA purification, and measurement of mRNA levels by reverse-transcription and quantitative polymerase chain reaction. For each assay, troubleshooting of common problems and critical controls are discussed. We present our optimized techniques and data from studies that measure specific and direct repression (i.e. negative regulation) of mRNAs by members of the PUF family of RNA binding proteins in cultured human cells.

  20. Loop structures in the 5' untranslated region and antisense RNA mediate pilE gene expression in Neisseria gonorrhoeae.

    PubMed

    Masters, Thao L; Wachter, Jenny; Hill, Stuart A

    2016-11-01

    Regulation of the Neisseria gonorrhoeae pilE gene is ill-defined. In this study, post-transcriptional effects on expression were assessed. In silico analysis predicts the formation of three putative stable stem-loop structures with favourable free energies within the 5' untranslated region of the pilE message. Using quantitative reverse transcriptase PCR analyses, we show that each loop structure forms, with introduced destabilizing stem-loop mutations diminishing loop stability. Utilizing a series of pilE translational fusions, deletion of either loop 1 or loop 2 caused a significant reduction of pilE mRNA resulting in reduced expression of the reporter gene. Consequently, the formation of the loops apparently protects the pilE transcript from degradation. Putative loop 3 contains the pilE ribosomal binding site. Consequently, its formation may influence translation. Analysis of a small RNA transcriptome revealed an antisense RNA being produced upstream of the pilE promoter that is predicted to hybridize across the 5' untranslated region loops. Insertional mutants were created where the antisense RNA is not transcribed. In these mutants, pilE transcript levels are greatly diminished, with any residual message apparently not being translated. Complementation of these insertion mutants in trans with the antisense RNA gene facilitates pilE translation yielding a pilus + phenotype. Overall, this study demonstrates a complex relationship between loop-dependent transcript protection and antisense RNA in modulating pilE expression levels.

  1. Computational identification of new structured cis-regulatory elements in the 3'-untranslated region of human protein coding genes.

    PubMed

    Chen, Xiaowei Sylvia; Brown, Chris M

    2012-10-01

    Messenger ribonucleic acids (RNAs) contain a large number of cis-regulatory RNA elements that function in many types of post-transcriptional regulation. These cis-regulatory elements are often characterized by conserved structures and/or sequences. Although some classes are well known, given the wide range of RNA-interacting proteins in eukaryotes, it is likely that many new classes of cis-regulatory elements are yet to be discovered. An approach to this is to use computational methods that have the advantage of analysing genomic data, particularly comparative data on a large scale. In this study, a set of structural discovery algorithms was applied followed by support vector machine (SVM) classification. We trained a new classification model (CisRNA-SVM) on a set of known structured cis-regulatory elements from 3'-untranslated regions (UTRs) and successfully distinguished these and groups of cis-regulatory elements not been strained on from control genomic and shuffled sequences. The new method outperformed previous methods in classification of cis-regulatory RNA elements. This model was then used to predict new elements from cross-species conserved regions of human 3'-UTRs. Clustering of these elements identified new classes of potential cis-regulatory elements. The model, training and testing sets and novel human predictions are available at: http://mRNA.otago.ac.nz/CisRNA-SVM.

  2. Distinct 3'-untranslated region elements regulate stage-specific mRNA accumulation and translation in Leishmania.

    PubMed

    McNicoll, François; Müller, Michaela; Cloutier, Serge; Boilard, Nathalie; Rochette, Annie; Dubé, Marthe; Papadopoulou, Barbara

    2005-10-21

    We recently characterized a large developmentally regulated gene family in Leishmania encoding the amastin surface proteins. While studying the regulation of these genes, we identified a region of 770 nucleotides (nt) within the 2055-nt 3'-untranslated region (3'-UTR) that regulates stage-specific gene expression at the level of translation. An intriguing feature of this 3'-UTR regulatory region is the presence of a approximately 450-nt element that is highly conserved among several Leishmania mRNAs. Here we show, using a luciferase reporter system and polysome profiling experiments, that the 450-nt element stimulates translation initiation of the amastin mRNA in response to heat shock, which is the main environmental change that the parasite encounters upon its entry into the mammalian host. Deletional analyses depicted a second region of approximately 100 nucleotides located at the 3'-end of several amastin transcripts, which also activates translation in response to elevated temperature. Both 3'-UTR regulatory elements act in an additive manner to stimulate amastin mRNA translation. In addition, we show that acidic pH encountered in the phagolysosomes of macrophages, the location of parasitic differentiation, triggers the accumulation of amastin transcripts by a distinct mechanism that is independent of the 450-nt and 100-nt elements. Overall, these important findings support the notion that stage-specific post-transcriptional regulation of the amastin mRNAs in Leishmania is complex and involves the coordination of distinct mechanisms controlling mRNA stability and translation that are independently triggered by key environmental signals inducing differentiation of the parasite within macrophages.

  3. Endothelin-1 expression is strongly repressed by AU-rich elements in the 3'-untranslated region of the gene.

    PubMed

    Reimunde, Francisco M; Castañares, Cristina; Redondo-Horcajo, Mariano; Lamas, Santiago; Rodríguez-Pascual, Fernando

    2005-05-01

    The regulation of the synthesis of the endothelial-derived vasoconstrictor ET-1 (endothelin-1) is a complex process that occurs mainly at the mRNA level. Transcription of the gene accounts for an important part of the regulation of expression, as already described for different modulators such as the cytokine TGF-beta (transforming growth factor-beta). However, very little is known about mechanisms governing ET-1 expression at the post-transcriptional level. The aim of the present study was to investigate the regulation of the ET-1 expression at this level. Since the 3'-UTR (3'-untranslated region) of mRNAs commonly contains genetic determinants for the post-transcriptional control of gene expression, we focused on the potential role of the 3'-UTR of ET-1 mRNA. Experiments performed with luciferase reporter constructs containing the 3'-UTR showed that this region exerts a potent destabilizing effect. Deletional analyses allowed us to locate this activity within a region at positions 924-1127. Some (but not all) of the AREs (AU-rich elements) present in this region were found to be essential for this mRNA-destabilizing activity. We also present evidence that cytosolic proteins from endothelial cells interact specifically with these RNA elements, and that a close correlation exists between the ability of the AREs to destabilize ET-1 mRNA and the binding of proteins to these elements. Our results are compatible with the existence of a strong repressional control of ET-1 expression mediated by destabilization of the mRNA exerted through the interaction of specific cytosolic proteins with AREs present in the 3'-UTR of the gene.

  4. The 5'-untranslated region of the mouse mammary tumor virus mRNA exhibits cap-independent translation initiation.

    PubMed

    Vallejos, Maricarmen; Ramdohr, Pablo; Valiente-Echeverría, Fernando; Tapia, Karla; Rodriguez, Felipe E; Lowy, Fernando; Huidobro-Toro, J Pablo; Dangerfield, John A; López-Lastra, Marcelo

    2010-01-01

    In this study, we demonstrate the identification of an internal ribosome entry site (IRES) within the 5'-untranslated region (5'-UTR) of the mouse mammary tumor virus (MMTV). The 5'-UTR of the full-length mRNA derived from the infectious, complete MMTV genome was cloned into a dual luciferase reporter construct containing an upstream Renilla luciferase gene (RLuc) and a downstream firefly luciferase gene (FLuc). In rabbit reticulocyte lysate, the MMTV 5'-UTR was capable of driving translation of the second cistron. In vitro translational activity from the MMTV 5'-UTR was resistant to the addition of m(7)GpppG cap-analog and cleavage of eIF4G by foot-and-mouth disease virus (FMDV) L-protease. IRES activity was also demonstrated in the Xenopus laevis oocyte by micro-injection of capped and polyadenylated bicistronic RNAs harboring the MMTV-5'-UTR. Finally, transfection assays showed that the MMTV-IRES exhibits cell type-dependent translational activity, suggesting a requirement for as yet unidentified cellular factors for its optimal function.

  5. Untranslated regions of mRNA and their role in regulation of gene expression in protozoan parasites.

    PubMed

    Rao, Shilpa J; Chatterjee, Sangeeta; Pal, Jayantapal K

    2017-03-01

    Protozoan parasites are one of the oldest living entities in this world that throughout their existence have shown excellent resilience to the odds of survival and have adapted beautifully to ever changing rigors of the environment. In view of the dynamic environment encountered by them throughout their life cycle, and in establishing pathogenesis, it is unsurprising that modulation of gene expression plays a fundamental role in their survival. In higher eukaryotes, untranslated regions (UTRs) of transcripts are one of the crucial regulators of gene expression (influencing mRNA stability and translation efficiency). Parasitic protozoan genome studies have led to the characterization (in silico, in vitro and in vivo) of a large number of their genes. Comparison of higher eukaryotic UTRs with parasitic protozoan UTRs reveals the existence of several similar and dissimilar facets of the UTRs. This review focuses on the elements of UTRs of medically important protozoan parasites and their regulatory role in gene expression. Such information may be useful to researchers in designing gene targeting strategies linked with perturbation of host-parasite relationships leading to control of specific parasites.

  6. Long 5' untranslated regions regulate the RNA stability of the deep-sea filamentous phage SW1.

    PubMed

    Jian, Huahua; Xiong, Lei; Xu, Guanpeng; Xiao, Xiang; Wang, Fengping

    2016-02-22

    Virus production in the deep-sea environment has been found to be high, and viruses have been suggested to play significant roles in the overall functioning of this ecosystem. Nevertheless, little is known about these viruses, including the mechanisms that control their production, which makes them one of the least understood biological entities on Earth. Previously, we isolated the filamentous phage SW1, whose virus production and gene transcription were found to be active at low temperatures, from a deep-sea bacterium, Shewanella piezotolerans WP3. In this study, the operon structure of phage SW1 is presented, which shows two operons with exceptionally long 5' and 3' untranslated regions (UTRs). In addition, the 5'UTR was confirmed to significantly influence the RNA stability of the SW1 transcripts. Our study revealed novel regulation of the operon and led us to propose a unique regulatory mechanism for Inoviruses. This type of RNA-based regulation may represent a mechanism for significant viral production in the cold deep biosphere.

  7. Long 5′ untranslated regions regulate the RNA stability of the deep-sea filamentous phage SW1

    PubMed Central

    Jian, Huahua; Xiong, Lei; Xu, Guanpeng; Xiao, Xiang; Wang, Fengping

    2016-01-01

    Virus production in the deep-sea environment has been found to be high, and viruses have been suggested to play significant roles in the overall functioning of this ecosystem. Nevertheless, little is known about these viruses, including the mechanisms that control their production, which makes them one of the least understood biological entities on Earth. Previously, we isolated the filamentous phage SW1, whose virus production and gene transcription were found to be active at low temperatures, from a deep-sea bacterium, Shewanella piezotolerans WP3. In this study, the operon structure of phage SW1 is presented, which shows two operons with exceptionally long 5′ and 3′ untranslated regions (UTRs). In addition, the 5′UTR was confirmed to significantly influence the RNA stability of the SW1 transcripts. Our study revealed novel regulation of the operon and led us to propose a unique regulatory mechanism for Inoviruses. This type of RNA-based regulation may represent a mechanism for significant viral production in the cold deep biosphere. PMID:26898180

  8. Identification of a nucleotide in 5′ untranslated region contributing to virus replication and virulence of Coxsackievirus A16

    PubMed Central

    Li, Zhaolong; Liu, Xin; Wang, Shaohua; Li, Jingliang; Hou, Min; Liu, Guanchen; Zhang, Wenyan; Yu, Xiao-Fang

    2016-01-01

    Coxsackievirus A16 (CA16) and enterovirus 71 (EV71) are two main causative pathogens of hand, foot and mouth disease (HFMD). Unlike EV71, virulence determinants of CA16, particularly within 5′ untranslated region (5′UTR), have not been investigated until now. Here, a series of nucleotides present in 5′UTR of lethal but not in non-lethal CA16 strains were screened by aligning nucleotide sequences of lethal circulating Changchun CA16 and the prototype G10 as well as non-lethal SHZH05 strains. A representative infectious clone based on a lethal Changchun024 sequence and infectious mutants with various nucleotide alterations in 5′UTR were constructed and further investigated by assessing virus replication in vitro and virulence in neonatal mice. Compared to the lethal infectious clone, the M2 mutant with a change from cytosine to uracil at nucleotide 104 showed weaker virulence and lower replication capacity. The predicted secondary structure of the 5′UTR of CA16 RNA showed that M2 mutant located between the cloverleaf and stem-loop II, affected interactions between the 5′UTR and the heterogeneous nuclear ribonucleoprotein K (hnRNP K) and A1 (hnRNP A1) that are important for translational activity. Thus, our research determined a virulence-associated site in the 5′UTR of CA16, providing a crucial molecular target for antiviral drug development. PMID:26861413

  9. MicroRNAs and Androgen Receptor 3′ Untranslated Region: A Missing Link in Castration-resistant Prostate Cancer?

    PubMed Central

    Sikand, Kavleen; Barik, Sailen; Shukla, Girish C.

    2012-01-01

    The ligand-activated transcription factor, androgen receptor (AR) plays a central role in the development and progression of prostate cancer. Prostate cancer initiates as an androgen-dependent disease and further accumulation of multiple sequential genetic and epigenetic alterations transform it into an aggressive, castration-resistant prostate cancer (CRPC). The molecular basis of the transition from androgen-dependent prostate cancer to CRPC remains unclear. However, it is apparent that AR plays a pivotal role in this alteration. The recent discovery that microRNAs (miRNAs) can target the function of AR suggests a functional role of these non-coding RNAs in the pathogenesis of prostate cancer. miRNAs usually function by targeting the 3′ untranslated region (UTR) of a mRNA by base-pairing interactions and modulate translation either by destabilizing the message or by repression of protein synthesis in actively translating ribosomes. Here, we discuss the potential molecular pathways through which AR targeting miRNAs may promote CRPC. Modulation of AR expression by miRNAs presents a novel therapeutic option for prostate cancer, albeit it will likely be used in combination with the existing therapies. PMID:22468168

  10. Pathway optimization by re-design of untranslated regions for L-tyrosine production in Escherichia coli

    PubMed Central

    Cheol Kim, Seong; Eun Min, Byung; Gyu Hwang, Hyun; Woo Seo, Sang; Yeol Jung, Gyoo

    2015-01-01

    L-tyrosine is a commercially important compound in the food, pharmaceutical, chemical, and cosmetic industries. Although several attempts have been made to improve L-tyrosine production, translation-level expression control and carbon flux rebalancing around phosphoenolpyruvate (PEP) node still remain to be achieved for optimizing the pathway. Here, we demonstrate pathway optimization by altering gene expression levels for L-tyrosine production in Escherichia coli. To optimize the L-tyrosine biosynthetic pathway, a synthetic constitutive promoter and a synthetic 5′-untranslated region (5′-UTR) were introduced for each gene of interest to allow for control at both transcription and translation levels. Carbon flux rebalancing was achieved by controlling the expression level of PEP synthetase using UTR Designer. The L-tyrosine productivity of the engineered E. coli strain was increased through pathway optimization resulting in 3.0 g/L of L-tyrosine titer, 0.0354 g L-tyrosine/h/g DCW of productivity, and 0.102 g L-tyrosine/g glucose yield. Thus, this work demonstrates that pathway optimization by 5′-UTR redesign is an effective strategy for the development of efficient L-tyrosine-producing bacteria. PMID:26346938

  11. A 40 kd protein binds specifically to the 5'-untranslated regions of yeast mitochondrial mRNAs.

    PubMed Central

    Papadopoulou, B; Dekker, P; Blom, J; Grivell, L A

    1990-01-01

    Using a gel mobility shift assay we show that a 40 kd protein (p40), present in extracts of yeast mitochondria, binds specifically to the 5'-untranslated leader of cytochrome c oxidase subunit II mRNA. Binding of p40 to coxII RNA protects an 8-10 nucleotide segment from diethylpyocarbonate modification, indicating that the protein interacts with only a restricted region of the 5'-leader. This segment is located at position -12 with respect to the initiation AUG. Deletion of 10 nucleotides encompassing this site completely abolishes protein binding. Nevertheless, Bal31 deletion analysis within the coxII leader shows that a major part of the leader is essential for p40 binding, suggesting that binding of the protein is also dependent on secondary structural features. p40 binds to other mitochondrial leader mRNAs including those for coxI, coxIII and cyt b. p40 is present in a cytoplasmic (rho0) petite mutant lacking mitochondrial protein synthesis. It is therefore presumably nuclear encoded. The possible biological function of the protein is discussed. Images Fig. 2. Fig. 3. Fig. 4. Fig. 5. Fig. 6. Fig. 8. Fig. 9. PMID:1701144

  12. CaMKIIalpha 3' untranslated region-directed mRNA translocation in living neurons: visualization by GFP linkage

    NASA Technical Reports Server (NTRS)

    Rook, M. S.; Lu, M.; Kosik, K. S.

    2000-01-01

    The CaMKIIalpha mRNA extends into distal hippocampal dendrites, and the 3' untranslated region (3'UTR) is sufficient to mediate this localization. We labeled the 3'UTR of the CaMKIIalpha mRNA in hippocampal cultures by using a green fluorescent protein (GFP)/MS2 bacteriophage tagging system. The CaMKIIalpha 3'UTR formed discrete granules throughout the dendrites of transfected cells. The identity of the fluorescent granules was verified by in situ hybridization. Over 30 min time periods these granules redistributed without a net increase in granule number; with depolarization there is a tendency toward increased numbers of granules in the dendrites. These observations suggest that finer time resolution of granule motility might reveal changes in the motility characteristics of granules after depolarization. So that motile granules could be tracked, shorter periods of observation were required. The movements of motile granules can be categorized as oscillatory, unidirectional anterograde, or unidirectional retrograde. Colocalization of CaMKIIalpha 3'UTR granules and synapses suggested that oscillatory movements allowed the granules to sample several local synapses. Neuronal depolarization increased the number of granules in the anterograde motile pool. Based on the time frame over which the granule number increased, the translocation of granules may serve to prepare the dendrite for mounting an adequate local translation response to future stimuli. Although the resident pool of granules can respond to signals that induce local translation, the number of granules in a dendrite might reflect its activation history.

  13. Translational control of the Xenopus laevis connexin-41 5'-untranslated region by three upstream open reading frames.

    PubMed

    Meijer, H A; Dictus, W J; Keuning, E D; Thomas, A A

    2000-10-06

    The Xenopus laevis Connexin-41 (Cx41) mRNA contains three upstream open reading frames (uORFs) in the 5'-untranslated region (UTR). We analyzed the translation efficiency of constructs containing the Cx41 5'-UTR linked to the green fluorescent protein reporter after injection of transcripts into one-cell stage Xenopus embryos. The translational efficiency of the wild-type Cx41 5'-UTR was only 2% compared with that of the beta-globin 5'-UTR. Mutation of each of the three uAUGs into AAG codons enhanced translation 82-, 9-, and 4-fold compared with the wild-type Cx41 5'-UTR. Based on these increased translation efficiencies, the percentages of ribosomes that recognized the uAUGs were calculated. Only 0.03% of the ribosomes that entered at the cap structure scanned the entire 5'-UTR and translated the main ORF. The results indicate that all uAUGs are recognized by the majority of the scanning ribosomes and that the three uAUGs strongly modulate translation efficiency in Xenopus laevis embryos. Based on these data, a model of ribosomal flow along the mRNA is postulated. We conclude that the three uORFs may play an important role in the regulation of Cx41 expression.

  14. The G-quadruplex augments translation in the 5' untranslated region of transforming growth factor β2.

    PubMed

    Agarwala, Prachi; Pandey, Satyaprakash; Mapa, Koyeli; Maiti, Souvik

    2013-03-05

    Transforming growth factor β2 (TGFβ2) is a versatile cytokine with a prominent role in cell migration, invasion, cellular development, and immunomodulation. TGFβ2 promotes the malignancy of tumors by inducing epithelial-mesenchymal transition, angiogenesis, and immunosuppression. As it is well-documented that nucleic acid secondary structure can regulate gene expression, we assessed whether any secondary motif regulates its expression at the post-transcriptional level. Bioinformatics analysis predicts an existence of a 23-nucleotide putative G-quadruplex sequence (PG4) in the 5' untranslated region (UTR) of TGFβ2 mRNA. The ability of this stretch of sequence to form a highly stable, intramolecular parallel quadruplex was demonstrated using ultraviolet and circular dichroism spectroscopy. Footprinting studies further validated its existence in the presence of a neighboring nucleotide sequence. Following structural characterization, we evaluated the biological relevance of this secondary motif using a dual luciferase assay. Although PG4 inhibits the expression of the reporter gene, its presence in the context of the entire 5' UTR sequence interestingly enhances gene expression. Mutation or removal of the G-quadruplex sequence from the 5' UTR of the gene diminished the level of expression of this gene at the translational level. Thus, here we highlight an activating role of the G-quadruplex in modulating gene expression of TGFβ2 at the translational level and its potential to be used as a target for the development of therapeutics against cancer.

  15. The WOR 1 5′ untranslated region regulates white‐opaque switching in C andida albicans by reducing translational efficiency

    PubMed Central

    Guan, Zhiyun

    2015-01-01

    Summary The human fungal pathogen C andida albicans undergoes white‐opaque phenotypic switching, which enhances its adaptation to host niches. Switching is controlled by a transcriptional regulatory network of interlocking feedback loops acting on the transcription of WOR 1, the master regulator of white‐opaque switching, but regulation of the network on the translational level is not yet explored. Here, we show that the long 5′ untranslated region of WOR 1 regulates the white‐opaque phenotype. Deletion of the WOR 1 5′ UTR promotes white‐to‐opaque switching and stabilizes the opaque state. The WOR 1 5′ UTR reduces translational efficiency and the association of the transcript with polysomes. Reduced polysome association was observed for additional key regulators of cell fate and morphology with long 5′ UTR as well. Overall, we find a novel regulatory step of white‐opaque switching at the translational level. This translational regulation is implicated for many key regulators of cell fate and morphology in C . albicans. PMID:25831958

  16. Characterization of untranslated regions of the salmonid alphavirus 3 (SAV3) genome and construction of a SAV3 based replicon.

    PubMed

    Karlsen, Marius; Villoing, Stephane; Rimstad, Espen; Nylund, Are

    2009-10-27

    Salmonid alphavirus (SAV) causes disease in farmed salmonid fish and is divided into different genetic subtypes (SAV1-6). Here we report the cloning and characterization of the 5'- and 3'- untranslated regions (UTR) of a SAV3 isolated from Atlantic salmon in Norway. The sequences of the UTRs are very similar to those of SAV1 and SAV2, but single nucleotide polymorphisms are present, also in the 3' - conserved sequence element (3'-CSE). Prediction of the RNA secondary structure suggested putative stem-loop structures in both the 5'- and 3'-ends, similar to those of alphaviruses from the terrestrial environment, indicating that the general genome replication initiation strategy for alphaviruses is also utilized by SAV. A DNA replicon vector, pmSAV3, based upon a pVAX1 backbone and the SAV3 genome was constructed, and the SAV3 non-structural proteins were used to express a reporter gene controlled by the SAV3 subgenomic promoter. Transfection of pmSAV3 into CHSE and BF2 cell lines resulted in expression of the reporter protein, confirming that the cloned SAV3 replication apparatus and UTRs are functional in fish cells.

  17. The 3' untranslated region of human Cyclin-Dependent Kinase 5 Regulatory subunit 1 contains regulatory elements affecting transcript stability

    PubMed Central

    Moncini, Silvia; Bevilacqua, Annamaria; Venturin, Marco; Fallini, Claudia; Ratti, Antonia; Nicolin, Angelo; Riva, Paola

    2007-01-01

    Background CDK5R1 plays a central role in neuronal migration and differentiation during central nervous system development. CDK5R1 has been implicated in neurodegenerative disorders and proposed as a candidate gene for mental retardation. The remarkable size of CDK5R1 3'-untranslated region (3'-UTR) suggests a role in post-transcriptional regulation of CDK5R1 expression. Results The bioinformatic study shows a high conservation degree in mammals and predicts several AU-Rich Elements (AREs). The insertion of CDK5R1 3'-UTR into luciferase 3'-UTR causes a decreased luciferase activity in four transfected cell lines. We identified 3'-UTR subregions which tend to reduce the reporter gene expression, sometimes in a cell line-dependent manner. In most cases the quantitative analysis of luciferase mRNA suggests that CDK5R1 3'-UTR affects mRNA stability. A region, leading to a very strong mRNA destabilization, showed a significantly low half-life, indicating an accelerated mRNA degradation. The 3' end of the transcript, containing a class I ARE, specifically displays a stabilizing effect in neuroblastoma cell lines. We also observed the interaction of the stabilizing neuronal RNA-binding proteins ELAV with the CDK5R1 transcript in SH-SY5Y cells and identified three 3'-UTR sub-regions showing affinity for ELAV proteins. Conclusion Our findings evince the presence of both destabilizing and stabilizing regulatory elements in CDK5R1 3'-UTR and support the hypothesis that CDK5R1 gene expression is post-transcriptionally controlled in neurons by ELAV-mediated mechanisms. This is the first evidence of the involvement of 3'-UTR in the modulation of CDK5R1 expression. The fine tuning of CDK5R1 expression by 3'-UTR may have a role in central nervous system development and functioning, with potential implications in neurodegenerative and cognitive disorders. PMID:18053171

  18. 5′ and 3′ Untranslated Regions Strongly Enhance Performance of Geminiviral Replicons in Nicotiana benthamiana Leaves

    PubMed Central

    Diamos, Andrew G.; Rosenthal, Sun H.; Mason, Hugh S.

    2016-01-01

    We previously reported a recombinant protein production system based on a geminivirus replicon that yields high levels of vaccine antigens and monoclonal antibodies in plants. The bean yellow dwarf virus (BeYDV) replicon generates massive amounts of DNA copies, which engage the plant transcription machinery. However, we noticed a disparity between transcript level and protein production, suggesting that mRNAs could be more efficiently utilized. In this study, we systematically evaluated genetic elements from human, viral, and plant sources for their potential to improve the BeYDV system. The tobacco extensin terminator enhanced transcript accumulation and protein production compared to other commonly used terminators, indicating that efficient transcript processing plays an important role in recombinant protein production. Evaluation of human-derived 5′ untranslated regions (UTRs) indicated that many provided high levels of protein production, supporting their cross-kingdom function. Among the viral 5′ UTRs tested, we found the greatest enhancement with the tobacco mosaic virus omega leader. An analysis of the 5′ UTRs from the Arabidopsis thaliana and Nicotinana benthamiana photosystem I K genes found that they were highly active when truncated to include only the near upstream region, providing a dramatic enhancement of transgene production that exceeded that of the tobacco mosaic virus omega leader. The tobacco Rb7 matrix attachment region inserted downstream from the gene of interest provided significant enhancement, which was correlated with a reduction in plant cell death. Evaluation of Agrobacterium strains found that EHA105 enhanced protein production and reduced cell death compared to LBA4301 and GV3101. We used these improvements to produce Norwalk virus capsid protein at >20% total soluble protein, corresponding to 1.8 mg/g leaf fresh weight, more than twice the highest level ever reported in a plant system. We also produced the monoclonal antibody

  19. A functional role for the conserved protonatable hairpins in the 5' untranslated region of turnip yellow mosaic virus RNA.

    PubMed Central

    Hellendoorn, K; Verlaan, P W; Pleij, C W

    1997-01-01

    The 5' untranslated region (UTR) of the RNA of several tymoviruses contains conserved hairpins with protonatable internal loops, consisting of C-C and C-A mismatches (K. Hellendoorn, P. J. A. Michiels, R. Buitenhuis, and C. W. A. Pleij, Nucleic Acids Res. 24, 4910-4917, 1996). Here, we present a functional analysis of the 5' UTR of turnip yellow mosaic virus (TYMV) RNA, which contains two protonatable hairpins with nearly identical internal loops. Mutations were introduced in an infectious cDNA clone, and T7 RNA transcripts were used to infect Chinese cabbage plants. Different symptoms were observed for the various mutants, pointing to a functional role of the C-C and C-A mismatches in the hairpins of the 5' UTR. The replication of the virus is influenced by the mutations made, while in vitro translation studies showed that the expression of the two overlapping reading frames of TYMV is not influenced by the secondary structure of the leader. Various mutants were propagated for up to five serial passages of infection, and the sequence of the 5' UTR was determined. This resulted in virus RNA with new non-wild-type sequences that produced the wild-type phenotype in infected plants. Remarkably, in all cases C-C or C-A mismatches were introduced. The internal loop of the 5'-proximal hairpin seems to be more important for the viral life cycle than that of the second hairpin. A deletion of 75% of the leader, including the two hairpins, resulted in a virus that was deficient in viral spread. Since the ratio between filled and empty capsids was changed drastically by this mutation, a role of the 5' UTR in viral packaging is proposed. PMID:9343237

  20. A eukaryotic-like 3′ untranslated region in Salmonella enterica hilD mRNA

    PubMed Central

    López-Garrido, Javier; Puerta-Fernández, Elena; Casadesús, Josep

    2014-01-01

    Long 3′ untranslated regions (3′UTRs) are common in eukaryotic mRNAs. In contrast, long 3′UTRs are rare in bacteria, and have not been characterized in detail. We describe a 3′UTR of 310 nucleotides in hilD mRNA, a transcript that encodes a transcriptional activator of Salmonella enterica pathogenicity island 1 (SPI-1). Deletion of the hilD 3′UTR increases the hilD mRNA level, suggesting that the hilD 3′UTR may play a role in hilD mRNA turnover. Cloning of the hilD 3′UTR downstream of the green fluorescent protein (gfp) gene decreases green fluorescent protein (GFP) activity in both Escherichia coli and S. enterica, indicating that the hilD 3′UTR can act as an independent module. S. enterica mutants lacking either ribonuclease E or polynucleotide phosphorylase contain similar amounts of hilD and hilD Δ3′UTR mRNAs, suggesting that the hilD 3′UTR is a target for hilD mRNA degradation by the degradosome. The hilD 3′UTR is also necessary for modulation of hilD and SPI-1 expression by the RNA chaperone Hfq. Overexpression of SPI-1 in the absence of the hilD 3′UTR retards Salmonella growth and causes uncontrolled invasion of epithelial cells. Based on these observations, we propose that the S. enterica hilD 3′UTR is a cis-acting element that contributes to cellular homeostasis by promoting hilD mRNA turnover. PMID:24682814

  1. Chikungunya Virus 3′ Untranslated Region: Adaptation to Mosquitoes and a Population Bottleneck as Major Evolutionary Forces

    PubMed Central

    Chen, Rubing; Wang, Eryu; Tsetsarkin, Konstantin A.; Weaver, Scott C.

    2013-01-01

    The 3′ untranslated genome region (UTR) of arthropod-borne viruses is characterized by enriched direct repeats (DRs) and stem-loop structures. Despite many years of theoretical and experimental study, on-going positive selection on the 3′UTR had never been observed in ‘real-time,’ and the role of the arbovirus 3′UTR remains poorly understood. We observed a lineage-specific 3′UTR sequence pattern in all available Asian lineage of the mosquito-borne alphavirus, chikungunya virus (CHIKV) (1958–2009), including complicated mutation and duplication patterns of the long DRs. Given that a longer genome is usually associated with less efficient replication, we hypothesized that the fixation of these genetic changes in the Asian lineage 3′UTR was due to their beneficial effects on adaptation to vectors or hosts. Using reverse genetic methods, we examined the functional importance of each direct repeat. Our results suggest that adaptation to mosquitoes, rather than to mammalian hosts, is a major evolutionary force on the CHIKV 3′UTR. Surprisingly, the Asian 3′UTR appeared to be inferior to its predicted ancestral sequence for replication in both mammals and mosquitoes, suggesting that its fixation in Asia was not a result of directional selection. Rather, it may have resulted from a population bottleneck during its introduction from Africa to Asia. We propose that this introduction of a 3′UTR with deletions led to genetic drift and compensatory mutations associated with the loss of structural/functional constraints, followed by two independent beneficial duplications and fixation due to positive selection. Our results provide further evidence that the limited epidemic potential of the Asian CHIKV strains resulted from founder effects that reduced its fitness for efficient transmission by mosquitoes there. PMID:24009512

  2. PTB Binds to the 3’ Untranslated Region of the Human Astrovirus Type 8: A Possible Role in Viral Replication

    PubMed Central

    Espinosa-Hernández, Wendy; Velez-Uriza, Dora; Valdés, Jesús; Vélez-Del Valle, Cristina; Salas-Benito, Juan; Martínez-Contreras, Rebeca; García-Espítia, Matilde; Salas-Benito, Mariana; Vega-Almeida, Tania; De Nova-Ocampo, Mónica

    2014-01-01

    The 3′ untranslated region (3′UTR) of human astroviruses (HAstV) consists of two hairpin structures (helix I and II) joined by a linker harboring a conserved PTB/hnRNP1 binding site. The identification and characterization of cellular proteins that interact with the 3′UTR of HAstV-8 virus will help to uncover cellular requirements for viral functions. To this end, mobility shift assays and UV cross-linking were performed with uninfected and HAstV-8-infected cell extracts and HAstV-8 3′UTR probes. Two RNA-protein complexes (CI and CII) were recruited into the 3′UTR. Complex CII formation was compromised with cold homologous RNA, and seven proteins of 35, 40, 45, 50, 52, 57/60 and 75 kDa were cross-linked to the 3′UTR. Supermobility shift assays indicated that PTB/hnRNP1 is part of this complex, and 3′UTR-crosslinked PTB/hnRNP1 was immunoprecipitated from HAstV-8 infected cell-membrane extracts. Also, immunofluorescence analyses revealed that PTB/hnRNP1 is distributed in the nucleus and cytoplasm of uninfected cells, but it is mainly localized perinuclearly in the cytoplasm of HAstV-8 infected cells. Furthermore, the minimal 3′UTR sequences recognized by recombinant PTB are those conforming helix I, and an intact PTB/hnRNP1-binding site. Finally, small interfering RNA-mediated PTB/hnRNP1 silencing reduced synthesis viral genome and virus yield in CaCo2 cells, suggesting that PTB/hnRNP1 is required for HAstV replication. In conclusion, PTB/hnRNP1 binds to the 3′UTR HAstV-8 and is required or participates in viral replication. PMID:25406089

  3. Destabilization of ERBB2 transcripts by targeting 3' untranslated region messenger RNA associated HuR and histone deacetylase-6.

    PubMed

    Scott, Gary K; Marx, Corina; Berger, Crystal E; Saunders, Laura R; Verdin, Eric; Schäfer, Stefan; Jung, Manfred; Benz, Christopher C

    2008-07-01

    In addition to repressing ERBB2 promoter function, histone deacetylase (HDAC) inhibitors induce the accelerated decay of mature ERBB2 transcripts; the mechanism mediating this transcript destabilization is unknown but depends on the 3' untranslated region (UTR) of ERBB2 mRNA. Using ERBB2-overexpressing human breast cancer cells (SKBR3), the mRNA stability factor HuR was shown to support ERBB2 transcript integrity, bind and endogenously associate with a conserved U-rich element within the ERBB2 transcript 3' UTR, coimmunoprecipitate with RNA-associated HDAC activity, and colocalize with HDAC6. HDAC6 also coimmunoprecipitates with HuR in an RNA-dependent manner and within 6 hours of exposure to a pan-HDAC inhibitor dose, that does not significantly alter cytosolic HuR levels or HuR binding to ERBB2 mRNA. Cellular ERBB2 transcript levels decline while remaining physically associated with HDAC6. Knockdown of HDAC6 protein by small interfering RNA partially suppressed the ERBB2 transcript decay induced by either pan-HDAC or HDAC6-selective enzymatic inhibitors. Three novel hydroxamates, ST71, ST17, and ST80 were synthesized and shown to inhibit HDAC6 with 14-fold to 31-fold greater selectivity over their binding and inhibition of HDAC1. Unlike more potent pan-HDAC inhibitors, these HDAC6-selective inhibitors produced dose-dependent growth arrest of ERBB2-overexpressing breast cancer cells by accelerating the decay of mature ERBB2 mRNA without repressing ERBB2 promoter function. In sum, these findings point to the therapeutic potential of HuR and HDAC6-selective inhibitors, contrasting ERBB2 stability effects induced by HDAC6 enzymatic inhibition and HDAC6 protein knockdown, and show that ERBB2 transcript stability mechanisms include exploitable targets for the development of novel anticancer therapies.

  4. Reselection of a genomic upstream open reading frame in mouse hepatitis coronavirus 5'-untranslated-region mutants.

    PubMed

    Wu, Hung-Yi; Guan, Bo-Jhih; Su, Yu-Pin; Fan, Yi-Hsin; Brian, David A

    2014-01-01

    An AUG-initiated upstream open reading frame (uORF) encoding a potential polypeptide of 3 to 13 amino acids (aa) is found within the 5' untranslated region (UTR) of >75% of coronavirus genomes based on 38 reference strains. Potential CUG-initiated uORFs are also found in many strains. The AUG-initiated uORF is presumably translated following genomic 5'-end cap-dependent ribosomal scanning, but its function is unknown. Here, in a reverse-genetics study with mouse hepatitis coronavirus, the following were observed. (i) When the uORF AUG-initiating codon was replaced with a UAG stop codon along with a U112A mutation to maintain a uORF-harboring stem-loop 4 structure, an unimpaired virus with wild-type (WT) growth kinetics was recovered. However, reversion was found at all mutated sites within five virus passages. (ii) When the uORF was fused with genomic (main) ORF1 by converting three in-frame stop codons to nonstop codons, a uORF-ORF1 fusion protein was made, and virus replicated at WT levels. However, a frameshifting G insertion at virus passage 7 established a slightly 5'-extended original uORF. (iii) When uAUG-eliminating deletions of 20, 30, or 51 nucleotides (nt) were made within stem-loop 4, viable but debilitated virus was recovered. However, a C80U mutation in the first mutant and an A77G mutation in the second appeared by passage 10, which generated alternate uORFs that correlated with restored WT growth kinetics. In vitro, the uORF-disrupting nondeletion mutants showed enhanced translation of the downstream ORF1 compared with the WT. These results together suggest that the uORF represses ORF1 translation yet plays a beneficial but nonessential role in coronavirus replication in cell culture.

  5. The Modulatory Effects of the Polymorphisms in GLA 5'-Untranslated Region Upon Gene Expression Are Cell-Type Specific.

    PubMed

    Ferreira, Susana; Reguenga, Carlos; Oliveira, João Paulo

    2015-01-01

    Lysosomal α-galactosidase A (αGal) is the enzyme deficient in Fabry disease (FD). The 5'-untranslated region (5'UTR) of the αGal gene (GLA) shows a remarkable degree of variation with three common single nucleotide polymorphisms at nucleotide positions c.-30G>A, c.-12G>A and c.-10C>T. We have recently identified in young Portuguese stroke patients a fourth polymorphism, at c.-44C>T, co-segregating in cis with the c.-12A allele. In vivo, the c.-30A allele is associated with higher enzyme activity in plasma, whereas c.-10T is associated with moderately decreased enzyme activity in leucocytes. Limited data suggest that c.-44T might be associated with increased plasma αGal activity. We have used a luciferase reporter system to experimentally assess the relative modulatory effects on gene expression of the different GLA 5'UTR polymorphisms, as compared to the wild-type sequence, in four different human cell lines. Group-wise, the relative luciferase expression patterns of the various GLA variant isoforms differed significantly in all four cell lines, as evaluated by non-parametric statistics, and were cell-type specific. Some of the post hoc pairwise statistical comparisons were also significant, but the observed effects of the GLA 5'UTR polymorphisms upon the luciferase transcriptional activity in vitro did not consistently replicate the in vivo observations.These data suggest that the GLA 5'UTR polymorphisms are possible modulators of the αGal expression. Further studies are needed to elucidate the biological and clinical implications of these observations, particularly to clarify the effect of these polymorphisms in individuals carrying GLA variants associated with high residual enzyme activity, with no or mild FD clinical phenotypes.

  6. Fragile X mental retardation protein interactions with a G quadruplex structure in the 3'-untranslated region of NR2B mRNA.

    PubMed

    Stefanovic, Snezana; DeMarco, Brett A; Underwood, Ayana; Williams, Kathryn R; Bassell, Gary J; Mihailescu, Mihaela Rita

    2015-12-01

    Fragile X syndrome, the most common cause of inherited intellectual disability, is caused by a trinucleotide CGG expansion in the 5'-untranslated region of the FMR1 gene, which leads to the loss of expression of the fragile X mental retardation protein (FMRP). FMRP, an RNA-binding protein that regulates the translation of specific mRNAs, has been shown to bind a subset of its mRNA targets by recognizing G quadruplex structures. It has been suggested that FMRP controls the local protein synthesis of several protein components of the post synaptic density (PSD) in response to specific cellular needs. We have previously shown that the interactions between FMRP and mRNAs of the PSD scaffold proteins PSD-95 and Shank1 are mediated via stable G-quadruplex structures formed within the 3'-untranslated regions of these mRNAs. In this study we used biophysical methods to show that a comparable G quadruplex structure forms in the 3'-untranslated region of the glutamate receptor subunit NR2B mRNA encoding for a subunit of N-methyl-d-aspartate (NMDA) receptors that is recognized specifically by FMRP, suggesting a common theme for FMRP recognition of its dendritic mRNA targets.

  7. Porcine SOX9 Gene Expression Is Influenced by an 18 bp Indel in the 5'-Untranslated Region.

    PubMed

    Brenig, Bertram; Duan, Yanyu; Xing, Yuyun; Ding, Nengshui; Huang, Lusheng; Schütz, Ekkehard

    2015-01-01

    Sex determining region Y-box 9 (SOX9) is an important regulator of sex and skeletal development and is expressed in a variety of embryonal and adult tissues. Loss or gain of function resulting from mutations within the coding region or chromosomal aberrations of the SOX9 locus lead to a plethora of detrimental phenotypes in humans and animals. One of these phenotypes is the so-called male-to-female or female-to-male sex-reversal which has been observed in several mammals including pig, dog, cat, goat, horse, and deer. In 38,XX sex-reversal French Large White pigs, a genome-wide association study suggested SOX9 as the causal gene, although no functional mutations were identified in affected animals. However, besides others an 18 bp indel had been detected in the 5'-untranslated region of the SOX9 gene by comparing affected animals and controls. We have identified the same indel (Δ18) between position +247 bp and +266 bp downstream the transcription start site of the porcine SOX9 gene in four other pig breeds; i.e., German Large White, Laiwu Black, Bamei, and Erhualian. These animals have been genotyped in an attempt to identify candidate genes for porcine inguinal and/or scrotal hernia. Because the 18 bp segment in the wild type 5'-UTR harbours a highly conserved cAMP-response element (CRE) half-site, we analysed its role in SOX9 expression in vitro. Competition and immunodepletion electromobility shift assays demonstrate that the CRE half-site is specifically recognized by CREB. Both binding of CREB to the wild type as well as the absence of the CRE half-site in Δ18 reduced expression efficiency in HEK293T, PK-15, and ATDC5 cells significantly. Transfection experiments of wild type and Δ18 SOX9 promoter luciferase constructs show a significant reduction of RNA and protein levels depending on the presence or absence of the 18 bp segment. Hence, the data presented here demonstrate that the 18 bp indel in the porcine SOX9 5'-UTR is of functional importance and may

  8. The 5′ untranslated regions (UTRs) of CCN1, CCN2, and CCN4 exhibit cryptic promoter activity

    PubMed Central

    Huang, Bau-Lin; Dornbach, Lisa M.

    2007-01-01

    CCNs are structurally related matricellular proteins that are highly expressed in many embryonic and adult tissues, including the skeletal system and tumors, where canonical cap-dependent translation is suppressed under hypoxic environments. CCNs are encoded by mRNAs containing long G/C rich 5′-untranslated regions (5′-UTRs). Given that they are expressed under conditions of cellular stress, it has been suggested that the long G/C-rich regions contain internal ribosomal entry sites (IRES) that allow these mRNAS to be translated under conditions where cap-dependent translation is suppressed. Previously published work supported this possibility. However, recent studies have shown that a number of previously reported cellular IRES elements do not in fact possess IRES activity. Here we aimed to reveal whether the 5′UTRs of CCNs harbor IRES activities. The 5′UTRs of CCN1, 2, and 4 were tested in this study. Our results showed that the 5′UTRs of these genes do not contain IRES elements, but instead appear to contain cryptic promoters. Both promoterless and hairpin-containing dicistronic tests showed that transcription was initiated by cryptic promoter elements in 5′UTRs of CCN1, 2, and 4. When dicistronic mRNAs were translated in vitro or in vivo, no IRES activities were detected in the 5′UTRs of CCN1, 2, and 4. Furthermore, these cryptic promoter activities from 5′UTRs of CCN1, 2, and 4 could be detected in various cell types, including chondrocytes, osteoblasts, and endothelial cells, where the cryptic promoter permitted varying degrees of activation. In addition, the core promoter element of the CCN2 5′UTR was identified. CCNs are expressed under conditions of cellular stress, and it has been suggested that some CCN family members utilize IRES-mediated translation initiation to facilitate this expression. We found no evidence for IRES activity, but rather found that the unusually long 5′UTRs of CCNs 1, 2, and 4 harbor cryptic promoters that showed

  9. In silico analysis of microRNAS targeting the HLA-G 3' untranslated region alleles and haplotypes.

    PubMed

    Castelli, Erick C; Moreau, Philippe; Oya e Chiromatzo, Alynne; Mendes-Junior, Celso Teixeira; Veiga-Castelli, Luciana C; Yaghi, Layale; Giuliatti, Silvana; Carosella, Edgardo D; Donadi, Eduardo Antonio

    2009-12-01

    It has been reported that microRNAs (miRNA) may have allele-specific targeting for the 3' untranslated region (3' UTR) of the HLA-G locus. In a previous study, we reported 11 3'UTR haplotypes encompassing the 14-bp insertion/deletion polymorphism and seven SNPs (+3003 T/C, +3010 C/G, +3027 C/A, +3035 C/T, +3142 C/G, +3187 A/G, and +3196 C/G), of which only the +3142 C/G SNP has been reported to influence the binding of miRNAs. Using bioinformatics analyses, we identified putative miRNA-binding sites considering the haplotypes encompassing these eight polymorphic sites, and we ranked the lowest free energies that could potentially lead to an mRNA degradation or translational repression. When a specific haplotype or a particular SNP was associated with a miRNA-binding site, we defined a free energy difference of 4 kcal/mol between alleles to classify them energetically distant. The best results were obtained for the miR-513a-5p, miR-518c*, miR-1262 and miR-92a-1*, miR-92a-2*, miR-661, miR-1224-5p, and miR-433 miRNAs, all influencing one or more of the +3003, +3010, +3027, and +3035 SNPs. The miR-2110, miR-93, miR-508-5p, miR-331-5p, miR-616, miR-513b, and miR-589* miRNAs targeted the 14-bp fragment region, and miR-148a, miR-19a*, miR-152, mir-148b, and miR-218-2 also influenced the +3142 C/G polymorphism. These results suggest that these miRNAs might play a relevant role on the HLA-G expression pattern.

  10. Partial deletion of stem-loop 2 in the 3' untranslated region of foot-and-mouth disease virus identifies a region that is dispensable for virus replication.

    PubMed

    Biswal, Jitendra K; Subramaniam, Saravanan; Ranjan, Rajeev; Pattnaik, Bramhadev

    2016-08-01

    The 3' untranslated region (3' UTR) of the foot-and-mouth disease virus (FMDV) genome plays an essential role in virus replication, but the properties of the 3' UTR are not completely defined. In order to determine the role of different regions of the 3' UTR in FMDV replication, we conducted site-directed mutagenesis of the 3' UTR of FMDV serotype O IND R2/1975 using a cDNA clone. Through independent serial deletions in various regions of the 3' UTR, we demonstrated that deletion of nucleotides between the stem-loop (SL) structures and in the beginning and the end regions of the SL2 structure could be lethal for FMDV replication. However, a block deletion of 20 nucleotides (nt 60 to 79) in the middle of SL2 did not affect the viability of FMDV in cultured cells. Characterisation of the deletion mutant virus (O(R2/1975-Δ3'UTR 60-79)) revealed no significant difference in growth kinetics or RNA replication ability compared to the parental virus. However, the mutant virus produced slightly larger plaques when compared to the parental virus. This is the first description of a dispensable 20-nucleotide region in SL2 of the FMDV 3' UTR.

  11. Functional Characterization of a Single Nucleotide Polymorphism in the 3' Untranslated Region of Sheep DLX3 Gene.

    PubMed

    Rong, Enguang; Zhang, Zhiwei; Qiao, Shupei; Yang, Hua; Yan, Xiaohong; Li, Hui; Wang, Ning

    2015-01-01

    The Distal-less 3 (homeobox protein DLX-3), a transcription factor, is critical for the development of hair follicle and hair formation and regeneration. We previously identified and found that four SNPs (c. *118T>C, c. *228T>C, c. *688A>G and c. *1,038_1,039 insC) in 3' untranslated region (UTR) of sheep DLX3 were in high linkage disequilibrium with each other and significantly associated with wool crimp (P<0.05), however, the underlying mechanisms by which these SNPs affect the wool crimp remains unknown. In the present study, we performed association analysis between these four identified SNPs and DLX3 gene expression in sheep skin using quantitative real-time RT-PCR. The results showed that these SNPs were significantly associated with sheep skin DLX3 mRNA expression levels. Then, we constructed DLX3 3'UTR luciferase reporters and validated the association. The reporter assays showed that the three major haplotypes, derived from the four SNPs, had significantly different effects on luciferase reporter activity and the four SNPs also had significantly different allelic effects on the luciferase reporter activity (p < 0.05). Bioinformatics analysis showed that the SNP (c. *1,038_1,039 insC) was located within a potential miR-188 binding site of the 3'UTR of sheep DLX3 mRNA. This SNP may affect miR-188-mediated DLX3 gene expression and result in phenotypic variation. To test the hypothesis, we investigated the effects of miR-188 mimic and inhibitor on the activity of the DLX3 3'UTR luciferase reporter with different SNP alleles. The results showed that in both sheep fetal fibroblasts (SFFs) and human HaCaT cells, miR-188 mimic could significantly decrease the allele D (deletion) luciferase reporter activity (p < 0.05), but miR-188 inhibitor could increased the reporter activitiy. However, neither miR-188 mimc nor inhibitor could influence the allele I (insertion) reporter activity. In addition, transfection of miR-188 mimic dramatically decreased the endogenous

  12. The parvovirus H-1 NS2 protein affects viral gene expression through sequences in the 3' untranslated region.

    PubMed

    Li, X; Rhode, S L

    1993-05-01

    We reported previously that an NS2 null mutant of parvovirus H-1 (H-1SA) was capable of lytic growth in human and hamster cells, but not in rat cells (Li and Rhode, 1991). The host-range phenotype of H-1SA was also manifested in newborn rats and was associated with a reduction of viral protein synthesis to about 10% of wild-type virus and an absence of virions in cultured rat fibroblasts. However, the H-1SA mRNAs for NS1 and capsid proteins, R1 and R3, accumulated to wild-type levels and translated well with a cell free rabbit reticulocyte lysate. These results indicate that NS2 plays an important role in the regulation of viral protein synthesis in rat cells in vivo and in vitro, but NS2 is largely dispensable in other types of cells, such as human and hamster cells. To analyze whether the 5' and 3' untranslated regions (UTR) of viral RNA are involved in the regulation by NS2, the viral VP2 gene was replaced by a reporter gene, firefly luciferase, in a plasmid clone of viral sequences and the protein synthesis under the control of P38 was evaluated by luciferase assay. Cells were transfected with luciferase expressing plasmids and subsequently infected with wild-type H-1 or H-1SA. We were able to mimic the defect in expression that we observed in cultured cells and animals with virus infection. Luciferase activity in H- 1SA-infected rat cells was about 10-fold lower than that in H-1-infected rat cells, but only 2-fold lower or less in H-1SA-infected human cells and hamster cells compared to wild-type H-1. These results are consistent with our previous data that NS2 has a host-range phenotype in the natural host of H-1, the rat. Deletion of 5' UTR sequences from P38 transcripts reduced the overall P38-luc expression but expression was NS2 independent, whereas deletion of the terminal 3' UTR sequences of viral RNA reduced NS2-dependent expression in rat cells. These results suggest that the regulation of viral protein synthesis by NS2 depends on RNA sequences in the

  13. The 5' untranslated region and Gag product of Idefix, a long terminal repeat-retrotransposon from Drosophila melanogaster, act together to initiate a switch between translated and untranslated states of the genomic mRNA.

    PubMed

    Meignin, Carine; Bailly, Jean-Luc; Arnaud, Frédérick; Dastugue, Bernard; Vaury, Chantal

    2003-11-01

    Idefix is a long terminal repeat (LTR)-retrotransposon present in Drosophila melanogaster which shares similarities with vertebrates retroviruses both in its genomic arrangement and in the mechanism of transposition. Like in retroviruses, its two LTRs flank a long 5' untranslated region (5'UTR) and three open reading frames referred to as the gag, pol, and env genes. Here we report that its 5'UTR, located upstream of the gag gene, can fold into highly structured domains that are known to be incompatible with efficient translation by ribosome scanning. Using dicistronic plasmids analyzed by both (i) in vitro transcription and translation in rabbit reticulocyte or wheat germ lysates and (ii) in vivo expression in transgenic flies, we show that the 5'UTR of Idefix exhibits an internal ribosome entry site (IRES) activity that is able to promote translation of a downstream cistron in a cap-independent manner. The functional state of this novel IRES depends on eukaryotic factors that are independent of their host origin. However, in vivo, its function can be down-regulated by trans-acting factors specific to tissues or developmental stages of its host. We identify one of these trans-acting factors as the Gag protein encoded by Idefix itself. Our data support a model in which nascent Gag is able to block translation initiated from the viral mRNA and thus its own translation. These data highlight the fact that LTR-retrotransposons may autoregulate their replication cycle through their Gag production.

  14. Is Single-Strand Conformation Polymorphism Analysis of the Full 5′ Untranslated Region an Adequate Approach To Study Hepatitis C Virus Quasispecies Distribution? ▿

    PubMed Central

    Vera-Otarola, Jorge; Barría, María Inés; León, Ursula; Carvallo, Pilar; Soza, Alejandro; López-Lastra, Marcelo

    2009-01-01

    Single-strand conformation polymorphism (SSCP) analysis is used by many laboratories to study the quasispecies distribution of the hepatitis C virus (HCV). Here we question the validity of this experimental approach, as conclusions are drawn from the analysis of the migration patterns of two ssDNA molecules and not from RNA. Using previously characterized mutants of the HCV 5′ untranslated regions, we show that contrary to what has been predicted, SSCP migration patterns of DNA amplicons with differences in their nucleotide sequences generated from the full 5′ UTR of HCV are not necessarily unique. PMID:19553315

  15. Analysis of the H gene, the central untranslated region and the proximal coding part of the F gene of wild-type and vaccine canine distemper viruses.

    PubMed

    Haas, L; Liermann, H; Harder, T C; Barrett, T; Löchelt, M; von Messling, V; Baumgärtner, W; Greiser-Wilke, I

    1999-09-01

    This paper summarizes the results of the genetic analysis of several parts of the genome of canine distemper virus (CDV) field isolates and vaccine viruses. The haemagglutinin (H) gene analysis showed that recent viruses did not differ significantly from vaccine strains. The analysis of the long untranslated region between the matrix (M) and fusion (F) gene revealed distinct genetic heterogeneity. The putative F protein start codon AUG461 of vaccine strain Onderstepoort was found to be mutated in all wild-type isolates and in another vaccine strain. The proximal coding part of the F gene was well conserved. Phylogenetic analysis of this segment showed the presence of several cocirculating CDV genotypes.

  16. Is single-strand conformation polymorphism analysis of the full 5' untranslated region an adequate approach to study hepatitis C virus quasispecies distribution?

    PubMed

    Vera-Otarola, Jorge; Barría, María Inés; León, Ursula; Carvallo, Pilar; Soza, Alejandro; López-Lastra, Marcelo

    2009-09-01

    Single-strand conformation polymorphism (SSCP) analysis is used by many laboratories to study the quasispecies distribution of the hepatitis C virus (HCV). Here we question the validity of this experimental approach, as conclusions are drawn from the analysis of the migration patterns of two ssDNA molecules and not from RNA. Using previously characterized mutants of the HCV 5' untranslated regions, we show that contrary to what has been predicted, SSCP migration patterns of DNA amplicons with differences in their nucleotide sequences generated from the full 5' UTR of HCV are not necessarily unique.

  17. Structure and expression of the guinea pig preproenkephalin gene: site-specific cleavage in the 3' untranslated region yields truncated mRNA transcripts in specific brain regions.

    PubMed Central

    LaForge, K S; Unterwald, E M; Kreek, M J

    1995-01-01

    We isolated the guinea pig preproenkephalin gene from a genomic library by hybridization to a rat cDNA probe. The entire nucleotide sequence of the gene was determined. Genomic Southern blot hybridization demonstrated that the gene exists in a single copy within the genome. On the basis of RNase protection transcript mapping and homology comparisons with known preproenkephalin sequences from other species and assuming a poly(A) tail length of 100 residues, we predicted an mRNA transcript of approximately 1,400 nucleotides encoded by three exons. Northern (RNA) blot analysis of total RNA from several brain regions showed high levels of preproenkephalin mRNA in the caudate putamen, nucleus accumbens, and hypothalamus, with detectable levels in the amygdala, ventral tegmental area, and central gray and also in the pituitary. Unexpectedly, in several brain regions, the mRNA appeared not only in the 1,400-nucleotide length but also in a shorter length of approximately 1,130 bases. Significant amounts of the shorter mRNA were found in the caudate putamen, nucleus accumbens, and amygdala. The longer, but not the shorter, transcripts from the caudate putamen were found to be polyadenylated, but the difference in size was not due solely to the presence of poly(A) tails. Northern gel analysis of total RNA from the caudate putamen with probes from each exon, together with RNase protection mapping of the 3' end of the mRNA demonstrated that the 1,400-base preproenkephalin mRNA transcripts are cleaved in a site-specific manner in some brain regions, yielding a 1,130-base transcript and a 165-base polyadenylated fragment derived from the terminal end of the 3' untranslated region of the mRNA. This cleavage may serve as a preliminary step in RNA degradation and provide a mechanism for control of preproenkephalin mRNA abundance through selective degradation. PMID:7891703

  18. Dis3- and exosome subunit-responsive 3 Prime mRNA instability elements

    SciTech Connect

    Kiss, Daniel L.; Hou, Dezhi; Gross, Robert H.; Andrulis, Erik D.

    2012-07-06

    Highlights: Black-Right-Pointing-Pointer Successful use of a novel RNA-specific bioinformatic tool, RNA SCOPE. Black-Right-Pointing-Pointer Identified novel 3 Prime UTR cis-acting element that destabilizes a reporter mRNA. Black-Right-Pointing-Pointer Show exosome subunits are required for cis-acting element-mediated mRNA instability. Black-Right-Pointing-Pointer Define precise sequence requirements of novel cis-acting element. Black-Right-Pointing-Pointer Show that microarray-defined exosome subunit-regulated mRNAs have novel element. -- Abstract: Eukaryotic RNA turnover is regulated in part by the exosome, a nuclear and cytoplasmic complex of ribonucleases (RNases) and RNA-binding proteins. The major RNase of the complex is thought to be Dis3, a multi-functional 3 Prime -5 Prime exoribonuclease and endoribonuclease. Although it is known that Dis3 and core exosome subunits are recruited to transcriptionally active genes and to messenger RNA (mRNA) substrates, this recruitment is thought to occur indirectly. We sought to discover cis-acting elements that recruit Dis3 or other exosome subunits. Using a bioinformatic tool called RNA SCOPE to screen the 3 Prime untranslated regions of up-regulated transcripts from our published Dis3 depletion-derived transcriptomic data set, we identified several motifs as candidate instability elements. Secondary screening using a luciferase reporter system revealed that one cassette-harboring four elements-destabilized the reporter transcript. RNAi-based depletion of Dis3, Rrp6, Rrp4, Rrp40, or Rrp46 diminished the efficacy of cassette-mediated destabilization. Truncation analysis of the cassette showed that two exosome subunit-sensitive elements (ESSEs) destabilized the reporter. Point-directed mutagenesis of ESSE abrogated the destabilization effect. An examination of the transcriptomic data from exosome subunit depletion-based microarrays revealed that mRNAs with ESSEs are found in every up-regulated mRNA data set but are

  19. Regulated expression of nuclear protein(s) in myogenic cells that binds to a conserved 3' untranslated region in pro alpha 1 (I) collagen cDNA.

    PubMed Central

    Herget, T; Burba, M; Schmoll, M; Zimmermann, K; Starzinski-Powitz, A

    1989-01-01

    We describe the identification and DNA-binding properties of nuclear proteins from rat L6 myoblasts which recognize an interspecies conserved 3' untranslated segment of pro alpha 1 (I) collagen cDNA. Levels of the two pro alpha 1 (I) collagen RNAs, present in L6 myoblasts, decreased drastically between 54 and 75 h after induction of myotube formation in serum-free medium. Both mRNAs contained a conserved sequence segment of 135 nucleotides (termed tame sequence) in the 3' untranslated region that had 96% homology to the human and murine pro alpha 1 (I) collagen genes. The cDNA of this tame sequence was specifically recognized by nuclear protein(s) from L6 myoblasts, as judged by gel retardation assays and DNase I footprints. The tame-binding protein(s) was able to recognize its target sequence on double-stranded DNA but bound also to the appropriate single-stranded oligonucleotide. Protein that bound to the tame sequence was undetectable in nuclear extracts of L6 myotubes that did not accumulate the two collagen mRNAs. Therefore, the activity of this nuclear protein seems to be linked to accumulation of the sequences that it recognizes in vitro. The collagen RNAs and the nuclear tame-binding proteins reappeared after a change of medium, which further suggests that the RNAs and the protein(s) are coordinately regulated. Images PMID:2779548

  20. High genetic diversity in the coat protein and 3 untranslated regions among geographical isolates of Cardamom mosaic virus from south India.

    PubMed

    Jacob, T; Jebasingh, T; Venugopal, M N; Usha, R

    2003-09-01

    A survey was conducted to study the biological and genetic diversity of Cardamom mosaic virus (CdMV) that causes the most widespread disease in the cardamom growing area in the Western Ghats of south India. Six distinct subgroups were derived based on their symptomatology and host range from the sixty isolates collected. The serological variability between the virus isolates was analysed by ELISA and Western blotting. The 3 terminal region consisting of the coat protein (CP) coding sequence and 3 untranslated region (3 UTR) was cloned and sequenced from seven isolates. Sequence comparisons revealed considerable genetic diversity among the isolates in their CP and 3 UTR, making CdMV one of the highly variable members of Potyviridae. The possible occurrence of recombination between the isolates and the movement of the virus in the cardamom tract of south India are discussed.

  1. miR-155* mediates suppressive effect of PTEN 3'-untranslated region on AP-1/NF-κB pathway in HTR-8/SVneo cells.

    PubMed

    Xue, P; Zheng, M; Diao, Z; Shen, L; Liu, M; Gong, P; Sun, H; Hu, Y

    2013-08-01

    Among miRNAs, miR-155 is a known regulator of immune system. Accumulating studies have revealed the connections between miR-155 and activator protein 1 (AP-1)/nuclear factor (NF)-κB. However, miR-155*, a miR-155 paralog, has so far been less studied. Here we demonstrated that miR-155*, induced by lipopolysaccharide (LPS) in an AP-1/NF-κB dependent manner, played a positive feedback role in AP-1/NF-κB pathway via targeting interleukin-1 receptor-associated kinase M (IRAKM) and NF-κB inhibitor interacting Ras-like 1 (NKIRAS1) in trophoblasts. Our study further proved that miR-155*-targeted PTEN 3'-untranslated region (3'UTR) increased IRAKM and NKIRAS1 expression by competing for miR-155* binding, thereby suppressing AP-1/NF-κB activation induced by LPS.

  2. GB Virus C/Hepatitis G Virus Groups and Subgroups: Classification by a Restriction Fragment Length Polymorphism Method Based on Phylogenetic Analysis of the 5′ Untranslated Region

    PubMed Central

    Quarleri, J. F.; Mathet, V. L.; Feld, M.; Ferrario, D.; della Latta, M. P.; Verdun, R.; Sánchez, D. O.; Oubiña, J. R.

    1999-01-01

    A phylogenetic tree based on 150 5′ untranslated region sequences deposited in GenBank database allowed segregation of the sequences into three major groups, including two subgroups, i.e., 1, 2a, 2b, and 3, supported by bootstrap analysis. Restriction site analysis of these sequences predicted that HinfI and either AatII or AciI could be used for genomic typing with 99.4% accuracy. cDNA sequencing and subsequent alignment of 21 Argentine GB virus C/hepatitis G virus strains confirmed restriction fragment length polymorphism patterns theoretically predicted. This method may be useful for a rapid screening of samples when either epidemiological or transmission studies of this agent are carried out. PMID:10203483

  3. Linkage disequilibrium between polymorphisms at the 5{prime} untranslated region and intron 5 (Dde I) of the antithrombin III (ATIII) gene in the Chinese

    SciTech Connect

    Tay, J.S.H.; Liu, Y.; Low, P.S.

    1994-09-01

    A length polymorphism at the 5{prime} untranslated region of exon 1 and an RFLP (Dde I) in intron 5 (nt 160) of the ATIII gene were amplified by polymerase chain reaction with primers of published sequences. DNA fragments were size-fractionated by agarose gel electrophoresis (3% NuSieve and 1% Seakem GTG) and photographed over a UV transilluminator. A strong linkage disequilibrium was observed between these two polymorphisms of the ATIII gene in the Chinese ({chi}{sup 2} = 63.7; {triangle} 0.42, P < 0.001). The estimated frequencies of the three haplotypes were found to be 0.37 for SD+, 0.40 for LD+ and 0.23 for LD-.

  4. The 5' untranslated region of the rbp1 mRNA is required for translation of its mRNA under low temperatures in the cyanobacterium Synechococcus elongatus.

    PubMed

    Hayashi, Rie; Sugita, Chieko; Sugita, Mamoru

    2017-01-01

    The unicellular cyanobacterium Synechococcus elongatus has three RNA-binding protein (Rbp) genes, rbp1, rbp2 and rbp3. The rbp1 gene was upregulated by cold treatment while rbp2 and rbp3 expression decreased remarkably after exposure to cold temperatures. To investigate the mechanism underlying cold-induced rbp1 expression, a series of rbp1-luxAB transcriptional fusion constructs were expressed in S. elongatus PCC 7942 under cold conditions. The results showed that the region from -33 to -3 of the transcription initiation site contains an essential sequence for basal transcription of the rbp1 gene and that the 120-bp region (-34 to -153) does not contain critical cis-elements required for cold-shock induction. In contrast, mutational analysis carrying the 5'-untranslated region (UTR) of rbp1-luxAB translational fusions indicated that the 5'-UTR of rbp1 plays an important role in cold induction of the rbp1 gene product. Taken together, we conclude that the cold induction of rbp1 may be regulated at a posttranscriptional level rather than at the transcriptional level.

  5. Molecular archaeology of Flaviviridae untranslated regions: duplicated RNA structures in the replication enhancer of flaviviruses and pestiviruses emerged via convergent evolution.

    PubMed

    Gritsun, Dmitri J; Jones, Ian M; Gould, Ernest A; Gritsun, Tamara S

    2014-01-01

    RNA secondary structures in the 3'untranslated regions (3'UTR) of the viruses of the family Flaviviridae, previously identified as essential (promoters) or beneficial (enhancers) for replication, have been analysed. Duplicated enhancer elements are revealed as a global feature in the evolution of the 3'UTR of distantly related viruses within the genera Flavivirus and Pestivirus. For the flaviviruses, duplicated structures occur in the 3'UTR of all four distantly related ecological virus subgroups (tick-borne, mosquito-borne, no known vector and insect-specific flaviviruses (ISFV). RNA structural differences distinguish tick-borne flaviviruses with discrete pathogenetic characteristics. For Aedes- and Culex-associated ISFV, secondary RNA structures with different conformations display numerous short ssRNA direct repeats, exposed as loops and bulges. Long quadruplicate regions comprise almost the entire 3'UTR of Culex-associated ISFV. Extended duplicated sequence and associated RNA structures were also discovered in the 3'UTR of pestiviruses. In both the Flavivirus and Pestivirus genera, duplicated RNA structures were localized to the enhancer regions of the 3'UTR suggesting an adaptive role predominantly in wild-type viruses. We propose sequence reiteration might act as a scaffold for dimerization of proteins involved in assembly of viral replicase complexes. Numerous nucleotide repeats exposed as loops/bulges might also interfere with host immune responses acting as a molecular sponge to sequester key host proteins or microRNAs.

  6. Genome-wide association identifies a deletion in the 3' untranslated region of striatin in a canine model of arrhythmogenic right ventricular cardiomyopathy.

    PubMed

    Meurs, Kathryn M; Mauceli, Evan; Lahmers, Sunshine; Acland, Gregory M; White, Stephen N; Lindblad-Toh, Kerstin

    2010-09-01

    Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a familial cardiac disease characterized by ventricular arrhythmias and sudden cardiac death. It is most frequently inherited as an autosomal dominant trait with incomplete and age-related penetrance and variable clinical expression. The human disease is most commonly associated with a causative mutation in one of several genes encoding desmosomal proteins. We have previously described a spontaneous canine model of ARVC in the boxer dog. We phenotyped adult boxer dogs for ARVC by performing physical examination, echocardiogram and ambulatory electrocardiogram. Genome-wide association using the canine 50k SNP array identified several regions of association, of which the strongest resided on chromosome 17. Fine mapping and direct DNA sequencing identified an 8-bp deletion in the 3' untranslated region (UTR) of the Striatin gene on chromosome 17 in association with ARVC in the boxer dog. Evaluation of the secondary structure of the 3' UTR demonstrated that the deletion affects a stem loop structure of the mRNA and expression analysis identified a reduction in Striatin mRNA. Dogs that were homozygous for the deletion had a more severe form of disease based on a significantly higher number of ventricular premature complexes. Immunofluorescence studies localized Striatin to the intercalated disc region of the cardiac myocyte and co-localized it to three desmosomal proteins, Plakophilin-2, Plakoglobin and Desmoplakin, all involved in the pathogenesis of ARVC in human beings. We suggest that Striatin may serve as a novel candidate gene for human ARVC.

  7. Molecular Archaeology of Flaviviridae Untranslated Regions: Duplicated RNA Structures in the Replication Enhancer of Flaviviruses and Pestiviruses Emerged via Convergent Evolution

    PubMed Central

    Gritsun, Dmitri J.; Jones, Ian M.; Gould, Ernest A.; Gritsun, Tamara S.

