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Sample records for 17s u2 snrnp

  1. An siRNA Screen Identifies the U2 snRNP Spliceosome as a Host Restriction Factor for Recombinant Adeno-associated Viruses

    PubMed Central

    Schreiber, Claire A.; Sakuma, Toshie; Izumiya, Yoshihiro; Holditch, Sara J.; Hickey, Raymond D.; Bressin, Robert K.; Basu, Upamanyu; Koide, Kazunori; Asokan, Aravind; Ikeda, Yasuhiro

    2015-01-01

    Adeno-associated viruses (AAV) have evolved to exploit the dynamic reorganization of host cell machinery during co-infection by adenoviruses and other helper viruses. In the absence of helper viruses, host factors such as the proteasome and DNA damage response machinery have been shown to effectively inhibit AAV transduction by restricting processes ranging from nuclear entry to second-strand DNA synthesis. To identify host factors that might affect other key steps in AAV infection, we screened an siRNA library that revealed several candidate genes including the PHD finger-like domain protein 5A (PHF5A), a U2 snRNP-associated protein. Disruption of PHF5A expression selectively enhanced transgene expression from AAV by increasing transcript levels and appears to influence a step after second-strand synthesis in a serotype and cell type-independent manner. Genetic disruption of U2 snRNP and associated proteins, such as SF3B1 and U2AF1, also increased expression from AAV vector, suggesting the critical role of U2 snRNP spliceosome complex in this host-mediated restriction. Notably, adenoviral co-infection and U2 snRNP inhibition appeared to target a common pathway in increasing expression from AAV vectors. Moreover, pharmacological inhibition of U2 snRNP by meayamycin B, a potent SF3B1 inhibitor, substantially enhanced AAV vector transduction of clinically relevant cell types. Further analysis suggested that U2 snRNP proteins suppress AAV vector transgene expression through direct recognition of intact AAV capsids. In summary, we identify U2 snRNP and associated splicing factors, which are known to be affected during adenoviral infection, as novel host restriction factors that effectively limit AAV transgene expression. Concurrently, we postulate that pharmacological/genetic manipulation of components of the spliceosomal machinery might enable more effective gene transfer modalities with recombinant AAV vectors. PMID:26244496

  2. 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.

  3. Cwc24p Is a General Saccharomyces cerevisiae Splicing Factor Required for the Stable U2 snRNP Binding to Primary Transcripts

    PubMed Central

    Coltri, Patricia P.; Oliveira, Carla C.

    2012-01-01

    Splicing of primary transcripts is an essential process for the control of gene expression. Specific conserved sequences in premature transcripts are important to recruit the spliceosome machinery. The Saccharomyces cerevisiae catalytic spliceosome is composed of about 60 proteins and 5 snRNAs (U1, U2, U4/U6 and U5). Among these proteins, there are core components and regulatory factors, which might stabilize or facilitate splicing of specific substrates. Assembly of a catalytic complex depends on the dynamics of interactions between these proteins and RNAs. Cwc24p is an essential S. cerevisiae protein, originally identified as a component of the NTC complex, and later shown to affect splicing in vivo. In this work, we show that Cwc24p also affects splicing in vitro. We show that Cwc24p is important for the U2 snRNP binding to primary transcripts, co-migrates with spliceosomes, and that it interacts with Brr2p. Additionally, we show that Cwc24p is important for the stable binding of Prp19p to the spliceosome. We propose a model in which Cwc24p is required for stabilizing the U2 association with primary transcripts, and therefore, especially important for splicing of RNAs containing non-consensus branchpoint sequences. PMID:23029180

  4. Mammalian splicing factor SF1 interacts with SURP domains of U2 snRNP-associated proteins.

    PubMed

    Crisci, Angela; Raleff, Flore; Bagdiul, Ivona; Raabe, Monika; Urlaub, Henning; Rain, Jean-Christophe; Krämer, Angela

    2015-12-01

    Splicing factor 1 (SF1) recognizes the branch point sequence (BPS) at the 3' splice site during the formation of early complex E, thereby pre-bulging the BPS adenosine, thought to facilitate subsequent base-pairing of the U2 snRNA with the BPS. The 65-kDa subunit of U2 snRNP auxiliary factor (U2AF65) interacts with SF1 and was shown to recruit the U2 snRNP to the spliceosome. Co-immunoprecipitation experiments of SF1-interacting proteins from HeLa cell extracts shown here are consistent with the presence of SF1 in early splicing complexes. Surprisingly almost all U2 snRNP proteins were found associated with SF1. Yeast two-hybrid screens identified two SURP domain-containing U2 snRNP proteins as partners of SF1. A short, evolutionarily conserved region of SF1 interacts with the SURP domains, stressing their role in protein-protein interactions. A reduction of A complex formation in SF1-depleted extracts could be rescued with recombinant SF1 containing the SURP-interaction domain, but only partial rescue was observed with SF1 lacking this sequence. Thus, SF1 can initially recruit the U2 snRNP to the spliceosome during E complex formation, whereas U2AF65 may stabilize the association of the U2 snRNP with the spliceosome at later times. In addition, these findings may have implications for alternative splicing decisions. PMID:26420826

  5. Mammalian splicing factor SF1 interacts with SURP domains of U2 snRNP-associated proteins.

    PubMed

    Crisci, Angela; Raleff, Flore; Bagdiul, Ivona; Raabe, Monika; Urlaub, Henning; Rain, Jean-Christophe; Krämer, Angela

    2015-12-01

    Splicing factor 1 (SF1) recognizes the branch point sequence (BPS) at the 3' splice site during the formation of early complex E, thereby pre-bulging the BPS adenosine, thought to facilitate subsequent base-pairing of the U2 snRNA with the BPS. The 65-kDa subunit of U2 snRNP auxiliary factor (U2AF65) interacts with SF1 and was shown to recruit the U2 snRNP to the spliceosome. Co-immunoprecipitation experiments of SF1-interacting proteins from HeLa cell extracts shown here are consistent with the presence of SF1 in early splicing complexes. Surprisingly almost all U2 snRNP proteins were found associated with SF1. Yeast two-hybrid screens identified two SURP domain-containing U2 snRNP proteins as partners of SF1. A short, evolutionarily conserved region of SF1 interacts with the SURP domains, stressing their role in protein-protein interactions. A reduction of A complex formation in SF1-depleted extracts could be rescued with recombinant SF1 containing the SURP-interaction domain, but only partial rescue was observed with SF1 lacking this sequence. Thus, SF1 can initially recruit the U2 snRNP to the spliceosome during E complex formation, whereas U2AF65 may stabilize the association of the U2 snRNP with the spliceosome at later times. In addition, these findings may have implications for alternative splicing decisions.

  6. Mapping of epitopes on U1 snRNP polypeptide A with synthetic peptides and autoimmune sera.

    PubMed Central

    Barakat, S; Briand, J P; Abuaf, N; van Regenmortel, M H; Muller, S

    1991-01-01

    The ability of synthetic peptides encompassing almost the entire sequence of snRNP U1A polypeptide to be recognized in ELISA by sera of autoimmune patients was investigated. Sera from 18 patients with mixed connective tissue disease (MCTD), 145 with systemic lupus erythematosus (SLE) and 120 with other rheumatic autoimmune diseases were tested with 13 overlapping peptides. Among them, peptide 257-282 and, to a lower extent, peptide 1-11 were recognized by MCTD, SLE and Sjögren's syndrome sera. In contrast, peptide 35-58 was recognized by 94% of MCTD and only 19% of SLE sera. It did not react with any of the other patient sera. The ELISA results were compared with the pattern of reactivity observed in immunoblotting. The results indicate that peptide 35-58 probably contains a major epitope recognized by MCTD autoantibodies. It is noteworthy that in snRNP particles, this region of U1A interacts with RNA and presents only limited homology with the corresponding sequence 32-50 of U2B''. Images Fig. 3 PMID:1717192

  7. Spliceosome SNRNP200 Promotes Viral RNA Sensing and IRF3 Activation of Antiviral Response

    PubMed Central

    Tremblay, Nicolas; Baril, Martin; Chatel-Chaix, Laurent; Es-Saad, Salwa; Park, Alex Young; Koenekoop, Robert K.; Lamarre, Daniel

    2016-01-01

    Spliceosomal SNRNP200 is a Ski2-like RNA helicase that is associated with retinitis pigmentosa 33 (RP33). Here we found that SNRNP200 promotes viral RNA sensing and IRF3 activation through the ability of its amino-terminal Sec63 domain (Sec63-1) to bind RNA and to interact with TBK1. We show that SNRNP200 relocalizes into TBK1-containing cytoplasmic structures upon infection, in contrast to the RP33-associated S1087L mutant, which is also unable to rescue antiviral response of SNRNP200 knockdown cells. This functional rescue correlates with the Sec63-1-mediated binding of viral RNA. The hindered IFN-β production of knockdown cells was further confirmed in peripheral blood cells of RP33 patients bearing missense mutation in SNRNP200 upon infection with Sendai virus (SeV). This work identifies a novel immunoregulatory role of the spliceosomal SNRNP200 helicase as an RNA sensor and TBK1 adaptor for the activation of IRF3-mediated antiviral innate response. PMID:27454487

  8. An unusual chemical reactivity of Sm site adenosines strongly correlates with proper assembly of core U snRNP particles.

    PubMed

    Hartmuth, K; Raker, V A; Huber, J; Branlant, C; Lührmann, R

    1999-01-01

    The small nuclear ribonucleoprotein particles (snRNP) U1, U2, U4, and U5 contain a common set of eight Sm proteins that bind to the conserved single-stranded 5'-PuAU3-6GPu-3' (Sm binding site) region of their constituent U snRNA (small nuclear RNA), forming the Sm core RNP. Using native and in vitro reconstituted U1 snRNPs, accessibility of the RNA within the Sm core RNP to chemical structure probes was analyzed. Hydroxyl radical footprinting of in vitro reconstituted U1 snRNP demonstrated that riboses within a large continuous RNA region, including the Sm binding site, were protected. This protection was dependent on the binding of the Sm proteins. In contrast with the riboses, the phosphate groups within the Sm core site were accessible to modifying reagents. The invariant adenosine residue at the 5' end, as well as an adenosine two nucleotides downstream of the Sm binding site, showed an unexpected reactivity with dimethyl sulfate. This novel reactivity could be attributed to N7-methylation of the adenosine and was not observed in naked RNA, indicating that it is an intrinsic property of the RNA- protein interactions within the Sm core RNP. Further, this reactivity was observed concomitantly with formation of the Sm subcore intermediate during Sm core RNP assembly. As the Sm subcore can be viewed as the commitment complex in this assembly pathway, these results suggest that the peculiar reactivity of the Sm site adenosine bases may be diagnostic for proper assembly of the Sm core RNP. Consistent with this idea, a strong correlation was found between the unusual N7-A methylation sensitivity of the Sm core RNP and its ability to be imported into the nucleus of Xenopus laevis oocytes. PMID:9878394

  9. Antibodies specific for N6-methyladenosine react with intact snRNPs U2 and U4/U6.

    PubMed

    Bringmann, P; Lührmann, R

    1987-03-23

    Antibodies specific for N6-methyladenosine (m6A) were elicited in rabbits and used to study the accessibility in intact snRNPs of the m6A residues present in the snRNAs U2, U4 and U6. The antibody quantitatively precipitates snRNPs U2 and U4/U6 from total nucleoplasmic snRNPs U1-U6 isolated from HeLa cells, which demonstrates that the m6A residues of the respective snRNAs are not protected by snRNP proteins in the snRNP particles. While the anti-m6A IgG does not react at all with U5 RNPs lacking m6A, a significant amount of U1 RNPs was co-precipitated despite the fact that U1 RNA does not contain m6A either. Since anti-m6A IgG does not react with purified U1 RNPs and co-precipitation of U1 RNPs is dependent on the presence of U2 RNPs but not of U4/U6 RNPs, these data indicate an interaction between snRNPs U1 and U2 in vitro. The anti-m6A precipitation pattern described above was also observed with snRNPs isolation from mouse Ehrlich ascites tumor cells, indicating similar three-dimensional arrangements of snRNAs in homologous snRNP particles from different organisms. PMID:2951275

  10. Sm protein methylation is dispensable for snRNP assembly in Drosophila melanogaster

    PubMed Central

    Gonsalvez, Graydon B.; Praveen, Kavita; Hicks, Amanda J.; Tian, Liping; Matera, A. Gregory

    2008-01-01

    Sm proteins form stable ribonucleoprotein (RNP) complexes with small nuclear (sn)RNAs and are core components of the eukaryotic spliceosome. In vivo, the assembly of Sm proteins onto snRNAs requires the survival motor neurons (SMN) complex. Several reports have shown that SMN protein binds with high affinity to symmetric dimethylarginine (sDMA) residues present on the C-terminal tails of SmB, SmD1, and SmD3. This post-translational modification is thought to play a crucial role in snRNP assembly. In human cells, two distinct protein arginine methyltransferases (PRMT5 and PRMT7) are required for snRNP biogenesis. However, in Drosophila, loss of Dart5 (the fruit fly PRMT5 ortholog) has little effect on snRNP assembly, and homozygous mutants are completely viable. To resolve these apparent differences, we examined this topic in detail and found that Drosophila Sm proteins are also methylated by two methyltransferases, Dart5/PRMT5 and Dart7/PRMT7. Unlike dart5, we found that dart7 is an essential gene. However, the lethality associated with loss of Dart7 protein is apparently unrelated to defects in snRNP assembly. To conclusively test the requirement for sDMA modification of Sm proteins in Drosophila snRNP assembly, we constructed a fly strain that exclusively expresses an isoform of SmD1 that cannot be sDMA modified. Interestingly, these flies were viable, and snRNP assays revealed no defects in comparison to wild type. In contrast, dart5 mutants displayed a strong synthetic lethal phenotype in the presence of a hypomorphic Smn mutation. We therefore conclude that dart5 is required for viability when SMN is limiting. PMID:18369183

  11. Internal Modification of U2 Small Nuclear (Snrna) Occurs in Nucleoli of Xenopus Oocytes

    PubMed Central

    Yu, Yi-Tao; Shu, Mei-Di; Narayanan, Aarthi; Terns, Rebecca M.; Terns, Michael P.; Steitz, Joan A.

    2001-01-01

    U2 small nuclear (sn)RNA contains a large number of posttranscriptionally modified nucleotides, including a 5′ trimethylated guanosine cap, 13 pseudouridines, and 10 2′-O-methylated residues. Using Xenopus oocytes, we demonstrated previously that at least some of these modified nucleotides are essential for biogenesis of a functional snRNP. Here we address the subcellular site of U2 internal modification. Upon injection into the cytoplasm of oocytes, G-capped U2 that is transported to the nucleus becomes modified, whereas A-capped U2 that remains in the cytoplasm is not modified. Furthermore, by injecting U2 RNA into isolated nuclei or enucleated oocytes, we observe that U2 internal modifications occur exclusively in the nucleus. Analysis of the intranuclear localization of fluorescently labeled RNAs shows that injected wild-type U2 becomes localized to nucleoli and Cajal bodies. Both internal modification and nucleolar localization of U2 are dependent on the Sm binding site. An Sm-mutant U2 is targeted only to Cajal bodies. The Sm binding site can be replaced by a nucleolar localization signal derived from small nucleolar RNAs (the box C/D motif), resulting in rescue of internal modification as well as nucleolar localization. Analysis of additional chimeric U2 RNAs reveals a correlation between internal modification and nucleolar localization. Together, our results suggest that U2 internal modification occurs within the nucleolus. PMID:11257127

  12. A physical map encompassing GP2B, EPB3, D17S183, D17S78, D17S1183, and D17S1184

    SciTech Connect

    Miki, Y.; Swensen, J.J.; Neuhausen, S.L.

    1995-01-01

    The q21 region of chromosome 17 contains the gene BRCA1, which is involved in familial early-onset breast and ovarian cancers. A physical map of a region that extends from a distal boundary of the BRCA1 region, D17S78, to GP2B has been constructed. The map consists of 30 STSs, including 2 new short tandem repeat polymorphic markers. The contig is composed of a mixture of 7 YACs, 5 P1 plasmids, and 14 cosmids and was ordered by STS-content mapping. 18 refs., 2 figs., 1 tab.

  13. The Drosophila 7SK snRNP and the essential role of dHEXIM in development

    PubMed Central

    Nguyen, Duy; Krueger, Brian J.; Sedore, Stanley C.; Brogie, John E.; Rogers, Jason T.; Rajendra, T. K.; Saunders, Abbie; Matera, A. Greg; Lis, John T.; Uguen, Patricia

    2012-01-01

    Regulation of the positive transcription elongation factor, P-TEFb, plays a major role in controlling mammalian transcription and this is accomplished in part by controlled release of P-TEFb from the 7SK snRNP that sequesters the kinase in an inactive state. We demonstrate here that a similar P-TEFb control system exists in Drosophila. We show that an RNA previously suggested to be a 7SK homolog is, in fact, associated with P-TEFb, through the action of a homolog of the human HEXIM1/2 proteins (dHEXIM). In addition, a Drosophila La related protein (now called dLARP7) is shown to be the functional homolog of human LARP7. The Drosophila 7SK snRNP (d7SK snRNP) responded to treatment of cells with P-TEFb inhibitors and to nuclease treatment of cell lysates by releasing P-TEFb. Supporting a critical role for the d7SK snRNP in Drosophila development, dLARP7 and dHEXIM were found to be ubiquitously expressed throughout embryos and tissues at all stages. Importantly, knockdown of dHEXIM was embryonic lethal, and reduction of dHEXIM in specific tissues led to serious developmental defects. Our results suggest that regulation of P-TEFb by the d7SK snRNP is essential for the growth and differentiation of tissues required during Drosophila development. PMID:22379134

  14. The PSI-U1 snRNP interaction regulates male mating behavior in Drosophila.

    PubMed

    Wang, Qingqing; Taliaferro, J Matthew; Klibaite, Ugne; Hilgers, Valérie; Shaevitz, Joshua W; Rio, Donald C

    2016-05-10

    Alternative pre-mRNA splicing (AS) is a critical regulatory mechanism that operates extensively in the nervous system to produce diverse protein isoforms. Fruitless AS isoforms have been shown to influence male courtship behavior, but the underlying mechanisms are unknown. Using genome-wide approaches and quantitative behavioral assays, we show that the P-element somatic inhibitor (PSI) and its interaction with the U1 small nuclear ribonucleoprotein complex (snRNP) control male courtship behavior. PSI mutants lacking the U1 snRNP-interacting domain (PSIΔAB mutant) exhibit extended but futile mating attempts. The PSIΔAB mutant results in significant changes in the AS patterns of ∼1,200 genes in the Drosophila brain, many of which have been implicated in the regulation of male courtship behavior. PSI directly regulates the AS of at least one-third of these transcripts, suggesting that PSI-U1 snRNP interactions coordinate the behavioral network underlying courtship behavior. Importantly, one of these direct targets is fruitless, the master regulator of courtship. Thus, PSI imposes a specific mode of regulatory control within the neuronal circuit controlling courtship, even though it is broadly expressed in the fly nervous system. This study reinforces the importance of AS in the control of gene activity in neurons and integrated neuronal circuits, and provides a surprising link between a pleiotropic pre-mRNA splicing pathway and the precise control of successful male mating behavior.

  15. A Proteomic Strategy Identifies Lysine Methylation of Splicing Factor snRNP70 by the SETMAR Enzyme.

    PubMed

    Carlson, Scott M; Moore, Kaitlyn E; Sankaran, Saumya M; Reynoird, Nicolas; Elias, Joshua E; Gozani, Or

    2015-05-01

    The lysine methyltransferase (KMT) SETMAR is implicated in the response to and repair of DNA damage, but its molecular function is not clear. SETMAR has been associated with dimethylation of histone H3 lysine 36 (H3K36) at sites of DNA damage. However, SETMAR does not methylate H3K36 in vitro. This and the observation that SETMAR is not active on nucleosomes suggest that H3K36 methylation is not a physiologically relevant activity. To identify potential non-histone substrates, we utilized a strategy on the basis of quantitative proteomic analysis of methylated lysine. Our approach identified lysine 130 of the mRNA splicing factor snRNP70 as a SETMAR substrate in vitro, and we show that the enzyme primarily generates monomethylation at this position. Furthermore, we show that SETMAR methylates snRNP70 Lys-130 in cells. Because snRNP70 is a key early regulator of 5' splice site selection, our results suggest a model in which methylation of snRNP70 by SETMAR regulates constitutive and/or alternative splicing. In addition, the proteomic strategy described here is broadly applicable and is a promising route for large-scale mapping of KMT substrates.

  16. A Proteomic Strategy Identifies Lysine Methylation of Splicing Factor snRNP70 by the SETMAR Enzyme*

    PubMed Central

    Carlson, Scott M.; Moore, Kaitlyn E.; Sankaran, Saumya M.; Reynoird, Nicolas; Elias, Joshua E.; Gozani, Or

    2015-01-01

    The lysine methyltransferase (KMT) SETMAR is implicated in the response to and repair of DNA damage, but its molecular function is not clear. SETMAR has been associated with dimethylation of histone H3 lysine 36 (H3K36) at sites of DNA damage. However, SETMAR does not methylate H3K36 in vitro. This and the observation that SETMAR is not active on nucleosomes suggest that H3K36 methylation is not a physiologically relevant activity. To identify potential non-histone substrates, we utilized a strategy on the basis of quantitative proteomic analysis of methylated lysine. Our approach identified lysine 130 of the mRNA splicing factor snRNP70 as a SETMAR substrate in vitro, and we show that the enzyme primarily generates monomethylation at this position. Furthermore, we show that SETMAR methylates snRNP70 Lys-130 in cells. Because snRNP70 is a key early regulator of 5′ splice site selection, our results suggest a model in which methylation of snRNP70 by SETMAR regulates constitutive and/or alternative splicing. In addition, the proteomic strategy described here is broadly applicable and is a promising route for large-scale mapping of KMT substrates. PMID:25795785

  17. Cooperative binding of TIA-1 and U1 snRNP in K-SAM exon splicing activation

    SciTech Connect

    Gesnel, Marie-Claude; Theoleyre, Sandrine; Del Gatto-Konczak, Fabienne; Breathnach, Richard . E-mail: breathna@nantes.inserm.fr

    2007-07-13

    In 293 cells, splicing of the human fibroblast growth factor receptor-2 K-SAM alternative exon is inefficient, but can be made efficient by provoking TIA-1 binding to the U-rich IAS1 sequence downstream from the exon's 5' splice site. We show here that TIA-1 domains known to interact with U1 snRNP and to recruit it to 5' splice sites in vitro are required for TIA-1 activation of K-SAM exon splicing in vivo. We further show that tethering downstream from the K-SAM exon a fusion between the U1 snRNP component U1C and the bacteriophage MS2 coat protein provokes IAS1-dependent exon splicing, and present evidence that the fusion functions after its incorporation into U1 snRNP. Our in vivo data, taken together with previous in vitro results, show that K-SAM splicing activation involves cooperative binding of TIA-1 and U1 snRNP to the exon's 5' splice site region.

  18. snRNP protein expression enhances the formation of Cajal bodies containing p80-coilin and SMN.

    PubMed

    Sleeman, J E; Ajuh, P; Lamond, A I

    2001-12-01

    Splicing snRNPs (small nuclear ribonucleoproteins) are essential sub-units of the spliceosome. Here we report the establishment of stable cell lines expressing fluorescently tagged SmB, a core snRNP protein. Analysis of these stable cell lines has allowed us to characterize the nuclear pathway that leads to snRNP accumulation in nuclear speckles and has identified a limiting nucleolar step in the pathway that can be saturated by overexpression of Sm proteins. After nuclear import, newly assembled snRNPs accumulate first in a subset of Cajal bodies that contain both p80-coilin and the survival of motor neurons protein (SMN) and not in bodies that contain p80-coilin but lack SMN. Treatment of cells with leptomycin B (LMB) inhibits both the accumulation of snRNPs in nuclear bodies and their subsequent accumulation in speckles. The formation of Cajal bodies is enhanced by Sm protein expression and the assembly of new snRNPs. Formation of heterokaryons between HeLa cell lines expressing Sm proteins and primary cells that usually lack Cajal bodies results in the detection of Cajal bodies in primary cell nuclei. Transient over-expression of exogenous SmB alone is sufficient to induce correspondingly transient Cajal body formation in primary cells. These data indicate that the level of snRNP protein expression and snRNP assembly, rather than the expression levels of p80-coilin or SMN, may be a key trigger for Cajal body formation. PMID:11792806

  19. The splicing factor U2AF65 stabilizes TRF1 protein by inhibiting its ubiquitin-dependent proteolysis

    SciTech Connect

    Kim, Jeonghee; Chung, In Kwon

    2014-01-17

    Highlights: •Identification of U2AF65 as a novel TRF1-interacting protein. •U2AF65 stabilizes TRF1 protein by inhibiting its ubiquitin-dependent proteolysis. •U2AF65 interferes with the interaction between TRF1 and Fbx4. •U2AF65 represents a new route for modulating TRF1 function at telomeres. -- Abstract: The human telomeric protein TRF1 is a component of the six-subunit protein complex shelterin, which provides telomere protection by organizing the telomere into a high-order structure. TRF1 functions as a negative regulator of telomere length by controlling the access of telomerase to telomeres. Thus, the cellular abundance of TRF1 at telomeres should be maintained and tightly regulated to ensure proper telomere function. Here, we identify U2 small nuclear ribonucleoprotein (snRNP) auxiliary factor 65 (U2AF65), an essential pre-mRNA splicing factor, as a novel TRF1-interacting protein. U2AF65 interacts with TRF1 in vitro and in vivo and is capable of stabilizing TRF1 protein by inhibiting its ubiquitin-dependent proteolysis. We also found that U2AF65 interferes with the interaction between TRF1 and Fbx4, an E3 ubiquitin ligase for TRF1. Depletion of endogenous U2AF65 expression by short interfering RNA (siRNA) reduced the stability of endogenous TRF1 whereas overexpression of U2AF65 significantly extended the half-life of TRF1. These findings demonstrate that U2AF65 plays a critical role in regulating the level of TRF1 through physical interaction and ubiquitin-mediated proteolysis. Hence, U2AF65 represents a new route for modulating TRF1 function at telomeres.

  20. Vought XO4U-2

    NASA Technical Reports Server (NTRS)

    1933-01-01

    Vought XO4U-2: A biplane scout, the Vought XO4U-2 was 'flown' in the NACA's 30 x 60 Full Scale Tunnel at the Langley Memorial Aeronautical Laboratory in early spring 1933. Part of these tests were to study the cooling of the Pratt & Whitney Twin Wasp radial engine. Other tests involved the relation of the slipstream to stability and control.

  1. Yeast ortholog of the Drosophila crooked neck protein promotes spliceosome assembly through stable U4/U6.U5 snRNP addition.

    PubMed Central

    Chung, S; McLean, M R; Rymond, B C

    1999-01-01

    Mutants in the Drosophila crooked neck (crn) gene show an embryonic lethal phenotype with severe developmental defects. The unusual crn protein consists of sixteen tandem repeats of the 34 amino acid tetratricopeptide (TPR) protein recognition domain. Crn-like TPR elements are found in several RNA processing proteins, although it is unknown how the TPR repeats or the crn protein contribute to Drosophila development. We have isolated a Saccharomyces cerevisiae gene, CLF1, that encodes a crooked neck-like factor. CLF1 is an essential gene but the lethal phenotype of a clf1::HIS3 chromosomal null mutant can be rescued by plasmid-based expression of CLF1 or the Drosophila crn open reading frame. Clf1p is required in vivo and in vitro for pre-mRNA 5' splice site cleavage. Extracts depleted of Clf1p arrest spliceosome assembly after U2 snRNP addition but prior to productive U4/U6.U5 association. Yeast two-hybrid analyses and in vitro binding studies show that Clf1p interacts specifically and differentially with the U1 snRNP-Prp40p protein and the yeast U2AF65 homolog, Mud2p. Intriguingly, Prp40p and Mud2p also bind the phylogenetically conserved branchpoint binding protein (BBP/SF1). Our results indicate that Clf1p acts as a scaffolding protein in spliceosome assembly and suggest that Clf1p may support the cross-intron bridge during the prespliceosome-to-spliceosome transition. PMID:10445879

  2. Cracking the control of RNA polymerase II elongation by 7SK snRNP and P-TEFb

    PubMed Central

    C. Quaresma, Alexandre J.; Bugai, Andrii; Barboric, Matjaz

    2016-01-01

    Release of RNA polymerase II (Pol II) from promoter-proximal pausing has emerged as a critical step regulating gene expression in multicellular organisms. The transition of Pol II into productive elongation requires the kinase activity of positive transcription elongation factor b (P-TEFb), which is itself under a stringent control by the inhibitory 7SK small nuclear ribonucleoprotein (7SK snRNP) complex. Here, we provide an overview on stimulating Pol II pause release by P-TEFb and on sequestering P-TEFb into 7SK snRNP. Furthermore, we highlight mechanisms that govern anchoring of 7SK snRNP to chromatin as well as means that release P-TEFb from the inhibitory complex, and propose a unifying model of P-TEFb activation on chromatin. Collectively, these studies shine a spotlight on the central role of RNA binding proteins (RBPs) in directing the inhibition and activation of P-TEFb, providing a compelling paradigm for controlling Pol II transcription with a non-coding RNA. PMID:27369380

  3. Reconstitution of the human U snRNP assembly machinery reveals stepwise Sm protein organization.

    PubMed

    Neuenkirchen, Nils; Englbrecht, Clemens; Ohmer, Jürgen; Ziegenhals, Thomas; Chari, Ashwin; Fischer, Utz

    2015-07-14

    The assembly of spliceosomal U snRNPs depends on the coordinated action of PRMT5 and SMN complexes in vivo. These trans-acting factors enable the faithful delivery of seven Sm proteins onto snRNA and the formation of the common core of snRNPs. To gain mechanistic insight into their mode of action, we reconstituted the assembly machinery from recombinant sources. We uncover a stepwise and ordered formation of distinct Sm protein complexes on the PRMT5 complex, which is facilitated by the assembly chaperone pICln. Upon completion, the formed pICln-Sm units are displaced by new pICln-Sm protein substrates and transferred onto the SMN complex. The latter acts as a Brownian machine that couples spontaneous conformational changes driven by thermal energy to prevent mis-assembly and to ensure the transfer of Sm proteins to cognate RNA. Investigation of mutant SMN complexes provided insight into the contribution of individual proteins to these activities. The biochemical reconstitution presented here provides a basis for a detailed molecular dissection of the U snRNP assembly reaction.

  4. Core structure of the U6 snRNP at 1.7 Å resolution

    PubMed Central

    Montemayor, Eric J.; Curran, Elizabeth C.; Liao, Hong Hong; Andrews, Kristie L.; Treba, Christine N.; Butcher, Samuel E.; Brow, David A.

    2014-01-01

    The spliceosome is a dynamic assembly of five small nuclear ribonucleoproteins (snRNPs) that removes introns from eukaryotic pre-mRNA. U6 is the most conserved of the spliceosomal snRNAs and participates directly in catalysis. Here, we report the crystal structure of the Saccharomyces cerevisiae U6 snRNP core, containing most of U6 snRNA and all four RRM domains of the Prp24 protein. It reveals a unique interlocked RNP architecture that sequesters the 5′ splice site-binding bases of U6 snRNA. RRMs 1, 2 and 4 of Prp24 form an electropositive groove that binds double-stranded RNA and may nucleate annealing of U4 and U6 snRNAs. Substitutions in Prp24 that suppress a mutation in U6 localize to direct RNA-protein contacts. Our results provide the most complete view to date of a multi-RRM protein bound to RNA, and reveal striking co-evolution of protein and RNA structure. PMID:24837192

  5. The human U5 snRNP 52K protein (CD2BP2) interacts with U5-102K (hPrp6), a U4/U6.U5 tri-snRNP bridging protein, but dissociates upon tri-snRNP formation

    PubMed Central

    LAGGERBAUER, BERNHARD; LIU, SUNBIN; MAKAROV, EVGENY; VORNLOCHER, HANS-PETER; MAKAROVA, OLGA; INGELFINGER, DIERK; ACHSEL, TILMANN; LÜHRMANN, REINHARD

    2005-01-01

    The U5 snRNP plays an essential role in both U2- and U12-dependent splicing. Here, we have characterized a 52-kDa protein associated with the human U5 snRNP, designated U5-52K. Protein sequencing revealed that U5-52K is identical to the CD2BP2, which interacts with the cytoplasmic portion of the human T-cell surface protein CD2. Consistent with it associating with an snRNP, immunofluorescence studies demonstrated that the 52K protein is predominantly located in the nucleoplasm of HeLa cells, where it overlaps, at least in part, with splicing-factor compartments (or “speckles”). We further demonstrate that the 52K protein is a constituent of the 20S U5 snRNP, but is not found in U4/U6.U5 tri-snRNPs. Thus, it is the only 20S U5-specific protein that is not integrated into the tri-snRNP and resembles, in this respect, the U4/U6 di-snRNP assembly factor Prp24p/p110. Yeast two-hybrid screening and pulldown assays revealed that the 52K protein interacts with the U5-specific 102K and 15K proteins, suggesting that these interactions are responsible for its integration into the U5 particle. The N-terminal two-thirds of 52K interact with the 102K protein, whereas its C-terminal GYF-domain binds the 15K protein. As the latter lacks a proline-rich tract, our data indicate that a GYF-domain can also engage in specific protein–protein interactions in a polyproline-independent manner. Interestingly, the U5-102K protein has been shown previously to play an essential role in tri-snRNP formation, binding the U4/U6-61K protein. The interaction of 52K with a tri-snRNP bridging protein, coupled with its absence from the tri-snRNP, suggests it might function in tri-snRNP assembly. PMID:15840814

  6. RAN/TC4 mutants identify a common requirement for snRNP and protein import into the nucleus

    PubMed Central

    1996-01-01

    Kinetic competition experiments have demonstrated that at least some factors required for the nuclear import of proteins and U snRNPs are distinct. Both import processes require energy, and in the case of protein import, the energy requirement is known to be at least partly met by GTP hydrolysis by the Ran GTPase. We have compared the effects of nonhydrolyzable GTP analogues and two mutant Ran proteins on the nuclear import of proteins and U snRNPs in vitro. The mutant Ran proteins have different defects; Q69L (glutamine 69 changed to leucine) is defective in GTP hydrolysis while T24N (threonine 24 changed to asparagine) is defective in binding GTP. Both protein and snRNP import are sensitive either to the presence of the two mutant Ran proteins, which act as dominant negative inhibitors of nuclear import, or to incubation with nonhydrolyzable GTP analogues. This demonstrates that there is a requirement for a GTPase activity for the import of U snRNPs, as well as proteins, into the nucleus. The dominant negative effects of the two mutant Ran proteins indicate that the pathways of protein and snRNP import share at lease one common component. PMID:8636225

  7. Functional organization of the Sm core in the crystal structure of human U1 snRNP

    PubMed Central

    Weber, Gert; Trowitzsch, Simon; Kastner, Berthold; Lührmann, Reinhard; Wahl, Markus C

    2010-01-01

    U1 small nuclear ribonucleoprotein (snRNP) recognizes the 5′-splice site early during spliceosome assembly. It represents a prototype spliceosomal subunit containing a paradigmatic Sm core RNP. The crystal structure of human U1 snRNP obtained from natively purified material by in situ limited proteolysis at 4.4 Å resolution reveals how the seven Sm proteins, each recognize one nucleotide of the Sm site RNA using their Sm1 and Sm2 motifs. Proteins D1 and D2 guide the snRNA into and out of the Sm ring, and proteins F and E mediate a direct interaction between the Sm site termini. Terminal extensions of proteins D1, D2 and B/B′, and extended internal loops in D2 and B/B′ support a four-way RNA junction and a 3′-terminal stem-loop on opposite sides of the Sm core RNP, respectively. On a higher organizational level, the core RNP presents multiple attachment sites for the U1-specific 70K protein. The intricate, multi-layered interplay of proteins and RNA rationalizes the hierarchical assembly of U snRNPs in vitro and in vivo. PMID:21113136

  8. Autoimmune response to U1 small nuclear ribonucleoprotein (U1 snRNP) associated with cytomegalovirus infection.

    PubMed

    Newkirk, M M; van Venrooij, W J; Marshall, G S

    2001-01-01

    The induction of autoantibodies to U1 small nuclear ribonucleoprotein (U1 snRNP) complexes is not well understood. We present evidence that healthy individuals with cytomegalovirus (CMV) infection have an increased frequency and quantity of antibodies to ribonucleoprotein, directed primarily against the U1-70k protein. A significant association between the presence of antibodies to CMV and antibodies to the total RNP targeted by the immune response to the spliceosome (to both the Sm and RNP; Sm/RNP) was found for patients with systemic lupus erythematosus (SLE) but not those with mixed connective-tissue disease. CMV thus may play a role in inducing autoimmune responses in a subset of patients with systemic lupus erythematosus.

  9. Autoimmune response to U1 small nuclear ribonucleoprotein (U1 snRNP) associated with cytomegalovirus infection.

    PubMed

    Newkirk, M M; van Venrooij, W J; Marshall, G S

    2001-01-01

    The induction of autoantibodies to U1 small nuclear ribonucleoprotein (U1 snRNP) complexes is not well understood. We present evidence that healthy individuals with cytomegalovirus (CMV) infection have an increased frequency and quantity of antibodies to ribonucleoprotein, directed primarily against the U1-70k protein. A significant association between the presence of antibodies to CMV and antibodies to the total RNP targeted by the immune response to the spliceosome (to both the Sm and RNP; Sm/RNP) was found for patients with systemic lupus erythematosus (SLE) but not those with mixed connective-tissue disease. CMV thus may play a role in inducing autoimmune responses in a subset of patients with systemic lupus erythematosus. PMID:11438044

  10. Structural Basis of Brr2-Prp8 Interactions and Implications for U5 snRNP Biogenesis and the Spliceosome Active Site

    PubMed Central

    Nguyen, Thi Hoang Duong; Li, Jade; Galej, Wojciech P.; Oshikane, Hiroyuki; Newman, Andrew J.; Nagai, Kiyoshi

    2013-01-01

    Summary The U5 small nuclear ribonucleoprotein particle (snRNP) helicase Brr2 disrupts the U4/U6 small nuclear RNA (snRNA) duplex and allows U6 snRNA to engage in an intricate RNA network at the active center of the spliceosome. Here, we present the structure of yeast Brr2 in complex with the Jab1/MPN domain of Prp8, which stimulates Brr2 activity. Contrary to previous reports, our crystal structure and mutagenesis data show that the Jab1/MPN domain binds exclusively to the N-terminal helicase cassette. The residues in the Jab1/MPN domain, whose mutations in human Prp8 cause the degenerative eye disease retinitis pigmentosa, are found at or near the interface with Brr2, clarifying its molecular pathology. In the cytoplasm, Prp8 forms a precursor complex with U5 snRNA, seven Sm proteins, Snu114, and Aar2, but after nuclear import, Brr2 replaces Aar2 to form mature U5 snRNP. Our structure explains why Aar2 and Brr2 are mutually exclusive and provides important insights into the assembly of U5 snRNP. PMID:23727230

  11. Electronic structure of U2PtC2 and U2RhC2

    DOE PAGES

    Ronning, F.; Zhu, J. -X.

    2015-03-18

    In this study, we present density functional theory calculations within the generalized gradient approximation of U2RhC2 and U2PtC2. We find the calculated density of states are significantly less than that measured by specific heat indicating the need for electronic correlations. The mass enhancement found for U2PtC2 is m*/mband ≈ 4.

  12. High intranuclear mobility and dynamic clustering of the splicing factor U1 snRNP observed by single particle tracking

    PubMed Central

    Kues, Thorsten; Dickmanns, Achim; Lührmann, Reinhard; Peters, Reiner; Kubitscheck, Ulrich

    2001-01-01

    Uridine-rich small nuclear ribonucleoproteins (U snRNPs) are components of the splicing machinery that removes introns from precursor mRNA. Like other splicing factors, U snRNPs are diffusely distributed throughout the nucleus and, in addition, are concentrated in distinct nuclear substructures referred to as speckles. We have examined the intranuclear distribution and mobility of the splicing factor U1 snRNP on a single-molecule level. Isolated U1 snRNPs were fluorescently labeled and incubated with digitonin-permeabilized 3T3 cells in the presence of Xenopus egg extract. By confocal microscopy, U1 snRNPs were found to be imported into nuclei, yielding a speckled intranuclear distribution. Employing a laser video-microscope optimized for high sensitivity and high speed, single U1 snRNPs were visualized and tracked at a spatial precision of 35 nm and a time resolution of 30 ms. The single-particle data revealed that U1 snRNPs occurred in small clusters that colocalized with speckles. In the clusters, U1 snRNPs resided for a mean decay time of 84 ms before leaving the optical slice in the direction of the optical axis, which corresponded to a mean effective diffusion coefficient of 1 μm2/s. An analysis of the trajectories of single U1 snRNPs revealed that at least three kinetic classes of low, medium, and high mobility were present. Moreover, the mean square displacements of these fractions were virtually independent of time, suggesting arrays of binding sites. The results substantiate the view that nuclear speckles are not rigid structures but highly dynamic domains characterized by a rapid turnover of U1 snRNPs and other splicing factors. PMID:11593012

  13. The conserved central domain of yeast U6 snRNA: importance of U2-U6 helix Ia in spliceosome assembly.

    PubMed Central

    Ryan, Daniel E; Abelson, John

    2002-01-01

    In the pre-mRNA processing machinery of eukaryotic cells, U6 snRNA is located at or near the active site for pre-mRNA splicing catalysis, and U6 is involved in catalyzing the first chemical step of splicing. We have further defined the roles of key features of yeast U6 snRNA in the splicing process. By assaying spliceosome assembly and splicing in yeast extracts, we found that mutations of yeast U6 nt 56 and 57 are similar to previously reported deletions of U2 nt 27 or 28, all within yeast U2-U6 helix Ia. These mutations lead to the accumulation of yeast A1 spliceosomes, which form just prior to the Prp2 ATPase step and the first chemical step of splicing. These results strongly suggest that, at a late stage of spliceosome assembly, the presence of U2-U6 helix Ia is important for promoting the first chemical step of splicing, presumably by bringing together the 5' splice site region of pre-mRNA, which is base paired to U6 snRNA, and the branchsite region of the intron, which is base paired to U2 snRNA, for activation of the first chemical step of splicing, as previously proposed by Madhani and Guthrie [Cell, 1992, 71: 803-817]. In the 3' intramolecular stem-loop of U6, mutation G81C causes an allele-specific accumulation of U6 snRNP. Base pairing of the U6 3' stem-loop in yeast spliceosomes does not extend as far as to include the U6 sequence of U2-U6 helix Ib, in contrast to the human U6 3' stem-loop structure. PMID:12212854

  14. U2 And NASA Create Video To Celebrate Collaboration

    NASA Video Gallery

    NASA and U2 released a commemorative video highlighting a year's worth of collaboration in space and on the Irish rock band's 360 Degree tour. U2 approached NASA in 2009 with an idea to include a d...

  15. Dinoflagellate 17S rRNA sequence inferred from the gene sequence: Evolutionary implications.

    PubMed

    Herzog, M; Maroteaux, L

    1986-11-01

    We present the complete sequence of the nuclear-encoded small-ribosomal-subunit RNA inferred from the cloned gene sequence of the dinoflagellate Prorocentrum micans. The dinoflagellate 17S rRNA sequence of 1798 nucleotides is contained in a family of 200 tandemly repeated genes per haploid genome. A tentative model of the secondary structure of P. micans 17S rRNA is presented. This sequence is compared with the small-ribosomal-subunit rRNA of Xenopus laevis (Animalia), Saccharomyces cerevisiae (Fungi), Zea mays (Planta), Dictyostelium discoideum (Protoctista), and Halobacterium volcanii (Monera). Although the secondary structure of the dinoflagellate 17S rRNA presents most of the eukaryotic characteristics, it contains sufficient archaeobacterial-like structural features to reinforce the view that dinoflagellates branch off very early from the eukaryotic lineage.

  16. Dinoflagellate 17S rRNA sequence inferred from the gene sequence: Evolutionary implications

    PubMed Central

    Herzog, Michel; Maroteaux, Luc

    1986-01-01

    We present the complete sequence of the nuclear-encoded small-ribosomal-subunit RNA inferred from the cloned gene sequence of the dinoflagellate Prorocentrum micans. The dinoflagellate 17S rRNA sequence of 1798 nucleotides is contained in a family of 200 tandemly repeated genes per haploid genome. A tentative model of the secondary structure of P. micans 17S rRNA is presented. This sequence is compared with the small-ribosomal-subunit rRNA of Xenopus laevis (Animalia), Saccharomyces cerevisiae (Fungi), Zea mays (Planta), Dictyostelium discoideum (Protoctista), and Halobacterium volcanii (Monera). Although the secondary structure of the dinoflagellate 17S rRNA presents most of the eukaryotic characteristics, it contains sufficient archaeobacterial-like structural features to reinforce the view that dinoflagellates branch off very early from the eukaryotic lineage. PMID:16578795

  17. Dinoflagellate 17S rRNA sequence inferred from the gene sequence: Evolutionary implications.

    PubMed

    Herzog, M; Maroteaux, L

    1986-11-01

    We present the complete sequence of the nuclear-encoded small-ribosomal-subunit RNA inferred from the cloned gene sequence of the dinoflagellate Prorocentrum micans. The dinoflagellate 17S rRNA sequence of 1798 nucleotides is contained in a family of 200 tandemly repeated genes per haploid genome. A tentative model of the secondary structure of P. micans 17S rRNA is presented. This sequence is compared with the small-ribosomal-subunit rRNA of Xenopus laevis (Animalia), Saccharomyces cerevisiae (Fungi), Zea mays (Planta), Dictyostelium discoideum (Protoctista), and Halobacterium volcanii (Monera). Although the secondary structure of the dinoflagellate 17S rRNA presents most of the eukaryotic characteristics, it contains sufficient archaeobacterial-like structural features to reinforce the view that dinoflagellates branch off very early from the eukaryotic lineage. PMID:16578795

  18. The human U1-70K snRNP protein: cDNA cloning, chromosomal localization, expression, alternative splicing and RNA-binding.

    PubMed Central

    Spritz, R A; Strunk, K; Surowy, C S; Hoch, S O; Barton, D E; Francke, U

    1987-01-01

    We have isolated and sequenced cDNA clones encoding the human U1-70K snRNP protein, and have mapped this locus (U1AP1) to human chromosome 19. The gene produces two size classes of RNA, a major 1.7-kb RNA and a minor 3.9-kb RNA. The 1.7-kb species appears to be the functional mRNA; the role of the 3.9-kb RNA, which extends further in the 5' direction, is unclear. The actual size of the hU1-70K protein is probably 52 kd, rather than 70 kd. The protein contains three regions similar to known nucleic acid-binding proteins, and it binds RNA in an in vitro assay. Comparison of the cDNA sequences indicates that there are multiple subclasses of mRNA that arise by alternative pre-mRNA splicing of at least four alternative exon segments. This suggests that multiple forms of the hU1-70K protein may exist, possibly with different functions in vivo. Images PMID:2447561

  19. A Natural Product from Polygonum cuspidatum Sieb. Et Zucc. Promotes Tat-Dependent HIV Latency Reversal through Triggering P-TEFb's Release from 7SK snRNP.

    PubMed

    Wang, Cong; Yang, Shuiyuan; Lu, Huasong; You, Hongchao; Ni, Man; Shan, Wenjun; Lin, Ting; Gao, Xiang; Chen, Haifeng; Zhou, Qiang; Xue, Yuhua

    2015-01-01

    The latent reservoirs of HIV represent a major impediment to eradication of HIV/AIDS. To overcome this problem, agents that can activate latent HIV proviruses have been actively sought after, as they can potentially be used in combination with the highly active antiretroviral therapy (HAART) to eliminate the latent reservoirs. Although several chemical compounds have been shown to activate latency, they are of limited use due to high toxicity and poor clinical outcomes. In an attempt to identify natural products as effective latency activators from traditional Chinese medicinal herbs that have long been widely used in human population, we have isolated procyanidin C-13,3',3"-tri-O-gallate (named as REJ-C1G3) from Polygonum cuspidatum Sieb. et Zucc., that can activate HIV in latently infected Jurkat T cells. REJ-C1G3 preferentially stimulates HIV transcription in a process that depends on the viral encoded Tat protein and acts synergistically with prostratin (an activator of the NF-κB pathway) or JQ1 (an inhibitor of Brd4) to activate HIV latency. Our mechanistic analyses further show that REJ-C1G3 accomplishes these tasks by inducing the release of P-TEFb, a host cofactor essential for Tat-activation of HIV transcription, from the cellular P-TEFb reservoir 7SK snRNP. PMID:26569506

  20. Building Customized University-to-Business (U2B) Partnerships

    ERIC Educational Resources Information Center

    Irvine, George; Verma, Lisa

    2013-01-01

    Continuing education (CE) units throughout the United States have successfully built University-to-Business (U2B) partnerships to provide greater value to their community partners and to increase revenue for the university. Our experience in building U2B partnerships and feedback from our partners--businesses, corporations, state agencies, and…

  1. General Approach for Tetramer Based Identification of Autoantigen Reactive B Cells: Characterization of La and snRNP Reactive B Cells in Autoimmune BXD2 Mice

    PubMed Central

    Hamilton, Jennie A.; Li, Jun; Wu, Qi; Yang, PingAr; Luo, Bao; Li, Hao; Bradley, John E.; Taylor, Justin J.; Randall, Troy D.; Mountz, John D.; Hsu, Hui-Chen

    2015-01-01

    Autoreactive B cells are associated with the development of several autoimmune diseases, including systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). The low frequency of these cells represents a major barrier to their analysis. Antigen-tetramers prepared from linear epitopes represent a promising strategy for the identification of small subsets of antigen-reactive immune cells. This is challenging given the requirement for identification and validation of linear epitopes and the complexity of autoantibody responses, including the broad spectrum of autoantibody specificities and the contribution of isotype to pathogenicity. We therefore tested a two-tiered peptide microarray approach, coupled with epitope mapping of known autoantigens, to identify and characterize autoepitopes using the BXD2 autoimmune mouse model. Microarray results were verified through comparison with established age-associated profiles of autoantigen specificities and autoantibody class switching in BXD2 and control (B6) mice and high-throughput ELISA and ELISPOT analyses of synthetic peptides. Tetramers were prepared from two linear peptides derived from two ribonucleic acid binding proteins (RBP): lupus La and 70 kDa U1 small nuclear ribonucleoprotein (snRNP). Flow cyotmetric analysis of tetramer-reactive B-cell subsets revealed a significantly higher frequency and greater numbers of RBP-reactive marginal zone precursor (MZ-P), transitional T3 and PDL-2+CD80+ memory B cells, with significantly elevated CD69 and CD86 observed in RBP+ MZ-P B cells in the spleens of BXD2 compared to B6 mice, suggesting a regulatory defect. This study establishes a feasible strategy for the characterization of autoantigen-specific B-cell subsets in different models of autoimmunity and, potentially, humans. PMID:25888644

  2. Eschewing obfuscation - MIL-HDBK-17's approach to composite material test methods development

    NASA Astrophysics Data System (ADS)

    Spigel, Barry

    Military Handbook 17, 'Polymer Matrix Composites', establishes guidelines for the use of composite material test methods. A discussion is presented of the rationale behind the test method guidelines, and the consequences of MIL-HDBK-17's contents on the use and standardization of test methods. Test method requirements feature rigorous review of precision and bias, interlaboratory testing, and publication in an open-literature document. The criteria also permit inclusion of 'common practice' test methods in the handbook as long as the test has begun the process towards standardization, thereby allowing an independent determination of precision and bias.

  3. Disruption of snRNP biogenesis factors Tgs1 and pICln induces phenotypes that mirror aspects of SMN-Gemins complex perturbation in Drosophila, providing new insights into spinal muscular atrophy.

    PubMed

    Borg, Rebecca M; Fenech Salerno, Benji; Vassallo, Neville; Bordonne, Rémy; Cauchi, Ruben J

    2016-10-01

    The neuromuscular disorder, spinal muscular atrophy (SMA), results from insufficient levels of the survival motor neuron (SMN) protein. Together with Gemins 2-8 and Unrip, SMN forms the large macromolecular SMN-Gemins complex, which is known to be indispensable for chaperoning the assembly of spliceosomal small nuclear ribonucleoproteins (snRNPs). It remains unclear whether disruption of this function is responsible for the selective neuromuscular degeneration in SMA. In the present study, we first show that loss of wmd, the Drosophila Unrip orthologue, has a negative impact on the motor system. However, due to lack of a functional relationship between wmd/Unrip and Gemin3, it is likely that Unrip joined the SMN-Gemins complex only recently in evolution. Second, we uncover that disruption of either Tgs1 or pICln, two cardinal players in snRNP biogenesis, results in viability and motor phenotypes that closely resemble those previously uncovered on loss of the constituent members of the SMN-Gemins complex. Interestingly, overexpression of both factors leads to motor dysfunction in Drosophila, a situation analogous to that of Gemin2. Toxicity is conserved in the yeast S. pombe where pICln overexpression induces a surplus of Sm proteins in the cytoplasm, indicating that a block in snRNP biogenesis is partly responsible for this phenotype. Importantly, we show a strong functional relationship and a physical interaction between Gemin3 and either Tgs1 or pICln. We propose that snRNP biogenesis is the pathway connecting the SMN-Gemins complex to a functional neuromuscular system, and its disturbance most likely leads to the motor dysfunction that is typical in SMA. PMID:27388936

  4. 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.

  5. Superconformal mechanics in S U (2 |1 ) superspace

    NASA Astrophysics Data System (ADS)

    Ivanov, E.; Sidorov, S.; Toppan, F.

    2015-04-01

    Using the worldline S U (2 |1 ) superfield approach, we construct N =4 superconformally invariant actions for the d =1 multiplets (1 ,4 ,3 ) and (2 ,4 ,2 ). The S U (2 |1 ) superfield framework automatically implies the trigonometric realization of the superconformal symmetry and the harmonic oscillator term in the corresponding component actions. We deal with the general N =4 superconformal algebra D (2 ,1 ;α ) and its central-extended α =0 and α =-1 p s u (1 ,1 |2 )⊕s u (2 ) descendants. We capitalize on the observation that D (2 ,1 ;α ) at α ≠0 can be treated as a closure of its two s u (2 |1 ) subalgebras, one of which defines the superisometry of the S U (2 |1 ) superspace, while the other is related to the first one through the reflection of μ , the parameter of contraction to the flat N =4 ,d =1 superspace. This closure property and its α =0 analog suggest a simple criterion for the S U (2 |1 ) invariant actions to be superconformal: they should be even functions of μ . We find that the superconformal actions of the multiplet (2 ,4 ,2 ) exist only at α =-1 ,0 and are reduced to a sum of the free sigma-model-type action and the conformal superpotential yielding, respectively, the oscillator potential ˜μ2 and the standard conformal inverse-square potential in the bosonic sector. The sigma-model action in this case can be constructed only on account of nonzero central charge in the superalgebra s u (1 ,1 |2 ).

  6. U-2 Spy Plane With Fictitious NASA Markings

    NASA Technical Reports Server (NTRS)

    1960-01-01

    After Francis Gary Powers was shot down over the Soviet Union during a CIA spy flight on May 1. 1960, NASA issued a press release with a cover story about a U-2 conducting weather research that may have strayed off course after the pilot reported difficulties with his oxygen equipment. To bolster the cover-up, a U-2 was quickly painted in NASA markings, with a fictitious NASA serial number, and put on display for the news media at the NASA Flight Research Center at Edwards Air Force Base on May 6, 1960. The U-2 cover story in 1956 was that it was an NACA plane to conduct high-altitude weather research. But various observers doubted this story from the beginning. Certainly the Soviets did not believe it once the aircraft began overflying their territory. The NASA cover story quickly blew up in the agency's face when both Gary Powers and aircraft wreckage were displayed by the Soviet Union, proving that it was a reconnaissance aircraft. This caused embarrassment for several top NASA officials.

  7. Ocean Chlorophyll Studies from a U-2 Aircraft Platform

    NASA Technical Reports Server (NTRS)

    Kim, H. H.; Mcclain, C. R.; Blaine, L. R.; Hart, W. D.; Atkinson, L. P.; Yoder, J. A.

    1979-01-01

    Chlorophyll gradient maps of large ocean areas were generated from U-2 ocean color scanner data obtained over test sites in the Pacific and Atlantic Oceans. The delineation of oceanic features using the upward radiant intensity relies on an analysis method which presupposes that radiation backscattered from the atmosphere and ocean surface can be properly modeled using a measurement made at 778 nm. An estimation of the chlorophyll concentration was performed by properly ratioing radiances measured at 472 nm and 548 nm after removing the atmospheric effects. The correlation between the remotely sensed data and in-situ surface chlorophyll measurements was validated in two sets of data. The results show that the correlation between the in-situ measured chlorophyll and the derived quantity is a negative exponential function and the correlation coefficient was calculated to be -0.965.

  8. Quick look data report for COMET Test U2

    SciTech Connect

    Farmer, M.T.; Spencer, B.W.; Kilsdonk, D.J.; Aeschlimann, R.W.

    1998-01-08

    Investigations are underway at Forschungszentrum Karlsruhe (FZK) addressing methods to terminate and stabilize a core melt accident situation ex-vessel. In this approach, the molten core-concrete interaction (MCCI) begins erosion of the concrete, and after erosion proceeds to some modest depth, it exposes and unseals an array of tubes. The tubes are connected to a water reservoir pressurized by static water head. Upon unsealing, the tubes direct a flow of water into the bottom of the corium layer. The water is forced up through the melt, cooling the melt and causing it to solidify in a form that allows continued permeation and heat removal by the water. Thus, the accident progression can be halted, and the debris may be permanently cooled. The key aspect of the passive ex-vessel core retention approach described above is the ability of water injected at the bottom of a corium melt layer to quench the melt forming a coolable debris bed in the process. This process has been tested using iron-alumina thermite as a corium simulant with promising results. As a part of a collaborative research agreement between FZK and the US DOE, two scoping tests are being conducted at Argonne National Laboratory to test the FZK core retention concept using real reactor materials. The second of these two tests, denoted COMET Test U2, was successfully conducted on December 17, 1997. The objectives of this data report are to: summarize the experiment facility and operating procedure for COMET Test U2, and present the test data.

  9. Wild-Type U2AF1 Antagonizes the Splicing Program Characteristic of U2AF1-Mutant Tumors and Is Required for Cell Survival

    PubMed Central

    Fei, Dennis Liang; Motowski, Hayley; Chatrikhi, Rakesh; Gao, Shaojian; Kielkopf, Clara L.; Varmus, Harold

    2016-01-01

    We have asked how the common S34F mutation in the splicing factor U2AF1 regulates alternative splicing in lung cancer, and why wild-type U2AF1 is retained in cancers with this mutation. A human lung epithelial cell line was genetically modified so that U2AF1S34F is expressed from one of the two endogenous U2AF1 loci. By altering levels of mutant or wild-type U2AF1 in this cell line and by analyzing published data on human lung adenocarcinomas, we show that S34F-associated changes in alternative splicing are proportional to the ratio of S34F:wild-type gene products and not to absolute levels of either the mutant or wild-type factor. Preferential recognition of specific 3′ splice sites in S34F-expressing cells is largely explained by differential in vitro RNA-binding affinities of mutant versus wild-type U2AF1 for those same 3′ splice sites. Finally, we show that lung adenocarcinoma cell lines bearing U2AF1 mutations do not require the mutant protein for growth in vitro or in vivo. In contrast, wild-type U2AF1 is required for survival, regardless of whether cells carry the U2AF1S34F allele. Our results provide mechanistic explanations of the magnitude of splicing changes observed in U2AF1-mutant cells and why tumors harboring U2AF1 mutations always retain an expressed copy of the wild-type allele. PMID:27776121

  10. A Natural Product from Polygonum cuspidatum Sieb. Et Zucc. Promotes Tat-Dependent HIV Latency Reversal through Triggering P-TEFb’s Release from 7SK snRNP

    PubMed Central

    Lu, Huasong; You, Hongchao; Ni, Man; Shan, Wenjun; Lin, Ting; Gao, Xiang; Chen, Haifeng; Zhou, Qiang; Xue, Yuhua

    2015-01-01

    The latent reservoirs of HIV represent a major impediment to eradication of HIV/AIDS. To overcome this problem, agents that can activate latent HIV proviruses have been actively sought after, as they can potentially be used in combination with the highly active antiretroviral therapy (HAART) to eliminate the latent reservoirs. Although several chemical compounds have been shown to activate latency, they are of limited use due to high toxicity and poor clinical outcomes. In an attempt to identify natural products as effective latency activators from traditional Chinese medicinal herbs that have long been widely used in human population, we have isolated procyanidin C-13,3',3"-tri-O-gallate (named as REJ-C1G3) from Polygonum cuspidatum Sieb. et Zucc., that can activate HIV in latently infected Jurkat T cells. REJ-C1G3 preferentially stimulates HIV transcription in a process that depends on the viral encoded Tat protein and acts synergistically with prostratin (an activator of the NF-κB pathway) or JQ1 (an inhibitor of Brd4) to activate HIV latency. Our mechanistic analyses further show that REJ-C1G3 accomplishes these tasks by inducing the release of P-TEFb, a host cofactor essential for Tat-activation of HIV transcription, from the cellular P-TEFb reservoir 7SK snRNP. PMID:26569506

  11. P elements inserted in the vicinity of or within the Drosophila snRNP SmD3 gene nested in the first intron of the Ornithine Decarboxylase Antizyme gene affect only the expression of SmD3.

    PubMed Central

    Schenkel, Heide; Hanke, Susanne; De Lorenzo, Cécilia; Schmitt, Rolf; Mechler, Bernard M

    2002-01-01

    The Drosophila gene for snRNP SmD3 (SmD3) is contained in reverse orientation within the first intron of the Ornithine Decarboxylase Antizyme (AZ) gene. Previous studies show that two closely linked P elements cause the gutfeeling phenotype characterized by embryonic lethality and aberrant neuronal and muscle cell differentiation. However, the exact nature of the gene(s) affected in the gutfeeling phenotype remained unknown. This study shows that a series of P inserts located within the 5'-untranslated region (5'-UTR) of SmD3 or its promoter affects only the expression of SmD3. Our analysis reveals that the gutfeeling phenotype associated with P elements inserted in the 5'-UTR of SmD3 results from amorphic or strongly hypomorphic mutations. In contrast, P inserts in the SmD3 promoter region reduce the expression of SmD3 without abolishing it and produce larval lethality with overgrown imaginal discs, brain hemispheres, and hematopoietic organs. The lethality of these mutations could be rescued by an SmD3+ transgene. Finally, inactivation of AZ was obtained by complementing with SmD3+ the deficiency Df(2R)guf(lex47) that uncovers both SmD3 and AZ. Interestingly, AZ inactivation causes a new phenotype characterized by late larval lethality and atrophy of the brain, imaginal discs, hematopoietic organs, and salivary glands. PMID:12072471

  12. R2U2: Monitoring and Diagnosis of Security Threats for Unmanned Aerial Systems

    NASA Technical Reports Server (NTRS)

    Schumann, Johann; Moosbruger, Patrick; Rozier, Kristin Y.

    2015-01-01

    We present R2U2, a novel framework for runtime monitoring of security properties and diagnosing of security threats on-board Unmanned Aerial Systems (UAS). R2U2, implemented in FPGA hardware, is a real-time, REALIZABLE, RESPONSIVE, UNOBTRUSIVE Unit for security threat detection. R2U2 is designed to continuously monitor inputs from the GPS and the ground control station, sensor readings, actuator outputs, and flight software status. By simultaneously monitoring and performing statistical reasoning, attack patterns and post-attack discrepancies in the UAS behavior can be detected. R2U2 uses runtime observer pairs for linear and metric temporal logics for property monitoring and Bayesian networks for diagnosis of security threats. We discuss the design and implementation that now enables R2U2 to handle security threats and present simulation results of several attack scenarios on the NASA DragonEye UAS.

  13. RLIM interacts with Smurf2 and promotes TGF-{beta} induced U2OS cell migration

    SciTech Connect

    Huang, Yongsheng; Yang, Yang; Gao, Rui; Yang, Xianmei; Yan, Xiaohua; Wang, Chenji; Jiang, Sirui; Yu, Long

    2011-10-14

    Highlights: {yields} RLIM directly binds to Smurf2. {yields} RLIM enhances TGF-{beta} responsiveness in U2OS cells. {yields} RLIM promotes TGF-{beta} driven migration of osteosarcoma U2OS cells. -- Abstract: TGF-{beta} (transforming growth factor-{beta}), a pleiotropic cytokine that regulates diverse cellular processes, has been suggested to play critical roles in cell proliferation, migration, and carcinogenesis. Here we found a novel E3 ubiquitin ligase RLIM which can directly bind to Smurf2, enhancing TGF-{beta} responsiveness in osteosarcoma U2OS cells. We constructed a U2OS cell line stably over-expressing RLIM and demonstrated that RLIM promoted TGF-{beta}-driven migration of U2OS cells as tested by wound healing assay. Our results indicated that RLIM is an important positive regulator in TGF-{beta} signaling pathway and cell migration.

  14. Mutant U2AF1 Expression Alters Hematopoiesis and Pre-mRNA Splicing In Vivo

    PubMed Central

    Shirai, Cara Lunn; Ley, James N.; White, Brian S.; Kim, Sanghyun; Tibbitts, Justin; Shao, Jin; Ndonwi, Matthew; Wadugu, Brian; Duncavage, Eric J.; Okeyo-Owuor, Theresa; Liu, Tuoen; Griffith, Malachi; McGrath, Sean; Magrini, Vincent; Fulton, Robert S.; Fronick, Catrina; O’Laughlin, Michelle; Graubert, Timothy A.; Walter, Matthew J.

    2015-01-01

    SUMMARY Heterozygous somatic mutations in the spliceosome gene U2AF1 occur in ~11% of patients with myelodysplastic syndromes (MDS), the most common adult myeloid malignancy. It is unclear how these mutations contribute to disease. We examined in vivo hematopoietic consequences of the most common U2AF1 mutation using a doxycycline-inducible transgenic mouse model. Mice expressing mutant U2AF1(S34F) display altered hematopoiesis and changes in pre-mRNA splicing in hematopoietic progenitor cells by whole transcriptome analysis (RNA-seq). Integration with human RNA-seq datasets determined that common mutant U2AF1-induced splicing alterations are enriched in RNA processing genes, ribosomal genes, and recurrently-mutated MDS and acute myeloid leukemia-associated genes. These findings support the hypothesis that mutant U2AF1 alters downstream gene isoform expression, thereby contributing to abnormal hematopoiesis in MDS patients. PMID:25965570

  15. BRCA1 maps proximal to D17S579 on chromosome 17q21 by genetic analysis

    SciTech Connect

    Chamberlain, J.S.; Boehnke, M.; Frank, T.S.; Kiousis, S.; Junxhe Xu; Sunwei Guo; Hauser, E.R.; Helmbold, E.A.; Markel, D.S.; Keshavarzi, S.M.; Calzone, K.; Weber, B.L. ); Norum, R.A. ); Jackson, C.E. Henry Ford Hospital, Detroit, MI ); Garber, J. ); Collins, F.S. Howard Hughes Medical Inst., Ann Arbor, MI )

    1993-04-01

    Previous studies have demonstrated linkage between early-onset breast cancer and ovarian cancer and genetic markers on chromosome 17q21. These markers define the location of a gene (BRCA1) which appears to be inherited as an autosomal dominant susceptibility allele. The authors analyzed five families with multiple affected individuals for evidence of linkage to the BRCA1 region. Two of the five families appear to be linked to BRCA1. One apparently linked family contains critical recombinants, suggesting that the gene is proximal to the marker D17S579 (Mfd188). These findings are consistent with the maximum-likelihood position estimated by the Breast Cancer Linkage Consortium and with recombination events detected in other linked families. Linkage analysis was greatly aided by PCR-based analysis of paraffin-embedded normal breast tissue from deceased family members, demonstrating the feasibility and importance of this approach. One of the two families with evidence of linkage between breast cancer and genetic markers flanking BRCA1 represents the first such family of African-American descent to be reported in detail. 16 refs., 4 figs., 2 tabs.

  16. Patterns of missplicing due to somatic U2AF1 mutations in myeloid neoplasms

    PubMed Central

    Przychodzen, Bartlomiej; Jerez, Andres; Guinta, Kathryn; Sekeres, Mikkael A.; Padgett, Richard; Maciejewski, Jaroslaw P.

    2013-01-01

    Recently, recurrent mutations of spliceosomal genes were frequently identified in myeloid malignancies, as well as other types of cancers. One of these spliceosomal genes, U2AF1, was affected by canonical somatic mutations in aggressive type of myeloid malignancies. We hypothesized that U2AF1 mutations causes defects of splicing (missplicing) in specific genes and that such misspliced genes might be important in leukemogenesis. We analyzed RNA deep sequencing to compare splicing patterns of 201 837 exons between the cases with U2AF1 mutations (n = 6) and wild type (n = 14). We identified different alternative splicing patterns in 35 genes comparing cells with mutant and wild-type U2AF1. U2AF1 mutations are associated with abnormal splicing of genes involved in functionally important pathways, such as cell cycle progression and RNA processing. In addition, many of these genes are somatically mutated or deleted in various cancers. Of note is that the alternative splicing patterns associated with U2AF1 mutations were associated with specific sequence signals at the affected splice sites. These novel observations support the hypothesis that U2AF1 mutations play a significant role in myeloid leukemogenesis due to selective missplicing of tumor-associated genes. PMID:23775717

  17. The flavonoid luteolin enhances doxorubicin-induced autophagy in human osteosarcoma U2OS cells

    PubMed Central

    Zhang, Baoliang; Yu, Xin; Xia, Hong

    2015-01-01

    Luteolin (LUT), a flavone, which is universally present as constituent of medicinal plants as well as some vegetables and spices, has been demonstrated display specific anti-carcinogenic effects. However, the mechanisms by which LUT inhibits human osteosarcoma growth remain unknown. The effects of LUT on cell growth in human osteosarcoma U2OS cells were measured by MTT assay and flowcytometry. The effects of LUT on morphological markers of autophagy in U2OS were analyzed by fluorescence microscopy and electron microscopy. Autophagic markers, beclin1 and LC3 were detected by western blotting. Here, we found that LUT induced autophagy in U2OS and acted as an enhancer to sensitize doxorubicin (DOX)-mediated autophagy signaling. The combined treatment of LUT and DOX greatly decreases the growth of U2OS, showing synergistic cytotoxicity. Our results indicate that LUT in combination with DOX maybe a novel strategy for the treatment of human osteosarcoma. PMID:26629003

  18. Unlimited Horizons: Design and Development of the U-2. [NASA Aeronautics Book Series

    NASA Technical Reports Server (NTRS)

    Merlin, Peter W.

    2015-01-01

    This book describes the creation, history, design, and research value of the U-2 program. It also describes the involvement of NACA, as a cover story, and the later use by NASA of these aircraft as environmental research platforms.

  19. 12. WORK CREW REMOVING CROSS BRACE AT U2, PART OF ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    12. WORK CREW REMOVING CROSS BRACE AT U2, PART OF CONRAIL'S PROGRAM TO INCREASE CLEARANCE ON ITS BRIDGES AND OVERPASSES FOR DOUBLE-STACK TRAINS. - Conrail Port Perry Bridge, Spanning Monongahela River, Elizabeth, Allegheny County, PA

  20. Transcription on lampbrush chromosome loops in the absence of U2 snRNA.

    PubMed Central

    Tsvetkov, A; Jantsch, M; Wu, Z; Murphy, C; Gall, J G

    1992-01-01

    The five small nuclear RNAs (snRNAs) involved in splicing occur on the loops of amphibian lampbrush chromosomes and in hundreds to thousands of extrachromosomal granules called B snurposomes. To assess the role of these snRNAs during transcription and to explore possible relationships between the loops and B snurposomes, we injected single-stranded antisense oligodeoxynucleotides (oligos) against U1 and U2 snRNA into toad and newt oocytes. As shown before, antisense U1 and U2 oligos caused truncation of U1 and complete destruction of U2 snRNAs, respectively. However, injection of any oligo, regardless of sequence, brought on dramatic cytological changes, including shortening of the chromosomes and retraction of the lateral loops, with concomitant shutdown of polymerase II transcription, as well as disappearance of some or all of the B snurposomes. When injected oocytes were incubated for 12 h or longer in physiological saline, these changes were reversible; that is, the chromosomes lengthened, transcription (detected by 3H-UTP incorporation) resumed on newly extended lateral loops, and B snurposomes reappeared. In situ hybridization showed that loops and B snurposomes had negligible amounts of U2 snRNA after recovery from injection of the anti-U2 oligo, whereas these structures had normal levels of U2 snRNA after recovery from a control oligo. Thus, the morphological integrity of B snurposomes and lampbrush chromosome loops is not dependent on the presence of U2 snRNA. Because transcription occurs in the absence of U2 snRNA, we conclude that splicing is not required for transcription on lampbrush chromosome loops. Images PMID:1627829

  1. Target discrimination by RNA-binding proteins: role of the ancillary protein U2A' and a critical leucine residue in differentiating the RNA-binding specificity of spliceosomal proteins U1A and U2B".

    PubMed Central

    Rimmele, M E; Belasco, J G

    1998-01-01

    The spliceosomal proteins U1A and U2B" each use a homologous RRM domain to bind specifically to their respective snRNA targets, U1hpll and U2hpIV, two stem-loops that are similar yet distinct in sequence. Previous studies have shown that binding of U2B" to U2hpIV is facilitated by the ancillary protein U2A', whereas specific binding of U1A to U1hpll requires no cofactor. Here we report that U2A' enables U2B" to distinguish the loop sequence of U2hpIV from that of U1hpll but plays no role in stem sequence discrimination. Although U2A' can also promote heterospecific binding of U1A to U2hpIV, a much higher concentration of the ancillary protein is required due to the approximately 500-fold greater affinity of U2A' for U2B". Additional experiments have identified a single leucine residue in U1A(Leu-44) that is critical for the intrinsic specificity of this protein for the loop sequence of U1 hpll in preference to that of U2hpIV. Our data suggest that most of the difference in RNA-binding specificity between U1A and U2B" can be accounted for by this leucine residue and by the contribution of the ancillary protein U2A' to the specificity of U2B". PMID:9814759

  2. Thermal and transport properties of U2Pt x Ir1-x C2

    NASA Astrophysics Data System (ADS)

    Kang, Mingu; Wakeham, N.; Ni, Ni; Bauer, E. D.; Kim, Jeehoon; Ronning, F.

    2015-09-01

    We report thermal and transport properties of U2Pt x Ir1-x C2 from which a magnetic phase diagram is obtained. Pure U2IrC2 is an antiferromagnet at 6.5 K, whose Néel temperature initially rises to 13.2 K at x = 0.2 and subsequently is suppressed to zero temperature with increasing Pt content near x = 0.6. Heat capacity divided by temperature at x = 0.6 shows an upturn at low temperature, consistent with the expectations of enhanced quantum fluctuations in the presence of an underlying quantum critical point. The entropy after the phonon contribution has been subtracted has a value of 0.24 Rln2 at the Néel temperature of U2IrC2, revealing an itinerant nature of the 5 f electrons in this compound. On the Pt rich side of the phase diagram, superconductivity is suppressed by x = 0.85. The residual resistivity increases by a factor of 10 from pure Pt (x = 1) to x = 0.85 where superconductivity is suppressed to zero. By comparing the phase diagram of Ir doped U2PtC2 with the phase diagram of pressure tuned and Rh doped U2PtC2 we demonstrate the role of electronic tuning in this system.

  3. Thermal and transport properties of U2Pt(x)Ir(1-x)C2.

    PubMed

    Kang, Mingu; Wakeham, N; Ni, Ni; Bauer, E D; Kim, Jeehoon; Ronning, F

    2015-09-16

    We report thermal and transport properties of U2Pt x Ir1-x C2 from which a magnetic phase diagram is obtained. Pure U2IrC2 is an antiferromagnet at 6.5 K, whose Néel temperature initially rises to 13.2 K at x = 0.2 and subsequently is suppressed to zero temperature with increasing Pt content near x = 0.6. Heat capacity divided by temperature at x = 0.6 shows an upturn at low temperature, consistent with the expectations of enhanced quantum fluctuations in the presence of an underlying quantum critical point. The entropy after the phonon contribution has been subtracted has a value of 0.24 Rln2 at the Néel temperature of U2IrC2, revealing an itinerant nature of the 5 f electrons in this compound. On the Pt rich side of the phase diagram, superconductivity is suppressed by x = 0.85. The residual resistivity increases by a factor of 10 from pure Pt (x = 1) to x = 0.85 where superconductivity is suppressed to zero. By comparing the phase diagram of Ir doped U2PtC2 with the phase diagram of pressure tuned and Rh doped U2PtC2 we demonstrate the role of electronic tuning in this system. PMID:26302330

  4. Partial base-methylation and other structural differences in the 17 S ribosomal RNA of sycamore cells during growth in cell culture.

    PubMed

    Miassod, R; Cecchini, J P

    1979-04-26

    Sycamore (Acer pseudoplatanus L.) cytoplasmic rRNA was investigated in rapidly dividing cells, cells starting mitosis after the lag phase of growth (4 days) induced by deconditioning of the culture medium and also in growth-arrested cells from 10 day-old cultures deprived of exogenous auxin (i.e. exponential, early exponential and 2,4-dichlorophenoxyacetic acid (2,4-D)-deprived cultures). rRNA was extracted and purified from mixed 14C-labelled exponential cultures and 3H-labelled early exponential cultures. A 14C-labelled exponential culture and a 3H-labelled 2,4-D-deprived culture were analyzed in the same way. The 17 S rRNA molecules from both early exponential and 2,4-D-deprived cultures displayed a lower electrophoretic mobility on polyacrylamide gels than those from exponential cultures. Alkaline and acid hydrolysates of purified 17 S rRNA labelled on the phosphate groups or the methyl groups were analyzed on ion-exchange resins. There was no change in the extent of ribose methylation of the molecule from the three different cultures. However, the base methylation of the 17 S rRNA was decreased in early exponential cultures and in 2,4-D-deprived cultures. Part of the molecules synthesized in early exponential cultures specifically lacked 7-methylguanine, N6-methyladenine and N6,N6-dimethyladenine. The possible significance of these changes in the 17 S rRNA were discussed.

  5. Evaluation of the disposable absorption containment trunk for female U-2 and TR-1 pilots.

    PubMed

    Barlow, J F; Richardson, S E

    1991-06-01

    Female U-2 and TR-1 pilots needed a urine collection device as part of their high flight pressure suit ensembles. The Disposable Absorption Containment Trunk (DACT) had been designed for short term use with the NASA Space Shuttle Program. This study evaluated the DACT for the extended and repetitive use required for integration into the U-2 and TR-1 aircraft. Six female subjects were tested wearing a custom-fit DACT and the 1030 full pressure suit ensemble with torso harness. They were strapped into the U-2 ejection seat with parachute and seat kit for 10 h and 45 min every 3 d for a total of 5 sessions. The DACT was evaluated for absorptive capability and comfort. Subjects were monitored for dermatitis, vaginitis, and urinary tract infections. The DACT reliably absorbed menstrual flow and urinary outputs to 950 cc. Higher urinary outputs resulted in minimal leakage. Dermatitis in the form of mild erythema and chafing was present, but cleared between sessions without medical intervention. Complaints concerning comfort were minor and limited to fit problems. One case of vaginitis developed. There were no cases of urinary tract infection. The DACT was found to be an acceptable method of urine collection for female U-2 and TR-1 pilots. Minor changes in design will enhance the comfort and performance of the DACT. PMID:1859348

  6. Evaluation of the disposable absorption containment trunk for female U-2 and TR-1 pilots.

    PubMed

    Barlow, J F; Richardson, S E

    1991-06-01

    Female U-2 and TR-1 pilots needed a urine collection device as part of their high flight pressure suit ensembles. The Disposable Absorption Containment Trunk (DACT) had been designed for short term use with the NASA Space Shuttle Program. This study evaluated the DACT for the extended and repetitive use required for integration into the U-2 and TR-1 aircraft. Six female subjects were tested wearing a custom-fit DACT and the 1030 full pressure suit ensemble with torso harness. They were strapped into the U-2 ejection seat with parachute and seat kit for 10 h and 45 min every 3 d for a total of 5 sessions. The DACT was evaluated for absorptive capability and comfort. Subjects were monitored for dermatitis, vaginitis, and urinary tract infections. The DACT reliably absorbed menstrual flow and urinary outputs to 950 cc. Higher urinary outputs resulted in minimal leakage. Dermatitis in the form of mild erythema and chafing was present, but cleared between sessions without medical intervention. Complaints concerning comfort were minor and limited to fit problems. One case of vaginitis developed. There were no cases of urinary tract infection. The DACT was found to be an acceptable method of urine collection for female U-2 and TR-1 pilots. Minor changes in design will enhance the comfort and performance of the DACT.

