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Sample records for al gen env

  1. Revision of J3Gen and Validity of the Attacks by Peinado et al.

    PubMed

    Peinado, Alberto; Munilla, Jorge; Fúster-Sabater, Amparo

    2015-05-22

    This letter is the reply to: Remarks on Peinado et al.'s Analysis of J3Gen by J. Garcia-Alfaro, J. Herrera-Joancomartí and J. Melià-Seguí published in Sensors 2015, 15, 6217-6220. Peinado et al. cryptanalyzed the pseudorandom number generator proposed by Melià-Seguí et al., describing two possible attacks. Later, Garcia-Alfaro claimed that one of this attack did not hold in practice because the assumptions made by Peinado et al. were not correct. This letter reviews those remarks, showing that J3Gen is anyway flawed and that, without further information, the interpretation made by Peinado et al. seems to be correct.

  2. 2nd Gen FeCrAl ODS Alloy Development For Accident-Tolerant Fuel Cladding

    SciTech Connect

    Dryepondt, Sebastien N.; Massey, Caleb P.; Edmondson, Philip D.

    2016-08-01

    Extensive research at ORNL aims at developing advanced low-Cr high strength FeCrAl alloys for accident tolerant fuel cladding. One task focuses on the fabrication of new low Cr oxide dispersion strengthened (ODS) FeCrAl alloys. The first Fe-12Cr-5Al+Y2O3 (+ ZrO2 or TiO2) ODS alloys exhibited excellent tensile strength up to 800 C and good oxidation resistance in steam up to 1400 C, but very limited plastic deformation at temperature ranging from room to 800 C. To improve alloy ductility, several fabrication parameters were considered. New Fe-10-12Cr-6Al gas-atomized powders containing 0.15 to 0.5wt% Zr were procured and ball milled for 10h, 20h or 40h with Y2O3. The resulting powder was then extruded at temperature ranging from 900 to 1050 C. Decreasing the ball milling time or increasing the extrusion temperature changed the alloy grain size leading to lower strength but enhanced ductility. Small variations of the Cr, Zr, O and N content did not seem to significantly impact the alloy tensile properties, and, overall, the 2nd gen ODS FeCrAl alloys showed significantly better ductility than the 1st gen alloys. Tube fabrication needed for fuel cladding will require cold or warm working associated with softening heat treatments, work was therefore initiated to assess the effect of these fabrications steps on the alloy microstructure and properties. This report has been submitted as fulfillment of milestone M3FT 16OR020202091 titled, Report on 2nd Gen FeCrAl ODS Alloy Development for the Department of Energy Office of Nuclear Energy, Advanced Fuel Campaign of the Fuel Cycle R&D program.

  3. Optimized Gen-II FeCrAl cladding production in large quantity for campaign testing

    SciTech Connect

    Yamamoto, Yukinori; Sun, Zhiqian; Pint, Bruce A.; Terrani, Kurt A.

    2016-06-03

    There are two major objectives in this report; (1) to optimize microstructure control of ATF FeCrAl alloys during tube drawing processes, and (2) to provide an update on the progress of ATF FeCrAl tube production via commercial manufacturers. Experimental efforts have been made to optimize the process parameters balancing the tube fabricability, especially for tube drawing processes, and microstructure control of the final tube products. Lab-scale sheet materials of Gen II FeCrAl alloys (Mo-containing and Nb-containing FeCrAl alloys) were used in the study, combined with a stepwise warm-rolling process and intermediate annealing, aiming to simulate the tube drawing process in a commercial tube manufacturer. The intermediate annealing at 650ºC for 1h was suggested for the tube-drawing process of Mo-containing FeCrAl alloys because it successfully softened the material by recovering the work hardening introduced through the rolling step, without inducing grain coarsening due to recrystallization. The final tube product is expected to have stabilized deformed microstructure providing the improved tensile properties with sufficient ductility. Optimization efforts on Nb-containing FeCrAl alloys focused on the effect of alloying additions and annealing conditions on the stability of deformed microstructure. Relationships between the second-phase precipitates (Fe2Nb-Laves phase) and microstructure stability are discussed. FeCrAl tube production through commercial tube manufacturers is currently in progress. Three different manufacturers, Century Tubes, Inc. (CTI), Rhenium Alloys, Inc. (RAI), and Superior Tube Company, Inc. (STC), are providing capabilities for cold-drawing, warm-drawing, and HPTR cold-pilgering, respectively. The first two companies are currently working on large quantity tube production (expected 250 ft length) of Gen I model FeCrAl alloy (B136Y3, at CTI) and Gen II (C35M4, at RAI), with the process parameters obtained from the experimental

  4. A proposal to reclassify Nocardia pinensis Blackall et al. as Skermania piniformis gen. nov., comb. nov.

    PubMed

    Chun, J; Blackall, L L; Kang, S O; Hah, Y C; Goodfellow, M

    1997-01-01

    The type strain of Nocardia pinensis was the subject of chemotaxonomic and 16S ribosomal DNA sequencing studies. The resultant nucleotide sequence was aligned with the sequences of representatives of the genera Corynebacterium, Dietzia, Gordona, Mycobacterium, Nocardia, Rhodococcus, and Tsukamurella, and phylogenetic trees were generated by using the Fitch-Margoliash, maximum-parsimony, maximum-likelihood, and neighbor-joining methods. It was evident from the phylogenetic analyses that N. pinensis represents a distinct phyletic line that is most closely associated with the Gordona clade. This genealogical evidence, together with chemotaxonomic and phenotypic data derived from this and previous studies, indicates that N. pinensis merits generic status within the family Nocardiaceae. Therefore, we propose that N. pinensis Blackall et al. 1989 be reclassified as Skermania piniformis gen. nov., comb. nov. The type strain of Skermania piniformis cleaved an array of conjugated substrates based on the fluorophores 7-amino-4-methylcoumarin and 4-methylumbelliferone.

  5. ALV-J GP37 molecular analysis reveals novel virus-adapted sites and three tyrosine-based Env species.

    PubMed

    Ye, Jianqiang; Fan, Zhonglei; Shang, Jianjun; Tian, Xiaoyan; Yang, Jialiang; Chen, Hongjun; Shao, Hongxia; Qin, Aijian

    2015-01-01

    Compared to other avian leukosis viruses (ALV), ALV-J primarily induces myeloid leukemia and hemangioma and causes significant economic loss for the poultry industry. The ALV-J Env protein is hypothesized to be related to its unique pathogenesis. However, the molecular determinants of Env for ALV-J pathogenesis are unclear. In this study, we compared and analyzed GP37 of ALV-J Env and the EAV-HP sequence, which has high homology to that of ALV-J Env. Phylogenetic analysis revealed five groups of ALV-J GP37 and two novel ALV-J Envs with endemic GP85 and EAV-HP-like GP37. Furthermore, at least 15 virus-adapted mutations were detected in GP37 compared to the EAV-HP sequence. Further analysis demonstrated that three tyrosine-based motifs (YxxM, ITIM (immune tyrosine-based inhibitory motif) and ITAM-like (immune tyrosine-based active motif like)) associated with immune disease and oncogenesis were found in the cytoplasmic tail of GP37. Based on the potential function and distribution of these motifs in GP37, ALV-J Env was grouped into three species, inhibitory Env, bifunctional Env and active Env. Accordingly, 36.91%, 61.74% and 1.34% of ALV-J Env sequences from GenBank are classified as inhibitory, bifunctional and active Env, respectively. Additionally, the Env of the ALV-J prototype strain, HPRS-103, and 17 of 18 EAV-HP sequences belong to the inhibitory Env. And models for signal transduction of the three ALV-J Env species were predicted. Our findings and models provide novel insights for identifying the roles and molecular mechanism of ALV-J Env in the unique pathogenesis of ALV-J.

  6. Youngimonas vesicularis gen. nov., sp. nov., of the family Rhodobacteraceae, isolated from surface seawater, reclassification of Donghicola xiamenensis Tan et al. 2009 as Pseudodonghicola xiamenensis gen. nov., comb. nov. and emended description of the genus Donghicola Yoon et al. 2007.

    PubMed

    Hameed, Asif; Shahina, Mariyam; Lin, Shih-Yao; Nakayan, Phanit; Liu, You-Cheng; Lai, Wei-An; Hsu, Yi-Han

    2014-08-01

    A Gram-staining-negative, non-pigmented, strictly aerobic, rod-shaped, non-spore-forming, non-motile bacterium, devoid of bacteriochlorophyll, designated strain CC-AMW-E(T), was isolated from surface seawater off the coast at Kending, Taiwan. Strain CC-AMW-E(T) shared 95.7 and 93.9% 16S rRNA gene sequence similarity, respectively, with the type strains of the type species of the genera Donghicola (Donghicola eburneus SW-277(T)) and Roseovarius (Roseovarius tolerans EL-172(T)). The predominant (>75% of the total) fatty acid was summed feature 8 (C(18 : 1)ω6c and/or C(18 : 1)ω7c). The polar lipid profile included major amounts of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine and an unidentified aminolipid. In addition, moderate amounts of an unidentified lipid and trace amounts of an unidentified phospholipid were detected. The DNA G+C content was 67.9 mol%. Ubiquinone Q-10 was the sole respiratory quinone. Based on its phylogenetic distinctiveness and distinguishing phenotypic characteristics (in particular its polar lipid pattern), we conclude that strain CC-AMW-E(T) represents a novel genus and species of the family Rhodobacteraceae, for which the name Youngimonas vesicularis gen. nov., sp. nov. is proposed. The type strain of Youngimonas vesicularis is CC-AMW-E(T) ( = JCM 18819(T) = BCRC 80549(T)). In addition, an emended description of the genus Donghicola Yoon et al. 2007 and the reclassification of Donghicola xiamenensis Tan et al. 2009 as Pseudodonghicola xiamenensis gen. nov., comb. nov. (type strain Y-2(T) = MCCC 1A00107(T) = LMG 24574(T) = CGMCC 1.7081(T)) are proposed.

  7. Reclassification of Geobacillus pallidus (Scholz et al. 1988) Banat et al. 2004 as Aeribacillus pallidus gen. nov., comb. nov.

    PubMed

    Miñana-Galbis, David; Pinzón, Dora L; Lorén, J Gaspar; Manresa, Angels; Oliart-Ros, Rosa M

    2010-07-01

    Although Anoxybacillus and Geobacillus, two genera of thermophilic bacteria close to the genus Bacillus, have only been described recently, the number of species in these genera has increased rapidly. Four thermophilic, lipolytic strains (DR01, DR02, DR03 and DR04) isolated from a hot spring in Veracruz (Mexico), which could not be identified phenotypically, were subjected to 16S rRNA gene sequence analysis. Three strains were identified as belonging to the genus Anoxybacillus, but strain DR03 was identified as Geobacillus pallidus. This result led us to perform a phylogenetic analysis of the genera Anoxybacillus and Geobacillus based on 16S rRNA gene sequences from all the type strains of these genera. Phylogenetic trees showed three major clusters, Anoxybacillus-Geobacillus tepidamans, Geobacillus sensu stricto and Geobacillus pallidus, while the 16S rRNA gene sequences of G. pallidus (DR03 and the type strain) showed low similarity to sequences of Anoxybacillus (92.5-95.1 %) and Geobacillus (92.8-94.5 %) species, as well as to Bacillus subtilis (92.2-92.4 %). In addition, G. pallidus could be differentiated from Anoxybacillus and Geobacillus on the basis of DNA G+C content and fatty acid and polar lipid profiles. From these results, it is proposed that Geobacillus pallidus should be classified in a novel genus, for which we propose the name Aeribacillus, as Aeribacillus pallidus gen. nov., comb. nov. The type strain of Aeribacillus pallidus is H12(T) (=ATCC 51176(T) =DSM 3670(T) =LMG 19006(T)).

  8. Roseitalea porphyridii gen. nov., sp. nov., isolated from a red alga, and reclassification of Hoeflea suaedae Chung et al. 2013 as Pseudohoeflea suaedae gen. nov., comb. nov.

    PubMed

    Hyeon, Jong Woo; Jeong, Sang Eun; Baek, Kyunghwa; Jeon, Che Ok

    2017-02-01

    A Gram-staining-negative, strictly aerobic bacterial strain, designated MA7-20T, was isolated from a marine alga, Porphyridium marinum, in Korea. Cells showing oxidase-positive and catalase-positive activities were motile rods with bipolar flagella. Growth of strain MA7-20T was observed at 15-45 °C (optimum, 30-37 °C), at pH 6.0-10.5 (optimum, pH 7.0-8.0) and in the presence of 0-7 % (w/v) NaCl (optimum, 2-3 %). Strain MA7-20T contained summed feature 8 (comprising C18 : 1ω7c/C18 : 1ω6c), 11-methyl C18 : 1ω7c and C18 : 0 as the major fatty acids and ubiquinone-10 as the sole isoprenoid quinone. The major polar lipids were phosphatidylcholine, phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine and phosphatidyl-N-methylethanolamine. The G+C content of the genomic DNA was 61.5 mol%. Strain MA7-20T was most closely related to Hoeflea suaedae YC6898T, Oricola cellulosilytica CC-AMH-0T and Nitratireductor basaltis J3T with 96.0, 95.8 and 95.8 % 16S rRNA gene sequence similarities, respectively, but the strain formed a distinct phylogenetic lineage from them within the family Phyllobacteriaceae with a low bootstrap value. H. suaedae also formed a clearly distinct phylogenetic lineage from other members of the genus Hoeflea and closely related genera. On the basis of phenotypic, chemotaxonomic and molecular properties, strain MA7-20T represents a novel species of a new genus of the family Phyllobacteriaceae, for which the name Roseitalea porphyridii gen. nov., sp. nov. is proposed. The type strain is MA7-20T (=KACC 18807T=JCM 31538T). In addition, H. suaedae is also reclassified as Pseudohoeflea suaedae gen. nov., comb. nov. (type strain YC6898T=KACC 14911T=NBRC 107700T).

  9. Leifsonia poae gen. nov., sp. nov., isolated from nematode galls on Poa annua, and reclassification of 'Corynebacterium aquaticum' Leifson 1962 as Leifsonia aquatica (ex Leifson 1962) gen. nov., nom. rev., comb. nov. and Clavibacter xyli Davis et al. 1984 with two subspecies as Leifsonia xyli (Davis et al. 1984) gen. nov., comb. nov.

    PubMed

    Evtushenko, L I; Dorofeeva, L V; Subbotin, S A; Cole, J R; Tiedje, J M

    2000-01-01

    The new genus Leifsonia gen. nov. with two new species, Leifsonia poae sp. nov. (type strain VKM Ac-1401T) and Leifsonia aquatica (ex Leifson 1962) nom. rev., comb. nov. (the type species, with VKM Ac-1400T = DSM 20146T = JCM 1368T as type strain), is proposed to accommodate bacteria found in Poa annua root gall, induced by the nematode Subanguina radicicola, and 'Corynebacterium aquaticum' Leifson 1962. Further, it is proposed to reclassify Clavibacter xyli Davis et al. 1984 with two subspecies in the new genus as Leifsonia xyli (Davis et al. 1984) comb. nov., Leifsonia xyli subsp. xyli (Davis et al. 1984) comb. nov. and Leifsonia xyli subsp. cynodontis (Davis et al. 1984) comb. nov. Members of the proposed genus are characterized by coryneform morphology, peptidoglycans based upon 2,4-diaminobutyric acid, the major menaquinone MK-11, phosphatidylglycerol and diphosphatidylglycerol as principal phospholipids, the high content of anteiso- and iso-branched saturated fatty acids, and a DNA G+C base composition of 66-73 mol%. They form a distinct phylogenetic branch attached to the line of descent of Agromyces spp. The new and reclassified species of the new genus clearly differ from each other phylogenetically and phenetically and can be recognized by their morphologies, the cell wall sugar composition, the requirement of complex media for growth, and numerous physiological characteristics, including the oxidase reaction.

  10. Mobilicoccus pelagius gen. nov., sp. nov. and Piscicoccus intestinalis gen. nov., sp. nov., two new members of the family Dermatophilaceae, and reclassification of Dermatophilus chelonae (Masters et al. 1995) as Austwickia chelonae gen. nov., comb. nov.

    PubMed

    Hamada, Moriyuki; Iino, Takao; Iwami, Takahiro; Harayama, Shigeaki; Tamura, Tomohiko; Suzuki, Ken-ichiro

    2010-01-01

    Two Gram-positive bacteria, designated strains Aji5-31(T) and Ngc37-23(T), were isolated from the intestinal tracts of fishes. 16S rRNA gene sequence analysis indicated that both strains were related to the members of the family Dermatophilaceae, with 95.6-96.9% 16S rRNA gene sequence similarities. The family Dermatophilaceae contains 2 genera and 3 species: Dermatophilus congolensis, Dermatophilus chelonae and Kineosphaera limosa. However, it has been suggested that the taxonomic position of D. chelonae should be reinvestigated using a polyphasic approach, because the chemotaxonomic characteristics are not known (Stackebrandt, 2006; Stackebrandt and Schumann, 2000). Our present study revealed that strains Aji5-31(T), Ngc37-23(T) and D. chelonae NBRC 105200(T) should be separated from the other members of the family Dermatophilaceae on the basis of the following characteristics: the predominant menaquinone of strain Aji5-31(T) is MK-8(H(2)), strain Ngc37-23(T) possesses iso- branched fatty acids as major components, and the menaquinone composition of D. chelonae is MK-8(H(4)), MK-8 and MK-8(H(2)) (5 : 3 : 2, respectively). On the basis of these distinctive phenotypic characteristics and phylogenetic analysis results, it is proposed that strains Aji5-31(T) and Ngc37-23(T) be classified as two novel genera and species of the family Dermatophilaceae. The names are Mobilicoccus pelagius gen. nov., sp. nov. and Piscicoccus intestinalis gen. nov., sp. nov., and the type strains are Aji5-31(T) (=NBRC 104925(T) =DSM 22762(T)) and Ngc37-23(T) (=NBRC 104926(T) =DSM 22761(T)), respectively. In addition, D. chelonae should be reassigned to a new genus of the family Dermatophilaceae with the name Austwickia chelonae gen. nov., comb. nov.

  11. Description of Alloprevotella rava gen. nov., sp. nov., isolated from the human oral cavity, and reclassification of Prevotella tannerae Moore et al. 1994 as Alloprevotella tannerae gen. nov., comb. nov.

    PubMed

    Downes, Julia; Dewhirst, Floyd E; Tanner, Anne C R; Wade, William G

    2013-04-01

    Five strains of anaerobic, gram-negative bacilli isolated from the human oral cavity were subjected to a comprehensive range of phenotypic and genotypic tests and were found to comprise a homogeneous group. Phylogenetic analysis of full-length 16S rRNA gene sequences showed that these strains represented a novel group within the family Prevotellaceae, and the most closely related species was Prevotella tannerae. P. tannerae and the novel taxon are deeply branched from the genus Prevotella, with sequence identities to the type strain of the type species of Prevotella, Prevotella melaninogenica, of 82.2 and 85.6 %, respectively. The novel genus Alloprevotella gen. nov. is proposed to accommodate the novel species Alloprevotella rava gen. nov., sp. nov. and the previously named Prevotella tannerae Moore et al. 1994 as Alloprevotella tannerae gen. nov., comb. nov. The type species is Alloprevotella tannerae. The type strain of Alloprevotella rava is 81/4-12(T) ( = DSM 22548(T)  = CCUG 58091(T)) and the type strain of Alloprevotella tannerae is ATCC 51259(T)  = CCUG 34292(T)  = CIP 104476(T)  = NCTC 13073(T). Alloprevotella rava is weakly to moderately saccharolytic and produces moderate amounts of acetic acid and major amounts of succinic acid as end products of fermentation. Strains are sensitive to 20 % bile and hydrolyse gelatin. The principal cellular long-chain fatty acids are anteiso-C15 : 0, iso-C15 : 0, C16 : 0, iso-C17 : 0 and iso-C17 : 0 3-OH. The G+C content of the DNA of the type strain is 47 mol%.

  12. Description of Mogibacterium pumilum gen. nov., sp. nov. and Mogibacterium vescum gen. nov., sp. nov., and reclassification of Eubacterium timidum (Holdeman et al. 1980) as Mogibacterium timidum gen. nov., comb. nov.

    PubMed

    Nakazawa, F; Sato, M; Poco, S E; Hashimura, T; Ikeda, T; Kalfas, S; Sundqvist, G; Hoshino, E

    2000-03-01

    A new genus, Mogibacterium, is proposed for anaerobic, non-spore-forming, Gram-positive, rod-shaped bacteria which have been isolated from the periodontal pockets of adult human patients with periodontal disease and infected root canals. The novel isolates, strains D2-18T, BA11a-f and D5-2T, were inert in most of the conventional biochemical tests and phenotypically resemble asaccharolytic Eubacterium species. The protein profiles of whole cells on SDS-PAGE gels and Western immunoblotting reaction analysis distinguished these organisms from type strains belonging to the previously described Eubacterium species. The G + C content of the DNA is 45-46 mol% for Mogibacterium pumilum and 46 mol% for Mogibacterium vescum. The levels of DNA-DNA relatedness of these new species to other Eubacterium species, including Eubacterium limosum, Eubacterium brachy, Eubacterium lentum, Eubacterium nodatum, Eubacterium saphenum, and the more recently proposed Eubacterium minutum and Eubacterium exiguum (reclassified as Slackia exigua), are less than 2%. The DNA-DNA hybridization value between M. pumilum and M. vescum was 30%. Eubacterium timidum exhibited DNA homologies with Mogibacterium species which were low (17 and 18%) but clearly higher than with all the other Eubacterium species. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the closest phylogenetic neighbour of Mogibacterium species was E. timidum, and that these three species represent a novel lineage distinct from the previously described genera of Gram-positive, rod-shaped bacteria. On the basis of phenotypic characteristics and 16S rRNA gene sequence comparisons, it is also proposed that E. timidum is transferred to the genus Mogibacterium gen. nov. as Mogibacterium timidum gen. nov., comb. nov. (type strain ATCC 33093T).

  13. Quantifying CD4/CCR5 Usage Efficiency of HIV-1 Env Using the Affinofile System.

    PubMed

    Webb, Nicholas E; Lee, Benhur

    2016-01-01

    Entry of HIV-1 into target cells involves the interaction of the HIV envelope (Env) with both a primary receptor (CD4) and a coreceptor (CXCR4 or CCR5). The relative efficiency with which a particular Env uses these receptors is a major component of cellular tropism in the context of entry and is related to a variety of pathological Env phenotypes (Chikere et al. Virology 435:81-91, 2013). The protocols outlined in this chapter describe the use of the Affinofile system, a 293-based dual-inducible cell line that expresses up to 25 distinct combinations of CD4 and CCR5, as well as the associated Viral Entry Receptor Sensitivity Assay (VERSA) metrics used to summarize the CD4/CCR5-dependent infectivity results. This system allows for high-resolution profiling of CD4 and CCR5 usage efficiency in the context of unique viral phenotypes.

  14. Proposal to replace the illegitimate genus name Prescottia Jones et al. 2013 with the genus name Prescottella gen. nov. and to replace the illegitimate combination Prescottia equi Jones et al. 2013 with Prescottella equi comb. nov.

    PubMed

    Jones, Amanda L; Sutcliffe, Iain C; Goodfellow, Michael

    2013-06-01

    Recently we proposed that Rhodococcus equi (Magnusson 1923) Goodfellow and Alderson 1977 be transferred to a novel genus, Prescottia, as Prescottia equi comb. nov. However, in accordance with Principle 2 and Rule 51b(4) of the Bacteriological Code (1990 Revision), the bacterial genus name Prescottia Jones et al. 2013 is deemed illegitimate as this name has been used previously for a plant genus within the family Orchidaceae. Consequently, a new genus name, Prescottella gen. nov. is proposed for the bacterial taxon and a new combination Prescottella equi comb. nov. is proposed for the type species.

  15. Retroviral env glycoprotein trafficking and incorporation into virions.

    PubMed

    Murakami, Tsutomu

    2012-01-01

    Together with the Gag protein, the Env glycoprotein is a major retroviral structural protein and is essential for forming infectious virus particles. Env is synthesized, processed, and transported to certain microdomains at the plasma membrane and takes advantage of the same host machinery for its trafficking as that used by cellular glycoproteins. Incorporation of Env into progeny virions is probably mediated by the interaction between Env and Gag, in some cases with the additional involvement of certain host factors. Although several general models have been proposed to explain the incorporation of retroviral Env glycoproteins into virions, the actual mechanism for this process is still unclear, partly because structural data on the Env protein cytoplasmic tail is lacking. This paper presents the current understanding of the synthesis, trafficking, and virion incorporation of retroviral Env proteins.

  16. Nucleotide Sequence of the Akv env Gene

    PubMed Central

    Lenz, Jack; Crowther, Robert; Straceski, Anthony; Haseltine, William

    1982-01-01

    The sequence of 2,191 nucleotides encoding the env gene of murine retrovirus Akv was determined by using a molecular clone of the Akv provirus. Deduction of the encoded amino acid sequence showed that a single open reading frame encodes a 638-amino acid precursor to gp70 and p15E. In addition, there is a typical leader sequence preceding the amino terminus of gp70. The locations of potential glycosylation sites and other structural features indicate that the entire gp70 molecule and most of p15E are located on the outer side of the membrane. Internal cleavage of the env precursor to generate gp70 and p15E occurs immediately adjacent to several basic amino acids at the carboxyl terminus of gp70. This cleavage generates a region of 42 uncharged, relatively hydrophobic amino acids at the amino terminus of p15E, which is located in a position analogous to the hydrophobic membrane fusion sequence of influenza virus hemagglutinin. The mature polypeptides are predicted to associate with the membrane via a region of 30 uncharged, mostly hydrophobic amino acids located near the carboxyl terminus of p15E. Distal to this membrane association region is a sequence of 35 amino acids at the carboxyl terminus of the env precursor, which is predicted to be located on the inner side of the membrane. By analogy to Moloney murine leukemia virus, a proteolytic cleavage in this region removes the terminal 19 amino acids, thus generating the carboxyl terminus of p15E. This leaves 15 amino acids at the carboxyl terminus of p15E on the inner side of the membrane in a position to interact with virion cores during budding. The precise location and order of the large RNase T1-resistant oligonucleotides in the env region were determined and compared with those from several leukemogenic viruses of AKR origin. This permitted a determination of how the differences in the leukemogenic viruses affect the primary structure of the env gene products. PMID:6283170

  17. Putative role of Tat-Env interaction in HIV infection.

    PubMed

    Poon, Selina; Moscoso, Carlos G; Xing, Li; Kan, Elaine; Sun, Yide; Kolatkar, Prasanna R; Vahlne, Anders G; Srivastava, Indresh K; Barnett, Susan W; Cheng, R Holland

    2013-09-24

    To study the complex formed between Tat protein and Env soluble trimeric immunogen, and compare with previously determined structures of Env native trimers and Env-CD4m complexes. The soluble Env trimer was used to mimic the spike glycoprotein on the virus surface for the study. To overcome limitations of other structural determination methods, cryoelectron microscopy was employed to image the complex, and single particle reconstruction was utilized to reconstruct the structure of the complex from collected micrographs. Molecular modeling of gp120-Tat was performed to provide atomic coordinates for docking. Images were preprocessed by multivariate statistical analysis to identify principal components of variation then submitted for reconstruction. Reconstructed structures were docked with modeled gp120-Tat atomic coordinates to study the positions of crucial epitopes. Analysis of the Env-Tat complex demonstrated an intermediate structure between Env native trimers and Env-CD4m structures. Docking results indicate that the CD4-binding site and the V3 loop are exposed in the Env-Tat complex. The integrin-binding sequence in Tat was also exposed in Env-Tat docking. The intermediate structure induced by Tat-interaction with Env could potentially provide an explanation for increased virus infection in the presence of Tat protein. Consequently, exposure of CD4-binding sites and a putative integrin-binding sequence on Tat in the complex may provide a new avenue for rational design of an effective HIV vaccine. © 2013 Wolters Kluwer Health | Lippincott Williams & Wilkins

  18. Proposal to replace the illegitimate genus name Bryantella Wolin et al. 2004VP with the genus name Marvinbryantia gen. nov. and to replace the illegitimate combination Bryantella formatexigens Wolin et al. 2004VP with Marvinbryantia formatexigens comb. nov.

    PubMed

    Wolin, Meyer J; Miller, Terry L; Lawson, Paul A

    2008-03-01

    The prokaryote generic name Bryantella Wolin et al. 2004(VP) is illegitimate because it is a later homonym of Bryantella Chickering, 1946 (Animalia, Arthropoda, Arachnida, Araneae, Salticidae, Dendryphantinae, Dendryphantini) and a later homonym of Bryantella Britton, 1957 (Animalia, Arthropoda, Scarabaeoidea, Scarabaeidae, Melolonthinae) [Principle 2, Rule 51b(4) of the Bacteriological Code (1990 Revision)]. Bryantella represents a genus of jumping spiders within the family Salticidae and a genus of melolonthine scarab beetles within the family Scarabaeidae. Therefore, a new genus name, Marvinbryantia gen. nov., is proposed for this taxon. As a result, a new combination, Marvinbryantia formatexigens comb. nov., is required for the type species to replace the illegitimate combination Bryantella formatexigens Wolin et al. 2004(VP).

  19. Streptacidiphilus gen. nov., acidophilic actinomycetes with wall chemotype I and emendation of the family Streptomycetaceae (Waksman and Henrici (1943)AL) emend. Rainey et al. 1997.

    PubMed

    Kim, Seung Bum; Lonsdale, John; Seong, Chi-Nam; Goodfellow, Michael

    2003-01-01

    The taxonomic position of acidophilic actinomycetes selectively isolated from acidic soils and litter was examined using a polyphasic approach. The distinct 16S rDNA phyletic branch formed by representative strains was equated with related monophyletic clades that corresponded to the genera Kitasatospora and Streptomyces. The acidophilic isolates also exhibited a distinctive pH profile, a unique 16S rDNA signature, and contained major amounts of LL-diaminopimelic acid, galactose and rhamnose in whole-organism hydrolysates. It is proposed that these acidophilic actinomycetes be assigned to a new genus, Streptacidiphilus gen. nov., on the basis of genotypic and phenotypic differences. Three species were defined on the basis of DNA:DNA pairing and phenotypic data, namely, Streptacidiphilus albus sp. nov., the type species, Streptacidiphilus neutrinimicus sp. nov. and Streptacidiphilus carbonis sp. nov. Members of the genera Kitasatospora, Streptacidiphilus and Streptomyces share a number of key characteristics and form a stable monophyletic branch in the 16S rDNA tree. It is, therefore, proposed that the description of the family Streptomycetaceae be emended to account for properties shown by Kitasatospora and Streptacidiphilus species.

  20. Reclassification of Lactobacillus catenaformis (Eggerth 1935) Moore and Holdeman 1970 and Lactobacillus vitulinus Sharpe et al. 1973 as Eggerthia catenaformis gen. nov., comb. nov. and Kandleria vitulina gen. nov., comb. nov., respectively.

    PubMed

    Salvetti, Elisa; Felis, Giovanna E; Dellaglio, Franco; Castioni, Anna; Torriani, Sandra; Lawson, Paul A

    2011-10-01

    The development of molecular tools and in particular the use of 16S rRNA gene sequencing has had a profound effect on the taxonomy of many bacterial groups. Gram-positive organisms that encompass the genera Lactobacillus and Clostridium within the Firmicutes are examples of taxa that have undergone major revisions based on phylogenetic information. A consequence of these reorganizations is that a number of organisms are now recognized as being misclassified. Previous studies have demonstrated that Lactobacillus catenaformis and Lactobacillus vitulinus are phylogenetically unrelated to Lactobacillus sensu stricto, being placed within the Clostridia rRNA cluster XVII. Based on the phenotypic, chemotaxonomic and phylogenetic data presented, it is proposed that L. catenaformis and L. vitulinus be reclassified in two new genera, named respectively Eggerthia gen. nov., with the type species Eggerthia catenaformis gen. nov., comb. nov. (type strain DSM 20559(T) = ATCC 25536(T) = CCUG 48174(T) = CIP 104817(T) = JCM 1121(T)) and Kandleria gen. nov., with the type species Kandleria vitulina gen. nov., comb. nov. (type strain LMG 18931(T) = ATCC 27783(T) = CCUG 32236(T) = DSM 20405(T) = JCM 1143(T)).

  1. Presence of env-like sequences in Quercus suber retrotransposons.

    PubMed

    Carvalho, M; Ribeiro, T; Viegas, W; Morais-Cecilio, L; Rocheta, M

    2010-01-01

    The main difference between LTR retrotransposons and retroviruses is the presence of the envelope (env) gene in the latter, downstream of the pol gene. The env gene is involved in their infectious capacity. Here we report the presence of env-like sequences in the genome of Quercus suber (cork oak), one of the most economically important Portuguese species. These gene sequences were isolated through DNA amplification between RNaseH conserved motifs and 3' LTR, based on the structure of copia retrotransposons. Phylogenetic analysis revealed that almost all the clones isolated are clustered with Cyclops-2, a Ty3-gypsy element identified in Pisum sativum, except one clustered with gypsy and copia retroelements found in different species. This suggests the existence of a potential ancestral sequence of the env gene, prior to the separation of Ty3-gypsy and Ty1-copia retrotransposons. Additionally, the isolated env-like sequences showed 26-39% of homology with env-like sequences characterized in viruses. The origin of env-like sequences in retrotransposons from host plant taxa is discussed.

  2. The family Coriobacteriaceae: reclassification of Eubacterium exiguum (Poco et al. 1996) and Peptostreptococcus heliotrinreducens (Lanigan 1976) as Slackia exigua gen. nov., comb. nov. and Slackia heliotrinireducens gen. nov., comb. nov., and Eubacterium lentum (Prevot 1938) as Eggerthella lenta gen. nov., comb. nov.

    PubMed

    Wade, W G; Downes, J; Dymock, D; Hiom, S J; Weightman, A J; Dewhirst, F E; Paster, B J; Tzellas, N; Coleman, B

    1999-04-01

    16S rRNA gene sequences were determined for Eubacterium exiguum and Peptostreptococcus heliotrinreducens. These species were found to be closely related and, together with Eubacterium lentum, to constitute a branch of the Coriobacteriaceae. Two new genera are proposed on the basis of phenotypic characteristics and 16S rRNA gene sequence comparisons: Slackia to include the bile-sensitive species Eubacterium exiguum and P. heliotrinreducens, and Eggerthella to include the bile-resistant Eubacterium lentum. It is proposed that Eubacterium exiguum and Peptostreptococcus heliotrinreducens are transferred to the genus Slackia gen. nov. as Slackia exigua gen. nov., comb. nov. (type strain ATCC 700122T) and Slackia heliotrinireducens gen. nov., comb. nov. (type strain NTCC 11029T), respectively, and Eubacterium lentum is transferred to the genus Eggerthella gen. nov. as Eggerthella lenta gen. nov., comb. nov. with Eggerthella lenta as the type species.

  3. Faecalicoccus acidiformans gen. nov., sp. nov., isolated from the chicken caecum, and reclassification of Streptococcus pleomorphus (Barnes et al. 1977), Eubacterium biforme (Eggerth 1935) and Eubacterium cylindroides (Cato et al. 1974) as Faecalicoccus pleomorphus comb. nov., Holdemanella biformis gen. nov., comb. nov. and Faecalitalea cylindroides gen. nov., comb. nov., respectively, within the family Erysipelotrichaceae.

    PubMed

    De Maesschalck, Celine; Van Immerseel, Filip; Eeckhaut, Venessa; De Baere, Siegrid; Cnockaert, Margo; Croubels, Siska; Haesebrouck, Freddy; Ducatelle, Richard; Vandamme, Peter

    2014-11-01

    Strains LMG 27428(T) and LMG 27427 were isolated from the caecal content of a chicken and produced butyric, lactic and formic acids as major metabolic end products. The genomic DNA G+C contents of strains LMG 27428(T) and LMG 27427 were 40.4 and 38.8 mol%. On the basis of 16S rRNA gene sequence similarity, both strains were most closely related to the generically misclassified Streptococcus pleomorphus ATCC 29734(T). Strain LMG 27428(T) could be distinguished from S. pleomorphus ATCC 29734(T) based on production of more lactic acid and less formic acid in M2GSC medium, a higher DNA G+C content and the absence of activities of acid phosphatase and leucine, arginine, leucyl glycine, pyroglutamic acid, glycine and histidine arylamidases, while strain LMG 27428 was biochemically indistinguishable from S. pleomorphus ATCC 29734(T). The novel genus Faecalicoccus gen. nov. within the family Erysipelotrichaceae is proposed to accommodate strains LMG 27428(T) and LMG 27427. Strain LMG 27428(T) ( =DSM 26963(T)) is the type strain of Faecalicoccus acidiformans sp. nov., and strain LMG 27427 ( =DSM 26962) is a strain of Faecalicoccus pleomorphus comb. nov. (type strain LMG 17756(T) =ATCC 29734(T) =DSM 20574(T)). Furthermore, the nearest phylogenetic neighbours of the genus Faecalicoccus are the generically misclassified Eubacterium cylindroides DSM 3983(T) (94.4% 16S rRNA gene sequence similarity to strain LMG 27428(T)) and Eubacterium biforme DSM 3989(T) (92.7% 16S rRNA gene sequence similarity to strain LMG 27428(T)). We present genotypic and phenotypic data that allow the differentiation of each of these taxa and propose to reclassify these generically misnamed species of the genus Eubacterium formally as Faecalitalea cylindroides gen. nov., comb. nov. and Holdemanella biformis gen. nov., comb. nov., respectively. The type strain of Faecalitalea cylindroides is DSM 3983(T) =ATCC 27803(T) =JCM 10261(T) and that of Holdemanella biformis is DSM 3989(T

  4. RRE-dependent HIV-1 Env RNA effects on Gag protein expression, assembly and release

    SciTech Connect

    López, Claudia S.; Sloan, Rachel; Cylinder, Isabel; Kozak, Susan L.; Kabat, David; Barklis, Eric

    2014-08-15

    The HIV-1 Gag proteins are translated from the full-length HIV-1 viral RNA (vRNA), whereas the envelope (Env) protein is translated from incompletely spliced Env mRNAs. Nuclear export of vRNAs and Env mRNAs is mediated by the Rev accessory protein which binds to the rev-responsive element (RRE) present on these RNAs. Evidence has shown there is a direct or indirect interaction between the Gag protein, and the cytoplasmic tail (CT) of the Env protein. Our current work shows that env gene expression impacts HIV-1 Gag expression and function in two ways. At the protein level, full-length Env expression altered Gag protein expression, while Env CT-deletion proteins did not. At the RNA level, RRE-containing Env mRNA expression reduced Gag expression, processing, and virus particle release from cells. Our results support models in which Gag is influenced by the Env CT, and Env mRNAs compete with vRNAs for nuclear export. - Highlights: • At the protein level, full-length HIV-1 Env alters Gag protein expression. • HIV-1 Env RNA expression reduces Gag levels and virus release. • Env RNA effects on Gag are dependent on the RRE. • RRE-containing Env RNAs compete with vRNAs for nuclear export.

  5. An amphipathic sequence in the cytoplasmic tail of HIV-1 Env alters cell tropism and modulates viral receptor specificity.

    PubMed

    Vzorov, A N; Yang, C; Compans, R W

    2015-09-01

    The human immunodeficiency virus type 1 (HIV-1) 92UG046 Env protein, obtained from a CD4-independent HIV-1 primary isolate (Zerhouni et al., 2004), has the ability to initiate an infection in HeLa cells expressing CD4 when carrying the full-length (FL) Env, but uses CD8 molecules for receptor-mediated entry when carrying a truncated Env (CT84). To determine whether a specific length or structure in the cytoplasmic tail (CT) is responsible for this alteration of tropism, we compared a series of Env constructs with different CT truncations and the presence or absence of an amphipathic alpha- helical sequence. We found that truncated constructs containing the alpha-helical LLP-2 structure in their CT domains conferred a switch from CD4 to CD8 tropism. The results support the conclusion that the structure of the CT domain can play an important role in determining receptor specificity.

  6. Evolution of B cell analysis and Env trimer redesign.

    PubMed

    Karlsson Hedestam, Gunilla B; Guenaga, Javier; Corcoran, Martin; Wyatt, Richard T

    2017-01-01

    HIV-1 and its surface envelope glycoproteins (Env), gp120 and gp41, have evolved immune evasion strategies that render the elicitation of effective antibody responses to the functional Env entry unit extremely difficult. HIV-1 establishes chronic infection and stimulates vigorous immune responses in the human host; forcing selection of viral variants that escape cellular and antibody (Ab)-mediated immune pressure, yet possess contemporary fitness. Successful survival of fit variants through the gauntlet of the human immune system make this virus and these glycoproteins a formidable challenge to target by vaccination, requiring a systematic approach to Env mimetic immunogen design and evaluation of elicited responses. Here, we review key aspects of HIV-1 Env immunogenicity and immunogen re-design, based on experimental data generated by us and others over the past decade or more. We further provide rationale and details regarding the use of newly evolving tools to analyze B cell responses, including approaches to use next generation sequencing for antibody lineage tracing and B cell fate mapping. Together, these developments offer opportunities to address long-standing questions about the establishment of effective B cell immunity elicited by vaccination, not just against HIV-1. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  7. Appreciating HIV-1 diversity: subtypic differences in ENV

    SciTech Connect

    Gnanakaran, S; Shen, Tongye; Lynch, Rebecca M; Derdeyn, Cynthia A

    2008-01-01

    Human immunodeficiency virus type 1 (HIV-1) group M is responsible for the current AIDS pandemic and exhibits exceedingly high levels of viral genetic diversity around the world, necessitating categorization of viruses into distinct lineages, or subtypes. These subtypes can differ by around 35% in the envelope (Env) glycoproteins of the virus, which are displayed on the surface of the virion and are targets for both neutralizing antibody and cell-mediated immune responses. This diversity reflects the remarkable ability of the virus to adapt to selective pressures, the bulk of which is applied by the host immune response, and represents a serious obstacle for developing an effective vaccine with broad coverage. Thus, it is important to understand the underlying biological consequences of inter-subtype diversity. Recent studies have revealed that the HIV-1 subtypes exhibit phenotypic differences that result from subtle differences in Env structure, particularly within the highly immunogenic V3 domain, which participates directly in viral entry. This review will therefore explore current research that describes subtypic differences in Env at the genetic and phenotypic level, focusing in particular on V3, and highlighting recent discoveries about the unique features of subtype C Env, which is the most prevalent subtype globally.

  8. A Highly Conserved gp120 Inner Domain Residue Modulates Env Conformation and Trimer Stability

    PubMed Central

    Ding, Shilei; Tolbert, William D.; Prévost, Jérémie; Pacheco, Beatriz; Coutu, Mathieu; Debbeche, Olfa; Xiang, Shi-Hua

    2016-01-01

    ABSTRACT Previous studies have shown that highly conserved residues in the inner domain of gp120 are required for HIV-1 envelope glycoprotein (Env) transitions to the CD4-bound conformation (A. Finzi, S. H. Xiang, B. Pacheco, L. Wang, J. Haight, et al., Mol Cell 37:656–667, 2010, http://dx.doi.org/10.1016/j.molcel.2010.02.012; A. Desormeaux, M. Coutu, H. Medjahed, B. Pacheco, A. Herschhorn, et al., J Virol 87:2549–2562, 2013, http://dx.doi.org/10.1128/JVI.03104-12). Moreover, W69, a highly conserved residue located at the interface between layer 1 and layer 2 of the inner domain, was recently shown to be important for efficient Env recognition by CD4-induced (CD4i) antibodies capable of potent antibody-dependent cellular cytotoxicity (W. D. Tolbert, N. Gohain, M. Veillette, J. P. Chapleau, C. Orlandi, et al., 2016, Structure 24:697–709, http://dx.doi.org/10.1016/j.str.2016.03.005; S. Ding, M. Veillette, M. Coutu, J. Prevost, L. Scharf, et al., 2016, J Virol 90:2127–2134, http://dx.doi.org/10.1128/JVI.02779-15). We evaluated the contribution of the hydrophobicity of W69 to conformational changes of Env by replacing it with a series of residues with aliphatic or aromatic side chains of decreasing chain length. We have found that the hydrophobicity of residue 69 is important for Env processing, CD4 binding, and its transition to the CD4-bound conformation. The most deleterious effect was observed when W69 was replaced with alanine or glycine residues. However, the functions lost due to W69 mutations could be progressively restored with amino acids of increasing aliphatic chain length and fully recovered with residues bearing an aromatic ring. Interestingly, poor CD4 binding of W69A could be fully restored by introducing a compensatory mutation within layer 2 (S115W). Structural studies of HIV-1 gp120 coree W69A/S115W mutant bound to the CD4 peptide mimetic M48U1 and Fab of anti-cluster A antibody N60-i3 revealed no perturbations to the overall structure of the

  9. Feline immunodeficiency virus env gene evolution in experimentally infected cats.

    PubMed

    Kraase, Martin; Sloan, Richard; Klein, Dieter; Logan, Nicola; McMonagle, Linda; Biek, Roman; Willett, Brian J; Hosie, Margaret J

    2010-03-15

    Feline immunodeficiency virus (FIV), an immunosuppressive lentivirus found in cats worldwide, is studied to illuminate mechanisms of lentiviral pathogenesis and to identify key components of protective immunity. During replication, lentiviruses accumulate errors of nucleotide mis-incorporation due to the low-fidelity of reverse transcriptase and recombination between viral variants, resulting in the emergence of a complex viral "quasispecies". In patients infected with HIV-1, env sequences may vary by up to 10% and the detection of quasispecies with greater heterogeneity is associated with higher viral loads and reduced CD4+ T cell numbers [1], indicating that transmission of more complex quasispecies may lead to disease progression. However, little is known about how FIV evolves as disease progresses, or why some cats develop AIDS rapidly while disease progression is slow in others. The aim of this study was to determine whether disease progression may be governed by viral evolution and to examine the diversity of viral variants emerging following infection with an infectious molecular clone. The FIV env gene encoding the envelope glycoprotein (Env) was examined at early (12 weeks) and late (322 weeks) stages of FIV infection in two groups of cats infected experimentally with the FIV-GL8 molecular clone. Viral variants were detected within quasispecies in cats in the late stages of FIV infection that contained differing amino acid compositions in several variable loops of Env, some of which were identified as determinants of receptor usage and resistance to neutralization. Therefore these results indicate that the FIV env gene evolves during the course of infection, giving rise to variants that resist neutralization and likely lead to disease progression.

  10. Panels of HIV-1 Subtype C Env Reference Strains for Standardized Neutralization Assessments.

    PubMed

    Hraber, Peter; Rademeyer, Cecilia; Williamson, Carolyn; Seaman, Michael S; Gottardo, Raphael; Tang, Haili; Greene, Kelli; Gao, Hongmei; LaBranche, Celia; Mascola, John R; Morris, Lynn; Montefiori, David C; Korber, Bette

    2017-10-01

    half of current HIV-1 infections, and is a focus for vaccine design and testing. Standard reference reagents, particularly virus panels to study neutralization by antibodies, are crucial for developing cost-effective and yet rigorous and reproducible assays against diverse examples of this variable virus. We developed clade C-specific panels for use as standardized reagents to monitor complex polyclonal sera for neutralization activity and to characterize the potency and breadth of cross-reactive neutralization by monoclonal antibodies, whether engineered or isolated from infected individuals. We chose from 200 southern African, clade C envelope-pseudotyped viruses with neutralization titers against 16 broadly neutralizing antibodies and 30 sera from chronic clade C infections. We selected panels to represent the diversity of bnAb neutralization profiles and Env neutralization sensitivities. Use of standard virus panels can facilitate comparison of results across studies and sites. Copyright © 2017 Hraber et al.

  11. Glycosylation Benchmark Profile for HIV-1 Envelope Glycoprotein Production Based on Eleven Env Trimers.

    PubMed

    Go, Eden P; Ding, Haitao; Zhang, Shijian; Ringe, Rajesh P; Nicely, Nathan; Hua, David; Steinbock, Robert T; Golabek, Michael; Alin, James; Alam, S Munir; Cupo, Albert; Haynes, Barton F; Kappes, John C; Moore, John P; Sodroski, Joseph G; Desaire, Heather

    2017-05-01

    HIV-1 envelope glycoprotein (Env) glycosylation is important because individual glycans are components of multiple broadly neutralizing antibody epitopes, while shielding other sites that might otherwise be immunogenic. The glycosylation on Env is influenced by a variety of factors, including the genotype of the protein, the cell line used for its expression, and the details of the construct design. Here, we used a mass spectrometry (MS)-based approach to map the complete glycosylation profile at every site in multiple HIV-1 Env trimers, accomplishing two goals. (i) We determined which glycosylation sites contain conserved glycan profiles across many trimeric Envs. (ii) We identified the variables that impact Env's glycosylation profile at sites with divergent glycosylation. Over half of the gp120 glycosylation sites on 11 different trimeric Envs have a conserved glycan profile, indicating that a native consensus glycosylation profile does indeed exist among trimers. We showed that some soluble gp120s and gp140s exhibit highly divergent glycosylation profiles compared to trimeric Env. We also assessed the impact of several variables on Env glycosylation: truncating the full-length Env; producing Env, instead of the more virologically relevant T lymphocytes, in CHO cells; and purifying Env with different chromatographic platforms, including nickel-nitrilotriacetic acid (Ni-NTA), 2G12, and PGT151 affinity. This report provides the first consensus glycosylation profile of Env trimers, which should serve as a useful benchmark for HIV-1 vaccine developers. This report also defines the sites where glycosylation may be impacted when Env trimers are truncated or produced in CHO cells.IMPORTANCE A protective HIV-1 vaccine will likely include a recombinant version of the viral envelope glycoprotein (Env). Env is highly glycosylated, and yet vaccine developers have lacked guidance on how to assess whether their immunogens have optimal glycosylation. The following important

  12. Structure and immune recognition of trimeric prefusion HIV-1 Env

    PubMed Central

    Pancera, Marie; Zhou, Tongqing; Druz, Aliaksandr; Georgiev, Ivelin S.; Soto, Cinque; Gorman, Jason; Huang, Jinghe; Acharya, Priyamvada; Chuang, Gwo-Yu; Ofek, Gilad; Stewart-Jones, Guillaume B. E.; Stuckey, Jonathan; Bailer, Robert T.; Joyce, M. Gordon; Louder, Mark K.; Tumba, Nancy; Yang, Yongping; Zhang, Baoshan; Cohen, Myron S.; Haynes, Barton F.; Mascola, John R.; Morris, Lynn; Munro, James B.; Blanchard, Scott C.; Mothes, Walther; Connors, Mark; Kwong, Peter D.

    2015-01-01

    The HIV-1-envelope (Env) spike, comprising three gp120 and three gp41 subunits, is a conformational machine that facilitates HIV-1 entry by rearranging from a mature unliganded state, through receptor-bound intermediates, to a postfusion state. As the sole viral antigen on the HIV-1-virion surface, Env is both the target of neutralizing antibodies and a focus of vaccine efforts. Here we report the structure at 3.5-Å resolution for an HIV-1-Env trimer captured in a mature closed state by antibodies PGT122 and 35O22. This structure reveals the prefusion conformation of gp41, indicates rearrangements needed for fusion activation, and defines parameters of immune evasion and immune recognition. Prefusion gp41 encircles N- and C-terminal strands of gp120 with four helices that form a membrane-proximal collar, fastened by insertion of a fusion peptide-proximal methionine into a gp41-tryptophan clasp. Spike rearrangements required for entry likely involve opening the clasp and expelling the termini. N-linked glycosylation and sequence-variable regions cover the prefusion closed spike: we used chronic cohorts to map the prevalence and location of effective HIV-1-neutralizing responses, which were distinguished by their recognition of N-linked glycan and tolerance for epitope-sequence variation. PMID:25296255

  13. Wautersia gen. nov., a novel genus accommodating the phylogenetic lineage including Ralstonia eutropha and related species, and proposal of Ralstonia [Pseudomonas] syzygii (Roberts et al. 1990) comb. nov.

    PubMed

    Vaneechoutte, Mario; Kämpfer, Peter; De Baere, Thierry; Falsen, Enevold; Verschraegen, Gerda

    2004-03-01

    Comparative 16S rDNA sequence analysis indicates that two distinct sublineages, with a sequence dissimilarity of >4 % (bootstrap value, 100 %), exist within the genus RALSTONIA: the Ralstonia eutropha lineage, which comprises Ralstonia basilensis, Ralstonia campinensis, R. eutropha, Ralstonia gilardii, Ralstonia metallidurans, Ralstonia oxalatica, Ralstonia paucula, Ralstonia respiraculi and Ralstonia taiwanensis; and the Ralstonia pickettii lineage, which comprises Ralstonia insidiosa, Ralstonia mannitolilytica, R. pickettii, Ralstonia solanacearum and Ralstonia syzygii comb. nov. (previously Pseudomonas syzygii). This phylogenetic discrimination is supported by phenotypic differences. Members of the R. eutropha lineage have peritrichous flagella, do not produce acids from glucose and are susceptible to colistin, in contrast to members of the R. pickettii lineage, which have one or more polar flagella, produce acid from several carbohydrates and are colistin-resistant. Members of the R. pickettii lineage are viable for up to 6 days on tryptic soy agar at 25 degrees C, whereas members of the R. eutropha lineage are viable for longer than 9 days. It is proposed that species of the R. eutropha lineage should be classified in a novel genus, Wautersia gen. nov. Finally, based on the literature and new DNA-DNA hybridization data, it is proposed that Pseudomonas syzygii should be renamed Ralstonia syzygii comb. nov.

  14. Proposal to unify Clostridium orbiscindens Winter et al. 1991 and Eubacterium plautii (Séguin 1928) Hofstad and Aasjord 1982, with description of Flavonifractor plautii gen. nov., comb. nov., and reassignment of Bacteroides capillosus to Pseudoflavonifractor capillosus gen. nov., comb. nov.

    PubMed

    Carlier, Jean-Philippe; Bedora-Faure, Marie; K'ouas, Guylène; Alauzet, Corentine; Mory, Francine

    2010-03-01

    We isolated several strains from various clinical samples (five samples of blood, four of intra-abdominal pus and one of infected soft tissue) that were anaerobic, motile or non-motile and Gram-positive rods. Some of the strains formed spores. Phylogenetic analysis of the 16S rRNA gene sequence showed that these organisms could be placed within clostridial cluster IV as defined by Collins et al. [(1994). Int J Syst Bacteriol 44, 812-826] and shared more than 99 % sequence similarity with Clostridium orbiscindens DSM 6740(T) and Eubacterium plautii DSM 4000(T). Together, they formed a distinct cluster, with Bacteroides capillosus ATCC 29799(T) branching off from this line of descent with sequence similarities of 97.1-97.4 %. The next nearest neighbours of these organisms were Clostridium viride, Oscillibacter valericigenes, Papillibacter cinnamivorans and Sporobacter termitidis, with sequence similarities to the respective type strains of 93.1-93.4, 91.2-91.4, 89.8-90 and 88.7-89.3 %. On the basis of biochemical properties, phylogenetic position, DNA G+C content and DNA-DNA hybridization, it is proposed to unify Clostridium orbiscindens and Eubacterium plautii in a new genus as Flavonifractor plautii gen. nov., comb. nov., with the type strain Prévot S1(T) (=ATCC 29863(T) =VPI 0310(T) =DSM 4000(T)), and to reassign Bacteroides capillosus to Pseudoflavonifractor capillosus gen. nov., comb. nov., with the type strain CCUG 15402A(T) (=ATCC 29799(T) =VPI R2-29-1(T)).

  15. Splicing of Friend Murine Leukemia Virus env-mRNA Enhances Its Ability to Form Polysomes

    PubMed Central

    Machinaga, Akihito; Ishihara, Syuhei; Shirai, Akiko; Takase-Yoden, Sayaka

    2016-01-01

    Friend murine leukemia virus (MLV) belongs to the gamma retroviruses of the Retroviridae family. The positive-sense RNA of its genome contains a 5′ long terminal repeat (LTR), 5′ leader sequence, gag, pol, env, and 3′ LTR. Transcription from proviral DNA begins from the R region of the 5′ LTR and ends at the polyadenylation signal located at the R region of the other end of the 3′ LTR. There is a 5′ splice site in the 5′ leader sequence and a 3′ splice site at the 3′ end of the pol region. Both full-length unspliced mRNAs and a singly spliced mRNA (env-mRNA) are produced in MLV-infected cells. The MLV Env protein plays important roles both in viral adsorption to host cells and in neuropathogenic disease in MLV-infected mice and rats. Understanding the regulatory mechanisms controlling Env expression is important for determining the functions of the Env protein. We have previously shown that splicing increases env-mRNA stability and translation efficiency. Generally, mRNA polysome formation correlates with translation efficiency. Therefore, here we investigated the effects of env-mRNA splicing on polysome formation to identify mechanisms for Env up-regulation due to splicing. We performed polysome profile analyses using Env-expression plasmids producing spliced or unspliced env-mRNA and showed that the former formed polysomes more efficiently than the latter. Thus, splicing of env-mRNA facilitated polysome formation, suggesting that this contributes to up-regulation of Env expression. We replaced the env region of the expression plasmids with a luciferase (luc) gene, and found that in this case both unspliced and spliced luc-mRNA formed polysomes to a similar extent. Thus, we conclude that whether mRNA polysome formation is affected by splicing depends on the structure of gene in question. PMID:26909075

  16. Telmatobacter bradus gen. nov., sp. nov., a cellulolytic facultative anaerobe from subdivision 1 of the Acidobacteria, and emended description of Acidobacterium capsulatum Kishimoto et al. 1991.

    PubMed

    Pankratov, Timofey A; Kirsanova, Lilia A; Kaparullina, Elena N; Kevbrin, Vadim V; Dedysh, Svetlana N

    2012-02-01

    A gram-negative, facultatively anaerobic, chemo-organotrophic, non-pigmented, slow-growing bacterium was isolated from acidic Sphagnum peat and designated strain TPB6017(T). Cells of this strain were long rods that multiplied by normal cell division and were motile by means of a single flagellum. Cells grew under reduced oxygen tension and under anoxic conditions and were able to ferment sugars and several polysaccharides, including amorphous and crystalline cellulose. Strain TPB6017(T) was a psychrotolerant acidophile capable of growth between pH 3.0 and 7.5 (optimum 4.5-5.0) and at 4-35 °C (optimum 20-28 °C). It was extremely sensitive to salt stress; growth was inhibited at NaCl concentrations above 0.1 % (w/v). The major fatty acids were iso-C(15 : 0) and iso-C(17 : 1)ω9c; the polar lipids were phosphatidylethanolamine and a number of phospholipids and aminophospholipids with an unknown structure. The quinone was MK-8. The DNA G+C content was 57.6 mol%. Comparative 16S rRNA gene sequence analysis revealed that strain TPB6017(T) was a member of subdivision 1 of the phylum Acidobacteria and belonged to a phylogenetic lineage defined by the acidophilic aerobic chemo-organotroph Acidobacterium capsulatum (92.3 % sequence similarity). However, cell morphology, type of flagellation, the absence of pigment, differences in fatty acid and polar lipid composition, possession of a cellulolytic capability, inability to grow under fully oxic conditions and good growth in anoxic conditions distinguished strain TPB6017(T) from A. capsulatum. Therefore, it is proposed that strain TPB6017(T) represents a novel acidobacterium species in a new genus, Telmatobacter bradus gen. nov., sp. nov.; strain TPB6017(T) ( = DSM 23630(T) = VKM B-2570(T)) is the type strain.

  17. Proposal for creation of a new genus Neomicrococcus gen. nov. to accommodate Zhihengliuella aestuarii Baik et al. 2011 and Micrococcus lactis Chittpurna et al. 2011 as Neomicrococcus aestuarii comb. nov. and Neomicrococcus lactis comb. nov.

    PubMed

    Prakash, Om; Sharma, Avinash; Nimonkar, Yogesh; Shouche, Yogesh S

    2015-11-01

    Micrococcus lactis and Zhihengliuella aestuarii were described independently in 2011. Their type strains showed high levels of 16S rRNA gene sequence similarity (99.3%). Phylogenetic analysis revealed that M. lactis MCC 2278T and Z. aestuarii JCM 16166T formed a monophyletic group and showed distant relationships to other members of closely related genera such as Micrococcus, Zhihengliuella, Arthrobacter and Citricoccus. The presence of large proportions of iso-C14:0 and iso-C16:0 with small amounts of iso-C15:0 distinguished M. lactis MCC 2278T and Z. aestuarii JCM 16166T from other members of the genera Micrococcus and Zhihengliuella. Unlike other members of the genera Zhihengliuella and Micrococcus, M. lactis MCC 2278T and Z. aestuarii JCM 16166T showed growth at low concentrations of NaCl. Thus, based on distinctive phylogenetic, chemotaxonomic and physiological features of these two organisms in comparison with other members of the genera Micrococcus and Zhihengliuella, it is clear that they do not fit within the existing classification and deserve separate status. DNA-DNA hybridization between the two type strains was 63%, indicating that they represent separate species. In this study, we propose the creation of a novel genus, Neomicrococcus gen. nov., to accommodate the two species with Neomicrococcus aestuarii gen. nov., comb. nov. (type strain JCM 16166T = KCTC 19557T) as the type species. Neomicrococcus lactis comb. nov. (type strain MCC 2278T = DSM 23694T) is also proposed.

  18. [Al

    PubMed

    Purath; Köppe; Schnöckel

    1999-10-04

    A "naked" aluminum atom links two aluminum tetrahedra in the [Al(7){N(SiMe(3))(2)}(6)](-) ion (see picture), which results from the reaction of a metastable AlCl solution with LiN(SiMe(3))(2) and crystallizes with [Li(OEt(2))(3)](+) as cation. This unique structure among molecular metal atom clusters represents a small but characteristic section of cubic close-packed aluminum.

  19. Identification of the major capsid protein of erythrocytic necrosis virus (ENV) and development of quantitative real-time PCR assays for quantification of ENV DNA.

    PubMed

    Purcell, Maureen K; Pearman-Gillman, Schuyler; Thompson, Rachel L; Gregg, Jacob L; Hart, Lucas M; Winton, James R; Emmenegger, Eveline J; Hershberger, Paul K

    2016-07-01

    Viral erythrocytic necrosis (VEN) is a disease of marine and anadromous fish that is caused by the erythrocytic necrosis virus (ENV), which was recently identified as a novel member of family Iridoviridae by next-generation sequencing. Phylogenetic analysis of the ENV DNA polymerase grouped ENV with other erythrocytic iridoviruses from snakes and lizards. In the present study, we identified the gene encoding the ENV major capsid protein (MCP) and developed a quantitative real-time PCR (qPCR) assay targeting this gene. Phylogenetic analysis of the MCP gene sequence supported the conclusion that ENV does not group with any of the currently described iridovirus genera. Because there is no information regarding genetic variation of the MCP gene across the reported host and geographic range for ENV, we also developed a second qPCR assay for a more conserved ATPase-like gene region. The MCP and ATPase qPCR assays demonstrated good analytical and diagnostic sensitivity and specificity based on samples from laboratory challenges of Pacific herring Clupea pallasii The qPCR assays had similar diagnostic sensitivity and specificity as light microscopy of stained blood smears for the presence of intraerythrocytic inclusion bodies. However, the qPCR assays may detect viral DNA early in infection prior to the formation of inclusion bodies. Both qPCR assays appear suitable for viral surveillance or as a confirmatory test for ENV in Pacific herring from the Salish Sea.

  20. Identification of the major capsid protein of erythrocytic necrosis virus (ENV) and development of quantitative real-time PCR assays for quantification of ENV DNA

    USGS Publications Warehouse

    Purcell, Maureen K.; Pearman-Gillman, Schuyler; Thompson, Rachel L.; Gregg, Jacob L.; Hart, Lucas M.; Winton, James R.; Emmenegger, Eveline J.; Hershberger, Paul K.

    2016-01-01

    Viral erythrocytic necrosis (VEN) is a disease of marine and anadromous fish that is caused by the erythrocytic necrosis virus (ENV), which was recently identified as a novel member of family Iridoviridae by next-generation sequencing. Phylogenetic analysis of the ENV DNA polymerase grouped ENV with other erythrocytic iridoviruses from snakes and lizards. In the present study, we identified the gene encoding the ENV major capsid protein (MCP) and developed a quantitative real-time PCR (qPCR) assay targeting this gene. Phylogenetic analysis of the MCP gene sequence supported the conclusion that ENV does not group with any of the currently described iridovirus genera. Because there is no information regarding genetic variation of the MCP gene across the reported host and geographic range for ENV, we also developed a second qPCR assay for a more conserved ATPase-like gene region. The MCP and ATPase qPCR assays demonstrated good analytical and diagnostic sensitivity and specificity based on samples from laboratory challenges of Pacific herring Clupea pallasii. The qPCR assays had similar diagnostic sensitivity and specificity as light microscopy of stained blood smears for the presence of intraerythrocytic inclusion bodies. However, the qPCR assays may detect viral DNA early in infection prior to the formation of inclusion bodies. Both qPCR assays appear suitable for viral surveillance or as a confirmatory test for ENV in Pacific herring from the Salish Sea.

  1. Transcriptional and functional studies of Human Endogenous Retrovirus envelope EnvP(b) and EnvV genes in human trophoblasts

    SciTech Connect

    Vargas, Amandine Thiery, Maxime Lafond, Julie Barbeau, Benoit

    2012-03-30

    HERV (Human Endogenous Retrovirus)-encoded envelope proteins are implicated in the development of the placenta. Indeed, Syncytin-1 and -2 play a crucial role in the fusion of human trophoblasts, a key step in placentation. Other studies have identified two other HERV env proteins, namely EnvP(b) and EnvV, both expressed in the placenta. In this study, we have fully characterized both env transcripts and their expression pattern and have assessed their implication in trophoblast fusion. Through RACE analyses, standard spliced transcripts were detected, while EnvV transcripts demonstrated alternative splicing at its 3 Prime end. Promoter activity and expression of both genes were induced in forskolin-stimulated BeWo cells and in primary trophoblasts. Although we have confirmed the fusogenic activity of EnvP(b), overexpression or silencing experiments revealed no impact of this protein on trophoblast fusion. Our results demonstrate that both env genes are expressed in human trophoblasts but are not required for syncytialization.

  2. SimEnvVis: A Climate Data Visualization Wizard

    NASA Astrophysics Data System (ADS)

    Heitzler, Magnus; Nocke, Thomas

    2013-04-01

    To efficiently make sense of complex climate data, climate scientists need to choose and utilize appropriate analysis tools in respect to specific sets of tasks. Among these, visual analysis tools, like those originating from the field of visual analytics, efficiently support to communicate such information by directly addressing human visual perception. However, climate scientists often are not aware of or not familiar with the large variety of available visual analysis tools or are underestimating their potential benefit for common research tasks and thus reducing the probability to use most suitable ones and therefore impairing the knowledge discovery process. To address this problem, SimEnvVis was developed as an easy-to-use wizard-based software system guiding the user step-by-step in choosing most appropriate visualization and visual analytics tools from a large and easily extendable repository consisting of script-based and interactive tools with different application foci (spatial, temporal or abstract data) and supported techniques (e.g. glyphs, isocontours, stream visualization). Considering the analysis context (e.g. data characteristics, user preferences and analysis tasks) SimEnvVis automatically evaluates the attached tools using a combination of a vector-based and a rule-based mechanism. Based on the users decision, the selected visual analysis tool is launched using a template which is dynamically parameterized by taking into account the analysis context. By displaying the session history in different modes as well as providing the possibility to start SimEnvVis in first-time-user mode to reduce GUI complexity and hide tools which are under development the wizard is in particular useful for novice users. This way, SimEnvVis increases the probability for the usage of appropriate visual analysis tools, lowers the obstacles of familiarization with them and therefore accelerates the knowledge discovery process as well as positively contributes to the

  3. Broadly Neutralizing Antibody 8ANC195 Recognizes Closed and Open States of HIV-1 Env.

    PubMed

    Scharf, Louise; Wang, Haoqing; Gao, Han; Chen, Songye; McDowall, Alasdair W; Bjorkman, Pamela J

    2015-09-10

    The HIV-1 envelope (Env) spike contains limited epitopes for broadly neutralizing antibodies (bNAbs); thus, most neutralizing antibodies are strain specific. The 8ANC195 epitope, defined by crystal and electron microscopy (EM) structures of bNAb 8ANC195 complexed with monomeric gp120 and trimeric Env, respectively, spans the gp120 and gp41 Env subunits. To investigate 8ANC195's gp41 epitope at higher resolution, we solved a 3.58 Å crystal structure of 8ANC195 complexed with fully glycosylated Env trimer, revealing 8ANC195 insertion into a glycan shield gap to contact gp120 and gp41 glycans and protein residues. To determine whether 8ANC195 recognizes the CD4-bound open Env conformation that leads to co-receptor binding and fusion, one of several known conformations of virion-associated Env, we solved EM structures of an Env/CD4/CD4-induced antibody/8ANC195 complex. 8ANC195 binding partially closed the CD4-bound trimer, confirming structural plasticity of Env by revealing a previously unseen conformation. 8ANC195's ability to bind different Env conformations suggests advantages for potential therapeutic applications.

  4. Description of Anaerobacterium chartisolvens gen. nov., sp. nov., an obligately anaerobic bacterium from Clostridium rRNA cluster III isolated from soil of a Japanese rice field, and reclassification of Bacteroides cellulosolvens Murray et al. 1984 as Pseudobacteroides cellulosolvens gen. nov., comb. nov.

    PubMed

    Horino, Haruka; Fujita, Takashi; Tonouchi, Akio

    2014-04-01

    An obligately anaerobic bacterial strain designated T-1-35(T) was isolated as a dominant cultivable cellulose-degrading bacterium from soil of a Japanese rice field as an anaerobic filter-paper degrader. Cells of strain T-1-35(T) stained Gram-positive and were non-spore-forming rods with rounded ends, 0.8-1.0×3.5-15.0 µm, and motile by means of two to four polar flagella. Cells of strain T-1-35(T) exhibited pleomorphism: in aged cultures (over 90 days of incubation), almost all cells were irregularly shaped. Although no spore formation was observed, cells tolerated high temperatures, up to 90 °C for 10 min. The temperature range for growth was 15-40 °C, with an optimum at 35 °C. The pH range for growth was 5.5-9.0, with an optimum at pH 8.0-8.5 (slightly alkaliphilic). Strain T-1-35(T) fermented some carbohydrates to produce ethanol and lactate as the major products. Major cellular fatty acids were iso-C16 : 0 and iso-C13 : 0 3-OH. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that strain T-1-35(T) belonged to Clostridium rRNA cluster III. The closest relative of strain T-1-35(T) was Bacteroides cellulosolvens WM2(T), with 16S rRNA gene sequence similarity of 93.4 %. Phenotypic, physiological and molecular genetic methods demonstrated that strain T-1-35(T) was distinct from its phylogenetic relatives (members of Clostridium rRNA cluster III) because it predominantly produced ethanol, iso-C13 : 0 3-OH was a major cellular fatty acid and it always exhibited pleomorphism. On the basis of the results of a polyphasic taxonomic study, strain T-1-35(T) is considered to represent a novel genus and species, Anaerobacterium chartisolvens gen. nov., sp. nov. The type strain of Anaerobacterium chartisolvens is T-1-35(T) ( = DSM 27016(T) = NBRC 109520(T)). In addition, from the results of our phylogenetic analysis and its phenotypic features, the species Bacteroides cellulosolvens Murray et al. 1984 is proposed to be reclassified

  5. Phylogenomic analysis of the family Peptostreptococcaceae (Clostridium cluster XI) and proposal for reclassification of Clostridium litorale (Fendrich et al. 1991) and Eubacterium acidaminophilum (Zindel et al. 1989) as Peptoclostridium litorale gen. nov. comb. nov. and Peptoclostridium acidaminophilum comb. nov.

    PubMed Central

    Brover, Vyacheslav; Tolstoy, Igor; Yutin, Natalya

    2016-01-01

    In 1994, analyses of clostridial 16S rRNA gene sequences led to the assignment of 18 species to Clostridium cluster XI, separating them from Clostridium sensu stricto (Clostridium cluster I). Subsequently, most cluster XI species have been assigned to the family Peptostreptococcaceae with some species being reassigned to new genera. However, several misclassified Clostridium species remained, creating a taxonomic conundrum and confusion regarding their status. Here, we have re-examined the phylogeny of cluster XI species by comparing the 16S rRNA gene-based trees with protein- and genome-based trees, where available. The resulting phylogeny of the Peptostreptococcaceae was consistent with the recent proposals on creating seven new genera within this family. This analysis also revealed a tight clustering of Clostridium litorale and Eubacterium acidaminophilum. Based on these data, we propose reassigning these two organisms to the new genus Peptoclostridium as Peptoclostridium litorale gen. nov. comb. nov. (the type species of the genus) and Peptoclostridium acidaminophilum comb. nov., respectively. As correctly noted in the original publications, the genera Acetoanaerobium and Proteocatella also fall within cluster XI, and can be assigned to the Peptostreptococcaceae. Clostridium sticklandii, which falls within radiation of genus Acetoanaerobium, is proposed to be reclassified as Acetoanaerobium sticklandii comb. nov. The remaining misnamed members of the Peptostreptococcaceae, [Clostridium] hiranonis, [Clostridium] paradoxum and [Clostridium] thermoalcaliphilum, still remain to be properly classified. PMID:27902180

  6. Phylogenomic analysis of the family Peptostreptococcaceae (Clostridium cluster XI) and proposal for reclassification of Clostridium litorale (Fendrich et al. 1991) and Eubacterium acidaminophilum (Zindel et al. 1989) as Peptoclostridium litorale gen. nov. comb. nov. and Peptoclostridium acidaminophilum comb. nov.

    PubMed

    Galperin, Michael Y; Brover, Vyacheslav; Tolstoy, Igor; Yutin, Natalya

    2016-12-01

    In 1994, analyses of clostridial 16S rRNA gene sequences led to the assignment of 18 species to Clostridium cluster XI, separating them from Clostridium sensu stricto (Clostridium cluster I). Subsequently, most cluster XI species have been assigned to the family Peptostreptococcaceae with some species being reassigned to new genera. However, several misclassified Clostridium species remained, creating a taxonomic conundrum and confusion regarding their status. Here, we have re-examined the phylogeny of cluster XI species by comparing the 16S rRNA gene-based trees with protein- and genome-based trees, where available. The resulting phylogeny of the Peptostreptococcaceae was consistent with the recent proposals on creating seven new genera within this family. This analysis also revealed a tight clustering of Clostridium litorale and Eubacterium acidaminophilum. Based on these data, we propose reassigning these two organisms to the new genus Peptoclostridium as Peptoclostridium litorale gen. nov. comb. nov. (the type species of the genus) and Peptoclostridium acidaminophilum comb. nov., respectively. As correctly noted in the original publications, the genera Acetoanaerobium and Proteocatella also fall within cluster XI, and can be assigned to the Peptostreptococcaceae. Clostridium sticklandii, which falls within radiation of genus Acetoanaerobium, is proposed to be reclassified as Acetoanaerobium sticklandii comb. nov. The remaining misnamed members of the Peptostreptococcaceae, [Clostridium] hiranonis, [Clostridium] paradoxum and [Clostridium] thermoalcaliphilum, still remain to be properly classified.

  7. NMobTec-EnvEdu: M-Learning System for Environmental Education

    ERIC Educational Resources Information Center

    Cavus, Nadire

    2008-01-01

    This paper introduced the implementation of a New Mobile Technologies and Environmental Education System (NMobTec-EnvEdu) designed for m-learning environments. The NMobTec-EnvEdu system has been developed to provide environmental education in a collaborative framework to undergraduate students through the Internet using mobile phones. The study…

  8. Development and characterization of an equine infectious anemia virus Env-pseudotyped reporter virus.

    PubMed

    Tallmadge, R L; Brindley, M A; Salmans, J; Mealey, R H; Maury, W; Carpenter, S

    2008-07-01

    We developed a replication-defective reporter virus pseudotyped with the envelope glycoprotein of equine infectious anemia virus (EIAV). The in vitro host range and neutralization phenotype of EIAV Env-pseudotyped virus were similar to those of replication-competent virus. An EIAV Env pseudovirus will improve antigenic characterization of viral variants and evaluation of lentivirus vaccines.

  9. Phenethyl Alcohol as a Suppressor of the EnvA Phenotype Associated with the envA Gene in Escherichia coli K-12

    PubMed Central

    Normark, Staffan

    1971-01-01

    In Escherichia coli K-12 the envA gene was previously shown to mediate chain formation and a decreased tolerance to several antibacterial agents. Phenethyl alcohol at low concentrations has now been found to increase the tolerance to actinomycin D, ampicillin, rifampin, and gentian violet in strains containing envA. The increased tolerance to gentian violet was correlated to a decreased uptake of the dye. A phenotype suppression of chain formation and colony morphology in envA mutants was also obtained. Except for an increase in palmitic acid, chemical analysis revealed no differences between an envA and its wild-type strain in the lipopolysaccharide part of the envelope. However, a decrease in the amount of phosphatidylglycerol and a C18: 1 fatty acid was observed in the extractable lipids of a strain containing envA. Growth in the presence of phenethyl alcohol reversed the changes in fatty acid and the phospholipid composition. Phenethyl alcohol was found to cause an immediate but transient inhibition of ribonucleic acid synthesis. It is suggested that this inhibition affects the penetrability barrier of the outer cell envelope layers in strains containing envA. Images PMID:4941568

  10. Nucleotide sequencing of an apparent proviral copy of env mRNA defines determinants of expression of the mouse mammary tumor virus env gene.

    PubMed Central

    Majors, J E; Varmus, H E

    1983-01-01

    To extend our understanding of the organization and expression of the mouse mammary tumor virus genome, we determined the nucleotide sequence of large regions of a cloned mouse mammary tumor virus strain C3H provirus that appears to be a DNA copy of env mRNA. In conjunction with analysis of several additional clones of integrated and unintegrated mouse mammary tumor virus DNAs, we came to the following conclusions: (i) the mRNA for env is generated by splicing mechanisms that recognize conventional eucaryotic signals at donor and acceptor sites with a leader of at least 289 bases in length; (ii) the first of three possible initiation codons for translation of env follows the splice junction by a single nucleotide and produces a signal peptide of 98 amino acids; (iii) the amino terminal sequence of the major virion glycoprotein gp52env is confirmed by nucleotide sequencing and is encoded by a sequence beginning 584 nucleotides from the 5' end of env mRNA; (iv) the final 17 amino acids at the carboxyl terminus of the primary product of env are encoded within the long terminal repeat by the 51 bases at the 5' end of the U3 domain; and (v) bases 2 through 4 at the 5' end of the long terminal repeat constitute an initiation codon that commences an open reading frame capable of directing the synthesis of a 36-kilodalton protein. PMID:6312081

  11. Functional and structural characterization of EnvZ, an osmosensing histidine kinase of E. coli.

    PubMed

    Yoshida, Takeshi; Phadtare, Sangita; Inouye, Masayori

    2007-01-01

    EnvZ is an osmosensing histidine kinase located in the inner membrane, and one of the most extensively studied Escherichia coli histidine kinases. Because of its structural complexity, functional and structural studies have been quite challenging. It is a multidomain transmembrane protein consisting of 450 amino acid residues. In addition, it must form a dimer to function as a histidine kinase like all the other histidine kinases. EnvZ consists of the 115-residue periplasmic domain, two transmembrane domains (TM1 and TM2), and the cytoplasmic domain consisting of the 43-residue linker (HAMP) domain and the 228-residue kinase domain. It has been shown that the kinase domain of EnvZ, responsible for its enzymatic activities, contains all of the conserved regions of histidine kinases such as H, F, N, G1, G2, and G3 boxes. Therefore, the 271-residue cytoplasmic domain of EnvZ (termed EnvZc) has been used as a model system to establish fundamental characteristics of histidine kinases. The DNA fragment encoding EnvZc was cloned in pET vector and EnvZc was expressed and purified. It is highly soluble and retains all the enzymatic activities of EnvZ. We demonstrated that it consists of two functional domains, domain A and domain B. NMR spectroscopic studies of these two domains revealed, for the first time, the structure of a histidine kinase. Domain A is responsible for dimerization of EnvZc forming a four-helical bundle containing two alpha-helical hairpin structures, while domain B is a monomer and has an ATP-binding pocket formed by regions conserved among the histidine kinases. In this chapter, we describe functional and structural studies of EnvZc, which can be applied to characterize other histidine kinases.

  12. Targeting HIV-1 Env gp140 to LOX-1 Elicits Immune Responses in Rhesus Macaques.

    PubMed

    Zurawski, Gerard; Zurawski, Sandra; Flamar, Anne-Laure; Richert, Laura; Wagner, Ralf; Tomaras, Georgia D; Montefiori, David C; Roederer, Mario; Ferrari, Guido; Lacabaratz, Christine; Bonnabau, Henri; Klucar, Peter; Wang, Zhiqing; Foulds, Kathryn E; Kao, Shing-Fen; Yates, Nicole L; LaBranche, Celia; Jacobs, Bertram L; Kibler, Karen; Asbach, Benedikt; Kliche, Alexander; Salazar, Andres; Reed, Steve; Self, Steve; Gottardo, Raphael; Galmin, Lindsey; Weiss, Deborah; Cristillo, Anthony; Thiebaut, Rodolphe; Pantaleo, Giuseppe; Levy, Yves

    2016-01-01

    Improved antigenicity against HIV-1 envelope (Env) protein is needed to elicit vaccine-induced protective immunity in humans. Here we describe the first tests in non-human primates (NHPs) of Env gp140 protein fused to a humanized anti-LOX-1 recombinant antibody for delivering Env directly to LOX-1-bearing antigen presenting cells, especially dendritic cells (DC). LOX-1, or 1ectin-like oxidized low-density lipoprotein (LDL) receptor-1, is expressed on various antigen presenting cells and endothelial cells, and is involved in promoting humoral immune responses. The anti-LOX-1 Env gp140 fusion protein was tested for priming immune responses and boosting responses in animals primed with replication competent NYVAC-KC Env gp140 vaccinia virus. Anti-LOX-1 Env gp140 vaccination elicited robust cellular and humoral responses when used for either priming or boosting immunity. Co-administration with Poly ICLC, a TLR3 agonist, was superior to GLA, a TLR4 agonist. Both CD4+ and CD8+ Env-specific T cell responses were elicited by anti-LOX-1 Env gp140, but in particular the CD4+ T cells were multifunctional and directed to multiple epitopes. Serum IgG and IgA antibody responses induced by anti-LOX-1 Env gp140 against various gp140 domains were cross-reactive across HIV-1 clades; however, the sera neutralized only HIV-1 bearing sequences most similar to the clade C 96ZM651 Env gp140 carried by the anti-LOX-1 vehicle. These data, as well as the safety of this protein vaccine, justify further exploration of this DC-targeting vaccine approach for protective immunity against HIV-1.