    2014-01-01

    RNA secondary structures in the 3′untranslated regions (3′UTR) of the viruses of the family Flaviviridae, previously identified as essential (promoters) or beneficial (enhancers) for replication, have been analysed. Duplicated enhancer elements are revealed as a global feature in the evolution of the 3′UTR of distantly related viruses within the genera Flavivirus and Pestivirus. For the flaviviruses, duplicated structures occur in the 3′UTR of all four distantly related ecological virus subgroups (tick-borne, mosquito-borne, no known vector and insect-specific flaviviruses (ISFV). RNA structural differences distinguish tick-borne flaviviruses with discrete pathogenetic characteristics. For Aedes- and Culex-associated ISFV, secondary RNA structures with different conformations display numerous short ssRNA direct repeats, exposed as loops and bulges. Long quadruplicate regions comprise almost the entire 3′UTR of Culex-associated ISFV. Extended duplicated sequence and associated RNA structures were also discovered in the 3′UTR of pestiviruses. In both the Flavivirus and Pestivirus genera, duplicated RNA structures were localized to the enhancer regions of the 3′UTR suggesting an adaptive role predominantly in wild-type viruses. We propose sequence reiteration might act as a scaffold for dimerization of proteins involved in assembly of viral replicase complexes. Numerous nucleotide repeats exposed as loops/bulges might also interfere with host immune responses acting as a molecular sponge to sequester key host proteins or microRNAs. PMID:24647143

  8. Complex Effects of Deletions in the 5′ Untranslated Region of Primate Foamy Virus on Viral Gene Expression and RNA Packaging

    PubMed Central

    Heinkelein, Martin; Thurow, Jana; Dressler, Marco; Imrich, Horst; Neumann-Haefelin, Dieter; McClure, Myra O.; Rethwilm, Axel

    2000-01-01

    Due to various advantageous features there is current interest in retroviral vectors derived from primate foamy viruses (PFVs). Two PFV cis-acting sequences have been mapped in the 5′ region of the RNA (pre-)genome and in the 3′ pol genomic region. In order to genetically separate PFV packaging constructs from vector constructs, we investigated the effect of deletions in the 5′ untranslated region (UTR) of PFV packaging constructs and vectors on gene expression and RNA incorporation into viral particles. Our results indicate that the 5′ UTR serves different previously unknown functions. First, the R region of the long terminal repeat was found to be required for PFV gag gene expression. This regulation of gene expression appeared to be mainly posttranscriptional. Second, constructs with sequence deletions between the R region and the gag gene start codon packaged as much viral mRNA into particles as the undeleted construct, and RNA from such a 5′-UTR-deleted packaging construct was copackaged into vector-virus particles, together with vector RNA which was preferentialy packaged. Finally, in the U5 region a sequence was identified that was required to allow cleavage of the Gag precursor protein by the pol gene-encoded protease, suggesting a role of RNA in PFV particle formation. Taken together, the results indicate that complex interactions of the viral RNA, capsid, and polymerase proteins take place during PFV particle formation and that a clear separation of PFV vector and packaging construct sequences may be difficult to achieve. PMID:10708430

  9. Functional equivalence of common and unique sequences in the 3' untranslated regions of alfalfa mosaic virus RNAs 1, 2, and 3.

    PubMed Central

    van Rossum, C M; Brederode, F T; Neeleman, L; Bol, J F

    1997-01-01

    The 3' untranslated regions (UTRs) of alfalfa mosaic virus (AMV) RNAs 1, 2, and 3 consist of a common 3'-terminal sequence of 145 nucleotides (nt) and upstream sequences of 18 to 34 nt that are unique for each RNA. The common sequence can be folded into five stem-loop structures, A to E, despite the occurrence of 22 nt differences between the three RNAs in this region. Exchange of the common sequences or full-length UTRs between the three genomic RNAs did not affect the replication of these RNAs in vivo, indicating that the UTRs are functionally equivalent. Mutations that disturbed base pairing in the stem of hairpin E reduced or abolished RNA replication, whereas compensating mutations restored RNA replication. In vitro, the 3' UTRs of the three RNAs were recognized with similar efficiencies by the AMV RNA-dependent RNA polymerase (RdRp). A deletion analysis of template RNAs indicated that a 3'-terminal sequence of 127 nt in each of the three AMV RNAs was not sufficient for recognition by the RdRp. Previously, it has been shown that this 127-nt sequence is sufficient for coat protein binding. Apparently, sequences required for recognition of AMV RNAs by the RdRp are longer than sequences required for CP binding. PMID:9094656

  10. Interactome analysis of the EV71 5' untranslated region in differentiated neuronal cells SH-SY5Y and regulatory role of FBP3 in viral replication.

    PubMed

    Huang, Hsing-I; Chang, Ying-Ying; Lin, Jhao-Yin; Kuo, Rei-Lin; Liu, Hao-Ping; Shih, Shin-Ru; Wu, Chih-Ching

    2016-09-01

    Enterovirus 71 (EV71), a single-stranded RNA virus, is one of the most serious neurotropic pathogens in the Asia-Pacific region. Through interactions with host proteins, the 5' untranslated region (5'UTR) of EV71 is important for viral replication. To gain a protein profile that interact with the EV71 5'UTR in neuronal cells, we performed a biotinylated RNA-protein pull-down assay in conjunction with LC-MS/MS analysis. A total of 109 proteins were detected and subjected to Database for Annotation, Visualization and Integrated Discovery (DAVID) analyses. These proteins were found to be highly correlated with biological processes including RNA processing/splicing, epidermal cell differentiation, and protein folding. A protein-protein interaction network was constructed using the STRING online database to illustrate the interactions of those proteins that are mainly involved in RNA processing/splicing or protein folding. Moreover, we confirmed that the far-upstream element binding protein 3 (FBP3) was able to bind to the EV71 5'UTR. The redistribution of FBP3 in subcellular compartments was observed after EV71 infection, and the decreased expression of FBP3 in host neuronal cells markedly inhibited viral replication. Our results reveal various host proteins that potentially interact with the EV71 5'UTR in neuronal cells, and we found that FBP3 could serve as a positive regulator in host cells.

  11. Correlation of the 5'untranslated region (5'UTR) and non-structural 5B (NS5B) nucleotide sequences in hepatitis C virus subtyping.

    PubMed

    Baclig, Michael O; Chan, Veronica F; Ramos, John Donnie A; Gopez-Cervantes, Juliet; Natividad, Filipinas F

    2010-07-07

    The 5'untranslated region (5'UTR) is often targeted to detect major genotypes in hepatitis C virus (HCV) but its insufficient sequence variation limits its usefulness for differentiating HCV subtypes. Subtyping has important implications to epidemiologic studies, clinical management, and vaccine development. Analysis of the nucleotide sequence of variable regions such as the non-structural 5B (NS5B) is considered the reference method for identifying HCV subtypes. We evaluated the accuracy of subtyping of HCV genotype 1 (HCV-1) samples from the Philippines by 5'UTR sequencing as compared with the NS5B sequence. A total of 30 patients infected with HCV-1 previously confirmed by PCR-RFLP and clinically diagnosed with chronic hepatitis C were analyzed. Nucleotide sequencing of the 5'UTR showed that 15 (50%) were identified as 1a and 15 (50%) were identified as 1b. Sequence analysis of the NS5B revealed that 13 (43%) belonged to subtype 1a while 17 (57%) belonged to subtype 1b. The most predominant subtype was 1b by NS5B sequencing. The predictive value of 5'UTR sequencing to subtype 1a was 73% while for subtype 1b, predictive value was 87%. Overall concordance between 5'UTR and NS5B sequencing was 80%. NS5B sequence and phylogenetic analysis is still the reference method for identifying HCV-1a and 1b subtypes.

  12. Correlation of the 5′untranslated region (5′UTR) and non-structural 5B (NS5B) nucleotide sequences in hepatitis C virus subtyping

    PubMed Central

    Baclig, Michael O; Chan, Veronica F; Ramos, John Donnie A; Gopez-Cervantes, Juliet; Natividad, Filipinas F

    2010-01-01

    The 5′untranslated region (5′UTR) is often targeted to detect major genotypes in hepatitis C virus (HCV) but its insufficient sequence variation limits its usefulness for differentiating HCV subtypes. Subtyping has important implications to epidemiologic studies, clinical management, and vaccine development. Analysis of the nucleotide sequence of variable regions such as the non-structural 5B (NS5B) is considered the reference method for identifying HCV subtypes. We evaluated the accuracy of subtyping of HCV genotype 1 (HCV-1) samples from the Philippines by 5′UTR sequencing as compared with the NS5B sequence. A total of 30 patients infected with HCV-1 previously confirmed by PCR-RFLP and clinically diagnosed with chronic hepatitis C were analyzed. Nucleotide sequencing of the 5′UTR showed that 15 (50%) were identified as 1a and 15 (50%) were identified as 1b. Sequence analysis of the NS5B revealed that 13 (43%) belonged to subtype 1a while 17 (57%) belonged to subtype 1b. The most predominant subtype was 1b by NS5B sequencing. The predictive value of 5′UTR sequencing to subtype 1a was 73% while for subtype 1b, predictive value was 87%. Overall concordance between 5′UTR and NS5B sequencing was 80%. NS5B sequence and phylogenetic analysis is still the reference method for identifying HCV-1a and 1b subtypes. PMID:21537395

  13. Haplotypes of the HLA-G 3’ Untranslated Region Respond to Endogenous Factors of HLA-G+ and HLA-G- Cell Lines Differentially

    PubMed Central

    Cagnin, Natalia F.; Sgorla de Almeida, Bibiana; Castelli, Erick C.; Carosella, Edgardo D.; Donadi, Eduardo A.; Moreau, Philippe

    2017-01-01

    The immune checkpoint HLA-G prevents maternal rejection of the fetus and contributes in cancer invasion and acceptance of allografts. The 5’ and 3’ regulatory regions of the HLA-G gene are polymorphic and balancing selection probably maintains this variability. It is proposed that nucleotide variations may affect the level of HLA-G expression. To investigate this issue we aimed to analyze how haplotypes of the 3’ untranslated region (3’UTR) with highest worldwide frequencies, namely UTR-1, UTR-2, UTR-3, UTR-4, UTR-5, UTR-18 and UTR-7, impact the expression of a luciferase reporter gene in vitro. Experiments performed with the HLA-G positive cell lines JEG-3 (choricarcinoma) and FON (melanoma), and with the HLA-G negative cell lines M8 (melanoma) and U251MG (glioblastoma) showed that the HLA-G 3’UTR polymorphism influences the response to endogenous cellular factors and may vary according to the cell type. UTR-5 and UTR-7 impact the activity of luciferase the most whereas UTR-2, UTR-3, UTR-4, and UTR-18 have intermediate impact, and UTR-1 has the lowest impact. These results corroborate the previous associations between amounts of plasma sHLA-G levels and 3’UTR haplotypes in healthy individuals and reinforce that 3’UTR typing may be a predictor of the genetic predisposition of an individual to express different levels of HLA-G. PMID:28045999

  14. Evaluation of a new-generation line-probe assay that detects 5' untranslated and core regions to genotype and subtype hepatitis C virus.

    PubMed

    Nadarajah, Rohan; Khan, G Yasmeen; Miller, Steven A; Brooks, Geo F

    2007-08-01

    The VERSANT HCV Genotype 2.0 Assay (LiPA 2.0; Innogenetics, Ghent, Belgium; distributed by Siemens Medical Solutions Diagnostics, Tarrytown, NY) is a new-generation line-probe assay that simultaneously detects sequences in the 5' untranslated (5'UTR) and core regions to genotype and subtype hepatitis C virus (HCV). We tested 60 specimens of known genotype and subtype and 2 specimens with mixed infections with the LiPA 2.0 assay. After arbitration based on genotype and subtype determined by sequencing, there was concordance in 58 of 60 specimens (specificity, 96.7%). Computer-assisted typing yielded comparable results, but much more rapidly. Of 67 clinical specimens, 64 readily yielded genotype and subtype; 3 indeterminate specimens were typed by sequencing and were uncommon types not in the database. The newgeneration line-probe assay that detects the 5'UTR and core regions to genotype and subtype HCV is applicable to more than 95% of specimens. Interpretation is facilitated by computer-assisted analysis.

  15. Haplotypes of the HLA-G 3' Untranslated Region Respond to Endogenous Factors of HLA-G+ and HLA-G- Cell Lines Differentially.

    PubMed

    Poras, Isabelle; Yaghi, Layale; Martelli-Palomino, Gustavo; Mendes-Junior, Celso T; Muniz, Yara Costa Netto; Cagnin, Natalia F; Sgorla de Almeida, Bibiana; Castelli, Erick C; Carosella, Edgardo D; Donadi, Eduardo A; Moreau, Philippe

    2017-01-01

    The immune checkpoint HLA-G prevents maternal rejection of the fetus and contributes in cancer invasion and acceptance of allografts. The 5' and 3' regulatory regions of the HLA-G gene are polymorphic and balancing selection probably maintains this variability. It is proposed that nucleotide variations may affect the level of HLA-G expression. To investigate this issue we aimed to analyze how haplotypes of the 3' untranslated region (3'UTR) with highest worldwide frequencies, namely UTR-1, UTR-2, UTR-3, UTR-4, UTR-5, UTR-18 and UTR-7, impact the expression of a luciferase reporter gene in vitro. Experiments performed with the HLA-G positive cell lines JEG-3 (choricarcinoma) and FON (melanoma), and with the HLA-G negative cell lines M8 (melanoma) and U251MG (glioblastoma) showed that the HLA-G 3'UTR polymorphism influences the response to endogenous cellular factors and may vary according to the cell type. UTR-5 and UTR-7 impact the activity of luciferase the most whereas UTR-2, UTR-3, UTR-4, and UTR-18 have intermediate impact, and UTR-1 has the lowest impact. These results corroborate the previous associations between amounts of plasma sHLA-G levels and 3'UTR haplotypes in healthy individuals and reinforce that 3'UTR typing may be a predictor of the genetic predisposition of an individual to express different levels of HLA-G.

  16. A Single Nucleotide in Stem Loop II of 5′-Untranslated Region Contributes to Virulence of Enterovirus 71 in Mice

    PubMed Central

    Yeh, Ming-Te; Wang, Shainn-Wei; Yu, Chun-Keung; Lin, Kuei-Hsiang; Lei, Huan-Yao; Su, Ih-Jen; Wang, Jen-Ren

    2011-01-01

    Background Enterovirus 71 (EV71) has emerged as a neuroinvasive virus responsible for several large outbreaks in the Asia-Pacific region while virulence determinant remains unexplored. Principal Findings In this report, we investigated increased virulence of unadapted EV71 clinical isolate 237 as compared with isolate 4643 in mice. A fragment 12 nucleotides in length in stem loop (SL) II of 237 5′-untranslated region (UTR) visibly reduced survival time and rate in mice was identified by constructing a series of infectious clones harboring chimeric 5′-UTR. In cells transfected with bicistronic plasmids, and replicon RNAs, the 12-nt fragment of isolate 237 enhanced translational activities and accelerated replication of subgenomic EV71. Finally, single nucleotide change from cytosine to uridine at base 158 in this short fragment of 5′-UTR was proven to reduce viral translation and EV71 virulence in mice. Results collectively indicated a pivotal role of novel virulence determinant C158 on virus translation in vitro and EV71 virulence in vivo. Conclusions These results presented the first reported virulence determinant in EV71 5′-UTR and first position discovered from unadapted isolates. PMID:22069490

  17. Characterization of the partial RNA1 and RNA2 3' untranslated region of tomato ringspot virus isolates from North America

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The 3' non-translated regions (NTRs) of RNA1 and RNA2 of Tomato ringspot virus (ToRSV) are long and virtually identical. In this study, sequences containing most of the 3’ NTRs (1168-1265 bp) were determined from 18 ToRSV isolates collected from fruit trees, small fruits, and grapevines in North Am...

  18. Potential control of human immunodeficiency virus type 1 asp expression by alternative splicing in the upstream untranslated region.

    PubMed

    Barbagallo, Michael S; Birch, Katherine E; Deacon, Nicholas J; Mosse, Jennifer A

    2012-07-01

    The negative-sense asp open reading frame (ORF) positioned opposite to the human immunodeficiency virus type 1 (HIV-1) env gene encodes the 189 amino acid, membrane-associated ASP protein. Negative-sense transcription, regulated by long terminal repeat sequences, has been observed early in HIV-1 infection in vitro. All subtypes of HIV-1 were scanned to detect the negative-sense asp ORF and to identify potential regulatory sequences. A series of highly conserved upstream short open reading frames (sORFs) was identified. This potential control region from HIV-1(NL4-3), containing six sORFs, was cloned upstream of the reporter gene EGFP. Expression by transfection of HEK293 cells indicated that the introduction of this sORF region inhibits EGFP reporter expression; analysis of transcripts revealed no significant changes in levels of EGFP mRNA. Reverse transcriptase-polymerase chain reaction analysis (RT-PCR) further demonstrated that the upstream sORF region undergoes alternative splicing in vitro. The most abundant product is spliced to remove sORFs I to V, leaving only the in-frame sORF VI upstream of asp. Sequence analysis revealed the presence of typical splice donor- and acceptor-site motifs. Mutation of the highly conserved splice donor and acceptor sites modulates, but does not fully relieve, inhibition of EGFP production. The strong conservation of asp and its sORFs across all HIV-1 subtypes suggests that the asp gene product may have a role in the pathogenesis of HIV-1. Alternative splicing of the upstream sORF region provides a potential mechanism for controlling expression of the asp gene.

  19. Unorthodox expression of an enzyme: evidence for an untranslated region within carA from Pseudomonas aeruginosa.

    PubMed Central

    Wong, S C; Abdelal, A T

    1990-01-01

    The genes encoding carbamoylphosphate synthetase from Pseudomonas aeruginosa PAO1 were cloned in Escherichia coli. Deletion and transposition analysis determined the locations of carA, encoding the small subunit, and carB, encoding the large subunit, on the chromosomal insert. The nucleotide sequence of carA and the flanking regions was determined. The derived amino acid sequence for the small subunit of carbamoylphosphate synthetase from P. aeruginosa exhibited 68% homology with its counterparts in E. coli and Salmonella typhimurium. The derived sequences in the three organisms were essentially identical in the three polypeptide segments that are conserved in glutamine amidotransferases but showed low homology at the amino- and carboxy-terminal regions. The amino-terminal amino acid sequences were determined for the large and small subunits. The first 15 amino acids of the large subunit were identical to those derived from the carB sequence. However, comparison of the derived sequence for carA with the amino-terminal amino acid sequence for the small subunit suggested that codons 5 to 8 are not translated. The DNA sequence for the region encompassing these four codons was confirmed by direct sequencing of chromosomal DNA after amplification by the polymerase chain reaction. The mRNA sequence was also deduced by in vitro synthesis of cDNA, enzymatic amplification, and sequencing, confirming that 12 nucleotides in the 5' terminal of carA are transcribed but are not translated. Images FIG. 2 FIG. 3 FIG. 7 FIG. 8 PMID:2153657

  20. Polymorphisms at the 3' untranslated region of SLC11A1 gene are associated with protection to Brucella infection in goats.

    PubMed

    Iacoboni, Paola A; Hasenauer, Flavia C; Caffaro, M Eugenia; Gaido, Analia; Rossetto, Cristina; Neumann, Roberto D; Salatin, Antonio; Bertoni, Emiliano; Poli, Mario A; Rossetti, Carlos A

    2014-08-15

    Goats are susceptible to brucellosis and the detection of Brucella-infected animals is carried out by serological tests. In other ruminant species, polymorphisms in microsatellites (Ms) of 3' untranslated region (3'UTR) of the solute carrier family 11 member A1 (SLC11A1) gene were associated with resistance to Brucella abortus infection. Goats present two polymorphic Ms at the 3'UTR end of SLC11A1 gene, called regions A and B. Here, we evaluated if polymorphisms in regions A and/or B are associated with Brucella infection in goats. Serum (for the detection of Brucella-specific antibodies) and hair samples (for DNA isolation and structure analysis of the SLC11A1 gene) were randomly collected from 229 adult native goats from the northwest of Argentina. Serological status was evaluated by buffer plate antigen test (BPAT) complemented by the fluorescent polarization assay (FPA), and the genotype of the 3'UTR of the SLC11A1 gene was determined by capillary electrophoresis and confirmed by sequence analysis. Polymorphisms in regions A and B of the 3'UTR SLC11A1 gene were found statistically significant associated with protection to Brucella infection. Specifically, the association study indicates statistical significance of the allele A15 and B7/B7 genotype with absence of Brucella-specific antibodies (p=0.0003 and 0.0088, respectively). These data open a promising opportunity for limiting goat brucellosis through selective breeding of animals based on genetic markers associated with natural resistance to B. melitensis infection.

  1. Endothelin-1 expression is strongly repressed by AU-rich elements in the 3′-untranslated region of the gene

    PubMed Central

    2004-01-01

    The regulation of the synthesis of the endothelial-derived vasoconstrictor ET-1 (endothelin-1) is a complex process that occurs mainly at the mRNA level. Transcription of the gene accounts for an important part of the regulation of expression, as already described for different modulators such as the cytokine TGF-β (transforming growth factor-β). However, very little is known about mechanisms governing ET-1 expression at the post-transcriptional level. The aim of the present study was to investigate the regulation of the ET-1 expression at this level. Since the 3′-UTR (3′-untranslated region) of mRNAs commonly contains genetic determinants for the post-transcriptional control of gene expression, we focused on the potential role of the 3′-UTR of ET-1 mRNA. Experiments performed with luciferase reporter constructs containing the 3′-UTR showed that this region exerts a potent destabilizing effect. Deletional analyses allowed us to locate this activity within a region at positions 924–1127. Some (but not all) of the AREs (AU-rich elements) present in this region were found to be essential for this mRNA-destabilizing activity. We also present evidence that cytosolic proteins from endothelial cells interact specifically with these RNA elements, and that a close correlation exists between the ability of the AREs to destabilize ET-1 mRNA and the binding of proteins to these elements. Our results are compatible with the existence of a strong repressional control of ET-1 expression mediated by destabilization of the mRNA exerted through the interaction of specific cytosolic proteins with AREs present in the 3′-UTR of the gene. PMID:15595926

  2. Ribosome binding to a 5' translational enhancer is altered in the presence of the 3' untranslated region in cap-independent translation of turnip crinkle virus.

    PubMed

    Stupina, Vera A; Yuan, Xuefeng; Meskauskas, Arturas; Dinman, Jonathan D; Simon, Anne E

    2011-05-01

    Plus-strand RNA viruses without 5' caps require noncanonical mechanisms for ribosome recruitment. A translational enhancer in the 3' untranslated region (UTR) of Turnip crinkle virus (TCV) contains an internal T-shaped structure (TSS) that binds to 60S ribosomal subunits. We now report that the 63-nucleotide (nt) 5' UTR of TCV contains a 19-nt pyrimidine-rich element near the initiation codon that supports translation of an internal open reading frame (ORF) independent of upstream 5' UTR sequences. Addition of 80S ribosomes to the 5' UTR reduced the flexibility of the polypyrimidine residues and generated a toeprint consistent with binding to this region. Binding of salt-washed 40S ribosomal subunits was reduced 6-fold when the pyrimidine-rich sequence was mutated. 40S subunit binding generated the same toeprint as 80S ribosomes but also additional ones near the 5' end. Generation of out-of-frame AUGs upstream of the polypyrimidine region reduced translation, which suggests that 5'-terminal entry of 40S subunits is followed by scanning and that the polypyrimidine region is needed for an alternative function that requires ribosome binding. No evidence for RNA-RNA interactions between 5' and 3' sequences was found, suggesting that TCV utilizes an alternative means for circularizing its genome. Combining 5' and 3' UTR fragments in vitro had no discernible effect on the structures of the RNAs. In contrast, when 80S ribosomes were added to both fragments, structural changes were found in the 5' UTR polypyrimidine tract that were not evident when ribosomes interacted with the individual fragments. This suggests that ribosomes can promote an interaction between the 5' and 3' UTRs of TCV.

  3. Targeting deoxyhypusine hydroxylase activity impairs cap-independent translation initiation driven by the 5'untranslated region of the HIV-1, HTLV-1, and MMTV mRNAs.

    PubMed

    Cáceres, C Joaquín; Angulo, Jenniffer; Contreras, Nataly; Pino, Karla; Vera-Otarola, Jorge; López-Lastra, Marcelo

    2016-10-01

    Replication of the human immunodeficiency virus type 1 (HIV-1) is dependent on eIF5A hypusination. Hypusine is formed post-translationally on the eIF5A precursor by two consecutive enzymatic steps; a reversible reaction involving the enzyme deoxyhypusine synthase (DHS) and an irreversible step involving the enzyme deoxyhypusine hydroxylase (DOHH). In this study we explored the effect of inhibiting DOHH activity and therefore eIF5A hypusination, on HIV-1 gene expression. Results show that the expression of proteins from an HIV-1 molecular clone is reduced when DOHH activity is inhibited by Deferiprone (DFP) or Ciclopirox (CPX). Next we evaluated the requirement of DOHH activity for internal ribosome entry site (IRES)-mediated translation initiation driven by the 5'untranslated region (5'UTR) of the full length HIV-1 mRNA. Results show that HIV-1 IRES activity relies on DOHH protein concentration and enzymatic activity. Similar results were obtained for IRES-dependent translation initiation mediated by 5'UTR of the human T-cell lymphotropic virus type 1 (HTLV-1) and the mouse mammary tumor virus (MMTV) mRNAs. Interestingly, activity of the poliovirus IRES, was less sensitive to the targeting of DOHH suggesting that not all viral IRESs are equally dependent on the cellular concentration or the activity of DOHH. In summary we present evidence indicating that the cellular concentration of DOHH and its enzymatic activity play a role in HIV-1, HTLV-1 and MMTV IRES-mediated translation initiation.

  4. The 5′-untranslated region of the mouse mammary tumor virus mRNA exhibits cap-independent translation initiation

    PubMed Central

    Vallejos, Maricarmen; Ramdohr, Pablo; Valiente-Echeverría, Fernando; Tapia, Karla; Rodriguez, Felipe E.; Lowy, Fernando; Huidobro-Toro, J. Pablo; Dangerfield, John A.; López-Lastra, Marcelo

    2010-01-01

    In this study, we demonstrate the identification of an internal ribosome entry site (IRES) within the 5′-untranslated region (5′-UTR) of the mouse mammary tumor virus (MMTV). The 5′-UTR of the full-length mRNA derived from the infectious, complete MMTV genome was cloned into a dual luciferase reporter construct containing an upstream Renilla luciferase gene (RLuc) and a downstream firefly luciferase gene (FLuc). In rabbit reticulocyte lysate, the MMTV 5′-UTR was capable of driving translation of the second cistron. In vitro translational activity from the MMTV 5′-UTR was resistant to the addition of m7GpppG cap-analog and cleavage of eIF4G by foot-and-mouth disease virus (FMDV) L-protease. IRES activity was also demonstrated in the Xenopus laevis oocyte by micro-injection of capped and polyadenylated bicistronic RNAs harboring the MMTV-5′-UTR. Finally, transfection assays showed that the MMTV-IRES exhibits cell type-dependent translational activity, suggesting a requirement for as yet unidentified cellular factors for its optimal function. PMID:19889724

  5. De novo computational identification of stress-related sequence motifs and microRNA target sites in untranslated regions of a plant translatome

    PubMed Central

    Munusamy, Prabhakaran; Zolotarov, Yevgen; Meteignier, Louis-Valentin; Moffett, Peter; Strömvik, Martina V.

    2017-01-01

    Gene regulation at the transcriptional and translational level leads to diversity in phenotypes and function in organisms. Regulatory DNA or RNA sequence motifs adjacent to the gene coding sequence act as binding sites for proteins that in turn enable or disable expression of the gene. Whereas the known DNA and RNA binding proteins range in the thousands, only a few motifs have been examined. In this study, we have predicted putative regulatory motifs in groups of untranslated regions from genes regulated at the translational level in Arabidopsis thaliana under normal and stressed conditions. The test group of sequences was divided into random subgroups and subjected to three de novo motif finding algorithms (Seeder, Weeder and MEME). In addition to identifying sequence motifs, using an in silico tool we have predicted microRNA target sites in the 3′ UTRs of the translationally regulated genes, as well as identified upstream open reading frames located in the 5′ UTRs. Our bioinformatics strategy and the knowledge generated contribute to understanding gene regulation during stress, and can be applied to disease and stress resistant plant development. PMID:28276452

  6. A Polymorphism in the 5′-Untranslated Region of the Porcine Cholecystokinin Type A Receptor Gene Affects Feed Intake and Growth

    PubMed Central

    Houston, R. D.; Haley, C. S.; Archibald, A. L.; Cameron, N. D.; Plastow, G. S.; Rance, K. A.

    2006-01-01

    The location and utilization of quantitative trait loci (QTL) and candidate genes with significant effects on economically important traits are becoming increasingly important in livestock breeding programs. The porcine cholecystokinin type A receptor (CCKAR) is a candidate gene for performance traits, due to its known role in the physiological control of feed intake, satiety, and obesity. We investigated the association of CCKAR polymorphisms with feeding, growth, and efficiency traits in an F2 population derived from a cross between Meishan and Large White founder animals and in lines of Large White pigs that had been divergently selected on the basis of lean growth efficiency traits. In the F2 population, CCKAR genotype was significantly associated with daily feed intake and average daily gain. The effects of the polymorphisms were then assessed in a larger-scale analysis of segregating commercial lines. A newly discovered single-nucleotide polymorphism (SNP) within the 5′-untranslated region (5′-UTR) had highly significant effects on feed intake, average daily gain, and days to 110 kg, which were not seen for a previously reported SNP within the CCKAR gene. Furthermore, we provide evidence that the novel SNP disrupts the binding of the YY1 transcription factor, which raises the possibility that it is the causal variant. The 5′-UTR SNP could be utilized as a molecular genetic test for increased feed intake, faster lean growth, and reduced days to market weight in segregating commercial lines. PMID:16951077

  7. The Effect of Poly(ADP-ribose) Polymerase-1 Gene 3'Untranslated Region Polymorphism in Colorectal Cancer Risk among Saudi Cohort.

    PubMed

    Alhadheq, Abdullah M; Purusottapatnam Shaik, Jilani; Alamri, Abdullah; Aljebreen, Abdulrahman M; Alharbi, Othman; Almadi, Majid A; Alhadeq, Faten; Azzam, Nahla A; Semlali, Abdelhabib; Alanazi, Mohammad; Bazzi, Mohammad D; Reddy Parine, Narasimha

    2016-01-01

    Background. DNA repair systems are essential for each cell to repair and maintain the genome integrity. Base excision repair pathway is one of the crucial pathways to maintain genome integrity and PARP-1 plays a key role in BER pathway. The purpose of this study is to evaluate the association between polymorphisms in PARP-1 3'untranslated region (3'UTR) SNP rs8679 and its expression in colorectal cancer. Methods. Genotyping and gene expression were performed using TaqMan assays. The effects of age, gender, and tumor location were evaluated in cases and controls regarding the genotyping results. Resulting data was analyzed using SPSS software. Results and Conclusions. Genotyping analysis for SNP rs8679 showed decreased susceptibility to colorectal cancer at heterozygous TC allele and at minor allele C. Further this protective association was also observed in younger age patients (≤57), in female patients, and also in patients with tumors located at colon and rectum. PARP-1 expression levels are significantly different in colorectal cancer compared to matched normal tissue. Our findings proved that the upregulation of PARP-1 is associated with tumor progression and poor prognosis in Saudi patients with colorectal cancer, suggesting that PARP-1 can be novel and valuable signatures for predicting the clinical outcome of patients with colorectal cancer.

  8. Inhibition of porcine reproductive and respiratory syndrome virus replication with exosome-transferred artificial microRNA targeting the 3' untranslated region.

    PubMed

    Zhu, Li; Bao, Liping; Zhang, Xinyu; Xia, Xiaoli; Sun, Huaichang

    2015-10-01

    Porcine reproductive and respiratory syndrome (PRRS) is an economically important swine disease. As part of the development of RNA interference (RNAi) strategy against the disease, in this study a recombinant adenovirus (rAd) expressing the artificial microRNA (amiRNA) targeting the 3' untranslated region (UTR) was used to investigate the exosome-mediated amiRNA transfer from different pig cell types to porcine alveolar macrophages (PAMs). Quantitative RT-PCR showed that the sequence-specific amiRNA was expressed in and secreted via exosomes from the rAd-transduced pig kidney cell line PK-15, PAM cell line 3D4/163, kidney fibroblast cells (PFCs) and endometrial endothelial cells (PEECs) with different secretion efficiencies. Fluorescent microscopy revealed that the dye-labeled amiRNA-containing exosomes of different cell origins were efficiently taken up by all of the five types of pig cells tested, including primary PAMs. Quantitative RT-PCR showed that the amiRNA-containing exosomes of different cell origins were taken up by primary PAMs in both time- and dose-dependent manners. Both quantitative RT-PCR and viral titration assays showed that the exosome-delivered amiRNA had potent anti-viral effects against three different PRRSV strains. These data suggest that the exosomes derived from pig cells could serve as an efficient miRNA transfer vehicle, and that the exosome-delivered amiRNA had potent anti-viral effects against different PRRSV strains.

  9. Human tumor necrosis factor-alpha gene 3' untranslated region confers inducible toxin responsiveness to homologous promoter in monocytic THP-1 cells.

    PubMed

    Seiler-Tuyns, A; Dufour, N; Spertini, F

    1999-07-30

    To better define the role of 3' untranslated region (3'UTR) on transcriptional regulation of the human tumor necrosis factor (TNF)-alpha gene, monocytic human THP-1 cells were transfected with two TNF-alpha promoter constructs spanning base pairs -1897/-1 and -1214/-1, respectively, and linked to the rabbit beta-globin gene. Quantitative globin gene expression of chimerae was measured by reverse transcription-polymerase chain reaction. A construct linking the chicken beta-actin promoter and a deleted portion of the beta-globin gene was cotransfected and used as internal standard. Unexpectedly, when THP-1 cells were stimulated with lipopolysaccharide or toxic shock syndrome toxin-1, gene regulation was hardly detected. In contrast, endogenous TNF-alpha gene regulation measured by the same reverse transcription-polymerase chain reaction procedure was vigorous. Remarkably, ligation of 3'UTR to chimeric constructs led to a drastic drop in the basal level of chimeric gene expression, resulting in a 15- to 40-fold induction of the reporter gene. Consistently, when the TNF-alpha promoter was replaced by the cytomegalovirus early immediate promoter, gene expression was also uniformly reduced but was no longer up-regulated upon stimulation with lipopolysaccharide and toxic shock syndrome toxin-1. These data provide the first line of evidence that, in addition to its role in TNF-alpha transcript stability and translation, human TNF-alpha 3'UTR also participates in modulating gene expression at the transcriptional level.

  10. Efficient inhibition of human immunodeficiency virus replication using novel modified microRNA-30a targeting 3′-untranslated region transcripts

    PubMed Central

    NEJATI, AHMAD; SHAHMAHMOODI, SHOHREH; AREFIAN, EHSAN; SHOJA, ZABIHOLLAH; MARASHI, SAYED-MAHDI; TABATABAIE, HAMIDEH; MOLLAEI-KANDELOUS, YAGHOUB; SOLEIMANI, MASOUD; NATEGH, RAKHSHANDEH

    2016-01-01

    RNA interference (RNAi)-based gene therapy is currently considered to be a combinatorial anti-human immunodeficiency virus-1 (HIV-1) therapy. Although artificial polycistronic microRNAs (miRs) can reduce HIV-1 escape mutant variants, this approach may increase the risk of side effects. The present study aimed to optimize the efficiency of anti-HIV RNAi gene therapy in order to reduce the cell toxicity induced by multi-short hairpin RNA expression. An artificial miR-30a-3′-untranslated region (miR-3-UTR) obtained from a single RNA polymerase II was used to simultaneously target all viral transcripts. The results of the present study demonstrated that HIV-1 replication was significantly inhibited in the cells with the miR-3-UTR construct, suggesting that miR-3′-UTR may serve as a promising tool for RNAi-based gene therapy in the treatment of HIV-1. PMID:27168813

  11. Genetic polymorphism of 3' untranslated region of zeta-chain associated protein kinase 70 kDa in southern Taiwanese patients with rheumatoid arthritis.

    PubMed

    Chen, Shih-Yao; Liu, Ming-Fei; Wang, Chrong-Reen

    2016-03-01

    T cell activation participates in the pathogenesis of rheumatoid arthritis (RA), and the signaling molecule zeta-chain-associated protein kinase 70 kDa (ZAP-70) plays a crucial role in this process. Different mutations in the coding sequence of ZAP-70 are involved in a variety of immunological phenotypes, and recent evidence indicates that genetic variations within the 3' untranslated regions (UTR) of microRNA binding sites may affect the hybridization with target mRNAs, leading to phenotype changes with disease status. In this study, we evaluated the possible effect of ZAP-70 polymorphism as a genetic risk factor in RA by examining the single-nucleotide polymorphism in 100 patients and 100 ethnicity- and sex-matched healthy individuals from southern Taiwan. In both groups, the genotype distribution of rs2278699 in the 3' UTR was in the Hardy-Weinberg equilibrium. In RA, there were higher frequencies of the G allele (15.5 versus 8.0 %, odds ratio 2.1, P = 0.020) and significant differences in the trend of various genotypes (P = 0.024). The results suggest that genetic polymorphism in the 3' UTR of ZAP-70 is associated with RA susceptibility in southern Taiwanese.

  12. A QPCR-based reporter system to study post-transcriptional regulation via the 3' untranslated region of mRNA in Saccharomyces cerevisiae.

    PubMed

    Lind, Kristina; Norbeck, Joakim

    2009-07-01

    Post-transcriptional regulation via the 3' untranslated region (3' UTR) of mRNA is an important factor in governing eukaryotic gene expression. Achieving detailed understanding of these processes requires highly quantitative systems in which comparative studies can be performed. To this end, we have developed a plasmid reporter system for Saccharomyces cerevisiae, in which the 3' UTR can be easily replaced and modified. Accurate quantification of the tandem affinity purification tag (TAP)-reporter protein and of TAP-mRNA is achieved by immuno-QPCR and by RT-QPCR, respectively. We have used our reporter system to evaluate the consequences on gene expression from varying the 3' UTR, a problem often encountered during C-terminal tagging of proteins. It was clear that the choice of 3' UTR was a strong determinant of the reporter expression, in a manner dependent on the growth conditions used. Mutations affecting either decapping (lsm1Delta) or deadenylation (pop2Delta) were also found to affect reporter gene expression in a highly 3' UTR-dependent manner. Our results using this set-up clearly indicate that the common strategy used for C-terminal tagging, with concomitant replacement of the native 3' UTR, will very likely provide incorrect conclusions on gene expression.

  13. Transcriptional effects of a lupus-associated polymorphism in the 5´ untranslated region (UTR) of human complement receptor 2 (CR2/CD21)

    PubMed Central

    Cruickshank, Mark N.; Karimi, Mahdad; Mason, Rhonda L.; Fenwick, Emily; Mercer, Tim; Tsao, Betty P.; Boackle, Susan A.; Ulgiati, Daniela

    2012-01-01

    Systemic lupus erythematosus (SLE) is a complex autoimmune disease with a strong genetic component that determines risk. A common three single-nucleotide polymorphism (SNP) haplotype of the complement receptor 2 (CR2) gene has been associated with increased risk of SLE (Wu et al., 2007) (Douglas et al., 2009), and a less common haplotype consisting of the major allele at SNP1 and minor alleles at SNP2 and 3 confers protection (Douglas et al., 2009). SNP1 (rs3813946), which is located in the 5´ untranslated region (UTR) of the CR2 gene, altered transcriptional activity of a CR2 promoter-luciferase reporter gene construct transiently transfected into a B cell line (Wu et al., 2007) and had an independent effect in the protective haplotype (Douglas et al., 2009). In this study, we show that this SNP alters transcriptional activity in a transiently transfected non B-cell line as well as in stably transfected cell lines, supporting its relevance in vivo. Furthermore, the allele at this SNP affects chromatin accessibility of the surrounding sequence and transcription factor binding. These data confirm the effects of rs3813946 on CR2 transcription, identifying the 5´UTR to be a novel regulatory element for the CR2 gene in which variation may alter gene function and modify the development of lupus. PMID:22673213

  14. Alternative splicing within the elk-1 5' untranslated region serves to modulate initiation events downstream of the highly conserved upstream open reading frame 2.

    PubMed

    Rahim, Gwendoline; Araud, Tanguy; Jaquier-Gubler, Pascale; Curran, Joseph

    2012-05-01

    The 5' untranslated region (UTR) plays a central role in the regulation of mammalian translation initiation. Key components include RNA structure, upstream AUGs (uAUGs), upstream open reading frames (uORFs), and internal ribosome entry site elements that can interact to modulate the readout. We previously reported the characterization of two alternatively spliced 5' UTR isoforms of the human elk-1 gene. Both contain two uAUGs and a stable RNA stem-loop, but the long form (5' UTR(L)) was more repressive than the short form (5' UTR(S)) for initiation at the ELK-1 AUG. We now demonstrate that ELK-1 expression arises by a combination of leaky scanning and reinitiation, with the latter mediated by the small uORF2 conserved in both spliced isoforms. In HEK293T cells, a considerable fraction of ribosomes scans beyond the ELK-1 AUG in a reinitiation mode. These are sequestered by a series of out-of-frame AUG codons that serve to prevent access to a second in-frame AUG start site used to express short ELK-1 (sELK-1), an N-terminally truncated form of ELK-1 that has been observed only in neuronal cells. We present evidence that all these events are fine-tuned by the nature of the 5' UTR and the activity of the α subunit of eukaryotic initiation factor 2 and provide insights into the neuronal specificity of sELK-1 expression.

  15. A Viral mRNA Motif at the 3′-Untranslated Region that Confers Translatability in a Cell-Specific Manner. Implications for Virus Evolution

    PubMed Central

    Garcia-Moreno, Manuel; Sanz, Miguel Angel; Carrasco, Luis

    2016-01-01

    Sindbis virus (SINV) mRNAs contain several motifs that participate in the regulation of their translation. We have discovered a motif at the 3′ untranslated region (UTR) of viral mRNAs, constituted by three repeated sequences, which is involved in the translation of both SINV genomic and subgenomic mRNAs in insect, but not in mammalian cells. These data illustrate for the first time that an element present at the 3′-UTR confers translatability to mRNAs from an animal virus in a cell-specific manner. Sequences located at the beginning of the 5′-UTR may also regulate SINV subgenomic mRNA translation in both cell lines in a context of infection. Moreover, a replicon derived from Sleeping disease virus, an alphavirus that have no known arthropod vector for transmission, is much more efficient in insect cells when the repeated sequences from SINV are inserted at its 3′-UTR, due to the enhanced translatability of its mRNAs. Thus, these findings provide a clue to understand, at the molecular level, the evolution of alphaviruses and their host range. PMID:26755446

  16. The Effect of Poly(ADP-ribose) Polymerase-1 Gene 3′Untranslated Region Polymorphism in Colorectal Cancer Risk among Saudi Cohort

    PubMed Central

    Alhadheq, Abdullah M.; Purusottapatnam Shaik, Jilani; Alamri, Abdullah; Aljebreen, Abdulrahman M.; Alharbi, Othman; Almadi, Majid A.; Alhadeq, Faten; Azzam, Nahla A.; Alanazi, Mohammad; Bazzi, Mohammad D.

    2016-01-01

    Background. DNA repair systems are essential for each cell to repair and maintain the genome integrity. Base excision repair pathway is one of the crucial pathways to maintain genome integrity and PARP-1 plays a key role in BER pathway. The purpose of this study is to evaluate the association between polymorphisms in PARP-1 3′untranslated region (3′UTR) SNP rs8679 and its expression in colorectal cancer. Methods. Genotyping and gene expression were performed using TaqMan assays. The effects of age, gender, and tumor location were evaluated in cases and controls regarding the genotyping results. Resulting data was analyzed using SPSS software. Results and Conclusions. Genotyping analysis for SNP rs8679 showed decreased susceptibility to colorectal cancer at heterozygous TC allele and at minor allele C. Further this protective association was also observed in younger age patients (≤57), in female patients, and also in patients with tumors located at colon and rectum. PARP-1 expression levels are significantly different in colorectal cancer compared to matched normal tissue. Our findings proved that the upregulation of PARP-1 is associated with tumor progression and poor prognosis in Saudi patients with colorectal cancer, suggesting that PARP-1 can be novel and valuable signatures for predicting the clinical outcome of patients with colorectal cancer. PMID:27746584

  17. Internal initiation of translation directed by the 5'-untranslated region of the tobamovirus subgenomic RNA I(2).

    PubMed

    Skulachev, M V; Ivanov, P A; Karpova, O V; Korpela, T; Rodionova, N P; Dorokhov, Y L; Atabekov, J G

    1999-10-10

    Previously we reported that, unlike RNA of typical tobamoviruses, the translation of the coat protein (CP) gene of a crucifer-infecting tobamovirus (crTMV) in vitro occurred by an internal ribosome entry mechanism mediated by the 148-nt region that contained an internal ribosome entry site (IRES(CP,148)(CR)). The equivalent 148-nt sequence from TMV U1 RNA (U1(CP,148)(SP)) was incapable of promoting internal initiation. In the present work, we have found that the 228-nt region upstream of the movement protein (MP) gene of crTMV RNA (IRES(MP,228)(CR)) contained an IRES element that directed in vitro translation of the 3'-proximal reporter genes from chimeric dicistronic transcripts. Surprisingly, the equivalent 228-nt sequence upstream from the MP gene of TMV U1 directed translation of the downstream gene of a dicistronic transcripts as well. Consequently this sequence was termed IRES(MP,228)(U1). It was shown that IRES(MP,228)(CR), IRES(MP,228)(U1), and IRES(CP,148)(CR) could mediate expression of the 3'-proximal GUS gene from dicistronic 35S promoter-based constructs in vivo in experiments on transfection of tobacco protoplasts and particle bombardment of Nicotiana benthamiana leaves. The results indicated that an IRES element was located within the 75-nt region upstream of MP gene (IRES(MP,75)), which corresponded closely to the length of the 5'UTR of TMV subgenomic RNA (sgRNA) I(2). The RNA transcripts structurally equivalent to I(2) sgRNAs of TMV U1 and crTMV, but containing a hairpin structure (H) immediately upstream of IRES(MP,75) (HIRES(MP), (75)(CR)-MP-CP-3'UTR; HIRES(MP,75)(U1)-MP-CP-3'UTR), were able to express the MP gene in vitro. The capacity of HIRES(MP,75)(CR) sequence for mediating internal translation of the 3'-proximal GUS gene in vivo, in tobacco protoplasts, was demonstrated. We suggested that expression of the MP gene from I(2) sgRNAs might proceed via internal ribosome entry pathway mediated by IRES(MP) element contained in the 75-nt 5'UTR

  18. A high affinity HSF-1 binding site in the 5'-untranslated region of the murine tumor necrosis factor-alpha gene is a transcriptional repressor.

    PubMed

    Singh, Ishwar S; He, Ju-Ren; Calderwood, Stuart; Hasday, Jeffrey D

    2002-02-15

    Tumor necrosis factor-alpha (TNFalpha) is a pivotal early mediator of host defenses that is essential for survival in infections. We previously reported that exposing macrophages to febrile range temperatures (FRT) (38.5-40 degrees C) markedly attenuates TNFalpha expression by causing abrupt and premature cessation of transcription. We showed that this inhibitory effect of FRT is mediated by an alternatively activated repressor form of heat shock factor 1 (HSF-1) and that a fragment of the TNFalpha gene comprising a minimal 85-nucleotide (nt) proximal promoter and the 138-nt 5'-untranslated region (UTR) was sufficient for mediating this effect. In the present study we have used an electrophoretic mobility shift assay (EMSA) to identify a high affinity binding site for HSF-1 in the 5'-UTR of the TNFalpha gene and have used a chromosome immunoprecipitation assay to show that HSF-1 binds to this region of the endogenous TNFalpha gene. Mutational inactivation of this site blocks the inhibitory effect of overexpressed HSF-1 on activity of the minimal TNFalpha promoter (-85/+138) in Raw 264.7 murine macrophages, identifying this site as an HSF-1-dependent repressor. However, the same mutation fails to block repression of a full-length (-1080/+138) TNFalpha promoter construct by HSF-1 overexpression, and HSF-1 binds to upstream sequences in the regions -1080/-845, -533/-196, and -326/-39 nt in EMSA, suggesting that additional HSF-1-dependent repressor elements are present upstream of the minimal -85-nt promoter. Furthermore, although mutation of the HSF-1 binding site in the minimal TNFalpha promoter construct abrogates HSF-1-mediated repression, the same mutation fails to abrogate repression of this construct by high levels of HSF-1 overexpression or exposure to 39.5 degrees C. This suggests that HSF-1 might repress TNFalpha transcription through redundant mechanisms, some of which might not require high affinity binding of HSF-1.

  19. Role of the 3'-untranslated region of human endothelin-1 in vascular endothelial cells. Contribution to transcript lability and the cellular heat shock response.

    PubMed

    Mawji, Imtiaz A; Robb, G Brett; Tai, Sharon C; Marsden, Philip A

    2004-03-05

    Endothelin-1 (ET-1) is a potent vasoconstrictor peptide expressed in the vascular endothelium. Stringent control over ET-1 expression is achieved through a highly regulated promoter and rapid mRNA turnover. Since little is known about mechanisms governing ET-1 post-transcriptional regulation, and changes in ET-1 mRNA stability are implicated in disease processes, we characterized these pathways using a variety of functional approaches. We expressed human ET-1 and luciferase transcripts with or without a wild type ET-1 3'-untranslated region (3'-UTR) and found that the 3'-UTR had potent mRNA destabilizing activity. Deletion analysis localized this activity to two domains of the 3'-UTR we have termed destabilizing elements 1 and 2 (DE1 and DE2). Mutational studies revealed that DE1 functions as an AU-rich element (ARE) dependent on a 100-nucleotide region. This activity was further localized to a 10-nucleotide region at position 978-987 of the 3'-UTR. Depletion of AUF1 by RNA interference up-regulated ET-1 in endothelial cells suggesting AUF1-dependent regulation. Since AUF1 functions through the ubiquitin-proteasome pathway, we disrupted this pathway with heat shock and proteasome inhibitor in endothelial cells and observed stabilization of endogenous ET-1 mRNA. Chimeric transcripts bearing wild type ET-1 3'-UTRs were also stabilized in response to proteasome inhibition whereas DE1 mutants failed to respond. Taken together, these findings suggest a complex model of ARE-mediated mRNA turnover dependent on two 3'-UTR domains, DE1 and DE2. Furthermore, DE1 functions as an ARE directing mRNA half-life through the proteasome. Finally, this data provides evidence for a novel pathway of ET-1 mRNA stabilization by heat shock.

  20. Genome-wide association identifies a deletion in the 3’ untranslated region of Striatin in a canine model of arrhythmogenic right ventricular cardiomyopathy

    PubMed Central

    Meurs, Kathryn M.; Mauceli, Evan; Lahmers, Sunshine; Acland, Gregory M.; White, Stephen N.; Lindblad-Toh, Kerstin

    2010-01-01

    Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a familial cardiac disease characterized by ventricular arrhythmias and sudden cardiac death. It is most frequently inherited as an autosomal dominant trait with incomplete and age-related penetrance and variable clinical expression. The human disease is most commonly associated with a causative mutation in one of several genes encoding desmosomal proteins. We have previously described a spontaneous canine model of ARVC in the boxer dog. We phenotyped adult boxer dogs for ARVC by performing physical examination, echocardiogram and ambulatory electrocardiogram. Genome-wide association using the canine 50k SNP array identified several regions of association, of which the strongest resided on chromosome 17. Fine-mapping and direct DNA sequencing identified an eight base pair deletion in the 3’ untranslated region (UTR) of the striatin (STRN) gene on chromosome 17 in association with ARVC in the boxer dog. Evaluation of the secondary structure of the 3’ UTR demonstrated that the deletion affects a stem loop structure of the mRNA and expression analysis identified a reduction in striatin mRNA. Dogs that were homozygous for the deletion had a more severe form of disease based on a significantly higher number of ventricular premature complexes. Immunofluorescence studies localized striatin to the intercalated disc region of the cardiac myocyte and co-localized it to three desmosomal proteins, plakophilin- 2, plakoglobin and desmoplakin, all involved in the pathogenesis of ARVC in human beings. We suggest that striatin may serve as a novel candidate gene for human ARVC. PMID:20596727

  1. A 205-Nucleotide Deletion in the 3′ Untranslated Region of Avian Leukosis Virus Subgroup J, Currently Emergent in China, Contributes to Its Pathogenicity

    PubMed Central

    Wang, Qi; Gao, Yulong; Wang, Yongqiang; Qin, Liting; Qi, Xiaole; Qu, Yue; Gao, Honglei

    2012-01-01

    In the past 5 years, an atypical clinical outbreak of avian leukosis virus subgroup J (ALV-J), which contains a unique 205-nucleotide deletion in its 3′ untranslated region (3′UTR), has become epidemic in chickens in China. To determine the role of the 205-nucleotide deletion in the pathogenicity of ALV-J, a pair of viruses were constructed and rescued. The first virus was an ALV-J Chinese isolate (designated HLJ09SH01) containing the 205-nucleotide deletion in its 3′UTR. The second virus was a chimeric clone in which the 3′UTR contains a 205-nucleotide sequence corresponding to a region of the ALV-J prototype virus. The replication and pathogenicity of the rescued viruses (rHLJ09SH01 and rHLJ09SH01A205) were investigated. Compared to rHLJ09SH01A205, rHLJ09SH01 showed a moderate growth advantage in vitro and in vivo, in addition to exhibiting a higher oncogenicity rate and lethality rate in layers and broilers. Increased vascular endothelial growth factor A (VEGF-A) and vascular endothelial growth receptor subtype 2 (VEGFR-2) expression was induced by rHLJ09SH01 more so than by rHLJ09SH01A205 during early embryonic vascular development, but this increased expression disappeared when the expression levels were normalized to the viral levels. This finding suggests that the expression of VEGF-A and VEGFR-2 is associated with viral replication and may also represent a novel molecular mechanism underlying the oncogenic potential of ALV-J. Overall, our findings not only indicate that the unique 205-nucleotide deletion in the ALV-J genome occurred naturally in China and contributes to increased pathogenicity but also point to the possible mechanism of ALV-J-induced oncogenicity. PMID:22993155

  2. Molecular Mechanism for Hypertensive Renal Disease: Differential Regulation of Chromogranin A Expression at 3'-Untranslated Region Polymorphism C+87T by MicroRNA-107.

    PubMed

    Zhang, Kuixing; Mir, Saiful A; Hightower, C Makena; Miramontes-Gonzalez, Jose Pablo; Maihofer, Adam X; Chen, Yuqing; Mahata, Sushil K; Nievergelt, Caroline M; Schork, Nicholas J; Freedman, Barry I; Vaingankar, Sucheta M; O'Connor, Daniel T

    2015-08-01

    Chromogranin A (CHGA) is coreleased with catecholamines from secretory vesicles in adrenal medulla and sympathetic axons. Genetic variation in the CHGA 3'-region has been associated with autonomic control of circulation, hypertension, and hypertensive nephropathy, and the CHGA 3'-untranslated region (3'-UTR) variant C+87T (rs7610) displayed peak associations with these traits in humans. Here, we explored the molecular mechanisms underlying these associations. C+87T occurred in a microRNA-107 (miR-107) motif (match: T>C), and CHGA mRNA expression varied inversely with miR-107 abundance. In cells transfected with chimeric luciferase/CHGA 3'-UTR reporters encoding either the T allele or the C allele, changes in miR-107 expression levels had much greater effects on expression of the T allele. Cotransfection experiments with hsa-miR-107 oligonucleotides and eukaryotic CHGA plasmids produced similar results. Notably, an in vitro CHGA transcription/translation experiment revealed that changes in hsa-miR-107 expression altered expression of the T allele variant only. Mice with targeted ablation of Chga exhibited greater eGFR. Using BAC transgenesis, we created a mouse model with a humanized CHGA locus (T/T genotype at C+87T), in which treatment with a hsa-miR-107 inhibitor yielded prolonged falls in SBP/DBP compared with wild-type mice. We conclude that the CHGA 3'-UTR C+87T disrupts an miR-107 motif, with differential effects on CHGA expression, and that a cis:trans (mRNA:miR) interaction regulates the association of CHGA with BP and hypertensive nephropathy. These results indicate new strategies for probing autonomic circulatory control and ultimately, susceptibility to hypertensive renal sequelae.

  3. Evolution and Emergence of Enteroviruses through Intra- and Inter-species Recombination: Plasticity and Phenotypic Impact of Modular Genetic Exchanges in the 5' Untranslated Region.

    PubMed

    Muslin, Claire; Joffret, Marie-Line; Pelletier, Isabelle; Blondel, Bruno; Delpeyroux, Francis

    2015-01-01

    Genetic recombination shapes the diversity of RNA viruses, including enteroviruses (EVs), which frequently have mosaic genomes. Pathogenic circulating vaccine-derived poliovirus (cVDPV) genomes consist of mutated vaccine poliovirus (PV) sequences encoding capsid proteins, and sequences encoding nonstructural proteins derived from other species' C EVs, including certain coxsackieviruses A (CV-A) in particular. Many cVDPV genomes also have an exogenous 5' untranslated region (5' UTR). This region is involved in virulence and includes the cloverleaf (CL) and the internal ribosomal entry site, which play major roles in replication and the initiation of translation, respectively. We investigated the plasticity of the PV genome in terms of recombination in the 5' UTR, by developing an experimental model involving the rescue of a bipartite PV/CV-A cVDPV genome rendered defective by mutations in the CL, following the co-transfection of cells with 5' UTR RNAs from each of the four human EV species (EV-A to -D). The defective cVDPV was rescued by recombination with 5' UTR sequences from the four EV species. Homologous and nonhomologous recombinants with large deletions or insertions in three hotspots were isolated, revealing a striking plasticity of the 5' UTR. By contrast to the recombination of the cVDPV with the 5' UTR of group II (EV-A and -B), which can decrease viral replication and virulence, recombination with the 5' UTRs of group I (EV-C and -D) appeared to be evolutionarily neutral or associated with a gain in fitness. This study illustrates how the genomes of positive-strand RNA viruses can evolve into mosaic recombinant genomes through intra- or inter-species modular genetic exchanges, favoring the emergence of new recombinant lineages.

  4. Identification and sexually dimorphic expression of vasa isoforms in Dabry's sturgeon (Acipenser dabryanus), and functional analysis of vasa 3'-untranslated region.

    PubMed

    Ye, Huan; Yue, Hua-Mei; Yang, Xiao-Ge; Li, Chuang-Ju; Wei, Qi-Wei

    2016-10-01

    Germ cells are set aside from somatic cells early in embryogenesis, and are responsible for transmitting genetic information through generations. Vasa is a highly conserved germ cell marker across animal phyla, and widely used to label primordial germ cells. Dabry's sturgeon is a rare and endangered species distributed solely in the Yangtze River basin. Here, seven vasa isoforms, named Advasa1-7, were isolated and characterized in Dabry's sturgeon. RT-PCR and western blot analyses revealed that vasa mRNA and protein were mainly restricted to the testis and ovary, but exhibited sexually dimorphic expression. Cellular and subcellular localization uncovered that Advasa mRNA and protein displayed mitotic and meiotic expression in females, and mainly showed mitotic expression in males; surprisingly, they exhibited both cytoplasmic and nuclear expression in the ovarian germ cells, while showing exclusively cytoplasmic expression in the testicular germ cells. By microinjecting chimeric RNA consisting of the red fluorescent protein coding region and the Advasa 3'-untranslated region into embryos of Dabry's sturgeon, zebrafish and medaka, we demonstrated that it had the ability to visualize primordial germ cells (PGCs) in Dabry's sturgeon and zebrafish but not in medaka. It seemed that the machinery of vasa 3'UTR RNA localization was conserved between Dabry's sturgeon and ostariophysan, while possibly changed during the divergence of euteleosts and ostariophysan. Finally, Dabry's sturgeon PGCs moved on the yolk ball, and migrated toward the genital ridge via mesenchyme. Taken together, these results provide new information for vasa expression pattern and function, and lay a foundation for PGC cryopreservation and conservation of Dabry's sturgeon.

  5. Evolution and Emergence of Enteroviruses through Intra- and Inter-species Recombination: Plasticity and Phenotypic Impact of Modular Genetic Exchanges in the 5’ Untranslated Region

    PubMed Central

    Muslin, Claire; Joffret, Marie-Line; Pelletier, Isabelle; Blondel, Bruno; Delpeyroux, Francis

    2015-01-01

    Genetic recombination shapes the diversity of RNA viruses, including enteroviruses (EVs), which frequently have mosaic genomes. Pathogenic circulating vaccine-derived poliovirus (cVDPV) genomes consist of mutated vaccine poliovirus (PV) sequences encoding capsid proteins, and sequences encoding nonstructural proteins derived from other species’ C EVs, including certain coxsackieviruses A (CV-A) in particular. Many cVDPV genomes also have an exogenous 5’ untranslated region (5’ UTR). This region is involved in virulence and includes the cloverleaf (CL) and the internal ribosomal entry site, which play major roles in replication and the initiation of translation, respectively. We investigated the plasticity of the PV genome in terms of recombination in the 5’ UTR, by developing an experimental model involving the rescue of a bipartite PV/CV-A cVDPV genome rendered defective by mutations in the CL, following the co-transfection of cells with 5’ UTR RNAs from each of the four human EV species (EV-A to -D). The defective cVDPV was rescued by recombination with 5’ UTR sequences from the four EV species. Homologous and nonhomologous recombinants with large deletions or insertions in three hotspots were isolated, revealing a striking plasticity of the 5’ UTR. By contrast to the recombination of the cVDPV with the 5’ UTR of group II (EV-A and -B), which can decrease viral replication and virulence, recombination with the 5’ UTRs of group I (EV-C and -D) appeared to be evolutionarily neutral or associated with a gain in fitness. This study illustrates how the genomes of positive-strand RNA viruses can evolve into mosaic recombinant genomes through intra- or inter-species modular genetic exchanges, favoring the emergence of new recombinant lineages. PMID:26562151

  6. Isolation of an insulin-like growth factor II cDNA with a unique 5 prime untranslated region from human placenta

    SciTech Connect

    Shen, Shujane; Daimon, Makoto; Wang, Chunyeh; Ilan, J. ); Jansen, M. )

    1988-03-01

    Human insulin-like growth factor II (IGF-II) cDNA from a placental library was isolated and sequenced. The 5{prime} untranslated region (5{prime}-UTR) sequence of this cDNA differs completely from that of adult human liver and has considerable base sequence identity to the same region of an IGF-II cDNA of a rat liver cell line, BRL-3A. Human placental poly(A){sup +} RNA was probed with either the 5{prime}-UTR of the isolated human placental IGF-II cDNA or the 5{prime}-UTR of the IGF-II cDNA obtained from adult human liver. No transcripts were detected by using the 5{prime}-UTR of the adult liver IGF-II as the probe. In contrast, three transcripts of 6.0, 3.2, and 2.2 kilobases were detected by using the 5{prime}-UTR of the placental IGF-II cDNA as the probe or the probe from the coding sequence. A fourth IGF-II transcript of 4.9 kilobases presumably containing a 5{prime}-UTR consisting of a base sequence dissimilar to that of either IGF-II 5{prime}-UTR was apparent. Therefore, IGF-II transcripts detected may be products of alternative splicing as their 5{prime}-UTR sequence is contained within the human IGF-II gene or they may be a consequence of alternative promoter utilization in placenta.

  7. The 3'-untranslated region length and AU-rich RNA location modulate RNA-protein interaction and translational control of β2-adrenergic receptor mRNA.

    PubMed

    Subramaniam, Kothandharaman; Kandasamy, Karthikeyan; Joseph, Kusumam; Spicer, Eleanor K; Tholanikunnel, Baby G

    2011-06-01

    Posttranscriptional controls play a major role in β(2)-adrenergic receptor (β(2)-AR) expression. We recently reported that β(2)-AR mRNA translation is suppressed by elements in its 3'-untranslated region (UTR). We also identified T-cell-restricted intracellular antigen-related protein (TIAR) and HuR as prominent AU-rich (ARE) RNA-binding proteins that associate with β(2)-AR mRNA 3'-UTR. In this study, we identified a poly(U) region at the distal end of the 3'-UTR as critical for TIAR binding to β(2)-AR mRNA and for translational suppression. Here, we also report that the locations of the poly(U) and ARE sequences within the 3'-UTR are important determinants that control the translation of β(2)-AR mRNA. Consistent with this finding, a 20-nucleotide ARE RNA from the proximal 3'-UTR that did not inhibit mRNA translation in its native position was able to suppress translation when re-located to the distal 3'-UTR of the receptor mRNA. Immunoprecipitation and polysome profile analysis demonstrated the importance of 3'-UTR length and the ARE RNA location within the 3'-UTR, as key determinants of RNA/protein interactions and translational control of β(2)-AR mRNA. Further, the importance of 3'-UTR length and ARE location in TIAR and HuR association with mRNA and translational suppression was demonstrated using a chimeric luciferase reporter gene.

  8. Characterisation of genotypes among bovine viral diarrhoea virus 2 strains according to palindromic nucleotide substitutions in the 5 untranslated genomic region.

    PubMed

    Giangaspero, M; Harasawa, R

    2004-01-01

    Pestivirus bovine viral diarrhea virus 2 (BVDV-2) strains from 61 isolates from cattle and sheep, and from some adventitious contaminants of biologicals, have been assessed using the palindromic nucleotide substitutions (PNS) method at three variable loci (V1, V2 and V3) located delin the 5' untranslated region (UTR) of genomic RNA. This genotyping procedure is new, simple and practical. Two characteristics of the base pairings common to BVDV-2 species, a C-G or U-A pairing at the V1 locus, and a G*U pairing at the V2 locus, were observed in isolates tested. The PNS method showed six genotypes: BVDV-2a, BVDV-2b, BVDV-2c, BVDV- 2d, BVDV-2e and BVDV-2f. Twenty-five strains showed the BVDV-2a genotype specific combination of three base pairings (A-U in position 1 and C-G or U*G in position 18 in V1 and U-A or U*G in position 4 in V2). Ten strains were identified by a single C-G pairing in position 4 from the bottom of the V2 stem region, characteristic to genotype BVDV-2b. Three strains were assigned to genotype BVDV-2c, due to their recognition by a G*U pairing at the bottom of the V1 stem region. A U-A pairing, characteristic of the BVDV-2d genotype when found in position 18 of the V1 stem region, was observed in fourteen strains. Genotype BVDV- 2e, present in only six South American cattle isolates, was characterized by G-C pairing in position 12, by U-A pairing in position 16 and G_G or G-_A bulges in position 18 in the V1 region. One strain from Argentina was classified as genotype BVDV-2f, showing: A-U pairing in position 9 and 12, U-A in position 16 and G_A bulge in position 18 in V1 region. Two strains were not characterized due to incomplete sequence of V1 locus.

  9. Polypyrimidine tract-binding protein binds to the 5' untranslated region of the mouse mammary tumor virus mRNA and stimulates cap-independent translation initiation.

    PubMed

    Cáceres, Carlos J; Contreras, Nataly; Angulo, Jenniffer; Vera-Otarola, Jorge; Pino-Ajenjo, Constanza; Llorian, Miriam; Ameur, Melissa; Lisboa, Francisco; Pino, Karla; Lowy, Fernando; Sargueil, Bruno; López-Lastra, Marcelo

    2016-05-01

    The 5' untranslated region (UTR) of the full-length mRNA of the mouse mammary tumor virus (MMTV) harbors an internal ribosomal entry site (IRES). In this study, we show that the polypyrimidine tract-binding protein (PTB), an RNA-binding protein with four RNA recognition motifs (RRMs), binds to the MMTV 5' UTR stimulating its IRES activity. There are three isoforms of PTB: PTB1, PTB2, and PTB4. Results show that PTB1 and PTB4, but not PTB2, stimulate MMTV-IRES activity. PTB1 promotes MMTV-IRES-mediated initiation more strongly than PTB4. When expressed in combination, PTB1 further enhanced PTB4 stimulation of the MMTV-IRES, while PTB2 fully abrogates PTB4-induced stimulation. PTB1-induced stimulation of MMTV-IRES was not altered in the presence of PTB4 or PTB2. Mutational analysis reveals that stimulation of MMTV-IRES activity is abrogated when PTB1 is mutated either in RRM1/RRM2 or RRM3/RRM4. In contrast, a PTB4 RRM1/RRM2 mutant has reduced effect over MMTV-IRES activity, while stimulation of the MMTV-IRES activity is still observed when the PTB4 RRM3/RMM4 mutant is used. Therefore, PTB1 and PTB4 differentially stimulate the IRES activity. In contrast, PTB2 acts as a negative modulator of PTB4-induced stimulation of MMTV-IRES. We conclude that PTB1 and PTB4 act as IRES trans-acting factors of the MMTV-IRES.

  10. A variable number of tandem repeats in the 3'-untranslated region of the dopamine transporter modulates striatal function during working memory updating across the adult age span.

    PubMed

    Sambataro, Fabio; Podell, Jamie E; Murty, Vishnu P; Das, Saumitra; Kolachana, Bhaskar; Goldberg, Terry E; Weinberger, Daniel R; Mattay, Venkata S

    2015-08-01

    Dopamine modulation of striatal function is critical for executive functions such as working memory (WM) updating. The dopamine transporter (DAT) regulates striatal dopamine signaling via synaptic reuptake. A variable number of tandem repeats in the 3'-untranslated region of SLC6A3 (DAT1-3'-UTR-VNTR) is associated with DAT expression, such that 9-repeat allele carriers tend to express lower levels (associated with higher extracellular dopamine concentrations) than 10-repeat homozygotes. Aging is also associated with decline of the dopamine system. The goal of the present study was to investigate the effects of aging and DAT1-3'-UTR-VNTR on the neural activity and functional connectivity of the striatum during WM updating. Our results showed both an age-related decrease in striatal activity and an effect of DAT1-3'-UTR-VNTR. Ten-repeat homozygotes showed reduced striatal activity and increased striatal-hippocampal connectivity during WM updating relative to the 9-repeat carriers. There was no age by DAT1-3'-UTR-VNTR interaction. These results suggest that, whereas striatal function during WM updating is modulated by both age and genetically determined DAT levels, the rate of the age-related decline in striatal function is similar across both DAT1-3'-UTR-VNTR genotype groups. They further suggest that, because of the baseline difference in striatal function based on DAT1-3'-UTR-VNTR polymorphism, 10-repeat homozygotes, who have lower levels of striatal function throughout the adult life span, may reach a threshold of decreased striatal function and manifest impairments in cognitive processes mediated by the striatum earlier in life than the 9-repeat carriers. Our data suggest that age and DAT1-3'-UTR-VNTR polymorphism independently modulate striatal function.