  7. Synthesis of phase-pure U2N3 microspheres and its decomposition into UN.

    PubMed

    Silva, Chinthaka M; Hunt, Rodney D; Snead, Lance L; Terrani, Kurt A

    2015-01-01

    Uranium mononitride (UN) is important as a nuclear fuel. Fabrication of UN in its microspherical form also has its own merits since the advent of the concept of accident-tolerant fuel, where UN is being considered as a potential fuel in the form of TRISO particles. However, not many processes have been well established to synthesize kernels of UN. Therefore, a process for synthesis of microspherical UN with a minimum amount of carbon is discussed herein. First, a series of single-phased microspheres of uranium sesquinitride (U2N3) were synthesized by nitridation of UO2+C microspheres at a few different temperatures. Resulting microspheres were of low-density U2N3 and decomposed into low-density UN. The variation of density of the synthesized sesquinitrides as a function of its chemical composition indicated the presence of extra (interstitial) nitrogen atoms corresponding to its hyperstoichiometry, which is normally indicated as α-U2N3. Average grain sizes of both U2N3 and UN varied in a range of 1-2.5 μm. These also had a considerably large amount of pore spacing, indicating the potential sinterability of UN toward its use as a nuclear fuel.

  8. Ferromagnetic spin-glass behaviour in single-crystalline U2 IrSi3.

    PubMed

    Szlawska, M; Majewicz, M; Kaczorowski, D

    2014-03-26

    A single crystal of the U-based ternary silicide U(2)IrSi(3) was investigated by means of magnetic, resistivity and heat-capacity measurements performed in wide ranges of temperature and external magnetic fields. The results hint at the formation of a non-trivial magnetic ground state in which ferromagnetic ordering coexists with spin-glass freezing. PMID:24594881

  9. The Transcription Map of Human Papillomavirus Type 18 during Genome Replication in U2OS Cells

    PubMed Central

    Toots, Mart; Männik, Andres; Kivi, Gaily; Ustav, Mart; Ustav, Ene; Ustav, Mart

    2014-01-01

    The human osteosarcoma cell line U2OS is useful for studying genome replication of human papillomavirus (HPVs) subtypes that belong to different phylogenetic genera. In this study, we defined the HPV18 transcription map in U2OS cells during transient replication, stable maintenance and vegetative amplification by identifying viral promoter regions, transcription polyadenylation and splicing sites during HPV18 genome replication. Mapping of the HPV18 transcription start sites in U2OS cells revealed five distinct promoter regions (P102, P520, P811, P1193 and P3000). With the exception of P3000, all of these regions have been previously identified during productive HPV18 infection. Collectively, the data suggest that U2OS cells are suitable for studying the replication and transcription properties of HPVs and to serve as a platform for conducting high-throughput drug screens to identify HPV replication inhibitors. In addition, we have identified mRNA species that are initiated from the promoter region P3000, which can encode two E2C regulator proteins that contain only the C-terminal hinge and DNA-binding and dimerization domains of E2. We show that these proteins regulate the initial amplification of HPV18 by modulating viral transcription. Moreover, we show that one of these proteins can act as a transcriptional activator of promoter P102. PMID:25548925

  10. 16. DETAIL, VERTICAL MEMBER L2U2 FROM BELOW AND EAST. UNUSUALLY ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    16. DETAIL, VERTICAL MEMBER L2-U2 FROM BELOW AND EAST. UNUSUALLY SHAPED DESIGN REFLECTS COLUMN STRESSES ALSO NOTE LACING, WHICH COMPRISES ANGLES RATHER THAN COMMON FLAT BARS - Coraopolis Bridge, Spanning Ohio River back channel at Ferree Street & Grand Avenue, Coraopolis, Allegheny County, PA

  11. Synthesis of phase-pure U2N3 microspheres and its decomposition into UN

    DOE PAGES

    Silva, Chinthaka M.; Hunt, Rodney Dale; Snead, Lance Lewis; Terrani, Kurt A.

    2014-12-12

    Uranium mononitride (UN) is important as a nuclear fuel. Fabrication of UN in its microspherical form also has its own merits since the advent of the concept of accident-tolerant fuel, where UN is being considered as a potential fuel in the form of TRISO particles. But, not many processes have been well established to synthesize kernels of UN. Therefore, a process for synthesis of microspherical UN with a minimum amount of carbon is discussed herein. First, a series of single-phased microspheres of uranium sesquinitride (U2N3) were synthesized by nitridation of UO2+C microspheres at a few different temperatures. Resulting microspheres weremore » of low-density U2N3 and decomposed into low-density UN. The variation of density of the synthesized sesquinitrides as a function of its chemical composition indicated the presence of extra (interstitial) nitrogen atoms corresponding to its hyperstoichiometry, which is normally indicated as α-U2N3. Average grain sizes of both U2N3 and UN varied in a range of 1–2.5 μm. In addition, these had a considerably large amount of pore spacing, indicating the potential sinterability of UN toward its use as a nuclear fuel.« less

  12. The ternary germanides UMnGe and U2Mn3Ge

    NASA Astrophysics Data System (ADS)

    Hoffmann, Rolf-Dieter; Pöttgen, Rainer; Chevalier, Bernard; Gaudin, Etienne; Matar, Samir F.

    2013-07-01

    The title compounds were prepared by induction levitation melting of the elemental components and subsequent annealing. UMnGe (Pnma, a = 686.12(9), b = 425.49(6) and c = 736.5(1) pm) adopts the orthorhombic structure of TiNiSi and U2Mn3Ge (P63/mmc, a = 524.3(2) and c = 799.2(3) pm) possesses the hexagonal Mg2Cu3Si-type structure (ordered variant of the hexagonal Laves phase MgZn2). Both structures were refined from X-ray powder data to residuals of RI = 0.021 and 0.014 for UMnGe and U2Mn3Ge, respectively. The manganese and germanium atoms in UMnGe build up a three-dimensional [MnGe] network of ordered Mn3Ge3 hexagons with Mn-Ge distances ranging from 248 to 259 pm. The uranium atoms are coordinated by two tilted Mn3Ge3 hexagons. The manganese atoms in U2Mn3Ge build up Kagomé networks with 252 and 272 pm Mn-Mn distances which are connected via the germanium atoms (254 pm Mn-Ge) to a three-dimensional network. A remarkable feature of the U2Mn3Ge structure is a short U-U distance of 278 pm between adjacent cavities of the [Mn3Ge] network. From DFT based electronic structure calculations both germanides are found more cohesive than the Laves phase UMn2, thus underpinning the substantial role of Mn-Ge bonding. Calculations for both germanides show ferrimagnetic ground states with antiparallel spin alignments between U and Mn. The valence bands show bonding characteristics for interactions of atoms of different chemical natures and significant Mn-Mn bonding in U2Mn3Ge. Preliminary investigation of UMnGe by magnetization measurements confirms an antiferromagnetic arrangement below TN = 240 K.

  13. Hydrogen isotope effect on storage behavior of U2Ti and UZr2.3

    NASA Astrophysics Data System (ADS)

    Jat, Ram Avtar; Sawant, S. G.; Rajan, M. B.; Dhanuskar, J. R.; Kaity, Santu; Parida, S. C.

    2013-11-01

    U2Ti and UZr2.3 alloys were prepared by arc melting method, vacuum annealed and characterized by XRD, SEM and EDX methods. Hydrogen isotope effect on the storage behavior of these alloys were studied by measuring the hydrogen/deuterium desorption pressure-composition-temperature (PCT) profiles in the temperature range of 573-678 K using a Sievert's type volumetric apparatus. It was observed that, in the temperature and pressure range of investigation, all the isotherms show a single desorption plateau. The PCT data reveals that both U2Ti and UZr2.3 alloys had normal isotope effects on hydrogen/deuterium desorption at all experimental temperatures. Thermodynamic parameters for dehydrogenation and dedeuteration reactions of the corresponding hydrides and deuterides of the above alloys were deduced from the PCT data.

  14. Erosion Analysis for the Misaligned U2 Nozzle and its Connector Block

    SciTech Connect

    Lee, S.Y.

    2002-11-19

    In support of an erosion evaluation for the misaligned mating surfaces of the U2 nozzle and its connector block in the 3H evaporator lift-separator jumper, the Engineering Modeling and Simulation Group (EMSG) has developed computational models to identify potential sites of high erosion. Two mechanisms were considered to evaluate high erosion locations representative of the actual flow process in the misaligned U2 nozzle, abrasive erosion which occurs by high wall shear of viscous liquid film, and chip-off erosion which is mainly governed by particle impingement. The results show that primary locations of the highest erosion due to particle impingement are at the occurrence of sudden change of flow direction, sudden contraction, and flow obstruction as expected. Potential damage sites due to the abrasive wall erosion are at the upstream and downstream regions of sudden changes of flow direction.

  15. Cleavage-induced termination in U2 snRNA gene expression

    SciTech Connect

    Nabavi, Sadeq; Nazar, Ross N.

    2010-03-12

    The maturation of many small nuclear RNAs is dependent on RNase III-like endonuclease mediated cleavage, which generates a loading site for the exosome complex that trims the precursor at its 3' end. Using a temperature sensitive Pac1 nuclease, here we show that the endonuclease cleavage is equally important in terminating the transcription of the U2 snRNA in Schizosaccharomyces pombe. Using a temperature sensitive Dhp1p 5' {yields} 3' exonuclease, we demonstrate that it also is an essential component of the termination pathway. Taken together the results support a 'reversed torpedoes' model for the termination and maturation of the U2 snRNA; the Pac1 endonuclease cleavage provides entry sites for the 3' and 5' exonuclease activities, leading to RNA maturation in one direction and transcript termination in the other.

  16. Quantum geometry and quantization on U(u(2)) background. Noncommutative Dirac monopole

    NASA Astrophysics Data System (ADS)

    Gurevich, Dimitri; Saponov, Pavel

    2016-08-01

    In our previous publications we introduced differential calculus on the enveloping algebras U(gl(m)) similar to the usual calculus on the commutative algebra Sym (gl(m)) . The main ingredients of our calculus are quantum partial derivatives which turn into the usual partial derivatives in the classical limit. In the particular case m = 2 we prolonged this calculus on a central extension A of the algebra U(gl(2)) . In the present paper we consider the problem of a further extension of the quantum partial derivatives on the skew-field of the algebra A and define the corresponding de Rham complex. As an application of the differential calculus we suggest a method of transferring dynamical models defined on an extended Sym (u(2)) to an extended algebra U(u(2)) . We call this procedure the quantization with noncommutative configuration space. In this sense we quantize the Dirac monopole and find a solution of this model.

  17. The TOR signaling pathway regulates starvation-induced pseudouridylation of yeast U2 snRNA.

    PubMed

    Wu, Guowei; Radwan, Mohamed K; Xiao, Mu; Adachi, Hironori; Fan, Jason; Yu, Yi-Tao

    2016-08-01

    Pseudouridine (Ψ) has been identified in various types of RNAs, including mRNA, rRNA, tRNA, snRNA, and many other noncoding RNAs. We have previously shown that RNA pseudouridylation, like DNA and protein modifications, can be induced by stress. For instance, growing yeast cells to saturation induces the formation of Ψ93 in U2 snRNA. Here, we further investigate this inducible RNA modification. We show that switching yeast cells from nutrient-rich medium to different nutrient-deprived media (including water) results in the formation of Ψ93 in U2 snRNA. Using gene deletion/conditional depletion as well as rapamycin treatment, we further show that the TOR signaling pathway, which controls cell entry into stationary phase, regulates Ψ93 formation. The RAS/cAMP signaling pathway, which parallels the TOR pathway, plays no role in this inducible modification.

  18. The gene for pancreatic polypeptide (PPY) and the anonymous marker D17S78 are within 45 kb of each other on chromosome 17q21

    SciTech Connect

    Chandrasekharappa, S.C.; King, S.E.; Lee, Y.H.; Weber, B.L.; Collins, F.S. ); Friedman, L.; King, M.C. ); Welsch, P.; Bowcock, A.M. )

    1994-05-15

    A gene for early-onset breast and ovarian cancer (BRCA1) has been localized to a small region of chromosome 17q21. A combination of genetic linkage studies, radiation-reduced hybrid analysis, and physical mapping by FISH has identified several genes/markers that lie in this interval. Among these are the gene encoding pancreatic polypeptide (PPY) and a polymorphic marker at locus D17S78. Efforts to construct a physical map of this region by isolating a large number of yeast artificial chromosome (YAC) and cosmid clones demonstrate that PPY and D17S78 are present within the same cosmid clone, and therefore no farther than 45 kb apart. This observation takes on particular significance since it excludes a recently described BRCA1 candidate gene from the interval defined by meiotic mapping. Although PPY and D17S78 were found to be no farther than 45 kb apart, identification of a smaller fragment that hybridizes to both probes would indicate that these two are much closer. The probe p131 and the gene PPY were previously mapped to 17q21-q23 and to the proximal long arm of chromosome 17, respectively. The demonstration of the close proximity of these markers should allow them to be treated as a single locus in terms of long-range genomic mapping of this region, and the genomic clones isolated should serve as useful resources for the identification of the BRCA1 gene. Analysis of a large number of a familial and spordic breast and ovarian cancers has identified frequent loss of heterozygosity near the BRCA1 locus. A recent report has suggested the responsible interval lies just telomeric to PPY, and a suggested candidate gene (MCD) for BRCA1 was found to be somatically rearranged in two of several hundred sporadic breast tumors.

  19. High altitude perspective. [cost-reimbursable services using NASA U-2 aircraft

    NASA Technical Reports Server (NTRS)

    1978-01-01

    The capabilities of the NASA Ames Center U-2 aircraft for research or experimental programs are described for such areas as Earth resources inventories; remote sensing data interpretation, electronic sensor research and development; satellite investigative support; stratospheric gas studies; and astronomy and astrophysics. The availability of this aircraft on a cost-reimbursable basis for use in high-altitude investigations that cannot be performed by the private sector is discussed.

  20. RCRA Part A permit characterization plan for the U-2bu subsidence crater. Revision 1

    SciTech Connect

    1998-04-01

    This plan presents the characterization strategy for Corrective Action Unit (CAU) 109, U-2bu Subsidence Crater (referred to as U-2bu) in Area 2 at the Nevada Test Site (NTS). The objective of the planned activities is to obtain sufficient characterization data for the crater soils and observed wastes under the conditions of the current Resource Conservation and Recovery Act (RCRA) Part A permit. The scope of the characterization plan includes collecting surface and subsurface soil samples with hand augers and for the purpose of site characterization. The sampling strategy is to characterize the study area soils and look for RCRA constituents. Observable waste soils and surrounding crater soils will be analyzed and evaluated according to RCRA closure criteria. Because of the status of the crater a RCRA Part A permit site, acquired radionuclide analyses will only be evaluated in regards to the health and safety of site workers and the disposition of wastes generated during site characterization. The U-2bu Subsidence Crater was created in 1971 by a Lawrence Livermore National Laboratory underground nuclear test, event name Miniata, and was used as a land-disposal unit for radioactive and hazardous waste from 1973 to 1988.

  1. Closure plan for Corrective Action Unit 109: U-2bu subsidence crater, Nevada Test Site, Nevada

    SciTech Connect

    1999-03-01

    The U-2bu subsidence crater, Corrective Action Unit 109, will be closed in accordance with the Resource Conservation and Recovery Act, the Nevada Division of Environmental Protection operational permit, and the Federal Facility Agreement and Consent Order. The U-2bu subsidence crater is located in Area 2 of the Nevada Test Site. It was created in 1971 by an underground nuclear test with the name Miniata. The crater has a diameter of 288 meters (944 feet) and an approximate depth of 35 meters (115 feet). Based on the results of the analyses reported in the site characterization report, the only constituents of concern in the U-2bu subsidence crater include leachable lead and total petroleum hydrocarbons. Closure activities will include the excavation and disposal of impacted soil from the top of the crater. Upon completion of excavation, verification samples will be collected to show that the leachable lead has been removed to concentrations below the regulatory action level. After sample results show that the lead has been removed, the excavated area will be backfilled and a soil flood diversion berm will be constructed as a best management practice. An independent registered professional engineer will certify the site was closed following the approved Closure Plan. Post-closure care is not warranted for this site because closure activities will involve removal of the Resource Conservation and Recovery Act constituents of concern.

  2. X-ray Structures of U2 snRNA-Branchpoint Duplexes Containing Conserved Pseudouridines

    SciTech Connect

    Lin,Y.; Kielkopf, C.

    2008-01-01

    A pseudouridine-modified region of the U2 small nuclear (sn)RNA anneals with the intronic branchpoint sequence and positions a bulged adenosine to serve as the nucleophile in the first chemical step of pre-mRNA splicing. We have determined three X-ray structures of RNA oligonucleotides containing the pseudouridylated U2 snRNA and the branchpoint consensus sequences. The expected adenosine branchpoint is extrahelical in a 1.65 Angstroms resolution structure containing the mammalian consensus sequence variant and in a 2.10 Angstroms resolution structure containing a shortened Saccharomyces cerevisiae consensus sequence. The adenosine adjacent to the expected branchpoint is extrahelical in a third structure, which contains the intact yeast consensus sequence at 1.57 Angstroms resolution. The hydration and base stacking interactions mediated by the U2 snRNA pseudouridines correlate with the identity of the unpaired adenosine. The expected adenosine bulge is associated with a well-stacked pseudouridine, which is linked via an ordered water molecule to a neighboring nucleotide. In contrast, the bulge of the adjacent adenosine shifts the base stacking and disrupts the water-mediated interactions of the pseudouridine. These structural differences may contribute to the ability of the pseudouridine modification to promote the bulged conformation of the branch site adenosine and to enhance catalysis by snRNAs. Furthermore, iodide binding sites are identified adjacent to the unconventional bulged adenosine, and the structure of the mammalian consensus sequence variant provides a high-resolution view of a hydrated magnesium ion bound in a similar manner to a divalent cation binding site of the group II intron.

  3. Sp6<=crSscrU2 symmetry of the fermion dynamical symmetry model

    NASA Astrophysics Data System (ADS)

    Chen, Jin-Quan; Chen, Xuan-Gen; Feng, Da Hsuan; Wu, Cheng-Li; Ginocchio, Joseph N.; Guidry, Mike W.

    1989-12-01

    The SP6×scrSscrU2 symmetry of the fermion dynamical symmetry model (FDSM) was studied. Analytic expressions are given for the wave functions and matrix elements of the pair creation operators S°, D°, and E2 transition operator for the heritage u=0, 1, 2 cases. Various Pauli effects which differentiate the FDSM SU3 from IBM SU3, as well as the relationship between the wave functions and matrix elements in the FDSM, the interacting boson model, the Sp (6,R) models, and the Elliott model are discussed. Extensive discussions of the physical implications of the model are presented.

  4. Pseudouridines in U2 snRNA stimulate the ATPase activity of Prp5 during spliceosome assembly.

    PubMed

    Wu, Guowei; Adachi, Hironori; Ge, Junhui; Stephenson, David; Query, Charles C; Yu, Yi-Tao

    2016-03-15

    Pseudouridine (Ψ) is the most abundant internal modification identified in RNA, and yet little is understood of its effects on downstream reactions. Yeast U2 snRNA contains three conserved Ψs (Ψ35, Ψ42, and Ψ44) in the branch site recognition region (BSRR), which base pairs with the pre-mRNA branch site during splicing. Here, we show that blocks to pseudouridylation at these positions reduce the efficiency of pre-mRNA splicing, leading to growth-deficient phenotypes. Restoration of pseudouridylation at these positions using designer snoRNAs results in near complete rescue of splicing and cell growth. These Ψs interact genetically with Prp5, an RNA-dependent ATPase involved in monitoring the U2 BSRR-branch site base-pairing interaction. Biochemical analysis indicates that Prp5 has reduced affinity for U2 snRNA that lacks Ψ42 and Ψ44 and that Prp5 ATPase activity is reduced when stimulated by U2 lacking Ψ42 or Ψ44 relative to wild type, resulting in inefficient spliceosome assembly. Furthermore, in vivo DMS probing analysis reveals that pseudouridylated U2, compared to U2 lacking Ψ42 and Ψ44, adopts a slightly different structure in the branch site recognition region. Taken together, our results indicate that the Ψs in U2 snRNA contribute to pre-mRNA splicing by directly altering the binding/ATPase activity of Prp5. PMID:26873591

  5. Closure Plan for Corrective Action Unit 109: U-2bu Subsidence Crater Nevada Test Site, Nevada

    SciTech Connect

    Shannon Parsons

    1999-03-01

    The U-2bu subsidence crater, Corrective Action Unit 109, will be closed in accordance with the Resource Conservation and Recovery Act, the Nevada Division of Environmental Protection operational permit, and the Federal Facilities Agreement and Consent Order. The U-2bu subsidence crater is located in Area 2 of the Nevada Test Site. It was created in 1971 by an underground nuclear test with the name Miniata. The crater has a diameter of 288 meters (944 feet) and an approximate depth of 35 meters (115 feet). The subsidence crater was used as a land disposal unit for radioactive and hazardous waste from 1973 to 1988. Site disposal history is supported by memorandums, letters, and personnel who worked at the Nevada Test Site at the time of active disposal. Closure activities will include the excavation and disposal of impacted soil form the tip of the crater. Upon completion of excavation, verification samples will be collected to show that lead has been removed to concentrations be low regulatory action level. The area will then be backfilled and a soil flood diversion berm will be constructed, and certified by an independent professional engineer as to having followed the approved Closure Plan.

  6. Subjective experiences of watching stereoscopic Avatar and U2 3D in a cinema

    NASA Astrophysics Data System (ADS)

    Pölönen, Monika; Salmimaa, Marja; Takatalo, Jari; Häkkinen, Jukka

    2012-01-01

    A stereoscopic 3-D version of the film Avatar was shown to 85 people who subsequently answered questions related to sickness, visual strain, stereoscopic image quality, and sense of presence. Viewing Avatar for 165 min induced some symptoms of visual strain and sickness, but the symptom levels remained low. A comparison between Avatar and previously published results for the film U2 3D showed that sickness and visual strain levels were similar despite the films' runtimes. The genre of the film had a significant effect on the viewers' opinions and sense of presence. Avatar, which has been described as a combination of action, adventure, and sci-fi genres, was experienced as more immersive and engaging than the music documentary U2 3D. However, participants in both studies were immersed, focused, and absorbed in watching the stereoscopic 3-D (S3-D) film and were pleased with the film environments. The results also showed that previous stereoscopic 3-D experience significantly reduced the amount of reported eye strain and complaints about the weight of the viewing glasses.

  7. Ground state of the U2Mo compound: Physical properties of the Ω-phase

    NASA Astrophysics Data System (ADS)

    Losada, E. L.; Garcés, J. E.

    2016-10-01

    Using ab initio calculations, unexpected structural instability was recently found in the ground state of the U2 Mo compound. Instead of the unstable I4/mmm and the Pmmn structures, in this work the P6/mmm (#191) space group, usually called Ω-phase, is proposed as the fundamental state. Total energy calculations using Wien2k code slightly favoured the last structure. Electronic and elastic properties are studied in this work in order to characterize the physical properties of this new phase. The stability of the Ω-phase is studied by means of its elastic constants calculation and phonon dispersion spectrum. Analysis of isotropic indices shows that the new phase is a ductile material with a minimal degree of anisotropy, suggesting that U2 Mo in the P6/mmm structure is an elastic isotropic material. Analysis of charge density, density of electronic states (DOS) and the character of the bands revealed a high level of hybridization between d-molybdenum electronic states and d- and f-uranium ones.

  8. U-2 with fictitious NASA markings to support CIA cover story for pilot Gary Powers, shot down over S

    NASA Technical Reports Server (NTRS)

    1960-01-01

    After Francis Gary Powers was shot down over the Soviet Union during a CIA spy flight on 1 May 1960, NASA issued a press release with a cover story about a U-2 conducting weather research that may have strayed off course after the pilot 'reported difficulties with his oxygen equipment.' To bolster the cover-up, a U-2 was quickly painted in NASA markings, with a fictitious NASA serial number, and put on display for the news media at the NASA Flight Research Center at Edwards Air Force Base on 6 May 1960. The next day, Soviet Premier Nikita Kruschev exposed the cover-up by revealing that the pilot had been captured, and espionage equipment had been recovered from the wreckage. 7 May 1956 - NACA Director Dr. Hugh L. Dryden issues a press release stating that U-2 aircraft are conducting weather research for NACA with Air Force support from Watertown, Nevada. 22 May 1956 - A second press release is issued with cover story for U-2 aircraft operating overseas. 1 May 1960 - Francis Gary Powers is shot down near Sverdlovsk. 6 May 1960 - U-2 with fictitious NASA serial number and NASA markings is shown to news media to bolster cover story of NASA weather research flights with U-2. 7 May 1960 - Soviet Premier Kruschev announces capture and confession of Powers. 1960 - Dr. Hugh L. Dryden tells senate committee that some 200 U-2 flights carrying NASA weather instrumentation have taken place since 1956. 2 April 1971 - NASA receives two U-2C aircraft for high-altitude research.

  9. Antibacterial Activity of Elephant Garlic and Its Effect against U2OS Human Osteosarcoma Cells

    PubMed Central

    Huang, Zehao; Ren, Jianwu

    2013-01-01

    Objective(s): The present study was designed to investigate the antibacterial function and pharmacological effect of elephant garlic (Allium ampeloprasum var. ampeloprasum) on U2OS human osteosarcoma cells. Materials and Methods: Seven kinds of bacteria were reconstituted, inoculated and tested in this research to evaluate elephant garlic antibacterial activity. By the means of FACS analysis, cell proliferation assay, confocal laser scanning microscopy and Transwell migration assays, the effect of elephant garlic against U2OS human osteosarcoma cells was unveiled. Rerults: The antimicrobial activity of elephant garlic was stronger than ampicillin when used against Escherichia coli, Bacillus subtilis, Bacillus thuringiensis, Staphylococcus actinomycetes, and gray actinomycetes. Even at a very low concentration (12.5%), elephant garlic still had an antibacterial effect on common bacteria E. coli and S. aureus. The G0/G1 ratio of elephant garlic treated group cells increased while S phase decreased. Elephant garlic extract inhibited the growth of human osteosarcoma cells, U2OS, through preventing the transition from G1 phase to S phase. It reduced osteosarcoma cell, U2OS, invasion ability and significantly increased the proportion of apoptosis. It significantly affected the cytoskeleton generation. Conclusion: Elephant garlic exhibits antibacterial property and has an inhibitory effect on osteosarcoma cells (U2OS) proliferation and cell activity, suggesting the mechanism of its anticancer effects on U2OS human osteosarcoma cells. PMID:24379966

  10. The splicing factor U2AF65 is functionally conserved in the thermotolerant deep-sea worm Alvinella pompejana.

    PubMed

    Henscheid, Kristy L; Shin, David S; Cary, S Craig; Berglund, J Andrew

    2005-03-10

    Due to their inherent stability, thermophilic bacteria and archaea serve as important resources for biochemical and biophysical analyses of many biological processes. Unfortunately, scientists characterizing eukaryote-specific processes, such as nuclear pre-mRNA splicing, are unable to take advantage of these sources of thermostable proteins. To identify and provide a source of thermostable eukaryotic proteins, we are characterizing splicing factors in the thermotolerant deep-sea vent polychaete, Alvinella pompejana. This worm, also known as the Pompeii worm, is found in the extreme environment of deep-sea hydrothermal vents, and is one of the most thermotolerant eukaryotic organisms known. We report on detailed analyses of U2AF65, the large subunit of the U2 small nuclear ribonucleoprotein auxiliary factor, an essential splicing factor important for intron definition and alternative splicing. The cloning and characterization of Pompeii U2AF65 show it is highly similar to human U2AF65 in sequence and function and is more thermostable than the human protein when bound to RNA in vitro. Notably, Pompeii U2AF65 can restore splicing in a human extract depleted of human U2AF. We also determine that the general splicing mechanisms and signal sequences are conserved in the Pompeii worm, an annelid which has previously been uncharacterized in terms of splicing factors and signals.

  11. RNA structure replaces the need for U2AF2 in splicing

    PubMed Central

    Lin, Chien-Ling; Taggart, Allison J.; Lim, Kian Huat; Cygan, Kamil J.; Ferraris, Luciana; Creton, Robbert; Huang, Yen-Tsung; Fairbrother, William G.

    2016-01-01

    RNA secondary structure plays an integral role in catalytic, ribosomal, small nuclear, micro, and transfer RNAs. Discovering a prevalent role for secondary structure in pre-mRNAs has proven more elusive. By utilizing a variety of computational and biochemical approaches, we present evidence for a class of nuclear introns that relies upon secondary structure for correct splicing. These introns are defined by simple repeat expansions of complementary AC and GT dimers that co-occur at opposite boundaries of an intron to form a bridging structure that enforces correct splice site pairing. Remarkably, this class of introns does not require U2AF2, a core component of the spliceosome, for its processing. Phylogenetic analysis suggests that this mechanism was present in the ancestral vertebrate lineage prior to the divergence of tetrapods from teleosts. While largely lost from land dwelling vertebrates, this class of introns is found in 10% of all zebrafish genes. PMID:26566657

  12. Complete Elastic Tensor through the First-Order Transformation in U2Rh3Si5

    NASA Astrophysics Data System (ADS)

    Leisure, R. G.; Kern, S.; Drymiotis, F. R.; Ledbetter, H.; Migliori, A.; Mydosh, J. A.

    2005-08-01

    The complete elastic tensor of U2Rh3Si5 has been determined over the temperature range of 5 300 K, including the dramatic first-order transition to an antiferromagnetic state at 25.5 K. Sharp upward steps in the elastic moduli as the temperature is decreased through the transition reveal the first-order nature of the phase change. In the antiferromagnetic state the temperature dependence of the elastic moduli scales with the square of the ordered moment on the uranium ion, demonstrating strong spin-lattice coupling. The temperature dependence of the moduli well above the transition indicates coupling of the ultrasonic waves to the crystal electric field levels of the uranium ion where the lowest state is a singlet. The elastic constant data suggest that the first-order phase change is magnetically driven by a bootstrap mechanism involving the ground state singlet and a magnetically active crystal electric field level.

  13. Structural/functional properties of a mammalian multi-component structure containing all major spliceosomal small nuclear ribonucleoprotein particles.

    PubMed Central

    Moraitou, M; Patrinou-Georgoula, M; Guialis, A

    1998-01-01

    An approx. 40 S multi-component structure, consisting of all major spliceosomal small nuclear ribonucleoprotein particles (snRNP) (U1, U2, U4/U6 and U5) in stable association with a large number of polypeptides, mainly in the range 50-210 kDa, has been reported to exist within rat liver nuclear extracts [Guialis, Moraitou, Patrinou-Georgoula and Dangli (1991) Nucleic Acids Res. 19, 287-296]. Using a new polyclonal antibody recognizing a 63 kDa protein component of the complex, this multi-snRNP assembly was detected within rat liver nuclear extracts as efficiently as with the antibody for the U2 snRNP-specific B' polypeptide. The 63 kDa protein was found to correspond to the 66 kDa subunit of the splicing factor SF3a, a known integral component of the HeLa 17 S U2 snRNP. Anti-2,2,7-trimethylguanosine affinity chromatography was an easy and efficient way of purifying the multi-snRNP complex from rat liver 40 S heterogeneous nuclear ribonucleoprotein particle (hnRNP)-containing sucrose gradient fractions. By subsequent glycerol-gradient sedimentation, all known snRNP forms active in RNA splicing were identified among its constituents. A complex structurally similar to the rat multi-snRNP was also identified in HeLa nuclear extracts. Preservation of hnRNP-snRNP interactions was observed within HeLa 40 S fractions. Moreover, these fractions were capable of restoring splicing activity when applied in reconstitution studies to supplement a micrococcal nuclease-treated splicing extract. PMID:9576861

  14. IL-17s adopt a cystine knot fold: structure and activity of a novel cytokine, IL-17F, and implications for receptor binding

    PubMed Central

    Hymowitz, Sarah G.; Filvaroff, Ellen H.; Yin, JianPing; Lee, James; Cai, Liping; Risser, Philip; Maruoka, Miko; Mao, Weiguang; Foster, Jessica; Kelley, Robert F.; Pan, Guohua; Gurney, Austin L.; de Vos, Abraham M.; Starovasnik, Melissa A.

    2001-01-01

    The proinflammatory cytokine interleukin 17 (IL-17) is the founding member of a family of secreted proteins that elicit potent cellular responses. We report a novel human IL-17 homolog, IL-17F, and show that it is expressed by activated T cells, can stimulate production of other cytokines such as IL-6, IL-8 and granulocyte colony-stimulating factor, and can regulate cartilage matrix turnover. Unexpectedly, the crystal structure of IL-17F reveals that IL-17 family members adopt a monomer fold typical of cystine knot growth factors, despite lacking the disulfide responsible for defining the canonical ‘knot’ structure. IL-17F dimerizes in a parallel manner like neurotrophins, and features an unusually large cavity on its surface. Remarkably, this cavity is located in precisely the same position where nerve growth factor binds its high affinity receptor, TrkA, suggesting further parallels between IL-17s and neurotrophins with respect to receptor recognition. PMID:11574464

  15. Useful to Usable (U2U): Transforming Climate Variability and Change Information for Cereal Crop Producers

    NASA Astrophysics Data System (ADS)

    Niyogi, D.; Andresen, J.

    2011-12-01

    Corn and soybean production contributes over $100 billion annually to the U.S. economy, most of which comes from the intensely cultivated corn-belt region of the Midwest. Successful crop production in this region is highly dependent on favorable temperatures and appropriate precipitation patters, making this industry vulnerable to changes in climate patterns. Though predictive models are constantly improving, there are gaps in our understanding of how different management practices can be used to help farmers adapt to changes in climate while maintaining economic viability. Furthermore, currently available tools and models are not meeting producers' needs, and little is known about the types of information they would like to access. Useful to Usable (U2U): Transforming Climate Variability and Change Information for Cereal Crop Producers is an integrated research and extension project that seeks to improve the resilience and profitability of farms in the North Central Region amid variable climate change through the development and dissemination of improved decision support tools, resource materials, and training. The goal is to work closely with producers to help them make better long-term plans on what, when and where to plant, and also how to manage crops for maximum yields and minimum environmental damage. The U2U team is composed of a uniquely qualified group of climatologists, crop modelers, agronomists, economists, and social scientists from 10 partner universities across the Midwest. Over the span of 5 years, collaborators will complete tasks associated with 5 objectives that will enhance the usability of climate information for the agricultural community and lead to more sustainable farming operations. First the team will produce research on the biophysical and economic impacts of different climate scenarios on corn and soybean yields in the North Central Region (objective 1) and conduct complementary research to understand how producers and advisors are

  16. CD82 suppresses CD44 alternative splicing-dependent melanoma metastasis by mediating U2AF2 ubiquitination and degradation

    PubMed Central

    Fu, Ailing; Zhu, Huifeng; Ren, Qiao; Wang, Bochu; Xu, Xingran; Bai, Huiyuan; Dong, Cheng

    2016-01-01

    Melanoma is one of the most lethal forms of skin cancer due to its early metastatic spread. The variant form of CD44 (CD44v), a cell surface glycoprotein, is highly expressed on metastatic melanoma. The mechanisms of regulation of CD44 alternative splicing in melanoma and its pathogenic contributions are so far poorly understood. Here, we investigated the expression level of CD44 in a large set of melanocytic lesions at different stages. We found that the expression of CD44v8-10 and a splicing factor, U2AF2, is significantly increased during melanoma progression, while CD82/KAI1, a tetraspanin family of tumor suppressor, is reduced in metastatic melanoma. CD44v8-10 and U2AF2 expressions which are negatively correlated with CD82 levels are dramatically elevated in primary melanoma compared with dysplastic nevi and further increased in metastatic melanoma. We also showed that patients with higher CD44v8-10 and U2AF2 expression levels tended to have shorter survival. By using both in vivo and in vitro assays, we demonstrated that CD82 inhibits the production of CD44v8-10 on melanoma. Mechanistically, U2AF2 is a downstream target of CD82 and in malignant melanoma facilitates CD44v8-10 alternative splicing. U2AF2-mediated CD44 isoform switch is required for melanoma migration in vitro and lung and liver metastasis in vivo. Notably, overexpression of CD82 suppresses U2AF2 activity by inducing U2AF2 ubiquitination. In addition, our data suggested that enhancement of melanoma migration by U2AF2-dependent CD44v8-10 splicing is mediated by Src/FAK/RhoA activation and formation of stress fibers as well as CD44-E-selectin binding reinforcement. These findings uncovered a hitherto unappreciated function of CD82 in severing the linkage between U2AF2-mediated CD44 alternative splicing and cancer aggressiveness, with potential prognostic and therapeutic implications in melanoma. PMID:27041584

  17. An extended U2AF65–RNA-binding domain recognizes the 3′ splice site signal

    PubMed Central

    Agrawal, Anant A.; Salsi, Enea; Chatrikhi, Rakesh; Henderson, Steven; Jenkins, Jermaine L.; Green, Michael R.; Ermolenko, Dmitri N.; Kielkopf, Clara L.

    2016-01-01

    How the essential pre-mRNA splicing factor U2AF65 recognizes the polypyrimidine (Py) signals of the major class of 3′ splice sites in human gene transcripts remains incompletely understood. We determined four structures of an extended U2AF65–RNA-binding domain bound to Py-tract oligonucleotides at resolutions between 2.0 and 1.5 Å. These structures together with RNA binding and splicing assays reveal unforeseen roles for U2AF65 inter-domain residues in recognizing a contiguous, nine-nucleotide Py tract. The U2AF65 linker residues between the dual RNA recognition motifs (RRMs) recognize the central nucleotide, whereas the N- and C-terminal RRM extensions recognize the 3′ terminus and third nucleotide. Single-molecule FRET experiments suggest that conformational selection and induced fit of the U2AF65 RRMs are complementary mechanisms for Py-tract association. Altogether, these results advance the mechanistic understanding of molecular recognition for a major class of splice site signals. PMID:26952537

  18. Unusual 5f magnetism in the U2Fe3Ge ternary Laves phase: a single crystal study

    NASA Astrophysics Data System (ADS)

    Henriques, M. S.; Gorbunov, D. I.; Waerenborgh, J. C.; Havela, L.; Shick, A. B.; Diviš, M.; Andreev, A. V.; Gonçalves, A. P.

    2013-02-01

    Magnetic properties of the intermetallic compound U2Fe3Ge were studied on a single crystal. The compound crystallizes in the hexagonal Mg2Cu3Si structure, an ordered variant of the MgZn2 Laves structure (C14). U2Fe3Ge displays ferromagnetic order below the Curie temperature TC = 55 K and presents an exception to the Hill rule, as the nearest inter-uranium distances do not exceed 3.2 Å. Magnetic moments lie in the basal plane of the hexagonal lattice, with the spontaneous magnetic moment Ms = 1.0 μB/f.u. at T = 2 K. No anisotropy within the basal plane is detected. In contrast to typical U-based intermetallics, U2Fe3Ge exhibits very low magnetic anisotropy, whose field does not exceed 10 T. The dominance of U in the magnetism of U2Fe3Ge is suggested by the 57Fe Mössbauer spectroscopy study, which indicates very low or even zero Fe moments. Electronic structure calculations are in agreement with the observed easy-plane anisotropy but fail to explain the lack of an Fe contribution to the magnetism of U2Fe3Ge.

  19. Facile CO Cleavage by a Multimetallic CsU2 Nitride Complex.

    PubMed

    Falcone, Marta; Kefalidis, Christos E; Scopelliti, Rosario; Maron, Laurent; Mazzanti, Marinella

    2016-09-26

    Uranium nitrides are important materials with potential for application as fuels for nuclear power generation, and as highly active catalysts. Molecular nitride compounds could provide important insight into the nature of the uranium-nitride bond, but currently little is known about their reactivity. In this study, we found that a complex containing a nitride bridging two uranium centers and a cesium cation readily cleaved the C≡O bond (one of the strongest bonds in nature) under ambient conditions. The product formed has a [CsU2 (μ-CN)(μ-O)] core, thus indicating that the three cations cooperate to cleave CO. Moreover, the addition of MeOTf to the nitride complex led to an exceptional valence disproportionation of the CsU(IV) -N-U(IV) core to yield CsU(III) (OTf) and [MeN=U(V) ] fragments. The important role of multimetallic cooperativity in both reactions is illustrated by the computed reaction mechanisms. PMID:27596570

  20. Ultracold atoms in U(2) non-Abelian gauge potentials preserving the Landau levels

    SciTech Connect

    Burrello, Michele; Trombettoni, Andrea

    2011-10-15

    We study ultracold atoms subjected to U(2) non-Abelian potentials: we consider gauge potentials having, in the Abelian limit, degenerate Landau levels and we then investigate the effect of general homogeneous non-Abelian terms. The conditions under which the structure of degenerate Landau levels is preserved are classified and discussed. The typical gauge potentials preserving the Landau levels are characterized by a fictitious magnetic field and by an effective spin-orbit interaction (e.g., obtained through the rotation of two-dimensional atomic gases coupled with a tripod scheme). The single-particle energy spectrum can be analytically determined for a class of gauge potentials, whose physical implementation is discussed. The corresponding Landau levels are deformed by the non-Abelian contribution of the potential and their spin degeneracy is split. The related deformed quantum Hall states for fermions and bosons (in the presence of strong intraspecies interaction) are determined far from and at the degeneracy points of the Landau levels, where non-Abelian states appear. We present a discussion of the effect of the angular momentum, as well as results for U(3) gauge potentials.

  1. An Atlas of extraterrestrial particles collected with NASA U-2 aircraft, 1974 - 1976

    NASA Technical Reports Server (NTRS)

    Brownlee, D. E.; Tomandl, D.; Blanchard, M. B.; Ferry, G. V.; Kyte, F.

    1976-01-01

    Extraterrestrial particles collected during U-2 flights in the stratosphere were divided into four groups: chondritic, iron-sulfur--nickel, mafic silicates, and others. The chondritic aggregates are typically composed of Fe, Mg, Si, C, S, Ca, and Ni. Detectable levels of He-4 implanted from the solar wind occur in some. Olivine, spinel, and possibly pyrrhotite and a hydrated layered-lattice silicate were identified. The chondritic ablation particles contain no sulfur and appear to have been melted. Magnetite, olivine, and pyroxene were identified. The iron-sulfur-nickel type particles resemble meteoritic iron sulfide with a small amount of nickel, and contain magnetite and troilite. The mafic silicate type particles are iron magnesium silicate grains with clumps of chondritic aggregate particles adhering to their surfaces. Olivine and possibly pyrrhotite and pyroxene were identified. Most of the iron-nickel type particles are spherules and include taenite and wustite. The other type particles include nickel-iron mounds on spheroidal glassy-like grains having chondritic-like elemental abundances.

  2. Description of work for 216-U-1 and 216-U-2 stainless steel pipeline integrity testing

    SciTech Connect

    Wasemiller, M.A.

    1994-06-30

    The objectives of this integrity test are to (1) inspect the interior of this pipeline by in-line camera survey and (2) if required, conduct a pressure test on a section of the pipeline. The U-1 and U-2 Cribs were constructed in 1951. From March 1952 to June 1967, the site received cell drainage from Tank 5-2 in the 221-U Building nd waste from the 224-U Building via the overflow from the 241-U-361 Settling Tank. From June 1957 to July 1957, the site received waste from the 224-U Building via the overflow from the 241-U-361 Settling Tank and contaminated solvent from the 276-U Settling Tank solvent storage area. The discharge of 221-U waste was discontinued during shutdown of production operations. From July 1957 to May 1967, the site received waste from the 224-U Building and equipment decontamination and reclamation wastes from operations in the 221-U Building canyon. The scope of work is encompassed in five steps: (1) obtaining access to the pipeline in order to perform an in-line camera survey of the line to the greatest extent possible, (2) evaluating the need for further investigation of the pipeline, (3) blanking the line, as needed, to perform a pressure test, (4) conducting the pressure test, as needed, and (5) documenting the ability of the line to maintain pressure.

  3. First Principles Study of a-U2N3 Surfaces and its Anti-oxidant Mechanism

    NASA Astrophysics Data System (ADS)

    Jin, Mengting; Zhang, Yanning

    With the advantages over oxides as fuel materials for fast nuclear reactors, actinide nitrides have been extensively studied in experiments. In particular, a-U2N3 is also the main composition of surface layer obtained by surface nitriding for the enhancement of oxidation resistance of uranium in ambient conditions.[1-4] However, the anti-oxidant mechanism behind is still unclear, which hinders the further development of surface treatment technologies for uranium. Here we perform extensive ab initio studies on the geometric, magnetic and electronic properties of a-U2N3 bulk and (001) surfaces. Then the adsorption and diffusion of O2 near the stable a-U2N3(001) surface will be discussed, focusing on the local atomic arrangements of U-N and U-N-O that cannot be observed easily in experiments. Our theoretical results may give some insights in understanding the anti-oxidant mechanism of surface nitriding.

  4. Genetic polymorphism study at four minisatellite loci (D1S80, D17S5, D19S20, and APOB) among five Indian population groups.

    PubMed

    Das, Birajalaxmi; Ghosh, Anu; Chauhan, P S; Seshadri, M

    2002-06-01

    The present study reports the genetic variation observed among five anthropologically distinct population groups of India, using four highly polymorphic minisatellite loci (D1S80, D17S5, D19S20, and APOB 3' VNTR) in order to examine the effect of geographical and linguistic affiliations on the genetic affinities among these groups. Random individuals from five ethnic groups were studied; the sample size ranged from 235 to 364. The population groups belong to two geographically separated regions of India, the state of Maharashtra (western India) and the state of Kerala (southern India). The two Maharashtrian groups (Konkanastha Brahmins and Marathas) speak "Marathi," an Indo-European language, whereas the three Kerala population groups (Nairs, Ezhavas, and Muslims) speak "Malayalam," an Indo-Dravidian language. Genomic DNA was extracted from peripheral blood samples and analyzed using amplified fragment length polymorphism (Amp-FLP) technique. All four loci displayed high heterozygosity with average heterozygosity in the range of 0.82 to 0.84. The Polymorphic Information Content and Power of Discrimination were > or = 0.75 and > or = 0.80, respectively. The coefficient of gene differentiation was found to be low (average G(ST) = 1.2%; range between 0.6% at D1S80 locus to 1.6% at APOB 3' VNTR locus) across the loci, indicating close affinity among the population groups. The neighbor-joining tree revealed two clear clusters, one for the two Maharashtrian population groups and the other for the three Kerala population groups. The results obtained are in conformity with the geographical and linguistic backgrounds of the studied populations.

  5. On the oscillator realization of conformal U(2, 2) quantum particles and their particle-hole coherent states

    SciTech Connect

    Calixto, M. Pérez-Romero, E.

    2014-08-01

    We revise the unireps. of U(2, 2) describing conformal particles with continuous mass spectrum from a many-body perspective, which shows massive conformal particles as compounds of two correlated massless particles. The statistics of the compound (boson/fermion) depends on the helicity h of the massless components (integer/half-integer). Coherent states (CS) of particle-hole pairs (“excitons”) are also explicitly constructed as the exponential action of exciton (non-canonical) creation operators on the ground state of unpaired particles. These CS are labeled by points Z (2×2 complex matrices) on the Cartan-Bergman domain D₄=U(2,2)/U(2)², and constitute a generalized (matrix) version of Perelomov U(1, 1) coherent states labeled by points z on the unit disk D₁=U(1,1)/U(1)². First, we follow a geometric approach to the construction of CS, orthonormal basis, U(2, 2) generators and their matrix elements and symbols in the reproducing kernel Hilbert space H{sub λ}(D₄) of analytic square-integrable holomorphic functions on D₄, which carries a unitary irreducible representation of U(2, 2) with index λϵN (the conformal or scale dimension). Then we introduce a many-body representation of the previous construction through an oscillator realization of the U(2, 2) Lie algebra generators in terms of eight boson operators with constraints. This particle picture allows us for a physical interpretation of our abstract mathematical construction in the many-body jargon. In particular, the index λ is related to the number 2(λ – 2) of unpaired quanta and to the helicity h = (λ – 2)/2 of each massless particle forming the massive compound.

  6. On the oscillator realization of conformal U(2, 2) quantum particles and their particle-hole coherent states

    NASA Astrophysics Data System (ADS)

    Calixto, M.; Pérez-Romero, E.

    2014-08-01

    We revise the unireps. of U(2, 2) describing conformal particles with continuous mass spectrum from a many-body perspective, which shows massive conformal particles as compounds of two correlated massless particles. The statistics of the compound (boson/fermion) depends on the helicity h of the massless components (integer/half-integer). Coherent states (CS) of particle-hole pairs ("excitons") are also explicitly constructed as the exponential action of exciton (non-canonical) creation operators on the ground state of unpaired particles. These CS are labeled by points Z (2 × 2 complex matrices) on the Cartan-Bergman domain {D}_4=U(2,2)/U(2)^2, and constitute a generalized (matrix) version of Perelomov U(1, 1) coherent states labeled by points z on the unit disk {D}_1=U(1,1)/U(1)^2. First, we follow a geometric approach to the construction of CS, orthonormal basis, U(2, 2) generators and their matrix elements and symbols in the reproducing kernel Hilbert space H_λ ({D}_4) of analytic square-integrable holomorphic functions on {D}_4, which carries a unitary irreducible representation of U(2, 2) with index λ in {N} (the conformal or scale dimension). Then we introduce a many-body representation of the previous construction through an oscillator realization of the U(2, 2) Lie algebra generators in terms of eight boson operators with constraints. This particle picture allows us for a physical interpretation of our abstract mathematical construction in the many-body jargon. In particular, the index λ is related to the number 2(λ - 2) of unpaired quanta and to the helicity h = (λ - 2)/2 of each massless particle forming the massive compound.

  7. Characterization of U-2 wt% Mo and U-10 wt% Mo alloy powders prepared by centrifugal atomization

    NASA Astrophysics Data System (ADS)

    Kim, Ki Hwan; Lee, Don Bac; Kim, Chang Kyu; Hofman, Gerard E.; Paik, Kyung Wook

    1997-06-01

    The characteristics of high density UMo alloy powder solidified rapidly by the centrifugal atomization process have been examined. The results indicate that most of the atomized UMo alloy particles have a smooth surface and near-perfect spherical shape. The atomized powder, irrespective of particle size, is found to be single phase γ-U alloy with isotropic structure and non-dendritic grain. The continuous cooling DSC trace of U-2 wt% Mo alloy shows a small, broad endothermic peak originated from the formation of α-U phase and U 2Mo phase, whereas that of U-10 wt% Mo alloy shows no peak over all temperature ranges associated with the decomposition of γ-U phase. The γ-U phase of U-2 wt% Mo powder is decomposed as the α-U phase and the U 2Mo phase after an annealing treatment at 400°C for 100 h. But the γ-U phase of atomized U-10 wt% Mo powder remains as it was.

  8. Useful to Usable (U2U): Transforming climate information into usable tools to support Midwestern agricultural production

    NASA Astrophysics Data System (ADS)

    Prokopy, L. S.; Widhalm, M.

    2014-12-01

    There is a close connection between weather and climate patterns and successful agricultural production. Therefore, incorporating climate information into farm management is likely to reduce the risk of economic losses and increase profitability. While weather and climate information is becoming ever more abundant and accessible, the use of such information in the agricultural community remains limited. Useful to Usable (U2U): Transforming Climate Variability and Change Information for Cereal Crop Producers is a USDA-NIFA funded research and extension project focused on improving the use of climate information for agricultural production in the Midwestern United States by developing user-driven decision tools and training resources. The U2U team is a diverse and uniquely qualified group of climatologists, crop modelers, agronomists, and social scientists from 9 Midwestern universities and two NOAA Regional Climate Centers. Together, we strive to help producers make better long-term plans on what, when and where to plant and also how to manage crops for maximum yields and minimum environmental damage. To ensure relevance and usability of U2U products, our social science team is using a number of techniques including surveys and focus groups to integrate stakeholder interests, needs, and concerns into all aspects of U2U research. It is through this coupling of physical and social science disciplines that we strive to transform existing climate information into actionable knowledge.

  9. Concerted evolution of the tandem array encoding primate U2 snRNA occurs in situ, without changing the cytological context of the RNU2 locus.

    PubMed Central

    Pavelitz, T; Rusché, L; Matera, A G; Scharf, J M; Weiner, A M

    1995-01-01

    In primates, the tandemly repeated genes encoding U2 small nuclear RNA evolve concertedly, i.e. the sequence of the U2 repeat unit is essentially homogeneous within each species but differs somewhat between species. Using chromosome painting and the NGFR gene as an outside marker, we show that the U2 tandem array (RNU2) has remained at the same chromosomal locus (equivalent to human 17q21) through multiple speciation events over > 35 million years leading to the Old World monkey and hominoid lineages. The data suggest that the U2 tandem repeat, once established in the primate lineage, contained sequence elements favoring perpetuation and concerted evolution of the array in situ, despite a pericentric inversion in chimpanzee, a reciprocal translocation in gorilla and a paracentric inversion in orang utan. Comparison of the 11 kb U2 repeat unit found in baboon and other Old World monkeys with the 6 kb U2 repeat unit in humans and other hominids revealed that an ancestral U2 repeat unit was expanded by insertion of a 5 kb retrovirus bearing 1 kb long terminal repeats (LTRs). Subsequent excision of the provirus by homologous recombination between the LTRs generated a 6 kb U2 repeat unit containing a solo LTR. Remarkably, both junctions between the human U2 tandem array and flanking chromosomal DNA at 17q21 fall within the solo LTR sequence, suggesting a role for the LTR in the origin or maintenance of the primate U2 array. Images PMID:7828589

  10. High similarity of U2 snDNA sequence between A and B chromosomes in the grasshopper Abracris flavolineata.

    PubMed

    Menezes-de-Carvalho, Nahanna Zimmermann; Palacios-Gimenez, Octavio Manuel; Milani, Diogo; Cabral-de-Mello, Diogo Cavalcanti

    2015-10-01

    B chromosomes are frequently enriched for a wide variety of repetitive DNAs. Among grasshoppers in the species Abracris flavolineata (Ommatolampidinae) the B chromosomes are submetacentric, C-negative and harbor repetitive DNAs such as, U2 snDNA, C 0 t-1 DNA, two Mariner-like elements and some microsatellites. Here, we provide evidence showing the intragenome similarity between the B chromosome and the A complement in A. flavolineata, combining analysis of microdissection and chromosome painting and B chromosome-specific amplification through polymerase chain reaction (PCR) of U2 snDNA. Chromosome painting revealed signals spread through the C-negative regions, including the A and B chromosomes. Moreover, significant clustered signals forming bands were observed in some A chromosomes, and for the B chromosome, significant signals were located on both arms, which could be caused by accumulation of repetitive DNA sequences. The C-positive regions did not reveal any signals. Sequence comparison of U2 snDNA between that obtained from a genome without the B chromosome and that from µB-DNA revealed high similarity with the occurrence of four shared haplotypes, one of them (i.e., Hap1) being highly prevalent and putatively ancestral. The highest divergence from Hap1 was observed for Hap3, which was caused by only six mutational steps. These data support an intraspecific origin of the B chromosome in A. flavolineata that is highly similar with the A complement, and the low U2 snDNA sequence diversity observed in the B chromosome could be related to its recent origin, besides intrachromosomal concerted evolution for U2 snDNA repeats in the B chromosome.

  11. U(2)⁵ flavor symmetry and lepton universality violation in W→τν̄τ

    DOE PAGES

    Filipuzzi, Alberto; Portolés, Jorge; González-Alonso, Martín

    2012-06-26

    The seeming violation of universality in the τ lepton coupling to the W boson suggested by LEP-II data is studied using an effective field theory (EFT) approach. Within this framework we explore how this feature fits into the current constraints from electroweak precision observables using different assumptions about the flavor structure of New Physics, namely [U(2)×U(1)]⁵ and U(2)⁵. We show the importance of leptonic and semileptonic tau decay measurements, giving 3–4 TeV bounds on the New Physics effective scale at 90% C.L. We conclude under very general assumptions that it is not possible to accommodate this deviation from universality inmore » the EFT framework, and thus such a signal could only be explained by the introduction of light degrees of freedom or New Physics strongly coupled at the electroweak scale.« less

  12. High-Pressure Synthesis and Characterization of the Actinide Borate Phosphate U2[BO4][PO4].

    PubMed

    Hinteregger, Ernst; Wurst, Klaus; Perfler, Lukas; Kraus, Florian; Huppertz, Hubert

    2013-10-14

    A new actinide borate phosphate, U2[BO4][PO4], was synthesized in a Walker-type multianvil apparatus at 12.5 GPa and 1000 °C. The crystal structure was determined from single-crystal X-ray diffraction data collected at room temperature. U2[BO4][PO4] crystallizes in the monoclinic space group P21/c with four formula units per unit cell and the lattice parameters a = 854.6(2), b = 775.3(2), c = 816.3(2) pm, and β = 102.52(3)°. The structure consists of double layers of linked uranium-oxygen polyhedra parallel to [100]. The borate tetrahedra are located between the uranium-oxygen layers inside the double layer. The phosphate groups link the double layers. PMID:24611029

  13. GROMOS polarizable charge-on-spring models for liquid urea: COS/U and COS/U2

    SciTech Connect

    Lin, Zhixiong; Bachmann, Stephan J.; Gunsteren, Wilfred F. van

    2015-03-07

    Two one-site polarizable urea models, COS/U and COS/U2, based on the charge-on-spring model are proposed. The models are parametrized against thermodynamic properties of urea-water mixtures in combination with the polarizable COS/G2 and COS/D2 models for liquid water, respectively, and have the same functional form of the inter-atomic interaction function and are based on the same parameter calibration procedure and type of experimental data as used to develop the GROMOS biomolecular force field. Thermodynamic, dielectric, and dynamic properties of urea-water mixtures simulated using the polarizable models are closer to experimental data than using the non-polarizable models. The COS/U and COS/U2 models may be used in biomolecular simulations of protein denaturation.

  14. Human PSENEN and U2AF1L4 genes are concertedly regulated by a genuine bidirectional promoter.

    PubMed

    Didych, D A; Shamsutdinov, M F; Smirnov, N A; Akopov, S B; Monastyrskaya, G S; Uspenskaya, N Y; Nikolaev, L G; Sverdlov, E D

    2013-02-15

    Head-to-head genes with a short distance between their transcription start sites may constitute up to 10% of all genes in the genomes of various species. It was hypothesized that this intergenic space may represent bidirectional promoters which are able to initiate transcription of both genes, but the true bidirectionality was proved only for a few of them. We present experimental evidence that, according to several criteria, a 269 bp region located between the PSENEN and U2AF1L4 human genes is a genuine bidirectional promoter regulating a concerted divergent transcription of these genes. Concerted transcription of PSENEN and U2AF1L4 can be necessary for regulation of T-cell activity. PMID:23246698

  15. U(2)⁵ flavor symmetry and lepton universality violation in W→τν̄τ

    SciTech Connect

    Filipuzzi, Alberto; Portolés, Jorge; González-Alonso, Martín

    2012-06-26

    The seeming violation of universality in the τ lepton coupling to the W boson suggested by LEP-II data is studied using an effective field theory (EFT) approach. Within this framework we explore how this feature fits into the current constraints from electroweak precision observables using different assumptions about the flavor structure of New Physics, namely [U(2)×U(1)]⁵ and U(2)⁵. We show the importance of leptonic and semileptonic tau decay measurements, giving 3–4 TeV bounds on the New Physics effective scale at 90% C.L. We conclude under very general assumptions that it is not possible to accommodate this deviation from universality in the EFT framework, and thus such a signal could only be explained by the introduction of light degrees of freedom or New Physics strongly coupled at the electroweak scale.

  16. The NASA Thunderstorm Overflight Program (TOP): Research in atmospheric electricity from an instrumented U-2 aircraft platform

    NASA Technical Reports Server (NTRS)

    Vaughan, O. H., Jr.

    1983-01-01

    An overview of the NASA Thunderstorm Overflight Program (TOP) is presented. The various instruments flown on the NASA U-2 aircraft, as well as the ground instrumentation used to collect optical and electronic signature from the lightning events, are discussed. Samples of some of the photographic and electronic signatures are presented. Approximately 6400 electronic data samples of optical pulses were collected and are being analyzed.

  17. Knockdown of PARP-1 Inhibits Proliferation and ERK Signals, Increasing Drug Sensitivity in Osteosarcoma U2OS Cells.

    PubMed

    Li, Sheng; Cui, Zhengli; Meng, Xianfeng

    2016-01-01

    Poly(ADP-ribose) polymerase 1 (PARP-1) is reported to be involved in DNA repair and is now recognized as a key regulator in carcinogenesis. However, the potential role and the molecular mechanism underlying the effect of PARP-1 on osteosarcoma (OS) cells have not been elucidated. In this study, the results showed that knockdown of PARP-1 resulted in decreased cell proliferation, increased cell apoptosis, and G0/G1 phase arrest in U2OS cells. In addition, increased expression of active caspase 3 and Bax, but reduced Bcl-2, cyclin D1, and phosphorylated extracellular signal regulated kinase 1/2 (pERK1/2) were observed in PARP-1 knockdown in U2OS cells. Moreover, knockdown of PARP-1 correlated with elevated chemosensitivity of U2OS cells to cisplatin through inactivation of the ERK1/2 signaling pathway. In conclusion, our findings demonstrated that PARP-1 plays an important role in regulating OS growth, combining PARP-1 gene therapy with traditional chemotherapy, and may serve as a promising approach to OS therapy. PMID:27656839

  18. Ancient Origin of the U2 Small Nuclear RNA Gene-Targeting Non-LTR Retrotransposons Utopia

    PubMed Central

    Kojima, Kenji K.

    2015-01-01

    Most non-long terminal repeat (non-LTR) retrotransposons encoding a restriction-like endonuclease show target-specific integration into repetitive sequences such as ribosomal RNA genes and microsatellites. However, only a few target-specific lineages of non-LTR retrotransposons are distributed widely and no lineage is found across the eukaryotic kingdoms. Here we report the most widely distributed lineage of target sequence-specific non-LTR retrotransposons, designated Utopia. Utopia is found in three supergroups of eukaryotes: Amoebozoa, SAR, and Opisthokonta. Utopia is inserted into a specific site of U2 small nuclear RNA genes with different strength of specificity for each family. Utopia families from oomycetes and wasps show strong target specificity while only a small number of Utopia copies from reptiles are flanked with U2 snRNA genes. Oomycete Utopia families contain an “archaeal” RNase H domain upstream of reverse transcriptase (RT), which likely originated from a plant RNase H gene. Analysis of Utopia from oomycetes indicates that multiple lineages of Utopia have been maintained inside of U2 genes with few copy numbers. Phylogenetic analysis of RT suggests the monophyly of Utopia, and it likely dates back to the early evolution of eukaryotes. PMID:26556480

  19. Ancient Origin of the U2 Small Nuclear RNA Gene-Targeting Non-LTR Retrotransposons Utopia.

    PubMed

    Kojima, Kenji K; Jurka, Jerzy

    2015-01-01

    Most non-long terminal repeat (non-LTR) retrotransposons encoding a restriction-like endonuclease show target-specific integration into repetitive sequences such as ribosomal RNA genes and microsatellites. However, only a few target-specific lineages of non-LTR retrotransposons are distributed widely and no lineage is found across the eukaryotic kingdoms. Here we report the most widely distributed lineage of target sequence-specific non-LTR retrotransposons, designated Utopia. Utopia is found in three supergroups of eukaryotes: Amoebozoa, SAR, and Opisthokonta. Utopia is inserted into a specific site of U2 small nuclear RNA genes with different strength of specificity for each family. Utopia families from oomycetes and wasps show strong target specificity while only a small number of Utopia copies from reptiles are flanked with U2 snRNA genes. Oomycete Utopia families contain an "archaeal" RNase H domain upstream of reverse transcriptase (RT), which likely originated from a plant RNase H gene. Analysis of Utopia from oomycetes indicates that multiple lineages of Utopia have been maintained inside of U2 genes with few copy numbers. Phylogenetic analysis of RT suggests the monophyly of Utopia, and it likely dates back to the early evolution of eukaryotes. PMID:26556480

  20. Ancient Origin of the U2 Small Nuclear RNA Gene-Targeting Non-LTR Retrotransposons Utopia.

    PubMed

    Kojima, Kenji K; Jurka, Jerzy

    2015-01-01

    Most non-long terminal repeat (non-LTR) retrotransposons encoding a restriction-like endonuclease show target-specific integration into repetitive sequences such as ribosomal RNA genes and microsatellites. However, only a few target-specific lineages of non-LTR retrotransposons are distributed widely and no lineage is found across the eukaryotic kingdoms. Here we report the most widely distributed lineage of target sequence-specific non-LTR retrotransposons, designated Utopia. Utopia is found in three supergroups of eukaryotes: Amoebozoa, SAR, and Opisthokonta. Utopia is inserted into a specific site of U2 small nuclear RNA genes with different strength of specificity for each family. Utopia families from oomycetes and wasps show strong target specificity while only a small number of Utopia copies from reptiles are flanked with U2 snRNA genes. Oomycete Utopia families contain an "archaeal" RNase H domain upstream of reverse transcriptase (RT), which likely originated from a plant RNase H gene. Analysis of Utopia from oomycetes indicates that multiple lineages of Utopia have been maintained inside of U2 genes with few copy numbers. Phylogenetic analysis of RT suggests the monophyly of Utopia, and it likely dates back to the early evolution of eukaryotes.

  1. Salinomycin simultaneously induces apoptosis and autophagy through generation of reactive oxygen species in osteosarcoma U2OS cells.

    PubMed

    Kim, Sang-Hun; Choi, Young-Jun; Kim, Kwang-Youn; Yu, Sun-Nyoung; Seo, Young-Kyo; Chun, Sung-Sik; Noh, Kyung-Tae; Suh, Jeung-Tak; Ahn, Soon-Cheol

    2016-04-29

    Salinomycin, a polyether antibiotic, acts as a highly selective potassium ionophore. It was reported to anticancer activity on various cancer cell lines. In this study, salinomycin was examined on apoptosis and autophagy through generation of reactive oxygen species (ROS) in osteosarcoma U2OS cells. Apoptosis, autophagy, mitochondrial membrane potential (MMP) and ROS were analyzed using flow cytometry. Also, expressions of apoptosis- and autophagy-related proteins were determined by western blotting. As a result, salinomycin triggered apoptosis of U2OS cells, which was accompanied by change of MMP and cleavage of caspases-3 and poly (ADP-ribose) polymerase. And salinomycin increased the expression of autophagy-related protein and accumulation of acidic vesicular organelles (AVO). Salinomycin-induced ROS production promotes both apoptosis and autophagy, as evidenced by the result that treatment of N-acetyl-l-cysteine (NAC), a ROS scavenger, attenuated both apoptosis and autophagy. In addition, inhibition of autophagy by 3-methyladenine (3 MA) enhanced the salinoymcin-induced apoptosis. Taken together, these results suggested that salinomycin-induced autophagy, as a survival mechanism, might be a potential strategy through ROS regulation in cancer therapy. PMID:27033598

  2. Theoretical investigations of structural, electronic and elastic properties of U2S3 type Nb2N3 under high pressure

    NASA Astrophysics Data System (ADS)

    Zhang, Ji-Dong; Huang, Xiao-Yu

    2015-01-01

    The structural, electronic and elastic properties of U2S3 type Nb2N3 under pressure ranging from 0 GPa to 100 GPa are investigated by plane-wave pseudopotential density functional theory with the generalized gradient approximation in this paper. The results at zero pressure are in good agreement with available theoretical results. The properties of U2S3 type Nb2N3 are compared to the new synthetic superhard material η-Ta2N3. The results show that U2S3 type Nb2N3 is hard to compress, and expected to have comparable hardness to η-Ta2N3.

  3. Ultrafast carrier relaxation through Auger recombination in the topological insulator B i1.5S b0.5T e1.7S e1.3

    NASA Astrophysics Data System (ADS)

    Onishi, Yoshito; Ren, Zhi; Segawa, Kouji; Kaszub, Wawrzyniec; Lorenc, Maciej; Ando, Yoichi; Tanaka, Koichiro

    2015-02-01

    Ultrafast carrier dynamics have great significance for our understanding of the transport properties of the surface state in topological insulator (TI) materials. We report midinfrared pump-probe measurements on the intrinsic TI material B i1.5S b0.5T e1.7S e1.3 and show that the change in photoinduced reflectivity can be decomposed into a fast negative part and a slow positive part. Calculations of the dielectric function made at various carrier temperatures and densities reveal that the fast negative component corresponds to the disappearance of the phase-space filling effect due to hot carriers around the probe energy and the decay component corresponds to the recombination of carriers near the band edge. The ratio of the fast negative component to the slow positive component is larger in the excitations conducted at the higher carrier densities, which suggests that the carrier temperature increases through Auger recombination. A qualitative analysis using rate equations reinforces this assumption, so we conclude that Auger recombination is the main cause of the population relaxation at carrier densities higher than 1018c m-3 and that we determined the Auger coefficient for B i1.5S b0.5T e1.7S e1.3 as C =0.4 ×10-26c m6/s .

  4. THRA1 and D17S183 flank an interval of <4 cM for the breast-ovarian cancer gene (BRCA1) on chromosome 17q21

    SciTech Connect

    Bowcock, A.M.; Osborne-Lawrence, S. ); Anderson, L.A.; Friedman, L.S.; Rowell, S.E.; Hall, J.M.; King, M.C. ); Black, D.M.; Solomon, E. )

    1993-04-01

    In order to pinpoint the locale of the gene for early-onset familial breast and ovarian cancer (BRCA1), polymorphisms were developed within the locus for thyroid hormone receptor alpha (THRA1) and for several anonymous sequences at chromosome 17q12-q21. The THRA1 polymorphism is a dinucleotide repeat with 10 alleles and heterozygosity .79. Gene mapping in extended families with inherited, early-onset breast and ovarian cancer indicates that BRCA1 is distal to THRA1 and proximal to D17S183 (SCG43), an interval of <4 cM. This locale excludes HER2, THRA1, WNT3, HOX2, NGFR, PHB, COLIA1, NME1, and NME2 as candidates for BRCA1 but does not exclude RARA or EDH17B. Resolving the remaining recombination events in these families by new polymorphisms in the THRA1-D17S183 interval will facilitate positional cloning of the breast-ovarian cancer gene on chromosome 17q12-q21. 16 refs., 3 figs., 1 tab.

  5. The new NCPSS BL19U2 beamline at the SSRF for small-angle X-ray scattering from biological macromolecules in solution1

    PubMed Central

    Li, Na; Li, Xiuhong; Wang, Yuzhu; Liu, Guangfeng; Zhou, Ping; Wu, Hongjin; Hong, Chunxia; Bian, Fenggang; Zhang, Rongguang

    2016-01-01

    The beamline BL19U2 is located in the Shanghai Synchrotron Radiation Facility (SSRF) and is its first beamline dedicated to biological material small-angle X-ray scattering (BioSAXS). The electrons come from an undulator which can provide high brilliance for the BL19U2 end stations. A double flat silicon crystal (111) monochromator is used in BL19U2, with a tunable monochromatic photon energy ranging from 7 to 15 keV. To meet the rapidly growing demands of crystallographers, biochemists and structural biologists, the BioSAXS beamline allows manual and automatic sample loading/unloading. A Pilatus 1M detector (Dectris) is employed for data collection, characterized by a high dynamic range and a short readout time. The highly automated data processing pipeline SASFLOW was integrated into BL19U2, with help from the BioSAXS group of the European Molecular Biology Laboratory (EMBL, Hamburg), which provides a user-friendly interface for data processing. The BL19U2 beamline was officially opened to users in March 2015. To date, feedback from users has been positive and the number of experimental proposals at BL19U2 is increasing. A description of the new BioSAXS beamline and the setup characteristics is given, together with examples of data obtained. PMID:27738413

  6. Deep recombination centers in C u2ZnSnS e4 revealed by screened-exchange hybrid density functional theory

    NASA Astrophysics Data System (ADS)

    Yee, Ye Sheng; Magyari-Köpe, Blanka; Nishi, Yoshio; Bent, Stacey F.; Clemens, Bruce M.

    2015-11-01

    We present a comprehensive study of the thermodynamic and electronic properties of intrinsic point defects in the solar energy conversion materials C u2ZnSnS e4 and CuInS e2 based on the screened-exchange hybrid density functional theory. A comparison between the defect transition levels for C u2ZnSnS e4 and CuInS e2 reveals that in C u2ZnSnS e4 , the S nCu and S nZn antisite defects can be recombination centers with defect states close to midgap, while the I nCu antisite defect has a shallow defect level in CuInS e2 . The resultant higher Shockley-Read-Hall recombination rate in C u2ZnSnS e4 reduces the steady-state concentration of minority carriers and quasi-Fermi level separation under illumination. This may explain the origin of the low open-circuit voltage values for C u2ZnSnS e4 solar cells compared to CuInS e2 solar cells.

  7. Alteration of the SETBP1 Gene and Splicing Pathway Genes SF3B1, U2AF1, and SRSF2 in Childhood Acute Myeloid Leukemia

    PubMed Central

    Choi, Hyun-Woo; Kim, Hye-Ran; Baek, Hee-Jo; Kook, Hoon; Cho, Duck; Shin, Jong-Hee; Suh, Soon-Pal; Ryang, Dong-Wook

    2015-01-01

    Background Recurrent somatic SET-binding protein 1 (SETBP1) and splicing pathway gene mutations have recently been found in atypical chronic myeloid leukemia and other hematologic malignancies. These mutations have been comprehensively analyzed in adult AML, but not in childhood AML. We investigated possible alteration of the SETBP1, splicing factor 3B subunit 1 (SF3B1), U2 small nuclear RNA auxiliary factor 1 (U2AF1), and serine/arginine-rich splicing factor 2 (SRSF2) genes in childhood AML. Methods Cytogenetic and molecular analyses were performed to reveal chromosomal and genetic alterations. Sequence alterations in the SETBP1, SF3B1, U2AF1, and SRSF2 genes were examined by using direct sequencing in a cohort of 53 childhood AML patients. Results Childhood AML patients did not harbor any recurrent SETBP1 gene mutations, although our study did identify a synonymous mutation in one patient. None of the previously reported aberrations in the mutational hotspot of SF3B1, U2AF1, and SRSF2 were identified in any of the 53 patients. Conclusions Alterations of the SETBP1 gene or SF3B1, U2AF1, and SRSF2 genes are not common genetic events in childhood AML, implying that the mutations are unlikely to exert a driver effect in myeloid leukemogenesis during childhood. PMID:25553291

  8. A genome landscape of SRSF3-regulated splicing events and gene expression in human osteosarcoma U2OS cells

    PubMed Central

    Ajiro, Masahiko; Jia, Rong; Yang, Yanqin; Zhu, Jun; Zheng, Zhi-Ming

    2016-01-01

    Alternative RNA splicing is an essential process to yield proteomic diversity in eukaryotic cells, and aberrant splicing is often associated with numerous human diseases and cancers. We recently described serine/arginine-rich splicing factor 3 (SRSF3 or SRp20) being a proto-oncogene. However, the SRSF3-regulated splicing events responsible for its oncogenic activities remain largely unknown. By global profiling of the SRSF3-regulated splicing events in human osteosarcoma U2OS cells, we found that SRSF3 regulates the expression of 60 genes including ERRFI1, ANXA1 and TGFB2, and 182 splicing events in 164 genes, including EP300, PUS3, CLINT1, PKP4, KIF23, CHK1, SMC2, CKLF, MAP4, MBNL1, MELK, DDX5, PABPC1, MAP4K4, Sp1 and SRSF1, which are primarily associated with cell proliferation or cell cycle. Two SRSF3-binding motifs, CCAGC(G)C and A(G)CAGCA, are enriched to the alternative exons. An SRSF3-binding site in the EP300 exon 14 is essential for exon 14 inclusion. We found that the expression of SRSF1 and SRSF3 are mutually dependent and coexpressed in normal and tumor tissues/cells. SRSF3 also significantly regulates the expression of at least 20 miRNAs, including a subset of oncogenic or tumor suppressive miRNAs. These data indicate that SRSF3 affects a global change of gene expression to maintain cell homeostasis. PMID:26704980

  9. A genome landscape of SRSF3-regulated splicing events and gene expression in human osteosarcoma U2OS cells.

    PubMed

    Ajiro, Masahiko; Jia, Rong; Yang, Yanqin; Zhu, Jun; Zheng, Zhi-Ming

    2016-02-29

    Alternative RNA splicing is an essential process to yield proteomic diversity in eukaryotic cells, and aberrant splicing is often associated with numerous human diseases and cancers. We recently described serine/arginine-rich splicing factor 3 (SRSF3 or SRp20) being a proto-oncogene. However, the SRSF3-regulated splicing events responsible for its oncogenic activities remain largely unknown. By global profiling of the SRSF3-regulated splicing events in human osteosarcoma U2OS cells, we found that SRSF3 regulates the expression of 60 genes including ERRFI1, ANXA1 and TGFB2, and 182 splicing events in 164 genes, including EP300, PUS3, CLINT1, PKP4, KIF23, CHK1, SMC2, CKLF, MAP4, MBNL1, MELK, DDX5, PABPC1, MAP4K4, Sp1 and SRSF1, which are primarily associated with cell proliferation or cell cycle. Two SRSF3-binding motifs, CCAGC(G)C and A(G)CAGCA, are enriched to the alternative exons. An SRSF3-binding site in the EP300 exon 14 is essential for exon 14 inclusion. We found that the expression of SRSF1 and SRSF3 are mutually dependent and coexpressed in normal and tumor tissues/cells. SRSF3 also significantly regulates the expression of at least 20 miRNAs, including a subset of oncogenic or tumor suppressive miRNAs. These data indicate that SRSF3 affects a global change of gene expression to maintain cell homeostasis.

