Sample records for abundant constitutive expression

  1. Abundant constitutive expression of the immediate-early 94K protein from cytomegalovirus (Colburn) in a DNA-transfected mouse cell line

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Jeang, K.T.; Cho, M.S.; Hayward, G.S.

    1984-10-01

    A 94-kilodalton phosphoprotein known as IE94 is the only viral polypeptide synthesized in abundance under immediate-early conditions after infection by cytomegalovirus (CMV) strain Colburn in either permissive primate or nonpermissive rodent cells. The authors isolated a clonal Ltk/sup +/ cell line which expressed the /sup 35/methionine-labeled IE94 polypeptide in sufficient abundance to be visualized directly in autoradiographs after gel electrophoresis of total-cell-culture protein extracts. The IE94 polypeptide synthesized in the transfected cells was indistinguishable in size and overall net charge from that produced in virus-infected cells. In addition, the IE94 protein expressed in LH/sub 2/p198-3 cells was phosphorylated (presumably bymore » a cellular protein kinase) and generated similar phosphopeptide patterns after partial tryptic digestion to those obtained with the CMV IE94 protein from infected cells. The cell line contained two to four stably integrated copies of the IE94 gene and synthesized a single virus-specific mRNA of 2.5 kilobases detectable on Northern blots. A new antigen, detectable by indirect anticomplement immunofluorescence with monoclonal antibody against the human CMV IE68 protein, was present in the nuclei of more than 95% of the LH/sub 2/l198-3 cells. This evidence suggests that (unlike most herpesvirus genes) the CMV IE94 gene, together with its complex promoter and spliced mRNA structure, may contain all of the regulatory elements necessary for strong constitutive expression in mammalian cells in the absence of other viral factors.« less

  2. Constitutive expression of Botrytis aclada laccase in Pichia pastoris

    PubMed Central

    Kittl, Roman; Gonaus, Christoph; Pillei, Christian; Haltrich, Dietmar; Ludwig, Roland

    2012-01-01

    The heterologous expression of laccases is important for their large-scale production and genetic engineering—a prerequisite for industrial application. Pichia pastoris is the preferred expression host for fungal laccases. The recently cloned laccase from the ascomycete Botrytis aclada (BaLac) has been efficiently expressed in P. pastoris under the control of the inducible alcohol oxidase (AOX1) promoter. In this study, we compare these results to the constitutive expression in the same organism using the glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter. The results show that the amounts of BaLac produced with the GAP system (517 mgL-1) and the AOX1 system (495 mgL-1) are comparable. The constitutive expression is, however, faster, and the specific activity of BaLac in the culture supernatant is higher (41.3 Umg-1 GAP, 14.2 Umg-1 AOX1). In microtiter plates, the constitutive expression provides a clear advantage due to easy manipulation (simple medium, no methanol feeding) and fast enzyme production (high-throughput screening assays can already be performed after 48 h). PMID:22705842

  3. Metaproteomics reveals abundant transposase expression in mutualistic endosymbionts

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kleiner, Manuel; Young, Jacque C; Shah, Manesh B

    2013-01-01

    Transposases, enzymes that catalyze the movement of mobile genetic elements, are the most abundant genes in nature. While many bacteria encode an abundance of transposases in their genomes, the current paradigm is that transposase gene expression is tightly regulated and generally low due to its severe mutagenic effects. In the current study, we detected the highest number of transposase proteins ever reported in bacteria, in symbionts of the gutless marine worm Olavius algarvensis using metaproteomics. At least 26 different transposases from 12 different families were detected and genomic and proteomic analyses suggest many of these are active. This high expressionmore » of transposases indicates that the mechanisms for their tight regulation have been disabled or destroyed. Based on recent studies on other symbionts and pathogens that showed high transposase transcription, we speculate that abundant transposase expression might be common in symbionts and pathogens.« less

  4. Constitutive nitrate reductase expression and inhibition in winged bean

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wu, Shenchuan; Harper, J.E.

    It was found that NO{sub 3}{sup {minus}} had no effect on winged bean nitrate reductase activity (NRA). Similar NRA was expressed in plants grown on NO{sub 3}{sup {minus}}, urea, NH{sub 4}{sup +}, and nil N. This indicated that the primary NR expressed in winged bean was constitutive, rather than substrate-inducible. Maximum NRA in winged bean was obtained in the light. KClO{sub 3} was capable of inhibiting NRA of leaves if added to the root growth medium or to the NR assay medium, indicating possible competition with NO{sub 3}{sup {minus}} at the reduction site. While it has previously been shown thatmore » either cycloheximide alone, or both cycloheximide and chloramphenicol impair the synthesis of NR protein, our data unexpectedly demonstrated that cycloheximide had little effect on NRA, whereas chloramphenicol greatly inhibited the expression of NRA in winged bean. One interpretation is that chloroplasts may influence the activity and/or synthesis of constitutive NR proteins.« less

  5. Constitutive and Stress-induced Expression of CCL5 Machinery in Rodent Retina

    PubMed Central

    Duncan, D'Anne S.; McLaughlin, William M.; Vasilakes, Noah; Echevarria, Franklin D.; Formichella, Cathryn R.; Sappington, Rebecca M.

    2017-01-01

    Signaling by inflammatory cytokines and chemokines is associated with neurodegeneration in disease and injury. Here we examined expression of the β-chemokine CCL5 and its receptors in the mouse retina and evaluated its relevance in glaucoma, a common optic neuropathy associated with sensitivity to intraocular pressure (IOP). Using quantitative PCR, fluorescent in situ hybridization, immunohistochemistry and quantitative image analysis, we found CCL5 mRNA and protein was constitutively expressed in the inner retina and synaptic layers. CCL5 appeared to associate with Müller cells and RGCs as well as synaptic connections between horizontal cells and bipolar cells in the OPL and amacrine cells, bipolar cells and RGCs in the IPL. Although all three high-affinity receptors (CCR5, CCR3, CCR1) for CCL5 were expressed constitutively, CCR5 expression was significantly higher than CCR3, which was also markedly greater than CCR1. Localization patterns for constitutive CCR5, CCR3 and CCR1 expression differed, particularly with respect to expression in inner retinal neurons. Stress-related expression of CCL5 was primarily altered in aged DBA/2 mice with elevated IOP. In contrast, changes in expression and localization of both CCR3 and CCR5 were evident not only in aged DBA/2 mice, but also in age-matched control mice and young DBA/2 mice. These groups do not exhibit elevated IOP, but possess either the aging stress (control mice) or the genetic predisposition to glaucoma (DBA/2 mice). Together, these data indicate that CCL5 and its high-affinity receptors are constitutively expressed in murine retina and differentially induced by stressors associated with glaucomatous optic neuropathy. Localization patterns further indicate that CCL5 signaling may be relevant for modulation of synapses in both health and disease, particularly in the inner plexiform layer. PMID:28936366

  6. Genetic requirements for high constitutive SOS expression in recA730 mutants of Escherichia coli.

    PubMed

    Vlašić, Ignacija; Šimatović, Ana; Brčić-Kostić, Krunoslav

    2011-09-01

    The RecA protein in its functional state is in complex with single-stranded DNA, i.e., in the form of a RecA filament. In SOS induction, the RecA filament functions as a coprotease, enabling the autodigestion of the LexA repressor. The RecA filament can be formed by different mechanisms, but all of them require three enzymatic activities essential for the processing of DNA double-stranded ends. These are helicase, 5'-3' exonuclease, and RecA loading onto single-stranded DNA (ssDNA). In some mutants, the SOS response can be expressed constitutively during the process of normal DNA metabolism. The RecA730 mutant protein is able to form the RecA filament without the help of RecBCD and RecFOR mediators since it better competes with the single-strand binding (SSB) protein for ssDNA. As a consequence, the recA730 mutants show high constitutive SOS expression. In the study described in this paper, we studied the genetic requirements for constitutive SOS expression in recA730 mutants. Using a β-galactosidase assay, we showed that the constitutive SOS response in recA730 mutants exhibits different requirements in different backgrounds. In a wild-type background, the constitutive SOS response is partially dependent on RecBCD function. In a recB1080 background (the recB1080 mutation retains only helicase), constitutive SOS expression is partially dependent on RecBCD helicase function and is strongly dependent on RecJ nuclease. Finally, in a recB-null background, the constitutive SOS expression of the recA730 mutant is dependent on the RecJ nuclease. Our results emphasize the importance of the 5'-3' exonuclease for high constitutive SOS expression in recA730 mutants and show that RecBCD function can further enhance the excellent intrinsic abilities of the RecA730 protein in vivo. Copyright © 2011, American Society for Microbiology. All Rights Reserved.

  7. A Constitutive Expression System for Cellulase Secretion in Escherichia coli and Its Use in Bioethanol Production

    PubMed Central

    Munjal, Neha; Jawed, Kamran; Wajid, Saima; Yazdani, Syed Shams

    2015-01-01

    The production of biofuels from lignocellulosic biomass appears to be attractive and viable due to the abundance and availability of this biomass. The hydrolysis of this biomass, however, is challenging because of the complex lignocellulosic structure. The ability to produce hydrolytic cellulase enzymes in a cost-effective manner will certainly accelerate the process of making lignocellulosic ethanol production a commercial reality. These cellulases may need to be produced aerobically to generate large amounts of protein in a short time or anaerobically to produce biofuels from cellulose via consolidated bioprocessing. Therefore, it is important to identify a promoter that can constitutively drive the expression of cellulases under both aerobic and anaerobic conditions without the need for an inducer. Using lacZ as reporter gene, we analyzed the strength of the promoters of four genes, namely lacZ, gapA, ldhA and pflB, and found that the gapA promoter yielded the maximum expression of the β-galactosidase enzyme under both aerobic and anaerobic conditions. We further cloned the genes for two cellulolytic enzymes, β-1,4-endoglucanase and β-1,4-glucosidase, under the control of the gapA promoter, and we expressed these genes in Escherichia coli, which secreted the products into the extracellular medium. An ethanologenic E. colistrain transformed with the secretory β-glucosidase gene construct fermented cellobiose in both defined and complex medium. This recombinant strain also fermented wheat straw hydrolysate containing glucose, xylose and cellobiose into ethanol with an 85% efficiency of biotransformation. An ethanologenic strain that constitutively secretes a cellulolytic enzyme is a promising platform for producing lignocellulosic ethanol. PMID:25768292

  8. Expression Profiling of Transcriptome and Its Associated Disease Risk in Yang Deficiency Constitution of Healthy Subjects

    PubMed Central

    Yu, Ruoxi; Yang, Yin; Han, Yuanyuan; Hou, Pengwei; Li, Yingshuai; Li, Siqi

    2016-01-01

    Objectives. Differences among healthy subjects and associated disease risks are of substantial interest in clinical medicine. According to the theory of “constitution-disease correlation” in traditional Chinese medicine, we try to find out if there is any connection between intolerance of cold in Yang deficiency constitution and molecular evidence and if there is any gene expression basis in specific disorders. Methods. Peripheral blood mononuclear cells were collected from Chinese Han individuals with Yang deficiency constitution (n = 20) and balanced constitution (n = 8) (aged 18–28) and global gene expression profiles were determined between them using the Affymetrix HG-U133 Plus 2.0 array. Results. The results showed that when the fold change was ≥1.2 and q ≤ 0.05, 909 genes were upregulated in the Yang deficiency constitution, while 1189 genes were downregulated. According to our research differential genes found in Yang deficiency constitution were usually related to lower immunity, metabolic disorders, and cancer tendency. Conclusion. Gene expression disturbance exists in Yang deficiency constitution, which corresponds to the concept of constitution and gene classification. It also suggests people with Yang deficiency constitution are susceptible to autoimmune diseases, enteritis, arthritis, metabolism disorders, and cancer, which provides molecular evidence for the theory of “constitution-disease correlation.” PMID:28484499

  9. Whole genome expression and biochemical correlates of extreme constitutional types defined in Ayurveda.

    PubMed

    Prasher, Bhavana; Negi, Sapna; Aggarwal, Shilpi; Mandal, Amit K; Sethi, Tav P; Deshmukh, Shailaja R; Purohit, Sudha G; Sengupta, Shantanu; Khanna, Sangeeta; Mohammad, Farhan; Garg, Gaurav; Brahmachari, Samir K; Mukerji, Mitali

    2008-09-09

    Ayurveda is an ancient system of personalized medicine documented and practiced in India since 1500 B.C. According to this system an individual's basic constitution to a large extent determines predisposition and prognosis to diseases as well as therapy and life-style regime. Ayurveda describes seven broad constitution types (Prakritis) each with a varying degree of predisposition to different diseases. Amongst these, three most contrasting types, Vata, Pitta, Kapha, are the most vulnerable to diseases. In the realm of modern predictive medicine, efforts are being directed towards capturing disease phenotypes with greater precision for successful identification of markers for prospective disease conditions. In this study, we explore whether the different constitution types as described in Ayurveda has molecular correlates. Normal individuals of the three most contrasting constitutional types were identified following phenotyping criteria described in Ayurveda in Indian population of Indo-European origin. The peripheral blood samples of these individuals were analysed for genome wide expression levels, biochemical and hematological parameters. Gene Ontology (GO) and pathway based analysis was carried out on differentially expressed genes to explore if there were significant enrichments of functional categories among Prakriti types. Individuals from the three most contrasting constitutional types exhibit striking differences with respect to biochemical and hematological parameters and at genome wide expression levels. Biochemical profiles like liver function tests, lipid profiles, and hematological parameters like haemoglobin exhibited differences between Prakriti types. Functional categories of genes showing differential expression among Prakriti types were significantly enriched in core biological processes like transport, regulation of cyclin dependent protein kinase activity, immune response and regulation of blood coagulation. A significant enrichment of

  10. Whole genome expression and biochemical correlates of extreme constitutional types defined in Ayurveda

    PubMed Central

    Prasher, Bhavana; Negi, Sapna; Aggarwal, Shilpi; Mandal, Amit K; Sethi, Tav P; Deshmukh, Shailaja R; Purohit, Sudha G; Sengupta, Shantanu; Khanna, Sangeeta; Mohammad, Farhan; Garg, Gaurav; Brahmachari, Samir K; Mukerji, Mitali

    2008-01-01

    Background Ayurveda is an ancient system of personalized medicine documented and practiced in India since 1500 B.C. According to this system an individual's basic constitution to a large extent determines predisposition and prognosis to diseases as well as therapy and life-style regime. Ayurveda describes seven broad constitution types (Prakritis) each with a varying degree of predisposition to different diseases. Amongst these, three most contrasting types, Vata, Pitta, Kapha, are the most vulnerable to diseases. In the realm of modern predictive medicine, efforts are being directed towards capturing disease phenotypes with greater precision for successful identification of markers for prospective disease conditions. In this study, we explore whether the different constitution types as described in Ayurveda has molecular correlates. Methods Normal individuals of the three most contrasting constitutional types were identified following phenotyping criteria described in Ayurveda in Indian population of Indo-European origin. The peripheral blood samples of these individuals were analysed for genome wide expression levels, biochemical and hematological parameters. Gene Ontology (GO) and pathway based analysis was carried out on differentially expressed genes to explore if there were significant enrichments of functional categories among Prakriti types. Results Individuals from the three most contrasting constitutional types exhibit striking differences with respect to biochemical and hematological parameters and at genome wide expression levels. Biochemical profiles like liver function tests, lipid profiles, and hematological parameters like haemoglobin exhibited differences between Prakriti types. Functional categories of genes showing differential expression among Prakriti types were significantly enriched in core biological processes like transport, regulation of cyclin dependent protein kinase activity, immune response and regulation of blood coagulation. A

  11. Constitutive gene expression and specification of tissue identity in adult planarian biology

    PubMed Central

    Reddien, Peter W.

    2011-01-01

    Planarians are flatworms that constitutively maintain adult tissues through cell turnover and can regenerate entire organisms from tiny body fragments. In addition to requiring new cells (from neoblasts), these feats require mechanisms that specify tissue identity in the adult. Critical roles for Wnt and BMP signaling in regeneration and maintenance of the body axes have been uncovered, among other regulatory factors. Available data indicate that genes involved in positional identity regulation at key embryonic stages in other animals display persisting regionalized expression in adult planarians. These expression patterns suggest that a constitutively active gene expression map exists for maintenance of the planarian body. Planarians therefore present a fertile ground for identification of factors regulating regionalization of the metazoan body plan and for study of the attributes of these factors that can lead to maintenance and regeneration of adult tissues. PMID:21680047

  12. Preprodynorphin-expressing neurons constitute a large subgroup of somatostatin-expressing GABAergic interneurons in the mouse neocortex.

    PubMed

    Sohn, Jaerin; Hioki, Hiroyuki; Okamoto, Shinichiro; Kaneko, Takeshi

    2014-05-01

    Dynorphins, leumorphin, and neoendorphins are preprodynorphin (PPD)-derived peptides and ligands for κ-opioid receptors. Using an antibody to PPD C-terminal, we investigated the chemical and molecular characteristics of PPD-expressing neurons in mouse neocortex. PPD-immunopositive neuronal somata were distributed most frequently in layer 5 and less frequently in layers 2-4 and 6 throughout neocortical regions. Combined labeling of immunofluorescence and fluorescent mRNA signals revealed that almost all PPD-immunopositive neurons expressed glutamic acid decarboxylase but not vesicular glutamate transporter, indicating their γ-aminobutyric acid (GABA)ergic characteristics, and that PPD-immunopositive neurons accounted for 15% of GABAergic interneurons in the primary somatosensory area. As GABAergic interneurons were divided into several groups by specific markers, we further examined the chemical characteristics of PPD-expressing neurons by the double immunofluorescence labeling method. More than 95% of PPD-immunopositive neurons were also somatostatin (SOM)-immunopositive in the primary somatosensory, primary motor, orbitofrontal, and primary visual areas, but only 24% were SOM-immunopositive in the medial prefrontal cortex. In the primary somatosensory area, PPD-immunopositive neurons constituted 50%, 79%, 55%, and 17% of SOM-immunopositive neurons in layers 2-3, 4, 5, and 6, respectively. Although SOM-expressing neurons contained calretinin-, neuropeptide Y-, nitric oxide synthase-, and reelin-expressing neurons as subgroups, only reelin immunoreactivity was detected in many PPD-immunopositive neurons. These results indicate that PPD-expressing neurons constitute a large subgroup of SOM-expressing cortical interneurons, and the PPD/SOM-expressing GABAergic neurons might serve not only as inhibitory elements in the local cortical circuit, but also as modulators for cortical neurons expressing κ-opioid and/or SOM receptors. Copyright © 2013 Wiley Periodicals

  13. Generation of a stable cell line for constitutive miRNA expression.

    PubMed

    Lieber, Diana

    2013-01-01

    miRNAs have in recent years emerged as novel players in virus-host interactions. While individual miRNAs are capable of regulating many targets simultaneously, not much is known about the role of distinct host or viral miRNAs in the context of infection. Analysis of the function of a miRNA is often hampered by the complexity of virus-host interactions and the enormous changes in the host cell during infection. Many viral miRNAs as for example from Kaposi sarcoma-associated Herpesvirus (KSHV) are probably exclusively expressed in latent infection. This might lead to a steady-state situation with offense and defense mechanisms counteracting each other. Cellular miRNAs involved in defense against pathogens on the other hand might be suppressed in infection. A cell culture system allowing for constitutive expression of individual miRNAs at high levels is a useful tool to enhance miRNA-specific functions and to uncouple viral miRNA function from other infection-related mechanisms. Here, a protocol is described to generate stable cell lines for constitutive expression of single cellular or viral miRNA precursors in absence of infection. The procedure comprises cloning of the precursor sequence, generation of the lentiviral expression vector, transduction of the cells of interest, selection for polyclonal cell lines, and isolation of monoclonal cell lines by limiting dilution.

  14. Hepatic cytochrome P450 activity, abundance, and expression throughout human development

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Sadler, Natalie C.; Nandhikonda, Premchendar; Webb-Robertson, Bobbie-Jo M.

    Cytochrome P450s are Phase I metabolic enzymes that play critical roles in the biotransformation of endogenous compounds and xenobiotics. The expression and activity of P450 enzymes can vary considerably throughout human development, especially when comparing fetal development to neonates, children, and adults. In an effort to develop a more comprehensive understanding of the ontogeny of P450 expression and activity we employed a multi-omic characterization of P450 transcript expression, protein abundance, and functional activity. To quantify the functional activity of individual P450s we employ activity-based protein profiling, which uses modified mechanism-based inhibitors of P450s as chemical probes, in tandem with proteomicmore » analyses to quantify activity. Our results reveal life-stage-dependent variability in P450 expression, abundance, and activity throughout human development and frequent discordant relationships between expression and activity. The results were used to distribute P450s into three general classes based upon developmental stage of expression and activity. We have significantly expanded the knowledge of P450 ontogeny, particularly at the level of individual P450 activity. We anticipate that our ontogeny results will be useful for enabling predictive therapeutic dosing, and for avoiding potentially adverse and harmful reactions during maturation from both therapeutic drugs and environmental xenobiotics.« less

  15. Constitutive Expression of Short Hairpin RNA in Vivo Triggers Buildup of Mature Hairpin Molecules

    PubMed Central

    Ahn, M.; Witting, S.R.; Ruiz, R.; Saxena, R.

    2011-01-01

    Abstract RNA interference (RNAi) has become the cornerstone technology for studying gene function in mammalian cells. In addition, it is a promising therapeutic treatment for multiple human diseases. Virus-mediated constitutive expression of short hairpin RNA (shRNA) has the potential to provide a permanent source of silencing molecules to tissues, and it is being devised as a strategy for the treatment of liver conditions such as hepatitis B and hepatitis C virus infection. Unintended interaction between silencing molecules and cellular components, leading to toxic effects, has been described in vitro. Despite the enormous interest in using the RNAi technology for in vivo applications, little is known about the safety of constitutively expressing shRNA for multiple weeks. Here we report the effects of in vivo shRNA expression, using helper-dependent adenoviral vectors. We show that gene-specific knockdown is maintained for at least 6 weeks after injection of 1 × 1011 viral particles. Nonetheless, accumulation of mature shRNA molecules was observed up to weeks 3 and 4, and then declined gradually, suggesting the buildup of mature shRNA molecules induced cell death with concomitant loss of viral DNA and shRNA expression. No evidence of well-characterized innate immunity activation (such as interferon production) or saturation of the exportin-5 pathway was observed. Overall, our data suggest constitutive expression of shRNA results in accumulation of mature shRNA molecules, inducing cellular toxicity at late time points, despite the presence of gene silencing. PMID:21780944

  16. Enhancing Antitumor Efficacy of Chimeric Antigen Receptor T Cells Through Constitutive CD40L Expression

    PubMed Central

    Curran, Kevin J; Seinstra, Beatrijs A; Nikhamin, Yan; Yeh, Raymond; Usachenko, Yelena; van Leeuwen, Dayenne G; Purdon, Terence; Pegram, Hollie J; Brentjens, Renier J

    2015-01-01

    Adoptive cell therapy with genetically modified T cells expressing a chimeric antigen receptor (CAR) is a promising therapy for patients with B-cell acute lymphoblastic leukemia. However, CAR-modified T cells (CAR T cells) have mostly failed in patients with solid tumors or low-grade B-cell malignancies including chronic lymphocytic leukemia with bulky lymph node involvement. Herein, we enhance the antitumor efficacy of CAR T cells through the constitutive expression of CD40 ligand (CD40L, CD154). T cells genetically modified to constitutively express CD40L (CD40L-modified T cells) demonstrated increased proliferation and secretion of proinflammatory TH1 cytokines. Further, CD40L-modified T cells augmented the immunogenicity of CD40+ tumor cells by the upregulated surface expression of costimulatory molecules (CD80 and CD86), adhesion molecules (CD54, CD58, and CD70), human leukocyte antigen (HLA) molecules (Class I and HLA-DR), and the Fas-death receptor (CD95). Additionally, CD40L-modified T cells induced maturation and secretion of the proinflammatory cytokine interleukin-12 by monocyte-derived dendritic cells. Finally, tumor-targeted CD19-specific CAR/CD40L T cells exhibited increased cytotoxicity against CD40+ tumors and extended the survival of tumor-bearing mice in a xenotransplant model of CD19+ systemic lymphoma. This preclinical data supports the clinical application of CAR T cells additionally modified to constitutively express CD40L with anticipated enhanced antitumor efficacy. PMID:25582824

  17. Genome-wide Determinants of Proviral Targeting, Clonal Abundance and Expression in Natural HTLV-1 Infection

    PubMed Central

    Melamed, Anat; Laydon, Daniel J.; Gillet, Nicolas A.; Tanaka, Yuetsu; Taylor, Graham P.; Bangham, Charles R. M.

    2013-01-01

    The regulation of proviral latency is a central problem in retrovirology. We postulate that the genomic integration site of human T lymphotropic virus type 1 (HTLV-1) determines the pattern of expression of the provirus, which in turn determines the abundance and pathogenic potential of infected T cell clones in vivo. We recently developed a high-throughput method for the genome-wide amplification, identification and quantification of proviral integration sites. Here, we used this protocol to test two hypotheses. First, that binding sites for transcription factors and chromatin remodelling factors in the genome flanking the proviral integration site of HTLV-1 are associated with integration targeting, spontaneous proviral expression, and in vivo clonal abundance. Second, that the transcriptional orientation of the HTLV-1 provirus relative to that of the nearest host gene determines spontaneous proviral expression and in vivo clonal abundance. Integration targeting was strongly associated with the presence of a binding site for specific host transcription factors, especially STAT1 and p53. The presence of the chromatin remodelling factors BRG1 and INI1 and certain host transcription factors either upstream or downstream of the provirus was associated respectively with silencing or spontaneous expression of the provirus. Cells expressing HTLV-1 Tax protein were significantly more frequent in clones of low abundance in vivo. We conclude that transcriptional interference and chromatin remodelling are critical determinants of proviral latency in natural HTLV-1 infection. PMID:23555266

  18. Constitutive type VI secretion system expression gives Vibrio cholerae intra- and interspecific competitive advantages.

    PubMed

    Unterweger, Daniel; Kitaoka, Maya; Miyata, Sarah T; Bachmann, Verena; Brooks, Teresa M; Moloney, Jessica; Sosa, Oscar; Silva, David; Duran-Gonzalez, Jorge; Provenzano, Daniele; Pukatzki, Stefan

    2012-01-01

    The type VI secretion system (T6SS) mediates protein translocation across the cell membrane of Gram-negative bacteria, including Vibrio cholerae - the causative agent of cholera. All V. cholerae strains examined to date harbor gene clusters encoding a T6SS. Structural similarity and sequence homology between components of the T6SS and the T4 bacteriophage cell-puncturing device suggest that the T6SS functions as a contractile molecular syringe to inject effector molecules into prokaryotic and eukaryotic target cells. Regulation of the T6SS is critical. A subset of V. cholerae strains, including the clinical O37 serogroup strain V52, express T6SS constitutively. In contrast, pandemic strains impose tight control that can be genetically disrupted: mutations in the quorum sensing gene luxO and the newly described regulator gene tsrA lead to constitutive T6SS expression in the El Tor strain C6706. In this report, we examined environmental V. cholerae isolates from the Rio Grande with regard to T6SS regulation. Rough V. cholerae lacking O-antigen carried a nonsense mutation in the gene encoding the global T6SS regulator VasH and did not display virulent behavior towards Escherichia coli and other environmental bacteria. In contrast, smooth V. cholerae strains engaged constitutively in type VI-mediated secretion and displayed virulence towards prokaryotes (E. coli and other environmental bacteria) and a eukaryote (the social amoeba Dictyostelium discoideum). Furthermore, smooth V. cholerae strains were able to outcompete each other in a T6SS-dependent manner. The work presented here suggests that constitutive T6SS expression provides V. cholerae with an advantage in intraspecific and interspecific competition.

  19. Constitutively expressed COX-2 in osteoblasts positively regulates Akt signal transduction via suppression of PTEN activity.

