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Sample records for chitinase-like protein-encoding atctl2

  1. Chitinase-like proteins as regulators of innate immunity and tissue repair: helpful lessons for asthma?

    Science.gov (United States)

    Sutherland, Tara E

    2018-02-19

    Chitinases and chitinase-like proteins (CLPs) belong to the glycoside hydrolase family 18 of proteins. Chitinases are expressed in mammals and lower organisms, facilitate chitin degradation, and hence act as host-defence enzymes. Gene duplication and loss-of-function mutations of enzymatically active chitinases have resulted in the expression of a diverse range of CLPs across different species. CLPs are genes that are increasingly associated with inflammation and tissue remodelling not only in mammals but also across distant species. While the focus has remained on understanding the functions and expression patterns of CLPs during disease in humans, studies in mouse and lower organisms have revealed important and overlapping roles of the CLP family during physiology, host defence and pathology. This review will summarise recent insights into the regulatory functions of CLPs on innate immune pathways and discuss how these effects are not only important for host defence and tissue injury/repair after pathogen invasion, but also how they have extensive implications for pathological processes involved in diseases such as asthma. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  2. The Drosophila chitinase-like protein IDGF3 is involved in protection against nematodes and woung healing

    Czech Academy of Sciences Publication Activity Database

    Kučerová, Lucie; Brož, Václav; Arefin, B.; Maaroufi, H. O.; Hurychová, J.; Strnad, Hynek; Žurovec, Michal; Theopold, U.

    2016-01-01

    Roč. 8, č. 2 (2016), s. 199-210 ISSN 1662-811X R&D Projects: GA ČR GA14-27816S Institutional support: RVO:60077344 ; RVO:68378050 Keywords : chitinase-like proteins * imaginal disc growth factor * hemolymph clot Subject RIV: CE - Biochemistry; EB - Genetics ; Molecular Biology (UMG-J) Impact factor: 3.938, year: 2016 http://www.karger.com/Article/FullText/442351

  3. The chitinase-like protein YKL-40 increases mucin5AC production in human bronchial epithelial cells

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    Liu, Chunyi; Li, Qi [Division of Respiratory Medicine, Second Affiliated Hospital, Chongqing Medical University, No. 74, Linjiang Road, Yuzhong District, Chongqing 400010 (China); Zhou, Xiangdong, E-mail: zxd999@263.net [Division of Respiratory Medicine, Second Affiliated Hospital, Chongqing Medical University, No. 74, Linjiang Road, Yuzhong District, Chongqing 400010 (China); Kolosov, Victor P.; Perelman, Juliy M. [Far Eastern Scientific Center of Physiology and Pathology of Respiration, Siberian Branch, Russian Academy of Medical Sciences, Blagoveshchensk (Russian Federation)

    2013-11-01

    Mucus overproduction is an important feature in patients with chronic inflammatory airway diseases. However, the regulatory mechanisms that mediate excessive mucin production remain elusive. Recently, the level of YKL-40, a chitinase-like protein, has been found to be significantly increased in chronic inflammatory airway diseases and has been shown to be associated with the severity of these diseases. In this study, we sought to explore the effect of YKL-40 on mucin5AC (MUC5AC) production in chronic inflammatory airway diseases and the potential signaling pathways involved in this process. We found that elevated YKL-40 levels increased the mRNA and protein expression of MUC5AC in a dose- and time-dependent manner, in association with the phosphorylation of extracellular signal-regulated kinase (ERK) and nuclear factor κB (NF-κB), reflecting their activation. These responses were significantly suppressed by the knockdown of protease-activating receptor 2 (PAR2) with specific small interfering RNA or the inhibitors of ERK and NF-κB. YKL-40-induced MUC5AC overproduction was also effectively attenuated by the inhibitor of focal adhesion kinase (FAK). Taken together, these results imply that YKL-40 can stimulate excessive MUC5AC production through PAR2- and FAK-mediated mechanisms. - Highlights: • MUC5AC is the major secreted mucin in chronic inflammatory airway diseases. • YKL-40 is a prototype of the chitinase-like protein in mammals. • YKL-40 is an active player in chronic inflammatory airway diseases. • YKL-40 can increase MUC5AC production via PAR2-mediated pathway. • FAK is another candidate to mediate YKL-40-induced MUC5AC overexpression.

  4. The chitinase-like protein YKL-40 increases mucin5AC production in human bronchial epithelial cells

    International Nuclear Information System (INIS)

    Liu, Chunyi; Li, Qi; Zhou, Xiangdong; Kolosov, Victor P.; Perelman, Juliy M.

    2013-01-01

    Mucus overproduction is an important feature in patients with chronic inflammatory airway diseases. However, the regulatory mechanisms that mediate excessive mucin production remain elusive. Recently, the level of YKL-40, a chitinase-like protein, has been found to be significantly increased in chronic inflammatory airway diseases and has been shown to be associated with the severity of these diseases. In this study, we sought to explore the effect of YKL-40 on mucin5AC (MUC5AC) production in chronic inflammatory airway diseases and the potential signaling pathways involved in this process. We found that elevated YKL-40 levels increased the mRNA and protein expression of MUC5AC in a dose- and time-dependent manner, in association with the phosphorylation of extracellular signal-regulated kinase (ERK) and nuclear factor κB (NF-κB), reflecting their activation. These responses were significantly suppressed by the knockdown of protease-activating receptor 2 (PAR2) with specific small interfering RNA or the inhibitors of ERK and NF-κB. YKL-40-induced MUC5AC overproduction was also effectively attenuated by the inhibitor of focal adhesion kinase (FAK). Taken together, these results imply that YKL-40 can stimulate excessive MUC5AC production through PAR2- and FAK-mediated mechanisms. - Highlights: • MUC5AC is the major secreted mucin in chronic inflammatory airway diseases. • YKL-40 is a prototype of the chitinase-like protein in mammals. • YKL-40 is an active player in chronic inflammatory airway diseases. • YKL-40 can increase MUC5AC production via PAR2-mediated pathway. • FAK is another candidate to mediate YKL-40-induced MUC5AC overexpression

  5. Chitinase-like (CTL) and cellulose synthase (CESA) gene expression in gelatinous-type cellulosic walls of flax (Linum usitatissimum L.) bast fibers.

    Science.gov (United States)

    Mokshina, Natalia; Gorshkova, Tatyana; Deyholos, Michael K

    2014-01-01

    Plant chitinases (EC 3.2.1.14) and chitinase-like (CTL) proteins have diverse functions including cell wall biosynthesis and disease resistance. We analyzed the expression of 34 chitinase and chitinase-like genes of flax (collectively referred to as LusCTLs), belonging to glycoside hydrolase family 19 (GH19). Analysis of the transcript expression patterns of LusCTLs in the stem and other tissues identified three transcripts (LusCTL19, LusCTL20, LusCTL21) that were highly enriched in developing bast fibers, which form cellulose-rich gelatinous-type cell walls. The same three genes had low relative expression in tissues with primary cell walls and in xylem, which forms a xylan type of secondary cell wall. Phylogenetic analysis of the LusCTLs identified a flax-specific sub-group that was not represented in any of other genomes queried. To provide further context for the gene expression analysis, we also conducted phylogenetic and expression analysis of the cellulose synthase (CESA) family genes of flax, and found that expression of secondary wall-type LusCESAs (LusCESA4, LusCESA7 and LusCESA8) was correlated with the expression of two LusCTLs (LusCTL1, LusCTL2) that were the most highly enriched in xylem. The expression of LusCTL19, LusCTL20, and LusCTL21 was not correlated with that of any CESA subgroup. These results defined a distinct type of CTLs that may have novel functions specific to the development of the gelatinous (G-type) cellulosic walls.

  6. A Phenotyping Method of Giant Cells from Root-Knot Nematode Feeding Sites by Confocal Microscopy Highlights a Role for CHITINASE-LIKE 1 in Arabidopsis

    Science.gov (United States)

    Cabrera, Javier; Olmo, Rocio; Ruiz-Ferrer, Virginia; Hermans, Christian; Martinez-Argudo, Isabel; Escobar, Carolina

    2018-01-01

    Most effective nematicides for the control of root-knot nematodes are banned, which demands a better understanding of the plant-nematode interaction. Understanding how gene expression in the nematode-feeding sites relates to morphological features may assist a better characterization of the interaction. However, nematode-induced galls resulting from cell-proliferation and hypertrophy hinders such observation, which would require tissue sectioning or clearing. We demonstrate that a method based on the green auto-fluorescence produced by glutaraldehyde and the tissue-clearing properties of benzyl-alcohol/benzyl-benzoate preserves the structure of the nematode-feeding sites and the plant-nematode interface with unprecedented resolution quality. This allowed us to obtain detailed measurements of the giant cells’ area in an Arabidopsis line overexpressing CHITINASE-LIKE-1 (CTL1) from optical sections by confocal microscopy, assigning a role for CTL1 and adding essential data to the scarce information of the role of gene repression in giant cells. Furthermore, subcellular structures and features of the nematodes body and tissues from thick organs formed after different biotic interactions, i.e., galls, syncytia, and nodules, were clearly distinguished without embedding or sectioning in different plant species (Arabidopsis, cucumber or Medicago). The combination of this method with molecular studies will be valuable for a better understanding of the plant-biotic interactions. PMID:29389847

  7. Sequence analysis and gene expression of putative oil palm chitinase and chitinase-like proteins in response to colonization of Ganoderma boninense and Trichoderma harzianum.

    Science.gov (United States)

    Yeoh, K-A; Othman, A; Meon, S; Abdullah, F; Ho, C-L

    2013-01-01

    Chitinases are glycosyl hydrolases that cleave the β-1,4-glycosidic linkages between N-acetylglucosamine residues in chitin which is a major component of fungal cell wall. Plant chitinases hydrolyze fungal chitin to chitin oligosaccharides that serve as elicitors of plant defense system against fungal pathogens. However, plants synthesize many chitinase isozymes and some of them are not pathogenesis-related. In this study, three full-length cDNA sequences encoding a putative chitinase (EgChit3-1) and two chitinase-like proteins (EgChit1-1 and EgChit5-1) have been cloned from oil palm (Elaeis guineensis) by polymerase chain reaction (PCR). The abundance of these transcripts in the roots and leaves of oil palm seedlings treated with Ganoderma boninense (a fungal pathogen) or Trichoderma harzianum (an avirulent symbiont), and a combination of both fungi at 3, 6 and 12 weeks post infection were profiled by real time quantitative reverse-transcription (qRT)-PCR. Our findings showed that the gene expression of EgChit3-1 increased significantly in the roots of oil palm seedlings treated with either G. boninense or T. harzianum and a combination of both; whereas the gene expression of EgChit1-1 in the treated roots of oil palm seedlings was not significantly higher compared to those of the untreated oil palm roots. The gene expression of EgChit5-1 was only higher in the roots of oil palm seedlings treated with T. harzianum compared to those of the untreated oil palm roots. In addition, the gene expression of EgChit1-1 and EgChit3-1 showed a significantly higher gene expression in the leaf samples of oil palm seedlings treated with either G. boninense or T. harzianum.

  8. The presence of two S-layer-protein-encoding genes is conserved among species related to Lactobacillus acidophilus

    NARCIS (Netherlands)

    Boot, H.J.; Kolen, C.P.A.M.; Pot, B.; Kersters, K.; Pouwels, P.H.

    1996-01-01

    Previously we have shown that the type strain of Lactobacillus acidophilus possesses two S-protein-encoding genes, one of which is silent, on a chromosomal segment of 6 kb. The S-protein-encoding gene in the expression site can be exchanged for the silent S-protein-encoding gene by inversion of this

  9. Molecular evolution of the Paramyxoviridae and Rhabdoviridae multiple-protein-encoding P gene.

    Science.gov (United States)

    Jordan, I K; Sutter, B A; McClure, M A

    2000-01-01

    Presented here is an analysis of the molecular evolutionary dynamics of the P gene among 76 representative sequences of the Paramyxoviridae and Rhabdoviridae RNA virus families. In a number of Paramyxoviridae taxa, as well as in vesicular stomatitis viruses of the Rhabdoviridae, the P gene encodes multiple proteins from a single genomic RNA sequence. These products include the phosphoprotein (P), as well as the C and V proteins. The complexity of the P gene makes it an intriguing locus to study from an evolutionary perspective. Amino acid sequence alignments of the proteins encoded at the P and N loci were used in independent phylogenetic reconstructions of the Paramyxoviridae and Rhabdoviridae families. P-gene-coding capacities were mapped onto the Paramyxoviridae phylogeny, and the most parsimonious path of multiple-coding-capacity evolution was determined. Levels of amino acid variation for Paramyxoviridae and Rhabdoviridae P-gene-encoded products were also analyzed. Proteins encoded in overlapping reading frames from the same nucleotides have different levels of amino acid variation. The nucleotide architecture that underlies the amino acid variation was determined in order to evaluate the role of selection in the evolution of the P gene overlapping reading frames. In every case, the evolution of one of the proteins encoded in the overlapping reading frames has been constrained by negative selection while the other has evolved more rapidly. The integrity of the overlapping reading frame that represents a derived state is generally maintained at the expense of the ancestral reading frame encoded by the same nucleotides. The evolution of such multicoding sequences is likely a response by RNA viruses to selective pressure to maximize genomic information content while maintaining small genome size. The ability to evolve such a complex genomic strategy is intimately related to the dynamics of the viral quasispecies, which allow enhanced exploration of the adaptive

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

    International Nuclear Information System (INIS)

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

    1988-01-01

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

  11. Relationships between protein-encoding gene abundance and corresponding process are commonly assumed yet rarely observed

    Science.gov (United States)

    Rocca, Jennifer D.; Hall, Edward K.; Lennon, Jay T.; Evans, Sarah E.; Waldrop, Mark P.; Cotner, James B.; Nemergut, Diana R.; Graham, Emily B.; Wallenstein, Matthew D.

    2015-01-01

    For any enzyme-catalyzed reaction to occur, the corresponding protein-encoding genes and transcripts are necessary prerequisites. Thus, a positive relationship between the abundance of gene or transcripts and corresponding process rates is often assumed. To test this assumption, we conducted a meta-analysis of the relationships between gene and/or transcript abundances and corresponding process rates. We identified 415 studies that quantified the abundance of genes or transcripts for enzymes involved in carbon or nitrogen cycling. However, in only 59 of these manuscripts did the authors report both gene or transcript abundance and rates of the appropriate process. We found that within studies there was a significant but weak positive relationship between gene abundance and the corresponding process. Correlations were not strengthened by accounting for habitat type, differences among genes or reaction products versus reactants, suggesting that other ecological and methodological factors may affect the strength of this relationship. Our findings highlight the need for fundamental research on the factors that control transcription, translation and enzyme function in natural systems to better link genomic and transcriptomic data to ecosystem processes.

  12. [Cloning, mutagenesis and symbiotic phenotype of three lipid transfer protein encoding genes from Mesorhizobium huakuii 7653R].

    Science.gov (United States)

    Li, Yanan; Zeng, Xiaobo; Zhou, Xuejuan; Li, Youguo

    2016-12-04

    Lipid transfer protein superfamily is involved in lipid transport and metabolism. This study aimed to construct mutants of three lipid transfer protein encoding genes in Mesorhizobium huakuii 7653R, and to study the phenotypes and function of mutations during symbiosis with Astragalus sinicus. We used bioinformatics to predict structure characteristics and biological functions of lipid transfer proteins, and conducted semi-quantitative and fluorescent quantitative real-time PCR to analyze the expression levels of target genes in free-living and symbiotic conditions. Using pK19mob insertion mutagenesis to construct mutants, we carried out pot plant experiments to observe symbiotic phenotypes. MCHK-5577, MCHK-2172 and MCHK-2779 genes encoding proteins belonged to START/RHO alpha_C/PITP/Bet_v1/CoxG/CalC (SRPBCC) superfamily, involved in lipid transport or metabolism, and were identical to M. loti at 95% level. Gene relative transcription level of the three genes all increased compared to free-living condition. We obtained three mutants. Compared with wild-type 7653R, above-ground biomass of plants and nodulenitrogenase activity induced by the three mutants significantly decreased. Results indicated that lipid transfer protein encoding genes of Mesorhizobium huakuii 7653R may play important roles in symbiotic nitrogen fixation, and the mutations significantly affected the symbiotic phenotypes. The present work provided a basis to study further symbiotic function mechanism associated with lipid transfer proteins from rhizobia.

  13. Overexpression of chitinase like protein YKL-40 in leukemia patients

    Directory of Open Access Journals (Sweden)

    Anil K. Hurmale

    2013-01-01

    Full Text Available YKL-40 is a member of mammalian chitanase (CHI3L1, expressed and secreted by several types of solid tumor cells, inflammatory cells and stem cells. The precise physiological role of YKL-40 in cancer is still not clear and it is suggested that it play a role in cancer cell proliferation, differentiation, metastatic potential, cell attachment and migration, reorganization and tissue remodeling.The aim of the study was to check the appearance of YKL-40 in leukemic cells and over-expression of YKL-40 in the plasma of leukemia patients in comparison to healthy controls, and find whether YKL-40 could serve as a peripheral biomarker for leukemia. The study was conducted between July 2012 and March 2013 and included 67 volunteers, 55 having leukemia at the stage of diagnosis ofthe disease and 12 normal healthy volunteers. YKL-40 levels were determined in all plasma samples using the YKL-40 enzyme-linked immunosorbent assay (ELISA kit and expression of YKL-40 was observed by using immunocytochemical (ICC analysis. YKL-40 plasma levels differed significantly between patients with leukemia and the normal healthy volunteers (P=<0.001 and YKL-40 was positively expressed in all four types of leukemia (AML, ALL, CLL and CML specimens.

  14. The protein encoded by the proto-oncogene DEK changes the topology of chromatin and reduces the efficiency of DNA replication in a chromatin-specific manner

    DEFF Research Database (Denmark)

    Alexiadis, V; Waldmann, T; Andersen, Jens S.

    2000-01-01

    The structure of chromatin regulates the genetic activity of the underlying DNA sequence. We report here that the protein encoded by the proto-oncogene DEK, which is involved in acute myelogenous leukemia, induces alterations of the superhelical density of DNA in chromatin. The change in topology...

  15. Diverse replication-associated protein encoding circular DNA viruses in guano samples of Central-Eastern European bats.

    Science.gov (United States)

    Kemenesi, Gábor; Kurucz, Kornélia; Zana, Brigitta; Földes, Fanni; Urbán, Péter; Vlaschenko, Anton; Kravchenko, Kseniia; Budinski, Ivana; Szodoray-Parádi, Farkas; Bücs, Szilárd; Jére, Csaba; Csősz, István; Szodoray-Parádi, Abigél; Estók, Péter; Görföl, Tamás; Boldogh, Sándor; Jakab, Ferenc

    2018-03-01

    Circular replication-associated protein encoding single-stranded DNA (CRESS DNA) viruses are increasingly recognized worldwide in a variety of samples. Representative members include well-described veterinary pathogens with worldwide distribution, such as porcine circoviruses or beak and feather disease virus. In addition, numerous novel viruses belonging to the family Circoviridae with unverified pathogenic roles have been discovered in different human samples. Viruses of the family Genomoviridae have also been described as being highly abundant in different faecal and environmental samples, with case reports showing them to be suspected pathogens in human infections. In order to investigate the genetic diversity of these viruses in European bat populations, we tested guano samples from Georgia, Hungary, Romania, Serbia and Ukraine. This resulted in the detection of six novel members of the family Circoviridae and two novel members of the family Genomoviridae. Interestingly, a gemini-like virus, namely niminivirus, which was originally found in raw sewage samples in Nigeria, was also detected in our samples. We analyzed the nucleotide composition of members of the family Circoviridae to determine the possible host origins of these viruses. This study provides the first dataset on CRESS DNA viruses of European bats, and members of several novel viral species were discovered.

  16. Molecular adaptation within the coat protein-encoding gene of Tunisian almond isolates of Prunus necrotic ringspot virus.

    Science.gov (United States)

    Boulila, Moncef; Ben Tiba, Sawssen; Jilani, Saoussen

    2013-04-01

    The sequence alignments of five Tunisian isolates of Prunus necrotic ringspot virus (PNRSV) were searched for evidence of recombination and diversifying selection. Since failing to account for recombination can elevate the false positive error rate in positive selection inference, a genetic algorithm (GARD) was used first and led to the detection of potential recombination events in the coat protein-encoding gene of that virus. The Recco algorithm confirmed these results by identifying, additionally, the potential recombinants. For neutrality testing and evaluation of nucleotide polymorphism in PNRSV CP gene, Tajima's D, and Fu and Li's D and F statistical tests were used. About selection inference, eight algorithms (SLAC, FEL, IFEL, REL, FUBAR, MEME, PARRIS, and GA branch) incorporated in HyPhy package were utilized to assess the selection pressure exerted on the expression of PNRSV capsid. Inferred phylogenies pointed out, in addition to the three classical groups (PE-5, PV-32, and PV-96), the delineation of a fourth cluster having the new proposed designation SW6, and a fifth clade comprising four Tunisian PNRSV isolates which underwent recombination and selective pressure and to which the name Tunisian outgroup was allocated.

  17. Domestication process of the goat revealed by an analysis of the nearly complete mitochondrial protein-encoding genes.

    Directory of Open Access Journals (Sweden)

    Koh Nomura

    Full Text Available Goats (Capra hircus are one of the oldest domesticated species, and they are kept all over the world as an essential resource for meat, milk, and fiber. Although recent archeological and molecular biological studies suggested that they originated in West Asia, their domestication processes such as the timing of population expansion and the dynamics of their selection pressures are little known. With the aim of addressing these issues, the nearly complete mitochondrial protein-encoding genes were determined from East, Southeast, and South Asian populations. Our coalescent time estimations suggest that the timing of their major population expansions was in the Late Pleistocene and significantly predates the beginning of their domestication in the Neolithic era (≈10,000 years ago. The ω (ratio of non-synonymous rate/synonymous substitution rate for each lineage was also estimated. We found that the ω of the globally distributed haplogroup A which is inherited by more than 90% of goats examined, turned out to be extremely low, suggesting that they are under severe selection pressure probably due to their large population size. Conversely, the ω of the Asian-specific haplogroup B inherited by about 5% of goats was relatively high. Although recent molecular studies suggest that domestication of animals may tend to relax selective constraints, the opposite pattern observed in our goat mitochondrial genome data indicates the process of domestication is more complex than may be presently appreciated and cannot be explained only by a simple relaxation model.

  18. The P0 protein encoded by cotton leafroll dwarf virus (CLRDV) inhibits local but not systemic RNA silencing.

    Science.gov (United States)

    Delfosse, Verónica C; Agrofoglio, Yamila C; Casse, María F; Kresic, Iván Bonacic; Hopp, H Esteban; Ziegler-Graff, Véronique; Distéfano, Ana J

    2014-02-13

    Plants employ RNA silencing as a natural defense mechanism against viruses. As a counter-defense, viruses encode silencing suppressor proteins (SSPs) that suppress RNA silencing. Most, but not all, the P0 proteins encoded by poleroviruses have been identified as SSP. In this study, we demonstrated that cotton leafroll dwarf virus (CLRDV, genus Polerovirus) P0 protein suppressed local silencing that was induced by sense or inverted repeat transgenes in Agrobacterium co-infiltration assay in Nicotiana benthamiana plants. A CLRDV full-length infectious cDNA clone that is able to infect N. benthamiana through Agrobacterium-mediated inoculation also inhibited local silencing in co-infiltration assays, suggesting that the P0 protein exhibits similar RNA silencing suppression activity when expressed from the full-length viral genome. On the other hand, the P0 protein did not efficiently inhibit the spread of systemic silencing signals. Moreover, Northern blotting indicated that the P0 protein inhibits the generation of secondary but not primary small interfering RNAs. The study of CLRDV P0 suppression activity may contribute to understanding the molecular mechanisms involved in the induction of cotton blue disease by CLRDV infection. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. Identification of human microRNA-like sequences embedded within the protein-encoding genes of the human immunodeficiency virus.

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    Bryan Holland

    Full Text Available BACKGROUND: MicroRNAs (miRNAs are highly conserved, short (18-22 nts, non-coding RNA molecules that regulate gene expression by binding to the 3' untranslated regions (3'UTRs of mRNAs. While numerous cellular microRNAs have been associated with the progression of various diseases including cancer, miRNAs associated with retroviruses have not been well characterized. Herein we report identification of microRNA-like sequences in coding regions of several HIV-1 genomes. RESULTS: Based on our earlier proteomics and bioinformatics studies, we have identified 8 cellular miRNAs that are predicted to bind to the mRNAs of multiple proteins that are dysregulated during HIV-infection of CD4+ T-cells in vitro. In silico analysis of the full length and mature sequences of these 8 miRNAs and comparisons with all the genomic and subgenomic sequences of HIV-1 strains in global databases revealed that the first 18/18 sequences of the mature hsa-miR-195 sequence (including the short seed sequence, matched perfectly (100%, or with one nucleotide mismatch, within the envelope (env genes of five HIV-1 genomes from Africa. In addition, we have identified 4 other miRNA-like sequences (hsa-miR-30d, hsa-miR-30e, hsa-miR-374a and hsa-miR-424 within the env and the gag-pol encoding regions of several HIV-1 strains, albeit with reduced homology. Mapping of the miRNA-homologues of env within HIV-1 genomes localized these sequence to the functionally significant variable regions of the env glycoprotein gp120 designated V1, V2, V4 and V5. CONCLUSIONS: We conclude that microRNA-like sequences are embedded within the protein-encoding regions of several HIV-1 genomes. Given that the V1 to V5 regions of HIV-1 envelopes contain specific, well-characterized domains that are critical for immune responses, virus neutralization and disease progression, we propose that the newly discovered miRNA-like sequences within the HIV-1 genomes may have evolved to self-regulate survival of the

  20. Variant Exported Blood-Stage Proteins Encoded by Plasmodium Multigene Families Are Expressed in Liver Stages Where They Are Exported into the Parasitophorous Vacuole.

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    Aurélie Fougère

    2016-11-01

    Full Text Available Many variant proteins encoded by Plasmodium-specific multigene families are exported into red blood cells (RBC. P. falciparum-specific variant proteins encoded by the var, stevor and rifin multigene families are exported onto the surface of infected red blood cells (iRBC and mediate interactions between iRBC and host cells resulting in tissue sequestration and rosetting. However, the precise function of most other Plasmodium multigene families encoding exported proteins is unknown. To understand the role of RBC-exported proteins of rodent malaria parasites (RMP we analysed the expression and cellular location by fluorescent-tagging of members of the pir, fam-a and fam-b multigene families. Furthermore, we performed phylogenetic analyses of the fam-a and fam-b multigene families, which indicate that both families have a history of functional differentiation unique to RMP. We demonstrate for all three families that expression of family members in iRBC is not mutually exclusive. Most tagged proteins were transported into the iRBC cytoplasm but not onto the iRBC plasma membrane, indicating that they are unlikely to play a direct role in iRBC-host cell interactions. Unexpectedly, most family members are also expressed during the liver stage, where they are transported into the parasitophorous vacuole. This suggests that these protein families promote parasite development in both the liver and blood, either by supporting parasite development within hepatocytes and erythrocytes and/or by manipulating the host immune response. Indeed, in the case of Fam-A, which have a steroidogenic acute regulatory-related lipid transfer (START domain, we found that several family members can transfer phosphatidylcholine in vitro. These observations indicate that these proteins may transport (host phosphatidylcholine for membrane synthesis. This is the first demonstration of a biological function of any exported variant protein family of rodent malaria parasites.

  1. CHIR99021 promotes self-renewal of mouse embryonic stem cells by modulation of protein-encoding gene and long intergenic non-coding RNA expression

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Yongyan [College of Veterinary Medicine, Northwest A and F University, Yangling 712100, Shaanxi (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A and F University, Yangling 712100, Shaanxi (China); Ai, Zhiying [Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A and F University, Yangling 712100, Shaanxi (China); College of Life Sciences, Northwest A and F University, Yangling 712100, Shaanxi (China); Yao, Kezhen [College of Veterinary Medicine, Northwest A and F University, Yangling 712100, Shaanxi (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A and F University, Yangling 712100, Shaanxi (China); Cao, Lixia; Du, Juan; Shi, Xiaoyan [Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A and F University, Yangling 712100, Shaanxi (China); College of Life Sciences, Northwest A and F University, Yangling 712100, Shaanxi (China); Guo, Zekun, E-mail: gzk@nwsuaf.edu.cn [College of Veterinary Medicine, Northwest A and F University, Yangling 712100, Shaanxi (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A and F University, Yangling 712100, Shaanxi (China); Zhang, Yong, E-mail: zhylab@hotmail.com [College of Veterinary Medicine, Northwest A and F University, Yangling 712100, Shaanxi (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A and F University, Yangling 712100, Shaanxi (China)

    2013-10-15

    Embryonic stem cells (ESCs) can proliferate indefinitely in vitro and differentiate into cells of all three germ layers. These unique properties make them exceptionally valuable for drug discovery and regenerative medicine. However, the practical application of ESCs is limited because it is difficult to derive and culture ESCs. It has been demonstrated that CHIR99021 (CHIR) promotes self-renewal and enhances the derivation efficiency of mouse (m)ESCs. However, the downstream targets of CHIR are not fully understood. In this study, we identified CHIR-regulated genes in mESCs using microarray analysis. Our microarray data demonstrated that CHIR not only influenced the Wnt/β-catenin pathway by stabilizing β-catenin, but also modulated several other pluripotency-related signaling pathways such as TGF-β, Notch and MAPK signaling pathways. More detailed analysis demonstrated that CHIR inhibited Nodal signaling, while activating bone morphogenetic protein signaling in mESCs. In addition, we found that pluripotency-maintaining transcription factors were up-regulated by CHIR, while several developmental-related genes were down-regulated. Furthermore, we found that CHIR altered the expression of epigenetic regulatory genes and long intergenic non-coding RNAs. Quantitative real-time PCR results were consistent with microarray data, suggesting that CHIR alters the expression pattern of protein-encoding genes (especially transcription factors), epigenetic regulatory genes and non-coding RNAs to establish a relatively stable pluripotency-maintaining network. - Highlights: • Combined use of CHIR with LIF promotes self-renewal of J1 mESCs. • CHIR-regulated genes are involved in multiple pathways. • CHIR inhibits Nodal signaling and promotes Bmp4 expression to activate BMP signaling. • Expression of epigenetic regulatory genes and lincRNAs is altered by CHIR.

  2. CHIR99021 promotes self-renewal of mouse embryonic stem cells by modulation of protein-encoding gene and long intergenic non-coding RNA expression

    International Nuclear Information System (INIS)

    Wu, Yongyan; Ai, Zhiying; Yao, Kezhen; Cao, Lixia; Du, Juan; Shi, Xiaoyan; Guo, Zekun; Zhang, Yong

    2013-01-01

    Embryonic stem cells (ESCs) can proliferate indefinitely in vitro and differentiate into cells of all three germ layers. These unique properties make them exceptionally valuable for drug discovery and regenerative medicine. However, the practical application of ESCs is limited because it is difficult to derive and culture ESCs. It has been demonstrated that CHIR99021 (CHIR) promotes self-renewal and enhances the derivation efficiency of mouse (m)ESCs. However, the downstream targets of CHIR are not fully understood. In this study, we identified CHIR-regulated genes in mESCs using microarray analysis. Our microarray data demonstrated that CHIR not only influenced the Wnt/β-catenin pathway by stabilizing β-catenin, but also modulated several other pluripotency-related signaling pathways such as TGF-β, Notch and MAPK signaling pathways. More detailed analysis demonstrated that CHIR inhibited Nodal signaling, while activating bone morphogenetic protein signaling in mESCs. In addition, we found that pluripotency-maintaining transcription factors were up-regulated by CHIR, while several developmental-related genes were down-regulated. Furthermore, we found that CHIR altered the expression of epigenetic regulatory genes and long intergenic non-coding RNAs. Quantitative real-time PCR results were consistent with microarray data, suggesting that CHIR alters the expression pattern of protein-encoding genes (especially transcription factors), epigenetic regulatory genes and non-coding RNAs to establish a relatively stable pluripotency-maintaining network. - Highlights: • Combined use of CHIR with LIF promotes self-renewal of J1 mESCs. • CHIR-regulated genes are involved in multiple pathways. • CHIR inhibits Nodal signaling and promotes Bmp4 expression to activate BMP signaling. • Expression of epigenetic regulatory genes and lincRNAs is altered by CHIR

  3. Mimic Phosphorylation of a βC1 Protein Encoded by TYLCCNB Impairs Its Functions as a Viral Suppressor of RNA Silencing and a Symptom Determinant.

    Science.gov (United States)

    Zhong, Xueting; Wang, Zhan Qi; Xiao, Ruyuan; Cao, Linge; Wang, Yaqin; Xie, Yan; Zhou, Xueping

    2017-08-15

    Phosphorylation of the βC1 protein encoded by the betasatellite of tomato yellow leaf curl China virus (TYLCCNB-βC1) by SNF1-related protein kinase 1 (SnRK1) plays a critical role in defense of host plants against geminivirus infection in Nicotiana benthamiana However, how phosphorylation of TYLCCNB-βC1 impacts its pathogenic functions during viral infection remains elusive. In this study, we identified two additional tyrosine residues in TYLCCNB-βC1 that are phosphorylated by SnRK1. The effects of TYLCCNB-βC1 phosphorylation on its functions as a viral suppressor of RNA silencing (VSR) and a symptom determinant were investigated via phosphorylation mimic mutants in N. benthamiana plants. Mutations that mimic phosphorylation of TYLCCNB-βC1 at tyrosine 5 and tyrosine 110 attenuated disease symptoms during viral infection. The phosphorylation mimics weakened the ability of TYLCCNB-βC1 to reverse transcriptional gene silencing and to suppress posttranscriptional gene silencing and abolished its interaction with N. benthamiana ASYMMETRIC LEAVES 1 in N. benthamiana leaves. The mimic phosphorylation of TYLCCNB-βC1 had no impact on its protein stability, subcellular localization, or self-association. Our data establish an inhibitory effect of phosphorylation of TYLCCNB-βC1 on its pathogenic functions as a VSR and a symptom determinant and provide a mechanistic explanation of how SnRK1 functions as a host defense factor. IMPORTANCE Tomato yellow leaf curl China virus (TYLCCNV), which causes a severe yellow leaf curl disease in China, is a monopartite geminivirus associated with the betasatellite (TYLCCNB). TYLCCNB encodes a single pathogenicity protein, βC1 (TYLCCNB-βC1), which functions as both a viral suppressor of RNA silencing (VSR) and a symptom determinant. Here, we show that mimicking phosphorylation of TYLCCNB-βC1 weakens its ability to reverse transcriptional gene silencing, to suppress posttranscriptional gene silencing, and to interact with N

  4. In silico Identification of Novel Chitinase-Like Proteins in the Silkworm, Bombyx mori, Genome

    Science.gov (United States)

    Pan, Ye; Lü, Peng; Wang, Yong; Yin, Lijing; Ma, Hexiang; Ma, Guohong; Chen, Keping; He, Yuanqing

    2012-01-01

    In insects, chitinases participate in the periodic shedding of old exoskeletons and the turnover of peritrophic membranes. Chitinase family members have been identified in dozens of species, including Tribolium castaneum, Drosophila melanogaster, and Anopheles gambiae. In this study, nine chitinases and three hypothetical chitinases have been identified in Bombyx mori L. (Lepidoptera: Bombycidae) through genome-wide searching. Phylogenetic analyses revealed that seven of them belong to the seven chitinase groups, respectively. BmCht25 and BmCht26 could not be grouped into the known chitinase groups, and might belong to two new groups of the chitinase family. BmCht10, BmCht25, and BmIDGF have glutamate amino acid substitutions in the active catalytic domain. Only BmCht5 and BmCht10 contain CBD domain and PEST sequences (rich in proline, glutamic acid, serine, and threonine). BmCht5 and BmCht26 are located on chromosome 7, and others (BmCht6, BmCht7, BmCht10, BmCht11, BmCht20, BmIDGF) are located on separate chromosomes of Bombyx mori, respectively. The present study provides important background information for future studies using Bombyx mori as a model organism for insect development and virus and host interaction. PMID:23461297

  5. Molecular mechanisms for protein-encoded inheritance

    Science.gov (United States)

    Wiltzius, Jed J. W.; Landau, Meytal; Nelson, Rebecca; Sawaya, Michael R.; Apostol, Marcin I.; Goldschmidt, Lukasz; Soriaga, Angela B.; Cascio, Duilio; Rajashankar, Kanagalaghatta; Eisenberg, David

    2013-01-01

    Strains are phenotypic variants, encoded by nucleic acid sequences in chromosomal inheritance and by protein “conformations” in prion inheritance and transmission. But how is a protein “conformation” stable enough to endure transmission between cells or organisms? Here new polymorphic crystal structures of segments of prion and other amyloid proteins offer structural mechanisms for prion strains. In packing polymorphism, prion strains are encoded by alternative packings (polymorphs) of β-sheets formed by the same segment of a protein; in a second mechanism, segmental polymorphism, prion strains are encoded by distinct β-sheets built from different segments of a protein. Both forms of polymorphism can produce enduring “conformations,” capable of encoding strains. These molecular mechanisms for transfer of information into prion strains share features with the familiar mechanism for transfer of information by nucleic acid inheritance, including sequence specificity and recognition by non-covalent bonds. PMID:19684598

  6. Molecular, Structural and Immunological Characterization of Der p 18, a Chitinase-Like House Dust Mite Allergen.

    Directory of Open Access Journals (Sweden)

    Yvonne Resch

    Full Text Available The house dust mite (HDM allergen Der p 18 belongs to the glycoside hydrolase family 18 chitinases. The relevance of Der p 18 for house dust mite allergic patients has only been partly investigated.To perform a detailed characterization of Der p 18 on a molecular, structural and immunological level.Der p 18 was expressed in E. coli, purified to homogeneity, tested for chitin-binding activity and its secondary structure was analyzed by circular dichroism. Der p 18-specific IgG antibodies were produced in rabbits to localize the allergen in mites using immunogold electron microscopy and to search for cross-reactive allergens in other allergen sources (i.e. mites, crustacea, mollusca and insects. IgE reactivity of rDer p 18 was tested with sera from clinically well characterized HDM-allergic patients (n = 98 and its allergenic activity was analyzed in basophil activation experiments.Recombinant Der p 18 was expressed and purified as a folded, biologically active protein. It shows weak chitin-binding activity and partial cross-reactivity with Der f 18 from D. farinae but not with proteins from the other tested allergen sources. The allergen was mainly localized in the peritrophic matrix of the HDM gut and to a lower extent in fecal pellets. Der p 18 reacted with IgE from 10% of mite allergic patients from Austria and showed allergenic activity when tested for basophil activation in Der p 18-sensitized patients.Der p 18 is a rather genus-specific minor allergen with weak chitin-binding activity but exhibits allergenic activity and therefore should be included in diagnostic test panels for HDM allergy.

  7. Development of embryo-like structures in the suspension cultures of flax coincides with secretion of chitinase-like proteins

    Czech Academy of Sciences Publication Activity Database

    Petrovská, Beáta; Salaj, T.; Moravčíková, J.; Libantová, J.; Salaj, J.

    2010-01-01

    Roč. 32, č. 4 (2010), s. 651-656 ISSN 0137-5881 Institutional research plan: CEZ:AV0Z50380511 Keywords : Conditioned media * Embryo-like structures * Linum usitatissimum L Subject RIV: EF - Botanics Impact factor: 1.344, year: 2010

  8. Molecular, Structural and Immunological Characterization of Der p 18, a Chitinase-Like House Dust Mite Allergen

    Czech Academy of Sciences Publication Activity Database

    Resch, A.; Blatt, K.; Malkus, U.; Fercher, CH.; Swoboda, I.; Focke-Tejkl, M.; Chen, K. W.; Seiberler, S.; Mittermann, I.; Lupinek, CH.; Rodriguez-Dominguez, A.; Zieglmayer, P.; Zieglmayer, R.; Keller, W.; Krzyžánek, Vladislav; Valent, P.; Valenta, R.; Vrtala, S.

    2016-01-01

    Roč. 11, č. 8 (2016), e0160641:1-19 E-ISSN 1932-6203 Institutional support: RVO:68081731 Keywords : dermatophagoides-pteronyssinus * peritrophic matrix * igebinding * protein Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering Impact factor: 2.806, year: 2016 http://journals.plos.org/plosone/article/asset?id=10.1371/journal.pone.0160641.PDF

  9. Properties of virion transactivator proteins encoded by primate cytomegaloviruses

    Directory of Open Access Journals (Sweden)

    Barry Peter A

    2009-05-01

    Full Text Available Abstract Background Human cytomegalovirus (HCMV is a betaherpesvirus that causes severe disease in situations where the immune system is immature or compromised. HCMV immediate early (IE gene expression is stimulated by the virion phosphoprotein pp71, encoded by open reading frame (ORF UL82, and this transactivation activity is important for the efficient initiation of viral replication. It is currently recognized that pp71 acts to overcome cellular intrinsic defences that otherwise block viral IE gene expression, and that interactions of pp71 with the cell proteins Daxx and ATRX are important for this function. A further property of pp71 is the ability to enable prolonged gene expression from quiescent herpes simplex virus type 1 (HSV-1 genomes. Non-human primate cytomegaloviruses encode homologs of pp71, but there is currently no published information that addresses their effects on gene expression and modes of action. Results The UL82 homolog encoded by simian cytomegalovirus (SCMV, strain Colburn, was identified and cloned. This ORF, named S82, was cloned into an HSV-1 vector, as were those from baboon, rhesus monkey and chimpanzee cytomegaloviruses. The use of an HSV-1 vector enabled expression of the UL82 homologs in a range of cell types, and permitted investigation of their abilities to direct prolonged gene expression from quiescent genomes. The results show that all UL82 homologs activate gene expression, and that neither host cell type nor promoter target sequence has major effects on these activities. Surprisingly, the UL82 proteins specified by non-human primate cytomegaloviruses, unlike pp71, did not direct long term expression from quiescent HSV-1 genomes. In addition, significant differences were observed in the intranuclear localization of the UL82 homologs, and in their effects on Daxx. Strikingly, S82 mediated the release of Daxx from nuclear domain 10 substructures much more rapidly than pp71 or the other proteins tested. All UL82 homologs stimulated the early release of ATRX from nuclear domain 10. Conclusion All of the UL82 homolog proteins analysed activated gene expression, but surprising differences in other aspects of their properties were revealed. The results provide new information on early events in infection with cytomegaloviruses.

  10. Molecular adaptation within the coat protein-encoding gene of ...

    Indian Academy of Sciences (India)

    Since failing to account for recombination can elevate the false positive error rate in positive ... †These authors contributed equally to this work. ucts, as .... The plant samples (leaves) were ground (1:5, w/v) in a PBS- ..... Sweet cherry. USA.

  11. Identification of physicochemical selective pressure on protein encoding nucleotide sequences

    Directory of Open Access Journals (Sweden)

    Sainudiin Raazesh

    2006-03-01

    Full Text Available Abstract Background Statistical methods for identifying positively selected sites in protein coding regions are one of the most commonly used tools in evolutionary bioinformatics. However, they have been limited by not taking the physiochemical properties of amino acids into account. Results We develop a new codon-based likelihood model for detecting site-specific selection pressures acting on specific physicochemical properties. Nonsynonymous substitutions are divided into substitutions that differ with respect to the physicochemical properties of interest, and those that do not. The substitution rates of these two types of changes, relative to the synonymous substitution rate, are then described by two parameters, γ and ω respectively. The new model allows us to perform likelihood ratio tests for positive selection acting on specific physicochemical properties of interest. The new method is first used to analyze simulated data and is shown to have good power and accuracy in detecting physicochemical selective pressure. We then re-analyze data from the class-I alleles of the human Major Histocompatibility Complex (MHC and from the abalone sperm lysine. Conclusion Our new method allows a more flexible framework to identify selection pressure on particular physicochemical properties.

  12. Chimeras taking shape: Potential functions of proteins encoded by chimeric RNA transcripts

    Science.gov (United States)

    Frenkel-Morgenstern, Milana; Lacroix, Vincent; Ezkurdia, Iakes; Levin, Yishai; Gabashvili, Alexandra; Prilusky, Jaime; del Pozo, Angela; Tress, Michael; Johnson, Rory; Guigo, Roderic; Valencia, Alfonso

    2012-01-01

    Chimeric RNAs comprise exons from two or more different genes and have the potential to encode novel proteins that alter cellular phenotypes. To date, numerous putative chimeric transcripts have been identified among the ESTs isolated from several organisms and using high throughput RNA sequencing. The few corresponding protein products that have been characterized mostly result from chromosomal translocations and are associated with cancer. Here, we systematically establish that some of the putative chimeric transcripts are genuinely expressed in human cells. Using high throughput RNA sequencing, mass spectrometry experimental data, and functional annotation, we studied 7424 putative human chimeric RNAs. We confirmed the expression of 175 chimeric RNAs in 16 human tissues, with an abundance varying from 0.06 to 17 RPKM (Reads Per Kilobase per Million mapped reads). We show that these chimeric RNAs are significantly more tissue-specific than non-chimeric transcripts. Moreover, we present evidence that chimeras tend to incorporate highly expressed genes. Despite the low expression level of most chimeric RNAs, we show that 12 novel chimeras are translated into proteins detectable in multiple shotgun mass spectrometry experiments. Furthermore, we confirm the expression of three novel chimeric proteins using targeted mass spectrometry. Finally, based on our functional annotation of exon organization and preserved domains, we discuss the potential features of chimeric proteins with illustrative examples and suggest that chimeras significantly exploit signal peptides and transmembrane domains, which can alter the cellular localization of cognate proteins. Taken together, these findings establish that some chimeric RNAs are translated into potentially functional proteins in humans. PMID:22588898

  13. Diverse circular replication-associated protein encoding viruses circulating in invertebrates within a lake ecosystem.

    Science.gov (United States)

    Dayaram, Anisha; Galatowitsch, Mark L; Argüello-Astorga, Gerardo R; van Bysterveldt, Katherine; Kraberger, Simona; Stainton, Daisy; Harding, Jon S; Roumagnac, Philippe; Martin, Darren P; Lefeuvre, Pierre; Varsani, Arvind

    2016-04-01

    Over the last five years next-generation sequencing has become a cost effective and efficient method for identifying known and unknown microorganisms. Access to this technique has dramatically changed the field of virology, enabling a wide range of environmental viral metagenome studies to be undertaken of organisms and environmental samples from polar to tropical regions. These studies have led to the discovery of hundreds of highly divergent single stranded DNA (ssDNA) virus-like sequences encoding replication-associated proteins. Yet, few studies have explored how viruses might be shared in an ecosystem through feeding relationships. Here we identify 169 circular molecules (160 CRESS DNA molecules, nine circular molecules) recovered from a New Zealand freshwater lake, that we have tentatively classified into 51 putatively novel species and five previously described species (DflaCV-3, -5, -6, -8, -10). The CRESS DNA viruses identified in this study were recovered from molluscs (Echyridella menzeisii, Musculium novaezelandiae, Potamopyrgus antipodarum and Physella acuta) and insect larvae (Procordulia grayi, Xanthocnemis zealandica, and Chironomus zealandicus) collected from Lake Sarah, as well as from the lake water and benthic sediments. Extensive diversity was observed across most CRESS DNA molecules recovered. The putative capsid protein of one viral species was found to be most similar to those of members of the Tombusviridae family, thus expanding the number of known RNA-DNA hybrid viruses in nature. We noted a strong association between the CRESS DNA viruses and circular molecules identified in the water and browser organisms (C. zealandicus, P. antipodarum and P. acuta), and between water sediments and undefended prey species (C. zealandicus). However, we were unable to find any significant correlation of viral assemblages to the potential feeding relationships of the host aquatic invertebrates. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. Comparative genomic analyses of transport proteins encoded within the genomes of Leptospira species.

    Science.gov (United States)

    Buyuktimkin, Bora; Saier, Milton H

    2015-11-01

    Select species of the bacterial genus Leptospira are causative agents of leptospirosis, an emerging global zoonosis affecting nearly one million people worldwide annually. We examined two Leptospira pathogens, Leptospira interrogans serovar Lai str. 56601 and Leptospira borgpetersenii serovar Hardjo-bovis str. L550, as well as the free-living leptospiral saprophyte, Leptospira biflexa serovar Patoc str. 'Patoc 1 (Ames)'. The transport proteins of these leptospires were identified and compared using bioinformatics to gain an appreciation for which proteins may be related to pathogenesis and saprophytism. L. biflexa possesses a disproportionately high number of secondary carriers for metabolite uptake and environmental adaptability as well as an increased number of inorganic cation transporters providing ionic homeostasis and effective osmoregulation in a rapidly changing environment. L. interrogans and L. borgpetersenii possess far fewer transporters, but those that they have are remarkably similar, with near-equivalent representation in most transporter families. These two Leptospira pathogens also possess intact sphingomyelinases, holins, and virulence-related outer membrane porins. These virulence-related factors, in conjunction with decreased transporter substrate versatility, indicate that pathogenicity was accompanied by progressively narrowing ecological niches and the emergence of a limited set of proteins responsible for host invasion. The variability of host tropism and mortality rates by infectious leptospires suggests that small differences in individual sets of proteins play important physiological and pathological roles. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Comparative analyses of transport proteins encoded within the genomes of Leptospira species.

    Science.gov (United States)

    Buyuktimkin, Bora; Saier, Milton H

    2016-09-01

    Select species of the bacterial genus Leptospira are causative agents of leptospirosis, an emerging global zoonosis affecting nearly one million people worldwide annually. We examined two Leptospira pathogens, Leptospira interrogans serovar Lai str. 56601 and Leptospira borgpetersenii serovar Hardjo-bovis str. L550, as well as the free-living leptospiral saprophyte, Leptospira biflexa serovar Patoc str. 'Patoc 1 (Ames)'. The transport proteins of these leptospires were identified and compared using bioinformatics to gain an appreciation for which proteins may be related to pathogenesis and saprophytism. L. biflexa possesses a disproportionately high number of secondary carriers for metabolite uptake and environmental adaptability as well as an increased number of inorganic cation transporters providing ionic homeostasis and effective osmoregulation in a rapidly changing environment. L. interrogans and L. borgpetersenii possess far fewer transporters, but those that they all have are remarkably similar, with near-equivalent representation in most transporter families. These two Leptospira pathogens also possess intact sphingomyelinases, holins, and virulence-related outer membrane porins. These virulence-related factors, in conjunction with decreased transporter substrate versatility, indicate that pathogenicity arose in Leptospira correlating to progressively narrowing ecological niches and the emergence of a limited set of proteins responsible for host invasion. The variability of host tropism and mortality rates by infectious leptospires suggests that small differences in individual sets of proteins play important physiological and pathological roles. Copyright © 2016. Published by Elsevier Ltd.

  16. Brain transcriptional stability upon prion protein-encoding gene invalidation in zygotic or adult mouse

    Directory of Open Access Journals (Sweden)

    Béringue Vincent

    2010-07-01

    Full Text Available Abstract Background The physiological function of the prion protein remains largely elusive while its key role in prion infection has been expansively documented. To potentially assess this conundrum, we performed a comparative transcriptomic analysis of the brain of wild-type mice with that of transgenic mice invalidated at this locus either at the zygotic or at the adult stages. Results Only subtle transcriptomic differences resulting from the Prnp knockout could be evidenced, beside Prnp itself, in the analyzed adult brains following microarray analysis of 24 109 mouse genes and QPCR assessment of some of the putatively marginally modulated loci. When performed at the adult stage, neuronal Prnp disruption appeared to sequentially induce a response to an oxidative stress and a remodeling of the nervous system. However, these events involved only a limited number of genes, expression levels of which were only slightly modified and not always confirmed by RT-qPCR. If not, the qPCR obtained data suggested even less pronounced differences. Conclusions These results suggest that the physiological function of PrP is redundant at the adult stage or important for only a small subset of the brain cell population under classical breeding conditions. Following its early reported embryonic developmental regulation, this lack of response could also imply that PrP has a more detrimental role during mouse embryogenesis and that potential transient compensatory mechanisms have to be searched for at the time this locus becomes transcriptionally activated.

  17. Translational Control of Host Gene Expression by a Cys-Motif Protein Encoded in a Bracovirus.

    Directory of Open Access Journals (Sweden)

    Eunseong Kim

    Full Text Available Translational control is a strategy that various viruses use to manipulate their hosts to suppress acute antiviral response. Polydnaviruses, a group of insect double-stranded DNA viruses symbiotic to some endoparasitoid wasps, are divided into two genera: ichnovirus (IV and bracovirus (BV. In IV, some Cys-motif genes are known as host translation-inhibitory factors (HTIF. The genome of endoparasitoid wasp Cotesia plutellae contains a Cys-motif gene (Cp-TSP13 homologous to an HTIF known as teratocyte-secretory protein 14 (TSP14 of Microplitis croceipes. Cp-TSP13 consists of 129 amino acid residues with a predicted molecular weight of 13.987 kDa and pI value of 7.928. Genomic DNA region encoding its open reading frame has three introns. Cp-TSP13 possesses six conserved cysteine residues as other Cys-motif genes functioning as HTIF. Cp-TSP13 was expressed in Plutella xylostella larvae parasitized by C. plutellae. C. plutellae bracovirus (CpBV was purified and injected into non-parasitized P. xylostella that expressed Cp-TSP13. Cp-TSP13 was cloned into a eukaryotic expression vector and used to infect Sf9 cells to transiently express Cp-TSP13. The synthesized Cp-TSP13 protein was detected in culture broth. An overlaying experiment showed that the purified Cp-TSP13 entered hemocytes. It was localized in the cytosol. Recombinant Cp-TSP13 significantly inhibited protein synthesis of secretory proteins when it was added to in vitro cultured fat body. In addition, the recombinant Cp-TSP13 directly inhibited the translation of fat body mRNAs in in vitro translation assay using rabbit reticulocyte lysate. Moreover, the recombinant Cp-TSP13 significantly suppressed cellular immune responses by inhibiting hemocyte-spreading behavior. It also exhibited significant insecticidal activities by both injection and feeding routes. These results indicate that Cp-TSP13 is a viral HTIF.

  18. Evaluating the silencing suppressor activity of proteins encoded by maize rayado fino virus

    Science.gov (United States)

    Maize rayado fino virus (MRFV), the type member of the genus Marafivirus, family Tymoviridae, is transmitted in a persistent, circulative manner by leafhoppers of the genus Dalbulus. Symptoms of MRFV infection on leaves of its maize host are small chlorotic spots that coalesce into short stripes. T...

  19. Molecular comparison of the structural proteins encoding gene clusters of two related Lactobacillus delbrueckii bacteriophages.

    Science.gov (United States)

    Vasala, A; Dupont, L; Baumann, M; Ritzenthaler, P; Alatossava, T

    1993-01-01

    Virulent phage LL-H and temperate phage mv4 are two related bacteriophages of Lactobacillus delbrueckii. The gene clusters encoding structural proteins of these two phages have been sequenced and further analyzed. Six open reading frames (ORF-1 to ORF-6) were detected. Protein sequencing and Western immunoblotting experiments confirmed that ORF-3 (g34) encoded the main capsid protein Gp34. The presence of a putative late promoter in front of the phage LL-H g34 gene was suggested by primer extension experiments. Comparative sequence analysis between phage LL-H and phage mv4 revealed striking similarities in the structure and organization of this gene cluster, suggesting that the genes encoding phage structural proteins belong to a highly conservative module. Images PMID:8497043

  20. Proteins Encoded in Genomic Regions Associated with Immune-Mediated Disease Physically Interact and Suggest Underlying Biology

    Science.gov (United States)

    Rossin, Elizabeth J.; Lage, Kasper; Raychaudhuri, Soumya; Xavier, Ramnik J.; Tatar, Diana; Benita, Yair

    2011-01-01

    Genome-wide association studies (GWAS) have defined over 150 genomic regions unequivocally containing variation predisposing to immune-mediated disease. Inferring disease biology from these observations, however, hinges on our ability to discover the molecular processes being perturbed by these risk variants. It has previously been observed that different genes harboring causal mutations for the same Mendelian disease often physically interact. We sought to evaluate the degree to which this is true of genes within strongly associated loci in complex disease. Using sets of loci defined in rheumatoid arthritis (RA) and Crohn's disease (CD) GWAS, we build protein–protein interaction (PPI) networks for genes within associated loci and find abundant physical interactions between protein products of associated genes. We apply multiple permutation approaches to show that these networks are more densely connected than chance expectation. To confirm biological relevance, we show that the components of the networks tend to be expressed in similar tissues relevant to the phenotypes in question, suggesting the network indicates common underlying processes perturbed by risk loci. Furthermore, we show that the RA and CD networks have predictive power by demonstrating that proteins in these networks, not encoded in the confirmed list of disease associated loci, are significantly enriched for association to the phenotypes in question in extended GWAS analysis. Finally, we test our method in 3 non-immune traits to assess its applicability to complex traits in general. We find that genes in loci associated to height and lipid levels assemble into significantly connected networks but did not detect excess connectivity among Type 2 Diabetes (T2D) loci beyond chance. Taken together, our results constitute evidence that, for many of the complex diseases studied here, common genetic associations implicate regions encoding proteins that physically interact in a preferential manner, in line with observations in Mendelian disease. PMID:21249183

  1. A Legionella pneumophila effector protein encoded in a region of genomic plasticity binds to Dot/Icm-modified vacuoles.

    Directory of Open Access Journals (Sweden)

    Shira Ninio

    2009-01-01

    Full Text Available Legionella pneumophila is an opportunistic pathogen that can cause a severe pneumonia called Legionnaires' disease. In the environment, L. pneumophila is found in fresh water reservoirs in a large spectrum of environmental conditions, where the bacteria are able to replicate within a variety of protozoan hosts. To survive within eukaryotic cells, L. pneumophila require a type IV secretion system, designated Dot/Icm, that delivers bacterial effector proteins into the host cell cytoplasm. In recent years, a number of Dot/Icm substrate proteins have been identified; however, the function of most of these proteins remains unknown, and it is unclear why the bacterium maintains such a large repertoire of effectors to promote its survival. Here we investigate a region of the L. pneumophila chromosome that displays a high degree of plasticity among four sequenced L. pneumophila strains. Analysis of GC content suggests that several genes encoded in this region were acquired through horizontal gene transfer. Protein translocation studies establish that this region of genomic plasticity encodes for multiple Dot/Icm effectors. Ectopic expression studies in mammalian cells indicate that one of these substrates, a protein called PieA, has unique effector activities. PieA is an effector that can alter lysosome morphology and associates specifically with vacuoles that support L. pneumophila replication. It was determined that the association of PieA with vacuoles containing L. pneumophila requires modifications to the vacuole mediated by other Dot/Icm effectors. Thus, the localization properties of PieA reveal that the Dot/Icm system has the ability to spatially and temporally control the association of an effector with vacuoles containing L. pneumophila through activities mediated by other effector proteins.

  2. The mechanosensory structure of the hair cell requires clarin-1, a protein encoded by Usher syndrome III causative gene.

    Science.gov (United States)

    Geng, Ruishuang; Melki, Sami; Chen, Daniel H-C; Tian, Guilian; Furness, David N; Oshima-Takago, Tomoko; Neef, Jakob; Moser, Tobias; Askew, Charles; Horwitz, Geoff; Holt, Jeffrey R; Imanishi, Yoshikazu; Alagramam, Kumar N

    2012-07-11

    Mutation in the clarin-1 gene (Clrn1) results in loss of hearing and vision in humans (Usher syndrome III), but the role of clarin-1 in the sensory hair cells is unknown. Clarin-1 is predicted to be a four transmembrane domain protein similar to members of the tetraspanin family. Mice carrying null mutation in the clarin-1 gene (Clrn1(-/-)) show loss of hair cell function and a possible defect in ribbon synapse. We investigated the role of clarin-1 using various in vitro and in vivo approaches. We show by immunohistochemistry and patch-clamp recordings of Ca(2+) currents and membrane capacitance from inner hair cells that clarin-1 is not essential for formation or function of ribbon synapse. However, reduced cochlear microphonic potentials, FM1-43 [N-(3-triethylammoniumpropyl)-4-(4-(dibutylamino)styryl) pyridinium dibromide] loading, and transduction currents pointed to diminished cochlear hair bundle function in Clrn1(-/-) mice. Electron microscopy of cochlear hair cells revealed loss of some tall stereocilia and gaps in the v-shaped bundle, although tip links and staircase arrangement of stereocilia were not primarily affected by Clrn1(-/-) mutation. Human clarin-1 protein expressed in transfected mouse cochlear hair cells localized to the bundle; however, the pathogenic variant p.N48K failed to localize to the bundle. The mouse model generated to study the in vivo consequence of p.N48K in clarin-1 (Clrn1(N48K)) supports our in vitro and Clrn1(-/-) mouse data and the conclusion that CLRN1 is an essential hair bundle protein. Furthermore, the ear phenotype in the Clrn1(N48K) mouse suggests that it is a valuable model for ear disease in CLRN1(N48K), the most prevalent Usher syndrome III mutation in North America.

  3. The Mechanosensory Structure of the Hair Cell Requires Clarin-1, a Protein Encoded by Usher Syndrome III Causative Gene

    Science.gov (United States)

    Geng, Ruishuang; Melki, Sami; Chen, Daniel H.-C.; Tian, Guilian; Furness, David; Oshima-Takago, Tomoko; Neef, Jakob; Moser, Tobias; Askew, Charles; Horwitz, Geoff; Holt, Jeffrey; Imanishi, Yoshikazu; Alagramam, Kumar N.

    2012-01-01

    Mutation in the clarin-1 gene results in loss of hearing and vision in humans (Usher syndrome III), but the role of clarin-1 in the sensory hair cells is unknown. Clarin-1 is predicted to be a four transmembrane domain protein similar to members of the tetraspanin family. Mice carrying null mutation in the clarin-1 (Clrn1−/−) gene show loss of hair cell function and a possible defect in ribbon synapse. We investigated the role of clarin-1 using various in vitro and in vivo approaches. We show by immunohistochemistry and patch-clamp recordings of Ca2+ currents and membrane capacitance from IHCs that clarin-1 is not essential for formation or function of ribbon synapse. However, reduced cochlear microphonic potentials, FM1-43 loading and transduction currents pointed to diminished cochlear hair bundle function in Clrn1−/− mice. Electron microscopy of cochlear hair cells revealed loss of some tall stereocilia and gaps in the v-shaped bundle, although tip-links and staircase arrangement of stereocilia were not primarily affected by Clrn1−/− mutation. Human clarin-1 protein expressed in transfected mouse cochlear hair cells localized to the bundle; however, the pathogenic variant, p.N48K, failed to localize to the bundle. The mouse model generated to study the in vivo consequence of p. N48K in clarin-1 (Clrn1N48K) supports our in vitro and Clrn1−/− mouse data and the conclusion that CLRN1 is an essential hair bundle protein. Further, the ear phenotype in the Clrn1N48K mouse suggests that it is a valuable model for ear disease in CLRN1N48K, the most prevalent Usher III mutation in North America. PMID:22787034

  4. Proteins Encoded in Genomic Regions Associated with Immune-Mediated Disease Physically Interact and Suggest Underlying Biology

    DEFF Research Database (Denmark)

    Rossin, Elizabeth J.; Hansen, Kasper Lage; Raychaudhuri, Soumya

    2011-01-01

    Genome-wide association studies (GWAS) have defined over 150 genomic regions unequivocally containing variation predisposing to immune-mediated disease. Inferring disease biology from these observations, however, hinges on our ability to discover the molecular processes being perturbed by these r......Genome-wide association studies (GWAS) have defined over 150 genomic regions unequivocally containing variation predisposing to immune-mediated disease. Inferring disease biology from these observations, however, hinges on our ability to discover the molecular processes being perturbed...... in rheumatoid arthritis (RA) and Crohn's disease (CD) GWAS, we build protein-protein interaction (PPI) networks for genes within associated loci and find abundant physical interactions between protein products of associated genes. We apply multiple permutation approaches to show that these networks are more...... that the RA and CD networks have predictive power by demonstrating that proteins in these networks, not encoded in the confirmed list of disease associated loci, are significantly enriched for association to the phenotypes in question in extended GWAS analysis. Finally, we test our method in 3 non...

  5. Phylogenetic analysis of fungal heterotrimeric G protein-encoding genes and their expression during dimorphism in Mucor circinelloides.

    Science.gov (United States)

    Valle-Maldonado, Marco Iván; Jácome-Galarza, Irvin Eduardo; Díaz-Pérez, Alma Laura; Martínez-Cadena, Guadalupe; Campos-García, Jesús; Ramírez-Díaz, Martha Isela; Reyes-De la Cruz, Homero; Riveros-Rosas, Héctor; Díaz-Pérez, César; Meza-Carmen, Víctor

    2015-12-01

    In fungi, heterotrimeric G proteins are key regulators of biological processes such as mating, virulence, morphology, among others. Mucor circinelloides is a model organism for many biological processes, and its genome contains the largest known repertoire of genes that encode putative heterotrimeric G protein subunits in the fungal kingdom: twelve Gα (McGpa1-12), three Gβ (McGpb1-3), and three Gγ (McGpg1-3). Phylogenetic analysis of fungal Gα showed that they are divided into four distinct groups as reported previously. Fungal Gβ and Gγ are also divided into four phylogenetic groups, and to our understanding this is the first report of a phylogenetic classification for fungal Gβ and Gγ subunits. Almost all genes that encode putative heterotrimeric G subunits in M. circinelloides are differentially expressed during dimorphic growth, except for McGpg1 (Gγ) that showed very low mRNA levels at all developmental stages. Moreover, several of the subunits are expressed in a similar pattern and at the same level, suggesting that they constitute discrete complexes. For example, McGpb3 (Gβ), and McGpg2 (Gγ), are co-expressed during mycelium growth, and McGpa1, McGpb2, and McGpg2, are co-expressed during yeast development. These findings provide the conceptual framework to study the biological role of these genes during M. circinelloides morphogenesis. Copyright © 2015 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  6. Optimized Mitochondrial Targeting of Proteins Encoded by Modified mRNAs Rescues Cells Harboring Mutations in mtATP6

    Directory of Open Access Journals (Sweden)

    Randall Marcelo Chin

    2018-03-01

    Full Text Available Summary: Mitochondrial disease may be caused by mutations in the protein-coding genes of the mitochondrial genome. A promising strategy for treating such diseases is allotopic expression—the translation of wild-type copies of these proteins in the cytosol, with subsequent translocation into the mitochondria, resulting in rescue of mitochondrial function. In this paper, we develop an automated, quantitative, and unbiased screening platform to evaluate protein localization and mitochondrial morphology. This platform was used to compare 31 mitochondrial targeting sequences and 15 3′ UTRs in their ability to localize up to 9 allotopically expressed proteins to the mitochondria and their subsequent impact on mitochondrial morphology. Taking these two factors together, we synthesized chemically modified mRNAs that encode for an optimized allotopic expression construct for mtATP6. These mRNAs were able to functionally rescue a cell line harboring the 8993T > G point mutation in the mtATP6 gene. : Allotopic expression of proteins normally encoded by mtDNA is a promising therapy for mitochondrial disease. Chin et al. use an unbiased and high-content imaging-based screening platform to optimize allotopic expression. Modified mRNAs encoding for the optimized allotopic expression constructs rescued the respiration and growth of mtATP6-deficient cells. Keywords: mitochondria, mitochondrial disease, mRNA, modified mRNA, ATP6, allotopic expression, rare disease, gene therapy, screening, high content imaging

  7. Thermal response of rat fibroblasts stably transfected with the human 70-kDa heat shock protein-encoding gene

    International Nuclear Information System (INIS)

    Li, G.C.; Li, Ligeng; Liu, Yunkang; Mak, J.Y.; Chen, Lili; Lee, W.M.F.

    1991-01-01

    The major heat shock protein hsp70 is synthesized by cells of a wide variety of organisms in response to heat shock or other environmental stresses and is assumed to play an important role in protecting cells from thermal stress. The authors have tested this hypothesis directly by transfecting a constitutively expressed recombinant human hsp70-encoding gene into rat fibroblasts and examining the relationship between the levels of human hsp70 expressed and thermal resistance of the stably transfected rat cells. Successful transfection and expression of the gene for human hsp70 were characterized by RNA hybridization analysis, low-dimensional gel electrophoresis, and immunoblot analysis. When individual cloned cell lines were exposed to 45C and their thermal survivals were determined by colony-formation assay, they found that the expression of human hsp70 conferred heat resistance to the rat cells. These results reinforce the hypothesis that hsp70 has a protective function against thermal stress

  8. Heterogeneous nuclear ribonucleoproteins H, H', and F are members of a ubiquitously expressed subfamily of related but distinct proteins encoded by genes mapping to different chromosomes

    DEFF Research Database (Denmark)

    Honoré, B; Rasmussen, H H; Vorum, H

    1995-01-01

    Molecular cDNA cloning, two-dimensional gel immunoblotting, and amino acid microsequencing identified three sequence-unique and distinct proteins that constitute a subfamily of ubiquitously expressed heterogeneous nuclear ribonucleoproteins corresponding to hnRNPs H, H', and F. These proteins share...... epitopes and sequence identity with two other proteins, isoelectric focusing sample spot numbers 2222 (37.6 kDa; pI 6.5) and 2326 (39.5 kDa; pI 6.6), indicating that the subfamily may contain additional members. The identity between hnRNPs H and H' is 96%, between H and F 78%, and between H' and F 75......%, respectively. The three proteins contain three repeats, which we denote quasi-RRMs (qRRMs) since they have a remote similarity to the RNA recognition motif (RRM). The three qRRMs of hnRNP H, with a few additional NH2-terminal amino acids, were constructed by polymerase chain reaction amplification and used...

  9. Safety evaluation of the phosphinothricin acetyltransferase proteins encoded by the pat and bar sequences that confer tolerance to glufosinate-ammonium herbicide in transgenic plants.

    Science.gov (United States)

    Hérouet, Corinne; Esdaile, David J; Mallyon, Bryan A; Debruyne, Eric; Schulz, Arno; Currier, Thomas; Hendrickx, Koen; van der Klis, Robert-Jan; Rouan, Dominique

    2005-03-01

    Transgenic plant varieties, which are tolerant to glufosinate-ammonium, were developed. The herbicide tolerance is based upon the presence of either the bar or the pat gene, which encode for two homologous phosphinothricin acetyltransferases (PAT), in the plant genome. Based on both a review of published literature and experimental studies, the safety assessment reviews the first step of a two-step-approach for the evaluation of the safety of the proteins expressed in plants. It can be used to support the safety of food or feed products derived from any crop that contains and expresses these PAT proteins. The safety evaluation supports the conclusion that the genes and the donor microorganisms (Streptomyces) are innocuous. The PAT enzymes are highly specific and do not possess the characteristics associated with food toxins or allergens, i.e., they have no sequence homology with any known allergens or toxins, they have no N-glycosylation sites, they are rapidly degraded in gastric and intestinal fluids, and they are devoid of adverse effects in mice after intravenous administration at a high dose level. In conclusion, there is a reasonable certainty of no harm resulting from the inclusion of the PAT proteins in human food or in animal feed.

  10. Combined virus-like particle and fusion protein-encoding DNA vaccination of cotton rats induces protection against respiratory syncytial virus without causing vaccine-enhanced disease

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Hye Suk; Lee, Young-Tae; Kim, Ki-Hye; Park, Soojin; Kwon, Young-Man; Lee, Youri; Ko, Eun-Ju; Jung, Yu-Jin [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); Lee, Jong Seok [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); National Institute of Biological Resources, Incheon (Korea, Republic of); Kim, Yu-Jin [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); Lee, Yu-Na; Kim, Min-Chul [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); Animal and Plant Quarantine Agency, Gyeonggi-do, Gimcheon, Gyeongsangbukdo (Korea, Republic of); Cho, Minkyoung [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); Kang, Sang-Moo, E-mail: skang24@gsu.edu [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States)

    2016-07-15

    A safe and effective vaccine against respiratory syncytial virus (RSV) should confer protection without causing vaccine-enhanced disease. Here, using a cotton rat model, we investigated the protective efficacy and safety of an RSV combination vaccine composed of F-encoding plasmid DNA and virus-like particles containing RSV fusion (F) and attachment (G) glycoproteins (FFG-VLP). Cotton rats with FFG-VLP vaccination controlled lung viral replication below the detection limit, and effectively induced neutralizing activity and antibody-secreting cell responses. In comparison with formalin inactivated RSV (FI-RSV) causing severe RSV disease after challenge, FFG-VLP vaccination did not cause weight loss, airway hyper-responsiveness, IL-4 cytokines, histopathology, and infiltrates of proinflammatory cells such as eosinophils. FFG-VLP was even more effective in preventing RSV-induced pulmonary inflammation than live RSV infections. This study provides evidence that FFG-VLP can be developed into a safe and effective RSV vaccine candidate. - Highlights: • Combined RSV FFG VLP vaccine is effective in inducing F specific responses. • FFG VLP vaccine confers RSV neutralizing activity and viral control in cotton rats. • Cotton rats with RSV FFG VLP vaccination do not show vaccine-enhanced disease. • Cotton rats with FFG VLP vaccine induce F specific antibody secreting cell responses. • Cotton rats with FFG VLP do not induce lung cellular infiltrates and Th2 cytokine.

  11. The Nance-Horan syndrome protein encodes a functional WAVE homology domain (WHD) and is important for co-ordinating actin remodelling and maintaining cell morphology.

    Science.gov (United States)

    Brooks, Simon P; Coccia, Margherita; Tang, Hao R; Kanuga, Naheed; Machesky, Laura M; Bailly, Maryse; Cheetham, Michael E; Hardcastle, Alison J

    2010-06-15

    Nance-Horan syndrome (NHS) is an X-linked developmental disorder, characterized by bilateral congenital cataracts, dental anomalies, facial dysmorphism and mental retardation. Null mutations in a novel gene, NHS, cause the syndrome. The NHS gene appears to have multiple isoforms as a result of alternative transcription, but a cellular function for the NHS protein has yet to be defined. We describe NHS as a founder member of a new protein family (NHS, NHSL1 and NHSL2). Here, we demonstrate that NHS is a novel regulator of actin remodelling and cell morphology. NHS localizes to sites of cell-cell contact, the leading edge of lamellipodia and focal adhesions. The N-terminus of isoforms NHS-A and NHS-1A, implicated in the pathogenesis of NHS, have a functional WAVE homology domain that interacts with the Abi protein family, haematopoietic stem/progenitor cell protein 300 (HSPC300), Nap1 and Sra1. NHS knockdown resulted in the disruption of the actin cytoskeleton. We show that NHS controls cell morphology by maintaining the integrity of the circumferential actin ring and controlling lamellipod formation. NHS knockdown led to a striking increase in cell spreading. Conversely, ectopic overexpression of NHS inhibited lamellipod formation. Remodelling of the actin cytoskeleton and localized actin polymerization into branched actin filaments at the plasma membrane are essential for mediating changes in cell shape, migration and cell contact. Our data identify NHS as a new regulator of actin remodelling. We suggest that NHS orchestrates actin regulatory protein function in response to signalling events during development.

  12. The Nance–Horan syndrome protein encodes a functional WAVE homology domain (WHD) and is important for co-ordinating actin remodelling and maintaining cell morphology

    Science.gov (United States)

    Brooks, Simon P.; Coccia, Margherita; Tang, Hao R.; Kanuga, Naheed; Machesky, Laura M.; Bailly, Maryse; Cheetham, Michael E.; Hardcastle, Alison J.

    2010-01-01

    Nance–Horan syndrome (NHS) is an X-linked developmental disorder, characterized by bilateral congenital cataracts, dental anomalies, facial dysmorphism and mental retardation. Null mutations in a novel gene, NHS, cause the syndrome. The NHS gene appears to have multiple isoforms as a result of alternative transcription, but a cellular function for the NHS protein has yet to be defined. We describe NHS as a founder member of a new protein family (NHS, NHSL1 and NHSL2). Here, we demonstrate that NHS is a novel regulator of actin remodelling and cell morphology. NHS localizes to sites of cell–cell contact, the leading edge of lamellipodia and focal adhesions. The N-terminus of isoforms NHS-A and NHS-1A, implicated in the pathogenesis of NHS, have a functional WAVE homology domain that interacts with the Abi protein family, haematopoietic stem/progenitor cell protein 300 (HSPC300), Nap1 and Sra1. NHS knockdown resulted in the disruption of the actin cytoskeleton. We show that NHS controls cell morphology by maintaining the integrity of the circumferential actin ring and controlling lamellipod formation. NHS knockdown led to a striking increase in cell spreading. Conversely, ectopic overexpression of NHS inhibited lamellipod formation. Remodelling of the actin cytoskeleton and localized actin polymerization into branched actin filaments at the plasma membrane are essential for mediating changes in cell shape, migration and cell contact. Our data identify NHS as a new regulator of actin remodelling. We suggest that NHS orchestrates actin regulatory protein function in response to signalling events during development. PMID:20332100

  13. Identification of herpes simplex virus type 1 proteins encoded within the first 1.5 kb of the latency-associated transcript.

    Science.gov (United States)

    Henderson, Gail; Jaber, Tareq; Carpenter, Dale; Wechsler, Steven L; Jones, Clinton

    2009-09-01

    Expression of the first 1.5 kb of the latency-associated transcript (LAT) that is encoded by herpes simplex virus type 1 (HSV-1) is sufficient for wild-type (wt) levels of reactivation from latency in small animal models. Peptide-specific immunoglobulin G (IgG) was generated against open reading frames (ORFs) that are located within the first 1.5 kb of LAT coding sequences. Cells stably transfected with LAT or trigeminal ganglionic neurons of mice infected with a LAT expressing virus appeared to express the L2 or L8 ORF. Only L2 ORF expression was readily detected in trigeminal ganglionic neurons of latently infected mice.

  14. RNA-Seq-based analysis of cold shock response in Thermoanaerobacter tengcongensis, a bacterium harboring a single cold shock protein encoding gene.

    Directory of Open Access Journals (Sweden)

    Bo Liu

    Full Text Available BACKGROUND: Although cold shock responses and the roles of cold shock proteins in microorganisms containing multiple cold shock protein genes have been well characterized, related studies on bacteria possessing a single cold shock protein gene have not been reported. Thermoanaerobacter tengcongensis MB4, a thermophile harboring only one known cold shock protein gene (TtescpC, can survive from 50° to 80 °C, but has poor natural competence under cold shock at 50 °C. We therefore examined cold shock responses and their effect on natural competence in this bacterium. RESULTS: The transcriptomes of T. tengcongensis before and after cold shock were analyzed by RNA-seq and over 1200 differentially expressed genes were successfully identified. These genes were involved in a wide range of biological processes, including modulation of DNA replication, recombination, and repair; energy metabolism; production of cold shock protein; synthesis of branched amino acids and branched-chain fatty acids; and sporulation. RNA-seq analysis also suggested that T. tengcongensis initiates cell wall and membrane remodeling processes, flagellar assembly, and sporulation in response to low temperature. Expression profiles of TtecspC and failed attempts to produce a TtecspC knockout strain confirmed the essential role of TteCspC in the cold shock response, and also suggested a role of this protein in survival at optimum growth temperature. Repression of genes encoding ComEA and ComEC and low energy metabolism levels in cold-shocked cells are the likely basis of poor natural competence at low temperature. CONCLUSION: Our study demonstrated changes in global gene expression under cold shock and identified several candidate genes related to cold shock in T. tengcongensis. At the same time, the relationship between cold shock response and poor natural competence at low temperature was preliminarily elucidated. These findings provide a foundation for future studies on genetic and molecular mechanisms associated with cold shock and acclimation at low temperature.

  15. Putative recombination events and evolutionary history of five economically important viruses of fruit trees based on coat protein-encoding gene sequence analysis.

    Science.gov (United States)

    Boulila, Moncef

    2010-06-01

    To enhance the knowledge of recombination as an evolutionary process, 267 accessions retrieved from GenBank were investigated, all belonging to five economically important viruses infecting fruit crops (Plum pox, Apple chlorotic leaf spot, Apple mosaic, Prune dwarf, and Prunus necrotic ringspot viruses). Putative recombinational events were detected in the coat protein (CP)-encoding gene using RECCO and RDP version 3.31beta algorithms. Based on RECCO results, all five viruses were shown to contain potential recombination signals in the CP gene. Reconstructed trees with modified topologies were proposed. Furthermore, RECCO performed better than the RDP package in detecting recombination events and exhibiting their evolution rate along the sequences of the five viruses. RDP, however, provided the possible major and minor parents of the recombinants. Thus, the two methods should be considered complementary.