  13. Targeting HIV-1 Env gp140 to LOX-1 Elicits Immune Responses in Rhesus Macaques

    PubMed Central

    Zurawski, Sandra; Flamar, Anne-Laure; Richert, Laura; Wagner, Ralf; Tomaras, Georgia D.; Montefiori, David C.; Roederer, Mario; Ferrari, Guido; Lacabaratz, Christine; Bonnabau, Henri; Klucar, Peter; Wang, Zhiqing; Foulds, Kathryn E.; Kao, Shing-Fen; Yates, Nicole L.; LaBranche, Celia; Jacobs, Bertram L.; Kibler, Karen; Asbach, Benedikt; Kliche, Alexander; Salazar, Andres; Reed, Steve; Self, Steve; Gottardo, Raphael; Galmin, Lindsey; Weiss, Deborah; Cristillo, Anthony; Thiebaut, Rodolphe; Pantaleo, Giuseppe; Levy, Yves

    2016-01-01

    Improved antigenicity against HIV-1 envelope (Env) protein is needed to elicit vaccine-induced protective immunity in humans. Here we describe the first tests in non-human primates (NHPs) of Env gp140 protein fused to a humanized anti-LOX-1 recombinant antibody for delivering Env directly to LOX-1-bearing antigen presenting cells, especially dendritic cells (DC). LOX-1, or 1ectin-like oxidized low-density lipoprotein (LDL) receptor-1, is expressed on various antigen presenting cells and endothelial cells, and is involved in promoting humoral immune responses. The anti-LOX-1 Env gp140 fusion protein was tested for priming immune responses and boosting responses in animals primed with replication competent NYVAC-KC Env gp140 vaccinia virus. Anti-LOX-1 Env gp140 vaccination elicited robust cellular and humoral responses when used for either priming or boosting immunity. Co-administration with Poly ICLC, a TLR3 agonist, was superior to GLA, a TLR4 agonist. Both CD4+ and CD8+ Env-specific T cell responses were elicited by anti-LOX-1 Env gp140, but in particular the CD4+ T cells were multifunctional and directed to multiple epitopes. Serum IgG and IgA antibody responses induced by anti-LOX-1 Env gp140 against various gp140 domains were cross-reactive across HIV-1 clades; however, the sera neutralized only HIV-1 bearing sequences most similar to the clade C 96ZM651 Env gp140 carried by the anti-LOX-1 vehicle. These data, as well as the safety of this protein vaccine, justify further exploration of this DC-targeting vaccine approach for protective immunity against HIV-1. PMID:27077384

  14. Pseudotyping incompatibility between HIV-1 and gibbon ape leukemia virus Env is modulated by Vpu.

    PubMed

    Lucas, Tiffany M; Lyddon, Terri D; Cannon, Paula M; Johnson, Marc C

    2010-03-01

    The Env protein from gibbon ape leukemia virus (GaLV) has been shown to be incompatible with human immunodeficiency virus type 1 (HIV-1) in the production of infectious pseudotyped particles. This incompatibility has been mapped to the C-terminal cytoplasmic tail of GaLV Env. Surprisingly, we found that the HIV-1 accessory protein Vpu modulates this incompatibility. The infectivity of HIV-1 pseudotyped with murine leukemia virus (MLV) Env was not affected by Vpu. However, the infectivity of HIV-1 pseudotyped with an MLV Env with the cytoplasmic tail from GaLV Env (MLV/GaLV Env) was restricted 50- to 100-fold by Vpu. A Vpu mutant containing a scrambled membrane-spanning domain, Vpu(RD), was still able to restrict MLV/GaLV Env, but mutation of the serine residues at positions 52 and 56 completely alleviated the restriction. Loss of infectivity appeared to be caused by reduced MLV/GaLV Env incorporation into viral particles. The mechanism of this downmodulation appears to be distinct from Vpu-mediated CD4 downmodulation because Vpu-expressing cells that failed to produce infectious HIV-1 particles nonetheless continued to display robust surface MLV/GaLV Env expression. In addition, if MLV and HIV-1 were simultaneously introduced into the same cells, only the HIV-1 particle infectivity was restricted by Vpu. Collectively, these data suggest that Vpu modulates the cellular distribution of MLV/GaLV Env, preventing its recruitment to HIV-1 budding sites.

  15. Clustering and Mobility of HIV-1 Env at Viral Assembly Sites Predict Its Propensity To Induce Cell-Cell Fusion

    PubMed Central

    Roy, Nathan H.; Chan, Jany; Lambelé, Marie

    2013-01-01

    HIV-1 Env mediates virus attachment to and fusion with target cell membranes, and yet, while Env is still situated at the plasma membrane of the producer cell and before its incorporation into newly formed particles, Env already interacts with the viral receptor CD4 on target cells, thus enabling the formation of transient cell contacts that facilitate the transmission of viral particles. During this first encounter with the receptor, Env must not induce membrane fusion, as this would prevent the producer cell and the target cell from separating upon virus transmission, but how Env's fusion activity is controlled remains unclear. To gain a better understanding of the Env regulation that precedes viral transmission, we examined the nanoscale organization of Env at the surface of producer cells. Utilizing superresolution microscopy (stochastic optical reconstruction microscopy [STORM]) and fluorescence recovery after photobleaching (FRAP), we quantitatively assessed the clustering and dynamics of Env upon its arrival at the plasma membrane. We found that Gag assembly induced the aggregation of small Env clusters into larger domains and that these domains were completely immobile. Truncation of the cytoplasmic tail (CT) of Env abrogated Gag's ability to induce Env clustering and restored Env mobility at assembly sites, both of which correlated with increased Env-induced fusion of infected and uninfected cells. Hence, while Env trapping by Gag secures Env incorporation into viral particles, Env clustering and its sequestration at assembly sites likely also leads to the repression of its fusion function, and thus, by preventing the formation of syncytia, Gag helps to secure efficient transfer of viral particles to target cells. PMID:23637402

  16. GenBank.

    PubMed

    Benson, Dennis A; Karsch-Mizrachi, Ilene; Lipman, David J; Ostell, James; Wheeler, David L

    2008-01-01

    GenBank (R) is a comprehensive database that contains publicly available nucleotide sequences for more than 260 000 named organisms, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects. Most submissions are made using the web-based BankIt or standalone Sequin programs and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the European Molecular Biology Laboratory Nucleotide Sequence Database in Europe and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through NCBI's retrieval system, Entrez, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI Homepage: www.ncbi.nlm.nih.gov.

  17. GenBank.

    PubMed

    Benson, Dennis A; Karsch-Mizrachi, Ilene; Lipman, David J; Ostell, James; Sayers, Eric W

    2011-01-01

    GenBank® is a comprehensive database that contains publicly available nucleotide sequences for more than 380,000 organisms named at the genus level or lower, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects, including whole genome shotgun (WGS) and environmental sampling projects. Most submissions are made using the web-based BankIt or standalone Sequin programs, and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the European Nucleotide Archive (ENA) and the DNA Data Bank of Japan (DDBJ) ensures worldwide coverage. GenBank is accessible through the NCBI Entrez retrieval system that integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI Homepage: www.ncbi.nlm.nih.gov.

  18. GenBank.

    PubMed

    Benson, Dennis A; Karsch-Mizrachi, Ilene; Lipman, David J; Ostell, James; Sayers, Eric W

    2010-01-01

    GenBank is a comprehensive database that contains publicly available nucleotide sequences for more than 300,000 organisms named at the genus level or lower, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects, including whole genome shotgun (WGS) and environmental sampling projects. Most submissions are made using the web-based BankIt or standalone Sequin programs, and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the European Molecular Biology Laboratory Nucleotide Sequence Database in Europe and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through the NCBI Entrez retrieval system, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bi-monthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI homepage: www.ncbi.nlm.nih.gov.

  19. Using GenBank.

    PubMed

    Wheeler, David

    2007-01-01

    GenBank(R) is a comprehensive database of publicly available DNA sequences for more than 205,000 named organisms and for more than 60,000 within the embryophyta, obtained through submissions from individual laboratories and batch submissions from large-scale sequencing projects. Daily data exchange with the European Molecular Biology Laboratory (EMBL) in Europe and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through the National Center for Biotechnology Information (NCBI) retrieval system, Entrez, which integrates data from the major DNA and protein sequence databases with taxonomy, genome, mapping, protein structure, and domain information and the biomedical journal literature through PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available through FTP. GenBank usage scenarios ranging from local analyses of the data available through FTP to online analyses supported by the NCBI Web-based tools are discussed. To access GenBank and its related retrieval and analysis services, go to the NCBI Homepage at http://www.ncbi.nlm.nih.gov.

  20. GenBank.

    PubMed

    Benson, Dennis A; Karsch-Mizrachi, Ilene; Lipman, David J; Ostell, James; Wheeler, David L

    2007-01-01

    GenBank (R) is a comprehensive database that contains publicly available nucleotide sequences for more than 240 000 named organisms, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects. Most submissions are made using the web-based BankIt or standalone Sequin programs and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the EMBL Data Library in Europe and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through NCBI's retrieval system, Entrez, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI Homepage (www.ncbi.nlm.nih.gov).

  1. GenBank.

    PubMed

    Benson, Dennis A; Karsch-Mizrachi, Ilene; Lipman, David J; Ostell, James; Wheeler, David L

    2006-01-01

    GenBank (R) is a comprehensive database that contains publicly available DNA sequences for more than 205 000 named organisms, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects. Most submissions are made using the Web-based BankIt or standalone Sequin programs and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the EMBL Data Library in Europe and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through NCBI's retrieval system, Entrez, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, go to the NCBI Homepage at www.ncbi.nlm.nih.gov.

  2. GenBank.

    PubMed

    Benson, Dennis A; Karsch-Mizrachi, Ilene; Lipman, David J; Ostell, James; Sayers, Eric W

    2009-01-01

    GenBank is a comprehensive database that contains publicly available nucleotide sequences for more than 300,000 organisms named at the genus level or lower, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects. Most submissions are made using the web-based BankIt or standalone Sequin programs, and accession numbers are assigned by GenBank(R) staff upon receipt. Daily data exchange with the European Molecular Biology Laboratory Nucleotide Sequence Database in Europe and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through the National Center for Biotechnology Information (NCBI) Entrez retrieval system, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI Homepage: www.ncbi.nlm.nih.gov.

  3. GenBank.

    PubMed

    Benson, Dennis A; Karsch-Mizrachi, Ilene; Lipman, David J; Ostell, James; Wheeler, David L

    2005-01-01

    GenBank is a comprehensive database that contains publicly available DNA sequences for more than 165,000 named organisms, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects. Most submissions are made using the web-based BankIt or standalone Sequin programs and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the EMBL Data Library in the UK and the DNA Data Bank of Japan helps to ensure worldwide coverage. GenBank is accessible through NCBI's retrieval system, Entrez, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, go to the NCBI Homepage at http://www.ncbi.nlm.nih.gov.

  4. GenBank

    PubMed Central

    Benson, Dennis A.; Clark, Karen; Karsch-Mizrachi, Ilene; Lipman, David J.; Ostell, James; Sayers, Eric W.

    2014-01-01

    GenBank® is a comprehensive database that contains publicly available nucleotide sequences for over 280 000 formally described species. These sequences are obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects, including whole-genome shotgun and environmental sampling projects. Most submissions are made using the web-based BankIt or standalone Sequin programs, and GenBank staff assign accession numbers upon data receipt. Daily data exchange with the European Nucleotide Archive and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through the National Center for Biotechnology Information (NCBI) Entrez retrieval system, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI home page: www.ncbi.nlm.nih.gov. PMID:24217914

  5. Using GenBank.

    PubMed

    Sayers, Eric W; Karsch-Mizrachi, Ilene

    2016-01-01

    GenBank(®) is a comprehensive database of publicly available DNA sequences for 300,000 named organisms, more than 110,000 within the embryophyta, obtained through submissions from individual laboratories and batch submissions from large-scale sequencing projects. Daily data exchange with the European Nucleotide Archive (ENA) in Europe and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through the NCBI Entrez retrieval system that integrates data from the major DNA and protein sequence databases with taxonomy, genome, mapping, protein structure and domain information, as well as the biomedical journal literature in PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. GenBank usage scenarios ranging from local analyses of the data available via FTP to online analyses supported by the NCBI web-based tools are discussed. To access GenBank and its related retrieval and analysis services, go to the NCBI home page at www.ncbi.nlm.nih.gov .

  6. GenBank

    PubMed Central

    Benson, Dennis A.; Cavanaugh, Mark; Clark, Karen; Karsch-Mizrachi, Ilene; Lipman, David J.; Ostell, James; Sayers, Eric W.

    2013-01-01

    GenBank® (http://www.ncbi.nlm.nih.gov) is a comprehensive database that contains publicly available nucleotide sequences for almost 260 000 formally described species. These sequences are obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects, including whole-genome shotgun (WGS) and environmental sampling projects. Most submissions are made using the web-based BankIt or standalone Sequin programs, and GenBank staff assigns accession numbers upon data receipt. Daily data exchange with the European Nucleotide Archive (ENA) and the DNA Data Bank of Japan (DDBJ) ensures worldwide coverage. GenBank is accessible through the NCBI Entrez retrieval system, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI home page: www.ncbi.nlm.nih.gov. PMID:23193287

  7. GenBank

    PubMed Central

    Clark, Karen; Karsch-Mizrachi, Ilene; Lipman, David J.; Ostell, James; Sayers, Eric W.

    2016-01-01

    GenBank® (www.ncbi.nlm.nih.gov/genbank/) is a comprehensive database that contains publicly available nucleotide sequences for over 340 000 formally described species. Recent developments include a new starting page for submitters, a shift toward using accession.version identifiers rather than GI numbers, a wizard for submitting 16S rRNA sequences, and an Identical Protein Report to address growing issues of data redundancy. GenBank organizes the sequence data received from individual laboratories and large-scale sequencing projects into 18 divisions, and GenBank staff assign unique accession.version identifiers upon data receipt. Most submitters use the web-based BankIt or standalone Sequin programs. Daily data exchange with the European Nucleotide Archive (ENA) and the DNA Data Bank of Japan (DDBJ) ensures worldwide coverage. GenBank is accessible through the nuccore, nucest, and nucgss databases of the Entrez retrieval system, which integrates these records with a variety of other data including taxonomy nodes, genomes, protein structures, and biomedical journal literature in PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. PMID:26590407

  8. GenBank

    PubMed Central

    Benson, Dennis A.; Karsch-Mizrachi, Ilene; Lipman, David J.; Ostell, James; Sayers, Eric W.

    2010-01-01

    GenBank® is a comprehensive database that contains publicly available nucleotide sequences for more than 300 000 organisms named at the genus level or lower, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects, including whole genome shotgun (WGS) and environmental sampling projects. Most submissions are made using the web-based BankIt or standalone Sequin programs, and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the European Molecular Biology Laboratory Nucleotide Sequence Database in Europe and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through the NCBI Entrez retrieval system, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bi-monthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI homepage: www.ncbi.nlm.nih.gov. PMID:19910366

  9. GenBank

    PubMed Central

    Benson, Dennis A.; Karsch-Mizrachi, Ilene; Clark, Karen; Lipman, David J.; Ostell, James; Sayers, Eric W.

    2012-01-01

    GenBank® is a comprehensive database that contains publicly available nucleotide sequences for more than 250 000 formally described species. These sequences are obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects, including whole-genome shotgun (WGS) and environmental sampling projects. Most submissions are made using the web-based BankIt or standalone Sequin programs, and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the European Nucleotide Archive (ENA) and the DNA Data Bank of Japan (DDBJ) ensures worldwide coverage. GenBank is accessible through the NCBI Entrez retrieval system, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI home page: www.ncbi.nlm.nih.gov. PMID:22144687

  10. GenBank

    PubMed Central

    Benson, Dennis A.; Karsch-Mizrachi, Ilene; Lipman, David J.; Ostell, James; Sayers, Eric W.

    2011-01-01

    GenBank® is a comprehensive database that contains publicly available nucleotide sequences for more than 380 000 organisms named at the genus level or lower, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects, including whole genome shotgun (WGS) and environmental sampling projects. Most submissions are made using the web-based BankIt or standalone Sequin programs, and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the European Nucleotide Archive (ENA) and the DNA Data Bank of Japan (DDBJ) ensures worldwide coverage. GenBank is accessible through the NCBI Entrez retrieval system that integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI Homepage: www.ncbi.nlm.nih.gov. PMID:21071399

  11. Effects of modification of the HIV-1 Env cytoplasmic tail on immunogenicity of VLP vaccines.

    PubMed

    Vzorov, Andrei N; Wang, Li; Chen, Jianjun; Wang, Bao-Zhong; Compans, Richard W

    2016-02-01

    We investigated the effects on assembly and antigenic properties of specific modifications of the transmembrane spanning (TMS) and cytoplasmic tail (CT) domains of HIV-1 Env from a transmitted/founder (T/F) ZM53 Env glycoprotein. A construct containing a short version of the TMS domain derived from the mouse mammary tumor virus (MMTV) Env with or without a GCN4 trimerization sequence in the CT exhibited the highest levels of incorporation into VLPs and induced the highest titers of anti-Env IgG immune responses in a VLP context. Sera from guinea pigs immunized by VLPs with high Env content, and containing the CT trimerization sequence, had increased neutralization activity and antibody avidity. A cross-clade prime-boost regimen with clade B SF162 or clade C ZM53 Env DNA priming and boosting with VLPs containing modified ZM53 Env further enhanced these immune responses. The modified VLPs demonstrate improved potential as HIV-1 vaccine antigens.

  12. Chenggangzhangella methanolivorans gen. nov., sp. nov., a member of the family Methylocystaceae, transfer of Methylopila helvetica Doronina et al. 2000 to Albibacter helveticus comb. nov. and emended description of the genus Albibacter.

    PubMed

    Yang, Li-Qiang; Liu, Lan; Salam, Nimaichand; Xiao, Min; Kim, Chang-Jin; Hozzein, Wael N; Park, Dong-Jin; Li, Wen-Jun; Zhang, Hui-Wen

    2016-09-01

    A Gram-stain-negative, rod-shaped, non-motile and aerobic bacterial strain, designated CHL1T, was isolated from a sludge sample collected from a sewage treatment tank of an agricultural chemical factory. The strain grew at salinities of 0.5-5 % (w/v) NaCl (optimum 2.5 %). Growth occurred at pH 6.0-8.0 (optimum pH 7.0) and 5-40 °C (optimum 28-30 °C). The genomic DNA G+C content was determined to be 70.4 mol%. Q-10 was detected as the respiratory quinone. The major fatty acids (>10 %) were C18 : 1ω7c and/or C18 : 1ω6c and C16 : 0. The polar lipids consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, two unidentified phospholipids and two unidentified aminophospholipids. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain CHL1T formed a distinct clade with Albibacter methylovorans DSM 22840T and Methylopila helvetica DM9T within the family Methylocystaceae. On the basis of phenotypic, chemotaxonomic and phylogenetic characteristics, the strain merits recognition as a representative of a novel species of a new genus within the family Methylocystaceae, for which the name Chenggangzhangella methanolivorans gen. nov., sp. nov. is proposed. The type strain of the type species is CHL1T (=KCTC 42661T=CCTCC AB 2015175T). In addition, the species Methylopila helveticaDoronina et al. (2000) is proposed to be transferred to the genus Albibacter as Albibacterhelveticus comb. nov. (type strain DM9T=CIP 106788=VKM B-2189) on the basis of the phylogenetic analysis. An emended description of the genus Albibacter is also provided.

  13. Rapid evolution of the env gene leader sequence in cats naturally infected with feline immunodeficiency virus.

    PubMed

    Bęczkowski, Paweł M; Hughes, Joseph; Biek, Roman; Litster, Annette; Willett, Brian J; Hosie, Margaret J

    2015-04-01

    Analysing the evolution of feline immunodeficiency virus (FIV) at the intra-host level is important in order to address whether the diversity and composition of viral quasispecies affect disease progression. We examined the intra-host diversity and the evolutionary rates of the entire env and structural fragments of the env sequences obtained from sequential blood samples in 43 naturally infected domestic cats that displayed different clinical outcomes. We observed in the majority of cats that FIV env showed very low levels of intra-host diversity. We estimated that env evolved at a rate of 1.16×10(-3) substitutions per site per year and demonstrated that recombinant sequences evolved faster than non-recombinant sequences. It was evident that the V3-V5 fragment of FIV env displayed higher evolutionary rates in healthy cats than in those with terminal illness. Our study provided the first evidence that the leader sequence of env, rather than the V3-V5 sequence, had the highest intra-host diversity and the highest evolutionary rate of all env fragments, consistent with this region being under a strong selective pressure for genetic variation. Overall, FIV env displayed relatively low intra-host diversity and evolved slowly in naturally infected cats. The maximum evolutionary rate was observed in the leader sequence of env. Although genetic stability is not necessarily a prerequisite for clinical stability, the higher genetic stability of FIV compared with human immunodeficiency virus might explain why many naturally infected cats do not progress rapidly to AIDS. © 2015 The Authors.

  14. Rapid evolution of the env gene leader sequence in cats naturally infected with feline immunodeficiency virus

    PubMed Central

    Hughes, Joseph; Biek, Roman; Litster, Annette; Willett, Brian J.; Hosie, Margaret J.

    2015-01-01

    Analysing the evolution of feline immunodeficiency virus (FIV) at the intra-host level is important in order to address whether the diversity and composition of viral quasispecies affect disease progression. We examined the intra-host diversity and the evolutionary rates of the entire env and structural fragments of the env sequences obtained from sequential blood samples in 43 naturally infected domestic cats that displayed different clinical outcomes. We observed in the majority of cats that FIV env showed very low levels of intra-host diversity. We estimated that env evolved at a rate of 1.16×10−3 substitutions per site per year and demonstrated that recombinant sequences evolved faster than non-recombinant sequences. It was evident that the V3–V5 fragment of FIV env displayed higher evolutionary rates in healthy cats than in those with terminal illness. Our study provided the first evidence that the leader sequence of env, rather than the V3–V5 sequence, had the highest intra-host diversity and the highest evolutionary rate of all env fragments, consistent with this region being under a strong selective pressure for genetic variation. Overall, FIV env displayed relatively low intra-host diversity and evolved slowly in naturally infected cats. The maximum evolutionary rate was observed in the leader sequence of env. Although genetic stability is not necessarily a prerequisite for clinical stability, the higher genetic stability of FIV compared with human immunodeficiency virus might explain why many naturally infected cats do not progress rapidly to AIDS. PMID:25535323

  15. env Gene of Chicken RNA Tumor Viruses: Extent of Conservation in Cellular and Viral Genomes

    PubMed Central

    Fujita, Donald J.; Tal, Jacov; Varmus, Harold E.; Bishop, J. Michael

    1978-01-01

    The env gene of avian sarcoma-leukosis viruses codes for envelope glycoproteins that determine viral host range, antigenic specificity, and interference patterns. We used molecular hybridization to analyze the natural distribution and possible origins of the nucleotide sequences that encode env; our work exploited the availability of radioactive DNA (cDNAgp) complementary to most or all of env. env sequences were detectable in the DNAs of chickens which synthesized an env gene product (chick helper factor positive) encoded by an endogenous viral gene and also in the DNAs of chickens which synthesized little or no env gene product (chick helper factor negative). env sequences were not detectable in DNAs from Japanese quail, ring-necked pheasant, golden pheasant, duck, squab, salmon sperm, or calf thymus. The detection of sequences closely related to viral env only in chicken DNA contrasts sharply with the demonstration that the transforming gene (src) of avian sarcoma viruses has readily detectable homologues in the DNAs of all avian species tested [D. Stehelin, H. E. Varmus, J. M. Bishop, and P. K. Vogt, Nature (London) 260: 170-173, 1976] and in the DNAs of other vertebrates (D. Spector, personal communication). Thermal denaturation studies on duplexes formed between cDNAgp and chicken DNA and also between cDNAgp and RNAs of subgroup A to E viruses derived from chickens indicated that these duplexes were well matched. In contrast, cDNAgp did not form stable hybrids with RNAs of viruses which were isolated from ring-necked and golden pheasants. We conclude that substantial portions of nucleotide sequences within the env genes of viruses of subgroups A to E are closely related and that these genes probably have a common, perhaps cellular, evolutionary origin. PMID:212576

  16. The signature of climate change on surface ozone: Using the Online integrated climate-chemistry model (EnvClimA)

    NASA Astrophysics Data System (ADS)

    Zakey, A. S.; Baklanov, A.; Solmon, F.; Giorgi, F.; Nuterman, R.; Sass, B. H.; Korsholm, U. S.; Nielsen, K. P.; Sørensen, J. H.; Mahura, A.

    2012-04-01

    The signature of climate change on European surface ozone was studied using the online integrated climate-chemistry model for Environmental applications (EnvClimA). The EnvClimA is an environmental version of the regional climate model of the International Centre for Theoretical Physics (ICTP) (RegCM-CHEM4, Shalapy et al., 2012). The model domain has a horizontal resolution of 50 × 50 km and 18 vertical sigma levels. In this study, a 20 year simulation was preformed for the selected European domain for the reference (2000-2009) and future (2040-2049) periods. For both simulations, the initial and boundary conditions for the meteorological fields are provided every six hours from the global ECHAM5-r3 model. The chemical boundary conditions over Europe are provided also every six hours by the Danish Eulerian Hemispheric Model (DEHM). The anthropogenic emissions of nitrogen oxides, sulfur dioxide, ammonia, non-methane volatile organic carbon and carbon monoxide were taken from the IPCC-RCP4.5 future emission scenario. In this simulation the biogenic isoprene emissions are not considered, because the MEGAN module (on-line coupled with the land surface scheme in EnvClimA) tends to overestimate (almost twice) the total emitted biogenic isoprene. The EnvClimA results indicated zonal behavior of average daily maximum concentrations for the surface ozone (O3). In winter, model has a substantial negative bias for both mean and daily maximum O3. This may be due to an underestimation of the winter air temperature over north-eastern Europe and due to feedback (included in the model) of O3 on the meteorological variables. Although the model spatial correlation is rather poor for diurnal average concentration, but for the average of daily maximum O3 concentrations the model showed correlation coefficients higher than 0.8 during summer. The model always showed the highest spatial correlation over central and southern Europe. The general pattern indicated an increase of surface

  17. Characterisation of env and gag gene fragments of bovine leukemia viruses (BLVs) from cattle in Turkey.

    PubMed

    Alkan, Feray; Oğuzoğlu, Tuba Çiğdem; Timurkan, Mehmet Ozkan; Karapınar, Zeynep

    2011-10-01

    The aim of this work was to investigate the molecular characteristics of bovine leukemia viruses (BLVs) in Turkey. The variability of env and gag fragments of BLVs was examined using DNA from blood samples obtained for sequence analysis of BLVs in four cattle herds from three different geographical areas in Turkey. The env gene sequences were highly similar to those of Brasilian, Argentine, and Japanese BLV strains, while gag genes from Turkish BLV isolates showed greatest similarity to those of Iranian isolates. This paper is the first report on the partial characterisation of env and gag genetic fragments of BLVs from Turkey.

  18. Structure-Guided Redesign Increases the Propensity of HIV Env To Generate Highly Stable Soluble Trimers

    PubMed Central

    Guenaga, Javier; Dubrovskaya, Viktoriya; de Val, Natalia; Sharma, Shailendra K.; Carrette, Barbara; Ward, Andrew B.

    2015-01-01

    ABSTRACT Due to high viral diversity, an effective HIV-1 vaccine will likely require Envs derived from multiple subtypes to generate broadly neutralizing antibodies (bNAbs). Soluble Env mimics, like the native flexibly linked (NFL) and SOSIP trimers, derived from the subtype A BG505 Env, form homogeneous, stable native-like trimers. However, other Env sequences, such as JRFL and 16055 from subtypes B and C, do so to a lesser degree. The high-resolution BG505 SOSIP crystal structures permit the identification and redesign of Env elements involved in trimer stability. Here, we identified structure trimer-derived (TD) residues that increased the propensity of the subtype B JRFL and subtype C 16055 Env sequences to form well-ordered, homogenous, and highly stable soluble trimers. The generation of these spike mimics no longer required antibody-based selection, positive or negative. Using the redesigned subtype B and C trimer representatives as respective foundations, we further stabilized the NFL TD trimers by engineering an intraprotomer disulfide linkage in the prebridging sheet, I201C-A433C (CC), that locks the gp120 in the receptor nontriggered state. We demonstrated that this disulfide pair prevented CD4 induced-conformational rearrangements in NFL trimers derived from the prototypic subtype A, B, and C representatives. Coupling the TD-based design with the engineered disulfide linkage, CC, increased the propensity of Env to form soluble highly stable spike mimics that are resistant to CD4-induced changes. These advances will allow testing of the hypothesis that such stabilized immunogens will more efficiently elicit neutralizing antibodies in small-animal models and primates. IMPORTANCE HIV-1 displays unprecedented global diversity circulating in the human population. Since the envelope glycoprotein (Env) is the target of neutralizing antibodies, Env-based vaccine candidates that address such diversity are needed. Soluble well-ordered Env mimics, typified by NFL

  19. Novel Feline Leukemia Virus Interference Group Based on the env Gene

    PubMed Central

    Miyake, Ariko; Watanabe, Shinya; Hiratsuka, Takahiro; Ito, Jumpei; Ngo, Minh Ha; Makundi, Isaac; Kawasaki, Junna; Endo, Yasuyuki; Tsujimoto, Hajime

    2016-01-01

    Feline leukemia virus (FeLV) subgroups have emerged in infected cats via the mutation or recombination of the env gene of subgroup A FeLV (FeLV-A), the primary virus. We report the isolation and characterization of a novel env gene, TG35-2, and report that the TG35-2 pseudotype can be categorized as a novel FeLV subgroup. The TG35-2 envelope protein displays strong sequence identity to FeLV-A Env, suggesting that selection pressure in cats causes novel FeLV subgroups to emerge. PMID:26889025

  20. Novel Feline Leukemia Virus Interference Group Based on the env Gene.

    PubMed

    Miyake, Ariko; Watanabe, Shinya; Hiratsuka, Takahiro; Ito, Jumpei; Ngo, Minh Ha; Makundi, Isaac; Kawasaki, Junna; Endo, Yasuyuki; Tsujimoto, Hajime; Nishigaki, Kazuo

    2016-05-01

    Feline leukemia virus (FeLV) subgroups have emerged in infected cats via the mutation or recombination of the env gene of subgroup A FeLV (FeLV-A), the primary virus. We report the isolation and characterization of a novel env gene, TG35-2, and report that the TG35-2 pseudotype can be categorized as a novel FeLV subgroup. The TG35-2 envelope protein displays strong sequence identity to FeLV-A Env, suggesting that selection pressure in cats causes novel FeLV subgroups to emerge. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  1. GenBank

    PubMed Central

    Benson, Dennis A.; Clark, Karen; Karsch-Mizrachi, Ilene; Lipman, David J.; Ostell, James; Sayers, Eric W.

    2015-01-01

    GenBank® (http://www.ncbi.nlm.nih.gov/genbank/) is a comprehensive database that contains publicly available nucleotide sequences for over 300 000 formally described species. These sequences are obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects, including whole-genome shotgun and environmental sampling projects. Most submissions are made using the web-based BankIt or standalone Sequin programs, and GenBank staff assign accession numbers upon data receipt. Daily data exchange with the European Nucleotide Archive and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through the NCBI Entrez retrieval system, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. PMID:25414350

  2. GenBank

    PubMed Central

    Benson, Dennis A.; Cavanaugh, Mark; Clark, Karen; Karsch-Mizrachi, Ilene; Lipman, David J.; Ostell, James; Sayers, Eric W.

    2017-01-01

    GenBank® (www.ncbi.nlm.nih.gov/genbank/) is a comprehensive database that contains publicly available nucleotide sequences for 370 000 formally described species. These sequences are obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects, including whole genome shotgun (WGS) and environmental sampling projects. Most submissions are made using the web-based BankIt or the NCBI Submission Portal. GenBank staff assign accession numbers upon data receipt. Daily data exchange with the European Nucleotide Archive (ENA) and the DNA Data Bank of Japan (DDBJ) ensures worldwide coverage. GenBank is accessible through the NCBI Nucleotide database, which links to related information such as taxonomy, genomes, protein sequences and structures, and biomedical journal literature in PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. Recent updates include changes to policies regarding sequence identifiers, an improved 16S submission wizard, targeted loci studies, the ability to submit methylation and BioNano mapping files, and a database of anti-microbial resistance genes. PMID:27899564

  3. Quantifying Selection against Synonymous Mutations in HIV-1 env Evolution

    PubMed Central

    Zanini, Fabio

    2013-01-01

    Intrapatient evolution of human immunodeficiency virus type 1 (HIV-1) is driven by the adaptive immune system resulting in rapid change of HIV-1 proteins. When cytotoxic CD8+ T cells or neutralizing antibodies target a new epitope, the virus often escapes via nonsynonymous mutations that impair recognition. Synonymous mutations do not affect this interplay and are often assumed to be neutral. We test this assumption by tracking synonymous mutations in longitudinal intrapatient data from the C2-V5 part of the env gene. We find that most synonymous variants are lost even though they often reach high frequencies in the viral population, suggesting a cost to the virus. Using published data from SHAPE (selective 2′-hydroxyl acylation analyzed by primer extension) assays, we find that synonymous mutations that disrupt base pairs in RNA stems flanking the variable loops of gp120 are more likely to be lost than other synonymous changes: these RNA hairpins might be important for HIV-1. Computational modeling indicates that, to be consistent with the data, a large fraction of synonymous mutations in this genomic region need to be deleterious with a cost on the order of 0.002 per day. This weak selection against synonymous substitutions does not result in a strong pattern of conservation in cross-sectional data but slows down the rate of evolution considerably. Our findings are consistent with the notion that large-scale patterns of RNA structure are functionally relevant, whereas the precise base pairing pattern is not. PMID:23986591

  4. Differential binding of neutralizing and non-neutralizing antibodies to native-like soluble HIV-1 Env trimers, uncleaved Env proteins, and monomeric subunits.

    PubMed

    Yasmeen, Anila; Ringe, Rajesh; Derking, Ronald; Cupo, Albert; Julien, Jean-Philippe; Burton, Dennis R; Ward, Andrew B; Wilson, Ian A; Sanders, Rogier W; Moore, John P; Klasse, Per Johan

    2014-05-29

    The trimeric envelope glycoproteins (Env) on the surface of HIV-1 virions are the targets for neutralizing antibodies (NAbs). No candidate HIV-1 immunogen has yet induced potent, broadly active NAbs (bNAbs). Part of the explanation may be that previously tested Env proteins inadequately mimic the functional, native Env complex. Trimerization and the proteolytic processing of Env precursors into gp120 and gp41 profoundly alter antigenicity, but soluble cleaved trimers are too unstable to serve as immunogens. By introducing stabilizing mutations (SOSIP), we constructed soluble, cleaved Env trimers derived from the HIV-1 subtype A isolate BG505 that resemble native Env spikes on virions both structurally and antigenically. We used surface plasmon resonance (SPR) to quantify antibody binding to different forms of BG505 Env: the proteolytically cleaved SOSIP.664 trimers, cleaved gp120-gp41ECTO protomers, and gp120 monomers. Non-NAbs to the CD4-binding site bound only marginally to the trimers but equally well to gp120-gp41ECTO protomers and gp120 monomers, whereas the bNAb VRC01, directed to the CD4bs, bound to all three forms. In contrast, bNAbs to V1V2 glycan-dependent epitopes bound preferentially (PG9 and PG16) or exclusively (PGT145) to trimers. We also explored the antigenic consequences of three different features of SOSIP.664 gp140 trimers: the engineered inter-subunit disulfide bond, the trimer-stabilizing I559P change in gp41ECTO, and proteolytic cleavage at the gp120-gp41ECTO junction. Each of these three features incrementally promoted native-like trimer antigenicity. We compared Fab and IgG versions of bNAbs and validated a bivalent model of IgG binding. The NAbs showed widely divergent binding kinetics and degrees of binding to native-like BG505 SOSIP.664. High off-rate constants and low stoichiometric estimates of NAb binding were associated with large amounts of residual infectivity after NAb neutralization of the corresponding BG505.T332N pseudovirus

  5. Differential binding of neutralizing and non-neutralizing antibodies to native-like soluble HIV-1 Env trimers, uncleaved Env proteins, and monomeric subunits

    PubMed Central

    2014-01-01

    Background The trimeric envelope glycoproteins (Env) on the surface of HIV-1 virions are the targets for neutralizing antibodies (NAbs). No candidate HIV-1 immunogen has yet induced potent, broadly active NAbs (bNAbs). Part of the explanation may be that previously tested Env proteins inadequately mimic the functional, native Env complex. Trimerization and the proteolytic processing of Env precursors into gp120 and gp41 profoundly alter antigenicity, but soluble cleaved trimers are too unstable to serve as immunogens. By introducing stabilizing mutations (SOSIP), we constructed soluble, cleaved Env trimers derived from the HIV-1 subtype A isolate BG505 that resemble native Env spikes on virions both structurally and antigenically. Results We used surface plasmon resonance (SPR) to quantify antibody binding to different forms of BG505 Env: the proteolytically cleaved SOSIP.664 trimers, cleaved gp120-gp41ECTO protomers, and gp120 monomers. Non-NAbs to the CD4-binding site bound only marginally to the trimers but equally well to gp120-gp41ECTO protomers and gp120 monomers, whereas the bNAb VRC01, directed to the CD4bs, bound to all three forms. In contrast, bNAbs to V1V2 glycan-dependent epitopes bound preferentially (PG9 and PG16) or exclusively (PGT145) to trimers. We also explored the antigenic consequences of three different features of SOSIP.664 gp140 trimers: the engineered inter-subunit disulfide bond, the trimer-stabilizing I559P change in gp41ECTO, and proteolytic cleavage at the gp120-gp41ECTO junction. Each of these three features incrementally promoted native-like trimer antigenicity. We compared Fab and IgG versions of bNAbs and validated a bivalent model of IgG binding. The NAbs showed widely divergent binding kinetics and degrees of binding to native-like BG505 SOSIP.664. High off-rate constants and low stoichiometric estimates of NAb binding were associated with large amounts of residual infectivity after NAb neutralization of the corresponding BG505.T

  6. Receptor Activation of HIV-1 Env Leads to Asymmetric Exposure of the gp41 Trimer

    PubMed Central

    2016-01-01

    Structural rearrangements of HIV-1 glycoprotein Env promote viral entry through membrane fusion. Env is a symmetric homotrimer with each protomer composed of surface subunit gp120 and transmembrane subunit gp41. Cellular CD4- and chemokine receptor-binding to gp120 coordinate conformational changes in gp41, first to an extended prehairpin intermediate (PHI) and, ultimately, into a fusogenic trimer-of-hairpins (TOH). HIV-1 fusion inhibitors target gp41 in the PHI and block TOH formation. To characterize structural transformations into and through the PHI, we employed asymmetric Env trimers containing both high and low affinity binding sites for individual fusion inhibitors. Asymmetry was achieved using engineered Env heterotrimers composed of protomers deficient in either CD4- or chemokine receptor-binding. Linking receptor engagement to inhibitor affinity allowed us to assess conformational changes of individual Env protomers in the context of a functioning trimer. We found that the transition into the PHI could occur symmetrically or asymmetrically depending on the stoichiometry of CD4 binding. Sequential engagement of multiple CD4s promoted progressive exposure of individual fusion inhibitor binding sites in a CD4-dependent fashion. By contrast, engagement of only a single CD4 molecule led to a delayed, but symmetric, exposure of the gp41 trimer. This complex coupling between Env-CD4 interaction and gp41 exposure explained the multiphasic fusion-inhibitor titration observed for a mutant Env homotrimer with a naturally asymmetric gp41. Our results suggest that the spatial and temporal exposure of gp41 can proceed in a nonconcerted, asymmetric manner depending on the number of CD4s that engage the Env trimer. The findings have important implications for the mechanism of viral membrane fusion and the development of vaccine candidates designed to elicit neutralizing antibodies targeting gp41 in the PHI. PMID:27992602

  7. Receptor Activation of HIV-1 Env Leads to Asymmetric Exposure of the gp41 Trimer.

    PubMed

    Khasnis, Mukta D; Halkidis, Konstantine; Bhardwaj, Anshul; Root, Michael J

    2016-12-01

    Structural rearrangements of HIV-1 glycoprotein Env promote viral entry through membrane fusion. Env is a symmetric homotrimer with each protomer composed of surface subunit gp120 and transmembrane subunit gp41. Cellular CD4- and chemokine receptor-binding to gp120 coordinate conformational changes in gp41, first to an extended prehairpin intermediate (PHI) and, ultimately, into a fusogenic trimer-of-hairpins (TOH). HIV-1 fusion inhibitors target gp41 in the PHI and block TOH formation. To characterize structural transformations into and through the PHI, we employed asymmetric Env trimers containing both high and low affinity binding sites for individual fusion inhibitors. Asymmetry was achieved using engineered Env heterotrimers composed of protomers deficient in either CD4- or chemokine receptor-binding. Linking receptor engagement to inhibitor affinity allowed us to assess conformational changes of individual Env protomers in the context of a functioning trimer. We found that the transition into the PHI could occur symmetrically or asymmetrically depending on the stoichiometry of CD4 binding. Sequential engagement of multiple CD4s promoted progressive exposure of individual fusion inhibitor binding sites in a CD4-dependent fashion. By contrast, engagement of only a single CD4 molecule led to a delayed, but symmetric, exposure of the gp41 trimer. This complex coupling between Env-CD4 interaction and gp41 exposure explained the multiphasic fusion-inhibitor titration observed for a mutant Env homotrimer with a naturally asymmetric gp41. Our results suggest that the spatial and temporal exposure of gp41 can proceed in a nonconcerted, asymmetric manner depending on the number of CD4s that engage the Env trimer. The findings have important implications for the mechanism of viral membrane fusion and the development of vaccine candidates designed to elicit neutralizing antibodies targeting gp41 in the PHI.

  8. Phylogenetics of HIV-1 subtype G env: Greater complexity and older origins than previously reported.

    PubMed

    Tongo, Marcel; Essomba, René G; Nindo, Frederick; Abrahams, Fatima; Nanfack, Aubin Joseph; Fokam, Joseph; Takou, Desire; Torimiro, Judith N; Mpoudi-Ngole, Eitel; Burgers, Wendy A; Martin, Darren P; Dorfman, Jeffrey R

    2015-10-01

    HIV-1 subtype G has played an early and central role in the emergent complexity of the HIV-1 group M (HIV-1M) epidemic in central/west Africa. Here, we analysed new subtype G env sequences sampled from 8 individuals in Yaoundé, Cameroon during 2007-2010, together with all publically available subtype G-attributed full-length env sequences with known sampling dates and locations. We inferred that the most recent common ancestor (MRCA) of the analysed subtype G env sequences most likely occurred in ∼1953 (95% Highest Posterior Density interval [HPD] 1939-1963): about 15 years earlier than previous estimates. We found that the subtype G env phylogeny has a complex structure including seven distinct lineages, each likely dating back to the late 1960s or early 1970s. Sequences from Angola, Gabon and the Democratic Republic of Congo failed to group consistently in these lineages, possibly because they are related to more ancient sequences that are poorly sampled. The circulating recombinant form (CRF), CRF06_cpx env sequences but not CRF25_cpx env sequences are phylogenetically nested within the subtype G clade. This confirms that the CRF06_cpx env plausibly was derived through recombination from a subtype G parent, and suggests that the CRF25_cpx env was likely derived from an HIV-1M lineage related to the MRCA of subtype G that has remained undiscovered and may be extinct. Overall, this fills important gaps in our knowledge of the early events in the spread of HIV-1M.

  9. Interrelationships of VEL1 and ENV1 in light response and development in Trichoderma reesei.

    PubMed

    Bazafkan, Hoda; Dattenböck, Christoph; Stappler, Eva; Beier, Sabrina; Schmoll, Monika

    2017-01-01

    Sexual development is regulated by a complex regulatory mechanism in fungi. For Trichoderma reesei, the light response pathway was shown to impact sexual development, particularly through the photoreceptor ENVOY. Moreover, T. reesei communicates chemically with a potential mating partner in its vicinity, a response which is mediated by the velvet family protein VEL1 and its impact on secondary metabolism. We therefore studied the regulatory interactions of ENV1 and VEL1 with a focus on sexual development. Although individual mutants in both genes are female sterile under standard crossing conditions (light-dark cycles), an altered light regime enabled sexual development, which we found to be due to conditional female sterility of Δenv1, but not Δvel1. Phenotypes of growth and asexual sporulation as well as regulation of the peptide pheromone precursors of double mutants suggested that ENV1 and VEL1 balance positive and negative regulators of these functions. Additionally, VEL1 contributed to the strong deregulation of the pheromone system observed in env1 mutants. Female sterility of Δvel1 was rescued by deletion of env1 in darkness in MAT1-1, indicating a block of sexual development by ENV1 in darkness that is balanced by VEL1 in the wild-type. We conclude that ENV1 and VEL1 exert complementing functions in development of T. reesei. Our results further showed that the different developmental phenotypes of vel1/veA mutants in T. reesei and Aspergillus nidulans are not due to the presence or function of ENV1 in the VELVET regulatory pathway in T. reesei.

  10. Diverse antibody genetic and recognition properties revealed following HIV-1 Env immunization

    PubMed Central

    Phad, Ganesh E.; Bernat, Néstor Vázquez; Feng, Yu; Ingale, Jidnyasa; Murillo, Paola Andrea Martinez; O’Dell, Sijy; Li, Yuxing; Mascola, John R.; Sundling, Christopher; Wyatt, Richard T.; Karlsson Hedestam, Gunilla B.

    2015-01-01

    Isolation of monoclonal antibodies (MAbs) elicited by vaccination provides opportunities to define the development of effective immunity. Ab responses elicited by current HIV-1 envelope glycoprotein (Env) immunogens display narrow neutralizing activity with limited capacity to block infection by tier 2 viruses. Intense work in the field suggests that improved Env immunogens are forthcoming and it is therefore important to concurrently develop approaches to investigate the quality of vaccine-elicited responses at a higher level of resolution. Here, we cloned a representative set of MAbs elicited by a model Env immunogen in rhesus macaques and comprehensively characterized their genetic and functional properties. The MAbs were genetically diverse, even within groups of Abs targeting the same sub-region of Env, consistent with a highly polyclonal response. MAbs directed against two sub-determinants of Env, the CD4 binding site (CD4bs) and the V3 region, could in part account for the neutralizing activity observed in the plasma of the animal from which they were cloned, demonstrating the power of MAb isolation for a detailed understanding of the elicited response. Finally, through comparative analyses of MAb binding and neutralizing capacity of HIV-1 using matched Envs, we demonstrate complex relationships between epitope recognition and accessibility, highlighting the protective quaternary packing of the HIV-1 spike relative to vaccine-induced MAbs. PMID:25964491

  11. Bryocella elongata gen. nov., sp. nov., a member of subdivision 1 of the Acidobacteria isolated from a methanotrophic enrichment culture, and emended description of Edaphobacter aggregans Koch et al. 2008.

    PubMed

    Dedysh, Svetlana N; Kulichevskaya, Irina S; Serkebaeva, Yulia M; Mityaeva, Maria A; Sorokin, Vladimir V; Suzina, Natalia E; Rijpstra, W Irene C; Damsté, Jaap S Sinninghe

    2012-03-01

    An aerobic, pink-pigmented, chemo-organotrophic bacterium, designated strain SN10(T), was isolated from a methanotrophic enrichment culture obtained from an acidic Sphagnum peat. This isolate was represented by Gram-negative, non-motile rods that multiply by normal cell division and form rosettes. Strain SN10(T) is an obligately acidophilic, mesophilic bacterium capable of growth at pH 3.2-6.6 (with an optimum at pH 4.7-5.2) and at 6-32 °C (with an optimum at 20-24 °C). The preferred growth substrates are sugars and several heteropolysaccharides of plant and microbial origin, such as pectin, lichenan, fucoidan and gellan gum. While not being capable of growth on C(1) compounds, strain SN10(T) can develop in co-culture with exopolysaccharide-producing methanotrophs by utilization of their capsular material. The major fatty acids determined in strain SN10(T) using the conventional lipid extraction procedure are iso-C(15:0) and C(16:1)ω7c. Upon hydrolysis of total cell material, substantial amounts of the uncommon membrane-spanning lipid 13,16-dimethyl octacosanedioic acid (isodiabolic acid) were also detected. The polar lipids are two phosphohexoses, phosphatidylethanolamine, phosphatidylglycerol and several phospholipids of unknown structure. The major quinone is MK-8. Pigments are carotenoids. The G+C content of the DNA is 60.7 mol%. Strain SN10(T) forms a separate lineage within subdivision 1 of the phylum Acidobacteria and displays 94.0-95.4% 16S rRNA gene sequence similarity to members of the genera Edaphobacter and Granulicella, 93.0-93.7% similarity to members of the genus Terriglobus and 92.2-92.3 % similarity to the type strains of Telmatobacter bradus and Acidobacterium capsulatum. Therefore, strain SN10(T) is classified within a novel genus and species, for which the name Bryocella elongata gen. nov., sp. nov. is proposed. Strain SN10(T) (=LMG 25276(T) =DSM 22489(T)) is the type strain of Bryocella elongata. An emended description of Edaphobacter

  12. Processing and amino acid sequence analysis of the mouse mammary tumor virus env gene product.

    PubMed Central

    Arthur, L O; Copeland, T D; Oroszlan, S; Schochetman, G

    1982-01-01

    The envelope proteins of mouse mammary tumor virus (MMTV) are synthesized from a subgenomic 24S mRNA as a 75,000-dalton glycosylated precursor polyprotein which is eventually processed to the mature glycoproteins gp52 and gp36. In vivo synthesis of this env precursor in the presence of the core glycosylation inhibitor tunicamycin yielded a precursor of approximately 61,000 daltons (P61env). However, a 67,000-dalton protein (P67env) was obtained from cell-free translation with the MMTV 24S mRNA as the template. To determine whether the portion of the protein cleaved from P67env to give P61env was removed from the NH2-terminal end of P67env and as such would represent a leader sequence, the NH2-terminal amino acid sequence of the terminal peptide gp52 was determined. Glutamic acid, and not methionine, was found to be the amino-terminal residue of gp52, indicating that the cleaved portion was derived from the NH2-terminal end of P67env. The NH2-terminal amino acid sequences of gp52's from endogenous and exogenous C3H MMTVs were determined though 46 residues and found to be identical. However, amino acid composition and type-specific gp52 radioimmunoassays from MMTVs grown in heterologous cells indicated primary structure differences between gp52's of the two viruses. The nucleic acid sequence of cloned MMTV DNA fragments (J. Majors and H. E. Varmus, personal communication) in conjunction with the NH2-terminal sequence of gp52 allowed localization of the env gene in the MMTV genome. Nucleotides coding for the NH2 terminus of gp52 begin approximately 0.8 kilobase to the 3' side of the single EcoRI cleavage site. Localization of the env gene at that point agrees with the proposed gene order -gag-pol-env- and also allows sufficient coding potential for the glycoprotein precursor without extending into the long terminal repeat. Images PMID:6281457

  13. Allostreptomyces psammosilenae gen. nov., sp. nov., an endophytic actinobacterium isolated from the roots of Psammosilene tunicoides and emended description of the family Streptomycetaceae [Waksman and Henrici (1943)AL] emend. Rainey et al. 1997, emend. Kim et al. 2003, emend. Zhi et al. 2009.

    PubMed

    Huang, Mei-Juan; Rao, Manik Prabhu Narsing; Salam, Nimaichand; Xiao, Min; Huang, Hai-Quan; Li, Wen-Jun

    2017-02-01

    A Gram-stain-positive actinobacterium, designated strain YIM DR4008T, was isolated from the root sample of Psammosilene tunicoides collected from Lijiang, Yunnan, China. Strain YIM DR4008T could grow at temperatures ranging from 10 to 50 °C (optimum 28-30 °C), at pH 5.0-11.0 (optimum pH 7.0) and in the presence of up to 4 % (w/v) NaCl. Sequence analysis of the 16S ribosomal RNA gene revealed that strain YIM DR4008T shared highest similarity (95.0 %) with Streptomyces griseoplanus NBRC 12779T and <95 % similarity with other known members of the genera Streptomyces, Kitasatospora and Streptacidiphilus. The diagnostic cell-wall diamino acid of strain YIM DR4008T was found to be ll-diaminopimelic acid. The whole-cell hydrolysates contained a major amount of galactose and mannose along with a small proportion of fucose, glucose, rhamnose and ribose. The polar lipids consisted of diphosphatidylglycerol, phosphatidylinositol mannosides and three unidentified phospholipids. The respiratory menaquinones were MK-9(H6) and MK-9(H8), while the major cellular fatty acids (>10 %) were anteiso-C15 : 0, C16 : 0, iso-C16 : 0, iso-C15 : 0 and anteiso-C17 : 0. The genomic DNA G+C content was determined to be 75.3 mol%. Based on the phenotypic, chemotaxonomic and molecular characteristics, strain YIM DR4008T is proposed to be recognized as a novel species of a new genus in the family Streptomycetaceae, with the name Allostreptomyces psammosilenae gen. nov., sp. nov. The type strain of the type species is YIM DR4008T (=DSM 42178T=CGMCC 4.7247T). An emended description of the family Streptomycetaceae is also provided.

  14. Antibody to gp41 MPER alters functional properties of HIV-1 Env without complete neutralization.

    PubMed

    Kim, Arthur S; Leaman, Daniel P; Zwick, Michael B

    2014-07-01

    Human antibody 10E8 targets the conserved membrane proximal external region (MPER) of envelope glycoprotein (Env) subunit gp41 and neutralizes HIV-1 with exceptional potency. Remarkably, HIV-1 containing mutations that reportedly knockout 10E8 binding to linear MPER peptides are partially neutralized by 10E8, producing a local plateau in the dose response curve. Here, we found that virus partially neutralized by 10E8 becomes significantly less neutralization sensitive to various MPER antibodies and to soluble CD4 while becoming significantly more sensitive to antibodies and fusion inhibitors against the heptad repeats of gp41. Thus, 10E8 modulates sensitivity of Env to ligands both pre- and post-receptor engagement without complete neutralization. Partial neutralization by 10E8 was influenced at least in part by perturbing Env glycosylation. With unliganded Env, 10E8 bound with lower apparent affinity and lower subunit occupancy to MPER mutant compared to wild type trimers. However, 10E8 decreased functional stability of wild type Env while it had an opposite, stabilizing effect on MPER mutant Envs. Clade C isolates with natural MPER polymorphisms also showed partial neutralization by 10E8 with altered sensitivity to various gp41-targeted ligands. Our findings suggest a novel mechanism of virus neutralization by demonstrating how antibody binding to the base of a trimeric spike cross talks with adjacent subunits to modulate Env structure and function. The ability of an antibody to stabilize, destabilize, partially neutralize as well as alter neutralization sensitivity of a virion spike pre- and post-receptor engagement may have implications for immunotherapy and vaccine design.

  15. Molecular identification of erythrocytic necrosis virus (ENV) from the blood of Pacific herring (Clupea pallasii)

    USGS Publications Warehouse

    Emmenegger, Eveline J.; Glenn, Jolene A.; Winton, James R.; Batts, William N.; Gregg, Jacob L.; Hershberger, Paul K.

    2014-01-01

    Viral erythrocytic necrosis (VEN) is a condition affecting the red blood cells of more than 20 species of marine and anadromous fishes in the North Atlantic and North Pacific Oceans. Among populations of Pacific herring (Clupea pallasii) on the west coast of North America the disease causes anemia and elevated mortality in periodic epizootics. Presently, VEN is diagnosed by observation of typical cytoplasmic inclusion bodies in stained blood smears from infected fish. The causative agent, erythrocytic necrosis virus (ENV), is unculturable and a presumed iridovirus by electron microscopy. In vivo amplification of the virus in pathogen-free laboratory stocks of Pacific herring with subsequent virus concentration, purification, DNA extraction, and high-throughput sequencing were used to obtain genomic ENV sequences. Fragments with the highest sequence identity to the family Iridoviridae were used to design four sets of ENV-specific polymerase chain reaction (PCR) primers. Testing of blood and tissue samples from experimentally and wild infected Pacific herring as well as DNA extracted from other amphibian and piscine iridoviruses verified the assays were specific to ENV with a limit of detection of 0.0003 ng. Preliminary phylogenetic analyses of a 1448 bp fragment of the putative DNA polymerase gene supported inclusion of ENV in a proposed sixth genus of the family Iridoviridae that contains other erythrocytic viruses from ectothermic hosts. This study provides the first molecular evidence of ENV's inclusion within the Iridoviridae family and offers conventional PCR assays as a means of rapidly surveying the ENV-status of wild and propagated Pacific herring stocks.

  16. Antibody to gp41 MPER Alters Functional Properties of HIV-1 Env without Complete Neutralization

    PubMed Central

    Kim, Arthur S.; Leaman, Daniel P.; Zwick, Michael B.

    2014-01-01

    Human antibody 10E8 targets the conserved membrane proximal external region (MPER) of envelope glycoprotein (Env) subunit gp41 and neutralizes HIV-1 with exceptional potency. Remarkably, HIV-1 containing mutations that reportedly knockout 10E8 binding to linear MPER peptides are partially neutralized by 10E8, producing a local plateau in the dose response curve. Here, we found that virus partially neutralized by 10E8 becomes significantly less neutralization sensitive to various MPER antibodies and to soluble CD4 while becoming significantly more sensitive to antibodies and fusion inhibitors against the heptad repeats of gp41. Thus, 10E8 modulates sensitivity of Env to ligands both pre- and post-receptor engagement without complete neutralization. Partial neutralization by 10E8 was influenced at least in part by perturbing Env glycosylation. With unliganded Env, 10E8 bound with lower apparent affinity and lower subunit occupancy to MPER mutant compared to wild type trimers. However, 10E8 decreased functional stability of wild type Env while it had an opposite, stabilizing effect on MPER mutant Envs. Clade C isolates with natural MPER polymorphisms also showed partial neutralization by 10E8 with altered sensitivity to various gp41-targeted ligands. Our findings suggest a novel mechanism of virus neutralization by demonstrating how antibody binding to the base of a trimeric spike cross talks with adjacent subunits to modulate Env structure and function. The ability of an antibody to stabilize, destabilize, partially neutralize as well as alter neutralization sensitivity of a virion spike pre- and post-receptor engagement may have implications for immunotherapy and vaccine design. PMID:25058619

  17. Molecular identification of erythrocytic necrosis virus (ENV) from the blood of Pacific herring (Clupea pallasii).

    PubMed

    Emmenegger, Eveline J; Glenn, Jolene A; Winton, James R; Batts, William N; Gregg, Jacob L; Hershberger, Paul K

    2014-11-07

    Viral erythrocytic necrosis (VEN) is a condition affecting the red blood cells of more than 20 species of marine and anadromous fishes in the North Atlantic and North Pacific Oceans. Among populations of Pacific herring (Clupea pallasii) on the west coast of North America the disease causes anemia and elevated mortality in periodic epizootics. Presently, VEN is diagnosed by observation of typical cytoplasmic inclusion bodies in stained blood smears from infected fish. The causative agent, erythrocytic necrosis virus (ENV), is unculturable and a presumed iridovirus by electron microscopy. In vivo amplification of the virus in pathogen-free laboratory stocks of Pacific herring with subsequent virus concentration, purification, DNA extraction, and high-throughput sequencing were used to obtain genomic ENV sequences. Fragments with the highest sequence identity to the family Iridoviridae were used to design four sets of ENV-specific polymerase chain reaction (PCR) primers. Testing of blood and tissue samples from experimentally and wild infected Pacific herring as well as DNA extracted from other amphibian and piscine iridoviruses verified the assays were specific to ENV with a limit of detection of 0.0003 ng. Preliminary phylogenetic analyses of a 1448 bp fragment of the putative DNA polymerase gene supported inclusion of ENV in a proposed sixth genus of the family Iridoviridae that contains other erythrocytic viruses from ectothermic hosts. This study provides the first molecular evidence of ENV's inclusion within the Iridoviridae family and offers conventional PCR assays as a means of rapidly surveying the ENV-status of wild and propagated Pacific herring stocks.

  18. GenBank

    PubMed Central

    Burks, Christian; Cinkosky, Michael J.; Fischer, William M.; Gilna, Paul; Hayden, Jamie E.-D.; Keen, Gifford M.; Kelly, Michael; Kristofferson, David; Lawrence, Julie

    1992-01-01

    The GenBank nucleotide sequence database now contains sequence data and associated annotation corresponding to 85,000,000 nucleotides in 67,000 entries from a total of 3,000 organisms. The input stream of data coming into the database is primarily as direct submissions from the scientific community on electronic media, with little or no data being keyboarded from the printed page by the databank staff. The data are maintained in a relational database management system and are made available in flatfile form through on-line access, and through various network and off-line computer-readable media. The data are also distributed in relational form through satellite copies at a number of institutions in the U.S. and elsewhere. In addition, GenBank provides the U.S. distribution center for the BIOSCI electronic bulletin board service. PMID:1598235

  19. Role of Virus Receptor Hyal2 in Oncogenic Transformation of Rodent Fibroblasts by Sheep Betaretrovirus Env Proteins

    PubMed Central

    Liu, Shan-Lu; Duh, Fuh-Mei; Lerman, Michael I.; Miller, A. Dusty

    2003-01-01

    The ovine betaretroviruses jaagsiekte sheep retrovirus (JSRV) and enzootic nasal tumor virus (ENTV) cause contagious cancers in the lungs and upper airways of sheep and goats. Oncogenic transformation assays using mouse and rat fibroblasts have localized the transforming activity to the Env proteins encoded by these viruses, which require the putative lung and breast cancer tumor suppressor hyaluronidase 2 (Hyal2) to promote virus entry into cells. These results suggested the hypothesis that the JSRV and ENTV Env proteins cause cancer by inhibiting the tumor suppressor activity of Hyal2. Consistent with this hypothesis, we show that human Hyal2 and other Hyal2 orthologs that can promote virus entry, including rat Hyal2, can suppress transformation by the Env proteins of JSRV and ENTV. Furthermore, we provide direct evidence for binding of the surface (SU) region of JSRV Env to human and rat Hyal2. However, mouse Hyal2 did not mediate entry of virions bearing JSRV or ENTV Env proteins, bound JSRV SU poorly if at all, and did not suppress transformation by the JSRV or ENTV Env proteins, indicating that mouse Hyal2 plays no role in transformation of mouse fibroblasts and that the Env proteins can transform at least some cells by a Hyal2-independent mechanism. Expression of human Hyal2 in mouse cells expressing JSRV Env caused a marked reduction in Env protein levels, indicating that human Hyal2 suppresses Env-mediated transformation in mouse cells by increasing Env degradation rather than by exerting a more general Env-independent tumor suppressor activity. PMID:12584308

  20. Intracellular trafficking of Gag and Env proteins and their interactions modulate pseudotyping of retroviruses.

    PubMed

    Sandrin, Virginie; Muriaux, Delphine; Darlix, Jean-Luc; Cosset, François-Loïc

    2004-07-01

    Glycoproteins derived from most retroviruses and from several families of enveloped viruses can form infectious pseudotypes with murine leukemia virus (MLV) and lentiviral core particles, like the MLV envelope glycoproteins (Env) that are incorporated on either virus type. However, coexpression of a given glycoprotein with heterologous core proteins does not always give rise to highly infectious viral particles, and restrictions on pseudotype formation have been reported. To understand the mechanisms that control the recruitment of viral surface glycoproteins on lentiviral and retroviral cores, we exploited the fact that the feline endogenous retrovirus RD114 glycoprotein does not efficiently pseudotype lentiviral cores derived from simian immunodeficiency virus, whereas it is readily incorporated onto MLV particles. Our results indicate that recruitment of glycoproteins by the MLV and lentiviral core proteins occurs in intracellular compartments and not at the cell surface. We found that Env and core protein colocalization in intracytoplasmic vesicles is required for pseudotype formation. By investigating MLV/RD114 Env chimeras, we show that signals in the cytoplasmic tail of either glycoprotein differentially influenced their intracellular localization; that of MLV allows endosomal localization and hence recruitment by both lentiviral and MLV cores. Furthermore, we found that upon membrane binding, MLV core proteins could relocalize Env glycoproteins in late endosomes and allow their incorporation on viral particles. Thus, intracellular colocalization, as well as interactions between Env and core proteins, may influence the recruitment of the glycoprotein onto viral particles and generate infectious pseudotyped viruses.

  1. Parenteral Administration of Capsule Depolymerase EnvD Prevents Lethal Inhalation Anthrax Infection

    PubMed Central

    Negus, David; Vipond, Julia; Hatch, Graham J.; Rayner, Emma L.

    2015-01-01

    Left untreated, inhalation anthrax is usually fatal. Vegetative forms of Bacillus anthracis survive in blood and tissues during infection due to elaboration of a protective poly-γ-d-glutamic acid (PDGA) capsule that permits uncontrolled bacterial growth in vivo, eventually leading to overwhelming bacillosis and death. As a measure to counter threats from multidrug-resistant strains, we are evaluating the prophylactic and therapeutic potential of the PDGA depolymerase EnvD, a stable and potent enzyme which rapidly and selectively removes the capsule from the surface of vegetative cells. Repeated intravenous administration of 10 mg/kg recombinant EnvD (rEnvD) to mice infected with lethal doses of B. anthracis Ames spores by inhalation prevented the emergence of symptoms of anthrax and death; all animals survived the 5-day treatment period, and 70% survived to the end of the 14-day observation period. In contrast to results in sham-treated animals, the lungs and spleen of rEnvD-dosed animals were free of gross pathological changes. We conclude that rEnvD has potential as an agent to prevent the emergence of inhalation anthrax in infected animals and is likely to be effective against drug-resistant forms of the pathogen. PMID:26438506

  2. Mutations in both env and gag genes are required for HIV-1 resistance to the polysulfonic dendrimer SPL2923, as corroborated by chimeric virus technology.

    PubMed

    Hantson, Anke; Fikkert, Valery; Van Remoortel, Barbara; Pannecouque, Chistophe; Cherepanov, Peter; Matthews, Barry; Holan, George; De Clercq, Erik; Vandamme, Anne-Mieke; Debyser, Zeger; Witvrouw, Myriam

    2005-01-01

    A drug-resistant NL4.3/SPL2923 strain has previously been generated by in vitro selection of HIV-1(NL4.3) in the presence of the polysulfonic dendrimer SPL2923 and mutations were reported in its gp120 gene (Witvrouw et al., 2000). Here, we further analysed the (cross) resistance profile of NL4.3/SPL2923. NL4.3/SPL2923 was found to contain additional mutations in gp41 and showed reduced susceptibility to SPL2923, dextran sulfate (DS) and enfuvirtide. To delineate to what extent the mutations in each env gene were accountable for the phenotypic (cross) resistance of NL4.3/SPL2923, the gp120-, gp41- and gp160-sequences derived from this strain were placed into a wild-type background using env chimeric virus technology (CVT). The cross resistance of NL4.3/SPL2923 towards DS was fully reproduced following gp160-recombination, while it was only partially reproduced following gp120- or gp41-recombination. The mutations in gp41 of NL4.3/SPL2923 were sufficient to reproduce the cross resistance to enfuvirtide. Unexpectedly, the reduced sensitivity towards SPL2923 was not fully reproduced after gp160-recombination. The search for mutations in NL4.3/SPL2923 in viral genes other than env revealed several mutations in the gene encoding the HIV p17 matrix protein (MA) and one mutation in the gene encoding the p24 capsid protein (CA). In order to analyse the impact of the gag mutations alone and in combination with the mutations in env on the phenotypic resistance towards SPL2923, we developed a novel p17- and p17/gp160-CVT. Phenotypic analysis of the NL4.3/SPL2923 p17- and p17/gp160-recombined strains indicated that the mutations in both env and gag have to be present to fully reproduce the resistance of NL4.3/SPL2923 towards SPL2923.

  3. TrafficGen Architecture Document

    DTIC Science & Technology

    2016-01-01

    distribution unlimited. 2 Fig. 1 TrafficGen user interface The TrafficGen user’s guide details specific features and their use. TrafficGen is available...model is the foundation for the timeline-based user interface and for generating commands for integration with external applications. 3. User... Interface The user interface of this application is comprised of several sets of MVC classes and other support classes. Working together, they present

  4. [VLP vaccines and effects of HIV-1 Env protein modifications on their antigenic properties].

    PubMed

    Vzorov, A N; Compans, R W

    2016-01-01

    An ideal protective HIV-1 vaccine can elicit broadly neutralizing antibodies, capable of preventing HIV transmission. The strategies of designing vaccines include generation of soluble recombinant proteins which mimic the native Env complex and are able to enhance the immunogenicity of gp120. Recent data indicate that the cytoplasmic tail (CT) of the Env protein has multiple functions, which can affect the early steps of infection, as well as viral assembly and antigenic properties. Modifications in the CT can be used to induce conformational changes in functional regions of gp120 and to stabilize the trimeric structure, avoiding immune misdirection and induction of non-neutralizing antibody responses. Env-trimers with modified CTs in virus-like particles (VLPs) are able to induce antibodies with broad spectrum neutralizing activity and high avidity and have the potential for developing an effective vaccine against HIV.

  5. env-encoded residues are not required for transformation by p48v-myb.

    PubMed Central

    Lipsick, J S; Ibanez, C E

    1987-01-01

    The v-myb oncogene of avian myeloblastosis virus induces acute myeloblastic leukemia in chickens and transforms avian myeloid cells in vitro. The protein product of this oncogene, p48v-myb, is partially encoded by the retroviral gag and env genes. We demonstrated that the env-encoded carboxyl terminus of p48v-myb is not required for transformation. Our results showed, in addition, that a coding region of c-myb which is not essential for transformation was transduced by avian myeloblastosis virus. Images PMID:3027417

  6. Direct evidence for intracellular anterograde co-transport of M-PMV Gag and Env on microtubules

    PubMed Central

    Pereira, Lara E.; Clark, Jasmine; Grznarova, Petra; Wen, Xiaoyun; LaCasse, Rachel; Ruml, Tomas; Spearman, Paul

    2014-01-01

    The intracellular transport of Mason-Pfizer monkey virus (M-PMV) assembled capsids from the pericentriolar region to the plasma membrane (PM) requires trafficking of envelope glycoprotein (Env) to the assembly site via the recycling endosome. However, it is unclear if Env-containing vesicles play a direct role in trafficking capsids to the PM. Using live cell microscopy, we demonstrate, for the first time, anterograde co-transport of Gag and Env. Nocodazole disruption of microtubules had differential effects on Gag and Env trafficking, with pulse-chase assays showing a delayed release of Env-deficient virions. Particle tracking demonstrated an initial loss of linear movement of GFP-tagged capsids and mCherry-tagged Env, followed by renewed movement of Gag but not Env at 4 h post-treatment. Thus, while delayed capsid trafficking can occur in the absence of microtubules, efficient anterograde transport of capsids appears to be mediated by microtubule-associated Env-containing vesicles. PMID:24418544

  7. Reclassification of Eubacterium rectale (Prévot et al., 1967) in a new genus Agathobacter gen. nov., as Agathobacter rectalis comb. nov., within the family Lachnospiraceae, and description of Agathobacter ruminis sp. nov., from the rumen.

    PubMed

    Rosero, Jaime A; Killer, Jiří; Sechovcová, Hana; Mrázek, Jakub; Benada, Oldřich; Fliegerová, Kateřina; Havlík, Jaroslav; Kopečný, Jan

    2015-11-30

    Three strains of a Gram-positive, butyrate producing bacteria were isolated from the rumen content of grazing sheep and cow. The strains were anaerobic, Gram-positive cell wall, straight to slightly curved rod-shaped, non-spore-forming and single flagellate. C14:1, C14:0, C16:0 and C16:1 were the predominant fatty acids. The type of cell-wall peptidoglycan is A1γ. The DNA G+C content varied from 41.4 to 42.2 mol%. The 16S rRNA gene sequence similarities between the isolates and Eubacterium rectale, Roseburia hominis and Roseburia intestinalis were found to be 96, 95 and 95%, respectively. The phylogenetic tree showed that the strains constituted a different taxon, separate from other taxa with validly published names and forming a cluster with strains of Eubacterium rectale. On the basis of phenotypic, chemotaxonomic and phylogenetic results (16S RNA, DnaK, GroEL, atpA genes), isolates are considered to represent a novel species of a novel genus of the family Lachnospiraceae for which we propose the name Agathobacter ruminis gen. nov., sp. nov. with the type strain Agathobacter ruminis JK623T (=DSM 29029T =LMG 28559T). We also propose the transfer of Eubacterium rectale to the new genus Agathobacter gen. nov. This genus represents saccharoclastic chemoorganotrophic non-spore forming rods, with Gram-positive membrane, obligatory anaerobic. Main fermentation products on PYG medium were butyrate, acetate, hydrogen and lactate. Peptidoglycan in all species is of A1γ type. Type species is Agathobacter rectalis, gen. nov., comb nov. (Egghert 1935) with type strain ATCC 33656T (==KCTC 5835T). Two species of the new genus, Agathobacter rectalis and Agathobacter ruminis has been defined.

  8. Repeated Vaccination of Cows with HIV Env gp140 during Subsequent Pregnancies Elicits and Sustains an Enduring Strong Env-Binding and Neutralising Antibody Response

    PubMed Central

    Center, Rob J.; Gonelli, Christopher; Muller, Brian; Mackenzie, Charlene; Khoury, Georges; Lichtfuss, Marit; Rawlin, Grant; Purcell, Damian F. J.

    2016-01-01

    An important feature of a potential vaccine against HIV is the production of broadly neutralising antibodies (BrNAbs) capable of potentially blocking infectivity of a diverse array of HIV strains. BrNAbs naturally arise in some HIV infected individuals after several years of infection and their serum IgG can neutralise various HIV strains across different subtypes. We previously showed that vaccination of cows with HIV gp140 AD8 trimers resulted in a high titre of serum IgG against HIV envelope (Env) that had strong BrNAb activity. These polyclonal BrNAbs concentrated into the colostrum during the late stage of pregnancy and can be harvested in vast quantities immediately after calving. In this study, we investigated the effect of prolonged HIV gp140 vaccination on bovine colostrum IgG HIV Env-binding and BrNAb activity over subsequent pregnancies. Repeated immunisation led to a maintained high titre of HIV Env specific IgG in the colostrum batches, but this did not increase through repeated cycles. Colostrum IgG from all batches also strongly competed with sCD4 binding to gp140 Env trimer and with human-derived monoclonal VRC01 and b12 BrNAbs that bind the CD4 binding site (CD4bs). Furthermore, competition neutralisation assays using RSC3 Env gp120 protein core and a derivative CD4bs mutant, RSC3 Δ371I/P363N, showed that CD4bs neutralising antibodies contribute to the neutralising activity of all batches of purified bovine colostrum IgG. This result indicates that the high IgG titre/avidity of anti-CD4bs antibodies with BrNAb activity was achieved during the first year of vaccination and was sustained throughout the years of repeated vaccinations in the cow tested. Although IgG of subsequent colostrum batches may have a higher avidity towards the CD4bs, the overall breadth in neutralisation was not enhanced. This implies that the boosting vaccinations over 4 years elicited a polyclonal antibody response that maintained the proportion of both neutralising and non

  9. Repeated Vaccination of Cows with HIV Env gp140 during Subsequent Pregnancies Elicits and Sustains an Enduring Strong Env-Binding and Neutralising Antibody Response.

    PubMed

    Heydarchi, Behnaz; Center, Rob J; Gonelli, Christopher; Muller, Brian; Mackenzie, Charlene; Khoury, Georges; Lichtfuss, Marit; Rawlin, Grant; Purcell, Damian F J

    2016-01-01

    An important feature of a potential vaccine against HIV is the production of broadly neutralising antibodies (BrNAbs) capable of potentially blocking infectivity of a diverse array of HIV strains. BrNAbs naturally arise in some HIV infected individuals after several years of infection and their serum IgG can neutralise various HIV strains across different subtypes. We previously showed that vaccination of cows with HIV gp140 AD8 trimers resulted in a high titre of serum IgG against HIV envelope (Env) that had strong BrNAb activity. These polyclonal BrNAbs concentrated into the colostrum during the late stage of pregnancy and can be harvested in vast quantities immediately after calving. In this study, we investigated the effect of prolonged HIV gp140 vaccination on bovine colostrum IgG HIV Env-binding and BrNAb activity over subsequent pregnancies. Repeated immunisation led to a maintained high titre of HIV Env specific IgG in the colostrum batches, but this did not increase through repeated cycles. Colostrum IgG from all batches also strongly competed with sCD4 binding to gp140 Env trimer and with human-derived monoclonal VRC01 and b12 BrNAbs that bind the CD4 binding site (CD4bs). Furthermore, competition neutralisation assays using RSC3 Env gp120 protein core and a derivative CD4bs mutant, RSC3 Δ371I/P363N, showed that CD4bs neutralising antibodies contribute to the neutralising activity of all batches of purified bovine colostrum IgG. This result indicates that the high IgG titre/avidity of anti-CD4bs antibodies with BrNAb activity was achieved during the first year of vaccination and was sustained throughout the years of repeated vaccinations in the cow tested. Although IgG of subsequent colostrum batches may have a higher avidity towards the CD4bs, the overall breadth in neutralisation was not enhanced. This implies that the boosting vaccinations over 4 years elicited a polyclonal antibody response that maintained the proportion of both neutralising and non

  10. Targeted Isolation of Antibodies Directed against Major Sites of SIV Env Vulnerability

    PubMed Central

    Mason, Rosemarie D.; Welles, Hugh C.; Adams, Cameron; Chakrabarti, Bimal K.; Gorman, Jason; Zhou, Tongqing; Nguyen, Richard; O’Dell, Sijy; Lusvarghi, Sabrina; Bewley, Carole A.; Li, Hui; Shaw, George M.; Sheng, Zizhang; Shapiro, Lawrence; Wyatt, Richard; Kwong, Peter D.; Mascola, John R.; Roederer, Mario

    2016-01-01

    The simian immunodeficiency virus (SIV) challenge model of lentiviral infection is often used as a model to human immunodeficiency virus type 1 (HIV-1) for studying vaccine mediated and immune correlates of protection. However, knowledge of the structure of the SIV envelope (Env) glycoprotein is limited, as is knowledge of binding specificity, function and potential efficacy of SIV antibody responses. In this study we describe the use of a competitive probe binding sort strategy as well as scaffolded probes for targeted isolation of SIV Env-specific monoclonal antibodies (mAbs). We isolated nearly 70 SIV-specific mAbs directed against major sites of SIV Env vulnerability analogous to broadly neutralizing antibody (bnAb) targets of HIV-1, namely, the CD4 binding site (CD4bs), CD4-induced (CD4i)-site, peptide epitopes in variable loops 1, 2 and 3 (V1, V2, V3) and potentially glycan targets of SIV Env. The range of SIV mAbs isolated includes those exhibiting varying degrees of neutralization breadth and potency as well as others that demonstrated binding but not neutralization. Several SIV mAbs displayed broad and potent neutralization of a diverse panel of 20 SIV viral isolates with some also neutralizing HIV-27312A. This extensive panel of SIV mAbs will facilitate more effective use of the SIV non-human primate (NHP) model for understanding the variables in development of a HIV vaccine or immunotherapy. PMID:27064278

  11. Env sequence determinants in CXCR4-using human immunodeficiency virus type-1 subtype C.

    PubMed

    Lin, Nina H; Becerril, Carlos; Giguel, Francoise; Novitsky, Vladimir; Moyo, Sikhulile; Makhema, Joseph; Essex, Myron; Lockman, Shahin; Kuritzkes, Daniel R; Sagar, Manish

    2012-11-25

    HIV-1 subtype C (HIV-1C) CXCR4-using virus is isolated infrequently and is poorly characterized. Understanding HIV-1C env characteristics has implications for the clinical use of antiretrovirals that target viral entry. A total of 209 env clones derived from 10 samples with mixed CCR5-(R5), CXCR4-using (X4) or dual-tropic HIV-1C were phenotyped for coreceptor usage. Intra-patient X4 and R5 variants generally formed distinct monophyletic phylogenetic clusters. X4 compared to R5 envs had significantly greater amino acid variability and insertions, higher net positive charge, fewer glycosylation sites and increased basic amino acid substitutions in the GPGQ crown. Basic amino acid substitution and/or insertion prior to the crown are highly sensitive characteristics for predicting X4 viruses. Chimeric env functional studies suggest that the V3 loop is necessary but often not sufficient to impart CXCR4 utilization. Our studies provide insights into the unique genotypic characteristics of X4 variants in HIV-1C. Copyright © 2012 Elsevier Inc. All rights reserved.

