All Small Nuclear RNAs (snRNAs) of the [U4/U6.U5] Tri-snRNP Localize to Nucleoli; Identification of the Nucleolar Localization Element of U6 snRNA

Science.gov (United States)

Gerbi, Susan A.; Lange, Thilo Sascha

2002-01-01

Previously, we showed that spliceosomal U6 small nuclear RNA (snRNA) transiently passes through the nucleolus. Herein, we report that all individual snRNAs of the [U4/U6.U5] tri-snRNP localize to nucleoli, demonstrated by fluorescence microscopy of nucleolar preparations after injection of fluorescein-labeled snRNA into Xenopus oocyte nuclei. Nucleolar localization of U6 is independent from [U4/U6] snRNP formation since sites of direct interaction of U6 snRNA with U4 snRNA are not nucleolar localization elements. Among all regions in U6, the only one required for nucleolar localization is its 3′ end, which associates with the La protein and subsequently during maturation of U6 is bound by Lsm proteins. This 3′-nucleolar localization element of U6 is both essential and sufficient for nucleolar localization and also required for localization to Cajal bodies. Conversion of the 3′ hydroxyl of U6 snRNA to a 3′ phosphate prevents association with the La protein but does not affect U6 localization to nucleoli or Cajal bodies. PMID:12221120

Crystallization and preliminary X-ray analysis of a U2AF65 variant in complex with a polypyrimidine-tract analogue by use of protein engineering

International Nuclear Information System (INIS)

Sickmier, E. Allen; Frato, Katherine E.; Kielkopf, Clara L.

2006-01-01

A complex of the essential splicing factor U2AF 65 and a deoxyuridine oligonucleotide has been crystallized by modification of an interdomain linker. The large subunit of the essential pre-mRNA splicing factor U2 auxiliary factor (U2AF 65 ) binds the polypyrimidine tract near the 3′ splice site of pre-mRNA introns and directs the association of the U2 small nuclear ribonucleoprotein particle (U2 snRNP) of the spliceosome with the pre-mRNA. Protein engineering, in which the flexible linker region connecting tandem RNA-recognition motifs (RRMs) within the U2AF 65 RNA-binding domain was partially deleted, allowed successful crystallization of the protein–nucleic acid complex. Cocrystals of a U2AF 65 variant with a deoxyuridine dodecamer diffract X-rays to 2.9 Å resolution and contain one complex per asymmetric unit

The Arabidopsis splicing factors, AtU2AF65, AtU2AF35, and AtSF1 shuttle between nuclei and cytoplasms

KAUST Repository

Park, Hyo-Young

2017-04-21

The Arabidopsis splicing factors, AtU2AF65, AtU2AF35, and AtSF1 shuttle between nuclei and cytoplasms. These proteins also move rapidly and continuously in the nuclei, and their movements are affected by ATP depletion. The U2AF65 proteins are splicing factors that interact with SF1 and U2AF35 proteins to promote U2snRNP for the recognition of the pre-mRNA 3\\' splice site during early spliceosome assembly. We have determined the subcellular localization and movement of these proteins\\' Arabidopsis homologs. It was found that Arabidopsis U2AF65 homologs, AtU2AF65a, and AtU2AF65b proteins interact with AtU2AF35a and AtU2AF35b, which are Arabidopsis U2AF35 homologs. We have examined the mobility of these proteins including AtSF1 using fluorescence recovery after photobleaching and fluorescence loss in photobleaching analyses. These proteins displayed dynamic movements in nuclei and their movements were affected by ATP depletion. We have also demonstrated that these proteins shuttle between nuclei and cytoplasms, suggesting that they may also function in cytoplasm. These results indicate that such splicing factors show very similar characteristics to their human counterparts, suggesting evolutionary conservation.

The Arabidopsis splicing factors, AtU2AF65, AtU2AF35, and AtSF1 shuttle between nuclei and cytoplasms

KAUST Repository

Park, Hyo-Young; Lee, Keh Chien; Jang, Yun Hee; Kim, SoonKap; Thu, May Phyo; Lee, Jeong Hwan; Kim, Jeong-Kook

2017-01-01

The Arabidopsis splicing factors, AtU2AF65, AtU2AF35, and AtSF1 shuttle between nuclei and cytoplasms. These proteins also move rapidly and continuously in the nuclei, and their movements are affected by ATP depletion. The U2AF65 proteins are splicing factors that interact with SF1 and U2AF35 proteins to promote U2snRNP for the recognition of the pre-mRNA 3' splice site during early spliceosome assembly. We have determined the subcellular localization and movement of these proteins' Arabidopsis homologs. It was found that Arabidopsis U2AF65 homologs, AtU2AF65a, and AtU2AF65b proteins interact with AtU2AF35a and AtU2AF35b, which are Arabidopsis U2AF35 homologs. We have examined the mobility of these proteins including AtSF1 using fluorescence recovery after photobleaching and fluorescence loss in photobleaching analyses. These proteins displayed dynamic movements in nuclei and their movements were affected by ATP depletion. We have also demonstrated that these proteins shuttle between nuclei and cytoplasms, suggesting that they may also function in cytoplasm. These results indicate that such splicing factors show very similar characteristics to their human counterparts, suggesting evolutionary conservation.

Variability in clinical phenotypes of PRPF8-linked autosomal dominant retinitis pigmentosa correlates with differential PRPF8/SNRNP200 interactions.

Science.gov (United States)

Escher, Pascal; Passarin, Olga; Munier, Francis L; Tran, Viet H; Vaclavik, Veronika

2018-01-01

To expand the genotype/phenotype correlations in patients with autosomal dominant retinitis pigmentosa (adRP) harboring PRPF8 variants. Two patients, a father and his daughter, harboring a novel p.PRPF8-Glu2331* variant, underwent ophthalmic examination at 3-year-interval, including fundus photography, fundus autofluorescence, optical coherence tomography, and ISCEV standard full field ERGs. All reported disease-causing PRPF8 variants were collected and localized in the PRPF8 and PRPF8/SNRNP200 protein structures. The p.PRPF8-Glu2331* variant results in a truncated PRPF8 protein lacking the last five C-terminal amino acids and caused in the two patients a severe clinical phenotype, with the macula being affected from the second decade on. All but two adRP-linked variants are located in the last exon 43 encoding the C-terminal tail of the C-terminal PRPF8 Jab1 domain. The p.PRPF8-Ser2118Phe and -Asn2280Lys variants encoded by exons 39 and 42, respectively, are located at the basis of the C-terminal tail. Frame-shift mutations and nonconservative amino acid changes in PRPF8 typically cause severe clinical phenotypes. The conservative missense variant p.PRPF8-Arg2310Lys that is not altering the global charge of the C-terminal tail, and variants located at the basis of the C-terminal tail show milder clinical phenotypes, in accordance with functional data on PRPF8/SNRNP200 interactions in yeast.

U bodies respond to nutrient stress in Drosophila

International Nuclear Information System (INIS)

Buckingham, Mickey; Liu, Ji-Long

2011-01-01

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.

U bodies respond to nutrient stress in Drosophila

Energy Technology Data Exchange (ETDEWEB)

Buckingham, Mickey; Liu, Ji-Long, E-mail: jilong.liu@dpag.ox.ac.uk

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.

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

Science.gov (United States)

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. © 2016 Schwer et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

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

Science.gov (United States)

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

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

OpenAIRE

Weber, G.; Trowitzsch, S.; Kastner, B.; Lührmann, R.; Wahl, M.

2010-01-01

The U1 small nuclear ribonucleoprotein initiates the assembly of the spliceosome. Here, the structure of the natively purified U1 small nuclear ribonucleoprotein particle reveals the core Sm protein ring and its interactions with the Sm site in the small nuclear RNA.

Cajal bodies and snRNPs friends with benefits

Czech Academy of Sciences Publication Activity Database

Staněk, David

2017-01-01

Roč. 14, č. 6 (2017), s. 671-679 ISSN 1547-6286 R&D Projects: GA ČR GA15-00790S Institutional support: RVO:68378050 Keywords : spinal muscular-atrophy * small nuclear-rna * u4/u6.u5 tri-snrnp * xenopus-laevis oocytes * u6 spliceosomal rna * coiled bodies * smn complex * u1 snrnp * u2 snrnp * in-vivo Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Cell biology Impact factor: 3.900, year: 2016

Dual RNA Processing Roles of Pat1b via Cytoplasmic Lsm1-7 and Nuclear Lsm2-8 Complexes

Directory of Open Access Journals (Sweden)

Caroline Vindry

2017-08-01

Full Text Available Pat1 RNA-binding proteins, enriched in processing bodies (P bodies, are key players in cytoplasmic 5′ to 3′ mRNA decay, activating decapping of mRNA in complex with the Lsm1-7 heptamer. Using co-immunoprecipitation and immunofluorescence approaches coupled with RNAi, we provide evidence for a nuclear complex of Pat1b with the Lsm2-8 heptamer, which binds to the spliceosomal U6 small nuclear RNA (snRNA. Furthermore, we establish the set of interactions connecting Pat1b/Lsm2-8/U6 snRNA/SART3 and additional U4/U6.U5 tri-small nuclear ribonucleoprotein particle (tri-snRNP components in Cajal bodies, the site of snRNP biogenesis. RNA sequencing following Pat1b depletion revealed the preferential upregulation of mRNAs normally found in P bodies and enriched in 3′ UTR AU-rich elements. Changes in >180 alternative splicing events were also observed, characterized by skipping of regulated exons with weak donor sites. Our data demonstrate the dual role of a decapping enhancer in pre-mRNA processing as well as in mRNA decay via distinct nuclear and cytoplasmic Lsm complexes.

Sequence Classification: 893627 [

Lifescience Database Archive (English)

Full Text Available Non-TMB Non-TMH Non-TMB Non-TMB Non-TMB Non-TMB >gi|6325043|ref|NP_015111.1| Component of U2 snRNP; disrupti...on causes reduced U2 snRNP levels; physically interacts with Msl1p; putative homolo

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

International Nuclear Information System (INIS)

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.

2008-01-01

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

Fanconi anemia FANCD2 and FANCI proteins regulate the nuclear dynamics of splicing factors.

Science.gov (United States)

Moriel-Carretero, María; Ovejero, Sara; Gérus-Durand, Marie; Vryzas, Dimos; Constantinou, Angelos

2017-12-04

Proteins disabled in the cancer-prone disorder Fanconi anemia (FA) ensure the maintenance of chromosomal stability during DNA replication. FA proteins regulate replication dynamics, coordinate replication-coupled repair of interstrand DNA cross-links, and mitigate conflicts between replication and transcription. Here we show that FANCI and FANCD2 associate with splicing factor 3B1 (SF3B1), a key spliceosomal protein of the U2 small nuclear ribonucleoprotein (U2 snRNP). FANCI is in close proximity to SF3B1 in the nucleoplasm of interphase and mitotic cells. Furthermore, we find that DNA replication stress induces the release of SF3B1 from nuclear speckles in a manner that depends on FANCI and on the activity of the checkpoint kinase ATR. In chromatin, both FANCD2 and FANCI associate with SF3B1, prevent accumulation of postcatalytic intron lariats, and contribute to the timely eviction of splicing factors. We propose that FANCD2 and FANCI contribute to the organization of functional domains in chromatin, ensuring the coordination of DNA replication and cotranscriptional processes. © 2017 Moriel-Carretero et al.

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

International Nuclear Information System (INIS)

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

Biochemical function of typical and variant Arabidopsis thaliana U-box E3 ubiquitin-protein ligases

DEFF Research Database (Denmark)

Wiborg, Jakob; O'Shea, Charlotte; Skriver, Karen

2008-01-01

of the distant U-box protein, AtPUB49, representing a large family of eukaryotic proteins containing a U-box linked to a cyclophilin-like peptidyl-prolyl cis-trans isomerase domain, was characterized biochemically. AtPUB49 functioned both as a prolyl isomerase and a chaperone by catalysing cis......The variance of the U-box domain in 64 Arabidopsis thaliana (thale cress) E3s (ubiquitin-protein ligases) was used to examine the interactions between E3s and E2s (ubiquitin-conjugating enzymes). E2s and E3s are components of the ubiquitin protein degradation pathway. Seven U-box proteins were...... analysed for their ability to ubiquitinate proteins in vitro in co-operation with different E2s. All U-box domains exhibited ubiquitination activity and interacted productively with UBC4/5-type E2s. Three and four of the U-box domains mediated ubiquitin addition in the presence of UBC13 and UBC7 E2s...

The translation initiation factor eIF4E regulates the sex-specific expression of the master switch gene Sxl in Drosophila melanogaster.

Directory of Open Access Journals (Sweden)

Patricia L Graham

2011-07-01

Full Text Available 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(2d--that have been directly implicated in Sxl splicing regulation.

The urokinase receptor (uPAR) and the uPAR-associated protein (uPARAP/Endo180)

DEFF Research Database (Denmark)

Behrendt, Niels

2004-01-01

The breakdown of the barriers formed by extracellular matrix proteins is a pre-requisite for all processes of tissue remodeling. Matrix degradation reactions take part in specific physiological events in the healthy organism but also represent a crucial step in cancer invasion. These degradation...... on the surface of various cell types that serves to bind the urokinase plasminogen activator and localize the activation reactions in the proteolytic cascade system of plasminogen activation. uPARAP is an integral membrane protein with a pronounced role in the internalization of collagen for intracellular...... degradation. Both receptors have additional functions that are currently being unraveled. The present discussion of uPAR and uPARAP is centered on their protein structure and molecular and cellular function....

Biochemical function of typical and variant Arabidopsis thaliana U-box E3 ubiquitin-protein ligases.

Science.gov (United States)

Wiborg, Jakob; O'Shea, Charlotte; Skriver, Karen

2008-08-01

The variance of the U-box domain in 64 Arabidopsis thaliana (thale cress) E3s (ubiquitin-protein ligases) was used to examine the interactions between E3s and E2s (ubiquitin-conjugating enzymes). E2s and E3s are components of the ubiquitin protein degradation pathway. Seven U-box proteins were analysed for their ability to ubiquitinate proteins in vitro in co-operation with different E2s. All U-box domains exhibited ubiquitination activity and interacted productively with UBC4/5-type E2s. Three and four of the U-box domains mediated ubiquitin addition in the presence of UBC13 and UBC7 E2s respectively, but no productive interaction was observed with the UBC15 E2 tested. The activity of AtPUB54 [Arabidopsis thaliana (thale cress) plant U-box 54 protein] was dependent on Trp(266) in the E2-binding cleft, and the E2 selectivity was changed by substitution of this position. The function of the distant U-box protein, AtPUB49, representing a large family of eukaryotic proteins containing a U-box linked to a cyclophilin-like peptidyl-prolyl cis-trans isomerase domain, was characterized biochemically. AtPUB49 functioned both as a prolyl isomerase and a chaperone by catalysing cis-trans isomerization of peptidyl-prolyl bonds and dissolving protein aggregates. In conclusion, both typical and atypical Arabidopsis U-box proteins were active E3s. The overlap in the E3/E2 selectivity suggests that in vivo specificity is not determined only by the E3-E2 interactions, but also by other parameters, e.g. co-existence or interactions with additional domains. The biochemical functions of AtPUB49 suggest that the protein can be involved in folding or degradation of protein substrates. Similar functions can also be retained within a protein complex with separate chaperone and U-box proteins.

Novel monoclonal autoantibody specificity associated with ribonucleoprotein complexes

International Nuclear Information System (INIS)

Winkler, A.; Watson-McKown, R.; Wise, K.

1986-01-01

The authors describe an IgG/sub 2a/, kappa monoclonal autoantibody (mAb) F78 derived from a 6-month old MRL-Mp lpr/lpr mouse that recognizes a novel epitope associated with small nuclear ribonuclear protein complexes (snRNP). Indirect immunofluorescent staining of HEp-2 cells with F78 showed a nonnucleolar speckled nuclear pattern characteristic of anti-RNP and anti-Sm mAbs which could be abrogated by pretreating fixed cells with 0.1M HCl prior to staining. Immunoblots of whole cell extracts (dissociated in SDS, urea and mercaptan at 4 0 C then subjected to SDS-PAGE) showed that F78 selectively bound to a component of M/sub r/ = 100,000 clearly distinct from components recognized by two mAbs described by Billings et al that detected, respectively, proteins of M/sub r/ = 70,000 associated with RNP and M/sub r/ = 13,000 associated with Sm. Incubation of extracts at 100 0 C prior to SDS-PAGE eliminated subsequent binding of F78 but not of the other nAbs. F78 as well as the other mAbs selectively immunoprecipitated characteristic patterns of small nuclear RNAs (U 1 , U 2 , U 4 , U 5 , U 6 ) from extracts of 32 P-phosphate labeled HeLa cells. These results suggest a new specificity associated with snRNP that is recognized in the MRL autoimmune response

Nuclear bodies: news insights into structure and function

Czech Academy of Sciences Publication Activity Database

Staněk, David; Fox, A.H.

2017-01-01

Roč. 46, léto (2017), s. 94-101 ISSN 0955-0674 R&D Projects: GA ČR GA15-00790S; GA MŠk LO1419 Institutional support: RVO:68378050 Keywords : rna-binding proteins * cajal bodies * smn complex * human-cells * in-vivo * human telomerase * coiled bodies * lncrna neat1 * u1 snrnp * body Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Cell biology Impact factor: 9.937, year: 2016

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

Directory of Open Access Journals (Sweden)

Marianna Rakszegi

2017-09-01

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

B-CELL EPITOPE ON THE U1 SNRNP-C AUTOANTIGEN CONTAINS A SEQUENCE SIMILAR TO THAT OF THE HERPES-SIMPLEX VIRUS PROTEIN

NARCIS (Netherlands)

MISAKI, Y; YAMAMOTO, K; YANAGI, K; MIURA, H; ICHIJO, H; KATO, T; MATO, T; WELLINGWESTER, S; NISHIOKA, K; ITO, K

The mechanism of autoantibody production in autoimmune diseases is not well understood. In the present study we performed the B cell epitope mapping of the U1 small nuclear ribonucleoprotein (snRNP)-C, one of the target molecules of anti-nRNP autoantibody to investigate how B cells respond to the

Changes in nuclear protein acetylation in u.v.-damaged human cells

International Nuclear Information System (INIS)

Ramanathan, B.; Smerdon, M.J.

1986-01-01

The levels of nuclear protein acetylation in u.v.-irradiated human fibroblasts have been investigated. Initially, we measured the levels of acetylation in total acid-soluble nuclear proteins and observed two distinct differences between the irradiated and unirradiated (control) cells. Immediately after irradiation, there is a 'wave' of protein hyperacetylation that lasts for 2-6 h, followed by a hypoacetylation phase, lasting for many hours, and the total level of acetylation does not return to that of control cells until 24-72 h after u.v. damage. Both the magnitude and duration of each phase is dependent on the dose of u.v. light used. The wave of hyperacetylation is more pronounced at low u.v. doses, while the wave of hypoacetylation is more pronounced at higher u.v. doses. Furthermore, the duration of each phase is prolonged when cells are exposed to 2 mM hydroxyurea, an agent which retards the rate of excision repair at u.v.-damaged sites. Examinations of the acetylation levels of the individual nuclear proteins indicated that acetylation of the core histones follows the same pattern observed for the total acid-soluble protein fractions. Furthermore, these were the only major proteins in the total acid-soluble fraction observed to undergo the early, rapid hyperacetylation immediately following u.v. damage. These results raise the possibility that a causal relationship exists between nuclear protein acetylation and nucleotide excision repair of DNA in human cells. (author)

Changes in nuclear protein acetylation in u. v. -damaged human cells

Energy Technology Data Exchange (ETDEWEB)

Ramanathan, B.; Smerdon, M.J.

1986-07-01

We have investigated the levels of nuclear protein acetylation in u.v.-irradiated human fibroblasts. We measured the levels of acetylation in total acid-soluble nuclear proteins and observed two distinct differences between the irradiated and unirradiated (control) cells. Immediately after irradiation, there is a wave of protein hyperacetylation (i.e. a total acetylation level greater than that of unirradiated cells) that lasts for 2-6 h depending on the experimental conditions. This hyperacetylation phase is then followed by a hypoacetylation phase, lasting for many hours, and the total level of acetylation does not return to that of control cells until 24-72 h after u.v. damage. Both the magnitude and duration of each phase is dependent on the dose of u.v. light used. The wave of hyperacetylation is more pronounced at low u.v. doses (i.e. less than 5 J/m2), while the wave of hypoacetylation is more pronounced at higher u.v. doses (greater than or equal to 8 J/m2). Furthermore, the duration of each phase is prolonged when cells are exposed to 2 mM hydroxyurea. Examination of the acetylation levels of the individual nuclear proteins indicated that acetylation of the core histones follows the same pattern observed for the total acid-soluble protein fractions. Furthermore, these were the only major proteins in the total acid-soluble fraction observed to undergo the early, rapid hyperacetylation immediately following u.v. damage. Acetylation of histone H1 was negligible in both damaged and control cells, while three prominent non-histone proteins were acetylated only after long labeling times (greater than 4 h) in each case, gradually becoming hyperacetylated in the u.v.-damaged cells. These results raise the possibility that a causal relationship exists between nuclear protein acetylation and nucleotide excision repair of DNA in human cells.

Crystallization and preliminary X-ray diffraction analysis of the C-terminal domain of the human spliceosomal DExD/H-box protein hPrp22

International Nuclear Information System (INIS)

Kudlinzki, Denis; Nagel, Christian; Ficner, Ralf

2009-01-01

The cloning, purification and crystallization of the C-terminal domain of human hPrp22 are reported. This communication also contains data for the preliminary X-ray diffraction analysis. The Homo sapiens DExD/H-box protein hPrp22 is a crucial component of the eukaryotic pre-mRNA splicing machinery. Within the splicing cycle, it is involved in the ligation of exons and generation of the lariat and it additionally catalyzes the release of mature mRNA from the spliceosomal U5 snRNP. The yeast homologue of this protein, yPrp22, shows ATP-dependent RNA-helicase activity and is capable of unwinding RNA/RNA duplex molecules. A truncated construct coding for residues 950–1183 of human Prp22, comprising the structurally and functionally uncharacterized C-terminal domain, was cloned into an Escherichia coli expression vector. The protein was subsequently overproduced, purified and crystallized. The crystals obtained diffracted to 2.1 Å resolution, belonged to the tetragonal space group P4 1 2 1 2 or P4 3 2 1 2, with unit-cell parameters a = b = 78.2, c = 88.4 Å, and contained one molecule in the asymmetric unit

Human parvovirus B19 VP1u Protein as inflammatory mediators induces liver injury in naïve mice.

Science.gov (United States)

Hsu, Tsai-Ching; Chiu, Chun-Ching; Chang, Shun-Chih; Chan, Hsu-Chin; Shi, Ya-Fang; Chen, Tzy-Yen; Tzang, Bor-Show

2016-01-01

Human parvovirus B19 (B19V) is a human pathogen known to be associated with many non-erythroid diseases, including hepatitis. Although B19V VP1-unique region (B19-VP1u) has crucial roles in the pathogenesis of B19V infection, the influence of B19-VP1u proteins on hepatic injury is still obscure. This study investigated the effect and possible inflammatory signaling of B19-VP1u in livers from BALB/c mice that were subcutaneously inoculated with VP1u-expressing COS-7 cells. The in vivo effects of B19-VP1u were analyzed by using live animal imaging system (IVIS), Haematoxylin-Eosin staining, gel zymography, and immunoblotting after inoculation. Markedly hepatocyte disarray and lymphocyte infiltration, enhanced matrix metalloproteinase (MMP)-9 activity and increased phosphorylation of p38, ERK, IKK-α, IκB and NF-κB (p-p65) proteins were observed in livers from BALB/c mice receiving COS-7 cells expressing B19-VP1u as well as the significantly increased CRP, IL-1β and IL-6. Notably, IFN-γ and phosphorylated STAT1, but not STAT3, were also significantly increased in the livers of BALB/c mice that were subcutaneously inoculated with VP1u-expressing COS-7 cells. These findings revealed the effects of B19-VP1u on liver injury and suggested that B19-VP1u may have a role as mediators of inflammation in B19V infection.

Effect of dietary protein on the excretion of. cap alpha. /sub 2u/, the sex-dependent protein of the adult male rat

Energy Technology Data Exchange (ETDEWEB)

Neuhaus, O W; Flory, W

1975-01-01

Adult male rates were maintained on normal (20 percent casein), protein-free (0 percent casein), high protein (50 percent casein), deficient protein (20 percent zein), and a supplemented, deficient protein (20 percent zein plus L-lysine and L-tryptophan) diets. Rats on a protein-free diet excreted approximately 1 mg ..cap alpha../sub 2u//24 h compared with a normal of 10-15 mg/24 h. Depleted rats placed on the normal diet showed a rapid restoration of the normal ..cap alpha../sub 2u/ excretion as well as total urinary proteins. Accumulation of ..cap alpha../sub 2u/ in the blood serum was measured in nephrectomized rats. Rats on the protein free diet accumulated only 30 percent of the ..cap alpha../sub 2u/ compared to normals. On a 50 precent casein diet, rats excreted 30-50 mg ..cap alpha../sub 2u//24 h. However, the accumulation was normal in the serum of nephrectomized rats. A high protein diet did not stimulate ..cap alpha../sub 2u/ synthesis but probably increased the renal loss of all urinary proteins. The excretion of ..cap alpha../sub 2u/ on a zein diet was reduced to the same degree as with the protein-free diet. Supplementation with lysine and tryptophan restored the capacity to eliminate ..cap alpha../sub 2u/ to near normal levels. Accumulation of ..cap alpha../sub 2u/ in the serum of nephrectomized rats kept on the zein diets showed that the effect was to suppress the synthesis of the ..cap alpha../sub 2u/. Supplementation restored the biosynthesis of ..cap alpha../sub 2u/. It is concluded that the effect of dietary protein on the excretion of urinary proteins in the adult male rat is caused in a large part by an influence on the hepatic biosynthesis of ..cap alpha../sub 2u/. The biosynthesis of this protein, which represents approximately 30 percent of the total urinary proteins, is dependent on an adequate supply of dietary protein.

A mutation linked to retinitis pigmentosa in HPRP31 causes protein instability and impairs its interactions with spliceosomal snRNPs

Czech Academy of Sciences Publication Activity Database

Huranová, Martina; Hnilicová, Jarmila; Fleischer, Branislav; Cvačková, Zuzana; Staněk, David

2009-01-01

Roč. 18, č. 11 (2009), s. 2014-2023 ISSN 0964-6906 R&D Projects: GA AV ČR KAN200520801 Grant - others: Max Planck Society(DE) Partner group program Institutional research plan: CEZ:AV0Z50520514 Keywords : retinitis pigmentosa * snRNP * splicing Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.386, year: 2009

7 CFR 51.3741 - U.S. Commercial.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Commercial. 51.3741 Section 51.3741 Agriculture... Standards for Grades of Honey Dew and Honey Ball Type Melons Grades § 51.3741 U.S. Commercial. “U.S. Commercial” consists of honey dew or honey ball type melons which meet the requirements of U.S. No. 1 grade...

7 CFR 51.2278 - U.S. Commercial.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Commercial. 51.2278 Section 51.2278 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing... Standards for Shelled English Walnuts (Juglans Regia) Grades § 51.2278 U.S. Commercial. “U.S. Commercial...

7 CFR 51.2647 - U.S. Commercial.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Commercial. 51.2647 Section 51.2647 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing... Standards for Grades for Sweet Cherries 1 Grades § 51.2647 U.S. Commercial. “U.S. Commercial” consists of...

7 CFR 51.1542 - U.S. Commercial.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Commercial. 51.1542 Section 51.1542 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing... Standards for Grades of Potatoes 1 Grades § 51.1542 U.S. Commercial. “U.S. Commercial” consists of potatoes...

Protein: FBA7 [TP Atlas

Lifescience Database Archive (English)

Full Text Available FBA7 claudin-zona occluden Tjp1 Zo1 Tight junction protein ZO-1 Tight junction protein 1, Zona occludens pr...otein 1, Zonula occludens protein 1 10090 Mus musculus 21872 P39447 2RRM P39447 21431884 ...

R7-binding protein targets the G protein β5/R7-regulator of G protein signaling complex to lipid rafts in neuronal cells and brain

Directory of Open Access Journals (Sweden)

Zhang Jian-Hua

2007-09-01

Full Text Available Abstract Background Heterotrimeric guanine nucleotide-binding regulatory proteins (G proteins, composed of Gα, Gβ, and Gγ subunits, are positioned at the inner face of the plasma membrane and relay signals from activated G protein-coupled cell surface receptors to various signaling pathways. Gβ5 is the most structurally divergent Gβ isoform and forms tight heterodimers with regulator of G protein signalling (RGS proteins of the R7 subfamily (R7-RGS. The subcellular localization of Gβ 5/R7-RGS protein complexes is regulated by the palmitoylation status of the associated R7-binding protein (R7BP, a recently discovered SNARE-like protein. We investigate here whether R7BP controls the targeting of Gβ5/R7-RGS complexes to lipid rafts, cholesterol-rich membrane microdomains where conventional heterotrimeric G proteins and some effector proteins are concentrated in neurons and brain. Results We show that endogenous Gβ5/R7-RGS/R7BP protein complexes are present in native neuron-like PC12 cells and that a fraction is targeted to low-density, detergent-resistant membrane lipid rafts. The buoyant density of endogenous raft-associated Gβ5/R7-RGS protein complexes in PC12 cells was similar to that of lipid rafts containing the palmitoylated marker proteins PSD-95 and LAT, but distinct from that of the membrane microdomain where flotillin was localized. Overexpression of wild-type R7BP, but not its palmitoylation-deficient mutant, greatly enriched the fraction of endogenous Gβ5/R7-RGS protein complexes in the lipid rafts. In HEK-293 cells the palmitoylation status of R7BP also regulated the lipid raft targeting of co-expressed Gβ5/R7-RGS/R7BP proteins. A fraction of endogenous Gβ5/R7-RGS/R7BP complexes was also present in lipid rafts in mouse brain. Conclusion A fraction of Gβ5/R7-RGS/R7BP protein complexes is targeted to low-density, detergent-resistant membrane lipid rafts in PC12 cells and brain. In cultured cells, the palmitoylation status of

7 CFR 51.883 - U.S. Fancy Export.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Fancy Export. 51.883 Section 51.883 Agriculture... Standards for Grades of Table Grapes (European or Vinifera Type) 1 Grades § 51.883 U.S. Fancy Export. “U.S. Fancy Export” consists of grapes which meet the requirements for U.S. Fancy Table, except that bunches...

Structural studies of the Enterococcus faecalis SufU [Fe-S] cluster protein

Directory of Open Access Journals (Sweden)

Frazzon Jeverson

2009-02-01

Full Text Available Abstract Background Iron-sulfur clusters are ubiquitous and evolutionarily ancient inorganic prosthetic groups, the biosynthesis of which depends on complex protein machineries. Three distinct assembly systems involved in the maturation of cellular Fe-S proteins have been determined, designated the NIF, ISC and SUF systems. Although well described in several organisms, these machineries are poorly understood in Gram-positive bacteria. Within the Firmicutes phylum, the Enterococcus spp. genus have recently assumed importance in clinical microbiology being considered as emerging pathogens for humans, wherein Enterococcus faecalis represents the major species associated with nosocomial infections. The aim of this study was to carry out a phylogenetic analysis in Enterococcus faecalis V583 and a structural and conformational characterisation of it SufU protein. Results BLAST searches of the Enterococcus genome revealed a series of genes with sequence similarity to the Escherichia coli SUF machinery of [Fe-S] cluster biosynthesis, namely sufB, sufC, sufD and SufS. In addition, the E. coli IscU ortholog SufU was found to be the scaffold protein of Enterococcus spp., containing all features considered essential for its biological activity, including conserved amino acid residues involved in substrate and/or co-factor binding (Cys50,76,138 and Asp52 and, phylogenetic analyses showed a close relationship with orthologues from other Gram-positive bacteria. Molecular dynamics for structural determinations and molecular modeling using E. faecalis SufU primary sequence protein over the PDB:1su0 crystallographic model from Streptococcus pyogenes were carried out with a subsequent 50 ns molecular dynamic trajectory. This presented a stable model, showing secondary structure modifications near the active site and conserved cysteine residues. Molecular modeling using Haemophilus influenzae IscU primary sequence over the PDB:1su0 crystal followed by a MD

Dimerization in the Grb7 Protein

OpenAIRE

Peterson, Tabitha A.; Benallie, Renee L.; Bradford, Andrew M.; Pias, Sally C.; Yazzie, Jaron.; Lor, Siamee N.; Haulsee, Zachary M.; Park, Chad K.; Johnson, Dennis L.; Rohrschneider, Larry R.; Spuches, Anne.; Lyons, Barbara A.

2012-01-01

In previous studies, we showed that the tyrosine phosphorylation state of growth factor receptor–bound protein 7 (Grb7) affects its ability to bind to the transcription regulator FHL2 and the cortactin-interacting protein, human HS-1-associated protein-1. Here, we present results describing the importance of dimerization in the Grb7–Src homology 2 (SH2) domain in terms of its structural integrity and the ability to bind phosphorylated tyrosine peptide ligands. A tyrosine phosphorylation-mimic...

7 CFR 51.2832 - U.S. Commercial.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Commercial. 51.2832 Section 51.2832 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing.... Commercial. U.S. Commercial consists of onions which meet the following requirements: (a) Basic requirements...

Regulator of G Protein Signaling 7 (RGS7) Can Exist in a Homo-oligomeric Form That Is Regulated by Gαo and R7-binding Protein.

Science.gov (United States)

Tayou, Junior; Wang, Qiang; Jang, Geeng-Fu; Pronin, Alexey N; Orlandi, Cesare; Martemyanov, Kirill A; Crabb, John W; Slepak, Vladlen Z

2016-04-22

RGS (regulator of G protein signaling) proteins of the R7 subfamily (RGS6, -7, -9, and -11) are highly expressed in neurons where they regulate many physiological processes. R7 RGS proteins contain several distinct domains and form obligatory dimers with the atypical Gβ subunit, Gβ5 They also interact with other proteins such as R7-binding protein, R9-anchoring protein, and the orphan receptors GPR158 and GPR179. These interactions facilitate plasma membrane targeting and stability of R7 proteins and modulate their activity. Here, we investigated RGS7 complexes using in situ chemical cross-linking. We found that in mouse brain and transfected cells cross-linking causes formation of distinct RGS7 complexes. One of the products had the apparent molecular mass of ∼150 kDa on SDS-PAGE and did not contain Gβ5 Mass spectrometry analysis showed no other proteins to be present within the 150-kDa complex in the amount close to stoichiometric with RGS7. This finding suggested that RGS7 could form a homo-oligomer. Indeed, co-immunoprecipitation of differentially tagged RGS7 constructs, with or without chemical cross-linking, demonstrated RGS7 self-association. RGS7-RGS7 interaction required the DEP domain but not the RGS and DHEX domains or the Gβ5 subunit. Using transfected cells and knock-out mice, we demonstrated that R7-binding protein had a strong inhibitory effect on homo-oligomerization of RGS7. In contrast, our data indicated that GPR158 could bind to the RGS7 homo-oligomer without causing its dissociation. Co-expression of constitutively active Gαo prevented the RGS7-RGS7 interaction. These results reveal the existence of RGS protein homo-oligomers and show regulation of their assembly by R7 RGS-binding partners. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

7 CFR 51.1141 - U.S. Fancy.

Science.gov (United States)

2010-01-01

... tolerances see § 51.1151. (f) Internal quality: Lots meeting the internal requirements for “U.S. Grade AA... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Fancy. 51.1141 Section 51.1141 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

7 CFR 51.2731 - U.S. Spanish Splits.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Spanish Splits. 51.2731 Section 51.2731... STANDARDS) United States Standards for Grades of Shelled Spanish Type Peanuts Grades § 51.2731 U.S. Spanish Splits. “U.S. Spanish Splits” consists of shelled Spanish type peanut kernels which are split or broken...

MMP-9, uPA and uPAR proteins expression and its prognostic significance in esophageal squamous cell carcinoma treated by radiotherapy

International Nuclear Information System (INIS)

Zhu Shuchai; Wang Yafei; Su Jingwei; Wang Yuxiang; Shen Wenbin; Li Juan

2008-01-01

Objective: To explore the the prognostic significance of MMP-9, uPA and uPAR protein expression and its relationship with clinical-pathologic factors in esophageal squamous cell carcinoma treated by radiotherapy. Methods: MMP-9, uPA and uPAR protein expression was measured in 59 esophageal carcinomas and 41 peri-carcinoma tissues with immunohistochemistry. The relationship between the protein expression and the clinical-pathological parameters was analyzed, and the prognostic factors in esophageal squamous cell carcinoma treated by radiotherapy alone was evaluated. Results: The rates of positive expression of MMP-9, uPA and uPAR were 85%, 76% and 78% in esophageal carcinoma and 39%, 49% and 44% in peri-carcinoma tissues (χ 2 =22.54, 8.04 and 12.18; P=0.000,0.005 and 0.000). The rates of positive expression of MMP-9 was 79% and 100% when the depth of tumor invasion was ≤2 cm and >2 cm(P= 0.048), respectively. The expression of uPA was significantly correlated with the status of fat interspace between the esophageal lesion and the vertebra in CT scanning image. When the fat interspace existed and disappeared, the rates of strong positive expression was 44% and 70%, respectively (χ 2 =4.21, P=0.040). The positive expression rate of uPA was significantly correlated with distant metastasis, which was 100% in patients with distant metastasis and 68.89% in those without distant metastasis(χ 2 =4.12, P=0.042). The positive expression rate of MMP-9, uPA and uPAR did not affect the prognosis and the short-term result of esophageal carcinoma treated by radiotherapy alone. Conclusions: The protein expression of MMP-9, uPA and uPAR may correlate with local infiltration and distant metastasis in esophageal squamous cell carcinoma. Protein expression may not influence the prognosis of esophageal carcinoma treated by radio therapy, though long time followed-up is still needed. (authors)

Mass Spectrometry of Intact Proteins Reveals +98 u Chemical Artifacts Following Precipitation in Acetone.

Science.gov (United States)

Güray, Melda Z; Zheng, Shi; Doucette, Alan A

2017-02-03

Protein precipitation in acetone is frequently employed ahead of mass spectrometry for sample preconcentration and purification. Unfortunately, acetone is not chemically inert; mass artifacts have previously been observed on glycine-containing peptides when exposed to acetone under acidic conditions. We herein report a distinct chemical modification occurring at the level of intact proteins when incubated in acetone. This artifact manifests as one or more satellite peaks in the MS spectrum of intact protein, spaced 98 u above the mass of the unmodified protein. Other artifacts (+84, +112 u) also appear upon incubation of proteins or peptides in acetone. The reaction is pH-sensitive, being suppressed when proteins are exposed to acetone under acidic conditions. The +98 u artifact is speculated to originate through an intermediate product of aldol condensation of acetone to form diacetone alcohol and mesityl oxide. A +98 u product could originate from nucleophilic attack on mesityl oxide or through condensation with diacetone alcohol. Given the extent of modification possible upon exposure of proteins to acetone, particularly following overnight solvent exposure or incubation at room temperature, an awareness of the variables influencing this novel modification is valued by proteomics researchers who employ acetone precipitation for protein purification.

The Ribosomal Protein uL22 Modulates the Shape of the Protein Exit Tunnel

DEFF Research Database (Denmark)

Wekselman, Itai; Zimmerman, Ella; Davidovich, Chen

2017-01-01

Erythromycin is a clinically useful antibiotic that binds to an rRNA pocket in the ribosomal exit tunnel. Commonly, resistance to erythromycin is acquired by alterations of rRNA nucleotides that interact with the drug. Mutations in the β hairpin of ribosomal protein uL22, which is rather distal...... of the β hairpin of the mutated uL22 toward the interior of the exit tunnel, triggering a cascade of structural alterations of rRNA nucleotides that propagate to the erythromycin binding pocket. Our findings support recent studies showing that the interactions between uL22 and specific sequences within...

7 CFR 51.1435 - U.S. Commercial Pieces.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Commercial Pieces. 51.1435 Section 51.1435 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards... STANDARDS) United States Standards for Grades of Shelled Pecans Grades § 51.1435 U.S. Commercial Pieces. The...

7 CFR 51.1433 - U.S. Commercial Halves.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Commercial Halves. 51.1433 Section 51.1433 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards... STANDARDS) United States Standards for Grades of Shelled Pecans Grades § 51.1433 U.S. Commercial Halves. The...

7 CFR 51.2833 - U.S. No. 1 Boilers.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 1 Boilers. 51.2833 Section 51.2833 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards...) Grades § 51.2833 U.S. No. 1 Boilers. U.S. No. 1 Boilers consists of onions which meet all the...

7 CFR 51.1148 - U.S. No. 2.

Science.gov (United States)

2010-01-01

... tolerances see § 51.1151. (e) Internal quality: Lots meeting the internal requirements for “U.S. Grade AA... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 2. 51.1148 Section 51.1148 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

snRNP proteins in health and disease

Czech Academy of Sciences Publication Activity Database

Krausová, Michaela; Staněk, David

S1084-9521, č. 17 (2017), s. 30150-30157 ISSN 1084-9521 R&D Projects: GA ČR GA15-00790S; GA MŠk LO1419 Institutional support: RVO:68378050 Keywords : Congenital craniofacial disorders * Haematological malignancies * Mutations * Retinopathy * Spliceosome Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Biochemistry and molecular biology Impact factor: 6.614, year: 2016

7 CFR 51.1142 - U.S. No. 1 Bright.

Science.gov (United States)

2010-01-01

...) Internal quality: Lots meeting the internal requirements for “U.S. Grade AA Juice (Double A)” or “U.S... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 1 Bright. 51.1142 Section 51.1142 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

7 CFR 51.1145 - U.S. No. 1 Bronze.

Science.gov (United States)

2010-01-01

...) Internal quality: Lots meeting the internal requirements for “U.S. Grade AA Juice (Double A)” or “U.S... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 1 Bronze. 51.1145 Section 51.1145 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

7 CFR 51.1147 - U.S. No. 2 Bright.

Science.gov (United States)

2010-01-01

...) Internal quality: Lots meeting the internal requirements for “U.S. Grade AA Juice (Double A)” or “U.S... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 2 Bright. 51.1147 Section 51.1147 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

Wongabel Rhabdovirus Accessory Protein U3 Targets the SWI/SNF Chromatin Remodeling Complex

Science.gov (United States)

Joubert, D. Albert; Rodriguez-Andres, Julio; Monaghan, Paul; Cummins, Michelle; McKinstry, William J.; Paradkar, Prasad N.; Moseley, Gregory W.

2014-01-01

ABSTRACT Wongabel virus (WONV) is an arthropod-borne rhabdovirus that infects birds. It is one of the growing array of rhabdoviruses with complex genomes that encode multiple accessory proteins of unknown function. In addition to the five canonical rhabdovirus structural protein genes (N, P, M, G, and L), the 13.2-kb negative-sense single-stranded RNA (ssRNA) WONV genome contains five uncharacterized accessory genes, one overlapping the N gene (Nx or U4), three located between the P and M genes (U1 to U3), and a fifth one overlapping the G gene (Gx or U5). Here we show that WONV U3 is expressed during infection in insect and mammalian cells and is required for efficient viral replication. A yeast two-hybrid screen against a mosquito cell cDNA library identified that WONV U3 interacts with the 83-amino-acid (aa) C-terminal domain of SNF5, a component of the SWI/SNF chromatin remodeling complex. The interaction was confirmed by affinity chromatography, and nuclear colocalization was established by confocal microscopy. Gene expression studies showed that SNF5 transcripts are upregulated during infection of mosquito cells with WONV, as well as West Nile virus (Flaviviridae) and bovine ephemeral fever virus (Rhabdoviridae), and that SNF5 knockdown results in increased WONV replication. WONV U3 also inhibits SNF5-regulated expression of the cytokine gene CSF1. The data suggest that WONV U3 targets the SWI/SNF complex to block the host response to infection. IMPORTANCE The rhabdoviruses comprise a large family of RNA viruses infecting plants, vertebrates, and invertebrates. In addition to the major structural proteins (N, P, M, G, and L), many rhabdoviruses encode a diverse array of accessory proteins of largely unknown function. Understanding the role of these proteins may reveal much about host-pathogen interactions in infected cells. Here we examine accessory protein U3 of Wongabel virus, an arthropod-borne rhabdovirus that infects birds. We show that U3 enters the

7 CFR 51.2732 - U.S. No. 2 Spanish.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 2 Spanish. 51.2732 Section 51.2732... STANDARDS) United States Standards for Grades of Shelled Spanish Type Peanuts Grades § 51.2732 U.S. No. 2 Spanish. “U.S. No. 2 Spanish” consists of shelled Spanish type peanut kernels which may be split or broken...

7 CFR 51.2730 - U.S. No. 1 Spanish.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 1 Spanish. 51.2730 Section 51.2730... STANDARDS) United States Standards for Grades of Shelled Spanish Type Peanuts Grades § 51.2730 U.S. No. 1 Spanish. “U.S. No. 1 Spanish” consists of shelled Spanish type peanut kernels which are whole and free...

AR-v7 protein expression is regulated by protein kinase and phosphatase

Science.gov (United States)

Li, Yinan; Xie, Ning; Gleave, Martin E.; Rennie, Paul S.; Dong, Xuesen

2015-01-01

Failure of androgen-targeted therapy and progression of castration-resistant prostate cancer (CRPC) are often attributed to sustained expression of the androgen receptor (AR) and its major splice variant, AR-v7. Although the new generation of anti-androgens such as enzalutamide effectively inhibits AR activity, accumulating pre-clinical and clinical evidence indicates that AR-v7 remains constitutively active in driving CRPC progression. However, molecular mechanisms which control AR-v7 protein expression remain unclear. We apply multiple prostate cancer cell models to demonstrate that enzalutamide induces differential activation of protein phosphatase-1 (PP-1) and Akt kinase depending on the gene context of cancer cells. The balance between PP-1 and Akt activation governs AR phosphorylation status and activation of the Mdm2 ubiquitin ligase. Mdm2 recognizes phosphorylated serine 213 of AR-v7, and induces AR-v7 ubiquitination and protein degradation. These findings highlight the decisive roles of PP-1 and Akt for AR-v7 protein expression and activities when AR is functionally blocked. PMID:26378044

Decomposition of the metastable phase γU in U-7% and U-7% Mo-0.9% Pt

International Nuclear Information System (INIS)

Arico, Sergio F.; Gribaudo, Luis M.

2004-01-01

The 'Reduced Enrichment for Research and Test Reactors' is an international project for the development of a nuclear fuel with high density in uranium capable to get a great neutron flux with good capacity for being reprocessed. One of the candidates is a fuel containing U-Mo alloy powder, as bcc metastable phase γ, dispersed in Al powder. In order to know the influence of Pt as a stabilizing element two U-7 wt.% Mo alloys are studied, one of them with 0.9 wt.% Pt. They were fabricated in an arc furnace and both homogenized in composition during 2 h at 1000 C degrees. Then, isothermal treatments at 480, 430 and 350 C degrees were performed at times between 1 and 177 h. The decomposition of the γ phase was studied by metallography and X-ray diffraction analysis. Adding Pt, the start of the decomposition of the γ phase is delayed, but the initial grain size of the alloys is an important variable which has also to be considered. (author) [es

Analysis of protein folds using protein contact networks

Indian Academy of Sciences (India)

is a well-recognized classification system of proteins, which is based on manual in- ... can easily correspond to the information in the 2D matrix. ..... [7] U K Muppirala and Zhijun Li, Protein Engineering, Design & Selection 19, 265 (2006).

In-pile test results of U-silicide or U-nitride coated U-7Mo particle dispersion fuel in Al

Science.gov (United States)

Kim, Yeon Soo; Park, J. M.; Lee, K. H.; Yoo, B. O.; Ryu, H. J.; Ye, B.

2014-11-01

U-silicide or U-nitride coated U-Mo particle dispersion fuel in Al (U-Mo/Al) was in-pile tested to examine the effectiveness of the coating as a diffusion barrier between the U-7Mo fuel kernels and Al matrix. This paper reports the PIE data and analyses focusing on the effectiveness of the coating in terms of interaction layer (IL) growth and general fuel performance. The U-silicide coating showed considerable success, but it also provided evidence for additional improvement for coating process. The U-nitride coated specimen showed largely inefficient results in reducing IL growth. From the test, important observations were also made that can be utilized to improve U-Mo/Al fuel performance. The heating process for coating turned out to be beneficial to suppress fuel swelling. The use of larger fuel particles confirmed favorable effects on fuel performance.

In-pile test results of U-silicide or U-nitride coated U-7Mo particle dispersion fuel in Al

Energy Technology Data Exchange (ETDEWEB)

Kim, Yeon Soo, E-mail: yskim@anl.gov [Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States); Park, J.M.; Lee, K.H.; Yoo, B.O. [Korea Atomic Energy Research Institute, 989-111 Daedeokdaero, Yuseong-gu, Daejeon 305-353 (Korea, Republic of); Ryu, H.J. [Dept. of Nuclear and Quantum Engineering, Korea Advanced Institute of Science and Technology, 291 Daehak-ro, Yuseong-gu, Daejeon 305-701 (Korea, Republic of); Ye, B. [Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States)

2014-11-15

U-silicide or U-nitride coated U-Mo particle dispersion fuel in Al (U-Mo/Al) was in-pile tested to examine the effectiveness of the coating as a diffusion barrier between the U-7Mo fuel kernels and Al matrix. This paper reports the PIE data and analyses focusing on the effectiveness of the coating in terms of interaction layer (IL) growth and general fuel performance. The U-silicide coating showed considerable success, but it also provided evidence for additional improvement for coating process. The U-nitride coated specimen showed largely inefficient results in reducing IL growth. From the test, important observations were also made that can be utilized to improve U-Mo/Al fuel performance. The heating process for coating turned out to be beneficial to suppress fuel swelling. The use of larger fuel particles confirmed favorable effects on fuel performance.