  10. Thermodynamic studies of studtite thermal decomposition pathways via amorphous intermediates UO3, U2O7, and UO4

    DOE PAGES

    Guo, Xiaofeng; Wu, Di; Xu, Hongwu; Burns, Peter C.; Navrotsky, Alexandra

    2016-06-08

    The thermal decomposition of studtite (UO2)O2(H2O)2·2H2O results in a series of intermediate X-ray amorphous materials with general composition UO3+x (x = 0, 0.5, 1). As an extension of a structural study on U2O7, this work provides detailed calorimetric data on these amorphous oxygen-rich materials since their energetics and thermal stability are unknown. These were characterized in situ by thermogravimetry, and mass spectrometry. Ex situ X-ray diffraction and infrared spectroscopy characterized their chemical bonding and local structures. This detailed characterization formed the basis for obtaining formation enthalpies by high temperature oxide melt solution calorimetry. The thermodynamic data demonstrate the metastability ofmore » the amorphous UO3+x materials, and explain their irreversible and spontaneous reactions to generate oxygen and form metaschoepite. Thus, formation of studtite in the nuclear fuel cycle, followed by heat treatment, can produce metastable amorphous UO3+x materials that pose the risk of significant O2 gas. Quantitative knowledge of the energy landscape of amorphous UO3+x was provided for stability analysis and assessment of conditions for decomposition.« less

  11. Thermodynamic studies of studtite thermal decomposition pathways via amorphous intermediates UO3, U2O7, and UO4

    NASA Astrophysics Data System (ADS)

    Guo, Xiaofeng; Wu, Di; Xu, Hongwu; Burns, Peter C.; Navrotsky, Alexandra

    2016-09-01

    The thermal decomposition of studtite (UO2)O2(H2O)2·2H2O results in a series of intermediate X-ray amorphous materials with general composition UO3+x (x = 0, 0.5, 1). As an extension of a structural study on U2O7, this work provides detailed calorimetric data on these amorphous oxygen-rich materials since their energetics and thermal stability are unknown. These were characterized in situ by thermogravimetry, and mass spectrometry. Ex situ X-ray diffraction and infrared spectroscopy characterized their chemical bonding and local structures. This detailed characterization formed the basis for obtaining formation enthalpies by high temperature oxide melt solution calorimetry. The thermodynamic data demonstrate the metastability of the amorphous UO3+x materials, and explain their irreversible and spontaneous reactions to generate oxygen and form metaschoepite. Thus, formation of studtite in the nuclear fuel cycle, followed by heat treatment, can produce metastable amorphous UO3+x materials that pose the risk of significant O2 gas. Quantitative knowledge of the energy landscape of amorphous UO3+x was provided for stability analysis and assessment of conditions for decomposition.

  12. Caenorhabditis elegans pseudouridine synthase 1 activity in vivo: tRNA is a substrate, but not U2 small nuclear RNA.

    PubMed

    Patton, Jeffrey R; Padgett, Richard W

    2003-06-01

    The formation of pseudouridine (Psi) from uridine is post-transcriptional and catalysed by pseudouridine synthases, several of which have been characterized from eukaryotes. Pseudouridine synthase 1 (Pus1p) has been well characterized from yeast and mice. In yeast, Pus1p has been shown to have dual substrate specificity, modifying uridines in tRNAs and at position 44 in U2 small nuclear RNA (U2 snRNA). In order to study the in vivo activity of a metazoan Pus1p, a knockout of the gene coding for the homologue of Pus1p in Caenorhabditis elegans was obtained. The deletion encompasses the first two putative exons and includes the essential aspartate that is required for activity in truA pseudouridine synthases. The locations of most modified nucleotides on small RNAs in C. elegans are not known, and the positions of Psi were determined on four tRNAs and U2 snRNA. The uridine at position 27 of tRNA(Val) (AAC), a putative Pus1p-modification site, was converted into Psi in the wild-type worms, but the tRNA(Val) (AAC) from mutant worms lacked the modification. Psi formation at positions 13, 32, 38 and 39, all of which should be modified by other pseudouridine synthases, was not affected by the loss of Pus1p. The absence of Pus1p in C. elegans had no effect on the modification of U2 snRNA in vivo, even though worm U2 snRNA has a Psi at position 45 (the equivalent of yeast U2 snRNA position 44) and at four other positions. This result was unexpected, given the known dual specificity of yeast Pus1p.

  13. DNA Methylation Is Linked to Deacetylation of Histone H3, but Not H4, on the Imprinted Genes Snrpn and U2af1-rs1

    PubMed Central

    Gregory, Richard I.; Randall, Tamzin E.; Johnson, Colin A.; Khosla, Sanjeev; Hatada, Izuho; O'Neill, Laura P.; Turner, Bryan M.; Feil, Robert

    2001-01-01

    The relationship between DNA methylation and histone acetylation at the imprinted mouse genes U2af1-rs1 and Snrpn is explored by chromatin immunoprecipitation (ChIP) and resolution of parental alleles using single-strand conformational polymorphisms. The U2af1-rs1 gene lies within a differentially methylated region (DMR), while Snrpn has a 5′ DMR (DMR1) with sequences homologous to the imprinting control center of the Prader-Willi/Angelman region. For both DMR1 of Snrpn and the 5′ untranslated region (5′-UTR) and 3′-UTR of U2af1-rs1, the methylated and nonexpressed maternal allele was underacetylated, relative to the paternal allele, at all H3 lysines tested (K14, K9, and K18). For H4, underacetylation of the maternal allele was exclusively (U2af1-rs1) or predominantly (Snrpn) at lysine 5. Essentially the same patterns of differential acetylation were found in embryonic stem (ES) cells, embryo fibroblasts, and adult liver from F1 mice and in ES cells from mice that were dipaternal or dimaternal for U2af1-rs1. In contrast, in a region within Snrpn that has biallelic methylation in the cells and tissues analyzed, the paternal (expressed) allele showed relatively increased acetylation of H4 but not of H3. The methyl-CpG-binding-domain (MBD) protein MeCP2 was found, by ChIP, to be associated exclusively with the maternal U2af1-rs1 allele. To ask whether DNA methylation is associated with histone deacetylation, we produced mice with transgene-induced methylation at the paternal allele of U2af1-rs1. In these mice, H3 was underacetylated across both the parental U2af1-rs1 alleles whereas H4 acetylation was unaltered. Collectively, these data are consistent with the hypothesis that CpG methylation leads to deacetylation of histone H3, but not H4, through a process that involves selective binding of MBD proteins. PMID:11463825

  14. Final Characterization Report for Corrective Action Unit 109: Area 2 U-2BU Crater, Nevada Test Site, Nevada, Revision 1

    SciTech Connect

    ITLV

    1998-12-01

    Corrective Action Unit 109, Area 2 U-2bu Crater, is an inactive Resource Conservation and Recovery Act Part A Permit disposal unit located in Area 2 at the Nevada Test Site, Nevada. The Corrective Action Unit has been characterized under the requirements of the Nevada Test Site Resource Conservation and Recovery Act, Part A Permit (NDEP, 1995) and Title 40 Code of Federal Regulations Part 265 (CFR, 1996). The site characterization was performed under the RCRA Part A Permit Characterization Plan for the U-2bu Subsidence Crater (DOE/NV, 1998c), as approved by the Nevada Division of Environmental Protection (Liebendorfer, 1998). The primary objective of the site characterization activities was to evaluate the presence, concentration, and extent of any Resource Conservation and Recovery Act contaminants in the crater. Surface soil samples were collected on April 22, 1998, and subsurface soil samples and geotechnical samples were collected from April 27-29, 1998. Soil samples were collected using a hand auger or a piston-type drive hammer to advance a 5-centimeter (2-inch) diameter steel sampling tool into the ground. The permit for the Nevada Test Site requires that Corrective Action Unit 109 be closed under 40 Code of Federal Regulations 265 Subpart G and 40 Code of Federal Regulations Part 265.310 (CFR, 1996). Analysis of the data collected during the characterization effort indicates that lead was detected in Study Area 1 at 5.7 milligrams per liter, above the regulatory level in 40 Code of Federal Regulations 261.24 of 5.0 milligrams per liter. Except for the lead detection at a single location within the crater, the original Resource Conservation Recovery Act constituents of potential concern determined between the U.S. Department of Energy, Nevada Operations Office and the Nevada Department of Environmental Protection during the Data Quality Objectives process (DOE/NV, 1998b) were not found to be present at Corrective Action Unit 109 above regulatory levels of

  15. Sm and U2B" proteins redistribute to different nuclear domains in dormant and proliferating onion cells.

    PubMed

    Cui, Ping; Moreno Díaz de la Espina, Susana

    2003-05-01

    Monoclonal antibodies against the spliceosomal proteins Sm and U2B", and against p105, a protein component of interchromatin granules, were used to investigate the nuclear distribution of the splicing factors in Allium cepa L. meristematic cells. Confocal microscopy showed that in steady-state proliferating cells, the spliceosomal components were distributed into two nuclear domains: (i) a diffuse nucleoplasmic network similar to that formed by interchromatin granules and (ii) numerous Cajal bodies. These domains were the counterpart of the perichromatin fibrils and granules, interchromatin granules and Cajal bodies observed by electron microscopy after EDTA and bismuth oxynitrate stainings. Dormant cells showed a nuclear distribution of the proteins in small Cajal bodies and numerous micro-speckles, correlated with the distribution of ribonucleoproteins (RNPs) observed by electron microscopy. The spliceosomal proteins relocated to the diffuse nucleoplasmic network and Cajal bodies when the cells were released from dormancy by water soaking and they re-started their proliferative activity. Inhibition of RNA synthesis by 5,6-dichloro-1-beta- d-ribofuranosylbenzimidazole (DRB) treatment in proliferating cells demonstrated that the micro-speckles were not the morphological expression of a transcription block. Fractionation and confocal microscopy studies showed a differential association of the splicing factors with the nuclear matrix depending not only on the protein, but also on nuclear activity. Our results suggest a reversible relocation of the spliceosomal proteins between different sub-nuclear domains in physiological conditions. We report here an unusual nuclear domain in dormant nuclei, the micro-speckles, corresponding to storage sites for RNPs, which were rapidly mobilised after water imbibition. PMID:12721845

  16. Carbon dioxide, hydrographic, and chemical data obtained in the Central South Pacific Ocean (WOCE sections P17S and P16S) during the tunes-2-expedition of the R/V Thomas Washington, July--August 1991

    SciTech Connect

    1991-12-31

    This data documentation discusses the procedures and methods used to measure total carbon dioxide (TCO{sub 2}), discrete partial pressure of TCO{sub 2} (pCO{sub 2}), and total alkalinity (TALK), during the Research Vessel (R/V) Thomas Washington TUNES Leg 2 Expedition in the central South Pacific Ocean. Conducted as part of the World Ocean Circulation Experiment (WOCE), the cruise began in Papeete, Tahiti, French Polynesia, on July 16, 1991, and returned to Papeete on August 25, 1991. WOCE Meridional Sections P17S along 135{degrees} W and P16S along 150{degrees} W were completed during the 40-day expedition. A total of 97 hydrographic stations were occupied. Hydrographic and chemical measurements made along WOCE Sections P17S and P16S included pressure, temperature, salinity, and oxygen measured by conductivity, temperature and depth sensor; bottle salinity; oxygen; phosphate; nitrate; nitrite; silicate; CFC-12; CFC- 11; TCO{sub 2}; TALK; and pCO{sub 2} measured at 20{degrees}C. The TCO{sub 2} concentration in 1000 seawater samples was determined with a coulometric analysis system, the pCO{sub 2} in 940 water samples was determined with an equilibrator/gas chromatograph system, while the TALK concentration in 139 samples was determined on shore at the laboratory of C. Goyet of Woods Hole Oceanographic Institution with an alkalinity titration system. In addition, 156 coulometric measurements for the Certified Reference Material (Batch {number_sign}6) were made and yielded a mean value of 2303.2 {plus_minus} 1.5 {mu}mol/kg. This mean value agrees within a standard deviation of the 2304.6 {plus_minus} 1.6 {mu}mol/kg (N=9) value determined with the manometer of C. D. Keeling at Scripps Institution of Oceanography (SIO). Replicate samples from 11 Niskin bottles at 4 stations were also collected for later shore-based reference analyses of TCO{sub 2} and TALK by vacuum extraction and manometry in the laboratory of C. D. Keeling of SIO.

  17. Draft Genome Sequence of Streptomyces sp. AVP053U2 Isolated from Styela clava, a Tunicate Collected in Long Island Sound

    PubMed Central

    deMayo, James A.; Maas, Kendra R.

    2016-01-01

    Streptomyces sp. AVP053U2 is a marine bacterium isolated from Styela clava, a tunicate collected in Long Island Sound. Here, we report a draft genome for this bacterium, which was found to contain a high capacity for secondary metabolite production based on analysis and identification of numerous biosynthetic gene clusters. PMID:27738023

  18. Inactive allele-specific methylation and chromatin structure of the imprinted gene U2af1-rs1 on mouse chromosome 11

    SciTech Connect

    Shibata, Hideo; Yoshino, Kiyoshi; Kamiya, Mamoru

    1996-07-01

    The imprinted U2Af1-rs1 gene that maps to mouse chromosome 11 is predominately expressed from the paternal allele. We examined the methylation of genomic sequences in and around the U2af1-rs1 locus to establish the extent of sequence modifications that accompanied the silencing of the maternal allele. The analysis of HapII or HhaI sites showed that the silent maternal allele was hypermethylated in a block of CpG sequences that covered more than 10 kb. By comparison, the expressed paternal allele was unmethylated from a CpG island upstream of the transcribed region through 2 kb. An analysis of DNaseI hypersensitivity of a putative promoter of U2af1-rs1 showed an open chromatin conformation only on the unmethylated, expressed paternal allele. These results suggest that allele-specific hypermethylation covering the gene and its upstream CpG island plays a role in maternal allele repression of U2af1-rs1, which is reflected in altered chromatin conformation of DNaseI hypersensitive sites. 9 refs., 2 figs.

  19. Molecular cloning and in silico characterization of knottin peptide, U2-SCRTX-Lit2, from brown spider (Loxosceles intermedia) venom glands.

    PubMed

    Meissner, Gabriel Otto; de Resende Lara, Pedro Túlio; Scott, Luis Paulo Barbour; Braz, Antônio Sérgio Kimus; Chaves-Moreira, Daniele; Matsubara, Fernando Hitomi; Soares, Eduardo Mendonça; Trevisan-Silva, Dilza; Gremski, Luiza Helena; Veiga, Silvio Sanches; Chaim, Olga Meiri

    2016-09-01

    Inhibitor cystine knots (ICKs) are a family of structural peptides with a large number of cysteine residues that form intramolecular disulfide bonds, resulting in a knot. These peptides are involved in a variety of biological functions including predation and defense, and are found in various species, such as spiders, scorpions, sea anemones, and plants. The Loxosceles intermedia venom gland transcriptome identified five groups of ICK peptides that represent more than 50 % of toxin-coding transcripts. Here, we describe the molecular cloning of U2-Sicaritoxin-Lit2 (U2-SCRTX-Lit2), bioinformatic characterization, structure prediction, and molecular dynamic analysis. The sequence of U2-SCRTX-Lit2 obtained from the transcriptome is similar to that of μ-Hexatoxin-Mg2, a peptide that inhibits the insect Nav channel. Bioinformatic analysis of sequences classified as ICK family members also showed a conservation of cysteine residues among ICKs from different spiders, with the three dimensional molecular model of U2-SCRTX-Lit2 similar in structure to the hexatoxin from μ-hexatoxin-Mg2a. Molecular docking experiments showed the interaction of U2-SCRTX-Lit2 to its predictable target-the Spodoptera litura voltage-gated sodium channel (SlNaVSC). After 200 ns of molecular dynamic simulation, the final structure of the complex showed stability in agreement with the experimental data. The above analysis corroborates the existence of a peptide toxin with insecticidal activity from a novel ICK family in L. intermedia venom and demonstrates that this peptide targets Nav channels.

  20. Molecular cloning and in silico characterization of knottin peptide, U2-SCRTX-Lit2, from brown spider (Loxosceles intermedia) venom glands.

    PubMed

    Meissner, Gabriel Otto; de Resende Lara, Pedro Túlio; Scott, Luis Paulo Barbour; Braz, Antônio Sérgio Kimus; Chaves-Moreira, Daniele; Matsubara, Fernando Hitomi; Soares, Eduardo Mendonça; Trevisan-Silva, Dilza; Gremski, Luiza Helena; Veiga, Silvio Sanches; Chaim, Olga Meiri

    2016-09-01

    Inhibitor cystine knots (ICKs) are a family of structural peptides with a large number of cysteine residues that form intramolecular disulfide bonds, resulting in a knot. These peptides are involved in a variety of biological functions including predation and defense, and are found in various species, such as spiders, scorpions, sea anemones, and plants. The Loxosceles intermedia venom gland transcriptome identified five groups of ICK peptides that represent more than 50 % of toxin-coding transcripts. Here, we describe the molecular cloning of U2-Sicaritoxin-Lit2 (U2-SCRTX-Lit2), bioinformatic characterization, structure prediction, and molecular dynamic analysis. The sequence of U2-SCRTX-Lit2 obtained from the transcriptome is similar to that of μ-Hexatoxin-Mg2, a peptide that inhibits the insect Nav channel. Bioinformatic analysis of sequences classified as ICK family members also showed a conservation of cysteine residues among ICKs from different spiders, with the three dimensional molecular model of U2-SCRTX-Lit2 similar in structure to the hexatoxin from μ-hexatoxin-Mg2a. Molecular docking experiments showed the interaction of U2-SCRTX-Lit2 to its predictable target-the Spodoptera litura voltage-gated sodium channel (SlNaVSC). After 200 ns of molecular dynamic simulation, the final structure of the complex showed stability in agreement with the experimental data. The above analysis corroborates the existence of a peptide toxin with insecticidal activity from a novel ICK family in L. intermedia venom and demonstrates that this peptide targets Nav channels. PMID:27488102

  1. The target of the DEAH-box NTP triphosphatase Prp43 in Saccharomyces cerevisiae spliceosomes is the U2 snRNP-intron interaction

    PubMed Central

    Fourmann, Jean-Baptiste; Dybkov, Olexandr; Agafonov, Dmitry E; Tauchert, Marcel J; Urlaub, Henning; Ficner, Ralf; Fabrizio, Patrizia; Lührmann, Reinhard

    2016-01-01

    The DEAH-box NTPase Prp43 and its cofactors Ntr1 and Ntr2 form the NTR complex and are required for disassembling intron-lariat spliceosomes (ILS) and defective earlier spliceosomes. However, the Prp43 binding site in the spliceosome and its target(s) are unknown. We show that Prp43 fused to Ntr1's G-patch motif (Prp43_Ntr1GP) is as efficient as the NTR in ILS disassembly, yielding identical dissociation products and recognizing its natural ILS target even in the absence of Ntr1’s C-terminal-domain (CTD) and Ntr2. Unlike the NTR, Prp43_Ntr1GP disassembles earlier spliceosomal complexes (A, B, Bact), indicating that Ntr2/Ntr1-CTD prevents NTR from disrupting properly assembled spliceosomes other than the ILS. The U2 snRNP-intron interaction is disrupted in all complexes by Prp43_Ntr1GP, and in the spliceosome contacts U2 proteins and the pre-mRNA, indicating that the U2 snRNP-intron interaction is Prp43’s major target. DOI: http://dx.doi.org/10.7554/eLife.15564.001 PMID:27115347

  2. Concerted evolution of the tandemly repeated genes encoding primate U2 small nuclear RNA (the RNU2 locus) does not prevent rapid diversification of the (CT){sub n} {center_dot} (GA){sub n} microsatellite embedded within the U2 repeat unit

    SciTech Connect

    Liao, D.; Weiner, A.M.

    1995-12-10

    The RNU2 locus encoding human U2 small nuclear RNA (snRNA) is organized as a nearly perfect tandem array containing 5 to 22 copies of a 5.8-kb repeat unit. Just downstream of the U2 snRNA gene in each 5.8-kb repeat unit lies a large (CT){sub n}{center_dot}(GA){sub n} dinucleotide repeat (n {approx} 70). This form of genomic organization, in which one repeat is embedded within another, provides an unusual opportunity to study the balance of forces maintaining the homogeneity of both kinds of repeats. Using a combination of field inversion gel electrophoresis and polymerase chain reaction, we have been able to study the CT microsatellites within individual U2 tandem arrays. We find that the CT microsatellites within an RNU2 allele exhibit significant length polymorphism, despite the remarkable homogeneity of the surrounding U2 repeat units. Length polymorphism is due primarily to loss or gain of CT dinucleotide repeats, but other types of deletions, insertions, and substitutions are also frequent. Polymorphism is greatly reduced in regions where pure (CT){sub n} tracts are interrupted by occasional G residues, suggesting that irregularities stabilize both the length and the sequence of the dinucleotide repeat. We further show that the RNU2 loci of other catarrhine primates (gorilla, chimpanzee, ogangutan, and baboon) contain orthologous CT microsatellites; these also exhibit length polymorphism, but are highly divergent from each other. Thus, although the CT microsatellite is evolving far more rapidly than the rest of the U2 repeat unit, it has persisted through multiple speciation events spanning >35 Myr. The persistence of the CT microsatellite, despite polymorphism and rapid evolution, suggests that it might play a functional role in concerted evolution of the RNU2 loci, perhaps as an initiation site for recombination and/or gene conversion. 70 refs., 5 figs.

  3. Dihydroptychantol A, a macrocyclic bisbibenzyl derivative, induces autophagy and following apoptosis associated with p53 pathway in human osteosarcoma U2OS cells

    SciTech Connect

    Li Xia; Wu, William K.K.; Sun Bin; Cui Min; Liu Shanshan; Gao Jian; Lou Hongxiang

    2011-03-01

    Dihydroptychantol A (DHA), a novel macrocyclic bisbibenzyl compound extracted from liverwort Asterella angusta, has antifungal and multi-drug resistance reversal properties. Here, the chemically synthesized DHA was employed to test its anti-cancer activities in human osteosarcoma U2OS cells. Our results demonstrated that DHA induced autophagy followed by apoptotic cell death accompanied with G{sub 2}/M-phase cell cycle arrest in U2OS cells. DHA-induced autophagy was morphologically characterized by the formation of double membrane-bound autophagic vacuoles recognizable at the ultrastructural level. DHA also increased the levels of LC3-II, a marker of autophagy. Surprisingly, DHA-mediated apoptotic cell death was potentiated by the autophagy inhibitor 3-methyladenine, suggesting that autophagy may play a protective role that impedes the eventual cell death. Furthermore, p53 was shown to be involved in DHA-meditated autophagy and apoptosis. In this connection, DHA increased nuclear expression of p53, induced p53 phosphorylation, and upregulated p53 target gene p21{sup Waf1/Cip1}. In contrast, cytoplasmic p53 was reduced by DHA, which contributed to the stimulation of autophagy. In relation to the cell cycle, DHA decreased the expression of cyclin B{sub 1}, a cyclin required for progression through the G{sub 2}/M phase. Taken together, DHA induces G{sub 2}/M-phase cell cycle arrest and apoptosis in U2OS cells. DHA-induced apoptosis was preceded by the induction of protective autophagy. DHA-mediated autophagy and apoptosis are associated with the cytoplasmic and nuclear functions of p53.

  4. How Are Short Exons Flanked by Long Introns Defined and Committed to Splicing?

    PubMed

    Hollander, Dror; Naftelberg, Shiran; Lev-Maor, Galit; Kornblihtt, Alberto R; Ast, Gil

    2016-10-01

    The splice sites (SSs) delimiting an intron are brought together in the earliest step of spliceosome assembly yet it remains obscure how SS pairing occurs, especially when introns are thousands of nucleotides long. Splicing occurs in vivo in mammals within minutes regardless of intron length, implying that SS pairing can instantly follow transcription. Also, factors required for SS pairing, such as the U1 small nuclear ribonucleoprotein (snRNP) and U2AF65, associate with RNA polymerase II (RNAPII), while nucleosomes preferentially bind exonic sequences and associate with U2 snRNP. Based on recent publications, we assume that the 5' SS-bound U1 snRNP can remain tethered to RNAPII until complete synthesis of the downstream intron and exon. An additional U1 snRNP then binds the downstream 5' SS, whereas the RNAPII-associated U2AF65 binds the upstream 3' SS to facilitate SS pairing along with exon definition. Next, the nucleosome-associated U2 snRNP binds the branch site to advance splicing complex assembly. This may explain how RNAPII and chromatin are involved in spliceosome assembly and how introns lengthened during evolution with a relatively minimal compromise in splicing.

  5. How Are Short Exons Flanked by Long Introns Defined and Committed to Splicing?

    PubMed

    Hollander, Dror; Naftelberg, Shiran; Lev-Maor, Galit; Kornblihtt, Alberto R; Ast, Gil

    2016-10-01

    The splice sites (SSs) delimiting an intron are brought together in the earliest step of spliceosome assembly yet it remains obscure how SS pairing occurs, especially when introns are thousands of nucleotides long. Splicing occurs in vivo in mammals within minutes regardless of intron length, implying that SS pairing can instantly follow transcription. Also, factors required for SS pairing, such as the U1 small nuclear ribonucleoprotein (snRNP) and U2AF65, associate with RNA polymerase II (RNAPII), while nucleosomes preferentially bind exonic sequences and associate with U2 snRNP. Based on recent publications, we assume that the 5' SS-bound U1 snRNP can remain tethered to RNAPII until complete synthesis of the downstream intron and exon. An additional U1 snRNP then binds the downstream 5' SS, whereas the RNAPII-associated U2AF65 binds the upstream 3' SS to facilitate SS pairing along with exon definition. Next, the nucleosome-associated U2 snRNP binds the branch site to advance splicing complex assembly. This may explain how RNAPII and chromatin are involved in spliceosome assembly and how introns lengthened during evolution with a relatively minimal compromise in splicing. PMID:27507607

  6. Electronic structure of crystalline uranium nitrides UN, U2N3 and UN2: LCAO calculations with the basis set optimization

    NASA Astrophysics Data System (ADS)

    Evarestov, R. A.; Panin, A. I.; Bandura, A. V.; Losev, M. V.

    2008-06-01

    The results of LCAO DFT calculations of lattice parameters, cohesive energy and bulk modulus of the crystalline uranium nitrides UN, U2N3 and UN2 are presented and discussed. The LCAO computer codes Gaussian03 and Crystal06 are applied. The calculations are made with the uranium atom relativistic effective small core potential by Stuttgart-Cologne group (60 electrons in the core). The calculations include the U atom basis set optimization. Powell, Hooke-Jeeves, conjugated gradient and Box methods are implemented in the author's optimization package, being external to the codes for molecular and periodic calculations. The basis set optimization in LCAO calculations improves the agreement of the lattice parameter and bulk modulus of UN crystal with the experimental data, the change of the cohesive energy due to the optimization is small. The mixed metallic-covalent chemical bonding is found both in LCAO calculations of UN and U2N3 crystals; UN2 crystal has the semiconducting nature.

  7. Endogenous U2·U5·U6 snRNA complexes in S. pombe are intron lariat spliceosomes

    PubMed Central

    Chen, Weijun; Shulha, Hennady P.; Ashar-Patel, Ami; Yan, Jing; Green, Karin M.; Query, Charles C.; Rhind, Nick; Weng, Zhiping; Moore, Melissa J.

    2014-01-01

    Excision of introns from pre-mRNAs is mediated by the spliceosome, a multi-megadalton complex consisting of U1, U2, U4/U6, and U5 snRNPs plus scores of associated proteins. Spliceosome assembly and disassembly are highly dynamic processes involving multiple stable intermediates. In this study, we utilized a split TAP-tag approach for large-scale purification of an abundant endogenous U2·U5·U6 complex from Schizosaccharomyces pombe. RNAseq revealed this complex to largely contain excised introns, indicating that it is primarily ILS (intron lariat spliceosome) complexes. These endogenous ILS complexes are remarkably resistant to both high-salt and nuclease digestion. Mass spectrometry analysis identified 68, 45, and 43 proteins in low-salt-, high-salt-, and micrococcal nuclease-treated preps, respectively. The protein content of a S. pombe ILS complex strongly resembles that previously reported for human spliced product (P) and Saccharomyces cerevisiae ILS complexes assembled on single pre-mRNAs in vitro. However, the ATP-dependent RNA helicase Brr2 was either substoichiometric in low-salt preps or completely absent from high-salt and MNase preps. Because Brr2 facilitates spliceosome disassembly, its relative absence may explain why the ILS complex accumulates logarithmically growing cultures and the inability of S. pombe extracts to support in vitro splicing. PMID:24442611

  8. Dosage of the Abcg1-U2af1 region modifies locomotor and cognitive deficits observed in the Tc1 mouse model of Down syndrome.

    PubMed

    Marechal, Damien; Lopes Pereira, Patricia; Duchon, Arnaud; Herault, Yann

    2015-01-01

    Down syndrome (DS) results from one extra copy of human chromosome 21 and leads to several alterations including intellectual disabilities and locomotor defects. The transchromosomic Tc1 mouse model carrying an extra freely-segregating copy of human chromosome 21 was developed to better characterize the relation between genotype and phenotype in DS. The Tc1 mouse exhibits several locomotor and cognitive deficits related to DS. In this report we analyzed the contribution of the genetic dosage of 13 conserved mouse genes located between Abcg1 and U2af1, in the telomeric part of Hsa21. We used the Ms2Yah model carrying a deletion of the corresponding interval in the mouse genome to rescue gene dosage in the Tc1/Ms2Yah compound mice to determine how the different behavioral phenotypes are affected. We detected subtle changes with the Tc1/Ms2Yah mice performing better than the Tc1 individuals in the reversal paradigm of the Morris water maze. We also found that Tc1/Ms2Yah compound mutants performed better in the rotarod than the Tc1 mice. This data support the impact of genes from the Abcg1-U2af1 region as modifiers of Tc1-dependent memory and locomotor phenotypes. Our results emphasize the complex interactions between triplicated genes inducing DS features.

  9. The monitoring of gene functions on a cell-defined siRNA microarray in human bone marrow stromal and U2OS cells

    PubMed Central

    Kim, Hi Chul; Kim, Gi-Hwan; Shum, David; Cho, Ssang-Goo; Lee, Eun Ju; Kwon, Yong-Jun

    2016-01-01

    Here, we developed a cell defined siRNA microarray (CDSM) for human bone marrow stromal cells (hBMSCs) designed to control the culture of cells inside the spot area without reducing the efficiency of siRNA silencing, “Development of a cell-defined siRNA microarray for analysis of gene functionin human bone marrow stromal cells” (Kim et al., 2016 [1]). First, we confirmed that p65 protein inhibition efficiency was maintained when hBMSCs were culture for 7 days on the siRNA spot, and siRNA spot activity remained in spite of long term storage (10 days and 2 months). Additionally, we confirmed p65 protein inhibition in U2OS cells after 48 h reverse transfection. PMID:27054175

  10. Functional dissection of an enhancer-like element located within the second intron of the human U2AF1L4 gene.

    PubMed

    Didych, D A; Smirnov, N A; Kotova, E S; Akopov, S B; Nikolaev, L G; Sverdlov, E D

    2011-08-01

    A detailed functional and evolutionary analysis of an enhancer element of the human genome (enhancer 12) located in the second intron of the U2AF1L4 gene, which we identified earlier, is presented. Overlapping fragments of the studied genome region were analyzed for enhancer activity, and the site responsible for the activity of this element was identified using transient transfections of HeLa cells. Comparison of the enhancer 12 sequence with orthologous sequences from seven primate species revealed the existence of evolutionarily conserved sequences within this element. One of the identified conservative regions is likely responsible for the enhancer activity and is able to specifically interact in vitro with proteins of HeLa cell nuclear extract. The ability of orthologous primate sequences to compete with enhancer 12 for binding with HeLa cell nuclear extract proteins and to enhance the activity of the reporter gene in transient transfection of HeLa cells is demonstrated. PMID:22022969

  11. Identification of cell cycle-regulated genes periodically expressed in U2OS cells and their regulation by FOXM1 and E2F transcription factors.

    PubMed

    Grant, Gavin D; Brooks, Lionel; Zhang, Xiaoyang; Mahoney, J Matthew; Martyanov, Viktor; Wood, Tammara A; Sherlock, Gavin; Cheng, Chao; Whitfield, Michael L

    2013-12-01

    We identify the cell cycle-regulated mRNA transcripts genome-wide in the osteosarcoma-derived U2OS cell line. This results in 2140 transcripts mapping to 1871 unique cell cycle-regulated genes that show periodic oscillations across multiple synchronous cell cycles. We identify genomic loci bound by the G2/M transcription factor FOXM1 by chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) and associate these with cell cycle-regulated genes. FOXM1 is bound to cell cycle-regulated genes with peak expression in both S phase and G2/M phases. We show that ChIP-seq genomic loci are responsive to FOXM1 using a real-time luciferase assay in live cells, showing that FOXM1 strongly activates promoters of G2/M phase genes and weakly activates those induced in S phase. Analysis of ChIP-seq data from a panel of cell cycle transcription factors (E2F1, E2F4, E2F6, and GABPA) from the Encyclopedia of DNA Elements and ChIP-seq data for the DREAM complex finds that a set of core cell cycle genes regulated in both U2OS and HeLa cells are bound by multiple cell cycle transcription factors. These data identify the cell cycle-regulated genes in a second cancer-derived cell line and provide a comprehensive picture of the transcriptional regulatory systems controlling periodic gene expression in the human cell division cycle.

  12. Molecular Cytogenetic Analysis of the European Hake Merluccius merluccius (Merlucciidae, Gadiformes): U1 and U2 snRNA Gene Clusters Map to the Same Location.

    PubMed

    García-Souto, Daniel; Troncoso, Tomás; Pérez, Montse; Pasantes, Juan José

    2015-01-01

    The European hake (Merluccius merluccius) is a highly valuable and intensely fished species in which a long-term alive stock has been established in captivity for aquaculture purposes. Due to their huge economic importance, genetic studies on hakes were mostly focused on phylogenetic and phylogeographic aspects; however chromosome numbers are still not described for any of the fifteen species in the genus Merluccius. In this work we report a chromosome number of 2n = 42 and a karyotype composed of three meta/submetacentric and 18 subtelo/telocentric chromosome pairs. Telomeric sequences appear exclusively at both ends of every single chromosome. Concerning rRNA genes, this species show a single 45S rDNA cluster at an intercalary location on the long arm of subtelocentric chromosome pair 12; the single 5S rDNA cluster is also intercalary to the long arm of chromosome pair 4. While U2 snRNA gene clusters map to a single subcentromeric position on chromosome pair 13, U1 snRNA gene clusters seem to appear on almost all chromosome pairs, but showing bigger clusters on pairs 5, 13, 16, 17 and 19. The brightest signals on pair 13 are coincident with the single U2 snRNA gene cluster signals. Therefore, the use of these probes allows the unequivocal identification of at least 7 of the chromosome pairs that compose the karyotype of Merluccius merluccius thus opening the way to integrate molecular genetics and cytological data on the study of the genome of this important species. PMID:26716701

  13. Molecular Cytogenetic Analysis of the European Hake Merluccius merluccius (Merlucciidae, Gadiformes): U1 and U2 snRNA Gene Clusters Map to the Same Location

    PubMed Central

    García-Souto, Daniel; Troncoso, Tomás; Pérez, Montse; Pasantes, Juan José

    2015-01-01

    The European hake (Merluccius merluccius) is a highly valuable and intensely fished species in which a long-term alive stock has been established in captivity for aquaculture purposes. Due to their huge economic importance, genetic studies on hakes were mostly focused on phylogenetic and phylogeographic aspects; however chromosome numbers are still not described for any of the fifteen species in the genus Merluccius. In this work we report a chromosome number of 2n = 42 and a karyotype composed of three meta/submetacentric and 18 subtelo/telocentric chromosome pairs. Telomeric sequences appear exclusively at both ends of every single chromosome. Concerning rRNA genes, this species show a single 45S rDNA cluster at an intercalary location on the long arm of subtelocentric chromosome pair 12; the single 5S rDNA cluster is also intercalary to the long arm of chromosome pair 4. While U2 snRNA gene clusters map to a single subcentromeric position on chromosome pair 13, U1 snRNA gene clusters seem to appear on almost all chromosome pairs, but showing bigger clusters on pairs 5, 13, 16, 17 and 19. The brightest signals on pair 13 are coincident with the single U2 snRNA gene cluster signals. Therefore, the use of these probes allows the unequivocal identification of at least 7 of the chromosome pairs that compose the karyotype of Merluccius merluccius thus opening the way to integrate molecular genetics and cytological data on the study of the genome of this important species. PMID:26716701

  14. Cinobufagin induces autophagy-mediated cell death in human osteosarcoma U2OS cells through the ROS/JNK/p38 signaling pathway.

    PubMed

    Ma, Kun; Zhang, Chuan; Huang, Man-Yu; Li, Wu-Yin; Hu, Guo-Qiang

    2016-07-01

    The main objective of this study was to explore whether autophagy could be triggered by cinobufagin, and to clarify the role of autophagy in the antitumor effects of cinobufagin on U2OS cells and the underlying mechanisms. U2OS cells were exposed to 15, 30, 60 and 120 mg/l cinobufagin for 0, 12, 24 and 48 h. An MTT assay was used to measure cell viability. FITC-Annexin Ⅴ/PI staining and flow cytometry were used to analyze the apoptotic ratio, while apoptotic morphological changes were assessed by PI and Hoechst 33258 viable cell staining. The effects of autophagy on the cells were investigated with GFP-LC3b green fluorescence plasmid transfection and transmission electron microscopy. The levels of caspase-3, -8, - 9, cleaved PARP, LC3-II/LC3-I, p62 and the activation of JNK/p-38 were detected by western blot analysis. Reactive oxygen species (ROS) fluorescence intensity was examined under fluorescence microscopy with an analysis software system. Cell proliferation was obviously inhibited by cinobufagin in a dose- and time-dependent manner. The apoptosis ratio was gradually increased with treatment time as evidenced by flow cytometric analysis and Hoechst 33258 staining. Exposure to cinobufagin resulted in the activation of caspase-3, -8, -9, as well as cleaved PARP which indicated that cinobufagin induced caspase-dependent apoptosis. Autophagy was confirmed in the cinobufagin-treated cells as evidenced by formation of autophagosomes, accumulation of GFP-LC3 fluorescence particles as well as the upregulation of LC3-II/LC3-I levels. Inhibition of autophagy diminished apoptosis as detected by the MTT assays. Moreover the percentage of apoptotic cells decreased following pretreatment with 3-MA, CQ and si-beclin-1. Cinobufagin also induced phosphorylation of the JNK and p38 signaling pathway as well as ROS generation. The JNK and p38 inhibitors significantly attenuated coexistence of apoptosis and autophagy-related proteins. The ROS scavenger also prevented

  15. Zoledronate blocks geranylgeranylation not farnesylation to suppress human osteosarcoma U2OS cells metastasis by EMT via Rho A activation and FAK-inhibited JNK and p38 pathways

    PubMed Central

    Cheng, Hsin-Lin; Lin, Chiao-Wen; Yang, Jia-Sin; Hsieh, Ming-Ju; Yang, Shun-Fa; Lu, Ko-Hsiu

    2016-01-01

    Zoledronate is a standard treatment for preventing skeletal complications of osteoporosis and some types of cancer associated with bone metastases, but we little know whether the effect of zoledronate on metastasis of osteosarcoma. Here, we investigated the inhibitory effects of zoledronate on cell viability, motility, migration and invasion of 4 osteosarcoma cell lines (Saos2, MG-63, HOS and U2OS) by affecting cell morphology, epithelial-mesenchymal transition (EMT) and cytoskeletal organization as well as induction of E-cadherin and reduction of N-cadherin with activation of transcription factors Slug and Twist, especially in U2OS cells. Zoledronate decreased JNK and p38 phosphorylation and upper streams of focal adhesion kinase (FAK) and Src to suppress the motility, invasiveness and migration of U2OS cells. In addition to zoledronate-inhibited Rho A and Cdc42 membrane translocation and GTPγS activities, the anti-metastatic effects in U2OS cells including inhibition of adhesion were reversed by geranylgeraniol, but not farnesol. In conclusion, Zoledronate blocks geranylgeranylation not farnesylation to suppress human osteosarcoma U2OS cell-matrix and cell-cell interactions, migration potential, the invasive activity, and the adhesive ability by EMT via Rho A activation and FAK-inhibited JNK and p38 pathways. PMID:26848867

  16. Zoledronate blocks geranylgeranylation not farnesylation to suppress human osteosarcoma U2OS cells metastasis by EMT via Rho A activation and FAK-inhibited JNK and p38 pathways.

    PubMed

    Cheng, Hsin-Lin; Lin, Chiao-Wen; Yang, Jia-Sin; Hsieh, Ming-Ju; Yang, Shun-Fa; Lu, Ko-Hsiu

    2016-03-01

    Zoledronate is a standard treatment for preventing skeletal complications of osteoporosis and some types of cancer associated with bone metastases, but we little know whether the effect of zoledronate on metastasis of osteosarcoma. Here, we investigated the inhibitory effects of zoledronate on cell viability, motility, migration and invasion of 4 osteosarcoma cell lines (Saos2, MG-63, HOS and U2OS) by affecting cell morphology, epithelial-mesenchymal transition (EMT) and cytoskeletal organization as well as induction of E-cadherin and reduction of N-cadherin with activation of transcription factors Slug and Twist, especially in U2OS cells. Zoledronate decreased JNK and p38 phosphorylation and upper streams of focal adhesion kinase (FAK) and Src to suppress the motility, invasiveness and migration of U2OS cells. In addition to zoledronate-inhibited Rho A and Cdc42 membrane translocation and GTPγS activities, the anti-metastatic effects in U2OS cells including inhibition of adhesion were reversed by geranylgeraniol, but not farnesol. In conclusion, Zoledronate blocks geranylgeranylation not farnesylation to suppress human osteosarcoma U2OS cell-matrix and cell-cell interactions, migration potential, the invasive activity, and the adhesive ability by EMT via Rho A activation and FAK-inhibited JNK and p38 pathways. PMID:26848867

  17. Microarray and ChIP-seq data analysis revealed changes in p53-mediated transcriptional regulation in Nutlin-3-treated U2OS cells.

    PubMed

    Zhao, Song; Niu, Feng; Xu, Chang-Yan; Ye, Long; Bi, Gui-Bin; Chen, Lin; Gong, Ping; Tian, Gang; Nie, Tian-Hong

    2015-09-01

    Integrative analysis of chromatin immunoprecipitation-sequencing (ChIP-seq) data and microarray data was performed to illustrate the effect of Nutlin‑3 on promoter selectivity and transcriptional regulation by the tumor suppressor p53 in U2OS human osteosarcoma cells. Raw data (accession number, GSE46642) were downloaded from Gene Expression Omnibus. Differential analyses were performed using package limma of R software. Gene ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were performed for the differentially expressed genes (DEGs) using the Database for Annotation, Visualization and Integration Discovery. Integrative analysis of ChIP‑seq data and microarray data were confirmed with ChIP‑Array. A total of 565 DEGs were identified, including 373 upregulated genes and 192 downregulated genes. Genes involved in the p53 signaling pathway, cell cycle, DNA replication, cytokine‑cytokine receptor interaction and melanoma were markedly over‑represented in the DEGs. A total of 39 DEGs were directly regulated by p53 and two were the transcription factors (TFs), E2F2 and HOXA1. E2F2 regulated 25 DEGs, while HOXA1 regulated one DEG. The cell cycle, p53 signaling pathway, melanoma and pathways involved in cancer were enriched in the direct and indirect target genes. Changes in the p53‑binding pattern induced by Nutlin‑3 were described in the present study, which may advance the understanding of the regulatory network of p53 in osteosarcoma and aid in the development of novel therapies.

  18. Analysis of a cDNA clone expressing a human autoimmune antigen: full-length sequence of the U2 small nuclear RNA-associated B antigen

    SciTech Connect

    Habets, W.J.; Sillekens, P.T.G.; Hoet, M.H.; Schalken, J.A.; Roebroek, A.J.M.; Leunissen, J.A.M.; Van de Ven, W.J.M.; Van Venrooij, W.J.

    1987-04-01

    A U2 small nuclear RNA-associated protein, designated B'', was recently identified as the target antigen for autoimmune sera from certain patients with systemic lupus erythematosus and other rheumatic diseases. Such antibodies enabled them to isolate cDNA clone lambdaHB''-1 from a phage lambdagt11 expression library. This clone appeared to code for the B'' protein as established by in vitro translation of hybrid-selected mRNA. The identity of clone lambdaHB''-1 was further confirmed by partial peptide mapping and analysis of the reactivity of the recombinant antigen with monospecific and monoclonal antibodies. Analysis of the nucleotide sequence of the 1015-base-pair cDNA insert of clone lambdaHB''-1 revealed a large open reading frame of 800 nucleotides containing the coding sequence for a polypeptide of 25,457 daltons. In vitro transcription of the lambdaHB''-1 cDNA insert and subsequent translation resulted in a protein product with the molecular size of the B'' protein. These data demonstrate that clone lambdaHB''-1 contains the complete coding sequence of this antigen. The deduced polypeptide sequence contains three very hydrophilic regions that might constitute RNA binding sites and/or antigenic determinants. These findings might have implications both for the understanding of the pathogenesis of rheumatic diseases as well as for the elucidation of the biological function of autoimmune antigens.

  19. Processing of U-2.5Zr-7.5Nb and U-3Zr-9Nb alloys by sintering process

    SciTech Connect

    Dos Santos, A. M. M.; Ferraz, W. B.; Lameiras, F. S.; Mazzeu, T. D. O.

    2012-07-01

    To minimize the risk of nuclear proliferation, there is worldwide interest in reducing fuel enrichment of research and test reactors. To achieve this objective while still guaranteeing criticality and cycle length requirements, there is need of developing high density uranium metallic fuels. Alloying elements such as Zr, Nb and Mo are added to uranium to improve fuel performance in reactors. In this context, the Centro de Desenvolvimento da Tecnologia Nuclear (CDTN) is developing the U-2.5Zr-7.5Nb and U-3Zr-9Nb (weight %) alloys by the innovative process of sintering that utilizes raw materials in the form of powders. The powders were pressed at 400 MPa and then sintered under a vacuum of about 1x10{sup -4} Torr at temperatures ranging from 1050 deg. to 1500 deg.C. The densities of the alloys were measured geometrically and by hydrostatic method and the phases identified by X ray diffraction (XRD). The microstructures of the pellets were observed by scanning electron microscopy (SEM) and the alloying elements were analyzed by energy dispersive X-ray spectroscopy (EDS). The results obtained showed the fuel density to slightly increase with the sintering temperature. The highest density achieved was approximately 80% of theoretical density. It was observed in the pellets a superficial oxide layer formed during the sintering process. (authors)

  20. The Activation-Induced Assembly of an RNA/Protein Interactome Centered on the Splicing Factor U2AF2 Regulates Gene Expression in Human CD4 T Cells.

    PubMed

    Whisenant, Thomas C; Peralta, Eigen R; Aarreberg, Lauren D; Gao, Nina J; Head, Steven R; Ordoukhanian, Phillip; Williamson, Jamie R; Salomon, Daniel R

    2015-01-01

    Activation of CD4 T cells is a reaction to challenges such as microbial pathogens, cancer and toxins that defines adaptive immune responses. The roles of T cell receptor crosslinking, intracellular signaling, and transcription factor activation are well described, but the importance of post-transcriptional regulation by RNA-binding proteins (RBPs) has not been considered in depth. We describe a new model expanding and activating primary human CD4 T cells and applied this to characterizing activation-induced assembly of splicing factors centered on U2AF2. We immunoprecipitated U2AF2 to identify what mRNA transcripts were bound as a function of activation by TCR crosslinking and costimulation. In parallel, mass spectrometry revealed the proteins incorporated into the U2AF2-centered RNA/protein interactome. Molecules that retained interaction with the U2AF2 complex after RNAse treatment were designated as "central" interactome members (CIMs). Mass spectrometry also identified a second class of activation-induced proteins, "peripheral" interactome members (PIMs), that bound to the same transcripts but were not in physical association with U2AF2 or its partners. siRNA knockdown of two CIMs and two PIMs caused changes in activation marker expression, cytokine secretion, and gene expression that were unique to each protein and mapped to pathways associated with key aspects of T cell activation. While knocking down the PIM, SYNCRIP, impacts a limited but immunologically important set of U2AF2-bound transcripts, knockdown of U2AF1 significantly impairs assembly of the majority of protein and mRNA components in the activation-induced interactome. These results demonstrated that CIMs and PIMs, either directly or indirectly through RNA, assembled into activation-induced U2AF2 complexes and play roles in post-transcriptional regulation of genes related to cytokine secretion. These data suggest an additional layer of regulation mediated by the activation-induced assembly of RNA

  1. The Activation-Induced Assembly of an RNA/Protein Interactome Centered on the Splicing Factor U2AF2 Regulates Gene Expression in Human CD4 T Cells

    PubMed Central

    Aarreberg, Lauren D.; Gao, Nina J.; Head, Steven R.; Ordoukhanian, Phillip; Williamson, Jamie R.; Salomon, Daniel R.

    2015-01-01

    Activation of CD4 T cells is a reaction to challenges such as microbial pathogens, cancer and toxins that defines adaptive immune responses. The roles of T cell receptor crosslinking, intracellular signaling, and transcription factor activation are well described, but the importance of post-transcriptional regulation by RNA-binding proteins (RBPs) has not been considered in depth. We describe a new model expanding and activating primary human CD4 T cells and applied this to characterizing activation-induced assembly of splicing factors centered on U2AF2. We immunoprecipitated U2AF2 to identify what mRNA transcripts were bound as a function of activation by TCR crosslinking and costimulation. In parallel, mass spectrometry revealed the proteins incorporated into the U2AF2-centered RNA/protein interactome. Molecules that retained interaction with the U2AF2 complex after RNAse treatment were designated as “central” interactome members (CIMs). Mass spectrometry also identified a second class of activation-induced proteins, “peripheral” interactome members (PIMs), that bound to the same transcripts but were not in physical association with U2AF2 or its partners. siRNA knockdown of two CIMs and two PIMs caused changes in activation marker expression, cytokine secretion, and gene expression that were unique to each protein and mapped to pathways associated with key aspects of T cell activation. While knocking down the PIM, SYNCRIP, impacts a limited but immunologically important set of U2AF2-bound transcripts, knockdown of U2AF1 significantly impairs assembly of the majority of protein and mRNA components in the activation-induced interactome. These results demonstrated that CIMs and PIMs, either directly or indirectly through RNA, assembled into activation-induced U2AF2 complexes and play roles in post-transcriptional regulation of genes related to cytokine secretion. These data suggest an additional layer of regulation mediated by the activation-induced assembly

  2. Fine structure of the "PcG body" in human U-2 OS cells established by correlative light-electron microscopy.

    PubMed

    Smigová, Jana; Juda, Pavel; Cmarko, Dušan; Raška, Ivan

    2011-01-01

    Polycomb group (PcG) proteins of the Polycomb repressive complex 1 (PRC1) are found to be diffusely distributed in nuclei of cells from various species. However they can also be localized in intensely fluorescent foci, whether imaged using GFP fusions to proteins of PRC1 complex, or by conventional immunofluorescence microscopy. Such foci are termed PcG bodies, and are believed to be situated in the nuclear intechromatin compartment. However, an ultrastructural description of the PcG body has not been reported to date. To establish the ultrastructure of PcG bodies in human U-2 OS cells stably expressing recombinant polycomb BMI1-GFP protein, we used correlative light-electron microscopy (CLEM) implemented with high-pressure freezing, cryosubstitution and on-section labeling of BMI1 protein with immunogold. This approach allowed us to clearly identify fluorescent PcG bodies, not as distinct nuclear bodies, but as nuclear domains enriched in separated heterochromatin fascicles. Importantly, high-pressure freezing and cryosubstitution allowed for a high and clear-cut immunogold BMI1 labeling of heterochromatin structures throughout the nucleus. The density of immunogold labeled BMI1 in the heterochromatin fascicles corresponding to fluorescent "PcG bodies" did not differ from the density of labeling of heterochromatin fascicles outside of the "PcG bodies". Accordingly, an appearance of the fluorescent "PcG bodies" seems to reflect a local accumulation of the labeled heterochromatin structures in the investigated cells. The results of this study should allow expansion of the knowledge about the biological relevance of the "PcG bodies" in human cells.

  3. Fine structure of the “PcG body” in human U-2 OS cells established by correlative light-electron microscopy

    PubMed Central

    Juda, Pavel; Cmarko, Dušan

    2011-01-01

    Polycomb group (PcG) proteins of the Polycomb repressive complex 1 (PRC1) are found to be diffusely distributed in nuclei of cells from various species. However they can also be localized in intensely fluorescent foci, whether imaged using GFP fusions to proteins of PRC1 complex, or by conventional immunofluorescence microscopy. Such foci are termed PcG bodies, and are believed to be situated in the nuclear intechromatin compartment. However, an ultrastructural description of the PcG body has not been reported to date. To establish the ultrastructure of PcG bodies in human U-2 OS cells stably expressing recombinant polycomb BMI1-GFP protein, we used correlative light-electron microscopy (CLEM) implemented with high-pressure freezing, cryosubstitution and on-section labeling of BMI1 protein with immunogold. This approach allowed us to clearly identify fluorescent PcG bodies, not as distinct nuclear bodies, but as nuclear domains enriched in separated heterochromatin fascicles. Importantly, high-pressure freezing and cryosubstitution allowed for a high and clear-cut immunogold BMI1 labeling of heterochromatin structures throughout the nucleus. The density of immunogold labeled BMI1 in the heterochromatin fascicles corresponding to fluorescent “PcG bodies” did not differ from the density of labeling of heterochromatin fascicles outside of the “PcG bodies”. Accordingly, an appearance of the fluorescent “PcG bodies” seems to reflect a local accumulation of the labeled heterochromatin structures in the investigated cells. The results of this study should allow expansion of the knowledge about the biological relevance of the “PcG bodies” in human cells. PMID:21818415

  4. Chromosomal Mapping of Repetitive DNA Sequences in Five Species of Astyanax (Characiformes, Characidae) Reveals Independent Location of U1 and U2 snRNA Sites and Association of U1 snRNA and 5S rDNA.

    PubMed

    Silva, Duilio M Z A; Utsunomia, Ricardo; Pansonato-Alves, José C; Oliveira, Cláudio; Foresti, Fausto

    2015-01-01

    Astyanax is a genus of Characidae fishes currently composed of 155 valid species. Previous cytogenetic studies revealed high chromosomal diversification among them, and several studies have been performed using traditional cytogenetic techniques to investigate karyotypes and chromosomal locations of 18S and 5S rDNA genes. However, only a few studies are currently available about other repetitive sequences. Here, the chromosomal location of small nuclear RNA genes, identified as U1 and U2 snRNA clusters, was established and compared to the distribution of 5S rDNA and histone clusters in 5 Astyanax species (A. paranae, A. fasciatus, A. bockmanni, A. altiparanae, and A. jordani) using FISH. The cytogenetic mapping of U1 and U2 snRNA demonstrated a conserved pattern in the number of sites per genome independent of the location in Astyanax species. The location of the U1 snRNA gene was frequently associated with 5S rDNA sequences, indicating a possible interaction between the distinct repetitive DNA families. Finally, comparisons involving the location of U1 and U2 snRNA clusters in the chromosomes of Astyanax species revealed a very diverse pattern, suggesting that many rearrangements have occurred during the diversification process of this group. PMID:26329975

  5. Chromosomal Mapping of Repetitive DNA Sequences in Five Species of Astyanax (Characiformes, Characidae) Reveals Independent Location of U1 and U2 snRNA Sites and Association of U1 snRNA and 5S rDNA.

    PubMed

    Silva, Duilio M Z A; Utsunomia, Ricardo; Pansonato-Alves, José C; Oliveira, Cláudio; Foresti, Fausto

    2015-01-01

    Astyanax is a genus of Characidae fishes currently composed of 155 valid species. Previous cytogenetic studies revealed high chromosomal diversification among them, and several studies have been performed using traditional cytogenetic techniques to investigate karyotypes and chromosomal locations of 18S and 5S rDNA genes. However, only a few studies are currently available about other repetitive sequences. Here, the chromosomal location of small nuclear RNA genes, identified as U1 and U2 snRNA clusters, was established and compared to the distribution of 5S rDNA and histone clusters in 5 Astyanax species (A. paranae, A. fasciatus, A. bockmanni, A. altiparanae, and A. jordani) using FISH. The cytogenetic mapping of U1 and U2 snRNA demonstrated a conserved pattern in the number of sites per genome independent of the location in Astyanax species. The location of the U1 snRNA gene was frequently associated with 5S rDNA sequences, indicating a possible interaction between the distinct repetitive DNA families. Finally, comparisons involving the location of U1 and U2 snRNA clusters in the chromosomes of Astyanax species revealed a very diverse pattern, suggesting that many rearrangements have occurred during the diversification process of this group.

  6. Dryofragin inhibits the migration and invasion of human osteosarcoma U2OS cells by suppressing MMP-2/9 and elevating TIMP-1/2 through PI3K/AKT and p38 MAPK signaling pathways.

    PubMed

    Su, Yan; Wan, Daqian; Song, Wenqi

    2016-08-01

    Dryofragin, a phloroglucinol derivative extracted from Dryopteris fragrans (L.) Schott, was found to inhibit proliferation and induce apoptosis of tumor cells. However, the mechanism involved in the suppression of cancer cell metastasis by dryofragin remains unclear. Our study investigated the mechanisms for the antitumor properties of dryofragin on the migration and invasion of human osteosarcoma U2OS cells. Dryofragin suppressed the migration and invasive ability of U2OS cells, and it decreased the expression of MMP-2 and MMP-9 and elevated the expression of TIMP-1 and TIMP-2. Western blotting assays indicated that dryofragin was effective in suppressing the phosphorylation of phosphatidylinositide-3 kinase (PI3K), Akt, and p38 MAPK. These results suggest that dryofragin inhibited U2OS cell migration and invasion by reducing the expression of MMP-2 and MMP-9 and elevating the expression of TIMP-1 and TIMP-2 through the PI3K/AKT and p38 MAPK signaling pathways. Above all, we conclude that dryofragin represents an anti-invasive agent and may potentially be applicable in osteosarcoma therapy. PMID:27243922

  7. Inactivation of the WNT5A Alternative Promoter B Is Associated with DNA Methylation and Histone Modification in Osteosarcoma Cell Lines U2OS and SaOS-2.

    PubMed

    Vaidya, Himani; Rumph, Candie; Katula, Karen S

    2016-01-01

    WNT5A is a secreted ligand involved in Wnt pathway signaling and has a role in cell movement and differentiation. Altered WNT5A expression is associated with various cancers, although in most studies the focus has been on only one of the known WNT5A isoforms. In this study, we analyzed expression from two of the major WNT5A promoters, termed promoter A and promoter B, in normal human osteoblasts, SaOS-2 and U2OS osteosarcoma cell lines, and osteosarcoma tumor tissue. We found that both promoters A and B are active in normal osteoblasts with nearly 11-fold more promoter B than A transcripts. Promoter B but not promoter A transcripts are decreased or nearly undetectable in the SaOS-2 and U2OS cell lines and osteosarcoma tumor tissues. Transient transfection of promoter A and promoter B reporter constructs confirmed that SaOS-2 cells have the necessary factors to transcribe both promoters. Bisulfite sequencing analysis revealed that three CpG enriched regions upstream of the promoter B exon 1βare highly methylated in both SaOS-2 and U2OS cells. The CpG island sub-region R6 located in promoter B exon 1β was approximately 51% methylated in SaOS-2 and 25% methylated in U2OS. Region 3 was approximately 28% methylated in normal osteoblasts, whereas the others were unmethylated. Promoter B was re-activated by treatment of SaOS-2 cells with 1 μM 5-azacytidine, which was associated with only a small insignificant change in methylation of sub-region R6. ChIP analysis of U2OS and SaOS-2 cells indicated that the promoter B region is less enriched in the active histone mark H3K4me3, in comparison to promoter A and that there is increased enrichment of the repressive mark H3K27me3 in association with the promoter B genomic region in the cell line SaOS-2. These findings show that epigenetic inactivation of the WNT5A promoter B involves both DNA methylation and histone modifications and suggest that differential expression of the WNT5A alternative promoters A and B is a

  8. Gas-phase reactions of the bare Th2+ and U2+ ions with small alkanes, CH4, C2H6, and C3H8: experimental and theoretical study of elementary organoactinide chemistry.

    PubMed

    Di Santo, Emanuela; Santos, Marta; Michelini, Maria C; Marçalo, Joaquim; Russo, Nino; Gibson, John K

    2011-02-16

    The gas-phase reactions of two dipositive actinide ions, Th(2+) and U(2+), with CH(4), C(2)H(6), and C(3)H(8) were studied by both experiment and theory. Fourier transform ion cyclotron resonance mass spectrometry was employed to study the bimolecular ion-molecule reactions; the potential energy profiles (PEPs) for the reactions, both observed and nonobserved, were computed by density functional theory (DFT). The experiments revealed that Th(2+) reacts with all three alkanes, including CH(4) to produce ThCH(2)(2+), whereas U(2+) reacts with C(2)H(6) and C(3)H(8), with different product distributions than for Th(2+). The comparative reactivities of Th(2+) and U(2+) toward CH(4) are well explained by the computed PEPs. The PEPs for the reactions with C(2)H(6) effectively rationalize the observed reaction products, ThC(2)H(2)(2+) and UC(2)H(4)(2+). For C(3)H(8) several reaction products were experimentally observed; these and additional potential reaction pathways were computed. The DFT results for the reactions with C(3)H(8) are consistent with the observed reactions and the different products observed for Th(2+) and U(2+); however, several exothermic products which emerge from energetically favorable PEPs were not experimentally observed. The comparison between experiment and theory reveals that DFT can effectively exclude unfavorable reaction pathways, due to energetic barriers and/or endothermic products, and can predict energetic differences in similar reaction pathways for different ions. However, and not surprisingly, a simple evaluation of the PEP features is insufficient to reliably exclude energetically favorable pathways. The computed PEPs, which all proceed by insertion, were used to evaluate the relationship between the energetics of the bare Th(2+) and U(2+) ions and the energies for C-H and C-C activation. It was found that the computed energetics for insertion are entirely consistent with the empirical model which relates insertion efficiency to the

  9. Isolation of eukaryotic ribosomal proteins. Purification and characterization of the 40 S ribosomal subunit proteins Sa, Sc, S3a, S3b, S5', S9, S10, S11, S12, S14, S15, S15', S16, S17, S18, S19, S20, S21, S26, S27', and S29.

    PubMed

    Collatz, E; Ulbrich, N; Tsurugi, K; Lightfoot, H N; MacKinlay, W; Lin, A; Wool, I G

    1977-12-25

    The proteins of the small subunit of rat liver ribosomes were separated into five main groups by stepwise elution from carboxymethylcellulose with LiCl at pH 6.5. Twenty-one proteins (Sa, Sc, S3a, S3b, S5', S9, S10, S11, S12, S14, S15, S15', S16, S17, S18, S19, S20, S21, S26, S27', and S29) were isolated from three groups (A40, C40, and D40) by ion exchange chromatography on DEAE-cellulose, carboxymethylcellulose, and phosphocellulose and by filtration through Sephadex. The amount of protein obtained varied from 0.1 to 11 mg. Six of the proteins (S5', S10, S11, S18, S19, and S27') had no detectable contamination; the impurities in the others were no greater than 9%. The molecular weight of the proteins was estimated by polyacrylamide gel electrophoresis in sodium dodecyl sulfate; the amino acid composition was determined.

  10. A 23Na magic angle spinning nuclear magnetic resonance, XANES, and high-temperature X-ray diffraction study of NaUO3, Na4UO5, and Na2U2O7.

    PubMed

    Smith, A L; Raison, P E; Martel, L; Charpentier, T; Farnan, I; Prieur, D; Hennig, C; Scheinost, A C; Konings, R J M; Cheetham, A K

    2014-01-01

    The valence state of uranium has been confirmed for the three sodium uranates NaU(V)O3/[Rn](5f(1)), Na4U(VI)O5/[Rn](5f(0)), and Na2U(VI)2O7/[Rn](5f(0)), using X-ray absorption near-edge structure (XANES) spectroscopy. Solid-state (23)Na magic angle spinning nuclear magnetic resonance (MAS NMR) measurements have been performed for the first time, yielding chemical shifts at -29.1 (NaUO3), 15.1 (Na4UO5), and -14.1 and -19 ppm (Na1 8-fold coordinated and Na2 7-fold coordinated in Na2U2O7), respectively. The [Rn]5f(1) electronic structure of uranium in NaUO3 causes a paramagnetic shift in comparison to Na4UO5 and Na2U2O7, where the electronic structure is [Rn]5f(0). A (23)Na multi quantum magic angle spinning (MQMAS) study on Na2U2O7 has confirmed a monoclinic rather than rhombohedral structure with evidence for two distinct Na sites. DFT calculations of the NMR parameters on the nonmagnetic compounds Na4UO5 and Na2U2O7 have permitted the differentiation between the two Na sites of the Na2U2O7 structure. The linear thermal expansion coefficients of all three compounds have been determined using high-temperature X-ray diffraction: αa = 22.7 × 10(-6) K(-1), αb = 12.9 × 10(-6) K(-1), αc = 16.2 × 10(-6) K(-1), and αvol = 52.8 × 10(-6) K(-1) for NaUO3 in the range 298-1273 K; αa = 37.1 × 10(-6) K(-1), αc = 6.2 × 10(-6) K(-1), and αvol = 81.8 × 10(-6) K(-1) for Na4UO5 in the range 298-1073 K; αa = 6.7 × 10(-6) K(-1), αb = 14.4 × 10(-6) K(-1), αc = 26.8 × 10(-6) K(-1), αβ = -7.8 × 10(-6) K(-1), and αvol = -217.6 × 10(-6) K(-1) for Na2U2O7 in the range 298-573 K. The α to β phase transition reported for the last compound above about 600 K was not observed in the present studies, either by high-temperature X-ray diffraction or by differential scanning calorimetry. PMID:24350659

  11. Transmission electron microscopy analysis for the process of crystallization of \\text{C}{{\\text{u}}_{2}}\\text{ZnSn}{{\\text{S}}_{4}} film from sputtered Zn/CuSn precursor

    NASA Astrophysics Data System (ADS)

    Li, Wei; Chen, Jian; Yan, Chang; Liu, Fangyang; Hao, Xiaojing

    2014-05-01

    The mechanism of crystallization of \\text{C}{{\\text{u}}_{2}}\\text{ZnSn}{{\\text{S}}_{4}} (CZTS) made by the sulfurization of a sputtered Zn/CuSn stack was studied by transmission electron microscopy. At 250\\;{}^\\circ \\text{C}, the Zn buried at the bottom of the metallic stack was found to be driven to the upper layer by alloying with Cu and reacting with S to form ZnS. At 500\\;{}^\\circ \\text{C}, CZTS was found to be formed and elemental Sn was observed in the vicinity of the back contact region, while large quantities of \\text{C}{{\\text{u}}_{2}}\\text{S} and ZnS were segregated at the film surface. At 575\\;{}^\\circ \\text{C}, \\text{C}{{\\text{u}}_{2}}\\text{Sn}{{\\text{S}}_{3}} was found, while all the elemental Sn had vanished. Upon extending the duration of the annealing to 10 min, at 575\\;{}^\\circ \\text{C}, all the secondary phases except ZnS were consumed and the CZTS formation was completed, while around 200 nm of \\text{Mo}{{\\text{S}}_{2}} was formed between the CZTS layer and Mo back contact. The finished solar cell exhibits an efficiency of 2.66%, an open-circuit voltage of 666.9 mV and a short-circuit current density of 9.14\\;\\text{mA}\\;\\text{c}{{\\text{m}}^{-2}}. The solar cell performance is possibly limited by the thick \\text{Mo}{{\\text{S}}_{2}} layer and the large density of voids in the back contact region due to the Sn loss at high sulfurization temperatures.

  12. Inhibition of fatty acid synthase suppresses U-2 OS cell invasion and migration via downregulating the activity of HER2/PI3K/AKT signaling pathway in vitro

    SciTech Connect

    Wang, Tao Fang; Wang, Heng; Peng, Ai Fen; Luo, Qing Feng; Liu, Zhi Li; Zhou, Rong Ping; Gao, Song; Zhou, Yang; Chen, Wen Zhao

    2013-10-18

    Highlights: •We investigate the relationship between FASN and HER2 or p-HER2 by IHC in OS tissues. •We construct FASN-specific RNAi plasmid. •Inhibiting FASN down-regulates HER2/PI3K/AKT cell signaling in U-2 OS. •Inhibiting FASN blocks U-2 OS cell invasion and migration. -- Abstract: FASN plays an important role in the malignant phenotype of various tumors. Our previous studies show that inhibition FASN could induce apoptosis and inhibit proliferation in human osteosarcoma (OS) cell in vivo and vitro. The aim in this study was to investigate the effect of inhibition FASN on the activity of HER2/PI3K/AKT axis and invasion and migration of OS cell. The expression of FASN, HER2 and p-HER2(Y1248) proteins was detected by immunohistochemistry in OS tissues from 24 patients with pulmonary metastatic disease, and the relationship between FASN and p-HER2 as well as HER2 was investigated. The results showed that there was a positive correlation between FASN and HER2 as well as p-HER2 protein expression. The U-2 OS cells were transfected with either the FASN specific RNAi plasmid or the negative control RNAi plasmid. FASN mRNA was measured by RT-PCR. Western blot assays was performed to examine the protein expression of FASN, HER2, p-HER2(Y1248), PI3K, Akt and p-Akt (Ser473). Migration and invasion of cells were investigated by wound healing and transwell invasion assays. The results showed that the activity of HER2/PI3K/AKT signaling pathway was suppressed by inhibiting FASN. Meanwhile, the U-2OS cells migration and invasion were also impaired by inhibiting the activity of FASN/HER2/PI3K/AKT. Our results indicated that inhibition of FASN suppresses OS cell invasion and migration via down-regulation of the “HER2/PI3K/AKT” axis in vitro. FASN blocker may be a new therapeutic strategy in OS management.

  13. The nuclear cap-binding complex interacts with the U4/U6·U5 tri-snRNP and promotes spliceosome assembly in mammalian cells.

    PubMed

    Pabis, Marta; Neufeld, Noa; Steiner, Michaela C; Bojic, Teodora; Shav-Tal, Yaron; Neugebauer, Karla M

    2013-08-01

    The nuclear cap-binding complex (CBC) binds to the 7-methyl guanosine cap present on every RNA polymerase II transcript. CBC has been implicated in many aspects of RNA biogenesis; in addition to roles in miRNA biogenesis, nonsense-mediated decay, 3'-end formation, and snRNA export from the nucleus, CBC promotes pre-mRNA splicing. An unresolved question is how CBC participates in splicing. To investigate CBC's role in splicing, we used mass spectrometry to identify proteins that copurify with mammalian CBC. Numerous components of spliceosomal snRNPs were specifically detected. Among these, three U4/U6·U5 snRNP proteins (hBrr2, hPrp4, and hPrp31) copurified with CBC in an RNA-independent fashion, suggesting that a significant fraction of CBC forms a complex with the U4/U6·U5 snRNP and that the activity of CBC might be associated with snRNP recruitment to pre-mRNA. To test this possibility, CBC was depleted from HeLa cells by RNAi. Chromatin immunoprecipitation and live-cell imaging assays revealed decreased cotranscriptional accumulation of U4/U6·U5 snRNPs on active transcription units, consistent with a requirement for CBC in cotranscriptional spliceosome assembly. Surprisingly, recruitment of U1 and U2 snRNPs was also affected, indicating that RNA-mediated interactions between CBC and snRNPs contribute to splicing. On the other hand, CBC depletion did not impair snRNP biogenesis, ruling out the possibility that decreased snRNP recruitment was due to changes in nuclear snRNP concentration. Taken together, the data support a model whereby CBC promotes pre-mRNA splicing through a network of interactions with and among spliceosomal snRNPs during cotranscriptional spliceosome assembly.

  14. Efficient production of lignocellulolytic enzymes xylanase, β-xylosidase, ferulic acid esterase and β-glucosidase by the mutant strain Aspergillus awamori 2B.361 U2/1

    PubMed Central

    Gottschalk, Leda Maria Fortes; de Sousa Paredes, Raquel; Teixeira, Ricardo Sposina Sobral; da Silva, Ayla Sant’Ana; da Silva Bon, Elba Pinto

    2013-01-01

    The production of xylanase, β-xylosidase, ferulic acid esterase and β-glucosidase by Aspergillus awamori 2B.361 U2/1, a hyper producer of glucoamylase and pectinase, was evaluated using selected conditions regarding nitrogen nutrition. Submerged cultivations were carried out at 30 °C and 200 rpm in growth media containing 30 g wheat bran/L as main carbon source and either yeast extract, ammonium sulfate, sodium nitrate or urea, as nitrogen sources; in all cases it was used a fixed molar carbon to molar nitrogen concentration of 10.3. The use of poor nitrogen sources favored the accumulation of xylanase, β-xylosidase and ferulic acid esterase to a peak concentrations of 44,880; 640 and 118 U/L, respectively, for sodium nitrate and of 34,580, 685 and 170 U/L, respectively, for urea. However, the highest β-glucosidase accumulation of 10,470 U/L was observed when the rich organic nitrogen source yeast extract was used. The maxima accumulation of filter paper activity, xylanase, β-xylosidase, ferulic acid esterase and β-glucosidase by A. awamori 2B.361 U2/1 was compared to that produced by Trichoderma reesei Rut-C30. The level of β-glucosidase was over 17-fold higher for the Aspergillus strain, whereas the levels of xylanase and β-xylosidase were over 2-fold higher. This strain also produced ferulic acid esterase (170 U/L), which was not detected in the T. reesei culture. PMID:24294256

  15. The LANDSAT story: Module U-2

    NASA Technical Reports Server (NTRS)

    1980-01-01

    A review of the various LANDSAT program elements which are relevant to user participation is provided. Sources for additional information and assistance where potential users may acquire more details and further guidance in using LANDSAT data are identified. The multispectral imagery capability of the LANDSAT satellites is emphasized.

  16. Molecular characterization and chromosomal mapping of the 5S rRNA gene in Solea senegalensis: a new linkage to the U1, U2, and U5 small nuclear RNA genes.

    PubMed

    Manchado, Manuel; Zuasti, Eugenia; Cross, Ismael; Merlo, Alejandro; Infante, Carlos; Rebordinos, Laureana

    2006-01-01

    Some units of the 5S rDNA of Solea senegalensis were amplified by PCR and sequenced. Three main PCR products (227, 441, and 2166 bp) were identified. The 227- and 441-bp fragments were characterized by highly divergent nontranscribed spacer sequences (referred to as NTS-I and NTS-II) that were 109 and 324 bp long, respectively, yet their coding sequences were nearly identical. The 2166-bp 5S rDNA unit was composed of two 5S rRNA genes separated by NTS-I and followed by a 1721-bp spacer containing the U2, U5, and U1 small nuclear RNA genes (snRNAs). They were inverted and arranged in the transcriptional direction opposite that of the 5S rRNA gene. This simultaneous linkage of 3 different snRNAs had never been observed before. The PCR products were used as probes in fluorescence in situ hybridization experiments to locate the corresponding loci on the chromosomes of S. senegalensis. A major 5S rDNA chromosomal site was located along most of the short arm of a submetacentric pair, while a minor site was detected near the centromeric region of an acrocentric pair.

  17. Dealing with Climate Change and Variability in the Growing Season: a U2U Decision Support Tool for Central United States Corn Producers Based on Corn Growing Degree Days

    NASA Astrophysics Data System (ADS)

    Angel, J. R.; Todey, D.; Massey, R.; Widhalm, M.; Biehl, L. L.; Andresen, J.

    2014-12-01

    Climate extremes are a major challenge for corn producers in the central United States. Among those extremes are wet springs that lead to planting delays, late spring and early fall frosts that can damage crops, and extreme summer temperatures either too warm or too cool. A newly-operational corn growing degree-day (CGDD) tool helps producers manage and adapt to these extremes. For example, a challenge in recent years has been exceptionally wet springs that have led to significant planting delays. Producers have been forced to re-assess their planting strategies on short notice, such as switching to a faster-growing but lower-yielding hybrids. With this pattern of wetter springs projected to continue or worsen in the central United States, the problem will remain and likely get worse. Another example is helping producers identify the risk of early or late frost/freezes. The CGDD tool puts current conditions into a 30-year historical perspective and offers trend projections (based on climatology or forecasts) through the end of the calendar year. Corn, or sometimes called modified, growing degree-days use a temperature base of 10 C (50 F) and a ceiling of 30 C (86 F) and is strongly correlated with the development of the corn crop. This tool was developed as part of USDA-supported U2U Useful to Usable Project for transforming climate variability and change information for cereal crop producers.

  18. On the Quasi-linear Elliptic PDE {-nabla \\cdot ( nabla{u}/ sqrt{1-| nabla{u} |^2}) = 4 π sum_k a_k δ_{s_k}} in Physics and Geometry

    NASA Astrophysics Data System (ADS)

    Kiessling, Michael K.-H.

    2012-09-01

    It is shown that for each finite number N of Dirac measures {δ_{s_n}} supported at points {s_n in {R}^3} with given amplitudes {a_n in {R} backslash\\{0\\}} there exists a unique real-valued function {u in C^{0, 1}({R}^3)}, vanishing at infinity, which distributionally solves the quasi-linear elliptic partial differential equation of divergence form {-nabla \\cdot ( nabla{u}/ sqrt{1-| nabla{u} |^2}) = 4 π sum_{n=1}^N a_n δ_{s_n}}. Moreover, {u in C^{ω}({R}^3backslash \\{s_n\\}_{n=1}^N)}. The result can be interpreted in at least two ways: (a) for any number N of point charges of arbitrary magnitude and sign at prescribed locations s n in three-dimensional Euclidean space there exists a unique electrostatic field which satisfies the Maxwell-Born-Infeld field equations smoothly away from the point charges and vanishes as | s| → ∞; (b) for any number N of integral mean curvatures assigned to locations {s_n in {R}^3 subset{R}^{1, 3}} there exists a unique asymptotically flat, almost everywhere space-like maximal slice with point defects of Minkowski spacetime {{R}^{1, 3}}, having lightcone singularities over the s n but being smooth otherwise, and whose height function vanishes as | s| → ∞. No struts between the point singularities ever occur.

  19. The crystal structure of a heptameric archaeal Sm protein: Implications for the eukaryotic snRNP core.

    PubMed

    Mura, C; Cascio, D; Sawaya, M R; Eisenberg, D S

    2001-05-01

    Sm proteins form the core of small nuclear ribonucleoprotein particles (snRNPs), making them key components of several mRNA-processing assemblies, including the spliceosome. We report the 1.75-A crystal structure of SmAP, an Sm-like archaeal protein that forms a heptameric ring perforated by a cationic pore. In addition to providing direct evidence for such an assembly in eukaryotic snRNPs, this structure (i) shows that SmAP homodimers are structurally similar to human Sm heterodimers, (ii) supports a gene duplication model of Sm protein evolution, and (iii) offers a model of SmAP bound to single-stranded RNA (ssRNA) that explains Sm binding-site specificity. The pronounced electrostatic asymmetry of the SmAP surface imparts directionality to putative SmAP-RNA interactions.

  20. Purification of Drosophila snRNPs and characterization of two populations of functional U1 particles.

    PubMed Central

    Labourier, E; Rio, D C

    2001-01-01

    U1 snRNP is required at an early stage during assembly of the spliceosome, the dynamic ribonucleoprotein (RNP) complex that performs nuclear pre-mRNA splicing. Here, we report the purification of U1 snRNP particles from Drosophila nuclear extracts and the characterization of their biochemical properties, polypeptide contents, and splicing activities. On the basis of their antigenicity, apparent molecular weight, and by peptide sequencing, the Drosophila 70K, SNF, B, U1-C, D1, D2, D3, E, F, and G proteins are shown to be integral components of these particles. Sequence database searches revealed that both the U1-specific and the Sm proteins are extensively conserved between human and Drosophila snRNPs. Furthermore, both species possess a conserved intrinsic U1-associated kinase activity with identical substrate specificity in vitro. Finally, our results demonstrate that a second type of functional U1 particle, completely lacking the U1/U2-specific protein SNF and the associated protein kinase activity, can be isolated from cultured Kc cell or Canton S embryonic nuclear extracts. This work describes the first characterization of a purified Drosophila snRNP particle and reinforces the view that their activity and composition, with the exception of the atypical bifunctional U1-A/U2-B" SNF protein, are highly conserved in metazoans. PMID:11333025

  1. Determination of film processing specifications for the Apollo 17 S-209 lunar sounder experiment

    NASA Technical Reports Server (NTRS)

    Weinstein, M. S.