  11. Post-transcriptional regulation of cytokine genes in fish: A role for conserved AU-rich elements located in the 3'-untranslated region of their mRNAs.

    PubMed

    Roca, Francisco J; Cayuela, María L; Secombes, Chris J; Meseguer, José; Mulero, Victoriano

    2007-01-01

    The overproduction of cytokines, such us interleukin-1beta (IL-1beta) and tumor necrosis factor alpha (TNFalpha), contributes to the pathological complications observed in many inflammatory diseases caused by bacterial endotoxins. The synthesis of these cytokines is tightly regulated at both transcriptional and post-transcriptional levels. Post-transcriptional regulation of gene expression depends on specific cis-acting sequences and trans-acting factors. Thus, the presence of adenylate- and uridylate-rich (AU-rich) elements (AREs) has been described in the 3'-untranslated regions (UTRs) of many unstable mammalian mRNAs. Although, it represents the most widespread, phylogenetically conserved and efficient determinant of mRNA stability among those so far characterized in mammalian cells, no studies are available on the functional relevance of this sequence in non-mammalian vertebrates. In this contribution, we study the enzymatic activity of various luciferase reporter constructs, containing or lacking the 3'UTR of IL-1beta and TNFalpha from different fish species, and report the finding that bony fish AREs are able to decrease luciferase activity but are less potent than their mammalian counterparts. Surprisingly, the 3'UTR of the IL-1beta from the cartilaginous fish small spotted catshark had the greatest ability to decrease luciferase activity. Lastly, the functional significance of the above was confirmed by measuring the half-life of IL-1beta and TNFalpha mRNAs in gilthead seabream leukocytes by blocking transcription with actinomycin D. Both cytokine mRNAs were unstable with an estimated half-life of about 45 min in control and activated cells.

  12. microRNA-558 facilitates the expression of hypoxia-inducible factor 2 alpha through binding to 5′-untranslated region in neuroblastoma

    PubMed Central

    Song, Huajie; Jiao, Wanju; Li, Dan; Fang, Erhu; Wang, Xiaojing; Mei, Hong; Pu, Jiarui; Huang, Kai; Tong, Qiangsong

    2016-01-01

    Neuroblastoma (NB) is the most common extracranial solid tumor in childhood. Our previous studies have shown that hypoxia-inducible factor 2 alpha (HIF-2α), one member of the bHLH-PAS transcription factor family, facilitates the progression of NB under non-hypoxic conditions. However, the mechanisms underlying HIF-2α expression in NB still remain largely unknown. Herein, through analyzing the computational algorithm programs, we identified microRNA-558 (miR-558) as a crucial regulator of HIF-2α expression in NB. We demonstrated that miR-558 promoted the expression of HIF-2α at translational levels in NB cells through recruiting Argonaute 2 (AGO2). Mechanistically, miR-558 directly bound with its complementary site within 5′-untranslated region (5′-UTR) to facilitate the binding of AGO2 to eukaryotic translation initiation factor 4E (eIF4E) binding protein 1, resulting in increased eIF4E enrichment and HIF-2α translation. In addition, miR-558 promoted the growth, invasion, metastasis, and angiogenesis of NB cells in vitro and in vivo, and these biological features were rescued by knockdown of AGO2, eIF4E, or HIF-2α. In clinical NB specimens, miR-558, AGO2, and eIF4E were highly expressed and positively correlated with HIF-2α expression. Patients with high miR-558, HIF-2α, AGO2, or eIF4E levels had lower survival probability. Taken together, these results demonstrate that miR-558 facilitates the expression of HIF-2α through bindingto its 5′-UTR, thus promoting the tumorigenesis and aggressiveness of NB. PMID:27276678

  13. Mini-genome rescue of Crimean-Congo hemorrhagic fever virus and research into the evolutionary patterns of its untranslated regions.

    PubMed

    Zhao, Jiuru; Xia, Han; Zhang, Yujiang; Yin, Shiyu; Zhang, Zhong; Tang, Shuang; Kou, Zheng; Yu, Jingfeng; Fan, Zhaojun; Li, Tianxian

    2013-10-01

    Crimean-Congo hemorrhagic fever virus (CCHFV) is a member of genus Nairovirus, family Bunyaviridae, which are distributed widely in Africa, Europe and Asia with several genotypes. As a BSL-4 level pathogen, the requirement of high-level biosafety facilities severely constrains researches on live virus manipulation. In this study, we developed a helper-virus-independent mini-genome rescue system for the Chinese YL04057 strain. Based on the enhanced green fluorescent protein (EGFP)-derived mini-genome plasmids, this polymerase I driven system permits easy observation and quantification. Unlike previous report, gradually reduced levels of activity of the CCHFV L, M and S untranslated regions (UTRs) were observed in our system. We also demonstrated that the UTRs at both ends were indispensable for mini-genome background expression. In addition, we phylogentically analyzed all six UTRs of CCHFV and showed that L-UTRs were clustered together approximately corresponding to their original geographical continents. The UTRs of M segment showed a similar branch structure to its open reading frames (ORFs), and nearly an identical tree was generated with 5' UTRs of S segment compared with its ORFs. However, the 3' UTRs of S segment formed new divergent groups. Compatibility tests of YL04057 strain nucleocapsid protein and L protein expression plasmids with Nigerian strain IbAr10200 mini-genomes revealed lower compatibility of L-UTRs without an obvious effect on M-UTRs. Moreover, we demonstrated that the L-UTRs could tolerate certain nucleotide mutations. This system may provide a foundation for future studies of the viral replication cycle, pathogenic mechanisms and evolutionary patterns of CCHFV.

  14. Spliceosomal introns in the 5′ untranslated region of plant BTL RING-H2 ubiquitin ligases are evolutionary conserved and required for gene expression

    PubMed Central

    2013-01-01

    Background Introns located close to the 5′ end of a gene or in the 5′ untranslated region often exert positive effects on gene expression. This effect, known as intron-mediated enhancement (IME), has been observed in diverse eukaryotic organisms, including plants. The sequences involved in IME seem to be spread across the intron and function in an additive manner. The IMEter algorithm was developed to predict plant introns that may enhance gene expression. We have identified several plant members of the BTL class of E3s, which may have orthologs across eukaryotes, that contain a 5′UTR intron. The RING finger E3 ligases are key enzymes of the ubiquitination system that mediate the transfer of ubiquitin to substrates. Results In this study, we retrieved BTL sequences from several angiosperm species and found that 5′UTR introns showing a strong IMEter score were predicted, suggesting that they may be conserved by lineage. Promoter-GUS fusion lines were used to confirm the IME effect of these 5′UTR introns on gene expression. IMEter scores of BTLs were compared with the 5′UTR introns of two gene families MHX and polyubiquitin genes. Conclusions Analysis performed in two Arabidopsis BTL E3 ligases genes indicated that the 5′UTR introns were essential for gene expression in all the tissues tested. Comparison of the average 5′UTR intron size on three gene families in ten angiosperm species suggests that a prevalent size for a 5′UTR intron is in the range of 600 nucleotides, and that the overall IMEter score within a gene family is preserved across several angiosperms. Our results indicated that gene expression dependent on a 5′UTR intron is an efficient regulatory mechanism in BTL E3 ligases that has been preserved throughout plant evolution. PMID:24228887

  15. Effect of β2-adrenergic receptor gene (ADRB2) 3′ untranslated region polymorphisms on inhaled corticosteroid/long-acting β2-adrenergic agonist response

    PubMed Central

    2012-01-01

    Background Evidence suggests that variation in the length of the poly-C repeat in the 3′ untranslated region (3′UTR) of the β2-adrenergic receptor gene (ADRB2) may contribute to interindividual variation in β-agonist response. However, methodology in previous studies limited the assessment of the effect of sequence variation in the context of poly-C repeat length. The objectives of this study were to design a novel genotyping method to fully characterize sequence variation in the ADRB2 3′UTR poly-C repeat in asthma patients treated with inhaled corticosteroid and long-acting β2-adrenergic agonist (ICS/LABA) combination therapy, and to analyze the effect of the poly-C repeat polymorphism on clinical response. Methods In 2,250 asthma patients randomized to treatment with budesonide/formoterol or fluticasone/salmeterol in a six-month study (AstraZeneca study code: SD-039-0735), sequence diversity in the ADRB2 poly-C repeat region was determined using a novel sequencing-based genotyping method. The relationship between the poly-C repeat polymorphism and the incidence of severe asthma exacerbations, and changes in pulmonary function and asthma symptoms from baseline to the average during the treatment period, were analyzed. Results Poly-C repeat genotypes were assigned in 97% (2,192/2,250) of patients. Of the 13 different poly-C repeat alleles identified, six alleles occurred at a frequency of >5% in one or more population in this study. The repeat length of these six common alleles ranged from 10 to 14 nucleotides. Twelve poly-C repeat genotypes were observed at a frequency of >1%. No evidence of an association between poly-C repeat genotype and the incidence of severe asthma exacerbations was observed. Patients’ pulmonary function measurements improved and asthma symptoms declined when treated with ICS/LABA combination therapy regardless of poly-C repeat genotype. Conclusions The extensive sequence diversity present in the poly-C repeat region of the ADRB2

  16. The (95)(Δ)G mutation in the 5'untranslated region of the norA gene increases efflux activity in Staphylococcus epidermidis isolates.

    PubMed

    García-Gómez, Elizabeth; Jaso-Vera, Marcos E; Juárez-Verdayes, Marco A; Alcántar-Curiel, María D; Zenteno, Juan C; Betanzos-Cabrera, Gabriel; Peralta, Humberto; Rodríguez-Martínez, Sandra; Cancino-Díaz, Mario E; Jan-Roblero, Janet; Cancino-Diaz, Juan C

    2017-02-01

    In the Staphylococcus aureus ATCC25923 strain, the flqB mutation in the 5'untranslated region (5'UTR) of the norA gene causes increased norA mRNA expression and high efflux activity (HEA). The involvement of the norA gene 5'UTR in HEA has not been explored in S. epidermidis; therefore, we examined the function of this region in S. epidermidis clinical isolates. The selection of isolates with HEA was performed based on ethidium bromide (EtBr) MIC values and efflux efficiency (EF) using the semi-automated fluorometric method. The function of the 5'UTR was studied by quantifying the levels of norA expression (RT-qPCR) and by identifying 5'UTR mutations by sequence analysis. Only 10 isolates from a total of 165 (6.1%) had HEA (EtBr MIC = 300 μg/ml and EF ranged from 48.4 to 97.2%). Eight of 10 isolates with HEA had the 5'UTR (95)(Δ)G mutation. Isolates carrying the (95)(Δ)G mutation had higher levels of norA expression compared with those that did not. To corroborate that the (95)(Δ)G mutation is involved in HEA, a strain adapted to EtBr was obtained in vitro. This strain also presented the (95)(Δ)G mutation and had a high level of norA expression and EF, indicating that the (95)(Δ)G mutation is important for the HEA phenotype. The (95)(Δ)G mutation produces a different structure in the Shine-Dalgarno region, which may promote better translation of norA mRNA. To our knowledge, this is the first report to demonstrate the participation of the 5'UTR (95)(Δ)G mutation of the norA gene in the HEA phenotype of S. epidermidis isolates. Here, we propose that the efflux of EtBr is caused by an increment in the transcription and/or translation of the norA gene.

  17. 5[prime] to 3[prime] nucleic acid synthesis using 3[prime]-photoremovable protecting group

    DOEpatents

    Pirrung, M.C.; Shuey, S.W.; Bradley, J.C.

    1999-06-01

    The present invention relates, in general, to a method of synthesizing a nucleic acid, and, in particular, to a method of effecting 5[prime] to 3[prime] nucleic acid synthesis. The method can be used to prepare arrays of oligomers bound to a support via their 5[prime] end. The invention also relates to a method of effecting mutation analysis using such arrays. The invention further relates to compounds and compositions suitable for use in such methods.

  18. Delayed translational silencing of ceruloplasmin transcript in gamma interferon-activated U937 monocytic cells: role of the 3' untranslated region

    NASA Technical Reports Server (NTRS)

    Mazumder, B.; Fox, P. L.

    1999-01-01

    Ceruloplasmin (Cp) is an acute-phase protein with ferroxidase, amine oxidase, and pro- and antioxidant activities. The primary site of Cp synthesis in human adults is the liver, but it is also synthesized by cells of monocytic origin. We have shown that gamma interferon (IFN-gamma) induces the synthesis of Cp mRNA and protein in monocytic cells. We now report that the induced synthesis of Cp is terminated by a mechanism involving transcript-specific translational repression. Cp protein synthesis in U937 cells ceased after 16 h even in the presence of abundant Cp mRNA. RNA isolated from cells treated with IFN-gamma for 24 h exhibited a high in vitro translation rate, suggesting that the transcript was not defective. Ribosomal association of Cp mRNA was examined by sucrose centrifugation. When Cp synthesis was high, i.e., after 8 h of IFN-gamma treatment, Cp mRNA was primarily associated with polyribosomes. However, after 24 h, when Cp synthesis was low, Cp mRNA was primarily in the nonpolyribosomal fraction. Cytosolic extracts from cells treated with IFN-gamma for 24 h, but not for 8 h, contained a factor which blocked in vitro Cp translation. Inhibitor expression was cell type specific and present in extracts of human cells of myeloid origin, but not in several nonmyeloid cells. The inhibitory factor bound to the 3' untranslated region (3'-UTR) of Cp mRNA, as shown by restoration of in vitro translation by synthetic 3'-UTR added as a "decoy" and detection of a binding complex by RNA gel shift analysis. Deletion mapping of the Cp 3'-UTR indicated an internal 100-nucleotide region of the Cp 3'-UTR that was required for complex formation as well as for silencing of translation. Although transcript-specific translational control is common during development and differentiation and global translational control occurs during responses to cytokines and stress, to our knowledge, this is the first report of translational silencing of a specific transcript following cytokine

  19. Untranslated region-dependent exclusive expression of high-sensitivity subforms of alpha4beta2 and alpha3beta2 nicotinic acetylcholine receptors.

    PubMed

    Briggs, Clark A; Gubbins, Earl J; Marks, Michael J; Putman, C Brent; Thimmapaya, Rama; Meyer, Michael D; Surowy, Carol S

    2006-07-01

    alpha4beta2 nicotinic acetylcholine receptors (nAChRs) are recognized as the principal nicotine binding site in brain. Recombinant alpha4beta2 nAChR demonstrate biphasic concentration-response relationships with low- and high-EC50 components. This study shows that untranslated regions (UTR) can influence expression of high-sensitivity subforms of alpha4beta2 and alpha3beta2 nAChR. Oocytes injected with alpha4 and beta2 RNA lacking UTR expressed biphasic concentration-response relationships for acetylcholine with high-sensitivity EC50 values of 0.5 to 2.5 microM (14-24% of the population) and low-sensitivity EC50 values of 110 to 180 microM (76-86%). In contrast, message with UTR expressed exclusively the high-sensitivity alpha4beta2 nAChR subform with an acetylcholine EC50 value of 2.2 microM. Additional studies revealed pharmacological differences between high- and low-sensitivity alpha4beta2 subforms. Whereas the antagonists dihydro-beta-erythroidine (IC50 of 3-6 nM) and methyllycaconitine (IC50 of 40-135 nM) were not selective between high- and low-sensitivity alpha4beta2, chlorisondamine, mecamylamine, and d-tubocurarine were, respectively, 100-, 8-, and 5-fold selective for the alpha4beta2 subform with low sensitivity to acetylcholine. Conversely, agonists that selectively activated the high-sensitivity alpha4beta2 subform with respect to efficacy as well as potency were identified. Furthermore, two of these agonists were shown to activate mouse brain alpha4beta2 as well as the ferret high-sensitivity alpha4beta2 expressed in Xenopus laevis oocytes. With the use of UTR-containing RNA, exclusive expression of a novel high-sensitivity alpha3beta2 nAChR was also achieved. These studies 1) provide further evidence for the existence of multiple subforms of alpha4beta2 nAChR, 2) extend that to alpha3beta2 nAChR, 3) demonstrate UTR influence on beta2-containing nAChR properties, and 4) reveal compounds that interact with alpha4beta2 in a subform-selective manner.

  20. A HuD-ZBP1 ribonucleoprotein complex localizes GAP-43 mRNA into axons through its 3' untranslated region AU-rich regulatory element.

    PubMed

    Yoo, Soonmoon; Kim, Hak H; Kim, Paul; Donnelly, Christopher J; Kalinski, Ashley L; Vuppalanchi, Deepika; Park, Michael; Lee, Seung J; Merianda, Tanuja T; Perrone-Bizzozero, Nora I; Twiss, Jeffery L

    2013-09-01

    Localized translation of axonal mRNAs contributes to developmental and regenerative axon growth. Although untranslated regions (UTRs) of many different axonal mRNAs appear to drive their localization, there has been no consensus RNA structure responsible for this localization. We recently showed that limited expression of ZBP1 protein restricts axonal localization of both β-actin and GAP-43 mRNAs. β-actin 3'UTR has a defined element for interaction with ZBP1, but GAP-43 mRNA shows no homology to this RNA sequence. Here, we show that an AU-rich regulatory element (ARE) in GAP-43's 3'UTR is necessary and sufficient for its axonal localization. Axonal GAP-43 mRNA levels increase after in vivo injury, and GAP-43 mRNA shows an increased half-life in regenerating axons. GAP-43 mRNA interacts with both HuD and ZBP1, and HuD and ZBP1 co-immunoprecipitate in an RNA-dependent fashion. Reporter mRNA with the GAP-43 ARE competes with endogenous β-actin mRNA for axonal localization and decreases axon length and branching similar to the β-actin 3'UTR competing with endogenous GAP-43 mRNA. Conversely, over-expressing GAP-43 coding sequence with its 3'UTR ARE increases axonal elongation and this effect is lost when just the ARE is deleted from GAP-43's 3'UTR. We have recently found that over-expression of GAP-43 using an axonally targeted construct with the 3'UTRs of GAP-43 promoted elongating growth of axons, while restricting the mRNA to the cell body with the 3'UTR of γ-actin had minimal effect on axon length. In this study, we show that the ARE in GAP-43's 3'UTR is responsible for localization of GAP-43 mRNA into axons and is sufficient for GAP-43 protein's role in elongating axonal growth.

  1. The 5′ Untranslated Region of the Human T-Cell Lymphotropic Virus Type 1 mRNA Enables Cap-Independent Translation Initiation

    PubMed Central

    Olivares, Eduardo; Landry, Dori M.; Cáceres, C. Joaquín; Pino, Karla; Rossi, Federico; Navarrete, Camilo; Huidobro-Toro, Juan Pablo; Thompson, Sunnie R.

    2014-01-01

    ABSTRACT The human T-cell leukemia virus type 1 (HTLV-1) is a complex human retrovirus that causes adult T cell leukemia and of HTLV-associated myelopathy/tropical spastic paraparesis. The mRNA of some complex retroviruses, including the human and simian immunodeficiency viruses (HIV and SIV), can initiate translation using a canonical cap-dependent mechanism or through an internal ribosome entry site (IRES). In this study, we present strong evidence showing that like HIV-1 and SIV, the 5′-untranslated region (5′UTR) of the HTLV-1 full-length mRNA harbors an IRES. Cap-independent translational activity was evaluated and demonstrated using dual luciferase bicistronic mRNAs in rabbit reticulocyte lysate, in mammalian cell culture, and in Xenopus laevis oocytes. Characterization of the HTLV-1 IRES shows that its activity is dependent on the ribosomal protein S25 (RPS25) and that its function is highly sensitive to the drug edeine. Together, these findings suggest that the 5′UTR of the HTLV-1 full-length mRNA enables internal recruitment of the eukaryotic translation initiation complex. However, the recognition of the initiation codon requires ribosome scanning. These results suggest that, after internal recruitment by the HTLV-1 IRES, a scanning step takes place for the 40S ribosomal subunit to be positioned at the translation initiation codon. IMPORTANCE The mechanism by which retroviral mRNAs recruit the 40S ribosomal subunit internally is not understood. This study provides new insights into the mechanism of translation initiation used by the human T-cell lymphotropic virus type 1 (HTLV-1). The results show that the HTLV-1 mRNA can initiate translation via a noncanonical mechanism mediated by an internal ribosome entry site (IRES). This study also provides evidence showing the involvement of cellular proteins in HTLV-1 IRES-mediated translation initiation. Together, the data presented in this report significantly contribute to the understanding of HTLV-1 gene

  2. The proximal region of the 3'-untranslated region of cyclooxygenase-2 is recognized by a multimeric protein complex containing HuR, TIA-1, TIAR, and the heterogeneous nuclear ribonucleoprotein U.

    PubMed

    Cok, Steven J; Acton, Stephen J; Morrison, Aubrey R

    2003-09-19

    Cyclooxygenase-2 (COX-2) is an early response gene induced in renal mesangial cells by interleukin-1beta (IL-1beta). The 3'-untranslated region (3'-UTR) of COX-2 mRNA plays an important role in IL-1beta induction by regulating message stability and translational efficiency. The first 60 nucleotides of the 3'-UTR of COX-2 are highly conserved and contain multiple copies of the regulatory sequence AUUUA. Introduction of the 60-nucleotide sequence into the 3'-UTR of a heterologous reporter gene resulted in a 70% decrease in reporter gene expression. Electrophoretic mobility shift assays (EMSAs) demonstrated that mesangial cell nuclear fractions contain a multimeric protein complex that bound this region of COX-2 mRNA in a sequence-specific manner. We identified four members of the protein-RNA complex as HuR, TIA-1, TIAR, and the heterogeneous nuclear ribonucleoprotein U (hnRNP U). Treatment of mesangial cells with IL-1beta caused an increase in cytosolic HuR, which was accompanied by an increase in COX-2 mRNA that co-immunoprecipitated with cytosolic HuR. Therefore, we propose that HuR binds to the proximal region of the 3'-UTR of COX-2 following stimulation by IL-1beta and increases the expression of COX-2 mRNA by facilitating its transport out of the nucleus.

  3. Comparative analysis of the base compositions of the pre-mRNA 3' cleaved-off region and the mRNA 3' untranslated region relative to the genomic base composition in animals and plants.

    PubMed

    Li, Xiu-Qing

    2014-01-01

    The precursor messenger RNA (pre-mRNA) three-prime cleaved-off region (3'COR) and the mRNA three-prime untranslated region (3'UTR) play critical roles in regulating gene expression. The differences in base composition between these regions and the corresponding genomes are still largely uncharacterized in animals and plants. In this study, the base compositions of non-redundant 3'CORs and 3'UTRs were compared with the corresponding whole genomes of eleven animals, four dicotyledonous plants, and three monocotyledonous (cereal) plants. Among the four bases (A, C, G, and U for adenine, cytosine, guanine, and uracil, respectively), U (which corresponds to T, for thymine, in DNA) was the most frequent, A the second most frequent, G the third most frequent, and C the least frequent in most of the species in both the 3'COR and 3'UTR regions. In comparison with the whole genomes, in both regions the U content was usually the most overrepresented (particularly in the monocotyledonous plants), and the C content was the most underrepresented. The order obtained for the species groups, when ranked from high to low according to the U contents in the 3'COR and 3'UTR was as follows: dicotyledonous plants, monocotyledonous plants, non-mammal animals, and mammals. In contrast, the genomic T content was highest in dicotyledonous plants, lowest in monocotyledonous plants, and intermediate in animals. These results suggest the following: 1) there is a mechanism operating in both animals and plants which is biased toward U and against C in the 3'COR and 3'UTR; 2) the 3'UTR and 3'COR, as functional units, minimized the difference between dicotyledonous and monocotyledonous plants, while the dicotyledonous and monocotyledonous genomes evolved into two extreme groups in terms of base composition.

  4. Role of the 2 adenine (g.11293_11294insAA) insertion polymorphism in the 3' untranslated region of the factor VII (FVII) gene: molecular characterization of a patient with severe FVII deficiency.

    PubMed

    Peyvandi, F; Garagiola, I; Palla, R; Marziliano, N; Mannucci, P M

    2005-11-01

    Polymorphic variants in the gene encoding factor VII (F7) affect the plasma levels of this coagulation protein and modify the clinical phenotype of FVII deficiency in some patients. In this study we report the in vitro functional analysis of a novel polymorphic variant located in the 3' untranslated region of F7: g.11293_11294insAA. To determine whether this variant regulates FVII expression, we initially compared an expression vector containing FVII cDNA with g.11293_11294insAA with the FVII wild-type (WT) construct. The kinetics of mRNA production showed that the insertion decreases the steady-state FVII mRNA levels. To assess whether the insertion influences the phenotype of FVII-deficient patients, we evaluated its effect on the expression of FVII in a patient with severe FVII deficiency (undetectable FVII activity and antigen) carrying two additional homozygous missense variations (p.Arg277Cys and p.Arg353Gln). The two substitutions alone reduced the expression of FVII activity and antigen in vitro, but with the insertion polymorphism in our expression vector the patient's phenotype of undetectable plasma FVII was recapitulated. The insertion polymorphism in the 3' untranslated region of F7 is another modifier of FVII expression that might explain the poor genotype-phenotype correlation in some FVII-deficient patients.

  5. The 3' untranslated region of the Andes hantavirus small mRNA functionally replaces the poly(A) tail and stimulates cap-dependent translation initiation from the viral mRNA.

    PubMed

    Vera-Otarola, Jorge; Soto-Rifo, Ricardo; Ricci, Emiliano P; Ohlmann, Théophile; Darlix, Jean-Luc; López-Lastra, Marcelo

    2010-10-01

    In the process of translation of eukaryotic mRNAs, the 5' cap and the 3' poly(A) tail interact synergistically to stimulate protein synthesis. Unlike its cellular counterparts, the small mRNA (SmRNA) of Andes hantavirus (ANDV), a member of the Bunyaviridae, lacks a 3' poly(A) tail. Here we report that the 3' untranslated region (3'UTR) of the ANDV SmRNA functionally replaces a poly(A) tail and synergistically stimulates cap-dependent translation initiation from the viral mRNA. Stimulation of translation by the 3'UTR of the ANDV SmRNA was found to be independent of viral proteins and of host poly(A)-binding protein.

  6. The 3′ Untranslated Region of the Andes Hantavirus Small mRNA Functionally Replaces the Poly(A) Tail and Stimulates Cap-Dependent Translation Initiation from the Viral mRNA ▿

    PubMed Central

    Vera-Otarola, Jorge; Soto-Rifo, Ricardo; Ricci, Emiliano P.; Ohlmann, Théophile; Darlix, Jean-Luc; López-Lastra, Marcelo

    2010-01-01

    In the process of translation of eukaryotic mRNAs, the 5′ cap and the 3′ poly(A) tail interact synergistically to stimulate protein synthesis. Unlike its cellular counterparts, the small mRNA (SmRNA) of Andes hantavirus (ANDV), a member of the Bunyaviridae, lacks a 3′ poly(A) tail. Here we report that the 3′ untranslated region (3′UTR) of the ANDV SmRNA functionally replaces a poly(A) tail and synergistically stimulates cap-dependent translation initiation from the viral mRNA. Stimulation of translation by the 3′UTR of the ANDV SmRNA was found to be independent of viral proteins and of host poly(A)-binding protein. PMID:20660206

  7. The chicken FMR1 gene is highly conserved with a CCT 5{prime} - untranslated repeat and encodes an RNA-binding protein

    SciTech Connect

    Price, D.K.; Zhang, F.; Ashley, C.T. Jr.; Warren, S.T.

    1996-01-01

    The transcriptional silencing of the human gene, fragile X metal retardation 1 (FMR1), is due to abnormal methylation in response to an expanded 5{prime}-untranslated CGG trinucleotide repeat and accounts for most cases of fragile X syndrome, a frequent inherited form of metal retardation. Although the encoded fragile X mental retardation protein (FMRP) is known to have properties of a RNA-binding protein, the precise function of FMRP remains to be elucidated. We report the cloning of the chicken homolog of FMR1 and show strong evolutionary conservation, with nucleotide and amino acid identities of 85 and 92%, respectively, between chicken and human. In place of the mammalian CGG trinucleotide repeat, a 99-nt tripartite repetitive element containing a CCT trinucleotide repeat flanked on both sides by dinucleotide repeats was identified. Blocks of highly conserved 3{prime}-untranslated sequence were also found. Within the coding region, two copies each of the highly conserved K homology motif and the Arg-Gly-Gly (RGG) box motif, both ribonucleotide particle family domains implicated in RNA binding, were identified. Chicken FMRP was found to bind RNA in vitro, and this activity correlated with the presence of the carboxy-terminal portion of the protein that includes the RGG motifs. 49 refs., 7 figs.

  8. Structural complexity of Dengue virus untranslated regions: cis-acting RNA motifs and pseudoknot interactions modulating functionality of the viral genome

    PubMed Central

    Sztuba-Solinska, Joanna; Teramoto, Tadahisa; Rausch, Jason W.; Shapiro, Bruce A.; Padmanabhan, Radhakrishnan; Le Grice, Stuart F. J.

    2013-01-01

    The Dengue virus (DENV) genome contains multiple cis-acting elements required for translation and replication. Previous studies indicated that a 719-nt subgenomic minigenome (DENV-MINI) is an efficient template for translation and (−) strand RNA synthesis in vitro. We performed a detailed structural analysis of DENV-MINI RNA, combining chemical acylation techniques, Pb2+ ion-induced hydrolysis and site-directed mutagenesis. Our results highlight protein-independent 5′–3′ terminal interactions involving hybridization between recognized cis-acting motifs. Probing analyses identified tandem dumbbell structures (DBs) within the 3′ terminus spaced by single-stranded regions, internal loops and hairpins with embedded GNRA-like motifs. Analysis of conserved motifs and top loops (TLs) of these dumbbells, and their proposed interactions with downstream pseudoknot (PK) regions, predicted an H-type pseudoknot involving TL1 of the 5′ DB and the complementary region, PK2. As disrupting the TL1/PK2 interaction, via ‘flipping’ mutations of PK2, previously attenuated DENV replication, this pseudoknot may participate in regulation of RNA synthesis. Computer modeling implied that this motif might function as autonomous structural/regulatory element. In addition, our studies targeting elements of the 3′ DB and its complementary region PK1 indicated that communication between 5′–3′ terminal regions strongly depends on structure and sequence composition of the 5′ cyclization region. PMID:23531545

  9. Sequence and spatial requirements for the tissue- and species-independent 3{prime}-end processing mechanism of plant mRNA

    SciTech Connect

    Wu, L.; Ueda, T.; Messing, J.

    1994-10-01

    Two cis-regulatory regions are required for efficient mRNA 3{prime}-end processing of the maize 27-kDa zein mRNA: a region containing a duplicated AAUGAA poly(A) signal and a region that is present upstream from it. Strict spatial positioning of these two regions is required for efficient mRNA 3{prime}-end processing. Insertions of a stuffer sequence as short as 17 or 18 bp either between the upstream region and the two AAUGAA motifs or between the two AAUGAA motifs drastically reduced the efficiency of 3{prime}-end processing. Mutational analyses of the nucleotide preference at the fourth position of the AAUGAA motif revealed the preference order G > A >> C or U, suggesting that AAUAAA is neither a defective nor an optimal poly(A) signal for the 27-kDa zein mRNA. As for the 3{prime} control region of the cauliflower mosaic virus (CaMV) transcription unit, the mRNA 3{prime}-end processing mechanism mediated by the 27-kDa zein 3{prime} control sequence is neither tissue nor species specific. The 3{prime} upstream sequence of the 27-kDa zein gene can functionally replace that of the CaMV transcription unit. Conversely, the CaMV upstream sequence can mediate mRNA polyadenylation in the presence of a duplicated 27-kDa zein poly(A) signal. However, instead of the proximal poly(A) signal normally used in the 27-kDa zein mRNA, the distal signal is utilized. These results suggest that a general mechanism controls the 3{prime}-end processing of plant mRNAs and that the cis-regulatory functions mediated by their upstream regions are interchangeable. 35 refs., 5 figs.

  10. Specific binding of Fusarium graminearum Hex1 protein to untranslated regions of the genomic RNA of Fusarium graminearum virus 1 correlates with increased accumulation of both strands of viral RNA.

    PubMed

    Son, Moonil; Choi, Hoseong; Kim, Kook-Hyung

    2016-02-01

    The HEX1 gene of Fusarium graminearum was previously reported to be required for the efficient accumulation of Fusarium graminearum virus 1 (FgV1) RNA in its host. To investigate the molecular mechanism underlying the production of FgHEX1 and the replication of FgV1 viral RNA, we conducted electrophoretic mobility shift assays (EMSA) with recombinant FgHex1 protein and RNA sequences derived from various regions of FgV1 genomic RNA. These analyses demonstrated that FgHex1 and both the 5'- and 3'-untranslated regions of plus-strand FgV1 RNA formed complexes. To determine whether FgHex1 protein affects FgV1 replication, we quantified accumulation viral RNAs in protoplasts and showed that both (+)- and (-)-strands of FgV1 RNAs were increased in the over-expression mutant and decreased in the deletion mutant. These results indicate that the FgHex1 functions in the synthesis of both strands of FgV1 RNA and therefore in FgV1 replication probably by specifically binding to the FgV1 genomic RNA.

  11. p190RhoGEF Binds to a destabilizing element in the 3' untranslated region of light neurofilament subunit mRNA and alters the stability of the transcript.

    PubMed

    Cañete-Soler, R; Wu, J; Zhai, J; Shamim, M; Schlaepfer, W W

    2001-08-24

    Stabilization of neurofilament (NF) mRNAs plays a major role in regulating levels of NF expression and in establishing axonal size and rate of axonal conduction. Previous studies have identified a 68-nucleotide destabilizing element at the junction of the coding region and 3' untranslated region of the light NF subunit (NF-L) mRNA. The present study has used the destabilizing element (probe A) to screen a rat brain cDNA library for interactive proteins. A cDNA clone encoding 1068 nucleotides in the C-terminal domain of p190RhoGEF (clone 39) was found to bind strongly and specifically to the RNA probe. The interaction was confirmed using a glutathione S-transferase/clone 39 fusion protein in Northwestern, gel-shift, and cross-linkage studies. The glutathione S-transferase/clone 39 fusion protein also enhanced the cross-linkage of a major 43-kDa protein in brain extract to the destabilizing element. Functional studies on stably transfected neuronal cells showed that p190RhoGEF expression increased the half-life of a wild-type NF-L mRNA but did not alter the half-life of a mutant NF-L mRNA lacking the destabilizing element. The findings reveal a novel interactive feature of p190RhoGEF that links the exchange factor with NF mRNA stability and regulation of the axonal cytoskeleton.

  12. Maize Mu transposons are targeted to the 5' untranslated region of the gl8 gene and sequences flanking Mu target-site duplications exhibit nonrandom nucleotide composition throughout the genome.

    PubMed Central

    Dietrich, Charles R; Cui, Feng; Packila, Mark L; Li, Jin; Ashlock, Daniel A; Nikolau, Basil J; Schnable, Patrick S

    2002-01-01

    The widespread use of the maize Mutator (Mu) system to generate mutants exploits the preference of Mu transposons to insert into genic regions. However, little is known about the specificity of Mu insertions within genes. Analysis of 79 independently isolated Mu-induced alleles at the gl8 locus established that at least 75 contain Mu insertions. Analysis of the terminal inverted repeats (TIRs) of the inserted transposons defined three new Mu transposons: Mu10, Mu 11, and Mu12. A large percentage (>80%) of the insertions are located in the 5' untranslated region (UTR) of the gl8 gene. Ten positions within the 5' UTR experienced multiple independent Mu insertions. Analyses of the nucleotide composition of the 9-bp TSD and the sequences directly flanking the TSD reveals that the nucleotide composition of Mu insertion sites differs dramatically from that of random DNA. In particular, the frequencies at which C's and G's are observed at positions -2 and +2 (relative to the TSD) are substantially higher than expected. Insertion sites of 315 RescueMu insertions displayed the same nonrandom nucleotide composition observed for the gl8-Mu alleles. Hence, this study provides strong evidence for the involvement of sequences flanking the TSD in Mu insertion-site selection. PMID:11861572

  13. Functional single nucleotide polymorphism in IL-17A 3' untranslated region is targeted by miR-4480 in vitro and may be associated with age-related macular degeneration.

    PubMed

    Popp, Nicholas A; Yu, Dianke; Green, Bridgett; Chew, Emily Y; Ning, Baitang; Chan, Chi-Chao; Tuo, Jingsheng

    2016-01-01

    Age-related macular degeneration (AMD) is a leading cause of irreversible central vision loss in the elderly. Genetic factors contributing to AMD include single nucleotide polymorphisms (SNPs) in immune-related genes including CFH, C2, CFI, C9, and C3, thus implicating these pathways in AMD pathogenesis. MicroRNAs (miRNAs) are powerful regulators of gene expression and execute this function by binding to the 3' untranslated region (3'UTR) of target mRNAs, leading to mRNA degradation. In this study, we searched for the possible association of SNPs in the 3'UTR region of IL-17A, a gene implicated in AMD pathogenesis without any previous SNP association with AMD. Using two independent sample cohorts of Caucasian subjects, six SNPs in the IL-17A 3'-UTR were selected for genotyping based on bioinformatic predictions of the SNP effect on microRNA binding. The SNP rs7747909 was found to be associated with AMD (P < 0.05) in the NEI cohort, using a dominant model logistic regression. Luciferase reporter gene assays and RNA electrophoretic mobility shift assays were performed using ARPE-19 cells to confirm the preferential binding of microRNAs to the major allele of the SNP. Our findings support the hypothesis that microRNA-mediated gene dysregulation may play a role in the pathogenesis of AMD.

  14. DNA Sequence Variants in the Five Prime Untranslated Region of the Cyclooxygenase-2 Gene Are Commonly Found in Healthy Dogs and Gray Wolves

    PubMed Central

    Safra, Noa; Hayward, Louisa J.; Aguilar, Miriam; Sacks, Benjamin N.; Westropp, Jodi L.; Mohr, F. Charles; Mellersh, Cathryn S.; Bannasch, Danika L.

    2015-01-01

    The aim of this study was to investigate the frequency of regional DNA variants upstream to the translation initiation site of the canine Cyclooxygenase-2 (Cox-2) gene in healthy dogs. Cox-2 plays a role in various disease conditions such as acute and chronic inflammation, osteoarthritis and malignancy. A role for Cox-2 DNA variants in genetic predisposition to canine renal dysplasia has been proposed and dog breeders have been encouraged to select against these DNA variants. We sequenced 272–422 bases in 152 dogs unaffected by renal dysplasia and found 19 different haplotypes including 11 genetic variants which had not been described previously. We genotyped 7 gray wolves to ascertain the wildtype variant and found that the wolves we analyzed had predominantly the second most common DNA variant found in dogs. Our results demonstrate an elevated level of regional polymorphism that appears to be a feature of healthy domesticated dogs. PMID:26244515

  15. SHAPE Analysis of the RNA Secondary Structure of the Mouse Hepatitis Virus 5′ Untranslated Region and N-Terminal Nsp1 Coding Sequences

    PubMed Central

    Yang, Dong; Liu, Pinghua; Wudeck, Elyse V.; Giedroc, David P.; Leibowitz, Julian L.

    2014-01-01

    SHAPE technology was used to analyze RNA secondary structure of the 5′ most 474 nts of the MHV-A59 genome encompassing the minimal 5′ cis-acting region required for defective interfering RNA replication. The structures generated were in agreement with previous characterizations of SL1 through SL4 and two recently predicted secondary structure elements, S5 and SL5A. SHAPE provided biochemical support for four additional stem-loops not previously functionally investigated in MHV. Secondary structure predictions for 5′ regions of MHV-A59, BCoV and SARS-CoV were similar despite high sequence divergence. The pattern of SHAPE reactivity of in virio genomic RNA, ex virio genomic RNA, and in vitro synthesized RNA were similar, suggesting that binding of N protein or other proteins to virion RNA fails to protect the RNA from reaction with lipid permeable SHAPE reagent. Reverse genetic experiments suggested that SL5C and SL6 within the nsp1 coding sequence are not required for viral replication. PMID:25462342

  16. A 3′ untranslated region variant in FMR1 eliminates neuronal activity-dependent translation of FMRP by disrupting binding of the RNA-binding protein HuR

    PubMed Central

    Suhl, Joshua A.; Muddashetty, Ravi S.; Anderson, Bart R.; Ifrim, Marius F.; Visootsak, Jeannie; Bassell, Gary J.; Warren, Stephen T.

    2015-01-01

    Fragile X syndrome is a common cause of intellectual disability and autism spectrum disorder. The gene underlying the disorder, fragile X mental retardation 1 (FMR1), is silenced in most cases by a CGG-repeat expansion mutation in the 5′ untranslated region (UTR). Recently, we identified a variant located in the 3′UTR of FMR1 enriched among developmentally delayed males with normal repeat lengths. A patient-derived cell line revealed reduced levels of endogenous fragile X mental retardation protein (FMRP), and a reporter containing a patient 3′UTR caused a decrease in expression. A control reporter expressed in cultured mouse cortical neurons showed an expected increase following synaptic stimulation that was absent when expressing the patient reporter, suggesting an impaired response to neuronal activity. Mobility-shift assays using a control RNA detected an RNA–protein interaction that is lost with the patient RNA, and HuR was subsequently identified as an associated protein. Cross-linking immunoprecipitation experiments identified the locus as an in vivo target of HuR, supporting our in vitro findings. These data suggest that the disrupted interaction of HuR impairs activity-dependent translation of FMRP, which may hinder synaptic plasticity in a clinically significant fashion. PMID:26554012

  17. Replacement of the yeast TRP4 3' untranslated region by a hammerhead ribozyme results in a stable and efficiently exported mRNA that lacks a poly(A) tail.

    PubMed Central

    Düvel, Katrin; Valerius, Oliver; Mangus, David A; Jacobson, Allan; Braus, Gerhard H

    2002-01-01

    The mRNA poly(A) tail serves different purposes, including the facilitation of nuclear export, mRNA stabilization, efficient translation, and, finally, specific degradation. The posttranscriptional addition of a poly(A) tail depends on sequence motifs in the 3' untranslated region (3' UTR) of the mRNA and a complex trans-acting protein machinery. In this study, we have replaced the 3' UTR of the yeast TRP4 gene with sequences encoding a hammerhead ribozyme that efficiently cleaves itself in vivo. Expression of the TRP4-ribozyme allele resulted in the accumulation of a nonpolyadenylated mRNA. Cells expressing the TRP4-ribozyme mRNA showed a reduced growth rate due to a reduction in Trp4p enzyme activity. The reduction in enzyme activity was not caused by inefficient mRNA export from the nucleus or mRNA destabilization. Rather, analyses of mRNA association with polyribosomes indicate that translation of the ribozyme-containing mRNA is impaired. This translational defect allows sufficient synthesis of Trp4p to support growth of trp4 cells, but is, nevertheless, of such magnitude as to activate the general control network of amino acid biosynthesis. PMID:12003493

  18. Molecular cloning of cDNA for the zeta isoform of the 14-3-3 protein: homologous sequences in the 3'-untranslated region of frog and human zeta isoforms.

    PubMed

    Miura, I; Nakajima, T; Ohtani, H; Kashiwagi, A; Nakamura, M

    1997-10-01

    14-3-3 proteins constitute a family of well-conserved eukaryotic proteins that possess diverse biochemical activities such as regulation of gene transcription, cell proliferation and activation of protein kinase C. At least 7 subtypes (alpha to theta) of 14-3-3 protein are known, but the zeta subtype of this protein has been cloned only in mammals. We cloned the zeta subtype of 14-3-3 protein (14-3-3 zeta) from the frog, Rana rugosa. The sequence encoded 245 amino acids that share 92% identity with rat and bovine 14-3-3 zeta s, and 92% with human phospholipase A2 (PLA2; 14-3-3 zeta). Northern blot analysis revealed a single band of about 1.8 kb in tadpoles at stage 25. The 14-3-3 zeta mRNA level was high in the brain, lung, spleen and kidney, and low in the heart and testis, as opposed to the mRNA level, which was only faintly detected in the liver, pancreas, ovary and muscle. Furthermore, high similarity in the 3'-untranslated region (3'-UTR) was observed between frog and human 14-3-3 zeta cDNA. The results suggest that 14-3-3 zeta is highly conserved throughout eukaryotic evolution, and that the homologous sequence in the 3'-UTR of 14-3-3 zeta cDNA may be conserved in frogs and humans.

  19. AU-rich elements in the mRNA 3'-untranslated region of the rat receptor for advanced glycation end products and their relevance to mRNA stability.

    PubMed

    Caballero, José Juan; Girón, María Dolores; Vargas, Alberto Manuel; Sevillano, Natalia; Suárez, María Dolores; Salto, Rafael

    2004-06-18

    Several putative polyadenylation sequences and an adenylate plus timidylate rich element (ARE) are present at the 3' end of the rat advanced glycation end products receptor (RAGE) gene. Two transcripts are generated by the use of alternative polyadenylation sequences, one containing the ARE sequence in its 3'-untranslated region (3'-UTR). Transfections of CHO-k1 or NRK cells with constructs expressing the 3'-UTRs of the transcripts fused to a green fluorescence protein mRNA show that the ARE sequence has a negative effect on protein expression correlating with a decrease in the amount of mRNA, as shown in CHO-k1 transfected cells. When transfected cells were incubated in the presence of Actinomycin D the amount of fluorescence decreased in cells transfected with the ARE sequence, indicating that this sequence induces lower mRNA stability. Thus, alternative polyadenylation signals and an ARE sequence provide a novel mechanism for the regulation of the rat RAGE gene expression.

  20. microRNA-4717 differentially interacts with its polymorphic target in the PD1 3' untranslated region: A mechanism for regulating PD-1 expression and function in HBV-associated liver diseases.

    PubMed

    Zhang, Guoyu; Li, Na; Li, Zhu; Zhu, Qianqian; Li, Fang; Yang, Cuiling; Han, Qunying; Lv, Yi; Zhou, Zhihua; Liu, Zhengwen

    2015-08-07

    Programmed cell death-1 (PD-1) is involved in hepatitis B virus (HBV) infection, the leading cause of hepatocellular carcinoma (HCC) worldwide. Single-nucleotide polymorphism, rs10204525, located in the PD1 3' untranslated regions (UTR), is associated with chronic HBV infection. MicroRNAs (miRNAs) regulate gene expression via specific binding to the target 3'UTR of mRNA. In this study, three miRNAs were predicted to putatively interact with PD1 rs10204525 polymorphic site of allele G. One of them, miRNA-4717, was demonstrated to allele-specifically affect luciferase activity in a dose-dependent manner in cells transfected with vectors containing different rs10204525 alleles. In lymphocytes from chronic HBV patients withrs10204525 genotype GG, miR-4717 mimics significantly decreased PD-1 expression and increased (TNF)-α and interferon (IFN)-γ production. miR-4717 inhibitor significantly increased PD-1 expression and decreased TNF-α and IFN-γ production although not significantly. In lymphocytes from chronic HBV patients with rs10204525 genotype AA, no similar effects were observed. miR-4717 levels in peripheral lymphocytes from patients with HBV-related chronic hepatitis, cirrhosis and HCC were significantly decreased. In conclusion, miR-4717 may allele-specifically regulate PD-1 expression through interaction with the 3' UTR of PD1 mRNA, leading to the alteration of immune regulation and affecting the susceptibility and disease course of chronic HBV infection.

  1. IL-1beta induces stabilization of IL-8 mRNA in malignant breast cancer cells via the 3' untranslated region: Involvement of divergent RNA-binding factors HuR, KSRP and TIAR.

    PubMed

    Suswam, Esther A; Nabors, L Burt; Huang, Yuanyuan; Yang, Xiuhua; King, Peter H

    2005-03-01

    IL-8 plays an integral role in promoting the malignant phenotype in breast cancer, and its production is directly influenced by inflammatory cytokines in the tumor microenvironment. Here, we show that activation of IL-1beta receptors on malignant HS578t and MDA-MB-231 breast cancer cells strongly induces IL-8 expression and that RNA stabilization is persistently activated at least 12-24 hr after stimulation. SB 203580 and rapamycin reversed the RNA stabilization effect of IL-1beta in a dose-dependent manner, suggesting involvement of the p38/MAP kinase and mTOR pathways. A luciferase reporter assay indicated that the stabilization effect was dependent on cis elements in the 3'-untranslated region (UTR) of the IL-8 transcript. By UV cross-linking, we identified multiple cellular factors that interact with the IL-8 3'UTR, ranging 34-76 kDa. Immunoprecipitation analysis indicated that HuR, KSRP and TIAR bound to one or more loci in the 3'UTR. While the cross-linking patterns were similar, quantitative immunoprecipitation of native IL-8 RNA from IL-1beta-stimulated cytoplasmic extract revealed a 20-fold greater association of transcript with the stabilizing factor HuR vs. the destabilizing factor KSRP. In conclusion, IL-1beta is a potent cytokine stimulus for IL-8 RNA stabilization in breast cancer cells, possibly by enhanced binding of cytoplasmic HuR to the 3'UTR.

  2. Most microRNAs in the single-cell alga Chlamydomonas reinhardtii are produced by Dicer-like 3-mediated cleavage of introns and untranslated regions of coding RNAs

    PubMed Central

    Valli, Adrian A.; Santos, Bruno A.C.M.; Hnatova, Silvia; Bassett, Andrew R.; Molnar, Attila; Chung, Betty Y.; Baulcombe, David C.

    2016-01-01

    We describe here a forward genetic screen to investigate the biogenesis, mode of action, and biological function of miRNA-mediated RNA silencing in the model algal species, Chlamydomonas reinhardtii. Among the mutants from this screen, there were three at Dicer-like 3 that failed to produce both miRNAs and siRNAs and others affecting diverse post-biogenesis stages of miRNA-mediated silencing. The DCL3-dependent siRNAs fell into several classes including transposon- and repeat-derived siRNAs as in higher plants. The DCL3-dependent miRNAs differ from those of higher plants, however, in that many of them are derived from mRNAs or from the introns of pre-mRNAs. Transcriptome analysis of the wild-type and dcl3 mutant strains revealed a further difference from higher plants in that the sRNAs are rarely negative switches of mRNA accumulation. The few transcripts that were more abundant in dcl3 mutant strains than in wild-type cells were not due to sRNA-targeted RNA degradation but to direct DCL3 cleavage of miRNA and siRNA precursor structures embedded in the untranslated (and translated) regions of the mRNAs. Our analysis reveals that the miRNA-mediated RNA silencing in C. reinhardtii differs from that of higher plants and informs about the evolution and function of this pathway in eukaryotes. PMID:26968199

  3. The mammalian target of rapamycin complex 1 regulates leptin biosynthesis in adipocytes at the level of translation: the role of the 5'-untranslated region in the expression of leptin messenger ribonucleic acid.

    PubMed

    Chakrabarti, Partha; Anno, Takatoshi; Manning, Brendan D; Luo, Zhijun; Kandror, Konstantin V

    2008-10-01

    Leptin production by adipose cells in vivo is increased after feeding and decreased by food deprivation. However, molecular mechanisms that control leptin expression in response to food intake remain unknown. Here, we test the hypothesis that leptin expression in adipose cells is regulated by nutrient- and insulin-sensitive mammalian target of rapamycin complex 1 (mTORC1)-mediated pathway. The activity of mTORC1 in 3T3-L1 adipocytes was up-regulated by stable expression of either constitutively active Rheb or dominant-negative AMP-activated protein kinase. In both cases, expression of endogenous leptin was significantly elevated at the level of translation. To investigate the role of leptin 5'-untranslated region (UTR) in the regulation of protein expression, we created bicistronic reporter constructs with and without the 5'-UTR. We found that the presence of leptin 5'-UTR renders mRNA resistant to regulation by mTORC1. It appears, therefore, that mTORC1 controls translation of leptin mRNA via a novel mechanism that does not require the presence of either the 5'-terminal oligopyrimidine tract or the 5'-UTR.

  4. The Rev protein of human immunodeficiency virus type 1 counteracts the effect of an AU-rich negative element in the human papillomavirus type 1 late 3' untranslated region.

    PubMed Central

    Tan, W; Schwartz, S

    1995-01-01

    We have identified a sequence in the late 3' untranslated region of human papillomavirus type 1 mRNAs that acts posttranscriptionally to repress gene expression. Deletion analysis localized the inhibitory element to an AU-rich sequence between nucleotides 6958 and 6984 on the human papillomavirus type 1 genome. This sequence inhibits gene expression in an orientation-dependent manner. Upon transfection of eucaryotic cells with plasmids containing this sequence, approximately 4-fold-lower cytoplasmic mRNA levels and 64- to 128-fold-lower protein levels were produced compared with those produced by plasmids lacking the inhibitory sequence. Interestingly, providing the constitutive transport element of simian retrovirus type 1 in sense orientation counteracted inhibition exerted by the human papillomavirus type 1 sequence. Inhibition could also be overcome by the presence of human immunodeficiency virus type 1 Rev protein in trans and its target sequence, the Rev-responsive element, in cis. Rev is a nuclear protein and acts by promoting nuclear export of human immunodeficiency virus type 1 mRNAs encoding structural proteins. Our results are consistent with a model for human papillomavirus type 1 late-gene expression in which mRNAs containing human papillomavirus type 1 inhibitory sequences enter a nonproductive route in the nucleus, resulting in inefficient mRNA utilization. Rev directs mRNA containing inhibitory sequences to a productive route by interacting with the Rev-responsive element. PMID:7707519

  5. The phosphorylation of protein S6 modulates the interaction of the 40 S ribosomal subunit with the 5'-untranslated region of a dictyostelium pre-spore-specific mRNA and controls its stability.

    PubMed

    Chiaberge, S; Cassarino, E; Mangiarotti, G

    1998-10-16

    AC914 mRNA, a pre-spore-specific mRNA that accumulates only in the post-aggregation stage of development, is transcribed constitutively as shown by nuclear run-off experiments and by fusing its promoter to the luciferase reporter gene. The same mRNA disappears quickly from disaggregated cells. If the 5'-untranslated region (5'UTR) of the constitutively expressed Actin 15 mRNA is substituted for the 5'UTR of AC914 mRNA, this can no longer be destabilized and accumulates both in growing and disaggregated cells. If the 5'UTR of AC914 mRNA is substituted for the 5'UTR of Actin 15 mRNA, the latter accumulates only in aggregated cells. Pactamycin, but not other inhibitors of protein synthesis, prevents AC914 mRNA from being destabilized in disaggregated cells, suggesting a role of 40 S subunits in the destabilization. This has been confirmed by using an in vitro system in which the in vivo stability of different mRNAs is reproduced. A protein kinase A-dependent phosphorylation of ribosomal protein S6 determines whether 40 S subunits are capable or not of destabilizing AC914 mRNA in the in vitro system.

  6. Genetic variation in the 3’ untranslated region of dengue virus serotype 3 strains isolated from mosquitoes and humans in Brazil

    PubMed Central

    2013-01-01

    Summary Background Dengue, a mosquito-borne viral infection caused by one of the four dengue virus (DENV) serotypes (DENV-1 to 4), replicate alternately on the mosquito vector and human host and are responsible for infections throughout tropical and subtropical regions of the world. In Brazil, the disease has become a major public health problem and the introduction of DENV-3 in 2000 in Rio de Janeiro (RJ) was associated with severe dengue epidemics. The potential emergence of strains associated with severe disease highlights the need for the surveillance of DENV in human host and vectors. Methods Aiming to contribute for DENV phylogenetic and vector-virus-human host studies, we sequenced the entire genome of one DENV-3 isolated from naturally infected Aedes aegypti from RJ in 2001 and characterized the 3’ UTR from strains isolated from mosquitoes and humans. Mosquitoes were pooled and submitted to virus isolation in Ae. albopictus C6/36 cells and the infecting serotype was identified by immunofluorescence using type-specific monoclonal antibody. Sequence analysis was performed using BioEdit software, the multiple alignments were performed using CLUSTAL W and the phylogenetic analysis by MEGA 5, using the Neighbor-joining method. Secondary structure prediction was performed by using the MFOLD program. Results Exclusive substitutions and a substitution leading to a stop codon on the NS5 gene were observed in the DENV-3 isolated from a naturally infected Ae. aegypti and fully sequenced. As an 8- nucleotides deletion was observed within the 11- nucleotides (nts) insertion on the variable region (VR) from the 3′UTR in this isolate, we further sequenced other DENV-3 from both mosquitoes and humans. The majority of DENV-3 from RJ analyzed were characterized by the 11-nts insertion in the VR of the 3′UTR, despite the observation of strains carrying the 8-nts deletion. The latter presented similar secondary structures, however not all strains presenting the 11-nts

  7. 5'-Untranslated region of the tryptophan hydroxylase-2 gene harbors an asymmetric bidirectional promoter but not internal ribosome entry site in vitro.

    PubMed

    Chen, Guo-Lin; Miller, Gregory M

    2009-04-15

    Tryptophan hydroxylase-2 (TPH2) catalyzes the synthesis of neuronal serotonin, a major neurotransmitter involved in many brain functions and psychiatric disorders. We have previously revealed a critical role of the human TPH2 (hTPH2) 5'-UTR in gene expression regulation. This study aimed to further characterize mechanism(s) by which the hTPH2 5'-UTR regulates gene expression. An internal ribosome entry site (IRES) activity in hTPH2 5'-UTR was suggested by the conventional bicistronic reporter assay; however, further stringent experiments, including in vitro translation, quantitative real-time PCR, Northern blot, ribonuclease protection assay, and monocistronic reporter assay, demonstrated that the hTPH2 5'-UTR harbors a bidirectional promoter, but not IRES, within its downstream segment (61-141). The antisense promoter is much stronger than the sense promoter, but the strength of both promoters are cell-line dependent, with the highest and lowest activities being observed in HEK-293T and SK-N-MC cells, respectively. In accordance with our previous findings, the upstream segment (1-60) of hTPH2 5'-UTR suppresses the neighboring promoter of both direction, independent of the cell line and its location in the 5'- or 3'-flanking regions of the gene. In summary, this study demonstrates that no IRES but an asymmetric bidirectional promoter is present in the downstream segment of hTPH2 5'-UTR, and this promoter is susceptible to a gene silencing effect caused by the upstream segment (1-60) of hTPH2 5'-UTR. Our findings point to the potential involvement of antisense transcription and non-coding RNA in the regulation of TPH2 gene expression.

  8. Next-Generation Sequencing of 5' Untranslated Region of Hepatitis C Virus in Search of Minor Viral Variant in a Patient Who Revealed New Genotype While on Antiviral Treatment.

    PubMed

    Caraballo Cortes, Kamila; Bukowska-Ośko, Iwona; Pawełczyk, Agnieszka; Perlejewski, Karol; Płoski, Rafał; Lechowicz, Urszula; Stawiński, Piotr; Demkow, Urszula; Laskus, Tomasz; Radkowski, Marek

    2016-01-01

    The role of mixed infections with different hepatitis C virus (HCV) genotypes in viral persistence, treatment effects, and tissue tropism is unclear. Next-generation sequencing (NGS), which is suitable for analysis of large, genetically diverse populations offers unparalleled advantages for the study of mixed infections. The aim of the study was to determine, using two different deep sequencing strategies (pyrosequencing - 454 Life Sciences/Roche and reversible terminator sequencing-by-synthesis by Illumina), the origin of a novel HCV genotype transiently detectable during antiviral therapy (pre-existing minor population vs. de novo superinfection). Secondly, we compared 5' untranslated region (5'-UTR) variants obtained by the two NGS approaches. 5' UTR amplification products from 9 samples collected from genotype 1b infected patient before, during, and after treatment (4 serum and 5 peripheral blood mononuclear cell - PBMC - samples) were subjected to the next-generation sequencing. The sequencing revealed the presence of two (454/Roche) and one (Illumina) genotype 4 variants in PBMC at Week 16. None of these variants were present either in the preceding or following samples as revealed by both platforms. 454/Roche sequencing detected 24 different 5'-UTR variants: 8 were present in serum and PBMC, 4 only in serum and 12 only in PBMC. Illumina sequencing detected 11 different 5'-UTR variants: 5 in serum and PBMC, 4 only in serum and 2 only in PBMC. Six variants were identical for both sequencing platforms. The difference in variants number was primarily due to variability in two 5'-UTR homopolymeric regions. In conclusion, longitudinal analysis of HCV variants, employing two independent deep sequencing methods, suggests that the transient presence of a different genotype strain in PBMC was a result of superinfection and not a selection of pre-existing minor variant.

  9. PCK1 is negatively regulated by bta-miR-26a, and a single-nucleotide polymorphism in the 3' untranslated region is involved in semen quality and longevity of Holstein bulls.

    PubMed

    Huang, Jinming; Guo, Fang; Zhang, Zebin; Zhang, Yuanpei; Wang, Xiuge; Ju, Zhihua; Yang, Chunhong; Wang, Changfa; Hou, Minghai; Zhong, Jifeng

    2016-03-01

    Phosphoenolpyruvate carboxykinase 1 (PCK1) is a multi-functional enzyme that plays important roles in physiological processes, including reproduction. We previously reported that the PCK1 transcript has five splice variants; PCK1-AS4, which lacks exon 5, is enriched in the testis of Holstein bulls. In the present study, we profiled select PCK1 transcript variants in the testis, epididymus, and semen of high- and low-performance bulls, and examined the possibility that microRNAs may be involved in single nucleotide polymorphism (SNP)-mediated modulation of PCK1 expression. PCK1-AS4 abundance is not significantly different between high- and low-performance bulls. Luciferase reporter assays, however, showed that bovine PCK1 expression is repressed by bta-miR-26a in HepG2 hepatocyte cells. One SNP (c. + 2183 G > T) at the miRNA-binding site of PCK1 does not influence PCK1 expression, but is associated with elevated ejaculation volume, fresh sperm motility, and genomic estimated breeding value of longevity, as well as with reduced values of composite index and calving ease. Collectively, the identified 3'-untranslated-region SNP variant highlights the importance of PCK1 in the fecundity of Holstein bulls, and implicates a role for bta-miR-26a in regulating PCK1 abundance. Further study is needed to assess the effects of other genetic variants in 5'-flanking region and exons of PCK1 on enzyme levels in the testis and sperm. Mol. Reprod. Dev. 83: 217-225, 2016. © 2016 Wiley Periodicals, Inc.

  10. An insertion/deletion polymorphism in the 3' untranslated region of type I collagen a2 (COL1A2) is associated with susceptibility for hepatocellular carcinoma in a Chinese population.

    PubMed

    Zhu, Zhansheng; Jiang, Yuting; Chen, Shougong; Jia, Shasha; Gao, Xueren; Dong, Dong; Gao, Yuzhen

    2011-05-01

    Hepatocellular carcinoma (HCC) is one of the most common and severe diseases in the world. Besides the influence of environmental factors, such as viral infection, an increasing number of novel genetic components identified by genome-wide association studies have been associated with predisposition to HCC. Thus, studies focusing on functional variants in these findings are indispensable. In the present study, based on in-silico analysis, we carried out a case-control study in a Chinese population (207 cases and 245 controls) to investigate the association between HCC susceptibility with a 7 base pair (bp) insertion/deletion polymorphism (rs3917) in the 3'UTR of COL1A2. Our results showed that the ins/del + del/del genotype had an odds ratio of 1.76 (95% C.I.=1.03-3.01; P=0.028) for developing HCC compared to the ins/ins genotype. Carriers for the "del" allele of rs3917 were associated with a 1.73-fold increased risk for HCC (95% C.I.=1.06-2.84; P(trend)=0.02). Computational modeling suggests that this polymorphism is located in the hsa-let-7 g potential target sequence in the COL1A2 3' untranslated region. Our data suggest that most likely, common genetic changes in COL1A2 may influence HCC risk, at least in part by let-7 g-mediated regulation, which is possibly involved in the pathogenesis of HCC. The replication of our studies in other populations will further strengthen our understanding of this association.

  11. XIAP 3'-untranslated region as a ceRNA promotes FSCN1 function in inducing the progression of breast cancer by binding endogenous miR-29a-5p.

    PubMed

    Wu, Qiang; Yan, Hong; Tao, Si-Qi; Wang, Xiao-Nan; Mou, Lang; Chen, Ping; Cheng, Xing-Wang; Wu, Wen-Yong; Wu, Zheng-Sheng

    2017-02-07

    The non-coding 3'-untranslated region (UTR) of genes play an important role in the regulation of microRNA (miRNA) functions, since it can bind and inactivate multiple miRNAs. Herein, we report that ectopic expression of XIAP 3'UTR increased human breast cancer cells proliferation, colony formation, migration, invasion and xenograft tumor growth and suppressed tumor cell death. To investigate this process, we further correlated the genome-wide transcriptional profiling with the gene expression alterations after transfecting XIAP 3'UTR in MCF-7 cells. We identified a robust, genome-wide mechanism of cell migration, motility and epithelial to mesenchymal transition by which mediated by a previously described cellular component movement factor FSCN1. Expression of XIAP and FSCN1 were up-regulated synergistically after transfecting XIAP 3'UTR in vitro and in vivo. Interactions between XIAP and FSCN1 appear to be a key determinant of these processes. Co-transfection with Dicer siRNA reversed the XIAP 3'UTR-mediated oncogenicity, suggesting the miRNAs might be involved in that process. Furthermore, we demonstrated that one miRNA, miR-29a-5p, can bind to both the XIAP and FSCN1 3'UTRs and play an important role in that interactions. We showed that the 3'UTR of XIAP was able to antagonize miR-29a-5p, and resulted in the increased translation of XIAP and FSCN1. Thus, our findings reveal important new insights into how XIAP 3'UTR works, suggesting that the non-coding XIAP 3'UTR serves as a competitor for miRNA binding and subsequently inactivates miRNA functions, by which XIAP 3'UTR frees the target mRNAs from being repressed.

  12. Mutant copper-zinc superoxide dismutase associated with amyotrophic lateral sclerosis binds to adenine/uridine-rich stability elements in the vascular endothelial growth factor 3′-untranslated region

    PubMed Central

    Li, Xuelin; Lu, Liang; Bush, Donald J.; Zhang, Xiaowen; Zheng, Lei; Suswam, Esther A.; King, Peter H.

    2009-01-01

    Vascular endothelial growth factor (VEGF) is a neurotrophic factor essential for maintenance of motor neurons. Loss of this factor produces a phenotype similar to amyotrophic lateral sclerosis (ALS). We recently showed that ALS-producing mutations of Cu/Zn-superoxide dismutase (SOD1) disrupt post-transcriptional regulation of VEGF mRNA, leading to significant loss of expression. Mutant SOD1 was present in the ribonucleoprotein complex associated with adenine/uridine-rich elements (ARE) of the VEGF 3′-untranslated region (UTR). Here, we show by electrophoretic mobility shift assay that mutant SOD1 bound directly to the VEGF 3′-UTR with a predilection for AREs similar to the RNA stabilizer HuR. SOD1 mutants A4V and G37R showed higher affinity for the ARE than L38V or G93A. Wild-type SOD1 bound very weakly with an apparent Kd 11- to 72-fold higher than mutant forms. Mutant SOD1 showed an additional lower shift with VEGF ARE that was accentuated in the metal-free state. A similar pattern of binding was observed with AREs of tumor necrosis factor-α and interleukin-8, except only a single shift predominated. Using an ELISA-based assay, we demonstrated that mutant SOD1 competes with HuR and neuronal HuC for VEGF 3′-UTR binding. To define potential RNA-binding domains, we truncated G37R, G93A and wild-type SOD1 and found that peptides from the N-terminal portion of the protein that included amino acids 32-49 could recapitulate the binding pattern of full-length protein. Thus, the strong RNA-binding affinity conferred by ALS-associated mutations of SOD1 may contribute to the post-transcriptional dysregulation of VEGF mRNA. PMID:19196430

  13. Oligomerizations of deoxyadenosine bis-phosphates and of their 3-prime-5-prime, 3-prime-3-prime, and 5-prime-5-prime dimers - Effects of a pyrophosphate-linked, poly(T) analog

    NASA Technical Reports Server (NTRS)

    Visscher, J.; Bakker, C. G.; Schwartz, Alan W.

    1990-01-01

    The effect of a 3-prime-5-prime pyrophosphate-linked oligomer of pTp on oligomerizations of pdAp and of its 3-prime-5-prime, 3-prime-3-prime, and 5-prime-5-prime dimers was investigated, using HPLC to separate the reaction mixtures; peak detection was by absorbance monitoring at 254 nm. It was expected that the dimers would form stable complexes with the template, with the degree of stability depending upon the internal linkage of each dimer. It was found that, although the isomers differ substantially in their oligomerization behavior in the absence of template, the analog-template catalyzes the oligomerization to about the same extent in all three cases.

  14. Reverse Stroop Effects with Untranslated Responses

    ERIC Educational Resources Information Center

    Blais, Chris; Besner, Derek

    2006-01-01

    Translation accounts have argued that the presence of a Stroop effect in the context of a nonvocal untranslated response is caused by verbal mediation. In its simplest form, color-labeled buttons are translated into a verbal code that interferes with color responses. On this logic, in the reverse Stroop task (identify the word; ignore the color),…

  15. rs78378222 polymorphism in the 3'-untranslated region of TP53 contributes to development of age-associated cataracts by modifying microRNA-125b-induced apoptosis of lens epithelial cells.