    PubMed

    Li, Ching-Ju; Chang, Je-Ken; Wang, Gwo-Jaw; Ho, Mei-Ling

    2011-02-01

    Cyclooxygenase-2 (COX-2) is thought to be an inducible enzyme, but increasing reports indicate that COX-2 is constitutively expressed in several organs. The status of COX-2 expression in bone and its physiological role remains undefined. Non-selective non-steroidal anti-inflammatory drugs (NSAIDs) and selective COX-2 inhibitors, which commonly suppress COX-2 activity, were reported to suppress osteoblast proliferation via Akt/FOXO3a/p27(Kip1) signaling, suggesting that COX-2 may be the key factor of the suppressive effects of NSAIDs on proliferation. Although Akt activation correlates with PTEN deficiency and cell viability, the role of COX-2 on PTEN/Akt regulation remains unclear. In this study, we hypothesized that COX-2 may be constitutively expressed in osteoblasts and regulate PTEN/Akt-related proliferation. We examined the localization and co-expression of COX-2 and p-Akt in normal mouse femurs and in cultured mouse (mOBs) and human osteoblasts (hOBs). Our results showed that osteoblasts adjacent to the trabeculae, periosteum and endosteum in mouse femurs constitutively expressed COX-2, while COX-2 co-expressed with p-Akt in osteoblasts sitting adjacent to trabeculae in vivo, and in mOBs and hOBs in vitro. We further used COX-2 siRNA to test the role of COX-2 in Akt signaling in hOBs; COX-2 silencing significantly inhibited PTEN phosphorylation, enhanced PTEN activity, and suppressed p-Akt level and proliferation. However, replenishment of the COX-2 enzymatic product, PGE2, failed to reverse COX-2-dependent Akt phosphorylation. Furthermore, transfection with recombinant human COX-2 (rhCOX-2) significantly reversed COX-2 siRNA-suppressed PTEN phosphorylation, but this effect was reduced when the enzymatic activity of rhCOX-2 was blocked. This finding indicated that the effect of COX-2 on PTEN/Akt signaling is not related to PGE2 but still dependent on COX-2 enzymatic activity. Conversely, COX-1 silencing did not affect PTEN/Akt signaling. Our findings provide

  20. Constitutive expression of human pancreatic lipase-related protein 1 in Pichia pastoris.

    PubMed

    Aloulou, Ahmed; Grandval, Philippe; De Caro, Josiane; De Caro, Alain; Carrière, Frédéric

    2006-06-01

    High-level constitutive expression of the human pancreatic lipase-related protein 1 (HPLRP1) was achieved using the methylotrophic yeast Pichia pastoris. The HPLRP1 cDNA, including its original leader sequence, was subcloned into the pGAPZB vector and further integrated into the genome of P. pastoris X-33 under the control of the glyceraldehyde 3-phosphate dehydrogenase (GAP) constitutive promoter. A major protein with a molecular mass of 50 kDa was found to be secreted into the culture medium and was identified using anti-HPLRP1 polyclonal antibodies as HPLRP1 recombinant protein. The level of expression reached 100-120 mg of HPLRP1 per liter of culture medium after 40 h, as attested by specific and quantitative enzyme-linked immunosorbent assay. A single cation-exchange chromatography sufficed to obtain a highly purified recombinant HPLRP1 after direct batch adsorption onto S-Sepharose of the HPLRP1 present in the culture medium, at pH 5.5. N-terminal sequencing and mass spectrometry analysis were carried out to monitor the production of the mature protein and to confirm that its signal peptide was properly processed.

  1. Constitutive expression of tert in thymocytes leads to increased incidence and dissemination of T-cell lymphoma in Lck-Tert mice.

    PubMed

    Canela, Andrés; Martín-Caballero, Juan; Flores, Juana M; Blasco, María A

    2004-05-01

    Here we describe a new mouse model with constitutive expression of the catalytic subunit of telomerase (Tert) targeted to thymocytes and peripheral T cells (Lck-Tert mice). Two independent Lck-Tert mouse lines showed higher incidences of spontaneous T-cell lymphoma than the corresponding age-matched wild-type controls, indicating that constitutive expression of Tert promotes lymphoma. Interestingly, T-cell lymphomas in Lck-Tert mice were more disseminated than those in wild-type controls and affected both lymphoid and nonlymphoid tissues, while nonlymphoid tissues were never affected with lymphoma in age-matched wild-type controls. Importantly, these roles of Tert constitutive expression in promoting tumor progression and dissemination were independent of the role of telomerase in telomere length maintenance, since telomere length distributions on a single-cell basis were identical in Lck-Tert and wild-type thymocytes. Finally, Tert constitutive expression did not interfere with telomere capping in Lck-Tert primary thymocytes, although it resulted in greater chromosomal instability upon gamma irradiation in Lck-Tert primary lymphocytes than in controls, suggesting that Tert overexpression may interfere with the cellular response to DNA damage.

  2. Slower skeletal muscle phenotypes are critical for constitutive expression of Hsp70 in overloaded rat plantaris muscle.

    PubMed

    O'Neill, David E T; Aubrey, F Kris; Zeldin, David A; Michel, Robin N; Noble, Earl G

    2006-03-01

    Heat shock protein 72 (Hsp70) is constitutively expressed in rat hindlimb muscles, reportedly in proportion to their content of type I myosin heavy chain. This distribution pattern has been suggested to result from the higher recruitment and activity of such muscles and/or a specific relationship between myosin phenotype and Hsp70 content. To differentiate between these possibilities, the fiber-specific distribution of Hsp70 was examined in male Sprague-Dawley rat plantaris under control conditions, following a fast-to-slow phenotypic shift in response to surgically induced overload (O) and in response to O when the phenotypic shift was prevented by 3,5,3'-triiodo-dl-thyronine administration. Constitutive expression of Hsp70 was restricted to type I and IIa fibers in plantaris from control rats, and this fiber-specific pattern of expression was maintained following O of up to 28 days, although Hsp70 content in the O muscle doubled. When O (for 40 days) of the plantaris was combined with 3,5,3'-triiodo-dl-thyronine administration, despite typical hypertrophy in the overloaded plantaris, prevention of the normal phenotypic transformation also blocked the increased expression of Hsp70 observed in euthyroid controls. Collectively, these data suggest that chronic changes in constitutive expression of Hsp70 with altered contractile activity appear critically dependent on fast-to-slow phenotypic remodeling.

  3. Constitutive expression of the active fragment of human vasostatin Vs30 in Pichia pastoris SMD1168H.

    PubMed

    Calderon-Salais, Sergio; Velazquez-Bernardino, Prisiliana; Balderas-Hernandez, Victor E; Barba de la Rosa, Ana P; De Leon-Rodriguez, Antonio

    2018-04-01

    Vasostatin 30 (Vs30) is an active fragment derived from the N-terminal region (135-164 aa) of human calreticulin and has the ability to inhibit angiogenesis. In this work, the expression of Vs30 was performed using a protease-deficient strain of the methylotrophic yeast Pichia pastoris. The vs30 gene was optimized for P. pastoris preferential codon usage and inserted into constitutive expression vector pGAPZαA. In addition, a plasmid with four copies of the expression cassette was obtained and transformed into P. pastoris. The flask fermentation conditions were: culture volume of 25 mL in 250 mL baffled flasks at 28 °C, pH 6 and harvest time of 48 h. Up to 21.07 mg/L Vs30 were attained and purified by ultrafiltration with a 30-kDa cut-off membrane and the recovery was 49.7%. Bioactivity of Vs30 was confirmed by the inhibition of cell proliferation, as well as the inhibition of the capillary-like structures formation of EA.hy926 cells in vitro. This work constitutes the first report on the expression of Vs30 in Pichia pastoris using a constitutive promoter and multi-copy approach such as strategies to improve the recombinant Vs30 expression. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Constitutional downregulation of SEMA5A expression in autism.

    PubMed

    Melin, M; Carlsson, B; Anckarsater, H; Rastam, M; Betancur, C; Isaksson, A; Gillberg, C; Dahl, N

    2006-01-01

    There is strong evidence for the importance of genetic factors in idiopathic autism. The results from independent twin and family studies suggest that the disorder is caused by the action of several genes, possibly acting epistatically. We have used cDNA microarray technology for the identification of constitutional changes in the gene expression profile associated with idiopathic autism. Samples were obtained and analyzed from 6 affected subjects belonging to multiplex autism families and from 6 healthy controls. We assessed the expression levels for approximately 7,700 genes by cDNA microarrays using mRNA derived from Epstein-Barr virus-transformed B lymphocytes. The microarray data were analyzed in order to identify up- or downregulation of specific genes. A common pattern with nine downregulated genes was identified among samples derived from individuals with autism when compared to controls. Four of these nine genes encode proteins involved in biological processes associated with brain function or the immune system, and are consequently considered as candidates for genes associated with autism. Quantitative real-time PCR confirms the downregulation of the gene encoding SEMA5A, a protein involved in axonal guidance. Epstein-Barr virus should be considered as a possible source for altered expression, but our consistent results make us suggest SEMA5A as a candidate gene in the etiology of idiopathic autism.

  5. Constitutional downregulation of SEMA5A expression in autism

    PubMed Central

    Melin, Malin; Carlsson, Birgit; Anckarsäter, Henrik; Rastam, Maria; Betancur, Catalina; Isaksson, Anders; Gillberg, Christopher; Dahl, Niklas

    2006-01-01

    There is strong evidence for the importance of genetic factors in idiopathic autism. The results from independent twin and family studies suggest that the disorder is caused by the action of several genes, possibly acting epistatically. We have used cDNA microarray technology for the identification of constitutional changes in the gene expression profile associated with idiopathic autism. Samples were obtained and analyzed from six affected subjects belonging to multiplex autism families and from six healthy controls. We assessed the expression levels for approximately 7,700 genes by cDNA microarrays using mRNA derived from Epstein Barr virus (EBV)-transformed B-lymphocytes. The microarray data was analyzed in order to identify up- or down-regulation of specific genes. A common pattern with nine down-regulated genes was identified among samples derived from individuals with autism when compared to controls. Four of these nine genes encode proteins involved in biological processes associated with brain function or the immune system, and are consequently considered as candidates for genes associated with autism. Quantitative realtime PCR confirms the down-regulation of the gene encoding SEMA5A, a protein involved in axonal guidance. EBV should be considered as a possible source for altered expression but our consistent results make us suggest SEMA5A a candidate gene in the etiology of idiopathic autism. PMID:17028446

  6. A synthetic promoter library for constitutive gene expression in Lactobacillus plantarum.

    PubMed

    Rud, Ida; Jensen, Peter Ruhdal; Naterstad, Kristine; Axelsson, Lars

    2006-04-01

    A synthetic promoter library (SPL) for Lactobacillus plantarum has been developed, which generalizes the approach for obtaining synthetic promoters. The consensus sequence, derived from rRNA promoters extracted from the L. plantarum WCFS1 genome, was kept constant, and the non-consensus sequences were randomized. Construction of the SPL was performed in a vector (pSIP409) previously developed for high-level, inducible gene expression in L. plantarum and Lactobacillus sakei. A wide range of promoter strengths was obtained with the approach, covering 3-4 logs of expression levels in small increments of activity. The SPL was evaluated for the ability to drive beta-glucuronidase (GusA) and aminopeptidase N (PepN) expression. Protein production from the synthetic promoters was constitutive, and the most potent promoters gave high protein production with levels comparable to those of native rRNA promoters, and production of PepN protein corresponding to approximately 10-15 % of the total cellular protein. High correlation was obtained between the activities of promoters when tested in L. sakei and L. plantarum, which indicates the potential of the SPL for other Lactobacillus species. The SPL enables fine-tuning of stable gene expression for various applications in L. plantarum.

  7. Constitutive expression of the nifA gene activates associative nitrogen fixation of Enterobacter gergoviae 57-7, an opportunistic endophytic diazotroph.

    PubMed

    An, Q; Dong, Y; Wang, W; Li, Y; Li, J

    2007-09-01

    This study was undertaken to investigate whether a nitrogen-fixing bacterium Enterobacter gergoviae 57-7, which was isolated from surface-sterilized maize (Zea mays L.) roots, can colonize in roots and whether constitutive expression of the nifA gene encoding the transcriptional activator of nitrogenase genes can activate nif gene expression in planta. Maize seedlings grown in an agar medium were inoculated with Ent. gergoviae strains containing the green fluorescent protein reporter gene. Root colonization and expression of the dinitrogenase reductase gene (nifH) by Ent. gergoviae were observed by confocal laser scanning microscopy. gfp-tagged Ent. gergoviae was observed to colonize predominantly in cortical aerenchyma of primary and lateral roots and in stellar parenchyma cells and xylem vessels of primary roots. In planta nifH :: gfp expression was not detected but after a constitutively expressed nifA gene was introduced into Ent. gergoviae. Enterobacter gergoviae 57-7 is an opportunistic endophyte because it can live in soil and colonize in maize roots in the gnotobiotic agar culture. In agreement with previous (15)N-dilution evidence that Ent. gergoviae 57-7 did not fix N(2) in association with maize in pots whereas a derivative E7 containing a constitutively expressed nifA gene promoted plant growth partly through associative nitrogen fixation, constitutive expression of the nifA gene can activate bacterial nif gene expression in planta. This study and our previous studies suggest that manipulation of the promoter of the nifA gene in a nitrogen-fixing bacterium having a high colonization competence is a practical and promising approach to achieve a stable associative nitrogen fixation for cereals.

  8. Constitutive androstane receptor activation evokes the expression of glycolytic genes

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yarushkin, Andrei A.; Kazantseva, Yuliya A.; Prokopyeva, Elena A.

    It is well-known that constitutive androstane receptor (CAR) activation by 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP) increases the liver-to-body weight ratio. CAR-mediated liver growth is correlated with increased expression of the pleiotropic transcription factor cMyc, which stimulates cell cycle regulatory genes and drives proliferating cells into S phase. Because glycolysis supports cell proliferation and cMyc is essential for the activation of glycolytic genes, we hypothesized that CAR-mediated up-regulation of cMyc in mouse livers might play a role in inducing the expression of glycolytic genes. The aim of the present study was to examine the effect of long-term CAR activation on glycolytic genes in amore » mouse model not subjected to metabolic stress. We demonstrated that long-term CAR activation by TCPOBOP increases expression of cMyc, which was correlated with reduced expression of gluconeogenic genes and up-regulation of glucose transporter, glycolytic and mitochondrial pyruvate metabolising genes. These changes in gene expression after TCPOBOP treatment were strongly correlated with changes in levels of glycolytic intermediates in mouse livers. Moreover, we demonstrated a significant positive regulatory effect of TCPOBOP-activated CAR on both mRNA and protein levels of Pkm2, a master regulator of glucose metabolism and cell proliferation. Thus, our findings provide evidence to support the conclusion that CAR activation initiates a transcriptional program that facilitates the coordinated metabolic activities required for cell proliferation. - Highlights: • CAR-mediated liver growth is correlated with increased expression of cMyc. • CAR activation increased the expression of glycolytic genes in mouse livers. • CAR activation increased the level of Pkm2 in mouse livers.« less

  9. Drought, Abscisic Acid and Transpiration Rate Effects on the Regulation of PIP Aquaporin Gene Expression and Abundance in Phaseolus vulgaris Plants

    PubMed Central

    AROCA, RICARDO; FERRANTE, ANTONIO; VERNIERI, PAOLO; CHRISPEELS, MAARTEN J.

    2006-01-01

    • Background and Aims Drought causes a decline of root hydraulic conductance, which aside from embolisms, is governed ultimately by aquaporins. Multiple factors probably regulate aquaporin expression, abundance and activity in leaf and root tissues during drought; among these are the leaf transpiration rate, leaf water status, abscisic acid (ABA) and soil water content. Here a study is made of how these factors could influence the response of aquaporin to drought. • Methods Three plasma membrane intrinsic proteins (PIPs) or aquaporins were cloned from Phaseolus vulgaris plants and their expression was analysed after 4 d of water deprivation and also 1 d after re-watering. The effects of ABA and of methotrexate (MTX), an inhibitor of stomatal opening, on gene expression and protein abundance were also analysed. Protein abundance was examined using antibodies against PIP1 and PIP2 aquaporins. At the same time, root hydraulic conductance (L), transpiration rate, leaf water status and ABA tissue concentration were measured. • Key Results None of the treatments (drought, ABA or MTX) changed the leaf water status or tissue ABA concentration. The three treatments caused a decline in the transpiration rate and raised PVPIP2;1 gene expression and PIP1 protein abundance in the leaves. In the roots, only the drought treatment raised the expression of the three PIP genes examined, while at the same time diminishing PIP2 protein abundance and L. On the other hand, ABA raised both root PIP1 protein abundance and L. • Conclusions The rise of PvPIP2;1 gene expression and PIP1 protein abundance in the leaves of P. vulgaris plants subjected to drought was correlated with a decline in the transpiration rate. At the same time, the increase in the expression of the three PIP genes examined caused by drought and the decline of PIP2 protein abundance in the root tissues were not correlated with any of the parameters measured. PMID:17028296

  10. Expression of a constitutively activated plasma membrane H+-ATPase in Nicotiana tabacum BY-2 cells results in cell expansion.

    PubMed

    Niczyj, Marta; Champagne, Antoine; Alam, Iftekhar; Nader, Joseph; Boutry, Marc

    2016-11-01

    Increased acidification of the external medium by an activated H + -ATPase results in cell expansion, in the absence of upstream activating signaling. The plasma membrane H + -ATPase couples ATP hydrolysis with proton transport outside the cell, and thus creates an electrochemical gradient, which energizes secondary transporters. According to the acid growth theory, this enzyme is also proposed to play a major role in cell expansion, by acidifying the external medium and so activating enzymes that are involved in cell wall-loosening. However, this theory is still debated. To challenge it, we made use of a plasma membrane H + -ATPase isoform from Nicotiana plumbaginifolia truncated from its C-terminal auto-inhibitory domain (ΔCPMA4), and thus constitutively activated. This protein was expressed in Nicotiana tabacum BY-2 suspension cells using a heat shock inducible promoter. The characterization of several independent transgenic lines showed that the expression of activated ΔCPMA4 resulted in a reduced external pH by 0.3-1.2 units, as well as in an increased H + -ATPase activity by 77-155 % (ATP hydrolysis), or 70-306 % (proton pumping) of isolated plasma membranes. In addition, ΔCPMA4-expressing cells were 17-57 % larger than the wild-type cells and displayed abnormal shapes. A proteomic comparison of plasma membranes isolated from ΔCPMA4-expressing and wild-type cells revealed the altered abundance of several proteins involved in cell wall synthesis, transport, and signal transduction. In conclusion, the data obtained in this work showed that H + -ATPase activation is sufficient to induce cell expansion and identified possible actors which intervene in this process.

  11. Seeing red; the development of pON.mCherry, a broad-host range constitutive expression plasmid for Gram-negative bacteria.

    PubMed

    Gebhardt, Michael J; Jacobson, Rachael K; Shuman, Howard A

    2017-01-01

    The development of plasmid-mediated gene expression control in bacteria revolutionized the field of bacteriology. Many of these expression control systems rely on the addition of small molecules, generally metabolites or non-metabolized analogs thereof, to the growth medium to induce expression of the genes of interest. The paradigmatic example of an expression control system is the lac system from Escherichia coli, which typically relies on the Ptac promoter and the Lac repressor, LacI. In many cases, however, constitutive gene expression is desired, and other experimental approaches require the coordinated control of multiple genes. While multiple systems have been developed for use in E. coli and its close relatives, the utility and/or functionality of these tools does not always translate to other species. For example, for the Gram-negative pathogen, Legionella pneumophila, a causative agent of Legionnaires' Disease, the aforementioned Ptac system represents the only well-established expression control system. In order to enhance the tools available to study bacterial gene expression in L. pneumophila, we developed a plasmid, pON.mCherry, which confers constitutive gene expression from a mutagenized LacI binding site. We demonstrate that pON.mCherry neither interferes with other plasmids harboring an intact LacI-Ptac expression system nor alters the growth of Legionella species during intracellular growth. Furthermore, the broad-host range plasmid backbone of pON.mCherry allows constitutive gene expression in a wide variety of Gram-negative bacterial species, making pON.mCherry a useful tool for the greater research community.

  12. Effect of chromosome constitution variations on the expression of Turner phenotype.

    PubMed

    Bispo, A V S; Dos Santos, L O; Burégio-Frota, P; Galdino, M B; Duarte, A R; Leal, G F; Araújo, J; Gomes, B; Soares-Ventura, E M; Muniz, M T C; Santos, N

    2013-03-13

    Turner syndrome (TS) is a chronic disease related to haploinsufficiency of genes that are normally expressed in both X chromosomes in patients with female phenotype that is associated with a wide range of somatic malformations. We made detailed cytogenetic and clinical analysis of 65 patients with TS from the region of Recife, Brazil, to determine the effects of different chromosome constitutions on expression of the TS phenotype. Overall, patients with X-monosomy exhibited a tendency to have more severe phenotypes with higher morbidity, showing its importance in TS prognosis. Additionally, we found rare genetic and phenotypic abnormalities associated with this syndrome. To the best of our knowledge, this is the first case of 45,X,t(11;12)(q22;q22) described as a TS karyotype. Turner patients usually have normal intelligence; however, moderate to severe levels of mental retardation were found in 5 TS cases, which is considerate a very uncommon feature in this syndrome.

  13. Constitutive and inducible expression of SKALP/elafin provides anti-elastase defense in human epithelia.

    PubMed Central

    Pfundt, R; van Ruissen, F; van Vlijmen-Willems, I M; Alkemade, H A; Zeeuwen, P L; Jap, P H; Dijkman, H; Fransen, J; Croes, H; van Erp, P E; Schalkwijk, J

    1996-01-01

    Skin-derived antileukoproteinase (SKALP), also known as elafin, is a serine proteinase inhibitor first discovered in keratinocytes from hyperproliferative human epidermis. In addition to the proteinase inhibiting domain which is directed against polymorphonuclear leukocyte (PMN) derived enzymes such as elastase and proteinase 3, SKALP contains multiple transglutaminase (TGase) substrate domains which enable crosslinking to extracellular and cell envelope proteins. Here we show that SKALP is constitutively expressed in several epithelia that are continuously subjected to inflammatory stimuli, such as the oral cavity and the vagina where it co-localizes with type 1 TGase. All epithelia from sterile body cavities are negative for SKALP. In general, stratified squamous epithelia are positive, whereas pseudostratified epithelia, simple/glandular epithelia and normal epidermis are negative. SKALP was found in fetal tissues of the oral cavity from 17 wk gestation onwards where it continued to be expressed up to adult life. Remarkably, in fetal epidermis SKALP was found from week 28 onwards, but was downregulated to undetectable levels in neonatal skin within three months, suggesting a role during pregnancy in feto-maternal interactions or in the early maturation phase of the epidermis. Immunoelectron microscopy revealed the presence of SKALP in secretory vesicles including the lamellar granules. In culture models for epidermal keratinocytes we found that expression of the endogenous SKALP gene provided protection against cell detachment caused by purified elastase or activated PMNs. Addition of exogenous recombinant SKALP fully protected the keratinocytes against PMN-dependent detachment whereas superoxide dismutase and catalase were only marginally effective. These findings strongly suggest that the constitutive expression of SKALP in squamous epithelia, and the inducible expression in epidermis participate in the control of epithelial integrity, by inhibiting PMN

  14. Microbial Gene Abundance and Expression Patterns across a River to Ocean Salinity Gradient

    PubMed Central

    Fortunato, Caroline S.; Crump, Byron C.

    2015-01-01

    Microbial communities mediate the biogeochemical cycles that drive ecosystems, and it is important to understand how these communities are affected by changing environmental conditions, especially in complex coastal zones. As fresh and marine waters mix in estuaries and river plumes, the salinity, temperature, and nutrient gradients that are generated strongly influence bacterioplankton community structure, yet, a parallel change in functional diversity has not been described. Metagenomic and metatranscriptomic analyses were conducted on five water samples spanning the salinity gradient of the Columbia River coastal margin, including river, estuary, plume, and ocean, in August 2010. Samples were pre-filtered through 3 μm filters and collected on 0.2 μm filters, thus results were focused on changes among free-living microbial communities. Results from metagenomic 16S rRNA sequences showed taxonomically distinct bacterial communities in river, estuary, and coastal ocean. Despite the strong salinity gradient observed over sampling locations (0 to 33), the functional gene profiles in the metagenomes were very similar from river to ocean with an average similarity of 82%. The metatranscriptomes, however, had an average similarity of 31%. Although differences were few among the metagenomes, we observed a change from river to ocean in the abundance of genes encoding for catabolic pathways, osmoregulators, and metal transporters. Additionally, genes specifying both bacterial oxygenic and anoxygenic photosynthesis were abundant and expressed in the estuary and plume. Denitrification genes were found throughout the Columbia River coastal margin, and most highly expressed in the estuary. Across a river to ocean gradient, the free-living microbial community followed three different patterns of diversity: 1) the taxonomy of the community changed strongly with salinity, 2) metabolic potential was highly similar across samples, with few differences in functional gene abundance

  15. Microbial Gene Abundance and Expression Patterns across a River to Ocean Salinity Gradient.

    PubMed

    Fortunato, Caroline S; Crump, Byron C

    2015-01-01

    Microbial communities mediate the biogeochemical cycles that drive ecosystems, and it is important to understand how these communities are affected by changing environmental conditions, especially in complex coastal zones. As fresh and marine waters mix in estuaries and river plumes, the salinity, temperature, and nutrient gradients that are generated strongly influence bacterioplankton community structure, yet, a parallel change in functional diversity has not been described. Metagenomic and metatranscriptomic analyses were conducted on five water samples spanning the salinity gradient of the Columbia River coastal margin, including river, estuary, plume, and ocean, in August 2010. Samples were pre-filtered through 3 μm filters and collected on 0.2 μm filters, thus results were focused on changes among free-living microbial communities. Results from metagenomic 16S rRNA sequences showed taxonomically distinct bacterial communities in river, estuary, and coastal ocean. Despite the strong salinity gradient observed over sampling locations (0 to 33), the functional gene profiles in the metagenomes were very similar from river to ocean with an average similarity of 82%. The metatranscriptomes, however, had an average similarity of 31%. Although differences were few among the metagenomes, we observed a change from river to ocean in the abundance of genes encoding for catabolic pathways, osmoregulators, and metal transporters. Additionally, genes specifying both bacterial oxygenic and anoxygenic photosynthesis were abundant and expressed in the estuary and plume. Denitrification genes were found throughout the Columbia River coastal margin, and most highly expressed in the estuary. Across a river to ocean gradient, the free-living microbial community followed three different patterns of diversity: 1) the taxonomy of the community changed strongly with salinity, 2) metabolic potential was highly similar across samples, with few differences in functional gene abundance

  16. A constitutive expression system for glycosyl hydrolase family 7 cellobiohydrolases in Hypocrea jecorina

    DOE PAGES

    Linger, Jeffrey G.; Taylor, II, Larry E.; Baker, John O.; ...

    2015-03-18

    One of the primary industrial-scale cellulase producers is the ascomycete fungus, Hypocrea jecorina, which produces and secretes large quantities of diverse cellulolytic enzymes. Perhaps the single most important biomass degrading enzyme is cellobiohydrolase I (cbh1or Cel7A) due to its enzymatic proficiency in cellulose depolymerization. However, production of Cel7A with native-like properties from heterologous expression systems has proven difficult. In this study, we develop a protein expression system in H. jecorina (Trichoderma reesei) useful for production and secretion of heterologous cellobiohydrolases from glycosyl hydrolase family 7. Building upon previous work in heterologous protein expression in filamentous fungi, we have integrated amore » native constitutive enolase promoter with the native cbh1 signal sequence. The results are the following: The constitutive eno promoter driving the expression of Cel7A allows growth on glucose and results in repression of the native cellulase system, severely reducing background endo- and other cellulase activity and greatly simplifying purification of the recombinant protein. Coupling this system to a Δcbh1 strain of H. jecorina ensures that only the recombinant Cel7A protein is produced. Two distinct transformant colony morphologies were observed and correlated with high and null protein production. Production levels in ‘fast’ transformants are roughly equivalent to those in the native QM6a strain of H. jecorina, typically in the range of 10 to 30 mg/L when grown in continuous stirred-tank fermenters. ‘Slow’ transformants showed no evidence of Cel7A production. Specific activity of the purified recombinant Cel7A protein is equivalent to that of native protein when assayed on pretreated corn stover, as is the thermal stability and glycosylation level. Purified Cel7A produced from growth on glucose demonstrated remarkably consistent specific activity. Purified Cel7A from the same strain grown on lactose

  17. Multiple Genes Repress Motility in Uropathogenic Escherichia coli Constitutively Expressing Type 1 Fimbriae▿ †

    PubMed Central

    Simms, Amy N.; Mobley, Harry L. T.