  12. Env sequence determinants in CXCR4-using human immunodeficiency virus type-1 subtype C

    PubMed Central

    Lin, Nina H.; Becerril, Carlos; Giguel, Francoise; Novitsky, Vladimir; Moyo, Sikhulile; Makhema, Joseph; Essex, Myron; Lockman, Shahin; Kuritzkes, Daniel R.; Sagar, Manish

    2012-01-01

    HIV-1 subtype C (HIV-1C) CXCR4-using virus is isolated infrequently and is poorly characterized. Understanding HIV-1C env characteristics has implications for the clinical use of antiretrovirals that target viral entry. A total of 209 env clones derived from 10 samples with mixed CCR5-(R5), CXCR4-using (X4) or dual-tropic HIV-1C were phenotyped for coreceptor usage. Intra-patient X4 and R5 variants generally formed distinct monophyletic phylogenetic clusters. X4 compared to R5 envs had significantly greater amino acid variability and insertions, higher net positive charge, fewer glycosylation sites and increased basic amino acid substitutions in the GPGQ crown. Basic amino acid substitution and/or insertion prior to the crown are highly sensitive characteristics for predicting X4 viruses. Chimeric env functional studies suggest that the V3 loop is necessary but often not sufficient to impart CXCR4 utilization. Our studies provide insights into the unique genotypic characteristics of X4 variants in HIV-1C. PMID:22954962

  13. Native Conformation and Canonical Disulfide Bond Formation Are Interlinked Properties of HIV-1 Env Glycoproteins

    PubMed Central

    Go, Eden P.; Cupo, Albert; Ringe, Rajesh; Pugach, Pavel; Moore, John P.

    2015-01-01

    ABSTRACT We investigated whether there is any association between a native-like conformation and the presence of only the canonical (i.e., native) disulfide bonds in the gp120 subunits of a soluble recombinant human immunodeficiency virus type 1 (HIV-1) envelope (Env) glycoprotein. We used a mass spectrometry (MS)-based method to map the disulfide bonds present in nonnative uncleaved gp140 proteins and native-like SOSIP.664 trimers based on the BG505 env gene. Our results show that uncleaved gp140 proteins were not homogeneous, in that substantial subpopulations (20 to 80%) contained aberrant disulfide bonds. In contrast, the gp120 subunits of the native-like SOSIP.664 trimer almost exclusively retained the canonical disulfide bond pattern. We also observed that the purification method could influence the proportion of an Env protein population that contained aberrant disulfide bonds. We infer that gp140 proteins may always contain a variable but substantial proportion of aberrant disulfide bonds but that the impact of this problem can be minimized via design and/or purification strategies that yield native-like trimers. The same factors may also be relevant to the production and purification of monomeric gp120 proteins that are free of aberrant disulfide bonds. IMPORTANCE It is widely thought that a successful HIV-1 vaccine will include a recombinant form of the Env protein, a trimer located on the virion surface. To increase yield and simplify purification, Env proteins are often made in truncated, soluble forms. A consequence, however, can be the loss of the native conformation concomitant with the virion-associated trimer. Moreover, some soluble recombinant Env proteins contain aberrant disulfide bonds that are not expected to be present in the native trimer. To assess whether these observations are linked, to determine the extent of disulfide bond scrambling, and to understand why scrambling occurs, we determined the disulfide bond profiles of two soluble Env

  14. Antigenic properties of a transport-competent influenza HA/HIV Env chimeric protein

    SciTech Connect

    Ye Ling; Sun Yuliang; Lin Jianguo; Bu Zhigao; Wu Qingyang; Jiang, Shibo; Steinhauer, David A.; Compans, Richard W.; Yang Chinglai . E-mail: chyang@emory.edu

    2006-08-15

    The transmembrane subunit (gp41) of the HIV Env glycoprotein contains conserved neutralizing epitopes which are not well-exposed in wild-type HIV Env proteins. To enhance the exposure of these epitopes, a chimeric protein, HA/gp41, in which the gp41 of HIV-1 89.6 envelope protein was fused to the C-terminus of the HA1 subunit of the influenza HA protein, was constructed. Characterization of protein expression showed that the HA/gp41 chimeric proteins were expressed on cell surfaces and formed trimeric oligomers, as found in the HIV Env as well as influenza HA proteins. In addition, the HA/gp41 chimeric protein expressed on the cell surface can also be cleaved into 2 subunits by trypsin treatment, similar to the influenza HA. Moreover, the HA/gp41 chimeric protein was found to maintain a pre-fusion conformation. Interestingly, the HA/gp41 chimeric proteins on cell surfaces exhibited increased reactivity to monoclonal antibodies against the HIV Env gp41 subunit compared with the HIV-1 envelope protein, including the two broadly neutralizing monoclonal antibodies 2F5 and 4E10. Immunization of mice with a DNA vaccine expressing the HA/gp41 chimeric protein induced antibodies against the HIV gp41 protein and these antibodies exhibit neutralizing activity against infection by an HIV SF162 pseudovirus. These results demonstrate that the construction of such chimeric proteins can provide enhanced exposure of conserved epitopes in the HIV Env gp41 and may represent a novel vaccine design strategy for inducing broadly neutralizing antibodies against HIV.

  15. Env7p Associates with the Golgin Protein Imh1 at the trans-Golgi Network in Candida albicans

    PubMed Central

    Rao, Kongara Hanumantha; Ghosh, Swagata

    2016-01-01

    ABSTRACT Vesicular dynamics is one of the very important aspects of cellular physiology, an imbalance of which leads to the disorders or diseases in higher eukaryotes. We report the functional characterization of a palmitoylated protein kinase from Candida albicans whose homologue in Saccharomyces cerevisiae has been reported to be involved in negative regulation of membrane fusion and was named Env7. However, the downstream target of this protein remains to be identified. Env7 in C. albicans (CaEnv7) could be isolated from the membrane fraction and localized to vesicular structures associated with the Golgi apparatus. Our work reports Env7 in C. albicans as a new player involved in maintaining the functional dynamics at the trans-Golgi network (TGN) by interacting with two other TGN-resident proteins, namely, Imh1p and Arl1p. Direct interaction could be detected between Env7p and the golgin protein Imh1p. Env7 is itself phosphorylated (Env7p) and phosphorylates Imh1 in vivo. An interaction between Env7 and Imh1 is required for the targeted localization of Imh1. CaEnv7 has a putative palmitoylation site toward both N and C termini. An N-terminal palmitoylation-defective strain retains its ability to phosphorylate Imh1 in vitro. An ENV7 homozygous mutant showed compromised filamentation in solid media and attenuated virulence, whereas an overexpressed strain affected cell wall integrity. Thus, Env7 plays a subtle but important role at the level of multitier regulation that exists at the TGN. IMPORTANCE A multitier regulation exists at the trans-Golgi network in all higher organisms. We report a palmitoylated protein kinase, Env7, that functions at the TGN interface by interacting with two more TGN-resident proteins, namely, Imh1 and Arl1. Palmitoylation seems to be important for the specific localization. This study focuses on the involvement of a ubiquitous protein kinase, whose substrates had not yet been reported from any organism, as an upstream signaling

  16. Reclassification of Leifsonia ginsengi (Qiu et al. 2007) as Herbiconiux ginsengi gen. nov., comb. nov. and description of Herbiconiux solani sp. nov., an actinobacterium associated with the phyllosphere of Solanum tuberosum L.

    PubMed

    Behrendt, Undine; Schumann, Peter; Hamada, Moriyuki; Suzuki, Ken-Ichiro; Spröer, Cathrin; Ulrich, Andreas

    2011-05-01

    In the context of studying the effects of transgenic fructan-producing potatoes on the community structure of phyllosphere bacteria, a group of strains closely related to the species Leifsonia ginsengi was isolated. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the new isolates and L. ginsengi DSM 19088(T) formed a lineage at the genus level and this finding was supported by chemotaxonomic characterization. The peptidoglycan type of the representative isolate, K134/01(T), and L. ginsengi DSM 19088(T) was B2γ, with d- and l-diaminobutyric acid as the diagnostic diamino acid and glycine, alanine and threo-3-hydroxyglutamic acid. The almost-complete substitution of glutamic acid by threo-3-hydroxyglutamic acid supported the differentiation of the new strains from recognized species of the genus Leifsonia. Furthermore, the detection of substantial amounts of the fatty acid cyclohexyl-C(17 : 0) in the new isolates and L. ginsengi DSM 19088(T) was a prominent chemotaxonomic feature for a clear demarcation of these strains from all genera of the family Microbacteriaceae that display the B2γ cell-wall type. Comparative phylogenetic and phenotypic analyses of the isolates and L. ginsengi DSM 19088(T) revealed the separate species status of the isolates. On the basis of these results, it is proposed that L. ginsengi should be classified as the type species of a novel genus, Herbiconiux gen. nov., with the name Herbiconiux ginsengi gen. nov., comb. nov. (type strain wged11(T) = CGMCC 4.3491(T) = JCM 13908(T) = DSM 19088(T) = NBRC 104580(T)). The phyllosphere isolates are assigned to a novel species, Herbiconiux solani sp. nov. (type strain K134/01(T) = DSM 19813(T) = LMG 24387(T) = NBRC 106740(T)).

  17. Sequencing of Gag/Env association with HIV genotyping resolution and HIV-related epidemiologic studies of HIV in China.

    PubMed

    Ren, L; Wang, H W; Xu, Y; Feng, Y; Zhang, H F; Wang, K H

    2016-10-24

    HIV genotyping has led to conflicting results between laboratories. Therefore, identifying the most accurate gene combinations to sequence remains a priority. Datasets of Chinese HIV subtypes based on several markers and deposited in PubMed, Metstr, CNKI, and VIP databases between 2000 and 2015 were studied. In total, 9177 cases of amplification-positive samples from 26 provinces of China were collected and used to classify HIV subtypes based on eight individual genes or a combination thereof. CRF01_AE, CRF07_BC, CRF08_BC and B were the prevalent HIV subtypes in China, accounting for 84.07% of all genotypes. Gag/Env sequencing classified a greater number of HIV subtypes compared to other genes or combination of gene fragments. The geographical distribution of Gag and Gag/Env genotypes was similar to that observed with all genetic markers. Further principal component analysis showed a significantly different geographical distribution pattern of HIV in China for HIV genotypes detected with Gag/Env, which was in line with the distribution of all HIV genotypes in China. Gag/Env sequences had the highest diversity of the eight markers studied, followed by Gag and Gag/Pol/Env; Pol/Env polymorphisms were the least divergent. Gag/Env can serve as a high-resolution marker for HIV genotyping.

  18. The inner membrane histidine kinase EnvZ senses osmolality via helix-coil transitions in the cytoplasm.

    PubMed

    Wang, Loo Chien; Morgan, Leslie K; Godakumbura, Pahan; Kenney, Linda J; Anand, Ganesh S

    2012-05-30

    Two-component systems mediate bacterial signal transduction, employing a membrane sensor kinase and a cytoplasmic response regulator (RR). Environmental sensing is typically coupled to gene regulation. Understanding how input stimuli activate kinase autophosphorylation remains obscure. The EnvZ/OmpR system regulates expression of outer membrane proteins in response to osmotic stress. To identify EnvZ conformational changes associated with osmosensing, we used HDXMS to probe the effects of osmolytes (NaCl, sucrose) on the cytoplasmic domain of EnvZ (EnvZ(c)). Increasing osmolality decreased deuterium exchange localized to the four-helix bundle containing the autophosphorylation site (His(243)). EnvZ(c) exists as an ensemble of multiple conformations and osmolytes favoured increased helicity. High osmolality increased autophosphorylation of His(243), suggesting that these two events are linked. In-vivo analysis showed that the cytoplasmic domain of EnvZ was sufficient for osmosensing, transmembrane domains were not required. Our results challenge existing claims of robustness in EnvZ/OmpR and support a model where osmolytes promote intrahelical H-bonding enhancing helix stabilization, increasing autophosphorylation and downstream signalling. The model provides a conserved mechanism for signalling proteins that respond to diverse physical and mechanical stimuli.

  19. Highly complex neutralization determinants on a monophyletic lineage of newly transmitted subtype C HIV-1 Env clones from India

    PubMed Central

    Kulkarni, Smita S.; Lapedes, Alan; Tang, Haili; Gnanakaran, S.; Daniels, Marcus G.; Zhang, Ming; Bhattacharya, Tanmoy; Li, Ming; Polonis, Victoria R.; McCutchan, Francine E.; Morris, Lynn; Ellenberger, Dennis; Butera, Salvatore T.; Bollinger, Robert C.; Korber, Bette T.; Paranjape, Ramesh S.; Montefiori, David C.

    2009-01-01

    Little is known about the neutralization properties of HIV-1 in India to optimally design and test vaccines. For this reason, a functional Env clone was obtained from each of ten newly acquired, heterosexually transmitted HIV-1 infections in Pune, Maharashtra. These clones formed a phylogenetically distinct genetic lineage within subtype C. As Env-pseudotyped viruses the clones were mostly resistant to IgG1b12, 2G12 and 2F5 but all were sensitive to 4E10. When compared to a large multi-subtype panel of Env-pseudotyped viruses (subtypes B, C and CRF02_AG) in neutralization assays with a multi-subtype panel of HIV-1-positive plasma samples, the Indian Envs were remarkably complex antigenically. With the exception of the Indian Envs, results of a hierarchical clustering analysis showed a strong subtype association with the patterns of neutralization susceptibility. From these patterns we were able to identify 19 neutralization cluster-associated amino acid signatures in gp120 and 14 signatures in the ectodomain and cytoplasmic tail of gp41. We conclude that newly transmitted Indian Envs are antigenically complex in spite of close genetic similarity. Delineation of neutralization-associated amino acid signatures provides a deeper understanding of the antigenic structure of HIV-1 Env. PMID:19167740

  20. Using GenCade to Create a Sediment Budget in SBAS

    DTIC Science & Technology

    2015-08-01

    change and sand transport model (Frey et al. 2012a), and the Sediment Budget Analysis System (SBAS) (Dopsovic et al. 2003). Two of the main GenCade...outputs are shoreline change and longshore transport rates, which can be analyzed to develop a sediment budget. A new, automated connection utilizes the...for those who have experience with GenCade and have at least a conceptual knowledge of how to construct a sediment budget. INTRODUCTION: Developed

  1. Panels of HIV-1 Subtype C Env Reference Strains for Standardized Neutralization Assessments

    PubMed Central

    Rademeyer, Cecilia; Williamson, Carolyn; Seaman, Michael S.; Gottardo, Raphael; Tang, Haili; Greene, Kelli; Gao, Hongmei; LaBranche, Celia; Mascola, John R.; Morris, Lynn; Montefiori, David C.

    2017-01-01

    ABSTRACT In the search for effective immunologic interventions to prevent and treat HIV-1 infection, standardized reference reagents are a cost-effective way to maintain robustness and reproducibility among immunological assays. To support planned and ongoing studies where clade C predominates, here we describe three virus panels, chosen from 200 well-characterized clade C envelope (Env)-pseudotyped viruses from early infection. All 200 Envs were expressed as a single round of replication pseudoviruses and were tested to quantify neutralization titers by 16 broadly neutralizing antibodies (bnAbs) and sera from 30 subjects with chronic clade C infections. We selected large panels of 50 and 100 Envs either to characterize cross-reactive breadth for sera identified as having potent neutralization activity based on initial screening or to evaluate neutralization magnitude-breadth distributions of newly isolated antibodies. We identified these panels by downselection after hierarchical clustering of bnAb neutralization titers. The resulting panels represent the diversity of neutralization profiles throughout the range of virus sensitivities identified in the original panel of 200 viruses. A small 12-Env panel was chosen to screen sera from vaccine trials or natural-infection studies for neutralization responses. We considered panels selected by previously described methods but favored a computationally informed method that enabled selection of viruses representing diverse neutralization sensitivity patterns, given that we do not a priori know what the neutralization-response profile of vaccine sera will be relative to that of sera from infected individuals. The resulting 12-Env panel complements existing panels. Use of standardized panels enables direct comparisons of data from different trials and study sites testing HIV-1 clade C-specific products. IMPORTANCE HIV-1 group M includes nine clades and many recombinants. Clade C is the most common lineage, responsible for

  2. Virion Background and Efficiency of Virion Incorporation Determine Susceptibility of Simian Immunodeficiency Virus Env-Driven Viral Entry to Inhibition by IFITM Proteins.

    PubMed

    Wrensch, Florian; Hoffmann, Markus; Gärtner, Sabine; Nehlmeier, Inga; Winkler, Michael; Pöhlmann, Stefan

    2017-01-15

    Interferon-induced transmembrane proteins (IFITMs) can inhibit the cellular entry of several enveloped viruses, including simian immunodeficiency virus (SIV). The blockade of SIV by IFITMs is isolate specific, raising the question of which parameters impact sensitivity to IFITM. We show that the virion context in which SIV-Env is presented and the efficiency of virion incorporation determine Env susceptibility to inhibition by IFITMs. Thus, determinants other than the nature of the envelope protein can impact the IFITM sensitivity of viral entry. The host cell-encoded IFITM proteins can block viral entry and are an important component of the innate defenses against viral infection. However, the determinants controlling whether a virus is susceptible to blockade by IFITM proteins are incompletely understood. Our study shows that the amount of envelope proteins incorporated into virions as well as the nature of the virion particle itself can impact the sensitivity of viral entry to IFITMs. These results show for the first time that determinants other than the viral envelope protein can impact sensitivity to IFITM and have implications for the interpretation of previously published data on inhibition of viruses by IFITM proteins. Moreover, our findings might help to define the mechanism underlying the antiviral activity of IFITM proteins. Copyright © 2017 Wrensch et al.

  3. Analysis of the human env-specific cytotoxic T-lymphocyte (CTL) response in natural human immunodeficiency virus type 1 infection: low prevalence of broadly cross-reactive env-specific CTL.

    PubMed Central

    Carmichael, A; Jin, X; Sissons, P

    1996-01-01

    Major histocompatibility complex-restricted cytotoxic T lymphocytes (CTL) are part of the cellular immune response to persistent virus infections. Candidate vaccines against human immunodeficiency virus type 1 (HIV-1) should elicit broad cross-reactive immunity to confer protection against different strains of HIV-1. As it is likely that candidate vaccines will include the envelope gene product Env, we determined the proportion of CTL clones which recognized variable and conserved determinants in three env variants during natural infection. Limiting dilution analysis was used to characterize numerous short-term CTL clones derived from peripheral blood of HIV-1-infected subjects, using split-well analysis to assay cytotoxicity against target cells expressing gp160env of HIV-1 strains IIIB, MN, and RF. In 9 of 12 HIV-1-infected subjects, at the clonal level most env-specific CTL recognized determinant(s) within one env variant but not in the other variants. In some subjects, CTL recognized multiple nonconserved determinants in different variants. The pattern of recognition of different env variants was relatively stable over time. In most of the patients studied, the proportion of CTL which showed cross-recognition of conserved determinants shared among the three strains was low. Two novel CTL epitopes within gp41 were identified by using 15-mer peptides of the HIV-SF2 sequence. When specific peptide was used to stimulate CTL precursors in vitro, the frequency of peptide-specific CTL precursors was very high, but the CTL elicited by this stimulation were highly strain specific. We conclude that the use of a single HIV env variant to detect CTL activity can underestimate the magnitude and complexity of the env-specific CTL response. The low prevalence of CTL clones which show cross-recognition of conserved determinants may have implications for immunization strategies based solely on env; to elicit broadly cross-reactive CTL other, more conserved viral antigens are

  4. Halotalea alkalilenta gen. nov., sp. nov., a novel osmotolerant and alkalitolerant bacterium from alkaline olive mill wastes, and emended description of the family Halomonadaceae Franzmann et al. 1989, emend. Dobson and Franzmann 1996.

    PubMed

    Ntougias, Spyridon; Zervakis, Georgios I; Fasseas, Constantinos

    2007-09-01

    A novel Gram-negative, motile, nonsporulating, rod-shaped bacterium isolated from alkaline sludge-like wastes ('alpeorujo' or 'alperujo') of two-phase olive oil extraction is described. The strain, designated AW-7(T), is an obligate aerobe that is halotolerant (tolerating up to 15 % w/v NaCl), sugar-tolerant (tolerating up to 45 % and 60 % w/v (+)-d-glucose and maltose respectively; these are the highest concentrations tolerated by any known members of the Bacteria domain) and alkalitolerant (growing at a broad pH range of 5-11). Strain AW-7(T) is chemo-organotrophic. Ubiquinone-9 was detected in the respiratory chain of strain AW-7(T). The major fatty acids present are C(18 : 1)omega7c, C(16 : 0), C(19 : 0) cyclo omega8c, C(12 : 0) 3-OH and C(16 : 1)omega7c/iso-C(15 : 0) 2-OH. Based on 16S rRNA gene sequence analysis, strain AW-7(T) showed almost equal phylogenetic distances from Zymobacter palmae (95.6 % similarity) and Carnimonas nigrificans (95.4 % similarity). In addition, low DNA-DNA relatedness values were found for strain AW-7(T) against Carnimonas nigrificans CECT 4437(T) (22.5-25.4 %) and Z. palmae DSM 10491(T) (11.9-14.4 %). The DNA G+C content of strain AW-7(T) is 64.4 mol%. Physiological and chemotaxonomic data further confirmed the differentiation of strain AW-7(T) from the genera Zymobacter and Carnimonas. Thus, strain AW-7(T) represents a novel bacterial genus within the family Halomonadaceae, for which the name Halotalea gen. nov. is proposed. Halotalea alkalilenta sp. nov. (type strain AW-7(T)=DSM 17697(T)=CECT 7134(T)) is proposed as the type species of the genus Halotalea gen. nov. A reassignment of the descriptive 16S rRNA signature characteristics of the family Halomonadaceae permitted the placement of the novel genus Halotalea into the family; in contrast, the genus Halovibrio possessed only 12 out of the 18 signature characteristics proposed, and hence it was excluded from the family Halomonadaceae.

  5. Crystallographic snapshot of the Escherichia coli EnvZ histidine kinase in an active conformation.

    PubMed

    Ferris, Hedda U; Coles, Murray; Lupas, Andrei N; Hartmann, Marcus D

    2014-06-01

    Sensor histidine kinases are important sensors of the extracellular environment and relay signals via conformational changes that trigger autophosphorylation of the kinase and subsequent phosphorylation of a response regulator. The exact mechanism and the regulation of this protein family are a matter of ongoing investigation. Here we present a crystal structure of a functional chimeric protein encompassing the entire catalytic part of the Escherichia coli EnvZ histidine kinase, fused to the HAMP domain of the Archaeoglobus fulgidus Af1503 receptor. The construct is thus equivalent to the full cytosolic part of EnvZ. The structure shows a putatively active conformation of the catalytic domain and gives insight into how this conformation could be brought about in response to sensory input. Our analysis suggests a sequential flip-flop autokinase mechanism.

  6. Comprehensive Characterization of the Transmitted/Founder env Genes From a Single MSM Cohort in China.

    PubMed

    Chen, Yue; Li, Ning; Zhang, Tong; Huang, Xiaojie; Cai, Fangping; Vandergrift, Nathan; Xin, Ruolei; Meng, Zhefeng; Zhang, Xiaoyan; Jiang, Chunlai; Xu, Xiaoning; Montefiori, David C; Gao, Feng; Wu, Hao

    2015-08-01

    The men having sex with men (MSM) population has become one of the major risk groups for HIV-1 infection in China. However, the epidemiological patterns, function of the env genes, and autologous and heterologous neutralization activity in the same MSM population have not been systematically characterized. The env gene sequences were obtained by the single genome amplification. The time to the most recent common ancestor was estimated for each genotype using the Bayesian Markov Chain Monte Carlo approach. Coreceptor usage was determined in NP-2 cells. Neutralization was analyzed using Env pseudoviruses in TZM-bl cells. We have obtained 547 full-length env gene sequences by single genome amplification from 30 acute/early HIV-1--infected individuals in the Beijing MSM cohort. Three genotypes (subtype B, CRF01_AE, and CRF07_BC) were identified and 20% of the individuals were infected with multiple transmitted/founder (T/F) viruses. The tight clusters of the MSM sequences regardless of geographic origins indicated nearly exclusive transmission within the MSM population and limited number of introductions. The time to the most recent common ancestor for each genotype was 10-15 years after each was first introduced in China. Disparate preferences for coreceptor usages among 3 genotypes might lead to the changes in percentage of different genotypes in the MSM population over time. The genotype-matched and genotype-mismatched neutralization activity varied among the 3 genotypes. The identification of unique characteristics for transmission, coreceptor usage, neutralization profile, and epidemic patterns of HIV-1 is critical for the better understanding of transmission mechanisms, development of preventive strategies, and evaluation of vaccine efficacy in the MSM population in China.

  7. EnvMine: a text-mining system for the automatic extraction of contextual information.

    PubMed

    Tamames, Javier; de Lorenzo, Victor

    2010-06-01

    For ecological studies, it is crucial to count on adequate descriptions of the environments and samples being studied. Such a description must be done in terms of their physicochemical characteristics, allowing a direct comparison between different environments that would be difficult to do otherwise. Also the characterization must include the precise geographical location, to make possible the study of geographical distributions and biogeographical patterns. Currently, there is no schema for annotating these environmental features, and these data have to be extracted from textual sources (published articles). So far, this had to be performed by manual inspection of the corresponding documents. To facilitate this task, we have developed EnvMine, a set of text-mining tools devoted to retrieve contextual information (physicochemical variables and geographical locations) from textual sources of any kind. EnvMine is capable of retrieving the physicochemical variables cited in the text, by means of the accurate identification of their associated units of measurement. In this task, the system achieves a recall (percentage of items retrieved) of 92% with less than 1% error. Also a Bayesian classifier was tested for distinguishing parts of the text describing environmental characteristics from others dealing with, for instance, experimental settings.Regarding the identification of geographical locations, the system takes advantage of existing databases such as GeoNames to achieve 86% recall with 92% precision. The identification of a location includes also the determination of its exact coordinates (latitude and longitude), thus allowing the calculation of distance between the individual locations. EnvMine is a very efficient method for extracting contextual information from different text sources, like published articles or web pages. This tool can help in determining the precise location and physicochemical variables of sampling sites, thus facilitating the performance

  8. EnvMine: A text-mining system for the automatic extraction of contextual information

    PubMed Central

    2010-01-01

    Background For ecological studies, it is crucial to count on adequate descriptions of the environments and samples being studied. Such a description must be done in terms of their physicochemical characteristics, allowing a direct comparison between different environments that would be difficult to do otherwise. Also the characterization must include the precise geographical location, to make possible the study of geographical distributions and biogeographical patterns. Currently, there is no schema for annotating these environmental features, and these data have to be extracted from textual sources (published articles). So far, this had to be performed by manual inspection of the corresponding documents. To facilitate this task, we have developed EnvMine, a set of text-mining tools devoted to retrieve contextual information (physicochemical variables and geographical locations) from textual sources of any kind. Results EnvMine is capable of retrieving the physicochemical variables cited in the text, by means of the accurate identification of their associated units of measurement. In this task, the system achieves a recall (percentage of items retrieved) of 92% with less than 1% error. Also a Bayesian classifier was tested for distinguishing parts of the text describing environmental characteristics from others dealing with, for instance, experimental settings. Regarding the identification of geographical locations, the system takes advantage of existing databases such as GeoNames to achieve 86% recall with 92% precision. The identification of a location includes also the determination of its exact coordinates (latitude and longitude), thus allowing the calculation of distance between the individual locations. Conclusion EnvMine is a very efficient method for extracting contextual information from different text sources, like published articles or web pages. This tool can help in determining the precise location and physicochemical variables of sampling sites, thus

  9. Quaternary structures of HIV Env immunogen exhibit conformational vicissitudes and interface diminution elicited by ligand binding

    PubMed Central

    Moscoso, Carlos G.; Sun, Yide; Poon, Selina; Xing, Li; Kan, Elaine; Martin, Loïc; Green, Dominik; Lin, Frank; Vahlne, Anders G.; Barnett, Susan; Srivastava, Indresh; Cheng, R. Holland

    2011-01-01

    The human immunodeficiency virus envelope protein is the key element mediating entry into host cells. Conformational rearrangement of Env upon binding to the host CD4 receptor and chemokine coreceptor drives membrane fusion. We elucidated the quaternary arrangement of the soluble Env trimeric immunogen o-gp140ΔV2TV1, in both its native (unliganded) and CD4-induced (liganded) states by cryoelectron microscopy and molecular modeling. The liganded conformation was elicited by binding gp140 to the synthetic CD4-mimicking miniprotein CD4m. Upon CD4m binding, an outward domain shift of the three gp120 subunits diminishes gp120–gp41 interactions, whereas a “flat open” concave trimer apex is observed consequent to gp120 tilting away from threefold axis, likely juxtaposing the fusion peptide with the host membrane. Additional features observed in the liganded conformation include rotations of individual gp120 subunits that may release gp41 for N- and C-helix refolding and also may lead to optimal exposure of the elicited coreceptor binding site. Such quaternary arrangements of gp140 lead to the metastable liganded conformation, with putative locations of exposed epitopes contributing to a description of sequential events occurring prior to membrane fusion. Our observations imply a mechanism whereby a soluble Env trimeric construct, as opposed to trimers extracted from virions, may better expose crucial epitopes such as the CD4 binding site and V3, as well as epitopes in the vicinity of gp41, subsequent to conjugation with CD4m. Structural features gleaned from our studies should aid the design of Env-based immunogens for inducement of potent broadly neutralizing antibodies against exposed conformational epitopes. PMID:21444771

  10. Comprehensive Characterization of the Transmitted/founder env Genes from a Single MSM Cohort in China

    PubMed Central

    Chen, Yue; Li, Ning; Zhang, Tong; Huang, Xiaojie; Cai, Fangping; Vandergrift, Nathan; Xin, Ruolei; Meng, Zhefeng; Zhang, Xiaoyan; Jiang, Chunlai; Xu, Xiaoning; Montefiori, David C; Gao, Feng; Wu, Hao

    2015-01-01

    Background The men having sex with men (MSM) population has become one of major risk groups for HIV-1 infection in China. However, the epidemiological patterns, function of the env genes, and autologous and heterologous neutralization activity in the same MSM population have not been systematically characterized. Methods The env gene sequences were obtained by the single genome amplification (SGA). The time to the most recent common ancestor (tMRCA) was estimated for each genotype using the Bayesian MCMC approach. Coreceptor usage was determined in NP-2 cells. Neutralization was analyzed using Env pseudoviruses in TZM-bl cells. Results We have obtained 547 full-length env gene sequences by SGA from 30 acute/early HIV-1-infected individuals in the Beijing MSM cohort. Three genotypes (Subtype B, CRF01_AE, and CRF07_BC) were identified and 20% of the individuals were infected with multiple transmitted/founder (T/F) viruses. The tight clusters of the MSM sequences regardless of geographic origins indicated nearly exclusive transmission within the MSM population and limited number of introductions. The tMRCA for each genotype was 10-15 years after each was first introduced in China. Disparate preferences for coreceptor usages among three genotypes might lead to the changes in percentage of different genotypes in the MSM population over time. The genotype-matched and -mismatched neutralization activity varied among the three genotypes. Conclusions Identification of unique characteristics for transmission, coreceptor usage, neutralization profile and epidemic patterns of HIV-1 is critical for the better understanding of transmission mechanisms, development of preventive strategies, and evaluation of vaccine efficacy in the MSM population in China. PMID:25886933

  11. Persian adaptation of a questionnaire of environmental risk factors in multiple sclerosis (EnvIMS-Q).

    PubMed

    Sahraian, Mohammad Ali; Naghshineh, Hoda; Shati, Mohsen; Jahromi, Soodeh Razeghi; Rezaei, Niloofar

    2016-11-01

    It seems that gene-environment interaction play most important role in Multiple Sclerosis development. Increasing the incidence and prevalence of MS during the recent decades in the low prevalence area such as Iran is explained better by environment factors. Environmental Risk Factors in Multiple Sclerosis (the 'EnvIMS-Q') is a 6-page self-administered questionnaire for case control studies. the objectives of study are validation and adaptation of the EnvIMS-Q' then development of a Persian version for case control studies in Persian population. This questionnaire translated literally and in culturally relevant form, then content validation process was done by three groups' experts. According to giving rating to each item, each section and the whole instrument, we calculated their content validation indexes and also added some new questions and a new section to EnvIMS-Q. Finally, we analyzed repeatability of the answers within a 4 weeks interval. Relevancy and clarity indexes of all items were more than 80%. Scale relevancy index equaled 99% and scale clarity index equaled 97%. Repeatability of most items was acceptable. the use of standardized validated questionnaires will assist the researchers to perform local studies on the role of environmental factors on the basis of reliable data. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Structure and immune recognition of trimeric pre-fusion HIV-1 Env

    SciTech Connect

    Pancera, Marie; Zhou, Tongqing; Druz, Aliaksandr; Georgiev, Ivelin S.; Soto, Cinque; Gorman, Jason; Huang, Jinghe; Acharya, Priyamvada; Chuang, Gwo-Yu; Ofek, Gilad; Stewart-Jones, Guillaume B. E.; Stuckey, Jonathan; Bailer, Robert T.; Joyce, M. Gordon; Louder, Mark K.; Tumba, Nancy; Yang, Yongping; Zhang, Baoshan; Cohen, Myron S.; Haynes, Barton F.; Morris, Lynn; Munro, James B.; Blanchard, Scott C.; Mothes, Walther; Connors, Mark; Kwong, Peter D.

    2014-10-08

    The human immunodeficiency virus type 1 (HIV-1) envelope (Env) spike, comprising three gp120 and three gp41 subunits, is a conformational machine that facilitates HIV-1 entry by rearranging from a mature unliganded state, through receptor-bound intermediates, to a post-fusion state. As the sole viral antigen on the HIV-1 virion surface, Env is both the target of neutralizing antibodies and a focus of vaccine efforts. Here we report the structure at 3.5 Å resolution for an HIV-1 Env trimer captured in a mature closed state by antibodies PGT122 and 35O22. This structure reveals the pre-fusion conformation of gp41, indicates rearrangements needed for fusion activation, and defines parameters of immune evasion and immune recognition. Pre-fusion gp41 encircles amino- and carboxy-terminal strands of gp120 with four helices that form a membrane-proximal collar, fastened by insertion of a fusion peptide-proximal methionine into a gp41-tryptophan clasp. Spike rearrangements required for entry involve opening the clasp and expelling the termini. In conclusion, N-linked glycosylation and sequence-variable regions cover the pre-fusion closed spike; we used chronic cohorts to map the prevalence and location of effective HIV-1-neutralizing responses, which were distinguished by their recognition of N-linked glycan and tolerance for epitope-sequence variation.

  13. Gene envY of Escherichia coli K-12 affects thermoregulation of major porin expression.

    PubMed Central

    Lundrigan, M D; Earhart, C F

    1984-01-01

    The temperature-dependent expression of OmpF and OmpC, the major channel-forming proteins of the Escherichia coli K-12 outer membrane, was studied. In wild-type cells, decreasing growth temperatures resulted in increased amounts of OmpF protein and correspondingly decreased quantities of OmpC protein. Bacteria deleted for the 13-min chromosomal region did not exhibit this temperature-dependent fluctuation in porin proteins. Plasmid pML22, which consists of pBR322 containing a 0.5-megadalton E. coli chromosomal DNA insert, complemented the thermoregulatory defect. The regulatory gene was named envY. In minicells, pML22 directed the synthesis of an envelope polypeptide (EnvY) having an apparent molecular weight of 25,000. The EnvY protein was synthesized in minicells in greater amounts at 27 degrees C than at 37 degrees C, and a reducing agent was necessary in the solubilization buffer for its subsequent detection on polyacrylamide gels. The results describe the initial characterization of a regulatory system which, along with proteins of the ompB operon, the cyclic AMP system, and the tolC gene product, is involved in a complex network affecting major porin expression. Images PMID:6317653

  14. Antibodies to a conformational epitope on gp41 neutralize HIV-1 by destabilizing the Env spike.

    PubMed

    Lee, Jeong Hyun; Leaman, Daniel P; Kim, Arthur S; Torrents de la Peña, Alba; Sliepen, Kwinten; Yasmeen, Anila; Derking, Ronald; Ramos, Alejandra; de Taeye, Steven W; Ozorowski, Gabriel; Klein, Florian; Burton, Dennis R; Nussenzweig, Michel C; Poignard, Pascal; Moore, John P; Klasse, Per Johan; Sanders, Rogier W; Zwick, Michael B; Wilson, Ian A; Ward, Andrew B

    2015-09-25

    The recent identification of three broadly neutralizing antibodies (bnAbs) against gp120-gp41 interface epitopes has expanded the targetable surface on the HIV-1 envelope glycoprotein (Env) trimer. By using biochemical, biophysical and computational methods, we map the previously unknown trimer epitopes of two related antibodies, 3BC315 and 3BC176. A cryo-EM reconstruction of a soluble Env trimer bound to 3BC315 Fab at 9.3 Å resolution reveals that the antibody binds between two gp41 protomers, and neutralizes the virus by accelerating trimer decay. In contrast, bnAb 35O22 binding to a partially overlapping quaternary epitope at the gp120-gp41 interface does not induce decay. A conserved gp41-proximal glycan at N88 was also shown to play a role in the binding kinetics of 3BC176 and 3BC315. Finally, our data suggest that the dynamic structure of the Env trimer influences exposure of bnAb epitopes.

  15. Antibodies to a conformational epitope on gp41 neutralize HIV-1 by destabilizing the Env spike

    PubMed Central

    Lee, Jeong Hyun; Leaman, Daniel P.; Kim, Arthur S.; Torrents de la Peña, Alba; Sliepen, Kwinten; Yasmeen, Anila; Derking, Ronald; Ramos, Alejandra; de Taeye, Steven W.; Ozorowski, Gabriel; Klein, Florian; Burton, Dennis R.; Nussenzweig, Michel C.; Poignard, Pascal; Moore, John P.; Klasse, Per Johan; Sanders, Rogier W.; Zwick, Michael B.; Wilson, Ian A.; Ward, Andrew B.

    2015-01-01

    The recent identification of three broadly neutralizing antibodies (bnAbs) against gp120–gp41 interface epitopes has expanded the targetable surface on the HIV-1 envelope glycoprotein (Env) trimer. By using biochemical, biophysical and computational methods, we map the previously unknown trimer epitopes of two related antibodies, 3BC315 and 3BC176. A cryo-EM reconstruction of a soluble Env trimer bound to 3BC315 Fab at 9.3 Å resolution reveals that the antibody binds between two gp41 protomers, and neutralizes the virus by accelerating trimer decay. In contrast, bnAb 35O22 binding to a partially overlapping quaternary epitope at the gp120–gp41 interface does not induce decay. A conserved gp41-proximal glycan at N88 was also shown to play a role in the binding kinetics of 3BC176 and 3BC315. Finally, our data suggest that the dynamic structure of the Env trimer influences exposure of bnAb epitopes. PMID:26404402

  16. Structure and immune recognition of trimeric pre-fusion HIV-1 Env

    DOE PAGES

    Pancera, Marie; Zhou, Tongqing; Druz, Aliaksandr; ...

    2014-10-08

    The human immunodeficiency virus type 1 (HIV-1) envelope (Env) spike, comprising three gp120 and three gp41 subunits, is a conformational machine that facilitates HIV-1 entry by rearranging from a mature unliganded state, through receptor-bound intermediates, to a post-fusion state. As the sole viral antigen on the HIV-1 virion surface, Env is both the target of neutralizing antibodies and a focus of vaccine efforts. Here we report the structure at 3.5 Å resolution for an HIV-1 Env trimer captured in a mature closed state by antibodies PGT122 and 35O22. This structure reveals the pre-fusion conformation of gp41, indicates rearrangements needed formore » fusion activation, and defines parameters of immune evasion and immune recognition. Pre-fusion gp41 encircles amino- and carboxy-terminal strands of gp120 with four helices that form a membrane-proximal collar, fastened by insertion of a fusion peptide-proximal methionine into a gp41-tryptophan clasp. Spike rearrangements required for entry involve opening the clasp and expelling the termini. In conclusion, N-linked glycosylation and sequence-variable regions cover the pre-fusion closed spike; we used chronic cohorts to map the prevalence and location of effective HIV-1-neutralizing responses, which were distinguished by their recognition of N-linked glycan and tolerance for epitope-sequence variation.« less

  17. Antibodies to a conformational epitope on gp41 neutralize HIV-1 by destabilizing the Env spike

    NASA Astrophysics Data System (ADS)

    Lee, Jeong Hyun; Leaman, Daniel P.; Kim, Arthur S.; Torrents de La Peña, Alba; Sliepen, Kwinten; Yasmeen, Anila; Derking, Ronald; Ramos, Alejandra; de Taeye, Steven W.; Ozorowski, Gabriel; Klein, Florian; Burton, Dennis R.; Nussenzweig, Michel C.; Poignard, Pascal; Moore, John P.; Klasse, Per Johan; Sanders, Rogier W.; Zwick, Michael B.; Wilson, Ian A.; Ward, Andrew B.

    2015-09-01

    The recent identification of three broadly neutralizing antibodies (bnAbs) against gp120-gp41 interface epitopes has expanded the targetable surface on the HIV-1 envelope glycoprotein (Env) trimer. By using biochemical, biophysical and computational methods, we map the previously unknown trimer epitopes of two related antibodies, 3BC315 and 3BC176. A cryo-EM reconstruction of a soluble Env trimer bound to 3BC315 Fab at 9.3 Å resolution reveals that the antibody binds between two gp41 protomers, and neutralizes the virus by accelerating trimer decay. In contrast, bnAb 35O22 binding to a partially overlapping quaternary epitope at the gp120-gp41 interface does not induce decay. A conserved gp41-proximal glycan at N88 was also shown to play a role in the binding kinetics of 3BC176 and 3BC315. Finally, our data suggest that the dynamic structure of the Env trimer influences exposure of bnAb epitopes.

  18. Aerobic Biodegradation of N-Nitrosodimethylamine by the Propanotroph Rhodococcus ruber ENV425▿

    PubMed Central

    Fournier, Diane; Hawari, Jalal; Halasz, Annamaria; Streger, Sheryl H.; McClay, Kevin R.; Masuda, Hisako; Hatzinger, Paul B.

    2009-01-01

    The propanotroph Rhodococcus ruber ENV425 was observed to rapidly biodegrade N-nitrosodimethylamine (NDMA) after growth on propane, tryptic soy broth, or glucose. The key degradation intermediates were methylamine, nitric oxide, nitrite, nitrate, and formate. Small quantities of formaldehyde and dimethylamine were also detected. A denitrosation reaction, initiated by hydrogen atom abstraction from one of the two methyl groups, is hypothesized to result in the formation of n-methylformaldimine and nitric oxide, the former of which decomposes in water to methylamine and formaldehyde and the latter of which is then oxidized further to nitrite and then nitrate. Although the strain mineralized more than 60% of the carbon in [14C]NDMA to 14CO2, growth of strain ENV425 on NDMA as a sole carbon and energy source could not be confirmed. The bacterium was capable of utilizing NDMA, as well as the degradation intermediates methylamine and nitrate, as sources of nitrogen during growth on propane. In addition, ENV425 reduced environmentally relevant microgram/liter concentrations of NDMA to <2 ng/liter in batch cultures, suggesting that the bacterium may have applications for groundwater remediation. PMID:19542346

  19. Novel Monoclonal Antibody Directed at the Receptor Binding Site on the Avian Sarcoma and Leukosis Virus Env Complex

    PubMed Central

    Ochsenbauer-Jambor, Christina; Delos, Sue E.; Accavitti, Mary Ann; White, Judith M.; Hunter, Eric

    2002-01-01

    We report here on the generation of a mouse monoclonal antibody directed against Rous sarcoma virus (RSV) subgroup A Env that will be useful in functional and structural analysis of RSV Env, as well as in approaches employing the RCAS/Tva system for gene targeting. BALB/c mice were primed and given boosters twice with EnvA-expressing NIH 3T3 cells. Resulting hybridomas were tested by enzyme-linked immunosorbent assay against RCANBP virions and SU-A-immunoglobulin G immunoadhesin. One highly reactive hybridoma clone, mc8C5, was subcloned and tested in immunofluorescence, immunoprecipitation (IP), and Western blotting assays. In all three assays, mc8C5-4 subgroup-specifically recognizes SR-A Env, through the SU domain, expressed from different vectors in both avian and mammalian cells. This multifunctionality is notable for a mouse monoclonal. We furthermore observed a preference for binding to terminally glycosylated Env over core-glycosylated Env precursor in IPs, suggesting that the epitope is at least partially conformational and dependent on glycosylation. Most importantly, we found mc8C5-4 inhibited Env function: in vitro, the monoclonal not only interferes with binding of the EnvA receptor, Tva, but it also blocks the Tva-induced conformational change required for activation of the fusion peptide, without inducing that change itself. Infection of Tva-expressing avian or mammalian cells by avian sarcoma and leukosis virus (ASLV) or EnvA-pseudotyped murine leukemia virus, respectively, is efficiently inhibited by mc8C5-4. The apparent interference of the monoclonal with the EnvA-Tva complex formation suggests that the epitope seen by mc8C5 overlaps with the receptor binding site. This is supported by the observation that mutations of basic residues in hr2 or of the downstream glycosylation site, which both impair Tva-binding to EnvA, have similar effects on the binding of mc8C5. Thus, anti-ASLV-SU-A mc8C5-4 proves to be a unique new immunoreagent that targets

  20. Study of the HIV-2 Env Cytoplasmic Tail Variability and Its Impact on Tat, Rev and Nef

    PubMed Central

    Bakouche, Nordine; Vandenbroucke, Anne-Thérèse; Goubau, Patrick; Ruelle, Jean

    2013-01-01

    Background The HIV-2 env’s 3’ end encodes the cytoplasmic tail (CT) of the Env protein. This genomic region also encodes the rev, Tat and Nef protein in overlapping reading frames. We studied the variability in the CT coding region in 46 clinical specimens and in 2 reference strains by sequencing and by culturing. The aims were to analyse the variability of Env CT and the evolution of proteins expressed from overlapping coding sequences. Results A 70% reduction of the length of the CT region affected the HIV-2 ROD and EHO strains in vitro due to a premature stop codon in the env gene. In clinical samples this wasn’t observed, but the CT length varied due to insertions and deletions. We noted 3 conserved and 3 variable regions in the CT. The conserved regions were those containing residues involved in Env endocytosis, the potential HIV-2 CT region implicated in the NF-kB activation and the potential end of the lentiviral lytic peptide one. The variable regions were the potential HIV-2 Kennedy region, the potential lentiviral lytic peptide two and the beginning of the potential lentiviral lytic peptide one. A very hydrophobic region was coded downstream of the premature stop codon observed in vitro, suggesting a membrane spanning region. Interestingly, the nucleotides that are responsible for the variability of the CT don’t impact rev and Nef. However, in the Kennedy-like coding region variability resulted only from nucleotide changes that impacted Env and Tat together. Conclusion The HIV-2 Env, Tat and Rev C-terminal part are subject to major length variations in both clinical samples and cultured strains. The HIV-2 Env CT contains variable and conserved regions. These regions don’t affect the rev and Nef amino acids composition which evolves independently. In contrast, Tat co-evolves with the Env CT. PMID:24223892

  1. Construction and characterization of the recombinant Moloney murine leukemia viruses bearing the mouse Fv-4 env gene.

    PubMed Central

    Masuda, M; Yoshikura, H

    1990-01-01

    A nucleotide sequence of the mouse Fv-4 env gene was completed. Structural comparison revealed a close relationship of Fv-4 to the ecotropic Cas-Br-E murine leukemia virus isolated from a wild mouse in southern California. Various portions of the env gene of Moloney murine leukemia virus were replaced by the corresponding Fv-4 env sequence to construct recombinant murine leukemia virus clones. Infectivity of these recombinants was checked by the S+L- cell focus induction assay and the XC cell syncytium formation assay. Recombinants bearing the following Fv-4 env sequence retained ecotropic infectivity; the AccI-BamHI and BamHI-BalI regions coding for the N- and C-terminal halves of Fv-4 gp70SU, respectively; and the BalI-NcoI region encoding the cleavage site between gp70SU and p15(E)TM of the Fv-4 env. However, when the Fv-4 sequence was substituted for the p15(E)TM-coding NcoI-EcoRV region or the AccI-EcoRV region covering almost the entire env gene, infectivity was undetectable in our assays. The recombinant clone containing the Fv-4 AccI-EcoRV region, i.e., almost the entire Fv-4 env sequence, was introduced with pSV2neo into NIH 3T3 cells, and a G418r cell line named NIH(Fv4)-2 was isolated. The NIH(Fv4)-2 cell released viral particles that contained reverse transcriptase, Fv-4 env molecules as well as the other viral proteins, and viral genomic RNA. However, proviral DNA synthesis was not detected upon inoculation of this virus in NIH 3T3 cells. The loss of infectivity of the recombinant virus bearing the Fv-4 AccI-EcoRV region appeared to be caused by failure in an early step of replication. Images PMID:2304138

  2. Crystal structure, conformational fixation and entry-related interactions of mature ligand-free HIV-1 Env

    DOE PAGES

    Do Kwon, Young; Pancera, Marie; Acharya, Priyamvada; ...

    2015-06-22

    As the sole viral antigen on the HIV-1–virion surface, trimeric Env is a focus of vaccine efforts. In this paper, we present the structure of the ligand-free HIV-1–Env trimer, fix its conformation and determine its receptor interactions. Epitope analyses revealed trimeric ligand-free Env to be structurally compatible with broadly neutralizing antibodies but not poorly neutralizing ones. We coupled these compatibility considerations with binding antigenicity to engineer conformationally fixed Envs, including a 201C 433C (DS) variant specifically recognized by broadly neutralizing antibodies. DS-Env retained nanomolar affinity for the CD4 receptor, with which it formed an asymmetric intermediate: a closed trimer boundmore » by a single CD4 without the typical antigenic hallmarks of CD4 induction. Finally, antigenicity-guided structural design can thus be used both to delineate mechanism and to fix conformation, with DS-Env trimers in virus-like-particle and soluble formats providing a new generation of vaccine antigens.« less

  3. Crystal structure, conformational fixation and entry-related interactions of mature ligand-free HIV-1 Env

    SciTech Connect

    Do Kwon, Young; Pancera, Marie; Acharya, Priyamvada; Georgiev, Ivelin S.; Crooks, Emma T.; Gorman, Jason; Joyce, M. Gordon; Guttman, Miklos; Ma, Xiaochu; Narpala, Sandeep; Soto, Cinque; Terry, Daniel S.; Yang, Yongping; Zhou, Tongqing; Ahlsen, Goran; Bailer, Robert T.; Chambers, Michael; Chuang, Gwo-Yu; Doria-Rose, Nicole A.; Druz, Aliaksandr; Hallen, Mark A.; Harned, Adam; Kirys, Tatsiana; Louder, Mark K.; O'Dell, Sijy; Ofek, Gilad; Osawa, Keiko; Prabhakaran, Madhu; Sastry, Mallika; Stewart-Jones, Guillaume B. E.; Stuckey, Jonathan; Thomas, Paul V.; Tittley, Tishina; Williams, Constance; Zhang, Baoshan; Zhao, Hong; Zhou, Zhou; Donald, Bruce R.; Lee, Lawrence K.; Zolla-Pazner, Susan; Baxa, Ulrich; Schön, Arne; Freire, Ernesto; Shapiro, Lawrence; Lee, Kelly K.; Arthos, James; Munro, James B.; Blanchard, Scott C.; Mothes, Walther; Binley, James M.; McDermott, Adrian B.; Mascola, John R.; Kwong, Peter D.

    2015-06-22

    As the sole viral antigen on the HIV-1–virion surface, trimeric Env is a focus of vaccine efforts. In this paper, we present the structure of the ligand-free HIV-1–Env trimer, fix its conformation and determine its receptor interactions. Epitope analyses revealed trimeric ligand-free Env to be structurally compatible with broadly neutralizing antibodies but not poorly neutralizing ones. We coupled these compatibility considerations with binding antigenicity to engineer conformationally fixed Envs, including a 201C 433C (DS) variant specifically recognized by broadly neutralizing antibodies. DS-Env retained nanomolar affinity for the CD4 receptor, with which it formed an asymmetric intermediate: a closed trimer bound by a single CD4 without the typical antigenic hallmarks of CD4 induction. Finally, antigenicity-guided structural design can thus be used both to delineate mechanism and to fix conformation, with DS-Env trimers in virus-like-particle and soluble formats providing a new generation of vaccine antigens.

  4. HIV-1 Nef responsiveness is determined by Env variable regions involved in trimer association and correlates with neutralization sensitivity.

    PubMed

    Usami, Yoshiko; Göttlinger, Heinrich

    2013-11-14

    HIV-1 Nef and the unrelated murine leukemia virus glycoGag similarly enhance the infectivity of HIV-1 virions. We now show that the effects of Nef and glycoGag are similarly determined by variable regions of HIV-1 gp120 that control Env trimer association and neutralization sensitivity. Whereas neutralization-sensitive X4-tropic Env proteins conferred high responsiveness to Nef and glycoGag, particles bearing neutralization-resistant R5-tropic Envs were considerably less affected. The profoundly different Nef/glycoGag responsiveness of a neutralization-resistant and a neutralization-sensitive R5-tropic Env could be switched by exchanging their gp120 V1/V2 regions, which also switches their neutralization sensitivity. Within V1/V2, the same determinants governed Nef/glycoGag responsiveness and neutralization sensitivity, indicating that these phenotypes are mechanistically linked. The V1/V2 and V3 regions, which form an apical trimer-association domain, together determined the Nef and glycoGag responsiveness of an X4-tropic Env. Our results suggest that Nef and glycoGag counteract the inactivation of Env spikes with relatively unstable apical trimer-association domains.

  5. Adenovirus prime, Env protein boost vaccine protects against neutralization-resistant SIVsmE660 variants in rhesus monkeys.

    PubMed

    Keele, Brandon F; Li, Wenjun; Borducchi, Erica N; Nkolola, Joseph P; Abbink, Peter; Chen, Bing; Seaman, Michael S; Barouch, Dan H

    2017-06-05

    Previous studies have shown that DNA prime, Ad5 boost vaccines protect against neutralization-sensitive but not neutralization-resistant virus variants within the SIVsmE660 swarm. Here we show that Ad prime, Env protein boost vaccines protect against neutralization-resistant SIVsmE660 variants. We perform two studies in rhesus monkeys with Ad35/Ad26 vectors expressing SIVmac239 Gag/Pol/Env with or without an AS01B-adjuvanted SIVmac32H gp140 protein boost. In a repetitive, low-dose challenge study, we observe robust protection against acquisition of infection by both Ad Alone and Ad/Env vaccines. In a single, high-dose challenge study, only the Ad/Env vaccine affords significant protection against acquisition of infection. Analysis of transmitted/founder (T/F) viruses from this study demonstrates that the Ad/Env vaccine blocks both neutralization-sensitive and neutralization-resistant SIVsmE660 variants in rhesus monkeys with restrictive TRIM5α alleles. These data demonstrate that the adjuvanted Env protein boost is critical for protecting against high-dose SIVsmE660 challenge and for blocking neutralization-resistant viruses within the SIVsmE660 swarm.

  6. Crystal structure, conformational fixation, and entry-related interactions of mature ligand-free HIV-1 Env

    PubMed Central

    Kwon, Young Do; Pancera, Marie; Acharya, Priyamvada; Georgiev, Ivelin S.; Crooks, Emma T.; Gorman, Jason; Joyce, M. Gordon; Guttman, Miklos; Ma, Xiaochu; Narpala, Sandeep; Soto, Cinque; Terry, Daniel S.; Yang, Yongping; Zhou, Tongqing; Ahlsen, Goran; Bailer, Robert T.; Chambers, Michael; Chuang, Gwo-Yu; Doria-Rose, Nicole A.; Druz, Aliaksandr; Hallen, Mark A.; Harned, Adam; Kirys, Tatsiana; Louder, Mark K.; O’Dell, Sijy; Ofek, Gilad; Osawa, Keiko; Prabhakaran, Madhu; Sastry, Mallika; Stewart-Jones, Guillaume B.E.; Stuckey, Jonathan; Thomas, Paul V.; Tittley, Tishina; Williams, Constance; Zhang, Baoshan; Zhao, Hong; Zhou, Zhou; Donald, Bruce R.; Lee, Lawrence K.; Zolla-Pazner, Susan; Baxa, Ulrich; Schön, Arne; Freire, Ernesto; Shapiro, Lawrence; Lee, Kelly K.; Arthos, James; Munro, James B.; Blanchard, Scott C.; Mothes, Walther; Binley, James M.; McDermott, Adrian B.; Mascola, John R.; Kwong, Peter D.

    2016-01-01

    As the sole viral antigen on the HIV-1-virion surface, trimeric Env is a focus of vaccine efforts. Here we present the structure of the ligand-free HIV-1-Env trimer, fix its conformation, and determine its receptor interactions. Epitope analyses revealed trimeric ligand-free Env to be structurally compatible with broadly neutralizing antibodies, but not poorly neutralizing ones. We coupled these compatibility considerations with binding antigenicity to engineer conformationally fixed Envs, including a 201C-433C (DS) variant, specifically recognized by broadly neutralizing antibodies. DS-Env retained nanomolar affinity for the CD4 receptor, with which it formed an asymmetric intermediate: a closed trimer bound by a single CD4 without the typical antigenic hallmarks of CD4 induction. Antigenicity-guided structural design can thus be used both to delineate mechanism and to fix conformation, with DS-Env trimers in virus-like particle and soluble formats providing a new generation of vaccine antigens. PMID:26098315

  7. Antibody potency relates to the ability to recognize the closed, pre-fusion form of HIV Env

    NASA Astrophysics Data System (ADS)

    Guttman, Miklos; Cupo, Albert; Julien, Jean-Philippe; Sanders, Rogier W.; Wilson, Ian A.; Moore, John P.; Lee, Kelly K.

    2015-02-01

    HIV’s envelope glycoprotein (Env) is the sole target for neutralizing antibodies. The structures of many broadly neutralizing antibodies (bNAbs) in complex with truncated Env subunits or components have been reported. However, their interaction with the intact Env trimer, and the structural determinants that underlie neutralization resistance in this more native context are less well understood. Here we use hydrogen/deuterium exchange to examine the interactions between a panel of bNAbs and native-like Env trimers (SOSIP.664 trimers). Highly potent bNAbs cause only localized effects at their binding interface, while the binding of less potent antibodies is associated with elaborate changes throughout the trimer. In conjunction with binding kinetics, our results suggest that poorly neutralizing antibodies can only bind when the trimer transiently samples an open state. We propose that the kinetics of such opening motions varies among isolates, with Env from neutralization-sensitive viruses opening more frequently than Env from resistant viruses.

  8. Panels of HIV-1 Subtype C Env Reference Strains for Standardized Neutralization Assessments

    DOE PAGES

    Hraber, Peter; Rademeyer, Cecilia; Williamson, Carolyn; ...

    2017-07-26

    In the search for effective immunologic interventions to prevent and treat HIV-1 infection, standardized reference reagents are a cost-effective way to maintain robustness and reproducibility among immunological assays. To support planned and ongoing studies where clade C predominates, here we describe three virus panels, chosen from 200 well-characterized clade C envelope (Env)-pseudotyped viruses from early infection. All 200 Envs were expressed as a single round of replication pseudoviruses and were tested to quantify neutralization titers by 16 broadly neutralizing antibodies (bnAbs) and sera from 30 subjects with chronic clade C infections. We selected large panels of 50 and 100 Envsmore » either to characterize cross-reactive breadth for sera identified as having potent neutralization activity based on initial screening or to evaluate neutralization magnitude-breadth distributions of newly isolated antibodies. We identified these panels by downselection after hierarchical clustering of bnAb neutralization titers. The resulting panels represent the diversity of neutralization profiles throughout the range of virus sensitivities identified in the original panel of 200 viruses. A small 12-Env panel was chosen to screen sera from vaccine trials or natural-infection studies for neutralization responses. We considered panels selected by previously described methods but favored a computationally informed method that enabled selection of viruses representing diverse neutralization sensitivity patterns, given that we do nota prioriknow what the neutralization-response profile of vaccine sera will be relative to that of sera from infected individuals. The resulting 12-Env panel complements existing panels. Use of standardized panels enables direct comparisons of data from different trials and study sites testing HIV-1 clade C-specific products. HIV-1 group M includes nine clades and many recombinants. Clade C is the most common lineage, responsible for roughly half

  9. Enteractinococcus coprophilus gen. nov., sp. nov., of the family Micrococcaceae, isolated from Panthera tigris amoyensis faeces, and transfer of Yaniella fodinae Dhanjal et al. 2011 to the genus Enteractinococcus as Enteractinococcus fodinae comb. nov.

    PubMed Central

    Cao, Yan-Ru; Jin, Rong-Xian; Han, Li; Li, You-Long; Huang, Xue-Shi; Xue, Quan-Hong

    2012-01-01

    A novel actinobacterium, designated strain YIM 100590T, was isolated from Panthera tigris amoyensis faeces collected from Yunnan Wild Animal Park in Yunnan province, south-west China. Phylogenetic analysis based on 16S rRNA gene sequence data showed that strain YIM 100590T is a member of the family Micrococcaceae. Cells were coccoid to oval (0.7–1.5 µm in diameter) occurring singly or in clusters. Growth was observed at 10–37 °C (optimum 28 °C) and at pH 7.0–11.0 (optimum pH 8.0). The major fatty acids were iso-C15 : 0 (32.22 %), anteiso-C15 : 0 (31.64 %) and iso-C16 : 0 (17.38 %). The peptidoglycan was of A4α type (l-Lys–Gly–l-Glu). The major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, phosphatidylinositol mannosides, dimannosyl diacylglycerol, an unknown glycolipid and two unknown phospholipids. The quinone system comprised menaquinones MK-7 (91.9 %) and MK-8 (8.3 %). The DNA G+C content of strain YIM 100590T was 56.2 mol%. Chemotaxonomic data indicated that the strain belongs to the family Micrococcaceae. On the basis of morphological and chemotaxonomic data and phylogenetic analysis, strain YIM 100590T is considered to represent a novel species of a new genus within the family Micrococcaceae, for which the name Enteractinococcus coprophilus gen. nov., sp. nov. is proposed. The type strain of Enteractinococcus coprophilus is YIM 100590T ( = DSM 24083T = JCM 17352T). Yaniella fodinae DSM 22966T was transferred to the new genus as Enteractinococcus fodinae comb. nov. (type strain G5T = DSM 22966T = JCM 17931T = MTCC 9846T). PMID:22228667

  10. Bordetella bronchialis sp. nov., Bordetella flabilis sp. nov. and Bordetella sputigena sp. nov., isolated from human respiratory specimens, and reclassification of Achromobacter sediminum Zhang et al. 2014 as Verticia sediminum gen. nov., comb. nov.

    PubMed

    Vandamme, Peter A; Peeters, Charlotte; Cnockaert, Margo; Inganäs, Elisabeth; Falsen, Enevold; Moore, Edward R B; Nunes, Olga C; Manaia, Célia M; Spilker, Theodore; LiPuma, John J

    2015-10-01

    The phenotypic and genotypic characteristics of four Bordetella hinzii-like strains from human respiratory specimens and representing nrdA gene sequence based genogroups 3, 14 and 15 were examined. In a 16S rRNA gene sequence based phylogenetic tree, the four strains consistently formed a single coherent lineage but their assignment to the genus Bordetella was equivocal. The respiratory quinone, polar lipid and fatty acid profiles generally conformed to those of species of the genus Bordetella and were characterized by the presence of ubiquinone 8, of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol and several aminolipids, and of high percentages of C16 : 0, cyclo-C17 : 0 and summed feature 2, as major chemotaxonomic marker molecules, respectively. The DNA G+C content was about 66 mol%, which corresponded with that of the high-percentage DNA G+C content genera of the family Alcaligenaceae including the genus Bordetella. DNA–DNA hybridization experiments revealed the presence of three distinct genomospecies and thus confirmed phenotypic differences as revealed by means of extensive biochemical characterization. We therefore propose to formally classify Bordetella genogroups 3, 14 and 15 as Bordetella bronchialis sp. nov. (type strain LMG 28640T = AU3182T = CCUG 56828T), Bordetella sputigena sp. nov. (type strain LMG 28641T = CCUG 56478T) and Bordetella flabilis sp. nov. (type strain LMG 28642T = AU10664T = CCUG 56827T). In addition, we propose to reclassify Achromobacter sediminum into the novel genus Verticia, as Verticia sediminum, gen. nov., comb. nov., on the basis of its unique phylogenetic position, its marine origin and its distinctive phenotypic, fatty acid and polar lipid profile.

  11. Altererythrobacter epoxidivorans gen. nov., sp. nov., an epoxide hydrolase-active, mesophilic marine bacterium isolated from cold-seep sediment, and reclassification of Erythrobacter luteolus Yoon et al. 2005 as Altererythrobacter luteolus comb. nov.

    PubMed

    Kwon, Kae Kyoung; Woo, Jung-Hee; Yang, Sung-Hyun; Kang, Ji-Hyun; Kang, Sung Gyun; Kim, Sang-Jin; Sato, Takako; Kato, Chiaki

    2007-10-01

    A novel marine bacterium, strain JCS350(T), was isolated from marine sediment samples collected from a cold-seep area. The 16S rRNA gene sequence of the isolate showed high similarity to that of Erythrobacter luteolus SW-109(T) (95.9 % sequence similarity). Lower 16S rRNA gene sequence similarities were shown to other members of the genus Erythrobacter (94.6-95.4 %) and members of the genus Porphyrobacter (94.5-95.2 %). Phylogenetic analysis with all members of the family Erythrobacteraceae and several members of the family Sphingomonadaceae revealed that the isolate formed a phyletic line with [Erythrobacter] luteolus that was distinct from other members of the family Erythrobacteraceae. The dominant fatty acids of strain JCS350(T) were 18 : 1omega7c, 16 : 1omega7c and cyclopropane 17 : 0. The major respiratory quinone was ubiquinone 10. The DNA G+C content was 54.5 mol%. The isolate did not contain bacteriochlorophyll a. Optimal growth required the presence of 2 % (w/v) NaCl with either 0.18 % CaCl(2) or 0.59 % MgCl(2), at pH 6.5 and at 35 degrees C. On the basis of the evidence of this polyphasic taxonomic study, strain JCS350(T) should be classified in a novel genus and species in the family Erythrobacteraceae, for which the name Altererythrobacter epoxidivorans gen. nov., sp. nov. is proposed. The misclassified species [Erythrobacter] luteolus is transferred to the new genus as Altererythrobacter luteolus comb. nov. The type strain of Altererythrobacter epoxidivorans is JCS350(T) (=KCCM 42314(T) =JCM 13815(T)) and the type strain of Altererythrobacter luteolus is SW-109(T) (=KCTC 12311(T) =JCM 12599(T)).

  12. Antagonism of BST-2/Tetherin Is a Conserved Function of the Env Glycoprotein of Primary HIV-2 Isolates.

    PubMed

    Chen, Chia-Yen; Shingai, Masashi; Welbourn, Sarah; Martin, Malcolm A; Borrego, Pedro; Taveira, Nuno; Strebel, Klaus

    2016-12-15

    Although HIV-2 does not encode a vpu gene, the ability to antagonize bone marrow stromal antigen 2 (BST-2) is conserved in some HIV-2 isolates, where it is controlled by the Env glycoprotein. We previously reported that a single-amino-acid difference between the laboratory-adapted ROD10 and ROD14 Envs controlled the enhancement of virus release (referred to here as Vpu-like) activity. Here, we investigated how conserved the Vpu-like activity is in primary HIV-2 isolates. We found that half of the 34 tested primary HIV-2 Env isolates obtained from 7 different patients enhanced virus release. Interestingly, most HIV-2 patients harbored a mixed population of viruses containing or lacking Vpu-like activity. Vpu-like activity and Envelope functionality varied significantly among Env isolates; however, there was no direct correlation between these two functions, suggesting they evolved independently. In comparing the Env sequences from one HIV-2 patient, we found that similar to the ROD10/ROD14 Envs, a single-amino-acid change (T568I) in the ectodomain of the TM subunit was sufficient to confer Vpu-like activity to an inactive Env variant. Surprisingly, however, absence of Vpu-like activity was not correlated with absence of BST-2 interaction. Taken together, our data suggest that maintaining the ability to antagonize BST-2 is of functional relevance not only to HIV-1 but also to HIV-2 as well. Our data show that as with Vpu, binding of HIV-2 Env to BST-2 is important but not sufficient for antagonism. Finally, as observed previously, the Vpu-like activity in HIV-2 Env can be controlled by single-residue changes in the TM subunit. Lentiviruses such as HIV-1 and HIV-2 encode accessory proteins whose function is to overcome host restriction mechanisms. Vpu is a well-studied HIV-1 accessory protein that enhances virus release by antagonizing the host restriction factor BST-2. HIV-2 does not encode a vpu gene. Instead, the HIV-2 Env glycoprotein was found to antagonize BST-2

  13. Saccharomyces cerevisiae Env7 Is a Novel Serine/Threonine Kinase 16-Related Protein Kinase and Negatively Regulates Organelle Fusion at the Lysosomal Vacuole

    PubMed Central

    Manandhar, Surya P.; Ricarte, Florante; Cocca, Stephanie M.

    2013-01-01

    Membrane fusion depends on conserved components and is responsible for organelle biogenesis and vesicular trafficking. Yeast vacuoles are dynamic structures analogous to mammalian lysosomes. We report here that yeast Env7 is a novel palmitoylated protein kinase ortholog that negatively regulates vacuolar membrane fusion. Microscopic and biochemical studies confirmed the localization of tagged Env7 at the vacuolar membrane and implicated membrane association via the palmitoylation of its N-terminal Cys13 to -15. In vitro kinase assays established Env7 as a protein kinase. Site-directed mutagenesis of the Env7 alanine-proline-glutamic acid (APE) motif Glu269 to alanine results in an unstable kinase-dead allele that is stabilized and redistributed to the detergent-resistant fraction by interruption of the proteasome system in vivo. Palmitoylation-deficient Env7C13-15S is also kinase dead and mislocalizes to the cytoplasm. Microscopy studies established that env7Δ is defective in maintaining fragmented vacuoles during hyperosmotic response and in buds. ENV7 function is not redundant with a similar role of vacuolar membrane kinase Yck3, as the two do not share a substrate, and ENV7 is not a suppressor of yck3Δ. Bayesian phylogenetic analyses strongly support ENV7 as an ortholog of the gene encoding human STK16, a Golgi apparatus protein kinase with undefined function. We propose that Env7 function in fusion/fission dynamics may be conserved within the endomembrane system. PMID:23166297

  14. Saccharomyces cerevisiae Env7 is a novel serine/threonine kinase 16-related protein kinase and negatively regulates organelle fusion at the lysosomal vacuole.

    PubMed

    Manandhar, Surya P; Ricarte, Florante; Cocca, Stephanie M; Gharakhanian, Editte

    2013-02-01

    Membrane fusion depends on conserved components and is responsible for organelle biogenesis and vesicular trafficking. Yeast vacuoles are dynamic structures analogous to mammalian lysosomes. We report here that yeast Env7 is a novel palmitoylated protein kinase ortholog that negatively regulates vacuolar membrane fusion. Microscopic and biochemical studies confirmed the localization of tagged Env7 at the vacuolar membrane and implicated membrane association via the palmitoylation of its N-terminal Cys13 to -15. In vitro kinase assays established Env7 as a protein kinase. Site-directed mutagenesis of the Env7 alanine-proline-glutamic acid (APE) motif Glu269 to alanine results in an unstable kinase-dead allele that is stabilized and redistributed to the detergent-resistant fraction by interruption of the proteasome system in vivo. Palmitoylation-deficient Env7C13-15S is also kinase dead and mislocalizes to the cytoplasm. Microscopy studies established that env7Δ is defective in maintaining fragmented vacuoles during hyperosmotic response and in buds. ENV7 function is not redundant with a similar role of vacuolar membrane kinase Yck3, as the two do not share a substrate, and ENV7 is not a suppressor of yck3Δ. Bayesian phylogenetic analyses strongly support ENV7 as an ortholog of the gene encoding human STK16, a Golgi apparatus protein kinase with undefined function. We propose that Env7 function in fusion/fission dynamics may be conserved within the endomembrane system.

  15. 25 Years of GenBank

    MedlinePlus

    ... of this page please turn Javascript on. Unique DNA database has helped advance scientific discoveries worldwide Since ... GenBank? Basically, GenBank is the biggest database of DNA in the world. Deoxyribonucleic acid (DNA) is the ...

  16. Real-time analysis of human immunodeficiency virus type 1 Env-mediated membrane fusion by fluorescence resonance energy transfer.

    PubMed

    Furuta, Rika A; Nishikawa, Masao; Fujisawa, Jun-ichi

    2006-02-01

    Human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env)-mediated membrane fusion occurs as a sequence of events that is triggered by CD4 binding to the Env gp120 subunit. In this study, we analyzed the dynamics of Env-mediated membrane fusion at the single-cell level using fluorescent fusion proteins and confocal laser fluorescent microscopy. Either enhanced cyan or yellow fluorescent protein (CFP and YFP, respectively) was fused to the end of the cytoplasmic regions of the HIV-1 receptors (CD4 and CCR5) and Env proteins. Real-time imaging of membrane fusion mediated by these recombinant proteins revealed that the kinetics of fusion in our system was faster than that previously reported. Analysis of the receptor interaction by fluorescence resonance energy transfer (FRET) at the single-cell level demonstrated a tendency for oligomerization of CD4-CD4, but not of CD4-CCR5, in the absence of Env-expressing cells. However, when Env-expressing cells attached to the receptor cells, FRET produced by CD4-CCR5 interaction was increased; the FRET intensity began to decline before the formation of the fusion pore. These changes in FRET may represent the temporal association of these receptors, triggered by gp120 binding, and their dissociation during the formation of the fusion pore. In addition, the FRET analysis of receptor interactions in the presence of fusion inhibitors showed that not only inhibitors acting on CCR5 but also the gp41-derived peptide T-20 interfered with CD4-CCR5 interaction during fusion. These data suggest that T-20 could affect the formation of Env-receptors complexes during the membrane fusion.

  17. Safety and Immunogenicity of a rAd35-EnvA Prototype HIV-1 Vaccine in Combination with rAd5-EnvA in Healthy Adults (VRC 012).

    PubMed

    Crank, Michelle C; Wilson, Eleanor M P; Novik, Laura; Enama, Mary E; Hendel, Cynthia S; Gu, Wenjuan; Nason, Martha C; Bailer, Robert T; Nabel, Gary J; McDermott, Adrian B; Mascola, John R; Koup, Richard A; Ledgerwood, Julie E; Graham, Barney S

    2016-01-01

    VRC 012 was a Phase I study of a prototype recombinant adenoviral-vector serotype-35 (rAd35) HIV vaccine, the precursor to two recently published clinical trials, HVTN 077 and 083. On the basis of prior evaluation of multiclade rAd5 HIV vaccines, Envelope A (EnvA) was selected as the standard antigen for a series of prototype HIV vaccines to compare various vaccine platforms. In addition, prior studies of rAd5-vectored vaccines suggested pre-existing human immunity may be a confounding factor in vaccine efficacy. rAd35 is less seroprevalent across human populations and was chosen for testing alone and in combination with a rAd5-EnvA vaccine in the present two-part phase I study. First, five subjects each received a single injection of 109, 1010, or 1011 particle units (PU) of rAd35-EnvA in an open-label, dose-escalation study. Next, 20 Ad5/Ad35-seronegative subjects were randomized to blinded, heterologous prime-boost schedules combining rAd5-EnvA and rAd35-EnvA with a three month interval. rAd35-EnvA was given at 1010 or 1011 PU to ten subjects each; all rAd5-EnvA injections were 1010 PU. EnvA-specific immunogenicity was assessed four weeks post-injection. Solicited reactogenicity and clinical safety were followed after each injection. Vaccinations were well tolerated at all dosages. Antibody responses measured by ELISA were detected at 4 weeks in 30% and 50% of subjects after single doses of 1010 or 1011 PU rAd35, respectively, and in 89% after a single rAd5-EnvA 1010 PU injection. EnvA-specific IFN-γ ELISpot responses were detected at four weeks in 0%, 70%, and 50% of subjects after the respective rAd35-EnvA dosages compared to 89% of subjects after rAd5. T cell responses were higher after a single rAd5-EnvA 1010 PU injection than after a single rAd35-EnvA 1010 PU injection, and humoral responses were low after a single dose of either vector. Of those completing the vaccine schedule, 100% of rAd5-EnvA recipients and 90% of rAd35-EnvA recipients had both T cell

  18. Safety and Immunogenicity of a rAd35-EnvA Prototype HIV-1 Vaccine in Combination with rAd5-EnvA in Healthy Adults (VRC 012)

    PubMed Central

    Crank, Michelle C.; Wilson, Eleanor M. P.; Novik, Laura; Enama, Mary E.; Hendel, Cynthia S.; Gu, Wenjuan; Nason, Martha C.; Bailer, Robert T.; Nabel, Gary J.; McDermott, Adrian B.; Mascola, John R.; Koup, Richard A.; Ledgerwood, Julie E.; Graham, Barney S.

    2016-01-01

    Background VRC 012 was a Phase I study of a prototype recombinant adenoviral-vector serotype-35 (rAd35) HIV vaccine, the precursor to two recently published clinical trials, HVTN 077 and 083. On the basis of prior evaluation of multiclade rAd5 HIV vaccines, Envelope A (EnvA) was selected as the standard antigen for a series of prototype HIV vaccines to compare various vaccine platforms. In addition, prior studies of rAd5-vectored vaccines suggested pre-existing human immunity may be a confounding factor in vaccine efficacy. rAd35 is less seroprevalent across human populations and was chosen for testing alone and in combination with a rAd5-EnvA vaccine in the present two-part phase I study. Methods First, five subjects each received a single injection of 109, 1010, or 1011 particle units (PU) of rAd35-EnvA in an open-label, dose-escalation study. Next, 20 Ad5/Ad35-seronegative subjects were randomized to blinded, heterologous prime-boost schedules combining rAd5-EnvA and rAd35-EnvA with a three month interval. rAd35-EnvA was given at 1010 or 1011 PU to ten subjects each; all rAd5-EnvA injections were 1010 PU. EnvA-specific immunogenicity was assessed four weeks post-injection. Solicited reactogenicity and clinical safety were followed after each injection. Results Vaccinations were well tolerated at all dosages. Antibody responses measured by ELISA were detected at 4 weeks in 30% and 50% of subjects after single doses of 1010 or 1011 PU rAd35, respectively, and in 89% after a single rAd5-EnvA 1010 PU injection. EnvA-specific IFN-γ ELISpot responses were detected at four weeks in 0%, 70%, and 50% of subjects after the respective rAd35-EnvA dosages compared to 89% of subjects after rAd5. T cell responses were higher after a single rAd5-EnvA 1010 PU injection than after a single rAd35-EnvA 1010 PU injection, and humoral responses were low after a single dose of either vector. Of those completing the vaccine schedule, 100% of rAd5-EnvA recipients and 90% of rAd35-Env

  19. Introduction to GenCade

    DTIC Science & Technology

    2012-10-01

    16-18 October 2012 5 GenCade Conceptual Processes: Coastal Sediment Dynamics Flood shoal ~150,000 m3/yr D50 = 0.27 (W) 0.38 (E) Moriches...offset • LMOVY 0.0 Leftward shoreline displacement velocity • D50 0.33 Grain size diameter in millimeters • BERMHT 2 Average berm height • DCLOS 8

  20. FutureGen Project Report

    SciTech Connect

    Cabe, Jim; Elliott, Mike

    2010-09-30

    This report summarizes the comprehensive siting, permitting, engineering, design, and costing activities completed by the FutureGen Industrial Alliance, the Department of Energy, and associated supporting subcontractors to develop a first of a kind near zero emissions integrated gasification combined cycle power plant and carbon capture and storage project (IGCC-CCS). With the goal to design, build, and reliably operate the first IGCC-CCS facility, FutureGen would have been the lowest emitting pulverized coal power plant in the world, while providing a timely and relevant basis for coal combustion power plants deploying carbon capture in the future. The content of this report summarizes key findings and results of applicable project evaluations; modeling, design, and engineering assessments; cost estimate reports; and schedule and risk mitigation from initiation of the FutureGen project through final flow sheet analyses including capital and operating reports completed under DOE award DE-FE0000587. This project report necessarily builds upon previously completed siting, design, and development work executed under DOE award DE-FC26- 06NT4207 which included the siting process; environmental permitting, compliance, and mitigation under the National Environmental Policy Act; and development of conceptual and design basis documentation for the FutureGen plant. For completeness, the report includes as attachments the siting and design basis documents, as well as the source documentation for the following: • Site evaluation and selection process and environmental characterization • Underground Injection Control (UIC) Permit Application including well design and subsurface modeling • FutureGen IGCC-CCS Design Basis Document • Process evaluations and technology selection via Illinois Clean Coal Review Board Technical Report • Process flow diagrams and heat/material balance for slurry-fed gasifier configuration • Process flow diagrams and heat/material balance

  1. env Gene typing of human immunodeficiency virus type 1 strains on electronic microarrays.

    PubMed

    Saunders, N A; Alexander, S; Tatt, I

    2005-04-01

    The NanoChip system was used for subtyping human immunodeficiency virus type 1 (HIV-1) strains using probes complementary to the V1 region of the env gene. Probes for six subtypes (A to D, F, and G) and two circulating recombinant forms (AG and AE) of HIV-1 group M were included. The specificity of these oligonucleotides had been evaluated previously in a DNA enzyme immunoassay. Samples from 112 patient sera were used as templates in a nested reverse transcription-PCR to produce amplicons that were applied to the array. The array was then hybridized successively to pairs of oligonucleotide probes. The strains were assigned a subtype on the basis of their probe hybridization patterns. One strain gave a contradictory pattern and was designated as untypeable by the NanoChip assay. Eighty-eight strains gave hybridization patterns that allowed a correct subtype designation to be made by the NanoChip assay compared to either the sequence or the heteroduplex mobility assay (HMA)-determined subtypes. Thirteen strains that reacted with the subtype A probe (SA2) were incorrectly assigned to subtype A, or to one of the related circulating recombinant types (AE or AG), on the basis of reactions with probe SAE1 or SAG1. The results indicate that these oligonucleotides have relatively low specificities. The probe subtypes of three strains matched the subtypes determined for the gag and pol genes but not the env gene, suggesting that a recombination event may have occurred within the env gene. Overall, the NanoChip assay gave results comparable to those for HMA and sequencing and provides a convenient and cost-effective means by which to subtype HIV-1.