    PubMed

    Zhao, Yang; Li, Xiao; Zhu, Siquan

    2016-09-01

    MicroRNAs (miRNAs) negatively regulate the expression of the target genes by binding to 'seed sequences' in the 3'‑untranslated region (3'‑UTR) mRNA transcripts, and the variants within or nearby 'seed sequences' may compromise or enhance miRNA/mRNA interaction leading to either 'loss‑of‑function' or 'gain‑of‑function' effects. Cataracts are the leading cause of blindness worldwide and are characterized by progressive aggregation and precipitation of lens proteins, and the development of age‑related cataracts is associated with dysregulated cellular activities of lens epithelial cells. Luciferase assays and online miRNA databases were used to validate that tumor protein p53 (TP53) is the target gene of miR‑125b. Furthermore, reverse transcription‑quantitative polymerase chain reaction and western blotting were conducted to detect expression levels of miR‑125b and TP53 in different groups of cells transfected with miR‑125b mimics or inhibitors. In addition, flow cytometry analysis and the MTT assay were conducted to detect the effects of miR‑125b on apoptosis and cell viability. The current study demonstrated that the rs78378222 polymorphism minor allele introduces a novel potential miR‑125b binding site in the TP53 3'‑UTR with a consecutive 8‑bp perfect match, creating a 'gain‑of‑function' variant and affecting the regulation of TP53 expression. A luciferase assay demonstrated that transfection of lens epithelial cells with wild type TP53 3'‑UTR significantly reduced the luciferase activity of the miR‑125b overexpressing cells compared with scramble controls. In addition, the luciferase activity of miR‑125b overexpressing cells transfected with the construct containing the rs78378222 polymorphism minor allele was also reduced compared with cells transfected with the wild type 3'‑UTR. Furthermore, it was demonstrated that the expression level of miR‑125 was comparable in epithelial cells from patients with age

  16. Comparative performance evaluation of hepatitis C virus genotyping based on the 5' untranslated region versus partial sequencing of the NS5B region of brazilian patients with chronic hepatitis C

    PubMed Central

    2011-01-01

    Background Genotyping of hepatitis C virus (HCV) has become an essential tool for prognosis and prediction of treatment duration. The aim of this study was to compare two HCV genotyping methods: reverse hybridization line probe assay (LiPA v.1) and partial sequencing of the NS5B region. Methods Plasma of 171 patients with chronic hepatitis C were screened using both a commercial method (LiPA HCV Versant, Siemens, Tarrytown, NY, USA) and different primers targeting the NS5B region for PCR amplification and sequencing analysis. Results Comparison of the HCV genotyping methods showed no difference in the classification at the genotype level. However, a total of 82/171 samples (47.9%) including misclassification, non-subtypable, discrepant and inconclusive results were not classified by LiPA at the subtype level but could be discriminated by NS5B sequencing. Of these samples, 34 samples of genotype 1a and 6 samples of genotype 1b were classified at the subtype level using sequencing of NS5B. Conclusions Sequence analysis of NS5B for genotyping HCV provides precise genotype and subtype identification and an accurate epidemiological representation of circulating viral strains. PMID:21967749

  17. Saccharomyces cerevisiae Hrq1 requires a long 3 Prime -tailed DNA substrate for helicase activity

    SciTech Connect

    Kwon, Sung-Hun; Choi, Do-Hee; Lee, Rina; Bae, Sung-Ho

    2012-10-26

    Highlights: Black-Right-Pointing-Pointer Hrq1 has intrinsic 3 Prime -5 Prime helicase and DNA strand annealing activities. Black-Right-Pointing-Pointer Hrq1 requires a long 3 Prime -tail for efficient DNA unwinding. Black-Right-Pointing-Pointer Helicase activity of Hrq1 is stimulated by a fork structure. Black-Right-Pointing-Pointer Hrq1 is a moderately processive helicase. -- Abstract: RecQ helicases are well conserved proteins from bacteria to human and function in various DNA metabolism for maintenance of genome stability. Five RecQ helicases are found in humans, whereas only one RecQ helicase has been described in lower eukaryotes. However, recent studies predicted the presence of a second RecQ helicase, Hrq1, in fungal genomes and verified it as a functional gene in fission yeast. Here we show that 3 Prime -5 Prime helicase activity is intrinsically associated with Hrq1 of Saccharomyces cerevisiae. We also determined several biochemical properties of Hrq1 helicase distinguishable from those of other RecQ helicase members. Hrq1 is able to unwind relatively long duplex DNA up to 120-bp and is significantly stimulated by a preexisting fork structure. Further, the most striking feature of Hrq1 is its absolute requirement for a long 3 Prime -tail ( Greater-Than-Or-Slanted-Equal-To 70-nt) for efficient unwinding of duplex DNA. We also found that Hrq1 has potent DNA strand annealing activity. Our results indicate that Hrq1 has vigorous helicase activity that deserves further characterization to expand our understanding of RecQ helicases.

  18. Hypoxia may increase rat insulin mRNA levels by promoting binding of the polypyrimidine tract-binding protein (PTB) to the pyrimidine-rich insulin mRNA 3'-untranslated region.

    PubMed Central

    Tillmar, Linda; Welsh, Nils

    2002-01-01

    BACKGROUND: Recent reports identify the 3'-UTR of insulin mRNA as crucial for control of insulin messenger stability. This region contains a pyrimidine-rich sequence, which is similar to the hypoxia-responsive mRNA-stabilizing element of tyrosine hydroxylase. This study aimed to determine whether hypoxia affects insulin mRNA levels. MATERIALS AND METHODS: Rat islets were incubated at normoxic or hypoxic conditions and with or without hydrogen peroxide and a nitric oxide donor. Insulin mRNA was determined by Northern hybridization. Islet homogenates were used for electrophoretic mobility shift assay with an RNA-oligonucleotide, corresponding to the pyrimidine-rich sequence of the 3'-UTR of rat insulin I mRNA. The expression of reporter gene mRNA, in islets transfected with reporter gene constructs containing the wild-type or mutated insulin mRNA pyrimidine-rich sequences, was measured by semiquantitive RT-PCR. RESULTS: Insulin mRNA was increased in response to hypoxia. This was paralleled by increased binding of the polypyrimidine tract-binding protein (PTB) to the pyrimidine-rich sequence of the 3'-UTR of insulin mRNA, which was counteracted by hydrogen peroxide. The reporter gene mRNA level containing the wild-type binding site was not increased in response to hypoxia, but mutation of the site resulted in a destabilization of the mRNA. CONCLUSIONS: The complete understanding of different diabetic conditions requires the elucidation of mechanisms that control insulin gene expression. Our data show that hypoxia may increase insulin mRNA levels by promoting the binding of PTB to the insulin mRNA 3'-UTR. Hydrogen peroxide abolishes the hypoxic effect indicating involvement of reactive oxygen species and/or the redox potential in the oxygen-signaling pathway. PMID:12359957

  19. Identification and genetic mapping of a homeobox gene to the 4p16. 1 region of human chromosome 4

    SciTech Connect

    Stadler, H.S.; Padanilam, B.J.; Solursh, M. ); Buetow, K. ); Murray, J.C. )

    1992-12-01

    A human craniofacial cDNA library was screened with a degenerate oligonucleotide probe based on the conserved third helix of homeobox genes. From this screening, we identified a homeobox gene, H6, which shared only 57-65% amino acid identity to previously reported homeodomains. H6 was physically mapped to the 4P16.1 region by using somatic cell hybrids containing specific deletions of human chromosome 4. Linkage data from a single-stranded conformational polymorphism derived from the 3[prime] untranslated region of the H6 cDNA placed this homeobox gene more than 20 centimorgans proximal of the previously mapped HOX7 gene on chromosome 4. Identity comparisons of the H6 Homeodomain with previously reported homeodomains reveal the highest identities to be with the Nk class of homeobox genes in Drosophila melanogaster. 53 refs., 5 figs., 2 tabs.

  20. An AU-Rich Sequence Element (UUUN[A/U]U) Downstream of the Edited C in Apolipoprotein B mRNA Is a High-Affinity Binding Site for Apobec-1: Binding of Apobec-1 to This Motif in the 3′ Untranslated Region of c-myc Increases mRNA Stability

    PubMed Central

    Anant, Shrikant; Davidson, Nicholas O.

    2000-01-01

    Apobec-1, the catalytic subunit of the mammalian apolipoprotein B (apoB) mRNA-editing enzyme, is a cytidine deaminase with RNA binding activity for AU-rich sequences. This RNA binding activity is required for Apobec-1 to mediate C-to-U RNA editing. Filter binding assays, using immobilized Apobec-1, demonstrate saturable binding to a 105-nt apoB RNA with a Kd of ∼435 nM. A series of AU-rich templates was used to identify a high-affinity (∼50 nM) binding site of consensus sequence UUUN[A/U]U, with multiple copies of this sequence constituting the high-affinity binding site. In order to determine whether this consensus site could be functionally demonstrated from within an apoB RNA, circular-permutation analysis was performed, revealing one major (UUUGAU) and one minor (UU) site located 3 and 16 nucleotides, respectively, downstream of the edited base. Secondary-structure predictions reveal a stem-loop flanking the edited base with Apobec-1 binding to the consensus site(s) at an open loop. A similar consensus (AUUUA) is present in the 3′ untranslated regions of several mRNAs, including that of c-myc, that are known to undergo rapid degradation. In this context, it is presumed that the consensus motif acts as a destabilizing element. As an independent test of the ability of Apobec-1 to bind to this sequence, F442A cells were transfected with Apobec-1 and the half-life of c-myc mRNA was determined following actinomycin D treatment. These studies demonstrated an increase in the half-life of c-myc mRNA from 90 to 240 min in control versus Apobec-1-expressing cells. Apobec-1 expression mutants, in which RNA binding activity is eliminated, failed to alter c-myc mRNA turnover. Taken together, the data establish a consensus binding site for Apobec-1 embedded in proximity to the edited base in apoB RNA. Binding to this site in other target RNAs raises the possibility that Apobec-1 may be involved in other aspects of RNA metabolism, independent of its role as an apoB RNA

  1. Novel adenosine 3 prime ,5 prime -cyclic monophosphate dependent protein kinases in a marine diatom

    SciTech Connect

    Lin, P.P.C.; Volcani, B.E. )

    1989-08-08

    Two novel adenosine 3{prime},5{prime}-cyclic monophosphate (cAMP) dependent protein kinases have been isolated from the diatom Cylindrotheca fusiformis. The kinases, designated I and II, are eluted from DEAE-Sephacel at 0.10 and 0.15 M NaCl. They have a high affinity for cAMP and are activated by micromolar cAMP. They exhibit maximal activity at 5 mM Mg{sup 2+} and pH 8 with the preferred phosphate donor ATP and phosphate acceptor histone H1. They phosphorylate sea urchin sperm histone H1 on a single serine site in the sequence Arg-Lys-Gly-Ser({sup 32}P)-Ser-Asn-Ala-Arg and have an apparent M{sub r} of 75,000 as determined by gel filtration and sucrose density sedimentation. In the kinase I preparation a single protein band with an apparent M{sub r} of about 78,000 is photolabeled with 8-azido({sup 32}P)cAMP and is also phosphorylated with ({gamma}-{sup 32}P)ATP in a cAMP-dependent manner, after autoradiography following sodium dodecyl sulfate gel electrophoresis. The rate of phosphorylation of the 78,000-dalton band is independent of the enzyme concentration. The results indicate that (i) these diatom cAMP-dependent protein kinases are monomeric proteins, possessing both the cAMP-binding regulatory and catalytic domains on the same polypeptide chain, (ii) the enzymes do not dissociate into smaller species upon activation by binding cAMP, and (iii) self-phosphorylation of the enzymes by an intrapeptide reaction is cAMP dependent. The two diatom cAMP kinases are refractory to the heat-stable protein kinase modulator from rabbit muscle, but they respond differently to proteolytic degradation and to inhibition by arachidonic acid and several microbial alkaloids.

  2. A phylogenetically conserved sequence within viral 3' untranslated RNA pseudoknots regulates translation.

    PubMed Central

    Leathers, V; Tanguay, R; Kobayashi, M; Gallie, D R

    1993-01-01

    Both the 68-base 5' leader (omega) and the 205-base 3' untranslated region (UTR) of tobacco mosaic virus (TMV) promote efficient translation. A 35-base region within omega is necessary and sufficient for the regulation. Within the 3' UTR, a 52-base region, composed of two RNA pseudoknots, is required for regulation. These pseudoknots are phylogenetically conserved among seven viruses from two different viral groups and one satellite virus. The pseudoknots contained significant conservation at the secondary and tertiary levels and at several positions at the primary sequence level. Mutational analysis of the sequences determined that the primary sequence in several conserved positions, particularly within the third pseudoknot, was essential for function. The higher-order structure of the pseudoknots was also required. Both the leader and the pseudoknot region were specifically recognized by, and competed for, the same proteins in extracts made from carrot cell suspension cells and wheat germ. Binding of the proteins is much stronger to omega than the pseudoknot region. Synergism was observed between the TMV 3' UTR and the cap and to a lesser extent between omega and the 3' UTR. The functional synergism and the protein binding data suggest that the cap, TMV 5' leader, and 3' UTR interact to establish an efficient level of translation. Images PMID:8355685

  3. A T-cell specific transcriptional enhancer element 3 prime of C sub. alpha. in the human T-cell receptor. alpha. locus

    SciTech Connect

    Ho, Icheng; Yang, Lihsuan; Morle, G.; Leiden, J.M. )

    1989-09-01

    A transcriptional enhancer element has been identified 4.5 kilobases 3{prime} of C{sub {alpha}} (constant region {alpha} chain) in the human T-cell receptor (TCR) {alpha}-chain locus. This enhancer is active on both a TCR V{sub {alpha}} (variable region {alpha} chain) promoter and the minimal simian virus 40 promoter in TCR {alpha}/{beta} Jurkat and EL4 cells but is inactive on a V{sub {alpha}} promoter TCR {gamma}/{delta} PEER and Molt-13 cells, clone 13 B cells, and HeLa fibroblasts. The enhancer has been localized to a 116-base-pair BstXI/Dra I restriction enzyme fragment, which lacks immunoglobulin octamer and {kappa}B enhancer motifs but does contain a consensus cAMP-response element (CRE). DNase I footprint analyses demonstrated that the minimal enhancer contains two binding sites for Jurkat nuclear proteins. One of these sites corresponds to the CRE, while the other does not correspond to a known transcriptional enhancer motif. These data support a model in which TCR {alpha} gene transcription is regulated by a unique set of cis-acting sequences and trans-acting factors, which are differentially active in cells of the TCR {alpha}/{beta} lineage. In addition, the TCR {alpha} enhancer may play a role in activating oncogene expression in T-lymphoblastoid tumors that have previously been shown to display chromosomal translocations into the human TCR {alpha} locus.

  4. Trans-activation function of a 3 prime truncated X gene-cell fusion product from integrated hepatitis B virus DNA in chronic hepatitis tissues

    SciTech Connect

    Takada, Shinako; Koike, Katsuro )

    1990-08-01

    To investigate the expression and transactivation function of the X gene in integrated hepatitis B virus (HBV) DNA from chronic hepatitis tissues, a series of transfectants containing cloned integrated HBV DNAs was made and analyzed for X mRNA expression and trans-activation activity by using a chloramphenicol acetyltransferase assay. Most of the integrated HBV DNAs expressed X mRNA and encoded a product with trans-activation activity in spite of the loss of the 3{prime} end region of the X gene due to integration. From cDNA cloning and sequence analysis of X mRNA transcribed from native or integrated HBV DNA, the X protein was found to be translated from the X open reading frame without splicing. For integrated HBV DNA, transcription was extended to a cellular flanking DNA and an X gene-cell fusion transcript was terminated by using a cellular poly(A) signal. The amino acid sequence deduced from an X-cell fusion transcript indicated truncation of the carboxyl-terminal five amino acids, but the upstream region of seven amino acids conserved among hepadnaviruses was retained in the integrated HBV DNA, suggesting that this conserved region is essential for the transactivation function of the X protein. These findings support the following explanation for hepatocarcinogenesis by HBV DNA integration: the expression of a cellular oncogene(s) is transactivated at the time of chronic infection by the increasing amounts of the integrated HBV gene product(s), such as the X-cell fusion product.

  5. Genetic variation at the ApoB 3[prime] HVR, D2S44, and D7S21 loci in the Ewondo ethnic group of Cameroon

    SciTech Connect

    Destro-Bisol, G.; d'Aloja, E.; Dobosz, M.; Pascali, V.L. ); Spedini, G. ); Presciuttini, S. )

    1994-07-01

    A sample of the Ewondo population (a Bantu-speaking group of Southern Cameroon) was analyzed for the polymorphism at three tandem repeated DNA loci (ApoB 3[prime] HVR, D2S44, and D7S21). The authors observed a greater number of ApoB 3[prime] HVR alleles (17) and a significantly higher estimated heterozygosity (.879[+-].011) than in previously surveyed populations, with the exception of U.S. Blacks. The higher genetic variability of Ewondo and U.S. Blacks was also shown by the ApoB 3[prime] HVR allele-frequency spectra. A method for measuring population distances, based on cumulative fragment-size distribution, is described. Interpopulation comparisons for ApoB 3[prime] HVR were carried out by this method and were compared with those obtained by a genetic distance measurement. The two sets of results showed a consistent pattern of population differentiation: the Ewondos and the U.S. Blacks clustered together and were apart from both a Caucasian cluster (Swedes, U.S. Whites, Italians, and Germans) and other well-defined populations (Sikhs of India and Pehuence Indians of Chile). Profile distances were then computed from D2S44 and D7S21 binned data. This analysis indicated a genetic affinity between Ewondos, U.S. Blacks, and Afro-Caribbean Blacks and outlined the genetic diversity between Ewondos, Caucasians, and Asian Indians. 56 refs., 5 figs., 1 tab.

  6. Cortisol metabolism in hepatocytes of rainbow trout treated with 3,3{prime},4,4{prime} tetrachlorobiphenyl

    SciTech Connect

    Vijayan, M.M.; Fiest, G.; Otto, D.; Moon, T.W.

    1995-12-31

    The objective of this study was to investigate the potential of hepatocytes for cortisol uptake and metabolism in 3,3{prime},4,4{prime}-tetrachlorobiphenyl (TCBP) treated trout. Two groups of rainbow trout (Oncorhynchus mykiss) were either given an intraperitoneal implant of peanut oil alone or peanut oil containing TCBP (10 mg.kg{sup {minus}1} body weight) and sampled six weeks later. The toxicant exposed fish had significantly lower condition factor and plasma glucose concentration, whereas plasma cortisol, protein and hepatocyte protein concentration and liver ethoxyresorufin-O-deethylase (EROD) activity were significantly higher in the TCBP compared to the sham group. There was no significant difference in plasma lactate and amino acid concentration, hepatocyte glycogen content or liver cytosolic cortisol binding affinity or capacity between the two groups. The uptake of [{sup 3}H] cortisol was significantly higher in the hepatocytes of TCBP treated fish compared to the sham fish. Also, there was enhanced catabolism of [{sup 3}H] cortisol by hepatocytes of TCBP treated fish; the major metabolite appeared to be tetrahydrocortisone. The results indicate that the potential for cortisol clearance is enhanced in hepatocytes of TCBP treated trout. The data also tend to suggest in vivo regulatory mechanisms that might possibly prevent the increased clearance of the hormone from circulation in toxicant exposed fish.

  7. Construction of a 2.8-megabase yeast artificial chromosome contig and cloning of the human methylthioadenosine phosphorylase gene from the tumor suppressor region on 9p21

    SciTech Connect

    Olopade, O.I.; Pomykala, H.M.; Hagos, F.

    1995-07-03

    Many human malignant cells lack methylthioadenosine phosphorylase (MTAP) enzyme activity. The gene (MTAP) encoding this enzyme was previously mapped to the short arm of chromosome 9, band p21-22, a region that is frequently deleted in multiple tumor types. To clone candidate tumor suppressor genes from the deleted region on 9p21-22, we have constructed a long-range physical map of 2.8 megabases for 9p21 by using overlapping yeast artificial chromosome and cosmid clones. This map includes the type I IFN gene cluster, the recently identified candidate tumor suppressor genes CDKN2 (p16{sup INK4A}) and CDKN2B (p15{sup INK4B}), and several CpG islands. In addition, we have identified other transcription units within the yeast artificial chromosome contig. Sequence analysis of a 2.5-kb cDNA clone isolated from a CpG island that maps between the IFN genes and CDKN2 reveals a predicted open reading frame of 283 amino acids followed by 1302 nucleotides of 3{prime} untranslated sequence. This gene is evolutionarily conserved and shows significant amino acid homologies to mouse and human purine nucleoside phosphorylases and to a hypothetical 25.8-kDa protein in the pet gene (coding for cytochrome bc{sub 1} complex) region of Rhodospirillum rubrum. The location, expression pattern, and nucleotide sequences of this gene suggest that it codes for the MTAP enzyme. 35 refs., 4 figs., 1 tab.

  8. Exclusion of candidate genes from the chromosome 1q juvenile glaucoma region and mapping of the peripheral cannabis receptor gene (CNR2) to chromosome 1

    SciTech Connect

    Sunden, S.L.F.; Nichols, B.E.; Alward, W.L.M.

    1994-09-01

    Juvenile onset primary open angle glaucoma has been mapped by linkage to 1q21-q31. Several candidate genes were evaluated in the same family used to identify the primary linkage. Atrionatriuretic peptide receptor A (NPR1) and laminin C1 (LAMC1) have been previously mapped to this region and could putatively play a role in the pathogenesis of glaucoma. A third gene, the peripheral cannabis receptor (CNR2) was not initially mapped in humans but was a candidate because of the relief that cannabis affords some patients with primary open angle glaucoma. Microsatellites associated with NPR1 and LAMC1 revealed multiple recombinations in affected members of this pedigree. CNR2 was shown to be on chromosome 1 by PCR amplification of a 150 bp fragment of the 3{prime} untranslated region in monochromosomal somatic cell hybrids (NIGMS panel No. 2). These primers also revealed a two allele single strand conformation polymorphism which showed multiple recombinants with juvenile onset primary open angle glaucoma in large pedigrees, segregating this disorder. The marker was then mapped to 1p34-p36 by linkage, with the most likely location between liver alkaline phosphatase (ALPL) and alpha-L-1 fucosidase (FUCA1).

  9. 3' Untranslated regions mediate transcriptional interference between convergent genes both locally and ectopically in Saccharomyces cerevisiae.

    PubMed

    Wang, Luwen; Jiang, Ning; Wang, Lin; Fang, Ou; Leach, Lindsey J; Hu, Xiaohua; Luo, Zewei

    2014-01-01

    Paired sense and antisense (S/AS) genes located in cis represent a structural feature common to the genomes of both prokaryotes and eukaryotes, and produce partially complementary transcripts. We used published genome and transcriptome sequence data and found that over 20% of genes (645 pairs) in the budding yeast Saccharomyces cerevisiae genome are arranged in convergent pairs with overlapping 3'-UTRs. Using published microarray transcriptome data from the standard laboratory strain of S. cerevisiae, our analysis revealed that expression levels of convergent pairs are significantly negatively correlated across a broad range of environments. This implies an important role for convergent genes in the regulation of gene expression, which may compensate for the absence of RNA-dependent mechanisms such as micro RNAs in budding yeast. We selected four representative convergent gene pairs and used expression assays in wild type yeast and its genetically modified strains to explore the underlying patterns of gene expression. Results showed that convergent genes are reciprocally regulated in yeast populations and in single cells, whereby an increase in expression of one gene produces a decrease in the expression of the other, and vice-versa. Time course analysis of the cell cycle illustrated the functional significance of this relationship for the three pairs with relevant functional roles. Furthermore, a series of genetic modifications revealed that the 3'-UTR sequence plays an essential causal role in mediating transcriptional interference, which requires neither the sequence of the open reading frame nor the translation of fully functional proteins. More importantly, transcriptional interference persisted even when one of the convergent genes was expressed ectopically (in trans) and therefore does not depend on the cis arrangement of convergent genes; we conclude that the mechanism of transcriptional interference cannot be explained by the transcriptional collision model, which postulates a clash between simultaneous transcriptional processes occurring on opposite DNA strands.

  10. Dynein light chain binding to a 3′-untranslated sequence mediates parathyroid hormone mRNA association with microtubules

    PubMed Central

    Epstein, Eyal; Sela-Brown, Alin; Ringel, Israel; Kilav, Rachel; King, Stephen M.; Benashski, Sharon E.; Yisraeli, Joel K.; Silver, Justin; Naveh-Many, Tally

    2000-01-01

    The 3′-untranslated region (UTR) of mRNAs binds proteins that determine mRNA stability and localization. The 3′-UTR of parathyroid hormone (PTH) mRNA specifically binds cytoplasmic proteins. We screened an expression library for proteins that bind the PTH mRNA 3′-UTR, and the sequence of 1 clone was identical to that of the dynein light chain LC8, a component of the dynein complexes that translocate cytoplasmic components along microtubules. Recombinant LC8 binds PTH mRNA 3′-UTR, as shown by RNA electrophoretic mobility shift assay. We showed that PTH mRNA colocalizes with microtubules in the parathyroid gland, as well as with a purified microtubule preparation from calf brain, and that this association was mediated by LC8. To our knowledge, this is the first report of a dynein complex protein binding an mRNA. The dynein complex may be the motor that is responsible for transporting mRNAs to specific locations in the cytoplasm and for the consequent is asymmetric distribution of translated proteins in the cell. PMID:10683380

  11. A mutation in an alternative untranslated exon of hexokinase 1 associated with Hereditary Motor and Sensory Neuropathy – Russe (HMSNR)

    PubMed Central

    Hantke, Janina; Chandler, David; King, Rosalind; Wanders, Ronald JA; Angelicheva, Dora; Tournev, Ivailo; McNamara, Elyshia; Kwa, Marcel; Guergueltcheva, Velina; Kaneva, Radka; Baas, Frank; Kalaydjieva, Luba

    2009-01-01

    Hereditary Motor and Sensory Neuropathy – Russe (HMSNR) is a severe autosomal recessive disorder, identified in the Gypsy population. Our previous studies mapped the gene to 10q22-q23 and refined the gene region to ∼70 kb. Here we report the comprehensive sequencing analysis and fine mapping of this region, reducing it to ∼26 kb of fully characterised sequence spanning the upstream exons of Hexokinase 1 (HK1). We identified two sequence variants in complete linkage disequilibrium, a G>C in a novel alternative untranslated exon (AltT2) and a G>A in the adjacent intron, segregating with the disease in affected families and present in the heterozygote state in only 5/790 population controls. Sequence conservation of the AltT2 exon in 16 species with invariable preservation of the G allele at the mutated site, strongly favour the exonic change as the pathogenic mutation. Analysis of the Hk1 upstream region in mouse mRNA from testis and neural tissues showed an abundance of AltT2-containing transcripts generated by extensive, developmentally regulated alternative splicing. Expression is very low compared with ubiquitous Hk1 and all transcripts skip exon1, which encodes the protein domain responsible for binding to the outer mitochondrial membrane, and regulation of energy production and apoptosis. Hexokinase activity measurement and immunohistochemistry of the peripheral nerve showed no difference between patients and controls. The mutational mechanism and functional effects remain unknown and could involve disrupted translational regulation leading to increased anti-apoptotic activity (suggested by the profuse regenerative activity in affected nerves), or impairment of an unknown HK1 function in the peripheral nervous system (PNS). PMID:19536174

  12. Translation of the Saccharomyces cerevisiae tcm1 gene in the absence of a 5'-untranslated leader.

    PubMed Central

    Maicas, E; Shago, M; Friesen, J D

    1990-01-01

    The role of eukaryotic 5'-untranslated messenger RNA leaders is not entirely clear, since they share little sequence similarity among each other. The importance of the leader in determining the efficiency of translation initiation was addressed here by examining the polyribosome distribution of several leader-deletion alleles of the yeast tcm1 gene (coding for ribosomal protein L3). Shortening of this 22-nucleotide leader, or complete removal of it (the first nucleotide of the mRNA becoming the A of the translation initiation codon AUG) permitted translation, albeit reduced. Further deletion of as few as the first two nucleotides of the initiation codon leads to a substantial reduction in ribosome loading, which is compatible with inefficient initiation at the next downstream, out-of-frame, AUG triplet. A second measure of translation initiation was obtained by assaying qualitatively for the production of biologically active L3 protein using growth-resistance to trichodermin. This experiment indicates that ribosomes can recognize the correct initiation codon even in the complete absence of a leader. We conclude that the 5'-untranslated leader of the yeast tcm1 gene is not essential for accurate translation initiation, but enhances its efficiency. Images PMID:2216774

  13. A{sup -2} {yields} G transition at the 3{prime} acceptor splice site of IVS17 characterizes the COL2A1 gene mutation in the original Stickler syndrome kindred

    SciTech Connect

    Williams, C.J.; Ganguly, A.; Considine, E.

    1996-06-14

    Hereditary progressive arthro-ophthalmopathy, or {open_quotes}Stickler syndrome,{close_quotes} is an autosomal dominant osteochondrodysplasia characterized by a variety of ocular and skeletal anomalies which frequently lead to retinal detachment and precocious osteoarthritis. A variety of mutations in the COL2A1 gene have been identified in {open_quotes}Stickler{close_quotes} families; in most cases studied thus far, the consequence of mutation is the premature generation of a stop codon. We report here the characterization of a COL2A1 gene mutation in the original kindred described by Stickler et al. Conformational sensitive gel electrophoresis (CSGE) was used to screen for mutations in the entire COL2A1 gene in an affected member from the kindred. A prominent heteroduplex species was noted in the polymerase chain reaction (PCR) product from a region of the gene including exons 17 to 20. Direct sequencing of PCR-amplified genomic DNA resulted in the identification of a base substitution at the A{sup -2} position of the 3{prime} splice acceptor site of IVS17. Sequencing of DNA from affected and unaffected family members confirmed that the mutation segregated with the disease phenotype. Reverse transcriptase-PCR analysis of poly A+ RNA demonstrated that the mutant allele utilized a cryptic splice site in exon 18 of the gene, eliminating 16 bp at the start of exon 18. This frameshift eventually results in a premature termination codon. These findings are the first report of a splice site mutation in classical Stickler syndrome and they provide a satisfying historical context in which to view COL2A1 mutations in this dysplasia. 25 refs., 3 figs., 1 tab.

  14. Mixed-function oxidase enzyme activity and oxidative stress in lake trout (Salvelinus namaycush) exposed to 3,3{prime},4,4{prime}5-pentachlorobiphenyl (PCB-126)

    SciTech Connect

    Palace, V.P.; Klaverkamp, J.F.; Lockhart, W.L. |; Metner, D.A.; Muir, D.C.G.; Brown, S.B.

    1996-06-01

    Juvenile lake trout were intraperitoneally injected with corn oil containing nominal concentrations of 0, 0.6, 6.3, or 25 {micro}g [{sup 14}C]-3,3{prime},4,4{prime},5-pentachlorobiphenyl (PCB-126) per gram of body weight. The PCB-126 accumulated in liver in a dose-dependent manner to a sustained concentration by 6 weeks and remained elevated for the 30-week experimental period. Mixed-function oxidase (MFO) enzyme activity was elevated in the two highest dose groups relative to the control group, but not in the low-dose group throughout the 30 weeks. Oxidative stress, measured by the thiobarbituric acid reactive substances test, was correlated with ethoxyresorufin O-deethylase and was elevated in liver of the two highest PCB dose groups but not the low-dose group. The activities of the enzymatic antioxidants superoxide dismutase, catalase, and glutathione peroxidase were unaffected by PCB-126 exposure. The nonenzymatic antioxidants superoxide dismutase, catalase, and glutathione peroxidase were unaffected by PCB-126 exposure. The nonenzymatic antioxidant tocopherol was depleted to approximately 75% of the control concentration in liver of all three PCB-dosed groups. Hepatic ascorbic acid levels were not different in any of the treatment groups. Retinol was depleted by greater than an order of magnitude in liver of the two highest dose groups but not in the los-dose group. This study demonstrates a correlation between hepatic MFO activity and oxidative stress in PCB-exposed lake trout. Tocopherol and retinol may be important mediators of oxidative stress but additional study is required to confirm the antioxidant activity of retinol.

  15. Nuclear Factor E2-Related Factor-2 Negatively Regulates NLRP3 Inflammasome Activity by Inhibiting Reactive Oxygen Species-Induced NLRP3 Priming

    PubMed Central

    Liu, Xiuting; Zhang, Xin; Ding, Yang; Zhou, Wei; Tao, Lei; Lu, Ping; Wang, Yajing

    2017-01-01

    Abstract Aims: The NLRP3 inflammasome is a multiprotein complex that protects hosts against a variety of pathogens. However, the molecular mechanisms of modulating NLRP3 inflammasome activation, especially at the priming step, are still poorly understood. This study was designed to elucidate the negative regulation of nuclear factor E2-related factor-2 (Nrf2) on the activation of NLRP3 inflammasome. Results: We reported that Nrf2 activation inhibited NLRP3 expression, caspase-1 cleavage, and subsequent IL-1β generation. Compared with normal cells, Nrf2-deficient cells showed upregulated cleaved caspase-1, which were attributed to the increased transcription of NLRP3 caused by excess reactive oxygen species (ROS). Furthermore, priming of the NLRP3 inflammasome was sensitive to the exogenous ROS levels induced by H2O2 or rotenone. Combined with adenosine triphosphate, rotenone triggered higher activity of the NLRP3 inflammasome compared with lipopolysaccharide, suggesting that ROS promoted the priming step. In addition, Nrf2-induced NQO1 was involved in the inhibition of the NLRP3 inflammasome. In an in vivo alum-induced peritonitis mouse model, Nrf2 activation suppressed typical IL-1 signaling-dependent inflammation, whereas Nrf2−/− mice exhibited a significant increase in the recruitment of immune cell and the generation of IL-1β compared with wild-type mice. Innovation: We elucidated the effects and possible mechanisms of Nrf2 activation-induced NQO1 expression on NLRP3 inflammasome inactivation and established a novel regulatory role of the Nrf2 pathway in ROS-induced NLRP3 priming. Conclusions: We demonstrated Nrf2 negatively regulating NLRP3 inflammasome activity by inhibiting the priming step and suggested that Nrf2 could be a potential target for some uncontrolled inflammasome activation-associated diseases. Antioxid. Redox Signal. 26, 28–43. PMID:27308893

  16. Lack of developmental and reproductive toxicity of 2,3,3{prime},4,4{prime}-pentachlorobiphenyl (PCB 105) in ring-necked pheasants

    SciTech Connect

    Hornung, M.W.; Miller, L.; Peterson, R.E.; Melancon, M.

    1995-12-31

    One of these PCBs, 2,3,3{prime},4,4{prime}-pentachlorobiphenyl (PCB 105) has the potential to produce toxicity by an Ah receptor-mediated mechanism. To determine the potency of PCB 105 for producing reproductive and developmental toxicity, adult ring-necked pheasant hens were orally dosed with 0, 0.06, 0.6 or 6 mg PCB 105/kg hen/week for 10 weeks after which hens were bred with control roosters once per week for 8 weeks. Eggs were collected daily and incubated until hatched, or for 28 days, after which embryo development was evaluated. Fertilized egg production, embryo mortality and chick mortality were not significantly different between treatment groups, nor were total body, liver and heart weights of chicks 1 day post-hatch (dph). To determine whether signs of PCB 105 toxicity were delayed, the first chick to hatch from each hen was evaluated at 21 dph for signs of toxicity. Chick total body, liver and heart weights at 21 dph were not significantly different between treatment groups. Three hepatic microsomal monooxygenase activities were significantly elevated in 1 day old chicks from hens given a cumulative PCB 105 dose of 6 mg/kg and in 21 day old chicks from hens given a cumulative PCB dose of 60 mg/kg as compared to respective control chicks. These results indicate that a cumulative PCB 105 dose up to 60 mg/kg hen does not decrease the production of fertilized eggs or increase embryo or chick mortality in ring-necked pheasants, but does increase chick hepatic monooxygenase activity.

  17. Effects of pre- and postnatal exposure to 3,3[prime],4,4-[prime],5-pentachlorobiphenyl on physical development, neurobehavior and xenobiotic metabolizing enzymes in rats

    SciTech Connect

    Bernhoft, A.; Nafstad, I.; Engen, P. ); Skaare, J.U. )

    1994-10-01

    An experiment was conducted to study the effects of the coplanar non-ortho-chlorinated congener 3,3[prime],4,4[prime],5-pentachlorobiphenyl (PCB-126) in rats exposed during fetal development and postnatal suckling period. Two groups of eight dams were administered by gavage six doses of 10 and 20 [mu]g/kg body weight of PCB-126 dissolved in corn oil every second day from days 9 to 19 of gestation. The corresponding control rats were treated with corn oil only. The physical development of the offspring was observed. The effects of PCB-126 on hepatic xenobiotic metabolizing enzyme activities and the concentrations of PCB in the liver and brain were investigated in samples from pups of different age and from their mothers. The litter size, the body weights, and the survival of the exposed sucklings were reduced, and the onset of spontaneous movement and neuromuscular maturation were delayed, whereas the development of reflexes was not affected. The body weight was still reduced in a dose-related manner up to 18 weeks postpartum. Also, the postpartum body weight of the PCB-exposed mothers was reduced as compared to controls, but the difference disappeared at weaning. The hepatic enzyme activities of cytochrome P450 1A1 examined by ethoxyresorufin O-deethylase (EROD) and glutathione S-transferase (GST) toward 1-chloro-2,4-dinitrobenzene (CDNB) were increased in both the exposed pups and their mothers, and the relative liver weight was increased in the exposed pups. Hepatic PCB-126 residues were detected in samples collected throughout the experiment, whereas no detectable concentration was found in the brain. The authors conclude that exposure of this PCB congener in utero and through lactation showed fetotoxic effects, delayed physical maturation, and induced liver xenobiotic metabolizing enzymes without causing neurobehavioral effects.

  18. Tributyltin potentiates 3,3{prime},4,4{prime},5-pentachlorobiphenyl-induced cytochrome P-4501A-related activity

    SciTech Connect

    DeLong, G.T.; Rice, C.D.

    1997-10-01

    Induction of cytochrome P-4501A protein and induction of related enzyme activity are hallmark physiological responses following exposure to planar halogenated aromatic hydrocarbons (HAHs) such as 3,3{prime},4,4{prime},5-pentachlorobiphenyl (PCB 126; PeCB). Environments contaminated by HAHs are often contaminated by mixtures of anthropogenic contaminants, including organometallic compounds. Both HAHs and organometallics easily bioconcentrate in aquatic food chains that may be linked to humans through seafood consumption. Tributyltin (TBT), a marine biocide, has been detected in many aquatic environments. Exposure to TBT, as well as several PCBs, has been associated with immunotoxicity, neurotoxicity, and endocrine disruption. Recently TBT has been shown to inhibit cytochrome P-4501A activity in vitro. Female mice were exposed to 0.07, 0.1, and 1.0 mg/kg PeCB, TBT or both. P-4501A levels and BaP-OHase activity were significantly elevated in mice exposed to PeCB alone. This effect was enhanced by coexposure to low levels of TBT; PeCB-induced P-4501A-related activity was potentiated at the low range of each. The highest dose of TBT, however, inhibited these activities when given in combination with PeCB. Thymic atrophy was evident only in mice exposed daily to 0.1 and 1.0 mg/kg PeCB alone, or to a combination of the lowest and highest dose of PeCB and TBT, respectively. Because environmental levels of. TBT are not expected to be as high as the highest level used in our toxicological studies, we conclude that environmental exposure to TBT may potentiate, rather than inhibit, the activity of environmental levels of HAHs that are associated with P-4501A induction. 31 refs., 8 figs.

  19. HPLC determination of tocopherol, retinol, dehydroretinol and retinyl palmitate in tissues of Lake Char (Salvelinus namaycush) exposed to coplanar 3,3[prime],4,4[prime],5-pentachlorobiphenyl

    SciTech Connect

    Palace, V.P. . Dept. of Zoology); Brown, S.B. . Dept. of Fisheries and Oceans)

    1994-03-01

    Tocopherol, retinol, dehydroretinol, and retinyl palmitate were measured by reversed-phase HPLC in liver, kidney, and plasma of lake char exposed to orally administered coplanar 3,3[prime],4,4[prime],5-pentachlorobiphenyl (PCB). Tocopherol concentrations were unaffected after eight weeks. Liver retinol, dehydroretinol, and retinyl palmitate concentrations were lower, whereas kidney retinyl palmitate was elevated in PCB-exposed groups. Tissue retinoid concentrations provide sensitive indicators of coplanar PCB exposure in fish.

  20. Stable, Microfabricated Thin Layer Chromatography Plates without Volume Distortion on Patterned, Carbon and Al2O3-Primed Carbon Nanotube Forests

    SciTech Connect

    Jensen, David S.; Kanyal, Supriya S.; Gupta, Vipul; Vail, Michael A.; Dadson, Andrew; Engelhard, Mark H.; Vanfleet, Richard; Davis, Robert C.; Linford, Matthew R.

    2012-09-28

    In a recent report (Song, J.; et al., Advanced Functional Materials 2011, 21, 1132-1139) some of us described the fabrication of thin layer chromatography (TLC) plates from patterned carbon nanotube (CNT) forests, which were directly infiltrated/coated with silicon by low pressure chemical vapor deposition (LPCVD) of silicon using SiH4. Following infiltration, the nanotubes were removed from the assemblies and the silicon simultaneously converted to SiO2 in a high temperature oxidation step. However, while straightforward, this process had some shortcomings, not the least of which was some distortion of the lithographically patterned features during the volume expansion that accompanied oxidation. Herein we overcome theis issue and also take substantial steps forward in the microfabrication of TLC plates by showing: (i) A new method for creating an adhesion promotion layer on CNT forests by depositing a few nanometers of carbon followed by atomic layer deposition (ALD) of Al2O3. This method for appears to be new, and X-ray photoelectron spectroscopy confirms the expected presence of oxygen after carbon deposition. ALD of Al2O3 alone and in combination with the carbon on patterned CNT forests was also explored as an adhesion promotion layer for CNT forest infiltration. (ii) Rapid, conformal deposition of an inorganic material that does not require subsequent oxidation: fast pseudo-ALD growth of SiO2 via alumina catalyzed deposition of tris(tert-butoxy)silanol onto the carbon/Al2O3-primed CNT forests. (iii) Faithful reproduction of the features in the masks used to microfabricate the TLC plates (M-TLC) this advance springs from the previous two points. (iv) A bonded (amino) phase on a CNT-templated microfabricated TLC plate. (v) Fast, highly efficient (125,000 - 225,000 N/m) separations of fluorescent dyes on M-TLC plates. (vi) Extensive characterization of our new materials by TEM, SEM, EDAX, DRIFT, and XPS. (vii) A substantially lower process temperature for the

  1. Genetic Variation in the 3'-Untranslated Region of NBN Gene Is Associated with Gastric Cancer Risk in a Chinese Population

    PubMed Central

    Zhu, Xun; Ren, Chuanli; Xie, Lan; Dai, Ningbin; Gu, Yayun; Yan, Caiwang; Dai, Juncheng; Ma, Hongxia; Jiang, Yue; Chen, Jiaping; Hu, Zhibin; Shen, Hongbing; Wu, Haorong; Jin, Guangfu

    2015-01-01

    NBN plays a crucial role in carcinogenesis as a core component for both homologous recombination (HR) and non-homologous end-joining (NHEJ) DNA double-strand breaks (DSBs) repair pathways. Genetic variants in the NBN gene have been associated with multiple cancers risk, suggesting pleiotropic effect on cancer. We hypothesized that genetic variants in the NBN gene may modify the risk of gastric cancer. To test this hypothesis, we evaluated the association between four potentially functional single nucleotide polymorphisms in NBN and gastric cancer risk in a case–control study of 1,140 gastric cancer cases and 1,547 controls in a Chinese population. We found that the A allele of rs10464867 (G>A) was significantly associated with a decreased risk of gastric cancer (odds ratio [OR] = 0.81, 95% confidence interval [95% CI] = 0.71–0.94; P = 4.71×10−3). Furthermore, the association between A allele of rs10464867 and decreased risk of gastric cancer was more significantly in elder individuals (per-allele OR = 0.72[0.59–0.88], P = 1.07×10−3), and male individuals (per-allele OR = 0.73[0.62–0.87], P = 3.68×10−4). We further conducted a haplotype analysis and identified that the NBN Ars10464867Grs14448Grs1063053 haplotype conferred stronger protective effect on gastric cancer (OR = 0.76[0.65–0.89], P = 6.39×10−4). In summary, these findings indicate that genetic variants at NBN gene may contribute to gastric cancer susceptibility and may further advance our understanding of NBN gene in cancer development. PMID:26402912

  2. Widespread Shortening of 3' Untranslated Regions and Increased Exon Inclusion Are Evolutionarily Conserved Features of Innate Immune Responses to Infection.

    PubMed

    Pai, Athma A; Baharian, Golshid; Pagé Sabourin, Ariane; Brinkworth, Jessica F; Nédélec, Yohann; Foley, Joseph W; Grenier, Jean-Christophe; Siddle, Katherine J; Dumaine, Anne; Yotova, Vania; Johnson, Zachary P; Lanford, Robert E; Burge, Christopher B; Barreiro, Luis B

    2016-09-01

    The contribution of pre-mRNA processing mechanisms to the regulation of immune responses remains poorly studied despite emerging examples of their role as regulators of immune defenses. We sought to investigate the role of mRNA processing in the cellular responses of human macrophages to live bacterial infections. Here, we used mRNA sequencing to quantify gene expression and isoform abundances in primary macrophages from 60 individuals, before and after infection with Listeria monocytogenes and Salmonella typhimurium. In response to both bacteria we identified thousands of genes that significantly change isoform usage in response to infection, characterized by an overall increase in isoform diversity after infection. In response to both bacteria, we found global shifts towards (i) the inclusion of cassette exons and (ii) shorter 3' UTRs, with near-universal shifts towards usage of more upstream polyadenylation sites. Using complementary data collected in non-human primates, we show that these features are evolutionarily conserved among primates. Following infection, we identify candidate RNA processing factors whose expression is associated with individual-specific variation in isoform abundance. Finally, by profiling microRNA levels, we show that 3' UTRs with reduced abundance after infection are significantly enriched for target sites for particular miRNAs. These results suggest that the pervasive usage of shorter 3' UTRs is a mechanism for particular genes to evade repression by immune-activated miRNAs. Collectively, our results suggest that dynamic changes in RNA processing may play key roles in the regulation of innate immune responses.

  3. Acute toxicity of cadmium, copper, zinc, ammonia, 3,3 prime -dichlorobenzidine, 2,6-dichloro-4-nitroaniline, methylene chloride, and 2,4,6-trichlorophenol to juvenile grass shrimp and killifish

    SciTech Connect

    Burton, D.T.; Fisher, D.J. )

    1990-05-01

    The acute toxicity of several compounds was investigated while performing a toxicity evaluation of a complex chemical effluent. The tests were conducted for one or more of the following reasons: (1) data were not available for the chemical; (2) data were not available for the species; or (3) data were not available for the juvenile life stage of the species. Forty-eight hour acute toxicity tests were run on juvenile grass shrimp (Palaemonetes pugio) and juvenile killifish (Fundulus heteroclitus) exposed to the following compounds: cadmium, copper, zinc, ammonia, 3,3{prime}-dichlorobenzidine, 2,6-dichloro-4-nitroaniline, methylene chloride (dichloromethane) and 2,4,6-trichlorophenol.

  4. Breed-dependent transcriptional regulation of 5'-untranslated GR (NR3C1) exon 1 mRNA variants in the liver of newborn piglets.

    PubMed

    Zou, Huafeng; Li, Runsheng; Jia, Yimin; Yang, Xiaojing; Ni, Yingdong; Cong, Rihua; Soloway, Paul D; Zhao, Ruqian

    2012-01-01

    Glucocorticoids are vital for life and regulate an array of physiological functions by binding to the ubiquitously expressed glucocorticoid receptor (GR, also known as NR3C1). Previous studies demonstrate striking breed differences in plasma cortisol levels in pigs. However, investigation into the breed-dependent GR transcriptional regulation is hampered by lacking porcine GR promoter information. In this study, we sequenced 5.3 kb upstream of the translation start codon of the porcine GR gene, and identified seven alternative 5'-untranslated exons 1-4, 1-5, 1-6, 1-7, 1-8, 1-9,10 and 1-11. Among all these mRNA variants, exons 1-4 and 1-5, as well as the total GR were expressed significantly (P<0.05) higher in the liver of newborn piglets of Large White (LW) compared with Erhualian, a Chinese indigenous breed. Overall level of CpG methylation in the region flanking exons 1-4 and 1-5 did not show breed difference. However, nuclear content of Sp1, p-CREB and GR in the liver was significantly (P<0.05) higher in LW piglets, associated with enhanced binding of p-CREB, and higher level of histone H3 acetylation in 1-4 and 1-5 promoters. In contrast, GR binding to promoters of exons 1-4 and 1-5 was significantly diminished in LW piglets, implicating the presence of negative GREs. These results indicate that the difference in the hepatic expression of GR transcript variants between two breeds of pigs is determined, at least partly, by the disparity in the binding of transcription factors and the enrichment of histone H3 acetylation to the promoters.

  5. Sequences of the 5' portion of the human c-sis gene: characterization of the transcriptional promoter and regulation of expression of the protein product by 5' untranslated mRNA sequences.

    PubMed Central

    Ratner, L; Thielan, B; Collins, T

    1987-01-01

    The c-sis gene encodes the B polypeptide chain of platelet-derived growth factor (PDGF), and is expressed in a number of normal and pathological conditions. In order to study the control of synthesis of the human c-sis product, we have initiated a study of two regions of this genetic locus which regulate transcription and translation. A clone of the 5' portion of the gene was obtained which included 1361 nucleotides upstream of the RNA initiation site. Transcriptional promoter activity of this region was demonstrated in normal and transformed cells using a plasmid with the sequences upstream of the c-sis RNA initiation site fused to an indicator gene, chloramphenicol acetyl transferase. Experiments were also performed to identify other possible regulatory regions of the c-sis gene. These data demonstrated that a portion of the c-sis first exon encoding the 5' untranslated region of the c-sis mRNA inhibited synthesis of the PDGF B product in vitro. These results define regions of the c-sis gene whose activity may be important in the regulation of transcription and translation under normal conditions and in the pathogenesis several human diseases. Images PMID:3627977

  6. Translation of vph mRNA in Streptomyces lividans and Escherichia coli after removal of the 5' untranslated leader.

    PubMed

    Wu, C J; Janssen, G R

    1996-10-01

    The Streptomyces vinaceus viomycin phosphotransferase (vph) mRNA contains an untranslated leader with a conventional Shine-Dalgarno homology. The vph leader was removed by ligation of the vph coding sequence to the transcriptional start site of a Streptomyces or an Escherichia coli promoter, such that transcription would initiate at the first position of the vph start codon. Analysis of mRNA demonstrated that transcription initiated primarily at the A of the vph AUG translational start codon in both Streptomyces lividans and E. coli; cells expressing the unleadered vph mRNA were resistant to viomycin indicating that the Shine-Dalgarno sequence, or other features contained within the leader, was not necessary for vph translation. Addition of four nucleotides (5'-AUGC-3') onto the 5' end of the unleadered vph mRNA resulted in translation initiation from the vph start codon and the AUG triplet contained within the added sequence. Translational fusions of vph sequence to a Tn5 neo reporter gene indicated that the first 16 codons of vph coding sequence were sufficient to specify the translational start site and reading frame for expression of neomycin resistance in both E. coli and S. lividans.

  7. Hydrothermal syntheses, crystal structures and luminescence properties of zinc(II) and cadmium(II) coordination polymers based on bifunctional 3,2 Prime :6 Prime ,3 Prime Prime -terpyridine-4 Prime -carboxylic acid

    SciTech Connect

    Li, Na; Guo, Hui-Lin; Hu, Huai-Ming; Song, Juan; Xu, Bing; Yang, Meng-Lin; Dong, Fa-Xin; Xue, Gang-Lin

    2013-02-15

    Five new coordination polymers, [Zn{sub 2}(ctpy){sub 2}Cl{sub 2}]{sub n} (1), [Zn{sub 2}(ctpy){sub 2}(ox)(H{sub 2}O){sub 2}]{sub n} (2), [Zn{sub 2}(ctpy)(3-btc)(H{sub 2}O)]{sub n}{center_dot}0.5nH{sub 2}O (3), [Cd(ctpy){sub 2}(H{sub 2}O)]{sub n} (4), [Cd{sub 4}(ctpy){sub 2}(2-btc){sub 2}(H{sub 2}O){sub 2}]{sub n}{center_dot}2nH{sub 2}O (5), (Hctpy=3,2 Prime :6 Prime ,3 Prime Prime -terpyridine-4 Prime -carboxylic acid, H{sub 2}ox=oxalic acid, H{sub 3}(3-btc)=1,3,5-benzenetricarboxylic acid, H{sub 3}(2-btc)=1,2,4-benzenetricarboxylic acid) have been synthesized under hydrothermal conditions and characterized by elemental analysis, IR spectroscopy, and single-crystal X-ray diffraction. Compounds 1-2 are a one-dimensional chain with weak interactions to form 3D supramolecular structures. Compound 3 is a 4-nodal 3D topology framework comprised of binuclear zinc units and (ctpy){sup -} anions. Compound 4 shows two dimensional net. Compound 5 is a (4,5,6)-connected framework with {l_brace}4{sup 4}{center_dot}6{sup 2}{r_brace}{l_brace}4{sup 6}{center_dot}6{sup 4}{r_brace}{sub 2}{l_brace}4{sup 9}{center_dot}6{sup 6}{r_brace} topology. In addition, the thermal stabilities and photoluminescence properties of 1-5 were also studied in the solid state. - Graphical abstract: Five new Zn/Cd compounds with 3,2 Prime :6 Prime ,3 Prime Prime -terpyridine-4 Prime -carboxylic acid were prepared. The photoluminescence and thermal stabilities properties of 1-5 were investigated in the solid state. Highlights: Black-Right-Pointing-Pointer Five new zinc/cadmium metal-organic frameworks have been hydrothermal synthesized. Black-Right-Pointing-Pointer The structural variation is attributed to the diverse metal ions and auxiliary ligand. Black-Right-Pointing-Pointer Compounds 1-5 exhibit 1D ring chain, 2D layer and 3D open-framework, respectively. Black-Right-Pointing-Pointer These compounds exhibit strong solid state luminescence emission at room temperature.

  8. A stem–loop structure in the 59 untranslated region of bean pod mottle virus RNA2 is specifically required for RNA2 accumulation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bean pod mottle virus (BPMV) is a bipartite, positive-sense (+) RNA plant virus of the family Secoviridae. Its RNA1 encodes all proteins needed for genome replication and is capable of autonomous replication. By contrast, BPMV RNA2 must utilize RNA1-encoded proteins for replication. Here, we sought ...

  9. HLA-G 3'-untranslated region polymorphisms are associated with HTLV-1 infection, proviral load and HTLV-associated myelopathy/tropical spastic paraparesis development.

    PubMed

    Cilião Alves, Daiani Cristina; Haddad, Rodrigo; Rocha-Júnior, Maurício Cristiano; de Deus Wagatsuma, Virgínia Mara; Martelli-Palomino, Gustavo; Marques, Adriana Aparecida; Takayanagui, Osvaldo Massaiti; Covas, Dimas Tadeu; Kashima, Simone; Donadi, Eduardo Antônio

    2016-10-01

    Most human T-lymphotropic virus type 1 (HTLV-1)-infected patients remain asymptomatic throughout life. The factors associated with HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) development have not been fully elucidated; immunological and genetic factors may be involved. The association of 14 bp INS/DEL HLA-G polymorphism with HTLV-1 infection susceptibility has been reported previously. Here, other polymorphic sites at the HLA-G 3'-UTR (14-bp D/I, +3003C/T, +3010C/G, +3027A/C, +3035C/T, +3142C/G, +3187A/G and +3196C/G) were evaluated in 37 HTLV-1-infected individuals exhibiting HAM/TSP, 45 HTLV-1 asymptomatic carriers (HAC) and 153 uninfected individuals, followed up at University Hospital of the Faculty of Medicine of Ribeirão Preto, University of São Paulo, Brazil. It was observed that: (i) 14bpDI genotype is a risk factor for HTLV-1 infection, while the 14bpDD and +3142CC genotypes were associated with protection against infection; (ii) the +3142C allele and the +3003CT and +3142CC genotypes were associated with susceptibility, while 14bpII and +3003TT genotypes were associated with protection against HAM/TSP development; and (iii) the 14bpII, +3010CC, +3142GG and +3187AA genotypes were associated with lower HTLV-1 proviral load compared to respective counterpart genotypes. Findings that HLA-G has a well-recognized immunomodulatory role and that the genetic variability at HLA-G 3'-UTR may post-transcriptionally modify HLA-G production indicate a differential genetic susceptibility to: (i) the development of HTLV-1 infection, (ii) the magnitude of HTLV-1 proviral load and (iii) HAM/TSP development.

  10. A single-nucleotide polymorphism in the 3' untranslated region of the LPIN1 gene and association analysis with performance traits in chicken.

    PubMed

    Zhang, S P; Li, S Y; Chen, W; Lu, W W; Huang, Y Q

    2013-06-01

    1. A single nucleotide polymorphism (SNP), c.*77C>G, was found in the 3' UTR of the chicken LPIN1 gene by DNA sequencing. In total, 860 chickens were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in a F2 resource population obtained by crossing F0 Gushi chickens and Anka broilers, and the associations of this polymorphism with chicken growth, carcass, muscle fibre traits and serum biochemistry parameters were analysed. 2. Significant associations were found between the polymorphism and breast muscle fibre diameter (FDB). Comparison of the different genotypes of c.*77C>G in the F2 resource population showed that the GG genotype had significantly higher values than that of CG genotype in FDB. c.*77C>G was predicted to cause changes to multiple microRNA (miRNA) binding sites. But the total mRNA level of chicken LPIN1, LPIN1-;α and LPIN1-β in liver and muscle tissues did not show significant difference among GG, CG and CC genotypes, respectively. 3. The results suggested that chicken LPIN1 has a potential effect on muscle fibre development, but no effect on other studied traits.

  11. Widespread Shortening of 3’ Untranslated Regions and Increased Exon Inclusion Are Evolutionarily Conserved Features of Innate Immune Responses to Infection

    PubMed Central

    Pagé Sabourin, Ariane; Nédélec, Yohann; Dumaine, Anne; Yotova, Vania; Johnson, Zachary P.; Lanford, Robert E.; Burge, Christopher B.

    2016-01-01

    The contribution of pre-mRNA processing mechanisms to the regulation of immune responses remains poorly studied despite emerging examples of their role as regulators of immune defenses. We sought to investigate the role of mRNA processing in the cellular responses of human macrophages to live bacterial infections. Here, we used mRNA sequencing to quantify gene expression and isoform abundances in primary macrophages from 60 individuals, before and after infection with Listeria monocytogenes and Salmonella typhimurium. In response to both bacteria we identified thousands of genes that significantly change isoform usage in response to infection, characterized by an overall increase in isoform diversity after infection. In response to both bacteria, we found global shifts towards (i) the inclusion of cassette exons and (ii) shorter 3’ UTRs, with near-universal shifts towards usage of more upstream polyadenylation sites. Using complementary data collected in non-human primates, we show that these features are evolutionarily conserved among primates. Following infection, we identify candidate RNA processing factors whose expression is associated with individual-specific variation in isoform abundance. Finally, by profiling microRNA levels, we show that 3’ UTRs with reduced abundance after infection are significantly enriched for target sites for particular miRNAs. These results suggest that the pervasive usage of shorter 3’ UTRs is a mechanism for particular genes to evade repression by immune-activated miRNAs. Collectively, our results suggest that dynamic changes in RNA processing may play key roles in the regulation of innate immune responses. PMID:27690314

  12. Variation in the coding and 3’ untranslated regions of the porcine prolactin receptor short form modifies protein expression and function

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The actions of prolactin (PRL) are mediated by both long (LF) and short isoforms (SF) of the PRL receptor (PRLR). Here, we report on a genetic and functional analysis of the porcine PRLR (pPRLR) SF. Three single nucleotide polymorphisms (SNPs) within exon 11 of the pPRLR-SF give rise to four amino a...

  13. Cooperative enhancement of translation by two adjacent microRNA-122/Argonaute 2 complexes binding to the 5' untranslated region of hepatitis C virus RNA.

    PubMed

    Nieder-Röhrmann, Anika; Dünnes, Nadia; Gerresheim, Gesche K; Shalamova, Lyudmila A; Herchenröther, Andreas; Niepmann, Michael

    2017-02-01

    The liver-specific microRNA-122 (miR-122) binds to two conserved binding sites in the 5' UTR of hepatitis C virus (HCV) RNA. This binding was reported to enhance HCV RNA replication, translation and stability. We have analysed binding of miR-122/Argonaute 2 (Ago2) complexes to these sites using anti-Ago2 co-immunoprecipitation of radioactively labelled HCV RNAs along with ectopic miR-122 in HeLa cells. Our results show that the miR-122 target sites can be addressed separately. When both target sites were addressed simultaneously, we observed a synergistic binding of both miR/Ago2 complexes. Consistently, simultaneous binding of both miR-122/Ago2 complexes results in cooperative translation stimulation. In the binding assays as well as in the translation assays, binding site 1 has a stronger effect than binding site 2. We also analysed the overall RNA stability as well as the 5' end integrity of these HCV RNAs in the presence of miR-122. Surprisingly, using short HCV reporter RNAs, we did not find effects of miR-122 binding on overall RNA stability or 5' end integrity over up to 36 h. In contrast, using full-length HCV genomes that are incapable of replication, we found a positive influence of miR-122 on RNA stability, indicating that features of the full-length HCV genome that do not reside in the 5' and 3' UTRs may render HCV RNA genome stability miR-122 dependent.

  14. Identification and characterization of upstream open reading frames (uORF) in the 5' untranslated regions (UTR) of genes in Saccharomyces cerevisiae.

    PubMed

    Zhang, Zhihong; Dietrich, Fred S

    2005-08-01

    We have taken advantage of recently sequenced hemiascomycete fungal genomes to computationally identify additional genes potentially regulated by upstream open reading frames (uORFs). Our approach is based on the observation that the structure, including the uORFs, of the post-transcriptionally uORF regulated Saccharomyces cerevisiae genes GCN4 and CPA1 is conserved in related species. Thirty-eight candidate genes for which uORFs were found in multiple species were identified and tested. We determined by 5' RACE that 15 of these 38 genes are transcribed. Most of these 15 genes have only a single uORF in their 5' UTR, and the length of these uORFs range from 3 to 24 codons. We cloned seven full-length UTR sequences into a luciferase (LUC) reporter system. Luciferase activity and mRNA level were compared between the wild-type UTR construct and a construct where the uORF start codon was mutated. The translational efficiency index (TEI) of each construct was calculated to test the possible regulatory function on translational level. We hypothesize that uORFs in the UTR of RPC11, TPK1, FOL1, WSC3, and MKK1 may have translational regulatory roles while uORFs in the 5' UTR of ECM7 and IMD4 have little effect on translation under the conditions tested.

  15. rs10719 Polymorphism Located within DROSHA 3′-Untranslated Region is Responsible for Development of Primary Hypertension by Disrupting Binding with microRNA-27b

    PubMed Central

    Zhang, Yabing; Cao, Ai-lin; Dong, Chun

    2017-01-01

    Background MiR-27b is reportedly involved with many diseases (e.g., gastric cancer) by acting on different signaling pathways. In this study, we aimed at understanding the relationship between miR-27b and hypertension and its underlying molecular mechanism. Material/Methods Peripheral blood was collected from patients with hypertension, and statistical analysis was performed to study the association between rs10719 and risk of hypertension. Tissue samples were collected from patients with lung cancer, and the expression of miR-27b and DROSHA was determined using Western blot analysis and real-time PCR. Results We first searched the miRNA database online, and identified DROSHA as a virtual target of miR-27b with the “seed sequence” located within the 3′-UTR of the target gene, and then validated DROSHA to be the direct gene via luciferase reporter assay system. We also established the negative regulatory relationship between miR-27b and DROSHA via studying the relative luciferase activity. We also conducted real-time PCR to study the mRNA and protein expression level of miR-27b among different groups. Furthermore, we conducted real-time PCR and densitometry analysis to study the mRNA and protein expression level of DROSHA among different groups of cells treated with scramble control, miR-27b mimics, DROSHA siRNA, and miR-27b inhibitors to verify the negative regulatory relationship between MiR-27b and DROSHA. Conclusions The presence of rs10719 disrupted the interaction between miR-27b and DROSHA, which might be the underlying mechanism of the observation that rs10719 is significantly associated with risk of primary hypertension. PMID:28214904

  16. Genetic variant in the 3'-untranslated region of VEGFR1 gene influences chronic obstructive pulmonary disease and lung cancer development in Chinese population.

    PubMed

    Wang, Hui; Yang, Lei; Deng, Jieqiong; Wang, Bo; Yang, Xiaorong; Yang, Rongrong; Cheng, Mei; Fang, Wenxiang; Qiu, Fuman; Zhang, Xin; Ji, Weidong; Ran, Pixin; Zhou, Yifeng; Lu, Jiachun

    2014-09-01

    Lung inflammation and epithelial to mesenchymal transition (EMT) are two pathogenic features for the two contextual diseases: chronic obstructive pulmonary disease (COPD) and lung cancer. VEGFR1 (or FLT1) plays a certain role in promoting tumour growth, inflammation and EMT. To simultaneously test the association between the single nucleotide polymorphisms (SNPs) in VEGFR1 and risk of COPD and lung cancer would reveal genetic mechanisms shared by these two diseases and joint aetiology. We conducted a two-population hospital-based case-control study. Three potential functional SNPs (rs664393, rs7326277 and rs9554314) were genotyped in southern Chinese and validated in eastern Chinese to explore their associations with COPD risk in 1511 COPD patients and 1677 normal lung function controls, and with lung cancer risk in 1559 lung cancer cases and 1679 cancer-free controls. We also detected the function of the promising SNP. Individuals carrying the rs7326277C (CT+CC) variant genotypes of VEGFR1 had a significant decrease in risk of both COPD (OR = 0.78; 95% CI = 0.68-0.90) and lung cancer (OR = 0.79; 95% CI = 0.64-0.98), compared with those carrying the rs7326277TT genotype. Functional assays further showed that the rs7326277C genotypes had lower transcriptional activity and caused decreased VEGFR expression, compared with the rs7326277TT genotype. However, no significant association was observed for the other two SNPs (rs664393 and rs9554314) and either COPD or lung cancer risk. Our data suggested that the rs7326277C variant of VEGFR1 could reduce both COPD and lung cancer risk by lowering VEGFR1 mRNA expression; the SNP might be a common susceptible locus for both COPD and lung cancer.

  17. Genome-wide association identifies a deletion in the 3’ untranslated region of Striatin in a canine model of arrhythmogenic right ventricular cardiomyopathy

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a familial cardiac disease characterized by rapid ventricular tachycardia and sudden cardiac death. It is most frequently inherited as an autosomal dominant trait with incomplete and age-related penetrance and variable clinical expression. Th...

  18. The 3' untranslated region of the two cytosolic glutamine synthetase (GS(1)) genes in alfalfa (Medicago sativa) regulates transcript stability in response to glutamine.

    PubMed

    Simon, Bindu; Sengupta-Gopalan, Champa

    2010-10-01

    Glutamine synthetase (GS) catalyzes the ATP-dependent condensation of ammonia with glutamate to produce glutamine. The GS enzyme is located either in the chloroplast (GS(2)) or in the cytoplasm (GS(1)). GS(1) is encoded by a small gene family and the members exhibit differential expression pattern mostly attributed to transcriptional regulation. Based on our recent finding that a soybean GS(1) gene, Gmglnβ ( 1 ) is subject to its 3'UTR-mediated post-transcriptional regulation as a transgene in alfalfa (Medicago sativa) we have raised the question of whether the 3'UTR-mediated transcript destabilization is a more universal phenomenon. Gene constructs consisting of the CaMV35S promoter driving the reporter gene, GUS, followed by the 3'UTRs of the two alfalfa GS(1) genes, MsGSa and MsGSb, were introduced into alfalfa and tobacco. The analysis of these transformants suggests that while both the 3'UTRs promote transcript turnover, the MsGSb 3'UTR is more effective than the MsGSa 3'UTR. However, both the 3'UTRs along with Gmglnβ ( 1 ) 3'UTR respond to nitrate as a trigger in transcript turnover. More detailed analysis points to glutamine rather than nitrate as the mediator of transcript turnover. Our data suggests that the 3'UTR-mediated regulation of GS(1) genes at the level of transcript turnover is probably universal and is used for fine-tuning the expression in keeping with the availability of the substrates.

  19. HLA-E coding and 3' untranslated region variability determined by next-generation sequencing in two West-African population samples.

    PubMed

    Castelli, Erick C; Mendes-Junior, Celso T; Sabbagh, Audrey; Porto, Iane O P; Garcia, André; Ramalho, Jaqueline; Lima, Thálitta H A; Massaro, Juliana D; Dias, Fabrício C; Collares, Cristhianna V A; Jamonneau, Vincent; Bucheton, Bruno; Camara, Mamadou; Donadi, Eduardo A

    2015-12-01

    HLA-E is a non-classical Human Leucocyte Antigen class I gene with immunomodulatory properties. Whereas HLA-E expression usually occurs at low levels, it is widely distributed amongst human tissues, has the ability to bind self and non-self antigens and to interact with NK cells and T lymphocytes, being important for immunosurveillance and also for fighting against infections. HLA-E is usually the most conserved locus among all class I genes. However, most of the previous studies evaluating HLA-E variability sequenced only a few exons or genotyped known polymorphisms. Here we report a strategy to evaluate HLA-E variability by next-generation sequencing (NGS) that might be used to other HLA loci and present the HLA-E haplotype diversity considering the segment encoding the entire HLA-E mRNA (including 5'UTR, introns and the 3'UTR) in two African population samples, Susu from Guinea-Conakry and Lobi from Burkina Faso. Our results indicate that (a) the HLA-E gene is indeed conserved, encoding mainly two different protein molecules; (b) Africans do present several unknown HLA-E alleles presenting synonymous mutations; (c) the HLA-E 3'UTR is quite polymorphic and (d) haplotypes in the HLA-E 3'UTR are in close association with HLA-E coding alleles. NGS has proved to be an important tool on data generation for future studies evaluating variability in non-classical MHC genes.

  20. RNA-Mediated Virus Resistance: Role of Repeated Transgenes and Delineation of Targeted Regions.

    PubMed Central

    Sijen, T.; Wellink, J.; Hiriart, J. B.; Van Kammen, A.