    2008-01-01

    Two surface organelles of uropathogenic Escherichia coli (UPEC), flagella and type 1 fimbriae, are critical for colonization of the urinary tract but mediate opposite actions. Flagella propel bacteria through urine and along mucus layers, while type 1 fimbriae allow bacteria to adhere to specific receptors present on uroepithelial cells. Constitutive expression of type 1 fimbriae leads to repression of motility and chemotaxis in UPEC strain CFT073, suggesting that UPEC may coordinately regulate motility and adherence. To identify genes involved in this regulation of motility by type 1 fimbriae, transposon mutagenesis was performed on a phase-locked type 1 fimbrial ON variant of strain CFT073 (CFT073 fim L-ON), followed by a screen for restoration of motility in soft agar. Functions of the genes identified included attachment, metabolism, transport, DNA mismatch repair, and transcriptional regulation, and a number of genes had hypothetical function. Isogenic deletion mutants of these genes were also constructed in CFT073 fim L-ON. Motility was partially restored in six of these mutants, including complementable mutations in four genes encoding known transcriptional regulators, lrhA, lrp, slyA, and papX; a mismatch repair gene, mutS; and one hypothetical gene, ydiV. Type 1 fimbrial expression in these mutants was unaltered, and the majority of these mutants expressed larger amounts of flagellin than the fim L-ON parental strain. Our results indicate that repression of motility in CFT073 fim L-ON is not solely due to the constitutive expression of type 1 fimbriae on the surfaces of the bacteria and that multiple genes may contribute to this repression. PMID:18359812

  18. Proton-pumping rhodopsins are abundantly expressed by microbial eukaryotes in a high-Arctic fjord.

    PubMed

    Vader, Anna; Laughinghouse, Haywood D; Griffiths, Colin; Jakobsen, Kjetill S; Gabrielsen, Tove M

    2018-02-01

    Proton-pumping rhodopsins provide an alternative pathway to photosynthesis by which solar energy can enter the marine food web. Rhodopsin genes are widely found in marine bacteria, also in the Arctic, and were recently reported from several eukaryotic lineages. So far, little is known about rhodopsin expression in Arctic eukaryotes. In this study, we used metatranscriptomics and 18S rDNA tag sequencing to examine the mid-summer function and composition of marine protists (size 0.45-10 µm) in the high-Arctic Billefjorden (Spitsbergen), especially focussing on the expression of microbial proton-pumping rhodopsins. Rhodopsin transcripts were highly abundant, at a level similar to that of genes involved in photosynthesis. Phylogenetic analyses placed the environmental rhodopsins within disparate eukaryotic lineages, including dinoflagellates, stramenopiles, haptophytes and cryptophytes. Sequence comparison indicated the presence of several functional types, including xanthorhodopsins and a eukaryotic clade of proteorhodopsin. Transcripts belonging to the proteorhodopsin clade were also abundant in published metatranscriptomes from other oceanic regions, suggesting a global distribution. The diversity and abundance of rhodopsins show that these light-driven proton pumps play an important role in Arctic microbial eukaryotes. Understanding this role is imperative to predicting the future of the Arctic marine ecosystem faced by a changing light climate due to diminishing sea-ice. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  19. Plasmodium yoelii yoelii 17XNL constitutively expressing GFP throughout the life cycle.

    PubMed

    Ono, Takeshi; Tadakuma, Takushi; Rodriguez, Ana

    2007-03-01

    Plasmodium yoelii is a rodent parasite commonly used as a model to study malaria infection. It is the preferred model parasite for liver-stage immunological studies and is also widely used to study hepatocyte, erythrocyte and mosquito infection. We have generated a P. yoelii yoelii 17XNL line that is stably transfected with the green fluorescent protein (gfp) gene. This parasite line constitutively expresses high levels of GFP during the complete parasite life cycle including liver, blood and mosquito stages. These fluorescent parasites can be used in combination with fluorescence activated cell sorting or live microscopy for a wide range of experimental applications.

  20. Amending the Constitution. Teaching Strategy.

    ERIC Educational Resources Information Center

    Pittman, Keith A.

    1998-01-01

    Presents a lesson for secondary students where they learn about the historical forces that have shaped the U.S. Constitution through the amendment process, examine the constitutional amending process, and discuss the freedoms of expression guaranteed by the First Amendment. Includes three student handouts. (CMK)

  1. IFN-ε Is Constitutively Expressed by Cells of the Reproductive Tract and Is Inefficiently Secreted by Fibroblasts and Cell Lines

    PubMed Central

    Hermant, Pascale; Francius, Cédric; Clotman, Frédéric; Michiels, Thomas

    2013-01-01

    Type-I interferons (IFNs) form a large family of cytokines that primarily act to control the early development of viral infections. Typical type-I IFN genes, such as those encoding IFN-α or IFN-β are upregulated by viral infection in many cell types. In contrast, the gene encoding IFN-ε was reported to be constitutively expressed by cells of the female reproductive tract and to contribute to the protection against vaginal infections with herpes simplex virus 2 and Chlamydia muridarum. Our data confirm the lack of induction of IFN-ε expression after viral infection and the constitutive expression of IFN-ε by cells of the female but also of the male reproductive organs. Interestingly, when expressed from transfected expression plasmids in 293T, HeLa or Neuro2A cells, the mouse and human IFN-ε precursors were inefficiently processed and secretion of IFN-ε was minimal. Analysis of chimeric constructs produced between IFN-ε and limitin (IFN-ζ) showed that both the signal peptide and the mature moiety of IFN-ε contribute to poor processing of the precursor. Immunofluorescent detection of FLAG-tagged IFN-ε in transfected cells suggested that IFN-ε and chimeric proteins were defective for progression through the secretory pathway. IFN-ε did not, however, act intracellularly and impart an antiviral state to producing cells. Given the constitutive expression of IFN-ε in specialized cells and the poor processing of IFN-ε precursor in fibroblasts and cell lines, we hypothesize that IFN-ε secretion may require a co-factor specifically expressed in cells of the reproductive organs, that might secure the system against aberrant release of this IFN. PMID:23951133

  2. Induction of hepatocyte-like cells from mouse embryonic stem cells by lentivirus-mediated constitutive expression of Foxa2/Hnf4a.

    PubMed

    Liu, Tao; Zhang, Shichang; Xiang, Dedong; Wang, Yingjie

    2013-11-01

    Hepatocytes can be generated from embryonic stem cells (ESCs) using inducers such as chemical compounds and cytokines, but issues related to low differentiation efficiencies remain to be resolved. Recent work has shown that overexpression of lineage-specific transcription factors can directly cause cells phenotypic changes, including differentiation, trans-differentiation, and de-differentiation. We hypothesized that lentivirus-mediated constitutive expression of forkhead box A2 (Foxa2) and hepatocyte nuclear factor 4 alpha (Hnf4a) could promote inducing mouse ESCs to hepatocyte-likes cells. First, ESC lines that stably expressed Foxa2, Hnf4a, or Foxa2/Hnf4a were constructed via lentiviral expression vectors. Second, observations of cell morphology changes were made during the cell culture process, followed by experiments examining teratoma formation. Then, the effects of constitutive expression of Foxa2 and Hnf4a on hepatic differentiation and maturation were determined by measuring the marker gene expression levels of Albumin, α-fetoprotein, Cytokeratin18, and α1-antitrypsin. The results indicate that constitutive expression of Foxa2 and Hnf4a does not affect ESCs culture, teratoma formation, or the expression levels of the specific hepatocyte genes under autonomous differentiation. However, with some assistance from inducing factors, Foxa2 significantly increased the hepatic differentiation of ESCs, whereas the expression of Hnf4a alone or Foxa2/Hnf4a could not. Differentiated CCE-Foxa2 cells were more superior in expressing several liver-specific markers and protein, storing glycogen than differentiated CCE cells. Therefore, our method employing the transduction of Foxa2 would be a valuable tool for the efficient generation of functional hepatocytes derived from ESCs. © 2013 Wiley Periodicals, Inc.

  3. Abundant Gene-by-Environment Interactions in Gene Expression Reaction Norms to Copper within Saccharomyces cerevisiae

    PubMed Central

    Hodgins-Davis, Andrea; Adomas, Aleksandra B.; Warringer, Jonas; Townsend, Jeffrey P.

    2012-01-01

    Genetic variation for plastic phenotypes potentially contributes phenotypic variation to populations that can be selected during adaptation to novel ecological contexts. However, the basis and extent of plastic variation that manifests in diverse environments remains elusive. Here, we characterize copper reaction norms for mRNA abundance among five Saccharomyces cerevisiae strains to 1) describe population variation across the full range of ecologically relevant copper concentrations, from starvation to toxicity, and 2) to test the hypothesis that plastic networks exhibit increased population variation for gene expression. We find that although the vast majority of the variation is small in magnitude (considerably <2-fold), not just some, but most genes demonstrate variable expression across environments, across genetic backgrounds, or both. Plastically expressed genes included both genes regulated directly by copper-binding transcription factors Mac1 and Ace1 and genes indirectly responding to the downstream metabolic consequences of the copper gradient, particularly genes involved in copper, iron, and sulfur homeostasis. Copper-regulated gene networks exhibited more similar behavior within the population in environments where those networks have a large impact on fitness. Nevertheless, expression variation in genes like Cup1, important to surviving copper stress, was linked with variation in mitotic fitness and in the breadth of differential expression across the genome. By revealing a broader and deeper range of population variation, our results provide further evidence for the interconnectedness of genome-wide mRNA levels, their dependence on environmental context and genetic background, and the abundance of variation in gene expression that can contribute to future evolution. PMID:23019066

  4. Diversity, abundance, and consistency of microbial oxygenase expression and biodegradation in a shallow contaminated aquifer

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yagi, J.M.; Madsen, E.L.

    The diversity of Rieske dioxygenase genes and short-term temporal variability in the abundance of two selected dioxygenase gene sequences were examined in a naphthalene-rich, coal tar waste-contaminated subsurface study site. Using a previously published PCR-based approach (S. M. Ni Chadhain, R. S. Norman, K. V. Pesce, J. J. Kukor, and G. J. Zylstra, Appl. Environ. Microbiol. 72: 4078-4087, 2006) a broad suite of genes was detected, ranging from dioxygenase sequences associated with Rhodococcus and Sphingomonas to 32 previously uncharacterized Rieske gene sequence clone groups. The nag genes appeared frequently (20% of the total) in two groundwater monitoring wells characterized bymore » low (similar to 10{sup 2} ppb; similar to 1 {mu} M) ambient concentrations of naphthalene. A quantitative competitive PCR assay was used to show that abundances of nag genes (and archetypal nah genes) fluctuated substantially over a 9-month period. To contrast short-term variation with long-term community stability, in situ community gene expression (dioxygenase mRNA) and biodegradation potential (community metabolism of naphthalene in microcosms) were compared to measurements from 6 years earlier. cDNA sequences amplified from total RNA extracts revealed that nah- and nag-type genes were expressed in situ, corresponding well with structural gene abundances. Despite evidence for short-term (9-month) shifts in dioxygenase gene copy number, agreement in field gene expression (dioxygenase mRNA) and biodegradation potential was observed in comparisons to equivalent assays performed 6 years earlier. Thus, stability in community biodegradation characteristics at the hemidecadal time frame has been documented for these subsurface microbial communities.« less

  5. Construction and properties of a cell line constitutively expressing the herpes simplex virus glycoprotein B dependent on functional alpha 4 protein synthesis.

    PubMed Central

    Arsenakis, M; Hubenthal-Voss, J; Campadelli-Fiume, G; Pereira, L; Roizman, B

    1986-01-01

    We report the construction of a cell line constitutively expressing the glycoprotein B (gB) of herpes simplex virus (HSV) 1. The cell line was constructed in two steps. In the first, a baby hamster kidney cell line was transfected with the DNA of a plasmid containing the neomycin phosphotransferase gene that confers resistance to the antibiotic G418 and the gene specifying a temperature-sensitive (ts-) alpha 4 protein of HSV-1, the major viral regulatory protein. A clonal cell line, alpha 4/c113, selected for resistance to the antibiotic G418, expressed high levels of alpha 4 protein constitutively. Superinfection of these cells with HSV-2 resulted in twofold induction of the resident HSV-1 alpha 4 gene. In the second step, alpha 4/c113 cells were transfected with the DNA of a plasmid carrying the gB gene and the mouse methotrexate resistance dihydrofolate reductase gene. A clonal cell line, alpha 4/c113/gB, selected for methotrexate resistance expressed gB constitutively. Expression of both gB and alpha 4 continued unabated for at least 32 serial passages. Cells passaged serially in medium containing both methotrexate and G418 after passage 10 contained a higher copy number of the alpha 4 gene and produced larger amounts of both gB and alpha 4 proteins than did cells maintained in medium containing methotrexate alone. Expression of gB was dependent on the presence of functional alpha 4 protein inasmuch as expression of gB ceased on shift up to nonpermissive temperatures, when shifted to permissive temperatures, the cell line reinitiated expression of gB after a delay commensurate with the length of incubation at the nonpermissive temperature, and the cell-resident HSV-1 gB gene was expressed at the nonpermissive temperature in cells infected with a recombinant expressing a ts+ alpha 4 protein and an HSV-2 gB. The properties of the alpha 4/c113 cell line suggest that it may express other viral genes induced by alpha 4 protein constitutively, provided that the

  6. Constitutive expression of HCA(2) in human retina and primary human retinal pigment epithelial cells.

    PubMed

    Yu, Alice L; Birke, Kerstin; Lorenz, Reinhard L; Welge-Lussen, Ulrich

    2014-05-01

    HCA2, a receptor of β-hydroxybutyrate and niacin, has recently been described in mouse retina and immortalized human retinal pigment epithelial (RPE) cell lines. As HCA2 might be a pharmacologic target, e.g. in diabetic retinopathy, we studied its expression in human retina and primary human RPE cells. Paraffin sections of human retina and primary human RPE cells were obtained from human donor eyes. Expression of HCA2 in human retina was investigated by immunohistochemistry of paraffin sections and by RT-PCR. HCA2 expression in primary human RPE cells was examined by immunocytochemistry and by Western-blot analysis. Positive immunohistochemical staining for HCA2 was found in paraffin sections of human retina, and positive immunocytochemical staining for HCA2 in primary human RPE cells. RT-PCR analysis detected mRNA expression of HCA2 in human retina. The expression of HCA2 protein was found in primary human RPE cells. Based on these results, HCA2 appears to be constitutively expressed in human retina and in primary human RPE cells. Although its functional role is still unknown, HCA2 may be potentially involved in the pathogenesis of various retinopathies and may offer a new therapeutic target.

  7. Constitutive expression of cell wall invertase genes increases grain yield and starch content in maize.

    PubMed

    Li, Bei; Liu, Hua; Zhang, Yue; Kang, Tao; Zhang, Li; Tong, Jianhua; Xiao, Langtao; Zhang, Hongxia

    2013-12-01

    Grain size, number and starch content are important determinants of grain yield and quality. One of the most important biological processes that determine these components is the carbon partitioning during the early grain filling, which requires the function of cell wall invertase. Here, we showed the constitutive expression of cell wall invertase-encoding gene from Arabidopsis, rice (Oryza sativa) or maize (Zea mays), driven by the cauliflower mosaic virus (CaMV) 35S promoter, all increased cell wall invertase activities in different tissues and organs, including leaves and developing seeds, and substantially improved grain yield up to 145.3% in transgenic maize plants as compared to the wild-type plants, an effect that was reproduced in our 2-year field trials at different locations. The dramatically increased grain yield is due to the enlarged ears with both enhanced grain size and grain number. Constitutive expression of the invertase-encoding gene also increased total starch content up to 20% in the transgenic kernels. Our results suggest that cell wall invertase gene can be genetically engineered to improve both grain yield and grain quality in crop plants. © 2013 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  8. Constitutive expression of transgenes encoding derivatives of the synthetic antimicrobial peptide BP100: impact on rice host plant fitness.

    PubMed

    Nadal, Anna; Montero, Maria; Company, Nuri; Badosa, Esther; Messeguer, Joaquima; Montesinos, Laura; Montesinos, Emilio; Pla, Maria

    2012-09-04

    The Biopeptide BP100 is a synthetic and strongly cationic α-helical undecapeptide with high, specific antibacterial activity against economically important plant-pathogenic bacteria, and very low toxicity. It was selected from a library of synthetic peptides, along with other peptides with activities against relevant bacterial and fungal species. Expression of the BP100 series of peptides in plants is of major interest to establish disease-resistant plants and facilitate molecular farming. Specific challenges were the small length, peptide degradation by plant proteases and toxicity to the host plant. Here we approached the expression of the BP100 peptide series in plants using BP100 as a proof-of-concept. Our design considered up to three tandemly arranged BP100 units and peptide accumulation in the endoplasmic reticulum (ER), analyzing five BP100 derivatives. The ER retention sequence did not reduce the antimicrobial activity of chemically synthesized BP100 derivatives, making this strategy possible. Transformation with sequences encoding BP100 derivatives (bp100der) was over ten-fold less efficient than that of the hygromycin phosphotransferase (hptII) transgene. The BP100 direct tandems did not show higher antimicrobial activity than BP100, and genetically modified (GM) plants constitutively expressing them were not viable. In contrast, inverted repeats of BP100, whether or not elongated with a portion of a natural antimicrobial peptide (AMP), had higher antimicrobial activity, and fertile GM rice lines constitutively expressing bp100der were produced. These GM lines had increased resistance to the pathogens Dickeya chrysanthemi and Fusarium verticillioides, and tolerance to oxidative stress, with agronomic performance comparable to untransformed lines. Constitutive expression of transgenes encoding short cationic α-helical synthetic peptides can have a strong negative impact on rice fitness. However, GM plants expressing, for example, BP100 based on inverted

  9. The low noise limit in gene expression

    DOE PAGES

    Dar, Roy D.; Weinberger, Leor S.; Cox, Chris D.; ...

    2015-10-21

    Protein noise measurements are increasingly used to elucidate biophysical parameters. Unfortunately noise analyses are often at odds with directly measured parameters. Here we show that these inconsistencies arise from two problematic analytical choices: (i) the assumption that protein translation rate is invariant for different proteins of different abundances, which has inadvertently led to (ii) the assumption that a large constitutive extrinsic noise sets the low noise limit in gene expression. While growing evidence suggests that transcriptional bursting may set the low noise limit, variability in translational bursting has been largely ignored. We show that genome-wide systematic variation in translational efficiencymore » can-and in the case of E. coli does-control the low noise limit in gene expression. Therefore constitutive extrinsic noise is small and only plays a role in the absence of a systematic variation in translational efficiency. Lastly, these results show the existence of two distinct expression noise patterns: (1) a global noise floor uniformly imposed on all genes by expression bursting; and (2) high noise distributed to only a select group of genes.« less

  10. Conditional expression of constitutively active estrogen receptor {alpha} in chondrocytes impairs longitudinal bone growth in mice

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Ikeda, Kazuhiro; Tsukui, Tohru; Imazawa, Yukiko

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer Conditional transgenic mice expressing constitutively active estrogen receptor {alpha} (caER{alpha}) in chondrocytes were developed. Black-Right-Pointing-Pointer Expression of caER{alpha} in chondrocytes impaired longitudinal bone growth in mice. Black-Right-Pointing-Pointer caER{alpha} affects chondrocyte proliferation and differentiation. Black-Right-Pointing-Pointer This mouse model is useful for understanding the physiological role of ER{alpha}in vivo. -- Abstract: Estrogen plays important roles in the regulation of chondrocyte proliferation and differentiation, which are essential steps for longitudinal bone growth; however, the mechanisms of estrogen action on chondrocytes have not been fully elucidated. In the present study, we generated conditional transgenic mice, designated as caER{alpha}{sup ColII}, expressing constitutively activemore » mutant estrogen receptor (ER) {alpha} in chondrocytes, using the chondrocyte-specific type II collagen promoter-driven Cre transgenic mice. caER{alpha}{sup ColII} mice showed retardation in longitudinal growth, with short bone lengths. BrdU labeling showed reduced proliferation of hypertrophic chondrocytes in the proliferating layer of the growth plate of tibia in caER{alpha}{sup ColII} mice. In situ hybridization analysis of type X collagen revealed that the maturation of hypertrophic chondrocytes was impaired in caER{alpha}{sup ColII} mice. These results suggest that ER{alpha} is a critical regulator of chondrocyte proliferation and maturation during skeletal development, mediating longitudinal bone growth in vivo.« less

  11. Constitutive expression of transgenes encoding derivatives of the synthetic antimicrobial peptide BP100: impact on rice host plant fitness

    PubMed Central

    2012-01-01

    Background The Biopeptide BP100 is a synthetic and strongly cationic α-helical undecapeptide with high, specific antibacterial activity against economically important plant-pathogenic bacteria, and very low toxicity. It was selected from a library of synthetic peptides, along with other peptides with activities against relevant bacterial and fungal species. Expression of the BP100 series of peptides in plants is of major interest to establish disease-resistant plants and facilitate molecular farming. Specific challenges were the small length, peptide degradation by plant proteases and toxicity to the host plant. Here we approached the expression of the BP100 peptide series in plants using BP100 as a proof-of-concept. Results Our design considered up to three tandemly arranged BP100 units and peptide accumulation in the endoplasmic reticulum (ER), analyzing five BP100 derivatives. The ER retention sequence did not reduce the antimicrobial activity of chemically synthesized BP100 derivatives, making this strategy possible. Transformation with sequences encoding BP100 derivatives (bp100der) was over ten-fold less efficient than that of the hygromycin phosphotransferase (hptII) transgene. The BP100 direct tandems did not show higher antimicrobial activity than BP100, and genetically modified (GM) plants constitutively expressing them were not viable. In contrast, inverted repeats of BP100, whether or not elongated with a portion of a natural antimicrobial peptide (AMP), had higher antimicrobial activity, and fertile GM rice lines constitutively expressing bp100der were produced. These GM lines had increased resistance to the pathogens Dickeya chrysanthemi and Fusarium verticillioides, and tolerance to oxidative stress, with agronomic performance comparable to untransformed lines. Conclusions Constitutive expression of transgenes encoding short cationic α-helical synthetic peptides can have a strong negative impact on rice fitness. However, GM plants expressing, for

  12. Identification of Abundantly Expressed Novel and Conserved Genes from the Infective Larval Stage of Toxocara canis by an Expressed Sequence Tag Strategy

    PubMed Central

    Tetteh, Kevin K. A.; Loukas, Alex; Tripp, Cindy; Maizels, Rick M.

    1999-01-01

    Larvae of Toxocara canis, a nematode parasite of dogs, infect humans, causing visceral and ocular larva migrans. In noncanid hosts, larvae neither grow nor differentiate but endure in a state of arrested development. Reasoning that parasite protein production is orientated to immune evasion, we undertook a random sequencing project from a larval cDNA library to characterize the most highly expressed transcripts. In all, 266 clones were sequenced, most from both 3′ and 5′ ends, and similarity searches against GenBank protein and dbEST nucleotide databases were conducted. Cluster analyses showed that 128 distinct gene products had been found, all but 3 of which represented newly identified genes. Ninety-five genes were represented by a single clone, but seven transcripts were present at high frequencies, each composing >2% of all clones sequenced. These high-abundance transcripts include a mucin and a C-type lectin, which are both major excretory-secretory antigens released by parasites. Four highly expressed novel gene transcripts, termed ant (abundant novel transcript) genes, were found. Together, these four genes comprised 18% of all cDNA clones isolated, but no similar sequences occur in the Caenorhabditis elegans genome. While the coding regions of the four genes are dissimilar, their 3′ untranslated tracts have significant homology in nucleotide sequence. The discovery of these abundant, parasite-specific genes of newly identified lectins and mucins, as well as a range of conserved and novel proteins, provides defined candidates for future analysis of the molecular basis of immune evasion by T. canis. PMID:10456930

  13. Highly abundant and stage-specific mRNAs in the obligate pathogen Bremia lactucae.

    PubMed

    Judelson, H S; Michelmore, R W

    1990-01-01

    Germinating spores of the obligate pathogen Bremia lactucae (lettuce downy mildew) contain several unusually abundant species of mRNA. Thirty-nine cDNA clones corresponding to prevalent transcripts were isolated from a library synthesized using poly(A)+ RNA from germinating spores; these clones represented only five distinct classes. Each corresponding mRNA accounted for from 0.4 to 9 percent by mass of poly(A)+ RNA from germinating spores and together represented greater than 20 percent of the mRNA. The expression of the corresponding genes, and a gene encoding Hsp70, was analyzed in spores during germination and during growth in planta. The Hsp70 mRNA and mRNA from one abundant cDNA clone (ham34) were expressed constitutively. Two clones (ham9 and ham12) hybridized only to mRNA from spores and germinating spores. Two clones (ham37 and ham27) showed hybridization specific to germinating spores. Quantification of the number of genes homologous to each cDNA clone indicated that four clones corresponded to one or two copies per haploid genome, and one hybridized to an approximately 11-member family of genes. A sequence of the gene corresponding to ham34 was obtained to investigate its function and to identify sequences conferring high levels of gene expression for use in constructing vectors for the transformation of B. lactucae.

  14. Constitutive IDO expression in human cancer is sustained by an autocrine signaling loop involving IL-6, STAT3 and the AHR

    PubMed Central

    Sahm, Felix; Rauschenbach, Katharina J.; Trump, Saskia; Winter, Marcus; Ott, Martina; Ochs, Katharina; Lutz, Christian; Liu, Xiangdong; Anastasov, Natasa; Lehmann, Irina; Höfer, Thomas; von Deimling, Andreas; Wick, Wolfgang; Platten, Michael

    2014-01-01

    Indoleamine-2,3-dioxygenase (IDO) inhibitors have entered clinical trials based on their ability to restore anti-tumor immunity in preclinical studies. However, the mechanisms leading to constitutive expression of IDO in human tumors are largely unknown. Here we analyzed the pathways mediating constitutive IDO expression in human cancer. IDO-positive tumor cells and tissues showed basal phosphorylation and acetylation of STAT3 as evidenced by western blotting and immunoprecipitation. Inhibition of IL-6 or STAT3 using siRNA and/or pharmacological inhibitors reduced IDO mRNA and protein expression as well as kynurenine formation. In turn, IDO enzymatic activity activated the AHR as shown by the induction of AHR target genes. IDO-mediated AHR activation induced IL-6 expression, while inhibition or knockdown of the AHR reduced IL-6 expression. IDO activity thus sustains its own expression via an autocrine AHR–IL-6–STAT3 signaling loop. Inhibition of the AHR–IL-6–STAT3 signaling loop restored T-cell proliferation in mixed leukocyte reactions performed in the presence of IDO-expressing human cancer cells. Identification of the IDO-AHR-IL-6-STAT3 signaling loop maintaining IDO expression in human cancers reveals novel therapeutic targets for the inhibition of this core pathway promoting immunosuppression of human cancers. The relevance of the IDO-AHR-IL-6-STAT3 transcriptional circuit is underscored by the finding that high expression of its members IDO, STAT3 and the AHR target gene CYP1B1 is associated with reduced relapse-free survival in lung cancer patients. PMID:24657910

  15. Constitutive Activity among Orphan Class-A G Protein Coupled Receptors.

    PubMed

    Martin, Adam L; Steurer, Michael A; Aronstam, Robert S

    2015-01-01

    The purpose of this study was to evaluate the extent of constitutive activity among orphan class-A G protein coupled receptors within the cAMP signaling pathway. Constitutive signaling was revealed by changes in gene expression under control of the cAMP response element. Gene expression was measured in Chinese hamster ovary cells transiently co-transfected with plasmids containing a luciferase reporter and orphan receptor. Criteria adopted for defining constitutive activation were: 1) 200% elevation over baseline reporter gene expression; 2) 40% inhibition of baseline expression; and 3) 40% inhibition of expression stimulated by 3 μM forskolin. Five patterns of activity were noted: 1) inhibition under both baseline and forskolin stimulated expression (GPR15, GPR17, GPR18, GPR20, GPR25, GPR27, GPR31, GPR32, GPR45, GPR57, GPR68, GPR83, GPR84, GPR132, GPR150, GPR176); 2) no effect on baseline expression, but inhibition of forskolin stimulated expression (GPR4, GPR26, GPR61, GPR62, GPR78, GPR101, GPR119); 3) elevation of baseline signaling coupled with inhibition of forskolin stimulated expression (GPR6, GPR12); 4) elevation of baseline signaling without inhibition of forskolin stimulated expression (GPR3, GPR21, GPR52, GPR65); and 5) no effect on expression (GPR1, GPR19, GPR22, GPR34, GPR35, GPR39, GPR63, GPR82, GPR85, GPR87). Constitutive activity was observed in 75% of the orphan class-A receptors examined (30 of 40). This constitutive signaling cannot be explained by simple overexpression of the receptor. Inhibition of cAMP mediated expression was far more common (65%) than stimulation of expression (15%). Orphan receptors that were closely related based on amino acid homology tended to have similar effects on gene expression. These results suggest that identification of inverse agonists may be a fruitful approach for categorizing these orphan receptors and targeting them for pharmacological intervention.

  16. Constitutive Activity among Orphan Class-A G Protein Coupled Receptors

    PubMed Central

    Martin, Adam L.; Steurer, Michael A.; Aronstam, Robert S.