  16. Cloning of human basic A1, a distinct 59-kDa dystrophin-associated protein encoded on chromosome 8q23-24

    Energy Technology Data Exchange (ETDEWEB)

    Ahn, A.H. [Harvard Medical School, Boston, MA (United States); Yoshida, Mikiharu; Hagiwara, Yasuko; Ozawa, Eijiro [National Institute of Neuroscience, Ogawa Higashi, Kodaira (Japan); Anderson, M.S.; Feener, C.A.; Selig, S. [Howard Hughes Medical Institute at Children`s Hospital, Boston, MA (United States); Kunkel, L.M. [Harvard Medical School, Boston, MA (United States)]|[Howard Hughes Medical Institute at Children`s Hosptial, Boston, MA (United States)

    1994-05-10

    Duchenne and Becker muscular dystrophies are caused by defects of dystrophin, which forms a part of the membrane cytoskeleton of specialized cells such as muscle. It has been previously shown that the dystrophin-associated protein A1 (59-kDa DAP) is actually a heterogeneous group of phosphorylated proteins consisting of an acidic ({alpha}-A1) and a distinct basic ({beta}-A1) component. Partial peptide sequence of the A1 complex purified from rabbit muscle permitted the design of oligonucleotide probes that were used to isolate a cDNA for one human isoform of A1. This cDNA encodes a basic A1 isoform that is distinct from the recently described syntrophins in Torpedo and mouse and is expressed in many tissues with at least five distinct mRNA species of 5.9, 4.8, 4.3, 3.1, and 1.5 kb. A comparison of the human cDNA sequence with the GenBank expressed sequence tag (EST) data base has identified a relative from human skeletal muscle, EST25263, which is probably a human homologue of the published mouse syntrophin 2. The authors have mapped the human basic component of A1 and EST25263 genes to chromosomes 8q23-24 and 16, respectively.

  17. Translational control and differential RNA decay are key elements regulating postsegregational expression of the killer protein encoded by the parB locus of plasmid R1

    DEFF Research Database (Denmark)

    Gerdes, K; Helin, K; Christensen, O W

    1988-01-01

    The parB locus of plasmid R1, which mediates plasmid stability via postsegregational killing of plasmid-free cells, encodes two genes, hok and sok. The hok gene product is a potent cell-killing protein. The hok gene is regulated at the translational level by the sok gene-encoded repressor, a small...

  18. MvaT Family Proteins Encoded on IncP-7 Plasmid pCAR1 and the Host Chromosome Regulate the Host Transcriptome Cooperatively but Differently.

    Science.gov (United States)

    Yun, Choong-Soo; Takahashi, Yurika; Shintani, Masaki; Takeda, Toshiharu; Suzuki-Minakuchi, Chiho; Okada, Kazunori; Yamane, Hisakazu; Nojiri, Hideaki

    2016-02-01

    MvaT proteins are members of the H-NS family of proteins in pseudomonads. The IncP-7 conjugative plasmid pCAR1 carries an mvaT-homologous gene, pmr. In Pseudomonas putida KT2440 bearing pCAR1, pmr and the chromosomally carried homologous genes, turA and turB, are transcribed at high levels, and Pmr interacts with TurA and TurB in vitro. In the present study, we clarified how the three MvaT proteins regulate the transcriptome of P. putida KT2440(pCAR1). Analyses performed by a modified chromatin immunoprecipitation assay with microarray technology (ChIP-chip) suggested that the binding regions of Pmr, TurA, and TurB in the P. putida KT2440(pCAR1) genome are almost identical; nevertheless, transcriptomic analyses using mutants with deletions of the genes encoding the MvaT proteins during the log and early stationary growth phases clearly suggested that their regulons were different. Indeed, significant regulon dissimilarity was found between Pmr and the other two proteins. Transcription of a larger number of genes was affected by Pmr deletion during early stationary phase than during log phase, suggesting that Pmr ameliorates the effects of pCAR1 on host fitness more effectively during the early stationary phase. Alternatively, the similarity of the TurA and TurB regulons implied that they might play complementary roles as global transcriptional regulators in response to plasmid carriage. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  19. Novel RepA-MCM proteins encoded in plasmids pTAU4, pORA1 and pTIK4 from Sulfolobus neozealandicus

    DEFF Research Database (Denmark)

    Greve, B.; Jensen, S.; Phan, H.

    2005-01-01

    Three plasmids isolated from the crenarchaeal thermoacidophile Sulfolobus neozealandicus were characterized. Plasmids pTAU4 (7,192 bp), pORA1 (9,689 bp) and pTIK4 (13,638 bp) show unusual properties that distinguish them from previously characterized cryptic plasmids of the genus Sulfolobus. Plas...

  20. Combinational deletion of three membrane protein-encoding genes highly attenuates yersinia pestis while retaining immunogenicity in a mouse model of pneumonic plague.

    Science.gov (United States)

    Tiner, Bethany L; Sha, Jian; Kirtley, Michelle L; Erova, Tatiana E; Popov, Vsevolod L; Baze, Wallace B; van Lier, Christina J; Ponnusamy, Duraisamy; Andersson, Jourdan A; Motin, Vladimir L; Chauhan, Sadhana; Chopra, Ashok K

    2015-04-01

    Previously, we showed that deletion of genes encoding Braun lipoprotein (Lpp) and MsbB attenuated Yersinia pestis CO92 in mouse and rat models of bubonic and pneumonic plague. While Lpp activates Toll-like receptor 2, the MsbB acyltransferase modifies lipopolysaccharide. Here, we deleted the ail gene (encoding the attachment-invasion locus) from wild-type (WT) strain CO92 or its lpp single and Δlpp ΔmsbB double mutants. While the Δail single mutant was minimally attenuated compared to the WT bacterium in a mouse model of pneumonic plague, the Δlpp Δail double mutant and the Δlpp ΔmsbB Δail triple mutant were increasingly attenuated, with the latter being unable to kill mice at a 50% lethal dose (LD50) equivalent to 6,800 LD50s of WT CO92. The mutant-infected animals developed balanced TH1- and TH2-based immune responses based on antibody isotyping. The triple mutant was cleared from mouse organs rapidly, with concurrent decreases in the production of various cytokines and histopathological lesions. When surviving animals infected with increasing doses of the triple mutant were subsequently challenged on day 24 with the bioluminescent WT CO92 strain (20 to 28 LD50s), 40 to 70% of the mice survived, with efficient clearing of the invading pathogen, as visualized in real time by in vivo imaging. The rapid clearance of the triple mutant, compared to that of WT CO92, from animals was related to the decreased adherence and invasion of human-derived HeLa and A549 alveolar epithelial cells and to its inability to survive intracellularly in these cells as well as in MH-S murine alveolar and primary human macrophages. An early burst of cytokine production in macrophages elicited by the triple mutant compared to WT CO92 and the mutant's sensitivity to the bactericidal effect of human serum would further augment bacterial clearance. Together, deletion of the ail gene from the Δlpp ΔmsbB double mutant severely attenuated Y. pestis CO92 to evoke pneumonic plague in a mouse model while retaining the required immunogenicity needed for subsequent protection against infection. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  1. Isolation and characterization of BetaM protein encoded by ATP1B4 - a unique member of the Na,K-ATPase β-subunit gene family

    International Nuclear Information System (INIS)

    Pestov, Nikolay B.; Zhao, Hao; Basrur, Venkatesha; Modyanov, Nikolai N.

    2011-01-01

    Highlights: → Structural properties of BetaM and Na,K-ATPase β-subunits are sharply different. → BetaM protein is concentrated in nuclear membrane of skeletal myocytes. → BetaM does not associate with a Na,K-ATPase α-subunit in skeletal muscle. → Polypeptide chain of the native BetaM is highly sensitive to endogenous proteases. → BetaM in neonatal muscle is a product of alternative splice mRNA variant B. -- Abstract: ATP1B4 genes represent a rare instance of the orthologous gene co-option that radically changed functions of encoded BetaM proteins during vertebrate evolution. In lower vertebrates, this protein is a β-subunit of Na,K-ATPase located in the cell membrane. In placental mammals, BetaM completely lost its ancestral role and through acquisition of two extended Glu-rich clusters into the N-terminal domain gained entirely new properties as a muscle-specific protein of the inner nuclear membrane possessing the ability to regulate gene expression. Strict temporal regulation of BetaM expression, which is the highest in late fetal and early postnatal myocytes, indicates that it plays an essential role in perinatal development. Here we report the first structural characterization of the native eutherian BetaM protein. It should be noted that, in contrast to structurally related Na,K-ATPase β-subunits, the polypeptide chain of BetaM is highly sensitive to endogenous proteases that greatly complicated its isolation. Nevertheless, using a complex of protease inhibitors, a sample of authentic BetaM was isolated from pig neonatal skeletal muscle by a combination of ion-exchange and lectin-affinity chromatography followed by SDS-PAGE. Results of the analysis of the BetaM tryptic digest using MALDI-TOF and ESI-MS/MS mass spectrometry have demonstrated that native BetaM in neonatal skeletal muscle is a product of alternative splice mRNA variant B and comprised of 351 amino acid residues. Isolated BetaM protein was also characterized by SELDI-TOF mass spectrometry before and after deglycosylation. This allowed us to determine that the carbohydrate moiety of BetaM has molecular mass 5.9 kDa and consists of short high-mannose type N-glycans. The results of direct analysis of the purified native eutherian BetaM protein provide first insights into structural properties underlying its entirely new evolutionarily acquired functions.

  2. Isolation and characterization of BetaM protein encoded by ATP1B4 - a unique member of the Na,K-ATPase {beta}-subunit gene family

    Energy Technology Data Exchange (ETDEWEB)

    Pestov, Nikolay B. [Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Ave., Toledo, OH 43614 (United States); Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Moscow 117997 (Russian Federation); Zhao, Hao [Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Ave., Toledo, OH 43614 (United States); Basrur, Venkatesha [Department of Pathology, University of Michigan Medical School, Ann Arbor, MI 48109 (United States); Modyanov, Nikolai N., E-mail: nikolai.modyanov@utoledo.edu [Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Ave., Toledo, OH 43614 (United States)

    2011-09-09

    Highlights: {yields} Structural properties of BetaM and Na,K-ATPase {beta}-subunits are sharply different. {yields} BetaM protein is concentrated in nuclear membrane of skeletal myocytes. {yields} BetaM does not associate with a Na,K-ATPase {alpha}-subunit in skeletal muscle. {yields} Polypeptide chain of the native BetaM is highly sensitive to endogenous proteases. {yields} BetaM in neonatal muscle is a product of alternative splice mRNA variant B. -- Abstract: ATP1B4 genes represent a rare instance of the orthologous gene co-option that radically changed functions of encoded BetaM proteins during vertebrate evolution. In lower vertebrates, this protein is a {beta}-subunit of Na,K-ATPase located in the cell membrane. In placental mammals, BetaM completely lost its ancestral role and through acquisition of two extended Glu-rich clusters into the N-terminal domain gained entirely new properties as a muscle-specific protein of the inner nuclear membrane possessing the ability to regulate gene expression. Strict temporal regulation of BetaM expression, which is the highest in late fetal and early postnatal myocytes, indicates that it plays an essential role in perinatal development. Here we report the first structural characterization of the native eutherian BetaM protein. It should be noted that, in contrast to structurally related Na,K-ATPase {beta}-subunits, the polypeptide chain of BetaM is highly sensitive to endogenous proteases that greatly complicated its isolation. Nevertheless, using a complex of protease inhibitors, a sample of authentic BetaM was isolated from pig neonatal skeletal muscle by a combination of ion-exchange and lectin-affinity chromatography followed by SDS-PAGE. Results of the analysis of the BetaM tryptic digest using MALDI-TOF and ESI-MS/MS mass spectrometry have demonstrated that native BetaM in neonatal skeletal muscle is a product of alternative splice mRNA variant B and comprised of 351 amino acid residues. Isolated BetaM protein was also characterized by SELDI-TOF mass spectrometry before and after deglycosylation. This allowed us to determine that the carbohydrate moiety of BetaM has molecular mass 5.9 kDa and consists of short high-mannose type N-glycans. The results of direct analysis of the purified native eutherian BetaM protein provide first insights into structural properties underlying its entirely new evolutionarily acquired functions.

  3. Suppressive effects of mycoviral proteins encoded by Magnaporthe oryzae chrysovirus 1 strain A on conidial germination of the rice blast fungus.

    Science.gov (United States)

    Urayama, Syun-Ichi; Kimura, Yuri; Katoh, Yu; Ohta, Tomoko; Onozuka, Nobuya; Fukuhara, Toshiyuki; Arie, Tsutomu; Teraoka, Tohru; Komatsu, Ken; Moriyama, Hiromitsu

    2016-09-02

    Magnaporthe oryzae chrysovirus 1 strain A (MoCV1-A) is the causal agent of growth repression and attenuated virulence (hypovirulence) of the rice blast fungus, Magnaporthe oryzae. We previously revealed that heterologous expression of the MoCV1-A ORF4 protein resulted in cytological damage to the yeasts Saccharomyces cerevisiae and Cryptococcus neoformans. Since the ORF4 protein is one of the components of viral particles, we evaluated the inhibitory effects of the purified virus particle against the conidial germination of M. oryzae, and confirmed its suppressive effects. Recombinant MoCV1-A ORF4 protein produced in Pichia pastoris was also effective for suppression of conidial germination of M. oryzae. MoCV1-A ORF4 protein sequence showed significant similarity to 6 related mycoviral proteins; Botrysphaeria dothidea chrysovirus 1, two Fusarium graminearum viruses, Fusarium oxysporum f. sp. dianthi mycovirus 1, Penicillium janczewski chrysovirus and Agaricus bisporus virus 1 in the Chrysoviridae family. Multiple alignments of the ORF4-related protein sequences showed that their central regions (210-591 aa in MoCV1-A ORF4) are relatively conserved. Indeed, yeast transformants expressing the conserved central region of MoCV1-A ORF4 protein (325-575 aa) showed similar impaired growth phenotypes as those observed in yeasts expressing the full-length MoCV1-A ORF4 protein. These data suggest that the mycovirus itself and its encoded viral protein can be useful as anti-fungal proteins to control rice blast disease caused by M. oryzae and other pathogenic fungi. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Identification of the major structural and nonstructural proteins encoded by human parvovirus B19 and mapping of their genes by procaryotic expression of isolated genomic fragments

    Energy Technology Data Exchange (ETDEWEB)

    Cotmore, S.F.; McKie, V.C.; Anderson, L.J.; Astell, C.R.; Tattersall, P.

    1986-11-01

    Plasma from a child with homozygous sickle-cell disease, sampled during the early phase of an aplastic crisis, contained human parvovirus B19 virions. Plasma taken 10 days later (during the convalescent phase) contained both immunoglobulin M and immunoglobulin G antibodies directed against two viral polypeptides with apparent molecular weights for 83,000 and 58,000 which were present exclusively in the particulate fraction of the plasma taken during the acute phase. These two protein species comigrated at 110S on neutral sucrose velocity gradients with the B19 viral DNA and thus appear to constitute the viral capsid polypeptides. The B19 genome was molecularly cloned into a bacterial plasmid vector. Two expression constructs containing B19 sequences from different halves of the viral genome were obtained, which directed the synthesis, in bacteria, of segments of virally encoded protein. These polypeptide fragments were then purified and used to immunize rabbits. Antibodies against a protein sequence specified between nucleotides 2897 and 3749 recognized both the 83- and 58-kilodalton capsid polypeptides in aplastic plasma taken during the acute phase and detected similar proteins in the similar proteins in the tissues of a stillborn fetus which had been infected transplacentally with B19. Antibodies against a protein sequence encoded in the other half of the B19 genome (nucleotides 1072 through 2044) did not react specifically with any protein in plasma taken during the acute phase but recognized three nonstructural polypeptides of 71, 63, and 52 kilodaltons present in the liver and, at lower levels, in some other tissues of the transplacentally infected fetus.

  5. Many Saccharomyces cerevisiae Cell Wall Protein Encoding Genes Are Coregulated by Mss11, but Cellular Adhesion Phenotypes Appear Only Flo Protein Dependent.

    Science.gov (United States)

    Bester, Michael C; Jacobson, Dan; Bauer, Florian F

    2012-01-01

    The outer cell wall of the yeast Saccharomyces cerevisiae serves as the interface with the surrounding environment and directly affects cell-cell and cell-surface interactions. Many of these interactions are facilitated by specific adhesins that belong to the Flo protein family. Flo mannoproteins have been implicated in phenotypes such as flocculation, substrate adhesion, biofilm formation, and pseudohyphal growth. Genetic data strongly suggest that individual Flo proteins are responsible for many specific cellular adhesion phenotypes. However, it remains unclear whether such phenotypes are determined solely by the nature of the expressed FLO genes or rather as the result of a combination of FLO gene expression and other cell wall properties and cell wall proteins. Mss11 has been shown to be a central element of FLO1 and FLO11 gene regulation and acts together with the cAMP-PKA-dependent transcription factor Flo8. Here we use genome-wide transcription analysis to identify genes that are directly or indirectly regulated by Mss11. Interestingly, many of these genes encode cell wall mannoproteins, in particular, members of the TIR and DAN families. To examine whether these genes play a role in the adhesion properties associated with Mss11 expression, we assessed deletion mutants of these genes in wild-type and flo11Δ genetic backgrounds. This analysis shows that only FLO genes, in particular FLO1/10/11, appear to significantly impact on such phenotypes. Thus adhesion-related phenotypes are primarily dependent on the balance of FLO gene expression.

  6. Combined virus-like particle and fusion protein-encoding DNA vaccination of cotton rats induces protection against respiratory syncytial virus without causing vaccine-enhanced disease

    International Nuclear Information System (INIS)

    Hwang, Hye Suk; Lee, Young-Tae; Kim, Ki-Hye; Park, Soojin; Kwon, Young-Man; Lee, Youri; Ko, Eun-Ju; Jung, Yu-Jin; Lee, Jong Seok; Kim, Yu-Jin; Lee, Yu-Na; Kim, Min-Chul; Cho, Minkyoung; Kang, Sang-Moo

    2016-01-01

    A safe and effective vaccine against respiratory syncytial virus (RSV) should confer protection without causing vaccine-enhanced disease. Here, using a cotton rat model, we investigated the protective efficacy and safety of an RSV combination vaccine composed of F-encoding plasmid DNA and virus-like particles containing RSV fusion (F) and attachment (G) glycoproteins (FFG-VLP). Cotton rats with FFG-VLP vaccination controlled lung viral replication below the detection limit, and effectively induced neutralizing activity and antibody-secreting cell responses. In comparison with formalin inactivated RSV (FI-RSV) causing severe RSV disease after challenge, FFG-VLP vaccination did not cause weight loss, airway hyper-responsiveness, IL-4 cytokines, histopathology, and infiltrates of proinflammatory cells such as eosinophils. FFG-VLP was even more effective in preventing RSV-induced pulmonary inflammation than live RSV infections. This study provides evidence that FFG-VLP can be developed into a safe and effective RSV vaccine candidate. - Highlights: • Combined RSV FFG VLP vaccine is effective in inducing F specific responses. • FFG VLP vaccine confers RSV neutralizing activity and viral control in cotton rats. • Cotton rats with RSV FFG VLP vaccination do not show vaccine-enhanced disease. • Cotton rats with FFG VLP vaccine induce F specific antibody secreting cell responses. • Cotton rats with FFG VLP do not induce lung cellular infiltrates and Th2 cytokine.

  7. DNA binding sites recognised in vitro by a knotted class 1 homeodomain protein encoded by the hooded gene, k, in barley (Hordeum vulgare)

    DEFF Research Database (Denmark)

    Krusell, L; Rasmussen, I; Gausing, K

    1997-01-01

    of knotted1 from maize was isolated from barley seedlings and expressed as a maltose binding protein fusion in E. coli. The purified HvH21-fusion protein selected DNA fragments with 1-3 copies of the sequence TGAC. Gel shift experiments showed that the TGAC element was required for binding and the results...

  8. [Cloning and expression analysis of a zinc-regulated transporters (ZRT), iron-regulated transporter (IRT)-like protein encoding gene in Dendrobium officinale].

    Science.gov (United States)

    Zhang, Gang; Li, Yi-Min; Li, Biao; Zhang, Da-Wei; Guo, Shun-Xing

    2015-01-01

    The zinc-regulated transporters (ZRT), iron-regulated transporter (IRT)-like protein (ZIP) plays an important role in the growth and development of plant. In this study, a full length cDNA of ZIP encoding gene, designed as DoZIP1 (GenBank accession KJ946203), was identified from Dendrobium officinale using RT-PCR and RACE. Bioinformatics analysis showed that DoZIP1 consisted of a 1,056 bp open reading frame (ORF) encoded a 351-aa protein with a molecular weight of 37.57 kDa and an isoelectric point (pI) of 6.09. The deduced DoZIP1 protein contained the conserved ZIP domain, and its secondary structure was composed of 50.71% alpha helix, 11.11% extended strand, 36.18% random coil, and beta turn 1.99%. DoZIP1 protein exhibited a signal peptide and eight transmembrane domains, presumably locating in cell membrane. The amino acid sequence had high homology with ZIP proteins from Arabidopsis, alfalfa and rice. A phylogenetic tree analysis demonstrated that DoZIP1 was closely related to AtZIP10 and OsZIP3, and they were clustered into one clade. Real time quantitative PCR analysis demonstrated that the transcription level of DoZIP1 in D. officinale roots was the highest (4.19 fold higher than that of stems), followed by that of leaves (1.12 fold). Molecular characters of DoZIP1 will be useful for further functional determination of the gene involving in the growth and development of D. officinale.

  9. Identification of the major structural and nonstructural proteins encoded by human parvovirus B19 and mapping of their genes by procaryotic expression of isolated genomic fragments

    International Nuclear Information System (INIS)

    Cotmore, S.F.; McKie, V.C.; Anderson, L.J.; Astell, C.R.; Tattersall, P.

    1986-01-01

    Plasma from a child with homozygous sickle-cell disease, sampled during the early phase of an aplastic crisis, contained human parvovirus B19 virions. Plasma taken 10 days later (during the convalescent phase) contained both immunoglobulin M and immunoglobulin G antibodies directed against two viral polypeptides with apparent molecular weights for 83,000 and 58,000 which were present exclusively in the particulate fraction of the plasma taken during the acute phase. These two protein species comigrated at 110S on neutral sucrose velocity gradients with the B19 viral DNA and thus appear to constitute the viral capsid polypeptides. The B19 genome was molecularly cloned into a bacterial plasmid vector. Two expression constructs containing B19 sequences from different halves of the viral genome were obtained, which directed the synthesis, in bacteria, of segments of virally encoded protein. These polypeptide fragments were then purified and used to immunize rabbits. Antibodies against a protein sequence specified between nucleotides 2897 and 3749 recognized both the 83- and 58-kilodalton capsid polypeptides in aplastic plasma taken during the acute phase and detected similar proteins in the similar proteins in the tissues of a stillborn fetus which had been infected transplacentally with B19. Antibodies against a protein sequence encoded in the other half of the B19 genome (nucleotides 1072 through 2044) did not react specifically with any protein in plasma taken during the acute phase but recognized three nonstructural polypeptides of 71, 63, and 52 kilodaltons present in the liver and, at lower levels, in some other tissues of the transplacentally infected fetus

  10. A Major Facilitator Superfamily protein encoded by TcMucK gene is not required for cuticle pigmentation, growth and development in Tribolium castaneum.

    Science.gov (United States)

    Mun, Seulgi; Noh, Mi Young; Osanai-Futahashi, Mizuko; Muthukrishnan, Subbaratnam; Kramer, Karl J; Arakane, Yasuyuki

    2014-06-01

    Insect cuticle pigmentation and sclerotization (tanning) are vital physiological processes for insect growth, development and survival. We have previously identified several colorless precursor molecules as well as enzymes involved in their biosynthesis and processing to yield the mature intensely colored body cuticle pigments. A recent study indicated that the Bombyx mori (silkmoth) gene, BmMucK, which encodes a protein orthologous to a Culex pipiens quiquefasciatus (Southern house mosquito) cis,cis, muconate transporter, is a member of the "Major Facilitator Superfamily" (MFS) of transporter proteins and is associated with the appearance of pigmented body segments of naturally occurring body color mutants of B. mori. While RNA interference of the BmMucK gene failed to result in any observable phenotype, RNAi using a dsRNA for an orthologous gene from the red flour beetle, Tribolium castaneum, was reported to result in molting defects and darkening of the cuticle and some body parts, leading to the suggestion that orthologs of MucK genes may differ in their functions among insects. To verify the role and essentiality of the ortholog of this gene in development and body pigmentation function in T. castaneum we obtained cDNAs for the orthologous gene (TcMucK) from RNA isolated from the GA-1 wild-type strain of T. castaneum. The sequence of a 1524 nucleotides-long cDNA for TcMucK which encodes the putatively full-length protein, was assembled from two overlapping RT-PCR fragments and the expression profile of this gene during development was analyzed by real-time PCR. This cDNA encodes a 55.8 kDa protein consisting of 507 amino acid residues and includes 11 putative transmembrane segments. Transcripts of TcMucK were detected throughout all of the developmental stages analyzed. The function of this gene was explored by injection of two different double-stranded RNAs targeting different regions of the TcMucK gene (dsTcMucKs) into young larvae to down-regulate transcripts during subsequent stages of insect development until the adult stage. RNA interference of TcMucK had no observable effects on larval, pupal or adult pigmentation. In addition, it did not affect larval-larval, larval-pupal and pupal-adult molting or survival. Thus, in contrast to the results of Zhao et al. (2012), our study demonstrates that TcMucK is not essential for growth, development or cuticle pigmentation of T. castaneum. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Pmr, a histone-like protein H1 (H-NS) family protein encoded by the IncP-7 plasmid pCAR1, is a key global regulator that alters host function.

    Science.gov (United States)

    Yun, Choong-Soo; Suzuki, Chiho; Naito, Kunihiko; Takeda, Toshiharu; Takahashi, Yurika; Sai, Fumiya; Terabayashi, Tsuguno; Miyakoshi, Masatoshi; Shintani, Masaki; Nishida, Hiromi; Yamane, Hisakazu; Nojiri, Hideaki

    2010-09-01

    Histone-like protein H1 (H-NS) family proteins are nucleoid-associated proteins (NAPs) conserved among many bacterial species. The IncP-7 plasmid pCAR1 is transmissible among various Pseudomonas strains and carries a gene encoding the H-NS family protein, Pmr. Pseudomonas putida KT2440 is a host of pCAR1, which harbors five genes encoding the H-NS family proteins PP_1366 (TurA), PP_3765 (TurB), PP_0017 (TurC), PP_3693 (TurD), and PP_2947 (TurE). Quantitative reverse transcription-PCR (qRT-PCR) demonstrated that the presence of pCAR1 does not affect the transcription of these five genes and that only pmr, turA, and turB were primarily transcribed in KT2440(pCAR1). In vitro pull-down assays revealed that Pmr strongly interacted with itself and with TurA, TurB, and TurE. Transcriptome comparisons of the pmr disruptant, KT2440, and KT2440(pCAR1) strains indicated that pmr disruption had greater effects on the host transcriptome than did pCAR1 carriage. The transcriptional levels of some genes that increased with pCAR1 carriage, such as the mexEF-oprN efflux pump genes and parI, reverted with pmr disruption to levels in pCAR1-free KT2440. Transcriptional levels of putative horizontally acquired host genes were not altered by pCAR1 carriage but were altered by pmr disruption. Identification of genome-wide Pmr binding sites by ChAP-chip (chromatin affinity purification coupled with high-density tiling chip) analysis demonstrated that Pmr preferentially binds to horizontally acquired DNA regions. The Pmr binding sites overlapped well with the location of the genes differentially transcribed following pmr disruption on both the plasmid and the chromosome. Our findings indicate that Pmr is a key factor in optimizing gene transcription on pCAR1 and the host chromosome.

  12. The effects of clobazam treatment in rats on the expression of genes and proteins encoding glucronosyltransferase 1A/2B (UGT1A/2B) and multidrug resistance‐associated protein-2 (MRP2), and development of thyroid follicular cell hypertrophy

    Energy Technology Data Exchange (ETDEWEB)

    Miyawaki, Izuru, E-mail: izuru-miyawaki@ds-pharma.co.jp; Tamura, Akitoshi; Matsumoto, Izumi; Inada, Hiroshi; Kunimatsu, Takeshi; Kimura, Juki; Funabashi, Hitoshi

    2012-12-15

    Clobazam (CLB) is known to increase hepatobiliary thyroxine (T4) clearance in Sprague–Dawley (SD) rats, which results in hypothyroidism followed by thyroid follicular cell hypertrophy. However, the mechanism of the acceleration of T4-clearance has not been fully investigated. In the present study, we tried to clarify the roles of hepatic UDP-glucronosyltransferase (UGT) isoenzymes (UGT1A and UGT2B) and efflux transporter (multidrug resistance–associated protein-2; MRP2) in the CLB-induced acceleration of T4-clearance using two mutant rat strains, UGT1A-deficient mutant (Gunn) and MRP2-deficient mutant (EHBR) rats, especially focusing on thyroid morphology, levels of circulating hormones (T4 and triiodothyronine (T3)) and thyroid-stimulating hormone (TSH), and mRNA or protein expressions of UGTs (Ugt1a1, Ugt1a6, and Ugt2b1/2) and MRP2 (Mrp). CLB induced thyroid morphological changes with increases in TSH in SD and Gunn rats, but not in EHBR rats. T4 was slightly decreased in SD and Gunn rats, and T3 was decreased in Gunn rats, whereas these hormones were maintained in EHBR rats. Hepatic Ugt1a1, Ugt1a6, Ugt2b1/2, and Mrp2 mRNAs were upregulated in SD rats. In Gunn rats, UGT1A mRNAs (Ugt1a1/6) and protein levels were quite low, but UGT2B mRNAs (Ugt2b1/2) and protein were prominently upregulated. In SD and Gunn rats, MRP2 mRNA and protein were upregulated to the same degree. These results suggest that MRP2 is an important contributor in development of the thyroid cellular hypertrophy in CLB-treated rats, and that UGT1A and UGT2B work in concert with MRP2 in the presence of MRP2 function to enable the effective elimination of thyroid hormones. -- Highlights: ► Role of UGT and MRP2 in thyroid pathology was investigated in clobazam-treated rats. ► Clobazam induced thyroid cellular hypertrophy in SD and Gunn rats, but not EHBR rats. ► Hepatic Mrp2 gene and protein were upregulated in SD and Gunn rats, but not EHBR rats. ► Neither serum thyroid hormones (T3/T4) nor thyroid pathology changed in EHBR rats. ► Mrp2 was implied to be a key molecule in clobazam-induced thyroid pathology in rats.

  13. The effects of clobazam treatment in rats on the expression of genes and proteins encoding glucronosyltransferase 1A/2B (UGT1A/2B) and multidrug resistance‐associated protein-2 (MRP2), and development of thyroid follicular cell hypertrophy

    International Nuclear Information System (INIS)

    Miyawaki, Izuru; Tamura, Akitoshi; Matsumoto, Izumi; Inada, Hiroshi; Kunimatsu, Takeshi; Kimura, Juki; Funabashi, Hitoshi

    2012-01-01

    Clobazam (CLB) is known to increase hepatobiliary thyroxine (T4) clearance in Sprague–Dawley (SD) rats, which results in hypothyroidism followed by thyroid follicular cell hypertrophy. However, the mechanism of the acceleration of T4-clearance has not been fully investigated. In the present study, we tried to clarify the roles of hepatic UDP-glucronosyltransferase (UGT) isoenzymes (UGT1A and UGT2B) and efflux transporter (multidrug resistance–associated protein-2; MRP2) in the CLB-induced acceleration of T4-clearance using two mutant rat strains, UGT1A-deficient mutant (Gunn) and MRP2-deficient mutant (EHBR) rats, especially focusing on thyroid morphology, levels of circulating hormones (T4 and triiodothyronine (T3)) and thyroid-stimulating hormone (TSH), and mRNA or protein expressions of UGTs (Ugt1a1, Ugt1a6, and Ugt2b1/2) and MRP2 (Mrp). CLB induced thyroid morphological changes with increases in TSH in SD and Gunn rats, but not in EHBR rats. T4 was slightly decreased in SD and Gunn rats, and T3 was decreased in Gunn rats, whereas these hormones were maintained in EHBR rats. Hepatic Ugt1a1, Ugt1a6, Ugt2b1/2, and Mrp2 mRNAs were upregulated in SD rats. In Gunn rats, UGT1A mRNAs (Ugt1a1/6) and protein levels were quite low, but UGT2B mRNAs (Ugt2b1/2) and protein were prominently upregulated. In SD and Gunn rats, MRP2 mRNA and protein were upregulated to the same degree. These results suggest that MRP2 is an important contributor in development of the thyroid cellular hypertrophy in CLB-treated rats, and that UGT1A and UGT2B work in concert with MRP2 in the presence of MRP2 function to enable the effective elimination of thyroid hormones. -- Highlights: ► Role of UGT and MRP2 in thyroid pathology was investigated in clobazam-treated rats. ► Clobazam induced thyroid cellular hypertrophy in SD and Gunn rats, but not EHBR rats. ► Hepatic Mrp2 gene and protein were upregulated in SD and Gunn rats, but not EHBR rats. ► Neither serum thyroid hormones (T3/T4) nor thyroid pathology changed in EHBR rats. ► Mrp2 was implied to be a key molecule in clobazam-induced thyroid pathology in rats.

  14. YKL-40: a novel marker shared by chronic inflammation and oncogenic transformation

    DEFF Research Database (Denmark)

    Roslind, Anne; Johansen, Julia S

    2009-01-01

    YKL-40, a member of 'mammalian chitinase-like proteins', is secreted by macrophages, neutrophils, chondrocytes, endothelial-, vascular smooth muscle-, and cancer cells. High serum YKL-40 is a biomarker of poor prognosis in patients with cancer, inflammation and increased tissue remodelling. High...

  15. Cerebrospinal fluid chitinase-3-like 2 and chitotriosidase are potential prognostic biomarkers in early multiple sclerosis

    DEFF Research Database (Denmark)

    Møllgaard, M; Degn, M; Sellebjerg, F

    2016-01-01

    : In a prospective cohort of 73 patients with ON as a first demyelinating episode and 26 age-matched healthy controls levels of CHI3L2 and chitotriosidase in CSF were explored by enzyme-linked immunosorbent assay. Associations with magnetic resonance imaging white matter lesions, CSF oligoclonal bands......BACKGROUND AND PURPOSE: The role of chitinases and chitinase-like proteins in multiple sclerosis (MS) is currently unknown; however, cerebrospinal fluid (CSF) levels of chitinase 3-like 1 (CHI3L1) predict prognosis in early MS. Whether this applies to other chitinases and chitinase-like proteins...... is yet to be established. Our objective was to investigate the potential of chitinase 3-like 2 (CHI3L2) and chitotriosidase as prognostic biomarkers in optic neuritis (ON) as the first demyelinating episode and to evaluate the ability of CHI3L2 to predict long-term MS risk and disability. METHODS...

  16. Simple method for identification of plasmid-coded proteins

    International Nuclear Information System (INIS)

    Sancar, A.; Hack, A.M.; Rupp, W.D.

    1979-01-01

    Proteins encoded by plasmid DNA are specifically labeled in uv-irradiated cells of Escherichia coli carrying recA and uvrA mutations because extensive degradation of the chromosome DNA occurs concurrently with amplification of plasmid DNA

  17. Journal of Genetics, Volume 92, 2013

    Indian Academy of Sciences (India)

    tropic mutants of Mendelian inheritance in Catharanthus roseus. (Research ... bioinformatics. Molecular adaptation within the coat protein-encoding gene of .... based NCII designs: a simulation study (Research article) 529 common bean.

  18. Journal of Genetics, Volume 92, 2013

    Indian Academy of Sciences (India)

    Molecular adaptation within the coat protein-encoding gene of Tunisian almond ... Neurospora crassa ncs-1, mid-1 and nca-2 double-mutant phe- notypes suggest .... based NCII designs: a simulation study (Research article) 529. Li, Shujuan.

  19. Sub classification and targeted characterization of prophage ...

    Indian Academy of Sciences (India)

    PRAKASH KUMAR

    2007-06-21

    Jun 21, 2007 ... structure-function characterization of proteins encoded by cryptic prophages will help understand the contribution of prophage .... distilled water, collected by centrifugation and suspended in. 1 ml of .... Pdb homolog. Acc no.

  20. Journal of Biosciences | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    Observations on sporozoite detection in naturally infected sibling species of the ... of green fluorescent protein and -glucuronidase protein-encoding genes as a .... Spatio-temporal tumour model for analysis and mechanism of action of ...

  1. The RAS Initiative

    Science.gov (United States)

    NCI established the RAS Initiative to explore innovative approaches for attacking the proteins encoded by mutant forms of RAS genes and to ultimately create effective, new therapies for RAS-related cancers.

  2. Topology of a Membrane Associated Regulator of Prokaryotic DNA Replication

    National Research Council Canada - National Science Library

    Firshein, William

    1998-01-01

    This proposal has focused on a broad host range plasmid, RK2, as a model system to study how a pair of initiation proteins encoded by the plasmid for DNA replication function when replication occurs...

  3. Regulation of YKL-40 expression during genotoxic or microenvironmental stress in human glioblastoma cells

    DEFF Research Database (Denmark)

    Junker, Nanna; Johansen, Julia S; Hansen, Lasse T

    2005-01-01

    YKL-40 is a 40 kDa secreted glycoprotein belonging to the family of 'mammalian chitinase-like proteins', but without chitinase activity. YKL-40 has a proliferative effect on fibroblasts, chondrocytes and synoviocytes, and chemotactic effect on endothelium and vascular smooth muscle cells. Elevated...... material from glioblastomas patients. We investigated the expression of YKL-40 in three human malignant glioma cell lines exposed to different types of stress. Whereas a polymerase chain reaction transcript was detectable in all three cell lines, only U87 produced measurable amounts of YKL-40 protein. In U...... is attenuated by p53. In contrast, both basic fibroblast growth factor and tumor necrosing factor-alpha repressed YKL-40. These are the first data on regulation of YKL-40 in cancer cells. Diverse types of stress resulted in YKL-40 elevation, which strongly supports an involvement of YKL-40 in the malignant...

  4. The genome of Paenibacillus sabinae T27 provides insight into evolution, organization and functional elucidation of nif and nif-like genes

    OpenAIRE

    Li, Xinxin; Deng, Zhiping; Liu, Zhanzhi; Yan, Yongliang; Wang, Tianshu; Xie, Jianbo; Lin, Min; Cheng, Qi; Chen, Sanfeng

    2014-01-01

    Background Most biological nitrogen fixation is catalyzed by the molybdenum nitrogenase. This enzyme is a complex which contains the MoFe protein encoded by nifDK and the Fe protein encoded by nifH. In addition to nifHDK, nifHDK-like genes were found in some Archaea and Firmicutes, but their function is unclear. Results We sequenced the genome of Paenibacillus sabinae T27. A total of 4,793 open reading frames were predicted from its 5.27 Mb genome. The genome of P. sabinae T27 contains fiftee...

  5. Materials and methods to increase plant growth and yield

    Science.gov (United States)

    Kirst, Matias

    2017-05-16

    The present invention relates to materials and methods for modulating growth rates, yield, and/or resistance to drought conditions in plants. In one embodiment, a method of the invention comprises increasing expression of an hc1 gene (or a homolog thereof that provides for substantially the same activity), or increasing expression or activity of the protein encoded by an hc1 gene thereof, in a plant, wherein expression of the hc1 gene or expression or activity of the protein encoded by an hc1 gene results in increased growth rate, yield, and/or drought resistance in the plant.

  6. Chitins and Chitosans as Immunoadjuvants and Non-Allergenic Drug Carriers

    Directory of Open Access Journals (Sweden)

    Riccardo A. A. Muzzarelli

    2010-02-01

    Full Text Available Due to the fact that some individuals are allergic to crustaceans, the presumed relationship between allergy and the presence of chitin in crustaceans has been investigated. In vivo, chitin is part of complex structures with other organic and inorganic compounds: in arthropods chitin is covalently linked to proteins and tanned by quinones, in fungi it is covalently linked to glucans, while in bacteria chitin is diversely combined according to Gram(+/- classification. On the other hand, isolated, purified chitin is a plain polysaccharide that, at the nano level, presents itself as a highly associated structure, recently refined in terms of regularity, nature of bonds, crystallinity degree and unusual colloidal behavior. Chitins and modified chitins exert a number of beneficial actions, i.e., (i they stimulate macrophages by interacting with receptors on the macrophage surface that mediate the internalization of chitin particles to be degraded by lysozyme and N-acetyl-β-glucosaminidase (such as Nod-like, Toll-like, lectin, Dectin-1, leukotriene 134 and mannose receptors; (ii the macrophages produce cytokines and other compounds that confer non-specific host resistance against bacterial and viral infections, and anti-tumor activity; (iii chitin is a strong Th1 adjuvant that up-regulates Th1 immunity induced by heat-killed Mycobacterium bovis, while down- regulating Th2 immunity induced by mycobacterial protein; (iv direct intranasal application of chitin microparticles into the lung was also able to significantly down-regulate allergic response to Dermatophagoids pteronyssinus and Aspergillus fumigatus in a murine model of allergy; (v chitin microparticles had a beneficial effect in preventing and treating histopathologic changes in the airways of asthmatic mice; (vi authors support the fact that chitin depresses the development of adaptive type 2 allergic responses. Since the expression of chitinases, chitrotriosidase and chitinase-like proteins

  7. Chitins and chitosans as immunoadjuvants and non-allergenic drug carriers.

    Science.gov (United States)

    Muzzarelli, Riccardo A A

    2010-02-21

    Due to the fact that some individuals are allergic to crustaceans, the presumed relationship between allergy and the presence of chitin in crustaceans has been investigated. In vivo, chitin is part of complex structures with other organic and inorganic compounds: in arthropods chitin is covalently linked to proteins and tanned by quinones, in fungi it is covalently linked to glucans, while in bacteria chitin is diversely combined according to Gram(+/-) classification. On the other hand, isolated, purified chitin is a plain polysaccharide that, at the nano level, presents itself as a highly associated structure, recently refined in terms of regularity, nature of bonds, crystallinity degree and unusual colloidal behavior. Chitins and modified chitins exert a number of beneficial actions, i.e., (i) they stimulate macrophages by interacting with receptors on the macrophage surface that mediate the internalization of chitin particles to be degraded by lysozyme and N-acetyl-beta-glucosaminidase (such as Nod-like, Toll-like, lectin, Dectin-1, leukotriene 134 and mannose receptors); (ii) the macrophages produce cytokines and other compounds that confer non-specific host resistance against bacterial and viral infections, and anti-tumor activity; (iii) chitin is a strong Th1 adjuvant that up-regulates Th1 immunity induced by heat-killed Mycobacterium bovis, while down- regulating Th2 immunity induced by mycobacterial protein; (iv) direct intranasal application of chitin microparticles into the lung was also able to significantly down-regulate allergic response to Dermatophagoids pteronyssinus and Aspergillus fumigatus in a murine model of allergy; (v) chitin microparticles had a beneficial effect in preventing and treating histopathologic changes in the airways of asthmatic mice; (vi) authors support the fact that chitin depresses the development of adaptive type 2 allergic responses. Since the expression of chitinases, chitrotriosidase and chitinase-like proteins is greatly

  8. From reverse transcription to human brain tumors

    Directory of Open Access Journals (Sweden)

    Dmitrenko V. V.

    2013-05-01

    Full Text Available Reverse transcriptase from avian myeloblastosis virus (AMV was the subject of the study, from which the investi- gations of the Department of biosynthesis of nucleic acids were started. Production of AMV in grams quantities and isolation of AMV reverse transcriptase were established in the laboratory during the seventies of the past cen- tury and this initiated research on the cDNA synthesis, cloning and investigation of the structure and functions of the eukaryotic genes. Structures of salmon insulin and insulin-like growth factor (IGF family genes and their transcripts were determined during long-term investigations. Results of two modern techniques, microarray-ba- sed hybridization and SAGE, were used for the identification of the genes differentially expressed in astrocytic gliomas and human normal brain. Comparison of SAGE results on the genes overexpressed in glioblastoma with the results of microarray analysis revealed a limited number of common genes. 105 differentially expressed genes, common to both methods, can be included in the list of candidates for the molecular typing of glioblastoma. The first experiments on the classification of glioblastomas based on the data of the 20 genes expression were conducted by using of artificial neural network analysis. The results of these experiments showed that the expression profiles of these genes in 224 glioblastoma samples and 74 normal brain samples could be according to the Koho- nen’s maps. The CHI3L1 and CHI3L2 genes of chitinase-like cartilage protein were revealed among the most overexpressed genes in glioblastoma, which could have prognostic and diagnostic potential. Results of in vitro experiments demonstrated that both proteins, CHI3L1 and CHI3L2, may initiate the phosphorylation of ERK1/ ERK2 and AKT kinases leading to the activation of MAPK/ERK1/2 and PI3K/AKT signaling cascades in human embryonic kidney 293 cells, human glioblastoma U87MG, and U373 cells. The new human cell line

  9. Fast neutron radiation induced Glu-B1 deficient lines of an elite bread wheat variety

    Science.gov (United States)

    Five isogenic wheat lines deficient in high-molecular weight subunit (HMW-GS) proteins encoded by the B-genome were identified from a fast-neutron radiation-mutagenized population of Summit, an elite variety of bread wheat (Triticum aestivum L.). The mutant lines differ from the wild-type progenit...

  10. Fine-scale distribution patterns of Synechococcus ecological diversity in the microbial mats of Mushroom Spring, Yellowstone National Park

    DEFF Research Database (Denmark)

    Becraft, E.; Cohan, F.; Kühl, Michael

    2011-01-01

    Past analyses of sequence diversity in high-resolution protein-encoding genes have identified putative ecological species of unicellular cyanobacteria in the genus Synechococcus, which are specialized to 60°C but not 65°C in Mushroom Spring microbial mats. Because these studies were limited to only...

  11. Exploring the link between MORF4L1 and risk of breast cancer

    NARCIS (Netherlands)

    Martrat, G.; Maxwell, C.A.; Tominaga, E.; Porta-de-la-Riva, M.; Bonifaci, N.; Gomez-Baldo, L.; Bogliolo, M.; Lazaro, C.; Blanco, I.; Brunet, J.; Aguilar, H.; Fernandez-Rodriguez, J.; Seal, S.; Renwick, A.; Rahman, N.; Kuehl, J.; Neveling, K.; Schindler, D.; Ramirez, M.J.; Castella, M.; Hernandez, G.; Easton, D.F.; Peock, S.; Cook, M.; Oliver, C.T.; Frost, D.; Platte, R.; Evans, D.G.; Lalloo, F.; Eeles, R.; Izatt, L.; Chu, C.; Davidson, R.; Ong, K.R.; Cook, J.; Douglas, F.; Hodgson, S.; Brewer, C.; Morrison, P.J.; Porteous, M.; Peterlongo, P.; Manoukian, S.; Peissel, B.; Zaffaroni, D.; Roversi, G.; Barile, M.; Viel, A.; Pasini, B.; Ottini, L.; Putignano, A.L.; Savarese, A.; Bernard, L.; Radice, P.; Healey, S.; Spurdle, A.; Chen, X.; Beesley, J.; Rookus, M.A.; Verhoef, S.; Tilanus-Linthorst, M.A.; Vreeswijk, M.P.; van Asperen, C.J.; Bodmer, D.; Ausems, M.G.E.M.; van Os, T.A.; Blok, M.J.; Meijers-Heijboer, H.E.J.; Hogervorst, F.B.L.; Goldgar, D.E.; Buys, S.; John, E.M.; Miron, A.; Southey, M.; Daly, M.B.; Harbst, K.; Borg, A.; Rantala, J.; Barbany-Bustinza, G.; Ehrencrona, H.; Stenmark-Askmalm, M.; Kaufman, B.; Laitman, Y.; Milgrom, R.; Friedman, E.; Domchek, S.M.; Nathanson, K.L.; Rebbeck, T.R.; Johannsson, O.T.; Couch, F.J.; Wang, X.; Fredericksen, Z.; Cuadras, D.; Moreno, V.; Pientka, F.K.; Depping, R.; Caldes, T.; Osorio, A.; Benitez, J.; Bueren, J.; Heikkinen, T.; Nevanlinna, H.; Hamann, U.; Torres, D.; Caligo, M.A.; Godwin, A.K.; Imyanitov, E.N.; Janavicius, R.; Sinilnikova, O.M.; Stoppa-Lyonnet, D.; Mazoyer, S.; Verny-Pierre, C.; Castera, L.; de Pauw, A.; Bignon, Y.J.; Uhrhammer, N.; Peyrat, J.P.; Vennin, P.; Ferrer, S.F.; Collonge-Rame, M.A.; Mortemousque, I.; McGuffog, L.; Chenevix-Trench, G.; Pereira-Smith, O.M.; Antoniou, A.C.; Ceron, J.; Tominaga, K.; Surralles, J.; Pujana, M.A.

    2011-01-01

    Introduction: Proteins encoded by Fanconi anemia (FA) and/or breast cancer (BrCa) susceptibility genes cooperate in a common DNA damage repair signaling pathway. To gain deeper insight into this pathway and its influence on cancer risk, we searched for novel components through protein physical

  12. Exploring the link between MORF4L1 and risk of breast cancer.

    NARCIS (Netherlands)

    Martrat, G.; Maxwell, C.M.; Tominaga, E.; Porta-de-la-Riva, M.; Bonifaci, N.; Gomez-Baldo, L.; Bogliolo, M.; Lazaro, C.; Blanco, I.; Brunet, J.; Aguilar, H.; Fernandez-Rodriguez, J.; Seal, S.; Renwick, A.; Rahman, N.; Kuhl, J.; Neveling, K.; Schindler, D.; Ramirez, M.J.; Castella, M.; Hernandez, G.; Easton, D.F.; Peock, S.; Cook, M.; Oliver, C.T.; Frost, D.; Platte, R.; Evans, D.G.; Lalloo, F.; Eeles, R.; Izatt, L.; Chu, C.; Davidson, R.; Ong, K.R.; Cook, J.; Douglas, F.; Hodgson, S.; Brewer, C.; Morrison, P.J.; Porteous, M.; Peterlongo, P.; Manoukian, S.; Peissel, B.; Zaffaroni, D.; Roversi, G.; Barile, M.; Viel, A.; Pasini, B.; Ottini, L.; Putignano, A.L.; Savarese, A.; Bernard, L.; Radice, P.; Healey, S.; Spurdle, A.; Chen, X.; Beesley, J.; Rookus, M.A.; Verhoef, S.; Tilanus-Linthorst, M.A.; Vreeswijk, M.P.; Asperen, C.J. van; Bodmer, D.; Ausems, M.G.; Os, T.A. van; Blok, M.J.; Meijers-Heijboer, H.E.; Hogervorst, F.B.L.; Goldgar, D.E.; Buys, S.; John, E.M.; Miron, A.; Southey, M.; Daly, M.B.; Harbst, K.; Borg, A.; Rantala, J.; Barbany-Bustinza, G.; Ehrencrona, H.; Stenmark-Askmalm, M.; Kaufman, B.; Laitman, Y.; Milgrom, R.; Friedman, E.; Domchek, S.M.; Nathanson, K.L.; Rebbeck, T.R.; Johannsson, O.T.; Couch, F.J.; Wang, X.; Fredericksen, Z.; Cuadras, D.; Moreno, V.; Pientka, F.K.; Depping, R.; Caldes, T.; Osorio, A.; Benitez, J.; Bueren, J.; Heikkinen, T.; Nevanlinna, H.; Hamann, U.; Torres, D.; Caligo, M.A.; Godwin, A.K.; Imyanitov, E.N.; Janavicius, R.; Sinilnikova, O.M.; Stoppa-Lyonnet, D.; Mazoyer, S.; Verny-Pierre, C.; Castera, L.; Pauw, A. de; Bignon, Y.J.; Uhrhammer, N.; Peyrat, J.P.; Vennin, P.; Ferrer, S.F.; Collonge-Rame, M.A.; Mortemousque, I.; McGuffog, L.; Chenevix-Trench, G.; Pereira-Smith, O.M.; Antoniou, A.C.; Ceron, J.; Tominaga, K.; Surralles, J.; Pujana, M.A.

    2011-01-01

    INTRODUCTION: Proteins encoded by Fanconi anemia (FA) and/or breast cancer (BrCa) susceptibility genes cooperate in a common DNA damage repair signaling pathway. To gain deeper insight into this pathway and its influence on cancer risk, we searched for novel components through protein physical

  13. Exploring the link between MORF4L1 and risk of breast cancer

    NARCIS (Netherlands)

    Martrat, Griselda; Maxwell, Christopher M.; Tominaga, Emiko; Porta-de-la-Riva, Montserrat; Bonifaci, Núria; Gómez-Baldó, Laia; Bogliolo, Massimo; Lázaro, Conxi; Blanco, Ignacio; Brunet, Joan; Aguilar, Helena; Fernández-Rodríguez, Juana; Seal, Sheila; Renwick, Anthony; Rahman, Nazneen; Kühl, Julia; Neveling, Kornelia; Schindler, Detlev; Ramírez, María J.; Castellà, María; Hernández, Gonzalo; Easton, Douglas F.; Peock, Susan; Cook, Margaret; Oliver, Clare T.; Frost, Debra; Platte, Radka; Evans, D. Gareth; Lalloo, Fiona; Eeles, Rosalind; Izatt, Louise; Chu, Carol; Davidson, Rosemarie; Ong, Kai-Ren; Cook, Jackie; Douglas, Fiona; Hodgson, Shirley; Brewer, Carole; Morrison, Patrick J.; Porteous, Mary; Peterlongo, Paolo; Manoukian, Siranoush; Peissel, Bernard; Zaffaroni, Daniela; Roversi, Gaia; Barile, Monica; Viel, Alessandra; Pasini, Barbara; Ottini, Laura; Putignano, Anna Laura; Savarese, Antonella; Bernard, Loris; Radice, Paolo; Healey, Sue; Spurdle, Amanda; Chen, Xiaoqing; Beesley, Jonathan; Rookus, Matti A.; Verhoef, Senno; Tilanus-Linthorst, Madeleine A.; Vreeswijk, Maaike P.; Asperen, Christi J.; Bodmer, Danielle; Ausems, Margreet G. E. M.; van Os, Theo A.; Blok, Marinus J.; Meijers-Heijboer, Hanne E. J.; Hogervorst, Frans B. L.; Goldgar, David E.; Buys, Saundra; John, Esther M.; Miron, Alexander; Southey, Melissa; Daly, Mary B.; Harbst, Katja; Borg, Ake; Rantala, Johanna; Barbany-Bustinza, Gisela; Ehrencrona, Hans; Stenmark-Askmalm, Marie; Kaufman, Bella; Laitman, Yael; Milgrom, Roni; Friedman, Eitan; Domchek, Susan M.; Nathanson, Katherine L.; Rebbeck, Timothy R.; Johannsson, Oskar Thor; Couch, Fergus J.; Wang, Xianshu; Fredericksen, Zachary; Cuadras, Daniel; Moreno, Víctor; Pientka, Friederike K.; Depping, Reinhard; Caldés, Trinidad; Osorio, Ana; Benítez, Javier; Bueren, Juan; Heikkinen, Tuomas; Nevanlinna, Heli; Hamann, Ute; Torres, Diana; Caligo, Maria Adelaide; Godwin, Andrew K.; Imyanitov, Evgeny N.; Janavicius, Ramunas; Sinilnikova, Olga M.; Stoppa-Lyonnet, Dominique; Mazoyer, Sylvie; Verny-Pierre, Carole; Castera, Laurent; de Pauw, Antoine; Bignon, Yves-Jean; Uhrhammer, Nancy; Peyrat, Jean-Philippe; Vennin, Philippe; Ferrer, Sandra Fert; Collonge-Rame, Marie-Agnès; Mortemousque, Isabelle; McGuffog, Lesley; Chenevix-Trench, Georgia; Pereira-Smith, Olivia M.; Antoniou, Antonis C.; Cerón, Julián; Tominaga, Kaoru; Surrallés, Jordi; Pujana, Miguel Angel

    2011-01-01

    Proteins encoded by Fanconi anemia (FA) and/or breast cancer (BrCa) susceptibility genes cooperate in a common DNA damage repair signaling pathway. To gain deeper insight into this pathway and its influence on cancer risk, we searched for novel components through protein physical interaction

  14. Exploring the link between MORF4L1 and risk of breast cancer

    NARCIS (Netherlands)

    G. Martrat (Griselda); C.A. Maxwell (Christopher); E. Tominaga (Emiko); M. Porta-de-la-Riva (Montserrat); N. Bonifaci (Núria); L. Gómez-Baldó (Laia); M. Bogliolo (Massimo); C. Lázaro (Conxi); I. Blanco (Ignacio); J. Brunet (Joan); H. Aguilar (Helena); J. Fernández-Rodríguez (Juana); S. Seal (Sheila); A. Renwick (Anthony); N. Rahman (Nazneen); J. Kühl (Julia); K. Neveling (Kornelia); D. Schindler (Detlev); M.J. Ramírez (María); M. Castellà (María); G. Hernández (Gonzalo); D.F. Easton (Douglas); S. Peock (Susan); M. Cook (Margaret); C.T. Oliver (Clare); D. Frost (Debra); R. Platte (Radka); D.G. Evans (Gareth); F. Lalloo (Fiona); R. Eeles (Rosalind); L. Izatt (Louise); C. Chu (Chengbin); R. Davidson (Rosemarie); K. Ong; F. Douglas (Fiona); S.V. Hodgson (Shirley); C. Brewer (Carole); P.J. Morrison (Patrick); M.E. Porteous (Mary); P. Peterlongo (Paolo); S. Manoukian (Siranoush); B. Peissel (Bernard); D. Zaffaroni (Daniela); G. Roversi (Gaia); M. Barile (Monica); A. Viel (Alessandra); B. Pasini (Barbara); L. Ottini (Laura); A.L. Putignano; A. Savarese (Antonella); L. Bernard (Loris); P. Radice (Paolo); S. Healey (Sue); A.B. Spurdle (Amanda); X. Chen (Xiaoqing); J. Beesley (Jonathan); M.A. Rookus (Matti); S. Verhoef; M.M.A. Tilanus-Linthorst (Madeleine); M.P. Vreeswijk (Maaike); D. Bodmer (Danielle); M.G.E.M. Ausems (Margreet); T.A.M. van Os (Theo); M.J. Blok (Marinus); E.J. Meijers-Heijboer (Hanne); F.B.L. Hogervorst (Frans); D. Goldgar (David); S.S. Buys (Saundra); E.M. John (Esther); A. Miron (Alexander); M.C. Southey (Melissa); M.J. Daly (Mark); K. Harbst (Katja); Å. Borg (Åke); J. Rantala (Johanna); G. Barbany-Bustinza (Gisela); H. Ehrencrona (Hans); M. Stenmark-Askmalm (M.); B. Kaufman (Bella); Y. Laitman (Yael); R. Milgrom (Roni); E. Friedman (Eitan); S.M. Domchek (Susan); K.L. Nathanson (Katherine); R. Rebbeck (Timothy); O.T. Johannson (Oskar); F.J. Couch (Fergus); X. Wang (Xing); Z. Fredericksen (Zachary); D. Cuadras (Daniel); V. Moreno (Víctor); F.K. Pientka (Friederike); R. Depping (Reinhard); T. Caldes (Trinidad); A. Osorio (Ana); J. Benítez (Javier); J. Bueren (Juan); T. Heikinen (Tuomas); H. Nevanlinna (Heli); U. Hamann (Ute); D. Torres (Diana); M.A. Caligo (Maria); A.K. Godwin (Andrew); E.N. Imyanitov (Evgeny); R. Janavicius (Ramunas); O. Sinilnikova (Olga); D. Stoppa-Lyonnet (Dominique); S. Mazoyer (Sylvie); C. Verny-Pierre (Carole); L. Castera (Laurent); A. de Pauw (Antoine); Y.-J. Bignon (Yves-Jean); N. Uhrhammer (Nancy); J.-P. Peyrat; P. Vennin (Philippe); S.F. Ferrer; M.-A. Collonge-Rame; I. Mortemousque (Isabelle); L. McGuffog (Lesley); G. Chenevix-Trench (Georgia); O.M. Pereira-Smith (Olivia); A.C. Antoniou (Antonis); J. Cerón (Julián); J. Surrallés (Jordi); M.A. Pujana (Miguel); C.J. van Asperen (Christi)

    2011-01-01

    textabstractIntroduction: Proteins encoded by Fanconi anemia (FA) and/or breast cancer (BrCa) susceptibility genes cooperate in a common DNA damage repair signaling pathway. To gain deeper insight into this pathway and its influence on cancer risk, we searched for novel components through protein

  15. Complete Genome Sequence of the Methanococcus maripaludis Type Strain JJ (DSM 2067), a Model for Selenoprotein Synthesis in Archaea.

    Science.gov (United States)

    Poehlein, Anja; Heym, Daniel; Quitzke, Vivien; Fersch, Julia; Daniel, Rolf; Rother, Michael

    2018-04-05

    Methanococcus maripaludis type strain JJ (DSM 2067) is an important organism because it serves as a model for primary energy metabolism and hydrogenotrophic methanogenesis and is amenable to genetic manipulation. The complete genome (1.7 Mb) harbors 1,815 predicted protein-encoding genes, including 9 encoding selenoproteins. Copyright © 2018 Poehlein et al.

  16. Clinical and pathological associations with p53 tumour-suppressor gene mutations and expression of p21WAF1/Cip1 in colorectal carcinoma

    NARCIS (Netherlands)

    Slebos, R. J.; Baas, I. O.; Clement, M.; Polak, M.; Mulder, J. W.; van den Berg, F. M.; Hamilton, S. R.; Offerhaus, G. J.

    1996-01-01

    Inactivation of the p53 tumour-suppressor gene is common in a wide variety of human neoplasms. In the majority of cases, single point mutations in the protein-encoding sequence of p53 lead to positive immunohistochemistry (IHC) for the p53 protein, and are accompanied by loss of the wild-type

  17. Polyethylene glycol (PEG)-mediated transformation of the fused egfp ...

    African Journals Online (AJOL)

    Yomi

    2012-03-06

    Mar 6, 2012 ... pUC19 vector. Enhanced green fluorescent protein-encoding gene (egfp) and a selection marker, the hygromycin ... agro-industrial waste bioconversion (Shabtay et al., 2009;. Salvachúa et al., 2011); and .... gene, two pairs of primers were designed based on the Lentinula edodes sequences published by ...

  18. Different Stationary Phase Selectivities and Morphologies for Intact Protein Separations

    NARCIS (Netherlands)

    Astefanei, A.; Dapic, I.; Camenzuli, M.

    The central dogma of biology proposed that one gene encodes for one protein. We now know that this does not reflect reality. The human body has approximately 20,000 protein-encoding genes; each of these genes can encode more than one protein. Proteins expressed from a single gene can vary in terms

  19. First Insights into the Genome Sequence of Clostridium thermopalmarium DSM 5974, a Butyrate-Producing Bacterium Isolated from Palm Wine.

    Science.gov (United States)

    Poehlein, Anja; Hettwer, Eva; Mohnike, Lennart; Daniel, Rolf

    2018-04-26

    Clostridium thermopalmarium is a moderate thermophilic, rod-shaped, and endospore-forming bacterium, which was isolated from palm wine in Senegal. Butyrate is produced from a broad variety of sugar substrates. Here, we present the draft genome sequence of C. thermopalmarium DSM 5974 (2.822 Mb) containing 2,665 predicted protein-encoding genes. Copyright © 2018 Poehlein et al.

  20. Protein folding on a chip

    CERN Multimedia

    2004-01-01

    "Scientists at the U.S. Department of Energy's Brookhaven National Laboratory are proposing to use a super- computer originally developed to simulate elementary particles in high- energy physics to help determine the structures and functions of proteins, including, for example, the 30,000 or so proteins encoded by the human genome" (1 page)

  1. First Insights into the Genome of Fructobacillus sp. EFB-N1, Isolated from Honey Bee Larva Infected with European Foulbrood

    OpenAIRE

    Djukic, Marvin; Daniel, Rolf; Poehlein, Anja

    2015-01-01

    European foulbrood is a worldwide disease affecting the honey bee brood. Here, we report the draft genome sequence of Fructobacillus sp. EFB-N1, which was isolated from an infected honey bee larva derived from a Swiss European foulbrood outbreak. The genome consists of 68 contigs and harbors 1,629 predicted protein-encoding genes.

  2. Recombinant constructs of Borrelia burgdorferi

    Energy Technology Data Exchange (ETDEWEB)

    Dattwyler, Raymond J. (Setauket, NY); Gomes-Solecki, Maria J. C. (New York, NY); Luft, Benjamin J. (East Setauket, NY); Dunn, John J.(Bellport, NY)

    2007-02-20

    Novel chimeric nucleic acids, encoding chimeric Borrelia proteins comprising OspC or an antigenic fragment thereof and OspA or an antigenic fragment thereof, are disclosed. Chimeric proteins encoded by the nucleic acid sequences are also disclosed. The chimeric proteins are useful as vaccine immunogens against Lyme borreliosis, as well as for immunodiagnostic reagents.

  3. Application of a widely-used tropical anti-worm agent, mebendazole ...

    African Journals Online (AJOL)

    well-known anti-parasitic drug widely used in the tropical areas, inhibits cancer cell growth. Preclinical .... Bcl-2 and related proteins, encoded by the Bcl-2 oncogene ... homodimer that acts as an apoptosis inducer, while the ... Figure 1: Schematic presentation of proposed mebendazole anticancer mechanisms of action in.

  4. UBA domain containing proteins in fission yeast

    DEFF Research Database (Denmark)

    Hartmann-Petersen, Rasmus; Semple, Colin A M; Ponting, Chris P

    2003-01-01

    characterised on both the functional and structural levels. One example of a widespread ubiquitin binding module is the ubiquitin associated (UBA) domain. Here, we discuss the approximately 15 UBA domain containing proteins encoded in the relatively small genome of the fission yeast Schizosaccharomyces pombe...

  5. Genetic and molecular analyses of Escherichia coli N-acetylneuraminate lyase gene.

    OpenAIRE

    Kawakami, B; Kudo, T; Narahashi, Y; Horikoshi, K

    1986-01-01

    Two plasmids containing the N-acetylneuraminate lyase (NALase) gene (nanA) of Escherichia coli, pNL1 and pNL4, were constructed. Immunoprecipitation analysis indicated that the 35,000-dalton protein encoded in pNL4 was NALase. The synthesis of NALase in E. coli carrying these plasmids was constitutive.

  6. HIV-1 Nef control of cell signalling molecules: multiple strategies

    Indian Academy of Sciences (India)

    HIV-1 has at its disposal numerous proteins encoded by its genome which provide the required arsenal to establish and maintain infection in its host for a considerable number of years. One of the most important and enigmatic of these proteins is Nef. The Nef protein of HIV-1 plays a fundamental role in the virus life cycle.

  7. Cytoplasmic localization of a functionally active Fanconi anemia group A green fluorescent protein chimera in human 293 cells

    NARCIS (Netherlands)

    Kruyt, FAE; Waisfisz, Q; Dijkmans, LM; Hermsen, M.A.; Youssoufian, H; Arwert, F; Joenje, H

    1997-01-01

    Hypersensitivity to cross-linking agents and predisposition to malignancy are characteristic of the genetically heterogeneous inherited bone marrow failure syndrome, Fanconi anemia (FA). The protein encoded by the recently cloned FA complementation group A gene, FAA, has been expected to localize in

  8. Similarity and functional analyses of expressed parasitism genes in Heterodera schachtii and Heterodera glycines

    Science.gov (United States)

    The secreted proteins encoded by “parasitism genes” expressed within the esophageal glands cells of cyst nematodes play important roles in plant parasitism. Homologous transcripts and encoded proteins of the Heterodera glycines pioneer parasitism genes Hgsyv46, Hg4e02 and Hg5d08 were identified and ...

  9. Naturally occurred frame-shift mutations in the tvb receptor gene are responsible for decreased susceptibility to subgroups B, D, and E avian leukosis virus infection in chicken

    Science.gov (United States)

    The group of highly related avian leukosis viruses (ALVs) in chickens were thought to have evolved from a common retroviral ancestor into six subgroups, A to E and J. These ALV subgroups use diverse cellular proteins encoded by four genetic loci in chickens as receptors to gain entry into host cells...

  10. 78 FR 35290 - Government-Owned Inventions; Availability for Licensing

    Science.gov (United States)

    2013-06-12

    ... surface markers: programmed cell death protein 1 (PD-1; CD279), 4-1BB (CD137), T cell Ig- and mucin-domain... lack of suitable methods for screening hundreds, or even thousands, of cells expressing such proteins...) encodes a mammalian secreted protein comprising the same amino acid sequence. Synalleles are variants of a...

  11. The blessing effect of an extra copy of chromosome 21

    African Journals Online (AJOL)

    Solaf M. Elsayed

    2014-02-25

    Feb 25, 2014 ... mented in neuroblastoma: The S-100b protein (encoded by a chromosome ... the growth and induce death of human and murine neuroblas- toma cell lines [3 .... et al. A lack of neuroblastoma in Down syndrome: a study from.

  12. Research Resource: A Dual Proteomic Approach Identifies Regulated Islet Proteins During β-Cell Mass Expansion In Vivo

    DEFF Research Database (Denmark)

    Horn, Signe; Kirkegaard, Jeannette S.; Hoelper, Soraya

    2016-01-01

    to be up regulated as a response to pregnancy. These included several proteins, not previously associated with pregnancy-induced islet expansion, such as CLIC1, STMN1, MCM6, PPIB, NEDD4, and HLTF. Confirming the validity of our approach, we also identified proteins encoded by genes known to be associated...

  13. Draft Genome Sequence of Bacillus mycoides M2E15, a Strain Isolated from the Endosphere of Potato

    NARCIS (Netherlands)

    Yi, Yanglei; de Jong, Anne; Spoelder, Jan; Elzenga, J Theo M; van Elsas, Jan Dirk; Kuipers, Oscar P

    2016-01-01

    We present the draft genome sequence of Bacillus mycoides M2E15, a bacterium isolated from potato endosphere. Analysis of the 6.08-Mbp draft genome sequence identified 6,386 protein-encoding sequences, including potential plant growth promoting genes. Specifically, genes for proteins involved in

  14. A plasma membrane zinc transporter from ¤Medicago truncatula¤ is up-regulated in roots by Zn fertilization, yet down-regulated by arbuscular mycorrhizal colonization

    DEFF Research Database (Denmark)

    Burleigh, S.H.; Kristensen, B.K.; Bechmann, I.E.

    2003-01-01

    of yeast implying that the protein encoded by this gene can transport Zn across the yeast's plasma membrane. The product of a MtZIP2-GFP fusion construct introduced into onion cells by particle bombardment likewise localized to the plasma membrane. The MtZIP2 gene was expressed in roots and stems...

  15. Inefficient exogenous loading of a tapasin-dependent peptide onto HLA-B*44:02 can be improved by acid treatment or fixation of target cells

    NARCIS (Netherlands)

    Stroobant, Vincent; Demotte, Nathalie; Luiten, Rosalie M.; Leonhardt, Ralf M.; Cresswell, Peter; Bonehill, Aude; Michaux, Alexandre; Ma, Wenbin; Mulder, Arend; van den Eynde, Benoît J.; van der Bruggen, Pierre; Vigneron, Nathalie

    2012-01-01

    Antitumor cytolytic T lymphocytes (CTLs) recognize peptides derived from cellular proteins and presented on MHC class I. One category of peptides recognized by these CTLs is derived from proteins encoded by cancer-germline genes, which are specifically expressed in tumors, and therefore represent

  16. Presence of potential allergy-related linear epitopes in novel proteins from conventional crops and the implication for the safety assessment of these crops with respect to the current testing of genetically modified crops

    NARCIS (Netherlands)

    Kleter, G.A.; Peijnenburg, A.A.C.M.

    2003-01-01

    Mitochondria of cytoplasmic male sterile crop plants contain novel, chimeric open reading frames. In addition, a number of crops carry endogenous double-stranded ribonucleic acid (dsRNA). In this study, the novel proteins encoded by these genetic components were screened for the presence of

  17. fruitless alternative splicing and sex behaviour in insects

    Indian Academy of Sciences (India)

    In Drosophila melanogaster, male courtship requires proteins encoded by the fruitless (fru) gene that are produced in different sex-specific isoforms via alternative splicing. Drosophila mutant flies with loss-of-function alleles of the fru gene exhibit blocked male courtship behaviour. However, various individual steps in the ...

  18. Effect of Q211 and K222 PRNP polymorphic variants in the susceptibility of goats to oral infections with Goat Bovine Spongiform Encephalopathy

    NARCIS (Netherlands)

    Aguilar-Calvo, Patricia; Fast, C.; Tauscher, Kerstin; Espinosa, J.C.; Groschup, M.H.; Muhammad, Nadeem; Goldmann, W.; Langeveld, J.P.M.; Bossers, A.; Andreoletti, O.

    2015-01-01

    Background. The prion protein-encoding gene (PRNP) is one of the major determinants for scrapie occurrence in sheep and goats. However, its effect on bovine spongiform encephalopathy (BSE) transmission to goats is not clear.

    Methods. Goats harboring wild-type, R/Q211 or Q/K222 PRNP

  19. Brain expressed microRNAs implicated in schizophrenia etiology

    DEFF Research Database (Denmark)

    Hansen, Thomas; Olsen, Line; Lindow, Morten

    2007-01-01

    Protein encoding genes have long been the major targets for research in schizophrenia genetics. However, with the identification of regulatory microRNAs (miRNAs) as important in brain development and function, miRNAs genes have emerged as candidates for schizophrenia-associated genetic factors...

  20. Usher syndrome: molecular links of pathogenesis, proteins and pathways.

    NARCIS (Netherlands)

    Kremer, H.; Wijk, E. van; Marker, T.; Wolfrum, U.; Roepman, R.

    2006-01-01

    Usher syndrome is the most common form of deaf-blindness. The syndrome is both clinically and genetically heterogeneous, and to date, eight causative genes have been identified. The proteins encoded by these genes are part of a dynamic protein complex that is present in hair cells of the inner ear

  1. Definition of domain boundaries and crystallization of the SMN Tudor domain

    NARCIS (Netherlands)

    Sprangers, Remco; Selenko, Philipp; Sattler, Michael; Sinning, Irmgard; Groves, Matthew R

    Spinal muscular atropy (SMA) is the major genetic disease leading to childhood mortality and is caused by mutations in or deletions of the smn1 gene. The human survival of motor neurons (SMN) protein encoded by this gene plays an important role in the assembly of snRNPs (small nuclear

  2. Novel Single-Stranded DNA Virus Genomes Recovered from Chimpanzee Feces Sampled from the Mambilla Plateau in Nigeria

    Science.gov (United States)

    Walters, Matthew; Bawuro, Musa; Christopher, Alfred; Knight, Alexander; Kraberger, Simona; Stainton, Daisy; Chapman, Hazel

    2017-01-01

    ABSTRACT Metagenomic approaches are rapidly expanding our knowledge of the diversity of viruses. In the fecal matter of Nigerian chimpanzees we recovered three gokushovirus genomes, one circular replication-associated protein encoding single-stranded DNA virus (CRESS), and a CRESS DNA molecule. PMID:28254982

  3. A site-specific endonuclease encoded by a typical archaeal intron

    DEFF Research Database (Denmark)

    Dalgaard, Jacob; Garrett, Roger Antony; Belfort, Malene

    1993-01-01

    The protein encoded by the archaeal intron in the 23S rRNA gene of the hyperthermophile Desulfurococcus mobilis is a double-strand DNase that, like group I intron homing endonucleases, is capable of cleaving an intronless allele of the gene. This enzyme, I-Dmo I, is unusual among the intron...

  4. Identification and cloning of two insecticidal protein genes from ...

    African Journals Online (AJOL)

    Bacillus thuringiensis (Bt) is the most widely applied type of microbial pesticide due to its high specificity and environmental safety. The activity of Bt is largely attributed to the insecticidal crystal protein encoded by the cry genes. Different insecticidal crystal proteins of Bt have different bioactivity against distinct agricultural ...

  5. Sequence and annotation of the 314-kb MT325 and the 321-kb FR483 viruses that infect Chlorella Pbi.

    Science.gov (United States)

    Fitzgerald, Lisa A; Graves, Michael V; Li, Xiao; Feldblyum, Tamara; Hartigan, James; Van Etten, James L

    2007-02-20

    Viruses MT325 and FR483, members of the family Phycodnaviridae, genus Chlorovirus, infect the fresh water, unicellular, eukaryotic, chlorella-like green alga, Chlorella Pbi. The 314,335-bp genome of MT325 and the 321,240-bp genome of FR483 are the first viruses that infect Chlorella Pbi to have their genomes sequenced and annotated. Furthermore, these genomes are the two smallest chlorella virus genomes sequenced to date, MT325 has 331 putative protein-encoding and 10 tRNA-encoding genes and FR483 has 335 putative protein-encoding and 9 tRNA-encoding genes. The protein-encoding genes are almost evenly distributed on both strands, and intergenic space is minimal. Approximately 40% of the viral gene products resemble entries in public databases, including some that are the first of their kind to be detected in a virus. For example, these unique gene products include an aquaglyceroporin in MT325, a potassium ion transporter protein and an alkyl sulfatase in FR483, and a dTDP-glucose pyrophosphorylase in both viruses. Comparison of MT325 and FR483 protein-encoding genes with the prototype chlorella virus PBCV-1 indicates that approximately 82% of the genes are present in all three viruses.

  6. Were protein internal repeats formed by "bricolage"?

    Science.gov (United States)

    Lavorgna, G; Patthy, L; Boncinelli, E

    2001-03-01

    Is evolution an engineer, or is it a tinkerer--a "bricoleur"--building up complex molecules in organisms by increasing and adapting the materials at hand? An analysis of completely sequenced genomes suggests the latter, showing that increasing repetition of modules within the proteins encoded by these genomes is correlated with increasing complexity of the organism.

  7. Enhanced functional and structural domain assignments using

    Indian Academy of Sciences (India)

    Unknown

    using remote similarity detection procedures for proteins encoded in the genome of Mycobacterium tuberculosis H37Rv” (J. Biosci. 29 (3) 245–. 259, 2004) by Seema Namboori, Natasha Mhatre, Sentivel Sujatha,. Narayanaswamy Srinivasan and Shashi Bhushan Pandit. The three-dimensional structure and subcellular ...

  8. Reconstructing the phylogeny of Scolytinae and close allies: Major obstacles and prospects for a solution

    Science.gov (United States)

    Bjarte H. Jordal

    2007-01-01

    To enable the resolution of deep phylogenetic divergence in Scolytinae and closely related weevils, several new molecular markers were screened for their phylogenetic potential. The nuclear protein encoding genes, CAD and Arginine Kinase, were particularly promising and will be added to future phylogenetic studies in combination with 28S, COI, and Elongation Factor 1...

  9. Identification of the chitinase genes from the diamondback moth, Plutella xylostella.

    Science.gov (United States)

    Liao, Z H; Kuo, T C; Kao, C H; Chou, T M; Kao, Y H; Huang, R N

    2016-12-01

    Chitinases have an indispensable function in chitin metabolism and are well characterized in numerous insect species. Although the diamondback moth (DBM) Plutella xylostella, which has a high reproductive potential, short generation time, and characteristic adaptation to adverse environments, has become one of the most serious pests of cruciferous plants worldwide, the information on the chitinases of the moth is presently limited. In the present study, using degenerated polymerase chain reaction (PCR) and rapid amplification of cDNA ends-PCR strategies, four chitinase genes of P. xylostella were cloned, and an exhaustive search was conducted for chitinase-like sequences from the P. xylostella genome and transcriptomic database. Based on the domain analysis of the deduced amino acid sequences and the phylogenetic analysis of the catalytic domain sequences, we identified 15 chitinase genes from P. xylostella. Two of the gut-specific chitinases did not cluster with any of the known phylogenetic groups of chitinases and might be in a new group of the chitinase family. Moreover, in our study, group VIII chitinase was not identified. The structures, classifications and expression patterns of the chitinases of P. xylostella were further delineated, and with this information, further investigations on the functions of chitinase genes in DBM could be facilitated.

  10. Chitinase 3-like 1 Regulates Cellular and Tissue Responses via IL-13 Receptor α2

    Directory of Open Access Journals (Sweden)

    Chuan Hua He

    2013-08-01

    Full Text Available Members of the 18 glycosyl hydrolase (GH 18 gene family have been conserved over species and time and are dysregulated in inflammatory, infectious, remodeling, and neoplastic disorders. This is particularly striking for the prototypic chitinase-like protein chitinase 3-like 1 (Chi3l1, which plays a critical role in antipathogen responses where it augments bacterial killing while stimulating disease tolerance by controlling cell death, inflammation, and remodeling. However, receptors that mediate the effects of GH 18 moieties have not been defined. Here, we demonstrate that Chi3l1 binds to interleukin-13 receptor α2 (IL-13Rα2 and that Chi3l1, IL-13Rα2, and IL-13 are in a multimeric complex. We also demonstrate that Chi3l1 activates macrophage mitogen-activated protein kinase, protein kinase B/AKT, and Wnt/β-catenin signaling and regulates oxidant injury, apoptosis, pyroptosis, inflammasome activation, antibacterial responses, melanoma metastasis, and TGF-β1 production via IL-13Rα2-dependent mechanisms. Thus, IL-13Rα2 is a GH 18 receptor that plays a critical role in Chi3l1 effector responses.