Mutations in spliceosomal proteins and retina degeneration

Czech Academy of Sciences Publication Activity Database

Růžičková, Šárka; Staněk, David

2017-01-01

Roč. 14, č. 5 (2017), s. 544-552 ISSN 1547-6286 R&D Projects: GA MŠk LO1419 Institutional support: RVO:68378050 Keywords : Retinitis pigmentosa * snRNP * splicing Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Genetics and heredity (medical genetics to be 3) Impact factor: 3.900, year: 2016

HT-7U superconducting tokamak: Physics design, engineering progress and schedule

International Nuclear Information System (INIS)

Wan Yuanxi

2002-01-01

The superconducting tokamak research program begun in China in ASIPP since 1994. The program is included in existent superconducting tokamak HT-7 and the next new superconducting tokamak HT-7U which is one of national key research projects in China. With the elongation cross-section, divertor and higher plasma parameter the main objectives of HT-7U are widely investigation both of the physics and technology for steady state advanced tokamak as well as the investigation of power and particle handle under steady-state operation condition. The physics and engineering design have been completed and significant progresses on R and D and fabrication have been achieved. HT-7U will begin assembly at 2003 and possible to get first plasma around 2004. (author)

7 CFR 51.1146 - U.S. No. 1 Russet.

Science.gov (United States)

2010-01-01

.... (a) For tolerances see § 51.1151. (b) Internal quality: Lots meeting the internal requirements for “U... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 1 Russet. 51.1146 Section 51.1146 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

7 CFR 51.1144 - U.S. No. 1 Golden.

Science.gov (United States)

2010-01-01

...) For tolerances see § 51.1151. (b) Internal quality: Lots meeting the internal requirements for “U.S... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 1 Golden. 51.1144 Section 51.1144 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

7 CFR 51.1149 - U.S. No. 2 Russet.

Science.gov (United States)

2010-01-01

... tolerances see § 51.1151. (b) Internal quality: Lots meeting the internal requirements for “U.S. Grade AA... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 2 Russet. 51.1149 Section 51.1149 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

Associations of Metabolic Syndrome, Elevated C-Reactive Protein, and Physical Activity in U.S. Adolescents.

Science.gov (United States)

Williams, Bethany D; Richardson, Michael R; Johnson, Tammie M; Churilla, James R

2017-12-01

The aim was to estimate the prevalence of metabolic syndrome (MetS) criteria, elevated C-reactive protein (CRP), and physical activity (PA) as well as the odds of MetS criteria in those active versus inactive utilizing a representative sample of U.S. adolescents. The study sample (n = 676) included male and female adolescent (12-17 years) participants in the 2007-2010 National Health and Nutrition Examination Survey. The criteria analyzed were based on a modified definition of MetS using the Third Report of the National Cholesterol Education Program Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults. Current adult cut points were used to determine elevated CRP. Activity was estimated using reported days per week and minutes per day of moderate/vigorous PA. The MetS criteria with the highest and lowest overall prevalence estimates were elevated fasting glucose and elevated blood pressure (20.7% [95% confidence interval, 17.02-24.38] and 5.7% [95% confidence interval, 3.70-7.70], respectively). The prevalence of elevated CRP was 7.1% (6.3% and 7.8% in males and females, respectively; p = .42). The prevalence of insufficient PA was 75.0%. Odds of low high-density lipoprotein cholesterol were significantly lower in active adolescents when compared with inactive adolescents (odds ratio = .39, p < .05). In a representative sample of U.S. adolescents, elevated fasting glucose is the most prevalent MetS criterion. One out of five U.S. adolescents has elevated fasting glucose, and three out four do not meet the daily federal PA recommendations. Adolescents meeting the federal PA recommendation demonstrate approximately 60% lower odds of having low high-density lipoprotein cholesterol. Copyright © 2017 Society for Adolescent Health and Medicine. Published by Elsevier Inc. All rights reserved.

TEM Characterization of High Burn-up Microstructure of U-7Mo Alloy

Energy Technology Data Exchange (ETDEWEB)

Jian Gan; Brandon Miller; Dennis Keiser; Adam Robinson; James Madden; Pavel Medvedev; Daniel Wachs

2014-04-01

As an essential part of global nuclear non-proliferation effort, the RERTR program is developing low enriched U-Mo fuels (< 20% U-235) for use in research and test reactors that currently employ highly enriched uranium fuels. One type of fuel being developed is a dispersion fuel plate comprised of U-7Mo particles dispersed in Al alloy matrix. Recent TEM characterizations of the ATR irradiated U-7Mo dispersion fuel plates include the samples with a local fission densities of 4.5, 5.2, 5.6 and 6.3 E+21 fissions/cm3 and irradiation temperatures of 101-136?C. The development of the irradiated microstructure of the U-7Mo fuel particles consists of fission gas bubble superlattice, large gas bubbles, solid fission product precipitates and their association to the large gas bubbles, grain subdivision to tens or hundreds of nanometer size, collapse of bubble superlattice, and amorphisation. This presentation will describe the observed microstructures specifically focusing on the U-7Mo fuel particles. The impact of the observed microstructure on the fuel performance and the comparison of the relevant features with that of the high burn-up UO2 fuels will be discussed.

Thermophysical properties of heat-treated U-7Mo/Al dispersion fuel

Energy Technology Data Exchange (ETDEWEB)

Cho, Tae Won; Kim, Yeon Soo; Park, Jong Man; Lee, Kyu Hong; Kim, Sunghwan; Lee, Chong Tak; Yang, Jae Ho; Oh, Jang Soo; Sohn, Dong-Seong

2018-04-01

In this study, the effects of interaction layer (IL) on thermophysical properties of U-7Mo/Al dispersion fuel were examined. Microstructural analyses revealed that ILs were formed uniformly on U-Mo particles during heating of U-7Mo/Al samples. The IL volume fraction was measured by applying image analysis methods. The uranium loadings of the samples were calculated based on the measured meat densities at 298 K. The density of the IL was estimated by using the measured density and IL volume fraction. Thermal diffusivity and heat capacity of the samples after the heat treatment were measured as a function of temperature and volume fractions of U-Mo and IL. The thermal conductivity of IL-formed U-7Mo/Al was derived by using the measured thermal diffusivity, heat capacity, and density. The thermal conductivity obtained in the present study was lower than that predicted by the modified Hashin–Shtrikman model due to the theoretical model’s inability to consider the thermal resistance at interfaces between the meat constituents.

Thermophysical properties of heat-treated U-7Mo/Al dispersion fuel

Science.gov (United States)

Cho, Tae Won; Kim, Yeon Soo; Park, Jong Man; Lee, Kyu Hong; Kim, Sunghwan; Lee, Chong Tak; Yang, Jae Ho; Oh, Jang Soo; Sohn, Dong-Seong

2018-04-01

In this study, the effects of interaction layer (IL) on thermophysical properties of U-7Mo/Al dispersion fuel were examined. Microstructural analyses revealed that ILs were formed uniformly on U-Mo particles during heating of U-7Mo/Al samples. The IL volume fraction was measured by applying image analysis methods. The uranium loadings of the samples were calculated based on the measured meat densities at 298 K. The density of the IL was estimated by using the measured density and IL volume fraction. Thermal diffusivity and heat capacity of the samples after the heat treatment were measured as a function of temperature and volume fractions of U-Mo and IL. The thermal conductivity of IL-formed U-7Mo/Al was derived by using the measured thermal diffusivity, heat capacity, and density. The thermal conductivity obtained in the present study was lower than that predicted by the modified Hashin-Shtrikman model due to the theoretical model's inability to consider the thermal resistance at interfaces between the meat constituents.

Promoter proximal polyadenylation sites reduce transcription activity

DEFF Research Database (Denmark)

Andersen, Pia Kjølhede; Lykke-Andersen, Søren; Jensen, Torben Heick

2012-01-01

Gene expression relies on the functional communication between mRNA processing and transcription. We previously described the negative impact of a point-mutated splice donor (SD) site on transcription. Here we demonstrate that this mutation activates an upstream cryptic polyadenylation (CpA) site......, which in turn causes reduced transcription. Functional depletion of U1 snRNP in the context of the wild-type SD triggers the same CpA event accompanied by decreased RNA levels. Thus, in accordance with recent findings, U1 snRNP can shield premature pA sites. The negative impact of unshielded pA sites...... on transcription requires promoter proximity, as demonstrated using artificial constructs and supported by a genome-wide data set. Importantly, transcription down-regulation can be recapitulated in a gene context devoid of splice sites by placing a functional bona fide pA site/transcription terminator within ∼500...

Reaction layer in U-7WT%MO/Al diffusion couples

International Nuclear Information System (INIS)

Mirandou, M.I.; Balart, S.N.; Ortiz, M.; Granovsky, M.S.

2003-01-01

New results of the reaction layer characterization between γ (U-7wt%Mo) alloy and Al, in chemical diffusion couples, are presented. The analysis was performed using optical and scanning electron microscopy with EDAX and X-ray diffraction techniques. Besides the main components (U, Mo)Al 3 and (U, Mo)Al 4 , already reported, two ternary compounds of high Al content have been identified in the reaction layer when it grew in retained or decomposed γ (U, Mo) phase, respectively. The drastic consequence on the interdiffusion behavior due to the thermal instability of the retained γ (U, Mo) phase is discussed. (author)

A large complement of the predicted Arabidopsis ARM repeat proteins are members of the U-box E3 ubiquitin ligase family.

Science.gov (United States)

Mudgil, Yashwanti; Shiu, Shin-Han; Stone, Sophia L; Salt, Jennifer N; Goring, Daphne R

2004-01-01

The Arabidopsis genome was searched to identify predicted proteins containing armadillo (ARM) repeats, a motif known to mediate protein-protein interactions in a number of different animal proteins. Using domain database predictions and models generated in this study, 108 Arabidopsis proteins were identified that contained a minimum of two ARM repeats with the majority of proteins containing four to eight ARM repeats. Clustering analysis showed that the 108 predicted Arabidopsis ARM repeat proteins could be divided into multiple groups with wide differences in their domain compositions and organizations. Interestingly, 41 of the 108 Arabidopsis ARM repeat proteins contained a U-box, a motif present in a family of E3 ligases, and these proteins represented the largest class of Arabidopsis ARM repeat proteins. In 14 of these U-box/ARM repeat proteins, there was also a novel conserved domain identified in the N-terminal region. Based on the phylogenetic tree, representative U-box/ARM repeat proteins were selected for further study. RNA-blot analyses revealed that these U-box/ARM proteins are expressed in a variety of tissues in Arabidopsis. In addition, the selected U-box/ARM proteins were found to be functional E3 ubiquitin ligases. Thus, these U-box/ARM proteins represent a new family of E3 ligases in Arabidopsis.

Design of the Cryostat for HT-7U Superconducting Tokamak

Science.gov (United States)

Yu, Jie; Wu, Song-tao; Song, Yun-tao; Weng, Pei-de

2002-06-01

The cryostat of HT-7U tokamak is a large vacuum vessel surrounding the entire basic machine with a cylindrical shell, a dished top and a flat bottom. The main function of HT-7U cryostat is to provide a thermal barrier between an ambient temperature test hall and a liquid helium-cooled superconducting magnet. The loads applied to the cryostat are from sources of vacuum pressure, dead weight, seismic events and electromagnetic forces originated by eddy currents. It also provides feed-through penetrations for all the connecting elements inside and outside the cryostat. The main material selected for the cryostat is stainless steel 304L. The structural analyses including buckling for the cryostat vessel under the plasma operation condition have been carried out by using a finite element code. Stress analysis results show that the maximum stress intensity was below the allowable value. In this paper, the structural analyses and design of HT-7U cryostat are emphasized.

The ribosomal protein uL22 modulates the shape of the nascent protein exit tunnel

DEFF Research Database (Denmark)

Wekselman, I.; Zimmerman, E.; Davidovich, C.

2017-01-01

in the entrance of theribosomal exit tunnel and interferes with the progression of nas-cent chains. Commonly, resistance to erythromycin is acquiredby alterations of rRNA nucleotides that interact with the drug.Mutations in theb-hairpin of ribosomal protein uL22, which israther distal to the erythromycin binding...... to erythromycin binding pocket and increases its flexi-bility. Based on our results, we suggest a feasble mechanism thatexplains how nanscent proteins can be translated when ery-thromycin is bound to the ribosome. Furthermore, our findingssupport recent studies showing that the interactions betweenuL22...

[Cloning, expression and transcriptional analysis of biotin carboxyl carrier protein gene (accA) from Amycolatopsis mediterranei U32 ].

Science.gov (United States)

Lu, Jie; Yao, Yufeng; Jiang, Weihong; Jiao, Ruishen

2003-02-01

Acetyl CoA carboxylase (EC 6.4.1.2, ACC) catalyzes the ATP-dependent carboxylation of acetyl CoA to yield malonyl CoA, which is the first committed step in fatty acid synthesis. A pair of degenerate PCR primers were designed according to the conserved amino acid sequence of AccA from M. tuberculosis and S. coelicolor. The product of the PCR amplification, a DNA fragment of 250bp was used as a probe for screening the U32 genomic cosmid library and its gene, accA, coding the biotinylated protein subunit of acetyl CoA carboxylase, was successfully cloned from U32. The accA ORF encodes a 598-amino-acid protein with the calculated molecular mass of 63.7kD, with 70.1% of G + C content. A typical Streptomyces RBS sequence, AGGAGG, was found at the - 6 position upstream of the start codon GTG. Analysis of the deduced amino acid sequence showed the presence of biotin-binding site and putative ATP-bicarbonate interaction region, which suggested the U32 AccA may act as a biotin carboxylase as well as a biotin carrier protein. Gene accA was then cloned into the pET28 (b) vector and expressed solubly in E. coli BL21 (DE3) by 0.1 mmol/L IPTG induction. Western blot confirmed the covalent binding of biotin with AccA. Northern blot analyzed transcriptional regulation of accA by 5 different nitrogen sources.

Phase development in a U-7 wt.% Mo vs. Al-7 wt.% Ge diffusion couple

Science.gov (United States)

Perez, E.; Keiser, D. D.; Sohn, Y. H.

2013-10-01

Fuel development for the Reduced Enrichment for Research and Test Reactors (RERTR) program has demonstrated that U-Mo alloys in contact with Al develop interaction regions with phases that have poor irradiation behavior. The addition of Si to the Al has been considered with positive results. In this study, compositional modification is considered by replacing Si with Ge to determine the effect on the phase development in the system. The microstructural and phase development of a diffusion couple of U-7 wt.% Mo in contact with Al-7 wt.% Ge was examined by transmission electron microscopy, scanning electron microscopy and energy dispersive spectroscopy. The interdiffusion zone developed a microstructure that included the cubic-UGe3 phase and amorphous phases. The UGe3 phase was observed with and without Mo and Al solid solution developing a (U,Mo)(Al,Ge)3 phase.

Aedes aegypti D7 Saliva Protein Inhibits Dengue Virus Infection.

Directory of Open Access Journals (Sweden)

Michael J Conway

2016-09-01

Full Text Available Aedes aegypti is the primary vector of several medically relevant arboviruses including dengue virus (DENV types 1-4. Ae. aegypti transmits DENV by inoculating virus-infected saliva into host skin during probing and feeding. Ae. aegypti saliva contains over one hundred unique proteins and these proteins have diverse functions, including facilitating blood feeding. Previously, we showed that Ae. aegypti salivary gland extracts (SGEs enhanced dissemination of DENV to draining lymph nodes. In contrast, HPLC-fractionation revealed that some SGE components inhibited infection. Here, we show that D7 proteins are enriched in HPLC fractions that are inhibitory to DENV infection, and that recombinant D7 protein can inhibit DENV infection in vitro and in vivo. Further, binding assays indicate that D7 protein can directly interact with DENV virions and recombinant DENV envelope protein. These data reveal a novel role for D7 proteins, which inhibits arbovirus transmission to vertebrates through a direct interaction with virions.

SEM and TEM Characterization of As-Fabricated U-7Mo Disperson Fuel Plates

International Nuclear Information System (INIS)

Keiser, D.D. Jr.; Yao, B.; Perez, E.; Sohn, Y.H.

2009-01-01

The starting microstructure of a dispersion fuel plate can have a dramatic impact on the overall performance of the plate during irradiation. To improve the understanding of the as-fabricated microstructures of dispersion fuel plates, SEM and TEM analysis have been performed on RERTR-9A archive fuel plates, which went through an additional hot isostatic procsssing (HIP) step during fabrication. The fuel plates had depleted U-7Mo fuel particles dispersed in either Al-2Si or 4043 Al alloy matrix. For the characterized samples, it was observed that a large fraction of the ?-phase U-7Mo alloy particles had decomposed during fabrication, and in areas near the fuel/matrix interface where the transformation products were present significant fuel/matrix interaction had occurred. Relatively thin Si-rich interaction layers were also observed around the U-7Mo particles. In the thick interaction layers, (U)(Al,Si)3 and U6Mo4Al43 were identified, and in the thin interaction layers U(Al,Si)3, U3Si3Al2, U3Si5, and USi1.88-type phases were observed. The U3Si3Al2 phase contained some Mo. Based on the results of this work, exposure of dispersion fuel plates to relatively high temperatures during fabrication impacts the overall microstructure, particularly the nature of the interaction layers around the fuel particles. The time and temperature of fabrication should be carefully controlled in order to produce the most uniform Si-rich layers around the U-7Mo particles.

Structure of a rare non-standard sequence k-turn bound by L7Ae protein

Science.gov (United States)

Huang, Lin; Lilley, David M.J.

2014-01-01

Kt-23 from Thelohania solenopsae is a rare RNA kink turn (k-turn) where an adenine replaces the normal guanine at the 2n position. L7Ae is a member of a strongly conserved family of proteins that bind a range of k-turn structures in the ribosome, box C/D and H/ACA small nucleolar RNAs and U4 small nuclear RNA. We have solved the crystal structure of T. solenopsae Kt-23 RNA bound to Archeoglobus fulgidus L7Ae protein at a resolution of 2.95 Å. The protein binds in the major groove displayed on the outer face of the k-turn, in a manner similar to complexes with standard k-turn structures. The k-turn adopts a standard N3 class conformation, with a single hydrogen bond from A2b N6 to A2n N3. This contrasts with the structure of the same sequence located in the SAM-I riboswitch, where it adopts an N1 structure, showing the inherent plasticity of k-turn structure. This potentially can affect any tertiary interactions in which the RNA participates. PMID:24482444

Characterization of the reaction layer in U-7wt%Mo/Al diffusion couples

Energy Technology Data Exchange (ETDEWEB)

Mirandou, M.I.; Balart, S.N.; Ortiz, M.; Granovsky, M.S. E-mail: granovsk@cnea.gov.ar

2003-11-15

The reaction layer in chemical diffusion couples U-7wt%Mo/Al was investigated using optical and scanning electron microscopy, electron probe microanalysis and X-ray diffraction (XRD) techniques. When the U-7wt%Mo alloy was previously homogenized and the {gamma}(U, Mo) phase was retained, the formation of (U, Mo)Al{sub 3} and (U, Mo)Al{sub 4} was observed at 580 deg. C. Also a very thin band was detected close to the Al side, the structure of the ternary compound Al{sub 20}UMo{sub 2} might be assigned to it. When the decomposition of the {gamma}(U, Mo) took place, a drastic change in the diffusion behavior was observed. In this case, XRD indicated the presence of phases with the structures of (U, Mo)Al{sub 3}, Al{sub 43}U{sub 6}Mo{sub 4}, {gamma}(U, Mo) and {alpha}(U) in the reaction layer.

The urokinase receptor associated protein (uPARAP/endo180)

DEFF Research Database (Denmark)

Engelholm, L H; Nielsen, B S; Danø, K

2001-01-01

The urokinase-mediated plasminogen activation system plays a central role in the extracellular proteolytic degradation reactions in cancer invasion. In this review article we discuss a number of recent findings identifying a new cellular receptor protein, uPARAP, that interacts with components of...... and their substrate degradation products and thus may add to the complicated interplay between several cell types in governing restricted tissue degradation....

Economic Optimizing Control for Single-Cell Protein Production in a U-Loop Reactor

DEFF Research Database (Denmark)

Drejer, André; Ritschel, Tobias Kasper Skovborg; Jørgensen, Sten Bay

2017-01-01

The production of single-cell protein (SCP) in a U-loop reactor by a methanotroph is a cost efficient sustainable alternative to protein from fish meal obtained by over-fishing the oceans. SCP serves as animal feed. In this paper, we present a mathematical model that describes the dynamics of SCP...

Oxidation of uraninite: does tetragonal U3O7 occur in nature?

International Nuclear Information System (INIS)

Janeczek, J.; Ewing, R.C.; Thomas, L.E.

1993-01-01

Samples of uraninite and pitchblende annealed at 1200 C in H 2 , and untreated pitchblende were sequentially oxidized in air at 180-190 C, 230 C, and 300 C. Uraninite and untreated pitchblende oxidized to the U 4 O 9 -type oxide, and their X-ray symmetry remained isometric up to 300 C. Reduced pitchblende after oxidation to UO 2+x and U 4 O 9 -type oxides transformed into α-U 3 O 8 at 300 C. Two major mechanisms control uraninite and untreated pitchblende stability during oxidation: (1) Th and/or REE maintain charge balance and block oxygen interstitials near impurity cations; (2) the uraninite structure saturates with respect to excess oxygen and radiation-induced oxygen interstitials. Untreated pitchblende during oxidation behaved similarly to irradiated UO 2 in spent nuclear fuel; whereas, reduced pitchblende resembled nonirradiated UO 2 . An analysis of the data in the literature, as well as our own efforts to identify U 3 O 7 in samples from Cigar Lake, Canada, failed to provide conclusive evidence of the natural occurrence of tetragonal α-U 3 O 7 . Most probably, reported occurrences of U 3 O 7 are mixtures of isometric uraninites of slightly different compositions. (orig.)

Antigenicity analysis of human parvovirus B19-VP1u protein in the induction of anti-phospholipid syndrome.

Science.gov (United States)

Lin, Chun-Yu; Chiu, Chun-Ching; Cheng, Ju; Lin, Chia-Yun; Shi, Ya-Fang; Tsai, Chun-Chou; Tzang, Bor-Show; Hsu, Tsai-Ching

2018-01-01

Mounting evidence suggests a connection between human parvovirus B19 (B19) and autoimmune diseases, and especially an association between the B19-VP1 unique region (VP1u) and anti-phospholipid syndrome (APS). However, little is known about the antigenicity of B19-VP1u in the induction of APS-like syndrome. To elucidate the antigenicity of B19-VP1u in the induction of APS, N-terminal truncated B19-VP1u (tVP1u) proteins were prepared to immunize Balb/c mice to generate antibodies against B19-tVP1u proteins. The secreted phospholipase A2 (sPLA2) activities and binding specificity of mice anti-B19-tVP1u antibodies with cardiolipin (CL) and beta-2-glycoprotein I (β2GPI) were evaluated by performing immunoblot, ELISA and absorption experiments. A mice model of passively induced APS was adopted. Although sPLA2 activities were identified in all B19-tVP1u proteins, only amino acid residues 61-227 B19-tVP1u exhibited a higher sPLA2 activity. Autoantibodies against CL and β2GPI exhibited binding activities with all B19-tVP1u proteins. IgG that was purified from mice that had been immunized with amino acid residues 21-227 to 121-227 B19-tVP1u proteins exhibited significantly higher binding activity with CL. IgG that was purified from mice that had been immunized with amino acid residues 21-227, 31-227, 82-227 and 91-227 B19-tVP1u proteins exhibited significantly higher binding activity with β2GPI. Accordingly, significantly higher binding inhibition of CL was detected in the presence of amino acid residues 61-227 and 101-227 B19-tVP1u. Significantly higher binding inhibition of β2GPI was detected in the presence of amino acid residues 21-227, 31-227, 82-227 and 91-227 B19-tVP1u. The mice that received amino acid residues 31-227 or 61-227 anti-tB19-VP1u IgG revealed significant thrombocytopenia and those that received amino acid residues 21-227, 31-227, 61-227, 71-227, 82-227, 91-227, 101-227 or 114-227 anti-tB19-VP1u IgG exhibited significantly prolonged aPTT. These

HSP60, a protein downregulated by IGFBP7 in colorectal carcinoma

Directory of Open Access Journals (Sweden)

Lin Jie

2010-04-01

Full Text Available Abstract Background In our previous study, it was well defined that IGFBP7 was an important tumor suppressor gene in colorectal cancer (CRC. We aimed to uncover the downstream molecules responsible for IGFBP7's behaviour in this study. Methods Differentially expressed protein profiles between PcDNA3.1(IGFBP7-transfected RKO cells and the empty vector transfected controls were generated by two-dimensional gel electrophoresis (2-DE and mass spectrometry (MS identification. The selected differentially expressed protein induced by IGFBP7 was confirmed by western blot and ELISA. The biological behaviour of the protein was explored by cell growth assay and colony formation assay. Results Six unique proteins were found differentially expressed in PcDNA3.1(IGFBP7-transfected RKO cells, including albumin (ALB, 60 kDa heat shock protein(HSP60, Actin cytoplasmic 1 or 2, pyruvate kinase muscle 2(PKM2, beta subunit of phenylalanyl-tRNA synthetase(FARSB and hypothetical protein. The downregulation of HSP60 by IGFBP7 was confirmed by western blot and ELISA. Recombinant human HSP60 protein could increase the proliferation rate and the colony formation ability of PcDNA3.1(IGFBP7-RKO cells. Conclusion HSP60 was an important downstream molecule of IGFBP7. The downregulation of HSP60 induced by IGFBP7 may be, at least in part, responsible for IGFBP7's tumor suppressive biological behaviour in CRC.

7 CFR 51.1143 - U.S. No. 1.

Science.gov (United States)

2010-01-01

..., (22) Other means. (d) For tolerances see § 51.1151. (e) Internal quality: Lots meeting the internal... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 1. 51.1143 Section 51.1143 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

A Large Complement of the Predicted Arabidopsis ARM Repeat Proteins Are Members of the U-Box E3 Ubiquitin Ligase Family1[w

Science.gov (United States)

Mudgil, Yashwanti; Shiu, Shin-Han; Stone, Sophia L.; Salt, Jennifer N.; Goring, Daphne R.

2004-01-01

The Arabidopsis genome was searched to identify predicted proteins containing armadillo (ARM) repeats, a motif known to mediate protein-protein interactions in a number of different animal proteins. Using domain database predictions and models generated in this study, 108 Arabidopsis proteins were identified that contained a minimum of two ARM repeats with the majority of proteins containing four to eight ARM repeats. Clustering analysis showed that the 108 predicted Arabidopsis ARM repeat proteins could be divided into multiple groups with wide differences in their domain compositions and organizations. Interestingly, 41 of the 108 Arabidopsis ARM repeat proteins contained a U-box, a motif present in a family of E3 ligases, and these proteins represented the largest class of Arabidopsis ARM repeat proteins. In 14 of these U-box/ARM repeat proteins, there was also a novel conserved domain identified in the N-terminal region. Based on the phylogenetic tree, representative U-box/ARM repeat proteins were selected for further study. RNA-blot analyses revealed that these U-box/ARM proteins are expressed in a variety of tissues in Arabidopsis. In addition, the selected U-box/ARM proteins were found to be functional E3 ubiquitin ligases. Thus, these U-box/ARM proteins represent a new family of E3 ligases in Arabidopsis. PMID:14657406

Mitochondrial Band-7 family proteins: scaffolds for respiratory chain assembly?

Directory of Open Access Journals (Sweden)

Bernadette eGehl

2014-04-01

Full Text Available The band-7 protein family comprises a diverse set of membrane-bound proteins characterised by the presence of a conserved domain. The exact function of this band-7 domain remains elusive, but examples from animal and bacterial stomatin-type proteins demonstrate binding to lipids and the ability to assemble into membrane-bound oligomers that form putative scaffolds. Some members, such as prohibitins and human stomatin-like protein 2 (HsSLP2, localise to the mitochondrial inner membrane where they function in cristae formation and hyperfusion. In Arabidopsis, the band-7 protein family has diversified and includes plant-specific members. Mitochondrial-localised members include prohibitins (AtPHBs and two stomatin-like proteins (AtSLP1 and -2. Studies into PHB function in plants have demonstrated an involvement in root meristem proliferation and putative scaffold formation for mAAA proteases, but it remains unknown how these roles are achieved at the molecular level. In this minireview we summarise the current status of band-7 protein functions in Arabidopsis, and speculate how the mitochondrial members might recruit specific lipids to form microdomains that could shape the organisation and functioning of the respiratory chain.

A novel uPAg-KPI fusion protein inhibits the growth and invasion of human ovarian cancer cells in vitro.

Science.gov (United States)

Zhao, Li-Ping; Xu, Tian-Min; Kan, Mu-Jie; Xiao, Ye-Chen; Cui, Man-Hua

2016-05-01

Urokinase-type plasminogen activator (uPA) acts by breaking down the basement membrane and is involved in cell proliferation, migration and invasion. These actions are mediated by binding to the uPA receptor (uPAR) via its growth factor domain (GFD). The present study evaluated the effects of uPAg-KPI, a fusion protein of uPA-GFD and a kunitz protease inhibitor (KPI) domain that is present in the amyloid β-protein precursor. Using SKOV-3 cells, an ovarian cancer cell line, we examined cell viability, migration, invasion and also protein expression. Furthermore, we examined wound healing, and migration and invasion using a Transwell assay. Our data showed that uPAg-KPI treatment reduced the viability of ovarian cancer SKOV-3 cells in both a concentration and time-dependent manner by arresting tumor cells at G1/G0 phase of the cell cycle. The IC50 of uPAg-KPI was 0.5 µg/µl after 48 h treatment. At this concentration, uPAg-KPI also inhibited tumor cell colony formation, wound closure, as well as cell migration and invasion capacity. At the protein level, western blot analysis demonstrated that uPAg-KPI exerted no significant effect on the expression of total extracellular signal-regulated kinase (ERK)1/ERK2 and AKT, whereas it suppressed levels of phosphorylated ERK1/ERK2 and AKT. Thus, we suggest that this novel uPAg-KPI fusion protein reduced cell viability, colony formation, wound healing and the invasive ability of human ovarian cancer SKOV-3 cells in vitro by regulating ERK and AKT signaling. Further studies using other cell lines will confirm these findings.

A role for complexes of survival of motor neurons (SMN) protein with gemins and profilin in neurite-like cytoplasmic extensions of cultured nerve cells

International Nuclear Information System (INIS)

Sharma, Aarti; Lambrechts, Anja; Le thi Hao; Le, Thanh T.; Sewry, Caroline A.; Ampe, Christophe; Burghes, Arthur H.M.; Morris, Glenn E.

2005-01-01

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 β-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

C to U RNA editing mediated by APOBEC1 requires RNA-binding protein RBM47.

Science.gov (United States)

Fossat, Nicolas; Tourle, Karin; Radziewic, Tania; Barratt, Kristen; Liebhold, Doreen; Studdert, Joshua B; Power, Melinda; Jones, Vanessa; Loebel, David A F; Tam, Patrick P L

2014-08-01

Cytidine (C) to Uridine (U) RNA editing is a post-transcriptional modification that is accomplished by the deaminase APOBEC1 and its partnership with the RNA-binding protein A1CF. We identify and characterise here a novel RNA-binding protein, RBM47, that interacts with APOBEC1 and A1CF and is expressed in tissues where C to U RNA editing occurs. RBM47 can substitute for A1CF and is necessary and sufficient for APOBEC1-mediated editing in vitro. Editing is further impaired in Rbm47-deficient mutant mice. These findings suggest that RBM47 and APOBEC1 constitute the basic machinery for C to U RNA editing. © 2014 The Authors.

Controlling cellular P-TEFb activity by the HIV-1 transcriptional transactivator Tat.

Directory of Open Access Journals (Sweden)

Lisa Muniz

Full Text Available The human immunodeficiency virus 1 (HIV-1 transcriptional transactivator (Tat is essential for synthesis of full-length transcripts from the integrated viral genome by RNA polymerase II (Pol II. Tat recruits the host positive transcription elongation factor b (P-TEFb to the HIV-1 promoter through binding to the transactivator RNA (TAR at the 5'-end of the nascent HIV transcript. P-TEFb is a general Pol II transcription factor; its cellular activity is controlled by the 7SK small nuclear RNA (snRNA and the HEXIM1 protein, which sequester P-TEFb into transcriptionally inactive 7SK/HEXIM/P-TEFb snRNP. Besides targeting P-TEFb to HIV transcription, Tat also increases the nuclear level of active P-TEFb through promoting its dissociation from the 7SK/HEXIM/P-TEFb RNP by an unclear mechanism. In this study, by using in vitro and in vivo RNA-protein binding assays, we demonstrate that HIV-1 Tat binds with high specificity and efficiency to an evolutionarily highly conserved stem-bulge-stem motif of the 5'-hairpin of human 7SK snRNA. The newly discovered Tat-binding motif of 7SK is structurally and functionally indistinguishable from the extensively characterized Tat-binding site of HIV TAR and importantly, it is imbedded in the HEXIM-binding elements of 7SK snRNA. We show that Tat efficiently replaces HEXIM1 on the 7SK snRNA in vivo and therefore, it promotes the disassembly of the 7SK/HEXIM/P-TEFb negative transcriptional regulatory snRNP to augment the nuclear level of active P-TEFb. This is the first demonstration that HIV-1 specifically targets an important cellular regulatory RNA, most probably to promote viral transcription and replication. Demonstration that the human 7SK snRNA carries a TAR RNA-like Tat-binding element that is essential for the normal transcriptional regulatory function of 7SK questions the viability of HIV therapeutic approaches based on small drugs blocking the Tat-binding site of HIV TAR.

Daily Distribution of Carbohydrate, Protein and Fat Intake in Elite Youth Academy Soccer Players Over a 7-Day Training Period.

Science.gov (United States)

Naughton, Robert J; Drust, Barry; O'Boyle, Andy; Morgans, Ryland; Abayomi, Julie; Davies, Ian G; Morton, James P; Mahon, Elizabeth

2016-10-01

While traditional approaches to dietary analysis in athletes have focused on total daily energy and macronutrient intake, it is now thought that daily distribution of these parameters can also influence training adaptations. Using 7-day food diaries, we quantified the total daily macronutrient intake and distribution in elite youth soccer players from the English Premier League in U18 (n = 13), U15/16 (n = 25) and U13/14 squads (n = 21). Total energy (43.1 ± 10.3, 32.6 ± 7.9, 28.1 ± 6.8 kcal·kg -1 ·day -1 ), CHO (6 ± 1.2, 4.7 ± 1.4, 3.2 ± 1.3 g·kg - 1 ·day -1 ) and fat (1.3 ± 0.5, 0.9 ± 0.3, 0.9 ± 0.3 g·kg- 1 ·day -1 ) intake exhibited hierarchical differences (p U15/16 > U18. In addition, CHO intake in U18s was lower (p lunch (~0.5 g·kg -1 ) > breakfast (~0.3 g·kg -1 ). We conclude elite youth soccer players do not meet current CHO guidelines. Although daily protein targets are achieved, we report a skewed daily distribution in all ages such that dinner > lunch > breakfast. Our data suggest that dietary advice for elite youth players should focus on both total daily macronutrient intake and optimal daily distribution patterns.

PTPBR7 binding proteins in myelinating neurons of the mouse brain

NARCIS (Netherlands)

Chesini, I.M.; Debyser, G.; Croes, H.J.E.; Dam, G.B. ten; Devreese, B.; Stoker, A.W.; Hendriks, W.J.A.J.

2011-01-01

Mouse protein tyrosine phosphatase PTPBR7 is a receptor-like, transmembrane protein that is localized on the surface of neuronal cells. Its protein phosphatase activity is reduced upon multimerization, and PTPBR7-deficient mice display motor coordination defects. Extracellular molecules that may

TEM characterization of irradiated U-7Mo/Mg dispersion fuel

Science.gov (United States)

Gan, J.; Keiser, D. D.; Miller, B. D.; Jue, J. F.; Robinson, A. B.; Madden, J.

2017-10-01

This paper presents the results of transmission electron microscopy (TEM) characterization on neutron-irradiated samples taken from the low-flux and high-flux sides of the same fuel plate with U-7Mo fuel particles dispersed in Mg matrix with aluminum alloy Al6061 as cladding material that was irradiated edge-on to the core in the Advanced Test Reactor. The corresponding local fission density and fission rate of the fuel particles and the average fuel-plate centerline temperature for the low-flux and high-flux samples are estimated to be 3.7 × 1021 f/cm3, 7.4 × 1014 f/cm3/s and 123 °C, and 5.5 × 1021 f/cm3, 11.0 × 1014 f/cm3/s and 158 °C, respectively. Complex interaction layers developed at the Al-Mg interface, consisting of Al3Mg2 and Al12Mg17 along with precipitates of MgO, Mg2Si and FeAl5.3. No interaction between Mg matrix and U-Mo fuel particle was identified. For the U-Mo fuel particles, at low fission density, small elongated bubbles wrapped around the clean areas with a fission gas bubble superlattice, which suggests that bubble coalescence is an important mechanism for converting the fission gas bubble superlattice to large bubbles. At high fission density, no bubbles or porosity were observed in the Mg matrix, and pockets of residual fission gas bubble superlattice were observed in the U-Mo fuel particle interior.

Protein: MPB2 [TP Atlas

Lifescience Database Archive (English)

Full Text Available MPB2 Ubiquitin ligases STUB1 CHIP STUB1 E3 ubiquitin-protein ligase CHIP Antigen NY...-CO-7, CLL-associated antigen KW-8, Carboxy terminus of Hsp70-interacting protein, STIP1 homology and U box-containing pr

7 CFR 1218.23 - U.S. Highbush Blueberry Council.

Science.gov (United States)

2010-01-01

... 7 Agriculture 10 2010-01-01 2010-01-01 false U.S. Highbush Blueberry Council. 1218.23 Section 1218... SERVICE (MARKETING AGREEMENTS AND ORDERS; MISCELLANEOUS COMMODITIES), DEPARTMENT OF AGRICULTURE BLUEBERRY PROMOTION, RESEARCH, AND INFORMATION ORDER Blueberry Promotion, Research, and Information Order Definitions...

Transmission electron microscopy characterization of irradiated U-7Mo/Al-2Si dispersion fuel

International Nuclear Information System (INIS)

Gan, J.; Keiser, D.D.; Wachs, D.M.; Robinson, A.B.; Miller, B.D.; Allen, T.R.

2010-01-01

The plate-type dispersion fuels, with the atomized U(Mo) fuel particles dispersed in the Al or Al alloy matrix, are being developed for use in research and test reactors worldwide. It is found that the irradiation performance of a plate-type dispersion fuel depends on the radiation stability of the various phases in a fuel plate. Transmission electron microscopy was performed on a sample (peak fuel mid-plane temperature ∼109 deg. C and fission density ∼4.5 x 10 27 f m -3 ) taken from an irradiated U-7Mo dispersion fuel plate with Al-2Si alloy matrix to investigate the role of Si addition in the matrix on the radiation stability of the phase(s) in the U-7Mo fuel/matrix interaction layer. A similar interaction layer that forms in irradiated U-7Mo dispersion fuels with pure Al matrix has been found to exhibit poor irradiation stability, likely as a result of poor fission gas retention. The interaction layer for both U-7Mo/Al-2Si and U-7Mo/Al fuels is observed to be amorphous. However, unlike the latter, the amorphous layer for the former was found to effectively retain fission gases in areas with high Si concentration. When the Si concentration becomes relatively low, the fission gas bubbles agglomerate into fewer large pores. Within the U-7Mo fuel particles, a bubble superlattice ordered as fcc structure and oriented parallel to the bcc metal lattice was observed where the average bubble size and the superlattice constant are 3.5 nm and 11.5 nm, respectively. The estimated fission gas inventory in the bubble superlattice correlates well with the fission density in the fuel.

Reaction layer between U-7WT%Mo and Al alloys in chemical diffusion couples

International Nuclear Information System (INIS)

Mirandou, M.; Granovsky, M.; Ortiz, M.; Balart, S.; Arico, S.; Gribaudo, L.

2005-01-01

Several failures in U-Mo dispersion fuel plates like pillowing and large porosities have been reported during irradiation experiments. These failures have been assigned to the formation of a large (U-Mo)/Al interaction product under high operating conditions. The modification of the matrix by alloying Al to change the interaction layer and improve its irradiation behavior, has been proposed. This paper reports diffusion experiments performed between U-7wt%Mo and various Al alloys containing Mg and / or Si. By the use of Optical Microscopy, SEM and X-Ray diffraction, it was found that with a concentration of 5.2wt% or 7.1 wt%Si the interaction layer is constituted mainly by (U,Mo)(Si,Al) 3 and no (U,Mo)Al 4 is detected. As part of the studies of properties of the U-Mo alloys the time for isothermal transformation start at different temperatures of the γ phase is being evaluated for the present U-7wt%Mo alloy. These results are used to plan the future diffusion program that will include diffusion under irradiation at CNEA RA3 reactor. (author)

Usage of U7 snRNA in gene therapy of hemoglobin C disorder ...

African Journals Online (AJOL)

Here, a bioinformatic analysis was performed to study the effect of co-expression between human Hb C b-globin chain gene and U7.623. The gene ontological results show that full recovery of hemoglobin function and biological process can be derived. This confirms that U7 snRNA can be a good tool for gene therapy in Hb ...

The reconstruction of HT-7 superconducting tokamak and the present status of HT-7U project

International Nuclear Information System (INIS)

Weng, P.D.

2000-01-01

The first Chinese superconducting tokamak HT-7 was reconstructed from T-7. The main purposes of reconstruction are to improve the accessibility of the device and to provide a possibility of long pulse operation with high performance. The reconstruction has been done successfully. The HT-7U project has been approved and funded as a National Project, the engineering design and R and D are on the way. (author)

ON-GOING STATUS OF KJRR FUEL (U-7MO) QUALIFICATION

Energy Technology Data Exchange (ETDEWEB)

Yim, J. S.; Tahk, Y. W.; Oh, J. Y.; Kim, H. J.; Kong, E. H.; Lee, B. H.; Park, J. M.; Jeong, Y. J.; Lee, K. H.; Kim, S. H.; Lee, C. T.; Beasley, A. A.; Choi, Y. J.; Crawford, D. S.; Nielsen, J. W.; Woolstenhulme, N. E.

2017-03-01

In order to cope with global shortage of Mo-99 supplies and with growing demand of neutron transmutation doping, KJRR construction plan has been launched since April 2012 to provide self-sufficiency of domestic RI demand, and to extend Si doping capacity for power device market growth. Through comprehensive surveillance of the fuels in-reactor behavior, KAERI has selected the fuel meat of U-7%Mo dispersion in an aluminum matrix with 5wt%Si for the KJRR fuel. As part of the efforts for fuel licensing and qualification of the KJRR fuel, an LTA irradiation test at the ATR started from November 2015 was successfully completed by reaching at 219 EFPD in the end of February 2017. Together with the results of HAMP-1 already completed irradiation and PIE, the successful irradiation of the LTA also demonstrates the fuel integrity under more rigorous conditions than the KJRR operation conditions. This paper updates the current status of the KJRR U7Mo (8 g-U/cm3) LTA irradiation and PIE plan up to date as of February 2017.

Large-scale purification and crystallization of the endoribonuclease XendoU: troubleshooting with His-tagged proteins

International Nuclear Information System (INIS)

Renzi, Fabiana; Panetta, Gianna; Vallone, Beatrice; Brunori, Maurizio; Arceci, Massimo; Bozzoni, Irene; Laneve, Pietro; Caffarelli, Elisa

2006-01-01

Recombinant His-tagged XendoU, a eukaryotic endoribonuclease, appeared to aggregate in the presence of divalent cations. Monodisperse protein which yielded crystals diffracting to 2.2 Å was obtained by addition of EDTA. XendoU is the first endoribonuclease described in higher eukaryotes as being involved in the endonucleolytic processing of intron-encoded small nucleolar RNAs. It is conserved among eukaryotes and its viral homologue is essential in SARS replication and transcription. The large-scale purification and crystallization of recombinant XendoU are reported. The tendency of the recombinant enzyme to aggregate could be reversed upon the addition of chelating agents (EDTA, imidazole): aggregation is a potential drawback when purifying and crystallizing His-tagged proteins, which are widely used, especially in high-throughput structural studies. Purified monodisperse XendoU crystallized in two different space groups: trigonal P3 1 21, diffracting to low resolution, and monoclinic C2, diffracting to higher resolution

7 CFR 51.2078 - U.S. No. 2 Mixed.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 2 Mixed. 51.2078 Section 51.2078 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946...

7 CFR 51.2076 - U.S. No. 1 Mixed.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. No. 1 Mixed. 51.2076 Section 51.2076 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946...

7 CFR 51.1177 - U.S. Grade A Juice.

Science.gov (United States)

2010-01-01

... STANDARDS) United States Standards for Grades of Florida Oranges and Tangelos Standards for Internal Quality... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Grade A Juice. 51.1177 Section 51.1177 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards...

Structure and thermal properties of as-fabricated U-7Mo/Mg and U-10Mo/Mg low-enriched uranium research reactor fuels

Energy Technology Data Exchange (ETDEWEB)

Kulakov, Mykola, E-mail: mykola.kulakov@cnl.ca [Fuel Development Branch, Canadian Nuclear Laboratories, Chalk River, ON K0J 1J0 Canada (Canada); Saoudi, Mouna [Fuel Development Branch, Canadian Nuclear Laboratories, Chalk River, ON K0J 1J0 Canada (Canada); Piro, Markus H.A. [Fuel and Fuel Channel Safety Branch, Canadian Nuclear Laboratories, Chalk River, ON K0J 1J0 Canada (Canada); Donaberger, Ronald L. [Canadian Neutron Beam Centre, Chalk River, ON K0J 1J0 Canada (Canada)

2017-02-15

Aluminum-clad U-7Mo/Mg and U-10Mo/Mg pin-type mini-elements (with a core uranium loading of 4.5 gU/cm{sup 3}) have been fabricated at the Canadian Nuclear Laboratories for experimental tests and ultimately for use in research and test reactors. In this study, the microstructure and phase composition of unirradiated U-7Mo/Mg and U-10Mo/Mg fuel cores were analyzed using optical and scanning electron microscopy, and neutron powder diffraction. Thermal properties were characterized using a combination of experimental measurements and thermodynamic calculations. The thermal diffusivity was measured using the laser flash method. The temperature-dependent specific heat capacities were calculated based on the linear rule of mixture using the weight fraction of different crystalline phases and their specific heat capacity values taken from the literature. The thermal conductivity was then calculated using the measured thermal diffusivity, the measured density and the calculated specific heat capacity. The resulting thermal conductivity is practically identical for both types of fuel. The in-reactor temperatures were predicted using conjugate heat transfer simulations. - Highlights: • Neutron diffraction analysis shows that most of the γ-U(Mo) phase was retained in as-fabricated U-7Mo/Mg and U-10Mo/Mg fuel cores. • The experimental thermal conductivity of both types of fuel is practically identical. • Based on conjugate heat transfer simulations, under normal operating conditions, the in-reactor fuel centreline temperature is about 510 K.

Disruption of endocytic trafficking protein Rab7 impairs invasiveness of cholangiocarcinoma cells.

Science.gov (United States)

Suwandittakul, Nantana; Reamtong, Onrapak; Molee, Pattamaporn; Maneewatchararangsri, Santi; Sutherat, Maleerat; Chaisri, Urai; Wongkham, Sopit; Adisakwattana, Poom

2017-09-07

Alterations and mutations of endo-lysosomal trafficking proteins have been associated with cancer progression. Identification and characterization of endo-lysosomal trafficking proteins in invasive cholangiocarcinoma (CCA) cells may benefit prognosis and drug design for CCA. To identify and characterize endo-lysosomal trafficking proteins in invasive CCA. A lysosomal-enriched fraction was isolated from a TNF-α induced invasive CCA cell line (KKU-100) and uninduced control cells and protein identification was performed with nano-LC MS/MS. Novel lysosomal proteins that were upregulated in invasive CCA cells were validated by real-time RT-PCR. We selected Rab7 for further studies of protein level using western blotting and subcellular localization using immunofluorescence. The role of Rab7 in CCA invasion was determined by siRNA gene knockdown and matrigel transwell assay. Rab7 mRNA and protein were upregulated in invasive CCA cells compared with non-treated controls. Immunofluorescence studies demonstrated that Rab7 was expressed predominantly in invasive CCA cells and was localized in the cytoplasm and lysosomes. Suppression of Rab7 translation significantly inhibited TNF-α-induced cell invasion compared to non-treated control (p= 0.044). Overexpression of Rab7 in CCA cells was associated with cell invasion, supporting Rab7 as a novel candidate for the development of diagnostic and therapeutic strategies for CCA.

K X-rays and nuclear reaction times in the deep inelastic reactions U+U and U+Pb at 7.5 MeV/amu

International Nuclear Information System (INIS)

Stoller, C.