    2015-01-01

    The purpose of this study was to evaluate the extent of constitutive activity among orphan class-A G protein coupled receptors within the cAMP signaling pathway. Constitutive signaling was revealed by changes in gene expression under control of the cAMP response element. Gene expression was measured in Chinese hamster ovary cells transiently co-transfected with plasmids containing a luciferase reporter and orphan receptor. Criteria adopted for defining constitutive activation were: 1) 200% elevation over baseline reporter gene expression; 2) 40% inhibition of baseline expression; and 3) 40% inhibition of expression stimulated by 3 μM forskolin. Five patterns of activity were noted: 1) inhibition under both baseline and forskolin stimulated expression (GPR15, GPR17, GPR18, GPR20, GPR25, GPR27, GPR31, GPR32, GPR45, GPR57, GPR68, GPR83, GPR84, GPR132, GPR150, GPR176); 2) no effect on baseline expression, but inhibition of forskolin stimulated expression (GPR4, GPR26, GPR61, GPR62, GPR78, GPR101, GPR119); 3) elevation of baseline signaling coupled with inhibition of forskolin stimulated expression (GPR6, GPR12); 4) elevation of baseline signaling without inhibition of forskolin stimulated expression (GPR3, GPR21, GPR52, GPR65); and 5) no effect on expression (GPR1, GPR19, GPR22, GPR34, GPR35, GPR39, GPR63, GPR82, GPR85, GPR87). Constitutive activity was observed in 75% of the orphan class-A receptors examined (30 of 40). This constitutive signaling cannot be explained by simple overexpression of the receptor. Inhibition of cAMP mediated expression was far more common (65%) than stimulation of expression (15%). Orphan receptors that were closely related based on amino acid homology tended to have similar effects on gene expression. These results suggest that identification of inverse agonists may be a fruitful approach for categorizing these orphan receptors and targeting them for pharmacological intervention. PMID:26384023

  17. Scavenger receptor B1, the HDL receptor, is expressed abundantly in liver sinusoidal endothelial cells

    PubMed Central

    Ganesan, Latha P.; Mates, Jessica M.; Cheplowitz, Alana M.; Avila, Christina L.; Zimmerer, Jason M.; Yao, Zhili; Maiseyeu, Andrei; Rajaram, Murugesan V. S.; Robinson, John M.; Anderson, Clark L.

    2016-01-01

    Cholesterol from peripheral tissue, carried by HDL, is metabolized in the liver after uptake by the HDL receptor, SR-B1. Hepatocytes have long been considered the only liver cells expressing SR-B1; however, in this study we describe two disparate immunofluorescence (IF) experiments that suggest otherwise. Using high-resolution confocal microscopy employing ultrathin (120 nm) sections of mouse liver, improving z-axis resolution, we identified the liver sinusoidal endothelial cells (LSEC), marked by FcγRIIb, as the cell within the liver expressing abundant SR-B1. In contrast, the hepatocyte, identified with β-catenin, expressed considerably weaker levels, although optical resolution of SR-B1 was inadequate. Thus, we moved to a different IF strategy, first separating dissociated liver cells by gradient centrifugation into two portions, hepatocytes (parenchymal cells) and LSEC (non-parenchymal cells). Characterizing both portions for the cellular expression of SR-B1 by flow cytometry, we found that LSEC expressed considerable amounts of SR-B1 while in hepatocytes SR-B1 expression was barely perceptible. Assessing mRNA of SR-B1 by real time PCR we found messenger expression in LSEC to be about 5 times higher than in hepatocytes. PMID:26865459

  18. Biallelic MLH1 SNP cDNA expression or constitutional promoter methylation can hide genomic rearrangements causing Lynch syndrome.

    PubMed

    Morak, Monika; Koehler, Udo; Schackert, Hans Konrad; Steinke, Verena; Royer-Pokora, Brigitte; Schulmann, Karsten; Kloor, Matthias; Höchter, Wilhelm; Weingart, Josef; Keiling, Cortina; Massdorf, Trisari; Holinski-Feder, Elke

    2011-08-01

    A positive family history, germline mutations in DNA mismatch repair genes, tumours with high microsatellite instability, and loss of mismatch repair protein expression are the hallmarks of hereditary non-polyposis colorectal cancer (Lynch syndrome). However, in ~10-15% of cases of suspected Lynch syndrome, no disease-causing mechanism can be detected. Oligo array analysis was performed to search for genomic imbalances in patients with suspected mutation-negative Lynch syndrome with MLH1 deficiency in their colorectal tumours. A deletion in the LRRFIP2 (leucine-rich repeat flightless-interacting protein 2) gene flanking the MLH1 gene was detected, which turned out to be a paracentric inversion on chromosome 3p22.2 creating two new stable fusion transcripts between MLH1 and LRRFIP2. A single-nucleotide polymorphism in MLH1 exon 8 was expressed from both alleles, initially pointing to appropriate MLH1 function at least in peripheral cells. In a second case, an inherited duplication of the MLH1 gene region resulted in constitutional MLH1 promoter methylation. Constitutional MLH1 promoter methylation may therefore in rare cases be a heritable disease mechanism and should not be overlooked in seemingly sporadic patients.

  19. Expression of endothelin-1 and constitutional nitric oxide synthase messenger RNA in saphenous vein endothelial cells exposed to arterial flow shear stress.

    PubMed

    Zhu, Z G; Li, H H; Zhang, B R

    1997-11-01

    It has long been speculated that increased blood flow shear stress might be one of the major factors affecting the patency of grafted saphenous vein in coronary artery bypass operations. The underlying cellular and molecular mechanisms for so-called "shear stress damage" have not yet been well elucidated. Endothelial cells harvested from human saphenous vein were cultured in vitro and then exposed to a high arterial level flow shear stress in the parallel flow chamber. The expression levels of endothelin-1 and constitutional nitric oxide synthase by the endothelial cells were evaluated semiquantitatively at the gene transcription (messenger RNA) level using reverse transcription polymerase chain reaction. After 7 hours of exposure to arterial level shear stress, the expression of constitutional nitric oxide synthase messenger RNA by saphenous vein endothelial cells was significantly reduced, whereas the expression of endothelin-1 messenger RNA was substantially increased. These changes were more predominant at 24 hours. Arterial level flow shear stress could cause important changes in the gene transcription level in saphenous vein endothelial cells within a short period of time. The functional alterations of saphenous vein endothelial cells, as manifested by the increased expression of endothelin-1 and decreased expression of nitric oxide synthase messenger RNA, might play a crucial role in the vein graft remodeling process.

  20. Defective interleukin-4/Stat6 activity correlates with increased constitutive expression of negative regulators SOCS-3, SOCS-7, and CISH in colon cancer cells.

    PubMed

    Liu, Xiao Hong; Xu, Shuang Bing; Yuan, Jia; Li, Ben Hui; Zhang, Yan; Yuan, Qin; Li, Pin Dong; Li, Feng; Zhang, Wen Jie

    2009-12-01

    Interleukin-4 (IL-4)-induced Stat6 activities (phenotypes) vary among human cancer cells, of which the HT-29 cell line carries an active Stat6(high) phenotype, while Caco-2 carries a defective Stat6(null) phenotype, respectively. Cancer cells with Stat6(high) show resistance to apoptosis and exaggerated metastasis, suggesting the clinical significance of Stat6 phenotypes. We previously showed that Stat6(high) HT-29 cells exhibited low constitutive expression of Stat6-negative regulators SOCS-1 and SHP-1 because of gene hypermethylation. This study further examined the constitutive expression of other closely related SOCS family numbers including SOCS-3, SOCS-5, SOCS-7, and CISH using RT-PCR. Similar to SOCS-1 and SHP-1, Stat6(high) HT-29 cells expressed low constitutive mRNA of SOCS-3, SOCS-7, and CISH than Stat6(null) Caco-2 cells. Interestingly, DNA demethylation using 5-aza-2'-deoxycytidine in HT-29 cells up-regulated mRNA expression of the above genes, indicating a hypermethylation status, which was confirmed by methylation-specific sequencing in selected SOCS-3 gene. Furthermore, defective Stat6(null) Caco-2 exhibited impaired phosphorylation of Stat6 after IL-4 stimulation by flow cytometry, in keeping with the notion of an over-performed negative regulation. The findings that IL-4/Stat6 phenotypes show differential expression of multiple negative regulators suggest a model that a collective force of powerful negative regulators, directly and indirectly, acts on Stat6 activation, which may result in differential Stat6 phenotypes.

  1. Establishment of Sf9 Transformants Constitutively Expressing PBAN Receptor Variants: Application to Functional Evaluation

    PubMed Central

    Lee, Jae Min; Hull, J. Joe; Kawai, Takeshi; Tsuneizumi, Kazuhide; Kurihara, Masaaki; Tanokura, Masaru; Nagata, Koji; Nagasawa, Hiromichi; Matsumoto, Shogo

    2012-01-01

    To facilitate further evaluation of pheromone biosynthesis activating neuropeptide receptor (PBANR) functionality and regulation, we generated cultured insect cell lines constitutively expressing green fluorescent protein chimeras of the recently identified Bombyx mori PBANR (BommoPBANR) and Pseudaletia separata PBANR (PsesePBANR) variants. Fluorescent chimeras included the BommoPBANR-A, -B, and -C variants and the PsesePBANR-B and -C variants. Cell lines expressing non-chimeric BommoPBANR-B and -C variants were also generated. Functional evaluation of these transformed cell lines using confocal laser microscopy revealed that a Rhodamine Red-labeled PBAN derivative (RR-C10PBANR2K) specifically co-localized with all of the respective PBANR variants at the plasma membrane. Near complete internalization of the fluorescent RR-C10PBANR2K ligand 30 min after binding was observed in all cell lines except those expressing the BommoPBANR-A variant, in which the ligand/receptor complex remained at the plasma membrane. Fluorescent Ca2+ imaging further showed that the BommoPBANR-A cell line exhibited drastically different Ca2+ mobilization kinetics at a number of RR-C10PBANR2K concentrations including 10 μM. These observations demonstrate a clear functional difference between the BommoPBANR-A variant and the BommoPBANR-B and -C variants in terms of receptor regulation and activation of downstream effector molecules. We also found that, contrary to previous reports, ligand-induced internalization of BommoPBANR-B and BommoPBANR-C in cell lines stably expressing these variants occurred in the absence of extracellular Ca2+. PMID:22654874

  2. [Learning and Memory Capacity and NMDA Receptor Expression in Shen Deficiency Constitution Rats].

    PubMed

    Sun, Yu-ru; Sun, Yao-guang; Zhang, Qi; Wang, Xiao-di; Wang, Xing; Sun, Li-jun

    2016-05-01

    To explore material bases and neurobiological mechanisms of "Shen storing will" by observing learning and memory capacities and N-methyl-D-aspartic acid (NMDA) receptor expressions in Shen deficiency constitution (SDC) rats. Totally 40 SD rats were randomly divided into the model group, the Zuogui Pill (ZP) group, the Yougui Pill (YP) group, the blank control group (consisting of normal pregnant rats), 10 in each group. SDC young rat model (inherent deficiency and postnatal malnutrition) was prepared by the classic way of "cat scaring rat". Medication started when they were scared by cat. Rats in the ZP group and the YP group were administered by gastrogavage with ZP suspension 0.1875 g/mL and YP suspension 0.0938 g/mL respectively. Equal volume of normal saline was administered to rats in the blank control group and the model group by gastrogavage. All medication was given once per day, 5 days in a week for 2 consecutive months. Learning and memory capacities were detected by Morris water maze test. Expressions of NMDA receptor subunits NR2A and NR2B in hippocamus were detected by immunohistochemical method. Compared with the blank control group, the latency period, total distance in Morris water maze test were longer in the model group (P < 0.05). All the aforesaid indices all decreased in the ZP group and the YP group, with statistical difference when compared with the model group (P < 0.05). The protein expressions of NR2A and NR2B in hippocamus were lower in the model group than in the blank control group (P < 0.05). But when compared with the model group, they were obviously higher in the ZP group and the YP group (P < 0.05). SDC rats had degenerated learning and memory capacities and lowered NMDA receptor expressions. ZP and YP could up-regulate learning and memory capacities and NMDA receptor expressions, thereby improving deterioration of brain functions in SDC rats.

  3. Identification of centrarchid hepcidins and evidence that 17β-estradiol disrupts constitutive expression of hepcidin-1 and inducible expression of hepcidin-2 in largemouth bass (Micropterus salmoides)

    USGS Publications Warehouse

    Robertson, L.S.; Iwanowicz, L.R.; Marranca, J.M.

    2009-01-01

    Hepcidin is a highly conserved antimicrobial peptide and iron-regulatory hormone. Here, we identify two hepcidin genes (hep-1 and hep-2) in largemouth bass (Micropterus salmoides) and smallmouth bass (Micropterus dolomieu). Hepcidin-1 contains a putative ATCUN metal-binding site in the amino-terminus that is missing in hepcidin-2, suggesting that hepcidin-1 may function as an iron-regulatory hormone. Both hepcidins are predominately expressed in the liver of largemouth bass, similar to other fish and mammals. Experimental exposure of pond-raised largemouth bass to 17β-estradiol and/or the bacteria Edwardsiella ictaluri led to distinct changes in expression of hep-1 and hep-2. Estradiol reduced the constitutive expression of hep-1 in the liver. Bacterial exposure induced expression of hep-2, suggesting that hepcidin-2 may have an antimicrobial function, and this induction was abolished by estradiol. To our knowledge, this is the first report of the regulation of hepcidin expression by estradiol in either fish or mammals.

  4. Identification of centrarchid hepcidins and evidence that 17beta-estradiol disrupts constitutive expression of hepcidin-1 and inducible expression of hepcidin-2 in largemouth bass (Micropterus salmoides).

    PubMed

    Robertson, Laura S; Iwanowicz, Luke R; Marranca, Jamie Marie

    2009-06-01

    Hepcidin is a highly conserved antimicrobial peptide and iron-regulatory hormone. Here, we identify two hepcidin genes (hep-1 and hep-2) in largemouth bass (Micropterus salmoides) and smallmouth bass (Micropterus dolomieu). Hepcidin-1 contains a putative ATCUN metal-binding site in the amino-terminus that is missing in hepcidin-2, suggesting that hepcidin-1 may function as an iron-regulatory hormone. Both hepcidins are predominately expressed in the liver of largemouth bass, similar to other fish and mammals. Experimental exposure of pond-raised largemouth bass to 17beta-estradiol and/or the bacteria Edwardsiella ictaluri led to distinct changes in expression of hep-1 and hep-2. Estradiol reduced the constitutive expression of hep-1 in the liver. Bacterial exposure induced expression of hep-2, suggesting that hepcidin-2 may have an antimicrobial function, and this induction was abolished by estradiol. To our knowledge, this is the first report of the regulation of hepcidin expression by estradiol in either fish or mammals.

  5. A molecular analysis of desiccation tolerance mechanisms in the anhydrobiotic nematode Panagrolaimus superbus using expressed sequenced tags

    PubMed Central

    2012-01-01

    Background Some organisms can survive extreme desiccation by entering into a state of suspended animation known as anhydrobiosis. Panagrolaimus superbus is a free-living anhydrobiotic nematode that can survive rapid environmental desiccation. The mechanisms that P. superbus uses to combat the potentially lethal effects of cellular dehydration may include the constitutive and inducible expression of protective molecules, along with behavioural and/or morphological adaptations that slow the rate of cellular water loss. In addition, inducible repair and revival programmes may also be required for successful rehydration and recovery from anhydrobiosis. Results To identify constitutively expressed candidate anhydrobiotic genes we obtained 9,216 ESTs from an unstressed mixed stage population of P. superbus. We derived 4,009 unigenes from these ESTs. These unigene annotations and sequences can be accessed at http://www.nematodes.org/nembase4/species_info.php?species=PSC. We manually annotated a set of 187 constitutively expressed candidate anhydrobiotic genes from P. superbus. Notable among those is a putative lineage expansion of the lea (late embryogenesis abundant) gene family. The most abundantly expressed sequence was a member of the nematode specific sxp/ral-2 family that is highly expressed in parasitic nematodes and secreted onto the surface of the nematodes' cuticles. There were 2,059 novel unigenes (51.7% of the total), 149 of which are predicted to encode intrinsically disordered proteins lacking a fixed tertiary structure. One unigene may encode an exo-β-1,3-glucanase (GHF5 family), most similar to a sequence from Phytophthora infestans. GHF5 enzymes have been reported from several species of plant parasitic nematodes, with horizontal gene transfer (HGT) from bacteria proposed to explain their evolutionary origin. This P. superbus sequence represents another possible HGT event within the Nematoda. The expression of five of the 19 putative stress response

  6. A constitutively expressed antifungal peptide protects Tenebrio molitor during a natural infection by the entomopathogenic fungus Beauveria bassiana.

    PubMed

    Maistrou, Sevasti; Paris, Véronique; Jensen, Annette B; Rolff, Jens; Meyling, Nicolai V; Zanchi, Caroline

    2018-09-01

    Antimicrobial peptides have been well studied in the context of bacterial infections. Antifungal peptides have received comparatively less attention. Fungal pathogens of insects and their hosts represent a unique opportunity to study host-pathogen interactions due to the million of years of co-evolution they share. In this study, we investigated role of a constitutively expressed thaumatin-like peptide with antifungal activity expressed by the mealworm beetle Tenebrio molitor, named Tenecin 3, during a natural infection with the entomopathogenic fungus Beauveria bassiana. We monitored the effect of the expression of Tenecin 3 on the survival of infected hosts as well as on the progression of the fungal infection inside the host. Finally, we tested the activity of Tenecin 3 against B. bassiana. These findings could help improving biocontrol strategies and help understanding the evolution of antifungal peptides as a defense mechanism. Copyright © 2018 Elsevier Ltd. All rights reserved.

  7. Regulation of the mouse Treacher Collins syndrome homolog (Tcof1) promoter through differential repression of constitutive expression.

    PubMed

    Shows, Kathryn H; Shiang, Rita

    2008-11-01

    Treacher Collins syndrome is an autosomal-dominant mandibulofacial dysostosis caused by haploinsufficiency of the TCOF1 gene product treacle. Mouse Tcof1 protein is approximately 61% identical and 71% similar to treacle, and heterozygous knockout of Tcof1 causes craniofacial malformation. Tcof1 expression is high in developing neural crest, but much lower in other tissues. To investigate this dual regulation, highly conserved regions upstream of TCOF1 homologs were tested through deletion and mutation reporter assays, and conserved predicted transcription factor binding sites were assessed through chromatin binding studies. Assays were performed in mouse P19 embryonic carcinoma cells and in HEK293 cells to determine differential activation in cell types at different stages of differentiation. Binding of Cebpb, Zfp161, and Sp1 transcription factors was specific to the Tcof1 regulatory region in P19 cells. The Zfp161 binding site demonstrated P19 cell-specific repression, while the Sp1/Sp3 candidate site demonstrated HEK293 cell-specific activation. Moreover, presence of c-myb and Zfp161 transcripts was specific to P19 cells. A minimal promoter fragment from -253 to +43 bp directs constitutive expression in both cell types, and dual regulation of Tcof1 appears to be through differential repression of this minimal promoter. The CpG island at the transcription start site remains unmethylated in P19 cells, 11.5 dpc mouse embryonic tissue, and adult mouse ear, which supports constitutive activation of the Tcof1 promoter.

  8. Utility of temporally distinct baculovirus promoters for constitutive and baculovirus-inducible transgene expression in transformed insect cells.

    PubMed

    Lin, Chi-Hung; Jarvis, Donald L

    2013-05-10

    Genetically transformed lepidopteran insect cell lines have biotechnological applications as constitutive recombinant protein production platforms and improved hosts for baculovirus-mediated recombinant protein production. Insect cell transformation is often accomplished with a DNA construct(s) encoding a foreign protein(s) under the transcriptional control of a baculovirus immediate early promoter, such as the ie1 promoter. However, the potential utility of increasingly stronger promoters from later baculovirus gene classes, such as delayed early (39K), late (p6.9), and very late (polh), has not been systematically assessed. Hence, we produced DNA constructs encoding secreted alkaline phosphatase (SEAP) under the transcriptional control of each of the four temporally distinct classes of baculovirus promoters, used them to transform insect cells, and compared the levels of SEAP RNA and protein production obtained before and after baculovirus infection. The ie1 construct was the only one that supported SEAP protein production by transformed insect cells prior to baculovirus infection, confirming that only immediate early promoters can be used to isolate transformed insect cells for constitutive recombinant protein production. However, baculovirus infection activated transgene expression by all four classes of baculovirus promoters. After infection, cells transformed with the very late (polh) and late (p6.9) promoter constructs produced the highest levels of SEAP RNA, but only low levels of SEAP protein. Conversely, cells transformed with the immediate early (ie1) and delayed early (39K) promoter constructs produced lower levels of RNA, but equal or higher levels of SEAP protein. Unexpectedly, the 39K promoter construct provided tightly regulated, baculovirus-inducible protein production at higher levels than the later promoter constructs. Thus, this study demonstrated the utility of the 39K promoter for insect cell engineering, particularly when one requires higher

  9. Constitutive Uncoupling of Pathways of Gene Expression That Control Growth and Differentiation in Myeloid Leukemia: A Model for the Origin and Progression of Malignancy

    NASA Astrophysics Data System (ADS)

    Sachs, Leo

    1980-10-01

    Chemical carcinogens and tumor promoters have pleiotropic effects. Tumor initiators can produce a variety of mutations and tumor promoters can regulate a variety of physiological molecules that control growth and differentiation. The appropriate mutation and the regulation of the appropriate molecules to induce cell growth can initiate and promote the sequence of changes required for transformation of normal cells into malignant cells. After this sequence of changes, some tumors can still be induced to revert with a high frequency from a malignant phenotype to a nonmalignant phenotype. Results obtained from analysis of regulation of growth and differentiation in normal and leukemic myeloid cells, the phenotypic reversion of malignancy by induction of normal differentiation in myeloid leukemia, and the blocks in differentiation-defective leukemic cell mutants have been used to propose a general model for the origin and progression of malignancy. The model states that malignancy originates by changing specific pathways of gene expression required for growth from inducible to constitutive in cells that can still be induced to differentiate normally by the physiological inducer of differentiation. The malignant cells, unlike the normal cells, then no longer require the physiological inducer for growth. This changes the requirements for growth and uncouples growth from differentiation. Constitutive expression of other specific pathways can uncouple other controls, which then causes blocks in differentiation and the further progression of malignancy. The existence of specific constitutive pathways of gene expression that uncouple controls in malignant cells can also explain the expression of fetal proteins, hormones, and some other specialized products of normal development in various types of tumors.

  10. Insight into small RNA abundance and expression in high- and low-temperature stress response using deep sequencing in Arabidopsis.

    PubMed

    Baev, Vesselin; Milev, Ivan; Naydenov, Mladen; Vachev, Tihomir; Apostolova, Elena; Mehterov, Nikolay; Gozmanva, Mariyana; Minkov, Georgi; Sablok, Gaurav; Yahubyan, Galina

    2014-11-01

    Small RNA profiling and assessing its dependence on changing environmental factors have expanded our understanding of the transcriptional and post-transcriptional regulation of plant stress responses. Insufficient data have been documented earlier to depict the profiling of small RNA classes in temperature-associated stress which has a wide implication for climate change biology. In the present study, we report a comparative assessment of the genome-wide profiling of small RNAs in Arabidopsis thaliana using two conditional responses, induced by high- and low-temperature. Genome-wide profiling of small RNAs revealed an abundance of 21 nt small RNAs at low temperature, while high temperature showed an abundance of 21 nt and 24 nt small RNAs. The two temperature treatments altered the expression of a specific subset of mature miRNAs and displayed differential expression of a number of miRNA isoforms (isomiRs). Comparative analysis demonstrated that a large number of protein-coding genes can give rise to differentially expressed small RNAs following temperature shifts. Low temperature caused accumulation of small RNAs, corresponding to the sense strand of a number of cold-responsive genes. In contrast, high temperature stimulated the production of small RNAs of both polarities from genes encoding functionally diverse proteins. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  11. Absolute quantification of microbial taxon abundances.

    PubMed

    Props, Ruben; Kerckhof, Frederiek-Maarten; Rubbens, Peter; De Vrieze, Jo; Hernandez Sanabria, Emma; Waegeman, Willem; Monsieurs, Pieter; Hammes, Frederik; Boon, Nico

    2017-02-01

    High-throughput amplicon sequencing has become a well-established approach for microbial community profiling. Correlating shifts in the relative abundances of bacterial taxa with environmental gradients is the goal of many microbiome surveys. As the abundances generated by this technology are semi-quantitative by definition, the observed dynamics may not accurately reflect those of the actual taxon densities. We combined the sequencing approach (16S rRNA gene) with robust single-cell enumeration technologies (flow cytometry) to quantify the absolute taxon abundances. A detailed longitudinal analysis of the absolute abundances resulted in distinct abundance profiles that were less ambiguous and expressed in units that can be directly compared across studies. We further provide evidence that the enrichment of taxa (increase in relative abundance) does not necessarily relate to the outgrowth of taxa (increase in absolute abundance). Our results highlight that both relative and absolute abundances should be considered for a comprehensive biological interpretation of microbiome surveys.

  12. Allergic constitution theory of Chinese medicine and its assessment criterion and related studies.

    PubMed

    Wang, Ji; Wang, Ting; Li, Ying-shuai; Li, Ling-ru; Zheng, Yan-fei; Wang, Qi

    2015-09-01

    Constitution factor plays an important role in the occurrence, development, and transformation of diseases. The occurrence of allergic diseases is mainly caused by the disorganized physiological function and suitability regulation of patients, except for their exposure to outside allergens. Moreover, it represents susceptibility and hypersensitivity to allergens. The current study expresses the concept of allergic constitution from the perspective of Chinese medicine (CM) and presents the criterion of allergic constitution. In addition, the distribution of allergic constitution in population, its factors, and its relation to health-related quality of life (HRQOL) were investigated. The HRQOL scores of allergic constitution were found to be lower than those of the Pinghe constitution. After making a study on the gene expression profile of allergic constitution, the characteristics of up-regulated or down-regulated genes were found. Finally, CM drug was researched and developed to improve allergic constitution. Based on clinical trials and animal experiments, CM is found to have good regulatory effects on allergic constitution.

  13. Understanding relationships among abundance, extirpation, and climate at ecoregional scales.

    PubMed

    Beever, Erik A; Dobrowski, S Z; Long, J; Mynsberge, A R; Piekielek, N B

    2013-07-01

    Recent research on mountain-dwelling species has illustrated changes in species distributional patterns in response to climate change. Abundance of a species will likely provide an earlier warning indicator of change than will occupancy, yet relationships between abundance and climatic factors have received less attention. We tested whether predictors of counts of American pikas (Ochotona princeps) during surveys from the Great Basin region in 1994-1999 and 2003-2008 differed between the two periods. Additionally, we tested whether various modeled aspects of ecohydrology better predicted relative density than did average annual precipitation, and whether risk of site-wide extirpation predicted subsequent population counts of pikas. We observed several patterns of change in pika abundance at range edges that likely constitute early warnings of distributional shifts. Predictors of pika abundance differed strongly between the survey periods, as did pika extirpation patterns previously reported from this region. Additionally, maximum snowpack and growing-season precipitation resulted in better-supported models than those using average annual precipitation, and constituted two of the top three predictors of pika density in the 2000s surveys (affecting pikas perhaps via vegetation). Unexpectedly, we found that extirpation risk positively predicted subsequent population size. Our results emphasize the need to clarify mechanisms underlying biotic responses to recent climate change at organism-relevant scales, to inform management and conservation strategies for species of concern.

  14. Understanding relationships among abundance, extirpation,and climate at ecoregional scales

    USGS Publications Warehouse

    Beever, Erik A.; Solomon Dubrowski,; ,; ,; J. Long,; ,; A. Mysnberge,; Piekielek, N. B.

    2014-01-01

    Recent research on mountain-dwelling species has illustrated changes in species’ distributional patterns in response to climate change. Abundance of a species will likely provide an earlier warning indicator of change than will occupancy, yet relationships between abundance and climatic factors have received less attention. We tested whether predictors of counts of American pikas (Ochotona princeps) during surveys from the Great Basin region in 1994–1999 and 2003–2008 differed between the two periods. Additionally, we tested whether various modeled aspects of ecohydrology better predicted relative density than did average annual precipitation, and whether risk of site-wide extirpation predicted subsequent population counts of pikas. We observed several patterns of change in pika abundance at range edges that likely constitute early warnings of distributional shifts. Predictors of pika abundance differed strongly between the survey periods, as did pika extirpation patterns previously reported from this region. Additionally, maximum snowpack and growing-season precipitation resulted in better-supported models than those using average annual precipitation, and constituted two of the top three predictors of pika density in the 2000s surveys (affecting pikas perhaps via vegetation). Unexpectedly, we found that extirpation risk positively predicted subsequent population size. Our results emphasize the need to clarify mechanisms underlying biotic responses to recent climate change at organism-relevant scales, to inform management and conservation strategies for species of concern.

  15. Differential requirements of two recA mutants for constitutive SOS expression in Escherichia coli K-12.

    PubMed

    Long, Jarukit Edward; Renzette, Nicholas; Centore, Richard C; Sandler, Steven J

    2008-01-01

    Repairing DNA damage begins with its detection and is often followed by elicitation of a cellular response. In E. coli, RecA polymerizes on ssDNA produced after DNA damage and induces the SOS Response. The RecA-DNA filament is an allosteric effector of LexA auto-proteolysis. LexA is the repressor of the SOS Response. Not all RecA-DNA filaments, however, lead to an SOS Response. Certain recA mutants express the SOS Response (recA(C)) in the absence of external DNA damage in log phase cells. Genetic analysis of two recA(C) mutants was used to determine the mechanism of constitutive SOS (SOS(C)) expression in a population of log phase cells using fluorescence of single cells carrying an SOS reporter system (sulAp-gfp). SOS(C) expression in recA4142 mutants was dependent on its initial level of transcription, recBCD, recFOR, recX, dinI, xthA and the type of medium in which the cells were grown. SOS(C) expression in recA730 mutants was affected by none of the mutations or conditions tested above. It is concluded that not all recA(C) alleles cause SOS(C) expression by the same mechanism. It is hypothesized that RecA4142 is loaded on to a double-strand end of DNA and that the RecA filament is stabilized by the presence of DinI and destabilized by RecX. RecFOR regulate the activity of RecX to destabilize the RecA filament. RecA730 causes SOS(C) expression by binding to ssDNA in a mechanism yet to be determined.