    1996-01-01

    Resistance to cowpea mosaic virus (CPMV) in transgenic Nicotiana benthamiana plants is RNA mediated. In resistant CPMV movement protein (MP) gene-transformed lines, transgene steady state mRNA levels were low, whereas nuclear transcription rates were high, implying that a post-transcriptional gene-silencing mechanism is at the base of the resistance. The silencing mechanism can also affect potato virus X (PVX) RNAs when they contain CPMV MP gene sequences. In particular, sequences situated in the 3[prime] part of the transcribed region of the MP transgene direct elimination of recombinant PVX genomes. Remarkably, successive portions of this 3[prime] part, which can be as small as 60 nucleotides, all tag PVX genomes for degradation. These observations suggest that the entire 3[prime] part of the MP transgene mRNA is the initial target of the silencing mechanism. The arrangement of transgenes in the plant genome plays an important role in establishing resistance because the frequency of resistant lines increased from 20 to 60% when transformed with a transgene containing a direct repeat of MP sequences rather than a single MP transgene. Interestingly, we detected strong methylation in all of the plants containing directly repeated MP sequences. In sensitive lines, only the promoter region was found to be heavily methylated, whereas in resistant lines, only the transcribed region was strongly methylated. PMID:12239378

  1. Synthesis of 25-hydroxyvitamin D sub 3 3. beta. -3 prime -(N-(4-azido-2-nitrophenyl)amino)propyl ether, a second-generation photoaffinity analogue of 25-hydroxyvitamin D sub 3 : Photoaffinity labeling of rat serum vitamin D binding protein

    SciTech Connect

    Ray, R.; Holick, M.F. ); Bouillon, R.; Van Baelen, H. )

    1991-05-14

    Vulnerability of 25-hydroxy-(26,27-{sup 3}H)vitamin D{sub 3} 3{beta}-N-(4-azido-2-nitrophenyl)glycinate, a photoaffinity analogue of 25-hydroxyvitamin D{sub 3} (25-OH-D{sub 3}) toward standard conditions of carboxymethylationin promoted the authors to synthesize 25-hydroxyvitamin D{sub 3} 3{beta}-3{prime}-(N-(4-azido-2-nitrophenyl)amino)propyl ether (25-ANE), a hydrolytically stable photoaffinity analogue of 25-OH-D{sub 3}, and 25-hydroxyvitamin D{sub 3} 3{beta}-3{prime}-(N-(4-azido-2-nitro-(3,5-{sup 3}H)phenyl)amino)propyl ether ({sup 3}H-25-ANE), the radiolabeled counterpart of 25-ANE competes for the 25-OH-D{sub 3} binding site in rat serum vitamin D binding protein (rDBP). On the other hand, UV exposure of a sample of purified rat DBP (rDBP), preincubated in the dark with {sup 3}H-25-ANE, covalently labeled the protein. However, very little covalent labeling was observed in the absence of UV light or in the presence of a large excess of 25-OH-D{sub 3}. These results provide strong evidence for the covalent labeling of the 25-OH-D{sub 3} binding site in rDPB by {sup 3}H-25-ANE.

  2. Differential accumulation of ribonucleotide reductase subunits in clam oocytes: the large subunit is stored as a polypeptide, the small subunit as untranslated mRNA

    PubMed Central

    1986-01-01

    Within minutes of fertilization of clam oocytes, translation of a set of maternal mRNAs is activated. One of the most abundant of these stored mRNAs encodes the small subunit of ribonucleotide reductase (Standart, N. M., S. J. Bray, E. L. George, T. Hunt, and J. V. Ruderman, 1985, J. Cell Biol., 100:1968-1976). Unfertilized oocytes do not contain any ribonucleotide reductase activity; such activity begins to appear shortly after fertilization. In virtually all organisms, this enzyme is composed of two dissimilar subunits with molecular masses of approximately 44 and 88 kD, both of which are required for activity. This paper reports the identification of the large subunit of clam ribonucleotide reductase isolated by dATP-Sepharose chromatography as a relatively abundant 86-kD polypeptide which is already present in oocytes, and whose level remains constant during early development. The enzyme activity of this large subunit was established in reconstitution assays using the small subunit isolated from embryos by virtue of its binding to the anti-tubulin antibody YL 1/2. Thus the two components of clam ribonucleotide reductase are differentially stored in the oocyte: the small subunit in the form of untranslated mRNA and the large subunit as protein. When fertilization triggers the activation of translation of the maternal mRNA, the newly synthesized small subunit combines with the preformed large subunit to generate active ribonucleotide reductase. PMID:3536960

  3. Linkage disequilibrium in the neurofibromatosis I (NF1) region: Implications for gene mapping

    SciTech Connect

    Jorde, L.B.; Watkins, W.S.; Viskochil, D.; Ward, K. ); O'Connell, P. )

    1993-11-01

    To test the usefulness of linkage disequilibrium for gene mapping, the authors compared physical distances and linkage disequilibrium among eight RFLPs in the neutrofibromatosis 1 (NF1) region. Seven of the polymorphisms span most of the NF1 gene, while the remaining polymorphism lies approximately 70 kb 3[prime] to a stop codon in exon 49. By using Centre d'Etude du Polymorphisme Humain (CEPH) kindreds, 91-110 unrelated parents were genotyped. A high degree of disequilibrium is maintained among the seven intragenic polymorphisms (r > .82, P < 10[sup [minus]7]), even though they are separated by as much as 340 kb. The 3[prime] polymorphism is only 68 kb distal to the next polymorphism, but disequilibrium between the 3[prime] polymorphism and all others is comparatively low ([vert bar]r[vert bar] < .33, P values .27-.001). This result was replicated in three sets of unrelated kindreds: the Utah CEPH families, the non-Utah CEPH families, and an independent set of NF1 families. Trigenic, quadrigenic, three-locus, and four-locus disequilibrium measures were also estimated. There was little evidence of higher-order linkage disequilibrium. As expected for a disease with multiple mutations, no disequilibrium was observed between the disease gene and any of the RFLPs. The observed pattern of high disequilibrium within the gene and a loss of disequilibrium 3[prime] to the stop codon could have implications for gene mapping studies. These are discussed, and guidelines for linkage disequilibrium studies are suggested. 80 refs., 2 figs., 5 tabs.

  4. Effects of 3,3{prime},4,4{prime},5-pentachlorobiphenyl (PCB 126), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), or an extract derived from field-collected cormorant eggs injected into double-crested cormorant (Phalacrocorax auritus) eggs

    SciTech Connect

    Powell, D.C.; Aulerich, R.J.; Powell, J.F.; Restum, J.C.; Giesy, J.P.; Bursian, S.J.; Meadows, J.C.; Tillitt, D.E.; Stromborg, K.L.

    1997-07-01

    Double-crested cormorant (Phalacrocorax auritus) eggs were injected with either 3,3{prime},4,4{prime},5-pentachlorobiphenyl (PCB 126), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), or an extract derived from field-collected double-crested cormorant eggs. These compounds were injected into the yolks of cormorant eggs from an isolated colony on Lake Winnipegosis, Manitoba, Canada. Upon hatching, chicks were necropsied. The brain, bursa, heart, liver, and spleen were removed and weighed. An approximate median lethal dose (LD50) of 158 {micro}g/kg egg was determined for PCB 126, which is 69 times greater than the LD50 determined for the chicken (Gallus domesticus) in a previous study. A significantly greater mortality occurred at the highest dose of TCDD when compared to the vehicle control. However, the mortality data did not provide sufficient information for the determination of an LD50. The cormorant egg extract did not adversely affect hatchability. No significant increases were observed in the incidence of developmental abnormalities, including pronounced edema, in any of the treatment groups, nor were there any relevant effects on body and organ weights. Based on the results from this study, the cormorant appears to be considerably less sensitive to polyhalogenated diaromatic hydrocarbons than the chicken, which has been the typical species used for egg injection studies.

  5. Interleukin-3 priming in acute myeloid leukaemia patients.

    PubMed

    Tafuri, A; de Felice, L; Goodacre, A; Fenu, S; Petrucci, M T; Valentini, T; Alimena, G; Petti, M C; Meloni, G; Mandelli, F

    1995-09-01

    Several studies have demonstrated that G-CSF, GM-CSF and, in particular, IL-3 can effectively recruit acute myeloid leukaemia (AML) blasts into the cell cycle, resulting in a significant increase in cytosine-arabinoside (Ara-C) mediated cytotoxicity in vitro. Since IL-3 has shown biological and clinical activity, we investigated the cell kinetic effects of rIL-3 and high-dose Ara-C/idarubicin in three patients with refractory AML selected for the presence of chromosome 7 monosomy; this enabled differentiation between the effects of IL-3 on leukaemic and on normal cells. The in vivo administration of rhIL-3 (250 micrograms/m2d s.c. for 6-10d) recruited AML blasts into the cell cycle in two of the three patients, and this effect resulted in an increase in in vitro growth of clonogenic cells (CFU-L) and of their S-phase fraction. The percentage of leukaemic cells with monosomy 7 increased only in the two cases who showed a proliferative response. Normal cells were not recruited, even when rhIL-3 was administered for up to 10 d. In vitro studies showed an increased Ara-C cytotoxicity on clonogenic AML cells, in particular with IL-3 plus GM-CSF, thus confirming the priming effects of IL-3 in the two responding cases. The results of this study suggest that rhIL-3 can selectively recruit leukaemic cells into the cell cycle. Although leukaemic blasts can be sensitized to Ara-C, other mechanisms of primary blast resistance may limit the clinical benefit of kinetic-based approaches.

  6. Higher order structures of the 5'-proximal region decrease the efficiency of translation of the porcine pro-opiomelanocortin mRNA.

    PubMed

    Chevrier, D; Vézina, C; Bastille, J; Linard, C; Sonenberg, N; Boileau, G

    1988-01-15

    The SP6 polymerase/promoter system was used to synthesize porcine pro-opiomelanocortin mRNAs with nucleotide sequence deletions in the 5'- as well as 3'-untranslated and coding regions. The translational efficiency of the mutant mRNAs was evaluated by cell-free translation or by monitoring the rate and extent of ribosome binding in the presence of sparsomycin. The results of these experiments indicate that specific nucleotide sequences in the 5'-untranslated and coding regions of the pro-opiomelanocortin mRNA decrease its rate of translation. Structure mapping of the mRNA with double-strand and single-strand specific nucleases suggests that these sequences can form stable secondary structures.

  7. High-density single-nucleotide polymorphism (SNP) map in the 96-kb region containing the entire human DiGeorge syndrome critical region 2 (DGCR2) gene at 22q11.2.

    PubMed

    Iida, A; Ohnishi, Y; Ozaki, K; Ariji, Y; Nakamura, Y; Tanaka, T

    2001-01-01

    We constructed a high-density single-nucleotide polymorphism (SNP) map in the 96-kb region containing the DiGeorge syndrome critical region 2 (DGCR2) gene at chromosome 22q11.2, a human counterpart of mouse seizure-related gene SEZ-12. A total of 102 SNPs were isolated from the region by systematic screening among 48 Japanese individuals: 9 SNPs in the 5' flanking region, 3 in the 5' untranslated region, 2 in the coding regions, 77 in introns, 7 in the 3' untranslated region, and 4 in the 3' flanking region. By a comparison of our data with SNPs deposited in the dbSNP database in the National Center for Biotechnology Information, 80 SNPs (78.4%) were considered to be novel. The ratio of transition to transversion was 3.08:1. In addition, eight other types of genetic variations (one GA dinucleotide polymorphism and seven insertion/deletion polymorphisms) were discovered. The high-resolution map that we constructed will be a useful resource for analyzing gene scans of complex diseases mapped to this local segment on chromosome 22.

  8. Analysis of the genetic diversity of the Plasmodium falciparum multidrug resistance gene 5' upstream region.

    PubMed

    Myrick, Alissa; Sarr, Ousmane; Dieng, Therese; Ndir, Omar; Mboup, Souleymane; Wirth, Dyann F

    2005-02-01

    Recent findings indicating a low level of polymorphism in the Plasmodium falciparum genome have led to the hypothesis that existent polymorphisms are likely to have functional significance. We tested this hypothesis by developing a map of the polymorphism in the P. falciparum multidrug resistance 1 (pfmdr1) gene 5' upstream region and assaying its correlation with drug resistance in a sample of field isolates from Dakar, Senegal. A comparison of six geographically diverse laboratory strains showed that the 1.94-kb 5'-untranslated region is highly monomorphic, with a total of four unique single nucleotide polymorphisms (SNPs) being identified. All of the mutations were localized to a 462-basepair region proximal to the transcription start point. Analysis of this region in field isolates shows the prevalence of one SNP throughout the entire population of parasites, irrespective of drug resistance status. The SNP frequency of the pfmdr1 upstream region is lower than that found in the noncoding region of other genes.

  9. Molecular cloning of the gene encoding the bovine brain ribonuclease and its expression in different regions of the brain.

    PubMed Central

    Sasso, M P; Carsana, A; Confalone, E; Cosi, C; Sorrentino, S; Viola, M; Palmieri, M; Russo, E; Furia, A

    1991-01-01

    In this paper we report the molecular cloning of the gene encoding the bovine brain ribonuclease. The nucleotide sequence determined in this work shows a high degree of identity to the homologous gene encoding the bovine pancreatic ribonuclease. Processing of the primary transcripts of these genes also follows a similar pathway, splicing of the unique intron in the 5' untranslated region occurs at corresponding positions. Expression of the bovine brain ribonuclease gene can be detected both at the transcriptional and translational levels in all the regions of the brain examined. Images PMID:1754384

  10. The turnip yellow mosaic virus tRNA-like structure cannot be replaced by generic tRNA-like elements or by heterologous 3' untranslated regions known to enhance mRNA expression and stability.

    PubMed Central

    Skuzeski, J M; Bozarth, C S; Dreher, T W

    1996-01-01

    The tRNA-like structure (TLS) at the 3' end of the turnip yellow mosaic virus genome was replaced with heterologous tRNA-like elements, and with a poly(A) tail, in order to assess its role. Replacement with the valylatable TLSs from two closely related tymoviruses resulted in infectious viruses. In contrast, no systemic symptoms on plants, and only low viral accumulations in protoplasts, were observed for three chimeric genomes with 3' sequences known to enhance mRNA stability and translatability. One of these chimeras had a poly(A) tail, and the others had the TLS with associated upstream pseudoknot tracts from the 3' ends of brome mosaic and tobacco mosaic viruses. The latter two chimeric RNAs were shown to be appropriately folded by demonstrating their aminoacylation in vitro with tyrosine and histidine, respectively. The results show that enhancement of genome stability or gene expression is not the major role of the turnip yellow mosaic virus TLS. The major role is likely to be replicational, dependent on features present in tymoviral TLSs but not in generic tRNA-like structures. PMID:8642631

  11. Naturally occurring human genetic variation in the 3’-untranslated region of the secretory protein chromogranin A (CHGA) is associated with autonomic blood pressure regulation and hypertension in sex-dependent fashion

    PubMed Central

    Chen, Yuqing; Rao, Fangwen; Rodriguez-Flores, Juan L.; Mahata, Manjula; Fung, Maple M.; Stridsberg, Mats; Vaingankar, Sucheta M.; Wen, Gen; Salem, Rany M.; Das, Madhusudan; Cockburn, Myles G.; Schork, Nicholas J.; Ziegler, Michael G.; Hamilton, Bruce A.; Mahata, Sushil K.; Taupenot, Laurent; O’Connor, Daniel T.

    2008-01-01

    Objective Determination whether common variation at the CHGA locus increases susceptibility to hypertension. Background Chromogranin A (CHGA) regulates catecholamine storage and release. Previously we systematically identified genetic variants across CHGA. Methods Dense genotyping across the CHGA locus in >1000 individuals with the most extreme BPs in the population, as well twin pairs with autonomic phenotypes. Characterizing function of a trait-associated 3'-UTR variant with transfected CHGA 3'-UTR/luciferase reporter plasmids. Results CHGA was overexpressed in patients with hypertension, especially hypertensive men, and CHGA predicted catecholamines. In individuals with extreme BPs, CHGA genetic variants predicted BP, especially in men, with a peak association occurred in the 3'-UTR at C+87T, accounting for up to ~12/~9 mmHg. The C+87T genotype predicted CHGA secretion in vivo, with the +87T allele (associated with lower BP) also diminishing plasma CHGA by ~10%. The C+87T 3'-UTR variant also predicted the BP response to environmental (cold) stress; the same allele (+87T) that diminished basal BP in the population also decreased the SBP response to stress by ~12 mmHg, and the response was smaller in women (by ~6 mmHg). In a chromaffin cell-transfected CHGA 3'-UTR/luciferase reporter plasmid, the +87T allele associated with lower BP also decreased reporter expression by ~30%. In cultured chromaffin cells, reducing endogenous Chga expression by si-RNA caused ~2/3 depletion of catecholamine storage vesicles. Conclusions Common variant C+87T in the CHGA 3'-UTR is a functional polymorphism causally associated with hypertension especially in men of the population, and propose steps ("intermediate phenotypes") whereby in sex-dependent fashion this genetic variant influences the ultimate disease trait. These observations suggest new molecular strategies to probe the pathophysiology, risk, and rational treatment of hypertension. PMID:19017515

  12. Introduction of mutations into the non-structural genes or 3' untranslated region of an attenuated dengue virus type 4 vaccine candidate further decreases replication in rhesus monkeys while retaining protective immunity.

    PubMed

    Hanley, Kathryn A; Manlucu, Luella R; Manipon, Gracielle G; Hanson, Christopher T; Whitehead, Stephen S; Murphy, Brian R; Blaney, Joseph E

    2004-09-03

    A dengue virus vaccine candidate, rDEN4Delta30, has been previously reported to be safe and immunogenic in humans, but a subset of vaccinees developed asymptomatic rash, elevation of liver enzymes and/or mild neutropenia. In the current study, mutations that had previously been shown to reduce replication of DEN4 virus in suckling mice and/or in SCID mice engrafted with human liver cells (SCID-HuH-7 mice) were introduced into rDEN4Delta30 in an attempt to further attenuate this virus. Three of the five resulting modified rDEN4Delta30 viruses showed decreased replication in SCID-HuH-7 mice relative to rDEN4Delta30. Moreover, in rhesus monkeys, two of the modified rDEN4Delta30 viruses showed a decrease in replication relative to rDEN4Delta30 while generating levels of neutralizing antibody similar to rDEN4Delta30 virus. All of the modified rDEN4Delta30 viruses completely protected immunized rhesus monkeys from challenge with wild-type DEN4 virus. Based on their attenuation for both human liver cells and rhesus monkeys, two of the modified rDEN4Delta30 vaccine candidates are currently being prepared for use in clinical trials. The application of these attenuating mutations to flavivirus vaccine development is discussed.

  13. Region between the canine distemper virus M and F genes modulates virulence by controlling fusion protein expression.

    PubMed

    Anderson, Danielle E; von Messling, Veronika

    2008-11-01

    Morbilliviruses, including measles and canine distemper virus (CDV), are nonsegmented, negative-stranded RNA viruses that cause severe diseases in humans and animals. The transcriptional units in their genomes are separated by untranslated regions (UTRs), which contain essential transcription and translation signals. Due to its increased length, the region between the matrix (M) protein and fusion (F) protein open reading frames is of particular interest. In measles virus, the entire F 5' region is untranslated, while several start codons are found in most other morbilliviruses, resulting in a long F protein signal peptide (Fsp). To characterize the role of this region in morbillivirus pathogenesis, we constructed recombinant CDVs, in which either the M-F UTR was replaced with that between the nucleocapsid (N) and phosphoprotein (P) genes, or 106 Fsp residues were deleted. The Fsp deletion alone had no effect in vitro and in vivo. In contrast, substitution of the UTR was associated with a slight increase in F gene and protein expression. Animals infected with this virus either recovered completely or experienced prolonged disease and death due to neuroinvasion. The combination of both changes resulted in a virus with strongly increased F gene and protein expression and complete attenuation. Taken together, our results provide evidence that the region between the morbillivirus M and F genes modulates virulence through transcriptional control of the F gene expression.

  14. Nucleotide diversity analysis highlights functionally important genomic regions.

    PubMed

    Tatarinova, Tatiana V; Chekalin, Evgeny; Nikolsky, Yuri; Bruskin, Sergey; Chebotarov, Dmitry; McNally, Kenneth L; Alexandrov, Nickolai

    2016-10-24

    We analyzed functionality and relative distribution of genetic variants across the complete Oryza sativa genome, using the 40 million single nucleotide polymorphisms (SNPs) dataset from the 3,000 Rice Genomes Project (http://snp-seek.irri.org), the largest and highest density SNP collection for any higher plant. We have shown that the DNA-binding transcription factors (TFs) are the most conserved group of genes, whereas kinases and membrane-localized transporters are the most variable ones. TFs may be conserved because they belong to some of the most connected regulatory hubs that modulate transcription of vast downstream gene networks, whereas signaling kinases and transporters need to adapt rapidly to changing environmental conditions. In general, the observed profound patterns of nucleotide variability reveal functionally important genomic regions. As expected, nucleotide diversity is much higher in intergenic regions than within gene bodies (regions spanning gene models), and protein-coding sequences are more conserved than untranslated gene regions. We have observed a sharp decline in nucleotide diversity that begins at about 250 nucleotides upstream of the transcription start and reaches minimal diversity exactly at the transcription start. We found the transcription termination sites to have remarkably symmetrical patterns of SNP density, implying presence of functional sites near transcription termination. Also, nucleotide diversity was significantly lower near 3' UTRs, the area rich with regulatory regions.

  15. Nucleotide diversity analysis highlights functionally important genomic regions

    PubMed Central

    Tatarinova, Tatiana V.; Chekalin, Evgeny; Nikolsky, Yuri; Bruskin, Sergey; Chebotarov, Dmitry; McNally, Kenneth L.; Alexandrov, Nickolai

    2016-01-01

    We analyzed functionality and relative distribution of genetic variants across the complete Oryza sativa genome, using the 40 million single nucleotide polymorphisms (SNPs) dataset from the 3,000 Rice Genomes Project (http://snp-seek.irri.org), the largest and highest density SNP collection for any higher plant. We have shown that the DNA-binding transcription factors (TFs) are the most conserved group of genes, whereas kinases and membrane-localized transporters are the most variable ones. TFs may be conserved because they belong to some of the most connected regulatory hubs that modulate transcription of vast downstream gene networks, whereas signaling kinases and transporters need to adapt rapidly to changing environmental conditions. In general, the observed profound patterns of nucleotide variability reveal functionally important genomic regions. As expected, nucleotide diversity is much higher in intergenic regions than within gene bodies (regions spanning gene models), and protein-coding sequences are more conserved than untranslated gene regions. We have observed a sharp decline in nucleotide diversity that begins at about 250 nucleotides upstream of the transcription start and reaches minimal diversity exactly at the transcription start. We found the transcription termination sites to have remarkably symmetrical patterns of SNP density, implying presence of functional sites near transcription termination. Also, nucleotide diversity was significantly lower near 3′ UTRs, the area rich with regulatory regions. PMID:27774999

  16. cDNA sequence, genomic organization, and evolutionary conservation of a novel gene from the WAGR region

    SciTech Connect

    Schwartz, F.; Eisenman, R.; Knoll, J.; Bruns, G.

    1995-09-20

    A new gene (239FB) with predominant and differential expression in fetal brain has recently been isolated from a chromosome 11p13-p14 boundary area near FSHB. The corresponding mRNA has an open reading frame of 294 amino acids, a 3` untranslated region of 1247 nucleotides, and a highly GC-rich 5` untranslated region. The coding and 3` UT sequence is specified by 6 exons within nearly 87 kb of isolated genomic locus. The 5` end region of the transcript maps adjacent to the only genomically defined CpG island in a chromosomal subregion that may be associated with part of the mental retardation of some WAGR (Wilms tumor, aniridia, genitourinary anomalies, and mental retardation) syndrome patients. In addition to nucleotide and amino acid similarity to an EST from a normalized infant brain cDNA library, the predicted protein has extensive similarity to Caenorhbditis elegans polypeptides of, as yet, unknown function. The 239FB locus is, therefore, likely part of a family of genes with two members expressed in human brain. The extensive conservation of the predicted protein suggests a fundamental function of the gene product and will enable evaluation of the role of the 239FB gene in neurogenesis in model organisms. 48 refs., 4 figs., 1 tab.

  17. High sequence turnover in the regulatory regions of the developmental gene hunchback in insects.

    PubMed

    Hancock, J M; Shaw, P J; Bonneton, F; Dover, G A

    1999-02-01

    Extensive sequence analysis of the developmental gene hunchback and its 5' and 3' regulatory regions in Drosophila melanogaster, Drosophila virilis, Musca domestica, and Tribolium castaneum, using a variety of computer algorithms, reveals regions of high sequence simplicity probably generated by slippage-like mechanisms of turnover. No regions are entirely refractory to the action of slippage, although the density and composition of simple sequence motifs varies from region to region. Interestingly, the 5' and 3' flanking regions share short repetitive motifs despite their separation by the gene itself, and the motifs are different in composition from those in the exons and introns. Furthermore, there are high levels of conservation of motifs in equivalent orthologous regions. Detailed sequence analysis of the P2 promoter and DNA footprinting assays reveal that the number, orientation, sequence, spacing, and protein-binding affinities of the BICOID-binding sites varies between species and that the 'P2' promoter, the nanos response element in the 3' untranslated region, and several conserved boxes of sequence in the gene (e.g., the two zinc-finger regions) are surrounded by cryptically-simple-sequence DNA. We argue that high sequence turnover and genetic redundancy permit both the general maintenance of promoter functions through the establishment of coevolutionary (compensatory) changes in cis- and trans-acting genetic elements and, at the same time, the possibility of subtle changes in the regulation of hunchback in the different species.

  18. Multiple pathways for steel regulation suggested by genomic and sequence analysis of the murine Steel gene

    SciTech Connect

    Bedell, M.A.; Copeland, N.G.; Jenkins, N.A.

    1996-03-01

    The Steel (Sl) locus encodes mast cell growth factor (Mgf) that is required for the development of germ cells, hematopoietic cells and melanocytes. Although the expression patterns of the Mgf gene are well characterized, little is known of the factors which regulate its expression. Here, we describe the cloning and sequence of the full-length transcription unit and the 5{prime} flanking region of the murine Mgf gene. The full-length Mgf mRNA consists of a short 5{prime} untranslated region (UTR), a 0.8-kb ORF and a long 3{prime} UTR. A single transcription initiation site is used in a number of mouse tissues and is located just downstream of binding sites for several known transcription factors. In the 5{prime} UTR, two ATGs were found upstream of the initiator methionine and are conserved among different species, suggesting that Mgf may be translationally regulated. At least two Mgf mRNAs are produced by alternative use of polyadenylation sites, but numerous other potential polyadenylation sites were found in the 3{prime} UTR. In addition, the 3{prime} UTR contains numerous sequence motifs that may regulate Mgf mRNA stability. These studies suggest multiple ways in which expression of Mgf may be regulated. 39 refs., 4 figs.

  19. Genomic organization of the neurofibromatosis 1 gene (NF1)

    SciTech Connect

    Li, Y.; O`Connell, P.; Huntsman Breidenbach, H.

    1995-01-01

    Neurofibromatosis 1 maps to chromosome band 17q11.2, and the NF1 locus has been partially characterized. Even though the full-length NF1 cDNA has been sequenced, the complete genomic structure of the NF1 gene has not been elucidated. The 5{prime} end of NF1 is embedded in a CpG island containing a NotI restriction site, and the remainder of the gene lies in the adjacent 350-kb NotI fragment. In our efforts to develop a comprehensive screen for NF1 mutations, we have isolated genomic DNA clones that together harbor the entire NF1 cDNA sequence. We have identified all intron-exon boundaries of the coding region and established that it is composed of 59 exons. Furthermore, we have defined the 3{prime}-untranslated region (3{prime}-UTR) of the NF1 gene; it spans approximately 3.5 kb of genomic DNA sequence and is continuous with the stop codon. Oligonucleotide primer pairs synthesized from exon-flanking DNA sequences were used in the polymerase chain reaction with cloned, chromosome 17-specific genomic DNA as template to amplify NF1 exons 1 through 27b and the exon containing the 3{prime}-UTR separately. This information should be useful for implementing a comprehensive NF1 mutation screen using genomic DNA as template. 41 refs., 3 figs., 2 tabs.

  20. Transgenic mouse model of hemifacial microsomia: Cloning and characterization of insertional mutation region on chromosome 10

    SciTech Connect

    Naora, Hiroyuki; Otani, Hiroki; Tanaka, Osamu

    1994-10-01

    The 643 transgenic mouse line carries an autosomal dominant insertional mutation that results in hemifacial microsomia (HFM), including microtia and/or abnormal biting. In this paper, we characterize the transgene integration site in transgenic mice and preintegration site of wildtype mice. The locus, designated Hfm (hemifacial microsomia-associated locus), was mapped to chromosome 10, B1-3, by chromosome in situ hybridization. We cloned the transgene insertion site from the transgenic DNA library. By using the 5{prime} and 3{prime} flanking sequences, the preintegration region was isolated. The analysis of these regions showed that a deletion of at least 23 kb DNA occurred in association with the transgene integration. Evolutionarily conserved regions were detected within and beside the deleted region. The result of mating between hemizygotes suggests that the phenotype of the homozygote is lethality in the prenatal period. These results suggests that the Hfm locus is necessary for prenatal development and that this strain is a useful animal model for investigating the genetic predisposition to HFM in humans.

  1. The transcriptionally active regions in the genome of Bacillus subtilis

    PubMed Central

    Rasmussen, Simon; Nielsen, Henrik Bjørn; Jarmer, Hanne

    2009-01-01

    The majority of all genes have so far been identified and annotated systematically through in silico gene finding. Here we report the finding of 3662 strand-specific transcriptionally active regions (TARs) in the genome of Bacillus subtilis by the use of tiling arrays. We have measured the genome-wide expression during mid-exponential growth on rich (LB) and minimal (M9) medium. The identified TARs account for 77.3% of the genes as they are currently annotated and additionally we find 84 putative non-coding RNAs (ncRNAs) and 127 antisense transcripts. One ncRNA, ncr22, is predicted to act as a translational control on cstA and an antisense transcript was observed opposite the housekeeping sigma factor sigA. Through this work we have discovered a long conserved 3′ untranslated region (UTR) in a group of membrane-associated genes that is predicted to fold into a large and highly stable secondary structure. One of the genes having this tail is efeN, which encodes a target of the twin-arginine translocase (Tat) protein translocation system. PMID:19682248

  2. NcoI RFLP at the creatine kinase-muscle type gene locus (CKMM, chromosome 19)

    SciTech Connect

    Coerwinkel-Driessen, M.; Schepens, J.; van Zandvoort, P.; van Oost, B.; Mariman, E.; Wieringa, B. )

    1988-09-12

    A 3.2 kbp human genomic DNA fragment (BamHI-Sau3A) of the 3{prime} untranslated and 3{prime} flanking region of the CKMM gene was isolated and subcloned into the BamHI site of vector pSP64. The CKMM 3{prime}-probe identifies a 2-allele polymorphism with bands at 2.3 and 1.0 kbp (allele A) and 3.3 kbp (allele B). In addition a weak constant 4.2 kbp band is observed. This probe also detects a 2-allele TaqI RFLP reported previously, as either a 4.3 kbp (A) or a 4.2 kbp (B) band. The CKMM locus previously has been assigned to 19q13.2-q13.3. By Southern blot analysis of human-rodent somatic cell hybrids containing unique subregional fragments of chromosome 19 of man the authors have assigned the gene to 19q13.2. Co-dominant segregation was observed in 8 families with 3 generations.

  3. Identification and genetic mapping of a homeobox gene to the 4p16.1 region of human chromosome 4.

    PubMed

    Stadler, H S; Padanilam, B J; Buetow, K; Murray, J C; Solursh, M

    1992-12-01

    A human craniofacial cDNA library was screened with a degenerate oligonucleotide probe based on the conserved third helix of homeobox genes. From this screening, we identified a homeobox gene, H6, which shared only 57-65% amino acid identity to previously reported homeodomains. H6 was physically mapped to the 4p16.1 region by using somatic cell hybrids containing specific deletions of human chromosome 4. Linkage data from a single-stranded conformational polymorphism derived from the 3' untranslated region of the H6 cDNA placed this homeobox gene more than 20 centimorgans proximal of the previously mapped HOX7 gene on chromosome 4. Identity comparisons of the H6 homeodomain with previously reported homeodomains reveal the highest identities to be with the Nk class of homeobox genes in Drosophila melanogaster.

  4. Identification and genetic mapping of a homeobox gene to the 4p16.1 region of human chromosome 4.

    PubMed Central

    Stadler, H S; Padanilam, B J; Buetow, K; Murray, J C; Solursh, M

    1992-01-01

    A human craniofacial cDNA library was screened with a degenerate oligonucleotide probe based on the conserved third helix of homeobox genes. From this screening, we identified a homeobox gene, H6, which shared only 57-65% amino acid identity to previously reported homeodomains. H6 was physically mapped to the 4p16.1 region by using somatic cell hybrids containing specific deletions of human chromosome 4. Linkage data from a single-stranded conformational polymorphism derived from the 3' untranslated region of the H6 cDNA placed this homeobox gene more than 20 centimorgans proximal of the previously mapped HOX7 gene on chromosome 4. Identity comparisons of the H6 homeodomain with previously reported homeodomains reveal the highest identities to be with the Nk class of homeobox genes in Drosophila melanogaster. Images PMID:1360670

  5. Sequence breakpoints in the aflatoxin biosynthesis gene cluster and flanking regions in nonaflatoxigenic Aspergillus flavus isolates.

    PubMed

    Chang, Perng-Kuang; Horn, Bruce W; Dorner, Joe W

    2005-11-01

    Aspergillus flavus populations are genetically diverse. Isolates that produce either, neither, or both aflatoxins and cyclopiazonic acid (CPA) are present in the field. We investigated defects in the aflatoxin gene cluster in 38 nonaflatoxigenic A. flavus isolates collected from southern United States. PCR assays using aflatoxin-gene-specific primers grouped these isolates into eight (A-H) deletion patterns. Patterns C, E, G, and H, which contain 40 kb deletions, were examined for their sequence breakpoints. Pattern C has one breakpoint in the cypA 3' untranslated region (UTR) and another in the verA coding region. Pattern E has a breakpoint in the amdA coding region and another in the ver1 5'UTR. Pattern G contains a deletion identical to the one found in pattern C and has another deletion that extends from the cypA coding region to one end of the chromosome as suggested by the presence of telomeric sequence repeats, CCCTAATGTTGA. Pattern H has a deletion of the entire aflatoxin gene cluster from the hexA coding region in the sugar utilization gene cluster to the telomeric region. Thus, deletions in the aflatoxin gene cluster among A. flavus isolates are not rare, and the patterns appear to be diverse. Genetic drift may be a driving force that is responsible for the loss of the entire aflatoxin gene cluster in nonaflatoxigenic A. flavus isolates when aflatoxins have lost their adaptive value in nature.

  6. Different mRNAs code for dopa decarboxylase in tissues of neuronal and nonneuronal origin

    SciTech Connect

    Krieger, M.; Coge, F.; Gros, F.; Thibault, J. )

    1991-03-15

    A cDNA clone for dopa decarboxylase has been isolated from a rat pheochromocytoma cDNA library and the cDNA sequence has been determined. It corresponds to an mRNA of 2094 nucleotides. The length of the mRNA was measured by primer-extension of rat pheochromocytoma RNA and the 5{prime} end of the sequence of the mRNA was confirmed by the PCR. A probe spanning the translation initiation site of the mRNA was used to hybridize with mRNAs from various organs of the rat. S1 nuclease digestion of the mRNAs annealed with this probe revealed two classes of mRNAs. The comparison of the cDNA sequence and published sequences for rat liver, human pheochromocytoma, and Droxophila dopa decarboxylase supported the conclusion that two mRNAs are produced: one is specific for tissue of neuronal origin and the other is specific for tissues of nonneuronal (mesodermal or endodermal) origin. The neuronal mRNA contains a 5{prime} untranslated sequence that is highly conserved between human and rat pheochromocytoma including a GA stretch. The coding sequence and the 3{prime} untranslated sequence of mRNAs from rat liver and pheochromocytoma are identical. The rat mRNA differs only in the 5{prime} untranslated region. Thus a unique gene codes for dopa decarboxylase and this gene gives rise to at least two transcripts presumably in response to different signals during development.

  7. Identification, characterization and functional analysis of regulatory region of nanos gene from half-smooth tongue sole (Cynoglossus semilaevis).

    PubMed

    Huang, Jinqiang; Li, Yongjuan; Shao, Changwei; Wang, Na; Chen, Songlin

    2017-03-25

    The nanos gene encodes an RNA-binding zinc finger protein, which is required in the development and maintenance of germ cells. However, there is very limited information about nanos in flatfish, which impedes its application in fish breeding. In this study, we report the molecular cloning, characterization and functional analysis of the 3'-untranslated region of the nanos gene (Csnanos) from half-smooth tongue sole (Cynoglossus semilaevis), which is an economically important flatfish in China. The 1233-bp cDNA sequence, 1709-bp genomic sequence and flanking sequences (2.8-kb 5'- and 1.6-kb 3'-flanking regions) of Csnanos were cloned and characterized. Sequence analysis revealed that CsNanos shares low homology with Nanos in other species, but the zinc finger domain of CsNanos is highly similar. Phylogenetic analysis indicated that CsNanos belongs to the Nanos2 subfamily. Csnanos expression was widely detected in various tissues, but the expression level was higher in testis and ovary. During early development and sex differentiation, Csnanos expression exhibited a clear sexually dimorphic pattern, suggesting its different roles in the migration and differentiation of primordial germ cells (PGCs). Higher expression levels of Csnanos mRNA in normal females and males than in neomales indicated that the nanos gene may play key roles in maintaining the differentiation of gonad. Moreover, medaka PGCs were successfully labeled by the microinjection of synthesized mRNA consisting of green fluorescence protein and the 3'-untranslated region of Csnanos. These findings provide new insights into nanos gene expression and function, and lay the foundation for further study of PGC development and applications in tongue sole breeding.

  8. A 3 Mb YAC contig in the region of Usher Ib on chromosome 11q

    SciTech Connect

    Kelley, P.M.; Overbeck, L.; Weston, M.

    1994-09-01

    Under syndrome type Ib, a recessive disorder characterized by deafness, retinitis pigmentosa, and vestibular dysfunction has been mapped to chromosome 11q13. A 3 Mb YAC contig has been constructed covering the critical region of Usher Ib and spanning over eight loci: D11S1321, D11S527, D11S533, OMP, D11S906, D11S911, D11S937, and D11S918. This contig was constructed by PCR screening using the above described DNA markers of the CEPH mega YAC library. Additional YACs were identified by data presented in the Genethon physical map. A long-range restriction map has been constructed from both YAC and genomic DNA using STS markers as probes. Cosmid libraries from a subset of YACs have been screened for the location of CpG islands. In addition, potential transcribed regions have been identified by 3{prime} exon trapping of cosmid pools and placed on the YAC physical map.

  9. Knockdown of pre-mRNA cleavage factor Im 25 kDa promotes neurite outgrowth

    SciTech Connect

    Fukumitsu, Hidefumi; Soumiya, Hitomi; Furukawa, Shoei

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer CFIm25 knockdown promoted NGF-induced neurite out growth from PC12 cells. Black-Right-Pointing-Pointer Depletion of CFIm25 did not influence the morphology of proliferating PC12 cells. Black-Right-Pointing-Pointer CFIm regulated NGF-induced neurite outgrowth via coordinating RhoA activity. Black-Right-Pointing-Pointer CFIm25 knockdown increase the number of primary dendrites of hippocampal neurons. -- Abstract: Mammalian precursor mRNA (pre-mRNA) cleavage factor I (CFIm) plays important roles in the selection of poly(A) sites in a 3 Prime -untranslated region (3 Prime -UTR), producing mRNAs with variable 3 Prime ends. Because 3 Prime -UTRs often contain cis elements that impact stability or localization of mRNA or translation, alternative polyadenylation diversifies utilization of primary transcripts in mammalian cells. However, the physiological role of CFIm remains unclear. CFIm acts as a heterodimer comprising a 25 kDa subunit (CFIm25) and one of the three large subunits-CFIm59, CFIm68, or CFIm72. CFIm25 binds directly to RNA and introduces and anchors the larger subunit. To examine the physiological roles of CFIm, we knocked down the CFIm25 gene in neuronal cells using RNA interference. Knockdown of CFIm25 increased the number of primary dendrites of developing hippocampal neurons and promoted nerve growth factor (NGF)-induced neurite extension from rat pheochromocytoma PC12 cells without affecting the morphology of proliferating PC12 cells. On the other hand, CFIm25 knockdown did not influence constitutively active or dominantly negative RhoA suppression or promotion of NGF-induced neurite extension from PC12 cells, respectively. Taken together, our results indicate that endogenous CFIm may promote neuritogenesis in developing neurons by coordinating events upstream of NGF-induced RhoA inactivation.

  10. Absence of mutations in the interspecies conserved regions of the CFTR promoter region in cystic fibrosis (CF) and CF related patients.

    PubMed Central

    Verlingue, C; Vuillaumier, S; Mercier, B; Le Gac, M; Elion, J; Férec, C; Denamur, E

    1998-01-01

    This study was aimed at testing if a 5.2 kb untranslated region on both sides of the first CFTR exon, shown to contain regulatory elements, could carry mutations responsible for cystic fibrosis (CF) or CF related phenotypes. Selection of the DNA segments studied within this region was based upon the identification of conserved sequences throughout evolution (phylogenetic footprints, PFs). Comparison of the CFTR sequences in eight species representing four orders of mammals (man, gibbon, rhesus monkey, squirrel, monkey, rabbit, cow, rat, and mouse) identified four clusters of PFs within the 3.9 kb of DNA sequence upstream from the initiation codon, as well as two nearby PFs at +1 kb within intron 1. Six DNA segments containing PFs were scanned for mutations by denaturing gradient gel electrophoresis (DGGE) in patients with CF (n = 29), congenital bilateral absence of the vas deferens (n = 143), or disseminated bronchiectasis (n = 33), for whom only one or no mutations had been identified despite extensive DGGE analysis of the 27 CFTR exons and exon/intron boundaries. Only one polymorphism (-966 T-->G) was identified with a frequency of 2.2% and no other sequence variations were found. This study reinforces the idea that the promoter region in the CFTR is not frequently mutated. PMID:9507393

  11. Rarity of DNA sequence alterations in the promoter region of the human androgen receptor gene.

    PubMed

    Cabral, D F; Santos, A; Ribeiro, M L; Mesquita, J C; Carvalho-Salles, A B; Hackel, C

    2004-12-01

    The human androgen receptor (AR) gene promoter lies in a GC-rich region containing two principal sites of transcription initiation and a putative Sp1 protein-binding site, without typical "TATA" and "CAAT" boxes. It has been suggested that mutations within the 5'untranslated region (5'UTR) may contribute to the development of prostate cancer by changing the rates of gene transcription and/or translation. In order to investigate this question, the aim of the present study was to search for the presence of mutations or polymorphisms at the AR-5'UTR in 92 prostate cancer patients, where histological diagnosis of adenocarcinoma was established in specimens obtained from transurethral resection or after prostatectomy. The AR-5'UTR was amplified by PCR from genomic DNA samples of the patients and of 100 healthy male blood donors, included as controls. Conformation-sensitive gel electrophoresis was used for DNA sequence alteration screening. Only one band shift was detected in one individual from the blood donor group. Sequencing revealed a new single nucleotide deletion (T) in the most conserved portion of the promoter region at position +36 downstream from the transcription initiation site I. Although the effect of this specific mutation remains unknown, its rarity reveals the high degree of sequence conservation of the human androgen promoter region. Moreover, the absence of detectable variation within the critical 5'UTR in prostate cancer patients indicates a low probability of its involvement in prostate cancer etiology.

  12. Identification of a missense mutation and several polymorphisms in the proenkephalin A gene of schizophrenic patients

    SciTech Connect

    Mikesell, M.J.; Sommer, S.S.; McMurray, C.T.

    1996-09-20

    Schizophrenia is a complex and severe disorder of unknown cause and pathophysiology. In this study, we examined the opioid hypothesis for schizophrenia at the molecular level, focusing on the dopamine-regulated proenkephalin A gene (chromosome 8q11.23-q12). We have screened 150 schizophrenic patients for sequence variations within the promoter region, entire coding sequence, and 3{prime}-untranslated region. We find one sequence change in a conserved amino acid that may be of functional significance. This mutation was found in a single schizophrenia patient but not in controls. Although several new, race-specific polymorphisms were identified, all other sequence changes appeared to be common polymorphisms, unlikely to contribute to the etiology of schizophrenia. 38 refs., 3 figs., 2 tabs.

  13. Stereoselective formation of a 2 prime (3 prime)- aminoacyl ester of a nucleotide

    NASA Technical Reports Server (NTRS)

    Weber, A. L.

    1986-01-01

    Reaction of DL-series and adenosine-5-phosphorimidazolide in the presence of adenosine-5'-(0-methylphosphate) and imidazole resulted in the stereoselective synthesis of the aminoacyl nucleotide ester, 2'(3')-0-seryl-adenosine-5'-(0-methylphosphate). The enantiomeric excess of D-serine incorporated into 2'(3')-0-seryl-adenosine-5'-(0-methylphosphate) was about 9%. Adenylyl-(5->N)-serine and an unknown product also incorporated an excess of D-serine, however, seryl-serine showed an excess of L-serine. The relationship of these results to the origin of the biological pairing of L-amino acids and nucleotides containing D-ribose is discussed.

  14. The 3 prime paradigm of the miR-200 family and other microRNAs

    PubMed Central

    Moschos, Sterghios; Spivack, Simon D

    2011-01-01

    The number of predicted human microRNAs in Sanger miRBase currently stands at over 1,000, with each of these in turn predicted to target numerous mRNAs. However, those microRNAs for which mRNA targets have been evaluated, verified and reported in the literature are still in the minority and the bulk of microRNA/mRNA interactions are yet to be confirmed. Confirmation of microRNA interaction with predicted mRNA targets represents a considerable undertaking, made more complex by potential synergistic effects of multiple microRNAs and the three possible outcomes (translational repression, degradation or a mixture of both). In addition, contrasting results obtained when either stably expressing or transiently transfecting members of the miR-200 family illustrate limitations in the verification methods currently in use. In this article we suggest that instead of allowing computational predictions to drive investigation, it would be desirable, when possible, to systematically evaluate microRNA targets using inducible, stable, ectopic expression. The advantage of stable lines ectopically expressing microRNA(s) is that they allow an analysis of changes to both the proteome and the transcriptome. This would allow verification of targets, improve the design of prediction algorithms and greatly increase our understanding of the outcome of microRNA/mRNA interaction. PMID:21242719

  15. The 3 prime paradigm of the miR-200 family and other microRNAs.

    PubMed

    Brock, Graham J; Moschos, Sterghios; Spivack, Simon D; Hurteau, Gregory J

    2011-03-01

    The number of predicted human microRNAs in Sanger miRBase currently stands at over a thousand, with each of these in turn predicted to target numerous mRNAs. However, those microRNAs for which mRNA targets have been evaluated, verified and reported in the literature are still in the minority and the bulk of microRNA/mRNA interactions are yet to be confirmed. Confirmation of microRNA interaction with predicted mRNA targets represents a considerable undertaking, made more complex by potential synergistic effects of multiple microRNAs and the three possible outcomes (translational repression, degradation or a mixture of both). In addition, contrasting results obtained when either stably expressing or transiently transfecting members of the miR-200 family illustrate limitations in the verification methods currently in use. In this article we suggest that instead of allowing computational predictions to drive investigation, it would be desirable, when possible, to systematically evaluate microRNA targets using inducible, stable, ectopic expression. The advantage of stable lines ectopically expressing microRNA(s) is that they allow an analysis of changes to both the proteome and the transcriptome. This would allow verification of targets, improve the design of prediction algorithms and greatly increase our understanding of the outcome of microRNA/mRNA interaction.

  16. 2[prime] and 3[prime] Carboranyl uridines and their diethyl ether adducts

    DOEpatents

    Soloway, A.H.; Barth, R.F.; Anisuzzaman, A.K.; Alam, F.; Tjarks, W.

    1992-12-15

    A process is described for preparing carboranyl uridine nucleoside compounds and their diethyl ether adducts, which exhibit a tenfold increase in boron content over prior art boron containing nucleoside compounds. The carboranyl uridine nucleoside compounds exhibit enhanced lipophilicity and hydrophilic properties adequate to enable solvation in aqueous media for subsequent incorporation of the compounds in methods for boron neutron capture therapy in mammalian tumor cells. No Drawings

  17. Returning "Region" to World Regional Geography

    ERIC Educational Resources Information Center

    Rees, Peter W.; Legates, Margaret

    2013-01-01

    World regional geography textbooks rarely focus on the process of region formation, despite frequent calls to reincorporate a regional approach to teaching global geography. An instructional strategy using problem-based learning in a small honors section of a large world regional geography course is described. Using a hypothetical scenario…

  18. Ionospheric research. [E region, F region, D region

    NASA Technical Reports Server (NTRS)

    1975-01-01

    Progress is reported in the following areas: D-region theory; E and F-region; wave propagation; mass spectrometer measurements; and atmospheric reactions. Various supporting operations are included: design and construction of instrumentation; and programming.

  19. The highly recombinogenic bz locus lies in an unusually gene-rich region of the maize genome.

    PubMed

    Fu, H; Park, W; Yan, X; Zheng, Z; Shen, B; Dooner, H K

    2001-07-17

    The bronze (bz) locus exhibits the highest rate of recombination of any gene in higher plants. To investigate the possible basis of this high rate of recombination, we have analyzed the physical organization of the region around the bz locus. Two adjacent bacterial artificial chromosome clones, comprising a 240-kb contig centered around the Bz-McC allele, were isolated, and 60 kb of contiguous DNA spanning the two bacterial artificial chromosome clones was sequenced. We find that the bz locus lies in an unusually gene-rich region of the maize genome. Ten genes, at least eight of which are shown to be transcribed, are contained in a 32-kb stretch of DNA that is uninterrupted by retrotransposons. We have isolated nearly full length cDNAs corresponding to the five proximal genes in the cluster. The average intertranscript distance between them is just 1 kb, revealing a surprisingly compact packaging of adjacent genes in this part of the genome. At least 11 small insertions, including several previously described miniature inverted repeat transposable elements, were detected in the introns and 3' untranslated regions of genes and between genes. The gene-rich region is flanked at the proximal and distal ends by retrotransposon blocks. Thus, the maize genome appears to have scattered regions of high gene density similar to those found in other plants. The unusually high rate of intragenic recombination seen in bz may be related to the very high gene density of the region.

  20. Sequence analysis of the ERCC2 gene regions in human, mouse, and hamster reveals three linked genes

    SciTech Connect

    Lamerdin, J.E.; Stilwagen, S.A.; Ramirez, M.H.

    1996-06-15

    The ERCC2 (excision repair cross-complementing rodent repair group 2) gene product is involved in transcription-coupled repair as an integral member of the basal transcription factor BTF2/TFIIH complex. Defects in this gene can result in three distinct human disorders, namely the cancer-prone syndrome xeroderma pigmentosum complementation group D, trichothiodystrophy, and Cockayne syndrome. We report the comparative analysis of 91.6 kb of new sequence including 54.3 kb encompassing the human ERCC2 locus, the syntenic region in the mouse (32.6 kb), and a further 4.7 kb of sequence 3{prime} of the previously reported ERCC2 region in the hamster. In addition to ERCC2, our analysis revealed the presence of two previously undescribed genes in all three species. The first is centromeric (in the human) to ERCC2 and is most similar to the kinesin light chain gene in sea urchin. The second gene is telomeric (in the human) to ERCC2 and contains a motif found in ankyrins, some cell proteins, and transcription factors. Multiple EST matches to this putative new gene indicate that it is expressed in several human tissues, including breast. The identification and description of two new genes provides potential candidate genes for disorders mapping to this region of 19q13.2. 42 refs., 6 figs., 3 tabs.

  1. Identification of the functional elements in the promoter region of human DNA topoisomerase IIIbeta gene.

    PubMed

    Cho, Young Hoon; Park, Jee Young; Han, Sang Youp; Chung, In Kwon

    2004-09-17

    In this study, we have isolated and characterized the promoter region of the human DNA topoisomerase IIIbeta (hTOP3beta) gene. The 5' RACE assay showed a short exon 1 encoding only the 35-bp untranslated region and suggested the presence of multiple transcription initiation sites. The hTOP3beta gene promoter lacks a canonical TATA box or initiation element and is moderately high in GC content. Transient expression of a luciferase reporter gene under the control of serially deleted 5'-flanking sequence identified an activator element between -141 and -119 upstream of the transcription initiation site and a second regulatory element between -91 and -71. On the basis of scanning mutations of triple nucleotides, we demonstrated that a 5'GGAACC3' element between -117 and -112 plays a critical role in the up-regulation of the basal transcription activity. Changing the 5'GGAACC3' sequence leads to markedly reduced promoter activity. Gel mobility shift assays revealed that the 5'GGAACC3' element is required for DNA binding by the transcription factor complex. These observations lead to the conclusion that the positive regulatory region including the 5'GGAACC3' core element is essential for efficient expression of the hTOP3beta gene as well as for the binding of as yet unidentified regulatory factor(s).

  2. Exon organization of the mouse entactin gene corresponds to the structural domains of the polypeptide and has regional homology to the low-density lipoprotein receptor gene

    SciTech Connect

    Durkin, M.E.; Chung, A.E.; Wewer, U.M.

    1995-03-20

    Entactin is a widespread basement membrane protein of 150 kDa that binds to type IV collagen and laminin. The complete exon-intron structure of the mouse entactin gene has been determined from {lambda} genomic DNA clones. The gene spans at least 65 kb and contains 20 exons. The exon organization of the mouse entactin gene closely corresponds to the organization of the polypeptide into distinct structural and functional domains. The two amino-terminal globular domains are encoded by three exons each. Single exons encode the two protease-sensitive, O-glycosylated linking regions. The six EGF-like repeats and the single thyroglobulin-type repeat are each encoded by separate exons. The carboxyl-terminal half of entactin displays sequence homology to the growth factor-like region of the low-density lipoprotein receptor, and in both genes this region is encoded by eight exons. The positions of four introns are also conserved in the homologous region of the two genes. These observations suggest that the entactin gene has evolved via exon shuffling. Finally, several sequence polymorphisms useful for gene linkage analysis were found in the 3{prime} noncoding region of the last exon. 52 refs., 8 figs.

  3. Hereditary vitamin D resistant rickets due to deletion of exon 3 of the vitamin D receptor

    SciTech Connect

    Rut, A.R.; O`Riordan, J.L.H.; Hughes, M.R.

    1994-09-01

    Hereditary vitamin D resistant rickets is an autosomal recessive disorder characterized by severe rickets, hypolcalcaemia, secondary hyperparathyroidism and occasionally, the absence of body hair. The pathological process involves resistance of target tissues to the actions of calcitriol [1,25(OH{sub 2}D{sub 3})], the hormonal form of vitamin D. Calcitriol mediates its actions through a nuclear receptor (VDR) which has been cloned and shown to be a member of the superfamily of steriod/thyroid/retinoic acid receptors. Skin fibroblasts were obtained from a Greek child with characteristic features of the condition. Total RNA was extracted from rapidly dividing cells and reverse transcribed. The coding region was amplified by PCR with primers 31a in the 5{prime} untranslated region and 31b in the 3{prime} untranslated region of the VDR cDNA sequence. The 5{prime} and 3{prime} halves of VDR were further amplified using primers tagged with M13 forward and reverse primer sequences. The whole process was carried out in duplicate starting with RNA. Sequence data was obtained using Taq dye primer cycle sequencing (ABI). Agarose gel electrophoresis revealed that the 5{prime} product was approximately 100 bp shorter than control. This was confirmed by sequencing which demonstrated a 131 bp deletion of the C-terminal part of the DNA binding domain (bases 147-277). Bases 147-277 are coded for by exon 3 and this deletion is bounded by the splice junctions. This is the first report of a deletion in VDR in any patient with vitamin D-resistant rickets. Such a deletion not only removes the second zinc finger but also results in a frameshift that corrupts the remainder of the receptor. Such a deletion may have arisen as a result of a microdeletion of genomic DNA or, more likely, as a result of defective splicing.

  4. miR-214 regulates lactoferrin expression and pro-apoptotic function in mammary epithelial cells.

    PubMed

    Liao, Yalin; Du, Xiaogu; Lönnerdal, Bo

    2010-09-01

    Lactoferrin (Lf) is an abundantly expressed protein in human milk. Lactoferrin exhibits several important biological functions, and its expression is regulated by multiple environmental factors. Cellular endogenous factors, however, have not been extensively studied with regard to lactoferrin gene expression. In this study, we showed that lactoferrin gene expression and function are directly targeted by miR-214 in HC11 and MCF7 cells. In the lactoferrin mRNA 3 prime untranslated region (UTR) of human, mouse, rat, pig, bovine, camel, and goat species, there is a conserved region that perfectly matches the seed region of miR-214. Transfection of miR-214 mimic in HEK293 cells dose-dependently inhibited the activity of pGL3-control vector containing lactoferrin mRNA 3 prime UTR downstream of the luciferase gene. In HC11 cells, miR-214 overexpression inhibited the induction of lactoferrin expression by beta -estradiol (E2) and dexamethasone-prolactin-insulin (DPI). Furthermore, in MCF7 cells, overexpression of miR-214 markedly decreased lactoferrin expression (P lt 0.05), and inhibition of endogenous miR-214 expression increased lactoferrin expression and cellular apoptotic activities (P lt 0.05). In summary, our data showed that miR-214 is directly involved in lactoferrin expression and lactoferrin mediated cancer susceptibility (proapoptotic activities) in mammary epithelial cells.

  5. NcoI and TaqI RFLPs for human M creatine kinase (CKM)

    SciTech Connect

    Perryman, M.B.; Hejtmancik, J.F.; Ashizawa, Tetsuo; Armstrong, R.; Lin, Sunchiang; Roberts, R.; Epstein, H.F. )

    1988-09-12

    Probe pHMCKUT contains a 135 bp cDNA fragment inserted into pGEM 3. The probe corresponds to nucleotides 1,201 to 1,336 located in the 3{prime} untranslated region of human M creatine kinase. The probe is specific for human M creatine kinase and does not hybridize to human B cretine kinase sequences. NcoI identifies a two allele polymorphism of a band at either 2.5 kb or 3.6 kb. TaqI identifies a two allele polymorphism at either 3.8 kb or 4.5 kb. Human M creatine has been localized to chromosome 19q. Autosomal co-dominant inheritance was shown in six informative Caucasian families.

  6. No association between schizophrenia and polymorphisms within the genes for debrisoquine 4-hydroxylase (CYP2D6) and the dopamine transporter (DAT)

    SciTech Connect

    Daniels, J.; Williams, J.; Asherson, P.; McGuffin, P.; Owen, M.

    1995-02-27

    It has been suggested that the cytochrome P450 mono-oxygenase, debrisoquine 4-hydroxylase, is involved in the catabolism and processing of neurotransmitters subsequent to their reuptake into target cells. It is also thought to be related to the dopamine transporter that acts to take released dopamine back up into presynaptic terminals. The present study used the association approach to test the hypothesis that mutations in the genes for debrisoquine 4-hydroxylase (CYP2D6) and the dopamine transporter (DAT) confer susceptibility to schizophrenia. There were no differences in allele or genotype frequencies between patients and controls in the mutations causing the poor metaboliser phenotype in CYP2D6. In addition there was no association found between schizophrenia and a 48 bp repeat within the 3{prime} untranslated region of DAT. 18 refs., 2 tabs.

  7. Aberrant splicing and transcription termination caused by P element insertion into the intron of a Drosophila gene

    SciTech Connect

    Horowitz, H.; Berg, C.A.

    1995-01-01

    Insertional mutagenesis screens using the P[lacZ, rosy{sup +}] (PZ) transposable element have provided thousands of mutant lines for analyzing genes of varied function in the fruitfly, Drosophila melanogaster. As has been observed with other P elements, many of the PZ-induced mutations result from insertion of the P element into the promoter or 5{prime} untranslated regions of the affected gene. We document here a novel mechanism for mutagenesis by this element. We show that sequences present within the element direct aberrant splicing and termination events that produce an mRNA composed of 5{prime} sequences from the mutated gene (in this case, pipsqueak) and 3{prime} sequences from within the P[lacZ, rosy{sup +}] element. These truncated RNAs could yield proteins with dominant mutant effects. 43 refs., 4 figs.

  8. Regional Sustainable Environmental Management

    EPA Science Inventory

    Regional sustainable environmental management is an interdisciplinary effort to develop a sufficient understanding of the interactions between ecosystems, the economy, law, and technology to formulate effective long-term management strategies on a regional scale. Regional sustai...

  9. Genome-wide analyses in bacteria show small-RNA enrichment for long and conserved intergenic regions.

    PubMed

    Tsai, Chen-Hsun; Liao, Rick; Chou, Brendan; Palumbo, Michael; Contreras, Lydia M

    2015-01-01

    Interest in finding small RNAs (sRNAs) in bacteria has significantly increased in recent years due to their regulatory functions. Development of high-throughput methods and more sophisticated computational algorithms has allowed rapid identification of sRNA candidates in different species. However, given their various sizes (50 to 500 nucleotides [nt]) and their potential genomic locations in the 5' and 3' untranslated regions as well as in intergenic regions, identification and validation of true sRNAs have been challenging. In addition, the evolution of bacterial sRNAs across different species continues to be puzzling, given that they can exert similar functions with various sequences and structures. In this study, we analyzed the enrichment patterns of sRNAs in 13 well-annotated bacterial species using existing transcriptome and experimental data. All intergenic regions were analyzed by WU-BLAST to examine conservation levels relative to species within or outside their genus. In total, more than 900 validated bacterial sRNAs and 23,000 intergenic regions were analyzed. The results indicate that sRNAs are enriched in intergenic regions, which are longer and more conserved than the average intergenic regions in the corresponding bacterial genome. We also found that sRNA-coding regions have different conservation levels relative to their flanking regions. This work provides a way to analyze how noncoding RNAs are distributed in bacterial genomes and also shows conserved features of intergenic regions that encode sRNAs. These results also provide insight into the functions of regions surrounding sRNAs and into optimization of RNA search algorithms.

  10. Genome-Wide Analyses in Bacteria Show Small-RNA Enrichment for Long and Conserved Intergenic Regions

    PubMed Central

    Tsai, Chen-Hsun; Liao, Rick; Chou, Brendan; Palumbo, Michael

    2014-01-01

    Interest in finding small RNAs (sRNAs) in bacteria has significantly increased in recent years due to their regulatory functions. Development of high-throughput methods and more sophisticated computational algorithms has allowed rapid identification of sRNA candidates in different species. However, given their various sizes (50 to 500 nucleotides [nt]) and their potential genomic locations in the 5′ and 3′ untranslated regions as well as in intergenic regions, identification and validation of true sRNAs have been challenging. In addition, the evolution of bacterial sRNAs across different species continues to be puzzling, given that they can exert similar functions with various sequences and structures. In this study, we analyzed the enrichment patterns of sRNAs in 13 well-annotated bacterial species using existing transcriptome and experimental data. All intergenic regions were analyzed by WU-BLAST to examine conservation levels relative to species within or outside their genus. In total, more than 900 validated bacterial sRNAs and 23,000 intergenic regions were analyzed. The results indicate that sRNAs are enriched in intergenic regions, which are longer and more conserved than the average intergenic regions in the corresponding bacterial genome. We also found that sRNA-coding regions have different conservation levels relative to their flanking regions. This work provides a way to analyze how noncoding RNAs are distributed in bacterial genomes and also shows conserved features of intergenic regions that encode sRNAs. These results also provide insight into the functions of regions surrounding sRNAs and into optimization of RNA search algorithms. PMID:25313390

  11. The leader region of Laminin B1 mRNA confers cap-independent translation.

    PubMed

    Petz, Michaela; Kozina, Daniela; Huber, Heidemarie; Siwiec, Tanja; Seipelt, Joachim; Sommergruber, Wolfgang; Mikulits, Wolfgang

    2007-01-01

    Translation initiation of eukaryotic mRNAs generally occurs by cap-dependent ribosome scanning. However, certain mRNAs contain internal ribosome entry sites (IRES) allowing cap-independent translation. Several of these IRES-competent transcripts and their corresponding proteins are involved in tumourigenesis. This study focused on IRES-driven translation control during the epithelial to mesenchymal transition (EMT) of hepatocytes that reflects crucial aspects of carcinoma progression. Expression profiling of EMT revealed Laminin B1 (LamB1) to be translationally upregulated. The 5'-untranslated region (UTR) of LamB1 was potent to direct IRES-dependent mRNA utilization of a bicistronic reporter construct. Stringent assays for cryptic promoter and splice sites showed no aberrantly expressed transcripts, suggesting that the reporter activity provided by the leader region of LamB1 mRNA exclusively depends on IRES. In accordance, LamB1 expression increased upon negative interference with cap-dependent translation by expression of human rhinovirus 2A protease or heat shock of cells. Finally, the enhanced expression of LamB1 during EMT correlated with an elevated IRES activity. Together, these data provide first evidence that the 5'-UTR of LamB1 contains a bona fide IRES that directs translational upregulation of LamB1 during stress conditions and neoplastic progression of hepatocytes.

  12. Regional differences in the genetic variability of Finno-Ugric speaking Komi populations.

    PubMed

    Khrunin, Andrey; Verbenko, Dmitry; Nikitina, Kseniya; Limborska, Svetlana

    2007-01-01

    The Komi (Komi-Zyryan) people are one of the most numerous ethnic groups belonging to the Finno-Ugric linguistic community. They occupy an extensive territory in north Russia to the west of the Ural Mountains, in the northeast of the East European Plain. This is an area of long-term interactions between Europeans and North Asians. Genetic variability was evaluated in two geographically distinct populations, the Izhemski and Priluzski Komi. We searched for polymorphisms of the TP53 gene (a 16-bp duplication in intron 3 and three RFLPs: for Bsh1236I at codon 72, for MspI in intron 6, and for BamHI in the 3' flanking region) and for variable number tandem repeat (VNTR) polymorphisms of locus D1S80 and of the 3' untranslated region of the gene for apolipoprotein B (ApoB). Some data from our previous studies of TP53, 3'ApoB, and D1S80 variability were involved in the comparison of Komi with other Eastern European populations. Multidimensional scaling analysis of genetic distances was used for the evaluation of genetic relationships between populations. The results revealed some affinity between Priluzski Komi and Eastern Slavonic populations, and significant segregation of Izhemski Komi from other ethnic groups studied. The unique genetic features of Izhemski Komi may have been determined by their ethnogenesis or the pressure of environmental factors, such as special nutrition and adaptation to extreme climatic conditions.

  13. Nucleotide sequence of the 3'-terminal region of potato virus YN RNA.

    PubMed

    van der Vlugt, R; Allefs, S; de Haan, P; Goldbach, R

    1989-01-01

    The sequence of the 3'-terminal 1611 nucleotides of the genome of the tobacco veinal necrosis strain of potato virus Y (PVYN) was determined. The sequence revealed an open reading frame of 1285 nucleotides, of which the start was not identified, and an untranslated region of 316 nucleotides upstream of a poly(A) tract. Comparison of the open reading frame with the amino-terminal sequence of the viral coat protein enabled mapping of the start of the coat protein at amino acid -267, and indicated that maturation of this protein requires proteolytic processing from a larger polyprotein precursor at a glutamine/glycine dipeptide sequence. The coat protein of PVYN displayed significant (51 to 63%) sequence homology to the coat proteins of four other potyviruses, tobacco etch virus, tobacco vein mottling virus, plum pox virus and sugarcane mosaic virus. Even higher sequence homology (91%) was detected with the coat protein of a fifth potyvirus, pepper mottle virus (PeMV). This homology was of the same level as found between the coat proteins of PVYN and a second strain of this virus, PVYD. Since, moreover, PVYN and PeMV were the only potyviruses displaying homology in the 3'-terminal, non-translated regions of their genomes, we conclude that PeMV should be regarded as a strain of PVY.

  14. Association study of the CNR1 gene exon 3 alternative promoter region polymorphisms and substance dependence

    PubMed Central

    Herman, Aryeh I.; Kranzler, Henry R.; Cubells, Joseph F.; Gelernter, Joel; Covault, Jonathan

    2008-01-01

    An alternative promoter producing a novel 5′-untranslated region of cannabinoid receptor mRNA has recently been described in CNR1, the gene encoding the cannabinoid receptor protein. Single nucleotide polymorphisms (SNPs) adjacent to this site were reported to be associated with polysubstance abuse (Zhang et al. 2004). We examined the association of 4 SNPs (rs6928499, rs806379, rs1535255, rs2023239) in the distal region of intron 2 of CNR1 both with individual substance dependence diagnoses (i.e., alcohol, cocaine, and opioids), as well as with polysubstance dependence. The study samples consisted of European American and African American subjects with drug and or alcohol dependence (n=895), and controls (n = 472). Subjects were grouped as polysubstance dependent, opioid dependent, cocaine dependent, cannabis dependent and alcohol dependent. There was a modest association of marker rs1535255 with alcohol dependence, respectively (P=0.04), though with correction for multiple phenotype comparisons, this effect was not considered statistically significant. These findings fail to replicate the original report of an association between SNPs adjacent to an alternative CNR1 exon 3 transcription start site and polysubstance abuse. PMID:16741937

  15. Analysis of the promoter region of the gene encoding NAD-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase.

    PubMed Central

    Bélanger, C; Peri, K G; MacKenzie, R E

    1991-01-01

    Sequence analysis of the 5'-flanking region of the gene encoding NAD-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase (NMDMC) revealed several putative cis-regulatory elements. To delineate the function of these regulatory elements, various deletion mutants of the 5'-flanking region were connected to the reporter gene chloramphenicol acetyltransferase (CAT) and promoter activity was measured in transient transfection assays. Transfection experiments performed with the sequence extending from -508 to +59 produced a high-level transient expression of the CAT gene in BALB/c 3T3-SV-T2 and NIH 3T3 cells. Removal of the sequence from +16 to +59 which includes the second transcription start point at +43, a TATA-like box and 5'-untranslated sequences abolished the promoter activity. Deletion analysis of 5'-upstream sequences revealed that the region from positions -55 to +59 is sufficient to mediate a high CAT activity comparable to the level obtained with the construct -508/+59. Within this region are found a CAAT box, a TATA-like box and two putative GC boxes. A functional analysis of the promoter showed that the sequence from -55 to +59 is sufficient to respond to stimulation by serum. Images PMID:1843253

  16. miR-148 targets human DNMT3b protein coding region.