1985-01-01

The K-shell ionisation probability of the heavy reaction products emerging from binary deep inelastic collisions of U + U and U + Pb at 7.5 MeV/amu has been measured as a function of the total kinetic energy loss - Q. After subtraction of the ionisation probability due to internal conversion of γ-rays, a strongly Q-dependent Psub(K) is found, in agreement with theoretical predictions relating the change in ionisation probability to the nuclear sticking time. The deduced nuclear reaction times are in qualitative agreement with predictions from nuclear models of deep inelastic reactions. (orig.)

Small kernel 1 encodes a pentatricopeptide repeat protein required for mitochondrial nad7 transcript editing and seed development in maize (Zea mays) and rice (Oryza sativa).

Science.gov (United States)

Li, Xiao-Jie; Zhang, Ya-Feng; Hou, Mingming; Sun, Feng; Shen, Yun; Xiu, Zhi-Hui; Wang, Xiaomin; Chen, Zong-Liang; Sun, Samuel S M; Small, Ian; Tan, Bao-Cai

2014-09-01

RNA editing modifies cytidines (C) to uridines (U) at specific sites in the transcripts of mitochondria and plastids, altering the amino acid specified by the DNA sequence. Here we report the identification of a critical editing factor of mitochondrial nad7 transcript via molecular characterization of a small kernel 1 (smk1) mutant in Zea mays (maize). Mutations in Smk1 arrest both the embryo and endosperm development. Cloning of Smk1 indicates that it encodes an E-subclass pentatricopeptide repeat (PPR) protein that is targeted to mitochondria. Loss of SMK1 function abolishes the C → U editing at the nad7-836 site, leading to the retention of a proline codon that is edited to encode leucine in the wild type. The smk1 mutant showed dramatically reduced complex-I assembly and NADH dehydrogenase activity, and abnormal biogenesis of the mitochondria. Analysis of the ortholog in Oryza sativa (rice) reveals that rice SMK1 has a conserved function in C → U editing of the mitochondrial nad7-836 site. T-DNA knock-out mutants showed abnormal embryo and endosperm development, resulting in embryo or seedling lethality. The leucine at NAD7-279 is highly conserved from bacteria to flowering plants, and analysis of genome sequences from many plants revealed a molecular coevolution between the requirement for C → U editing at this site and the existence of an SMK1 homolog. These results demonstrate that Smk1 encodes a PPR-E protein that is required for nad7-836 editing, and this editing is critical to NAD7 function in complex-I assembly in mitochondria, and hence to embryo and endosperm development in maize and rice. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

The calcium-dependent protein kinase CPK7 acts on root hydraulic conductivity.

Science.gov (United States)

Li, Guowei; Boudsocq, Marie; Hem, Sonia; Vialaret, Jérôme; Rossignol, Michel; Maurel, Christophe; Santoni, Véronique

2015-07-01

The hydraulic conductivity of plant roots (Lp(r)) is determined in large part by the activity of aquaporins. Mechanisms occurring at the post-translational level, in particular phosphorylation of aquaporins of the plasma membrane intrinsic protein 2 (PIP2) subfamily, are thought to be of critical importance for regulating root water transport. However, knowledge of protein kinases and phosphatases acting on aquaporin function is still scarce. In the present work, we investigated the Lp(r) of knockout Arabidopsis plants for four Ca(2+)-dependent protein kinases. cpk7 plants showed a 30% increase in Lp(r) because of a higher aquaporin activity. A quantitative proteomic analysis of wild-type and cpk7 plants revealed that PIP gene expression and PIP protein quantity were not correlated and that CPK7 has no effect on PIP2 phosphorylation. In contrast, CPK7 exerts a negative control on the cellular abundance of PIP1s, which likely accounts for the higher Lp(r) of cpk7. In addition, this study revealed that the cellular amount of a few additional proteins including membrane transporters is controlled by CPK7. The overall work provides evidence for CPK7-dependent stability of specific membrane proteins. © 2014 John Wiley & Sons Ltd.

Phylogeny and evolution of Rab7 and Rab9 proteins

Directory of Open Access Journals (Sweden)

Wyroba Elżbieta

2009-05-01

Full Text Available Abstract Background An important role in the evolution of intracellular trafficking machinery in eukaryotes played small GTPases belonging to the Rab family known as pivotal regulators of vesicle docking, fusion and transport. The Rab family is very diversified and divided into several specialized subfamilies. We focused on the VII functional group comprising Rab7 and Rab9, two related subfamilies, and analysed 210 sequences of these proteins. Rab7 regulates traffic from early to late endosomes and from late endosome to vacuole/lysosome, whereas Rab9 participates in transport from late endosomes to the trans-Golgi network. Results Although Rab7 and Rab9 proteins are quite small and show heterogeneous rates of substitution in different lineages, we found a phylogenetic signal and inferred evolutionary relationships between them. Rab7 proteins evolved before radiation of main eukaryotic supergroups while Rab9 GTPases diverged from Rab7 before split of choanoflagellates and metazoans. Additional duplication of Rab9 and Rab7 proteins resulting in several isoforms occurred in the early evolution of vertebrates and next in teleost fishes and tetrapods. Three Rab7 lineages emerged before divergence of monocots and eudicots and subsequent duplications of Rab7 genes occurred in particular angiosperm clades. Interestingly, several Rab7 copies were identified in some representatives of excavates, ciliates and amoebozoans. The presence of many Rab copies is correlated with significant differences in their expression level. The diversification of analysed Rab subfamilies is also manifested by non-conserved sequences and structural features, many of which are involved in the interaction with regulators and effectors. Individual sites discriminating different subgroups of Rab7 and Rab9 GTPases have been identified. Conclusion Phylogenetic reconstructions of Rab7 and Rab9 proteins were performed by a variety of methods. These Rab GTPases show diversification

ORF7-encoded accessory protein 7a of feline infectious peritonitis virus as a counteragent against IFN-α-induced antiviral response.

Science.gov (United States)

Dedeurwaerder, Annelike; Olyslaegers, Dominique A J; Desmarets, Lowiese M B; Roukaerts, Inge D M; Theuns, Sebastiaan; Nauwynck, Hans J

2014-02-01

The type I IFN-mediated immune response is the first line of antiviral defence. Coronaviruses, like many other viruses, have evolved mechanisms to evade this innate response, ensuring their survival. Several coronavirus accessory genes play a central role in these pathways, but for feline coronaviruses this has never to our knowledge been studied. As it has been demonstrated previously that ORF7 is essential for efficient replication in vitro and virulence in vivo of feline infectious peritonitis virus (FIPV), the role of this ORF in the evasion of the IFN-α antiviral response was investigated. Deletion of ORF7 from FIPV strain 79-1146 (FIPV-Δ7) rendered the virus more susceptible to IFN-α treatment. Given that ORF7 encodes two proteins, 7a and 7b, it was further explored which of these proteins is active in this mechanism. Providing 7a protein in trans rescued the mutant FIPV-Δ7 from IFN sensitivity, which was not achieved by addition of 7b protein. Nevertheless, addition of protein 7a to FIPV-Δ3Δ7, a FIPV mutant deleted in both ORF3 and ORF7, could no longer increase the replication capacity of this mutant in the presence of IFN. These results indicate that FIPV 7a protein is a type I IFN antagonist and protects the virus from the antiviral state induced by IFN, but it needs the presence of ORF3-encoded proteins to exert its antagonistic function.

Fission Product Yields of 233U, 235U, 238U and 239Pu in Fields of Thermal Neutrons, Fission Neutrons and 14.7-MeV Neutrons

Science.gov (United States)

Laurec, J.; Adam, A.; de Bruyne, T.; Bauge, E.; Granier, T.; Aupiais, J.; Bersillon, O.; Le Petit, G.; Authier, N.; Casoli, P.

2010-12-01

The yields of more than fifteen fission products have been carefully measured using radiochemical techniques, for 235U(n,f), 239Pu(n,f) in a thermal spectrum, for 233U(n,f), 235U(n,f), and 239Pu(n,f) reactions in a fission neutron spectrum, and for 233U(n,f), 235U(n,f), 238U(n,f), and 239Pu(n,f) for 14.7 MeV monoenergetic neutrons. Irradiations were performed at the EL3 reactor, at the Caliban and Prospero critical assemblies, and at the Lancelot electrostatic accelerator in CEA-Valduc. Fissions were counted in thin deposits using fission ionization chambers. The number of fission products of each species were measured by gamma spectrometry of co-located thick deposits.

Effects of Silicide Coating on the Interdiffusion between U-7Mo and Al

Energy Technology Data Exchange (ETDEWEB)

Nam, Ji Min; Kim, Ji Hyun; Kim, Sunghwan; Lee, Kyu Hong; Park, Jong Man; Jeong, Yong Jin; Kim, Ki Nam [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

2015-05-15

The excessive interaction between the U-Mo alloys and their surrounding Al matrix lead to and excessive local swelling called 'pillowing'. For this reason, KAERI suggested several remedies such as alloying U-Mo with Ti, or Al matrix with Si. In addition, silicide, or nitride coatings on the surface of U-Mo particles have also been proposed to hinder the growth of interaction layer. In this study, centrifugally atomized U-7Mo alloy powders were coated with silicide layers at varying T (T = 900 and 1000 .deg. C) for 30 min, respectively. U-Mo alloy powder was blended with Si powders and subsequently heat-treated to form uranium-silicide coating layers on the surface of U-Mo alloy particles. For an annealing test, silicide-coated U-Mo alloy powders were made into a compact, and Al powders were used as a matrix. From EDS results, transformed uranium aluminide intermetallic compounds were mainly U(Al,Si)3. U(Al,Si)3 phase left the silicide coating layer behind, and formed inside of U-7Mo particles, as shown in Fig. 3(a) and (b). In the case of sample B, Al could not penetrate the silicide coating layer and the coating layers were remained constant, as shown in Fig. 3(c) and (d). From the results, we made a comparison between the compacts of sample A and B, and it was shown that Al can easily diffuse into unreacted Si and U{sub 3}Si{sub 5} mixed layer while U{sub 3}Si{sub 2} acted as a good diffusion barrier at 550 .deg. C though those layers had the same thickness.

Effects of Silicide Coating on the Interdiffusion between U-7Mo and Al

International Nuclear Information System (INIS)

Nam, Ji Min; Kim, Ji Hyun; Kim, Sunghwan; Lee, Kyu Hong; Park, Jong Man; Jeong, Yong Jin; Kim, Ki Nam

2015-01-01

The excessive interaction between the U-Mo alloys and their surrounding Al matrix lead to and excessive local swelling called 'pillowing'. For this reason, KAERI suggested several remedies such as alloying U-Mo with Ti, or Al matrix with Si. In addition, silicide, or nitride coatings on the surface of U-Mo particles have also been proposed to hinder the growth of interaction layer. In this study, centrifugally atomized U-7Mo alloy powders were coated with silicide layers at varying T (T = 900 and 1000 .deg. C) for 30 min, respectively. U-Mo alloy powder was blended with Si powders and subsequently heat-treated to form uranium-silicide coating layers on the surface of U-Mo alloy particles. For an annealing test, silicide-coated U-Mo alloy powders were made into a compact, and Al powders were used as a matrix. From EDS results, transformed uranium aluminide intermetallic compounds were mainly U(Al,Si)3. U(Al,Si)3 phase left the silicide coating layer behind, and formed inside of U-7Mo particles, as shown in Fig. 3(a) and (b). In the case of sample B, Al could not penetrate the silicide coating layer and the coating layers were remained constant, as shown in Fig. 3(c) and (d). From the results, we made a comparison between the compacts of sample A and B, and it was shown that Al can easily diffuse into unreacted Si and U 3 Si 5 mixed layer while U 3 Si 2 acted as a good diffusion barrier at 550 .deg. C though those layers had the same thickness

Gradient elution behavior of proteins in hydrophobic interaction chromatography with U-shaped retention factor curves.

Science.gov (United States)

Creasy, Arch; Lomino, Joseph; Barker, Gregory; Khetan, Anurag; Carta, Giorgio

2018-04-27

Protein retention in hydrophobic interaction chromatography is described by the solvophobic theory as a function of the kosmostropic salt concentration. In general, an increase in salt concentration drives protein partitioning to the hydrophobic surface while a decrease reduces it. In some cases, however, protein retention also increases at low salt concentrations resulting in a U-shaped retention factor curve. During gradient elution the salt concentration is gradually decreased from a high value thereby reducing the retention factor and increasing the protein chromatographic velocity. For these conditions, a steep gradient can overtake the protein in the column, causing it to rebind. Two dynamic models, one based on the local equilibrium theory and the other based on the linear driving force approximation, are presented. We show that the normalized gradient slope determines whether the protein elutes in the gradient, partially elutes, or is trapped in the column. Experimental results are presented for two different monoclonal antibodies and for lysozyme on Capto Phenyl (High Sub) resin. One of the mAbs and lysozyme exhibit U-shaped retention factor curves and for each, we determine the critical gradient slope beyond which 100% recovery is no longer possible. Elution with a reverse gradient is also demonstrated at low salt concentrations for these proteins. Understanding this behavior has implications in the design of gradient elution since the gradient slope impacts protein recovery. Copyright © 2018 Elsevier B.V. All rights reserved.

Expression of an IRF-3 fusion protein and mouse estrogen receptor, inhibits hepatitis C viral replication in RIG-I-deficient Huh 7.5 cells

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Liu Chen

2011-09-01

Full Text Available Abstract Interferon Regulatory Factor-3 (IRF-3 plays a central role in the induction of interferon (IFN production and succeeding interferon-stimulated genes (ISG expression en route for restraining hepatitis C virus (HCV infection. Here, we established a stable Huh7.5-IRF3ER cell line expressing a fusion protein of IRF-3 and mouse estrogen receptor (ER to examine IFN production and anti-HCV effects of IRF-3 in retinoic acid inducible-gene-I (RIG-I deficient Huh 7.5 cells. Homodimerization of the IRF-3ER fusion protein was detected by Western blotting after treatment with the estrogen receptor agonist 4-hydrotamoxifen (4-HT in Huh7.5-IRF3ER cells. Expression of IFN-α, IFN-β, and their inhibitory effects on HCV replication were demonstrated by real-time polymerase chain reaction (PCR. Peak expression of IFN-α and IFN-β was achieved 24-hours post 4-HT treatment, coinciding with the appearance of phosphorylated signal transducer and activator of transcription (STAT proteins. Additionally, HCV viral replication declined in time-dependent fashion. In previous studies, a novel IFN-mediated pathway regulating expression of 1-8U and heterogeneous nuclear ribonucleoprotein M (hnRNP M inhibited HCV internal ribosomal entry site (IRES-dependent translation. When expression of ISGs such as 1-8U and hnRNP M were measured in 4-HT-treated Huh7.5-IRF3ER cells, both genes were positively regulated by activation of the IRF-3ER fusion protein. In conclusion, the anti-HCV effects of IRF-3ER homodimerization inhibited HCV RNA replication as well as HCV IRES-dependent translation in Huh7.5-IRF3ER cells. The results of this study indicate that IRF-3ER homodimerization is a key step to restore IFN expression in Huh7.5-IRF3ER cells and in achieving its anti-HCV effects.

The Transient Receptor Potential Melastatin 7 Channel Regulates Pancreatic Cancer Cell Invasion through the Hsp90α/uPA/MMP2 pathway

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Pierre Rybarczyk

2017-04-01

Full Text Available Pancreatic ductal adenocarcinoma (PDAC is an aggressive malignancy with a very poor prognosis. There is an urgent need to better understand the molecular mechanisms that regulate PDAC cell aggressiveness. The transient receptor potential melastatin 7 (TRPM7 is a nonselective cationic channel that mainly conducts Ca2+ and Mg2+. TRPM7 is overexpressed in numerous malignancies including PDAC. In the present study, we used the PANC-1 and MIA PaCa-2 cell lines to specifically assess the role of TRPM7 in cell invasion and matrix metalloproteinase secretion. We show that TRPM7 regulates Mg2+ homeostasis and constitutive cation entry in both PDAC cell lines. Moreover, cell invasion is strongly reduced by TRPM7 silencing without affecting the cell viability. Conditioned media were further studied, by gel zymography, to detect matrix metalloproteinase (MMP secretion in PDAC cells. Our results show that MMP-2, urokinase plasminogen activator (uPA, and heat-shock protein 90α (Hsp90α secretions are significantly decreased in TRPM7-deficient PDAC cells. Moreover, TRPM7 expression in human PDAC lymph node metastasis is correlated to the channel expression in primary tumor. Taken together, our results show that TRPM7 is involved in PDAC cell invasion through regulation of Hsp90α/uPA/MMP-2 proteolytic axis, confirming that this channel could be a promising biomarker and possibly a target for PDAC metastasis therapy.

26 CFR 1.871-7 - Taxation of nonresident alien individuals not engaged in U.S. business.

Science.gov (United States)

2010-04-01

... 26 Internal Revenue 9 2010-04-01 2010-04-01 false Taxation of nonresident alien individuals not engaged in U.S. business. 1.871-7 Section 1.871-7 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF... Corporations § 1.871-7 Taxation of nonresident alien individuals not engaged in U.S. business. (a) Imposition...

Corium Spreading Over Concrete: The Vulcano VE-U7 and VE-U8 Tests

International Nuclear Information System (INIS)

Journeau, Christophe; Boccaccio, Eric; Fouquart, Pascal; Jegou, Claude; Piluso, Pascal

2002-01-01

Two experiments have been performed in the VULCANO facility in which prototypic corium has been spread over concrete. In the VE-U7 test, a mixture representative of what can be expected at the opening of EPR reactor-pit gate has been spread on siliceous concrete and on a reference channel in inert refractory ceramic. The spreading progression was not much affected by the presence of concrete and sparging gases. In the VE-U8 test, a UO 2 -ZrO 2 mixture, prototypic of in-vessel corium, has been spread over a lime-siliceous concrete. Although residual power was not simulated in this experiment, up to 2 cm of concrete have been eroded during the test. Results in terms of spreading behaviour, effects of gases, concrete erosion and thermal attack are presented and discussed. (authors)

Whirlin and PDZ domain-containing 7 (PDZD7) proteins are both required to form the quaternary protein complex associated with Usher syndrome type 2.

Science.gov (United States)

Chen, Qian; Zou, Junhuang; Shen, Zuolian; Zhang, Weiping; Yang, Jun

2014-12-26

Usher syndrome (USH) is the leading genetic cause of combined hearing and vision loss. Among the three USH clinical types, type 2 (USH2) occurs most commonly. USH2A, GPR98, and WHRN are three known causative genes of USH2, whereas PDZD7 is a modifier gene found in USH2 patients. The proteins encoded by these four USH genes have been proposed to form a multiprotein complex, the USH2 complex, due to interactions found among some of these proteins in vitro, their colocalization in vivo, and mutual dependence of some of these proteins for their normal in vivo localizations. However, evidence showing the formation of the USH2 complex is missing, and details on how this complex is formed remain elusive. Here, we systematically investigated interactions among the intracellular regions of the four USH proteins using colocalization, yeast two-hybrid, and pull-down assays. We show that multiple domains of the four USH proteins interact among one another. Importantly, both WHRN and PDZD7 are required for the complex formation with USH2A and GPR98. In this USH2 quaternary complex, WHRN prefers to bind to USH2A, whereas PDZD7 prefers to bind to GPR98. Interaction between WHRN and PDZD7 is the bridge between USH2A and GPR98. Additionally, the USH2 quaternary complex has a variable stoichiometry. These findings suggest that a non-obligate, short term, and dynamic USH2 quaternary protein complex may exist in vivo. Our work provides valuable insight into the physiological role of the USH2 complex in vivo and informs possible reconstruction of the USH2 complex for future therapy. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Role of bone morphogenetic protein-7 in renal fibrosis

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Rui Xi eLi

2015-04-01

Full Text Available Renal fibrosis is final common pathway of end stage renal disease. Irrespective of the primary cause, renal fibrogenesis is a dynamic process which involves a large network of cellular and molecular interaction, including pro-inflammatory cell infiltration and activation, matrix-producing cell accumulation and activation, and secretion of profibrogenic factors that modulate extracellular matrix (ECM formation and cell-cell interaction. Bone morphogenetic protein-7 is a protein of the TGF-β super family and increasingly regarded as a counteracting molecule against TGF-β. A large variety of evidence shows an anti-fibrotic role of BMP-7 in chronic kidney disease, and this effect is largely mediated via counterbalancing the profibrotic effect of TGF-β. Besides, BMP-7 reduced ECM formation by inactivating matrix-producing cells and promoting mesenchymal-to-epithelial transition (MET. BMP-7 also increased ECM degradation. Despite these observations, the anti-fibrotic effect of BMP-7 is still controversial such that fine regulation of BMP-7 expression in vivo might be a great challenge for its ultimate clinical application.

Role of bone morphogenetic protein-7 in renal fibrosis

Science.gov (United States)

Li, Rui Xi; Yiu, Wai Han; Tang, Sydney C. W.

2015-01-01

Renal fibrosis is final common pathway of end stage renal disease. Irrespective of the primary cause, renal fibrogenesis is a dynamic process which involves a large network of cellular and molecular interaction, including pro-inflammatory cell infiltration and activation, matrix-producing cell accumulation and activation, and secretion of profibrogenic factors that modulate extracellular matrix (ECM) formation and cell-cell interaction. Bone morphogenetic protein-7 is a protein of the TGF-β super family and increasingly regarded as a counteracting molecule against TGF-β. A large variety of evidence shows an anti-fibrotic role of BMP-7 in chronic kidney disease, and this effect is largely mediated via counterbalancing the profibrotic effect of TGF-β. Besides, BMP-7 reduced ECM formation by inactivating matrix-producing cells and promoting mesenchymal-to-epithelial transition (MET). BMP-7 also increased ECM degradation. Despite these observations, the anti-fibrotic effect of BMP-7 is still controversial such that fine regulation of BMP-7 expression in vivo might be a great challenge for its ultimate clinical application. PMID:25954203

Towards steady-state operational design for the data and PF control systems of the HT-7U

International Nuclear Information System (INIS)

Luo, J.R.; Zhu, L.; Wang, H.Z.; Ji, Z.S.; Wang, F.

2003-01-01

Fusion energy is an ultimate and inexhaustible source of energy for mankind and is expected to be obtained in controlled operation within this century. Among various possible candidates for fusion, the tokamak is presently the most qualified one, and since it uses superconducting magnetic coils, it will be adequate for steady-state operation. The HT-7U superconducting tokamak is a part of national project in China on fusion research, scheduled to become available on-line by the end of 2004 (Wan Y.X. and HT-7 and HT-7U Groups 2000 Overview of steady state operation of HT-7 and present status of the HT-7U project Nucl. Fusion 40 1057). The control system of the HT-7U is designed as a distributed control system (HT7UDCS), including many subsystems that provide the various functions of supervision, remote control, real-time monitoring, data acquisition and data handling. The major features of the HT-7U tokamak, which make long-pulse (∼1000 s) operation possible are the flexible poloidal field (PF) system, an auxiliary heating system, the current-driving system and a divertor system. In order to realize these features simultaneously, real-time data handling and analysis, along with a significant control capability is required. This paper discusses the design of the HT7UDCS. (author)

RNA-binding domain of the A protein component of the U1 small nuclear ribonucleoprotein analyzed by NMR spectroscopy is structurally similar to ribosomal proteins

International Nuclear Information System (INIS)

Hoffman, D.W.; Query, C.C.; Golden, B.L.; White, S.W.; Keene, J.D.

1991-01-01

An RNA recognition motif (RRM) of ∼80 amino acids constitutes the core of RNA-binding domains found in a large family of proteins involved in RNA processing. The U1 RNA-binding domain of the A protein component of the human U1 small nuclear ribonucleoprotein (RNP), which encompasses the RRM sequence, was analyzed by using NMR spectroscopy. The domain of the A protein is a highly stable monomer in solution consisting of four antiparallel β-strands and two α-helices. The highly conserved RNP1 and RNP2 consensus sequences, containing residues previously suggested to be involved in nucleic acid binding, are juxtaposed in adjacent β-strands. Conserved aromatic side chains that are critical for RNA binding are clustered on the surface to the molecule adjacent to a variable loop that influences recognition of specific RNA sequences. The secondary structure and topology of the RRM are similar to those of ribosomal proteins L12 and L30, suggesting a distant evolutionary relationship between these two types of RNA-associated proteins

Supraspliceosomes at Defined Functional States Portray the Pre-Assembled Nature of the Pre-mRNA Processing Machine in the Cell Nucleus

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Hani Kotzer-Nevo

2014-06-01

Full Text Available 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.

Small amounts of functional ATP7A protein permit mild phenotype

DEFF Research Database (Denmark)

Møller, Lisbeth Birk

2015-01-01

concentrations, ATP7A shifts to the post-Golgi compartments or to the plasma membrane to export copper out of the cell. Impaired copper-regulation trafficking has been observed for ATP7A mutants, but its impact on the clinical outcome is not clear. The major problem in patients with MD seems to be insufficient...... of missense mutations on structural models of the ATP7A protein suggests that affected conserved residues generally lead to a severe phenotype. The ATP7A protein traffics within the cells. At low copper levels, ATP7A locates to the Trans-Golgi Network (TGN) to load cuproenzymes with copper, whereas at higher...

The Sso7d protein of Sulfolobus solfataricus: in vitro relationship among different activities

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Annamaria Guagliardi

2002-01-01

Full Text Available The physiological role of the nonspecific DNA-binding protein Sso7d from the crenarchaeon Sulfolobus solfataricus is unknown. In vitro studies have shown that Sso7d promotes annealing of complementary DNA strands (Guagliardi et al. 1997, induces negative supercoiling (Lopez-Garcia et al. 1998, and chaperones the disassembly and renaturation of protein aggregates in an ATP hydrolysis-dependent manner (Guagliardi et al. 2000. In this study, we examined the relationships among the binding of Sso7d to double-stranded DNA, its interaction with protein aggregates, and its ATPase activity. Experiments with 1-anilinonaphthalene-8-sulfonic acid as probe demonstrated that exposed hydrophobic surfaces in Sso7d are responsible for interactions with protein aggregates and double-stranded DNA, whereas the site of ATPase activity has a non-hydrophobic character. The interactions of Sso7d with double-stranded DNA and with protein aggregates are mutually exclusive events, suggesting that the disassembly activity and the DNA-related activities of Sso7d may be competitive in vivo. In contrast, the hydrolysis of ATP by Sso7d is independent of the binding of Sso7d to double-stranded DNA or protein aggregates.

Molecular evolution of a-kinase anchoring protein (AKAP-7: implications in comparative PKA compartmentalization

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Johnson Keven R

2012-07-01

Full Text Available Abstract Background A-Kinase Anchoring Proteins (AKAPs are molecular scaffolding proteins mediating the assembly of multi-protein complexes containing cAMP-dependent protein kinase A (PKA, directing the kinase in discrete subcellular locations. Splice variants from the AKAP7 gene (AKAP15/18 are vital components of neuronal and cardiac phosphatase complexes, ion channels, cardiac Ca2+ handling and renal water transport. Results Shown in evolutionary analyses, the formation of the AKAP7-RI/RII binding domain (required for AKAP/PKA-R interaction corresponds to vertebrate-specific gene duplication events in the PKA-RI/RII subunits. Species analyses of AKAP7 splice variants shows the ancestral AKAP7 splice variant is AKAP7α, while the ancestral long form AKAP7 splice variant is AKAP7γ. Multi-species AKAP7 gene alignments, show the recent formation of AKAP7δ occurs with the loss of native AKAP7γ in rats and basal primates. AKAP7 gene alignments and two dimensional Western analyses indicate that AKAP7γ is produced from an internal translation-start site that is present in the AKAP7δ cDNA of mice and humans but absent in rats. Immunofluorescence analysis of AKAP7 protein localization in both rat and mouse heart suggests AKAP7γ replaces AKAP7δ at the cardiac sarcoplasmic reticulum in species other than rat. DNA sequencing identified Human AKAP7δ insertion-deletions (indels that promote the production of AKAP7γ instead of AKAP7δ. Conclusions This AKAP7 molecular evolution study shows that these vital scaffolding proteins developed in ancestral vertebrates and that independent mutations in the AKAP7 genes of rodents and early primates has resulted in the recent formation of AKAP7δ, a splice variant of likely lesser importance in humans than currently described.

Association with the Plasma Membrane Is Sufficient for Potentiating Catalytic Activity of Regulators of G Protein Signaling (RGS) Proteins of the R7 Subfamily.

Science.gov (United States)

Muntean, Brian S; Martemyanov, Kirill A

2016-03-25

Regulators of G protein Signaling (RGS) promote deactivation of heterotrimeric G proteins thus controlling the magnitude and kinetics of responses mediated by G protein-coupled receptors (GPCR). In the nervous system, RGS7 and RGS9-2 play essential role in vision, reward processing, and movement control. Both RGS7 and RGS9-2 belong to the R7 subfamily of RGS proteins that form macromolecular complexes with R7-binding protein (R7BP). R7BP targets RGS proteins to the plasma membrane and augments their GTPase-accelerating protein (GAP) activity, ultimately accelerating deactivation of G protein signaling. However, it remains unclear if R7BP serves exclusively as a membrane anchoring subunit or further modulates RGS proteins to increase their GAP activity. To directly answer this question, we utilized a rapidly reversible chemically induced protein dimerization system that enabled us to control RGS localization independent from R7BP in living cells. To monitor kinetics of Gα deactivation, we coupled this strategy with measuring changes in the GAP activity by bioluminescence resonance energy transfer-based assay in a cellular system containing μ-opioid receptor. This approach was used to correlate changes in RGS localization and activity in the presence or absence of R7BP. Strikingly, we observed that RGS activity is augmented by membrane recruitment, in an orientation independent manner with no additional contributions provided by R7BP. These findings argue that the association of R7 RGS proteins with the membrane environment provides a major direct contribution to modulation of their GAP activity. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

IL-7 Induces an Epitope Masking of γc Protein in IL-7 Receptor Signaling Complex

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Tae Sik Goh

2017-01-01

Full Text Available IL-7 signaling via IL-7Rα and common γ-chain (γc is necessary for the development and homeostasis of T cells. Although the delicate mechanism in which IL-7Rα downregulation allows the homeostasis of T cell with limited IL-7 has been well known, the exact mechanism behind the interaction between IL-7Rα and γc in the absence or presence of IL-7 remains unclear. Additionally, we are still uncertain as to how only IL-7Rα is separately downregulated by the binding of IL-7 from the IL-7Rα/γc complex. We demonstrate here that 4G3, TUGm2, and 3E12 epitope masking of γc protein are induced in the presence of IL-7, indicating that the epitope alteration is induced by IL-7 binding to the preassembled receptor core. Moreover, the epitope masking of γc protein is inversely correlated with the expression of IL-7Rα upon IL-7 binding, implying that the structural alteration of γc might be involved in the regulation of IL-7Rα expression. The conformational change in γc upon IL-7 binding may contribute not only to forming the functional IL-7 signaling complex but also to optimally regulating the expression of IL-7Rα.

IL-7 Induces an Epitope Masking of γc Protein in IL-7 Receptor Signaling Complex

Science.gov (United States)

Goh, Tae Sik; Jo, Yuna; Lee, Byunghyuk; Kim, Geona; Hwang, Hyunju; Ko, Eunhee; Kang, Seung Wan; Oh, Sae-Ock; Baek, Sun-Yong; Yoon, Sik; Lee, Jung Sub

2017-01-01

IL-7 signaling via IL-7Rα and common γ-chain (γc) is necessary for the development and homeostasis of T cells. Although the delicate mechanism in which IL-7Rα downregulation allows the homeostasis of T cell with limited IL-7 has been well known, the exact mechanism behind the interaction between IL-7Rα and γc in the absence or presence of IL-7 remains unclear. Additionally, we are still uncertain as to how only IL-7Rα is separately downregulated by the binding of IL-7 from the IL-7Rα/γc complex. We demonstrate here that 4G3, TUGm2, and 3E12 epitope masking of γc protein are induced in the presence of IL-7, indicating that the epitope alteration is induced by IL-7 binding to the preassembled receptor core. Moreover, the epitope masking of γc protein is inversely correlated with the expression of IL-7Rα upon IL-7 binding, implying that the structural alteration of γc might be involved in the regulation of IL-7Rα expression. The conformational change in γc upon IL-7 binding may contribute not only to forming the functional IL-7 signaling complex but also to optimally regulating the expression of IL-7Rα. PMID:28127156

Structure of the Aeropyrum pernix L7Ae multifunctional protein and insight into its extreme thermostability

International Nuclear Information System (INIS)

Bhuiya, Mohammad Wadud; Suryadi, Jimmy; Zhou, Zholi; Brown, Bernard Andrew II

2013-01-01

The crystal structure of A. pernix L7Ae is reported, providing insight into the extreme thermostability of this protein. Archaeal ribosomal protein L7Ae is a multifunctional RNA-binding protein that directs post-transcriptional modification of archaeal RNAs. The L7Ae protein from Aeropyrum pernix (Ap L7Ae), a member of the Crenarchaea, was found to have an extremely high melting temperature (>383 K). The crystal structure of Ap L7Ae has been determined to a resolution of 1.56 Å. The structure of Ap L7Ae was compared with the structures of two homologs: hyperthermophilic Methanocaldococcus jannaschii L7Ae and the mesophilic counterpart mammalian 15.5 kD protein. The primary stabilizing feature in the Ap L7Ae protein appears to be the large number of ion pairs and extensive ion-pair network that connects secondary-structural elements. To our knowledge, Ap L7Ae is among the most thermostable single-domain monomeric proteins presently observed

Identification of salivary mucin MUC7 binding proteins from Streptococcus gordonii

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Thornton David J

2009-08-01

Full Text Available Abstract Background The salivary mucin MUC7 (previously known as MG2 can adhere to various strains of streptococci that are primary colonizers and predominant microorganisms of the oral cavity. Although there is a growing interest in interaction between oral pathogens and salivary mucins, studies reporting the specific binding sites on the bacteria are rather limited. Identification and characterization of the specific interacting proteins on the bacterial cell surface, termed adhesins, are crucial to further understand host-pathogen interactions. Results We demonstrate here, using purified MUC7 to overlay blots of SDS-extracts of Streptococcus gordonii cell surface proteins, 4 MUC7-binding bands, with apparent molecular masses of 62, 78, 84 and 133 kDa from the Streptococcus gordonii strain, PK488. Putative adhesins were identified by in-gel digestion and subsequent nanoLC-tandem mass spectrometry analysis of resultant peptides. The 62 kDa and 84 kDa bands were identified as elongation factor (EF Tu and EF-G respectively. The 78 kDa band was a hppA gene product; the 74 kDa oligopeptide-binding lipoprotein. The 133 kDa band contained two proteins; alpha enolase and DNA-directed RNA polymerase, beta' subunit. Some of these proteins, for example alpha enolase are expected to be intracellular, however, flow cytometric analysis confirmed its location on the bacterial surface. Conclusion Our data demonstrated that S. gordonii expressed a number of putative MUC7 recognizing proteins and these contribute to MUC7 mucin binding of this streptococcal strain.

Identification of karyopherin α1 and α7 interacting proteins in porcine tissue.

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Ki-Eun Park

Full Text Available Specialized trafficking systems in eukaryotic cells serve a critical role in partitioning intracellular proteins between the nucleus and cytoplasm. Cytoplasmic proteins (including chromatin remodeling enzymes and transcription factors must gain access to the nucleus to exert their functions to properly program fundamental cellular events ranging from cell cycle progression to gene transcription. Knowing that nuclear import mediated by members of the karyopherin α family of transport receptors plays a critical role in regulating development and differentiation, we wanted to determine the identity of proteins that are trafficked by this karyopherin α pathway. To this end, we performed a GST pull-down assay using porcine orthologs of karyopherin α1 (KPNA1 and karyopherin α7 (KPNA7 and prey protein derived from porcine fibroblast cells and used a liquid chromatography and tandem mass spectrometry (LC-MS/MS approach to determine the identity of KPNA1 and KPNA7 interacting proteins. Our screen revealed that the proteins that interact with KPNA1 and KPNA7 are generally nuclear proteins that possess nuclear localization signals. We further validated two candidate proteins from this screen and showed that they are able to be imported into the nucleus in vivo and also interact with members of the karyopherin α family of proteins in vitro. Our results also reveal the utility of using a GST pull-down approach coupled with LC-MS/MS to screen for protein interaction partners in a non-traditional model system.

Insulating process for HT-7U central solenoid model coils

International Nuclear Information System (INIS)

Cui Yimin; Pan Wanjiang; Wu Songtao; Wan Yuanxi

2003-01-01

The HT-7U superconducting Tokamak is a whole superconducting magnetically confined fusion device. The insulating system of its central solenoid coils is critical to its properties. In this paper the forming of the insulating system and the vacuum-pressure-impregnating (VPI) are introduced, and the whole insulating process is verified under the super-conducting experiment condition

Crystal structures of the ligand-binding region of uPARAP

DEFF Research Database (Denmark)

Yuan, Cai; Jürgensen, Henrik J; Engelholm, Lars H

2016-01-01

The proteins of the mannose receptor (MR) family share a common domain organization and have a broad range of biological functions. Urokinase plasminogen activator receptor-associated protein (uPARAP) (or Endo180) is a member of this family and plays an important role in extracellular matrix...... remodelling through interaction with its ligands, including collagens and urokinase plasminogen activator receptor (uPAR). We report the crystal structures of the first four domains of uPARAP (also named the ligand-binding region, LBR) at pH 7.4 in Ca(2+)-bound and Ca(2+)-free forms. The first domain....... These LLRs undergo a Ca(2+)-dependent conformational change, and this is likely to be the key structural determinant affecting the overall conformation of uPARAP. Our results provide a molecular mechanism to support the structural flexibility of uPARAP, and shed light on the structural flexibility of other...

In vivo inhibition of incorporation of (U-/sup 14/C)glucose into proteins in experimental focal epilepsy

Energy Technology Data Exchange (ETDEWEB)

Coutinho-Netto, J.; Boyar, M.M.; Abdul-Ghani, A.S.; Bradford, H.F.

1982-08-01

The in vivo incorporation of (/sup 14/C) from (U-/sup 14/C)-glucose into rat brain proteins from different cortical areas was examined in three different experimental focal epilepsies: cobalt, freeze-lesions, and tityustoxin. When (U-/sup 14/C)-glucose was injected intraperitoneally into awake and unrestrained animals with marked signs of epileptic hyperactivity, the inhibition of incorporation of (/sup 14/C)-amino acids into trichloracetic acid (TCA)-insoluble proteins was highest in the focal (sensorimotor) area when compared with distant regions (approx. 60%), but less when compared with the contralateral (sensorimotor) region (approx. 23%). Greatly decreased incorporation caused by both cobalt and freeze-lesion-induced epilepsies was also observed in the contralateral area when a comparison was made with distant regions (approx. 50%), but there were no significant differences in protein-specific radioactivity between the different distant areas.

Effect of the Zr elements with thermal properties changes of U-7Mo-xZr/Al dispersion fuel

International Nuclear Information System (INIS)

Supardjo; Agoeng Kadarjono; Boybul; Aslina Br Ginting

2016-01-01

Thermal properties data of nuclear fuel is required as input data to predict material properties change phenomenon during the fabrication process and irradiated in a nuclear reactor. Study the influence of Zr element in the U-7Mo-xZr/Al (x = 1%, 2% and 3%) fuel dispersion to changes in the thermal properties at various temperatures have been stiffened. Thermal analysis includes determining the melting temperature, enthalpy, and phase changes made using Differential Thermal Analysis (DTA) in the temperature range between 30 °C up to 1400 °C, while the heat capacity of U-7Mo-xZr alloy and U-7Mo-xZr/Al dispersion fuel using Differential Scanning Calorimeter (DSC) at room temperature up to 450 °C. Thermal analyst data DTA shows that Zr levels of all three compositions showed a similar phenomenon. At temperatures between 565.60 °C - 584.98 °C change becomes α + δ to α + γ phase and at 649.22 °C – 650.13 °C happen smelting Al matrix Occur followed by a reaction between Al matrix with U-7Mo-xZr on 670.38 °C - 673.38 °C form U (Al, Mo)x Zr. Furthermore a phase change α + β becomes β + γ Occurs at temperatures 762.08 °C - 776.33 °C and diffusion between the matrix by U-7Mo-xZr/Al on 853.55 °C - 875.20 °C. Every phenomenon that Occurs, enthalpy posed a relative stable. Consolidation of uranium Occur in 1052.42 °C - 1104.99 °C and decomposition reaction of U (Al, Mo)x and U (Al, Zr)_x becomes (UAl_4, UAl_3, UAl_2), U-Mo, and UZr on 1328,34 °C - 1332,06 °C , The existence of Zr in U-Mo alloy increases the heat capacity of the U-7Mo-xZr/Al, dispersion fuel and the higher heat capacity of Zr levels increased due to interactions between the atoms of Zr with Al matrix so that the heat absorbed by the fuel increase. (author)

7 CFR 51.1176 - U.S. Grade AA Juice (Double A).

Science.gov (United States)

2010-01-01

... STANDARDS) United States Standards for Grades of Florida Oranges and Tangelos Standards for Internal Quality... 7 Agriculture 2 2010-01-01 2010-01-01 false U.S. Grade AA Juice (Double A). 51.1176 Section 51.1176 Agriculture Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards...

Accuracy issues involved in modeling in vivo protein structures using PM7.

Science.gov (United States)

Martin, Benjamin P; Brandon, Christopher J; Stewart, James J P; Braun-Sand, Sonja B

2015-08-01

Using the semiempirical method PM7, an attempt has been made to quantify the error in prediction of the in vivo structure of proteins relative to X-ray structures. Three important contributory factors are the experimental limitations of X-ray structures, the difference between the crystal and solution environments, and the errors due to PM7. The geometries of 19 proteins from the Protein Data Bank that had small R values, that is, high accuracy structures, were optimized and the resulting drop in heat of formation was calculated. Analysis of the changes showed that about 10% of this decrease in heat of formation was caused by faults in PM7, the balance being attributable to the X-ray structure and the difference between the crystal and solution environments. A previously unknown fault in PM7 was revealed during tests to validate the geometries generated using PM7. Clashscores generated by the Molprobity molecular mechanics structure validation program showed that PM7 was predicting unrealistically close contacts between nonbonding atoms in regions where the local geometry is dominated by very weak noncovalent interactions. The origin of this fault was traced to an underestimation of the core-core repulsion between atoms at distances smaller than the equilibrium distance. © 2015 The Authors. Proteins: Structure, Function, and Bioinformatics Published By Wiley Periodicals, Inc.

Overexpression and purification of U24 from human herpesvirus type-6 in E. coli: unconventional use of oxidizing environments with a maltose binding protein-hexahistine dual tag to enhance membrane protein yield

Directory of Open Access Journals (Sweden)

Straus Suzana K

2011-06-01

Full Text Available Abstract Background Obtaining membrane proteins in sufficient quantity for biophysical study and biotechnological applications has been a difficult task. Use of the maltose binding protein/hexahistidine dual tag system with E.coli as an expression host is emerging as a high throughput method to enhance membrane protein yield, solubility, and purity, but fails to be effective for certain proteins. Optimizing the variables in this system to fine-tune for efficiency can ultimately be a daunting task. To identify factors critical to success in this expression system, we have selected to study U24, a novel membrane protein from Human Herpesvirus type-6 with potent immunosuppressive ability and a possible role in the pathogenesis of the disease multiple sclerosis. Results We expressed full-length U24 as a C-terminal fusion to a maltose binding protein/hexahistidine tag and examined the effects of temperature, growth medium type, cell strain type, oxidizing vs. reducing conditions and periplasmic vs. cytoplasmic expression location. Temperature appeared to have the greatest effect on yield; at 37°C full-length protein was either poorly expressed (periplasm or degraded (cytoplasm whereas at 18°C, expression was improved especially in the periplasm of C41(DE3 cells and in the cytoplasm of oxidizing Δtrx/Δgor mutant strains, Origami 2 and SHuffle. Expression of the fusion protein in these strains were estimated to be 3.2, 5.3 and 4.3 times greater, respectively, compared to commonly-used BL21(DE3 cells. We found that U24 is isolated with an intramolecular disulfide bond under these conditions, and we probed whether this disulfide bond was critical to high yield expression of full-length protein. Expression analysis of a C21SC37S cysteine-free mutant U24 demonstrated that this disulfide was not critical for full-length protein expression, but it is more likely that strained metabolic conditions favour factors which promote protein expression. This

Thermal properties of KUO3(s) and K2U2O7 - by high temperature Calvet calorimeter

International Nuclear Information System (INIS)

Jayanthi, K.; Iyer, V.S.; Venugopal, V.

1998-01-01

The thermal properties of KUO 3 (s) and K 2 U 2 O 7 (s) were determined using a high temperature Calvet calorimeter by drop method. The enthalpy increments, (H T o - H 298.15 0 ), in kJ/mol for KUO 3 (s) and K 2 U 2 O 7 (s) can be represented by, H T o - H 298.15 0 KUO 3 (s) kJ/mol ± 0.7 = -39.15 + 0.129T + 0.1005x10 -4 T 2 (369-714K) and H T o -H 298.15 0 K 2 U 2 O 7 (s) kJ/mol ± 0.7 = -52.99 + 0.1361T + 0.146x10 -3 T 2 (391 - 683K). (author)

Interaction of the iron–sulfur cluster assembly protein IscU with the Hsc66/Hsc20 molecular chaperone system of Escherichia coli

Science.gov (United States)

Hoff, Kevin G.; Silberg, Jonathan J.; Vickery, Larry E.

2000-01-01

The iscU gene in bacteria is located in a gene cluster encoding proteins implicated in iron–sulfur cluster assembly and an hsc70-type (heat shock cognate) molecular chaperone system, iscSUA-hscBA. To investigate possible interactions between these systems, we have overproduced and purified the IscU protein from Escherichia coli and have studied its interactions with the hscA and hscB gene products Hsc66 and Hsc20. IscU and its iron–sulfur complex (IscU–Fe/S) stimulated the basal steady-state ATPase activity of Hsc66 weakly in the absence of Hsc20 but, in the presence of Hsc20, increased the ATPase activity up to 480-fold. Hsc20 also decreased the apparent Km for IscU stimulation of Hsc66 ATPase activity, and surface plasmon resonance studies revealed that Hsc20 enhances binding of IscU to Hsc66. Surface plasmon resonance and isothermal titration calorimetry further showed that IscU and Hsc20 form a complex, and Hsc20 may thereby aid in the targeting of IscU to Hsc66. These results establish a direct and specific role for the Hsc66/Hsc20 chaperone system in functioning with isc gene components for the assembly of iron–sulfur cluster proteins. PMID:10869428

Localization of PDZD7 to the stereocilia ankle-link associates this scaffolding protein with the Usher syndrome protein network.

Science.gov (United States)

Grati, M'hamed; Shin, Jung-Bum; Weston, Michael D; Green, James; Bhat, Manzoor A; Gillespie, Peter G; Kachar, Bechara

2012-10-10

Usher syndrome is the leading cause of genetic deaf-blindness. Monoallelic mutations in PDZD7 increase the severity of Usher type II syndrome caused by mutations in USH2A and GPR98, which respectively encode usherin and GPR98. PDZ domain-containing 7 protein (PDZD7) is a paralog of the scaffolding proteins harmonin and whirlin, which are implicated in Usher type 1 and type 2 syndromes. While usherin and GPR98 have been reported to form hair cell stereocilia ankle-links, harmonin localizes to the stereocilia upper tip-link density and whirlin localizes to both tip and ankle-link regions. Here, we used mass spectrometry to show that PDZD7 is expressed in chick stereocilia at a comparable molecular abundance to GPR98. We also show by immunofluorescence and by overexpression of tagged proteins in rat and mouse hair cells that PDZD7 localizes to the ankle-link region, overlapping with usherin, whirlin, and GPR98. Finally, we show in LLC-PK1 cells that cytosolic domains of usherin and GPR98 can bind to both whirlin and PDZD7. These observations are consistent with PDZD7 being a modifier and candidate gene for USH2, and suggest that PDZD7 is a second scaffolding component of the ankle-link complex.

Heterogeneous genetic diversity pattern in Plasmodium vivax genes encoding merozoite surface proteins (MSP) -7E, -7F and -7L.

Science.gov (United States)

Garzón-Ospina, Diego; Forero-Rodríguez, Johanna; Patarroyo, Manuel A

2014-12-13

The msp-7 gene has become differentially expanded in the Plasmodium genus; Plasmodium vivax has the highest copy number of this gene, several of which encode antigenic proteins in merozoites. DNA sequences from thirty-six Colombian clinical isolates from P. vivax (pv) msp-7E, -7F and -7L genes were analysed for characterizing and studying the genetic diversity of these pvmsp-7 members which are expressed during the intra-erythrocyte stage; natural selection signals producing the variation pattern so observed were evaluated. The pvmsp-7E gene was highly polymorphic compared to pvmsp-7F and pvmsp-7L which were seen to have limited genetic diversity; pvmsp-7E polymorphism was seen to have been maintained by different types of positive selection. Even though these copies seemed to be species-specific duplications, a search in the Plasmodium cynomolgi genome (P. vivax sister taxon) showed that both species shared the whole msp-7 repertoire. This led to exploring the long-term effect of natural selection by comparing the orthologous sequences which led to finding signatures for lineage-specific positive selection. The results confirmed that the P. vivax msp-7 family has a heterogeneous genetic diversity pattern; some members are highly conserved whilst others are highly diverse. The results suggested that the 3'-end of these genes encode MSP-7 proteins' functional region whilst the central region of pvmsp-7E has evolved rapidly. The lineage-specific positive selection signals found suggested that mutations occurring in msp-7s genes during host switch may have succeeded in adapting the ancestral P. vivax parasite population to humans.