  16. Altered focal adhesion regulation correlates with cardiomyopathy in mice expressing constitutively active rac1

    PubMed Central

    Sussman, Mark A.; Welch, Sara; Walker, Angela; Klevitsky, Raisa; Hewett, Timothy E.; Price, Robert L.; Schaefer, Erik; Yager, Karen

    2000-01-01

    The ras family of small GTP-binding proteins exerts powerful effects upon cell structure and function. One member of this family, rac, induces actin cytoskeletal reorganization in nonmuscle cells and hypertrophic changes in cultured cardiomyocytes. To examine the effect of rac1 activation upon cardiac structure and function, transgenic mice were created that express constitutively activated rac1 specifically in the myocardium. Transgenic rac1 protein was expressed at levels comparable to endogenous rac levels, with activation of the rac1 signaling pathway resulting in two distinct cardiomyopathic phenotypes: a lethal dilated phenotype associated with neonatal activation of the transgene and a transient cardiac hypertrophy seen among juvenile mice that resolved with age. Neither phenotype showed myofibril disarray and hypertrophic hearts were hypercontractilein working heart analyses. The rac1 target p21-activated kinase translocated from a cytosolic to a cytoskeletal distribution, suggesting that rac1 activation was inducing focal adhesion reorganization. Corroborating results showed altered localizations of src in dilated cardiomyopathy and paxillin in both cardiomyopathic phenotypes. This study, the first examination of rac1-mediated cardiac effects in vivo, demonstrates that dilation and hypertrophy can share a common molecular origin and presents evidence that both timing and concurrent signaling from multiple pathways can influence cardiac remodeling. PMID:10749567

  17. Regulation of the Mouse Treacher Collins Syndrome Homolog (Tcof1) Promoter Through Differential Repression of Constitutive Expression

    PubMed Central

    Shiang, Rita

    2008-01-01

    Treacher Collins syndrome is an autosomal-dominant mandibulofacial dysostosis caused by haploinsufficiency of the TCOF1 gene product treacle. Mouse Tcof1 protein is approximately 61% identical and 71% similar to treacle, and heterozygous knockout of Tcof1 causes craniofacial malformation. Tcof1 expression is high in developing neural crest, but much lower in other tissues. To investigate this dual regulation, highly conserved regions upstream of TCOF1 homologs were tested through deletion and mutation reporter assays, and conserved predicted transcription factor binding sites were assessed through chromatin binding studies. Assays were performed in mouse P19 embryonic carcinoma cells and in HEK293 cells to determine differential activation in cell types at different stages of differentiation. Binding of Cebpb, Zfp161, and Sp1 transcription factors was specific to the Tcof1 regulatory region in P19 cells. The Zfp161 binding site demonstrated P19 cell–specific repression, while the Sp1/Sp3 candidate site demonstrated HEK293 cell–specific activation. Moreover, presence of c-myb and Zfp161 transcripts was specific to P19 cells. A minimal promoter fragment from −253 to +43 bp directs constitutive expression in both cell types, and dual regulation of Tcof1 appears to be through differential repression of this minimal promoter. The CpG island at the transcription start site remains unmethylated in P19 cells, 11.5 dpc mouse embryonic tissue, and adult mouse ear, which supports constitutive activation of the Tcof1 promoter. PMID:18771418

  18. Bayesian Inference of Allele-Specific Gene Expression Indicates Abundant Cis-Regulatory Variation in Natural Flycatcher Populations

    PubMed Central

    Wang, Mi

    2017-01-01

    Abstract Polymorphism in cis-regulatory sequences can lead to different levels of expression for the two alleles of a gene, providing a starting point for the evolution of gene expression. Little is known about the genome-wide abundance of genetic variation in gene regulation in natural populations but analysis of allele-specific expression (ASE) provides a means for investigating such variation. We performed RNA-seq of multiple tissues from population samples of two closely related flycatcher species and developed a Bayesian algorithm that maximizes data usage by borrowing information from the whole data set and combines several SNPs per transcript to detect ASE. Of 2,576 transcripts analyzed in collared flycatcher, ASE was detected in 185 (7.2%) and a similar frequency was seen in the pied flycatcher. Transcripts with statistically significant ASE commonly showed the major allele in >90% of the reads, reflecting that power was highest when expression was heavily biased toward one of the alleles. This would suggest that the observed frequencies of ASE likely are underestimates. The proportion of ASE transcripts varied among tissues, being lowest in testis and highest in muscle. Individuals often showed ASE of particular transcripts in more than one tissue (73.4%), consistent with a genetic basis for regulation of gene expression. The results suggest that genetic variation in regulatory sequences commonly affects gene expression in natural populations and that it provides a seedbed for phenotypic evolution via divergence in gene expression. PMID:28453623

  19. Measuring the expressed abundance of the three phases of water with an imaging spectrometer over melting snow

    NASA Astrophysics Data System (ADS)

    Green, Robert O.; Painter, Thomas H.; Roberts, Dar A.; Dozier, Jeff

    2006-10-01

    From imaging spectrometer data, we simultaneously estimate the abundance of the three phases of water in an environment that includes melting snow, basing the analysis on the spectral shift in the absorption coefficient between water vapor, liquid water, and ice at 940, 980, and 1030 nm respectively. We apply a spectral fitting algorithm that measures the expressed abundance of the three phases of water to a data set acquired by the Airborne Visible/Infrared Imaging Spectrometer (AVIRIS) over Mount Rainier, Washington, on 14 June 1996. Precipitable water vapor varies from 1 mm over the summit of Mount Rainier to 10 mm over the lower valleys to the northwest. Equivalent path absorption of liquid water varies from 0 to 13 mm, with the zero values over rocky areas and high-elevation snow and the high values associated with liquid water held in vegetation canopies and in melting snow. Ice abundance varies from 0 to 30 mm equivalent path absorption in the snow- and glacier-covered portions of Mount Rainier. The water and ice abundances are related to the amount of liquid water and the sizes of the ice grains in the near-surface layer. Precision of the estimates, calculated over locally homogeneous areas, indicates an uncertainty of better than 1.5% for all three phases, except for liquid water in vegetation, where an optimally homogeneous site was not found. The analysis supports new strategies for hydrological research and applications as imaging spectrometers become more available.

  20. Genome resequencing and transcriptome profiling reveal structural diversity and expression patterns of constitutive disease resistance genes in Huanglongbing-tolerant Poncirus trifoliata and its hybrids

    PubMed Central

    Rawat, Nidhi; Kumar, Brajendra; Albrecht, Ute; Du, Dongliang; Huang, Ming; Yu, Qibin; Zhang, Yi; Duan, Yong-Ping; Bowman, Kim D; Gmitter, Fred G; Deng, Zhanao

    2017-01-01

    Huanglongbing (HLB) is the most destructive bacterial disease of citrus worldwide. While most citrus varieties are susceptible to HLB, Poncirus trifoliata, a close relative of Citrus, and some of its hybrids with Citrus are tolerant to HLB. No specific HLB tolerance genes have been identified in P. trifoliata but recent studies have shown that constitutive disease resistance (CDR) genes were expressed at much higher levels in HLB-tolerant Poncirus hybrids and the expression of CDR genes was modulated by Candidatus Liberibacter asiaticus (CLas), the pathogen of HLB. The current study was undertaken to mine and characterize the CDR gene family in Citrus and Poncirus and to understand its association with HLB tolerance in Poncirus. We identified 17 CDR genes in two citrus genomes, deduced their structures, and investigated their phylogenetic relationships. We revealed that the expansion of the CDR family in Citrus seems to be due to segmental and tandem duplication events. Through genome resequencing and transcriptome sequencing, we identified eight CDR genes in the Poncirus genome (PtCDR1-PtCDR8). The number of SNPs was the highest in PtCDR2 and the lowest in PtCDR7. Most of the deletion and insertion events were observed in the UTR regions of Citrus and Poncirus CDR genes. PtCDR2 and PtCDR8 were in abundance in the leaf transcriptomes of two HLB-tolerant Poncirus genotypes and were also upregulated in HLB-tolerant, Poncirus hybrids as revealed by real-time PCR analysis. These two CDR genes seem to be good candidate genes for future studies of their role in citrus-CLas interactions. PMID:29152310

  1. Establishment of a quantitative PCR system for discriminating chitinase-like proteins: catalytically inactive breast regression protein-39 and Ym1 are constitutive genes in mouse lung.

    PubMed

    Ohno, Misa; Kida, Yuta; Sakaguchi, Masayoshi; Sugahara, Yasusato; Oyama, Fumitaka

    2014-10-08

    Mice and humans produce chitinase-like proteins (CLPs), which are highly homologous to chitinases but lack chitinolytic activity. Mice express primarily three CLPs, including breast regression protein-39 (BRP-39) [chitinase 3-like-1 (Chi3l1) or 38-kDa glycoprotein (gp38k)], Ym1 (Chi3l3) and Ym2 (Chi3l4). Recently, CLPs have attracted considerable attention due to their increased expression in a number of pathological conditions, including asthma, allergies, rheumatoid arthritis and malignant tumors. Although the exact functions of CLPs are largely unknown, the significance of their increased expression levels during pathophysiological states needs to be determined. The quantification of BRP-39, Ym1 and Ym2 is an important step in gaining insight into the in vivo regulation of the CLPs. We constructed a standard DNA for quantitative real-time PCR (qPCR) by containing three CLPs target fragments and five reference genes cDNA in a one-to-one ratio. We evaluated this system by analyzing the eight target cDNA sequences. Tissue cDNAs obtained by reverse transcription from total RNA from four embryonic stages and eight adult tissues were analyzed using the qPCR system with the standard DNA. We established a qPCR system detecting CLPs and comparing their expression levels with those of five reference genes using the same scale in mouse tissues. We found that BRP-39 and Ym1 were abundant in the mouse lung, whereas Ym2 mRNA was abundant in the stomach, followed by lung. The expression levels of BRP-39 and Ym1 in the mouse lung were higher than those of two active chitinases and were comparable to glyceraldehyde-3-phosphate dehydrogenase, a housekeeping gene which is constitutively expressed in all tissues. Our results indicate that catalytically inactive BRP-39 and Ym1 are constitutive genes in normal mouse lung.

  2. Patterns of rare and abundant marine microbial eukaryotes.

    PubMed

    Logares, Ramiro; Audic, Stéphane; Bass, David; Bittner, Lucie; Boutte, Christophe; Christen, Richard; Claverie, Jean-Michel; Decelle, Johan; Dolan, John R; Dunthorn, Micah; Edvardsen, Bente; Gobet, Angélique; Kooistra, Wiebe H C F; Mahé, Frédéric; Not, Fabrice; Ogata, Hiroyuki; Pawlowski, Jan; Pernice, Massimo C; Romac, Sarah; Shalchian-Tabrizi, Kamran; Simon, Nathalie; Stoeck, Thorsten; Santini, Sébastien; Siano, Raffaele; Wincker, Patrick; Zingone, Adriana; Richards, Thomas A; de Vargas, Colomban; Massana, Ramon

    2014-04-14

    Biological communities are normally composed of a few abundant and many rare species. This pattern is particularly prominent in microbial communities, in which most constituent taxa are usually extremely rare. Although abundant and rare subcommunities may present intrinsic characteristics that could be crucial for understanding community dynamics and ecosystem functioning, microbiologists normally do not differentiate between them. Here, we investigate abundant and rare subcommunities of marine microbial eukaryotes, a crucial group of organisms that remains among the least-explored biodiversity components of the biosphere. We surveyed surface waters of six separate coastal locations in Europe, independently considering the picoplankton, nanoplankton, and microplankton/mesoplankton organismal size fractions. Deep Illumina sequencing of the 18S rRNA indicated that the abundant regional community was mostly structured by organismal size fraction, whereas the rare regional community was mainly structured by geographic origin. However, some abundant and rare taxa presented similar biogeography, pointing to spatiotemporal structure in the rare microeukaryote biosphere. Abundant and rare subcommunities presented regular proportions across samples, indicating similar species-abundance distributions despite taxonomic compositional variation. Several taxa were abundant in one location and rare in other locations, suggesting large oscillations in abundance. The substantial amount of metabolically active lineages found in the rare biosphere suggests that this subcommunity constitutes a diversity reservoir that can respond rapidly to environmental change. We propose that marine planktonic microeukaryote assemblages incorporate dynamic and metabolically active abundant and rare subcommunities, with contrasting structuring patterns but fairly regular proportions, across space and time. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. Constitutive expression of tdTomato protein as a cytotoxicity and proliferation marker for space radiation biology

    NASA Astrophysics Data System (ADS)

    Chishti, Arif A.; Hellweg, Christine E.; Berger, Thomas; Baumstark-Khan, Christa; Feles, Sebastian; Kätzel, Thorben; Reitz, Günther

    2015-01-01

    The radiation risk assessment for long-term space missions requires knowledge on the biological effectiveness of different space radiation components, e.g. heavy ions, on the interaction of radiation and other space environmental factors such as microgravity, and on the physical and biological dose distribution in the human body. Space experiments and ground-based experiments at heavy ion accelerators require fast and reliable test systems with an easy readout for different endpoints. In order to determine the effect of different radiation qualities on cellular proliferation and the biological depth dose distribution after heavy ion exposure, a stable human cell line expressing a novel fluorescent protein was established and characterized. tdTomato, a red fluorescent protein of the new generation with fast maturation and high fluorescence intensity, was selected as reporter of cell proliferation. Human embryonic kidney (HEK/293) cells were stably transfected with a plasmid encoding tdTomato under the control of the constitutively active cytomegalovirus (CMV) promoter (ptdTomato-N1). The stably transfected cell line was named HEK-ptdTomato-N1 8. This cytotoxicity biosensor was tested by ionizing radiation (X-rays and accelerated heavy ions) exposure. As biological endpoints, the proliferation kinetics and the cell density reached 100 h after irradiation reflected by constitutive expression of the tdTomato were investigated. Both were reduced dose-dependently after radiation exposure. Finally, the cell line was used for biological weighting of heavy ions of different linear energy transfer (LET) as space-relevant radiation quality. The relative biological effectiveness of accelerated heavy ions in reducing cellular proliferation peaked at an LET of 91 keV/μm. The results of this study demonstrate that the HEK-ptdTomato-N1 reporter cell line can be used as a fast and reliable biosensor system for detection of cytotoxic damage caused by ionizing radiation.

  4. Constitutive Expression of Mammalian Nitric Oxide Synthase in Tobacco Plants Triggers Disease Resistance to Pathogens

    PubMed Central

    Chun, Hyun Jin; Park, Hyeong Cheol; Koo, Sung Cheol; Lee, Ju Huck; Park, Chan Young; Choi, Man Soo; Kang, Chang Ho; Baek, Dongwon; Cheong, Yong Hwa; Yun, Dae-Jin; Chung, Woo Sik; Cho, Moo Je; Kim, Min Chul

    2012-01-01

    Nitric oxide (NO) is known for its role in the activation of plant defense responses. To examine the involvement and mode of action of NO in plant defense responses, we introduced calmodulin-dependent mammalian neuronal nitric oxide synthase (nNOS), which controls the CaMV35S promoter, into wild-type and NahG tobacco plants. Constitutive expression of nNOS led to NO production and triggered spontaneous induction of leaf lesions. Transgenic plants accumulated high amounts of H2O2, with catalase activity lower than that in the wild type. nNOS transgenic plants contained high levels of salicylic acid (SA), and they induced an array of SA-, jasmonic acid (JA)-, and/or ethylene (ET)-related genes. Consequently, NahG co-expression blocked the induction of systemic acquired resistance (SAR)-associated genes in transgenic plants, implying SA is involved in NO-mediated induction of SAR genes. The transgenic plants exhibited enhanced resistance to a spectrum of pathogens, including bacteria, fungi, and viruses. Our results suggest a highly ranked regulatory role for NO in SA-, JA-, and/or ET-dependent pathways that lead to disease resistance. PMID:23124383

  5. Constitutive relations in optics in terms of geometric algebra

    NASA Astrophysics Data System (ADS)

    Dargys, A.

    2015-11-01

    To analyze the electromagnetic wave propagation in a medium the Maxwell equations should be supplemented by constitutive relations. At present the classification of linear constitutive relations is well established in tensorial-matrix and exterior p-form calculus. Here the constitutive relations are found in the context of Clifford geometric algebra. For this purpose Cl1,3 algebra that conforms with relativistic 4D Minkowskian spacetime is used. It is shown that the classification of linear optical phenomena with the help of constitutive relations in this case comes from the structure of Cl1,3 algebra itself. Concrete expressions for constitutive relations which follow from this algebra are presented. They can be applied in calculating the propagation properties of electromagnetic waves in any anisotropic, linear and nondissipative medium.

  6. Constitutive expression of active microbial transglutaminase in Escherichia coli and comparative characterization to a known variant.

    PubMed

    Javitt, Gabe; Ben-Barak-Zelas, Zohar; Jerabek-Willemsen, Moran; Fishman, Ayelet

    2017-02-28

    Microbial transglutaminase (mTG) is a robust enzyme catalyzing the formation of an isopeptide bond between glutamine and lysine residues. It has found use in food applications, pharmaceuticals, textiles, and biomedicine. Overexpression of soluble and active mTG in E. coli has been limited due to improper protein folding and requirement for proteolytic cleavage of the pro-domain. Furthermore, to integrate mTG more fully industrially and academically, thermostable and solvent-stable variants may be imperative. A novel expression system constitutively producing active mTG was designed. Wild-type (WT) mTG and a S2P variant had similar expression levels, comparable to previous studies. Kinetic constants were determined by a glutamate dehydrogenase-coupled assay, and the S2P variant showed an increased affinity and a doubled enzyme efficiency towards Z-Gln-Gly. The melting temperature (T m ) of the WT was determined by intrinsic fluorescence measurements to be 55.8 ± 0.1 °C and of the S2P variant to be 56.3 ± 0.4 °C and 45.5 ± 0.1 °C, showing a moderately different thermostability profile. Stability in water miscible organic solvents was determined for both the WT and S2P variant. Of the solvents tested, incubation of mTG in isopropanol for 24 h at 4 °C showed the strongest stabilizing effect with mTG retaining 61 and 72% activity for WT and S2P respectively in 70% isopropanol. Both enzymes also showed an increased initial activity in the presence of organic solvents with the highest activity increase in 40% DMSO. Nevertheless, both enzymes were inactivated in 70% of all organic solvents tested. A constitutive expression system of active mTG in E. coli without downstream proteolytic cleavage processing was used for overexpression and characterization. High throughput techniques for testing thermostability and kinetics were useful in streamlining analysis and could be used in the future for quickly identifying beneficial mutants. Hitherto untested

  7. Application of symbolic computations to the constitutive modeling of structural materials

    NASA Technical Reports Server (NTRS)

    Arnold, Steven M.; Tan, H. Q.; Dong, X.

    1990-01-01

    In applications involving elevated temperatures, the derivation of mathematical expressions (constitutive equations) describing the material behavior can be quite time consuming, involved and error-prone. Therefore intelligent application of symbolic systems to faciliate this tedious process can be of significant benefit. Presented here is a problem oriented, self contained symbolic expert system, named SDICE, which is capable of efficiently deriving potential based constitutive models in analytical form. This package, running under DOE MACSYMA, has the following features: (1) potential differentiation (chain rule), (2) tensor computations (utilizing index notation) including both algebraic and calculus; (3) efficient solution of sparse systems of equations; (4) automatic expression substitution and simplification; (5) back substitution of invariant and tensorial relations; (6) the ability to form the Jacobian and Hessian matrix; and (7) a relational data base. Limited aspects of invariant theory were also incorporated into SDICE due to the utilization of potentials as a starting point and the desire for these potentials to be frame invariant (objective). The uniqueness of SDICE resides in its ability to manipulate expressions in a general yet pre-defined order and simplify expressions so as to limit expression growth. Results are displayed, when applicable, utilizing index notation. SDICE was designed to aid and complement the human constitutive model developer. A number of examples are utilized to illustrate the various features contained within SDICE. It is expected that this symbolic package can and will provide a significant incentive to the development of new constitutive theories.

  8. Selection for low dormancy in annual ryegrass (Lolium rigidum) seeds results in high constitutive expression of a glucose-responsive α-amylase isoform

    PubMed Central

    Goggin, Danica E.; Powles, Stephen B.

    2012-01-01

    Background and Aims α-Amylase in grass caryopses (seeds) is usually expressed upon commencement of germination and is rarely seen in dry, mature seeds. A heat-stable α-amylase activity was unexpectedly selected for expression in dry annual ryegrass (Lolium rigidum) seeds during targeted selection for low primary dormancy. The aim of this study was to characterize this constitutive activity biochemically and determine if its presence conferred insensitivity to the germination inhibitors abscisic acid and benzoxazolinone. Methods α-Amylase activity in developing, mature and germinating seeds from the selected (low-dormancy) and a field-collected (dormant) population was characterized by native activity PAGE. The response of seed germination and α-amylase activity to abscisic acid and benzoxazolinone was assessed. Using an alginate affinity matrix, α-amylase was purified from dry and germinating seeds for analysis of its enzymatic properties. Key Results The constitutive α-amylase activity appeared late during seed development and was mainly localized in the aleurone; in germinating seeds, this activity was responsive to both glucose and gibberellin. It migrated differently on native PAGE compared with the major activities in germinating seeds of the dormant population, but the enzymatic properties of α-amylase purified from the low-dormancy and dormant seeds were largely indistinguishable. Seed imbibition on benzoxazolinone had little effect on the low-dormancy seeds but greatly inhibited germination and α-amylase activity in the dormant population. Conclusions The constitutive α-amylase activity in annual ryegrass seeds selected for low dormancy is electrophoretically different from that in germinating seeds and its presence confers insensitivity to benzoxazolinone. The concurrent selection of low dormancy and constitutive α-amylase activity may help to enhance seedling establishment under competitive conditions. PMID:23002268

  9. Evaluation of constitutive iron reductase (AtFRO2) expression on mineral accumulation and distribution in soybean (Glycine max. L)

    PubMed Central

    Vasconcelos, Marta W.; Clemente, Thomas E.; Grusak, Michael A.

    2014-01-01

    Iron is an important micronutrient in human and plant nutrition. Adequate iron nutrition during crop production is central for assuring appropriate iron concentrations in the harvestable organs, for human food or animal feed. The whole-plant movement of iron involves several processes, including the reduction of ferric to ferrous iron at several locations throughout the plant, prior to transmembrane trafficking of ferrous iron. In this study, soybean plants that constitutively expressed the AtFRO2 iron reductase gene were analyzed for leaf iron reductase activity, as well as the effect of this transgene’s expression on root, leaf, pod wall, and seed mineral concentrations. High Fe supply, in combination with the constitutive expression of AtFRO2, resulted in significantly higher concentrations of different minerals in roots (K, P, Zn, Ca, Ni, Mg, and Mo), pod walls (Fe, K, P, Cu, and Ni), leaves (Fe, P, Cu, Ca, Ni, and Mg) and seeds (Fe, Zn, Cu, and Ni). Leaf and pod wall iron concentrations increased as much as 500% in transgenic plants, while seed iron concentrations only increased by 10%, suggesting that factors other than leaf and pod wall reductase activity were limiting the translocation of iron to seeds. Protoplasts isolated from transgenic leaves had three-fold higher reductase activity than controls. Expression levels of the iron storage protein, ferritin, were higher in the transgenic leaves than in wild-type, suggesting that the excess iron may be stored as ferritin in the leaves and therefore unavailable for phloem loading and delivery to the seeds. Also, citrate and malate levels in the roots and leaves of transgenic plants were significantly higher than in wild-type, suggesting that organic acid production could be related to the increased accumulation of minerals in roots, leaves, and pod walls, but not in the seeds. All together, these results suggest a more ubiquitous role for the iron reductase in whole-plant mineral accumulation and

  10. Constitutional epimutation as a mechanism for cancer causality and heritability?

    PubMed

    Hitchins, Megan P

    2015-10-01

    Constitutional epimutation, which is an aberration in gene expression due to an altered epigenotype that is widely distributed in normal tissues (albeit frequently mosaic), provides an alternative mechanism to genetic mutation for cancer predisposition. Observational studies in cancer-affected families have revealed intergenerational inheritance of constitutional epimutation, providing unique insights into the heritability of epigenetic traits in humans. In this Opinion article, the potential contribution of constitutional epimutation to the 'missing' causality and heritability of cancer is explored.

  11. The Arabidopsis E3 Ubiquitin Ligase HOS1 Negatively Regulates CONSTANS Abundance in the Photoperiodic Control of Flowering[W

    PubMed Central

    Lazaro, Ana; Valverde, Federico; Piñeiro, Manuel; Jarillo, Jose A.

    2012-01-01

    The Arabidopsis thaliana early in short days6 (esd6) mutant was isolated in a screen for mutations that accelerate flowering time. Among other developmental alterations, esd6 displays early flowering in both long- and short-day conditions. Fine mapping of the mutation showed that the esd6 phenotype is caused by a lesion in the HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES1 (HOS1) locus, which encodes a RING finger–containing E3 ubiquitin ligase. The esd6/hos1 mutation causes decreased FLOWERING LOCUS C expression and requires CONSTANS (CO) protein for its early flowering phenotype under long days. Moreover, CO and HOS1 physically interact in vitro and in planta, and HOS1 regulates CO abundance, particularly during the daylight period. Accordingly, hos1 causes a shift in the regular long-day pattern of expression of FLOWERING LOCUS T (FT) transcript, starting to rise 4 h after dawn in the mutant. In addition, HOS1 interacts synergistically with CONSTITUTIVE PHOTOMORPHOGENIC1, another regulator of CO protein stability, in the regulation of flowering time. Taken together, these results indicate that HOS1 is involved in the control of CO abundance, ensuring that CO activation of FT occurs only when the light period reaches a certain length and preventing precocious flowering in Arabidopsis. PMID:22408073

  12. Constitutive cylindrospermopsin pool size in Cylindrospermopsis raciborskii under different light and CO2 partial pressure conditions.

    PubMed

    Pierangelini, Mattia; Sinha, Rati; Willis, Anusuya; Burford, Michele A; Orr, Philip T; Beardall, John; Neilan, Brett A

    2015-05-01

    Cylindrospermopsin (CYN) and 7-deoxy-cylindrospermopsin (dCYN) are potent hepatotoxic alkaloids produced by numerous species of cyanobacteria, including the freshwater Cylindrospermopsis raciborskii. C. raciborskii is an invasive cyanobacterium, and the study of how environmental parameters drive CYN production has received significant interest from water managers and health authorities. Light and CO2 affect cell growth and physiology in photoautotrophs, and these are potential regulators of cyanotoxin biosynthesis. In this study, we investigated how light and CO2 affect CYN and dCYN pool size as well as the expression of the key genes, cyrA and cyrK, involved in CYN biosynthesis in a toxic C. raciborskii strain. For cells growing at different light intensities (10 and 100 μmol photons m(-2) s(-1)), we observed that the rate of CYN pool size production (μCYN) was coupled to the cell division rate (μc) during batch culture. This indicated that CYN pool size under our experimental conditions is constant and cell quotas of CYN (QCYN) and dCYN (QdCYN) are fixed. Moreover, a lack of correlation between expression of cyrA and total CYN cell quotas (QCYNs) suggests that the CYN biosynthesis is regulated posttranscriptionally. Under elevated CO2 (1,300 ppm), we observed minor effects on QCYN and no effects on expression of cyrA and cyrK. We conclude that the CYN pool size is constitutive and not affected by light and CO2 conditions. Thus, C. raciborskii bloom toxicity is determined by the absolute abundance of C. raciborskii cells within the water column and the relative abundance of toxic and nontoxic strains. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  13. Regulatory elements involved in constitutive and phorbol ester-inducible expression of the plasminogen activator inhibitor type 2 gene promoter.