    1972-01-01

    The lunar sounder is described as a radar system operating at carrier frequencies of 5, 15, and 150 MHz. The radar echoes are recorded onto Kodak type S0-394 film through the use of an optical recorder utilizing a cathode ray tube as the exposing device. A processing configuration is determined with regard to linearity, dynamic range, and noise.

  2. The intronic splicing code: multiple factors involved in ATM pseudoexon definition.

    PubMed

    Dhir, Ashish; Buratti, Emanuele; van Santen, Maria A; Lührmann, Reinhard; Baralle, Francisco E

    2010-02-17

    Abundance of pseudo splice sites in introns can potentially give rise to innumerable pseudoexons, outnumbering the real ones. Nonetheless, these are efficiently ignored by the splicing machinery, a process yet to be understood completely. Although numerous 5' splice site-like sequences functioning as splicing silencers have been found to be enriched in predicted human pseudoexons, the lack of active pseudoexons pose a fundamental challenge to how these U1snRNP-binding sites function in splicing inhibition. Here, we address this issue by focusing on a previously described pathological ATM pseudoexon whose inhibition is mediated by U1snRNP binding at intronic splicing processing element (ISPE), composed of a consensus donor splice site. Spliceosomal complex assembly demonstrates inefficient A complex formation when ISPE is intact, implying U1snRNP-mediated unproductive U2snRNP recruitment. Furthermore, interaction of SF2/ASF with its motif seems to be dependent on RNA structure and U1snRNP interaction. Our results suggest a complex combinatorial interplay of RNA structure and trans-acting factors in determining the splicing outcome and contribute to understanding the intronic splicing code for the ATM pseudoexon.

  3. 322-R2U2 Engineering Assessment - August 2015

    SciTech Connect

    Abri, M.; Griffin, D.

    2015-08-13

    This Engineering Assessment and Certification of Integrity of retention tank system 322-R2 has been prepared for tank systems that store and neutralizes hazardous waste and have secondary containment. The regulations require that this assessment be completed periodically and certified by an independent, qualified, California-registered professional engineer. Abri Environmental Engineering performed an inspection of the 322-R2 Tank system at the Lawrence Livermore National Laboratory (LLNL) in Livermore, CA. Mr. William W. Moore, P.E., conducted this inspection on March 16, 2015. Mr. Moore is a California Registered Civil Engineer, with extensive experience in civil engineering, and hazardous waste management.

  4. U1/U2 crib groundwater biological treatment demonstration project

    SciTech Connect

    Koegler, S.S.; Brouns, T.M.; Heath, W.O.

    1989-11-01

    The primary objective of the biological treatment project is to develop and demonstrate a process for Hanford groundwater remediation. Biodenitrification using facultative anaerobic microorganisms is a promising technology for the simultaneous removal of nitrates and organics from contaminated aqueous streams. During FY 1988, a consortium of Hanford groundwater microorganisms was shown to degrade both nitrates and carbon tetrachloride (CC1{sub 4}). A pilot-scale treatment system was designed and constructed based on the results of laboratory-and-bench-scale testing. This report summarizes the results of biological groundwater treatment studies performed during FY 1989 at the pilot-scale. These tests were conducted using a simulated Hanford groundwater with a continuous stirred-tank bioreactor, and a fluidized-bed bioreactor that was added to the pilot-scale treatment system in FY 1989. The pilot-scale system demonstrated continuous degradation of nitrates and CC1{sub 4} in a simulated groundwater. 4 refs., 7 figs., 1 tab.

  5. Purification and characterization of native spliceosomes suitable for three-dimensional structural analysis.

    PubMed Central

    Jurica, Melissa S; Licklider, Lawrence J; Gygi, Steven R; Grigorieff, Nikolaus; Moore, Melissa J

    2002-01-01

    We describe characterization of spliceosomes affinity purified under native conditions. These spliceosomes consist largely of C complex containing splicing intermediates. After C complex assembly on an MS2 affinity-tagged pre-mRNA substrate containing a 3' splice site mutation, followed by RNase H digestion of earlier complexes, spliceosomes were purified by size exclusion and affinity selection. This protocol yielded 40S C complexes in sufficient quantities to visualize in negative stain by electron microscopy. Complexes purified in this way contain U2, U5, and U6 snRNAs, but very little U1 or U4 snRNA. Analysis by tandem mass spectrometry confirmed the presence of core snRNP proteins (SM and LSM), U2 and U5 snRNP-specific proteins, and the second step factors Prp16, Prp17, Slu7, and Prp22. In contrast, proteins specific to earlier splicing complexes, such as U2AF and U1 snRNP components, were not detected in C complex, but were present in similarly purified H complex. Images of these spliceosomes revealed single particles with dimensions of approximately 270 x 240 A that assort into well-defined classes. These images represent an important first step toward attaining a comprehensive three-dimensional understanding of pre-mRNA splicing. PMID:11991638

  6. Genome-wide RNAi screens in human brain tumor isolates reveal a novel viability requirement for PHF5A

    PubMed Central

    Hubert, Christopher G.; Bradley, Robert K.; Ding, Yu; Toledo, Chad M.; Herman, Jacob; Skutt-Kakaria, Kyobi; Girard, Emily J.; Davison, Jerry; Berndt, Jason; Corrin, Philip; Hardcastle, Justin; Basom, Ryan; Delrow, Jeffery J.; Webb, Thomas; Pollard, Steven M.; Lee, Jeongwu; Olson, James M.; Paddison, Patrick J.

    2013-01-01

    To identify key regulators of human brain tumor maintenance and initiation, we performed multiple genome-wide RNAi screens in patient-derived glioblastoma multiforme (GBM) stem cells (GSCs). These screens identified the plant homeodomain (PHD)-finger domain protein PHF5A as differentially required for GSC expansion, as compared with untransformed neural stem cells (NSCs) and fibroblasts. Given PHF5A's known involvement in facilitating interactions between the U2 snRNP complex and ATP-dependent helicases, we examined cancer-specific roles in RNA splicing. We found that in GSCs, but not untransformed controls, PHF5A facilitates recognition of exons with unusual C-rich 3′ splice sites in thousands of essential genes. PHF5A knockdown in GSCs, but not untransformed NSCs, astrocytes, or fibroblasts, inhibited splicing of these genes, leading to cell cycle arrest and loss of viability. Notably, pharmacologic inhibition of U2 snRNP activity phenocopied PHF5A knockdown in GSCs and also in NSCs or fibroblasts overexpressing MYC. Furthermore, PHF5A inhibition compromised GSC tumor formation in vivo and inhibited growth of established GBM patient-derived xenograft tumors. Our results demonstrate a novel viability requirement for PHF5A to maintain proper exon recognition in brain tumor-initiating cells and may provide new inroads for novel anti-GBM therapeutic strategies. PMID:23651857

  7. Genome-wide RNAi screens in human brain tumor isolates reveal a novel viability requirement for PHF5A.

    PubMed

    Hubert, Christopher G; Bradley, Robert K; Ding, Yu; Toledo, Chad M; Herman, Jacob; Skutt-Kakaria, Kyobi; Girard, Emily J; Davison, Jerry; Berndt, Jason; Corrin, Philip; Hardcastle, Justin; Basom, Ryan; Delrow, Jeffery J; Webb, Thomas; Pollard, Steven M; Lee, Jeongwu; Olson, James M; Paddison, Patrick J

    2013-05-01

    To identify key regulators of human brain tumor maintenance and initiation, we performed multiple genome-wide RNAi screens in patient-derived glioblastoma multiforme (GBM) stem cells (GSCs). These screens identified the plant homeodomain (PHD)-finger domain protein PHF5A as differentially required for GSC expansion, as compared with untransformed neural stem cells (NSCs) and fibroblasts. Given PHF5A's known involvement in facilitating interactions between the U2 snRNP complex and ATP-dependent helicases, we examined cancer-specific roles in RNA splicing. We found that in GSCs, but not untransformed controls, PHF5A facilitates recognition of exons with unusual C-rich 3' splice sites in thousands of essential genes. PHF5A knockdown in GSCs, but not untransformed NSCs, astrocytes, or fibroblasts, inhibited splicing of these genes, leading to cell cycle arrest and loss of viability. Notably, pharmacologic inhibition of U2 snRNP activity phenocopied PHF5A knockdown in GSCs and also in NSCs or fibroblasts overexpressing MYC. Furthermore, PHF5A inhibition compromised GSC tumor formation in vivo and inhibited growth of established GBM patient-derived xenograft tumors. Our results demonstrate a novel viability requirement for PHF5A to maintain proper exon recognition in brain tumor-initiating cells and may provide new inroads for novel anti-GBM therapeutic strategies. PMID:23651857

  8. Spliceosome assembly in the absence of stable U4/U6 RNA pairing

    PubMed Central

    Burke, Jordan E.; Butcher, Samuel E.; Brow, David A.

    2015-01-01

    The cycle of spliceosome assembly, intron excision, and spliceosome disassembly involves large-scale structural rearrangements of U6 snRNA that are functionally important. U6 enters the splicing pathway bound to the Prp24 protein, which chaperones annealing of U6 to U4 RNA to form a U4/U6 di-snRNP. During catalytic activation of the assembled spliceosome, U4 snRNP is released and U6 is paired to U2 snRNA. Here we show that point mutations in U4 and U6 that decrease U4/U6 base-pairing in vivo are lethal in combination. However, this synthetic phenotype is rescued by a mutation in U6 that alters a U6–Prp24 contact and stabilizes U2/U6. Remarkably, the resulting viable triple mutant strain lacks detectable U4/U6 base-pairing and U4/U6 di-snRNP. Instead, this strain accumulates free U4 snRNP, protein-free U6 RNA, and a novel complex containing U2/U6 di-snRNP. Further mutational analysis indicates that disruption of the U6–Prp24 interaction rather than stabilization of U2/U6 renders stable U4/U6 di-snRNP assembly nonessential. We propose that an essential function of U4/U6 pairing is to displace Prp24 from U6 RNA, and thus a destabilized U6–Prp24 complex renders stable U4/U6 pairing nonessential. PMID:25762536

  9. Achieving targeted and quantifiable alteration of mRNA splicing with Morpholino oligos

    SciTech Connect

    Morcos, Paul A. . E-mail: pmorcos@gene-tools.com

    2007-06-29

    This work represents the first guide for using steric-block antisense oligos as tools for effective and targeted modification of RNA splicing. Comparison of several steric-block oligo types shows the properties of Morpholinos provide significant advantages over other potential splice-blocking oligos. The procedures and complications of designing effective splice-blocking Morpholino oligos are described. The design process requires complete pre-mRNA sequence for defining suitable targets, which usually generate specific predictable messengers. To validate the targeting procedure, the level and nature of transcript alteration is characterized by RT-PCR analysis of splice modification in a {beta}-globin splice model system. An oligo-walking study reveals that while U1 and U2 small nuclear RiboNucleoProtein (snRNP) binding sites are the most effective targets for blocking splicing, inclusion of these sites is not required to achieve effective splice modifications. The most effective targeting strategy employs simultaneously blocking snRNP binding sites and splice-junctions. The work presented here continues to be the basis for most of the successful Morpholino oligos designed for the worldwide research community to block RNA splicing.

  10. The alternative splicing program of differentiated smooth muscle cells involves concerted non-productive splicing of post-transcriptional regulators

    PubMed Central

    Llorian, Miriam; Gooding, Clare; Bellora, Nicolas; Hallegger, Martina; Buckroyd, Adrian; Wang, Xiao; Rajgor, Dipen; Kayikci, Melis; Feltham, Jack; Ule, Jernej; Eyras, Eduardo; Smith, Christopher W.J.

    2016-01-01

    Alternative splicing (AS) is a key component of gene expression programs that drive cellular differentiation. Smooth muscle cells (SMCs) are important in the function of a number of physiological systems; however, investigation of SMC AS has been restricted to a handful of events. We profiled transcriptome changes in mouse de-differentiating SMCs and observed changes in hundreds of AS events. Exons included in differentiated cells were characterized by particularly weak splice sites and by upstream binding sites for Polypyrimidine Tract Binding protein (PTBP1). Consistent with this, knockdown experiments showed that that PTBP1 represses many smooth muscle specific exons. We also observed coordinated splicing changes predicted to downregulate the expression of core components of U1 and U2 snRNPs, splicing regulators and other post-transcriptional factors in differentiated cells. The levels of cognate proteins were lower or similar in differentiated compared to undifferentiated cells. However, levels of snRNAs did not follow the expression of splicing proteins, and in the case of U1 snRNP we saw reciprocal changes in the levels of U1 snRNA and U1 snRNP proteins. Our results suggest that the AS program in differentiated SMCs is orchestrated by the combined influence of auxiliary RNA binding proteins, such as PTBP1, along with altered activity and stoichiometry of the core splicing machinery. PMID:27317697

  11. The Translation Initiation Factor eIF4E Regulates the Sex-Specific Expression of the Master Switch Gene Sxl in Drosophila melanogaster

    PubMed Central

    Graham, Patricia L.; Yanowitz, Judith L.; Penn, Jill K. M.; Deshpande, Girish; Schedl, Paul

    2011-01-01

    In female fruit flies, Sex-lethal (Sxl) turns off the X chromosome dosage compensation system by a mechanism involving a combination of alternative splicing and translational repression of the male specific lethal-2 (msl-2) mRNA. A genetic screen identified the translation initiation factor eif4e as a gene that acts together with Sxl to repress expression of the Msl-2 protein. However, eif4e is not required for Sxl mediated repression of msl-2 mRNA translation. Instead, eif4e functions as a co-factor in Sxl-dependent female-specific alternative splicing of msl-2 and also Sxl pre-mRNAs. Like other factors required for Sxl regulation of splicing, eif4e shows maternal-effect female-lethal interactions with Sxl. This female lethality can be enhanced by mutations in other co-factors that promote female-specific splicing and is caused by a failure to properly activate the Sxl-positive autoregulatory feedback loop in early embryos. In this feedback loop Sxl proteins promote their own synthesis by directing the female-specific alternative splicing of Sxl-Pm pre-mRNAs. Analysis of pre-mRNA splicing when eif4e activity is compromised demonstrates that Sxl-dependent female-specific splicing of both Sxl-Pm and msl-2 pre-mRNAs requires eif4e activity. Consistent with a direct involvement in Sxl-dependent alternative splicing, eIF4E is associated with unspliced Sxl-Pm pre-mRNAs and is found in complexes that contain early acting splicing factors—the U1/U2 snRNP protein Sans-fils (Snf), the U1 snRNP protein U1-70k, U2AF38, U2AF50, and the Wilms' Tumor 1 Associated Protein Fl(2)d—that have been directly implicated in Sxl splicing regulation. PMID:21829374

  12. Scaling properties for the first RE-like mixed valence examples in uranium compounds: U 2Ru 2Sn and U 2RuGa 8

    NASA Astrophysics Data System (ADS)

    Troć, Robert

    2006-05-01

    The present study was motivated by the scaling characterization of the first example of mixed valence (MV) RE-like behaviour found recently among intermetallic ternary uranium compounds. The χ( T) function for both title compounds has been fitted to the interconfigurational fluctuation (ICF) model of Sales and Wohlleben in order to determine the characteristic fluctuation temperatures Tsf and interconfigurational excitation energies Eex. A good scaling, with similar values of Tsf like from those derived from the ICF model, has been achieved for both these ternaries by plotting Tχ( T)/ C against the reduced T/ Tsf . Moreover, this scaling follows almost exactly those found earlier in a number of MV- RE compounds.

  13. Functional genomic screening reveals splicing of the EWS-FLI1 fusion transcript as a vulnerability in Ewing sarcoma

    PubMed Central

    Grohar, Patrick J.; Kim, Suntae; Rangel Rivera, Guillermo O.; Sen, Nirmalya; Haddock, Sara; Harlow, Matt L.; Maloney, Nichole K.; Zhu, Jack; O’Neill, Maura; Jones, Tamara L.; Huppi, Konrad; Grandin, Magdalena; Gehlhaus, Kristen; Klumpp-Thomas, Carleen A.; Buehler, Eugen; Helman, Lee J.; Martin, Scott E.; Caplen, Natasha J.

    2016-01-01

    Summary Ewing sarcoma cells depend on the EWS-FLI1 fusion transcription factor for cell survival. Using an assay of EWS-FLI1 activity and genome-wide RNAi screening, we have identified proteins required for the processing of the EWS-FLI1 pre-mRNA. We show Ewing sarcoma cells harboring a genomic breakpoint that retains exon 8 of EWSR1 require the RNA-binding protein HNRNPH1 to express in-frame EWS-FLI1. We also demonstrate the sensitivity of EWS-FLI1 fusion transcripts to the loss-of-function of the U2 snRNP component, SF3B1. Disrupted splicing of the EWS-FLI1 transcript alters EWS-FLI1 protein expression and EWS-FLI1 driven expression. Our results show that the processing of the EWS-FLI1 fusion RNA is a potentially targetable vulnerability in Ewing sarcoma cells. PMID:26776507

  14. Functional Genomic Screening Reveals Splicing of the EWS-FLI1 Fusion Transcript as a Vulnerability in Ewing Sarcoma.

    PubMed

    Grohar, Patrick J; Kim, Suntae; Rangel Rivera, Guillermo O; Sen, Nirmalya; Haddock, Sara; Harlow, Matt L; Maloney, Nichole K; Zhu, Jack; O'Neill, Maura; Jones, Tamara L; Huppi, Konrad; Grandin, Magdalena; Gehlhaus, Kristen; Klumpp-Thomas, Carleen A; Buehler, Eugen; Helman, Lee J; Martin, Scott E; Caplen, Natasha J

    2016-01-26

    Ewing sarcoma cells depend on the EWS-FLI1 fusion transcription factor for cell survival. Using an assay of EWS-FLI1 activity and genome-wide RNAi screening, we have identified proteins required for the processing of the EWS-FLI1 pre-mRNA. We show that Ewing sarcoma cells harboring a genomic breakpoint that retains exon 8 of EWSR1 require the RNA-binding protein HNRNPH1 to express in-frame EWS-FLI1. We also demonstrate the sensitivity of EWS-FLI1 fusion transcripts to the loss of function of the U2 snRNP component, SF3B1. Disrupted splicing of the EWS-FLI1 transcript alters EWS-FLI1 protein expression and EWS-FLI1-driven expression. Our results show that the processing of the EWS-FLI1 fusion RNA is a potentially targetable vulnerability in Ewing sarcoma cells.

  15. Structure–function analysis and genetic interactions of the SmG, SmE, and SmF subunits of the yeast Sm protein ring

    PubMed Central

    Schwer, Beate; Kruchten, Joshua; Shuman, Stewart

    2016-01-01

    A seven-subunit Sm protein ring forms a core scaffold of the U1, U2, U4, and U5 snRNPs that direct pre-mRNA splicing. Using human snRNP structures to guide mutagenesis in Saccharomyces cerevisiae, we gained new insights into structure–function relationships of the SmG, SmE, and SmF subunits. An alanine scan of 19 conserved amino acids of these three proteins, comprising the Sm RNA binding sites or inter-subunit interfaces, revealed that, with the exception of Arg74 in SmF, none are essential for yeast growth. Yet, for SmG, SmE, and SmF, as for many components of the yeast spliceosome, the effects of perturbing protein–RNA and protein–protein interactions are masked by built-in functional redundancies of the splicing machine. For example, tests for genetic interactions with non-Sm splicing factors showed that many benign mutations of SmG, SmE, and SmF (and of SmB and SmD3) were synthetically lethal with null alleles of U2 snRNP subunits Lea1 and Msl1. Tests of pairwise combinations of SmG, SmE, SmF, SmB, and SmD3 alleles highlighted the inherent redundancies within the Sm ring, whereby simultaneous mutations of the RNA binding sites of any two of the Sm subunits are lethal. Our results suggest that six intact RNA binding sites in the Sm ring suffice for function but five sites may not. PMID:27417296

  16. Structure-function analysis and genetic interactions of the SmG, SmE, and SmF subunits of the yeast Sm protein ring.

    PubMed

    Schwer, Beate; Kruchten, Joshua; Shuman, Stewart

    2016-09-01

    A seven-subunit Sm protein ring forms a core scaffold of the U1, U2, U4, and U5 snRNPs that direct pre-mRNA splicing. Using human snRNP structures to guide mutagenesis in Saccharomyces cerevisiae, we gained new insights into structure-function relationships of the SmG, SmE, and SmF subunits. An alanine scan of 19 conserved amino acids of these three proteins, comprising the Sm RNA binding sites or inter-subunit interfaces, revealed that, with the exception of Arg74 in SmF, none are essential for yeast growth. Yet, for SmG, SmE, and SmF, as for many components of the yeast spliceosome, the effects of perturbing protein-RNA and protein-protein interactions are masked by built-in functional redundancies of the splicing machine. For example, tests for genetic interactions with non-Sm splicing factors showed that many benign mutations of SmG, SmE, and SmF (and of SmB and SmD3) were synthetically lethal with null alleles of U2 snRNP subunits Lea1 and Msl1. Tests of pairwise combinations of SmG, SmE, SmF, SmB, and SmD3 alleles highlighted the inherent redundancies within the Sm ring, whereby simultaneous mutations of the RNA binding sites of any two of the Sm subunits are lethal. Our results suggest that six intact RNA binding sites in the Sm ring suffice for function but five sites may not.

  17. Interactions of SR45, an SR-like protein, with spliceosomal proteins and an intronic sequence: insights into regulated splicing.

    PubMed

    Day, Irene S; Golovkin, Maxim; Palusa, Saiprasad G; Link, Alicia; Ali, Gul S; Thomas, Julie; Richardson, Dale N; Reddy, Anireddy S N

    2012-09-01

    SR45 is a serine/arginine-rich (SR)-like protein with two arginine/serine-rich (RS) domains. We have previously shown that SR45 regulates alternative splicing (AS) by differential selection of 5' and 3' splice sites. However, it is unknown how SR45 regulates AS. To gain mechanistic insights into the roles of SR45 in splicing, we screened a yeast two-hybrid library with SR45. This screening resulted in the isolation of two spliceosomal proteins, U1-70K and U2AF(35) b that are known to function in 5' and 3' splice site selection, respectively. This screen not only confirmed our prior observation that U1-70K and SR45 interact, but also helped to identify an additional interacting partner (U2AF(35) ). In vitro and in vivo analyses revealed an interaction of SR45 with both paralogs of U2AF(35) . Furthermore, we show that the RS1 and RS2 domains of SR45, and not the RNA recognition motif (RRM) domain, associate independently with both U2AF(35) proteins. Interaction studies among U2AF(35) paralogs and between U2AF(35) and U1-70K revealed that U2AF(35) can form homo- or heterodimers and that U2AF(35) proteins can associate with U1-70K. Using RNA probes from SR30 intron 10, whose splicing is altered in the sr45 mutant, we show that SR45 and U2AF(35) b bind to different parts of the intron, with a binding site for SR45 in the 5' region and two binding regions, each ending with a known 3' splice site, for U2AF(35) b. These results suggest that SR45 recruits U1snRNP and U2AF to 5' and 3' splice sites, respectively, by interacting with pre-mRNA, U1-70K and U2AF(35) and modulates AS. PMID:22563826

  18. Materials Data on U2Ga8Ru (SG:123) by Materials Project

    DOE Data Explorer

    Kristin Persson

    2016-02-10

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  19. Critical experiments on low-enriched uranium oxide systems with H/U = 2. 03

    SciTech Connect

    Rothe, R E; Goebel, G R

    1982-02-01

    Seven critical experiments were performed on a horizontal split table machine using 4.48% enriched /sup 235/U uranium oxide (U/sub 3/O/sub 8/). The oxide was compacted to a density of 4.68 g/cm/sup 3/ and placed in 152-mm cubical aluminum cans. Water was added to achieve an H/U atomic ratio of 2.03. Various arrays of oxide cans were distributed on each half of the split table and the separation between halves reduced until criticality occurred. The critical table separation varied from 4.3 mm to 29.3 mm. These experiments were performed in both plastic and concrete reflectors. The first five experiments required the addition of a high-enriched (approx. 93% /sup 235/U) metal driver to achieve criticality. Critical uranium driver masses ranged from 2.765 kg to 13.730 kg for 5 x 5 x 5 arrays of uranium oxide cans. In all five cases, the center can of the array was deleted to accommodate the driver. The uranium oxide mass was 1859.6 kg. Two additional experiments in the plastic reflector contained either 9.3-mm- or 24.3-mm-thick plastic moderator material between the oxide cans. These latter experiments did not require a driver to achieve criticality; and the uranium oxide mass was 723.9 kg for the configuration having the thinner interstitial moderator and 452.4 kg for the other.

  20. Materials Data on U2S3 (SG:62) by Materials Project

    SciTech Connect

    Kristin Persson

    2014-11-02

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  1. Materials Data on U2Bi2O7 (SG:51) by Materials Project

    DOE Data Explorer

    Kristin Persson

    2016-02-10

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  2. Materials Data on Rb6U2W4O21 (SG:2) by Materials Project

    SciTech Connect

    Kristin Persson

    2014-07-09

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  3. Materials Data on U2CuP2(HO)24 (SG:126) by Materials Project

    SciTech Connect

    Kristin Persson

    2014-07-09

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  4. Materials Data on U2CuAs2(HO)24 (SG:126) by Materials Project

    SciTech Connect

    Kristin Persson

    2014-07-09

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  5. Materials Data on K2U2O7 (SG:2) by Materials Project

    SciTech Connect

    Kristin Persson

    2014-07-09

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  6. Materials Data on Rb2U2O7 (SG:2) by Materials Project

    SciTech Connect

    Kristin Persson

    2014-07-09

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  7. Materials Data on U2F9 (SG:217) by Materials Project

    DOE Data Explorer

    Kristin Persson

    2016-02-10

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  8. Magnetism and superconductivity in U2PtxRh1-xC2

    DOE PAGES

    Wakeham, N.; Ni, Ni; Bauer, E. D.; Thompson, J. D.; Tegtmeier, E.; Ronning, F.

    2015-01-09

    We report the phase diagram of the doping series U₂PtxRh(1–x)C₂, studied through measurements of resistivity, specific heat, and magnetic susceptibility. The Néel temperature of U₂Rh₂C₂ of ~ 22 K is suppressed with increasing Pt content, reaching zero temperature close to x = 0.7, where we observed signatures of increased quantum fluctuations. Additionally, evidence is presented that the antiferromagnetic state undergoes a spin-reorientation transition upon application of an applied magnetic field. This transition shows non-monotonic behavior as a function of x, peaking at around x = 0.3. Superconductivity is observed for x ≥ 0.9, with Tc increasing with increasing x. Themore » reduction in Tc and increase in residual resistivity with decreasing Pt content is inconsistent with the extension of the Abrikosov-Gor'kov theory to unconventional superconductivity.« less

  9. The Z"k^(^s^u^(^2^)^,^3^/^2^) parafermions [rapid communication

    NASA Astrophysics Data System (ADS)

    Jacob, P.; Mathieu, P.

    2005-10-01

    We introduce a novel parafermionic theory for which the conformal dimension of the basic parafermion is 3/2 (1 -1/k), with k even. The structure constants and the central charges are obtained from mode-type associativity calculations. The spectrum of the completely reducible representations is also determined. The primary fields turns out to be labeled by two positive integers instead of a single one for the usual parafermionic models. The simplest singular vectors are also displayed. It is argued that these models are equivalent to the non-unitary minimal Wk (k + 1 , k + 3) models. More generally, we expect all Wk (k + 1 , k + 2 β) models to be identified with generalized parafermionic models whose lowest-dimensional parafermion has dimension β (1 - 1 / k).

  10. 46 CFR 54.01-5 - Scope (modifies U-1 and U-2).

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... Other: (a) Fired thermal fluid heaters: All 52 63 (b) Unfired steam boiler: More than 206 kPa (30 psig... requirements, section VIII of the ASME Boiler and Pressure Vessel Code (incorporated by reference, see 46 CFR... section VIII of the ASME Boiler and Pressure Vessel Code, and numbered (1), (2), etc. 2 See 46 CFR...

  11. 46 CFR 54.01-5 - Scope (modifies U-1 and U-2).

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... Other: (a) Fired thermal fluid heaters: All 52 63 (b) Unfired steam boiler: More than 206 kPa (30 psig... requirements, section VIII of the ASME Boiler and Pressure Vessel Code (incorporated by reference, see 46 CFR... section VIII of the ASME Boiler and Pressure Vessel Code, and numbered (1), (2), etc. 2 See 46 CFR...

  12. 46 CFR 54.01-5 - Scope (modifies U-1 and U-2).

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... Other: (a) Fired thermal fluid heaters: All 52 63 (b) Unfired steam boiler: More than 206 kPa (30 psig... requirements, section VIII of the ASME Boiler and Pressure Vessel Code (incorporated by reference, see 46 CFR... section VIII of the ASME Boiler and Pressure Vessel Code, and numbered (1), (2), etc. 2 See 46 CFR...

  13. 46 CFR 54.01-5 - Scope (modifies U-1 and U-2).

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... Other: (a) Fired thermal fluid heaters: All 52 63 (b) Unfired steam boiler: More than 206 kPa (30 psig... requirements, section VIII of the ASME Boiler and Pressure Vessel Code (incorporated by reference, see 46 CFR... section VIII of the ASME Boiler and Pressure Vessel Code, and numbered (1), (2), etc. 2 See 46 CFR...

  14. 46 CFR 54.01-5 - Scope (modifies U-1 and U-2).

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... Other: (a) Fired thermal fluid heaters: All 52 63 (b) Unfired steam boiler: More than 206 kPa (30 psig... requirements, section VIII of the ASME Boiler and Pressure Vessel Code (incorporated by reference, see 46 CFR... section VIII of the ASME Boiler and Pressure Vessel Code, and numbered (1), (2), etc. 2 See 46 CFR...

  15. Midinfrared beamline at the National Synchrotron Light Source port U2B

    SciTech Connect

    Carr, G.L. ); Hanfland, M. ); Williams, G.P. )

    1995-02-01

    A new infrared beamline has been developed on a conventional dipole bending magnet port of the vacuum ultraviolet ring at the National Synchrotron Light Source. The port provides approximately 12 mrad horizontal and 8 mrad vertical aperture, which limits the useful spectral range to wavelengths less than 20 [mu]m. Though the total flux across the midinfrared is less than that from a globar source, the calculated brightness is at least two orders of magnitude greater. Also, the synchrotron source delivers light in subnanosecond pulses. The developing experimental programs include studies of hydrogen and other materials at extremely high pressures, and time-resolved studies of infrared sensor materials. The measurement results presented here, characterizing the actual brightness advantage and spectroscopic performance, demonstrate the synchrotron's remarkable advantage for microspectroscopic studies.

  16. Altered mRNA Splicing, Chondrocyte Gene Expression and Abnormal Skeletal Development due to SF3B4 Mutations in Rodriguez Acrofacial Dysostosis

    PubMed Central

    Nevarez, Lisette; Pogue, Robert; Krakow, Deborah; Cohn, Daniel H.

    2016-01-01

    The acrofacial dysostoses (AFD) are a genetically heterogeneous group of inherited disorders with craniofacial and limb abnormalities. Rodriguez syndrome is a severe, usually perinatal lethal AFD, characterized by severe retrognathia, oligodactyly and lower limb abnormalities. Rodriguez syndrome has been proposed to be a severe form of Nager syndrome, a non-lethal AFD that results from mutations in SF3B4, a component of the U2 small nuclear ribonucleoprotein particle (U2 snRNP). Furthermore, a case with a phenotype intermediate between Rodriguez and Nager syndromes has been shown to have an SF3B4 mutation. We identified heterozygosity for SF3B4 mutations in Rodriguez syndrome, confirming that the phenotype is a dominant disorder that is allelic with Nager syndrome. The mutations led to reduced SF3B4 synthesis and defects in mRNA splicing, primarily exon skipping. The mutations also led to reduced expression in growth plate chondrocytes of target genes, including the DLX5, DLX6, SOX9, and SOX6 transcription factor genes, which are known to be important for skeletal development. These data provide mechanistic insight toward understanding how SF3B4 mutations lead to the skeletal abnormalities observed in the acrofacial dysostoses. PMID:27622494

  17. A targeted oligonucleotide enhancer of SMN2 exon 7 splicing forms competing quadruplex and protein complexes in functional conditions.

    PubMed

    Smith, Lindsay D; Dickinson, Rachel L; Lucas, Christian M; Cousins, Alex; Malygin, Alexey A; Weldon, Carika; Perrett, Andrew J; Bottrill, Andrew R; Searle, Mark S; Burley, Glenn A; Eperon, Ian C

    2014-10-01

    The use of oligonucleotides to activate the splicing of selected exons is limited by a poor understanding of the mechanisms affected. A targeted bifunctional oligonucleotide enhancer of splicing (TOES) anneals to SMN2 exon 7 and carries an exonic splicing enhancer (ESE) sequence. We show that it stimulates splicing specifically of intron 6 in the presence of repressing sequences in intron 7. Complementarity to the 5' end of exon 7 increases U2AF65 binding, but the ESE sequence is required for efficient recruitment of U2 snRNP. The ESE forms at least three coexisting discrete states: a quadruplex, a complex containing only hnRNP F/H, and a complex enriched in the activator SRSF1. Neither hnRNP H nor quadruplex formation contributes to ESE activity. The results suggest that splicing limited by weak signals can be rescued by rapid exchange of TOES oligonucleotides in various complexes and raise the possibility that SR proteins associate transiently with ESEs. PMID:25263560

  18. αCP binding to a cytosine-rich subset of polypyrimidine tracts drives a novel pathway of cassette exon splicing in the mammalian transcriptome

    PubMed Central

    Ji, Xinjun; Park, Juw Won; Bahrami-Samani, Emad; Lin, Lan; Duncan-Lewis, Christopher; Pherribo, Gordon; Xing, Yi; Liebhaber, Stephen A.

    2016-01-01

    Alternative splicing (AS) is a robust generator of mammalian transcriptome complexity. Splice site specification is controlled by interactions of cis-acting determinants on a transcript with specific RNA binding proteins. These interactions are frequently localized to the intronic U-rich polypyrimidine tracts (PPT) located 5′ to the majority of splice acceptor junctions. αCPs (also referred to as polyC-binding proteins (PCBPs) and hnRNPEs) comprise a subset of KH-domain proteins with high affinity and specificity for C-rich polypyrimidine motifs. Here, we demonstrate that αCPs promote the splicing of a defined subset of cassette exons via binding to a C-rich subset of polypyrimidine tracts located 5′ to the αCP-enhanced exonic segments. This enhancement of splice acceptor activity is linked to interactions of αCPs with the U2 snRNP complex and may be mediated by cooperative interactions with the canonical polypyrimidine tract binding protein, U2AF65. Analysis of αCP-targeted exons predicts a substantial impact on fundamental cell functions. These findings lead us to conclude that the αCPs play a direct and global role in modulating the splicing activity and inclusion of an array of cassette exons, thus driving a novel pathway of splice site regulation within the mammalian transcriptome. PMID:26896798

  19. Cloning of the cDNA for U1 small nuclear ribonucleoprotein particle 70K protein from Arabidopsis thaliana

    NASA Technical Reports Server (NTRS)

    Reddy, A. S.; Czernik, A. J.; An, G.; Poovaiah, B. W.

    1992-01-01

    We cloned and sequenced a plant cDNA that encodes U1 small nuclear ribonucleoprotein (snRNP) 70K protein. The plant U1 snRNP 70K protein cDNA is not full length and lacks the coding region for 68 amino acids in the amino-terminal region as compared to human U1 snRNP 70K protein. Comparison of the deduced amino acid sequence of the plant U1 snRNP 70K protein with the amino acid sequence of animal and yeast U1 snRNP 70K protein showed a high degree of homology. The plant U1 snRNP 70K protein is more closely related to the human counter part than to the yeast 70K protein. The carboxy-terminal half is less well conserved but, like the vertebrate 70K proteins, is rich in charged amino acids. Northern analysis with the RNA isolated from different parts of the plant indicates that the snRNP 70K gene is expressed in all of the parts tested. Southern blotting of genomic DNA using the cDNA indicates that the U1 snRNP 70K protein is coded by a single gene.

  20. Relationships between chlorophyll density and ocean radiance as measured by U2/OCS: Algorithms, examples and comparison

    NASA Technical Reports Server (NTRS)

    Kim, H. H.; Hart, W. D.

    1983-01-01

    An ocean atmosphere radiative transfer process computation method which is suitable for determining lower boundary ocean albedo and other radiation components from spectral measurements of upwelling radiance taken from a high altitude platform is described. The method was applied to a set of color scanner data taken from slope water of the South Atlantic Bight to determine the influence of cholorophyll-a pigments in the sea on the ratio of upwelling radiance to down welling irradiance as a function of wavelength. The resulting chlorophyll concentrations are compared with measurements made by ships stationed along the flight path.

  1. Direct containment heating experiments in Zion Nuclear Power Plant Geometry using prototypic core materials, the U2 test

    SciTech Connect

    Binder, J.L.; McUmber, L.M.; Spencer, B.W.

    1993-05-01

    A third Direct Containment Heating (DCH) experiments has been completed which utilizes prototypic core materials. The reactor material tests are a follow on to the Integral Effects Testing (IET) DCH program. The IET series of tests primarily addressed the effect of scale on DCH phenomena. This was accomplished by completing a series of counterpart tests in 1/40 and 1/10th linear scale DCH facilities at Argonne National Laboratory (ANL) and Sandia National Laboratories (SNL), respectively. The IET experiments modeled the Zion Nuclear Power Plant Geometry. The scale models included representations of the primary system volume, RPV lower head, cavity and instrument tunnel, and the lower containment structures. The experiments were steam driven at nominally 6.2 MPa. Iron-alumina thermite with chromium was used as a core melt simulant in the IET experiments. While the IET experiments at ANL and SNL provided useful data on the effect of scale on DCH phenomena, a significant question concerns the potential experiment distortions introduced by the use of non-prototypic iron/alumina thermite. Therefore, further testing with prototypic materials has been carried out at ANL. A prototypic core melt was produced for the experiment by first mixing powders of uranium, zirconium, iron oxide (Fe{sub 2}O{sub 3}), and chromium trioxide (CrO{sub 3}). When ignited the powders react exothermically to produce a molten mixture. The amounts of each powder were selected to produce the anticipated composition for a core melt following a station blackout: 57.8 mass% UO{sub 2} 10.5 mass% ZrO{sub 2} 14.3 mass% Fe, 13.7 mass% Zr, and 3.7 mass% Cr. Development tests measured the initial melt temperature to be in the range of 2600 - 2700 K. The total thermal specific energy content of the melt at 2700 K is 1.2 MJ/kg compared to 2.25 MJ/kg for the iron-alumina simulant at its measured initial temperature of 2500 K.

  2. Materials Data on U2Si(H2O5)2 (SG:70) by Materials Project

    SciTech Connect

    Kristin Persson

    2014-11-02

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  3. Materials Data on CsU2H3(C2O7)2 (SG:11) by Materials Project

    SciTech Connect

    Kristin Persson

    2014-07-09

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  4. Materials Data on RbU2H5C4O15 (SG:11) by Materials Project

    SciTech Connect

    Kristin Persson

    2014-10-02

    Computed materials data using density functional theory calculations. These calculations determine the electronic structure of bulk materials by solving approximations to the Schrodinger equation. For more information, see https://materialsproject.org/docs/calculations

  5. Recognition of Trimethylated Histone H3 Lysine 4 Facilitates the Recruitment of Transcription Post-Initiation Factors and pre-mRNA Splicing

    PubMed Central

    Sims, Robert J.; Millhouse, Scott; Chen, Chi-Fu; Lewis, Brian A.; Erdjument-Bromage, Hediye; Tempst, Paul; Manley, James L.; Reinberg, Danny

    2007-01-01

    Tri-methylation of histone H3 on lysine 4 (H3K4me3) localizes near the 5′ region of genes and is tightly associated with active loci. Several proteins, such as CHD1, BPTF, JMJD2A, and the ING tumor suppressor family, directly recognize this lysine methyl mark. However, how H3K4me3 recognition participates in active transcription remains poorly characterized. Here we identify specific CHD1-interacting proteins via H3K4me3 affinity purification, including numerous factors mediating post-initiation events. Conventional biochemical purification revealed a stable complex between CHD1 and components of the spliceosome. Depletion of CHD1 in extracts dramatically reduced splicing efficiency in vitro, indicating a functional link between CHD1 and the spliceosome. Knockdown of CHD1 and H3K4me3 levels by siRNA reduced association of U2 snRNP components with chromatin, and more importantly, altered the efficiency of pre-mRNA splicing on active genes in vivo. These findings suggest that methylated H3K4 serves to facilitate the competency of pre-mRNA maturation through the bridging of spliceosomal components to H3K4me3 via CHD1. PMID:18042460

  6. A Broad Set of Chromatin Factors Influences Splicing

    PubMed Central

    Allemand, Eric; Myers, Michael P.; Garcia-Bernardo, Jose; Harel-Bellan, Annick; Krainer, Adrian R.; Muchardt, Christian

    2016-01-01

    Several studies propose an influence of chromatin on pre-mRNA splicing, but it is still unclear how widespread and how direct this phenomenon is. We find here that when assembled in vivo, the U2 snRNP co-purifies with a subset of chromatin-proteins, including histones and remodeling complexes like SWI/SNF. Yet, an unbiased RNAi screen revealed that the outcome of splicing is influenced by a much larger variety of chromatin factors not all associating with the spliceosome. The availability of this broad range of chromatin factors impacting splicing further unveiled their very context specific effect, resulting in either inclusion or skipping, depending on the exon under scrutiny. Finally, a direct assessment of the impact of chromatin on splicing using an in vitro co-transcriptional splicing assay with pre-mRNAs transcribed from a nucleosomal template, demonstrated that chromatin impacts nascent pre-mRNP in their competence for splicing. Altogether, our data show that numerous chromatin factors associated or not with the spliceosome can affect the outcome of splicing, possibly as a function of the local chromatin environment that by default interferes with the efficiency of splicing. PMID:27662573

  7. Structure and novel functional mechanism of Drosophila SNF in sex-lethal splicing.

    PubMed

    Hu, Jicheng; Cui, Gaofeng; Li, Congmin; Liu, Cong; Shang, Erchang; Lai, Luhua; Jin, Changwen; Wang, Jiwu; Xia, Bin

    2009-09-03

    Sans-fille (SNF) is the Drosophila homologue of mammalian general splicing factors U1A and U2B'', and it is essential in Drosophila sex determination. We found that, besides its ability to bind U1 snRNA, SNF can also bind polyuridine RNA tracts flanking the male-specific exon of the master switch gene Sex-lethal (Sxl) pre-mRNA specifically, similar to Sex-lethal protein (SXL). The polyuridine RNA binding enables SNF directly inhibit Sxl exon 3 splicing, as the dominant negative mutant SNF(1621) binds U1 snRNA but not polyuridine RNA. Unlike U1A, both RNA recognition motifs (RRMs) of SNF can recognize polyuridine RNA tracts independently, even though SNF and U1A share very high sequence identity and overall structure similarity. As SNF RRM1 tends to self-associate on the opposite side of the RNA binding surface, it is possible for SNF to bridge the formation of super-complexes between two introns flanking Sxl exon 3 or between a intron and U1 snRNP, which serves the molecular basis for SNF to directly regulate Sxl splicing. Taken together, a new functional model for SNF in Drosophila sex determination is proposed. The key of the new model is that SXL and SNF function similarly in promoting Sxl male-specific exon skipping with SNF being an auxiliary or backup to SXL, and it is the combined dose of SXL and SNF governs Drosophila sex determination.

  8. Genetic interaction mapping reveals a role for the SWI/SNF nucleosome remodeler in spliceosome activation in fission yeast.

    PubMed

    Patrick, Kristin L; Ryan, Colm J; Xu, Jiewei; Lipp, Jesse J; Nissen, Kelly E; Roguev, Assen; Shales, Michael; Krogan, Nevan J; Guthrie, Christine

    2015-03-01

    Although numerous regulatory connections between pre-mRNA splicing and chromatin have been demonstrated, the precise mechanisms by which chromatin factors influence spliceosome assembly and/or catalysis remain unclear. To probe the genetic network of pre-mRNA splicing in the fission yeast Schizosaccharomyces pombe, we constructed an epistatic mini-array profile (E-MAP) and discovered many new connections between chromatin and splicing. Notably, the nucleosome remodeler SWI/SNF had strong genetic interactions with components of the U2 snRNP SF3 complex. Overexpression of SF3 components in ΔSWI/SNF cells led to inefficient splicing of many fission yeast introns, predominantly those with non-consensus splice sites. Deletion of SWI/SNF decreased recruitment of the splicing ATPase Prp2, suggesting that SWI/SNF promotes co-transcriptional spliceosome assembly prior to first step catalysis. Importantly, defects in SWI/SNF as well as SF3 overexpression each altered nucleosome occupancy along intron-containing genes, illustrating that the chromatin landscape both affects--and is affected by--co-transcriptional splicing.

  9. All 17 S-locus F-box proteins of the S2 - and S3 -haplotypes of Petunia inflata are assembled into similar SCF complexes with a specific function in self-incompatibility.

    PubMed

    Li, Shu; Williams, Justin S; Sun, Penglin; Kao, Teh-Hui

    2016-09-01

    The collaborative non-self-recognition model for S-RNase-based self-incompatibility predicts that multiple S-locus F-box proteins (SLFs) produced by pollen of a given S-haplotype collectively mediate ubiquitination and degradation of all non-self S-RNases, but not self S-RNases, in the pollen tube, thereby resulting in cross-compatible pollination but self-incompatible pollination. We had previously used pollen extracts containing GFP-fused S2 -SLF1 (SLF1 with an S2 -haplotype) of Petunia inflata for co-immunoprecipitation (Co-IP) and mass spectrometry (MS), and identified PiCUL1-P (a pollen-specific Cullin1), PiSSK1 (a pollen-specific Skp1-like protein) and PiRBX1 (a conventional Rbx1) as components of the SCF(S) (2-) (SLF) (1) complex. Using pollen extracts containing PiSSK1:FLAG:GFP for Co-IP/MS, we identified two additional SLFs (SLF4 and SLF13) that were assembled into SCF(SLF) complexes. As 17 SLF genes (SLF1 to SLF17) have been identified in S2 and S3 pollen, here we examined whether all 17 SLFs are assembled into similar complexes and, if so, whether these complexes are unique to SLFs. We modified the previous Co-IP/MS procedure, including the addition of style extracts from four different S-genotypes to pollen extracts containing PiSSK1:FLAG:GFP, to perform four separate experiments. The results taken together show that all 17 SLFs and an SLF-like protein, SLFLike1 (encoded by an S-locus-linked gene), co-immunoprecipitated with PiSSK1:FLAG:GFP. Moreover, of the 179 other F-box proteins predicted by S2 and S3 pollen transcriptomes, only a pair with 94.9% identity and another pair with 99.7% identity co-immunoprecipitated with PiSSK1:FLAG:GFP. These results suggest that SCF(SLF) complexes have evolved specifically to function in self-incompatibility.

  10. SPF30 is an essential human splicing factor required for assembly of the U4/U5/U6 tri-small nuclear ribonucleoprotein into the spliceosome.

    PubMed

    Rappsilber, J; Ajuh, P; Lamond, A I; Mann, M

    2001-08-17

    Spliceosome assembly involves the sequential recruitment of small nuclear ribonucleoproteins (snRNPs) onto a pre-mRNA substrate. Although several non-snRNP proteins function during the binding of U1 and U2 snRNPs, little is known about the subsequent binding of the U4/U5/U6 tri-snRNP. A recent proteomic analysis of the human spliceosome identified SPF30 (Neubauer, G., King, A., Rappsilber, J., Calvio, C., Watson, M., Ajuh, P., Sleeman, J., Lamond, A., and Mann, M. (1998) Nat. Genet. 20, 46-50), a homolog of the survival of motor neurons (SMN) protein, as a spliceosome factor. We show here that SPF30 is a nuclear protein that associates with both U4/U5/U6 and U2 snRNP components. In the absence of SPF30, the preformed tri-snRNP fails to assemble into the spliceosome. Mass spectrometric analysis shows that a recombinant glutathione S-transferase-SPF30 fusion protein associates with complexes containing core Sm and U4/U5/U6 tri-snRNP proteins when added to HeLa nuclear extract, most strongly to U4/U6-90. The data indicate that SPF30 is an essential human splicing factor that may act to dock the U4/U5/U6 tri-snRNP to the A complex during spliceosome assembly or, alternatively, may act as a late assembly factor in both the tri-snRNP and the A-complex. PMID:11331295

  11. Cell cycle gene-specific control of transcription has a critical role in proliferation of primordial germ cells

    PubMed Central

    Okamura, Daiji; Maeda, Ikuma; Taniguchi, Hirofumi; Tokitake, Yuko; Ikeda, Makiko; Ozato, Keiko; Mise, Nathan; Abe, Kuniya; Noce, Toshiaki; Izpisua Belmonte, Juan Carlos; Matsui, Yasuhisa

    2012-01-01

    Transcription elongation is stimulated by positive transcription elongation factor b (P-TEFb), for which activity is repressed in the 7SK small nuclear ribonucleoprotein (7SK snRNP) complex. We show here a critical role of 7SK snRNP in growth control of primordial germ cells (PGCs). The expression of p15INK4b, a cyclin-dependent kinase inhibitor (CDKI) gene, in PGCs is selectively activated by P-TEFb and its recruiting molecule, Brd4, when the amount of active P-TEFb is increased due to reduction of the 7SK snRNP, and PGCs consequently undergo growth arrest. These results indicate that CDKI gene-specific control of transcription by 7SK snRNP plays a pivotal role in the maintenance of PGC proliferation. PMID:23154982

  12. U1 small nuclear ribonucleoproteins (snRNPs) aggregate in Alzheimer’s disease due to autosomal dominant genetic mutations and trisomy 21

    PubMed Central

    2014-01-01

    Background We recently identified U1 small nuclear ribonucleoprotein (snRNP) tangle-like aggregates and RNA splicing abnormalities in sporadic Alzheimer’s disease (AD). However little is known about snRNP biology in early onset AD due to autosomal dominant genetic mutations or trisomy 21 in Down syndrome. Therefore we investigated snRNP biochemical and pathologic features in these disorders. Findings We performed quantitative proteomics and immunohistochemistry in postmortem brain from genetic AD cases. Electron microscopy was used to characterize ultrastructural features of pathologic aggregates. U1-70k and other snRNPs were biochemically enriched in the insoluble fraction of human brain from subjects with presenilin 1 (PS1) mutations. Aggregates of U1 snRNP-immunoreactivity formed cytoplasmic tangle-like structures in cortex of AD subjects with PS1 and amyloid precursor protein (APP) mutations as well as trisomy 21. Ultrastructural analysis with electron microscopy in an APP mutation case demonstrated snRNP immunogold labeling of paired helical filaments (PHF). Conclusions These studies identify U1 snRNP pathologic changes in brain of early onset genetic forms of AD. Since dominant genetic mutations and trisomy 21 result in dysfunctional amyloid processing, the findings suggest that aberrant β-amyloid processing may influence U1 snRNP aggregate formation. PMID:24773620

  13. Pressure-induced electronic and magnetic phase transitions in a Mott insulator: Ti-doped C a3R u2O7 bilayer ruthenate

    NASA Astrophysics Data System (ADS)

    Zou, T.; Cao, H. B.; Liu, G. Q.; Peng, J.; Gottschalk, M.; Zhu, M.; Zhao, Y.; Leão, J. B.; Tian, W.; Mao, Z. Q.; Ke, X.

    2016-07-01

    We report the hydrostatic pressure-induced electronic and magnetic phase transitions in a Mott insulator, a bilayer ruthenate C a3(Ru0.97Ti0.03 ) 2O7 , via electronic transport and single crystal neutron diffraction measurements. The system undergoes an insulator-metal transition at a very small hydrostatic pressure ≈0.04 GPa, followed by a magnetic phase transition around 0.3 GPa, suggesting that the low energy charge fluctuation and magnetic ordering couple to the pressure separately in this compound. The a b initio calculations show that the suppressed Ru O6 flattening induced by the pressure reduces the orbital polarization and gives rise to an insulator-metal transition preceding the magnetic phase transition.

  14. 2,2,2-Trichloroethanol lengthens the circadian period of Bmal1-driven circadian bioluminescence rhythms in U2OS cells.

    PubMed

    Gao, Jie; Yang, Caixia; Ding, Hui; Zhang, Yanli; Xue, Jinhua; Cai, Yanning

    2015-07-01

    2,2,2-Trichloroethanol (TCOH) is responsible for the pharmacological actions of chloral hydrate (CH), and is a major metabolite of trichloroethylene. Human exposure to TCOH is known to be increasing. Recently, it was reported that TCOH causes a significant phase delay of Per2 expression in mouse liver when injected daily over the course of several days. However, it is not clear whether TCOH directly modulates the molecular clock. In the present study we used a cell-based assay system to test this possibility. We found that the daily oscillation period of Bmal1 was lengthened to 3 h following treatment with 1.5 mM TCOH, and increased to 5 h with 3 mM TCOH treatment. However, low concentrations of TCOH had no noticeable effects. The effect of TCOH on Per2 oscillation was marginal. Interestingly, serum from rats anesthetized with CH also modulated Bmal1 period, suggesting that exposure to anesthesia should be taken into consideration for circadian rhythm studies. In summary, our study reveals a direct regulation of TCOH on molecular clock. PMID:25956065

  15. Human STAGA complex is a chromatin-acetylating transcription coactivator that interacts with pre-mRNA splicing and DNA damage-binding factors in vivo.

    PubMed

    Martinez, E; Palhan, V B; Tjernberg, A; Lymar, E S; Gamper, A M; Kundu, T K; Chait, B T; Roeder, R G

    2001-10-01

    GCN5 is a histone acetyltransferase (HAT) originally identified in Saccharomyces cerevisiae and required for transcription of specific genes within chromatin as part of the SAGA (SPT-ADA-GCN5 acetylase) coactivator complex. Mammalian cells have two distinct GCN5 homologs (PCAF and GCN5L) that have been found in three different SAGA-like complexes (PCAF complex, TFTC [TATA-binding-protein-free TAF(II)-containing complex], and STAGA [SPT3-TAF(II)31-GCN5L acetylase]). The composition and roles of these mammalian HAT complexes are still poorly characterized. Here, we present the purification and characterization of the human STAGA complex. We show that STAGA contains homologs of most yeast SAGA components, including two novel human proteins with histone-like folds and sequence relationships to yeast SPT7 and ADA1. Furthermore, we demonstrate that STAGA has acetyl coenzyme A-dependent transcriptional coactivator functions from a chromatin-assembled template in vitro and associates in HeLa cells with spliceosome-associated protein 130 (SAP130) and DDB1, two structurally related proteins. SAP130 is a component of the splicing factor SF3b that associates with U2 snRNP and is recruited to prespliceosomal complexes. DDB1 (p127) is a UV-damaged-DNA-binding protein that is involved, as part of a complex with DDB2 (p48), in nucleotide excision repair and the hereditary disease xeroderma pigmentosum. Our results thus suggest cellular roles of STAGA in chromatin modification, transcription, and transcription-coupled processes through direct physical interactions with sequence-specific transcription activators and with components of the splicing and DNA repair machineries. PMID:11564863

  16. hnRNP A1 controls HIV-1 mRNA splicing through cooperative binding to intron and exon splicing silencers in the context of a conserved secondary structure.

    PubMed Central

    Damgaard, Christian Kroun; Tange, Thomas Ostergaard; Kjems, Jørgen

    2002-01-01

    The removal of the second intron in the HIV-1 rev/tat pre-mRNAs, which involves the joining of splice site SD4 to SA7, is inhibited by hnRNP A1 by a mechanism that requires the intronic splicing silencer (ISS) and the exon splicing silencer (ESS3). In this study, we have determined the RNA secondary structure and the hnRNP A1 binding sites within the 3' splice site region by phylogenetic comparison and chemical/enzymatic probing. A biochemical characterization of the RNA/protein complexes demonstrates that hnRNP A1 binds specifically to primarily three sites, the ISS, a novel UAG motif in the exon splicing enhancer (ESE) and the ESS3 element, which are all situated in experimentally supported stem loop structures. A mutational analysis of the ISS region revealed that the core hnRNP A1 binding site directly overlaps with a major branchpoint used in splicing to SA7, thereby providing a direct explanation for the inhibition of U2 snRNP association with the pre-mRNA by hnRNP A1. Binding of hnRNP A1 to the ISS core site is inhibited by RNA structure but strongly stimulated by the exonic silencer, ESS3. Moreover, the ISS also stimulate binding of hnRNP A1 to the exonic splicing regulators ESS3 and the ESE. Our results suggest a model where a network is formed between hnRNP A1 molecules situated at discrete sites in the intron and exon and that these interactions preclude the recognition of essential splicing signals including the branch point. PMID:12458794

  17. U1 small nuclear ribonucleoprotein particle-specific proteins interact with the first and second stem-loops of U1 RNA, with the A protein binding directly to the RNA independently of the 70K and Sm proteins.

    PubMed Central

    Patton, J R; Habets, W; van Venrooij, W J; Pederson, T

    1989-01-01

    The U1 small nuclear ribonucleoprotein particle (U1 snRNP), a cofactor in pre-mRNA splicing, contains three proteins, termed 70K, A, and C, that are not present in the other spliceosome-associated snRNPs. We studied the binding of the A and C proteins to U1 RNA, using a U1 snRNP reconstitution system and an antibody-induced nuclease protection technique. Antibodies that reacted with the A and C proteins induced nuclease protection of the first two stem-loops of U1 RNA in reconstituted U1 snRNP. Detailed analysis of the antibody-induced nuclease protection patterns indicated the existence of relatively long-range protein-protein interactions in the U1 snRNP, with the 5' end of U1 RNA and its associated specific proteins interacting with proteins bound to the Sm domain near the 3' end. UV cross-linking experiments in conjunction with an A-protein-specific antibody demonstrated that the A protein bound directly to the U1 RNA rather than assembling in the U1 snRNP exclusively via protein-protein interactions. This conclusion was supported by additional experiments revealing that the A protein could bind to U1 RNA in the absence of bound 70K and Sm core proteins. Images PMID:2529425

  18. Functional analysis of the sea urchin U7 small nuclear RNA

    SciTech Connect

    Gilmartin, G.M.; Schaufele, F.; Schaffner, G.; Birnstiel, M.L.

    1988-03-01

    U7 small nuclear RNA (snRNA) is an essential component of the RNA-processing machinery which generates the 3' end of mature histone mRNA in the sea urchin. The U7 small nuclear ribonucleoprotein particle (snRNP) is classified as a member of the Sm-type U snRNP family by virtue of its recognition by both anti-trimethylguanosine and anti-Sm antibodies. The authors analyzed the function-structure relationship of the U7 snRNP by mutagenesis experiments. These suggested that the U7 snRNP of the sea urchin is composed of three important domains. The fist domain encompasses the 5'-terminal sequence, up to about nucleotides 7, which are accessible to micrococcal nuclease, while the remainder of the RNA is highly protected and hence presumably bound by proteins. This region contains the sequence complementarities between the U7 snRNA and the histone pre-mRNA which have previously been shown to be required for 3' processing. Nucleotides 9 to 20 constitute a second domain which includes sequences for Sm protein binding. The complementarities between the U7 snRNA sequences in this region and the terminal palindrome fo the historne mRNA appear to be fortuitous and play only a secondary, if any, role in 3' processing. The third domain is composed of the terminal palindrome of U7 snRNA, the secondary structure of which must be maintained for the U7 snRNP to function, but its sequence can be drastically altered without any observable effect on snRNP assembly or 3' processing.

  19. (1'S,12'R,13'S,17'S)-15',15'-Dimethyl-1,2-dihydro-11',14',16',18'-tetra-oxa-7'-aza-spiro-[indole-3,8'-penta-cyclo-[10.6.0.0(2,9).0(3,7).0(13,17)]octa-deca-ne]-2,10'-dione.

    PubMed

    Sabari, V; Ponnusamy, R; Prasanna, R; Raghunathan, R; Aravindhan, S

    2013-04-01

    In the title compound, C22H24N2O6, the indole ring has a twist conformation and the tetra-hydro-2H-pyran-2-one ring a half-chair conformation. One of the pyrrolidine rings adopts an envelope conformation on the N atom, while the other has a twist conformation; the 'butterfly' angle between their mean planes is 62.98 (11)°. The dioxolane ring adopts a twist conformation and the tetra-hydro-furan ring has an envelope conformation on the C atom in the fused tetra-hydro-2H-pyran-2-one ring adjacent to the O atom of the tetra-hydro-furan ring. The 'butterfly' angle between the mean planes of these two five-membered rings is 69.14 (10)°. In the crystal, mol-ecules are linked by N-H⋯O hydrogen bonds, forming chains along the a axis.

  20. Crystallization and biochemical characterization of the human spliceosomal Aar2-Prp8(RNaseH) complex.

    PubMed

    Santos, Karine; Preussner, Marco; Heroven, Anna Christina; Weber, Gert

    2015-11-01

    In eukaryotes, the removal of nuclear noncoding sequences (pre-mRNA splicing) is catalyzed by the spliceosome, which consists of five ribonucleoprotein particles (U1, U2, U4, U5 and U6 snRNPs, each with a respective snRNA) and a plethora of protein factors that aid spliceosomal maturation, assembly, activation and disassembly. Recently, the U5 snRNP maturation factor Aar2p from baker's yeast has been characterized structurally and biochemically. Aar2p binds to the RNaseH (RH) and Jab1/MPN domains of the highly conserved U5-specific Prp8p, which forms a framework for the spliceosomal catalytic centre. Thereby, Aar2p sterically excludes Brr2p, a helicase essential for the catalytic activation of the spliceosome, from Prp8p binding. At the same time, Aar2p blocks U4/U6 di-snRNA binding to Prp8p. Aar2p therefore prevents premature spliceosome activation and its functions are regulated by reversible phosphorylation. To date, little is known about the hypothetical human Aar2 (hsAar2) orthologue C20ORF4. This study identifies C20ORF4 (i) as part of the HeLa proteome by Western blotting and (ii) as a true Aar2 orthologue which binds to the RH domain (hsRH) of Prp8 and corroborates an evolutionary link between yeast and human Aar2 function. An elaborate strategy was devised to crystallize hsAar2 in complex with hsRH. The analysis of initial weakly diffracting crystals obtained by in situ proteolysis and homology modelling guided the design of an hsAar2 construct in which an internal loop was replaced by three serines (hsAar2(Δloop)). A complex of hsAar2(Δloop) and hsRH crystallized in space group C2; the crystals diffracted to 2.35 Å resolution and were suitable for structure determination by molecular-replacement approaches. The study presented here suggests a connection between Aar2 and the spliceosome in human cells and paves the way for structural studies of human Aar2. PMID:26527271

  1. The 68 kDa subunit of mammalian cleavage factor I interacts with the U7 small nuclear ribonucleoprotein and participates in 3′-end processing of animal histone mRNAs

    PubMed Central

    Ruepp, Marc-David; Vivarelli, Silvia; Pillai, Ramesh S.; Kleinschmidt, Nicole; Azzouz, Teldja N.; Barabino, Silvia M. L.; Schümperli, Daniel

    2010-01-01

    Metazoan replication-dependent histone pre-mRNAs undergo a unique 3′-cleavage reaction which does not result in mRNA polyadenylation. Although the cleavage site is defined by histone-specific factors (hairpin binding protein, a 100-kDa zinc-finger protein and the U7 snRNP), a large complex consisting of cleavage/polyadenylation specificity factor, two subunits of cleavage stimulation factor and symplekin acts as the effector of RNA cleavage. Here, we report that yet another protein involved in cleavage/polyadenylation, mammalian cleavage factor I 68-kDa subunit (CF Im68), participates in histone RNA 3′-end processing. CF Im68 was found in a highly purified U7 snRNP preparation. Its interaction with the U7 snRNP depends on the N-terminus of the U7 snRNP protein Lsm11, known to be important for histone RNA processing. In vivo, both depletion and overexpression of CF Im68 cause significant decreases in processing efficiency. In vitro 3′-end processing is slightly stimulated by the addition of low amounts of CF Im68, but inhibited by high amounts or by anti-CF Im68 antibody. Finally, immunoprecipitation of CF Im68 results in a strong enrichment of histone pre-mRNAs. In contrast, the small CF Im subunit, CF Im25, does not appear to be involved in histone RNA processing. PMID:20634199

  2. U1 small nuclear ribonucleoprotein complex and RNA splicing alterations in Alzheimer’s disease

    PubMed Central

    Bai, Bing; Hales, Chadwick M.; Chen, Ping-Chung; Gozal, Yair; Dammer, Eric B.; Fritz, Jason J.; Wang, Xusheng; Xia, Qiangwei; Duong, Duc M.; Street, Craig; Cantero, Gloria; Cheng, Dongmei; Jones, Drew R.; Wu, Zhiping; Li, Yuxin; Diner, Ian; Heilman, Craig J.; Rees, Howard D.; Wu, Hao; Lin, Li; Szulwach, Keith E.; Gearing, Marla; Mufson, Elliott J.; Bennett, David A.; Montine, Thomas J.; Seyfried, Nicholas T.; Wingo, Thomas S.; Sun, Yi E.; Jin, Peng; Hanfelt, John; Willcock, Donna M.; Levey, Allan; Lah, James J.; Peng, Junmin

    2013-01-01

    Deposition of insoluble protein aggregates is a hallmark of neurodegenerative diseases. The universal presence of β-amyloid and tau in Alzheimer’s disease (AD) has facilitated advancement of the amyloid cascade and tau hypotheses that have dominated AD pathogenesis research and therapeutic development. However, the underlying etiology of the disease remains to be fully elucidated. Here we report a comprehensive study of the human brain-insoluble proteome in AD by mass spectrometry. We identify 4,216 proteins, among which 36 proteins accumulate in the disease, including U1-70K and other U1 small nuclear ribonucleoprotein (U1 snRNP) spliceosome components. Similar accumulations in mild cognitive impairment cases indicate that spliceosome changes occur in early stages of AD. Multiple U1 snRNP subunits form cytoplasmic tangle-like structures in AD but not in other examined neurodegenerative disorders, including Parkinson disease and frontotemporal lobar degeneration. Comparison of RNA from AD and control brains reveals dysregulated RNA processing with accumulation of unspliced RNA species in AD, including myc box-dependent-interacting protein 1, clusterin, and presenilin-1. U1-70K knockdown or antisense oligonucleotide inhibition of U1 snRNP increases the protein level of amyloid precursor protein. Thus, our results demonstrate unique U1 snRNP pathology and implicate abnormal RNA splicing in AD pathogenesis. PMID:24023061

  3. High-density genetic map of the BRCA1 region of chromosome 17q12-q21

    SciTech Connect

    Anderson, L.A.; Friedman, L.; Lynch, E.; King, M.C. ); Osborne-Lawrence, S.; Bowcock, A. ); Weissenbach, J. )

    1993-09-01

    To facilitate the positional cloning of the breast-ovarian cancer gene BRCA1, the authors constructed a high-density genetic map of the 8.3-cM interval between D17S250 and GIP on chromosome 17q12-q21. Markers were mapped by linkage in the CEPH and in extended kindreds in the breast cancer series. The map comprises 33 ordered polymorphisms, including 12 genes and 21 anonymous markers, yielding an average of one polymorphism every 250 kb. Twenty-five of the markers are PCR-based systems. The order of polymorphic genes and markers is cen-D17S250-D17S518-HER2-THRA1-RARA-D17S80-KRT10-[D17S800-D17S857]-GAS-D17S856-EDH17B-D17S855-D17S859-D17S858-[PPY-D17S78]-D17S183-EPB3-D17S579-D17S509-[D17S508-D17S190 = D17S810]-D17S791-[D17S181 = D17S806]-D17S797-HOX2B-GP3A-[D17S507 = GIP]-qter. BRCA1 lies in the middle of the interval, between THRA1 and D17S183. Markers from this map can be used to determine whether cancer is linked to BRCA1 in families, to evaluate whether tumors have lost heterozygosity at loci in the region, and to identify probes for characterizing chromosomal rearrangements from patients and from tumors. 21 refs., 1 fig., 3 tabs.

  4. Highly variable cancer subpopulations that exhibit enhanced transcriptome variability and metastatic fitness

    PubMed Central

    Nguyen, Alexander; Yoshida, Mitsukuni; Goodarzi, Hani; Tavazoie, Sohail F.

    2016-01-01

    Individual cells within a tumour can exhibit distinct genetic and molecular features. The impact of such diversification on metastatic potential is unknown. Here we identify clonal human breast cancer subpopulations that display different levels of morphological and molecular diversity. Highly variable subpopulations are more proficient at metastatic colonization and chemotherapeutic survival. Through single-cell RNA-sequencing, inter-cell transcript expression variability is identified as a defining feature of the highly variable subpopulations that leads to protein-level variation. Furthermore, we identify high variability in the spliceosomal machinery gene set. Engineered variable expression of the spliceosomal gene SNRNP40 promotes metastasis, attributable to cells with low expression. Clinically, low SNRNP40 expression is associated with metastatic relapse. Our findings reveal transcriptomic variability generation as a mechanism by which cancer subpopulations can diversify gene expression states, which may allow for enhanced fitness under changing environmental pressures encountered during cancer progression. PMID:27138336

  5. Purification of noncoding RNA and bound proteins using FLAG peptide-conjugated antisense-oligonucleotides.

    PubMed

    Adachi, Shungo; Natsume, Tohru

    2015-01-01

    To understand the function of certain RNAs, including noncoding RNAs, it is important to identify the proteins that interact with the RNAs. Here we describe the method for purification of ribonucleoprotein (RNP) complexes composed of specific cellular RNAs by pull-down with FLAG peptide-conjugated antisense oligonucleotide (ASO). Using this method, we identified a novel protein component of U7 snRNP complex.

  6. A Bidirectional SF2/ASF- and SRp40-Dependent Splicing Enhancer Regulates Human Immunodeficiency Virus Type 1 rev, env, vpu, and nef Gene Expression

    PubMed Central

    Caputi, Massimo; Freund, Marcel; Kammler, Susanne; Asang, Corinna; Schaal, Heiner

    2004-01-01

    The integrated human immunodeficiency virus type 1 (HIV-1) genome is transcribed in a single pre-mRNA that is alternatively spliced into more than 40 mRNAs. We characterized a novel bidirectional exonic splicing enhancer (ESE) that regulates the expression of the HIV-1 env, vpu, rev, and nef mRNAs. The ESE is localized downstream of the vpu-, env-, and nef-specific 3′ splice site no. 5. SF2/ASF and SRp40 activate the ESE and are required for efficient 3′ splice site usage and binding of the U1 snRNP to the downstream 5′ splice site no. 4. U1 snRNP binding to the 5′ splice site no. 4 is required for splicing of the rev and nef mRNAs and to increase expression of the partially spliced env mRNA. Finally, our results indicate that this ESE is necessary for the recruitment of the U1 snRNP to the 5′ splice site no. 4, even when the 5′ splice site and the U1 snRNA have been mutated to obtain a perfect complementary match. The ESE characterized here is highly conserved in most viral subtypes. PMID:15163745

  7. Calpain 2 Activation of P-TEFb Drives Megakaryocyte Morphogenesis and Is Disrupted by Leukemogenic GATA1 Mutation

    PubMed Central

    Elagib, Kamaleldin E.; Rubinstein, Jeremy D.; Delehanty, Lorrie L.; Ngoh, Valerie S.; Greer, Peter A.; Li, Shuran; Lee, Jae K.; Li, Zhe; Orkin, Stuart H.; Mihaylov, Ivailo S.; Goldfarb, Adam N.

    2014-01-01

    SUMMARY Megakaryocyte morphogenesis employs a “hypertrophy-like” developmental program, dependent on P-TEFb kinase activation and cytoskeletal remodeling. P-TEFb activation classically occurs by a feedback regulated process of signal-induced, reversible release of active Cdk9-cyclin T modules from large inactive 7SK snRNP complexes. Here we have identified an alternative pathway of irreversible P-TEFb activation in megakaryopoiesis, mediated by dissolution of the 7SK snRNP complex. In this pathway calpain 2 cleavage of the core 7SK snRNP component MePCE promoted P-TEFb release and consequent upregulation of a cohort of cytoskeleton remodeling factors, including α-actinin-1. In a subset of human megakaryocytic leukemias, the transcription factor GATA1 undergoes truncating mutation (GATA1s). Here we linked the GATA1s mutation to defects in megakaryocytic upregulation of calpain 2 and of P-TEFb-dependent cytoskeletal remodeling factors. Restoring calpain 2 expression in GATA1s-mutant megakaryocytes rescued normal development, implicating this morphogenetic pathway as a target in human leukemogenesis. PMID:24369834

  8. Crystal structure of (acetato-κO)(ethanol-κO)[(9S,17S,21S,29S)-9,17,21,29-tetra-hydroxy-18,30-dioxa-octa-cyclo-[18.10.0.0(2,7).0(8,19).0(9,17).0(11,16).0(21,29).0(23,28)]triaconta-1,3,5,7,11(16),12,14,19,23(28),24,26-undeca-ene-10,22-dione-κ(3) O (18),O (21),O (22)]caesium ethanol monosolvate.

    PubMed

    Bengiat, Ravell; Gil, Maayan; Klein, Asne; Bogoslavsky, Benny; Cohen, Shmuel; Yardeni, Guy; Zilbermann, Israel; Almog, Joseph

    2016-07-01

    The title compound, [Cs(CH3COO)(C28H16O8)(C2H5OH)]·C2H5OH, is the product of the complexation between one vasarene analogue [1], bis ninhydrin naphthalene-1,3-diol and CsF, where the F(-) ion has reacted with residual acetic acid (AcOH), to form a [1]·CsOAc complex. The inter-molecular inter-actions with the multiple oxygen-containing functional groups of the ligand, as well as O-H⋯O hydrogen bonds involving the ethanol solvent mol-ecules, stabilize the complex, forming a chain along [100]. Additional parallel-displaced π-π stacking, with an inter-planar distance of 3.669 (1) Å, connect several unit cells in a three-dimensional supra-molecular structure, though, the larger size of AcO(-) (1.60 Å) compared to F(-) (1.33 Å) prevents the tight packing that was once achieved with other vasarene complexes of CsF.

  9. Crystal structure of (acetato-κO)(ethanol-κO)[(9S,17S,21S,29S)-9,17,21,29-tetra­hydroxy-18,30-dioxa­octa­cyclo­[18.10.0.02,7.08,19.09,17.011,16.021,29.023,28]triaconta-1,3,5,7,11(16),12,14,19,23(28),24,26-undeca­ene-10,22-dione-κ3 O 18,O 21,O 22]caesium ethanol monosolvate

    PubMed Central

    Bengiat, Ravell; Gil, Maayan; Klein, Asne; Bogoslavsky, Benny; Cohen, Shmuel; Yardeni, Guy; Zilbermann, Israel; Almog, Joseph

    2016-01-01

    The title compound, [Cs(CH3COO)(C28H16O8)(C2H5OH)]·C2H5OH, is the product of the complexation between one vasarene analogue [1], bis ninhydrin naphthalene-1,3-diol and CsF, where the F− ion has reacted with residual acetic acid (AcOH), to form a [1]·CsOAc complex. The inter­molecular inter­actions with the multiple oxygen-containing functional groups of the ligand, as well as O—H⋯O hydrogen bonds involving the ethanol solvent mol­ecules, stabilize the complex, forming a chain along [100]. Additional parallel-displaced π–π stacking, with an inter­planar distance of 3.669 (1) Å, connect several unit cells in a three-dimensional supra­molecular structure, though, the larger size of AcO− (1.60 Å) compared to F− (1.33 Å) prevents the tight packing that was once achieved with other vasarene complexes of CsF. PMID:27555923

  10. Crystal structure of (acetato-κO)(ethanol-κO)[(9S,17S,21S,29S)-9,17,21,29-tetra-hydroxy-18,30-dioxa-octa-cyclo-[18.10.0.0(2,7).0(8,19).0(9,17).0(11,16).0(21,29).0(23,28)]triaconta-1,3,5,7,11(16),12,14,19,23(28),24,26-undeca-ene-10,22-dione-κ(3) O (18),O (21),O (22)]caesium ethanol monosolvate.

    PubMed

    Bengiat, Ravell; Gil, Maayan; Klein, Asne; Bogoslavsky, Benny; Cohen, Shmuel; Yardeni, Guy; Zilbermann, Israel; Almog, Joseph

    2016-07-01

    The title compound, [Cs(CH3COO)(C28H16O8)(C2H5OH)]·C2H5OH, is the product of the complexation between one vasarene analogue [1], bis ninhydrin naphthalene-1,3-diol and CsF, where the F(-) ion has reacted with residual acetic acid (AcOH), to form a [1]·CsOAc complex. The inter-molecular inter-actions with the multiple oxygen-containing functional groups of the ligand, as well as O-H⋯O hydrogen bonds involving the ethanol solvent mol-ecules, stabilize the complex, forming a chain along [100]. Additional parallel-displaced π-π stacking, with an inter-planar distance of 3.669 (1) Å, connect several unit cells in a three-dimensional supra-molecular structure, though, the larger size of AcO(-) (1.60 Å) compared to F(-) (1.33 Å) prevents the tight packing that was once achieved with other vasarene complexes of CsF. PMID:27555923

  11. Studies of the 5' exonuclease and endonuclease activities of CPSF-73 in histone pre-mRNA processing.

    PubMed

    Yang, Xiao-cui; Sullivan, Kelly D; Marzluff, William F; Dominski, Zbigniew

    2009-01-01

    Processing of histone pre-mRNA requires a single 3' endonucleolytic cleavage guided by the U7 snRNP that binds downstream of the cleavage site. Following cleavage, the downstream cleavage product (DCP) is rapidly degraded in vitro by a nuclease that also depends on the U7 snRNP. Our previous studies demonstrated that the endonucleolytic cleavage is catalyzed by the cleavage/polyadenylation factor CPSF-73. Here, by using RNA substrates with different nucleotide modifications, we characterize the activity that degrades the DCP. We show that the degradation is blocked by a 2'-O-methyl nucleotide and occurs in the 5'-to-3' direction. The U7-dependent 5' exonuclease activity is processive and continues degrading the DCP substrate even after complete removal of the U7-binding site. Thus, U7 snRNP is required only to initiate the degradation. UV cross-linking studies demonstrate that the DCP and its 5'-truncated version specifically interact with CPSF-73, strongly suggesting that in vitro, the same protein is responsible for the endonucleolytic cleavage of histone pre-mRNA and the subsequent degradation of the DCP. By using various RNA substrates, we define important space requirements upstream and downstream of the cleavage site that dictate whether CPSF-73 functions as an endonuclease or a 5' exonuclease. RNA interference experiments with HeLa cells indicate that degradation of the DCP does not depend on the Xrn2 5' exonuclease, suggesting that CPSF-73 degrades the DCP both in vitro and in vivo. PMID:18955505

  12. Crystal structures of the Lsm complex bound to the 3' end sequence of U6 small nuclear RNA.

    PubMed

    Zhou, Lijun; Hang, Jing; Zhou, Yulin; Wan, Ruixue; Lu, Guifeng; Yin, Ping; Yan, Chuangye; Shi, Yigong

    2014-02-01

    Splicing of precursor messenger RNA (pre-mRNA) in eukaryotic cells is carried out by the spliceosome, which consists of five small nuclear ribonucleoproteins (snRNPs) and a number of accessory factors and enzymes. Each snRNP contains a ring-shaped subcomplex of seven proteins and a specific RNA molecule. The U6 snRNP contains a unique heptameric Lsm protein complex, which specifically recognizes the U6 small nuclear RNA at its 3' end. Here we report the crystal structures of the heptameric Lsm complex, both by itself and in complex with a 3' fragment of U6 snRNA, at 2.8 Å resolution. Each of the seven Lsm proteins interacts with two neighbouring Lsm components to form a doughnut-shaped assembly, with the order Lsm3-2-8-4-7-5-6. The four uridine nucleotides at the 3' end of U6 snRNA are modularly recognized by Lsm3, Lsm2, Lsm8 and Lsm4, with the uracil base specificity conferred by a highly conserved asparagine residue. The uracil base at the extreme 3' end is sandwiched by His 36 and Arg 69 from Lsm3, through π-π and cation-π interactions, respectively. The distinctive end-recognition of U6 snRNA by the Lsm complex contrasts with RNA binding by the Sm complex in the other snRNPs. The structural features and associated biochemical analyses deepen mechanistic understanding of the U6 snRNP function in pre-mRNA splicing.