Regulation of neurite morphogenesis by interaction between R7 regulator of G protein signaling complexes and G protein subunit Gα13.

Science.gov (United States)

Scherer, Stephanie L; Cain, Matthew D; Kanai, Stanley M; Kaltenbronn, Kevin M; Blumer, Kendall J

2017-06-16

The R7 regulator of G protein signaling family (R7-RGS) critically regulates nervous system development and function. Mice lacking all R7-RGS subtypes exhibit diverse neurological phenotypes, and humans bearing mutations in the retinal R7-RGS isoform RGS9-1 have vision deficits. Although each R7-RGS subtype forms heterotrimeric complexes with Gβ 5 and R7-RGS-binding protein (R7BP) that regulate G protein-coupled receptor signaling by accelerating deactivation of G i/o α-subunits, several neurological phenotypes of R7-RGS knock-out mice are not readily explained by dysregulated G i/o signaling. Accordingly, we used tandem affinity purification and LC-MS/MS to search for novel proteins that interact with R7-RGS heterotrimers in the mouse brain. Among several proteins detected, we focused on Gα 13 because it had not been linked to R7-RGS complexes before. Split-luciferase complementation assays indicated that Gα 13 in its active or inactive state interacts with R7-RGS heterotrimers containing any R7-RGS isoform. LARG (leukemia-associated Rho guanine nucleotide exchange factor (GEF)), PDZ-RhoGEF, and p115RhoGEF augmented interaction between activated Gα 13 and R7-RGS heterotrimers, indicating that these effector RhoGEFs can engage Gα 13 ·R7-RGS complexes. Because Gα 13 /R7-RGS interaction required R7BP, we analyzed phenotypes of neuronal cell lines expressing RGS7 and Gβ 5 with or without R7BP. We found that neurite retraction evoked by Gα 12/13 -dependent lysophosphatidic acid receptors was augmented in R7BP-expressing cells. R7BP expression blunted neurite formation evoked by serum starvation by signaling mechanisms involving Gα 12/13 but not Gα i/o These findings provide the first evidence that R7-RGS heterotrimers interact with Gα 13 to augment signaling pathways that regulate neurite morphogenesis. This mechanism expands the diversity of functions whereby R7-RGS complexes regulate critical aspects of nervous system development and function. © 2017 by

Biochemical and structural characterization of Cren7, a novel chromatin protein conserved among Crenarchaea

OpenAIRE

Guo, Li; Feng, Yingang; Zhang, Zhenfeng; Yao, Hongwei; Luo, Yuanming; Wang, Jinfeng; Huang, Li

2007-01-01

Archaea contain a variety of chromatin proteins consistent with the evolution of different genome packaging mechanisms. Among the two main kingdoms in the Archaea, Euryarchaeota synthesize histone homologs, whereas Crenarchaeota have not been shown to possess a chromatin protein conserved at the kingdom level. We report the identification of Cren7, a novel family of chromatin proteins highly conserved in the Crenarchaeota. A small, basic, methylated and abundant protein, Cren7 displays a high...

Potential proteins targeted by let-7f-5p in HeLa cells.

Science.gov (United States)

Wang, Yu; Chen, Xiujuan; Zhang, Yi; Song, Jiandong

2017-07-24

MicroRNAs are a class of small, endogenous, non-coding RNAs mediating posttranscriptional gene silencing. The current authors hypothesized that let-7f-5p is likely involved in cell invasion and proliferation by regulating the expression of target genes. The current study combined let-7f-5p with iTRAQ to assess its effect on gene expression in HeLa cells. Results indicated that 164 proteins were expressed at different levels in HeLa cells overexpressing let-7f-5p and negative controls and that 172 proteins were expressed at different levels in let-7f-5p-silenced HeLa cells and negative controls. Results indicated that let-7f-5p may suppress insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1) in HeLa cells.

Human herpesvirus 6B U19 protein is a PML-regulated transcriptional activator that localizes to nuclear foci in a PML-independent manner

DEFF Research Database (Denmark)

Kofod-Olsen, Emil; Ross-Hansen, Katrine; Mikkelsen, Jacob Giehm

2008-01-01

Human herpesvirus 6B (HHV-6B) contains an IE-B domain spanning open reading frames U16/17-U19, based on homology with human cytomegalovirus. Here, the protein product, U19, of the HHV-6B U19 gene is identified as a 47 kDa transcriptional activator. HHV-6B infection or overexpression of U19...... transactivated the RANTES promoter. Mutational analysis of the promoter indicated that transactivation was not critically dependent on the promoter sites CRE, NF-kappaB, ISRE or NF-IL6. ND10 are nuclear substructures that are involved in several cellular regulatory pathways, including those controlling gene...... structure, U19 also localized to the centre of ND10. Knockdown of PML by small interfering RNA did not prevent U19 localization to ND10-like foci, but instead led to a fourfold increase in U19-induced transcription from the RANTES promoter. Generation of four truncated U19 proteins indicated that the N...

Manufacture of the rectifier of the HT-7U PFPS

International Nuclear Information System (INIS)

Gao Ge; Fu Peng; Tang Lunjun; Wang Linsen

2005-01-01

The rectifiers of the HT-7U poloidal field power supply (PFPS) are introduced. A new control method, four quadrants converter, is brought forward, which overcomes the short-coming of both the circulating current mode and the non-circulating current mode. This control mode also resolves the problem of DC circulating current in the identical phase anti-parallel connection rectifiers when these rectifiers run in the circulating current mode. (authors)

Design and realization on function of pre-forming and continuous winding for HT-7U special winding machine

International Nuclear Information System (INIS)

Yu Jie; Gao Daming; Wen Jun; Zhu Wenhua; Cheng Leping; Tao Yuming

2000-05-01

The winding machine is one of the critical facilities for R and D of HT-7U construction. The machine mainly consists of five parts, CICC pay-off spool, a four-rollers straightening assembly, a four-roller forming/bending assembly, continuous winding structure and CNC control system with three-axis CNC control. The facility is needed for CICC magnet fabrication of HT-7U. The main requirements of the winding machine are: continuous winding to reduce number of joints inside the coils; pre-forming CICC conductor to avoid winding with tension; suitable for all TF and PF coils within the scope of various coil shape and dimension limit; improving the configuration tolerance, specially flatness of the CICC conductor. The author emphasizes on the design and realization on function of Pre-forming and Continuous Winding for HT-7U special winding machine. The winding machine with high accuracy has just been developed and applied to the construction of HT-7U model coils

Efficiency of a solid-phase chemiluminescence immunoassay for detection of antinuclear and cytoplasmic autoantibodies compared with gold standard immunoprecipitation.

Science.gov (United States)

Gelpí, Carmen; Pérez, Elena; Roldan, Cristina

2014-09-01

The aim of this study was to compare the degree of agreement of a novel Zenit RA chemiluminescent immunoassay (CLIA) from A. Menarini Diagnostics (Florence, Italy) and the gold standard immunoprecipitation assay to screen for the presence of specific anti-U1snRNP, anti-Sm, anti-Ro/SS-A, anti-La/SS-B, anti-Jo-1((his)tRNA-Synthetase) and anti-Scl-70(Topo I) antibodies. We studied 114 sera, 98 from patients with well-defined autoimmune connective tissue diseases and 16 from blood donor volunteers. All samples were fully characterized using the new chemiluminescent immunoassay and immunoprecipitation. In addition, all the samples were analyzed by indirect immunofluorescence (IIF) and anti-Scl-70(Topo I) antibodies were analyzed by immunoblot (IB) assay. Discrepant samples were analyzed using a commercial dot blot technique (Recomline from Mikrogen). The simple Kappa coefficient was used to measure the level of agreement between the results of Zenit RA CLIA and the gold standard. The Kappa agreement between Zenit RA CLIA and gold standard immunoprecipitation, as well as IB and IIFassays for the presence of anti-Scl-70(Topo I)(0.948) was excellent. The concordance between Zenit RA CLIA and gold standard immunoprecipitation for the presence of anti-U1snRNP (0.883), anti-Ro/SS-A (0.878), anti-Jo-1((his)tRNA-Synthetase) (0.791) and anti-Sm (0.786) was good, and excellent when the cut-off was raised to 14 U/ml (arbitrary units/ml). Between Zenit RA CLIA and gold standard immunoprecipitation for the presence of anti-La/SS-B, the Kappa agreement had a value of 0.689, but this improved to 0.775 when the cut-off was raised to14 U/ml. Precision was good based on the evaluation of replicate samples. Inter-assay coefficient variation was lower than 3.4 % (CV in %) in all the kits and <1.2 % (CV in  %) for intra-assay measurements. Our findings show that Zenit RA CLIA was specific and sensitive to detect anti-U1snRNP, anti-Sm, anti-Ro/SS-A, anti-La/SS-B, anti-Jo-1((his

Human papillomavirus (HPV) type 16 E7 protein bodies cause tumour regression in mice

International Nuclear Information System (INIS)

Whitehead, Mark; Öhlschläger, Peter; Almajhdi, Fahad N; Alloza, Leonor; Marzábal, Pablo; Meyers, Ann E; Hitzeroth, Inga I; Rybicki, Edward P

2014-01-01

Human papillomaviruses (HPV) are the causative agents of cervical cancer in women, which results in over 250 000 deaths per year. Presently there are two prophylactic vaccines on the market, protecting against the two most common high-risk HPV types 16 and 18. These vaccines remain very expensive and are not generally affordable in developing countries where they are needed most. Additionally, there remains a need to treat women that are already infected with HPV, and who have high-grade lesions or cervical cancer. In this paper, we characterize the immunogenicity of a therapeutic vaccine that targets the E7 protein of the most prevalent high-risk HPV - type 16 – the gene which has previously been shown to be effective in DNA vaccine trials in mice. The synthetic shuffled HPV-16 E7 (16E7SH) has lost its transforming properties but retains all naturally-occurring CTL epitopes. This was genetically fused to Zera®, a self-assembly domain of the maize γ-zein able to induce the accumulation of recombinant proteins into protein bodies (PBs), within the endoplasmic reticulum in a number of expression systems. High-level expression of the HPV 16E7SH protein fused to Zera® in plants was achieved, and the protein bodies could be easily and cost-effectively purified. Immune responses comparable to the 16E7SH DNA vaccine were demonstrated in the murine model, with the protein vaccine successfully inducing a specific humoral as well as cell mediated immune response, and mediating tumour regression. The fusion of 16E7SH to the Zera® peptide was found to enhance the immune responses, presumably by means of a more efficient antigen presentation via the protein bodies. Interestingly, simply mixing the free PBs and 16E7SH also enhanced immune responses, indicating an adjuvant activity for the Zera® PBs

AMP-activated protein kinase downregulates Kv7.1 cell surface expression

DEFF Research Database (Denmark)

Andersen, Martin N; Krzystanek, Katarzyna; Jespersen, Thomas

2012-01-01

in response to polarization of the epithelial Madin-Darby canine kidney (MDCK) cell line and that this was mediated by activation of protein kinase C (PKC). In this study, the pathway downstream of PKC, which leads to internalization of Kv7.1 upon cell polarization, is elucidated. We show by confocal...... microscopy that Kv7.1 is endocytosed upon initiation of the polarization process and sent for degradation by the lysosomal pathway. The internalization could be mimicked by pharmacological activation of the AMP-activated protein kinase (AMPK) using three different AMPK activators. We demonstrate...

Human herpesvirus 6B U19 protein is a PML-regulated transcriptional activator that localizes to nuclear foci in a PML-independent manner

DEFF Research Database (Denmark)

Kofod-Olsen, Emil; Ross-Hansen, Katrine; Mikkelsen, Jacob Giehm

2008-01-01

Human herpesvirus 6B (HHV-6B) contains an IE-B domain spanning open reading frames U16/17-U19, based on homology with human cytomegalovirus. Here, the protein product, U19, of the HHV-6B U19 gene is identified as a 47 kDa transcriptional activator. HHV-6B infection or overexpression of U19...

How to find the optimal partner--studies of snurportin 1 interactions with U snRNA 5' TMG-cap analogues containing modified 2-amino group of 7-methylguanosine.

Science.gov (United States)

Piecyk, Karolina; Niedzwiecka, Anna; Ferenc-Mrozek, Aleksandra; Lukaszewicz, Maciej; Darzynkiewicz, Edward; Jankowska-Anyszka, Marzena

2015-08-01

Snurportin 1 is an adaptor protein that mediates the active nuclear import of uridine-rich small nuclear RNAs (U snRNA) by the importin-β receptor pathway. Its cellular activity influences the overall transport yield of small ribonucleoprotein complexes containing N(2),N(2),7-trimethylguanosine (TMG) capped U snRNA. So far little is still known about structural requirements related to molecular recognition of the trimethylguanosine moiety by snurportin in solution. Since these interactions are of a great biomedical importance, we synthesized a series of new 7-methylguanosine cap analogues with extended substituents at the exocyclic 2-amino group to gain a deeper insight into how the TMG-cap is adapted into the snurportin cap-binding pocket. Prepared chemical tools were applied in binding assays using emission spectroscopy. Surprisingly, our results revealed strict selectivity of snurportin towards the TMG-cap structure that relied mainly on its structural stiffness and compactness. Copyright © 2015 Elsevier Ltd. All rights reserved.

Removal of U(VI) from aqueous solutions using Shewanella sp. RCRI7, isolated from Qurugoel Lake in Iran

Energy Technology Data Exchange (ETDEWEB)

Abdehvand, Adib Zaheri; Keshtkar, Alireza; Fatemi, Faezeh [Nuclear Science and Technology Research Institute, Tehran (Iran, Islamic Republic of). Nuclear Fuel Cycle Research School; Tarhiz, Vahideh; Hejazi, Mohammad Saeid [Tabriz Univ. of Medical Sciences (Iran, Islamic Republic of). Molecular Medicine Research Center

2017-04-01

Isolation, genotypic and phenotypic characterization of an aqueous bacterium, Shewanella sp RCRI7, from Qurugoel Lake in Iran and uranium removal from aqueous solutions using the isolate is described. Based on 16S rRNA gene sequence analysis and phylogenetic tree, strain RCRI7{sup T} falls into genus Shewanella. Closely related type strains include Shewanella xiamenensis S4{sup T} KJ542801, Shewanella profunda DSM15900{sup T} FR733713, Shewanella putrefaciens LMG 26268{sup T} X81623 and Shewanella oneidensis MR-1{sup T} AE014299. Anaerobic incubation of the bacteria in the presence of U(VI) led to uranium removal from the solution and formation of a black precipitate. Analysis of the precipitate using UV-vis confirmed the reduction of U(VI) to U(IV). The effects of pH, temperature, U(VI) concentration and cell density on uranium removal were elucidated. The maximum uranium removal was 97%. As a conclusion, the findings revealed the ability of the local strain RCRI7 for U(VI) bioreduction as an effective bacterium for uranium immobilization.

Removal of U(VI) from aqueous solutions using Shewanella sp. RCRI7, isolated from Qurugoel Lake in Iran

International Nuclear Information System (INIS)

Abdehvand, Adib Zaheri; Keshtkar, Alireza; Fatemi, Faezeh; Tarhiz, Vahideh; Hejazi, Mohammad Saeid

2017-01-01

Isolation, genotypic and phenotypic characterization of an aqueous bacterium, Shewanella sp RCRI7, from Qurugoel Lake in Iran and uranium removal from aqueous solutions using the isolate is described. Based on 16S rRNA gene sequence analysis and phylogenetic tree, strain RCRI7 T falls into genus Shewanella. Closely related type strains include Shewanella xiamenensis S4 T KJ542801, Shewanella profunda DSM15900 T FR733713, Shewanella putrefaciens LMG 26268 T X81623 and Shewanella oneidensis MR-1 T AE014299. Anaerobic incubation of the bacteria in the presence of U(VI) led to uranium removal from the solution and formation of a black precipitate. Analysis of the precipitate using UV-vis confirmed the reduction of U(VI) to U(IV). The effects of pH, temperature, U(VI) concentration and cell density on uranium removal were elucidated. The maximum uranium removal was 97%. As a conclusion, the findings revealed the ability of the local strain RCRI7 for U(VI) bioreduction as an effective bacterium for uranium immobilization.

Evaluation of antibodies against feline coronavirus 7b protein for diagnosis of feline infectious peritonitis in cats.

Science.gov (United States)

Kennedy, Melissa A; Abd-Eldaim, Mohamed; Zika, Sarah E; Mankin, Joseph M; Kania, Stephen A

2008-09-01

To determine whether expression of feline coronavirus (FCoV) 7b protein, as indicated by the presence of specific serum antibodies, consistently correlated with occurrence of feline infectious peritonitis (FIP) in cats. 95 serum samples submitted for various diagnostic assays and 20 samples from specific-pathogen-free cats tested as negative control samples. The 7b gene from a virulent strain of FCoV was cloned into a protein expression vector. The resultant recombinant protein was produced and used in antibody detection assays via western blot analysis of serum samples. Results were compared with those of an immunofluorescence assay (IFA) for FCoV-specific antibody and correlated with health status. Healthy IFA-seronegative cats were seronegative for antibodies against the 7b protein. Some healthy cats with detectable FCoV-specific antibodies as determined via IFA were seronegative for antibodies against the 7b protein. Serum from cats with FIP had antibodies against the 7b protein, including cats with negative results via conventional IFA. However, some healthy cats, as well as cats with conditions other than FIP that were seropositive to FCoV via IFA, were also seropositive for the 7b protein. Expression of the 7b protein, as indicated by detection of antibodies against the protein, was found in most FCoV-infected cats. Seropositivity for this protein was not specific for the FCoV virulent biotype or a diagnosis of FIP.

Rice MEL2, the RNA recognition motif (RRM) protein, binds in vitro to meiosis-expressed genes containing U-rich RNA consensus sequences in the 3'-UTR.

Science.gov (United States)

Miyazaki, Saori; Sato, Yutaka; Asano, Tomoya; Nagamura, Yoshiaki; Nonomura, Ken-Ichi

2015-10-01

Post-transcriptional gene regulation by RNA recognition motif (RRM) proteins through binding to cis-elements in the 3'-untranslated region (3'-UTR) is widely used in eukaryotes to complete various biological processes. Rice MEIOSIS ARRESTED AT LEPTOTENE2 (MEL2) is the RRM protein that functions in the transition to meiosis in proper timing. The MEL2 RRM preferentially associated with the U-rich RNA consensus, UUAGUU[U/A][U/G][A/U/G]U, dependently on sequences and proportionally to MEL2 protein amounts in vitro. The consensus sequences were located in the putative looped structures of the RNA ligand. A genome-wide survey revealed a tendency of MEL2-binding consensus appearing in 3'-UTR of rice genes. Of 249 genes that conserved the consensus in their 3'-UTR, 13 genes spatiotemporally co-expressed with MEL2 in meiotic flowers, and included several genes whose function was supposed in meiosis; such as Replication protein A and OsMADS3. The proteome analysis revealed that the amounts of small ubiquitin-related modifier-like protein and eukaryotic translation initiation factor3-like protein were dramatically altered in mel2 mutant anthers. Taken together with transcriptome and gene ontology results, we propose that the rice MEL2 is involved in the translational regulation of key meiotic genes on 3'-UTRs to achieve the faithful transition of germ cells to meiosis.

Amino Acid Substitutions Improve the Immunogenicity of H7N7HA Protein and Protect Mice against Lethal H7N7 Viral Challenge.

Directory of Open Access Journals (Sweden)

Subaschandrabose Rajesh Kumar

Full Text Available Avian influenza A H7N7/NL/219/03 virus creates a serious pandemic threat to human health because it can transmit directly from domestic poultry to humans and from human to human. Our previous vaccine study reported that mice when immunized intranasally (i.n with live Bac-HA were protected from lethal H7N7/NL/219/03 challenge, whereas incomplete protection was obtained when administered subcutaneously (s.c due to the fact that H7N7 is a poor inducer of neutralizing antibodies. Interestingly, our recent vaccine studies reported that mice when vaccinated subcutaneously with Bac-HA (H7N9 was protected against both H7N9 (A/Sh2/2013 and H7N7 virus challenge. HA1 region of both H7N7 and H7N9 viruses are differ at 15 amino acid positions. Among those, we selected three amino acid positions (T143, T198 and I211 in HA1 region of H7N7. These amino acids are located within or near the receptor binding site. Following the selection, we substituted the amino acid at these three positions with amino acids found on H7N9HA wild-type. In this study, we evaluate the impact of amino acid substitutions in the H7N7 HA-protein on the immunogenicity. We generated six mutant constructs from wild-type influenza H7N7HA cDNA by site directed mutagenesis, and individually expressed mutant HA protein on the surface of baculovirus (Bac-HAm and compared their protective efficacy of the vaccines with Bac-H7N7HA wild-type (Bac-HA by lethal H7N7 viral challenge in a mouse model. We found that mice immunized subcutaneously with Bac-HAm constructs T143A or T198A-I211V or I211V-T143A serum showed significantly higher hemagglutination inhibition and neutralization titer against H7N7 and H7N9 viruses when compared to Bac-HA vaccinated mice groups. We also observed low level of lung viral titer, negligible weight loss and complete protection against lethal H7N7 viral challenge. Our results indicated that amino acid substitution at position 143 or 211 improve immunogenicity of H7N7HA

TEM investigation of irradiated U-7 weight percent Mo dispersion fuel

International Nuclear Information System (INIS)

Van den Berghe, S.

2009-01-01

In the FUTURE experiment, fuel plates containing U-7 weight percent Mo atomized powder were irradiated in the BR2 reactor. At a burn-up of approximately 33 percent 235 U (6.5 percent FIMA or 1.41 10 21 fissions/cm 3 meat), the fuel plates showed an important deformation and the irradiation was stopped. The plates were submitted to detailed PIE at the Laboratory for High and Medium level Activity. The results of these examinations were reported in the scientific report of last year and published in open literature. Since then, the microstructural aspects of the FUTURE fuel were studied in more detail using transmission electron microscopy (TEM), in an attempt to understand the nature of the interaction phase and the fission gas behavior in the atomized U(Mo) fuel. The FUTURE experiment is regarded as the definitive proof that the classical atomized U(Mo) dispersion fuel is not stable under irradiation, at least in the conditions required for normal operation of plate-type fuel. The main cause for the instability was identified to be the irradiation behavior of the U(Mo)-Al interaction phase which is formed between the U(Mo) particles and the pure aluminum matrix during irradiation. It is assumed to become amorphous under irradiation and as such cannot retain the fission gas in stable bubbles. As a consequence, gas filled voids are generated between the interaction layer and the matrix, resulting in fuel plate pillowing and failure. The objective of the TEM investigation was the confirmation of this assumption of the amorphisation of the interaction phase. A deeper understanding of the actual nature of this layer and the fission gas behaviour in these fuels in general can allow a more oriented search for a solution to the fuel failures

Optical spectroscopy and crystal-field analysis of U3+: Ba2YCl7

International Nuclear Information System (INIS)

Karbowiak, M.; Mech, A.; Drozdzyndki, J.; Gajek, Z.; Edelstein, N.M.

2002-01-01

High resolution absorption spectra of a U 3+ (0.3%): Ba 2 YCl 7 single crystal were recorded in the 4000-50 000 cm -1 range at 7 K. The observed crystal-field levels were assigned and fit to the parameters of the simplified angular overlap model (AOM) as well as a semi-empirical Hamiltonian representing the combined atomic and one-electron crystal-field interactions. The starting values of the AOM parameters were obtained from ab initio calculations. The analysis of the spectra allowed the assignment of 65 crystal-field levels with a relatively small rms deviation of 25 cm -1 and has shown that the AOM approach can predict quite well the B q k crystal-field parameters. The value determined for the crystal-field strength parameter, N v , corresponds well with those determined for U 3+ in other chloride single crystals. (authors)

12 CFR 7.5009 - Location under 12 U.S.C. 85 of national banks operating exclusively through the Internet.

Science.gov (United States)

2010-01-01

... operating exclusively through the Internet. 7.5009 Section 7.5009 Banks and Banking COMPTROLLER OF THE... under 12 U.S.C. 85 of national banks operating exclusively through the Internet. For purposes of 12 U.S.C. 85, the main office of a national bank that operates exclusively through the Internet is the...

U-8 wt %Mo and 7 wt %Mo alloys powder obtained by an hydride-de hydride process; Obtencion de polvo de aleaciones U-8% Mo y U-7% Mo (en peso) mediante hidruracion

Energy Technology Data Exchange (ETDEWEB)

Balart, Silvia N; Bruzzoni, Pablo; Granovsky, Marta S; Gribaudo, Luis M.J.; Hermida, Jorge D; Ovejero, Jose; Rubiolo, Gerardo H; Vicente, Eduardo E [Comision Nacional de Energia Atomica, General San Martin (Argentina). Dept. de Materiales

2000-07-01

Uranium-molybdenum alloys are been tested as a component in high-density LEU dispersion fuels with very good performances. These alloys need to be transformed to powder due to the manufacturing requirements of the fuels. One method to convert ductile alloys into powder is the hydride-de hydride process, which takes advantage of the ability of the U-{alpha} phase to transform to UH{sub 3}: a brittle and relatively low-density compound. U-Mo alloys around 7 and 8 wt % Mo were melted and heat treated at different temperature ranges in order to partially convert {gamma} -phase to {alpha} -phase. Subsequent hydriding transforms this {alpha} -phase to UH{sub 3}. The volume change associated to the hydride formation embrittled the material which ends up in a powdered alloy. Results of the optical metallography, scanning electron microscopy, X-ray diffraction during different steps of the process are shown. (author)

Stress corrosion of the alloy U-7.5 Nb-2.5 Zr

International Nuclear Information System (INIS)

Lepoutre, D.; Nomine, A.M.; Miannay, D.

1983-09-01

Oxide formed on U-7.5 Nb-2.5 Zr at room temperature during stress corrosion cracking in oxygen is identical to the natural oxide of the alloy. It is formed by UO 2 with Nb and Zr and is associated with an increased Nb content at the interface. This oxide would be responsible for cracking [fr

Biochemical and structural characterization of Cren7, a novel chromatin protein conserved among Crenarchaea.

Science.gov (United States)

Guo, Li; Feng, Yingang; Zhang, Zhenfeng; Yao, Hongwei; Luo, Yuanming; Wang, Jinfeng; Huang, Li

2008-03-01

Archaea contain a variety of chromatin proteins consistent with the evolution of different genome packaging mechanisms. Among the two main kingdoms in the Archaea, Euryarchaeota synthesize histone homologs, whereas Crenarchaeota have not been shown to possess a chromatin protein conserved at the kingdom level. We report the identification of Cren7, a novel family of chromatin proteins highly conserved in the Crenarchaeota. A small, basic, methylated and abundant protein, Cren7 displays a higher affinity for double-stranded DNA than for single-stranded DNA, constrains negative DNA supercoils and is associated with genomic DNA in vivo. The solution structure and DNA-binding surface of Cren7 from the hyperthermophilic crenarchaeon Sulfolobus solfataricus were determined by NMR. The protein adopts an SH3-like fold. It interacts with duplex DNA through a beta-sheet and a long flexible loop, presumably resulting in DNA distortions through intercalation of conserved hydrophobic residues into the DNA structure. These data suggest that the crenarchaeal kingdom in the Archaea shares a common strategy in chromatin organization.

Design and performance of U7B beamline and X-ray diffraction and scattering station at NSRL and its preliminary experiments in protein crystallography

International Nuclear Information System (INIS)

Pan Guoqiang; Xu, Chaoyin; Fan Rong; Gao Chen; Lou Xiaohua; Teng Maikun; Huang Qingqiu; Niu Liwen

2005-01-01

This publication describes the design and performance of the U7B beamline and X-ray diffraction and diffuse scattering station at National Synchrotron Radiation Laboratory (NSRL). The beamline optics comprise a Pt-coated toroidal focusing mirror and a double-crystal Si(1 1 1) monochromator. A preliminary experiment of diffraction data collection and processing was carried out using a commercial imaging plate detector system (Mar345). The data collected from one single crystal of acutohaemolysin, a Lys49-type PLA2 from Agkistrodon acutus venom, are of high quality

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

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Jicheng Hu

Full Text Available 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.

Screening for proteins interacting with MCM7 in human lung cancer library using yeast two hybrid system

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Yuchen HAN

2008-08-01

Full Text Available Background and objective MCM7 is a subunit of the MCM complex that plays a key role in DNA replication initiation. But little is known about its interaction proteins. In this study yeast two hybrid screening was used to identify the MCM7 interacting proteins. Methods Yeast expression vector containing human full length MCM7-pGBKT7 plasmid was constructed, and with a library of cDNAs from human lung cancer-pACT2 plasmid was transformed into yeast strain AH109, and was electively grew in X-a-gal auxotrophy medium SD/-Trp-Leu-His-Ade, and the blue colonies were picked up, the plasmid of the yeast colonies was extracted , and transformed into E. Coli to extract DNA and performed sequence analysis. Results Eleven proteins were identified which could specifically interact with MCM7 proteins, among these five were cytoskeleton proteins, six were enzymes, kinases and related receptors. Conclusion The investigation provides functional clues for further exploration of MCM7 gene.

Functional analysis of U1-70K interacting SR proteins in pre-mRNA splicing in Arabidopsis

International Nuclear Information System (INIS)

Reddy, A.S.N.

2008-01-01

Proteins of a serine/arginine-rich (SR) family are part of the spliceosome and are implicated in both constitutive and alternative splicing of pre-mRNAs. With the funding from DOE we have been studying alternative of splicing of genes encoding serine/arginine-rich (SR) proteins and the roles of SR proteins that interact with U1-70K in regulating basic and alternative splicing. Alternative splicing of pre-mRNAs of Arabidopsis serine/arginine-rich proteins and its regulation by hormones and stresses: We analyzed the splicing of all 19 Arabidopsis genes in different tissues, during different seedling stages and in response to various hormonal and stress treatments. Remarkably, about 90 different transcripts are produced from 15 SR genes, thereby increasing the transcriptome complexity of SR genes by about five fold. Using the RNA isolated from polysomes we have shown that most of the splice variants are recruited for translation. Alternative splicing of some SR genes is controlled in a developmental and tissue-specific manner (Palusa et al., 2007). Interestingly, among the various hormones and abiotic stresses tested, temperature stress (cold and heat) and ultraviolet light dramatically altered alternative splicing of pre-mRNAs of several SR genes whereas hormones altered the splicing of only two SR genes (Palusa et al., 2007). Localization and dynamics of a novel serine/arginine-rich protein that interacts with U1-70K: We analyzed the intranuclear movement of SR45 fused to GFP by fluorescence recovery after photobleaching (FRAP) and fluorescence loss in photobleaching (FLIP). We demonstrate that the movement of GFP-SR45 is ATP-dependent. Interestingly, inhibition of transcription or phosphorylation slowed the mobility of GFP-SR45 (Ali et al., 2006). Our studies have revealed that the nuclear localization signals are located in arg/ser-rich domains (RS) 1 and 2, whereas the speckle targeting signals are exclusively present in RS2 (Ali et al., 2006). The regulation of

VP7: an attachment protein of bluetongue virus for cellular receptors in Culicoides variipennis.

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Xu, G; Wilson, W; Mecham, J; Murphy, K; Zhou, E M; Tabachnick, W

1997-07-01

The importance of VP7 of bluetongue virus (BTV) in the binding of BTV to membrane proteins of the BTV vector Culicoides variipennis was investigated. Core BTV particles, prepared from whole viruses, lacked outer proteins VP2 and VP5 and had VP7 exposed. More core particles and whole viruses bound to membrane preparations of adults of C. variipennis and KC cells, which were cultured from this vector insect, than to membrane preparations of Manduca sexta larvae. More core particles than whole viruses bound to membrane preparations of adults of C. variipennis and KC cells. Polyclonal anti-idiotypic antibodies (anti-Id), which were made against an antigen-combining region of an anti-BTV-10 VP7 antibody and functionally mimicked VP7, bound more to the membrane preparations of adults of C. variipennis and KC cells, and less to cytosol preparations. In Western overalay analysis, the Culicoides plasma membrane preparation reduced binding of an anti-VP7 monoclonal antibody to VP7. Whole and core BTV particles and the anti-Id bound to a membrane protein with a molecular mass of 23 kDa that was present predominantly in membrane preparations of adults of C. variipennis and KC cells. This protein was present in much lower concentrations in membrane preparations of C6/36 and DM-2 insect cells.

Structural and functional analysis of the human spliceosomal DEAD-box helicase Prp28

Energy Technology Data Exchange (ETDEWEB)

Möhlmann, Sina [Georg-August-University Göttingen, Justus-von-Liebig Weg 11, 37077 Göttingen (Germany); Mathew, Rebecca [Max-Planck-Institute for Biophysical Chemistry, Am Fassberg, 37077 Göttingen (Germany); Neumann, Piotr; Schmitt, Andreas [Georg-August-University Göttingen, Justus-von-Liebig Weg 11, 37077 Göttingen (Germany); Lührmann, Reinhard [Max-Planck-Institute for Biophysical Chemistry, Am Fassberg, 37077 Göttingen (Germany); Ficner, Ralf, E-mail: rficner@uni-goettingen.de [Georg-August-University Göttingen, Justus-von-Liebig Weg 11, 37077 Göttingen (Germany)

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.

Study on microstructure change of Uranium nitride coated U-7wt%Mo powder by heat treatment

Energy Technology Data Exchange (ETDEWEB)

Cho, Woo Hyoung; Park, Jae Soon; Lee, Hae In; Kim, Woo Jeong; Yang, Jae Ho; Park, Jong Man [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

2011-10-15

Uranium-molybdenum alloy particle dispersion fuel in an aluminum matrix with a high uranium density has been developed for a high performance research reactor in the RERTR program. In order to retard the fuel-matrix interaction in U-Mo/Al dispersion fuel in which the U-Mo fuel particles were dispersed in Al matrix, nitride layer coated U-Mo fuel particle has been designed and techniques to fabricate nitride-layer coated U-7wt%Mo particles have been developed in our lab. In this study, uranium nitride coated U-Mo particle has heat treatment for several times and degree. And we suggested for interaction layer remedy in U-Mo dispersion fuel. We investigate effect of heat treatment interaction layer evolution on uranium nitride coated U-Mo powder. The EDS and XRD analysis to investigate the phase evolution in uranium nitride coated layer is also a part of the present work

7 CFR 51.2541 - U.S. Fancy, U.S. Extra No. 1, U.S. No. 1 And U.S. Select Grades.

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2010-01-01

... PRODUCTS 1,2 (INSPECTION, CERTIFICATION, AND STANDARDS) United States Standards for Grades of Pistachio.... Fancy,” “U.S. Extra No. 1,” “U.S. No. 1,” and “U.S. Select” consists of pistachio nuts in the shell...

Nanodiamond enhances immune responses in mice against recombinant HA/H7N9 protein.

Science.gov (United States)

Pham, Ngoc Bich; Ho, Thuong Thi; Nguyen, Giang Thu; Le, Thuy Thi; Le, Ngoc Thu; Chang, Huan-Cheng; Pham, Minh Dinh; Conrad, Udo; Chu, Ha Hoang

2017-10-05

The continuing spread of the newly emerged H7N9 virus among poultry in China, as well as the possibility of human-to-human transmission, has attracted numerous efforts to develop an effective vaccine against H7N9. The use of nanoparticles in vaccinology is inspired by the fact that most pathogens have a dimension within the nano-size range and therefore can be processed efficiently by the immune system, which leads to a potent immune response. Herein, we report a facile approach to increase antigen size to achieve not only fast but also effective responses against the recombinant HA/H7N9 protein via a simple conjugation of the protein onto the surface of nanodiamond particles. In this study, trimeric Haemagglutinin (H7) that is transiently expressed in N. benthamiana was purified using affinity chromatography, and its trimeric state was revealed successfully by the cross-linking reaction. The trimeric H7 solution was subsequently mixed with a nanodiamond suspension in different ratios. The successful conjugation of the trimeric H7 onto the surface of nanodiamond particles was demonstrated by the changes in size and Zeta-potential of the particles before and after protein coating, Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and Western-blot analysis. Next, biofunction of the protein-nanodiamond conjugates was screened using a haemagglutination assay. A mixture containing 5 µg of trimeric H7 and 60 µg of nanodiamond corresponds to a ratio of 1:12 (w/w) of agglutinated chicken red blood cells at HA titer of 1024, which is 512-fold higher than the HA titer of free trimeric H7. After the 2nd and 3rd immunization in mice, ELISA and Western blot analyses demonstrated that the physical mixture of trimeric H7 protein and nanodiamond (1:12, w/w) elicited statistically significant stronger H7-specific-IgG response demonstrated by higher amounts of H7N9-specific IgG (over 15.4-fold with P < 0.05 after the second immunization). These results

Circulating IGF-binding protein 7 (IGFBP7 levels are elevated in patients with endometriosis or undergoing diabetic hemodialysis

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Sasajima Koji

2008-11-01

Full Text Available Abstract Background Insulin-like growth factor-binding protein-7 (IGFBP7 is a secretory protein with a molecular mass of approximately 30 kDa. It is abundantly expressed in the uterine endometrium during the secretory phase of the menstrual cycle. Decreased IGFBP7 expression has been observed in some cancers and leiomyomata. Methods To determine whether serum IGFBP7 levels reflect changes in uterine IGFBP7 expression in humans during the menstrual cycle, and to examine whether serum IGFBP7 levels are altered in patients with various disorders, we developed a novel, dual-antibody sandwich enzyme-linked immunosorbent assay (ELISA. Firstly, concentrations of IGFBP7 released into the medium were determined in cultured endometrial stromal and glandular cells. Blood samples were collected from women who had normal menstrual cycles and who had been diagnosed with endometriosis. Serum from hemodialysis patients and gastrointestinal cancers was also used to determine the IGFBP7 levels. Results Using this new ELISA, we demonstrated that cultured uterine cells secrete IGFBP7 into the medium. Patients with endometriosis and those with type II diabetes mellitus undergoing hemodialysis had significantly higher serum concentrations of IGFBP7 than the relevant control subjects. There were no differences in serum IGFBP7 levels in women at different stages of the menstrual cycle. Furthermore, serum IGFBP7 levels in patients with colorectal, esophageal, or endometrial cancer were not different than normal healthy subjects. Conclusion Our observations suggest that IGFBP7 is associated with the pathophysiology of endometriosis and diabetes mellitus, and that serum IGFBP7 levels do not reflect enhanced uterine expression of IGFBP7 mRNA during the menstrual cycle.

7 CFR 318.13-12 - Movement by the U.S. Department of Agriculture.

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2010-01-01

... 7 Agriculture 5 2010-01-01 2010-01-01 false Movement by the U.S. Department of Agriculture. 318.13-12 Section 318.13-12 Agriculture Regulations of the Department of Agriculture (Continued) ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE STATE OF HAWAII AND TERRITORIES QUARANTINE...

A Study on Silicide Coatings as Diffusion barrier for U-7Mo Dispersion Fuel

Energy Technology Data Exchange (ETDEWEB)

Won, Ju Jin; Kim, Sung Hwan; Lee, Kyu Hong; Jeong, Yong Jin; Kim, Ki Nam; Park, Jong Man; Lee, Chong Tak [KAERI, Daejeon (Korea, Republic of)

2016-05-15

Gamma phase U-Mo alloys are regarded as one of the promising candidates for advanced research reactor fuel when it comes to the irradiation performance. However, it has been reported that interaction layer formation between the UMo alloys and Al matrix degrades the irradiation performance of U-Mo dispersion fuel. The excessive interaction between the U-Mo alloys and their surrounding Al matrix lead to excessive local swelling called 'pillowing'. For this reason, KAERI suggested several remedies such as alloying U-Mo with Al matrix with Si. In addition, silicide or nitride coatings on the surface of U-Mo particles have also been proposed to hinder the growth of the interaction layer. In this study, centrifugally atomized U-7Mo alloy powders were coated with silicide layers at 900 .deg. C for 1hr. U-Mo alloy powder was mixed with MoSi{sub 2}, Si and ZrSi{sub 2} powders and subsequently heat-treated to form uranium-silicide coating layers on the surface of U-Mo alloy particles. Silicide coated U-Mo powders and characterized using scanning electron microscopy (SEM), energy dispersive x-ray spectroscopy (EDS) and X-ray diffractometer (XRD). The ZrSi{sub 2} coating layers has a thickness of about 1∼ 2μm. The surface of a silicide coated particle was very rough and silicide powder attached to the surface of the coating layer. 3. The XRD analysis of the coating layers showed that, they consisted of compounds such as U3Si{sub 2}, USi{sub 2}.

Bone morphogenetic protein-7 promotes chondrogenesis in human amniotic epithelial cells.

Science.gov (United States)

Zhou, Junjie; Yu, Guangrong; Cao, Chengfu; Pang, Jinhui; Chen, Xianqi

2011-06-01

Bone morphogenetic proteins (BMPs) play important roles at multiple stages of chondrogenesis. This study was undertaken to investigate the potential role of bone morphogenetic protein-7 (BMP-7) in the differentiation of chondrocytes using tissue engineering techniques. The impact of BMP-7 on human amniotic epithelial cells (hAECs) was tested. The hAECs were treated either with recombinant human BMP-7 cDNA or with transforming growth factor beta 1 (TGF-β1) as a positive control for three weeks in vitro. Cartilaginous differentiation and proliferation were assayed by quantitative RT-PCR, histology, and in situ hybridization. Our results were such that hAECs treated with either BMP-7 or TGF-β1 expressed cartilage markers (aggrecan, Sox9, CEP-68, and type II and X collagens) within three weeks. Compared with a control vector, BMP-7 induced a decrease in type I collagen expression, while the transcription of the cartilage-specific type II collagen remained stable. In induction experiments, BMP-7 transgenic hAECs exhibited the largest amount of matrix synthesis. In conclusion, these data indicate that BMP-7 plays an important role in inducing the production of cartilage by hAECs in vitro. Cartilage differentiation and matrix maturation can be promoted by BMPs in a cartilage engineering paradigm. These properties make BMPs promising tools in the engineering of cartilaginous joint bio-prostheses and as candidate biological agents or genes for cartilage stabilisation.

Interaction of uranyl ions with snake venom proteins from Lachesis muta muta

International Nuclear Information System (INIS)

MacCordick, H.J.; Taghva, F.

1997-01-01

The reaction product of uranyl nitrate with whole-protein Bushmaster snake venom in nitrate buffer at pH 3.5 has been studied. The maximum uptake of uranium was 291 μmol U x g -1 of venom. The infrared spectrum of the product showed an asymmetric O-U-O vibration at 921 cm -1 typical of complex formation with the uranyl ion. Stability measurements with the UO 2 2+ -protein complex in neutral medium indicated moderate hydrolytic stability, with 14% dissociation after 16 hours at 0 deg C. Neutron irradiation and desorption studies with a 235 U-labelled complex showed that generated fission products such as lanthanides and barium were readily lixiviated at pH 7, whereas Ru and Zr were highly retained by the protein substrate. (author)

Molecular Characterization of LRB7 Gene and a Water Channel Protein TIP2 in Chorispora bungeana

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Ming Li

2016-01-01

Full Text Available Background. Water channel proteins, also called aquaporins, are integral membrane proteins from major intrinsic protein (MIP family and involved in several pathways including not only water transport but also cell signaling, reproduction, and photosynthesis. The full cDNA and protein sequences of aquaporin in Chorispora bungeana Fisch. & C.A. Mey (C. bungeana are still unknown. Results. In this study, PCR and rapid amplification of cDNA ends approaches were used to clone the full cDNA of LRB7 (GenBank accession number: EU636988 of C. bungeana. Sequence analysis indicated that it was 1235 bp, which had two introns and encoded a protein of 250 amino acids. Structure analysis revealed that the protein had two conserved NPA motifs, one of which is MIP signature sequence (SGxHxNPAVT, six membrane helix regions, and additional membrane-embedded domains. Phylogenetic analysis suggested that the protein was from TIP2 subgroup. Surprisingly, semiquantitative RT-PCR experiment and western blot analysis showed that LRB7 and TIP2 were only detectable in roots, unlike Arabidopsis and Raphanus. Connecting with our previous studies, LRB7 was supported to associate with chilling-tolerance in C. bungeana. Conclusion. This is the first time to characterize the full sequences of LRB7 gene and water channel protein in C. bungeana. Our findings contribute to understanding the water transports in plants under low temperatures.

Cleanup Summary Report for the Defense Threat Reduction Agency Fiscal Year 2007, Task 6.7, U12u-Tunnel (Legacy Site), Nevada Test Site, Nevada

International Nuclear Information System (INIS)

2008-01-01

This letter serves as notice of completion for cleanup of the U12u-Tunnel (Legacy Site) as specified in the Defense Threat Reduction Agency (DTRA) Fiscal Year 2007 Statement of Work, Task 6.7. The U12u-Tunnel Legacy Site is located near the intersection of the U12u-Tunnel access road and the U12n-Tunnel access road in Area 12 of the Nevada Test Site (see Figure 1). The site encompasses 1.2 acres and was used to store miscellaneous mining equipment and materials that were used to support DTRA testing in Area 12. Field activities commenced February 11, 2008, and were completed February 20, 2008. Radiological surveys were performed on a drill jumbo and all material stored at the site. The drill jumbo was relocated to U12p-Tunnel portal and consolidated with other critical mining equipment for future use or storage. Ten truck loads of solid waste (53 tons) were shipped to the Nevada Test Site, Area 9 U10c Sanitary Landfill for disposal. No hazardous or radiological waste was generated at this site

7 CFR 51.303 - U.S. Utility.

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2010-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... Standards for Grades of Apples Grades § 51.303 U.S. Utility. “U.S. Utility” consists of apples of one...

7 CFR 51.342 - U.S. Cider.

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2010-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... Standards for Grades of Apples for Processing Grades § 51.342 U.S. Cider. “U.S. Cider” consists of apples...

Identification of novel putative-binding proteins for cellular prion protein and a specific interaction with the STIP1 homology and U-Box-containing protein 1

Science.gov (United States)

Gimenez, Ana Paula Lappas; Richter, Larissa Morato Luciani; Atherino, Mariana Campos; Beirão, Breno Castello Branco; Fávaro, Celso; Costa, Michele Dietrich Moura; Zanata, Silvio Marques; Malnic, Bettina; Mercadante, Adriana Frohlich

2015-01-01

ABSTRACT Prion diseases involve the conversion of the endogenous cellular prion protein, PrPC, into a misfolded infectious isoform, PrPSc. Several functions have been attributed to PrPC, and its role has also been investigated in the olfactory system. PrPC is expressed in both the olfactory bulb (OB) and olfactory epithelium (OE) and the nasal cavity is an important route of transmission of diseases caused by prions. Moreover, Prnp−/− mice showed impaired behavior in olfactory tests. Given the high PrPC expression in OE and its putative role in olfaction, we screened a mouse OE cDNA library to identify novel PrPC-binding partners. Ten different putative PrPC ligands were identified, which were involved in functions such as cellular proliferation and apoptosis, cytoskeleton and vesicle transport, ubiquitination of proteins, stress response, and other physiological processes. In vitro binding assays confirmed the interaction of PrPC with STIP1 homology and U-Box containing protein 1 (Stub1) and are reported here for the first time. Stub1 is a co-chaperone with ubiquitin E3-ligase activity, which is associated with neurodegenerative diseases characterized by protein misfolding and aggregation. Physiological and pathological implications of PrPC-Stub1 interaction are under investigation. The PrPC-binding proteins identified here are not exclusive to the OE, suggesting that these interactions may occur in other tissues and play general biological roles. These data corroborate the proposal that PrPC is part of a multiprotein complex that modulates several cellular functions and provide a platform for further studies on the physiological and pathological roles of prion protein. PMID:26237451

Dicty_cDB: Contig-U01056-1 [Dicty_cDB

Lifescience Database Archive (English)

Full Text Available AJ928354 ) Theileria annulata EST, clone tam025d03_q1k. 32 5.7 3 ( Z32534 ) Human rotavirus (St Thomas) gene... 5 (1566bp). 44 6.0 1 ( U11492 ) Human rotavirus ST3 non-structural protein NSP1 mRN... 44 6.0 1 ( EF672613 ) Human rotavirus

Evening primrose (Oenothera paradoxa) cake as an unconventional protein source.

Science.gov (United States)

Golabczak, J; Py, R

2003-01-01

An efficient procedure of a protein isolate production from the evening primrose cake was developed. The cake is a by-product of oil extraction from seeds by using the cold pressing method. The evening primrose cake contains 22.7% of protein. Its content in the protein isolate derived from the cake is 74%. Proteins present in evening primrose seeds are rich in Trp (7%) and Met (3%), but Lys-deficient (1.3%) as compared to the FAO protein standard. Apart from the proteins, the protein isolate contains 8.5% (w/w per s.s.) dietary fiber, that negatively affects its digestibility. To enhance the bio-availability of the protein isolate, it was partially hydrolyzed with commercial preparations of trypsin and other proteases (Alcalase and Flavourzyme, Novozymes. Denmark). The most advanced proteolysis (52%) was achieved by 6 h digestion of 2% protein suspension with a mixture of Flavourzyme and Alcalase (350 and 600 U per g of protein, respectively) at 50 degrees C and pH 9.0.