    PubMed

    Duursma, Anja M; Kedde, Martijn; Schrier, Mariette; le Sage, Carlos; Agami, Reuven

    2008-05-01

    MicroRNAs (miRNAs) are small noncoding RNA molecules of 20-24 nucleotides that regulate gene expression. In animals, miRNAs form imperfect interactions with sequences in the 3' Untranslated region (3'UTR) of mRNAs, causing translational inhibition and mRNA decay. In contrast, plant miRNAs mostly associate with protein coding regions. Here we show that human miR-148 represses DNA methyltransferase 3b (Dnmt3b) gene expression through a region in its coding sequence. This region is evolutionary conserved and present in the Dnmt3b splice variants Dnmt3b1, Dnmt3b2, and Dnmt3b4, but not in the abundantly expressed Dnmt3b3. Whereas overexpression of miR-148 results in decreased DNMT3b1 expression, short-hairpin RNA-mediated miR-148 repression leads to an increase in DNMT3b1 expression. Interestingly, mutating the putative miR-148 target site in Dnmt3b1 abolishes regulation by miR-148. Moreover, endogenous Dnmt3b3 mRNA, which lacks the putative miR-148 target site, is resistant to miR-148-mediated regulation. Thus, our results demonstrate that the coding sequence of Dnmt3b mediates regulation by the miR-148 family. More generally, we provide evidence that coding regions of human genes can be targeted by miRNAs, and that such a mechanism might play a role in determining the relative abundance of different splice variants.

  17. Mutations in noncoding regions of GJB1 are a major cause of X-linked CMT

    PubMed Central

    Tomaselli, Pedro J.; Rossor, Alexander M.; Horga, Alejandro; Jaunmuktane, Zane; Carr, Aisling; Saveri, Paola; Piscosquito, Giuseppe; Pareyson, Davide; Laura, Matilde; Blake, Julian C.; Poh, Roy; Polke, James; Houlden, Henry

    2017-01-01

    Objective: To determine the prevalence and clinical and genetic characteristics of patients with X-linked Charcot-Marie-Tooth disease (CMT) due to mutations in noncoding regions of the gap junction β-1 gene (GJB1). Methods: Mutations were identified by bidirectional Sanger sequence analysis of the 595 bases of the upstream promoter region, and 25 bases of the 3′ untranslated region (UTR) sequence in patients in whom mutations in the coding region had been excluded. Clinical and neurophysiologic data were retrospectively collected. Results: Five mutations were detected in 25 individuals from 10 kindreds representing 11.4% of all cases of CMTX1 diagnosed in our neurogenetics laboratory between 1996 and 2016. Four pathogenic mutations, c.-17G>A, c.-17+1G>T, c.-103C>T, and c.-146-90_146-89insT were detected in the 5′UTR. A novel mutation, c.*15C>T, was detected in the 3′ UTR of GJB1 in 2 unrelated families with CMTX1 and is the first pathogenic mutation in the 3′UTR of any myelin-associated CMT gene. Mutations segregated with the phenotype, were at sites predicted to be pathogenic, and were not present in the normal population. Conclusions: Mutations in noncoding DNA are a major cause of CMTX1 and highlight the importance of mutations in noncoding DNA in human disease. Next-generation sequencing platforms for use in inherited neuropathy should therefore include coverage of these regions. PMID:28283593

  18. Epidermal surface antigen (MS17S1) is highly conserved between mouse and human

    SciTech Connect

    Cho, Y.J.; Chema, D.; Cho, M.

    1995-05-20

    A mouse monoclonal antibody ECS-1 raised to human keratinocytes detects a 35-kDa epidermal surface antigen (ESA) and causes keratinocyte dissociation in vitro. ECS-1 stains skin of 16-day mouse embryo and 8- to 9-week human fetus. Mouse Esa cDNA encodes a 379-amino-acid protein that is 99.2% identical to the human, differing at only 3 amino acids. The gene (M17S1) was mapped to mouse chromosome 11, highlighting the conserved linkage synteny existing between human chromosome 17 and mouse chromosome 11. Although the nude locus has been mapped to the same region of chromosome 11, no abnormalities in protein, mRNA, or cDNA or genomic sequences were detected in nude mice. However, both nude and control mice were found to have a second Esa mRNA transcript that conserves amino acid sequence and molecular weight. The mouse and human 5{prime} and 3{prime} untranslated sequences are conserved. Similar RNA folding patterns of the 5{prime} untranslated region are predicted despite a 91-bp insertion in the mouse. These data suggest that both the function and the regulation of ESA protein are of importance and that Esa (M17S1) is not the nude locus gene. 42 refs., 7 figs., 3 tabs.

  19. Identification of brain-derived neurotrophic factor promoter regions mediating tissue-specific, axotomy-, and neuronal activity-induced expression in transgenic mice

    PubMed Central

    1995-01-01

    The structure of rat brain-derived neurotrophic factor (BDNF) gene is complex; four 5' exons are linked to separate promoters and one 3' exon is encoding the BDNF protein. To analyze the relative importance of the regulatory regions in vivo, we have generated transgenic mice with six different promoter constructs of the BDNF gene fused to the chloramphenicol acetyl transferase reporter gene. High level and neuronal expression of the reporter gene, that in many respects recapitulated BDNF gene expression, was achieved by using 9 kb of genomic sequences covering the promoter regions that lie adjacent to each other in the genome (promoters I and II and promoters III and IV, respectively) and by including sequences of BDNF intron-exon splice junctions and 3' untranslated region in the constructs. The genomic regions responsible for the in vivo upregulation of BDNF expression in the axotomized sciatic nerve and in the brain after kainic acid-induced seizures and KCl-induced spreading depression were mapped. These data show that regulation of the different aspects of BDNF expression is controlled by different regions in vivo, and they suggest that these promoter constructs may be useful for targeted expression of heterologous genes to specific regions of the central and peripheral nervous systems in an inducible manner. PMID:7822414

  20. Colonial Museums in the Us (Un)translated

    ERIC Educational Resources Information Center

    Valdeón, Roberto A.

    2015-01-01

    This paper examines the role of museums in the creation of anglophone stories in the USA, and how the (non-)translation of signs contributes to create a narrative of exclusion vis-à-vis other groups, notably native Americans, the Spanish, and the French. Particular attention is paid to open-air museums that preserve old buildings and areas…

  1. Characterization of direct selected cDNAs from the BRCA1 region of 17q21

    SciTech Connect

    Welcsh, P.L.; Osborne-Lawrence, S.L.; Spillman, M.A.

    1994-09-01

    A gene involved in the development of early-onset familial breast and ovarian cancer, BRCA1, has been mapped to human chromosome 17q21. Polymorphisms closely linked to BRCA1 has been sublocalized to a region of 17q21 which is defined by the markers D17S856 and D17S78. A physical map of this region, that consists of yeast artificial chromosome (YAC) and cosmid contigs, has been constructed and used to isolate potential coding sequences via direct selection. We have identified at least 23 unique transcripts in a 600 kb interval corresponding to approximately one gene every 30 kb. We have determined the expression profile of these cDNAs by generating cDNA-specific primers which have been used in a screen of cDNAs derived from wide variety of tissues and cell types. Full length cDNA clones are being obtained from cDNA libraries in which the genes have been shown to be expressed by a variety of techniques which include direct screening, 5{prime} and 3{prime} RACE, anchor PCR as well as modified selection procedures. We are currently screening for mutations in these candidate cDNAs in affected family members known to harbor a germ-line BRCA1 mutation and in sporadic breast and ovarian tumors. Mutation screening is being performed by Southern and Northern blotting, DNA sequencing, and SSCP analysis of germline DNA and cDNA. Finally, we are analyzing these candidate cDNAs in a number of breast and ovarian cancer cell lines for induction by known mitogenic factors such as estrogen and progesterone by Northern blotting and RT-PCR.

  2. ERLN Regional Support

    EPA Pesticide Factsheets

    Regional labs play important roles in the Environmental Response Laboratory Network. They can serve as point of contact; coordinate sample flow, special analytical service requests, or training exercises; and partner with regional emergency/disaster staff.

  3. Region 9 RTOC Charter

    EPA Pesticide Factsheets

    U.S. EPA Pacific Southwest (Region 9) Regional Tribal Operations Committee (RTOC) Charter as amended 11/13/2014: Mission, Goals, Scope, Structure & Membership, Meetings, Administration, Charter Amendment/Review, and Current Working Draft.

  4. Utah: Salt Lake Region

    Atmospheric Science Data Center

    2014-05-15

    article title:  Winter and Summer Views of the Salt Lake Region     View Larger Image Magnificent views of the region surrounding Salt Lake City, Utah are captured in these winter and summer images from the ...

  5. Region 9 Tribal Program

    EPA Pesticide Factsheets

    EPA Region 9 helps tribes strengthen management of environmental programs in Indian country, and ensure that tribes have a voice in decisions. The region serves 148 federally recognized tribes with the states of Arizona, California, and Nevada.

  6. Regional Smart Growth Alliances

    EPA Pesticide Factsheets

    This page describes the Urban Land Institute regional smart growth alliances that received funding from EPA to help support economic development, accommodate growth, enhance quality of, and protect the environment in regions across the country.

  7. Hepatitis C virus 3'X region interacts with human ribosomal proteins.

    PubMed

    Wood, J; Frederickson, R M; Fields, S; Patel, A H

    2001-02-01

    To identify proteins that can bind the 3' untranslated region (UTR) of hepatitis C virus (HCV) we screened human cDNA libraries using the Saccharomyces cerevisiae three-hybrid system. Screening with an RNA sequence derived from the 3'-terminal 98 nucleotides (3'X region) of an infectious clone of HCV (H77c) yielded clones of human ribosomal proteins L22, L3, S3, and mL3, a mitochondrial homologue of L3. We performed preliminary characterization of the binding between the 3'X region and these proteins by a three-hybrid mating assay using mutant 3'X sequences. We have further characterized the interaction between 3'X and L22, since this protein is known to be associated with two small Epstein-Barr virus (EBV)-encoded RNA species (EBERs) which are abundantly produced in cells latently infected with EBV. The EBERs, which have similar predicted secondary structure to the HCV 3'X, assemble into ribonucleoprotein particles that include L22 and La protein. To confirm that L22 binds HCV 3'X we performed in vitro binding assays using recombinant L22 (expressed as a glutathione S-transferase [GST] fusion protein) together with a 3'X riboprobe. The 3'X region binds to the GST-L22 fusion protein (but not to GST alone), and this interaction is subject to competition with unlabeled 3'X RNA. To establish the functional role played by L22 in internal ribosome entry site (IRES)-mediated translation of HCV sequences we performed translational analysis in HuH-7 cells using monocistronic and bicistronic reporter constructs. The relative amount of core-chloramphenicol acetyltransferase reporter protein translated under the control of the HCV IRES was stimulated in the presence of L22 and La when these proteins were supplied in trans.

  8. [Regional aging in Germany].

    PubMed

    Bucher, H

    1996-01-01

    Elderly people in Germany have a specific regional distribution. Recent regional population projections show that these patterns will change. The most dynamic process of aging will take place in the suburban parts of the large western Germany agglomerations, whereas in eastern Germany aging concentrates in regions with a lower density. There will be a regional deconcentration of elderly people with consequences for the planning of infrastructure.

  9. Learning Regions in Germany

    ERIC Educational Resources Information Center

    Thinesse-Demel, Jutta

    2010-01-01

    In 2000, the German Federal Ministry of Education and Research (BMBF) launched the programme "Learning Regions--Providing Support for Networks'" in cooperation with the Lander. It was co-financed by the European Social Fund (ESF). Some 90 regions were selected and financially supported. After one year, 71 regions continued to build-up…

  10. Genetic diversity of bovine viral diarrhea viruses from the Galicia region of Spain

    PubMed Central

    Factor, C.; Yus, E.; Eiras, C.; Sanjuan, M. L.; Cerviño, M.; Arnaiz, I.; Diéguez, F. J.

    2016-01-01

    This study examined the frequency and diversity of bovine viral diarrhoea viruses (BVDVs) infecting cattle in Galicia (northwestern Spain). A total of 86 BVDV strains were typed in samples of serum from 79 persistently infected animals and 3 viraemic animals and of abomasal fluid from 4 fetuses. Samples came from 73 farms participating in a voluntary BVDV control programme. Typing was based on a 288-bp sequence from the 5′ untranslated region amplified using primers 324 and 326. Of the 86 strains, 85 (98.8 per cent) belonged to species BVDV-1 and 1 (1.2 per cent) belonged to BVDV-2; 73 strains (84.9 per cent) were typed as BVDV-1b, 2 as BVDV-1e and 6 as BVDV-1d. One strain each was typed as belonging to 1a, 1h, 1k and 1l. The sole BVDV-2 strain was classified as 2a. These results identify BVDV-1b as the predominant species, and they indicate the presence of viral types not previously described anywhere in Spain. This is also the first report of BVDV-2 in Galicia and only the second report of BVDV-2 in Spain. PMID:27843559

  11. Mutations in the promoter region of the aldolase B gene that cause hereditary fructose intolerance.

    PubMed

    Coffee, Erin M; Tolan, Dean R

    2010-12-01

    Hereditary fructose intolerance (HFI) is a potentially fatal inherited metabolic disease caused by a deficiency of aldolase B activity in the liver and kidney. Over 40 disease-causing mutations are known in the protein-coding region of ALDOB. Mutations upstream of the protein-coding portion of ALDOB are reported here for the first time. DNA sequence analysis of 61 HFI patients revealed single base mutations in the promoter, intronic enhancer, and the first exon, which is entirely untranslated. One mutation, g.-132G>A, is located within the promoter at an evolutionarily conserved nucleotide within a transcription factor-binding site. A second mutation, IVS1+1G>C, is at the donor splice site of the first exon. In vitro electrophoretic mobility shift assays show a decrease in nuclear extract-protein binding at the g.-132G>A mutant site. The promoter mutation results in decreased transcription using luciferase reporter plasmids. Analysis of cDNA from cells transfected with plasmids harboring the IVS1+1G>C mutation results in aberrant splicing leading to complete retention of the first intron (~5 kb). The IVS1+1G>C splicing mutation results in loss of luciferase activity from a reporter plasmid. These novel mutations in ALDOB represent 2% of alleles in American HFI patients, with IVS1+1G>C representing a significantly higher allele frequency (6%) among HFI patients of Hispanic and African-American ethnicity.

  12. Genetic diversity of bovine viral diarrhea viruses from the Galicia region of Spain.

    PubMed

    Factor, C; Yus, E; Eiras, C; Sanjuan, M L; Cerviño, M; Arnaiz, I; Diéguez, F J

    2016-01-01

    This study examined the frequency and diversity of bovine viral diarrhoea viruses (BVDVs) infecting cattle in Galicia (northwestern Spain). A total of 86 BVDV strains were typed in samples of serum from 79 persistently infected animals and 3 viraemic animals and of abomasal fluid from 4 fetuses. Samples came from 73 farms participating in a voluntary BVDV control programme. Typing was based on a 288-bp sequence from the 5' untranslated region amplified using primers 324 and 326. Of the 86 strains, 85 (98.8 per cent) belonged to species BVDV-1 and 1 (1.2 per cent) belonged to BVDV-2; 73 strains (84.9 per cent) were typed as BVDV-1b, 2 as BVDV-1e and 6 as BVDV-1d. One strain each was typed as belonging to 1a, 1h, 1k and 1l. The sole BVDV-2 strain was classified as 2a. These results identify BVDV-1b as the predominant species, and they indicate the presence of viral types not previously described anywhere in Spain. This is also the first report of BVDV-2 in Galicia and only the second report of BVDV-2 in Spain.

  13. Triptolide inhibits COX-2 expression by regulating mRNA stability in TNF-{alpha}-treated A549 cells

    SciTech Connect

    Sun, Lixin; Zhang, Shuang; Jiang, Zhenzhou; Huang, Xin; Wang, Tao; Huang, Xiao; Li, Han; Zhang, Luyong

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer Triptolide inhibited COX-2 expression and the half-life of COX-2 mRNA is decreased. Black-Right-Pointing-Pointer The HuR protein shuttling from nucleus to cytoplasm is inhibited by triptolide. Black-Right-Pointing-Pointer Triptolide inhibited 3 Prime -UTR fluorescence reporter gene activity. Black-Right-Pointing-Pointer COX-2 mRNA binding to HuR is decreased by triptolide in pull-down experiments. -- Abstract: Cyclooxygenase-2 (COX-2) over-expression is frequently associated with human non-small-cell lung cancer (NSCLC) and involved in tumor proliferation, invasion, angiogenesis and resistance to apoptosis. In the present study, the effects of triptolide on COX-2 expression in A549 cells were investigated and triptolide was found to inhibit TNF-{alpha}-induced COX-2 expression. In our further studies, it was found that triptolide decreased the half-life of COX-2 mRNA dramatically and that it inhibited 3 Prime -untranslated region (3 Prime -UTR) fluorescence reporter gene activity. Meanwhile, triptolide inhibited the HuR shuttling from nucleus to cytoplasm. After triptolide treatment, decreased COX-2 mRNA in pull-down experiments with anti-HuR antibodies was observed, indicating that the decreased cytoplasmic HuR is responsible for the decreased COX-2 mRNA. Taken together, our results provided evidence for the first time that triptolide inhibited COX-2 expression by COX-2 mRNA stability modulation and post-transcriptional regulation. These results provide a novel mechanism of action for triptolide which may be important in the treatment of lung cancer.

  14. MiR-214 inhibits cell growth in hepatocellular carcinoma through suppression of {beta}-catenin

    SciTech Connect

    Wang, Xiaojun; Chen, Ji; Li, Feng; Lin, Yanting; Zhang, Xiaoping; Lv, Zhongwei; Jiang, Jiaji

    2012-11-30

    Highlights: Black-Right-Pointing-Pointer miR-214 is frequently downregulated in human HCC cell lines and tissues. Black-Right-Pointing-Pointer miR-214 overexpression inhibits HCC cell growth in vitro and in vivo. Black-Right-Pointing-Pointer miR-214 directly targets {beta}-catenin 3 Prime -UTR in HCC cells. Black-Right-Pointing-Pointer miR-214 regulates {beta}-catenin downstream signaling molecules. -- Abstract: Mounting evidence has shown that microRNAs (miRNAs) are implicated in carcinogenesis and can function as oncogenes or tumor suppressor genes in human cancers. Recent profile studies of miRNA expression have documented a deregulation of miRNA (miR-214) in hepatocellular carcinoma (HCC). However, its potential functions and underlying mechanisms in hepatocarcinogenesis remain largely unknown. Here, we confirmed that miR-214 is significantly downregulated in HCC cells and specimens. Ectopic overexpression of miR-214 inhibited proliferation of HCC cells in vitro and tumorigenicity in vivo. Further studies revealed that miR-214 could directly target the 3 Prime -untranslated region (3 Prime -UTR) of {beta}-catenin mRNA and suppress its protein expression. Similar to the restoring miR-214 expression, {beta}-catenin downregulation inhibited cell growth, whereas restoring the {beta}-catenin expression abolished the function of miR-214. Moreover, miR-214-mediated reduction of {beta}-catenin resulted in suppression of several downstream genes including c-Myc, cyclinD1, TCF-1, and LEF-1. These findings indicate that miR-214 serves as tumor suppressor and plays substantial roles in inhibiting the tumorigenesis of HCC through suppression of {beta}-catenin. Given these, miR-214 may serve as a useful prognostic or therapeutic target for treatment of HCC.

  15. Reversed-polarity regions

    NASA Technical Reports Server (NTRS)

    Tang, F.

    1982-01-01

    It is found by a statistical study of 58 reversed-polarity regions (RPRs) covering the 11-year period 1969-1979 that RPRs (1) have a lifespan comparable to normal active regions, (2) do not show a tendency to rotate toward a more normal alignment, and (3) have stable configurations that do not suggest stress due to their anomalous magnetic alignment. As in normal regions, RPR magnetic complexity is found to be the primary factor in flare productivity. Weak-field RPRs produce no flares, and regions with complex spots produce more flares than regions with non-complex spots by a factor of five. The main difference between RPRs and normal regions lies in complex spot frequency, with less that 17% of normal active regions having such spots and fewer than 1.8% having long-lived complex ones, while 41% of RPRs have complex spots and 24% have long-lived complex spots.

  16. Genetic and physical mapping of 2q35 in the region of NRAMP and IL8R genes: Identification of a polymorphic repeat in exon 2 of NRAMP

    SciTech Connect

    White, J.K.; Shaw, M.A.; Barton, C.H.

    1994-11-15

    Recent interest has focused on the region of conserved synteny between mouse chromosome 1 and human 2q33-q37, particularly over the region encoding the murine macrophage resistance gene Ity/Lsh/Bcg (candidate Nramp) and members of the Il8r interleukin-8 (IL8) receptor gene cluster. In this paper, identification of a restriction fragment length polymorphism in the Il8RB gene in 35 pedigrees previously typed for markers in the 2q33-37 interval provided evidence (lod scores > 3) for linkage between Il8RB and the 2q34-135 markers FN1, TNP1, VIL1, and DES. Physical mapping, using yeast artificial chromosomes isolated with VIL1, confirmed that IL8RA, IL8RB and the IL8RB pseudogene map within the NRAMP-VIL1 interval, with the physical distance (155 kb) from 5{prime} LSH to 3{prime} VIL1 representing {approx}3-fold that observed in the mouse. Partial sequencing of NRAMP confirmed the presence of the N-terminal proline/serine-rich putative SH3 binding domain in exon 2 of the human gene. Further analysis of Brazilian leprosy and visceral leishmaniasis pedigrees identified a rare second allele varying in a 9-nucleotide repeat motif of the exon 2 sequence but segregating independently of the disease phenotype. 38 refs., 4 figs., 3 tabs.

  17. Isolation and characterization of two overlapping cosmid clones from the 4q35 region, near the facioscapulohumeral muscular dystrophy locus

    SciTech Connect

    Deidda, G.; Grisanti, P.; Vigneti, E.

    1994-09-01

    The gene for facioscapulohumeral muscular dystrophy (FSHD) has been localized by linkage analysis to the 4q35 region. The most telomeric p13E-11 prove has been shown to detect 4q35 DNA rearrangements in both sporadic and familial cases of the disease. With the aim of constructing a detailed physical map of the 4q35 region and searching for the mutant gene, we used p13E-11 probe to isolate cosmid clones from a human genomic library in a pCos-EMBL 2 vector. Two positive clones were isolated, clones 3 and 5, which partially overlap and carry human genomic inserts of 42 and 45 kb, respectively. The cosmids share a common region containing the p13E-11 region and a stretch of KpnI units consisting of 3.2 kb tandemly repeated sequences (about 10). The restriction maps were constructed using the following enzymes: Bam HI, BgIII, Eco RI, EcoRV, KpnI and Sfi I. Clone 3 extends 4 kb upstream of C5 and stops within the Kpn repeats. Clone 5 extends 4 kb downstream from the Kpn repeats and it presents an additional EcoRI site. Clone 5 contains a stretch of Kpn sequences of nearly 32 kb, corresponding to 10 Kpn repeats; clone 3 contains a stretch of 29 kb corresponding to 9 Kpn repeats, as determined by PFGE analysis of partial digestion of the clones. Clone 5 seems to contain the entire Eco RI region prone to rearrangements in FSHD patients. From clone 5 several subclones were obtained, from the Kpn region and from the region spanning from the last Kpn repeat to the cloning site. No single copy sequences were detected. Subclones from the 3{prime} end region contain beta-satellite or Sau3A-like sequences. In situ hybridization with the whole C5 cosmid shows hybridization signals at the tip of chromosome 4 (4q35) and chromosome 10 (10q26), in the pericentromeric region of chromosome 1 (1q12) and in the p12 region of the acrocentric chromosomes (chr. 21, 22, 13, 14, 15).

  18. Characterization of the Promoter Regions of Two Sheep Keratin-Associated Protein Genes for Hair Cortex-Specific Expression.

    PubMed

    Zhao, Zhichao; Liu, Guangbin; Li, Xinyun; Huang, Ji; Xiao, Yujing; Du, Xiaoyong; Yu, Mei

    2016-01-01

    The keratin-associated proteins (KAPs) are the structural proteins of hair fibers and are thought to play an important role in determining the physical properties of hair fibers. These proteins are activated in a striking sequential and spatial pattern in the keratinocytes of hair fibers. Thus, it is important to elucidate the mechanism that underlies the specific transcriptional activity of these genes. In this study, sheep KRTAP 3-3 and KRTAP11-1 genes were found to be highly expressed in wool follicles in a tissue-specific manner. Subsequently, the promoter regions of the two genes that contained the 5' flanking/5' untranslated regions and the coding regions were cloned. Using an in vivo transgenic approach, we found that the promoter regions from the two genes exhibited transcriptional activity in hair fibers. A much stronger and more uniformly expressed green fluorescent signal was observed in the KRTAP11-1-ZsGreen1 transgenic mice. In situ hybridization revealed the symmetrical expression of sheep KRTAP11-1 in the entire wool cortex. Consistently, immunohistochemical analysis demonstrated that the pattern of ZsGreen1 expression in the hair cortex of transgenic mice matches that of the endogenous KRTAP11-1 gene, indicating that the cloned promoter region contains elements that are sufficient to govern the wool cortex-specific transcription of KRTAP11-1. Furthermore, regulatory regions in the 5' upstream sequence of the sheep KRTAP11-1 gene that may regulate the observed hair keratinocyte specificity were identified using in vivo reporter assays.

  19. Characterization of the Promoter Regions of Two Sheep Keratin-Associated Protein Genes for Hair Cortex-Specific Expression

    PubMed Central

    Zhao, Zhichao; Liu, Guangbin; Li, Xinyun; Huang, Ji; Xiao, Yujing; Du, Xiaoyong; Yu, Mei

    2016-01-01

    The keratin-associated proteins (KAPs) are the structural proteins of hair fibers and are thought to play an important role in determining the physical properties of hair fibers. These proteins are activated in a striking sequential and spatial pattern in the keratinocytes of hair fibers. Thus, it is important to elucidate the mechanism that underlies the specific transcriptional activity of these genes. In this study, sheep KRTAP 3–3 and KRTAP11-1 genes were found to be highly expressed in wool follicles in a tissue-specific manner. Subsequently, the promoter regions of the two genes that contained the 5′ flanking/5′ untranslated regions and the coding regions were cloned. Using an in vivo transgenic approach, we found that the promoter regions from the two genes exhibited transcriptional activity in hair fibers. A much stronger and more uniformly expressed green fluorescent signal was observed in the KRTAP11-1-ZsGreen1 transgenic mice. In situ hybridization revealed the symmetrical expression of sheep KRTAP11-1 in the entire wool cortex. Consistently, immunohistochemical analysis demonstrated that the pattern of ZsGreen1 expression in the hair cortex of transgenic mice matches that of the endogenous KRTAP11-1 gene, indicating that the cloned promoter region contains elements that are sufficient to govern the wool cortex-specific transcription of KRTAP11-1. Furthermore, regulatory regions in the 5′ upstream sequence of the sheep KRTAP11-1 gene that may regulate the observed hair keratinocyte specificity were identified using in vivo reporter assays. PMID:27100288

  20. Transition Region Models

    NASA Astrophysics Data System (ADS)

    Hansteen, V.; Murdin, P.

    2000-11-01

    The SOLAR TRANSITION REGION comprises the PLASMA between the CHROMOSPHERE and the CORONA. In both of these regions the temperature is fairly uniform. The transition region, by contrast, is believed to be characterized by a very steep temperature rise from a chromospheric temperature of slightly less than 104 K to coronal temperatures on the order of 106 K. The goal of modeling the transition regi...

  1. Structure of the coding region and mRNA variants of the apyrase gene from pea (Pisum sativum)

    NASA Technical Reports Server (NTRS)

    Shibata, K.; Abe, S.; Davies, E.

    2001-01-01

    Partial amino acid sequences of a 49 kDa apyrase (ATP diphosphohydrolase, EC 3.6.1.5) from the cytoskeletal fraction of etiolated pea stems were used to derive oligonucleotide DNA primers to generate a cDNA fragment of pea apyrase mRNA by RT-PCR and these primers were used to screen a pea stem cDNA library. Two almost identical cDNAs differing in just 6 nucleotides within the coding regions were found, and these cDNA sequences were used to clone genomic fragments by PCR. Two nearly identical gene fragments containing 8 exons and 7 introns were obtained. One of them (H-type) encoded the mRNA sequence described by Hsieh et al. (1996) (DDBJ/EMBL/GenBank Z32743), while the other (S-type) differed by the same 6 nucleotides as the mRNAs, suggesting that these genes may be alleles. The six nucleotide differences between these two alleles were found solely in the first exon, and these mutation sites had two types of consensus sequences. These mRNAs were found with varying lengths of 3' untranslated regions (3'-UTR). There are some similarities between the 3'-UTR of these mRNAs and those of actin and actin binding proteins in plants. The putative roles of the 3'-UTR and alternative polyadenylation sites are discussed in relation to their possible role in targeting the mRNAs to different subcellular compartments.

  2. Two bi-allelic single nucleotide polymorphisms within the promoter region of the horse tumour necrosis factor alpha gene.

    PubMed

    Matiasovic, J; Lukeszová, L; Horín, P

    2002-08-01

    Primers based on GenBank sequences within the 5' untranslated region (UTR) of the human and horse tumour necrosis factor alpha (TNF-alpha) genes were designed and used to amplify a 522-bp product. Sequencing of five clones derived from five independent PCRs obtained from three different animals of three different breeds (Old Kladruber, Akhal-Teke and Shetland Pony) revealed a high level of sequence identity to the TNF-alpha promoter regions of other species. The existing GenBank horse sequences were confirmed and extended upstream by 230 nucleotides. Based on the sequence obtained, a new horse-specific forward primer was designed to amplify a 213-bp PCR product, which was screened for polymorphism using single-strand conformation polymorphism (SSCP). Three allelic variants of the horse TNF-alpha gene were identified and sequenced (GenBank accession numbers ADF 349558-60). Two single nucleotide polymorphisms explained the existence of the three SSCP alleles detected: C/T and T/C single base pair substitutions at positions 137 and 147, respectively. Differences in allelic frequencies between Old Kladruber and Akhal-Teke breeds were observed.

  3. Deletions of a differentially methylated CpG island at SNRPN define a putative imprinting control region

    SciTech Connect

    Sutcliffe, J.S.,; Nakao, M.; Beaudet, A.L.

    1994-09-01

    Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are associated with paternal and maternal deficiencies, respectively, of gene expression within human chromosome 15q11-q13, and are caused by deletion, uniparental disomy, or other mutations. Four transcripts designated PAR-5, PAR-7, PAR-1 and PAR-4 were isolated and localized to a region within 300 kb telomeric to the gene encoding small nuclear ribonucleoprotein-associated polypeptide N (SNRPN). Analysis of the transcripts in cultured fibroblasts and lymphoblasts from deletion patients demonstrated that SNRPN, PAR-5 and PAR-1 are expressed exclusively from the paternal chromosome, defining an imprinted domain that spans at least 200 kb. All three imprinted transcripts were absent in cells from three PWS patients (one pair of sibs and one sporadic case) with small deletions that involve a differentially methylated CpG island containing a previously undescribed 5{prime} untranslated exon ({alpha}) of SNRPN. Methylation of the CpG island is specific for the maternal chromosome consistent with paternal expression of the imprinted domain. One deletion, which is benign when maternally transmitted, extends upstream <30 kb from the CpG island, and is associated with altered methylation centromeric to SNRPN, and loss of transcription telomeric to SNRPN, implying the presence of an imprinting control region around the CpG island containing exon {alpha}.

  4. Genetic diversity within 5′upstream region of Toll-like receptor 8 gene reveals differentiation of riverine and swamp buffaloes☆

    PubMed Central

    Dubey, P.K.; Goyal, S.; Kumari, N.; Mishra, S.K.; Arora, R.; Kataria, R.S.

    2013-01-01

    In this study the nucleotide diversity in the 5′untranslated region (UTR) of TLR8 gene in riverine as well as swamp buffaloes has been described. Analysis of the 5′UTR of TLR8 gene showed presence of two SNPs in this region, g.-139G>T and g.-128A>G. A PCR–RFLP assay designed for genotyping of g.-139G>T SNP across 667 samples from 2 buffalo populations revealed a striking difference in allele frequency distribution across the swamp and riverine buffaloes. The frequency of T allele was higher in swamp buffalo as compared to riverine buffalo, ranging from 0.71 to 1. The G allele on the other hand exhibited a higher frequency across all the Indian riverine buffalo breeds/populations. The principal component analysis revealed separate clusters for the riverine and swamp buffaloes, as expected; however, the riverine type Assamese buffalo population of eastern India formed a distinct cluster. Since most of the buffalo populations in the eastern region are swamp type, this demarcation may be related to the difference in immune response in riverine and swamp buffaloes. These preliminary results indicate that the genetic variation observed in 5′upstream region of TLR8 gene, which differentiates swamp and riverine buffalo, needs to be further explored for association with disease susceptibility in buffalo, an important dairy and meat animal of Southeast Asia. PMID:25606371

  5. The Regional Educational Laboratories.

    ERIC Educational Resources Information Center

    Office of Educational Research and Improvement (ED), Washington, DC. Office of Reform Assistance and Dissemination.

    The Regional Educational Laboratory Program is the U.S. Department of Education's largest research and development investment designed to help educators, policymakers, and communities improve schools and help all students attain their potential. The network of 10 regional laboratories works to ensure that those involved in education improvement at…

  6. Ad Hoc Rural Regionalism

    ERIC Educational Resources Information Center

    Hamin, Elisabeth M.; Marcucci, Daniel J.

    2008-01-01

    A new regionalism has been much documented and researched for metropolitan areas; this article documents that there is a new rural regionalism as well. In the United States, these groups appear most likely to emerge in areas that are challenged by outcomes characterizing globalization's effects on the rural condition: namely, exurban or…

  7. REGIONAL EMAP PROPOSALS

    EPA Science Inventory

    The US EPA's Environmental Assessment and Monitoring Program (EMAP) annually funds regional EMAP (REMAP) projects through each of the regions to support the improvement of monitoring activities by the states. The last call for proposals emphasized the need to support biological m...

  8. MC-6 Casius Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-6 quadrangle, Casius region of Mars. Except for the highly dissected southwestern part, which contains faults, mesas, and buttes of Nilosyrtis Mensae, the Casius region is dominated by light-colored and dark, relatively smooth plains. Latitude range 30 to 65 degrees, longitude range -120 to -60 degrees.

  9. Regions and Western Europe.

    ERIC Educational Resources Information Center

    Brunt, Barry M.

    1995-01-01

    Maintains that regional geography is undergoing important changes in its method of study to achieve a greater degree of relevancy in the context of a global system. Presents Western Europe as a case study to reflect this new approach. Includes 11 maps illustrating 6 generalizations applied to regional patterns. (CFR)

  10. Reversed-polarity regions

    NASA Technical Reports Server (NTRS)

    Tang, F.

    1980-01-01

    The 58 RPRS studied have a lifespan comparable to normal active regions and have no tendency to rotate toward a more normal alignment. They seem to have stable configurations with no apparent evidence suggesting stress due to their anomalous magnetic alignment. Magnetic complexity in RPRs is the key to flare productivity just as it is in normal regions - weak field RPRs produced no flares and regions with complex spots produced more flares than regions with noncomplex spots by a factor of 5. The RPRs however, differ from normal regions in the frequency of having complex spots, particularly the long lived complex spots, in them. Less than 17 percent of normal ARs have complex spots; less than 1.8 percent have long lived complex spots. In contrast, 41 percent of RPRs have complex spots and 24 percent have long lived complex spots.

  11. Identification of the first gene (FRG1) from the FSHD region on human chromosome 4q35.

    PubMed

    van Deutekom, J C; Lemmers, R J; Grewal, P K; van Geel, M; Romberg, S; Dauwerse, H G; Wright, T J; Padberg, G W; Hofker, M H; Hewitt, J E; Frants, R R

    1996-05-01

    Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant, neuromuscular disorder characterized by progressive weakness of muscles in the face, shoulder and upper arm. Deletion of integral copies of a 3.3 kb repeated unit from the subtelomeric region on chromosome 4q35 has been shown to be associated with FSHD. These repeated units which are apparently not transcribed, map very close to the 4q telomere and belong to a 3.3 kb repeat family dispersed over heterochromatic regions of the genome. Hence, position effect variegation (PEV), inducing allele-specific transcriptional repression of a gene located more centromeric, has been postulated as the underlying genetic mechanism of FSHD. This hypothesis has directed the search for the FSHD gene to the region centromeric to the repeated units. A CpG island was identified and found to be associated with the 5' untranslated region of a novel human gene, FRG1 (FSHD Region Gene 1). This evolutionary conserved gene is located about 100 kb proximal to the repeated units and belongs to a multigene family with FRG1 related sequences on multiple chromosomes. The mature chromosome 4 FRG1 transcript is 1042 bp in length and contains nine exons which encode a putative protein of 258 amino acid residues. Transcription of FRG1 was detected in several human tissues including placenta, lymphocytes, brain and muscle. To investigate a possible PEV mechanism, allele-specific FRG1 steady-state transcript levels were determined using RNA-based single-strand conformation polymorphism (SSCP) analysis. A polymorphic fragment contained within the first exon of FRG1 was amplified from reverse transcribed RNA from lymphocytes and muscle biopsies of patients and controls. No evidence for PEV mediated repression of allelic transcription was obtained in these tissues. However, detection of PEV in FSHD patients may require analysis of more specific cell types at particular developmental stages.

  12. Structural and mutational analysis of a conserved gene (DGSI) from the minimal DiGeorge syndrome critical region.

    PubMed

    Gong, W; Emanuel, B S; Galili, N; Kim, D H; Roe, B; Driscoll, D A; Budarf, M L

    1997-02-01

    The majority of patients with DiGeorge syndrome (DGS), velocardiofacial syndrome (VCFS), conotruncal anomaly face syndrome (CTAFS) and some individuals with familial or sporadic conotruncal cardiac defects have hemizygous deletions of chromosome 22. Most patients with these disorders share a common large deletion, spanning > 1.5 Mb within 22q11.21-q11.23. Recently, the smallest region of deletion overlap has been narrowed to a 250 kb area, the minimal DGS critical region (MDGCR), which includes the locus D22S75 (N25). We have isolated and characterized a novel, highly conserved gene, DGSI, within the MDGCR. DGSI has 10 exons and nine introns encompassing 1702 bp of cDNA sequence and 11 kb of genomic DNA. The encoded protein has 476 amino acids with a predicted mol. wt of 52.6 kDa. The intron-exon boundaries have been analyzed and conform to the consensus GT/AG motif. The corresponding murine Dgsi has been isolated and localized to proximal mouse chromosome 16. The mouse gene contains the same number of exons and introns, and the predicted protein has 479 amino acids with 93.2% identity to that of the human DGSI gene. By database searching, both genes have significant homology to a Caenorhabditis elegans hypothetical protein, F42H10.7. Further, mutation analysis has been performed in 16 patients, who have no detectable 22q11.2 deletion and some of the characteristic clinical features of DGS/VCFS. We have detected eight sequence variants in DGSI. These occurred in the 5'-untranslated region, the coding region and the intronic regions adjacent to the intron-exon boundaries of the gene. Seven of the eight variants were also present in normal controls or unaffected family members, suggesting they may not be of etiologic significance.

  13. A 1. 8-Mb YAC contig in Xp11. 23: Identification of Cpg islands and physical mapping of CA repeats in a region of high gene density

    SciTech Connect

    Coleman, M.P.; Nemeth, A.H.; Campbell, L.; Raut, C.P.; Davies, K.E. ); Weissenbach, J. )

    1994-05-15

    The genes ARAF1, SYN1, TIMP, and PFC are clustered within 70 kb of one another, and, as reported in the accompanying paper, at least four more genes map within 400 kb: a cluster of Krueppel-type zinc finger genes (including ZNF21, ZNF41, and ZNF81) and ELK-1, a member of the ets oncogene superfamily. This gene-rich region is of particular interest because of the large number of disease genes mapping to Xp11.23: At least three eye diseases (retinitis pigmentosa type 2, congenital stationary night blindness CSNB1, and Aland Island eye disease), Wiskott-Aldrich syndrome, X-linked nephrolithiasis, and a translocation breakpoint associated with synovial sarcoma. The authors have constructed a 1.8-Mb YAC contig in this region, confirming the link between TIMP and OATL1 reported by Knight et al. (1994) and extending the map in the distal direction. To investigate the likelihood that more genes are located within this region, they have carried out detailed mapping of rare-cutter restriction sites in these YACs and identified seven CpG islands. At least six of these islands are located over 50 kb from any known gene locations, suggesting that the region contains at least this many as yet unidentified genes. They have also mapped the physical locations of six highly polymorphic CA repeats within the contig, thus integrating the physical, genetic, and transcriptional maps of the region and facilitating the mapping and identification of disease genes. Together with the report by Knight et al., these data indicate the following order of loci: Xpter-DXS1264-DXS1055-DXS1003-DXS1146-DXS1266-(ZNF41, ARAF1)-SYN1 CA repeat-SYN1 (3[prime] end)-TIMP-SYN1 (5[prime] end)-PFC CA repeat-PFC-(DXS426, ELK1)-(DXS1265, ZNF81)-ZNF21-DXS1267-OATL1-Xcen. 40 refs., 4 figs., 3 tabs.

  14. Regional ocean data assimilation.

    PubMed

    Edwards, Christopher A; Moore, Andrew M; Hoteit, Ibrahim; Cornuelle, Bruce D

    2015-01-01

    This article reviews the past 15 years of developments in regional ocean data assimilation. A variety of scientific, management, and safety-related objectives motivate marine scientists to characterize many ocean environments, including coastal regions. As in weather prediction, the accurate representation of physical, chemical, and/or biological properties in the ocean is challenging. Models and observations alone provide imperfect representations of the ocean state, but together they can offer improved estimates. Variational and sequential methods are among the most widely used in regional ocean systems, and there have been exciting recent advances in ensemble and four-dimensional variational approaches. These techniques are increasingly being tested and adapted for biogeochemical applications.

  15. Drycleaner Database - Region 7

    EPA Pesticide Factsheets

    THIS DATA ASSET NO LONGER ACTIVE: This is metadata documentation for the Region 7 Drycleaner Database (R7DryClnDB) which tracks all Region7 drycleaners who notify Region 7 subject to Maximum Achievable Control Technologiy (MACT) standards. The Air and Waste Management Division is the primary managing entity for this database. This work falls under objectives for EPA's 2003-2008 Strategic Plan (Goal 4) for Healthy Communities & Ecosystems, which are to reduce chemical and/or pesticide risks at facilities.

  16. Upper Extremity Regional Anesthesia

    PubMed Central

    Neal, Joseph M.; Gerancher, J.C.; Hebl, James R.; Ilfeld, Brian M.; McCartney, Colin J.L.; Franco, Carlo D.; Hogan, Quinn H.

    2009-01-01

    Brachial plexus blockade is the cornerstone of the peripheral nerve regional anesthesia practice of most anesthesiologists. As part of the American Society of Regional Anesthesia and Pain Medicine’s commitment to providing intensive evidence-based education related to regional anesthesia and analgesia, this article is a complete update of our 2002 comprehensive review of upper extremity anesthesia. The text of the review focuses on (1) pertinent anatomy, (2) approaches to the brachial plexus and techniques that optimize block quality, (4) local anesthetic and adjuvant pharmacology, (5) complications, (6) perioperative issues, and (6) challenges for future research. PMID:19282714

  17. Structural analysis of the 5 prime flanking region of the. beta. -globin gene in African sickle cell anemia patients: Further evidence for three origins of the sickle cell mutation in Africa

    SciTech Connect

    Chebloune, Y.; Pagnier, J.; Trabuchet, G.; Faure, C.; Verdier, G.; Labie, D.; Nigon, V. )

    1988-06-01

    Haplotype analysis of the {beta}-globin gene cluster shows two regions of DNA characterized by nonrandom association of restriction site polymorphisms. These regions are separated by a variable segment containing the repeated sequences (ATTTT){sub n} and (AT){sub x}T{sub y}, which might be involved in recombinational events. Studies of haplotypes linked to the sickle cell gene in Africa provide strong argument for three origins of the mutation: Benin, Senegal, and the Central African Republic. The structure of the variable segment in the three African populations was studied by S1 nuclease mapping of genomic DNA, which allows a comparison of several samples. A 1080-base-pair DNA segment was sequenced for one sample from each population. S1 nuclease mapping confirmed the homogeneity of each population with regard to both (ATTTT){sub n} and (AT){sub x}T{sub y} repeats. The authors found three additional structures for (AT){sub x}T{sub y} correlating with the geographic origin of the patients. Ten other nucleotide positions, 5{prime} and 3{prime} to the (AT){sub x}T{sub y} copies, were found to be variable when compared to homologous sequences from human and monkey DNAs. These results allow us to propose an evolutionary scheme for the polymorphisms in the 5{prime} flanking region of the {beta}-globin gene. The results strongly support the hypothesis of three origins for the sickle mutation in Africa.

  18. Regional Instrumentation Centers.

    ERIC Educational Resources Information Center

    Cromie, William J.

    1980-01-01

    Focuses on the activities of regional instrumentation centers that utilize the state-of-the-art instruments and methodology in basic scientific research. The emphasis is on the centers involved in mass spectroscopy, magnetic resonance spectroscopy, lasers, and accelerators. (SA)

  19. Mercury's South Polar Region

    NASA Video Gallery

    This animation shows 89 wide-angle camera (WAC) images of Mercury’s south polar region acquired by the Mercury Dual Imaging System (MDIS) over one complete Mercury solar day (176 Earth days). Thi...

  20. Regional Hearing Clerk

    EPA Pesticide Factsheets

    The Regional Hearing Clerk receives filings for proceedings under the Consolidated Rules of Practice Governing the Administrative Assessment of Civil Penalties and the Revocation/Termination or Suspension of Permits, 40 Code of Federal Regulations Part 22

  1. Complex Regional Pain Syndrome

    MedlinePlus

    Complex regional pain syndrome (CRPS) is a chronic pain condition. It causes intense pain, usually in the arms, hands, legs, or feet. ... in skin temperature, color, or texture Intense burning pain Extreme skin sensitivity Swelling and stiffness in affected ...

  2. Regional Screening Levels (RSLs)

    EPA Pesticide Factsheets

    Regional Screening Level RSL Home Page introduces risk assessors to Chemical Risk Assessment preliminary remediation goals PRG risk based concentration RBC and risk calculations for the assessment of human Health.

  3. Complex Regional Pain Syndrome

    MedlinePlus

    ... regional pain syndrome is an uncommon form of chronic pain that usually affects an arm or a leg. ... exercises may be. Transcutaneous electrical nerve stimulation (TENS) . Chronic pain is sometimes eased by applying electrical impulses to ...

  4. Regional Health Information Systems

    PubMed Central

    Fuller, Sherrilynne

    1997-01-01

    Abstract In general, there is agreement that robust integrated information systems are the foundation for building successful regional health care delivery systems. Integrated Advanced Information Management System (IAIMS) institutions that, over the years, have developed strategies for creating cohesive institutional information systems and services are finding that IAIMS strategies work well in the even more complex regional environment. The key elements of IAIMS planning are described and lessons learned are discussed in the context of regional health information systems developed. The challenges of aligning the various information agencies and agendas in support of a regional health information system are complex ; however, the potential rewards for health care in quality, efficacy, and cost savings are enormous. PMID:9067887

  5. Transcription-Associated R-Loop Formation across the Human FMR1 CGG-Repeat Region

    PubMed Central

    Loomis, Erick W.; Sanz, Lionel A.; Chédin, Frédéric; Hagerman, Paul J.

    2014-01-01

    Expansion of a trinucleotide (CGG) repeat element within the 5′ untranslated region (5′UTR) of the human FMR1 gene is responsible for a number of heritable disorders operating through distinct pathogenic mechanisms: gene silencing for fragile X syndrome (>200 CGG) and RNA toxic gain-of-function for FXTAS (∼55–200 CGG). Existing models have focused almost exclusively on post-transcriptional mechanisms, but co-transcriptional processes could also contribute to the molecular dysfunction of FMR1. We have observed that transcription through the GC-rich FMR1 5′UTR region favors R-loop formation, with the nascent (G-rich) RNA forming a stable RNA:DNA hybrid with the template DNA strand, thereby displacing the non-template DNA strand. Using DNA:RNA (hybrid) immunoprecipitation (DRIP) of genomic DNA from cultured human dermal fibroblasts with both normal (∼30 CGG repeats) and premutation (55

  6. Regionalism and Secession

    DTIC Science & Technology

    2015-03-01

    strong independence movements in Spain, but not in Gennany? How have state fonna.tion and the relationship betv.re.en the center and the region...affected independence movements ? This thesis provides a historical analysis of Bavarian and Catalan regionalism and argues that autonomy mitigates...Catalonians struggle actively and adamantly for autonomy. Why are there strong independence movements in Spain, but not in Germany? How have state

  7. MC-16 Memnonia Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-16 quadrangle, Memnonia region of Mars. Heavily cratered highlands in the southern two-thirds are cut in the northeastern part by a large outflow channel, Mangala Vallis. The highlands are bounded to the north by undulating wind-eroded deposits and to the east by lava flows of the Tharsis region. Latitude range -30 to 0 degrees, longitude range 135 to 180 degrees.

  8. On regional geomagnetic charts

    USGS Publications Warehouse

    Alldredge, L.R.

    1987-01-01

    When regional geomagnetic charts for areas roughly the size of the US were compiled by hand, some large local anomalies were displayed in the isomagnetic lines. Since the late 1960s, when the compilation of charts using computers and mathematical models was started, most of the details available in the hand drawn regional charts have been lost. One exception to this is the Canadian magnetic declination chart for 1980. This chart was constructed using a 180 degrees spherical harmonic model. -from Author

  9. Modeling the transition region

    NASA Astrophysics Data System (ADS)

    Singer, Bart A.

    1994-04-01

    The calculation of engineering flows undergoing laminar-turbulent transition presents special problems. Mean-flow quantities obey neither the fully laminar nor the fully turbulent correlations. In addition, local maxima in skin friction, wall temperature, and heat transfer often occur near the end of the transition region. Traditionally, modeling this region has been important for the design of turbine blades, where the transition region is long in relation to the chord length of the blade. More recently, the need for better transition-region models has been recognized by designers of hypersonic vehicles where the high Mach number, the low Reynolds number, and the low-disturbance flight environment emphasize the importance of the transition region. Needless to say, a model that might work well for the transitional flows typically found in gas turbines will not necessarily work well for the external surface of a hypersonic vehicle. In Section 2 of this report, some of the important flow features that control the transition region will be discussed. In Section 3, different approaches to the modeling problem will be summarized and cataloged. Fully turbulent flow models will be discussed in detail in Section 4; models specifically designed for transitional flow, in Section 5; and the evaluation of models, in Section 6.

  10. The near-naked hairless (Hr(N)) mutation disrupts hair formation but is not due to a mutation in the Hairless coding region.

    PubMed

    Liu, Yutao; Das, Suchita; Olszewski, Robert E; Carpenter, Donald A; Culiat, Cymbeline T; Sundberg, John P; Soteropoulos, Patricia; Liu, Xiaochen; Doktycz, Mitchel J; Michaud, Edward J; Voy, Brynn H

    2007-07-01

    Near-naked hairless (Hr(N)) is a semi-dominant, spontaneous mutation that was suggested by allelism testing to be allelic with mouse Hairless (Hr). Hr(N) mice differ from other Hr mutants in that hair loss appears as the postnatal coat begins to emerge, rather than as an inability to regrow hair after the first catagen and that the mutation displays semi-dominant inheritance. We sequenced the Hr cDNA in Hr(N)/Hr(N) mice and characterized the pathological and molecular phenotypes to identify the basis for hair loss in this model. Hr(N)/Hr(N) mice exhibit dystrophic hairs that are unable to emerge consistently from the hair follicle, whereas Hr(N)/+ mice display a sparse coat of hair and a milder degree of follicular dystrophy than their homozygous littermates. DNA microarray analysis of cutaneous gene expression demonstrates that numerous genes are downregulated in Hr(N)/Hr(N) mice, primarily genes important for hair structure. By contrast, Hr expression is significantly increased. Sequencing the Hr-coding region, intron-exon boundaries, 5'- and 3'-untranslated region, and immediate upstream region did not reveal the underlying mutation. Therefore, Hr(N) does not appear to be an allele of Hr but may result from a mutation in a closely linked gene or from a regulatory mutation in Hr.

  11. The vasa regulatory region mediates germline expression and maternal transmission of proteins in the malaria mosquito Anopheles gambiae: a versatile tool for genetic control strategies

    PubMed Central

    Papathanos, Philippos A; Windbichler, Nikolai; Menichelli, Miriam; Burt, Austin; Crisanti, Andrea

    2009-01-01

    Background Germline specific promoters are an essential component of potential vector control strategies which function by genetic drive, however suitable promoters are not currently available for the main human malaria vector Anopheles gambiae. Results We have identified the Anopheles gambiae vasa-like gene and found its expression to be specifically localized to both the male and female gonads in adult mosquitoes. We have functionally characterised using transgenic reporter lines the regulatory regions required for driving transgene expression in a pattern mirroring that of the endogenous vasa locus. Two reporter constructs indicate the existence of distinct vasa regulatory elements within the 5' untranslated regions responsible not only for the spatial and temporal but also for the sex specific germline expression. vasa driven eGFP expression in the ovary of heterozygous mosquitoes resulted in the progressive accumulation of maternal protein and transcript in developing oocytes that were then detectable in all embryos and neonatal larvae. Conclusion We have characterized the vasa regulatory regions that are not only suited to drive transgenes in the early germline of both sexes but could also be utilized to manipulate the zygotic genome of developing embryos via maternal deposition of active molecules. We have used computational models to show that a homing endonuclease-based gene drive system can function in the presence of maternal deposition and describe a novel non-invasive control strategy based on early vasa driven homing endonuclease expression. PMID:19573226

  12. Human gene encoding prostacyclin synthase (PTGIS): Genomic organization, chromosomal localization, and promoter activity

    SciTech Connect

    Yokoyama, Chieko; Yabuki, Tomoko; Inoue, Hiroyasu

    1996-09-01

    The prostacyclin synthase gene isolated from human genomic libraries (PTGIS) consists of 10 exons spanning approximately 60 kb. All the splice donor and acceptor sites conform to the GT/AG rule. Genomic Southern blot and fluorescence in situ hybridization analyses revealed that the human prostacyclin synthase gene is present as a single copy per haploid genome and is localized on chromosome 20q13.11-q13.13. The 1.5-kb sequence of the 5{prime} of the translational initiation site contained both GC-rich and pyrimidine-rich regions and consensus sequences of the transcription factor recognition sites such as Sp1, AP-2, the interferon-{gamma} response element, GATA, NF-{kappa}B, the CACCC box, and the glucocorticoid response element. The core binding sequence (GAGACC) of the shear stress responsive element was also found in the 5{prime}-flanking region of the gene. The major product of the primer extension analysis suggested that the transcription of the gene started from the positions around 49 bp upstream of the translational initiation codon. Transient transfection experiments using human aortic and bovine arterial endothelial cells demonstrated that the GC-rich region (positions -145 to -10) possessed a significant promoter activity. The 6-kb downstream sequence of the translational termination codon contained multiple polyadenylation signals, Alu repeat sequences, and the consensus sequence of the primate-repetitive DNA element, MER1. Two sizes of the prostacyclin synthase mRNAs (approximately 6 and 3.3 kb) were detected with the human aorta and lung. RNA blot hybridization analysis using the 3{prime}-untranslated region as probe indicated that the sizes of the 3{prime}-flanking regions were different in the major 6-kb and minor 3.3-kb mRNAs. 54 refs., 7 figs.

  13. Northeast Regional Biomass Program

    SciTech Connect

    Lusk, P.D.

    1992-12-01

    The Northeast Regional Biomass Program has been in operation for a period of nine years. During this time, state managed programs and technical programs have been conducted covering a wide range of activities primarily aim at the use and applications of wood as a fuel. These activities include: assessments of available biomass resources; surveys to determine what industries, businesses, institutions, and utility companies use wood and wood waste for fuel; and workshops, seminars, and demonstrations to provide technical assistance. In the Northeast, an estimated 6.2 million tons of wood are used in the commercial and industrial sector, where 12.5 million cords are used for residential heating annually. Of this useage, 1504.7 mw of power has been generated from biomass. The use of wood energy products has had substantial employment and income benefits in the region. Although wood and woodwaste have received primary emphasis in the regional program, the use of municipal solid waste has received increased emphasis as an energy source. The energy contribution of biomass will increase as potentia users become more familiar with existing feedstocks, technologies, and applications. The Northeast Regional Biomass Program is designed to support region-specific to overcome near-term barriers to biomass energy use.

  14. Ultrasound in regional anaesthesia.

    PubMed

    Griffin, J; Nicholls, B

    2010-04-01

    Ultrasound guidance is rapidly becoming the gold standard for regional anaesthesia. There is an ever growing weight of evidence, matched with improving technology, to show that the use of ultrasound has significant benefits over conventional techniques, such as nerve stimulation and loss of resistance. The improved safety and efficacy that ultrasound brings to regional anaesthesia will help promote its use and realise the benefits that regional anaesthesia has over general anaesthesia, such as decreased morbidity and mortality, superior postoperative analgesia, cost-effectiveness, decreased postoperative complications and an improved postoperative course. In this review we consider the evidence behind the improved safety and efficacy of ultrasound-guided regional anaesthesia, before discussing its use in pain medicine, paediatrics and in the facilitation of neuraxial blockade. The Achilles' heel of ultrasound-guided regional anaesthesia is that anaesthetists are far more familiar with providing general anaesthesia, which in most cases requires skills that are achieved faster and more reliably. To this ends we go on to provide practical advice on ultrasound-guided techniques and the introduction of ultrasound into a department.

  15. The role of the 3' region of mammalian gonadotropin β subunit gene in the luteinizing hormone to chorionic gonadotropin evolution.

    PubMed

    Gabay, Reut; Rozen, Shelly; Samokovlisky, Albena; Amor, Yehudit; Rosenfeld, Rakefet; Kohen, Fortune; Amsterdam, Abraham; Berger, Peter; Ben-Menahem, David

    2014-02-15

    CGβ subunits comprise a unique carboxyl-terminal peptide (CTP) that has multiple O-linked glycans and extends serum half-life of the protein. It has evolved by incorporating a previously untranslated region of the LHβ gene into the reading frame. Although CTP-like sequences are encrypted in the LHβ genes of several mammals, the CGβ subunit developed only in primates and equids. To study this restriction in evolution, we examined whether the cryptic CTP decoded from the bovine LHβ gene (boCTP) possesses key characteristics of the human (h) CGβ-CTP. The boCTP does not impede several crucial aspects of hormone biosynthesis, but compared to the hCGβ-CTP, the stretch lacks O-glycans and determinants for circulatory survival. O-glycan deficiency and the associated incapacity to extend serum half-life is a major drawback of the boCTP. This may explain why LH did not evolve into CG in ruminants and consequently alternative mechanisms evolved to delay luteolysis early in gestation.

  16. Ground- and excited-state tautomerism in 2-(3{prime}-hydroxy-2{prime}-pyridyl)benzimidazole

    SciTech Connect

    Prieto, F.R.; Rodriguez, M.C.R.; Gonzalez, M.M.; Fernandez, M.A.R.

    1994-09-01

    Ground-state HPyBI is determined to have keto-enol equilibrium in water, and the enol form predominates in nonaqueous solutions. The keto form is the only excited form in all the solvents considered. Ultrafast intramolecular proton transfer creates the enol form from the keto form. 47 refs., 6 figs., 3 tabs.

  17. Regional anesthesia at home

    PubMed Central

    Cheng, Gloria S.; Choy, Lynna P.; Ilfeld, Brian M.

    2009-01-01

    Purpose of review To review the recently published peer-reviewed literature involving regional anesthesia and analgesia in patients at home. Recent findings The potential benefits and risks of regional anesthesia and analgesia at home are pertinent queries, and increased data regarding these topics are rapidly becoming available. Of particular interest is the use of continuous peripheral nerve blocks at home and their potential effect upon hospitalization duration and recovery profile. Summary Advantages of regional techniques include site-specific anesthesia and decreased postoperative opioid use. For shoulder surgeries, the interscalene block provides effective analgesia with minimal complications, whereas the impact and risks of intraarticular injections remain unclear. Perineural catheters are an analgesic option that offer improved pain relief among other benefits. They are now being used at home in both adult and pediatric populations. PMID:18660659

  18. NV PFA Regional Data

    SciTech Connect

    James Faulds

    2015-10-28

    This project focused on defining geothermal play fairways and development of a detailed geothermal potential map of a large transect across the Great Basin region (96,000 km2), with the primary objective of facilitating discovery of commercial-grade, blind geothermal fields (i.e. systems with no surface hot springs or fumaroles) and thereby accelerating geothermal development in this promising region. Data included in this submission consists of: structural settings (target areas, recency of faulting, slip and dilation potential, slip rates, quality), regional-scale strain rates, earthquake density and magnitude, gravity data, temperature at 3 km depth, permeability models, favorability models, degree of exploration and exploration opportunities, data from springs and wells, transmission lines and wilderness areas, and published maps and theses for the Nevada Play Fairway area.

  19. Noncoding regions of C. elegans mRNA undergo selective adenosine to inosine deamination and contain a small number of editing sites per transcript.

    PubMed

    Wheeler, Emily C; Washburn, Michael C; Major, Francois; Rusch, Douglas B; Hundley, Heather A

    2015-01-01

    ADARs (Adenosine deaminases that act on RNA) "edit" RNA by converting adenosines to inosines within double-stranded regions. The primary targets of ADARs are long duplexes present within noncoding regions of mRNAs, such as introns and 3' untranslated regions (UTRs). Because adenosine and inosine have different base-pairing properties, editing within these regions can alter splicing and recognition by small RNAs. However, despite numerous studies identifying multiple editing sites in these genomic regions, little is known about the extent to which editing sites co-occur on individual transcripts or the functional output of these combinatorial editing events. To begin to address these questions, we performed an ultra-deep sequencing analysis of 4 Caenorhabditis elegans 3' UTRs that are known ADAR targets. Synchronous editing events were determined for the long duplexes in vivo. Furthermore, the validity of each editing event was confirmed by sequencing the same regions of mRNA from worms that lack A-to-I editing. This analysis identified a large number of editing sites that can occur within each 3' UTR, but interestingly, each individual transcript contained only a small fraction of these A-to-I editing events. In addition, editing patterns were not random, indicating that an editing event can affect the efficiency of editing at subsequent adenosines. Furthermore, we identified specific sites that can be both positively and negatively correlated with additional sites leading to mutually exclusive editing patterns. These results suggest that editing in noncoding regions is selective and hyper-editing of cellular RNAs is rare.

  20. In situ optical sequencing and structure analysis of a trinucleotide repeat genome region by localization microscopy after specific COMBO-FISH nano-probing

    NASA Astrophysics Data System (ADS)

    Stuhlmüller, M.; Schwarz-Finsterle, J.; Fey, E.; Lux, J.; Bach, M.; Cremer, C.; Hinderhofer, K.; Hausmann, M.; Hildenbrand, G.

    2015-10-01

    Trinucleotide repeat expansions (like (CGG)n) of chromatin in the genome of cell nuclei can cause neurological disorders such as for example the Fragile-X syndrome. Until now the mechanisms are not clearly understood as to how these expansions develop during cell proliferation. Therefore in situ investigations of chromatin structures on the nanoscale are required to better understand supra-molecular mechanisms on the single cell level. By super-resolution localization microscopy (Spectral Position Determination Microscopy; SPDM) in combination with nano-probing using COMBO-FISH (COMBinatorial Oligonucleotide FISH), novel insights into the nano-architecture of the genome will become possible. The native spatial structure of trinucleotide repeat expansion genome regions was analysed and optical sequencing of repetitive units was performed within 3D-conserved nuclei using SPDM after COMBO-FISH. We analysed a (CGG)n-expansion region inside the 5' untranslated region of the FMR1 gene. The number of CGG repeats for a full mutation causing the Fragile-X syndrome was found and also verified by Southern blot. The FMR1 promotor region was similarly condensed like a centromeric region whereas the arrangement of the probes labelling the expansion region seemed to indicate a loop-like nano-structure. These results for the first time demonstrate that in situ chromatin structure measurements on the nanoscale are feasible. Due to further methodological progress it will become possible to estimate the state of trinucleotide repeat mutations in detail and to determine the associated chromatin strand structural changes on the single cell level. In general, the application of the described approach to any genome region will lead to new insights into genome nano-architecture and open new avenues for understanding mechanisms and their relevance in the development of heredity diseases.

  1. Climatic Concepts and Regions.

    ERIC Educational Resources Information Center

    Thomas, Paul F.

    Designed for students in grades 7 through 12, this teaching unit presents illustrative resource materials depicting concepts related to climate and geographic regions. Emphasis is on giving students an understanding of climatic elements and factors, not as isolated, disjointed entities, but as a dynamic interplay of forces having a very definite…

  2. Benchmarks: WICHE Region 2012

    ERIC Educational Resources Information Center

    Western Interstate Commission for Higher Education, 2013

    2013-01-01

    Benchmarks: WICHE Region 2012 presents information on the West's progress in improving access to, success in, and financing of higher education. The information is updated annually to monitor change over time and encourage its use as a tool for informed discussion in policy and education communities. To establish a general context for the…

  3. Regional cancer pain syndromes.

    PubMed

    Chang, Victor T; Janjan, Nora; Jain, Subash; Chau, Chi

    2006-12-01

    Cancer pain often presents in a body region. This review summarizes articles from 1999-2004 relevant to cancer pain syndromes in the head and neck, chest, back, abdomen, pelvis, and limbs. Although the evidence is limited, progress is being made in further development of the evidence base to support and guide current practice.

  4. Norway's Regional Colleges

    ERIC Educational Resources Information Center

    Kintzer, Frederick C.

    1974-01-01

    Created in 1969 as a 3-college system coordinated by the Regional College section of the Ministry of Education, the current 6 institutions represent Norway's attempt to extend equal opportunity and employment-oriented education to rural and remote areas. (Editor)

  5. REGIONAL CONFERENCE SUMMARIES, 1966.

    ERIC Educational Resources Information Center

    Bureau of Adult, Vocational, and Technical Education (DHEW/OE), Washington, DC. Div. of Vocational and Technical Education.

    AN AVERAGE OF 200 TEACHER EDUCATORS, STATE DIRECTORS, LAYMEN, AND REPRESENTATIVES OF VARIOUS AGENCIES ATTENDED EACH OF NINE REGIONAL CONFERENCES CONDUCTED THROUGHOUT THE UNITED STATES TO DISCUSS THE INFLUENCE OF SOCIAL AND ECONOMIC CHANGES AND PROBLEMS IN PLANNING AND CONDUCTING VOCATIONAL AND TECHNICAL EDUCATION PROGRAMS. MAJOR SPEECHES PRESENTED…

  6. Australia's Regional Youth Exodus.

    ERIC Educational Resources Information Center

    Gabriel, M.

    2002-01-01

    Examines media coverage of youth outmigration from Tasmania in the context of Australia's regional crisis. Focuses on how young people are constructed by others and positioned in others' visions of their rural home towns. Discusses two recurring narratives: strategies to keep youth at home, and preoccupation with the "best and brightest"…

  7. TVA and regional development

    SciTech Connect

    Not Available

    1983-01-01

    Although TVA is best known as a producer of electric power, its mission, in the words of Franklin D. Roosevelt, called for a program that touches and gives life to all forms of human concerns. This booklet provides general information about the scores of ways - beyond the electric power generation - in which the agency serves the region and nation.

  8. MC-24 Phaethontis Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-24 quadrangle, Phaethontis region of Mars. The Phaethontis quadrangle is dominated by heavily cratered highlands and low-lying areas forming relatively smooth plains. Latitude range -65 to -30 degrees, longitude range 120 to 180.

  9. MC-2 Diacria Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-2 quadrangle, Diacria region of Mars. The northern two-thirds is dominated by relatively smooth plains. The southeastern part is marked by aureole deposits of the largest known volcano in the solar system, Olympus Mons. Latitude range 30 to 65 degrees, longitude range 120 to 180.

  10. MC-14 Amenthes Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-14 quadrangle, Amenthes region of Mars. The southern part includes heavily cratered highlands. The northern part is dominated by relatively smooth plains of Elysium Planitia and the eastern half of the Isidis basin. Latitude range 0 to 30 degrees, longitude range -135 to -90 degrees.

  11. MC-3 Arcadia Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-3 quadrangle, Arcadia region of Mars. The southern part contains the large shield volcano, Alba Patera, and the highly faulted Tempe Terra province, which includes many small volcanoes. The northern part is dominated by relatively smooth plains. Latitude range 30 to 65 degrees, longitude range 60 to 120 degrees.

  12. Regionalism. Clip and Save.

    ERIC Educational Resources Information Center

    Hubbard, Guy

    2002-01-01

    Focuses on the art movement, called Regionalism, discussing the painters involved and describing the characteristics of the art movement. Provides a set of learning activities and background information on John Steuart Curry. Includes a discussion of Curry's painting, "Tornado Over Kansas," and a reproduction of the painting. (CMK)

  13. Regional Norms for English.

    ERIC Educational Resources Information Center

    Kachru, Braj B.

    The debate continues about regional norms for English usage around the world, although the discussion has become more realistic and less didactic. Educated non-native varieties are increasingly accepted, distinctions are being made between national and international language uses, and localized varieties are no longer considered as necessarily…

  14. Recipe for Regional Development.

    ERIC Educational Resources Information Center

    Baldwin, Fred D.