  2. env Gene Typing of Human Immunodeficiency Virus Type 1 Strains on Electronic Microarrays

    PubMed Central

    Saunders, N. A.; Alexander, S.; Tatt, I.

    2005-01-01

    The NanoChip system was used for subtyping human immunodeficiency virus type 1 (HIV-1) strains using probes complementary to the V1 region of the env gene. Probes for six subtypes (A to D, F, and G) and two circulating recombinant forms (AG and AE) of HIV-1 group M were included. The specificity of these oligonucleotides had been evaluated previously in a DNA enzyme immunoassay. Samples from 112 patient sera were used as templates in a nested reverse transcription-PCR to produce amplicons that were applied to the array. The array was then hybridized successively to pairs of oligonucleotide probes. The strains were assigned a subtype on the basis of their probe hybridization patterns. One strain gave a contradictory pattern and was designated as untypeable by the NanoChip assay. Eighty-eight strains gave hybridization patterns that allowed a correct subtype designation to be made by the NanoChip assay compared to either the sequence or the heteroduplex mobility assay (HMA)-determined subtypes. Thirteen strains that reacted with the subtype A probe (SA2) were incorrectly assigned to subtype A, or to one of the related circulating recombinant types (AE or AG), on the basis of reactions with probe SAE1 or SAG1. The results indicate that these oligonucleotides have relatively low specificities. The probe subtypes of three strains matched the subtypes determined for the gag and pol genes but not the env gene, suggesting that a recombination event may have occurred within the env gene. Overall, the NanoChip assay gave results comparable to those for HMA and sequencing and provides a convenient and cost-effective means by which to subtype HIV-1. PMID:15815017

  3. Phylogenetic and structural diversity in the feline leukemia virus env gene.

    PubMed

    Watanabe, Shinya; Kawamura, Maki; Odahara, Yuka; Anai, Yukari; Ochi, Haruyo; Nakagawa, So; Endo, Yasuyuki; Tsujimoto, Hajime; Nishigaki, Kazuo

    2013-01-01

    Feline leukemia virus (FeLV) belongs to the genus Gammaretrovirus, and causes a variety of neoplastic and non-neoplastic diseases in cats. Alteration of viral env sequences is thought to be associated with disease specificity, but the way in which genetic diversity of FeLV contributes to the generation of such variants in nature is poorly understood. We isolated FeLV env genes from naturally infected cats in Japan and analyzed the evolutionary dynamics of these genes. Phylogenetic reconstructions separated our FeLV samples into three distinct genetic clusters, termed Genotypes I, II, and III. Genotype I is a major genetic cluster and can be further classified into Clades 1-7 in Japan. Genotypes were correlated with geographical distribution; Genotypes I and II were distributed within Japan, whilst FeLV samples from outside Japan belonged to Genotype III. These results may be due to geographical isolation of FeLVs in Japan. The observed structural diversity of the FeLV env gene appears to be caused primarily by mutation, deletion, insertion and recombination, and these variants may be generated de novo in individual cats. FeLV interference assay revealed that FeLV genotypes did not correlate with known FeLV receptor subgroups. We have identified the genotypes which we consider to be reliable for evaluating phylogenetic relationships of FeLV, which embrace the high structural diversity observed in our sample. Overall, these findings extend our understanding of Gammaretrovirus evolutionary patterns in the field, and may provide a useful basis for assessing the emergence of novel strains and understanding the molecular mechanisms of FeLV transmission in cats.

  4. Biochem-Env: a platform of biochemistry for research in environmental and agricultural sciences.

    PubMed

    Cheviron, Nathalie; Grondin, Virginie; Mougin, Christian

    2017-04-07

    Biochemical indicators are potent tools to assess ecosystem functioning under anthropic and global pressures. Nevertheless, additional work is needed to improve the methods used for the measurement of these indicators, and for a more relevant interpretation of the obtained results. To face these challenges, the platform Biochem-Env aims at providing innovative and standardized measurement protocols, as well as database and information system favoring result interpretation and opening. Its skills and tools are also offered for expertise, consulting, training, and standardization. In addition, the platform is a service of a French Research Infrastructure for Analysis and Experimentation on Ecosystems, for research in environmental and agricultural sciences.

  5. Drosophila germline invasion by the endogenous retrovirus gypsy: involvement of the viral env gene.

    PubMed

    Pelisson, A; Mejlumian, L; Robert, V; Terzian, C; Bucheton, A

    2002-10-01

    The endogenous retrovirus gypsy is expressed at high levels in mutant flamenco female flies. Gypsy viral particles extracted from such flies can infect naive flamenco individuals raised in the presence of these extracts mixed into their food. This results in the integration of new proviruses into the germline genome. These proviruses can then increase their copy number by (1) expression in the flamenco female somatic cells, (2) transfer into the oocyte and (3) integration into the genome of the progeny. Surprisingly, unlike the infection observed in the feeding experiments, this strategy of endogenous proviral multiplication does not seem to involve the expression of the viral env gene.

  6. Reclassification of Rhodospirillum photometricum Molisch 1907, Rhodospirillum sulfurexigens Anil Kumar et al. 2008 and Rhodospirillum oryzae Lakshmi et al. 2013 in a new genus, Pararhodospirillum gen. nov., as Pararhodospirillum photometricum comb. nov., Pararhodospirillum sulfurexigens comb. nov. and Pararhodospirillum oryzae comb. nov., respectively, and emended description of the genus Rhodospirillum.

    PubMed

    Lakshmi, K V N S; Divyasree, B; Ramprasad, E V V; Sasikala, Ch; Ramana, Ch V

    2014-04-01

    The genus Rhodospirillum is represented by four species, with three of them showing phylogenetic divergence compared to the type species, Rhodospirillum rubrum. Differences in the major diagnostic properties such as internal photosynthetic membranes, quinones, fatty acids, carotenoid composition and a few other phenotypic properties warrant the reclassification of members of this genus. Resultantly, a new genus, Pararhodospirillum gen. nov., is proposed based on the analysis of nine strains to accommodate Rhodospirillum photometricum, Rhodospirillum sulfurexigens and Rhodospirillum oryzae as Pararhodospirillum photometricum comb. nov., Pararhodospirillum sulfurexigens comb. nov. and Pararhodospirillum oryzae comb. nov., respectively. The type species of the genus is Pararhodospirillum photometricum comb. nov. An emended description of the genus Rhodospirillum is also proposed.

  7. Early Induction and Maintenance of Env-Specific T-Helper Cells following Human Immunodeficiency Virus Type 1 Infection

    PubMed Central

    Malhotra, Uma; Holte, Sarah; Zhu, Tuofu; Delpit, Elizabeth; Huntsberry, Claire; Sette, Alessandro; Shankarappa, Raj; Maenza, Janine; Corey, Lawrence; McElrath, M. Juliana

    2003-01-01

    Mounting evidence points to a role for CD4+ T-helper (Th) cell activities in controlling human immunodeficiency virus type 1 (HIV-1) infection. To determine the induction and evolution of Th responses following acute infection, we prospectively analyzed Env- and Gag-specific Th responses longitudinally for 92 patients with acute (n = 28) or early (n = 64) HIV-1 infection (median, 55 days postinfection [DPI]). The probability of detecting HIV-1-specific lymphoproliferative responses was remarkably low, and when present, the responses were more likely to be Gag specific than Env specific (16 versus 5%). Env-specific responses were significantly more common in patients presenting at <30 DPI than in those presenting at 30 to 365 DPI (21 versus 0.5%, P = 0.001). By contrast, Gag-specific responses occurred with similar frequencies among subjects presenting at <30 DPI and 30 to 365 DPI (13 versus 17%, P = 0.6). After treatment, and regardless of the duration of infection before therapy, Gag-specific Th responses predominated. Furthermore, some acutely infected subjects lost detectable Env-specific Th proliferative responses, which failed to reemerge upon treatment. Detailed analysis for one such subject revealed Env-specific lymphoproliferation at 11 DPI but no detectable Env-specific lymphoproliferation or ex vivo gamma interferon (IFN-γ) secretion at multiple subsequent time points. Env-specific CD4+ T-cell clones from 11 DPI recognized six epitopes in both conserved and variable regions within gp120 and gp41, exhibited major histocompatibility complex-restricted cytotoxicity, and secreted high levels of antiviral cytokines. T-cell receptor clonal transcript analyses and autologous virus sequencing revealed that Th cells induced during acute infection were maintained and there were no Th escape mutations. Subsequent analysis for this subject and six of seven others revealed detectable IFN-γ-secreting cells, but only following in vitro gp160 stimulation. In summary

  8. Direct Probing of Germinal Center Responses Reveals Immunological Features and Bottlenecks for Neutralizing Antibody Responses to HIV Env Trimer.

    PubMed

    Havenar-Daughton, Colin; Carnathan, Diane G; Torrents de la Peña, Alba; Pauthner, Matthias; Briney, Bryan; Reiss, Samantha M; Wood, Jennifer S; Kaushik, Kirti; van Gils, Marit J; Rosales, Sandy L; van der Woude, Patricia; Locci, Michela; Le, Khoa M; de Taeye, Steven W; Sok, Devin; Mohammed, Ata Ur Rasheed; Huang, Jessica; Gumber, Sanjeev; Garcia, AnaPatricia; Kasturi, Sudhir P; Pulendran, Bali; Moore, John P; Ahmed, Rafi; Seumois, Grégory; Burton, Dennis R; Sanders, Rogier W; Silvestri, Guido; Crotty, Shane

    2016-11-22

    Generating tier 2 HIV-neutralizing antibody (nAb) responses by immunization remains a challenging problem, and the immunological barriers to induction of such responses with Env immunogens remain unclear. Here, some rhesus monkeys developed autologous tier 2 nAbs upon HIV Env trimer immunization (SOSIP.v5.2) whereas others did not. This was not because HIV Env trimers were immunologically silent because all monkeys made similar ELISA-binding antibody responses; the key difference was nAb versus non-nAb responses. We explored the immunological barriers to HIV nAb responses by combining a suite of techniques, including longitudinal lymph node fine needle aspirates. Unexpectedly, nAb development best correlated with booster immunization GC B cell magnitude and Tfh characteristics of the Env-specific CD4 T cells. Notably, these factors distinguished between successful and unsuccessful antibody responses because GC B cell frequencies and stoichiometry to GC Tfh cells correlated with nAb development, but did not correlate with total Env Ab binding titers. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  9. HIV-1 Env associates with HLA-C free-chains at the cell membrane modulating viral infectivity

    PubMed Central

    Serena, Michela; Parolini, Francesca; Biswas, Priscilla; Sironi, Francesca; Blanco Miranda, Almudena; Zoratti, Elisa; Scupoli, Maria Teresa; Ziglio, Serena; Valenzuela-Fernandez, Agustin; Gibellini, Davide; Romanelli, Maria Grazia; Siccardi, Antonio; Malnati, Mauro; Beretta, Alberto; Zipeto, Donato

    2017-01-01

    HLA-C has been demonstrated to associate with HIV-1 envelope glycoprotein (Env). Virions lacking HLA-C have reduced infectivity and increased susceptibility to neutralizing antibodies. Like all others MHC-I molecules, HLA-C requires β2-microglobulin (β2m) for appropriate folding and expression on the cell membrane but this association is weaker, thus generating HLA-C free-chains on the cell surface. In this study, we deepen the understanding of HLA-C and Env association by showing that HIV-1 specifically increases the amount of HLA-C free chains, not bound to β2m, on the membrane of infected cells. The association between Env and HLA-C takes place at the cell membrane requiring β2m to occur. We report that the enhanced infectivity conferred to HIV-1 by HLA-C specifically involves HLA-C free chain molecules that have been correctly assembled with β2m. HIV-1 Env-pseudotyped viruses produced in the absence of β2m are less infectious than those produced in the presence of β2m. We hypothesize that the conformation and surface expression of HLA-C molecules could be a discriminant for the association with Env. Binding stability to β2m may confer to HLA-C the ability to preferentially act either as a conventional immune-competent molecule or as an accessory molecule involved in HIV-1 infectivity. PMID:28051183

  10. λ Light Chain Bias Associated With Enhanced Binding and Function of Anti-HIV Env Glycoprotein Antibodies

    PubMed Central

    Sajadi, Mohammad M.; Farshidpour, Maham; Brown, Eric P.; Ouyang, Xin; Seaman, Michael S.; Pazgier, Marzena; Ackerman, Margaret E.; Robinson, Harriet; Tomaras, Georgia; Parsons, Matthew S.; Charurat, Manhattan; DeVico, Anthony L.; Redfield, Robert R.; Lewis, George K.

    2016-01-01

    The humoral response to human immunodeficiency virus (HIV) remains incompletely understood. In this report, we describe biased λ light chain use during the HIV Env glycoprotein (Env) response in HIV infection and vaccination. We examined HIV Env binding (and neutralization) in the context of light chain use in subjects with acute HIV infection, chronic HIV infection, and among HIV vaccinees. In all populations tested, there was a λ chain bias for HIV Env binding antibodies, compared with other HIV antigens (such as p24) or tetanus toxoid. In subjects with chronic HIV infection, a λ bias was noted for neutralization, with λ antibodies accounting for up to 90% of all neutralization activity observed. This is the first report of antibody function in a human infection being tied to light chain use. In HIV infection, antibodies expressing λ light chains tended to have longer CDRL3s, increased light chain contact with HIV Env, and less hypermutation in the heavy chain, compared with antibodies using the κ light chain. These data also support an evolutionary model for the understanding the various κ to λ light chain ratios observed across species and suggest that the λ light chain bias against HIV provides the host an advantage in developing a more efficient humoral response. PMID:26347575

  11. λ Light Chain Bias Associated With Enhanced Binding and Function of Anti-HIV Env Glycoprotein Antibodies.

    PubMed

    Sajadi, Mohammad M; Farshidpour, Maham; Brown, Eric P; Ouyang, Xin; Seaman, Michael S; Pazgier, Marzena; Ackerman, Margaret E; Robinson, Harriet; Tomaras, Georgia; Parsons, Matthew S; Charurat, Manhattan; DeVico, Anthony L; Redfield, Robert R; Lewis, George K

    2016-01-01

    The humoral response to human immunodeficiency virus (HIV) remains incompletely understood. In this report, we describe biased λ light chain use during the HIV Env glycoprotein (Env) response in HIV infection and vaccination. We examined HIV Env binding (and neutralization) in the context of light chain use in subjects with acute HIV infection, chronic HIV infection, and among HIV vaccinees. In all populations tested, there was a λ chain bias for HIV Env binding antibodies, compared with other HIV antigens (such as p24) or tetanus toxoid. In subjects with chronic HIV infection, a λ bias was noted for neutralization, with λ antibodies accounting for up to 90% of all neutralization activity observed. This is the first report of antibody function in a human infection being tied to light chain use. In HIV infection, antibodies expressing λ light chains tended to have longer CDRL3s, increased light chain contact with HIV Env, and less hypermutation in the heavy chain, compared with antibodies using the κ light chain. These data also support an evolutionary model for the understanding the various κ to λ light chain ratios observed across species and suggest that the λ light chain bias against HIV provides the host an advantage in developing a more efficient humoral response. © The Author 2015. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  12. HIV-1 Env associates with HLA-C free-chains at the cell membrane modulating viral infectivity.

    PubMed

    Serena, Michela; Parolini, Francesca; Biswas, Priscilla; Sironi, Francesca; Blanco Miranda, Almudena; Zoratti, Elisa; Scupoli, Maria Teresa; Ziglio, Serena; Valenzuela-Fernandez, Agustin; Gibellini, Davide; Romanelli, Maria Grazia; Siccardi, Antonio; Malnati, Mauro; Beretta, Alberto; Zipeto, Donato

    2017-01-04

    HLA-C has been demonstrated to associate with HIV-1 envelope glycoprotein (Env). Virions lacking HLA-C have reduced infectivity and increased susceptibility to neutralizing antibodies. Like all others MHC-I molecules, HLA-C requires β2-microglobulin (β2m) for appropriate folding and expression on the cell membrane but this association is weaker, thus generating HLA-C free-chains on the cell surface. In this study, we deepen the understanding of HLA-C and Env association by showing that HIV-1 specifically increases the amount of HLA-C free chains, not bound to β2m, on the membrane of infected cells. The association between Env and HLA-C takes place at the cell membrane requiring β2m to occur. We report that the enhanced infectivity conferred to HIV-1 by HLA-C specifically involves HLA-C free chain molecules that have been correctly assembled with β2m. HIV-1 Env-pseudotyped viruses produced in the absence of β2m are less infectious than those produced in the presence of β2m. We hypothesize that the conformation and surface expression of HLA-C molecules could be a discriminant for the association with Env. Binding stability to β2m may confer to HLA-C the ability to preferentially act either as a conventional immune-competent molecule or as an accessory molecule involved in HIV-1 infectivity.

  13. Mucosal immunization of sheep with a Maedi-Visna virus (MVV) env DNA vaccine protects against early MVV productive infection.

    PubMed

    González, Belén; Reina, Ramsés; García, Iker; Andrés, Sara; Glaria, Idoia; Alzueta, María; Mora, María Isabel; Jugo, Begoña M; Arrieta-Aguirre, Inés; de la Lastra, José M Pérez; Rodríguez, Dolores; Rodríguez, Juan Ramón; Esteban, Mariano; Grilló, María Jesús; Blacklaws, Barbara A; Harkiss, Gordon D; Chebloune, Yahia; Luján, Lluís; de Andrés, Damián; Amorena, Beatriz

    2005-07-29

    Gene gun mucosal DNA immunization of sheep with a plasmid expressing the env gene of Maedi-Visna virus (MVV) was used to examine the protection against MVV infection in sheep from a naturally infected flock. For immunization, sheep were primed with a pcDNA plasmid (pcDNA-env) encoding the Env glycoproteins of MVV and boosted with combined pcDNA-env and pCR3.1-IFN-gamma plasmid inoculations. The pcDNA plasmid used in the control group contained the lacZ coding sequences instead of the env gene. Within a month post-challenge, the viral load in the vaccinated group was lower (p < or = 0.05) and virus was only detected transiently compared with the control group. Furthermore, 2 months later, neutralizing antibodies (NtAb) were detected in all the control animals and none of the vaccinated animals (p < or = 0.01). These results demonstrated a significant early protective effect of this immunization strategy against MVV infection that restricts the virus replication following challenge in the absence of NtAb production. This vaccine protective effect against MVV infection disappeared after two years post-challenge, when active replication of MVV challenge strain was observed. Protection conferred by the vaccine could not be explained by OLA DRB1 allele or genotype differences. Most of the individuals were DRB1 heterozygous and none was totally resistant to infection.

  14. Activation of HERV-K Env protein is essential for tumorigenesis and metastasis of breast cancer cells.

    PubMed

    Zhou, Fuling; Li, Ming; Wei, Yongchang; Lin, Kevin; Lu, Yue; Shen, Jianjun; Johanning, Gary L; Wang-Johanning, Feng

    2016-12-20

    Human endogenous retrovirus type K (HERV-K) Env protein was previously demonstrated to be overexpressed in human breast cancer (BC) cells and tissues. However, the molecular pathways driving the specific alterations are unknown. We now show that knockdown of its expression with an shRNA (shRNAenv) blocked BC cell proliferation, migration, and invasion. shRNAenv transduction also attenuated the ability of BC cells to form tumors, and notably prevented metastasis. Mechanistically, downregulation of HERV-K blocked expression of tumor-associated genes that included Ras, p-RSK, and p-ERK. The major upstream regulators influenced by HERV-K knockdown were p53, TGF- β1, and MYC. Of interest, when the HERV-K env gene was overexpressed in shRNAenv-transduced BC cells using an HERV-K env expression vector, Ras/Raf/MEK/ERK pathway signaling was restored. CDK5, which alters p53 phosphorylation in some cancers, was upregulated and p53 was downregulated when HERV-K was overexpressed. CDK5 is also a mediator of TGF-β1-induced epithelial-mesenchymal transition and migration in cancer cells, and is involved in tumor formation. Importantly, reductions in migration, invasion, and transformation of BC cells stably transduced with shRNAenv was reversed after adding back a vector with a synonymous mutation of HERV-K env. Taken together, these results indicate that HERV-K Env protein plays an important role in tumorigenesis and metastasis of BC.

  15. Superiority in Rhesus Macaques of Targeting HIV-1 Env Gp140 to CD40 Versus LOX-1 in Combination with Replication Competent NYVAC-KC for Induction of Env-Specific Antibody and T Cell Responses.

    PubMed

    Zurawski, Gerard; Shen, Xiaoying; Zurawski, Sandra; Tomaras, Georgia D; Montefiori, David C; Roederer, Mario; Ferrari, Guido; Lacabaratz, Christine; Klucar, Peter; Wang, Zhiqing; Foulds, Kathryn E; Kao, Shing-Fen; Yu, Xuesong; Sato, Alicia; Yates, Nicole L; LaBranche, Celia; Stanfield-Oakley, Sherry; Kibler, Karen; Jacobs, Bertram; Salazar, Andres; Self, Steve; Fulp, Jimmy; Gottardo, Raphael; Galmin, Lindsey; Weiss, Deborah; Cristillo, Anthony; Pantaleo, Giuseppe; Levy, Yves

    2017-02-15

    We compared the HIV-1-specific immune responses generated by targeting HIV-1 envelope protein (Env gp140) to either CD40 or LOX-1, two endocytic receptors on dendritic cells (DCs), in Rhesus macaques primed with a poxvirus vector (NYVAC-KC) expressing Env gp140. The DC-targeting vaccines, humanized recombinant monoclonal antibodies fused to Env gp140, were administered as a boost with poly ICLC adjuvant either alone or co-administered with the NYVAC-KC vector. All the DC-targeting vaccine administrations with poly ICLC increased the low-level serum anti-Env IgG responses elicited by NYVAC-KC priming significantly more (up to P =0.01) than a group without poly ICLC. The responses were robust, cross-reactive, and contained antibodies specific to multiple epitopes within gp140 including the C1, C2, V1-3, C4, C5, and gp41 immuno-dominant regions. The DC-targeting vaccines also elicited modest serum Env-specific IgA responses. All groups gave serum neutralization activity limited to Tier 1 viruses and antibody dependent cytotoxicity responses (ADCC) after DC-targeting boosts. Furthermore, CD4(+) and CD8(+) T cell responses specific to multiple Env epitopes were strongly boosted by the DC-targeting vaccines + poly ICLC. Together, these results indicate that prime/boost immunization via NYVAC-KC and either αCD40.Env gp140/poly ICLC or αLOX-1.Env gp140/poly ICLC induced balanced antibody and T cell responses against HIV-1 Env. Co-administration of NYVAC-KC with the DC-targeting vaccines increased T cell responses, but had minimal effects on antibody responses except for suppressing serum IgA responses. Overall, compared to LOX-1, targeting Env to CD40 gave more robust T cell and serum antibody responses with broader epitope representation and greater durability.IMPORTANCE An effective vaccine to prevent HIV-1 infection does not yet exist. An approach to elicit strong protective antibody development is to direct virus protein antigens specifically to dendritic cells

  16. Distributed Generation Market Demand Model (dGen): Documentation

    SciTech Connect

    Sigrin, Benjamin; Gleason, Michael; Preus, Robert; Baring-Gould, Ian; Margolis, Robert

    2016-02-01

    The Distributed Generation Market Demand model (dGen) is a geospatially rich, bottom-up, market-penetration model that simulates the potential adoption of distributed energy resources (DERs) for residential, commercial, and industrial entities in the continental United States through 2050. The National Renewable Energy Laboratory (NREL) developed dGen to analyze the key factors that will affect future market demand for distributed solar, wind, storage, and other DER technologies in the United States. The new model builds off, extends, and replaces NREL's SolarDS model (Denholm et al. 2009a), which simulates the market penetration of distributed PV only. Unlike the SolarDS model, dGen can model various DER technologies under one platform--it currently can simulate the adoption of distributed solar (the dSolar module) and distributed wind (the dWind module) and link with the ReEDS capacity expansion model (Appendix C). The underlying algorithms and datasets in dGen, which improve the representation of customer decision making as well as the spatial resolution of analyses (Figure ES-1), also are improvements over SolarDS.

  17. Summary of CPAS Gen II Parachute Analysis

    NASA Technical Reports Server (NTRS)

    Morris, Aaron L.; Bledsoe, Kristin J.; Fraire, Usbaldo, Jr.; Moore, James W.; Olson, Leah M.; Ray, Eric

    2011-01-01

    The Orion spacecraft is currently under development by NASA and Lockheed Martin. Like Apollo, Orion will use a series of parachutes to slow its descent and splashdown safely. The Orion parachute system, known as the CEV Parachute Assembly System (CPAS), is being designed by NASA, the Engineering and Science Contract Group (ESCG), and Airborne Systems. The first generation (Gen I) of CPAS testing consisted of thirteen tests and was executed in the 2007-2008 timeframe. The Gen I tests provided an initial understanding of the CPAS parachutes. Knowledge gained from Gen I testing was used to plan the second generation of testing (Gen II). Gen II consisted of six tests: three singleparachute tests, designated as Main Development Tests, and three Cluster Development Tests. Gen II required a more thorough investigation into parachute performance than Gen I. Higher fidelity instrumentation, enhanced analysis methods and tools, and advanced test techniques were developed. The results of the Gen II test series are being incorporated into the CPAS design. Further testing and refinement of the design and model of parachute performance will occur during the upcoming third generation of testing (Gen III). This paper will provide an overview of the developments in CPAS analysis following the end of Gen I, including descriptions of new tools and techniques as well as overviews of the Gen II tests.

  18. Ultrastructural evidence of an interaction between Env and Gag proteins during assembly of HIV type 1.

    PubMed

    Bugelski, P J; Maleeff, B E; Klinkner, A M; Ventre, J; Hart, T K

    1995-01-01

    Assembly and budding of retroviruses is believed to involve a complex interaction of envelope and capsid proteins at the host cell membrane. The nature of these interactions is, however, incompletely understood. Studies of the topography of the surface of HIV-1 have shown that the envelope glycoprotein projections (knobs) are arranged in a T = 7 levo rotational symmetry. Similarly, an icosahedral structure has been suggested for the p17 matrix of HIV-1. In an effort to investigate whether there is a structural interaction between these molecules, virions whose maturation was blocked by an inhibitor of HIV protease were studied using cytochemistry, morphometry, and 2D fast Fourier transform image enhancement. Analysis of the relationship between core morphology and the topographic distribution of envelope glycoprotein projections on HIV-1 provided structural evidence of an interaction between Env and Gag proteins. Furthermore, image enhancement revealed a periodic substructure in the Pr55gag plaque. Taken together, the data suggest an interaction between Pr55gag and the gp120-gp41 complex during assembly and budding of HIV-1. This interaction may, in part, contribute to determining the amount of Env glycoprotein that will be incorporated into a virion, and therefore play a role in the biology of HIV-1.

  19. Avian hemangioma retrovirus induces cell proliferation via the envelope (env) gene.

    PubMed

    Alian, A; Sela-Donenfeld, D; Panet, A; Eldor, A

    2000-10-10

    Several years ago, a field strain retrovirus, avian hemangioma virus (AHV), was isolated from hemangioma tumors in layer hens. Sequence analysis indicated that the AHV genome contains the three prototypic retroviral genes, gag, pol, and env, and is devoid of an oncogene. In cultured endothelial cells, however, AHV induced a significant cytopathic effect through a typical apoptotic cascade. We now demonstrate that AHV also induces cell proliferation and anchorage-independent growth of BSC-1 epithelial cells and NIH-3T3 fibroblasts. This was shown by measurements of (1) cell viability, (2) DNA synthesis, (3) flow cytometry analysis of the cell DNA content, and (4) clonogenic efficiency of the infected cells. Anchorage-independent cell growth was demonstrated by colony formation in soft agar. Moreover, the AHV env gene was cloned into a MuLV-based retroviral vector, and infection of NIH-3T3 cells with this vector induced cell proliferation as well as clonogenic growth. These results suggest that AHV, which is devoid of an oncogene, is a pleiotropic activator capable of inducing either apoptosis or cellular proliferation, depending on the infected cell type. Copyright 2000 Academic Press.

  20. Structural insights into key sites of vulnerability on HIV-1 Env and influenza HA.

    PubMed

    Julien, Jean-Philippe; Lee, Peter S; Wilson, Ian A

    2012-11-01

    Human immunodeficiency virus-1 (HIV-1) envelope protein (Env) and influenza hemagglutinin (HA) are the surface glycoproteins responsible for viral entry into host cells, the first step in the virus life cycle necessary to initiate infection. These glycoproteins exhibit a high degree of sequence variability and glycosylation, which are used as strategies to escape host immune responses. Nonetheless, antibodies with broadly neutralizing activity against these viruses have been isolated that have managed to overcome these barriers. Here, we review recent advances in the structural characterization of these antibodies with their viral antigens that defines a few sites of vulnerability on these viral spikes. These broadly neutralizing antibodies tend to focus their recognition on the sites of similar function between the two viruses: the receptor-binding site and membrane fusion machinery. However, some sites of recognition are unique to the virus neutralized, such as the dense shield of oligomannose carbohydrates on HIV-1 Env. These observations are discussed in the context of structure-based design strategies to aid in vaccine design or development of antivirals.

  1. The Streptomycin-Treated Mouse Intestine Selects Escherichia coli envZ Missense Mutants That Interact with Dense and Diverse Intestinal Microbiota

    PubMed Central

    Leatham-Jensen, Mary P.; Frimodt-Møller, Jakob; Adediran, Jimmy; Mokszycki, Matthew E.; Banner, Megan E.; Caughron, Joyce E.; Krogfelt, Karen A.; Conway, Tyrrell

    2012-01-01

    Previously, we reported that the streptomycin-treated mouse intestine selected nonmotile Escherichia coli MG1655 flhDC deletion mutants of E. coli MG1655 with improved colonizing ability that grow 15% faster in vitro in mouse cecal mucus and 15 to 30% faster on sugars present in mucus (M. P. Leatham et al., Infect. Immun. 73:8039–8049, 2005). Here, we report that the 10 to 20% remaining motile E. coli MG1655 are envZ missense mutants that are also better colonizers of the mouse intestine than E. coli MG1655. One of the flhDC mutants, E. coli MG1655 ΔflhD, and one of the envZ missense mutants, E. coli MG1655 mot-1, were studied further. E. coli MG1655 mot-1 is more resistant to bile salts and colicin V than E. coli MG1655 ΔflhD and grows ca. 15% slower in vitro in mouse cecal mucus and on several sugars present in mucus compared to E. coli MG1655 ΔflhD but grows 30% faster on galactose. Moreover, E. coli MG1655 mot-1 and E. coli MG1655 ΔflhD appear to colonize equally well in one intestinal niche, but E. coli MG1655 mot-1 appears to use galactose to colonize a second, smaller intestinal niche either not colonized or colonized poorly by E. coli MG1655 ΔflhD. Evidence is also presented that E. coli MG1655 is a minority member of mixed bacterial biofilms in the mucus layer of the streptomycin-treated mouse intestine. We offer a hypothesis, which we call the “Restaurant” hypothesis, that explains how nutrient acquisition in different biofilms comprised of different anaerobes can account for our results. PMID:22392928

  2. HIV-1 Env- and Vpu-Specific Antibody-Dependent Cellular Cytotoxicity Responses Associated with Elite Control of HIV.

    PubMed

    Madhavi, Vijaya; Wines, Bruce D; Amin, Janaki; Emery, Sean; Lopez, Ester; Kelleher, Anthony; Center, Rob J; Hogarth, P Mark; Chung, Amy W; Kent, Stephen J; Stratov, Ivan

    2017-09-15

    Studying HIV-infected individuals who control HIV replication (elite controllers [ECs]) enables exploration of effective anti-HIV immunity. HIV Env-specific and non-Env-specific antibody-dependent cellular cytotoxicity (ADCC) may contribute to protection from progressive HIV infection, but the evidence is limited. We recruited 22 ECs and matched them with 44 viremic subjects. HIV Env- and Vpu-specific ADCC responses in sera were studied using a novel enzyme-linked immunosorbent assay (ELISA)-based dimeric recombinant soluble FcγRIIIa (rsFcγRIIIa)-binding assay, surface plasmon resonance, antibody-dependent natural killer (NK) cell activation assays, and ADCC-mediated killing assays. ECs had higher levels of HIV Env-specific antibodies capable of binding FcγRIIIa, activating NK cells, and mediating granzyme B activity (all P < 0.01) than viremic subjects. ECs also had higher levels of antibodies against a C-terminal 13-mer Vpu peptide capable of mediating FcγRIIIa binding and NK cell activation than viremic subjects (both P < 0.05). Our data associate Env-specific and Vpu epitope-specific ADCC in effective immune responses against HIV among ECs. Our findings have implications for understanding the role of ADCC in HIV control.IMPORTANCE Understanding immune responses associated with elite control of HIV may aid the development of immunotherapeutic and vaccine strategies for controlling HIV infection. Env is a major HIV protein target of functional antibody responses that are heightened in ECs. Interestingly, EC antibodies also target Vpu, an accessory protein crucial to HIV, which degrades CD4 and antagonizes tetherin. Antibodies specific to Vpu are a common feature of the immune response of ECs that may prove to be of functional importance to the design of improved ADCC-based immunotherapy and preventative HIV vaccines. Copyright © 2017 American Society for Microbiology.

  3. Antidiabetic, antidyslipidemic and toxicity profile of ENV-2: A potent pyrazole derivative against diabetes and related diseases.

    PubMed

    Hernández-Vázquez, Eduardo; Ocampo-Montalban, Hugo; Cerón-Romero, Litzia; Cruz, Miguel; Gómez-Zamudio, Jaime; Hiriart-Valencia, Guadalupe; Villalobos-Molina, Rafael; Flores-Flores, Angelica; Estrada-Soto, Samuel

    2017-03-18

    Diabetes is a major health problem and a predisposition factor for further degenerative complications and, therefore, novel therapies are urgently needed. Currently, cannabinoid receptor 1 (CB1 receptor) antagonists have been considered as promissory entities for metabolic disorders treatment. Accordingly, the purpose of this work was the evaluation of the sub-acute antidiabetic, anti-hyperglycemic, antidyslipidemic and toxicological profile of ENV-2, a potent hypoglycemic and antioxidant CB1 receptor antagonist. In this study, ENV-2 showed a pronounced anti-hyperglycemic effect even at a dose of 5mg/kg (P<0.05) in a glucose tolerance test on normoglycemic rats. Moreover, after administration of ENV-2 (16mg/kg) to diabetic rats, a prominent antidiabetic activity was observed (P<0.05), which was higher than glibenclamide. Sub-acute treatment (10 days) of ENV-2 resulted in a significant reduction of plasma glucose (P<0.05). Also, the levels of peripheral lipids were improved; blood triacylglycerols (TG) and cholesterol (CHOL) were diminished (P<0.05). In addition, it was found that ENV-2 reduced IL-1β and IL-18 mRNA expression in adipose tissue (P<0.05). Due to the satisfactory outcomes, we were interested in evaluating the toxicity of ENV-2 in both acute and sub-chronic approaches. Regarding the acute administration, the compound resulted to be non-toxic and was grouped in category 5 according to OECD. It was also found that sub-chronic administration did not increase the size of the studied organs, while no structural damage was observed in heart, lung, liver and kidney tissues. Finally, neither AST nor ALT damage hepatic markers were augmented.

  4. From AWE-GEN to AWE-GEN-2d: a high spatial and temporal resolution weather generator

    NASA Astrophysics Data System (ADS)

    Peleg, Nadav; Fatichi, Simone; Paschalis, Athanasios; Molnar, Peter; Burlando, Paolo

    2016-04-01

    A new weather generator, AWE-GEN-2d (Advanced WEather GENerator for 2-Dimension grid) is developed following the philosophy of combining physical and stochastic approaches to simulate meteorological variables at high spatial and temporal resolution (e.g. 2 km x 2 km and 5 min for precipitation and cloud cover and 100 m x 100 m and 1 h for other variables variable (temperature, solar radiation, vapor pressure, atmospheric pressure and near-surface wind). The model is suitable to investigate the impacts of climate variability, temporal and spatial resolutions of forcing on hydrological, ecological, agricultural and geomorphological impacts studies. Using appropriate parameterization the model can be used in the context of climate change. Here we present the model technical structure of AWE-GEN-2d, which is a substantial evolution of four preceding models (i) the hourly-point scale Advanced WEather GENerator (AWE-GEN) presented by Fatichi et al. (2011, Adv. Water Resour.) (ii) the Space-Time Realizations of Areal Precipitation (STREAP) model introduced by Paschalis et al. (2013, Water Resour. Res.), (iii) the High-Resolution Synoptically conditioned Weather Generator developed by Peleg and Morin (2014, Water Resour. Res.), and (iv) the Wind-field Interpolation by Non Divergent Schemes presented by Burlando et al. (2007, Boundary-Layer Meteorol.). The AWE-GEN-2d is relatively parsimonious in terms of computational demand and allows generating many stochastic realizations of current and projected climates in an efficient way. An example of model application and testing is presented with reference to a case study in the Wallis region, a complex orography terrain in the Swiss Alps.

  5. Putative Phosphatidylinositol 3-Kinase (PI3K) Binding Motifs in Ovine Betaretrovirus Env Proteins Are Not Essential for Rodent Fibroblast Transformation and PI3K/Akt Activation

    PubMed Central

    Liu, Shan-Lu; Lerman, Michael I.; Miller, A. Dusty

    2003-01-01

    Jaagsiekte sheep retrovirus (JSRV) and enzootic nasal tumor virus (ENTV) are simple betaretroviruses that cause epithelial cell tumors in the lower and upper airways of sheep and goats. The envelope (Env) glycoproteins of both viruses can transform rodent and chicken fibroblasts, indicating that they play an essential role in oncogenesis. Previous studies found that a YXXM motif in the Env cytoplasmic tail, a putative docking site for phosphatidylinositol 3-kinase (PI3K) after tyrosine phosphorylation, was necessary for rodent cell transformation but was not required for transformation of DF-1 chicken fibroblasts. Here we show that JSRV and ENTV Env proteins with tyrosine or methionine mutations in the YXXM motif can still transform rodent fibroblasts, albeit with reduced efficiency. Akt was activated in cells transformed by JSRV or ENTV Env proteins and in cells transformed by the proteins with tyrosine mutations. Furthermore, the PI3K-specific inhibitor LY294002 could inhibit Akt activation and cell transformation in all cases, indicating that Akt activation and transformation is PI3K dependent. However, we could not detect tyrosine phosphorylation of JSRV or ENTV Env proteins or an interaction between the Env proteins and PI3K in the transformed cells. We found no evidence for mitogen-activated protein kinase activation in cells that were transformed by the JSRV or ENTV Env proteins. We conclude that ovine betaretrovirus Env proteins transform the rodent fibroblasts by indirectly activating the PI3K/Akt pathway. PMID:12829832

  6. Impact of antibody quality and anamnestic response on viremia control post-challenge in a combined Tat/Env vaccine regimen in rhesus macaques

    PubMed Central

    Demberg, Thorsten; Brocca-Cofano, Egidio; Kuate, Seraphin; Aladi, Stanley; Vargas-Inchaustegui, Diego A.; Venzon, David; Kalisz, Irene; Kalyanaraman, V.S.; Lee, Eun Mi; Pal, Ranajit; DiPasquale, Janet; Ruprecht, Ruth M.; Montefiori, David C.; Srivastava, Indresh; Barnett, Susan W.; Robert-Guroff, Marjorie

    2013-01-01

    Previously, priming rhesus macaques with Adenovirus type 5 host range mutant-recombinants encoding Tat and Env and boosting with Tat and Env protein in MPL-SE controlled chronic viremia by 4 logs following homologous intravenous SHIV89.6P challenge. Here we evaluated Tat, Env, and Tat/Env regimens for immunogenicity and protective efficacy using clade C Env, alum adjuvant, and a heterologous intrarectal SHIV1157ipd3N4 challenge. Despite induction of strong cellular and humoral immunity, Tat/Env group T and B-cell memory responses were not significantly enhanced over Tat- or Env-only groups. Lack of viremia control post-challenge was attributed to lower avidity Env antibodies and no anamnestic ADCC response or SHIV1157ipd3N4 neutralizing antibody development post-challenge. Poor biologic activity of the Tat immunogen may have impaired Tat immunity. In the absence of sterilizing immunity, strong anamnestic responses to heterologous virus can help control viremia. Both antibody breadth and optimal adjuvanticity are needed to elicit high-quality antibody for protective efficacy. PMID:23528732

  7. Dense display of HIV-1 envelope spikes on the lambda phage scaffold does not result in the generation of improved antibody responses to HIV-1 Env.

    PubMed

    Mattiacio, Jonelle; Walter, Scott; Brewer, Matt; Domm, William; Friedman, Alan E; Dewhurst, Stephen

    2011-03-21

    The generation of strong, virus-neutralizing antibody responses to the HIV-1 envelope spike (Env) is a major goal in HIV-1 vaccine research. To try to enhance the Env-specific response, we displayed oligomeric gp140 on a virus-like scaffold provided by the lambda phage capsid. To do this, an in vitro complementation system was used to "decorate" phage particles with glycosylated, mammalian cell-derived envelope oligomers. We compared the immune response to lambda phage particles displaying HIV-1 Env to that elicited by soluble oligomeric gp140 in rabbits. Env-binding antibody titers were higher in animals that received oligomeric gp140 as compared to Env decorated phage particles, as were virus neutralizing antibody responses. The Env decorated phage particles were, however, able to efficiently boost a protein-primed humoral response to levels equivalent to those elicited by high-dose adjuvanted Env oligomers. These results show that display of HIV-1 envelope spikes on the bacteriophage lambda capsid does not result in an improved, Env-specific humoral immune response. Copyright © 2011 Elsevier Ltd. All rights reserved.

  8. Effect of point mutations in the N terminus of the lentivirus lytic peptide-1 sequence of human immunodeficiency virus type 1 transmembrane protein gp41 on Env stability.

    PubMed

    Lee, Sheau-Fen; Ko, Chiung-Yuan; Wang, Chin-Tien; Chen, Steve S-L

    2002-05-03

    To understand the role of the lentivirus lytic peptide-1 region of the human immunodeficiency virus type 1 transmembrane glycoprotein (gp) 41 in viral infection, we examined the effects on virus replication of single amino acid deletions spanning this region in an infectious provirus of the HXB2 strain. Among the mutants analyzed, only the deletion of one of the two adjacent valine residues located at positions 832 and 833 (termed the Delta 833 mutant for simplicity) greatly reduced the steady-state, cell-associated levels of the Env precursor and gp120, as opposed to the wild-type virus. The altered Env phenotype resulted in severely impaired virus infectivity and gp120 incorporation into this mutant virion. Analyses of additional mutants with deletions at Ile-830, Ala-836, and Ile-840 demonstrated that the Delta 830 mutant exhibited the most significant inhibitory effect on Env steady-state expression. These results indicate that the N terminus of the lentivirus lytic peptide-1 region is critical for Env steady-state expression. Among the mutant viruses encoding Env proteins in which residues Val-832 and Val-833 were individually substituted by nonconserved amino acids Ala, Ser, or Pro, which were expected to disrupt the alpha-helical structure in the increasingly severe manner of Pro > Ser > Ala, only the 833P mutant exhibited significantly reduced steady-state Env expression. Pulse labeling and pulse-chase studies demonstrated that the Delta 830, Delta 833, and 833P mutants of Env proteins degraded more rapidly in a time-dependent manner after biosynthesis than did the wild-type Env. The results indicate that residue 830 and 833 mutations are likely to induce a conformational change in Env that targets the mutant protein for cellular degradation. Our study has implications about the structural determinants located at the N terminus of the lentivirus lytic peptide-1 sequence of gp41 that affect the fate of Env in virus-infected cells.

  9. A novel gene that interferes with the phosphotransfer signal transduction mediated by the EnvZ osmosensor in Escherichia coli.

    PubMed

    Hirokawa, K; Ogino, T; Aiba, H; Mizuno, T

    1996-10-01

    In Escherichia coli, expression of the major outer membrane proteins, OmpC and OmpF, is regulated in response to the medium osmolarity and other environmental stimuli. A two-component signal transduction system, mediated by EnvZ and OmpR, is crucially responsible for this osmotic regulation of the ompC and ompF genes. In this study, an E. coli gene was cloned, which interferes with expression of both the ompC and ompF genes at the level of transcription, provided that the cloned gene was introduced in E. coli cells by a multicopy plasmid. The gene product was identified as F107, which was previously characterized as a hypothetical protein in E. coli genome databases. F107 containing 107 amino acids appears to be highly hydrophobic, and has a sequence similarity to the eukaryotic type of cytochrome-c oxidase subunit III. The mechanism by which F107 inhibits transcription of ompC and ompF was examined extensively, mainly by using a set of envZ and ompR mutants. These results suggested that F107 interferes specifically with a function of the EnvZ osmosensory kinase. Possible mechanisms by which F107 affects the EnvZ function are discussed.

  10. Specifically modified Env immunogens activate B-cell precursors of broadly neutralizing HIV-1 antibodies in transgenic mice

    PubMed Central

    McGuire, Andrew T.; Gray, Matthew D.; Dosenovic, Pia; Gitlin, Alexander D.; Freund, Natalia T.; Petersen, John; Correnti, Colin; Johnsen, William; Kegel, Robert; Stuart, Andrew B.; Glenn, Jolene; Seaman, Michael S.; Schief, William R.; Strong, Roland K.; Nussenzweig, Michel C.; Stamatatos, Leonidas

    2016-01-01

    VRC01-class broadly neutralizing HIV-1 antibodies protect animals from experimental infection and could contribute to an effective vaccine response. Their predicted germline forms (gl) bind Env inefficiently, which may explain why they are not elicited by HIV-1 Env-immunization. Here we show that an optimized Env immunogen can engage multiple glVRC01-class antibodies. Furthermore, this immunogen activates naive B cells expressing the human germline heavy chain of 3BNC60, paired with endogenous mouse light chains in vivo. To address whether it activates B cells expressing the fully humanized gl3BNC60 B-cell receptor (BCR), we immunized mice carrying both the heavy and light chains of gl3BNC60. B cells expressing this BCR display an autoreactive phenotype and fail to respond efficiently to soluble forms of the optimized immunogen, unless it is highly multimerized. Thus, specifically designed Env immunogens can activate naive B cells expressing human BCRs corresponding to precursors of broadly neutralizing HIV-1 antibodies even when the B cells display an autoreactive phenotype. PMID:26907590

  11. Specifically modified Env immunogens activate B-cell precursors of broadly neutralizing HIV-1 antibodies in transgenic mice.

    PubMed

    McGuire, Andrew T; Gray, Matthew D; Dosenovic, Pia; Gitlin, Alexander D; Freund, Natalia T; Petersen, John; Correnti, Colin; Johnsen, William; Kegel, Robert; Stuart, Andrew B; Glenn, Jolene; Seaman, Michael S; Schief, William R; Strong, Roland K; Nussenzweig, Michel C; Stamatatos, Leonidas

    2016-02-24

    VRC01-class broadly neutralizing HIV-1 antibodies protect animals from experimental infection and could contribute to an effective vaccine response. Their predicted germline forms (gl) bind Env inefficiently, which may explain why they are not elicited by HIV-1 Env-immunization. Here we show that an optimized Env immunogen can engage multiple glVRC01-class antibodies. Furthermore, this immunogen activates naive B cells expressing the human germline heavy chain of 3BNC60, paired with endogenous mouse light chains in vivo. To address whether it activates B cells expressing the fully humanized gl3BNC60 B-cell receptor (BCR), we immunized mice carrying both the heavy and light chains of gl3BNC60. B cells expressing this BCR display an autoreactive phenotype and fail to respond efficiently to soluble forms of the optimized immunogen, unless it is highly multimerized. Thus, specifically designed Env immunogens can activate naive B cells expressing human BCRs corresponding to precursors of broadly neutralizing HIV-1 antibodies even when the B cells display an autoreactive phenotype.

  12. Natively glycosylated HIV-1 Env structure reveals new mode for antibody recognition of the CD4-binding site

    PubMed Central

    West, Anthony P; Schamber, Michael; Gazumyan, Anna; Golijanin, Jovana; Seaman, Michael S; Fätkenheuer, Gerd; Klein, Florian; Nussenzweig, Michel C; Bjorkman, Pamela J

    2016-01-01

    HIV-1 vaccine design is informed by structural studies elucidating mechanisms by which broadly neutralizing antibodies (bNAbs) recognize and/or accommodate N-glycans on the trimeric envelope glycoprotein (Env). Variability in high-mannose and complex-type Env glycoforms leads to heterogeneity that usually precludes visualization of the native glycan shield. We present 3.5-Å- and 3.9-Å-resolution crystal structures of the HIV-1 Env trimer with fully processed and native glycosylation, revealing a glycan shield of high-mannose and complex-type N-glycans, which we used to define complete epitopes of two bNAbs. Env trimer was complexed with 10-1074 (against the V3-loop) and IOMA, a new CD4-binding site (CD4bs) antibody. Although IOMA derives from VH1-2*02, the germline gene of CD4bs-targeting VRC01-class bNAbs, its light chain lacks the short CDRL3 that defines VRC01-class bNAbs. Thus IOMA resembles 8ANC131-class/VH1-46–derived CD4bs bNAbs, which have normal-length CDRL3s. The existence of bNAbs that combine features of VRC01-class and 8ANC131-class antibodies has implications for immunization strategies targeting VRC01-like bNAbs. PMID:27617431

  13. Hypothyroid Graves' disease complicated with elephantiasis nostras verrucosa (ENV): a case report and review of the literature.

    PubMed

    Ukinç, Kubilay; Bayraktar, Miyase; Gedik, Arzu

    2009-08-01

    Thyroid dermopathy is not a frequent feature of hyperthyroid Graves' disease, being present in less than 5% of the patients. Graves' disease has been shown to exist in euthyroid or hypothyroid forms in untreated patients. Here, we describe a case of hypothyroid Graves' disease with elephantiasis nostras verrucosa (ENV), which is an extreme form of thyroid dermopathy (TD). A 58-year-old female patient was admitted to the emergency department with somnolence, hypothermia, and bradycardia. Her mental status gradually worsened, resulting in a deep coma. She was intubated and followed in the intensive care unit, as she needed mechanical ventilatory assistance due to respiratory failure. She also had bilateral non-pitting edema, a cobblestone-like appearance, and hyperkeratotic greenish-brown-colored lesions in the pretibial and dorsal regions of the feet that were compatible with ENV. Hypothyroid Graves' disease is a very rare condition among autoimmune thyroid disorders, and ENV is an extremely rare form of TD. Here, we present a patient with hypothyroid Graves' disease and ENV.

  14. HIV-1 Env-Specific Memory and Germinal Center B Cells in C57BL/6 Mice

    PubMed Central

    Soldemo, Martina; Pedersen, Gabriel K.; Hedestam, Gunilla B. Karlsson

    2014-01-01

    Continued efforts to define the immunogenic properties of the HIV-1 envelope glycoproteins (Env) are needed to elicit effective antibody (Ab) responses by vaccination. HIV-1 is a highly neutralization-resistant virus due to conformational and glycan shielding of conserved Ab determinants on the virus spike. Elicitation of broadly neutralizing Abs that bind poorly accessible epitope regions on Env is therefore extremely challenging and will likely require selective targeting of specific sub-determinants. To evaluate such approaches there is a pressing need for in vivo studies in both large and small animals, including mice. Currently, most mouse immunization studies are performed in the BALB/c strain; however, the C57BL/6 strain offers improved possibilities for mechanistic studies due to the availability of numerous knock-out strains on this genetic background. Here, we compared Env immunogenicity in BALB/c and C57BL/6 mice and found that the magnitude of the antigen-specific response was somewhat lower in C57BL/6 than in BALB/c mice by ELISA but not significantly different by B cell ELISpot measurements. We then established protocols for the isolation of single Env-specific memory B cells and germinal center (GC) B cells from immunized C57BL/6 mice to facilitate future studies of the elicited response at the monoclonal Ab level. We propose that these protocols can be used to gain an improved understanding of the early recruitment of Env-specific B cells to the GC as well as the archiving of such responses in the memory B cell pool following immunization. PMID:25198199

  15. Structural and functional studies of the HAMP domain of EnvZ, an osmosensing transmembrane histidine kinase in Escherichia coli.

    PubMed

    Kishii, Ryuta; Falzon, Liliana; Yoshida, Takeshi; Kobayashi, Hiroshi; Inouye, Masayori

    2007-09-07

    The HAMP domain plays an essential role in signal transduction not only in histidine kinase but also in a number of other signal-transducing receptor proteins. Here we expressed the EnvZ HAMP domain (Arg(180)-Thr(235)) with the R218K mutation (termed L(RK)) or with L(RK) connected with domain A (Arg(180)-Arg(289)) (termed LA(RK)) of EnvZ, an osmosensing transmembrane histidine kinase in Escherichia coli, by fusing it with protein S. The L(RK) and LA(RK) proteins were purified after removing protein S. The CD analysis of the isolated L protein revealed that it consists of a random structure or is unstructured. This suggests that the EnvZ HAMP domain by itself is unable to form a stable structure and that this structural fragility may be important for its role in signal transduction. Interestingly the substitution of Ala(193) in the EnvZ HAMP domain with valine or leucine in Tez1A1, a chimeric protein of Tar and EnvZ, caused a constitutive OmpC phenotype. The CD analysis of LA(RK)(A193L) revealed that this mutated HAMP domain possesses considerable secondary structures and that the thermostability of this entire LA(RK)(A193L) became substantially lower than that of LA(RK) or just domain A, indicating that the structure of the HAMP domain with the A193L mutation affects the stability of downstream domain A. This results in cooperative thermodenaturation of domain A with the mutated HAMP domain. These results are discussed in light of the recently solved NMR structure of the HAMP domain from a thermophilic bacterium (Hulko, M., Berndt, F., Gruber, M., Linder, J. U., Truffault, V., Schultz, A., Martin, J., Schultz, J. E., Lupas, A. N., and Coles, M. (2006) Cell 126, 929-940).

  16. Genetic attributes of blood-derived subtype-C HIV-1 tat and env in India and neurocognitive function.

    PubMed

    Tilghman, Myres W; Bhattacharya, Jayanta; Deshpande, Suprit; Ghate, Manisha; Espitia, Stephen; Grant, Igor; Marcotte, Thomas D; Smith, Davey; Mehendale, Sanjay

    2014-01-01

    Genetic elements in HIV-1 subtype B tat and env are associated with neurotoxicity yet less is known about other subtypes. HIV-1 subtype C tat and env sequences were analyzed to determine viral genetic elements associated with neurocognitive impairment in a large Indian cohort. Population-based sequences of HIV-1 tat (exon 1) and env (C2-V3 coding region) were generated from blood plasma of HIV-infected patients in Pune, India. Participants were classified as cognitively normal or impaired based on neuropsychological assessment. Tests for signature residues, positive and negative selection, entropy, and ambiguous bases were performed using tools available through Los Alamos National Laboratory (http://www.hiv.lanl.gov) and Datamonkey (http://www.datamonkey.org). HIV-1 subtype C tat and env sequences were analyzed for 155 and 160 participants, of which 34-36% were impaired. Two signature residues were unique to impaired participants in exon 1 of tat at codons 29 (arginine) and 68 (proline). Positive selection was noted at codon 29 among normal participants and at codon 68 in both groups. The signature at codon 29 was also a signature for low CD4+ (<200 cells/mm(3)) counts but remained associated with impairment after exclusion of those with low CD4+ counts. No unique genetic signatures were noted in env. In conclusion, two signature residues were identified in exon 1 of HIV-1 subtype C tat that were associated with neurocognitive impairment in India and not completely accounted for by HIV disease progression. These signatures support a linkage between diversifying selection in HIV-1 subtype C tat and neurocognitive impairment. © 2013 Wiley Periodicals, Inc.

  17. A biosensor assay for studying ligand-membrane receptor interactions: Binding of antibodies and HIV-1 Env to chemokine receptors

    PubMed Central

    Hoffman, Trevor L.; Canziani, Gabriela; Jia, Li; Rucker, Joseph; Doms, Robert W.

    2000-01-01

    The HIV envelope (Env) protein mediates entry into cells by binding CD4 and an appropriate coreceptor, which triggers structural changes in Env that lead to fusion between the viral and cellular membranes. The major HIV-1 coreceptors are the seven transmembrane domain chemokine receptors CCR5 and CXCR4. The type of coreceptor used by a virus strain is an important determinant of viral tropism and pathogenesis, and virus-receptor interactions can be therapeutic targets. However, Envs from many virus strains interact with CXCR4 and CCR5 with low affinity such that direct study of this important interaction is difficult if not impossible using standard cell-surface binding techniques. We have developed an approach that makes it possible to study ligand binding to membrane proteins, including Env–coreceptor interactions, using an optical biosensor. CCR5, CXCR4, and other membrane proteins were incorporated into retrovirus particles, which were purified and attached to the biosensor surface. Binding of conformationally sensitive antibodies as well as Env to these receptors was readily detected. The equilibrium dissociation constant for the interaction between an Env derived from the prototype HIV-1 strain IIIB for CXCR4 was approximately 500 nM, explaining the difficulty in measuring this interaction using standard equilibrium binding techniques. Retroviral pseudotypes represent easily produced, stable, homogenous structures that can be used to present a wide array of single and multiple membrane-spanning proteins in a native lipid environment for biosensor studies, thus avoiding the need for detergent solubilization, purification, and reconstitution. The approach should have general applicability and can be used to correlate Env–receptor binding constants to viral tropism and pathogenesis. PMID:11005830

  18. Stabilizing the Native Trimer of HIV-1 Env by Destabilizing the Heterodimeric Interface of the gp41 Postfusion Six-Helix Bundle

    PubMed Central

    Kesavardhana, Sannula

    2014-01-01

    ABSTRACT The HIV-1 envelope glycoprotein (Env) is a trimer of gp120-gp41 heterodimers and is essential for viral entry. The gp41 subunit in native, prefusion trimeric Env exists in a metastable conformation and attains a stable six-helix bundle (6-HB) conformation comprised of a trimer of N-heptad repeat (NHR) and C-heptad repeat (CHR) heterodimers, that drives the fusion of viral and cellular membranes. We attempted to stabilize native Env trimers by incorporation of mutations at the NHR-CHR interface that disrupt the postfusion 6-HB of gp41. The mutations V570D and I573D stabilize native Env of the HIV-1 JRFL strain and occlude nonneutralizing epitopes to a greater extent than the previously identified I559P mutation that is at the interface of the NHR trimers in the 6-HB. The mutations prevent soluble-CD4 (sCD4)-induced gp120 shedding and 6-HB formation. In the context of cell surface-expressed JRFL Env, introduction of a previously reported additional disulfide between residues A501 and T605 perturbs the native conformation, though this effect is partially alleviated by furin coexpression. The data suggest that positions 570 and 573 are surface proximal in native Env and that the NHR homotrimeric coiled coil in native Env terminates before or close to residue 573. Aspartic acid substitutions at these positions stabilize native trimers through destabilization of the postfusion 6-HB conformation. These mutations can be used to stabilize Env in a DNA vaccine format. IMPORTANCE The major protein on the surface of HIV-1 is the envelope (Env) glycoprotein. Env is a trimer of gp120-gp41 heterodimers. gp120 is involved in receptor/coreceptor binding and gp41 in the fusion of viral and cellular membranes. Like many other viral fusion proteins, the gp41 subunit in native trimeric Env exists in a metastable conformation. gp41 readily forms a stable six-helix bundle (6-HB) conformation comprised of a trimer of N-heptad repeat (NHR) and C-heptad repeat (CHR) heterodimers

  19. Gen-X : Science Objectives

    NASA Astrophysics Data System (ADS)

    Heyl, J. S.; Generation-X Team

    2004-08-01

    Charting the development of the first objects from very early times into the structure we see in the Universe today is a sweeping goal of cosmology. X-ray observations are a key factor in achieving this goal: these first objects are expected to be powerful sources of X-rays and X-rays penetrate both the haze of the high z intergalactic medium, and the dust and gas expected around high z objects. These scientific goals drive the mission parameters of Generation-X, giving it an unrivaled combination of spatial, spectral resolution and sensitivity. Not only will Gen-X observe the birth of the first black holes, stars and galaxies, trace their evolution and the elements they produce. It will also probe more nearby phenomena with unparalleled precision, exploring both the most extreme objects in the universe (such as supermassive black holes, magnetars and microquasars) and the more mundane but equally important (such as protoplanetary disks and sun-like stars). Gen-X will be an extraordinarily powerful X-ray observatory for all of astrophysics.

  20. Macrophage entry mediated by HIV Envs from brain and lymphoid tissues is determined by the capacity to use low CD4 levels and overall efficiency of fusion

    SciTech Connect

    Thomas, Elaine R.; Dunfee, Rebecca L.; Stanton, Jennifer; Bogdan, Derek; Taylor, Joann; Kunstman, Kevin; Bell, Jeanne E.; Wolinsky, Steven M.; Gabuzda, Dana . E-mail: dana_gabuzda@dfci.harvard.edu

    2007-03-30

    HIV infects macrophages and microglia in the central nervous system (CNS), which express lower levels of CD4 than CD4+ T cells in peripheral blood. To investigate mechanisms of HIV neurotropism, full-length env genes were cloned from autopsy brain and lymphoid tissues from 4 AIDS patients with HIV-associated dementia (HAD). Characterization of 55 functional Env clones demonstrated that Envs with reduced dependence on CD4 for fusion and viral entry are more frequent in brain compared to lymphoid tissue. Envs that mediated efficient entry into macrophages were frequent in brain but were also present in lymphoid tissue. For most Envs, entry into macrophages correlated with overall fusion activity at all levels of CD4 and CCR5. gp160 nucleotide sequences were compartmentalized in brain versus lymphoid tissue within each patient. Proline at position 308 in the V3 loop of gp120 was associated with brain compartmentalization in 3 patients, but mutagenesis studies suggested that P308 alone does not contribute to reduced CD4 dependence or macrophage-tropism. These results suggest that HIV adaptation to replicate in the CNS selects for Envs with reduced CD4 dependence and increased fusion activity. Macrophage-tropic Envs are frequent in brain but are also present in lymphoid tissues of AIDS patients with HAD, and entry into macrophages in the CNS and other tissues is dependent on the ability to use low receptor levels and overall efficiency of fusion.

  1. Identification and characterization of the Escherichia coli envC gene encoding a periplasmic coiled-coil protein with putative peptidase activity.

    PubMed

    Hara, Hiroshi; Narita, Setsuko; Karibian, Doris; Park, James T; Yamamoto, Yoshihiro; Nishimura, Yukinobu

    2002-07-02

    PM61 is a chain-forming envC strain of Escherichia coli with a leaky outer membrane. It was found to have an oversized penicillin-binding protein 3, which was the result of an IS4 insertion in the prc gene. The other properties of PM61 were caused by the envC mutation. We cloned the envC (yibP) gene and identified the mutation site, causing a single residue substitution, H366Y, in the PM61 envC allele. The gene product was predicted to be a periplasmic protein having coiled-coil structure in the N-terminal region and homology to lysostaphin in the C-terminal region. Overexpression of envC inhibited cell growth, and overexpression of the PM61 mutant allele caused cell lysis. Disruption of the chromosomal envC caused the same defects as the envC point mutation, indicating the gene is dispensable for growth but important for normal septation/separation and cell envelope integrity.

  2. Nature of the Penetration Barrier in Escherichia coli K-12: Effect of Macromolecular Inhibition on Penetrability in Strains Containing the envA Gene

    PubMed Central

    Normark, Staffan; Westling, Britta

    1971-01-01

    The envA mutation in Escherichia coli K-12, which maps at 1.5 min, was previously shown to mediate sensitivity to gentian violet as well as to several antibiotics. Moreover, strains containing the envA gene were recently found to be lysed by lysozyme in the absence of ethylenediaminetetraacetate. It is here reported that the envA mutation mediates an increased uptake of gentian violet. The uptake of the dye was markedly affected by growth with different antibiotics interfering with macromolecular synthesis. Amino acid starvation of a strain containing envA with a stringent control of ribonucleic acid (RNA) synthesis resulted in a decreased uptake of gentian violet. However, no decrease in dye uptake was found during starvation in an envA transductant with a relaxed control of RNA synthesis. Inhibition of deoxyribonucleic acid (DNA) synthesis by nalidixic acid decreased the uptake of gentian violet of envA cells and, in addition, rendered the cells insensitive to the lytic action of lysozyme. Chloramphenicol treatment increased penetrability in wild-type and starved envA cells. In most instances, this effect of chloramphenicol was prevented by selectively interfering with DNA or RNA synthesis. A coordinate regulation of nucleic acid synthesis and penetrability is suggested. PMID:4941566

  3. Relationship of the env genes and the endonuclease domain of the pol genes of simian foamy virus type 1 and human foamy virus.

    PubMed Central

    Mergia, A; Shaw, K E; Lackner, J E; Luciw, P A

    1990-01-01

    We have molecularly cloned and sequenced a portion of the simian foamy virus type 1 (SFV-1); open reading frames representing the endonuclease domain of the polymerase (pol) and the envelope (env) genes were identified by comparison with the human foamy virus (HFV). Unlike the HFV genomic organization, the SFV-1 pol gene overlaps the env gene; thus, the open reading frames reported for HFV between pol and env is not present in SFV-1. Comparisons of predicted amino acid sequences of HFV and SFV-1 reveal that the endonuclease domains of the pol genes are about 84% related. The region predicted to encode the SFV-1 extracellular env domain is 569 codons; SFV-1 and HFV have 64% amino acid similarity in this env domain. The predicted hydrophobic transmembrane env proteins of both HFV and SFV-1 show about 73% similarity. A total of 16 potential glycosylation sites are found in SFV-1 env, and 15 are found in HFV; 11 are shared. SFV-1 has 25 cysteine residues, and HFV has 23 residues; all 23 cysteine residues of HFV are conserved in SFV-1. This sequence analysis reveals that the human and simian foamy viruses are highly related. PMID:2152825

  4. Unique N-linked glycosylation of CasBrE Env influences its stability, processing, and viral infectivity but not its neurotoxicity.

    PubMed

    Renszel, Krystal M; Traister, Russell S; Lynch, William P

    2013-08-01

    The envelope protein (Env) from the CasBrE murine leukemia virus (MLV) can cause acute spongiform neurodegeneration analogous to that induced by prions. Upon central nervous system (CNS) infection, Env is expressed as multiple isoforms owing to differential asparagine (N)-linked glycosylation. Because N-glycosylation can affect protein folding, stability, and quality control, we explored whether unique CasBrE Env glycosylation features could influence neurovirulence. CasBrE Env possesses 6/8 consensus MLV glycosylation sites (gs) but is missing gs3 and gs5 and contains a putative site (gs*). Twenty-nine mutants were generated by modifying these three sites, individually or in combination, to mimic the amino acid sequence in the nonneurovirulent Friend 57 MLV. Three basic viral phenotypes were observed: replication defective (dead; titer < 1 focus-forming unit [FFU]/ml), replication compromised (RC) (titer = 10(2) to 10(5) FFU/ml); and wild-type-like (WTL) (titer > 10(5) FFU/ml). Env protein was undetectable in dead mutants, while RC and WTL mutants showed variations in Env expression, processing, virus incorporation, virus entry, and virus spread. The newly introduced gs3 and gs5 sites were glycosylated, whereas gs* was not. Six WTL mutants tested in mice showed no clear attenuation in disease onset or severity versus controls. Furthermore, three RC viruses tested by neural stem cell (NSC)-mediated brainstem dissemination also induced acute spongiosis. Thus, while unique N-glycosylation affected structural features of Env involved in protein stability, proteolytic processing, and virus assembly and entry, these changes had minimal impact on CasBrE Env neurotoxicity. These findings suggest that the Env protein domains responsible for spongiogenesis represent highly stable elements upon which the more variable viral functional domains have evolved.

  5. Life+ EnvEurope DEIMS - improving access to long-term ecosystem monitoring data in Europe

    NASA Astrophysics Data System (ADS)

    Kliment, Tomas; Peterseil, Johannes; Oggioni, Alessandro; Pugnetti, Alessandra; Blankman, David

    2013-04-01

    Long-term ecological (LTER) studies aim at detecting environmental changes and analysing its related drivers. In this respect LTER Europe provides a network of about 450 sites and platforms. However, data on various types of ecosystems and at a broad geographical scale is still not easily available. Managing data resulting from long-term observations is therefore one of the important tasks not only for an LTER site itself but also on the network level. Exchanging and sharing the information within a wider community is a crucial objective in the upcoming years. Due to the fragmented nature of long-term ecological research and monitoring (LTER) in Europe - and also on the global scale - information management has to face several challenges: distributed data sources, heterogeneous data models, heterogeneous data management solutions and the complex domain of ecosystem monitoring with regard to the resulting data. The Life+ EnvEurope project (2010-2013) provides a case study for a workflow using data from the distributed network of LTER-Europe sites. In order to enhance discovery, evaluation and access to data, the EnvEurope Drupal Ecological Information Management System (DEIMS) has been developed. This is based on the first official release of the Drupal metadata editor developed by US LTER. EnvEurope DEIMS consists of three main components: 1) Metadata editor: a web-based client interface to manage metadata of three information resource types - datasets, persons and research sites. A metadata model describing datasets based on Ecological Metadata Language (EML) was developed within the initial phase of the project. A crosswalk to the INSPIRE metadata model was implemented to convey to the currently on-going European activities. Person and research site metadata models defined within the LTER Europe were adapted for the project needs. The three metadata models are interconnected within the system in order to provide easy way to navigate the user among the related

  6. Global Panel of HIV-1 Env Reference Strains for Standardized Assessments of Vaccine-Elicited Neutralizing Antibodies

    PubMed Central

    deCamp, Allan; Hraber, Peter; Bailer, Robert T.; Seaman, Michael S.; Ochsenbauer, Christina; Kappes, John; Gottardo, Raphael; Edlefsen, Paul; Self, Steve; Tang, Haili; Greene, Kelli; Gao, Hongmei; Daniell, Xiaoju; Sarzotti-Kelsoe, Marcella; Gorny, Miroslaw K.; Zolla-Pazner, Susan; LaBranche, Celia C.; Mascola, John R.; Korber, Bette T.

    2014-01-01

    ABSTRACT Standardized assessments of HIV-1 vaccine-elicited neutralizing antibody responses are complicated by the genetic and antigenic variability of the viral envelope glycoproteins (Envs). To address these issues, suitable reference strains are needed that are representative of the global epidemic. Several panels have been recommended previously, but no clear answers have been available on how many and which strains are best suited for this purpose. We used a statistical model selection method to identify a global panel of reference Env clones from among 219 Env-pseudotyped viruses assayed in TZM-bl cells with sera from 205 HIV-1-infected individuals. The Envs and sera were sampled globally from diverse geographic locations and represented all major genetic subtypes and circulating recombinant forms of the virus. Assays with a panel size of only nine viruses adequately represented the spectrum of HIV-1 serum neutralizing activity seen with the larger panel of 219 viruses. An optimal panel of nine viruses was selected and augmented with three additional viruses for greater genetic and antigenic coverage. The spectrum of HIV-1 serum neutralizing activity seen with the final 12-virus panel closely approximated the activity seen with subtype-matched viruses. Moreover, the final panel was highly sensitive for detection of many of the known broadly neutralizing antibodies. For broader assay applications, all 12 Env clones were converted to infectious molecular clones using a proviral backbone carrying a Renilla luciferase reporter gene (Env.IMC.LucR viruses). This global panel should facilitate highly standardized assessments of vaccine-elicited neutralizing antibodies across multiple HIV-1 vaccine platforms in different parts of the world. IMPORTANCE An effective HIV-1 vaccine will need to overcome the extraordinary genetic variability of the virus, where most variation occurs in the viral envelope glycoproteins that are the sole targets for neutralizing antibodies

  7. A rev1-vpu polymorphism unique to HIV-1 subtype A and C strains impairs envelope glycoprotein expression from rev-vpu-env cassettes and reduces virion infectivity in pseudotyping assays

    SciTech Connect

    Kraus, Matthias H.; Parrish, Nicholas F.; Shaw, Katharina S.; Decker, Julie M.; Keele, Brandon F.; Salazar-Gonzalez, Jesus F.; Grayson, Truman; McPherson, David T.; Ping, Li-Hua; Anderson, Jeffrey A.; Swanstrom, Ronald; Williamson, Carolyn; Shaw, George M.; Hahn, Beatrice H.

    2010-02-20

    Functional studies of HIV-1 envelope glycoproteins (Envs) commonly include the generation of pseudoviruses, which are produced by co-transfection of rev-vpu-env cassettes with an env-deficient provirus. Here, we describe six Env constructs from transmitted/founder HIV-1 that were defective in the pseudotyping assay, although two produced infectious virions when expressed from their cognate proviruses. All of these constructs exhibited an unusual gene arrangement in which the first exon of rev (rev1) and vpu were in the same reading frame without an intervening stop codon. Disruption of the rev1-vpu fusion gene by frameshift mutation, stop codon, or abrogation of the rev initiation codon restored pseudovirion infectivity. Introduction of the fusion gene into wildtype Env cassettes severely compromised their function. The defect was not due to altered env and rev transcription or a dominant negative effect of the expressed fusion protein, but seemed to be caused by inefficient translation at the env initiation codon. Although the rev1-vpu polymorphism affects Env expression only in vitro, it can cause problems in studies requiring Env complementation, such as analyses of co-receptor usage and neutralization properties, since 3% of subtype A, 20% of subtype C and 5% of CRF01{sub A}/E viruses encode the fusion gene. A solution is to eliminate the rev initiation codon when amplifying rev-vpu-env cassettes since this increases Env expression irrespective of the presence of the polymorphism.

  8. Isolation and sequencing of infectious clones of feline foamy virus and a human/feline foamy virus Env chimera.

    PubMed

    Hatama, S; Otake, K; Omoto, S; Murase, Y; Ikemoto, A; Mochizuki, M; Takahashi, E; Okuyama, H; Fujii, Y

    2001-12-01

    Full-length DNAs of the Coleman and S7801 strains (pSKY3.0, pSKY5.0) of infectious feline foamy viruses (FFVs) were cloned and sequenced. Parental viruses, designated SKY3.0 and SKY5.0, were secreted following transfection of Crandell feline kidney (CRFK) cells. Production of the rescued parental viruses was enhanced in the presence of trichostatin A. Amino acid sequence similarities between FFV and human foamy virus (HFV) are extremely low for the envelope protein and capsid antigen, as predicted from the two clones. However, a chimeric FFV clone was constructed with the HFV Env substituted for the FFV Env. The chimeric virus (HFFV, SKY4.0) was able to infect and replicate in CRFK cells as well as in peripheral blood mononuclear cells of cats in vivo. Consequently, the chimeric HFFV may be useful for the creation of FV vectors for gene transfer strategies.

  9. Variation in simian immunodeficiency virus env is confined to V1 and V4 during progression to simian AIDS.

    PubMed Central

    Overbaugh, J; Rudensey, L M; Papenhausen, M D; Benveniste, R E; Morton, W R

    1991-01-01

    We have monitored changes in the simian immunodeficiency virus (SIV) envelope (env) gene in two macaques which developed AIDS after inoculation with a molecular clone of SIV. As the animals progressed to AIDS, selection occurred for viruses with variation in two discrete regions (V1 and V4) but not for viruses with changes in the region of SIV env that corresponds to the immunodominant, V3 loop of human immunodeficiency virus. Within the highly variable domains, the vast majority of nucleotide changes encoded an amino acid change (98%), suggesting that these envelope variants had evolved as a result of phenotypic selection. Analysis of the biological properties of these variants, which have been selected for in the host, may be useful in defining the mechanisms underlying viral persistence and progression to simian AIDS. PMID:1942255

  10. Effects of murine leukemia virus env gene proteins on macrophage-mediated cytotoxicity in vitro

    NASA Technical Reports Server (NTRS)

    Chapes, S. K.; Takemoto, L. J.; Spooner, B. S. (Principal Investigator)

    1991-01-01

    F5b Tumor cells were incubated with concentrated culture supernatants taken from cells resistant (F5m) or sensitive (F5b) to contact-dependent macrophage cytotoxicity. Macrophage cell line B6MP102 and murine peritoneal macrophages killed targets incubated with supernatants taken from sensitive cells but poorly killed cells incubated in supernatants isolated from resistant cells. Membranes from cells resistant to macrophage killing, F5m, were fused into F5b cells. The fused F5b cells were killed significantly less than F5b cells fused with F5b cell membranes or untreated F5b cells. The decreased killing of F5b cells corresponded to increased concentrations of gp70(a) molecules on F5b cells. Affinity purified gp70(a) was added to cytotoxicity assays but failed to inhibit macrophage cytotoxicity. P15E molecules were detectable on both F5b and F5m cells. In addition, a synthetic peptide found to exhibit the inhibitory properties of p15E was added to cytotoxicity assays. P15E synthetic peptide also did not inhibit macrophage cytotoxicity. Therefore, env gene proteins of murine leukemia virus do not appear responsible for inducing tumor cell resistance to activated macrophage contact-dependent cytotoxicity.

  11. Obacunone Represses Salmonella Pathogenicity Islands 1 and 2 in an envZ-Dependent Fashion

    PubMed Central

    Vikram, Amit; Jayaprakasha, Guddadarangavvanahally K.; Jesudhasan, Palmy R.

    2012-01-01

    Obacunone belongs to a class of unique triterpenoids called limonoids, present in Citrus species. Previous studies from our laboratory suggested that obacunone possesses antivirulence activity and demonstrates inhibition of cell-cell signaling in Vibrio harveyi and Escherichia coli O157:H7. The present work sought to determine the effect of obacunone on the food-borne pathogen Salmonella enterica serovar Typhimurium LT2 by using a cDNA microarray. Transcriptomic studies indicated that obacunone represses Salmonella pathogenicity island 1 (SPI1), the maltose transporter, and the hydrogenase operon. Furthermore, phenotypic data for the Caco-2 infection assay and maltose utilization were in agreement with microarray data suggesting repression of SPI1 and maltose transport. Further studies demonstrated that repression of SPI1 was plausibly mediated through hilA. Additionally, obacunone seems to repress SPI2 under SPI2-inducing conditions as well as in Caco-2 infection models. Furthermore, obacunone seems to repress hilA in an EnvZ-dependent fashion. Altogether, the results of the study seems to suggest that obacunone exerts an antivirulence effect on S. Typhimurium and may serve as a lead compound for development of antivirulence strategies for S. Typhimurium. PMID:22843534

  12. αEnv-decorated phosphatidylserine liposomes trigger phagocytosis of HIV-virus-like particles in macrophages.

    PubMed

    Gramatica, Andrea; Petazzi, Roberto A; Lehmann, Maik J; Ziomkowska, Joanna; Herrmann, Andreas; Chiantia, Salvatore

    2014-07-01

    Macrophages represent an important cellular target of HIV-1. Interestingly, they are also believed to play a potential role counteracting its infection. However, HIV-1 is known to impair macrophage immune functions such as antibody-mediated phagocytosis. Here, we present immunoliposomes that can bind HIV-1 virus-like particles (HIV-VLPs) while being specifically phagocytosed by macrophages, thus allowing the co-internalization of HIV-VLPs. These liposomes are decorated with anti-Env antibodies and contain phosphatidylserine (PS). PS mediates liposome internalization by macrophages via a mechanism not affected by HIV-1. Hence, PS-liposomes mimic apoptotic cells and are internalized into the macrophages due to specific recognition, carrying the previously bound HIV-VLPs. With a combination of flow cytometry, confocal live-cell imaging and electron microscopy we demonstrate that the PS-immunoliposomes presented here are able to elicit efficient HIV-VLPs phagocytosis by macrophages and might represent a new nanotechnological approach to enhance HIV-1 antigen presentation and reduce the ongoing inflammation processes. This team of authors demonstrate that specific phosphatidylserin immunoliposomes are able to elicit efficient phagocytosis of HIV-virus-like particle by macrophages and might represent a new nanomedicine approach to enhance HIV-1 antigen presentation and reduce ongoing inflammation processes. Copyright © 2014 Elsevier Inc. All rights reserved.

  13. Increased HIV-1 vaccine efficacy against viruses with genetic signatures in Env V2.

    PubMed

    Rolland, Morgane; Edlefsen, Paul T; Larsen, Brendan B; Tovanabutra, Sodsai; Sanders-Buell, Eric; Hertz, Tomer; deCamp, Allan C; Carrico, Chris; Menis, Sergey; Magaret, Craig A; Ahmed, Hasan; Juraska, Michal; Chen, Lennie; Konopa, Philip; Nariya, Snehal; Stoddard, Julia N; Wong, Kim; Zhao, Hong; Deng, Wenjie; Maust, Brandon S; Bose, Meera; Howell, Shana; Bates, Adam; Lazzaro, Michelle; O'Sullivan, Annemarie; Lei, Esther; Bradfield, Andrea; Ibitamuno, Grace; Assawadarachai, Vatcharain; O'Connell, Robert J; deSouza, Mark S; Nitayaphan, Sorachai; Rerks-Ngarm, Supachai; Robb, Merlin L; McLellan, Jason S; Georgiev, Ivelin; Kwong, Peter D; Carlson, Jonathan M; Michael, Nelson L; Schief, William R; Gilbert, Peter B; Mullins, James I; Kim, Jerome H

    2012-10-18

    The RV144 trial demonstrated 31% vaccine efficacy at preventing human immunodeficiency virus (HIV)-1 infection. Antibodies against the HIV-1 envelope variable loops 1 and 2 (Env V1 and V2) correlated inversely with infection risk. We proposed that vaccine-induced immune responses against V1/V2 would have a selective effect against, or sieve, HIV-1 breakthrough viruses. A total of 936 HIV-1 genome sequences from 44 vaccine and 66 placebo recipients were examined. We show that vaccine-induced immune responses were associated with two signatures in V2 at amino acid positions 169 and 181. Vaccine efficacy against viruses matching the vaccine at position 169 was 48% (confidence interval 18% to 66%; P = 0.0036), whereas vaccine efficacy against viruses mismatching the vaccine at position 181 was 78% (confidence interval 35% to 93%; P = 0.0028). Residue 169 is in a cationic glycosylated region recognized by broadly neutralizing and RV144-derived antibodies. The predicted distance between the two signature sites (21 ± 7 Å) and their match/mismatch dichotomy indicate that multiple factors may be involved in the protection observed in RV144. Genetic signatures of RV144 vaccination in V2 complement the finding of an association between high V1/V2-binding antibodies and reduced risk of HIV-1 acquisition, and provide evidence that vaccine-induced V2 responses plausibly had a role in the partial protection conferred by the RV144 regimen.