Centromere Protein (CENP)-W Interacts with Heterogeneous Nuclear Ribonucleoprotein (hnRNP) U and May Contribute to Kinetochore-Microtubule Attachment in Mitotic Cells

Science.gov (United States)

Chun, Younghwa; Kim, Raehyung; Lee, Soojin

2016-01-01

Background Recent studies have shown that heterogeneous nuclear ribonucleoprotein U (hnRNP U), a component of the hnRNP complex, contributes to stabilize the kinetochore-microtubule interaction during mitosis. CENP-W was identified as an inner centromere component that plays crucial roles in the formation of a functional kinetochore complex. Results We report that hnRNP U interacts with CENP-W, and the interaction between hnRNP U and CENP-W mutually increased each other’s protein stability by inhibiting the proteasome-mediated degradation. Further, their co-localization was observed chiefly in the nuclear matrix region and at the microtubule-kinetochore interface during interphase and mitosis, respectively. Both microtubule-stabilizing and microtubule-destabilizing agents significantly decreased the protein stability of CENP-W. Furthermore, loss of microtubules and defects in microtubule organization were observed in CENP-W-depleted cells. Conclusion Our data imply that CENP-W plays an important role in the attachment and interaction between microtubules and kinetochore during mitosis. PMID:26881882

7 CFR 51.301 - U.S. Fancy.

Science.gov (United States)

2010-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... Standards for Grades of Apples Grades § 51.301 U.S. Fancy. “U.S. Fancy” consists of apples of one variety...

Structure of Rotavirus Outer-Layer Protein VP7 Bound with a Neutralizing Fab

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Aoki, Scott T.; Settembre, Ethan C.; Trask, Shane D.; Greenberg, Harry B.; Harrison, Stephen C.; Dormitzer, Philip R.; (Stanford-MED); (CH-Boston)

2009-06-17

Rotavirus outer-layer protein VP7 is a principal target of protective antibodies. Removal of free calcium ions (Ca{sup 2+}) dissociates VP7 trimers into monomers, releasing VP7 from the virion, and initiates penetration-inducing conformational changes in the other outer-layer protein, VP4. We report the crystal structure at 3.4 angstrom resolution of VP7 bound with the Fab fragment of a neutralizing monoclonal antibody. The Fab binds across the outer surface of the intersubunit contact, which contains two Ca{sup 2+} sites. Mutations that escape neutralization by other antibodies suggest that the same region bears the epitopes of most neutralizing antibodies. The monovalent Fab is sufficient to neutralize infectivity. We propose that neutralizing antibodies against VP7 act by stabilizing the trimer, thereby inhibiting the uncoating trigger for VP4 rearrangement. A disulfide-linked trimer is a potential subunit immunogen.

C-terminal region of MAP7 domain containing protein 3 (MAP7D3 promotes microtubule polymerization by binding at the C-terminal tail of tubulin.

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Saroj Yadav

Full Text Available MAP7 domain containing protein 3 (MAP7D3, a newly identified microtubule associated protein, has been shown to promote microtubule assembly and stability. Its microtubule binding region has been reported to consist of two coiled coil motifs located at the N-terminus. It possesses a MAP7 domain near the C-terminus and belongs to the microtubule associated protein 7 (MAP7 family. The MAP7 domain of MAP7 protein has been shown to bind to kinesin-1; however, the role of MAP7 domain in MAP7D3 remains unknown. Based on the bioinformatics analysis of MAP7D3, we hypothesized that the MAP7 domain of MAP7D3 may have microtubule binding activity. Indeed, we found that MAP7 domain of MAP7D3 bound to microtubules as well as enhanced the assembly of microtubules in vitro. Interestingly, a longer fragment MDCT that contained the MAP7 domain (MD with the C-terminal tail (CT of the protein promoted microtubule polymerization to a greater extent than MD and CT individually. MDCT stabilized microtubules against dilution induced disassembly. MDCT bound to reconstituted microtubules with an apparent dissociation constant of 3.0 ± 0.5 µM. An immunostaining experiment showed that MDCT localized along the length of the preassembled microtubules. Competition experiments with tau indicated that MDCT shares its binding site on microtubules with tau. Further, we present evidence indicating that MDCT binds to the C-terminal tail of tubulin. In addition, MDCT could bind to tubulin in HeLa cell extract. Here, we report a microtubule binding region in the C-terminal region of MAP7D3 that may have a role in regulating microtubule assembly dynamics.

37 CFR 7.31 - Requirements for transformation of an extension of protection to the United States into a U.S...

Science.gov (United States)

2010-07-01

... transformation of an extension of protection to the United States into a U.S. application. 7.31 Section 7.31... transformation of an extension of protection to the United States into a U.S. application. If the International... transformation within three months of the date of cancellation of the international registration and include: (1...

Insulin-like Growth Factor Binding Protein 7 Mediates Glioma Cell Growth and Migration

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Wei Jiang

2008-12-01

Full Text Available Insulin-like growth factor binding protein 7 (IGFBP-7 is the only member of the IGFBP superfamily that binds strongly to insulin, suggesting that IGFBP-7 may have different functions from other IGFBPs. Unlike other IGFBPs, the expression and functions of IGFBP-7 in glioma tumors have not been reported. Using cDNA microarray analysis, we found that expression of IGFBP-7 correlated with the grade of glioma tumors and the overall patient survival. This finding was further validated by real-time reverse transcription-polymerase chain reaction and Western blot analysis. We used RNAi to examine the role of IGFBP-7 in glioma cells, inhibiting IGFBP-7 expression by short interfering RNA transfection. Cell proliferation was suppressed after IGFBP-7 expression was inhibited for 5 days, and glioma cell growth was stimulated consistently by the addition of recombinant IGFBP-7 protein. Moreover, glioma cell migration was attenuated by IGFBP-7 depletion but enhanced by IGFBP-7 overexpression and addition. Overexpression of AKT1 in IGFBP-7-overxpressed cells attenuated the IGFBP-7-promoted migration and further enhanced inhibition of IGFBP-7 depletion on the migration. Phosphorylation of AKT and Erk1/2 was also inversely regulated by IGFBP-7 expression. These two factors together suggest that IGFBP-7 can regulate glioma cell migration through the AKT-ERK pathway, thereby playing an important role in glioma growth and migration.

Assembly and dynamics of the U4/U6 di-snRNP by single-molecule FRET

Science.gov (United States)

Hardin, John W.; Warnasooriya, Chandani; Kondo, Yasushi; Nagai, Kiyoshi; Rueda, David

2015-01-01

In large ribonucleoprotein machines, such as ribosomes and spliceosomes, RNA functions as an assembly scaffold as well as a critical catalytic component. Protein binding to the RNA scaffold can induce structural changes, which in turn modulate subsequent binding of other components. The spliceosomal U4/U6 di-snRNP contains extensively base paired U4 and U6 snRNAs, Snu13, Prp31, Prp3 and Prp4, seven Sm and seven LSm proteins. We have studied successive binding of all protein components to the snRNA duplex during di-snRNP assembly by electrophoretic mobility shift assay and accompanying conformational changes in the U4/U6 RNA 3-way junction by single-molecule FRET. Stems I and II of the duplex were found to co-axially stack in free RNA and function as a rigid scaffold during the entire assembly, but the U4 snRNA 5′ stem-loop adopts alternative orientations each stabilized by Prp31 and Prp3/4 binding accounting for altered Prp3/4 binding affinities in presence of Prp31. PMID:26503251

7 CFR 1221.7 - Customs.

Science.gov (United States)

2010-01-01

... 7 Agriculture 10 2010-01-01 2010-01-01 false Customs. 1221.7 Section 1221.7 Agriculture... INFORMATION ORDER Sorghum Promotion, Research, and Information Order Definitions § 1221.7 Customs. Customs means the U.S. Customs and Border Protection of the U.S. Department of Homeland Security. ...

Temperature field and thermal stress analysis of the HT-7U vacuum vessel

International Nuclear Information System (INIS)

Song Yuntao; Yao Damao; Wu Songtao; Weng Peide

2000-01-01

The HT-7U vacuum vessel is an all-metal-welded double-wall interconnected with toroidal and poloidal stiffening ribs. The channels formed between the ribs and walls are filled with boride water as a nuclear shielding. On the vessel surface facing the plasma are installed cable-based Ohmic heaters. Prior to plasma operation the vessel is to be baked out and discharge cleaned at about 250 degree C. During baking out the non-uniformity of temperature distribution on the vacuum vessel will bring about serious thermal stress that can damage the vessel. In order to determine and optimize the design of the HT-7U vacuum vessel, a three-dimensional finite element model was performed to analyse its temperature field and thermal stress. the maximal thermal stress appeared on the round of lower vertical port and maximal deformation located just on the region between the upper vertical port and the horizontal port. The results show that the reinforced structure has a good capability of withstanding the thermal loads

Role of Bunyamwera Orthobunyavirus NSs protein in infection of mosquito cells.

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Szemiel, Agnieszka M; Failloux, Anna-Bella; Elliott, Richard M

2012-01-01

Bunyamwera orthobunyavirus is both the prototype and study model of the Bunyaviridae family. The viral NSs protein seems to contribute to the different outcomes of infection in mammalian and mosquito cell lines. However, only limited information is available on the growth of Bunyamwera virus in cultured mosquito cells other than the Aedes albopictus C6/36 line. To determine potential functions of the NSs protein in mosquito cells, replication of wild-type virus and a recombinant NSs deletion mutant was compared in Ae. albopictus C6/36, C7-10 and U4.4 cells, and in Ae. aegypti Ae cells by monitoring N protein production and virus yields at various times post infection. Both viruses established persistent infections, with the exception of NSs deletion mutant in U4.4 cells. The NSs protein was nonessential for growth in C6/36 and C7-10 cells, but was important for productive replication in U4.4 and Ae cells. Fluorescence microscopy studies using recombinant viruses expressing green fluorescent protein allowed observation of three stages of infection, early, acute and late, during which infected cells underwent morphological changes. In the absence of NSs, these changes were less pronounced. An RNAi response efficiently reduced virus replication in U4.4 cells transfected with virus specific dsRNA, but not in C6/36 or C7/10 cells. Lastly, Ae. aegypti mosquitoes were exposed to blood-meal containing either wild-type or NSs deletion virus, and at various times post-feeding, infection and disseminated infection rates were measured. Compared to wild-type virus, infection rates by the mutant virus were lower and more variable. If the NSs deletion virus was able to establish infection, it was detected in salivary glands at 6 days post-infection, 3 days later than wild-type virus. Bunyamwera virus NSs is required for efficient replication in certain mosquito cell lines and in Ae. aegypti mosquitoes.

Structural basis for antagonizing a host restriction factor by C7 family of poxvirus host-range proteins.

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Meng, Xiangzhi; Krumm, Brian; Li, Yongchao; Deng, Junpeng; Xiang, Yan

2015-12-01

Human sterile alpha motif domain-containing 9 (SAMD9) protein is a host restriction factor for poxviruses, but it can be overcome by some poxvirus host-range proteins that share homology with vaccinia virus C7 protein. To understand the mechanism of action for this important family of host-range factors, we determined the crystal structures of C7 and myxoma virus M64, a C7 family member that is unable to antagonize SAMD9. Despite their different functions and only 23% sequence identity, the two proteins have very similar overall structures, displaying a previously unidentified fold comprised of a compact 12-stranded antiparallel β-sandwich wrapped in two short α helices. Extensive structure-guided mutagenesis of C7 identified three loops clustered on one edge of the β sandwich as critical for viral replication and binding with SAMD9. The loops are characterized with functionally important negatively charged, positively charged, and hydrophobic residues, respectively, together forming a unique "three-fingered molecular claw." The key residues of the claw are not conserved in two C7 family members that do not antagonize SAMD9 but are conserved in distantly related C7 family members from four poxvirus genera that infect diverse mammalian species. Indeed, we found that all in the latter group of proteins bind SAMD9. Taken together, our data indicate that diverse mammalian poxviruses use a conserved molecular claw in a C7-like protein to target SAMD9 and overcome host restriction.

39 CFR 7.7 - Enforcement.

Science.gov (United States)

2010-07-01

... 39 Postal Service 1 2010-07-01 2010-07-01 false Enforcement. 7.7 Section 7.7 Postal Service UNITED STATES POSTAL SERVICE THE BOARD OF GOVERNORS OF THE U.S. POSTAL SERVICE PUBLIC OBSERVATION (ARTICLE VII) § 7.7 Enforcement. (a) Under 5 U.S.C. 552b(g), any person may bring a proceeding in the United States...

The prion protein has RNA binding and chaperoning properties characteristic of nucleocapsid protein NCP7 of HIV-1.

Science.gov (United States)

Gabus, C; Derrington, E; Leblanc, P; Chnaiderman, J; Dormont, D; Swietnicki, W; Morillas, M; Surewicz, W K; Marc, D; Nandi, P; Darlix, J L

2001-06-01

Transmissible spongiform encephalopathies are fatal neurodegenerative diseases associated with the accumulation of a protease-resistant form of the prion protein (PrP). Although PrP is conserved in vertebrates, its function remains to be identified. In vitro PrP binds large nucleic acids causing the formation of nucleoprotein complexes resembling human immunodeficiency virus type 1 (HIV-1) nucleocapsid-RNA complexes and in vivo MuLV replication accelerates the scrapie infectious process, suggesting possible interactions between retroviruses and PrP. Retroviruses, including HIV-1 encode a major nucleic acid binding protein (NC protein) found within the virus where 2000 NC protein molecules coat the dimeric genome. NC is required in virus assembly and infection to chaperone RNA dimerization and packaging and in proviral DNA synthesis by reverse transcriptase (RT). In HIV-1, 5'-leader RNA/NC interactions appear to control these viral processes. This prompted us to compare and contrast the interactions of human and ovine PrP and HIV-1 NCp7 with HIV-1 5'-leader RNA. Results show that PrP has properties characteristic of NCp7 with respect to viral RNA dimerization and proviral DNA synthesis by RT. The NC-like properties of huPrP map to the N-terminal region of huPrP. Interestingly, PrP localizes in the membrane and cytoplasm of PrP-expressing cells. These findings suggest that PrP is a multifunctional protein possibly participating in nucleic acid metabolism.

Cloning and Expression of Plantaricin W Produced by Lactobacillus plantarum U10 Isolate from "Tempoyak" Indonesian Fermented Food as Immunity Protein in Lactococcus lactis.

Science.gov (United States)

Lages, Aksar Chair; Mustopa, Apon Zaenal; Sukmarini, Linda; Suharsono

2015-10-01

Plantaricins, one of bacteriocin produced by Lactobacillus plantarum, are already known to have activities against several pathogenic bacterium. L. plantarum U10 isolated from "tempoyak," an Indonesian fermented food, produced one kind of plantaricin designated as plantaricin W (plnW). The plnW is suggested as a putative membrane location of protein and has similar conserved motif which is important as immunity to bacteriocin itself. Thus, due to study about this plantaricin, several constructs have been cloned and protein was analyzed in Lactococcus lactis. In this study, plnW gene was successfully cloned into vector NICE system pNZ8148 and created the transformant named L. lactis NZ3900 pNZ8148-WU10. PlnW protein was 25.3 kDa in size. The concentration of expressed protein was significantly increased by 10 ng/mL nisin induction. Furthermore, PlnW exhibited protease activity with value of 2.22 ± 0.05 U/mL and specific activity about 1.65 ± 0.03 U/mg protein with 50 ng/mL nisin induction. Immunity study showed that the PlnW had immunity activity especially against plantaricin and rendered L. lactis recombinant an immunity broadly to other bacteriocins such as pediocin, fermentcin, and acidocin.

Immunization with mutant HPV16 E7 protein inhibits the growth of TC-1 cells in tumor-bearing mice.

Science.gov (United States)

Li, Yan-Li; Ma, Zhong-Liang; Zhao, Yue; Zhang, Jing

2015-04-01

Two human papillomavirus (HPV) 16 oncogenic proteins, E6 and E7, are co-expressed in the majority of HPV16-induced cervical cancer cells. Thus, the E6 and E7 proteins are good targets for developing therapeutic vaccines for cervical cancer. In the present study, immunization with the mutant non-transforming HPV16 E7 (mE7) protein was demonstrated to inhibit the growth of TC-1 cells in the TC-1 mouse model. The HPV16 mE7 gene was amplified by splicing overlap extension polymerase chain reaction using pET-28a(+)-E7 as a template, and the gene was cloned into pET-28a(+) to form pET-28a(+)-mE7. Compared with the E7 protein, mE7 lacks amino acid residues 94-98, and at residue 24, there is a Cys to Gly substitution. pET-28a(+)-mE7 was then introduced into Escherichia coli Rosetta. The expression of mE7 was induced by isopropyl β-D-1-thiogalactopyranoside. The mE7 protein was purified using Ni-NTA agarose and detected by SDS-PAGE and western blot analysis. In the tumor prevention model, no tumor was detected in the mice vaccinated with the mE7 protein. After 40 days, the tumor-free mice and control mice were challenged with 2×10 5 TC-1 cells. All control mice developed tumors six days later, but mE7 immunized mice were tumor free until 90 days. In the tumor therapy model, the TC-1 cells were initially injected subcutaneously, and the mice were subsequently vaccinated. Vaccination against the mE7 protein may significantly inhibit TC-1 cell growth compared to the control. These results demonstrated that immunization with the HPV16 mE7 protein elicited a long-term protective immunity against TC-1 tumor growth and generated a significant inhibition of TC-1 growth in a TC-1 mouse model.

Uniform 15N- and 15N/13C-labeling of proteins in mammalian cells and solution structure of the amino terminal fragment of u-PA

International Nuclear Information System (INIS)

Hansen, A.P.; Petros, A.M.; Meadows, R.P.; Mazar, A.P.; Nettesheim, D.G.; Pederson, T.M.; Fesik, S.W.

1994-01-01

Urokinase-type plasminogen activator (u-PA) is a 54-kDa glycoprotein that catalyzes the conversion of plasminogen to plasmin, a broad-specificity protease responsible for the degradation of fibrin clots and extracellular matrix components. The u-PA protein consists of three individual modules: a growth factor domain (GFD), a kringle, and a serine protease domain. The amino terminal fragment (ATF) includes the GFD-responsible for u-PA binding to its receptor-and the kringle domains. This protein was expressed and uniformly 15 N-and 15 N/ 13 C-labeled in mammalian cells by methods that will be described. In addition, we present the three-dimensional structure of ATF that was derived from 1299 NOE-derived distance restraints along with the φ angle and hydrogen bonding restraints. Although the individual domains in the structures were highly converged, the two domains are structurally independent. The overall structures of the individual domains are very similar to the structures of homologous proteins. However, important structural differences between the growth factor domain of u-PA and other homologous proteins were observed in the region that has been implicated in binding the urokinase receptor. These results may explain, in part, why other growth factors show no appreciable affinity for the urokinase receptor

Protein P7 of the cystovirus φ6 is located at the three-fold axis of the unexpanded procapsid.

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Garrett Katz

Full Text Available The objective of this study was to determine the location of protein P7, the RNA packaging factor, in the procapsid of the φ6 cystovirus. A comparison of cryo-electron microscopy high-resolution single particle reconstructions of the φ6 complete unexpanded procapsid, the protein P2-minus procapsid (P2 is the RNA directed RNA-polymerase, and the P7-minus procapsid, show that prior to RNA packaging the P7 protein is located near the three-fold axis of symmetry. Difference maps highlight the precise position of P7 and demonstrate that in P7-minus particles the P2 proteins are less localized with reduced densities at the three-fold axes. We propose that P7 performs the mechanical function of stabilizing P2 on the inner protein P1 shell which ensures that entering viral single-stranded RNA is replicated.

Interactions of β-Conglycinin (7S with Different Phenolic Acids—Impact on Structural Characteristics and Proteolytic Degradation of Proteins

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Jing Gan

2016-10-01

Full Text Available p-Coumalic acid (PCA, caffeic acid (CA, gallic acid (GA and chlorogenic acid (CGA are the major phenolic acids that co-exist with soy protein components in foodstuffs. Surprisingly, there are only a handful of reports that describe their interaction with β-Conglycinin (7S, a major soy protein. In this report, we investigated the interaction between phenolic acids and soy protein 7S and observed an interaction between each of these phenolic acids and soy protein 7S, which was carried out by binding. Further analysis revealed that the binding activity of the phenolic acids was structure dependent. Here, the binding affinity of CA and GA towards 7S was found to be stronger than that of PCA, because CA and GA have one more hydroxyl group. Interestingly, the binding of phenolic acids with soy protein 7S did not affect protein digestion by pepsin and trypsin. These findings aid our understanding of the relationship between different phenolic acids and proteins in complex food systems.

The AMPA receptor-associated protein Shisa7 regulates hippocampal synaptic function and contextual memory

NARCIS (Netherlands)

Schmitz, Leanne J M; Klaassen, Remco V; Ruiperez-Alonso, Marta; Zamri, Azra Elia; Stroeder, Jasper; Rao-Ruiz, Priyanka; Lodder, Johannes C; van der Loo, Rolinka J; Mansvelder, Huib D; Smit, August B; Spijker, Sabine; Verhage, Matthijs

2017-01-01

Glutamatergic synapses rely on AMPA receptors (AMPARs) for fast synaptic transmission and plasticity. AMPAR auxiliary proteins regulate receptor trafficking, and modulate receptor mobility and its biophysical properties. The AMPAR auxiliary protein Shisa7 (CKAMP59) has been shown to interact with

Analysis of Protein–Protein Interactions in MCF-7 and MDA-MB-231 Cell Lines Using Phthalic Acid Chemical

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Shih-Shin Liang

2014-11-01

Full Text Available Phthalates are a class of plasticizers that have been characterized as endocrine disrupters, and are associated with genital diseases, cardiotoxicity, hepatotoxicity, and nephrotoxicity in the GeneOntology gene/protein database. In this study, we synthesized phthalic acid chemical probes and demonstrated differing protein–protein interactions between MCF-7 cells and MDA-MB-231 breast cancer cell lines. Phthalic acid chemical probes were synthesized using silicon dioxide particle carriers, which were modified using the silanized linker 3-aminopropyl triethoxyslane (APTES. Incubation with cell lysates from breast cancer cell lines revealed interactions between phthalic acid and cellular proteins in MCF-7 and MDA-MB-231 cells. Subsequent proteomics analyses indicated 22 phthalic acid-binding proteins in both cell types, including heat shock cognate 71-kDa protein, ATP synthase subunit beta, and heat shock protein HSP 90-beta. In addition, 21 MCF-7-specific and 32 MDA-MB-231 specific phthalic acid-binding proteins were identified, including related proteasome proteins, heat shock 70-kDa protein, and NADPH dehydrogenase and ribosomal correlated proteins, ras-related proteins, and members of the heat shock protein family, respectively.

The VULCANO VE-U7 Corium spreading benchmark

Energy Technology Data Exchange (ETDEWEB)

Journeau, Christophe; Haquet, Jean-Francois [CEA Cadarache, Severe Accident Mastering experimental Laboratory (DEN/DTN/STRI/LMA), 13108 St Paul lez Durance (France); Spindler, Bertrand [CEA Grenoble, Physicochemistry and Multiphasic Thermalhydraulics Laboratory (DEN/DTN/SE2T/LPTM), 17 rue des Martyrs, F-38054 Grenoble CEDEX 9 (France); Spengler, Claus [Gesellschaft fuer Reaktorsicherheit mbH, Department for Thermohydraulics/Process Engineering, Schwertnergasse 1, D-50667 Koeln (Germany); Foit, Jerzy [Forschungszentrum Karlsruhe GmbH, Institut fuer Kern nd Energietechnik (IKET), P.O. Box 3640, D-76021 Karlsruhe (Germany)

2006-07-01

In a hypothetical nuclear reactor severe accident, corium spreading is one possible mitigation measure that has been selected for the EPR design. A post-test benchmark exercise has been organized on the VULCANO VE-U7 corium spreading experiment. In this test, a prototypic corium mixture representative of what could be expected at the opening of EPR reactor-pit gate has been spread on siliceous concrete and on a reference channel in inert refractory ceramic. The spreading progression was not much affected by the presence of concrete and sparging gases. The procedure used to estimate the corium physical properties from its composition and temperature provided a satisfactory data set. The CORFLOW, LAVA and THEMA codes provide satisfactory calculations of the spreading front evolution and of its final length. LAVA and THEMA estimations of the substrate temperatures, which are the initial conditions for longer term Molten Core Concrete Interaction or Corium Ceramic Interaction computations, are also close to the measured data, within the experimental uncertainties. (authors)

The VULCANO VE-U7 Corium spreading benchmark

International Nuclear Information System (INIS)

Journeau, Christophe; Haquet, Jean-Francois; Spindler, Bertrand; Spengler, Claus; Foit, Jerzy

2006-01-01

In a hypothetical nuclear reactor severe accident, corium spreading is one possible mitigation measure that has been selected for the EPR design. A post-test benchmark exercise has been organized on the VULCANO VE-U7 corium spreading experiment. In this test, a prototypic corium mixture representative of what could be expected at the opening of EPR reactor-pit gate has been spread on siliceous concrete and on a reference channel in inert refractory ceramic. The spreading progression was not much affected by the presence of concrete and sparging gases. The procedure used to estimate the corium physical properties from its composition and temperature provided a satisfactory data set. The CORFLOW, LAVA and THEMA codes provide satisfactory calculations of the spreading front evolution and of its final length. LAVA and THEMA estimations of the substrate temperatures, which are the initial conditions for longer term Molten Core Concrete Interaction or Corium Ceramic Interaction computations, are also close to the measured data, within the experimental uncertainties. (authors)

Properties measurements of (U{sub 0.7}Pu{sub 0.3})O{sub 2-x} in PO{sub 2}-controlled atmosphere

Energy Technology Data Exchange (ETDEWEB)

Kato, M.; Murakami, T.; Sunaoshi, T. [Advanced Nuclear System Research and Development Directorate, Japan Atomic Energy Agency, Muramatsu Tokai-mura Ibaraki, 319-1194 (Japan); Nelson, A.T.; McClellan, K.J. [Los Alamos National Laboratory, Los Alamos, New Mexico 87545 (United States)

2013-07-01

The investigation of physical properties of uranium and plutonium mixed oxide (MOX) fuels is important for the development of fast reactor fuels. It is well known that MOX is a nonstoichiometric oxide, and the physical properties change drastically with the Oxygen-to-Metal (O/M) ratio. A control technique for O/M ratio was established for measurements of high temperature properties of uranium and plutonium mixed oxide fuels. Sintering behavior, thermal expansion and O/M change of (U{sub 0.7}Pu{sub 0.3})O{sub 2.00} and (U{sub 0.7}Pu{sub 0.3})O{sub 1.99} were investigated in PO{sub 2}-controlled atmosphere which was controlled by H{sub 2}/H{sub 2}O gas system. Sintering behavior changed drastically with O/M ratio, and shrinkage of (U{sub 0.7}Pu{sub 0.3})O{sub 2.00} was faster and more advanced at lower temperatures as compared with (U{sub 0.7}Pu{sub 0.3})O{sub 1.99}. Thermal expansion was observed to be slightly increased with decreasing O/M ratio. (authors)

Regulator of G-Protein Signaling 7 Regulates Reward Behavior by Controlling Opioid Signaling in the Striatum.

Science.gov (United States)

Sutton, Laurie P; Ostrovskaya, Olga; Dao, Maria; Xie, Keqiang; Orlandi, Cesare; Smith, Roy; Wee, Sunmee; Martemyanov, Kirill A

2016-08-01

Morphine mediates its euphoric and analgesic effects by acting on the μ-opioid receptor (MOR). MOR belongs to the family of G-protein coupled receptors whose signaling efficiency is controlled by the regulator of G-protein signaling (RGS) proteins. Our understanding of the molecular diversity of RGS proteins that control MOR signaling, their circuit specific actions, and underlying cellular mechanisms is very limited. We used genetic approaches to ablate regulator of G-protein signaling 7 (RGS7) both globally and in specific neuronal populations. We used conditioned place preference and self-administration paradigms to examine reward-related behavior and a battery of tests to assess analgesia, tolerance, and physical dependence to morphine. Electrophysiology approaches were applied to investigate the impact of RGS7 on morphine-induced alterations in neuronal excitability and plasticity of glutamatergic synapses. At least three animals were used for each assessment. Elimination of RGS7 enhanced reward, increased analgesia, delayed tolerance, and heightened withdrawal in response to morphine administration. RGS7 in striatal neurons was selectively responsible for determining the sensitivity of rewarding and reinforcing behaviors to morphine without affecting analgesia, tolerance, and withdrawal. In contrast, deletion of RGS7 in dopaminergic neurons did not influence morphine reward. RGS7 exerted its effects by controlling morphine-induced changes in excitability of medium spiny neurons in nucleus accumbens and gating the compositional plasticity of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid and N-methyl-D-aspartate receptors. This study identifies RGS7 as a novel regulator of MOR signaling by dissecting its circuit specific actions and pinpointing its role in regulating morphine reward by controlling the activity of nucleus accumbens neurons. Copyright © 2016 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

Mpn1, Mutated in Poikiloderma with Neutropenia Protein 1, Is a Conserved 3′-to-5′ RNA Exonuclease Processing U6 Small Nuclear RNA

Directory of Open Access Journals (Sweden)

Vadim Shchepachev

2012-10-01

Full Text Available Clericuzio-type poikiloderma with neutropenia (PN is a rare genodermatosis associated with mutations in the C16orf57 gene, which codes for the uncharacterized protein hMpn1. We show here that, in both fission yeasts and humans, Mpn1 processes the spliceosomal U6 small nuclear RNA (snRNA posttranscriptionally. In Mpn1-deficient cells, U6 molecules carry 3′ end polyuridine tails that are longer than those in normal cells and lack a terminal 2′,3′ cyclic phosphate group. In mpn1Δ yeast cells, U6 snRNA and U4/U6 di-small nuclear RNA protein complex levels are diminished, leading to precursor messenger RNA splicing defects, which are reverted by expression of either yeast or human Mpn1 and by overexpression of U6. Recombinant hMpn1 is a 3′-to-5′ RNA exonuclease that removes uridines from U6 3′ ends, generating terminal 2′,3′ cyclic phosphates in vitro. Finally, U6 degradation rates increase in mpn1Δ yeasts and in lymphoblasts established from individuals affected by PN. Our data indicate that Mpn1 promotes U6 stability through 3′ end posttranscriptional processing and implicate altered U6 metabolism as a potential mechanism for PN pathogenesis.

7 CFR 51.2542 - U.S. Artificially Opened.

Science.gov (United States)

2010-01-01

... STANDARDS) United States Standards for Grades of Pistachio Nuts in the Shell § 51.2542 U.S. Artificially Opened. “U.S. Artificially Opened” consists of artificially opened pistachio nuts in the shell which meet...

Lysine-Less Variants of Spinal Muscular Atrophy SMN and SMNΔ7 Proteins Are Degraded by the Proteasome Pathway

Directory of Open Access Journals (Sweden)

Raúl Sánchez-Lanzas

2017-12-01

Full Text Available Spinal muscular atrophy is due to mutations affecting the SMN1 gene coding for the full-length protein (survival motor neuron; SMN and the SMN2 gene that preferentially generates an exon 7-deleted protein (SMNΔ7 by alternative splicing. To study SMN and SMNΔ7 degradation in the cell, we have used tagged versions at the N- (Flag or C-terminus (V5 of both proteins. Transfection of those constructs into HeLa cells and treatment with cycloheximide showed that those protein constructs were degraded. Proteasomal degradation usually requires prior lysine ubiquitylation. Surprisingly, lysine-less variants of both proteins tagged either at N- (Flag or C-terminus (V5 were also degraded. The degradation of the endogenous SMN protein, and the protein constructs mentioned above, was mediated by the proteasome, as it was blocked by lactacystin, a specific and irreversible proteasomal inhibitor. The results obtained allowed us to conclude that SMN and SMNΔ7 proteasomal degradation did not absolutely require internal ubiquitylation nor N-terminal ubiquitylation (prevented by N-terminal tagging. While the above conclusions are firmly supported by the experimental data presented, we discuss and justify the need of deep proteomic techniques for the study of SMN complex components (orphan and bound turn-over to understand the physiological relevant mechanisms of degradation of SMN and SMNΔ7 in the cell.

The VP7 Outer Capsid Protein of Rotavirus Induces Polyclonal B-Cell Activation

Science.gov (United States)

Blutt, Sarah E.; Crawford, Sue E.; Warfield, Kelly L.; Lewis, Dorothy E.; Estes, Mary K.; Conner, Margaret E.

2004-01-01

The early response to a homologous rotavirus infection in mice includes a T-cell-independent increase in the number of activated B lymphocytes in the Peyer's patches. The mechanism of this activation has not been previously determined. Since rotavirus has a repetitively arranged triple-layered capsid and repetitively arranged antigens can induce activation of B cells, one or more of the capsid proteins could be responsible for the initial activation of B cells during infection. To address this question, we assessed the ability of rotavirus and virus-like particles to induce B-cell activation in vivo and in vitro. Using infectious rotavirus, inactivated rotavirus, noninfectious but replication-competent virus, and virus-like particles, we determined that neither infectivity nor RNA was necessary for B-cell activation but the presence of the rotavirus outer capsid protein, VP7, was sufficient for murine B-cell activation. Preincubation of the virus with neutralizing VP7 antibodies inhibited B-cell activation. Polymyxin B treatment and boiling of the virus preparation were performed, which ruled out possible lipopolysaccharide contamination as the source of activation and confirmed that the structural conformation of VP7 is important for B-cell activation. These findings indicate that the structure and conformation of the outer capsid protein, VP7, initiate intestinal B-cell activation during rotavirus infection. PMID:15194774

The herpes simplex virus 1 UL51 protein interacts with the UL7 protein and plays a role in its recruitment into the virion.

Science.gov (United States)

Roller, Richard J; Fetters, Rachel

2015-03-01

The alphaherpesvirus UL51 protein is a tegument component that interacts with the viral glycoprotein E and functions at multiple steps in virus assembly and spread in epithelial cells. We show here that pUL51 forms a complex in infected cells with another conserved tegument protein, pUL7. This complex can form in the absence of other viral proteins and is largely responsible for recruitment of pUL7 to cytoplasmic membranes and into the virion tegument. Incomplete colocalization of pUL51 and pUL7 in infected cells, however, suggests that a significant fraction of the population of each protein is not complexed with the other and that they may accomplish independent functions. The ability of herpesviruses to spread from cell to cell in the face of an immune response is critical for disease and shedding following reactivation from latency. Cell-to-cell spread is a conserved ability of herpesviruses, and the identification of conserved viral genes that mediate this process will aid in the design of attenuated vaccines and of novel therapeutics. The conserved UL51 gene of herpes simplex virus 1 plays important roles in cell-to-cell spread and in virus assembly in the cytoplasm, both of which likely depend on specific interactions with other viral and cellular proteins. Here we identify one of those interactions with the product of another conserved herpesvirus gene, UL7, and show that formation of this complex mediates recruitment of UL7 to membranes and to the virion. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

7 CFR 51.2543 - U.S. Non-Split.

Science.gov (United States)

2010-01-01

... Standards for Grades of Pistachio Nuts in the Shell § 51.2543 U.S. Non-Split. “U.S. Non-Split” consists of non-split pistachio nuts in the shell which meet the following requirements: (a) Basic requirements...

The Verticillium-specific protein VdSCP7 localizes to the plant nucleus and modulates immunity to fungal infections.

Science.gov (United States)

Zhang, Lisha; Ni, Hao; Du, Xuan; Wang, Sheng; Ma, Xiao-Wei; Nürnberger, Thorsten; Guo, Hui-Shan; Hua, Chenlei

2017-07-01

Fungal pathogens secrete effector proteins to suppress plant basal defense for successful colonization. Resistant plants, however, can recognize effectors by cognate R proteins to induce effector-triggered immunity (ETI). By analyzing secretomes of the vascular fungal pathogen Verticillium dahliae, we identified a novel secreted protein VdSCP7 that targets the plant nucleus. The green fluorescent protein (GFP)-tagged VdSCP7 gene with either a mutated nuclear localization signal motif or with additional nuclear export signal was transiently expressed in Nicotiana benthamiana, and investigated for induction of plant immunity. The role of VdSCP7 in V. dahliae pathogenicity was characterized by gene knockout and complementation, and GFP labeling. Expression of the VdSCP7 gene in N. benthamiana activated both salicylic acid and jasmonate signaling, and altered the plant's susceptibility to the pathogens Botrytis cinerea and Phytophthora capsici. The immune response activated by VdSCP7 was highly dependent on its initial extracellular secretion and subsequent nuclear localization in plants. Knockout of the VdSCP7 gene significantly enhanced V. dahliae aggressiveness on cotton. GFP-labeled VdSCP7 is secreted by V. dahliae and accumulates in the plant nucleus. We conclude that VdSCP7 is a novel effector protein that targets the host nucleus to modulate plant immunity, and suggest that plants can recognize VdSCP7 to activate ETI during fungal infection. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Identification of intracellular proteins and signaling pathways in human endothelial cells regulated by angiotensin-(1-7).

Science.gov (United States)

Meinert, Christian; Gembardt, Florian; Böhme, Ilka; Tetzner, Anja; Wieland, Thomas; Greenberg, Barry; Walther, Thomas

2016-01-01

The study aimed to identify proteins regulated by the cardiovascular protective peptide angiotensin-(1-7) and to determine potential intracellular signaling cascades. Human endothelial cells were stimulated with Ang-(1-7) for 1 h, 3 h, 6 h, and 9 h. Peptide effects on intracellular signaling were assessed via antibody microarray, containing antibodies against 725 proteins. Bioinformatics software was used to identify affected intracellular signaling pathways. Microarray data was verified exemplarily by Western blot, Real-Time RT-PCR, and immunohistochemical studies. The microarray identified 110 regulated proteins after 1 h, 119 after 3 h, 31 after 6 h, and 86 after 9 h Ang-(1-7) stimulation. Regulated proteins were associated with high significance to several metabolic pathways like “Molecular Mechanism of Cancer” and “p53 signaling” in a time dependent manner. Exemplarily, Western blots for the E3-type small ubiquitin-like modifier ligase PIAS2 confirmed the microarray data and displayed a decrease by more than 50% after Ang-(1-7) stimulation at 1 h and 3 h without affecting its mRNA. Immunohistochemical studies with PIAS2 in human endothelial cells showed a decrease in cytoplasmic PIAS2 after Ang-(1-7) treatment. The Ang-(1-7) mediated decrease of PIAS2 was reproduced in other endothelial cell types. The results suggest that angiotensin-(1-7) plays a role in metabolic pathways related to cell death and cell survival in human endothelial cells.

Interaction of bacteriophage T4 and T7 single-stranded DNA-binding proteins with DNA

International Nuclear Information System (INIS)

Shokri, Leila; Williams, Mark C; Rouzina, Ioulia

2009-01-01

Bacteriophages T4 and T7 are well-studied model replication systems, which have allowed researchers to determine the roles of many proteins central to DNA replication, recombination and repair. Here we summarize and discuss the results from two recently developed single-molecule methods to determine the salt-dependent DNA-binding kinetics and thermodynamics of the single-stranded DNA (ssDNA)-binding proteins (SSBs) from these systems. We use these methods to characterize both the equilibrium double-stranded DNA (dsDNA) and ssDNA binding of the SSBs T4 gene 32 protein (gp32) and T7 gene 2.5 protein (gp2.5). Despite the overall two-orders-of-magnitude weaker binding of gp2.5 to both forms of DNA, we find that both proteins exhibit four-orders-of-magnitude preferential binding to ssDNA relative to dsDNA. This strong preferential ssDNA binding as well as the weak dsDNA binding is essential for the ability of both proteins to search dsDNA in one dimension to find available ssDNA-binding sites at the replication fork

Assembly of the U5 snRNP component PRPF8 is controlled by the HSP90/R2TP chaperones

Czech Academy of Sciences Publication Activity Database

Malinová, Anna; Cvačková, Zuzana; Matějů, Daniel; Hořejší, Zuzana; Abeza, C.; Vandermoere, F.; Bertrand, E.; Staněk, David; Verheggen, C.

2017-01-01

Roč. 216, č. 6 (2017), s. 1579-1596 ISSN 0021-9525 R&D Projects: GA ČR GPP301/12/P425; GA ČR GA15-00790S; GA ČR(CZ) GA14-34264S; GA MŠk LO1419 Institutional support: RVO:68378050 Keywords : dominant retinitis-pigmentosa * splicing factor prp8 * rna-polymerase-ii * structural basis * spliceosomal snrnps * coiled bodies * cajal bodies * r2tp complex * mutations * biogenesis Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Biochemistry and molecular biology Impact factor: 7.955, year: 2016

Variation in bull beef quality due to ultimate muscle pH is correlated to endopeptidase and small heat shock protein levels.

Science.gov (United States)

Pulford, D J; Dobbie, P; Fraga Vazquez, S; Fraser-Smith, E; Frost, D A; Morris, C A

2009-09-01

This study set out to determine if ultimate pH (pH(u)) affected the performance of intracellular small heat shock protein and endopeptidase dynamics in muscle during beef ageing. Longissimus dorsi muscles from 39 Angus or Limousin×Angus bulls were examined to see if pH(u) achieved at 22h post mortem (rigor) affected tenderness and water holding capacity of beef. Samples were segregated into three pH(u) groups termed high (pH>6.3), intermediate (5.7pHpHpH(u) beef. More than 30% of bull beef did not achieve acceptable tenderness at 8 days post mortem with this ageing regime. No significant differences in calpain or cathepsin enzyme levels due to meat pH were observed until after 22h post mortem, but low pH(u) beef had elevated caspase 3/7 activity soon after slaughter. At 22h post mortem, greater levels of μ-calpain enzyme were found in the high and intermediate pH(u) beef and cathepsin B levels were superior in the low pH(u) beef after 2 days post mortem. Different rates of desmin and troponin T protein degradation were also observed in aged bull beef. Both proteins were degraded within 6h post mortem for high pH(u) beef, but took >3 days post mortem for intermediate pH(u) beef. High levels of alpha β-crystallin (aβC) at 22h post mortem coincided with delayed muscle protein degradation for low pH(u) beef. Our results support the hypothesis that aβC shields myofibrils and buffers against endopeptidase degradation of beef structure during ageing.

The collagen receptor uPARAP/Endo180

DEFF Research Database (Denmark)

Engelholm, Lars H; Ingvarsen, Signe; Jürgensen, Henrik J

2009-01-01

The uPAR-associated protein (uPARAP/Endo180), a type-1 membrane protein belonging to the mannose receptor family, is an endocytic receptor for collagen. Through this endocytic function, the protein takes part in a previously unrecognized mechanism of collagen turnover. uPARAP/Endo180 can bind...... and internalize both intact and partially degraded collagens. In some turnover pathways, the function of the receptor probably involves an interplay with certain matrix-degrading proteases whereas, in other physiological processes, redundant mechanisms involving both endocytic and pericellular collagenolysis seem...... in collagen breakdown seems to be involved in invasive tumor growth Udgivelsesdato: 2009...

Depletion of nuclear import protein karyopherin alpha 7 (KPNA7) induces mitotic defects and deformation of nuclei in cancer cells.

Science.gov (United States)

Vuorinen, Elisa M; Rajala, Nina K; Ihalainen, Teemu O; Kallioniemi, Anne

2018-03-27

Nucleocytoplasmic transport is a tightly regulated process carried out by specific transport machinery, the defects of which may lead to a number of diseases including cancer. Karyopherin alpha 7 (KPNA7), the newest member of the karyopherin alpha nuclear importer family, is expressed at a high level during embryogenesis, reduced to very low or absent levels in most adult tissues but re-expressed in cancer cells. We used siRNA-based knock-down of KPNA7 in cancer cell lines, followed by functional assays (proliferation and cell cycle) and immunofluorescent stainings to determine the role of KPNA7 in regulation of cancer cell growth, proper mitosis and nuclear morphology. In the present study, we show that the silencing of KPNA7 results in a dramatic reduction in pancreatic and breast cancer cell growth, irrespective of the endogenous KPNA7 expression level. This growth inhibition is accompanied by a decrease in the fraction of S-phase cells as well as aberrant number of centrosomes and severe distortion of the mitotic spindles. In addition, KPNA7 depletion leads to reorganization of lamin A/C and B1, the main nuclear lamina proteins, and drastic alterations in nuclear morphology with lobulated and elongated nuclei. Taken together, our data provide new important evidence on the contribution of KPNA7 to the regulation of cancer cell growth and the maintenance of nuclear envelope environment, and thus deepens our understanding on the impact of nuclear transfer proteins in cancer pathogenesis.

Role of Bone Morphogenetic Protein 7 (BMP7 in the Modulation of Corneal Stromal and Epithelial Cell Functions

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Bhavani S. Kowtharapu

2018-05-01

Full Text Available In the cornea, healing of the wounded avascular surface is an intricate process comprising the involvement of epithelial, stromal and neuronal cell interactions. These interactions result to the release of various growth factors that play prominent roles during corneal wound healing response. Bone morphogenetic proteins (BMPs are unique multi-functional potent growth factors of the transforming growth factor-beta (TGF-β superfamily. Treatment of corneal epithelial cells with substance P and nerve growth factor resulted to an increase in the expression of BMP7 mRNA. Since BMP7 is known to modulate the process of corneal wound healing, in this present study, we investigated the influence of exogenous rhBMP7 on human corneal epithelial cell and stromal cell (SFs function. To obtain a high-fidelity expression profiling of activated biomarkers and pathways, transcriptome-wide gene-level expression profiling of epithelial cells in the presence of BMP7 was performed. Gene ontology analysis shows BMP7 stimulation activated TGF-β signaling and cell cycle pathways, whereas biological processes related to cell cycle, microtubule and intermediate filament cytoskeleton organization were significantly impacted in corneal epithelial cells. Scratch wound healing assay showed increased motility and migration of BMP7 treated epithelial cells. BMP7 stimulation studies show activation of MAPK cascade proteins in epithelial cells and SFs. Similarly, a difference in the expression of claudin, Zink finger E-box-binding homeobox 1 was observed along with phosphorylation levels of cofilin in epithelial cells. Stimulation of SFs with BMP7 activated them with increased expression of α-smooth muscle actin. In addition, an elevated phosphorylation of epidermal growth factor receptor following BMP7 stimulation was also observed both in corneal epithelial cells and SFs. Based on our transcriptome analysis data on epithelial cells and the results obtained in SFs, we

Ribonucleoprotein organization of eukaryotic RNA. XXXII. U2 small nuclear RNA precursors and their accurate 3' processing in vitro as ribonucleoprotein particles.

Science.gov (United States)

Wieben, E D; Nenninger, J M; Pederson, T

1985-05-05

Biosynthetic precursors of U2 small nuclear RNA have been identified in cultured human cells by hybrid-selection of pulse-labeled RNA with cloned U2 DNA. These precursor molecules are one to approximately 16 nucleotides longer than mature U2 RNA and contain 2,2,7-trimethylguanosine "caps". The U2 RNA precursors are associated with proteins that react with a monoclonal antibody for antigens characteristic of small nuclear ribonucleoprotein particles. Like previously described precursors of U1 and U4 small nuclear RNAs, the pre-U2 RNAs are recovered in cytoplasmic fractions, although it is not known if this is their location in vivo. The precursors are processed to mature-size U2 RNA when cytoplasmic extracts are incubated in vitro at 37 degrees C. Mg2+ is required but ATP is not. The ribonucleoprotein structure of the pre-U2 RNA is maintained during the processing reaction in vitro, as are the 2,2,7-trimethylguanosine caps. The ribonucleoprotein organization is of major importance, as exogenous, protein-free U2 RNA precursors are degraded rapidly in the in vitro system. Two lines of evidence indicate that the conversion of U2 precursors to mature-size U2 RNA involves a 3' processing reaction. First, the reaction is unaffected by a large excess of mature U2 small nuclear RNP, whose 5' trimethylguanosine caps would be expected to compete for a 5' processing activity. Second, when pre-U2 RNA precursors are first stoichiometrically decorated with an antibody specific for 2,2,7-trimethylguanosine, the extent of subsequent processing in vitro is unaffected. These results provide the first demonstration of a eukaryotic RNA processing reaction in vitro occurring within a ribonucleoprotein particle.

Molecular dynamics simulations of the Nip7 proteins from the marine deep- and shallow-water Pyrococcus species.

Science.gov (United States)

Medvedev, Kirill E; Alemasov, Nikolay A; Vorobjev, Yuri N; Boldyreva, Elena V; Kolchanov, Nikolay A; Afonnikov, Dmitry A

2014-10-15

The identification of the mechanisms of adaptation of protein structures to extreme environmental conditions is a challenging task of structural biology. We performed molecular dynamics (MD) simulations of the Nip7 protein involved in RNA processing from the shallow-water (P. furiosus) and the deep-water (P. abyssi) marine hyperthermophylic archaea at different temperatures (300 and 373 K) and pressures (0.1, 50 and 100 MPa). The aim was to disclose similarities and differences between the deep- and shallow-sea protein models at different temperatures and pressures. The current results demonstrate that the 3D models of the two proteins at all the examined values of pressures and temperatures are compact, stable and similar to the known crystal structure of the P. abyssi Nip7. The structural deviations and fluctuations in the polypeptide chain during the MD simulations were the most pronounced in the loop regions, their magnitude being larger for the C-terminal domain in both proteins. A number of highly mobile segments the protein globule presumably involved in protein-protein interactions were identified. Regions of the polypeptide chain with significant difference in conformational dynamics between the deep- and shallow-water proteins were identified. The results of our analysis demonstrated that in the examined ranges of temperatures and pressures, increase in temperature has a stronger effect on change in the dynamic properties of the protein globule than the increase in pressure. The conformational changes of both the deep- and shallow-sea protein models under increasing temperature and pressure are non-uniform. Our current results indicate that amino acid substitutions between shallow- and deep-water proteins only slightly affect overall stability of two proteins. Rather, they may affect the interactions of the Nip7 protein with its protein or RNA partners.