  11. Cloning of the Bacillus thuringiensis serovar sotto chitinase (Schi gene and characterization of its protein

    Directory of Open Access Journals (Sweden)

    Wan-Fang Zhong

    2005-12-01

    Full Text Available Chitinase plays a positive role in the pathogenicity of Bacillus thuringiensis to insect pests. We used touchdown PCR to clone the chitinase (Schi gene from Bacillus thuringiensis serovar sotto (Bt sotto chromosomal DNA. Our DNA sequencing analysis revealed that the Bt sotto Schi gene consists of an open reading frame (ORF of 2067 nucleotides with codes for the chitinase precursor. We also found that the putative promoter consensus sequences (the -35 and -10 regions of the Bt soto Schi gene are identical to those of the chiA71 gene from Bt Pakistani, the chiA74 gene from Bt kenyae and the ichi gene from Bt israelensis. The Schi chitinase precursor is 688 amino acids long with an estimated molecular mass of 75.75 kDa and a theoretical isoelectric point of 5.74, and contains four domains, which are, in sequence, a signal peptide, an N-terminal catalytic domain, a fibronectin type III like domain and a C-terminal chitin-binding domain. Sequence comparison and the evolutionary relationship of the Bt sotto Schi chitinase to other chitinase and chitinase-like proteins are also discussed.

  12. Multifunctional Receptor Stabilin-1 in Homeostasis and Disease

    Directory of Open Access Journals (Sweden)

    Julia Kzhyshkowska

    2010-01-01

    Full Text Available The multifunctional scavenger receptor stabilin-1 (STAB1, FEEL-1, CLEVER-1, KIAA0246 is expressed on tissue macrophages and sinusoidal endothelial cells in healthy organisms, and its expression on both macrophages and different subtypes of endothelial cells is induced during chronic inflammation and tumor progression. Stabilin-1 is a type-1 transmembrane receptor that mediates endocytic and phagocytic clearance of “unwanted-self” components, intracellular sorting of the endogenously synthesized chitinase-like protein SI-CLP, and transcytosis of the growth hormone family member placental lactogen. The central sorting station for stabilin-1 trafficking seems to be the trans-Golgi network (TGN. Transport of stabilin-1 in the TGN requires interaction with GGA adaptors that bind to the classical DDSLL motif and a novel acidic cluster in its cytoplasmic tail. Degradation of stabilin-1 seems to depend on the interaction with sorting nexin 17. However, the mechanisms keeping stabilin-1 on the cell surface remain to be identified. This issue deserves specific attention due to the growing amount of data indicating that function of stabilin-1 in cell adhesion events is essential for inflammation and metastasis. Taking into consideration the complexity of stabilin-1—mediated processes, investigation of stabilin-1 functions in the animal models, as well as mathematic modeling of intracellular trafficking and extracellular contact, would enable prediction of stabilin-1 behavior in complex biological systems and would open perspectives for therapeutic targeting of stabilin-1 pathways in chronic inflammation and carcinogenesis.

  13. Sex determination in insects: a binary decision based on alternative splicing.

    Science.gov (United States)

    Salz, Helen K

    2011-08-01

    The gene regulatory networks that control sex determination vary between species. Despite these differences, comparative studies in insects have found that alternative splicing is reiteratively used in evolution to control expression of the key sex-determining genes. Sex determination is best understood in Drosophila where activation of the RNA binding protein-encoding gene Sex-lethal is the central female-determining event. Sex-lethal serves as a genetic switch because once activated it controls its own expression by a positive feedback splicing mechanism. Sex fate choice in is also maintained by self-sustaining positive feedback splicing mechanisms in other dipteran and hymenopteran insects, although different RNA binding protein-encoding genes function as the binary switch. Studies exploring the mechanisms of sex-specific splicing have revealed the extent to which sex determination is integrated with other developmental regulatory networks. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. Disruption and analysis of the clpB, clpC, and clpE genes in Lactococcus lactis: ClpE, a new Clp family in gram-positive bacteria

    DEFF Research Database (Denmark)

    Ingmer, Hanne; Vogensen, Finn K.; Hammer, Karin

    1999-01-01

    In the genome of the gram-positive bacterium Lactococcus lactis MG1363, we have identified three genes (clpC, clpE, and clpB) which encode Clp proteins containing two conserved ATP binding domains. The proteins encoded by two of the genes belong to the previously described ClpB and ClpC families....... The clpE gene, however, encodes a member of a new Clp protein family that is characterized by a short N-terminal domain including a putative zinc binding domain (-CX2CX22CX2C-). Expression of the 83-kDa ClpE protein as well as of the two proteins encoded by clpB was strongly induced by heat shock and...... was shown to participate in the degradation of randomly folded proteins in L. lactis, could be necessary for degrading proteins generated by certain types of stress....

  15. Molecular characterization and expression of the M6 gene of grass carp hemorrhage virus (GCHV), an aquareovirus.

    Science.gov (United States)

    Qiu, T; Lu, R H; Zhang, J; Zhu, Z Y

    2001-07-01

    The complete nucleotide sequence of M6 gene of grass carp hemorrhage virus (GCHV) was determined. It is 2039 nucleotides in length and contains a single large open reading frame that could encode a protein of 648 amino acids with predicted molecular mass of 68.7 kDa. Amino acid sequence comparison revealed that the protein encoded by GCHV M6 is closely related to the protein mu1 of mammalian reovirus. The M6 gene, encoding the major outer-capsid protein, was expressed using the pET fusion protein vector in Escherichia coli and detected by Western blotting using chicken anti-GCHV immunoglobulin (IgY). The result indicates that the protein encoded by M6 may share a putative Asn-42-Pro-43 proteolytic cleavage site with mu1.

  16. Unexplored therapeutic opportunities in the human genome

    DEFF Research Database (Denmark)

    Oprea, Tudor I; Bologa, Cristian G; Brunak, Søren

    2018-01-01

    A large proportion of biomedical research and the development of therapeutics is focused on a small fraction of the human genome. In a strategic effort to map the knowledge gaps around proteins encoded by the human genome and to promote the exploration of currently understudied, but potentially d...... as well as key drug target classes, including G protein-coupled receptors, protein kinases and ion channels, which illustrate the nature of the unexplored opportunities for biomedical research and therapeutic development....

  17. Sex Determination in Insects: a binary decision based on alternative splicing

    OpenAIRE

    Salz, Helen K.

    2011-01-01

    The gene regulatory networks that control sex determination vary between species. Despite these differences, comparative studies in insects have found that alternative splicing is reiteratively used in evolution to control expression of the key sex determining genes. Sex determination is best understood in Drosophila where activation of the RNA binding protein encoding gene Sex-lethal is the central female-determining event. Sex-lethal serves as a genetic switch because once activated it cont...

  18. Target Site Recognition by a Diversity-Generating Retroelement

    OpenAIRE

    Guo, Huatao; Tse, Longping V.; Nieh, Angela W.; Czornyj, Elizabeth; Williams, Steven; Oukil, Sabrina; Liu, Vincent B.; Miller, Jeff F.

    2011-01-01

    Diversity-generating retroelements (DGRs) are in vivo sequence diversification machines that are widely distributed in bacterial, phage, and plasmid genomes. They function to introduce vast amounts of targeted diversity into protein-encoding DNA sequences via mutagenic homing. Adenine residues are converted to random nucleotides in a retrotransposition process from a donor template repeat (TR) to a recipient variable repeat (VR). Using the Bordetella bacteriophage BPP-1 element as a prototype...

  19. Draft Genome Sequence of Ezakiella peruensis Strain M6.X2, a Human Gut Gram-Positive Anaerobic Coccus.

    Science.gov (United States)

    Diop, Awa; Diop, Khoudia; Tomei, Enora; Raoult, Didier; Fenollar, Florence; Fournier, Pierre-Edouard

    2018-03-01

    We report here the draft genome sequence of Ezakiella peruensis strain M6.X2 T The draft genome is 1,672,788 bp long and harbors 1,589 predicted protein-encoding genes, including 26 antibiotic resistance genes with 1 gene encoding vancomycin resistance. The genome also exhibits 1 clustered regularly interspaced short palindromic repeat region and 333 genes acquired by horizontal gene transfer. Copyright © 2018 Diop et al.

  20. Phylogenetic reconstruction of South American felids defined by protein electrophoresis

    OpenAIRE

    Pecon Slattery, J.; Johnson, W. E.; Goldman, D.; O'Brien, S. J.

    1994-01-01

    Phylogenetic associations among six closely related South American felid species were defined by changes in protein-encoding gene loci. We analyzed proteins isolated from skin fibroblasts using two-dimensional electrophoresis and allozymes extracted from blood cells. Genotypes were determined for multiple individuals of ocelot, margay, tigrina, Geoffroy's cat, kodkod, and pampas cat at 548 loci resolved by two-dimensional electrophoresis and 44 allozyme loci. Phenograms were constructed using...

  1. Fanconi anemia (cross)linked to DNA repair.

    Science.gov (United States)

    Niedernhofer, Laura J; Lalai, Astrid S; Hoeijmakers, Jan H J

    2005-12-29

    Fanconi anemia is characterized by hypersensitivity to DNA interstrand crosslinks (ICLs) and susceptibility to tumor formation. Despite the identification of numerous Fanconi anemia (FANC) genes, the mechanism by which proteins encoded by these genes protect a cell from DNA interstrand crosslinks remains unclear. The recent discovery of two DNA helicases that, when defective, cause Fanconi anemia tips the balance in favor of the direct involvement of the FANC proteins in DNA repair and the bypass of DNA lesions.

  2. Evolutionary genomics of archaeal viruses: unique viral genomes in the third domain of life

    DEFF Research Database (Denmark)

    Prangishvili, D.; Garrett, R. A.; Koonin, E.

    2006-01-01

    In terms of virion morphology, the known viruses of archaea fall into two distinct classes: viruses of mesophilic and moderately thermophilic Eueryarchaeota closely resemble head-and-tail bacteriophages whereas viruses of hyperthermophilic Crenarchaeota show a variety of unique morphotypes...... of bacteriophages. The proteins encoded by the genes belonging to this pool include predicted transcription regulators, ATPases implicated in viral DNA replication and packaging, enzymes of DNA precursor metabolism, RNA modification enzymes, and glycosylases. In addition, each of the crenarchaeal viruses encodes...

  3. Comprehensive Evaluation of Streptococcus sanguinis Cell Wall-Anchored Proteins in Early Infective Endocarditis▿ †

    OpenAIRE

    Turner, Lauren Senty; Kanamoto, Taisei; Unoki, Takeshi; Munro, Cindy L.; Wu, Hui; Kitten, Todd

    2009-01-01

    Streptococcus sanguinis is a member of the viridans group of streptococci and a leading cause of the life-threatening endovascular disease infective endocarditis. Initial contact with the cardiac infection site is likely mediated by S. sanguinis surface proteins. In an attempt to identify the proteins required for this crucial step in pathogenesis, we searched for surface-exposed, cell wall-anchored proteins encoded by S. sanguinis and then used a targeted signature-tagged mutagenesis (STM) a...

  4. Serum immune response to Shigella protein antigens in rhesus monkeys and humans infected with Shigella spp.

    OpenAIRE

    Oaks, E V; Hale, T L; Formal, S B

    1986-01-01

    The serum antibody response to proteins encoded by the virulence-associated plasmid of Shigella flexneri was determined in monkeys challenged with virulent S. flexneri serotype 2a. With water-extractable antigen in an enzyme-linked immunosorbent assay, a significant increase in antibody titer against proteins from a plasmid-carrying, virulent strain of S. flexneri serotype 5 could be demonstrated in convalescent sera. There were minimal antibody titers against proteins from an avirulent (plas...

  5. The E5 Proteins

    OpenAIRE

    DiMaio, Daniel; Petti, Lisa

    2013-01-01

    The E5 proteins are short transmembrane proteins encoded by many animal and human papillomaviruses. These proteins display transforming activity in cultured cells and animals, and they presumably also play a role in the productive virus life cycle. The E5 proteins are thought to act by modulating the activity of cellular proteins. Here, we describe the biological activities of the best-studied E5 proteins and discuss the evidence implicating specific protein targets and pathways in mediating ...

  6. Usher syndrome: Hearing loss, retinal degeneration and associated abnormalities

    OpenAIRE

    Mathur, Pranav; Yang, Jun

    2015-01-01

    Usher syndrome (USH), clinically and genetically heterogeneous, is the leading genetic cause of combined hearing and vision loss. USH is classified into three types, based on the hearing and vestibular symptoms observed in patients. Sixteen loci have been reported to be involved in the occurrence of USH and atypical USH. Among them, twelve have been identified as causative genes and one as a modifier gene. Studies on the proteins encoded by these USH genes suggest that USH proteins interact a...

  7. Adaptive evolution of mitochondrial energy metabolism genes associated with increased energy demand in flying insects.

    Science.gov (United States)

    Yang, Yunxia; Xu, Shixia; Xu, Junxiao; Guo, Yan; Yang, Guang

    2014-01-01

    Insects are unique among invertebrates for their ability to fly, which raises intriguing questions about how energy metabolism in insects evolved and changed along with flight. Although physiological studies indicated that energy consumption differs between flying and non-flying insects, the evolution of molecular energy metabolism mechanisms in insects remains largely unexplored. Considering that about 95% of adenosine triphosphate (ATP) is supplied by mitochondria via oxidative phosphorylation, we examined 13 mitochondrial protein-encoding genes to test whether adaptive evolution of energy metabolism-related genes occurred in insects. The analyses demonstrated that mitochondrial DNA protein-encoding genes are subject to positive selection from the last common ancestor of Pterygota, which evolved primitive flight ability. Positive selection was also found in insects with flight ability, whereas no significant sign of selection was found in flightless insects where the wings had degenerated. In addition, significant positive selection was also identified in the last common ancestor of Neoptera, which changed its flight mode from direct to indirect. Interestingly, detection of more positively selected genes in indirect flight rather than direct flight insects suggested a stronger selective pressure in insects having higher energy consumption. In conclusion, mitochondrial protein-encoding genes involved in energy metabolism were targets of adaptive evolution in response to increased energy demands that arose during the evolution of flight ability in insects.

  8. Identification of New Single Nucleotide Polymorphism-Based Markers for Inter- and Intraspecies Discrimination of Obligate Bacterial Parasites (Pasteuria spp.) of Invertebrates ▿ †

    Science.gov (United States)

    Mauchline, Tim H.; Knox, Rachel; Mohan, Sharad; Powers, Stephen J.; Kerry, Brian R.; Davies, Keith G.; Hirsch, Penny R.

    2011-01-01

    Protein-encoding and 16S rRNA genes of Pasteuria penetrans populations from a wide range of geographic locations were examined. Most interpopulation single nucleotide polymorphisms (SNPs) were detected in the 16S rRNA gene. However, in order to fully resolve all populations, these were supplemented with SNPs from protein-encoding genes in a multilocus SNP typing approach. Examination of individual 16S rRNA gene sequences revealed the occurrence of “cryptic” SNPs which were not present in the consensus sequences of any P. penetrans population. Additionally, hierarchical cluster analysis separated P. penetrans 16S rRNA gene clones into four groups, and one of which contained sequences from the most highly passaged population, demonstrating that it is possible to manipulate the population structure of this fastidious bacterium. The other groups were made from representatives of the other populations in various proportions. Comparison of sequences among three Pasteuria species, namely, P. penetrans, P. hartismeri, and P. ramosa, showed that the protein-encoding genes provided greater discrimination than the 16S rRNA gene. From these findings, we have developed a toolbox for the discrimination of Pasteuria at both the inter- and intraspecies levels. We also provide a model to monitor genetic variation in other obligate hyperparasites and difficult-to-culture microorganisms. PMID:21803895

  9. Identification of new single nucleotide polymorphism-based markers for inter- and intraspecies discrimination of obligate bacterial parasites (Pasteuria spp.) of invertebrates.

    Science.gov (United States)

    Mauchline, Tim H; Knox, Rachel; Mohan, Sharad; Powers, Stephen J; Kerry, Brian R; Davies, Keith G; Hirsch, Penny R

    2011-09-01

    Protein-encoding and 16S rRNA genes of Pasteuria penetrans populations from a wide range of geographic locations were examined. Most interpopulation single nucleotide polymorphisms (SNPs) were detected in the 16S rRNA gene. However, in order to fully resolve all populations, these were supplemented with SNPs from protein-encoding genes in a multilocus SNP typing approach. Examination of individual 16S rRNA gene sequences revealed the occurrence of "cryptic" SNPs which were not present in the consensus sequences of any P. penetrans population. Additionally, hierarchical cluster analysis separated P. penetrans 16S rRNA gene clones into four groups, and one of which contained sequences from the most highly passaged population, demonstrating that it is possible to manipulate the population structure of this fastidious bacterium. The other groups were made from representatives of the other populations in various proportions. Comparison of sequences among three Pasteuria species, namely, P. penetrans, P. hartismeri, and P. ramosa, showed that the protein-encoding genes provided greater discrimination than the 16S rRNA gene. From these findings, we have developed a toolbox for the discrimination of Pasteuria at both the inter- and intraspecies levels. We also provide a model to monitor genetic variation in other obligate hyperparasites and difficult-to-culture microorganisms.

  10. Glycoprotein YKL-40: A potential biomarker of disease activity in rheumatoid arthritis during intensive treatment with csDMARDs and infliximab. Evidence from the randomised controlled NEO-RACo trial.

    Directory of Open Access Journals (Sweden)

    Tuija Väänänen

    Full Text Available YKL-40, a chitinase-like glycoprotein associated with inflammation and tissue remodeling, is produced by joint tissues and recognized as a candidate auto-antigen in rheumatoid arthritis (RA. In the present study, we investigated YKL-40 as a potential biomarker of disease activity in patients with early RA at baseline and during intensive treatment aiming for early remission.Ninety-nine patients with early DMARD-naïve RA participated in the NEO-RACo study. For the first four weeks, the patients were treated with the combination of sulphasalazine, methotrexate, hydroxychloroquine and low dose prednisolone (FIN-RACo DMARD combination, and subsequently randomized to receive placebo or infliximab added on the treatment for further 22 weeks. Disease activity was evaluated using the 28-joint disease activity score and plasma YKL-40 concentrations were measured by immunoassay.At the baseline, plasma YKL-40 concentration was 57 ± 37 (mean ± SD ng/ml. YKL-40 was significantly associated with the disease activity score, interleukin-6 and erythrocyte sedimentation rate both at the baseline and during the 26 weeks' treatment. The csDMARD combination decreased YKL-40 levels already during the first four weeks of treatment, and there was no further reduction when the tumour necrosis factor-α antagonist infliximab was added on the combination treatment.High YKL-40 levels were found to be associated with disease activity in early DMARD-naïve RA and during intensive treat-to-target therapy. The present results suggest YKL-40 as a useful biomarker of disease activity in RA to be used to steer treatment towards remission.

  11. Ozone-Induced Nasal Type 2 Immunity in Mice Is Dependent on Innate Lymphoid Cells.

    Science.gov (United States)

    Kumagai, Kazuyoshi; Lewandowski, Ryan; Jackson-Humbles, Daven N; Li, Ning; Van Dyken, Steven J; Wagner, James G; Harkema, Jack R

    2016-06-01

    Epidemiological studies suggest that elevated ambient concentrations of ozone are associated with activation of eosinophils in the nasal airways of atopic and nonatopic children. Mice repeatedly exposed to ozone develop eosinophilic rhinitis and type 2 immune responses. In this study, we determined the role of innate lymphoid cells (ILCs) in the pathogenesis of ozone-induced eosinophilic rhinitis by using lymphoid-sufficient C57BL/6 mice, Rag2(-/-) mice that are devoid of T cells and B cells, and Rag2(-/-)Il2rg(-/-) mice that are depleted of all lymphoid cells including ILCs. The animals were exposed to 0 or 0.8 ppm ozone for 9 consecutive weekdays (4 h/d). Mice were killed 24 hours after exposure, and nasal tissues were selected for histopathology and gene expression analysis. ILC-sufficient C57BL/6 and Rag2(-/-) mice exposed to ozone developed marked eosinophilic rhinitis and epithelial remodeling (e.g., epithelial hyperplasia and mucous cell metaplasia). Chitinase-like proteins and alarmins (IL-33, IL-25, and thymic stromal lymphopoietin) were also increased morphometrically in the nasal epithelium of ozone-exposed C57BL/6 and Rag2(-/-) mice. Ozone exposure elicited increased expression of Il4, Il5, Il13, St2, eotaxin, MCP-2, Gob5, Arg1, Fizz1, and Ym2 mRNA in C57BL/6 and Rag2(-/-) mice. In contrast, ozone-exposed ILC-deficient Rag2(-/-)Il2rg(-/-) mice had no nasal lesions or overexpression of Th2- or ILC2-related transcripts. These results indicate that ozone-induced eosinophilic rhinitis, nasal epithelial remodeling, and type 2 immune activation are dependent on ILCs. To the best of our knowledge, this is the first study to demonstrate that ILCs play an important role in the nasal pathology induced by repeated ozone exposure.

  12. Glycoprotein YKL-40 Levels in Plasma Are Associated with Fibrotic Changes on HRCT in Asbestos-Exposed Subjects

    Directory of Open Access Journals (Sweden)

    Tuija Väänänen

    2017-01-01

    Full Text Available YKL-40 is a chitinase-like glycoprotein produced by alternatively activated macrophages that are associated with wound healing and fibrosis. Asbestosis is a chronic asbestos-induced lung disease, in which injury of epithelial cells and activation of alveolar macrophages lead to enhanced collagen production and fibrosis. We studied if YKL-40 is related to inflammation, fibrosis, and/or lung function in subjects exposed to asbestosis. Venous blood samples were collected from 85 men with moderate or heavy occupational asbestos exposure and from 28 healthy, age-matched controls. Levels of plasma YKL-40, CRP, IL-6, adipsin, and MMP-9 were measured with enzyme-linked immunosorbent assay (ELISA. Plasma YKL-40 levels were significantly higher in subjects with asbestosis (n=19 than in those with no fibrotic findings in HRCT following asbestos exposure (n=66 or in unexposed healthy controls. In asbestos-exposed subjects, plasma YKL-40 correlated negatively with lung function capacity parameters FVC (Pearson’s r −0.259, p=0.018 and FEV1 (Pearson’s r −0.240, p=0.028 and positively with CRP (Spearman’s rho 0.371, p<0.001, IL-6 (Spearman’s rho 0.314, p=0.003, adipsin (Spearman’s rho 0.459, p<0.001, and MMP-9 (Spearman’s rho 0.243, p=0.025. The present finding suggests YKL-40 as a biomarker associated with fibrosis and inflammation in asbestos-exposed subjects.

  13. YKL-40 tissue expression and plasma levels in patients with ovarian cancer

    International Nuclear Information System (INIS)

    Høgdall, Estrid VS; Christensen, Lise H; Ringsholt, Merete; Høgdall, Claus K; Christensen, Ib Jarle; Johansen, Julia S; Kjaer, Susanne K; Blaakaer, Jan; Ostenfeld-Møller, Lene; Price, Paul A

    2009-01-01

    YKL-40 (chitinase-3-like-1) is a member of 'mammalian chitinase-like proteins'. The protein is expressed in many types of cancer cells and the highest plasma YKL-40 levels have been found in patients with metastatic disease, short recurrence/progression-free intervals, and short overall survival. The aim of the study was to determine the expression of YKL-40 in tumor tissue and plasma in patients with borderline ovarian tumor or epithelial ovarian cancer (OC), and investigate prognostic value of this marker. YKL-40 protein expression was determined by immunohistochemistry in tissue arrays from 181 borderline tumors and 473 OC. Plasma YKL-40 was determined by ELISA in preoperative samples from 19 patients with borderline tumor and 76 OC patients. YKL-40 protein expression was found in cancer cells, tumor associated macrophages, neutrophils and mast cells. The tumor cell expression was higher in OC than in borderline tumors (p = 0.001), and associated with FIGO stage (p < 0.0001) and histological subtype (p = 0.0009). Positive YKL-40 expression (≥ 5% staining) was not associated with reduced survival. Plasma YKL-40 was also higher in patients with OC than in patients with borderline tumors (p < 0.0001), and it was positively correlated to serum CA-125 (p < 0.0001) and FIGO stage (p = 0.0001). Univariate Cox analysis of plasma YKL-40 showed association with overall survival (p < 0.0001). Multivariate Cox analysis, including plasma YKL-40, serum CA125, FIGO stage, age and radicality after primary surgery as variables, showed that elevated plasma YKL-40 was associated with a shorter survival (HR = 2.13, 95% CI: 1.40–3.25, p = 0.0004). YKL-40 in OC tissue and plasma are related to stage and histology, but only plasma YKL-40 is a prognostic biomarker in patients with OC

  14. Characterization of the Carbohydrate Binding Module 18 gene family in the amphibian pathogen Batrachochytrium dendrobatidis.

    Science.gov (United States)

    Liu, Peng; Stajich, Jason E

    2015-04-01

    Batrachochytrium dendrobatidis (Bd) is the causative agent of chytridiomycosis responsible for worldwide decline in amphibian populations. Previous analysis of the Bd genome revealed a unique expansion of the carbohydrate-binding module family 18 (CBM18) predicted to be a sub-class of chitin recognition domains. CBM expansions have been linked to the evolution of pathogenicity in a variety of fungal species by protecting the fungus from the host. Based on phylogenetic analysis and presence of additional protein domains, the gene family can be classified into 3 classes: Tyrosinase-, Deacetylase-, and Lectin-like. Examination of the mRNA expression levels from sporangia and zoospores of nine of the cbm18 genes found that the Lectin-like genes had the highest expression while the Tyrosinase-like genes showed little expression, especially in zoospores. Heterologous expression of GFP-tagged copies of four CBM18 genes in Saccharomyces cerevisiae demonstrated that two copies containing secretion signal peptides are trafficked to the cell boundary. The Lectin-like genes cbm18-ll1 and cbm18-ll2 co-localized with the chitinous cell boundaries visualized by staining with calcofluor white. In vitro assays of the full length and single domain copies from CBM18-LL1 demonstrated chitin binding and no binding to cellulose or xylan. Expressed CBM18 domain proteins were demonstrated to protect the fungus, Trichoderma reeseii, in vitro against hydrolysis from exogenously added chitinase, likely by binding and limiting exposure of fungal chitin. These results demonstrate that cbm18 genes can play a role in fungal defense and expansion of their copy number may be an important pathogenicity factor of this emerging infectious disease of amphibians. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. The Spanish biology/disease initiative within the human proteome project: Application to rheumatic diseases.

    Science.gov (United States)

    Ruiz-Romero, Cristina; Calamia, Valentina; Albar, Juan Pablo; Casal, José Ignacio; Corrales, Fernando J; Fernández-Puente, Patricia; Gil, Concha; Mateos, Jesús; Vivanco, Fernando; Blanco, Francisco J

    2015-09-08

    The Spanish Chromosome 16 consortium is integrated in the global initiative Human Proteome Project, which aims to develop an entire map of the proteins encoded following a gene-centric strategy (C-HPP) in order to make progress in the understanding of human biology in health and disease (B/D-HPP). Chromosome 16 contains many genes encoding proteins involved in the development of a broad range of diseases, which have a significant impact on the health care system. The Spanish HPP consortium has developed a B/D platform with five programs focused on selected medical areas: cancer, obesity, cardiovascular, infectious and rheumatic diseases. Each of these areas has a clinical leader associated to a proteomic investigator with the responsibility to get a comprehensive understanding of the proteins encoded by Chromosome 16 genes. Proteomics strategies have enabled great advances in the area of rheumatic diseases, particularly in osteoarthritis, with studies performed on joint cells, tissues and fluids. In this manuscript we describe how the Spanish HPP-16 consortium has developed a B/D platform with five programs focused on selected medical areas: cancer, obesity, cardiovascular, infectious and rheumatic diseases. Each of these areas has a clinical leader associated to a proteomic investigator with the responsibility to get a comprehensive understanding of the proteins encoded by Chromosome 16 genes. We show how the Proteomic strategy has enabled great advances in the area of rheumatic diseases, particularly in osteoarthritis, with studies performed on joint cells, tissues and fluids. This article is part of a Special Issue entitled: HUPO 2014. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Multilocus sequence analysis of Treponema denticola strains of diverse origin

    Directory of Open Access Journals (Sweden)

    Mo Sisu

    2013-02-01

    Full Text Available Abstract Background The oral spirochete bacterium Treponema denticola is associated with both the incidence and severity of periodontal disease. Although the biological or phenotypic properties of a significant number of T. denticola isolates have been reported in the literature, their genetic diversity or phylogeny has never been systematically investigated. Here, we describe a multilocus sequence analysis (MLSA of 20 of the most highly studied reference strains and clinical isolates of T. denticola; which were originally isolated from subgingival plaque samples taken from subjects from China, Japan, the Netherlands, Canada and the USA. Results The sequences of the 16S ribosomal RNA gene, and 7 conserved protein-encoding genes (flaA, recA, pyrH, ppnK, dnaN, era and radC were successfully determined for each strain. Sequence data was analyzed using a variety of bioinformatic and phylogenetic software tools. We found no evidence of positive selection or DNA recombination within the protein-encoding genes, where levels of intraspecific sequence polymorphism varied from 18.8% (flaA to 8.9% (dnaN. Phylogenetic analysis of the concatenated protein-encoding gene sequence data (ca. 6,513 nucleotides for each strain using Bayesian and maximum likelihood approaches indicated that the T. denticola strains were monophyletic, and formed 6 well-defined clades. All analyzed T. denticola strains appeared to have a genetic origin distinct from that of ‘Treponema vincentii’ or Treponema pallidum. No specific geographical relationships could be established; but several strains isolated from different continents appear to be closely related at the genetic level. Conclusions Our analyses indicate that previous biological and biophysical investigations have predominantly focused on a subset of T. denticola strains with a relatively narrow range of genetic diversity. Our methodology and results establish a genetic framework for the discrimination and phylogenetic

  17. Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

    OpenAIRE

    In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A

    1997-01-01

    Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, delet...

  18. Somatic mutations in the transcriptional corepressor gene BCORL1 in adult acute myelogenous leukemia

    OpenAIRE

    Li, Meng; Collins, Roxane; Jiao, Yuchen; Ouillette, Peter; Bixby, Dale; Erba, Harry; Vogelstein, Bert; Kinzler, Kenneth W.; Papadopoulos, Nickolas; Malek, Sami N.

    2011-01-01

    To further our understanding of the genetic basis of acute myelogenous leukemia (AML), we determined the coding exon sequences of ∼ 18 000 protein-encoding genes in 8 patients with secondary AML. Here we report the discovery of novel somatic mutations in the transcriptional corepressor gene BCORL1 that is located on the X-chromosome. Analysis of BCORL1 in an unselected cohort of 173 AML patients identified a total of 10 mutated cases (6%) with BCORL1 mutations, whereas analysis of 19 AML cell...

  19. Molecular cloning of a Candida albicans gene (SSB1) coding for a protein related to the Hsp70 family.

    Science.gov (United States)

    Maneu, V; Cervera, A M; Martinez, J P; Gozalbo, D

    1997-06-15

    We have cloned and sequenced a Candida albicans gene (SSB1) encoding a potential member of the heat-shock protein seventy (hsp70) family. The protein encoded by this gene contains 613 amino acids and shows a high degree (85%) of sequence identity to the ssb subfamily (ssb1 and ssb2) of the Saccharomyces cerevisiae hsp70 family. The transcribed mRNA (2.1 kb) is present in similar amounts both in yeast and germ tube cells of C. albicans.

  20. Gene therapy and its implications in Periodontics

    Science.gov (United States)

    Mahale, Swapna; Dani, Nitin; Ansari, Shumaila S.; Kale, Triveni

    2009-01-01

    Gene therapy is a field of Biomedicine. With the advent of gene therapy in dentistry, significant progress has been made in the control of periodontal diseases and reconstruction of dento-alveolar apparatus. Implementation in periodontics include: -As a mode of tissue engineering with three approaches: cell, protein-based and gene delivery approach. -Genetic approach to Biofilm Antibiotic Resistance. Future strategies of gene therapy in preventing periodontal diseases: -Enhances host defense mechanism against infection by transfecting host cells with an antimicrobial peptide protein-encoding gene. -Periodontal vaccination. Gene therapy is one of the recent entrants and its applications in the field of periodontics are reviewed in general here. PMID:20376232

  1. The Search for an Effective Therapy to Treat Fragile X Syndrome: Dream or Reality?

    OpenAIRE

    Castagnola, Sara; Bardoni, Barbara; Maurin, Thomas

    2017-01-01

    Fragile X Syndrome (FXS) is the most common form of intellectual disability and a primary cause of autism. It originates from the lack of the Fragile X Mental Retardation Protein (FMRP), which is an RNA-binding protein encoded by the Fragile X Mental Retardation Gene 1 (FMR1) gene. Multiple roles have been attributed to this protein, ranging from RNA transport (from the nucleus to the cytoplasm, but also along neurites) to translational control of mRNAs. Over the last 20 years many studies ha...

  2. The p21 ras C-terminus is required for transformation and membrane association

    DEFF Research Database (Denmark)

    Willumsen, B M; Christensen, A; Hubbert, N L

    1984-01-01

    The Harvey murine sarcoma virus (Ha-MuSV) transforming gene, v-rasH, encodes a 21,000 molecular weight protein (p21) that is closely related to the p21 proteins encoded by the cellular transforming genes of the ras gene family. The primary translation product (prop21), which is found in the cytosol...... of these biochemical features of the protein, we have now studied a series of deletion mutants located at or near the C-terminus of the viral p21 protein. Our tissue culture studies indicate that amino acids located at or near the C-terminus are required for cellular transformation, membrane association and lipid...

  3. Genome-Wide Identification and Expression Analysis of the UGlcAE Gene Family in Tomato

    OpenAIRE

    Xing Ding; Jinhua Li; Yu Pan; Yue Zhang; Lei Ni; Yaling Wang; Xingguo Zhang

    2018-01-01

    The UGlcAE has the capability of interconverting UDP-d-galacturonic acid and UDP-d-glucuronic acid, and UDP-d-galacturonic acid is an activated precursor for the synthesis of pectins in plants. In this study, we identified nine UGlcAE protein-encoding genes in tomato. The nine UGlcAE genes that were distributed on eight chromosomes in tomato, and the corresponding proteins contained one or two trans-membrane domains. The phylogenetic analysis showed that SlUGlcAE genes could be divided into s...

  4. Identification of protective antigens for vaccination against systemic salmonellosis

    Directory of Open Access Journals (Sweden)

    Dirk eBumann

    2014-08-01

    Full Text Available There is an urgent medical need for improved vaccines with broad serovar coverage and high efficacy against systemic salmonellosis. Subunit vaccines offer excellent safety profiles but require identification of protective antigens, which remains a challenging task. Here, I review crucial properties of Salmonella antigens that might help to narrow down the number of potential candidates from more than 4000 proteins encoded in Salmonella genomes, to a more manageable number of 50-200 most promising antigens. I also discuss complementary approaches for antigen identification and potential limitations of current pre-clinical vaccine testing.

  5. Involvement of human endogenous retroviral syncytin-1 in human osteoclast fusion

    DEFF Research Database (Denmark)

    Søe, Kent; Andersen, Thomas Lykke; Hobolt-Pedersen, Anne-Sofie

    2011-01-01

    fusion of the lipid bilayers of their cell membranes are still unknown. Syncytin-1 is a protein encoded by a human endogenous retroviral gene which was stably integrated into the human ancestor genome more than 24 million years ago. Upon activation, syncytin-1 is able to destabilize the lipid bilayer....... This was documented through Q-PCR, Western blot and immunofluorescence analyses. These in vitro findings were confirmed by immunohistochemical stainings in human iliac crest biopsies. A syncytin-1 inhibitory peptide reduced the number of nuclei per osteoclast by 30%, as well as TRACP activity. From a mechanistic...

  6. Virus-specific proteins in cells infected with tomato black ring nepovirus: evidence for proteolytic processing in vivo.

    OpenAIRE

    Demangeat, Gerard; Hemmer, O; Reinbolt, J; Mayo, M A; Fritsch, Coralie

    1992-01-01

    The synthesis of proteins encoded by the RNA of tomato black ring virus (TBRV) in vivo was studied in protoplasts by direct labelling with [35S]methionine, and in protoplasts and plants by immunoblotting experiments with specific antisera. Comparison of the proteins synthesized in infected and mock-inoculated protoplasts suggested that proteins of M(r) 120K, 90K, 80K, 57K and 46K were virus-specific. The proteins derived from the RNA-1-encoded polyprotein detected by immunoblotting were a sta...

  7. Generation of a human embryonic stem cell line, NERCe003-A-1, with lentivirus vector-mediated inducible CTNNB1 overexpression

    Directory of Open Access Journals (Sweden)

    Yang Wang

    2018-04-01

    Full Text Available The human embryonic stem cell (hESC line NERCe003-A-1 was generated by introducing lentiviral-vector–mediated tetracycline-inducible β-catenin expression into a normal hESC line, NERCe003-A. The resulting cell line can overexpress the β-catenin protein, encoded by the CTNNB1 gene, after exposure to doxycycline (Dox. CTNNB1 gene expression was confirmed by quantitative PCR (qPCR and immunofluorescence assays. Further characterization confirmed that the NERCe003-A-1 cell line expresses typical pluripotency markers and has the ability to form the three germ layers both in vitro and in vivo.

  8. Electrotransformation and clonal isolation of Rickettsia species

    Science.gov (United States)

    Riley, Sean P; Macaluso, Kevin R; Martinez, Juan J

    2015-01-01

    Genetic manipulation of obligate intracellular bacteria of the genus Rickettsia is currently undergoing a rapid period of change. The development of viable genetic tools, including replicative plasmids, transposons, homologous recombination, fluorescent protein-encoding genes, and antibiotic selectable markers has provided the impetus for future research development. This unit is designed to coalesce the basic methods pertaining to creation of genetically modified Rickettsia. The unit describes a series of methods, from inserting exogenous DNA into Rickettsia to the final isolation of genetically modified bacterial clones. Researchers working towards genetic manipulation of Rickettsia or similar obligate intracellular bacteria will find these protocols to be a valuable reference. PMID:26528784

  9. Expression of a highly basic peroxidase gene in NaCl-adapted tomato cell suspensions.

    Science.gov (United States)

    Medina, M I; Botella, M A; Quesada, M A; Valpuesta, V

    1997-05-05

    A tomato peroxidase gene, TPX2, that is only weakly expressed in the roots of young tomato seedlings is highly expressed in tomato suspension cells adapted to high external NaCl concentration. The protein encoded by this gene, with an isolectric point value of approximately 9.6, is found in the culture medium of the growing cells. Our data suggest that the expression of TPX2 in the salt-adapted cells is not the result of the elicitation imposed by the in vitro culture or the presence of high NaCl concentration in the medium.

  10. Identification and characterization of a novel Chlamydia trachomatis reticulate body protein

    DEFF Research Database (Denmark)

    Shaw, Allan C; Larsen, Martin Røssel; Roepstorff, Peter

    2002-01-01

    The genome of the obligate intracellular bacterium Chlamydia trachomatis comprises 894 genes predicted by computer-based analysis. As part of a large-scale proteome analysis of C. trachomatis, a small abundant protein encoded by a previously unrecognized novel 204-bp open reading frame was identi...... cycle. The protein is rapidly degraded and is only present in reticulate or intermediate bodies, suggesting a possible function in the intracellular stage of C. trachomatis development. We have termed the protein '7-kDa reticulate body protein'....

  11. E2F-dependent induction of p14ARF during cell cycle re-entry in human T cells

    DEFF Research Database (Denmark)

    del Arroyo, Ana Gutierrez; El Messaoudi, Selma; Clark, Paula A

    2007-01-01

    The ARF protein, encoded by alternate exon usage within the CDKN2A locus, provides a link between the retinoblastoma (pRb) and p53 tumor suppressor pathways. Agents that disable pRb or otherwise impinge on the E2F family of transcription factors induce expression of ARF, resulting in stabilization...... of p53 and activation of p53-regulated genes. However, in some cell types ARF is not induced upon cell cycle re-entry, as expected of a conventional E2F target gene, leading to the suggestion that the ARF promoter only responds to supra-physiological or aberrant levels of E2F. These properties have...

  12. The Gly482Ser genotype at the PPARGC1A gene and elevated blood pressure: a meta-analysis involving 13,949 individuals

    DEFF Research Database (Denmark)

    Vimaleswaran, Karani Santhanakrishnan; Luan, Jian'an; Andersen, Gitte

    2008-01-01

    The protein encoded by the PPARGC1A gene is expressed at high levels in metabolically active tissues and is involved in the control of oxidative stress via reactive oxygen species detoxification. Several recent reports suggest that the PPARGC1A Gly482Ser (rs8192678) missense polymorphism may relate...... inversely with blood pressure. We used conventional meta-analysis methods to assess the association between Gly482Ser and systolic (SBP) or diastolic blood pressures (DBP) or hypertension in 13,949 individuals from 17 studies, of which 6,042 were previously unpublished observations. The studies comprised...

  13. [Expression and significance of respiratory chain enzyme of cells in urine sediment in MELAS syndrome].

    Science.gov (United States)

    Wu, Hai-rong; Ma, Yi-nan; Qi, Yu; Liu, Hong-gang

    2013-04-23

    To explore the expression and significance of respiratory chain enzyme of cells in urine sediment in mitochondrial encephalopathy myopathy, lactic acidosis and stroke-like episodes (MELAS) syndrome. Through enzyme histochemistry, the authors analyzed the changes of respiratory chain enzyme in urine sediment in 20 MELAS patients due to mitochondrial A3243G mutation (MELAS group) and 20 health peoples (control group). And the impact on the expression of protein encoded by nuclear DNA (A21347) and mitochondrial DNA (A6404) was detected by immunochemistry. Image pro Plus 6.0 software was used for analysis of absorbance (A) of staining images as staining intensity. The data were expressed as M (Q1, Q3) and analyzed through statistical software. The staining intensity of complexes Iin the MELAS group was lower than that in the control group (0.06(0.01, 0.12) vs 0.12(0.01, 0.62), P = 0.010). The intergroup staining intensity of complex II showed no marked difference. Increased density of blue particle and cytoplasmic gathering was found in 13 cased (65%) of the MELAS group under light microscope. The staining intensity of complexes IV was expressed at a low level in the MELAS group (0.14(0.03, 0.32) vs 0.23(0.06, 0.43), P = 0.038). The expression of protein encoded by nuclear DNA (A21347) was lower than that in the control group (0.05(0.02, 0.45) vs 0.17(0.03, 0.70), P = 0.000). The expression of protein encoded by mitochondrial DNA (A6404) was also lower than that in the control group (0.03(0.01, 0.07) vs 0.15 (0.09, 0.23), P = 0.000). Abnormal change of respiratory chain enzyme in urine sediment in MELAS due to mitochondrial A3243G mutation and a low expression of proteins encoded by two kinds of DNA in complexes IV can help to confirm the genetic diagnosis of mitochondrial encephalomyopathies so that different subtypes may be classified and its pathogenesis elucidated.

  14. Systematic Characterisation of Cellular Localisation and Expression Profiles of Proteins Containing MHC Ligands

    DEFF Research Database (Denmark)

    Juncker, Agnieszka; Larsen, Mette Voldby; Weinhold, Nils

    2009-01-01

    Background: Presentation of peptides on Major Histocompatibility Complex (MHC) molecules is the cornerstone in immune system activation and increased knowledge of the characteristics of MHC ligands and their source proteins is highly desirable. Methodology/Principal Finding: In the present large......-scale study, we used a large data set of proteins containing experimentally identified MHC class I or II ligands and examined the proteins according to their expression profiles at the mRNA level and their Gene Ontology (GO) classification within the cellular component ontology. Proteins encoded by highly...

  15. Adenovirus E4-ORF1 Dysregulates Epidermal Growth Factor and Insulin/Insulin-Like Growth Factor Receptors To Mediate Constitutive Myc Expression

    OpenAIRE

    Kong, Kathleen; Kumar, Manish; Taruishi, Midori; Javier, Ronald T.

    2015-01-01

    The E4-ORF1 protein encoded by human adenovirus stimulates viral replication in human epithelial cells by binding and activating cellular phosphatidylinositol 3-kinase (PI3K) at the plasma membrane and cellular Myc in the nucleus. In this study, we showed that E4-ORF1 hijacks the tyrosine kinase activities of cellular epidermal growth factor receptor (EGFR) and insulin receptor (InsR)/insulin-like growth factor receptor 1 (IGF1R), as well as the lipid kinase activity of PI3K, to mediate const...

  16. Link of the unique oncogenic properties of adenovirus type 9 E4-ORF1 to a select interaction with the candidate tumor suppressor protein ZO-2

    OpenAIRE

    Glaunsinger, Britt A.; Weiss, Robert S.; Lee, Siu Sylvia; Javier, Ronald

    2001-01-01

    Adenovirus type 9 (Ad9) is distinct among human adenoviruses because it elicits solely mammary tumors in animals and its primary oncogenic determinant is the E4 region-encoded ORF1 (E4-ORF1) protein. We report here that the PDZ domain-containing protein ZO-2, which is a candidate tumor suppressor protein, is a cellular target for tumorigenic Ad9 E4-ORF1 but not for non-tumorigenic wild-type E4-ORF1 proteins encoded by adenovirus types 5 and 12. Complex formation was mediated by the C-terminal...

  17. Nucleotide sequence of a chickpea chlorotic stunt virus relative that infects pea and faba bean in China.

    Science.gov (United States)

    Zhou, Cui-Ji; Xiang, Hai-Ying; Zhuo, Tao; Li, Da-Wei; Yu, Jia-Lin; Han, Cheng-Gui

    2012-07-01

    We determined the genome sequence of a new polerovirus that infects field pea and faba bean in China. Its entire nucleotide sequence (6021 nt) was most closely related (83.3% identity) to that of an Ethiopian isolate of chickpea chlorotic stunt virus (CpCSV-Eth). With the exception of the coat protein (encoded by ORF3), amino acid sequence identities of all gene products of this virus to those of CpCSV-Eth and other poleroviruses were Polerovirus, and the name pea mild chlorosis virus is proposed.

  18. Characterization of large-insert DNA libraries from soil for environmental genomic studies of Archaea

    DEFF Research Database (Denmark)

    Treusch, Alexander H; Kletzin, Arnulf; Raddatz, Guenter

    2004-01-01

    Complex genomic libraries are increasingly being used to retrieve complete genes, operons or large genomic fragments directly from environmental samples, without the need to cultivate the respective microorganisms. We report on the construction of three large-insert fosmid libraries in total...... (approximately 1% each) have been captured in our libraries. The diversity of putative protein-encoding genes, as reflected by their distribution into different COG clusters, was comparable to that encoded in complete genomes of cultivated microorganisms. A huge variety of genomic fragments has been captured...

  19. Genetic and epigenetic regulation of YKL-40 in childhood.

    Science.gov (United States)

    Guerra, Stefano; Melén, Erik; Sunyer, Jordi; Xu, Cheng-Jian; Lavi, Iris; Benet, Marta; Bustamante, Mariona; Carsin, Anne-Elie; Dobaño, Carlota; Guxens, Mònica; Tischer, Christina; Vrijheid, Martine; Kull, Inger; Bergström, Anna; Kumar, Ashish; Söderhäll, Cilla; Gehring, Ulrike; Dijkstra, Dorieke J; van der Vlies, Pieter; Wickman, Magnus; Bousquet, Jean; Postma, Dirkje S; Anto, Josep M; Koppelman, Gerard H

    2018-03-01

    Circulating levels of the chitinase-like protein YKL-40 are influenced by genetic variation in its encoding gene (chitinase 3-like 1 [CHI3L1]) and are increased in patients with several diseases, including asthma. Epigenetic regulation of circulating YKL-40 early in life is unknown. We sought to determine (1) whether methylation levels at CHI3L1 CpG sites mediate the association of CHI3L1 single nucleotide polymorphisms (SNPs) with YKL-40 levels in the blood and (2) whether these biomarkers (CHI3L1 SNPs, methylation profiles, and YKL-40 levels) are associated with asthma in early childhood. We used data from up to 2405 participants from the Spanish Infancia y Medio Ambiente; the Swedish Barn/Children, Allergy, Milieu, Stockholm, Epidemiological survey; and the Dutch Prevention and Incidence of Asthma and Mite Allergy birth cohorts. Associations between 68 CHI3L1 SNPs, methylation levels at 14 CHI3L1 CpG sites in whole-blood DNA, and circulating YKL-40 levels at 4 years of age were tested by using correlation analysis, multivariable regression, and mediation analysis. Each of these biomarkers was also tested for association with asthma at 4 years of age by using multivariable logistic regression. YKL-40 levels were significantly associated with 7 SNPs and with methylation at 5 CpG sites. Consistent associations between these 7 SNPs (particularly rs10399931 and rs4950928) and 5 CpG sites were observed. Alleles linked to lower YKL-40 levels were associated with higher methylation levels. Participants with high YKL-40 levels (defined as the highest YKL-40 tertile) had increased odds for asthma compared with subjects with low YKL-40 levels (meta-analyzed adjusted odds ratio, 1.90 [95% CI, 1.08-3.36]). In contrast, neither SNPs nor methylation levels at CpG sites in CHI3L1 were associated with asthma. The effects of CHI3L1 genetic variation on circulating YKL-40 levels are partly mediated by methylation profiles. In our study YKL-40 levels, but not CHI3L1 SNPs or

  20. Differential expression and function of breast regression protein 39 (BRP-39 in murine models of subacute cigarette smoke exposure and allergic airway inflammation

    Directory of Open Access Journals (Sweden)

    Coyle Anthony J

    2011-04-01

    Full Text Available Abstract Background While the presence of the chitinase-like molecule YKL40 has been reported in COPD and asthma, its relevance to inflammatory processes elicited by cigarette smoke and common environmental allergens, such as house dust mite (HDM, is not well understood. The objective of the current study was to assess expression and function of BRP-39, the murine equivalent of YKL40 in a murine model of cigarette smoke-induced inflammation and contrast expression and function to a model of HDM-induced allergic airway inflammation. Methods CD1, C57BL/6, and BALB/c mice were room air- or cigarette smoke-exposed for 4 days in a whole-body exposure system. In separate experiments, BALB/c mice were challenged with HDM extract once a day for 10 days. BRP-39 was assessed by ELISA and immunohistochemistry. IL-13, IL-1R1, IL-18, and BRP-39 knock out (KO mice were utilized to assess the mechanism and relevance of BRP-39 in cigarette smoke- and HDM-induced airway inflammation. Results Cigarette smoke exposure elicited a robust induction of BRP-39 but not the catalytically active chitinase, AMCase, in lung epithelial cells and alveolar macrophages of all mouse strains tested. Both BRP-39 and AMCase were increased in lung tissue after HDM exposure. Examining smoke-exposed IL-1R1, IL-18, and IL-13 deficient mice, BRP-39 induction was found to be IL-1 and not IL-18 or IL-13 dependent, while induction of BRP-39 by HDM was independent of IL-1 and IL-13. Despite the importance of BRP-39 in cellular inflammation in HDM-induced airway inflammation, BRP-39 was found to be redundant for cigarette smoke-induced airway inflammation and the adjuvant properties of cigarette smoke. Conclusions These data highlight the contrast between the importance of BRP-39 in HDM- and cigarette smoke-induced inflammation. While functionally important in HDM-induced inflammation, BRP-39 is a biomarker of cigarette smoke induced inflammation which is the byproduct of an IL-1

  1. Transcriptional responses in honey bee larvae infected with chalkbrood fungus.

    Science.gov (United States)

    Aronstein, Katherine A; Murray, Keith D; Saldivar, Eduardo

    2010-06-21

    Diseases and other stress factors working synergistically weaken honey bee health and may play a major role in the losses of bee populations in recent years. Among a large number of bee diseases, chalkbrood has been on the rise. We present here the experimental identification of honey bee genes that are differentially expressed in response to infection of honey bee larvae with the chalkbrood fungus, Ascosphaera apis. We used cDNA-AFLP Technology to profile transcripts in infected and uninfected bee larvae. From 64 primer combinations, over 7,400 transcriptionally-derived fragments were obtained A total of 98 reproducible polymorphic cDNA-AFLP fragments were excised and sequenced, followed by quantitative real-time RT-PCR (qRT-PCR) analysis of these and additional samples.We have identified a number of differentially-regulated transcripts that are implicated in general mechanisms of stress adaptation, including energy metabolism and protein transport. One of the most interesting differentially-regulated transcripts is for a chitinase-like enzyme that may be linked to anti-fungal activities in the honey bee larvae, similarly to gut and fat-body specific chitinases found in mosquitoes and the red flour beetle. Surprisingly, we did not find many components of the well-characterized NF-kappaB intracellular signaling pathways to be differentially-regulated using the cDNA-AFLP approach. Therefore, utilizing qRT-PCR, we probed some of the immune related genes to determine whether the lack of up-regulation of their transcripts in our analysis can be attributed to lack of immune activation or to limitations of the cDNA-AFLP approach. Using a combination of cDNA-AFLP and qRT-PCR analyses, we were able to determine several key transcriptional events that constitute the overall effort in the honey bee larvae to fight natural fungal infection. Honey bee transcripts identified in this study are involved in critical functions related to transcriptional regulation, apoptotic

  2. Human microcephaly protein RTTN interacts with STIL and is required to build full-length centrioles.

    Science.gov (United States)

    Chen, Hsin-Yi; Wu, Chien-Ting; Tang, Chieh-Ju C; Lin, Yi-Nan; Wang, Won-Jing; Tang, Tang K

    2017-08-15

    Mutations in many centriolar protein-encoding genes cause primary microcephaly. Using super-resolution and electron microscopy, we find that the human microcephaly protein, RTTN, is recruited to the proximal end of the procentriole at early S phase, and is located at the inner luminal walls of centrioles. Further studies demonstrate that RTTN directly interacts with STIL and acts downstream of STIL-mediated centriole assembly. CRISPR/Cas9-mediated RTTN gene knockout in p53-deficient cells induce amplification of primitive procentriole bodies that lack the distal-half centriolar proteins, POC5 and POC1B. Additional analyses show that RTTN serves as an upstream effector of CEP295, which mediates the loading of POC1B and POC5 to the distal-half centrioles. Interestingly, the naturally occurring microcephaly-associated mutant, RTTN (A578P), shows a low affinity for STIL binding and blocks centriole assembly. These findings reveal that RTTN contributes to building full-length centrioles and illuminate the molecular mechanism through which the RTTN (A578P) mutation causes primary microcephaly.Mutations in many centriolar protein-encoding genes cause primary microcephaly. Here the authors show that human microcephaly protein RTTN directly interacts with STIL and acts downstream of STIL-mediated centriole assembly, contributing to building full-length centrioles.

  3. TED, an Autonomous and Rare Maize Transposon of the Mutator Superfamily with a High Gametophytic Excision Frequency[W

    Science.gov (United States)

    Li, Yubin; Harris, Linda; Dooner, Hugo K.

    2013-01-01

    Mutator (Mu) elements, one of the most diverse superfamilies of DNA transposons, are found in all eukaryotic kingdoms, but are particularly numerous in plants. Most of the present knowledge on the transposition behavior of this superfamily comes from studies of the maize (Zea mays) Mu elements, whose transposition is mediated by the autonomous Mutator-Don Robertson (MuDR) element. Here, we describe the maize element TED (for Transposon Ellen Dempsey), an autonomous cousin that differs significantly from MuDR. Element excision and reinsertion appear to require both proteins encoded by MuDR, but only the single protein encoded by TED. Germinal excisions, rare with MuDR, are common with TED, but arise in one of the mitotic divisions of the gametophyte, rather than at meiosis. Instead, transposition-deficient elements arise at meiosis, suggesting that the double-strand breaks produced by element excision are repaired differently in mitosis and meiosis. Unlike MuDR, TED is a very low-copy transposon whose number and activity do not undergo dramatic changes upon inbreeding or outcrossing. Like MuDR, TED transposes mostly to unlinked sites and can form circular transposition products. Sequences closer to TED than to MuDR were detected only in the grasses, suggesting a rather recent evolutionary split from a common ancestor. PMID:24038653

  4. Functional substitution for TAF(II)250 by a retroposed homolog that is expressed in human spermatogenesis.

    Science.gov (United States)

    Wang, P Jeremy; Page, David C

    2002-09-15

    TAF(II)250, the largest subunit of the general transcription factor TFIID, is expressed from the human X chromosome, at least in somatic cells. In male meiosis, however, the sex chromosomes are transcriptionally silenced, while the autosomes remain active. How then are protein-encoding genes transcribed during human male meiosis? Here we present a novel autosomal human gene, TAF1L, which is homologous to TAF(II)250 and is expressed specifically in the testis, apparently in germ cells. We hypothesize that during male meiosis, transcription of protein-encoding genes relies upon TAF1L as a functional substitute for TAF(II)250. Like TAF(II)250, the human TAF1L protein can bind directly to TATA-binding protein, an essential component of TFIID. Most importantly, transfection with human TAF1L rescued the temperature-sensitive lethality of a hamster cell line mutant in TAF(II)250. TAF1L lacks introns and evidently arose by retroposition of a processed TAF(II)250 mRNA during primate evolution. The observation that TAF1L can functionally replace TAF(II)250 provides experimental support for the hypothesis that during male meiosis, autosomes provide cellular functions usually supplied by the X chromosome in somatic cells.

  5. Functional characterization of MAT1-1-specific mating-type genes in the homothallic ascomycete Sordaria macrospora provides new insights into essential and nonessential sexual regulators.

    Science.gov (United States)

    Klix, V; Nowrousian, M; Ringelberg, C; Loros, J J; Dunlap, J C; Pöggeler, S

    2010-06-01

    Mating-type genes in fungi encode regulators of mating and sexual development. Heterothallic ascomycete species require different sets of mating-type genes to control nonself-recognition and mating of compatible partners of different mating types. Homothallic (self-fertile) species also carry mating-type genes in their genome that are essential for sexual development. To analyze the molecular basis of homothallism and the role of mating-type genes during fruiting-body development, we deleted each of the three genes, SmtA-1 (MAT1-1-1), SmtA-2 (MAT1-1-2), and SmtA-3 (MAT1-1-3), contained in the MAT1-1 part of the mating-type locus of the homothallic ascomycete species Sordaria macrospora. Phenotypic analysis of deletion mutants revealed that the PPF domain protein-encoding gene SmtA-2 is essential for sexual reproduction, whereas the alpha domain protein-encoding genes SmtA-1 and SmtA-3 play no role in fruiting-body development. By means of cross-species microarray analysis using Neurospora crassa oligonucleotide microarrays hybridized with S. macrospora targets and quantitative real-time PCR, we identified genes expressed under the control of SmtA-1 and SmtA-2. Both genes are involved in the regulation of gene expression, including that of pheromone genes.

  6. Functional Characterization of MAT1-1-Specific Mating-Type Genes in the Homothallic Ascomycete Sordaria macrospora Provides New Insights into Essential and Nonessential Sexual Regulators▿†

    Science.gov (United States)

    Klix, V.; Nowrousian, M.; Ringelberg, C.; Loros, J. J.; Dunlap, J. C.; Pöggeler, S.

    2010-01-01

    Mating-type genes in fungi encode regulators of mating and sexual development. Heterothallic ascomycete species require different sets of mating-type genes to control nonself-recognition and mating of compatible partners of different mating types. Homothallic (self-fertile) species also carry mating-type genes in their genome that are essential for sexual development. To analyze the molecular basis of homothallism and the role of mating-type genes during fruiting-body development, we deleted each of the three genes, SmtA-1 (MAT1-1-1), SmtA-2 (MAT1-1-2), and SmtA-3 (MAT1-1-3), contained in the MAT1-1 part of the mating-type locus of the homothallic ascomycete species Sordaria macrospora. Phenotypic analysis of deletion mutants revealed that the PPF domain protein-encoding gene SmtA-2 is essential for sexual reproduction, whereas the α domain protein-encoding genes SmtA-1 and SmtA-3 play no role in fruiting-body development. By means of cross-species microarray analysis using Neurospora crassa oligonucleotide microarrays hybridized with S. macrospora targets and quantitative real-time PCR, we identified genes expressed under the control of SmtA-1 and SmtA-2. Both genes are involved in the regulation of gene expression, including that of pheromone genes. PMID:20435701

  7. How do eubacterial organisms manage aggregation-prone proteome? [v1; ref status: indexed, http://f1000r.es/3k3

    Directory of Open Access Journals (Sweden)

    Rishi Das Roy

    2014-06-01

    Full Text Available Eubacterial genomes vary considerably in their nucleotide composition. The percentage of genetic material constituted by guanosine and cytosine (GC nucleotides ranges from 20% to 70%.  It has been posited that GC-poor organisms are more dependent on protein folding machinery. Previous studies have ascribed this to the accumulation of mildly deleterious mutations in these organisms due to population bottlenecks. This phenomenon has been supported by protein folding simulations, which showed that proteins encoded by GC-poor organisms are more prone to aggregation than proteins encoded by GC-rich organisms. To test this proposition using a genome-wide approach, we classified different eubacterial proteomes in terms of their aggregation propensity and chaperone-dependence using multiple machine learning models. In contrast to the expected decrease in protein aggregation with an increase in GC richness, we found that the aggregation propensity of proteomes increases with GC content. A similar and even more significant correlation was obtained with the GroEL-dependence of proteomes: GC-poor proteomes have evolved to be less dependent on GroEL than GC-rich proteomes. We thus propose that a decrease in eubacterial GC content may have been selected in organisms facing proteostasis problems.

  8. Direct interactions of the five known Fanconi anaemia proteins suggest a common functional pathway.

    Science.gov (United States)

    Medhurst, A L; Huber, P A; Waisfisz, Q; de Winter , J P; Mathew, C G

    2001-02-15

    Fanconi anaemia (FA) is an autosomal recessive inherited disorder associated with a progressive aplastic anaemia, diverse congenital abnormalities and cancer. The condition is genetically heterogeneous, with at least seven complementation groups (A-G) described. Cells from individuals who are homozygous for mutations in FA genes are characterized by chromosomal instability and hypersensitivity to DNA interstrand crosslinking agents. These features suggest a possible role for the encoded proteins in the recognition or repair of these lesions, but neither their function nor whether they operate in a concerted or discrete functional pathways is known. The recent cloning of the FANCF and FANCE genes has allowed us to investigate the interaction of the proteins encoded by five of the seven complementation groups of FA. We used the yeast two-hybrid system and co-immunoprecipitation analysis to test the 10 possible pairs of proteins for direct interaction. In addition to the previously described binding of FANCA to FANCG, we now demonstrate direct interaction of FANCF with FANCG, of FANCC with FANCE and a weaker interaction of FANCE with both FANCA and FANCG. These findings show that the newly identified FANCE protein is an integral part of the FA pathway, and support the concept of a functional link between all known proteins encoded by the genes that are mutated in this disorder. These proteins may act either as a multimeric complex or by sequential recruitment of subsets of the proteins in a common pathway that protects the genomic integrity of mammalian cells.

  9. Proteome and membrane fatty acid analyses on Oligotropha carboxidovorans OM5 grown under chemolithoautotrophic and heterotrophic conditions.

    Directory of Open Access Journals (Sweden)

    Debarati Paul

    Full Text Available Oligotropha carboxidovorans OM5 T. (DSM 1227, ATCC 49405 is a chemolithoautotrophic bacterium able to utilize CO and H(2 to derive energy for fixation of CO(2. Thus, it is capable of growth using syngas, which is a mixture of varying amounts of CO and H(2 generated by organic waste gasification. O. carboxidovorans is capable also of heterotrophic growth in standard bacteriologic media. Here we characterize how the O. carboxidovorans proteome adapts to different lifestyles of chemolithoautotrophy and heterotrophy. Fatty acid methyl ester (FAME analysis of O. carboxidovorans grown with acetate or with syngas showed that the bacterium changes membrane fatty acid composition. Quantitative shotgun proteomic analysis of O. carboxidovorans grown in the presence of acetate and syngas showed production of proteins encoded on the megaplasmid for assimilating CO and H(2 as well as proteins encoded on the chromosome that might have contributed to fatty acid and acetate metabolism. We found that adaptation to chemolithoautotrophic growth involved adaptations in cell envelope, oxidative homeostasis, and metabolic pathways such as glyoxylate shunt and amino acid/cofactor biosynthetic enzymes.

  10. Comparative Analysis of 37 Acinetobacter Bacteriophages

    Directory of Open Access Journals (Sweden)

    Dann Turner

    2017-12-01

    Full Text Available Members of the genus Acinetobacter are ubiquitous in the environment and the multiple-drug resistant species A. baumannii is of significant clinical concern. This clinical relevance is currently driving research on bacterial viruses infecting A. baumannii, in an effort to implement phage therapy and phage-derived antimicrobials. Initially, a total of 42 Acinetobacter phage genome sequences were available in the international nucleotide sequence databases, corresponding to a total of 2.87 Mbp of sequence information and representing all three families of the order Caudovirales and a single member of the Leviviridae. A comparative bioinformatics analysis of 37 Acinetobacter phages revealed that they form six discrete clusters and two singletons based on genomic organisation and nucleotide sequence identity. The assignment of these phages to clusters was further supported by proteomic relationships established using OrthoMCL. The 4067 proteins encoded by the 37 phage genomes formed 737 groups and 974 orphans. Notably, over half of the proteins encoded by the Acinetobacter phages are of unknown function. The comparative analysis and clustering presented enables an updated taxonomic framing of these clades.

  11. Prioritization of candidate disease genes by combining topological similarity and semantic similarity.

    Science.gov (United States)

    Liu, Bin; Jin, Min; Zeng, Pan

    2015-10-01

    The identification of gene-phenotype relationships is very important for the treatment of human diseases. Studies have shown that genes causing the same or similar phenotypes tend to interact with each other in a protein-protein interaction (PPI) network. Thus, many identification methods based on the PPI network model have achieved good results. However, in the PPI network, some interactions between the proteins encoded by candidate gene and the proteins encoded by known disease genes are very weak. Therefore, some studies have combined the PPI network with other genomic information and reported good predictive performances. However, we believe that the results could be further improved. In this paper, we propose a new method that uses the semantic similarity between the candidate gene and known disease genes to set the initial probability vector of a random walk with a restart algorithm in a human PPI network. The effectiveness of our method was demonstrated by leave-one-out cross-validation, and the experimental results indicated that our method outperformed other methods. Additionally, our method can predict new causative genes of multifactor diseases, including Parkinson's disease, breast cancer and obesity. The top predictions were good and consistent with the findings in the literature, which further illustrates the effectiveness of our method. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Nucleotide sequence of the human N-myc gene

    International Nuclear Information System (INIS)

    Stanton, L.W.; Schwab, M.; Bishop, J.M.

    1986-01-01

    Human neuroblastomas frequently display amplification and augmented expression of a gene known as N-myc because of its similarity to the protooncogene c-myc. It has therefore been proposed that N-myc is itself a protooncogene, and subsequent tests have shown that N-myc and c-myc have similar biological activities in cell culture. The authors have now detailed the kinship between N-myc and c-myc by determining the nucleotide sequence of human N-myc and deducing the amino acid sequence of the protein encoded by the gene. The topography of N-myc is strikingly similar to that of c-myc: both genes contain three exons of similar lengths; the coding elements of both genes are located in the second and third exons; and both genes have unusually long 5' untranslated regions in their mRNAs, with features that raise the possibility that expression of the genes may be subject to similar controls of translation. The resemblance between the proteins encoded by N-myc and c-myc sustains previous suspicions that the genes encode related functions

  13. Synergistic interactions between RAD5, RAD16, and RAD54, three partially homologous yeast DNA repair genes each in a different repair pathway

    International Nuclear Information System (INIS)

    Glassner, B.J.; Mortimer, R.K.

    1994-01-01

    Considerable homology has recently been noted between the proteins encoded by the RAD5, RAD16 and RAD54 genes of Saccharomyces cerevisiae. These genes are members of the RAD6, RAD3 and RAD50 epistasis groups, respectively, which correspond to the three major DNA repair pathways in yeast. These proteins also share homology with other eucaryotic proteins, including those encoded by SNF2 and MO1 of yeast, brahma and lodestar of Drosophila and the human ERCC6 gene. The homology shares features with known helicases, suggesting a newly identified helicase subfamily. We have constructed a series of congenic single-, double- and triple-deletion mutants involving RAD5, RAD16 and RAD54 to examine the interactions between these genes. Each deletion mutation alone has only a moderate effect on survival after exposure to UV radiation. Each pairwise-double mutant exhibits marked synergism. The triple-deletion mutant displays further synergism. These results confirm the assignment of the RAD54 gene to the RAD50 epistasis group and suggest that the RAD16 gene plays a larger role in DNA repair after exposure to UV radiation than has been suggested previously. Additionally, the proteins encoded by RAD5, RAD16, and RAD54 may compete for the same substrate after damage induced by UV radiation, possibly at an early step in their respective pathways. 49 refs., 6 figs., 2 tabs

  14. TED, an autonomous and rare maize transposon of the mutator superfamily with a high gametophytic excision frequency.

    Science.gov (United States)

    Li, Yubin; Harris, Linda; Dooner, Hugo K

    2013-09-01

    Mutator (Mu) elements, one of the most diverse superfamilies of DNA transposons, are found in all eukaryotic kingdoms, but are particularly numerous in plants. Most of the present knowledge on the transposition behavior of this superfamily comes from studies of the maize (Zea mays) Mu elements, whose transposition is mediated by the autonomous Mutator-Don Robertson (MuDR) element. Here, we describe the maize element TED (for Transposon Ellen Dempsey), an autonomous cousin that differs significantly from MuDR. Element excision and reinsertion appear to require both proteins encoded by MuDR, but only the single protein encoded by TED. Germinal excisions, rare with MuDR, are common with TED, but arise in one of the mitotic divisions of the gametophyte, rather than at meiosis. Instead, transposition-deficient elements arise at meiosis, suggesting that the double-strand breaks produced by element excision are repaired differently in mitosis and meiosis. Unlike MuDR, TED is a very low-copy transposon whose number and activity do not undergo dramatic changes upon inbreeding or outcrossing. Like MuDR, TED transposes mostly to unlinked sites and can form circular transposition products. Sequences closer to TED than to MuDR were detected only in the grasses, suggesting a rather recent evolutionary split from a common ancestor.

  15. Isolation and characterization of a copalyl diphosphate synthase gene promoter from Salvia miltiorrhiza

    Directory of Open Access Journals (Sweden)

    Piotr Szymczyk

    2016-09-01

    Full Text Available The promoter, 5' UTR, and 34-nt 5' fragments of protein encoding region of the Salvia miltiorrhiza copalyl diphosphate synthase gene were cloned and characterized. No tandem repeats, miRNA binding sites, or CpNpG islands were observed in the promoter, 5' UTR, or protein encoding fragments. The entire isolated promoter and 5' UTR is 2235 bp long and contains repetitions of many cis-active elements, recognized by homologous transcription factors, found in Arabidopsis thaliana and other plant species. A pyrimidine-rich fragment with only 6 non-pyrimidine bases was localized in the 33-nt stretch from nt 2185 to 2217 in the 5' UTR. The observed cis-active sequences are potential binding sites for trans-factors that could regulate spatio-temporal CPS gene expression in response to biotic and abiotic stress conditions. Obtained results are initially verified by in silico and co-expression studies based on A. thaliana microarray data. The quantitative RT-PCR analysis confirmed that the entire 2269-bp copalyl diphosphate synthase gene fragment has the promoter activity. Quantitative RT-PCR analysis was used to study changes in CPS promoter activity occurring in response to the application of four selected biotic and abiotic regulatory factors; auxin, gibberellin, salicylic acid, and high-salt concentration.

  16. Agrobacterium T-DNA-encoded protein Atu6002 interferes with the host auxin response

    Science.gov (United States)

    Lacroix, Benoît; Gizatullina, Diana I.; Babst, Benjamin A.; Gifford, Andrew N.; Citovsky, Vitaly

    2013-01-01

    Summary Several genes in the Agrobacterium tumefaciens transferred (T) DNA encode proteins that are involved in developmental alterations leading to the formation of tumors in infected plants. We investigated the role of the protein encoded by the Atu6002 gene, the function of which is completely unknown. The Atu6002 expression occurs in Agrobacterium-induced tumors, and is also activated upon activation of plant cell division by growth hormones. Within the expressing plant cells, the Atu6002 protein is targeted to the plasma membrane. Interestingly, constitutive ectopic expression of Atu6002 in transgenic tobacco plants lead to a severe developmental phenotype characterized by stunted growth, shorter internodes, lanceolate leaves, increased branching, and modified flower morphology. These Atu6002-expressing plants also displayed impaired response to auxin. However, auxin cellular uptake and polar transport were not significantly inhibited in these plants, suggesting that Atu6002 interferes with auxin perception or signaling pathways. PMID:24128370

  17. Phylogenetic characterization of the ubiquitous electron transfer flavoprotein families ETF-alpha and ETF-beta.

    Science.gov (United States)

    Tsai, M H; Saier, M H

    1995-06-01

    Electron transfer flavoproteins (ETF) are alpha beta-heterodimers found in eukaryotic mitochondria and bacteria. We have identified currently sequenced protein members of the ETF-alpha and ETF-beta families. Members of these two families include (a) the ETF subunits of mammals and bacteria, (b) homologous pairs of proteins (FixB/FixA) that are essential for nitrogen fixation in some bacteria, and (c) a pair of carnitine-inducible proteins encoded by two open reading frames in Escherichia coli (YaaQ and YaaR). These three groups of proteins comprise three clusters on both the ETF-alpha and ETF-beta phylogenetic trees, separated from each other by comparable phylogenetic distances. This fact suggests that these two protein families evolved with similar overall rates of evolutionary divergence. Relative regions of sequence conservation are evaluated, and signature sequences for both families are derived.

  18. Ataxin-1 with an expanded glutamine tract alters nuclear matrix-associated structures

    DEFF Research Database (Denmark)

    Skinner, P J; Koshy, B T; Cummings, C J

    1997-01-01

    a similar pattern of nuclear localization; with expanded ataxin-1 occurring in larger structures that are fewer in number than those of normal ataxin-1. Colocalization studies show that mutant ataxin-1 causes a specific redistribution of the nuclear matrix-associated domain containing promyelocytic...... the subcellular localization of wild-type human ataxin-1 (the protein encoded by the SCA1 gene) and mutant ataxin-1 in the Purkinje cells of transgenic mice. We found that ataxin-1 localizes to the nuclei of cerebellar Purkinje cells. Normal ataxin-1 localizes to several nuclear structures approximately 0.......5 microm across, whereas the expanded ataxin-1 localizes to a single approximately 2-microm structure, before the onset of ataxia. Mutant ataxin-1 localizes to a single nuclear structure in affected neurons of SCA1 patients. Similarly, COS-1 cells transfected with wild-type or mutant ataxin-1 show...

  19. Allelic deletions of cell growth regulators during progression of bladder cancer

    DEFF Research Database (Denmark)

    Primdahl, H; von der Maase, H; Christensen, M

    2000-01-01

    Cell growth regulators include proteins of the p53 pathway encoded by the genes CDKN2A (p16, p14arf), MDM2, TP53, and CDKN1A (p21) as well as proteins encoded by genes like RB1, E2F, and MYCL. In the present study we investigated allelic deletions of all these genes in each recurrent bladder tumor...... difference in the numbers of gene loci hit by deletions muscle-invasive versus noninvasive tumors (P = 0.0000002), with the genes most often hit by deletions in muscle-invasive tumors being TP53, RB1, and MYCL. A number of novel findings were made. Losses of MYCL and RB1 alleles were more pronounced...... that a characteristic difference between recurrent noninvasive and recurrent progressing bladder tumors is loss of cell cycle-regulatory genes in the latter group....

  20. Somatic mutations in the transcriptional corepressor gene BCORL1 in adult acute myelogenous leukemia.

    Science.gov (United States)

    Li, Meng; Collins, Roxane; Jiao, Yuchen; Ouillette, Peter; Bixby, Dale; Erba, Harry; Vogelstein, Bert; Kinzler, Kenneth W; Papadopoulos, Nickolas; Malek, Sami N

    2011-11-24

    To further our understanding of the genetic basis of acute myelogenous leukemia (AML), we determined the coding exon sequences of ∼ 18 000 protein-encoding genes in 8 patients with secondary AML. Here we report the discovery of novel somatic mutations in the transcriptional corepressor gene BCORL1 that is located on the X-chromosome. Analysis of BCORL1 in an unselected cohort of 173 AML patients identified a total of 10 mutated cases (6%) with BCORL1 mutations, whereas analysis of 19 AML cell lines uncovered 4 (21%) BCORL1 mutated cell lines. The majority (87%) of the mutations in BCORL1 were predicted to inactivate the gene product as a result of nonsense mutations, splice site mutation, or out-of-frame insertions or deletions. These results indicate that BCORL1 by genetic criteria is a novel candidate tumor suppressor gene, joining the growing list of genes recurrently mutated in AML.

  1. Further delineation of facioaudiosymphalangism syndrome

    DEFF Research Database (Denmark)

    Bayat, Allan; Fijalkowski, Igor; Nygaard, Tobias

    2016-01-01

    Mutations in the NOG gene give rise to a wide range of clinical phenotypes. Noggin, the protein encoded by this gene is a secreted modulator of multiple pathways involved in both bone and joint development. Proximal symphalangism is commonly observed in patients bearing mutations in this gene, ho...... by affected individuals but no hearing loss, further adding to the phenotypic variability of the syndrome. With these findings we broaden the understanding of NOG-related-symphalangism spectrum disorder. © 2016 Wiley Periodicals, Inc......., however secondary symptomes are often found including typical facies with hemicylindrical nose with bulbous tip, hyperopia, reduced mobility of multiple joints, hearing loss due to stapes fixation, and recurrent pain from affected joints. With large variation of the phenotype both within and between...

  2. Identification of Molecular and Cellular Responses of Desulfovibrio vulgaris Biofilms under Culture Conditions Relevant to Field Conditions for Bioreduction of Toxic Metals and Radionuclides. Final report

    International Nuclear Information System (INIS)

    Wall, Judy D.

    2011-01-01

    Our findings demonstrated that D. vulgaris surface-adhered populations produce extracellular structures, and that the cells have altered carbon and energy flux compared to planktonic cells. Biofilms did not have greatly increased carbohydrate accumulation. Interestingly genes present on the native plasmid found in D. vulgaris Hildenborough were necessary for wild type biofilm formation. In addition, extracellular appendages dependent on functions or proteins encoded by flaG or fliA also contributed to biofilm formation. Studies with SRB biofilms have indicated that the reduction and precipitation of metals can occur within the biofilm matrix; however, little work has been done to elucidate the physiological state of surface-adhered cells during metal reduction (Cr6+, U6+) and how this process is affected by nutrient feed levels (i.e., the stimulant).

  3. Structural Insights Into The Bacterial Carbon-Phosphorus Lyase Machinery

    DEFF Research Database (Denmark)

    Brodersen, Ditlev Egeskov

    the proteins encoded in the phn operon act in concert to catabolise phosphonate remain unknown. We have determined the crystal structure of a 240 kDa Escherichia coli carbon-phosphorus lyase core complex at 1.7 Å and show that it comprises a highly intertwined network of subunits with several unexpected......Phosphonate compounds act as a nutrient source for some microorganisms when phosphate is limiting but require a specialised enzymatic machinery due to the presence of the highly stable carbon-phosphorus bond. Despite the fundamental importance to microbial metabolism, the details of how...... structural features. The complex contains at least two different active sites and suggest a revision of current models of carbon-phosphorus bond cleavage. Using electron microscopy, we map the binding site of an additional protein subunit, which may use ATP for driving conformational changes during...

  4. The genes coding for the hsp70(dnaK) molecular chaperone machine occur in the moderate thermophilic archaeon Methanosarcina thermophila TM-1

    DEFF Research Database (Denmark)

    Hofman-Bang, H Jacob Peider; Lange, Marianne; Ahring, Birgitte Kiær

    1999-01-01

    The hsp70 (dnaK) locus of the moderate thermophilic archaeon Methanosarcina thermophila TM-1 was cloned, sequenced, and tested in vitro to measure gene induction by heat and ammonia, i.e., stressors pertinent to the biotechnological ecosystem of this methanogen that plays a key role in anaerobic...... thermoautotrophicum Delta H, from another genus, in which trkA is not part of the locus. The proteins encoded in the TM-1 genes are very similar to the S-6 homologs, but considerably less similar to the Delta H proteins. The TM-1 Hsp70(DnaK) protein has the 23-amino acid deletion-by comparison with homologs from Gram...

  5. Genome sequence of the tsetse fly (Glossina morsitans ): Vector of African trypanosomiasis

    KAUST Repository

    Watanabe, Junichi

    2014-04-24

    Tsetse flies are the sole vectors of human African trypanosomiasis throughout sub-Saharan Africa. Both sexes of adult tsetse feed exclusively on blood and contribute to disease transmission. Notable differences between tsetse and other disease vectors include obligate microbial symbioses, viviparous reproduction, and lactation. Here, we describe the sequence and annotation of the 366-megabase Glossina morsitans morsitans genome. Analysis of the genome and the 12,308 predicted protein-encoding genes led to multiple discoveries, including chromosomal integrations of bacterial (Wolbachia) genome sequences, a family of lactation-specific proteins, reduced complement of host pathogen recognition proteins, and reduced olfaction/chemosensory associated genes. These genome data provide a foundation for research into trypanosomiasis prevention and yield important insights with broad implications for multiple aspects of tsetse biology.