  13. Functional interplay between PPM1G and the transcription elongation machinery

    PubMed Central

    Gudipaty, Swapna Aravind; D’Orso, Iván

    2016-01-01

    Transcription elongation is a critical regulatory step in the gene expression cycle. One key regulator of the switch between transcription initiation and elongation is the P-TEFb kinase, which phosphorylates RNA polymerase II (Pol II) and several negative elongation factors to relieve the elongation block at paused promoters to facilitate productive elongation. Here, we highlight recent findings signifying the role of the PPM1G/PP2Cγ phosphatase in activating and maintaining the active transcription elongation state by regulating the availability of P-TEFb and blocking its assembly into the catalytic inactive 7SK small nuclear ribonucleoprotein (snRNP) complex. PMID:27088130

  14. Spliceosomal snRNA modifications and their function

    PubMed Central

    Karijolich, John; Yu, Yi-Tao

    2014-01-01

    Spliceosomal snRNAs are extensively 2'-O-methylated and pseudouridylated. The modified nucleotides are relatively highly conserved across species, and are often clustered in regions of functional importance in pre-mRNA splicing. Over the past decade, the study of the mechanisms and functions of spliceosomal snRNA modifications has intensified. Two independent mechanisms behind these modifications, RNA-independent (protein-only) and RNA-dependent (RNA-guided), have been discovered. The role of spliceosomal snRNA modifications in snRNP biogenesis and spliceosome assembly has also been verified. PMID:20215871

  15. Cancer-Associated Perturbations in Alternative Pre-messenger RNA Splicing.

    PubMed

    Shkreta, Lulzim; Bell, Brendan; Revil, Timothée; Venables, Julian P; Prinos, Panagiotis; Elela, Sherif Abou; Chabot, Benoit

    2013-01-01

    For most of our 25,000 genes, the removal of introns by pre-messenger RNA (pre-mRNA) splicing represents an essential step toward the production of functional messenger RNAs (mRNAs). Alternative splicing of a single pre-mRNA results in the production of different mRNAs. Although complex organisms use alternative splicing to expand protein function and phenotypic diversity, patterns of alternative splicing are often altered in cancer cells. Alternative splicing contributes to tumorigenesis by producing splice isoforms that can stimulate cell proliferation and cell migration or induce resistance to apoptosis and anticancer agents. Cancer-specific changes in splicing profiles can occur through mutations that are affecting splice sites and splicing control elements, and also by alterations in the expression of proteins that control splicing decisions. Recent progress in global approaches that interrogate splicing diversity should help to obtain specific splicing signatures for cancer types. The development of innovative approaches for annotating and reprogramming splicing events will more fully establish the essential contribution of alternative splicing to the biology of cancer and will hopefully provide novel targets and anticancer strategies. Metazoan genes are usually made up of several exons interrupted by introns. The introns are removed from the pre-mRNA by RNA splicing. In conjunction with other maturation steps, such as capping and polyadenylation, the spliced mRNA is then transported to the cytoplasm to be translated into a functional protein. The basic mechanism of splicing requires accurate recognition of each extremity of each intron by the spliceosome. Introns are identified by the binding of U1 snRNP to the 5' splice site and the U2AF65/U2AF35 complex to the 3' splice site. Following these interactions, other proteins and snRNPs are recruited to generate the complete spliceosomal complex needed to excise the intron. While many introns are constitutively

  16. CYCLIN-DEPENDENT KINASE G1 is associated with the spliceosome to regulate CALLOSE SYNTHASE5 splicing and pollen wall formation in Arabidopsis.

    PubMed

    Huang, Xue-Yong; Niu, Jin; Sun, Ming-Xi; Zhu, Jun; Gao, Ju-Fang; Yang, Jun; Zhou, Que; Yang, Zhong-Nan

    2013-02-01

    Arabidopsis thaliana CYCLIN-DEPEDENT KINASE G1 (CDKG1) belongs to the family of cyclin-dependent protein kinases that were originally characterized as cell cycle regulators in eukaryotes. Here, we report that CDKG1 regulates pre-mRNA splicing of CALLOSE SYNTHASE5 (CalS5) and, therefore, pollen wall formation. The knockout mutant cdkg1 exhibits reduced male fertility with impaired callose synthesis and abnormal pollen wall formation. The sixth intron in CalS5 pre-mRNA, a rare type of intron with a GC 5' splice site, is abnormally spliced in cdkg1. RNA immunoprecipitation analysis suggests that CDKG1 is associated with this intron. CDKG1 contains N-terminal Ser/Arg (RS) motifs and interacts with splicing factor Arginine/Serine-Rich Zinc Knuckle-Containing Protein33 (RSZ33) through its RS region to regulate proper splicing. CDKG1 and RS-containing Zinc Finger Protein22 (SRZ22), a splicing factor interacting with RSZ33 and U1 small nuclear ribonucleoprotein particle (snRNP) component U1-70k, colocalize in nuclear speckles and reside in the same complex. We propose that CDKG1 is recruited to U1 snRNP through RSZ33 to facilitate the splicing of the sixth intron of CalS5.

  17. The Ribosome Cooperates with the Assembly Chaperone pICln to Initiate Formation of snRNPs.

    PubMed

    Paknia, Elham; Chari, Ashwin; Stark, Holger; Fischer, Utz

    2016-09-20

    The formation of macromolecular complexes within the crowded environment of cells often requires aid from assembly chaperones. PRMT5 and SMN complexes mediate this task for the assembly of the common core of pre-mRNA processing small nuclear ribonucleoprotein particles (snRNPs). Core formation is initiated by the PRMT5-complex subunit pICln, which pre-arranges the core proteins into spatial positions occupied in the assembled snRNP. The SMN complex then accepts these pICln-bound proteins and unites them with small nuclear RNA (snRNA). Here, we have analyzed how newly synthesized snRNP proteins are channeled into the assembly pathway to evade mis-assembly. We show that they initially remain bound to the ribosome near the polypeptide exit tunnel and dissociate upon association with pICln. Coincident with its release activity, pICln ensures the formation of cognate heterooligomers and their chaperoned guidance into the assembly pathway. Our study identifies the ribosomal quality control hub as a site where chaperone-mediated assembly of macromolecular complexes can be initiated. PMID:27653676

  18. Deletion of SNURF/SNRPN U1B and U1B* upstream exons in a child with developmental delay and excessive weight.

    PubMed

    Koufaris, Costas; Alexandrou, Angelos; Papaevripidou, Ioannis; Alexandrou, Ioanna; Christophidou-Anastasiadou, Violetta; Sismani, Carolina

    2016-09-01

    Prader-Willi syndrome is a rare syndrome characterized by hypotonia, developmental delay and excessive appetite. This syndrome is caused by the loss of function of paternally-expressed genes located in an imprinting centre in 15q11-q13. Here, we report the case of a patient who was referred to us with Prader-Willi syndrome-like symptoms including obesity and developmental delay. Examination of this patient revealed that he was a carrier of a paternally inherited deletion that affected the U1B and U1B* upstream exons of the SNURF-SNRNP gene within the 15q11-q13 imprinted region. Mutations localized within this genomic region have not been previously reported in Prader-Willi syndrome patients. It is possible that disruption of upstream exons of SNURF-SNRNP could contribute to Prader-Willi phenotype by disrupting brain-specific alternative transcripts, although, case reports from further patients with a comparable phenotype are required. PMID:27659333

  19. Interaction between RNA helicase ROOT INITIATION DEFECTIVE 1 and GAMETOPHYTIC FACTOR 1 is involved in female gametophyte development in Arabidopsis

    PubMed Central

    Zhu, Dong Zi; Zhao, Xue Fang; Liu, Chang Zhen; Ma, Fang Fang; Wang, Fang; Gao, Xin-Qi; Zhang, Xian Sheng

    2016-01-01

    ROOT INITIATION DEFECTIVE 1 (RID1) is an Arabidopsis DEAH/RHA RNA helicase. It functions in hypocotyl de-differentiation, de novo meristem formation, and cell specification of the mature female gametophyte (FG). However, it is unclear how RID1 regulates FG development. In this study, we observed that mutations to RID1 disrupted the developmental synchrony and retarded the progression of FG development. RID1 exhibited RNA helicase activity, with a preference for unwinding double-stranded RNA in the 3′ to 5′ direction. Furthermore, we found that RID1 interacts with GAMETOPHYTIC FACTOR 1 (GFA1), which is an integral protein of the spliceosome component U5 small nuclear ribonucleoprotein (snRNP) particle. Substitution of specific RID1 amino acids (Y266F and T267I) inhibited the interaction with GFA1. In addition, the mutated RID1 could not complement the seed-abortion phenotype of the rid1 mutant. The rid1 and gfa1 mutants exhibited similar abnormalities in pre-mRNA splicing and down-regulated expression of some genes involved in FG development. Our results suggest that an interaction between RID1 and the U5 snRNP complex regulates essential pre-mRNA splicing of the genes required for FG development. This study provides new information regarding the mechanism underlying the FG developmental process. PMID:27683728

  20. The Dengue Virus NS5 Protein Intrudes in the Cellular Spliceosome and Modulates Splicing.

    PubMed

    De Maio, Federico A; Risso, Guillermo; Iglesias, Nestor G; Shah, Priya; Pozzi, Berta; Gebhard, Leopoldo G; Mammi, Pablo; Mancini, Estefania; Yanovsky, Marcelo J; Andino, Raul; Krogan, Nevan; Srebrow, Anabella; Gamarnik, Andrea V

    2016-08-01

    Dengue virus NS5 protein plays multiple functions in the cytoplasm of infected cells, enabling viral RNA replication and counteracting host antiviral responses. Here, we demonstrate a novel function of NS5 in the nucleus where it interferes with cellular splicing. Using global proteomic analysis of infected cells together with functional studies, we found that NS5 binds spliceosome complexes and modulates endogenous splicing as well as minigene-derived alternative splicing patterns. In particular, we show that NS5 alone, or in the context of viral infection, interacts with core components of the U5 snRNP particle, CD2BP2 and DDX23, alters the inclusion/exclusion ratio of alternative splicing events, and changes mRNA isoform abundance of known antiviral factors. Interestingly, a genome wide transcriptome analysis, using recently developed bioinformatics tools, revealed an increase of intron retention upon dengue virus infection, and viral replication was improved by silencing specific U5 components. Different mechanistic studies indicate that binding of NS5 to the spliceosome reduces the efficiency of pre-mRNA processing, independently of NS5 enzymatic activities. We propose that NS5 binding to U5 snRNP proteins hijacks the splicing machinery resulting in a less restrictive environment for viral replication. PMID:27575636

  1. Splicing enhancement in the yeast rp51b intron.

    PubMed Central

    Libri, D; Lescure, A; Rosbash, M

    2000-01-01

    Splicing enhancement in higher eukaryotes has been linked to SR proteins, to U1 snRNP, and to communication between splice sites across introns or exons mediated by protein-protein interactions. It has been previously shown that, in yeast, communication mediated by RNA-RNA interactions between the two ends of introns is a basis for splicing enhancement. We designed experiments of randomization-selection to isolate splicing enhancers that would work independently from RNA secondary structures. Surprisingly, one of the two families of sequences selected was essentially composed of 5' splice site variants. We show that this sequence enhances splicing independently of secondary structure, is exportable to heterologous contexts, and works in multiple copies with additive effects. The data argue in favor of an early role for splicing enhancement, possibly coincident with commitment complex formation. Genetic compensation experiments with U1 snRNA mutants suggest that U1 snRNP binding to noncanonical locations is required for splicing enhancement. PMID:10744020

  2. The Dengue Virus NS5 Protein Intrudes in the Cellular Spliceosome and Modulates Splicing.

    PubMed

    De Maio, Federico A; Risso, Guillermo; Iglesias, Nestor G; Shah, Priya; Pozzi, Berta; Gebhard, Leopoldo G; Mammi, Pablo; Mancini, Estefania; Yanovsky, Marcelo J; Andino, Raul; Krogan, Nevan; Srebrow, Anabella; Gamarnik, Andrea V

    2016-08-01

    Dengue virus NS5 protein plays multiple functions in the cytoplasm of infected cells, enabling viral RNA replication and counteracting host antiviral responses. Here, we demonstrate a novel function of NS5 in the nucleus where it interferes with cellular splicing. Using global proteomic analysis of infected cells together with functional studies, we found that NS5 binds spliceosome complexes and modulates endogenous splicing as well as minigene-derived alternative splicing patterns. In particular, we show that NS5 alone, or in the context of viral infection, interacts with core components of the U5 snRNP particle, CD2BP2 and DDX23, alters the inclusion/exclusion ratio of alternative splicing events, and changes mRNA isoform abundance of known antiviral factors. Interestingly, a genome wide transcriptome analysis, using recently developed bioinformatics tools, revealed an increase of intron retention upon dengue virus infection, and viral replication was improved by silencing specific U5 components. Different mechanistic studies indicate that binding of NS5 to the spliceosome reduces the efficiency of pre-mRNA processing, independently of NS5 enzymatic activities. We propose that NS5 binding to U5 snRNP proteins hijacks the splicing machinery resulting in a less restrictive environment for viral replication.

  3. Structural requirements for protein-catalyzed annealing of U4 and U6 RNAs during di-snRNP assembly

    PubMed Central

    Didychuk, Allison L.; Montemayor, Eric J.; Brow, David A.; Butcher, Samuel E.

    2016-01-01

    Base-pairing of U4 and U6 snRNAs during di-snRNP assembly requires large-scale remodeling of RNA structure that is chaperoned by the U6 snRNP protein Prp24. We investigated the mechanism of U4/U6 annealing in vitro using an assay that enables visualization of ribonucleoprotein complexes and faithfully recapitulates known in vivo determinants for the process. We find that annealing, but not U6 RNA binding, is highly dependent on the electropositive character of a 20 Å-wide groove on the surface of Prp24. During annealing, we observe the formation of a stable ternary complex between U4 and U6 RNAs and Prp24, indicating that displacement of Prp24 in vivo requires additional factors. Mutations that stabilize the U6 ‘telestem’ helix increase annealing rates by up to 15-fold, suggesting that telestem formation is rate-limiting for U4/U6 pairing. The Lsm2–8 complex, which binds adjacent to the telestem at the 3′ end of U6, provides a comparable rate enhancement. Collectively, these data identify domains of the U6 snRNP that are critical for one of the first steps in assembly of the megaDalton U4/U6.U5 tri-snRNP complex, and lead to a dynamic model for U4/U6 pairing that involves a striking degree of evolved cooperativity between protein and RNA. PMID:26673715

  4. CYCLIN-DEPENDENT KINASE G1 Is Associated with the Spliceosome to Regulate CALLOSE SYNTHASE5 Splicing and Pollen Wall Formation in Arabidopsis[C][W][OA

    PubMed Central

    Huang, Xue-Yong; Niu, Jin; Sun, Ming-Xi; Zhu, Jun; Gao, Ju-Fang; Yang, Jun; Zhou, Que; Yang, Zhong-Nan

    2013-01-01

    Arabidopsis thaliana CYCLIN-DEPEDENT KINASE G1 (CDKG1) belongs to the family of cyclin-dependent protein kinases that were originally characterized as cell cycle regulators in eukaryotes. Here, we report that CDKG1 regulates pre-mRNA splicing of CALLOSE SYNTHASE5 (CalS5) and, therefore, pollen wall formation. The knockout mutant cdkg1 exhibits reduced male fertility with impaired callose synthesis and abnormal pollen wall formation. The sixth intron in CalS5 pre-mRNA, a rare type of intron with a GC 5′ splice site, is abnormally spliced in cdkg1. RNA immunoprecipitation analysis suggests that CDKG1 is associated with this intron. CDKG1 contains N-terminal Ser/Arg (RS) motifs and interacts with splicing factor Arginine/Serine-Rich Zinc Knuckle-Containing Protein33 (RSZ33) through its RS region to regulate proper splicing. CDKG1 and RS-containing Zinc Finger Protein22 (SRZ22), a splicing factor interacting with RSZ33 and U1 small nuclear ribonucleoprotein particle (snRNP) component U1-70k, colocalize in nuclear speckles and reside in the same complex. We propose that CDKG1 is recruited to U1 snRNP through RSZ33 to facilitate the splicing of the sixth intron of CalS5. PMID:23404887

  5. The Dengue Virus NS5 Protein Intrudes in the Cellular Spliceosome and Modulates Splicing

    PubMed Central

    Shah, Priya; Pozzi, Berta; Gebhard, Leopoldo G.; Mammi, Pablo; Yanovsky, Marcelo J.; Andino, Raul; Krogan, Nevan; Srebrow, Anabella; Gamarnik, Andrea V.

    2016-01-01

    Dengue virus NS5 protein plays multiple functions in the cytoplasm of infected cells, enabling viral RNA replication and counteracting host antiviral responses. Here, we demonstrate a novel function of NS5 in the nucleus where it interferes with cellular splicing. Using global proteomic analysis of infected cells together with functional studies, we found that NS5 binds spliceosome complexes and modulates endogenous splicing as well as minigene-derived alternative splicing patterns. In particular, we show that NS5 alone, or in the context of viral infection, interacts with core components of the U5 snRNP particle, CD2BP2 and DDX23, alters the inclusion/exclusion ratio of alternative splicing events, and changes mRNA isoform abundance of known antiviral factors. Interestingly, a genome wide transcriptome analysis, using recently developed bioinformatics tools, revealed an increase of intron retention upon dengue virus infection, and viral replication was improved by silencing specific U5 components. Different mechanistic studies indicate that binding of NS5 to the spliceosome reduces the efficiency of pre-mRNA processing, independently of NS5 enzymatic activities. We propose that NS5 binding to U5 snRNP proteins hijacks the splicing machinery resulting in a less restrictive environment for viral replication. PMID:27575636

  6. Knock-out mutations of Arabidopsis SmD3-b induce pleotropic phenotypes through altered transcript splicing.

    PubMed

    Swaraz, A M; Park, Young-Doo; Hur, Yoonkang

    2011-05-01

    SmD3 is a core protein of small nuclear ribonucleoprotein (snRNP) essential for splicing of primary transcripts. To elucidate function of SmD3 protein in plants, phenotypes and gene expression of SmD3 knock-out and overexpressing mutants in Arabidopsis have been analyzed. smd3-a knock-out mutant or SmD3-a and SmD3-b overexpressors did not show phenotypic alteration. Knock-out of SmD3-b resulted in the pleotropic phenotypes of delayed flowering time and completion of life cycle, reduced root growth, partially defective leaf venation, abnormal numbers of trichome branches, and changed numbers of floral organs. Microarray data revealed that the smd3-b mutant had altered expression of genes related to the above phenotypes, indirectly suggesting that changed splicing of these genes may cause the observed phenotypes. Splicing of selected genes was either totally blocked or reduced in the smd3-b mutant, indicating the important role of SmD3-b in the process. A double knock-out mutant of smd3-a and smd3-b could not be generated, indicating possible redundant function of these two genes. All data indicate that SmD3-b may be major component of the spliceosomal snRNP in Arabidopsis, but the function of SmD3-a may be redundant.

  7. Cajal bodies are linked to genome conformation

    PubMed Central

    Wang, Qiuyan; Sawyer, Iain A.; Sung, Myong-Hee; Sturgill, David; Shevtsov, Sergey P.; Pegoraro, Gianluca; Hakim, Ofir; Baek, Songjoon; Hager, Gordon L.; Dundr, Miroslav

    2016-01-01

    The mechanisms underlying nuclear body (NB) formation and their contribution to genome function are unknown. Here we examined the non-random positioning of Cajal bodies (CBs), major NBs involved in spliceosomal snRNP assembly and their role in genome organization. CBs are predominantly located at the periphery of chromosome territories at a multi-chromosome interface. Genome-wide chromosome conformation capture analysis (4C-seq) using CB-interacting loci revealed that CB-associated regions are enriched with highly expressed histone genes and U small nuclear or nucleolar RNA (sn/snoRNA) loci that form intra- and inter-chromosomal clusters. In particular, we observed a number of CB-dependent gene-positioning events on chromosome 1. RNAi-mediated disassembly of CBs disrupts the CB-targeting gene clusters and suppresses the expression of U sn/snoRNA and histone genes. This loss of spliceosomal snRNP production results in increased splicing noise, even in CB-distal regions. Therefore, we conclude that CBs contribute to genome organization with global effects on gene expression and RNA splicing fidelity. PMID:26997247

  8. Tissue-specific expression and cDNA cloning of small nuclear ribonucleoprotein-associated polypeptide N

    SciTech Connect

    McAllister, G.; Amara, S.G.; Lerner, M.R. )

    1988-07-01

    Sera from some patients with systemic lupus erythematosus and other autoimmune diseases have antibodies against nuclear antigens. An example is anti-Sm sera, which recognize proteins associated with small nuclear RNA molecules (small nuclear ribonucleoprotein (snRNP) particles). In this paper anti-Sm sera were used to probe immunoblots of various rat tissues. A previously unidentified M{sub r} 28,000 polypeptide was recognized by these anti-Sm sera. This polypeptide, referred to as N, is expressed in a tissue-specific manner, being most abundant in rat brain, less so in heart, and undetectable in the other tissues examined. Immunoprecipitation experiments using antibodies directed against the cap structure of small nuclear RNAs have demonstrated that N is a snRNP-associated polypeptide. Anti-Sm serum was also used to isolate a partial cDNA clone ({lambda}rb91) from a rat brain phage {lambda}gt11 cDNA expression library. A longer cDNA clone was obtained by rescreening the library with {lambda}rb91. In vitro transcription and subsequent translation of this subcloned, longer insert (pGMA2) resulted in a protein product with the same electrophoretic and immunological properties as N, confirming that pGMA2 encodes N. The tissue distribution of N and the involvement of snRNP particles in nuclear pre-mRNA processing may imply a role for N in tissue-specific pre-mRNA splicing.

  9. Recruitment of the NineTeen Complex to the activated spliceosome requires AtPRMT5

    PubMed Central

    Deng, Xian; Lu, Tiancong; Wang, Lulu; Gu, Lianfeng; Sun, Jing; Kong, Xiangfeng; Liu, Chunyan; Cao, Xiaofeng

    2016-01-01

    Protein arginine methylation, catalyzed by protein arginine methyltransferases (PRMTs), is involved in a multitude of biological processes in eukaryotes. Symmetric arginine dimethylation mediated by PRMT5 modulates constitutive and alternative pre-mRNA splicing of diverse genes to regulate normal growth and development in multiple species; however, the underlying molecular mechanism remains largely unknown. A genetic screen for suppressors of an Arabidopsis symmetric arginine dimethyltransferase mutant, atprmt5, identified two gain-of-function alleles of pre-mRNA processing factor 8 gene (prp8-8 and prp8-9), the highly conserved core component of the U5 small nuclear ribonucleoprotein (snRNP) and the spliceosome. These two atprmt5 prp8 double mutants showed suppression of the developmental and splicing alterations of atprmt5 mutants. In atprmt5 mutants, the NineTeen complex failed to be assembled into the U5 snRNP to form an activated spliceosome; this phenotype was restored in the atprmt5 prp8-8 double mutants. We also found that loss of symmetric arginine dimethylation of Sm proteins prevents recruitment of the NineTeen complex and initiation of spliceosome activation. Together, our findings demonstrate that symmetric arginine dimethylation has important functions in spliceosome assembly and activation, and uncover a key molecular mechanism for arginine methylation in pre-mRNA splicing that impacts diverse developmental processes. PMID:27114555

  10. Inhibition of SNW1 association with spliceosomal proteins promotes apoptosis in breast cancer cells

    PubMed Central

    Sato, Naoki; Maeda, Masao; Sugiyama, Mai; Ito, Satoko; Hyodo, Toshinori; Masuda, Akio; Tsunoda, Nobuyuki; Kokuryo, Toshio; Hamaguchi, Michinari; Nagino, Masato; Senga, Takeshi

    2015-01-01

    RNA splicing is a fundamental process for protein synthesis. Recent studies have reported that drugs that inhibit splicing have cytotoxic effects on various tumor cell lines. In this report, we demonstrate that depletion of SNW1, a component of the spliceosome, induces apoptosis in breast cancer cells. Proteomics and biochemical analyses revealed that SNW1 directly associates with other spliceosome components, including EFTUD2 (Snu114) and SNRNP200 (Brr2). The SKIP region of SNW1 interacted with the N-terminus of EFTUD2 as well as two independent regions in the C-terminus of SNRNP200. Similar to SNW1 depletion, knockdown of EFTUD2 increased the numbers of apoptotic cells. Furthermore, we demonstrate that exogenous expression of either the SKIP region of SNW1 or the N-terminus region of EFTUD2 significantly promoted cellular apoptosis. Our results suggest that the inhibition of SNW1 or its associating proteins may be a novel therapeutic strategy for cancer treatment. PMID:25450007

  11. Identification and characterisation of a nuclear localisation signal in the SMN associated protein, Gemin4

    SciTech Connect

    Lorson, Monique A.; Dickson, Alexa M.; Shaw, Debra J.; Todd, Adrian G.; Young, Elizabeth C.; Morse, Robert; Wolstencroft, Catherine; Lorson, Christian L.; Young, Philip J.

    2008-10-10

    Gemin4 is a ubiquitously expressed multifunctional protein that is involved in U snRNP assembly, apoptosis, nuclear/cytoplasmic transportation, transcription, and RNAi pathways. Gemin4 is one of the core components of the Gemin-complex, which also contains survival motor neuron (SMN), the seven Gemin proteins (Gemin2-8), and Unrip. Mutations in the SMN1 gene cause the autosomal recessive disorder spinal muscular atrophy (SMA). Although the functions assigned to Gemin4 predominantly occur in the nucleus, the mechanisms that mediate the nuclear import of Gemin4 remain unclear. Here, using a novel panel of Gemin4 constructs we identify a canonical nuclear import sequence (NLS) in the N-terminus of Gemin4. The Gemin4 NLS is necessary and independently sufficient to mediate nuclear import of Gemin4. This is the first functional NLS identified within the SMN-Gemin complex.

  12. Defective minor spliceosome mRNA processing results in isolated familial growth hormone deficiency

    PubMed Central

    Argente, Jesús; Flores, Raquel; Gutiérrez-Arumí, Armand; Verma, Bhupendra; Martos-Moreno, Gabriel Á; Cuscó, Ivon; Oghabian, Ali; Chowen, Julie A; Frilander, Mikko J; Pérez-Jurado, Luis A

    2014-01-01

    The molecular basis of a significant number of cases of isolated growth hormone deficiency remains unknown. We describe three sisters affected with severe isolated growth hormone deficiency and pituitary hypoplasia caused by biallelic mutations in the RNPC3 gene, which codes for a minor spliceosome protein required for U11/U12 small nuclear ribonucleoprotein (snRNP) formation and splicing of U12-type introns. We found anomalies in U11/U12 di-snRNP formation and in splicing of multiple U12-type introns in patient cells. Defective transcripts include preprohormone convertases SPCS2 and SPCS3 and actin-related ARPC5L genes, which are candidates for the somatotroph-restricted dysfunction. The reported novel mechanism for familial growth hormone deficiency demonstrates that general mRNA processing defects of the minor spliceosome can lead to very narrow tissue-specific consequences. Subject Categories Genetics, Gene Therapy ' Genetic Disease; Metabolism PMID:24480542

  13. Drosophila SMN complex proteins Gemin2, Gemin3, and Gemin5 are components of U bodies

    SciTech Connect

    Cauchi, Ruben J.; Sanchez-Pulido, Luis; Liu, Ji-Long

    2010-08-15

    Uridine-rich small nuclear ribonucleoproteins (U snRNPs) play key roles in pre-mRNA processing in the nucleus. The assembly of most U snRNPs takes place in the cytoplasm and is facilitated by the survival motor neuron (SMN) complex. Discrete cytoplasmic RNA granules called U bodies have been proposed to be specific sites for snRNP assembly because they contain U snRNPs and SMN. U bodies invariably associate with P bodies, which are involved in mRNA decay and translational control. However, it remains unknown whether other SMN complex proteins also localise to U bodies. In Drosophila there are four SMN complex proteins, namely SMN, Gemin2/CG10419, Gemin3 and Gemin5/Rigor mortis. Drosophila Gemin3 was originally identified as the Drosophila orthologue of human and yeast Dhh1, a component of P bodies. Through an in silico analysis of the DEAD-box RNA helicases we confirmed that Gemin3 is the bona fide Drosophila orthologue of vertebrate Gemin3 whereas the Drosophila orthologue of Dhh1 is Me31B. We then made use of the Drosophila egg chamber as a model system to study the subcellular distribution of the Gemin proteins as well as Me31B. Our cytological investigations show that Gemin2, Gemin3 and Gemin5 colocalise with SMN in U bodies. Although they are excluded from P bodies, as components of U bodies, Gemin2, Gemin3 and Gemin5 are consistently found associated with P bodies, wherein Me31B resides. In addition to a role in snRNP biogenesis, SMN complexes residing in U bodies may also be involved in mRNP assembly and/or transport.

  14. Gemin4. A novel component of the SMN complex that is found in both gems and nucleoli.

    PubMed

    Charroux, B; Pellizzoni, L; Perkinson, R A; Yong, J; Shevchenko, A; Mann, M; Dreyfuss, G

    2000-03-20

    The survival of motor neurons (SMN) protein, the product of the neurodegenerative disease spinal muscular atrophy (SMA) gene, is localized both in the cytoplasm and in discrete nuclear bodies called gems. In both compartments SMN is part of a large complex that contains several proteins including Gemin2 (formerly SIP1) and the DEAD box protein Gemin3. In the cytoplasm, the SMN complex is associated with snRNP Sm core proteins and plays a critical role in spliceosomal snRNP assembly. In the nucleus, SMN is required for pre-mRNA splicing by serving in the regeneration of spliceosomes. These functions are likely impaired in cells of SMA patients because they have reduced levels of functional SMN. Here, we report the identification by nanoelectrospray mass spectrometry of a novel component of the SMN complex that we name Gemin4. Gemin4 is associated in vivo with the SMN complex through a direct interaction with Gemin3. The tight interaction of Gemin4 with Gemin3 suggests that it could serve as a cofactor of this DEAD box protein. Gemin4 also interacts directly with several of the Sm core proteins. Monoclonal antibodies against Gemin4 efficiently immunoprecipitate the spliceosomal U snRNAs U1 and U5 from Xenopus oocytes cytoplasm. Immunolocalization experiments show that Gemin4 is colocalized with SMN in the cytoplasm and in gems. Interestingly, Gemin4 is also detected in the nucleoli, suggesting that the SMN complex may also function in preribosomal RNA processing or ribosome assembly. PMID:10725331

  15. B and T cell immune response to small nuclear ribonucleoprotein particles in lupus mice: autoreactive CD4(+) T cells recognize a T cell epitope located within the RNP80 motif of the 70K protein.

    PubMed

    Monneaux, F; Briand, J P; Muller, S

    2000-08-01

    Systemic lupus erythematosus is characterized by the presence of high titers of autoantibodies reacting with various components of the U1 small nuclear ribonucleoprotein particle (snRNP). It has been suggested that these antibodies are produced by an antigen-driven mechanism under the dependence of antigen-specific T cells. To investigate the role of T cell help in this process, we sought, with 20 overlapping peptides, the Th epitopes on the U1-70K snRNP in unprimed H-2(k) MRL / lpr lupus mice and immunized CBA normal mice. The peptide 131 - 151 was recognized by both IgG autoantibodies and CD4(+) T cells from 7 - 9-week-old MRL / lpr mice. In this test, antigen-presenting cells (APC) from MRL / lpr mice were required; APC from naive CBA mice failed to stimulate CD4(+) cells from MRL / lpr mice. The potential role of MRL / lpr B cells as APC, the expression of MHC class II molecules at their surface and their activation state (expression of CD69, CD80 / B7-1 and CD86 / B7-2 molecules) were studied. Peptide 131 - 151 bound both I-A(k) and I-E(k) class II molecules and favored an IL-2-positive T cell response but not IFN-gamma, IL-6 and IL-10 secretion. Segment 131 - 151 is localized within the RNP80 motif and contains residues that are highly conserved in many nuclear, nucleolar and cytoplasmic RNA binding proteins.

  16. Effect of aging on mechanical properties of aluminum-alloy rivets

    NASA Technical Reports Server (NTRS)

    Roop, Frederick C

    1941-01-01

    Curves and tabular data present the results of strength tests made during and after 2 1/2 years of aging on rivets and rivet wire of 3/16-inch nominal diameter. The specimens were of aluminum alloy: 24S, 17S, and A17S of the duralumin type and 53S of the magnesium-silicide type.

  17. A breast-ovarian cancer susceptibility gene maps to chromosome 17q21

    SciTech Connect

    Feunteun, J. ); Narod, S.A.; Parboosingh, J. ); Lynch, H.T.; Watson, P.; Conway, T.; Lynch, J. ); O'Connell, P.; White, R. ); Lenoir, G.M. )

    1993-04-01

    Nineteen North American Caucasian families that contain a minimum of four confirmed cases of breast or ovarian cancer have been studied. Four polymorphisms (cLB17.1, D17S579, D17S588, and D17S74), which span a region of approximately 15 cM on chromosome 17q12, were typed. The data confirm the location of a dominant gene conferring susceptibility to breast and ovarian cancer (maximum lod = 9.78) and suggest that the breast-ovarian cancer syndrome is genetically heterogeneous. Two recombinants in one large family suggest that the breast-ovarian cancer locus lies between D17S588 and D17S579. 14 refs., 3 figs., 3 tabs.

  18. Priming by Rhizobacterium Protects Tomato Plants from Biotrophic and Necrotrophic Pathogen Infections through Multiple Defense Mechanisms

    PubMed Central

    Ahn, Il-Pyung; Lee, Sang-Woo; Kim, Min Gab; Park, Sang-Ryeol; Hwang, Duk-Ju; Bae, Shin-Chul

    2011-01-01

    A selected strain of rhizobacterium, Pseudomonas putida strain LSW17S (LSW17S), protects tomato plants (Lycopersicon esculentum L. cv. Seokwang) from bacterial speck by biotrophic Pseudomonas syringae pv. tomato strain DC3000 (DC3000) and bacterial wilt by necrotrophic Ralstonia solanacearum KACC 10703 (Rs10703). To investigate defense mechanisms induced by LSW17S in tomato plants, transcription patterns of pathogenesis-related (PR) genes and H2O2 production were analyzed in plants treated with LSW17S and subsequent pathogen inoculation. LSW17S alone did not induce transcriptions of employed PR genes in leaves and roots. DC3000 challenge following LSW17S triggered rapid transcriptions of PR genes and H2O2 production in leaves and roots. Catalase infiltration with DC3000 attenuated defense-related responses and resistance against DC3000 infection. Despite depriving H2O2 production and PR1b transcription by the same treatment, resistance against Rs10703 infection was not deterred significantly. H2O2 is indispensable for defense signaling and/or mechanisms primed by LSW17S and inhibition of bacterial speck, however, it is not involved in resistance against bacterial wilt. PMID:21710203

  19. Caesium diuranium hexa-telluride.

    PubMed

    Mesbah, Adel; Ibers, James A

    2012-10-01

    Single crystals of CsU(2)Te(6) were synthesized from the reaction of U, Te, and Cs(2)Te(3) at 1273 K. CsU(2)Te(6) crystallizes in the space group Cmcm in the CsTh(2)Te(6) structure type. The asymmetric unit comprises one U (site symmetry m2m), one Cs (m2m; half-occupancy) and two Te atoms (m.. and m2m). The structure of CsU(2)Te(6) consists of infinite [U(2)Te(6)] layers perpendicular to [010] separated by Cs atoms. There are infinite Te-Te-Te linear chains along [001].

  20. Modeling Sorption and Degradation of 17β-Estradiol-17-Sulfate in Agricultural Soils

    NASA Astrophysics Data System (ADS)

    Bai, X.; Casey, F. X.; Hakk, H.; Shrestha, S. L.; DeSutter, T.; Khan, E.; Oduor, P. G.

    2011-12-01

    The natural steroid hormone, 17β-estradiol (E2), can be an endocrine disruptor at part-per trillion levels. Laboratory studies indicate a low potential for E2 persistence and mobility in the environment; however, field studies consistently indicate the presence of E2 and its primary metabolite, estrone, at levels sufficiently high to impact water quality. To facilitate urine excretion, animals may release E2 as a sulfated conjugate, which would have a higher aqueous solubility than the parent compound. We hypothesize that E2 conjugates contribute to the detection of free estrogens in the environment. The objective of this study was to determine the sorption, degradation, and mobility of a model conjugate, 17β-estradiol-17-sulfate (E2-17S), in agricultural soils. Radiolabeled E2-17S ([14C]E2-17S) was chemically synthesized in a three-step process, and then batch experiments were conducted in natural and sterile soils. Additionally, soil organic carbon (OC) was varied (1.29 and 0.26%) to investigate its effect on the fate of [14C]E2-17S. Liquid scintillation counting (LSC) was used in concert with high performance liquid chromatography (HPLC) to detect and quantitate parent compound and metabolites of E2-17S in the aqueous and bound phases. Residual soil was combusted to determine non-extractable levels of 14C. The E2-17S was relatively stable in the aqueous phase for natural and sterile soils. Mono- and di- hydroxyl E2-17S were detected as metabolites of E2-17S in the aqueous phase above both sterile and natural soil. Deconjugation to form E2 was not observed in aqueous phase; however, E2 and estrone were extracted from both natural and sterile soils. A conceptual model was developed to simulate and identify the fate and transport processes of E2-17S. Organic carbon was found to be an important factor affecting the sorption and degradation of E2-17S in soils.

  1. Members of a family of proteins (the RD family) detected by a U1 70K monoclonal antibody are present in spliceosomal complexes.

    PubMed Central

    Staknis, D; Reed, R

    1995-01-01

    We have characterized a monoclonal antibody (mAb) to the U1 snRNP component U1 70K. We find that this antibody recognizes several proteins, in addition to U1 70K, in purified spliceosomal complexes and in total HeLa cell nuclear extract preparations. The novel mAb U1 70K antigens can also be specifically immunoprecipitated by the antibody. Similarly to U1 70K, many of the mAb U1 70K antigens can be phosphorylated by a co-purifying kinase activity. The epitope recognized by mAb U1 70K was previously shown to be a repeating arginine/aspartate (RD) dipeptide. Thus we have designated the novel mAb U1 70K antigens the RD family. Comparison of mAb U1 70K with a recently characterized antibody, mAb 16H3, whose epitope is a repeating R/D or R/E motif, showed that a large subset of the antigens are common. In contrast, most of the mAb U1 70K antigens are distinct from the proteins detected by mAb 104, an antibody to the SR family of splicing factors. Images PMID:7479068

  2. Molecular Basis and Therapeutic Strategies to Rescue Factor IX Variants That Affect Splicing and Protein Function.

    PubMed

    Tajnik, Mojca; Rogalska, Malgorzata Ewa; Bussani, Erica; Barbon, Elena; Balestra, Dario; Pinotti, Mirko; Pagani, Franco

    2016-05-01

    Mutations that result in amino acid changes can affect both pre-mRNA splicing and protein function. Understanding the combined effect is essential for correct diagnosis and for establishing the most appropriate therapeutic strategy at the molecular level. We have identified a series of disease-causing splicing mutations in coagulation factor IX (FIX) exon 5 that are completely recovered by a modified U1snRNP particle, through an SRSF2-dependent enhancement mechanism. We discovered that synonymous mutations and missense substitutions associated to a partial FIX secretion defect represent targets for this therapy as the resulting spliced-corrected proteins maintains normal FIX coagulant specific activity. Thus, splicing and protein alterations contribute to define at the molecular level the disease-causing effect of a number of exonic mutations in coagulation FIX exon 5. In addition, our results have a significant impact in the development of splicing-switching therapies in particular for mutations that affect both splicing and protein function where increasing the amount of a correctly spliced protein can circumvent the basic functional defects. PMID:27227676

  3. Global Analysis of Chlorella variabilis NC64A mRNA Profiles during the Early Phase of Paramecium bursaria Chlorella Virus-1 Infection

    PubMed Central

    Rowe, Janet M.; Jeanniard, Adrien; Gurnon, James R.; Xia, Yuannan; Dunigan, David D.; Van Etten, James L.; Blanc, Guillaume

    2014-01-01

    The PBCV-1/Chlorella variabilis NC64A system is a model for studies on interactions between viruses and algae. Here we present the first global analyses of algal host transcripts during the early stages of infection, prior to virus replication. During the course of the experiment stretching over 1 hour, about a third of the host genes displayed significant changes in normalized mRNA abundance that either increased or decreased compared to uninfected levels. The population of genes with significant transcriptional changes gradually increased until stabilizing at 40 minutes post infection. Functional categories including cytoplasmic ribosomal proteins, jasmonic acid biosynthesis and anaphase promoting complex/cyclosomes had a significant excess in upregulated genes, whereas spliceosomal snRNP complexes and the shikimate pathway had significantly more down-regulated genes, suggesting that these pathways were activated or shut-down in response to the virus infection. Lastly, we examined the expression of C. varibilis RNA polymerase subunits, as PBCV-1 transcription depends on host RNA polymerases. Two subunits were up-regulated, RPB10 and RPC34, suggesting that they may function to support virus transcription. These results highlight genes and pathways, as well as overall trends, for further refinement of our understanding of the changes that take place during the early stages of viral infection. PMID:24608695

  4. Association of HLA-DRB1 alleles with susceptibility to mixed connective tissue disease in Polish patients.

    PubMed

    Paradowska-Gorycka, A; Stypińska, B; Olesińska, M; Felis-Giemza, A; Mańczak, M; Czuszynska, Z; Zdrojewski, Z; Wojciechowicz, J; Jurkowska, M

    2016-01-01

    Mixed connective tissue disease (MCTD) is a systemic autoimmune disease, originally defined as a connective tissue inflammatory syndrome with overlapping features of systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), polymyositis/dermatomyositis (PM/DM) and systemic sclerosis (SSc), characterized by the presence of antibodies against components of the U1 small nuclear ribonucleoprotein (U1snRNP). The aim of the study was to assess the frequency of (high-resolution-typed) DRB1 alleles in a cohort of Polish patients with MCTD (n = 103). Identification of the variants potentially associated with risk and protection was carried out by comparison with the DKMS Polish Bone Marrow Donor Registry (41306 alleles). DRB1*15:01 (odds ratio (OR): 6.06; 95% confidence interval (CI) 4.55-8.06), DRB1*04 (OR: 3.69; 95% CI 2.69-5.01) and *09:01 (OR: 8.12; 95% CI 2.15-21.75) were identified as risk alleles for MCTD, while HLA-DRB1*07:01 allele was found to be protective (OR: 0.50; 95% CI 0.28-0.83). The carrier frequency of the DRB1*01 was higher in MCTD patients compared with controls, although the differences were not statistically significant. Our results confirm the modulating influence of HLA-DRB1 genotypes on development of connective tissue diseases such as MCTD.

  5. Pathogenesis of Systemic Lupus Erythematosus Revisited 2011: End Organ Resistance to Damage, Autoantibody Initiation and Diversification, and HLA-DR

    PubMed Central

    Fu, Shu Man; Deshmukh, Umesh S; Gaskin, Felicia

    2011-01-01

    Systemic lupus erythematosus (SLE) is a multi-system disorder resulting from interaction of susceptibility genes and environmental factors. SLE has protean clinical presentations at the initial diagnosis and relapses. SLE-related autoantibodies have unique patterns of diversification to linked proteins such as the snRNP particle and the diversification takes years before clinical diagnosis. There are both clinical and experimental evidence to indicate that separate genes contribute to autoimmunity and end organ damage and these genes are independent and interactive. Among the numerous susceptibility genes, HLA-D complex is dominant. Results from the authors’ laboratories led us to postulate a unified hypothesis for SLE pathogenesis. This hypothesis states that SLE-autoantibodies are initiated by environmental T cell epitope mimics of the SLE-related autoantigens in hosts with susceptible HLA-D alleles. These autoantibodies diversify over a period of years due the accumulation of cross-reactive T cells. This process ultimately leads to the generation of organ specific autoantibodies and autoreactive effector T cells due to the polyreactive nature of T and B cell receptors from hosts with susceptibility genes to end organ damage, resulting in protean clinical presentations. This hypothesis accounts for most of the features unique to SLE and has clinical implications as to how patients should be treated. PMID:21632208

  6. Origins of De Novo Genes in Human and Chimpanzee

    PubMed Central

    Ruiz-Orera, Jorge; Hernandez-Rodriguez, Jessica; Chiva, Cristina; Sabidó, Eduard; Kondova, Ivanela; Bontrop, Ronald; Marqués-Bonet, Tomàs; Albà, M.Mar

    2015-01-01

    The birth of new genes is an important motor of evolutionary innovation. Whereas many new genes arise by gene duplication, others originate at genomic regions that did not contain any genes or gene copies. Some of these newly expressed genes may acquire coding or non-coding functions and be preserved by natural selection. However, it is yet unclear which is the prevalence and underlying mechanisms of de novo gene emergence. In order to obtain a comprehensive view of this process, we have performed in-depth sequencing of the transcriptomes of four mammalian species—human, chimpanzee, macaque, and mouse—and subsequently compared the assembled transcripts and the corresponding syntenic genomic regions. This has resulted in the identification of over five thousand new multiexonic transcriptional events in human and/or chimpanzee that are not observed in the rest of species. Using comparative genomics, we show that the expression of these transcripts is associated with the gain of regulatory motifs upstream of the transcription start site (TSS) and of U1 snRNP sites downstream of the TSS. In general, these transcripts show little evidence of purifying selection, suggesting that many of them are not functional. However, we find signatures of selection in a subset of de novo genes which have evidence of protein translation. Taken together, the data support a model in which frequently-occurring new transcriptional events in the genome provide the raw material for the evolution of new proteins. PMID:26720152

  7. U bodies respond to nutrient stress in Drosophila

    SciTech Connect

    Buckingham, Mickey; Liu, Ji-Long

    2011-12-10

    The neurodegenerative disease spinal muscular atrophy (SMA) is caused by mutation of the survival motor neuron 1 (SMN1) gene. Cytoplasmic SMN protein-containing granules, known as U snRNP bodies (U bodies), are thought to be responsible for the assembly and storage of small nuclear ribonucleoproteins (snRNPs) which are essential for pre-mRNA splicing. U bodies exhibit close association with cytoplasmic processing bodies (P bodies), which are involved in mRNA decay and translational repression. The close association of the U body and P body in Drosophila resemble that of the stress granule and P body in yeast and mammalian cells. However, it is unknown whether the U body is responsive to any stress. Using Drosophila oogenesis as a model, here we show that U bodies increase in size following nutritional deprivation. Despite nutritional stress, U bodies maintain their close association with P bodies. Our results show that U bodies are responsive to nutrition changes, presumably through the U body-P body pathway.

  8. [The role of RNA splicing in the pathogenesis of spinal muscular atrophy and development of its therapeutics].

    PubMed

    Sahashi, Kentaro; Sobue, Gen

    2014-12-01

    Loss-of-function mutations in SMN1 cause spinal muscular atrophy (SMA), a leading genetic cause of infant mortality. Degeneration of alpha-motor neurons that results in progressive paralysis is a pathological hallmark of SMA. Recently, peripheral-tissue involvement has also been reported in SMA. Patients have low levels of functional SMN which is attributed to alternative splicing in SMN2, a gene closely-related to SMN1. This decrease in the expression of SMN, a ubiquitously expressed protein involved in promoting snRNP assembly required for splicing, is responsible for SMA. However, the mechanism through which decrease in SMN levels causes SMA remains unclear. Currently, no curative treatment is available for SMA, but SMN restoration is thought to be necessary and sufficient for cure. Antisense oligonucleotides (ASOs) can be designed to specifically alter splicing patterns of target pre-mRNAs. We identified an ASO that redirects SMN2 splicing and is currently in clinical trials for use as RNA-targeting therapeutics. Further, we have also reported a novel application of splicing-modulating ASOs--creation of animal phenocopy models of diseases by inducing mis-splicing. Exploring the relationship between the spatial and temporal effects of therapeutic and pathogenic ASOs yields relevant insights into the roles of SMN in SMA pathogenesis and into its normal physiological functions. This knowledge, in turn, contributes to the ongoing development of targeted therapeutics.

  9. Origins of De Novo Genes in Human and Chimpanzee.

    PubMed

    Ruiz-Orera, Jorge; Hernandez-Rodriguez, Jessica; Chiva, Cristina; Sabidó, Eduard; Kondova, Ivanela; Bontrop, Ronald; Marqués-Bonet, Tomàs; Albà, M Mar

    2015-12-01

    The birth of new genes is an important motor of evolutionary innovation. Whereas many new genes arise by gene duplication, others originate at genomic regions that did not contain any genes or gene copies. Some of these newly expressed genes may acquire coding or non-coding functions and be preserved by natural selection. However, it is yet unclear which is the prevalence and underlying mechanisms of de novo gene emergence. In order to obtain a comprehensive view of this process, we have performed in-depth sequencing of the transcriptomes of four mammalian species--human, chimpanzee, macaque, and mouse--and subsequently compared the assembled transcripts and the corresponding syntenic genomic regions. This has resulted in the identification of over five thousand new multiexonic transcriptional events in human and/or chimpanzee that are not observed in the rest of species. Using comparative genomics, we show that the expression of these transcripts is associated with the gain of regulatory motifs upstream of the transcription start site (TSS) and of U1 snRNP sites downstream of the TSS. In general, these transcripts show little evidence of purifying selection, suggesting that many of them are not functional. However, we find signatures of selection in a subset of de novo genes which have evidence of protein translation. Taken together, the data support a model in which frequently-occurring new transcriptional events in the genome provide the raw material for the evolution of new proteins.

  10. Cloning of mDEAH9, a putative RNA helicase and mammalian homologue of Saccharomyces cerevisiae splicing factor Prp43.

    PubMed

    Gee, S; Krauss, S W; Miller, E; Aoyagi, K; Arenas, J; Conboy, J G

    1997-10-28

    Yeast splicing factor Prp43, a DEAH box protein of the putative RNA helicase/RNA-dependent NTPase family, is a splicing factor that functions late in the pre-mRNA splicing pathway to facilitate spliceosome disassembly. In this paper we report cDNA cloning and characterization of mDEAH9, an apparent mammalian homologue of Prp43. Amino acid sequence comparison revealed that the two proteins are approximately 65% identical over a 500-aa region spanning the central helicase domain and the C-terminal region. Expression of mDEAH9 in S. cerevisiae bearing a temperature-sensitive mutation in prp43 was sufficient to restore growth at the nonpermissive temperature. This functional complementation was specific, as mouse mDEAH9 failed to complement mutations in related splicing factor genes prp16 or prp22. Finally, double label immunofluorescence experiments performed with mammalian cells revealed colocalization of mDEAH9 and splicing factor SC35 in punctate nuclear speckles. Thus, the hypothesis that mDEAH9 represents the mammalian homologue of yeast Prp43 is supported by its high sequence homology, functional complementation, and colocalization with a known splicing factor in the nucleus. Our results provide additional support for the hypothesis that the spliceosomal machinery that mediates regulated, dynamic changes in conformation of pre-mRNA and snRNP RNAs has been highly conserved through evolution.

  11. NUFIP and the HSP90/R2TP chaperone bind the SMN complex and facilitate assembly of U4-specific proteins.

    PubMed

    Bizarro, Jonathan; Dodré, Maxime; Huttin, Alexandra; Charpentier, Bruno; Schlotter, Florence; Branlant, Christiane; Verheggen, Céline; Massenet, Séverine; Bertrand, Edouard

    2015-10-15

    The Sm proteins are loaded on snRNAs by the SMN complex, but how snRNP-specific proteins are assembled remains poorly characterized. U4 snRNP and box C/D snoRNPs have structural similarities. They both contain the 15.5K and proteins with NOP domains (PRP31 for U4, NOP56/58 for snoRNPs). Biogenesis of box C/D snoRNPs involves NUFIP and the HSP90/R2TP chaperone system and here, we explore the function of this machinery in U4 RNP assembly. We show that yeast Prp31 interacts with several components of the NUFIP/R2TP machinery, and that these interactions are separable from each other. In human cells, PRP31 mutants that fail to stably associate with U4 snRNA still interact with components of the NUFIP/R2TP system, indicating that these interactions precede binding of PRP31 to U4 snRNA. Knock-down of NUFIP leads to mislocalization of PRP31 and decreased association with U4. Moreover, NUFIP is associated with the SMN complex through direct interactions with Gemin3 and Gemin6. Altogether, our data suggest a model in which the NUFIP/R2TP system is connected with the SMN complex and facilitates assembly of U4 snRNP-specific proteins. PMID:26275778

  12. Splicing-Mediated Autoregulation Modulates Rpl22p Expression in Saccharomyces cerevisiae

    PubMed Central

    Gabunilas, Jason

    2016-01-01

    In Saccharomyces cerevisiae, splicing is critical for expression of ribosomal protein genes (RPGs), which are among the most highly expressed genes and are tightly regulated according to growth and environmental conditions. However, knowledge of the precise mechanisms by which RPG pre-mRNA splicing is regulated on a gene-by-gene basis is lacking. Here we show that Rpl22p has an extraribosomal role in the inhibition of splicing of the RPL22B pre-mRNA transcript. A stem loop secondary structure within the intron is necessary for pre-mRNA binding by Rpl22p in vivo and splicing inhibition in vivo and in vitro and can rescue splicing inhibition in vitro when added in trans to splicing reactions. Splicing inhibition by Rpl22p may be partly attributed to the reduction of co-transcriptional U1 snRNP recruitment to the pre-mRNA at the RPL22B locus. We further demonstrate that the inhibition of RPL22B pre-mRNA splicing contributes to the down-regulation of mature transcript during specific stress conditions, and provide evidence hinting at a regulatory role for this mechanism in conditions of suppressed ribosome biogenesis. These results demonstrate an autoregulatory mechanism that fine-tunes the expression of the Rpl22 protein and by extension Rpl22p paralog composition according to the cellular demands for ribosome biogenesis. PMID:27097027

  13. Supraspliceosomes at defined functional states portray the pre-assembled nature of the pre-mRNA processing machine in the cell nucleus.

    PubMed

    Kotzer-Nevo, Hani; de Lima Alves, Flavia; Rappsilber, Juri; Sperling, Joseph; Sperling, Ruth

    2014-06-30

    When isolated from mammalian cell nuclei, all nuclear pre-mRNAs are packaged in multi-subunit large ribonucleoprotein complexes-supraspliceosomes-composed of four native spliceosomes interconnected by the pre-mRNA. Supraspliceosomes contain all five spliceosomal U snRNPs, together with other splicing factors, and are functional in splicing. Supraspliceosomes studied thus far represent the steady-state population of nuclear pre-mRNAs that were isolated at different stages of the splicing reaction. To analyze specific splicing complexes, here, we affinity purified Pseudomonas aeruginosa phage 7 (PP7)-tagged splicing complexes assembled in vivo on Adenovirus Major Late (AdML) transcripts at specific functional stages, and characterized them using molecular techniques including mass spectrometry. First, we show that these affinity purified splicing complexes assembled on PP7-tagged AdML mRNA or on PP7-tagged AdML pre-mRNA are assembled in supraspliceosomes. Second, similar to the general population of supraspliceosomes, these defined supraspliceosomes populations are assembled with all five U snRNPs at all splicing stages. This study shows that dynamic changes in base-pairing interactions of U snRNA:U snRNA and U snRNA:pre-mRNA that occur in vivo during the splicing reaction do not require changes in U snRNP composition of the supraspliceosome. Furthermore, there is no need to reassemble a native spliceosome for the splicing of each intron, and rearrangements of the interactions will suffice.

  14. Nuclear localization and in vivo dynamics of a plant-specific serine/arginine-rich protein.

    PubMed

    Ali, Gul Shad; Golovkin, Maxim; Reddy, Anireddy S N

    2003-12-01

    Serine/arginine-rich (SR) proteins in non-plant systems are known to play important roles in both constitutive and alternative splicing of pre-messenger RNAs (pre-mRNAs). Recently, we isolated a novel SR protein (SR45), which interacts with U1 snRNP 70K protein, a key protein involved in 5' splice site recognition. SR45 is found only in plants and is unique in having two SR domains separated by an RNA recognition motif (RRM). To study the localization and dynamics of SR45, we expressed it as a fusion to green fluorescent protein (GFP) in cultured cells and transgenic Arabidopsis plants. The SR45 is localized exclusively to nuclei. In interphase nuclei, GFP-SR45 was found both in speckles and nucleoplasm. The speckles exhibited intranuclear movements and changes in morphology. Inhibition of transcription and protein phosphorylation resulted in redistribution of SR45 to bigger speckles. The change in the number and morphology of speckles caused by inhibition of transcription was blocked by an inhibitor of phosphatases. These results indicate that transcription activity of the cell and protein (de)phosphorylation regulate the intranuclear distribution of SR45. PMID:14675452

  15. The spliceosomal phosphopeptide P140 controls the lupus disease by interacting with the HSC70 protein and via a mechanism mediated by gammadelta T cells.

    PubMed

    Page, Nicolas; Schall, Nicolas; Strub, Jean-Marc; Quinternet, Marc; Chaloin, Olivier; Décossas, Marion; Cung, Manh Thong; Van Dorsselaer, Alain; Briand, Jean-Paul; Muller, Sylviane

    2009-01-01

    The phosphopeptide P140 issued from the spliceosomal U1-70K snRNP protein is recognized by lupus CD4(+) T cells, transiently abolishes T cell reactivity to other spliceosomal peptides in P140-treated MRL/lpr mice, and ameliorates their clinical features. P140 modulates lupus patients' T cell response ex vivo and is currently included in phase IIb clinical trials. Its underlying mechanism of action remains elusive. Here we show that P140 peptide binds a unique cell-surface receptor, the constitutively-expressed chaperone HSC70 protein, known as a presenting-protein. P140 induces apoptosis of activated MRL/lpr CD4(+) T cells. In P140-treated mice, it increases peripheral blood lymphocyte apoptosis and decreases B cell, activated T cell, and CD4(-)CD8(-)B220(+) T cell counts via a specific mechanism strictly depending on gammadelta T cells. Expression of inflammation-linked genes is rapidly regulated in CD4(+) T cells. This work led us to identify a powerful pathway taken by a newly-designed therapeutic peptide to immunomodulate lupus autoimmunity.

  16. 3'-end-dependent formation of U6 small nuclear ribonucleoprotein particles in Xenopus laevis oocyte nuclei.

    PubMed

    Terns, M P; Lund, E; Dahlberg, J E

    1992-07-01

    We have identified and characterized a U6 small nuclear (sn) ribonucleoprotein particle (RNP) present in the nuclei of Xenopus laevis oocytes. The structure of this U6 snRNP was investigated by native gel shift analysis and a combination of RNA-protein UV cross-linking, RNase T1 fingerprinting, and immunoprecipitation assays. These analyses demonstrate that certain forms of U6 snRNA associate with the 50-kDa nuclear antigen La both in vivo and in vitro. The La protein binds the stretch of uridylates at the 3' hydroxyl end of newly synthesized U6 snRNA. La does not bind to mature U6 snRNAs that have 2',3'-cyclic phosphate (greater than p) groups at their 3' ends (E. Lund and J. E. Dahlberg, Science 255:327-330, 1992) or to U6 snRNAs in anti-Sm-precipitable U4/U6 snRNPs. We propose that 3'-end modification, including posttranscriptional UMP addition, modulates the binding of La protein to U6 snRNA which, in turn, may affect the function of this RNA. PMID:1535684

  17. Splicing-Mediated Autoregulation Modulates Rpl22p Expression in Saccharomyces cerevisiae.

    PubMed

    Gabunilas, Jason; Chanfreau, Guillaume

    2016-04-01

    In Saccharomyces cerevisiae, splicing is critical for expression of ribosomal protein genes (RPGs), which are among the most highly expressed genes and are tightly regulated according to growth and environmental conditions. However, knowledge of the precise mechanisms by which RPG pre-mRNA splicing is regulated on a gene-by-gene basis is lacking. Here we show that Rpl22p has an extraribosomal role in the inhibition of splicing of the RPL22B pre-mRNA transcript. A stem loop secondary structure within the intron is necessary for pre-mRNA binding by Rpl22p in vivo and splicing inhibition in vivo and in vitro and can rescue splicing inhibition in vitro when added in trans to splicing reactions. Splicing inhibition by Rpl22p may be partly attributed to the reduction of co-transcriptional U1 snRNP recruitment to the pre-mRNA at the RPL22B locus. We further demonstrate that the inhibition of RPL22B pre-mRNA splicing contributes to the down-regulation of mature transcript during specific stress conditions, and provide evidence hinting at a regulatory role for this mechanism in conditions of suppressed ribosome biogenesis. These results demonstrate an autoregulatory mechanism that fine-tunes the expression of the Rpl22 protein and by extension Rpl22p paralog composition according to the cellular demands for ribosome biogenesis. PMID:27097027

  18. Coilin association with Box C/D scaRNA suggests a direct role for the Cajal body marker protein in scaRNP biogenesis

    PubMed Central

    Enwerem, Isioma I.; Velma, Venkatramreddy; Broome, Hanna J.; Kuna, Marija; Begum, Rowshan A.; Hebert, Michael D.

    2014-01-01

    ABSTRACT Spliceosomal small nuclear ribonucleoproteins (snRNPs) are enriched in the Cajal body (CB). Guide RNAs, known as small Cajal body-specific RNAs (scaRNAs), direct modification of the small nuclear RNA (snRNA) component of the snRNP. The protein WRAP53 binds a sequence motif (the CAB box) found in many scaRNAs and the RNA component of telomerase (hTR) and targets these RNAs to the CB. We have previously reported that coilin, the CB marker protein, associates with certain non-coding RNAs. For a more comprehensive examination of the RNAs associated with coilin, we have sequenced the RNA isolated from coilin immunocomplexes. A striking preferential association of coilin with the box C/D scaRNAs 2 and 9, which lack a CAB box, was observed. This association varied by treatment condition and WRAP53 knockdown. In contrast, reduction of WRAP53 did not alter the level of coilin association with hTR. Additional studies showed that coilin degrades/processes scaRNA 2 and 9, associates with active telomerase and can influence telomerase activity. These findings suggest that coilin plays a novel role in the biogenesis of box C/D scaRNPs and telomerase. PMID:24659245

  19. Association of HLA-DRB1 alleles with susceptibility to mixed connective tissue disease in Polish patients.

    PubMed

    Paradowska-Gorycka, A; Stypińska, B; Olesińska, M; Felis-Giemza, A; Mańczak, M; Czuszynska, Z; Zdrojewski, Z; Wojciechowicz, J; Jurkowska, M

    2016-01-01

    Mixed connective tissue disease (MCTD) is a systemic autoimmune disease, originally defined as a connective tissue inflammatory syndrome with overlapping features of systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), polymyositis/dermatomyositis (PM/DM) and systemic sclerosis (SSc), characterized by the presence of antibodies against components of the U1 small nuclear ribonucleoprotein (U1snRNP). The aim of the study was to assess the frequency of (high-resolution-typed) DRB1 alleles in a cohort of Polish patients with MCTD (n = 103). Identification of the variants potentially associated with risk and protection was carried out by comparison with the DKMS Polish Bone Marrow Donor Registry (41306 alleles). DRB1*15:01 (odds ratio (OR): 6.06; 95% confidence interval (CI) 4.55-8.06), DRB1*04 (OR: 3.69; 95% CI 2.69-5.01) and *09:01 (OR: 8.12; 95% CI 2.15-21.75) were identified as risk alleles for MCTD, while HLA-DRB1*07:01 allele was found to be protective (OR: 0.50; 95% CI 0.28-0.83). The carrier frequency of the DRB1*01 was higher in MCTD patients compared with controls, although the differences were not statistically significant. Our results confirm the modulating influence of HLA-DRB1 genotypes on development of connective tissue diseases such as MCTD. PMID:26818120

  20. The reciprocal regulation between splicing and 3′‐end processing

    PubMed Central

    2016-01-01

    Most eukaryotic precursor mRNAs are subjected to RNA processing events, including 5′‐end capping, splicing and 3′‐end processing. These processing events were historically studied independently; however, since the early 1990s tremendous efforts by many research groups have revealed that these processing factors interact with each other to control each other's functions. U1 snRNP and its components negatively regulate polyadenylation of precursor mRNAs. Importantly, this function is necessary for protecting the integrity of the transcriptome and for regulating gene length and the direction of transcription. In addition, physical and functional interactions occur between splicing factors and 3′‐end processing factors across the last exon. These interactions activate or inhibit splicing and 3′‐end processing depending on the context. Therefore, splicing and 3′‐end processing are reciprocally regulated in many ways through the complex protein–protein interaction network. Although interesting questions remain, future studies will illuminate the molecular mechanisms underlying the reciprocal regulation. WIREs RNA 2016, 7:499–511. doi: 10.1002/wrna.1348 For further resources related to this article, please visit the WIREs website. PMID:27019070

  1. GEMINs: potential therapeutic targets for spinal muscular atrophy?

    PubMed Central

    Borg, Rebecca; Cauchi, Ruben J.

    2014-01-01

    The motor neuron degenerative disease spinal muscular atrophy (SMA) remains one of the most frequently inherited causes of infant mortality. Afflicted patients loose the survival motor neuron 1 (SMN1) gene but retain one or more copies of SMN2, a homolog that is incorrectly spliced. Primary treatment strategies for SMA aim at boosting SMN protein levels, which are insufficient in patients. SMN is known to partner with a set of diverse proteins collectively known as GEMINs to form a macromolecular complex. The SMN-GEMINs complex is indispensible for chaperoning the assembly of small nuclear ribonucleoproteins (snRNPs), which are key for pre-mRNA splicing. Pharmaceutics that alleviate the neuromuscular phenotype by restoring the fundamental function of SMN without augmenting its levels are also crucial in the development of an effective treatment. Their use as an adjunct therapy is predicted to enhance benefit to patients. Inspired by the surprising discovery revealing a premier role for GEMINs in snRNP biogenesis together with in vivo studies documenting their requirement for the correct function of the motor system, this review speculates on whether GEMINs constitute valid targets for SMA therapeutic development. PMID:25360080

  2. Dual function of C/D box small nucleolar RNAs in rRNA modification and alternative pre-mRNA splicing.

    PubMed

    Falaleeva, Marina; Pages, Amadis; Matuszek, Zaneta; Hidmi, Sana; Agranat-Tamir, Lily; Korotkov, Konstantin; Nevo, Yuval; Eyras, Eduardo; Sperling, Ruth; Stamm, Stefan

    2016-03-22

    C/D box small nucleolar RNAs (SNORDs) are small noncoding RNAs, and their best-understood function is to target the methyltransferase fibrillarin to rRNA (for example, SNORD27 performs 2'-O-methylation of A27 in 18S rRNA). Unexpectedly, we found a subset of SNORDs, including SNORD27, in soluble nuclear extract made under native conditions, where fibrillarin was not detected, indicating that a fraction of the SNORD27 RNA likely forms a protein complex different from canonical snoRNAs found in the insoluble nuclear fraction. As part of this previously unidentified complex,SNORD27 regulates the alternative splicing of the transcription factor E2F7p re-mRNA through direct RNA-RNA interaction without methylating the RNA, likely by competing with U1 small nuclear ribonucleoprotein (snRNP). Furthermore, knockdown of SNORD27 activates previously "silent" exons in several other genes through base complementarity across the entire SNORD27 sequence, not just the antisense boxes. Thus, some SNORDs likely function in both rRNA and pre-mRNA processing, which increases the repertoire of splicing regulators and links both processes. PMID:26957605

  3. Global analysis of Chlorella variabilis NC64A mRNA profiles during the early phase of Paramecium bursaria chlorella virus-1 infection.

    PubMed

    Rowe, Janet M; Jeanniard, Adrien; Gurnon, James R; Xia, Yuannan; Dunigan, David D; Van Etten, James L; Blanc, Guillaume

    2014-01-01

    The PBCV-1/Chlorella variabilis NC64A system is a model for studies on interactions between viruses and algae. Here we present the first global analyses of algal host transcripts during the early stages of infection, prior to virus replication. During the course of the experiment stretching over 1 hour, about a third of the host genes displayed significant changes in normalized mRNA abundance that either increased or decreased compared to uninfected levels. The population of genes with significant transcriptional changes gradually increased until stabilizing at 40 minutes post infection. Functional categories including cytoplasmic ribosomal proteins, jasmonic acid biosynthesis and anaphase promoting complex/cyclosomes had a significant excess in upregulated genes, whereas spliceosomal snRNP complexes and the shikimate pathway had significantly more down-regulated genes, suggesting that these pathways were activated or shut-down in response to the virus infection. Lastly, we examined the expression of C. varibilis RNA polymerase subunits, as PBCV-1 transcription depends on host RNA polymerases. Two subunits were up-regulated, RPB10 and RPC34, suggesting that they may function to support virus transcription. These results highlight genes and pathways, as well as overall trends, for further refinement of our understanding of the changes that take place during the early stages of viral infection.

  4. Screening of genes related to sulfide metabolism in Urechis unicinctus (Echiura, Urechidae) using suppression subtractive hybridization and cDNA microarray analysis.

    PubMed

    Shi, Xiaoli; Shao, Mingyu; Zhang, Litao; Ma, Yubin; Zhang, Zhifeng

    2012-09-01

    Exogenous sulfide can generally induce metabolic injuries in most organisms and even cause death. However, organisms inhabiting intertidal zones, hydrothermal vents, and cold seeps, can tolerate, metabolize, and utilize sulfide. In this study, both suppression subtractive hybridization and cDNA microarray analysis were employed to screen sulfide metabolism-related genes from the body wall in echiuran worm Urechis unicinctus, a marine sediment species. A total of 3456 monoclones were isolated and 82 were identified as differentially expressed genes in worms exposed to 50 μM sulfide for 24 h, compared to controls. The identified genes encoded proteins with multiple processes, including metabolism, cellular process, biological regulation, response to stimulus, multicellular organismal process, localization, development, and cellular component organization. Eight genes, serase, vacuolar protein, src tyrosine kinase, sulfide oxidase-like oxidoreductase, aprataxin, SN-RNP, aminopeptidase, and predicted protein, were selected to verify expression in the worm using qRT-PCR. The agreement of gene expression evaluation was 62.5% between the results of microarray analysis and qRT-PCR. These new data will provide clues for further probing of the molecular mechanism of sulfide metabolism. PMID:22591583

  5. Profiling of Epstein-Barr virus latent RNA expression in clinical specimens by gene-specific multiprimed cDNA synthesis and PCR.

    PubMed

    Stevens, Servi J C; Brink, Antoinette A T P; Middeldorp, Jaap M

    2005-01-01

    We describe a two-step RT-PCR method for simultaneous detection of EBNA-1 (QK and Y3K splice variants), EBNA-2, LMP-1, LMP-2a and -2b, ZEBRA, and BARTs RNA encoded by Epstein-Barr virus. As a control for RNA integrity, the low-copy-number transcript derived from U1A snRNP, a cellular housekeeping gene, is coamplified. Copy DNA (cDNA) for these nine targets is simultaneously synthesized in a gene-specific, multiprimed cDNA reaction, which strongly reduces the amount of required clinical specimen and allows more sensitive detection than random hexamer or oligo-dT priming. For amplification, cDNA synthesis is followed by nine separate PCRs for the mentioned targets. Primers were designed either as intron-flanking, to avoid background DNA amplification, or in different exons, allowing identification of differentially spliced RNA molecules. To increase specificity, PCR products are detected by autoradiography after hybridization with radiolabeled internal oligonucleotide probes. The method described is highly suitable for profiling EBV latent RNA expression in tissue biopsies, cultured or isolated cells, and unfractionated whole blood and for definition of EBV latency type I, II, or III gene expression in these samples.

  6. The spliceosomal PRP19 complex of trypanosomes

    PubMed Central

    Ambrósio, Daniela L.; Badjatia, Nitika; Günzl, Arthur

    2015-01-01

    Summary In trypanosomes, mRNAs are processed by spliced leader (SL) trans splicing, in which a capped SL, derived from SL RNA, is spliced onto the 5′ end of each mRNA. This process is mediated by the spliceosome, a large and dynamic RNA-protein machinery consisting of small nuclear ribonucleoproteins (snRNPs) and non-snRNP proteins. Due to early evolutionary divergence, the amino acid sequences of trypanosome splicing factors exhibit limited similarity to those of their eukaryotic orthologs making their bioinformatic identification challenging. Most of the ~ 60 protein components that have been characterized thus far are snRNP proteins because, in contrast to individual snRNPs, purification of intact spliceosomes has not been achieved yet. Here, we characterize the non-snRNP PRP19 complex of Trypanosoma brucei. We identified a complex that contained the core subunits PRP19, CDC5, PRL1, and SPF27, as well as PRP17, SKIP and PPIL1. Three of these proteins were newly annotated. The PRP19 complex was associated primarily with the activated spliceosome and, accordingly, SPF27 silencing blocked the first splicing step. Interestingly, SPF27 silencing caused an accumulation of SL RNA with a hypomethylated cap that closely resembled the defect observed previously upon depletion of the cyclin-dependent kinase CRK9, indicating that both proteins may function in spliceosome activation. PMID:25524563

  7. Gemin3 Is an Essential Gene Required for Larval Motor Function and Pupation in Drosophila

    PubMed Central

    Shpargel, Karl B.; Praveen, Kavita; Rajendra, T. K.

    2009-01-01

    The assembly of metazoan Sm-class small nuclear ribonucleoproteins (snRNPs) is an elaborate, step-wise process that takes place in multiple subcellular compartments. The initial steps, including formation of the core RNP, are mediated by the survival motor neuron (SMN) protein complex. Loss-of-function mutations in human SMN1 result in a neuromuscular disease called spinal muscular atrophy. The SMN complex is comprised of SMN and a number of tightly associated proteins, collectively called Gemins. In this report, we identify and characterize the fruitfly ortholog of the DEAD box protein, Gemin3. Drosophila Gemin3 (dGem3) colocalizes and interacts with dSMN in vitro and in vivo. RNA interference for dGem3 codepletes dSMN and inhibits efficient Sm core assembly in vitro. Transposon insertion mutations in Gemin3 are larval lethals and also codeplete dSMN. Transgenic overexpression of dGem3 rescues lethality, but overexpression of dSMN does not, indicating that loss of dSMN is not the primary cause of death. Gemin3 mutant larvae exhibit motor defects similar to previously characterized Smn alleles. Remarkably, appreciable numbers of Gemin3 mutants (along with one previously undescribed Smn allele) survive as larvae for several weeks without pupating. Our results demonstrate the conservation of Gemin3 protein function in metazoan snRNP assembly and reveal that loss of either Smn or Gemin3 can contribute to neuromuscular dysfunction. PMID:18923150

  8. Molecular Basis and Therapeutic Strategies to Rescue Factor IX Variants That Affect Splicing and Protein Function

    PubMed Central

    Bussani, Erica; Barbon, Elena; Pinotti, Mirko; Pagani, Franco

    2016-01-01

    Mutations that result in amino acid changes can affect both pre-mRNA splicing and protein function. Understanding the combined effect is essential for correct diagnosis and for establishing the most appropriate therapeutic strategy at the molecular level. We have identified a series of disease-causing splicing mutations in coagulation factor IX (FIX) exon 5 that are completely recovered by a modified U1snRNP particle, through an SRSF2-dependent enhancement mechanism. We discovered that synonymous mutations and missense substitutions associated to a partial FIX secretion defect represent targets for this therapy as the resulting spliced-corrected proteins maintains normal FIX coagulant specific activity. Thus, splicing and protein alterations contribute to define at the molecular level the disease-causing effect of a number of exonic mutations in coagulation FIX exon 5. In addition, our results have a significant impact in the development of splicing-switching therapies in particular for mutations that affect both splicing and protein function where increasing the amount of a correctly spliced protein can circumvent the basic functional defects. PMID:27227676

  9. Dual function of C/D box small nucleolar RNAs in rRNA modification and alternative pre-mRNA splicing

    PubMed Central

    Falaleeva, Marina; Pages, Amadis; Matuszek, Zaneta; Hidmi, Sana; Agranat-Tamir, Lily; Korotkov, Konstantin; Nevo, Yuval; Sperling, Ruth; Stamm, Stefan

    2016-01-01

    C/D box small nucleolar RNAs (SNORDs) are small noncoding RNAs, and their best-understood function is to target the methyltransferase fibrillarin to rRNA (for example, SNORD27 performs 2′-O-methylation of A27 in 18S rRNA). Unexpectedly, we found a subset of SNORDs, including SNORD27, in soluble nuclear extract made under native conditions, where fibrillarin was not detected, indicating that a fraction of the SNORD27 RNA likely forms a protein complex different from canonical snoRNAs found in the insoluble nuclear fraction. As part of this previously unidentified complex, SNORD27 regulates the alternative splicing of the transcription factor E2F7 pre-mRNA through direct RNA–RNA interaction without methylating the RNA, likely by competing with U1 small nuclear ribonucleoprotein (snRNP). Furthermore, knockdown of SNORD27 activates previously “silent” exons in several other genes through base complementarity across the entire SNORD27 sequence, not just the antisense boxes. Thus, some SNORDs likely function in both rRNA and pre-mRNA processing, which increases the repertoire of splicing regulators and links both processes. PMID:26957605

  10. Non-coding RNAs, the cutting edge of histone messages

    PubMed Central

    Köhn, Marcel; Hüttelmaier, Stefan

    2016-01-01

    ABSTRACT In metazoan the 3′-end processing of histone mRNAs is a conserved process involving the concerted action of many protein factors and the non-coding U7 snRNA. Recently, we identified that the processing of histone pre-mRNAs is promoted by an additional ncRNA, the Y3-derived Y3** RNA. U7 modulates the association of the U7 snRNP whereas Y3** promotes recruitment of CPSF (cleavage and polyadenylation specific factor) proteins to nascent histone transcripts at histone locus bodies (HLBs) in mammals. This enhances the 3′-end cleavage of nascent histone pre-mRNAs and modulates HLB assembly. Here we discuss new insights in the role of ncRNAs in the spatiotemporal control of histone synthesis. We propose that ncRNAs scaffold the formation of functional protein-RNA complexes and their sequential deposition on nascent histone pre-mRNAs at HLBs. These findings add to the multiple roles of ncRNAs in controlling gene expression and may provide new avenues for targeting histone synthesis in cancer. PMID:26909464

  11. The Retroviruses Human Immunodeficiency Virus Type 1 and Moloney Murine Leukemia Virus Adopt Radically Different Strategies To Regulate Promoter-Proximal Polyadenylation

    PubMed Central

    Furger, Andre; Monks, Joan; Proudfoot, Nick J.

    2001-01-01

    Maximal gene expression in retroviruses requires that polyadenylation in the 5′ long terminal repeat (LTR) is suppressed. In human immunodeficiency virus type 1 (HIV-1) the promoter-proximal poly(A) site is blocked by interaction of U1 snRNP with the closely positioned major splice donor site (MSD) 200 nucleotides downstream. Here we investigated whether the same mechanism applies to down-regulate 5′ LTR polyadenylation in Moloney murine leukemia virus (MoMLV). Although the same molecular architecture is present in both viruses, the MoMLV poly(A) signal in the 5′ LTR is active whether or not the MSD is mutated. This surprising difference between the two retroviruses is not due to their actual poly(A) signals or MSD sequences, since exchange of either element between the two viral sequences does not alter their ability to regulate 5′ LTR poly(A) site use. Instead we demonstrate that sequence between the cap and AAUAAA is required for MSD-dependent poly(A) regulation in HIV-1, indicating a key role for this part of the LTR in poly(A) site suppression. We also show that the MoMLV poly(A) signal is an intrinsically weak RNA-processing signal. This suggests that in the absence of a poly(A) site suppression mechanism, MoMLV is forced to use a weak poly(A) signal. PMID:11689654

  12. hnRNP H binding at the 5′ splice site correlates with the pathological effect of two intronic mutations in the NF-1 and TSHβ genes

    PubMed Central

    Buratti, Emanuele; Baralle, Marco; De Conti, Laura; Baralle, Diana; Romano, Maurizio; Ayala, Youhna M.; Baralle, Francisco E.