    1994-01-01

    The Ceramics Corridor has created new jobs in New York's Appalachian region by fostering ceramics research and product development by small private companies. Corridor business incubators offer tenants low overhead costs, fiber-optic connections to Alfred University's mainframe computer, rental of lab space, and use of equipment small companies…

  15. MicroRNA-101 mediates the suppressive effect of laminar shear stress on mTOR expression in vascular endothelial cells

    SciTech Connect

    Chen, Kui; Fan, Wendong; Wang, Xing; Ke, Xiao; Wu, Guifu; Hu, Chengheng

    2012-10-12

    Highlights: Black-Right-Pointing-Pointer Laminar shear stress upregulates miR-101 expression in vascular endothelial cells. Black-Right-Pointing-Pointer miR-101 represses mTOR expression through a specific 3 Prime UTR binding site. Black-Right-Pointing-Pointer Overexpression of miR-101 inhibits G1/S transition and endothelial cell proliferation. Black-Right-Pointing-Pointer Blockade of miR-101 attenuates the suppressive effect of laminar flow on mTOR expression. -- Abstract: Shear stress associated with blood flow plays an important role in regulating gene expression and cell function in endothelial cells (ECs). MicroRNAs (miRNAs) are highly conserved, small non-coding RNAs that negatively regulate the expression of target genes by binding to the mRNA 3 Prime -untranslated region (3 Prime UTR) at the posttranscriptional level involved in diverse cellular processes. This study demonstrates that microRNA-101 in response to laminar shear stress (LSS) is involved in the flow regulation of gene expression in ECs. qRT-PCR analysis showed that miR-101 expression was significantly upregulated in human umbilical vein endothelial cells (HUVECs) exposed to 12 dyn/cm{sup 2} laminar shear stress for 12 h. We found that transfection of miR-101 significantly decreased the luciferase activity of plasmid reporter containing the 3 Prime UTR of mammalian target of rapamycin (mTOR) gene. Western analysis revealed that the protein level of mTOR was significantly reduced in ECs transfected with miR-101. Furthermore, miR-101 overexpression induced cell cycle arrest at the G1/S transition and suppressed endothelial cell proliferation. Finally, transfection of miR-101 inhibitors attenuated the suppressive effects of LSS on mTOR expression, which identified the efficacy of loss-of-function of miR-101 in laminar flow-treated ECs. In conclusion, we have demonstrated that upregulation of miR-101 in response to LSS contributes to the suppressive effects of LSS on mTOR expression and EC

  16. Systematic screening for mutations in the promoter and the coding region of the 5-HT{sub 1A} gene

    SciTech Connect

    Erdmann, J.; Shimron-Abarbanell, D.; Cichon, S.

    1995-10-09

    In the present study we sought to identify genetic variation in the 5-HT{sub 1A} receptor gene which through alteration of protein function or level of expression might contribute to the genetic predisposition to neuropsychiatric diseases. Genomic DNA samples from 159 unrelated subjects (including 45 schizophrenic, 46 bipolar affective, and 43 patients with Tourette`s syndrome, as well as 25 healthy controls) were investigated by single-strand conformation analysis. Overlapping PCR (polymerase chain reaction) fragments covered the whole coding sequence as well as the 5{prime} untranslated region of the 5-HT{sub 1A} gene. The region upstream to the coding sequence we investigated contains a functional promoter. We found two rare nucleotide sequence variants. Both mutations are located in the coding region of the gene: a coding mutation (A{yields}G) in nucleotide position 82 which leads to an amino acid exchange (Ile{yields}Val) in position 28 of the receptor protein and a silent mutation (C{yields}T) in nucleotide position 549. The occurrence of the Ile-28-Val substitution was studied in an extended sample of patients (n = 352) and controls (n = 210) but was found in similar frequencies in all groups. Thus, this mutation is unlikely to play a significant role in the genetic predisposition to the diseases investigated. In conclusion, our study does not provide evidence that the 5-HT{sub 1A} gene plays either a major or a minor role in the genetic predisposition to schizophrenia, bipolar affective disorder, or Tourette`s syndrome. 29 refs., 4 figs., 1 tab.

  17. Secondary structure and the role in translation initiation of the 5'-terminal region of p53 mRNA.

    PubMed

    Błaszczyk, Leszek; Ciesiołka, Jerzy

    2011-08-23

    The p53 protein is one of the major factors involved in cell cycle control, DNA repair, and induction of apoptosis. We determined the secondary structure of the 5'-terminal region of p53 mRNA that includes two major translation initiation codons AUG1 and AUG2, responsible for the synthesis of p53 and its N-truncated isoform ΔN-p53. It turned out that a part of the coding sequence was involved in the folding of the 5' untranslated region for p53. The most characteristic structural elements in the 5'-terminal region of p53 mRNA were two hairpin motifs. In one of them, the initiation codon AUG1 was embedded while the other hairpin has been earlier shown to bind the Mdm2 protein. Alternative mechanisms of p53 mRNA translation initiation were investigated in vitro using model mRNA templates. The results confirmed that initiation from AUG1 was mostly cap-dependent. The process was stimulated by a cap structure and strongly inhibited by a stable hairpin at the template 5' end. Upon inhibition, the remaining protein fraction was synthesized in a cap-independent process, which was strongly stimulated by the addition of a cap analogue. The translation initiation from AUG2 showed a largely cap-independent character. The 5' cap structure actually decreased initiation from this site which argues against a leaky scanning mechanism but might suggest the presence of an IRES. Moreover, blocking cap-dependent translation from AUG1 by the stable hairpin did not change the level of initiation from AUG2. Upon addition of the cap analogue, translation from this site was even increased.

  18. The regulatory region controlling the nitrate-responsive expression of a nitrate reductase gene, NIA1, in Arabidopsis.

    PubMed

    Konishi, Mineko; Yanagisawa, Shuichi

    2011-05-01

    Nitrate reductase (NR) is the enzyme that catalyzes the first step of nitrate assimilation. It is well known that the expression of NR genes is rapidly induced in various plants by nitrate. Previously, the activity of a tobacco NR gene promoter was reported to be high in tobacco plants grown on medium containing ammonium as the sole nitrogen source, but low in tobacco plants grown on nitrate-containing medium. This cast some doubt on the role of the NR gene promoter in the nitrate-inducible expression of this gene. Furthermore, in previous studies, transformation with genomic fragments containing NR loci restored the reduced NR activity in NR mutants to a limited extent, suggesting a complex regulation of NR gene expression. Here, we show that although the 1.9 kb promoter of an NR gene in Arabidopsis, NIA1, is not activated by nitrate, the expression of a GUS (β-glucuronidase) reporter gene inserted between the 5'- and 3'-flanking sequences of the NIA1 coding region is strongly induced by nitrate. When the 3'-flanking sequence was fused downstream of the GUS gene under the control of the 35S minimal promoter, its expression was also strongly induced by nitrate. Furthermore, dissection analysis of the 3'-flanking region revealed that the sequence downstream of the transcriptional terminator rather than the 3'-untranslated region plays a role in nitrate-inducible expression, indicating a requirement for the 3'-flanking sequence for the nitrate-inducible transcription of NIA1. We also show that the 2.7 kb promoter sequence of NIA2, another NR gene of Arabidopsis, cannot direct nitrate-inducible expression.

  19. Isolation of cosmid and cDNA clones in the region surrounding the BTK gene at Xq21.3-q22

    SciTech Connect

    Vorechovsky, I.; Zhou, J.N.; Hammarstroem, L.

    1994-06-01

    A regional physical and transcription map involving yeast artificial chromosomes (YACs), cosmids, and cDNAs has been constructed for Xq21.3-q22 around the gene BTK (formerly atk or BPK) defective in X-linked agammaglobulinemia (XLA). With a positional cloning strategy employing direct cDNA selection, novel cDNAs were found to cluster in the region of approximately 100 kb flanking the XLA and {alpha}-galactosidase A loci. While these widely expressed transcripts are in the area known to contain CpG islands, a less evolutionarily conserved gene, located more than 130 kb distal of DXS178, maps to cosmid clones that could not be digested with rare-cutting restriction enzymes. The presence of transcribed sequences flanking the BTK allowed investigation of their involvement in complex XLA phenotypes. Southern blot analysis using cDNA clones isolated from this region permitted exclusion of a contiguous deletion syndrome as an underlying defect in three patients with XLA and associated growth hormone deficiency. A single XLA patient with torsion dystonia and cosegregating X-linked deafness has been found with a deletion in the 3{prime} part of BTK extending centromerically into the flanking expressed sequence DXS1274E. This suggests a possible involvement of the DXS1274E in this phenotype. The GenBank accession numbers for novel cDNA sequences are as follows: DXS1269E (L20773), DXS1271E (UO1923), DXS1273E (UO1925), and DXS1274E (UO1922). 51 refs., 4 figs., 1 tab.

  20. Active region seismology

    NASA Technical Reports Server (NTRS)

    Bogdan, Tom; Braun, D. C.

    1995-01-01

    Active region seismology is concerned with the determination and interpretation of the interaction of the solar acoustic oscillations with near-surface target structures, such as magnetic flux concentration, sunspots, and plage. Recent observations made with a high spatial resolution and a long temporal duration enabled measurements of the scattering matrix for sunspots and solar active regions to be carried out as a function of the mode properties. Based on this information, the amount of p-mode absorption, partial-wave phase shift, and mode mixing introduced by the sunspot, could be determined. In addition, the possibility of detecting the presence of completely submerged magnetic fields was raised, and new procedures for performing acoustic holography of the solar interior are being developed. The accumulating evidence points to the mode conversion of p-modes to various magneto-atmospheric waves within the magnetic flux concentration as being the unifying physical mechanism responsible for these diverse phenomena.

  1. The immediate upstream region of the 5′-UTR from the AUG start codon has a pronounced effect on the translational efficiency in Arabidopsis thaliana

    PubMed Central

    Kim, Younghyun; Lee, Goeun; Jeon, Eunhyun; Sohn, Eun ju; Lee, Yongjik; Kang, Hyangju; Lee, Dong wook; Kim, Dae Heon; Hwang, Inhwan

    2014-01-01

    The nucleotide sequence around the translational initiation site is an important cis-acting element for post-transcriptional regulation. However, it has not been fully understood how the sequence context at the 5′-untranslated region (5′-UTR) affects the translational efficiency of individual mRNAs. In this study, we provide evidence that the 5′-UTRs of Arabidopsis genes showing a great difference in the nucleotide sequence vary greatly in translational efficiency with more than a 200-fold difference. Of the four types of nucleotides, the A residue was the most favourable nucleotide from positions −1 to −21 of the 5′-UTRs in Arabidopsis genes. In particular, the A residue in the 5′-UTR from positions −1 to −5 was required for a high-level translational efficiency. In contrast, the T residue in the 5′-UTR from positions −1 to −5 was the least favourable nucleotide in translational efficiency. Furthermore, the effect of the sequence context in the −1 to −21 region of the 5′-UTR was conserved in different plant species. Based on these observations, we propose that the sequence context immediately upstream of the AUG initiation codon plays a crucial role in determining the translational efficiency of plant genes. PMID:24084084

  2. Light-associated and processing-dependent protein binding to 5' regions of rbcL mRNA in the chloroplasts of a C4 plant.

    PubMed

    McCormac, D J; Litz, H; Wang, J; Gollnick, P D; Berry, J O

    2001-02-02

    In amaranth, a C(4) dicotyledonous plant, the plastid rbcL gene (encoding the large subunit of ribulose-1,5-bisphosphate carboxylase) is regulated post-transcriptionally during many developmental processes, including light-mediated development. To identify post-transcriptional regulators of rbcL expression, three types of analyses (polysome heel printing, gel retardation, and UV cross-linking) were utilized. These approaches revealed that multiple proteins interact with 5' regions of rbcL mRNA in light-grown, but not etiolated, amaranth plants. Light-associated binding of a 47-kDa protein (p47), observed by UV cross-linking, was highly specific for the rbcL 5' RNA. Binding of p47 occurred only with RNAs corresponding to mature processed rbcL transcripts (5'-untranslated region (UTR) terminating at -66); transcripts with longer 5'-UTRs did not associate with p47 in vitro. Variations in the length of the rbcL 5'-UTR were found to occur in vivo, and these different 5' termini may prevent or enhance light-associated p47 binding, possibly affecting rbcL expression as well. p47 binding correlates with light-dependent rbcL polysome association of the fully processed transcripts in photosynthetic leaves and cotyledons but not with cell-specific rbcL mRNA accumulation in bundle sheath and mesophyll chloroplasts.

  3. Enhanced translation by Nucleolin via G-rich elements in coding and non-coding regions of target mRNAs.

    PubMed

    Abdelmohsen, Kotb; Tominaga, Kumiko; Lee, Eun Kyung; Srikantan, Subramanya; Kang, Min-Ju; Kim, Mihee M; Selimyan, Roza; Martindale, Jennifer L; Yang, Xiaoling; Carrier, France; Zhan, Ming; Becker, Kevin G; Gorospe, Myriam

    2011-10-01

    RNA-binding proteins (RBPs) regulate gene expression at many post-transcriptional levels, including mRNA stability and translation. The RBP nucleolin, with four RNA-recognition motifs, has been implicated in cell proliferation, carcinogenesis and viral infection. However, the subset of nucleolin target mRNAs and the influence of nucleolin on their expression had not been studied at a transcriptome-wide level. Here, we globally identified nucleolin target transcripts, many of which encoded cell growth- and cancer-related proteins, and used them to find a signature motif on nucleolin target mRNAs. Surprisingly, this motif was very rich in G residues and was not only found in the 3'-untranslated region (UTR), but also in the coding region (CR) and 5'-UTR. Nucleolin enhanced the translation of mRNAs bearing the G-rich motif, since silencing nucleolin did not change target mRNA stability, but decreased the size of polysomes forming on target transcripts and lowered the abundance of the encoded proteins. In summary, nucleolin binds G-rich sequences in the CR and UTRs of target mRNAs, many of which encode cancer proteins, and enhances their translation.

  4. Exploring the 5'-UTR DNA region as a target for optimizing recombinant gene expression from the strong and inducible Pm promoter in Escherichia coli.

    PubMed

    Berg, Laila; Kucharova, Veronika; Bakke, Ingrid; Valla, Svein; Brautaset, Trygve

    2012-04-30

    By using the strong and inducible Pm promoter as a model, we recently reported that the β-lactamase production (encoded by bla) can be stimulated up to 20-fold in Escherichia coli by mutating the DNA region corresponding to the 5'-untranslated region of mRNA (UTR). One striking observation was the unexpected large stimulatory effect some of these UTR variants had on the bla transcript production level. We here demonstrate that such UTR variants can also be used to improve the expression level of the alternative genes celB (encoding phosphoglucomutase) and inf-α2b (encoding human cytokine interferon α2b), which both can be expressed to high levels even with the wild-type Pm UTR DNA sequence. Our data indicated some degree of context dependency between the UTR DNA and concomitant recombinant gene sequences. By constructing and using a synthetic operon, we demonstrated that UTR variants optimized for high-level expression of probably any recombinant gene can be efficiently selected from large UTR mutant libraries. The stimulation affected both the transcript production and translational level, and such modified UTR sequences therefore clearly have a significant applied potential for improvement of recombinant gene expression processes.

  5. Regional Seismic Wave Propagation

    DTIC Science & Technology

    1979-07-31

    Baikal to the Pamirs, earthquakes occuring in the Baikal region, Sinkiang , the Gobi desert, southwest China and the Himalayas generated Lg/P ratios...data were obtained from stations within the USSR from earthquake events occuring in Baikal, Sinkiang , the Gobi desert, Southwest China and the...earthquakes originating in the Sinkiang province and recorded by seismo- graphic stations along the Pamir-Lena River profile [25] 0 - recorded by short

  6. Northwest Regional Climate Assessment

    NASA Technical Reports Server (NTRS)

    Lipschultz, Fred

    2011-01-01

    Objectives are to establish a continuing, inclusive National process that: 1) synthesizes relevant science and information 2) increases understanding of what is known & not known 3) identifies information needs related to preparing for climate variability and change, and reducing climate impacts and vulnerability 4) evaluates progress of adaptation & mitigation activities 5) informs science priorities 6) builds assessment capacity in regions and sectors 7) builds understanding & skilled use of findings

  7. MC-7 Cebrenia Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-7 quadrangle, Cebrenia region of Mars. The northwestern two-thirds is dominated by light-colored and dark, relatively smooth plains. The southeastern part is marked by one of three prominent Elysium shield volcanoes, Hecates Tholus, and the ridge system of Phlegra Montes. Latitude range 30 to 65 degrees, longitude range -180 to -120 degrees.

  8. MC-23 Aeolis Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-23 quadrangle, Aeolis region of Mars. The southern part is dominated by heavily cratered highlands that are cut by two large channels having features characteristic of terrestrial river beds. The highlands are separated from the northern plains of Elysium Planitia by a highly dissected, discontinuous northwest trending scarp. The northeastern part is marked by a large shield volcano, Apollinaris Patera. Latitude range - 30 to 0 degrees, longitude range -180 to -135 degrees.

  9. MC-12 Arabia Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-12 quadrangle, Arabia region of Mars. Heavily cratered highlands dominate the Arabia quadrangle. The northeastern part is marked by a large impact crater, Cassini. Cassini is an ancient remnant of the many large impact events that occurred during the period of heavy bombardment. Latitude range 0 to 30 degrees, longitude range -45 to 0 degrees.

  10. Mc-9 Tharsis Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-9 quadrangle, Tharsis region of Mars. Three of the four largest shield volcanoes on Mars--Olympus, Ascraeus, and Pavonis Montes--lie within the Tharsis quadrangle, together with several smaller shields. The north-central part is marked by highly faulted terrain of Ceraunius Fossae. Latitude range 0 to 30 degrees, longitude range 90 to 135 degrees.

  11. MC-26 Argyre Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-26 quadrangle, Argyre region of Mars. The west-central part is marked by the large Argyre impact basin, defined by a rim of rugged mountain blocks that surrounds a nearly circular expanse of light-colored plains 800 km across. The large basin is surrounded by heavily cratered highlands. Latitude range -65 to -30 degrees, longitude range 0 to 60 degrees.

  12. Regional Renewable Energy Cooperatives

    NASA Astrophysics Data System (ADS)

    Hazendonk, P.; Brown, M. B.; Byrne, J. M.; Harrison, T.; Mueller, R.; Peacock, K.; Usher, J.; Yalamova, R.; Kroebel, R.; Larsen, J.; McNaughton, R.

    2014-12-01

    We are building a multidisciplinary research program linking researchers in agriculture, business, earth science, engineering, humanities and social science. Our goal is to match renewable energy supply and reformed energy demands. The program will be focused on (i) understanding and modifying energy demand, (ii) design and implementation of diverse renewable energy networks. Geomatics technology will be used to map existing energy and waste flows on a neighbourhood, municipal, and regional level. Optimal sites and combinations of sites for solar and wind electrical generation (ridges, rooftops, valley walls) will be identified. Geomatics based site and grid analyses will identify best locations for energy production based on efficient production and connectivity to regional grids and transportation. Design of networks for utilization of waste streams of heat, water, animal and human waste for energy production will be investigated. Agriculture, cities and industry produce many waste streams that are not well utilized. Therefore, establishing a renewable energy resource mapping and planning program for electrical generation, waste heat and energy recovery, biomass collection, and biochar, biodiesel and syngas production is critical to regional energy optimization. Electrical storage and demand management are two priorities that will be investigated. Regional scale cooperatives may use electric vehicle batteries and innovations such as pump storage and concentrated solar molten salt heat storage for steam turbine electrical generation. Energy demand management is poorly explored in Canada and elsewhere - our homes and businesses operate on an unrestricted demand. Simple monitoring and energy demand-ranking software can easily reduce peaks demands and move lower ranked uses to non-peak periods, thereby reducing the grid size needed to meet peak demands. Peak demand strains the current energy grid capacity and often requires demand balancing projects and

  13. Transition Region Flows

    NASA Astrophysics Data System (ADS)

    Brekke, P.; Murdin, P.

    2000-11-01

    Ultraviolet emission lines emitted from the SOLAR TRANSITION REGION are often shifted from their expected rest wavelengths. Shifts of spectral lines are due to the so-called DOPPLER EFFECT, where the source of emission is moving either away from or towards the observer, causing a change in the apparent wavelength. The shifted emission lines are most often interpreted as a flow of plasma along ...

  14. Regional technical cooperation.

    PubMed

    Sullivan, P H

    1997-01-01

    The AIDS epidemic threatens economic development in Asia because Asia offers fertile conditions for unchecked transmission and because the epidemic has the most impact on young adults who make up a large sector of the work force. Prevention is still possible, however, and should be viewed as an investment in the future. Effective prevention strategies will have regional as well as domestic components and will recognize the hierarchy of interventions and spread the burden among the public sector, nongovernmental organizations (NGOs), and the private sector in each country. The public sector should 1) ensure that markets function well and do not discriminate against infected individuals; 2) provide a supportive macroeconomic framework of fiscal, trade, and credit policies; and 3) provide public and quasipublic goods, such as information and training. The contribution of NGOs should are vital for reducing the suffering involved with HIV/AIDS. Private sector contributions can include care facilities, research and development, and funding. The private sector must realize that the threat to the stock of human capital posed by HIV/AIDS will reduce profits. The regional dimensions of the HIV/AIDS epidemic relate 1) to factors that contribute to transmission and 2) to approaches that can be taken to prevent transmission and curb its impact. The Greater Mekong Subregion Work Program on HIV/AIDS is a good example of a cooperative regional effort to prevent and control HIV/AIDS. The epidemic requires cooperation among sectors and among countries.

  15. Regional Shelter Analysis Methodology

    SciTech Connect

    Dillon, Michael B.; Dennison, Deborah; Kane, Jave; Walker, Hoyt; Miller, Paul

    2015-08-01

    The fallout from a nuclear explosion has the potential to injure or kill 100,000 or more people through exposure to external gamma (fallout) radiation. Existing buildings can reduce radiation exposure by placing material between fallout particles and exposed people. Lawrence Livermore National Laboratory was tasked with developing an operationally feasible methodology that could improve fallout casualty estimates. The methodology, called a Regional Shelter Analysis, combines the fallout protection that existing buildings provide civilian populations with the distribution of people in various locations. The Regional Shelter Analysis method allows the consideration of (a) multiple building types and locations within buildings, (b) country specific estimates, (c) population posture (e.g., unwarned vs. minimally warned), and (d) the time of day (e.g., night vs. day). The protection estimates can be combined with fallout predictions (or measurements) to (a) provide a more accurate assessment of exposure and injury and (b) evaluate the effectiveness of various casualty mitigation strategies. This report describes the Regional Shelter Analysis methodology, highlights key operational aspects (including demonstrating that the methodology is compatible with current tools), illustrates how to implement the methodology, and provides suggestions for future work.

  16. Region counting algorithm based on region labeling automaton

    NASA Astrophysics Data System (ADS)

    Yang, Sudi; Gu, Guoqing

    2007-12-01

    Region counting is a conception in computer graphics and image analysis, and it has many applications in medical area recently. The existing region-counting algorithms are almost based on filling method. Although filling algorithm has been improved well, the speed of these algorithms used to count regions is not satisfied. A region counting algorithm based on region labeling automaton is proposed in this paper. By tracing the boundaries of the regions, the number of the region can be obtained fast. And the proposed method was found to be fastest and requiring less memory.

  17. Cloning and functional characterization of the 5' regulatory region of ovine Hormone Sensitive Lipase (HSL) gene.

    PubMed

    Lampidonis, Antonis D; Stravopodis, Dimitrios J; Voutsinas, Gerassimos E; Messini-Nikolaki, Niki; Stefos, George C; Margaritis, Lukas H; Argyrokastritis, Alexandros; Bizelis, Iosif; Rogdakis, Emmanuel

    2008-12-31

    Hormone Sensitive Lipase (HSL) catalyzes the rate-limiting step in the mobilization of fatty acids from adipose tissue, thus determining the supply of energy substrates in the body. HSL enzymatic activity is increased by adrenergic agonists, such as catecholamines and glucagons, which induce cyclic AMP (cAMP) intracellular production, subsequently followed by the activation of Protein Kinase A (PKA) and its downstream signaling cascade reactions. HSL constitutes the critical enzyme in the modulation of lipid stores and the only component being subjected to hormonal control in terms of the recently identified Adipose Triglyceride Lipase (ATGL). In order to acquire detailed knowledge with regard to the mechanisms regulating ovine HSL (ovHSL) gene transcription activity, we initially isolated and cloned the 5' proximal and distal promoter regions through a genome walking approach, with the utilization of the already characterized ovHSL cDNAs. As evinced by BLAST analysis and a multiple alignment procedure, the isolated genomic fragment of 2.744 kb appeared to contain the already specified 5'-untranslated region (5'-UTR), which was interrupted by a relatively large intron of 1.448 kb. Regarding the upstream remaining part of 1.224 kb, it was demonstrated to represent a TATA-less promoter area, harboring several cis-regulatory elements that could be putatively recognized by relatively more general transcription factors, mainly including Stimulating protein 1 (Sp1), CCAAT-box Binding Factors (CBFs), Activator Protein 2 (AP2) and Glucocorticoid Receptor (GR), as well as other cis-acting regions denominated as Insulin Response Element (IRE), Glucose Response Element (GRE), Fat Specific Element (FSE) and cAMP Response Element (CRE), which could likely function in a nourishment (i.e. glucose)-/hormone-dependent fashion. When different genomic fragments were directionally (5' to 3') cloned into a suitable reporter vector upstream of a promoter-less luciferase gene and

  18. Southeast Regional Experiment Station

    NASA Astrophysics Data System (ADS)

    1994-08-01

    This is the final report of the Southeast Regional Experiment Station project. The Florida Solar Energy Center (FSEC), a research institute of the University of Central Florida (UCF), has operated the Southeast Regional Experiment Station (SE RES) for the US Department of Energy (DOE) since September 1982. Sandia National Laboratories, Albuquerque (SNLA) provides technical program direction for both the SE RES and the Southwest Regional Experiment Station (SW RES) located at the Southwest Technology Development Institute at Las Cruces, New Mexico. This cooperative effort serves a critical role in the national photovoltaic program by conducting system evaluations, design assistance and technology transfer to enhance the cost-effective utilization and development of photovoltaic technology. Initially, the research focus of the SE RES program centered on utility-connected PV systems and associated issues. In 1987, the SE RES began evaluating amorphous silicon (a-Si) thin-film PV modules for application in utility-interactive systems. Stand-alone PV systems began receiving increased emphasis at the SE RES in 1986. Research projects were initiated that involved evaluation of vaccine refrigeration, water pumping and other stand-alone power systems. The results of this work have led to design optimization techniques and procedures for the sizing and modeling of PV water pumping systems. Later recent research at the SE RES included test and evaluation of batteries and charge controllers for stand-alone PV system applications. The SE RES project provided the foundation on which FSEC achieved national recognition for its expertise in PV systems research and related technology transfer programs. These synergistic products of the SE RES illustrate the high visibility and contributions the FSEC PV program offers to the DOE.

  19. [Ultrasound and regional anaesthesia].

    PubMed

    Delaunay, L; Plantet, F; Jochum, D

    2009-02-01

    The use of ultrasound is the latest major evolution in regional anaesthesia. Review of available literature shows significant changes in clinical practice. Ultrasound guidance allows the visualization of anatomical variations or unsuspected intraneural injections, reduces the volume of local anaesthetic injections and confirms correct local anaesthetic distribution or catheter placement. No study has found a statistical difference in success rates and safety because all studies were underpowered. However, the ability to visualize an invasive procedure that has been performed blindly in the past is an undeniable progress in terms of safety. The necessity to be familiar with the machine and the learning curve can be repulsive. The aim of this article is to demystify ultrasound guidance by explaining the fundamentals of the clinical use of ultrasound. With the help of different chapters, the authors explain the different adjustments and possible artefacts and give easy solutions for the use of bedside ultrasound. Training is essential and can be performed on manikins or training phantom. For each region the main anatomical landmarks are explained. One must be familiar with several imaging techniques: short axis (transverse) or long axis (longitudinal) nerve imaging, in-plane or out-of-plane imaging and hydrolocalization. Viewing the needle's tip position during its progression remains the main safety endpoint. Therefore, electrical nerve stimulation and ultrasound guidance should be combined, especially for beginners, to confirm proximity to neural structures and to help in case of difficulty. Optimizing safety and clinical results must remain a key priority in regional anaesthesia. Finally, specific regulations concerning the transducers are described. Paediatric specificities are also mentioned.

  20. Regional river sulfur runoff

    SciTech Connect

    Husar, R.B.; Husar, J.D.

    1985-01-20

    The water and sulfur runoff data for 54 large river basins were assembled, covering 65% of the nondesert land area of the world. The sulfur concentration ranges from 0.5 mg S/L for the West African rivers Niger and Volta to 100 mg S/L in the Colorado River; the world average is 3.2 mg S/L. The concentrations in central and eastern Europe as well as central and eastern North America exceed 8 mg S/L. The sulfur runoff density is also highest in the river basins over these industrialized regions, exceeding 2 g S/m/sup 2//yr. However, high sulfur runoff density in excess of 3 g S/m/sup 2//yr is also measured over the Pacific islands New Zealand and New Guinea and the archipelagos of Indonesia and the Philippines. The natural background sulfur runoff was estimated by assuming that South America, Africa, Australia, and the Pacific Islands are unperturbed by man and that the average river sulfur concentration is in the range 1--3 mg S/L. Taking these background concentration values, the man-induced sulfur runoff for Europe ranges between 2 and 8 times the natural flow, and over North America, man's contribution ranges between 1 and 5 times the natural runoff. The global sulfur flow from nondesert land to the oceans and the Caspian Sea is estimated as 131 Tg S/yr, of which 46--85 Tg S/yr is attributed to natural causes. The regional river sulfur runoff pattern discussed in this paper does not have enough spatial resolution to be directly applicable to studies of the environmental effects of man-induced sulfur flows. However, it points to the continental-size regions where those perturbations are most evident and to the magnitude of the perturbations as expressed in units of the natural flows.

  1. Regional river sulfur runoff

    NASA Astrophysics Data System (ADS)

    Husar, Rudolf B.; Husar, Janja Djukic

    1985-01-01

    The water and sulfur runoff data for 54 large river basins were assembled, covering 65% of the nondesert land area of the world. The sulfur concentration ranges from 0.5 mg S/L for the West African rivers Niger and Volta to 100 mg S/L in the Colorado River; the world average is 3.2 mg S/L. The concentrations in central and eastern Europe as well as central and eastern North America exceed 8 mg S/L. The sulfur runoff density is also highest in the river basins over these industrialized regions, exceeding 2 g S/m2/yr. However, high sulfur runoff density in excess of 3 g S/m2/yr is also measured over the Pacific islands New Zealand and New Guinea and the archipelagos of Indonesia and the Philippines. The natural background sulfur runoff was estimated by assuming that South America, Africa, Australia, and the Pacific Islands are unperturbed by man and that the average river sulfur concentration is in the range 1-3 mg S/L. Taking these background concentration values, the man-induced sulfur runoff for Europe ranges between 2 and 8 times the natural flow, and over North America, man's contribution ranges between 1 and 5 times the natural runoff. The global sulfur flow from nondesert land to the oceans and the Caspian Sea is estimated as 131 Tg S/yr, of which 46-85 Tg S/yr is attributed to natural causes. The regional river sulfur runoff pattern discussed in this paper does not have enough spatial resolution to be directly applicable to studies of the environmental effects of man-induced sulfur flows. However, it points to the continental-size regions where those perturbations are most evident and to the magnitude of the perturbations as expressed in units of the natural flows.

  2. Complex regional pain syndrome

    PubMed Central

    Sebastin, Sandeep J

    2011-01-01

    Complex regional pain syndrome (CRPS) previously known as reflex sympathetic dystrophy is a chronic neurological disorder involving the limbs characterized by disabling pain, swelling, vasomotor instability, sudomotor abnormality, and impairment of motor function. CRPS is not uncommon after hand surgery and may complicate post-operative care. There is no specific diagnostic test for CRPS and the diagnosis is based on history, clinical examination, and supportive laboratory findings. Recent modifications to diagnostic criteria have enabled clinicians to diagnose this disease more consistently. This review gives a synopsis of CRPS and discusses the diagnosis, pathophysiology, and treatment options based on the limited evidence in the literature. PMID:22022040

  3. MC-27 Noachis Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-27 quadrangle, Noachis region of Mars. Heavily cratered highlands dominate the Noachis quadrangle. The northeastern part is marked by the western half of the ancient large Hellas basin, defined by a rim of rugged mountain blocks that surrounds a nearly circular expanse of light-colored plains. The southeastern part is marked by the Peneus caldera and part of the Amphitrites shield volcano and associated ridged plains that may be basaltic flows. Latitude range -65 to -30 degrees, longitude range -60 to 0 degrees.

  4. MC-8 Amazonis Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-8 quadrangle, Amazonis region of Mars. The central part, which is dominated by light-colored, relatively smooth to hummocky plains of Amazonis Planitia, is partly bounded to the east by the western flank of the largest known volcano in the solar system, Olympus Mons, and its associated aureole deposits. Moderately cratered knobby terrain is west of the plains of Amazonis Planitia. Latitude range 0 to 30 degrees, longitude range 135 to 180 degrees.

  5. MC-15 Elysium Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-15 quadrangle, Elysium region of Mars. The Elysium quadrangle includes relatively smooth lowland plains immediately north of the more cratered highlands. The plains are interrupted on the northwest by two large shield volcanoes, Elysium Mons and Albor Tholus. The plains are also marked by an elongate crater, Orcus Patera, at the east boundary and a band of knobby terrain that extends northeastward through the eastern part. Latitude range 0 to 30 degrees, longitude range -180 to -135 degrees.

  6. MC-18 Coprates Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-18 quadrangle, Coprates region of Mars. Moderately cratered and faulted highland ridged plains in the northern and southern parts are cut by the prominent Valles Marineris chasma system, which reaches depths of 10 km and extends in an east-southeast direction for about 2,500 km across the quadrangle. The long, central canyons appear to be large, fault-bounded rifts, whereas some of the isolated, northern canyons are the sources of large outflow channels. Latitude range -30 to 0 degrees, longitude range 45 to 90 degrees.

  7. MC-21 Iapygia Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-21 quadrangle, Iapygia region of Mars. Heavily cratered and in places dissected highlands dominate the Iapygia quadrangle. The west-central part is marked by a large impact crater, Huygens. Huygens is an ancient remnant of the many large impact events that occurred during the period of heavy bombardment. The southern one-third is characterized by mountainous and knobby terrain of the northern rim of the enormous Hellas impact basin. Latitude range -30 to 0 degrees, longitude range -90 to -45 degrees.

  8. MC-29 Eridania Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-29 quadrangle, Eridania region of Mars. The quadrangle is dominated by heavily cratered highlands, with some moderately cratered plains in the central part and large ridge systems in the southern part. The west-central part is marked by a large impact crater, Kepler. Kepler is an ancient remnant of the many large impact events that occurred during the period of heavy bombardment. Latitude range -65 to 30 degrees, longitude range -180 to -120 degrees.

  9. Regional flood probabilities

    USGS Publications Warehouse

    Troutman, B.M.; Karlinger, M.R.

    2003-01-01

    The T-year annual maximum flood at a site is defined to be that streamflow, that has probability 1/T of being exceeded in any given year, and for a group of sites the corresponding regional flood probability (RFP) is the probability that at least one site will experience a T-year flood in any given year. The RFP depends on the number of sites of interest and on the spatial correlation of flows among the sites. We present a Monte Carlo method for obtaining the RFP and demonstrate that spatial correlation estimates used in this method may be obtained with rank transformed data and therefore that knowledge of the at-site peak flow distribution is not necessary. We examine the extent to which the estimates depend on specification of a parametric form for the spatial correlation function, which is known to be nonstationary for peak flows. It is shown in a simulation study that use of a stationary correlation function to compute RFPs yields satisfactory estimates for certain nonstationary processes. Application of asymptotic extreme value theory is examined, and a methodology for separating channel network and rainfall effects on RFPs is suggested. A case study is presented using peak flow data from the state of Washington. For 193 sites in the Puget Sound region it is estimated that a 100-year flood will occur on the average every 4,5 years.

  10. Colorado Regional Faults

    DOE Data Explorer

    Hussein, Khalid

    2012-02-01

    Citation Information: Originator: Earth Science &Observation Center (ESOC), CIRES, University of Colorado at Boulder Originator: Colorado Geological Survey (CGS) Publication Date: 2012 Title: Regional Faults Edition: First Publication Information: Publication Place: Earth Science & Observation Center, Cooperative Institute for Research in Environmental Science, University of Colorado, Boulder Publisher: Earth Science &Observation Center (ESOC), CIRES, University of Colorado at Boulder Description: This layer contains the regional faults of Colorado Spatial Domain: Extent: Top: 4543192.100000 m Left: 144385.020000 m Right: 754585.020000 m Bottom: 4094592.100000 m Contact Information: Contact Organization: Earth Science &Observation Center (ESOC), CIRES, University of Colorado at Boulder Contact Person: Khalid Hussein Address: CIRES, Ekeley Building Earth Science & Observation Center (ESOC) 216 UCB City: Boulder State: CO Postal Code: 80309-0216 Country: USA Contact Telephone: 303-492-6782 Spatial Reference Information: Coordinate System: Universal Transverse Mercator (UTM) WGS’1984 Zone 13N False Easting: 500000.00000000 False Northing: 0.00000000 Central Meridian: -105.00000000 Scale Factor: 0.99960000 Latitude of Origin: 0.00000000 Linear Unit: Meter Datum: World Geodetic System 1984 (WGS ’984) Prime Meridian: Greenwich Angular Unit: Degree Digital Form: Format Name: Shape file

  11. Mountain regions in peril

    NASA Astrophysics Data System (ADS)

    Showstack, Randy

    The United Nations has declared 2002 the International Year of Mountains (IYM) to bring attention to a number of threats that affect ecosystems and human populations in mountainous and highland regions around the world.“More than half of humanity—3 billion people—relies on mountains for safe, fresh water, water to grow food, to produce electricity to sustain industries and, most important, water to drink,“ said Jacques Diouf, director-general of the U.N. Food and Agriculture Organization, the primary IYM sponsor. “Yet, mountain glaciers, the source of water for many of the world's river systems and people, are melting at unprecedented rates.”

  12. Complex regional pain syndrome

    PubMed Central

    Carr, Emily S.; De La Cerda, Ashley

    2016-01-01

    Complex regional pain syndrome (CRPS) is a neurologic disorder that often results in debilitating chronic pain, but the diagnosis may elude providers as it is one of exclusion. A history of trauma may be elucidated. We report a case of CRPS and review the clinical findings, appropriate workup, and treatment options for the patient. The patient we describe went through an extensive workup before receiving the correct diagnosis. Delay in diagnosis leads to prolonged suffering for the patient and, at times, unnecessary invasive debridement procedures. Raising awareness of this entity may help physicians make the correct diagnosis early, as well as initiate a collaborative effort between neurology, anesthesiology, and dermatology to provide the patient the most favorable outcome. PMID:27365892

  13. MC-28 Hellas Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-28 quadrangle, Hellas region of Mars. The northwestern part includes the eastern half of the ancient large Hellas basin, defined by a rim of rugged mountain blocks that surrounds a nearly circular expanse of light-colored plains. The southwestern part is marked by part of the Amphitrites shield volcano and associated ridged plains that may be basaltic lava flows. The northern part includes a low, dissected shield volcano, Hadriaca Patera, and associated plains that may be basaltic lava flows. These plains are dissected in places by large sinuous channels. The eastern part is dominated by high-standing heavily cratered highlands, perhaps uplifted by the Hellas impact event. Latitude range -65 to -30 degrees, longitude range -120 to -60 degrees.

  14. MC-25 Thaumasia Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-25 quadrangle, Thaumasia region of Mars. In the northern part, a prominent physiographic feature, Thaumasia plateau, includes a complex array of small- and large-scale faults and ridges and ancient volcanoes. Channel systems, upturned beds, and large troughs occur in places along the southern edge of the plateau. In the southern part, the lowlands that surround the plateau include heavily cratered highland terrain and relatively smooth, low plains. The east-central part is marked by a relatively young large impact crater, Lowell. Lowell is one of several large, relatively young impact events on Mars that followed the period of heavy bombardment. Latitude range -65 to -30 degrees, longitude range 60 to 120 degrees.

  15. MC-17 Phoenicis Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-17 quadrangle, Phoenicis Lacus region of Mars. Two of the four largest shield volcanoes on Mars are within the northwestern part, the south half of Pavonis Mons and Arsia Mons. The eastern part includes Syria and Sinai Plana. Most of the quadrangle forms the Tharsis plateau--the highest plateau on Mars; its elevation, 10 km, is twice that of the Tibetan Plateau, the highest plateau on Earth. Also in the northeastern part is Noctis Labyrinthus, a complex system of fault valleys at the west end of Valles Marineris. The south-central part is marked by the large fault system, Claritas Fossae. Latitude range -30 to 0 degrees, longitude range 90 to 135 degrees.

  16. Strengthening regional safeguards

    SciTech Connect

    Palhares, L.; Almeida, G.; Mafra, O.

    1996-08-01

    Nuclear cooperation between Argentina and Brazil has been growing since the early 1980`s and as it grew, so did cooperation with the US Department of Energy (DOE). The Brazilian-Argentine Agency for Accounting and Control of Nuclear Materials (ABACC) was formed in December 1991 to operate the Common System of Accounting and Control of Nuclear Materials (SCCC). In April 1994, ABACC and the DOE signed an Agreement of Cooperation in nuclear material safeguards. This cooperation has included training safeguards inspectors, exchanging nuclear material measurement and containment and surveillance technology, characterizing reference materials, and studying enrichment plant safeguards. The goal of the collaboration is to exchange technology, evaluate new technology in Latin American nuclear facilities, and strengthen regional safeguards. This paper describes the history of the cooperation, its recent activities, and future projects. The cooperation is strongly supported by all three governments: the Republics of Argentina and Brazil and the United States.

  17. Reull Vallis Source Region

    NASA Technical Reports Server (NTRS)

    2002-01-01

    [figure removed for brevity, see original site] (Released 1 July 2002) The jumbled, chaotic terrain in this THEMIS image may represent a source region for the Reull Vallis, one of the larger channel systems in the southern hemisphere of Mars. Such regions of chaos are thought to form by the catastrophic release of groundwater. If this was the case, then the water would have flowed down gradient to the south and may have contributed to the formation of the Reull Vallis. The top of the image shows two short segments of channels that are interrupted by the chaos, demonstrating that there was a channel system in place before the ground foundered to produce the chaos. One of the more intriguing features seen among the jumbled blocks are narrow ledges that vaguely resemble bath tub rings in the way they conform to the topography. Two good examples are seen running roughly left-right across the image about a fourth of the way down. At first they appear to be layers protruding from the cliff faces, but upon closer inspection a more ledge-like character is evident. Note how they appear different between the south-facing and north facing cliffs. The occurrence of one of these features on the south-facing interior rim of the largest crater in the image but nowhere else around the rim argues against the idea that the ledges are due to a layer of rock cropping out throughout the landscape. Instead, they appear more like the edges of a layer of sediment that drapes the topography. It is possible that the sediment is mixed with ice and is best preserved in the shadowed portions of the terrain. There is no easy explanation for these unusual features. They represent one more Martian enigma.

  18. Variants in the 1q21 risk region are associated with a visual endophenotype of autism and schizophrenia.

    PubMed

    Goodbourn, P T; Bosten, J M; Bargary, G; Hogg, R E; Lawrance-Owen, A J; Mollon, J D

    2014-02-01

    Deficits in sensitivity to visual stimuli of low spatial frequency and high temporal frequency (so-called frequency-doubled gratings) have been demonstrated both in schizophrenia and in autism spectrum disorder (ASD). Such basic perceptual functions are ideal candidates for molecular genetic study, because the underlying neural mechanisms are well characterized; but they have sometimes been overlooked in favor of cognitive and neurophysiological endophenotypes, for which neural substrates are often unknown. Here, we report a genome-wide association study of a basic visual endophenotype associated with psychological disorder. Sensitivity to frequency-doubled gratings was measured in 1060 healthy young adults, and analyzed for association with genotype using linear regression at 642 758 single nucleotide polymorphism (SNP) markers. A significant association (P = 7.9 × 10(-9) ) was found with the SNP marker rs1797052, situated in the 5'-untranslated region of PDZK1; each additional copy of the minor allele was associated with an increase in sensitivity equivalent to more than half a standard deviation. A permutation procedure, which accounts for multiple testing, showed that the association was significant at the α = 0.005 level. The region on chromosome 1q21.1 surrounding PDZK1 is an established susceptibility locus both for schizophrenia and for ASD, mirroring the common association of the visual endophenotype with the two disorders. PDZK1 interacts with N-methyl-d-aspartate receptors and neuroligins, which have been implicated in the etiologies of schizophrenia and ASD. These findings suggest that perceptual abnormalities observed in two different disorders may be linked by common genetic elements.

  19. Identification and characterization of Kaposi's sarcoma-associated herpesvirus open reading frame 11 promotor activation

    SciTech Connect

    Chen, Lei

    2008-01-01

    Open reading frame 11 (ORF11) of Kaposi's sarcoma-associated herpesvirus belongs to a herpesviral homologous protein family shared by some members of the gamma- herpesvirus subfamily. Little is known about this ORF11 homologous protein family. We have characterized an unknown open reading frame, ORF11, located adjacent and in the opposite orientation to a well-characterized viral IL-6 gene. Northern blot analysis reveals that ORF11 is expressed during the KSHV lytic cycle with delayed-early transcription kinetics. We have determined the 5{prime} and 3{prime} untranslated region of the unspliced ORF11 transcript and identified both the transcription start site and the transcription termination site. Core promoter region, representing ORF11 promoter activity, was mapped to a 159nt fragment 5{prime} most proximal to the transcription start site. A functional TATA box was identified in the core promoter region. Interestingly, we found that ORF11 transcriptional activation is not responsive to Rta, the KSHV lytic switch protein. We also discovered that part of the ORF11 promoter region, the 209nt fragment upstream of the transcription start site, was repressed by phorbol esters. Our data help to understand transcription regulation of ORF11 and to elucidate roles of ORF11 in KSHV pathogenesis and life cycle.

  20. Molecular weight abnormalities of the CTCF transcription factor: CTCF migrates aberrantly in SDS-PAGE and the size of the expressed protein is affected by the UTRs and sequences within the coding region of the CTCF gene.

    PubMed

    Klenova, E M; Nicolas, R H; U, S; Carne, A F; Lee, R E; Lobanenkov, V V; Goodwin, G H

    1997-02-01

    CTCF belongs to the Zn finger transcription factors family and binds to the promoter region of c-myc. CTCF is highly conserved between species, ubiquitous and localised in nuclei. The endogenous CTCF migrates as a 130 kDa (CTCF-130) protein on SDS-PAGE, however, the open reading frame (ORF) of the CTCF cDNA encodes only a 82 kDa protein (CTCF-82). In the present study we investigate this phenomenon and show with mass-spectra analysis that this occurs due to aberrant mobility of the CTCF protein. Another paradox is that our original cDNA, composed of the ORF and 3'-untranslated region (3'-UTR), produces a protein with the apparent molecular weight of 70 kDa (CTCF-70). This paradox has been found to be an effect of the UTRs and sequences within the coding region of the CTCF gene resulting in C-terminal truncation of CTCF-130. The potential attenuator has been identified and point-mutated. This restored the electrophoretic mobility of the CTCF protein to 130 kDa. CTCF-70, the aberrantly migrating CTCF N-terminus per se, is also detected in some cell types and therefore may have some biological implications. In particular, CTCF-70 interferes with CTCF-130 normal function, enhancing transactivation induced by CTCF-130 in COS6 cells. The mechanism of CTCF-70 action and other possible functions of CTCF-70 are discussed.

  1. Molecular weight abnormalities of the CTCF transcription factor: CTCF migrates aberrantly in SDS-PAGE and the size of the expressed protein is affected by the UTRs and sequences within the coding region of the CTCF gene.

    PubMed Central

    Klenova, E M; Nicolas, R H; U, S; Carne, A F; Lee, R E; Lobanenkov, V V; Goodwin, G H

    1997-01-01

    CTCF belongs to the Zn finger transcription factors family and binds to the promoter region of c-myc. CTCF is highly conserved between species, ubiquitous and localised in nuclei. The endogenous CTCF migrates as a 130 kDa (CTCF-130) protein on SDS-PAGE, however, the open reading frame (ORF) of the CTCF cDNA encodes only a 82 kDa protein (CTCF-82). In the present study we investigate this phenomenon and show with mass-spectra analysis that this occurs due to aberrant mobility of the CTCF protein. Another paradox is that our original cDNA, composed of the ORF and 3'-untranslated region (3'-UTR), produces a protein with the apparent molecular weight of 70 kDa (CTCF-70). This paradox has been found to be an effect of the UTRs and sequences within the coding region of the CTCF gene resulting in C-terminal truncation of CTCF-130. The potential attenuator has been identified and point-mutated. This restored the electrophoretic mobility of the CTCF protein to 130 kDa. CTCF-70, the aberrantly migrating CTCF N-terminus per se, is also detected in some cell types and therefore may have some biological implications. In particular, CTCF-70 interferes with CTCF-130 normal function, enhancing transactivation induced by CTCF-130 in COS6 cells. The mechanism of CTCF-70 action and other possible functions of CTCF-70 are discussed. PMID:9016583

  2. Regional brain hypometabolism is unrelated to regional amyloid plaque burden

    PubMed Central

    Altmann, Andre; Ng, Bernard; Landau, Susan M.; Jagust, William J.

    2015-01-01

    See Sorg and Grothe (doi:10.1093/brain/awv302) for a scientific commentary on this article. In its original form, the amyloid cascade hypothesis of Alzheimer’s disease holds that fibrillar deposits of amyloid are an early, driving force in pathological events leading ultimately to neuronal death. Early clinicopathological investigations highlighted a number of inconsistencies leading to an updated hypothesis in which amyloid plaques give way to amyloid oligomers as the driving force in pathogenesis. Rather than focusing on the inconsistencies, amyloid imaging studies have tended to highlight the overlap between regions that show early amyloid plaque signal on positron emission tomography and that also happen to be affected early in Alzheimer’s disease. Recent imaging studies investigating the regional dependency between metabolism and amyloid plaque deposition have arrived at conflicting results, with some showing regional associations and other not. We extracted multimodal neuroimaging data from the Alzheimer’s disease neuroimaging database for 227 healthy controls and 434 subjects with mild cognitive impairment. We analysed regional patterns of amyloid deposition, regional glucose metabolism and regional atrophy using florbetapir (18F) positron emission tomography, 18F-fluordeoxyglucose positron emission tomography and T1-weighted magnetic resonance imaging, respectively. Specifically, we derived grey matter density and standardized uptake value ratios for both positron emission tomography tracers in 404 functionally defined regions of interest. We examined the relation between regional glucose metabolism and amyloid plaques using linear models. For each region of interest, correcting for regional grey matter density, age, education and disease status, we tested the association of regional glucose metabolism with (i) cortex-wide florbetapir uptake; (ii) regional (i.e. in the same region of interest) florbetapir uptake; and (iii) regional florbetapir uptake

  3. Characterization of a human X-linked gene from the DXS732E locus in the candidate region for the anhidrotic ectodermal dysplasia (EDA) gene (Xq13.1)

    SciTech Connect

    Gault, J.; Zonana, J.; Zeltinger, J.

    1994-09-01

    A conserved mouse genomic clone was used to identify a homologous human genomic clone (the DXS732E locus), which was subsequently employed to isolate cDNAs from a human fetal brain library. Nine unique overlapping cDNAs were isolated, and sequences analysis of 3.9 kb identified a putative 1 kb ORF. GRAIL analysis of the sequence supported the hypothesis that the putative ORF was coding sequence, and Prosite analysis of the putative ORF identified potential glycosylation and phosphorylation sites. The 5{prime} end of the gene maps within a CpG island, and comparison of cDNA sequences indicate the gene is alternatively spliced at its 3{prime} end. Northern analysis and RT-PCR indicate that two different sized messages appear to be expressed with the gene expressed in human fetal kidney, intestine, brain, and muscle. The gene is expressed in 77 day human skin, a time when hair follicle formation occurs. Anhidrotic ectodermal dysplasia (EDA) results in the abnormal morphogenesis of hair, teeth and eccrine sweat glands. A positional cloning strategy towards cloning the EDA gene had been used, and deletion and X-autosome translocation patients have been useful in further delimiting the EDA region. The present gene at the DXS732E locus is partially deleted in one EDA patient who does not have other apparent abnormalities. No rearrangements of the gene have been detected in two female X-autosome translocation EDA patients, nor in four additional male patients with submicroscopic molecular deletions.

  4. The gene for human U2 snRNP auxiliary factor small 35-kDa subunit (U2AF1) maps to the progressive myoclonus epilepsy (EPM1) critical region on chromosome 21q22.3

    SciTech Connect

    Lalioti, M.D.; Rossier, C.; Antonarakis, S.E.

    1996-04-15

    We used targeted exon trapping to clone portions of genes from human chromosome 21q22.3. One trapped sequence showed complete homology with the cDNA of human U2AF{sup 35} (M96982; HGM-approved nomenclature U2AF1), which encodes for the small 35-kDa subunit of the U2 snRNP auxiliary factor. Using the U2AF1 cDNA as a probe, we mapped this gene to cosmid Q15D2, a P1, and YAC 350F7 of the Chumakov et al. contig, close to the cystathionine-{beta}-synthase gene (CBS) on 21q22.3. This localization was confirmed by PCR using oligonucleotides from the 3{prime} UTR and by FISH. As U2AF1 associated with a number of different factors during mRNA splicing, overexpression in trisomy 21 individuals could contribute to some Down syndrome phenotypes by interfering with the splicing process. Furthermore, because this gene maps in the critical region for the progressive myoclonus epilepsy I locus (EPM1), mutation analysis will be carried out in patients to evaluate the potential role of U2AF1 as a candidate for EPM1. 24 refs., 1 fig.

  5. Landslides of Palestinian Region

    NASA Astrophysics Data System (ADS)

    Alwahsh, H.

    2013-12-01

    Natural disasters are extreme sudden events caused by environmental and natural actors that take away the lives of many thousands of people each year and damage large amount of properties. They strike anywhere on earth, often without any warning. A risk maps of natural disaster are very useful to identify the places that might be adversely affected in the event of natural disaster. The earthquakes are one of natural disaster that have the greatest hazards and will cause loss of life and properties due to damaging the structures of building, dams, bridges. In addition, it will affect local geology and soil conditions. The site effects play an important role in earthquake risk because of its amplification or damping simulation. Another parameter in developing risk map is landslide, which is also one of the most important topics in site effect hazards. Palestine region has been suffering landslide hazards because of the topographical and geological conditions of this region. Most Palestine consists of mountainous area, which has great steep slopes and the type of soil is mainly grayish to yellowish silty clay (Marl Soil). Due to the above mentioned factors many landslides have been occurred from Negev south to the northern borders of Palestine. An example of huge and destruction landslide in a Palestine authority is the landslide in the White Mountain area in the city of Nablus, which occurred in 1997. The geotechnical and geophysical investigation as well as slope stability analysis should be considered in making landslide maps that are necessary to develop risk levels of the natural disaster. Landslides occurred in slopes that are created naturally or by human beings. Failure of soil mass occurs, and hence landslide of soil mass happen due to sliding of soil mass along a plane or curved surface. In general, the slopes become unstable when the shear stresses (driving force) generated in the soil mass exceed the available shearing resistance on the rupture surface

  6. USArray Regional Phase Analysis

    NASA Astrophysics Data System (ADS)

    Buehler, J. S.; Shearer, P. M.

    2014-12-01

    The regional Pn and Sn phases, which are typically described as headwaves that propagate in the uppermost mantle, are sensitive to heterogeneities in the mantle lid and complement other seismic studies with poorer vertical resolution at this depth. We have experimented with a variety of approaches to image the velocity structure and anisotropy in the western U.S., starting with separate Pn and Sn time-term tomographies, but also localized cross-correlation and stacking approaches that benefit from the regular USArray station arrangement. Later we combined the data sets for joint Pn-Sn inversions and the resulting Vp/Vs maps provide further insight into the nature of the seismic anomalies. Now that USArray has reached the east coast, we are updating our models to include the cumulative station footprint. The sparser source distribution in the eastern U.S., and the resulting longer ray paths, provide new challenges and justify the inclusion of additional parameters that account for the velocity gradient in the mantle lid. Our results show generally higher Pn velocities in the eastern U.S., but we observe patches of lower velocities around the New Madrid seismic zone and below the eastern Appalachians. We find that the Pn fast axes generally do not agree with SKS splitting orientations, suggesting significant vertical changes in anisotropy in the upper mantle. For example, the circular pattern of the fast polarization direction of SKS in the western U.S. is much less pronounced in the Pn results, and in the eastern U.S. the dominant Pn fast direction is approximately north-south, whereas the SKS fast polarizations are oriented roughly parallel to the absolute plate motion direction. Since Pn and Sn travel through the crust, they can provide additional information on crustal thickness. In several regions our results and estimates from receiver function studies are inconsistent. For example, beneath the Colorado Plateau our crustal thickness estimates are about 35-40 km

  7. A Regional Resource: Appalachian Campuses

    ERIC Educational Resources Information Center

    Roesch, Harry

    1975-01-01

    An Appalachian Regional Commission survey of 180 institutions of higher education in the Appalachian Region pinpoints which institutions offer technical assistance to state and local governments and officals. (Author)

  8. On Titan's Xanadu region

    NASA Astrophysics Data System (ADS)

    Brown, Robert H.; Barnes, Jason W.; Melosh, H. Jay

    2011-08-01

    A large, circular marking ˜1800 km across is seen in near-infrared images of Titan. The feature is centered at 10°S, 120°W on Titan, encompasses much of Titan's western Xanadu region, and has an off-center, quasi-circular, inner margin about 700 km across, with lobate outer margins extending 200-500 km from the inner margin. On the feature's southern flank is Tui Regio, an area that has very high reflectivity at 5 μm, and is hypothesized to exhibit geologically recent cryovolcanic flows (Barnes, J.W. et al. [2006]. Geophys. Res. Lett. 33), similar to flows seen in Hotei Regio, a cryovolcanic area whose morphology may be controlled by pre-existing, crustal fractures resulting from an ancient impact (Soderblom, L.A. et al. [2009]. Icarus, 204). The spectral reflectivity of the large, circular feature is quite different than that of its surroundings, making it compositionally distinct, and radar measurements of its topography, brightness temperature and volume scattering also suggest that the feature is quite distinct from its surroundings. These and several other lines of evidence, in addition to the feature's morphology, suggest that it may occupy the site of an ancient impact.

  9. Sudurnes Regional Heating Corp.

    SciTech Connect

    Lienau, P.J.

    1996-11-01

    The Svartsengi geothermal area is close to the town of Grindavik on the Rekjanes peninsula and is part of an active fissure swarm, lined with crater-rows and open fissures and faults. The high-temperature area has an area of 2 sq. km and shows only limited signs of geothermal activity at the surface. The reservoir, however, contains lots of energy and at least 8 wells supply the Svartsengi Power Plant with steam. The steam is not useable for domestic heating purposes so that heat exchangers are used to heat cold groundwater with the steam. Some steam is also used for producing 16.4 MW{sub e} of electrical power. The article shows the distribution system piping hot water to nine towns and the Keflavik International Airport. The effluent brine from the Svartsengi Plant is disposed of into a surface pond, called the Blue Lagoon, popular to tourists and people suffering from psoriasis and other forms of eczema seeking therapeutic effects from the silica rich brine. This combined power plant and regional district heating system (cogeneration) is an interesting and unique design for the application of geothermal energy.

  10. A duplicated gene in the breakpoint regions of the 7q11.23 Williams-Beuren syndrome deletion encodes the initiator binding protein TFII-I and BAP-135, a phosphorylation target of BTK.

    PubMed

    Pérez Jurado, L A; Wang, Y K; Peoples, R; Coloma, A; Cruces, J; Francke, U

    1998-03-01

    Williams-Beuren syndrome (WBS) is a neurodevelopmental disorder with multisystemic manifestations caused by heterozygosity for a partial deletion of chromosome band 7q11.23. The breakpoints cluster within regions located approximately 1 cM either side of the elastin (ELN) locus. We have characterized a duplicated region near the common deletion breakpoints, which includes a transcribed gene. The centromeric (C) and telomeric (T) copies are almost identical in the duplicated 3[prime] portions but diverge at their 5[prime]-ends. C-specific 4.3 kb mRNA and T-specific 5.4 kb mRNA are widely expressed in embryonic and adult tissues. The telomeric gene gives rise to several alternatively spliced forms and is deleted in all WBS individuals who have documented ELN deletions. Database searches revealed that this gene encodes BAP-135, a protein phosphorylated by Bruton's tyrosine kinase in B cells, as well as the multifunctional transcription factor TFII-I, hence the gene name GTF2I. The centromeric gene is not deleted in WBS and appears to be a partially truncated expressed pseudogene with no protein product (gene name GTF2IP1). Both loci map to different genomic clone contigs that also contain other deleted and non-deleted loci. A probe from the shared region recognizes a >3 Mb Not I junction fragment that is unique to individuals with the WBS deletion. Therefore, the duplicated region containing GTF2I and GTF2IP1 respectively is located close to the deletion breakpoints and may predispose to unequal meiotic recombination between chromosome 7 homologs and/or to intrachromosomal rearrangements. Hemizygosity for GTF2I may also contribute to the WBS phenotype.

  11. Europa Wedge Region

    NASA Technical Reports Server (NTRS)

    1998-01-01

    This image shows an area of crustal separation on Jupiter's moon, Europa. Lower resolution pictures taken earlier in the tour of NASA's Galileo spacecraft revealed that dark wedge-shaped bands in this region are areas where the icy crust has completely pulled apart. Dark material has filled up from below and filled the void created by this separation.

    In the lower left corner of this image, taken by Galileo's onboard camera on December 16, 1997, a portion of one dark wedge area is visible, revealing a linear texture along the trend of the wedge. The lines of the texture change orientation slightly and reflect the fact that we are looking at a bend in the wedge. The older, bright background, visible on the right half of the image, is criss-crossed with ridges. A large, bright ridge runs east-west through the upper part of the image, cutting across both the older background plains and the wedge. This ridge is rough in texture, with numerous small terraces and troughs containing dark material.

    North is to the top of the picture and the sun illuminates the surface from the northwest. This image, centered at approximately 16.5 degrees south latitude and 196.5 degrees west longitude, covers an area approximately 10 kilometers square (about 6.5 miles square). The resolution of this image is about 26 meters per picture element. This image was taken by the solid state imaging system from a distance of 1250 kilometers (750 miles).

    The Jet Propulsion Laboratory, Pasadena, CA manages the Galileo mission for NASA's Office of Space Science, Washington, DC. JPL is an operating division of California Institute of Technology (Caltech).

    This image and other images and data received from Galileo are posted on the World Wide Web, on the Galileo mission home page at URL http://www.jpl.nasa.gov/ galileo.

  12. Regions. [Africa, Middle East].

    PubMed

    1985-03-01

    This discussion of population focuses on the regions of Africa and the Middle East. In South Africa more white women are working but fewer black women work. The overall result is that the percentage of women who work is declining. Marita de Beer, research liaison executive at the South African Advertising Research Foundation, reports that the female population grew by 31% in the past 10 years while the number of working women has grown by only 11%. Among blacks the female population rose by 36%, but the number of workers among them declined by about 1%. Married women are among the fastest growing groups of working women in South Africa. The most recent estimate of the population of Nigeria is 92 million. According to Professor Vremudia Diejomaoh, Nigeria's population will probably reach 155 million by 2000 with 33% living in urban areas. In Saudi Arabia the Pan Arab Research Center recently completed a census of retail outlets in 3 metropolitan areas: Jeddah, Riyadh, and Dammam. The types of outlets surveyed include large supermarkets, small supermarkets, groceries with and without deep freeze, tobacco shops, meat shop/delis, small cafeterias, large restaurants/hotels, cosmetics shops or perfumeries, camera stores, toy shops, pharmacies, watch and gift shop, newsstands, department store, and appliance outlets. Using the Census of Retail Outlets as a base, Pan Arab Research Center also has a new distribution audit system that will cover 500 outlets. By plotting Arab countries according to their population policies and their current growth rates, it is possible to project where the middle class will grow fastest in the Arab world. The countries that have declining growth rates and strong population programs designed to encourage lower fertility rates among women are Egypt, Tunisia, Morocco, Algeria, and Lebanon. The countries most likely to have a better per capita distribution of resources within this decade are those where governments encourage reductions in

  13. Identification of a novel gene expressed in activated natural killer cells and T cells

    SciTech Connect

    Dahl, C.A.; Schall, R.P.; He, H.; Cairns, J.S. )

    1992-01-15

    The authors have isolated a cDNA clone from a human activated NK cell-derived cDNA library that identifies a transcript [NK4] that is selectively expressed in lymphocytes. The expression of this transcript is increased after activation of T cells by mitogens or activation of NK cells by IL-2 (lymphokine-activated killer cells). The transcript levels demonstrated by Northern blot analysis increase by 12 h after activation, remain high for at least 48 h, and require protein synthesis for expression. Southern blot analysis of B lymphoblastoid lines derived from 18 unrelated individuals reveal variable banding patterns suggestive of polymorphism within the NK4 gene. No homology was found between the sequence of the coding region of this transcript and any sequences in the GenBank data base. Sequence homology to the U1 small nuclear RNA was found within the 3[prime] untranslated region immediately upstream of the site of polyadenylation, suggesting a possible role for U1 in the polyadenylation process. Sequence analysis indicates the transcript would encode a protein having a mass of 27 kDa. The presence of a signal sequence and lack of a transmembrane region suggests that the protein is secreted. In addition, the protein contains an RGD sequence that may be involved in cellular adhesion. This transcript appears to encode a novel product common to the activation pathways of both NK cells and T cells. 50 refs., 8 figs.

  14. Regional governance: strategies and disputes in health region management

    PubMed Central

    dos Santos, Adriano Maia; Giovanella, Ligia

    2014-01-01

    OBJECTIVE To analyze the regional governance of the health systemin relation to management strategies and disputes. METHODOLOGICAL PROCEDURES A qualitative study with health managers from 19 municipalities in the health region of Bahia, Northeastern Brazil. Data were drawn from 17 semi-structured interviews of state, regional, and municipal health policymakers and managers; a focus group; observations of the regional interagency committee; and documents in 2012. The political-institutional and the organizational components were analyzed in the light of dialectical hermeneutics. RESULTS The regional interagency committee is the chief regional governance strategy/component and functions as a strategic tool for strengthening governance. It brings together a diversity of members responsible for decision making in the healthcare territories, who need to negotiate the allocation of funding and the distribution of facilities for common use in the region. The high turnover of health secretaries, their lack of autonomy from the local executive decisions, inadequate technical training to exercise their function, and the influence of party politics on decision making stand as obstacles to the regional interagency committee’s permeability to social demands. Funding is insufficient to enable the fulfillment of the officially integrated agreed-upon program or to boost public supply by the system, requiring that public managers procure services from the private market at values higher than the national health service price schedule (Brazilian Unified Health System Table). The study determined that “facilitators” under contract to health departments accelerated access to specialized (diagnostic, therapeutic and/or surgical) services in other municipalities by direct payment to physicians for procedure costs already covered by the Brazilian Unified Health System. CONCLUSIONS The characteristics identified a regionalized system with a conflictive pattern of governance and

  15. An interactive region merging method

    NASA Astrophysics Data System (ADS)

    Wu, Rongteng

    2011-11-01

    The paper presents a novel region merging method based on the interactive information from users. An image firstly is partitioned into homogeneous regions by using an initial segmentation and the regions will be label by taking an interactive scheme. In this scheme, the users only roughly specify the position and main features of the object and background, then any region will belong to non-label region or label region i.e. object or background. A similarity rule is used to guide the merging process with the help of the users' markers. And then the object of interest is extracted from the image. Experiment results show that the proposed method is efficient for us to extract the object of interest from the complex background.

  16. Sensitive detection of Tomato ringspot virus by real-time TaqMan RT-PCR targeting the highly conserved 3'-UTR region.

    PubMed

    Tang, Joe; Khan, Subuhi; Delmiglio, Catia; Ward, Lisa I

    2014-06-01

    A real-time TaqMan RT-PCR assay was developed for the rapid and sensitive detection of Tomato ringspot virus (ToRSV), an important plant virus which infects a wide range of fruit and ornamental crops. Primers and a probe were designed based on the highly conserved 3'-untranslated region (UTR) sequences of ToRSV, to amplify a 182bp fragment of this region of RNA-1 and RNA-2. The assay was demonstrated to reliably amplify all ToRSV isolates tested. The detection limit was estimated to be about 12 copies of the ToRSV target region. No amplification was observed from the RNA of other nepoviruses or healthy host species. A comparison with a published conventional RT-PCR and a SYBR-based qRT-PCR indicated that both of the published assays lacked reliability and sensitivity, as neither were able to amplify all ToRSV isolates tested, and both were approximately 1000 times less sensitive than the novel TaqMan real-time assay. This TaqMan real-time assay was tested using four different reagent kits and was shown to be robust and stable, with no significant differences in sensitivity between kits. It is expected that the implementation of this TaqMan real-time RT-PCR assay will facilitate efficient phytosanitary certification of nursery stock requiring testing for ToRSV by regulatory agencies, and will also have wider uses for the general detection of ToRSV in a range of hosts.

  17. Regional Climate Tutorial: Assessing Regional Climate Change and Its Impacts

    NASA Astrophysics Data System (ADS)

    Barron, E.; Fisher, A.

    2002-05-01

    Recent scientific progress now enables credible projections of global changes in climate over long time periods. But people will experience global climate change where they live and work, and have difficulty thinking of a future beyond their grandchildren's lifetime. Although the task of projecting climate change and its impacts is far more challenging for regional and relatively near-term time scales, these are the scales at which actions most easily can be taken to moderate negative impacts. This tutorial will summarize what is known about projecting changes in regional climate, and about assessing the impacts for sectors such as forests, agriculture, fresh water quantity and quality, coastal zones, human health, and ecosystems. The Mid-Atlantic Regional Assessment (MARA) is used to provide context and illustrate how adaptation within the region and feedback from other regions influence the impacts that might be experienced.

  18. Global and regional air quality responses to regional CO reductions

    NASA Astrophysics Data System (ADS)

    Fry, M. M.; Adelman, Z.; Dolwick, P.; Jang, C.; West, J. J.

    2011-12-01

    Ozone (O3) precursor emissions influence global and regional air quality and climate through changes in the tropospheric concentrations of O3, methane (CH4), and aerosols. Here we examine the influence of regional carbon monoxide (CO) emissions on air quality by simulating 50% reductions in anthropogenic CO emissions from 10 world regions (Australia/New Zealand, Southeast Asia, East Asia, India, Southern Africa, Northern Africa/Middle East, Former Soviet Union, Europe, South America, and North America), using the global chemical transport model MOZART-4. The IPCC AR5 Representative Concentration Pathway 8.5 (RCP 8.5) emissions inventory for 2005 and global meteorology from the Goddard Earth Observing System Model, version 5 (GEOS-5) for 2004-2005 are used as inputs to MOZART-4, run at 1.9 x 2.5 degree horizontal resolution. Base case global air quality is first simulated for the year 2005, and the resulting distributions of tropospheric O3 and related species are compared with observations. Then CO emission reductions from each of the 10 regions are simulated individually. We quantify global and regional changes in O3 and PM2.5 at the surface and within the troposphere, including the influence of each regional reduction on long-term O3 concentrations via CH4 and the long-range transport of O3 and CO. This analysis shows the sensitivity of global and regional air quality to anthropogenic CO emissions from many world regions, in contrast to previous studies of only a few regions. Beyond this study, these simulations will be used to estimate the net radiative forcing due to CO emission reductions from these world regions.