  6. Crystal structure of Homo sapiens protein LOC79017

    Energy Technology Data Exchange (ETDEWEB)

    Bae, Euiyoung; Bingman, Craig A.; Aceti, David J.; Phillips, Jr., George N. (UW)

    2010-02-08

    LOC79017 (MW 21.0 kDa, residues 1-188) was annotated as a hypothetical protein encoded by Homo sapiens chromosome 7 open reading frame 24. It was selected as a target by the Center for Eukaryotic Structural Genomics (CESG) because it did not share more than 30% sequence identity with any protein for which the three-dimensional structure is known. The biological function of the protein has not been established yet. Parts of LOC79017 were identified as members of uncharacterized Pfam families (residues 1-95 as PB006073 and residues 104-180 as PB031696). BLAST searches revealed homologues of LOC79017 in many eukaryotes, but none of them have been functionally characterized. Here, we report the crystal structure of H. sapiens protein LOC79017 (UniGene code Hs.530024, UniProt code O75223, CESG target number go.35223).

  7. Mutations in Cas9 Enhance the Rate of Acquisition of Viral Spacer Sequences during the CRISPR-Cas Immune Response.

    Science.gov (United States)

    Heler, Robert; Wright, Addison V; Vucelja, Marija; Bikard, David; Doudna, Jennifer A; Marraffini, Luciano A

    2017-01-05

    CRISPR loci and their associated (Cas) proteins encode a prokaryotic immune system that protects against viruses and plasmids. Upon infection, a low fraction of cells acquire short DNA sequences from the invader. These sequences (spacers) are integrated in between the repeats of the CRISPR locus and immunize the host against the matching invader. Spacers specify the targets of the CRISPR immune response through transcription into short RNA guides that direct Cas nucleases to the invading DNA molecules. Here we performed random mutagenesis of the RNA-guided Cas9 nuclease to look for variants that provide enhanced immunity against viral infection. We identified a mutation, I473F, that increases the rate of spacer acquisition by more than two orders of magnitude. Our results highlight the role of Cas9 during CRISPR immunization and provide a useful tool to study this rare process and develop it as a biotechnological application. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Crystallization and preliminary crystallographic studies of FoxE from Rhodobacter ferrooxidans SW2, an FeII oxidoreductase involved in photoferrotrophy

    International Nuclear Information System (INIS)

    Pereira, L.; Saraiva, I. H.; Coelho, R.; Newman, D. K.; Louro, R. O.; Frazão, C.

    2012-01-01

    Crystals of the R. ferrooxidans SW2 iron oxidoreductase FoxE were obtained and the phase problem was solved by Fe SAD at 2.44 Å resolution. FoxE is a protein encoded by the foxEYZ operon of Rhodobacter ferrooxidans SW2 that is involved in Fe II -based anoxygenic photosynthesis (‘photoferrotrophy’). It is thought to reside in the periplasm, where it stimulates light-dependent Fe II oxidation. It contains 259 residues, including two haem c-binding motifs. As no three-dimensional model is available and there is no structure with a similar sequence, crystals of FoxE were produced. They diffracted to 2.44 Å resolution using synchrotron radiation at the Fe edge. The phase problem was solved by SAD using SHELXC/D/E and the experimental maps confirmed the presence of two haems per molecule

  9. Identification of T1D susceptibility genes within the MHC region by combining protein interaction networks and SNP genotyping data

    DEFF Research Database (Denmark)

    Brorsson, C.; Hansen, Niclas Tue; Hansen, Kasper Lage

    2009-01-01

    genes. We have developed a novel method that combines single nucleotide polymorphism (SNP) genotyping data with protein-protein interaction (ppi) networks to identify disease-associated network modules enriched for proteins encoded from the MHC region. Approximately 2500 SNPs located in the 4 Mb MHC......To develop novel methods for identifying new genes that contribute to the risk of developing type 1 diabetes within the Major Histocompatibility Complex (MHC) region on chromosome 6, independently of the known linkage disequilibrium (LD) between human leucocyte antigen (HLA)-DRB1, -DQA1, -DQB1...... region were analysed in 1000 affected offspring trios generated by the Type 1 Diabetes Genetics Consortium (T1DGC). The most associated SNP in each gene was chosen and genes were mapped to ppi networks for identification of interaction partners. The association testing and resulting interacting protein...

  10. Candidate chromosome 1 disease susceptibility genes for Sjogren’s syndrome xerostomia are narrowed by novel NOD.B10 congenic mice

    Science.gov (United States)

    Mongini, Patricia K. A.; Kramer, Jill M.; Ishikawa, Tomo-o; Herschman, Harvey; Esposito, Donna

    2014-01-01

    Sjogren’s syndrome (SS) is characterized by salivary gland leukocytic infiltrates and impaired salivation (xerostomia). Cox-2 (Ptgs2) is located on chromosome 1 within the span of the Aec2 region. In an attempt to demonstrate that COX-2 drives antibody-dependent hyposalivation, NOD.B10 congenic mice bearing a Cox-2flox gene were generated. A congenic line with non-NOD alleles in Cox-2-flanking genes failed manifest xerostomia. Further backcrossing yielded disease-susceptible NOD.B10 Cox-2flox lines; fine genetic mapping determined that critical Aec2 genes lie within a 1.56 to 2.17 Mb span of DNA downstream of Cox-2. Bioinformatics analysis revealed that susceptible and non-susceptible lines exhibit non-synonymous coding SNPs in 8 protein-encoding genes of this region, thereby better delineating candidate Aec2 alleles needed for SS xerostomia. PMID:24685748

  11. Identification and characterization of the grape WRKY family.

    Science.gov (United States)

    Zhang, Ying; Feng, Jian Can

    2014-01-01

    WRKY transcription factors have functions in plant growth and development and in response to biotic and abiotic stresses. Many studies have focused on functional identification of WRKY transcription factors, but little is known about the molecular phylogeny or global expression patterns of the complete WRKY family. In this study, we identified 80 WRKY proteins encoded in the grape genome. Based on the structural features of these proteins, the grape WRKY genes were classified into three groups (groups 1-3). Analysis of WRKY genes expression profiles indicated that 28 WRKY genes were differentially expressed in response to biotic stress caused by grape whiterot and/or salicylic acid (SA). In that 16 WRKY genes upregulated both by whiterot pathogenic bacteria and SA. The results indicated that 16 WRKY proteins participated in SA-dependent defense signal pathway. This study provides a basis for cloning genes with specific functions from grape.

  12. Identification and Characterization of the Grape WRKY Family

    Directory of Open Access Journals (Sweden)

    Ying Zhang

    2014-01-01

    Full Text Available WRKY transcription factors have functions in plant growth and development and in response to biotic and abiotic stresses. Many studies have focused on functional identification of WRKY transcription factors, but little is known about the molecular phylogeny or global expression patterns of the complete WRKY family. In this study, we identified 80 WRKY proteins encoded in the grape genome. Based on the structural features of these proteins, the grape WRKY genes were classified into three groups (groups 1–3. Analysis of WRKY genes expression profiles indicated that 28 WRKY genes were differentially expressed in response to biotic stress caused by grape whiterot and/or salicylic acid (SA. In that 16 WRKY genes upregulated both by whiterot pathogenic bacteria and SA. The results indicated that 16 WRKY proteins participated in SA-dependent defense signal pathway. This study provides a basis for cloning genes with specific functions from grape.

  13. The Polycomb group proteins bind throughout the INK4A-ARF locus and are disassociated in senescent cells

    DEFF Research Database (Denmark)

    Bracken, Adrian P; Kleine-Kohlbrecher, Daniela; Dietrich, Nikolaj

    2007-01-01

    The p16INK4A and p14ARF proteins, encoded by the INK4A-ARF locus, are key regulators of cellular senescence, yet the mechanisms triggering their up-regulation are not well understood. Here, we show that the ability of the oncogene BMI1 to repress the INK4A-ARF locus requires its direct association...... and is dependent on the continued presence of the EZH2-containing Polycomb-Repressive Complex 2 (PRC2) complex. Significantly, EZH2 is down-regulated in stressed and senescing populations of cells, coinciding with decreased levels of associated H3K27me3, displacement of BMI1, and activation of transcription...

  14. Direct Pathogenic Effects of HERV-encoded Proteins

    DEFF Research Database (Denmark)

    Hansen, Dorte Tranberg; Møller-Larsen, Anné; Petersen, Thor

    Background: Multiple sclerosis (MS) is a demyelinating, inflammatory disease of the central nervous system (CNS). MS is mediated by the immune system but the etiology of the disease remains unknown. Retroviral envelope (Env) proteins, encoded by human endogenous retroviruses (HERVs), are expressed...... in increased amounts on B cells from MS patients. Furthermore, the amount of anti-HERV antibodies in serum and cerebrospinal fluid from patients with MS is increased when compared with healthy controls. Aim: The overall aim of this project is to investigate the potential role of HERVs in the development of MS...... and the possible direct pathogenic effects of HERV-encoded Env proteins on the CNS. Methods: Construction and characterization of a panel of recombinant Env-proteins is initiated and their pathogenic potential will be investigated: Fusiogenic potential analyzed by flow cytometry and confocal microscopy. Analysis...

  15. Conventional and real time RT-PCR assays for the detection and differentiation of variant rabbit hemorrhagic disease virus (RHDVb) and its recombinants.

    Science.gov (United States)

    Dalton, K P; Arnal, J L; Benito, A A; Chacón, G; Martín Alonso, J M; Parra, F

    2018-01-01

    Since its emergence, variant RHDV (RHDVb/RHDV2) has spread throughout the Iberian Peninsula aided by the apparent lack of cross protection provided by classic (genogroup 1; G1) strain derived vaccines. In addition to RHDVb, full-length genome sequencing of RHDV strains has recently revealed the circulation of recombinant viruses on the Iberian Peninsula. These recombinant viruses contain the RHDVb structural protein encoding sequences and the non-structural coding regions of either pathogenic RHDV-G1 strains or non-pathogenic (np) rabbit caliciviruses. The aim of the work was twofold: firstly to validate a diagnostic real time RT-PCR developed in 2012 for the detection of RHDVb strains and secondly, to design a conventional RT-PCR for the differentiation of RHDVb strains from RHDVb recombinants by subsequent sequencing of the amplicon. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. Expression dynamics and ultrastructural localization of epitope-tagged Abutilon mosaic virus nuclear shuttle and movement proteins in Nicotiana benthamiana cells

    International Nuclear Information System (INIS)

    Kleinow, Tatjana; Tanwir, Fariha; Kocher, Cornelia; Krenz, Bjoern; Wege, Christina; Jeske, Holger

    2009-01-01

    The geminivirus Abutilon mosaic virus (AbMV) encodes two proteins which are essential for viral spread within plants. The nuclear shuttle protein (NSP) transfers viral DNA between the nucleus and cytoplasm, whereas the movement protein (MP) facilitates transport between cells through plasmodesmata and long-distance via phloem. An inducible overexpression system for epitope-tagged NSP and MP in plants yielded unprecedented amounts of both proteins. Western blots revealed extensive posttranslational modification and truncation for MP, but not for NSP. Ultrastructural examination of Nicotiana benthamiana tissues showed characteristic nucleopathic alterations, including fibrillar rings, when epitope-tagged NSP and MP were simultaneously expressed in leaves locally infected with an AbMV DNA A in which the coat protein gene was replaced by a green fluorescent protein encoding gene. Immunogold labelling localized NSP in the nucleoplasm and in the fibrillar rings. MP appeared at the cell periphery, probably the plasma membrane, and plasmodesmata.

  17. Architecture of the large subunit of the mammalian mitochondrial ribosome.

    Science.gov (United States)

    Greber, Basil J; Boehringer, Daniel; Leitner, Alexander; Bieri, Philipp; Voigts-Hoffmann, Felix; Erzberger, Jan P; Leibundgut, Marc; Aebersold, Ruedi; Ban, Nenad

    2014-01-23

    Mitochondrial ribosomes synthesize a number of highly hydrophobic proteins encoded on the genome of mitochondria, the organelles in eukaryotic cells that are responsible for energy conversion by oxidative phosphorylation. The ribosomes in mammalian mitochondria have undergone massive structural changes throughout their evolution, including ribosomal RNA shortening and acquisition of mitochondria-specific ribosomal proteins. Here we present the three-dimensional structure of the 39S large subunit of the porcine mitochondrial ribosome determined by cryo-electron microscopy at 4.9 Å resolution. The structure, combined with data from chemical crosslinking and mass spectrometry experiments, reveals the unique features of the 39S subunit at near-atomic resolution and provides detailed insight into the architecture of the polypeptide exit site. This region of the mitochondrial ribosome has been considerably remodelled compared to its bacterial counterpart, providing a specialized platform for the synthesis and membrane insertion of the highly hydrophobic protein components of the respiratory chain.

  18. Partial sequence determination of metabolically labeled radioactive proteins and peptides

    International Nuclear Information System (INIS)

    Anderson, C.W.

    1982-01-01

    The author has used the sequence analysis of radioactive proteins and peptides to approach several problems during the past few years. They, in collaboration with others, have mapped precisely several adenovirus proteins with respect to the nucleotide sequence of the adenovirus genome; identified hitherto missed proteins encoded by bacteriophage MS2 and by simian virus 40; analyzed the aminoterminal maturation of several virus proteins; determined the cleavage sites for processing of the poliovirus polyprotein; and analyzed the mechanism of frameshifting by excess normal tRNAs during cell-free protein synthesis. This chapter is designed to aid those without prior experience at protein sequence determinations. It is based primarily on the experience gained in the studies cited above, which made use of the Beckman 890 series automated protein sequencers

  19. FERM proteins in animal morphogenesis.

    Science.gov (United States)

    Tepass, Ulrich

    2009-08-01

    Proteins containing a FERM domain are ubiquitous components of the cytocortex of animal cells where they are engaged in structural, transport, and signaling functions. Recent years have seen a wealth of genetic studies in model organisms that explore FERM protein function in development and tissue organization. In addition, mutations in several FERM protein-encoding genes have been associated with human diseases. This review will provide a brief overview of the FERM domain structure and the FERM protein superfamily and then discuss recent advances in our understanding of the mechanism of function and developmental requirement of several FERM proteins including Moesin, Myosin-VIIA, Myosin-XV, Coracle/Band4.1 as well as Yurt and its vertebrate homologs Mosaic Eyes and EPB41L5/YMO1/Limulus.

  20. Recent discoveries in the molecular pathogenesis of the inherited bone marrow failure syndrome Fanconi anemia.

    Science.gov (United States)

    Mamrak, Nicholas E; Shimamura, Akiko; Howlett, Niall G

    2017-05-01

    Fanconi anemia (FA) is a rare autosomal and X-linked genetic disease characterized by congenital abnormalities, progressive bone marrow failure (BMF), and increased cancer risk during early adulthood. The median lifespan for FA patients is approximately 33years. The proteins encoded by the FA genes function together in the FA-BRCA pathway to repair DNA damage and to maintain genome stability. Within the past two years, five new FA genes have been identified-RAD51/FANCR, BRCA1/FANCS, UBE2T/FANCT, XRCC2/FANCU, and REV7/FANCV-bringing the total number of disease-causing genes to 21. This review summarizes the discovery of these new FA genes and describes how these proteins integrate into the FA-BRCA pathway to maintain genome stability and critically prevent early-onset BMF and cancer. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Molecular characterization of two prunus necrotic ringspot virus isolates from Canada.

    Science.gov (United States)

    Cui, Hongguang; Hong, Ni; Wang, Guoping; Wang, Aiming

    2012-05-01

    We determined the entire RNA1, 2 and 3 sequences of two prunus necrotic ringspot virus (PNRSV) isolates, Chr3 from cherry and Pch12 from peach, obtained from an orchard in the Niagara Fruit Belt, Canada. The RNA1, 2 and 3 of the two isolates share nucleotide sequence identities of 98.6%, 98.4% and 94.5%, respectively. Their RNA1- and 2-encoded amino acid sequences are about 98% identical to the corresponding sequences of a cherry isolate, CH57, the only other PNRSV isolate with complete RNA1 and 2 sequences available. Phylogenetic analysis of the coat protein and movement protein encoded by RNA3 of Pch12 and Chr3 and published PNRSV isolates indicated that Chr3 belongs to the PV96 group and Pch12 belongs to the PV32 group.

  2. Differentiation of mycoplasmalike organisms (MLOs) in European fruit trees by PCR using specific primers derived from the sequence of a chromosomal fragment of the apple proliferation MLO.

    Science.gov (United States)

    Jarausch, W; Saillard, C; Dosba, F; Bové, J M

    1994-01-01

    A 1.8-kb chromosomal DNA fragment of the mycoplasmalike organism (MLO) associated with apple proliferation was sequenced. Three putative open reading frames were observed on this fragment. The protein encoded by open reading frame 2 shows significant homologies with bacterial nitroreductases. From the nucleotide sequence four primer pairs for PCR were chosen to specifically amplify DNA from MLOs associated with European diseases of fruit trees. Primer pairs specific for (i) Malus-affecting MLOs, (ii) Malus- and Prunus-affecting MLOs, and (iii) Malus-, Prunus-, and Pyrus-affecting MLOs were obtained. Restriction enzyme analysis of the amplification products revealed restriction fragment length polymorphisms between Malus-, Prunus, and Pyrus-affecting MLOs as well as between different isolates of the apple proliferation MLO. No amplification with either primer pair could be obtained with DNA from 12 different MLOs experimentally maintained in periwinkle. Images PMID:7916180

  3. Self-assembling peptide and protein nanodiscs for studies of membrane proteins

    DEFF Research Database (Denmark)

    Midtgaard, Søren Roi

    Particles containing both lipids and proteins (so-called lipoproteins) are vital to study. They are selfassembling particles that, in the human body, are responsible for the transport of lipids and cholesterol. Due to the increasing problems of obesity and related illnesses in the world, obtaining...... more knowledge about the cholesterol and lipid metabolism is paramount. As an example, in 2012, cardiovascular disease was still the main cause of death in the U.S. This means that the study of lipoproteins is not only of pure academic interest but vital to current world problems. Another reason...... of proteins encoded by the human genome. G-protein coupled receptors mediate the majority of hormone and neurotransmitter signals as well as being responsible for perception of light, smell and taste in the human body, and a number of Nobel prizes has been awarded based on their study. Structural...

  4. S. pombe placed on the prion map

    Directory of Open Access Journals (Sweden)

    Jacqueline Hayles

    2017-02-01

    Full Text Available Schizosaccharomyces pombe has been used extensively as a model organism, however it is only recently that the first prion in this organism, a copper transporter protein encoded by ctr4, has been conclusively demonstrated. Prions are found in a wide range of organisms and have been implicated in a number of human neurodegenerative diseases. Research into the biology of prions has been carried out mainly in the budding yeast Saccharomyces cerevisiae, however there are many questions still to be addressed. Now, with the identification of the Ctr4 prion in S. pombe, further work in the two yeasts and comparisons of prion biology in these organisms should lead to a greater understanding of prions and their role in disease.

  5. Culicoides variipennis (Diptera: Ceratopogonidae) complex in California.

    Science.gov (United States)

    Holbrook, F R; Tabachnick, W J

    1995-07-01

    Genetic relationships were examined among 24 collections, representing 23 populations of Culicoides variipennis (Coquillett) using isozyme electrophoresis of 11 protein encoding loci. The populations were collected from alkaline or fresh water larval habitats in California. Distance analysis demonstrated that C. v. occidentalis Wirth and Jones and C. v. sonorensis Wirth and Jones are genetically distinct. All C. v. occidentalis were geographically isolated from each other in highly alkaline or saline larval habitats, whereas C. v. sonorensis populations were collected from artificial freshwater habitats that were polluted with organic wastes. Higher levels of gene flow were found between C. v. sonorensis populations than from C. v. sonorensis populations to nearby C. v. occidentalis populations, indicative of genetic isolation between subspecies. Northern California C. v. sonorensis were genetically distinguishable from southern California C. v. sonorensis. The relationship between this variation and bluetongue disease epidemiology in California is discussed.

  6. Crystallization and preliminary X-ray analysis of Ebola VP35 interferon inhibitory domain mutant proteins

    International Nuclear Information System (INIS)

    Leung, Daisy W.; Borek, Dominika; Farahbakhsh, Mina; Ramanan, Parameshwaran; Nix, Jay C.; Wang, Tianjiao; Prins, Kathleen C.; Otwinowski, Zbyszek; Honzatko, Richard B.; Helgeson, Luke A.; Basler, Christopher F.; Amarasinghe, Gaya K.

    2010-01-01

    Three mutant forms of Ebola VP35 interferon inhibitory domain were crystallized in three different space groups. VP35 is one of seven structural proteins encoded by the Ebola viral genome and mediates viral replication, nucleocapsid formation and host immune suppression. The C-terminal interferon inhibitory domain (IID) of VP35 is critical for dsRNA binding and interferon inhibition. The wild-type VP35 IID structure revealed several conserved residues that are important for dsRNA binding and interferon antagonism. Here, the expression, purification and crystallization of recombinant Zaire Ebola VP35 IID mutants R312A, K319A/R322A and K339A in space groups P6 1 22, P2 1 2 1 2 1 and P2 1 , respectively, are described. Diffraction data were collected using synchrotron sources at the Advanced Light Source and the Advanced Photon Source

  7. Euglena in time: Evolution, control of central metabolic processes and multi-domain proteins in carbohydrate and natural product biochemistry

    Directory of Open Access Journals (Sweden)

    Ellis C. O’Neill

    2015-12-01

    Full Text Available Euglena gracilis is a eukaryotic microalgae that has been the subject of scientific study for hundreds of years. It has a complex evolutionary history, with traces of at least four endosymbiotic genomes and extensive horizontal gene transfer. Given the importance of Euglena in terms of evolutionary cell biology and its unique taxonomic position, we initiated a de novo transcriptome sequencing project in order to understand this intriguing organism. By analysing the proteins encoded in this transcriptome, we can identify an extremely complex metabolic capacity, rivalling that of multicellular organisms. Many genes have been acquired from what are now very distantly related species. Herein we consider the biology of Euglena in different time frames, from evolution through control of cell biology to metabolic processes associated with carbohydrate and natural products biochemistry.

  8. New technologies accelerate the exploration of non-coding RNAs in horticultural plants

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Degao; Mewalal, Ritesh; Hu, Rongbin; Tuskan, Gerald A.; Yang, Xiaohan

    2017-07-05

    Non-coding RNAs (ncRNAs), that is, RNAs not translated into proteins, are crucial regulators of a variety of biological processes in plants. While protein-encoding genes have been relatively well-annotated in sequenced genomes, accounting for a small portion of the genome space in plants, the universe of plant ncRNAs is rapidly expanding. Recent advances in experimental and computational technologies have generated a great momentum for discovery and functional characterization of ncRNAs. Here we summarize the classification and known biological functions of plant ncRNAs, review the application of next-generation sequencing (NGS) technology and ribosome profiling technology to ncRNA discovery in horticultural plants and discuss the application of new technologies, especially the new genome-editing tool clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) systems, to functional characterization of plant ncRNAs.

  9. Zea mI, the maize homolog of the allergen-encoding Lol pI gene of rye grass.

    Science.gov (United States)

    Broadwater, A H; Rubinstein, A L; Chay, C H; Klapper, D G; Bedinger, P A

    1993-09-15

    Sequence analysis of a pollen-specific cDNA from maize has identified a homolog (Zea mI) of the gene (Lol pI) encoding the major allergen of rye-grass pollen. The protein encoded by the partial cDNA sequence is 59.3% identical and 72.7% similar to the comparable region of the reported amino acid sequence of Lol pIA. Southern analysis indicates that this cDNA represents a member of a small multigene family in maize. Northern analysis shows expression only in pollen, not in vegetative or female floral tissues. The timing of expression is developmentally regulated, occurring at a low level prior to the first pollen mitosis and at a high level after this postmeiotic division. Western analysis detects a protein in maize pollen lysates using polyclonal antiserum and monoclonal antibodies directed against purified Lolium perenne allergen.

  10. Complexity and diversity of the NKR-P1:Clr (Klrb1:Clec2 recognition systems

    Directory of Open Access Journals (Sweden)

    Christina L Kirkham

    2014-06-01

    Full Text Available The NKR-P1 receptors were identified as prototypical natural killer (NK cell surface antigens and later shown to be conserved from rodents to humans on NK cells and subsets of T cells. C-type lectin-like in nature, they were originally shown to be capable of activating NK cell function and to recognize ligands on tumour cells. However, certain family members have subsequently been shown to be capable of inhibiting NK cell activity, and to recognize proteins encoded by a family of genetically linked C-type lectin-related (Clr ligands. Some of these ligands are expressed by normal, healthy cells, and modulated during transformation, infection, and cellular stress, while other ligands are upregulated during the immune response and during pathological circumstances. Here, we discuss historical and recent developments in NKR-P1 biology that demonstrate this NK receptor-ligand system to be far more complex and diverse than originally anticipated.

  11. Bioinformatics decoding the genome

    CERN Multimedia

    CERN. Geneva; Deutsch, Sam; Michielin, Olivier; Thomas, Arthur; Descombes, Patrick

    2006-01-01

    Extracting the fundamental genomic sequence from the DNA From Genome to Sequence : Biology in the early 21st century has been radically transformed by the availability of the full genome sequences of an ever increasing number of life forms, from bacteria to major crop plants and to humans. The lecture will concentrate on the computational challenges associated with the production, storage and analysis of genome sequence data, with an emphasis on mammalian genomes. The quality and usability of genome sequences is increasingly conditioned by the careful integration of strategies for data collection and computational analysis, from the construction of maps and libraries to the assembly of raw data into sequence contigs and chromosome-sized scaffolds. Once the sequence is assembled, a major challenge is the mapping of biologically relevant information onto this sequence: promoters, introns and exons of protein-encoding genes, regulatory elements, functional RNAs, pseudogenes, transposons, etc. The methodological ...

  12. CRISPR-based immune systems of the Sulfolobales: complexity and diversity

    DEFF Research Database (Denmark)

    Garrett, Roger Antony; Shah, Shiraz Ali; Vestergaard, Gisle Alberg

    2011-01-01

    CRISPR (cluster of regularly interspaced palindromic repeats)/Cas and CRISPR/Cmr systems of Sulfolobus, targeting DNA and RNA respectively of invading viruses or plasmids are complex and diverse. We address their classification and functional diversity, and the wide sequence diversity of RAMP...... (repeat-associated mysterious protein)-motif containing proteins encoded in Cmr modules. Factors influencing maintenance of partially impaired CRISPR-based systems are discussed. The capacity for whole CRISPR transcripts to be generated despite the uptake of transcription signals within spacer sequences...... is considered. Targeting of protospacer regions of invading elements by Cas protein-crRNA (CRISPR RNA) complexes exhibit relatively low sequence stringency, but the integrity of protospacer-associated motifs appears to be important. Different mechanisms for circumventing or inactivating the immune systems...

  13. Morganella psychrotolerans sp. nov., a histamine-producing bacterium isolated from various seafoods

    DEFF Research Database (Denmark)

    Emborg, Jette; Dalgaard, Paw; Ahrens, Peter

    2006-01-01

    Morganella morganii subsp. morganii (strain LMG 7874T) and Morganella morganii subsp. sibonii (strain DSM 14850T), respectively. Analysis of the 16S rRNA gene sequences showed a similarity of 98.6 % between mesophilic and psychrotolerant isolates. However, fragments of seven protein-encoding housekeeping...... genes (atpD, dnaN, gyrB, hdc, infB, rpoB and tuf) all showed less than 90.9 % sequence similarity between the two groups. The psychrotolerant isolates grew at 0-2 {degrees}C and also differed from the mesophilic M. morganii isolates with respect to growth at 37 {degrees}C and in 8.5 % (w/v) Na......Cl and fermentation of D-galactose. The psychrotolerant strains appear to represent a novel species, for which the name Morganella psychrotolerans sp. nov. is proposed. The type strain is U2/3T (=LMG 23374T=DSM 17886T)....

  14. First Case Report of Turcot Syndrome Type 1 in Colombia

    Directory of Open Access Journals (Sweden)

    Vallejo Dora

    2012-01-01

    Full Text Available Turcot syndrome is an autosomal recessive disorder clinically characterized by the occurrence of primary tumors of the central nervous system and adenomatous colonic polyps during the first or second decades of life, with a spectrum of clinical features such as “café-au-lait” spots, axillary freckling, and hyperpigmented spots. Currently its prevalence globally and in Colombia remains unknown. We present the case of a 20-year-old male with a clinical presentation of both glioblastoma multiforme and multiple adenomatous colonic polyps. The molecular genetics study revealed a mutation in KrasAsp12 gene and altered expression of HMSH2 and HMSH6 proteins encoded by the DNA mismatch repair genes in two of the colonic polyps. Even though this clinical presentation may suggest a shorter survival rate, this patient is still alive after seven months of treatment. A literature review complements this report.

  15. Molecular genetic analysis of cereal β-amylase genes using exon-primed intron-crossing (EPIC PCR

    Directory of Open Access Journals (Sweden)

    Stratula Olga

    2014-01-01

    Full Text Available The proteins encoded by cereal β-amylase genes Bamy1 and Bamy2 genes play an important role in seedling germination and in the brewing process. Here, we use exon-primed intron-crossing (EPIC to analyse Bamy1 and Bamy2 genetic diversity among 38 accessions belonging to six Poaceae tribes. DNA sequence alignment of multiple Poaceae species β-amylase sequences allowed design of EPIC primers that simultaneously amplify Bamy1 and Bamy2 in all the cereal species investigated. The genetic variation observed in the samples investigated is analysed and discussed, and illustrates the effectiveness of this approach for intra- and interspecific analysis in plant species.

  16. Characterization of the nuclear localization signal of the hepatitis delta virus antigen

    International Nuclear Information System (INIS)

    Alves, Carolina; Freitas, Natalia; Cunha, Celso

    2008-01-01

    The delta antigen (HDAg) is the only protein encoded by the hepatitis delta virus (HDV) RNA genome. The HDAg contains an RNA binding domain, a dimerization domain, and a nuclear localization signal (NLS). The nuclear import of HDV RNPs is thought to be one of the first tasks of the HDAg during the HDV replication cycle. Using c-myc-PK fusions with several regions of the HDAg in transfection assays in Huh7 cells, we found that the HDAg NLS consists of a single stretch of 10 amino acids, EGAPPAKRAR, located in positions 66-75. Deletion and mutation analysis of this region showed that both the acidic glutamic acid residue at position 66 and the basic arginine residue at position 75 are essential for promoting nuclear import

  17. Examination of the requirement for ucp-4, a putative homolog of mammalian uncoupling proteins, for stress tolerance and longevity in C. elegans.

    Science.gov (United States)

    Iser, Wendy B; Kim, Daemyung; Bachman, Eric; Wolkow, Catherine

    2005-10-01

    Reactive oxygen species (ROS) are generated by mitochondrial respiration and can react with and damage cellular components. According to the free radical theory of aging, oxidative damage from mitochondrial ROS is a major cause of cellular decline during aging. Mitochondrial uncoupling proteins (UCPs) uncouple ATP production from electron transport and can be stimulated by free radicals, suggesting UCPs may perform a cytoprotective function. The nematode, Caenorhabditis elegans, contains one UCP-like protein, encoded by the ucp-4 gene. We have investigated the genetic requirement for ucp-4 in normal aging and stress resistance. Consistent with the hypothesis that ucp-4 encodes a putative uncoupling protein, animals lacking ucp-4 function contained elevated ATP levels. However, the absence of ucp-4 function did not affect adult lifespan or survival in the presence of thermal or oxidative stress. Together, these results demonstrate that ucp-4 is a negative regulator of ATP production in C. elegans, but is not required for normal lifespan.

  18. Damaged mitochondria in Fanconi anemia - an isolated event or a general phenomenon?

    Science.gov (United States)

    Pagano, Giovanni; Shyamsunder, Pavithra; Verma, Rama S; Lyakhovich, Alex

    2014-01-01

    Fanconi anemia (FA) is known as an inherited bone marrow failure syndrome associated with cancer predisposition and susceptibility to a number of DNA damaging stimuli, along with a number of clinical features such as upper limb malformations, increased diabetes incidence and typical anomalies in skin pigmentation. The proteins encoded by FA-defective genes (FANC proteins) display well-established roles in DNA damage and repair pathways. Moreover, some independent studies have revealed that mitochondrial dysfunction (MDF) is also involved in FA phenotype. Unconfined to FA, we have shown that other syndromes featuring DNA damage and repair (such as ataxia-telangiectasia, AT, and Werner syndrome, WS) display MDF-related phenotypes, along with oxidative stress (OS) that, altogether, may play major roles in these diseases. Experimental and clinical studies are warranted in the prospect of future therapies to be focused on compounds scavenging reactive oxygen species (ROS) as well as protecting mitochondrial functions.

  19. IQCJ-SCHIP1, a novel fusion transcript encoding a calmodulin-binding IQ motif protein

    International Nuclear Information System (INIS)

    Kwasnicka-Crawford, Dorota A.; Carson, Andrew R.; Scherer, Stephen W.

    2006-01-01

    The existence of transcripts that span two adjacent, independent genes is considered rare in the human genome. This study characterizes a novel human fusion gene named IQCJ-SCHIP1. IQCJ-SCHIP1 is the longest isoform of a complex transcriptional unit that bridges two separate genes that encode distinct proteins, IQCJ, a novel IQ motif containing protein and SCHIP1, a schwannomin interacting protein that has been previously shown to interact with the Neurofibromatosis type 2 (NF2) protein. IQCJ-SCHIP1 is located on the chromosome 3q25 and comprises a 1692-bp transcript encompassing 11 exons spanning 828 kb of the genomic DNA. We show that IQCJ-SCHIP1 mRNA is highly expressed in the brain. Protein encoded by the IQCJ-SCHIP1 gene was localized to cytoplasm and actin-rich regions and in differentiated PC12 cells was also seen in neurite extensions

  20. Insulin and insulin-like growth factor receptors and responses

    International Nuclear Information System (INIS)

    Roth, R.A.; Steele-Perkins, G.; Hari, J.; Stover, C.; Pierce, S.; Turner, J.; Edman, J.C.; Rutter, W.J.

    1988-01-01

    Insulin is a member of a family of structurally related hormones with diverse physiological functions. In humans, the best-characterized members of this family include insulin, insulin-like growth factor (IGF)-I, and IGF-II. Each of these three polypeptide hormones has its own distinct receptor. The structures of each of these receptors have now been deduced from analyses of isolated cDNA clones. To study further the responses mediated through these three different receptors, the authors have been studying cells expressing the proteins encoded by these three cDNAs. The isolated cDNAs have been transfected into Chinese hamster ovary (CHO) cells, and the resulting transfected cell lines have been characterized as to the ligand-binding activities and signal-transducing activities of the expressed proteins

  1. Nucleotide sequence and genetic organization of Hungarian grapevine chrome mosaic nepovirus RNA2.

    Science.gov (United States)

    Brault, V; Hibrand, L; Candresse, T; Le Gall, O; Dunez, J

    1989-10-11

    The complete nucleotide sequence of hungarian grapevine chrome mosaic nepovirus (GCMV) RNA2 has been determined. The RNA sequence is 4441 nucleotides in length, excluding the poly(A) tail. A polyprotein of 1324 amino acids with a calculated molecular weight of 146 kDa is encoded in a single long open reading frame extending from nucleotides 218 to 4190. This polyprotein is homologous with the protein encoded by the S strain of tomato black ring virus (TBRV) RNA2, the only other nepovirus sequenced so far. Direct sequencing of the viral coat protein and in vitro translation of transcripts derived from cDNA sequences demonstrate that, as for comoviruses, the coat protein is located at the carboxy terminus of the polyprotein. A model for the expression of GCMV RNA2 is presented.

  2. How Next-Generation Sequencing Has Aided Our Understanding of the Sequence Composition and Origin of B Chromosomes

    Directory of Open Access Journals (Sweden)

    Alevtina Ruban

    2017-10-01

    Full Text Available Accessory, supernumerary, or—most simply—B chromosomes, are found in many eukaryotic karyotypes. These small chromosomes do not follow the usual pattern of segregation, but rather are transmitted in a higher than expected frequency. As increasingly being demonstrated by next-generation sequencing (NGS, their structure comprises fragments of standard (A chromosomes, although in some plant species, their sequence also includes contributions from organellar genomes. Transcriptomic analyses of various animal and plant species have revealed that, contrary to what used to be the common belief, some of the B chromosome DNA is protein-encoding. This review summarizes the progress in understanding B chromosome biology enabled by the application of next-generation sequencing technology and state-of-the-art bioinformatics. In particular, a contrast is drawn between a direct sequencing approach and a strategy based on a comparative genomics as alternative routes that can be taken towards the identification of B chromosome sequences.

  3. Genome sequence of the tsetse fly (Glossina morsitans ): Vector of African trypanosomiasis

    KAUST Repository

    Watanabe, Junichi; Hattori, Masahira; Berriman, Matthew; Lehane, Michael J.; Hall, Neil; Solano, Philippe; Aksoy, Serap; Hide, Winston; Touré , Yé ya Tié moko; Attardo, Geoffrey M.; Darby, Alistair Charles; Toyoda, Atsushi; Hertz-Fowler, Christiane; Larkin, Denis M.; Cotton, James A.; Sanders, Mandy J.; Swain, Martin T.; Quail, Michael A.; Inoue, Noboru; Ravel, Sophie; Taylor, Todd Duane; Srivastava, Tulika P.; Sharma, Vineet Kumar; Warren, Wesley C.; Wilson, Richard K.; Suzuki, Yutaka; Lawson, Daniel; Hughes, Daniel Seth Toney; Megy, Karyn; Masiga, Daniel K.; Mireji, Paul Odhiambo; Hansen, Immo Alex; Van Den Abbeele, Jan; Benoit, Joshua B.; Bourtzis, Kostas; Obiero, George F O; Robertson, Hugh M.; Jones, Jeffery W.; Zhou, Jingjiang; Field, Linda M.; Friedrich, Markus; Nyanjom, Steven R G; Telleria, Erich Loza; Caljon, Guy; Ribeiro, José M. C.; Acosta-Serrano, Alvaro; Ooi, Cherpheng; Rose, Clair; Price, David P.; Haines, Lee Rafuse; Christoffels, Alan G.; Sim, Cheolho; Pham, Daphne Q D; Denlinger, David L.; Geiser, Dawn L.; Omedo, Irene A.; Winzerling, Joy J.; Peyton, Justin T.; Marucha, Kevin K.; Jonas, Mario; Meuti, Megan E.; Rawlings, Neil David; Zhang, Qirui; Macharia, Rosaline Wanjiru; Michalkova, Veronika; Dashti, Zahra Jalali Sefid; Baumann, Aaron A.; Gä de, Gerd; Marco, Heather G.; Caers, Jelle; Schoofs, Liliane; Riehle, Michael A.; Hu, Wanqi; Tu, Zhijian; Tarone, Aaron M.; Malacrida, Anna Rodolfa; Kibet, Caleb K.; Scolari, Francesca; Koekemoer, J. J. O.; Willis, Judith H.; Gomulski, Ludvik M.; Falchetto, Marco; Scott, Maxwell J.; Fu, Shuhua; Sze, Singhoi; Luiz, Thiago; Weiss, Brian L.; Walshe, Deirdre P.; Wang, Jingwen; Wamalwa, Mark; Mwangi, Sarah; Ramphul, Urvashi N.; Snyder, Anna K.; Brelsfoard, Corey L.; Thomas, Gavin H.; Tsiamis, George; Arensburger, Peter; Rio, Rita V M; Macdonald, Sandy J.; Panji, Sumir; Kruger, Adele F.; Benkahla, Alia; Balyeidhusa, Apollo Simon Peter; Msangi, Atway R.; Okoro, Chinyere K.; Stephens, Dawn; Stanley, Eleanor J.; Mpondo, Feziwe; Wamwiri, Florence N.; Mramba, Furaha; Siwo, Geoffrey H.; Githinji, George; Harkins, Gordon William; Murilla, Grace Adira; Lehvä slaiho, Heikki; Malele, Imna I.; Auma, Joanna Eseri; Kinyua, Johnson K.; Ouma, Johnson O.; Okedi, Loyce M A; Manga, Lucien; Aslett, Martin A.; Koffi, Mathurin; Gaunt, Michael W.; Makgamathe, Mmule; Mulder, Nicola Jane; Manangwa, Oliver; Abila, Patrick P.; Wincker, Patrick; Gregory, Richard I.; Bateta, Rosemary; Sakate, Ryuichi; Ommeh, Sheila; Lehane, Stella M.; Imanishi, Tadashi; Osamor, Victor Chukwudi; Kawahara, Yoshihiro

    2014-01-01

    Tsetse flies are the sole vectors of human African trypanosomiasis throughout sub-Saharan Africa. Both sexes of adult tsetse feed exclusively on blood and contribute to disease transmission. Notable differences between tsetse and other disease vectors include obligate microbial symbioses, viviparous reproduction, and lactation. Here, we describe the sequence and annotation of the 366-megabase Glossina morsitans morsitans genome. Analysis of the genome and the 12,308 predicted protein-encoding genes led to multiple discoveries, including chromosomal integrations of bacterial (Wolbachia) genome sequences, a family of lactation-specific proteins, reduced complement of host pathogen recognition proteins, and reduced olfaction/chemosensory associated genes. These genome data provide a foundation for research into trypanosomiasis prevention and yield important insights with broad implications for multiple aspects of tsetse biology.

  4. Connective tissue growth factor (CTGF) and cancer progression.

    Science.gov (United States)

    Chu, Chia-Yu; Chang, Cheng-Chi; Prakash, Ekambaranellore; Kuo, Min-Liang

    2008-11-01

    Connective tissue growth factor (CTGF) is a member of the CCN family of secreted, matrix-associated proteins encoded by immediate early genes that play various roles in angiogenesis and tumor growth. CCN family proteins share uniform modular structure which mediates various cellular functions such as regulation of cell division, chemotaxis, apoptosis, adhesion, motility, angiogenesis, neoplastic transformation, and ion transport. Recently, CTGF expression has been shown to be associated with tumor development and progression. There is growing body of evidence that CTGF may regulate cancer cell migration, invasion, angiogenesis, and anoikis. In this review, we will highlight the influence of CTGF expression on the biological behavior and progression of various cancer cells, as well as its regulation on various types of protein signals and their mechanisms.

  5. [Replication of Streptomyces plasmids: the DNA nucleotide sequence of plasmid pSB 24.2].

    Science.gov (United States)

    Bolotin, A P; Sorokin, A V; Aleksandrov, N N; Danilenko, V N; Kozlov, Iu I

    1985-11-01

    The nucleotide sequence of DNA in plasmid pSB 24.2, a natural deletion derivative of plasmid pSB 24.1 isolated from S. cyanogenus was studied. The plasmid amounted by its size to 3706 nucleotide pairs. The G-C composition was equal to 73 per cent. The analysis of the DNA structure in plasmid pSB 24.2 revealed the protein-encoding sequence of DNA, the continuity of which was significant for replication of the plasmid containing more than 1300 nucleotide pairs. The analysis also revealed two A-T-rich areas of DNA, the G-C composition of which was less than 55 per cent and a DNA area with a branched pin structure. The results may be of value in investigation of plasmid replication in actinomycetes and experimental cloning of DNA with this plasmid as a vector.

  6. EasyClone: method for iterative chromosomal integration of multiple genes in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Jensen, Niels Bjerg; Strucko, Tomas; Kildegaard, Kanchana Rueksomtawin

    2014-01-01

    of multiple genes with an option of recycling selection markers. The vectors combine the advantage of efficient uracil excision reaction-based cloning and Cre-LoxP-mediated marker recycling system. The episomal and integrative vector sets were tested by inserting genes encoding cyan, yellow, and red...... fluorescent proteins into separate vectors and analyzing for co-expression of proteins by flow cytometry. Cells expressing genes encoding for the three fluorescent proteins from three integrations exhibited a much higher level of simultaneous expression than cells producing fluorescent proteins encoded...... on episomal plasmids, where correspondingly 95% and 6% of the cells were within a fluorescence interval of Log10 mean ± 15% for all three colors. We demonstrate that selective markers can be simultaneously removed using Cre-mediated recombination and all the integrated heterologous genes remain...

  7. Crystal structures of the methyltransferase and helicase from the ZIKA 1947 MR766 Uganda strain

    Energy Technology Data Exchange (ETDEWEB)

    Bukrejewska, Malgorzata; Derewenda, Urszula; Radwanska, Malwina; Engel, Daniel A.; Derewenda, Zygmunt S.

    2017-08-15

    Two nonstructural proteins encoded byZika virusstrain MR766 RNA, a methyltransferase and a helicase, were crystallized and their structures were solved and refined at 2.10 and 2.01 Å resolution, respectively. The NS5 methyltransferase contains a boundS-adenosyl-L-methionine (SAM) co-substrate. The NS3 helicase is in the apo form. Comparison with published crystal structures of the helicase in the apo, nucleotide-bound and single-stranded RNA (ssRNA)-bound states suggests that binding of ssRNA to the helicase may occur through conformational selection rather than induced fit.

  8. RASTtk: A modular and extensible implementation of the RAST algorithm for building custom annotation pipelines and annotating batches of genomes

    Energy Technology Data Exchange (ETDEWEB)

    Brettin, Thomas; Davis, James J.; Disz, Terry; Edwards, Robert A.; Gerdes, Svetlana; Olsen, Gary J.; Olson, Robert; Overbeek, Ross; Parrello, Bruce; Pusch, Gordon D.; Shukla, Maulik; Thomason, James A.; Stevens, Rick; Vonstein, Veronika; Wattam, Alice R.; Xia, Fangfang

    2015-02-10

    The RAST (Rapid Annotation using Subsystem Technology) annotation engine was built in 2008 to annotate bacterial and archaeal genomes. It works by offering a standard software pipeline for identifying genomic features (i.e., protein-encoding genes and RNA) and annotating their functions. Recently, in order to make RAST a more useful research tool and to keep pace with advancements in bioinformatics, it has become desirable to build a version of RAST that is both customizable and extensible. In this paper, we describe the RAST tool kit (RASTtk), a modular version of RAST that enables researchers to build custom annotation pipelines. RASTtk offers a choice of software for identifying and annotating genomic features as well as the ability to add custom features to an annotation job. RASTtk also accommodates the batch submission of genomes and the ability to customize annotation protocols for batch submissions. This is the first major software restructuring of RAST since its inception.

  9. Identification of Molecular and Cellular Responses of Desulfovibrio vulgaris Biofilms under Culture Conditions Relevant to Field Conditions for Bioreduction of Toxic Metals and Radionuclides

    Energy Technology Data Exchange (ETDEWEB)

    Judy D. Wall

    2011-06-09

    Our findings demonstrated that D. vulgaris surface-adhered populations produce extracellular structures, and that that the cells have altered carbon and energy flux compared to planktonic cells. Biofilms did not have greatly increased carbohydrate accumulation. Interestingly genes present on the native plasmid found in D. vulgaris Hildenborough were necessary for wild type biofilm formation. In addition, extracellular appendages dependent on functions or proteins encoded by flaG or fliA also contributed to biofilm formation. Studies with SRB biofilms have indicated that the reduction and precipitation of metals can occur within the biofilm matrix; however, little work has been done to elucidate the physiological state of surface-adhered cells during metal reduction (Cr6+, U6+) and how this process is affected by nutrient feed levels (i.e., the stimulant).

  10. First insight into the genome of an uncultivated crenarchaeote from soil

    DEFF Research Database (Denmark)

    Quaiser, Achim; Ochsenreiter, Torsten; Klenk, Hans-Peter

    2002-01-01

    RNA genes and of several protein encoding genes (e.g. DNA polymerase, FixAB, glycosyl transferase) confirmed the specific affiliation of the genomic fragment with the non-thermophilic clade of the crenarchaeota. Content and structure of the genomic fragment indicated that the archaea from soil differ......Molecular phylogenetic surveys based on the characterization of 16S rRNA genes have revealed that soil is an environment particularly rich in microbial diversity. A clade of crenarchaeota (archaea) has frequently been detected among many other novel lineages of uncultivated bacteria. In this study...... we have initiated a genomic approach for the characterization of uncultivated microorganisms from soil. We have developed a procedure based on a two-phase electrophoresis technique that allows the fast and reliable purification of concentrated and clonable, high molecular weight DNA. From this DNA we...

  11. A novel member of the split betaalphabeta fold: Solution structure of the hypothetical protein YML108W from Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Pineda-Lucena, Antonio; Liao, Jack; Cort, John R.; Yee, Adelinda; Kennedy, Michael A.; Edwards, Aled M.

    2003-05-01

    As part of the Northeast Structural Genomics Consortium pilot project focused on small eukaryotic proteins and protein domains, we have determined the NMR structure of the protein encoded by open reading frame YML108W from Saccharomyces cerevisiae. YML108W belongs to one of the numerous structural proteomics targets whose biological function is unknown. Moreover, this protein does not have sequence similarity to any other protein. The NMR structure of YML108W consists of a four-stranded b-sheet with strand order 2143 and two a-helices, with an overall topology of bbabba. Strand b1 runs parallel to b4, and b2:b1 and b4:b3 pairs are arranged in an antiparallel fashion. While this fold belongs to the split bab family, it appears to be unique among this family; it is a novel arrangement of secondary structure, thereby expanding the universe of protein folds

  12. The complete sequence of human chromosome 5

    Energy Technology Data Exchange (ETDEWEB)

    Schmutz, Jeremy; Martin, Joel; Terry, Astrid; Couronne, Olivier; Grimwood, Jane; Lowry, State; Gordon, Laurie A.; Scott, Duncan; Xie, Gary; Huang, Wayne; Hellsten, Uffe; Tran-Gyamfi, Mary; She, Xinwei; Prabhakar, Shyam; Aerts, Andrea; Altherr, Michael; Bajorek, Eva; Black, Stacey; Branscomb, Elbert; Caoile, Chenier; Challacombe, Jean F.; Chan, Yee Man; Denys, Mirian; Detter, Chris; Escobar, Julio; Flowers, Dave; Fotopulos, Dea; Glavina, Tijana; Gomez, Maria; Gonzales, Eidelyn; Goodstenin, David; Grigoriev, Igor; Groza, Matthew; Hammon, Nancy; Hawkins, Trevor; Haydu, Lauren; Israni, Sanjay; Jett, Jamie; Kadner, Kristen; Kimbal, Heather; Kobayashi, Arthur; Lopez, Frederick; Lou, Yunian; Martinez, Diego; Medina, Catherine; Morgan, Jenna; Nandkeshwar, Richard; Noonan, James P.; Pitluck, Sam; Pollard, Martin; Predki, Paul; Priest, James; Ramirez, Lucia; Rash, Sam; Retterer, James; Rodriguez, Alex; Rogers, Stephanie; Salamov, Asaf; Salazar, Angelica; Thayer, Nina; Tice, Hope; Tsai, Ming; Ustaszewska, Anna; Vo, Nu; Wheeler, Jeremy; Wu, Kevin; Yang, Joan; Dickson, Mark; Cheng, Jan-Fang; Eichler, Evan E.; Olsen, Anne; Pennacchio, Len A.; Rokhsar, Daniel S.; Richardson, Paul; Lucas, Susan M.; Myers, Richard M.; Rubin, Edward M.

    2004-04-15

    Chromosome 5 is one of the largest human chromosomes yet has one of the lowest gene densities. This is partially explained by numerous gene-poor regions that display a remarkable degree of noncoding and syntenic conservation with non-mammalian vertebrates, suggesting they are functionally constrained. In total, we compiled 177.7 million base pairs of highly accurate finished sequence containing 923 manually curated protein-encoding genes including the protocadherin and interleukin gene families and the first complete versions of each of the large chromosome 5 specific internal duplications. These duplications are very recent evolutionary events and play a likely mechanistic role, since deletions of these regions are the cause of debilitating disorders including spinal muscular atrophy (SMA).

  13. NATO Advanced Study Institute on Evolving Methods for Macromolecular Gystallography

    CERN Document Server

    Read, Randy J

    2007-01-01

    X-ray crystallography is the pre-eminent technique for visualizing the structures of macromolecules at atomic resolution. These structures are central to understanding the detailed mechanisms of biological processes, and to discovering novel therapeutics using a structure-based approach. As yet, structures are known for only a small fraction of the proteins encoded by human and pathogenic genomes. To counter the myriad modern threats of disease, there is an urgent need to determine the structures of the thousands of proteins whose structure and function remain unknown. This volume draws on the expertise of leaders in the field of macromolecular crystallography to illuminate the dramatic developments that are accelerating progress in structural biology. Their contributions span the range of techniques from crystallization through data collection, structure solution and analysis, and show how modern high-throughput methods are contributing to a deeper understanding of medical problems.

  14. Cloning, expression, and chromosome mapping of human galectin-7

    DEFF Research Database (Denmark)

    Madsen, Peder; Rasmussen, H H; Flint, T

    1995-01-01

    The galectins are a family of beta-galactoside-binding proteins implicated in modulating cell-cell and cell-matrix interactions. Here we report the cloning and expression of a novel member of this family (galectin-7) that correspond to IEF (isoelectric focusing) 17 (12,700 Da; pI, 7.6) in the human...... keratinocyte protein data base, and that is strikingly down-regulated in SV40 transformed keratinocytes (K14). The cDNA was cloned from a lambda gt11 cDNA expression library using degenerated oligodeoxyribonucleotides back-translated from an IEF 17 peptide sequence. The protein encoded by the galectin-7 clone......14 keratinocytes imply a role in cell-cell and/or cell-matrix interactions necessary for normal growth control. The galectin-7 gene was mapped to chromosome 19. Udgivelsesdato: 1995-Mar-17...

  15. Evolutionary relationship and structural characterization of the EPF/EPFL gene family.

    Directory of Open Access Journals (Sweden)

    Naoki Takata

    Full Text Available EPF1-EPF2 and EPFL9/Stomagen act antagonistically in regulating leaf stomatal density. The aim of this study was to elucidate the evolutionary functional divergence of EPF/EPFL family genes. Phylogenetic analyses showed that AtEPFL9/Stomagen-like genes are conserved only in vascular plants and are closely related to AtEPF1/EPF2-like genes. Modeling showed that EPF/EPFL peptides share a common 3D structure that is constituted of a scaffold and loop. Molecular dynamics simulation suggested that AtEPF1/EPF2-like peptides form an additional disulfide bond in their loop regions and show greater flexibility in these regions than AtEPFL9/Stomagen-like peptides. This study uncovered the evolutionary relationship and the conformational divergence of proteins encoded by the EPF/EPFL family genes.

  16. Evolutionary relationship and structural characterization of the EPF/EPFL gene family.

    Science.gov (United States)

    Takata, Naoki; Yokota, Kiyonobu; Ohki, Shinya; Mori, Masashi; Taniguchi, Toru; Kurita, Manabu

    2013-01-01

    EPF1-EPF2 and EPFL9/Stomagen act antagonistically in regulating leaf stomatal density. The aim of this study was to elucidate the evolutionary functional divergence of EPF/EPFL family genes. Phylogenetic analyses showed that AtEPFL9/Stomagen-like genes are conserved only in vascular plants and are closely related to AtEPF1/EPF2-like genes. Modeling showed that EPF/EPFL peptides share a common 3D structure that is constituted of a scaffold and loop. Molecular dynamics simulation suggested that AtEPF1/EPF2-like peptides form an additional disulfide bond in their loop regions and show greater flexibility in these regions than AtEPFL9/Stomagen-like peptides. This study uncovered the evolutionary relationship and the conformational divergence of proteins encoded by the EPF/EPFL family genes.

  17. Chatty Mitochondria: Keeping Balance in Cellular Protein Homeostasis.

    Science.gov (United States)

    Topf, Ulrike; Wrobel, Lidia; Chacinska, Agnieszka

    2016-08-01

    Mitochondria are multifunctional cellular organelles that host many biochemical pathways including oxidative phosphorylation (OXPHOS). Defective mitochondria pose a threat to cellular homeostasis and compensatory responses exist to curtail the source of stress and/or its consequences. The mitochondrial proteome comprises proteins encoded by the nuclear and mitochondrial genomes. Disturbances in protein homeostasis may originate from mistargeting of nuclear encoded mitochondrial proteins. Defective protein import and accumulation of mistargeted proteins leads to stress that triggers translation alterations and proteasomal activation. These cytosolic pathways are complementary to the mitochondrial unfolded protein response (UPRmt) that aims to increase the capacity of protein quality control mechanisms inside mitochondria. They constitute putative targets for interventions aimed at increasing the fitness, stress resistance, and longevity of cells and organisms. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Mechanistic basis of an epistatic interaction reducing age at onset in hereditary spastic paraplegia.

    Science.gov (United States)

    Newton, Timothy; Allison, Rachel; Edgar, James R; Lumb, Jennifer H; Rodger, Catherine E; Manna, Paul T; Rizo, Tania; Kohl, Zacharias; Nygren, Anders O H; Arning, Larissa; Schüle, Rebecca; Depienne, Christel; Goldberg, Lisa; Frahm, Christiane; Stevanin, Giovanni; Durr, Alexandra; Schöls, Ludger; Winner, Beate; Beetz, Christian; Reid, Evan

    2018-05-01

    Many genetic neurological disorders exhibit variable expression within affected families, often exemplified by variations in disease age at onset. Epistatic effects (i.e. effects of modifier genes on the disease gene) may underlie this variation, but the mechanistic basis for such epistatic interactions is rarely understood. Here we report a novel epistatic interaction between SPAST and the contiguous gene DPY30, which modifies age at onset in hereditary spastic paraplegia, a genetic axonopathy. We found that patients with hereditary spastic paraplegia caused by genomic deletions of SPAST that extended into DPY30 had a significantly younger age at onset. We show that, like spastin, the protein encoded by SPAST, the DPY30 protein controls endosomal tubule fission, traffic of mannose 6-phosphate receptors from endosomes to the Golgi, and lysosomal ultrastructural morphology. We propose that additive effects on this pathway explain the reduced age at onset of hereditary spastic paraplegia in patients who are haploinsufficient for both genes.

  19. RASTtk: a modular and extensible implementation of the RAST algorithm for building custom annotation pipelines and annotating batches of genomes.

    Science.gov (United States)

    Brettin, Thomas; Davis, James J; Disz, Terry; Edwards, Robert A; Gerdes, Svetlana; Olsen, Gary J; Olson, Robert; Overbeek, Ross; Parrello, Bruce; Pusch, Gordon D; Shukla, Maulik; Thomason, James A; Stevens, Rick; Vonstein, Veronika; Wattam, Alice R; Xia, Fangfang

    2015-02-10

    The RAST (Rapid Annotation using Subsystem Technology) annotation engine was built in 2008 to annotate bacterial and archaeal genomes. It works by offering a standard software pipeline for identifying genomic features (i.e., protein-encoding genes and RNA) and annotating their functions. Recently, in order to make RAST a more useful research tool and to keep pace with advancements in bioinformatics, it has become desirable to build a version of RAST that is both customizable and extensible. In this paper, we describe the RAST tool kit (RASTtk), a modular version of RAST that enables researchers to build custom annotation pipelines. RASTtk offers a choice of software for identifying and annotating genomic features as well as the ability to add custom features to an annotation job. RASTtk also accommodates the batch submission of genomes and the ability to customize annotation protocols for batch submissions. This is the first major software restructuring of RAST since its inception.

  20. Assessment of a DNA vaccine encoding an anchored-glycosylphosphatidylinositol tegumental antigen complexed to protamine sulphate on immunoprotection against murine schistosomiasis

    Directory of Open Access Journals (Sweden)

    Eduardo JM Nascimento

    2007-02-01

    Full Text Available Protamine sulphate/DNA complexes have been shown to protect DNA from DNase digestion in a lipid system for gene transfer. A DNA-based vaccine complexed to protamine sulphate was used to induce an immune response against Schistosoma mansoni anchored-glycosylphosphatidylinositol tegumental antigen in BALB/c mice. The protection elicited ranged from 33 to 44%. The spectrum of the elicited immune response induced by the vaccine formulation without protamine was characterized by a high level of IgG (IgG1> IgG2a. Protamine sulphate added to the DNA vaccine formulation retained the green fluorescent protein encoding-plasmid longer in muscle and spleen. The experiments in vivo showed that under protamine sulphate effect, the scope of protection remained unchanged, but a modulation in antibody production (IgG1= IgG2a was observed.

  1. Computational Analysis of Uncharacterized Proteins of Environmental Bacterial Genome

    Science.gov (United States)

    Coxe, K. J.; Kumar, M.

    2017-12-01

    Betaproteobacteria strain CB is a gram-negative bacterium in the phylum Proteobacteria and are found naturally in soil and water. In this complex environment, bacteria play a key role in efficiently eliminating the organic material and other pollutants from wastewater. To investigate the process of pollutant removal from wastewater using bacteria, it is important to characterize the proteins encoded by the bacterial genome. Our study combines a number of bioinformatics tools to predict the function of unassigned proteins in the bacterial genome. The genome of Betaproteobacteria strain CB contains 2,112 proteins in which function of 508 proteins are unknown, termed as uncharacterized proteins (UPs). The localization of the UPs with in the cell was determined and the structure of 38 UPs was accurately predicted. These UPs were predicted to belong to various classes of proteins such as enzymes, transporters, binding proteins, signal peptides, transmembrane proteins and other proteins. The outcome of this work will help better understand wastewater treatment mechanism.

  2. No Love Lost Between Viruses and Interferons.

    Science.gov (United States)

    Fensterl, Volker; Chattopadhyay, Saurabh; Sen, Ganes C

    2015-11-01

    The interferon system protects mammals against virus infections. There are several types of interferons, which are characterized by their ability to inhibit virus replication and resultant pathogenesis by triggering both innate and cell-mediated immune responses. Virus infection is sensed by a variety of cellular pattern-recognition receptors and triggers the synthesis of interferons, which are secreted by the infected cells. In uninfected cells, cell surface receptors recognize the secreted interferons and activate intracellular signaling pathways that induce the expression of interferon-stimulated genes; the proteins encoded by these genes inhibit different stages of virus replication. To avoid extinction, almost all viruses have evolved mechanisms to defend themselves against the interferon system. Consequently, a dynamic equilibrium of survival is established between the virus and its host, an equilibrium that can be shifted to the host's favor by the use of exogenous interferon as a therapeutic antiviral agent.

  3. Mutation Study of Two Thymidine Kinases 

    DEFF Research Database (Denmark)

    Skovgaard, Tine; Munch-Petersen, Birgitte; Eklund, Hans

    that phosphorylates all the natural deoxyribonucleosides and like insects, C. elegans only contains a single deoxyribonucleoside kinase-like gene. In contrast to the insects, however, the protein encoded by the elegans gene is 46 % identical to human TK1 (HuTK1) and have no homology to the insect kinase. Like HuTK1...... the C. elegans kinase (CeTK1) has thymidine as the preferred substrate, but it also displays activity with deoxyguanosine, though with high Km. A number of point mutations have been introduced in the active site of both the human and elegans TK's in order to change the substrate specificity away from...... not phosphorylate the anticancer analog 1-β-D-arabinofuranosylcytosine (AraC), however. The HuTK1 mutant has been crystallized, and azidothymidine monophosphate has been modelled into the active site....

  4. Recognitional specificity and evolution in the tomato-Cladosporium fulvum pathosystem.

    Science.gov (United States)

    Wulff, B B H; Chakrabarti, A; Jones, D A

    2009-10-01

    The interactions between plants and many biotrophic or hemibiotrophic pathogens are controlled by receptor proteins in the host and effector proteins delivered by the pathogen. Pathogen effectors facilitate pathogen growth through the suppression of host defenses and the manipulation of host metabolism, but recognition of a pathogen-effector protein by a host receptor enables the host to activate a suite of defense mechanisms that limit pathogen growth. In the tomato (Lycopersicon esculentum syn. Solanum lycopersicum)-Cladosporium fulvum (leaf mold fungus syn. Passalora fulva) pathosystem, the host receptors are plasma membrane-anchored, leucine-rich repeat, receptor-like proteins encoded by an array of Cf genes conferring resistance to C. fulvum. The pathogen effectors are mostly small, secreted, cysteine-rich, but otherwise largely dissimilar, extracellular proteins encoded by an array of avirulence (Avr) genes, so called because of their ability to trigger resistance and limit pathogen growth when the corresponding Cf gene is present in tomato. A number of Cf and Avr genes have been isolated, and details of the complex molecular interplay between tomato Cf proteins and C. fulvum effector proteins are beginning to emerge. Each effector appears to have a different role; probably most bind or modify different host proteins, but at least one has a passive role masking the pathogen. It is, therefore, not surprising that each effector is probably detected in a distinct and specific manner, some by direct binding, others as complexes with host proteins, and others via their modification of host proteins. The two papers accompanying this review contribute further to our understanding of the molecular specificity underlying effector perception by Cf proteins. This review, therefore, focuses on our current understanding of recognitional specificity in the tomato-C. fulvum pathosystem and highlights some of the critical questions that remain to be addressed. It also

  5. Canine distemper virus in the Serengeti ecosystem: molecular adaptation to different carnivore species.

    Science.gov (United States)

    Nikolin, Veljko M; Olarte-Castillo, Ximena A; Osterrieder, Nikolaus; Hofer, Heribert; Dubovi, Edward; Mazzoni, Camila J; Brunner, Edgar; Goller, Katja V; Fyumagwa, Robert D; Moehlman, Patricia D; Thierer, Dagmar; East, Marion L

    2017-04-01

    Was the 1993/1994 fatal canine distemper virus (CDV) epidemic in lions and spotted hyaenas in the Serengeti ecosystem caused by the recent spillover of a virulent domestic dog strain or one well adapted to these noncanids? We examine this question using sequence data from 13 'Serengeti' strains including five complete genomes obtained between 1993 and 2011. Phylogenetic and haplotype network analyses reveal that strains from noncanids during the epidemic were more closely related to each other than to those from domestic or wild canids. All noncanid 'Serengeti' strains during the epidemic encoded: (1) one novel substitution G134S in the CDV-V protein; and (2) the rare amino acid combination 519I/549H at two sites under positive selection in the region of the CDV-H protein that binds to SLAM (CD 150) host cell receptors. Worldwide, only a few noncanid strains in the America II lineage encode CDV-H 519I/549H. All canid 'Serengeti' strains during the epidemic coded CDV-V 134G, and CDV-H 519R/549Y, or 519R/549H. A functional assay of cell entry revealed the highest performance by CDV-H proteins encoding 519I/549H in cells expressing lion SLAM receptors, and the highest performance by proteins encoding 519R/549Y, typical of dog strains worldwide, in cells expressing dog SLAM receptors. Our findings are consistent with an epidemic in lions and hyaenas caused by CDV variants better adapted to noncanids than canids, but not with the recent spillover of a dog strain. Our study reveals a greater complexity of CDV molecular epidemiology in multihost environments than previously thought. © 2016 John Wiley & Sons Ltd.

  6. Comparative genomics of the type VI secretion systems of Pantoea and Erwinia species reveals the presence of putative effector islands that may be translocated by the VgrG and Hcp proteins

    Directory of Open Access Journals (Sweden)

    De Maayer Pieter

    2011-11-01

    Full Text Available Abstract Background The Type VI secretion apparatus is assembled by a conserved set of proteins encoded within a distinct locus. The putative effector proteins Hcp and VgrG are also encoded within these loci. We have identified numerous distinct Type VI secretion system (T6SS loci in the genomes of several ecologically diverse Pantoea and Erwinia species and detected the presence of putative effector islands associated with the hcp and vgrG genes. Results Between two and four T6SS loci occur among the Pantoea and Erwinia species. While two of the loci (T6SS-1 and T6SS-2 are well conserved among the various strains, the third (T6SS-3 locus is not universally distributed. Additional orthologous loci are present in Pantoea sp. aB-valens and Erwinia billingiae Eb661. Comparative analysis of the T6SS-1 and T6SS-3 loci showed non-conserved islands associated with the vgrG and hcp, and vgrG genes, respectively. These regions had a G+C content far lower than the conserved portions of the loci. Many of the proteins encoded within the hcp and vgrG islands carry conserved domains, which suggests they may serve as effector proteins for the T6SS. A number of the proteins also show homology to the C-terminal extensions of evolved VgrG proteins. Conclusions Extensive diversity was observed in the number and content of the T6SS loci among the Pantoea and Erwinia species. Genomic islands could be observed within some of T6SS loci, which are associated with the hcp and vgrG proteins and carry putative effector domain proteins. We propose new hypotheses concerning a role for these islands in the acquisition of T6SS effectors and the development of novel evolved VgrG and Hcp proteins.

  7. CEBPA exerts a specific and biologically important proapoptotic role in pancreatic β cells through its downstream network targets

    Science.gov (United States)

    Barbagallo, Davide; Condorelli, Angelo Giuseppe; Piro, Salvatore; Parrinello, Nunziatina; Fløyel, Tina; Ragusa, Marco; Rabuazzo, Agata Maria; Størling, Joachim; Purrello, Francesco; Di Pietro, Cinzia; Purrello, Michele

    2014-01-01

    Transcription factor CEBPA has been widely studied for its involvement in hematopoietic cell differentiation and causal role in hematological malignancies. We demonstrate here that it also performs a causal role in cytokine-induced apoptosis of pancreas β cells. Treatment of two mouse pancreatic α and β cell lines (αTC1-6 and βTC1) with proinflammatory cytokines IL-1β, IFN-γ, and TNF-α at doses that specifically induce apoptosis of βTC1 significantly increased the amount of mRNA and protein encoded by Cebpa and its proapoptotic targets, Arl6ip5 and Tnfrsf10b, in βTC1 but not in αTC1-6. Cebpa knockdown in βTC1 significantly decreased cytokine-induced apoptosis, together with the amount of Arl6ip5 and Tnfrsf10b. Analysis of the network comprising CEBPA, its targets, their first interactants, and proteins encoded by genes known to regulate cytokine-induced apoptosis in pancreatic β cells (genes from the apoptotic machinery and from MAPK and NFkB pathways) revealed that CEBPA, ARL6IP5, TNFRSF10B, TRAF2, and UBC are the top five central nodes. In silico analysis further suggests TRAF2 as trait d'union node between CEBPA and the NFkB pathway. Our results strongly suggest that Cebpa is a key regulator within the apoptotic network activated in pancreatic β cells during insulitis, and Arl6ip5, Tnfrsf10b, Traf2, and Ubc are key executioners of this program. PMID:24943845

  8. How to kill the honey bee larva: genomic potential and virulence mechanisms of Paenibacillus larvae.

    Directory of Open Access Journals (Sweden)

    Marvin Djukic

    Full Text Available Paenibacillus larvae, a Gram positive bacterial pathogen, causes American Foulbrood (AFB, which is the most serious infectious disease of honey bees. In order to investigate the genomic potential of P. larvae, two strains belonging to two different genotypes were sequenced and used for comparative genome analysis. The complete genome sequence of P. larvae strain DSM 25430 (genotype ERIC II consisted of 4,056,006 bp and harbored 3,928 predicted protein-encoding genes. The draft genome sequence of P. larvae strain DSM 25719 (genotype ERIC I comprised 4,579,589 bp and contained 4,868 protein-encoding genes. Both strains harbored a 9.7 kb plasmid and encoded a large number of virulence-associated proteins such as toxins and collagenases. In addition, genes encoding large multimodular enzymes producing nonribosomally peptides or polyketides were identified. In the genome of strain DSM 25719 seven toxin associated loci were identified and analyzed. Five of them encoded putatively functional toxins. The genome of strain DSM 25430 harbored several toxin loci that showed similarity to corresponding loci in the genome of strain DSM 25719, but were non-functional due to point mutations or disruption by transposases. Although both strains cause AFB, significant differences between the genomes were observed including genome size, number and composition of transposases, insertion elements, predicted phage regions, and strain-specific island-like regions. Transposases, integrases and recombinases are important drivers for genome plasticity. A total of 390 and 273 mobile elements were found in strain DSM 25430 and strain DSM 25719, respectively. Comparative genomics of both strains revealed acquisition of virulence factors by horizontal gene transfer and provided insights into evolution and pathogenicity.

  9. DNA repair in the c-myc proto-oncogene locus: Possible involvement in susceptibility or resistance to plasmacytoma induction in BALB/c mice

    International Nuclear Information System (INIS)

    Beecham, E.J.; Mushinski, J.F.; Shacter, E.; Potter, M.; Bohr, V.A.

    1991-01-01

    This report describes an unexpected difference in the efficiency of removal of UV-induced DNA damage in the c-myc locus in splenic B lymphoblasts from two inbred strains of mice. In cells from plasmacytoma-resistant DBA/2N mice, 35% of UV-induced damage in the regulatory and 5' flank of c-myc is removed by 12 h. However, in cells from plasmacytoma-susceptible BALB/cAn mice, damage is not removed from this region. In the protein-encoding region and 3' flank of c-myc as well as in two dihydrofolate reductase gene fragments, UV damage is repaired with similar efficiency in B lymphoblasts from both strains of mice. Furthermore, in the protein-encoding portion and 3' flank of c-myc, damage is selectively removed from only the transcribed strand. No repair is detected in the nontranscribed strand. In contrast, DNA repair in the 5' flank of c-myc is not strand specific; in DNA from DBA/2N cells, UV damage is rapidly removed from both the transcribed and nontranscribed strands. In BALB/cAn cells no repair was detected in either strand in the 5'flank, consistent with the results with double-stranded, nick-translated probes to this region of c-myc. In addition to the repair studies, we have detected post-UV-damage formation: in most of the genes studied, we find that additional T4 endonuclease-sensitive sites are formed in the DNA 2 h after irradiation. Our findings provide new insights into the details of gene-specific and strand-specific DNA repair and suggest that there may be close links between DNA repair and B-cell neoplastic development

  10. Biochemical and genetic analyses of the oomycete Pythium insidiosum provide new insights into clinical identification and urease-based evolution of metabolism-related traits

    Directory of Open Access Journals (Sweden)

    Theerapong Krajaejun

    2018-06-01

    Full Text Available The oomycete microorganism, Pythium insidiosum, causes the life-threatening infectious condition, pythiosis, in humans and animals worldwide. Affected individuals typically endure surgical removal of the infected organ(s. Detection of P. insidiosum by the established microbiological, immunological, or molecular methods is not feasible in non-reference laboratories, resulting in delayed diagnosis. Biochemical assays have been used to characterize P. insidiosum, some of which could aid in the clinical identification of this organism. Although hydrolysis of maltose and sucrose has been proposed as the key biochemical feature useful in discriminating P. insidiosum from other oomycetes and fungi, this technique requires a more rigorous evaluation involving a wider selection of P. insidiosum strains. Here, we evaluated 10 routinely available biochemical assays for characterization of 26 P. insidiosum strains, isolated from different hosts and geographic origins. Initial assessment revealed diverse biochemical characteristics across the P. insidiosum strains tested. Failure to hydrolyze sugars is observed, especially in slow-growing strains. Because hydrolysis of maltose and sucrose varied among different strains, use of the biochemical assays for identification of P. insidiosum should be cautioned. The ability of P. insidiosum to hydrolyze urea is our focus, because this metabolic process relies on the enzyme urease, an important virulence factor of other pathogens. The ability to hydrolyze urea varied among P. insidiosum strains and was not associated with growth rates. Genome analyses demonstrated that urease- and urease accessory protein-encoding genes are present in both urea-hydrolyzing and non-urea-hydrolyzing strains of P. insidiosum. Urease genes are phylogenetically conserved in P. insidiosum and related oomycetes, while the presence of urease accessory protein-encoding genes is markedly diverse in these organisms. In summary, we dissected

  11. How to kill the honey bee larva: genomic potential and virulence mechanisms of Paenibacillus larvae.

    Science.gov (United States)

    Djukic, Marvin; Brzuszkiewicz, Elzbieta; Fünfhaus, Anne; Voss, Jörn; Gollnow, Kathleen; Poppinga, Lena; Liesegang, Heiko; Garcia-Gonzalez, Eva; Genersch, Elke; Daniel, Rolf

    2014-01-01

    Paenibacillus larvae, a Gram positive bacterial pathogen, causes American Foulbrood (AFB), which is the most serious infectious disease of honey bees. In order to investigate the genomic potential of P. larvae, two strains belonging to two different genotypes were sequenced and used for comparative genome analysis. The complete genome sequence of P. larvae strain DSM 25430 (genotype ERIC II) consisted of 4,056,006 bp and harbored 3,928 predicted protein-encoding genes. The draft genome sequence of P. larvae strain DSM 25719 (genotype ERIC I) comprised 4,579,589 bp and contained 4,868 protein-encoding genes. Both strains harbored a 9.7 kb plasmid and encoded a large number of virulence-associated proteins such as toxins and collagenases. In addition, genes encoding large multimodular enzymes producing nonribosomally peptides or polyketides were identified. In the genome of strain DSM 25719 seven toxin associated loci were identified and analyzed. Five of them encoded putatively functional toxins. The genome of strain DSM 25430 harbored several toxin loci that showed similarity to corresponding loci in the genome of strain DSM 25719, but were non-functional due to point mutations or disruption by transposases. Although both strains cause AFB, significant differences between the genomes were observed including genome size, number and composition of transposases, insertion elements, predicted phage regions, and strain-specific island-like regions. Transposases, integrases and recombinases are important drivers for genome plasticity. A total of 390 and 273 mobile elements were found in strain DSM 25430 and strain DSM 25719, respectively. Comparative genomics of both strains revealed acquisition of virulence factors by horizontal gene transfer and provided insights into evolution and pathogenicity.

  12. Yeast Interspecies Comparative Proteomics Reveals Divergence in Expression Profiles and Provides Insights into Proteome Resource Allocation and Evolutionary Roles of Gene Duplication*

    Science.gov (United States)

    Kito, Keiji; Ito, Haruka; Nohara, Takehiro; Ohnishi, Mihoko; Ishibashi, Yuko; Takeda, Daisuke

    2016-01-01

    Omics analysis is a versatile approach for understanding the conservation and diversity of molecular systems across multiple taxa. In this study, we compared the proteome expression profiles of four yeast species (Saccharomyces cerevisiae, Saccharomyces mikatae, Kluyveromyces waltii, and Kluyveromyces lactis) grown on glucose- or glycerol-containing media. Conserved expression changes across all species were observed only for a small proportion of all proteins differentially expressed between the two growth conditions. Two Kluyveromyces species, both of which exhibited a high growth rate on glycerol, a nonfermentative carbon source, showed distinct species-specific expression profiles. In K. waltii grown on glycerol, proteins involved in the glyoxylate cycle and gluconeogenesis were expressed in high abundance. In K. lactis grown on glycerol, the expression of glycolytic and ethanol metabolic enzymes was unexpectedly low, whereas proteins involved in cytoplasmic translation, including ribosomal proteins and elongation factors, were highly expressed. These marked differences in the types of predominantly expressed proteins suggest that K. lactis optimizes the balance of proteome resource allocation between metabolism and protein synthesis giving priority to cellular growth. In S. cerevisiae, about 450 duplicate gene pairs were retained after whole-genome duplication. Intriguingly, we found that in the case of duplicates with conserved sequences, the total abundance of proteins encoded by a duplicate pair in S. cerevisiae was similar to that of protein encoded by nonduplicated ortholog in Kluyveromyces yeast. Given the frequency of haploinsufficiency, this observation suggests that conserved duplicate genes, even though minor cases of retained duplicates, do not exhibit a dosage effect in yeast, except for ribosomal proteins. Thus, comparative proteomic analyses across multiple species may reveal not only species-specific characteristics of metabolic processes under

  13. Direct enzyme assay evidence confirms aldehyde reductase function of Ydr541cp and Ygl039wp from Saccharomyces cerevisiae.

    Science.gov (United States)

    Moon, Jaewoong; Liu, Z Lewis

    2015-04-01

    The aldehyde reductase gene ARI1 is a recently characterized member of an intermediate subfamily within the short-chain dehydrogenase/reductase (SDR) superfamily that clarified mechanisms of in situ detoxification of 2-furaldehyde and 5-hydroxymethyl-2-furaldehyde by Saccharomyces cerevisiae. Uncharacterized open reading frames (ORFs) are common among tolerant candidate genes identified for lignocellulose-to-advanced biofuels conversion. This study presents partially purified proteins of two ORFs, YDR541C and YGL039W, and direct enzyme assay evidence against aldehyde-inhibitory compounds commonly encountered during lignocellulosic biomass fermentation processes. Each of the partially purified proteins encoded by these ORFs showed a molecular mass of approximately 38 kDa, similar to Ari1p, a protein encoded by aldehyde reductase gene. Both proteins demonstrated strong aldehyde reduction activities toward 14 aldehyde substrates, with high levels of reduction activity for Ydr541cp toward both aromatic and aliphatic aldehydes. While Ydr541cp was observed to have a significantly higher specific enzyme activity at 20 U/mg using co-factor NADPH, Ygl039wp displayed a NADH preference at 25 U/mg in reduction of butylaldehyde. Amino acid sequence analysis identified a characteristic catalytic triad, Ser, Tyr and Lys; a conserved catalytic motif of Tyr-X-X-X-Lys; and a cofactor-binding sequence motif, Gly-X-X-Gly-X-X-Ala, near the N-terminus that are shared by Ydr541cp, Ygl039wp, Yol151wp/GRE2 and Ari1p. Findings of aldehyde reductase genes contribute to the yeast gene annotation and aids development of the next-generation biocatalyst for advanced biofuels production. Copyright © 2015 John Wiley & Sons, Ltd.