    PubMed Central

    Cousin, E; Medcalf, R L; Bergonzelli, G E; Kruithof, E K

    1991-01-01

    Gene transcription rates and mRNA levels of plasminogen activator inhibitor type 2 (PAI-2) are markedly induced by the tumor promoting agent phorbol 12-myristate 13-acetate (PMA) in human HT1080 fibrosarcoma cells. To identify promoter elements required for basal-, and phorbol ester-inducible expression, deletion mutants of the PAI-1 promoter fused to the chloramphenicol acetyl transferase (CAT) reporter gene, were transiently expressed in HT1080 cells. Constitutive CAT activity was expressed from constructs containing more than 215 bp of promoter sequence, whereas deletion to position -91 bp abolished CAT gene expression. Treatment of transfected cells with PMA resulted in a three- to ten-fold increase in CAT expression from all constructs except from the construct shortened to position -91. DNAse1 protection analysis of the promoter region between -215 and the transcription initiation site revealed numerous protected regions, including two AP1-like binding sites (AP1a and AP1b) and one CRE-like element. Site-directed mutagenesis of the AP1a site or of the CRE-like site resulted in the loss of basal CAT activity and abolished the PMA effect, whereas mutagenesis of AP1b only partially inhibited basal and PMA-mediated expression. Our results suggest that the PAI-2 promoter contains at least two elements required for basal gene transcription and PMA-mediated induction. Images PMID:1650454

  14. Comparison of Gut Microbiota between Sasang Constitutions

    PubMed Central

    Bae, Hyo Sang; Lim, Chi-yeon; Kim, Mi Jeong; Seo, Jae-gu; Kim, Jong Yeol; Kim, Jai-eun

    2013-01-01

    The Sasang constitutional medicine has long been applied to diagnose and treat patients with various diseases. Studies have been conducted for establishment of scientific evidence supporting Sasang Constitutional (SC) diagnosis. Recent human microbiome studies have demonstrated individual variations of gut microbiota which can be dependent on lifestyle and health conditions. We hypothesized that gut microbial similarities and discrepancies may exist across SC types. We compared the difference of gut microbiota among three constitutions (So-Yang, So-Eum, and Tae-Eum), along with the investigation of anthropometric and biochemical parameters. Firmicutes and Bacteroidetes were predominant phyla in all SC types. The median plot analysis suggested that Firmicutes and Bacteroidetes appeared more abundant in SE and TE, respectively, in the male subjects of 20–29 years old. At the genus level, Bifidobacterium and Bacteroides manifested the difference between SE and TE types. For anthropometry, body weight, body mass index, and waist circumference of the TE type were significantly higher than those of the other types. Overall, findings indicated a possible link between SC types and gut microbiota within a narrow age range. Further investigations are deemed necessary to elucidate the influences of age, gender, and other factors in the context of SC types and gut microbiota. PMID:24454486

  15. Constitutive expression of thymidylate synthase from LCDV-C induces a transformed phenotype in fish cells.

    PubMed

    Zhao, Zhe; Shi, Yan; Ke, Fei; Wei, Sun; Gui, Jianfang; Zhang, Qiya

    2008-03-01

    Thymidylate synthase (TS), an essential enzyme in DNA synthesis and repair, plays a key role in the events of cell cycle regulation and tumor formation. Here, an investigation was presented about subcellular location and biological function of viral TS from lymphocystis disease virus from China (LCDV-C) in fish cells. Fluorescence microscopy revealed that LCDV-C TS was predominantly localized in the cytoplasm in fish cells. Cell cycle analysis demonstrated that LCDV-C TS promoted cell cycle progression into S and G2/M phase in the constitutive expressed cells. As a result, the cells have a faster growth rate compared with the control cells as revealed by cell growth curves. For foci assay, the TS-expressed cells gave rise to foci 4-5 weeks after incubation. Microscopic examination of the TS-induced foci revealed multilayered growth and crisscross morphology characteristic of transformed cells. Moreover, LCDV-C TS predisposed the transfected cells to acquire an anchorage-independent phenotype and could grow in 0.3% soft agar. So the data reveal LCDV-C TS is sufficient to induce a transformed phenotype in fish cells in vitro and exhibits its potential ability in cell transformation. To our knowledge, it is the first report on viral TS sequences associated with transforming activity.

  16. Model reduction for stochastic chemical systems with abundant species.

    PubMed

    Smith, Stephen; Cianci, Claudia; Grima, Ramon

    2015-12-07

    Biochemical processes typically involve many chemical species, some in abundance and some in low molecule numbers. We first identify the rate constant limits under which the concentrations of a given set of species will tend to infinity (the abundant species) while the concentrations of all other species remains constant (the non-abundant species). Subsequently, we prove that, in this limit, the fluctuations in the molecule numbers of non-abundant species are accurately described by a hybrid stochastic description consisting of a chemical master equation coupled to deterministic rate equations. This is a reduced description when compared to the conventional chemical master equation which describes the fluctuations in both abundant and non-abundant species. We show that the reduced master equation can be solved exactly for a number of biochemical networks involving gene expression and enzyme catalysis, whose conventional chemical master equation description is analytically impenetrable. We use the linear noise approximation to obtain approximate expressions for the difference between the variance of fluctuations in the non-abundant species as predicted by the hybrid approach and by the conventional chemical master equation. Furthermore, we show that surprisingly, irrespective of any separation in the mean molecule numbers of various species, the conventional and hybrid master equations exactly agree for a class of chemical systems.

  17. Constitutive Smad signaling and Smad-dependent collagen gene expression in mouse embryonic fibroblasts lacking peroxisome proliferator-activated receptor-{gamma}

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Ghosh, Asish K; Wei, Jun; Wu, Minghua

    2008-09-19

    Transforming growth factor-{beta} (TGF-{beta}), a potent inducer of collagen synthesis, is implicated in pathological fibrosis. Peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) is a nuclear hormone receptor that regulates adipogenesis and numerous other biological processes. Here, we demonstrate that collagen gene expression was markedly elevated in mouse embryonic fibroblasts (MEFs) lacking PPAR-{gamma} compared to heterozygous control MEFs. Treatment with the PPAR-{gamma} ligand 15d-PGJ{sub 2} failed to down-regulate collagen gene expression in PPAR-{gamma} null MEFs, whereas reconstitution of these cells with ectopic PPAR-{gamma} resulted in their normalization. Compared to control MEFs, PPAR-{gamma} null MEFs displayed elevated levels of the Type I TGF-{beta} receptor (T{beta}RI),more » and secreted more TGF-{beta}1 into the media. Furthermore, PPAR-{gamma} null MEFs showed constitutive phosphorylation of cellular Smad2 and Smad3, even in the absence of exogenous TGF-{beta}, which was abrogated by the ALK5 inhibitor SB431542. Constitutive Smad2/3 phosphorylation in PPAR-{gamma} null MEFs was associated with Smad3 binding to its cognate DNA recognition sequences, and interaction with coactivator p300 previously implicated in TGF-{beta} responses. Taken together, these results indicate that loss of PPAR-{gamma} in MEFs is associated with upregulation of collagen synthesis, and activation of intracellular Smad signal transduction, due, at least in part, to autocrine TGF-{beta} stimulation.« less

  18. Herpes simplex virus 1 microRNAs expressed abundantly during latent infection are not essential for latency in mouse trigeminal ganglia

    PubMed Central

    Kramer, Martha F.; Jurak, Igor; Pesola, Jean M.; Boissel, Sandrine; Knipe, David M.; Coen, Donald M.

    2013-01-01

    Several herpes simplex virus 1 microRNAs are encoded within or near the latency associated transcript (LAT) locus, and are expressed abundantly during latency. Some of these microRNAs can repress the expression of important viral proteins and are hypothesized to play important roles in establishing and/or maintaining latent infections. We found that in lytically infected cells and in acutely infected mouse ganglia, expression of LAT-encoded microRNAs was weak and unaffected by a deletion that includes the LAT promoter. In mouse ganglia latently infected with wild type virus, the microRNAs accumulated to high levels, but deletions of the LAT promoter markedly reduced expression of LAT-encoded microRNAs and also miR-H6, which is encoded upstream of LAT and can repress expression of ICP4. Because these LAT deletion mutants establish and maintain latent infections, these microRNAs are not essential for latency, at least in mouse trigeminal ganglia, but may help promote it. PMID:21782205

  19. The abundant world: Paul Feyerabend's metaphysics of science.

    PubMed

    Brown, Matthew J

    2016-06-01

    The goal of this paper is to provide an interpretation of Feyerabend's metaphysics of science as found in late works like Conquest of Abundance and Tyranny of Science. Feyerabend's late metaphysics consists of an attempt to criticize and provide a systematic alternative to traditional scientific realism, a package of views he sometimes referred to as "scientific materialism." Scientific materialism is objectionable not only on metaphysical grounds, nor because it provides a poor ground for understanding science, but because it implies problematic claims about the epistemic and cultural authority of science, claims incompatible with situating science properly in democratic societies. I show how Feyerabend's metaphysical view, which I call "the abundant world" or "abundant realism," constitute a sophisticated and challenging form of ontological pluralism that makes interesting connections with contemporary philosophy of science and issues of the political and policy role of science in a democratic society. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Label-free quantitative proteomics for the extremely thermophilic bacterium Caldicellulosiruptor obsidiansis reveal distinct abundance patterns upon growth on cellobiose, crystalline cellulose, and switchgrass.

    PubMed

    Lochner, Adriane; Giannone, Richard J; Keller, Martin; Antranikian, Garabed; Graham, David E; Hettich, Robert L

    2011-12-02

    Mass spectrometric analysis of Caldicellulosiruptor obsidiansis cultures grown on four different carbon sources identified 65% of the cells' predicted proteins in cell lysates and supernatants. Biological and technical replication together with sophisticated statistical analysis were used to reliably quantify protein abundances and their changes as a function of carbon source. Extracellular, multifunctional glycosidases were significantly more abundant on cellobiose than on the crystalline cellulose substrates Avicel and filter paper, indicating either disaccharide induction or constitutive protein expression. Highly abundant flagellar, chemotaxis, and pilus proteins were detected during growth on insoluble substrates, suggesting motility or specific substrate attachment. The highly abundant extracellular binding protein COB47_0549 together with the COB47_1616 ATPase might comprise the primary ABC-transport system for cellooligosaccharides, while COB47_0096 and COB47_0097 could facilitate monosaccharide uptake. Oligosaccharide degradation can occur either via extracellular hydrolysis by a GH1 β-glycosidase or by intracellular phosphorolysis using two GH94 enzymes. When C. obsidiansis was grown on switchgrass, the abundance of hemicellulases (including GH3, GH5, GH51, and GH67 enzymes) and certain sugar transporters increased significantly. Cultivation on biomass also caused a concerted increase in cytosolic enzymes for xylose and arabinose fermentation.

  1. Nutrients Can Enhance the Abundance and Expression of Alkane Hydroxylase CYP153 Gene in the Rhizosphere of Ryegrass Planted in Hydrocarbon-Polluted Soil

    PubMed Central

    Arslan, Muhammad; Afzal, Muhammad; Amin, Imran; Iqbal, Samina; Khan, Qaiser M.

    2014-01-01

    Plant-bacteria partnership is a promising strategy for the remediation of soil and water polluted with hydrocarbons. However, the limitation of major nutrients (N, P and K) in soil affects the survival and metabolic activity of plant associated bacteria. The objective of this study was to explore the effects of nutrients on survival and metabolic activity of an alkane degrading rhizo-bacterium. Annual ryegrass (Lolium multiflorum) was grown in diesel-contaminated soil and inoculated with an alkane degrading bacterium, Pantoea sp. strain BTRH79, in greenhouse experiments. Two levels of nutrients were applied and plant growth, hydrocarbon removal, and gene abundance and expression were determined after 100 days of sowing of ryegrass. Results obtained from these experiments showed that the bacterial inoculation improved plant growth and hydrocarbon degradation and these were further enhanced by nutrients application. Maximum plant biomass production and hydrocarbon mineralization was observed by the combined use of inoculum and higher level of nutrients. The presence of nutrients in soil enhanced the colonization and metabolic activity of the inoculated bacterium in the rhizosphere. The abundance and expression of CYP153 gene in the rhizosphere of ryegrass was found to be directly associated with the level of applied nutrients. Enhanced hydrocarbon degradation was associated with the population of the inoculum bacterium, the abundance and expression of CYP153 gene in the rhizosphere of ryegrass. It is thus concluded that the combination between vegetation, inoculation with pollutant-degrading bacteria and nutrients amendment was an efficient approach to reduce hydrocarbon contamination. PMID:25360680

  2. Constitutional dynamic self-sensing in a zinc(II)/polyiminofluorenes system.

    PubMed

    Giuseppone, Nicolas; Lehn, Jean-Marie

    2004-09-22

    The interaction of an external effector, ZnII ions, with a constitutional dynamic library of fluorescent polyiminofluorenes leads to component exchange, which generates an entity responding by a change in emission to the effector that has induced its formation. The overall coupled system displays a tuning of optical signal, resulting from two synergistic processes: adaptative constitutional reorganization and self-sensing. In broader terms, this work highlights the perspectives opened by constitutional dynamic chemistry toward the design of smart materials, capable of expressing different latent properties in response to environmental conditions.

  3. Model reduction for stochastic chemical systems with abundant species

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Smith, Stephen; Cianci, Claudia; Grima, Ramon

    2015-12-07

    Biochemical processes typically involve many chemical species, some in abundance and some in low molecule numbers. We first identify the rate constant limits under which the concentrations of a given set of species will tend to infinity (the abundant species) while the concentrations of all other species remains constant (the non-abundant species). Subsequently, we prove that, in this limit, the fluctuations in the molecule numbers of non-abundant species are accurately described by a hybrid stochastic description consisting of a chemical master equation coupled to deterministic rate equations. This is a reduced description when compared to the conventional chemical master equationmore » which describes the fluctuations in both abundant and non-abundant species. We show that the reduced master equation can be solved exactly for a number of biochemical networks involving gene expression and enzyme catalysis, whose conventional chemical master equation description is analytically impenetrable. We use the linear noise approximation to obtain approximate expressions for the difference between the variance of fluctuations in the non-abundant species as predicted by the hybrid approach and by the conventional chemical master equation. Furthermore, we show that surprisingly, irrespective of any separation in the mean molecule numbers of various species, the conventional and hybrid master equations exactly agree for a class of chemical systems.« less

  4. Constitutive non-inducible expression of the Arabidopsis thaliana Nia 2 gene in two nitrate reductase mutants of Nicotiana plumbaginifolia.

    PubMed

    Kaye, C; Crawford, N M; Malmberg, R L

    1997-04-01

    We have isolated a haploid cell line of N. plumbaginifolia, hNP 588, that is constitutive and not inducible for nitrate reductase. Nitrate reductase mutants were isolated from hNP 588 protoplasts upon UV irradiation. Two of these nitrate reductase-deficient cell lines, nia 3 and nia 25, neither of which contained any detectable nitrate reductase activity, were selected for complementation studies. A cloned Arabidopsis thaliana nitrate reductase gene Nia 2 was introduced into each of the two mutants resulting in 56 independent kanamycin-resistant cell lines. Thirty of the 56 kanamycin-resistant cell lines were able to grow on nitrate as the sole nitrogen source. Eight of these were further analyzed for nitrate reductase enzyme activity and nitrate reductase mRNA production. All eight lines had detectable nitrate reductase activity ranging from 7% to 150% of wild-type hNP 588 callus. The enzyme activity levels were not influenced by the nitrogen source in the medium. The eight lines examined expressed a constitutive, non-inducible 3.2 kb mRNA species that was not present in untransformed controls.

  5. Transgenic expression of fungal accessory hemicellulases in Arabidopsis thaliana triggers transcriptional patterns related to biotic stress and defense response

    DOE PAGES

    Tsai, Alex Yi-Lin; Chan, Kin; Ho, Chi-Yip; ...

    2017-03-02

    The plant cell wall is an abundant and renewable resource for lignocellulosic applications such as the production of biofuel. Due to structural and compositional complexities, the plant cell wall is, however, recalcitrant to hydrolysis and extraction of platform sugars. A cell wall engineering strategy to reduce this recalcitrance makes use of microbial cell wall modifying enzymes that are expressed directly in plants themselves. Previously, we constructed transgenic Arabidopsis thaliana constitutively expressing the fungal hemicellulases: Phanerochaete carnosa glucurnoyl esterase (PcGCE) and Aspergillus nidulans α-arabinofuranosidase (AnAF54). While the PcGCE lines demonstrated improved xylan extractability, they also displayed chlorotic leaves leading to themore » hypothesis that expression of such enzymes in planta resulted in plant stress. The objective of this study is to investigate the impact of transgenic expression of the aforementioned microbial hemicellulases in planta on the host arabidopsis. More specifically, we investigated transcriptome profiles by short read high throughput sequencing (RNAseq) from developmentally distinct parts of the plant stem. When compared to non-transformed wild-type plants, a subset of genes was identified that showed differential transcript abundance in all transgenic lines and tissues investigated. Intriguingly, this core set of genes was significantly enriched for those involved in plant defense and biotic stress responses. While stress and defense-related genes showed increased transcript abundance in the transgenic plants regardless of tissue or genotype, genes involved in photosynthesis (light harvesting) were decreased in their transcript abundance potentially reflecting wide-spread effects of heterologous microbial transgene expression and the maintenance of plant homeostasis. Additionally, an increase in transcript abundance for genes involved in salicylic acid signaling further substantiates our finding that

  6. Transgenic expression of fungal accessory hemicellulases in Arabidopsis thaliana triggers transcriptional patterns related to biotic stress and defense response

    PubMed Central

    Tsai, Alex Yi-Lin; Chan, Kin; Ho, Chi-Yip; Canam, Thomas; Capron, Resmi; Master, Emma R.; Bräutigam, Katharina

    2017-01-01

    The plant cell wall is an abundant and renewable resource for lignocellulosic applications such as the production of biofuel. Due to structural and compositional complexities, the plant cell wall is, however, recalcitrant to hydrolysis and extraction of platform sugars. A cell wall engineering strategy to reduce this recalcitrance makes use of microbial cell wall modifying enzymes that are expressed directly in plants themselves. Previously, we constructed transgenic Arabidopsis thaliana constitutively expressing the fungal hemicellulases: Phanerochaete carnosa glucurnoyl esterase (PcGCE) and Aspergillus nidulans α-arabinofuranosidase (AnAF54). While the PcGCE lines demonstrated improved xylan extractability, they also displayed chlorotic leaves leading to the hypothesis that expression of such enzymes in planta resulted in plant stress. The objective of this study is to investigate the impact of transgenic expression of the aforementioned microbial hemicellulases in planta on the host arabidopsis. More specifically, we investigated transcriptome profiles by short read high throughput sequencing (RNAseq) from developmentally distinct parts of the plant stem. When compared to non-transformed wild-type plants, a subset of genes was identified that showed differential transcript abundance in all transgenic lines and tissues investigated. Intriguingly, this core set of genes was significantly enriched for those involved in plant defense and biotic stress responses. While stress and defense-related genes showed increased transcript abundance in the transgenic plants regardless of tissue or genotype, genes involved in photosynthesis (light harvesting) were decreased in their transcript abundance potentially reflecting wide-spread effects of heterologous microbial transgene expression and the maintenance of plant homeostasis. Additionally, an increase in transcript abundance for genes involved in salicylic acid signaling further substantiates our finding that transgenic

  7. Transgenic expression of fungal accessory hemicellulases in Arabidopsis thaliana triggers transcriptional patterns related to biotic stress and defense response

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tsai, Alex Yi-Lin; Chan, Kin; Ho, Chi-Yip

    The plant cell wall is an abundant and renewable resource for lignocellulosic applications such as the production of biofuel. Due to structural and compositional complexities, the plant cell wall is, however, recalcitrant to hydrolysis and extraction of platform sugars. A cell wall engineering strategy to reduce this recalcitrance makes use of microbial cell wall modifying enzymes that are expressed directly in plants themselves. Previously, we constructed transgenic Arabidopsis thaliana constitutively expressing the fungal hemicellulases: Phanerochaete carnosa glucurnoyl esterase (PcGCE) and Aspergillus nidulans α-arabinofuranosidase (AnAF54). While the PcGCE lines demonstrated improved xylan extractability, they also displayed chlorotic leaves leading to themore » hypothesis that expression of such enzymes in planta resulted in plant stress. The objective of this study is to investigate the impact of transgenic expression of the aforementioned microbial hemicellulases in planta on the host arabidopsis. More specifically, we investigated transcriptome profiles by short read high throughput sequencing (RNAseq) from developmentally distinct parts of the plant stem. When compared to non-transformed wild-type plants, a subset of genes was identified that showed differential transcript abundance in all transgenic lines and tissues investigated. Intriguingly, this core set of genes was significantly enriched for those involved in plant defense and biotic stress responses. While stress and defense-related genes showed increased transcript abundance in the transgenic plants regardless of tissue or genotype, genes involved in photosynthesis (light harvesting) were decreased in their transcript abundance potentially reflecting wide-spread effects of heterologous microbial transgene expression and the maintenance of plant homeostasis. Additionally, an increase in transcript abundance for genes involved in salicylic acid signaling further substantiates our finding that

  8. Constitutive expression of the promyelocytic leukemia-associated oncogene PML-RARalpha in TF1 cells: isoform-specific and retinoic acid-dependent effects on growth, bcl-2 expression, and apoptosis.

    PubMed

    Slack, J L; Yu, M

    1998-05-01

    Two major isoforms of PML-RARalpha are associated with (15;17)-positive acute promyelocytic leukemia (APL); however, functional differences between these isoforms have been difficult to define, and the molecular mechanism by which each isoform contributes to the pathogenesis of APL is not fully understood. To address these issues, the 'short' (S) and 'long' (L) isoforms of PML-RARalpha were constitutively expressed in the factor-dependent human erythroleukemia cell line, TF1. Expression of the L, but not the S, isoform inhibited growth of these cells in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF). In the absence of GM-CSF, the S isoform partially protected against apoptosis, while the L isoform accelerated cell death. Treatment with all-trans retinoic acid (ATRA) inhibited cell growth and caused apoptosis only in PML-RARalpha-expressing cells, and these effects of ATRA were more marked in cells expressing the L isoform. ATRA treatment also led to downregulation of bcl-2 and endogenous RARalpha in PML-RARalpha-expressing cells, but had little effect on the level of exogenously expressed PML-RARalpha. We conclude that (1) subtle differences exist in the biologic activities of the L and S isoforms of PML-RARalpha, and (2) both isoforms are capable of transducing an ATRA-mediated signal that leads to downregulation of bcl-2 and induction of programmed cell death.

  9. Linking Changes in Snow Cover with Nitrogen Cycling and Microbial Abundance and Functional Gene Expression in Agricultural Soils

    NASA Astrophysics Data System (ADS)

    Goyer, C.; Brin, L.; Zebarth, B.; Burton, D.; Wertz, S.; Chantigny, M.

    2016-12-01

    In eastern Canada, climate change-related warming and increased precipitation may alter winter snow cover, with potential consequences for soil conditions, microbes, and N2O fluxes. We conducted a two-year field study with snow removal, passive snow addition, and ambient treatments in a potato-barley crop system. We measured in situ greenhouse gas (N2O and CO2) fluxes and belowground gas accumulation, and quantified abundance and expression of denitrifier (nirS, nirK, nosZ) and nitrifier (ammonium oxidizing archaeal (AOA) and bacterial (AOB) amoA) genes. Soil gas accumulated throughout winter, and surface fluxes were greatest during spring thaw. Greatest mid-winter soil N2O accumulation and spring thaw N2O fluxes were associated with snow removal in winter 1 and ambient snow in winter 2. High N2O accumulation and fluxes may have been due to increased substrate availability with increased frost intensity in removal plots in winter 1, but with greatest water content in ambient plots in winter 2. In each winter, greatest abundances of nirS, nirK gene denitrifiers and/or amoA gene of AOA were observed in the treatments with the greatest N2O accumulation and fluxes. Gene expression did not vary with treatment, but highest expression of amoA gene of AOA and AOB, and nosZ gene was measured near 0ºC, indicating activity during periods of stable snow cover and spring thaw. Results suggest that the magnitude of fluxes during spring thaw were related to soil conditions and microbial communities present during the prior winter, and not solely those during thaw. Furthermore, the effects of changing snow cover on microbes and N2O fluxes were not a straightforward effect of snow depth, but were likely mediated by temperature and moisture.

  10. Constitutive stimulatory G protein activity in limb mesenchyme impairs bone growth.

    PubMed

    Karaca, Anara; Malladi, Vijayram Reddy; Zhu, Yan; Tafaj, Olta; Paltrinieri, Elena; Wu, Joy Y; He, Qing; Bastepe, Murat

    2018-05-01

    GNAS mutations leading to constitutively active stimulatory G protein alpha-subunit (Gsα) cause different tumors, fibrous dysplasia of bone, and McCune-Albright syndrome, which are typically not associated with short stature. Enhanced signaling of the parathyroid hormone/parathyroid hormone-related peptide receptor, which couples to multiple G proteins including Gsα, leads to short bones with delayed endochondral ossification. It has remained unknown whether constitutive Gsα activity also impairs bone growth. Here we generated mice expressing a constitutively active Gsα mutant (Gsα-R201H) conditionally upon Cre recombinase (cGsα R201H mice). Gsα-R201H was expressed in cultured bone marrow stromal cells from cGsα R201H mice upon adenoviral-Cre transduction. When crossed with mice in which Cre is expressed in a tamoxifen-regulatable fashion (CAGGCre-ER™), tamoxifen injection resulted in mosaic expression of the transgene in double mutant offspring. We then crossed the cGsα R201H mice with Prx1-Cre mice, in which Cre is expressed in early limb-bud mesenchyme. The double mutant offspring displayed short limbs at birth, with narrow hypertrophic chondrocyte zones in growth plates and delayed formation of secondary ossification center. Consistent with enhanced Gsα signaling, bone marrow stromal cells from these mice demonstrated increased levels of c-fos mRNA. Our findings indicate that constitutive Gsα activity during limb development disrupts endochondral ossification and bone growth. Given that Gsα haploinsufficiency also leads to short bones, as in patients with Albright's hereditary osteodystrophy, these results suggest that a tight control of Gsα activity is essential for normal growth plate physiology. Copyright © 2018 Elsevier Inc. All rights reserved.

  11. Constitutive activity of the δ-opioid receptor expressed in C6 glioma cells: identification of non-peptide δ-inverse agonists

    PubMed Central

    Neilan, Claire L; Akil, Huda; Woods, James H; Traynor, John R

    1999-01-01

    G-protein coupled receptors can exhibit constitutive activity resulting in the formation of active ternary complexes in the absence of an agonist. In this study we have investigated constitutive activity in C6 glioma cells expressing either the cloned δ-(OP1) receptor (C6δ), or the cloned μ-(OP3) opioid receptor (C6μ).Constitutive activity was measured in the absence of Na+ ions to provide an increased signal. The degree of constitutive activity was defined as the level of [35S]-GTPγS binding that could be inhibited by pre-treatment with pertussis toxin (PTX). In C6δ cells the level of basal [35S]-GTPγS binding was reduced by 51.9±6.1 fmols mg−1 protein, whereas in C6μ and C6 wild-type cells treatment with PTX reduced basal [35S]-GTPγS binding by only 10.0±3.5 and 8.6±3.1 fmols mg−1 protein respectively.The δ-antagonists N,N-diallyl-Tyr-Aib-Aib-Phe-Leu-OH (ICI 174,864), 7-benzylidenenaltrexone (BNTX) and naltriben (NTB), in addition to clocinnamox (C-CAM), acted as δ-opioid receptor inverse agonists. Naloxone, buprenorphine, and naltrindole were neutral antagonists. Furthermore, naltrindole blocked the reduction in [35S]-GTPγS binding caused by the inverse agonists. The inverse agonists did not inhibit basal [35S]-GTPγS binding in C6μ or C6 wild-type cell membranes.Competition binding assays in C6δ cell membranes revealed a leftward shift in the displacement curve of [3H]-naltrindole by ICI 174,864 and C-CAM in the presence of NaCl and the GTP analogue, GppNHp. There was no change in the displacement curve for BNTX or NTB under these conditions.These data confirm the presence of constitutive activity associated with the δ-opioid receptor and identify three novel, non-peptide, δ-opioid inverse agonists. PMID:10516632

  12. Production of dengue virus envelope protein domain III-based antigens in tobacco chloroplasts using inducible and constitutive expression systems.

    PubMed

    Gottschamel, Johanna; Lössl, Andreas; Ruf, Stephanie; Wang, Yanliang; Skaugen, Morten; Bock, Ralph; Clarke, Jihong Liu

    2016-07-01

    Dengue fever is a disease in many parts of the tropics and subtropics and about half the world's population is at risk of infection according to the World Health Organization. Dengue is caused by any of the four related dengue virus serotypes DEN-1, -2, -3 and -4, which are transmitted to people by Aedes aegypti mosquitoes. Currently there is only one vaccine (Dengvaxia(®)) available (limited to a few countries) on the market since 2015 after half a century's intensive efforts. Affordable and accessible vaccines against dengue are hence still urgently needed. The dengue envelop protein domain III (EDIII), which is capable of eliciting serotype-specific neutralizing antibodies, has become the focus for subunit vaccine development. To contribute to the development of an accessible and affordable dengue vaccine, in the current study we have used plant-based vaccine production systems to generate a dengue subunit vaccine candidate in tobacco. Chloroplast genome engineering was applied to express serotype-specific recombinant EDIII proteins in tobacco chloroplasts using both constitutive and ethanol-inducible expression systems. Expression of a tetravalent antigen fusion construct combining EDIII polypeptides from all four serotypes was also attempted. Transplastomic EDIII-expressing tobacco lines were obtained and homoplasmy was verified by Southern blot analysis. Northern blot analyses showed expression of EDIII antigen-encoding genes. EDIII protein accumulation levels varied for the different recombinant EDIII proteins and the different expression systems, and reached between 0.8 and 1.6 % of total cellular protein. Our study demonstrates the suitability of the chloroplast compartment as a production site for an EDIII-based vaccine candidate against dengue fever and presents a Gateway(®) plastid transformation vector for inducible transgene expression.