  14. Comparing the flammability of fabrics in accordance with EN 531 and ENV 50354.

    PubMed

    Mäkinen, Helena; Mustonen, Suvi Sanna

    2004-01-01

    The purpose of protective clothing and other personal protective equipment (PPE) is to provide escape time, to reduce the burn injury level, and to prevent aggravation of the consequences to workers during exposure to an electric arc. In this study the flammability properties of 12 different types of flame-retardant fabrics were compared with the normally used flame spread test method (EN 532:1994) and electric arc test method (ENV 50354:2001). In the arc test at the lower testing current level of 4 kA, the requirement was passed by materials which did not pass the flame spread test. These materials contained a large amount of melting fibres, and therefore tended to shrink or melt. In order to meet the current level of 7 kA, a rather thick and heavy flame-retardant fabric is needed to pass the requirement. Lighter fabrics tended to break open in the tests. The flame retardancy of the under layer fabric is therefore important to ensure the needed protection.

  15. A machine learning approach for identifying amino acid signatures in the HIV env gene predictive of dementia.

    PubMed

    Holman, Alexander G; Gabuzda, Dana

    2012-01-01

    The identification of nucleotide sequence variations in viral pathogens linked to disease and clinical outcomes is important for developing vaccines and therapies. However, identifying these genetic variations in rapidly evolving pathogens adapting to selection pressures unique to each host presents several challenges. Machine learning tools provide new opportunities to address these challenges. In HIV infection, virus replicating within the brain causes HIV-associated dementia (HAD) and milder forms of neurocognitive impairment in 20-30% of patients with unsuppressed viremia. HIV neurotropism is primarily determined by the viral envelope (env) gene. To identify amino acid signatures in the HIV env gene predictive of HAD, we developed a machine learning pipeline using the PART rule-learning algorithm and C4.5 decision tree inducer to train a classifier on a meta-dataset (n = 860 env sequences from 78 patients: 40 HAD, 38 non-HAD). To increase the flexibility and biological relevance of our analysis, we included 4 numeric factors describing amino acid hydrophobicity, polarity, bulkiness, and charge, in addition to amino acid identities. The classifier had 75% predictive accuracy in leave-one-out cross-validation, and identified 5 signatures associated with HAD diagnosis (p<0.05, Fisher's exact test). These HAD signatures were found in the majority of brain sequences from 8 of 10 HAD patients from an independent cohort. Additionally, 2 HAD signatures were validated against env sequences from CSF of a second independent cohort. This analysis provides insight into viral genetic determinants associated with HAD, and develops novel methods for applying machine learning tools to analyze the genetics of rapidly evolving pathogens.

  16. A Machine Learning Approach for Identifying Amino Acid Signatures in the HIV Env Gene Predictive of Dementia

    PubMed Central

    Holman, Alexander G.; Gabuzda, Dana

    2012-01-01

    The identification of nucleotide sequence variations in viral pathogens linked to disease and clinical outcomes is important for developing vaccines and therapies. However, identifying these genetic variations in rapidly evolving pathogens adapting to selection pressures unique to each host presents several challenges. Machine learning tools provide new opportunities to address these challenges. In HIV infection, virus replicating within the brain causes HIV-associated dementia (HAD) and milder forms of neurocognitive impairment in 20–30% of patients with unsuppressed viremia. HIV neurotropism is primarily determined by the viral envelope (env) gene. To identify amino acid signatures in the HIV env gene predictive of HAD, we developed a machine learning pipeline using the PART rule-learning algorithm and C4.5 decision tree inducer to train a classifier on a meta-dataset (n = 860 env sequences from 78 patients: 40 HAD, 38 non-HAD). To increase the flexibility and biological relevance of our analysis, we included 4 numeric factors describing amino acid hydrophobicity, polarity, bulkiness, and charge, in addition to amino acid identities. The classifier had 75% predictive accuracy in leave-one-out cross-validation, and identified 5 signatures associated with HAD diagnosis (p<0.05, Fisher’s exact test). These HAD signatures were found in the majority of brain sequences from 8 of 10 HAD patients from an independent cohort. Additionally, 2 HAD signatures were validated against env sequences from CSF of a second independent cohort. This analysis provides insight into viral genetic determinants associated with HAD, and develops novel methods for applying machine learning tools to analyze the genetics of rapidly evolving pathogens. PMID:23166702

  17. Comparison of intradermal and intramuscular delivery followed by in vivo electroporation of SIV Env DNA in macaques

    PubMed Central

    Kulkarni, Viraj; Rosati, Margherita; Bear, Jenifer; Pilkington, Guy R; Jalah, Rashmi; Bergamaschi, Cristina; Singh, Ashish K; Alicea, Candido; Chowdhury, Bhabadeb; Zhang, Gen-Mu; Kim, Eun-Young; Wolinsky, Steven M; Huang, Wensheng; Guan, Yongjun; LaBranche, Celia; Montefiori, David C; Broderick, Kate E; Sardesai, Niranjan Y; Valentin, Antonio; Felber, Barbara K; Pavlakis, George N

    2013-01-01

    A panel of SIVmac251 transmitted Env sequences were tested for expression, function and immunogenicity in mice and macaques. The immunogenicity of a DNA vaccine cocktail expressing SIVmac239 and three transmitted SIVmac251 Env sequences was evaluated upon intradermal or intramuscular injection followed by in vivo electroporation in macaques using sequential vaccination of gp160, gp120 and gp140 expressing DNAs. Both intradermal and intramuscular vaccination regimens using the gp160 expression plasmids induced robust humoral immune responses, which further improved using the gp120 expressing DNAs. The responses showed durability of binding and neutralizing antibody titers and high avidity for > 1 y. The intradermal DNA delivery regimen induced higher cross-reactive responses able to neutralize the heterologous tier 1B-like SIVsmE660_CG7V. Analysis of cellular immune responses showed induction of Env-specific memory responses and cytotoxic granzyme B+ T cells in both vaccine groups, although the magnitude of the responses were ~10x higher in the intramuscular/electroporation group. The cellular responses induced by both regimens were long lasting and could be detected ~1 y after the last vaccination. These data show that both DNA delivery methods are able to induce robust and durable immune responses in macaques. PMID:23811579

  18. Production of Mucosally Transmissible SHIV Challenge Stocks from HIV-1 Circulating Recombinant Form 01_AE env Sequences.

    PubMed

    Tartaglia, Lawrence J; Chang, Hui-Wen; Lee, Benjamin C; Abbink, Peter; Ng'ang'a, David; Boyd, Michael; Lavine, Christy L; Lim, So-Yon; Sanisetty, Srisowmya; Whitney, James B; Seaman, Michael S; Rolland, Morgane; Tovanabutra, Sodsai; Ananworanich, Jintanat; Robb, Merlin L; Kim, Jerome H; Michael, Nelson L; Barouch, Dan H

    2016-02-01

    Simian-human immunodeficiency virus (SHIV) challenge stocks are critical for preclinical testing of vaccines, antibodies, and other interventions aimed to prevent HIV-1. A major unmet need for the field has been the lack of a SHIV challenge stock expressing circulating recombinant form 01_AE (CRF01_AE) env sequences. We therefore sought to develop mucosally transmissible SHIV challenge stocks containing HIV-1 CRF01_AE env derived from acutely HIV-1 infected individuals from Thailand. SHIV-AE6, SHIV-AE6RM, and SHIV-AE16 contained env sequences that were >99% identical to the original HIV-1 isolate and did not require in vivo passaging. These viruses exhibited CCR5 tropism and displayed a tier 2 neutralization phenotype. These challenge stocks efficiently infected rhesus monkeys by the intrarectal route, replicated to high levels during acute infection, and established chronic viremia in a subset of animals. SHIV-AE16 was titrated for use in single, high dose as well as repetitive, low dose intrarectal challenge studies. These SHIV challenge stocks should facilitate the preclinical evaluation of vaccines, monoclonal antibodies, and other interventions targeted at preventing HIV-1 CRF01_AE infection.

  19. Structures of HIV-1-Env V1V2 with broadly neutralizing antibodies reveal commonalities that enable vaccine design

    PubMed Central

    Gorman, Jason; Soto, Cinque; Yang, Max M.; Davenport, Thaddeus M.; Guttman, Miklos; Bailer, Robert T.; Chambers, Michael; Chuang, Gwo-Yu; DeKosky, Brandon J.; Doria-Rose, Nicole A.; Druz, Aliaksandr; Ernandes, Michael J.; Georgiev, Ivelin S.; Jarosinski, Marissa C.; Joyce, M. Gordon; Lemmin, Thomas M.; Leung, Sherman; Louder, Mark K.; McDaniel, Jonathan R.; Narpala, Sandeep; Pancera, Marie; Stuckey, Jonathan; Wu, Xueling; Yang, Yongping; Zhang, Baoshan; Zhou, Tongqing; Mullikin, James C.; Baxa, Ulrich; Georgiou, George; McDermott, Adrian B.; Bonsignori, Mattia; Haynes, Barton F.; Moore, Penny L.; Morris, Lynn; Lee, Kelly K.; Shapiro, Lawrence; Mascola, John R.; Kwong, Peter D.

    2016-01-01

    Broadly neutralizing antibodies (bNAbs) against HIV-1-Env V1V2 arise in multiple donors. However, atomic-level interactions had only been determined with antibodies from a single donor, making commonalities in recognition uncertain. Here we report the co-crystal structure of V1V2 with antibody CH03 from a second donor and model Env interactions of antibody CAP256-VRC26 from a third. These V1V2-directed bNAbs utilized strand-strand interactions between a protruding antibody loop and a V1V2 strand, but differed in their N-glycan recognition. Ontogeny analysis indicated protruding loops to develop early, with glycan interactions maturing over time. Altogether, the multidonor information suggested V1V2-directed bNAbs to form an ‘extended class’, for which we engineered ontogeny-specific antigens: Env trimers with chimeric V1V2s that interacted with inferred ancestor and intermediate antibodies. The ontogeny-based design of vaccine antigens described here may provide a general means for eliciting antibodies of a desired class. PMID:26689967

  20. Effects of different promoters on the virulence and immunogenicity of a HIV-1 Env-expressing recombinant vaccinia vaccine.

    PubMed

    Isshiki, Mao; Zhang, Xianfeng; Sato, Hirotaka; Ohashi, Takashi; Inoue, Makoto; Shida, Hisatoshi

    2014-02-07

    Previously, we developed a vaccination regimen that involves priming with recombinant vaccinia virus LC16m8Δ (rm8Δ) strain followed by boosting with a Sendai virus-containing vector. This protocol induced both humoral and cellular immune responses against the HIV-1 envelope protein. The current study aims to optimize this regimen by comparing the immunogenicity and safety of two rm8Δ strains that express HIV-1 Env under the control of a moderate promoter, p7.5, or a strong promoter, pSFJ1-10. m8Δ-p7.5-JRCSFenv synthesized less gp160 but showed significantly higher growth potential than m8Δ-pSFJ-JRCSFenv. The two different rm8Δ strains induced antigen-specific immunity; however, m8Δ-pSFJ-JRCSFenv elicited a stronger anti-Env antibody response whereas m8Δ-p7.5-JRCSFenv induced a stronger Env-specific cytotoxic T lymphocyte response. Both strains were less virulent than the parental m8Δ strain, suggesting that they would be safe for use in humans. These findings indicate the vaccine can be optimized to induce favorable immune responses (either cellular or humoral), and forms the basis for the rational design of an AIDS vaccine using recombinant vaccinia as the delivery vector. Copyright © 2013 Elsevier Ltd. All rights reserved.

  1. A novel human immunodeficiency virus type 1 protein, tev, shares sequences with tat, env, and rev proteins.

    PubMed Central

    Benko, D M; Schwartz, S; Pavlakis, G N; Felber, B K

    1990-01-01

    We have characterized a novel 28-kilodalton protein, p28tev, detected in human immunodeficiency virus type 1-infected cells. tev is recognized by both tat and rev monospecific antibodies. tev is initiated at the tat AUG and contains the first exon of tat at its amino terminus, a small portion of env in the middle, and the second exon of rev at its carboxy terminus. A cDNA clone producing tev was cloned and expressed in human cells. Sequence analysis revealed that the tev mRNA is generated by splicing to a novel exon located in the env region. This identifies a fourth class of multiply spliced human immunodeficiency virus mRNAs, produced in infected and transfected cells. tev is regulated during the virus life cycle similarly to the other regulatory proteins, tat, rev, and nef, and displays both tat and rev activities in functional assays. Since tev contains important functional domains of tat and rev and is produced very early after transfection, it may be an important regulator in the initial phase of virus expression. Another rev-related protein, p18(6)Drev, containing env and rev sequences, was characterized and was found not to have detectable rev activity. Images PMID:2186172

  2. Proviral amplification of the Gypsy endogenous retrovirus of Drosophila melanogaster involves env-independent invasion of the female germline.

    PubMed

    Chalvet, F; Teysset, L; Terzian, C; Prud'homme, N; Santamaria, P; Bucheton, A; Pélisson, A

    1999-05-04

    Gypsy is an infectious endogenous retrovirus of Drosophila melanogaster. The gypsy proviruses replicate very efficiently in the genome of the progeny of females homozygous for permissive alleles of the flamenco gene. This replicative transposition is correlated with derepression of gypsy expression, specifically in the somatic cells of the ovaries of the permissive mothers. The determinism of this amplification was studied further by making chimeric mothers containing different permissive/restrictive and somatic/germinal lineages. We show here that the derepression of active proviruses in the permissive soma is necessary and sufficient to induce proviral insertions in the progeny, even if the F1 flies derive from restrictive germ cells devoid of active proviruses. Therefore, gypsy endogenous multiplication results from the transfer of some gypsy-encoded genetic material from the soma towards the germen of the mother and its subsequent insertion into the chromosomes of the progeny. This transfer, however, is not likely to result from retroviral infection of the germline. Indeed, we also show here that the insertion of a tagged gypsy element, mutant for the env gene, occurs at high frequency, independently of the production of gypsy Env proteins by any transcomplementing helper. The possible role of the env gene for horizontal transfer to new hosts is discussed.

  3. Increased T cell breadth and antibody response elicited in prime-boost regimen by viral vector encoded homologous SIV Gag/Env in outbred CD1 mice.

    PubMed

    Andersson, Anne-Marie Carola; Holst, Peter Johannes

    2016-12-20

    A major obstacle for the development of HIV vaccines is the virus' worldwide sequence diversity. Nevertheless, the presence of T cell epitopes within conserved regions of the virus' structural Gag protein and conserved structures in the envelope (env) sequence raises the possibility that cross-reactive responses may be induced by vaccination. In this study, the aim was to investigate the importance of antigenic match on immunodominance and breadth of obtainable T cell responses. Outbred CD1 mice were immunized with either heterologous (SIVmac239 and HIV-1 clade B consensus) or homologous (SIVmac239) gag sequences using adenovirus (Ad5) and MVA vectors. Env (SIVmac239) was co-encoded in the vectors to study the induction of antibodies, which is a primary target of current HIV vaccine designs. All three vaccines were designed as virus-encoded virus-like particle vaccines. Antibody responses were analysed by ELISA, avidity ELISA, and neutralization assay. T cell responses were determined by intracellular cytokine staining of splenocytes. The homologous Env/Gag prime-boost regimen induced higher Env binding antibodies, and induced stronger and broader Gag specific CD8+ T cell responses than the homologous Env/heterologous Gag prime-boost regimen. Homologous Env/heterologous Gag immunization resulted in selective boosting of Env specific CD8+ T cell responses and consequently a paradoxical decreased recognition of variant sequences including conserved elements of p24 Gag. These results contrast with related studies using Env or Gag as the sole antigen and suggest that prime-boost immunizations based on homologous SIVmac239 Gag inserts is an efficient component of genetic VLP vaccines-both for induction of potent antibody responses and cross-reactive CD8+ T cell responses.

  4. HIV cell-to-cell transmission requires the production of infectious virus particles and does not proceed through env-mediated fusion pores.

    PubMed

    Monel, Blandine; Beaumont, Elodie; Vendrame, Daniela; Schwartz, Olivier; Brand, Denys; Mammano, Fabrizio

    2012-04-01

    Direct cell-to-cell transmission of human immunodeficiency virus (HIV) is a more potent and efficient means of virus propagation than infection by cell-free virus particles. The aim of this study was to determine whether cell-to-cell transmission requires the assembly of enveloped virus particles or whether nucleic acids with replication potential could translocate directly from donor to target cells through envelope glycoprotein (Env)-induced fusion pores. To this end, we characterized the transmission properties of viruses carrying mutations in the matrix protein (MA) that affect the incorporation of Env into virus particles but do not interfere with Env-mediated cell-cell fusion. By use of cell-free virus, the infectivity of MA mutant viruses was below the detection threshold both in single-cycle and in multiple-cycle assays. Truncation of the cytoplasmic tail (CT) of Env restored the incorporation of Env into MA mutant viruses and rescued their cell-free infectivity to different extents. In cell-to-cell transmission assays, MA mutations prevented HIV transmission from donor to target cells, despite efficient Env-dependent membrane fusion. HIV transmission was blocked at the level of virus core translocation into the cytosol of target cells. As in cell-free assays, rescue of Env incorporation by truncation of the Env CT restored the virus core translocation and cell-to-cell infectivity of MA mutant viruses. These data show that HIV cell-to-cell transmission requires the assembly of enveloped virus particles. The increased efficiency of this infection route may thus be attributed to the high local concentrations of virus particles at sites of cellular contacts rather than to a qualitatively different transmission process.

  5. Proposal of the genera Anaerococcus gen. nov., Peptoniphilus gen. nov. and Gallicola gen. nov. for members of the genus Peptostreptococcus.

    PubMed

    Ezaki, T; Kawamura, Y; Li, N; Li, Z Y; Zhao, L; Shu, S

    2001-07-01

    Members of genus Peptostreptococcus have previously been found to be distantly related to the type species, Peptostreptococcus anaerobius, on the basis of 16S rDNA sequence similarities. They were divided into three major phylogenetic groups, and their peptidoglycan structure and biochemical traits differed between groups. The reclassification of the species of these three groups into three new genera, Peptoniphilus gen. nov., Anaerococcus gen. nov. and Gallicola gen. nov., is proposed. The genus Peptoniphilus gen. nov. includes the following butyrate-producing, non-saccharolytic species that use peptone and amino acids as major energy sources: Peptoniphilus asaccharolyticus comb. nov. (type species), Peptoniphilus lacrimaris comb. nov., Peptoniphilus harei comb. nov., Peptoniphilus indolicus comb. nov. and Peptoniphilus ivorii comb. nov. The genus Anaerococcus gen. nov. contains the saccharolytic, butyrate-producing species Anaerococcus prevotii comb. nov. (type species), Anaerococcus tetradius comb. nov., Anaerococcus lactolyticus comb. nov., Anaerococcus hydrogenalis comb. nov., Anaerococcus vaginalis comb. nov. and Anaerococcus octavius sp. nov. The genus Gallicola gen. nov. contains a single species, Gallicola barnesae comb. nov.

  6. Human Ubc9 Is Involved in Intracellular HIV-1 Env Stability after Trafficking out of the Trans-Golgi Network in a Gag Dependent Manner

    PubMed Central

    Bohl, Christopher R.; Abrahamyan, Levon G.; Wood, Charles

    2013-01-01

    The cellular E2 Sumo conjugase, Ubc9 interacts with HIV-1 Gag, and is important for the assembly of infectious HIV-1 virions. In the previous study we demonstrated that in the absence of Ubc9, a defect in virion assembly was associated with decreased levels of mature intracellular Envelope (Env) that affected Env incorporation into virions and virion infectivity. We have further characterized the effect of Ubc9 knockdown on HIV Env processing and assembly. We found that gp160 stability in the endoplasmic reticulum (ER) and its trafficking to the trans-Golgi network (TGN) were unaffected, indicating that the decreased intracellular mature Env levels in Ubc9-depleted cells were due to a selective degradation of mature Env gp120 after cleavage from gp160 and trafficked out of the TGN. Decreased levels of Gag and mature Env were found to be associated with the plasma membrane and lipid rafts, which suggest that these viral proteins were not trafficked correctly to the assembly site. Intracellular gp120 were partially rescued when treated with a combination of lysosome inhibitors. Taken together our results suggest that in the absence of Ubc9, gp120 is preferentially degraded in the lysosomes likely before trafficking to assembly sites leading to the production of defective virions. This study provides further insight in the processing and packaging of the HIV-1 gp120 into mature HIV-1 virions. PMID:23861967

  7. Chimeric adenovirus type 5/35 vector encoding SIV gag and HIV env genes affords protective immunity against the simian/human immunodeficiency virus in monkeys.

    PubMed

    Someya, Kenji; Xin, Ke-Qin; Ami, Yasushi; Izumi, Yasuyuki; Mizuguchi, Hiroyuki; Ohta, Shinrai; Yamamoto, Naoki; Honda, Mitsuo; Okuda, Kenji

    2007-10-25

    Replication-defective adenovirus type 5 (Ad5) vector-based vaccines are widely known to induce strong immunity against immunodeficiency viruses. To exploit this immunogenicity while overcoming the potential problem of preexisting immunity against human adenoviruses type 5, we developed a recombinant chimeric adenovirus type 5 with type 35 fiber vector (rAd5/35). We initially produced a simian immunodeficiency virus (SIV) gag DNA plasmid (rDNA-Gag), a human immunodeficiency virus type 1 (HIV-1) 89.6 env DNA plasmid (rDNA-Env) and a recombinant Ad5/35 vector encoding the SIV gag and HIV env gene (rAd5/35-Gag and rAd5/35-Env). Prime-boost vaccination with rDNA-Gag and -Env followed by high doses of rAd5/35-Gag and -Env elicited higher levels of cellular immune responses than did rDNAs or rAd5/35s alone. When challenged with a pathogenic simian human immunodeficiency virus (SHIV), animals receiving a prime-boost regimen or rAd5/35s alone maintained a higher number of CD4(+) T cells and remarkably suppressed plasma viral RNA loads. These findings suggest the clinical promise of an rAd5/35 vector-based vaccine.

  8. AutoGen Version 5.0

    NASA Technical Reports Server (NTRS)

    Gladden, Roy E.; Khanampornpan, Teerapat; Fisher, Forest W.

    2010-01-01

    Version 5.0 of the AutoGen software has been released. Previous versions, variously denoted Autogen and autogen, were reported in two articles: Automated Sequence Generation Process and Software (NPO-30746), Software Tech Briefs (Special Supplement to NASA Tech Briefs), September 2007, page 30, and Autogen Version 2.0 (NPO- 41501), NASA Tech Briefs, Vol. 31, No. 10 (October 2007), page 58. To recapitulate: AutoGen (now signifying automatic sequence generation ) automates the generation of sequences of commands in a standard format for uplink to spacecraft. AutoGen requires fewer workers than are needed for older manual sequence-generation processes, and greatly reduces sequence-generation times. The sequences are embodied in spacecraft activity sequence files (SASFs). AutoGen automates generation of SASFs by use of another previously reported program called APGEN. AutoGen encodes knowledge of different mission phases and of how the resultant commands must differ among the phases. AutoGen also provides means for customizing sequences through use of configuration files. The approach followed in developing AutoGen has involved encoding the behaviors of a system into a model and encoding algorithms for context-sensitive customizations of the modeled behaviors. This version of AutoGen addressed the MRO (Mars Reconnaissance Orbiter) primary science phase (PSP) mission phase. On previous Mars missions this phase has more commonly been referred to as mapping phase. This version addressed the unique aspects of sequencing orbital operations and specifically the mission specific adaptation of orbital operations for MRO. This version also includes capabilities for MRO s role in Mars relay support for UHF relay communications with the MER rovers and the Phoenix lander.

  9. Exosomes carring gag/env of ALV-J possess negative effect on immunocytes.

    PubMed

    Wang, Guihua; Wang, Zhenzhen; Zhuang, Pingping; Zhao, Xiaomin; Cheng, Ziqiang

    2017-09-12

    J subgroup avian leukosis virus (ALV-J) is an exogenous retrovirus of avian. A key feature of ALV-J infection is leading to severe immunosuppressive characteristic of diseases. Viral components of retrovirus were reported closely associated with immunosuppression, and several similarities between exosomes and retrovirus preparations have lead to the hypotheses of retrovirus hijacker exosomes pathway. In this study, we purified exosomes from DF-1 cells infected and uninfected by ALV-J. Electron microscopy and mass spectrometry (MS) analysis showed that ALV-J not only increased the production of exosomes from ALV-J infected DF-1 cells (Exo-J) but also stimulated some proteins expression, especially ALV-J components secreted in exosomes. Immunosuppressive domain peptide (ISD) of envelope subunit transmembrane (TM) and gag of ALV-J were secreted in Exo-J. It has been reported that HIV gag was budded from endosome-like domains of the T cell plasma membrane. But env protein was first detected in exosomes from retrovirus infected cells. We found that Exo-J caused negative effects on splenocytes in a dose-dependant manner by flow cytometric analysis. And low dose of Exo-J activated immune activity of splenocytes, while high dose possessed immunosuppressive properties. Interestingly, Exo-J has no significant effects on the immunosuppression induced by ALV-J, and the immunosuppressive effects induced by Exo-J lower than that by ALV-J. Taken together, our data indicated that Exo-J supplied a microenvironment for the replication and transformation of ALV-J. Copyright © 2017. Published by Elsevier Ltd.

  10. Debroyerella gen. nov. and Ulladulla gen. nov., two new lysianassoid genera (Crustacea, Amphipoda, Lysianassoidea).

    PubMed

    Lowry, J K; Kilgallen, N M

    2015-02-19

    Two new genera and a new species of lysianassoid amphipods are described. Debroyerella gen. nov. is described for three Antarctic species previously assigned to the genus Cheirimedon. Ulladulla gen. nov. is described to accommodate the new species U. selje, from Australian waters. Diagnostic descriptions are given for the genera and all species are described in full.

  11. Unleashing Gen Y: Marketing Mars to Millennials

    NASA Technical Reports Server (NTRS)

    Leahy, Bart D.; Hidalgo, Loretta; Kloberdanz, Cassie

    2007-01-01

    Space advocates need to engage Generation Y (born 1977-1999).This outreach is necessary to recruit the next generation of scientists and engineers to explore Mars. Space advocates in the non-profit, private, and government sectors need to use a combination of technical communication, marketing, and politics, to develop messages that resonate with Gen Y. Until now, space messages have been generated by and for college-educated white males; Gen Y is much more diverse, including as much as one third minorities. Young women, too, need to be reached. My research has shown that messages emphasizing technology, fun, humor, and opportunity are the best means of reaching the Gen Y audience of 60 million (US population is 300 million). The important things space advocates must avoid are talking down to this generation, making false promises, or expecting them to "wait their turn" before they can participate. This is the MTV generation! We need to find ways of engaging Gen Y now to build a future where human beings can live and work on the planet Mars. In addition to the messages themselves, advocates need to keep up with Gen Y' s social networking and use of iPods, cell phones, and the Internet. NASA and space advocacy groups can use these tools for "viral marketing," where young people share targeted space-related information via cell phones or the Internet because they like it. Overall, Gen Y is a socially dynamic and media-savvy group; advocates' space messages need to be sincere, creative, and placed in locations where Gen Y lives. Mars messages must be memorable!

  12. A pilot study comparing the development of EIAV Env-specific antibodies induced by DNA/recombinant vaccinia-vectored vaccines and an attenuated Chinese EIAV vaccine.

    PubMed

    Meng, Qinglai; Lin, Yuezhi; Ma, Jian; Ma, Yan; Zhao, Liping; Li, Shenwei; Yang, Kai; Zhou, Jianhua; Shen, Rongxian; Zhang, Xiaoyan; Shao, Yiming

    2012-12-01

    Data from successful attenuated lentiviral vaccine studies indicate that fully mature Env-specific antibodies characterized by high titer, high avidity, and the predominant recognition of conformational epitopes are associated with protective efficacy. Although vaccination with a DNA prime/recombinant vaccinia-vectored vaccine boost strategy has been found to be effective in some trials with non-human primate/simian/human immunodeficiency virus (SHIV) models, it remains unclear whether this vaccination strategy could elicit mature equine infectious anemia virus (EIAV) Env-specific antibodies, thus protecting vaccinated horses against EIAV infection. Therefore, in this pilot study we vaccinated horses using a strategy based on DNA prime/recombinant Tiantan vaccinia (rTTV)-vectored vaccines encoding EIAV env and gag genes, and observed the development of Env-specific antibodies, neutralizing antibodies, and p26-specific antibodies. Vaccination with DNA induced low titer, low avidity, and the predominant recognition of linear epitopes by Env-specific antibodies, which was enhanced by boosting vaccinations with rTTV vaccines. However, the maturation levels of Env-specific antibodies induced by the DNA/rTTV vaccines were significantly lower than those induced by the attenuated vaccine EIAV(FDDV). Additionally, DNA/rTTV vaccines did not elicit broadly neutralizing antibodies. After challenge with a virulent EIAV strain, all of the vaccinees and control horses died from EIAV disease. These data indicate that the regimen of DNA prime/rTTV vaccine boost did not induce mature Env-specific antibodies, which might have contributed to immune protection failure.

  13. Single Amino Acid Substitution N659D in HIV-2 Envelope Glycoprotein (Env) Impairs Viral Release and Hampers BST-2 Antagonism.

    PubMed

    Dufrasne, François E; Lombard, Catherine; Goubau, Patrick; Ruelle, Jean

    2016-10-14

    BST-2 or tetherin is a host cell restriction factor that prevents the budding of enveloped viruses at the cell surface, thus impairing the viral spread. Several countermeasures to evade this antiviral factor have been positively selected in retroviruses: the human immunodeficiency virus type 2 (HIV-2) relies on the envelope glycoprotein (Env) to overcome BST-2 restriction. The Env gp36 ectodomain seems involved in this anti-tetherin activity, however residues and regions interacting with BST-2 are not clearly defined. Among 32 HIV-2 ROD Env mutants tested, we demonstrated that the asparagine residue at position 659 located in the gp36 ectodomain is mandatory to exert the anti-tetherin function. Viral release assays in cell lines expressing BST-2 showed a loss of viral release ability for the HIV-2 N659D mutant virus compared to the HIV-2 wild type virus. In bst-2 inactivated H9 cells, those differences were lost. Subtilisin treatment of infected cells demonstrated that the N659D mutant was more tethered at the cell surface. Förster resonance energy transfer (FRET) experiments confirmed a direct molecular link between Env and BST-2 and highlighted an inability of the mutant to bind BST-2. We also tested a virus presenting a truncation of 109 amino acids at the C-terminal part of Env, a cytoplasmic tail partial deletion that is spontaneously selected in vitro. Interestingly, viral release assays and FRET experiments indicated that a full Env cytoplasmic tail was essential in BST-2 antagonism. In HIV-2 infected cells, an efficient Env-mediated antagonism of BST-2 is operated through an intermolecular link involving the asparagine 659 residue as well as the C-terminal part of the cytoplasmic tail.

  14. A Pilot Study Comparing the Development of EIAV Env-Specific Antibodies Induced by DNA/Recombinant Vaccinia-Vectored Vaccines and an Attenuated Chinese EIAV Vaccine

    PubMed Central

    Meng, Qinglai; Lin, Yuezhi; Ma, Jian; Ma, Yan; Zhao, Liping; Li, Shenwei; Yang, Kai; Zhou, Jianhua; Shen, Rongxian

    2012-01-01

    Abstract Data from successful attenuated lentiviral vaccine studies indicate that fully mature Env-specific antibodies characterized by high titer, high avidity, and the predominant recognition of conformational epitopes are associated with protective efficacy. Although vaccination with a DNA prime/recombinant vaccinia-vectored vaccine boost strategy has been found to be effective in some trials with non-human primate/simian/human immunodeficiency virus (SHIV) models, it remains unclear whether this vaccination strategy could elicit mature equine infectious anemia virus (EIAV) Env-specific antibodies, thus protecting vaccinated horses against EIAV infection. Therefore, in this pilot study we vaccinated horses using a strategy based on DNA prime/recombinant Tiantan vaccinia (rTTV)-vectored vaccines encoding EIAV env and gag genes, and observed the development of Env-specific antibodies, neutralizing antibodies, and p26-specific antibodies. Vaccination with DNA induced low titer, low avidity, and the predominant recognition of linear epitopes by Env-specific antibodies, which was enhanced by boosting vaccinations with rTTV vaccines. However, the maturation levels of Env-specific antibodies induced by the DNA/rTTV vaccines were significantly lower than those induced by the attenuated vaccine EIAVFDDV. Additionally, DNA/rTTV vaccines did not elicit broadly neutralizing antibodies. After challenge with a virulent EIAV strain, all of the vaccinees and control horses died from EIAV disease. These data indicate that the regimen of DNA prime/rTTV vaccine boost did not induce mature Env-specific antibodies, which might have contributed to immune protection failure. PMID:23171359

  15. Identification of a HERV-K env surface peptide highly recognized in Rheumatoid Arthritis (RA) patients: a cross-sectional case-control study.

    PubMed

    Mameli, G; Erre, G L; Caggiu, E; Mura, S; Cossu, D; Bo, M; Cadoni, M L; Piras, A; Mundula, N; Colombo, E; Buscetta, G; Passiu, G; Sechi, L A

    2017-07-01

    Endogenous retroviruses (HERV) are believed to be pathogenic in several autoimmune diseases. Among them, HERV-K viruses have been reported recently to be involved in the pathogenesis of rheumatoid arthritis (RA). In this study we have explored the role of humoral immune response against HERV-K as a potential pathogenetic mechanism in RA. Four different peptides from the extracellular portion of the env protein of HERV-K (env-su19-37 , env-su109-126 , env-su164-186 , env-su209-226 ) were selected by bioinformatic analysis on the basis of their putative immunogenicity. Indirect enzyme-linked immunosorbent assay (ELISA) was then carried out to quantify antibodies against those peptides on blood samples of 70 consecutive RA patients and 71 healthy controls (HC). Differences between the two groups were analysed using the Mann-Whitney test. Potential correlations between RA laboratory, clinical descriptors and immunoglobulin (Ig)G levels were explored by bivariate regression analysis. Serum autoantibodies against one of four tested peptides of HERV-K (env-su19-37 ) were significantly higher in RA than in HC (19 versus 3%, P = 0·0025). Subgroup analysis showed no association between anti-HERV-K peptide humoral response and clinical, serological and clinimetric RA disease descriptors. Serum from RA patients in our series reacted significantly against HERV-K env-su19-37 peptide in comparison to the general population suggesting a role for the HERV-K- related, secondary antigenic-driven immune response in the pathogenesis of RA. Further studies are needed to confirm these results and to explore the role of this HERV-K surface peptide as a potential therapeutic target. © 2017 British Society for Immunology.

  16. Enhanced cell surface expression, immunogenicity and genetic stability resulting from a spontaneous truncation of HIV Env expressed by a recombinant MVA

    SciTech Connect

    Wyatt, Linda S. Belyakov, Igor M.; Earl, Patricia L.; Berzofsky, Jay A.; Moss, Bernard

    2008-03-15

    During propagation of modified vaccinia virus Ankara (MVA) encoding HIV 89.6 Env, a few viral foci stained very prominently. Virus cloned from such foci replicated to higher titers than the parent and displayed enhanced genetic stability on passage. Sequence analysis showed a single nucleotide deletion in the 89.6 env gene of the mutant that caused a frame shift and truncation of 115 amino acids from the cytoplasmic domain. The truncated Env was more highly expressed on the cell surface, induced higher antibody responses than the full-length Env, reacted with HIV neutralizing monoclonal antibodies and mediated CD4/co-receptor-dependent fusion. Intramuscular (IM), intradermal (ID) needleless, and intrarectal (IR) catheter inoculations gave comparable serum IgG responses. However, intraoral (IO) needleless injector route gave the highest IgA in lung washings and IR gave the highest IgA and IgG responses in fecal extracts. Induction of CTL responses in the spleens of individual mice as assayed by intracellular cytokine staining was similar with both the full-length and truncated Env constructs. Induction of acute and memory CTL in the spleens of mice immunized with the truncated Env construct by ID, IO, and IR routes was comparable and higher than by the IM route, but only the IR route induced CTL in the gut-associated lymphoid tissue. Thus, truncation of Env enhanced genetic stability as well as serum and mucosal antibody responses, suggesting the desirability of a similar modification in MVA-based candidate HIV vaccines.

  17. Three new anascosporic genera of the Saccharomycotina: Danielozyma gen. nov., Deakozyma gen. nov. and Middelhovenomyces gen. nov.

    PubMed

    Kurtzman, Cletus P; Robnett, Christie J

    2014-05-01

    Three new non-ascosporic, ascomycetous yeast genera are proposed based on their isolation from currently described species and genera. Phylogenetic placement of the genera was determined from analysis of nuclear gene sequences for D1/D2 large subunit rRNA, small subunit rRNA, translation elongation factor-1α and RNA polymerase II, subunits B1 and B2. The new taxa are: Deakozyma gen. nov., type species Deakozyma indianensis sp. nov. (type strain NRRL YB-1937, CBS 12903); Danielozyma gen. nov., type species Danielozyma ontarioensis comb. nov. (type strain NRRL YB-1246, CBS 8502); D. litseae comb. nov. (type strain NRRL YB-3246, CBS 8799); Middelhovenomyces gen. nov., type species Middelhovenomyces tepae comb. nov. (type strain NRRL Y-17670, CBS 5115) and M. petrohuensis comb. nov. (type strain NRRL Y-17663, CBS 8173).

  18. Enhancing Transport of Hydrogenophaga flava ENV735 for Bioaugmentation of Aquifers Contaminated with Methyl tert-Butyl Ether

    PubMed Central

    Streger, Sheryl H.; Vainberg, Simon; Dong, Hailiang; Hatzinger, Paul B.

    2002-01-01

    The gasoline oxygenate methyl tert-butyl ether (MTBE) has become a widespread contaminant in groundwater throughout the United States. Bioaugmentation of aquifers with MTBE-degrading cultures may be necessary to enhance degradation of the oxygenate in some locations. However, poor cell transport has sometimes limited bioaugmentation efforts in the past. The objective of this study was to evaluate the transport characteristics of Hydrogenophaga flava ENV735, a pure culture capable of growth on MTBE, and to improve movement of the strain through aquifer solids. The wild-type culture moved only a few centimeters in columns of aquifer sediment. An adhesion-deficient variant (H. flava ENV735:24) of the wild-type strain that moved more readily through sediments was obtained by sequential passage of cells through columns of sterile sediment. Hydrophobic and electrostatic interaction chromatography revealed that the wild-type strain is much more hydrophobic than the adhesion-deficient variant. Electrophoretic mobility assays and transmission electron microscopy showed that the wild-type bacterium contains two distinct subpopulations, whereas the adhesion-deficient strain has only a single, homogeneous population. Both the wild-type strain and adhesion-deficient variant degraded MTBE, and both were identified by 16S rRNA analysis as pure cultures of H. flava. The effectiveness of surfactants for enhancing transport of the wild-type strain was also evaluated. Many of the surfactants tested were toxic to ENV735; however, one nonionic surfactant, Tween 20, enhanced cell transport in sand columns. Improving microbial transport may lead to a more effective bioaugmentation strategy for MTBE-contaminated sites where indigenous oxygenate degraders are absent. PMID:12406751

  19. Detection of the human endogenous retrovirus ERV3-encoded Env-protein in human tissues using antibody-based proteomics.

    PubMed

    Fei, Chen; Atterby, Christina; Edqvist, Per-Henrik; Pontén, Fredrik; Zhang, Wei Wei; Larsson, Erik; Ryan, Frank P

    2014-01-01

    There is growing evidence to suggest that human endogenous retroviruses (HERVs) have contributed to human evolution, being expressed in development, normal physiology and disease. A key difficulty in the scientific evaluation of this potential viral contribution is the accurate demonstration of virally expressed protein in specific human cells and tissues. In this study, we have adopted the endogenous retrovirus, ERV3, as our test model in developing a reliable high-capacity methodology for the expression of such endogenous retrovirus-coded protein. Two affinity-purified polyclonal antibodies to ERV3 Env-encoded protein were generated to detect the corresponding protein expression pattern in specific human cells, tissues and organs. Sampling included normal tissues from 144 individuals ranging from childhood to old age. This included more than forty different tissues and organs and some 216 different cancer tissues representing the twenty commonest forms of human cancer. The Rudbeck Laboratory, Uppsala University and Uppsala University Hospital, Uppsala, Sweden. The potential expression at likely physiological level of the ERV3Env encoded protein in a wide range of human cells, tissues and organs. We found that ERV3 encoded Env protein is expressed at substantive levels in placenta, testis, adrenal gland, corpus luteum, Fallopian tubes, sebaceous glands, astrocytes, bronchial epithelium and the ducts of the salivary glands. Substantive expression was also seen in a variety of epithelial cells as well as cells known to undergo fusion in inflammation and in normal physiology, including fused macrophages, myocardium and striated muscle. This contrasted strongly with the low levels expressed in other tissues types. These findings suggest that this virus plays a significant role in human physiology and may also play a possible role in disease. This technique can now be extended to the study of other HERV genomes within the human chromosomes that may have contributed to

  20. The V3 Loop of HIV-1 Env Determines Viral Susceptibility to IFITM3 Impairment of Viral Infectivity.

    PubMed

    Wang, Yimeng; Pan, Qinghua; Ding, Shilei; Wang, Zhen; Yu, Jingyou; Finzi, Andrés; Liu, Shan-Lu; Liang, Chen

    2017-04-01

    Interferon-inducible transmembrane proteins (IFITMs) inhibit a broad spectrum of viruses, including HIV-1. IFITM proteins deter HIV-1 entry when expressed in target cells and also impair HIV-1 infectivity when expressed in virus producer cells. However, little is known about how viruses resist IFITM inhibition. In this study, we have investigated the susceptibilities of different primary isolates of HIV-1 to the inhibition of viral infectivity by IFITMs. Our results demonstrate that the infectivity of different HIV-1 primary isolates, including transmitted founder viruses, is diminished by IFITM3 to various levels, with strain AD8-1 exhibiting strong resistance. Further mutagenesis studies revealed that HIV-1 Env, and the V3 loop sequence in particular, determines the extent of inhibition of viral infectivity by IFITM3. IFITM3-sensitive Env proteins are also more susceptible to neutralization by soluble CD4 or the 17b antibody than are IFITM3-resistant Env proteins. Together, data from our study suggest that the propensity of HIV-1 Env to sample CD4-bound-like conformations modulates viral sensitivity to IFITM3 inhibition.IMPORTANCE Results of our study have revealed the key features of the HIV-1 envelope protein that are associated with viral resistance to the IFITM3 protein. IFITM proteins are important effectors in interferon-mediated antiviral defense. A variety of viruses are inhibited by IFITMs at the virus entry step. Although it is known that envelope proteins of several different viruses resist IFITM inhibition, the detailed mechanisms are not fully understood. Taking advantage of the fact that envelope proteins of different HIV-1 strains exhibit different degrees of resistance to IFITM3 and that these HIV-1 envelope proteins share the same domain structure and similar sequences, we performed mutagenesis studies and determined the key role of the V3 loop in this viral resistance phenotype. We were also able to associate viral resistance to IFITM3

  1. Overlapping cis sites used for splicing of HIV-1 env/nef and rev mRNAs.

    PubMed

    Swanson, A K; Stoltzfus, C M

    1998-12-18

    Alternative splicing is used to generate more than 30 human immunodeficiency virus type 1 (HIV-1) spliced and unspliced mRNAs from a single primary transcript. The abundance of HIV-1 mRNAs is determined by the efficiencies with which its different 5' and 3' splice sites are used. Three splice sites (A4c, A4a, and A4b) are upstream of the rev initiator AUG. RNAs spliced at A4c, A4a, and A4b are used as mRNAs for Rev. Another 3' splice site (A5) is immediately downstream of the rev initiator. RNAs spliced at A5 are used as mRNAs for Env and Nef. In this report, primer extension analysis of splicing intermediates was used to show that there are eight branch points in this region, all of which map to adenosine residues. In addition, cis elements recognized by the cellular splicing machinery overlap; the two most 3' branch points overlap with the AG dinucleotides at rev 3' splice sites A4a and A4b. Competition of the overlapping cis sites for different splicing factors may play a role in maintaining the appropriate balance of mRNAs in HIV-1-infected cells. In support of this possibility, mutations at rev 3' splice site A4b AG dinucleotide dramatically increased splicing of the env/nef 3' splice site A5. This correlated with increased usage of the four most 3' branch points, which include those within the rev 3' splice site AG dinucleotides. Consistent with these results, analysis of a mutant in which three of the four env/nef branch points were inactivated indicated that use of splice site A5 was inhibited and splicing was shifted predominantly to the most 5' rev 3' splice site A4c with preferential use of the two most 5' branch points. Our results suggest that spliceosomes formed at rev A4a-4b, rev A4c, and env/nef A5 3' splice sites each recognize different subsets of the eight branch point sequences.

  2. ATLAS Versus NextGen Model Atmospheres: A Combined Analysis of Synthetic Spectral Energy Distributions

    NASA Astrophysics Data System (ADS)

    Bertone, E.; Buzzoni, A.; Chávez, M.; Rodríguez-Merino, L. H.

    2004-08-01

    We carried out a critical appraisal of the two theoretical models, Kurucz' ATLAS9 and PHOENIX/NextGen, for stellar atmosphere synthesis. Our tests relied on the theoretical fit of spectral energy distributions (SEDs) for a sample of 334 target stars along the whole spectral-type sequence, from the classical optical catalogs of Gunn & Stryker and Jacoby et al. The best-fitting physical parameters (Teff, logg) of stars allowed an independent calibration of the temperature and bolometric scale versus empirical classification parameters (i.e., spectral type and MK luminosity class); in addition, the comparison of the synthetic templates from the ATLAS and NextGen grids allowed us to probe the capability of the models to match spectrophotometric properties of real stars and assess the impact of the different input physics. We can sketch the following main conclusions of our analysis: (1) Fitting accuracy of both theoretical libraries drastically degrades at low Teff at which both ATLAS and NextGen models still fail to properly account for the contribution of molecular features in the observed SED of K-M stars. (2) Compared with empirical calibrations, both ATLAS and NextGen fits tend, on average, to predict slightly warmer (by 4%-8%) Teff for both giant and dwarf stars of fixed spectral type, but ATLAS provides, in general, a sensibly better fit (a factor of 2 lower σ of flux residuals) than NextGen. (3) There is a striking tendency of NextGen to label target stars with an effective temperature and surface gravity higher than that of ATLAS. The effect is especially evident for MK I-III objects for which about one in four stars is clearly misclassified by NextGen in logg. This is a consequence of some ``degeneracy'' in the solution space, partly induced by the different input physics and geometry constraints in the computation of the integrated emerging flux (ATLAS model atmospheres assume standard plane-parallel layers, while NextGen adopts, for low-gravity stars, a

  3. Changes in Structure and Antigenicity of HIV-1 Env Trimers Resulting from Removal of a Conserved CD4 Binding Site-Proximal Glycan

    PubMed Central

    Liang, Yu; Guttman, Miklos; Williams, James A.; Verkerke, Hans; Alvarado, Daniel

    2016-01-01

    ABSTRACT The envelope glycoprotein (Env) is the major target for HIV-1 broadly neutralizing antibodies (bNAbs). One of the mechanisms that HIV has evolved to escape the host's immune response is to mask conserved epitopes on Env with dense glycosylation. Previous studies have shown that the removal of a particular conserved glycan at N197 increases the neutralization sensitivity of the virus to antibodies targeting the CD4 binding site (CD4bs), making it a site of significant interest from the perspective of vaccine design. At present, the structural consequences that result from the removal of the N197 glycan have not been characterized. Using native-like SOSIP trimers, we examine the effects on antigenicity and local structural dynamics resulting from the removal of this glycan. A large increase in the binding of CD4bs and V3-targeting antibodies is observed for the N197Q mutant in trimeric Env, while no changes are observed with monomeric gp120. While the overall structure and thermostability are not altered, a subtle increase in the flexibility of the variable loops at the trimeric interface of adjacent protomers is evident in the N197Q mutant by hydrogen-deuterium exchange mass spectrometry. Structural modeling of the glycan chains suggests that the spatial occupancy of the N197 glycan leads to steric clashes with CD4bs antibodies in the Env trimer but not monomeric gp120. Our results indicate that the removal of the N197 glycan enhances the exposure of relevant bNAb epitopes on Env with a minimal impact on the overall trimeric structure. These findings present a simple modification for enhancing trimeric Env immunogens in vaccines. IMPORTANCE The HIV-1 Env glycoprotein presents a dense patchwork of host cell-derived N-linked glycans. This so-called glycan shield is considered to be a major protective mechanism against immune recognition. While the positions of many N-linked glycans are isolate specific, some are highly conserved and are believed to play key

  4. Early Steps of Jaagsiekte Sheep Retrovirus-Mediated Cell Transformation Involve the Interaction between Env and the RALBP1 Cellular Protein

    PubMed Central

    Monot, Margaux; Erny, Alexandra; Gineys, Barbara; Desloire, Sophie; Dolmazon, Christine; Aublin-Gex, Anne; Lotteau, Vincent; Archer, Fabienne

    2015-01-01

    ABSTRACT Ovine pulmonary adenocarcinoma is a naturally occurring lung cancer in sheep induced by the Jaagsiekte sheep retrovirus (JSRV). Its envelope glycoprotein (Env) carries oncogenic properties, and its expression is sufficient to induce in vitro cell transformation and in vivo lung adenocarcinoma. The identification of cellular partners of the JSRV envelope remains crucial for deciphering mechanisms leading to cell transformation. We initially identified RALBP1 (RalA binding protein 1; also known as RLIP76 or RIP), a cellular protein implicated in the ras pathway, as a partner of JSRV Env by yeast two-hybrid screening and confirmed formation of RALBP1/Env complexes in mammalian cells. Expression of the RALBP1 protein was repressed in tumoral lungs and in tumor-derived alveolar type II cells. Through its inhibition using specific small interfering RNA (siRNA), we showed that RALBP1 was involved in envelope-induced cell transformation and in modulation of the mTOR (mammalian target of rapamycin)/p70S6K pathway by the retroviral envelope. IMPORTANCE JSRV-induced lung adenocarcinoma is of importance for the sheep industry. While the envelope has been reported as the oncogenic determinant of the virus, the cellular proteins directly interacting with Env are still not known. Our report on the formation of RALBP/Env complexes and the role of this interaction in cell transformation opens up a new hypothesis for the dysregulation observed upon virus infection in sheep. PMID:26041289

  5. Characterization of Env antigenicity of feline foamy virus (FeFV) using FeFV-infected cat sera and a monoclonal antibody.

    PubMed

    Phung, Hang T T; Tohya, Yukinobu; Miyazawa, Takayuki; Akashi, Hiroomi

    2005-04-10

    To characterize neutralizing antigenicity in relation to env genotypes of feline foamy virus (FeFV), serological analyses were performed using FeFV-infected cat sera and several field isolates including two env genotypes (F17- and FUV-types). Since three cats from which FeFV were isolated were found to have undetectable titers of virus neutralization (VN) antibodies, even to the homologous virus, VN antibodies were further examined with complement supplementation as an enhancement factor. With the presence of complement, the VN titers of FeFV-infected cat sera increased drastically. Although most of serum samples neutralized strains of either env genotype, sera sampled from two cats neutralized all the strains examined at similar titers, suggesting that superinfection with both env genotypes of FeFV might have occurred in the two cats. Further, we produced a monoclonal antibody (mAb) specifically neutralizing FeFV strains of FUV-type. The mAb was shown to have higher affinity to an epitope on Env of FUV-type than that of F17-type by immunoprecipitation assay. This study supplies basic information important for studies on FeFV vector development as well as on the relationship between the virus and the host immune response.

  6. Immunization with an SIV-based IDLV Expressing HIV-1 Env 1086 Clade C Elicits Durable Humoral and Cellular Responses in Rhesus Macaques

    PubMed Central

    Negri, Donatella; Blasi, Maria; LaBranche, Celia; Parks, Robert; Balachandran, Harikrishnan; Lifton, Michelle; Shen, Xiaoying; Denny, Thomas; Ferrari, Guido; Vescio, Maria Fenicia; Andersen, Hanne; Montefiori, David C; Tomaras, Georgia D; Liao, Hua-Xin; Santra, Sampa; Haynes, Barton F; Klotman, Mary E; Cara, Andrea

    2016-01-01

    The design of an effective HIV-1 vaccine remains a major challenge. Several vaccine strategies based on viral vectors have been evaluated in preclinical and clinical trials, with largely disappointing results. Integrase defective lentiviral vectors (IDLV) represent a promising vaccine candidate given their ability to induce durable and protective immune responses in mice after a single immunization. Here, we evaluated the immunogenicity of a SIV-based IDLV in nonhuman primates. Six rhesus monkeys were primed intramuscularly with IDLV-Env and boosted with the same vector after 1 year. A single immunization with IDLV-Env induced broad humoral and cellular immune responses that waned over time but were still detectable at 1 year postprime. The boost with IDLV-Env performed at 1 year from the prime induced a remarkable increase in both antibodies and T-cell responses. Antibody binding specificity showed a predominant cross-clade gp120-directed response. Monkeys' sera efficiently blocked anti-V2 and anti-CD4 binding site antibodies, neutralized the tier 1 MW965.26 pseudovirus and mediated antibody-dependent cellular cytotoxicity (ADCC). Durable polyfunctional Env-specific T-cell responses were also elicited. Our study demonstrates that an IDLV-Env-based vaccine induces functional, comprehensive, and durable immune responses in Rhesus macaques. These results support further evaluation of IDLV as a new HIV-1 vaccine delivery platform. PMID:27455880

  7. The conserved His8 of the Moloney murine leukemia virus Env SU subunit directs the activity of the SU-TM disulphide bond isomerase

    SciTech Connect

    Li Kejun; Zhang, Shujing; Kronqvist, Malin; Ekstroem, Maria; Wallin, Michael; Garoff, Henrik . E-mail: henrik.garoff@cbt.ki.se

    2007-04-25

    Murine leukemia virus (MLV) fusion is controlled by isomerization of the disulphide bond between the receptor-binding surface (SU) and fusion-active transmembrane subunits of the Env-complex. The bond is in SU linked to a CXXC motif. This carries a free thiol that upon receptor binding can be activated (ionized) to attack the disulphide and rearrange it into a disulphide isomer within the motif. To find out whether His8 in the conserved SPHQ sequence of Env directs thiol activation, we analyzed its ionization in MLV vectors with wtEnv and Env with His8 deleted or substituted for Tyr or Arg, which partially or completely arrests fusion. The ionization was monitored by following the pH effect on isomerization in vitro by Ca{sup 2+} depletion or in vivo by receptor binding. We found that wtEnv isomerized optimally at slightly basic pH whereas the partially active mutant required higher and the inactive mutants still higher pH. This suggests that His8 directs the ionization of the CXXC thiol.

  8. GenEng: A dialogue-based natural language interface to the GenBank

    SciTech Connect

    Karimi, E.

    1993-12-31

    To sequence all the genes in human DNA and analyze their functions, the Human Genome Project has been collecting a large body of data in data banks such as GenBank{copyright}. One of the important issues concerning computational biologists is the availability of the tools which can provide easy and efficient access to these data banks. In this paper, the author will give a brief overview of different approaches to the design of a relational database interface. The paper concentrates on the design of GenEng, a dialogue based natural language interface for information retrieval from the GenBank relational database.

  9. Colon Cancer on The Rise Among Gen Xers, Millennials

    MedlinePlus

    ... Colon Cancer on the Rise Among Gen Xers, Millennials And an old adversary -- the obesity epidemic -- may ... their early 50s and younger -- Gen Xers and millennials -- are experiencing significant increases in colon and rectal ...

  10. Variability of the env gene in cynomolgus macaques persistently infected with human immunodeficiency virus type 2 strain ben.

    PubMed Central

    Tolle, T; Petry, H; Bachmann, B; Hunsmann, G; Lüke, W

    1994-01-01

    The sequence variability of distinct regions of the proviral env gene of human immunodeficiency virus type 2 strain ben (HIV-2ben) isolated sequentially over 3 to 4 years from six experimentally infected macaques was studied. The regions investigated were homologous to the V1, V2, V3, V4, V5, and V7 hypervariable regions identified in the env genes of HIV-1 and simian immunodeficiency virus SIVmac, respectively. In contrast to findings with HIV-1 and SIVmac, the V1- and V2-homologous regions were found to be highly conserved during the course of the HIV-2ben infection in macaques. The V3-homologous region showed a degree of variation comparable to that of HIV-1 but not of SIV. In the V4-, V5-, and V7-homologous regions, mutation hot spots were detected in most reisolates of the infected monkeys. Most of these mutations occurred during the first 10 weeks after infection. After 50 weeks, new mutations were rarely detected. At most mutation sites, a dynamic equilibrium between the mutated viral isotype and the infecting predominant wild type was present. This equilibrium might prevent an accumulation of mutations in isolates later in the course of infection. PMID:8139054

  11. In-line alignment and Mg2+ coordination at the cleavage site of the env22 twister ribozyme

    PubMed Central

    Ren, Aiming; Košutić, Marija; Rajashankar, Kanagalaghatta R.; Frener, Marina; Santner, Tobias; Westhof, Eric; Micura, Ronald; Patel, Dinshaw J.

    2015-01-01

    Small self-cleaving nucleolytic ribozymes contain catalytic domains that accelerate site-specific cleavage/ligation of phosphodiester backbones. We report on the 2.9-Å crystal structure of the env22 twister ribozyme, which adopts a compact tertiary fold stabilized by co-helical stacking, double-pseudoknot formation and long-range pairing interactions. The U-A cleavage site adopts a splayed-apart conformation with the modeled 2′-O of U positioned for in-line attack on the adjacent to-be-cleaved P-O5′ bond. Both an invariant guanosine and a Mg2+ are directly coordinated to the non-bridging phosphate oxygens at the U-A cleavage step, with the former positioned to contribute to catalysis and the latter to structural integrity. The impact of key mutations on cleavage activity identified an invariant guanosine that contributes to catalysis. Our structure of the in-line aligned env22 twister ribozyme is compared with two recently-reported twister ribozymes structures, which adopt similar global folds, but differ in conformational features around the cleavage site. PMID:25410397

  12. Position-specific automated processing of V3 env ultra-deep pyrosequencing data for predicting HIV-1 tropism

    PubMed Central

    Jeanne, Nicolas; Saliou, Adrien; Carcenac, Romain; Lefebvre, Caroline; Dubois, Martine; Cazabat, Michelle; Nicot, Florence; Loiseau, Claire; Raymond, Stéphanie; Izopet, Jacques; Delobel, Pierre

    2015-01-01

    HIV-1 coreceptor usage must be accurately determined before starting CCR5 antagonist-based treatment as the presence of undetected minor CXCR4-using variants can cause subsequent virological failure. Ultra-deep pyrosequencing of HIV-1 V3 env allows to detect low levels of CXCR4-using variants that current genotypic approaches miss. However, the computation of the mass of sequence data and the need to identify true minor variants while excluding artifactual sequences generated during amplification and ultra-deep pyrosequencing is rate-limiting. Arbitrary fixed cut-offs below which minor variants are discarded are currently used but the errors generated during ultra-deep pyrosequencing are sequence-dependant rather than random. We have developed an automated processing of HIV-1 V3 env ultra-deep pyrosequencing data that uses biological filters to discard artifactual or non-functional V3 sequences followed by statistical filters to determine position-specific sensitivity thresholds, rather than arbitrary fixed cut-offs. It allows to retain authentic sequences with point mutations at V3 positions of interest and discard artifactual ones with accurate sensitivity thresholds. PMID:26585833

  13. N-terminally myristoylated feline foamy virus Gag allows Env-independent budding of sub-viral particles.

    PubMed

    Liu, Yang; Kim, Yong-Boum; Löchelt, Martin

    2011-11-01

    Foamy viruses (FVs) are distinct retroviruses classified as Spumaretrovirinae in contrast to the other retroviruses, the Orthoretrovirinae. As a unique feature of FVs, Gag is not sufficient for sub-viral particle (SVP) release. In primate and feline FVs (PFV and FFV), particle budding completely depends on the cognate FV Env glycoproteins. It was recently shown that an artificially added N-terminal Gag myristoylation signal (myr-signal) overcomes this restriction in PFV inducing an Orthoretrovirus-like budding phenotype. Here we show that engineered, heterologous N-terminal myr-signals also induce budding of the distantly related FFV Gag. The budding efficiency depends on the myr-signal and its location relative to the N-terminus of Gag. When the first nine amino acid residues of FFV Gag were replaced by known myr-signals, the budding efficiency as determined by the detection of extracellular SVPs was low. In contrast, adding myr-signals to the intact N-terminus of FFV Gag resulted in a more efficient SVP release. Importantly, budding of myr-Gag proteins was sensitive towards inhibition of cellular N-myristoyltransferases. As expected, the addition or insertion of myr-signals that allowed Env-independent budding of FFV SVPs also retargeted Gag to plasma membrane-proximal sites and other intracellular membrane compartments. The data confirm that membrane-targeted FV Gag has the capacity of SVP formation.

  14. [Functional analysis of Grp and Iris, the gag and env domesticated errantivirus genes, in the Drosophila melanogaster genome].

    PubMed

    Makhnovskii, P A; Kuzmin, I V; Nefedova, L N; Kima, A I

    2016-01-01

    Drosophila melanogaster is the only invertebrate that contains endogenous retroviruses, which are called errantiviruses. Two domesticated genes, Grp and Iris, which originate from errantivirus gag and env, respectively, have been found in the D. melanogaster genome. The functions performed by the genes in Drosophila are still unclear. To identify the functions of domesticated gag and env in the D. melanogaster genome, expression of Iris and Grp was studied in strains differing by the presence or absence of the functional gypsy errantivirus. In addition, the expression levels were measured after injection of gram-positive and gram-negative bacteria, which activate different immune response pathways, and exposure to various abiotic stress factors. The presence of functional D. melanogaster retrovirus gypsy was found to increase the Grp expression level in somatic tissues of the carcass, while exerting no effect on the Iris expression level. Activation of the immune response in D. melanogaster by bacteria Bacillus cereus increased the Grp expression level and did not affect Iris expression. As for the effects of abiotic stress factors (oxidative stress, starvation, and heat and cold stress), the Grp expression level increased in response to starvation in D. melanogaster females, and the Iris expression level was downregulated in heat shock and oxidative stress. Based on the findings, Grp was assumed to play a direct role in the immune response in D. melanogaster; Iris is not involved in immune responses, but and apparently performs a cell function that is inhibited in stress.

  15. Coexpression of the simian immunodeficiency virus Env and Rev proteins by a recombinant human adenovirus host range mutant.

    PubMed Central

    Cheng, S M; Lee, S G; Ronchetti-Blume, M; Virk, K P; Mizutani, S; Eichberg, J W; Davis, A; Hung, P P; Hirsch, V M; Chanock, R M

    1992-01-01

    Recombinant human adenoviruses (Ads) that replicate in the intestinal tract offer a novel, yet practical, means of immunoprophylaxis against a wide variety of viral and bacterial pathogens. For some infectious agents such as human immunodeficiency virus (HIV), the potential for residual infectious material in vaccine preparations must be eliminated. Therefore, recombinant human Ads that express noninfectious HIV or other microbial proteins are attractive vaccine candidates. To test such an approach for HIV, we chose an experimental model of AIDS based on simian immunodeficiency virus (SIV) infection of macaques. Our data demonstrate that the SIV Env gene products are expressed in cultured cells after infection with a recombinant Ad containing both SIV env and rev genes. An E3 deletion vector derived from a mutant of human Ad serotype 5 that efficiently replicates in both human and monkey cells was used to bypass the usual host range restriction of Ad infection. In addition, we show that the SIV rev gene is properly spliced from a single SIV subgenomic DNA fragment and that the Rev protein is expressed in recombinant Ad-SIV-infected human as well as monkey cells. The expression of SIV gene products in suitable live Ad vectors provides an excellent system for studying the regulation of SIV gene expression in cultured cells and evaluating the immunogenicity and protective efficacy of SIV proteins in macaques. Images PMID:1404612

  16. Model Building and Refinement of a Natively Glycosylated HIV-1 Env Protein by High-Resolution Cryoelectron Microscopy.

    PubMed

    Lee, Jeong Hyun; de Val, Natalia; Lyumkis, Dmitry; Ward, Andrew B

    2015-10-06

    Secretory and membrane proteins from mammalian cells undergo post-translational modifications, including N-linked glycosylation, which can result in a large number of possible glycoforms. This sample heterogeneity can be problematic for structural studies, particularly X-ray crystallography. Thus, crystal structures of heavily glycosylated proteins such as the HIV-1 Env viral spike protein have been determined by removing the majority of glycans. This step is most frequently carried out using Endoglycosidase H (EndoH) and requires that all expressed glycans be in the high-mannose form, which is often not the native glycoform. With significantly improved technologies in single-particle cryoelectron microscopy, we demonstrate that it is now possible to refine and build natively glycosylated HIV-1 Env structures in solution to 4.36 Å resolution. At this resolution we can now analyze the complete epitope of a broadly neutralizing antibody (bnAb), PGT128, in the context of the trimer expressed with native glycans.

  17. Robust antigen-specific humoral immune responses to sublingually delivered adenoviral vectors encoding HIV-1 Env: association with mucoadhesion and efficient penetration of the sublingual barrier.

    PubMed

    Domm, William; Brooks, Lauren; Chung, Hung Li; Feng, Changyong; Bowers, William J; Watson, Gene; McGrath, James L; Dewhurst, Stephen

    2011-09-16

    The efficient induction of virus-specific mucosal antibodies is an important unmet objective in Human Immunodeficiency Virus Type-1 (HIV-1) vaccine research. One promising approach is sublingual (SL) immunization. We examined the effectiveness of SL delivery of two different viral vectors: (i) a recombinant adenovirus (rAd5), and (ii) a Herpes Simplex Virus Type-1 amplicon vector (HSV-1). Initial in vitro videomicroscopy experiments showed that rAd5 particles were trapped in saliva (i.e., that Ad5 was mucoadhesive) - unlike HSV-1 virions, which migrated freely in both saliva and water. In vivo imaging studies in mice revealed that only the rAd5 vector efficiently transduced the SL epithelium. Consistent with this, SL delivery of an rAd5 encoding HIV-1 envelope glycoprotein (Env) resulted in robust antigen-specific antibody responses in plasma and in vaginal washes, whereas SL delivery of a HSV-1 amplicon vector encoding HIV-1 Env failed to elicit Env-specific antibodies. In contrast, both vectors elicited equivalent humoral responses following intramuscular (IM) delivery. Finally, SL delivery of the rAd5:Env vector resulted in elevated levels of Env-specific serum IgA, and vaginal IgA and IgG, when compared to IM delivery of the same vector. These results findings shed light on vector properties (mucoadhesion, penetration of the sublingual barrier) which may be important for the induction of potent humoral immune responses following sublingual vector administration. Our data also show that SL delivery of an Env-encoding rAd5 vector can elicit a potent antigen-specific mucosal antibody response in the absence of adjuvant. Overall, these findings support the further exploration of the SL delivery route for HIV-1 vaccine delivery.

  18. T-Cell Immune Responses Against Env from CRF12_BF and Subtype B HIV-1 Show High Clade-Specificity that Can Be Overridden by Multiclade Immunizations

    PubMed Central

    Mónaco, Daniela C.; Rodríguez, Ana M.; Pascutti, María F.; Carobene, Mauricio; Falivene, Juliana; Gómez, Alejandro; Maeto, Cynthia; Turk, Gabriela; Nájera, José L.; Esteban, Mariano; Gherardi, M. Magdalena

    2011-01-01

    Background The extreme genetic diversity of the human immunodeficiency virus type 1 (HIV-1) poses a daunting challenge to the generation of an effective AIDS vaccine. In Argentina, the epidemic is characterized by the high prevalence of infections caused by subtype B and BF variants. The aim of this study was to characterize in mice the immunogenic and antigenic properties of the Env protein from CRF12_BF in comparison with clade B, employing prime-boost schemes with the combination of recombinant DNA and vaccinia virus (VV) vectors. Methodology/Principal Findings As determined by ELISPOT from splenocytes of animals immunized with either EnvBF or EnvB antigens, the majority of the cellular responses to Env were found to be clade-specific. A detailed peptide mapping of the responses reveal that when there is cross-reactivity, there are no amino acid changes in the peptide sequence or were minimal and located at the peptide ends. In those cases, analysis of T cell polifunctionality and affinity indicated no differences with respect to the cellular responses found against the original homologous sequence. Significantly, application of a mixed immunization combining both clades (B and BF) induced a broader cellular response, in which the majority of the peptides targeted after the single clade vaccinations generated a positive response. In this group we could also find significant cellular and humoral responses against the whole gp120 protein from subtype B. Conclusions/Significance This work has characterized for the first time the immunogenic peptides of certain EnvBF regions, involved in T cell responses. It provides evidence that to improve immune responses to HIV there is a need to combine Env antigens from different clades, highlighting the convenience of the inclusion of BF antigens in future vaccines for geographic regions where these HIV variants circulate. PMID:21364754

  19. Mucosal Immunization of Lactating Female Rhesus Monkeys with a Transmitted/Founder HIV-1 Envelope Induces Strong Env-Specific IgA Antibody Responses in Breast Milk

    PubMed Central

    Fouda, Genevieve G. A.; Amos, Joshua D.; Wilks, Andrew B.; Pollara, Justin; Ray, Caroline A.; Chand, Anjali; Kunz, Erika L.; Liebl, Brooke E.; Whitaker, Kaylan; Carville, Angela; Smith, Shannon; Colvin, Lisa; Pickup, David J.; Staats, Herman F.; Overman, Glenn; Eutsey-Lloyd, Krissey; Parks, Robert; Chen, Haiyan; LaBranche, Celia; Barnett, Susan; Tomaras, Georgia D.; Ferrari, Guido; Montefiori, David C.; Liao, Hua-Xin; Letvin, Norman L.; Haynes, Barton F.

    2013-01-01

    We previously demonstrated that vaccination of lactating rhesus monkeys with a DNA prime/vector boost strategy induces strong T-cell responses but limited envelope (Env)-specific humoral responses in breast milk. To improve vaccine-elicited antibody responses in milk, hormone-induced lactating rhesus monkeys were vaccinated with a transmitted/founder (T/F) HIV Env immunogen in a prime-boost strategy modeled after the moderately protective RV144 HIV vaccine. Lactating rhesus monkeys were intramuscularly primed with either recombinant DNA (n = 4) or modified vaccinia virus Ankara (MVA) poxvirus vector (n = 4) expressing the T/F HIV Env C.1086 and then boosted twice intramuscularly with C.1086 gp120 and the adjuvant MF59. The vaccines induced Env-binding IgG and IgA as well as neutralizing and antibody-dependent cellular cytotoxicity (ADCC) responses in plasma and milk of most vaccinated animals. Importantly, plasma neutralization titers against clade C HIV variants MW965 (P = 0.03) and CAP45 (P = 0.04) were significantly higher in MVA-primed than in DNA-primed animals. The superior systemic prime-boost regimen was then compared to a mucosal-boost regimen, in which animals were boosted twice intranasally with C.1086 gp120 and the TLR 7/8 agonist R848 following the same systemic prime. While the systemic and mucosal vaccine regimens elicited comparable levels of Env-binding IgG antibodies, mucosal immunization induced significantly stronger Env-binding IgA responses in milk (P = 0.03). However, the mucosal regimen was not as potent at inducing functional IgG responses. This study shows that systemic MVA prime followed by either intranasal or systemic protein boosts can elicit strong humoral responses in breast milk and may be a useful strategy to interrupt postnatal HIV-1 transmission. PMID:23596289

  20. Accurate predictions of population-level changes in sequence and structural properties of HIV-1 Env using a volatility-controlled diffusion model

    PubMed Central

    DeLeon, Orlando; Hodis, Hagit; O’Malley, Yunxia; Johnson, Jacklyn; Salimi, Hamid; Zhai, Yinjie; Winter, Elizabeth; Remec, Claire; Eichelberger, Noah; Van Cleave, Brandon; Puliadi, Ramya; Harrington, Robert D.; Stapleton, Jack T.; Haim, Hillel

    2017-01-01

    The envelope glycoproteins (Envs) of HIV-1 continuously evolve in the host by random mutations and recombination events. The resulting diversity of Env variants circulating in the population and their continuing diversification process limit the efficacy of AIDS vaccines. We examined the historic changes in Env sequence and structural features (measured by integrity of epitopes on the Env trimer) in a geographically defined population in the United States. As expected, many Env features were relatively conserved during the 1980s. From this state, some features diversified whereas others remained conserved across the years. We sought to identify “clues” to predict the observed historic diversification patterns. Comparison of viruses that cocirculate in patients at any given time revealed that each feature of Env (sequence or structural) exists at a defined level of variance. The in-host variance of each feature is highly conserved among individuals but can vary between different HIV-1 clades. We designate this property “volatility” and apply it to model evolution of features as a linear diffusion process that progresses with increasing genetic distance. Volatilities of different features are highly correlated with their divergence in longitudinally monitored patients. Volatilities of features also correlate highly with their population-level diversification. Using volatility indices measured from a small number of patient samples, we accurately predict the population diversity that developed for each feature over the course of 30 years. Amino acid variants that evolved at key antigenic sites are also predicted well. Therefore, small “fluctuations” in feature values measured in isolated patient samples accurately describe their potential for population-level diversification. These tools will likely contribute to the design of population-targeted AIDS vaccines by effectively capturing the diversity of currently circulating strains and addressing properties

  1. Robust Antigen-Specific Humoral Immune Responses to Sublingually Delivered Adenoviral Vectors Encoding HIV-1 Env: Association with Mucoadhesion and Efficient Penetration of the Sublingual Barrier

    PubMed Central

    Domm, William; Brooks, Lauren; Chung, Hung Li; Feng, Changyong; Bowers, William J.; Watson, Gene; McGrath, James L.; Dewhurst, Stephen

    2011-01-01

    The efficient induction of virus-specific mucosal antibodies is an important unmet objective in Human Immunodeficiency Virus Type-1 (HIV-1) vaccine research. One promising approach is sublingual (SL) immunization. We examined the effectiveness of SL delivery of two different viral vectors: (i) a recombinant adenovirus (rAd5), and (ii) a Herpes Simplex Virus Type-1 amplicon vector (HSV-1). Initial in vitro videomicroscopy experiments showed that rAd5 particles were trapped in saliva (i.e., that Ad5 was mucoadhesive) - unlike HSV-1 virions, which migrated freely in both saliva and water. In vivo imaging studies in mice revealed that only the rAd5 vector efficiently transduced the SL epithelium. Consistent with this, SL delivery of an rAd5 encoding HIV-1 envelope glycoprotein (Env) resulted in robust antigen-specific antibody responses in plasma and in vaginal washes, whereas SL delivery of a HSV-1 amplicon vector encoding HIV-1 Env failed to elicit Env-specific antibodies. In contrast, both vectors elicited equivalent humoral responses following intramuscular (IM) delivery. Finally, SL delivery of the rAd5:Env vector resulted in elevated levels of Env-specific serum IgA, and vaginal IgA and IgG, when compared to IM delivery of the same vector. These results findings shed light on vector properties (mucoadhesion, penetration of the sublingual barrier) which may be important for the induction of potent humoral immune responses following sublingual vector administration. Our data also show that SL delivery of an Env-encoding rAd5 vector can elicit a potent antigen-specific mucosal antibody response in the absence of adjuvant. Overall, these findings support the further exploration of the SL delivery route for HIV-1 vaccine delivery. PMID:21801777

  2. VIEW OF GEN. LOGAN STATUE LOOKING SOUTHWEST WITH VISTA AS ...

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

    VIEW OF GEN. LOGAN STATUE LOOKING SOUTHWEST WITH VISTA AS TO THE GEN. THOMAS STATUE DOWN VERMONT AVENUE AND THE GEN. SCOTT STATUE DOWN RHODE ISLAND AVENUE IN THE BACKGROUND - Logan Circle, Vermont Avenue, Rhode Island Avenue, & Thirteenth Street, Washington, District of Columbia, DC

  3. 75 FR 56654 - RTCA NextGen Advisory Committee (NAC)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-09-16

    ... Federal Aviation Administration RTCA NextGen Advisory Committee (NAC) AGENCY: Federal Aviation Administration (FAA), DOT. ACTION: Notice of RTCA NextGen Advisory Committee (NAC) SUMMARY: The FAA published a... public of a meeting of RTCA NextGen Advisory Committee. The Agenda in that notice has been revised....

  4. 75 FR 54221 - RTCA NextGen Advisory Committee (NAC)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-09-03

    ...] [FR Doc No: 2010-22098] DEPARTMENT OF TRANSPORTATION Federal Aviation Administration RTCA NextGen Advisory Committee (NAC) AGENCY: Federal Aviation Administration (FAA), DOT. ACTION: Notice of RTCA NextGen... NextGen Advisory Committee (NAC). DATES: The meeting will be held September 23, 2010, from 8:30 a.m....

  5. 77 FR 2342 - Fifth Meeting: RTCA, Next Gen Advisory Committee

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-01-17

    ... TRANSPORTATION Federal Aviation Administration Fifth Meeting: RTCA, Next Gen Advisory Committee AGENCY: Federal Aviation Administration (FAA), U.S. Department of Transportation (DOT). ACTION: Notice of RTCA, NextGen... RTCA, NextGen Advisory Committee. DATES: The meeting will be held February 3, 2012, from 9:30...

  6. VitisGen: accelerating grape cultivar improvement

    USDA-ARS?s Scientific Manuscript database

    VitisGen is a multiple institute collaborative project funded by the USDA SCRI program, with a long term goal to accelerate grape cultivar improvement by using cutting-edge molecular marker technologies, rigorous centralized facilities to characterize traits, and molecular breeding expertise. The ge...

  7. Implication of the env Gene of the Human Endogenous Retrovirus W Family in the Expression of BDNF and DRD3 and Development of Recent-Onset Schizophrenia

    PubMed Central

    Huang, WenJie; Li, Shan; Hu, YuanMing; Yu, Honglian; Luo, Feng; Zhang, Qi; Zhu, Fan

    2011-01-01

    Objective: Retrovirus has been suggested as one of agents involved in the development of schizophrenia. In the present study, we examined the role of the human endogenous retrovirus W family (HERV-W) env gene in the etiopathogenesis of recent-onset schizophrenia, using molecular and epidemiological approaches. Methods: Nested RT-PCR was used to detect the messenger RNA (mRNA) of the HERV-w env gene in plasmas. Quantitative real-time polymerase chain reaction (PCR) was employed to detect the viral reverse transcriptase activity in human sera. Human U251 glioma cells were used to study the potential role of the HERV-W env gene in the etiopathogenesis of recent-onset schizophrenia. Results: We identified genes with mRNA sequences homologous to HERV-W env gene from plasmas of 42 out of 118 individuals with recent-onset schizophrenia but not from any of 106 normal persons (P < .01, t test). Quantitative real-time PCR showed a significantly increase in the reverse transcriptase activity in the sera of patients (by 35.59%) compared with controls (by 2.83%) (P < .05, t test). Overexpression of HERV-w env in human U251 glioma cells upregulated brain-derived neurotrophic factor (BDNF), an important schizophrenia-associated gene, neurotrophic tyrosine kinase receptor type 2 (NTRK2, also called TrkB), and dopamine receptor D3 and increased the phosphorylation of cyclic adenosine monophosphate response element–binding (CREB) protein. BDNF promoter reporter gene assays showed that the HERV-W env triggers BDNF production in human U251 glioma cells. Using gene knockdown, we found that CREB is required for the expression of BDNF that is regulated by env. Conclusion: Our data revealed that the transcriptional activation of HERV is associated with the development of schizophrenia in some patients and indicated that HERV-W env regulates the expression of schizophrenia-associated genes. This report is the first to elucidate the signaling pathway responsible for the upregulation of

  8. The Integrated Airport: Building a Successful NextGen Testbed

    SciTech Connect

    Frederick-Recascino, Christina; Sweigard, Doug; Lester, Wade

    2009-02-18

    This presentation will describe a unique public-private partnership - the Integrated Airport - that was created to engage in research and testing related to NextGen Technology deployment. NextGen refers to the program that will be initiated to modernize the US National Airspace. As with any major, multi-decade initiative, such as NextGen, integration of work efforts by multiple partners in the modernization is critical for success. This talk will focus on the development of the consortium, how the consortium plans for NextGen initiatives, the series of technology demonstrations we have produced and plans for the future of NextGen testing and implementation.