  14. Diversity and evolution of ABC proteins in mycorrhiza-forming fungi.

    Science.gov (United States)

    Kovalchuk, Andriy; Kohler, Annegret; Martin, Francis; Asiegbu, Fred O

    2015-12-28

    Transporter proteins are predicted to have an important role in the mycorrhizal symbiosis, due to the fact that this type of an interaction between plants and fungi requires a continuous nutrient and signalling exchange. ABC transporters are one of the large groups of transporter proteins found both in plants and in fungi. The crucial role of plant ABC transporters in the formation of the mycorrhizal symbiosis has been demonstrated recently. Some of the fungal ABC transporter-encoding genes are also induced during the mycorrhiza formation. However, no experimental evidences of the direct involvement of fungal ABC transporters in this process are available so far. To facilitate the identification of fungal ABC proteins with a potential role in the establishment of the mycorrhizal symbiosis, we have performed an inventory of the ABC protein-encoding genes in the genomes of 25 species of mycorrhiza-forming fungi. We have identified, manually annotated and curated more than 1300 gene models of putative ABC protein-encoding genes. Out of those, more than 1000 models are predicted to encode functional proteins, whereas about 300 models represent gene fragments or putative pseudogenes. We have also performed the phylogenetic analysis of the identified sequences. The sets of ABC proteins in the mycorrhiza-forming species were compared to the related saprotrophic or plant-pathogenic fungal species. Our results demonstrate the high diversity of ABC genes in the genomes of mycorrhiza-forming fungi. Via comparison of transcriptomics data from different species, we have identified candidate groups of ABC transporters that might have a role in the process of the mycorrhiza formation. Results of our inventory will facilitate the identification of fungal transporters with a role in the mycorrhiza formation. We also provide the first data on ABC protein-coding genes for the phylum Glomeromycota and for orders Pezizales, Atheliales, Cantharellales and Sebacinales, contributing to

  15. A History of the Discovery of Random X Chromosome Inactivation in the Human Female and its Significance

    Directory of Open Access Journals (Sweden)

    Sophia Balderman

    2011-07-01

    Full Text Available Genetic determinants of sex in placental mammals developed by the evolution of primordial autosomes into the male and female sex chromosomes. The Y chromosome determines maleness by the action of the gene SRY, which encodes a protein that initiates a sequence of events prompting the embryonic gonads to develop into testes. The X chromosome in the absence of a Y chromosome results in a female by permitting the conversion of the embryonic gonads into ovaries. We trace the historical progress that resulted in the discovery that one X chromosome in the female is randomly inactivated in early embryogenesis, accomplishing approximate equivalency of X chromosome gene dosage in both sexes. This event results in half of the somatic cells in a tissue containing proteins encoded by the genes of the maternal X chromosome and half having proteins encoded by the genes of the paternal X chromosome, on average, accounting for the phenotype of a female heterozygote with an X chromosome mutation. The hypothesis of X chromosome inactivation as a random event early in embryogenesis was first described as a result of studies of variegated coat color in female mice. Similar results were found in women using the X chromosome-linked gene, glucose-6-phosphate dehydrogenase, studied in red cells. The random inactivation of the X chromosome-bearing genes for isoenzyme types A and B of glucose-6-phosphate dehydrogenase was used to establish the clonal origin of neoplasms in informative women with leiomyomas. Behind these discoveries are the stories of the men and women scientists whose research enlightened these aspects of X chromosome function and their implication for medicine.

  16. Functional characterization of Bombyx mori nucleopolyhedrovirus mutant lacking late expression factor 9.

    Science.gov (United States)

    Zhang, Y; Shi, Y; Yu, H; Li, J; Quan, Y; Shu, T; Nie, Z; Zhang, Y; Yu, W

    Baculoviridae is a family of invertebrate viruses with large double-stranded DNA genomes. Proteins encoded by some late expression factor (lef ) genes are involved in the regulation of viral gene expression. Lef-9 is one of four transcription-specific Lefs, which are components of the virus-encoded RNA polymerase, and can initiate and transcribe late and very late genes. As a multifunctional protein encoded by the Bombyx mori nucleopolyhedrovirus (BmNPV), Lef-9 may be involved in the regulation of viral propagation. However, the underlying mechanism remains unclear. To determine the role of lef-9 in baculovirus infection, lef-9-knockout virus (lef-9-KO-Bacmid virus) was constructed using the Red recombination system, and the Bac-to-Bac system was used to prepare lef-9-repaired virus (lef-9-Re-Bacmid virus). The lef-9-KO virus did not produce infectious viruses or show infection activity, while the lef-9-repaired virus recovered both. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis of the transcription levels in wild-type-Bacmid, lef-9-KO-Bacmid, and lef-9-Re-Bacmid viruses showed that the lef-9-KO bacmid had little effect on viral genome replication. However, the transcription levels of the early and late viral genes, lef-3, ie-1, vp39, and p10, were significantly lower in BmN cells transfected with lef-9-KO-Bacmids than in the controls. Electron microscopy showed no visible enveloped virions in cells transfected with lef-9-KO-Bacmids, while many mature virions in cells transfected with lef-9-Re-Bacmid and wt-Bacmid were present. Thus, lef-9 was not essential for viral genome replication, but significantly affected viral gene transcription and expression in all periods of cell life cycle.

  17. Distribution and Evolution of Yersinia Leucine-Rich Repeat Proteins

    Science.gov (United States)

    Hu, Yueming; Huang, He; Hui, Xinjie; Cheng, Xi; White, Aaron P.

    2016-01-01

    Leucine-rich repeat (LRR) proteins are widely distributed in bacteria, playing important roles in various protein-protein interaction processes. In Yersinia, the well-characterized type III secreted effector YopM also belongs to the LRR protein family and is encoded by virulence plasmids. However, little has been known about other LRR members encoded by Yersinia genomes or their evolution. In this study, the Yersinia LRR proteins were comprehensively screened, categorized, and compared. The LRR proteins encoded by chromosomes (LRR1 proteins) appeared to be more similar to each other and different from those encoded by plasmids (LRR2 proteins) with regard to repeat-unit length, amino acid composition profile, and gene expression regulation circuits. LRR1 proteins were also different from LRR2 proteins in that the LRR1 proteins contained an E3 ligase domain (NEL domain) in the C-terminal region or an NEL domain-encoding nucleotide relic in flanking genomic sequences. The LRR1 protein-encoding genes (LRR1 genes) varied dramatically and were categorized into 4 subgroups (a to d), with the LRR1a to -c genes evolving from the same ancestor and LRR1d genes evolving from another ancestor. The consensus and ancestor repeat-unit sequences were inferred for different LRR1 protein subgroups by use of a maximum parsimony modeling strategy. Structural modeling disclosed very similar repeat-unit structures between LRR1 and LRR2 proteins despite the different unit lengths and amino acid compositions. Structural constraints may serve as the driving force to explain the observed mutations in the LRR regions. This study suggests that there may be functional variation and lays the foundation for future experiments investigating the functions of the chromosomally encoded LRR proteins of Yersinia. PMID:27217422

  18. Systems-level analysis of risk genes reveals the modular nature of schizophrenia.

    Science.gov (United States)

    Liu, Jiewei; Li, Ming; Luo, Xiong-Jian; Su, Bing

    2018-05-19

    Schizophrenia (SCZ) is a complex mental disorder with high heritability. Genetic studies (especially recent genome-wide association studies) have identified many risk genes for schizophrenia. However, the physical interactions among the proteins encoded by schizophrenia risk genes remain elusive and it is not known whether the identified risk genes converge on common molecular networks or pathways. Here we systematically investigated the network characteristics of schizophrenia risk genes using the high-confidence protein-protein interactions (PPI) from the human interactome. We found that schizophrenia risk genes encode a densely interconnected PPI network (P = 4.15 × 10 -31 ). Compared with the background genes, the schizophrenia risk genes in the interactome have significantly higher degree (P = 5.39 × 10 -11 ), closeness centrality (P = 7.56 × 10 -11 ), betweeness centrality (P = 1.29 × 10 -11 ), clustering coefficient (P = 2.22 × 10 -2 ), and shorter average shortest path length (P = 7.56 × 10 -11 ). Based on the densely interconnected PPI network, we identified 48 hub genes and 4 modules formed by highly interconnected schizophrenia genes. We showed that the proteins encoded by schizophrenia hub genes have significantly more direct physical interactions. Gene ontology (GO) analysis revealed that cell adhesion, cell cycle, immune system response, and GABR-receptor complex categories were enriched in the modules formed by highly interconnected schizophrenia risk genes. Our study reveals that schizophrenia risk genes encode a densely interconnected molecular network and demonstrates the modular nature of schizophrenia. Copyright © 2018 Elsevier B.V. All rights reserved.

  19. Viroporin Activity of the Foot-and-Mouth Disease Virus Non-Structural 2B Protein.

    Directory of Open Access Journals (Sweden)

    Da Ao

    Full Text Available Viroporins are a family of low-molecular-weight hydrophobic transmembrane proteins that are encoded by various animal viruses. Viroporins form transmembrane pores in host cells via oligomerization, thereby destroying cellular homeostasis and inducing cytopathy for virus replication and virion release. Among the Picornaviridae family of viruses, the 2B protein encoded by enteroviruses is well understood, whereas the viroporin activity of the 2B protein encoded by the foot-and-mouth disease virus (FMDV has not yet been described. An analysis of the FMDV 2B protein domains by computer-aided programs conducted in this study revealed that this protein may contain two transmembrane regions. Further biochemical, biophysical and functional studies revealed that the protein possesses a number of features typical of a viroporin when it is overexpressed in bacterial and mammalian cells as well as in FMDV-infected cells. The protein was found to be mainly localized in the endoplasmic reticulum (ER, with both the N- and C-terminal domains stretched into the cytosol. It exhibited cytotoxicity in Escherichia coli, which attenuated 2B protein expression. The release of virions from cells infected with FMDV was inhibited by amantadine, a viroporin inhibitor. The 2B protein monomers interacted with each other to form both intracellular and extracellular oligomers. The Ca(2+ concentration in the cells increased, and the integrity of the cytoplasmic membrane was disrupted in cells that expressed the 2B protein. Moreover, the 2B protein induced intense autophagy in host cells. All of the results of this study demonstrate that the FMDV 2B protein has properties that are also found in other viroporins and may be involved in the infection mechanism of FMDV.

  20. The apicoplast genomes of two taxonomic units of Babesia from sheep.

    Science.gov (United States)

    Wang, Tao; Guan, Guiquan; Korhonen, Pasi K; Koehler, Anson V; Hall, Ross S; Young, Neil D; Yin, Hong; Gasser, Robin B

    2017-01-15

    The apicoplast (ap) is a unique, non-photosynthetic organelle found in most apicomplexan parasites. Due to the essential roles that this organelle has, it has been widely considered as target for drugs against diseases caused by apicomplexans. Exploring the ap genomes of such parasites would provide a better understanding of their systematics and their basic molecular biology for therapeutics. However, there is limited information available on the ap genomes of apicomplexan parasites. In the present study, the ap genomes of two operational taxonomic units of Babesia (known as Babesia sp. Lintan [Bl] and Babesia sp. Xinjiang [Bx]) from sheep were sequenced, assembled and annotated using a massive parallel sequencing-based approach. Then, the gene content and gene order in these ap genomes (∼30.7kb in size) were defined and compared, and the genetic differences were assessed. In addition, a phylogenetic analysis of ap genomic data sets was carried out to assess the relationships of these taxonomic units with other apicomplexan parasites for which complete ap genomic data sets were publicly available. The results showed that the ap genomes of Bl and Bx encode 59 and 57 genes, respectively, including 2 ribosomal RNA genes, 25 transfer RNA genes and 30-32 protein-encoding genes, being similar in content to those of Babesia bovis and B. orientalis. Ap gene regions that might serve as markers for future epidemiological and population genetic studies of Babesia species were identified. Using sequence data for a subset of six protein-encoding genes, a close relationship of Bl and Bx with Babesia bovis from cattle and B. orientalis from water buffalo was inferred. Although the focus of the present study was on Babesia, we propose that the present sequencing-bioinformatic approach should be applicable to organellar genomes of a wide range of apicomplexans of veterinary importance. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. The architecture of mammalian ribosomal protein promoters

    Directory of Open Access Journals (Sweden)

    Perry Robert P

    2005-02-01

    Full Text Available Abstract Background Mammalian ribosomes contain 79 different proteins encoded by widely scattered single copy genes. Coordinate expression of these genes at transcriptional and post-transcriptional levels is required to ensure a roughly equimolar accumulation of ribosomal proteins. To date, detailed studies of only a very few ribosomal protein (rp promoters have been made. To elucidate the general features of rp promoter architecture, I made a detailed sequence comparison of the promoter regions of the entire set of orthologous human and mouse rp genes. Results A striking evolutionarily conserved feature of most rp genes is the separation by an intron of the sequences involved in transcriptional and translational regulation from the sequences with protein encoding function. Another conserved feature is the polypyrimidine initiator, which conforms to the consensus (Y2C+1TY(T2(Y3. At least 60 % of the rp promoters contain a largely conserved TATA box or A/T-rich motif, which should theoretically have TBP-binding capability. A remarkably high proportion of the promoters contain conserved binding sites for transcription factors that were previously implicated in rp gene expression, namely upstream GABP and Sp1 sites and downstream YY1 sites. Over 80 % of human and mouse rp genes contain a transposable element residue within 900 bp of 5' flanking sequence; very little sequence identity between human and mouse orthologues was evident more than 200 bp upstream of the transcriptional start point. Conclusions This analysis has provided some valuable insights into the general architecture of mammalian rp promoters and has identified parameters that might coordinately regulate the transcriptional activity of certain subsets of rp genes.

  2. A genetically engineered prime-boost vaccination strategy for oculonasal delivery with poly(D,L-lactic-co-glycolic acid) microparticles against infection of turkeys with avian Metapneumovirus.

    Science.gov (United States)

    Liman, Martin; Peiser, Lieselotte; Zimmer, Gert; Pröpsting, Marcus; Naim, Hassan Y; Rautenschlein, Silke

    2007-11-14

    In this study we demonstrated the use of an oculonasally delivered poly(D,L-lactic-co-glycolic acid) microparticle (PLGA-MP)-based and genetically engineered vaccination strategy in the avian system. An avian Metapneumovirus (aMPV) fusion (F) protein-encoding plasmid vaccine and the corresponding recombinant protein vaccine were produced and bound to or encapsulated by PLGA-MP, respectively. The PLGA-MP as the controlled release system was shown in vitro to not induce any cytopathic effects and to efficiently deliver the F protein-based aMPV-vaccines to avian cells for further processing. Vaccination of turkeys was carried out by priming with an MP-bound F protein-encoding plasmid vaccine and a booster-vaccination with an MP-encapsulated recombinant F protein. Besides the prime-boost F-specific vaccinated birds, negative control birds inoculated with a mock-MP prime-boost regimen as well as non-vaccinated birds and live vaccinated positive control birds were included in the study. The MP-based immunization of turkeys via the oculonasal route induced systemic humoral immune reactions as well as local and systemic cellular immune reactions, and had no adverse effects on the upper respiratory tract. The F protein-specific prime-boost strategy induced partial protection. After challenge the F protein-specific MP-vaccinated birds showed less clinical signs and histopathological lesions than control birds of mock MP-vaccinated and non-vaccinated groups did. The vaccination improved viral clearance and induced accumulation of local and systemic CD4+ T cells when compared to the mock MP-vaccination. It also induced systemic aMPV-neutralizing antibodies. The comparison of mock- and F protein-specific MP-vaccinated birds to non-vaccinated control birds suggests that aMPV-specific effects as well as adjuvant effects mediated by MP may have contributed to the overall protective effect.

  3. Topological and organizational properties of the products of house-keeping and tissue-specific genes in protein-protein interaction networks.

    Science.gov (United States)

    Lin, Wen-Hsien; Liu, Wei-Chung; Hwang, Ming-Jing

    2009-03-11

    Human cells of various tissue types differ greatly in morphology despite having the same set of genetic information. Some genes are expressed in all cell types to perform house-keeping functions, while some are selectively expressed to perform tissue-specific functions. In this study, we wished to elucidate how proteins encoded by human house-keeping genes and tissue-specific genes are organized in human protein-protein interaction networks. We constructed protein-protein interaction networks for different tissue types using two gene expression datasets and one protein-protein interaction database. We then calculated three network indices of topological importance, the degree, closeness, and betweenness centralities, to measure the network position of proteins encoded by house-keeping and tissue-specific genes, and quantified their local connectivity structure. Compared to a random selection of proteins, house-keeping gene-encoded proteins tended to have a greater number of directly interacting neighbors and occupy network positions in several shortest paths of interaction between protein pairs, whereas tissue-specific gene-encoded proteins did not. In addition, house-keeping gene-encoded proteins tended to connect with other house-keeping gene-encoded proteins in all tissue types, whereas tissue-specific gene-encoded proteins also tended to connect with other tissue-specific gene-encoded proteins, but only in approximately half of the tissue types examined. Our analysis showed that house-keeping gene-encoded proteins tend to occupy important network positions, while those encoded by tissue-specific genes do not. The biological implications of our findings were discussed and we proposed a hypothesis regarding how cells organize their protein tools in protein-protein interaction networks. Our results led us to speculate that house-keeping gene-encoded proteins might form a core in human protein-protein interaction networks, while clusters of tissue-specific gene

  4. Genotypic and phenotypic characterization of multidrug resistant Salmonella Typhimurium and Salmonella Kentucky strains recovered from chicken carcasses.

    Directory of Open Access Journals (Sweden)

    Rizwana Tasmin

    Full Text Available Salmonella Typhimurium is the leading cause of human non-typhoidal gastroenteritis in the US. S. Kentucky is one the most commonly recovered serovars from commercially processed poultry carcasses. This study compared the genotypic and phenotypic properties of two Salmonella enterica strains Typhimurium (ST221_31B and Kentucky (SK222_32B recovered from commercially processed chicken carcasses using whole genome sequencing, phenotype characterizations and an intracellular killing assay. Illumina MiSeq platform was used for sequencing of two Salmonella genomes. Phylogenetic analysis employing homologous alignment of a 1,185 non-duplicated protein-coding gene in the Salmonella core genome demonstrated fully resolved bifurcating patterns with varying levels of diversity that separated ST221_31B and SK222_32B genomes into distinct monophyletic serovar clades. Single nucleotide polymorphism (SNP analysis identified 2,432 (ST19 SNPs within 13 Typhimurium genomes including ST221_31B representing Sequence Type ST19 and 650 (ST152 SNPs were detected within 13 Kentucky genomes including SK222_32B representing Sequence Type ST152. In addition to serovar-specific conserved coding sequences, the genomes of ST221_31B and SK222_32B harbor several genomic regions with significant genetic differences. These included phage and phage-like elements, carbon utilization or transport operons, fimbriae operons, putative membrane associated protein-encoding genes, antibiotic resistance genes, siderophore operons, and numerous hypothetical protein-encoding genes. Phenotype microarray results demonstrated that ST221_31B is capable of utilizing certain carbon compounds more efficiently as compared to SK222_3B; namely, 1,2-propanediol, M-inositol, L-threonine, α-D-lactose, D-tagatose, adonitol, formic acid, acetoacetic acid, and L-tartaric acid. ST221_31B survived for 48 h in macrophages, while SK222_32B was mostly eliminated. Further, a 3-fold growth of ST221_31B was

  5. Md-miR156ab and Md-miR395 Target WRKY Transcription Factors to Influence Apple Resistance to Leaf Spot Disease.

    Science.gov (United States)

    Zhang, Qiulei; Li, Yang; Zhang, Yi; Wu, Chuanbao; Wang, Shengnan; Hao, Li; Wang, Shengyuan; Li, Tianzhong

    2017-01-01

    MicroRNAs (miRNAs) are key regulators of gene expression that post-transcriptionally regulate transcription factors involved in plant physiological activities. Little is known about the effects of miRNAs in disease resistance in apple ( Malus × domestica ). We globally profiled miRNAs in the apple cultivar Golden Delicious (GD) infected or not with the apple leaf spot fungus Alternaria alternaria f. sp. mali (ALT1), and identified 58 miRNAs that exhibited more than a 2-fold upregulation upon ALT1 infection. We identified a pair of miRNAs that target protein-coding genes involved in the defense response against fungal pathogens; Md-miR156ab targets a novel WRKY transcription factor, MdWRKYN1, which harbors a TIR and a WRKY domain. Md-miR395 targets another transcription factor, MdWRKY26, which contains two WRKY domains. Real-time PCR analysis showed that Md-miR156ab and Md-miR395 levels increased, while MdWRKYN1 and MdWRKY26 expression decreased in ALT1-inoculated GD leaves; furthermore, the overexpression of Md-miR156ab and Md-miR395 resulted in a significant reduction in MdWRKYN1 and MdWRKY26 expression. To investigate whether these miRNAs and their targets play a crucial role in plant defense, we overexpressed MdWRKYN1 or knocked down Md-miR156ab activity, which in both cases enhanced the disease resistance of the plants by upregulating the expression of the WRKY-regulated pathogenesis-related (PR) protein-encoding genes MdPR3-1, MdPR3-2, MdPR4, MdPR5, MdPR10-1 , and MdPR10-2 . In a similar analysis, we overexpressed MdWRKY26 or suppressed Md-miR395 activity, and found that many PR protein-encoding genes were also regulated by MdWRKY26 . In GD, ALT-induced Md-miR156ab and Md-miR395 suppress MdWRKYN1 and MdWRKY26 expression, thereby decreasing the expression of some PR genes, and resulting in susceptibility to ALT1.

  6. Functionally relevant microsatellites in sugarcane unigenes

    Directory of Open Access Journals (Sweden)

    Singh Nagendra K

    2010-11-01

    Full Text Available Abstract Background Unigene sequences constitute a rich source of functionally relevant microsatellites. The present study was undertaken to mine the microsatellites in the available unigene sequences of sugarcane for understanding their constitution in the expressed genic component of its complex polyploid/aneuploid genome, assessing their functional significance in silico, determining the extent of allelic diversity at the microsatellite loci and for evaluating their utility in large-scale genotyping applications in sugarcane. Results The average frequency of perfect microsatellite was 1/10.9 kb, while it was 1/44.3 kb for the long and hypervariable class I repeats. GC-rich trinucleotides coding for alanine and the GA-rich dinucleotides were the most abundant microsatellite classes. Out of 15,594 unigenes mined in the study, 767 contained microsatellite repeats and for 672 of these putative functions were determined in silico. The microsatellite repeats were found in the functional domains of proteins encoded by 364 unigenes. Its significance was assessed by establishing the structure-function relationship for the beta-amylase and protein kinase encoding unigenes having repeats in the catalytic domains. A total of 726 allelic variants (7.42 alleles per locus with different repeat lengths were captured precisely for a set of 47 fluorescent dye labeled primers in 36 sugarcane genotypes and five cereal species using the automated fragment analysis system, which suggested the utility of designed primers for rapid, large-scale and high-throughput genotyping applications in sugarcane. Pair-wise similarity ranging from 0.33 to 0.84 with an average of 0.40 revealed a broad genetic base of the Indian varieties in respect of functionally relevant regions of the large and complex sugarcane genome. Conclusion Microsatellite repeats were present in 4.92% of sugarcane unigenes, for most (87.6% of which functions were determined in silico. High level of

  7. West German Study Group Phase III PlanB Trial: First Prospective Outcome Data for the 21-Gene Recurrence Score Assay and Concordance of Prognostic Markers by Central and Local Pathology Assessment.

    Science.gov (United States)

    Gluz, Oleg; Nitz, Ulrike A; Christgen, Matthias; Kates, Ronald E; Shak, Steven; Clemens, Michael; Kraemer, Stefan; Aktas, Bahriye; Kuemmel, Sherko; Reimer, Toralf; Kusche, Manfred; Heyl, Volker; Lorenz-Salehi, Fatemeh; Just, Marianne; Hofmann, Daniel; Degenhardt, Tom; Liedtke, Cornelia; Svedman, Christer; Wuerstlein, Rachel; Kreipe, Hans H; Harbeck, Nadia

    2016-07-10

    The 21-gene Recurrence Score (RS) assay is a validated prognostic/predictive tool in early hormone receptor-positive breast cancer (BC); however, only a few prospective outcome results have been available so far. In the phase III PlanB trial, RS was prospectively used to define a subset of patients who received only endocrine therapy. We present 3-year outcome data and concordance analysis (among biomarkers/RS). Central tumor bank was established prospectively from PlanB (intermediate and high-risk, locally human epidermal growth factor receptor 2-negative BC). After an early amendment, HR-positive, pN0-1 patients with RS ≤ 11 were recommended to omit chemotherapy. From 2009 to 2011, PlanB enrolled 3,198 patients with a median age of 56 years; 41.1% had node-positive and 32.5% grade 3 disease. In 348 patients (15.3%), chemotherapy was omitted based on RS ≤ 11. After 35 months median follow-up, 3-year disease-free survival in patients with RS ≤ 11 and endocrine therapy alone was 98% versus 92% and 98% in RS > 25 and RS 12 to 25 in chemotherapy-treated patients, respectively. Nodal status, central and local grade, the Ki-67 protein encoded by the MKI67 gene, estrogen receptor, progesterone receptor, tumor size, and RS were univariate prognostic factors for disease-free survival; only nodal status, both central and local grade, and RS were independent multivariate factors. Histologic grade was discordant between central and local laboratories in 44%. RS was positively but moderately correlated with the Ki-67 protein encoded by the MKI67 gene and grade and negatively correlated with progesterone receptor and estrogen receptor. In this prospective trial, patients with enhanced clinical risk and omitted chemotherapy on the basis of RS ≤ 11 had excellent 3-year survival. The substantial discordance observed between traditional prognostic markers and RS emphasizes the need for standardized assessment and supports the potential integration of standardized, well

  8. The genome of Paenibacillus sabinae T27 provides insight into evolution, organization and functional elucidation of nif and nif-like genes.

    Science.gov (United States)

    Li, Xinxin; Deng, Zhiping; Liu, Zhanzhi; Yan, Yongliang; Wang, Tianshu; Xie, Jianbo; Lin, Min; Cheng, Qi; Chen, Sanfeng

    2014-08-27

    Most biological nitrogen fixation is catalyzed by the molybdenum nitrogenase. This enzyme is a complex which contains the MoFe protein encoded by nifDK and the Fe protein encoded by nifH. In addition to nifHDK, nifHDK-like genes were found in some Archaea and Firmicutes, but their function is unclear. We sequenced the genome of Paenibacillus sabinae T27. A total of 4,793 open reading frames were predicted from its 5.27 Mb genome. The genome of P. sabinae T27 contains fifteen nitrogen fixation (nif) genes, including three nifH, one nifD, one nifK, four nifB, two nifE, two nifN, one nifX and one nifV. Of the 15 nif genes, eight nif genes (nifB, nifH, nifD, nifK, nifE, nifN, nifX and nifV) and two non-nif genes (orf1 and hesA) form a complete nif gene cluster. In addition to the nif genes, there are nitrogenase-like genes, including two nifH-like genes and five pairs of nifDK-like genes. IS elements on the flanking regions of nif and nif-like genes imply that these genes might have been obtained by horizontal gene transfer. Phylogenies of the concatenated 8 nif gene (nifB, nifH, nifD, nifK, nifE, nifN, nifX and nifV) products suggest that P. sabinae T27 is closely related to Frankia. RT-PCR analysis showed that the complete nif gene cluster is organized as an operon. We demonstrated that the complete nif gene cluster under the control of σ70-dependent promoter enabled Escherichia coli JM109 to fix nitrogen. Also, here for the first time we demonstrated that unlike nif genes, the transcriptions of nifHDK-like genes were not regulated by ammonium and oxygen, and nifH-like or nifD-like gene could not restore the nitrogenase activity of Klebsiella pneumonia nifH- and nifD- mutant strains, respectively, suggesting that nifHDK-like genes were not involved in nitrogen fixation. Our data and analysis reveal the contents and distribution of nif and nif-like genes and contribute to the study of evolutionary history of nitrogen fixation in Paenibacillus. For the first time we

  9. A comparative gene analysis with rice identified orthologous group II HKT genes and their association with Na(+) concentration in bread wheat.

    Science.gov (United States)

    Ariyarathna, H A Chandima K; Oldach, Klaus H; Francki, Michael G

    2016-01-19

    Although the HKT transporter genes ascertain some of the key determinants of crop salt tolerance mechanisms, the diversity and functional role of group II HKT genes are not clearly understood in bread wheat. The advanced knowledge on rice HKT and whole genome sequence was, therefore, used in comparative gene analysis to identify orthologous wheat group II HKT genes and their role in trait variation under different saline environments. The four group II HKTs in rice identified two orthologous gene families from bread wheat, including the known TaHKT2;1 gene family and a new distinctly different gene family designated as TaHKT2;2. A single copy of TaHKT2;2 was found on each homeologous chromosome arm 7AL, 7BL and 7DL and each gene was expressed in leaf blade, sheath and root tissues under non-stressed and at 200 mM salt stressed conditions. The proteins encoded by genes of the TaHKT2;2 family revealed more than 93% amino acid sequence identity but ≤52% amino acid identity compared to the proteins encoded by TaHKT2;1 family. Specifically, variations in known critical domains predicted functional differences between the two protein families. Similar to orthologous rice genes on chromosome 6L, TaHKT2;1 and TaHKT2;2 genes were located approximately 3 kb apart on wheat chromosomes 7AL, 7BL and 7DL, forming a static syntenic block in the two species. The chromosomal region on 7AL containing TaHKT2;1 7AL-1 co-located with QTL for shoot Na(+) concentration and yield in some saline environments. The differences in copy number, genes sequences and encoded proteins between TaHKT2;2 homeologous genes and other group II HKT gene families within and across species likely reflect functional diversity for ion selectivity and transport in plants. Evidence indicated that neither TaHKT2;2 nor TaHKT2;1 were associated with primary root Na(+) uptake but TaHKT2;1 may be associated with trait variation for Na(+) exclusion and yield in some but not all saline environments.

  10. Properties and genomic analysis of Lactococcus garvieae lysogenic bacteriophage PLgT-1, a new member of Siphoviridae, with homology to Lactococcus lactis phages.

    Science.gov (United States)

    Hoai, Truong Dinh; Nishiki, Issei; Yoshida, Terutoyo

    2016-08-15

    The lysogenic phage PLgT-1 is highly prevalent in Lactococcus garvieae, which is a serious bacterial pathogen in marine fish. Therefore, information regarding this phage is one of the key factors to predict the evolution of this bacterium. However, many properties of this phage, its complete genome sequence, and its relationship with other viral communities has not been investigated to date. Here, we demonstrated that the phage PLgT-1 was not only induced by an induction agent (Mitomycin C), but could be released frequently during cell division in a nutrient-rich environment or in natural seawater. Integration of PLgT-1 into non-lysogenic bacteria via transduction changed the genotype, resulting in the diversification of L. garvieae. The complete DNA sequence of PLgT-1 was also determined. This phage has a dsDNA genome of 40,273bp with 66 open reading frames (ORFs). Of these, the biological functions of 24 ORFs could be predicted but those of 42 ORFs are unknown. Thus, PLgT-1 is a novel phage with several novel proteins encoded in its genome. The strict MegaBLAST search program for the PLgT-1 genome revealed that this phage had no similarities with other previously investigated phages specific to L. garvieae (WP-2 and GE1). Notably, PLgT-1 was relatively homologous with several phages of Lactococcus lactis and 17 of the 24 predicted proteins encoded in PLgT-1 were homologous with the deduced proteins of various phages from these dairy bacteria. Comparative genome analysis revealed that the L. garvieae phage PLgT-1 was most closely related to the L. lactis phage TP712. However, they differed from each other in genome size and gene arrangement. The results obtained in this study suggest that the lysogenic phage PLgT-1 is a new member of the family Siphoviridae and has been involved in horizontal gene exchange with microbial communities, especially with L. lactis and its phages. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

    Science.gov (United States)

    In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A; Rubin, P; Kemp, J; Israel, E; Busse, W; Ledford, D; Murray, J J; Segal, A; Tinkleman, D; Drazen, J M

    1997-03-01

    Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, deletion of two, or addition of one zinc finger (Sp1/Egr-1) binding sites in the region 176 to 147 bp upstream from the ATG translation start site where there are normally 5 Sp1 binding motifs in tandem. Reporter gene activity directed by any of the mutant forms of the transcription factor binding region was significantly (P < 0.05) less effective than the activity driven by the wild type transcription factor binding region. Electrophoretic mobility shift assays (EMSAs) demonstrated the capacity of wild type and mutant transcription factor binding regions to bind nuclear extracts from human umbilical vein endothelial cells (HUVECs). These data are consistent with a family of mutations in the 5-LO gene that can modify reporter gene transcription possibly through differences in Sp1 and Egr-1 transactivation.

  12. Molecular cloning, expression and characterization of a serine proteinase inhibitor gene from Entamoeba histolytica.

    Science.gov (United States)

    Riahi, Yael; Siman-Tov, Rama; Ankri, Serge

    2004-02-01

    Serine proteinase inhibitors (serpins) are irreversible suicide inhibitors of proteinases that regulate a wide range of biological processes, including pathogen evasion of the host defence system. We report the cloning and characterization of a gene encoding a serpin from the protozoan parasite Entamoeba histolytica (Ehserp) that may function in this manner. The protein encoded by Ehserp contains 371 amino acids with a predicted mass of 42.6 kDa. Antibodies to a 42 kDa recombinant Ehserp react specifically with two bands of 42 and 49 kDa in trophozoite extracts. Ehserp has a cytoplasmic localization and is secreted by trophozoites incubated in the presence of mammalian cells, but not by resting trophozoites. A panel of mammalian serine proteinases was screened, but none of them was inhibited by the recombinant Ehserp. In contrast, the 49 kDa Ehserp present in the secretion product (SP) of activated macrophages interacted with human neutrophil cathepsin G to form a complex resistant to sodium dodecyl sulphate. We discuss the nature of the 42 and 49 kDa Ehserp and the possible roles that Ehserp may play in the survival of the parasite inside the host.

  13. A chemokine-binding domain in the tumor necrosis factor receptor from variola (smallpox) virus.

    Science.gov (United States)

    Alejo, Alí; Ruiz-Argüello, M Begoña; Ho, Yin; Smith, Vincent P; Saraiva, Margarida; Alcami, Antonio

    2006-04-11

    Variola virus (VaV) is the causative agent of smallpox, one of the most devastating diseases encountered by man, that was eradicated in 1980. The deliberate release of VaV would have catastrophic consequences on global public health. However, the mechanisms that contribute to smallpox pathogenesis are poorly understood at the molecular level. The ability of viruses to evade the host defense mechanisms is an important determinant of viral pathogenesis. Here we show that the tumor necrosis factor receptor (TNFR) homologue CrmB encoded by VaV functions not only as a soluble decoy TNFR but also as a highly specific binding protein for several chemokines that mediate recruitment of immune cells to mucosal surfaces and the skin, sites of virus entry and viral replication at late stages of smallpox. CrmB binds chemokines through its C-terminal domain, which is unrelated to TNFRs, was named smallpox virus-encoded chemokine receptor (SECRET) domain and uncovers a family of poxvirus chemokine inhibitors. An active SECRET domain was found in another viral TNFR (CrmD) and three secreted proteins encoded by orthopoxviruses. These findings identify a previously undescribed chemokine-binding and inhibitory domain unrelated to host chemokine receptors and a mechanism of immune modulation in VaV that may influence smallpox pathogenesis.

  14. Induction of senescence and identification of differentially expressed genes in tomato in response to monoterpene.

    Directory of Open Access Journals (Sweden)

    Sumit Ghosh

    Full Text Available Monoterpenes, which are among the major components of plant essential oils, are known for their ecological roles as well for pharmaceutical properties. Geraniol, an acyclic monoterpene induces cell cycle arrest and apoptosis/senescence in various cancer cells and plants; however, the genes involved in the process and the underlying molecular mechanisms are not well understood. In this study, we demonstrate that treatment of tomato plants with geraniol results in induction of senescence due to a substantial alteration in transcriptome. We have identified several geraniol-responsive protein encoding genes in tomato using suppression subtractive hybridization (SSH approach. These genes comprise of various components of signal transduction, cellular metabolism, reactive oxygen species (ROS, ethylene signalling, apoptosis and DNA damage response. Upregulation of NADPH oxidase and antioxidant genes, and increase in ROS level after geraniol treatment point towards the involvement of ROS in geraniol-mediated senescence. The delayed onset of seedling death and induced expression of geraniol-responsive genes in geraniol-treated ethylene receptor mutant (Nr suggest that geraniol-mediated senescence involves both ethylene dependent and independent pathways. Moreover, expression analysis during tomato ripening revealed that geraniol-responsive genes are also associated with the natural organ senescence process.

  15. Induction of Senescence and Identification of Differentially Expressed Genes in Tomato in Response to Monoterpene

    Science.gov (United States)

    Kumar, Vinay; Kumar, Anil; Irfan, Mohammad; Chakraborty, Niranjan; Chakraborty, Subhra; Datta, Asis

    2013-01-01

    Monoterpenes, which are among the major components of plant essential oils, are known for their ecological roles as well for pharmaceutical properties. Geraniol, an acyclic monoterpene induces cell cycle arrest and apoptosis/senescence in various cancer cells and plants; however, the genes involved in the process and the underlying molecular mechanisms are not well understood. In this study, we demonstrate that treatment of tomato plants with geraniol results in induction of senescence due to a substantial alteration in transcriptome. We have identified several geraniol-responsive protein encoding genes in tomato using suppression subtractive hybridization (SSH) approach. These genes comprise of various components of signal transduction, cellular metabolism, reactive oxygen species (ROS), ethylene signalling, apoptosis and DNA damage response. Upregulation of NADPH oxidase and antioxidant genes, and increase in ROS level after geraniol treatment point towards the involvement of ROS in geraniol-mediated senescence. The delayed onset of seedling death and induced expression of geraniol-responsive genes in geraniol-treated ethylene receptor mutant (Nr) suggest that geraniol-mediated senescence involves both ethylene dependent and independent pathways. Moreover, expression analysis during tomato ripening revealed that geraniol-responsive genes are also associated with the natural organ senescence process. PMID:24098759

  16. Characterization and functional analysis of four HYH splicing variants in Arabidopsis hypocotyl elongation.

    Science.gov (United States)

    Li, Chen; Zheng, Lanlan; Zhang, Jingxuan; Lv, Yanxia; Liu, Jianping; Wang, Xuanbin; Palfalvi, Gergo; Wang, Guodong; Zhang, Yonghong

    2017-07-01

    Arabidopsis thaliana LONG HYPOCOTYL5 (HY5) is a positive regulator of the light signaling pathway. The hy5 mutant has an elongated hypocotyl in all light conditions, whereas the hy5 homolog (hyh) mutant has a very weak phenotype, but only in blue light. However, overexpression of HYH rescues the elongated hypocotyl phenotype in the hy5 null mutant. Here, we report the identification of four HYH splicing variants in Arabidopsis. Alternative splicing in the 5' region of the HYH gene occurred such that the proteins encoded by all four HYH variants retained their bZIP domain. In hypocotyl tissue, transcript levels of HYH.2, HYH.3, and HYH.4 were higher than those of HYH.1. Like HY5, all HYH variants were induced by light. Functional analysis of the four HYH variants, based on their abilities to complement the hy5 mutant, indicated that they have similar roles in hypocotyl development, and may function redundantly with HY5. Our results indicate that the bZIP domain in HYH is critical for the function of four variants in the compensation of hy5 mutant in hypocotyl development. Additionally, while HY5/HYH is found in plant species ranging from green algae to flowering plants, the potential alternative splicing events are distinct in different species, with certain HYH variants found with greater frequency in some species than others. Copyright © 2017. Published by Elsevier B.V.

  17. Characterization of the major formamidopyrimidine-DNA glycosylase homolog in Mycobacterium tuberculosis and its linkage to variable tandem repeats.

    Science.gov (United States)

    Olsen, Ingrid; Balasingham, Seetha V; Davidsen, Tonje; Debebe, Ephrem; Rødland, Einar A; van Soolingen, Dick; Kremer, Kristin; Alseth, Ingrun; Tønjum, Tone

    2009-07-01

    The ability to repair DNA damage is likely to play an important role in the survival of facultative intracellular parasites because they are exposed to high levels of reactive oxygen species and nitrogen intermediates inside phagocytes. Correcting oxidative damage in purines and pyrimidines is the primary function of the enzymes formamidopyrimidine (faPy)-DNA glycosylase (Fpg) and endonuclease VIII (Nei) of the base excision repair pathway, respectively. Four gene homologs, belonging to the fpg/nei family, have been identified in Mycobacterium tuberculosis H37Rv. The recombinant protein encoded by M. tuberculosis Rv2924c, termed Mtb-Fpg1, was overexpressed, purified and biochemically characterized. The enzyme removed faPy and 5-hydroxycytosine lesions, as well as 8-oxo-7,8-dihydroguanine (8oxoG) opposite to C, T and G. Mtb-Fpg1 thus exhibited substrate specificities typical for Fpg enzymes. Although Mtb-fpg1 showed nearly complete nucleotide sequence conservation in 32 M. tuberculosis isolates, the region upstream of Mtb-fpg1 in these strains contained tandem repeat motifs of variable length. A relationship between repeat length and Mtb-fpg1 expression level was demonstrated in M. tuberculosis strains, indicating that an increased length of the tandem repeats positively influenced the expression levels of Mtb-fpg1. This is the first example of such a tandem repeat region of variable length being linked to the expression level of a bacterial gene.

  18. MadR1, a Mycobacterium tuberculosis cell cycle stress response protein that is a member of a widely conserved protein class of prokaryotic, eukaryotic and archeal origin.

    Science.gov (United States)

    Crew, Rebecca; Ramirez, Melissa V; England, Kathleen; Slayden, Richard A

    2015-05-01

    Stress-induced molecular programs designed to stall division progression are nearly ubiquitous in bacteria, with one well-known example being the participation of the SulA septum inhibiting protein in the SOS DNA damage repair response. Mycobacteria similarly demonstrate stress-altered growth kinetics, however no such regulators have been found in these organisms. We therefore set out to identify SulA-like regulatory proteins in Mycobacterium tuberculosis. A bioinformatics modeling-based approach led to the identification of rv2216 as encoding for a protein with weak similarity to SulA, further analysis distinguished this protein as belonging to a group of uncharacterized growth promoting proteins. We have named the mycobacterial protein encoded by rv2216 morphology altering division regulator protein 1, MadR1. Overexpression of madR1 modulated cell length while maintaining growth kinetics similar to wild-type, and increased the proportion of bent or V-form cells in the population. The presence of MadR1-GFP at regions of cellular elongation (poles) and morphological differentiation (V-form) suggests MadR1 involvement in phenotypic heterogeneity and longitudinal cellular growth. Global transcriptional analysis indicated that MadR1 functionality is linked to lipid editing programs required for growth and persistence. This is the first report to differentiate the larger class of these conserved proteins from SulA proteins and characterizes MadR1 effects on the mycobacterial cell. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Mining secreted proteins that function in pepper fruit development and ripening using a yeast secretion trap (YST)

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Je Min, E-mail: jemin@knu.ac.kr [Department of Plant Science, College of Agriculture and Life Sciences, Seoul National University, Seoul (Korea, Republic of); Department of Horticultural Science, Kyungpook National University, Daegu (Korea, Republic of); Lee, Sang-Jik [Biotechnology Institute, Nongwoo Bio Co, Ltd, Yeoju (Korea, Republic of); Department of Plant Biology, Cornell University, Ithaca, NY (United States); Rose, Jocelyn K.C. [Department of Plant Biology, Cornell University, Ithaca, NY (United States); Yeam, Inhwa [Department of Horticulture and Breeding, Andong National University, Andong (Korea, Republic of); Kim, Byung-Dong [Department of Plant Science, College of Agriculture and Life Sciences, Seoul National University, Seoul (Korea, Republic of)

    2014-04-18

    Highlights: • Yeast secretion trap (YST) is a valuable tool for mining secretome. • A total of 80 secreted proteins are newly identified via YST in pepper fruits. • The secreted proteins are differentially regulated during pepper development and ripening. • Transient GFP-fusion assay and in planta secretion trap can effectively validate the secretion of proteins. - Abstract: Plant cells secrete diverse sets of constitutively- and conditionally-expressed proteins under various environmental and developmental states. Secreted protein populations, or secretomes have multiple functions, including defense responses, signaling, metabolic processes, and developmental regulation. To identify genes encoding secreted proteins that function in fruit development and ripening, a yeast secretion trap (YST) screen was employed using pepper (Capsicum annuum) fruit cDNAs. The YST screen revealed 80 pepper fruit-related genes (CaPFRs) encoding secreted proteins including cell wall proteins, several of which have not been previously described. Transient GFP-fusion assay and an in planta secretion trap were used to validate the secretion of proteins encoded by selected YST clones. In addition, RNA gel blot analyses provided further insights into their expression and regulation during fruit development and ripening. Integrating our data, we conclude that the YST provides a valuable functional genomics tool for the identification of substantial numbers of novel secreted plant proteins that are associated with biological processes, including fruit development and ripening.

  20. First description of a novel mitochondrial mutation in the MT-TI gene associated with multiple mitochondrial DNA deletion and depletion in family with severe dilated mitochondrial cardiomyopathy.

    Science.gov (United States)

    Alila-Fersi, Olfa; Tabebi, Mouna; Maalej, Marwa; Belguith, Neila; Keskes, Leila; Mkaouar-Rebai, Emna; Fakhfakh, Faiza

    2018-03-18

    Mitochondria are essential for early cardiac development and impaired mitochondrial function was described associated with heart diseases such as hypertrophic or dilated mitochondrial cardiomyopathy. In this study, we report a family including two individuals with severe dilated mitochondrial cardiomyopathy. The whole mitochondrial genome screening showed the presence of several variations and a novel homoplasmic mutation m.4318-4322delC in the MT-TI gene shared by the two patients and their mother and leading to a disruption of the tRNA Ile secondary structure. In addition, a mitochondrial depletion was present in blood leucocyte of the two affected brother whereas a de novo heteroplasmic multiple deletion in the major arc of mtDNA was present in blood leucocyte and mucosa of only one of them. These deletions in the major arc of the mtDNA resulted to the loss of several protein-encoding genes and also some tRNA genes. The mtDNA deletion and depletion could result to an impairment of the oxidative phosphorylation and energy metabolism in the respiratory chain in the studied patients. Our report is the first description of a family with severe lethal dilated mitochondrial cardiomyopathy and presenting several mtDNA abnormalities including punctual mutation, deletion and depletion. Copyright © 2018 Elsevier Inc. All rights reserved.

  1. Comparative genomics of Lactobacillus salivarius strains focusing on their host adaptation.

    Science.gov (United States)

    Lee, Jun-Yeong; Han, Geon Goo; Kim, Eun Bae; Choi, Yun-Jaie

    2017-12-01

    Lactobacillus salivarius is an important member of the animal gut microflora and is a promising probiotic bacterium. However, there is a lack of research on the genomic diversity of L. salivarius species. In this study, we generated 21 L. salivarius draft genomes, and investigated the pan-genome of L. salivarius strains isolated from humans, pigs and chickens using all available genomes, focusing on host adaptation. Phylogenetic clustering showed a distinct categorization of L. salivarius strains depending on their hosts. In the pan-genome, 15 host-specific genes and 16 dual-host-shared genes that only one host isolate did not possess were identified. Comparison of 56 extracellular protein encoding genes and 124 orthologs related to exopolysaccharide production in the pan-genome revealed that extracellular components of the assayed bacteria have been globally acquired and mutated under the selection pressure for host adaptation. We also found the three host-specific genes that are responsible for energy production in L. salivarius. These results showed that L. salivarius has evolved to adapt to host habitats in two ways, by gaining the abilities for niche adhesion and efficient utilization of nutrients. Our study offers a deeper understanding of the probiotic species L. salivarius, and provides a basis for future studies on L. salivarius and other mutualistic bacteria. Copyright © 2017 Elsevier GmbH. All rights reserved.

  2. Differential expression of the lethal gene Luteus-Pa in cacao of the Parinari series.

    Science.gov (United States)

    Rehem, B C; Almeida, A-A F; Figueiredo, G S F; Gesteira, A S; Santos, S C; Corrêa, R X; Yamada, M M; Valle, R R

    2016-02-22

    The recessive lethal character Luteus-Pa is found in cacao (Theobroma cacao) genotypes of the Parinari series (Pa) and is characterized by expression of leaf chlorosis and seedling death. Several genotypes of the Pa series are bearers of the gene responsible for the expression of the Luteus-Pa character, which can be used as a tool for determining relationships between genotypes of this group. To evaluate this phenomenon, we analyzed the differential expression of genes between mutant seedlings and wild-type hybrid Pa 30 x 169 seedlings, with the aim of elucidating the possible lethal mechanisms of the homozygous recessive character Luteus-Pa. Plant material was harvested from leaves of wild and mutant seedlings at different periods to construct a subtractive library and perform quantitative analysis using real-time PCR. The 649 sequences obtained from the subtractive library had an average length of 500 bp, forming 409 contigs. The probable proteins encoded were grouped into 10 functional categories. Data from ESTs identified genes associated with Rubisco, peroxidases, and other proteins and enzymes related to carbon assimilation, respiration, and photosystem 2. Mutant seedlings were characterized by synthesizing defective PsbO and PsbA proteins, which were overexpressed from 15 to 20 days after seedling emergence.

  3. Candidate Essential Genes in Burkholderia cenocepacia J2315 Identified by Genome-Wide TraDIS

    KAUST Repository

    Wong, Yee-Chin

    2016-08-22

    Burkholderia cenocepacia infection often leads to fatal cepacia syndrome in cystic fibrosis patients. However, antibiotic therapy rarely results in complete eradication of the pathogen due to its intrinsic resistance to many clinically available antibiotics. Recent attention has turned to the identification of essential genes as the proteins encoded by these genes may serve as potential targets for development of novel antimicrobials. In this study, we utilized TraDIS (Transposon Directed Insertion-site Sequencing) as a genome-wide screening tool to facilitate the identification of B. cenocepacia genes essential for its growth and viability. A transposon mutant pool consisting of approximately 500,000 mutants was successfully constructed, with more than 400,000 unique transposon insertion sites identified by computational analysis of TraDIS datasets. The saturated library allowed for the identification of 383 genes that were predicted to be essential in B. cenocepacia. We extended the application of TraDIS to identify conditionally essential genes required for in vitro growth and revealed an additional repertoire of 439 genes to be crucial for B. cenocepacia growth under nutrient-depleted conditions. The library of B. cenocepacia mutants can subsequently be subjected to various biologically related conditions to facilitate the discovery of genes involved in niche adaptation as well as pathogenicity and virulence.

  4. Suppression of cell division by pKi-67 antisense-RNA and recombinant protein.

    Science.gov (United States)

    Duchrow, M; Schmidt, M H; Zingler, M; Anemüller, S; Bruch, H P; Broll, R

    2001-01-01

    The human antigen defined by the monoclonal antibody Ki-67 (pKi-67) is a human nuclear protein strongly associated with cell proliferation and found in all tissues studied. It is widely used as a marker of proliferating cells, yet its function is unknown. To investigate its function we suppressed pKi-67 expression by antisense RNA and overexpressed a partial structure of pKi-67 in HeLa cells. A BrdU-incorporation assay showed a significant decrease in DNA synthesis after antisense inhibition. Cell cycle analysis indicated a higher proportion of cells in G1 phase and a lower proportion of cells in S phase while the number of G(2)/M phase cells remained constant. Overexpression of a recombinant protein encoding three of the repetitive elements from exon 13 of pKi-67 had a similar effect to that obtained by antisense inhibition. The similarity of the effect of expressing 'Ki-67 repeats' and pKi-67 antisense RNA could be explained by a negative effect on the folding of the endogenous protein in the endoplasmatic reticulum. Furthermore excessive self-association of pKi-67 via the repeat structure could inhibit its nuclear transport, preventing it from getting to its presumptive site of action. We conclude that the Ki-67 protein has an important role in the regulation of the cell cycle, which is mediated in part by its repetitive elements. Copyright 2001 S. Karger AG, Basel

  5. Structural biology of the sequestration and transport of heavy metal toxins: NMR structure determination of proteins containing the -Cys-X-Y-Cys-metal binding motifs. 1998 annual progress report

    International Nuclear Information System (INIS)

    Opella, S.J.

    1998-01-01

    'The overall goal of the research is to apply the methods of structural biology, which have been previously used primarily in biomedical applications, to bioremediation. The authors are doing this by using NMR spectroscopy to determine the structures of proteins involved in the bacterial mercury detoxification system. The research is based on the premise that the proteins encoded in the genes of the bacterial detoxification system are an untapped source of reagents and, more fundamentally, chemical strategies that can be used to remove heavy metal toxins from the environment. The initial goals are to determine the structures of the proteins of the bacterial mercury detoxification systems responsible for the sequestration and transport of the Hg(II) ions in to the cell where reduction to Hg(O) occurs. These proteins are meP, which is water soluble and can be investigated with multidimensional solution NMR methods, and merT, the transport protein in the membrane that requires solid-state NMR methods. As of June 1998, this report summarizes work after about one and half years of the three-year award. The authors have made significant accomplishments in three aspects of the NMR studies of the proteins of the bacterial mercury detoxification system.'

  6. Transcriptional and Epigenetic Regulatory Mechanisms Affecting HTLV-1 Provirus

    Directory of Open Access Journals (Sweden)

    Paola Miyazato

    2016-06-01

    Full Text Available Human T-cell leukemia virus type 1 (HTLV-1 is a retrovirus associated with human diseases, such as adult T-cell leukemia (ATL and HTLV-1-associated myelopathy/Tropic spastic paraparesis (HAM/TSP. As a retrovirus, its life cycle includes a step where HTLV-1 is integrated into the host genomic DNA and forms proviral DNA. In the chronic phase of the infection, HTLV‑1 is known to proliferate as a provirus via the mitotic division of the infected host cells. There are generally tens of thousands of infected clones within an infected individual. They exist not only in peripheral blood, but also in various lymphoid organs. Viral proteins encoded in HTLV-1 genome play a role in the proliferation and survival of the infected cells. As is the case with other chronic viral infections, HTLV-1 gene expression induces the activation of the host immunity against the virus. Thus, the transcription from HTLV-1 provirus needs to be controlled in order to evade the host immune surveillance. There should be a dynamic and complex regulation in vivo, where an equilibrium between viral antigen expression and host immune surveillance is achieved. The mechanisms regulating viral gene expression from the provirus are a key to understanding the persistent/latent infection with HTLV-1 and its pathogenesis. In this article, we would like to review our current understanding on this topic.

  7. Molecular Determinants of Human T-lymphotropic Virus Type 1 Transmission and Spread

    Directory of Open Access Journals (Sweden)

    Patrick L. Green

    2011-07-01

    Full Text Available Human T-lymphotrophic virus type-1 (HTLV-1 infects approximately 15 to 20 million people worldwide, with endemic areas in Japan, the Caribbean, and Africa. The virus is spread through contact with bodily fluids containing infected cells, most often from mother to child through breast milk or via blood transfusion. After prolonged latency periods, approximately 3 to 5% of HTLV-1 infected individuals will develop either adult T-cell leukemia/lymphoma (ATL, or other lymphocyte-mediated disorders such as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP. The genome of this complex retrovirus contains typical gag, pol, and env genes, but also unique nonstructural proteins encoded from the pX region. These nonstructural genes encode the Tax and Rex regulatory proteins, as well as novel proteins essential for viral spread in vivo such as, p30, p12, p13 and the antisense encoded HBZ. While progress has been made in the understanding of viral determinants of cell transformation and host immune responses, host and viral determinants of HTLV-1 transmission and spread during the early phases of infection are unclear. Improvements in the molecular tools to test these viral determinants in cellular and animal models have provided new insights into the early events of HTLV-1 infection. This review will focus on studies that test HTLV-1 determinants in context to full length infectious clones of the virus providing insights into the mechanisms of transmission and spread of HTLV-1.

  8. Diversity of extracellular proteins during the transition from the ‘proto-apicomplexan’ alveolates to the apicomplexan obligate parasites

    KAUST Repository

    Templeton, Thomas J.

    2015-11-20

    The recent completion of high-coverage draft genome sequences for several alveolate protozoans – namely, the chromerids, Chromera velia and Vitrella brassicaformis ; the perkinsid Perkinsus marinus ; the apicomplexan, Gregarina niphandrodes , as well as high coverage transcriptome sequence information for several colpodellids, allows for new genome-scale comparisons across a rich landscape of apicomplexans and other alveolates. Genome annotations can now be used to help interpret fine ultrastructure and cell biology, and guide new studies to describe a variety of alveolate life strategies, such as symbiosis or free living, predation, and obligate intracellular parasitism, as well to provide foundations to dissect the evolutionary transitions between these niches. This review focuses on the attempt to identify extracellular proteins which might mediate the physical interface of cell–cell interactions within the above life strategies, aided by annotation of the repertoires of predicted surface and secreted proteins encoded within alveolate genomes. In particular, we discuss what descriptions of the predicted extracellular proteomes reveal regarding a hypothetical last common ancestor of a pre-apicomplexan alveolate – guided by ultrastructure, life strategies and phylogenetic relationships – in an attempt to understand the evolution of obligate parasitism in apicomplexans.

  9. Lethal distemper in badgers (Meles meles) following epidemic in dogs and wolves.

    Science.gov (United States)

    Di Sabatino, Daria; Di Francesco, Gabriella; Zaccaria, Guendalina; Malatesta, Daniela; Brugnola, Luca; Marcacci, Maurilia; Portanti, Ottavio; De Massis, Fabrizio; Savini, Giovanni; Teodori, Liana; Ruggieri, Enzo; Mangone, Iolanda; Badagliacca, Pietro; Lorusso, Alessio

    2016-12-01

    Canine distemper virus (CDV) represents an important conservation threat to many wild carnivores. A large distemper epidemic sustained by an Arctic-lineage strain occurred in Italy in 2013, mainly in the Abruzzi region, causing overt disease in domestic and shepherd dogs, Apennine wolves (Canis lupus) and other wild carnivores. Two badgers were collected by the end of September 2015 in a rural area of the Abruzzi region and were demonstrated to be CDV-positive by real time RT-PCR and IHC in several tissues. The genome of CDV isolates from badgers showed Y549H substitution in the mature H protein. By employing all publicly available Arctic-lineage H protein encoding gene sequences, six amino acid changes in recent Italian strains with respect to Italian strains of dogs from 2000 to 2008, were observed. A CDV strain belonging to the European-wildlife lineage was also identified in a fox found dead in the same region in 2016, proving co-circulation of an additional CDV lineage. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. cDNA cloning of human DNA topoisomerase I. Catalytic activity of a 67.7-kDa carboxyl-terminal fragment

    International Nuclear Information System (INIS)

    D'Arpa, P.; Machlin, P.S.; Ratrie, H. III; Rothfield, N.F.; Cleveland, D.W.; Earnshaw, W.C.

    1988-01-01

    cDNA clones encoding human topoisomerase I were isolated from an expression vector library (λgt11) screened with autoimmune anti-topoisomerase I serum. One of these clones has been expressed as a fusion protein comprised of a 32-kDa fragment of the bacterial TrpE protein linked to 67.7 kDa of protein encoded by the cDNA. Three lines of evidence indicate that the cloned cDNA encodes topoisomerase I. (i) Proteolysis maps of the fusion protein and human nuclear topoisomerase I are essentially identical. (ii) The fusion protein relaxes supercoiled DNA, an activity that can be immunoprecipitated by anti-topoisomerase I serum. (iii) Sequence analysis has revealed that the longest cDNA clone (3645 base pairs) encodes a protein of 765 amino acids that shares 42% identity with Saccharomyces cerevisiae topoisomerase I. The sequence data also show that the catalytically active 67.7-kDa fragment is comprised of the carboxyl terminus

  11. A calcineurin inhibitory protein overexpressed in Down's syndrome interacts with the product of a ubiquitously expressed transcript

    Directory of Open Access Journals (Sweden)

    H.C.S. Silveira

    2004-06-01

    Full Text Available The Down's syndrome candidate region 1 (DSCR1 protein, encoded by a gene located in the human chromosome 21, interacts with calcineurin and is overexpressed in Down's syndrome patients. As an approach to clarifying a putative function for this protein, in the present study we used the yeast two-hybrid system to identify DSCR1 partners. The two-hybrid system is a method that allows the identification of protein-protein interactions through reconstitution of the activity of the yeast GAL 4 transcriptional activator. The gene DSCR1 fused to the GAL 4 binding domain (BD was used to screen a human fetal brain cDNA library cloned in fusion with the GAL 4 activation domain (AD. Three positive clones were found and sequence analysis revealed that all the plasmids coded for the ubiquitously expressed transcript (UXT. UXT, which is encoded in human Xp11, is a 157-amino acid protein present in both cytosol and nucleus of the cells. This positive interaction of DSCR1 and UXT was confirmed in vivo by mating the yeast strain AH109 (MATaexpressing AD-UXT with the strain Y187 (MATalpha expressing BD-DSCR1, and in vitro by co-immunoprecipitation experiments. These results may help elucidate a new function for DSCR1 and its participation in Down's syndrome pathogenesis.

  12. Immunogenicity of 60 novel latency-related antigens of Mycobacterium tuberculosis.

    Science.gov (United States)

    Serra-Vidal, Mᵃdel Mar; Latorre, Irene; Franken, Kees L C M; Díaz, Jéssica; de Souza-Galvão, Maria Luiza; Casas, Irma; Maldonado, José; Milà, Cèlia; Solsona, Jordi; Jimenez-Fuentes, M Ángeles; Altet, Neus; Lacoma, Alícia; Ruiz-Manzano, Juan; Ausina, Vicente; Prat, Cristina; Ottenhoff, Tom H M; Domínguez, José

    2014-01-01

    The aim of our work here was to evaluate the immunogenicity of 60 mycobacterial antigens, some of which have not been previously assessed, notably a novel series of in vivo-expressed Mycobacterium tuberculosis (IVE-TB) antigens. We enrolled 505 subjects and separated them in individuals with and without latent tuberculosis infection (LTBI) vs. patients with active tuberculosis (TB). Following an overnight and 7 days stimulation of whole blood with purified recombinant M. tuberculosis antigens, interferon-γ (IFN-γ) levels were determined by ELISA. Several antigens could statistically significantly differentiate the groups of individuals. We obtained promising antigens from all studied antigen groups [dormancy survival regulon (DosR regulon) encoded antigens; resuscitation-promoting factors (Rpf) antigens; IVE-TB antigens; reactivation associated antigens]. Rv1733, which is a probable conserved transmembrane protein encoded in DosR regulon, turned out to be very immunogenic and able to discriminate between the three defined TB status, thus considered a candidate biomarker. Rv2389 and Rv2435n, belonging to Rpf family and IVE-TB group of antigens, respectively, also stood out as LTBI biomarkers. Although more studies are needed to support our findings, the combined use of these antigens would be an interesting approach to TB immunodiagnosis candidates.

  13. Immunogenicity of 60 novel latency-related antigens of Mycobacterium tuberculosis

    Directory of Open Access Journals (Sweden)

    Mªdel Mar eSerra Vidal

    2014-10-01

    Full Text Available The aim of our work here was to evaluate the immunogenicity of 60 mycobacterial antigens, some of which have not been previously assessed, notably a novel series of in vivo-expressed M.tuberculosis (IVE-TB antigens. We enrolled 505 subjects and separated them in individuals with and without latent tuberculosis infection (LTBI versus patients with active tuberculosis. Following an overnight and 7 day stimulation of whole blood with purified recombinant M.tb antigens, interferon-γ (IFN-γ levels were determined by ELISA. Several antigens could statistically significantly differentiate the groups of individuals. We obtained promising antigens from all studied antigen groups (DosR regulon encoded antigens; resuscitation-promoting factors (Rpf antigens; IVE-TB antigens; reactivation asociated antigens. Rv1733, which is a probable conserved transmembrane protein encoded in DosR regulon, turned out to be very immunogenic and able to discriminate between the three defined TB status, thus considered a candidate biomarker. Rv2389 and Rv2435n, belonging to Rpf family and IVE-TB group of antigens, respectively, also stood out as LTBI biomarkers. Although more studies are needed to support our findings, the combined use of these antigens would be an interesting approach to tuberculosis immunodiagnosis candidates.

  14. A user-friendly web portal for analyzing conformational changes in structures of Mycobacterium tuberculosis.

    Science.gov (United States)

    Hassan, Sameer; Thangam, Manonanthini; Vasudevan, Praveen; Kumar, G Ramesh; Unni, Rahul; Devi, P K Gayathri; Hanna, Luke Elizabeth

    2015-10-01

    Initiation of the Tuberculosis Structural Consortium has resulted in the expansion of the Mycobacterium tuberculosis (MTB) protein structural database. Currently, 969 experimentally solved structures are available for 354 MTB proteins. This includes multiple crystal structures for a given protein under different functional conditions, such as the presence of different ligands or mutations. In depth analysis of the multiple structures reveal that subtle differences exist in conformations of a given protein under varied conditions. Therefore, it is immensely important to understand the conformational differences between the multiple structures of a given protein in order to select the most suitable structure for molecular docking and structure-based drug designing. Here, we introduce a web portal ( http://bmi.icmr.org.in/mtbsd/torsion.php ) that we developed to provide comparative data on the ensemble of available structures of MTB proteins, such as Cα root means square deviation (RMSD), sequence identity, presence of mutations and torsion angles. Additionally, torsion angles were used to perform principal component analysis (PCA) to identify the conformational differences between the structures. Additionally, we present a few case studies to demonstrate this database. Graphical Abstract Conformational changes seen in the structures of the enoyl-ACP reductase protein encoded by the Mycobacterial gene inhA.

  15. The microbial diversity of an industrially produced lambic beer shares members of a traditionally produced one and reveals a core microbiota for lambic beer fermentation.

    Science.gov (United States)

    Spitaels, Freek; Wieme, Anneleen D; Janssens, Maarten; Aerts, Maarten; Van Landschoot, Anita; De Vuyst, Luc; Vandamme, Peter

    2015-08-01

    The microbiota involved in lambic beer fermentations in an industrial brewery in West-Flanders, Belgium, was determined through culture-dependent and culture-independent techniques. More than 1300 bacterial and yeast isolates from 13 samples collected during a one-year fermentation process were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry followed by sequence analysis of rRNA and various protein-encoding genes. The bacterial and yeast communities of the same samples were further analyzed using denaturing gradient gel electrophoresis of PCR-amplified V3 regions of the 16S rRNA genes and D1/D2 regions of the 26S rRNA genes, respectively. In contrast to traditional lambic beer fermentations, there was no Enterobacteriaceae phase and a larger variety of acetic acid bacteria were found in industrial lambic beer fermentations. Like in traditional lambic beer fermentations, Saccharomyces cerevisiae, Saccharomyces pastorianus, Dekkera bruxellensis and Pediococcus damnosus were the microorganisms responsible for the main fermentation and maturation phases. These microorganisms originated most probably from the wood of the casks and were considered as the core microbiota of lambic beer fermentations. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. Naturally Occurring Off-Switches for CRISPR-Cas9.

    Science.gov (United States)

    Pawluk, April; Amrani, Nadia; Zhang, Yan; Garcia, Bianca; Hidalgo-Reyes, Yurima; Lee, Jooyoung; Edraki, Alireza; Shah, Megha; Sontheimer, Erik J; Maxwell, Karen L; Davidson, Alan R

    2016-12-15

    CRISPR-Cas9 technology would be enhanced by the ability to inhibit Cas9 function spatially, temporally, or conditionally. Previously, we discovered small proteins encoded by bacteriophages that inhibit the CRISPR-Cas systems of their host bacteria. These "anti-CRISPRs" were specific to type I CRISPR-Cas systems that do not employ the Cas9 protein. We posited that nature would also yield Cas9 inhibitors in response to the evolutionary arms race between bacteriophages and their hosts. Here, we report the discovery of three distinct families of anti-CRISPRs that specifically inhibit the CRISPR-Cas9 system of Neisseria meningitidis. We show that these proteins bind directly to N. meningitidis Cas9 (NmeCas9) and can be used as potent inhibitors of genome editing by this system in human cells. These anti-CRISPR proteins now enable "off-switches" for CRISPR-Cas9 activity and provide a genetically encodable means to inhibit CRISPR-Cas9 genome editing in eukaryotes. VIDEO ABSTRACT. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Nance-Horan syndrome protein, NHS, associates with epithelial cell junctions.

    Science.gov (United States)

    Sharma, Shiwani; Ang, Sharyn L; Shaw, Marie; Mackey, David A; Gécz, Jozef; McAvoy, John W; Craig, Jamie E

    2006-06-15

    Nance-Horan syndrome, characterized by congenital cataracts, craniofacial, dental abnormalities and mental disturbances, is an X-linked disorder with significant phenotypic heterogeneity. Affected individuals have mutations in the NHS (Nance-Horan syndrome) gene typically resulting in premature truncation of the protein. This report underlines the complexity of the regulation of the NHS gene that transcribes several isoforms. We demonstrate the differential expression of the two NHS isoforms, NHS-A and NHS-1A, and differences in the subcellular localization of the proteins encoded by these isoforms. This may in part explain the pleiotropic features of the syndrome. We show that the endogenous and exogenous NHS-A isoform localizes to the cell membrane of mammalian cells in a cell-type-dependent manner and that it co-localizes with the tight junction (TJ) protein ZO-1 in the apical aspect of cell membrane in epithelial cells. We also show that the NHS-1A isoform is a cytoplasmic protein. In the developing mammalian lens, we found continuous expression of NHS that became restricted to the lens epithelium in pre- and postnatal lens. Consistent with the in vitro findings, the NHS-A isoform associates with the apical cell membrane in the lens epithelium. This study suggests that disturbances in intercellular contacts underlie cataractogenesis in the Nance-Horan syndrome. NHS is the first gene localized at TJs that has been implicated in congenital cataracts.

  18. The expanding phenotype of mitochondrial myopathy.

    Science.gov (United States)

    DiMauro, Salvatore; Gurgel-Giannetti, Juliana

    2005-10-01

    Our understanding of mitochondrial diseases (defined restrictively as defects in the mitochondrial respiratory chain) continues to progress apace. In this review we provide an update of information regarding disorders that predominantly or exclusively affect skeletal muscle. Most recently described mitochondrial myopathies are due to defects in nuclear DNA, including coenzyme Q10 deficiency, and mutations in genes that control mitochondrial DNA (mtDNA) abundance and structure such as POLG and TK2. Barth syndrome, an X-linked recessive mitochondrial myopathy/cardiopathy, is associated with altered lipid composition of the inner mitochondrial membrane, but a putative secondary impairment of the respiratory chain remains to be documented. Concerning the 'other genome', the role played by mutations in protein encoding genes of mtDNA in causing isolated myopathies has been confirmed. It has also been confirmed that mutations in tRNA genes of mtDNA can cause predominantly myopathic syndromes and - contrary to conventional wisdom - these mutations can be homoplasmic. Defects in the mitochondrial respiratory chain impair energy production and almost invariably involve skeletal muscle, causing exercise intolerance, myalgia, cramps, or fixed weakness, which often affects extraocular muscles and results in droopy eyelids (ptosis) and progressive external ophthalmoplegia.

  19. Usher syndrome: hearing loss, retinal degeneration and associated abnormalities

    Science.gov (United States)

    Mathur, Pranav; Yang, Jun

    2014-01-01

    Usher syndrome (USH), clinically and genetically heterogeneous, is the leading genetic cause of combined hearing and vision loss. USH is classified into three types, based on the hearing and vestibular symptoms observed in patients. Sixteen loci have been reported to be involved in the occurrence of USH and atypical USH. Among them, twelve have been identified as causative genes and one as a modifier gene. Studies on the proteins encoded by these USH genes suggest that USH proteins interact among one another and function in multiprotein complexes in vivo. Although their exact functions remain enigmatic in the retina, USH proteins are required for the development, maintenance and function of hair bundles, which are the primary mechanosensitive structure of inner ear hair cells. Despite the unavailability of a cure, progress has been made to develop effective treatments for this disease. In this review, we focus on the most recent discoveries in the field with an emphasis on USH genes, protein complexes and functions in various tissues as well as progress toward therapeutic development for USH. PMID:25481835

  20. Transgene-mediated suppression of dengue viruses in the salivary glands of the yellow fever mosquito, Aedes aegypti.

    Science.gov (United States)

    Mathur, G; Sanchez-Vargas, I; Alvarez, D; Olson, K E; Marinotti, O; James, A A

    2010-12-01

    Controlled sex-, stage- and tissue-specific expression of antipathogen effector molecules is important for genetic engineering strategies to control mosquito-borne diseases. Adult female salivary glands are involved in pathogen transmission to human hosts and are target sites for expression of antipathogen effector molecules. The Aedes aegypti 30K a and 30K b genes are expressed exclusively in adult female salivary glands and are transcribed divergently from start sites separated by 263 nucleotides. The intergenic, 5'- and 3'-end untranslated regions of both genes are sufficient to express simultaneously two different transgene products in the distal-lateral lobes of the female salivary glands. An antidengue effector gene, membranes no protein (Mnp), driven by the 30K b promoter, expresses an inverted-repeat RNA with sequences derived from the premembrane protein-encoding region of the dengue virus serotype 2 genome and reduces significantly the prevalence and mean intensities of viral infection in mosquito salivary glands and saliva. © 2010 The Authors. Insect Molecular Biology © 2010 The Royal Entomological Society.

  1. Rubber gloves biodegradation by a consortium, mixed culture and pure culture isolated from soil samples.

    Science.gov (United States)

    Nawong, Chairat; Umsakul, Kamontam; Sermwittayawong, Natthawan

    2018-02-03

    An increasing production of natural rubber (NR) products has led to major challenges in waste management. In this study, the degradation of rubber latex gloves in a mineral salt medium (MSM) using a bacterial consortium, a mixed culture of the selected bacteria and a pure culture were studied. The highest 18% weight loss of the rubber gloves were detected after incubated with the mixed culture. The increased viable cell counts over incubation time indicated that cells used rubber gloves as sole carbon source leading to the degradation of the polymer. The growth behavior of NR-degrading bacteria on the latex gloves surface was investigated using the scanning electron microscope (SEM). The occurrence of the aldehyde groups in the degradation products was observed by Fourier Transform Infrared Spectroscopy analysis. Rhodococcus pyridinivorans strain F5 gave the highest weight loss of rubber gloves among the isolated strain and posses latex clearing protein encoded by lcp gene. The mixed culture of the selected strains showed the potential in degrading rubber within 30 days and is considered to be used efficiently for rubber product degradation. This is the first report to demonstrate a strong ability to degrade rubber by Rhodococcus pyridinivorans. Copyright © 2018. Published by Elsevier Editora Ltda.

  2. Partial duplication of the APBA2 gene in chromosome 15q13 corresponds to duplicon structures

    Directory of Open Access Journals (Sweden)

    Kesterson Robert A

    2003-04-01

    Full Text Available Abstract Background Chromosomal abnormalities affecting human chromosome 15q11-q13 underlie multiple genomic disorders caused by deletion, duplication and triplication of intervals in this region. These events are mediated by highly homologous segments of DNA, or duplicons, that facilitate mispairing and unequal cross-over in meiosis. The gene encoding an amyloid precursor protein-binding protein (APBA2 was previously mapped to the distal portion of the interval commonly deleted in Prader-Willi and Angelman syndromes and duplicated in cases of autism. Results We show that this gene actually maps to a more telomeric location and is partially duplicated within the broader region. Two highly homologous copies of an interval containing a large 5' exon and downstream sequence are located ~5 Mb distal to the intact locus. The duplicated copies, containing the first coding exon of APBA2, can be distinguished by single nucleotide sequence differences and are transcriptionally inactive. Adjacent to APBA2 maps a gene termed KIAA0574. The protein encoded by this gene is weakly homologous to a protein termed X123 that in turn maps adjacent to APBA1 on 9q21.12; APBA1 is highly homologous to APBA2 in the C-terminal region and is distinguished from APBA2 by the N-terminal region encoded by this duplicated exon. Conclusion The duplication of APBA2 sequences in this region adds to a complex picture of different low copy repeats present across this region and elsewhere on the chromosome.

  3. VapC toxins drive cellular dormancy under uranium stress for the extreme thermoacidophile Metallosphaera prunae.

    Science.gov (United States)

    Mukherjee, Arpan; Wheaton, Garrett H; Counts, James A; Ijeomah, Brenda; Desai, Jigar; Kelly, Robert M

    2017-07-01

    When abruptly exposed to toxic levels of hexavalent uranium, the extremely thermoacidophilic archaeon Metallosphaera prunae, originally isolated from an abandoned uranium mine, ceased to grow, and concomitantly exhibited heightened levels of cytosolic ribonuclease activity that corresponded to substantial degradation of cellular RNA. The M. prunae transcriptome during 'uranium-shock' implicated VapC toxins as possible causative agents of the observed RNA degradation. Identifiable VapC toxins and PIN-domain proteins encoded in the M. prunae genome were produced and characterized, three of which (VapC4, VapC7, VapC8) substantially degraded M. prunae rRNA in vitro. RNA cleavage specificity for these VapCs mapped to motifs within M. prunae rRNA. Furthermore, based on frequency of cleavage sequences, putative target mRNAs for these VapCs were identified; these were closely associated with translation, transcription, and replication. It is interesting to note that Metallosphaera sedula, a member of the same genus and which has a nearly identical genome sequence but not isolated from a uranium-rich biotope, showed no evidence of dormancy when exposed to this metal. M. prunae utilizes VapC toxins for post-transcriptional regulation under uranium stress to enter a cellular dormant state, thereby providing an adaptive response to what would otherwise be a deleterious environmental perturbation. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  4. Haploinsufficiency of a spliceosomal GTPase encoded by EFTUD2 causes mandibulofacial dysostosis with microcephaly.

    Science.gov (United States)

    Lines, Matthew A; Huang, Lijia; Schwartzentruber, Jeremy; Douglas, Stuart L; Lynch, Danielle C; Beaulieu, Chandree; Guion-Almeida, Maria Leine; Zechi-Ceide, Roseli Maria; Gener, Blanca; Gillessen-Kaesbach, Gabriele; Nava, Caroline; Baujat, Geneviève; Horn, Denise; Kini, Usha; Caliebe, Almuth; Alanay, Yasemin; Utine, Gulen Eda; Lev, Dorit; Kohlhase, Jürgen; Grix, Arthur W; Lohmann, Dietmar R; Hehr, Ute; Böhm, Detlef; Majewski, Jacek; Bulman, Dennis E; Wieczorek, Dagmar; Boycott, Kym M

    2012-02-10

    Mandibulofacial dysostosis with microcephaly (MFDM) is a rare sporadic syndrome comprising craniofacial malformations, microcephaly, developmental delay, and a recognizable dysmorphic appearance. Major sequelae, including choanal atresia, sensorineural hearing loss, and cleft palate, each occur in a significant proportion of affected individuals. We present detailed clinical findings in 12 unrelated individuals with MFDM; these 12 individuals compose the largest reported cohort to date. To define the etiology of MFDM, we employed whole-exome sequencing of four unrelated affected individuals and identified heterozygous mutations or deletions of EFTUD2 in all four. Validation studies of eight additional individuals with MFDM demonstrated causative EFTUD2 mutations in all affected individuals tested. A range of EFTUD2-mutation types, including null alleles and frameshifts, is seen in MFDM, consistent with haploinsufficiency; segregation is de novo in all cases assessed to date. U5-116kD, the protein encoded by EFTUD2, is a highly conserved spliceosomal GTPase with a central regulatory role in catalytic splicing and post-splicing-complex disassembly. MFDM is the first multiple-malformation syndrome attributed to a defect of the major spliceosome. Our findings significantly extend the range of reported spliceosomal phenotypes in humans and pave the way for further investigation in related conditions such as Treacher Collins syndrome. Copyright © 2012 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

  5. Cloning, purification and preliminary crystallographic analysis of a conserved hypothetical protein, SA0961 (YlaN), from Staphylococcus aureus

    International Nuclear Information System (INIS)

    Xu, Ling; Sedelnikova, Svetlana E.; Baker, Patrick J.; Rice, David W.

    2006-01-01

    SA0961 is an unknown hypothetical protein from Staphylococcus aureus that can be identified in the Firmicutes division of Gram-positive bacteria. SA0961 was cloned and the protein was overexpressed in Escherichia coli, purified and subsequently crystallized. SA0961 is an unknown hypothetical protein from Staphylococcus aureus that can be identified in the Firmicutes division of Gram-positive bacteria. The gene for the homologue of SA0961 in Bacillus subtilis, ylaN, has been shown to be essential for cell survival, thus identifying the protein encoded by this gene as a potential target for the development of novel antibiotics. SA0961 was cloned and the protein was overexpressed in Escherichia coli, purified and subsequently crystallized. Crystals of selenomethionine-labelled SA0961 diffract to beyond 2.4 Å resolution and belong to the monoclinic space group P2 1 , with unit-cell parameters a = 31.5, b = 42.7, c = 62.7 Å, β = 92.4° and two molecules in the asymmetric unit. A full structure determination is under way to provide insights into the function of this protein

  6. Cloning, purification and preliminary crystallographic analysis of a conserved hypothetical protein, SA0961 (YlaN), from Staphylococcus aureus

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Ling; Sedelnikova, Svetlana E.; Baker, Patrick J.; Rice, David W., E-mail: d.rice@sheffield.ac.uk [Krebs Institute for Biomolecular Research, Department of Molecular Biology and Biotechnology, The University of Sheffield, Sheffield S10 2TN (United Kingdom)

    2006-08-01

    SA0961 is an unknown hypothetical protein from Staphylococcus aureus that can be identified in the Firmicutes division of Gram-positive bacteria. SA0961 was cloned and the protein was overexpressed in Escherichia coli, purified and subsequently crystallized. SA0961 is an unknown hypothetical protein from Staphylococcus aureus that can be identified in the Firmicutes division of Gram-positive bacteria. The gene for the homologue of SA0961 in Bacillus subtilis, ylaN, has been shown to be essential for cell survival, thus identifying the protein encoded by this gene as a potential target for the development of novel antibiotics. SA0961 was cloned and the protein was overexpressed in Escherichia coli, purified and subsequently crystallized. Crystals of selenomethionine-labelled SA0961 diffract to beyond 2.4 Å resolution and belong to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 31.5, b = 42.7, c = 62.7 Å, β = 92.4° and two molecules in the asymmetric unit. A full structure determination is under way to provide insights into the function of this protein.