  13. Phytate addition to soil induces changes in the abundance and expression of Bacillus β-propeller phytase genes in the rhizosphere.

    PubMed

    Jorquera, Milko A; Saavedra, Nicolás; Maruyama, Fumito; Richardson, Alan E; Crowley, David E; del C Catrilaf, Rosa; Henriquez, Evelyn J; de la Luz Mora, María

    2013-02-01

    Phytate-mineralizing rhizobacteria (PMR) perform an essential function for the mineralization of organic phosphorus but little is known about their ecology in soils and rhizosphere. In this study, PCR-based methods were developed for detection and quantification of the Bacillus β-propeller phytase (BPP) gene. Experiments were conducted to monitor the presence and persistence of a phytate-mineralizing strain, Bacillus sp. MQH19, after inoculation of soil microcosms and within the rhizosphere. The occurrence of the BPP gene in natural pasture soils from Chilean Andisols was also examined. The results showed that the Bacillus BPP gene was readily detected in sterile and nonsterile microcosms, and that the quantitative PCR (qPCR) methods could be used to monitor changes in the abundance of the BPP gene over time. Our results also show that the addition of phytate to nonsterile soils induced the expression of the BPP gene in the rhizosphere of ryegrass and the BPP gene was detected in all pasture soils sampled. This study shows that phytate addition soils induced changes in the abundance and expression of Bacillus BPP to genes in the rhizosphere and demonstrates that Bacillus BPP gene is cosmopolitan in pasture soils from Chilean Andisols. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  14. Acute hypoxia stress induced abundant differential expression genes and alternative splicing events in heart of tilapia.

    PubMed

    Xia, Jun Hong; Li, Hong Lian; Li, Bi Jun; Gu, Xiao Hui; Lin, Hao Ran

    2018-01-10

    Hypoxia is one of the critical environmental stressors for fish in aquatic environments. Although accumulating evidences indicate that gene expression is regulated by hypoxia stress in fish, how genes undergoing differential gene expression and/or alternative splicing (AS) in response to hypoxia stress in heart are not well understood. Using RNA-seq, we surveyed and detected 289 differential expressed genes (DEG) and 103 genes that undergo differential usage of exons and splice junctions events (DUES) in heart of a hypoxia tolerant fish, Nile tilapia, Oreochromis niloticus following 12h hypoxic treatment. The spatio-temporal expression analysis validated the significant association of differential exon usages in two randomly selected DUES genes (fam162a and ndrg2) in 5 tissues (heart, liver, brain, gill and spleen) sampled at three time points (6h, 12h, and 24h) under acute hypoxia treatment. Functional analysis significantly associated the differential expressed genes with the categories related to energy conservation, protein synthesis and immune response. Different enrichment categories were found between the DEG and DUES dataset. The Isomerase activity, Oxidoreductase activity, Glycolysis and Oxidative stress process were significantly enriched for the DEG gene dataset, but the Structural constituent of ribosome and Structural molecule activity, Ribosomal protein and RNA binding protein were significantly enriched only for the DUES genes. Our comparative transcriptomic analysis reveals abundant stress responsive genes and their differential regulation function in the heart tissues of Nile tilapia under acute hypoxia stress. Our findings will facilitate future investigation on transcriptome complexity and AS regulation during hypoxia stress in fish. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Oncogenic JAK2V617F requires an intact SH2-like domain for constitutive activation and induction of a myeloproliferative disease in mice.

    PubMed

    Gorantla, Sivahari P; Dechow, Tobias N; Grundler, Rebekka; Illert, Anna Lena; Zum Büschenfelde, Christian Meyer; Kremer, Marcus; Peschel, Christian; Duyster, Justus

    2010-11-25

    The oncogenic JAK2V617F mutation is found in myeloproliferative neoplasms (MPNs) and is believed to be critical for leukemogenesis. Here we show that JAK2V617F requires an intact SH2 domain for constitutive activation of downstream signaling pathways. In addition, there is a strict requirement of cytokine receptor expression for the activation of this oncogene. Further analysis showed that the SH2 domain mutation did not interfere with JAK2 membrane distribution. However, coimmunoprecipitated experiments revealed a role for the SH2 domain in the aggregation and cross-phosphorylation of JAK2V617F at the cell membrane. Forced overexpression of cytokine receptors could rescue the JAK2V617F SH2 mutant supporting a critical role of JAK2V617F abundance for constitutive activation. However, under physiologic cytokine receptor expression the SH2 domain is absolutely necessary for oncogenic JAK2V617F activation. This is demonstrated in a bone marrow transplantation model, in which an intact SH2 domain in JAK2V617F is required for the induction of an MPN-like disease. Thus, our results points to an indispensable role of the SH2 domain in JAK2V617F-induced MPNs.

  16. Stage 3 immature human natural killer cells found in secondary lymphoid tissue constitutively and selectively express the TH17 cytokine interleukin-22

    PubMed Central

    Hughes, Tiffany; Becknell, Brian; McClory, Susan; Briercheck, Edward; Freud, Aharon G.; Zhang, Xiaoli; Mao, Hsiaoyin; Nuovo, Gerard; Yu, Jianhua

    2009-01-01

    Considerable functional heterogeneity within human natural killer (NK) cells has been revealed through the characterization of distinct NK-cell subsets. Accordingly, a small subset of CD56+NKp44+NK cells, termed NK-22 cells, was recently described within secondary lymphoid tissue (SLT) as IL-22− when resting, with a minor fraction of this population becoming IL-22+ when activated. Here we discover that the vast majority of stage 3 immature NK (iNK) cells in SLT constitutively and selectively express IL-22, a TH17 cytokine important for mucosal immunity, whereas earlier and later stages of NK developmental intermediates do not express IL-22. These iNK cells have a surface phenotype of CD34−CD117+CD161+CD94−, largely lack expression of NKp44 and CD56, and do not produce IFN-γ or possess cytolytic activity. In summary, stage 3 iNK cells are highly enriched for IL-22 and IL-26 messenger RNA, and IL-22 protein production, but do not express IL-17A or IL-17F. PMID:19244159

  17. Constitutional 3p26.3 terminal microdeletion in an adolescent with neuroblastoma.

    PubMed

    Pezzolo, Annalisa; Sementa, Angela Rita; Lerone, Margherita; Morini, Martina; Ognibene, Marzia; Defferrari, Raffaella; Mazzocco, Katia; Conte, Massimo; Gigliotti, Anna Rita; Garaventa, Alberto; Pistoia, Vito; Varesio, Luigi

    2017-05-04

    Neuroblastoma (NB) is a common and often lethal cancer of early childhood that accounts for 10% of pediatric cancer mortality. Incidence peaks in infancy and then rapidly declines, with less than 5% of cases diagnosed in children and adolescents ≥ 10 y. There is increasing evidence that NB has unique biology and an chronic disease course in older children and adolescents, but ultimately dismal survival. We describe a rare constitutional 3p26.3 terminal microdeletion which occurred in an adolescent with NB, with apparently normal phenotype without neurocognitive defects. We evaluated the association of expression of genes involved in the microdeletion with NB patient outcomes using R2 platform. We screened NB patient's tumor cells for CHL1 protein expression using immunofluorescence. Constitutional and tumor DNA were tested by array-comparative genomic hybridization and single nucleotide-polymorphism-array analyses. Peripheral blood mononuclear cells from the patient showed a 2.54 Mb sub-microscopic constitutional terminal 3p deletion that extended to band p26.3. The microdeletion 3p disrupted the CNTN4 gene and the neighboring CNTN6 and CHL1 genes were hemizygously deleted, each of these genes encode neuronal cell adhesion molecules. Low expression of CNTN6 and CNTN4 genes did not stratify NB patients, whereas low CHL1 expression characterized 417 NB patients having worse overall survival. CHL1 protein expression on tumor cells from the patient was weaker than positive control. This is the first report of a constitutional 3p26.3 deletion in a NB patient. Since larger deletions of 3p, indicative of the presence of one or more tumor suppressor genes in this region, occur frequently in neuroblastoma, our results pave the way to the identification of one putative NB suppressor genes mapping in 3p26.3.

  18. Gene co-expression network analysis in Rhodobacter capsulatus and application to comparative expression analysis of Rhodobacter sphaeroides

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Pena-Castillo, Lourdes; Mercer, Ryan; Gurinovich, Anastasia

    2014-08-28

    The genus Rhodobacter contains purple nonsulfur bacteria found mostly in freshwater environments. Representative strains of two Rhodobacter species, R. capsulatus and R. sphaeroides, have had their genomes fully sequenced and both have been the subject of transcriptional profiling studies. Gene co-expression networks can be used to identify modules of genes with similar expression profiles. Functional analysis of gene modules can then associate co-expressed genes with biological pathways, and network statistics can determine the degree of module preservation in related networks. In this paper, we constructed an R. capsulatus gene co-expression network, performed functional analysis of identified gene modules, and investigatedmore » preservation of these modules in R. capsulatus proteomics data and in R. sphaeroides transcriptomics data. Results: The analysis identified 40 gene co-expression modules in R. capsulatus. Investigation of the module gene contents and expression profiles revealed patterns that were validated based on previous studies supporting the biological relevance of these modules. We identified two R. capsulatus gene modules preserved in the protein abundance data. We also identified several gene modules preserved between both Rhodobacter species, which indicate that these cellular processes are conserved between the species and are candidates for functional information transfer between species. Many gene modules were non-preserved, providing insight into processes that differentiate the two species. In addition, using Local Network Similarity (LNS), a recently proposed metric for expression divergence, we assessed the expression conservation of between-species pairs of orthologs, and within-species gene-protein expression profiles. Conclusions: Our analyses provide new sources of information for functional annotation in R. capsulatus because uncharacterized genes in modules are now connected with groups of genes that constitute a joint functional

  19. Employing conservation of co-expression to improve functional inference

    PubMed Central

    Daub, Carsten O; Sonnhammer, Erik LL

    2008-01-01

    Background Observing co-expression between genes suggests that they are functionally coupled. Co-expression of orthologous gene pairs across species may improve function prediction beyond the level achieved in a single species. Results We used orthology between genes of the three different species S. cerevisiae, D. melanogaster, and C. elegans to combine co-expression across two species at a time. This led to increased function prediction accuracy when we incorporated expression data from either of the other two species and even further increased when conservation across both of the two other species was considered at the same time. Employing the conservation across species to incorporate abundant model organism data for the prediction of protein interactions in poorly characterized species constitutes a very powerful annotation method. Conclusion To be able to employ the most suitable co-expression distance measure for our analysis, we evaluated the ability of four popular gene co-expression distance measures to detect biologically relevant interactions between pairs of genes. For the expression datasets employed in our co-expression conservation analysis above, we used the GO and the KEGG PATHWAY databases as gold standards. While the differences between distance measures were small, Spearman correlation showed to give most robust results. PMID:18808668

  20. Constitutive expression, purification and characterisation of pectin methylesterase from Aspergillus niger in Pichia pastoris for potential application in the fruit juice industry.

    PubMed

    Jiang, Xiuping; Chen, Peng; Yin, Maolu; Yang, Qing

    2013-01-01

    Pectin methylesterase (PME) catalyses the hydrolysis of the methyl ester of pectin, yielding free carboxyl groups and methanol. PME is widely used in the food, cosmetic and pharmaceutical industries. PME from Aspergillus niger was constitutively expressed to a high level in the yeast Pichia pastoris. The recombinant PME was purified by a combination of ammonium sulfate fractionation and ion exchange chromatography, giving an overall yield of 28.0%. It appeared as a single band in sodium dodecyl sulfate polyacrylamide gel electrophoresis, with a molecular mass of about 45 kDa. Optimal activity of the enzyme occurred at a temperature of 50 °C and a pH of 4.7. The K(m), V(max) and k(cat) values of the enzyme with respect to pectin were 8.6 mmol L⁻¹ [Formula: See Text], 1.376 mmol min⁻¹ mg⁻¹ and 8.26 × 10² s⁻¹ respectively. Cations such as K⁺, Mg²⁺, Ni²⁺, Mn²⁺ and Co²⁺ slightly inhibited its activity, whereas Na⁺ had no effect. PME from A. niger was constitutively expressed to a high level in P. pastoris without methanol induction. The recombinant PME was purified and characterised and shown to be a good candidate for potential application in the fruit juice industry. Copyright © 2012 Society of Chemical Industry.

  1. Generalized estimators of avian abundance from count survey data

    USGS Publications Warehouse

    Royle, J. Andrew

    2004-01-01

    I consider modeling avian abundance from spatially referenced bird count data collected according to common protocols such as capture?recapture, multiple observer, removal sampling and simple point counts. Small sample sizes and large numbers of parameters have motivated many analyses that disregard the spatial indexing of the data, and thus do not provide an adequate treatment of spatial structure. I describe a general framework for modeling spatially replicated data that regards local abundance as a random process, motivated by the view that the set of spatially referenced local populations (at the sample locations) constitute a metapopulation. Under this view, attention can be focused on developing a model for the variation in local abundance independent of the sampling protocol being considered. The metapopulation model structure, when combined with the data generating model, define a simple hierarchical model that can be analyzed using conventional methods. The proposed modeling framework is completely general in the sense that broad classes of metapopulation models may be considered, site level covariates on detection and abundance may be considered, and estimates of abundance and related quantities may be obtained for sample locations, groups of locations, unsampled locations. Two brief examples are given, the first involving simple point counts, and the second based on temporary removal counts. Extension of these models to open systems is briefly discussed.

  2. Combining Constitutively Active Rheb Expression and Chondroitinase Promotes Functional Axonal Regeneration after Cervical Spinal Cord Injury.

    PubMed

    Wu, Di; Klaw, Michelle C; Connors, Theresa; Kholodilov, Nikolai; Burke, Robert E; Côté, Marie-Pascale; Tom, Veronica J

    2017-12-06

    After spinal cord injury (SCI), severed axons in the adult mammalian CNS are unable to mount a robust regenerative response. In addition, the glial scar at the lesion site further restricts the regenerative potential of axons. We hypothesized that a combinatorial approach coincidentally targeting these obstacles would promote axonal regeneration. We combined (1) transplantation of a growth-permissive peripheral nerve graft (PNG) into an incomplete, cervical lesion cavity; (2) transduction of neurons rostral to the SCI site to express constitutively active Rheb (caRheb; a Ras homolog enriched in brain), a GTPase that directly activates the growth-promoting pathway mammalian target of rapamycin (mTOR) via AAV-caRheb injection; and (3) digestion of growth-inhibitory chondroitin sulfate proteoglycans within the glial scar at the distal PNG interface using the bacterial enzyme chondroitinase ABC (ChABC). We found that expressing caRheb in neurons post-SCI results in modestly yet significantly more axons regenerating across a ChABC-treated distal graft interface into caudal spinal cord than either treatment alone. Excitingly, we found that caRheb+ChABC treatment significantly potentiates the formation of synapses in the host spinal cord and improves the animals' ability to use the affected forelimb. Thus, this combination strategy enhances functional axonal regeneration following a cervical SCI. Copyright © 2017 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.

  3. Growth-rate dependent global effects on gene expression in bacteria

    PubMed Central

    Klumpp, Stefan; Zhang, Zhongge; Hwa, Terence

    2010-01-01

    Summary Bacterial gene expression depends not only on specific regulations but also directly on bacterial growth, because important global parameters such as the abundance of RNA polymerases and ribosomes are all growth-rate dependent. Understanding these global effects is necessary for a quantitative understanding of gene regulation and for the robust design of synthetic genetic circuits. The observed growth-rate dependence of constitutive gene expression can be explained by a simple model using the measured growth-rate dependence of the relevant cellular parameters. More complex growth dependences for genetic circuits involving activators, repressors and feedback control were analyzed, and salient features were verified experimentally using synthetic circuits. The results suggest a novel feedback mechanism mediated by general growth-dependent effects and not requiring explicit gene regulation, if the expressed protein affects cell growth. This mechanism can lead to growth bistability and promote the acquisition of important physiological functions such as antibiotic resistance and tolerance (persistence). PMID:20064380

  4. Genomic Repeat Abundances Contain Phylogenetic Signal

    PubMed Central

    Dodsworth, Steven; Chase, Mark W.; Kelly, Laura J.; Leitch, Ilia J.; Macas, Jiří; Novák, Petr; Piednoël, Mathieu; Weiss-Schneeweiss, Hanna; Leitch, Andrew R.

    2015-01-01

    A large proportion of genomic information, particularly repetitive elements, is usually ignored when researchers are using next-generation sequencing. Here we demonstrate the usefulness of this repetitive fraction in phylogenetic analyses, utilizing comparative graph-based clustering of next-generation sequence reads, which results in abundance estimates of different classes of genomic repeats. Phylogenetic trees are then inferred based on the genome-wide abundance of different repeat types treated as continuously varying characters; such repeats are scattered across chromosomes and in angiosperms can constitute a majority of nuclear genomic DNA. In six diverse examples, five angiosperms and one insect, this method provides generally well-supported relationships at interspecific and intergeneric levels that agree with results from more standard phylogenetic analyses of commonly used markers. We propose that this methodology may prove especially useful in groups where there is little genetic differentiation in standard phylogenetic markers. At the same time as providing data for phylogenetic inference, this method additionally yields a wealth of data for comparative studies of genome evolution. PMID:25261464

  5. Inventory of high-abundance mRNAs in skeletal muscle of normal men.

    PubMed

    Welle, S; Bhatt, K; Thornton, C A

    1999-05-01

    G42875rial analysis of gene expression (SAGE) method was used to generate a catalog of 53,875 short (14 base) expressed sequence tags from polyadenylated RNA obtained from vastus lateralis muscle of healthy young men. Over 12,000 unique tags were detected. The frequency of occurrence of each tag reflects the relative abundance of the corresponding mRNA. The mRNA species that were detected 10 or more times, each comprising >/=0.02% of the mRNA population, accounted for 64% of the mRNA mass but <10% of the total number of mRNA species detected. Almost all of the abundant tags matched mRNA or EST sequences cataloged in GenBank. Mitochondrial transcripts accounted for approximately 20% of the polyadenylated RNA. Transcripts encoding proteins of the myofibrils were the most abundant nuclear-encoded mRNAs. Transcripts encoding ribosomal proteins, and those encoding proteins involved in energy metabolism, also were very abundant. The database can be used as a reference for investigations of alterations in gene expression associated with conditions that influence muscle function, such as muscular dystrophies, aging, and exercise.

  6. Identification of chemical modulators of the constitutive activated receptor (CAR) in a gene expression compendium

    PubMed Central

    Oshida, Keiyu; Vasani, Naresh; Jones, Carlton; Moore, Tanya; Hester, Susan; Nesnow, Stephen; Auerbach, Scott; Geter, David R.; Aleksunes, Lauren M.; Thomas, Russell S.; Applegate, Dawn; Klaassen, Curtis D.; Corton, J. Christopher

    2015-01-01

    The nuclear receptor family member constitutive activated receptor (CAR) is activated by structurally diverse drugs and environmentally-relevant chemicals leading to transcriptional regulation of genes involved in xenobiotic metabolism and transport. Chronic activation of CAR increases liver cancer incidence in rodents, whereas suppression of CAR can lead to steatosis and insulin insensitivity. Here, analytical methods were developed to screen for chemical treatments in a gene expression compendium that lead to alteration of CAR activity. A gene expression biomarker signature of 83 CAR-dependent genes was identified using microarray profiles from the livers of wild-type and CAR-null mice after exposure to three structurally-diverse CAR activators (CITCO, phenobarbital, TCPOBOP). A rank-based algorithm (Running Fisher’s algorithm (p-value ≤ 10-4)) was used to evaluate the similarity between the CAR biomarker signature and a test set of 28 and 32 comparisons positive or negative, respectively, for CAR activation; the test resulted in a balanced accuracy of 97%. The biomarker signature was used to identify chemicals that activate or suppress CAR in an annotated mouse liver/primary hepatocyte gene expression database of ~1850 comparisons. CAR was activated by 1) activators of the aryl hydrocarbon receptor (AhR) in wild-type but not AhR-null mice, 2) pregnane X receptor (PXR) activators in wild-type and to lesser extents in PXR-null mice, and 3) activators of PPARα in wild-type and PPARα-null mice. CAR was consistently activated by five conazole fungicides and four perfluorinated compounds. Comparison of effects in wild-type and CAR-null mice showed that the fungicide propiconazole increased liver weight and hepatocyte proliferation in a CAR-dependent manner, whereas the perfluorinated compound perfluorooctanoic acid (PFOA) increased these endpoints in a CAR-independent manner. A number of compounds suppressed CAR coincident with increases in markers of

  7. On Using the Color-Magnitude Diagram Morphology of M67 to Test Solar Abundances

    NASA Astrophysics Data System (ADS)

    Magic, Z.; Serenelli, A.; Weiss, A.; Chaboyer, B.

    2010-08-01

    The open cluster M67 has solar metallicity and an age of about 4 Gyr. The turnoff (TO) mass is close to the minimum mass for which solar metallicity stars develop a convective core during main sequence evolution as a result of the development of hydrogen burning through the CNO cycle. The morphology of the color-magnitude diagram (CMD) of M67 around the TO shows a clear hook-like feature, a direct sign that stars close to the TO have convective cores. VandenBerg et al. investigated the possibility of using the morphology of the M67 TO to put constraints on the solar metallicity, particularly CNO elements, for which solar abundances have been revised downward by more than 30% over the last few years. Here, we extend their work, filling the gaps in their analysis. To this aim, we compute isochrones appropriate for M67 using new (low metallicity) and old (high metallicity) solar abundances and study whether the characteristic TO in the CMD of M67 can be reproduced or not. We also study the importance of other constitutive physics on determining the presence of such a hook, particularly element diffusion, overshooting and nuclear reaction rates. We find that using the new solar abundance determinations, with low CNO abundances, makes it more difficult to reproduce the characteristic CMD of M67. This result is in agreement with results by VandenBerg et al. However, changes in the constitutive physics of the models, particularly overshooting, can influence and alter this result to the extent that isochrones constructed with models using low CNO solar abundances can also reproduce the TO morphology in M67. We conclude that only if all factors affecting the TO morphology are completely under control (and this is not the case), M67 could be used to put constraints on solar abundances.

  8. Drought response transcriptomes are altered in poplar with reduced tonoplast sucrose transporter expression

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Xue, Liang-Jiao; Frost, Christopher J.; Tsai, Chung-Jui

    Transgenic Populus tremula x alba (717-1B4) plants with reduced expression of a tonoplast sucrose efflux transporter, PtaSUT4, exhibit reduced shoot growth compared to wild type (WT) under sustained mild drought. The present study was undertaken to determine whether SUT4-RNAi directly or indirectly altered poplar predisposition and/or response to changes in soil water availability. While sucrose and hexose levels were constitutively elevated in shoot organs, expression responses to drought were most altered in the root tips of SUT4-RNAi plants. Prior to any drought treatment, constitutively elevated transcript levels of abscisic acid biosynthetic genes and bark/vegetative storage proteins suggested altered metabolism inmore » root tips of RNAi plants. Stronger drought-stimulation of stress-inducible genes encoding late-embryogenesis-abundant proteins in transgenic roots was consistent with increased vulnerability to soil drying. Transcript evidence suggested an RNAi effect on intercellular water trafficking by aquaporins in stem xylem during soil drying and recovery. Co-expression network analysis predicted altered integration of abscisic acid sensing/signaling with ethylene and jasmonate sensing/signaling in RNAi compared to WT roots. The overall conclusion is that steepened shoot-root sugar gradient in RNAi plants increased sensitivity of root tips to decreasing soil water availability.« less

  9. Drought response transcriptomes are altered in poplar with reduced tonoplast sucrose transporter expression

    DOE PAGES

    Xue, Liang-Jiao; Frost, Christopher J.; Tsai, Chung-Jui; ...

    2016-09-19

    Transgenic Populus tremula x alba (717-1B4) plants with reduced expression of a tonoplast sucrose efflux transporter, PtaSUT4, exhibit reduced shoot growth compared to wild type (WT) under sustained mild drought. The present study was undertaken to determine whether SUT4-RNAi directly or indirectly altered poplar predisposition and/or response to changes in soil water availability. While sucrose and hexose levels were constitutively elevated in shoot organs, expression responses to drought were most altered in the root tips of SUT4-RNAi plants. Prior to any drought treatment, constitutively elevated transcript levels of abscisic acid biosynthetic genes and bark/vegetative storage proteins suggested altered metabolism inmore » root tips of RNAi plants. Stronger drought-stimulation of stress-inducible genes encoding late-embryogenesis-abundant proteins in transgenic roots was consistent with increased vulnerability to soil drying. Transcript evidence suggested an RNAi effect on intercellular water trafficking by aquaporins in stem xylem during soil drying and recovery. Co-expression network analysis predicted altered integration of abscisic acid sensing/signaling with ethylene and jasmonate sensing/signaling in RNAi compared to WT roots. The overall conclusion is that steepened shoot-root sugar gradient in RNAi plants increased sensitivity of root tips to decreasing soil water availability.« less

  10. Constitutive Activation of the G-Protein Subunit G[alpha]s within Forebrain Neurons Causes PKA-Dependent Alterations in Fear Conditioning and Cortical "Arc" mRNA Expression

    ERIC Educational Resources Information Center

    Kelly, Michele P.; Cheung, York-Fong; Favilla, Christopher; Siegel, Steven J.; Kanes, Stephen J.; Houslay, Miles D.; Abel, Ted

    2008-01-01

    Memory formation requires cAMP signaling; thus, this cascade has been of great interest in the search for cognitive enhancers. Given that medications are administered long-term, we determined the effects of chronically increasing cAMP synthesis in the brain by expressing a constitutively active isoform of the G-protein subunit G[alpha]s…

  11. Contribution of MLH1 constitutional methylation for Lynch syndrome diagnosis in patients with tumor MLH1 downregulation.

    PubMed

    Pinto, Diana; Pinto, Carla; Guerra, Joana; Pinheiro, Manuela; Santos, Rui; Vedeld, Hege Marie; Yohannes, Zeremariam; Peixoto, Ana; Santos, Catarina; Pinto, Pedro; Lopes, Paula; Lothe, Ragnhild; Lind, Guro Elisabeth; Henrique, Rui; Teixeira, Manuel R

    2018-02-01

    Constitutional epimutation of the two major mismatch repair genes, MLH1 and MSH2, has been identified as an alternative mechanism that predisposes to the development of Lynch syndrome. In the present work, we aimed to investigate the prevalence of MLH1 constitutional methylation in colorectal cancer (CRC) patients with abnormal expression of the MLH1 protein in their tumors. In a series of 38 patients who met clinical criteria for Lynch syndrome genetic testing, with loss of MLH1 expression in the tumor and with no germline mutations in the MLH1 gene (35/38) or with tumors presenting the BRAF p.Val600Glu mutation (3/38), we screened for constitutional methylation of the MLH1 gene promoter using methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) in various biological samples. We found four (4/38; 10.5%) patients with constitutional methylation in the MLH1 gene promoter. RNA studies demonstrated decreased MLH1 expression in the cases with constitutional methylation when compared with controls. We could infer the mosaic nature of MLH1 constitutional hypermethylation in tissues originated from different embryonic germ layers, and in one family we could show that it occurred de novo. We conclude that constitutional MLH1 methylation occurs in a significant proportion of patients who have loss of MLH1 protein expression in their tumors and no MLH1 pathogenic germline mutation. Furthermore, we provide evidence that MLH1 constitutional hypermethylation is the molecular mechanism behind about 3% of Lynch syndrome families diagnosed in our institution, especially in patients with early onset or multiple primary tumors without significant family history. © 2018 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  12. Analysis of expressed sequence tags (ESTs) from avocado seed (Persea americana var. drymifolia) reveals abundant expression of the gene encoding the antimicrobial peptide snakin.

    PubMed

    Guzmán-Rodríguez, Jaquelina J; Ibarra-Laclette, Enrique; Herrera-Estrella, Luis; Ochoa-Zarzosa, Alejandra; Suárez-Rodríguez, Luis María; Rodríguez-Zapata, Luis C; Salgado-Garciglia, Rafael; Jimenez-Moraila, Beatriz; López-Meza, Joel E; López-Gómez, Rodolfo

    2013-09-01

    Avocado is one of the most important fruits in the world. Avocado "native mexicano" (Persea americana var. drymifolia) seeds are widely used in the propagation of this plant and are the primary source of rootstocks globally for a variety of avocado cultivars, such as the Hass avocado. Here, we report the isolation of 5005 ESTs from the 5' ends of P. americana var. drymifolia seed cDNA clones representing 1584 possible unigenes. These avocado seed ESTs were compared with the avocado flower EST library, and we detected several genes that are expressed either in both tissues or only in the seed. The snakin gene, which encodes an element of the innate immune response in plants, was one of those most frequently found among the seed ESTs, and this suggests that it is abundantly expressed in the avocado seed. We expressed the snakin gene in a heterologous system, namely the bovine endothelial cell line BVE-E6E7. Conditioned media from transfected BVE-E6E7 cells showed antimicrobial activity against strains of Escherichia coli and Staphylococcus aureus. This is the first study of the function of the snakin gene in plant seed tissue, and our observations suggest that this gene might play a protective role in the avocado seed. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  13. Computer simulation of the mathematical modeling involved in constitutive equation development: Via symbolic computations

    NASA Technical Reports Server (NTRS)

    Arnold, S. M.; Tan, H. Q.; Dong, X.