  19. Hawaii Regional Sediment Management: Regional Sediment Budget for the Poipu Region of Kauai, HI

    DTIC Science & Technology

    2013-06-01

    represent regular maintenance activities such as dredging, bypassing, or nourishment . The region addressed in this study has experienced one or more of these...Hawaii Coastal Geology Group US United States USACE US Army Corps of Engineers USGS US Geological Survey WIS Wave Information Study ERDC/CHL...This Coastal and Hydraulics Engineering Technical Note (CHETN) reviews the development of a regional sediment budget for the Poipu Region on the

  20. What's Happening to Regional Policy?

    ERIC Educational Resources Information Center

    Ravenhall, Mark

    2005-01-01

    Back in November, voters in the North East of England overwhelmingly rejected the move towards an elected regional assembly. The scale of the defeat (three to one) of a Government-backed scheme was a rude awakening for the Office of the Deputy Prime Minister and the range of regional agencies created since 1997. After all, it was felt that the…

  1. About the REL Pacific Region

    ERIC Educational Resources Information Center

    Regional Educational Laboratory Pacific, 2014

    2014-01-01

    REL Pacific is one of ten Regional Educational Laboratories established and funded by the U.S. Department of Education's Institute of Education Sciences. Their region encompasses approximately 4.9 million square miles and serves seven Pacific island entities, including American Samoa; the Commonwealth of the Northern Mariana Islands; the Federated…

  2. CLIMATE IMPACTS ON REGIONAL WATER

    EPA Science Inventory

    The New England region (including the 6 New England
    states plus upstate New York) offers a very diverse geography,
    matched by an equally diverse economy and human
    population. Livelihoods throughout the region are based
    on service industries that depend heavily on comm...

  3. Regional Early Childhood Policy Review

    ERIC Educational Resources Information Center

    Evans, Judith

    2008-01-01

    The UNESCO-UNICEF joint regional policy review project was launched in September 2006 with the aim to support the countries of Asia-Pacific region in meeting the first goal of Education For All (EFA) on Early Childhood Care and Education (ECCE) by identifying, documenting and sharing good practices as well as constraints and challenges in early…

  4. MISR Regional SAMUM Imagery Overview

    Atmospheric Science Data Center

    2016-08-24

    ... View Data  |  Download Data About this Web Site: Visualizations of select MISR Level 3 data for special regional ... in the northern hemisphere (see the table at right). By design, the images show data for a fixed geographical region, with a fixed ...

  5. MISR Regional VBBE Imagery Overview

    Atmospheric Science Data Center

    2016-08-24

    ... View Data  |  Download Data About this Web Site: Visualizations of select MISR Level 3 data for special regional ... in the northern hemisphere (see the table at right). By design, the images show data for a fixed geographical region, with a fixed ...

  6. Polymorphism and genetic mapping of the human oxytocin receptor gene on chromosome 3

    SciTech Connect

    Michelini, S.; Urbanek, M.; Goldman, D.

    1995-06-19

    Centrally administered oxytocin has been reported to facilitate affiliative and social behaviors, in functional harmony with its well-known peripheral effects on uterine contraction and milk ejection. The biological effects of oxytocin could be perturbed by mutations occurring in the sequence of the oxytocin receptor gene, and it would be of interest to establish the position of this gene on the human linkage map. Therefore we identified a polymorphism at the human oxytocin receptor gene. A portion of the 3{prime} untranslated region containing a 30 bp CA repeat was amplified by polymerase chain reaction (PCR), revealing a polymorphism with two alleles occurring with frequencies of 0.77 and 0.23 in a sample of Caucasian CEPH parents (n = 70). The CA repeat polymorphism we detected was used to map the human oxytocin receptor to chromosome 3p25-3p26, in a region which contains several important genes, including loci for Von Hippel-Lindau disease (VHL) and renal cell carcinoma. 53 refs., 2 figs., 1 tab.

  7. Identical homologs of the Galanthus nivalis agglutinin in Zea mays and Fusarium verticillioides.

    PubMed

    Fouquaert, Elke; Peumans, Willy J; Gheysen, Godelieve; Van Damme, Els J M

    2011-01-01

    The structural domain corresponding to the Galanthus nivalis agglutinin (GNA) is a mannose-binding motif that was originally discovered in plants but according to recent data also occurs in other eukaryotes and prokaryotes. Transcriptome analyses revealed that Fusarium verticillioides expresses a protein (FvGLLc1) identical to a recently identified cytoplasmic/nuclear GNA-like lectin from maize (ZmGLLc). The FvGLLc1 and ZmGLLc gene sequences are nearly identical in the coding region as well as in the intron and the 5 and 3 prime untranslated regions. However, whereas the Fusarium genome contains only a single gene with an intron, both an intronless and an intron containing lectin gene can be amplified from maize DNA. Southern blot analysis confirmed the presence of this cytoplasmic GNA-like gene in the maize and rice genome. A comparative analysis of the products amplified by different PCRs using genomic DNA from Fusarium species and maize DNA samples from sterile as well as contaminated plant material strongly indicated that the GNA-like sequence found in maize grown under sterile conditions is not derived from a contaminating Fusarium species. Furthermore, using a PCR-based approach it could be demonstrated that this particular type of lectin occurs also in other plants from distant taxa and is markedly conserved.

  8. Localization of the human fibromodulin gene (FMOD) to chromosome 1q32 and completion of the cDNA sequence

    SciTech Connect

    Sztrolovics, R.; Grover, J.; Roughley, P.J.

    1994-10-01

    This report describes the cloning of the 3{prime}-untranslated region of the human fibromodulin cDNA and its use to map the gene. For somatic cell hybrids, the generation of the PCR product was concordant with the presence of chromosome 1 and discordant with the presence of all other chromosomes, confirming that the fibromodulin gene is located within region q32 of chromosome 1. The physical mapping of genes is a critical step in the process of identifying which genes may be responsible for various inherited disorders. Specifically, the mapping of the fibromodulin gene now provides the information necessary to evaluate its potential role in genetic disorders of connective tissues. The analysis of previously reported diseases mapped to chromosome 1 reveals two genes located in the proximity of the fibromodulin locus. These are Usher syndrome type II, a recessive disorder characterized by hearing loss and retinitis pigmentosa, and Van der Woude syndrome, a dominant condition associated with abnormalities such as cleft lip and palate and hyperdontia. The genes for both of these disorders have been projected to be localized to 1q32 of a physical map that integrates available genetic linkage and physical data. However, it seems improbable that either of these disorders, exhibiting restricted tissue involvement, could be linked to the fibromodulin gene, given the wide tissue distribution of the encoded proteoglycan, although it remains possible that the relative importance of the quantity and function of the proteoglycan may avry between tissues. 11 refs., 1 fig.

  9. Global forcing and regional interactions

    USGS Publications Warehouse

    1992-01-01

    The Climate System Modeling Program (CSMP) sponsored a “Global Forcing and Regional Interaction Workshop” from October 21 to 23, 1991, at Colorado State University's Pingree Park campus, to evaluate the relationship between global climate forcing and the response of the land surface on a regional scale. The general aim of the workshop was to develop specific action plans and preliminary science research strategies for regional-global interactions. Each participant was invited to identify tractable, high pay-off science issues related to global forcing and regional interactions. The workshop, with twenty-six participants about evenly split between atmospheric scientists, hydrologists, and ecologists, was also designed to facilitate a network of collaborators to prepare multidisciplinary research proposals. Discussion also focused on regional climate over the last 200 years and included the influence of atmosphere-land surface processes on natural climate variability. Several major recommendations were made on topics discussed.

  10. About Region 8’s Central Regional Laboratory

    EPA Pesticide Factsheets

    The Region 8 laboratory plays a critical role in protecting people's health and the environment through the analysis of air, water, soil, and biota samples (plant, fish, and occasionally, mammalian tissue).

  11. Regional strategies for global leadership.

    PubMed

    Ghemawat, Pankaj

    2005-12-01

    The leaders of such global powerhouses as GE, Wal-Mart, and Toyota seem to have grasped two crucial truths: First, far from becoming submerged by the rising tide of globalization, geographic and other regional distinctions may in fact be increasing in importance. Second, regionally focused strategies, used in conjunction with local and global initiatives, can significantly boost a company's performance. The business and economic data reveal a highly regionalized world. For example, trade within regions, rather than across them, drove the surge of international commerce in the second half of the twentieth century. Regionalization is also apparent in foreign direct investment, companies' international sales, and competition among the world's largest multinationals. Harvard Business School Professor Pankaj Ghemawat says that the most successful companies employ five types of regional strategies in addition to--or even instead of--global ones: home base, portfolio, hub, platform, and mandate. Some companies adopt the strategies in sequence, but the most nimble switch from one to another and combine approaches as their markets and businesses evolve. At Toyota, for example, exports from the home base continue to be substantial even as the company builds up an international manufacturing presence. And as Toyota achieves economies of scale and scope with a strong network of hubs, the company also pursues economies of specialization through interregional mandates. Embracing regional strategies requires flexibility and creativity. A company must decide what constitutes a region, choose the most appropriate strategies, and mesh those strategies with the organization's existing structures. In a world that is neither truly global nor truly local, finding ways of coordinating within and across regions can deliver a powerful competitive advantage.

  12. Finding Distant Galactic HII Regions

    NASA Astrophysics Data System (ADS)

    Anderson, L. D.; Armentrout, W. P.; Johnstone, B. M.; Bania, T. M.; Balser, Dana S.; Wenger, Trey V.; Cunningham, V.

    2015-12-01

    The WISE Catalog of Galactic H ii Regions contains ˜2000 H ii region candidates lacking ionized gas spectroscopic observations. All candidates have the characteristic H ii region mid-infrared morphology of WISE 12 μ {{m}} emission surrounding 22 μ {{m}} emission, and additionally have detected radio continuum emission. We here report Green Bank Telescope hydrogen radio recombination line and radio continuum detections in the X-band (9 GHz; 3 cm) of 302 WISE H ii region candidates (out of 324 targets observed) in the zone 225^\\circ ≥slant {\\ell }≥slant -20^\\circ , | {\\text{}}b| ≤slant 6^\\circ . Here we extend the sky coverage of our H ii region Discovery Survey, which now contains nearly 800 H ii regions distributed across the entire northern sky. We provide LSR velocities for the 302 detections and kinematic distances for 131 of these. Of the 302 new detections, 5 have ({\\ell },{\\text{}}b,v) coordinates consistent with the Outer Scutum-Centaurus Arm (OSC), the most distant molecular spiral arm of the Milky Way. Due to the Galactic warp, these nebulae are found at Galactic latitudes >1° in the first Galactic quadrant, and therefore were missed in previous surveys of the Galactic plane. One additional region has a longitude and velocity consistent with the OSC but lies at a negative Galactic latitude (G039.183-01.422 -54.9 {km} {{{s}}}-1). With Heliocentric distances >22 kpc and Galactocentric distances >16 kpc, the OSC H ii regions are the most distant known in the Galaxy. We detect an additional three H ii regions near {\\ell }≃ 150^\\circ whose LSR velocities place them at Galactocentric radii >19 kpc. If their distances are correct, these nebulae may represent the limit to Galactic massive star formation.

  13. Active Region Release Two CMEs

    NASA Video Gallery

    Solar material can be seen blowing off the sun in this video captured by NASA’s Solar Dynamics Observatory (SDO) on the night of Feb. 5, 2013. This active region on the sun sent out two coronal ...

  14. Conformity Adequacy Review: Region 3

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 3 (DE, DC, MD, PA, VA, WV) is provided here.

  15. Conformity Adequacy Review: Region 10

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 10 (AK, ID, OR, WA) is provided here.

  16. Conformity Adequacy Review: Region 5

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the conformity SIP adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 5 (IL, IN, MI, OH, WI) is provided here.

  17. Conformity Adequacy Review: Region 7

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 7 (IA, KS, MO, NE) is provided here.

  18. Conformity Adequacy Review: Region 6

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 6 (AR, LA, NM, OK, TX) is provided here.

  19. Conformity Adequacy Review: Region 4

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 4 (AL, FL, GA, KY, MS, NC, SC, TN) is provided here.

  20. Conformity Adequacy Review: Region 2

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 2 (NJ, NY, PR, VI) is provided here.

  1. Conformity Adequacy Review: Region 8

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 3 (CO, MT, ND, SD, UT, WY) is provided here.

  2. Conformity Adequacy Review: Region 1

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 1 (CT, ME, MA, NH, RI, VT) is provided here.

  3. Boundary Preserving Dense Local Regions.

    PubMed

    Kim, Jaechul; Grauman, Kristen

    2015-05-01

    We propose a dense local region detector to extract features suitable for image matching and object recognition tasks. Whereas traditional local interest operators rely on repeatable structures that often cross object boundaries (e.g., corners, scale-space blobs), our sampling strategy is driven by segmentation, and thus preserves object boundaries and shape. At the same time, whereas existing region-based representations are sensitive to segmentation parameters and object deformations, our novel approach to robustly sample dense sites and determine their connectivity offers better repeatability. In extensive experiments, we find that the proposed region detector provides significantly better repeatability and localization accuracy for object matching compared to an array of existing feature detectors. In addition, we show our regions lead to excellent results on two benchmark tasks that require good feature matching: weakly supervised foreground discovery and nearest neighbor-based object recognition.

  4. Conformity Adequacy Review: Region 9

    EPA Pesticide Factsheets

    Resources are for air quality and transportation government and community leaders. Information on the adequacy/inadequacy of state implementation plans (SIPs) in EPA Region 9 (AZ, CA, HI, NV, AS, GU) is provided here.

  5. [Regional anesthesia and cancer immunology].

    PubMed

    Igarashi, Toru; Mori, Katsuya; Inoue, Kei; Suzuki, Takeshi; Morisaki, Hiroshi

    2013-09-01

    Regional anesthesia has been widely applied as an excellent method for perioperative analgesia. Recent studies suggested that regional anesthesia is a promising approach to minimize the risk of surgical site infection and postoperative cancer recurrence, subsequently providing the benefits to the long-term outcome. In particular, it is of great interest that regional anesthesia might be able to reduce cancer recurrence. In cancer patients, innate immunity against cancer could be depressed, resulting in the predisposition to evoke metastasis. Besides, during the perioperative periods, tumor immunity is significantly depressed due to surgical pain, activation of sympathetic nervous system, inflammatory responses, and others. In this review article, we discuss the tumor immunity during the perioperative period, with focus on the alterations of tumor immunity and regional anesthesia.

  6. Multimodal analgesia and regional anaesthesia.

    PubMed

    Tornero Tornero, C; Fernández Rodríguez, L E; Orduña Valls, J

    2017-03-24

    Multimodal analgesia provides quality analgesia, with fewer side effects due to the use of combined analgesics or analgesic techniques. Regional anaesthesia plays a fundamental role in achieving this goal. The different techniques of regional anaesthesia that include both peripheral and central blocks in either a single dose or in continuous infusion help to modulate the nociceptive stimuli that access the central level. The emergence of the ultrasound as an effective system to perform regional anaesthesia techniques has allowed the development of new regional anaesthesia techniques that formerly could not be carried out since only neurostimulation or skin references were used. It is essential to take into account that even with effective blocking it is advisable to associate other drugs by other routes, in this way we will be able to reduce the required doses individually and attempt to achieve a synergistic, not purely additive, effect.

  7. A regional technology transfer program

    NASA Technical Reports Server (NTRS)

    Chenery, P. J.

    1972-01-01

    The activities of the NC/STRC are reported. The background and organization of the regional dissemination center, and marketing methods are discussed along with the services provided, and available information resources.

  8. Variant in the 3' region of the IkappaBalpha gene associated with insulin resistance in Hispanic Americans: The IRAS Family Study.

    PubMed

    Miller, Melissa R; Zhang, Weiming; Sibbel, Scott P; Langefeld, Carl D; Bowden, Donald W; Haffner, Steven M; Bergman, Richard N; Norris, Jill M; Fingerlin, Tasha E

    2010-03-01

    The IKKbeta/NF-kappaB pathway is known to play an important role in inflammatory response and has also recently been implicated in the process of insulin resistance. We hypothesized that one or more variants in the IkappaBalpha gene (NFKBIA) or surrounding untranslated regions would be associated with insulin sensitivity (S(I)) in Hispanic-American families. We tested for association between 25 single-nucleotide polymorphisms (SNPs) in and near NFKBIA and S(I) in 981 individuals in 90 Hispanic-American families from the Insulin Resistance Atherosclerosis (IRAS) Family Study. SNP rs1951276 in the 3' flanking region of NFKBIA was associated with S(I) in the San Antonio (SA) sample after adjusting for age, gender, and admixture (uncorrected P = 1.69 x 10(-5); conservative Bonferroni correction P = 3.38 x 10(-4)). Subjects with at least one A allele for NFKBIA rs1951276 had approximately 29% lower S(I) compared to individuals homozygous for the G allele in the SA sample. Although not statistically significant, the effect was in the same direction in the San Luis Valley (SLV) sample alone (P = 0.348) and was significant in the combined SA and SLV samples (P = 5.37 x 10(-4); presence of A allele associated with approximately 20% lower S(I)). In SA, when adjusted for subcutaneous adipose tissue area (SAT, cm(2)), the association was modestly attenuated (P = 1.25 x 10(-3)), but the association remained highly significant after adjustment for visceral adipose tissue area (VAT, cm(2); P = 4.41 x 10(-6)). These results provide corroborating evidence that the NF-kappaB/IKKbeta pathway may mediate obesity-induced insulin resistance in humans.

  9. The rate-limiting step in yeast PGK1 mRNA degradation is an endonucleolytic cleavage in the 3'-terminal part of the coding region.

    PubMed Central

    Vreken, P; Raué, H A

    1992-01-01

    Insertion of an 18-nucleotide-long poly(G) tract into the 3'-terminal untranslated region of yeast phosphoglycerate kinase (PGK1) mRNA increases its chemical half-life by about a factor of 2 (P. Vreken, R. Van der Veen, V. C. H. F. de Regt, A. L. de Maat, R. J. Planta, and H. A. Raué, Biochimie 73:729-737, 1991). In this report, we show that this insertion also causes the accumulation of a degradation intermediate extending from the poly(G) sequence down to the transcription termination site. Reverse transcription and S1 nuclease mapping experiments demonstrated that this intermediate is the product of shorter-lived primary fragments resulting from endonucleolytic cleavage immediately downstream from the U residue of either of two 5'-GGUG-3' sequences present between positions 1100 and 1200 close to the 3' terminus (position 1251) of the coding sequence. Similar endonucleolytic cleavages appear to initiate degradation of wild-type PGK1 mRNA. Insertion of a poly(G) tract just upstream from the AUG start codon resulted in the accumulation of a 5'-terminal degradation intermediate extending from the insertion to the 1100-1200 region. RNase H degradation in the presence of oligo(dT) demonstrated that the wild-type and mutant PGK1 mRNAs are deadenylated prior to endonucleolytic cleavage and that the half-life of the poly(A) tail is three- to sixfold lower than that of the remainder of the mRNA. Thus, the endonucleolytic cleavage constitutes the rate-limiting step in degradation of both wild-type and mutant PGK1 transcripts, and the resulting fragments are degraded by a 5'----3' exonuclease, which appears to be severely retarded by a poly(G) sequence. Images PMID:1320194

  10. MC-10 Lanae Palus Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-10 quadrangle, Lanae Palus region of Mars. The western part is dominated by lava flows of the Tharsis region. The central part includes ridged terrain of Lunae Planum. The west and north borders of Lunae Planum are dissected by the large, relatively young outflow channel, Kasei Vallis, which terminates in Chryse Planitia. Latitude range 0 to 30 degrees, longitude range 45 to 90 degrees.

  11. ESPC Regional Arctic Prediction System

    DTIC Science & Technology

    2014-09-30

    the Navy the capability to conduct short-term (1 week) to extended (2 weeks) coupled weather forecasts for the Arctic region. APPROACH To...sensitivity of the Arctic weather forecast to key numerical parameters; and 5) conduct extensive validation and verification of the coupled system and...SEP 2014 2. REPORT TYPE 3. DATES COVERED 00-00-2014 to 00-00-2014 4. TITLE AND SUBTITLE ESPC Regional Arctic Prediction System 5a. CONTRACT

  12. MC-13 Syrtis Major Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-13 quadrangle, Syrtis Major region of Mars. The central part is dominated by dark dust and lava flows of the Syrtis Major Planitia region. These lava flows are partly bounded to the east by a large depression, Isidis basin, which contains smooth plains, and to the west and north by heavily cratered and moderately faulted highlands. Latitude range 0 to 30 degrees, longitude range -90 to -45 degrees.

  13. Revisiting Health Regionalization in Canada.

    PubMed

    Barker, Paul; Church, John

    2017-04-01

    Twenty years ago, many of Canada's provinces began to introduce regional health authorities to address problems with their health care systems. With this action, the provinces sought to achieve advances in community decision-making, the integration of health services, and the provision of care in the home and community. The authorities were also to help restrict health care costs. An assessment of the authorities indicates, however, that over the past two decades they have been unable to meet their objectives. Community representatives continue to play little role in determining the appropriate health services for their regions. Gains have been made towards integrating health services, but the plan for a near seamless set of health services has not been realized. Funding for health services remains focused on hospital and physician care, and health care expenditures have until very recently been little affected by regional authorities. This disappointing performance has caused some provinces to abandon their regional authorities, but this article argues that the provision of greater autonomy and a better public appreciation of their role and potential may lead to more successful regional authorities. Accordingly, the objective of this article is to reveal the shortcomings of regional health authorities in Canada while at the same time arguing that changes can be made to increase the chances of more workable authorities.

  14. Regional sinkhole susceptibility maps: The Latium Region case (central Italy)

    NASA Astrophysics Data System (ADS)

    La Vigna, F.; Teoli, P.; Mazza, R.; Leoni, G.; Capelli, G.

    2012-04-01

    Several and frequent studies were internationally presented about landslide susceptibility, meanwhile in literature is missing a broad diffusion of studies regarding sinkhole susceptibility. That's why sinkhole recurrence depends on several geological conditions related to specific geological and hydrogeological context (sinkhole prone area) that vary case by case. Notwithstanding this regionalization problem of sinkhole recurrence, in the central Appenine sedimentary basins (Italy) a certain number of geological, geomorphologic and hydrogeological conditions (sinkhole predisposing issues) can be considered in common between the surveyed sinkholes. Eventually this could be compared with similar geological conditions and sinkhole occurrence in the rest of Italy or in other countries. In this case study is presented a probabilistic approach regarding the Latium Region deriving from the comparison between the regional sinkhole inventory realized during a precedent project and the dataset of the new Hydrogeological Map of Latium Region (scale 1:100.000). Indexed elements, chosen because associated to the majority of sinkhole phenomena, are: outcropping lithologies, water table depth, main faults (even if buried), hydrothermal springs, land use and the epicentres of recent earthquakes. These indexed elements were weighted and combined in a matrix which preliminary result is the sinkhole susceptibility map of Latium Region. When definitively validated, this approach could be suitable for local authorities to planning more targeted studies in major hazard areas.

  15. Ig Constant Region Effects on Variable Region Structure and Function

    PubMed Central

    Janda, Alena; Bowen, Anthony; Greenspan, Neil S.; Casadevall, Arturo

    2016-01-01

    The adaptive humoral immune response is responsible for the generation of antimicrobial proteins known as immunoglobulin molecules or antibodies. Immunoglobulins provide a defense system against pathogenic microbes and toxins by targeting them for removal and/or destruction. Historically, antibodies have been thought to be composed of distinct structural domains known as the variable and constant regions that are responsible for antigen binding and mediating effector functions such as opsonization and complement activation, respectively. These domains were thought to be structurally and functionally independent. Recent work has revealed however, that in some families of antibodies, the two regions can influence each other. We will discuss the body of work that led to these observations, as well as the mechanisms that have been proposed to explain how these two different antibody regions may interact in the function of antigen binding. PMID:26870003

  16. Regional waveform calibration in the Pamir-Hindu Kush region

    NASA Astrophysics Data System (ADS)

    Zhu, Lupei; Helmberger, Donald V.; Saikia, Chandan K.; Woods, Bradley B.

    1997-10-01

    Twelve moderate-magnitude earthquakes (mb 4-5.5) in the Pamir-Hindu Kush region are investigated to determine their focal mechanisms and to relocate them using their regional waveform records at two broadband arrays, the Kyrgyzstan Regional Network (KNET), and the 1992 Pakistan Himalayas seismic experiment array (PAKH) in northern Pakistan. We use the "cut-and-paste" source estimation technique to invert the whole broadband waveforms for mechanisms and depths, assuming a one-dimensional velocity model developed for the adjacent Tibetan plateau. For several large events the source mechanisms obtained agree with those available from the Harvard centroid moment tensor (CMT) solutions. An advantage of using regional broadband waveforms is that focal depths can be better constrained either from amplitude ratios of Pnl to surface waves for crustal events or from time separation between the direct P and the shear-coupled P wave (sPn + sPmP) for mantle events. All the crustal events are relocated at shallower depths compared with their International Seismological Centre bulletin or Harvard CMT depths. After the focal depths are established, the events are then relocated horizontally using their first-arrival times. Only minor offsets in epicentral location are found for all mantle events and the bigger crustal events, while rather large offsets (up to 30 km) occur for the smaller crustal events. We also tested the performance of waveform inversion using only two broadband stations, one from the KNET array in the north of the region and one from the PAKH array in the south. We found that this geometry is adequate for determining focal depths and mechanisms of moderate size earthquakes in the Pamir-Hindu Kush region.

  17. Regional Transmission Projects: Finding Solutions

    SciTech Connect

    The Keystone Center

    2005-06-15

    The Keystone Center convened and facilitated a year-long Dialogue on "Regional Transmission Projects: Finding Solutions" to develop recommendations that will help address the difficult and contentious issues related to expansions of regional electric transmission systems that are needed for reliable and economic transmission of power within and across regions. This effort brought together a cross-section of affected stakeholders and thought leaders to address the problem with the collective wisdom of their experience and interests. Transmission owners sat at the table with consumer advocates and environmental organizations. Representatives from regional transmission organizations exchanged ideas with state and federal regulators. Generation developers explored common interests with public power suppliers. Together, the Dialogue participants developed consensus solutions about how to begin unraveling some of the more intractable issues surrounding identification of need, allocation of costs, and reaching consensus on siting issues that can frustrate the development of regional transmission infrastructure. The recommendations fall into three broad categories: 1. Recommendations on appropriate institutional arrangements and processes for achieving regional consensus on the need for new or expanded transmission infrastructure 2. Recommendations on the process for siting of transmission lines 3. Recommendations on the tools needed to support regional planning, cost allocation, and siting efforts. List of Dialogue participants: List of Dialogue Participants: American Electric Power American Transmission Company American Wind Energy Association California ISO Calpine Corporation Cinergy Edison Electric Institute Environmental Defense Federal Energy Regulatory Commission Great River Energy International Transmission Company ISO-New England Iowa Public Utility Board Kanner & Associates Midwest ISO National Association of Regulatory Utility Commissioners National Association

  18. Hawaii Regional Sediment Management: Regional Sediment Budget for the Kekaha Region of Kauai, HI

    DTIC Science & Technology

    2013-06-01

    study regions, have been developed by the University of Hawaii Coastal Geology Group (UH CGG) (Fletcher et al. 2012) for the US Geological Survey...maintenance activities such as dredging, bypassing, or nourishment . The region addressed in this study has experienced one or more of these activities... Coastal Geology Group US United States USACE US Army Corps of Engineers USGS US Geological Survey WIS Wave Information Study ERDC/CHL CHETN

  19. The genes encoding gonadal and nongonadal forms of 3[beta]-hydroxysteroid dehydrogenase/[Delta][sup 5]-[Delta][sup 4] isomerase are closely linked on mouse chromosome 3

    SciTech Connect

    Bain, P.A.; Meisler, M.H.; Payne, A.H. ); Taylor, B.A. )

    1993-04-01

    The biosynthesis of steroid hormones in the gonads and adrenal glands requires the activities of the enzyme 3[beta]-hydroxysteriod dehydrogenanse/isomerase (3[beta]HSD) which catalyzes the NAD[sup +]-dependant dehydrogenation and subsequent [Delta][sup 5] [r arrow] [Delta][sup 4] isomerization of[Delta][sup 5]-3[beta]-hydroxysteriods to [Delta][sup 4]-3-ketosteroids. The mouse expresses four isoforms of 3[beta]HSD. 3[beta]HSD I is expressed in gonads and adrenal glands and appears to be the major steroidogenic form, 3[beta]HSDs II and III are expressed in both liver and kidneys, and 3[beta]HSD IV has been detected only in kidneys. To determine the genetic relationship between the 3[beta]HSD isoforms, the authors have mapped the chromosomal locations of the four genes by linkage analysis using gene-specific process derived from the 3[prime] untranslated regions of 3 [beta]HSD cDNA clones. The four 3[beta]HSD structural genes (Hsd3b) are closely linked within a segment of chromosome 3 that is conserved on human chromosome 1. The order of markers on Chr 3 surrounding Hsd3b is: centromere-Gba-(4.4 [+-] 2.2)-Hsd3b-(3.3 [+-] 1.9)-Tshb-(6.7 [+-] 2.7)-Amy-1. 28 refs., 5 figs.

  20. cDNA isolated from a human T-cell library encodes a member of the protein-tyrosine-phosphatase family

    SciTech Connect

    Cool, D.E.; Tonks, N.K.; Charbonneau, H.; Walsh, K.A.; Fischer, E.H.; Krebs, E.G. )

    1989-07-01

    A human peripheral T-cell cDNA library was screened with two labeled synthetic oligonucleotides encoding regions of a human placenta protein-tyrosine-phosphatase. One positive clone was isolated and the nucleotide sequence was determined. It contained 1,305 base pairs of open reading frame followed by a TAA stop codon and 978 base pairs of 3{prime} untranslated end, although a poly(A){sup +} tail was not found. An initiator methionine residue was predicted at position 61, which would result in a protein of 415 amino acid residues. This was supported by the synthesis of a M{sub r} 48,000 protein in an in vitro reticulocyte lysate translation system using RNA transcribed from the cloned cDNA and T7 RNA polymerase. The deduced amino acid sequence was compared to other known proteins revealing 65% identity to the low M{sub r} PTPase 1B isolated from placenta. In view of the high degree of similarity, the T-cell cDNA likely encodes a newly discovered protein-tyrosine-phosphatase, thus expanding this family of genes.

  1. Data concatenation, Bayesian concordance and coalescent-based analyses of the species tree for the rapid radiation of Triturus newts.

    PubMed

    Wielstra, Ben; Arntzen, Jan W; van der Gaag, Kristiaan J; Pabijan, Maciej; Babik, Wieslaw

    2014-01-01

    The phylogenetic relationships for rapid species radiations are difficult to disentangle. Here we study one such case, namely the genus Triturus, which is composed of the marbled and crested newts. We analyze data for 38 genetic markers, positioned in 3-prime untranslated regions of protein-coding genes, obtained with 454 sequencing. Our dataset includes twenty Triturus newts and represents all nine species. Bayesian analysis of population structure allocates all individuals to their respective species. The branching patterns obtained by data concatenation, Bayesian concordance analysis and coalescent-based estimations of the species tree differ from one another. The data concatenation based species tree shows high branch support but branching order is considerably affected by allele choice in the case of heterozygotes in the concatenation process. Bayesian concordance analysis expresses the conflict between individual gene trees for part of the Triturus species tree as low concordance factors. The coalescent-based species tree is relatively similar to a previously published species tree based upon morphology and full mtDNA and any conflicting internal branches are not highly supported. Our findings reflect high gene tree discordance due to incomplete lineage sorting (possibly aggravated by hybridization) in combination with low information content of the markers employed (as can be expected for relatively recent species radiations). This case study highlights the complexity of resolving rapid radiations and we acknowledge that to convincingly resolve the Triturus species tree even more genes will have to be consulted.

  2. Human GluR6 kainate receptor (GRIK2): Molecular cloning, expression, polymorphism, and chromosomal assignment

    SciTech Connect

    Paschen, W.; Blackstone, C.D.; Huganir, R.L. ); Ross, C.A. Max-Planck-Institute for Neurological Research, Koeln )

    1994-04-01

    Glutamate receptors mediate the majority of excitatory neurotransmission in the brain, and molecular cloning studies have revealed several distinct families. Because neuropathological states and possibly human disorders may involve kainate-preferring glutamate receptors, the authors have isolated a cDNA clone for the human GluR6 kainate-preferring receptor. This clone shows a very high sequence similarity with that of the rat, except for a part of the 3[prime] untranslated region in which there is a TAA triplet repeat. When the protein was overexpressed in human embryonic kidney 293 cells, it had a molecular weight, an antibody recognition, and a glutamate ligand-binding profile similar to those of the rate GluR6 receptor. Northern analysis showed expression in both human cerebral and cerebellar cortices. By PCR analysis of rodent-human monochromosomal cell lines, the human GluR6 could be assigned to chromosome 6. The length of the TAA triplet repeat was polymorphic in the normal population, with at least four alleles and an observed heterozygosity of about 45%. These studies should provide the basis for expression or linkage studies of the GluR6 kainate receptor in human disease or neuropathologic states. 53 refs., 7 figs.

  3. Cloning of the canine GALC cDNA and identification of the mutation causing globoid cell leukodystrophy in West Highland White and Cairn terriers

    SciTech Connect

    Victoria, T.; Rafi, M.A.; Wenger, D.A.

    1996-05-01

    Globoid cell leukodystrophy, or Krabbe disease, is a severe, autosomal recessive disorder resulting from a deficiency of galactocerebrosidase (GALC) activity. GALC is responsible for the lysosomal catabolism of certain galactolipids, including galactosylceramide and psychosine. In addition to the human patients, there are several naturally occurring animal models for this disease, including the twitcher mouse, West Highland White terriers (WHWT), and Cairn terriers. All species have deficient GALC activity and have the characteristic pathological findings in the nervous system. We now describe the cloning of the canine GALC cDNA and the identification of the disease-causing mutation in both terrier breeds. The 2007-bp open reading frame is 88% identical to that in human, and the deduced amino acid sequence is about 90% identical. However, the 3{prime}-untranslated region is about 1 kb shorter than that in the human. Two nucleotide changes were found in affected dogs, an A to C transversion at cDNA position 473 (Y158S) and a C to T transition at position 1915 (P639S). Expression studies in COS-1 cells demonstrated that the A rapid test for the identification of the genotype at that position has been developed, and over 100 WHWT and Cairn terriers have been screened. This will allow breeders to mate their dogs selectivity and will permit the establishment of a colony of dogs for use in therapy trials. 30 refs., 4 figs., 2 tabs.

  4. The complete exon-intron structure of the 156-kb human gene NFKB1, which encodes the p105 and p50 proteins of transcription factors NF-{kappa}B and I{kappa}B-{gamma}: Implications for NF-{kappa}B-mediated signal transduction

    SciTech Connect

    Heron, E.; Deloukas, P.; van Loon, A.P.G.M.

    1995-12-10

    The NFKB1 gene encodes three proteins of the NF-{kappa}/Rel and I{kappa}B families: p105, p50, and (in mouse) I{kappa}B-{gamma}. We determined the complete genomic structure of human NFKB1. NFKB1 spans 156 kb and has 24 exons with introns varying between 40,000 and 323 bp in length. Although NFKB2, which encodes p100 and p52, also has 24 exons and has a comparable exon-intron structure, it is 20 times shorter than NFKB1. We propose that the long size of NFKB1 is important for transient activation of NF-{kappa}B complexes containing p50. I{kappa}B-{gamma} corresponds to the carboxyl-terminal half of p105. DNA sequence analysis showed that the 3{prime}-end of human intron 11 and the 5{prime}-end of exon 12 of NFKB1 are colinear with the 5{prime}-untranslated region of mouse I{kappa}B-{gamma} cDNA. I{kappa}B-{gamma} is thus likely to be generated by transcription starting within intron 11 and not by alternative splicing of the mouse mRNA encoding p105 and p50. 71 refs., 5 figs., 1 tab.

  5. Phosphoenolpyruvate carboxykinase (GTP): Characterization of the human PCK1 gene and localization distal to MODY on chromosome 20

    SciTech Connect

    Ting, Chao Nan; Burgess, D.L.; Chamberlain, J.S.; Meisler, M.H. ); Keith, T.P.; Falls, K. )

    1993-06-01

    The human PCK 1 gene encoding phosphoenolpyruvate carboxykinase (GTP) (PEPCK) was isolated and sequenced. There is 91% amino acid sequence identity (567/622 residues) between the human and the rat proteins, with conservation of intron/exon borders. A polymorphic dinucleotide microsatellite with the structure (CA)[sub 16](TA)[sub 5](CA) was identified in the 3[prime] untranslated region of the cloned human PCK1 gene. This highly informative genetic marker has an estimated PIC value of 0.79 and heterozygosity of 0.81. Analysis of the RW pedigree demonstrated recombination between PCK1 and the MODY gene on chromosome 20. Multipoint linkage analysis of the reference pedigrees of the Centre d'Etude du Polymorphisme Humain localized PCK1 on the genetic map of chromosome 20 at a position distal to markers that are closely linked to MODY. PCK1 is part of a conserved linkage group on mouse Chromosome 2 with identical gene order but expanded length in the human genome. 34ref., 7 figs., 1 tab.

  6. COL5A1: Genetic mapping and exclusion as candidate gene in families with nail-patella syndrome, tuberous sclerosis 1, hereditary hemorrhagic telangiectasia, and Ehlers-Danlos syndrome type II

    SciTech Connect

    Greenspan, D.S.; Northrup, H.; Au, K.S.

    1995-02-10

    COL5A1, the gene for the {alpha}1 chain of type V collagen, has been considered a candidate gene for certain diseases based on chromosomal location and/or disease phenotype. We have employed 3{prime}-untranslated region RFLPs to exclude COL5A1 as a candidate gene in families with tuberous sclerosis 1, Ehlers-Danlos syndrome type H, and nail-patella syndrome. In addition, we describe a polymorphic simple sequence repeat (SSR) within a COL5A1 intron. This SSR is used to exclude COL5A1 as a candidate gene in hereditary hemorrhagic telangiectasia (Osler-Rendu-Weber disease) and to add COL5A1 to the existing map of {open_quotes}index{close_quotes} markers of chromosome 9 by evaluation of the COL5A1 locus on the CEPH 40-family reference pedigree set. This genetic mapping places COL5A1 between markers D9S66 and D9S67. 14 refs., 1 fig., 2 tabs.

  7. Correlation of regional breath sound with regional ventilation in emphysema

    SciTech Connect

    Ploysongsang, Y.; Pare, J.A.; Macklem, P.T.

    1982-09-01

    We measured regional breath sound intensities (Ib) by a microphone amplifier system in 8 subjects with emphysema. We also measured regional white noise transmissions (Tn) from the same areas in all subjects. The recorded areas were 5, 10, 15, and 20 cm from the apex of the lung just lateral to the right anterior midclavicular line. Xenon ventilation indexes (xenon tidal raw counts, an index of total regional ventilation; xenon equilibration raw counts, an index of ventilating lung volume; xenon ventilation per unit volume (Vr), an index of ventilation per unit volume) were also recorded from the same areas. The Ib, Tn, Ib/Tn (an index of sound generation), and xenon ventilation indexes were all expressed as a fraction of the mean value of all four recorded areas. The Ib and Ib/Tn correlated best with the xenon tidal raw counts, correlated well with the xenon equilibration raw counts, and correlated poorly with Vr. We conclude that Ib and Ib/Tn can be used to quantify regional ventilation in subjects with emphysema.

  8. Northeast Regional Biomass Energy Program

    SciTech Connect

    O'Connell, R.A.

    1992-04-01

    The Northeast Regional Biomass Program (NRBP) is entering its ninth year of operation. The management and the objectives have virtually remained unchanged and are stated as follows. The program conducted by NRBP has three basic features: (1) a state grant component that provides funds (with a 50 percent matching requirement) to each of the states in the region to strengthen and integrate the work of state agencies involved in biomass energy; (2) a series of technical reports and studies in areas that have been identified as being of critical importance to the development of biomass energy in the region; and (3) a continuous long range planning component with heavy private sector involvement that helps to identify activities necessary to spur greater development and use of biomass energy in the Northeast.

  9. Northeast Regional Biomass Energy Program

    SciTech Connect

    O'Connell, R.A.

    1992-02-01

    The Northeast Regional Biomass Program (NRBP) is entering its ninth year of operation. The management and the objectives have virtually remained unchanged and are stated as follows. The program conducted by NRBP has three basic features: (1) a state grant component that provides funds (with a 50 percent matching requirement) to each of the states in the region to strengthen and integrate the work of state agencies involved in biomass energy; (2) a series of technical reports and studies in areas that have been identified as being of critical importance to the development of biomass energy in the region; and (3) a continuous long range planning component with heavy private sector involvement that helps to identify activities necessary to spur greater development and use of biomass energy in the Northeast.

  10. Satellites monitor Atlanta regional development

    USGS Publications Warehouse

    Todd, William J.; Blackmon, C.C.; Rudasill, R.G.

    1979-01-01

    Since the adoption of a Regional Development Plan in 1975, the Atlanta Regional Commission has investigated methods for monitoring regional development patterns in a periodic, efficient manner. A promising approach appears to be the use of Landsat satellite data. In cooperation with the Earth Resources Observation Systems (EROS) Data Center, the commission used machine processing of digital temporal overlays of Landsat data collected in 1972, 1974 and 1976 to detect land use and land cover changes in the Atlanta metropolitan area. Results of the analysis revealed the conversion of forested and open space areas to residential, commercial and industrial land use in the urban-rural fringe zone from 1972 to 1974 and from 1974 to 1976. The study indicated that a land use and land cover change-detection program may be used to revise small-area forecasts of land use, population and employment made by planning models.

  11. Regional Cooperation to Strengthen Safeguards

    SciTech Connect

    Minnini, Margot; Elkhamri, Oksana O.

    2016-06-06

    President Obama’s decision over four years ago to ”pivot” toward Asia represented an important strategic shift in American foreign policy and a rebalancing of U.S. economic and security engagement in the Asia-Pacific countries. The United States has since supported a variety of regional initiatives aimed at promoting nuclear security and safeguards. When a new regional organization, the Asia-Pacific Safeguards Network (APSN) was established in 2010, DOE/NNSA became an early member and enthusiastic advocate. Launched on the initiative of Australia, Japan, Republic of Korea, and Indonesia, the APSN aims to strengthen the quality and effectiveness of safeguards implementation in the Asia-Pacific region.

  12. Regional sea level change in the Thailand-Indonesia region

    NASA Astrophysics Data System (ADS)

    Fenoglio-Marc, L.; Becker, M. H.; Buchhaupt, C.

    2013-12-01

    It is expected that the regional sea level rise will strongly affect particular regions with direct impacts including submergence of coastal zones, rising water tables and salt intrusion into groundwaters. It can possibly also exacerbate other factors as floodings, associated to storms and hurricanes, as well as ground subsidence of anthropogenic nature. The Thailand-Vietnam-Indonesian region is one of those zones. On land, the Chao-Praya and Mekong Delta are fertile alluvial zones. The potential for sea level increases and extreme floodings due to global warming makes the Deltas a place where local, regional, and global environmental changes are converging. We investigate the relative roles of regional and global mechanisms resulting in multidecadal variations and inflections in the rate of sea level change. Altimetry and GRACE data are used to investigate the variation of land floodings. The land surface water extent is evaluated at 25 km sampling intervals over fifteen years (1993-2007) using a multisatellite methodology which captures the extent of episodic and seasonal inundations, wetlands, rivers, lakes, and irrigated agriculture, using passive and active (microwaves and visible observations. The regional sea level change is analysed during the period 1993-2012 using satellite altimetry, wind and ocean model data, tide gauge data and GPS. The rates of absolute eustatic sea level rise derived from satellite altimetry through 19-year long precise altimeter observations are in average higher than the global mean rate. Several tide gauge records indicate an even higher sea level rise relative to land. We show that the sea level change is closely linked to the ENSO mode of variability and strongly affected by changes in wind forcing and ocean circulation. We have determined the vertical crustal motion at a given tide gauge location by differencing the tide gauge sea level time-series with an equivalent time-series derived from satellite altimetry and by computing

  13. Floristic study of Mirabad region.

    PubMed

    Hasanzadeh Gorttapeh, A; Panahy, J

    2007-10-15

    The study area (Mirabad) is located between 36 degrees, 55' to 37 degrees north latitude and 45 degrees, 05' to 55 degrees, 44' east longitude in west Azerbaijan province. In this study, Flora of this region was determined by using available references. We encountered 192 species that belongs to 126 genera and 41 families. The largest family of region is Asteraceae with 31 sp. and the largest Genera is Astragalus with 7 sp. The main life forms are: Hemicryptophyte with 30.2% and Therophyte with 28.1%. The most extended chorotype with 62.5% is related to: Irano_Turanian.

  14. Hydrodynamics of post CHF region

    SciTech Connect

    Ishii, M.; De Jarlais, G.

    1984-04-01

    Among various two-phase flow regimes, the inverted flow in the post-dryout region is relatively less well understood due to its special heat transfer conditions. The review of existing data indicates further research is needed in the areas of basic hydrodynamics related to liquid core disintegration mechanisms, slug and droplet formations, entrainment, and droplet size distributions. In view of this, the inverted flow is studied in detail both analytically and experimentally. Criteria for initial flow regimes in the post-dryout region are given. Preliminary models for subsequent flow regime transition criteria are derived together with correlations for a mean droplet diameter based on the adiabatic simulation data.

  15. Solitary neurilemmoma in postaural region.

    PubMed

    Adhikary, Bivas; Sen, Indranil; Basu, Asim Jiban; Bandyopadhyay, Saumyendra Nath; Saha, Debdas; Basu, Sumit Kumar

    2007-05-01

    Neurilemmoma in postaural region arising from great auricular nerve is an extremely rare tumour. An 11 years boy presented with pain and swelling behind his left ear for last 3-4 years. The clinical examination revealed the swelling appeared to be diffuse with the margin being ill defined. On radiological examination a diffuse homogeneous mass was seen in the postaural region of the left side. The tumour was completely removed by an incision through postaural route. Histopathological study revealed neurilemmoma. Postoperative period was uneventful.

  16. Regional High School Senior Survey.

    ERIC Educational Resources Information Center

    Day, Philip R., Jr.

    In order to identify the educational needs and aspirations of graduating high school seniors in the service region of the University of Maine at Augusta, a survey instrument was designed and administered to 1,950 seniors at 19 institutions. In all, 1,744 completed surveys were returned, a 92 percent response rate. The data are sub-grouped into…

  17. Microinstabilities in the pedestal region

    NASA Astrophysics Data System (ADS)

    Dickinson, David; Dudson, Benjamin; Wilson, Howard; Roach, Colin

    2014-10-01

    The regulation of transport at the pedestal top is important for the inter-ELM pedestal dynamics. Linear gyrokinetic analysis of the pedestal region during an ELM cycle on MAST has shown kinetic ballooning modes to be unstable at the knee of the pressure profile and in the steep pedestal region whilst microtearing modes (MTMs) dominate in the shallow gradient region inboard of the pedestal top. The transition between these instabilities at the pedestal knee has been observed in low and high collisionality MAST pedestals, and is likely to play an important role in the broadening of the pedestal. Nonlinear simulations are needed in this region to understand the microturbulence, the corresponding transport fluxes, and to gain further insight into the processes underlying the pedestal evolution. Such gyrokinetic simulations are numerically challenging and recent upgrades to the GS2 gyrokinetic code help improve their feasibility. We are also exploring reduced models that capture the relevant physics using the plasma simulation framework BOUT + + . An electromagnetic gyrofluid model has recently been implemented with BOUT + + that has significantly reduced computational cost compared to the gyrokinetic simulations against which it will be benchmarked. This work was funded by the RCUK Energy programme, EURATOM and a EUROFusion fellowship WP14-FRF-CCFE/Dickinson and was carried out using: HELIOS at IFERC, Japan; ARCHER (EPSRC Grant No. EP/L000237/1); HECToR (EPSRC Grant No. EP/H002081/1).

  18. Future and present regional wind

    NASA Astrophysics Data System (ADS)

    Salameh, T.; Vrac, M.; Drobinski, P.; Naveau, P.

    2009-04-01

    Accurate and rapid determination of near-surface wind fields in a complex area (orography, inhomogeneous surface properties) is a challenge for applications like the evaluation of wind energy production, the prediction of pollution transport and hazardous conditions for aeronautics and ship navigation, among others. This work presents a statistical downscaling approach for regional near-surface wind field in the region of southern France (characterized by the presence of major mountain ranges). It is based on generalized additive models (GAM, Salameh et al. 2008), relating large-scale upper air to local-scale surface atmospheric fields. We apply our statistical downscaling model conditionally on regional circulation patterns defined from measurements. Hence, near-surface wind components in southern France are simulated based on large-scale information from ERA-40 reanalyses (1991-2001) and from IPCC scenarios (1991-2001 and 2040-2050). The performances of our method are evaluated, (1) by comparing downscaled wind from ERA-40 and from IPCC, and (2) by comparing them with measurements, for the period 1991-2001. Then, we evaluate the change in regional atmospheric circulations in southern France, by comparing future and present downscaled wind.

  19. Biocapacity optimization in regional planning

    NASA Astrophysics Data System (ADS)

    Guo, Jianjun; Yue, Dongxia; Li, Kai; Hui, Cang

    2017-01-01

    Ecological overshoot has been accelerating across the globe. Optimizing biocapacity has become a key to resolve the overshoot of ecological demand in regional sustainable development. However, most literature has focused on reducing ecological footprint but ignores the potential of spatial optimization of biocapacity through regional planning of land use. Here we develop a spatial probability model and present four scenarios for optimizing biocapacity of a river basin in Northwest China. The potential of enhanced biocapacity and its effects on ecological overshoot and water consumption in the region were explored. Two scenarios with no restrictions on croplands and water use reduced the overshoot by 29 to 53%, and another two scenarios which do not allow croplands and water use to increase worsened the overshoot by 11 to 15%. More spatially flexible transition rules of land use led to higher magnitude of change after optimization. However, biocapacity optimization required a large amount of additional water resources, casting considerable pressure on the already water-scarce socio-ecological system. Our results highlight the potential for policy makers to manage/optimize regional land use which addresses ecological overshoot. Investigation on the feasibility of such spatial optimization complies with the forward-looking policies for sustainable development and deserves further attention.

  20. Regional Security Application and Checkmate!

    DTIC Science & Technology

    2000-04-01

    30 F. Time, Geo- Spatial , and Resource Constraints ................................................ 32 G. Summary...leverage human interactions and computer modeling and tools to provide participants with useful insights on complex multi -disciplinary issues. It’s...and interdependent multi -disciplinary issues associated with such challenges. In this regard: (a) A regional S.E.N.S.E. application that addresses

  1. Biocapacity optimization in regional planning

    PubMed Central

    Guo, Jianjun; Yue, Dongxia; Li, Kai; Hui, Cang

    2017-01-01

    Ecological overshoot has been accelerating across the globe. Optimizing biocapacity has become a key to resolve the overshoot of ecological demand in regional sustainable development. However, most literature has focused on reducing ecological footprint but ignores the potential of spatial optimization of biocapacity through regional planning of land use. Here we develop a spatial probability model and present four scenarios for optimizing biocapacity of a river basin in Northwest China. The potential of enhanced biocapacity and its effects on ecological overshoot and water consumption in the region were explored. Two scenarios with no restrictions on croplands and water use reduced the overshoot by 29 to 53%, and another two scenarios which do not allow croplands and water use to increase worsened the overshoot by 11 to 15%. More spatially flexible transition rules of land use led to higher magnitude of change after optimization. However, biocapacity optimization required a large amount of additional water resources, casting considerable pressure on the already water-scarce socio-ecological system. Our results highlight the potential for policy makers to manage/optimize regional land use which addresses ecological overshoot. Investigation on the feasibility of such spatial optimization complies with the forward-looking policies for sustainable development and deserves further attention. PMID:28112224

  2. SDO Sees Active Region Outbursts

    NASA Video Gallery

    This close up video by NASA’s Solar Dynamics Observatory shows an active region near the right-hand edge of the sun’s disk, which erupted with at least a dozen minor events over a 30-hour period fr...

  3. PREL Pacific Region Language Cards.

    ERIC Educational Resources Information Center

    Pacific Region Educational Lab., Honolulu, HI.

    This collection of 10 cue cards presents English translations of common English words and expressions into 10 Pacific Region languages: Palauan, Samoan, Chamorro, Hawaiian, Carolinian, Chuukese, Pohnpeian, Marshallese, Yapese, and Kosraean. The cards translate the following: hello, good morning, good afternoon, good night, thank you, you're…

  4. Education and Italian Regional Development

    ERIC Educational Resources Information Center

    Di Liberto, Adriana

    2008-01-01

    In this paper, we study the connection between growth and human capital in a convergence regression for the panel of Italian regions. We include measures of average primary, secondary and tertiary education. We find that increased education seems to contribute to growth only in the South. Decomposing total schooling into its three constituent…

  5. Regional Needs Analysis Report. 2011

    ERIC Educational Resources Information Center

    Washington Higher Education Coordinating Board, 2011

    2011-01-01

    The Higher Education Coordinating Board (HECB) is required to develop a comprehensive and ongoing needs assessment process to analyze demand for additional degrees and programs [RCW 28B.76.230 (1)]. This report fulfills a portion of that mandate by focusing on employer demand on the regional level, but also includes additional information on…

  6. MC-1 Mare Boreum Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-1 quadrangle, Mare Boreum region of Mars. The central part is covered by a residual ice cap that is cut by spiral-patterned troughs exposing layered terrain. The cap is surrounded by broad flat plains and large dune fields. Latitude range 65 to 90, longitude range -180 to 180.

  7. MC-5 Ismenius Lacus Region

    NASA Technical Reports Server (NTRS)

    1994-01-01

    Mars digital-image mosaic merged with color of the MC-5 quadrangle, Ismenius Lacus region of Mars. Heavily cratered highlands of the southern part are separated from the relatively smooth plains of the northern part by a belt of dissected terrain, containing mesas and buttes. Latitude range 30 to 65 degrees, longitude range -60 to 0 degrees.

  8. Regional Expertise and Culture Proficiency

    DTIC Science & Technology

    2012-09-01

    for culture and language training and education Abbe (2008) Social initiative Willingness to communicate in cross-cultural settings; interest in... intercultural skills during officer formal education Montgomery, AL: Air War College, Air University. Russell et al. (1995, July). Intercultural communication ...JPEC) community with foreign language and regional expertise capabilities, integrate the capabilities into all force planning activities, and obtain

  9. 32 CFR 1605.7 - Region Manager.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 32 National Defense 6 2010-07-01 2010-07-01 false Region Manager. 1605.7 Section 1605.7 National... ORGANIZATION Region Administration § 1605.7 Region Manager. (a) Subject to the direction and control of the Director of Selective Service, the Region Manager of Selective Service for each region shall be...

  10. Algorithms for Labeling Focus Regions.

    PubMed

    Fink, M; Haunert, Jan-Henrik; Schulz, A; Spoerhase, J; Wolff, A

    2012-12-01

    In this paper, we investigate the problem of labeling point sites in focus regions of maps or diagrams. This problem occurs, for example, when the user of a mapping service wants to see the names of restaurants or other POIs in a crowded downtown area but keep the overview over a larger area. Our approach is to place the labels at the boundary of the focus region and connect each site with its label by a linear connection, which is called a leader. In this way, we move labels from the focus region to the less valuable context region surrounding it. In order to make the leader layout well readable, we present algorithms that rule out crossings between leaders and optimize other characteristics such as total leader length and distance between labels. This yields a new variant of the boundary labeling problem, which has been studied in the literature. Other than in traditional boundary labeling, where leaders are usually schematized polylines, we focus on leaders that are either straight-line segments or Bezier curves. Further, we present algorithms that, given the sites, find a position of the focus region that optimizes the above characteristics. We also consider a variant of the problem where we have more sites than space for labels. In this situation, we assume that the sites are prioritized by the user. Alternatively, we take a new facility-location perspective which yields a clustering of the sites. We label one representative of each cluster. If the user wishes, we apply our approach to the sites within a cluster, giving details on demand.

  11. Genetic Analysis of the Pestivirus Nonstructural Coding Region: Defects in the NS5A Unit Can Be Complemented in trans

    PubMed Central

    Grassmann, Claus W.; Isken, Olaf; Tautz, Norbert; Behrens, Sven-Erik

    2001-01-01

    The functional analysis of molecular determinants which control the replication of pestiviruses was considerably facilitated by the finding that subgenomic forms of the positive-strand RNA genome of BVDV (bovine viral diarrhea virus) are capable of autonomous replication in transfected host cells. The prototype replicon, BVDV DI9c, consists of the genomic 5′ and 3′ untranslated regions and a truncated open reading frame (ORF) encoding mainly the nonstructural proteins NS3, NS4A, NS4B, NS5A, and NS5B. To gain insight into which of these proteins are essential for viral replication and whether they act in cis or in trans, we introduced a large spectrum of in-frame mutations into the DI9c ORF. Tests of the mutant RNAs in terms of their replication capacity and their ability to support translation and cleavage of the nonstructural polyprotein, and whether defects could be rescued in trans, yielded the following results. (i) RNA replication was found to be dependent on the expression of each of the DI9c-encoded mature proteins NS3 to NS5B (and the known associated enzymatic activities). In the same context, a finely balanced molar ratio of the diverse proteolytic processing products was indicated to be crucial for the formation of an active catalytic replication complex. (ii) Synthesis of negative-strand intermediate and progeny positive-strand RNA was observed to be strictly coupled with all functional DI9c ORF derivatives. NS3 to NS5B were hence suggested to play a pivotal role even during early steps of the viral replication pathway. (iii) Mutations in the NS3 and NS4B units which generated nonfunctional or less functional RNAs were determined to be cis dominant. Likewise, lethal alterations in the NS4A and NS5B regions were invariably noncomplementable. (iv) In surprising contrast, replication of functional and nonfunctional NS5A mutants could be clearly enhanced and restored, respectively. In summary, our data provide initial insights into the organization of

  12. Regional Sediment Budgets for the Haleiwa Region, Oahu, Hawaii

    DTIC Science & Technology

    2014-06-01

    Hawaii by Jessica H. Podoski PURPOSE: This Coastal and Hydraulics Engineering Technical Note (CHETN) reviews the development of conceptual...morphology and coastal processes have on sediment pathways and transport volumes. In the Southeast Oahu Region, Mokapu Point to Makapuu Point RSB...and Development Center, Coastal and Hydraulics Laboratory,3909 Halls Ferry Road,Vicksburg,MS,39180-6199 8. PERFORMING ORGANIZATION REPORT NUMBER 9

  13. Numerical Simulation of Regional Circulation in the Monterey Bay Region

    NASA Technical Reports Server (NTRS)

    Tseng, Y. H.; Dietrich, D. E.; Ferziger, J. H.

    2003-01-01

    The objective of this study is to produce a high-resolution numerical model of Mon- terey Bay area in which the dynamics are determined by the complex geometry of the coastline, steep bathymetry, and the in uence of the water masses that constitute the CCS. Our goal is to simulate the regional-scale ocean response with realistic dynamics (annual cycle), forcing, and domain. In particular, we focus on non-hydrostatic e ects (by comparing the results of hydrostatic and non-hydrostatic models) and the role of complex geometry, i.e. the bay and submarine canyon, on the nearshore circulation. To the best of our knowledge, the current study is the rst to simulate the regional circulation in the vicinity of Monterey Bay using a non-hydrostatic model. Section 2 introduces the high resolution Monterey Bay area regional model (MBARM). Section 3 provides the results and veri cation with mooring and satellite data. Section 4 compares the results of hydrostatic and non-hydrostatic models.

  14. [Regional clinical audit, guideline targets, and local and regional benchmarks].

    PubMed

    Casino, F G; Lopez, T

    2005-01-01

    Regional clinical Audit, guideline Targets and local and regional Benchmarks In order to improve the quality of dialysis treatment, we have devised some routines, particularly suitable for electronic data management systems. First, we suggest a systematic monthly analysis of 10 common clinical performance measures (CPM), with the following guideline based targets: predialysis systolic blood pressure (SBP) < 140 mmHg; session length >/= 240 min; dialysis dose (spKt/V) >/=1.3; normalized protein catabolic rate (NPCR) >/=1.2 g/kg/d; hemoglobin (Hb) >/=11 g/dL; serum calcium (Ca) 8.4-9.5 mg/dL; serum phosphorus (P) 3.5-5.5 mg/dL; Ca x P /=20 mmol/L; serum potassium (K) 3.5-6.0 mmol/L. The Hb target should be reached in at least 85% of all maintenance hemodialysis (HD) patients in the unit; for all other targets, an arbitrary >/=80% is proposed. Since the above percentages are quite difficult to reach on a short-term basis, an intermediate local or regional standard (benchmark) could be devised as an average of the percentage of patients who actually reach the targets for each CPM at any dialysis unit in a given regional area; and therefore, from truly comparable patients. As an example, we simulated a regional audit by using the above targets with available data from 398 patients from southern Italy. A further step in this process was to find the cause(s) of failure in each patient who did not reach the targets. To this end, we suggest a systematic search of the well-known factors that could affect each CPM, for each failed patient. As an example, we screened all patients with Hb < 11 g/dL at a single unit, to establish the presence/absence of any common cause associated with inadequate response to epoetin treatment. Moreover, by using criteria for prescribing iron therapy or increasing epoetin dose, we found that some patients did not receive the appropriate therapy after blood sampling results. To avoid this possible

  15. Enhanced Salt Tolerance Conferred by the Complete 2.3 kb cDNA of the Rice Vacuolar Na(+)/H(+) Antiporter Gene Compared to 1.9 kb Coding Region with 5' UTR in Transgenic Lines of Rice.

    PubMed

    Amin, U S M; Biswas, Sudip; Elias, Sabrina M; Razzaque, Samsad; Haque, Taslima; Malo, Richard; Seraj, Zeba I

    2016-01-01

    Soil salinity is one of the most challenging problems that restricts the normal growth and production of rice worldwide. It has therefore become very important to produce more saline tolerant rice varieties. This study shows constitutive over-expression of the vacuolar Na(+)/H(+) antiporter gene (OsNHX1) from the rice landrace (Pokkali) and attainment of enhanced level of salinity tolerance in transgenic rice plants. It also shows that inclusion of the complete un-translated regions (UTRs) of the alternatively spliced OsNHX1 gene provides a higher level of tolerance to the transgenic rice. Two separate transformation events of the OsNHX1 gene, one with 1.9 kb region containing the 5' UTR with CDS and the other of 2.3 kb, including 5' UTR, CDS, and the 3' UTR regions were performed. The transgenic plants with these two different constructs were advanced to the T3 generation and physiological and molecular screening of homozygous plants was conducted at seedling and reproductive stages under salinity (NaCl) stress. Both transgenic lines were observed to be tolerant compared to WT plants at both physiological stages. However, the transgenic lines containing the CDS with both the 5' and 3' UTR were significantly more tolerant compared to the transgenic lines containing OsNHX1 gene without the 3' UTR. At the seedling stage at 12 dS/m stress, the chlorophyll content was significantly higher (P < 0.05) and the electrolyte leakage significantly lower (P < 0.05) in the order 2.3 kb > 1.9 kb > and WT lines. Yield in g/plant in the best line from the 2.3 kb plants was significantly more (P < 0.01) compared, respectively, to the best 1.9 kb line and WT plants at stress of 6 dS/m. Transformation with the complete transcripts rather than the CDS may therefore provide more durable level of tolerance.

  16. Complete nucleotide sequence and genome structure of a Japanese isolate of hibiscus latent Fort Pierce virus, a unique tobamovirus that contains an internal poly(A) region in its 3' end.

    PubMed

    Yoshida, Tetsuya; Kitazawa, Yugo; Komatsu, Ken; Neriya, Yutaro; Ishikawa, Kazuya; Fujita, Naoko; Hashimoto, Masayoshi; Maejima, Kensaku; Yamaji, Yasuyuki; Namba, Shigetou

    2014-11-01

    In this study, we detected a Japanese isolate of hibiscus latent Fort Pierce virus (HLFPV-J), a member of the genus Tobamovirus, in a hibiscus plant in Japan and determined the complete sequence and organization of its genome. HLFPV-J has four open reading frames (ORFs), each of which shares more than 98 % nucleotide sequence identity with those of other HLFPV isolates. Moreover, HLFPV-J contains a unique internal poly(A) region of variable length, ranging from 44 to 78 nucleotides, in its 3'-untranslated region (UTR), as is the case with hibiscus latent Singapore virus (HLSV), another hibiscus-infecting tobamovirus. The length of the HLFPV-J genome was 6431 nucleotides, including the shortest internal poly(A) region. The sequence identities of ORFs 1, 2, 3 and 4 of HLFPV-J to other tobamoviruses were 46.6-68.7, 49.9-70.8, 31.0-70.8 and 39.4-70.1 %, respectively, at the nucleotide level and 39.8-75.0, 43.6-77.8, 19.2-70.4 and 31.2-74.2 %, respectively, at the amino acid level. The 5'- and 3'-UTRs of HLFPV-J showed 24.3-58.6 and 13.0-79.8 % identity, respectively, to other tobamoviruses. In particular, when compared to other tobamoviruses, each ORF and UTR of HLFPV-J showed the highest sequence identity to those of HLSV. Phylogenetic analysis showed that HLFPV-J, other HLFPV isolates and HLSV constitute a malvaceous-plant-infecting tobamovirus cluster. These results indicate that the genomic structure of HLFPV-J has unique features similar to those of HLSV. To our knowledge, this is the first report of the complete genome sequence of HLFPV.

  17. Sulfur-Responsive Elements in the 3′-Nontranscribed Intergenic Region Are Essential for the Induction of SULFATE TRANSPORTER 2;1 Gene Expression in Arabidopsis Roots under Sulfur Deficiency

    PubMed Central

    Maruyama-Nakashita, Akiko; Watanabe-Takahashi, Akiko; Inoue, Eri; Yamaya, Tomoyuki; Saito, Kazuki; Takahashi, Hideki

    2015-01-01

    Under sulfur deficiency (−S), plants induce expression of the sulfate transport systems in roots to increase uptake and root-to-shoot transport of sulfate. The low-affinity sulfate transporter SULTR2;1 is predominantly expressed in xylem parenchyma and pericycle cells in Arabidopsis thaliana roots under –S. The mechanisms underlying –S-inducible expression of SULTR2;1 in roots have remained unclear, despite the possible significance of SULTR2;1 for acclimation to low-sulfur conditions. In this investigation, examination of deletions and base substitutions in the 3′-intergenic region of SULTR2;1 revealed novel sulfur-responsive elements, SURE21A (5′-CAATGTATC-3′) and SURE21B (5′-CTAGTAC-3′), located downstream of the SULTR2;1 3′-untranslated region. SURE21A and SULTR21B effectively induced reporter gene expression from fusion constructs under –S in combination with minimal promoters or promoters not inducible by –S, suggesting their versatility in controlling transcription. T-DNA insertions near SURE21A and SULTR21B abolished −S-inducible expression of SULTR2;1 in roots and reduced the uptake and root-to-shoot transport of sulfate. In addition, these mutations partially suppressed SULTR2;1 expression in shoots, without changing its –S-responsive expression. These findings indicate that SULTR2;1 contributes to the increase in uptake and internal translocation of sulfate driven by gene expression induced under the control of sulfur-responsive elements in the 3′-nontranscribed intergenic region of SULTR2;1. PMID:25855406

  18. Marine cloud brightening: regional applications.

    PubMed

    Latham, John; Gadian, Alan; Fournier, Jim; Parkes, Ben; Wadhams, Peter; Chen, Jack

    2014-12-28

    The general principle behind the marine cloud brightening (MCB) climate engineering technique is that seeding marine stratocumulus clouds with substantial concentrations of roughly monodisperse sub-micrometre-sized seawater particles might significantly enhance cloud albedo and longevity, thereby producing a cooling effect. This paper is concerned with preliminary studies of the possible beneficial application of MCB to three regional issues: (1) recovery of polar ice loss, (2) weakening of developing hurricanes and (3) elimination or reduction of coral bleaching. The primary focus is on Item 1. We focus discussion herein on advantages associated with engaging in limited-area seeding, regional effects rather than global; and the levels of seeding that may be required to address changing current and near-term conditions in the Arctic. We also mention the possibility that MCB might be capable of producing a localized cooling to help stabilize the West Antarctic Ice Sheet.

  19. Marine cloud brightening: regional applications

    PubMed Central

    Latham, John; Gadian, Alan; Fournier, Jim; Parkes, Ben; Wadhams, Peter; Chen, Jack

    2014-01-01

    The general principle behind the marine cloud brightening (MCB) climate engineering technique is that seeding marine stratocumulus clouds with substantial concentrations of roughly monodisperse sub-micrometre-sized seawater particles might significantly enhance cloud albedo and longevity, thereby producing a cooling effect. This paper is concerned with preliminary studies of the possible beneficial application of MCB to three regional issues: (1) recovery of polar ice loss, (2) weakening of developing hurricanes and (3) elimination or reduction of coral bleaching. The primary focus is on Item 1. We focus discussion herein on advantages associated with engaging in limited-area seeding, regional effects rather than global; and the levels of seeding that may be required to address changing current and near-term conditions in the Arctic. We also mention the possibility that MCB might be capable of producing a localized cooling to help stabilize the West Antarctic Ice Sheet. PMID:25404682

  20. [Health in Andean regional integration].

    PubMed

    Agudelo, Carlos A

    2007-01-01

    Despite their shared history, the Andean countries are socially and politically diverse, with heterogeneous health realities and complex integration processes. General developments such as the Latin American Free Trade Association and Latin American Integration Association have existed for decades, along with others of a regional scope, like the Andean Community of Nations, Caribbean Community, and Central American Common Market. The health field has a specific instrument in the Andean Region called the Hipólito Unánue Agreement, created in 1971. Integration processes have concentrated on economic aspects, based on preferential customs agreements that have led to an important long-term increase in trade. Less progress has been made in the field of health in terms of sharing national experiences, knowledge, and capabilities. Analysis of experiences in health has shown that integration depends on the countries' respective strengths and to a major extent on national political processes.