  7. A plasmodesmata-associated beta-1,3-glucanase in Arabidopsis.

    Science.gov (United States)

    Levy, Amit; Erlanger, Michael; Rosenthal, Michal; Epel, Bernard L

    2007-02-01

    Plasmodesmal conductivity is regulated in part by callose turnover, which is hypothesized to be determined by beta-1,3-glucan synthase versus glucanase activities. A proteomic analysis of an Arabidopsis thaliana plasmodesmata (Pd)-rich fraction identified a beta-1,3-glucanase as present in this fraction. The protein encoded by the putative plasmodesmal associated protein (ppap) gene, termed AtBG_ppap, had previously been found to be a post-translationally modified glycosylphosphatidylinositol (GPI) lipid-anchored protein. When fused to green fluorescent protein (GFP) and expressed in tobacco (Nicotiana tabacum) or Nicotiana benthamiana epidermal cells, this protein displays fluorescence patterns in the endoplasmic reticulum (ER) membrane system, along the cell periphery and in a punctate pattern that co-localizes with aniline blue-stained callose present around the Pd. Plasma membrane localization was verified by co-localization of AtBG_ppap:GFP together with a plasma membrane marker N-[3-triethylammoniumpropyl]-4-[p-diethylaminophenylhexatrienyl] pyridinium dibromide (FM4-64) in plasmolysed cells. In Arabidopsis T-DNA insertion mutants that do not transcribe AtBG_ppap, functional studies showed that GFP cell-to-cell movement between epidermal cells is reduced, and the conductivity coefficient of Pd is lower. Measurements of callose levels around Pd after wounding revealed that callose accumulation in the mutant plants was higher. Taken together, we suggest that AtBG_ppap is a Pd-associated membrane protein involved in plasmodesmal callose degradation, and functions in the gating of Pd.

  8. α-Hydroxyketone Synthesis and Sensing by Legionella and Vibrio

    Directory of Open Access Journals (Sweden)

    Hubert Hilbi

    2012-03-01

    Full Text Available Bacteria synthesize and sense low molecular weight signaling molecules, termed autoinducers, to measure their population density and community complexity. One class of autoinducers, the α-hydroxyketones (AHKs, is produced and detected by the water-borne opportunistic pathogens Legionella pneumophila and Vibrio cholerae, which cause Legionnaires’ disease and cholera, respectively. The “Legionella quorum sensing” (lqs or “cholera quorum sensing” (cqs genes encode enzymes that produce and sense the AHK molecules “Legionella autoinducer-1” (LAI-1; 3-hydroxypentadecane-4-one or cholera autoinducer-1 (CAI-1; 3-hydroxytridecane-4-one. AHK signaling regulates the virulence of L. pneumophila and V. cholerae, pathogen-host cell interactions, formation of biofilms or extracellular filaments, expression of a genomic “fitness island” and competence. Here, we outline the processes, wherein AHK signaling plays a role, and review recent insights into the function of proteins encoded by the lqs and cqs gene clusters. To this end, we will focus on the autoinducer synthases catalysing the biosynthesis of AHKs, on the cognate trans-membrane sensor kinases detecting the signals, and on components of the down-stream phosphorelay cascade that promote the transmission and integration of signaling events regulating gene expression.

  9. Long Noncoding RNAs in Lung Cancer.

    Science.gov (United States)

    Roth, Anna; Diederichs, Sven

    2016-01-01

    Despite great progress in research and treatment options, lung cancer remains the leading cause of cancer-related deaths worldwide. Oncogenic driver mutations in protein-encoding genes were defined and allow for personalized therapies based on genetic diagnoses. Nonetheless, diagnosis of lung cancer mostly occurs at late stages, and chronic treatment is followed by a fast onset of chemoresistance. Hence, there is an urgent need for reliable biomarkers and alternative treatment options. With the era of whole genome and transcriptome sequencing technologies, long noncoding RNAs emerged as a novel class of versatile, functional RNA molecules. Although for most of them the mechanism of action remains to be defined, accumulating evidence confirms their involvement in various aspects of lung tumorigenesis. They are functional on the epigenetic, transcriptional, and posttranscriptional level and are regulators of pathophysiological key pathways including cell growth, apoptosis, and metastasis. Long noncoding RNAs are gaining increasing attention as potential biomarkers and a novel class of druggable molecules. It has become clear that we are only beginning to understand the complexity of tumorigenic processes. The clinical integration of long noncoding RNAs in terms of prognostic and predictive biomarker signatures and additional cancer targets could provide a chance to increase the therapeutic benefit. Here, we review the current knowledge about the expression, regulation, biological function, and clinical relevance of long noncoding RNAs in lung cancer.

  10. Nucleotide sequences of immunoglobulin eta genes of chimpanzee and orangutan: DNA molecular clock and hominoid evolution

    Energy Technology Data Exchange (ETDEWEB)

    Sakoyama, Y.; Hong, K.J.; Byun, S.M.; Hisajima, H.; Ueda, S.; Yaoita, Y.; Hayashida, H.; Miyata, T.; Honjo, T.

    1987-02-01

    To determine the phylogenetic relationships among hominoids and the dates of their divergence, the complete nucleotide sequences of the constant region of the immunoglobulin eta-chain (C/sub eta1/) genes from chimpanzee and orangutan have been determined. These sequences were compared with the human eta-chain constant-region sequence. A molecular clock (silent molecular clock), measured by the degree of sequence divergence at the synonymous (silent) positions of protein-encoding regions, was introduced for the present study. From the comparison of nucleotide sequences of ..cap alpha../sub 1/-antitrypsin and ..beta..- and delta-globulin genes between humans and Old World monkeys, the silent molecular clock was calibrated: the mean evolutionary rate of silent substitution was determined to be 1.56 x 10/sup -9/ substitutions per site per year. Using the silent molecular clock, the mean divergence dates of chimpanzee and orangutan from the human lineage were estimated as 6.4 +/- 2.6 million years and 17.3 +/- 4.5 million years, respectively. It was also shown that the evolutionary rate of primate genes is considerably slower than those of other mammalian genes.

  11. Design and construction of a first-generation high-throughput integrated robotic molecular biology platform for bioenergy applications.

    Science.gov (United States)

    Hughes, Stephen R; Butt, Tauseef R; Bartolett, Scott; Riedmuller, Steven B; Farrelly, Philip

    2011-08-01

    The molecular biological techniques for plasmid-based assembly and cloning of gene open reading frames are essential for elucidating the function of the proteins encoded by the genes. High-throughput integrated robotic molecular biology platforms that have the capacity to rapidly clone and express heterologous gene open reading frames in bacteria and yeast and to screen large numbers of expressed proteins for optimized function are an important technology for improving microbial strains for biofuel production. The process involves the production of full-length complementary DNA libraries as a source of plasmid-based clones to express the desired proteins in active form for determination of their functions. Proteins that were identified by high-throughput screening as having desired characteristics are overexpressed in microbes to enable them to perform functions that will allow more cost-effective and sustainable production of biofuels. Because the plasmid libraries are composed of several thousand unique genes, automation of the process is essential. This review describes the design and implementation of an automated integrated programmable robotic workcell capable of producing complementary DNA libraries, colony picking, isolating plasmid DNA, transforming yeast and bacteria, expressing protein, and performing appropriate functional assays. These operations will allow tailoring microbial strains to use renewable feedstocks for production of biofuels, bioderived chemicals, fertilizers, and other coproducts for profitable and sustainable biorefineries. Published by Elsevier Inc.

  12. Trypanosoma brucei Tb927.2.6100 Is an Essential Protein Associated with Kinetoplast DNA

    KAUST Repository

    Beck, K.

    2013-05-06

    The mitochondrial DNA of trypanosomatid protozoa consists of a complex, intercatenated network of tens of maxicircles and thousands of minicircles. This structure, called kinetoplast DNA (kDNA), requires numerous proteins and multiprotein complexes for replication, segregation, and transcription. In this study, we used a proteomic approach to identify proteins that are associated with the kDNA network. We identified a novel protein encoded by Tb927.2.6100 that was present in a fraction enriched for kDNA and colocalized the protein with kDNA by fluorescence microscopy. RNA interference (RNAi) knockdown of its expression resulted in a growth defect and changes in the proportion of kinetoplasts and nuclei in the cell population. RNAi also resulted in shrinkage and loss of the kinetoplasts, loss of maxicircle and minicircle components of kDNA at similar rates, and (perhaps secondarily) loss of edited and pre-edited mRNA. These results indicate that the Tb927.2.6100 protein is essential for the maintenance of kDNA.

  13. High-yield nontoxic gene transfer through conjugation of the CM₁₈-Tat₁₁ chimeric peptide with nanosecond electric pulses.

    Science.gov (United States)

    Salomone, Fabrizio; Breton, Marie; Leray, Isabelle; Cardarelli, Francesco; Boccardi, Claudia; Bonhenry, Daniel; Tarek, Mounir; Mir, Lluis M; Beltram, Fabio

    2014-07-07

    We report a novel nontoxic, high-yield, gene delivery system based on the synergistic use of nanosecond electric pulses (NPs) and nanomolar doses of the recently introduced CM18-Tat11 chimeric peptide (sequence of KWKLFKKIGAVLKVLTTGYGRKKRRQRRR, residues 1-7 of cecropin-A, 2-12 of melittin, and 47-57 of HIV-1 Tat protein). This combined use makes it possible to drastically reduce the required CM18-Tat11 concentration and confines stable nanopore formation to vesicle membranes followed by DNA release, while no detectable perturbation of the plasma membrane is observed. Two different experimental assays are exploited to quantitatively evaluate the details of NPs and CM18-Tat11 cooperation: (i) cytofluorimetric analysis of the integrity of synthetic 1,2-dioleoyl-sn-glycero-3-phosphocholine giant unilamellar vesicles exposed to CM18-Tat11 and NPs and (ii) the in vitro transfection efficiency of a green fluorescent protein-encoding plasmid conjugated to CM18-Tat11 in the presence of NPs. Data support a model in which NPs induce membrane perturbation in the form of transient pores on all cellular membranes, while the peptide stabilizes membrane defects selectively within endosomes. Interestingly, atomistic molecular dynamics simulations show that the latter activity can be specifically attributed to the CM18 module, while Tat11 remains essential for cargo binding and vector subcellular localization. We argue that this result represents a paradigmatic example that can open the way to other targeted delivery protocols.

  14. Partial Least Squares Based Gene Expression Analysis in EBV- Positive and EBV-Negative Posttransplant Lymphoproliferative Disorders.

    Science.gov (United States)

    Wu, Sa; Zhang, Xin; Li, Zhi-Ming; Shi, Yan-Xia; Huang, Jia-Jia; Xia, Yi; Yang, Hang; Jiang, Wen-Qi

    2013-01-01

    Post-transplant lymphoproliferative disorder (PTLD) is a common complication of therapeutic immunosuppression after organ transplantation. Gene expression profile facilitates the identification of biological difference between Epstein-Barr virus (EBV) positive and negative PTLDs. Previous studies mainly implemented variance/regression analysis without considering unaccounted array specific factors. The aim of this study is to investigate the gene expression difference between EBV positive and negative PTLDs through partial least squares (PLS) based analysis. With a microarray data set from the Gene Expression Omnibus database, we performed PLS based analysis. We acquired 1188 differentially expressed genes. Pathway and Gene Ontology enrichment analysis identified significantly over-representation of dysregulated genes in immune response and cancer related biological processes. Network analysis identified three hub genes with degrees higher than 15, including CREBBP, ATXN1, and PML. Proteins encoded by CREBBP and PML have been reported to be interact with EBV before. Our findings shed light on expression distinction of EBV positive and negative PTLDs with the hope to offer theoretical support for future therapeutic study.

  15. Streptococcus iniae SF1: complete genome sequence, proteomic profile, and immunoprotective antigens.

    Directory of Open Access Journals (Sweden)

    Bao-cun Zhang

    Full Text Available Streptococcus iniae is a Gram-positive bacterium that is reckoned one of the most severe aquaculture pathogens. It has a broad host range among farmed marine and freshwater fish and can also cause zoonotic infection in humans. Here we report for the first time the complete genome sequence as well as the host factor-induced proteomic profile of a pathogenic S. iniae strain, SF1, a serotype I isolate from diseased fish. SF1 possesses a single chromosome of 2,149,844 base pairs, which contains 2,125 predicted protein coding sequences (CDS, 12 rRNA genes, and 45 tRNA genes. Among the protein-encoding CDS are genes involved in resource acquisition and utilization, signal sensing and transduction, carbohydrate metabolism, and defense against host immune response. Potential virulence genes include those encoding adhesins, autolysins, toxins, exoenzymes, and proteases. In addition, two putative prophages and a CRISPR-Cas system were found in the genome, the latter containing a CRISPR locus and four cas genes. Proteomic analysis detected 21 secreted proteins whose expressions were induced by host serum. Five of the serum-responsive proteins were subjected to immunoprotective analysis, which revealed that two of the proteins were highly protective against lethal S. iniae challenge when used as purified recombinant subunit vaccines. Taken together, these results provide an important molecular basis for future study of S. iniae in various aspects, in particular those related to pathogenesis and disease control.

  16. Molecular epidemiology of coxsackievirus A16: intratype and prevalent intertype recombination identified.

    Directory of Open Access Journals (Sweden)

    Xiangpeng Chen

    Full Text Available Coxsackievirus A16 (CVA16 is responsible for nearly 50% of all the confirmed hand, foot, and mouth disease (HFMD cases in mainland China, sometimes it could also cause severe complications, and even death. To clarify the genetic characteristics and the epidemic patterns of CVA16 in mainland China, comprehensive bioinfomatics analyses were performed by using 35 CVA16 whole genome sequences from 1998 to 2011, 593 complete CVA16 VP1 sequences from 1981 to 2011, and prototype strains of human enterovirus species A (EV-A. Analysis on complete VP1 sequences revealed that subgenotypes B1a and B1b were prevalent strains and have been co-circulating in many Asian countries since 2000, especially in mainland China for at least 13 years. While the prevalence of subgenotype B1c (totally 20 strains was much limited, only found in Malaysia from 2005 to 2007 and in France in 2010. Genotype B2 only caused epidemic in Japan and Malaysia from 1981 to 2000. Both subgenotypes B1a and B1b were potential recombinant viruses containing sequences from other EV-A donors in the 5'-untranslated region and P2, P3 non-structural protein encoding regions.

  17. Transverse relaxation dispersion of the p7 membrane channel from hepatitis C virus reveals conformational breathing

    Energy Technology Data Exchange (ETDEWEB)

    Dev, Jyoti; Brüschweiler, Sven [Harvard Medical School, Department of Biological Chemistry and Molecular Pharmacology (United States); Ouyang, Bo [Chinese Academy of Sciences, State Key Laboratory of Molecular Biology, Shanghai Institute of Biochemistry and Cell Biology (China); Chou, James J., E-mail: james-chou@hms.harvard.edu [Harvard Medical School, Department of Biological Chemistry and Molecular Pharmacology (United States)

    2015-04-15

    The p7 membrane protein encoded by hepatitis C virus (HCV) assembles into a homo-hexamer that selectively conducts cations. An earlier solution NMR structure of the hexameric complex revealed a funnel-like architecture and suggests that a ring of conserved asparagines near the narrow end of the funnel are important for cation interaction. NMR based drug-binding experiments also suggest that rimantadine can allosterically inhibit ion conduction via a molecular wedge mechanism. These results suggest the presence of dilation and contraction of the funnel tip that are important for channel activity and that the action of the drug is attenuating this motion. Here, we determined the conformational dynamics and solvent accessibility of the p7 channel. The proton exchange measurements show that the cavity-lining residues are largely water accessible, consistent with the overall funnel shape of the channel. Our relaxation dispersion data show that residues Val7 and Leu8 near the asparagine ring are subject to large chemical exchange, suggesting significant intrinsic channel breathing at the tip of the funnel. Moreover, the hinge regions connecting the narrow and wide regions of the funnel show strong relaxation dispersion and these regions are the binding sites for rimantadine. Presence of rimantadine decreases the conformational dynamics near the asparagine ring and the hinge area. Our data provide direct observation of μs–ms dynamics of the p7 channel and support the molecular wedge mechanism of rimantadine inhibition of the HCV p7 channel.

  18. Whole Genome and Global Gene Expression Analyses of the Model Mushroom Flammulina velutipes Reveal a High Capacity for Lignocellulose Degradation

    Science.gov (United States)

    Park, Young-Jin; Baek, Jeong Hun; Lee, Seonwook; Kim, Changhoon; Rhee, Hwanseok; Kim, Hyungtae; Seo, Jeong-Sun; Park, Hae-Ran; Yoon, Dae-Eun; Nam, Jae-Young; Kim, Hong-Il; Kim, Jong-Guk; Yoon, Hyeokjun; Kang, Hee-Wan; Cho, Jae-Yong; Song, Eun-Sung; Sung, Gi-Ho; Yoo, Young-Bok; Lee, Chang-Soo; Lee, Byoung-Moo; Kong, Won-Sik

    2014-01-01

    Flammulina velutipes is a fungus with health and medicinal benefits that has been used for consumption and cultivation in East Asia. F. velutipes is also known to degrade lignocellulose and produce ethanol. The overlapping interests of mushroom production and wood bioconversion make F. velutipes an attractive new model for fungal wood related studies. Here, we present the complete sequence of the F. velutipes genome. This is the first sequenced genome for a commercially produced edible mushroom that also degrades wood. The 35.6-Mb genome contained 12,218 predicted protein-encoding genes and 287 tRNA genes assembled into 11 scaffolds corresponding with the 11 chromosomes of strain KACC42780. The 88.4-kb mitochondrial genome contained 35 genes. Well-developed wood degrading machinery with strong potential for lignin degradation (69 auxiliary activities, formerly FOLymes) and carbohydrate degradation (392 CAZymes), along with 58 alcohol dehydrogenase genes were highly expressed in the mycelium, demonstrating the potential application of this organism to bioethanol production. Thus, the newly uncovered wood degrading capacity and sequential nature of this process in F. velutipes, offer interesting possibilities for more detailed studies on either lignin or (hemi-) cellulose degradation in complex wood substrates. The mutual interest in wood degradation by the mushroom industry and (ligno-)cellulose biomass related industries further increase the significance of F. velutipes as a new model. PMID:24714189

  19. Direct transfer of viral and cellular proteins from varicella-zoster virus-infected non-neuronal cells to human axons.

    Science.gov (United States)

    Grigoryan, Sergei; Yee, Michael B; Glick, Yair; Gerber, Doron; Kepten, Eldad; Garini, Yuval; Yang, In Hong; Kinchington, Paul R; Goldstein, Ronald S

    2015-01-01

    Varicella Zoster Virus (VZV), the alphaherpesvirus that causes varicella upon primary infection and Herpes zoster (shingles) following reactivation in latently infected neurons, is known to be fusogenic. It forms polynuclear syncytia in culture, in varicella skin lesions and in infected fetal human ganglia xenografted to mice. After axonal infection using VZV expressing green fluorescent protein (GFP) in compartmentalized microfluidic cultures there is diffuse filling of axons with GFP as well as punctate fluorescence corresponding to capsids. Use of viruses with fluorescent fusions to VZV proteins reveals that both proteins encoded by VZV genes and those of the infecting cell are transferred in bulk from infecting non-neuronal cells to axons. Similar transfer of protein to axons was observed following cell associated HSV1 infection. Fluorescence recovery after photobleaching (FRAP) experiments provide evidence that this transfer is by diffusion of proteins from the infecting cells into axons. Time-lapse movies and immunocytochemical experiments in co-cultures demonstrate that non-neuronal cells fuse with neuronal somata and proteins from both cell types are present in the syncytia formed. The fusogenic nature of VZV therefore may enable not only conventional entry of virions and capsids into axonal endings in the skin by classical entry mechanisms, but also by cytoplasmic fusion that permits viral protein transfer to neurons in bulk.

  20. Computational analysis of EBNA1 ``druggability'' suggests novel insights for Epstein-Barr virus inhibitor design

    Science.gov (United States)

    Gianti, Eleonora; Messick, Troy E.; Lieberman, Paul M.; Zauhar, Randy J.

    2016-04-01

    The Epstein-Barr Nuclear Antigen 1 (EBNA1) is a critical protein encoded by the Epstein-Barr Virus (EBV). During latent infection, EBNA1 is essential for DNA replication and transcription initiation of viral and cellular genes and is necessary to immortalize primary B-lymphocytes. Nonetheless, the concept of EBNA1 as drug target is novel. Two EBNA1 crystal structures are publicly available and the first small-molecule EBNA1 inhibitors were recently discovered. However, no systematic studies have been reported on the structural details of EBNA1 "druggable" binding sites. We conducted computational identification and structural characterization of EBNA1 binding pockets, likely to accommodate ligand molecules (i.e. "druggable" binding sites). Then, we validated our predictions by docking against a set of compounds previously tested in vitro for EBNA1 inhibition (PubChem AID-2381). Finally, we supported assessments of pocket druggability by performing induced fit docking and molecular dynamics simulations paired with binding affinity predictions by Molecular Mechanics Generalized Born Surface Area calculations for a number of hits belonging to druggable binding sites. Our results establish EBNA1 as a target for drug discovery, and provide the computational evidence that active AID-2381 hits disrupt EBNA1:DNA binding upon interacting at individual sites. Lastly, structural properties of top scoring hits are proposed to support the rational design of the next generation of EBNA1 inhibitors.

  1. A novel missense mutation in the gene EDARADD associated with an unusual phenotype of hypohidrotic ectodermal dysplasia.

    Science.gov (United States)

    Wohlfart, Sigrun; Söder, Stephan; Smahi, Asma; Schneider, Holm

    2016-01-01

    Hypohidrotic ectodermal dysplasia (HED) is a rare disorder characterized by deficient development of structures derived from the ectoderm including hair, nails, eccrine glands, and teeth. HED forms that are caused by mutations in the genes EDA, EDAR, or EDARADD may show almost identical phenotypes, explained by a common signaling pathway. Proper interaction of the proteins encoded by these three genes is important for the activation of the NF-κB signaling pathway and subsequent transcription of the target genes. Mutations in the gene EDARADD are most rarely implicated in HED. Here we describe a novel missense mutation, c.367G>A (p.Asp123Asn), in this gene which did not appear to influence the interaction between EDAR and EDARADD proteins, but led to an impaired ability to activate NF-κB signaling. Female members of the affected family showed either unilateral or bilateral amazia. In addition, an affected girl developed bilateral ovarian teratomas, possibly associated with her genetic condition. © 2015 Wiley Periodicals, Inc.

  2. The Gcn4 transcription factor reduces protein synthesis capacity and extends yeast lifespan.

    Science.gov (United States)

    Mittal, Nitish; Guimaraes, Joao C; Gross, Thomas; Schmidt, Alexander; Vina-Vilaseca, Arnau; Nedialkova, Danny D; Aeschimann, Florian; Leidel, Sebastian A; Spang, Anne; Zavolan, Mihaela

    2017-09-06

    In Saccharomyces cerevisiae, deletion of large ribosomal subunit protein-encoding genes increases the replicative lifespan in a Gcn4-dependent manner. However, how Gcn4, a key transcriptional activator of amino acid biosynthesis genes, increases lifespan, is unknown. Here we show that Gcn4 acts as a repressor of protein synthesis. By analyzing the messenger RNA and protein abundance, ribosome occupancy and protein synthesis rate in various yeast strains, we demonstrate that Gcn4 is sufficient to reduce protein synthesis and increase yeast lifespan. Chromatin immunoprecipitation reveals Gcn4 binding not only at genes that are activated, but also at genes, some encoding ribosomal proteins, that are repressed upon Gcn4 overexpression. The promoters of repressed genes contain Rap1 binding motifs. Our data suggest that Gcn4 is a central regulator of protein synthesis under multiple perturbations, including ribosomal protein gene deletions, calorie restriction, and rapamycin treatment, and provide an explanation for its role in longevity and stress response.The transcription factor Gcn4 is known to regulate yeast amino acid synthesis. Here, the authors show that Gcn4 also acts as a repressor of protein biosynthesis in a range of conditions that enhance yeast lifespan, such as ribosomal protein knockout, calorie restriction or mTOR inhibition.

  3. Porcine bocavirus NP1 negatively regulates interferon signaling pathway by targeting the DNA-binding domain of IRF9

    International Nuclear Information System (INIS)

    Zhang, Ruoxi; Fang, Liurong; Wang, Dang; Cai, Kaimei; Zhang, Huan; Xie, Lilan; Li, Yi; Chen, Huanchun; Xiao, Shaobo

    2015-01-01

    To subvert host antiviral immune responses, many viruses have evolved countermeasures to inhibit IFN signaling pathway. Porcine bocavirus (PBoV), a newly identified porcine parvovirus, has received attention because it shows clinically high co-infection prevalence with other pathogens in post-weaning multisystemic wasting syndrome (PWMS) and diarrheic piglets. In this study, we screened the structural and non-structural proteins encoded by PBoV and found that the non-structural protein NP1 significantly suppressed IFN-stimulated response element (ISRE) activity and subsequent IFN-stimulated gene (ISG) expression. However, NP1 affected neither the activation and translocation of STAT1/STAT2, nor the formation of the heterotrimeric transcription factor complex ISGF3 (STAT1/STAT2/IRF9). Detailed analysis demonstrated that PBoV NP1 blocked the ISGF3 DNA-binding activity by combining with the DNA-binding domain (DBD) of IRF9. In summary, these results indicate that PBoV NP1 interferes with type I IFN signaling pathway by blocking DNA binding of ISGF3 to attenuate innate immune responses. - Highlights: • Porcine bocavirus (PBoV) NP1 interferes with the IFN α/β signaling pathway. • PBoV NP1 does not prevent STAT1/STAT2 phosphorylation and nuclear translocation. • PBoV NP1 inhibits the DNA-binding activity of ISGF3. • PBoV NP1 interacts with the DNA-binding domain of IRF9.

  4. Porcine bocavirus NP1 negatively regulates interferon signaling pathway by targeting the DNA-binding domain of IRF9

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Ruoxi [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China); Fang, Liurong, E-mail: fanglr@mail.hzau.edu.cn [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China); Wang, Dang; Cai, Kaimei; Zhang, Huan [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China); Xie, Lilan; Li, Yi [College of Life Science and Technology, Wuhan Institute of Bioengineering, Wuhan 430415 (China); Chen, Huanchun; Xiao, Shaobo [State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070 (China); The Cooperative Innovation Center for Sustainable Pig Production, Wuhan 430070 (China)

    2015-11-15

    To subvert host antiviral immune responses, many viruses have evolved countermeasures to inhibit IFN signaling pathway. Porcine bocavirus (PBoV), a newly identified porcine parvovirus, has received attention because it shows clinically high co-infection prevalence with other pathogens in post-weaning multisystemic wasting syndrome (PWMS) and diarrheic piglets. In this study, we screened the structural and non-structural proteins encoded by PBoV and found that the non-structural protein NP1 significantly suppressed IFN-stimulated response element (ISRE) activity and subsequent IFN-stimulated gene (ISG) expression. However, NP1 affected neither the activation and translocation of STAT1/STAT2, nor the formation of the heterotrimeric transcription factor complex ISGF3 (STAT1/STAT2/IRF9). Detailed analysis demonstrated that PBoV NP1 blocked the ISGF3 DNA-binding activity by combining with the DNA-binding domain (DBD) of IRF9. In summary, these results indicate that PBoV NP1 interferes with type I IFN signaling pathway by blocking DNA binding of ISGF3 to attenuate innate immune responses. - Highlights: • Porcine bocavirus (PBoV) NP1 interferes with the IFN α/β signaling pathway. • PBoV NP1 does not prevent STAT1/STAT2 phosphorylation and nuclear translocation. • PBoV NP1 inhibits the DNA-binding activity of ISGF3. • PBoV NP1 interacts with the DNA-binding domain of IRF9.

  5. Non-dispersive phloem-protein bodies (NPBs of Populus trichocarpa consist of a SEOR protein and do not respond to cell wounding and Ca2+

    Directory of Open Access Journals (Sweden)

    Daniel L. Mullendore

    2018-04-01

    Full Text Available Differentiating sieve elements in the phloem of angiosperms produce abundant phloem-specific proteins before their protein synthesis machinery is degraded. These P-proteins initially form dense bodies, which disperse into individual filaments when the sieve element matures. In some cases, however, the dense protein agglomerations remain intact and are visible in functional sieve tubes as non-dispersive P-protein bodies, or NPBs. Species exhibiting NPBs are distributed across the entire angiosperm clade. We found that NPBs in the model tree, Populus trichocarpa, resemble the protein bodies described from other species of the order Malpighiales as they all consist of coaligned tubular fibrils bundled in hexagonal symmetry. NPBs of all Malpighiales tested proved unresponsive to sieve tube wounding and Ca2+. The P. trichocarpa NPBs consisted of a protein encoded by a gene that in the genome database of this species had been annotated as a homolog of SEOR1 (sieve element occlusion-related 1 in Arabidopsis. Sequencing of the gene in our plants corroborated this interpretation, and we named the gene PtSEOR1. Previously characterized SEOR proteins form irregular masses of P-protein slime in functional sieve tubes. We conclude that a subgroup of these proteins is involved in the formation of NPBs at least in the Malpighiales, and that these protein bodies have no role in rapid wound responses of the sieve tube network.

  6. BIIDXI, the At4g32460 DUF642 gene, is involved in pectin methyl esterase regulation during Arabidopsis thaliana seed germination and plant development.

    Science.gov (United States)

    Zúñiga-Sánchez, Esther; Soriano, Diana; Martínez-Barajas, Eleazar; Orozco-Segovia, Alma; Gamboa-deBuen, Alicia

    2014-12-02

    DUF642 proteins constitute a highly conserved family of proteins that are associated with the cell wall and are specific to spermatophytes. Transcriptome studies have suggested that members of this family are involved in seed development and germination processes. Previous in vitro studies have revealed that At4g32460- and At5g11420-encoded proteins interact with the catalytic domain of pectin methyl esterase 3 (AtPME3, which is encoded by At3g14310). PMEs play an important role in plant development, including seed germination. The aim of this study was to evaluate the function of the DUF642 gene At4g32460 during seed germination and plant development and to determine its relation to PME activity regulation. Our results indicated that the DUF642 proteins encoded by At4g32460 and At5g11420 could be positive regulators of PME activity during several developmental processes. Transgenic lines overexpressing these proteins showed increased PME activity during seed germination, and improved seed germination performance. In plants expressing At4g32460 antisense RNA, PME activity was decreased in the leaves, and the siliques were very short and contained no seeds. This phenotype was also present in the SALK_142260 and SALK_054867 lines for At4g32460. Our results suggested that the DUF642 family contributes to the complexity of the methylesterification process by participating in the fine regulation of pectin status during plant development.

  7. Pheromones and pheromone receptors are required for proper sexual development in the homothallic ascomycete Sordaria macrospora.

    Science.gov (United States)

    Mayrhofer, Severine; Weber, Jan M; Pöggeler, Stefanie

    2006-03-01

    The homothallic, filamentous ascomycete Sordaria macrospora is self-fertile and produces sexual fruiting bodies (perithecia) without a mating partner. Even so, S. macrospora transcriptionally expresses two pheromone-precursor genes (ppg1 and ppg2) and two pheromone-receptor genes (pre1 and pre2). The proteins encoded by these genes are similar to alpha-factor-like and a-factor-like pheromones and to G-protein-coupled pheromone receptors of the yeast Saccharomyces cerevisiae. It has been suggested that in S. macrospora, PPG1/PRE2 and PPG2/PRE1 form two cognate pheromone-receptor pairs. To investigate their function, we deleted (delta) pheromone-precursor genes (delta ppg1, delta ppg2) and receptor genes (delta pre1, delta pre2) and generated single- as well as double-knockout strains. No effect on vegetative growth, fruiting-body, and ascospore development was seen in the single pheromone-mutant and receptor-mutant strains, respectively. However, double-knockout strains lacking any compatible pheromone-receptor pair (delta pre2/delta ppg2, delta pre1/delta ppg1) and the double-pheromone mutant (delta ppg1/delta ppg2) displayed a drastically reduced number of perithecia and sexual spores, whereas deletion of both receptor genes (delta pre1/delta pre2) completely eliminated fruiting-body and ascospore formation. The results suggest that pheromones and pheromone receptors are required for optimal sexual reproduction of the homothallic S. macrospora.

  8. Quantitative Proteomic Analysis of Staphylococcus aureus Treated With Punicalagin, a Natural Antibiotic From Pomegranate That Disrupts Iron Homeostasis and Induces SOS.

    Science.gov (United States)

    Cooper, Bret; Islam, Nazrul; Xu, Yunfeng; Beard, Hunter S; Garrett, Wesley M; Gu, Ganyu; Nou, Xiangwu

    2018-05-01

    Staphylococcus aureus, a bacterial, food-borne pathogen of humans, can contaminate raw fruits and vegetables. While physical and chemical methods are available to control S. aureus, scientists are searching for inhibitory phytochemicals from plants. One promising compound from pomegranate is punicalagin, a natural antibiotic. To get a broader understanding of the inhibitory effect of punicalagin on S. aureus growth, high-throughput mass spectrometry and quantitative isobaric labeling was used to investigate the proteome of S. aureus after exposure to a sublethal dose of punicalagin. Nearly half of the proteins encoded by the small genome were interrogated, and nearly half of those exhibited significant changes in accumulation. Punicalagin treatment altered the accumulation of proteins and enzymes needed for iron acquisition, and it altered amounts of enzymes for glycolysis, citric acid cycling, protein biosynthesis, and purine and pyrimidine biosynthesis. Punicalagin treatment also induced an SOS cellular response to damaged DNA. Transcriptional comparison of marker genes shows that the punicalagin-induced iron starvation and SOS responses resembles those produced by EDTA and ciprofloxacin. These results show that punicalagin adversely alters bacterial growth by disrupting iron homeostasis and that it induces SOS, possibly through DNA biosynthesis inhibition. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Structure of a short-chain dehydrogenase/reductase (SDR) within a genomic island from a clinical strain of Acinetobacter baumannii

    Energy Technology Data Exchange (ETDEWEB)

    Shah, Bhumika S., E-mail: bhumika.shah@mq.edu.au; Tetu, Sasha G. [Macquarie University, Research Park Drive, Sydney, NSW 2109 (Australia); Harrop, Stephen J. [University of New South Wales, Sydney, NSW 2052 (Australia); Paulsen, Ian T.; Mabbutt, Bridget C. [Macquarie University, Research Park Drive, Sydney, NSW 2109 (Australia)

    2014-09-25

    The structure of a short-chain dehydrogenase encoded within genomic islands of A. baumannii strains has been solved to 2.4 Å resolution. This classical SDR incorporates a flexible helical subdomain. The NADP-binding site and catalytic side chains are identified. Over 15% of the genome of an Australian clinical isolate of Acinetobacter baumannii occurs within genomic islands. An uncharacterized protein encoded within one island feature common to this and other International Clone II strains has been studied by X-ray crystallography. The 2.4 Å resolution structure of SDR-WM99c reveals it to be a new member of the classical short-chain dehydrogenase/reductase (SDR) superfamily. The enzyme contains a nucleotide-binding domain and, like many other SDRs, is tetrameric in form. The active site contains a catalytic tetrad (Asn117, Ser146, Tyr159 and Lys163) and water molecules occupying the presumed NADP cofactor-binding pocket. An adjacent cleft is capped by a relatively mobile helical subdomain, which is well positioned to control substrate access.

  10. Structure of a short-chain dehydrogenase/reductase (SDR) within a genomic island from a clinical strain of Acinetobacter baumannii

    International Nuclear Information System (INIS)

    Shah, Bhumika S.; Tetu, Sasha G.; Harrop, Stephen J.; Paulsen, Ian T.; Mabbutt, Bridget C.

    2014-01-01

    The structure of a short-chain dehydrogenase encoded within genomic islands of A. baumannii strains has been solved to 2.4 Å resolution. This classical SDR incorporates a flexible helical subdomain. The NADP-binding site and catalytic side chains are identified. Over 15% of the genome of an Australian clinical isolate of Acinetobacter baumannii occurs within genomic islands. An uncharacterized protein encoded within one island feature common to this and other International Clone II strains has been studied by X-ray crystallography. The 2.4 Å resolution structure of SDR-WM99c reveals it to be a new member of the classical short-chain dehydrogenase/reductase (SDR) superfamily. The enzyme contains a nucleotide-binding domain and, like many other SDRs, is tetrameric in form. The active site contains a catalytic tetrad (Asn117, Ser146, Tyr159 and Lys163) and water molecules occupying the presumed NADP cofactor-binding pocket. An adjacent cleft is capped by a relatively mobile helical subdomain, which is well positioned to control substrate access

  11. Identification and Characterization of Pathogen-Response Genes (repat) in Spodoptera frugiperda (Lepidoptera: Noctuidae).

    Science.gov (United States)

    Machado, Vilmar; Serrano, Jose; Galián, Jose

    2016-01-01

    The fall armyworm (Spodoptera frugiperda, Noctuidae, Lepidoptera) is one of the most important crop pests in the Americas, causing significant damage to maize, rice and sorghum. The mechanisms that determine its defences against pathogens are particularly relevant for the development of management and control strategies. We used an in silico approach to identify and characterize pathogen response genes (repat) present in different tissue libraries of S. fugiperda. The analyses revealed complete cDNA for nine repat genes; of these, repat15 and repat39 were found in libraries from a specific tissue--the midgut of larvae fed with xenobiotic substances. High expression levels of some genes were found in different libraries: 39 hits in repat30 in challenged hemocytes, 16 hits in repat31 in fat body, 10 hits in repat32 in fat body and 10 in challenged hemocytes, and 10 hits in repat38 in midgut of non-treated larvae and midgut of larvae fed with natural and xenobiotic substances. The genes corresponded to two ontology categories, stress response and immune response, and their phylogenetic relationships, nucleotide similarity, number of amino acid residues and molecular weights agree with what has been described for repat genes. It is noteworthy that proteins encoded by the repat genes of S. frugiperda have important defence functions in other tissues beyond midgut and that their functional categories are likely diverse, as they are related to cell envelope structure, energy metabolism, transport and binding.

  12. A super-family of transcriptional activators regulates bacteriophage packaging and lysis in Gram-positive bacteria

    Science.gov (United States)

    Quiles-Puchalt, Nuria; Tormo-Más, María Ángeles; Campoy, Susana; Toledo-Arana, Alejandro; Monedero, Vicente; Lasa, Íñigo; Novick, Richard P.; Christie, Gail E.; Penadés, José R.

    2013-01-01

    The propagation of bacteriophages and other mobile genetic elements requires exploitation of the phage mechanisms involved in virion assembly and DNA packaging. Here, we identified and characterized four different families of phage-encoded proteins that function as activators required for transcription of the late operons (morphogenetic and lysis genes) in a large group of phages infecting Gram-positive bacteria. These regulators constitute a super-family of proteins, here named late transcriptional regulators (Ltr), which share common structural, biochemical and functional characteristics and are unique to this group of phages. They are all small basic proteins, encoded by genes present at the end of the early gene cluster in their respective phage genomes and expressed under cI repressor control. To control expression of the late operon, the Ltr proteins bind to a DNA repeat region situated upstream of the terS gene, activating its transcription. This involves the C-terminal part of the Ltr proteins, which control specificity for the DNA repeat region. Finally, we show that the Ltr proteins are the only phage-encoded proteins required for the activation of the packaging and lysis modules. In summary, we provide evidence that phage packaging and lysis is a conserved mechanism in Siphoviridae infecting a wide variety of Gram-positive bacteria. PMID:23771138

  13. Identification and isolation of stimulator of interferon genes (STING): an innate immune sensory and adaptor gene from camelids.

    Science.gov (United States)

    Premraj, A; Aleyas, A G; Nautiyal, B; Rasool, T J

    2013-10-01

    The mechanism by which type I interferon-mediated antiviral response is mounted by hosts against invading pathogen is an intriguing one. Of late, an endoplasmic reticulum transmembrane protein encoded by a gene called stimulator of interferon genes (STING) is implicated in the innate signalling pathways and has been identified and cloned in few mammalian species including human, mouse and pig. In this article, we report the identification of STING from three different species of a highly conserved family of mammals - the camelids. cDNAs encoding the STING of Old World camels - dromedary camel (Camelus dromedarius) and bactrian camel (Camelus bactrianus) and a New World camel - llama (Llama glama) were amplified using conserved primers and RACE. The complete STING cDNA of dromedary camel is 2171 bp long with a 706-bp 5' untranslated regions (UTR), an 1137-bp open reading frame (ORF) and a 328-bp 3' UTR. Sequence and phylogenetic analysis of the ORF of STING from these three camelids indicate high level of similarity among camelids and conservation of critical amino acid residues across different species. Quantitative real-time PCR analysis revealed high levels of STING mRNA expression in blood, spleen, lymph node and lung. The identification of camelid STING will help in better understanding of the role of this molecule in the innate immunity of the camelids and other mammals. © 2013 John Wiley & Sons Ltd.

  14. BRD4 Phosphorylation Regulates HPV E2-Mediated Viral Transcription, Origin Replication, and Cellular MMP-9 Expression

    Directory of Open Access Journals (Sweden)

    Shwu-Yuan Wu

    2016-08-01

    Full Text Available Post-translational modification can modulate protein conformation and alter binding partner recruitment within gene regulatory regions. Here, we report that bromodomain-containing protein 4 (BRD4, a transcription co-factor and chromatin regulator, uses a phosphorylation-induced switch mechanism to recruit E2 protein encoded by cancer-associated human papillomavirus (HPV to viral early gene and cellular matrix metalloproteinase-9 (MMP-9 promoters. Enhanced MMP-9 expression, induced upon keratinocyte differentiation, occurs via BRD4-dependent recruitment of active AP-1 and NF-κB to their target sequences. This is triggered by replacement of AP-1 family members JunB and JunD by c-Jun and by re-localization of NF-κB from the cytoplasm to the nucleus. In addition, BRD4 phosphorylation is critical for E2- and origin-dependent HPV DNA replication. A class of phospho-BRD4-targeting compounds, distinct from the BET bromodomain inhibitors, effectively blocks BRD4 phosphorylation-specific functions in transcription and factor recruitment.

  15. De novo 14q24.2q24.3 microdeletion including IFT43 is associated with intellectual disability, skeletal anomalies, cardiac anomalies, and myopia.

    Science.gov (United States)

    Stokman, Marijn F; Oud, Machteld M; van Binsbergen, Ellen; Slaats, Gisela G; Nicolaou, Nayia; Renkema, Kirsten Y; Nijman, Isaac J; Roepman, Ronald; Giles, Rachel H; Arts, Heleen H; Knoers, Nine V A M; van Haelst, Mieke M

    2016-06-01

    We report an 11-year-old girl with mild intellectual disability, skeletal anomalies, congenital heart defect, myopia, and facial dysmorphisms including an extra incisor, cup-shaped ears, and a preauricular skin tag. Array comparative genomic hybridization analysis identified a de novo 4.5-Mb microdeletion on chromosome 14q24.2q24.3. The deleted region and phenotype partially overlap with previously reported patients. Here, we provide an overview of the literature on 14q24 microdeletions and further delineate the associated phenotype. We performed exome sequencing to examine other causes for the phenotype and queried genes present in the 14q24.2q24.3 microdeletion that are associated with recessive disease for variants in the non-deleted allele. The deleted region contains 65 protein-coding genes, including the ciliary gene IFT43. Although Sanger and exome sequencing did not identify variants in the second IFT43 allele or in other IFT complex A-protein-encoding genes, immunocytochemistry showed increased accumulation of IFT-B proteins at the ciliary tip in patient-derived fibroblasts compared to control cells, demonstrating defective retrograde ciliary transport. This could suggest a ciliary defect in the pathogenesis of this disorder. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  16. Bacteria of the human gut microbiome catabolize red seaweed glycans with carbohydrate-active enzyme updates from extrinsic microbes.

    Science.gov (United States)

    Hehemann, Jan-Hendrik; Kelly, Amelia G; Pudlo, Nicholas A; Martens, Eric C; Boraston, Alisdair B

    2012-11-27

    Humans host an intestinal population of microbes--collectively referred to as the gut microbiome--which encode the carbohydrate active enzymes, or CAZymes, that are absent from the human genome. These CAZymes help to extract energy from recalcitrant polysaccharides. The question then arises as to if and how the microbiome adapts to new carbohydrate sources when modern humans change eating habits. Recent metagenome analysis of microbiomes from healthy American, Japanese, and Spanish populations identified putative CAZymes obtained by horizontal gene transfer from marine bacteria, which suggested that human gut bacteria evolved to degrade algal carbohydrates-for example, consumed in form of sushi. We approached this hypothesis by studying such a polysaccharide utilization locus (PUL) obtained by horizontal gene transfer by the gut bacterium Bacteroides plebeius. Transcriptomic and growth experiments revealed that the PUL responds to the polysaccharide porphyran from red algae, enabling growth on this carbohydrate but not related substrates like agarose and carrageenan. The X-ray crystallographic and biochemical analysis of two proteins encoded by this PUL, BACPLE_01689 and BACPLE_01693, showed that they are β-porphyranases belonging to glycoside hydrolase families 16 and 86, respectively. The product complex of the GH86 at 1.3 Å resolution highlights the molecular details of porphyran hydrolysis by this new porphyranase. Combined, these data establish experimental support for the argument that CAZymes and associated genes obtained from extrinsic microbes add new catabolic functions to the human gut microbiome.

  17. Battle Against Cancer: An Everlasting Saga of p53

    Directory of Open Access Journals (Sweden)

    Qian Hao

    2014-12-01

    Full Text Available Cancer is one of the most life-threatening diseases characterized by uncontrolled growth and spread of malignant cells. The tumor suppressor p53 is the master regulator of tumor cell growth and proliferation. In response to various stress signals, p53 can be activated and transcriptionally induces a myriad of target genes, including both protein-encoding and non-coding genes, controlling cell cycle progression, DNA repair, senescence, apoptosis, autophagy and metabolism of tumor cells. However, around 50% of human cancers harbor mutant p53 and, in the majority of the remaining cancers, p53 is inactivated through multiple mechanisms. Herein, we review the recent progress in understanding the molecular basis of p53 signaling, particularly the newly identified ribosomal stress—p53 pathway, and the development of chemotherapeutics via activating wild-type p53 or restoring mutant p53 functions in cancer. A full understanding of p53 regulation will aid the development of effective cancer treatments.

  18. Deinococcus gobiensis cold shock protein improves salt stress tolerance of escherichia coli

    International Nuclear Information System (INIS)

    Jiang Shijie; Wang Jin; Yang Mingkun; Chen Ming; Zhang Wei; Luo Xuegang

    2013-01-01

    The Deinococcus gobiensis I-0, an extremely radiation-resistant bacterium, isolated from the Gobi, has superior resistance to abiotic stress (e.g radiation, oxidation, dehydration and so on). The two cold-shock proteins encoded by csp1 (Dgo_CA1136) and csp2 (Dgo_PA0041) were identified in the complete genome sequence of D. gobiensis. In this study, we showed that D. gobiensis Csp1 protected Escherichia coli cells against cold shock and other abiotic stresses such as salt and osmotic shocks. The quantitative real-time PCR assay shows that the expression of trehalose synthase (otsA, otsB) was up-regulated remarkably under salt stress in the csp1-expressing strain, while no difference in the expression of the genes involved in trehalose degradation (treB and treC). The results suggested that Csp1 caused the accumulation of the trehalose was a major feature for improving tolerance to salt stress in E. coli. (authors)

  19. Robust heat-inducible gene expression by two endogenous hsp70-derived promoters in transgenic Aedes aegypti

    Science.gov (United States)

    Carpenetti, Tiffany L. G.; Aryan, Azadeh; Myles, Kevin M.; Adelman, Zach N.

    2011-01-01

    Aedes aegypti is an important vector of the viruses that cause dengue fever, dengue hemorrhagic fever, and yellow fever. Reverse genetic approaches to the study of gene function in this mosquito have been limited by the lack of a robust inducible promoter to allow precise temporal control over a protein-encoding or hairpin RNA transgene. Likewise, investigations into the molecular and biochemical basis of vector competence would benefit from the ability to activate an anti-pathogen molecule at specific times during infection. We have characterized the ability of genomic sequences derived from two Ae. aegypti hsp70 genes to drive heat-inducible expression of a reporter in both transient and germline transformation contexts. AaHsp70-luciferase transcripts accumulated specifically after heat shock, and displayed a pattern of rapid induction and decay similar to endogenous AaHsp70 genes. Luciferase expression in transgenic Ae. aegypti increased by ∼25-50 fold in whole adults by four hours after heat-shock, with significant activity (∼20 fold) remaining at 24 hr. Heat-induced expression was even more dramatic in midgut tissues, with one strain showing a ∼2500-fold increase in luciferase activity. The AaHsp70 promoters described could be valuable for gene function studies as well as for the precise timing of the expression of anti-pathogen molecules. PMID:22142225

  20. New perspectives on host-parasite interplay by comparative transcriptomic and proteomic analyses of Schistosoma japonicum.

    Directory of Open Access Journals (Sweden)

    Feng Liu

    2006-04-01

    Full Text Available Schistosomiasis remains a serious public health problem with an estimated 200 million people infected in 76 countries. Here we isolated ~ 8,400 potential protein-encoding cDNA contigs from Schistosoma japonicum after sequencing circa 84,000 expressed sequence tags. In tandem, we undertook a high-throughput proteomics approach to characterize the protein expression profiles of a number of developmental stages (cercariae, hepatic schistosomula, female and male adults, eggs, and miracidia and tissues at the host-parasite interface (eggshell and tegument by interrogating the protein database deduced from the contigs. Comparative analysis of these transcriptomic and proteomic data, the latter including 3,260 proteins with putative identities, revealed differential expression of genes among the various developmental stages and sexes of S. japonicum and localization of putative secretory and membrane antigens, enzymes, and other gene products on the adult tegument and eggshell, many of which displayed genetic polymorphisms. Numerous S. japonicum genes exhibited high levels of identity with those of their mammalian hosts, whereas many others appeared to be conserved only across the genus Schistosoma or Phylum Platyhelminthes. These findings are expected to provide new insights into the pathophysiology of schistosomiasis and for the development of improved interventions for disease control and will facilitate a more fundamental understanding of schistosome biology, evolution, and the host-parasite interplay.

  1. Three regulators of G protein signaling differentially affect mating, morphology and virulence in the smut fungus Ustilago maydis.

    Science.gov (United States)

    Moretti, Marino; Wang, Lei; Grognet, Pierre; Lanver, Daniel; Link, Hannes; Kahmann, Regine

    2017-09-01

    Regulators of G protein signaling (RGS) proteins modulate heterotrimeric G protein signaling negatively. To broaden an understanding of the roles of RGS proteins in fungal pathogens, we functionally characterized the three RGS protein-encoding genes (rgs1, rgs2 and rgs3) in the phytopathogenic fungus Ustilago maydis. It was found that RGS proteins played distinct roles in the regulation of development and virulence. rgs1 had a minor role in virulence when deleted in a solopathogenic strain. In crosses, rgs1 was dispensable for mating and filamentation, but was required for teliospore production. Haploid rgs2 mutants were affected in cell morphology, growth, mating and were unable to cause disease symptoms in crosses. However, virulence was unaffected when rgs2 was deleted in a solopathogenic strain, suggesting an exclusive involvement in pre-fusion events. These rgs2 phenotypes are likely connected to elevated intracellular cAMP levels. rgs3 mutants were severely attenuated in mating, in their response to pheromone, virulence and formation of mature teliospores. The mating defect could be traced back to reduced expression of the transcription factor rop1. It was speculated that the distinct roles of the three U. maydis RGS proteins were achieved by direct modulation of the Gα subunit-activated signaling pathways as well as through Gα-independent functions. © 2017 John Wiley & Sons Ltd.

  2. A copper-induced quinone degradation pathway provides protection against combined copper/quinone stress in Lactococcus lactis IL1403.

    Science.gov (United States)

    Mancini, Stefano; Abicht, Helge K; Gonskikh, Yulia; Solioz, Marc

    2015-02-01

    Quinones are ubiquitous in the environment. They occur naturally but are also in widespread use in human and industrial activities. Quinones alone are relatively benign to bacteria, but in combination with copper, they become toxic by a mechanism that leads to intracellular thiol depletion. Here, it was shown that the yahCD-yaiAB operon of Lactococcus lactis IL1403 provides resistance to combined copper/quinone stress. The operon is under the control of CopR, which also regulates expression of the copRZA copper resistance operon as well as other L. lactis genes. Expression of the yahCD-yaiAB operon is induced by copper but not by quinones. Two of the proteins encoded by the operon appear to play key roles in alleviating quinone/copper stress: YaiB is a flavoprotein that converts p-benzoquinones to less toxic hydroquinones, using reduced nicotinamide adenine dinucleotide phosphate (NADPH) as reductant; YaiA is a hydroquinone dioxygenase that converts hydroquinone putatively to 4-hydroxymuconic semialdehyde in an oxygen-consuming reaction. Hydroquinone and methylhydroquinone are both substrates of YaiA. Deletion of yaiB causes increased sensitivity of L. lactis to quinones and complete growth arrest under combined quinone and copper stress. Copper induction of the yahCD-yaiAB operon offers protection to copper/quinone toxicity and could provide a growth advantage to L. lactis in some environments. © 2014 John Wiley & Sons Ltd.

  3. Synucleins: are they two-edged swords?

    Science.gov (United States)

    Surguchov, Andrei

    2013-02-01

    The synuclein family consists of three distinct highly homologous genes, α-synuclein, β-synuclein, and γ-synuclein, which have so far been found only in vertebrates. Proteins encoded by these genes are characterized by an acidic C-terminal region and five or six imperfect repeat motifs (KTKEGV) distributed throughout the highly conserved N-terminal region. Numerous data demonstrate that synucleins are implicated in two groups of the most devastating human disorders, i.e., neurodegenerative diseases (NDDs) and cancer. Mutations in the α-synuclein gene are associated with familial forms of Parkinson's disease (PD), and accumulation of α-synuclein inclusions is a hallmark of this disorder. In breast cancer, increased expression of γ-synuclein correlates with disease progression. Conversely, some results indicate that the members of the synuclein family may have a protective effect. How might these small proteins combine such controversial properties? We present evidence that synuclein's features are basically regulated by two mechanisms, i.e., posttranslational modifications (PTMs) and the level of their expression. We also discuss a new, emerging area of investigation of synucleins, namely, their role in the cell-to-cell propagation of pathology. Copyright © 2012 Wiley Periodicals, Inc.

  4. Independent feeding and metabolic actions of orexins in mice.

    Science.gov (United States)

    Lubkin, M; Stricker-Krongrad, A

    1998-12-18

    Orexin-A and orexin-B (OX peptides) are two putative products of a newly discovered secreted protein encoded by a mRNA restricted to neuronal cell bodies of the lateral hypothalamus (LH). Because the activation of the LH can induce changes in energy balance, we wanted to investigate the actions of OX peptides on energy metabolism in mice. We injected male C57BL/6J mice with different doses (1, 3, and 10 nmol) of orexin-A and orexin-B into the third ventricle (i3vt). A single i3vt injection of orexin-A 3 h into the light period slightly stimulated feeding at the lowest dose only over the following 4 h (11 +/- 09 mg/mouse vs 80 +/- 13 mg/mouse, p energy utilization using indirect calorimetry. Single i3vt injection 3 h after light on, or just before dark onset, or in 4-h fasted mice resulted in increases in the metabolic rate. These effects were associated with decreases or increases in the respiratory quotient regarding the time of injection or the underlying metabolic state of the mice. The present findings provide direct evidence that OX peptides are more likely to be involved in the control of energy metabolism than of food intake in mice. Copyright 1998 Academic Press.

  5. Elevated Fibroblast Growth Factor Signaling Is Critical for the Pathogenesis of the Dwarfism in Evc2/Limbin Mutant Mice.

    Science.gov (United States)

    Zhang, Honghao; Kamiya, Nobuhiro; Tsuji, Takehito; Takeda, Haruko; Scott, Greg; Rajderkar, Sudha; Ray, Manas K; Mochida, Yoshiyuki; Allen, Benjamin; Lefebvre, Veronique; Hung, Irene H; Ornitz, David M; Kunieda, Tetsuo; Mishina, Yuji

    2016-12-01

    Ellis-van Creveld (EvC) syndrome is a skeletal dysplasia, characterized by short limbs, postaxial polydactyly, and dental abnormalities. EvC syndrome is also categorized as a ciliopathy because of ciliary localization of proteins encoded by the two causative genes, EVC and EVC2 (aka LIMBIN). While recent studies demonstrated important roles for EVC/EVC2 in Hedgehog signaling, there is still little known about the pathophysiological mechanisms underlying the skeletal dysplasia features of EvC patients, and in particular why limb development is affected, but not other aspects of organogenesis that also require Hedgehog signaling. In this report, we comprehensively analyze limb skeletogenesis in Evc2 mutant mice and in cell and tissue cultures derived from these mice. Both in vivo and in vitro data demonstrate elevated Fibroblast Growth Factor (FGF) signaling in Evc2 mutant growth plates, in addition to compromised but not abrogated Hedgehog-PTHrP feedback loop. Elevation of FGF signaling, mainly due to increased Fgf18 expression upon inactivation of Evc2 in the perichondrium, critically contributes to the pathogenesis of limb dwarfism. The limb dwarfism phenotype is partially rescued by inactivation of one allele of Fgf18 in the Evc2 mutant mice. Taken together, our data uncover a novel pathogenic mechanism to understand limb dwarfism in patients with Ellis-van Creveld syndrome.

  6. Elevated Fibroblast Growth Factor Signaling Is Critical for the Pathogenesis of the Dwarfism in Evc2/Limbin Mutant Mice.

    Directory of Open Access Journals (Sweden)

    Honghao Zhang

    2016-12-01

    Full Text Available Ellis-van Creveld (EvC syndrome is a skeletal dysplasia, characterized by short limbs, postaxial polydactyly, and dental abnormalities. EvC syndrome is also categorized as a ciliopathy because of ciliary localization of proteins encoded by the two causative genes, EVC and EVC2 (aka LIMBIN. While recent studies demonstrated important roles for EVC/EVC2 in Hedgehog signaling, there is still little known about the pathophysiological mechanisms underlying the skeletal dysplasia features of EvC patients, and in particular why limb development is affected, but not other aspects of organogenesis that also require Hedgehog signaling. In this report, we comprehensively analyze limb skeletogenesis in Evc2 mutant mice and in cell and tissue cultures derived from these mice. Both in vivo and in vitro data demonstrate elevated Fibroblast Growth Factor (FGF signaling in Evc2 mutant growth plates, in addition to compromised but not abrogated Hedgehog-PTHrP feedback loop. Elevation of FGF signaling, mainly due to increased Fgf18 expression upon inactivation of Evc2 in the perichondrium, critically contributes to the pathogenesis of limb dwarfism. The limb dwarfism phenotype is partially rescued by inactivation of one allele of Fgf18 in the Evc2 mutant mice. Taken together, our data uncover a novel pathogenic mechanism to understand limb dwarfism in patients with Ellis-van Creveld syndrome.

  7. Targeted disruption in mice of a neural stem cell-maintaining, KRAB-Zn finger-encoding gene that has rapidly evolved in the human lineage.

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    Huan-Chieh Chien

    Full Text Available Understanding the genetic basis of the physical and behavioral traits that separate humans from other primates is a challenging but intriguing topic. The adaptive functions of the expansion and/or reduction in human brain size have long been explored. From a brain transcriptome project we have identified a KRAB-Zn finger protein-encoding gene (M003-A06 that has rapidly evolved since the human-chimpanzee separation. Quantitative RT-PCR analysis of different human tissues indicates that M003-A06 expression is enriched in the human fetal brain in addition to the fetal heart. Furthermore, analysis with use of immunofluorescence staining, neurosphere culturing and Western blotting indicates that the mouse ortholog of M003-A06, Zfp568, is expressed mainly in the embryonic stem (ES cells and fetal as well as adult neural stem cells (NSCs. Conditional gene knockout experiments in mice demonstrates that Zfp568 is both an NSC maintaining- and a brain size-regulating gene. Significantly, molecular genetic analyses show that human M003-A06 consists of 2 equilibrated allelic types, H and C, one of which (H is human-specific. Combined contemporary genotyping and database mining have revealed interesting genetic associations between the different genotypes of M003-A06 and the human head sizes. We propose that M003-A06 is likely one of the genes contributing to the uniqueness of the human brain in comparison to other higher primates.

  8. DCD – a novel plant specific domain in proteins involved in development and programmed cell death

    Directory of Open Access Journals (Sweden)

    Doerks Tobias

    2005-07-01

    Full Text Available Abstract Background Recognition of microbial pathogens by plants triggers the hypersensitive reaction, a common form of programmed cell death in plants. These dying cells generate signals that activate the plant immune system and alarm the neighboring cells as well as the whole plant to activate defense responses to limit the spread of the pathogen. The molecular mechanisms behind the hypersensitive reaction are largely unknown except for the recognition process of pathogens. We delineate the NRP-gene in soybean, which is specifically induced during this programmed cell death and contains a novel protein domain, which is commonly found in different plant proteins. Results The sequence analysis of the protein, encoded by the NRP-gene from soybean, led to the identification of a novel domain, which we named DCD, because it is found in plant proteins involved in development and cell death. The domain is shared by several proteins in the Arabidopsis and the rice genomes, which otherwise show a different protein architecture. Biological studies indicate a role of these proteins in phytohormone response, embryo development and programmed cell by pathogens or ozone. Conclusion It is tempting to speculate, that the DCD domain mediates signaling in plant development and programmed cell death and could thus be used to identify interacting proteins to gain further molecular insights into these processes.

  9. The Replacement of five Consecutive Amino Acids in the Cyt1A Protein of Bacillus thuringiensis Enhances its Cytotoxic Activity against Lung Epithelial Cancer Cells

    Directory of Open Access Journals (Sweden)

    Kavita Nair

    2018-03-01

    Full Text Available Cyt1A protein is a cytolytic protein encoded by the cyt gene of Bacillus thuringiensis subsp. israelensis (Bti as part of the parasporal crystal proteins produced during the sporulation. Cyt1A protein is unique compared to the other endotoxins present in these parasporal crystals. Unlike δ-endotoxins, Cyt1A protein does not require receptors to bind to the target cell and activate the toxicity. It has the ability to affect a broad range of cell types and organisms, due to this characteristic. Cyt1A has been recognized to not only target the insect cells directly, but also recruit other endotoxins by acting as receptors. Due to these mode of actions, Cyt1A has been studied for its cytolytic activity against human cancer cell lines, although not extensively. In this study, we report a novel Cyt1A protein produced by a Bti strain QBT229 isolated from Qatar. When tested for its cytotoxicity against lung cancer cells, this local strain showed considerably higher activity compared to that of the reference Bti and other strains tested. The possible reasons for such enhanced activity were explored at the gene and protein levels. It was evidenced that five consecutive amino acid replacements in the β8 sheet of the Cyt1A protein enhanced the cytotoxicity against the lung epithelial cancer cells. Such novel Cyt1A protein with high cytotoxicity against lung cancer cells has been characterized and reported through this study.

  10. ToxR regulates the production of lipoproteins and the expression of serum resistance in Vibrio cholerae

    International Nuclear Information System (INIS)

    Parsot, C.; Taxman, E.; Mekalanos, J.J.

    1991-01-01

    The genes encoding three lipoproteins of Vibrio cholerae were identified by a combination of DNA sequence analysis and [ 3 H]palmitate labeling of hybrid proteins encoded by TnphoA gene fusions. The expression of these three lipoproteins, TagA, AcfD, and TcpC, was controlled by ToxR, the cholera toxin transcriptional activator. The involvement of other bacterial lipoproteins in conferring resistance ot the bactericidal effects of complement prompted us to examine this possibility in V. cholerae. Remarkably, mutations in toxR and tcp genes (including tcpC), involved in the biogenesis of the toxin coregulated pili, rendered V. cholerae about 10 4 - 10 6 times more sensitive to the vibriocidal activity of antibody and complement. Since V. cholerae is a noninvasive organism and toxR and tcp mutants are highly defective in intestinal colonization in animals and humans, these results raise the possibility that resistance to a gut-associated, complement-like bactericidal activity may be a major virulence determinant of V. cholerae and other enterobacterial species

  11. Diversity of extracellular proteins during the transition from the ‘proto-apicomplexan’ alveolates to the apicomplexan obligate parasites

    KAUST Repository

    Templeton, Thomas J.; Pain, Arnab

    2015-01-01

    The recent completion of high-coverage draft genome sequences for several alveolate protozoans – namely, the chromerids, Chromera velia and Vitrella brassicaformis ; the perkinsid Perkinsus marinus ; the apicomplexan, Gregarina niphandrodes , as well as high coverage transcriptome sequence information for several colpodellids, allows for new genome-scale comparisons across a rich landscape of apicomplexans and other alveolates. Genome annotations can now be used to help interpret fine ultrastructure and cell biology, and guide new studies to describe a variety of alveolate life strategies, such as symbiosis or free living, predation, and obligate intracellular parasitism, as well to provide foundations to dissect the evolutionary transitions between these niches. This review focuses on the attempt to identify extracellular proteins which might mediate the physical interface of cell–cell interactions within the above life strategies, aided by annotation of the repertoires of predicted surface and secreted proteins encoded within alveolate genomes. In particular, we discuss what descriptions of the predicted extracellular proteomes reveal regarding a hypothetical last common ancestor of a pre-apicomplexan alveolate – guided by ultrastructure, life strategies and phylogenetic relationships – in an attempt to understand the evolution of obligate parasitism in apicomplexans.

  12. Sequence analysis of the genome of carnation (Dianthus caryophyllus L.).

    Science.gov (United States)

    Yagi, Masafumi; Kosugi, Shunichi; Hirakawa, Hideki; Ohmiya, Akemi; Tanase, Koji; Harada, Taro; Kishimoto, Kyutaro; Nakayama, Masayoshi; Ichimura, Kazuo; Onozaki, Takashi; Yamaguchi, Hiroyasu; Sasaki, Nobuhiro; Miyahara, Taira; Nishizaki, Yuzo; Ozeki, Yoshihiro; Nakamura, Noriko; Suzuki, Takamasa; Tanaka, Yoshikazu; Sato, Shusei; Shirasawa, Kenta; Isobe, Sachiko; Miyamura, Yoshinori; Watanabe, Akiko; Nakayama, Shinobu; Kishida, Yoshie; Kohara, Mitsuyo; Tabata, Satoshi

    2014-06-01

    The whole-genome sequence of carnation (Dianthus caryophyllus L.) cv. 'Francesco' was determined using a combination of different new-generation multiplex sequencing platforms. The total length of the non-redundant sequences was 568,887,315 bp, consisting of 45,088 scaffolds, which covered 91% of the 622 Mb carnation genome estimated by k-mer analysis. The N50 values of contigs and scaffolds were 16,644 bp and 60,737 bp, respectively, and the longest scaffold was 1,287,144 bp. The average GC content of the contig sequences was 36%. A total of 1050, 13, 92 and 143 genes for tRNAs, rRNAs, snoRNA and miRNA, respectively, were identified in the assembled genomic sequences. For protein-encoding genes, 43 266 complete and partial gene structures excluding those in transposable elements were deduced. Gene coverage was ∼ 98%, as deduced from the coverage of the core eukaryotic genes. Intensive characterization of the assigned carnation genes and comparison with those of other plant species revealed characteristic features of the carnation genome. The results of this study will serve as a valuable resource for fundamental and applied research of carnation, especially for breeding new carnation varieties. Further information on the genomic sequences is available at http://carnation.kazusa.or.jp. © The Author 2013. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

  13. Quality control methodology for high-throughput protein-protein interaction screening.

    Science.gov (United States)

    Vazquez, Alexei; Rual, Jean-François; Venkatesan, Kavitha

    2011-01-01

    Protein-protein interactions are key to many aspects of the cell, including its cytoskeletal structure, the signaling processes in which it is involved, or its metabolism. Failure to form protein complexes or signaling cascades may sometimes translate into pathologic conditions such as cancer or neurodegenerative diseases. The set of all protein interactions between the proteins encoded by an organism constitutes its protein interaction network, representing a scaffold for biological function. Knowing the protein interaction network of an organism, combined with other sources of biological information, can unravel fundamental biological circuits and may help better understand the molecular basics of human diseases. The protein interaction network of an organism can be mapped by combining data obtained from both low-throughput screens, i.e., "one gene at a time" experiments and high-throughput screens, i.e., screens designed to interrogate large sets of proteins at once. In either case, quality controls are required to deal with the inherent imperfect nature of experimental assays. In this chapter, we discuss experimental and statistical methodologies to quantify error rates in high-throughput protein-protein interactions screens.

  14. Evasion of antiviral innate immunity by Theiler's virus L* protein through direct inhibition of RNase L.

    Directory of Open Access Journals (Sweden)

    Frédéric Sorgeloos

    Full Text Available Theiler's virus is a neurotropic picornavirus responsible for chronic infections of the central nervous system. The establishment of a persistent infection and the subsequent demyelinating disease triggered by the virus depend on the expression of L*, a viral accessory protein encoded by an alternative open reading frame of the virus. We discovered that L* potently inhibits the interferon-inducible OAS/RNase L pathway. The antagonism of RNase L by L* was particularly prominent in macrophages where baseline oligoadenylate synthetase (OAS and RNase L expression levels are elevated, but was detectable in fibroblasts after IFN pretreatment. L* mutations significantly affected Theiler's virus replication in primary macrophages derived from wild-type but not from RNase L-deficient mice. L* counteracted the OAS/RNase L pathway through direct interaction with the ankyrin domain of RNase L, resulting in the inhibition of this enzyme. Interestingly, RNase L inhibition was species-specific as Theiler's virus L* protein blocked murine RNase L but not human RNase L or RNase L of other mammals or birds. Direct RNase L inhibition by L* and species specificity were confirmed in an in vitro assay performed with purified proteins. These results demonstrate a novel viral mechanism to elude the antiviral OAS/RNase L pathway. By targeting the effector enzyme of this antiviral pathway, L* potently inhibits RNase L, underscoring the importance of this enzyme in innate immunity against Theiler's virus.

  15. Purification, crystallization and preliminary X-ray crystallographic analysis of a methanol dehydrogenase from the marine bacterium Methylophaga aminisulfidivorans MPT

    International Nuclear Information System (INIS)

    Choi, Jin Myung; Kim, Hee Gon; Kim, Jeong-Sun; Youn, Hyung-Seop; Eom, Soo Hyun; Yu, Sung-Lim; Kim, Si Wouk; Lee, Sung Haeng

    2011-01-01

    In order to obtain molecular insights into the methanol-oxidizing system of M. aminisulfidivorans, a native heterotetrameric α 2 β 2 methanol dehydrogenase complex was directly purified from M. aminisulfidivorans MP T grown in the presence of methanol and crystallized. Methylophaga aminisulfidivorans MP T is a marine methylotrophic bacterium that utilizes C 1 compounds such as methanol as a carbon and energy source. The released electron from oxidation flows through a methanol-oxidizing system (MOX) consisting of a series of electron-transfer proteins encoded by the mox operon. One of the key enzymes in the pathway is methanol dehydrogenase (MDH), which contains the prosthetic group pyrroloquinoline quinone (PQQ) and converts methanol to formaldehyde in the periplasm by transferring two electrons from the oxidation of one methanol molecule to the electron acceptor cytochrome c L . In order to obtain molecular insights into the oxidation mechanism, a native heterotetrameric α 2 β 2 MDH complex was directly purified from M. aminisulfidivorans MP T grown in the presence of methanol and crystallized. The crystal diffracted to 1.7 Å resolution and belonged to the monoclinic space group P2 1 (unit-cell parameters a = 63.9, b = 109.5, c = 95.6 Å, β = 100.5°). The asymmetric unit of the crystal contained one heterotetrameric complex, with a calculated Matthews coefficient of 2.24 Å 3 Da −1 and a solvent content of 45.0%

  16. Discovery and genomic characterization of a novel ovine partetravirus and a new genotype of bovine partetravirus.

    Directory of Open Access Journals (Sweden)

    Herman Tse

    Full Text Available Partetravirus is a recently described group of animal parvoviruses which include the human partetravirus, bovine partetravirus and porcine partetravirus (previously known as human parvovirus 4, bovine hokovirus and porcine hokovirus respectively. In this report, we describe the discovery and genomic characterization of partetraviruses in bovine and ovine samples from China. These partetraviruses were detected by PCR in 1.8% of bovine liver samples, 66.7% of ovine liver samples and 71.4% of ovine spleen samples. One of the bovine partetraviruses detected in the present samples is phylogenetically distinct from previously reported bovine partetraviruses and likely represents a novel genotype. The ovine partetravirus is a novel partetravirus and phylogenetically most related to the bovine partetraviruses. The genome organization is conserved amongst these viruses, including the presence of a putative transmembrane protein encoded by an overlapping reading frame in ORF2. Results from the present study provide further support to the classification of partetraviruses as a separate genus in Parvovirinae.

  17. Genome comparison of barley and maize smut fungi reveals targeted loss of RNA silencing components and species-specific presence of transposable elements.

    Science.gov (United States)

    Laurie, John D; Ali, Shawkat; Linning, Rob; Mannhaupt, Gertrud; Wong, Philip; Güldener, Ulrich; Münsterkötter, Martin; Moore, Richard; Kahmann, Regine; Bakkeren, Guus; Schirawski, Jan

    2012-05-01

    Ustilago hordei is a biotrophic parasite of barley (Hordeum vulgare). After seedling infection, the fungus persists in the plant until head emergence when fungal spores develop and are released from sori formed at kernel positions. The 26.1-Mb U. hordei genome contains 7113 protein encoding genes with high synteny to the smaller genomes of the related, maize-infecting smut fungi Ustilago maydis and Sporisorium reilianum but has a larger repeat content that affected genome evolution at important loci, including mating-type and effector loci. The U. hordei genome encodes components involved in RNA interference and heterochromatin formation, normally involved in genome defense, that are lacking in the U. maydis genome due to clean excision events. These excision events were possibly a result of former presence of repetitive DNA and of an efficient homologous recombination system in U. maydis. We found evidence of repeat-induced point mutations in the genome of U. hordei, indicating that smut fungi use different strategies to counteract the deleterious effects of repetitive DNA. The complement of U. hordei effector genes is comparable to the other two smuts but reveals differences in family expansion and clustering. The availability of the genome sequence will facilitate the identification of genes responsible for virulence and evolution of smut fungi on their respective hosts.

  18. Genome Comparison of Barley and Maize Smut Fungi Reveals Targeted Loss of RNA Silencing Components and Species-Specific Presence of Transposable Elements[W

    Science.gov (United States)

    Laurie, John D.; Ali, Shawkat; Linning, Rob; Mannhaupt, Gertrud; Wong, Philip; Güldener, Ulrich; Münsterkötter, Martin; Moore, Richard; Kahmann, Regine; Bakkeren, Guus; Schirawski, Jan

    2012-01-01

    Ustilago hordei is a biotrophic parasite of barley (Hordeum vulgare). After seedling infection, the fungus persists in the plant until head emergence when fungal spores develop and are released from sori formed at kernel positions. The 26.1-Mb U. hordei genome contains 7113 protein encoding genes with high synteny to the smaller genomes of the related, maize-infecting smut fungi Ustilago maydis and Sporisorium reilianum but has a larger repeat content that affected genome evolution at important loci, including mating-type and effector loci. The U. hordei genome encodes components involved in RNA interference and heterochromatin formation, normally involved in genome defense, that are lacking in the U. maydis genome due to clean excision events. These excision events were possibly a result of former presence of repetitive DNA and of an efficient homologous recombination system in U. maydis. We found evidence of repeat-induced point mutations in the genome of U. hordei, indicating that smut fungi use different strategies to counteract the deleterious effects of repetitive DNA. The complement of U. hordei effector genes is comparable to the other two smuts but reveals differences in family expansion and clustering. The availability of the genome sequence will facilitate the identification of genes responsible for virulence and evolution of smut fungi on their respective hosts. PMID:22623492

  19. Isolation and characterisation of cDNA clones representing the genes encoding the major tuber storage protein (dioscorin) of yam (Dioscorea cayenensis Lam.).

    Science.gov (United States)

    Conlan, R S; Griffiths, L A; Napier, J A; Shewry, P R; Mantell, S; Ainsworth, C

    1995-06-01

    cDNA clones encoding dioscorins, the major tuber storage proteins (M(r) 32,000) of yam (Dioscorea cayenesis) have been isolated. Two classes of clone (A and B, based on hybrid release translation product sizes and nucleotide sequence differences) which are 84.1% similar in their protein coding regions, were identified. The protein encoded by the open reading frame of the class A cDNA insert is of M(r) 30,015. The difference in observed and calculated molecular mass might be attributed to glycosylation. Nucleotide sequencing and in vitro transcription/translation suggest that the class A dioscorin proteins are synthesised with signal peptides of 18 amino acid residues which are cleaved from the mature peptide. The class A and class B proteins are 69.6% similar with respect to each other, but show no sequence identity with other plant proteins or with the major tuber storage proteins of potato (patatin) or sweet potato (sporamin). Storage protein gene expression was restricted to developing tubers and was not induced by growth conditions known to induce expression of tuber storage protein genes in other plant species. The codon usage of the dioscorin genes suggests that the Dioscoreaceae are more closely related to dicotyledonous than to monocotyledonous plants.

  20. Characteristics of the Lotus japonicus gene repertoire deduced from large-scale expressed sequence tag (EST) analysis.

    Science.gov (United States)

    Asamizu, Erika; Nakamura, Yasukazu; Sato, Shusei; Tabata, Satoshi

    2004-02-01

    To perform a comprehensive analysis of genes expressed in a model legume, Lotus japonicus, a total of 74472 3'-end expressed sequence tags (EST) were generated from cDNA libraries produced from six different organs. Clustering of sequences was performed with an identity criterion of 95% for 50 bases, and a total of 20457 non-redundant sequences, 8503 contigs and 11954 singletons were generated. EST sequence coverage was analyzed by using the annotated L. japonicus genomic sequence and 1093 of the 1889 predicted protein-encoding genes (57.9%) were hit by the EST sequence(s). Gene content was compared to several plant species. Among the 8503 contigs, 471 were identified as sequences conserved only in leguminous species and these included several disease resistance-related genes. This suggested that in legumes, these genes may have evolved specifically to resist pathogen attack. The rate of gene sequence divergence was assessed by comparing similarity level and functional category based on the Gene Ontology (GO) annotation of Arabidopsis genes. This revealed that genes encoding ribosomal proteins, as well as those related to translation, photosynthesis, and cellular structure were more abundantly represented in the highly conserved class, and that genes encoding transcription factors and receptor protein kinases were abundantly represented in the less conserved class. To make the sequence information and the cDNA clones available to the research community, a Web database with useful services was created at http://www.kazusa.or.jp/en/plant/lotus/EST/.

  1. Immunogenicity and protective effect of recombinant Brucella abortus Ndk (rNdk) against a virulent strain B. abortus 544 infection in BALB/c mice.

    Science.gov (United States)

    Hop, Huynh Tan; Simborio, Hannah Leah; Reyes, Alisha Wehdnesday Bernardo; Arayan, Lauren Togonon; Min, WonGi; Lee, Hu Jang; Kim, Dong Hee; Chang, Hong Hee; Kim, Suk

    2015-02-01

    In this study, we particularly evaluated the protective effect of recombinant protein encoded by Brucella abortus 544 ndk (nucleoside diphosphate kinase) gene against B. abortus infection in the BALB/c mice. Cloning and expression of B. abortus Ndk was accomplished by PCR amplification into a pMAL expression system, and purification of a recombinant Ndk (rNdk). As for the determination of IgG responses, rNdk induced vigorous IgG production, especially higher in IgG2a compared to IgG1 with titers of 5.2 and 4.8, respectively, whereas titers of these in mice immunized with MBP were 2.4 of IgG2a and 2.6 of IgG1. The analysis of cytokine has revealed that rNdk can strongly induce production of IFN-γ as well as proinflammatory cytokines (TNF, MCP1 and IL-6) but not much IL-10, suggesting rNdk elicited predominantly cell-mediated immune responses. Furthermore, the spleen proliferation and bacterial burden in the spleen of rNdk immunized mice were significantly lower than those of MBP-immunized mice against virulent B. abortus challenge (P abortus might be a useful candidate for subunit vaccine for brucellosis in animals. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  2. Transcript and proteomic analysis of developing white lupin (Lupinus albus L. roots

    Directory of Open Access Journals (Sweden)

    Watson Bonnie

    2009-01-01

    Full Text Available Abstract Background White lupin (Lupinus albus L. roots efficiently take up and accumulate (heavy metals, adapt to phosphate deficiency by forming cluster roots, and secrete antimicrobial prenylated isoflavones during development. Genomic and proteomic approaches were applied to identify candidate genes and proteins involved in antimicrobial defense and (heavy metal uptake and translocation. Results A cDNA library was constructed from roots of white lupin seedlings. Eight thousand clones were randomly sequenced and assembled into 2,455 unigenes, which were annotated based on homologous matches in the NCBInr protein database. A reference map of developing white lupin root proteins was established through 2-D gel electrophoresis and peptide mass fingerprinting. High quality peptide mass spectra were obtained for 170 proteins. Microsomal membrane proteins were separated by 1-D gel electrophoresis and identified by LC-MS/MS. A total of 74 proteins were putatively identified by the peptide mass fingerprinting and the LC-MS/MS methods. Genomic and proteomic analyses identified candidate genes and proteins encoding metal binding and/or transport proteins, transcription factors, ABC transporters and phenylpropanoid biosynthetic enzymes. Conclusion The combined EST and protein datasets will facilitate the understanding of white lupin's response to biotic and abiotic stresses and its utility for phytoremediation. The root ESTs provided 82 perfect simple sequence repeat (SSR markers with potential utility in breeding white lupin for enhanced agronomic traits.