Resistance to Inhibitors of Cholinesterase 3 (Ric-3 Expression Promotes Selective Protein Associations with the Human α7-Nicotinic Acetylcholine Receptor Interactome.

Directory of Open Access Journals (Sweden)

Matthew J Mulcahy

Full Text Available The α7-nicotinic acetylcholine receptor (α7-nAChR is a ligand-gated ion channel widely expressed in vertebrates and is associated with numerous physiological functions. As transmembrane ion channels, α7-nAChRs need to be expressed on the surface of the plasma membrane to function. The receptor has been reported to associate with proteins involved with receptor biogenesis, modulation of receptor properties, as well as intracellular signaling cascades and some of these associated proteins may affect surface expression of α7-nAChRs. The putative chaperone resistance to inhibitors of cholinesterase 3 (Ric-3 has been reported to interact with, and enhance the surface expression of, α7-nAChRs. In this study, we identified proteins that associate with α7-nAChRs when Ric-3 is expressed. Using α-bungarotoxin (α-bgtx, we isolated and compared α7-nAChR-associated proteins from two stably transfected, human tumor-derived cell lines: SH-EP1-hα7 expressing human α7-nAChRs and the same cell line further transfected to express Ric-3, SH-EP1-hα7-Ric-3. Mass spectrometric analysis of peptides identified thirty-nine proteins that are associated with α7-nAChRs only when Ric-3 was expressed. Significantly, and consistent with reports of Ric-3 function in the literature, several of the identified proteins are involved in biological processes that may affect nAChR surface expression such as post-translational processing of proteins, protein trafficking, and protein transport. Additionally, proteins affecting the cell cycle, the cytoskeleton, stress responses, as well as cyclic AMP- and inositol triphosphate-dependent signaling cascades were identified. These results illuminate how α-bgtx may be used to isolate and identify α7-nAChRs as well as how the expression of chaperones such as Ric-3 can influence proteins associating with α7-nAChRs. These associating proteins may alter activities of α7-nAChRs to expand their functionally-relevant repertoire as

The Sur7 protein regulates plasma membrane organization and prevents intracellular cell wall growth in Candida albicans.

Science.gov (United States)

Alvarez, Francisco J; Douglas, Lois M; Rosebrock, Adam; Konopka, James B

2008-12-01

The Candida albicans plasma membrane plays important roles in cell growth and as a target for antifungal drugs. Analysis of Ca-Sur7 showed that this four transmembrane domain protein localized to stable punctate patches, similar to the plasma membrane subdomains known as eisosomes or MCC that were discovered in S. cerevisiae. The localization of Ca-Sur7 depended on sphingolipid synthesis. In contrast to S. cerevisiae, a C. albicans sur7Delta mutant displayed defects in endocytosis and morphogenesis. Septins and actin were mislocalized, and cell wall synthesis was very abnormal, including long projections of cell wall into the cytoplasm. Several phenotypes of the sur7Delta mutant are similar to the effects of inhibiting beta-glucan synthase, suggesting that the abnormal cell wall synthesis is related to activation of chitin synthase activity seen under stress conditions. These results expand the roles of eisosomes by demonstrating that Sur7 is needed for proper plasma membrane organization and cell wall synthesis. A conserved Cys motif in the first extracellular loop of fungal Sur7 proteins is similar to a characteristic motif of the claudin proteins that form tight junctions in animal cells, suggesting a common role for these tetraspanning membrane proteins in forming specialized plasma membrane domains.

The Sur7 Protein Regulates Plasma Membrane Organization and Prevents Intracellular Cell Wall Growth in Candida albicans

Science.gov (United States)

Alvarez, Francisco J.; Douglas, Lois M.; Rosebrock, Adam

2008-01-01

The Candida albicans plasma membrane plays important roles in cell growth and as a target for antifungal drugs. Analysis of Ca-Sur7 showed that this four transmembrane domain protein localized to stable punctate patches, similar to the plasma membrane subdomains known as eisosomes or MCC that were discovered in S. cerevisiae. The localization of Ca-Sur7 depended on sphingolipid synthesis. In contrast to S. cerevisiae, a C. albicans sur7Δ mutant displayed defects in endocytosis and morphogenesis. Septins and actin were mislocalized, and cell wall synthesis was very abnormal, including long projections of cell wall into the cytoplasm. Several phenotypes of the sur7Δ mutant are similar to the effects of inhibiting β-glucan synthase, suggesting that the abnormal cell wall synthesis is related to activation of chitin synthase activity seen under stress conditions. These results expand the roles of eisosomes by demonstrating that Sur7 is needed for proper plasma membrane organization and cell wall synthesis. A conserved Cys motif in the first extracellular loop of fungal Sur7 proteins is similar to a characteristic motif of the claudin proteins that form tight junctions in animal cells, suggesting a common role for these tetraspanning membrane proteins in forming specialized plasma membrane domains. PMID:18799621

7 CFR 51.300 - U.S. Extra Fancy.

Science.gov (United States)

2010-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... Standards for Grades of Apples Grades § 51.300 U.S. Extra Fancy. “U.S. Extra Fancy” consists of apples of...

7 CFR 51.3053 - U.S. No. 3.

Science.gov (United States)

2010-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... Standards for Florida Avocados Grades § 51.3053 U.S. No. 3. “U.S. No. 3” consists of avocados of similar...

7 CFR 51.3052 - U.S. No. 2.

Science.gov (United States)

2010-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... Standards for Florida Avocados Grades § 51.3052 U.S. No. 2. “U.S. No. 2” consists of avocados of similar...

7 CFR 51.3050 - U.S. No. 1.

Science.gov (United States)

2010-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... Standards for Florida Avocados Grades § 51.3050 U.S. No. 1. “U.S. No. 1” consists of avocados of similar...

Identification and characterization of a phospholipase A1 activity type three secreted protein, PP_ExoU from Pseudomonas plecoglossicida NB2011, the causative agent of visceral granulomas disease in large yellow croaker (Larimichthys crocea).

Science.gov (United States)

Zhang, J; Wang, Y; Guo, H; Mao, Z; Ge, C

2017-06-01

Pseudomonas plecoglossicida NB2011, the causative agent of visceral granulomas disease in farmed Larimichthys crocea in China, encodes a predicted type three effector PP_ExoU, a homolog of the cytotoxin ExoU of Pseudomonas aeruginosa. In this study, secretion of PP_ExoU was tested in various broth, the protein was expressed with the pET30a prokaryotic system, the phospholipase A (PLA) activity of the recombinant protein was determined with fluorogenic phospholipid substrates, fusion expression with green fluorescent protein in transfected HeLa cells was investigated, and the lactate dehydrogenase (LDH) level was measured. The results showed the protein was type three secreted in several media; the recombinant protein displayed significant PLA1 activity with ubiquitin. Fluorescence was observed on the cell membrane and scattered in the cytoplasm of HeLa cells expressing catalytic wild-type PP_ExoU, blebbing and stretching developed in the cell membranes indicating of membrane damage. Fluorescence scattered in the cytoplasm of cells expressing the catalytic inactive protein. A significant LDH level was detected in HeLa cells expressing wild-type PP_exoU, but not in the Ser/Asp-mutated protein, suggestion mutation of predicted catalytic residues abolished the PLA activity. This is the first report on the function of a secreted type three protein from P. plecoglossicida. © 2016 John Wiley & Sons Ltd.

The C-terminal domain of the Arabidopsis AtMBD7 protein confers strong chromatin binding activity

International Nuclear Information System (INIS)

Zemach, Assaf; Paul, Laju K.; Stambolsky, Perry; Efroni, Idan; Rotter, Varda; Grafi, Gideon

2009-01-01

The Arabidopsis MBD7 (AtMBD7) - a naturally occurring poly MBD protein - was previously found to be functional in binding methylated-CpG dinucleotides in vitro and localized to highly methylated chromocenters in vivo. Furthermore, AtMBD7 has significantly lower mobility within the nucleus conferred by cooperative activity of its three MBD motifs. Here we show that besides the MBD motifs, AtMBD7 possesses a strong chromatin binding domain located at its C-terminus designated sticky-C (StkC). Mutational analysis showed that a glutamic acid residue near the C-terminus is essential though not sufficient for the StkC function. Further analysis demonstrated that this motif can render nuclear proteins highly immobile both in plant and animal cells, without affecting their native subnuclear localization. Thus, the C-terminal, StkC motif plays an important role in fastening AtMBD7 to its chromosomal, CpG-methylated sites. It may be possible to utilize this motif for fastening nuclear proteins to their chromosomal sites both in plant and animal cells for research and gene therapy applications.

Hydrophysical Evaluation of Wells TW-B, TW-7, UE-6d, U-2gg PSE 3A, U-10L 1, and UE-6e in Yucca Flat

Energy Technology Data Exchange (ETDEWEB)

Pohlmann, Karl [Desert Research Inst. (DRI), Las Vegas, NV (United States); Healey, John [Desert Research Inst. (DRI), Las Vegas, NV (United States); Lyles, Bred [Desert Research Inst. (DRI), Reno, NV (United States); Cooper, Clay [Desert Research Inst. (DRI), Reno, NV (United States); Hershey, Ronald L. [Desert Research Inst. (DRI), Reno, NV (United States)

2017-05-01

This study evaluated six wells in Yucca Flat in support of the Underground Test Area (UGTA) Activity conducted by the U.S. Department of Energy (DOE) at the Nevada National Security Site (NNSS). Accessibility and groundwater sampling conditions were assessed and if conditions permitted, samples were collected for tritium analysis. Four of the wells, TW-B, UE-6d, UE-6e, and TW-7 were sampled in support of UGTA responses to recommendations made by the Yucca Flat/Climax Mine External Peer Review Committee (Navarro, 2016). In addition to its role in support of these responses, TW-7 was included because it is listed in the NNSS Integrated Groundwater Sampling Plan (DOE, 2014) as a required sampling location, although it had not been sampled since 1994. U-2gg PSE 3A and U-10L 1 were evaluated to determine whether deteriorating well conditions can be addressed so that these wells can be used as additional sampling points in Yucca Flat.

7 CFR 51.302 - U.S. No. 1.

Science.gov (United States)

2010-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... Standards for Grades of Apples Grades § 51.302 U.S. No. 1. “U.S. No. 1” consists of apples which meet the...

Hsc66 substrate specificity is directed toward a discrete region of the iron-sulfur cluster template protein IscU.

Science.gov (United States)

Hoff, Kevin G; Ta, Dennis T; Tapley, Tim L; Silberg, Jonathan J; Vickery, Larry E

2002-07-26

Hsc66 and Hsc20 comprise a specialized chaperone system important for the assembly of iron-sulfur clusters in Escherchia coli. Only a single substrate, the Fe/S template protein IscU, has been identified for the Hsc66/Hsc20 system, but the mechanism by which Hsc66 selectively binds IscU is unknown. We have investigated Hsc66 substrate specificity using phage display and a peptide array of IscU. Screening of a heptameric peptide phage display library revealed that Hsc66 prefers peptides with a centrally located Pro-Pro motif. Using a cellulose-bound peptide array of IscU we determined that Hsc66 interacts specifically with a region (residues 99-103, LPPVK) that is invariant among all IscU family members. A synthetic peptide (ELPPVKIHC) corresponding to IscU residues 98-106 behaves in a similar manner to native IscU, stimulating the ATPase activity of Hsc66 with similar affinity as IscU, preventing Hsc66 suppression of bovine rhodanese aggregation, and interacting with the peptide-binding domain of Hsc66. Unlike native IscU, however, the synthetic peptide is not bound by Hsc20 and does not synergistically stimulate Hsc66 ATPase activity with Hsc20. Our results indicate that Hsc66 and Hsc20 recognize distinct regions of IscU and further suggest that Hsc66 will not bind LPPVK motifs with high affinity in vivo unless they are in the context of native IscU and can be directed to Hsc66 by Hsc20.

Expression of human protein S100A7 (psoriasin, preparation of antibody and application to human larynx squamous cell carcinoma

Directory of Open Access Journals (Sweden)

Barbieri Manuela R

2011-11-01

Full Text Available Abstract Background Up-regulation of S100A7 (Psoriasin, a small calcium-binding protein, is associated with the development of several types of carcinomas, but its function and possibility to serve as a diagnostic or prognostic marker have not been fully defined. In order to prepare antibodies to the protein for immunohistochemical studies we produced the recombinant S100A7 protein in E. coli. mRNA extracted from human tracheal tumor tissue which was amplified by RT-PCR to provide the region coding for the S100A7 gene. The amplified fragment was cloned in the vector pCR2.1-TOPO and sub-cloned in the expression vector pAE. The protein rS100A7 (His-tag was expressed in E. coli BL21::DE3, purified by affinity chromatography on an Ni-NTA column, recovered in the 2.0 to 3.5 mg/mL range in culture medium, and used to produce a rabbit polyclonal antibody anti-rS100A7 protein. The profile of this polyclonal antibody was evaluated in a tissue microarray. Results The rS100A7 (His-tag protein was homogeneous by SDS-PAGE and mass spectrometry and was used to produce an anti-recombinant S100A7 (His-tag rabbit serum (polyclonal antibody anti-rS100A7. The molecular weight of rS100A7 (His-tag protein determined by linear MALDI-TOF-MS was 12,655.91 Da. The theoretical mass calculated for the nonapeptide attached to the amino terminus is 12,653.26 Da (delta 2.65 Da. Immunostaining with the polyclonal anti-rS100A7 protein generated showed reactivity with little or no background staining in head and neck squamous cell carcinoma cells, detecting S100A7 both in nucleus and cytoplasm. Lower levels of S100A7 were detected in non-neoplastic tissue. Conclusions The polyclonal anti-rS100A7 antibody generated here yielded a good signal-to-noise contrast and should be useful for immunohistochemical detection of S100A7 protein. Its potential use for other epithelial lesions besides human larynx squamous cell carcinoma and non-neoplastic larynx should be explored in future.

7 CFR 51.1150 - U.S. No. 3.

Science.gov (United States)

2010-01-01

...) Sunburn; and (18) Other means. (d) For tolerances see § 51.1151. (e) Internal quality: Lots meeting the internal requirements for “U.S. Grade AA Juice (Double A)” or “U.S. Grade A Juice” may be so specified in... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

PENGARUH SERBUK U-Mo HASIL PROSES MEKANIK DAN HYDRIDE – DEHYDRIDE – GRINDING MILL TERHADAP KUALITAS PELAT ELEMEN BAKAR U-Mo/Al

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Supardjo Supardjo

2015-07-01

Full Text Available PENGARUH SERBUK U-Mo HASIL PROSES MEKANIK DAN HYDRIDE – DEHYDRIDE – GRINDING MILL TERHADAP KUALITAS PELAT ELEMEN BAKAR U-Mo/Al. Penelitian bahan bakar U-7Mo/Al tipe pelat dilakukan dalam rangka pengembangan bahan bakar U3Si2/Al untuk mendapatkan bahan bakar baru yang memiliki densitas uranium lebih tinggi, stabil selama digunakan sebagai bahan bakar di dalam reaktor dan mudah dilakukan proses olah ulangnya. Lingkup penelitian meliputi pembuatan: paduan U-7Mo dengan teknik peleburan, pembuatan serbuk U-7Mo dengan dikikir dan hydride - dehydride - grinding mill, IEB U-7Mo/Al dengan teknik kompaksi pada tekanan 20 bar, dan PEB U-7Mo/Al dengan teknik pengerolan panas pada temperatur 425oC. Paduan U-7Mo hasil proses peleburan cukup homogen, berat jenis 16,34 g/cm3 dan bersifat ulet, kemudian dibuat menjadi serbuk dengan cara dikikir dan hydride - dehydride - grinding mill. Serbuk U-7Mo hasil proses kikir berbentuk pipih, kontaminan Fe cukup tinggi, sedangkan serbuk hasil proses hydride - dehydride - grinding mill, cenderung equiaxial dengan kontaminan yang rendah. Kedua jenis serbuk U-7Mo tersebut digunakan sebagai bahan baku pembuatan IEB U-7Mo/Al dan PEB U-7Mo/Al dengan densitas uranium 7 gU/cm3 dan diperoleh produk dengan kualitas yang hampir sama. Hasil uji IEB U-7Mo/Al berukuran 25 x 15 x 3,15±0,05 mm, tidak terdapat cacat/retak, distribusi U-7Mo di dalam matriks cukup homogen dan tidak terdapat pengelompokan/aglomerasi U-7Mo yang berdimensi >1 mm. PEB U-7Mo/Al hasil pengerolan dengan tebal akhir 1,45 mm, memiliki ketebalan meat rerata 0,60 mm dan tebal kelongsong 0,4 mm dan terdapat 1 titik pengukuran kelongsong dengan ketebalan 0,15 mm. Dengan membandingkan penggunaan kedua jenis serbuk U-7Mo tersebut, IEB U-7Mo/Al dan PEB U-7Mo/Al yang dihasilkan memiliki kualitas hampir sama. Namun demikian penggunaan serbuk U- 7Mo hasil proses hydride - dehydride - grinding mill lebih baik karena proses pengerjaannya lebih cepat dan impuritas dalam

Rhythmic expression of DEC2 protein in vitro and in vivo.

Science.gov (United States)

Sato, Fuyuki; Muragaki, Yasuteru; Kawamoto, Takeshi; Fujimoto, Katsumi; Kato, Yukio; Zhang, Yanping

2016-06-01

Basic helix-loop-helix (bHLH) transcription factor DEC2 (bHLHE41/Sharp1) is one of the clock genes that show a circadian rhythm in various tissues. DEC2 regulates differentiation, sleep length, tumor cell invasion and apoptosis. Although studies have been conducted on the rhythmic expression of DEC2 mRNA in various tissues, the precise molecular mechanism of DEC2 expression is poorly understood. In the present study, we examined whether DEC2 protein had a rhythmic expression. Western blot analysis for DEC2 protein revealed a rhythmic expression in mouse liver, lung and muscle and in MCF-7 and U2OS cells. In addition, AMP-activated protein kinase (AMPK) activity (phosphorylation of AMPK) in mouse embryonic fibroblasts (MEFs) exhibited a rhythmic expression under the condition of medium change or glucose-depleted medium. However, the rhythmic expression of DEC2 in MEF gradually decreased in time under these conditions. The medium change affected the levels of DEC2 protein and phosphorylation of AMPK. In addition, the levels of DEC2 protein showed a rhythmic expression in vivo and in MCF-7 and U2OS cells. The results showed that the phosphorylation of AMPK immunoreactivity was strongly detected in the liver and lung of DEC2 knockout mice compared with that of wild-type mice. These results may provide new insights into rhythmic expression and the regulation between DEC2 protein and AMPK activity.

Uniform {sup 15}N- and {sup 15}N/{sup 13}C-labeling of proteins in mammalian cells and solution structure of the amino terminal fragment of u-PA

Energy Technology Data Exchange (ETDEWEB)

Hansen, A.P.; Petros, A.M.; Meadows, R.P.; Mazar, A.P.; Nettesheim, D.G.; Pederson, T.M.; Fesik, S.W. [Abbott Laboratories, Abbott Park, IL (United States)

1994-12-01

Urokinase-type plasminogen activator (u-PA) is a 54-kDa glycoprotein that catalyzes the conversion of plasminogen to plasmin, a broad-specificity protease responsible for the degradation of fibrin clots and extracellular matrix components. The u-PA protein consists of three individual modules: a growth factor domain (GFD), a kringle, and a serine protease domain. The amino terminal fragment (ATF) includes the GFD-responsible for u-PA binding to its receptor-and the kringle domains. This protein was expressed and uniformly {sup 15}N-and {sup 15}N/{sup 13}C-labeled in mammalian cells by methods that will be described. In addition, we present the three-dimensional structure of ATF that was derived from 1299 NOE-derived distance restraints along with the {phi} angle and hydrogen bonding restraints. Although the individual domains in the structures were highly converged, the two domains are structurally independent. The overall structures of the individual domains are very similar to the structures of homologous proteins. However, important structural differences between the growth factor domain of u-PA and other homologous proteins were observed in the region that has been implicated in binding the urokinase receptor. These results may explain, in part, why other growth factors show no appreciable affinity for the urokinase receptor.

Spliceosomal protein U1A is involved in alternative splicing and salt stress tolerance in Arabidopsis thaliana

KAUST Repository

Gu, Jinbao

2017-12-01

Soil salinity is a significant threat to sustainable agricultural production worldwide. Plants must adjust their developmental and physiological processes to cope with salt stress. Although the capacity for adaptation ultimately depends on the genome, the exceptional versatility in gene regulation provided by the spliceosome-mediated alternative splicing (AS) is essential in these adaptive processes. However, the functions of the spliceosome in plant stress responses are poorly understood. Here, we report the in-depth characterization of a U1 spliceosomal protein, AtU1A, in controlling AS of pre-mRNAs under salt stress and salt stress tolerance in Arabidopsis thaliana. The atu1a mutant was hypersensitive to salt stress and accumulated more reactive oxygen species (ROS) than the wild-type under salt stress. RNA-seq analysis revealed that AtU1A regulates AS of many genes, presumably through modulating recognition of 5′ splice sites. We showed that AtU1A is associated with the pre-mRNA of the ROS detoxification-related gene ACO1 and is necessary for the regulation of ACO1 AS. ACO1 is important for salt tolerance because ectopic expression of ACO1 in the atu1a mutant can partially rescue its salt hypersensitive phenotype. Our findings highlight the critical role of AtU1A as a regulator of pre-mRNA processing and salt tolerance in plants.

Estradiol attenuates EGF-induced rapid uPAR mobilization and cell migration via the G-protein-coupled receptor 30 in ovarian cancer cells

DEFF Research Database (Denmark)

Henic, Emir; Noskova, Vera; Høyer-Hansen, Gunilla

2009-01-01

: rapid mobilization of uPAR from detergent-resistant domains, increased mRNA, and decreased degradation. G-protein-coupled receptor 30 (GPR30) is a newly identified membrane estrogen receptor (ER).The objective of this study was to explore the effects of 17beta-estradiol (E(2)) on uPAR expression...... for ERalpha, and quantitative polymerase chain reaction. Estradiol attenuates the stimulatory effect of EGF on cell migration and uPAR expression. Specifically, E(2) reduces the very rapid increase of detergent extractable uPAR, which occurs within minutes of EGF stimulation and probably represents...... agonist G1, mimicked the effect of E(2) on uPAR expression and cell migration. OVCAR-3 cells express mRNA for GPR30.Estradiol attenuates EGF-induced mobilization of ligated uPAR from detergent-resistant domains and subsequent migration in ovarian cancer cells. The response to various ER ligands indicates...

Double demonstration of oncogenic high risk human papilloma virus DNA and HPV-E7 protein in oral cancers.

Science.gov (United States)

Pannone, G; Santoro, A; Carinci, F; Bufo, P; Papagerakis, S M; Rubini, C; Campisi, G; Giovannelli, L; Contaldo, M; Serpico, R; Mazzotta, M; Lo Muzio, L

2011-01-01

Oncogenic HPVs are necessarily involved in cervical cancer but their role in oral carcinogenesis is debated. To detect HPV in oral cancer, 38 cases of formalin fixed-paraffin embedded OSCC were studied by both DNA genotyping (MY09/11 L1 consensus primers in combination with GP5-GP6 primer pair followed by sequencing) and immunohistochemistry (monoclonal Abs against capsid protein and HPV-E7 protein, K1H8 DAKO and clone 8C9 INVITROGEN, respectively). HPV-16 tonsil cancer was used as positive control. The overall prevalence of HPV infection in OSCCs was 10.5%. Amplification of DNA samples showed single HPV DNA infection in 3 cases (HPV16; HPV53; HPV70) and double infection in one case of cheek cancer (HPV31/HPV44). The overall HR-HPV prevalence was 7.5%. E-7 antigen was immunohistochemically detected in all HPV-positive cases. HPV+ OSCC cases showed an overall better outcome than HPV negative oral cancers, as evaluated by Kaplan-Meier curves. HPVs exert their oncogenic role after DNA integration, gene expression of E5, E6 and E7 loci and p53/pRb host proteins suppression. This study showed that HPV-E7 protein inactivating pRb is expressed in oral cancer cells infected by oncogenic HPV other than classical HR-HPV-16/18. Interestingly HPV-70, considered a low risk virus with no definite collocation in oncogenic type category, gives rise to the expression of HPV-E7 protein and inactivate pRb in oral cancer. HPV-70, as proved in current literature, is able to inactivates also p53 protein, promoting cell immortalization. HPV-53, classified as a possible high risk virus, expresses E7 protein in OSCC, contributing to oral carcinogenesis. We have identified among OSCCs, a subgroup characterized by HPV infection (10.5%). Finally, we have proved the oncogenic potential of some HPV virus types, not well known in literature.

Development and characterization of monolithic fuel miniplate alloy U-2.5Zr-7.5Nb, coated in zircaloy; Desenvolvimento e caracterizacao do combustivel nuclear tipo placa monolitico da liga U-2,5Zr-7,5Nb revestido em zircaloy

Energy Technology Data Exchange (ETDEWEB)

Machado, Geraldo Correa

2014-06-01

The autocthonal production of nuclear fuel in Brazil for test and research reactors is restricted to MTR (Material Test Reactor) fuel type dispersion plate, using U3Si2 alloy, coated and dispersed in aluminum, developed by IPEN-SP for use in IEA-R1 reactor. Moreover, the UO{sub 2} fuel rod type for power reactors is manufactured by Rezende (RJ) with a German technology by INB under license. Currently, Brazil is performing two programs of developing reactors. Currently, Brazil is developing two reactors. One of them is the development, by CNEN, the Brazilian Multipurpose Reactor (RMB), for testing, research and radioisotope production. The other one is the development a power reactor for naval propulsion, conducted by the Brazilian Navy. This dissertation presents the development and characterization of monolithic fuel miniplate alloy U-2.5Zr-7.5Nb, coated in zircaloy (ZRY), on a laboratory scale. Due to its innovative features and properties, this fuel can be used as fuel in both test reactors, research and producing radioisotopes for power reactors as small and medium sizes. Thus, this high potential fuel can be used in domestic reactors currently under development. The development of monolithic fuel plate type is made using the technique called 'picture-frame' where a sandwich composed of a monolith alloy U-2.5Zr- 7.5Nb coupled to a frame and coated sheets of Zry is obtained. The alloy U-2.5Zr-7.5Nb was obtained by melting in an induction furnace and then was cast into rectangular ingots of graphite, thus achieving an ingot with approximate dimensions of 170 x 50 x 60 mm. The obtained ingot was hot rolled at 850 ºC, with a 50 % reduction in thickness, in order to refine the raw structure of fusion. Samples cut from the alloy U-2.5Zr-7.5Nb, with dimensions 20 x 20 x 6 mm were placed in frames and plates Zry and joined by TIG (Tungsten Inert Gas) under an atmosphere of argon, obtaining a set of 10 mm thick, 45 mm wide and 100 mm long. The sandwiches were

Surface expression and subunit specific control of steady protein levels by the Kv7.2 helix A-B linker.

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Paloma Aivar

Full Text Available Kv7.2 and Kv7.3 are the main components of the neuronal voltage-dependent M-current, which is a subthreshold potassium conductance that exerts an important control on neuronal excitability. Despite their predominantly intracellular distribution, these channels must reach the plasma membrane in order to control neuronal activity. Thus, we analyzed the amino acid sequence of Kv7.2 to identify intrinsic signals that may control its surface expression. Removal of the interlinker connecting helix A and helix B of the intracellular C-terminus produces a large increase in the number of functional channels at the plasma membrane. Moreover, elimination of this linker increased the steady-state amount of protein, which was not associated with a decrease of protein degradation. The magnitude of this increase was inversely correlated with the number of helix A - helix B linkers present in the tetrameric channel assemblies. In contrast to the remarkable effect on the amount of Kv7.2 protein, removal of the Kv7.2 linker had no detectable impact on the steady-state levels of Kv7.3 protein.

238U, 234U and 232Th in seawater

International Nuclear Information System (INIS)

Chen, J.H.; Edwards, R.L.; Wasserburg, G.J.

1986-01-01

We have developed techniques to determine 238 U, 234 U and 232 Th concentrations in seawater by isotope dilution mass spectrometry. Using these techniques, we have measured 238 U, 234 U and 232 Th in vertical profiles of unfiltered, acidified seawater from the Atlantic and 238 U and 234 U in vertical profiles from the Pacific. Determinations of 234 U/ 238 U at depths ranging from 0 to 4900 m in the Atlantic (7 0 44'N, 40 0 43'W) and the Pacific (14 0 41'N, 160 0 01'W) Oceans are the same within experimental error (±5per mille, 2σ). The average of these 234 U/ 238 U measurements is 144±2per mille (2σ) higher than the equilibrium ratio of 5.472 x 10 -5 . U concentrations, normalized to 35per mille salinity, range from 3.162 to 3.281 ng/g, a range of 3.8%. The average concentration of the Pacific samples (31 0 4'N, 159 0 1'W) is ∝1% higher than that of the Atlantic (7 0 44'N, 40 0 43'W and 31 0 49'N, 64 0 6'W). 232 Th concentrations from an Atlantic profile range from 0.092 to 0.145 pg/g. The observed constancy of the 234 U/ 238 U ratio is consistent with the predicted range of 234 U/ 238 U using a simple two-box model and the residence time of deep water in the ocean determined from 14 C. The variation in salinity-normalized U concentrations suggests that U may be much more reactive in the marine environment than previously thought. (orig./WB)

KARAKTERISASI PROTEIN MIOFIBRIL DARI IKAN KUNIRAN (Upeneus moluccensis DAN IKAN MATA BESAR (Selar crumenophthalmus [Characterization of Myofibrillar Protein from Goldband Goat Fish (Upeneus moluccensis and Bigeye Scad Fish (Selar crumenophthalmus

Directory of Open Access Journals (Sweden)

Achmad Subagio

2004-04-01

Full Text Available Characteristics of myofibrillar protein from goldband goat fish (U. moluccensis and bigeye scad fish (S. crumenophthalmus were studied for their development as food ingredient. Color analysis using chromameter showed that myfibrillar protein from goldband goat fish was light colored, while that of bigeye scad was slightly drak colored. Proximate analysis showed that their contents were similar by crude protein 7-10%, crude fat 0.2-0.5%, and ash 0.4-0.7%. Amino acid compositions of both myofibrillar proteins were very close, dominated by glutamic acid (20%, aspartic acid (10% and lysine (9%. However, comparing with bigeye scad, myofibrillar proteins from goldband goatfish were easily aggregated, had higher gelation capacity and higher emulsion activity, but lower solubility. Based on these result, myofibrillar protein from goldband goatfish has good charachteristics as food ingredient especially for restructured products comparing with bigeye scad

Urokinase receptor-associated protein (uPARAP) is expressed in connection with malignant as well as benign lesions of the human breast and occurs in specific populations of stromal cells

DEFF Research Database (Denmark)

Schnack Nielsen, Boye; Rank, Fritz; Engelholm, Lars H

2002-01-01

The urokinase-type plasminogen activator (uPA) and the uPA receptor (uPAR) are key components in the plasminogen activation system, serving to promote specific events of extracellular matrix degradation in connection with tissue remodeling and cancer invasion. We recently described a new u......PAR-associated protein (uPARAP), an internalization receptor that interacts with the pro-uPA:uPAR complex. In our study, we generated a specific polyclonal peptide antibody against human uPARAP and used it for the localization of uPARAP in different breast lesions. The affinity-purified antibodies specifically...

Guanine nucleotide-binding protein subunit beta-2-like 1, a new Annexin A7 interacting protein

International Nuclear Information System (INIS)

Du, Yue; Meng, Jinyi; Huang, Yuhong; Wu, Jun; Wang, Bo; Ibrahim, Mohammed M.; Tang, Jianwu

2014-01-01

Highlights: • RACK1 formed a complex with Annexin A7. • Depletion of RACK1 inhibited the proliferation, migration and invasion. • RACK1 RNAi abolished RACK1-Annexin A7 interaction. • RACK1-Annexin A7 may play a role in regulating the metastatic potentials. - Abstract: We report for the first time that Guanine nucleotide-binding protein subunit beta-2-like 1 (RACK1) formed a complex with Annexin A7. Hca-F and Hca-P are a pair of syngeneic mouse hepatocarcinoma cell lines established and maintained in our laboratory. Our previous study showed that both Annexin A7 and RACK1 were expressed higher in Hca-F (lymph node metastasis >70%) than Hca-P (lymph node metastasis <30%). Suppression of Annexin A7 expression in Hca-F cells induced decreased migration and invasion ability. In this study, knockdown of RACK1 by RNA interference (RNAi) had the same impact on metastasis potential of Hca-F cells as Annexin A7 down-regulation. Furthermore, by co-immunoprecipitation and double immunofluorescence confocal imaging, we found that RACK1 was in complex with Annexin A7 in control cells, but not in the RACK1-down-regulated cells, indicating the abolishment of RACK1-Annexin A7 interaction in Hca-F cells by RACK1 RNAi. Taken together, these results suggest that RACK1-Annexin A7 interaction may be one of the means by which RACK1 and Annexin A7 influence the metastasis potential of mouse hepatocarcinoma cells in vitro

Structure and biochemical function of a prototypical Arabidopsis U-box domain

DEFF Research Database (Denmark)

Andersen, Pernille; Kragelund, Birthe B; Olsen, Addie N

2004-01-01

U-box proteins, as well as other proteins involved in regulated protein degradation, are apparently over-represented in Arabidopsis compared with other model eukaryotes. The Arabidopsis protein AtPUB14 contains a typical U-box domain followed by an Armadillo repeat region, a domain organization t...

The Plasmodium falciparum exported protein PF3D7_0402000 binds to erythrocyte ankyrin and band 4.1

Energy Technology Data Exchange (ETDEWEB)

Shakya, Bikash; Penn, Wesley D.; Nakayasu, Ernesto S.; Lacount, Douglas J.

2017-09-01

Plasmodium falciparum extensively modifies the infected red blood cell (RBC), resulting in changes in deformability, shape and surface properties. These alterations suggest that the RBC cytoskeleton is a major target for modification during infection. However, the molecular mechanisms leading to these changes are largely unknown. To begin to address this question, we screened for exported P. falciparum proteins that bound to the erythrocyte cytoskeleton proteins ankyrin 1 (ANK1) and band 4.1 (4.1R), which form critical interactions with other cytoskeletal proteins that contribute to the deformability and stability of RBCs. Yeast two-hybrid screens with ANK1 and 4.1R identified eight interactions with P. falciparum exported proteins, including an interaction between 4.1R and PF3D7_0402000 (PFD0090c). This interaction was first identified in a large-scale screen (Vignali et al., Malaria J, 7:211, 2008), which also reported an interaction between PF3D7_0402000 and ANK1. We confirmed the interactions of PF3D7_0402000 with 4.1R and ANK1 in pair-wise yeast two-hybrid and co-precipitation assays. In both cases, an intact PHIST domain in PF3D7_0402000 was required for binding. Complex purification followed by mass spectrometry analysis provided additional support for the interaction of PF3D7_0402000 with ANK1 and 4.1R. RBC ghost cells loaded with maltose-binding protein (MBP)-PF3D7_0402000 passed through a metal microsphere column less efficiently than mock- or MBP-loaded controls, consistent with an effect of PF3D7_0402000 on RBC rigidity or membrane stability. This study confirmed the interaction of PF3D7_0402000 with 4.1R in multiple independent assays, provided the first evidence that PF3D7_0402000 also binds to ANK1, and suggested that PF3D7_0402000 affects deformability or membrane stability of uninfected RBC ghosts.

Interdiffusion between U(Mo,Pt) or U(Mo,Zr) and Al or Al A356 alloy

International Nuclear Information System (INIS)

Komar Varela, C.; Mirandou, M.; Arico, S.; Balart, S.; Gribaudo, L.

2009-01-01

Solid state reactions in chemical diffusion couples U-7 wt.%Mo-0.9 wt.%Pt/Al at 580 deg. C and U-7 wt.%Mo-0.9 wt.%Pt/Al A356 alloy, U-7 wt.%Mo-1 wt.%Zr/Al and U-7 wt.%Mo-1 wt.%Zr/Al A356 alloy at 550 deg. C were characterized. Results were obtained from optical and scanning electron microscopy, electron probe microanalysis and X-ray diffraction. The UAl 3, UAl 4 and Al 20 Mo 2 U phases were identified in the interaction layers of γU(Mo,Pt)/Al and γU(Mo,Zr)/Al diffusion couples. Al 43 Mo 4 U 6 ternary compound was also identified in γU(Mo,Zr)/Al due to the decomposition of γU(Mo,Zr) phase. The U(Al,Si) 3 and U 3 Si 5 phases were identified in the interaction layers of γU(Mo,Pt)/Al A356 and γU(Mo,Zr)/Al A356 diffusion couples. These phases are formed due to the migration of Si to the interaction layer. In the diffusion couple U(Mo,Zr)/Al A356, Zr 5 Al 3 phase was also identified in the interaction layer. The use of synchrotron radiation at Brazilian Synchrotron Light Laboratory (LNLS, CNPq, Campinas, Brazil) was necessary to achieve a complete crystallographic characterization.

Search for KPNA7 cargo proteins in human cells reveals MVP and ZNF414 as novel regulators of cancer cell growth.

Science.gov (United States)

Vuorinen, Elisa M; Rajala, Nina K; Rauhala, Hanna E; Nurminen, Anssi T; Hytönen, Vesa P; Kallioniemi, Anne

2017-01-01

Karyopherin alpha 7 (KPNA7) belongs to a family of nuclear import proteins that recognize and bind nuclear localization signals (NLSs) in proteins to be transported to the nucleus. Previously we found that KPNA7 is overexpressed in a subset of pancreatic cancer cell lines and acts as a critical regulator of growth in these cells. This characteristic of KPNA7 is likely to be mediated by its cargo proteins that are still mainly unknown. Here, we used protein affinity chromatography in Hs700T and MIA PaCa-2 pancreatic cancer cell lines and identified 377 putative KPNA7 cargo proteins, most of which were known or predicted to localize to the nucleus. The interaction was confirmed for two of the candidates, MVP and ZNF414, using co-immunoprecipitation, and their transport to the nucleus was hindered by siRNA based KPNA7 silencing. Most importantly, silencing of MVP and ZNF414 resulted in marked reduction in Hs700T cell growth. In conclusion, these data uncover two previously unknown human KPNA7 cargo proteins with distinct roles as novel regulators of pancreatic cancer cell growth, thus deepening our understanding on the contribution of nuclear transport in cancer pathogenesis. Copyright © 2016 Elsevier B.V. All rights reserved.

Optical spectroscopy and crystal-field analysis of U{sup 3+}: Ba{sub 2}YCl{sub 7}

Energy Technology Data Exchange (ETDEWEB)

Karbowiak, M.; Mech, A.; Drozdzyndki, J. [Wroclaw Univ., Faculty of Chemistry (Poland); Gajek, Z. [Polish Academy of Sciences, W. Trzebiatowski Institute of Low Temperature and Structure Research, Wroclaw (Poland); Edelstein, N.M. [Lawrence Berkeley National Lab., Chemical Sciences Div., CA (United States)

2002-11-01

High resolution absorption spectra of a U{sup 3+}(0.3%): Ba{sub 2}YCl{sub 7} single crystal were recorded in the 4000-50 000 cm{sup -1} range at 7 K. The observed crystal-field levels were assigned and fit to the parameters of the simplified angular overlap model (AOM) as well as a semi-empirical Hamiltonian representing the combined atomic and one-electron crystal-field interactions. The starting values of the AOM parameters were obtained from ab initio calculations. The analysis of the spectra allowed the assignment of 65 crystal-field levels with a relatively small rms deviation of 25 cm{sup -1} and has shown that the AOM approach can predict quite well the B{sub q}{sup k} crystal-field parameters. The value determined for the crystal-field strength parameter, N{sub v}, corresponds well with those determined for U{sup 3+} in other chloride single crystals. (authors)

Extraction chromatography of U(VI) and Pu(IV) adsorbed on amberlite XAD-7/dibutyloctanamide

International Nuclear Information System (INIS)

Prabhu, D.R.; Mahajan, G.R.; Nair, G.M.; Subramanian, M.S.

1992-01-01

The adsorption of U(VI) and Pu(IV) into the neutral poly acrylic resin Amberlite XAD-7, impregnated with dibutyloctanamide was found to be maximum at around 6M HNO 3 . Both these ions were found to be adsorbed as their monosolvates. The thermodynamic parameters obtained from the data at different temperatures indicated that the adsorption reaction was enthalpy favoured and entropy disfavored. (author). 5 refs., 1 tab

7 CFR 1499.7 - Transportation of goods.

Science.gov (United States)

2010-01-01

... 7 Agriculture 10 2010-01-01 2010-01-01 false Transportation of goods. 1499.7 Section 1499.7... Transportation of goods. (a) Shipments of donated commodities are subject to the requirements of 46 U.S.C. 55305 and 55314, regarding carriage on U.S.-flag vessels. (b) Transportation of donated commodities and...

7 CFR 1599.7 - Transportation of goods.

Science.gov (United States)

2010-01-01

... 7 Agriculture 10 2010-01-01 2010-01-01 false Transportation of goods. 1599.7 Section 1599.7... Transportation of goods. (a) Shipments of donated commodities are subject to the requirements of 46 U.S.C. 55305 and 55314, regarding carriage on U.S.-flag vessels. (b) Transportation of donated commodities and...

Rebalance between 7S and 11S globulins in soybean seeds of differing protein content and 11SA4.

Science.gov (United States)

Yang, A; Yu, X; Zheng, A; James, A T

2016-11-01

Protein content and globulin subunit composition of soybean seeds affect the quality of soy foods. In this proteomic study, the protein profile of soybean seeds with high (∼45.5%) or low (∼38.6%) protein content and with or without the glycinin (11S) subunit 11SA4 was examined. 44 unique proteins and their homologues were identified and showed that both protein content and 11SA4 influenced the abundance of a number of proteins. The absence of 11SA4 exerted a greater impact than the protein content, and led to a decreased abundance of glycinin G2/A2B1 and G5/A5A4B3 subunits, which resulted in lower total 11S with a concomitant higher total β-conglycinin (7S). Low protein content was associated with higher glycinin G3/A1aB1b and lower glycinin G4/A5A4B3. Using the proteomic approach, it was demonstrated that 11SA4 deficiency induced compensatory accumulation of 7S globulins and led to a similar total abundance for 7S+11S irrespective of protein content or 11SA4. Copyright © 2016 Elsevier Ltd. All rights reserved.

康奈尔净碳水化合物-蛋白质体系预测小肠可消化粗蛋白质含量%Predicting Utilizable Crude Protein Content in Small Intestine by Cornell Net Carbohydrate and Protein System

Institute of Scientific and Technical Information of China (English)

王燕; 杨方; 陈常栋; 王海威; 张微微; 张永根

2012-01-01

This study was conducted to establish mathematical models of predicting utilizable crude protein (uCP) content in feed for dairy cows using Cornell net carbohydrate and protein system (CNCPS). Contents of protein fractions of 13 kinds of common feeds were analyzed using CNCPS, and uCP of feeds was determined using mobile nylon bag method, and three Holstein dairy cows with permanent ruminal and duodenal cannulas were used in a single factor experimental design. The results showed as follows: 1) contents of uCP in soybean meal, cottonseed meal, rapeseed meal, sunflower seed meal, sesame meal, corn germ meal, rice bran, rice bran cake, rice bran meal, barley, wheat bran, corn and corn gluten feed were 390. 32, 321. 90, 297.21, 230.50, 388.62, 177.49, 85.53, 116.78, 134.74, 80.47, 128.26, 70.28 and 66. 65 g/kg, respectively. 2) The regression equation of contents of measured uCP and CNCPS fractions in all experimental feeds was uCP = -4.11 + 6.48PA+7.73PB, + 5.72PB2 +8.26PB3 +5. UPC (R2 =0.997 2, P<0.01) ; the regression equation of contents of measured uCP and CNCPS fractions in protein feed was uCP =12. 79 + 5. 47PA +7. 04PBl +9. 74PB2 +8. 14PB3 (R2 = 0. 998 7, P < 0. 01) ; the regression equation of contents of measured uCP and CNCPS fractions in energy feed was uCP = 14. 80 +8. 55PA +6. 27PB2 +17. 64PB3(R2 = 0.987 4, P<0.01). 3) Regression coefficients (R2) between CNCPS fractions contents and measured uCP contents in protein feed, energy feed and all experimental feed were all higher than 0. 95, and regression coefficient between measured uCP content and CNCPS fractions contents in protein feed was higher than that in energy feed. In conclusion, it is feasible that using contents of protein components subdivided by CNCPS to predict uCP content, and the prediction result in protein feed is more accurate than that in energy feed.%本试验旨在建立奶牛饲料的康奈尔净碳水化合物-蛋白质体系(Cornell net carbohydrate and protein system,CNCPS)预测小肠可消化粗蛋白质(u

Direct association between the Ret receptor tyrosine kinase and the Src homology 2-containing adapter protein Grb7.

Science.gov (United States)

Pandey, A; Liu, X; Dixon, J E; Di Fiore, P P; Dixit, V M

1996-05-03

Adapter proteins containing Src homology 2 (SH2) domains link transmembrane receptor protein-tyrosine kinases to downstream signal transducing molecules. A family of SH2 containing adapter proteins including Grb7 and Grb10 has been recently identified. We had previously shown that Grb10 associates with Ret via its SH2 domain in an activation-dependent manner (Pandey, A., Duan, H., Di Fiore, P.P., and Dixit, V.M. (1995) J. Biol, Chem. 270, 21461-21463). We now demonstrate that the related adapter molecule Grb7 also associates with Ret in vitro and in vivo, and that the binding of the SH2 domain of Grb7 to Ret is direct. This binding is dependent upon Ret autophosphorylation since Grb7 is incapable of binding a kinase-defective mutant of Ret. Thus two members of the Grb family, Grb7 and Grb10, likely relay signals emanating from Ret to other, as yet, unidentified targets within the cell.

Functional analyses of GB virus B p13 protein: development of a recombinant GB virus B hepatitis virus with a p7 protein

DEFF Research Database (Denmark)

Takikawa, Shingo; Engle, Ronald E; Emerson, Suzanne U

2006-01-01

GB virus B (GBV-B), which infects tamarins, is the virus most closely related to hepatitis C virus (HCV). HCV has a protein (p7) that is believed to form an ion channel. It is critical for viability. In vitro studies suggest that GBV-B has an analogous but larger protein (p13). We found...... plus part of p7) was nonviable. However, a mutant lacking amino acid 614-669 (p6) produced high titer viremia and acute resolving hepatitis; viruses recovered from both animals lacked the deleted sequence and had no other mutations. Thus, p6 was dispensable but p7 was essential for infectivity...... processing at both sites, suggesting that p13 is processed into two components (p6 and p7). Mutants with substitution at amino acid 669 or 681 were viable in vivo, but the recovered viruses had changes at amino acid 669 and 681, respectively, which restored cleavage. A mutant lacking amino acid 614-681 (p6...

Functional analyses of GB virus B p13 protein: Development of a recombinant GB virus B hepatitis virus with a p7 protein

DEFF Research Database (Denmark)

Takikawa, Shingo; Engle, Ronald E; Emerson, Suzanne U

2006-01-01

GB virus B (GBV-B), which infects tamarins, is the virus most closely related to hepatitis C virus (HCV). HCV has a protein (p7) that is believed to form an ion channel. It is critical for viability. In vitro studies suggest that GBV-B has an analogous but larger protein (p13). We found...... plus part of p7) was nonviable. However, a mutant lacking amino acid 614-669 (p6) produced high titer viremia and acute resolving hepatitis; viruses recovered from both animals lacked the deleted sequence and had no other mutations. Thus, p6 was dispensable but p7 was essential for infectivity...... processing at both sites, suggesting that p13 is processed into two components (p6 and p7). Mutants with substitution at amino acid 669 or 681 were viable in vivo, but the recovered viruses had changes at amino acid 669 and 681, respectively, which restored cleavage. A mutant lacking amino acid 614-681 (p6...

Kinetic study of the thermal decomposition of uranium metaphosphate, U(PO{sub 3}){sub 4}, into uranium pyrophosphate, UP{sub 2}O{sub 7}

Energy Technology Data Exchange (ETDEWEB)

Yang, Hee-Chul, E-mail: nhcyang@kaeri.re.kr; Kim, Hyung-Ju; Lee, Si-Young; Yang, In-Hwan; Chung, Dong-Yong

2017-06-15

The thermochemical properties of uranium compounds have attracted much interest in relation to thermochemical treatments and the safe disposal of radioactive waste bearing uranium compounds. The characteristics of the thermal decomposition of uranium metaphosphate, U(PO{sub 3}){sub 4}, into uranium pyrophosphate, UP{sub 2}O{sub 7}, have been studied from the view point of reaction kinetics and acting mechanisms. A mixture of U(PO{sub 3}){sub 4} and UP{sub 2}O{sub 7} was prepared from the pyrolysis residue of uranium-bearing spent TBP. A kinetic analysis of the reaction of U(PO{sub 3}){sub 4} into UP{sub 2}O{sub 7} was conducted using an isoconversional method and a master plot method on the basis of data from a non-isothermal thermogravimetric analysis. The thermal decomposition of U(PO{sub 3}){sub 4} into UP{sub 2}O{sub 7} followed a single-step reaction with an activation energy of 175.29 ± 1.58 kJ mol{sup −1}. The most probable kinetic model was determined as a type of nucleation and nuclei-growth models, the Avrami-Erofeev model (A3), which describes that there are certain restrictions on nuclei growth of UP{sub 2}O{sub 7} during the solid-state decomposition of U(PO{sub 3}){sub 4}. - Highlights: •Thermal decomposition kinetics of U(PO{sub 3}){sub 4} into UP{sub 2}O{sub 7} was investigated. •The thermal decomposition followed a single-step reaction with an activation energy of 175.3 ± 1.6 kJ mol{sup −1}. •The most probable kinetic model was determined as a type of nucleation and nuclei-growth models, the Avrami-Erofeev (A3).