    2004-01-01

    We have recently reported a disease-causing substitution (+5G > C) at the donor site of NF-1 exon 3 that produces its skipping. We have now studied in detail the splicing mechanism involved in analyzing RNA–protein complexes at several 5′ splice sites. Characteristic protein patterns were observed by pulldown and band-shift/super-shift analysis. Here, we show that hnRNP H binds specifically to the wild-type GGGgu donor sequence of the NF-1 exon 3. Depletion analyses shows that this protein restricts the accessibility of U1 small nuclear ribonucleoprotein (U1snRNA) to the donor site. In this context, the +5G > C mutation abolishes both U1snRNP base pairing and the 5′ splice site (5′ss) function. However, exon recognition in the mutant can be rescued by disrupting the binding of hnRNP H, demonstrating that this protein enhances the effects of the +5G > C substitution. Significantly, a similar situation was found for a second disease-causing +5G > A substitution in the 5′ss of TSHβ exon 2, which harbors a GGgu donor sequence. Thus, the reason why similar nucleotide substitutions can be either neutral or very disruptive of splicing function can be explained by the presence of specific binding signatures depending on local contexts. PMID:15299088

  13. Splicing diversity revealed by reduced spliceosomes in C. merolae and other organisms

    PubMed Central

    Hudson, Andrew J; Stark, Martha R; Fast, Naomi M; Russell, Anthony G; Rader, Stephen D

    2015-01-01

    Pre-mRNA splicing has been considered one of the hallmarks of eukaryotes, yet its diversity is astonishing: the number of substrate introns for splicing ranges from hundreds of thousands in humans to a mere handful in certain parasites. The catalytic machinery that carries out splicing, the spliceosome, is similarly diverse, with over 300 associated proteins in humans to a few tens in other organisms. In this Point of View, we discuss recent work characterizing the reduced spliceosome of the acidophilic red alga Cyanidioschyzon merolae, which further highlights the diversity of splicing in that it does not possess the U1 snRNP that is characteristically responsible for 5′ splice site recognition. Comparisons to other organisms with reduced spliceosomes, such as microsporidia, trypanosomes, and Giardia, help to identify the most highly conserved splicing factors, pointing to the essential core of this complex machine. These observations argue for increased exploration of important biochemical processes through study of a wider ranger of organisms. PMID:26400738

  14. cpsf1 is required for definitive HSC survival in zebrafish

    PubMed Central

    Payne, Elspeth M.; Rhodes, Jennifer; Gjini, Evisa; Johnston, Adam B.; Guo, Feng; Lee, Jeong-Soo; Stewart, Rodney A.; Kanki, John P.; Chen, Aye T.; Zhou, Yi; Zon, Leonard I.

    2011-01-01

    A comprehensive understanding of the genes and pathways regulating hematopoiesis is needed to identify genes causally related to bone marrow failure syndromes, myelodysplastic syndromes, and hematopoietic neoplasms. To identify novel genes involved in hematopoiesis, we performed an ethyl-nitrosourea mutagenesis screen in zebrafish (Danio rerio) to search for mutants with defective definitive hematopoiesis. We report the recovery and analysis of the grechetto mutant, which harbors an inactivating mutation in cleavage and polyadenylation specificity factor 1 (cpsf1), a gene ubiquitously expressed and required for 3′ untranslated region processing of a subset of pre-mRNAs. grechetto mutants undergo normal primitive hematopoiesis and specify appropriate numbers of definitive HSCs at 36 hours postfertilization. However, when HSCs migrate to the caudal hematopoietic tissue at 3 days postfertilization, their numbers start decreasing as a result of apoptotic cell death. Consistent with Cpsf1 function, c-myb:EGFP+ cells in grechetto mutants also show defective polyadenylation of snrnp70, a gene required for HSC development. By 5 days postfertilization, definitive hematopoiesis is compromised and severely decreased blood cell numbers are observed across the myeloid, erythroid, and lymphoid cell lineages. These studies show that cpsf1 is essential for HSC survival and differentiation in caudal hematopoietic tissue. PMID:21330472

  15. In vivo disruption of Xenopus U3 snRNA affects ribosomal RNA processing.

    PubMed Central

    Savino, R; Gerbi, S A

    1990-01-01

    DNA oligonucleotide complementary to sequences in the 5' third of U3 snRNA were injected into Xenopus oocyte nuclei to disrupt endogenous U3 snRNA. The effect of this treatment on rRNA processing was examined. We found that some toads have a single rRNA processing pathway, whereas in other toads, two rRNA processing pathways can coexist in a single oocyte. U3 snRNA disruption in toads with the single rRNA processing pathway caused a reduction in 20S and '32S' pre-rRNA. In addition, in toads with two rRNA processing pathways, an increase in '36S' pre-rRNA of the second pathway is observed. This is the first in vivo demonstration that U3 snRNA plays a role in rRNA processing. Cleavage site #3 is at the boundary of ITS 1 and 5.8S and links all of the affected rRNA intermediates: 20S and '32S' are the products of site #3 cleavage in the first pathway and '36S' is the substrate for cleavage at site #3 in the second pathway. We postulate that U3 snRNP folds pre-rRNA into a conformation dictating correct cleavage at processing site #3. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 5. Fig. 6. Fig. 7. Fig. 8. PMID:2357971

  16. Extensive interactions of PRP8 protein with the 5' and 3' splice sites during splicing suggest a role in stabilization of exon alignment by U5 snRNA.

    PubMed Central

    Teigelkamp, S; Newman, A J; Beggs, J D

    1995-01-01

    Precursor RNAs containing 4-thiouridine at specific sites were used with UV-crosslinking to map the binding sites of the yeast protein splicing factor PRP8. PRP8 protein interacts with a region of at least eight exon nucleotides at the 5' splice site and a minimum of 13 exon nucleotides and part of the polypyrimidine tract in the 3' splice site region. Crosslinking of PRP8 to mutant and duplicated 3' splice sites indicated that the interaction is not sequence specific, nor does it depend on the splice site being functional. Binding of PRP8 to the 5' exon was established before step 1 and to the 3' splice site region after step 1 of splicing. These interactions place PRP8 close to the proposed catalytic core of the spliceosome during both transesterification reactions. To date, this represents the most extensive mapping of the binding site(s) of a splicing factor on the substrate RNA. We propose that the large binding sites of PRP8 stabilize the intrinsically weaker interactions of U5 snRNA with both exons at the splice sites for exon alignment by the U5 snRNP. Images PMID:7781612

  17. Vent Processes and Deposits of a Hiatus in a Violent Eruption: Quilotoa Volcano, Ecuador

    NASA Astrophysics Data System (ADS)

    Best, J. A.; Bustillos, J.; Ort, M. H.; Cashman, K. V.; Mothes, P. A.; di Muro, A.; Rosi, M.

    2010-12-01

    The 800 BP eruption of Quilotoa volcano, Ecuador, produced two plinian eruptions separated by a short (days-weeks) hiatus. Units 1 and 3 (U1 and U3) of the eruption correspond to the first and second Plinian eruptions, respectively, and produced fallout and pyroclastic density currents. Unit 2 (U2) records processes during the hiatus and consists of three subunits: U2a, U2b, and U2c. 147 tephra samples of U1, U2, and U3 were collected from 25 sites from around the volcano. Thickness and grain-size features were described, with particular attention paid to U2, in order to characterize the processes that occurred during the eruptive hiatus. Grain-size and componentry analysis of a subset of these samples reveals a number of trends. The upper part of U1 is massive and normally graded at its top, 32-45 % dominantly vitric ash ≤ 3.0 φ, and likely represents the clearing of the air at the end of the first plinian eruption. U2a, present out to a maximum of 7 km from the vent, has a polymodal distribution with a large fraction of 4.0 φ and finer vitric material. Dune forms occur in this unit, which are interpreted to be the product of surges. The areal distribution of U2a is constrained by topography, whereas U2b is not. U2b is coarser overall with alternating fine- (2-3φ) and coarse- (1-2φ) grained layers. The beds, both coarse and fine, have a near-bimodal grain-size distribution and normal grading. U2b is interpreted as a fall deposit. The U2a/U2b contact is gradational in that 0-2 beds of U2b material occur within the uppermost U2a beds at proximal localities, indicating vent conditions for both briefly coexisted. U2c is a <2-cm-thick vitric ash bed with sparse dense juvenile vitric lapilli. These lapilli also occur in the overlying basal U3 fallout, which has a polymodal grain-size distribution. U2b is characteristically orange in color due to the dust that loosely covers the grains. Hydrothermal activity within the vent is likely the source of this staining

  18. 51. Photographic copy of the original construction drawing, 192627, by ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    51. Photographic copy of the original construction drawing, 1926-27, by Harrington, Howard, and Ash, Consulting Engineers, from microfilm copy at Bridge Division, Missouri Highway and Transportation Department, Jefferson City, Missouri. 671 FT. SPAN, JOINTS L0, U2 TO U4, PORTAL AT U2 - Cape Girardeau Bridge, Spanning Mississippi River at State Highway 146, Cape Girardeau, Cape Girardeau County, MO

  19. 40 CFR 98.214 - Monitoring and QA/QC requirements.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... requirements. (a) The annual mass of carbonate consumed (for Equation U-1 of this subpart) or carbonate inputs (for Equation U-2 of this subpart) must be determined annually from monthly measurements using the same... weigh hoppers or weigh belt feeders. (b) The annual mass of carbonate outputs (for Equation U-2 of...

  20. 40 CFR 98.214 - Monitoring and QA/QC requirements.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... requirements. (a) The annual mass of carbonate consumed (for Equation U-1 of this subpart) or carbonate inputs (for Equation U-2 of this subpart) must be determined annually from monthly measurements using the same... weigh hoppers or weigh belt feeders. (b) The annual mass of carbonate outputs (for Equation U-2 of...

  1. 77 FR 37265 - Lending Limits

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-06-21

    ... to the use of external credit ratings. See 77 FR 35253 (June 13, 2012). The interim final rule also...)(1) of the Home Owners' Loan Act (HOLA), 12 U.S.C. 1464(u)(1), provides that section 5200 of the... applies to ] national banks.'' In addition, section 5(u)(2) of HOLA, 12 U.S.C. 1464(u)(2),...

  2. Genetic homogeneity in Sjoegren-Larsson syndrome: Linkage to chromosome 17p in families of different non-Swedish ethnic origins

    SciTech Connect

    Rogers, G.R.; Lee, M.; Compton, J.G.

    1995-11-01

    Sjoegren-Larsson syndrome (SLS) is a rare, autosomal recessive disorder that is characterized by congenital ichthyosis, mental retardation, and spastic diplegia or tetraplegia. Three United States families, three Egyptian families, and one Israeli Arab family were investigated for linkage of the SLS gene to a region of chromosome 17. Pairwise and multipoint linkage analysis with nine markers mapped the SLS gene to the same region of the genome as that reported in Swedish SLS pedigrees. Examination of recombinants by haplotype analysis showed that the gene lies in the region containing the markers D17S953, D17S805, D17S689, and D17S842. D17S805 is pericentromeric on 17p. Patients in two consanguineous Egyptian families were homozygous at the nine marker loci tested, and another patient from a third family was homozygous for eight of the nine, suggesting that within each of these families the region of chromosome 17 carrying the SLS gene is identical by descent. Linkage of the SLS gene to chromosome 17p in families of Arabic, mixed European, Native American, and Swedish descent provides evidence for a single SLS locus and should prove useful for diagnosis and carrier detection in worldwide cases. 25 refs., 4 figs., 1 tab.

  3. A gene for autosomal dominant progressive cone dystrophy (CORD5) maps to chromosome 17p12-p13

    SciTech Connect

    Balciuniene, J.; Holmgren, G.; Forsman, K.

    1995-11-20

    Inherited retinal dystrophy is a common cause of visual impairment. Cone dystrophy affects the cone function and is manifested as progressive loss of the central vision, defective color vision, and photophobia. Linkage was demonstrated between progressive cone dystrophy (CORD5) and genetic markers on chromosome 17p12-p13 in a five-generation family. Multipoint analysis gave a maximum lod score of 7.72 at the marker D17S938. Recombinant haplotypes in the family suggest that the cone dystrophy locus is located in a 25-cM interval between the markers D17S926/D17S849 and D17S804/D17S945. Furthermore, one recombination was detected between the disease locus and a microsatellite marker in the candidate gene RCV1, encoding the retinal protein recoverin. Two additional candidate genes encoding retinal guanylate cyclase (GUC2D) and pigment epithelium-derived factor (PEDF) are located at 17p13.1. Moreover, loci for retinitis pigmentosa and Leber congenital amaurosis have been mapped to the same region. Identification of the cone dystrophy locus may be of importance not only for identifying functional genes in the cone system, but also for identifying genes for other retinal disorders. 34 refs., 3 figs., 2 tabs.

  4. Vent Processes and Deposits of a Hiatus in a Violent Eruption: Quilotoa Volcano, Ecuador

    NASA Astrophysics Data System (ADS)

    Best, J. A.; Bustillos, J.; Ort, M. H.; Cashman, K. V.; Mothes, P. A.; di Muro, A.; Rosi, M.

    2009-12-01

    The 800 BP eruption of Quilotoa volcano, Ecuador, produced two plinian eruptions separated by a short (days-weeks) hiatus. We examine the tephra produced both during this hiatus and erupted at the onset of the second Plinian eruption. Units 1 and 3 (U1 and U3) of the eruption correspond to the first and second Plinian eruptions, respectively, and produced fallout and pyroclastic density currents. Unit 2 (U2) records processes during the hiatus and consists of two subunits: U2a, a vitric ashfall, and U2b, a crystal and lithic-rich fallout. 130 individual tephra samples of U1, U2, and U3 were collected from 24 sites along three radial transects from the volcano in January 2009. Thickness and grain-size features were described, with particular attention paid to U2. Grain-size and componentry analysis of a subset of these samples reveals a number of trends. The upper part of U1 is massive and normally graded at its top. This part of U1 is dominantly vitric ash smaller than 3.0 φ and likely represents the clearing of the air at the end of the first plinian eruption. U2a has a polymodal distribution with a large fraction of 4.0 φ and finer vitric material. Dune forms occur in this unit, which is interpreted to be the product of surges. U2b is coarser overall with alternating fine- (2-3φ) and coarse- (1-2φ) grained layers. The beds have a unimodal grain-size distribution and normal grading. U2b is interpreted as a fall deposit. The U2a/U2b contact is gradational in that 0-2 beds of U2b material occur within the uppermost U2a beds, indicating vent conditions for both briefly coexisted. U2c is a <2-cm-thick vitric ash with sparse crystal-rich lava lapilli. These lapilli also occur in the overlying basal U3 fallout, which has a polymodal grain-size distribution. Some U2b pumice fragments and crystals are stained orange, which gives U2b its characteristic color. Stained grains are also present but rare in other units and may have been sourced from the conduit walls. The

  5. Triphosphate residues at the 5' ends of rRNA precursor and 5S RNA from Dictyostelium discoideum.

    PubMed Central

    Batts-Young, B; Lodish, H F

    1978-01-01

    We present here direct evidence for the preservation of a transcriptional initiation sequence in a eukaryotic rRNA precursor: the 5'-end group for precursor to 17S rRNA (p17S RNA) from Dictyostelium discoideum is identified as the triphosphate residue pppA-. We also show that mature 5S RNA form Dictyostelium bears a different triphosphate residue, pppG-. In contrast, we find no evidence for more than one phosphate at the 5' end of the 25S rRNA precursor (p25S RNA). These observations indicate that synthesis of the large ribosomal RNAs of Dictyostelium begins with the 5'-terminal sequence of the p17S RNA, and that 5S RNA transcription must be initiated independently, despite the close association of the 5S and rRNA coding segments. Images PMID:204930

  6. Autosomal dominant zonular cataract with sutural opacities localized to chromosome 17q11-12

    SciTech Connect

    Padma, T.; Ayyagari, R.; Murty, J.S.

    1995-10-01

    Congenital cataracts constitute a morphologically and genetically heterogeneous group of diseases that are a major cause of childhood blindness. Different loci for hereditary congenital cataracts have been mapped to chromosomes 1, 2, 16, and 17q24. We report linkage of a gene causing a unique form of autosomal dominant zonular cataracts with Y-sutural opacities to chromosome 17q11-12 in a three-generation family exhibiting a maximum lod score of 3.9 at D17S805. Multipoint analysis gave a Mod confidence interval of 17 cM. This interval is bounded by the markers D17S799 and D17S798, a region that would encompass a number of candidate genes including that coding for {Beta}A3/A1-crystallin. 30 refs., 2 figs., 1 tab.

  7. Ribosomal cistrons in higher plant cells. II. Sequence homology between the two mature rRNAs of sycamore cells and intracistronic reiteration. A DNA - rRNA hybridization study.

    PubMed

    Miassod, R; Cecchini, J P

    1976-01-01

    1. Uniformly labelled rRNA of sycamore cells has been annealed with homologous DNA. The fractions of DNA complementary to the 17S, or 26S, or 17S + 26S rRNAs are found to be 0.19%, 0.15% and 0.23%. They are not in the ratio of the molecular weight values (0.8, 1.2 and 2 - 10(6), respectively for the 17S, 26S and 17S + 26S rRNAs). This result is compatible with the large hybridization competition observed between the two rRNAs (53 and 72%) and with the shift-down of saturation curves when DNA is presaturated with unlabelled rRNA before the incubation with the other labelled rRNA. 2. Under the selected experimental procedure, the DNA - rRNA hybrids formed appear to be specific. Since there is an equal number of structural genes for the 17S and 26S rRNAs, these results mean the occurrence of a great sequence homology, strictly restricted to the two rRNAs. Homologous and specific sequences have been estimated to 0.1 and 0.7, or 0.85 and 0.35 million daltons, respectively in the 17S or 26S structural genes. 3. From the calculated lengths of homologous sequences, an intracistronic reiteration of some ribosomal sequences can be deduced. This internal reiteration is directly evidenced by the complex pattern of DNA - rRNA annealing curves. As demonstrated by base-composition analysis, the internal reiteration is heterogeneous and concerns both the homologous and specific sequences. In addition, the DNA saturation values allow the calculation of 4000 copies for the ribosomal cistron in the whole sycamore genome.

  8. Genomic characterization and expression analysis of five novel IL-17 genes in the Pacific oyster, Crassostrea gigas.

    PubMed

    Li, Jun; Zhang, Yang; Zhang, Yuehuan; Xiang, Zhiming; Tong, Ying; Qu, Fufa; Yu, Ziniu

    2014-10-01

    Interleukin-17 (IL-17) is a proinflammatory cytokine that plays an important role in clearing extracellular bacteria and contributes to the pathology of many autoimmune and allergic conditions. In the present study, five novel IL-17 homologs were identified by searching and analyzing the Pacific oyster genome. All six CgIL-17 members (including a previously reported homolog) contained four conserved cysteines that were used in the formation of disulfide bonds. Phylogenetic analysis showed that all invertebrate IL-17s were clustered into one group, implying that invertebrate IL-17s evolved from one common ancestral gene and subsequently diversified. All CgIL-17s shared the same genomic structure, containing two exons and one intron, except for the CgIL-17-3 and CgIL-17-5 genes, which each had only one exon. The expression pattern of the CgIL-17 genes was analyzed by qRT-PCR in a variety of tissues and at different developmental stages, and these genes were highly expressed in the gill and digestive gland tissues. Moreover, the expression of the CgIL-17 family genes was significantly up-regulated in hemocytes challenged with Pathogen-Associated Molecular Patterns (PAMPs). CgIL-17-3 had a strong response to lipopolysaccharide (LPS) and heat-killed Vibrio alginolyticus (HKVA) challenge, while CgIL-17-5 and CgIL-17-6 can be activated by peptidoglycan (PGN), but not by heat-killed Listeria monocytogenes (HKLM). The distinct, up-regulated transcript levels of the CgIL-17s in response to PAMPs challenge further indicate that CgIL-17s are likely to be significant components of immune responses by playing diversified roles in host defense in the Pacific oyster. These findings suggest that CgIL-17s are involved in innate immune responses and further supports their conserved function in mollusks immunity.

  9. A conserved intronic U1 snRNP-binding sequence promotes trans-splicing in Drosophila.

    PubMed

    Gao, Jun-Li; Fan, Yu-Jie; Wang, Xiu-Ye; Zhang, Yu; Pu, Jia; Li, Liang; Shao, Wei; Zhan, Shuai; Hao, Jianjiang; Xu, Yong-Zhen

    2015-04-01

    Unlike typical cis-splicing, trans-splicing joins exons from two separate transcripts to produce chimeric mRNA and has been detected in most eukaryotes. Trans-splicing in trypanosomes and nematodes has been characterized as a spliced leader RNA-facilitated reaction; in contrast, its mechanism in higher eukaryotes remains unclear. Here we investigate mod(mdg4), a classic trans-spliced gene in Drosophila, and report that two critical RNA sequences in the middle of the last 5' intron, TSA and TSB, promote trans-splicing of mod(mdg4). In TSA, a 13-nucleotide (nt) core motif is conserved across Drosophila species and is essential and sufficient for trans-splicing, which binds U1 small nuclear RNP (snRNP) through strong base-pairing with U1 snRNA. In TSB, a conserved secondary structure acts as an enhancer. Deletions of TSA and TSB using the CRISPR/Cas9 system result in developmental defects in flies. Although it is not clear how the 5' intron finds the 3' introns, compensatory changes in U1 snRNA rescue trans-splicing of TSA mutants, demonstrating that U1 recruitment is critical to promote trans-splicing in vivo. Furthermore, TSA core-like motifs are found in many other trans-spliced Drosophila genes, including lola. These findings represent a novel mechanism of trans-splicing, in which RNA motifs in the 5' intron are sufficient to bring separate transcripts into close proximity to promote trans-splicing. PMID:25838544

  10. A role for complexes of survival of motor neurons (SMN) protein with gemins and profilin in neurite-like cytoplasmic extensions of cultured nerve cells

    SciTech Connect

    Sharma, Aarti; Lambrechts, Anja; Le thi Hao; Le, Thanh T.; Sewry, Caroline A.; Ampe, Christophe; Burghes, Arthur H.M.; Morris, Glenn E. . E-mail: glenn.morris@rjah.nhs.uk

    2005-09-10

    Spinal muscular atrophy (SMA) is caused by reduced levels of SMN (survival of motor neurons protein) and consequent loss of motor neurons. SMN is involved in snRNP transport and nuclear RNA splicing, but axonal transport of SMN has also been shown to occur in motor neurons. SMN also binds to the small actin-binding protein, profilin. We now show that SMN and profilin II co-localise in the cytoplasm of differentiating rat PC12 cells and in neurite-like extensions, especially at their growth cones. Many components of known SMN complexes were also found in these extensions, including gemin2 (SIP-1), gemin6, gemin7 and unrip (unr-interacting protein). Coilin p80 and Sm core protein immunoreactivity, however, were seen only in the nucleus. SMN is known to associate with {beta}-actin mRNA and specific hnRNPs in axons and in neurite extensions of cultured nerve cells, and SMN also stimulates neurite outgrowth in cultures. Our results are therefore consistent with SMN complexes, rather than SMN alone, being involved in the transport of actin mRNPs along the axon as in the transport of snRNPs into the nucleus by similar SMN complexes. Antisense knockdown of profilin I and II isoforms inhibited neurite outgrowth of PC12 cells and caused accumulation of SMN and its associated proteins in cytoplasmic aggregates. BIAcore studies demonstrated a high affinity interaction of SMN with profilin IIa, the isoform present in developing neurons. Pathogenic missense mutations in SMN, or deletion of exons 5 and 7, prevented this interaction. The interaction is functional in that SMN can modulate actin polymerisation in vitro by reducing the inhibitory effect of profilin IIa. This suggests that reduced SMN in SMA might cause axonal pathfinding defects by disturbing the normal regulation of microfilament growth by profilins.

  11. Structure and interactions of the first three RNA recognition motifs of splicing factor Prp24

    PubMed Central

    Bae, Euiyoung; Reiter, Nicholas J.; Bingman, Craig A.; Kwan, Sharon S.; Lee, Donghan; Phillips, George N.; Butcher, Samuel E.; Brow, David A.

    2007-01-01

    Summary The essential Saccharomyces cerevisiae pre-messenger RNA splicing protein 24 (Prp24) has four RNA recognition motifs (RRMs) and facilitates U6 RNA base-pairing with U4 RNA during spliceosome assembly. Prp24 is a component of the free U6 small nuclear ribonucleoprotein particle (snRNP) but not the U4/U6 bi-snRNP, and so is thought to be displaced from U6 by U4/U6 base-pairing. The interaction partners of each of the four RRMs of Prp24 and how these interactions direct U4/U6 pairing are not known. Here we report the crystal structure of the first three RRMs and the solution structure of the first two RRMs of Prp24. Strikingly, RRM 2 forms extensive inter-domain contacts with RRMs 1 and 3. These contacts occupy much of the canonical RNA-binding faces (β-sheets) of RRMs 1 and 2, but leave the β-sheet of RRM 3 exposed. Previously identified substitutions in Prp24 that suppress mutations in U4 and U6 spliceosomal RNAs cluster primarily in the β-sheet of RRM 3, but also in a conserved loop of RRM 2. RNA binding assays and chemical shift mapping indicate that a large basic patch evident on the surface of RRMs 1 and 2 is part of a high affinity U6 RNA binding site. Our results suggest that Prp24 binds free U6 RNA primarily with RRMs 1 and 2, which may remodel the U6 secondary structure. The β-sheet of RRM 3 then influences U4/U6 pairing through interaction with an unidentified ligand. PMID:17320109

  12. Crystal structure, mutational analysis and RNA-dependent ATPase activity of the yeast DEAD-box pre-mRNA splicing factor Prp28

    SciTech Connect

    Jacewicz, Agata; Schwer, Beate; Smith, Paul; Shuman, Stewart

    2014-10-10

    Yeast Prp28 is a DEAD-box pre-mRNA splicing factor implicated in displacing U1 snRNP from the 5' splice site. Here we report that the 588-aa Prp28 protein consists of a trypsin-sensitive 126-aa N-terminal segment (of which aa 1–89 are dispensable for Prp28 function in vivo) fused to a trypsin-resistant C-terminal catalytic domain. Purified recombinant Prp28 and Prp28-(127–588) have an intrinsic RNA-dependent ATPase activity, albeit with a low turnover number. The crystal structure of Prp28-(127–588) comprises two RecA-like domains splayed widely apart. AMPPNP•Mg2+ is engaged by the proximal domain, with proper and specific contacts from Phe194 and Gln201 (Q motif) to the adenine nucleobase. The triphosphate moiety of AMPPNP•Mg2+ is not poised for catalysis in the open domain conformation. Guided by the Prp28•AMPPNP structure, and that of the Drosophila Vasa•AMPPNP•Mg2+•RNA complex, we targeted 20 positions in Prp28 for alanine scanning. ATP-site components Asp341 and Glu342 (motif II) and Arg527 and Arg530 (motif VI) and RNA-site constituent Arg476 (motif Va) are essential for Prp28 activity in vivo. Synthetic lethality of double-alanine mutations highlighted functionally redundant contacts in the ATP-binding (Phe194-Gln201, Gln201-Asp502) and RNA-binding (Arg264-Arg320) sites. As a result, overexpression of defective ATP-site mutants, but not defective RNA-site mutants, elicited severe dominant-negative growth defects.

  13. A conserved intronic U1 snRNP-binding sequence promotes trans-splicing in Drosophila.

    PubMed

    Gao, Jun-Li; Fan, Yu-Jie; Wang, Xiu-Ye; Zhang, Yu; Pu, Jia; Li, Liang; Shao, Wei; Zhan, Shuai; Hao, Jianjiang; Xu, Yong-Zhen

    2015-04-01

    Unlike typical cis-splicing, trans-splicing joins exons from two separate transcripts to produce chimeric mRNA and has been detected in most eukaryotes. Trans-splicing in trypanosomes and nematodes has been characterized as a spliced leader RNA-facilitated reaction; in contrast, its mechanism in higher eukaryotes remains unclear. Here we investigate mod(mdg4), a classic trans-spliced gene in Drosophila, and report that two critical RNA sequences in the middle of the last 5' intron, TSA and TSB, promote trans-splicing of mod(mdg4). In TSA, a 13-nucleotide (nt) core motif is conserved across Drosophila species and is essential and sufficient for trans-splicing, which binds U1 small nuclear RNP (snRNP) through strong base-pairing with U1 snRNA. In TSB, a conserved secondary structure acts as an enhancer. Deletions of TSA and TSB using the CRISPR/Cas9 system result in developmental defects in flies. Although it is not clear how the 5' intron finds the 3' introns, compensatory changes in U1 snRNA rescue trans-splicing of TSA mutants, demonstrating that U1 recruitment is critical to promote trans-splicing in vivo. Furthermore, TSA core-like motifs are found in many other trans-spliced Drosophila genes, including lola. These findings represent a novel mechanism of trans-splicing, in which RNA motifs in the 5' intron are sufficient to bring separate transcripts into close proximity to promote trans-splicing.

  14. Crystal structure, mutational analysis and RNA-dependent ATPase activity of the yeast DEAD-box pre-mRNA splicing factor Prp28

    DOE PAGES

    Jacewicz, Agata; Schwer, Beate; Smith, Paul; Shuman, Stewart

    2014-10-10

    Yeast Prp28 is a DEAD-box pre-mRNA splicing factor implicated in displacing U1 snRNP from the 5' splice site. Here we report that the 588-aa Prp28 protein consists of a trypsin-sensitive 126-aa N-terminal segment (of which aa 1–89 are dispensable for Prp28 function in vivo) fused to a trypsin-resistant C-terminal catalytic domain. Purified recombinant Prp28 and Prp28-(127–588) have an intrinsic RNA-dependent ATPase activity, albeit with a low turnover number. The crystal structure of Prp28-(127–588) comprises two RecA-like domains splayed widely apart. AMPPNP•Mg2+ is engaged by the proximal domain, with proper and specific contacts from Phe194 and Gln201 (Q motif) to themore » adenine nucleobase. The triphosphate moiety of AMPPNP•Mg2+ is not poised for catalysis in the open domain conformation. Guided by the Prp28•AMPPNP structure, and that of the Drosophila Vasa•AMPPNP•Mg2+•RNA complex, we targeted 20 positions in Prp28 for alanine scanning. ATP-site components Asp341 and Glu342 (motif II) and Arg527 and Arg530 (motif VI) and RNA-site constituent Arg476 (motif Va) are essential for Prp28 activity in vivo. Synthetic lethality of double-alanine mutations highlighted functionally redundant contacts in the ATP-binding (Phe194-Gln201, Gln201-Asp502) and RNA-binding (Arg264-Arg320) sites. As a result, overexpression of defective ATP-site mutants, but not defective RNA-site mutants, elicited severe dominant-negative growth defects.« less

  15. Primary structure of a human arginine-rich nuclear protein that colocalizes with spliceosome components

    SciTech Connect

    Chaudhary, N.; McMahon, C.; Blobel, G. )

    1991-09-15

    The cDNA for a 54-kDa nuclear protein (p54) has been cloned from a human hepatoma expression library. Contained within p54 is an arginine/serine-rich region similar to segments of several proteins that participate in pre-mRNA splicing including the 70-kDa component of U1 small nuclear ribonucleoprotein particle (snRNP) and the Drosophila transformer and suppressor-of-white-apricot proteins. The arginine/serine-rich region is dominated by a series of 8-amino acid imperfect repetitive motifs (consensus sequence, Arg-Arg-Ser-Arg-Ser-Arg-Ser-Arg). Antibodies raised against synthetic peptides of p54 react with an {approximately}70-kDa protein on immunoblots of HeLa cell and rat liver nuclear proteins. This apparent discrepancy in mass is also observed when p54 mRNA is translated in vitro. Indirect immunofluorescence studies in HeLa cells show that p54 is distributed throughout the nucleus in a speckled pattern, with an additional diffuse labeling of the nucleus excluding the nucleoli. Double immunofluorescence experiments indicate that these punctate regions are coincident with the speckles seen in cells stained with antibodies against several constituents of the pre-mRNA splicing machinery. Sedimentation analysis of HeLa cell extracts on sucrose gradients showed that p54 migrates at 4-6 S, indicating that the protein is not a tightly associated component of snRNPs. Although the function of p54 is not yet known, the structure and immunolocalization data suggest that this protein may have a role in pre-mRNA processing.

  16. Modulation of deregulated chaperone-mediated autophagy by a phosphopeptide

    PubMed Central

    Macri, Christophe; Wang, Fengjuan; Tasset, Inmaculada; Schall, Nicolas; Page, Nicolas; Briand, Jean-Paul; Cuervo, Ana Maria; Muller, Sylviane

    2015-01-01

    The P140 peptide, a 21-mer linear peptide (sequence 131–151) generated from the spliceosomal SNRNP70/U1–70K protein, contains a phosphoserine residue at position 140. It significantly ameliorates clinical manifestations in autoimmune patients with systemic lupus erythematosus and enhances survival in MRL/lpr lupus-prone mice. Previous studies showed that after P140 treatment, there is an accumulation of autophagy markers sequestosome 1/p62 and MAP1LC3-II in MRL/lpr B cells, consistent with a downregulation of autophagic flux. We now identify chaperone-mediated autophagy (CMA) as a target of P140 and demonstrate that its inhibitory effect on CMA is likely tied to its ability to alter the composition of HSPA8/HSC70 heterocomplexes. As in the case of HSPA8, expression of the limiting CMA component LAMP2A, which is increased in MRL/lpr B cells, is downregulated after P140 treatment. We also show that P140, but not the unphosphorylated peptide, uses the clathrin-dependent endo-lysosomal pathway to enter into MRL/lpr B lymphocytes and accumulates in the lysosomal lumen where it may directly hamper lysosomal HSPA8 chaperoning functions, and also destabilize LAMP2A in lysosomes as a result of its effect on HSP90AA1. This dual effect may interfere with the endogenous autoantigen processing and loading to major histocompatibility complex class II molecules and as a consequence, lead to lower activation of autoreactive T cells. These results shed light on mechanisms by which P140 can modulate lupus disease and exert its tolerogenic activity in patients. The unique selective inhibitory effect of the P140 peptide on CMA may be harnessed in other pathological conditions in which reduction of CMA activity would be desired. PMID:25719862

  17. Suppressors of the cdc-25.1(gf)-associated intestinal hyperplasia reveal important maternal roles for prp-8 and a subset of splicing factors in C. elegans

    PubMed Central

    Hebeisen, Michaël; Drysdale, John; Roy, Richard

    2008-01-01

    The maternal contribution of gene products enables embryos to initiate their developmental program in the absence of zygotic gene expression. In Caenorhabditis elegans, maternal CDC-25.1 levels are tightly regulated to promote early cell divisions, while stabilization of this phosphatase by gain-of-function mutations gives rise to intestinal-specific hyperplasia. To identify regulators of CDC-25.1 levels and/or function, we performed a modifier screen of the cdc-25.1(gf)-dependent hyperplasia. One of the isolated suppressor mutants possesses a donor splice site mutation in prp-8, a key splicing factor of the U5-specific snRNP. prp-8(rr40) produces aberrant prp-8 splice variants that generate C-terminal truncations at the expense of wild-type prp-8. Levels of maternal transcripts are reduced, including cdc-25.1, while zygotic transcripts appear unperturbed, suggesting a germ-line-specific role for this splicing factor in regulating the splicing, and consequently, the steady-state levels of maternal transcripts. Using a novel feeding RNAi strategy we found that only a subset of splicing factors suppress cdc-25.1(gf), suggesting that they too may play specific roles in germ-line spliceosome function. In humans, mutations in the corresponding hPrp8 C-terminal domain result in retinitis pigmentosa, a retinal-specific disorder. Intriguingly, despite affecting the general splicing apparatus, both human and C. elegans show tissue-specific defects resulting from mutations in this key splicing component. Our findings suggest that in addition to its important regulatory function in the C. elegans germ line, prp-8(rr40) may provide further insight into the etiology of this splicing-associated human disorder. PMID:18945809

  18. Assessment of the Frequency of Autoantibodies in Chronic Viral Hepatitis

    PubMed Central

    Acay, Akif; Demir, Kasim; Asik, Gulsah; Tunay, Havva; Acarturk, Gursel

    2015-01-01

    Objective: To examine the occurrence frequency of auto-antibodies and autoimmune diseases in patients with chronic hepatitis B or C. Methods: A total of 67 patients diagnosed with chronic hepatitis B and 77 patients diagnosed with chronic hepatitis C infection based on HBs Ag, Anti HCV, HBe Ag, Anti HBe Ag, HBV DNA, HCV RNA, liver ultrasound, and liver biopsy results as well as 48 healthy individuals were included in this study. ANA, anti dsDNA, anti LKM, Anti-SMA, AMA, C-ANCA, P-ANCA, anti-SSA, anti-SSB, anti-Scl-70, anti Jo-1, anti-U1snRNP, anti-centromere, anti-Jo-1, anti tpo, and anti tg were studied in all individuals in each study group. Results: ANA positivity was detected in 8 (12%), 15 (19%) and 2 (4%) individuals in HBV, HCV and control groups, respectively. The difference between the groups was significant (P=0.04). Similarly, anti Tg was positive in one subject in HBV group, in 6 subjects (7%) in HCV group, and in one subject among controls, the difference being significant (P=0.04). There were no significant differences between the study groups in the frequency of other auto-antibodies. Conclusion: Similar to studies involving patients who received interferon and/or antiviral agents, an increased frequency of auto-antibodies was also detected in our patient group consisting of interferon and anti-viral naive subjects. The increase in the frequency of auto-antibodies reached statistical significance among individuals with HCV infection. Thus, pre-treatment assessment of auto-antibodies in newly diagnosed cases of chronic hepatitis B or hepatitis C infection may provide beneficial information on the future occurrence of auto-immune responses in these patients. PMID:25878633

  19. DDX6 transfers P-TEFb kinase to the AF4/AF4N (AFF1) super elongation complex

    PubMed Central

    Mück, Fabian; Bracharz, Silvia; Marschalek, Rolf

    2016-01-01

    AF4/AFF1 and AF5/AFF4 are both backbones for the assembly of “super elongation complexes” (SECs) that exert 2 distinct functions after the recruitment of P-TEFb from the 7SK snRNP: (1) initiation and elongation of RNA polymerase II gene transcription, and (2) modification of transcribed gene regions by distinct histone methylation patterns. In this study we aimed to investigate one of the initial steps, namely how P-TEFb is transferred from 7SK snRNPs to the SECs. In particular, we were interested in the role of DDX6 that we have recently identified as part of the AF4 complex. DDX6 is an evolutionarily conserved member of the DEAD-box RNA helicase family that is known to control miRNA and mRNA biology (translation, storage and degradation). Overexpressed DDX6 is associated with different cancer types and with c-Myc protein overexpression. We could demonstrate that DDX6 binds to 7SK snRNA and causes the release and transfer of P-TEFb to the AF4/AF4N SEC. DDX6 also binds stably to AF4 and AF4N as demonstrated by GST pull-down and co-immunoprecipitation experiments. As a consequence, overexpression of either AF4/AF4N or DDX6 resulted in a strong increase of mRNA production (5-6 fold), while their simultaneous expression increased the cellular mRNA production by 11-fold. Conversely, the corresponding knockdown of DDX6 decreased mRNA production by 70%. In conclusion, AF4/AF4N and DDX6 represent key molecules for the elongation process of gene transcription and a model will be proposed for the hand-over process of P-TEFb to SECs. PMID:27679741

  20. The crystal structure of S. cerevisiae Sad1, a catalytically inactive deubiquitinase that is broadly required for pre-mRNA splicing

    PubMed Central

    Hadjivassiliou, Haralambos; Rosenberg, Oren S.; Guthrie, Christine

    2014-01-01

    Sad1 is an essential splicing factor initially identified in a genetic screen in Saccharomyces cerevisiae for snRNP assembly defects. Based on sequence homology, Sad1, or USP39 in humans, is predicted to comprise two domains: a zinc finger ubiquitin binding domain (ZnF-UBP) and an inactive ubiquitin-specific protease (iUSP) domain, both of which are well conserved. The role of these domains in splicing and their interaction with ubiquitin are unknown. We first used splicing microarrays to analyze Sad1 function in vivo and found that Sad1 is critical for the splicing of nearly all yeast intron-containing genes. By using in vitro assays, we then showed that it is required for the assembly of the active spliceosome. To gain structural insights into Sad1 function, we determined the crystal structure of the full-length protein at 1.8 Å resolution. In the structure, the iUSP domain forms the characteristic ubiquitin binding pocket, though with an amino acid substitution in the active site that results in complete inactivation of the enzymatic activity of the domain. The ZnF-UBP domain of Sad1 shares high structural similarly to other ZnF-UBPs; however, Sad1's ZnF-UBP does not possess the canonical ubiquitin binding motif. Given the precedents for ZnF-UBP domains to function as activators for their neighboring USP domains, we propose that Sad1's ZnF-UBP acts in a ubiquitin-independent capacity to recruit and/or activate Sad1's iUSP domain to interact with the spliceosome. PMID:24681967

  1. The large N-terminal region of the Brr2 RNA helicase guides productive spliceosome activation

    PubMed Central

    Absmeier, Eva; Wollenhaupt, Jan; Mozaffari-Jovin, Sina; Becke, Christian; Lee, Chung-Tien; Preussner, Marco; Heyd, Florian; Urlaub, Henning; Lührmann, Reinhard; Santos, Karine F.; Wahl, Markus C.

    2015-01-01

    The Brr2 helicase provides the key remodeling activity for spliceosome catalytic activation, during which it disrupts the U4/U6 di-snRNP (small nuclear RNA protein), and its activity has to be tightly regulated. Brr2 exhibits an unusual architecture, including an ∼500-residue N-terminal region, whose functions and molecular mechanisms are presently unknown, followed by a tandem array of structurally similar helicase units (cassettes), only the first of which is catalytically active. Here, we show by crystal structure analysis of full-length Brr2 in complex with a regulatory Jab1/MPN domain of the Prp8 protein and by cross-linking/mass spectrometry of isolated Brr2 that the Brr2 N-terminal region encompasses two folded domains and adjacent linear elements that clamp and interconnect the helicase cassettes. Stepwise N-terminal truncations led to yeast growth and splicing defects, reduced Brr2 association with U4/U6•U5 tri-snRNPs, and increased ATP-dependent disruption of the tri-snRNP, yielding U4/U6 di-snRNP and U5 snRNP. Trends in the RNA-binding, ATPase, and helicase activities of the Brr2 truncation variants are fully rationalized by the crystal structure, demonstrating that the N-terminal region autoinhibits Brr2 via substrate competition and conformational clamping. Our results reveal molecular mechanisms that prevent premature and unproductive tri-snRNP disruption and suggest novel principles of Brr2-dependent splicing regulation. PMID:26637280

  2. Splicing Factors Associate with Hyperphosphorylated RNA Polymerase II in the Absence of Pre-mRNA

    PubMed Central

    Kim, Euikyung; Du, Lei; Bregman, David B.; Warren, Stephen L.

    1997-01-01

    The carboxy-terminal domain (CTD) of the largest subunit of RNA polymerase II (Pol II) contains multiple tandem copies of the consensus heptapeptide, TyrSerProThrSerProSer. Concomitant with transcription initiation the CTD is phosphorylated. Elongating polymerase has a hyperphosphorylated CTD, but the role of this modification is poorly understood. A recent study revealed that some hyperphosphorylated polymerase molecules (Pol IIo) are nonchromosomal, and hence transcriptionally unengaged (Bregman, D.B., L. Du, S. van der Zee, S.L. Warren. 1995. J. Cell Biol. 129: 287–298). Pol IIo was concentrated in discrete splicing factor domains, suggesting a possible relationship between CTD phosphorylation and splicing factors, but no evidence beyond immunolocalization data was provided to support this idea. Here, we show that Pol IIo co-immunoprecipitates with members of two classes of splicing factors, the Sm snRNPs and non-snRNP SerArg (SR) family proteins. Significantly, Pol IIo's association with splicing factors is maintained in the absence of pre-mRNA, and the polymerase need not be transcriptionally engaged. We also provide definitive evidence that hyperphosphorylation of Pol II's CTD is poorly correlated with its transcriptional activity. Using monoclonal antibodies (mAbs) H5 and H14, which are shown here to recognize phosphoepitopes on Pol II's CTD, we have quantitated the level of Pol IIo at different stages of the cell cycle. The level of Pol IIo is similar in interphase and mitotic cells, which are transcriptionally active and inactive, respectively. Finally, complexes containing Pol IIo and splicing factors can be prepared from mitotic as well as interphase cells. The experiments reported here establish that hyperphosphorylation of the CTD is a good indicator of polymerase's association with snRNP and SR splicing factors, but not of its transcriptional activity. Most importantly, the present study suggests that splicing factors may associate with the

  3. Modeling of autosomal-dominant retinitis pigmentosa in Caenorhabditis elegans uncovers a nexus between global impaired functioning of certain splicing factors and cell type-specific apoptosis.

    PubMed

    Rubio-Peña, Karinna; Fontrodona, Laura; Aristizábal-Corrales, David; Torres, Silvia; Cornes, Eric; García-Rodríguez, Francisco J; Serrat, Xènia; González-Knowles, David; Foissac, Sylvain; Porta-De-La-Riva, Montserrat; Cerón, Julián

    2015-12-01

    Retinitis pigmentosa (RP) is a rare genetic disease that causes gradual blindness through retinal degeneration. Intriguingly, seven of the 24 genes identified as responsible for the autosomal-dominant form (adRP) are ubiquitous spliceosome components whose impairment causes disease only in the retina. The fact that these proteins are essential in all organisms hampers genetic, genomic, and physiological studies, but we addressed these difficulties by using RNAi in Caenorhabditis elegans. Our study of worm phenotypes produced by RNAi of splicing-related adRP (s-adRP) genes functionally distinguishes between components of U4 and U5 snRNP complexes, because knockdown of U5 proteins produces a stronger phenotype. RNA-seq analyses of worms where s-adRP genes were partially inactivated by RNAi, revealed mild intron retention in developing animals but not in adults, suggesting a positive correlation between intron retention and transcriptional activity. Interestingly, RNAi of s-adRP genes produces an increase in the expression of atl-1 (homolog of human ATR), which is normally activated in response to replicative stress and certain DNA-damaging agents. The up-regulation of atl-1 correlates with the ectopic expression of the pro-apoptotic gene egl-1 and apoptosis in hypodermal cells, which produce the cuticle, but not in other cell types. Our model in C. elegans resembles s-adRP in two aspects: The phenotype caused by global knockdown of s-adRP genes is cell type-specific and associated with high transcriptional activity. Finally, along with a reduced production of mature transcripts, we propose a model in which the retina-specific cell death in s-adRP patients can be induced through genomic instability.

  4. Nuclear speckles are involved in nuclear aggregation of PABPN1 and in the pathophysiology of oculopharyngeal muscular dystrophy.

    PubMed

    Bengoechea, Rocío; Tapia, Olga; Casafont, Iñigo; Berciano, José; Lafarga, Miguel; Berciano, María T

    2012-04-01

    Nuclear speckles are essential nuclear compartments involved in the assembly, delivery and recycling of pre-mRNA processing factors, and in the post-transcriptional processing of pre-mRNAs. Oculopharyngeal muscular dystrophy (OPMD) is caused by a small expansion of the polyalanine tract in the poly(A)-binding protein nuclear 1 (PABPN1). Aggregation of expanded PABPN1 into intranuclear inclusions (INIs) in skeletal muscle fibers is the pathological hallmark of OPMD. In this study what we have analyzed in muscle fibers of OPMD patients and in primary cultures of human myoblasts are the relationships between nuclear speckles and INIs, and the contribution of the former to the biogenesis of the latter. While nuclear speckles concentrate snRNP splicing factors and PABPN1 in control muscle fibers, they are depleted of PABPN1 and appear closely associated with INIs in muscle fibers of OPMD patients. The induction of INI formation in human myoblasts expressing either wild type GFP-PABPN1 or expanded GFP-PABPN1-17ala demonstrates that the initial aggregation of PABPN1 proteins and their subsequent growth in INIs occurs at the edges of the nuclear speckles. Moreover, the growing of INIs gradually depletes PABPN1 proteins and poly(A) RNA from nuclear speckles, although the existence of these nuclear compartments is preserved. Time-lapse experiments in cultured myoblasts confirm nuclear speckles as biogenesis sites of PABPN1 inclusions. Given the functional importance of nuclear speckles in the post-transcriptional processing of pre-mRNAs, the INI-dependent molecular reorganization of these nuclear compartments in muscle fibers may cause a severe dysfunction in nuclear trafficking and processing of polyadenylated mRNAs, thereby contributing to the molecular pathophysiology of OPMD. Our results emphasize the potential importance of nuclear speckles as nuclear targets of neuromuscular disorders.

  5. Assembly, nuclear import and function of U7 snRNPs studied by microinjection of synthetic U7 RNA into Xenopus oocytes.

    PubMed

    Stefanovic, B; Hackl, W; Lührmann, R; Schümperli, D

    1995-08-25

    In Xenopus oocytes in vitro transcribed mouse U7 RNA is assembled into small nuclear ribonucleoproteins (snRNPs) that are functional in histone RNA 3' processing. If the special Sm binding site of U7 (AAUUUGUCUAG, U7 Sm WT) is converted into the canonical Sm sequence derived from the major snRNAs (AAUUUUUGGAG, U7 Sm OPT) the RNA assembles into a particle which accumulates more efficiently in the nucleus, but which is non-functional. U7 RNA with a heavily mutated Sm binding site (AACGCGUCAUG, U7 Sm MUT) is deficient in nuclear accumulation and function. By UV cross-linking U7 Sm WT RNA can be linked to three proteins, i.e. the common snRNP proteins G and B/B' and an apparently U7-specific protein of 40 kDa. As a result of altering the Sm binding site, U7 Sm OPT RNA cannot be cross-linked to the 40 kDa protein and no cross-links are obtained with U7 Sm MUT RNA. The fact that the Sm site also interacts with at least one U7-specific protein is so far unique to U7 RNA and may provide an explanation for the atypical sequence of this site. All described RNA-protein interactions, including that with the 40 kDa protein, already occur in the cytoplasm. An additional cytoplasmic photoadduct obtained with U7 Sm WT and U7 Sm OPT, but not U7 Sm MUT, RNAs is indicative of a protein of 60-80 kDa. The m7G cap structure of U7 Sm WT and U7 Sm OPT RNA becomes hypermethylated. However, the 3mG cap enhances, but is not required for, nuclear accumulation. Finally, U7 Sm WT RNA is functional in histone RNA processing even when bearing an ApppG cap. PMID:7667090

  6. Structural and functional analysis of the human spliceosomal DEAD-box helicase Prp28

    SciTech Connect

    Möhlmann, Sina; Mathew, Rebecca; Neumann, Piotr; Schmitt, Andreas; Lührmann, Reinhard; Ficner, Ralf

    2014-06-01

    The crystal structure of the helicase domain of the human spliceosomal DEAD-box protein Prp28 was solved by SAD. The binding of ADP and ATP by Prp28 was studied biochemically and analysed with regard to the crystal structure. The DEAD-box protein Prp28 is essential for pre-mRNA splicing as it plays a key role in the formation of an active spliceosome. Prp28 participates in the release of the U1 snRNP from the 5′-splice site during association of the U5·U4/U6 tri-snRNP, which is a crucial step in the transition from a pre-catalytic spliceosome to an activated spliceosome. Here, it is demonstrated that the purified helicase domain of human Prp28 (hPrp28ΔN) binds ADP, whereas binding of ATP and ATPase activity could not be detected. ATP binding could not be observed for purified full-length hPrp28 either, but within an assembled spliceosomal complex hPrp28 gains ATP-binding activity. In order to understand the structural basis for the ATP-binding deficiency of isolated hPrp28, the crystal structure of hPrp28ΔN was determined at 2.0 Å resolution. In the crystal the helicase domain adopts a wide-open conformation, as the two RecA-like domains are extraordinarily displaced from the productive ATPase conformation. Binding of ATP is hindered by a closed conformation of the P-loop, which occupies the space required for the γ-phosphate of ATP.

  7. Antiribonucleoprotein antibodies in children with HIV infection: a comparative study with childhood-onset systemic lupus erythematosus.

    PubMed

    González, C M; López-Longo, F J; Samson, J; Monteagudo, I; Grau, R; Rodríguez-Mahou, M; St-Cyr, C; Lapointe, N; Carreño, L

    1998-01-01

    A number of clinical and laboratory features of HIV infection are found in systemic lupus erythematosus (SLE). The objective of this study was to analyze the presence of circulating antibodies to small nuclear ribonucleoproteins (snRNP) in both diseases. Sera from 44 HIV-infected children, from 22 patients with childhood-onset SLE, and from 50 healthy children were studied. Anti-snRNP antibodies were detected by ELISA using recombinant and affinity-purified nuclear antigens, by counterimmunoelectrophoresis (CIE), and by immunoblotting using extractable nuclear antigens. Results included the detection of anti-snRNP antibodies by ELISA in 30 HIV-infected patients (68.1%) and 19 SLE patients (86.3%). These antibodies were directed against U1-RNP (61.3% and 77.2%, respectively), Sm (29.5% and 54.5%, respectively), 60 kDa Ro/SS-A (47.7% and 50%, respectively), and La/SS-B proteins (18.1% and 9%, respectively). None of the HIV-infected children and 11 SLE patients (50%) showed anti-snRNP antibodies by CIE. None of the HIV-infected patients showed anti-70 kDa U1-RNP or anti-D-Sm antibodies by immunoblotting. No differences between the two groups were noted on the presence of nonprecipitating anti-snRNP antibodies. No such reactivities were observed among the normal sera tested. The authors concluded that nonprecipitating anti-snRNP antibodies in HIV-infected children are as frequent as in childhood-onset SLE. The significance of these antibodies is not clear at present. Although polyreactive and low-affinity antibodies and a mechanism of molecular mimicry may explain these results, a specific stimulation of B cells by nuclear antigens could not be excluded.

  8. PRP4K is a HER2-regulated modifier of taxane sensitivity

    PubMed Central

    Corkery, Dale P; Le Page, Cécile; Meunier, Liliane; Provencher, Diane; Mes-Masson, Anne-Marie; Dellaire, Graham

    2015-01-01

    The taxanes are used alone or in combination with anthracyclines or platinum drugs to treat breast and ovarian cancer, respectively. Taxanes target microtubules in cancer cells and modifiers of taxane sensitivity have been identified in vitro, including drug efflux and mitotic checkpoint proteins. Human epidermal growth factor receptor 2 (HER2/ERBB2) gene amplification is associated with benefit from taxane therapy in breast cancer yet high HER2 expression also correlates with poor survival in both breast and ovarian cancer. The pre-mRNA splicing factor 4 kinase PRP4K (PRPF4B), which we identified as a component of the U5 snRNP also plays a role in regulating the spindle assembly checkpoint (SAC) in response to microtubule-targeting drugs. In this study, we found a positive correlation between PRP4K expression and HER2 status in breast and ovarian cancer patient tumors, which we determined was a direct result of PRP4K regulation by HER2 signaling. Knock-down of PRP4K expression reduced the sensitivity of breast and ovarian cancer cell lines to taxanes, and low PRP4K levels correlated with in vitro-derived and patient acquired taxane resistance in breast and ovarian cancer. Patients with high-grade serous ovarian cancer and high HER2 levels had poor overall survival; however, better survival in the low HER2 patient subgroup treated with platinum/taxane-based therapy correlated positively with PRP4K expression (HR = 0.37 [95% CI 0.15-0.88]; p = 0.03). Thus, PRP4K functions as a HER2-regulated modifier of taxane sensitivity that may have prognostic value as a marker of better overall survival in taxane-treated ovarian cancer patients. PMID:25602630

  9. A conserved intronic U1 snRNP-binding sequence promotes trans-splicing in Drosophila

    PubMed Central

    Gao, Jun-Li; Fan, Yu-Jie; Wang, Xiu-Ye; Zhang, Yu; Pu, Jia; Li, Liang; Shao, Wei; Zhan, Shuai; Hao, Jianjiang

    2015-01-01

    Unlike typical cis-splicing, trans-splicing joins exons from two separate transcripts to produce chimeric mRNA and has been detected in most eukaryotes. Trans-splicing in trypanosomes and nematodes has been characterized as a spliced leader RNA-facilitated reaction; in contrast, its mechanism in higher eukaryotes remains unclear. Here we investigate mod(mdg4), a classic trans-spliced gene in Drosophila, and report that two critical RNA sequences in the middle of the last 5′ intron, TSA and TSB, promote trans-splicing of mod(mdg4). In TSA, a 13-nucleotide (nt) core motif is conserved across Drosophila species and is essential and sufficient for trans-splicing, which binds U1 small nuclear RNP (snRNP) through strong base-pairing with U1 snRNA. In TSB, a conserved secondary structure acts as an enhancer. Deletions of TSA and TSB using the CRISPR/Cas9 system result in developmental defects in flies. Although it is not clear how the 5′ intron finds the 3′ introns, compensatory changes in U1 snRNA rescue trans-splicing of TSA mutants, demonstrating that U1 recruitment is critical to promote trans-splicing in vivo. Furthermore, TSA core-like motifs are found in many other trans-spliced Drosophila genes, including lola. These findings represent a novel mechanism of trans-splicing, in which RNA motifs in the 5′ intron are sufficient to bring separate transcripts into close proximity to promote trans-splicing. PMID:25838544

  10. Import and export of nuclear proteins: focus on the nucleocytoplasmic movements of two different species of mammalian estrogen receptor.

    PubMed

    Sebastian, Thomas; Sreeja, S; Thampan, Raghava Varman

    2004-05-01

    There is a wealth of information regarding the import and export of nuclear proteins in general. Nevertheless, the available data that deals with the nucleocytoplasmic movement of steroid hormone receptors remains highly limited. Some research findings reported during the past five years have succeeded in identifying proteins related to the movement of estrogen receptor alpha from the cytoplasm to the nucleus. What is striking in these findings is the facilitatory role of estradiol in the transport process. A similar conclusion has been drawn from the studies on the plasma membrane-to nucleus movement of the alternative form of estrogen receptor, the non-activated estrogen receptor (naER). The internalization of naER from the plasma membrane takes place only in the presence of estradiol. While the gene regulatory functions of ER alpha appear to get terminated following its ubiquitinization within the nucleus, the naER, through its deglycosylated form, the nuclear estrogen receptor II (nER II) continues to remain functional even beyond its existence within the nucleus. Recent studies have indicated the possibility that the estrogen receptor that regulates the nucleo cytoplasmic transport of m RNP is the nERII. This appears to be the result of the interaction between nERII and three proteins belonging to a group of small nuclear ribonucleo proteins (snRNP). The interaction of nERII with two of this protein appears to activate the inherent Mg2+ ATPase activity of the complex, which leads to the exit of the RNP through the nuclear pore complex. PMID:15228090

  11. Physical and genetic interactions of yeast Cwc21p, an ortholog of human SRm300/SRRM2, suggest a role at the catalytic center of the spliceosome.

    PubMed

    Grainger, Richard J; Barrass, J David; Jacquier, Alain; Rain, Jean-Christophe; Beggs, Jean D

    2009-12-01

    In Saccharomyces cerevisiae, Cwc21p is a protein of unknown function that is associated with the NineTeen Complex (NTC), a group of proteins involved in activating the spliceosome to promote the pre-mRNA splicing reaction. Here, we show that Cwc21p binds directly to two key splicing factors-namely, Prp8p and Snu114p-and becomes the first NTC-related protein known to dock directly to U5 snRNP proteins. Using a combination of proteomic techniques we show that the N-terminus of Prp8p contains an intramolecular fold that is a Snu114p and Cwc21p interacting domain (SCwid). Cwc21p also binds directly to the C-terminus of Snu114p. Complementary chemical cross-linking experiments reveal reciprocal protein footprints between the interacting Prp8 and Cwc21 proteins, identifying the conserved cwf21 domain in Cwc21p as a Prp8p binding site. Genetic and functional interactions between Cwc21p and Isy1p indicate that they have related functions at or prior to the first catalytic step of splicing, and suggest that Cwc21p functions at the catalytic center of the spliceosome, possibly in response to environmental or metabolic changes. We demonstrate that SRm300, the only SR-related protein known to be at the core of human catalytic spliceosomes, is a functional ortholog of Cwc21p, also interacting directly with Prp8p and Snu114p. Thus, the function of Cwc21p is likely conserved from yeast to humans.

  12. Nuclear structures in Tribolium castaneum oocytes.

    PubMed

    Bogolyubov, Dmitry S; Batalova, Florina M; Kiselyov, Artyom M; Stepanova, Irina S

    2013-10-01

    The first ultrastructural and immunomorphological characteristics of the karyosphere (karyosome) and extrachromosomal nuclear bodies in the red flour beetle, Tribolium castaneum, are presented. The karyosphere forms early in the diplotene stage of meiotic prophase by the gathering of all oocyte chromosomes in a limited nuclear volume. Using the BrUTP assay, T. castaneum oocyte chromosomes united in the karyosphere maintain their transcriptional activity until the end of oocyte growth. Hyperphosphorylated RNA polymerase II and basal transcription factors (TFIID and TFIIH) were detected in the perichromatin region of the karyosphere. The T. castaneum karyosphere has an extrachromosomal capsule that separates chromosomes from the rest of the nucleoplasm. Certain structural proteins (F-actin, lamin B) were found in the capsule. Unexpectedly, the karyosphere capsule in T. castaneum oocytes was found to be enriched in TMG-capped snRNAs, which suggests that the capsule is not only a structural support for the karyosphere, but may be involved in biogenesis of snRNPs. We also identified the counterparts of 'universal' extrachromosomal nuclear domains, Cajal bodies (CBs) and interchromatin granule clusters (IGCs). Nuclear bodies containing IGC marker protein SC35 display some features unusual for typical IGCs. SC35 domains in T. castaneum oocytes are predominantly fibrillar complex bodies that do not contain trimethyl guanosine (TMG)-capped small nuclear (sn) RNAs. Microinjections of 2'-O-methyl (U)22 probes into the oocytes allowed revealing poly(A)+ RNAs in these nuclear domains. Several proteins related to mRNA export (heterogeneous ribonucleoprotein core protein A1, export adapters Y14 and Aly and export receptor NXF1) were also detected there. We believe that unusual SC35 nuclear domains of T. castaneum oocytes are possibly involved in mRNP but not snRNP biogenesis.

  13. Mixed connective tissue disease.

    PubMed

    Gunnarsson, Ragnar; Hetlevik, Siri Opsahl; Lilleby, Vibke; Molberg, Øyvind

    2016-02-01

    The concept of mixed connective tissue disease (MCTD) as a separate connective tissue disease (CTD) has persisted for more than four decades. High titers of antibodies targeting the U1 small nuclear ribonucleoprotein particle (U1 snRNP) in peripheral blood are a sine qua non for the diagnosis of MCTD, in addition to distinct clinical features including Raynaud's phenomenon (RP), "puffy hands," arthritis, myositis, pleuritis, pericarditis, interstitial lung disease (ILD), and pulmonary hypertension (PH). Recently, population-based epidemiology data from Norway estimated the point prevalence of adult-onset MCTD to be 3.8 per 100,000 and the mean annual incidence to be 2.1 per million per year, supporting the notion that MCTD is the least common CTD. Little is known about the etiology of MCTD, but recent genetic studies have confirmed that MCTD is a strongly HLA (​human leukocyte antigen)-linked disease, as the HLA profiles of MCTD differ distinctly from the corresponding profiles of ethnically matched healthy controls and other CTDs. In the first section of this review, we provide an update on the clinical, immunological, and genetic features of MCTD and discuss the relationship between MCTD and the other CTDs. Then we proceed to discuss the recent advances in therapy and our current understanding of prognosis and prognostic factors, especially those that are associated with the more serious pulmonary and cardiovascular complications of the disease. In the final section, we discuss some of the key, unresolved questions related to anti-RNP-associated diseases and indicate how these questions may be approached in future studies. PMID:27421219

  14. DDX6 transfers P-TEFb kinase to the AF4/AF4N (AFF1) super elongation complex.

    PubMed

    Mück, Fabian; Bracharz, Silvia; Marschalek, Rolf

    2016-01-01

    AF4/AFF1 and AF5/AFF4 are both backbones for the assembly of "super elongation complexes" (SECs) that exert 2 distinct functions after the recruitment of P-TEFb from the 7SK snRNP: (1) initiation and elongation of RNA polymerase II gene transcription, and (2) modification of transcribed gene regions by distinct histone methylation patterns. In this study we aimed to investigate one of the initial steps, namely how P-TEFb is transferred from 7SK snRNPs to the SECs. In particular, we were interested in the role of DDX6 that we have recently identified as part of the AF4 complex. DDX6 is an evolutionarily conserved member of the DEAD-box RNA helicase family that is known to control miRNA and mRNA biology (translation, storage and degradation). Overexpressed DDX6 is associated with different cancer types and with c-Myc protein overexpression. We could demonstrate that DDX6 binds to 7SK snRNA and causes the release and transfer of P-TEFb to the AF4/AF4N SEC. DDX6 also binds stably to AF4 and AF4N as demonstrated by GST pull-down and co-immunoprecipitation experiments. As a consequence, overexpression of either AF4/AF4N or DDX6 resulted in a strong increase of mRNA production (5-6 fold), while their simultaneous expression increased the cellular mRNA production by 11-fold. Conversely, the corresponding knockdown of DDX6 decreased mRNA production by 70%. In conclusion, AF4/AF4N and DDX6 represent key molecules for the elongation process of gene transcription and a model will be proposed for the hand-over process of P-TEFb to SECs. PMID:27679741

  15. Estrogen receptor alpha (ESR1)-signaling regulates the expression of the taxane-response biomarker PRP4K.

    PubMed

    Lahsaee, Sara; Corkery, Dale P; Anthes, Livia E; Holly, Alice; Dellaire, Graham

    2016-01-01

    The pre-mRNA splicing factor 4 kinase PRP4K (PRPF4B), is an essential kinase that is a component of the U5 snRNP and functions in spliceosome assembly. We demonstrated that PRP4K is a novel biological marker for taxane response in ovarian cancer patients and reduced levels of PRP4K correlate with intrinsic and acquired taxane resistance in both breast and ovarian cancer. Breast cancer treatments are chosen based on hormone and growth factor receptor status, with HER2 (ERBB2) positive breast cancer patients receiving anti-HER2 agents and taxanes and estrogen receptor alpha (ESR1) positive (ER+) breast cancer patients receiving anti-estrogen therapies such as tamoxifen. Here we demonstrate that PRP4K is expressed in the normal mammary duct epithelial cells of the mouse, and that estrogen induces PRP4K gene and protein expression in ER+ human MCF7 breast cancer cells. Estrogen acts through ESR1 to regulate PRP4K expression, as over-expression of ESR1 in the ER-negative MDA-MB-231 breast cancer cell line increased the expression of this kinase, and knock-down of ESR1 in ER+ T47D breast cancer cells reduced PRP4K levels. Furthermore, treatment with 4-hydroxytamoxifen (4-OHT) resulted in a dose-dependent decrease in PRP4K protein expression in MCF7 cells. Consistent with our previous studies identifying PRP4K as a taxane-response biomarker, reduced PRP4K expression in 4-OHT-treated cells correlated with reduced sensitivity to paclitaxel. Thus, PRP4K is novel estrogen regulated kinase, and its levels can be reduced by 4-OHT in ER+ breast cancer cells altering their response to taxanes. PMID:26712520

  16. U1 snRNA as an effective vector for stable expression of antisense molecules and for the inhibition of the splicing reaction.

    PubMed

    Martone, Julie; De Angelis, Fernanda Gabriella; Bozzoni, Irene

    2012-01-01

    We report the use of the U1 snRNA as a vector for the stable expression of antisense molecules against the splice junctions of specific dystrophin exons. The single-stranded 5' terminus of U1 can be replaced by unrelated sequences as long as 50 nucleotides without affecting both the stability and the ability to assemble into snRNP particles. Effective exon skipping has been obtained for different dystrophin exons by antisense sequences against 5' and 3' splice sites alone or in combination with ESE sequences. The efficacy of these molecules has been studied both in in vitro systems and in animals. In both cases the chimeric molecules, delivered as part of lentiviral or AAV vectors (De Angelis et al. Proc Natl Acad Sci USA 99:9456-9461, 2002; Denti et al. Proc Natl Acad Sci USA 103: 3758-3763, 2006; Denti et al. Hum Gene Ther 17: 565-743, 2006; Denti et al. Hum Gene Ther 19: 601-608, 2008; Incitti et al. Mol Ther 18: 1675-1682, 2010), provided high skipping activity and efficient rescue of dystrophin synthesis. Moreover, the U1-antisense molecules, delivered to mice via systemic injection of recombinant AAV viruses, displayed body wide transduction, long-term expression, dystrophin rescue as well as morphological and functional benefit (Denti et al. Hum Gene Ther 19: 601-608, 2008). In this Chapter we report methods for producing U1-antisense expression cassettes in the backbone of lentiviral constructs and for testing their activity both in patients' derived myoblasts as well as in fibroblasts reprogrammed to muscle differentiation.

  17. Temporal Profiling of Lapatinib-suppressed Phosphorylation Signals in EGFR/HER2 Pathways*

    PubMed Central

    Imami, Koshi; Sugiyama, Naoyuki; Imamura, Haruna; Wakabayashi, Masaki; Tomita, Masaru; Taniguchi, Masatoshi; Ueno, Takayuki; Toi, Masakazu; Ishihama, Yasushi

    2012-01-01

    Lapatinib is a clinically potent kinase inhibitor for breast cancer patients because of its outstanding selectivity for epidermal growth factor receptor (EGFR) and EGFR2 (also known as HER2). However, there is only limited information about the in vivo effects of lapatinib on EGFR/HER2 and downstream signaling targets. Here, we profiled the lapatinib-induced time- and dose-dependent phosphorylation dynamics in SKBR3 breast cancer cells by means of quantitative phosphoproteomics. Among 4953 identified phosphopeptides from 1548 proteins, a small proportion (5–7%) was regulated at least twofold by 1–10 μm lapatinib. We obtained a comprehensive phosphorylation map of 21 sites on EGFR/HER2, including nine novel sites on HER2. Among them, serine/threonine phosphosites located in a small region of HER2 (amino acid residues 1049–1083) were up-regulated by the drug, whereas all other sites were down-regulated. We show that cAMP-dependent protein kinase is involved in phosphorylation of this particular region of HER2 and regulates HER2 tyrosine kinase activity. Computational analyses of quantitative phosphoproteome data indicated for the first time that protein-protein networks related to cytoskeletal organization and transcriptional/translational regulation, such as RNP complexes (i.e. hnRNP, snRNP, telomerase, ribosome), are linked to EGFR/HER2 signaling networks. To our knowledge, this is the first report to profile the temporal response of phosphorylation dynamics to a kinase inhibitor. The results provide new insights into EGFR/HER2 regulation through region-specific phosphorylation, as well as a global view of the cellular signaling networks associated with the anti-breast cancer action of lapatinib. PMID:22964224

  18. Modeling of autosomal-dominant retinitis pigmentosa in Caenorhabditis elegans uncovers a nexus between global impaired functioning of certain splicing factors and cell type-specific apoptosis.

    PubMed

    Rubio-Peña, Karinna; Fontrodona, Laura; Aristizábal-Corrales, David; Torres, Silvia; Cornes, Eric; García-Rodríguez, Francisco J; Serrat, Xènia; González-Knowles, David; Foissac, Sylvain; Porta-De-La-Riva, Montserrat; Cerón, Julián

    2015-12-01

    Retinitis pigmentosa (RP) is a rare genetic disease that causes gradual blindness through retinal degeneration. Intriguingly, seven of the 24 genes identified as responsible for the autosomal-dominant form (adRP) are ubiquitous spliceosome components whose impairment causes disease only in the retina. The fact that these proteins are essential in all organisms hampers genetic, genomic, and physiological studies, but we addressed these difficulties by using RNAi in Caenorhabditis elegans. Our study of worm phenotypes produced by RNAi of splicing-related adRP (s-adRP) genes functionally distinguishes between components of U4 and U5 snRNP complexes, because knockdown of U5 proteins produces a stronger phenotype. RNA-seq analyses of worms where s-adRP genes were partially inactivated by RNAi, revealed mild intron retention in developing animals but not in adults, suggesting a positive correlation between intron retention and transcriptional activity. Interestingly, RNAi of s-adRP genes produces an increase in the expression of atl-1 (homolog of human ATR), which is normally activated in response to replicative stress and certain DNA-damaging agents. The up-regulation of atl-1 correlates with the ectopic expression of the pro-apoptotic gene egl-1 and apoptosis in hypodermal cells, which produce the cuticle, but not in other cell types. Our model in C. elegans resembles s-adRP in two aspects: The phenotype caused by global knockdown of s-adRP genes is cell type-specific and associated with high transcriptional activity. Finally, along with a reduced production of mature transcripts, we propose a model in which the retina-specific cell death in s-adRP patients can be induced through genomic instability. PMID:26490224

  19. DDX6 transfers P-TEFb kinase to the AF4/AF4N (AFF1) super elongation complex

    PubMed Central

    Mück, Fabian; Bracharz, Silvia; Marschalek, Rolf

    2016-01-01

    AF4/AFF1 and AF5/AFF4 are both backbones for the assembly of “super elongation complexes” (SECs) that exert 2 distinct functions after the recruitment of P-TEFb from the 7SK snRNP: (1) initiation and elongation of RNA polymerase II gene transcription, and (2) modification of transcribed gene regions by distinct histone methylation patterns. In this study we aimed to investigate one of the initial steps, namely how P-TEFb is transferred from 7SK snRNPs to the SECs. In particular, we were interested in the role of DDX6 that we have recently identified as part of the AF4 complex. DDX6 is an evolutionarily conserved member of the DEAD-box RNA helicase family that is known to control miRNA and mRNA biology (translation, storage and degradation). Overexpressed DDX6 is associated with different cancer types and with c-Myc protein overexpression. We could demonstrate that DDX6 binds to 7SK snRNA and causes the release and transfer of P-TEFb to the AF4/AF4N SEC. DDX6 also binds stably to AF4 and AF4N as demonstrated by GST pull-down and co-immunoprecipitation experiments. As a consequence, overexpression of either AF4/AF4N or DDX6 resulted in a strong increase of mRNA production (5-6 fold), while their simultaneous expression increased the cellular mRNA production by 11-fold. Conversely, the corresponding knockdown of DDX6 decreased mRNA production by 70%. In conclusion, AF4/AF4N and DDX6 represent key molecules for the elongation process of gene transcription and a model will be proposed for the hand-over process of P-TEFb to SECs.

  20. Solitary solutions for a class of Schrödinger equations in R^3

    NASA Astrophysics Data System (ADS)

    Wang, Youjun

    2016-08-01

    In this paper, we consider a model problem arising from a classical planar Heisenberg ferromagnetic spin chain -Δ u + (λ + ɛ') u ∓ Δ √{1-u2}u/√{1- u2} - ɛ'u/√{1 - u2} = 0, x in R3, where {λ} and {ɛ'}are real constants. By variational methods and perturbation arguments, we study the existence of positive classical solutions. Our results generalize the previous results in one-dimensional space given by Brüll et al. [4].

  1. Strong correlation of major earthquakes with solid-earth tides in part of the eastern United States

    USGS Publications Warehouse

    Weems, R.E.; Perry, W.H.

    1989-01-01

    East of the eastern American continental divide and south of lat. 42.5??N, moderate to large historic earthquakes correlate strongly with times of high and low solid-earth tides. This effect is most pronounced when solar declination lies between 17??N and 17??S. Significant correlation also exist between major earthquakes, time of day, lunar declinations, and lunar phase. -Authors

  2. Frequent loss of heterozygosity on chromosome 17 at 17q11.2-q12 in Barrett's adenocarcinoma.

    PubMed Central

    Swift, A.; Risk, J. M.; Kingsnorth, A. N.; Wright, T. A.; Myskow, M.; Field, J. K.

    1995-01-01

    Allelic loss on chromosome 17 in 18 Barrett's oesophageal tumours was analysed with 17 polymorphic microsatellite markers. Loss of heterozygosity (LOH) of one or more markers was seen in 72% (13 of 18) tumours on 17p and 56% (10 of 18) on 17q. The highest 17p losses were found at D17S799 (62%, five of eight) and D17S261 (55%, five of nine), while loss at the p53 locus was 31% (5 of 16). The highest loss on 17q was found at the TCF-2 (17q11.2-q12) locus with 66% (8 of 12) LOH. TCF-2 was the only marker lost in two of the tumour samples; furthermore, TCF-2 was lost in four other tumours which retained heterozygosity at the markers on either side of it, D17S261 and D17S740. Six markers were used to assess LOH at 17q11.2-q12, and five of eight of the tumour specimens which had LOH at TCF-2 had no other loss on 17q. No statistically significant correlations were found between loss on 17q or 17p and any clinicopathological parameters. We propose from these data that the 17q11.2-q12 region contains a novel predisposing gene in Barrett's adenocarcinomas and may represent the site of a tumour-suppressor gene. PMID:7734326

  3. 76 FR 56790 - Notice of Intent To Prepare an Amendment to the Caliente Resource Management Plan and Associated...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-09-14

    ... Delta Water Conservation District, and ARC Vineyards, LLC to purchase the following public lands... Kaweah Delta Water Conservation District, is described as: Mount Diablo Meridian T. 17 S., R. 28 E, Sec... suitable for management by other agencies (e.g., for conservation purposes) or land tenure adjustment...

  4. Viral infection correlated with superoxide anion radicals production and natural and synthetic copper complexes.

    PubMed

    Tomas, E; Popescu, A; Titire, A; Cajal, N; Cristescu, C; Tomas, S

    1989-01-01

    Studies conducted on asymmetric triazine derivatives synthetized at the Chemical and Pharmaceutical Research Institute showed that products S1, S16, S17, S19, S20 and S22 have a remarkable O2- radical scavenger activity. Among these derivatives, the product S1 is the most efficient as an antiviral agent.

  5. Testing for gene-gene interaction controlling total IgE in families from Barbados: evidence of sensitivity regarding linkage heterogeneity among families.

    PubMed

    Barnes, K C; Mathias, R A; Nickel, R; Freidhoff, L R; Stockton, M L; Xue, X; Naidu, R P; Levett, P N; Casolaro, V; Beaty, T H

    2001-01-15

    Genetic heterogeneity has been proposed as a hallmark feature of allergic disease. To test the hypothesis that total IgE levels are jointly influenced by a locus on chromosome 12q21.1-q21.31 and a locus on 17q11.2-q21.2, we conducted multipoint allele-sharing analyses using nonparametric linkage (NPL) methods on Afro-Caribbean families from Barbados to test for evidence of gene-gene interactions. Significant correlations were observed between NPL scores at D12S1052 and both D17S1293 and D17S1299 for a dichotomized phenotype of total IgE. An analysis of family-specific NPL scores revealed that evidence for interaction was being driven largely by one multiplex pedigree (NPL = 12.01, 12.23, and 12.16 at D12S1052, D17S1293, and D17S1299, respectively). Using the programs SIMWALK (v2.0) and GOLD, a different set of haplotypes in this influential family was observed around D12S1052 and the 17q loci compared to the other Barbados pedigrees. Our findings are a classic example of founder effect, provide evidence for sensitivity of this type of linkage analysis to unusual pedigrees, and highlight an element of genetic heterogeneity that has been given little attention in the study of complex traits.