  9. The environmental-data automated track annotation (Env-DATA) system: linking animal tracks with environmental data.

    PubMed

    Dodge, Somayeh; Bohrer, Gil; Weinzierl, Rolf; Davidson, Sarah C; Kays, Roland; Douglas, David; Cruz, Sebastian; Han, Jiawei; Brandes, David; Wikelski, Martin

    2013-01-01

    The movement of animals is strongly influenced by external factors in their surrounding environment such as weather, habitat types, and human land use. With advances in positioning and sensor technologies, it is now possible to capture animal locations at high spatial and temporal granularities. Likewise, scientists have an increasing access to large volumes of environmental data. Environmental data are heterogeneous in source and format, and are usually obtained at different spatiotemporal scales than movement data. Indeed, there remain scientific and technical challenges in developing linkages between the growing collections of animal movement data and the large repositories of heterogeneous remote sensing observations, as well as in the developments of new statistical and computational methods for the analysis of movement in its environmental context. These challenges include retrieval, indexing, efficient storage, data integration, and analytical techniques. This paper contributes to movement ecology research by presenting a new publicly available system, Environmental-Data Automated Track Annotation (Env-DATA), that automates annotation of movement trajectories with ambient atmospheric observations and underlying landscape information. Env-DATA provides a free and easy-to-use platform that eliminates technical difficulties of the annotation processes and relieves end users of a ton of tedious and time-consuming tasks associated with annotation, including data acquisition, data transformation and integration, resampling, and interpolation. The system is illustrated with a case study of Galapagos Albatross (Phoebastria irrorata) tracks and their relationship to wind, ocean productivity and chlorophyll concentration. Our case study illustrates why adult albatrosses make long-range trips to preferred, productive areas and how wind assistance facilitates their return flights while their outbound flights are hampered by head winds. The new Env-DATA system enhances

  10. Gen IV Materials Handbook Implementation Plan

    SciTech Connect

    Rittenhouse, P.; Ren, W.

    2005-03-29

    A Gen IV Materials Handbook is being developed to provide an authoritative single source of highly qualified structural materials information and materials properties data for use in design and analyses of all Generation IV Reactor Systems. The Handbook will be responsive to the needs expressed by all of the principal government, national laboratory, and private company stakeholders of Gen IV Reactor Systems. The Gen IV Materials Handbook Implementation Plan provided here addresses the purpose, rationale, attributes, and benefits of the Handbook and will detail its content, format, quality assurance, applicability, and access. Structural materials, both metallic and ceramic, for all Gen IV reactor types currently supported by the Department of Energy (DOE) will be included in the Gen IV Materials Handbook. However, initial emphasis will be on materials for the Very High Temperature Reactor (VHTR). Descriptive information (e.g., chemical composition and applicable technical specifications and codes) will be provided for each material along with an extensive presentation of mechanical and physical property data including consideration of temperature, irradiation, environment, etc. effects on properties. Access to the Gen IV Materials Handbook will be internet-based with appropriate levels of control. Information and data in the Handbook will be configured to allow search by material classes, specific materials, specific information or property class, specific property, data parameters, and individual data points identified with materials parameters, test conditions, and data source. Details on all of these as well as proposed applicability and consideration of data quality classes are provided in the Implementation Plan. Website development for the Handbook is divided into six phases including (1) detailed product analysis and specification, (2) simulation and design, (3) implementation and testing, (4) product release, (5) project/product evaluation, and (6) product

  11. Pharmacologic Inhibition of Nedd8 Activation Enzyme Exposes CD4-Induced Epitopes within Env on Cells Expressing HIV-1.

    PubMed

    Tokarev, Andrey; Stoneham, Charlotte; Lewinski, Mary K; Mukim, Amey; Deshmukh, Savitha; Vollbrecht, Thomas; Spina, Celsa A; Guatelli, John

    2015-12-16

    HIV-1 Vpu decreases the exposure of epitopes within the viral envelope glycoprotein (Env) on the surface of infected cells by downregulating both BST2 and CD4. To test the hypothesis that inhibiting Vpu activity would increase the exposure of these epitopes and sensitize infected cells to antibody-dependent cellular cytotoxicity (ADCC), we treated cells with the Nedd8 activation enzyme (NAE) inhibitor MLN4924, which inhibits the cullin1-based ubiquitin ligase complex coopted by Vpu to degrade cellular targets. Treatment of HeLa cells with MLN4924 or expression of a dominant negative mutant of cullin1 inhibited the Vpu-mediated downregulation of CD4 but not the downregulation of BST2. NAE inhibition also increased the surface exposure of CD4-induced epitopes within Env on HEK293 cells containing an inducible HIV genome, on infected CEM T cells, and on infected primary T cells. In contrast, the Vpu-mediated downregulation of BST2 was substantially inhibited by MLN4924 only when T cells were treated with alpha interferon (IFN-α) to induce high levels of BST2 expression. As reported previously, the absence of vpu or nef and even more so the combined absence of these two genes sensitized infected cells to ADCC. However, NAE inhibition affected ADCC minimally. Paradoxically, even in infected, IFN-treated cells in which NAE inhibition substantially rescued the surface level of BST2, the surface level of Env detected with an antibody recognizing a CD4-independent epitope (2G12) was minimally increased. Mutation of the C-terminal Vpu residue W76, which supports the ability of Vpu to stimulate virion release by displacing BST2 from assembly sites on the plasma membrane by a cullin1-independent mechanism, increased the exposure of Env detected by 2G12 on infected T cells. Thus, inhibiting the displacement function of Vpu together with its ability to degrade CD4 and BST2 may be required to sensitize infected cells to ADCC. Pathogenic viruses encode gene products that enable

  12. Yersinia pestis requires the 2-component regulatory system OmpR-EnvZ to resist innate immunity during the early and late stages of plague.

    PubMed

    Reboul, Angéline; Lemaître, Nadine; Titecat, Marie; Merchez, Maud; Deloison, Gaspard; Ricard, Isabelle; Pradel, Elizabeth; Marceau, Michaël; Sebbane, Florent

    2014-11-01

    Plague is transmitted by fleas or contaminated aerosols. To successfully produce disease, the causal agent (Yersinia pestis) must rapidly sense and respond to rapid variations in its environment. Here, we investigated the role of 2-component regulatory systems (2CSs) in plague because the latter are known to be key players in bacterial adaptation to environmental change. Along with the previously studied PhoP-PhoQ system, OmpR-EnvZ was the only one of Y. pestis' 23 other 2CSs required for production of bubonic, septicemic, and pneumonic plague. In vitro, OmpR-EnvZ was needed to counter serum complement and leukocytes but was not required for the secretion of antiphagocyte exotoxins. In vivo, Y. pestis lacking OmpR-EnvZ did not induce an early immune response in the skin and was fully virulent in neutropenic mice. We conclude that, throughout the course of Y. pestis infection, OmpR-EnvZ is required to counter toxic effectors secreted by polymorphonuclear leukocytes in the tissues. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  13. Comparison of Antibody-Dependent Cell-Mediated Cytotoxicity and Virus Neutralization by HIV-1 Env-Specific Monoclonal Antibodies

    PubMed Central

    von Bredow, Benjamin; Arias, Juan F.; Heyer, Lisa N.; Moldt, Brian; Le, Khoa; Robinson, James E.; Burton, Dennis R.

    2016-01-01

    ABSTRACT Although antibodies to the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein have been studied extensively for their ability to block viral infectivity, little data are currently available on nonneutralizing functions of these antibodies, such as their ability to eliminate virus-infected cells by antibody-dependent cell-mediated cytotoxicity (ADCC). HIV-1 Env-specific antibodies of diverse specificities, including potent broadly neutralizing and nonneutralizing antibodies, were therefore tested for ADCC against cells infected with a lab-adapted HIV-1 isolate (HIV-1NL4-3), a primary HIV-1 isolate (HIV-1JR-FL), and a simian-human immunodeficiency virus (SHIV) adapted for pathogenic infection of rhesus macaques (SHIVAD8-EO). In accordance with the sensitivity of these viruses to neutralization, HIV-1NL4-3-infected cells were considerably more sensitive to ADCC, both in terms of the number of antibodies and magnitude of responses, than cells infected with HIV-1JR-FL or SHIVAD8-EO. ADCC activity generally correlated with antibody binding to Env on the surfaces of virus-infected cells and with viral neutralization; however, neutralization was not always predictive of ADCC, as instances of ADCC in the absence of detectable neutralization, and vice versa, were observed. These results reveal incomplete overlap in the specificities of antibodies that mediate these antiviral activities and provide insights into the relationship between ADCC and neutralization important for the development of antibody-based vaccines and therapies for combating HIV-1 infection. IMPORTANCE This study provides fundamental insights into the relationship between antibody-dependent cell-mediated cytotoxicity (ADCC) and virus neutralization that may help to guide the development of antibody-based vaccines and immunotherapies for the prevention and treatment of HIV-1 infection. PMID:27122574

  14. Inhibition of HIV-1 Env-Mediated Cell-Cell Fusion by Lectins, Peptide T-20, and Neutralizing Antibodies

    PubMed Central

    Yee, Michael; Konopka, Krystyna; Balzarini, Jan; Düzgüneş, Nejat

    2011-01-01

    Background: Broadly cross-reactive, neutralizing human monoclonal antibodies, including 2F5, 2G12, 4E10 and IgG1 b12, can inhibit HIV-1 infection in vitro at very low concentrations. We examined the ability of these antibodies to inhibit cell-cell fusion between Clone69TRevEnv cells induced to express the viral envelope proteins, gp120/gp41 (Env), and highly CD4-positive SupT1 cells. The cells were loaded with green and red-orange cytoplasmic fluorophores, and fusion was monitored by fluorescence microscopy. Results: Cell-cell fusion was inhibited completely by the carbohydrate binding proteins (CBPs), Hippeastrum hybrid (Amaryllis) agglutinin (HHA), and Galanthus nivalis (Snowdrop) agglutinin (GNA), and by the peptide, T-20, at relatively low concentrations. Anti-gp120 and anti-gp41 antibodies, at concentrations much higher than those required for neutralization, were not particularly effective in inhibiting fusion. Monoclonal antibodies b12, m14 IgG and 2G12 had moderate inhibitory activity; the IC50 of 2G12 was about 80 µg/ml. Antibodies 4E10 and 2F5 had no inhibitory activity at the concentrations tested. Conclusions: These observations raise concerns about the ability of neutralizing antibodies to inhibit the spread of viral genetic material from infected cells to uninfected cells via cell-cell fusion. The interaction of gp120/gp41 with cell membrane CD4 may be different in cell-cell and virus-cell membrane fusion reactions, and may explain the differential effects of antibodies in these two systems. The fluorescence assay described here may be useful in high throughput screening of potential HIV fusion inhibitors. PMID:21660189

  15. Inhibition of HIV-1 Env-Mediated Cell-Cell Fusion by Lectins, Peptide T-20, and Neutralizing Antibodies.

    PubMed

    Yee, Michael; Konopka, Krystyna; Balzarini, Jan; Düzgüneş, Nejat

    2011-01-01

    Broadly cross-reactive, neutralizing human monoclonal antibodies, including 2F5, 2G12, 4E10 and IgG1 b12, can inhibit HIV-1 infection in vitro at very low concentrations. We examined the ability of these antibodies to inhibit cell-cell fusion between Clone69TRevEnv cells induced to express the viral envelope proteins, gp120/gp41 (Env), and highly CD4-positive SupT1 cells. The cells were loaded with green and red-orange cytoplasmic fluorophores, and fusion was monitored by fluorescence microscopy. Cell-cell fusion was inhibited completely by the carbohydrate binding proteins (CBPs), Hippeastrum hybrid (Amaryllis) agglutinin (HHA), and Galanthus nivalis (Snowdrop) agglutinin (GNA), and by the peptide, T-20, at relatively low concentrations. Anti-gp120 and anti-gp41 antibodies, at concentrations much higher than those required for neutralization, were not particularly effective in inhibiting fusion. Monoclonal antibodies b12, m14 IgG and 2G12 had moderate inhibitory activity; the IC(50) of 2G12 was about 80 µg/ml. Antibodies 4E10 and 2F5 had no inhibitory activity at the concentrations tested. These observations raise concerns about the ability of neutralizing antibodies to inhibit the spread of viral genetic material from infected cells to uninfected cells via cell-cell fusion. The interaction of gp120/gp41 with cell membrane CD4 may be different in cell-cell and virus-cell membrane fusion reactions, and may explain the differential effects of antibodies in these two systems. The fluorescence assay described here may be useful in high throughput screening of potential HIV fusion inhibitors.

  16. Comparison of Antibody-Dependent Cell-Mediated Cytotoxicity and Virus Neutralization by HIV-1 Env-Specific Monoclonal Antibodies.

    PubMed

    von Bredow, Benjamin; Arias, Juan F; Heyer, Lisa N; Moldt, Brian; Le, Khoa; Robinson, James E; Zolla-Pazner, Susan; Burton, Dennis R; Evans, David T

    2016-07-01

    Although antibodies to the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein have been studied extensively for their ability to block viral infectivity, little data are currently available on nonneutralizing functions of these antibodies, such as their ability to eliminate virus-infected cells by antibody-dependent cell-mediated cytotoxicity (ADCC). HIV-1 Env-specific antibodies of diverse specificities, including potent broadly neutralizing and nonneutralizing antibodies, were therefore tested for ADCC against cells infected with a lab-adapted HIV-1 isolate (HIV-1NL4-3), a primary HIV-1 isolate (HIV-1JR-FL), and a simian-human immunodeficiency virus (SHIV) adapted for pathogenic infection of rhesus macaques (SHIVAD8-EO). In accordance with the sensitivity of these viruses to neutralization, HIV-1NL4-3-infected cells were considerably more sensitive to ADCC, both in terms of the number of antibodies and magnitude of responses, than cells infected with HIV-1JR-FL or SHIVAD8-EO ADCC activity generally correlated with antibody binding to Env on the surfaces of virus-infected cells and with viral neutralization; however, neutralization was not always predictive of ADCC, as instances of ADCC in the absence of detectable neutralization, and vice versa, were observed. These results reveal incomplete overlap in the specificities of antibodies that mediate these antiviral activities and provide insights into the relationship between ADCC and neutralization important for the development of antibody-based vaccines and therapies for combating HIV-1 infection. This study provides fundamental insights into the relationship between antibody-dependent cell-mediated cytotoxicity (ADCC) and virus neutralization that may help to guide the development of antibody-based vaccines and immunotherapies for the prevention and treatment of HIV-1 infection. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  17. Safety Assurance in NextGen

    NASA Technical Reports Server (NTRS)

    HarrisonFleming, Cody; Spencer, Melissa; Leveson, Nancy; Wilkinson, Chris

    2012-01-01

    The generation of minimum operational, safety, performance, and interoperability requirements is an important aspect of safely integrating new NextGen components into the Communication Navigation Surveillance and Air Traffic Management (CNS/ATM) system. These requirements are used as part of the implementation and approval processes. In addition, they provide guidance to determine the levels of design assurance and performance that are needed for each element of the new NextGen procedures, including aircraft, operator, and Air Navigation and Service Provider. Using the enhanced Airborne Traffic Situational Awareness for InTrail Procedure (ATSA-ITP) as an example, this report describes some limitations of the current process used for generating safety requirements and levels of required design assurance. An alternative process is described, as well as the argument for why the alternative can generate more comprehensive requirements and greater safety assurance than the current approach.

  18. A Comparative Phase I Study of Combination, Homologous Subtype-C DNA, MVA, and Env gp140 Protein/Adjuvant HIV Vaccines in Two Immunization Regimes

    PubMed Central

    Joseph, Sarah; Quinn, Killian; Greenwood, Aldona; Cope, Alethea V.; McKay, Paul F.; Hayes, Peter J.; Kopycinski, Jakub T.; Gilmour, Jill; Miller, Aleisha N.; Geldmacher, Christof; Nadai, Yuka; Ahmed, Mohamed I. M.; Montefiori, David C.; Dally, Len; Bouliotis, George; Lewis, David J. M.; Tatoud, Roger; Wagner, Ralf; Esteban, Mariano; Shattock, Robin J.; McCormack, Sheena; Weber, Jonathan

    2017-01-01

    There remains an urgent need for a prophylactic HIV vaccine. We compared combined MVA and adjuvanted gp140 to sequential MVA/gp140 after DNA priming. We expected Env-specific CD4+ T-cells after DNA and MVA priming, and Env-binding antibodies in 100% individuals after boosting with gp140 and that combined vaccines would not compromise safety and might augment immunogenicity. Forty volunteers were primed three times with DNA plasmids encoding (CN54) env and (ZM96) gag-pol-nef at 0, 4 and 8 weeks then boosted with MVA-C (CN54 env and gag-pol-nef) and glucopyranosyl lipid adjuvant—aqueous formulation (GLA-AF) adjuvanted CN54gp140. They were randomised to receive them in combination at the same visit at 16 and 20 weeks (accelerated) or sequentially with MVA-C at 16, 20, and GLA-AF/gp140 at 24 and 28 weeks (standard). All vaccinations were intramuscular. Primary outcomes included ≥grade 3 safety events and the titer of CN54gp140-specific binding IgG. Other outcomes included neutralization, binding antibody specificity and T-cell responses. Two participants experienced asymptomatic ≥grade 3 transaminitis leading to discontinuation of vaccinations, and three had grade 3 solicited local or systemic reactions. A total of 100% made anti-CN54gp140 IgG and combining vaccines did not significantly alter the response; geometric mean titer 6424 (accelerated) and 6578 (standard); neutralization of MW965.2 Tier 1 pseudovirus was superior in the standard group (82 versus 45% responders, p = 0.04). T-cell ELISpot responses were CD4+ and Env-dominant; 85 and 82% responding in the accelerated and standard groups, respectively. Vaccine-induced IgG responses targeted multiple regions within gp120 with the V3 region most immunodominant and no differences between groups detected. Combining MVA and gp140 vaccines did not result in increased adverse events and did not significantly impact upon the titer of Env-specific binding antibodies, which were seen in 100% individuals

  19. Cocirculation of Two env Molecular Variants, of Possible Recombinant Origin, in Gorilla and Chimpanzee Simian Foamy Virus Strains from Central Africa

    PubMed Central

    Richard, Léa; Rua, Réjane; Betsem, Edouard; Mouinga-Ondémé, Augustin; Kazanji, Mirdad; Leroy, Eric; Njouom, Richard; Buseyne, Florence; Afonso, Philippe V.

    2015-01-01

    ABSTRACT Simian foamy virus (SFV) is a ubiquitous retrovirus in nonhuman primates (NHPs) that can be transmitted to humans, mostly through severe bites. In the past few years, our laboratory has identified more than 50 hunters from central Africa infected with zoonotic SFVs. Analysis of the complete sequences of five SFVs obtained from these individuals revealed that env was the most variable gene. Furthermore, recombinant SFV strains, some of which involve sequences in the env gene, were recently identified. Here, we investigated the variability of the env genes of zoonotic SFV strains and searched for possible recombinants. We sequenced the complete env gene or its surface glycoprotein region (SU) from DNA amplified from the blood of (i) a series of 40 individuals from Cameroon or Gabon infected with a gorilla or chimpanzee foamy virus (FV) strain and (ii) 1 gorilla and 3 infected chimpanzees living in the same areas as these hunters. Phylogenetic analyses revealed the existence of two env variants among both the gorilla and chimpanzee FV strains that were present in zoonotic and NHP strains. These variants differ greatly (>30% variability) in a 753-bp-long region located in the receptor-binding domain of SU, whereas the rest of the gene is very conserved. Although the organizations of the Env protein sequences are similar, the potential glycosylation patterns differ between variants. Analysis of recombination suggests that the variants emerged through recombination between different strains, although all parental strains could not be identified. IMPORTANCE SFV infection in humans is a great example of a zoonotic retroviral infection that has not spread among human populations, in contrast to human immunodeficiency viruses (HIVs) and human T-lymphotropic viruses (HTLVs). Recombination was a major mechanism leading to the emergence of HIV. Here, we show that two SFV molecular envelope gene variants circulate among ape populations in Central Africa and that both

  20. GenCade: Introduction, Background, and Formulation

    DTIC Science & Technology

    2012-03-07

    Draft, ft Wood Wood chips Flour Other US Army Corps of Engineers BUILDING STRONG® GenCade Matagorda Ship Channel, TX CMS Shark River Inlet...morphology change Habitat Change • Regional Sediment Management • Beach fills • Inlet bypassing • Channel maintenance • Unifying technology for...to SLR • Habitat evolution (Piping Plover; vegetation) Project Planning & Design 13th Annual CIRP Technology-Transfer Workshop, March 7, 2012 7

  1. JPDO Portfolio Analysis of NextGen

    DTIC Science & Technology

    2009-09-01

    consideration of environmental issues MITRE Fleet Forecast + new aircraft (Continuous Low Emissions Energy and Noise (CLEEN) MITRE Fleet Forecast...measures and monetizes the impacts of local air quality, climate change and noise levels near major airports and compares a NextGen scenario to a baseline...assumptions seem to be consistent with existing practices for defining new Area Navigation ( RNAV )/RNP and CDA procedures today. Note that this approach

  2. Single-Chain Soluble BG505.SOSIP gp140 Trimers as Structural and Antigenic Mimics of Mature Closed HIV-1 Env

    PubMed Central

    Georgiev, Ivelin S.; Joyce, M. Gordon; Yang, Yongping; Sastry, Mallika; Zhang, Baoshan; Baxa, Ulrich; Chen, Rita E.; Druz, Aliaksandr; Lees, Christopher R.; Narpala, Sandeep; Schön, Arne; Van Galen, Joseph; Chuang, Gwo-Yu; Gorman, Jason; Harned, Adam; Pancera, Marie; Stewart-Jones, Guillaume B. E.; Cheng, Cheng; Freire, Ernesto; McDermott, Adrian B.; Mascola, John R.

    2015-01-01

    ABSTRACT Similar to other type I fusion machines, the HIV-1 envelope glycoprotein (Env) requires proteolytic activation; specifically, cleavage of a gp160 precursor into gp120 and gp41 subunits creates an N-terminal gp41 fusion peptide and permits folding from an immature uncleaved state to a mature closed state. While the atomic-level consequences of cleavage for HIV-1 Env are still being determined, the uncleaved state is antigenically distinct from the mature closed state, and cleavage has been reported to be essential for mimicry of the mature viral spike by soluble versions of Env. Here we report the redesign of a current state-of-the-art soluble Env mimic, BG505.SOSIP, to make it cleavage independent. Specifically, we replaced the furin cleavage site between gp120 and gp41 with Gly-Ser linkers of various lengths. The resultant linked gp120-gp41 constructs, termed single-chain gp140 (sc-gp140), exhibited different levels of structural and antigenic mimicry of the parent cleaved BG505.SOSIP. When constructs were subjected to negative selection to remove subspecies recognized by poorly neutralizing antibodies, trimers of high antigenic mimicry of BG505.SOSIP could be obtained; negative-stain electron microscopy indicated these to resemble the mature closed state. Higher proportions of BG505.SOSIP-trimer mimicry were observed in sc-gp140s with linkers of 6 or more residues, with a linker length of 15 residues exhibiting especially promising traits. Overall, flexible linkages between gp120 and gp41 in BG505.SOSIP can thus substitute for cleavage, and sc-gp140s that closely mimicked the vaccine-preferred mature closed state of Env could be obtained. IMPORTANCE The trimeric HIV-1 envelope glycoprotein (Env) is the sole target of virus-directed neutralizing antibody responses and a primary focus of vaccine design. Soluble mimics of Env have proven challenging to obtain and have been thought to require proteolytic cleavage into two-component subunits, gp120 and gp41

  3. Single-Chain Soluble BG505.SOSIP gp140 Trimers as Structural and Antigenic Mimics of Mature Closed HIV-1 Env.

    PubMed

    Georgiev, Ivelin S; Joyce, M Gordon; Yang, Yongping; Sastry, Mallika; Zhang, Baoshan; Baxa, Ulrich; Chen, Rita E; Druz, Aliaksandr; Lees, Christopher R; Narpala, Sandeep; Schön, Arne; Van Galen, Joseph; Chuang, Gwo-Yu; Gorman, Jason; Harned, Adam; Pancera, Marie; Stewart-Jones, Guillaume B E; Cheng, Cheng; Freire, Ernesto; McDermott, Adrian B; Mascola, John R; Kwong, Peter D

    2015-05-01

    Similar to other type I fusion machines, the HIV-1 envelope glycoprotein (Env) requires proteolytic activation; specifically, cleavage of a gp160 precursor into gp120 and gp41 subunits creates an N-terminal gp41 fusion peptide and permits folding from an immature uncleaved state to a mature closed state. While the atomic-level consequences of cleavage for HIV-1 Env are still being determined, the uncleaved state is antigenically distinct from the mature closed state, and cleavage has been reported to be essential for mimicry of the mature viral spike by soluble versions of Env. Here we report the redesign of a current state-of-the-art soluble Env mimic, BG505.SOSIP, to make it cleavage independent. Specifically, we replaced the furin cleavage site between gp120 and gp41 with Gly-Ser linkers of various lengths. The resultant linked gp120-gp41 constructs, termed single-chain gp140 (sc-gp140), exhibited different levels of structural and antigenic mimicry of the parent cleaved BG505.SOSIP. When constructs were subjected to negative selection to remove subspecies recognized by poorly neutralizing antibodies, trimers of high antigenic mimicry of BG505.SOSIP could be obtained; negative-stain electron microscopy indicated these to resemble the mature closed state. Higher proportions of BG505.SOSIP-trimer mimicry were observed in sc-gp140s with linkers of 6 or more residues, with a linker length of 15 residues exhibiting especially promising traits. Overall, flexible linkages between gp120 and gp41 in BG505.SOSIP can thus substitute for cleavage, and sc-gp140s that closely mimicked the vaccine-preferred mature closed state of Env could be obtained. The trimeric HIV-1 envelope glycoprotein (Env) is the sole target of virus-directed neutralizing antibody responses and a primary focus of vaccine design. Soluble mimics of Env have proven challenging to obtain and have been thought to require proteolytic cleavage into two-component subunits, gp120 and gp41, to achieve

  4. Diagnosing ALS

    MedlinePlus

    ... that a person diagnosed with ALS seek a second opinion from an ALS "expert" - someone who diagnoses and treats many ALS patients and has training in this medical specialty. The ALS Association maintains a list of recognized experts in the field of ALS. See ALS Association Certified Centers of ...

  5. GEN3D Ver. 1.37

    SciTech Connect

    2012-01-04

    GEN3D is a three-dimensional mesh generation program. The three-dimensional mesh is generated by mapping a two-dimensional mesh into threedimensions according to one of four types of transformations: translating, rotating, mapping onto a spherical surface, and mapping onto a cylindrical surface. The generated three-dimensional mesh can then be reoriented by offsetting, reflecting about an axis, and revolving about an axis. GEN3D can be used to mesh geometries that are axisymmetric or planar, but, due to three-dimensional loading or boundary conditions, require a three-dimensional finite element mesh and analysis. More importantly, it can be used to mesh complex three-dimensional geometries composed of several sections when the sections can be defined in terms of transformations of two dimensional geometries. The code GJOIN is then used to join the separate sections into a single body. GEN3D reads and writes twodimensional and threedimensional mesh databases in the GENESIS database format; therefore, it is compatible with the preprocessing, postprocessing, and analysis codes used by the Engineering Analysis Department at Sandia National Laboratories, Albuquerque, NM.

  6. A functional interaction between gp41 and gp120 is observed for monomeric but not oligomeric, uncleaved HIV-1 Env gp140.

    PubMed

    Guttman, Miklos; Lee, Kelly K

    2013-11-01

    The envelope glycoprotein (Env) is the sole antigenic feature on the surface of HIV and the target for the humoral immune system. Soluble, uncleaved gp140 Env constructs truncated at the transmembrane domain are being investigated intensively as potential vaccine immunogens by many groups, and understanding their structural properties is essential. We used hydrogen/deuterium-exchange mass spectrometry and small-angle X-ray scattering to probe structural order in a panel of commonly used gp140 constructs and matched gp120 monomers. We observed that oligomeric forms of uncleaved gp140, generally presumed to be trimeric, contain a protease-resistant form of gp41 akin to the postfusion, helical bundle conformation and appear to lack specific interactions between gp120 and gp41. In contrast, the monomeric form of gp140 shows significant stabilization of the gp120 inner domain imparted by the gp41 region, demonstrating excellent agreement with past mutagenesis studies. Moreover, the gp140 monomers respond to CD4 binding in manner that is consistent with the initial stages of Env activation: CD4 binding induces structural ordering throughout gp120 while loosening its association with gp41. The results indicate that uncleaved gp140 oligomers do not represent an authentic prefusion form of Env, whereas gp140 monomers isolated from the same glycoprotein preparations in many ways exhibit function and internal structural order that are consistent with expectations for certain aspects of native Env. gp140 monomers may thus be a useful reagent for advancing structural and functional studies.

  7. Recombinant pro-apoptotic Mycobacterium tuberculosis generates CD8+ T cell responses against human immunodeficiency virus type 1 Env and M. tuberculosis in neonatal mice.

    PubMed

    Ranganathan, Uma Devi K; Larsen, Michelle H; Kim, John; Porcelli, Steven A; Jacobs, William R; Fennelly, Glenn J

    2009-12-10

    Mycobacterium bovis BCG is an attractive vaccine vector against breast milk HIV transmission because it elicits Th1-type responses in newborns. However, BCG causes disease in HIV-infected infants. Genetically attenuated Mycobacterium tuberculosis (Mtb) mutants represent a safer alternative for immunocompromised populations. In the current study, we compared the immunogenicity in mice of three different recombinant attenuated Mtb strains expressing an HIV envelope (Env) antigen construct. Two of these strains (DeltalysA DeltapanCD Mtb and DeltaRD1 DeltapanCD Mtb) failed to induce significant levels of HIV Env-specific CD8(+) T cell responses. In striking contrast, an HIV-1 Env-expressing attenuated DeltalysA Mtb containing a deletion in secA2, which encodes a virulence-related secretion system involved in evading adaptive immunity, generated consistently measurable Env-specific CD8(+) T cell responses that were significantly greater than those observed after immunization with BCG expressing HIV Env. Similarly, another strain of DeltalysA DeltasecA2 Mtb expressing SIV Gag induced Gag- and Mtb-specific CD8(+) T cells producing perforin or IFNgamma, and Gag-specific CD4(+) T cells producing IFNgamma within 3 weeks after immunization in adult mice; in addition, IFNgamma-producing Gag-specific CD8(+) T cells and Mtb-specific CD4(+) T cells were observed in neonatal mice within 1 week of immunization. We conclude that DeltalysA DeltasecA2 Mtb is a promising vaccine platform to construct a safe combination HIV-TB vaccine for use in neonates.

  8. Downregulation of Human Endogenous Retrovirus Type K (HERV-K) Viral env RNA in Pancreatic Cancer Cells Decreases Cell Proliferation and Tumor Growth.

    PubMed

    Li, Ming; Radvanyi, Laszlo; Yin, Bingnan; Li, Jia; Chivukula, Raghavender; Lin, Kevin; Lu, Yue; Shen, JianJun; Chang, David Z; Li, Donghui; Johanning, Gary L; Wang-Johanning, Feng

    2017-10-01

    Purpose: We investigated the role of the human endogenous retrovirus type K (HERV-K) envelope (env) gene in pancreatic cancer.Experimental Design: shRNA was employed to knockdown (KD) the expression of HERV-K in pancreatic cancer cells.Results: HERV-K env expression was detected in seven pancreatic cancer cell lines and in 80% of pancreatic cancer patient biopsies, but not in two normal pancreatic cell lines or uninvolved normal tissues. A new HERV-K splice variant was discovered in several pancreatic cancer cell lines. Reverse transcriptase activity and virus-like particles were observed in culture media supernatant obtained from Panc-1 and Panc-2 cells. HERV-K viral RNA levels and anti-HERV-K antibody titers were significantly higher in pancreatic cancer patient sera (N = 106) than in normal donor sera (N = 40). Importantly, the in vitro and in vivo growth rates of three pancreatic cancer cell lines were significantly reduced after HERV-K KD by shRNA targeting HERV-K env, and there was reduced metastasis to lung after treatment. RNA-Seq results revealed changes in gene expression after HERV-K env KD, including RAS and TP53. Furthermore, downregulation of HERV-K Env protein expression by shRNA also resulted in decreased expression of RAS, p-ERK, p-RSK, and p-AKT in several pancreatic cancer cells or tumors.Conclusions: These results demonstrate that HERV-K influences signal transduction via the RAS-ERK-RSK pathway in pancreatic cancer. Our data highlight the potentially important role of HERV-K in tumorigenesis and progression of pancreatic cancer, and indicate that HERV-K viral proteins may be attractive biomarkers and/or tumor-associated antigens, as well as potentially useful targets for detection, diagnosis, and immunotherapy of pancreatic cancer. Clin Cancer Res; 23(19); 5892-911. ©2017 AACR. ©2017 American Association for Cancer Research.

  9. HIV-1 Receptor Binding Site-Directed Antibodies Using a VH1-2 Gene Segment Orthologue Are Activated by Env Trimer Immunization

    PubMed Central

    Bale, Shridhar; Phad, Ganesh E.; Guenaga, Javier; Wilson, Richard; Soldemo, Martina; McKee, Krisha; Sundling, Christopher; Mascola, John; Li, Yuxing; Wyatt, Richard T.; Karlsson Hedestam, Gunilla B.

    2014-01-01

    Broadly neutralizing antibodies (bNAbs) isolated from chronically HIV-1 infected individuals reveal important information regarding how antibodies target conserved determinants of the envelope glycoprotein (Env) spike such as the primary receptor CD4 binding site (CD4bs). Many CD4bs-directed bNAbs use the same heavy (H) chain variable (V) gene segment, VH1-2*02, suggesting that activation of B cells expressing this allele is linked to the generation of this type of Ab. Here, we identify the rhesus macaque VH1.23 gene segment to be the closest macaque orthologue to the human VH1-2 gene segment, with 92% homology to VH1-2*02. Of the three amino acids in the VH1-2*02 gene segment that define a motif for VRC01-like antibodies (W50, N58, flanking the HCDR2 region, and R71), the two identified macaque VH1.23 alleles described here encode two. We demonstrate that immunization with soluble Env trimers induced CD4bs-specific VH1.23-using Abs with restricted neutralization breadth. Through alanine scanning and structural studies of one such monoclonal Ab (MAb), GE356, we demonstrate that all three HCDRs are involved in neutralization. This contrasts to the highly potent CD4bs-directed VRC01 class of bNAb, which bind Env predominantly through the HCDR2. Also unlike VRC01, GE356 was minimally modified by somatic hypermutation, its light (L) chain CDRs were of average lengths and it displayed a binding footprint proximal to the trimer axis. These results illustrate that the Env trimer immunogen used here activates B cells encoding a VH1-2 gene segment orthologue, but that the resulting Abs interact distinctly differently with the HIV-1 Env spike compared to VRC01. PMID:25166308

  10. BLyS-mediated modulation of naïve B cell subsets impacts HIV Env-induced antibody responses1

    PubMed Central

    Dosenovic, Pia; Soldemo, Martina; Scholz, Jean L.; O’Dell, Sijy; Grasset, Emilie K.; Pelletier, Nadège; Karlsson, Mikael C. I.; Mascola, John R.; Wyatt, Richard T.; Cancro, Michael P.; Karlsson Hedestam, Gunilla B.

    2012-01-01

    Neutralizing Abs provide the protective effect of the majority of existing human vaccines. For a prophylactic vaccine against HIV-1, broadly neutralizing Abs (bNAbs) targeting conserved epitopes of the viral envelope glycoproteins (Env) are likely required, as the pool of circulating HIV-1 variants is extremely diverse. The failure to efficiently induce bNAbs by vaccination may be due to the use of sub-optimal immunogens or immunization regimens, or it may indicate that B cells specific for broadly neutralizing Env determinants are selected against during peripheral checkpoints, either before or after antigen encounter. To investigate if perturbation of B cell subsets prior to immunization with recombinant Env protein affects the vaccine-induced Ab response in mice, we used B Lymphocyte Stimulator (BLyS), a cytokine that regulates survival and selection of peripheral B cells. We show that the transient BLyS treatment used here substantially affected naïve B cell populations; in particular, it resulted in an increased number of B cells surviving counter-selection at the transitional stages. We also observed an increased number of mature naïve B cells, especially marginal zone B cells, in BLyS-treated mice. Intriguingly, provision of excess BLyS prior to immunization led to a consistent improvement in the frequency and potency of HIV-1 Env vaccine-induced neutralizing Ab responses, without increasing the number of Env-specific Ab-secreting cells or the Ab binding titers measured after boosting. The results presented here suggest that an increased understanding of BLyS-regulated processes may help the design of vaccine regimens aimed at eliciting improved neutralizing Ab responses against HIV-1. PMID:22561155

  11. Structures and stability of metal-doped GenM (n = 9, 10) clusters

    NASA Astrophysics Data System (ADS)

    Qin, Wei; Lu, Wen-Cai; Xia, Lin-Hua; Zhao, Li-Zhen; Zang, Qing-Jun; Wang, C. Z.; Ho, K. M.

    2015-06-01

    The lowest-energy structures of neutral and cationic GenM (n = 9, 10; M = Si, Li, Mg, Al, Fe, Mn, Pb, Au, Ag, Yb, Pm and Dy) clusters were studied by genetic algorithm (GA) and first-principles calculations. The calculation results show that doping of the metal atoms and Si into Ge9 and Ge10 clusters is energetically favorable. Most of the metal-doped Ge cluster structures can be viewed as adding or substituting metal atom on the surface of the corresponding ground-state Gen clusters. However, the neutral and cationic FeGe9,10,MnGe9,10 and Ge10Al are cage-like with the metal atom encapsulated inside. Such cage-like transition metal doped Gen clusters are shown to have higher adsorption energy and thermal stability. Our calculation results suggest that Ge9,10Fe and Ge9Si would be used as building blocks in cluster-assembled nanomaterials because of their high stabilities.

  12. Generic classification of the Archiborborinae (Diptera: Sphaeroceridae), with a revision of Antrops Enderlein, Coloantrops gen. nov., Maculantrops gen. nov., Photoantrops gen. nov., and Poecilantrops gen. nov.

    PubMed

    Kits, Joel H; Marshall, Stephen A

    2013-01-01

    The Archiborborinae comprise a diverse clade of flies in the family Sphaeroceridae. We here revise the generic classification, redefining the genus Antrops Enderlein and naming 5 new genera: Boreantrops gen. nov., Coloantrops gen. nov., Maculantrops gen. nov., Photoantrops gen. nov., and Poecilantrops gen. nov. The genus Archiborborus, until recently a paraphyletic assemblage including most of the described species in the subfamily, is treated as a junior synonym of Antrops (syn. nov.) We revise the genera Antrops (53 species, including 40 sp. nov.: Antrops anovariegatus, Antrops aurantifemur, Antrops baeza, Antrops bellavista, Antrops biflavus, Antrops bucki, Antrops carpishensis, Antrops cochabamba, Antrops cochinoca, Antrops coniobaptos, Antrops coroico, Antrops cotopaxi, Antrops didactylos, Antrops diversipennis, Antrops eurus, Antrops fulgiceps, Antrops fuliginosus, Antrops guandera, Antrops guaramacalensis, Antrops inca, Antrops juninensis, Antrops mucarensis, Antrops niger, Antrops papallacta, Antrops pecki, Antrops podocarpus, Antrops quadrilobus, Antrops siberia, Antrops sierrazulensis, Antrops tachira, Antrops tequendama, Antrops tetrastichus, Antrops tumbrensis, Antrops unduavi, Antrops variegatus, Antrops versabilis, Antrops vittatus, Antrops yungas, and Antrops zongo and the following comb. nov.: Antrops annulatus (Richards), Antrops chaetosus (Richards), Antrops femoralis (Blanchard), Antrops hirtus (Bigot), Antrops maculipennis (Duda), Antrops maximus (Richards), Antrops microphthalmus (Richards), Antrops quadrinotus (Bigot), Antrops setosus (Duda), Antrops simplicimanus (Richards), Antrops nitidicollis (Becker), and Antrops orbitalis (Duda)), Coloantrops (1 species: Coloantrops daedalus, sp. nov.), Maculantrops (2 species, Maculantrops hirtipes (Macquart) comb. nov. and Maculantrops altiplanus, sp. nov.), Photoantrops (1 species: Pho-toantrops echinus sp. nov.), and Poecilantrops (10 species: Poecilantrops baorucensis, Poecilantrops boraceiensis

  13. An Gen2 Based Security Authentication Protocol for RFID System

    NASA Astrophysics Data System (ADS)

    Yi, Xiaoluo; Wang, Liangmin; Mao, Dongmei; Zhan, Yongzhao

    EPC Class-1 Generation-2 specification(Gen2 in brief) has been accepted as the standard for RFID tags under grant number ISO18000-6C. However, Gen2 does not pay due attention to security. For this reason, a Gen2 based security authentication protocol is developed in this paper. In details, we study the security requirements presented in the current Gen2 based RFID authentication protocols[7-13]. Then we point out the security flaws of Chien's mutual authentication protocol[7], and improve the protocol based on a 11 security requirements. Our improved protocol merely uses CRC and PRNG operations supported by Gen2 and meets the 11 security requirements. In contrast to the similar work [14,15] on Chien's protocol or other Gen2 based schemes, our protocol is more secure and our security analysis is much more comprehensive and qualitative.

  14. Erythropoietin receptor (EpoR)-dependent mitogenicity of spleen focus-forming virus correlates with viral pathogenicity and processing of env protein but not with formation of gp52-EpoR complexes in the endoplasmic reticulum.

    PubMed Central

    Wang, Y; Kayman, S C; Li, J P; Pinter, A

    1993-01-01

    Recent evidence suggests that interactions between spleen focus-forming virus (SFFV) env products and the erythropoietin receptor (EpoR) are responsible for viral pathogenicity. Infection of factor-dependent cell lines expressing epoR (the cloned gene for EpoR) with SFFVP is mitogenic, generating cell lines that are no longer dependent on added growth factor, and an immunoprecipitable complex between EpoR and immature env protein in the endoplasmic reticulum has been identified. The dependence of these in vitro activities on env protein processing and their relationship to pathogenicity of SFFV were explored by using glycosylation site mutants of SFFV env. Mutants carrying Asn-->Asp mutations at each of the two consensus signals for N-linked glycosylation in the N-terminal domain of SFFVAP-L env (gs1 and gs2), the gs1-2- double mutant, and the gs0 quadruple mutant (mutated at all four signals utilized for N-linked glycosylation in SFFVAP-L env) were made. The primary translation products (gp52) of single-site mutant envs were processed into more highly glycosylated forms, and the corresponding viruses induced splenomegaly in susceptible mice, whereas the gs1-2- and gs0 proteins were not processed, and these viruses were not pathogenic. Unprocessed env proteins of both pathogenic and nonpathogenic mutants coprecipitated with EpoR. In the BaF3 cell assay for epoR-dependent mitogenicity, the pathogenic single mutants induced factor-independent growth efficiently whereas the nonpathogenic gs1-2- and gs0 mutants did not. These data demonstrate that the ability of gp52 to form complexes with EpoR in the endoplasmic reticulum is not sufficient for either mitogenicity in cell culture or induction of splenomegaly in mice while supporting the hypothesis that pathogenicity and mitogenicity of SFFV both result from an interaction between EpoR and SFFV env protein. Images PMID:8437218

  15. TidGen Power System Commercialization Project

    SciTech Connect

    Sauer, Christopher R.; McEntee, Jarlath

    2013-12-30

    ORPC Maine, LLC, a wholly-owned subsidiary of Ocean Renewable Power Company, LLC (collectively ORPC), submits this Final Technical Report for the TidGen® Power System Commercialization Project (Project), partially funded by the U.S. Department of Energy (DE-EE0003647). The Project was built and operated in compliance with the Federal Energy Regulatory Commission (FERC) pilot project license (P-12711) and other permits and approvals needed for the Project. This report documents the methodologies, activities and results of the various phases of the Project, including design, engineering, procurement, assembly, installation, operation, licensing, environmental monitoring, retrieval, maintenance and repair. The Project represents a significant achievement for the renewable energy portfolio of the U.S. in general, and for the U.S. marine hydrokinetic (MHK) industry in particular. The stated Project goal was to advance, demonstrate and accelerate deployment and commercialization of ORPC’s tidal-current based hydrokinetic power generation system, including the energy extraction and conversion technology, associated power electronics, and interconnection equipment capable of reliably delivering electricity to the domestic power grid. ORPC achieved this goal by designing, building and operating the TidGen® Power System in 2012 and becoming the first federally licensed hydrokinetic tidal energy project to deliver electricity to a power grid under a power purchase agreement in North America. Located in Cobscook Bay between Eastport and Lubec, Maine, the TidGen® Power System was connected to the Bangor Hydro Electric utility grid at an on-shore station in North Lubec on September 13, 2012. ORPC obtained a FERC pilot project license for the Project on February 12, 2012 and the first Maine Department of Environmental Protection General Permit issued for a tidal energy project on January 31, 2012. In addition, ORPC entered into a 20-year agreement with Bangor Hydro Electric

  16. ALS Association

    MedlinePlus

    ... toward a world without ALS! Walk to Defeat ALS® Walk to Defeat ALS® draws people of all ... We need your help. I Will Advocate National ALS Registry The National ALS Registry is a congressionally ...

  17. Gen IV Materials Handbook Functionalities and Operation

    SciTech Connect

    Ren, Weiju

    2009-12-01

    This document is prepared for navigation and operation of the Gen IV Materials Handbook, with architecture description and new user access initiation instructions. Development rationale and history of the Handbook is summarized. The major development aspects, architecture, and design principles of the Handbook are briefly introduced to provide an overview of its past evolution and future prospects. Detailed instructions are given with examples for navigating the constructed Handbook components and using the main functionalities. Procedures are provided in a step-by-step fashion for Data Upload Managers to upload reports and data files, as well as for new users to initiate Handbook access.

  18. Back to the FutureGen?

    SciTech Connect

    Buchsbaum, L.

    2009-04-15

    After years of political wrangling, Democrats may green-light the experimental clean coal power plants. The article relates how the project came to be curtailed, how Senator Dick Durbin managed to protect $134 million in funding for FutureGen in Mattoon, and how once Obama was in office a $2 billion line item to fund a 'near zero emissions power plant(s)' was placed in the Senate version of the Stimulus Bill. The final version of the legislation cut the funding to $1 billion for 'fossil energy research and development'. In December 2008 the FutureGen Alliance and the City of Mattoon spent $6.5 billion to purchase the plants eventual 440 acre site. A report by the Government Accountability Office (GAO) said that Bush's inaction may have set back clean coal technology in the US by as much as a decade. If additional funding comes through construction of the plant could start in 2010. 1 fig., 1 photo.

  19. Antibodies against Gag are diagnostic markers for feline foamy virus infections while Env and Bet reactivity is undetectable in a substantial fraction of infected cats.

    PubMed

    Romen, Fabian; Pawlita, Michael; Sehr, Peter; Bachmann, Silke; Schröder, Johannes; Lutz, Hans; Löchelt, Martin

    2006-02-20

    Spumaretroviruses or foamy viruses constitute a distinct subfamily of retroviruses. The biology of foamy viruses within the authentic host, their mode of transmission, and disease potential in the authentic host or after zoonotic transmission into human or other species are almost unknown. Using feline foamy virus (FFV) as model system, we established modular enzyme-linked immunosorbent assays (ELISA) suited to determine feline IgG and IgM antibody responses against structural and non-structural FFV proteins. We validated the ELISAs with standard reference sera. In 99 cats admitted to a Swiss veterinary hospital, overall FFV Gag antibody prevalence was 36%, reactivity against Env and the non-structural protein Bet each was about 25%, and 19% of the sera were directed against all three FFV antigens. With one exception, all Bet- and/or Env-positive sera were also positive for Gag. In this small epidemiological pilot study, FFV antibodies were not significantly associated with clinical disease.

  20. Level of education and multiple sclerosis risk after adjustment for known risk factors: The EnvIMS study.

    PubMed

    Bjørnevik, Kjetil; Riise, Trond; Cortese, Marianna; Holmøy, Trygve; Kampman, Margitta T; Magalhaes, Sandra; Myhr, Kjell-Morten; Wolfson, Christina; Pugliatti, Maura

    2016-01-01

    Several recent studies have found a higher risk of multiple sclerosis (MS) among people with a low level of education. This has been suggested to reflect an effect of smoking and lower vitamin D status in the social class associated with lower levels of education. The objective of this paper is to investigate the association between level of education and MS risk adjusting for the known risk factors smoking, infectious mononucleosis, indicators of vitamin D levels and body size. Within the case-control study on Environmental Factors In MS (EnvIMS), 953 MS patients and 1717 healthy controls from Norway reported educational level and history of exposure to putative environmental risk factors. Higher level of education were associated with decreased MS risk (p trend = 0.001) with an OR of 0.53 (95% CI 0.41-0.68) when comparing those with the highest and lowest level of education. This association was only moderately reduced after adjusting for known risk factors (OR 0.61, 95% CI 0.44-0.83). The estimates remained similar when cases with disease onset before age 28 were excluded. These findings suggest that factors related to lower socioeconomic status other than established risk factors are associated with MS risk. © The Author(s), 2015.

  1. The Environmental-Data Automated Track Annotation (Env-DATA) System: Linking Animal Tracks with Environmental Data

    NASA Astrophysics Data System (ADS)

    Bohrer, G.; Dodge, S.; Weinzierl, R.; Davidson, S. C.; Kays, R.; Douglas, D. C.; Brandes, D.; Bildstein, K.; Wikelski, M.

    2013-12-01

    The movement of animals is strongly influenced by external factors in their surrounding environment such as weather, habitat types, and human land use. With the advances in positioning and sensor technologies, it is now possible to capture data of animal locations at high spatial and temporal granularities. Likewise, modern technology provides us with an increasing access to large volumes of environmental data, some of which changes on an hourly basis. Although there have been strong developments in computational methods for the analysis of movement in its environmental context, there remain challenges in efficiently linking the spatiotemporal locations of animals with the appropriate environmental conditions along their trajectories. To this end, our new Environmental-Data Automated Track Annotation (Env-DATA) system enhances Movebank, an open portal of animal tracking data, by automating access to environmental variables from global remote sensing, weather, and ecosystem products. The system automates the download and decryption of the data from open web resources of remote sensing and weather data, and provides several interpolation methods from the native grid resolution and structure to a global regular grid linked with the movement tracks in space and time. The system is open and free to any user with movement data. The aim is to facilitate new understanding and predictive capabilities of spatiotemporal patterns of animal movement in response to dynamic and changing environments from local to global scales. The system is illustrated with a series of case studies of pan-American migrations of turkey vultures, and foraging flights of Galapagos Albatross.

  2. Level of education and multiple sclerosis risk after adjustment for known risk factors: The EnvIMS study

    PubMed Central

    Bjørnevik, Kjetil; Riise, Trond; Cortese, Marianna; Holmøy, Trygve; Kampman, Margitta T; Magalhaes, Sandra; Myhr, Kjell-Morten; Wolfson, Christina; Pugliatti, Maura

    2016-01-01

    Background: Several recent studies have found a higher risk of multiple sclerosis (MS) among people with a low level of education. This has been suggested to reflect an effect of smoking and lower vitamin D status in the social class associated with lower levels of education. Objective: The objective of this paper is to investigate the association between level of education and MS risk adjusting for the known risk factors smoking, infectious mononucleosis, indicators of vitamin D levels and body size. Methods: Within the case-control study on Environmental Factors In MS (EnvIMS), 953 MS patients and 1717 healthy controls from Norway reported educational level and history of exposure to putative environmental risk factors. Results: Higher level of education were associated with decreased MS risk (p trend = 0.001) with an OR of 0.53 (95% CI 0.41–0.68) when comparing those with the highest and lowest level of education. This association was only moderately reduced after adjusting for known risk factors (OR 0.61, 95% CI 0.44–0.83). The estimates remained similar when cases with disease onset before age 28 were excluded. Conclusion: These findings suggest that factors related to lower socioeconomic status other than established risk factors are associated with MS risk. PMID:26014605

  3. The 17 Nucleotides Downstream from the env Gene Stop Codon Are Important for Murine Leukemia Virus Packaging

    PubMed Central

    Shin Yu, Seung; Kim, Jong-Mook; Kim, Sunyoung

    2000-01-01

    We have identified a previously unknown nucleotide sequence important for the packaging of murine leukemia virus. This nucleotide sequence is located downstream from the stop codon of the env gene but does not overlap the polypurine tract. Deletion of 17 bp from this region resulted in a more than 10-fold decrease in viral titer. Consistent with this result, the deletion mutant showed a 20- to 30-fold drop in the amount of virion RNA in the culture supernatant. The total amount of virion protein in the culture supernatant was comparable for the deletion mutant and the parental virus, suggesting that the mutant construct could release the empty viral particles. These results suggested that the packaging signal sequence might be present at the two extreme sites of the viral genome, one in the region around the splice donor sequence downstream from the 5′ long terminal repeat (LTR) and the other immediately upstream from the 3′ LTR. Implications for gene therapy, especially in regard to construction of retroviral vectors and packaging constructs, are discussed. PMID:10954583

  4. Timing of use of cod liver oil, a vitamin D source, and multiple sclerosis risk: The EnvIMS study.

    PubMed

    Cortese, Marianna; Riise, Trond; Bjørnevik, Kjetil; Holmøy, Trygve; Kampman, Margitta T; Magalhaes, Sandra; Pugliatti, Maura; Wolfson, Christina; Myhr, Kjell-Morten

    2015-12-01

    Low vitamin D levels have been associated with an increased risk of multiple sclerosis (MS), although it remains unknown whether this relationship varies by age. The objective of this paper is to investigate the association between vitamin D3 supplementation through cod liver oil at different postnatal ages and MS risk. In the Norwegian component of the multinational case-control study Environmental Factors In Multiple Sclerosis (EnvIMS), a total of 953 MS patients with maximum disease duration of 10 years and 1717 controls reported their cod liver oil use from childhood to adulthood. Self-reported supplement use at ages 13-18 was associated with a reduced risk of MS (OR 0.67, 95% CI 0.52-0.86), whereas supplementation during childhood was not found to alter MS risk (OR 1.01, 95% CI 0.81-1.26), each compared to non-use during the respective period. An inverse association was found between MS risk and the dose of cod liver oil during adolescence, suggesting a dose-response relationship (p trend = 0.001) with the strongest effect for an estimated vitamin D3 intake of 600-800 IU/d (OR 0.46, 95% CI 0.31-0.70). These findings not only support the hypothesis relating to low vitamin D as a risk factor for MS, but further point to adolescence as an important susceptibility period for adult-onset MS. © The Author(s), 2015.

  5. Identification of the hASCT2-binding domain of the Env ERVWE1/syncytin-1 fusogenic glycoprotein

    PubMed Central

    Cheynet, Valérie; Oriol, Guy; Mallet, François

    2006-01-01

    The cellular HERV-W envelope/syncytin-1 protein, encoded by the envelope gene of the ERVWE1 proviral locus is a fusogenic glycoprotein probably involved in the formation of the placental syncytiotrophoblast layer. Syncytin-1-induced in vitro cell-cell fusion is dependent on the interaction with hASCT2. As no receptor binding domain has been clearly defined in the SU of neither the HERV-W Env nor the retroviruses of the same interference group, we designed an in vitro binding assay to evaluate the interaction of the HERV-W envelope with the hASCT2 receptor. Using truncated HERV-W SU subunits, a region consisting of the N-terminal 124 amino acids of the mature SU glycoprotein was determined as the minimal receptor-binding domain. This domain contains several sub-domains which are poorly conserved among retroviruses of this interference group but a region of 18 residus containing the SDGGGX2DX2R conserved motif was proved to be essential for syncytin-1-hASCT2 interaction. PMID:16820059

  6. env Sequences of Simian Immunodeficiency Viruses from Chimpanzees in Cameroon Are Strongly Related to Those of Human Immunodeficiency Virus Group N from the Same Geographic Area

    PubMed Central

    Corbet, Sylvie; Müller-Trutwin, Michaela C.; Versmisse, Pierre; Delarue, Severine; Ayouba, Ahidjo; Lewis, John; Brunak, Soren; Martin, Paul; Brun-Vezinet, Françoise; Simon, François; Barre-Sinoussi, Françoise; Mauclere, Philippe

    2000-01-01

    Human immunodeficiency virus type 1 (HIV-1) group N from Cameroon is phylogenetically close, in env, to the simian immunodeficiency virus (SIV) cpz-gab from Gabon and SIVcpz-US of unknown geographic origin. We screened 29 wild-born Cameroonian chimpanzees and found that three (Cam3, Cam4, and Cam5) were positive for HIV-1 by Western blotting. Mitochondrial DNA sequence analysis demonstrated that Cam3 and Cam5 belonged to Pan troglodytes troglodytes and that Cam4 belonged to P. t. vellerosus. Genetic analyses of the viruses together with serological data demonstrated that at least one of the two P. t. troglodytes chimpanzees (Cam5) was infected in the wild, and revealed a horizontal transmission between Cam3 and Cam4. These data confirm that P. t. troglodytes is a natural host for HIV-1-related viruses. Furthermore, they show that SIVcpz can be transmitted in captivity, from one chimpanzee subspecies to another. All three SIVcpz-cam viruses clustered with HIV-1 N in env. The full Cam3 SIVcpz genome sequence showed a very close phylogenetic relationship with SIVcpz-US, a virus identified in a P. t. troglodytes chimpanzee captured nearly 40 years earlier. Like SIVcpz-US, SIVcpz-cam3 was closely related to HIV-1 N in env, but not in pol, supporting the hypothesis that HIV-1 N results from a recombination event. SIVcpz from chimpanzees born in the wild in Cameroon are thus strongly related in env to HIV-1 N from Cameroon, demonstrating the geographic coincidence of these human and simian viruses and providing a further strong argument in favor of the origin of HIV-1 being in chimpanzees. PMID:10590144

  7. Towards an International Culture: Gen Y Students and SNS?

    ERIC Educational Resources Information Center

    Lichy, Jessica

    2012-01-01

    This article reports the findings of a small-scale investigation into the Internet user behaviour of generation Y (Gen Y) students, with particular reference to social networking sites. The study adds to the literature on cross-cultural Internet user behaviour with specific reference to Gen Y and social networking. It compares how a cohort of…

  8. Towards an International Culture: Gen Y Students and SNS?

    ERIC Educational Resources Information Center

    Lichy, Jessica

    2012-01-01

    This article reports the findings of a small-scale investigation into the Internet user behaviour of generation Y (Gen Y) students, with particular reference to social networking sites. The study adds to the literature on cross-cultural Internet user behaviour with specific reference to Gen Y and social networking. It compares how a cohort of…

  9. The EnvIMS Study: Design and Methodology of an International Case-Control Study of Environmental Risk Factors in Multiple Sclerosis.

    PubMed

    Magalhaes, Sandra; Pugliatti, Maura; Casetta, Ilaria; Drulovic, Jelena; Granieri, Enrico; Holmøy, Trygve; Kampman, Margitta T; Landtblom, Anne-Marie; Lauer, Klaus; Myhr, Kjell-Morten; Parpinel, Maria; Pekmezovic, Tatjana; Riise, Trond; Wolfson, David; Zhu, Bin; Wolfson, Christina

    2015-01-01

    Multiple sclerosis (MS) is a chronic disease of the central nervous system, often resulting in significant neurological disability. The causes of MS are not known; however, the incidence of MS is increasing, thereby suggesting that changes in lifestyle and/or environmental factors may be responsible. On this background, the Environmental Risk Factors in MS Study or EnvIMS study was designed to further explore the etiology of MS. The design and methodology are described, providing details to enable investigators to (i) use our experiences to design their own studies; (ii) take advantage of, and build on the methodological work completed for, the EnvIMS study; (iii) become aware of this data source that is available for use by the research community. EnvIMS is a multinational case-control study, enrolling 2,800 cases with MS and 5,012 population-based controls in Canada, Italy, Norway, Serbia and Sweden. The study was designed to investigate the most commonly implicated risk factors for MS etiology using a self-report questionnaire. The use of a common methodology to study MS etiology across several countries enhances the comparability of results in different geographic regions and research settings, reduces the resources required for study design and enhances the opportunity for data harmonization. © 2015 S. Karger AG, Basel.

  10. Independent variation and positive selection in env V1 and V2 domains within maternal-infant strains of human immunodeficiency virus type 1 in vivo.

    PubMed Central

    Lamers, S L; Sleasman, J W; She, J X; Barrie, K A; Pomeroy, S M; Barrett, D J; Goodenow, M M

    1993-01-01

    Multiple targets for immune recognition and cellular tropism are localized to the V1 and V2 hypervariable regions in the amino portion of human immunodeficiency virus type 1 (HIV-1) gp120env. We have assessed genetic diversity in env V1 and V2 hypervariable domains in vivo within epidemiologically related strains of HIV-1. Our strategy was to analyze longitudinal samples from two seropositive mothers and multiple children infected by perinatal transmission. Although the V1 and V2 domains are closely linked in the HIV-1 genome, nucleotide sequences in V1 and in V2 evolved independently in maternal-infant viruses in vivo. A high proportion of the nucleotide substitutions would introduce amino acid diversity in V1 and in V2. A significant excess of nonsynonymous over synonymous substitutions was identified in HIV-1 env V1 and V2 peptides in the mothers and in two older children but was not generally apparent in HIV-1 sequences in infants. An excess of nonsynonymous over synonymous substitutions indicated that there is positive selection for independent genetic variation in the V1 and V2 domains in vivo. It is likely that there are host responses to complex determinants in the V1 or V2 hypervariable domain of HIV-1 gp120. PMID:8510212

  11. Construction of infectious feline foamy virus genomes: cat antisera do not cross-neutralize feline foamy virus chimera with serotype-specific Env sequences.

    PubMed

    Zemba, M; Alke, A; Bodem, J; Winkler, I G; Flower, R L; Pfrepper, K; Delius, H; Flügel, R M; Löchelt, M

    2000-01-05

    Full-length genomes of the feline foamy virus (FFV or FeFV) isolate FUV were constructed. DNA clone pFeFV-7 stably directed the expression of infectious FFV progeny virus indistinguishable from wild-type, uncloned FFV isolate FUV. The env and bel 1 genes of pFeFV-7 were substituted for by corresponding sequences of the FFV serotype 951 since previous studies implicated a defined part of FFV Env protein as responsible for serotype-specific differences in serum neutralization (I. G. Winkler, R. M. Flügel, M. Löchelt, and R. L. P. Flower, 1998. Virology 247: 144-151). Recombinant virus derived from chimeric plasmid pFeFV-7/951 containing the hybrid env gene and the parental clone pFeFV-7 were used for neutralization studies. By means of a rapid titration assay for FFV infectivity, we show that progeny virus derived from plasmid pFeFV-7 was neutralized by FUV- but not by 951-specific antisera, whereas pFeFV-7/951-derived chimeric virus was neutralized by 951-specific antisera only. Both recombinant proviruses will be useful for repeated delivery of foreign genes for therapeutic gene applications into cats.

  12. Description of Quercorhabditis rajouriensis gen. n., sp. n. (Nematoda: Rhabditidae).

    PubMed

    Shah, Ali Asghar; Hussain, Abid; Vaid, Shavish

    2013-01-01

    Quercorhabditis rajouriensis gen. n., sp. n. is described and illustrated. Distinctive characters include a labial region set off by a constriction and wider than adjoining body, heavily sclerotized cheilostom with arched rhabdions, barely differentiable stegostom, amphidelphic gonads, spicules with a free dorsal arm, and a leptoderan bursa with ten pairs of bursal papillae. The new genus resembles Diploscapteriodes Rahm, 1928 in the shape of cheilostom, presence of amphidelphic gonads, leptoderan bursa and long conoid tail. However, it can be differentiated from the latter genus in the shape of labial region, cheilostomal sclerotization, absence of ridge-like tooth in gymnostom, having spicules with a free dorsal arm, somewhat boat-shaped gubernaculum and in having ten pairs of bursal papillae. The new genus also resembles Curviditis (Dougherty, 1953) Andrássy, 1983, Rhabditella (Cobb, 1929) Chitwood, 1933 and Metarhabditis Tahseen et. al., 2004 in having spicules with free dorsal arm. However, it can be differentiated from these related genera by its heavily cuticularised cheilostom with arched rhabdions and barely differentiable stegostom. The new genus can further be differentiated from Curviditis and Rhabditella in having a well developed bursa and from Metarhabditis by its leptoderan bursa with ten pairs of bursal papillae arranged in 3+4+3 arrangement.

  13. The Integrated Airport: Building a Successful NextGen Testbed

    ScienceCinema

    Frederick-Recascino, Christina [Embry-Riddle Aeronautical University, Daytona Beach, Florida, United States; Sweigard, Doug [Lockheed Martin Corporation; Lester, Wade [ERAU

    2016-07-12

    This presentation will describe a unique public-private partnership - the Integrated Airport - that was created to engage in research and testing related to NextGen Technology deployment.  NextGen refers to the program that will be initiated to modernize the US National Airspace.  As with any major, multi-decade initiative, such as NextGen, integration of work efforts by multiple partners in the modernization is critical for success.  This talk will focus on the development of the consortium, how the consortium plans for NextGen initiatives, the series of technology demonstrations we have produced and plans for the future of NextGen testing and implementation. 

  14. The Integrated Airport: Building a Successful NextGen Testbed

    SciTech Connect

    Frederick-Recascino, Christina; Sweigard, Doug; Lester, Wade

    2009-02-18

    This presentation will describe a unique public-private partnership - the Integrated Airport - that was created to engage in research and testing related to NextGen Technology deployment.  NextGen refers to the program that will be initiated to modernize the US National Airspace.  As with any major, multi-decade initiative, such as NextGen, integration of work efforts by multiple partners in the modernization is critical for success.  This talk will focus on the development of the consortium, how the consortium plans for NextGen initiatives, the series of technology demonstrations we have produced and plans for the future of NextGen testing and implementation. 

  15. Critical Role for Env as well as Gag-Pol in Control of a Simian-Human Immunodeficiency Virus 89.6P Challenge by a DNA Prime/Recombinant Modified Vaccinia Virus Ankara Vaccine

    PubMed Central

    Amara, Rama Rao; Smith, James M.; Staprans, Silvija I.; Montefiori, David C.; Villinger, Francois; Altman, John D.; O'Neil, Shawn P.; Kozyr, Natalia L.; Xu, Yan; Wyatt, Linda S.; Earl, Patricia L.; Herndon, James G.; McNicholl, Janet M.; McClure, Harold M.; Moss, Bernard; Robinson, Harriet L.

    2002-01-01

    Cellular immune responses against epitopes in conserved Gag and Pol sequences of human immunodeficiency virus type 1 have become popular targets for candidate AIDS vaccines. Recently, we used a simian-human immunodeficiency virus model (SHIV 89.6P) with macaques to demonstrate the control of a pathogenic mucosal challenge by priming with Gag-Pol-Env-expressing DNA and boosting with Gag-Pol-Env-expressing recombinant modified vaccinia virus Ankara (rMVA). Here we tested Gag-Pol DNA priming and Gag-Pol rMVA boosting to evaluate the contribution of anti-Env immune responses to viral control. The Gag-Pol vaccine raised frequencies of Gag-specific T cells similar to those raised by the Gag-Pol-Env vaccine. Following challenge, these rapidly expanded to counter the challenge infection. Despite this, the control of the SHIV 89.6P challenge was delayed and inconsistent in the Gag-Pol-vaccinated group and all of the animals underwent severe and, in most cases, sustained loss of CD4+ cells. Interestingly, most of the CD4+ cells that were lost in the Gag-Pol-vaccinated group were uninfected cells. We suggest that the rapid appearance of binding antibody for Env in Gag-Pol-Env-vaccinated animals helped protect uninfected CD4+ cells from Env-induced apoptosis. Our results highlight the importance of immune responses to Env, as well as to Gag-Pol, in the control of immunodeficiency virus challenges and the protection of CD4+ cells. PMID:12021347

  16. Proposal for two new genera, Brevibacillus gen. nov. and Aneurinibacillus gen. nov.

    PubMed

    Shida, O; Takagi, H; Kadowaki, K; Komagata, K

    1996-10-01

    16S rRNA gene sequences of the type strains of 11 species belonging to the Bacillus brevis and Bacillus aneurinolyticus groups were determined. On the basis of the results of gene sequence analyses, these species were separated into two clusters. The B. brevis cluster included 10 species, namely, Bacillus brevis, Bacillus agri, Bacillus centrosporus, Bacillus choshinensis, Bacillus parabrevis, Bacillus reuszeri, Bacillus formosus, Bacillus borstelensis, Bacillus laterosporus, and Bacillus thermoruber. Bacillus aneurinolyticus and Bacillus migulanus belonged to the B. aneurinolyticus cluster. Moreover, the two clusters were phylogenetically distinct from other Bacillus, Amphibacillus, Sporolactobacillus, Paenibacillus, and Alicyclobacillus species. On the basis of our data, we propose reclassification of the B. brevis cluster as Brevibacillus gen. nov. and reclassification of the B. aneurinolyticus cluster as Aneurinibacillus gen. nov. By using 16S rRNA gene sequence alignments, two specific PCR amplification primers were designed for differentiating the two new genera from each other and from other aerobic, endospore-forming organisms.

  17. HIV-1 Env C2-V4 Diversification in a Slow-Progressor Infant Reveals a Flat but Rugged Fitness Landscape

    PubMed Central

    Smith, S. Abigail; Wood, Charles; West, John T.

    2013-01-01

    Human immunodeficiency virus type-1 (HIV-1) fitness has been associated with virus entry, a process mediated by the envelope glycoprotein (Env). We previously described Env genetic diversification in a Zambian, subtype C infected, slow-progressor child (1157i) in parallel with an evolving neutralizing antibody response. Because of the role the Variable-3 loop (V3) plays in transmission, cell tropism, neutralization sensitivity, and fitness, longitudinally isolated 1157i C2-V4 alleles were cloned into HIV-1NL4-3-eGFP and -DsRed2 infectious molecular clones. The fluorescent reporters allowed for dual-infection competitions between all patient-derived C2-V4 chimeras to quantify the effect of V3 diversification and selection on fitness. ‘Winners’ and ‘losers’ were readily discriminated among the C2-V4 alleles. Exceptional sensitivity for detection of subtle fitness differences was revealed through analysis of two alleles differing in a single synonymous amino acid. However, when the outcomes of N = 33 competitions were averaged for each chimera, the aggregate analysis showed that despite increasing diversification and divergence with time, natural selection of C2-V4 sequences in this individual did not appear to be producing a ‘survival of the fittest’ evolutionary pattern. Rather, we detected a relatively flat fitness landscape consistent with mutational robustness. Fitness outcomes were then correlated with individual components of the entry process. Env incorporation into particles correlated best with fitness, suggesting a role for Env avidity, as opposed to receptor/coreceptor affinity, in defining fitness. Nevertheless, biochemical analyses did not identify any step in HIV-1 entry as a dominant determinant of fitness. Our results lead us to conclude that multiple aspects of entry contribute to maintaining adequate HIV-1 fitness, and there is no surrogate analysis for determining fitness. The capacity for subtle polymorphisms in Env to nevertheless

  18. Nedd4-Mediated Increase in HIV-1 Gag and Env Proteins and Immunity following DNA-Vaccination of BALB/c Mice

    PubMed Central

    Lewis, Brad; Whitney, Stephen; Hudacik, Lauren; Galmin, Lindsey; Huaman, Maria Cecilia; Cristillo, Anthony D.

    2014-01-01

    The late assembly domain of many viruses is critical for budding. Within these domains, encoded in viral structural proteins, are the conserved motifs PTAP, PPxY and YPxL. These sequences are the key determinants for association of viral proteins with intracellular molecules such as Tsg101, Nedd4 and AIP1/ALIX. While roles for Tsg101 and AIP1/ALIX in HIV-1 budding have been well established, less is known about the role of Nedd4. Recent studies, however, have identified a function for Nedd4-like protein in HIV-1 release. In this study, we investigated post-transcriptional changes of Nedd4 following SHIVSF162P3 infection of rhesus macaques, its role on HIV-1 p24 and gp120 levels in vitro and its potential as an immune modulator in HIV vaccination of BALB/c mice. Increased Nedd4 protein levels were noted in both CD4+ and CD8+ T cells following SHIVSF162P3-infection of naïve macaques. Transient co-transfection studies in 293 cells with HXB2 and Nedd4 demonstrated a Nedd4-mediated increase in p24 and gp120 levels. This increase was found to be dependent on the Ca2+/calmodulin-regulated phospholipid binding C2 domain and not ubiquitin ligase activity or HIV LTR activity. Co-transfection of Nedd4 with plasmid DNA expressing Gag or Env was further shown to augment both intracellular and extracellular Gag or Env proteins. To assess the potential of Nedd4 as an immune modulator, BALB/c mice were immunized intramuscularly with plasmid DNA encoding HIV gag, env and Nedd4. Nedd4 co-administration was found to increase serum anti-p24 but not anti-gp120 antibodies. Nedd4 co-injection was found to have no affect on Gag- or Env-specific IFNγ but had a trend of increased Gag-specific IL-6, IL-17A and TNFα that was not seen following Env stimulation. Based on our initial findings, Nedd4-mediated changes in HIV protein levels and its potential use in HIV-1 vaccine development warrants further investigation. PMID:24614057

  19. Increased Sensitivity to Broadly Neutralizing Antibodies of End-Stage Disease R5 HIV-1 Correlates with Evolution in Env Glycosylation and Charge

    PubMed Central

    Sterjovski, Jasminka; Uchtenhagen, Hannes; Churchill, Melissa J.; Karlsson, Anders; Albert, Jan; Achour, Adnane; Gorry, Paul R.; Fenyö, Eva Maria; Jansson, Marianne

    2011-01-01

    Background Induction of broadly neutralizing antibodies, such as the monoclonal antibodies IgGb12, 2F5 and 2G12, is the objective of most antibody-based HIV-1 vaccine undertakings. However, despite the relative conserved nature of epitopes targeted by these antibodies, mechanisms underlying the sensitivity of circulating HIV-1 variants to broadly neutralizing antibodies are not fully understood. Here we have studied sensitivity to broadly neutralizing antibodies of HIV-1 variants that emerge during disease progression in relation to molecular alterations in the viral envelope glycoproteins (Env), using a panel of primary R5 HIV-1 isolates sequentially obtained before and after AIDS onset. Principal Findings HIV-1 R5 isolates obtained at end-stage disease, after AIDS onset, were found to be more sensitive to neutralization by TriMab, an equimolar mix of the IgGb12, 2F5 and 2G12 antibodies, than R5 isolates from the chronic phase. The increased sensitivity correlated with low CD4+ T cell count at time of virus isolation and augmented viral infectivity. Subsequent sequence analysis of multiple env clones derived from the R5 HIV-1 isolates revealed that, concomitant with increased TriMab neutralization sensitivity, end-stage R5 variants displayed envelope glycoproteins (Envs) with reduced numbers of potential N-linked glycosylation sites (PNGS), in addition to increased positive surface charge. These molecular changes in Env also correlated to sensitivity to neutralization by the individual 2G12 monoclonal antibody (mAb). Furthermore, results from molecular modeling suggested that the PNGS lost at end-stage disease locate in the proximity to the 2G12 epitope. Conclusions Our study suggests that R5 HIV-1 variants with increased sensitivity to broadly neutralizing antibodies, including the 2G12 mAb, may emerge in an opportunistic manner during severe immunodeficiency as a consequence of adaptive molecular Env changes, including loss of glycosylation and gain of positive

  20. Nedd4-mediated increase in HIV-1 Gag and Env proteins and immunity following DNA-vaccination of BALB/c mice.

    PubMed

    Lewis, Brad; Whitney, Stephen; Hudacik, Lauren; Galmin, Lindsey; Huaman, Maria Cecilia; Cristillo, Anthony D

    2014-01-01

    The late assembly domain of many viruses is critical for budding. Within these domains, encoded in viral structural proteins, are the conserved motifs PTAP, PPxY and YPxL. These sequences are the key determinants for association of viral proteins with intracellular molecules such as Tsg101, Nedd4 and AIP1/ALIX. While roles for Tsg101 and AIP1/ALIX in HIV-1 budding have been well established, less is known about the role of Nedd4. Recent studies, however, have identified a function for Nedd4-like protein in HIV-1 release. In this study, we investigated post-transcriptional changes of Nedd4 following SHIVSF162P3 infection of rhesus macaques, its role on HIV-1 p24 and gp120 levels in vitro and its potential as an immune modulator in HIV vaccination of BALB/c mice. Increased Nedd4 protein levels were noted in both CD4+ and CD8+ T cells following SHIVSF162P3-infection of naïve macaques. Transient co-transfection studies in 293 cells with HXB2 and Nedd4 demonstrated a Nedd4-mediated increase in p24 and gp120 levels. This increase was found to be dependent on the Ca2+/calmodulin-regulated phospholipid binding C2 domain and not ubiquitin ligase activity or HIV LTR activity. Co-transfection of Nedd4 with plasmid DNA expressing Gag or Env was further shown to augment both intracellular and extracellular Gag or Env proteins. To assess the potential of Nedd4 as an immune modulator, BALB/c mice were immunized intramuscularly with plasmid DNA encoding HIV gag, env and Nedd4. Nedd4 co-administration was found to increase serum anti-p24 but not anti-gp120 antibodies. Nedd4 co-injection was found to have no affect on Gag- or Env-specific IFNγ but had a trend of increased Gag-specific IL-6, IL-17A and TNFα that was not seen following Env stimulation. Based on our initial findings, Nedd4-mediated changes in HIV protein levels and its potential use in HIV-1 vaccine development warrants further investigation.

  1. NextGen Future Safety Assessment Game

    NASA Technical Reports Server (NTRS)

    Ancel, Ersin; Gheorghe, Adian; Jones, Sharon Monica

    2010-01-01

    The successful implementation of the next generation infrastructure systems requires solid understanding of their technical, social, political and economic aspects along with their interactions. The lack of historical data that relate to the long-term planning of complex systems introduces unique challenges for decision makers and involved stakeholders which in turn result in unsustainable systems. Also, the need to understand the infrastructure at the societal level and capture the interaction between multiple stakeholders becomes important. This paper proposes a methodology in order to develop a holistic approach aiming to provide an alternative subject-matter expert (SME) elicitation and data collection method for future sociotechnical systems. The methodology is adapted to Next Generation Air Transportation System (NextGen) decision making environment in order to demonstrate the benefits of this holistic approach.

  2. NextGen Future Safety Assessment Game

    NASA Technical Reports Server (NTRS)

    Ancel, Ersin; Gheorghe, Adrian; Jones, Sharon Monica

    2011-01-01

    The successful implementation of the next generation infrastructure systems requires solid understanding of their technical, social, political and economic aspects along with their interactions. The lack of historical data that relate to the long-term planning of complex systems introduces unique challenges for decision makers and involved stakeholders which in turn result in unsustainable systems. Also, the need to understand the infrastructure at the societal level and capture the interaction between multiple stakeholders becomes important. This paper proposes a methodology in order to develop a holistic approach aiming to provide an alternative subject-matter expert (SME) elicitation and data collection method for future sociotechnical systems. The methodology is adapted to Next Generation Air Transportation System (NextGen) decision making environment in order to demonstrate the benefits of this holistic approach.

  3. ALS - resources

    MedlinePlus

    Resources - ALS ... The following organizations are good resources for information on amyotrophic lateral sclerosis : Muscular Dystrophy Association -- www.mda.org/disease/amyotrophic-lateral-sclerosis National Amyotrophic Lateral Sclerosis (ALS) ...

  4. RxGen General Optical Model Prescription Generator

    NASA Technical Reports Server (NTRS)

    Sigrist, Norbert

    2012-01-01

    RxGen is a prescription generator for JPL's in-house optical modeling software package called MACOS (Modeling and Analysis for Controlled Optical Systems), which is an expert optical analysis software package focusing on modeling optics on dynamic structures, deformable optics, and controlled optics. The objectives of RxGen are to simplify and automate MACOS prescription generations, reducing errors associated with creating such optical prescriptions, and improving user efficiency without requiring MACOS proficiency. RxGen uses MATLAB (a high-level language and interactive environment developed by MathWorks) as the development and deployment platform, but RxGen can easily be ported to another optical modeling/analysis platform. Running RxGen within the modeling environment has the huge benefit that variations in optical models can be made an integral part of the modeling state. For instance, optical prescription parameters determined as external functional dependencies, optical variations by controlling the in-/exclusion of optical components like sub-systems, and/or controlling the state of all components. Combining the mentioned capabilities and flexibilities with RxGen's optical abstraction layer completely eliminates the hindering aspects for requiring proficiency in writing/editing MACOS prescriptions, allowing users to focus on the modeling aspects of optical systems, i.e., increasing productivity and efficiency. RxGen provides significant enhancements to MACOS and delivers a framework for fast prototyping as well as for developing very complex controlled optical systems.

  5. Head-Worn Displays for NextGen

    NASA Technical Reports Server (NTRS)

    Bailey, Randall E.; Shelton, Kevin J.; Arthur, J. J.

    2011-01-01

    The operating concepts emerging under the Next Generation air transportation system (NextGen) require new technology and procedures - not only on the ground-side - but also on the flight deck. Flight deck display and decision support technologies are specifically targeted to overcome aircraft safety barriers that might otherwise constrain the full realization of NextGen. One such technology is the very lightweight, unobtrusive head-worn display (HWD). HWDs with an integrated head-tracking system are being researched as they offer significant potential benefit under emerging NextGen operational concepts. Two areas of benefit for NextGen are defined. First, the HWD may be designed to be equivalent to the Head-Up Display (HUD) using Virtual HUD concepts. As such, these operational credits may be provided to significantly more aircraft for which HUD installation is neither practical nor possible. Second, the HWD provides unique display capabilities, such as an unlimited field-of-regard. These capabilities may be integral to emerging NextGen operational concepts, eliminating safety issues which might otherwise constrain the full realization of NextGen. The paper details recent research results, current HWD technology limitations, and future technology development needed to realize HWDs as a enabling technology for NextGen.