Spinal 5-HT7 Receptors and Protein Kinase A Constrain Intermittent Hypoxia-Induced Phrenic Long-term Facilitation

Science.gov (United States)

Hoffman, M.S.; Mitchell, G.S.

2013-01-01

Phrenic long-term facilitation (pLTF) is a form of serotonin-dependent respiratory plasticity induced by acute intermittent hypoxia (AIH). pLTF requires spinal Gq protein-coupled serotonin-2 receptor (5-HT2) activation, new synthesis of brain-derived neurotrophic factor (BDNF) and activation of its high-affinity receptor, TrkB. Intrathecal injections of selective agonists for Gs protein-coupled receptors (adenosine 2A and serotonin-7; 5-HT7) also induce long-lasting phrenic motor facilitation via TrkB “trans-activation.” Since serotonin release near phrenic motor neurons may activate multiple serotonin receptor subtypes, we tested the hypothesis that 5-HT7 receptor activation contributes to AIH-induced pLTF. A selective 5-HT7 receptor antagonist (SB-269970, 5mM, 12μl) was administered intrathecally at C4 to anesthetized, vagotomized and ventilated rats prior to AIH (3, 5-min episodes, 11% O2). Contrary to predictions, pLTF was greater in SB-269970 treated versus control rats (80±11% vs 45±6% 60 min post-AIH; p<0.05). Hypoglossal LTF was unaffected by spinal 5-HT7 receptor inhibition, suggesting that drug effects were localized to the spinal cord. Since 5-HT7 receptors are coupled to protein kinase A (PKA), we tested the hypothesis that PKA inhibits AIH-induced pLTF. Similar to 5-HT7 receptor inhibition, spinal PKA inhibition (KT-5720, 100μM, 15μl) enhanced pLTF (99±15% 60 min post-AIH; p<0.05). Conversely, PKA activation (8-br-cAMP, 100μM, 15μl) blunted pLTF versus control rats (16±5% vs 45±6% 60 min post-AIH; p<0.05). These findings suggest a novel mechanism whereby spinal Gs protein-coupled 5-HT7 receptors constrain AIH-induced pLTF via PKA activity. PMID:23850591

Methyl isobutyl ketone exposure-related increases in specific measures of α2u-globulin (α2u) nephropathy in male rats along with in vitro evidence of reversible protein binding

International Nuclear Information System (INIS)

Borghoff, S.J.; Poet, T.S.; Green, S.; Davis, J.; Hughes, B.; Mensing, T.; Sarang, S.S.; Lynch, A.M.; Hard, G.C.

2015-01-01

Chronic exposure to methyl isobutyl ketone (MIBK) resulted in an increase in the incidence of renal tubule adenomas and occurrence of renal tubule carcinomas in male, but not female Fischer 344 rats. Since a number of chemicals have been shown to cause male rat renal tumors through the α2u nephropathy-mediated mode of action, the objective of this study is to evaluate the ability of MIBK to induce measures of α2u nephropathy including renal cell proliferation in male and female F344 rats following exposure to the same inhalation concentrations used in the National Toxicology Program (NTP) cancer bioassay (0, 450, 900, or 1800 ppm). Rats were exposed 6 h/day for 1 or 4 weeks and kidneys excised approximately 18 h post exposure to evaluate hyaline droplet accumulation (HDA), α2u staining of hyaline droplets, renal cell proliferation, and to quantitate renal α2u concentration. There was an exposure-related increase in all measures of α2u nephropathy in male, but not female rat kidneys. The hyaline droplets present in male rat kidney stained positively for α2u. The changes in HDA and α2u concentration were comparable to D-limonene, an acknowledged inducer of α2u nephropathy. In a separate in vitro study using a two-compartment vial equilibration model to assess the interaction between MIBK and α2u, the dissociation constant (K d ) was estimated to be 1.27 × 10 −5 M. This K d is within the range of other chemicals known to bind to α2u and cause nephropathy. Together, the exposure-related increase in measures of α2u nephropathy, sustained increase in renal cell proliferation along with an indication of reversible binding of MIBK to α2u, support the inclusion of MIBK in the category of chemicals exerting renal effects through a protein droplet α2u nephropathy-mediated mode of action (MoA)

Probing slowly exchanging protein systems via {sup 13}C{sup {alpha}}-CEST: monitoring folding of the Im7 protein

Energy Technology Data Exchange (ETDEWEB)

Hansen, Alexandar L.; Bouvignies, Guillaume; Kay, Lewis E., E-mail: kay@pound.med.utoronto.ca [University of Toronto, Departments of Molecular Genetics, Biochemistry and Chemistry (Canada)

2013-03-15

A {sup 13}C{sup {alpha}} chemical exchange saturation transfer based experiment is presented for the study of protein systems undergoing slow interconversion between an 'observable' ground state and one or more 'invisible' excited states. Here a labeling strategy whereby [2-{sup 13}C]-glucose is the sole carbon source is exploited, producing proteins with {sup 13}C at the C{sup {alpha}} position, while the majority of residues remain unlabeled at CO or C{sup {beta}}. The new experiment is demonstrated with an application to the folding reaction of the Im7 protein that involves an on-pathway excited state. The obtained excited state {sup 13}C{sup {alpha}} chemical shifts are cross validated by comparison to values extracted from analysis of CPMG relaxation dispersion profiles, establishing the utility of the methodology.

Development and characterization of monolithic fuel miniplate alloy U-2.5Zr-7.5Nb, coated in zircaloy

International Nuclear Information System (INIS)

Machado, Geraldo Correa

2014-01-01

The autocthonal production of nuclear fuel in Brazil for test and research reactors is restricted to MTR (Material Test Reactor) fuel type dispersion plate, using U3Si2 alloy, coated and dispersed in aluminum, developed by IPEN-SP for use in IEA-R1 reactor. Moreover, the UO 2 fuel rod type for power reactors is manufactured by Rezende (RJ) with a German technology by INB under license. Currently, Brazil is performing two programs of developing reactors. Currently, Brazil is developing two reactors. One of them is the development, by CNEN, the Brazilian Multipurpose Reactor (RMB), for testing, research and radioisotope production. The other one is the development a power reactor for naval propulsion, conducted by the Brazilian Navy. This dissertation presents the development and characterization of monolithic fuel miniplate alloy U-2.5Zr-7.5Nb, coated in zircaloy (ZRY), on a laboratory scale. Due to its innovative features and properties, this fuel can be used as fuel in both test reactors, research and producing radioisotopes for power reactors as small and medium sizes. Thus, this high potential fuel can be used in domestic reactors currently under development. The development of monolithic fuel plate type is made using the technique called 'picture-frame' where a sandwich composed of a monolith alloy U-2.5Zr- 7.5Nb coupled to a frame and coated sheets of Zry is obtained. The alloy U-2.5Zr-7.5Nb was obtained by melting in an induction furnace and then was cast into rectangular ingots of graphite, thus achieving an ingot with approximate dimensions of 170 x 50 x 60 mm. The obtained ingot was hot rolled at 850 ºC, with a 50 % reduction in thickness, in order to refine the raw structure of fusion. Samples cut from the alloy U-2.5Zr-7.5Nb, with dimensions 20 x 20 x 6 mm were placed in frames and plates Zry and joined by TIG (Tungsten Inert Gas) under an atmosphere of argon, obtaining a set of 10 mm thick, 45 mm wide and 100 mm long. The sandwiches were hot rolled to

Analyses of pea necrotic yellow dwarf virus-encoded proteins.

Science.gov (United States)

Krenz, Björn; Schießl, Ingrid; Greiner, Eva; Krapp, Susanna

2017-06-01

Pea necrotic yellow dwarf virus (PNYDV) is a multipartite, circular, single-stranded DNA plant virus. PNYDV encodes eight proteins and the function of three of which remains unknown-U1, U2, and U4. PNYDV proteins cellular localization was analyzed by GFP tagging and bimolecular fluorescence complementation (BiFC) studies. The interactions of all eight PNYDV proteins were tested pairwise in planta (36 combinations in total). Seven interactions were identified and two (M-Rep with CP and MP with U4) were characterized further. MP and U4 complexes appeared as vesicle-like spots and were localized at the nuclear envelope and cell periphery. These vesicle-like spots were associated with the endoplasmatic reticulum. In addition, a nuclear localization signal (NLS) was mapped for U1, and a mutated U1 with NLS disrupted localized at plasmodesmata and therefore might also have a role in movement. Taken together, this study provides evidence for previously undescribed nanovirus protein-protein interactions and their cellular localization with novel findings not only for those proteins with unknown function, but also for characterized proteins such as the CP.

Sodium arsenite alters cell cycle and MTHFR, MT1/2, and c-Myc protein levels in MCF-7 cells

International Nuclear Information System (INIS)

Ruiz-Ramos, Ruben; Lopez-Carrillo, Lizbeth; Albores, Arnulfo; Hernandez-Ramirez, Raul U.; Cebrian, Mariano E.

2009-01-01

There is limited available information on the effects of arsenic on enzymes participating in the folate cycle. Therefore, our aim was to evaluate the effects of sodium arsenite on the protein levels of methylenetetrahydrofolate reductase (MTHFR) and dihydrofolate reductase (DHFR) and its further relationship with the expression MT1/2 and c-myc in MCF-7 cells. Arsenite treatment (0-10 μM) for 4 h decreased MTHFR levels in a concentration-dependent fashion without significant effects on DHFR. The effects on MTHFR were observed at arsenite concentrations not significantly affecting cell viability. We also observed an increase in S-phase recruitment at all concentrations probed. Lower concentrations (< 5 μM) induced cell proliferation, showing a high proportion of BrdU-stained cells, indicating a higher DNA synthesis rate. However, higher concentrations (≥ 5 μM) or longer treatment periods induced apoptosis. Arsenite also induced dose-dependent increases in MT1/2 and c-Myc protein levels. The levels of MTHFR were inversely correlated to MT1/2 and c-Myc overexpression and increased S-phase recruitment. Our findings indicate that breast epithelial cells are responsive to arsenite and suggest that exposure may pose a risk for breast cancer. The reductions in MTHFR protein levels contribute to understand the mechanisms underlying the induction of genes influencing growth regulation, such as c-myc and MT1/2. However, further research is needed to ascertain if the effects here reported following short-time and high-dose exposure are relevant for human populations chronically exposed to low arsenic concentrations.

Erwinia amylovora Enfeksiyonu Sonrası Elma, Armut ve Ayva Çeşitlerinde Konukçu Protein Miktarlarının Belirlenmesi

Directory of Open Access Journals (Sweden)

Şerife Çetin

2014-11-01

Full Text Available Ateş yanıklığı hastalığına neden olan Erwinia amylovora, Rosaceae familyasından başta armut, elma ve ayvalarda zararlı bakteriyel bir patojendir. Bu çalışmada, E. amylovora’ nın 2 virülent izolatının (Ea234-1 ve Ea240-3 farklı elma (Braeburn, Fuji, Gala ve Golden Delicious, armut (Santa Maria ve Williams ve ayva (Eşme ve Ekmek çeşitlerinde enfeksiyonu sonucu zamana bağlı olarak toplam protein miktarlarının belirlenmesi amaçlanmıştır. E. amylovora (108 hücre ml-1 ile yaprak inokulasyonundan sonraki 24, 36 ve 72. saatlerde örnekler alınmıştır. Enfeksiyonların doğrulanması için, bakteri inokule edilen bitkilerden re-izolasyonlar yapılmış ve etmen biyokimyasal ve moleküler testlerle E. amylovora olarak tanılanmıştır. Toplam protein miktarlarının belirlenmesinde Bradford yöntemi ve SDS-PAGE analizleri için Laemmli yöntemleri kullanılmış, protein ekstraktlarının 595 nm’ de absorbans değerleri elde edilmiştir. Elde edilen bulgulara göre, elma çeşitlerinde E. amylovora enfeksiyonu sonrası kontrole kıyasla 24. saatte toplam protein miktarının arttığı, 36. ve 72. saatte miktarın azaldığı, Braeburn çeşidinin en yüksek protein miktarına sahip olduğu belirlenmiştir. Armut çeşitlerinde, 24. ve 36. saatlerde toplam protein miktarı artarken, 72. saatte protein miktarının azaldığı ve Santa Maria çeşidinin en yüksek protein miktarına sahip olduğu tespit edilmiştir. Ayva çeşitlerinde, toplam protein miktarının 72. saatte arttığı ve Eşme çeşidinin en yüksek protein miktarına sahip olduğu belirlenmiştir. SDS-PAGE analizi sonucunda, farklı molekül ağırlıklarına sahip protein fraksiyonları elde edilmiştir. Elma ve ayva çeşitlerinde sırasıyla yaklaşık 55-70 ve 35-55 kDa, armut çeşitlerinde ise yaklaşık 55-70 kDa molekül ağırlığında protein bantları belirlenmiştir.

Preparation of 199Tl radionuclide on U-120 (R-7M) cyclotron

International Nuclear Information System (INIS)

Glukhov, G.G.; Komov, A.I.; Maslennikov, Yu.S.; Malinin, A.B.; Skuridin, V.S.

1989-01-01

The possibility of preparation of 199 Tl radionuclide, which can be successfully used instead of thallium-201 in radiopharmaceutical compound for medicine diagnosis due to nuclear-physical characteristics, is studied. It is established that thallium-199 free from thallium-298, but with thallium-200 impurity, is formed under irradiation of gold-197 target by 27.2-28 MeV energy α-particles. The construction of a thin-layer gold target allowing to decrease thallium-200 impurity up to >0.5 % is developed and tested. Perspectivity of thallium-199 production at domestic cyclotrons P-7M and u-120 is shown. 4 refs.; 2 figs.; 3 tabs

Protein-enriched familiar foods and drinks improve protein intake of hospitalized older patients: A randomized controlled trial.

Science.gov (United States)

Beelen, Janne; Vasse, Emmelyne; Janssen, Nancy; Janse, André; de Roos, Nicole M; de Groot, Lisette C P G M

2017-05-18

Adequate protein intake is important in preventing and treating undernutrition. Hospitalized older patients are recommended to consume 1.2-1.5 g of protein per kg body weight per day (g/kg/d) but most of them fail to do so. Therefore, we investigated whether a range of newly developed protein-enriched familiar foods and drinks were effective in increasing protein intake of hospitalized older patients. This randomized controlled trial involved 147 patients of ≥65 years (mean age: 78.5 ± 7.4 years). The control group (n = 80) received the standard energy and protein rich hospital menu. The intervention group (n = 67) received the same menu with various protein-enriched intervention products replacing regular products or added to the menu. Macronutrient intake on the fourth day of hospitalization, based on food ordering data, was compared between the two groups by using Independent T-tests and Mann Whitney U-tests. In the intervention group 30% of total protein was provided by the intervention products. The intervention group consumed 105.7 ± 34.2 g protein compared to 88.2 ± 24.4 g in the control group (p intervention group than in the control group reached a protein intake of 1.2 g/kg/d (79.1% vs 47.5%). Protein intake was significantly higher in the intervention group at breakfast, during the morning between breakfast and lunch, and at dinner. This study shows that providing protein-enriched familiar foods and drinks, as replacement of regular products or as additions to the hospital menu, better enables hospitalized older patients to reach protein intake recommendations. This trial is registered on ClinicalTrials.gov, Identifier: NCT02213393. Copyright © 2017 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

Biochemical and Structural Analysis of Hormone-sensitive Lipase Homolog EstE7: Insight into the Stabilized Dimerization of HSL-Homolog Proteins

International Nuclear Information System (INIS)

Nam, Ki Hyun; Park, Sung Ha; Lee, Won Ho; Hwang, Kwang Yeon

2010-01-01

Hormone sensitive lipase (HSL) plays a major role in energy homeostasis and lipid metabolism. Several crystal structures of HSL-homolog proteins have been identified, which has led to a better understanding of its molecular function. HSLhomolog proteins exit as both monomer and dimer, but the biochemical and structural basis for such oligomeric states has not been successfully elucidated. Therefore, we determined the crystal structure of HSL-homolog protein EstE7 from a metagenome library at 2.2 A resolution and characterized the oligomeric states of EstE7 both structurally and biochemically. EstE7 protein prefers the dimeric state in solution, which is supported by its higher enzymatic activity in the dimeric state. In the crystal form, EstE7 protein shows two-types of dimeric interface. Specifically, dimerization via the external β8-strand occurred through tight association between two pseudosymmetric folds via salt bridges, hydrogen bonds and van der Waals interactions. This dimer formation was similar to that of other HSL-homolog protein structures such as AFEST, BEFA, and EstE1. We anticipate that our results will provide insight into the oligomeric state of HSLhomolog proteins

Binding of 7-dehydrocholesterol to sterol carrier protein and vitamin D3 effect

International Nuclear Information System (INIS)

Takase, Sachiko; Oizumi, Kumiko; Moriuchi, Sachiko; Hosoya, Norimasa

1975-01-01

It was confirmed that deltasup(5,7)-sterol delta 7 -reductase activity was suppressed by cholecalciferol (vitamin D 3 ) in the enzyme system consisted of microsomes and sterol carrier protein (SCP). The enzyme activity was significantly decreased in the combination with microsomes obtained from either vitamin D-deficient or vitamin D 3 -treated rat liver and with SCP obtained from vitamin D 3 -treated rat. It was also demonstrated by the binding assay of the dextran-charcoal technique that 7-dehydrocholesterol binding to SCP could be specifically displaced by vitamin D 3 . The inhibition of cholecalciferol on 7-dehydro-cholesterol binding to liver SCP was confirmed to be non-competitive inhibition. (auth.)

Unlipidated Outer Membrane Protein Omp16 (U-Omp16) from Brucella spp. as Nasal Adjuvant Induces a Th1 Immune Response and Modulates the Th2 Allergic Response to Cow’s Milk Proteins

Science.gov (United States)

Ibañez, Andrés E.; Smaldini, Paola; Coria, Lorena M.; Delpino, María V.; Pacífico, Lucila G. G.; Oliveira, Sergio C.; Risso, Gabriela S.; Pasquevich, Karina A.; Fossati, Carlos Alberto; Giambartolomei, Guillermo H.; Docena, Guillermo H.; Cassataro, Juliana

2013-01-01

The discovery of novel mucosal adjuvants will help to develop new formulations to control infectious and allergic diseases. In this work we demonstrate that U-Omp16 from Brucella spp. delivered by the nasal route (i.n.) induced an inflammatory immune response in bronchoalveolar lavage (BAL) and lung tissues. Nasal co-administration of U-Omp16 with the model antigen (Ag) ovalbumin (OVA) increased the amount of Ag in lung tissues and induced OVA-specific systemic IgG and T helper (Th) 1 immune responses. The usefulness of U-Omp16 was also assessed in a mouse model of food allergy. U-Omp16 i.n. administration during sensitization ameliorated the hypersensitivity responses of sensitized mice upon oral exposure to Cow’s Milk Protein (CMP), decreased clinical signs, reduced anti-CMP IgE serum antibodies and modulated the Th2 response in favor of Th1 immunity. Thus, U-Omp16 could be used as a broad Th1 mucosal adjuvant for different Ag formulations. PMID:23861971

Cardiovascular Small Heat Shock Protein HSPB7 Is a Kinetically Privileged Reactive Electrophilic Species (RES) Sensor.

Science.gov (United States)

Surya, Sanjna L; Long, Marcus J C; Urul, Daniel A; Zhao, Yi; Mercer, Emily J; EIsaid, Islam M; Evans, Todd; Aye, Yimon

2018-02-08

Small heat shock protein (sHSP)-B7 (HSPB7) is a muscle-specific member of the non-ATP-dependent sHSPs. The precise role of HSPB7 is enigmatic. Here, we disclose that zebrafish Hspb7 is a kinetically privileged sensor that is able to react rapidly with native reactive electrophilic species (RES), when only substoichiometric amounts of RES are available in proximity to Hspb7 expressed in living cells. Among the two Hspb7-cysteines, this RES sensing is fulfilled by a single cysteine (C117). Purification and characterizations in vitro reveal that the rate for RES adduction is among the most efficient reported for protein-cysteines with native carbonyl-based RES. Covalent-ligand binding is accompanied by structural changes (increase in β-sheet-content), based on circular dichroism analysis. Among the two cysteines, only C117 is conserved across vertebrates; we show that the human ortholog is also capable of RES sensing in cells. Furthermore, a cancer-relevant missense mutation reduces this RES-sensing property. This evolutionarily conserved cysteine-biosensor may play a redox-regulatory role in cardioprotection.

Structural Basis for the Binding of the Neutralizing Antibody, 7D11, to the Poxvirus L1 Protein

Science.gov (United States)

2007-08-01

pCR- 7D11-vHC and pCR-7D11- vLC , respectively. Crystallization of the complex between L1 and 7D11-Fab VACV L1 protein was expressed and purified as...2005. Vaccinia virus H3L envelope protein is a major target of neutralizing antibodies in humans and elicits protection against lethal challenge in...D.M., Schmaljohn, C., Schmaljohn, A., 2000. DNA vaccination with vaccinia virus L1R and A33R genes protects mice against a lethal poxvirus challenge

Urokinase-type plasminogen activator (uPA) stimulates triglyceride synthesis in Huh7 hepatoma cells via p38-dependent upregulation of DGAT2.

Science.gov (United States)

Paland, Nicole; Gamliel-Lazarovich, Aviva; Coleman, Raymond; Fuhrman, Bianca

2014-11-01

The liver is the central organ of fatty acid and triglyceride metabolism. Oxidation and synthesis of fatty acids and triglycerides is under the control of peroxisome-proliferator-activated receptors (PPAR) α. Impairment of these receptors' function contributes to the accumulation of triglycerides in the liver resulting in non-alcoholic fatty liver disease. Urokinase-type plasminogen activator (uPA) was shown to regulate gene expression in the liver involving PPARγ transcriptional activity. In this study we questioned whether uPA modulates triglyceride metabolism in the liver, and investigated the mechanisms involved in the observed processes. Huh7 hepatoma cells were incubated with increasing concentrations of uPA for 24 h uPA dose-dependently increased the cellular triglyceride mass, and this effect resulted from increased de novo triglyceride synthesis mediated by the enzyme diglyceride acyltransferase 2 (DGAT2). Also, the amount of free fatty acids was highly up regulated by uPA through activation of the transcription factor SREBP-1. Chemical activation of PPARα further increased uPA-stimulated triglyceride synthesis, whereas inhibition of p38, an upstream activator of PPARα, completely abolished the stimulatory effect of uPA on both triglyceride synthesis and DGAT2 upregulation. The effect of uPA on triglyceride synthesis in Huh7 cells was mediated via binding to its receptor, the uPAR. In vivo studies in uPAR(-/-) mice demonstrated that no lipid droplets were observed in their livers compared to C57BL/6 mice and the triglyceride levels were significantly lower. This study presents a new biological function of the uPA/uPAR system in the metabolism of triglycerides and might present a new target for an early therapeutic intervention for NAFLD. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Effect of lattice distortion on uranium magnetic moments in U4Ru7Ge6 studied by polarized neutron diffraction

Science.gov (United States)

Vališka, Michal; Klicpera, Milan; Doležal, Petr; Fabelo, Oscar; Stunault, Anne; Diviš, Martin; Sechovský, Vladimír

2018-03-01

In a cubic ferromagnet, small spontaneous lattice distortions are expected below the Curie temperature, but the phenomenon is usually neglected. This study focuses on such an effect in the U4Ru7Ge6 compound. Based on DFT calculations, we propose a lattice distortion from the cubic I m -3 m space group to a lower, rhombohedral, symmetry described by the R -3 m space group. The strong spin-orbit coupling of the uranium ions plays an essential role in lowering the symmetry, giving rise to two different U sites (U1 and U2). Using polarized neutron diffraction in applied magnetic fields of 1 and 9 T in the ordered state (1.9 K ) and in the paramagnetic state (20 K ), we bring convincing experimental evidence of this splitting of the U sites, with different magnetic moments. The data have been analyzed both by maximum entropy calculations and by a direct fit in the dipolar approximation. In the ordered phase, the μL/μS ratio of the orbital and spin moments on the U2 site is remarkably lower than for the free U3 + or U4 + ion, which points to a strong hybridization of the U 5 f wave functions with the 4 d wave functions of the surrounding Ru. On the U1 site, the μL/μS ratio exhibits an unexpectedly low value: the orbital moment is almost quenched, like in metallic α -uranium. As a further evidence of the 5 f -4 d hybridization in the U4Ru7Ge6 system, we observe the absence of a magnetic moment on the Ru1 site, but a rather large induced moment on the Ru2 site, which is in closer coordination with both U positions. Very similar results are obtained at 20 K in the ferromagnetic regime induced by the magnetic field of 9 T . This shows that applying a strong magnetic field above the Curie temperature also leads to the splitting of the uranium sites, which further demonstrates the intimate coupling of the magnetic ordering and structural distortion. We propose that the difference between the magnetic moment on the U1 and U2 sites results from the strong spin

Production of recombinant proteins GST L1, E6 and E7 tag HPV 16 ...

African Journals Online (AJOL)

STORAGESEVER

2009-02-04

Feb 4, 2009 ... targeting the viral oncoproteins E6 and E7 are markers for HPV-associated ... Luminex XYP plate handler, Luminex SD sheath fluid delivery system, a Pentium 4 .... expression mediated by a potato virus X derived vector of the E7 protein .... inflammation, and antioxidant nutrients – assessing their roles as.

7 CFR 51.1001 - U.S. Combination.

Science.gov (United States)

2010-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... grade requirements only because of blanching shall be designated as “U.S. Combination, Mixed Color”: And...

Structural basis for antagonizing a host restriction factor by C7 family of poxvirus host-range proteins

OpenAIRE

Meng, Xiangzhi; Krumm, Brian; Li, Yongchao; Deng, Junpeng; Xiang, Yan

2015-01-01

Productive viral replication requires overcoming many barriers posed by the host innate immune system. Human sterile alpha motif domain-containing 9 (SAMD9) is a newly identified antiviral factor that is specifically targeted by poxvirus proteins belonging to the C7 family of host-range factors. Here we provide the first, to our knowledge, atomic view of two functionally divergent proteins from the C7 family and determine the molecular basis that dictates whether they can target SAMD9 effecti...

Protein kinase D2 regulates migration and invasion of U87MG glioblastoma cells in vitro

Energy Technology Data Exchange (ETDEWEB)

Bernhart, Eva; Damm, Sabine; Wintersperger, Andrea [Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz (Austria); DeVaney, Trevor [Institute of Biophysics, Medical University of Graz (Austria); Zimmer, Andreas [Institute of Pharmaceutical Sciences, Department of Pharmaceutical Technology, Karl-Franzens University, Graz (Austria); Raynham, Tony; Ireson, Christopher [Cancer Research Technology Ltd, London (United Kingdom); Sattler, Wolfgang, E-mail: wolfgang.sattler@medunigraz.at [Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz (Austria)

2013-08-01

Glioblastoma multiforme (GBM) is the most common malignant brain tumor, which, despite combined modality treatment, reoccurs and is invariably fatal for affected patients. Recently, a member of the serine/threonine protein kinase D (PRKD) family, PRKD2, was shown to be a potent mediator of glioblastoma growth. Here we studied the role of PRKD2 in U87MG glioblastoma cell migration and invasion in response to sphingosine-1-phosphate (S1P), an activator of PRKD2 and a GBM mitogen. Time-lapse microscopy demonstrated that random cell migration was significantly diminished in response to PRKD2 silencing. The pharmacological PRKD family inhibitor CRT0066101 decreased chemotactic migration and invasion across uncoated or matrigel-coated Transwell inserts. Silencing of PRKD2 attenuated migration and invasion of U87MG cells even more effectively. In terms of downstream signaling, CRT0066101 prevented PRKD2 autophosphorylation and inhibited p44/42 MAPK and to a smaller extent p54/46 JNK and p38 MAPK activation. PRKD2 silencing impaired activation of p44/42 MAPK and p54/46 JNK, downregulated nuclear c-Jun protein levels and decreased c-Jun{sup S73} phosphorylation without affecting the NFκB pathway. Finally, qPCR array analyses revealed that silencing of PRKD2 downregulates mRNA levels of integrin alpha-2 and -4 (ITGA2 and -4), plasminogen activator urokinase (PLAU), plasminogen activator urokinase receptor (PLAUR), and matrix metallopeptidase 1 (MMP1). Findings of the present study identify PRKD2 as a potential target to interfere with glioblastoma cell migration and invasion, two major determinants contributing to recurrence of glioblastoma after multimodality treatment. Highlights: • Sphingosine-1-phosphate induces glioma cell migration and invasion. • Part of the effects is mediated by protein kinase D2 (PRKD2) activation. • Inactivation of PRKD2 attenuates glioblastoma cell migration and invasion. • Both, RNAi and pharmacological inhibition of PRKD2 inhibits MAPK

Protein kinase D2 regulates migration and invasion of U87MG glioblastoma cells in vitro

International Nuclear Information System (INIS)

Bernhart, Eva; Damm, Sabine; Wintersperger, Andrea; DeVaney, Trevor; Zimmer, Andreas; Raynham, Tony; Ireson, Christopher; Sattler, Wolfgang

2013-01-01

Glioblastoma multiforme (GBM) is the most common malignant brain tumor, which, despite combined modality treatment, reoccurs and is invariably fatal for affected patients. Recently, a member of the serine/threonine protein kinase D (PRKD) family, PRKD2, was shown to be a potent mediator of glioblastoma growth. Here we studied the role of PRKD2 in U87MG glioblastoma cell migration and invasion in response to sphingosine-1-phosphate (S1P), an activator of PRKD2 and a GBM mitogen. Time-lapse microscopy demonstrated that random cell migration was significantly diminished in response to PRKD2 silencing. The pharmacological PRKD family inhibitor CRT0066101 decreased chemotactic migration and invasion across uncoated or matrigel-coated Transwell inserts. Silencing of PRKD2 attenuated migration and invasion of U87MG cells even more effectively. In terms of downstream signaling, CRT0066101 prevented PRKD2 autophosphorylation and inhibited p44/42 MAPK and to a smaller extent p54/46 JNK and p38 MAPK activation. PRKD2 silencing impaired activation of p44/42 MAPK and p54/46 JNK, downregulated nuclear c-Jun protein levels and decreased c-Jun S73 phosphorylation without affecting the NFκB pathway. Finally, qPCR array analyses revealed that silencing of PRKD2 downregulates mRNA levels of integrin alpha-2 and -4 (ITGA2 and -4), plasminogen activator urokinase (PLAU), plasminogen activator urokinase receptor (PLAUR), and matrix metallopeptidase 1 (MMP1). Findings of the present study identify PRKD2 as a potential target to interfere with glioblastoma cell migration and invasion, two major determinants contributing to recurrence of glioblastoma after multimodality treatment. Highlights: • Sphingosine-1-phosphate induces glioma cell migration and invasion. • Part of the effects is mediated by protein kinase D2 (PRKD2) activation. • Inactivation of PRKD2 attenuates glioblastoma cell migration and invasion. • Both, RNAi and pharmacological inhibition of PRKD2 inhibits MAPK

The p38 mitogen activated protein kinase regulates β-amyloid protein internalization through the α7 nicotinic acetylcholine receptor in mouse brain.

Science.gov (United States)

Ma, Kai-Ge; Lv, Jia; Yang, Wei-Na; Chang, Ke-Wei; Hu, Xiao-Dan; Shi, Li-Li; Zhai, Wan-Ying; Zong, Hang-Fan; Qian, Yi-Hua

2018-03-01

Alzheimer's disease (AD) is one of the most devastating neurodegenerative disorders. Intracellular β-amyloid protein (Aβ) is an early event in AD. It induces the formation of amyloid plaques and neuron damage. The α7 nicotinic acetylcholine receptor (α7nAChR) has been suggested to play an important role in Aβ caused cognition. It has high affinity with Aβ and could mediate Aβ internalization in vitro. However, whether in mouse brain the p38 MAPK signaling pathway is involved in the regulation of the α7nAChR mediated Aβ internalization and their role in mitochondria remains little known. Therefore, in this study, we revealed that Aβ is internalized by cholinergic and GABAergic neurons. The internalized Aβ were found deposits in lysosomes/endosomes and mitochondria. Aβ could form Aβ-α7nAChR complex with α7nAChR, activates the p38 mitogen activated protein kinase (MAPK). And the increasing of α7nAChR could in return mediate Aβ internalization in the cortex and hippocampus. In addition, by using the α7nAChR agonist PNU282987, the p38 phosphorylation level decreases, rescues the biochemical changes which are tightly associated with Aβ-induced apoptosis, such as Bcl2/Bax level, cytochrome c (Cyt c) release. Collectively, the p38 MAPK signaling pathway could regulate the α7nAChR-mediated internalization of Aβ. The activation of α7nAChR or the inhibition of p38 MAPK signaling pathway may be a beneficial therapy to AD. Copyright © 2017 Elsevier Inc. All rights reserved.

Low- and high-risk human papillomavirus E7 proteins regulate p130 differently

International Nuclear Information System (INIS)

Barrow-Laing, Lisa; Chen Wei; Roman, Ann

2010-01-01

The E7 protein of high-risk human papillomaviruses (HR HPVs) targets pRb family members (pRb, p107 and p130) for degradation; low-risk (LR) HPV E7 only targets p130 for degradation. The effect of HR HPV 16 E7 and LR HPV 6 E7 on p130 intracellular localization and half-life was examined. Nuclear/cytoplasmic fractionation and immunofluorescence showed that, in contrast to control and HPV 6 E7-expressing cells, a greater amount of p130 was present in the cytoplasm in the presence of HPV 16 E7. The half-life of p130, relative to control cells, was decreased in the cytoplasm in the presence of HPV 6 E7 or HPV 16 E7, but only decreased by HPV 6 E7 in the nucleus. Inhibition of proteasomal degradation extended the half-life of p130, regardless of intracellular localization. These results suggest that there may be divergent mechanisms by which LR and HR HPV E7 target p130 for degradation.

The human nucleolar protein FTSJ3 associates with NIP7 and functions in pre-rRNA processing.

Directory of Open Access Journals (Sweden)

Luis G Morello

Full Text Available NIP7 is one of the many trans-acting factors required for eukaryotic ribosome biogenesis, which interacts with nascent pre-ribosomal particles and dissociates as they complete maturation and are exported to the cytoplasm. By using conditional knockdown, we have shown previously that yeast Nip7p is required primarily for 60S subunit synthesis while human NIP7 is involved in the biogenesis of 40S subunit. This raised the possibility that human NIP7 interacts with a different set of proteins as compared to the yeast protein. By using the yeast two-hybrid system we identified FTSJ3, a putative ortholog of yeast Spb1p, as a human NIP7-interacting protein. A functional association between NIP7 and FTSJ3 is further supported by colocalization and coimmunoprecipitation analyses. Conditional knockdown revealed that depletion of FTSJ3 affects cell proliferation and causes pre-rRNA processing defects. The major pre-rRNA processing defect involves accumulation of the 34S pre-rRNA encompassing from site A' to site 2b. Accumulation of this pre-rRNA indicates that processing of sites A0, 1 and 2 are slower in cells depleted of FTSJ3 and implicates FTSJ3 in the pathway leading to 18S rRNA maturation as observed previously for NIP7. The results presented in this work indicate a close functional interaction between NIP7 and FTSJ3 during pre-rRNA processing and show that FTSJ3 participates in ribosome synthesis in human cells.

LSD1 demethylase and the methyl-binding protein PHF20L1 prevent SET7 methyltransferase-dependent proteolysis of the stem-cell protein SOX2.

Science.gov (United States)

Zhang, Chunxiao; Hoang, Nam; Leng, Feng; Saxena, Lovely; Lee, Logan; Alejo, Salvador; Qi, Dandan; Khal, Anthony; Sun, Hong; Lu, Fei; Zhang, Hui

2018-03-09

The pluripotency-controlling stem-cell protein SRY-box 2 (SOX2) plays a pivotal role in maintaining the self-renewal and pluripotency of embryonic stem cells and also of teratocarcinoma or embryonic carcinoma cells. SOX2 is monomethylated at lysine 119 (Lys-119) in mouse embryonic stem cells by the SET7 methyltransferase, and this methylation triggers ubiquitin-dependent SOX2 proteolysis. However, the molecular regulators and mechanisms controlling SET7-induced SOX2 proteolysis are unknown. Here, we report that in human ovarian teratocarcinoma PA-1 cells, methylation-dependent SOX2 proteolysis is dynamically regulated by the LSD1 lysine demethylase and a methyl-binding protein, PHD finger protein 20-like 1 (PHF20L1). We found that LSD1 not only removes the methyl group from monomethylated Lys-117 (equivalent to Lys-119 in mouse SOX2), but it also demethylates monomethylated Lys-42 in SOX2, a reaction that SET7 also regulated and that also triggered SOX2 proteolysis. Our studies further revealed that PHF20L1 binds both monomethylated Lys-42 and Lys-117 in SOX2 and thereby prevents SOX2 proteolysis. Down-regulation of either LSD1 or PHF20L1 promoted SOX2 proteolysis, which was prevented by SET7 inactivation in both PA-1 and mouse embryonic stem cells. Our studies also disclosed that LSD1 and PHF20L1 normally regulate the growth of pluripotent mouse embryonic stem cells and PA-1 cells by preventing methylation-dependent SOX2 proteolysis. In conclusion, our findings reveal an important mechanism by which the stability of the pluripotency-controlling stem-cell protein SOX2 is dynamically regulated by the activities of SET7, LSD1, and PHF20L1 in pluripotent stem cells. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Overexpressed Proteins in Hypervirulent Clade 8 and Clade 6 Strains of Escherichia coli O157:H7 Compared to E. coli O157:H7 EDL933 Clade 3 Strain.

Directory of Open Access Journals (Sweden)

Natalia Amigo

Full Text Available Escherichia coli O157:H7 is responsible for severe diarrhea and hemolytic uremic syndrome (HUS, and predominantly affects children under 5 years. The major virulence traits are Shiga toxins, necessary to develop HUS and the Type III Secretion System (T3SS through which bacteria translocate effector proteins directly into the host cell. By SNPs typing, E. coli O157:H7 was separated into nine different clades. Clade 8 and clade 6 strains were more frequently associated with severe disease and HUS. In this study, we aimed to identify differentially expressed proteins in two strains of E. coli O157:H7 (clade 8 and clade 6, obtained from cattle and compared them with the well characterized reference EDL933 strain (clade 3. Clade 8 and clade 6 strains show enhanced pathogenicity in a mouse model and virulence-related properties. Proteins were extracted and analyzed using the TMT-6plex labeling strategy associated with two dimensional liquid chromatography and mass spectrometry in tandem. We detected 2241 proteins in the cell extract and 1787 proteins in the culture supernatants. Attention was focused on the proteins related to virulence, overexpressed in clade 6 and 8 strains compared to EDL933 strain. The proteins relevant overexpressed in clade 8 strain were the curli protein CsgC, a transcriptional activator (PchE, phage proteins, Stx2, FlgM and FlgD, a dienelactone hydrolase, CheW and CheY, and the SPATE protease EspP. For clade 6 strain, a high overexpression of phage proteins was detected, mostly from Stx2 encoding phage, including Stx2, flagellin and the protease TagA, EDL933_p0016, dienelactone hydrolase, and Haemolysin A, amongst others with unknown function. Some of these proteins were analyzed by RT-qPCR to corroborate the proteomic data. Clade 6 and clade 8 strains showed enhanced transcription of 10 out of 12 genes compared to EDL933. These results may provide new insights in E. coli O157:H7 mechanisms of pathogenesis.

7 CFR 51.475 - U.S. Fancy.

Science.gov (United States)

2010-01-01

... meet the requirements of U.S. No. 1 grade except that the cantaloups have very good internal quality... shall not apply to the requirements relating to internal quality and uniformity of appearance: (1) At... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing...

Equilibrium optimization code OPEQ and results of applying it to HT-7U

International Nuclear Information System (INIS)

Zha Xuejun; Zhu Sizheng; Yu Qingquan

2003-01-01

The plasma equilibrium configuration has a strong impact on the confinement and MHD stability in tokamaks. For designing a tokamak device, it is an important issue to determine the sites and currents of poloidal coils which have some constraint conditions from physics and engineering with a prescribed equilibrium shape of the plasma. In this paper, an effective method based on multi-variables equilibrium optimization is given. The method can optimize poloidal coils when the previously prescribed plasma parameters are treated as an object function. We apply it to HT-7U equilibrium calculation, and obtain good results

LDL receptor-related protein 1 regulates the abundance of diverse cell-signaling proteins in the plasma membrane proteome.

Science.gov (United States)

Gaultier, Alban; Simon, Gabriel; Niessen, Sherry; Dix, Melissa; Takimoto, Shinako; Cravatt, Benjamin F; Gonias, Steven L

2010-12-03

LDL receptor-related protein 1 (LRP1) is an endocytic receptor, reported to regulate the abundance of other receptors in the plasma membrane, including uPAR and tissue factor. The goal of this study was to identify novel plasma membrane proteins, involved in cell-signaling, that are regulated by LRP1. Membrane protein ectodomains were prepared from RAW 264.7 cells in which LRP1 was silenced and control cells using protease K. Peptides were identified by LC-MS/MS. By analysis of spectral counts, 31 transmembrane and secreted proteins were regulated in abundance at least 2-fold when LRP1 was silenced. Validation studies confirmed that semaphorin4D (Sema4D), plexin domain-containing protein-1 (Plxdc1), and neuropilin-1 were more abundant in the membranes of LRP1 gene-silenced cells. Regulation of Plxdc1 by LRP1 was confirmed in CHO cells, as a second model system. Plxdc1 coimmunoprecipitated with LRP1 from extracts of RAW 264.7 cells and mouse liver. Although Sema4D did not coimmunoprecipitate with LRP1, the cell-surface level of Sema4D was increased by RAP, which binds to LRP1 and inhibits binding of other ligands. These studies identify Plxdc1, Sema4D, and neuropilin-1 as novel LRP1-regulated cell-signaling proteins. Overall, LRP1 emerges as a generalized regulator of the plasma membrane proteome.

Bromodomain-containing Protein 4 Activates Voltage-gated Sodium Channel 1.7 Transcription in Dorsal Root Ganglia Neurons to Mediate Thermal Hyperalgesia in Rats.

Science.gov (United States)

Hsieh, Ming-Chun; Ho, Yu-Cheng; Lai, Cheng-Yuan; Wang, Hsueh-Hsiao; Lee, An-Sheng; Cheng, Jen-Kun; Chau, Yat-Pang; Peng, Hsien-Yu

2017-11-01

Bromodomain-containing protein 4 binds acetylated promoter histones and promotes transcription; however, the role of bromodomain-containing protein 4 in inflammatory hyperalgesia remains unclear. Male Sprague-Dawley rats received hind paw injections of complete Freund's adjuvant to induce hyperalgesia. The dorsal root ganglia were examined to detect changes in bromodomain-containing protein 4 expression and the activation of genes involved in the expression of voltage-gated sodium channel 1.7, which is a key pain-related ion channel. The intraplantar complete Freund's adjuvant injections resulted in thermal hyperalgesia (4.0 ± 1.5 s; n = 7). The immunohistochemistry and immunoblotting results demonstrated an increase in the bromodomain-containing protein 4-expressing dorsal root ganglia neurons (3.78 ± 0.38 fold; n = 7) and bromodomain-containing protein 4 protein levels (2.62 ± 0.39 fold; n = 6). After the complete Freund's adjuvant injection, histone H3 protein acetylation was enhanced in the voltage-gated sodium channel 1.7 promoter, and cyclin-dependent kinase 9 and phosphorylation of RNA polymerase II were recruited to this area. Furthermore, the voltage-gated sodium channel 1.7-mediated currents were enhanced in neurons of the complete Freund's adjuvant rats (55 ± 11 vs. 19 ± 9 pA/pF; n = 4 to 6 neurons). Using bromodomain-containing protein 4-targeted antisense small interfering RNA to the complete Freund's adjuvant-treated rats, the authors demonstrated a reduction in the expression of bromodomain-containing protein 4 (0.68 ± 0.16 fold; n = 7), a reduction in thermal hyperalgesia (7.5 ± 1.5 s; n = 7), and a reduction in the increased voltage-gated sodium channel 1.7 currents (21 ± 4 pA/pF; n = 4 to 6 neurons). Complete Freund's adjuvant triggers enhanced bromodomain-containing protein 4 expression, ultimately leading to the enhanced excitability of nociceptive neurons and thermal hyperalgesia. This effect is

Cellulolytic (cel) genes of Clostridium thermocellum F7 and the proteins encoded by them

International Nuclear Information System (INIS)

Piruzyan, E.S.; Mogutov, M.A.; Velikodvorskaya, G.A.; Pushkarskaya, T.A.

1988-01-01

This study is concerned with genes cell, ce12, and ce13 encoding the endoglucanase of the cellulolytic complex of the anaerobic thermophilic Clostridium thermocellum F7 bacteria, these genes having been closed by us earlier. The authors present the characteristics of proteins synthesized by the cel genes in the minicell system of the strain Escherichia coli K-12 X925. The molecular weights of the proteins encoded by genes cell, ce12, and ce13 are 30,000, 45,000, and 50,000 dalton, respectively. The study of the homology of the cloned section of the C. thermocellum DNA containing the endoglucanase genes, using Southern's blot-hybridization method, did not reveal their physical linkage in the genome. The authors detected a plasmid with a size of about 30 kb in the cells of the C. thermocellum F7 strain investigated

Muscle function recovery in golden retriever muscular dystrophy after AAV1-U7 exon skipping.

Science.gov (United States)

Vulin, Adeline; Barthélémy, Inès; Goyenvalle, Aurélie; Thibaud, Jean-Laurent; Beley, Cyriaque; Griffith, Graziella; Benchaouir, Rachid; le Hir, Maëva; Unterfinger, Yves; Lorain, Stéphanie; Dreyfus, Patrick; Voit, Thomas; Carlier, Pierre; Blot, Stéphane; Garcia, Luis

2012-11-01

Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder resulting from lesions of the gene encoding dystrophin. These usually consist of large genomic deletions, the extents of which are not correlated with the severity of the phenotype. Out-of-frame deletions give rise to dystrophin deficiency and severe DMD phenotypes, while internal deletions that produce in-frame mRNAs encoding truncated proteins can lead to a milder myopathy known as Becker muscular dystrophy (BMD). Widespread restoration of dystrophin expression via adeno-associated virus (AAV)-mediated exon skipping has been successfully demonstrated in the mdx mouse model and in cardiac muscle after percutaneous transendocardial delivery in the golden retriever muscular dystrophy dog (GRMD) model. Here, a set of optimized U7snRNAs carrying antisense sequences designed to rescue dystrophin were delivered into GRMD skeletal muscles by AAV1 gene transfer using intramuscular injection or forelimb perfusion. We show sustained correction of the dystrophic phenotype in extended muscle areas and partial recovery of muscle strength. Muscle architecture was improved and fibers displayed the hallmarks of mature and functional units. A 5-year follow-up ruled out immune rejection drawbacks but showed a progressive decline in the number of corrected muscle fibers, likely due to the persistence of a mild dystrophic process such as occurs in BMD phenotypes. Although AAV-mediated exon skipping was shown safe and efficient to rescue a truncated dystrophin, it appears that recurrent treatments would be required to maintain therapeutic benefit ahead of the progression of the disease.

On the efficient bio-incorporation of 5-hydroxy-tryptophan in recombinant proteins expressed in Escherichia coli with T7 RNA polymerase-based vectors.

Science.gov (United States)

Oliveira-Souza, Wellington P; Bronze, Fellipe; Broos, Jaap; Marcondes, Marcelo F M; Oliveira, Vitor

2017-10-21

Biosynthetic incorporation of non-canonic amino acids is an attractive strategy to introduce new properties in recombinant proteins. Trp analogs can be incorporated in recombinant proteins replacing regular Trp during protein translation into a Trp-auxotrophic cell host. This straightforward method however, is limited to few analogs recognized and accepted by the cellular protein production machinery. 5-hydroxy-tryptophan (5OH-Trp) can be bio-incorporated using E. coli as expression host however; we have experienced very low incorporation yields - amount of protein containing regular Trp/amount of protein containing the Trp analog - during expressions of 5OH-Trp labeled proteins. Furthermore, this low incorporation yield were verified especially when the widely-used vectors based on the T7 RNA polymerase were used. Testing different 5OH-Trp incorporation protocols we verified that in these T7-based systems, the production of the T7 RNA polymerase is driven by the same elements - lac promoter/IPTG - as the target protein. Consequently, the bio-incorporation of the 5OH-Trp residues also occurs in this crucial enzyme, but, the produced T7 RNA polymerase labeled with 5OH-Trp is inactive or much less active. In the present work, we describe an efficient method to overcome this mentioned problem and bio-incorporate 5OH-Trp in proteins expressed in E. coli., using vectors based on the T7 RNA polymerase-T7 promoter. The two-step induction protocol here described showed incorporation efficiencies of 5OH-Trp higher than 90%. Copyright © 2017 Elsevier Inc. All rights reserved.

Systematic identification of anti-interferon function on hepatitis C virus genome reveals p7 as an immune evasion protein.