  3. Viewing the human microbiome through three-dimensional glasses: integrating structural and functional studies to better define the properties of myriad carbohydrate-active enzymes

    International Nuclear Information System (INIS)

    Turnbaugh, Peter J.; Henrissat, Bernard; Gordon, Jeffrey I.

    2010-01-01

    Metagenomics has unleashed a deluge of sequencing data describing the organismal, genetic, and transcriptional diversity of the human microbiome. To better understand the precise functions of the myriad proteins encoded by the microbiome, including carbohydrate-active enzymes, it will be critical to combine structural studies with functional analyses. Recent studies have provided an unprecedented view of the trillions of microbes associated with the human body. The human microbiome harbors tremendous diversity at multiple levels: the species that colonize each individual and each body habitat; the genes that are found in each organism’s genome; the expression of these genes and the interactions and activities of their protein products. The sources of this diversity are wide-ranging and reflect both environmental and host factors. A major challenge moving forward is defining the precise functions of members of various families of proteins represented in our microbiomes, including the highly diverse carbohydrate-active enzymes (CAZymes) involved in numerous biologically important chemical transformations, such as the degradation of complex dietary polysaccharides. Coupling metagenomic analyses to structural genomics initiatives and to biochemical and other functional assays of CAZymes will be essential for determining how these as well as other microbiome-encoded proteins operate to shape the properties of microbial communities and their human hosts

  4. A physical complex of the Fanconi anemia proteins FANCG/XRCC9 and FANCA

    Science.gov (United States)

    Waisfisz, Quinten; de Winter, Johan P.; Kruyt, Frank A. E.; de Groot, Jan; van der Weel, Laura; Dijkmans, Lonneke M.; Zhi, Yu; Arwert, Fré; Scheper, Rik J.; Youssoufian, Hagop; Hoatlin, Maureen E.; Joenje, Hans

    1999-01-01

    Fanconi anemia (FA) is a recessively inherited disease characterized at the cellular level by spontaneous chromosomal instability and specific hypersensitivity to cross-linking agents. FA is genetically heterogeneous, comprising at least eight complementation groups (A-H). We report that the protein encoded by the gene mutated in complementation group G (FANCG) localizes to the cytoplasm and nucleus of the cell and assembles in a molecular complex with the FANCA protein, both in vivo and in vitro. Endogenous FANCA/FANCG complex was detected in both non-FA cells and in FA cells from groups D and E. By contrast, no complex was detected in specific cell lines belonging to groups A and G, whereas reduced levels were found in cells from groups B, C, F, and H. Wild-type levels of FANCA/FANCG complex were restored upon correction of the cellular phenotype by transfection or cell fusion experiments, suggesting that this complex is of functional significance in the FA pathway. These results indicate that the cellular FA phenotype can be connected to three biochemical subtypes based on the levels of FANCA/FANCG complex. Disruption of the complex may provide an experimental strategy for chemosensitization of neoplastic cells. PMID:10468606

  5. Stress tolerances of nullmutants of function-unknown genes encoding menadione stress-responsive proteins in Aspergillus nidulans.

    Science.gov (United States)

    Leiter, Éva; Bálint, Mihály; Miskei, Márton; Orosz, Erzsébet; Szabó, Zsuzsa; Pócsi, István

    2016-07-01

    A group of menadione stress-responsive function-unkown genes of Aspergillus nidulans (Locus IDs ANID_03987.1, ANID_06058.1, ANID_10219.1, and ANID_10260.1) was deleted and phenotypically characterized. Importantly, comparative and phylogenetic analyses of the tested A. nidulans genes and their orthologs shed light only on the presence of a TANGO2 domain with NRDE protein motif in the translated ANID_06058.1 gene but did not reveal any recognizable protein-encoding domains in other protein sequences. The gene deletion strains were subjected to oxidative, osmotic, and metal ion stress and, surprisingly, only the ΔANID_10219.1 mutant showed an increased sensitivity to 0.12 mmol l(-1) menadione sodium bisulfite. The gene deletions affected the stress sensitivities (tolerances) irregularly, for example, some strains grew more slowly when exposed to various oxidants and/or osmotic stress generating agents, meanwhile the ΔANID_10260.1 mutant possessed a wild-type tolerance to all stressors tested. Our results are in line with earlier studies demonstrating that the deletions of stress-responsive genes do not confer necessarily any stress-sensitivity phenotypes, which can be attributed to compensatory mechanisms based on other elements of the stress response system with overlapping functions. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Quantification and Classification of E. coli Proteome Utilization and Unused Protein Costs across Environments.

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    Edward J O'Brien

    2016-06-01

    Full Text Available The costs and benefits of protein expression are balanced through evolution. Expression of un-utilized protein (that have no benefits in the current environment incurs a quantifiable fitness costs on cellular growth rates; however, the magnitude and variability of un-utilized protein expression in natural settings is unknown, largely due to the challenge in determining environment-specific proteome utilization. We address this challenge using absolute and global proteomics data combined with a recently developed genome-scale model of Escherichia coli that computes the environment-specific cost and utility of the proteome on a per gene basis. We show that nearly half of the proteome mass is unused in certain environments and accounting for the cost of this unused protein expression explains >95% of the variance in growth rates of Escherichia coli across 16 distinct environments. Furthermore, reduction in unused protein expression is shown to be a common mechanism to increase cellular growth rates in adaptive evolution experiments. Classification of the unused protein reveals that the unused protein encodes several nutrient- and stress- preparedness functions, which may convey fitness benefits in varying environments. Thus, unused protein expression is the source of large and pervasive fitness costs that may provide the benefit of hedging against environmental change.

  7. Potyvirus helper component-proteinase self-interaction in the yeast two-hybrid system and delineation of the interaction domain involved.

    Science.gov (United States)

    Urcuqui-Inchima, S; Walter, J; Drugeon, G; German-Retana, S; Haenni, A L; Candresse, T; Bernardi, F; Le Gall, O

    1999-05-25

    Using the yeast two-hybrid system, a screen was performed for possible interactions between the proteins encoded by the 5' region of potyviral genomes [P1, helper component-proteinase (HC-Pro), and P3]. A positive self-interaction involving HC-Pro was detected with lettuce mosaic virus (LMV) and potato virus Y (PVY). The possibility of heterologous interaction between the HC-Pro of LMV and of PVY was also demonstrated. No interaction involving either the P1 or the P3 proteins was detected. A series of ordered deletions from either the N- or C-terminal end of the LMV HC-Pro was used to map the domain involved in interaction to the 72 N-terminal amino acids of the protein, a region known to be dispensable for virus viability but necessary for aphid transmission. A similar but less detailed analysis mapped the interacting domain to the N-terminal half of the PVY HC-Pro. Copyright 1999 Academic Press.

  8. Two Crinivirus-specific proteins of Lettuce infectious yellows virus (LIYV), P26 and P9, are self-interacting.

    Science.gov (United States)

    Stewart, Lucy R; Hwang, Min Sook; Falk, Bryce W

    2009-11-01

    Interactions of Lettuce infectious yellows virus (LIYV)-encoded proteins were tested by yeast-two-hybrid (Y2H) assays. LIYV-encoded P34, Hsp70h, P59, CP, CPm, and P26 were tested in all possible pairwise combinations. Interaction was detected only for the P26-P26 combination. P26 self-interaction domains were mapped using a series of N- and C-terminal truncations. Orthologous P26 proteins from the criniviruses Beet pseudoyellows virus (BPYV), Cucurbit yellow stunting disorder virus (CYSDV), and Lettuce chlorosis virus (LCV) were also tested, and each exhibited strong self-interaction but no interaction with orthologous proteins. Two small putative proteins encoded by LIYV RNA2, P5 and P9, were also tested for interactions with the six aforementioned LIYV proteins and each other. No interactions were detected for P5, but P9-P9 self-interaction was detected. P26- and P9-encoding genes are present in all described members of the genus Crinivirus, but are not present in other members of the family Closteroviridae. LIYV P26 has previously been demonstrated to induce a unique LIYV cytopathology, plasmalemma deposits (PLDs), but no role is yet known for P9.

  9. Effect of metal sulfide pulp density on gene expression of electron transporters in Acidithiobacillus sp. FJ2.

    Science.gov (United States)

    Fatemi, Faezeh; Miri, Saba; Jahani, Samaneh

    2017-05-01

    In Acidithiobacillus ferrooxidans, one of the most important bioleaching bacterial species, the proteins encoded by the rus operon are involved in the electron transfer from Fe 2+ to O 2 . To obtain further knowledge about the mechanism(s) involved in the adaptive responses of the bacteria to growth on the different uranium ore pulp densities, we analyzed the expression of the four genes from the rus operon by real-time PCR, when Acidithiobacillus sp. FJ2 was grown in the presence of different uranium concentrations. The uranium bioleaching results showed the inhibitory effects of the metal pulp densities on the oxidation activity of the bacteria which can affect Eh, pH, Fe oxidation and uranium extractions. Gene expression analysis indicated that Acidithiobacillus sp. FJ2 tries to survive in the stress with increasing in the expression levels of cyc2, cyc1, rus and coxB, but the metal toxicity has a negative effect on the gene expression in different pulp densities. These results indicated that Acidithiobacillus sp. FJ2 could leach the uranium even in high pulp density (50%) by modulation in rus operon gene responses.

  10. Combined DECS Analysis and Next-Generation Sequencing Enable Efficient Detection of Novel Plant RNA Viruses

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    Hironobu Yanagisawa

    2016-03-01

    Full Text Available The presence of high molecular weight double-stranded RNA (dsRNA within plant cells is an indicator of infection with RNA viruses as these possess genomic or replicative dsRNA. DECS (dsRNA isolation, exhaustive amplification, cloning, and sequencing analysis has been shown to be capable of detecting unknown viruses. We postulated that a combination of DECS analysis and next-generation sequencing (NGS would improve detection efficiency and usability of the technique. Here, we describe a model case in which we efficiently detected the presumed genome sequence of Blueberry shoestring virus (BSSV, a member of the genus Sobemovirus, which has not so far been reported. dsRNAs were isolated from BSSV-infected blueberry plants using the dsRNA-binding protein, reverse-transcribed, amplified, and sequenced using NGS. A contig of 4,020 nucleotides (nt that shared similarities with sequences from other Sobemovirus species was obtained as a candidate of the BSSV genomic sequence. Reverse transcription (RT-PCR primer sets based on sequences from this contig enabled the detection of BSSV in all BSSV-infected plants tested but not in healthy controls. A recombinant protein encoded by the putative coat protein gene was bound by the BSSV-antibody, indicating that the candidate sequence was that of BSSV itself. Our results suggest that a combination of DECS analysis and NGS, designated here as “DECS-C,” is a powerful method for detecting novel plant viruses.

  11. Gene expression of a two-component regulatory system associated with sunscreen biosynthesis in the cyanobacterium Nostoc punctiforme ATCC 29133.

    Science.gov (United States)

    Janssen, Jacob; Soule, Tanya

    2016-01-01

    Long-wavelength ultraviolet radiation (UVA) can damage cells through photooxidative stress, leading to harmful photosensitized proteins and pigments in cyanobacteria. To mitigate damage, some cyanobacteria secrete the UVA-absorbing pigment scytonemin into their extracellular sheath. Comparative genomic analyses suggest that scytonemin biosynthesis is regulated by the two-component regulatory system (TCRS) proteins encoded by Npun_F1277 and Npun_F1278 in the cyanobacterium Nostoc punctiforme ATCC 29133. To understand the dynamics of these genes, their expression was measured following exposure to UVA, UVB, high visible (VIS) irradiance and oxidative stress for 20, 40 and 60 min. Overall, both genes had statistically similar patterns of expression for all four conditions and were generally upregulated, except for those exposed to UVB by 60 min and for the cells under oxidative stress. The greatest UVA response was an upregulation by 20 min, while the response to UVB was the most dramatic and persisted through 40 min. High VIS irradiance resulted in a modest upregulation, while oxidative stress caused a slight downregulation. Both genes were also found to occur on the same transcript. These results demonstrate that these genes are positively responding to several light-associated conditions, which suggests that this TCRS may regulate more than just scytonemin biosynthesis under UVA stress. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  12. Unravelling the cross-talk between iron starvation and oxidative stress responses highlights the key role of PerR (alr0957) in peroxide signalling in the cyanobacterium Nostoc PCC 7120.

    Science.gov (United States)

    Yingping, Fan; Lemeille, Sylvain; Talla, Emmanuel; Janicki, Annick; Denis, Yann; Zhang, Cheng-Cai; Latifi, Amel

    2014-10-01

    The cyanobacterial phylum includes oxygenic photosynthetic prokaryotes of a wide variety of morphologies, metabolisms and ecologies. Their adaptation to their various ecological niches is mainly achieved by sophisticated regulatory mechanisms and depends on a fine cross-talk between them. We assessed the global transcriptomic response of the filamentous cyanobacterium Nostoc PCC 7120 to iron starvation and oxidative stress. More than 20% of the differentially expressed genes in response to iron stress were also responsive to oxidative stress. These transcripts include antioxidant proteins-encoding genes that confirms that iron depletion leads to reactive oxygen accumulation. The activity of the Fe-superoxide dismutase was not significantly decreased under iron starvation, indicating that the oxidative stress generated under iron deficiency is not a consequence of (SOD) deficiency. The transcriptional data indicate that the adaptation of Nostoc to iron-depleted conditions displays important differences with what has been shown in unicellular cyanobacteria. While the FurA protein that regulates the response to iron deprivation has been well characterized in Nostoc, the regulators in charge of the oxidative stress response are unknown. Our study indicates that the alr0957 (perR) gene encodes the master regulator of the peroxide stress. PerR is a peroxide-sensor repressor that senses peroxide by metal-catalysed oxidation.

  13. Approaches and Perspectives for Development of African Swine Fever Virus Vaccines

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    Marisa Arias

    2017-10-01

    Full Text Available African swine fever (ASF is a complex disease of swine, caused by a large DNA virus belonging to the family Asfarviridae. The disease shows variable clinical signs, with high case fatality rates, up to 100%, in the acute forms. ASF is currently present in Africa and Europe where it circulates in different scenarios causing a high socio-economic impact. In most affected regions, control has not been effective in part due to lack of a vaccine. The availability of an effective and safe ASFV vaccines would support and enforce control–eradication strategies. Therefore, work leading to the rational development of protective ASF vaccines is a high priority. Several factors have hindered vaccine development, including the complexity of the ASF virus particle and the large number of proteins encoded by its genome. Many of these virus proteins inhibit the host’s immune system thus facilitating virus replication and persistence. We review previous work aimed at understanding ASFV–host interactions, including mechanisms of protective immunity, and approaches for vaccine development. These include live attenuated vaccines, and “subunit” vaccines, based on DNA, proteins, or virus vectors. In the shorter to medium term, live attenuated vaccines are the most promising and best positioned candidates. Gaps and future research directions are evaluated.

  14. Human case of bacteremia caused by Streptococcus canis sequence type 9 harboring the scm gene.

    Science.gov (United States)

    Taniyama, Daisuke; Abe, Yoshihiko; Sakai, Tetsuya; Kikuchi, Takahide; Takahashi, Takashi

    2017-01-01

    Streptococcus canis (Sc) is a zoonotic pathogen that is transferred mainly from companion animals to humans. One of the major virulence factors in Sc is the M-like protein encoded by the scm gene, which is involved in anti-phagocytic activities, as well as the recruitment of plasminogen to the bacterial surface in cooperation with enolase, and the consequent enhancement of bacterial transmigration and survival. This is the first reported human case of uncomplicated bacteremia following a dog bite, caused by Streptococcus canis harboring the scm gene. The similarity of the 16S rRNA from the infecting species to that of the Sc type strain, as well as the amplification of the species-specific cfg gene, encoding a co-hemolysin, was used to confirm the species identity. Furthermore, the isolate was confirmed as sequence type 9. The partial scm gene sequence harbored by the isolate was closely related to those of other two Sc strains. While this isolate did not possess the erm (A), erm (B), or mef (A), macrolide/lincosamide resistance genes, it was not susceptible to azithromycin: its susceptibility was intermediate. Even though human Sc bacteremia is rare, clinicians should be aware of this microorganism, as well as Pasteurella sp., Prevotella sp., and Capnocytophaga sp., when examining and treating patients with fever who maintain close contact with companion animals.

  15. Tuberculosis: looking beyond BCG vaccines.

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    Mustafa Abu S

    2003-01-01

    Full Text Available Tuberculosis (TB is an infectious disease of international importance and ranks among the top 10 causes of death in the World. About one-third of the world′s population is infected with Mycobacterium tuberculosis. Every year, approximately eight million people develop active disease and two million die of TB. The currently used BCG vaccines have shown variable protective efficacies against TB in different parts of the world. Moreover, being a live vaccine, BCG can be pathogenic in immunocompromised recipients. Therefore, there is an urgent need to develop new vaccines against TB. The comparative genome analysis has revealed the existence of several M. tuberculosis-specific regions that are deleted in BCG. The work carried out to determine the immunological reactivity of proteins encoded by genes located in these regions revealed several major antigens of M. tuberculosis, including the 6 kDa early secreted antigen target (ESAT6. Immunization with ESAT6 and its peptide (aa51-70 protects mice challenged with M. tuberculosis. The protective efficacy of immunization further improves when ESAT6 is recombinantly fused with M. tuberculosis antigen 85B. In addition, ESAT6 delivered as a DNA vaccine is also protective in mice. Whether these vaccines would be safe or not cannot be speculated. The answer regarding the safety and efficacy of these vaccines has to await human trials in different parts of the world.

  16. Plant Adaptation to Acid Soils: The Molecular Basis for Crop Aluminum Resistance.

    Science.gov (United States)

    Kochian, Leon V; Piñeros, Miguel A; Liu, Jiping; Magalhaes, Jurandir V

    2015-01-01

    Aluminum (Al) toxicity in acid soils is a significant limitation to crop production worldwide, as approximately 50% of the world's potentially arable soil is acidic. Because acid soils are such an important constraint to agriculture, understanding the mechanisms and genes conferring resistance to Al toxicity has been a focus of intense research interest in the decade since the last article on crop acid soil tolerance was published in this journal. An impressive amount of progress has been made during that time that has greatly increased our understanding of the diversity of Al resistance genes and mechanisms, how resistance gene expression is regulated and triggered by Al and Al-induced signals, and how the proteins encoded by these genes function and are regulated. This review examines the state of our understanding of the physiological, genetic, and molecular bases for crop Al tolerance, looking at the novel Al resistance genes and mechanisms that have been identified over the past ten years. Additionally, it examines how the integration of molecular and genetic analyses of crop Al resistance is starting to be exploited for the improvement of crop plants grown on acid soils via both molecular-assisted breeding and biotechnology approaches.

  17. Reversible Heat-Induced Inactivation of Chimeric β-Glucuronidase in Transgenic Plants1

    Science.gov (United States)

    Almoguera, Concepción; Rojas, Anabel; Jordano, Juan

    2002-01-01

    We compared the expression patterns in transgenic tobacco (Nicotiana tabacum) of two chimeric genes: a translational fusion to β-glucuronidase (GUS) and a transcriptional fusion, both with the same promoter and 5′-flanking sequences of Ha hsp17.7 G4, a small heat shock protein (sHSP) gene from sunflower (Helianthus annuus). We found that immediately after heat shock, the induced expression from the two fusions in seedlings was similar, considering chimeric mRNA or GUS protein accumulation. Surprisingly, we discovered that the chimeric GUS protein encoded by the translational fusion was mostly inactive in such conditions. We also found that this inactivation was fully reversible. Thus, after returning to control temperature, the GUS activity was fully recovered without substantial changes in GUS protein accumulation. In contrast, we did not find differences in the in vitro heat inactivation of the respective GUS proteins. Insolubilization of the chimeric GUS protein correlated with its inactivation, as indicated by immunoprecipitation analyses. The inclusion in another chimeric gene of the 21 amino-terminal amino acids from a different sHSP lead to a comparable reversible inactivation. That effect not only illustrates unexpected post-translational problems, but may also point to sequences involved in interactions specific to sHSPs and in vivo heat stress conditions. PMID:12011363

  18. Transcription and translation products of the cytolysin gene psm-mec on the mobile genetic element SCCmec regulate Staphylococcus aureus virulence.

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    Chikara Kaito

    Full Text Available The F region downstream of the mecI gene in the SCCmec element in hospital-associated methicillin-resistant Staphylococcus aureus (HA-MRSA contains two bidirectionally overlapping open reading frames (ORFs, the fudoh ORF and the psm-mec ORF. The psm-mec ORF encodes a cytolysin, phenol-soluble modulin (PSM-mec. Transformation of the F region into the Newman strain, which is a methicillin-sensitive S. aureus (MSSA strain, or into the MW2 (USA400 and FRP3757 (USA300 strains, which are community-acquired MRSA (CA-MRSA strains that lack the F region, attenuated their virulence in a mouse systemic infection model. Introducing the F region to these strains suppressed colony-spreading activity and PSMα production, and promoted biofilm formation. By producing mutations into the psm-mec ORF, we revealed that (i both the transcription and translation products of the psm-mec ORF suppressed colony-spreading activity and promoted biofilm formation; and (ii the transcription product of the psm-mec ORF, but not its translation product, decreased PSMα production. These findings suggest that both the psm-mec transcript, acting as a regulatory RNA, and the PSM-mec protein encoded by the gene on the mobile genetic element SCCmec regulate the virulence of Staphylococcus aureus.

  19. The role of HLA-E polymorphism in immunological response

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    Milena Iwaszko

    2011-09-01

    Full Text Available The HLA-E protein is one of the most extensively studied MHC class Ib antigens and the least polymorphic one compared to other MHC class I molecules. In the human population there have been reported just ten alleles encoding three different peptides. Only two of these alleles, namely HLA-E*0101 and HLA-E*0103, are widely distributed (around 50�0each. The proteins encoded by these alleles differ from each other in one amino acid at position 107. In HLA-E*0101 it is arginine and in HLA-E*0103 it is glycine. The difference between these proteins manifests itself in surface expression levels, affinities to leader peptides and thermal stabilities of their complexes.The HLA-E molecule is a ligand for CD94/NKG2 receptors on NK cells and TCR receptors on NK-CTL (NK-cytotoxic T lymphocyte cells, so it plays a double role in both innate and adaptive immunity. This paper reviews the knowledge on the role of the HLA-E molecule in the immunological response. Aspects related to polymorphism of the HLA-E gene and the course of several diseases including type I diabetes, ankylosing spondylitis, HCV and HIV infections, nasopharyngeal cancer and recurrent spontaneous abortions, as well as the outcome of hematopoietic stem cell transplantation, are presented and discussed in more detail.

  20. The Mitochondrion-Located Protein OsB12D1 Enhances Flooding Tolerance during Seed Germination and Early Seedling Growth in Rice

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    Dongli He

    2014-07-01

    Full Text Available B12D belongs to a function unknown subgroup of the Balem (Barley aleurone and embryo proteins. In our previous work on rice seed germination, we identified a B12D-like protein encoded by LOC_Os7g41350 (named OsB12D1. OsB12D1 pertains to an ancient protein family with an amino acid sequence highly conserved from moss to angiosperms. Among the six OsB12Ds, OsB12D1 is one of the major transcripts and is primarily expressed in germinating seed and root. Bioinformatics analyses indicated that OsB12D1 is an anoxic or submergence resistance-related gene. RT-PCR results showed OsB12D1 is induced remarkably in the coleoptiles or roots by flooding during seed germination and early seedling growth. The OsB12D1-overexpressed rice seeds could protrude radicles in 8 cm deep water, further exhibiting significant flooding tolerance compared to the wild type. Moreover, this tolerance was not affected by the gibberellin biosynthesis inhibitor paclobutrazol. OsB12D1 was identified in the mitochondrion by subcellular localization analysis and possibly enhances electron transport through mediating Fe and oxygen availability under flooded conditions. This work indicated that OsB12D1 is a promising gene that can help to enhance rice seedling establishment in farming practices, especially for direct seeding.

  1. A new member of a family of ATPases is essential for assembly of mitochondrial respiratory chain and ATP synthetase complexes in Saccharomyces cerevisiae.

    Science.gov (United States)

    Tzagoloff, A; Yue, J; Jang, J; Paul, M F

    1994-10-21

    Respiration-defective pet mutants of Saccharomyces cerevisiae, assigned to complementation group G25, are grossly deficient in mitochondrial respiratory and ATPase complexes. This phenotype is usually found in strains impaired in mitochondrial protein synthesis. The G25 mutants, however, synthesize all of the proteins encoded by mitochondrial DNA. The mutants are also able to import and process cytoplasmically derived subunits of these enzymes. These results are most compatible with the idea that the gene defined by G25 mutants (RCA1) codes for a protein essential for the assembly of functional respiratory and ATPase complexes. The RCA1 gene has been cloned by complementation of an rca1 mutant with a yeast genomic library. The sequence of the encoded product shows Rca1 protein to be a new member of a recently described family of ATPases. The Rca1 protein is a mitochondrial membrane protein and is the third known member of this family implicated to function in the biogenesis of mitochondria. The primary structure of Rca1 protein indicates several distinct domains in addition to the common purine nucleotide binding region shared by all members of this protein family. One, located in the amino-terminal half, contains two hydrophobic stretches of sufficient length to span a membrane lipid bilayer.

  2. Four regulatory elements in the human c-fos promoter mediate transactivation by HTLV-1 Tax protein.

    Science.gov (United States)

    Alexandre, C; Verrier, B

    1991-04-01

    Expression of the human c-fos proto-oncogene is activated in trans by the Tax protein encoded by human T-cell leukemia virus type-1 (HTLV-1). Indeed, we show here that a HeLa clone stably transfected by Tax expresses Fos at a high level. We also show that multiple elements of the human c-fos promoter, i.e. the v-sis conditioned medium inducible element (SIE), the dyad symmetry element (DSE) necessary for growth factor induction, the octanucleotide direct repeat element (DR), and the cyclic AMP response element (CRE) centred at -60, can all mediate Tax transactivation. In the DSE, the 10bp central core that binds the serum response factor (SRF) is, by itself, sufficient to mediate Tax transactivation. Moreover, a CRE-binding protein is involved in Tax activation through the CRE-60 element. Since Fos is a transregulator of cellular genes, our results suggest that the oncoprotein plays a crucial role in T-cell transformation by HTLV-1 in conjunction with other Tax-inducible genes.

  3. Toscana virus NSs protein promotes degradation of double-stranded RNA-dependent protein kinase.

    Science.gov (United States)

    Kalveram, Birte; Ikegami, Tetsuro

    2013-04-01

    Toscana virus (TOSV), which is transmitted by Phlebotomus spp. sandflies, is a major etiologic agent of aseptic meningitis and encephalitis in the Mediterranean. Like other members of the genus Phlebovirus of the family Bunyaviridae, TOSV encodes a nonstructural protein (NSs) in its small RNA segment. Although the NSs of Rift Valley fever virus (RVFV) has been identified as an important virulence factor, which suppresses host general transcription, inhibits transcription from the beta interferon promoter, and promotes the proteasomal degradation of double-stranded RNA-dependent protein kinase (PKR), little is known about the functions of NSs proteins encoded by less-pathogenic members of this genus. In this study we report that TOSV is able to downregulate PKR with similar efficiency as RVFV, while infection with the other phleboviruses-i.e., Punta Toro virus, sandfly fever Sicilian virus, or Frijoles virus-has no effect on cellular PKR levels. In contrast to RVFV, however, cellular transcription remains unaffected during TOSV infection. TOSV NSs protein promotes the proteasome-dependent downregulation of PKR and is able to interact with kinase-inactive PKR in infected cells.

  4. MicroRNAs: From Female Fertility, Germ Cells, and Stem Cells to Cancer in Humans

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    Irma Virant-Klun

    2016-01-01

    Full Text Available MicroRNAs are a family of naturally occurring small noncoding RNA molecules that play an important regulatory role in gene expression. They are suggested to regulate a large proportion of protein encoding genes by mediating the translational suppression and posttranscriptional control of gene expression. Recent findings show that microRNAs are emerging as important regulators of cellular differentiation and dedifferentiation, and are deeply involved in developmental processes including human preimplantation development. They keep a balance between pluripotency and differentiation in the embryo and embryonic stem cells. Moreover, it became evident that dysregulation of microRNA expression may play a fundamental role in progression and dissemination of different cancers including ovarian cancer. The interest is still increased by the discovery of exosomes, that is, cell-derived vesicles, which can carry different proteins but also microRNAs between different cells and are involved in cell-to-cell communication. MicroRNAs, together with exosomes, have a great potential to be used for prognosis, therapy, and biomarkers of different diseases including infertility. The aim of this review paper is to summarize the existent knowledge on microRNAs related to female fertility and cancer: from primordial germ cells and ovarian function, germinal stem cells, oocytes, and embryos to embryonic stem cells.

  5. A Survey of Protein Structures from Archaeal Viruses

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    Nikki Dellas

    2013-01-01

    Full Text Available Viruses that infect the third domain of life, Archaea, are a newly emerging field of interest. To date, all characterized archaeal viruses infect archaea that thrive in extreme conditions, such as halophilic, hyperthermophilic, and methanogenic environments. Viruses in general, especially those replicating in extreme environments, contain highly mosaic genomes with open reading frames (ORFs whose sequences are often dissimilar to all other known ORFs. It has been estimated that approximately 85% of virally encoded ORFs do not match known sequences in the nucleic acid databases, and this percentage is even higher for archaeal viruses (typically 90%–100%. This statistic suggests that either virus genomes represent a larger segment of sequence space and/or that viruses encode genes of novel fold and/or function. Because the overall three-dimensional fold of a protein evolves more slowly than its sequence, efforts have been geared toward structural characterization of proteins encoded by archaeal viruses in order to gain insight into their potential functions. In this short review, we provide multiple examples where structural characterization of archaeal viral proteins has indeed provided significant functional and evolutionary insight.

  6. Transposases are the most abundant, most ubiquitous genes in nature.

    Science.gov (United States)

    Aziz, Ramy K; Breitbart, Mya; Edwards, Robert A

    2010-07-01

    Genes, like organisms, struggle for existence, and the most successful genes persist and widely disseminate in nature. The unbiased determination of the most successful genes requires access to sequence data from a wide range of phylogenetic taxa and ecosystems, which has finally become achievable thanks to the deluge of genomic and metagenomic sequences. Here, we analyzed 10 million protein-encoding genes and gene tags in sequenced bacterial, archaeal, eukaryotic and viral genomes and metagenomes, and our analysis demonstrates that genes encoding transposases are the most prevalent genes in nature. The finding that these genes, classically considered as selfish genes, outnumber essential or housekeeping genes suggests that they offer selective advantage to the genomes and ecosystems they inhabit, a hypothesis in agreement with an emerging body of literature. Their mobile nature not only promotes dissemination of transposable elements within and between genomes but also leads to mutations and rearrangements that can accelerate biological diversification and--consequently--evolution. By securing their own replication and dissemination, transposases guarantee to thrive so long as nucleic acid-based life forms exist.

  7. Sexual selection, genetic conflict, selfish genes, and the atypical patterns of gene expression in spermatogenic cells.

    Science.gov (United States)

    Kleene, Kenneth C

    2005-01-01

    This review proposes that the peculiar patterns of gene expression in spermatogenic cells are the consequence of powerful evolutionary forces known as sexual selection. Sexual selection is generally characterized by intense competition of males for females, an enormous variety of the strategies to maximize male reproductive success, exaggerated male traits at all levels of biological organization, co-evolution of sexual traits in males and females, and conflict between the sexual advantage of the male trait and the reproductive fitness of females and the individual fitness of both sexes. In addition, spermatogenesis is afflicted by selfish genes that promote their transmission to progeny while causing deleterious effects. Sexual selection, selfish genes, and genetic conflict provide compelling explanations for many atypical features of gene expression in spermatogenic cells including the gross overexpression of certain mRNAs, transcripts encoding truncated proteins that cannot carry out basic functions of the proteins encoded by the same genes in somatic cells, the large number of gene families containing paralogous genes encoding spermatogenic cell-specific isoforms, the large number of testis-cancer-associated genes that are expressed only in spermatogenic cells and malignant cells, and the overbearing role of Sertoli cells in regulating the number and quality of spermatozoa.

  8. Association of Serum Adropin Concentrations with Diabetic Nephropathy

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    Wenchao Hu

    2016-01-01

    Full Text Available Objective. Adropin is a newly identified regulatory protein encoded by the Enho gene and is critically involved in energy homeostasis and insulin sensitivity. This study aims to determine the correlation of serum adropin concentrations with diabetic nephropathy (DN. Methods. This study consisted of 245 patients with type 2 diabetes mellitus (T2DM and 81 healthy subjects. Then T2DM patients were divided into normoalbuminuria, microalbuminuria, and macroalbuminuria subgroups based on urine albumin to creatinine ratio (ACR. Results. T2DM patients showed significantly lower serum adropin concentrations than those in the controls. T2DM patients with macroalbuminuria had significantly decreased serum adropin concentrations compared with the other three groups. In addition, T2DM patients with microalbuminuria showed lower serum adropin concentrations than those in patients with normoalbuminuria. Logistic regression analysis showed that serum adropin was correlated with decreased risk of developing T2DM and DN. Pearson correlation analysis indicated that serum adropin was negatively correlated with body mass index (BMI, blood urea nitrogen, creatinine, and ACR and positively correlated with glomerular filtration rate. Furthermore, multiple linear regression analysis showed that BMI and ACR were negatively correlated with serum adropin levels. Conclusion. Serum adropin concentrations are negatively associated with renal function. Adropin may be implicated in the pathogenesis of DN development.

  9. Chlamydia trachomatis protein CT009 is a structural and functional homolog to the key morphogenesis component RodZ and interacts with division septal plane localized MreB.

    Science.gov (United States)

    Kemege, Kyle E; Hickey, John M; Barta, Michael L; Wickstrum, Jason; Balwalli, Namita; Lovell, Scott; Battaile, Kevin P; Hefty, P Scott

    2015-02-01

    Cell division in Chlamydiae is poorly understood as apparent homologs to most conserved bacterial cell division proteins are lacking and presence of elongation (rod shape) associated proteins indicate non-canonical mechanisms may be employed. The rod-shape determining protein MreB has been proposed as playing a unique role in chlamydial cell division. In other organisms, MreB is part of an elongation complex that requires RodZ for proper function. A recent study reported that the protein encoded by ORF CT009 interacts with MreB despite low sequence similarity to RodZ. The studies herein expand on those observations through protein structure, mutagenesis and cellular localization analyses. Structural analysis indicated that CT009 shares high level of structural similarity to RodZ, revealing the conserved orientation of two residues critical for MreB interaction. Substitutions eliminated MreB protein interaction and partial complementation provided by CT009 in RodZ deficient Escherichia coli. Cellular localization analysis of CT009 showed uniform membrane staining in Chlamydia. This was in contrast to the localization of MreB, which was restricted to predicted septal planes. MreB localization to septal planes provides direct experimental observation for the role of MreB in cell division and supports the hypothesis that it serves as a functional replacement for FtsZ in Chlamydia. © 2014 John Wiley & Sons Ltd.

  10. Acute guttate psoriasis patients have positive streptococcus hemolyticus throat cultures and elevated antistreptococcal M6 protein titers.

    Science.gov (United States)

    Zhao, Guang; Feng, Xiaoling; Na, Aihua; Yongqiang, Jiang; Cai, Qing; Kong, Jian; Ma, Huijun

    2005-02-01

    To further study the role of Streptococci hemolyticus infection and streptococcal M6 protein in the pathogenesis of acute guttate psoriasis, streptococcal cultures were taken from the throats of 68 patients with acute guttate psoriasis. PCR technique was applied to detect M6 protein encoding DNA from those cultured streptococci. Pure M6 protein was obtained by Sephacry/S-200HR and Mono-Q chromatography from proliferated Streptococcus hemolyticus. Antistreptococcal M6 protein titers were measured in the serum of patients with acute guttate psoriasis, plaque psoriasis and healthy controls by ELISA. A high incidence of Streptococcus hemolyticus culture was observed in the guttate psoriatic group compared with the plaque psoriasis and control groups. Fourteen strains of Streptococcus hemolyticus were cultured from the throats of 68 acute guttate psoriasis patients. Of these, 5 strains contain DNA encoding the M6 protein gene as confirmed by PCR technique. More than 85% purification of M6 protein was obtained from Streptococcus pyogenes. Applying our pure M6 protein with the ELISA methods, we found that the titer of antistreptococcal M6 protein was significantly higher in the serum of guttate psoriasis patients than in the control or plaque psoriasis groups (P M6 protein in their sera.

  11. Evidence for systems-level molecular mechanisms of tumorigenesis

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    Capellá Gabriel

    2007-06-01

    Full Text Available Abstract Background Cancer arises from the consecutive acquisition of genetic alterations. Increasing evidence suggests that as a consequence of these alterations, molecular interactions are reprogrammed in the context of highly connected and regulated cellular networks. Coordinated reprogramming would allow the cell to acquire the capabilities for malignant growth. Results Here, we determine the coordinated function of cancer gene products (i.e., proteins encoded by differentially expressed genes in tumors relative to healthy tissue counterparts, hereafter referred to as "CGPs" defined as their topological properties and organization in the interactome network. We show that CGPs are central to information exchange and propagation and that they are specifically organized to promote tumorigenesis. Centrality is identified by both local (degree and global (betweenness and closeness measures, and systematically appears in down-regulated CGPs. Up-regulated CGPs do not consistently exhibit centrality, but both types of cancer products determine the overall integrity of the network structure. In addition to centrality, down-regulated CGPs show topological association that correlates with common biological processes and pathways involved in tumorigenesis. Conclusion Given the current limited coverage of the human interactome, this study proposes that tumorigenesis takes place in a specific and organized way at the molecular systems-level and suggests a model that comprises the precise down-regulation of groups of topologically-associated proteins involved in particular functions, orchestrated with the up-regulation of specific proteins.

  12. Foldability of a Natural De Novo Evolved Protein.

    Science.gov (United States)

    Bungard, Dixie; Copple, Jacob S; Yan, Jing; Chhun, Jimmy J; Kumirov, Vlad K; Foy, Scott G; Masel, Joanna; Wysocki, Vicki H; Cordes, Matthew H J

    2017-11-07

    The de novo evolution of protein-coding genes from noncoding DNA is emerging as a source of molecular innovation in biology. Studies of random sequence libraries, however, suggest that young de novo proteins will not fold into compact, specific structures typical of native globular proteins. Here we show that Bsc4, a functional, natural de novo protein encoded by a gene that evolved recently from noncoding DNA in the yeast S. cerevisiae, folds to a partially specific three-dimensional structure. Bsc4 forms soluble, compact oligomers with high β sheet content and a hydrophobic core, and undergoes cooperative, reversible denaturation. Bsc4 lacks a specific quaternary state, however, existing instead as a continuous distribution of oligomer sizes, and binds dyes indicative of amyloid oligomers or molten globules. The combination of native-like and non-native-like properties suggests a rudimentary fold that could potentially act as a functional intermediate in the emergence of new folded proteins de novo. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Phylogenetic reconstruction of South American felids defined by protein electrophoresis.

    Science.gov (United States)

    Slattery, J P; Johnson, W E; Goldman, D; O'Brien, S J

    1994-09-01

    Phylogenetic associations among six closely related South American felid species were defined by changes in protein-encoding gene loci. We analyzed proteins isolated from skin fibroblasts using two-dimensional electrophoresis and allozymes extracted from blood cells. Genotypes were determined for multiple individuals of ocelot, margay, tigrina, Geoffroy's cat, kodkod, and pampas cat at 548 loci resolved by two-dimensional electrophoresis and 44 allozyme loci. Phenograms were constructed using the methods of Fitch-Margoliash and neighbor-joining on a matrix of Nei's unbiased genetic distances for all pairs of species. Results of a relative-rate test indicate changes in two-dimensional electrophoresis data are constant among all South American felids with respect to a hyena outgroup. Allelic frequencies were transformed to discrete character states for maximum parsimony analysis. Phylogenetic reconstruction indicates a major split occurred approximately 5-6 million years ago, leading to three groups within the ocelot lineage. The earliest divergence led to Leopardus tigrina, followed by a split between an ancestor of an unresolved trichotomy of three species (Oncifelis guigna, O. geoffroyi, and Lynchailuris colocolo) and a recent common ancestor of Leopardus pardalis and L. wiedii. The results suggest that modern South American felids are monophyletic and evolved rapidly after the formation of the Panama land bridge between North and South America.

  14. The FANTASTIC FOUR proteins influence shoot meristem size in Arabidopsis thaliana

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    Brand Luise H

    2010-12-01

    Full Text Available Abstract Background Throughout their lives plants produce new organs from groups of pluripotent cells called meristems, located at the tips of the shoot and the root. The size of the shoot meristem is tightly controlled by a feedback loop, which involves the homeodomain transcription factor WUSCHEL (WUS and the CLAVATA (CLV proteins. This regulatory circuit is further fine-tuned by morphogenic signals such as hormones and sugars. Results Here we show that a family of four plant-specific proteins, encoded by the FANTASTIC FOUR (FAF genes, has the potential to regulate shoot meristem size in Arabidopsis thaliana. FAF2 and FAF4 are expressed in the centre of the shoot meristem, overlapping with the site of WUS expression. Consistent with a regulatory interaction between the FAF gene family and WUS, our experiments indicate that the FAFs can repress WUS, which ultimately leads to an arrest of meristem activity in FAF overexpressing lines. The finding that meristematic expression of FAF2 and FAF4 is under negative control by CLV3 further supports the hypothesis that the FAFs are modulators of the genetic circuit that regulates the meristem. Conclusion This study reports the initial characterization of the Arabidopsis thaliana FAF gene family. Our data indicate that the FAF genes form a plant specific gene family, the members of which have the potential to regulate the size of the shoot meristem by modulating the CLV3-WUS feedback loop.

  15. The FANTASTIC FOUR proteins influence shoot meristem size in Arabidopsis thaliana.

    Science.gov (United States)

    Wahl, Vanessa; Brand, Luise H; Guo, Ya-Long; Schmid, Markus

    2010-12-22

    Throughout their lives plants produce new organs from groups of pluripotent cells called meristems, located at the tips of the shoot and the root. The size of the shoot meristem is tightly controlled by a feedback loop, which involves the homeodomain transcription factor WUSCHEL (WUS) and the CLAVATA (CLV) proteins. This regulatory circuit is further fine-tuned by morphogenic signals such as hormones and sugars. Here we show that a family of four plant-specific proteins, encoded by the FANTASTIC FOUR (FAF) genes, has the potential to regulate shoot meristem size in Arabidopsis thaliana. FAF2 and FAF4 are expressed in the centre of the shoot meristem, overlapping with the site of WUS expression. Consistent with a regulatory interaction between the FAF gene family and WUS, our experiments indicate that the FAFs can repress WUS, which ultimately leads to an arrest of meristem activity in FAF overexpressing lines. The finding that meristematic expression of FAF2 and FAF4 is under negative control by CLV3 further supports the hypothesis that the FAFs are modulators of the genetic circuit that regulates the meristem. This study reports the initial characterization of the Arabidopsis thaliana FAF gene family. Our data indicate that the FAF genes form a plant specific gene family, the members of which have the potential to regulate the size of the shoot meristem by modulating the CLV3-WUS feedback loop.

  16. An optimal method of iron starvation of the obligate intracellular pathogen, Chlamydia trachomatis

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    Christopher C. Thompson

    2011-02-01

    Full Text Available Iron is an essential cofactor in a number of critical biochemical reactions, and as such, its acquisition, storage, and metabolism is highly regulated in most organisms. The obligate intracellular bacterium, Chlamydia trachomatis experiences a developmental arrest when iron within the host is depleted. The nature of the iron starvation response in Chlamydia is relatively uncharacterized because of the likely inefficient method of iron depletion, which currently relies on the compound deferoxamine mesylate (DFO. Inefficient induction of the iron starvation response precludes the identification of iron-regulated genes. This report evaluated DFO with another iron chelator, 2,2’-bipyridyl (Bpdl and presented a systematic comparison of the two across a range of criteria in a single-treatment time-of-infection regimen. We demonstrate that the membrane permeable Bpdl was superior to DFO in the inhibition of chlamydia development, the induction of aberrant morphology, and the induction of an iron starvation transcriptional response in both host and bacteria. Furthermore, iron starvation using Bpdl identified the periplasmic iron binding protein-encoding ytgA gene as iron- responsive. Overall, the data present a compelling argument for the use of Bpdl, rather than DFO, in future iron starvation studies of chlamydia and other intracellular bacteria.

  17. Alterations in cellular energy metabolism associated with the antiproliferative effects of the ATM inhibitor KU-55933 and with metformin.

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    Mahvash Zakikhani

    Full Text Available KU-55933 is a specific inhibitor of the kinase activity of the protein encoded by Ataxia telangiectasia mutated (ATM, an important tumor suppressor gene with key roles in DNA repair. Unexpectedly for an inhibitor of a tumor suppressor gene, KU-55933 reduces proliferation. In view of prior preliminary evidence suggesting defective mitochondrial function in cells of patients with Ataxia Telangiectasia (AT, we examined energy metabolism of cells treated with KU-55933. The compound increased AMPK activation, glucose uptake and lactate production while reducing mitochondrial membrane potential and coupled respiration. The stimulation of glycolysis by KU-55933 did not fully compensate for the reduction in mitochondrial functions, leading to decreased cellular ATP levels and energy stress. These actions are similar to those previously described for the biguanide metformin, a partial inhibitor of respiratory complex I. Both compounds decreased mitochondrial coupled respiration and reduced cellular concentrations of fumarate, malate, citrate, and alpha-ketogluterate. Succinate levels were increased by KU-55933 levels and decreased by metformin, indicating that the effects of ATM inhibition and metformin are not identical. These observations suggest a role for ATM in mitochondrial function and show that both KU-55933 and metformin perturb the TCA cycle as well as oxidative phosphorylation.

  18. Quantification of gene expression modulation at the human genome wide induced by low doses of ionizing radiations

    International Nuclear Information System (INIS)

    Nosel, Ingrid

    2013-01-01

    The main goal of this study is to identify the molecular mechanisms involved in the response to low doses of ionizing radiation by using oligonucleotide micro-arrays. The results presented in this study demonstrate the relevance of this tool in the characterization of changes in gene expression at low doses of g radiation. All experimentations conducted were conducted on T CD4+ human lymphocytes. From this model we tried to characterize in 600 minutes after irradiation the molecular players in response to a dose range of 5 and 500 mGy. In this study, we highlight two types of molecular response. First, a dose-dependent response to irradiation involving groups of genes whose proteins are implicated in the response to DNA damage and in the p53 signaling pathway. The expression of the majority of the genes is modulated from 100 mGy. The second type of response is independent of the irradiation dose and the proteins encoded are involved in the mechanism of oxidative phosphorylation. Some of these genes could potentially be regulated by a family of transcription factor with an ETS domain. These ETS factors are known to be potential targets of the MAPKinase pathway. All these genes are modulated from 5 mGy and are therefore potential molecular players involved in the cellular response to ionizing stress with a low intensity. (author)

  19. Translation of the shallot virus X TGB3 gene depends on non-AUG initiation and leaky scanning.

    Science.gov (United States)

    Lezzhov, Alexander A; Gushchin, Vladimir A; Lazareva, Ekaterina A; Vishnichenko, Valery K; Morozov, Sergey Y; Solovyev, Andrey G

    2015-10-01

    Triple gene block (TGB), a conserved gene module found in the genomes of many filamentous and rod-shaped plant viruses, encodes three proteins, TGB1, TGB2 and TGB3, required for viral cell-to-cell movement through plasmodesmata and systemic transport via the phloem. The genome of Shallot virus X, the type species of the genus Allexivirus, includes TGB1 and TGB2 genes, but contains no canonical ORF for TGB3 protein. However, a TGB3-like protein-encoding sequence lacking an AUG initiator codon has been found in the shallot virus X (ShVX) genome in a position typical for TGB3 genes. This putative TGB3 gene is conserved in all allexiviruses. Here, we carried out sequence analysis to predict possible non-AUG initiator codons in the ShVX TGB3-encoding sequence. We further used an agroinfiltration assay in Nicotiana benthamiana to confirm this prediction. Site-directed mutagenesis was used to demonstrate that the ShVX TGB3 could be translated on a bicistronic mRNA template via a leaky scanning mechanism.

  20. Short hairpin RNA-mediated knockdown of protein expression in Entamoeba histolytica

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    Singh Upinder

    2009-02-01

    Full Text Available Abstract Background Entamoeba histolytica is an intestinal protozoan parasite of humans. The genome has been sequenced, but the study of individual gene products has been hampered by the lack of the ability to generate gene knockouts. We chose to test the use of RNA interference to knock down gene expression in Entamoeba histolytica. Results An episomal vector-based system, using the E. histolytica U6 promoter to drive expression of 29-basepair short hairpin RNAs, was developed to target protein-encoding genes in E. histolytica. The short hairpin RNAs successfully knocked down protein levels of all three unrelated genes tested with this system: Igl, the intermediate subunit of the galactose- and N-acetyl-D-galactosamine-inhibitable lectin; the transcription factor URE3-BP; and the membrane binding protein EhC2A. Igl levels were reduced by 72%, URE3-BP by 89%, and EhC2A by 97%. Conclusion Use of the U6 promoter to drive expression of 29-basepair short hairpin RNAs is effective at knocking down protein expression for unrelated genes in Entamoeba histolytica, providing a useful tool for the study of this parasite.

  1. Short hairpin RNA-mediated knockdown of protein expression in Entamoeba histolytica.

    Science.gov (United States)

    Linford, Alicia S; Moreno, Heriberto; Good, Katelyn R; Zhang, Hanbang; Singh, Upinder; Petri, William A

    2009-02-17

    Entamoeba histolytica is an intestinal protozoan parasite of humans. The genome has been sequenced, but the study of individual gene products has been hampered by the lack of the ability to generate gene knockouts. We chose to test the use of RNA interference to knock down gene expression in Entamoeba histolytica. An episomal vector-based system, using the E. histolytica U6 promoter to drive expression of 29-basepair short hairpin RNAs, was developed to target protein-encoding genes in E. histolytica. The short hairpin RNAs successfully knocked down protein levels of all three unrelated genes tested with this system: Igl, the intermediate subunit of the galactose- and N-acetyl-D-galactosamine-inhibitable lectin; the transcription factor URE3-BP; and the membrane binding protein EhC2A. Igl levels were reduced by 72%, URE3-BP by 89%, and EhC2A by 97%. Use of the U6 promoter to drive expression of 29-basepair short hairpin RNAs is effective at knocking down protein expression for unrelated genes in Entamoeba histolytica, providing a useful tool for the study of this parasite.

  2. Structural, Dynamical, and Energetical Consequences of Rett Syndrome Mutation R133C in MeCP2

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    Tugba G. Kucukkal

    2015-01-01

    Full Text Available Rett Syndrome (RTT is a progressive neurodevelopmental disease affecting females. RTT is caused by mutations in the MECP2 gene and various amino acid substitutions have been identified clinically in different domains of the multifunctional MeCP2 protein encoded by this gene. The R133C variant in the methylated-CpG-binding domain (MBD of MeCP2 is the second most common disease-causing mutation in the MBD. Comparative molecular dynamics simulations of R133C mutant and wild-type MBD have been performed to understand the impact of the mutation on structure, dynamics, and interactions of the protein and subsequently understand the disease mechanism. Two salt bridges within the protein and two critical hydrogen bonds between the protein and DNA are lost upon the R133C mutation. The mutation was found to weaken the interaction with DNA and also cause loss of helicity within the 141-144 region. The structural, dynamical, and energetical consequences of R133C mutation were investigated in detail at the atomic resolution. Several important implications of this have been shown regarding protein stability and hydration dynamics as well as its interaction with DNA. The results are in agreement with previous experimental studies and further provide atomic level understanding of the molecular origin of RTT associated with R133C variant.

  3. Wilson's disease in Southern Brazil: genotype-phenotype correlation and description of two novel mutations in ATP7B gene

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    Ricardo Schmitt de Bem

    2013-08-01

    Full Text Available OBJECTIVE: Wilson's disease (WD is an inborn error of metabolism caused by abnormalities of the copper-transporting protein encoding gene ATP7B. In this study, we examined ATP7B for mutations in a group of patients living in southern Brazil. METHODS: 36 WD subjects were studied and classified according to their clinical and epidemiological data. In 23 subjects the ATP7B gene was analyzed. RESULTS: Fourteen distinct mutations were detected in at least one of the alleles. The c.3207C>A substitution at exon 14 was the most common mutation (allelic frequency=37.1% followed by the c.3402delC at exon 15 (allelic frequency=11.4%. The mutations c.2018-2030del13 at exon 7 and c.4093InsT at exon 20 are being reported for the first time. CONCLUSION: The c.3207C>A substitution at exon 14, was the most common mutation, with an allelic frequency of 37.1%. This mutation is the most common mutation described in Europe.

  4. A simple RT-PCR-based strategy for screening connexin identity

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    M. Urban

    1999-08-01

    Full Text Available Vertebrate gap junctions are aggregates of transmembrane channels which are composed of connexin (Cx proteins encoded by at least fourteen distinct genes in mammals. Since the same Cx type can be expressed in different tissues and more than one Cx type can be expressed by the same cell, the thorough identification of which connexin is in which cell type and how connexin expression changes after experimental manipulation has become quite laborious. Here we describe an efficient, rapid and simple method by which connexin type(s can be identified in mammalian tissue and cultured cells using endonuclease cleavage of RT-PCR products generated from "multi primers" (sense primer, degenerate oligonucleotide corresponding to a region of the first extracellular domain; antisense primer, degenerate oligonucleotide complementary to the second extracellular domain that amplify the cytoplasmic loop regions of all known connexins except Cx36. In addition, we provide sequence information on RT-PCR primers used in our laboratory to screen individual connexins and predictions of extension of the "multi primer" method to several human connexins.

  5. Isolation and characterization of φkm18p, a novel lytic phage with therapeutic potential against extensively drug resistant Acinetobacter baumannii.

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    Gwan-Han Shen

    Full Text Available AIMS: To isolate phages against extensively drug resistant Acinetobacter baumannii (XDRAB and characterize the highest lytic capability phage as a model to evaluate the potential on phage therapy. METHODS AND RESULTS: Eight phages were isolated from hospital sewage and showed narrow host spectrum. Phage φkm18p was able to effectively lyse the most XDRAB. It has a dsDNA genome of 45 kb in size and hexagonal head of about 59 nm in diameter and no tail. Bacterial population decreased quickly from 10(8 CFU ml(-1 to 10(3 CFU ml(-1 in 30 min by φkm18p. The 185 kDa lysis protein encoded by φkm18p genome was detected when the extracted protein did not boil before SDS-PAGE; it showed that the lysis protein is a complex rather than a monomer. Phage φkm18p improved human lung epithelial cells survival rates when they were incubated with A. baumannii. Combination of phages (φkm18p, φTZ1 and φ314 as a cocktail could lyse all genotype-varying XDRAB isolates. CONCLUSION: Infections with XDRAB are extremely difficult to treat and development of a phage cocktails therapy could be a therapeutic alternative in the future. Phage φkm18p is a good candidate for inclusion in phage cocktails.

  6. Differential quantitative proteomics of Porphyromonas gingivalis by linear ion trap mass spectrometry: Non-label methods comparison, q-values and LOWESS curve fitting

    Science.gov (United States)

    Xia, Qiangwei; Wang, Tiansong; Park, Yoonsuk; Lamont, Richard J.; Hackett, Murray

    2007-01-01

    Differential analysis of whole cell proteomes by mass spectrometry has largely been applied using various forms of stable isotope labeling. While metabolic stable isotope labeling has been the method of choice, it is often not possible to apply such an approach. Four different label free ways of calculating expression ratios in a classic "two-state" experiment are compared: signal intensity at the peptide level, signal intensity at the protein level, spectral counting at the peptide level, and spectral counting at the protein level. The quantitative data were mined from a dataset of 1245 qualitatively identified proteins, about 56% of the protein encoding open reading frames from Porphyromonas gingivalis, a Gram-negative intracellular pathogen being studied under extracellular and intracellular conditions. Two different control populations were compared against P. gingivalis internalized within a model human target cell line. The q-value statistic, a measure of false discovery rate previously applied to transcription microarrays, was applied to proteomics data. For spectral counting, the most logically consistent estimate of random error came from applying the locally weighted scatter plot smoothing procedure (LOWESS) to the most extreme ratios generated from a control technical replicate, thus setting upper and lower bounds for the region of experimentally observed random error.

  7. Culture medium, gas atmosphere and MAPK inhibition affect regulation of RNA-binding protein targets during mouse preimplantation development.

    Science.gov (United States)

    Calder, Michele D; Watson, Patricia H; Watson, Andrew J

    2011-11-01

    During oogenesis, mammalian oocytes accumulate maternal mRNAs that support the embryo until embryonic genome activation. RNA-binding proteins (RBP) may regulate the stability and turnover of maternal and embryonic mRNAs. We hypothesised that varying embryo culture conditions, such as culture medium, oxygen tension and MAPK inhibition, affects regulation of RBPs and their targets during preimplantation development. STAU1, ELAVL1, KHSRP and ZFP36 proteins and mRNAs were detected throughout mouse preimplantation development, whereas Elavl2 mRNA decreased after the two-cell stage. Potential target mRNAs of RBP regulation, Gclc, Slc2a1 and Slc7a1 were detected during mouse preimplantation development. Gclc mRNA was significantly elevated in embryos cultured in Whitten's medium compared with embryos cultured in KSOMaa, and Gclc mRNA was elevated under high-oxygen conditions. Inhibition of the p38 MAPK pathway reduced Slc7a1 mRNA expression while inhibition of ERK increased Slc2a1 mRNA expression. The half-lives of the potential RBP mRNA targets are not regulated in parallel; Slc2a1 mRNA displayed the longest half-life. Our results indicate that mRNAs and proteins encoding five RBPs are present during preimplantation development and more importantly, demonstrate that expression of RBP target mRNAs are regulated by culture medium, gas atmosphere and MAPK pathways.

  8. Bridging two scholarly islands enriches both: COI DNA barcodes for species identification versus human mitochondrial variation for the study of migrations and pathologies.

    Science.gov (United States)

    Thaler, David S; Stoeckle, Mark Y

    2016-10-01

    DNA barcodes for species identification and the analysis of human mitochondrial variation have developed as independent fields even though both are based on sequences from animal mitochondria. This study finds questions within each field that can be addressed by reference to the other. DNA barcodes are based on a 648-bp segment of the mitochondrially encoded cytochrome oxidase I. From most species, this segment is the only sequence available. It is impossible to know whether it fairly represents overall mitochondrial variation. For modern humans, the entire mitochondrial genome is available from thousands of healthy individuals. SNPs in the human mitochondrial genome are evenly distributed across all protein-encoding regions arguing that COI DNA barcode is representative. Barcode variation among related species is largely based on synonymous codons. Data on human mitochondrial variation support the interpretation that most - possibly all - synonymous substitutions in mitochondria are selectively neutral. DNA barcodes confirm reports of a low variance in modern humans compared to nonhuman primates. In addition, DNA barcodes allow the comparison of modern human variance to many other extant animal species. Birds are a well-curated group in which DNA barcodes are coupled with census and geographic data. Putting modern human variation in the context of intraspecies variation among birds shows humans to be a single breeding population of average variance.

  9. Ecological niche modelling and nDNA sequencing support a new, morphologically cryptic beetle species unveiled by DNA barcoding.

    Science.gov (United States)

    Hawlitschek, Oliver; Porch, Nick; Hendrich, Lars; Balke, Michael

    2011-02-09

    DNA sequencing techniques used to estimate biodiversity, such as DNA barcoding, may reveal cryptic species. However, disagreements between barcoding and morphological data have already led to controversy. Species delimitation should therefore not be based on mtDNA alone. Here, we explore the use of nDNA and bioclimatic modelling in a new species of aquatic beetle revealed by mtDNA sequence data. The aquatic beetle fauna of Australia is characterised by high degrees of endemism, including local radiations such as the genus Antiporus. Antiporus femoralis was previously considered to exist in two disjunct, but morphologically indistinguishable populations in south-western and south-eastern Australia. We constructed a phylogeny of Antiporus and detected a deep split between these populations. Diagnostic characters from the highly variable nuclear protein encoding arginine kinase gene confirmed the presence of two isolated populations. We then used ecological niche modelling to examine the climatic niche characteristics of the two populations. All results support the status of the two populations as distinct species. We describe the south-western species as Antiporus occidentalis sp.n. In addition to nDNA sequence data and extended use of mitochondrial sequences, ecological niche modelling has great potential for delineating morphologically cryptic species.

  10. Transverse relaxation dispersion of the p7 membrane channel from hepatitis C virus reveals conformational breathing

    International Nuclear Information System (INIS)

    Dev, Jyoti; Brüschweiler, Sven; Ouyang, Bo; Chou, James J.

    2015-01-01

    The p7 membrane protein encoded by hepatitis C virus (HCV) assembles into a homo-hexamer that selectively conducts cations. An earlier solution NMR structure of the hexameric complex revealed a funnel-like architecture and suggests that a ring of conserved asparagines near the narrow end of the funnel are important for cation interaction. NMR based drug-binding experiments also suggest that rimantadine can allosterically inhibit ion conduction via a molecular wedge mechanism. These results suggest the presence of dilation and contraction of the funnel tip that are important for channel activity and that the action of the drug is attenuating this motion. Here, we determined the conformational dynamics and solvent accessibility of the p7 channel. The proton exchange measurements show that the cavity-lining residues are largely water accessible, consistent with the overall funnel shape of the channel. Our relaxation dispersion data show that residues Val7 and Leu8 near the asparagine ring are subject to large chemical exchange, suggesting significant intrinsic channel breathing at the tip of the funnel. Moreover, the hinge regions connecting the narrow and wide regions of the funnel show strong relaxation dispersion and these regions are the binding sites for rimantadine. Presence of rimantadine decreases the conformational dynamics near the asparagine ring and the hinge area. Our data provide direct observation of μs–ms dynamics of the p7 channel and support the molecular wedge mechanism of rimantadine inhibition of the HCV p7 channel

  11. Dissecting the calcium-induced differentiation of human primary keratinocytes stem cells by integrative and structural network analyses.

    Directory of Open Access Journals (Sweden)

    Kiana Toufighi

    2015-05-01

    Full Text Available The molecular details underlying the time-dependent assembly of protein complexes in cellular networks, such as those that occur during differentiation, are largely unexplored. Focusing on the calcium-induced differentiation of primary human keratinocytes as a model system for a major cellular reorganization process, we look at the expression of genes whose products are involved in manually-annotated protein complexes. Clustering analyses revealed only moderate co-expression of functionally related proteins during differentiation. However, when we looked at protein complexes, we found that the majority (55% are composed of non-dynamic and dynamic gene products ('di-chromatic', 19% are non-dynamic, and 26% only dynamic. Considering three-dimensional protein structures to predict steric interactions, we found that proteins encoded by dynamic genes frequently interact with a common non-dynamic protein in a mutually exclusive fashion. This suggests that during differentiation, complex assemblies may also change through variation in the abundance of proteins that compete for binding to common proteins as found in some cases for paralogous proteins. Considering the example of the TNF-α/NFκB signaling complex, we suggest that the same core complex can guide signals into diverse context-specific outputs by addition of time specific expressed subunits, while keeping other cellular functions constant. Thus, our analysis provides evidence that complex assembly with stable core components and competition could contribute to cell differentiation.

  12. Mitochondrial introgression suggests extensive ancestral hybridization events among Saccharomyces species.

    Science.gov (United States)

    Peris, David; Arias, Armando; Orlić, Sandi; Belloch, Carmela; Pérez-Través, Laura; Querol, Amparo; Barrio, Eladio

    2017-03-01

    Horizontal gene transfer (HGT) in eukaryotic plastids and mitochondrial genomes is common, and plays an important role in organism evolution. In yeasts, recent mitochondrial HGT has been suggested between S. cerevisiae and S. paradoxus. However, few strains have been explored given the lack of accurate mitochondrial genome annotations. Mitochondrial genome sequences are important to understand how frequent these introgressions occur, and their role in cytonuclear incompatibilities and fitness. Indeed, most of the Bateson-Dobzhansky-Muller genetic incompatibilities described in yeasts are driven by cytonuclear incompatibilities. We herein explored the mitochondrial inheritance of several worldwide distributed wild Saccharomyces species and their hybrids isolated from different sources and geographic origins. We demonstrated the existence of several recombination points in mitochondrial region COX2-ORF1, likely mediated by either the activity of the protein encoded by the ORF1 (F-SceIII) gene, a free-standing homing endonuclease, or mostly facilitated by A+T tandem repeats and regions of integration of GC clusters. These introgressions were shown to occur among strains of the same species and among strains of different species, which suggests a complex model of Saccharomyces evolution that involves several ancestral hybridization events in wild environments. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Identification of a Chlorophyll Dephytylase Involved in Chlorophyll Turnover in Arabidopsis[OPEN

    Science.gov (United States)

    2016-01-01

    Chlorophyll turns over in green organs during photosystem repair and is salvaged via de- and rephytylation, but the enzyme involved in dephytylation is unknown. We have identified an Arabidopsis thaliana thylakoid protein with a putative hydrolase domain that can dephytylate chlorophyll in vitro and in vivo. The corresponding locus, CHLOROPHYLL DEPHYTYLASE1 (CLD1), was identified by mapping a semidominant, heat-sensitive, missense allele (cld1-1). CLD1 is conserved in oxygenic photosynthetic organisms, sharing structural similarity with pheophytinase, which functions in chlorophyll breakdown during leaf senescence. Unlike pheophytinase, CLD1 is predominantly expressed in green organs and can dephytylate chlorophyll in vitro. The specific activity is significantly higher for the mutant protein encoded by cld1-1 than the wild-type enzyme, consistent with the semidominant nature of the cld1-1 mutation. Supraoptimal CLD1 activities in cld1-1 mutants and transgenic seedlings led to the proportional accumulation of chlorophyllides derived from chlorophyll dephytylation after heat shock, which resulted in light-dependent cotyledon bleaching. Reducing CLD1 expression diminished thermotolerance and the photochemical efficiency of photosystem II under prolonged moderate heat stress. Taken together, our results suggest that CLD1 is the long-sought enzyme for removing the phytol chain from chlorophyll during its turnover at steady state within the chloroplast. PMID:27920339

  14. Identification of a Chlorophyll Dephytylase Involved in Chlorophyll Turnover in Arabidopsis.

    Science.gov (United States)

    Lin, Yao-Pin; Wu, Meng-Chen; Charng, Yee-Yung

    2016-12-01

    Chlorophyll turns over in green organs during photosystem repair and is salvaged via de- and rephytylation, but the enzyme involved in dephytylation is unknown. We have identified an Arabidopsis thaliana thylakoid protein with a putative hydrolase domain that can dephytylate chlorophyll in vitro and in vivo. The corresponding locus, CHLOROPHYLL DEPHYTYLASE1 (CLD1), was identified by mapping a semidominant, heat-sensitive, missense allele (cld1-1). CLD1 is conserved in oxygenic photosynthetic organisms, sharing structural similarity with pheophytinase, which functions in chlorophyll breakdown during leaf senescence. Unlike pheophytinase, CLD1 is predominantly expressed in green organs and can dephytylate chlorophyll in vitro. The specific activity is significantly higher for the mutant protein encoded by cld1-1 than the wild-type enzyme, consistent with the semidominant nature of the cld1-1 mutation. Supraoptimal CLD1 activities in cld1-1 mutants and transgenic seedlings led to the proportional accumulation of chlorophyllides derived from chlorophyll dephytylation after heat shock, which resulted in light-dependent cotyledon bleaching. Reducing CLD1 expression diminished thermotolerance and the photochemical efficiency of photosystem II under prolonged moderate heat stress. Taken together, our results suggest that CLD1 is the long-sought enzyme for removing the phytol chain from chlorophyll during its turnover at steady state within the chloroplast. © 2016 American Society of Plant Biologists. All rights reserved.

  15. Genetic and Hormonal Regulation of Chlorophyll Degradation during Maturation of Seeds with Green Embryos.

    Science.gov (United States)

    Smolikova, Galina; Dolgikh, Elena; Vikhnina, Maria; Frolov, Andrej; Medvedev, Sergei

    2017-09-16

    The embryos of some angiosperms (usually referred to as chloroembryos) contain chlorophylls during the whole period of embryogenesis. Developing embryos have photochemically active chloroplasts and are able to produce assimilates, further converted in reserve biopolymers, whereas at the late steps of embryogenesis, seeds undergo dehydration, degradation of chlorophylls, transformation of chloroplast in storage plastids, and enter the dormancy period. However, in some seeds, the process of chlorophyll degradation remains incomplete. These residual chlorophylls compromise the quality of seed material in terms of viability, nutritional value, and shelf life, and represent a serious challenge for breeders and farmers. The mechanisms of chlorophyll degradation during seed maturation are still not completely understood, and only during the recent decades the main pathways and corresponding enzymes could be characterized. Among the identified players, the enzymes of pheophorbide a oxygenase pathway and the proteins encoded by STAY GREEN ( SGR ) genes are the principle ones. On the biochemical level, abscisic acid (ABA) is the main regulator of seed chlorophyll degradation, mediating activity of corresponding catabolic enzymes on the transcriptional level. In general, a deep insight in the mechanisms of chlorophyll degradation is required to develop the approaches for production of chlorophyll-free high quality seeds.

  16. Role of the gerA operon in L-alanine germination of Bacillus licheniformis spores

    Directory of Open Access Journals (Sweden)

    Løvdal Irene S

    2012-03-01

    Full Text Available Abstract Background The genome of Bacillus licheniformis DSM 13 harbours three neighbouring open reading frames showing protein sequence similarities to the proteins encoded from the Bacillus subtilis subsp. subtilis 168 gerA operon, GerAA, GerAB and GerAC. In B. subtilis, these proteins are assumed to form a germinant receptor involved in spore germination induced by the amino acid L-alanine. Results In this study we show that disruption of the gerAA gene in B. licheniformis MW3 hamper L-alanine and casein hydrolysate-triggered spore germination, measured by absorbance at 600 nm and confirmed by phase contrast microscopy. This ability was restored by complementation with a plasmid-borne copy of the gerA locus. Addition of D-alanine in the casein hydrolysate germination assay abolished germination of both B. licheniformis MW3 and the complementation mutant. Germination of both B. licheniformis MW3 and the gerA disruption mutant was induced by the non-nutrient germinant Ca2+-Dipicolinic acid. Conclusions These results demonstrate that the B. licheniformis MW3 gerA locus is involved in germination induced by L-alanine and potentially other components present in casein hydrolysate.

  17. Role of the gerA operon in L-alanine germination of Bacillus licheniformis spores

    Science.gov (United States)

    2012-01-01

    Background The genome of Bacillus licheniformis DSM 13 harbours three neighbouring open reading frames showing protein sequence similarities to the proteins encoded from the Bacillus subtilis subsp. subtilis 168 gerA operon, GerAA, GerAB and GerAC. In B. subtilis, these proteins are assumed to form a germinant receptor involved in spore germination induced by the amino acid L-alanine. Results In this study we show that disruption of the gerAA gene in B. licheniformis MW3 hamper L-alanine and casein hydrolysate-triggered spore germination, measured by absorbance at 600 nm and confirmed by phase contrast microscopy. This ability was restored by complementation with a plasmid-borne copy of the gerA locus. Addition of D-alanine in the casein hydrolysate germination assay abolished germination of both B. licheniformis MW3 and the complementation mutant. Germination of both B. licheniformis MW3 and the gerA disruption mutant was induced by the non-nutrient germinant Ca2+-Dipicolinic acid. Conclusions These results demonstrate that the B. licheniformis MW3 gerA locus is involved in germination induced by L-alanine and potentially other components present in casein hydrolysate. PMID:22420404

  18. Involvement of C4 protein of beet severe curly top virus (family Geminiviridae in virus movement.

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    Kunling Teng

    Full Text Available BACKGROUND: Beet severe curly top virus (BSCTV is a leafhopper transmitted geminivirus with a monopartite genome. C4 proteins encoded by geminivirus play an important role in virus/plant interaction. METHODS AND FINDINGS: To understand the function of C4 encoded by BSCTV, two BSCTV mutants were constructed by introducing termination codons in ORF C4 without affecting the amino acids encoded by overlapping ORF Rep. BSCTV mutants containing disrupted ORF C4 retained the ability to replicate in Arabidopsis protoplasts and in the agro-inoculated leaf discs of N. benthamiana, suggesting C4 is not required for virus DNA replication. However, both mutants did not accumulate viral DNA in newly emerged leaves of inoculated N. benthamiana and Arabidopsis, and the inoculated plants were asymptomatic. We also showed that C4 expression in plant could help C4 deficient BSCTV mutants to move systemically. C4 was localized in the cytosol and the nucleus in both Arabidopsis protoplasts and N. benthamiana leaves and the protein appeared to bind viral DNA and ds/ssDNA nonspecifically, displaying novel DNA binding properties. CONCLUSIONS: Our results suggest that C4 protein in BSCTV is involved in symptom production and may facilitate virus movement instead of virus replication.

  19. Cell-Free Expression and In Situ Immobilization of Parasite Proteins from Clonorchis sinensis for Rapid Identification of Antigenic Candidates.

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    Christy Catherine

    Full Text Available Progress towards genetic sequencing of human parasites has provided the groundwork for a post-genomic approach to develop novel antigens for the diagnosis and treatment of parasite infections. To fully utilize the genomic data, however, high-throughput methodologies are required for functional analysis of the proteins encoded in the genomic sequences. In this study, we investigated cell-free expression and in situ immobilization of parasite proteins as a novel platform for the discovery of antigenic proteins. PCR-amplified parasite DNA was immobilized on microbeads that were also functionalized to capture synthesized proteins. When the microbeads were incubated in a reaction mixture for cell-free synthesis, proteins expressed from the microbead-immobilized DNA were instantly immobilized on the same microbeads, providing a physical linkage between the genetic information and encoded proteins. This approach of in situ expression and isolation enables streamlined recovery and analysis of cell-free synthesized proteins and also allows facile identification of the genes coding antigenic proteins through direct PCR of the microbead-bound DNA.

  20. Cell-free translational screening of an expression sequence tag library of Clonorchis sinensis for novel antigen discovery.

    Science.gov (United States)

    Kasi, Devi; Catherine, Christy; Lee, Seung-Won; Lee, Kyung-Ho; Kim, Yu Jung; Ro Lee, Myeong; Ju, Jung Won; Kim, Dong-Myung

    2017-05-01

    The rapidly evolving cloning and sequencing technologies have enabled understanding of genomic structure of parasite genomes, opening up new ways of combatting parasite-related diseases. To make the most of the exponentially accumulating genomic data, however, it is crucial to analyze the proteins encoded by these genomic sequences. In this study, we adopted an engineered cell-free protein synthesis system for large-scale expression screening of an expression sequence tag (EST) library of Clonorchis sinensis to identify potential antigens that can be used for diagnosis and treatment of clonorchiasis. To allow high-throughput expression and identification of individual genes comprising the library, a cell-free synthesis reaction was designed such that both the template DNA and the expressed proteins were co-immobilized on the same microbeads, leading to microbead-based linkage of the genotype and phenotype. This reaction configuration allowed streamlined expression, recovery, and analysis of proteins. This approach enabled us to identify 21 antigenic proteins. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:832-837, 2017. © 2017 American Institute of Chemical Engineers.

  1. Identification of mud crab reovirus VP12 and its interaction with the voltage-dependent anion-selective channel protein of mud crab Scylla paramamosain.

    Science.gov (United States)

    Xu, Hai-Dong; Su, Hong-Jun; Zou, Wei-Bin; Liu, Shan-Shan; Yan, Wen-Rui; Wang, Qian-Qian; Yuan, Li-Li; Chan, Siuming Francis; Yu, Xiao-Qiang; He, Jian-Guo; Weng, Shao-Ping

    2015-05-01

    Mud crab reovirus (MCRV) is the causative agent of a severe disease in cultured mud crab (Scylla paramamosain), which has caused huge economic losses in China. MCRV is a double-stranded RNA virus with 12 genomic segments. In this paper, SDS-PAGE, mass spectrometry and Western blot analyses revealed that the VP12 protein encoded by S12 gene is a structural protein of MCRV. Immune electron microscopy assay indicated that MCRV VP12 is a component of MCRV outer shell capsid. Yeast two hybrid cDNA library of mud crab was constructed and mud crab voltage-dependent anion-selective channel (mcVDAC) was obtained by MCRV VP12 screening. The full length of mcVDAC was 1180 bp with an open reading frame (ORF) of 849 bp encoding a 282 amino acid protein. The mcVDAC had a constitutive expression pattern in different tissues of mud crab. The interaction between MCRV VP12 and mcVDAC was determined by co-immunoprecipitation assay. The results of this study have provided an insight on the mechanisms of MCRV infection and the interactions between the virus and mud crab. Copyright © 2015 Elsevier Ltd. All rights reserved.

  2. The NB-LRR gene Pm60 confers powdery mildew resistance in wheat.

    Science.gov (United States)

    Zou, Shenghao; Wang, Huan; Li, Yiwen; Kong, Zhaosheng; Tang, Dingzhong

    2018-04-01

    Powdery mildew is one of the most devastating diseases of wheat. To date, few powdery mildew resistance genes have been cloned from wheat due to the size and complexity of the wheat genome. Triticum urartu is the progenitor of the A genome of wheat and is an important source for powdery mildew resistance genes. Using molecular markers designed from scaffolds of the sequenced T. urartu accession and standard map-based cloning, a powdery mildew resistance locus was mapped to a 356-kb region, which contains two nucleotide-binding and leucine-rich repeat domain (NB-LRR) protein-encoding genes. Virus-induced gene silencing, single-cell transient expression, and stable transformation assays demonstrated that one of these two genes, designated Pm60, confers resistance to powdery mildew. Overexpression of full-length Pm60 and two allelic variants in Nicotiana benthamiana leaves induced hypersensitive cell death response, but expression of the coiled-coil domain alone was insufficient to induce hypersensitive response. Yeast two-hybrid, bimolecular fluorescence complementation and luciferase complementation imaging assays showed that Pm60 protein interacts with its neighboring NB-containing protein, suggesting that they might be functionally related. The identification and cloning of this novel wheat powdery mildew resistance gene will facilitate breeding for disease resistance in wheat. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  3. Diagnostic Challenges of Female Genital Tuberculosis

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    Chaudhary Vigi

    2017-02-01

    Full Text Available Background: India accounts for one fifth of the global incidence of tuberculosis (TB annually. Genital tract TB is one of the extra pulmonary presentations of TB leading to infertility among Indian women. Genital TB is a chronic disease and often asymptomatic with very few specific complaints. Infertility is the most common clinical presentation of genital TB. Herein, we report a case of 32-year-old female patient suffering from abdominal pain and infertility for the last 8 months. Methods: Hysterosalpingography (HSG and ultrasonography (USG did not reveal characteristic radiological appearances of TB although USG detected the presence of a large fibroid in the right uterine wall. Histology, microscopy for acid fast bacilli, liquid culture and nucleic acid amplification assay targeting 64kDa protein encoding gene, the IS6110 element of endometrium biopsy were negative for tubercle bacilli. Results: Since the diagnosis of genital TB is elusive, antitubercular treatment (ATT using isoniazid, pyrazinamide, rifampicin, and ethambutol was prescribed for two months followed by maintenance therapy with isoniazid and rifampicin for four months without any pregnancy outcome. Conclusion: However, the patient conceived spontaneously after surgical removal of fibroid. Relating infertility to female genital tuberculosis due to high prevalence of TB in the country and ignoring the presence of uterine fibroid might not have been the right decision taken by the gynaecologist. This suggests the urgent need for an accurate method intended for diagnosis of female genital tuberculosis.

  4. A homozygous mutation in the endothelin-3 gene associated with a combined Waardenburg type 2 and Hirschsprung phenotype (Shah-Waardenburg syndrome).