  6. Daily Sampling of an HIV-1 Patient with Slowly Progressing Disease Displays Persistence of Multiple env Subpopulations Consistent with Neutrality

    PubMed Central

    Wilbe Ramsay, Karin; Alaeus, Annette; Albert, Jan; Leitner, Thomas

    2011-01-01

    The molecular evolution of HIV-1 is characterized by frequent substitutions, indels and recombination events. In addition, a HIV-1 population may adapt through frequency changes of its variants. To reveal such population dynamics we analyzed HIV-1 subpopulation frequencies in an untreated patient with stable, low plasma HIV-1 RNA levels and close to normal CD4+ T-cell levels. The patient was intensively sampled during a 32-day period as well as approximately 1.5 years before and after this period (days −664, 1, 2, 3, 11, 18, 25, 32 and 522). 77 sequences of HIV-1 env (approximately 3100 nucleotides) were obtained from plasma by limiting dilution with 7–11 sequences per time point, except day −664. Phylogenetic analysis using maximum likelihood methods showed that the sequences clustered in six distinct subpopulations. We devised a method that took into account the relatively coarse sampling of the population. Data from days 1 through 32 were consistent with constant within-patient subpopulation frequencies. However, over longer time periods, i.e. between days 1…32 and 522, there were significant changes in subpopulation frequencies, which were consistent with evolutionarily neutral fluctuations. We found no clear signal of natural selection within the subpopulations over the study period, but positive selection was evident on the long branches that connected the subpopulations, which corresponds to >3 years as the subpopulations already were established when we started the study. Thus, selective forces may have been involved when the subpopulations were established. Genetic drift within subpopulations caused by de novo substitutions could be resolved after approximately one month. Overall, we conclude that subpopulation frequencies within this patient changed significantly over a time period of 1.5 years, but that this does not imply directional or balancing selection. We show that the short-term evolution we study here is likely representative for many

  7. Genetic shift of env V3 loop viral sequences in patients with HIV-associated neurocognitive disorder during antiretroviral therapy.

    PubMed

    Eggers, Christian; Müller, Oliver; Thordsen, Ingo; Schreiber, Michael; Methner, Axel

    2013-12-01

    The development of human immunodeficiency virus type 1 (HIV)-associated neurocognitive disorder (HAND) involves the adaptation of viral sequences coding for the V3 loop of the env protein. The plasma and cerebrospinal fluid (CSF) may contain viral populations from various cellular sources and with differing pathogenicity. Combination antiretroviral therapy (cART) may alter the relative abundance of these viral populations, leading to a genetic shift. We characterized plasma and CNS viral populations prior to and during cART and relate the findings to viral elimination kinetics and the clinical phenotype. Longitudinal plasma and CSF samples of five chronically infected HIV patients, four of whom had HAND, and one seroconverter were analyzed for V3 sequences by RT-PCR and sequence analysis. In the chronically infected patients, pre-cART plasma and CSF viral sequences were different irrespective of viral elimination kinetics and clinical phenotype. cART induced replacement of plasma viral populations in all subjects. CSF viral populations underwent a clear genetic shift in some patients but remained stable in others. This was not dependent on the presence of HAND. The genetic shift of CSF V3 sequences was absent in the two subjects whose CSF viral load initially increased during cART. In one patient, pre- and post-treatment CSF sequences were closely related to the post-treatment plasma sequences, suggesting a common cellular source. We found heterogeneous patterns of genetic compartmentalization and genetic shift over time. Although these did not closely match viral elimination kinetics and clinical phenotype, the results imply different patterns of the dynamics and relative contribution of compartment-specific virus populations in chronic HIV infection.

  8. 76 FR 22162 - Third Meeting RTCA NextGen Advisory Committee (NAC)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-20

    ... Recommendations to be Considered for Submission to FAA, NextGen Performance Metrics. Preliminary Reports on Key NextGen Performance Indicators. NextGen Measurement Methodology. Review Recommendations to be... Federal Aviation Administration Third Meeting RTCA NextGen Advisory Committee (NAC) AGENCY: Federal...

  9. A new genus and species of isanthid sea anemone (Cnidaria: Anthozoa: Actiniaria) from Chilean Patagonia, Anthoparactis fossii n. gen. et sp

    NASA Astrophysics Data System (ADS)

    Häussermann, Verena; Rodríguez, Estefanía

    2014-09-01

    We describe a new genus and species of sea anemone from Chilean Patagonia. Anthoparactis fossii n. gen. et sp. adds another acontiate genus and species to the family Isanthidae Carlgren, 1938. Anthoparactis n. gen. differs from the other isanthid genera in having the same number of mesenteries distally and proximally, acontia with basitrichs only, and a column with verrucae distally. Anthoparactis fossii n. sp. differs from the most similar species, Isoparactis fionae Lauretta et al., 2013, in the number of cycles of mesenteries and tentacles, structures of the column, colour pattern of the oral disc, cnidae, and geographical distribution. Isanthidae now includes seven genera and 11 species.

  10. Env-2dCD4 S60C complexes act as super immunogens and elicit potent, broadly neutralizing antibodies against clinically relevant human immunodeficiency virus type 1 (HIV-1).

    PubMed

    Killick, Mark A; Grant, Michelle L; Cerutti, Nichole M; Capovilla, Alexio; Papathanasopoulos, Maria A

    2015-11-17

    The ability to induce a broadly neutralizing antibody (bNAb) response following vaccination is regarded as a crucial aspect in developing an effective vaccine against human immunodeficiency virus type 1 (HIV-1). The bNAbs target the HIV-1 envelope glycoprotein (Env) which is exposed on the virus surface, thereby preventing cell entry. To date, conventional vaccine approaches such as the use of Env-based immunogens have been unsuccessful. We expressed, purified, characterized and evaluated the immunogenicity of several unique HIV-1 subtype C Env immunogens in small animals. Here we report that vaccine immunogens based on Env liganded to a two domain CD4 variant, 2dCD4(S60C) are capable of consistently eliciting potent, broadly neutralizing antibody responses in New Zealand white rabbits against a panel of clinically relevant HIV-1 pseudoviruses. This was irrespective of the Env protein subtype and context. Importantly, depletion of the anti-CD4 antibodies appeared to abrogate the neutralization activity in the rabbit sera. Taken together, this data suggests that the Env-2dCD4(S60C) complexes described here are "super" immunogens, and potentially immunofocus antibody responses to a unique epitope spanning the 2dCD4(60C). Recent data from the two available anti-CD4 monoclonal antibodies, Ibalizumab and CD4-Ig (and bispecific variants thereof) have highlighted that the use of these broad and potent entry inhibitors could circumvent the need for a conventional vaccine targeting HIV-1. Overall, the ability of the unique Env-2dCD4(S60C) complexes to elicit potent bNAb responses has not been described previously, reinforcing that further investigation for their utility in preventing and controlling HIV-1/SIV infection is warranted.

  11. Site-directed mutagenesis of the codon for Ile-25 in gPr80env alters the neurovirulence of ts1, a mutant of Moloney murine leukemia virus TB.

    PubMed Central

    Szurek, P F; Floyd, E; Yuen, P H; Wong, P K

    1990-01-01

    ts1, a spontaneous temperature-sensitive mutant of Moloney murine leukemia virus TB, causes hind-limb paralysis in mice. A Val-25----Ile substitution in gPr80env is responsible for temperature sensitivity, inefficient processing of gPr80env, and neurovirulence. In this study, the Ile-25 in gPr80env was replaced with Thr, Ala, Leu, Gly, and Glu by site-directed mutagenesis of the codon for Ile-25 to generate a new set of mutant viruses, i.e., ts1-T, -A, -L, -G, and -E, respectively. The phenotypic characteristics of these mutant viruses differed from those of ts1. For each mutant, the degree of temperature sensitivity was correlated with the degree of inefficient processing of gPr80env, and the following rank order was observed for both parameters: ts1-E greater than ts1-G greater than ts1-L greater than ts1-A greater than ts1 greater than ts1-T. In FVB/N mice, mutant viruses of low and intermediate temperature sensitivity and inefficiency in processing of gPr80env were neurovirulent and consistently caused mutant-specific disease profiles: ts1-T caused severe whole-body tremor, ts1-A generally caused hind-limb paralysis, and ts1-L generally caused a delayed-onset paraparesis. By 150 days postinfection, FVB/N mice that were infected with ts1-G and -E, mutants of high temperature sensitivity and inefficiency in processing of gPr80env, had lymphoid leukemia instead of a neurological disease. These results suggest that the dynamics of gPr80env processing are important in determining the neurovirulent phenotype in vivo. Images PMID:2214016

  12. Molecular phylogeny of Urosomoida agilis, and new combinations: Hemiurosomoida longa gen. nov., comb. nov., and Heterourosomoida lanceolata gen. nov., comb. nov. (Ciliophora, Hypotricha).

    PubMed

    Singh, Jasbir; Kamra, Komal

    2015-02-01

    For years, systematics of three species, Urosomoida agilis (Engelmann, 1862) Hemberger in Foissner, 1982, Urosomoida longa (Gelei and Szabados, 1950) Foissner et al., 1991 and Oxytricha lanceolata Shibuya, 1930, has remained unresolved due to lack of adequate molecular data. Though, it is known since several years that the three species are not very closely related. In the present paper, 18S rRNA gene sequences for two key species, U. agilis and U. longa, and their morphology and morphometry have been analyzed. Molecular phylogeny inferred from maximum likelihood, neighbour joining and maximum parsimony methods has adequately removed ambiguity over their systematics. In phylogenetic trees, U. agilis clustered consistently with non-stylonychine oxytrichids. Both Urosomoida longa and Oxytricha lanceolata clustered consistently away from U. agilis and O. granulifera, the type species of the genera Urosomoida and Oxytricha, respectively. As a result of the current molecular phylogenetic investigation and based on previously inferred morphological and morphogenetic data it is proposed to remove Urosomoida longa and Oxytricha lanceolata from Urosomoida and incertae sedis in Oxytricha, respectively, and establish two new generic combinations, Hemiurosomoida longa gen. nov., comb. nov. and Heterourosomoida lanceolata gen. nov., comb. nov. for them.

  13. Structures and stability of metal-doped GenM (n = 9, 10) clusters

    DOE PAGES

    Qin, Wei; Lu, Wen-Cai; Xia, Lin-Hua; ...

    2015-06-26

    The lowest-energy structures of neutral and cationic Ge nM (n = 9, 10; M = Si, Li, Mg, Al, Fe, Mn, Pb, Au, Ag, Yb, Pm and Dy) clusters were studied by genetic algorithm (GA) and first-principles calculations. The calculation results show that doping of the metal atoms and Si into Ge9 and Ge10 clusters is energetically favorable. Most of the metal-doped Ge cluster structures can be viewed as adding or substituting metal atom on the surface of the corresponding ground-state Gen clusters. However, the neutral and cationic FeGe9,10,MnGe9,10 and Ge10Al are cage-like with the metal atom encapsulated inside. Suchmore » cage-like transition metal doped Gen clusters are shown to have higher adsorption energy and thermal stability. Our calculation results suggest that Ge9,10Fe and Ge9Si would be used as building blocks in cluster-assembled nanomaterials because of their high stabilities.« less

  14. EPCGen2 Pseudorandom Number Generators: Analysis of J3Gen

    PubMed Central

    Peinado, Alberto; Munilla, Jorge; Fúster-Sabater, Amparo

    2014-01-01

    This paper analyzes the cryptographic security of J3Gen, a promising pseudo random number generator for low-cost passive Radio Frequency Identification (RFID) tags. Although J3Gen has been shown to fulfill the randomness criteria set by the EPCglobal Gen2 standard and is intended for security applications, we describe here two cryptanalytic attacks that question its security claims: (i) a probabilistic attack based on solving linear equation systems; and (ii) a deterministic attack based on the decimation of the output sequence. Numerical results, supported by simulations, show that for the specific recommended values of the configurable parameters, a low number of intercepted output bits are enough to break J3Gen. We then make some recommendations that address these issues. PMID:24721767

  15. 98. SWITCH HOUSE MAIN LOBBY, GEN 3 CIRCUIT BREAKER, VIEW ...

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

    98. SWITCH HOUSE MAIN LOBBY, GEN 3 CIRCUIT BREAKER, VIEW OF OPPOSITE SIDE FROM HAER No. PA-505-97 - Delaware County Electric Company, Chester Station, Delaware River at South end of Ward Street, Chester, Delaware County, PA

  16. 1. GENERAL VIEW. Statues: Maj. Gen George Meade by Daniel ...

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

    1. GENERAL VIEW. Statues: Maj. Gen George Meade by Daniel Chester French, south side; Maj. Gen. John Reynolds by Charles Grafly, north side. Equestrian Statues: Maj. Gen George B. McClellan by Edward C. Potter, south side; Maj. Gen Winfield S. Hancock by J.Q.A. Ward, north side. The statue at the base of northern inner pedestal is Richard Smith, a type founder and donor of the Memorial. The niches are filled with eight colossal busts including Union generals, admirals, Pennsylvania governor, Memorial's architects (John T. and James H. Windrim), and executor of Smith's will. The frieze is carved with the names of eighty-four prominent Pennsylvania participants in the Civil War. - Smith Memorial Arch, Philadelphia, Philadelphia County, PA

  17. The haemosporidian parasites of bats with description of Sprattiella alecto gen. nov., sp. nov.

    PubMed

    Landau, I; Chavatte, J M; Karadjian, G; Chabaud, A; Beveridge, I

    2012-05-01

    Four species of Haemoproteidae were found in Pteropus alecto Temminck, 1837 in Queensland, Australia: i) Johnsprentia copemani, Landau et al., 2012; ii) Sprattiella alecto gen. nov., sp. nov., characterised by schizonts in the renal vessels; iii) Hepatocystis levinei, Landau et al., 1985, originally described from Pteropus poliocephalus Temminck, 1825 and, experimentally from Culicoides nubeculosus and found in this new host and for which features of the hepatic schizonts are reported; iv) gametocytes of Hepatocystis sp. which are illustrated but cannot be assigned to a known species. A tentative interpretation of phylogenetic characters of haemosporidians of bats is provided from the morphology of the gametocytes and localisation of the tissue stages with respect to recent data on the phylogeny of bats.

  18. CD4 down-modulation during infection of human T cells with human immunodeficiency virus type 1 involves independent activities of vpu, env, and nef.

    PubMed Central

    Chen, B K; Gandhi, R T; Baltimore, D

    1996-01-01

    The human immunodeficiency virus type 1 (HIV-1) genes vpu, env, and nef have all been implicated in modulating the levels of cell surface CD4 on infected cells. To quantitatively assess the relative contribution of each gene product to the regulation of CD4 during HIV infection of Jurkat T cells and peripheral blood mononuclear cells, we have developed an infectious HIV reporter system which expresses different combinations of these genes. To distinguish infected cells in the early or late stages of infection from uninfected cells, these viruses were designed to express human placental alkaline phosphatase with the kinetics of either early or late viral genes. Flow cytometry to detect placental alkaline phosphatase and CD4 in infected cells showed that vpu, env, and nef are independently capable of down-modulation of CD4. As predicted by their respective expression patterns, nef down-modulated CD4 rapidly during the early phase of virus infection whereas vpu and env functioned late in the infection. In both Jurkat cells and peripheral blood mononuclear cells, a combination of the three genes was more efficient than any one or two genes, demonstrating that all three genes are required to achieve maximal CD4 down-modulation. In primary cells, down-modulation of CD4 was less efficient than in Jurkat cells and there was a stronger dependence on nef function for reducing cell surface CD4. HIV therefore has three genes that are able to independently down-modulate CD4; together, they can eliminate the bulk of cell surface CD4. PMID:8709227

  19. An adenovirus-simian immunodeficiency virus env vaccine elicits humoral, cellular, and mucosal immune responses in rhesus macaques and decreases viral burden following vaginal challenge.

    PubMed Central

    Buge, S L; Richardson, E; Alipanah, S; Markham, P; Cheng, S; Kalyan, N; Miller, C J; Lubeck, M; Udem, S; Eldridge, J; Robert-Guroff, M

    1997-01-01

    Six female rhesus macaques were immunized orally and intranasally at 0 weeks and intratracheally at 12 weeks with an adenovirus type 5 host range mutant (Ad5hr)-simian immunodeficiency virus SIVsm env recombinant and at 24 and 36 weeks with native SIVmac251 gp120 in Syntex adjuvant. Four macaques received the Ad5hr vector and adjuvant alone; two additional controls were naive. In vivo replication of the Ad5hr wild-type and recombinant vectors occurred with detection of Ad5 DNA in stool samples and/or nasal secretions in all macaques and increases in Ad5 neutralizing antibody in 9 of 10 macaques following Ad administrations. SIV-specific neutralizing antibodies appeared after the second recombinant immunization and rose to titers > 10,000 following the second subunit boost. Immunoglobulin G (IgG) and IgA antibodies able to bind gp120 developed in nasal and rectal secretions, and SIV-specific IgGs were also observed in vaginal secretions and saliva. T-cell proliferative responses to SIV gp140 and T-helper epitopes were sporadically detected in all immunized macaques. Following vaginal challenge with SIVmac251, transient or persistent infection resulted in both immunized and control monkeys. The mean viral burden in persistently infected immunized macaques was significantly decreased in the primary infection period compared to that of control macaques. These results establish in vivo use of the Ad5hr vector, which overcomes the host range restriction of human Ads for rhesus macaques, thereby providing a new model for evaluation of Ad-based vaccines. In addition, they show that a vaccine regimen using the Ad5hr-SIV env recombinant and gp120 subunit induces strong humoral, cellular, and mucosal immunity in rhesus macaques. The reduced viral burden achieved solely with an env-based vaccine supports further development of Ad-based vaccines comprising additional viral components for immune therapy and AIDS vaccine development. PMID:9343211

  20. Pre-existing neutralizing antibody mitigates B cell dysregulation and enhances the Env-specific antibody response in SHIV-infected rhesus macaques

    PubMed Central

    Jaworski, Juan Pablo; Bryk, Peter; Brower, Zachary; Zheng, Bo; Hessell, Ann J.; Rosenberg, Alexander F.; Wu, Tong Tong; Sanz, Ignacio; Keefer, Michael C.; Haigwood, Nancy L.

    2017-01-01

    Our central hypothesis is that protection against HIV infection will be powerfully influenced by the magnitude and quality of the B cell response. Although sterilizing immunity, mediated by pre-formed abundant and potent antibodies is the ultimate goal for B cell-targeted HIV vaccine strategies, scenarios that fall short of this may still confer beneficial defenses against viremia and disease progression. We evaluated the impact of sub-sterilizing pre-existing neutralizing antibody on the B cell response to SHIV infection. Adult male rhesus macaques received passive transfer of a sub-sterilizing amount of polyclonal neutralizing immunoglobulin (Ig) purified from previously infected animals (SHIVIG) or control Ig prior to intra-rectal challenge with SHIVSF162P4 and extensive longitudinal sampling was performed. SHIVIG treated animals exhibited significantly reduced viral load and increased de novo Env-specific plasma antibody. Dysregulation of the B cell profile was grossly apparent soon after infection in untreated animals; exemplified by a ≈50% decrease in total B cells in the blood evident 2–3 weeks post-infection which was not apparent in SHIVIG treated animals. IgD+CD5+CD21+ B cells phenotypically similar to marginal zone-like B cells were highly sensitive to SHIV infection, becoming significantly decreased as early as 3 days post-infection in control animals, while being maintained in SHIVIG treated animals, and were highly correlated with the induction of Env-specific plasma antibody. These results suggest that B cell dysregulation during the early stages of infection likely contributes to suboptimal Env-specific B cell and antibody responses, and strategies that limit this dysregulation may enhance the host’s ability to eliminate HIV. PMID:28222180

  1. EarthEnv-DEM90: A nearly-global, void-free, multi-scale smoothed, 90m digital elevation model from fused ASTER and SRTM data

    NASA Astrophysics Data System (ADS)

    Robinson, Natalie; Regetz, James; Guralnick, Robert P.

    2014-01-01

    A variety of DEM products are available to the public at no cost, though all are characterized by trade-offs in spatial coverage, data resolution, and quality. The absence of a high-resolution, high-quality, well-described and vetted, free, global consensus product was the impetus for the creation of a new DEM product described here, 'EarthEnv-DEM90'. This new DEM is a compilation dataset constructed via rigorous techniques by which ASTER GDEM2 and CGIAR-CSI v4.1 products were fused into a quality-enhanced, consistent grid of elevation estimates that spans ∼91% of the globe. EarthEnv-DEM90 was assembled using methods for seamlessly merging input datasets, thoroughly filling voids, and smoothing data irregularities (e.g. those caused by DEM noise) from the approximated surface. The result is a DEM product in which elevational artifacts are strongly mitigated from the input data fusion zone, substantial voids are filled in the northern-most regions of the globe, and the entire DEM exhibits reduced terrain noise. As important as the final product is a well defined methodology, along with new processing techniques and careful attention to final outputs, that extends the value and usability of the work beyond just this single product. Finally, we outline EarthEnv-DEM90 acquisition instructions and metadata availability, so that researchers can obtain this high-resolution, high-quality, nearly-global new DEM product for the study of wide-ranging global phenomena.

  2. New sequestrate fungi from Guyana: Jimtrappea guyanensis gen. sp. nov., Castellanea pakaraimophila gen. sp. nov., and Costatisporus cyanescens gen. sp. nov. (Boletaceae, Boletales).

    PubMed

    Smith, Matthew E; Amses, Kevin R; Elliott, Todd F; Obase, Keisuke; Aime, M Catherine; Henkel, Terry W

    2015-12-01

    Jimtrappea guyanensis gen. sp. nov., Castellanea pakaraimophila gen. sp. nov., and Costatisporus cyanescens gen. sp. nov. are described as new to science. These sequestrate, hypogeous fungi were collected in Guyana under closed canopy tropical forests in association with ectomycorrhizal (ECM) host tree genera Dicymbe (Fabaceae subfam. Caesalpinioideae), Aldina (Fabaceae subfam. Papilionoideae), and Pakaraimaea (Dipterocarpaceae). Molecular data place these fungi in Boletaceae (Boletales, Agaricomycetes, Basidiomycota) and inform their relationships to other known epigeous and sequestrate taxa within that family. Macro- and micromorphological characters, habitat, and multi-locus DNA sequence data are provided for each new taxon. Unique morphological features and a molecular phylogenetic analysis of 185 taxa across the order Boletales justify the recognition of the three new genera.

  3. Marinagarivorans algicola gen. nov., sp. nov., isolated from marine algae.

    PubMed

    Guo, Ling-Yun; Li, Dong-Qi; Sang, Jin; Chen, Guan-Jun; Du, Zong-Jun

    2016-01-27

    Novel agar-degrading, Gram-staining-negative, motile, heterotrophic, facultatively anaerobic and pale yellow-pigmented bacterial strains, designated Z1T and JL1, were isolated from marine algae Gelidium amansii (Lamouroux) and Gracilaria verrucosa, respectively. Growth of the isolates was optimal at 28-30 °C, pH 7.0-7.5 and 1-3% (w/v) NaCl. Both strains contained Q-8 as the sole respiratory quinone. The major cellular fatty acids in strain Z1T were C18:1 ω7c, C16:0 and summed feature 3 (C16:1 ω7c and/or iso-C15:0 2-OH). The predominant polar lipids in strain Z1T were phosphatidylethanolamine (PE), phosphatidylglycerol (PG) and aminolipid (AL). The genomic DNA G+C content of both strains was 45.1 mol%. Strains Z1T and JL1 were closely related, with 99.9% 16S rRNA gene sequence similarity. The average nucleotide identity (ANI) value between strains Z1T and JL1 was 99.3%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strains Z1T and JL1 form a distinct phyletic line within the class Gammaproteobacteria, with less than 92.3% similarity to their closest relatives. Based on data from the current polyphasic study, the isolates are proposed to belong to a new genus and species designated Marinagarivorans algicola gen. nov., sp. nov. The type strain of Marinagarivorans algicola is Z1T (=ATCC BAA-2617T=CICC 10859T).

  4. Skryjelites auritus gen. et sp. nov. and Quasimolites quasimodo gen. et sp. nov.--two new middle Cambrian hyolithids (?Mollusca) from the Czech Republic.

    PubMed

    Valent, Martin; Fatka, Oldřich; Szabad, Michal; Micka, Václav; Marek, Ladislav

    2015-08-28

    Two new endemic genera and species of extinct group of Hyolitha, Skryjelites auritus gen. et sp. nov. and Quasimolites quasimodo gen. et sp. nov. are described and illustrated from the Buchava Formation of the Barrandian area (Czech Republic).

  5. Targeted N-glycan deletion at the receptor-binding site retains HIV Env NFL trimer integrity and accelerates the elicited antibody response.

    PubMed

    Dubrovskaya, Viktoriya; Guenaga, Javier; de Val, Natalia; Wilson, Richard; Feng, Yu; Movsesyan, Arlette; Karlsson Hedestam, Gunilla B; Ward, Andrew B; Wyatt, Richard T

    2017-09-13

    Extensive shielding by N-glycans on the surface of the HIV envelope glycoproteins (Env) restricts B cell recognition of conserved neutralizing determinants. Elicitation of broadly neutralizing antibodies (bNAbs) in selected HIV-infected individuals reveals that Abs capable of penetrating the glycan shield can be generated by the B cell repertoire. Accordingly, we sought to determine if targeted N-glycan deletion might alter antibody responses to Env. We focused on the conserved CD4 binding site (CD4bs) since this is a known neutralizing determinant that is devoid of glycosylation to allow CD4 receptor engagement, but is ringed by surrounding N-glycans. We selectively deleted potential N-glycan sites (PNGS) proximal to the CD4bs on well-ordered clade C 16055 native flexibly linked (NFL) trimers to potentially increase recognition by naïve B cells in vivo. We generated glycan-deleted trimer variants that maintained native-like conformation and stability. Using a panel of CD4bs-directed bNAbs, we demonstrated improved accessibility of the CD4bs on the N-glycan-deleted trimer variants. We showed that pseudoviruses lacking these Env PNGSs were more sensitive to neutralization by CD4bs-specific bNAbs but remained resistant to non-neutralizing mAbs. We performed rabbit immunogenicity experiments using two approaches comparing glycan-deleted to fully glycosylated NFL trimers. The first was to delete 4 PNGS sites and then boost with fully glycosylated Env; the second was to delete 4 sites and gradually re-introduce these N-glycans in subsequent boosts. We demonstrated that the 16055 PNGS-deleted trimers more rapidly elicited serum antibodies that more potently neutralized the CD4bs-proximal-PNGS-deleted viruses in a statistically significant manner and strongly trended towards increased neutralization of fully glycosylated autologous virus. This approach elicited serum IgG capable of cross-neutralizing selected tier 2 viruses lacking N-glycans at residue N276 (natural or

  6. Trimeric gp120-specific bovine monoclonal antibodies require cysteine and aromatic residues in CDRH3 for high affinity binding to HIV Env.

    PubMed

    Heydarchi, Behnaz; Center, Rob J; Bebbington, Jonathan; Cuthbertson, Jack; Gonelli, Christopher; Khoury, Georges; Mackenzie, Charlene; Lichtfuss, Marit; Rawlin, Grant; Muller, Brian; Purcell, Damian

    2017-04-01

    We isolated HIV-1 Envelope (Env)-specific memory B cells from a cow that had developed high titer polyclonal immunoglobulin G (IgG) with broad neutralizing activity after a long duration vaccination with HIV-1AD8 Env gp140 trimers. We cloned the bovine IgG matched heavy (H) and light (L) chain variable (V) genes from these memory B cells and constructed IgG monoclonal antibodies (mAbs) with either a human constant (C)-region/bovine V-region chimeric or fully bovine C and V regions. Among 42 selected Ig+ memory B cells, two mAbs (6A and 8C) showed high affinity binding to gp140 Env. Characterization of both the fully bovine and human chimeric isoforms of these two mAbs revealed them as highly type-specific and capable of binding only to soluble AD8 uncleaved gp140 trimers and covalently stabilized AD8 SOSIP gp140 cleaved trimers, but not monomeric gp120. Genomic sequence analysis of the V genes showed the third heavy complementarity-determining region (CDRH3) of 6A mAb was 21 amino acids in length while 8C CDRH3 was 14 amino acids long. The entire V heavy (VH) region was 27% and 25% diverged for 6A and 8C, respectively, from the best matched germline V genes available, and the CDRH3 regions of 6A and 8C were 47.62% and 78.57% somatically mutated, respectively, suggesting a high level of somatic hypermutation compared with CDRH3 of other species. Alanine mutagenesis of the VH genes of 6A and 8C, showed that CDRH3 cysteine and tryptophan amino acids were crucial for antigen binding. Therefore, these bovine vaccine-induced anti-HIV antibodies shared some of the notable structural features of elite human broadly neutralizing antibodies, such as CDRH3 size and somatic mutation during affinity-maturation. However, while the 6A and 8C mAbs inhibited soluble CD4 binding to gp140 Env, they did not recapitulate the neutralizing activity of the polyclonal antibodies against HIV infection.

  7. Polymorphisms in the HIV-1 gp41 env gene, natural resistance to enfuvirtide (T-20) and pol resistance among pregnant Brazilian women.

    PubMed

    Reis, Mônica Nogueira da Guarda; de Alcântara, Keila Correa; Cardoso, Ludimila Paula Vaz; Stefani, Mariane Martins Araújo

    2014-01-01

    The selective pressure of antiretroviral drugs (ARVs) targeting HIV-1 pol can promote drug resistance mutations in other genomic regions, such as env. Drug resistance among women should be monitored to avoid horizontal and mother-to-child transmission. To describe natural resistance to T-20 (enfuvirtide), gp41 env polymorphisms, mutations in pol and HIV-1 subtypes, 124 pregnant women were recruited. For 98 patients, the gp41 env, protease (PR) and reverse transcriptase (RT) fragments were sequenced. The patients were ARV naïve (n = 30), taking mother-to-child transmission prophylaxis (n = 50), or being treated with highly active ARV therapy/HAART (n = 18). The Stanford and IAS/USA databases and other sources were used to analyze PR/RT, gp41 env resistance mutations. The HIV-1 genetic diversity was analyzed by REGA/phylogenetic analyses. The patients' median age was 25 years (range, 16-42), 18.4% had AIDS. The frequency of natural resistance to T-20 (N42D, L44M, and R46M-low-impact mutations) was 6.1% (6/98); 20.4% (20/98) had compensatory mutations in HR2. The prevalence of transmitted drug resistance in the pol was 13.3% (4/30), and the prevalence of secondary drug resistance was 33.3% (6/18). Two patients were infected with multidrug resistant/MDR viruses. The analysis of HIV-1 subtypes (PR/RT/gp41) revealed that 61.2% (60/98) were subtype B, 12.2% (12/98) were subtype C, 4.1% (4/98) were subtype F1, and 22.4% (22/98) were possible recombinants (BF1 = 20.4%; BC = 2%). Natural resistance to T-20 was not associated with pol resistance or previous ARV use. The high rate of secondary resistance, including MDR, indicates that the number of women that may need T-20 salvage therapy may be higher than anticipated.

  8. Trimeric gp120-specific bovine monoclonal antibodies require cysteine and aromatic residues in CDRH3 for high affinity binding to HIV Env

    PubMed Central

    Center, Rob J.; Bebbington, Jonathan; Cuthbertson, Jack; Khoury, Georges; Lichtfuss, Marit; Rawlin, Grant; Purcell, Damian

    2017-01-01

    ABSTRACT We isolated HIV-1 Envelope (Env)-specific memory B cells from a cow that had developed high titer polyclonal immunoglobulin G (IgG) with broad neutralizing activity after a long duration vaccination with HIV-1AD8 Env gp140 trimers. We cloned the bovine IgG matched heavy (H) and light (L) chain variable (V) genes from these memory B cells and constructed IgG monoclonal antibodies (mAbs) with either a human constant (C)-region/bovine V-region chimeric or fully bovine C and V regions. Among 42 selected Ig+ memory B cells, two mAbs (6A and 8C) showed high affinity binding to gp140 Env. Characterization of both the fully bovine and human chimeric isoforms of these two mAbs revealed them as highly type-specific and capable of binding only to soluble AD8 uncleaved gp140 trimers and covalently stabilized AD8 SOSIP gp140 cleaved trimers, but not monomeric gp120. Genomic sequence analysis of the V genes showed the third heavy complementarity-determining region (CDRH3) of 6A mAb was 21 amino acids in length while 8C CDRH3 was 14 amino acids long. The entire V heavy (VH) region was 27% and 25% diverged for 6A and 8C, respectively, from the best matched germline V genes available, and the CDRH3 regions of 6A and 8C were 47.62% and 78.57% somatically mutated, respectively, suggesting a high level of somatic hypermutation compared with CDRH3 of other species. Alanine mutagenesis of the VH genes of 6A and 8C, showed that CDRH3 cysteine and tryptophan amino acids were crucial for antigen binding. Therefore, these bovine vaccine-induced anti-HIV antibodies shared some of the notable structural features of elite human broadly neutralizing antibodies, such as CDRH3 size and somatic mutation during affinity-maturation. However, while the 6A and 8C mAbs inhibited soluble CD4 binding to gp140 Env, they did not recapitulate the neutralizing activity of the polyclonal antibodies against HIV infection. PMID:27996375

  9. Roles of SLX1-SLX4, MUS81-EME1, and GEN1 in avoiding genome instability and mitotic catastrophe.

    PubMed

    Sarbajna, Shriparna; Davies, Derek; West, Stephen C

    2014-05-15

    The resolution of recombination intermediates containing Holliday junctions (HJs) is critical for genome maintenance and proper chromosome segregation. Three pathways for HJ processing exist in human cells and involve the following enzymes/complexes: BLM-TopoIIIα-RMI1-RMI2 (BTR complex), SLX1-SLX4-MUS81-EME1 (SLX-MUS complex), and GEN1. Cycling cells preferentially use the BTR complex for the removal of double HJs in S phase, with SLX-MUS and GEN1 acting at temporally distinct phases of the cell cycle. Cells lacking SLX-MUS and GEN1 exhibit chromosome missegregation, micronucleus formation, and elevated levels of 53BP1-positive G1 nuclear bodies, suggesting that defects in chromosome segregation lead to the transmission of extensive DNA damage to daughter cells. In addition, however, we found that the effects of SLX4, MUS81, and GEN1 depletion extend beyond mitosis, since genome instability is observed throughout all phases of the cell cycle. This is exemplified in the form of impaired replication fork movement and S-phase progression, endogenous checkpoint activation, chromosome segmentation, and multinucleation. In contrast to SLX4, SLX1, the nuclease subunit of the SLX1-SLX4 structure-selective nuclease, plays no role in the replication-related phenotypes associated with SLX4/MUS81 and GEN1 depletion. These observations demonstrate that the SLX1-SLX4 nuclease and the SLX4 scaffold play divergent roles in the maintenance of genome integrity in human cells. © 2014 Sarbajna et al.; Published by Cold Spring Harbor Laboratory Press.

  10. An anti-human immunodeficiency virus multiple antigen peptide encompassing the cleavage region of the env precursor interferes with membrane fusion at a post-CD4 binding step.

    PubMed

    Barbouche, R; Decroly, E; Kieny, M P; Fenouillet, E

    2000-07-20

    CLIV is a multiple antigen peptide ([PTKAKRRVVQREKR](4)-K(2)-K-betaA) that encompasses the cleavage region of the human immunodeficiency virus type 1 (HIV-1) envelope precursor. It displays an antiviral activity against HIV-1 and HIV-2 and inhibits HIV-1 Env-mediated cell-to-cell fusion. This effect has previously been attributed to interference with Env processing, resulting in the expression of a nonfusogenic envelope [Virology (1998) 247, 137]. However, we show here that CLIV does not alter the status of Env cleavage at steady state. Using various aggregation/syncytium assays that allow us to discriminate between gp120/CD4 binding and binding followed by gp41-mediated fusion, we demonstrate that CLIV inhibits a step of the cell-to-cell fusion process after CD4 binding. We demonstrate also that CLIV binds at 37 degrees C to a single class of protein present at the CD4(+) cell surface (Scatchard analysis: K(d) = 8 nM; B(max) = 10(4) sites/cell) and that the fusion inhibition activity seems to correlate with binding to this proteic component. In contrast, CLIV interacts with neither membrane-inserted nor CD4-associated Env. We therefore propose that CLIV interferes after Env/CD4 binding with a step of the membrane fusion process that may involve the C-terminal domain of gp120. Copyright 2000 Academic Press.

  11. Primate immune responses to HIV-1 Env formulated in the saponin-based adjuvant AbISCO-100 in the presence or absence of TLR9 co-stimulation

    PubMed Central

    Martinez, Paola; Sundling, Christopher; O'Dell, Sijy; Mascola, John R.; Wyatt, Richard T.; Karlsson Hedestam, Gunilla B.

    2015-01-01

    Protein-based vaccines require adjuvants to achieve optimal responses. Toll-like receptor (TLR) 9 agonists were previously shown to improve responses to protein-based vaccines, such as the Hepatitis B virus vaccine formulated in alum. Here, we used CpG-C together with the clinically relevant saponin-based adjuvant AbISCO-100/Matrix-M (AbISCO), to assess if TLR9 co-stimulation would quantitatively or qualitatively modulate HIV-1 envelope glycoprotein (Env)-specific B and T cell responses in rhesus macaques. The macaques were inoculated with soluble Env trimers in AbISCO, with or without the addition of CpG-C, using an interval similar to the Hepatitis B virus vaccine. Following a comprehensive evaluation of antigen-specific responses in multiple immune compartments, we show that the Env-specific circulating IgG, memory B cells and plasma cells displayed similar kinetics and magnitude in the presence or absence of CpG-C and that there was no apparent difference between the two groups in the elicited HIV-1 neutralizing antibody titers or antigen-specific CD4+ T cell responses. Importantly, the control of SHIV viremia was significantly improved in animals from both Env-immunized groups relative to adjuvant alone controls, demonstrating the potential of AbISCO to act as a stand-alone adjuvant for Env-based vaccines. PMID:25762407

  12. SEQ-GEN: A comprehensive multimission sequencing system

    NASA Technical Reports Server (NTRS)

    Salcedo, Jose; Starbird, Thomas J.

    1994-01-01

    SEQ-GEN is a user-interactive computer program used to plan and generate a sequence of commands for spacecraft. Desired activities are specified by the user of SEQ-GEN; SEQ-GEN in turn expands these activities, deriving the spacecraft commands necessary to accomplish the desired activities. SEQ-GEN models the effects on the spacecraft of the commands, predicting the state as a function of time, flagging any conflicts and rule violations. These states, conflicts, and violations are viewable both graphically and textually at the user's request. SEQ-GEN also displays the entire sequence graphically, showing each requested activity as a bar on its graphical timeline. SEQ-GEN immediately revalidates the sequence, updating its models and calculations along with its displays based on these changes. Because it has the ability to recalculate spacecraft states immediately, the user is able to perform 'what-if' sessions easily. SEQ-GEN, a multimission tool, is adaptable to any flight project. A flight project writes its adaptation files containing project unique information including in its simplest form, only spacecraft commands. For more involved projects the adaptation files may also contain flight and mission rules, description of the spacecraft and ground models, and the definition of activities. SEQ-GEN operates at whatever level of detail the adaptation files imply. Simple adaptations are straight forward to do. There is, however, no limit to the complexity of activity definitions or of spacecraft models: both may involve unlimited logical decision points. Commands and activities may involve any number of parameters of a wide variety of data types, including integer, float, time, boolean, and character strings. SEQ-GEN will be used by the Mars Pathfinder, Cassini, and VIM (Voyager Interstellar Mission) project in an effort to speed up adaptation time and to keep sequence generation costs down. SEQ-GEN is hosted on UNIX workstations. It uses MOTIF and X for windowing

  13. Taxonomy and phylogeny of Laburnicola gen. nov. and Paramassariosphaeria gen. nov. (Didymosphaeriaceae, Massarineae, Pleosporales).

    PubMed

    Wanasinghe, Dhanushka N; Jones, Evan Benjamin Gareth; Camporesi, Erio; Dissanayake, Asha J; Kamolhan, Sutakorn; Mortimer, Peter E; Xu, Jianchu; Abd-Elsalam, Kamel A; Hyde, Kevin D

    2016-11-01

    Laburnicola gen. nov., with four new species, Paramassariosphaeria gen. nov., with a new species and Montagnula saikhuensis sp. nov. are introduced in this paper. The phylogeny of these taxa and genera of Didymosphaeriaceae are resolved based on analysis of combined LSU, SSU, and ITS sequence data. Laburnicola is characterized by obpyriform, immersed ascomata, with a peridium fused to the host tissues, long pedicellate asci and ellipsoidal to fusoid ascospores, whose central cells have longitudinal septa and a rugged surface. Paramassariosphaeria is characterized by globose to subglobose ascomata, papillate ostioles, a peridium comprising pseudoparenchymatous cells of textura angularis, cylindric-clavate to clavate, pedicellate asci and brown, curved-fusoid ascospores, with multiple transverse septa and a mucilaginous sheath. Montagnula saikhuensis has immersed ascomata, with a peridium fused to the host tissues, long pedicellate asci and blackish brown, ellipsoidal ascospores with a single septum. The relationship of these taxa with other Didymosphaeriaceae species is discussed based on comparative morphology and phylogenetic analysis. Munkovalsaria appendiculata and Munkovalsaria donacina are transferred to the genus Montagnula.

  14. Reflexión bioética sobre el uso de organismos genéticamente modificados

    PubMed Central

    Yunta, Eduardo Rodríguez

    2011-01-01

    El presente artículo reflexiona desde los 4 principios de la bioética el uso comercial de organismos genéticamente modificados. Se cuestiona fundamentalmente la falta de transferencia de tecnología entre el mundo desarrollado y en desarrollo y el que el presente sistema de patentamiento de organismos vivos modificados fomenta intereses comerciales y no da debida importancia al desarrollo sostenible de la agricultura y ganadería en los países en desarrollo, donde más se necesita. Se reflexiona sobre la importancia que tiene evaluar los riesgos antes de introducirse en el mercado organismos genéticamente modificados y la necesidad de regulación en los países. PMID:21927675

  15. Cross-talk Suppression between the CpxA-CpxR and EnvZ-OmpR Two-Component Systems in E. coli

    PubMed Central

    Siryaporn, Albert; Goulian, Mark

    2009-01-01

    Many bacteria possess large numbers of two-component signaling systems, which are composed of histidine kinase-response regulator pairs. The high level of sequence similarity between some systems raises the possibility of undesired cross-talk between a histidine kinase and a non-cognate response regulator. Although molecular specificity ensures that phospho-transfer occurs primarily between correct partners, even a low level of inappropriate cross-talk could lead to unacceptable levels of noise or interference in signal transduction. To explore mechanisms that provide insulation against such interference, we have examined cross-talk between the histidine kinase CpxA and non-cognate response regulator OmpR in Escherichia coli. Our results show that there are two mechanisms that suppress cross-talk between these two proteins, which depend on the corresponding cognate partners CpxR and EnvZ and on the bifunctional nature of the histidine kinases CpxA and EnvZ. When cross-talk is detectable, we find it is independent of CpxA stimulus. We also show that cross-talk suppression leads to mutational robustness, i.e. it masks the effects of mutations that would otherwise lead to increased cross-talk. The mechanisms that provide insulation against interference described here may be applicable to many other two-component systems. PMID:18761686

  16. Different HIV-1 env frames: gp120 and ASP (antisense protein) biosynthesis, and theirs co-variation tropic amino acid signatures in X4- and R5-viruses.

    PubMed

    Dimonte, Salvatore

    2017-01-01

    Antisense protein (ASP) is the new actor of viral life of Human Immunodeficiency Virus type 1 (HIV-1) although proposed above 20 years ago. The asp ORF is into complementary strand of the gp120/gp41 junction of env gene. The ASP biological role remains little known. Knowing the Env markers of viral tropism, a dataset of sequences (660 strains) was used to analyze the hypothetical ASP involvement in CCR5 (R5) and/or CXCR4 (X4) co-receptor interaction. Preliminarily, prevalence of ASP and gp120V3 mutations was performed; following association among mutations were elaborate. The classical V3 tropic-signatures were confirmed, and 36 R5- and 22 X4-tropic ASP mutations were found. Moreover, by analyzing the ASP sequences, 36 out of 179 amino acid positions significantly associated with different co-receptor usage were found. Several statistically significant associations between gp120V3 and ASP mutations were observed. The dendrogram showed the existence of a cluster associated with R5-usage and a large cluster associated with X4-usage. These results show that gp120V3 and specific amino acid changes in ASP are associated together with CXCR4 and/or CCR5-usage. These findings implement previous observations on unclear ASP functions. J. Med. Virol. 89:112-122, 2017. © 2016 Wiley Periodicals, Inc.

  17. Virion Background and Efficiency of Virion Incorporation Determine Susceptibility of Simian Immunodeficiency Virus Env-Driven Viral Entry to Inhibition by IFITM Proteins

    PubMed Central

    Wrensch, Florian; Hoffmann, Markus; Gärtner, Sabine; Nehlmeier, Inga; Winkler, Michael

    2016-01-01

    ABSTRACT Interferon-induced transmembrane proteins (IFITMs) can inhibit the cellular entry of several enveloped viruses, including simian immunodeficiency virus (SIV). The blockade of SIV by IFITMs is isolate specific, raising the question of which parameters impact sensitivity to IFITM. We show that the virion context in which SIV-Env is presented and the efficiency of virion incorporation determine Env susceptibility to inhibition by IFITMs. Thus, determinants other than the nature of the envelope protein can impact the IFITM sensitivity of viral entry. IMPORTANCE The host cell-encoded IFITM proteins can block viral entry and are an important component of the innate defenses against viral infection. However, the determinants controlling whether a virus is susceptible to blockade by IFITM proteins are incompletely understood. Our study shows that the amount of envelope proteins incorporated into virions as well as the nature of the virion particle itself can impact the sensitivity of viral entry to IFITMs. These results show for the first time that determinants other than the viral envelope protein can impact sensitivity to IFITM and have implications for the interpretation of previously published data on inhibition of viruses by IFITM proteins. Moreover, our findings might help to define the mechanism underlying the antiviral activity of IFITM proteins. PMID:27807233

  18. Peptide Triazole Inactivators of HIV-1 Utilize a Conserved Two-Cavity Binding Site at the Junction of the Inner and Outer Domains of Env gp120

    PubMed Central

    Aneja, Rachna; Rashad, Adel A.; Li, Huiyuan; Sundaram, Ramalingam Venkat Kalyana; Duffy, Caitlin; Bailey, Lauren D.; Chaiken, Irwin

    2015-01-01

    We used coordinated mutagenesis, synthetic design, and flexible docking to investigate the structural mechanism of Env gp120 encounter by peptide triazole (PT) inactivators of HIV-1. Prior results demonstrated that the PT class of inhibitors suppresses binding at both CD4 and coreceptor sites on Env and triggers gp120 shedding, leading to cell-independent irreversible virus inactivation. Despite these enticing anti-HIV-1 phenotypes, structural understanding of the PT–gp120 binding mechanism has been incomplete. Here we found that PT engages two inhibitor ring moieties at the junction between the inner and outer domains of the gp120 protein. The results demonstrate how combined occupancy of two gp120 cavities can coordinately suppress both receptor and coreceptor binding and conformationally entrap the protein in a destabilized state. The two-cavity model has common features with small molecule gp120 inhibitor binding sites and provides a guide for further design of peptidomimetic HIV-1 inactivators based on the PT pharmacophore. PMID:25860784

  19. Identification of a new epitope for HIV-neutralizing antibodies in the gp41 membrane proximal external region by an Env-tailored phage display library.

    PubMed

    Zhou, Mingkui; Meyer, Torsten; Koch, Stefanie; Koch, Joachim; von Briesen, Hagen; Benito, José M; Soriano, Vincent; Haberl, Annette; Bickel, Markus; Dübel, Stefan; Hust, Michael; Dietrich, Ursula

    2013-02-01

    HIV controllers are a valuable source for the identification of HIV-neutralizing antibodies, as chronic infection over decades allows extensive affinity maturation of antibodies for improved Ag recognition. We analyzed a small cohort of elite controllers (ECs) for HIV-neutralizing antibodies using a panel of standardized HIV-1 pseudovirions on TZM-bl cells. An HIV-1 Env-tailored phage display library was generated to select epitopes targeted by neutralizing antibodies in the EC26 plasma sample showing the broadest neutralizing activity. Selected Env fragments were mostly allocated to the membrane proximal external region of gp41. After preabsorbing the EC26 plasma with the selected phage EC26-2A4, we achieved 50% depletion of its neutralizing activity. Furthermore, antibodies affinity-purified with the EC26-2A4 epitope from EC26 plasma showed neutralizing activity, proving that the selected phage indeed contains an epitope targeted by neutralizing plasma antibodies. Epitope fine mapping of the purified plasma antibodies on peptide arrays identified a new epitope overlapping, but clearly distinct, from the prominent 2F5 epitope. Of note, the purified antibodies did not show autoreactivity with cardiolipin, whereas low reactivity with phosphatidylserine comparable to mAb 2F5 was observed. Thus, this new epitope represents a promising candidate for further analysis in view of HIV vaccine development. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Structure-function analysis of the LytM domain of EnvC, an activator of cell wall remodeling at the Escherichia coli division site

    PubMed Central

    Peters, Nick T.; Morlot, Cécile; Yang, Desirée C.; Uehara, Tsuyoshi; Vernet, Thierry; Bernhardt, Thomas G.

    2013-01-01

    Proteins with LytM (Peptidase_M23) domains are broadly distributed in bacteria and have been implicated in a variety of important processes, including cell division and cell-shape determination. Most LytM-like proteins that have been structurally and/or biochemically characterized are metallo-endopeptidases that cleave crosslinks in the peptidoglycan (PG) cell wall matrix. Notable exceptions are the Escherichia coli cell division proteins EnvC and NlpD. These LytM factors are not hydrolases themselves, but instead serve as activators that stimulate PG cleavage by target enzymes called amidases to promote cell separation. Here we report the structure of the LytM domain from EnvC, the first structure of a LytM factor implicated in the regulation of PG hydrolysis. As expected, the fold is highly similar to that of other LytM proteins. However, consistent with its role as a regulator, the active site region is degenerate and lacks a catalytic metal ion. Importantly, genetic analysis indicates that residues in and around this degenerate active site are critical for amidase activation in vivo and in vitro. Thus, in the regulatory LytM factors, the apparent substrate binding pocket conserved in active metallo-endopeptidases has been adapted to control PG hydrolysis by another set of enzymes. PMID:23796240

  1. Fc Receptor-Mediated Activities of Env-Specific Human Monoclonal Antibodies Generated from Volunteers Receiving the DNA Prime-Protein Boost HIV Vaccine DP6-001.

    PubMed

    Costa, Matthew R; Pollara, Justin; Edwards, Regina Whitney; Seaman, Michael S; Gorny, Miroslaw K; Montefiori, David C; Liao, Hua-Xin; Ferrari, Guido; Lu, Shan; Wang, Shixia

    2016-11-15

    HIV-1 is able to elicit broadly potent neutralizing antibodies in a very small subset of individuals only after several years of infection, and therefore, vaccines that elicit these types of antibodies have been difficult to design. The RV144 trial showed that moderate protection is possible and that this protection may correlate with antibody-dependent cellular cytotoxicity (ADCC) activity. Our previous studies demonstrated that in an HIV vaccine phase I trial, the DP6-001 trial, a polyvalent Env DNA prime-protein boost formulation could elicit potent and broadly reactive, gp120-specific antibodies with positive neutralization activities. Here we report on the production and analysis of HIV-1 Env-specific human monoclonal antibodies (hMAbs) isolated from vaccinees in the DP6-001 trial. For this initial report, 13 hMAbs from four vaccinees in the DP6-001 trial showed broad binding to gp120 proteins of diverse subtypes both autologous and heterologous to vaccine immunogens. Equally cross-reactive Fc receptor-mediated functional activities, including ADCC and antibody-dependent cellular phagocytosis (ADCP) activities, were present with both immune sera and isolated MAbs, confirming the induction of nonneutralizing functional hMAbs by the DNA prime-protein boost vaccination. Elicitation of broadly reactive hMAbs by vaccination in healthy human volunteers confirms the value of the polyvalent formulation in this HIV vaccine design.

  2. Co-regulation of polysaccharide production, motility, and expression of type III secretion genes by EnvZ/OmpR and GrrS/GrrA systems in Erwinia amylovora.

    PubMed

    Li, Wenting; Ancona, Veronica; Zhao, Youfu

    2014-02-01

    The EnvZ/OmpR and GrrS/GrrA systems, two widely distributed two-component systems in gamma-Proteobacteria, negatively control amylovoran biosynthesis in Erwinia amylovora, and the two systems regulate motility in an opposing manner. In this study, we examined the interplay of EnvZ/OmpR and GrrS/GrrA systems in controlling various virulence traits in E. amylovora. Results showed that amylovoran production was significantly higher when both systems were inactivated, indicating that the two systems act as negative regulators and their combined effect on amylovoran production appears to be enhanced. In contrast, reduced motility was observed when both systems were deleted as compared to that of grrA/grrS mutants and WT strain, indicating that the two systems antagonistically regulate motility in E. amylovora. In addition, glycogen accumulation was much higher in envZ/ompR and two triple mutants than that of grrS/grrA mutants and WT strain, suggesting that EnvZ/OmpR plays a dominant role in regulating glycogen accumulation, whereas levan production was significantly lower in the grrS/grrA and two triple mutants as compared with that of WT and envZ/ompR mutants, indicating that GrrS/GrrA system dominantly controls levan production. Furthermore, both systems negatively regulated expression of three type III secretion (T3SS) genes and their combined negative effect on hrp-T3SS gene expression increased when both systems were deleted. These results demonstrated that EnvZ/OmpR and GrrS/GrrA systems co-regulate various virulence factors in E. amylovora by still unknown mechanisms or through different target genes, sRNAs, or proteins, indicating that a complex regulatory network may be involved, which needs to be further explored.

  3. Viable but non-culturable state (VBNC) of Escherichia coli related to EnvZ under the effect of pH, starvation and osmotic stress in sea water.

    PubMed

    Darcan, Cihan; Ozkanca, Reşit; Idil, Onder; Flint, Ken P

    2009-01-01

    When exposed extreme environmental conditions such as sea water, bacteria have been shown different survival strategy for continue their life. One of this strategy known as viable but nonculturable (VBNC) state which is very important for nondifferiation bacteria. VBNC cells cause serious human health problems. Little is known, however, about the genetic mechanisms underlying the VBNC state. Under different environmental conditions, porins are important in the survival strategy of bacteria. EnvZ/OmpR work together as regulators of ompF and ompC gene expression. It is known that the EnvZ system has a role in VBNC state. In this study we tried to find out the viability of EnvZ, OmpC and OmpF mutant E. coli under stress effect of osmolarity, pH and starvation. Bacteria were suspended in filtered-autoclaved sea water microcosms and numbers determined over 25 day incubation periods by plate count (PC), direct viable count (DVC) and count of cells capable of respiration (RCC). As regard to results, alkaline pH affected E. coli more than acidic pH, which led to decline in number. On the contrary glycine betaine addition to sea water protected E. coli porin mutants and also reduced the death rate of bacteria. Under the effect of pH, osmotic stress and starvation stress, wild type E. coli and porin mutants entered a dormant state or became VBNC with the exception of MSZ31 (envZ mutant) E. coli cells which did not enter the VBNC state under the three tested stress conditions. This study is the first report to demonstrate that E. coli could not enter the VBNC state in the lack of EnvZ product under the stress of osmolarity, pH and starvation and the relationship between EnvZ and VBNC state are not affected by pH, osmolarity and starvation.

  4. Human Endogenous Retrovirus K(HML-2) Gag- and Env-Specific T-Cell Responses Are Infrequently Detected in HIV-1-Infected Subjects Using Standard Peptide Matrix-Based Screening

    PubMed Central

    John, Vivek M.; Hunter, Diana V.; Martin, Eric; Mujib, Shariq; Mihajlovic, Vesna; Burgers, Peter C.; Luider, Theo M.; Gyenes, Gabor; Sheppard, Neil C.; SenGupta, Devi; Tandon, Ravi; Yue, Feng-Yun; Benko, Erika; Kovacs, Colin; Nixon, Douglas F.; Ostrowski, Mario A.

    2012-01-01

    T-cell responses to human endogenous retrovirus (HERV) K(HML-2) Gag and Env were mapped in HIV-1-infected subjects using 15mer peptides. Small peptide pools and high concentrations were used to maximize sensitivity. In the 23 subjects studied, only three bona fide HERV-K(HML-2)-specific responses were detected. At these high peptide concentrations, we detected false-positive responses, three of which were mapped to an HIV-1 Gag peptide contaminant. Thus, HERV-K(HML-2) Gag- and Env-specific T-cell responses are infrequently detected by 15mer peptide mapping. PMID:22205657

  5. GenAnneal: Genetically modified Simulated Annealing

    NASA Astrophysics Data System (ADS)

    Tsoulos, Ioannis G.; Lagaris, Isaac E.

    2006-05-01

    A modification of the standard Simulated Annealing (SA) algorithm is presented for finding the global minimum of a continuous multidimensional, multimodal function. We report results of computational experiments with a set of test functions and we compare to methods of similar structure. The accompanying software accepts objective functions coded both in Fortran 77 and C++. Program summaryTitle of program:GenAnneal Catalogue identifier:ADXI_v1_0 Program summary URL:http://cpc.cs.qub.ac.uk/summaries/ADXI_v1_0 Program available from: CPC Program Library, Queen's University of Belfast, N. Ireland Computer for which the program is designed and others on which it has been tested: The tool is designed to be portable in all systems running the GNU C++ compiler Installation: University of Ioannina, Greece on Linux based machines Programming language used:GNU-C++, GNU-C, GNU Fortran 77 Memory required to execute with typical data: 200 KB No. of bits in a word: 32 No. of processors used: 1 Has the code been vectorized or parallelized?: No No. of bytes in distributed program, including test data, etc.:84 885 No. of lines in distributed program, including test data, etc.:14 896 Distribution format: tar.gz Nature of physical problem: A multitude of problems in science and engineering are often reduced to minimizing a function of many variables. There are instances that a local optimum does not correspond to the desired physical solution and hence the search for a better solution is required. Local optimization techniques are frequently trapped in local minima. Global optimization is hence the appropriate tool. For example, solving a non-linear system of equations via optimization, employing a "least squares" type of objective, one may encounter many local minima that do not correspond to solutions (i.e. they are far from zero). Typical running time: Depending on the objective function. Method of solution: We modified the process of step selection that the traditional Simulated

  6. Genes and proteins of Escherichia coli (GenProtEc).

    PubMed

    Riley, M; Space, D B

    1996-01-01

    GenProtEc is a database of Escherichia coli genes and their gene products, classified by type of function and physiological role and with citations to the literature for each. Also present are data on sequence similarities among E.coli proteins with PAM values, percent identity of amino acids, length of alignment and percent aligned. The database is available as a PKZip file by ftp from mbl.edu/pub/ecoli.exe. The program runs under MS-DOS on IMB-compatible machines. GenProtEc can also be accessed through the World Wide Web at URL http://mbl.edu/html/ecoli.html.

  7. Genes and proteins of Escherichia coli (GenProtEc).

    PubMed Central

    Riley, M; Space, D B

    1996-01-01

    GenProtEc is a database of Escherichia coli genes and their gene products, classified by type of function and physiological role and with citations to the literature for each. Also present are data on sequence similarities among E.coli proteins with PAM values, percent identity of amino acids, length of alignment and percent aligned. The database is available as a PKZip file by ftp from mbl.edu/pub/ecoli.exe. The program runs under MS-DOS on IMB-compatible machines. GenProtEc can also be accessed through the World Wide Web at URL http://mbl.edu/html/ecoli.html. PMID:8594596

  8. Dural tear post mastoidectomy repaired with Dura Gen.

    PubMed

    Wong, C Y; Khairi, M D M; Mohamed, S A; Irfan, M

    2010-12-01

    Dural exposure may occur during the course of thinning the tegmen tympani and tegmen mastoideum in mastoid procedure. If large area of dura is exposed or lacerated, cerebrospinal fluid and brain herniation may enter the mastoid cavity. We report a case of a patient with injured dura mater and tegmen mastoideum during mastoidectomy for chronic suppurative otitis media with cholesteatoma managed by using DuraGen. The dura mater and tegmen defect healed totally showing the success of the procedure. A collagen matrix like DuraGen is an option for repairing dural tear in mastoid region.

  9. Introducing AstroGen: the Astronomy Genealogy Project

    NASA Astrophysics Data System (ADS)

    Tenn, Joseph S.

    2016-12-01

    The Astronomy Genealogy Project (AstroGen), a project of the Historical Astronomy Division of the American Astronomical Society (AAS), will soon appear on the AAS website. Ultimately, it will list the world's astronomers with their highest degrees, theses for those who wrote them, academic advisors (supervisors), universities, and links to the astronomers or their obituaries, their theses when online, and more. At present the AstroGen team is working on those who earned doctorates with astronomy-related theses. We show what can be learned already, with just ten countries essentially completed.

  10. Identification of three feline immunodeficiency virus (FIV) env gene subtypes and comparison of the FIV and human immunodeficiency virus type 1 evolutionary patterns.

    PubMed Central

    Sodora, D L; Shpaer, E G; Kitchell, B E; Dow, S W; Hoover, E A; Mullins, J I

    1994-01-01

    Feline immunodeficiency virus (FIV) is a lentivirus associated with AIDS-like illnesses in cats. As such, FIV appears to be a feline analog of human immunodeficiency virus (HIV). A hallmark of HIV infection is the large degree of viral genetic diversity that can develop within an infected individual and the even greater and continually increasing level of diversity among virus isolates from different individuals. Our goal in this study was to determine patterns of FIV genetic diversity by focusing on a 684-nucleotide region encompassing variable regions V3, V4, and V5 of the FIV env gene in order to establish parallels and distinctions between FIV and HIV type 1 (HIV-1). Our data demonstrate that, like HIV-1, FIV can be separated into distinct envelope sequence subtypes (three are described here). Similar to that found for HIV-1, the pairwise sequence divergence within an FIV subtype ranged from 2.5 to 15.0%, whereas that between subtypes ranged from 17.8 to 26.2%. However, the high number of synonymous nucleotide changes among FIV V3 to V5 env sequences may also include a significant number of back mutations and suggests that the evolutionary distances among FIV subtypes are underestimated. Although only a few subtype B viruses were available for examination, the pattern of diversity between the FIV A and B subtypes was found to be significantly distinct; subtype B sequences had proportionally fewer mutations that changed amino acids, compared with silent changes, suggesting a more advanced state of adaptation to the host. No similar distinction was evident for HIV-1 subtypes. The diversity of FIV genomes within individual infected cats was found to be as high as 3.7% yet twofold lower than that within HIV-1-infected people over a comparable region of the env gene. Despite these differences, significant parallels between patterns of FIV evolution and HIV-1 evolution exist, indicating that a wide array of potentially divergent virus challenges need to be considered

  11. Recombination between feline leukemia virus subgroup B or C and endogenous env elements alters the in vitro biological activities of the viruses.

    PubMed Central

    Pandey, R; Ghosh, A K; Kumar, D V; Bachman, B A; Shibata, D; Roy-Burman, P

    1991-01-01

    An important question in feline leukemia virus (FeLV) pathogenesis is whether, as in murine leukemia virus infection, homologous recombination between the infecting FeLV and the noninfectious endogenous FeLV-like proviruses serves as a significant base for the generation of proximal pathogens. To begin an analysis of this issue, several recombinant FeLVs were produced by using two different approaches: (i) the regions of the viral envelope (env) gene of a cloned FeLV (subgroup B virus [FeLV-B], Gardner-Arnstein strain) and those of two different endogenous proviral loci were exchanged to create specific FeLV chimeras, and (ii) vectors containing endogenous env and molecularly cloned infectious FeLV-C (Sarma strain) DNA sequences were coexpressed by transfection in nonfeline cells to facilitate recombination. The results of these combined approaches showed that up to three-fourths of the envelope glycoprotein (gp70), beginning from the N-terminal end, could be replaced by endogenous FeLV sequences to produce biologically active chimeric FeLVs. The in vitro replication efficiency or cell tropism of the recombinants appeared to be influenced by the amount of gp70 sequences replaced by the endogenous partner as well as by the locus of origin of the endogenous sequences. Additionally, a characteristic biological effect, aggregation of feline T-lymphoma cells (3201B cell line), was found to be specifically induced by replicating FeLV-C or FeLV-C-based recombinants. Multiple crossover sites in the gp70 protein selected under the conditions used for coexpression were identified. The results of induced coexpression were also supported by rapid generation of FeLV recombinants when FeLV-C was used to infect the feline 3201B cell line that constitutively expresses high levels of endogenous FeLV-specific mRNAs. Furthermore, a large, highly conserved open reading frame in the pol gene of an endogenous FeLV provirus was identified. This observation, particularly in reference to

  12. Viability of thin wall tube forming of ATF FeCrAl

    SciTech Connect

    Maloy, Stuart Andrew; Aydogan, Eda; Anderoglu, Osman; Lavender, Curt; Yamamoto, Yukinori

    2016-09-16

    Fabrication of thin walled tubing of FeCrAl alloys is critical to its success as a candidate enhanced accident-tolerant fuel cladding material. Alloys that are being investigated are Generation I and Generation II FeCrAl alloys produced at ORNL and an ODS FeCrAl alloy, MA-956 produced by Special Metals. Gen I and Gen II FeCrAl alloys were provided by ORNL and MA-956 was provided by LANL (initially produced by Special Metals). Three tube development efforts were undertaken. ORNL led the FeCrAl Gen I and Gen II alloy development and tube processing studies through drawing tubes at Rhenium Corporation. LANL received alloys from ORNL and led tube processing studies through drawing tubes at Century Tubing. PNNL led the development of tube processing studies on MA-956 through pilger processing working with Sandvik Corporation. A summary of the recent progress on tube development is provided in the following report and a separate ORNL report: ORNL/TM-2015/478, “Development and Quality Assessments of Commercial Heat Production of ATF FeCrAl Tubes”.

  13. GenR, an IclR-Type Regulator, Activates and Represses the Transcription of gen Genes Involved in 3-Hydroxybenzoate and Gentisate Catabolism in Corynebacterium glutamicum

    PubMed Central

    Chao, Hongjun

    2013-01-01

    The genes required for 3-hydroxybenzoate and gentisate catabolism in Corynebacterium glutamicum are closely clustered in three operons. GenR, an IclR-type regulator, can activate the transcription of genKH and genDFM operons in response to 3-hydroxybenzoate and gentisate, and it can repress its own expression. Footprinting analyses demonstrated that GenR bound to four sites with different affinities. Two GenR-binding sites (DFMn01 and DFMn02) were found to be located between positions −41 and −84 upstream of the −35 and −10 regions of the genDFM promoter, which was involved in positive regulation of genDFM transcription. The GenR binding site R-KHn01 (located between positions −47 and −16) overlapped the −35 region of the genKH promoter sequence and is involved in positive regulation of its transcription. The binding site R-KHn02, at which GenR binds to its own promoter, was found within a footprint extending from position −44 to −67. It appeared to be involved in negative regulation of the activity of the genR promoter. A consensus motif with a 5-bp imperfect palindromic sequence [ATTCC-N7(5)-GGAAT] was identified among all four GenR binding sites and found to be necessary to GenR regulation through site-directed mutagenesis. The results reveal a new regulatory function of the IclR family in the catabolism of aromatic compounds. PMID:23354754

  14. A Virtual Reality Framework to Optimize Design, Operation and Refueling of GEN-IV Reactors.

    SciTech Connect

    Rizwan-uddin; Nick Karancevic; Stefano Markidis; Joel Dixon; Cheng Luo; Jared Reynolds

    2008-04-23

    many GEN-IV candidate designs are currently under investigation. Technical issues related to material, safety and economics are being addressed at research laboratories, industry and in academia. After safety, economic feasibility is likely to be the most important crterion in the success of GEN-IV design(s). Lessons learned from the designers and operators of GEN-II (and GEN-III) reactors must play a vital role in achieving both safety and economic feasibility goals.

  15. HIV-specific humoral and cellular immunity in rabbits vaccinated with recombinant human immunodeficiency virus-like gag-env particles

    SciTech Connect

    Haffar, O.K.; Smithgall, M.D.; Moran, P.A.; Travis, B.M.; Zarling, J.M.; Hu, S.L. )

    1991-08-01

    Recombinant human immunodeficiency virus type-1 (HIV-1)-like gag-env particles produced in mammalian cells were inoculated into two New Zealand white rabbits. In parallel, two control rabbits were inoculated with the homologous HIV-1 virions inactivated by ultraviolet light (uv) and psoralen treatments. The humoral and cellular immune responses to HIV-1 were evaluated for both groups of animals. Recombinant particles elicited humoral immunity that was specific for all the viral structural proteins. The antibodies recognized both denatured and nondenatured proteins. Moreover, the sera neutralized the in vitro infectivity of the homologous virus in CEM cells. Importantly, the recombinant particles also generated a T helper response by priming with the HIV proteins. Similar results were observed with inactivated virus immunization. Therefore, the authors results suggest that the recombinant HIV-like particles elicit functional humoral immunity as well as cellular immunity and represent a novel vaccine candidate for AIDS.

  16. Reduced duration of breastfeeding is associated with a higher risk of multiple sclerosis in both Italian and Norwegian adult males: the EnvIMS study.

    PubMed

    Ragnedda, Giammario; Leoni, Stefania; Parpinel, Maria; Casetta, Ilaria; Riise, Trond; Myhr, Kjell-Morten; Wolfson, Christina; Pugliatti, Maura

    2015-05-01

    Breastfeeding for at least 4 months has been found to be associated with a reduced risk of immune-mediated diseases including multiple sclerosis (MS). Using data from a large multinational case-control study (EnvIMS), the association between MS and breastfeeding was investigated in two distinct populations. A questionnaire (EnvIMS-Q) which included a section on feeding during the first year of life was administered to MS cases and to age and sex frequency-matched controls from Italy and Norway. Logistic regression was used to estimate the odds ratio (ORs) and 95% confidence intervals (95% CIs) as a measure of the association between MS and exposure to prolonged breastfeeding (4 months or more, used as the reference category), vs. no breastfeeding or breastfeeding for less than 4 months (reduced exposure). Education, smoking habits, smoking in mother's pregnancy, and other types of milk used in infant feeding were included as covariates. A total of 547 cases and 1039 controls in Italy, and 737 cases and 1335 controls in Norway were studied. The distribution of prolonged (reference) breastfeeding differed between the Norwegian (65.4%) and the Italian (48.9%) study participants. A significant association between MS and reduced/no exposure to breastfeeding was found overall for Italy (OR(adj) = 1.37; 95% CI 1.09, 1.73), but not for Norway (OR(adj) = 1.14; 95% CI 0.92, 1.40). However, only in men, significant associations were observed for both populations (OR(Italy) = 2.33; 95% CI 1.50, 3.65, OR(Norway) = 2.13; 95% CI 1.37, 3.30). Reduced exposure to breastfeeding in males was found to be associated with increased risk of MS in Italy and in Norway.

  17. The Intracytoplasmic Domain of the Env Transmembrane Protein Is a Locus for Attenuation of Simian Immunodeficiency Virus SIVmac in Rhesus Macaques

    PubMed Central

    Shacklett, Barbara L.; Weber, Claudia Jo; Shaw, Karen E. S.; Keddie, Elise M.; Gardner, Murray B.; Sonigo, Pierre; Luciw, Paul A.

    2000-01-01

    The human and simian immunodeficiency virus (HIV-1 and SIVmac) transmembrane proteins contain unusually long intracytoplasmic domains (ICD-TM). These domains are suggested to play a role in envelope fusogenicity, interaction with the viral matrix protein during assembly, viral infectivity, binding of intracellular calmodulin, disruption of membranes, and induction of apoptosis. Here we describe a novel mutant virus, SIVmac-M4, containing multiple mutations in the coding region for the ICD-TM of pathogenic molecular clone SIVmac239. Parental SIVmac239-Nef+ produces high-level persistent viremia and simian AIDS in both juvenile and newborn rhesus macaques. The ICD-TM region of SIVmac-M4 contains three stop codons, a +1 frameshift, and mutation of three highly conserved, charged residues in the conserved C-terminal alpha-helix referred to as lentivirus lytic peptide 1 (LLP-1). Overlapping reading frames for tat, rev, and nef are not affected by these changes. In this study, four juvenile macaques received SIVmac-M4 by intravenous injection. Plasma viremia, as measured by branched-DNA (bDNA) assay, reached a peak at 2 weeks postinoculation but dropped to below detectable levels by 12 weeks. At over 1.5 years postinoculation, all four juvenile macaques remain healthy and asymptomatic. In a subsequent experiment, four neonatal rhesus macaques were given SIVmac-M4 intravenously. These animals exhibited high levels of viremia in the acute phase (2 weeks postinoculation) but are showing a relatively low viral load in the chronic phase of infection, with no clinical signs of disease for 1 year. These findings demonstrated that the intracytoplasmic domain of the transmembrane Env (Env-TM) is a locus for attenuation in rhesus macaques. PMID:10846063

  18. Reduced potency and incomplete neutralization of broadly neutralizing antibodies against cell-to-cell transmission of HIV-1 with transmitted founder Envs.

    PubMed

    Li, Hongru; Zony, Chati; Chen, Ping; Chen, Benjamin K

    2017-02-01

    Broadly neutralizing antibodies (bNAbs) have been isolated from HIV-1 patients and can potently block infection of a wide spectrum of HIV-1 subtypes. These antibodies define common epitopes shared by many viral isolates. While bNAbs potently antagonize infection with cell-free virus, inhibition of HIV-1 transmission from infected to uninfected CD4(+) T cells through virological synapses (VS), has been found to require greater amounts of antibody. In this study, we examined two well-studied molecular clones and two transmitted founder (T/F) viruses for their sensitivities to a panel of bNAbs in cell-free and cell-to-cell infection assays. We observed a relative resistance of cell-to-cell transmission to antibody neutralization that is reflected not only by reductions of antibody potency, but also by decreases in maximum neutralization capacity relative to cell-free infections. BNAbs targeting different epitopes exhibited incomplete neutralization against cell-associated virus with T/F Envs, which was not observed with cell-free form of the same virus. We further identified the membrane proximal internal tyrosine-based sorting motif as a determinant that can affect the incomplete neutralization of these T/F clones in cell-to-cell infection. These findings indicate that the signal that affects surface expression and/or internalization of Env from the plasma membrane can modulate the presentation of neutralizing epitopes on infected cells. These findings highlight that a fraction of virus can escape from high concentrations of antibody through cell-to-cell infection while maintaining sensitivity to neutralization in cell-free infection. The ability to fully inhibit cell-to-cell transmission may represent an important consideration in development of antibodies for treatment or prophylaxis.

  19. Mutations of a residue within the polyproline-rich region of Env alter the replication rate and level of cytopathic effects in chimeric avian retroviral vectors.

    PubMed

    Chang, Kevin W; Barsov, Eugene V; Ferris, Andrea L; Hughes, Stephen H

    2005-08-01

    Previous attempts to extend the host range of the avian sarcoma/leukosis virus (ASLV)-based RCASBP vectors produced two viral vectors, RCASBP M2C (4070A) and RCASBP M2C (797-8), which replicate using the amphotropic murine leukemia virus 4070A Env protein (2). Both viruses were adapted to replicate efficiently in the avian cell line DF-1, but RCASBP M2C (4070A) caused extensive cytopathic effects (CPE) in DF-1 cells whereas RCASBP M2C (797-8) induced low levels of CPE. The two viruses differed only at amino acid 242 of the polyproline-rich region in the surface (SU) subunit of the Env protein. In RCASBP M2C (4070A), an isoleucine replaced the wild-type proline residue, whereas a threonine residue was found in RCASBP M2C (797-8). In the present study, we show that other amino acid substitutions at position 242 strongly influence the CPE and replication rate of the chimeric viruses. There was a correlation between the amount of unintegrated linear retroviral DNA present in infected DF-1 cells and the level of CPE. This suggests that there may be a role for superinfection in the CPE. The treatment of RCASBP M2C (4070A)-infected cells with dantrolene, which inhibits the release of calcium from the endoplasmic reticulum (ER), reduced the amount of CPE seen during infection with the highly cytotoxic virus. Dantrolene treatment did not appear to affect virus production, suggesting that Ca2+ release from the ER had a role in the CPE caused by these viruses.

  20. Design and structure of two HIV-1 clade C SOSIP.664 trimers that increase the arsenal of native-like Env immunogens.

    PubMed

    Julien, Jean-Philippe; Lee, Jeong Hyun; Ozorowski, Gabriel; Hua, Yuanzi; Torrents de la Peña, Alba; de Taeye, Steven W; Nieusma, Travis; Cupo, Albert; Yasmeen, Anila; Golabek, Michael; Pugach, Pavel; Klasse, P J; Moore, John P; Sanders, Rogier W; Ward, Andrew B; Wilson, Ian A

    2015-09-22

    A key challenge in the quest toward an HIV-1 vaccine is design of immunogens that can generate a broadly neutralizing antibody (bnAb) response against the enormous sequence diversity of the HIV-1 envelope glycoprotein (Env). We previously demonstrated that a recombinant, soluble, fully cleaved SOSIP.664 trimer based on the clade A BG505 sequence is a faithful antigenic and structural mimic of the native trimer in its prefusion conformation. Here, we sought clade C native-like trimers with comparable properties. We identified DU422 and ZM197M SOSIP.664 trimers as being appropriately thermostable (Tm of 63.4 °C and 62.7 °C, respectively) and predominantly native-like, as determined by negative-stain electron microscopy (EM). Size exclusion chromatography, ELISA, and surface plasmon resonance further showed that these trimers properly display epitopes for all of the major bnAb classes, including quaternary-dependent, trimer-apex (e.g., PGT145) and gp120/gp41 interface (e.g., PGT151) epitopes. A cryo-EM reconstruction of the ZM197M SOSIP.664 trimer complexed with VRC01 Fab against the CD4 binding site at subnanometer resolution revealed a striking overall similarity to its BG505 counterpart with expected local conformational differences in the gp120 V1, V2, and V4 loops. These stable clade C trimers contribute additional diversity to the pool of native-like Env immunogens as key components of strategies to induce bnAbs to HIV-1.

  1. 78 FR 54509 - Tenth Meeting: RTCA Next Gen Advisory Committee (NAC)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-09-04

    ... Federal Aviation Administration Tenth Meeting: RTCA Next Gen Advisory Committee (NAC) AGENCY: Federal Aviation Administration (FAA), U.S. Department of Transportation (DOT). ACTION: Notice of RTCA NextGen... of the RTCA NextGen Advisory Committee (NAC). DATES: The meeting will be held September 19, 2013...

  2. 77 FR 25524 - Sixth Meeting: RTCA, NextGen Advisory Committee

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-04-30

    ... Federal Aviation Administration Sixth Meeting: RTCA, NextGen Advisory Committee AGENCY: Federal Aviation Administration (FAA), U.S. Department of Transportation (DOT). ACTION: Notice of a meeting of RTCA, NextGen... RTCA, NextGen Advisory Committee. DATES: The meeting will be held May 24, 2012, from 9:00 a.m.-3:00...

  3. 78 FR 5860 - Eighth Meeting: RTCA Next Gen Advisory Committee (NAC)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-01-28

    ... Federal Aviation Administration Eighth Meeting: RTCA Next Gen Advisory Committee (NAC) AGENCY: Federal Aviation Administration (FAA), U.S. Department of Transportation (DOT). ACTION: Notice of RTCA NextGen... meeting of the RTCA NextGen Advisory Committee (NAC). DATES: The meeting will be held February 7,...

  4. 76 FR 54526 - Fourth Meeting RTCA NextGen Advisory Committee (NAC)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-09-01

    ... Federal Aviation Administration Fourth Meeting RTCA NextGen Advisory Committee (NAC) AGENCY: Federal Aviation Administration (FAA), DOT. ACTION: RTCA NextGen Advisory Committee (NAC). SUMMARY: The FAA is issuing this notice to advise the public of a meeting of RTCA NextGen Advisory Committee (NAC). DATES:...

  5. 76 FR 3931 - Second Meeting RTCA NextGen Advisory Committee (NAC)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-01-21

    ... Federal Aviation Administration Second Meeting RTCA NextGen Advisory Committee (NAC) AGENCY: Federal Aviation Administration (FAA), DOT. ACTION: RTCA NextGen Advisory Committee (NAC). SUMMARY: The FAA is issuing this notice to advise the public of a meeting of RTCA NextGen Advisory Committee (NAC). DATES:...

  6. Substrate preference of Gen endonucleases highlights the importance of branched structures as DNA damage repair intermediates

    PubMed Central

    Bellendir, Stephanie P.; Rognstad, Danielle J.; Morris, Lydia P.; Zapotoczny, Grzegorz; Walton, William G.; Redinbo, Matthew R.; Ramsden, Dale A.

    2017-01-01

    Abstract Human GEN1 and yeast Yen1 are endonucleases with the ability to cleave Holliday junctions (HJs), which are proposed intermediates in recombination. In vivo, GEN1 and Yen1 function secondarily to Mus81, which has weak activity on intact HJs. We show that the genetic relationship is reversed in Drosophila, with Gen mutants having more severe defects than mus81 mutants. In vitro, DmGen, like HsGEN1, efficiently cleaves HJs, 5΄ flaps, splayed arms, and replication fork structures. We find that the cleavage rates for 5΄ flaps are significantly higher than those for HJs for both DmGen and HsGEN1, even in vast excess of enzyme over substrate. Kinetic studies suggest that the difference in cleavage rates results from a slow, rate-limiting conformational change prior to HJ cleavage: formation of a productive dimer on the HJ. Despite the stark difference in vivo that Drosophila uses Gen over Mus81 and humans use MUS81 over GEN1, we find the in vitro activities of DmGen and HsGEN1 to be strikingly similar. These findings suggest that simpler branched structures may be more important substrates for Gen orthologs in vivo, and highlight the utility of using the Drosophila model system to further understand these enzymes. PMID:28369583

  7. Substrate preference of Gen endonucleases highlights the importance of branched structures as DNA damage repair intermediates.