Science.gov (United States)

Qi, Hangfei; Chu, Virginia; Wu, Nicholas C; Chen, Zugen; Truong, Shawna; Brar, Gurpreet; Su, Sheng-Yao; Du, Yushen; Arumugaswami, Vaithilingaraja; Olson, C Anders; Chen, Shu-Hua; Lin, Chung-Yen; Wu, Ting-Ting; Sun, Ren

2017-02-21

Hepatitis C virus (HCV) encodes mechanisms to evade the multilayered antiviral actions of the host immune system. Great progress has been made in elucidating the strategies HCV employs to down-regulate interferon (IFN) production, impede IFN signaling transduction, and impair IFN-stimulated gene (ISG) expression. However, there is a limited understanding of the mechanisms governing how viral proteins counteract the antiviral functions of downstream IFN effectors due to the lack of an efficient approach to identify such interactions systematically. To study the mechanisms by which HCV antagonizes the IFN responses, we have developed a high-throughput profiling platform that enables mapping of HCV sequences critical for anti-IFN function at high resolution. Genome-wide profiling performed with a 15-nt insertion mutant library of HCV showed that mutations in the p7 region conferred high levels of IFN sensitivity, which could be alleviated by the expression of WT p7 protein. This finding suggests that p7 protein of HCV has an immune evasion function. By screening a liver-specific ISG library, we identified that IFI6-16 significantly inhibits the replication of p7 mutant viruses without affecting WT virus replication. In contrast, knockout of IFI6-16 reversed the IFN hypersensitivity of p7 mutant virus. In addition, p7 was found to be coimmunoprecipitated with IFI6-16 and to counteract the function of IFI6-16 by depolarizing the mitochondria potential. Our data suggest that p7 is a critical immune evasion protein that suppresses the antiviral IFN function by counteracting the function of IFI6-16.

Obtaining of U-2.5Zr7.5Nb and U-3Zr-9Nb alloys by sintering process

International Nuclear Information System (INIS)

Mazzeu, Thiago de Oliveira; Paula, Joao Bosco de; Ferraz, Wilmar Barbosa; Santos, Ana Maria Matildes dos; Brina, Jose Giovanni Mascarenhas

2011-01-01

The development of metallic fuels with low enrichment to be used in research and test reactors, as well in the future pressurized water reactors, focuses on the search for uranium alloys of high density. 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) in Belo Horizonte 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 400MPa and then sintered under a vacuum of about 5 x 10-6 Torr at temperatures ranging from 1050 deg to 1300 deg C. The densities of the alloys were measured geometrically and by hydrostatic method using water. The microstructures of the pellets were observed by scanning electron microscopy (SEM) and the elements of alloying were identified by energy dispersive X-ray spectroscopy (SEM/EDS) analysis. The obtained results showed a small increasing density with rising sintering temperature. The highest density achieved was approximately 80% of theoretical density. It was also qualitatively observed that the superficial oxidation of the pellets increased with increasing sintering temperature thus avoiding the fusion of the alloys at higher temperatures. (author)

SEM Characterization of the High Burn-up Microstructure of U-7Mo Alloy

Energy Technology Data Exchange (ETDEWEB)

Dennis D. Keiser, Jr.; Jan-Fong Jue; Jian Gan; Brandon Miller; Adam Robinson; Pavel Medvedev; James Madden; Dan Wachs; M. Teague

2014-04-01

During irradiation, the microstructure of U-7Mo evolves until at a fission density near 5x1021 f/cm3 a high-burnup microstructure exists that is very different than what was observed at lower fission densities. This microstructure is dominated by randomly distributed, relatively large, homogeneous fission gas bubbles. The bubble superlattice has collapsed in many microstructural regions, and the fuel grain sizes, in many areas, become sub-micron in diameter with both amorphous fuel and crystalline fuel present. Solid fission product precipitates can be found inside the fission gas bubbles. To generate more information about the characteristics of the high-fission density microstructure, three samples irradiated in the RERTR-7 experiment have been characterized using a scanning electron microscope equipped with a focused ion beam. The FIB was used to generate samples for SEM imaging and to perform 3D reconstruction of the microstructure, which can be used to look for evidence of possible fission gas bubble interlinkage.

Structural basis for the binding of the neutralizing antibody, 7D11, to the poxvirus L1 protein

International Nuclear Information System (INIS)

Su, Hua-Poo; Golden, Joseph W.; Gittis, Apostolos G.; Hooper, Jay W.; Garboczi, David N.

2007-01-01

Medical countermeasures to prevent or treat smallpox are needed due to the potential use of poxviruses as biological weapons. Safety concerns with the currently available smallpox vaccine indicate a need for research on alternative poxvirus vaccine strategies. Molecular vaccines involving the use of proteins and/or genes and recombinant antibodies are among the strategies under current investigation. The poxvirus L1 protein, encoded by the L1R open reading frame, is the target of neutralizing antibodies and has been successfully used as a component of both protein subunit and DNA vaccines. L1-specific monoclonal antibodies (e.g., mouse monoclonal antibody mAb-7D11, mAb-10F5) with potent neutralizing activity bind L1 in a conformation-specific manner. This suggests that proper folding of the L1 protein used in molecular vaccines will affect the production of neutralizing antibodies and protection. Here, we co-crystallized the Fab fragment of mAb-7D11 with the L1 protein. The crystal structure of the complex between Fab-7D11 and L1 reveals the basis for the conformation-specific binding as recognition of a discontinuous epitope containing two loops that are held together by a disulfide bond. The structure of this important conformational epitope of L1 will contribute to the development of molecular poxvirus vaccines and also provides a novel target for anti-poxvirus drugs. In addition, the sequence and structure of Fab-7D11 will contribute to the development of L1-targeted immunotherapeutics

The Effect of a High-Protein Diet and Exercise on Cardiac AQP7 and GLUT4 Gene Expression.

Science.gov (United States)

Palabiyik, Orkide; Karaca, Aziz; Taştekin, Ebru; Yamasan, Bilge Eren; Tokuç, Burcu; Sipahi, Tammam; Vardar, Selma Arzu

2016-10-01

High-protein (HP) diets are commonly consumed by athletes despite their potential health hazard, which is postulated to enforce a negative effect on bone and renal health. However, its effects on heart have not been known yet. Aquaporin-7 (AQP7) is an aquaglyceroporin that facilitates glycerol and water transport. Glycerol is an important cardiac energy production substrate, especially during exercise, in conjunction with fatty acids and glucose. Glucose transporter 4 (GLUT4) is an insulin-sensitive glucose transporter in heart. We aimed to investigate the effect of HPD on AQP7 and GLUT4 levels in the rat heart subjected to exercise. Male Sprague-Dawley rats were divided into control (n = 12), exercise (E) training (n = 10), HPD (n = 12), and HPD-E training (n = 9) groups. The HPD groups were fed a 45 % protein-containing diet 5 weeks. The HPD-E and E groups were performed the treadmill exercise during the 5-week study period. Real-time polymerase chain reaction and immunohistochemistry techniques were used to determine the gene expression and localization of AQP7 and GLUT4 in heart tissue. Results of relative gene expression were calculated by the 'Pfaffl' mathematical method using the REST program. Differences in AQP7 and GLUT4 gene expression were expressed as fold change compared to the control group. Heart weight/tibia ratio and ventricular wall thickness were evaluated as markers of cardiac hypertrophy. Further, serum glucose, glycerol, and insulin levels were also measured. AQP7 gene expression was found to be increased in the E (3.47-fold, p protein expression was also increased in the HPD and HPD-E groups (p protein expression was significantly increased in the E, HPD, and HPD-E groups compared to the control group (p = 0.024, p protein diet groups (C and E). Serum insulin levels were higher for HPD groups compared with the normal-protein diet groups (p < 0.001), whereas no differences were observed between the exercise and sedentary

Ck2-Dependent Phosphorylation Is Required to Maintain Pax7 Protein Levels in Proliferating Muscle Progenitors.

Directory of Open Access Journals (Sweden)

Natalia González

Full Text Available Skeletal muscle regeneration and long term maintenance is directly link to the balance between self-renewal and differentiation of resident adult stem cells known as satellite cells. In turn, satellite cell fate is influenced by a functional interaction between the transcription factor Pax7 and members of the MyoD family of muscle regulatory factors. Thus, changes in the Pax7-to-MyoD protein ratio may act as a molecular rheostat fine-tuning acquisition of lineage identity while preventing precocious terminal differentiation. Pax7 is expressed in quiescent and proliferating satellite cells, while its levels decrease sharply in differentiating progenitors Pax7 is maintained in cells (reacquiring quiescence. While the mechanisms regulating Pax7 levels based on differentiation status are not well understood, we have recently described that Pax7 levels are directly regulated by the ubiquitin-ligase Nedd4, thus promoting proteasome-dependent Pax7 degradation in differentiating satellite cells. Here we show that Pax7 levels are maintained in proliferating muscle progenitors by a mechanism involving casein kinase 2-dependent Pax7 phosphorylation at S201. Point mutations preventing S201 phosphorylation or casein kinase 2 inhibition result in decreased Pax7 protein in proliferating muscle progenitors. Accordingly, this correlates directly with increased Pax7 ubiquitination. Finally, Pax7 down regulation induced by casein kinase 2 inhibition results in precocious myogenic induction, indicating early commitment to terminal differentiation. These observations highlight the critical role of post translational regulation of Pax7 as a molecular switch controlling muscle progenitor fate.

Molecular Cloning and Function of FAS/APO1 Associated Protein in Breast Cancer.

Science.gov (United States)

1996-06-01

Ariyama T, Abe T, Druck T, Ohta M, Huebner K, Yanagisawa J, Reed JC, Sato T: PTPN13, a Fas-associated protein tyrosine phosphatase, is located on...20. Yang, Q., and Tonks, N. K. (1991). Isolation of a cDNA clone encoding a human protein-tyrosine phosphatase with homology 7. Huebner, K., Druck , T...Acad. Sci. U.S.A. 91, 7477 (1994). Res. 53, 1945 (1993).(Fig. 3D ). In contrast to Jurkat cells which 13. The original description of PTP-BAS (12

Effects of irradiation on the microstructure of U-7Mo dispersion fuel with Al-2Si matrix

Science.gov (United States)

Keiser, Dennis D.; Jue, Jan-Fong; Robinson, Adam B.; Medvedev, Pavel; Gan, Jian; Miller, Brandon D.; Wachs, Daniel M.; Moore, Glenn A.; Clark, Curtis R.; Meyer, Mitchell K.; Ross Finlay, M.

2012-06-01

The Reduced Enrichment for Research and Test Reactor (RERTR) program is developing low-enriched uranium U-Mo dispersion fuels for application in research and test reactors around the world. As part of this development, fuel plates have been irradiated in the Advanced Test Reactor and then characterized using optical metallography (OM) and scanning electron microscopy (SEM) to determine the as-irradiated microstructure. To demonstrate the irradiation performance of U-7Mo dispersion fuel plates with 2 wt.% Si added to the matrix, fuel plates were tested to moderate burnups at intermediate fission rates as part of the RERTR-6 experiment. Further testing was performed to higher fission rates as part of the RERTR-7A experiment, and very aggressive testing (high temperature, high fission density, and high fission rate) was performed in the RERTR-9A, RERTR-9B, and AFIP-1 experiments. As-irradiated microstructures were compared to those observed after fabrication to determine the effects of irradiation on the microstructure. Based on comparison of the microstructural characterization results for each irradiated sample, some general conclusions can be drawn about how the microstructure evolves during irradiation: there is growth during irradiation of the fuel/matrix interaction (FMI) layer created during fabrication; Si diffuses from the FMI layer to deeper depths in the U-7Mo particles as the irradiation conditions are made more aggressive; lowering of the Si content in the FMI layer results in an increase in the size of the fission gas bubbles; as the FMI layer grows during irradiation, more Si diffuses from the matrix to the FMI layer/matrix interface; and interlinking of fission gas bubbles in the fuel plate microstructure that may indicate breakaway swelling is not observed.

Effects of irradiation on the microstructure of U-7Mo dispersion fuel with Al-2Si matrix

Energy Technology Data Exchange (ETDEWEB)

Keiser, Dennis D., E-mail: Dennis.Keiser@inl.gov [Nuclear Fuels and Materials Division, Idaho National Laboratory, P.O. Box 1625, Idaho Falls, ID 83415-6188 (United States); Jue, Jan-Fong; Robinson, Adam B.; Medvedev, Pavel; Gan, Jian; Miller, Brandon D.; Wachs, Daniel M.; Moore, Glenn A.; Clark, Curtis R.; Meyer, Mitchell K. [Nuclear Fuels and Materials Division, Idaho National Laboratory, P.O. Box 1625, Idaho Falls, ID 83415-6188 (United States); Ross Finlay, M. [Australian Nuclear Science and Technology Organization, PMB 1, Menai, NSW 2234 (Australia)

2012-06-15

The Reduced Enrichment for Research and Test Reactor (RERTR) program is developing low-enriched uranium U-Mo dispersion fuels for application in research and test reactors around the world. As part of this development, fuel plates have been irradiated in the Advanced Test Reactor and then characterized using optical metallography (OM) and scanning electron microscopy (SEM) to determine the as-irradiated microstructure. To demonstrate the irradiation performance of U-7Mo dispersion fuel plates with 2 wt.% Si added to the matrix, fuel plates were tested to moderate burnups at intermediate fission rates as part of the RERTR-6 experiment. Further testing was performed to higher fission rates as part of the RERTR-7A experiment, and very aggressive testing (high temperature, high fission density, and high fission rate) was performed in the RERTR-9A, RERTR-9B, and AFIP-1 experiments. As-irradiated microstructures were compared to those observed after fabrication to determine the effects of irradiation on the microstructure. Based on comparison of the microstructural characterization results for each irradiated sample, some general conclusions can be drawn about how the microstructure evolves during irradiation: there is growth during irradiation of the fuel/matrix interaction (FMI) layer created during fabrication; Si diffuses from the FMI layer to deeper depths in the U-7Mo particles as the irradiation conditions are made more aggressive; lowering of the Si content in the FMI layer results in an increase in the size of the fission gas bubbles; as the FMI layer grows during irradiation, more Si diffuses from the matrix to the FMI layer/matrix interface; and interlinking of fission gas bubbles in the fuel plate microstructure that may indicate breakaway swelling is not observed.

Ubiquitinated proteins enriched from tumor cells by a ubiquitin binding protein Vx3(A7) as a potent cancer vaccine.

Science.gov (United States)

Aldarouish, Mohanad; Wang, Huzhan; Zhou, Meng; Hu, Hong-Ming; Wang, Li-Xin

2015-04-16

Our previous studies have demonstrated that autophagosome-enriched vaccine (named DRibbles: DRiPs-containing blebs) induce a potent anti-tumor efficacy in different murine tumor models, in which DRibble-containing ubiquitinated proteins are efficient tumor-specific antigen source for the cross-presentation after being loaded onto dendritic cells. In this study, we sought to detect whether ubiquitinated proteins enriched from tumor cells could be used directly as a novel cancer vaccine. The ubiquitin binding protein Vx3(A7) was used to isolate ubiquitinated proteins from EL4 and B16-F10 tumor cells after blocking their proteasomal degradation pathway. C57BL/6 mice were vaccinated with different doses of Ub-enriched proteins via inguinal lymph nodes or subcutaneous injection and with DRibbles, Ub-depleted proteins and whole cell lysate as comparison groups, respectively. The lymphocytes from the vaccinated mice were re-stimulated with inactivated tumor cells and the levels of IFN-γ in the supernatant were detected by ELISA. Anti-tumor efficacy of Ub-enriched proteins vaccine was evaluated by monitoring tumor growth in established tumor mice models. Graphpad Prism 5.0 was used for all statistical analysis. We found that after stimulation with inactivated tumor cells, the lymphocytes from the Ub-enriched proteins-vaccinated mice secreted high level of IFN-γ in dose dependent manner, in which the priming vaccination via inguinal lymph nodes injection induced higher IFN-γ level than that via subcutaneous injection. Moreover, the level of secreted IFN-γ in the Ub-enriched proteins group was markedly higher than that in the whole cell lysate and Ub-depleted proteins. Interestingly, the lymphocytes from mice vaccinated with Ub-enriched proteins, but not Ub-depleted proteins and whole cell lysates, isolated from EL4 or B16-F10 tumor cells also produced an obvious level of IFN-γ when stimulated alternately with inactivated B16-F10 or EL4 tumor cells. Furthermore, Ub

Analysis of the chloroplast protein kinase Stt7 during state transitions.

Directory of Open Access Journals (Sweden)

Sylvain Lemeille

2009-03-01

Full Text Available State transitions allow for the balancing of the light excitation energy between photosystem I and photosystem II and for optimal photosynthetic activity when photosynthetic organisms are subjected to changing light conditions. This process is regulated by the redox state of the plastoquinone pool through the Stt7/STN7 protein kinase required for phosphorylation of the light-harvesting complex LHCII and for the reversible displacement of the mobile LHCII between the photosystems. We show that Stt7 is associated with photosynthetic complexes including LHCII, photosystem I, and the cytochrome b6f complex. Our data reveal that Stt7 acts in catalytic amounts. We also provide evidence that Stt7 contains a transmembrane region that separates its catalytic kinase domain on the stromal side from its N-terminal end in the thylakoid lumen with two conserved Cys that are critical for its activity and state transitions. On the basis of these data, we propose that the activity of Stt7 is regulated through its transmembrane domain and that a disulfide bond between the two lumen Cys is essential for its activity. The high-light-induced reduction of this bond may occur through a transthylakoid thiol-reducing pathway driven by the ferredoxin-thioredoxin system which is also required for cytochrome b6f assembly and heme biogenesis.

Characterization of the interaction layer in diffusion couples U-Mo-Zr/Al and U-Mo-Zr/Al-A356 at 550 C degrees; Caracterizacion de la zona de interaccion en pares de difusion a 550 grados C U-Mo-Zr/Al y U-Mo-Zr/Al-A356

Energy Technology Data Exchange (ETDEWEB)

Komar Varela, Carolina; Arico, Sergio; Mirandou, Marcela; Balart, Silvia; Gribaudo, Luis [Comision Nacional de Energia Atomica, General San Martin (Argentina). Dept. de Materiales; com, carolinakomar@gmail

2007-07-01

Out-of-pile diffusion experiments were performed between U-7 wt.% Mo-1 wt.% Zr and Al or Al A356 (7,1 wt.% Si) at 550 C degrees. In this work morphological characterization and phase identification on both interaction layers are presented. They were carried out by the use of different techniques: optical and scanning electron microscopy, X-ray diffraction and WDS microanalysis. In the interaction layer U-7 wt.% Mo-1 wt.% Zr/Al, the phases UAl{sub 3}, UAl{sub 4}, Al{sub 20}Mo{sub 2}U and Al{sub 43}Mo{sub 4}U{sub 6} were identified. Similar results in the interaction layer of the U-7 % Mo/Al at 580 C degrees were previously obtained. In the interaction layer U-7 wt.% Mo-1 wt.% Zr/Al A356, the phases U(Al,Si){sub 3} with 25 at.% Si and Si{sub 5}U{sub 3} were identified. This last phase, with a higher Si concentration, was identified with X-ray diffraction synchrotron radiation performed at the National Synchrotron Light Laboratory, Campinas, Brazil. (author) [Spanish] Se realizaron experiencias fuera de reactor en pares de difusion quimica U-7 % Mo-1 % Zr/Al y U-7 % Mo-1 % Zr/Al A356. En este trabajo se presentan los resultados de la caracterizacion morfologica e identificacion de fases presentes en la zona de interaccion que se forma al ser sometidos a un tratamiento isotermico de 1,5 h a 550 grados C. Las tecnicas utilizadas fueron: microscopia optica y electronica de barrido, difraccion de rayos X y microanalisis cuantitativo por sonda electronica. En la zona de interaccion correspondiente al par U-7 % Mo-1 % Zr/Al se identificaron las fases UAl{sub 3}, UAl{sub 4}, Al{sub 20}Mo{sub 2}U y Al{sub 43}Mo{sub 4}U{sub 6}. Estas cuatro fases fueron identificadas en pares U-7 % Mo/Al a 580 grados C en trabajos anteriores. En la zona de interaccion correspondiente al par U-7 % Mo-1 % Zr/Al A356 se identificaron las fases U(Al,Si){sub 3} (con una concentracion de 25 %at.Si) y Si{sub 5}U{sub 3}. Este compuesto rico en Si solo pudo ser identificado mediante la utilizacion de

Analysis list: CBX7 [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available CBX7 Breast,Epidermis + hg19 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/target.../CBX7.1.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/target/CBX7.5.tsv http://dbarchive.biosciencedbc....jp/kyushu-u/hg19/target/CBX7.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/colo/CBX7.Breast.tsv,http://dbarchive.bioscience...dbc.jp/kyushu-u/hg19/colo/CBX7.Epidermis.tsv http://dbarchive.bioscience...dbc.jp/kyushu-u/hg19/colo/Breast.gml,http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/colo/Epidermis.gml ...

A docking study of enhanced intracellular survival protein from Mycobacterium tuberculosis with human DUSP16/MKP-7

International Nuclear Information System (INIS)

Yoon, Hye-Jin; Kim, Kyoung Hoon; Yang, Jin Kuk; Suh, Se Won; Kim, Hyunsik; Jang, Soonmin

2013-01-01

A docking study of Mtb Eis with its substrate DUSP16/MKP-7 was performed. The docking model suggests dissociation of hexameric Mtb Eis into dimers or monomers. The intracellular pathogen Mycobacterium tuberculosis (Mtb) causes tuberculosis, and one of its secreted effector proteins, called enhanced intracellular survival (Eis) protein, enhances its survival in macrophages. Mtb Eis activates JNK-specific dual-specificity protein phosphatase 16 (DUSP16)/mitogen-activated protein kinase phosphatase-7 (MKP-7) through the acetylation on Lys55, thus inactivating JNK by dephosphorylation. Based on the recently reported crystal structure of Mtb Eis, a docking model for the binding of Mtb Eis to DUSP16/MKP-7 was generated. In the docking model, the substrate helix containing Lys55 of DUSP16/MKP-7 fits nicely into the active-site cleft of Mtb Eis; the twisted β-sheet of Eis domain II embraces the substrate helix from one side. Most importantly, the side-chain of Lys55 is inserted toward acetyl-CoA and the resulting distance is 4.6 Å between the NZ atom of Lys55 and the carbonyl carbon of the acetyl group in acetyl-CoA. The binding of Mtb Eis and DUSP16/MKP-7 is maintained by strong electrostatic interactions. The active-site cleft of Mtb Eis has a negatively charged surface formed by Asp25, Glu138, Asp286, Glu395 and the terminal carboxylic group of Phe396. In contrast, DUSP16/MKP-7 contains five basic residues, Lys52, Lys55, Arg56, Arg57 and Lys62, which point toward the negatively charged surface of the active-site pocket of Mtb Eis. Thus, the current docking model suggests that the binding of DUSP16/MKP-7 to Mtb Eis should be established by charge complementarity in addition to a very favorable geometric arrangement. The suggested mode of binding requires the dissociation of the hexameric Mtb Eis into dimers or monomers. This study may be useful for future studies aiming to develop inhibitors of Mtb Eis as a new anti-tuberculosis drug candidate

A docking study of enhanced intracellular survival protein from Mycobacterium tuberculosis with human DUSP16/MKP-7

Energy Technology Data Exchange (ETDEWEB)

Yoon, Hye-Jin, E-mail: yoonhj@snu.ac.kr; Kim, Kyoung Hoon [Seoul National University, Seoul 151-747 (Korea, Republic of); Yang, Jin Kuk [Soongsil University, Seoul 156-743 (Korea, Republic of); Suh, Se Won [Seoul National University, Seoul 151-747 (Korea, Republic of); Seoul National University, Seoul 151-747 (Korea, Republic of); Kim, Hyunsik; Jang, Soonmin, E-mail: yoonhj@snu.ac.kr [Sejong University, Seoul 143-747 (Korea, Republic of)

2013-11-01

A docking study of Mtb Eis with its substrate DUSP16/MKP-7 was performed. The docking model suggests dissociation of hexameric Mtb Eis into dimers or monomers. The intracellular pathogen Mycobacterium tuberculosis (Mtb) causes tuberculosis, and one of its secreted effector proteins, called enhanced intracellular survival (Eis) protein, enhances its survival in macrophages. Mtb Eis activates JNK-specific dual-specificity protein phosphatase 16 (DUSP16)/mitogen-activated protein kinase phosphatase-7 (MKP-7) through the acetylation on Lys55, thus inactivating JNK by dephosphorylation. Based on the recently reported crystal structure of Mtb Eis, a docking model for the binding of Mtb Eis to DUSP16/MKP-7 was generated. In the docking model, the substrate helix containing Lys55 of DUSP16/MKP-7 fits nicely into the active-site cleft of Mtb Eis; the twisted β-sheet of Eis domain II embraces the substrate helix from one side. Most importantly, the side-chain of Lys55 is inserted toward acetyl-CoA and the resulting distance is 4.6 Å between the NZ atom of Lys55 and the carbonyl carbon of the acetyl group in acetyl-CoA. The binding of Mtb Eis and DUSP16/MKP-7 is maintained by strong electrostatic interactions. The active-site cleft of Mtb Eis has a negatively charged surface formed by Asp25, Glu138, Asp286, Glu395 and the terminal carboxylic group of Phe396. In contrast, DUSP16/MKP-7 contains five basic residues, Lys52, Lys55, Arg56, Arg57 and Lys62, which point toward the negatively charged surface of the active-site pocket of Mtb Eis. Thus, the current docking model suggests that the binding of DUSP16/MKP-7 to Mtb Eis should be established by charge complementarity in addition to a very favorable geometric arrangement. The suggested mode of binding requires the dissociation of the hexameric Mtb Eis into dimers or monomers. This study may be useful for future studies aiming to develop inhibitors of Mtb Eis as a new anti-tuberculosis drug candidate.

A 7-mer knowledge-based potential for detecting native protein structures from decoys

DEFF Research Database (Denmark)

Røgen, Peter

for faster sampling methods. Background: The C-alpha atoms define a polygonal curve in 3-space which is smoothened by the method presented in [1] and is illustrated below. The geometry of a 7-mer is described by two numbers that describe how stretched and curved the smoothening of the 7-mer is. These two...... numbers are called length and distance excess, c.f. [2], and give one point in the length - distance excess - plane, LDE-plane. Method: Given a sequence of amino acids, we break it down to all its 7-mers and search a database of known 3d-structures for similar 7-mer sequences. For the query 7-mer we...... define an energy function in the LDE-plane. This energy is given by the 7-mer found and depends linearly on some design parameters. The energy function of the full query sequence, F, is then a sum over all 7-mers. For a protein P and a decoy D we ideally want F(D)-F(P)=constant.RMSD( D , P ), where 0...

Metaproteomics Identifies the Protein Machinery Involved in Metal and Radionuclide Reduction in Subsurface Microbiomes and Elucidates Mechanisms and U(VI) Reduction Immobilization

Energy Technology Data Exchange (ETDEWEB)

Pfiffner, Susan M. [Univ. of Tennessee, Knoxville, TN (United States); Löffler, Frank [Univ. of Tennessee, Knoxville, TN (United States); Ritalahti, Kirsti [Univ. of Tennessee, Knoxville, TN (United States); Sayler, Gary [Univ. of Tennessee, Knoxville, TN (United States); Layton, Alice [Univ. of Tennessee, Knoxville, TN (United States); Hettich, Robert [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)

2015-08-31

radionuclide reduction activity, are suitable biomarkers, (2) identify new biomarker targets for site assessment and bioremediation monitoring, and (3) provide new information about specific proteins and mechanisms involved in U(VI) reduction and immobilization. This expanded metagenomic and metaproteomic toolbox contributed to implementing science-driven site management with broad benefits to the DOE mission.

A Chimeric Protein PTEN-L-p53 Enters U251 Cells to Repress Proliferation and Invasion.

Science.gov (United States)

Xiao, Man; An, Yang; Wang, Fengling; Yao, Chao; Zhang, Chu; Xin, Junfang; Duan, Yongjian; Zhao, Xiaofang; Fang, Na; Ji, Shaoping

2018-05-23

PTEN, a well-known tumor suppressor, dephosphorylates PIP3 and inhibits AKT activity. A translational variant of PTEN has been identified and termed PTEN-Long (PTEN-L). The additional 173 amino acids (PTEN-L leader) at the N-terminal constitute a potential signal peptide. Differing from canonical PTEN, PTEN-L is secreted into the extracellular fluid and re-enters recipient cells, playing the similar roles as PTEN in vivo and in vitro. This character confers the PTEN-L a therapeutic ability via directly protein delivering instead of traditional DNA and RNA vector options. In the present study, we employed PTEN-L leader to assemble a fusion protein, PTEN-L-p53, inosculated with the transcriptional regulator TP53, which is another powerful tumor suppressor. We overexpressed PTEN-L-p53 in HEK293T cells and detected it in both the cytoplasm and nucleus. Subsequently, we found that PTEN-L-p53 was secreted outside of the cells and detected in the culture media by immunoblotting. Furthermore, we demonstrated that PTEN-L-p53 freely entered the cells and suppressed the viability of U251cells (p53 R273H , a cell line with p53 R273H-mutation). PTEN-L-p53 is composed of endogenous protein/peptide bearing low immunogenicity, and only the junction region between PTEN-L leader and p53 can act as a new immune epitope. Accordingly, this fusion protein can potentially be used as a therapeutic option for TP53-abnormality cancers. Copyright © 2018. Published by Elsevier Inc.

The glomuvenous malformation protein Glomulin binds Rbx1 and regulates cullin RING ligase-mediated turnover of Fbw7.

Science.gov (United States)

Tron, Adriana E; Arai, Takehiro; Duda, David M; Kuwabara, Hiroshi; Olszewski, Jennifer L; Fujiwara, Yuko; Bahamon, Brittany N; Signoretti, Sabina; Schulman, Brenda A; DeCaprio, James A

2012-04-13

Fbw7, a substrate receptor for Cul1-RING-ligase (CRL1), facilitates the ubiquitination and degradation of several proteins, including Cyclin E and c-Myc. In spite of much effort, the mechanisms underlying Fbw7 regulation are mostly unknown. Here, we show that Glomulin (Glmn), a protein found mutated in the vascular disorder glomuvenous malformation (GVM), binds directly to the RING domain of Rbx1 and inhibits its E3 ubiquitin ligase activity. Loss of Glmn in a variety of cells, tissues, and GVM lesions results in decreased levels of Fbw7 and increased levels of Cyclin E and c-Myc. The increased turnover of Fbw7 is dependent on CRL and proteasome activity, indicating that Glmn modulates the E3 activity of CRL1(Fbw7). These data reveal an unexpected functional connection between Glmn and Rbx1 and demonstrate that defective regulation of Fbw7 levels contributes to GVM. Copyright © 2012 Elsevier Inc. All rights reserved.

Conditional RNA interference achieved by Oct-1 POU/rtTA fusion protein activator and a modified TRE-mouse U6 promoter

International Nuclear Information System (INIS)

Fei Zhaoliang; Chen Zheng; Wang Zhugang; Fei Jian

2007-01-01

RNA interference (RNAi) is a powerful technique and is widely used to down-regulate expression of specific genes in cultured cells and in vivo. In this paper, we report our development of a new tetracycline-inducible RNAi expression using a modified TRE-mouse U6 promoter in which the distal sequence element (DSE) was replaced by the tetracycline-responsive element (TRE). The modified TRE-mouse U6 promoter can be activated by a Tet-on version tetracycline-regulated artificial activator rTetOct which was constructed by fusing the rtTA DNA binding domain with the Oct-1 POU activation domain. This rTetOct/TRE-U6 system was successfully applied to conditionally and reversibly down-regulate the expression of endogenous p53 gene in MCF7 cells, and the expression of β-defensin gene (mBin1b) either transiently expressed in COS7 cells or stably expressed in CHO cells

Meet-U: Educating through research immersion.

Science.gov (United States)

Abdollahi, Nika; Albani, Alexandre; Anthony, Eric; Baud, Agnes; Cardon, Mélissa; Clerc, Robert; Czernecki, Dariusz; Conte, Romain; David, Laurent; Delaune, Agathe; Djerroud, Samia; Fourgoux, Pauline; Guiglielmoni, Nadège; Laurentie, Jeanne; Lehmann, Nathalie; Lochard, Camille; Montagne, Rémi; Myrodia, Vasiliki; Opuu, Vaitea; Parey, Elise; Polit, Lélia; Privé, Sylvain; Quignot, Chloé; Ruiz-Cuevas, Maria; Sissoko, Mariam; Sompairac, Nicolas; Vallerix, Audrey; Verrecchia, Violaine; Delarue, Marc; Guérois, Raphael; Ponty, Yann; Sacquin-Mora, Sophie; Carbone, Alessandra; Froidevaux, Christine; Le Crom, Stéphane; Lespinet, Olivier; Weigt, Martin; Abboud, Samer; Bernardes, Juliana; Bouvier, Guillaume; Dequeker, Chloé; Ferré, Arnaud; Fuchs, Patrick; Lelandais, Gaëlle; Poulain, Pierre; Richard, Hugues; Schweke, Hugo; Laine, Elodie; Lopes, Anne

2018-03-01

We present a new educational initiative called Meet-U that aims to train students for collaborative work in computational biology and to bridge the gap between education and research. Meet-U mimics the setup of collaborative research projects and takes advantage of the most popular tools for collaborative work and of cloud computing. Students are grouped in teams of 4-5 people and have to realize a project from A to Z that answers a challenging question in biology. Meet-U promotes "coopetition," as the students collaborate within and across the teams and are also in competition with each other to develop the best final product. Meet-U fosters interactions between different actors of education and research through the organization of a meeting day, open to everyone, where the students present their work to a jury of researchers and jury members give research seminars. This very unique combination of education and research is strongly motivating for the students and provides a formidable opportunity for a scientific community to unite and increase its visibility. We report on our experience with Meet-U in two French universities with master's students in bioinformatics and modeling, with protein-protein docking as the subject of the course. Meet-U is easy to implement and can be straightforwardly transferred to other fields and/or universities. All the information and data are available at www.meet-u.org.

Nuclear-specific AR-V7 Protein Localization is Necessary to Guide Treatment Selection in Metastatic Castration-resistant Prostate Cancer.

Science.gov (United States)

Scher, Howard I; Graf, Ryon P; Schreiber, Nicole A; McLaughlin, Brigit; Lu, David; Louw, Jessica; Danila, Daniel C; Dugan, Lyndsey; Johnson, Ann; Heller, Glenn; Fleisher, Martin; Dittamore, Ryan

2017-06-01

Circulating tumor cells (CTCs) expressing AR-V7 protein localized to the nucleus (nuclear-specific) identify metastatic castration-resistant prostate cancer (mCRPC) patients with improved overall survival (OS) on taxane therapy relative to the androgen receptor signaling inhibitors (ARSi) abiraterone acetate, enzalutamide, and apalutamide. To evaluate if expanding the positivity criteria to include both nuclear and cytoplasmic AR-V7 localization ("nuclear-agnostic") identifies more patients who would benefit from a taxane over an ARSi. The study used a cross-sectional cohort. Between December 2012 and March 2015, 193 pretherapy blood samples, 191 of which were evaluable, were collected and processed from 161 unique mCRPC patients before starting a new line of systemic therapy for disease progression at the Memorial Sloan Kettering Cancer Center. The association between two AR-V7 scoring criteria, post-therapy prostate-specific antigen (PSA) change (PTPC) and OS following ARSi or taxane treatment, was explored. One criterion required nuclear-specific AR-V7 localization, and the other required an AR-V7 signal but was agnostic to protein localization in CTCs. Correlation of AR-V7 status to PTPC and OS was investigated. Relationships with survival were analyzed using multivariable Cox regression and log-rank analyses. A total of 34 (18%) samples were AR-V7-positive using nuclear-specific criteria, and 56 (29%) were AR-V7-positive using nuclear-agnostic criteria. Following ARSi treatment, none of the 16 nuclear-specific AR-V7-positive samples and six of the 32 (19%) nuclear-agnostic AR-V7-positive samples had ≥50% PTPC at 12 weeks. The strongest baseline factor influencing OS was the interaction between the presence of nuclear-specific AR-V7-positive CTCs and treatment with a taxane (hazard ratio 0.24, 95% confidence interval 0.078-0.79; p=0.019). This interaction was not significant when nuclear-agnostic criteria were used. To reliably inform treatment selection

Protein enrichment, cellulase production and in vitro digestion improvement of pangolagrass with solid state fermentation.

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Hu, Chan-Chin; Liu, Li-Yun; Yang, Shang-Shyng

2012-02-01

Pangolagrass, Digitaria decumbens Stent, is a major grass for cow feeding, and may be a good substrate for protein enrichment. To improve the quality of pangolagrass for animal feeding, cellulolytic microbes were isolated from various sources and cultivated with solid state fermentation to enhance the protein content, cellulase production and in vitro digestion. The microbes, culture conditions and culture media were studied. Cellulolytic microbes were isolated from pangolagrass and its extracts, and composts. Pangolagrass supplemented with nitrogen and minerals was used to cultivate the cellulolytic microbes with solid state fermentation. The optimal conditions for protein enrichment and cellulase activity were pangolagrass substrate at initial moisture 65-70%, initial pH 6.0-8.0, supplementation with 2.5% (NH(4))(2)SO(4), 2.5% KH(2)PO(4) and K(2)HPO(4) mixture (2:1, w/w) and 0.3% MgSO(4).7H(2)O and cultivated at 30(o)C for 6 days. The protein content of fermented pangolagrass increased from 5.97-6.28% to 7.09-16.96% and the in vitro digestion improved from 4.11-4.38% to 6.08-19.89% with the inoculation of cellulolytic microbes by solid state fermentation. Each 1 g of dried substrate yielded Avicelase 0.93-3.76 U, carboxymethylcellulase 1.39-4.98 U and β-glucosidase 1.20-6.01 U. The isolate Myceliophthora lutea CL3 was the strain found to be the best at improving the quality of pangolagrass for animal feeding with solid state fermentation. Solid state fermentation of pangolagrass inoculated with appropriate microbes is a feasible process to enrich protein content, increase in vitro digestibility and improve the quality for animal feeding. Copyright © 2011. Published by Elsevier B.V.

CXC chemokine receptor 7 (CXCR7 regulates CXCR4 protein expression and capillary tuft development in mouse kidney.

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Sammy Haege

Full Text Available BACKGROUND: The CXCL12/CXCR4 axis is involved in kidney development by regulating formation of the glomerular tuft. Recently, a second CXCL12 receptor was identified and designated CXCR7. Although it is established that CXCR7 regulates heart and brain development in conjunction with CXCL12 and CXCR4, little is known about the influence of CXCR7 on CXCL12 dependent kidney development. METHODOLOGY/PRINCIPAL FINDINGS: We provided analysis of CXCR7 expression and function in the developing mouse kidney. Using in situ hybridization, we identified CXCR7 mRNA in epithelial cells including podocytes at all nephron stages up to the mature glomerulus. CXCL12 mRNA showed a striking overlap with CXCR7 mRNA in epithelial structures. In addition, CXCL12 was detected in stromal cells and the glomerular tuft. Expression of CXCR4 was complementary to that of CXCR7 as it occurred in mesenchymal cells, outgrowing ureteric buds and glomerular endothelial cells but not in podocytes. Kidney examination in CXCR7 null mice revealed ballooning of glomerular capillaries as described earlier for CXCR4 null mice. Moreover, we detected a severe reduction of CXCR4 protein but not CXCR4 mRNA within the glomerular tuft and in the condensed mesenchyme. Malformation of the glomerular tuft in CXCR7 null mice was associated with mesangial cell clumping. CONCLUSIONS/SIGNIFICANCE: We established that there is a similar glomerular pathology in CXCR7 and CXCR4 null embryos. Based on the phenotype and the anatomical organization of the CXCL12/CXCR4/CXCR7 system in the forming glomerulus, we propose that CXCR7 fine-tunes CXCL12/CXCR4 mediated signalling between podocytes and glomerular capillaries.

CXC Chemokine Receptor 7 (CXCR7) Regulates CXCR4 Protein Expression and Capillary Tuft Development in Mouse Kidney

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Haege, Sammy; Mueller, Wiebke; Nietzsche, Sandor; Lupp, Amelie; Mackay, Fabienne; Schulz, Stefan; Stumm, Ralf

2012-01-01

Background The CXCL12/CXCR4 axis is involved in kidney development by regulating formation of the glomerular tuft. Recently, a second CXCL12 receptor was identified and designated CXCR7. Although it is established that CXCR7 regulates heart and brain development in conjunction with CXCL12 and CXCR4, little is known about the influence of CXCR7 on CXCL12 dependent kidney development. Methodology/Principal Findings We provided analysis of CXCR7 expression and function in the developing mouse kidney. Using in situ hybridization, we identified CXCR7 mRNA in epithelial cells including podocytes at all nephron stages up to the mature glomerulus. CXCL12 mRNA showed a striking overlap with CXCR7 mRNA in epithelial structures. In addition, CXCL12 was detected in stromal cells and the glomerular tuft. Expression of CXCR4 was complementary to that of CXCR7 as it occurred in mesenchymal cells, outgrowing ureteric buds and glomerular endothelial cells but not in podocytes. Kidney examination in CXCR7 null mice revealed ballooning of glomerular capillaries as described earlier for CXCR4 null mice. Moreover, we detected a severe reduction of CXCR4 protein but not CXCR4 mRNA within the glomerular tuft and in the condensed mesenchyme. Malformation of the glomerular tuft in CXCR7 null mice was associated with mesangial cell clumping. Conclusions/Significance We established that there is a similar glomerular pathology in CXCR7 and CXCR4 null embryos. Based on the phenotype and the anatomical organization of the CXCL12/CXCR4/CXCR7 system in the forming glomerulus, we propose that CXCR7 fine-tunes CXCL12/CXCR4 mediated signalling between podocytes and glomerular capillaries. PMID:22880115

Protein Data Bank (PDB)

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U.S. Department of Health & Human Services — The Protein Data Bank (PDB) archive is the single worldwide repository of information about the 3D structures of large biological molecules, including proteins and...

Large-scale analysis of protein expression changes in human keratinocytes immortalized by human papilloma virus type 16 E6 and E7 oncogenes

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Arnouk Hilal

2009-08-01

Full Text Available Abstract Background Infection with high-risk type human papilloma viruses (HPVs is associated with cervical carcinomas and with a subset of head and neck squamous cell carcinomas. Viral E6 and E7 oncogenes cooperate to achieve cell immortalization by a mechanism that is not yet fully understood. Here, human keratinocytes were immortalized by long-term expression of HPV type 16 E6 or E7 oncoproteins, or both. Proteomic profiling was used to compare expression levels for 741 discrete protein features. Results Six replicate measurements were performed for each group using two-dimensional difference gel electrophoresis (2D-DIGE. The median within-group coefficient of variation was 19–21%. Significance of between-group differences was tested based on Significance Analysis of Microarray and fold change. Expression of 170 (23% of the protein features changed significantly in immortalized cells compared to primary keratinocytes. Most of these changes were qualitatively similar in cells immortalized by E6, E7, or E6/7 expression, indicating convergence on a common phenotype, but fifteen proteins (~2% were outliers in this regulatory pattern. Ten demonstrated opposite regulation in E6- and E7-expressing cells, including the cell cycle regulator p16INK4a; the carbohydrate binding protein Galectin-7; two differentially migrating forms of the intermediate filament protein Cytokeratin-7; HSPA1A (Hsp70-1; and five unidentified proteins. Five others had a pattern of expression that suggested cooperativity between the co-expressed oncoproteins. Two of these were identified as forms of the small heat shock protein HSPB1 (Hsp27. Conclusion This large-scale analysis provides a framework for understanding the cooperation between E6 and E7 oncoproteins in HPV-driven carcinogenesis.

Schedule-dependent inhibition of hypoxia-inducible factor-1alpha protein accumulation, angiogenesis, and tumor growth by topotecan in U251-HRE glioblastoma xenografts.

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Rapisarda, Annamaria; Zalek, Jessica; Hollingshead, Melinda; Braunschweig, Till; Uranchimeg, Badarch; Bonomi, Carrie A; Borgel, Suzanne D; Carter, John P; Hewitt, Stephen M; Shoemaker, Robert H; Melillo, Giovanni

2004-10-01

We have previously shown that topotecan, a topoisomerase I poison, inhibits hypoxia-inducible factor (HIF)-1alpha protein accumulation by a DNA damage-independent mechanism. Here, we report that daily administration of topotecan inhibits HIF-1alpha protein expression in U251-HRE glioblastoma xenografts. Concomitant with HIF-1alpha inhibition, topotecan caused a significant tumor growth inhibition associated with a marked decrease of angiogenesis and expression of HIF-1 target genes in tumor tissue. These results provide a compelling rationale for testing topotecan in clinical trials to target HIF-1 in cancer patients.

Transformation temperatures γU → δUZr2 Y γU →αU in U-Zr-Nb alloys

International Nuclear Information System (INIS)

Komar Varela, Carolina L.; Arico, Sergio F.; Gribaudo, Luis M.; Comision Nacional de Energia Atomica, General San Martin

2009-01-01

The international RERTR program has as primary objective the development of new fuels for research and test reactors to satisfy the requirement of low enrichment in 235 U (lower than 20 %). It is known that the cubic-phase (γU) has an excellent behaviour under irradiation. In this context, in the Materials Department (GIDAT-GAEN-CNEA) U-Zr-Nb alloys are considered candidates for the development of a high-density monolithic-type nuclear fuel. It is necessary to evaluate the thermodynamic parameters that allow to obtain a range of concentrations of the U-Zr-Nb system in which this phase can be retained in the metastable condition with the required 235 U density. In this work, eight U alloys with concentrations ranging from 13.9 to 43.7 wt.% Zr and from 0 to 6.4 wt.% Nb, were fabricated. Dynamical measurements of electrical resistivity, with a cooling rate of 4 o C/min, were performed and the results were analyzed. Considering this cooling rate, a Nb concentration of at least 17.8 wt. % would inhibit the transformation γU→ δUZr 2 and a concentration of al least 23.3 wt % would inhibit the γU → αU transformation. (author)

Genomic organization of the rat alpha 2u-globulin gene cluster.

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McFadyen, D A; Addison, W; Locke, J

1999-05-01

The alpha 2u-globulin are a group of similar proteins, belonging to the lipocalin superfamily of proteins, that are synthesized in a subset of secretory tissues in rats. The many alpha 2u-globulin isoforms are encoded by a multigene family that exhibits extensive homology. Despite a high degree of sequence identity, individual family members show diverse expression patterns involving complex hormonal, tissue-specific, and developmental regulation. Analysis suggests that there are approximately 20 alpha 2u-globulin genes in the rat genome. We have used fluorescence in situ hybridization (FISH) to show that the alpha 2u-globulin genes are clustered at a single site on rat Chromosome (Chr) 5 (5q22-24). Southern blots of rat genomic DNA separated by pulsed field gel electrophoresis indicated that the alpha 2u-globulin genes are contained on two NruI fragments with a total size of 880 kbp. Analysis of three P1 clones containing alpha 2u-globulin genes indicated that the alpha 2u-globulin genes are tandemly arranged in a head-to-tail fashion. The organization of the alpha 2u-globulin genes in the rat as a tandem array of single genes differs from the homologous major urinary protein genes in the mouse, which are organized as tandem arrays of divergently oriented gene pairs. The structure of these gene clusters may have consequences for the proposed function, as a pheromone transporter, for the protein products encoded by these genes.

Vaccinia virus protein C6 is a virulence factor that binds TBK-1 adaptor proteins and inhibits activation of IRF3 and IRF7.

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Leonie Unterholzner

2011-09-01

Full Text Available Recognition of viruses by pattern recognition receptors (PRRs causes interferon-β (IFN-β induction, a key event in the anti-viral innate immune response, and also a target of viral immune evasion. Here the vaccinia virus (VACV protein C6 is identified as an inhibitor of PRR-induced IFN-β expression by a functional screen of select VACV open reading frames expressed individually in mammalian cells. C6 is a member of a family of Bcl-2-like poxvirus proteins, many of which have been shown to inhibit innate immune signalling pathways. PRRs activate both NF-κB and IFN regulatory factors (IRFs to activate the IFN-β promoter induction. Data presented here show that C6 inhibits IRF3 activation and translocation into the nucleus, but does not inhibit NF-κB activation. C6 inhibits IRF3 and IRF7 activation downstream of the kinases TANK binding kinase 1 (TBK1 and IκB kinase-ε (IKKε, which phosphorylate and activate these IRFs. However, C6 does not inhibit TBK1- and IKKε-independent IRF7 activation or the induction of promoters by constitutively active forms of IRF3 or IRF7, indicating that C6 acts at the level of the TBK1/IKKε complex. Consistent with this notion, C6 immunoprecipitated with the TBK1 complex scaffold proteins TANK, SINTBAD and NAP1. C6 is expressed early during infection and is present in both nucleus and cytoplasm. Mutant viruses in which the C6L gene is deleted, or mutated so that the C6 protein is not expressed, replicated normally in cell culture but were attenuated in two in vivo models of infection compared to wild type and revertant controls. Thus C6 contributes to VACV virulence and might do so via the inhibition of PRR-induced activation of IRF3 and IRF7.

Design and simulation for the pulse high-voltage DC power supply (HVPS) of 1.2 MW/2.45 GHz HT-7U lower hybrid current drive system

International Nuclear Information System (INIS)

Huang Yiyun; Kuang Guangli; Xu Weihua; Liu Baohua; Lin Jianan; Wu Junshuan; Zheng Guanghua; Yang Chunshen

2000-01-01