  6. MASTER: PSN in interacting galaxy, dwarf nova and possible YSO objects

    NASA Astrophysics Data System (ADS)

    Gress, O.; Shurpakov, S.; Pogrosheva, T.; Lipunov, V.; Lopez, R. Rebolo; Buckley, D.; Tlatov, A.; Gorbovskoy, E.; Tiurina, N.; Balanutsa, P.; Kuznetsov, A.; Kornilov, V.; Vlasenko, D.; Popova, E.; Ricart, M. Serra; Israelian, G.; Lodieu, N.; Potter, S.; Kniazev, A.; Senik, V.; Dormidontov, D.; Parkhomenko, A.; Kochutina, N.; Shumkov, V.

    2016-07-01

    MASTER-IAC auto-detection system ( Lipunov et al., "MASTER Global Robotic Net", Advances in Astronomy, 2010, 349171 ) discovered OT source at (RA, Dec) = 15h 16m 47.17s +28d 37m 42.8s on 2016-06-28.97168 UT. The OT unfiltered magnitude is 17.8m (limit 20.2m).

  7. A radiation hybrid map of the BRCA1 region of chromosome 17q12-q21

    SciTech Connect

    Abel, K.J.; Boehnke, M.; Prahalad, M.; Flejter, W.L.; Watkins, M.; Chandrasekharappa, S.C.; Glover, T.W. Howard Hughes Medical Institute, Ann Arbor, MI ); Ho, P.; VanderStoep, J.; Weber, B.L. ); Collins, F.S. Michigan Human Genome Center, Ann Arbor, MI Howard Hughes Medical Institute, Ann Arbor, MI )

    1993-09-01

    The chromosomal region 17q12-q21 contains a gene (BRCA1) conferring susceptibility to early-onset familial breast and ovarian cancer. An 8000-rad radiation-reduced hybrid (RH) panel was constructed to provide a resource for long-range mapping of this region. A large fraction of the hybrids ([approximately]90%) retained detectable human chromosome 17 sequences. The complete panel of 76 hybrids was scored for the presence or absence of 22 markers from this chromosomal region, including 14 cloned genes, seven microsatellite repeats, and one anonymous DNA segment. Statistical analysis of the marker retention data employing multipoint methods provided both comprehensive and framework maps of this chromosomal region, including distance estimates between adjacent markers. The comprehensive RH map includes 17 loci and spans 179 cRays[sub (8000)]. Likelihood ratios of at least 1000:1 support the 10-locus framework order: cen-D17S250-ERBB2-(THRA1, TOP2A)-D17S855-PPY-D17S190-MTBT1-GP3A-BTR-D17S588-tel. The order obtained from RH mapping, when used in conjunction with other methods, will be useful in linkage analysis of breast cancer families and will facilitate the development of a physical map of this region. 42 refs., 3 figs., 2 tabs.

  8. Linkage to markers for the chromosome region 17q12-q21 in 13 Dutch breast cancer kindreds

    SciTech Connect

    Devilee, P.; Cornelis, R.S.; Bardoel, A.; Vliet, M. van; Leeuwen, I. van; Cleton, F.J.; Vasen, H.F.A.; Cornelisse, C.J.; Meera Khan, P. ); Bootsma, A.; Klein, A. de; Lindhout, D. )

    1993-04-01

    The authors have performed linkage analysis with five markers for the chromosome region 17q12-q21 in 13 Dutch breast cancer kindreds in order to find support for the claim by Hall et al. that a gene in this region, termed [open quotes]BRCA1,[close quotes] is associated with predisposition to early-onset familial breast cancer. This work is part of a collaborative study, the results of which are published elsewhere in this issue. Best evidence for linkage was observed with the marker CMM86 (D17S74) in pedigrees with an average age at onset of [le]47 years (LOD score = 1.77 at 1% recombination). In one breast-ovarian cancer family with a high probability of being linked to 17q, they observed one putative recombinant between D17S250 and D17S579, which suggests that BRCA1 is proximal to D17S579. 32 refs., 2 figs., 2 tabs.

  9. Feminist and Multicultural Collaboration in Counseling Supervision: Voices from Two African American Women

    ERIC Educational Resources Information Center

    Steward, Robbie J.; Phelps, Rosemary E.

    2004-01-01

    This article highlights 2 African-American women participants' unique experiences and perspectives in the Supervision and Training work group at the Division 17's Section for the Advancement of Women (SAW) Michigan conference. Differences between the authors' and White group members' experiences and perceptions are presented and discussed.

  10. 45 CFR 5b.2 - Purpose and scope.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... entered into pursuant to sections 1816 and 1842 of the Social Security Act, 42 U.S.C. 1395h and 1395u. (2... Title 5, United States Code; volunteers where acceptance of their services are authorized by law;...

  11. 75 FR 3668 - Conditional Approval and Promulgation of State Implementation Plans; Ohio; Carbon Monoxide and...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-01-22

    ... economic feasibility. On March 23, 1995, (60 FR 15235), EPA inadvertently approved a version of paragraph... this version of paragraph (U)(2)(f) on July 28, 2009, at 74 FR 37171. Because this version...

  12. Indentifying environmental features for land management decisions. [Uinta Basin, Davis County foothills, and Farmington Bay in Utah

    NASA Technical Reports Server (NTRS)

    1980-01-01

    The wetlands and water-related land use in the Uinta Basin were classified and mapped using photointerpretation of U-2 infrared photography and digital LANDSAT data. The digital maps were used to augment photointerpretations. A highly effective diagnostic tool emerged when the LANDSAT digital print was photoreduced to a film positive at the same scale as the U-2 film and overlain on the U-2 color film. As a result of this merging technique, cover types can be identified more accurately and probablistic statements can be made about the relative amounts of water being consumed in one pasture vs. another. The hazards to urban development on sensitive and unstable land in the foothills of Davis County were studied using NASA U-2 photography. Shoreline fluctuations were mapped in the Farmington Bay using LANDSAT digital data.

  13. 49 CFR 107.117 - Emergency processing.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ..., Hazardous Materials Standards Division, Office of Operating and Environmental Standards, U.S. Coast Guard, U...). (2) Noncertificate-Holding Aircraft (Those Which Operate Under 14 CFR Part 91): The Federal...

  14. Task 1: Correlation of satellite and ground data in air pollution studies. Task 2: Investigation to relate the chlorophyll and suspended sediment content in the waters of the lower Chesapeake Bay to ERTS-1 imagery. Task 3: The use of ERTS-1 to more fully utilize and apply marine station data to the study of productivity along the Eastern Shelf expanded waters of the United States

    NASA Technical Reports Server (NTRS)

    Copeland, G. E. (Principal Investigator); Bandy, A. R.; Fleischer, P.; Ludwick, J. C. (Principal Investigator); Hanna, W. J.; Gosink, T. A.; Bowker, D. W.; Marshall, H. G. (Principal Investigator)

    1972-01-01

    The author has identified the following significant results. Analysis of U-2 imagery of CARETS site indicates smoke plumes can be easily detected. First look at selected ERTS-1 color composites demonstrates plumes from forest fires can be detected.

  15. Local discrimination scheme for some unitary operations

    NASA Astrophysics Data System (ADS)

    Cao, TianQing; Gao, Fei; Tian, GuoJing; Xie, ShuCui; Wen, QiaoYan

    2016-09-01

    It has been shown that for two different multipartite unitary operations U 1 and U 2, when tr( U 1 † U 2) = 0, they can always be perfectly distinguished by local operations and classical communication in the single-run scenario. However, how to find the detailed scheme to complete the local discrimination is still a fascinating problem. In this paper, aiming at some U 1 and U 2 acting on the bipartite and tripartite space respectively, especially for U 1 † U 2 locally unitary equivalent to the high dimensional X-type hermitian unitary matrix V with tr V = 0, we put forward the explicit local distinguishing schemes in the single-run scenario.

  16. Thermal properties of nonstoichiometry uranium dioxide

    NASA Astrophysics Data System (ADS)

    Kavazauri, R.; Pokrovskiy, S. A.; Baranov, V. G.; Tenishev, A. V.

    2016-04-01

    In this paper, was developed a method of oxidation pure uranium dioxide to a predetermined deviation from the stoichiometry. Oxidation was carried out using the thermogravimetric method on NETZSCH STA 409 CD with a solid electrolyte galvanic cell for controlling the oxygen potential of the environment. 4 samples uranium oxide were obtained with a different ratio of oxygen-to-metal: O / U = 2.002, O / U = 2.005, O / U = 2.015, O / U = 2.033. For the obtained samples were determined basic thermal characteristics of the heat capacity, thermal diffusivity, thermal conductivity. The error of heat capacity determination is equal to 5%. Thermal diffusivity and thermal conductivity of the samples decreased with increasing deviation from stoichiometry. For the sample with O / M = 2.033, difference of both values with those of stoichiometric uranium dioxide is close to 50%.

  17. Noncommutative 3 Dimensional Soliton from Multi-instantons

    NASA Astrophysics Data System (ADS)

    Correa, D. H.; Forgacs, P.; Moreno, E. F.; Schaposnik, F. A.; Silva, G. A.

    2004-07-01

    We extend the relation between instanton and monopole solutions of the selfduality equations in SU(2) gauge theory to noncommutative space-times. Using this approach and starting from a noncommutative multi-instanton solution we construct a U(2) monopole configuration which lives in 3 dimensional ordinary space. This configuration resembles the Wu-Yang monopole and satisfies the selfduality (Bogomol'nyi) equations for a U(2) Yang-Mills-Higgs system.

  18. Compacton-like wave and kink-like wave solutions of the generalized KP-MEW (2, 2) equation

    NASA Astrophysics Data System (ADS)

    Li, Shaoyong; Song, Ming

    2014-03-01

    Using a bifurcation method and a numerical simulation approach of dynamical systems, we study the generalized KP-MEW(2, 2) equation (ut + (u2)x + (u2)xxt)x + uyy = 0. Two types of bounded traveling waves are found, that is, a compacton-like wave and a kink-like wave. The planar graphs of the compacton-like and kink-like waves are simulated using the software Maple. Exact implicit or parameter expressions of these solutions are given.

  19. Localization of genes for autosomal dominant congenital cataracts to chromosomes 2 and 17

    SciTech Connect

    Ayyagari, R.; Scott, M.; Wozencraft, L.

    1994-09-01

    Linkage analysis was performed in a seven generation family in which 28 of 52 individuals examined had autosomal dominant congenital pulverulent cataracts and a five generation family in which 10 of 17 individuals examined had autosomal dominant congenital zonular cataracts with sutural opacities. Initial analysis with 21 microsatellite markers in 7 candidate gene regions localized the pulverulent cataract locus to the long arm of chromosome 2 near the {beta}B2-crystallin gene. A lod score of 3.6 was obtained with D2S72 ({theta}=0.12), 3.5 with CRYG ({theta}=0.06), 3.4 with ({theta}=0.05), 2.0 with D2S117 ({theta}=0.22) and 6.6 with D2S128 ({theta}=0.05). Multipoint linkage analysis gave Zmax=4.2 at D2S157 with a one lod confidence interval covering 19 cM. The closest flanking markers showing obligate recombinants are D2S157 and D2S173. The zonular cataract locus was mapped to chromosome 2 near the {gamma}-crystallin gene cluster. A maximum lod score of 3.8 was obtained with D17S805 ({theta}=0.0), 2.1 with D17S798 ({theta}=0.60), and 3.7 with NF1 ({theta}=0.0). Multipoint analysis showed Zmax=3.81 at D17S805 with a one lod confidence interval covering 17 cM based on the Genethon map, localizing cataracts between markers D17S799 and D17S800. Further efforts are being directed at refining the localization of these cataract loci and examining the nearby crystallin genes for possible mutations.

  20. Microdeletions of chromosome 17p13.3 markers in an unselected survey of probands with type I lissencephaly

    SciTech Connect

    Giannakoudis, J.; Wrisch, A.; Farber, C.

    1994-09-01

    Type I lissencephaly (MIM No.247200, McKusick, 1992), a brain malformation characterized by a smooth cerebral surface, exhibits a four-layered cortex and leads to mental retardation and other neurological anomalies. Lissencephaly, type I occurs either isolated (ILS) or in association with dysmorphic facial features (Miller-Dieker syndrome, MDS). Microdeletions were detected within a 350 kb critical segment in 17p13.3 in about 13% of patients with ILS and about 90% with MDS. Most of these patients were selected for molecular analysis, however, by an already known abnormal karyotype. Therefore, the diagnostic value of microsatellite and VNTR markers to identify deletions in unselected ILS/MDS patients is still unknown. We have tested the respective significance of a novel (CA)17 VNDR element (D17S379) and of the VNTR marker YNZ22 (D17S5) to identify deletions in an unselected survey of 28 ILS/MDS patients. For D17S379, 50% of our patients were heterozygous, while 46% were uninformative with respect to segregation of alleles within their family. One patient (3.6%) was shown to be deleted for a paternal allele. PCR for D17S5, which maps proximal to the ILS region, disclosed a deletion in 3 patients (10.7%), including the one seen also by D17S379. Altogether, 75% were heterozygous and only 14% uninformative for this locus. Our results suggest that the combined PCR analysis for two of the most significant markers within the ILS/MDS region disclose a deletion in about 10% of unselected patients with features of type I lissencephaly. The low frequency of deletions detected may reflect different mutation mechanisms, genetic heterogeneity, the need for more densely spaced markers around the critical region, and/or more strict clinical criteria for defining the study group.

  1. Molecular analysis of the Smith-Magenis syndrome: a possible contiguous-gene syndrome associated with del(17)(p11.2).

    PubMed Central

    Greenberg, F; Guzzetta, V; Montes de Oca-Luna, R; Magenis, R E; Smith, A C; Richter, S F; Kondo, I; Dobyns, W B; Patel, P I; Lupski, J R

    1991-01-01

    We undertook clinical evaluation (32 cases) and molecular evaluation (31 cases) of unrelated patients affected with Smith-Magenis syndrome (SMS) associated with an interstitial deletion of band p11.2 of chromosome 17. Patients were evaluated both clinically and electrophysiologically for peripheral neuropathy, since markers showing close linkage to one form of Charcot-Marie-Tooth disease (CMT1A) map to this chromosomal region. The common clinical findings were broad flat midface with brachycephaly, broad nasal bridge, brachydactyly, speech delay, and hoarse, deep voice. Fifty-five percent of the patients showed clinical signs (e.g., decreased or absent deep tendon reflexes, pes planus or pes cavus, decreased sensitivity to pain, and decreased leg muscle mass) suggestive of peripheral neuropathy. However, unlike patients with CMT1A, these patients demonstrated normal nerve conduction velocities. Self-destructive behaviors, primarily onychotillomania and polyembolokoilamania, were observed in 67% of the patients, and significant symptoms of sleep disturbance were observed in 62%. The absence of REM sleep was demonstrated by polysomnography in two patients. Southern analysis indicated that most patients were deleted for five 17p11.2 markers--FG1 (D17S446), 1516 (D17S258), pYNM67-R5 (D17S29), pA10-41 (D17S71), and pS6.1-HB2 (D17S445)--thus defining a region which appears to be critical to SMS. The deletion was determined to be of paternal origin in nine patients and of maternal origin in six patients. The apparent random parental origin of deletion documented in 15 patients suggests that genomic imprinting does not play a role in the expression of the SMS clinical phenotype. Our findings suggest that SMS is likely a contiguous-gene deletion syndrome which comprises characteristic clinical features, developmental delay, clinical signs of peripheral neuropathy, abnormal sleep function, and specific behavioral anomalies. Images Figure 2 Figure 3 PMID:1746552

  2. Molecular analysis of the Smith-Magenis syndrome: a possible contiguous-gene syndrome associated with del(17)(p11.2).

    PubMed

    Greenberg, F; Guzzetta, V; Montes de Oca-Luna, R; Magenis, R E; Smith, A C; Richter, S F; Kondo, I; Dobyns, W B; Patel, P I; Lupski, J R

    1991-12-01

    We undertook clinical evaluation (32 cases) and molecular evaluation (31 cases) of unrelated patients affected with Smith-Magenis syndrome (SMS) associated with an interstitial deletion of band p11.2 of chromosome 17. Patients were evaluated both clinically and electrophysiologically for peripheral neuropathy, since markers showing close linkage to one form of Charcot-Marie-Tooth disease (CMT1A) map to this chromosomal region. The common clinical findings were broad flat midface with brachycephaly, broad nasal bridge, brachydactyly, speech delay, and hoarse, deep voice. Fifty-five percent of the patients showed clinical signs (e.g., decreased or absent deep tendon reflexes, pes planus or pes cavus, decreased sensitivity to pain, and decreased leg muscle mass) suggestive of peripheral neuropathy. However, unlike patients with CMT1A, these patients demonstrated normal nerve conduction velocities. Self-destructive behaviors, primarily onychotillomania and polyembolokoilamania, were observed in 67% of the patients, and significant symptoms of sleep disturbance were observed in 62%. The absence of REM sleep was demonstrated by polysomnography in two patients. Southern analysis indicated that most patients were deleted for five 17p11.2 markers--FG1 (D17S446), 1516 (D17S258), pYNM67-R5 (D17S29), pA10-41 (D17S71), and pS6.1-HB2 (D17S445)--thus defining a region which appears to be critical to SMS. The deletion was determined to be of paternal origin in nine patients and of maternal origin in six patients. The apparent random parental origin of deletion documented in 15 patients suggests that genomic imprinting does not play a role in the expression of the SMS clinical phenotype. Our findings suggest that SMS is likely a contiguous-gene deletion syndrome which comprises characteristic clinical features, developmental delay, clinical signs of peripheral neuropathy, abnormal sleep function, and specific behavioral anomalies.

  3. Haplotype analysis of BRCA1 intragenic markers in Iranian patients with familial breast and ovarian cancer

    PubMed Central

    Miresmaeili, Seyed Mohsen; Kordi Tamandani, Dor Mohammad; Kalantar, Seyed Mehdi; Moshtaghioun, Seyed Mohammad

    2016-01-01

    Background: Breast cancer is the most common malignancy in women. Breast Cancer Type 1 Susceptibility gene (BRCA1) is a tumor suppressor gene, involved in DNA damage repair and in 81% of the breast-ovarian cancer families were due to BRCA1. In some clinically investigated genes, the intragenic marker polymorphism is important and the screening of such mutations is faster by using short tandem repeat (STR) polymorphism. Individual polymorphism of STR is a good evidence for following inheritance of repeat polymorphism. Objective: The aim of this study was to evaluate three intragenic BRCA1 marker polymorphisms in families, which have two or more patients with breast/ovarian cancer in comparison to healthy women. Materials and Methods: A total of 107 breast and/or ovarian cancer patients and 93 unrelated healthy women with no clinical phenotype of any malignancy or familial cancer history constitute the study groups. Haplotyping analysis, at 3 intragenic BRCA1 microsatellite markers (D17S855, D17S1322 and D17S1323), were performed for all subject and control groups using labeled primers. Results: After fragment analysis, significance differences were observed as follows: two alleles of D17S855; allele 146 (p=0.02) and 150 (p=0.006), and two alleles of D17S1322, allele 121 (p=0.015) and 142 (p=0.043). These differences were compared with control group. There was significance difference in 8 di/tri allelic haplotypes in present experimental subjects. Some haplotypes were observed to have approximately twice the relation risk for breast cancer. Conclusion: According to recent results, assessment of presence or absence of mentioned alleles in BRCA1 microsatellite can be used for prognosis in individuals, suspected of having or not having the breast cancer. PMID:27351029

  4. Consistency and consensus models for group decision-making with uncertain 2-tuple linguistic preference relations

    NASA Astrophysics Data System (ADS)

    Zhang, Zhen; Guo, Chonghui

    2016-08-01

    Due to the uncertainty of the decision environment and the lack of knowledge, decision-makers may use uncertain linguistic preference relations to express their preferences over alternatives and criteria. For group decision-making problems with preference relations, it is important to consider the individual consistency and the group consensus before aggregating the preference information. In this paper, consistency and consensus models for group decision-making with uncertain 2-tuple linguistic preference relations (U2TLPRs) are investigated. First of all, a formula which can construct a consistent U2TLPR from the original preference relation is presented. Based on the consistent preference relation, the individual consistency index for a U2TLPR is defined. An iterative algorithm is then developed to improve the individual consistency of a U2TLPR. To help decision-makers reach consensus in group decision-making under uncertain linguistic environment, the individual consensus and group consensus indices for group decision-making with U2TLPRs are defined. Based on the two indices, an algorithm for consensus reaching in group decision-making with U2TLPRs is also developed. Finally, two examples are provided to illustrate the effectiveness of the proposed algorithms.

  5. Coevolution of Drosophila snf protein and its snRNA targets.

    PubMed

    Williams, Sandra G; Hall, Kathleen B

    2010-06-01

    SNF is a protein that is found in the U1 and U2 snRNPs (small nuclear ribonucleoproteins) of Drosophila. Its mammalian counterparts are two homologous proteins, U1A and U2B''. In vivo, these proteins segregate to the U1 and U2 snRNPs, respectively, where they bind distinct RNA hairpins. The RNA binding properties and mechanism of U1A have been studied extensively, but much less is known about SNF and U2B'' binding to their RNA targets. By comparing thermodynamic aspects of SNF-RNA interactions with those of U1A-RNA interactions, we find that SNF binds its RNA targets in a manner that is distinct from that of U1A. In vitro, SNF is able to bind both Drosophila U1 stem-loop II and U2 stem-loop IV with high affinity, although it binds stem-loop II more tightly than it binds stem-loop IV. Intriguingly, SNF is unable to bind human U2 stem-loop IV, which suggests that both the protein and RNAs have coevolved to interact with each other such that a single protein can bind RNAs that are more commonly bound by two distinct proteins.

  6. Hypoxia promotes drug resistance in osteosarcoma cells via activating AMP-activated protein kinase (AMPK) signaling

    PubMed Central

    Zhao, Changfu; Zhang, Qiao; Yu, Tao; Sun, Shudong; Wang, Wenjun; Liu, Guangyao

    2016-01-01

    Purpose Drug resistance has been recognized to be a major obstacle to the chemotherapy for osteosarcoma. And the potential importance of hypoxia as a target to reverse drug resistance in osteosarcoma has been indicated, though the mechanism underlining such role is not clarified. The present study aims to investigate the role of hypoxia in the drug resistance in osteosarcoma cells via activating AMP-activated protein kinase (AMPK) signaling. Experimental design We investigated the promotion of the resistance to doxorubicin of osteosarcoma MG-63 and U2-os cells in vitro, and then determined the role of hypoxia-inducible factor-1 (HIF-1)α and HIF-1β, the activation and regulatory role of AMPK in the osteosarcoma U2-os cells which were treated with doxorubicin under hypoxia. Results It was demonstrated that hypoxia significantly reduced the sensitivity of MG-63 and U2-os cells to doxorubicin, indicating an inhibited viability reduction and a reduced apoptosis promotion. And such reduced sensitivity was not associated with HIF-1α, though it was promoted by hypoxia in U2-os cells. Interestingly, the AMPK signaling was significantly promoted by hypoxia in the doxorubicin-treated U2-os cells, with a marked upregulation of phosphorylated AMPK (Thr 172) and phosphorylated acetyl-CoA carboxylase (ACC) (Ser 79), which were sensitive to the AMPK activator, AICAR and the AMPK inhibitor, Compound C. Moreover, the promoted AMPK activity by AICAR or the downregulated AMPK activity by Compound C significantly reduced or promoted the sensitivity of U2-os cells to doxorubicin. Conclusion The present study confirmed the AMPK signaling activation in the doxorubicin-treated osteosarcoma cells, in response to hypoxia, and the chemical upregulation or downregulation of AMPK signaling reduced or increased the chemo-sensitivity of osteosarcoma U2-os cells in vitro. Our study implies that AMPK inhibition might be a effective strategy to sensitize osteocarcoma cells to chemotherapy. PMID

  7. Ethnic differentiation at VNTR loci, with special reference to forensic applications

    SciTech Connect

    Devlin, B.; Risch, N. )

    1992-09-01

    Allele-rich VNTR loci provide valuable information for forensic inference. Interpretation of this information is complicated by measurement error, which renders discrete alleles difficult to distinguish. Two methods have been used to circumvent this difficulty-i.e, binning methods and direct evaluation of allele frequencies, the latter achieved by modeling the data as a mixture distribution. The authors use this modeling approach to estimate the allele frequency distributions for two loci-D17S79 and D2S44-for black, Caucasian, and Hispanic samples from the Lifecodes and FBI data bases. The databases are differentiated by the restriction enzyme used: PstI (Lifecodes) and HaeIII (FBI). The results show that alleles common in one ethnic group are almost always common in all ethnic groups, and likewise for rare alleles; this pattern holds for both loci. Gene diversity, or heterozygosity, measured as one minus the sum of the squared allele frequencies, is greater for D2S44 than for D17S79, in both data bases. The average gene diversity across ethnic groups when PstI (HaeIII) is used is .918 (.918) for D17S79 and is .985 (.983) for D2S44. The variance in gene diversity among ethnic groups is greater for D17S79 than for D2S44. The number of alleles, like the gene diversity, is greater for D2S44 than for D17S79. The mean numbers of alleles across ethnic groups, estimated from the PstI (HaeIII) data, are 40.25 (41.5) for D 17S79 and 104 (103) for D2S44. The number of alleles is correlated with sample size. The authors use the estimated allele frequency distributions for each ethnic group to explore the effects of unwittingly mixing populations and thereby violating independence assumptions. They show that, even in extreme cases of mixture, the estimated genotype probabilities are good estimates of the true probabilities, contradicting recent claims. 35 refs., 9 figs., 3 tabs.

  8. Systematic search for markers linked to insulin-dependent diabetes on chromosome 17

    SciTech Connect

    Williams, S.R.; Shephard, J.M.; Berger, J.S.

    1994-09-01

    We tested 19 microsatellite markers on chromosome 17 for linkage with insulin-dependent diabetes mellitus (IDDM). Nuclear families (N = 235) with at least two affected offspring were provided by the British Diabetic Association, Human Biological Data Interchange, and our own lab. The mean interval ({plus_minus}sd) between markers was 8 {plus_minus} 3 cM. For each parent heterozygous at the marker locus being studied, we determined whether the same or different alleles were transmitted to the two affected sibs. The {open_quotes}degree of sharing{close_quotes} for each marker is the frequency with which the same parental allele is transmitted to both affected sibs. Linkage with IDDM susceptibility leads to values higher than 0.50 for degree of sharing. Unlike lods, this approach makes no assumption about mode of inheritance. Mean sharing for the 19 markers was 0.51 (range 0.460 to 0.557; sd = .03). Three markers on 17q (all mutually unlinked) showed sharing of {approximately}0.55. The degree of sharing was 228/414 = 0.551 for D17A807 ({chi}{sup 2} = 4.26, p = .04), 172/309 = 0.557 for D17S784 ({chi}{sup 2} = 3.96, p = .04), and 218/398 = 0.548 for D17S798 ({chi}{sup 2} = 3.63, p =.06). We also looked for evidence of linkage disequilibrium between IDDM and alleles of each of these markers, by means of the transmission/disequilibrium test of Spielman. No significant linkage disequilibrium was found for D17S784 or D17S798. However, allele 6 of D17S807 was transmitted from heterozygous parents to IDDM offspring with frequency 131/230 = 0.57 ({chi}{sup 2} = 4.45, p = .035), supporting linkage with IDDM. We are currently investigating other markers and candidate genes in the region of D17S807.

  9. Evidence of coexistence of change of caged dynamics at T(g) and the dynamic transition at T(d) in solvated proteins.

    PubMed

    Capaccioli, S; Ngai, K L; Ancherbak, S; Paciaroni, A

    2012-02-16

    Mössbauer spectroscopy and neutron scattering measurements on proteins embedded in solvents including water and aqueous mixtures have emphasized the observation of the distinctive temperature dependence of the atomic mean square displacements, <u(2)>, commonly referred to as the dynamic transition at some temperature T(d). At low temperatures, <u(2)> increases slowly, but it assumes stronger temperature dependence after crossing T(d), which depends on the time/frequency resolution of the spectrometer. Various authors have made connection of the dynamics of solvated proteins, including the dynamic transition to that of glass-forming substances. Notwithstanding, no connection is made to the similar change of temperature dependence of <u(2)> obtained by quasielastic neutron scattering when crossing the glass transition temperature T(g), generally observed in inorganic, organic, and polymeric glass-formers. Evidences are presented here to show that such a change of the temperature dependence of <u(2)> from neutron scattering at T(g) is present in hydrated or solvated proteins, as well as in the solvent used, unsurprisingly since the latter is just another organic glass-former. If unaware of the existence of such a crossover of <u(2)> at T(g), and if present, it can be mistaken as the dynamic transition at T(d) with the ill consequences of underestimating T(d) by the lower value T(g) and confusing the identification of the origin of the dynamic transition. The <u(2)> obtained by neutron scattering at not so low temperatures has contributions from the dissipation of molecules while caged by the anharmonic intermolecular potential at times before dissolution of cages by the onset of the Johari-Goldstein β-relaxation or of the merged α-β relaxation. The universal change of <u(2)> at T(g) of glass-formers, independent of the spectrometer resolution, had been rationalized by sensitivity to change in volume and entropy of the dissipation of the caged molecules and its

  10. An in vitro investigation of bacteria-osteoblast competition on oxygen plasma-modified PEEK.

    PubMed

    Rochford, Edward T J; Subbiahdoss, Guruprakash; Moriarty, T Fintan; Poulsson, Alexandra H C; van der Mei, Henny C; Busscher, Henk J; Richards, R Geoff

    2014-12-01

    Polyetheretherketone (PEEK) films were oxygen plasma treated to increase surface free energy and characterized by X-ray photoelectron microscopy, atomic force microscopy, and water contact angles. A parallel plate flow chamber was used to measure Staphylococcus epidermidis, Staphylococcus aureus, and U-2 OS osteosarcomal cell-line adhesion to the PEEK films in separate monocultures. In addition, bacteria and U-2 OS cells were cocultured to model competition between osteoblasts and contaminating bacteria for the test surfaces. Plasma treatment of the surfaces increased surface oxygen content and decreased the hydrophobicity of the materials, but did not lead to a significant difference in bacterial or U-2 OS cell adhesion in the monocultures. In the S. epidermidis coculture experiments, the U-2 OS cells adhered in greater numbers on the treated surfaces compared to the untreated PEEK and spread to a similar extent. However, in the presence of S. aureus, cell death of the U-2 OS occurred within 10 h on all surfaces. The results of this study suggest that oxygen plasma treatment of PEEK may maintain the ability of osteoblast-like cells to adhere and spread, even in the presence of S. epidermidis contamination, without increasing the risk of preoperative bacterial adhesion. Therefore, oxygen plasma-treated PEEK remains a promising method to improve implant surface free energy for osseointegration.

  11. Using the fluorescence red edge effect to assess the long-term stability of lyophilized protein formulations.

    PubMed

    Qian, Ken K; Grobelny, Pawel J; Tyagi, Madhusudan; Cicerone, Marcus T

    2015-04-01

    Nanosecond relaxation processes in sugar matrices are causally linked through diffusional processes to protein stability in lyophilized formulations. Long-term protein degradation rates track mean-squared displacement (⟨u(2)⟩) of hydrogen atoms in sugar glasses, a parameter describing dynamics on a time scale of picoseconds to nanoseconds. However, measurements of ⟨u(2)⟩ are usually performed by neutron scattering, which is not conducive to rapid formulation screening in early development. Here, we present a benchtop technique to derive a ⟨u(2)⟩ surrogate based on the fluorescence red edge effect. Glycerol, lyophilized trehalose, and lyophilized sucrose were used as model systems. Samples containing 10(-6) mole fraction of rhodamine 6G, a fluorophore, were excited at either 532 nm (main peak) or 566 nm (red edge), and the ⟨u(2)⟩ surrogate was determined based the corresponding Stokes shifts. Results showed reasonable agreement between ⟨u(2)⟩ from neutron scattering and the surrogate from fluorescence, although deviations were observed at very low temperatures. We discuss the sources of the deviations and suggest technique improvements to ameliorate these. We expect that this method will be a valuable tool to evaluate lyophilized sugar matrices with respect to their ability to protect proteins from diffusion-limited degradation processes during long-term storage. Additionally, the method may have broader applications in amorphous pharmaceutical solids. PMID:25786057

  12. Three-dimensional evaluation of maxillary anterior alveolar bone for optimal placement of miniscrew implants

    PubMed Central

    Choi, Jin Hwan; Lee, Kee Joon; Park, Young Chel

    2014-01-01

    Objective This study aimed to propose clinical guidelines for placing miniscrew implants using the results obtained from 3-dimensional analysis of maxillary anterior interdental alveolar bone by cone-beam computed tomography (CBCT). Methods By using CBCT data from 52 adult patients (17 men and 35 women; mean age, 27.9 years), alveolar bone were measured in 3 regions: between the maxillary central incisors (U1-U1), between the maxillary central incisor and maxillary lateral incisor (U1-U2), and between the maxillary lateral incisor and the canine (U2-U3). Cortical bone thickness, labio-palatal thickness, and interdental root distance were measured at 4 mm, 6 mm, and 8 mm apical to the interdental cementoenamel junction (ICEJ). Results The cortical bone thickness significantly increased from the U1-U1 region to the U2-U3 region (p < 0.05). The labio-palatal thickness was significantly less in the U1-U1 region (p < 0.05), and the interdental root distance was significantly less in the U1-U2 region (p < 0.05). Conclusions The results of this study suggest that the interdental root regions U2-U3 and U1-U1 are the best sites for placing miniscrew implants into maxillary anterior alveolar bone. PMID:24696821

  13. Gauged twistor formulation of a massive spinning particle in four dimensions

    NASA Astrophysics Data System (ADS)

    Deguchi, Shinichi; Okano, Satoshi

    2016-02-01

    We present a gauged twistor model of a free massive spinning particle in four-dimensional Minkowski space. This model is governed by an action, referred to here as the gauged generalized Shirafuji (GGS) action, that consists of twistor variables, auxiliary variables, and U (1 ) and S U (2 ) gauge fields on the one-dimensional parameter space of a particle's worldline. The GGS action remains invariant under reparametrization and the local U (1 ) and S U (2 ) transformations of the relevant variables, although the S U (2 ) symmetry is nonlinearly realized. We consider the canonical Hamiltonian formalism based on the GGS action in the unitary gauge by following Dirac's recipe for constrained Hamiltonian systems. It is shown that just sufficient constraints for the twistor variables are consistently derived by virtue of the gauge symmetries of the GGS action. In the subsequent quantization procedure, these constraints turn into simultaneous differential equations for a twistor function. We perform the Penrose transform of this twistor function to define a massive spinor field of arbitrary rank, demonstrating that the spinor field satisfies generalized Dirac-Fierz-Pauli equations with S U (2 ) indices. We also investigate the rank-one spinor fields in detail to clarify the physical meanings of the U (1 ) and S U (2 ) symmetries.

  14. Heterologous expression and localization of gentisate transporter Ncg12922 from Corynebacterium glutamicum ATCC 13032

    SciTech Connect

    Xu Ying; Yan Dazhong; Zhou Ningyi . E-mail: n.zhou@pentium.whiov.ac.cn

    2006-07-28

    Ralstonia sp. strain U2 metabolizes naphthalene via gentisate (2,5-dihydroxybenzoate) to central metabolites, but it was found unable to utilize gentisate as growth substrate. A putative gentisate transporter encoded by ncg12922 from Corynebacterium glutamicum ATCC 13032 was functionally expressed in Ralstonia sp. strain U2, converting strain U2 to a gentisate utilizer. After ncg12922 was inserted into plasmid pGFPe with green fluorescence protein gene gfp, the expressed fusion protein Ncg12922-GFP could be visualized in the periphery of Escherichia coli cells under confocal microscope, consistent with a cytoplasmic membrane location. In contrast, GFP was ubiquitous in the cytoplasm of E. coli cells carrying pGFPe only. Gentisate 1,2-dioxygenase activity was present in the cell extract from strain U2 induced with gentisate but at a much lower level (one-fifth) than that obtained with salicylate. However, it exhibited a similar level in strain U2 containing Ncg12922 induced either by salicylate or gentisate.

  15. Exploring a new S U (4 ) symmetry of meson interpolators

    NASA Astrophysics Data System (ADS)

    Glozman, L. Ya.; Pak, M.

    2015-07-01

    In recent lattice calculations it has been discovered that mesons upon truncation of the quasizero modes of the Dirac operator obey a symmetry larger than the S U (2 )L×S U (2 )R×U (1 )A symmetry of the QCD Lagrangian. This symmetry has been suggested to be S U (4 )⊃S U (2 )L×S U (2 )R×U (1 )A that mixes not only the u- and d-quarks of a given chirality, but also the left- and right-handed components. Here it is demonstrated that bilinear q ¯q interpolating fields of a given spin J ≥1 transform into each other according to irreducible representations of S U (4 ) or, in general, S U (2 NF). This fact together with the coincidence of the correlation functions establishes S U (4 ) as a symmetry of the J ≥1 mesons upon quasizero mode reduction. It is shown that this symmetry is a symmetry of the confining instantaneous charge-charge interaction in QCD. Different subgroups of S U (4 ) as well as the S U (4 ) algebra are explored.

  16. Associations between antinuclear antibody staining patterns and clinical features of systemic lupus erythematosus: analysis of a regional Swedish register

    PubMed Central

    Frodlund, Martina; Dahlström, Örjan; Kastbom, Alf; Skogh, Thomas; Sjöwall, Christopher

    2013-01-01

    Objective Antinuclear antibody (ANA) analysis by immunofluorescence (IF) microscopy remains a diagnostic hallmark of systemic lupus erythematosus (SLE). The clinical relevance of ANA fine-specificities in SLE has been addressed repeatedly, whereas studies on IF-ANA staining patterns in relation to disease manifestations are very scarce. This study was performed to elucidate whether different staining patterns associate with distinct SLE phenotypes. Design Observational cohort study. Setting One university hospital rheumatology unit in Sweden. Participants The study population consisted of 222 cases (89% women; 93% Caucasians), where of 178 met ≥4/11 of the 1982 American College of Rheumatology (ACR-82) criteria. The remaining 20% had an SLE diagnosis based on positive IF-ANA (HEp-2 cells) and ≥2 typical organ manifestations at the time of diagnosis (Fries’ criteria). Outcome measures The IF-ANA staining patterns homogenous (H-ANA), speckled (S-ANA), combined homogenous and speckled (HS-ANA), centromeric (C-ANA), nucleolar (N-ANA)±other patterns and other nuclear patterns (oANA) were related to disease manifestations and laboratory measures. Antigen-specificities were also considered regarding double-stranded DNA (Crithidia luciliae) and the following extractable nuclear antigens: Ro/SSA, La/SSB, Smith antigen (Sm), small nuclear RNP (snRNP), Scl-70 and Jo-1 (immunodiffusion and/or line-blot technique). Results 54% of the patients with SLE displayed H-ANA, 22% S-ANA, 11% HS-ANA, 9% N-ANA, 1% C-ANA, 2% oANA and 1% were never IF-ANA positive. Staining patterns among patients meeting Fries’ criteria alone did not differ from those fulfilling ACR-82. H-ANA was significantly associated with the 10th criterion according to ACR-82 (‘immunological disorder’). S-ANA was inversely associated with arthritis, ‘immunological disorder’ and signs of organ damage. Conclusions H-ANA is the dominant IF-ANA pattern among Swedish patients with SLE, and was found to

  17. A Pre-mRNA-Splicing Factor Is Required for RNA-Directed DNA Methylation in Arabidopsis

    PubMed Central

    Huang, Chao-Feng; Miki, Daisuke; Tang, Kai; Zhou, Hao-Ran; Zheng, Zhimin; Chen, Wei; Ma, Ze-Yang; Yang, Lan; Zhang, Heng; Liu, Renyi; He, Xin-Jian; Zhu, Jian-Kang

    2013-01-01

    Cytosine DNA methylation is a stable epigenetic mark that is frequently associated with the silencing of genes and transposable elements (TEs). In Arabidopsis, the establishment of DNA methylation is through the RNA-directed DNA methylation (RdDM) pathway. Here, we report the identification and characterization of RDM16, a new factor in the RdDM pathway. Mutation of RDM16 reduced the DNA methylation levels and partially released the silencing of a reporter gene as well as some endogenous genomic loci in the DNA demethylase ros1-1 mutant background. The rdm16 mutant had morphological defects and was hypersensitive to salt stress and abscisic acid (ABA). Map-based cloning and complementation test led to the identification of RDM16, which encodes a pre-mRNA-splicing factor 3, a component of the U4/U6 snRNP. RNA-seq analysis showed that 308 intron retention events occurred in rdm16, confirming that RDM16 is involved in pre-mRNA splicing in planta. RNA-seq and mRNA expression analysis also revealed that the RDM16 mutation did not affect the pre-mRNA splicing of known RdDM genes, suggesting that RDM16 might be directly involved in RdDM. Small RNA expression analysis on loci showing RDM16-dependent DNA methylation suggested that unlike the previously reported putative splicing factor mutants, rdm16 did not affect small RNA levels; instead, the rdm16 mutation caused a decrease in the levels of Pol V transcripts. ChIP assays revealed that RDM16 was enriched at some Pol V target loci. Our results suggest that RDM16 regulates DNA methylation through influencing Pol V transcript levels. Finally, our genome-wide DNA methylation analysis indicated that RDM16 regulates the overall methylation of TEs and gene-surrounding regions, and preferentially targets Pol IV-dependent DNA methylation loci and the ROS1 target loci. Our work thus contributes to the understanding of RdDM and its interactions with active DNA demethylation. PMID:24068953

  18. The C-terminal repressor region of herpes simplex virus type 1 ICP27 is required for the redistribution of small nuclear ribonucleoprotein particles and splicing factor SC35; however, these alterations are not sufficient to inhibit host cell splicing.

    PubMed Central

    Sandri-Goldin, R M; Hibbard, M K; Hardwicke, M A

    1995-01-01

    Herpes simplex virus type 1 infection results in a reorganization of antigens associated with the small nuclear ribonucleoprotein particles (snRNPs), resulting in the formation of prominent clusters near the nuclear periphery. In this study, we show that the immediate-early protein ICP27, which is involved in the impairment of host cell splicing and in the changes in the distribution of snRNPs, is also required for reassorting the SR domain splicing factor SC35. Other viral processes, such as adsorption and penetration, shutoff of host protein synthesis, early and late gene expression, and DNA replication, do not appear to play a role in changing the staining pattern of splicing antigens. Furthermore, the C-terminal repressor region of ICP27, which is required for the inhibitory effects on splicing, also is involved in redistributing the snRNPs and SC35. During infection or transfection with five different repressor mutants, the speckled staining pattern characteristic of uninfected cells was seen and the level of a spliced target mRNA was not reduced. Infections in the presence of activator mutants showed a redistributed snRNP pattern and a decreased accumulation of spliced target mRNA. Moreover, two arginine-rich regions in the N-terminal half of ICP27 were not required for the redistribution of snRNPs or SC35. Substitution of these regions with a lysine-rich sequence from simian virus 40 large-T antigen resulted in a redistribution of splicing antigens. Unexpectedly, a repressor mutant with a ts phenotype showed a redistributed staining pattern like that seen with wild-type infected cells. During infections with this ts mutant, splicing was not inhibited, as shown in this and previous studies, confirming its repressor phenotype. Furthermore, both the mutant and the wild-type protein colocalized with snRNPs. Therefore, the redistribution of snRNPs and SC35 correlates with ICP27-mediated impairment of host cell splicing, but these alterations are not sufficient to

  19. Different Stability and Proteasome-Mediated Degradation Rate of SMN Protein Isoforms

    PubMed Central

    Locatelli, Denise; Terao, Mineko; Kurosaki, Mami; Zanellati, Maria Clara; Pletto, Daniela Rita; Finardi, Adele; Colciaghi, Francesca; Garattini, Enrico; Battaglia, Giorgio Stefano

    2015-01-01

    The key pathogenic steps leading to spinal muscular atrophy (SMA), a genetic disease characterized by selective motor neuron degeneration, are not fully clarified. The full-length SMN protein (FL-SMN), the main protein product of the disease gene SMN1, plays an established role in the cytoplasm in snRNP biogenesis ultimately leading to mRNA splicing within the nucleus. It is also involved in the mRNA axonal transport. However, to what extent the impairment of these two SMN functions contributes to SMA pathogenesis remains unknown. A shorter SMN isoform, axonal-SMN or a-SMN, with more specific axonal localization, has been discovered, but whether it might act in concert with FL-SMN in SMA pathogenesis is not known. As a first step in defining common or divergent intracellular roles of FL-SMN vs a-SMN proteins, we here characterized the turn-over of both proteins and investigated which pathway contributed to a-SMN degradation. We performed real time western blot and confocal immunofluorescence analysis in easily controllable in vitro settings. We analyzed co-transfected NSC34 and HeLa cells and cell clones stably expressing both a-SMN and FL-SMN proteins after specific blocking of transcript or protein synthesis and inhibition of known intracellular degradation pathways. Our data indicated that whereas the stability of both FL-SMN and a-SMN transcripts was comparable, the a-SMN protein was characterized by a much shorter half-life than FL-SMN. In addition, as already demonstrated for FL-SMN, the Ub/proteasome pathway played a major role in the a-SMN protein degradation. We hypothesize that the faster degradation rate of a-SMN vs FL-SMN is related to the protection provided by the protein complex in which FL-SMN is assembled. The diverse a-SMN vs FL-SMN C-terminus may dictate different protein interactions and complex formation explaining the different localization and role in the neuronal compartment, and the lower expression and stability of a-SMN. PMID:26214005

  20. Solution structure of the phosphocarrier protein HPr from Bacillus subtilis by two-dimensional NMR spectroscopy.

    PubMed Central

    Wittekind, M.; Rajagopal, P.; Branchini, B. R.; Reizer, J.; Saier, M. H.; Klevit, R. E.

    1992-01-01

    The solution structure of the phosphocarrier protein, HPr, from Bacillus subtilis has been determined by analysis of two-dimensional (2D) NMR spectra acquired for the unphosphorylated form of the protein. Inverse-detected 2D (1H-15N) heteronuclear multiple quantum correlation nuclear Overhauser effect (HMQC NOESY) and homonuclear Hartmann-Hahn (HOHAHA) spectra utilizing 15N assignments (reported here) as well as previously published 1H assignments were used to identify cross-peaks that are not resolved in 2D homonuclear 1H spectra. Distance constraints derived from NOESY cross-peaks, hydrogen-bonding patterns derived from 1H-2H exchange experiments, and dihedral angle constraints derived from analysis of coupling constants were used for structure calculations using the variable target function algorithm, DIANA. The calculated models were refined by dynamical simulated annealing using the program X-PLOR. The resulting family of structures has a mean backbone rmsd of 0.63 A (N, C alpha, C', O atoms), excluding the segments containing residues 45-59 and 84-88. The structure is comprised of a four-stranded antiparallel beta-sheet with two antiparallel alpha-helices on one side of the sheet. The active-site His 15 residue serves as the N-cap of alpha-helix A, with its N delta 1 atom pointed toward the solvent to accept the phosphoryl group during the phosphotransfer reaction with enzyme I. The existence of a hydrogen bond between the side-chain oxygen atom of Tyr 37 and the amide proton of Ala 56 is suggested, which may account for the observed stabilization of the region that includes the beta-turn comprised of residues 37-40. If the beta alpha beta beta alpha beta (alpha) folding topology of HPr is considered with the peptide chain polarity reversed, the protein fold is identical to that described for another group of beta alpha beta beta alpha beta proteins that include acylphosphatase and the RNA-binding domains of the U1 snRNP A and hnRNP C proteins. PMID:1303754

  1. Electric dipole moments of actinide atoms and RaO molecule

    SciTech Connect

    Flambaum, V. V.

    2008-02-15

    We have calculated the atomic electric dipole moments (EDMs) induced in {sup 229}Pa and {sup 225}Ac by their respective nuclear Schiff moments S. The results are d({sup 229}Pa)=-9.5x10{sup -17} [S/(e fm)]e cm=-1.1x10{sup -20}{eta} e cm and d({sup 225}Ac)=-8.6x10{sup -17} [S/(e fm)]e cm=-0.8x10{sup -21}{eta} e cm. EDM of {sup 229}Pa is 3x10{sup 4} times larger than {sup 199}Hg EDM and 40 times larger than {sup 225}Ra EDM. Possible use of actinides in solid state experiments is also discussed. The (T,P)-odd spin-axis interaction in RaO molecule is 500 times larger than in TlF.

  2. Localization of disinhibition-dementia-parkinsonism-amyotrophy complex to 17q21-22.

    PubMed Central

    Wilhelmsen, K. C.; Lynch, T.; Pavlou, E.; Higgins, M.; Nygaard, T. G.

    1994-01-01

    Disinhibition-dementia-parkinsonism-amyotrophy complex (DDPAC) is defined by familial adult-onset behavioral disturbance, followed by frontal lobe dementia, parkinsonism, and amyotrophy in variable proportions. A genetic etiology of DDPAC was suspected because of the familial clustering in family Mo, despite their wide geographic distribution. We have mapped the DDPAC locus to a 12-cM (sex averaged) region between D17S800 and D17S787 on chromosome 17q21-22. The basis for the variability of the clinical findings and pathology in DDPAC is unknown but suggests that the DDPAC locus should be screened as a candidate locus in family studies of conditions with behavioral abnormalities and neurological degeneration. PMID:7977375

  3. Characterization of 10 new polymorphic dinucleotide repeats and generation of a high-density microsatellite-based physical map of the BRCA1 region of chromosome 17q21

    SciTech Connect

    Couch, F.J.; Xu, J.; Weber, B.L.

    1994-12-01

    A familial early onset breast cancer gene (BRCA1) has been localized to chromosome 17q21. To aid in the identification of this gene a number of new microsatellite markers from the D17S857 to D17S78 region were isolated and characterized. These markers, along with previously published markers from the region, were localized on a physical map by STS content mapping of cosmids from the BRCA1 interval. This high-density STS map of the BRCA1 region will be useful for linkage studies of families with apparent inherited breast cancer and for loss of heterozygosity analysis of breast tumor DNAs. 19 refs., 2 figs., 2 tabs.

  4. Localization of disinhibition-dementia-parkinsonism-amyotrophy complex to 17q21-22

    SciTech Connect

    Wilhelmsen, K.C.; Lynch, T.; Pavlou, E.; Higgins, M.; Nygaard, T.G.

    1994-12-01

    Disinhibition-dementia-parkinsonism-amyotrophy complex (DDPAC) is defined by familial adult-onset behavioral disturbance, followed by frontal lobe dementia, parkinsonism, and amyotrophy in variable proportions. A genetic etiology of DDPAC was suspected because of the familial clustering in family Mo, despite their wide geographic distribution. We have mapped the DDPAC locus to a 12-cM (sex averaged) region between D17S800 and D17S787 on chromosome 17q21-22. The basis for the variability of the clinical findings and pathology in DDPAC is unknown but suggests that the DDPAC locus should be screened as a candidate locus in family studies of conditions with behavioral abnormalities and neurological degeneration.

  5. [Intracellular transport of nuclear ribosomal RNA in Acetabularia mediterranea].

    PubMed

    Naumova, L P; Pressman, E K; Sandakhchiev, L S

    1976-01-01

    The ribosomal RNA transport from a nucleus to a perinuclear cytoplasm and its following distribution in the cytoplasm of Acetabularia mediterranea cells were studied using transplantation of RNA-labeled rhizoid into unlabeled stalk. In addition rifamycin treatment was used for inhibition of cytoplasmic RNA synthesis. Acetabularia nuclei contain the stable RNA fractions similar to those present in some other eukaryotes. Nuclear 25S and 17S ribosomal RNA rapidly enter the rhizoid cytoplasm whereas the following trasfer of them to other regions of the cell is a very slow process. Within two days only an insignificant part of 25S and 17S ribosomal RNA is transferred from the rhizoid to the stalk and is distributed there over the base-apical gradient. No preferential transfer of the nuclear ribosomal RNA to the apical region was observed.

  6. The gene of an early onset progressive cataract (cerulean cataract) maps to 17q24

    SciTech Connect

    Armitage, M.M.; Ferrell, R.E.; Kivlin, J.D.

    1994-09-01

    Cerulean cataract is an autosomal dominant, fully penetrant, early onset, progressive cataract characterized by blue or white opacifications in the nucleus and cortex of the lens. A five generation family with 44 available affected members in three generations allowed exclusion of linkage of the cerulean cataract phenotype to lens structural protein genes and to all of the chromosomal regions to which autosomal dominant cataract phenotypes have previously been mapped. Exclusion of the plausible candidate instigated a genome-wide search utilizing short tandem repeat polymorphims. The genome search localized the cerulean cataract disease gene to chromosomal region 17q24. The three markers closest to the disease gene are D17S802 [Z({theta})=9.20 at ({theta})=0.086], D17S836 [Z({theta})=4.22 at ({theta})=0.061], and AFMa238yb5 [Z({theta})=7.11 at ({theta})=0.032]. Multipoint analysis yielded a maximum lod score of Z({theta})=11.4 between D17S802 and D17S836 at recombination rates of 0.048 and 0.013 respectively. Three genes that map near the 17q24 chromosomal region and are known to contain highly polymorphic microsatellites were tested for linkage. The genes, DHP-sensitive calcium channel gamma subunit (CACNLG), human somatastatin receptor (SSTR2), and the skeletal muscle sodium channel alpha subunit (SCN4A), were all excluded [Z({theta})=-{infinity} at ({theta})=0] as the gene causing cerulean cataract. The galactokinase (GK1) gene has not been cloned, but its map location is 17q23-q25. Galactokinase deficiency is characterized by a recessive, progressive, early onset cataract. Because of the map location of galactokinase, the age-at-onset, and progressive nature of cataracts associated with galactokinase deficiency, galactokinase is being investigated as a candidate gene for the cerulean cataract phenotype.

  7. A gene responsible for profound congenital nonsyndromal recessive deafness maps to the pericentromeric region of chromosome 17

    SciTech Connect

    Friedman, T.B.; Liang, Y.; Asher, J.H. Jr.

    1994-09-01

    Autosomal recessive deafness is the most common form of human hereditary hearing loss. Two percent of the 2,185 residents of Bengkala, Bali, Indonesia have profound congenital neurosensory nonsyndromal hereditary deafness due to a fully penetrant autosomal recessive mutation (NARD1). Families, identified through children with profound congenital deafness having hearing parents, give the expected 25% deaf progeny when corrected for ascertainment bias. Congenitally deaf individuals from Bengkala show no response to pure tone audiological examination. Obligate heterozygotes for autosomal recessive deafness in Bengkala have normal or borderline normal hearing. A chromosomal location for NARD1 was assigned directly using a linkage strategy that combines allele-frequency dependent homozygosity mapping (AHM) followed by an analysis of historical recombinants to position NARD1 relative to flanking markers. Thirteen deaf Bengkala villagers of hearing parents were typed initially for 148 STRPs distributed across the human genome and a cluster of tightly linked 17p markers with a significantly higher number of homozygotes than expected under Hardy-Weinberg and linkage equilibrium were identified. NARD1 maps closest to STRPs for D17S261 (Mfd41) and D17S805 (AFM234ta1) that are 3.2 cM apart. Recombinant genotypes for the flanking markers, D17S122 (VAW409) and D17S783 (AFM026vh7), in individuals homozygous for NARD1 place NARD1 in a 5.3 cM interval of the pericentromeric region of chromosome 17 on a refined 17p-17q12 genetic map.

  8. Cipacinoids A-D, Four Limonoids with Spirocyclic Skeletons from Cipadessa cinerascens.

    PubMed

    Yu, Jin-Hai; Liu, Qun-Fang; Sheng, Li; Wang, Guo-Cai; Li, Jia; Yue, Jian-Min

    2016-02-01

    Four limonoids, cipacinoids A-D (1-4), with spirocyclic skeletons were isolated from Cipadessa cinerascens. It is particularly notable that compounds 1-3 had a 17S-configuration for the first time in the limonoid family. Their structures with absolute configurations were assigned by spectroscopic data, X-ray crystallography, and CD analysis. Compound 1 showed moderate protein tyrosine phosphatase 1B (PTP1B) inhibition. PMID:26760061

  9. Molecular analyses of 17p11.2 deletions in 62 Smith-Magenis syndrome patients.

    PubMed Central

    Juyal, R. C.; Figuera, L. E.; Hauge, X.; Elsea, S. H.; Lupski, J. R.; Greenberg, F.; Baldini, A.; Patel, P. I.

    1996-01-01

    Smith-Magenis syndrome (SMS) is a clinically recognizable, multiple congenital anomalies/mental retardation syndrome caused by an interstitial deletion involving band p11.2 of chromosome 17. Toward the molecular definition of the interval defining this microdeletion syndrome, 62 unrelated SMS patients in conjunction with 70 available unaffected parents were molecularly analyzed with respect to the presence or absence of 14 loci in the proximal region of the short arm of chromosome 17. A multifaceted approach was used to determine deletion status at the various loci that combined (i) FISH analysis, (ii)PCR and Southern analysis of somatic cell hybrids retaining the deleted chromosome 17 from selected patients, and (iii) genotype determination of patients for whom a parent(s) was available at four microsatellite marker loci and at four loci with associated RFLPs. The relative order of two novel anonymous markers and a new microsatellite marker was determined in 17p11.2. The results confirmed that the proximal deletion breakpoint in the majority of SMS patients is located between markers D17S58 (EW301) and D17S446 (FG1) within the 17p11.1-17p11.2 region. The common distal breakpoint was mapped between markers cCI17-638, which lies distal to D17S71, and cCI17-498, which lies proximal to the Charcot Marie-Tooth disease type 1A locus. The locus D17S258 was found to be deleted in all 62 patients, and probes from this region can be used for diagnosis of the SMS deletion by FISH. Ten patients demonstrated molecularly distinct deletions; of these, two patients had smaller deletions and will enable the definition of the critical interval for SMS. Images Figure 2 PMID:8651284

  10. AIPL1 implicated in the pathogenesis of two cases of autosomal recessive retinal degeneration

    PubMed Central

    Li, David; Jin, Chongfei; Jiao, Xiaodong; Li, Lin; Bushra, Tahmina; Naeem, Muhammad Asif; Butt, Nadeem H.; Husnain, Tayyab; Sieving, Paul A.; Riazuddin, Sheikh; Riazuddin, S. Amer

    2014-01-01

    Purpose To localize and identify the gene and mutations causing autosomal recessive retinal dystrophy in two consanguineous Pakistani families. Methods Consanguineous families from Pakistan were ascertained to be affected with autosomal recessive retinal degeneration. All affected individuals underwent thorough ophthalmologic examinations. Blood samples were collected, and genomic DNA was extracted using a salting out procedure. Genotyping was performed using microsatellite markers spaced at approximately 10 cM intervals. Two-point linkage analysis was performed with the lod score method. Direct DNA sequencing of amplified genomic DNA was performed for mutation screening of candidate genes. Results Genome-wide linkage scans yielded a lod score of 3.05 at θ=0 for D17S1832 and 3.82 at θ=0 for D17S938, localizing the disease gene to a 12.22 cM (6.64 Mb) region flanked by D17S1828 and D17S1852 for family 61032 and family 61227, which contains aryl hydrocarbon receptor interacting protein-like 1 (AIPL1), a gene previously implicated in recessive Leber congenital amaurosis and autosomal dominant cone-rod dystrophy. Sequencing of AIPL1 showed a homozygous c.773G>C (p.Arg258Pro) sequence change in all affected individuals of family 61032 and a homozygous c.465G>T (p.(H93_Q155del)) change in all affected members of family 61227. Conclusions The results strongly suggest that the c.773G>C (p.R258P) and c.465G>T (p.(H93_Q155del)) mutations in AIPL1 cause autosomal recessive retinal degeneration in these consanguineous Pakistani families. PMID:24426771

  11. Role of HIV-1 matrix protein p17 variants in lymphoma pathogenesis.

    PubMed

    Dolcetti, Riccardo; Giagulli, Cinzia; He, Wangxiao; Selleri, Marina; Caccuri, Francesca; Eyzaguirre, Lindsay M; Mazzuca, Pietro; Corbellini, Silvia; Campilongo, Federica; Marsico, Stefania; Giombini, Emanuela; Muraro, Elena; Rozera, Gabriella; De Paoli, Paolo; Carbone, Antonino; Capobianchi, Maria Rosaria; Ippolito, Giuseppe; Fiorentini, Simona; Blattner, William A; Lu, Wuyuan; Gallo, Robert C; Caruso, Arnaldo

    2015-11-17

    Although in decline after successful anti-HIV therapy, B-cell lymphomas are still elevated in HIV-1-seropositive (HIV+) persons, and the mechanisms are obscure. The HIV-1 matrix protein p17 persists in germinal centers long after HIV-1 drug suppression, and some p17 variants (vp17s) activate Akt signaling and promote growth of transformed B cells. Here we show that vp17s derived from four of five non-Hodgkin lymphoma (NHL) tissues from HIV+ subjects display potent B-cell growth-promoting activity. They are characterized by amino acid insertions at position 117-118 (Ala-Ala) or 125-126 (Gly-Asn or Gly-Gln-Ala-Asn-Gln-Asn) among some other mutations throughout the sequence. Identical dominant vp17s are found in both tumor and plasma. Three of seven plasma samples from an independent set of NHL cases manifested multiple Ala insertions at position 117-118, and one with the Ala-Ala profile also promoted B-cell growth and activated Akt signaling. Ultradeep pyrosequencing showed that vp17s with C-terminal insertions are more frequently detected in plasma of HIV+ subjects with than without NHL. Insertion of Ala-Ala at position 117-118 into reference p17 (refp17) was sufficient to confer B-cell growth-promoting activity. In contrast, refp17 bearing the Gly-Asn insertion at position 125-126 did not, suggesting that mutations not restricted to the C terminus can also account for this activity. Biophysical analysis revealed that the Ala-Ala insertion mutant is destabilized compared with refp17, whereas the Gly-Asn form is stabilized. This finding provides an avenue for further exploration of structure function relationships and new treatment strategies in combating HIV-1-related NHL. PMID:26578780

  12. WITHDRAWN: (438) Efficacy of CL-108 compared to hydrocodone 7.5 mg/acetaminophen 325 mg in preventing vomiting and the use of anti-emetics, Opioid-Induced Nausea and Vomiting (OINV).

    PubMed

    Hersh, Elliot; Schachtel, Bernard; Kozarek, William; Schachtel, Emily; Marino, Mark

    2016-04-01

    The Publisher regrets that this abstract is an accidental duplication of abstract (431), also published in the 2016 American Pain Society Scientific Meeting abstracts supplement: J Pain 17:S82, 2016, http://dx.doi.org/10.1016/j.jpain.2016.01.408. The duplicate abstract (438) has therefore been withdrawn. The full Elsevier Policy on Article Withdrawal can be found at http://www.elsevier.com/locate/withdrawalpolicy. PMID:27105853

  13. WITHDRAWN: (440) Opioid-Induced Nausea and Vomiting (OINV) in Post-Operative Patients: A Comparison of CL-108 to Hydrocodone 7.5 mg/Acetaminophen 325 mg.

    PubMed

    Richardson, Stephen; Patrick, Kyle; Daniels, Stephen; Royall, Steven; Schachtel, Emily; Zhang, Bing; Marino, Mark; Schachtel, Bernard

    2016-04-01

    The Publisher regrets that this abstract is an accidental duplication of abstract (436), also published in the 2016 American Pain Society Scientific Meeting abstracts supplement: J Pain 17:S83, 2016, http://dx.doi.org/10.1016/j.jpain.2016.01.413. The duplicate abstract (440) has therefore been withdrawn. The full Elsevier Policy on Article Withdrawal can be found at http://www.elsevier.com/locate/withdrawalpolicy. PMID:27105852

  14. Nephropathic cystinosis (CTNS-LSB): construction of a YAC contig comprising the refined critical region on chromosome 17p13.

    PubMed

    Peters, U; Senger, G; Rählmann, M; Du Chesne, I; Stec, I; Köhler, M R; Weissenbach, J; Leal, S M; Koch, H G; Deufel, T; Harms, E

    1997-01-01

    A yeast artificial chromosome (YAC) contig was constructed encompassing the entire region on chromosome 17p13 where the autosomal recessive disorder infantile nephropathic cystinosis (MIM 21980, CTNS-LSB) has been genetically mapped. It comprises seven clones ordered by their content of a series of six sequence-tagged sites (STSs). Fluorescence in situ hybridisation (FISH) revealed two chimaeric clones. The order of four polymorphic STSs mapped with the contig was consistent with that of the known genetic map with the exception of markers D17S1583 (AFMb307zg5) and D17S1798 (AFMa202xf5) where a telomeric location of D17S1583 was inferred from the contig; two non-polymorphic STSs were localised within the marker frame-work. From the analysis of recombination events in an unaffected individual as defined by leucocyte cystine levels we support the high-resolution mapping of this region to a small genetic interval and show that it is entirely represented on a single, non-chimaeric YAC clone in the contig.

  15. Condensation nuclei measurement in the stratosphere for the NASA ACE program

    NASA Astrophysics Data System (ADS)

    Wilson, James Charles

    1994-11-01

    A condensation nucleus counter which operated at stratospheric pressures was developed, designed, and constructed. It was calibrated in the laboratory. Its response as a function of particle size and concentration was reported. This was the first time that the response of such an instrument was verified in the laboratory. An inlet was constructed which provided near isokinetic sampling. The resulting instrument, the U-2 CNC, was deployed on NASA U-2 aircraft in the study of the climatic effects of aerosol. These studies occurred in March, April, May, July, November, and December of 1992 and in April, May, June, and December of 1983. The U-2 CNC was used in the study of the aerosol cloud resulting from the eruption of El Chichon. It permitted the observation of new particle formation in the stratosphere.

  16. Condensation nuclei measurement in the stratosphere for the NASA ACE program. Final report, 1 February 1980-31 January 1987

    SciTech Connect

    Wilson, J.C.

    1994-11-01

    A condensation nucleus counter which operated at stratospheric pressures was developed, designed, and constructed. It was calibrated in the laboratory. Its response as a function of particle size and concentration was reported. This was the first time that the response of such an instrument was verified in the laboratory. An inlet was constructed which provided near isokinetic sampling. The resulting instrument, the U-2 CNC, was deployed on NASA U-2 aircraft in the study of the climatic effects of aerosol. These studies occurred in March, April, May, July, November, and December of 1992 and in April, May, June, and December of 1983. The U-2 CNC was used in the study of the aerosol cloud resulting from the eruption of El Chichon. It permitted the observation of new particle formation in the stratosphere.

  17. Condensation nuclei measurement in the stratosphere for the NASA ACE program

    NASA Technical Reports Server (NTRS)

    Wilson, James Charles

    1994-01-01

    A condensation nucleus counter which operated at stratospheric pressures was developed, designed, and constructed. It was calibrated in the laboratory. Its response as a function of particle size and concentration was reported. This was the first time that the response of such an instrument was verified in the laboratory. An inlet was constructed which provided near isokinetic sampling. The resulting instrument, the U-2 CNC, was deployed on NASA U-2 aircraft in the study of the climatic effects of aerosol. These studies occurred in March, April, May, July, November, and December of 1992 and in April, May, June, and December of 1983. The U-2 CNC was used in the study of the aerosol cloud resulting from the eruption of El Chichon. It permitted the observation of new particle formation in the stratosphere.

  18. Mott lobes evolution of the spin-1 Bose-Hubbard model

    NASA Astrophysics Data System (ADS)

    Hincapie-F, A. F.; Franco, R.; Silva-Valencia, J.

    2016-02-01

    We study spin-1 bosons confined in a one-dimensional optical lattice, taking into consideration both ferromagnetic and antiferromagnetic interaction. Using the density matrix renormalization group, we determine the phase diagram for the two firsts lobes and report the evolution of the first and second Mott lobes with respect to the spin-exchange interaction parameter (U 2). We determine that for the antiferromagnetic case, the first lobe is suppressed while the second grows as |U 2| increases. For the ferromagnetic case, the first and second Mott lobes are suppressed by the spin-exchange interaction parameter. We propose an expresion to describe the evolution of the critical point with the increase in |U 2| for both cases.

  19. Effects of polydeoxyribonucleotides (PDRN) on wound healing: Electric cell-substrate impedance sensing (ECIS).

    PubMed

    Koo, Youngmi; Yun, Yeoheung

    2016-12-01

    Polydeoxyribonucleotides (PDRN) have been explored as an effective treatment for tissue repair in peripheral artery occlusive disease, diabetic foot ulcers, and eye lotion. We report on the effect of polydeoxyribonucleotides (PDRN) on wound healing by using the electric cell-substrate impedance sensing (ECIS) system and viability testing. Human osteoblasts (U2OS) and primary human dermal fibroblasts (HDF) were used to study the effect of PDRN on migration and proliferation. ECIS allowed the creation of a wound by applying high current, and then monitoring the healing process by measuring impedance in real time. The traditional culture-insert gap-closure migration assay was performed and compared with the ECIS wound assay. PDRN-treated U2OS and HDF cells affected cell motilities to wounding site. Viability test results show that HDF and U2OS proliferation depended on PDRN concentration. Based on the results, a PDRN compound can be useful in wound healing associated with bone and skin.

  20. New mixing angles in the left-right symmetric model

    NASA Astrophysics Data System (ADS)

    Kokado, Akira; Saito, Takesi

    2015-12-01

    In the left-right symmetric model neutral gauge fields are characterized by three mixing angles θ12,θ23,θ13 between three gauge fields Bμ,WLμ 3,WRμ 3, which produce mass eigenstates Aμ,Zμ,Zμ', when G =S U (2 )L×S U (2 )R×U (1 )B-L×D is spontaneously broken down until U (1 )em . We find a new mixing angle θ', which corresponds to the Weinberg angle θW in the standard model with the S U (2 )L×U (1 )Y gauge symmetry, from these mixing angles. It is then shown that any mixing angle θi j can be expressed by ɛ and θ', where ɛ =gL/gR is a ratio of running left-right gauge coupling strengths. We observe that light gauge bosons are described by θ' only, whereas heavy gauge bosons are described by two parameters ɛ and θ'.

  1. Effects of polydeoxyribonucleotides (PDRN) on wound healing: Electric cell-substrate impedance sensing (ECIS).

    PubMed

    Koo, Youngmi; Yun, Yeoheung

    2016-12-01

    Polydeoxyribonucleotides (PDRN) have been explored as an effective treatment for tissue repair in peripheral artery occlusive disease, diabetic foot ulcers, and eye lotion. We report on the effect of polydeoxyribonucleotides (PDRN) on wound healing by using the electric cell-substrate impedance sensing (ECIS) system and viability testing. Human osteoblasts (U2OS) and primary human dermal fibroblasts (HDF) were used to study the effect of PDRN on migration and proliferation. ECIS allowed the creation of a wound by applying high current, and then monitoring the healing process by measuring impedance in real time. The traditional culture-insert gap-closure migration assay was performed and compared with the ECIS wound assay. PDRN-treated U2OS and HDF cells affected cell motilities to wounding site. Viability test results show that HDF and U2OS proliferation depended on PDRN concentration. Based on the results, a PDRN compound can be useful in wound healing associated with bone and skin. PMID:27612747

  2. Oleocanthal Modulates Estradiol-Induced Gene Expression Involving Estrogen Receptor α.

    PubMed

    Keiler, Annekathrin Martina; Djiogue, Sefirin; Ehrhardt, Tino; Zierau, Oliver; Skaltsounis, Leandros; Halabalaki, Maria; Vollmer, Günter

    2015-09-01

    Oleocanthal is a bioactive compound from olive oil. It has attracted considerable attention as it is anti-inflammatory, antiproliferative, and has been shown to possess neuroprotective properties in vitro and in vivo. Delineated from its polyphenolic structure, the aim of this study was to characterize oleocanthal towards estrogenic properties. This might contribute to partly explain the beneficial effects described for the Mediterranean diet. Estrogenic properties of oleocanthal were assessed by different methods: a) stimulation of reporter gene activity in MVLN or RNDA cells either expressing estrogen receptor α or β, b) stimulation of luciferase reporter gene activity in U2OS osteosarcoma cells expressing estrogen receptor α or β, and c) elucidation of the impact on estradiol-induced gene expression in U2OS cells transduced with both estrogen receptors. Depending on the cell line origin, oleocanthal inhibited luciferase activity (MVLN, U2OS-estrogen receptor β) or weakly induced reporter gene activity at 10 µM in U2OS-estrogen receptor α cells. However, oleocanthal inhibited stimulation of luciferase activity by estradiol from both estrogen receptors. Oleocanthal, if given alone, did not stimulate gene expression in U2OS cells, but it significantly modulated the response of estradiol. Oleocanthal enhanced the effect of estradiol on the regulation of those genes, which are believed to be regulated through heterodimeric estrogen receptors. As the estrogenic response pattern of oleocanthal is rather unique, we compared the results obtained with oleacein. Oleocanthal binds to both estrogen receptors inducing estradiol-agonistic or antiagonistic effects depending on the cell line. Regarding regulation of gene expression in U2OS-estrogen receptor α/β cells, oleocanthal and oleacein enhanced estradiol-mediated regulation of heterodimer-regulated genes. PMID:26166135

  3. Initial Incidence of White Matter Hyperintensities on MRI in Astronauts

    NASA Technical Reports Server (NTRS)

    Norcross, Jason; Sherman, Paul; McGuire, Steve; Kochunov, Peter

    2016-01-01

    Introduction: Previous literature has described the increase in white matter hyperintensity (WMH) burden associated with hypobaric exposure in the U-2 and altitude chamber operating personnel. Although astronauts have similar hypobaric exposure pressures to the U2 pilot population, astronauts have far fewer exposures and each exposure would be associated with a much lower level of decompression stress due to rigorous countermeasures to prevent decompression sickness. Therefore, we postulated that the WMH burden in the astronaut population would be less than in U2 pilots. Methods: Twenty-one post-flight de-identified astronaut MRIs (5 mm slice thickness FLAIR sequences) were evaluated for WMH count and volume. The only additional data provided was an age range of the astronauts (43-57) and if they had ever performed an EVA (13 yes, 8 no). Results: WMH count in these 21 astronaut MRI was 21.0 +/- 24.8 (mean+/- SD) and volume was 0.382 +/- 0.602 ml, which was significantly higher than previously published results for the U2 pilots. No significant differences between EVA and no EVA groups existed. Age range of astronaut population is not directly comparable to the U2 population. Discussion: With significantly less frequent (sometimes none) and less stressful hypobaric exposures, yet a much higher incidence of increased WMH, this indicates the possibility of additional mechanisms beyond hypobaric exposure. This increase unlikely to be attributable just to the differences in age between astronauts and U2 pilots. Forward work includes continuing review of post-flight MRI and evaluation of pre to post flight MRI changes if available. Data mining for potential WMH risk factors includes collection of age, sex, spaceflight experience, EVA hours, other hypobaric exposures, hyperoxic exposures, radiation, high performance aircraft experience and past medical history. Finally, neurocognitive and vision/eye results will be evaluated for any evidence of impairment linked to

  4. Chiral Lagrangians from lattice gauge theories in the strong coupling limit

    SciTech Connect

    Nagao, Taro; Nishigaki, Shinsuke M.

    2001-07-01

    We derive nonlinear {sigma} models (chiral Lagrangians) over symmetric spaces U(n), U(2n)/Sp(2n), and U(2n)/O(2n) from U(N), O(N), and Sp(2N) lattice gauge theories coupled to n flavors of staggered fermions, in the large-N and g{sup 2}N limit. To this end, we employ Zirnbauer{close_quote}s color-flavor transformation. We prove the spatial homogeneity of the vacuum configurations of mesons by explicitly solving the large-N saddle point equations, and thus establish these patterns of spontaneous chiral symmetry breaking in the above limit.

  5. A search variability in the UV spectrum of Pi Aquarii and Fe 3 shell lines of Be stars

    NASA Technical Reports Server (NTRS)

    1984-01-01

    Several short U1 and U2 observations of Be stars are obtained with the Copernicus satellite. Pi Aquarii (B1 IV-Ve) is observed with the U1 and U2 spectrometers. These scans are compared with earlier observations. Variations in the strengths and profiles of selected shell and photospheric features are examined. In order to study possible changes in the temperature of the circumstellar envelope, features covering a wide range in ionization are observed. Included in the observing program are lines of O VI, N V, Si IV, Si III, S III, Fe III, and N I.

  6. NOSL experiment support

    NASA Technical Reports Server (NTRS)

    Brook, M.

    1986-01-01

    An optical lightning detector was constructed and flown, along with Vinton cameras and a Fairchild Line Scan Spectrometer, on a U-2 during the summer of 1979. The U-2 lightning data was obtained in daylight, and was supplemented with ground truth taken at Langmuir Laboratory. Simulations were prepared as required to establish experiment operating procedures and science training for the astronauts who would operate the Night/Day Optical Survey of Thunderstorm Lightning (NOSL) equipment during the STS-2 NOSL experiment on the Space Shuttle. Data was analyzed and papers were prepared for publication.

  7. Stratospheric air sampling platform/sensor tradeoffs

    NASA Technical Reports Server (NTRS)

    Arno, R. D.; Page, W.

    1976-01-01

    Results of a study are described in which in-situ and remote sensing instrumentation are considered for accommodation on airborne platforms capable of reaching stratospheric altitudes. The instrumentation measures trace species of importance to present concerns regarding stratospheric pollution and possible ozone depletion. The platforms examined were the U-2, modified U-2, balloon, rocket, F-15 flown in a zoom-climb maneuver, YF-12, and remotely piloted vehicle (RPV). The sensors and performance characteristics of the platforms are described and special problems of sensor-platform integration are discussed. A typical latitudinal sampling mission is utilized to describe platform logistics problems and how the platforms might perform such missions.

  8. Cycloalkyl-based unsymmetrical unsaturated (U₂)-NHC ligands: flexibility and dissymmetry in ruthenium-catalysed olefin metathesis.

    PubMed

    Rouen, Mathieu; Borré, Etienne; Falivene, Laura; Toupet, Loic; Berthod, Mikaël; Cavallo, Luigi; Olivier-Bourbigou, Hélène; Mauduit, Marc

    2014-05-21

    Air-stable Ru-indenylidene and Hoveyda-type complexes bearing new unsymmetrical unsaturated N-heterocyclic carbene (U2-NHC) ligands combining a mesityl unit and a flexible cycloalkyl moiety as N-substituents were synthesised. Structural features, chemical stabilities and catalytic profiles in olefin metathesis of this new library of cycloalkyl-based U2-NHC Ru complexes were studied and compared with their unsymmetrical saturated NHC-Ru homologues as well as a set of commercially available Ru-catalysts bearing either symmetrical SIMes or IMes NHC ligands. PMID:24647372

  9. Cycloalkyl-based unsymmetrical unsaturated (U₂)-NHC ligands: flexibility and dissymmetry in ruthenium-catalysed olefin metathesis.

    PubMed

    Rouen, Mathieu; Borré, Etienne; Falivene, Laura; Toupet, Loic; Berthod, Mikaël; Cavallo, Luigi; Olivier-Bourbigou, Hélène; Mauduit, Marc

    2014-05-21

    Air-stable Ru-indenylidene and Hoveyda-type complexes bearing new unsymmetrical unsaturated N-heterocyclic carbene (U2-NHC) ligands combining a mesityl unit and a flexible cycloalkyl moiety as N-substituents were synthesised. Structural features, chemical stabilities and catalytic profiles in olefin metathesis of this new library of cycloalkyl-based U2-NHC Ru complexes were studied and compared with their unsymmetrical saturated NHC-Ru homologues as well as a set of commercially available Ru-catalysts bearing either symmetrical SIMes or IMes NHC ligands.