    PubMed

    Bellendir, Stephanie P; Rognstad, Danielle J; Morris, Lydia P; Zapotoczny, Grzegorz; Walton, William G; Redinbo, Matthew R; Ramsden, Dale A; Sekelsky, Jeff; Erie, Dorothy A

    2017-05-19

    Human GEN1 and yeast Yen1 are endonucleases with the ability to cleave Holliday junctions (HJs), which are proposed intermediates in recombination. In vivo, GEN1 and Yen1 function secondarily to Mus81, which has weak activity on intact HJs. We show that the genetic relationship is reversed in Drosophila, with Gen mutants having more severe defects than mus81 mutants. In vitro, DmGen, like HsGEN1, efficiently cleaves HJs, 5΄ flaps, splayed arms, and replication fork structures. We find that the cleavage rates for 5΄ flaps are significantly higher than those for HJs for both DmGen and HsGEN1, even in vast excess of enzyme over substrate. Kinetic studies suggest that the difference in cleavage rates results from a slow, rate-limiting conformational change prior to HJ cleavage: formation of a productive dimer on the HJ. Despite the stark difference in vivo that Drosophila uses Gen over Mus81 and humans use MUS81 over GEN1, we find the in vitro activities of DmGen and HsGEN1 to be strikingly similar. These findings suggest that simpler branched structures may be more important substrates for Gen orthologs in vivo, and highlight the utility of using the Drosophila model system to further understand these enzymes. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  8. GenStar: A method for de novo drug design

    NASA Astrophysics Data System (ADS)

    Rotstein, Sergio H.; Murcko, Mark A.

    1993-02-01

    A novel method, which we call GenStar, has been developed to suggest chemically reasonable structures which fill the active sites of enzymes. The proposed molecules provide good steric contact with the enzyme and exist in low-energy conformations. These structures are composed entirely of sp3 carbons which are grown sequentially, but which can also branch or form rings. User-selected enzyme seed atoms may be used to determine the area in which structure generation begins. Alternatively, GenStar may begin with a predocked `inhibitor core' from which atoms are grown. For each new atom generated by the program, several hundred candidate positions representing a range of reasonable bond lengths, bond angles, and torsion angles are considered. Each of these candidates is scored, based on a simple enzyme contact model. The selected position is chosen at random from among the highest scoring cases. Duplicate structures may be removed using a variety of criteria. The compounds may be energy minimized and displayed using standard modeling programs. Also, it is possible to analyze the collection of all structures created by GenStar and locate binding motifs for common fragments such as benzene and naphthylene. Tests of the method using HIV protease, FK506 binding protein (FKBP-12) and human carbonic anhydrase (HCA-II) demonstrated that structures similar to known potent inhibitors may be generated with GenStar.

  9. A New Parent Generation: Meet Mr. and Mrs. Gen X

    ERIC Educational Resources Information Center

    Howe, Neil

    2010-01-01

    Slowly but surely, Generation Xers have been taking over from Baby Boomers as the majority of parents in elementary and secondary education. Gen-X parents and Boomer parents belong to two neighboring generations, each possessing its own location in history and its own peer personality. They are similar in some respects, but clearly different in…

  10. ORPC RivGen controller performance raw data - Igiugig 2015

    DOE Data Explorer

    Jarlath McEntee

    2015-12-18

    Contains raw data for operations of Ocean Renewable Power Company (ORPC) RivGen Power System in Igiugig 2015 in Matlab data file format. Two data files capture the data and timestamps for data, including power in, voltage, rotation rate, and velocity.

  11. An electronic flight bag for NextGen avionics

    NASA Astrophysics Data System (ADS)

    Zelazo, D. Eyton

    2012-06-01

    The introduction of the Next Generation Air Transportation System (NextGen) initiative by the Federal Aviation Administration (FAA) will impose new requirements for cockpit avionics. A similar program is also taking place in Europe by the European Organisation for the Safety of Air Navigation (Eurocontrol) called the Single European Sky Air Traffic Management Research (SESAR) initiative. NextGen will require aircraft to utilize Automatic Dependent Surveillance-Broadcast (ADS-B) in/out technology, requiring substantial changes to existing cockpit display systems. There are two ways that aircraft operators can upgrade their aircraft in order to utilize ADS-B technology. The first is to replace existing primary flight displays with new displays that are ADS-B compatible. The second, less costly approach is to install an advanced Class 3 Electronic Flight Bag (EFB) system. The installation of Class 3 EFBs in the cockpit will allow aircraft operators to utilize ADS-B technology in a lesser amount of time with a decreased cost of implementation and will provide additional benefits to the operator. This paper describes a Class 3 EFB, the NexisTM Flight-Intelligence System, which has been designed to allow users a direct interface with NextGen avionics sensors while additionally providing the pilot with all the necessary information to meet NextGen requirements.

  12. Meet Mr. and Mrs. Gen X: A New Parent Generation

    ERIC Educational Resources Information Center

    Howe, Neil

    2010-01-01

    Slowly but surely, Generation Xers have been taking over from Baby Boomers as the majority of parents in elementary and secondary education. In the early 1990s, Gen Xers began joining parent-teacher associations in the nation's elementary schools. Around 2005, they became the majority of middle school parents. By the fall of 2008, they took over…

  13. A New Parent Generation: Meet Mr. and Mrs. Gen X

    ERIC Educational Resources Information Center

    Howe, Neil

    2010-01-01

    Slowly but surely, Generation Xers have been taking over from Baby Boomers as the majority of parents in elementary and secondary education. Gen-X parents and Boomer parents belong to two neighboring generations, each possessing its own location in history and its own peer personality. They are similar in some respects, but clearly different in…

  14. Meet Mr. and Mrs. Gen X: A New Parent Generation

    ERIC Educational Resources Information Center

    Howe, Neil

    2010-01-01

    Slowly but surely, Generation Xers have been taking over from Baby Boomers as the majority of parents in elementary and secondary education. In the early 1990s, Gen Xers began joining parent-teacher associations in the nation's elementary schools. Around 2005, they became the majority of middle school parents. By the fall of 2008, they took over…

  15. Safer Systems: A NextGen Aviation Safety Strategic Goal

    NASA Technical Reports Server (NTRS)

    Darr, Stephen T.; Ricks, Wendell R.; Lemos, Katherine A.

    2008-01-01

    The Joint Planning and Development Office (JPDO), is charged by Congress with developing the concepts and plans for the Next Generation Air Transportation System (NextGen). The National Aviation Safety Strategic Plan (NASSP), developed by the Safety Working Group of the JPDO, focuses on establishing the goals, objectives, and strategies needed to realize the safety objectives of the NextGen Integrated Plan. The three goal areas of the NASSP are Safer Practices, Safer Systems, and Safer Worldwide. Safer Practices emphasizes an integrated, systematic approach to safety risk management through implementation of formalized Safety Management Systems (SMS) that incorporate safety data analysis processes, and the enhancement of methods for ensuring safety is an inherent characteristic of NextGen. Safer Systems emphasizes implementation of safety-enhancing technologies, which will improve safety for human-centered interfaces and enhance the safety of airborne and ground-based systems. Safer Worldwide encourages coordinating the adoption of the safer practices and safer systems technologies, policies and procedures worldwide, such that the maximum level of safety is achieved across air transportation system boundaries. This paper introduces the NASSP and its development, and focuses on the Safer Systems elements of the NASSP, which incorporates three objectives for NextGen systems: 1) provide risk reducing system interfaces, 2) provide safety enhancements for airborne systems, and 3) provide safety enhancements for ground-based systems. The goal of this paper is to expose avionics and air traffic management system developers to NASSP objectives and Safer Systems strategies.

  16. Boomers, Gen-Xers, and Millennials: Understanding the "New Students."

    ERIC Educational Resources Information Center

    Oblinger, Diana

    2003-01-01

    Describes characteristics of the "new" college student, who may be an adult learner from the Baby Boomer era, a high school member of the "Millennial" generation, or a "Gen-Xer." Explores the learning styles of each type of student and discusses the importance of technology to each group. (SLD)

  17. Early detection of dominant Env-specific and subdominant Gag-specific CD8+ lymphocytes in equine infectious anemia virus-infected horses using major histocompatibility complex class I/peptide tetrameric complexes

    PubMed Central

    Mealey, Robert H.; Sharif, Amin; Ellis, Shirley A.; Littke, Matt H.; Leib, Steven R.; McGuire, Travis C.

    2012-01-01

    Cytotoxic T lymphocytes (CTL) are critical for control of lentiviruses, including equine infectious anemia virus (EIAV). Measurement of equine CTL responses has relied on chromium-release assays, which do not allow accurate quantitation. Recently, the equine MHC class I molecule 7-6, associated with the ELA-A1 haplotype, was shown to present both the Gag-GW12 and Env-RW12 EIAV CTL epitopes. In this study, 7-6/Gag-GW12 and 7-6/Env-RW12 MHC class I/peptide tetrameric complexes were constructed and used to analyze Gag-GW12- and Env-RW12-specific CTL responses in two EIAV-infected horses (A2164 and A2171). Gag-GW12 and Env-RW12 tetramer-positive CD8+ cells were identified in nonstimulated peripheral blood mononuclear cells as early as 14 days post-EIAV inoculation, and frequencies of tetramer-positive cells ranged from 0.4% to 6.7% of nonstimulated peripheral blood CD8+ cells during the 127-day study period. Although both horses terminated the initial viremic peak, only horse A2171 effectively controlled viral load. Neutralizing antibody was present during the initial control of viral load in both horses, but the ability to maintain control correlated with Gag-GW12-specific CD8+ cells in A2171. Despite Env-RW12 dominance, Env-RW12 escape viral variants were identified in both horses and there was no correlation between Env-RW12-specific CD8+ cells and control of viral load. Although Gag-GW12 CTL escape did not occur, a Gag-GW12 epitope variant arose in A2164 that was recognized less efficiently than the original epitope. These data indicate that tetramers are useful for identification and quantitation of CTL responses in horses, and suggest that the observed control of EIAV replication and clinical disease was associated with sustained CTL recognition of Gag-specific epitopes. PMID:15979679

  18. Early detection of dominant Env-specific and subdominant Gag-specific CD8+ lymphocytes in equine infectious anemia virus-infected horses using major histocompatibility complex class I/peptide tetrameric complexes.

    PubMed

    Mealey, Robert H; Sharif, Amin; Ellis, Shirley A; Littke, Matt H; Leib, Steven R; McGuire, Travis C

    2005-08-15

    Cytotoxic T lymphocytes (CTL) are critical for control of lentiviruses, including equine infectious anemia virus (EIAV). Measurement of equine CTL responses has relied on chromium-release assays, which do not allow accurate quantitation. Recently, the equine MHC class I molecule 7-6, associated with the ELA-A1 haplotype, was shown to present both the Gag-GW12 and Env-RW12 EIAV CTL epitopes. In this study, 7-6/Gag-GW12 and 7-6/Env-RW12 MHC class I/peptide tetrameric complexes were constructed and used to analyze Gag-GW12- and Env-RW12-specific CTL responses in two EIAV-infected horses (A2164 and A2171). Gag-GW12 and Env-RW12 tetramer-positive CD8+ cells were identified in nonstimulated peripheral blood mononuclear cells as early as 14 days post-EIAV inoculation, and frequencies of tetramer-positive cells ranged from 0.4% to 6.7% of nonstimulated peripheral blood CD8+ cells during the 127-day study period. Although both horses terminated the initial viremic peak, only horse A2171 effectively controlled viral load. Neutralizing antibody was present during the initial control of viral load in both horses, but the ability to maintain control correlated with Gag-GW12-specific CD8+ cells in A2171. Despite Env-RW12 dominance, Env-RW12 escape viral variants were identified in both horses and there was no correlation between Env-RW12-specific CD8+ cells and control of viral load. Although Gag-GW12 CTL escape did not occur, a Gag-GW12 epitope variant arose in A2164 that was recognized less efficiently than the original epitope. These data indicate that tetramers are useful for identification and quantitation of CTL responses in horses, and suggest that the observed control of EIAV replication and clinical disease was associated with sustained CTL recognition of Gag-specific epitopes.

  19. AL Amyloidosis

    PubMed Central

    2012-01-01

    Definition of the disease AL amyloidosis results from extra-cellular deposition of fibril-forming monoclonal immunoglobulin (Ig) light chains (LC) (most commonly of lambda isotype) usually secreted by a small plasma cell clone. Most patients have evidence of isolated monoclonal gammopathy or smoldering myeloma, and the occurrence of AL amyloidosis in patients with symptomatic multiple myeloma or other B-cell lymphoproliferative disorders is unusual. The key event in the development of AL amyloidosis is the change in the secondary or tertiary structure of an abnormal monoclonal LC, which results in instable conformation. This conformational change is responsible for abnormal folding of the LC, rich in β leaves, which assemble into monomers that stack together to form amyloid fibrils. Epidemiology AL amyloidosis is the most common type of systemic amyloidois in developed countries with an estimated incidence of 9 cases/million inhabitant/year. The average age of diagnosed patients is 65 years and less than 10% of patients are under 50. Clinical description The clinical presentation is protean, because of the wide number of tissues or organs that may be affected. The most common presenting symptoms are asthenia and dyspnoea, which are poorly specific and may account for delayed diagnosis. Renal manifestations are the most frequent, affecting two thirds of patients at presentation. They are characterized by heavy proteinuria, with nephrotic syndrome and impaired renal function in half of the patients. Heart involvement, which is present at diagnosis in more than 50% of patients, leading to restrictive cardiopathy, is the most serious complication and engages prognosis. Diagnostic methods The diagnosis relies on pathological examination of an involved site showing Congo red-positive amyloid deposits, with typical apple-green birefringence under polarized light, that stain positive with an anti-LC antibody by immunohistochemistry and/or immunofluorescence. Due to the

  20. DOE/NNSA perspective safeguard by design: GEN III/III+ light water reactors and beyond

    SciTech Connect

    Pan, Paul Y

    2010-12-10

    An overview of key issues relevant to safeguards by design (SBD) for GEN III/IV nuclear reactors is provided. Lessons learned from construction of typical GEN III+ water reactors with respect to SBD are highlighted. Details of SBD for safeguards guidance development for GEN III/III+ light water reactors are developed and reported. This paper also identifies technical challenges to extend SBD including proliferation resistance methodologies to other GEN III/III+ reactors (except HWRs) and GEN IV reactors because of their immaturity in designs.

  1. Comparison of telogen hair analyses: genRES MPX-2SP kit versus genRES MPX-SP1 and genRES MPX-SP2 kits.

    PubMed

    Schmid, D; Bayer, B; Anslinger, K

    2008-12-01

    STR investigations of telogen hair are invariably difficult due to the small amounts of nuclear DNA and its degradation products. However, in recent years there has been a considerable improvement. This study examined the suitability of a new STR kit with shortened amplicons for the investigation of hair in routine casework. This kit allows the simultaneous amplification of the eight STR-loci D3S1358, VWA, FGA, TH01, SE33, D8S1179, D18S51, and D21S11, and the sex-determining amelogenin system. It was tested against the genRES MPX-SP1 and genRES MPX-SP2 kits. The sensitivity of the new genRES MPX-2SP kit was demonstrated to be inferior to that of the genRES MPX-SP1, but almost equal to that of the genRES MPX-SP2 kit.

  2. Advanced Vehicle Concepts and Implications for NextGen

    NASA Technical Reports Server (NTRS)

    Blake, Matt; Smith, Jim; Wright, Ken; Mediavilla Ricky; Kirby, Michelle; Pfaender, Holger; Clarke, John-Paul; Volovoi, Vitali; Dorbian, Christopher; Ashok, Akshay; Reynolds, Tom; Waitz, Ian; Hileman, James; Arunachalam, Sarav; Hedrick, Matt; Vempati, Lakshmi; Laroza, Ryan; denBraven, Wim; Henderson, Jeff

    2010-01-01

    This report presents the results of a major NASA study of advanced vehicle concepts and their implications for the Next Generation Air Transportation System (NextGen). Comprising the efforts of dozens of researchers at multiple institutions, the analyses presented here cover a broad range of topics including business-case development, vehicle design, avionics, procedure design, delay, safety, environmental impacts, and metrics. The study focuses on the following five new vehicle types: Cruise-efficient short takeoff and landing (CESTOL) vehicles Large commercial tiltrotor aircraft (LCTRs) Unmanned aircraft systems (UAS) Very light jets (VLJs) Supersonic transports (SST). The timeframe of the study spans the years 2025-2040, although some analyses are also presented for a 3X scenario that has roughly three times the number of flights as today. Full implementation of NextGen is assumed.

  3. SimGen: A General Simulation Method for Large Systems.

    PubMed

    Taylor, William R

    2017-02-03

    SimGen is a stand-alone computer program that reads a script of commands to represent complex macromolecules, including proteins and nucleic acids, in a structural hierarchy that can then be viewed using an integral graphical viewer or animated through a high-level application programming interface in C++. Structural levels in the hierarchy range from α-carbon or phosphate backbones through secondary structure to domains, molecules, and multimers with each level represented in an identical data structure that can be manipulated using the application programming interface. Unlike most coarse-grained simulation approaches, the higher-level objects represented in SimGen can be soft, allowing the lower-level objects that they contain to interact directly. The default motion simulated by SimGen is a Brownian-like diffusion that can be set to occur across all levels of representation in the hierarchy. Links can also be defined between objects, which, when combined with large high-level random movements, result in an effective search strategy for constraint satisfaction, including structure prediction from predicted pairwise distances. The implementation of SimGen makes use of the hierarchic data structure to avoid unnecessary calculation, especially for collision detection, allowing it to be simultaneously run and viewed on a laptop computer while simulating large systems of over 20,000 objects. It has been used previously to model complex molecular interactions including the motion of a myosin-V dimer "walking" on an actin fibre, RNA stem-loop packing, and the simulation of cell motion and aggregation. Several extensions to this original functionality are described.

  4. Technology evaluation: C242-DM1, ImmunoGen Inc.

    PubMed

    Smith, S

    2001-04-01

    C242-DM1 is a tumor-activated immunotoxin under development by GlaxoSmithKline plc (formerly SmithKline Beecham plc), under licence from ImmunoGen Inc, as a potential treatment for colon tumor. It consists of a colon cancer-specific humanized antibody, C242, conjugated to the maytansine derivative DM1. In preclinical studies, C242-DM1 caused complete tumor regression in animal models of both human pancreatic and non-small cell lung cancer (NSCLC) at non-toxic doses. C242-DM1 has also been evaluated in an immunoconjugate combination with J-591 (Cornell University). The J591-DM1 immunoconjugate demonstrated effective, antigen-specific delivery of a highly cytotoxic drug to PSMA-positive Pca cells in vitro and in vivo with low systemic toxicity. Results from studies in monkeys showed that C242-DM1 had no significant toxicity or side effects, when administered at doses higher than those that were previously shown to completely eradicate human colon tumors in mice [271420]. ImmunoGen acquired the right to evaluate, and an option to license, technology related to maytansines from Takeda. In February 1999, ImmunoGen and SmithKline Beecham signed a US $45 million development and commercialization agreement for C242-DM1 [313493]. In August 1997, Immunogen received an SBIR grant to advance development of huC242-DM1 [258356]. EP-00425235, held by ImmunoGen, covers conjugated forms of ansamitocin (maytansine) derivatives. Takeda holds several patents for the production of ansamitocin and its analogs, the first one being JP-53124692.

  5. The GenABEL Project for statistical genomics

    PubMed Central

    Karssen, Lennart C.; van Duijn, Cornelia M.; Aulchenko, Yurii S.

    2016-01-01

    Development of free/libre open source software is usually done by a community of people with an interest in the tool. For scientific software, however, this is less often the case. Most scientific software is written by only a few authors, often a student working on a thesis. Once the paper describing the tool has been published, the tool is no longer developed further and is left to its own device. Here we describe the broad, multidisciplinary community we formed around a set of tools for statistical genomics. The GenABEL project for statistical omics actively promotes open interdisciplinary development of statistical methodology and its implementation in efficient and user-friendly software under an open source licence. The software tools developed withing the project collectively make up the GenABEL suite, which currently consists of eleven tools. The open framework of the project actively encourages involvement of the community in all stages, from formulation of methodological ideas to application of software to specific data sets. A web forum is used to channel user questions and discussions, further promoting the use of the GenABEL suite. Developer discussions take place on a dedicated mailing list, and development is further supported by robust development practices including use of public version control, code review and continuous integration. Use of this open science model attracts contributions from users and developers outside the “core team”, facilitating agile statistical omics methodology development and fast dissemination. PMID:27347381

  6. The GenABEL Project for statistical genomics.

    PubMed

    Karssen, Lennart C; van Duijn, Cornelia M; Aulchenko, Yurii S

    2016-01-01

    Development of free/libre open source software is usually done by a community of people with an interest in the tool. For scientific software, however, this is less often the case. Most scientific software is written by only a few authors, often a student working on a thesis. Once the paper describing the tool has been published, the tool is no longer developed further and is left to its own device. Here we describe the broad, multidisciplinary community we formed around a set of tools for statistical genomics. The GenABEL project for statistical omics actively promotes open interdisciplinary development of statistical methodology and its implementation in efficient and user-friendly software under an open source licence. The software tools developed withing the project collectively make up the GenABEL suite, which currently consists of eleven tools. The open framework of the project actively encourages involvement of the community in all stages, from formulation of methodological ideas to application of software to specific data sets. A web forum is used to channel user questions and discussions, further promoting the use of the GenABEL suite. Developer discussions take place on a dedicated mailing list, and development is further supported by robust development practices including use of public version control, code review and continuous integration. Use of this open science model attracts contributions from users and developers outside the "core team", facilitating agile statistical omics methodology development and fast dissemination.

  7. GEN IV reactors: Where we are, where we should go

    SciTech Connect

    Locatelli, G.; Mancini, M.; Todeschini, N.

    2012-07-01

    GEN IV power plants represent the mid-long term option of the nuclear sector. International literature proposes many papers and reports dealing with these reactors, but there is an evident difference of type and shape of information making impossible each kind of detailed comparison. Moreover, authors are often strongly involved in some particular design; this creates many difficulties in their super-partes position. Therefore it is necessary to put order in the most relevant information to understand strengths and weaknesses of each design and derive an overview useful for technicians and policy makers. This paper presents the state-of the art for GEN IV nuclear reactors providing a comprehensive literature review of the different designs with a relate taxonomy. It presents the more relevant references, data, advantages, disadvantages and barriers to the adoptions. In order to promote an efficient and wide adoption of GEN IV reactors the paper provides the pre-conditions that must be accomplished, enabling factors promoting the implementation and barriers limiting the extent and intensity of its implementation. It concludes outlying the state of the art of the most important R and D areas and the future achievements that must be accomplished for a wide adoption of these technologies. (authors)

  8. Description of Teunomyces gen. nov. for the Candida kruisii clade, Suhomyces gen. nov. for the Candida tanzawaensis clade and Suhomyces kilbournensis sp. nov.

    USDA-ARS?s Scientific Manuscript database

    DNA sequence analysis has shown that species of the Candida kruisii clade and species of the C. tanzawaensis clade represent phylogenetically circumscribed genera, which are described as Teunomyces gen. nov., type species T. kruisii, and Suhomyces gen. nov., type species S. tanzawaensis. Many of the...

  9. Detection of HIV type 1 env subtypes A, B, C, and E in Asia using dried blood spots: a new surveillance tool for molecular epidemiology.

    PubMed

    Cassol, S; Weniger, B G; Babu, P G; Salminen, M O; Zheng, X; Htoon, M T; Delaney, A; O'Shaughnessy, M; Ou, C Y

    1996-10-10

    Global surveillance of HIV-1 subtypes for genetic characterization is hampered by the biohazard of processing and the difficulties of shipping whole blood or cells from many developing country regions. We developed a technique for the direct automated sequencing of viral DNA from dried blood spot (DBS) specimens collected on absorbent paper, which can be mailed unrefrigerated in sturdy paper envelopes with low biohazard risk. DBS were collected nonrandomly from HIV-1-infected, mostly asymptomatic, patients in five Asian countries in 1991, and shipped via airmail or hand carried without refrigeration to Bangkok, and then transshipped to North America for processing. After more than 2 years of storage, including 6 months at ambient temperatures, proviral DNA in the DBS was amplified by nested PCR, and a 389-nucleotide segment of the C2-V3 env gene region was sequenced, from which 287 base pairs were aligned and subtyped by phylogenetic analysis with neighbor-joining and other methods. From southern India, there were 25 infections with subtype C and 2 with subtype A. From Myanmar (Burma), we identified the first subtype E infection, as well as six subtype BB, a distinct cluster within subtype B that was first discovered in Thailand and that has now appeared in China, Malaysia, and Japan. From southwest China, one BB was identified, while a "classical" B typical of North American and European strains was found in Indonesia. From Thailand, five DBS of ambiguous serotype were identified as three B, one BB, and one E. A blinded control serotype E specimen was correctly identified, but a serotype BB control was not tested. Most HIV-1 in southern India appears to be env subtype C, with rare A, as others have reported in western and northern India. The subtypes BB and E in Myanmar, and the BB in China, suggest epidemiological linkage with these subtypes in neighboring Thailand. DBS are a practical, economical technique for conducting large-scale molecular epidemiological

  10. Equine viperin restricts equine infectious anemia virus replication by inhibiting the production and/or release of viral Gag, Env, and receptor via distortion of the endoplasmic reticulum.

    PubMed

    Tang, Yan-Dong; Na, Lei; Zhu, Chun-Hui; Shen, Nan; Yang, Fei; Fu, Xian-Qiu; Wang, Yu-Hong; Fu, Li-Hua; Wang, Jia-Yi; Lin, Yue-Zhi; Wang, Xue-Feng; Wang, Xiaojun; Zhou, Jian-Hua; Li, Cheng-Yao

    2014-11-01

    Viperin is an endoplasmic reticulum (ER)-associated multifunctional protein that regulates virus replication and possesses broad antiviral activity. In many cases, viperin interferes with the trafficking and budding of viral structural proteins by distorting the membrane transportation system. The lentivirus equine infectious anemia virus (EIAV) has been studied extensively. In this study, we examined the restrictive effect of equine viperin (eViperin) on EIAV replication and investigated the possible molecular basis of this restriction to obtain insights into the effect of this cellular factor on retroviruses. We demonstrated that EIAV infection of primary equine monocyte-derived macrophages (eMDMs) upregulated the expression of eViperin. The overexpression of eViperin significantly inhibited the replication of EIAV in eMDMs, and knockdown of eViperin transcription enhanced the replication of EIAV in eMDMs by approximately 45.8%. Further experiments indicated that eViperin restricts EIAV at multiple steps of viral replication. The overexpression of eViperin inhibited EIAV Gag release. Both the α-helix domain and radical S-adenosylmethionine (SAM) domain were required for this activity. However, the essential motifs in SAM were different from those reported for the inhibition of HIV-1 Gag by human viperin. Furthermore, eViperin disrupted the synthesis of both EIAV Env and receptor, which consequently inhibited viral production and entry, respectively, and this disruption was dependent on the eViperin α-helix domain. Using immunofluorescence assays and electron microscopy, we demonstrated that the α-helix domain is responsible for the distortion of the endoplasmic reticulum (ER). Finally, EIAV did not exhibit counteracting eViperin at the protein level. In previous studies, viperin was indicated as restricting virus replications primarily by the inhibition of virus budding. Here, we show that viperin may have multiple antiviral mechanisms, including the reduction

  11. GenCade Version 1 Quick-Start Guide: How to Start a Successful GenCade Project

    DTIC Science & Technology

    2015-03-01

    modeler can easily make changes in the conceptual model and quickly generate new alternatives. The modeler should save new alternatives in different...to double-check that all features defined in the conceptual model are represented in the grid. When the project is saved, changes will be saved in...the control file. Changes made within the GenCade menu will be lost if the grid is regenerated, but changes made in the conceptual model will be

  12. Pantanalinema gen. nov. and Alkalinema gen. nov.: novel pseudanabaenacean genera (Cyanobacteria) isolated from saline-alkaline lakes.

    PubMed

    Vieira Vaz, Marcelo Gomes Marçal; Genuário, Diego Bonaldo; Andreote, Ana Paula Dini; Malone, Camila Francieli Silva; Sant'Anna, Célia Leite; Barbiero, Laurent; Fiore, Marli Fátima

    2015-01-01

    The genus Leptolyngbya Anagnostidis & Komárek (1988) was described from a set of strains identified as 'LPP-group B'. The morphology within this group is not particularly informative and underestimates the group's genetic diversity. In the present study, two new pseudanabaenacean genera related to Leptolyngbya morphotypes, Pantanalinema gen. nov. and Alkalinema gen. nov., are described under the provisions of the International Code of Nomenclature for Algae, Fungi and Plants, based on a polyphasic approach. Pantanalinema gen. nov. (type species Pantanalinema rosaneae sp. nov.) has sheaths and trichomes with slight gliding motility, which distinguish this genus from Alkalinema gen. nov. (type species Alkalinema pantanalense sp. nov.), which possesses trichomes arranged in an ornate (interwoven) pattern. 16S rRNA gene sequences of strains of Pantanalinema and Alkalinema exhibited low identity to each other (≤91.6 %) and to other sequences from known pseudanabaenacean genera (≤94.3 and 93.7 %, respectively). In a phylogenetic reconstruction, six sequences from strains of Pantanalinema and four from strains of Alkalinema formed two separate and robust clades (99 % bootstrap value), with the genera Oculatella and Phormidesmis, respectively, as the closest related groups. 16S-23S rRNA intergenic spacer sequences and secondary structures of strains of Pantanalinema and Alkalinema did not correspond to any previous descriptions. The strains of Pantanalinema and Alkalinema were able to survive and produce biomass at a range of pH (pH 4-11) and were also able to alter the culture medium to pH values ranging from pH 8.4 to 9.9. These data indicate that cyanobacterial communities in underexplored environments, such as the Pantanal wetlands, are promising sources of novel taxa.

  13. Xerochrysium gen. nov. and Bettsia, genera encompassing xerophilic species of Chrysosporium.

    PubMed

    Pitt, John I; Lantz, Henrik; Pettersson, Olga Vinnere; Leong, Su-Lin L

    2013-12-01

    On the basis of a study of ITS sequences, Vidal et al. (Rev. Iber. Micol. 17: 22, 2000) recommended that the genus Chrysosporium be restricted to species belonging to Onygenales. Using nrLSU genes, we studied the majority of clades examined by Vidal et al. and showed that currently accepted species in Chrysosporium phylogenetically belong in six clades in three orders. Surprisingly, the xerophilic species of Chrysosporium, long thought to be a single grouping away from the majority of Chrysosporium species, occupy two clades, one in Leotiales, the other in Eurotiales. Species accepted in Leotiales are related to the sexual genus Bettsia. One is the type species B. alvei, and related asexual strains classified as C. farinicola, the second is C. fastidium transferred to Bettsia as B. fastidia. Species in the Eurotiales are transferred to Xerochrysium gen. nov., where the accepted species are X. xerophilum and X. dermatitidis, the correct name for C. inops on transfer to Xerochrysium. All accepted species are extreme xerophiles, found in dried and concentrated foods.

  14. Paring Down HIV Env: Design and Crystal Structure of a Stabilized Inner Domain of HIV-1 gp120 Displaying a Major ADCC Target of the A32 Region.

    PubMed

    Tolbert, William D; Gohain, Neelakshi; Veillette, Maxime; Chapleau, Jean-Philippe; Orlandi, Chiara; Visciano, Maria L; Ebadi, Maryam; DeVico, Anthony L; Fouts, Timothy R; Finzi, Andrés; Lewis, George K; Pazgier, Marzena

    2016-05-03

    Evidence supports a role of antibody-dependent cellular cytotoxicity (ADCC) toward transitional epitopes in the first and second constant (C1-C2) regions of gp120 (A32-like epitopes) in preventing HIV-1 infection and in vaccine-induced protection. Here, we describe the first successful attempt at isolating the inner domain (ID) of gp120 as an independent molecule that encapsulates the A32-like region within a minimal structural unit of the HIV-1 Env. Through structure-based design, we developed ID2, which consists of the ID expressed independently of the outer domain and stabilized in the CD4-bound conformation by an inter-layer disulfide bond. ID2 expresses C1-C2 epitopes in the context of CD4-triggered full-length gp120 but without any known neutralizing epitope present. Thus, ID2 represents a novel probe for the analysis and/or selective induction of antibody responses to the A32 epitope region. We also present the crystal structure of ID2 complexed with mAb A32, which defines its epitope.

  15. Sequence analysis of selected regions of the env (V3 loop and gp41) and gag (p7) reading frames of Ethiopian human immunodeficiency virus type 1 strains.

    PubMed

    Ayehunie, S; Johansson, B; Sönnerborg, A; Zewdie, D W; Britton, S; Strannegård, O

    1993-01-01

    Amplified polymerase chain reaction (PCR) products, corresponding to the V3 loop and gp41 of the env, and p7 of the gag region, from proviral DNA of several Ethiopian and Swedish HIV-1 strains were sequenced. Of the six amino acids (GPGRAF) that constitute the principal neutralizing determinant (PND) within the V3 loop, the Ethiopian isolates all showed two amino acid changes (GPGQTF). Four to five other substitutions were found in the amino acids flanking the PND. Substitution of alanine (A) for threonine (T) should result in a change in the predicted secondary structure, i.e., disappearance of a coil structure. Percentage similarity data on a stretch of 22 amino acids within the V3 loop showed a concordance of the Ethiopian HIV-1 isolates with the sequences of published macrophage-T-cell tropic HIV isolates. Additionally derived protein sequences in two other regions showed two common substitutions in p7 and one to two substitutions in gp41 compared to a recent consensus sequence. These changes are hitherto unique for the Ethiopian strains, and suggest the presence of a clustering of a divergent HIV-1 strain in Addis Ababa, Ethiopia.

  16. The opgC gene is required for OPGs succinylation and is osmoregulated through RcsCDB and EnvZ/OmpR in the phytopathogen Dickeya dadantii

    PubMed Central

    Bontemps-Gallo, Sébastien; Madec, Edwige; Robbe-Masselot, Catherine; Souche, Erika; Dondeyne, Jacqueline; Lacroix, Jean-Marie

    2016-01-01

    Osmoregulated periplasmic glucans (OPGs) are a family of periplasmic oligosaccharides found in the envelope of most Proteobacteria. They are required for virulence of zoo- and phyto-pathogens. The glucose backbone of OPGs is substituted by various kinds of molecules depending on the species, O-succinyl residues being the most widely distributed. In our model, Dickeya dadantii, a phytopathogenic bacteria causing soft rot disease in a wide range of plant species, the backbone of OPGs is substituted by O-succinyl residues in media of high osmolarity and by O-acetyl residues whatever the osmolarity. The opgC gene encoding a transmembrane protein required for the succinylation of the OPGs in D. dadantii was found after an in silico search of a gene encoding a protein with the main characteristics recovered in the two previously characterized OpgC of E. coli and R. sphaeroides, i.e. 10 transmembrane segments and one acyl-transferase domain. Characterization of the opgC gene revealed that high osmolarity expression of the succinyl transferase is controlled by both the EnvZ-OmpR and RcsCDB phosphorelay systems. The loss of O-succinyl residue did not affect the virulence of D. dadantii, suggesting that only the glucose backbone of OPGs is required for virulence. PMID:26790533

  17. Prevalence of fowl glioma-inducing virus in chickens of zoological gardens in Japan and nucleotide variation in the env gene.

    PubMed

    Hatai, Hitoshi; Ochiai, Kenji; Murakami, Mariko; Imanishi, Syunsuke; Tomioka, Yukiko; Toyoda, Takeshi; Ohashi, Kazuhiko; Umemura, Takashi

    2008-05-01

    Fowl glioma-inducing virus (FGV), which belongs to subgroup A of avian leukosis virus (ALV), is tumorigenic in the nervous system. In a zoological garden in Japan, approximately 40% of chickens, including Japanese fowls, were infected with FGV. Because this zoological garden plays a role as a major supplier of Japanese fowl for other zoological gardens, FGV infection is suspected to have spread among ornamental chickens. In this study, the prevalence of the disease was examined in a total of 129 chickens in three other zoological gardens by nested polymerase chain reaction (PCR), reverse transcription nested PCR and enzyme-linked immunosorbent assay. Twenty-six to 56 percent of the fowls in each of the examined gardens were positive by nested PCR. The phylogenetic analysis revealed that the 3' untranslated region, including the specific sequence of FGV, of the 14 isolated ALVs showed high sequence identity and a close relationship with FGV. In addition, the env gene of the isolates frequently showed mutations and deletions of nucleotides. These results suggest that FGV is prevalent among ornamental chickens kept in zoological gardens in Japan.

  18. Amino acid mutations in the env gp90 protein that modify N-linked glycosylation of the Chinese EIAV vaccine strain enhance resistance to neutralizing antibodies.

    PubMed

    Han, Xiue; Zhang, Ping; Yu, Wei; Xiang, Wenhua; Li, Xiaodong

    2016-12-01

    The Chinese EIAV vaccine is an attenuated live virus vaccine obtained by serial passage of a virulent horse isolate (EIAVL) in donkeys (EIAVD) and, subsequently, in donkey cells in vitro. In this study, we compare the env gene of the original horse virulent virus (EIAVL) with attenuated strains serially passaged in donkey MDM (EIAVDLV) and donkey dermal cells (EIAVFDDV). Genetic comparisons among parental and attenuated strains found that vaccine strains contained amino acid substitutions/deletions in gp90 that resulted in a loss of three potential N-linked glycosylation sites, designated g5, g9, and g10. To investigate the biological significance of these changes, reverse-mutated viruses were constructed in the backbone of the EIAVFDDV infectious molecular clone (pLGFD3). The resulting virus stocks were characterized for replication efficiency in donkey dermal cells and donkey MDM, and were tested for sensitivity to neutralization using sera from two ponies experimentally infected with EIAVFDDV. Results clearly show that these mutations generated by site-directed mutagenesis resulted in cloned viruses with enhanced resistance to serum neutralizing antibodies that were also able to recognize parental viruses. This study indicates that these mutations played an important role in the attenuation of the EIAV vaccine strains.

  19. Phylogenetic analysis of env, gag, and tat genes of HIV type 1 detected among the injecting drug users in West Bengal, India.

    PubMed

    Mullick, Ranajoy; Sengupta, Satarupa; Sarkar, Kamalesh; Saha, M K; Chakrabarti, Sekhar

    2006-12-01

    A recent occurrence of HIV-1 seropositivity among a group of injecting drug users (IDUs) in Darjeeling, a hilly district in northern West Bengal, revealed overall 11.8% HIV seroprevalence. Our study based on env (C2-V3), gag (p24-p7), and tat (exon-1) genomic regions of HIV-1 detected among this population showed that Darjeeling IDU sequences belonged to subtype C. Interestingly, the IDU sequences from Darjeeling were again found to be closer to the C strains from Manipur, a northeastern state in India, which is linked to the Golden Triangle via the Manipur-Myanmar border, rather than the IDU C sequences from Nepal, a neighboring country of India. The outgroup reference strains from different sites of IDU-driven epidemics in the world like Russia, Vietnam, Thailand, and Spain belonged to the nonsubtype C group and formed separate clusters from the subtype C cluster in our analysis. These results indicate a rapid spread of HIV-1 by possible drug trafficking along international boundaries, which might also help in the invasion of HIV-1 among IDUs of Darjeeling through the Manipur-Myanmar border of India.

  20. HSV-1 amplicon vectors elicit polyfunctional T cell responses to HIV-1 Env, and strongly boost responses to an adenovirus prime

    PubMed Central

    Duke, Cindy M.P.; Maguire, Casey A.; Keefer, Michael C.; Federoff, Howard J.; Bowers, William J.; Dewhurst, Stephen

    2007-01-01

    HSV-1 amplicon vectors elicit strong T-cell responses to encoded antigens but the qualitative nature of these responses is poorly understood. Antigen-specific CD4+ and CD8+ T-cell responses to amplicon and adenovirus (rAd5) vectors encoding HIV-1 gp120 were assessed following immunization of mice, by performing intracellular cytokine staining for IFNγ, IL-2 and TNFα, following stimulation of splenocytes with a HIV-1 Env peptide pool. The quality of the primary T-cell response to amplicon and rAd5 vectors was strikingly similar, but there were qualitative differences in responses to amplicon vectors that incorporated different promoters upstream of gp120 - suggesting that promoters can significantly influence immune response quality. When prime-boost combinations were studied, a rAd5 prime and amplicon boost elicited the highest T-cell response. Furthermore, protocols that incorporated a rAd5 prime consistently elicited a greater proportion of polyfunctional CD4+ T-cells - regardless of boost. This suggests that initial priming can shape immune response quality after a boost. Overall, these findings provide insight into effective vector combinations for HIV-1 vaccine development. PMID:17868958

  1. Detection of immunoreactive epitopes in proteins encoded by gag, env, and pol genes of human T-lymphotropic virus type I using synthetic peptides

    SciTech Connect

    Yaroslavtseva, N.G.; Kornilaeva, G.V.; Pashkova, T.A.

    1995-10-01

    Reactivity of 26 synthetic peptides that comprise 12 to 26 amino acid residues corresponding to segments of the p19 (gag), gp46 (env), and pol proteins (pol) of human T-lymphotropic virus type I toward 31 positive sera was studied using enzyme-linked immunosorbent assay. Specific reactivity with high titers of antibodies (presented in reciprocal dilution values) was detected for the synthetic peptides corresponding to fragments 110-130 and 100-130 (titers up to 4050) of p19, 174-197 (up to 800), 186-201 (up to to 4050), 191-215 (up to 1350), 242-257 (up to 800), and 272-292 (up to 450) of gp46. Immunoreactivity of seven peptides, fragments of pol-proteins, was weak. New linear epitopes in the regions 145-158, 272-277, and 292-300 of gp46 were detected. In addition, location of the known linear epitopes in p19 and gp46 was refined on the basis of comparative study of overlapping peptides from these proteins. 25 refs., 4 tabs.

  2. The Madagascan endemic myrmicine ants related to Eutetramorium (Hymenoptera: Formicidae): taxonomy of the genera Eutetramorium Emery, Malagidris nom. n., Myrmisaraka gen. n., Royidris gen. n., and Vitsika gen. n.

    PubMed

    Bolton, Barry; Fisher, Brian L

    2014-04-24

    The monophyletic group of myrmicine ant genera related to Eutetramorium is described and its taxonomy is documented. The group is endemic in Madagascar and contains five genera: Eutetramorium Emery, 1899 (3 species, 1 of which is new); Malagidris nom. n., a replacement name for Brunella Forel, 1917, junior homonym of Brunella Smith, G.W. 1909 (Crustacea) (6 species, 5 of which are new); Myrmisaraka gen. n. (2 species, both new); Royidris gen. n. (15 species, 11 of which are new); Vitsika gen. n. (14 species, all of which are new). Keys to the worker caste are provided for all genera, and provisional keys to known males are given for Malagidris and Vitsika.

  3. Potential impact of viral load and genetic makeup of HIV type 1 on mother-to-child transmission: characterization of env-C2V3C3 and nef sequences.

    PubMed

    Pádua, Elizabeth; Parreira, Ricardo; Tendeiro, Rita; Nunes, Baltazar; Castela, João; Soares, Isabel; Mouzinho, Ana; Reis, Eduarda; Paixão, Maria Teresa

    2009-11-01

    HIV-1 mother-to-child transmission (MTCT) was evaluated in terms of the molecular characterization of the env and nef genomic regions and quantification of maternal RNA viral loads. Assignment of viral subtype was achieved by direct sequencing of PCR 1172 products amplified from proviral DNA in 45 HIV-1-nontransmitting mothers (NTM), along with 13 pairs of HIV-1-transmitting mothers (TM) and their infected children (C). Analysis of the env C2V3C3 and nef sequences revealed that subtypes G and B, and their genetic combinations (AG, BG), accounted for over 84.5% of all viruses identified. The genetic structure form envA-nefG was the most commonly observed, with a lower frequency in the NTM (13.3%) compared to the TM (23.1%) group. A greater number of genetic forms was observed among NTM, namely the presence of sequences assigned to subtypes D and F, as well as the intergenetic A/J, and C/U, recombinant forms, along with a mosaic provirus with a complex putative envA-nefEGE genetic structure. No significant differences were found when RNA viral loads were evaluated as a function of the viral subtypes. Nevertheless, a relatively high quantification of HIV-1 RNA was obtained in the NTM group, emphasizing the importance of the compliance and effectiveness of therapeutic schemes to control viral replication and reduce the risk of HIV vertical transmission. V3 sequences displaying features associated with the R5 phenotype dominated in both groups. Both C2V3C3 and Nef's functional domains were conserved during HIV-1 vertical transmission.

  4. Complete epitopes for vaccine design derived from a crystal structure of the broadly neutralizing antibodies PGT128 and 8ANC195 in complex with an HIV-1 Env trimer.

    PubMed

    Kong, Leopold; Torrents de la Peña, Alba; Deller, Marc C; Garces, Fernando; Sliepen, Kwinten; Hua, Yuanzi; Stanfield, Robyn L; Sanders, Rogier W; Wilson, Ian A

    2015-10-01

    The HIV-1 envelope gp160 glycoprotein (Env) is a trimer of gp120 and gp41 heterodimers that mediates cell entry and is the primary target of the humoral immune response. Broadly neutralizing antibodies (bNAbs) to HIV-1 have revealed multiple epitopes or sites of vulnerability, but mapping of most of these sites is incomplete owing to a paucity of structural information on the full epitope in the context of the Env trimer. Here, a crystal structure of the soluble BG505 SOSIP gp140 trimer at 4.6 Å resolution with the bNAbs 8ANC195 and PGT128 reveals additional interactions in comparison to previous antibody-gp120 structures. For 8ANC195, in addition to previously documented interactions with gp120, a substantial interface with gp41 is now elucidated that includes extensive interactions with the N637 glycan. Surprisingly, removal of the N637 glycan did not impact 8ANC195 affinity, suggesting that the antibody has evolved to accommodate this glycan without loss of binding energy. PGT128 indirectly affects the N262 glycan by a domino effect, in which PGT128 binds to the N301 glycan, which in turn interacts with and repositions the N262 glycan, thereby illustrating the important role of neighboring glycans on epitope conformation and stability. Comparisons with other Env trimer and gp120 structures support an induced conformation for glycan N262, suggesting that the glycan shield is allosterically modified upon PGT128 binding. These complete epitopes of two broadly neutralizing antibodies on the Env trimer can now be exploited for HIV-1 vaccine design.

  5. Cross-Neutralizing Antibodies in HIV-1 Individuals Infected by Subtypes B, F1, C or the B/Bbr Variant in Relation to the Genetics and Biochemical Characteristics of the env Gene

    PubMed Central

    de Almeida, Dalziza Victalina; Macieira, Karine Venegas; Grinsztejn, Beatriz Gilda Jegerhorn; Veloso dos Santos, Valdiléa Gonçalves; Guimarães, Monick Lindenmeyer

    2016-01-01

    Various HIV-1 env genetic and biochemical features impact the elicitation of cross-reactive neutralizing antibodies in natural infections. Thus, we aimed to investigate cross-neutralizing antibodies in individuals infected with HIV-1 env subtypes B, F1, C or the B/Bbr variant as well as env characteristics. Therefore, plasma samples from Brazilian chronically HIV-1 infected individuals were submitted to the TZM-bl neutralization assay. We also analyzed putative N-glycosylation sites (PNGLs) and the size of gp120 variable domains in the context of HIV-1 subtypes prevalent in Brazil. We observed a greater breadth and potency of the anti-Env neutralizing response in individuals infected with the F1 or B HIV-1 subtypes compared with the C subtype and the variant B/Bbr. We observed greater V1 B/Bbr and smaller V4 F1 than those of other subtypes (p<0.005), however neither was there a correlation verified between the variable region length and neutralization potency, nor between PNLG and HIV-1 subtypes. The enrichment of W at top of V3 loop in weak neutralizing response viruses and the P in viruses with higher neutralization susceptibility was statistically significant (p = 0.013). Some other signatures sites were associated to HIV-1 subtype-specific F1 and B/Bbr samples might influence in the distinct neutralizing response. These results indicate that a single amino acid substitution may lead to a distinct conformational exposure or load in the association domain of the trimer of gp120 and interfere with the induction power of the neutralizing response, which affects the sensitivity of the neutralizing antibody and has significant implications for vaccine design. PMID:27936047

  6. J3Gen: A PRNG for Low-Cost Passive RFID

    PubMed Central

    Melià-Seguí, Joan; Garcia-Alfaro, Joaquin; Herrera-Joancomartí, Jordi

    2013-01-01

    Pseudorandom number generation (PRNG) is the main security tool in low-cost passive radio-frequency identification (RFID) technologies, such as EPC Gen2. We present a lightweight PRNG design for low-cost passive RFID tags, named J3Gen. J3Gen is based on a linear feedback shift register (LFSR) configured with multiple feedback polynomials. The polynomials are alternated during the generation of sequences via a physical source of randomness. J3Gen successfully handles the inherent linearity of LFSR based PRNGs and satisfies the statistical requirements imposed by the EPC Gen2 standard. A hardware implementation of J3Gen is presented and evaluated with regard to different design parameters, defining the key-equivalence security and nonlinearity of the design. The results of a SPICE simulation confirm the power-consumption suitability of the proposal. PMID:23519344

  7. J3Gen: a PRNG for low-cost passive RFID.

    PubMed

    Melià-Seguí, Joan; Garcia-Alfaro, Joaquin; Herrera-Joancomartí, Jordi

    2013-03-19

    Pseudorandom number generation (PRNG) is the main security tool in low-cost passive radio-frequency identification (RFID) technologies, such as EPC Gen2. We present a lightweight PRNG design for low-cost passive RFID tags, named J3Gen. J3Gen is based on a linear feedback shift register (LFSR) configured with multiple feedback polynomials. The polynomials are alternated during the generation of sequences via a physical source of randomness. J3Gen successfully handles the inherent linearity of LFSR based PRNGs and satisfies the statistical requirements imposed by the EPC Gen2 standard. A hardware implementation of J3Gen is presented and evaluated with regard to different design parameters, defining the key-equivalence security and nonlinearity of the design. The results of a SPICE simulation confirm the power-consumption suitability of the proposal.

  8. Specific sequences commonly found in the V3 domain of HIV-1 subtype C isolates affect the overall conformation of native Env and induce a neutralization-resistant phenotype independent of V1/V2 masking.

    PubMed

    Salomon, Aidy; Krachmarov, Chavdar; Lai, Zhong; Honnen, William; Zingman, Barry S; Sarlo, Julie; Gorny, Miroslaw K; Zolla-Pazner, Susan; Robinson, James E; Pinter, Abraham

    2014-01-05

    Primary HIV-1 isolates are relatively resistant to neutralization by antibodies commonly induced after infection or vaccination. This is generally attributed to masking of sensitive epitopes by the V1/V2 domain and/or glycans situated at various positions in Env. Here we identified a novel masking effect mediated by subtype C-specific V3 sequences that contributes to the V1/V2-independent and glycan-independent neutralization resistance of chimeric and primary Envs to antibodies directed against multiple neutralization domains. Positions at several conserved charged and hydrophobic sites in the V3 crown and stem were also shown to affect neutralization phenotype. These results indicated that substitutions typically present in subtype C and related V3 sequences influence the overall conformation of native Env in a way that occludes multiple neutralization targets located both within and outside of the V3 domain, and may reflect an alternative mechanism for neutralization resistance that is particularly active in subtype C and related isolates.

  9. Molecular subtypes of env sequences around V3 region of human immunodeficiency virus type 1 in Taiwan.

    PubMed

    Chang, K S; Lin, C I; Ling, P; Lin, K H; Lin, H C; Twu, S J

    1994-06-01

    Samples of peripheral blood mononuclear cells (PBMC) were collected during 1990-91 from seropositive healthy, male HIV-1 carriers visiting Taipei Venereal Disease Control Center, and a male AIDS patient admitted to a general hospital. The V3 and its flanking nucleotide (nt) sequences in their DNA were amplified by polymerase chain reaction (PCR) and compared with those of known HIV-1 prototypes. The nt sequences obtained from 21 individuals (e.g., TW92) clustered as Group A, which were highly homologous (95.6-99.5%) to that of HXB2 virus while those from 6 individuals (TW90, TW91, TW97, TW99, TW102 and TW104) were classified as Group B showing low similarities (73.2-84.2%) to those of HXB2 and moderate similarities (80.7-90.0%) to those of SC and Bangkok (BK) viruses. By comparison of their deduced amino acid sequences with those of consensus sequences for subtypes A-F as defined by Myers et al. (1993), both Groups A and B viruses (except TW102) together with those of HXB2, SC and BK viruses could be identified as members or variants of subtype B, and the TW102 virus as a member of subtype E viruses. Individuals with the Group A viruses included 4 homosexual and 17 heterosexual Taiwanese males, 2 of the latter having a history of i.v. drug abuse. Among individuals with Group B viruses, those with TW97, TW99, TW104 and TW91, who was an AIDS patient, were heterosexual Taiwanese males, whereas both TW90 and TW102 viruses were from individuals who were overseas heterosexual Chinese from Thailand, the former with a history of i.v. drug abuse and the latter without.

  10. Recent changes in the GenBank On-line Service.

    PubMed Central

    Benton, D

    1990-01-01

    The GenBank On-line Service provides access to the GenBank and EMBL nucleic acid sequence databases and to the Swiss-Prot and GenPept protein sequence databases. Users can query the databases by sequence similarity and annotation keywords and retrieve entries of interest. This access is available through e-mail servers, anonymous FTP, anonymous interactive login, and login to established, password-protected, individual accounts. PMID:2326192

  11. Arctocypris fuhrmanni, n. gen., n. sp. (Crustacea, Ostracoda, Eucypridinae) from Spitsbergen (Norway).

    PubMed

    Petkovski, Trajan K; Scharf, Burkhard; Keyser, Dietmar

    2016-01-14

    Material from Spitsbergen (Norway) collected by Spitzenberger (1996) was reinvestigated. A new genus Arctocypris and a new species Arctocypris. fuhrmanni n. gen. n. sp. are described in the present paper. A key to the genera of the subfamily Eucypridinae is provided. At the moment Arctocypris n. gen. comprises four species: Arctocypris arctica (Olofsson, 1918) comb. nov.; A. dulcifons (Diebel & Pietrzeniuk, 1969) comb. nov.; A. foveata (Delorme, 1968) comb. nov. and Arctocypris fuhrmanni n. gen., n. sp.

  12. Measurement of the neutron electric form factor GEn in quasielastic scattering

    SciTech Connect

    Donal Day

    2003-07-15

    We have measured the electric form factor of the neutron, GEn, at two momentum transfers (Q2= 0.5 and Q2= 1.0 GeV/c2) through quasielastic scattering in Jefferson Lab's Hall C. Longitudinally polarized electrons scattered from polarized deuterated ammonia and GEn was extracted from the beam-target asymmetry AVed which, in quasielastic kinematics, is particularly sensitive to GEn and insensitive to MEC and FSI.

  13. Proposal to reclassify Brenneria quercina (Hildebrand and Schroth 1967) Hauben et al. 1999 into a new genus, Lonsdalea gen. nov., as Lonsdalea quercina comb. nov., descriptions of Lonsdalea quercina subsp. quercina comb. nov., Lonsdalea quercina subsp. iberica subsp. nov. and Lonsdalea quercina subsp. britannica subsp. nov., emendation of the description of the genus Brenneria, reclassification of Dickeya dieffenbachiae as Dickeya dadantii subsp. dieffenbachiae comb. nov., and emendation of the description of Dickeya dadantii.

    PubMed

    Brady, Carrie L; Cleenwerck, Ilse; Denman, Sandra; Venter, Stephanus N; Rodríguez-Palenzuela, Pablo; Coutinho, Teresa A; De Vos, Paul

    2012-07-01

    Bacterial isolates from oak trees in Spain and Britain, showing symptoms of bark canker and Acute Oak Decline (AOD), respectively, were examined by a polyphasic approach. Both 16S rRNA gene sequencing and multilocus sequence analysis (MLSA), based on partial sequences of gyrB, rpoB, infB and atpD genes, revealed that the isolates were separated into two genetic groups according to their origin. Their closest phylogenetic relative was Brenneria quercina, the causal agent of drippy nut disease of oak, which clustered distant to the other species of the genus Brenneria. MLSA data for species of the genera Brenneria, Pectobacterium, Dickeya, Erwinia, Pantoea and Samsonia confirmed the polyphyletic nature of the genus Brenneria and indicated synonymy of Dickeya dadantii and Dickeya dieffenbachiae. DNA-DNA hybridization experiments confirmed this synonymy and also revealed DNA-DNA relatedness values of 58-73% between the new oak isolates and B. quercina. Phenotypic and/or chemotaxonomic methods allowed B. quercina and the two genetic groups of new oak isolates to be discriminated from other recognized species of the genus Brenneria and from members of the closely related genera Dickeya, Pectobacterium and Samsonia. Based on the data obtained, the following taxonomic proposals are made: (1) reclassification of B. quercina as the type species of a novel genus, Lonsdalea gen. nov., as Lonsdalea quercina comb. nov. (type strain LMG 2724(T)=ATCC 29281(T)=CCUG 48867(T)=CFBP 3617(T)=CIP 105201(T)=DSM 4561(T)=ICMP 1845(T)), (2) classification of the oak isolates as Lonsdalea quercina subsp. iberica subsp. nov. (type strain LMG26264(T)=NCPPB 4490(T)) and Lonsdalea quercina subsp. britannica subsp. nov. (type strain LMG 26267(T)=NCPPB 4481(T)) and leading to the automatic creation of Lonsdalea quercina subsp. quercina subsp. nov. (type strain LMG 2724(T)=ATCC 29281(T)), (3) emendation of the description of the genus Brenneria, and (4) reclassification of Dickeya dieffenbachiae as

  14. Resolvase OsGEN1 Mediates DNA Repair by Homologous Recombination1[OPEN

    PubMed Central

    Lu, Pingli

    2017-01-01

    Yen1/GEN1 are canonical Holliday junction resolvases that belong to the RAD2/XPG family. In eukaryotes, such as budding yeast, mice, worms, and humans, Yen1/GEN1 work together with Mus81-Mms4/MUS81-EME1 and Slx1-Slx4/SLX1-SLX4 in DNA repair by homologous recombination to maintain genome stability. In plants, the biological function of Yen1/GEN1 remains largely unclear. In this study, we characterized the loss of function mutants of OsGEN1 and OsSEND1, a pair of paralogs of Yen1/GEN1 in rice (Oryza sativa). We first investigated the role of OsGEN1 during meiosis and found a reduction in chiasma frequency by ∼6% in osgen1 mutants, compared to the wild type, suggesting a possible involvement of OsGEN1 in the formation of crossovers. Postmeiosis, OsGEN1 foci were detected in wild-type microspore nuclei, but not in the osgen1 mutant concomitant with an increase in double-strand breaks. Persistent double-strand breaks led to programmed cell death of the male gametes and complete male sterility. In contrast, depletion of OsSEND1 had no effects on plant development and did not enhance osgen1 defects. Our results indicate that OsGEN1 is essential for homologous recombinational DNA repair at two stages of microsporogenesis in rice. PMID:28049740

  15. Phen-Gen: combining phenotype and genotype to analyze rare disorders.

    PubMed

    Javed, Asif; Agrawal, Saloni; Ng, Pauline C

    2014-09-01

    We introduce Phen-Gen, a method that combines patients' disease symptoms and sequencing data with prior domain knowledge to identify the causative genes for rare disorders. Simulations revealed that the causal variant was ranked first in 88% of cases when it was a coding variant-a 52% advantage over a genotype-only approach-and Phen-Gen outperformed other existing prediction methods by 13-58%. If disease etiology was unknown, the causal variant was assigned the top rank in 71% of simulations. Phen-Gen is available at http://phen-gen.org/.

  16. GRAIL and GenQuest Sequence Annotation Tools

    SciTech Connect

    Xu, Ying; Shah, Manesh B.; Einstein, J. Ralph; Parang, Morey; Snoddy, Jay; Petrov, Sergey; Olman, Victor; Zhang, Ge; Mural, Richard J.; Uberbacher, Edward C.

    1997-12-31

    Our goal is to develop and implement an integrated intelligent system which can recognize biologically significant features in DNA sequence and provide insight into the organization and function of regions of genomic DNA. GRAIL is a modular expert system which facilitates the recognition of gene features and provides an environment for the construction of sequence annotation. The last several years have seen a rapid evolution of the technology for analyzing genomic DNA sequences. The current GRAIL systems (including the e-mail, XGRAIL, JAVA-GRAIL and genQuest systems) are perhaps the most widely used, comprehensive, and user friendly systems available for computational characterization of genomic DNA sequence.

  17. GenCade Application at Onslow Bay, North Carolina

    DTIC Science & Technology

    2012-08-01

    by Ashley E. Frey, Sophie Munger, Greg L. Williams , Michael J. Wutkowski, and Kevin B. Conner PURPOSE: This Coastal and Hydraulics Engineering...Texas A&M University Corpus Christi, and Greg L. Williams , Michael J. Wutkowski, and Kevin B. Conner of the U.S. Army Engineer District, Wilmington. Dr...follows: Frey, A.E., Munger, S., Williams , G.L., Wutkowski, M.J. and Conner, K.B. 2012. GenCade Application at Onslow Bay, North Carolina. Coastal and

  18. Review of the taxonomy of the genus Arthrobacter, emendation of the genus Arthrobacter sensu lato, proposal to reclassify selected species of the genus Arthrobacter in the novel genera Glutamicibacter gen. nov., Paeniglutamicibacter gen. nov., Pseudoglutamicibacter gen. nov., Paenarthrobacter gen. nov. and Pseudarthrobacter gen. nov., and emended description of Arthrobacter roseus.

    PubMed

    Busse, Hans-Jürgen

    2016-01-01

    In this paper, the taxonomy of the genus Arthrobacter is discussed, from its first description in 1947 to the present state. Emphasis is given to intrageneric phylogeny and chemotaxonomic characteristics, concentrating on quinone systems, peptidoglycan compositions and polar lipid profiles. Internal groups within the genus Arthrobacter indicated from homogeneous chemotaxonomic traits and corresponding to phylogenetic grouping and/or high 16S rRNA gene sequence similarities are highlighted. Furthermore, polar lipid profiles and quinone systems of selected species are shown, filling some gaps concerning these chemotaxonomic traits. Based on phylogenetic groupings, 16S rRNA gene sequence similarities and homogeneity in peptidoglycan types, quinone systems and polar lipid profiles, a description of the genus Arthrobacter sensu lato and an emended description of Arthrobacter roseus are provided. Furthermore, reclassifications of selected species of the genus Arthrobacter into novel genera are proposed, namely Glutamicibacter gen. nov. (nine species), Paeniglutamicibacter gen. nov. (six species), Pseudoglutamicibacter gen. nov. (two species), Paenarthrobacter gen. nov. (six species) and Pseudarthrobacter gen. nov. (ten species).

  19. Worldwide genetic features of HIV-1 Env α4β7 binding motif: the Local Dissemination Impact of the LDI tripeptide

    PubMed Central

    Hait, Sabrina H.; Soares, Esmeralda A.; Sprinz, Eduardo; Arthos, James; Machado, Elizabeth S.; Soares, Marcelo A.

    2015-01-01

    Background HIV-1 gp120 binds to integrin α4β7, a homing receptor of lymphocytes to gut-associated lymphoid tissues. This interaction is mediated by the LDI/V tripeptide encoded in the V2-loop. This tripeptide mimics similar motifs in MAdCAM and VCAM, the natural ligands of α4β7. In this study we explored the association of V2-loop LDI/V mimotopes with transmission routes and patterns of disease progression in HIV-infected adult and pediatric patients. HIV-1 env sequences available in the Los Alamos HIV Sequence Database were included in the analyses. Methodology HIV-1 V2-loop sequences generated from infected adults and infants from South and Southeast Brazil, and also retrieved from the Los Alamos Database, were assessed for α4β7 binding tripeptide composition. Chi-Square/Fisher’s Exact test and Mann Whitney U-test were used for tripeptide comparisons. Shannon entropy was assessed for conservancy of the α4β7 tripeptide mimotope. Results We observed no association between the tripeptide composition or conservation and virus transmission route or disease progression. However, LDI was linked to successful epidemic dissemination of HIV-1 subtype C in South America, and further to other expanding non-B subtypes in Europe and Asia. In Africa, subtypes showing increased LDV prevalence evidenced an ongoing process of selection towards LDI expansion, an observation also extended to subtype B in the Americas and Western Europe. Conclusions The V2-loop LDI mimotope was conserved in HIV-1C from South America and other expanding subtypes across the globe, which suggests that LDI may promote successful dissemination of HIV at local geographic levels by means of increased transmission fitness. PMID:26569174

  20. Evaluation of moisture and heat transport in the fast-response building-resolving urban transport code QUIC EnvSim

    NASA Astrophysics Data System (ADS)

    Briggs, Kevin A.

    QUIC EnvSim (QES) is a complete building-resolving urban microclimate modeling system developed to rapidly compute mass, momentum, and heat transport for the design of sustainable cities. One of the more computationally intensive components of this type of modeling system is the transport and dispersion of scalars. In this paper, we describe and evaluate QESTransport, a Reynolds-averaged Navier-Stokes (RANS) scalar transport model. QESTransport makes use of light-weight methods and modeling techniques. It is parallelized for Graphics Processing Units (GPUs), utilizing NVIDIA's OptiX application programming interfaces (APIs). QESTransport is coupled with the well-validated QUIC Dispersion Modeling system. To couple the models, a new methodology was implemented to efficiently prescribe surface flux boundary conditions on both vertical walls and flat surfaces. In addition, a new internal boundary layer parameterization was introduced into QUIC to enable the representation of momentum advection across changing surface conditions. QESTransport is validated against the following three experimental test cases designed to evaluate the model's performance under idealized conditions: (i) flow over a step change in moisture, roughness, and temperature, (ii) flow over an isolated heated building, and (iii) flow through an array of heated buildings. For all three cases, the model is compared against published simulation results. QESTransport produces velocity, temperature, and moisture fields that are comparable to much more complex numerical models for each case. The code execution time performance is evaluated and demonstrates linear scaling on a single GPU for problem sizes up to 4.5 x 4.5 km at 5 m grid resolution, and is found to produce results at much better than real time for a 1.2 x 1.2 km section of downtown Salt Lake City, Utah.

  1. Genetic analyses of HIV-1 env sequences demonstrate limited compartmentalization in breast milk and suggest viral replication within the breast that increases with mastitis.

    PubMed

    Gantt, Soren; Carlsson, Jacquelyn; Heath, Laura; Bull, Marta E; Shetty, Avinash K; Mutsvangwa, Junior; Musingwini, Georgina; Woelk, Godfrey; Zijenah, Lynn S; Katzenstein, David A; Mullins, James I; Frenkel, Lisa M

    2010-10-01

    The concentration of human immunodeficiency virus type 1 (HIV-1) is generally lower in breast milk than in blood. Mastitis, or inflammation of the breast, is associated with increased levels of milk HIV-1 and risk of mother-to-child transmission through breastfeeding. We hypothesized that mastitis facilitates the passage of HIV-1 from blood into milk or stimulates virus production within the breast. HIV-1 env sequences were generated from single amplicons obtained from breast milk and blood samples in a cross-sectional study. Viral compartmentalization was evaluated using several statistical methods, including the Slatkin and Maddison (SM) test. Mastitis was defined as an elevated milk sodium (Na(+)) concentration. The association between milk Na(+) and the pairwise genetic distance between milk and blood viral sequences was modeled using linear regression. HIV-1 was compartmentalized within milk by SM testing in 6/17 (35%) specimens obtained from 9 women, but all phylogenetic clades included viral sequences from milk and blood samples. Monotypic sequences were more prevalent in milk samples than in blood samples (22% versus 13%; P = 0.012), which accounted for half of the compartmentalization observed. Mastitis was not associated with compartmentalization by SM testing (P = 0.621), but Na(+) was correlated with greater genetic distance between milk and blood HIV-1 populations (P = 0.041). In conclusion, local production of HIV-1 within the breast is suggested by compartmentalization of virus and a higher prevalence of monotypic viruses in milk specimens. However, phylogenetic trees demonstrate extensive mixing of viruses between milk and blood specimens. HIV-1 replication in breast milk appears to increase with inflammation, contributing to higher milk viral loads during mastitis.

  2. Hunting for new restriction enzymes in GenBank

    SciTech Connect

    Roberts, R.J.

    1997-12-01

    Restriction enzyme genes are hard to identify unless their surrounding sequences are available. This is because the best definition of a restriction enzyme gene is an open reading frame, that looks like nothing else in GenBank, but lies close to a DNA methylase gene. There are other clues too, such as nearby reading frames that code control proteins or the DNA specificity subunits of Type I restriction enzymes. We are developing software that searches the daily updates of GenBank to find candidate restriction enzyme genes. This is being followed by bench experiments to see of the predictions are correct. More than 50 potential new restriction enzymes have been predicted and it is quite remarkable that the density of restriction enzyme genes in microbial DNA is greater than one system per 200 Kb. The software development is emphasizing the graphic presentation of the search results. The approach could be used for other situations where a molecular biologist is interested to find new examples of their favorite genes.

  3. Next gen wavelets down-sampling preserving statistics

    NASA Astrophysics Data System (ADS)

    Szu, Harold; Miao, Lidan; Chanyagon, Pornchai; Cader, Masud

    2007-04-01

    We extend the 2 nd Gen Discrete Wavelet Transform (DWT) of Swelden to the Next Generations (NG) Digital Wavelet Transform (DWT) preserving the statistical salient features. The lossless NG DWT accomplishes the data compression of "wellness baseline profiles (WBP)" of aging population at homes. For medical monitoring system at home fronts we translate the military experience to dual usage of veterans & civilian alike with the following three requirements: (i) Data Compression: The necessary down sampling reduces the immense amount of data of individual WBP from hours to days and to weeks for primary caretakers in terms of moments, e.g. mean value, variance, etc., without the artifacts caused by FFT arbitrary windowing. (ii) Lossless: our new NG_DWT must preserve the original data sets. (iii) Phase Transition: NG_DWT must capture the critical phase transition of the wellness toward the sickness with simultaneous display of local statistical moments. According to the Nyquist sampling theory, assuming a band-limited wellness physiology, we must sample the WBP at least twice per day since it is changing diurnally and seasonally. Since NG_DWT, like the 2 nd Gen, is lossless, we can reconstruct the original time series for the physicians' second looks. This technique of NG_DWT can also help stock market day-traders monitoring the volatility of multiple portfolios without artificial horizon artifacts.

  4. GenGIS: A geospatial information system for genomic data

    PubMed Central

    Parks, Donovan H.; Porter, Michael; Churcher, Sylvia; Wang, Suwen; Blouin, Christian; Whalley, Jacqueline; Brooks, Stephen; Beiko, Robert G.

    2009-01-01

    The increasing availability of genetic sequence data associated with explicit geographic and ecological information is offering new opportunities to study the processes that shape biodiversity. The generation and testing of hypotheses using these data sets requires effective tools for mathematical and visual analysis that can integrate digital maps, ecological data, and large genetic, genomic, or metagenomic data sets. GenGIS is a free and open-source software package that supports the integration of digital map data with genetic sequences and environmental information from multiple sample sites. Essential bioinformatic and statistical tools are integrated into the software, allowing the user a wide range of analysis options for their sequence data. Data visualizations are combined with the cartographic display to yield a clear view of the relationship between geography and genomic diversity, with a particular focus on the hierarchical clustering of sites based on their similarity or phylogenetic proximity. Here we outline the features of GenGIS and demonstrate its application to georeferenced microbial metagenomic, HIV-1, and human mitochondrial DNA data sets. PMID:19635847

  5. GenDecoder: genetic code prediction for metazoan mitochondria

    PubMed Central

    Abascal, Federico; Zardoya, Rafael; Posada, David

    2006-01-01

    Although the majority of the organisms use the same genetic code to translate DNA, several variants have been described in a wide range of organisms, both in nuclear and organellar systems, many of them corresponding to metazoan mitochondria. These variants are usually found by comparative sequence analyses, either conducted manually or with the computer. Basically, when a particular codon in a query-species is linked to positions for which a specific amino acid is consistently found in other species, then that particular codon is expected to translate as that specific amino acid. Importantly, and despite the simplicity of this approach, there are no available tools to help predicting the genetic code of an organism. We present here GenDecoder, a web server for the characterization and prediction of mitochondrial genetic codes in animals. The analysis of automatic predictions for 681 metazoans aimed us to study some properties of the comparative method, in particular, the relationship among sequence conservation, taxonomic sampling and reliability of assignments. Overall, the method is highly precise (99%), although highly divergent organisms such as platyhelminths are more problematic. The GenDecoder web server is freely available from . PMID:16845034

  6. VitisGen on the road: mapping the way to the next generation of grapes

    USDA-ARS?s Scientific Manuscript database

    The USDA-NIFA VitisGen project (Award No. 2011-51181-30635) aims to optimize grape cultivar improvement through the integration of grape breeding, cutting edge genomics technology, precision phenotyping, socioeconomic research, and extension. VitisGen is a five-year project involving 25 co-PIs at 11...

  7. 77 FR 54648 - Seventh Meeting: RTCA NextGen Advisory Committee (NAC)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-09-05

    ... Meeting: RTCA NextGen Advisory Committee (NAC) AGENCY: Federal Aviation Administration (FAA), U.S. Department of Transportation (DOT). ACTION: Meeting Notice of RTCA NextGen Advisory Committee (NAC). SUMMARY: The FAA is issuing this notice to advise the public of the seventh meeting of the RTCA...

  8. How is the Xpert MRSA Gen 3 assay (Cepheid) performing on pooled eSwab medium?

    PubMed

    Jonckheere, Stijn; Van Vaerenbergh, Kristien; Boel, An; Vankeerberghen, Anne; De Beenhouwer, Hans

    2015-11-01

    The performance of the Xpert MRSA Gen 3 was compared to the Xpert MRSA on pooled eSwab media from nose, throat, and perineum using broth enriched cultured as gold standard. A lower specificity was found for the Xpert MRSA Gen 3 compared to the Xpert MRSA (91.8% versus 97.9%; P<0.05).

  9. CottonGen: a genomics, genetics and breeding database for cotton research

    USDA-ARS?s Scientific Manuscript database

    CottonGen (http://www.cottongen.org) is a curated and integrated web-based relational database providing access to publicly available genomic, genetic and breeding data for cotton. CottonGen supercedes CottonDB and the Cotton Marker Database, with enhanced tools for easier data sharing, mining, vis...

  10. Moryella indoligenes gen. nov., sp. nov., an anaerobic bacterium isolated from clinical specimens.

    PubMed

    Carlier, Jean-Philippe; K'ouas, Guylène; Han, Xiang Y

    2007-04-01

    Three Gram-positive, anaerobic, non-spore-forming, rod-shaped bacteria with pointed ends were isolated from clinical specimens. The organisms were weakly saccharolytic and produced indole, acetate, butyrate and lactate as major metabolic end products. 16S rRNA gene sequence analysis indicated that the isolates had no known close relatives among recognized bacteria but that they exhibited a phylogenetic association with Clostridium rRNA cluster XIVa [as defined by Collins, M. D. et al. (1994). Int J Syst Bacteriol 44, 812-826]. The closest recognized relatives were the type strains of Clostridium clostridioforme, Clostridium bolteae and Clostridium asparagiforme (16S rRNA gene sequence similarity values of 90.2-91.4 %). These results suggest that these three clinical isolates represent a novel species of a new genus, for which the name Moryella indoligenes gen. nov., sp. nov. is proposed. The type strain of Moryella indoligenes is AIP 220.04(T) (=CIP 109174(T)=CCUG 52648(T)).

  11. Afrojoyeuxia gen. n. and Hunkeleriella gen. n., two new genera of cestodes (Cyclophyllidea: Anoplocephalidae) from African rodents.

    PubMed

    Haukisalmi, Voitto

    2013-11-01

    Based on the study of type material, two new genera of cestodes (Cyclophyllidea: Anoplocephalidae) are proposed for Paranoplocephala Lüihe, 1910 sensu lato species from African rodents. Afrojoyeuxia gen. n., proposed for A. gundii (Joyeux, 1923) comb. n. from Ctenodactylus gundi (Rothmann) (Hystricomorpha: Ctenodactylidae), is characterized by a high length/width ratio of mature proglottids, longitudinally extensive testicular field positioned anterior to the female glands, an ovoid or subspherical cirrus-sac and a thick, conical cirrus. Hunkeleriella gen. n., proposed for H. dasymidis (Hunkeler, 1972) comb. n. from Dasymys incomtus (Sundevall) (Myomorpha: Muridae), differs from related genera mainly by its short (10-20 mm) and wide strobila and neck, unilateral genital pores (exceptionally with a few changes per strobila), the position of the genital pores (slightly anterior to the middle of proglottid margin) and initially tube-like early uterus (later reticulated). Parandrya Gulyaev et Chechulin, 1996, earlier suggested to be a junior synonym of Paranoplocephala, is considered to be a valid, independent genus. Evidence of non-monophyly and need for a taxonomic revision of Paranoplocephala sensu lato, as well as the phylogenetic position of A. gundii and H. dasymidis are discussed.

  12. Abu al-Layth al-Libi

    DTIC Science & Technology

    2015-02-01

    jihadi doctrine, al-Libi enrolled himself in the recently erected and highly popular al-Faruq 2 “Al-Sahab Releases ‘Winds of Paradise , Part III...February 2008, http://news.bbc.co.uk/2/hi/south_asia/7220823.stm. 3 “Al-Sahab Releases ‘Winds of Paradise , Part III,’” Global Terrorism Research Project...5 “Al-Sahab Releases ‘Winds of Paradise , Part III,’” Global Terrorism Research Project. 6 Ibid. The date provided in the video is 1410

  13. What Is ALS?

    MedlinePlus

    ... Javits, actor David Niven, “Sesame Street” creator Jon Stone, boxing champion Ezzard Charles, NBA Hall of Fame ... Help for People with ALS and Caregivers Read stories from families living with ALS Forms of ALS ...

  14. What Can a Historian Do with AstroGen?

    NASA Astrophysics Data System (ADS)

    Tenn, Joseph S.

    2015-01-01

    "Astrogen", the Astronomy Genealogy Project, is in the development stage. Patterned after the Mathematics Genealogy Project at http://genealogy.math.ndsu.nodak.edu, it will eventually include most of the world's astronomers, past and present, and provide information about their years of life, highest degrees, universities, and thesis titles. There will also be links to online theses, home pages, and obituaries when these are available. Although a few details remain to be worked out before it becomes public, it is possible to make some use of what has already been compiled. I will give an example, comparing graduates of Harvard University, the University of California at Berkeley, and the University of Chicago from different decades, with information about their professional careers and publication records. The author welcomes queries about AstroGen and is seeking more participants.

  15. Two-dimensional MHD generator model. [GEN code

    SciTech Connect

    Geyer, H. K.; Ahluwalia, R. K.; Doss, E. D.

    1980-09-01

    A steady state, two-dimensional MHD generator code, GEN, is presented. The code solves the equations of conservation of mass, momentum, and energy, using a Von Mises transformation and a local linearization of the equations. By splitting the source terms into a part proportional to the axial pressure gradient and a part independent of the gradient, the pressure distribution along the channel is easily obtained to satisfy various criteria. Thus, the code can run effectively in both design modes, where the channel geometry is determined, and analysis modes, where the geometry is previously known. The code also employs a mixing length concept for turbulent flows, Cebeci and Chang's wall roughness model, and an extension of that model to the effective thermal diffusities. Results on code validation, as well as comparisons of skin friction and Stanton number calculations with experimental results, are presented.

  16. Commissioning and Performance Analysis of WhisperGen Stirling Engine

    NASA Astrophysics Data System (ADS)

    Pradip, Prashant Kaliram

    Stirling engine based cogeneration systems have potential to reduce energy consumption and greenhouse gas emission, due to their high cogeneration efficiency and emission control due to steady external combustion. To date, most studies on this unit have focused on performance based on both experimentation and computer models, and lack experimental data for diversified operating ranges. This thesis starts with the commissioning of a WhisperGen Stirling engine with components and instrumentation to evaluate power and thermal performance of the system. Next, a parametric study on primary engine variables, including air, diesel, and coolant flowrate and temperature were carried out to further understand their effect on engine power and efficiency. Then, this trend was validated with the thermodynamic model developed for the energy analysis of a Stirling cycle. Finally, the energy balance of the Stirling engine was compared without and with heat recovery from the engine block and the combustion chamber exhaust.

  17. Tracking the Emergence of Host-Specific Simian Immunodeficiency Virus env and nef Populations Reveals nef Early Adaptation and Convergent Evolution in Brain of Naturally Progressing Rhesus Macaques.

    PubMed

    Lamers, Susanna L; Nolan, David J; Rife, Brittany D; Fogel, Gary B; McGrath, Michael S; Burdo, Tricia H; Autissier, Patrick; Williams, Kenneth C; Goodenow, Maureen M; Salemi, Marco

    2015-08-01

    While a clear understanding of the events leading to successful establishment of host-specific viral populations and productive infection in the central nervous system (CNS) has not yet been reached, the simian immunodeficiency virus (SIV)-infected rhesus macaque provides a powerful model for the study of human immunodeficiency virus (HIV) intrahost evolution and neuropathogenesis. The evolution of the gp120 and nef genes, which encode two key proteins required for the establishment and maintenance of infection, was assessed in macaques that were intravenously inoculated with the same viral swarm and allowed to naturally progress to simian AIDS and potential SIV-associated encephalitis (SIVE). Longitudinal plasma samples and immune markers were monitored until terminal illness. Single-genome sequencing was employed to amplify full-length env through nef transcripts from plasma over time and from brain tissues at necropsy. nef sequences diverged from the founder virus faster than gp120 diverged. Host-specific sequence populations were detected in nef (~92 days) before they were detected in gp120 (~182 days). At necropsy, similar brain nef sequences were found in different macaques, indicating convergent evolution, while gp120 brain sequences remained largely host specific. Molecular clock and selection analyses showed weaker clock-like behavior and stronger selection pressure in nef than in gp120, with the strongest nef selection in the macaque with SIVE. Rapid nef diversification, occurring prior to gp120 diversification, indicates that early adaptation of nef in the new host is essential for successful infection. Moreover, the convergent evolution of nef sequences in the CNS suggests a significant role for nef in establishing neurotropic strains. The SIV-infected rhesus macaque model closely resembles HIV-1 immunopathogenesis, neuropathogen