    Science.gov (United States)

    Hofstra, R M; Osinga, J; Tan-Sindhunata, G; Wu, Y; Kamsteeg, E J; Stulp, R P; van Ravenswaaij-Arts, C; Majoor-Krakauer, D; Angrist, M; Chakravarti, A; Meijers, C; Buys, C H

    1996-04-01

    Hirschsprung disease (HSCR) or colonic aganglionosis is a congenital disorder characterized by an absence of intramural ganglia along variable lengths of the colon resulting in intestinal obstruction. The incidence of HSCR is 1 in 5,000 live births. Mutations in the RET gene, which codes for a receptor tyrosine kinase, and in EDNRB which codes for the endothelin-B receptor, have been shown to be associated with HSCR in humans. The lethal-spotted mouse which has pigment abnormalities, but also colonic aganglionosis, carries a mutation in the gene coding for endothelin 3 (Edn3), the ligand for the receptor protein encoded by EDNRB. Here, we describe a mutation of the human gene for endothelin 3 (EDN3), homozygously present in a patient with a combined Waardenburg syndrome type 2 (WS2) and HSCR phenotype (Shah-Waardenburg syndrome). The mutation, Cys159Phe, in exon 3 in the ET-3 like domain of EDN3, presumably affects the proteolytic processing of the preproendothelin to the mature peptide EDN3. The patient's parents were first cousins. A previous child in this family had been diagnosed with a similar combination of HSCR, depigmentation and deafness. Depigmentation and deafness were present in other relatives. Moreover, we present a further indication for the involvement of EDNRB in HSCR by reporting a novel mutation detected in one of 40 unselected HSCR patients.

  5. Novel and recurrent MYO7A mutations in Usher syndrome type 1 and type 2.

    Science.gov (United States)

    Rong, Weining; Chen, Xue; Zhao, Kanxing; Liu, Yani; Liu, Xiaoxing; Ha, Shaoping; Liu, Wenzhou; Kang, Xiaoli; Sheng, Xunlun; Zhao, Chen

    2014-01-01

    Usher syndrome (USH) is a group of disorders manifested as retinitis pigmentosa and bilateral sensorineural hearing loss, with or without vestibular dysfunction. Here, we recruited three Chinese families affected with autosomal recessive USH for detailed clinical evaluations and for mutation screening in the genes associated with inherited retinal diseases. Using targeted next-generation sequencing (NGS) approach, three new alleles and one known mutation in MYO7A gene were identified in the three families. In two families with USH type 1, novel homozygous frameshift variant p.Pro194Hisfs*13 and recurrent missense variant p.Thr165Met were demonstrated as the causative mutations respectively. Crystal structural analysis denoted that p.Thr165Met would very likely change the tertiary structure of the protein encoded by MYO7A. In another family affected with USH type 2, novel biallelic mutations in MYO7A, c.[1343+1G>A];[2837T>G] or p.[?];[Met946Arg], were identified with clinical significance. Because MYO7A, to our knowledge, has rarely been correlated with USH type 2, our findings therefore reveal distinguished clinical phenotypes associated with MYO7A. We also conclude that targeted NGS is an effective approach for genetic diagnosis for USH, which can further provide better understanding of genotype-phenotype relationship of the disease.

  6. Clarin-1, encoded by the Usher Syndrome III causative gene, forms a membranous microdomain: possible role of clarin-1 in organizing the actin cytoskeleton.

    Science.gov (United States)

    Tian, Guilian; Zhou, Yun; Hajkova, Dagmar; Miyagi, Masaru; Dinculescu, Astra; Hauswirth, William W; Palczewski, Krzysztof; Geng, Ruishuang; Alagramam, Kumar N; Isosomppi, Juha; Sankila, Eeva-Marja; Flannery, John G; Imanishi, Yoshikazu

    2009-07-10

    Clarin-1 is the protein product encoded by the gene mutated in Usher syndrome III. Although the molecular function of clarin-1 is unknown, its primary structure predicts four transmembrane domains similar to a large family of membrane proteins that include tetraspanins. Here we investigated the role of clarin-1 by using heterologous expression and in vivo model systems. When expressed in HEK293 cells, clarin-1 localized to the plasma membrane and concentrated in low density compartments distinct from lipid rafts. Clarin-1 reorganized actin filament structures and induced lamellipodia. This actin-reorganizing function was absent in the modified protein encoded by the most prevalent North American Usher syndrome III mutation, the N48K form of clarin-1 deficient in N-linked glycosylation. Proteomics analyses revealed a number of clarin-1-interacting proteins involved in cell-cell adhesion, focal adhesions, cell migration, tight junctions, and regulation of the actin cytoskeleton. Consistent with the hypothesized role of clarin-1 in actin organization, F-actin-enriched stereocilia of auditory hair cells evidenced structural disorganization in Clrn1(-/-) mice. These observations suggest a possible role for clarin-1 in the regulation and homeostasis of actin filaments, and link clarin-1 to the interactive network of Usher syndrome gene products.

  7. The Drosophila melanogaster DmCK2beta transcription unit encodes for functionally non-redundant protein isoforms.

    Science.gov (United States)

    Jauch, Eike; Wecklein, Heike; Stark, Felix; Jauch, Mandy; Raabe, Thomas

    2006-06-07

    Genes encoding for the two evolutionary highly conserved subunits of a heterotetrameric protein kinase CK2 holoenzyme are present in all examined eukaryotic genomes. Depending on the organism, multiple transcription units encoding for a catalytically active CK2alpha subunit and/or a regulatory CK2beta subunit may exist. The phosphotransferase activity of members of the protein kinase CK2alpha family is thought to be independent of second messengers but is modulated by interaction with CK2beta-like proteins. In the genome of Drosophila melanogaster, one gene encoding for a CK2alpha subunit and three genes encoding for CK2beta-like proteins are present. The X-linked DmCK2beta transcription unit encodes for several CK2beta protein isoforms due to alternative splicing of its primary transcript. We addressed the question whether CK2beta-like proteins are redundant in function. Our in vivo experiments show that variations of the very C-terminal tail of CK2beta isoforms encoded by the X-linked DmCK2beta transcription unit influence their functional properties. In addition, we find that CK2beta-like proteins encoded by the autosomal D. melanogaster genes CK2betates and CK2beta' cannot fully substitute for a loss of CK2beta isoforms encoded by DmCK2beta.

  8. A Fungal Effector With Host Nuclear Localization and DNA-Binding Properties Is Required for Maize Anthracnose Development.

    Science.gov (United States)

    Vargas, Walter A; Sanz-Martín, José M; Rech, Gabriel E; Armijos-Jaramillo, Vinicio D; Rivera, Lina P; Echeverria, María Mercedes; Díaz-Mínguez, José M; Thon, Michael R; Sukno, Serenella A

    2016-02-01

    Plant pathogens have the capacity to manipulate the host immune system through the secretion of effectors. We identified 27 putative effector proteins encoded in the genome of the maize anthracnose pathogen Colletotrichum graminicola that are likely to target the host's nucleus, as they simultaneously contain sequence signatures for secretion and nuclear localization. We functionally characterized one protein, identified as CgEP1. This protein is synthesized during the early stages of disease development and is necessary for anthracnose development in maize leaves, stems, and roots. Genetic, molecular, and biochemical studies confirmed that this effector targets the host's nucleus and defines a novel class of double-stranded DNA-binding protein. We show that CgEP1 arose from a gene duplication in an ancestor of a lineage of monocot-infecting Colletotrichum spp. and has undergone an intense evolution process, with evidence for episodes of positive selection. We detected CgEP1 homologs in several species of a grass-infecting lineage of Colletotrichum spp., suggesting that its function may be conserved across a large number of anthracnose pathogens. Our results demonstrate that effectors targeted to the host nucleus may be key elements for disease development and aid in the understanding of the genetic basis of anthracnose development in maize plants.

  9. Dicer-like Proteins Regulate the Growth, Conidiation, and Pathogenicity of Colletotrichum gloeosporioides from Hevea brasiliensis

    Directory of Open Access Journals (Sweden)

    Qiannan Wang

    2018-01-01

    Full Text Available Colletotrichum gloeosporioides from Hevea brasiliensis is the hemibiotrophic fungi which could cause anthracnose in rubber trees. Dicer like proteins (DCL were the core enzymes for generation of small RNAs. In the present study, the knocking-out mutants of two dicer like proteins encoding genes of C. gloeosporioides were constructed; and functions of two proteins were investigated. The results showed that DCL play important roles in regulating the growth, conidiation and pathogenicity of C. gloeosporioides; and there is a functional redundancy between DCL1 and DCL2. Microscopy analysis and DAB staining revealed that loss of penetration ability into the host cells, instead of the decreased growth rate, was the main cause for the impaired pathogenicity of the ΔDcl1ΔDcl2 double mutant. Proteomics analysis suggested that DCL proteins affected the expression of functional proteins to regulating multiple biological processes of C. gloeosporioides. These data lead to a better understanding of the functions of DCL proteins in regulating the development and pathogenesis of C. gloeosporioides.

  10. Down-regulation of a calmodulin-related gene during transformation of human mammary epithelial cells

    International Nuclear Information System (INIS)

    Yaswen, P.; Smoll, A.; Stampfer, M.R.; Peehl, D.M.; Trask, D.K.; Sager, R.

    1990-01-01

    A human cDNA library obtained from cultured normal mammary epithelial cells (HMECs) was searched by subtractive hybridization for genes whose decrease in expression might be relevant to epithelial transformation. One clone identified by this procedure corresponded to a 1.4 kilobase mRNA, designated NB-1, whose expression was decreased >50-fold in HMECs tumorigenically transformed in vitro after exposure to benzo[α]pyrene and Kirsten sarcoma virus. Sequence analysis of NB-1 cDNA revealed an open reading frame with a high degree of homology to calmodulin. NB-1 expression could be demonstrated by polymerase chain reaction amplification in normal breast, prostate, cervix, and epidermal tissues. The presence of NB-1 transcripts was variable in primary breast carcinoma tissues and undetectable in tumor-derived cell lines of breast, prostate, or other origins. NB-1 mRNA expression could be down-regulated in cultured HMECs by exposure to reconstituted extracellular matrix material, while exposure to transforming growth factor type β increased its relative abundance. The protein encoded by NB-1 may have Ca 2 plus binding properties and perform functions similar to those of authentic calmodulin. Its possible roles in differentiation and/or suppression of tumorigenicity in epithelial tissues remain to be examined

  11. Peroxisome proliferator-activated receptor-γ agonists inhibit the replication of respiratory syncytial virus (RSV) in human lung epithelial cells

    International Nuclear Information System (INIS)

    Arnold, Ralf; Koenig, Wolfgang

    2006-01-01

    We have previously shown that peroxisome proliferator-activated receptor-γ (PPARγ) agonists inhibited the inflammatory response of RSV-infected human lung epithelial cells. In this study, we supply evidence that specific PPARγ agonists (15d-PGJ 2 , ciglitazone, troglitazone, Fmoc-Leu) efficiently blocked the RSV-induced cytotoxicity and development of syncytia in tissue culture (A549, HEp-2). All PPARγ agonists under study markedly inhibited the cell surface expression of the viral G and F protein on RSV-infected A549 cells. This was paralleled by a reduced cellular amount of N protein-encoding mRNA determined by real-time RT-PCR. Concomitantly, a reduced release of infectious progeny virus into the cell supernatants of human lung epithelial cells (A549, normal human bronchial epithelial cells (NHBE)) was observed. Similar results were obtained regardless whether PPARγ agonists were added prior to RSV infection or thereafter, suggesting that the agonists inhibited viral gene expression and not the primary adhesion or fusion process

  12. D1 dopamine receptor signaling is modulated by the R7 RGS protein EAT-16 and the R7 binding protein RSBP-1 in Caenoerhabditis elegans motor neurons.

    Directory of Open Access Journals (Sweden)

    Khursheed A Wani

    Full Text Available Dopamine signaling modulates voluntary movement and reward-driven behaviors by acting through G protein-coupled receptors in striatal neurons, and defects in dopamine signaling underlie Parkinson's disease and drug addiction. Despite the importance of understanding how dopamine modifies the activity of striatal neurons to control basal ganglia output, the molecular mechanisms that control dopamine signaling remain largely unclear. Dopamine signaling also controls locomotion behavior in Caenorhabditis elegans. To better understand how dopamine acts in the brain we performed a large-scale dsRNA interference screen in C. elegans for genes required for endogenous dopamine signaling and identified six genes (eat-16, rsbp-1, unc-43, flp-1, grk-1, and cat-1 required for dopamine-mediated behavior. We then used a combination of mutant analysis and cell-specific transgenic rescue experiments to investigate the functional interaction between the proteins encoded by two of these genes, eat-16 and rsbp-1, within single cell types and to examine their role in the modulation of dopamine receptor signaling. We found that EAT-16 and RSBP-1 act together to modulate dopamine signaling and that while they are coexpressed with both D1-like and D2-like dopamine receptors, they do not modulate D2 receptor signaling. Instead, EAT-16 and RSBP-1 act together to selectively inhibit D1 dopamine receptor signaling in cholinergic motor neurons to modulate locomotion behavior.

  13. Protannotator: a semiautomated pipeline for chromosome-wise functional annotation of the "missing" human proteome.

    Science.gov (United States)

    Islam, Mohammad T; Garg, Gagan; Hancock, William S; Risk, Brian A; Baker, Mark S; Ranganathan, Shoba

    2014-01-03

    The chromosome-centric human proteome project (C-HPP) aims to define the complete set of proteins encoded in each human chromosome. The neXtProt database (September 2013) lists 20,128 proteins for the human proteome, of which 3831 human proteins (∼19%) are considered "missing" according to the standard metrics table (released September 27, 2013). In support of the C-HPP initiative, we have extended the annotation strategy developed for human chromosome 7 "missing" proteins into a semiautomated pipeline to functionally annotate the "missing" human proteome. This pipeline integrates a suite of bioinformatics analysis and annotation software tools to identify homologues and map putative functional signatures, gene ontology, and biochemical pathways. From sequential BLAST searches, we have primarily identified homologues from reviewed nonhuman mammalian proteins with protein evidence for 1271 (33.2%) "missing" proteins, followed by 703 (18.4%) homologues from reviewed nonhuman mammalian proteins and subsequently 564 (14.7%) homologues from reviewed human proteins. Functional annotations for 1945 (50.8%) "missing" proteins were also determined. To accelerate the identification of "missing" proteins from proteomics studies, we generated proteotypic peptides in silico. Matching these proteotypic peptides to ENCODE proteogenomic data resulted in proteomic evidence for 107 (2.8%) of the 3831 "missing proteins, while evidence from a recent membrane proteomic study supported the existence for another 15 "missing" proteins. The chromosome-wise functional annotation of all "missing" proteins is freely available to the scientific community through our web server (http://biolinfo.org/protannotator).

  14. The frequency of genes encoding three putative group B streptococcal virulence factors among invasive and colonizing isolates

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    Borchardt Stephanie M

    2006-07-01

    Full Text Available Abstract Background Group B Streptococcus (GBS causes severe infections in very young infants and invasive disease in pregnant women and adults with underlying medical conditions. GBS pathogenicity varies between and within serotypes, with considerable variation in genetic content between strains. Three proteins, Rib encoded by rib, and alpha and beta C proteins encoded by bca and bac, respectively, have been suggested as potential vaccine candidates for GBS. It is not known, however, whether these genes occur more frequently in invasive versus colonizing GBS strains. Methods We screened 162 invasive and 338 colonizing GBS strains from different collections using dot blot hybridization to assess the frequency of bca, bac and rib. All strains were defined by serotyping for capsular type, and frequency differences were tested using the Chi square test. Results Genes encoding the beta C protein (bac and Rib (rib occurred at similar frequencies among invasive and colonizing isolates, bac (20% vs. 23%, and rib (28% vs. 20%, while the alpha (bca C protein was more frequently found in colonizing strains (46% vs, invasive (29%. Invasive strains were associated with specific serotype/gene combinations. Conclusion Novel virulence factors must be identified to better understand GBS disease.

  15. The Genome of the Epsilonproteobacterial Chemolithoautotroph Sulfurimonas dentrificans

    Energy Technology Data Exchange (ETDEWEB)

    USF Genomics Class; Sievert, Stefan M.; Scott, Kathleen M.; Klotz, Martin G.; Chain, Patrick S.G.; Hauser, Loren J.; Hemp, James; Hugler, Michael; Land, Miriam; Lapidus, Alla; Larimer, Frank W.; Lucas, Susan; Malfatti, Stephanie A.; Meyer, Folker; Paulsen, Ian T.; Ren, Qinghu; Simon, Jorg

    2007-08-08

    Sulfur-oxidizing epsilonproteobacteria are common in a variety of sulfidogenic environments. These autotrophic and mixotrophic sulfur-oxidizing bacteria are believed to contribute substantially to the oxidative portion of the global sulfur cycle. In order to better understand the ecology and roles of sulfur-oxidizing epsilonproteobacteria, in particular those of the widespread genus Sulfurimonas, in biogeochemical cycles, the genome of Sulfurimonas denitrificans DSM1251 was sequenced. This genome has many features, including a larger size (2.2 Mbp), that suggest a greater degree of metabolic versatility or responsiveness to the environment than seen for most of the other sequenced epsilonproteobacteria. A branched electron transport chain is apparent, with genes encoding complexes for the oxidation of hydrogen, reduced sulfur compounds, and formate and the reduction of nitrate and oxygen. Genes are present for a complete, autotrophic reductive citric acid cycle. Many genes are present that could facilitate growth in the spatially and temporally heterogeneous sediment habitat from where Sulfurimonas denitrificans was originally isolated. Many resistance-nodulation-development family transporter genes (10 total) are present; of these, several are predicted to encode heavy metal efflux transporters. An elaborate arsenal of sensory and regulatory protein-encoding genes is in place, as are genes necessary to prevent and respond to oxidative stress.

  16. Gene expression analysis of FABP4 in gastric cancer

    Directory of Open Access Journals (Sweden)

    Abdulkarim Yasin Karim

    2016-06-01

    Full Text Available Purpose: Gastric cancer has high incidence and mortality rate in several countries and is still one of the most frequent and lethal disease. In this study, we aimed to determine diagnostic markers in gastric cancer by molecular techniques; include mRNA expression analysis of FABP4 gene. Fatty acid binding protein 4 (FABP4 gene encodes the fatty acid binding protein found in adipocytes. The protein encoded by FABP4 are a family of small, highly conserved, cytoplasmic proteins that bind long-chain fatty acids and other hydrophobic ligands. It is thought that FABPs roles include fatty acid uptake, transport, and metabolism. Material and Methods: Total RNA were extracted from paired tumor and normal tissues of 47 gastric cancer. The mRNA expression level of FABP4 was measured employing semi- quantitative reverse transcription- polymerase chain reaction (RT- PCR. Results: The mRNA expression level of FABP4 was significantly decreased (down- regulated. Conclusion: Down-regulation of FABP4 gene seems to occur at the initial steps of gastric cancer development. In order to confirm the relationship between the gastric tumor and FABP4 gene, further analysis like immunohistochemistry and epigenetc techniques are necessary. [Cukurova Med J 2016; 41(2.000: 248-252

  17. Cloning and molecular characterization of the glyceraldehyde-3-phosphate dehydrogenase-encoding gene and cDNA from the plant pathogenic fungus Glomerella cingulata.

    Science.gov (United States)

    Templeton, M D; Rikkerink, E H; Solon, S L; Crowhurst, R N

    1992-12-01

    The glyceraldehyde-3-phosphate dehydrogenase gene (gpdA) has been identified from a genomic DNA library prepared from the plant pathogenic fungus Glomerella cingulata. Nucleotide sequence data revealed that this gene codes for a putative 338-amino-acid protein encoded by two exons of 129 and 885 bp, separated by an intron 216 bp long. The 5' leader sequence is also spliced by an intron of 156 bp. A cDNA clone was prepared using the polymerase chain reaction, the sequence of which was used to confirm the presence of the intron in the coding sequence and the splicing of the 5' leader sequence. The transcriptional start point (tsp) was mapped at -253 nt from the site of the initiation of translation by primer extension and is adjacent to a 42-bp pyrimidine-rich region. The general structure of the 5' flanking region shows similarities to gpdA from Aspergillus nidulans. The putative protein product is 71-86% identical at the aa level to GPDs from Aspergillus nidulans, Cryphonectria parasitica, Curvularia lunata, Podospora anserina and Ustilago maydis.

  18. Expression of a novel cardiac-specific tropomyosin isoform in humans

    International Nuclear Information System (INIS)

    Denz, Christopher R.; Narshi, Aruna; Zajdel, Robert W.; Dube, Dipak K.

    2004-01-01

    Tropomyosins are a family of actin binding proteins encoded by a group of highly conserved genes. Humans have four tropomyosin-encoding genes: TPM1, TPM2, TPM3, and TPM4, each of which is known to generate multiple isoforms by alternative splicing, promoters, and 3 ' end processing. TPM1 is the most versatile and encodes a variety of tissue specific isoforms. The TPM1 isoform specific to striated muscle, designated TPM1α, consists of 10 exons: 1a, 2b, 3, 4, 5, 6b, 7, 8, and 9a/b. In this study, using RT-PCR with adult and fetal human RNAs, we present evidence for the expression of a novel isoform of the TPM1 gene that is specifically expressed in cardiac tissues. The new isoform is designated TPM1κ and contains exon 2a instead of 2b. Ectopic expression of human GFP.TPM1κ fusion protein can promote myofibrillogenesis in cardiac mutant axolotl hearts that are lacking in tropomyosin

  19. Isolation and Characterization of a Bacteriophage Preying an Antifungal Bacterium

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    Aryan Rahimi-Midani

    2016-12-01

    Full Text Available Several Bacillus species were isolated from rice field soils, and 16S rRNA gene sequence analysis showed that Bacillus cereus was the most abundant. A strain named BC1 showed antifungal activity against Rhizoctonia solani. Bacteriophages infecting strain BC1 were isolated from the same soil sample. The isolated phage PK16 had an icosahedral head of 100 ± 5 nm and tail of 200 ± 5 nm, indicating that it belonged to the family Myoviridae. Analysis of the complete linear dsDNA genome revealed a 158,127-bp genome with G + C content of 39.9% comprising 235 open reading frames as well as 19 tRNA genes (including 1 pseudogene. Blastp analysis showed that the proteins encoded by the PK16 genome had the closest hits to proteins of seven different bacteriophages. A neighbor-joining phylogenetic tree based on the major capsid protein showed a robust clustering of phage PK16 with phage JBP901 and BCP8-2 isolated from Korean fermented food.

  20. Standard guidelines for the chromosome-centric human proteome project.

    Science.gov (United States)

    Paik, Young-Ki; Omenn, Gilbert S; Uhlen, Mathias; Hanash, Samir; Marko-Varga, György; Aebersold, Ruedi; Bairoch, Amos; Yamamoto, Tadashi; Legrain, Pierre; Lee, Hyoung-Joo; Na, Keun; Jeong, Seul-Ki; He, Fuchu; Binz, Pierre-Alain; Nishimura, Toshihide; Keown, Paul; Baker, Mark S; Yoo, Jong Shin; Garin, Jerome; Archakov, Alexander; Bergeron, John; Salekdeh, Ghasem Hosseini; Hancock, William S

    2012-04-06

    The objective of the international Chromosome-Centric Human Proteome Project (C-HPP) is to map and annotate all proteins encoded by the genes on each human chromosome. The C-HPP consortium was established to organize a collaborative network among the research teams responsible for protein mapping of individual chromosomes and to identify compelling biological and genetic mechanisms influencing colocated genes and their protein products. The C-HPP aims to foster the development of proteome analysis and integration of the findings from related molecular -omics technology platforms through collaborations among universities, industries, and private research groups. The C-HPP consortium leadership has elicited broad input for standard guidelines to manage these international efforts more efficiently by mobilizing existing resources and collaborative networks. The C-HPP guidelines set out the collaborative consensus of the C-HPP teams, introduce topics associated with experimental approaches, data production, quality control, treatment, and transparency of data, governance of the consortium, and collaborative benefits. A companion approach for the Biology and Disease-Driven HPP (B/D-HPP) component of the Human Proteome Project is currently being organized, building upon the Human Proteome Organization's organ-based and biofluid-based initiatives (www.hupo.org/research). The common application of these guidelines in the participating laboratories is expected to facilitate the goal of a comprehensive analysis of the human proteome.

  1. Trypanosoma brucei Tb927.2.6100 Is an Essential Protein Associated with Kinetoplast DNA

    KAUST Repository

    Beck, K.; Acestor, N.; Schulfer, A.; Anupama, A.; Carnes, J.; Panigrahi, A. K.; Stuart, K.

    2013-01-01

    The mitochondrial DNA of trypanosomatid protozoa consists of a complex, intercatenated network of tens of maxicircles and thousands of minicircles. This structure, called kinetoplast DNA (kDNA), requires numerous proteins and multiprotein complexes for replication, segregation, and transcription. In this study, we used a proteomic approach to identify proteins that are associated with the kDNA network. We identified a novel protein encoded by Tb927.2.6100 that was present in a fraction enriched for kDNA and colocalized the protein with kDNA by fluorescence microscopy. RNA interference (RNAi) knockdown of its expression resulted in a growth defect and changes in the proportion of kinetoplasts and nuclei in the cell population. RNAi also resulted in shrinkage and loss of the kinetoplasts, loss of maxicircle and minicircle components of kDNA at similar rates, and (perhaps secondarily) loss of edited and pre-edited mRNA. These results indicate that the Tb927.2.6100 protein is essential for the maintenance of kDNA.

  2. A soluble diacylglycerol acyltransferase is involved in triacylglycerol biosynthesis in the oleaginous yeast Rhodotorula glutinis.

    Science.gov (United States)

    Rani, Sapa Hima; Saha, Saikat; Rajasekharan, Ram

    2013-01-01

    The biosynthesis of triacylglycerol (TAG) occurs in the microsomal membranes of eukaryotes. Here, we report the identification and functional characterization of diacylglycerol acyltransferase (DGAT), a member of the 10 S cytosolic TAG biosynthetic complex (TBC) in Rhodotorula glutinis. Both a full-length and an N-terminally truncated cDNA clone of a single gene were isolated from R. glutinis. The DGAT activity of the protein encoded by RgDGAT was confirmed in vivo by the heterologous expression of cDNA in a Saccharomyces cerevisiae quadruple mutant (H1246) that is defective in TAG synthesis. RgDGAT overexpression in yeast was found to be capable of acylating diacylglycerol (DAG) in an acyl-CoA-dependent manner. Quadruple mutant yeast cells exhibit growth defects in the presence of oleic acid, but wild-type yeast cells do not. In an in vivo fatty acid supplementation experiment, RgDGAT expression rescued quadruple mutant growth in an oleate-containing medium. We describe a soluble acyl-CoA-dependent DAG acyltransferase from R. glutinis that belongs to the DGAT3 class of enzymes. The study highlights the importance of an alternative TAG biosynthetic pathway in oleaginous yeasts.

  3. Modeling of the Ebola Virus Delta Peptide Reveals a Potential Lytic Sequence Motif

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    William R. Gallaher

    2015-01-01

    Full Text Available Filoviruses, such as Ebola and Marburg viruses, cause severe outbreaks of human infection, including the extensive epidemic of Ebola virus disease (EVD in West Africa in 2014. In the course of examining mutations in the glycoprotein gene associated with 2014 Ebola virus (EBOV sequences, a differential level of conservation was noted between the soluble form of glycoprotein (sGP and the full length glycoprotein (GP, which are both encoded by the GP gene via RNA editing. In the region of the proteins encoded after the RNA editing site sGP was more conserved than the overlapping region of GP when compared to a distant outlier species, Tai Forest ebolavirus. Half of the amino acids comprising the “delta peptide”, a 40 amino acid carboxy-terminal fragment of sGP, were identical between otherwise widely divergent species. A lysine-rich amphipathic peptide motif was noted at the carboxyl terminus of delta peptide with high structural relatedness to the cytolytic peptide of the non-structural protein 4 (NSP4 of rotavirus. EBOV delta peptide is a candidate viroporin, a cationic pore-forming peptide, and may contribute to EBOV pathogenesis.

  4. Sorbitol dehydrogenase is a cytosolic protein required for sorbitol metabolism in Arabidopsis thaliana.

    Science.gov (United States)

    Aguayo, María Francisca; Ampuero, Diego; Mandujano, Patricio; Parada, Roberto; Muñoz, Rodrigo; Gallart, Marta; Altabella, Teresa; Cabrera, Ricardo; Stange, Claudia; Handford, Michael

    2013-05-01

    Sorbitol is converted to fructose in Rosaceae species by SORBITOL DEHYDROGENASE (SDH, EC 1.1.1.14), especially in sink organs. SDH has also been found in non-Rosaceae species and here we show that the protein encoded by At5g51970 in Arabidopsis thaliana (L.) Heynh. possesses the molecular characteristics of an SDH. Using a green fluorescent protein-tagged version and anti-SDH antisera, we determined that SDH is cytosolically localized, consistent with bioinformatic predictions. We also show that SDH is widely expressed, and that SDH protein accumulates in both source and sink organs. In the presence of NAD+, recombinant SDH exhibited greatest oxidative activity with sorbitol, ribitol and xylitol as substrates; other sugar alcohols were oxidized to a lesser extent. Under standard growth conditions, three independent sdh- mutants developed as wild-type. Nevertheless, all three exhibited reduced dry weight and primary root length compared to wild-type when grown in the presence of sorbitol. Additionally, under short-day conditions, the mutants were more resistant to dehydration stress, as shown by a reduced loss of leaf water content when watering was withheld, and a greater survival rate on re-watering. This evidence suggests that limitations in the metabolism of sugar alcohols alter the growth of Arabidopsis and its response to drought. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  5. The Central Conserved Region (CCR) of Respiratory Syncytial Virus (RSV) G Protein Modulates Host miRNA Expression and Alters the Cellular Response to Infection.

    Science.gov (United States)

    Bakre, Abhijeet A; Harcourt, Jennifer L; Haynes, Lia M; Anderson, Larry J; Tripp, Ralph A

    2017-07-03

    Respiratory Syncytial Virus (RSV) infects respiratory epithelial cells and deregulates host gene expression by many mechanisms including expression of RSV G protein (RSV G). RSV G protein encodes a central conserved region (CCR) containing a CX3C motif that functions as a fractalkine mimic. Disruption of the CX3C motif (a.a. 182-186) located in the CCR of the G protein has been shown to affect G protein function in vitro and the severity of RSV disease pathogenesis in vivo. We show that infection of polarized Calu3 respiratory cells with recombinant RSV having point mutations in Cys173 and 176 (C173/176S) (rA2-GC12), or Cys186 (C186S) (rA2-GC4) is associated with a decline in the integrity of polarized Calu-3 cultures and decreased virus production. This is accompanied with downregulation of miRNAs let-7f and miR-24 and upregulation of interferon lambda (IFNλ), a primary antiviral cytokine for RSV in rA2-GC12/rA2-GC4 infected cells. These results suggest that residues in the cysteine noose region of RSV G protein can modulate IFN λ expression accompanied by downregulation of miRNAs, and are important for RSV G protein function and targeting.

  6. Structures and Corresponding Functions of Five Types of Picornaviral 2A Proteins

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    Xiaoyao Yang

    2017-07-01

    Full Text Available Among the few non-structural proteins encoded by the picornaviral genome, the 2A protein is particularly special, irrespective of structure or function. During the evolution of the Picornaviridae family, the 2A protein has been highly non-conserved. We believe that the 2A protein in this family can be classified into at least five distinct types according to previous studies. These five types are (A chymotrypsin-like 2A, (B Parechovirus-like 2A, (C hepatitis-A-virus-like 2A, (D Aphthovirus-like 2A, and (E 2A sequence of the genus Cardiovirus. We carried out a phylogenetic analysis and found that there was almost no homology between each type. Subsequently, we aligned the sequences within each type and found that the functional motifs in each type are highly conserved. These different motifs perform different functions. Therefore, in this review, we introduce the structures and functions of these five types of 2As separately. Based on the structures and functions, we provide suggestions to combat picornaviruses. The complexity and diversity of the 2A protein has caused great difficulties in functional and antiviral research. In this review, researchers can find useful information on the 2A protein and thus conduct improved antiviral research.

  7. Repetitive DNA and Plant Domestication: Variation in Copy Number and Proximity to Genes of LTR-Retrotransposons among Wild and Cultivated Sunflower (Helianthus annuus) Genotypes.

    Science.gov (United States)

    Mascagni, Flavia; Barghini, Elena; Giordani, Tommaso; Rieseberg, Loren H; Cavallini, Andrea; Natali, Lucia

    2015-11-24

    The sunflower (Helianthus annuus) genome contains a very large proportion of transposable elements, especially long terminal repeat retrotransposons. However, knowledge on the retrotransposon-related variability within this species is still limited. We used next-generation sequencing (NGS) technologies to perform a quantitative and qualitative survey of intraspecific variation of the retrotransposon fraction of the genome across 15 genotypes--7 wild accessions and 8 cultivars--of H. annuus. By mapping the Illumina reads of the 15 genotypes onto a library of sunflower long terminal repeat retrotransposons, we observed considerable variability in redundancy among genotypes, at both superfamily and family levels. In another analysis, we mapped Illumina paired reads to two sets of sequences, that is, long terminal repeat retrotransposons and protein-encoding sequences, and evaluated the extent of retrotransposon proximity to genes in the sunflower genome by counting the number of paired reads in which one read mapped to a retrotransposon and the other to a gene. Large variability among genotypes was also ascertained for retrotransposon proximity to genes. Both long terminal repeat retrotransposon redundancy and proximity to genes varied among retrotransposon families and also between cultivated and wild genotypes. Such differences are discussed in relation to the possible role of long terminal repeat retrotransposons in the domestication of sunflower. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  8. Cadherin-8 expression, synaptic localization, and molecular control of neuronal form in prefrontal corticostriatal circuits.

    Science.gov (United States)

    Friedman, Lauren G; Riemslagh, Fréderike W; Sullivan, Josefa M; Mesias, Roxana; Williams, Frances M; Huntley, George W; Benson, Deanna L

    2015-01-01

    Neocortical interactions with the dorsal striatum support many motor and executive functions, and such underlying functional networks are particularly vulnerable to a variety of developmental, neurological, and psychiatric brain disorders, including autism spectrum disorders, Parkinson's disease, and Huntington's disease. Relatively little is known about the development of functional corticostriatal interactions, and in particular, virtually nothing is known of the molecular mechanisms that control generation of prefrontal cortex-striatal circuits. Here, we used regional and cellular in situ hybridization techniques coupled with neuronal tract tracing to show that Cadherin-8 (Cdh8), a homophilic adhesion protein encoded by a gene associated with autism spectrum disorders and learning disability susceptibility, is enriched within striatal projection neurons in the medial prefrontal cortex and in striatal medium spiny neurons forming the direct or indirect pathways. Developmental analysis of quantitative real-time polymerase chain reaction and western blot data show that Cdh8 expression peaks in the prefrontal cortex and striatum at P10, when cortical projections start to form synapses in the striatum. High-resolution immunoelectron microscopy shows that Cdh8 is concentrated at excitatory synapses in the dorsal striatum, and Cdh8 knockdown in cortical neurons impairs dendritic arborization and dendrite self-avoidance. Taken together, our findings indicate that Cdh8 delineates developing corticostriatal circuits where it is a strong candidate for regulating the generation of normal cortical projections, neuronal morphology, and corticostriatal synapses. © 2014 Wiley Periodicals, Inc.

  9. Overexpression of three TaEXPA1 homoeologous genes with distinct expression divergence in hexaploid wheat exhibit functional retention in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Zhaorong Hu

    Full Text Available Common wheat is a hexaploid species with most of the genes present as triplicate homoeologs. Expression divergences of homoeologs are frequently observed in wheat as well as in other polyploid plants. However, little is known about functional variances among homologous genes arising from polyploidy. Expansins play diverse roles in plant developmental processes related to the action of cell wall loosening. Expression of the three TaEXPA1 homoeologs varied dynamically at different stages and organs, and epigenetic modifications contribute to the expression divergence of three TaEXPA1 homoeologs during wheat development. Nevertheless, their functions remain to be clarified. We found that over expression of TaEXPA1-A, -B and -D produced similar morphological changes in transgenic Arabidopsis plants, including increased germination and growth rate during seedling and adult stages, indicating that the proteins encoded by these three wheat TaEXPA1 homoeologs have similar (or conserved functions in Arabidopsis. Collectively, our present study provided an example of a set of homoeologous genes expression divergence in different developmental stages and organs in hexaploid wheat but functional retention in transgenic Arabidopsis plants.

  10. TRPP2-dependent Ca2+ signaling in dorso-lateral mesoderm is required for kidney field establishment in Xenopus.

    Science.gov (United States)

    Futel, Mélinée; Leclerc, Catherine; Le Bouffant, Ronan; Buisson, Isabelle; Néant, Isabelle; Umbhauer, Muriel; Moreau, Marc; Riou, Jean-François

    2015-03-01

    In Xenopus laevis embryos, kidney field specification is dependent on retinoic acid (RA) and coincides with a dramatic increase of Ca(2+) transients, but the role of Ca(2+) signaling in the kidney field is unknown. Here, we identify TRPP2, a member of the transient receptor potential (TRP) superfamily of channel proteins encoded by the pkd2 gene, as a central component of Ca(2+) signaling in the kidney field. TRPP2 is strongly expressed at the plasma membrane where it might regulate extracellular Ca(2+) entry. Knockdown of pkd2 in the kidney field results in the downregulation of pax8, but not of other kidney field genes (lhx1, osr1 and osr2). We further show that inhibition of Ca(2+) signaling with an inducible Ca(2+) chelator also causes downregulation of pax8, and that pkd2 knockdown results in a severe inhibition of Ca(2+) transients in kidney field explants. Finally, we show that disruption of RA results both in an inhibition of intracellular Ca(2+) signaling and of TRPP2 incorporation into the plasma membrane of kidney field cells. We propose that TRPP2-dependent Ca(2+) signaling is a key component of pax8 regulation in the kidney field downstream of RA-mediated non-transcriptional control of TRPP2. © 2015. Published by The Company of Biologists Ltd.

  11. Protein-protein association and cellular localization of four essential gene products encoded by tellurite resistance-conferring cluster "ter" from pathogenic Escherichia coli.

    Science.gov (United States)

    Valkovicova, Lenka; Vavrova, Silvia Minarikova; Mravec, Jozef; Grones, Jozef; Turna, Jan

    2013-12-01

    Gene cluster "ter" conferring high tellurite resistance has been identified in various pathogenic bacteria including Escherichia coli O157:H7. However, the precise mechanism as well as the molecular function of the respective gene products is unclear. Here we describe protein-protein association and localization analyses of four essential Ter proteins encoded by minimal resistance-conferring fragment (terBCDE) by means of recombinant expression. By using a two-plasmid complementation system we show that the overproduced single Ter proteins are not able to mediate tellurite resistance, but all Ter members play an irreplaceable role within the cluster. We identified several types of homotypic and heterotypic protein-protein associations among the Ter proteins by in vitro and in vivo pull-down assays and determined their cellular localization by cytosol/membrane fractionation. Our results strongly suggest that Ter proteins function involves their mutual association, which probably happens at the interface of the inner plasma membrane and the cytosol.

  12. Mos1 transposon-based transformation of fish cell lines using baculoviral vectors

    International Nuclear Information System (INIS)

    Yokoo, Masako; Fujita, Ryosuke; Nakajima, Yumiko; Yoshimizu, Mamoru; Kasai, Hisae; Asano, Shin-ichiro; Bando, Hisanori

    2013-01-01

    Highlights: •The baculovirus vector infiltrates the cells of economic important fishes. •Drosophila Mos1 transposase expressed in fish cells maintains its ability to localize to the nucleus. •The baculoviral vector carrying Mos1 is a useful tool to stably transform fish cells. -- Abstract: Drosophila Mos1 belongs to the mariner family of transposons, which are one of the most ubiquitous transposons among eukaryotes. We first determined nuclear transportation of the Drosophila Mos1-EGFP fusion protein in fish cell lines because it is required for a function of transposons. We next constructed recombinant baculoviral vectors harboring the Drosophila Mos1 transposon or marker genes located between Mos1 inverted repeats. The infectivity of the recombinant virus to fish cells was assessed by monitoring the expression of a fluorescent protein encoded in the viral genome. We detected transgene expression in CHSE-214, HINAE, and EPC cells, but not in GF or RTG-2 cells. In the co-infection assay of the Mos1-expressing virus and reporter gene-expressing virus, we successfully transformed CHSE-214 and HINAE cells. These results suggest that the combination of a baculovirus and Mos1 transposable element may be a tool for transgenesis in fish cells

  13. FANCL ubiquitinates β-catenin and enhances its nuclear function.

    Science.gov (United States)

    Dao, Kim-Hien T; Rotelli, Michael D; Petersen, Curtis L; Kaech, Stefanie; Nelson, Whitney D; Yates, Jane E; Hanlon Newell, Amy E; Olson, Susan B; Druker, Brian J; Bagby, Grover C

    2012-07-12

    Bone marrow failure is a nearly universal complication of Fanconi anemia. The proteins encoded by FANC genes are involved in DNA damage responses through the formation of a multisubunit nuclear complex that facilitates the E3 ubiquitin ligase activity of FANCL. However, it is not known whether loss of E3 ubiquitin ligase activity accounts for the hematopoietic stem cell defects characteristic of Fanconi anemia. Here we provide evidence that FANCL increases the activity and expression of β-catenin, a key pluripotency factor in hematopoietic stem cells. We show that FANCL ubiquitinates β-catenin with atypical ubiquitin chain extension known to have nonproteolytic functions. Specifically, β-catenin modified with lysine-11 ubiquitin chain extension efficiently activates a lymphocyte enhancer-binding factor-T cell factor reporter. We also show that FANCL-deficient cells display diminished capacity to activate β-catenin leading to reduced transcription of Wnt-responsive targets c-Myc and Cyclin D1. Suppression of FANCL expression in normal human CD34(+) stem and progenitor cells results in fewer β-catenin active cells and inhibits expansion of multilineage progenitors. Together, these results suggest that diminished Wnt/β-catenin signaling may be an underlying molecular defect in FANCL-deficient hematopoietic stem cells leading to their accelerated loss.

  14. Isoforms of U1-70k control subunit dynamics in the human spliceosomal U1 snRNP.

    Directory of Open Access Journals (Sweden)

    Helena Hernández

    2009-09-01

    Full Text Available Most human protein-encoding genes contain multiple exons that are spliced together, frequently in alternative arrangements, by the spliceosome. It is established that U1 snRNP is an essential component of the spliceosome, in human consisting of RNA and ten proteins, several of which are post-translationally modified and exist as multiple isoforms. Unresolved and challenging to investigate are the effects of these post translational modifications on the dynamics, interactions and stability of the particle. Using mass spectrometry we investigate the composition and dynamics of the native human U1 snRNP and compare native and recombinant complexes to isolate the effects of various subunits and isoforms on the overall stability. Our data reveal differential incorporation of four protein isoforms and dynamic interactions of subunits U1-A, U1-C and Sm-B/B'. Results also show that unstructured post-translationally modified C-terminal tails are responsible for the dynamics of Sm-B/B' and U1-C and that their interactions with the Sm core are controlled by binding to different U1-70k isoforms and their phosphorylation status in vivo. These results therefore provide the important functional link between proteomics and structure as well as insight into the dynamic quaternary structure of the native U1 snRNP important for its function.

  15. Biochemical and Genetic Characterization of Coagulin, a New Antilisterial Bacteriocin in the Pediocin Family of Bacteriocins, Produced by Bacillus coagulans I4

    Science.gov (United States)

    Le Marrec, Claire; Hyronimus, Bertrand; Bressollier, Philippe; Verneuil, Bernard; Urdaci, Maria C.

    2000-01-01

    A plasmid-linked antimicrobial peptide, named coagulin, produced by Bacillus coagulans I4 has recently been reported (B. Hyronimus, C. Le Marrec and M. C. Urdaci, J. Appl. Microbiol. 85:42–50, 1998). In the present study, the complete, unambiguous primary amino acid sequence of the peptide was obtained by a combination of both N-terminal sequencing of purified peptide and the complete sequence deduced from the structural gene harbored by plasmid I4. Data revealed that this peptide of 44 residues has an amino acid sequence similar to that described for pediocins AcH and PA-1, produced by different Pediococcus acidilactici strains and 100% identical. Coagulin and pediocin differed only by a single amino acid at their C terminus. Analysis of the genetic determinants revealed the presence, on the pI4 DNA, of the entire 3.5-kb operon of four genes described for pediocin AcH and PA-1 production. No extended homology was observed between pSMB74 from P. acidilactici and pI4 when analyzing the regions upstream and downstream of the operon. An oppositely oriented gene immediately dowstream of the bacteriocin operon specifies a 474-amino-acid protein which shows homology to Mob-Pre (plasmid recombination enzyme) proteins encoded by several small plasmids extracted from gram-positive bacteria. This is the first report of a pediocin-like peptide appearing naturally in a non-lactic acid bacterium genus. PMID:11097892

  16. Molecular cloning and expression of cDNA encoding a lumenal calcium binding glycoprotein from sarcoplasmic reticulum

    International Nuclear Information System (INIS)

    Leberer, E.; Charuk, J.H.M.; MacLennan, D.H.; Green, N.M.

    1989-01-01

    Antibody screening was used to isolate a cDNA encoding the 160-kDa glycoprotein of rabbit skeletal muscle sarcoplasmic reticulum. The cDNA is identical to that encoding the 53-kDa glycoprotein except that it contains an in-frame insertion of 1,308 nucleotides near its 5' end, apparently resulting from alternative splicing. The protein encoded by the cDNA would contain a 19-residue NH 2 -terminal signal sequence and a 453-residue COOH-terminal sequence identical to the 53-kDa glycoprotein. It would also contain a 436-amino acid insert between these sequences. This insert would be highly acidic, suggesting that it might bind Ca 2+ . The purified 160-kDa glycoprotein and the glycoprotein expressed in COS-1 cells transfected with cDNA encoding the 160-kDa glycoprotein were shown to bind 45 C 2+ in a gel overlay assay. The protein was shown to be located in the lumen of the sarcoplasmic reticulum and to be associated through Ca 2+ with the membrane. The authors propose that this lumenal Ca 2+ binding glycoprotein of the sarcoplasmic reticulum be designated sarcalumenin

  17. Characterization of hypothetical proteins Cpn0146, 0147, 0284 & 0285 that are predicted to be in the Chlamydia pneumoniae inclusion membrane

    Directory of Open Access Journals (Sweden)

    Liu Kaiyang

    2007-05-01

    Full Text Available Abstract Background Although more than 100 Chlamydia pneumoniae hypothetical proteins have been predicted to be inclusion membrane proteins, only a few have been experimentally demonstrated to be in the inclusion membrane. Using antibodies raised with fusion proteins, we characterized four such hypothetical proteins encoded by two gene clusters (Cpn0146-147 and Cpn0284-285 in the C. pneumoniae genome. Results Cpn0146 and 0147 were detected in the inclusion membrane while Cpn0284 and 0285 inside inclusion and mainly associated with reticulate bodies although all four proteins contain an N-terminal bi-lobed hydrophobic region, a signature motif assigned to inclusion membrane proteins. These four hypothetical proteins were only detected in cells infected with C. pneumoniae but not other chlamydial species, with Cpn0147 at 6 hours and Cpn0146, 0284 & 0285 at 24 hours after infection. Cpn0146 & 147 but not Cpn0284 and 285 co-localized with a host cell endoplasmic reticulum marker, a property known to be possessed by some chlamydial inclusion membrane proteins, when expressed in the host cell cytosol via transgenes. However, the endoplasmic reticulum localization of the C. pneumoniae inclusion membrane proteins did not result in inhibition of the subsequent C. pneumoniae infection. Conclusion The hypothetical proteins Cpn0146 & 0147 were localized in the C. pneumoniae inclusion membrane while Cpn0284 & 0285 within the inclusion although all four were predicted to be Inc proteins, suggesting the need to experimentally characterize the predicted Inc proteins.

  18. Computational investigation of small RNAs in the establishment of root nodules and arbuscular mycorrhiza in leguminous plants.

    Science.gov (United States)

    Jin, Danfeng; Meng, Xianwen; Wang, Yue; Wang, Jingjing; Zhao, Yuhua; Chen, Ming

    2018-01-03

    Many small RNAs have been confirmed to play important roles in the development of root nodules and arbuscular mycorrhiza. In this study, we carried out the identification of certain small RNAs in leguminous plants (Medicago truncatula, soybean, peanut and common bean), such as miRNAs, tRFs and srRNAs, as well as the computational investigation of their regulations. Thirty miRNAs were predicted to be involved in establishing root nodules and mycorrhiza, and 12 of them were novel in common bean and peanut. The generation of tRFs in M. truncatula was not associated with tRNA gene frequencies and codon usage. Six tRFs exhibited different expressions in mycorrhiza and root nodules. Moreover, srRNA 5.8S in M. truncatula was generated from the regions with relatively low conservation at the rRNA 3' terminal. The protein-protein interactions between the proteins encoded by the target genes of miRNAs, tRFs and srRNAs were computed. The regulation of these three types of sRNAs in the symbiosis between leguminous plants and microorganisms is not a single regulation of certain signaling or metabolic pathways but a global regulation for the plants to own growth or specific events in symbiosis.

  19. A strong loss-of-function mutation in RAN1 results in constitutive activation of the ethylene response pathway as well as a rosette-lethal phenotype

    Science.gov (United States)

    Woeste, K. E.; Kieber, J. J.; Evans, M. L. (Principal Investigator)

    2000-01-01

    A recessive mutation was identified that constitutively activated the ethylene response pathway in Arabidopsis and resulted in a rosette-lethal phenotype. Positional cloning of the gene corresponding to this mutation revealed that it was allelic to responsive to antagonist1 (ran1), a mutation that causes seedlings to respond in a positive manner to what is normally a competitive inhibitor of ethylene binding. In contrast to the previously identified ran1-1 and ran1-2 alleles that are morphologically indistinguishable from wild-type plants, this ran1-3 allele results in a rosette-lethal phenotype. The predicted protein encoded by the RAN1 gene is similar to the Wilson and Menkes disease proteins and yeast Ccc2 protein, which are integral membrane cation-transporting P-type ATPases involved in copper trafficking. Genetic epistasis analysis indicated that RAN1 acts upstream of mutations in the ethylene receptor gene family. However, the rosette-lethal phenotype of ran1-3 was not suppressed by ethylene-insensitive mutants, suggesting that this mutation also affects a non-ethylene-dependent pathway regulating cell expansion. The phenotype of ran1-3 mutants is similar to loss-of-function ethylene receptor mutants, suggesting that RAN1 may be required to form functional ethylene receptors. Furthermore, these results suggest that copper is required not only for ethylene binding but also for the signaling function of the ethylene receptors.

  20. Protein Structure Prediction by Protein Threading

    Science.gov (United States)

    Xu, Ying; Liu, Zhijie; Cai, Liming; Xu, Dong

    The seminal work of Bowie, Lüthy, and Eisenberg (Bowie et al., 1991) on "the inverse protein folding problem" laid the foundation of protein structure prediction by protein threading. By using simple measures for fitness of different amino acid types to local structural environments defined in terms of solvent accessibility and protein secondary structure, the authors derived a simple and yet profoundly novel approach to assessing if a protein sequence fits well with a given protein structural fold. Their follow-up work (Elofsson et al., 1996; Fischer and Eisenberg, 1996; Fischer et al., 1996a,b) and the work by Jones, Taylor, and Thornton (Jones et al., 1992) on protein fold recognition led to the development of a new brand of powerful tools for protein structure prediction, which we now term "protein threading." These computational tools have played a key role in extending the utility of all the experimentally solved structures by X-ray crystallography and nuclear magnetic resonance (NMR), providing structural models and functional predictions for many of the proteins encoded in the hundreds of genomes that have been sequenced up to now.

  1. Myristylation of gag-onc fusion proteins in mammalian transforming retroviruses

    International Nuclear Information System (INIS)

    Schultz, A.; Oroszlan, S.

    1984-01-01

    Four cell lines producing transforming proteins encoded by three mammalian oncogenes (fes, abl, and ras) were investigated for incorporation of [ 3 H]myristate into gag-onc fusion proteins. Using 5-min pulse-labelings, fusion proteins of Abelson murine leukemia virus, Gardner-Arnstein strain of feline sarcoma virus (FeSV), and Snyder-Theilen strain of FeSV were shown to be myristylated. In a 4-hr pulse, p29gag-ras of rat sarcoma virus (RaSV) was also shown to incorporate radiolabel. The fatty acid was recovered from this labeled protein by acid hydrolysis, and identified by reverse-phase thin-layer chromatography to be [ 3 H]myristic acid. The results indicate that substitution of viral gag sequences by cellular oncogene sequences does not abolish their ability to become post-translationally modified by this long chain fatty acid. It is assumed that in the fusion proteins the myristyl moiety is linked through an amide linkage to the amino-terminal glycine as previously found for several retroviral gag precursor polyproteins. The possible role of myristylation of transforming proteins is discussed

  2. Myristylation of gag-onc fusion proteins in mammalian transforming retroviruses

    Energy Technology Data Exchange (ETDEWEB)

    Schultz, A.; Oroszlan, S.

    1984-03-01

    Four cell lines producing transforming proteins encoded by three mammalian oncogenes (fes, abl, and ras) were investigated for incorporation of (/sup 3/H)myristate into gag-onc fusion proteins. Using 5-min pulse-labelings, fusion proteins of Abelson murine leukemia virus, Gardner-Arnstein strain of feline sarcoma virus (FeSV), and Snyder-Theilen strain of FeSV were shown to be myristylated. In a 4-hr pulse, p29gag-ras of rat sarcoma virus (RaSV) was also shown to incorporate radiolabel. The fatty acid was recovered from this labeled protein by acid hydrolysis, and identified by reverse-phase thin-layer chromatography to be (/sup 3/H)myristic acid. The results indicate that substitution of viral gag sequences by cellular oncogene sequences does not abolish their ability to become post-translationally modified by this long chain fatty acid. It is assumed that in the fusion proteins the myristyl moiety is linked through an amide linkage to the amino-terminal glycine as previously found for several retroviral gag precursor polyproteins. The possible role of myristylation of transforming proteins is discussed.

  3. Unexplored therapeutic opportunities in the human genome.

    Science.gov (United States)

    Oprea, Tudor I; Bologa, Cristian G; Brunak, Søren; Campbell, Allen; Gan, Gregory N; Gaulton, Anna; Gomez, Shawn M; Guha, Rajarshi; Hersey, Anne; Holmes, Jayme; Jadhav, Ajit; Jensen, Lars Juhl; Johnson, Gary L; Karlson, Anneli; Leach, Andrew R; Ma'ayan, Avi; Malovannaya, Anna; Mani, Subramani; Mathias, Stephen L; McManus, Michael T; Meehan, Terrence F; von Mering, Christian; Muthas, Daniel; Nguyen, Dac-Trung; Overington, John P; Papadatos, George; Qin, Jun; Reich, Christian; Roth, Bryan L; Schürer, Stephan C; Simeonov, Anton; Sklar, Larry A; Southall, Noel; Tomita, Susumu; Tudose, Ilinca; Ursu, Oleg; Vidovic, Dušica; Waller, Anna; Westergaard, David; Yang, Jeremy J; Zahoránszky-Köhalmi, Gergely

    2018-05-01

    A large proportion of biomedical research and the development of therapeutics is focused on a small fraction of the human genome. In a strategic effort to map the knowledge gaps around proteins encoded by the human genome and to promote the exploration of currently understudied, but potentially druggable, proteins, the US National Institutes of Health launched the Illuminating the Druggable Genome (IDG) initiative in 2014. In this article, we discuss how the systematic collection and processing of a wide array of genomic, proteomic, chemical and disease-related resource data by the IDG Knowledge Management Center have enabled the development of evidence-based criteria for tracking the target development level (TDL) of human proteins, which indicates a substantial knowledge deficit for approximately one out of three proteins in the human proteome. We then present spotlights on the TDL categories as well as key drug target classes, including G protein-coupled receptors, protein kinases and ion channels, which illustrate the nature of the unexplored opportunities for biomedical research and therapeutic development.

  4. Transcriptional Responses in the Hemiparasitic Plant Triphysaria versicolor to Host Plant Signals1[w

    Science.gov (United States)

    Matvienko, Marta; Torres, Manuel J.; Yoder, John I.

    2001-01-01

    Parasitic plants in the Scrophulariaceae use chemicals released by host plant roots to signal developmental processes critical for heterotrophy. Haustoria, parasitic plant structures that attach to and invade host roots, develop on roots of the hemiparasitic plant Triphysaria versicolor within a few hours of exposure to either maize (Zea mays) root exudate or purified haustoria-inducing factors. We prepared a normalized, subtractive cDNA library enriched for transcripts differentially abundant in T. versicolor root tips treated with the allelopathic quinone 2,6-dimethoxybenzoquinone (DMBQ). Northern analyses estimated that about 10% of the cDNAs represent transcripts strongly up-regulated in roots exposed to DMBQ. Northern and reverse northern analyses demonstrated that most DMBQ-responsive messages were similarly up-regulated in T. versicolor roots exposed to maize root exudates. From the cDNA sequences we assembled a unigene set of 137 distinct transcripts and assigned functions by homology comparisons. Many of the proteins encoded by the transcripts are predicted to function in quinone detoxification, whereas others are more likely associated with haustorium development. The identification of genes transcriptionally regulated by haustorium-inducing factors provides a framework for dissecting genetic pathways recruited by parasitic plants during the transition to heterotrophic growth. PMID:11553755

  5. Flock House virus subgenomic RNA3 is replicated and its replication correlates with transactivation of RNA2

    International Nuclear Information System (INIS)

    Eckerle, Lance D.; Albarino, Cesar G.; Ball, L. Andrew.

    2003-01-01

    The nodavirus Flock House virus has a bipartite genome composed of RNAs 1 and 2, which encode the catalytic component of the RNA-dependent RNA polymerase (RdRp) and the capsid protein precursor, respectively. In addition to catalyzing replication of the viral genome, the RdRp also transcribes from RNA1 a subgenomic RNA3, which is both required for and suppressed by RNA2 replication. Here, we show that in the absence of RNA1 replication, FHV RdRp replicated positive-sense RNA3 transcripts fully and copied negative-sense RNA3 transcripts into positive strands. The two nonstructural proteins encoded by RNA3 were dispensable for replication, but sequences in the 3'-terminal 58 nucleotides were required. RNA3 variants that failed to replicate also failed to transactivate RNA2. These results imply that RNA3 is naturally produced both by transcription from RNA1 and by subsequent RNA1-independent replication and that RNA3 replication may be necessary for transactivation of RNA2

  6. Mos1 transposon-based transformation of fish cell lines using baculoviral vectors

    Energy Technology Data Exchange (ETDEWEB)

    Yokoo, Masako [Laboratory of Applied Molecular Entomology, Division of Applied Bioscience, Research Faculty of Agriculture, Hokkaido University, Sapporo 060-8589 (Japan); Fujita, Ryosuke [Laboratory of Applied Molecular Entomology, Division of Applied Bioscience, Research Faculty of Agriculture, Hokkaido University, Sapporo 060-8589 (Japan); Innate Immunity Laboratory, Graduate School of Life Science and Creative Research Institution, Hokkaido University, Sapporo 001-0021 (Japan); Nakajima, Yumiko [Functional Genomics Group, COMB, Tropical Biosphere Research Center, University of the Ryukyus, Okinawa 903-0213 (Japan); Yoshimizu, Mamoru; Kasai, Hisae [Faculty of Fisheries Sciences, Hokkaido University, Hakodate 041-8611 (Japan); Asano, Shin-ichiro [Laboratory of Applied Molecular Entomology, Division of Applied Bioscience, Research Faculty of Agriculture, Hokkaido University, Sapporo 060-8589 (Japan); Bando, Hisanori, E-mail: hban@abs.agr.hokudai.ac.jp [Laboratory of Applied Molecular Entomology, Division of Applied Bioscience, Research Faculty of Agriculture, Hokkaido University, Sapporo 060-8589 (Japan)

    2013-09-13

    Highlights: •The baculovirus vector infiltrates the cells of economic important fishes. •Drosophila Mos1 transposase expressed in fish cells maintains its ability to localize to the nucleus. •The baculoviral vector carrying Mos1 is a useful tool to stably transform fish cells. -- Abstract: Drosophila Mos1 belongs to the mariner family of transposons, which are one of the most ubiquitous transposons among eukaryotes. We first determined nuclear transportation of the Drosophila Mos1-EGFP fusion protein in fish cell lines because it is required for a function of transposons. We next constructed recombinant baculoviral vectors harboring the Drosophila Mos1 transposon or marker genes located between Mos1 inverted repeats. The infectivity of the recombinant virus to fish cells was assessed by monitoring the expression of a fluorescent protein encoded in the viral genome. We detected transgene expression in CHSE-214, HINAE, and EPC cells, but not in GF or RTG-2 cells. In the co-infection assay of the Mos1-expressing virus and reporter gene-expressing virus, we successfully transformed CHSE-214 and HINAE cells. These results suggest that the combination of a baculovirus and Mos1 transposable element may be a tool for transgenesis in fish cells.

  7. Elevated Fibroblast Growth Factor Signaling Is Critical for the Pathogenesis of the Dwarfism in Evc2/Limbin Mutant Mice

    Science.gov (United States)

    Zhang, Honghao; Kamiya, Nobuhiro; Tsuji, Takehito; Takeda, Haruko; Scott, Greg; Ray, Manas K.; Mochida, Yoshiyuki; Lefebvre, Veronique; Hung, Irene H.; Kunieda, Tetsuo; Mishina, Yuji

    2016-01-01

    Ellis-van Creveld (EvC) syndrome is a skeletal dysplasia, characterized by short limbs, postaxial polydactyly, and dental abnormalities. EvC syndrome is also categorized as a ciliopathy because of ciliary localization of proteins encoded by the two causative genes, EVC and EVC2 (aka LIMBIN). While recent studies demonstrated important roles for EVC/EVC2 in Hedgehog signaling, there is still little known about the pathophysiological mechanisms underlying the skeletal dysplasia features of EvC patients, and in particular why limb development is affected, but not other aspects of organogenesis that also require Hedgehog signaling. In this report, we comprehensively analyze limb skeletogenesis in Evc2 mutant mice and in cell and tissue cultures derived from these mice. Both in vivo and in vitro data demonstrate elevated Fibroblast Growth Factor (FGF) signaling in Evc2 mutant growth plates, in addition to compromised but not abrogated Hedgehog-PTHrP feedback loop. Elevation of FGF signaling, mainly due to increased Fgf18 expression upon inactivation of Evc2 in the perichondrium, critically contributes to the pathogenesis of limb dwarfism. The limb dwarfism phenotype is partially rescued by inactivation of one allele of Fgf18 in the Evc2 mutant mice. Taken together, our data uncover a novel pathogenic mechanism to understand limb dwarfism in patients with Ellis-van Creveld syndrome. PMID:28027321

  8. Development of a single-dose recombinant CAMP factor entrapping poly(lactide-co-glycolide) microspheres-based vaccine against Streptococcus agalactiae.

    Science.gov (United States)

    Liu, Gang; Yin, Jinhua; Barkema, Herman W; Chen, Liben; Shahid, Muhammad; Szenci, Otto; De Buck, Jeroen; Kastelic, John P; Han, Bo

    2017-03-01

    Streptococcus agalactiae is an important contagious bovine mastitis pathogen. Although it is well controlled and even eradicated in most Northern European and North American dairy herds, the prevalence of this pathogen remains very high in China. However, research on development of a vaccine against S. agalactiae mastitis is scarce. The aims of the present study were to: (1) develop a single-dose vaccine against S. agalactiae based on poly(lactic-co-glycolic acid) (PLGA) microspheres (MS) encapsulated CAMP factor, a conserved virulent protein encoded by S. agalactiae's cfb gene; and (2) evaluate its immunogenicity and protective efficacy in a mouse model. The cfb gene was cloned and expressed in a recombinant Escherichia coli strain Trans1-T1. The CAMP factor was tested to determine a safe dose range and then encapsulated in MS of PLGA (50:50) to assess its release pattern in vitro and immune reaction in vivo. Furthermore, a mouse model and a histopathological assay were developed to evaluate bacterial burden and vaccine efficacy. In the low dosage range (S. agalactiae challenge. Additionally, no pathological lesions were detected in the vaccinated group. Therefore, PLGA-CAMP conferred protective efficacy against S. agalactiae in our mouse model, indicating its potential as a vaccine against S. agalactiae mastitis. Furthermore, the slow-release kinetics of PLGA MS warranted optimism for development of a single-dose vaccine. Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.

  9. Extensive overproduction of the AdhE protein by rng mutations depends on mutations in the cra gene or in the Cra-box of the adhE promoter.

    Science.gov (United States)

    Kaga, Naoko; Umitsuki, Genryou; Clark, David P; Nagai, Kazuo; Wachi, Masaaki

    2002-07-05

    Escherichia coli RNase G encoded by the rng gene is involved in degradation of adhE mRNA. Overproduction of the AdhE protein by rng mutants was found to depend on the genetic background of strains derived from DC272 (adhC81) or MC1061. We found that DC272 carried a point mutation in the Cra-binding site of the adhE promoter. The Cra protein encoded by the cra gene is known to act as a repressor of adhE. P1-phage-mediated transduction and lacZ fusion analysis with the mutant adhE promoter confirmed that this mutation is responsible for overproduction. On the other hand, Southern hybridization revealed that MC1061 had a 0.85-kb deletion of the cra gene. Overproduction of AdhE in the MC1061 background was reversed to the wild-type levels by introduction of a plasmid carrying the cra(+) gene. These results indicated that expression of the adhE gene was regulated transcriptionally by Cra and posttranscriptionally by RNase G. (c) 2002 Elsevier Science (USA).

  10. Crystal structure of the toxin Msmeg_6760, the structural homolog of Mycobacterium tuberculosis Rv2035, a novel type II toxin involved in the hypoxic response

    Energy Technology Data Exchange (ETDEWEB)

    Bajaj, R. Alexandra; Arbing, Mark A.; Shin, Annie; Cascio, Duilio; Miallau, Linda (UCLA)

    2016-11-19

    The structure of Msmeg_6760, a protein of unknown function, has been determined. Biochemical and bioinformatics analyses determined that Msmeg_6760 interacts with a protein encoded in the same operon, Msmeg_6762, and predicted that the operon is a toxin–antitoxin (TA) system. Structural comparison of Msmeg_6760 with proteins of known function suggests that Msmeg_6760 binds a hydrophobic ligand in a buried cavity lined by large hydrophobic residues. Access to this cavity could be controlled by a gate–latch mechanism. The function of the Msmeg_6760 toxin is unknown, but structure-based predictions revealed that Msmeg_6760 and Msmeg_6762 are homologous to Rv2034 and Rv2035, a predicted novel TA system involved inMycobacterium tuberculosislatency during macrophage infection. The Msmeg_6760 toxin fold has not been previously described for bacterial toxins and its unique structural features suggest that toxin activation is likely to be mediated by a novel mechanism.

  11. Baculovirus proteins IE-1, LEF-3, and P143 interact with DNA in vivo: a formaldehyde cross-linking study

    International Nuclear Information System (INIS)

    Ito, Emma; Sahri, Daniela; Knippers, Rolf; Carstens, Eric B.

    2004-01-01

    IE-1, LEF-3, and P143 are three of six proteins encoded by Autographa californica nucleopolyhedrovirus (AcMNPV) essential for baculovirus DNA replication in transient replication assays. IE-1 is the major baculovirus immediate early transcription regulator. LEF-3 is a single-stranded DNA binding protein (SSB) and P143 is a DNA helicase protein. To investigate their interactions in vivo, we treated AcMNPV-infected Spodoptera frugiperda cells with formaldehyde and separated soluble proteins from chromatin by cell fractionation and cesium chloride equilibrium centrifugation. Up to 70% of the total LEF-3 appeared in the fraction of soluble, probably nucleoplasmic proteins, while almost all P143 and IE-1 were associated with viral chromatin in the nucleus. This suggests that LEF-3 is produced in quantities that are higher than needed for the coverage of single stranded regions that arise during viral DNA replication and is consistent with the hypothesis that LEF-3 has other functions such as the localization of P143 to the nucleus. Using a chromatin immunoprecipitation procedure, we present the first direct evidence of LEF-3, P143, and IE-1 proteins binding to closely linked sites on viral chromatin in vivo, suggesting that they may form replication complexes on viral DNA in infected cells

  12. Antigenic characterisation of yeast-expressed lyssavirus nucleoproteins.

    Science.gov (United States)

    Kucinskaite, Indre; Juozapaitis, Mindaugas; Serva, Andrius; Zvirbliene, Aurelija; Johnson, Nicholas; Staniulis, Juozas; Fooks, Anthony R; Müller, Thomas; Sasnauskas, Kestutis; Ulrich, Rainer G

    2007-12-01

    In Europe, three genotypes of the genus Lyssavirus, family Rhabdoviridae, are present, classical rabies virus (RABV, genotype 1), European bat lyssavirus type 1 (EBLV-1, genotype 5) and European bat lyssavirus type 2 (EBLV-2, genotype 6). The entire authentic nucleoprotein (N protein) encoding sequences of RABV (challenge virus standard, CVS, strain), EBLV-1 and EBLV-2 were expressed in yeast Saccharomyces cerevisiae at high level. Purification of recombinant N proteins by caesium chloride gradient centrifugation resulted in yields between 14-17, 25-29 and 18-20 mg/l of induced yeast culture for RABV-CVS, EBLV-1 and EBLV-2, respectively. The purified N proteins were evaluated by negative staining electron microscopy, which revealed the formation of nucleocapsid-like structures. The antigenic conformation of the N proteins was investigated for their reactivity with monoclonal antibodies (mAbs) directed against different lyssaviruses. The reactivity pattern of each mAb was virtually identical between immunofluorescence assay with virus-infected cells, and ELISA and dot blot assay using the corresponding recombinant N proteins. These observations lead us to conclude that yeast-expressed lyssavirus N proteins share antigenic properties with naturally expressed virus protein. These recombinant proteins have the potential for use as components of serological assays for lyssaviruses.

  13. Overexpression of DgWRKY4 Enhances Salt Tolerance in Chrysanthemum Seedlings

    Directory of Open Access Journals (Sweden)

    Ke Wang

    2017-09-01

    Full Text Available High salinity seriously affects the production of chrysanthemum, so improving the salt tolerance of chrysanthemum becomes the focus and purpose of our research. The WRKY transcription factor (TF family is highly associated with a number of processes of abiotic stress responses. We isolated DgWRKY4 from Dendranthema grandiflorum, and a protein encoded by this new gene contains two highly conserved WRKY domains and two C2H2 zinc-finger motifs. Then, we functionally characterized that DgWRKY4 was induced by salt, and DgWRKY4 overexpression in chrysanthemum resulted in increased tolerance to high salt stress compared to wild-type (WT. Under salt stress, the transgenic chrysanthemum accumulated less malondialdehyde, hydrogen peroxide (H2O2, and superoxide anion (O2− than WT, accompanied by more proline, soluble sugar, and activities of antioxidant enzymes than WT; in addition, a stronger photosynthetic capacity and a series of up-regulated stress-related genes were also found in transgenic chrysanthemum. All results demonstrated that DgWRKY4 is a positive regulatory gene responding to salt stress, via advancing photosynthetic capacity, promoting the operation of reactive oxygen species-scavenging system, maintaining membrane stability, enhancing the osmotic adjustment, and up-regulating transcript levels of stress-related genes. So, DgWRKY4 can serve as a new candidate gene for salt-tolerant plant breeding.

  14. Complete nucleotide sequence of the RNA-2 of grapevine deformation and Grapevine Anatolian ringspot viruses.

    Science.gov (United States)

    Ghanem-Sabanadzovic, Nina Abou; Sabanadzovic, Sead; Digiaro, Michele; Martelli, Giovanni P

    2005-05-01

    The nucleotide sequence of RNA-2 of Grapevine Anatolian ringspot virus (GARSV) and Grapevine deformation virus (GDefV), two recently described nepoviruses, has been determined. These RNAs are 3753 nt (GDefV) and 4607 nt (GARSV) in size and contain a single open reading frame encoding a polyprotein of 122 kDa (GDefV) and 150 kDa (GARSV). Full-length nucleotide sequence comparison disclosed 71-73% homology between GDefV RNA-2 and that of Grapevine fanleaf virus (GFLV) and Arabis mosaic virus (ArMV), and 62-64% homology between GARSV RNA-2 and that of Grapevine chrome mosaic virus (GCMV) and Tomato black ring virus (TBRV). As previously observed in other nepoviruses, the 5' non-coding regions of both RNAs are capable of forming stem-loop structures. Phylogenetic analysis of the three proteins encoded by RNA-2 (i.e. protein 2A, movement protein and coat protein) confirmed that GDefV and GARSV are distinct viruses which can be assigned as definitive species in subgroup A and subgroup B of the genus Nepovirus, respectively.

  15. A newly identified protein of Leptospira interrogans mediates binding to laminin.

    Science.gov (United States)

    Longhi, Mariana T; Oliveira, Tatiane R; Romero, Eliete C; Gonçales, Amane P; de Morais, Zenaide M; Vasconcellos, Silvio A; Nascimento, Ana L T O

    2009-10-01

    Pathogenic Leptospira is the aetiological agent of leptospirosis, a life-threatening disease that affects populations worldwide. The search for novel antigens that could be relevant in host-pathogen interactions is being pursued. These antigens have the potential to elicit several activities, including adhesion. This study focused on a hypothetical predicted lipoprotein of Leptospira, encoded by the gene LIC12895, thought to mediate attachment to extracellular matrix (ECM) components. The gene was cloned and expressed in Escherichia coli BL21 Star (DE3)pLys by using the expression vector pAE. The recombinant protein tagged with N-terminal hexahistidine was purified by metal-charged chromatography and characterized by circular dichroism spectroscopy. The capacity of the protein to mediate attachment to ECM components was evaluated by binding assays. The leptospiral protein encoded by LIC12895, named Lsa27 (leptospiral surface adhesin, 27 kDa), bound strongly to laminin in a dose-dependent and saturable fashion. Moreover, Lsa27 was recognized by antibodies from serum samples of confirmed leptospirosis specimens in both the initial and the convalescent phases of the disease. Lsa27 is most likely a surface protein of Leptospira as revealed in liquid-phase immunofluorescence assays with living organisms. Taken together, these data indicate that this newly identified membrane protein is expressed during natural infection and may play a role in mediating adhesion of L. interrogans to its host.

  16. Characterization of three novel adhesins of Leptospira interrogans.

    Science.gov (United States)

    Siqueira, Gabriela H; Atzingen, Marina V; Alves, Ivy J; de Morais, Zenaide M; Vasconcellos, Silvio A; Nascimento, Ana L T O

    2013-12-01

    We report cloning, expression, purification, and characterization of three predicted leptospiral membrane proteins (LIC11360, LIC11009, and LIC11975). In silico analysis and proteinase K accessibility data suggest that these proteins might be surface exposed. We show that proteins encoded by LIC11360, LIC11009 and LIC11975 genes interact with laminin in a dose-dependent and saturable manner. The proteins are referred to as leptospiral surface adhesions 23, 26, and 36 (Lsa23, Lsa26, and Lsa36), respectively. These proteins also bind plasminogen and generate active plasmin. Attachment of Lsa23 and Lsa36 to fibronectin occurs through the involvement of the 30-kDa and 70-kDa heparin-binding domains of the ligand. Dose-dependent, specific-binding of Lsa23 to the complement regulator C4BP and to a lesser extent, to factor H, suggests that this protein may interfere with the complement cascade pathways. Leptospira spp. may use these interactions as possible mechanisms during the establishment of infection.

  17. Functional identification of conserved residues involved in Lactobacillus rhamnosus strain GG sortase specificity and pilus biogenesis.

    Science.gov (United States)

    Douillard, François P; Rasinkangas, Pia; von Ossowski, Ingemar; Reunanen, Justus; Palva, Airi; de Vos, Willem M

    2014-05-30

    In Gram-positive bacteria, sortase-dependent pili mediate the adhesion of bacteria to host epithelial cells and play a pivotal role in colonization, host signaling, and biofilm formation. Lactobacillus rhamnosus strain GG, a well known probiotic bacterium, also displays on its cell surface mucus-binding pilus structures, along with other LPXTG surface proteins, which are processed by sortases upon specific recognition of a highly conserved LPXTG motif. Bioinformatic analysis of all predicted LPXTG proteins encoded by the L. rhamnosus GG genome revealed a remarkable conservation of glycine residues juxtaposed to the canonical LPXTG motif. Here, we investigated and defined the role of this so-called triple glycine (TG) motif in determining sortase specificity during the pilus assembly and anchoring. Mutagenesis of the TG motif resulted in a lack or an alteration of the L. rhamnosus GG pilus structures, indicating that the TG motif is critical in pilus assembly and that they govern the pilin-specific and housekeeping sortase specificity. This allowed us to propose a regulatory model of the L. rhamnosus GG pilus biogenesis. Remarkably, the TG motif was identified in multiple pilus gene clusters of other Gram-positive bacteria, suggesting that similar signaling mechanisms occur in other, mainly pathogenic, species. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Screening of Genes Specifically Expressed in Males of Fenneropenaeus chinensis and Their Potential as Sex Markers

    Directory of Open Access Journals (Sweden)

    Shihao Li

    2013-01-01

    Full Text Available The androgenic gland (AG, playing an important role in sex differentiation of male crustacean, is a target candidate to understand the mechanism of male development and to mine male-specific sex markers. An SSH library (designated as male reproduction-related tissues—SSH library, MRT-SSH library for short was constructed using cDNA from tissues located at the basal part of the 5th pereiopods, including AG and part of spermatophore sac, as tester, and the cDNA from the basal part of the 4th pereiopods of these male shrimp as driver. 402 ESTs from the SSH library were sequenced and assembled into 48 contigs and 104 singlets. Twelve contigs and 14 singlets were identified as known genes. The proteins encoded by the identified genes were categorized, according to their proposed functions, into neuropeptide hormone and hormone transporter, RNA posttranscriptional regulation, translation, cell growth and death, metabolism, genetic information processing, signal transduction/transport, or immunity-related proteins. Eleven highly expressed contigs in the SSH library were selected for validation of the MRT-SSH library and screening sex markers of shrimp. One contig, specifically expressed in male shrimp, had a potential to be developed as a transcriptomic sex marker in shrimp.

  19. Increased methylation and decreased expression of homeobox genes TLX1, HOXA10 and DLX5 in human placenta are associated with trophoblast differentiation.

    Science.gov (United States)

    Novakovic, Boris; Fournier, Thierry; Harris, Lynda K; James, Joanna; Roberts, Claire T; Yong, Hannah E J; Kalionis, Bill; Evain-Brion, Danièle; Ebeling, Peter R; Wallace, Euan M; Saffery, Richard; Murthi, Padma

    2017-07-03

    Homeobox genes regulate embryonic and placental development, and are widely expressed in the human placenta, but their regulatory control by DNA methylation is unclear. DNA methylation analysis was performed on human placentae from first, second and third trimesters to determine methylation patterns of homeobox gene promoters across gestation. Most homeobox genes were hypo-methylated throughout gestation, suggesting that DNA methylation is not the primary mechanism involved in regulating HOX genes expression in the placenta. Nevertheless, several genes showed variable methylation patterns across gestation, with a general trend towards an increase in methylation over gestation. Three genes (TLX1, HOXA10 and DLX5) showed inverse gains of methylation with decreasing mRNA expression throughout pregnancy, supporting a role for DNA methylation in their regulation. Proteins encoded by these genes were primarily localised to the syncytiotrophoblast layer, and showed decreased expression later in gestation. siRNA mediated downregulation of DLX5, TLX1 and HOXA10 in primary term villous cytotrophoblast resulted in decreased proliferation and increased expression of differentiation markers, including ERVW-1. Our data suggest that loss of DLX5, TLX1 and HOXA10 expression in late gestation is required for proper placental differentiation and function.

  20. The KNOXI Transcription Factor SHOOT MERISTEMLESS Regulates Floral Fate in Arabidopsis.

    Science.gov (United States)

    Roth, Ohad; Alvarez, John; Levy, Matan; Bowman, John L; Ori, Naomi; Shani, Eilon

    2018-05-09

    Plants have evolved a unique and conserved developmental program that enables the conversion of leaves into floral organs. Elegant genetic and molecular work has identified key regulators of flower meristem identity. However, further understanding of flower meristem specification has been hampered by redundancy and by pleiotropic effects. The KNOXI transcription factor SHOOT MERISTEMLESS (STM) is a well-characterized regulator of shoot apical meristem maintenance. Arabidopsis thaliana stm loss-of-function mutants arrest shortly after germination, and therefore the knowledge on later roles of STM in later processes, including flower development, is limited. Here, we uncover a role for STM in the specification of flower meristem identity. Silencing STM in the APETALA1 (AP1) expression domain in the ap1-4 mutant background resulted in a leafy-flower phenotype, and an intermediate stm-2 allele enhanced the flower meristem identity phenotype of ap1-4. Transcriptional profiling of STM perturbation suggested that STM activity affects multiple floral fate genes, among them the F-Box protein-encoding gene UNUSUAL FLORAL ORGANS (UFO). In agreement with this notion, stm-2 enhanced the ufo-2 floral fate phenotype, and ectopic UFO expression rescued the leafy flowers in genetic backgrounds with compromised AP1 and STM activities. This work suggests a genetic mechanism that underlies the activity of STM in the specification of flower meristem identity. © 2018 American Society of Plant Biologists. All rights reserved.