  10. New Dust Belts of Uranus: One Ring, Two Ring, Red Ring, Blue Ring

    SciTech Connect

    de Pater, I; Hammel, H B; Gibbard, S G; Showalter, M R

    2006-02-02

    We compare near-infrared observations of the recently discovered outer rings of Uranus with HST results. We find that the inner ring, R/2003 U 2, is red, whereas the outer ring, R/2003 U 1, is very blue. Blue is an unusual color for rings; Saturn's enigmatic E ring is the only other known example. By analogy to the E ring, R/2003 U 1 is probably produced via impacts into the embedded moon Mab, which apparently orbits at a location where non-gravitational perturbations favor the survival and spreading of sub-micron sized dust. R/2003 U 2 more closely resembles Saturn's G ring.

  11. Ocean color imagery: Coastal zone color scanner

    NASA Technical Reports Server (NTRS)

    Hovis, W. A.

    1975-01-01

    Investigations into the feasibility of sensing ocean color from high altitude for determination of chlorophyll and sediment distributions were carried out using sensors on NASA aircraft, coordinated with surface measurements carried out by oceanographic vessels. Spectrometer measurements in 1971 and 1972 led to development of an imaging sensor now flying on a NASA U-2 and the Coastal Zone Color Scanner to fly on Nimbus G in 1978. Results of the U-2 effort show the imaging sensor to be of great value in sensing pollutants in the ocean.

  12. The Impact of Relationship Education on Adolescents of Diverse Backgrounds

    ERIC Educational Resources Information Center

    Adler-Baeder, Francesca; Kerpelman, Jennifer L.; Schramm, David G.; Higginbotham, Brian; Paulk, Amber

    2007-01-01

    Adolescent-focused marriage education is a relatively uncharted research area. Using a quasi-experimental design, this study examined the effectiveness of an adapted version of the curriculum entitled, "Love U2: Increasing Your Relationship Smarts" with an economically, geographically, and racially diverse sample of 340 high school students.…

  13. miR-1 Inhibits Cell Growth, Migration, and Invasion by Targeting VEGFA in Osteosarcoma Cells

    PubMed Central

    Niu, Junjie; Guo, Qiaoge; Niu, Dongju; Liu, Bo

    2016-01-01

    microRNAs (miRNAs) are small noncoding RNAs and have been shown to play a crucial role in the osteosarcoma (OS) tumorigenesis and progression. VEGFA is a key regulator of angiogenesis and plays an important role in regulation of tumor metastasis. The objective of this study was to determine whether VEGFA was involved in miR-1-mediated suppression of proliferation, migration, and invasion of OS cells. The expression levels of miR-1 were significantly lower in OS tumor tissues than those in adjacent normal tissues and in SAOS-2 and U2OS cell lines compared to a normal osteoblast (NHOst) cell line. VEGFA was upregulated in OS tumor tissues and SAOS-2 and U2OS cell lines. The results of CCK-8 assay and transwell assay showed that miR-1 acted as a tumor suppressor by inhibiting cell proliferation, migration, and invasion in U2OS cells. Dual luciferase reporter assay demonstrated that VEGFA was a direct and functional target gene of miR-1. miR-1 directly inhibits the protein expression of VEGFA via its 3′-UTR. Knockdown of VEGFA by siRNA inhibited proliferation, migration, and invasion of U2OS cells. Our study suggested the potential inhibitory function of miR-1 in OS cell proliferation, migration, and invasion via inhibiting VEGFA. PMID:27777493

  14. Evolution of a tissue-specific splicing network.

    PubMed

    Taliaferro, J Matthew; Alvarez, Nehemiah; Green, Richard E; Blanchette, Marco; Rio, Donald C

    2011-03-15

    Alternative splicing of precursor mRNA (pre-mRNA) is a strategy employed by most eukaryotes to increase transcript and proteomic diversity. Many metazoan splicing factors are members of multigene families, with each member having different functions. How these highly related proteins evolve unique properties has been unclear. Here we characterize the evolution and function of a new Drosophila splicing factor, termed LS2 (Large Subunit 2), that arose from a gene duplication event of dU2AF(50), the large subunit of the highly conserved heterodimeric general splicing factor U2AF (U2-associated factor). The quickly evolving LS2 gene has diverged from the splicing-promoting, ubiquitously expressed dU2AF(50) such that it binds a markedly different RNA sequence, acts as a splicing repressor, and is preferentially expressed in testes. Target transcripts of LS2 are also enriched for performing testes-related functions. We therefore propose a path for the evolution of a new splicing factor in Drosophila that regulates specific pre-mRNAs and contributes to transcript diversity in a tissue-specific manner.

  15. Defect Induced Electronic Structure of Uranofullerene

    PubMed Central

    Dai, Xing; Cheng, Cheng; Zhang, Wei; Xin, Minsi; Huai, Ping; Zhang, Ruiqin; Wang, Zhigang

    2013-01-01

    The interaction between the inner atoms/cluster and the outer fullerene cage is the source of various novel properties of endohedral metallofullerenes. Herein, we introduce an adatom-type spin polarization defect on the surface of a typical endohedral stable U2@C60 to predict the associated structure and electronic properties of U2@C61 based on the density functional theory method. We found that defect induces obvious changes in the electronic structure of this metallofullerene. More interestingly, the ground state of U2@C61 is nonet spin in contrast to the septet of U2@C60. Electronic structure analysis shows that the inner U atoms and the C ad-atom on the surface of the cage contribute together to this spin state, which is brought about by a ferromagnetic coupling between the spin of the unpaired electrons of the U atoms and the C ad-atom. This discovery may provide a possible approach to adapt the electronic structure properties of endohedral metallofullerenes. PMID:23439318

  16. Spindle structure and function changes of the human cells after simulated microgravity

    NASA Astrophysics Data System (ADS)

    Li, Yu; Wei, Lijun; Qi, Jing; Yan, Xing; Wang, Hongying; Feng, Hui; Yue, Lei; Zhang, Yao

    The main induce factors of the space environment are sustaining microgravity and ionizing radiation which can influence cell structure and function. In this study, for investigating the damage to human proliferation cells under microgravity effects, human osteosarcoma cell lines (MG-63 and U-2 OS) and peripheral blood lymphocytes were analyzed after ground-based simulated microgravity effect with clinostat. The present results showed that under the simu-lated microgravity effect for 72 hours the cytoskeleton both in human osteosarcoma cell lines and peripheral blood lymphocytes were in disorder and the spindle structure changed. The multiple polar spindle rates in human osteosarcoma cell lines MG-63 and U-2 OS increased in a linear relationship with simulated microgravity hours (0 to 96 hours). The mitotic index increased in human osteosarcoma cell lines MG-63 and U-2 OS, while decreased in human pe-ripheral blood lymphocytes. And the cell cycle of MG-63 arrest at G2/M phase. The mode number of chromosomes also varied in MG-63, while the chromosome number of human periph-eral blood lymphocytes didn't show significant difference. The Real-time quantitative reverse transcription PCR (qRT-PCR) and Western blot showed that the spindle assembly checkpoint protein MAD2 and BUB1 in MG-63 and U-2 OS changed with the spindle structure change and chromosome number change.

  17. Nobiletin inhibits human osteosarcoma cells metastasis by blocking ERK and JNK-mediated MMPs expression.

    PubMed

    Cheng, Hsin-Lin; Hsieh, Ming-Ju; Yang, Jia-Sin; Lin, Chiao-Wen; Lue, Ko-Haung; Lu, Ko-Hsiu; Yang, Shun-Fa

    2016-06-01

    Nobiletin, a polymethoxyflavone, has a few pharmacological activities, including anti-inflammation and anti-cancer effects. However, its effect on human osteosarcoma progression remains uninvestigated. Therefore, we examined the effectiveness of nobiletin against cellular metastasis of human osteosarcoma and the underlying mechanisms. Nobiletin, up to 100 μM without cytotoxicity, significantly decreased motility, migration and invasion as well as enzymatic activities, protein levels and mRNA expressions of matrix metalloproteinase (MMP)-2 and MMP-9 in U2OS and HOS cells. In addition to inhibition of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), the inhibitory effect of nobiletin on the DNA-binding activity of the transcription factor nuclear factor-kappa B (NF-κB), cAMP response element-binding protein (CREB), and specificity protein 1 (SP-1) in U2OS and HOS cells. Co-treatment with ERK and JNK inhibitors and nobiletin further reduced U2OS cells migration and invasion. These results indicated that nobiletin inhibits human osteosarcoma U2OS and HOS cells motility, migration and invasion by down-regulating MMP-2 and MMP-9 expressions via ERK and JNK pathways and through the inactivation of downstream NF-κB, CREB, and SP-1. Nobiletin has the potential to serve as an anti-metastatic agent for treating osteosarcoma.

  18. Osteoblast integration of dental implant materials after challenge by sub-gingival pathogens: a co-culture study in vitro.

    PubMed

    Zhao, Bingran; van der Mei, Henny C; Rustema-Abbing, Minie; Busscher, Henk J; Ren, Yijin

    2015-12-01

    Sub-gingival anaerobic pathogens can colonize an implant surface to compromise osseointegration of dental implants once the soft tissue seal around the neck of an implant is broken. In vitro evaluations of implant materials are usually done in monoculture studies involving either tissue integration or bacterial colonization. Co-culture models, in which tissue cells and bacteria battle simultaneously for estate on an implant surface, have been demonstrated to provide a better in vitro mimic of the clinical situation. Here we aim to compare the surface coverage by U2OS osteoblasts cells prior to and after challenge by two anaerobic sub-gingival pathogens in a co-culture model on differently modified titanium (Ti), titanium-zirconium (TiZr) alloys and zirconia surfaces. Monoculture studies with either U2OS osteoblasts or bacteria were also carried out and indicated significant differences in biofilm formation between the implant materials, but interactions with U2OS osteoblasts were favourable on all materials. Adhering U2OS osteoblasts cells, however, were significantly more displaced from differently modified Ti surfaces by challenging sub-gingival pathogens than from TiZr alloys and zirconia variants. Combined with previous work employing a co-culture model consisting of human gingival fibroblasts and supra-gingival oral bacteria, results point to a different material selection to stimulate the formation of a soft tissue seal as compared to preservation of osseointegration under the unsterile conditions of the oral cavity. PMID:26674427

  19. Higgs-like boson at 750 GeV and genesis of baryons

    NASA Astrophysics Data System (ADS)

    Davoudiasl, Hooman; Giardino, Pier Paolo; Zhang, Cen

    2016-07-01

    We propose that the diphoton excess at 750 GeV reported by ATLAS and CMS is due to the decay of an exo-Higgs scalar η associated with the breaking of a new S U (2 )e symmetry, dubbed exo-spin. New fermions, exo-quarks and exo-leptons, get TeV-scale masses through Yukawa couplings with η and generate its couplings to gluons and photons at one loop. The matter content of our model yields a B -L anomaly under S U (2 )e, whose breaking we assume entails a first-order phase transition. A nontrivial B -L asymmetry may therefore be generated in the early Universe, potentially providing a baryogenesis mechanism through the Standard Model (SM) sphaleron processes. The spontaneous breaking of S U (2 )e can, in principle, directly lead to electroweak symmetry breaking, thereby accounting for the proximity of the mass scales of the SM Higgs and the exo-Higgs. Our model can be distinguished from those comprising a singlet scalar and vector fermions by the discovery of TeV scale exo-vector bosons, corresponding to the broken S U (2 )e generators, at the LHC.

  20. 57. Photocopy of construction drawing, Arizona Highway Department, May 1927, ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    57. Photocopy of construction drawing, Arizona Highway Department, May 1927, microfiche of original drawing located at Arizona Department of Transportation, Phoenix AZ). ARCH DETAIL, U0 - U2. - Navajo Bridge, Spanning Colorado River at U.S. Highway 89 Alternate, Page, Coconino County, AZ

  1. 10 CFR 455.71 - Eligibility.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... on the State's requirement for determining cost effectiveness) of any energy conservation measure... payback has been selected as the cost-effectiveness test pursuant to § 455.20(u)(2), the simple payback...) Have no plan or intention at the time of application to close or otherwise dispose of the building...

  2. 10 CFR 455.71 - Eligibility.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... on the State's requirement for determining cost effectiveness) of any energy conservation measure... payback has been selected as the cost-effectiveness test pursuant to § 455.20(u)(2), the simple payback...) Have no plan or intention at the time of application to close or otherwise dispose of the building...

  3. Remote sensing research for agricultural applications

    NASA Technical Reports Server (NTRS)

    Colwell, R. N. (Principal Investigator)

    1983-01-01

    The thematic mapper simulator (TMS) flown by the U-2/ER-2 aircraft is being used as a surrogate for LANDSAT-4TM data. Progress is reported on spectral data acquisition including TMS, color infrared high altitude aerial photography, and LANDSAT 3 MSS and ground data collection to support classification and testing. A test site in San Joaquin County was selected for analysis.

  4. Evaluation of remote sensing in control of pink cotton bollworm

    NASA Technical Reports Server (NTRS)

    Lewis, L. N. (Principal Investigator); Coleman, V. B.

    1973-01-01

    The author has identified the following significant results. The U-2 underflight photography has shown that the critical stages in cotton plow down (defoliation, shredding, and plowing) can be identified. This result will prove invaluable to a user agency whose purpose is to monitor the cotton season for compliance with California State law.

  5. Effect of different compounds on the induction of laccase production by Agaricus blazei.

    PubMed

    Valle, J S; Vandenberghe, L P S; Oliveira, A C C; Tavares, M F; Linde, G A; Colauto, N B; Soccol, C R

    2015-12-03

    Laccases are polyphenol oxidases produced by many fungi and have many applications in textile, food and beverage, and pulp and paper industries. Laccase production can be induced using aromatic or phenolic compounds that mostly affect the transcription of laccase-encoding genes. In this study, we analyzed laccase and biomass production by Agaricus blazei in the presence of different concentrations of nitrogen, copper, and inducers such as pyrogallol, veratryl alcohol, xylidine, vanillin, guaiacol, and ethanol. Laccase production by A. blazei U2-4 reached 43.8 U/mL in the presence of 2.8 g/L nitrogen and 150 μM copper. However, addition of copper to the cultivation medium decreased biomass production. Different compounds differentially induced laccase production by A. blazei. Moreover, different concentrations of these inducers exerted different effects on laccase activity. Ethanol (1.0 mM), guaiacol (0.5 mM), and vanillin (0.5 mM) were the best inducers and increased laccase activity by 120% (A. blazei U2-2), 30% (A. blazei U2-3), and 9% (A. blazei U2-4), respectively. In contrast, pyrogallol and xylidine decreased laccase activity but increased biomass production.

  6. Quantum abacus

    NASA Astrophysics Data System (ADS)

    Cheon, Taksu; Tsutsui, Izumi; Fülöp, Tamás

    2004-09-01

    We show that the point interactions on a line can be utilized to provide U(2) family of qubit operations for quantum information processing. Qubits are realized as states localized in either side of the point interaction which represents a controllable gate. The qubit manipulation proceeds in a manner analogous to the operation of an abacus.

  7. Conservation of functional domains involved in RNA binding and protein-protein interactions in human and Saccharomyces cerevisiae pre-mRNA splicing factor SF1.

    PubMed

    Rain, J C; Rafi, Z; Rhani, Z; Legrain, P; Krämer, A

    1998-05-01

    The modular structure of splicing factor SF1 is conserved from yeast to man and SF1 acts at early stages of spliceosome assembly in both organisms. The hnRNP K homology (KH) domain of human (h) SF1 is the major determinant for RNA binding and is essential for the activity of hSF1 in spliceosome assembly, supporting the view that binding of SF1 to RNA is essential for its function. Sequences N-terminal to the KH domain mediate the interaction between hSF1 and U2AF65, which binds to the polypyrimidine tract upstream of the 3' splice site. Moreover, yeast (y) SF1 interacts with Mud2p, the presumptive U2AF65 homologue in yeast, and the interaction domain is conserved in ySF1. The C-terminal degenerate RRMs in U2AF65 and Mud2p mediate the association with hSF1 and ySF1, respectively. Analysis of chimeric constructs of hSF1 and ySF indicates that the KH domain may serve a similar function in both systems, whereas sequences C-terminal to the KH domain are not exchangeable. Thus, these results argue for hSF1 and ySF1, as well as U2AF65 and Mud2p, being functional homologues.

  8. Thermal compatibility studies of unirradiated UMo alloys dispersed in aluminum

    NASA Astrophysics Data System (ADS)

    Lee, Don Bae; Kim, Ki Hwan; Kim, Chang Kyu

    1997-11-01

    The thermal compatibility of centrifugally atomized UMo alloys with aluminium has been studied. The results of the investigations show that the U-2 wt% Mo/aluminum dispersions increase in volume by 26% at 400°C after 2000 h. This large volume change is mainly due to the formation of voids and cracks resulting from nearly complete interdiffusion of UMo and aluminum. No significant dimensional changes occur in the U-10 wt% Mo/aluminum dispersions. Interdiffusion between U-10 wt% Mo and aluminum is found to be minimal. The different diffusion behavior is primarily due to the fact that U-10 wt% Mo particles are much more supersaturated with substitutional molybdenum than U-2 wt% Mo particles. The aluminum diffuses into the U-2 wt% Mo particles relatively rapidly along grain boundary with nearly pure uranium, forming UAl 3 almost fully throughout the 2000 h anneal, whereas the molybdenum supersaturated in the U-10 wt% Mo particles inhibits the diffusion of aluminum atoms. U-10 wt% Mo displays superior thermal compatibility with aluminum compared to U-2 wt% Mo.

  9. The Excitement of Collaboration.

    ERIC Educational Resources Information Center

    McCoy, Claire W.

    2000-01-01

    Describes the project called "U2U," a collaboration among the University of Minnesota School of Music, The University of Minnesota dance department, and two urban public school districts in Minneapolis-St. Paul (Minnesota). Presents strategies for collaboratively developing an interdisciplinary curriculum around William Walton's "Facade." Includes…

  10. NASA Aircraft on ramp (Aerial view) Sides: (L) QSRA (R) C-8A AWJSRA - Back to Front: CV-990 (711)

    NASA Technical Reports Server (NTRS)

    1981-01-01

    NASA Aircraft on ramp (Aerial view) Sides: (L) QSRA (R) C-8A AWJSRA - Back to Front: CV-990 (711) C-141 KAO, CV-990 (712) Galileo, T-38, YO-3A, Lear Jet, X-14, U-2, OH-6, CH-47, SH-3G, RSRA, AH-1G, XV-15, UH-1H

  11. A natural orbital analysis of the long range behavior of chemical bonding and van der Waals interaction in singlet H2: the issue of zero natural orbital occupation numbers.

    PubMed

    Sheng, X W; Mentel, Ł M; Gritsenko, O V; Baerends, E J

    2013-04-28

    This paper gives a natural orbital (NO) based analysis of the van der Waals interaction in (singlet) H2 at long distance. The van der Waals interaction, even if not leading to a distinct van der Waals well, affects the shape of the interaction potential in the van der Waals distance range of 5-9 bohrs and can be clearly distinguished from chemical bonding effects. In the NO basis the van der Waals interaction can be quantitatively covered with, apart from the ground state configurations (1σ(g))(2) and (1σ(u))(2), just the 4 configurations (2σ(g))(2) and (2σ(u))(2), and (1π(u))(2) and (1π(g))(2). The physics of the dispersion interaction requires and explains the peculiar relatively large positive CI coefficients of the doubly excited electron configurations (2σ(u))(2) and (1π(g))(2) (the occupancy amplitudes of the 2σ(u) and 1π(gx, y) NOs) in the distance range 5-9 bohrs, which have been observed before by Cioslowski and Pernal [Chem. Phys. Lett. 430, 188 (2006)]. We show that such positive occupancy amplitudes do not necessarily lead to the existence of zero occupation numbers at some H-H distances.

  12. Dynamics of the 1st order phase transition between the nuclear ordered phases of solid 3He

    NASA Astrophysics Data System (ADS)

    Tanaka, Takayoshi; Ito, Hideaki; Sasaki, Yutaka; Mizusaki, Takao

    2005-08-01

    Dynamics of the 1st order phase transition between the U2D2 and the high field phases (HFP) was studied by field-cycling method between these phases by using ultra low temperature magnetic resonance imaging (ULT-MRI). Single Crystal of U2D2 3He was produced at the bottom of compressional cell in superfluid 3He-B at about 0.5 mK. Domain distribution in the U2D2 crystal was examined by ULT-MRI. We have measured the NMR signal intensity to extract the time-evolution of the HFP, after the static magnetic field was swept quickly through the critical field BC1 and was stayed at B=BC1+ΔB. The volume concentration of the U2D2 decreased exponentially in time during the early stage of the phase transition. The rate constant depended positively on ΔB. After the phase transition to the HFP was completed, the static field decreased through BC1 and was fixed at B=BC1-ΔB. The observed rate constant was similar to the value in the opposite direction with identical ΔB. This exponential evolution and ΔB dependence of its rate suggest that the early stage of the phase transition is controlled by the nucleation process.

  13. Diboson excesses in an anomaly free leptophobic left-right model

    NASA Astrophysics Data System (ADS)

    Das, Kasinath; Li, Tianjun; Nandi, S.; Rai, Santosh Kumar

    2016-01-01

    The resonant excesses around 2 TeV reported by the ATLAS Collaboration can be explained in the left-right model, and the tight constraints from lepton plus missing energy searches can be evaded if the S U (2 )R gauge symmetry is leptophobic. We, for the first time, propose an anomaly- free leptophobic left-right model with gauge symmetry S U (3 )C×S U (2 )L×S U (2 )R×U (1 )X , where the SM leptons are singlets under S U (2 )R . The gauge anomalies are cancelled by introducing extra vectorlike quarks. The mass of the Z' gauge boson, which cannot be leptophobic, is assumed to be around or above 2.5 TeV so that the constraint on the dilepton final state can be avoided. Moreover, we find that the W'→W Z channel cannot explain the ATLAS diboson excess due to the tension with the constraint on the W'→j j decay mode. We solve this problem by considering the mixings between the SM quarks and vectorlike quarks. We show explicitly that the ATLAS diboson excess can be explained in the viable parameter space of our model, which is consistent with all the current experimental constraints.

  14. Bosonization, cocycles, and the D1-D5 CFT on the covering surface

    NASA Astrophysics Data System (ADS)

    Burrington, Benjamin A.; Peet, Amanda W.; Zadeh, Ida G.

    2016-01-01

    We consider the D1-D5 CFT near the orbifold point, specifically the computation of correlators involving twist sector fields using covering surface techniques. As is well known, certain twists introduce spin fields on the cover. Here we consider the bosonization of fermions to facilitate computations involving the spin fields. We find a set of cocycle operators that satisfy constraints coming from various S U (2 ) symmetries, including the S U (2 )L×S U (2 )R R-symmetry. Using these cocycles, we consider the correlator of four spin fields on the cover, and show that it is invariant under all of the S U (2 ) symmetries of the theory. We consider the mutual locality of operators, and compute several three-point functions. These computations lead us to a notion of radial ordering on the cover that is inherited from the original computation before lifting. Further, we note that summing over orbifold images sets certain branch-cut ambiguous correlators to zero.

  15. High Resolution Seismic Study of the Holocene Infill of the Elkhorn Slough, Central California

    EPA Science Inventory

    The seismic analysis of the sedimentary infill of the Elkhorn Slough, central California, reveals a succession of three main seismic units: U1, U2, U3, with their correspondent discontinuities d2, d3. These units are deposited over a paleorelief representing the channel location ...

  16. Applied Remote Sensing Program (ARSP). [photomapping arid land in Arizona for land and resources management

    NASA Technical Reports Server (NTRS)

    1977-01-01

    Imagery from U-2 flight or Skylab is used to produce maps of Arizona for resource management and land use. Color photography and thermal mapping techniques are described for studying vegetation growth, natural resources, flood plains, soil erosion, and heat loss from buildings.

  17. Osteoblast integration of dental implant materials after challenge by sub-gingival pathogens: a co-culture study in vitro.

    PubMed

    Zhao, Bingran; van der Mei, Henny C; Rustema-Abbing, Minie; Busscher, Henk J; Ren, Yijin

    2015-12-18

    Sub-gingival anaerobic pathogens can colonize an implant surface to compromise osseointegration of dental implants once the soft tissue seal around the neck of an implant is broken. In vitro evaluations of implant materials are usually done in monoculture studies involving either tissue integration or bacterial colonization. Co-culture models, in which tissue cells and bacteria battle simultaneously for estate on an implant surface, have been demonstrated to provide a better in vitro mimic of the clinical situation. Here we aim to compare the surface coverage by U2OS osteoblasts cells prior to and after challenge by two anaerobic sub-gingival pathogens in a co-culture model on differently modified titanium (Ti), titanium-zirconium (TiZr) alloys and zirconia surfaces. Monoculture studies with either U2OS osteoblasts or bacteria were also carried out and indicated significant differences in biofilm formation between the implant materials, but interactions with U2OS osteoblasts were favourable on all materials. Adhering U2OS osteoblasts cells, however, were significantly more displaced from differently modified Ti surfaces by challenging sub-gingival pathogens than from TiZr alloys and zirconia variants. Combined with previous work employing a co-culture model consisting of human gingival fibroblasts and supra-gingival oral bacteria, results point to a different material selection to stimulate the formation of a soft tissue seal as compared to preservation of osseointegration under the unsterile conditions of the oral cavity.

  18. Photograph and Speech Related to the Cuban Missile Crisis

    ERIC Educational Resources Information Center

    Bredhoff, Stacey; Potter, Lee Ann

    2012-01-01

    On Sunday, October 14, 1962, an American U-2 aircraft, flying a photographic reconnaissance mission over Cuba, took 928 images (one is included with this article). The next day, analysts at the National Photographic Interpretation Center concluded that the photographs showed evidence of Soviet missile site construction in Cuba and conveyed their…

  19. CryoEM structures of two spliceosomal complexes: starter and dessert at the spliceosome feast

    PubMed Central

    Nguyen, Thi Hoang Duong; Galej, Wojciech P; Fica, Sebastian M; Lin, Pei-Chun; Newman, Andrew J; Nagai, Kiyoshi

    2016-01-01

    The spliceosome is formed on pre-mRNA substrates from five small nuclear ribonucleoprotein particles (U1, U2, U4/U6 and U5 snRNPs), and numerous non-snRNP factors. Saccharomyces cerevisiae U4/U6.U5 tri-snRNP comprises U5 snRNA, U4/U6 snRNA duplex and approximately 30 proteins and represents a substantial part of the spliceosome before activation. Schizosaccharomyces pombe U2.U6.U5 spliceosomal complex is a post-catalytic intron lariat spliceosome containing U2 and U5 snRNPs, NTC (nineteen complex), NTC-related proteins (NTR), U6 snRNA, and an RNA intron lariat. Two recent papers describe near-complete atomic structures of these complexes based on cryoEM single-particle analysis. The U4/U6.U5 tri-snRNP structure provides crucial insight into the activation mechanism of the spliceosome. The U2.U6.U5 complex reveals the striking architecture of NTC and NTR and important features of the group II intron-like catalytic RNA core remaining after spliced mRNA is released. These two structures greatly advance our understanding of the mechanism of pre-mRNA splicing. PMID:26803803

  20. Nobiletin inhibits human osteosarcoma cells metastasis by blocking ERK and JNK-mediated MMPs expression

    PubMed Central

    Cheng, Hsin-Lin; Hsieh, Ming-Ju; Yang, Jia-Sin; Lin, Chiao-Wen; Lue, Ko-Haung; Lu, Ko-Hsiu; Yang, Shun-Fa

    2016-01-01

    Nobiletin, a polymethoxyflavone, has a few pharmacological activities, including anti-inflammation and anti-cancer effects. However, its effect on human osteosarcoma progression remains uninvestigated. Therefore, we examined the effectiveness of nobiletin against cellular metastasis of human osteosarcoma and the underlying mechanisms. Nobiletin, up to 100 μM without cytotoxicity, significantly decreased motility, migration and invasion as well as enzymatic activities, protein levels and mRNA expressions of matrix metalloproteinase (MMP)-2 and MMP-9 in U2OS and HOS cells. In addition to inhibition of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), the inhibitory effect of nobiletin on the DNA-binding activity of the transcription factor nuclear factor-kappa B (NF-κB), cAMP response element-binding protein (CREB), and specificity protein 1 (SP-1) in U2OS and HOS cells. Co-treatment with ERK and JNK inhibitors and nobiletin further reduced U2OS cells migration and invasion. These results indicated that nobiletin inhibits human osteosarcoma U2OS and HOS cells motility, migration and invasion by down-regulating MMP-2 and MMP-9 expressions via ERK and JNK pathways and through the inactivation of downstream NF-κB, CREB, and SP-1. Nobiletin has the potential to serve as an anti-metastatic agent for treating osteosarcoma. PMID:27144433

  1. 40 CFR 98.213 - Calculating GHG emissions.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... the process emissions of CO2 using actual mass of output carbonates with Equation U-2 of this section... = Annual mass of input carbonate type k (tons). EFk = Emission factor for the carbonate type k, as specified in Table U-1 of this subpart (metric tons CO2/metric ton carbonate input). Mj = Annual mass...

  2. Modules and supporting hardware for FASTBUS test and diagnostic purposes

    SciTech Connect

    Bertolucci, B.

    1981-10-01

    This paper contains detailed descriptions and circuitry of some modules and supporting hardware for the FASTBUS System developed at SLAC. A fast slave-only Memory Module (PRIMO), a Dummy Module (U2), a FASTBUS Test Box (LAIKA), and a Bus Display Bar (BBD) have been built, tested and used for test and diagnostic purposes for FASTBUS.

  3. Comparative evaluation of modified trichrome and Uvitex 2B stains for detection of low numbers of microsporidial spores in stool specimens.

    PubMed

    Ignatius, R; Henschel, S; Liesenfeld, O; Mansmann, U; Schmidt, W; Köppe, S; Schneider, T; Heise, W; Futh, U; Riecken, E O; Hahn, H; Ullrich, R

    1997-09-01

    At present, the laboratory diagnosis of intestinal infections caused by microsporidia depends on the detection of the typical spores either with a modified trichrome stain (MTS) or by staining with fluorochromes. The purpose of the present study was (i) to compare staining with MTS (MTS method) and the staining with the fluorochrome Uvitex 2B (U2B method) with respect to their sensitivities and specificities, particularly in the presence of low numbers of spores, and (ii) to evaluate their reliabilities under routine laboratory conditions. First, 30 negative human stool specimens as well as 30 specimens enriched with a low concentration of microsporidial spores were examined. The U2B and MTS methods detected 27 and 30, of the positive samples, respectively (95% confidence intervals for sensitivity, 0.73 to 0.98 for the U2B method and 0.88 to 1.00 for the MTS method) without yielding false-positive results (95% confidence intervals for specificity, 0.88 to 1.00 for the MTS and U2B methods). In addition, analysis of serial dilutions of 17 stool specimens from AIDS patients containing microsporidia revealed comparable detection thresholds (P = 0.52) for both methods. Finally, 40 slides prepared from one stool specimen containing very few microsporidia and 40 negative slides were included in the routine diagnostic program during 1 month in order to monitor laboratory handling and run-to-run variations. Again, both methods exhibited comparable sensitivities (95% confidence intervals, 0.83 to 0.99 for the MTS method and 0.91 to 1.00 for the U2B method) and specificities (95% confidence intervals, 0.91 to 1.00 for the MTS and U2B methods). In conclusion, MTS and U2B methods are equally useful in the diagnosis of microsporidiosis. However, since detection thresholds for both methods differed considerably in all diluted stool specimens, performance of a combination of both methods may be more sensitive than the performance of only one procedure in the event of very low

  4. Anemone altaica Induces Apoptosis in Human Osteosarcoma Cells.

    PubMed

    Chang, I-Chang; Chiang, Tsay-I; Lo, Chun; Lai, Yi-Hua; Yue, Chia-Herng; Liu, Jer-Yuh; Hsu, Li-Sung; Lee, Chia-Jen

    2015-01-01

    In the past decade, no significant improvement has been made in chemotherapy for osteosarcoma (OS). To develop improved agents against OS, we screened 70 species of medicinal plants and treated two human OS cell lines with different agent concentrations. We then examined cell viability using the MTT assay. Results showed that a candidate plant, particularly the rhizomes of Anemone altaica Fisch. ex C. A. Mey aqueous extract (AAE), suppressed the viability of HOS and U2OS cells in a concentration-dependent manner. Flow cytometry analysis revealed that AAE significantly increased the amount of cell shrinkage (Sub-G1 fragments) in HOS and U2OS cells. Moreover, AAE increased cytosolic cytochrome c and Bax, but decreased Bcl-2. The amount of cleaved caspase-3 and poly-(ADP-ribose) polymerase-1 (PARP-1) were significantly increased. AAE suppressed the growth of HOS and U2OS through the intrinsic apoptotic pathway. Data suggest that AAE is cytotoxic to HOS and U2OS cells and has no significant influence on human osteoblast hFOB cells. The high mRNA levels of apoptosis-related factors (PPP1R15A, SQSTM1, HSPA1B, and DDIT4) and cellular proliferation markers (SKA2 and BUB1B) were significantly altered by the AAE treatment of HOS and U2OS cells. Results show that the anticancer activity of AAE could up-regulate the expression of a cluster of genes, especially those in the apoptosis-related factor family and caspase family. Thus, AAE has great potential as a useful therapeutic drug for human OS. PMID:26224029

  5. Higgs boson spectra in supersymmetric left-right models

    NASA Astrophysics Data System (ADS)

    Babu, K. S.; Patra, Ayon

    2016-03-01

    We present a comprehensive analysis of the Higgs boson spectra in several versions of the supersymmetric left-right model based on the gauge symmetry S U (3 )c×S U (2 )L×S U (2 )R×U (1 )B-L. A variety of symmetry breaking sectors are studied, with a focus on the constraints placed on model parameters by the lightest neutral C P even Higgs boson mass Mh. The breaking of S U (2 )R symmetry is achieved by Higgs fields transforming either as triplets or doublets, and the electroweak symmetry breaking is triggered by either bi-doublets or doublets. The Higgs potential is analyzed with or without a gauge singlet Higgs field present. Seesaw models of Type I and Type II, inverse seesaw models, universal seesaw models and an E6 inspired alternate left-right model are included in our analysis. Several of these models lead to the tree-level relation Mh≤√{2 }mW (rather than Mh≤mZ that arises in the MSSM), realized when the S U (2 )R symmetry breaking scale is of order TeV. With such an enhanced upper limit, it becomes possible to accommodate a Higgs boson of mass 126 GeV with relatively light stops that mix negligibly. In models with Higgs triplets, a doubly charged scalar remains light below a TeV with its mass arising entirely from radiative corrections. We carry out the complete one-loop calculation for its mass induced by the Majorana Yukawa couplings and show the consistency of the framework. We argue that these models prefer a low S U (2 )R breaking scale. Other theoretical and phenomenological implications of these models are briefly discussed.

  6. Working Together to Deliver Usable Climate Information to Agricultural Producers and Advisors in the U.S. Corn Belt

    NASA Astrophysics Data System (ADS)

    Prokopy, L. S.; Widhalm, M.

    2012-12-01

    Weather and climate patterns are a driving force behind the success or failure of cropping systems. With U.S. corn and soybean production accounting for nearly one-third of global supplies and contributing $100 billion annually to the national economy, the ability to successfully produce crops under more variable climate conditions is critical. Useful to Usable (U2U): Transforming Climate Variability and Change Information for Cereal Crop Producers is a USDA-funded research and extension project focused on delivering to producers and farm advisors the resources and training they need to more effectively manage variable climate conditions. The U2U team is a diverse and uniquely qualified group of climatologists, crop modelers, agronomists, economists, and social scientists from 10 Midwestern universities and two NOAA Regional Climate Centers. Together, we strive to help producers make better long-term plans on what, when and where to plant and also how to manage crops for maximum yields and minimum environmental damage. Under development are tools that will allow the agricultural community to examine the financial, production, and environmental outcomes of different management options and climate scenarios so farmers can choose strategies that fit their capabilities and acceptable levels of risk. Researchers are currently using existing data and agro-climate models to investigate the impact of climate conditions on key topics such as crop yields, fieldwork opportunities, nitrogen management, and the cost-effectiveness of irrigation and tiling. To ensure relevance and usability of U2U products, our social science team is using a number of techniques including surveys and focus groups to integrate stakeholder interests, needs, and concerns into all aspects of U2U research. It is through this coupling of physical and social science disciplines that we strive to transform existing climate information into actionable knowledge. This presentation will elaborate on U2U

  7. A note on Hardy-Weinberg equilibrium of VNTR data by using the Federal Bureau of Investigation's fixed-bin method

    SciTech Connect

    Devlin, B.; Risch, N. )

    1992-09-01

    To fully utilize the information of VNTR data for forensic inference, the probability of observing the matching suspect and evidentiary profile in a reference population is estimated, usually by assuming independence of alleles within and between loci. This assumption has been challenged on the basis of the observation that there is frequently an excess of single-band phenotypes (SBP) in forensic data bases, which could indicate lack of independence. Nevertheless, another explanation is that the excess SBP are artifacts of laboratory methods. In this report the authors examine the excess of SBP for three VNTR loci studied by the FBI (D17S79 and D2S44, for blacks, and D14S13, for Caucasians). The FBI claims that the excess is due to the effect of null alleles; the null alleles are suspected to be small to be detected. The authors estimate the frequency of null alleles for two loci (D17S79 and D14A13) by comparing, for these loci, the data from the FBI data base and the data from the Lifecodes data base. These comparisons yield information on small fragments because Lifecodes uses the restriction enzyme PstI, which yields larger fragments than does HaeIII, which the FBI uses. For D17S19 in blacks, the authors estimate a null allele frequency of 4.4%, and, for D14S13 in Caucasians, they estimate a frequency of 3.0%. The null-allele frequency for D2S44 in blacks is derived similarly, again being based on analysis of DNA cut with HaeIII and PstI; the estimate of the null-allele frequency for this locus is 1.5%. Using these null-allele frequency estimates and a goodness-of-fit test, the authors show that there is no evidence for deviations from Hardy-Weinberg expectations of genotype probabilities at these loci. 20 refs., 1 fig.

  8. Structural Insights into the Catalytic Mechanism of Escherichia coli Selenophosphate Synthetase

    SciTech Connect

    Noinaj, Nicholas; Wattanasak, Rut; Lee, Duck-Yeon; Wally, Jeremy L.; Piszczek, Grzegorz; Chock, P. Boon; Stadtman, Thressa C.; Buchanan, Susan K.

    2012-03-26

    Selenophosphate synthetase (SPS) catalyzes the synthesis of selenophosphate, the selenium donor for the biosynthesis of selenocysteine and 2-selenouridine residues in seleno-tRNA. Selenocysteine, known as the 21st amino acid, is then incorporated into proteins during translation to form selenoproteins which serve a variety of cellular processes. SPS activity is dependent on both Mg{sup 2+} and K{sup +} and uses ATP, selenide, and water to catalyze the formation of AMP, orthophosphate, and selenophosphate. In this reaction, the gamma phosphate of ATP is transferred to the selenide to form selenophosphate, while ADP is hydrolyzed to form orthophosphate and AMP. Most of what is known about the function of SPS has derived from studies investigating Escherichia coli SPS (EcSPS) as a model system. Here we report the crystal structure of the C17S mutant of SPS from E. coli (EcSPS{sup C17S}) in apo form (without ATP bound). EcSPS{sup C17S} crystallizes as a homodimer, which was further characterized by analytical ultracentrifugation experiments. The glycine-rich N-terminal region (residues 1 through 47) was found in the open conformation and was mostly ordered in both structures, with a magnesium cofactor bound at the active site of each monomer involving conserved aspartate residues. Mutating these conserved residues (D51, D68, D91, and D227) along with N87, also found at the active site, to alanine completely abolished AMP production in our activity assays, highlighting their essential role for catalysis in EcSPS. Based on the structural and biochemical analysis of EcSPS reported here and using information obtained from similar studies done with SPS orthologs from Aquifex aeolicus and humans, we propose a catalytic mechanism for EcSPS-mediated selenophosphate synthesis.

  9. Stage-specific excretory-secretory small heat shock proteins from the parasitic nematode Strongyloides ratti--putative links to host's intestinal mucosal defense system.

    PubMed

    Younis, Abuelhassan Elshazly; Geisinger, Frank; Ajonina-Ekoti, Irene; Soblik, Hanns; Steen, Hanno; Mitreva, Makedonka; Erttmann, Klaus D; Perbandt, Markus; Liebau, Eva; Brattig, Norbert W

    2011-09-01

    In a search for molecules involved in the interaction between intestinal nematodes and mammalian mucosal host cells, we performed MS to identify excretory-secretory proteins from Strongyloides ratti. In the excretory-secretory proteins of the parasitic female stage, we detected, in addition to other peptides, peptides homologous with the Caenorhabditis elegans heat shock protein (HSP)-17, named Sra-HSP-17.1 (∼ 19 kDa) and Sra-HSP-17.2 (∼ 18 kDa), with 49% amino acid identity. The full-length cDNAs (483 bp and 474 bp, respectively) were identified, and the genomic organization was analyzed. To allow further characterization, the proteins were recombinantly expressed and purified. Profiling of transcription by quantitative real-time-PCR and of protein by ELISA in various developmental stages revealed parasitic female-specific expression. Sequence analyses of both the DNA and amino acid sequences showed that the two proteins share a conserved α-crystallin domain and variable N-terminals. The Sra-HSP-17s showed the highest homology with the deduced small HSP sequence of the human pathogen Strongyloides stercoralis. We observed strong immunogenicity of both proteins, leading to strong IgG responses following infection of rats. Flow cytometric analysis indicated the binding of Sra-HSP-17s to the monocyte-macrophage lineage but not to peripheral lymphocytes or neutrophils. A rat intestinal epithelial cell line showed dose-dependent binding to Sra-HSP-17.1, but not to Sra-HSP-17.2. Exposed monocytes released interleukin-10 but not tumor necrosis factor-α in response to Sra-HSP-17s, suggesting the possible involvement of secreted female proteins in host immune responses.

  10. Effect of breastfeeding piperine on the learning of offspring mice: interaction with caffeine and diazepam

    PubMed Central

    Moghadamnia, Ali Akbar; Zangoori, Vahid; Zargar-Nattaj, Seyed Sadegh; Tayebi, Pooya; Moghadamnia, Yasaman; Jorsaraei, Seyed Gholam Ali

    2010-01-01

    Piperine, the main alkaloid of black pepper (Piper nigrum), has been suggested to display several pharmacological properties, including pain relief, anticonvulsant, antidepressant-like, antianxiety, sedative, and anti-inflammatory effects. This study was designed to investigate the effect of piperine on learning in mice and the interaction of the effect with caffeine and diazepam. Piperine (100 mg/kg intraperitoneally) was injected into the mouse mothers or nursing dams during breastfeeding for 25 days at five-day intervals. After feeding the newborn mice, their learning was evaluated using a step-through passive avoidance task. Mouse learning was assessed 1 hr and 24 hr and 1 week after a training session. Piperine increased learning in the first (1 hr: 243.33 s vs 55.17 s, P = 0.002) and third assessments (1 week: 226 s vs 97 s, P < 0.05) post-training, but no significant change was seen at the second (24 hr) assessment. Piperine improved the effect of a low dose of caffeine (25 mg/kg intraperitoneally after a shock of 2 s duration) in a first assessment (295.17 s vs 149.17 s, P = 0.026) compared to a higher dose of caffeine. Piperine reversed diazepam (1 mg/kg intraperitoneally) suppression of learning 24 hours after training by a 4 s shock (298 s vs 135.67 s, P = 0.03). According to the results, piperine alone significantly increased learning 1 hour and 1 week after training assessments, and learning can be improved in the short term when followed by piperine administration. It was also shown that piperine can potentiate the effect of a low dose of caffeine and can reverse the effect of diazepam. PMID:27186097

  11. BRCA1-linked marker in postmenopausal breast cancer families

    SciTech Connect

    Folsom, A.R.; Chen, P.L.; Sellers, T.A.

    1994-09-01

    A majority of breast and ovarian cancer families and half of the early-onset breast cancer families are linked to markers on 17q (BRCA1). While linkage has been demonstrated in families with premenopausal disease, few studies have tested these markers in families with postmenopausal breast cancer. In the Iowa Women`s Health Study, a population-based study of over 42,000 women, an association of waist-to-hip ratio (WHR) with the risk of postmenopausal breast cancer was found predominantly in women with a positive family history -- this interaction was associated with a 3.2-fold elevated risk. This effect was even more pronounced when the definition of family history included breast and ovarian cancer, known to be linked to 17q markers. We evaluated evidence for linkage with D17S579, a BRCA-1-linked marker, in 13 families in which the index case had postmenopausal breast cancer. Genotyping for alleles at D17S579 was performed on 84 blood samples. Linkage analysis assumed that the breast cancer trait had an autosomal dominant mode of inheritance with a penetrance of 80%. For the 13 families studied, the maximum lod score was 0.29 at a theta of 0.27. There was significant evidence against tight linkage of breast cancer with D17S579 (theta<0.4). Heterogeneity analysis suggested evidence for the presence of both linked and unlinked families. Partitioning informative families on WHR of the index case suggested heterogeneity. These data suggest that, in a subset of families identified by a postmenopausal breast cancer proband, risk of breast cancer may be mediated by BRCA1, with heterogeneity defined by WHR.

  12. Incorporation of polyaniline nanofibres on graphene oxide by interfacial polymerization pathway for supercapacitor

    NASA Astrophysics Data System (ADS)

    Male, Umashankar; Srinivasan, Palaniappan; Singu, Bal Sydulu

    2015-09-01

    The aim of this work is to improve the supercapacitor performance of polyaniline (PANI). Polyaniline nano fibres are incorporated into graphene oxide (GO) layers by interfacial polymerization pathway, wherein PANI fibres are intercalated into GO layers and also cover the GO. PANI-GO hybrid composite is obtained in semi-crystalline form with good conductivity (1.7 S cm-1). The specific capacitance for PANI-GO (365 F g-1) is found to be higher than PANI (280 F g-1). At the energy density of 15 W h kg-1, the power density of PANI-GO (632 W kg-1) is higher than PANI (283 W kg-1).

  13. An Investigation of the Effectiveness of Stiffeners on Shear-resistant Plate-girder Webs

    NASA Technical Reports Server (NTRS)

    Moore, R L

    1942-01-01

    The results of 60 different tests on 2 aluminum alloy 17S-T plate girders are presented to show the influence of size and spacing of stiffeners upon the buckling characteristics of shear-resistant webs within the elastic range. It is demonstrated that stiffeners increase the stability of a web by retarding the formation subdivided panels. An empirical method of proportioning stiffeners is proposed which recognizes both of these stiffener functions, and comparisons are made with design procedures based upon theoretical considerations of the buckling problem. Also, some experimental data are provided to show the effect of stiffener size and spacing upon ultimate web strengths.

  14. Double-peak structure in the cyclic optical outbursts of blazar OJ 287.

    NASA Astrophysics Data System (ADS)

    Sillanpaa, A.; Takalo, L. O.; Pursimo, T.; Nilsson, K.; Heinamaki, P.; Katajainen, S.; Pietila, H.; Hanski, M.; Rekola, R.; Kidger, M.; Boltwood, P.; Turner, G. W.; Robertson, J. W.; Honeycut, R. K.; Efimov, Yu. S.; Shakhovskoy, N.; Fiorucci, M.; Tosti, G.; Ghisellini, G.; Raiteri, C. M.; Villata, M.; de Francesco, G.; Lanteri, L.; Chiaberge, M.; Peila, A.; Heidt, J.

    1996-11-01

    An international monitoring project, called OJ-94, was set up with the aim of monitoring blazar OJ 287 during the predicted optical outburst (in fall 1994, Sillanpaa et al. 1988ApJ...325..628S) and the predicted secondary outburst (in winter 1995-1996, Sillanpaa et al. 1996A&A...305L..17S). The most prominent features in the OJ 287 light curve during the period covered by the project are the large outburst in September-December 1994, reaching the maximum at the beginning of November, and the secondary outburst peaking just at Christmas 1995. Both these outbursts occurred almost exactly at the predicted times.

  15. Rheological behavior of Slide Ring Gels.

    NASA Astrophysics Data System (ADS)

    Sharma, Vivek; Park, Jong Seung; Park, Jung O.; Srinivasarao, Mohan

    2006-03-01

    Slide ring gels were synthesized by chemically crosslinking, sparsely populated α-cyclodextrin (α-CD) present on the polyrotaxanes consisting of α-CD and polyethylene glycol (PEG). [1] Unlike physically or chemically crosslinked gels, slide ring gels are topological gels where crosslinks can slide along the chain. [2] We investigate the rheological behavior of these gels swollen in water and compare their viscoelastic properties to those of physical and chemical gels. We also study the equilibrium swelling behavior of these gels. [1] Okumura and Ito, Adv. Mater. 2001, 13, 485 [2] C. Zhao et al, J. Phys. Cond. Mat. 2005, 17, S2841

  16. Localization of the gene (SYM1) for proximal symphalangism to human chromosome 17q21-q22

    SciTech Connect

    Polymeropoulos, M.H.; Rubenstein, J.R.; Poush, J.

    1995-05-20

    Proximal symphalangism, or Cushing symphalangism (MIM 185800), is an autosomal dominant disorder characterized by ankylosis of the proximal interphalangeal joints. Conductive deafness and reduced flexibility of the ankles have also been observed in affected individuals. The authors have used polymorphic markers throughout the genome to perform genetic linkage analysis in subsequent generations of the family originally described by Harvey Cushing. They have established linkage for this disorder to markers on chromosome 17 (17q21-q22), with Z{sub max} = 6.98 at {theta} = 0.05 with marker D17S790. 15 refs., 2 figs., 2 tabs.

  17. 49 CFR 595.7 - Requirements for vehicle modifications to accommodate people with disabilities.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ..., S7.4, S14, S15, S16, S17, S18, S19, S20, S21, S22, S23, S24, S25, S26 and S27 of 49 CFR 571.208 for... the type and in the manner described in paragraph (e) of this section. (c)(1) 49 CFR 571.101, except for S5.2.1, S5.3.1, S5.3.4, S5.4.1, and S5.4.3 of that section. (2) S5.1.1.5 of 49 CFR 571.108, in...

  18. Genetic linkage analysis in familial breast and ovarian cancer: Results from 214 families

    SciTech Connect

    Easton, D.F.; Ford, D. ); Bishop, D.T.; Crockford, G.P. )

    1993-04-01

    This paper reports the results of a collaborative linkage study involving 214 breast cancer families, including 57 breast-ovarian cancer families; this represents almost all the known families with 17q linkage data. Six markers on 17q, spanning approximately 30 cM, were typed in the families. The aims of the study were to define more precisely the localization of the disease gene, the extent of genetic heterogeneity and the characteristics of linked families and to estimate the penetrance of the 17q gene. Under the assumption of no genetic heterogeneity, the strongest linkage evidence was obtained with D17S588. Multipoint linkage analysis allowing for genetic heterogeneity provided evidence that the predisposing gene lies between the markers D17S588 and D17S250, an interval whose genetic length is estimated to be 8.3 cM in males and 18.0 cM in females. This position was supported over other intervals by odds of 66:1. The location of the gene with respect to D17S579 could not be determined unequivocally. Under the genetic model used in the analysis, the best estimate of the proportion of linked breast-ovarian cancer families was 1.0 (lower LOD -- 1 limit 0.79). In contrast, there was significant evidence of genetic heterogeneity among the families without ovarian cancer, with an estimated 45% being linked. These results suggest that a gene(s) on chromosome 17q accounts for the majority of families in which both early-onset breast cancer and ovarian cancer occur but that other genes predisposing to breast cancer exist. By examining the fit of the linkage data to different penetrance functions, the cumulative risk associated with the 17q gene was estimated to be 59% by age 50 years and 82% by age 70 years. The corresponding estimates for the breast-ovary families were 67% and 76%, and those for the families without ovarian cancer were 49% and 90%; these penetrance functions did not differ significantly from one another. 42 refs., 5 figs., 2 tabs.

  19. ATOMIC AND MOLECULAR PHYSICS: Bose-Einstein condensation on an atom chip

    NASA Astrophysics Data System (ADS)

    Yan, Bo; Cheng, Feng; Ke, Min; Li, Xiao-Lin; Tang, Jiu-Yao; Wang, Yu-Zhu

    2009-10-01

    This paper reports an experiment of creating Bose-Einstein condensate (BEC) on an atom chip. The chip-based Z-wire current with a homogeneous bias magnetic field creates a tight magnetic trap, which allows for a fast production of BEC. After a 4.17-s forced radio frequency evaporative cooling, a condensate with about 3000 atoms appears. The transition temperature is about 300 nK. This compact system is quite robust, allowing for versatile extensions and further studying of BEC.

  20. Reduced IgG anti-small nuclear ribonucleoprotein autoantibody production in systemic lupus erythematosus patients with positive IgM anti-cytomegalovirus antibodies

    PubMed Central

    Palafox Sánchez, Claudia Azucena; Satoh, Minoru; Chan, Edward KL; Carcamo, Wendy C; Muñoz Valle, José Francisco; Orozco Barocio, Gerardo; Oregon Romero, Edith; Navarro Hernández, Rosa Elena; Salazar Páramo, Mario; Cabral Castañeda, Antonio; Vázquez del Mercado, Mónica

    2009-01-01

    Introduction Systemic lupus erythematosus is characterized by production of autoantibodies to RNA or DNA–protein complexes such as small nuclear ribonucleoproteins (snRNPs). A role of Epstein–Barr virus in the pathogenesis has been suggested. Similar to Epstein–Barr virus, cytomegalovirus (CMV) infects the majority of individuals at a young age and establishes latency with a potential for reactivation. Homology of CMV glycoprotein B (UL55) with the U1snRNP-70 kDa protein (U1–70 k) has been described; however, the role of CMV infection in production of anti-snRNPs is controversial. We investigated the association of CMV serology and autoantibodies in systemic lupus erythematosus. Methods Sixty-one Mexican patients with systemic lupus erythematosus were tested for CMV and Epstein–Barr virus serology (viral capsid antigen, IgG, IgM) and autoantibodies by immunoprecipitation and ELISA (IgG and IgM class, U1RNP/Sm, U1–70 k, P peptide, rheumatoid factor, dsDNA, β2-glycoprotein I). Results IgG anti-CMV and IgM anti-CMV were positive in 95% (58/61) and 33% (20/61), respectively, and two cases were negative for both. Clinical manifestation and autoantibodies in the IgM anti-CMV(+) group (n = 20) versus the IgM anti-CMV(-)IgG (+) (n = 39) group were compared. Most (19/20) of the IgM anti-CMV(+) cases were IgG anti-CMV(+), consistent with reactivation or reinfection. IgM anti-CMV was unrelated to rheumatoid factor or IgM class autoantibodies and none was positive for IgM anti-Epstein–Barr virus–viral capsid antigen, indicating that this is not simply due to false positive results caused by rheumatoid factor or nonspecific binding by certain IgM. The IgM anti-CMV(+) group has significantly lower levels of IgG anti-U1RNP/Sm and IgG anti-U1–70 k (P = 0.0004 and P = 0.0046, respectively). This finding was also confirmed by immunoprecipitation. Among the IgM anti-CMV(-) subset, anti-Su was associated with anti-U1RNP and anti-Ro (P < 0.05). High levels of Ig

  1. Differential gene expression in neurospora crassa cell types: amplification of rRNA genes. Progress report, July 1979-30 June 1980

    SciTech Connect

    Dutta, S.K.

    1980-01-01

    The significant results obtained during 1979 to 1980 of the current research program are as follows: (1) the differential rRNA gene amplification in germinated conidia of N.crassa was confirmed. N.crassa rDNAs showed differences in degrees of homology with isolated DNAs from other Neurospora species which could be due to heterogeneity in internal spacers. Studies with N.crassa rDNA clones were initiated to study their heterogeneities. The organization of the Institutional Biohazard Committee (IBC) for Recombinant DNA research was completed and necessary certifications for the laboratory and the workers were obtained in accordance with the P/sub 2/EK/sub 1/ containment regulation of N.I.H. Known 17S and 26S N.crassa rDNA probes are being used to detect differences, if any, in restriction cleavage sites in rDNAs of different cell types and developmental mutants of N.crassa. DNAs from these N.crassa cells are restricted with EcoR/sub 1/ and Hind III and cleaved fragments separated by gel electrophoresis are transferred into nitrocellulose papers. Experiments are underway now to see if there are any changes in cleavage sites by annealing with /sup 32/P or /sup 3/H-17S or 26S rDNA probes followed by autoradiography.

  2. Functional analysis of transcribed spacers of yeast ribosomal DNA.

    PubMed Central

    Musters, W; Boon, K; van der Sande, C A; van Heerikhuizen, H; Planta, R J

    1990-01-01

    Making use of an rDNA unit, containing oligonucleotide tags in both the 17S and 26S rRNA gene, we have analyzed the effect of various deletions in the External Transcribed Spacer (ETS) and in one of the Internal Transcribed Spacers 1 (ITS1) on the process of ribosome formation in yeast. By following the fate of the tagged transcripts of this rDNA unit in vivo by Northern hybridization we found that deleting various parts of the ETS prevents the accumulation of tagged 17S rRNA and its assembly into 40S subunits, but not the formation of 60S subunits. Deleting the central region of ITS1, including a processing site that is used in an early stage of the maturation process, was also found to prevent the accumulation of functional 49 S subunits, whereas no effect on the formation of 60S subunits was detected. The implications of these findings for yeast pre-rRNA processing are discussed. Images Fig. 3. Fig. 4. Fig. 6. Fig. 7. PMID:2249660

  3. Physical and transcript map of the minimally deleted region III on 17p implicated in the early development of Barrett's oesophageal adenocarcinoma.

    PubMed

    Dunn, Julie R; Risk, Janet M; Langan, Joanne E; Marlee, Damian; Ellis, Anthony; Campbell, Fiona; Watson, Alastair J M; Field, John K

    2003-06-26

    Allelic imbalance (AI) studies on chromosome 17 (C17) in Barrett's oesophageal adenocarcinoma (BOA) tumours strongly suggest that a minimally deleted region on C17p harbours a BOA-associated gene with tumour suppressor function. This deleted region, designated minimal region III (MRIII), lies between the two microsatellite markers D17S1852 and D17S954. Computational sequence analysis techniques, BLAST and NIX, were used to assemble a physical map of MRIII, consisting of three overlapping bacterial artificial chromosome (BAC) clones, 297N7, 963H4 and 795F17, from the RPCI-11 library. The 270 kb genomic sequence of MRIII was analysed using the computational gene prediction methods NIX and TAP to identify putative BOA genes. A transcript map of MRIII has been generated and contains 25 candidate BOA genes, four of which are the named genes MYH3, SCO1, x006 and MAGOH-LIKE. The other candidates consist of seven genes predicted by TAP with associated ESTs identified by NIX, two genes predicted by TAP alone and 12 genes/ESTs (or pairs of ESTs) identified by NIX alone. No disease-specific mutations were identified in x006 or MAGOH-LIKE, although expression analysis of these genes suggests that they may show alternative splicing or be altered epigenetically or in regulatory regions in oesophageal cancer.

  4. Genetic polymorphism of 13 non-CODIS STR loci in three national populations from China.

    PubMed

    Liu, Qiu-Ling; Huang, Kai-Kai; Wu, Ye-Da; Zhao, Hu; Li, Cheng-Tao; Lu, De-Jian

    2014-12-01

    The aim of this study was to investigate a 13 non-CODIS STR loci database using three national populations from China. A new multiplex PCR system that simultaneously amplified 13 loci in the same PCR reaction was developed. This multiplex system included the 13 STR markers (D3S2402, D3S2452, D3S1766, D3S4554, D3S2388, D3S3051, D3S3053, D4S2364, D4S2404, AC001348A, AC001348B, D17S975, and D17S1294), which were successfully analyzed by using 441 DNA samples from three national populations in China (154 Mongol, 177 Kazakh, and 110 Uigur). Allele frequencies and mutation rates of the 13 non-CODIS STR loci were investigated. A total of 4-10 alleles at each locus were observed and altogether 84, 88, and 87 alleles for the all selected loci were found in the Mongol, Kazakh, and Uigur, respectively. Eight mutations were detected from the 13 selected loci in 9880 meioses in kinship cases. These results indicate that this multiplex system may provide significant polymorphic information for kinship testing and relationship investigations.

  5. A novel mutation within the 2B rod domain of keratin 9 in a Chinese pedigree with epidermolytic palmoplantar keratoderma combined with knuckle pads and camptodactyly.

    PubMed

    Du, Zhen-Fang; Wei, Wei; Wang, Yi-Fan; Chen, Xiao-Ling; Chen, Chun-Yue; Liu, Wen-Ting; Lu, Jia-Jun; Mao, Lian-Gen; Xu, Chen-Ming; Fang, Hong; Zhang, Xian-Ning

    2011-01-01

    Knuckle pads and camptodactyly are overlapping symptoms associated with many genetic and environmental factors. To the best of our knowledge, all reported cases of epidermolytic palmoplantar keratoderma (EPPK) with knuckle pads have been without accompanying camptodactyly. We here report a novel KRT9 mutation-EPPK family with combined knuckle pads and camptodactyly. All the EPPK-affected individuals in this southern Chinese pedigree suffered severe diffuse palmar and plantar hyperkeratosis including hyperhidrosis and cuticle splitting: 3 females presented EPPK only, 8 adult males had notably severe knuckle pads and camptodactyly as well as EPPK, and one 6-year-old boy manifested EPPK with knuckle pads. Haplotype analysis excluded the known candidate loci for camptodactyly and/or knuckle pad-like phenotypes on chromosomes 13q12, 3q11.2-q13.12, 1q24-q25, 4p16.3 and 16q11.1-q22, while only the markers D17S1787 and D17S579 flanking KRT9 showed co-segregation with EPPK. Then a novel c.T1373C (p.L458P) mutation within the sixth exon of KRT9 was validated, and this mutation presented a more severe pathogenicity than the previously reported p.L458F. We speculated that KRT9 plays a complicated role in the genesis of EPPK with knuckle pads and camptodactyly, which needs to be further investigated.

  6. DNA homologies of ribosomal RNA genes of Neurospora species

    SciTech Connect

    Mukhopadhyay, D.K.; Mimiko, R.; Dutta, S.K.

    1980-01-01

    Ribosomal RNA genes (rDNAs) of Neurospora crassa contain DNA sequences which code for 17S, 5.8S, and 26S rRNAs, in addition to internal and external spacers. As has been reported for many eukaryotes, the DNA sequences which code for 17S, 5.8S, and 26S rRNAs in Neurospora species are probably conserved while the internal and external spacer regions are probably variable sequences. Extensive electron microscopic studies of 45S precursor rRNA of several cold and warm blooded animals confirm that spacer regions vary extensively from species to species. It was desirable to know whether such differences in rDNA sequences exist between Neurospora species. Any such difference should be detectable using standard procedures for DNA homology studies rDNA sequences were isolated from N. crassa mycelial cells using the procedure described previously. The purified rDNA was /sup 3/H-labeled (by nick translation) and reassociated with total DNA isolated from the heterothallic species N. crassa and from three homothalliospecies: N. dodgei, N. lineolata, and N. africana. In addition, /sup 32/P-labeled total DNA of N. crassa was reannealed with unlabeled bulk DNA from N. crassa, N. dodgei, and N. lineolata.

  7. Rhodopsin and Melanopsin Contributions to the Early Redilation Phase of the Post-Illumination Pupil Response (PIPR)

    PubMed Central

    Adhikari, Prakash; Feigl, Beatrix; Zele, Andrew J.

    2016-01-01

    Melanopsin expressing intrinsically photosensitive Retinal Ganglion Cells (ipRGCs) entirely control the post-illumination pupil response (PIPR) from 6 s post-stimulus to the plateau during redilation after light offset. However, the photoreceptor contributions to the early redilation phase of the PIPR (< 6 s post-stimulus) have not been reported. Here, we evaluated the photoreceptor contributions to the early phase PIPR (0.6 s to 5.0 s) by measuring the spectral sensitivity of the criterion PIPR amplitude in response to 1 s light pulses at five narrowband stimulus wavelengths (409, 464, 508, 531 and 592 nm). The retinal irradiance producing a criterion PIPR was normalised to the peak and fitted by either a single photopigment nomogram or the combined melanopsin and rhodopsin spectral nomograms with the +L+M cone photopic luminous efficiency (Vλ) function. We show that the PIPR spectral sensitivity at times ≥ 1.7 s after light offset is best described by the melanopsin nomogram. At times < 1.7 s, the peak PIPR sensitivity shifts to longer wavelengths (range: 482 to 498 nm) and is best described by the combined photoreceptor nomogram, with major contributions from melanopsin and rhodopsin. This first report of melanopsin and rhodopsin contributions to the early phase PIPR is in line with the electrophysiological findings of ipRGC and rod signalling after the cessation of light stimuli and provides a cut-off time for isolating photoreceptor specific function in healthy and diseased eyes. PMID:27548480

  8. Modeling coupled sorption and transformation of 17β-estradiol-17-sulfate in soil-water systems

    NASA Astrophysics Data System (ADS)

    Bai, Xuelian; Shrestha, Suman L.; Casey, Francis X. M.; Hakk, Heldur; Fan, Zhaosheng

    2014-11-01

    Animal manure is the primary source of exogenous free estrogens in the environment, which are known endocrine-disrupting chemicals to disorder the reproduction system of organisms. Conjugated estrogens can act as precursors to free estrogens, which may increase the total estrogenicity in the environment. In this study, a comprehensive model was used to simultaneously simulate the coupled sorption and transformation of a sulfate estrogen conjugate, 17β-estradiol-17-sulfate (E2-17S), in various soil-water systems (non-sterile/sterile; topsoil/subsoil). The simulated processes included multiple transformation pathways (i.e. hydroxylation, hydrolysis, and oxidation) and mass transfer between the aqueous, reversibly sorbed, and irreversibly sorbed phases of all soils for E2-17S and its metabolites. The conceptual model was conceived based on a series of linear sorption and first-order transformation expressions. The model was inversely solved using finite difference to estimate process parameters. A global optimization method was applied for the inverse analysis along with variable model restrictions to estimate 36 parameters. The model provided a satisfactory simultaneous fit (R2adj = 0.93 and d = 0.87) of all the experimental data and reliable parameter estimates. This modeling study improved the understanding on fate and transport of estrogen conjugates under various soil-water conditions.

  9. Identification of neighboring protein pairs cross-linked with dimethyl 3,3'-dithiobispropionimidate in rat liver 40S ribosomal subunits.

    PubMed

    Uchiumi, T; Terao, K; Ogata, K

    1981-07-01

    Rat liver 40S ribosomal subunits were treated with a bifunctional imidoester, dimethyl 3,3'-dithiobispropionimidate (DTP), and the neighboring protein pairs were identified. The cross-linked proteins were analyzed by acrylamide/SDS diagonal gel electrophoresis (Sommer & Traut (1974) Proc. Natl. Acad. Sci. U.S. 71, 3946-3950). The cross-linked components that fell off the diagonal upon adding 2-mercaptoethanol in the second dimension were labeled with 125I in the acrylamide gel and identified by two-dimensional acrylamide/urea gel electrophoresis, followed by radioautography. Considering these results and the molecular weights, we propose the following ten pairs, according to our numbering system (Terao & Ogata (1975) Biochim. Biophys. Acta 402, 219-229): S3-S5 (S3/S3a-S4), S3-S14 (S3/S3a-S14), S3-S17 (S3/S3a-S16), S5-S22 (S4-S23/S24), S10-S12 (S8-S11), S9-S16 (S9-S18), S9-S22 (S9-S23/S24), S6-S23 (S5-S25), S17-S21 (S16-S19), and S16-S26 (S18-S27). The designation according to the proposed uniform nomenclature (McConkey et al. (1979) Mol. Gen. Genet. 169, 1-6) are given in parentheses.

  10. Loss of heterozygosity (LOH) in familial tumors of BRCA1 kindreds

    SciTech Connect

    Neuhausen, S.L.; Marshall, C.J.; Goldgar, D.E.

    1994-09-01

    In this study, we examined two families previously linked to BRCA1. Identification of recombinants in kindreds K2082 and K2035 had narrowed the region to between D17S776 and D17S1184, respectively. Further delineation of the region may be accomplished by examining loss of heterozygosity (LOH) in familial tumors. Five ovarian and six breast tumors from kindred K2082 (including two from sporadic breast cases) and three breast tumors from kindred K2035 were tested for LOH using six BRCA1-linked polymorphic STR markers and one marker at TP53. The two tumors from K2082 which did not share the linked haplotype did not show LOH in the BRCA1 region, nor did one of the tumors from a 17q-linked breast cancer case. Four ovarian and three breast tumors showed complete loss for the BRCA1 region. One ovarian and three breast tumors showed LOH only within the BRCA1 region. In all instances where LOH in the BRCA1 region was observed, the wild type allele was lost. These results therefore confirm the observation of Smith et al. (1992), and provide additional evidence that BRCA1 is a tumor suppressor gene. One of the breast tumors which showed loss within the BRCA1 region allowed us to narrow the region further.

  11. Chromosome 17q linkage studies of 18 Utah breast cancer kindreds

    SciTech Connect

    Goldgar, D.E.; Cannon-Albright, L.A.; Oliphant, A.; Ward, J.H.; Linker, G.; Swensen, J.; Tran, T.D.; Fields, P.; Uharriet, P.; Skolnick, M.H. )

    1993-04-01

    In this paper the authors present linkage results from the analysis of 18 Utah breast cancer kindreds, for three 17q markers. Four kindreds had LOD scores greater than 1.0 for at least one of the marker loci. One of these kindreds has a LOD score of 6.07 with D17S579, and they believe it to be the most informative 17q family reported to date. Among the kindreds which appear unlinked to 17q were an early-onset breast cancer family, a large breast-ovarian family, and a kindred with mixed age at onset. Analysis of individual recombinants in the linked families localizes the BRCA1 gene between THRA1 and D17S579 (Mfd188). A comparison of the Cancer and Steroid Hormone Study (CASH) model and a model which assumes a rare dominant susceptibility locus with low penetrance and no phenocopies stresses the difficulties in assessing linkage if the assumptions of the CASH model in terms of age at onset of breast cancer are not appropriate for the BRCA1 locus. A hypothetical breast cancer pedigree is used to calculate gene carrier probabilities under the CASH model, thereby illustrating some of the concerns regarding the use of this model to detect and exclude 17q linkage in breast cancer families.

  12. The human insulin-like growth factor-binding protein 4 gene maps to chromosome region 17q12-q21. 1 and is close to the gene for hereditary breast-ovarian cancer

    SciTech Connect

    Tonin, P.; Vivier, A.; Morgan, K.; Narod, S.; Pollack, M. ); Ehrenborg, E.; Zazzi, H.; Luthman, H.; Larsson, C. ); Lenoir, G. )

    1993-11-01

    The gene for insulin-like growth factor-binding protein 4 (IGFBP4) codes for a serum protein that binds to the family of insulin-like growth factors and modulates their activity. It has been mapped by in situ hybridization to chromosome region 17q12-q21.1. The authors have developed a CA-repeat polymorphism from a cosmid clone containing IGFBP4. By linkage analysis, IGFBP4 maps to the chromosome 17q interval THRA1-D17S579. This interval also contains the gene for hereditary breast-ovarian cancer, BRCA1. Genetic recombination between IGFBP4 and BRCA1 places IGFBP4 centromeric to the cancer susceptibility gene and effectively excludes it as a candidate gene for BRCA1. IGFBP4 is, however, one of the closest known centromeric markers for BRCA1; the estimated recombination fraction is 0.015. IGFBP4 and D17S579 together define a 2.8-cM interval that contains BRCA1. 18 refs., 3 figs., 1 tab.

  13. Accurate quantification of sphingosine-1-phosphate in normal and Fabry disease plasma, cells and tissues by LC-MS/MS with (13)C-encoded natural S1P as internal standard.

    PubMed

    Mirzaian, Mina; Wisse, Patrick; Ferraz, Maria J; Marques, André R A; Gabriel, Tanit L; van Roomen, Cindy P A A; Ottenhoff, Roelof; van Eijk, Marco; Codée, Jeroen D C; van der Marel, Gijsbert A; Overkleeft, Herman S; Aerts, Johannes M

    2016-08-01

    We developed a mass spectrometric procedure to quantify sphingosine-1-phosphate (S1P) in biological materials. The use of newly synthesized (13)C5 C18-S1P and commercial C17-S1P as internal standards rendered very similar results with respect to linearity, limit of detection and limit of quantitation. Caution is warranted with determination of plasma S1P levels. Earlier it was reported that S1P is elevated in plasma of Fabry disease patients. We investigated this with the improved quantification. No clear conclusion could be drawn for patient plasma samples given the lack of uniformity of blood collection and plasma preparation. To still obtain insight, plasma and tissues were identically collected from α-galactosidase A deficient Fabry mice and matched control animals. No significant difference was observed in plasma S1P levels. A significant 2.3 fold increase was observed in kidney of Fabry mice, but not in liver and heart. Comparative analysis of S1P in cultured fibroblasts from normal subjects and classically affected Fabry disease males revealed no significant difference. In conclusion, accurate quantification of S1P in biological materials is feasible by mass spectrometry using the internal standards (13)C5 C18-S1P or C17-S1P. Significant local increases of S1P in the kidney might occur in Fabry disease as suggested by the mouse model. PMID:27221202

  14. Berezinskii-Kosterlitz-Thouless phase transitions in two-dimensional non-Abelian spin models.

    PubMed

    Borisenko, Oleg; Chelnokov, Volodymyr; Cuteri, Francesca; Papa, Alessandro

    2016-07-01

    It is argued that two-dimensional U(N) spin models for any N undergo a Berezinskii-Kosterlitz-Thouless (BKT)-like phase transition, similarly to the famous XY model. This conclusion follows from the Berezinskii-like calculation of the two-point correlation function in U(N) models, approximate renormalization group analysis, and numerical investigations of the U(2) model. It is shown, via Monte Carlo simulations, that the universality class of the U(2) model coincides with that of the XY model. Moreover, preliminary numerical results point out that two-dimensional SU(N) spin models with the fundamental and adjoint terms and N>4 exhibit two phase transitions of BKT type, similarly to Z(N) vector models. PMID:27575078

  15. Berezinskii-Kosterlitz-Thouless phase transitions in two-dimensional non-Abelian spin models

    NASA Astrophysics Data System (ADS)

    Borisenko, Oleg; Chelnokov, Volodymyr; Cuteri, Francesca; Papa, Alessandro

    2016-07-01

    It is argued that two-dimensional U(N ) spin models for any N undergo a Berezinskii-Kosterlitz-Thouless (BKT)-like phase transition, similarly to the famous X Y model. This conclusion follows from the Berezinskii-like calculation of the two-point correlation function in U(N ) models, approximate renormalization group analysis, and numerical investigations of the U(2 ) model. It is shown, via Monte Carlo simulations, that the universality class of the U(2 ) model coincides with that of the X Y model. Moreover, preliminary numerical results point out that two-dimensional SU(N ) spin models with the fundamental and adjoint terms and N >4 exhibit two phase transitions of BKT type, similarly to Z (N ) vector models.

  16. Severe storm electricity

    NASA Technical Reports Server (NTRS)

    Rust, W. D.; Macgorman, D. R.

    1985-01-01

    During FY-85, Researchers conducted a field program and analyzed data. The field program incorporated coordinated measurements made with a NASA U2. Results include the following: (1) ground truth measurements of lightning for comparison with those obtained by the U2; (2) analysis of dual-Doppler radar and dual-VHF lightning mapping data from a supercell storm; (3) analysis of synoptic conditions during three simultaneous storm systems on 13 May 1983 when unusually large numbers of positive cloud-to-ground (+CG) flashes occurred; (4) analysis of extremely low frequency (ELF) wave forms; and (5) an assessment of a cloud -ground strike location system using a combination of mobile laboratory and fixed-base TV video data.

  17. Global Membrane Protein Interactome Analysis using In vivo Crosslinking and Mass Spectrometry-based Protein Correlation Profiling*

    PubMed Central

    Larance, Mark; Kirkwood, Kathryn J.; Tinti, Michele; Brenes Murillo, Alejandro; Ferguson, Michael A. J.; Lamond, Angus I.

    2016-01-01

    We present a methodology using in vivo crosslinking combined with HPLC-MS for the global analysis of endogenous protein complexes by protein correlation profiling. Formaldehyde crosslinked protein complexes were extracted with high yield using denaturing buffers that maintained complex solubility during chromatographic separation. We show this efficiently detects both integral membrane and membrane-associated protein complexes,in addition to soluble complexes, allowing identification and analysis of complexes not accessible in native extracts. We compare the protein complexes detected by HPLC-MS protein correlation profiling in both native and formaldehyde crosslinked U2OS cell extracts. These proteome-wide data sets of both in vivo crosslinked and native protein complexes from U2OS cells are freely available via a searchable online database (www.peptracker.com/epd). Raw data are also available via ProteomeXchange (identifier PXD003754). PMID:27114452

  18. Optimization of small long-life PWR based on thorium fuel

    SciTech Connect

    Subkhi, Moh Nurul; Suud, Zaki Waris, Abdul; Permana, Sidik

    2015-09-30

    A conceptual design of small long-life Pressurized Water Reactor (PWR) using thorium fuel has been investigated in neutronic aspect. The cell-burn up calculations were performed by PIJ SRAC code using nuclear data library based on JENDL 3.2, while the multi-energy-group diffusion calculations were optimized in three-dimension X-Y-Z geometry of core by COREBN. The excess reactivity of thorium nitride with ZIRLO cladding is considered during 5 years of burnup without refueling. Optimization of 350 MWe long life PWR based on 5% {sup 233}U & 2.8% {sup 231}Pa, 6% {sup 233}U & 2.8% {sup 231}Pa and 7% {sup 233}U & 6% {sup 231}Pa give low excess reactivity.

  19. Exploring flavor structure of supersymmetry breaking from rare B decays and the unitarity triangle

    NASA Astrophysics Data System (ADS)

    Goto, Toru; Okada, Yasuhiro; Shimizu, Yasuhiro; Shindou, Tetsuo; Tanaka, Minoru

    2004-08-01

    We study effects of supersymmetric particles in various rare B decay processes as well as in the unitarity triangle analysis. We consider three different supersymmetric models, the minimal supergravity, SU(5) SUSY GUT with right-handed neutrinos, and the minimal supersymmetric standard model with U(2) flavor symmetry. In the SU(5) SUSY GUT with right-handed neutrinos, we consider two cases of the mass matrix of the right-handed neutrinos. We calculate direct and mixing-induced CP asymmetries in the b→sγ decay and CP asymmetry in Bd→φKS as well as the Bs Bs mixing amplitude for the unitarity triangle analysis in these models. We show that large deviations are possible for the SU(5) SUSY GUT and the U(2) model. The patterns and correlations of deviations from the standard model will be useful to discriminate the different SUSY models in future B experiments.

  20. Theory of high Tc ferrimagnetism in a multiorbital Mott insulator.

    PubMed

    Meetei, O Nganba; Erten, Onur; Randeria, Mohit; Trivedi, Nandini; Woodward, Patrick

    2013-02-22

    We propose a model for the multiorbital material Sr(2)CrOsO(6), an insulator with remarkable magnetic properties and the highest T(c) ~/= 725 K among all perovskites with a net moment. We derive a new criterion for the Mott transition (U(1)U(2))(1/2)>2.5W by using slave-rotor mean field theory, where W is the bandwidth and U(1(2)) are the effective Coulomb interactions on Cr(Os) including Hund's coupling. We show that Sr(2)CrOsO(6) is a Mott insulator, where the large Cr U(1) compensates for the small Os U(2). The spin sector is described by a frustrated antiferromagnetic Heisenberg model that naturally explains the net moment arising from canting and also the observed nonmonotonic magnetization M(T). We predict characteristic magnetic structure factor peaks that can be probed by neutron experiments.