    1989-01-01

    Development of new material models for describing the high temperature constitutive behavior of real materials represents an important area of research in engineering disciplines. Derivation of mathematical expressions (constitutive equations) which describe this high temperature material behavior can be quite time consuming, involved and error prone; thus intelligent application of symbolic systems to facilitate this tedious process can be of significant benefit. A computerized procedure (SDICE) capable of efficiently deriving potential based constitutive models, in analytical form is presented. This package, running under MACSYMA, has the following features: partial differentiation, tensor computations, automatic grouping and labeling of common factors, expression substitution and simplification, back substitution of invariant and tensorial relations and a relational data base. Also limited aspects of invariant theory were incorporated into SDICE due to the utilization of potentials as a starting point and the desire for these potentials to be frame invariant (objective). Finally not only calculation of flow and/or evolutionary laws were accomplished but also the determination of history independent nonphysical coefficients in terms of physically measurable parameters, e.g., Young's modulus, was achieved. The uniqueness of SDICE resides in its ability to manipulate expressions in a general yet predefined order and simplify expressions so as to limit expression growth. Results are displayed when applicable utilizing index notation.

  14. Differentially enhanced insect resistance, at a cost, in Arabidopsis thaliana constitutively expressing a transcription factor of defensive metabolites.

    PubMed

    Johnson, Eric T; Dowd, Patrick F

    2004-08-11

    A transgenic line of Arabidopsis thaliana constitutively expressing a conserved MYB transcription factor of phenylpropanoid biosynthesis resulting in solid-purple leaves had significantly increased resistance to leaf feeding by first instar fall armyworms (Spodoptera frugiperda), but no enhanced resistance to cabbage looper (Trichoplusia ni) larvae, when compared to wild type plants. However, inflorescence and silique (seed pod) production were significantly reduced by 22 and 52%, respectively, in the transgenic line compared to wild type plants. Reduction in feeding by S. frugiperda was significantly positively correlated with reduction in weights of survivors, but both were negatively correlated with the concentration of anthocyanins. These results indicate that a single gene regulator can activate a defensive pathway sufficient to produce increased resistance to insects but that this activation confers a cost in plant productivity.

  15. Tax-Independent Constitutive IκB Kinase Activation in Adult T-Cell Leukemia Cells1

    PubMed Central

    Hironaka, Noriko; Mochida, Kanako; Mori, Naoki; Maeda, Michiyuki; Yamamoto, Naoki; Yamaoka, Shoji

    2004-01-01

    Abstract Adult T-cell leukemia (ATL) is a fatal T-cell malignancy that arises long after infection with human T-cell leukemia virus type I (HTLV-I). We reported previously that nuclear factor-κB (NF-κB) was constitutively activated in ATL cells, although expression of the viral proteins was barely detectable, including Tax, which was known to persistently activate NF-κB. Here we demonstrate that ATL cells that do not express detectable Tax protein exhibit constitutive IκB kinase (IKK) activity. Transfection studies revealed that a dominant-negative form of IKK1, and not of IKK2 or NF-κB essential modulator (NEMO), suppressed constitutive NFκB activity in ATL cells. This IKK activity was accompanied by elevated expression of p52, suggesting that the recently described noncanonical pathway of NF-κB activation operates in ATL cells. We finally show that specific inhibition of NF-κB by a super-repressor form of IκBα (SR-IκBα) in HTLV-I-infected T cells results in cell death regardless of Tax expression, providing definitive evidence of an essential role for NF-κB in the survival of ATL cells. In conclusion, the IKK complex is constitutively activated in ATL cells through a cellular mechanism distinct from that of Tax-mediated IKK activation. Further elucidation of this cellular mechanism should contribute to establishing a rationale for treatment of ATL. PMID:15153339

  16. Screening for MPL mutations in essential thrombocythemia and primary myelofibrosis: normal Mpl expression and absence of constitutive STAT3 and STAT5 activation in MPLW515L-positive platelets.

    PubMed

    Glembotsky, Ana C; Korin, Laura; Lev, Paola R; Chazarreta, Carlos D; Marta, Rosana F; Molinas, Felisa C; Heller, Paula G

    2010-05-01

    To evaluate the frequency of MPL W515L, W515K and S505N mutations in essential thrombocythemia (ET) and primary myelofibrosis (PMF) and to determine whether MPLW515L leads to impaired Mpl expression, constitutive STAT3 and STAT5 activation and enhanced response to thrombopoietin (TPO). Mutation detection was performed by allele-specific PCR and sequencing. Platelet Mpl expression was evaluated by flow cytometry, immunoblotting and real-time RT-PCR. Activation of STAT3 and STAT5 before and after stimulation with increasing concentrations of TPO was studied by immunoblotting. Plasma TPO was measured by ELISA. MPLW515L was detected in 1 of 100 patients with ET and 1 of 11 with PMF. Platelets from the PMF patient showed 100% mutant allele, which was <50% in platelets from the ET patient, who also showed the mutation in granulocytes, monocytes and B cells. Mpl surface and total protein expression were normal, and TPO levels were mildly increased in the MPLW515L-positive ET patient, while MPL transcripts did not differ from controls in both MPLW515L-positive patients. Constitutive STAT3 and STAT5 phosphorylation was absent and dose response to TPO-induced phosphorylation was not enhanced. The low frequency of MPL mutations in this cohort is in agreement with previous studies. The finding of normal Mpl levels in MPLW515L-positive platelets indicates this mutation does not lead to dysregulated Mpl expression, as frequently shown for myeloproliferative neoplasms. The lack of spontaneous STAT3 and STAT5 activation and the normal response to TPO is unexpected as MPLW515L leads to constitutive receptor activation and hypersensitivity to TPO in experimental models.

  17. Constitutive TL1A expression under colitogenic conditions modulates the severity and location of gut mucosal inflammation and induces fibrostenosis.

    PubMed

    Barrett, Robert; Zhang, Xiaolan; Koon, Hon Wai; Vu, Michelle; Chang, Jyh-Yau; Yeager, Nicole; Nguyen, Mary Ann; Michelsen, Kathrin S; Berel, Dror; Pothoulakis, Charalabos; Targan, Stephan R; Shih, David Q

    2012-02-01

    Intestinal fibrostenosis is a hallmark of severe Crohn's disease and can lead to multiple surgeries. Patients with certain TNFSF15 variants overexpress TL1A. The aim of this study was to determine the effect of TL1A overexpression on intestinal inflammation and the development of fibrostenosis. We assessed the in vivo consequences of constitutive TL1A expression on gut mucosal inflammation and fibrostenosis using two murine models of chronic colitis. In the dextran sodium sulfate (DSS) and adoptive T-cell transfer models, there was proximal migration of colonic inflammation, worsened patchy intestinal inflammation, and long gross intestinal strictures in Tl1a transgenic compared to wild-type littermates. In the DSS model, myeloid- and T-cell-expressing Tl1a transgenic mice had increased T-cell activation markers and interleukin-17 expression compared to wild-type mice. In the T-cell transfer model, Rag1(-/-) mice receiving Tl1a transgenic T cells had increased interferon-γ expression but reduced T-helper 17 cells and IL-17 production. Narrowed ureters with hydronephrosis were found only in the Tl1a transgenic mice in all chronic colitis models. In human translational studies, Crohn's disease patients with higher peripheral TL1A expression also exhibited intestinal fibrostenosis and worsened ileocecal inflammation with relative sparing of rectosigmoid inflammation. These data show that TL1A is an important cytokine that not only modulates the location and severity of mucosal inflammation, but also induces fibrostenosis. Copyright © 2012 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  18. Cloning and characterization of full length of a novel zebrafish gene Zsrg abundantly expressed in the germline stem cells.

    PubMed

    Lv, Daoyuan; Song, Ping; Chen, Yungui; Gong, Wuming; Mo, Saijun

    2005-04-08

    Using the digital differential display program of the National Center for Biotechnology Information, we identified a contig of expression sequence tags (ESTs) (Accession No. BM316936), which came from zebrafish ovary and testis libraries. The full-length cDNA of this transcript was cloned and further confirmed by polymerase chain reaction and sequencing. The full-length cDNA of the novel gene is 807bp and encodes a novel protein of 187 amino acids, which shares no significant homology with any other known proteins. Characterization of genomic sequences of the gene revealed that it spans 6kb on the linkage group 3 and is composed of five exons and four introns. RT-PCR analysis showed that it was expressed in mature oocytes and one-cell stage, and persisted until 24h of development. RT-PCR also revealed that it is expressed in gonad and kidney, with the highest level of expression in the testis. The expression sites of the novel gene in adult gonad were further localized by in situ hybridization to oogonia and growing oocytes in ovary and to spermatogonia, spermatocytes but not to spermatids in testis. Based on its abundance in testis and the germline stem cell-spermatogonia and oogonia, we hypothesize that it may function as a testicular development and gametogenesis related gene that plays important roles in spermatogenesis, and named it Zsrg (zebrafish testis spermatogenesis related gene, Zsrg).

  19. Ayurvedic genomics, constitutional psychology, and endocrinology: the missing connection.

    PubMed

    Rizzo-Sierra, Carlos V

    2011-05-01

    A recent methodological approach for human classification, diagnosis, and therapeutics through the combination of current Western constitutional psychology somatotypes and traditional Indian medicine (prakriti) body types and mind (manas) is herein presented. The striking similarities between psychologic somatotypes and Indian medicine body types permits proposal of a finite genopsycho-somatotyping of humans. Genopsycho-somatotyping of humans consists of a set of common physiologic, physical, and psychologic attributes related to a common basic birth constitution that remains somewhat permanent during human lifetime, since it is proposed that this birth constitution is programmed in the person's DNA (genes). This mainly provides a tool for classifying the human population based on broad and finite phenotype clusters across different ethnicity, languages, geographical location, or self-reported ancestry. In spite of any social or environmental traumatic event, I propose for males that every basic constitution has an associated identification organ, a measured property or marker, a soma, and some psyche general tendencies suggesting specific behavior or recurrent conduct. Three (3) basic extreme genopsycho-somatotypes or birth constitutions are enunciated: mesomorphic or andrus (Pitta), endomorphic or thymus (Khapa), and ectomorphic or thyrus (Vata). The method further predicts that male andrus constitution across races shares similarities in androgen (An) nuclear receptor behavior, whereas thymus constitutions are mainly regulated by T-cells (Tc) nuclear receptor behavior. Moreover, it suggests that thyrus constitutions share similarities in thyroxine (Th) nuclear receptor behavior. These proposed nuclear receptors are expected to regulate the expression of specific genes, thereby controlling the embryonic development, adult homeostasis, and metabolism of the human organism in a very profound way. The method finally predicts small differences in measured property

  20. Two novel heat-soluble protein families abundantly expressed in an anhydrobiotic tardigrade.

    PubMed

    Yamaguchi, Ayami; Tanaka, Sae; Yamaguchi, Shiho; Kuwahara, Hirokazu; Takamura, Chizuko; Imajoh-Ohmi, Shinobu; Horikawa, Daiki D; Toyoda, Atsushi; Katayama, Toshiaki; Arakawa, Kazuharu; Fujiyama, Asao; Kubo, Takeo; Kunieda, Takekazu

    2012-01-01

    Tardigrades are able to tolerate almost complete dehydration by reversibly switching to an ametabolic state. This ability is called anhydrobiosis. In the anhydrobiotic state, tardigrades can withstand various extreme environments including space, but their molecular basis remains largely unknown. Late embryogenesis abundant (LEA) proteins are heat-soluble proteins and can prevent protein-aggregation in dehydrated conditions in other anhydrobiotic organisms, but their relevance to tardigrade anhydrobiosis is not clarified. In this study, we focused on the heat-soluble property characteristic of LEA proteins and conducted heat-soluble proteomics using an anhydrobiotic tardigrade. Our heat-soluble proteomics identified five abundant heat-soluble proteins. All of them showed no sequence similarity with LEA proteins and formed two novel protein families with distinct subcellular localizations. We named them Cytoplasmic Abundant Heat Soluble (CAHS) and Secretory Abundant Heat Soluble (SAHS) protein families, according to their localization. Both protein families were conserved among tardigrades, but not found in other phyla. Although CAHS protein was intrinsically unstructured and SAHS protein was rich in β-structure in the hydrated condition, proteins in both families changed their conformation to an α-helical structure in water-deficient conditions as LEA proteins do. Two conserved repeats of 19-mer motifs in CAHS proteins were capable to form amphiphilic stripes in α-helices, suggesting their roles as molecular shield in water-deficient condition, though charge distribution pattern in α-helices were different between CAHS and LEA proteins. Tardigrades might have evolved novel protein families with a heat-soluble property and this study revealed a novel repertoire of major heat-soluble proteins in these anhydrobiotic animals.

  1. Human eosinophils constitutively express a unique serine protease, PRSS33.

    PubMed

    Toyama, Sumika; Okada, Naoko; Matsuda, Akio; Morita, Hideaki; Saito, Hirohisa; Fujisawa, Takao; Nakae, Susumu; Karasuyama, Hajime; Matsumoto, Kenji

    2017-07-01

    Eosinophils play important roles in asthma, especially airway remodeling, by producing various granule proteins, chemical mediators, cytokines, chemokines and proteases. However, protease production by eosinophils is not fully understood. In the present study, we investigated the production of eosinophil-specific proteases/proteinases by transcriptome analysis. Human eosinophils and other cells were purified from peripheral blood by density gradient sedimentation and negative/positive selections using immunomagnetic beads. Protease/proteinase expression in eosinophils and release into the supernatant were evaluated by microarray analysis, qPCR, ELISA, flow cytometry and immunofluorescence staining before and after stimulation with eosinophil-activating cytokines and secretagogues. mRNAs for extracellular matrix proteins in human normal fibroblasts were measured by qPCR after exposure to recombinant protease serine 33 (PRSS33) protein (rPRSS33), created with a baculovirus system. Human eosinophils expressed relatively high levels of mRNA for metalloproteinase 25 (MMP25), a disintegrin and metalloprotease 8 (ADAM8), ADAM10, ADAM19 and PRSS33. Expression of PRSS33 was the highest and eosinophil-specific. PRSS33 mRNA expression was not affected by eosinophil-activating cytokines. Immunofluorescence staining showed that PRSS33 was co-localized with an eosinophil granule protein. PRSS33 was not detected in the culture supernatant of eosinophils even after stimulation with secretagogues, but its cell surface expression was increased. rPRSS33 stimulation of human fibroblasts increased expression of collagen and fibronectin mRNAs, at least in part via protease-activated receptor-2 activation. Activated eosinophils may induce fibroblast extracellular matrix protein synthesis via cell surface expression of PRSS33, which would at least partly explain eosinophils' role(s) in airway remodeling. Copyright © 2017 Japanese Society of Allergology. Production and hosting by Elsevier

  2. Complementary DNA cloning and constitutive expression of cytochrome P450 1C1 in the gills of carp (Cyprinus carpio).

    PubMed

    Itakura, Takao; El-Kady, Mohamed; Mitsuo, Ryoichi; Kaminishi, Yoshio

    2005-01-01

    Cytochrome P450 (CYP) enzymes constitute a multigene family of many endogenous and xenobiotic substances. The CYP1 family is of particular interest in environmental toxicology because its members are dominant in the metabolism of polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), and aryl amines. A new complementary DNA of the CYP1C subfamily encoding CYP1C1 was isolated from carp liver after intraperitoneal injection of beta-napthoflavone (BNF). The full-length cDNA obtained contained a 5' noncoding region of 244 bp, an open reading frame of 1572 bp coding for 524 amino acids, a stop codon, and a 3' noncoding region of 965 bp. The predicted molecular weight of the protein was approximately 59.3 kDa. The deduced amino acid sequence of this cDNA was 82.1% and 80.2% similar to Japanese eel and scup CYP1C1 sequences, respectively, while it exhibited a similarity of 74.9% with the scup CYP1C2 sequence. The deduced amino acid sequence of carp CYP1C1 showed similarities with those of the reported CYP1B1s of teleosts and mammals of 48.4, 48.8, 48.2, 48.6, 45.3, and 45.5% for carp CYP1B1, carp CYP1B2, plaice CYP1B1, and human, rat, and mouse CYP1B1, respectively. The phylogenetic tree constructed using fish and mammalian CYP1 sequences suggested a closer relationship of the CYP1C subfamily to CYP1B than to CYP1A. The tree showed the possibility of the existence of CYP1C subfamily genes in mammalian species. Northern blot analysis for the liver, intestine, gills, and kidney showed no detectable induced expression but constitutive expression in the gill organs.

  3. Carbon Costs of Constitutive and Expressed Resistance to a Non-Native Pathogen in Limber Pine.

    PubMed

    Vogan, Patrick J; Schoettle, Anna W

    2016-01-01

    Increasing the frequency of resistance to the non-native fungus Cronartium ribicola (causative agent of white pine blister rust, WPBR) in limber pine populations is a primary management objective to sustain high-elevation forest communities. However, it is not known to what extent genetic disease resistance is costly to plant growth or carbon economy. In this study, we measured growth and leaf-level physiology in (1) seedling families from seed trees that have previously been inferred to carry or not carry Cr4, the dominant R gene allele conferring complete, gene-for-gene resistance to WPBR in limber pine, and (2) populations that were and were not infected with C. ribicola. We found that, in the absence of C. ribicola exposure, there was no significant difference in carbon relations between families born from seed trees that harbor the resistance allele compared to those that lack it, either to plant growth and phenology or leaf-level photosynthetic traits. However, post-infection with C. ribicola, growth was significantly reduced in inoculation survivors expressing complete resistance compared to uninoculated seedlings. Furthermore, inoculation survivors exhibited significant increases in a suite of traits including photosynthetic rate, respiration rate, leaf N, and stomatal conductance and a decrease in photosynthetic water-use efficiency. The lack of constitutive carbon costs associated with Cr4 resistance in non-stressed limber pine is consistent with a previous report that the R gene allele is not under selection in the absence of C. ribicola and suggests that host resistance may not bear a constitutive cost in pathosystems that have not coevolved. However, under challenge by C. ribicola, complete resistance to WPBR in limber pine has a significant cost to plant growth, though enhanced carbon acquisition post-infection may offset this somewhat. These costs and effects on performance further complicate predictions of this species' response in warmer future

  4. Carbon Costs of Constitutive and Expressed Resistance to a Non-Native Pathogen in Limber Pine

    PubMed Central

    2016-01-01

    Increasing the frequency of resistance to the non-native fungus Cronartium ribicola (causative agent of white pine blister rust, WPBR) in limber pine populations is a primary management objective to sustain high-elevation forest communities. However, it is not known to what extent genetic disease resistance is costly to plant growth or carbon economy. In this study, we measured growth and leaf-level physiology in (1) seedling families from seed trees that have previously been inferred to carry or not carry Cr4, the dominant R gene allele conferring complete, gene-for-gene resistance to WPBR in limber pine, and (2) populations that were and were not infected with C. ribicola. We found that, in the absence of C. ribicola exposure, there was no significant difference in carbon relations between families born from seed trees that harbor the resistance allele compared to those that lack it, either to plant growth and phenology or leaf-level photosynthetic traits. However, post-infection with C. ribicola, growth was significantly reduced in inoculation survivors expressing complete resistance compared to uninoculated seedlings. Furthermore, inoculation survivors exhibited significant increases in a suite of traits including photosynthetic rate, respiration rate, leaf N, and stomatal conductance and a decrease in photosynthetic water-use efficiency. The lack of constitutive carbon costs associated with Cr4 resistance in non-stressed limber pine is consistent with a previous report that the R gene allele is not under selection in the absence of C. ribicola and suggests that host resistance may not bear a constitutive cost in pathosystems that have not coevolved. However, under challenge by C. ribicola, complete resistance to WPBR in limber pine has a significant cost to plant growth, though enhanced carbon acquisition post-infection may offset this somewhat. These costs and effects on performance further complicate predictions of this species’ response in warmer future

  5. Field tolerance to fungal pathogens of Brassica napus constitutively expressing a chimeric chitinase gene

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Grison, R.; Grezes-Besset, B.; Lucante, N.

    1996-05-01

    Constitutive overexpression of a protein involved in plant defense mechanisms to disease is one of the strategies proposed to increase plant tolerance to fungal pathogens. A hybrid endochitinase gene under a constitutive promoter was introduced by Agrobacterium-mediated transformation into a winter-type oilseed rape (Brassica napus var. oleifera) inbred line. Progeny from transformed plants was challenged using three different fungal pathogens (Cylindrosporium concentricum, Phoma lingam, Sclerotinia sclerotiorum) in field trials at two different geographical locations. These plants exhibited an increased tolerance to disease as compared with the nontransgenic parental plants. 31 refs., 1 fig., 2 tabs.

  6. Implications of abundant hygroscopic minerals in the Martian regolith

    NASA Technical Reports Server (NTRS)

    Clark, B. C.

    1978-01-01

    Converging lines of evidence suggest that a significant portion of the Martian surface fines may consist of salts and smectite clays. Salts can form stoichiometric hydrates as well as eutectic solutions with depressed freezing points; clays contain bound water of constitution and adsorb significant quantities of water from the vapor phase. The formation of ice may be suppressed by these minerals in some regions on Mars, and their presence in abundance would imply important consequences for atmospheric and geologic processes and the prospects for exobiology.

  7. Acetaminophen Modulates P-Glycoprotein Functional Expression at the Blood-Brain Barrier by a Constitutive Androstane Receptor–Dependent Mechanism

    PubMed Central

    Thompson, Brandon J.; Sanchez-Covarrubias, Lucy; Zhang, Yifeng; Laracuente, Mei-Li; Vanderah, Todd W.; Ronaldson, Patrick T.; Davis, Thomas P.

    2013-01-01

    Effective pharmacologic treatment of pain with opioids requires that these drugs attain efficacious concentrations in the central nervous system (CNS). A primary determinant of CNS drug permeation is P-glycoprotein (P-gp), an endogenous blood-brain barrier (BBB) efflux transporter that is involved in brain-to-blood transport of opioid analgesics (i.e., morphine). Recently, the nuclear receptor constitutive androstane receptor (CAR) has been identified as a regulator of P-gp functional expression at the BBB. This is critical to pharmacotherapy of pain/inflammation, as patients are often administered acetaminophen (APAP), a CAR-activating ligand, in conjunction with an opioid. Our objective was to investigate, in vivo, the role of CAR in regulation of P-gp at the BBB. Following APAP treatment, P-gp protein expression was increased up to 1.4–1.6-fold in a concentration-dependent manner. Additionally, APAP increased P-gp transport of BODIPY-verapamil in freshly isolated rat brain capillaries. This APAP-induced increase in P-gp expression and activity was attenuated in the presence of CAR pathway inhibitor okadaic acid or transcriptional inhibitor actinomycin D, suggesting P-gp regulation is CAR-dependent. Furthermore, morphine brain accumulation was enhanced by P-gp inhibitors in APAP-treated animals, suggesting P-gp–mediated transport. A warm-water (50°C) tail-flick assay revealed a significant decrease in morphine analgesia in animals treated with morphine 3 or 6 hours after APAP treatment, as compared with animals treated concurrently. Taken together, our data imply that inclusion of APAP in a pain treatment regimen activates CAR at the BBB and increases P-gp functional expression, a clinically significant drug-drug interaction that modulates opioid analgesic efficacy. PMID:24019224

  8. Predicting the Dynamics of Protein Abundance

    PubMed Central

    Mehdi, Ahmed M.; Patrick, Ralph; Bailey, Timothy L.; Bodén, Mikael

    2014-01-01

    Protein synthesis is finely regulated across all organisms, from bacteria to humans, and its integrity underpins many important processes. Emerging evidence suggests that the dynamic range of protein abundance is greater than that observed at the transcript level. Technological breakthroughs now mean that sequencing-based measurement of mRNA levels is routine, but protocols for measuring protein abundance remain both complex and expensive. This paper introduces a Bayesian network that integrates transcriptomic and proteomic data to predict protein abundance and to model the effects of its determinants. We aim to use this model to follow a molecular response over time, from condition-specific data, in order to understand adaptation during processes such as the cell cycle. With microarray data now available for many conditions, the general utility of a protein abundance predictor is broad. Whereas most quantitative proteomics studies have focused on higher organisms, we developed a predictive model of protein abundance for both Saccharomyces cerevisiae and Schizosaccharomyces pombe to explore the latitude at the protein level. Our predictor primarily relies on mRNA level, mRNA–protein interaction, mRNA folding energy and half-life, and tRNA adaptation. The combination of key features, allowing for the low certainty and uneven coverage of experimental observations, gives comparatively minor but robust prediction accuracy. The model substantially improved the analysis of protein regulation during the cell cycle: predicted protein abundance identified twice as many cell-cycle-associated proteins as experimental mRNA levels. Predicted protein abundance was more dynamic than observed mRNA expression, agreeing with experimental protein abundance from a human cell line. We illustrate how the same model can be used to predict the folding energy of mRNA when protein abundance is available, lending credence to the emerging view that mRNA folding affects translation

  9. Predicting the dynamics of protein abundance.

    PubMed

    Mehdi, Ahmed M; Patrick, Ralph; Bailey, Timothy L; Bodén, Mikael

    2014-05-01

    Protein synthesis is finely regulated across all organisms, from bacteria to humans, and its integrity underpins many important processes. Emerging evidence suggests that the dynamic range of protein abundance is greater than that observed at the transcript level. Technological breakthroughs now mean that sequencing-based measurement of mRNA levels is routine, but protocols for measuring protein abundance remain both complex and expensive. This paper introduces a Bayesian network that integrates transcriptomic and proteomic data to predict protein abundance and to model the effects of its determinants. We aim to use this model to follow a molecular response over time, from condition-specific data, in order to understand adaptation during processes such as the cell cycle. With microarray data now available for many conditions, the general utility of a protein abundance predictor is broad. Whereas most quantitative proteomics studies have focused on higher organisms, we developed a predictive model of protein abundance for both Saccharomyces cerevisiae and Schizosaccharomyces pombe to explore the latitude at the protein level. Our predictor primarily relies on mRNA level, mRNA-protein interaction, mRNA folding energy and half-life, and tRNA adaptation. The combination of key features, allowing for the low certainty and uneven coverage of experimental observations, gives comparatively minor but robust prediction accuracy. The model substantially improved the analysis of protein regulation during the cell cycle: predicted protein abundance identified twice as many cell-cycle-associated proteins as experimental mRNA levels. Predicted protein abundance was more dynamic than observed mRNA expression, agreeing with experimental protein abundance from a human cell line. We illustrate how the same model can be used to predict the folding energy of mRNA when protein abundance is available, lending credence to the emerging view that mRNA folding affects translation efficiency

  10. Constitutional and Non-Constitutional Governments...Similarities and Differences throughout History. Resource Packet.

    ERIC Educational Resources Information Center

    Pallasch, Brian Thomas

    This civic education resource packet is designed to provide teachers, community leaders, and other civic educators with an understanding of the differences between constitutional and non-constitutional governments. Six papers discussing the topic are included: "The Differences bewteen Constitutional and Non-Constitutional Governments" (John…

  11. Identification of chimeric antigen receptors that mediate constitutive or inducible proliferation of T cells

    PubMed Central

    Frigault, Matthew J; Lee, Jihyun; Basil, Maria Ciocca; Carpenito, Carmine; Motohashi, Shinichiro; Scholler, John; Kawalekar, Omkar U.; Guedan, Sonia; McGettigan, Shannon E.; Posey, Avery D.; Ang, Sonny; Cooper, Laurence J. N.; Platt, Jesse M.; Johnson, F. Brad; Paulos, Chrystal M; Zhao, Yangbing; Kalos, Michael; Milone, Michael C.; June, Carl H.

    2015-01-01

    This study compared second generation chimeric antigen receptors encoding signaling domains composed of CD28, ICOS and 4-1BB. Here we report that certain CARs endow T cells with the ability to undergo long-term autonomous proliferation. Transduction of primary human T-cell with lentiviral vectors encoding some of the CARs resulted in sustained proliferation for up to three months following a single stimulation through the TCR. Sustained numeric expansion was independent of cognate antigen and did not require the addition of exogenous cytokines or feeder cells after a single stimulation of the TCR and CD28. Results from gene array and functional assays linked sustained cytokine secretion and expression of T-bet, EOMES and GATA-3 to the effect. Sustained expression of the endogenous IL2 locus has not been reported in primary T cells. Sustained proliferation was dependent on CAR structure and high expression, the latter of which was necessary but not sufficient. The mechanism involves constitutive signaling through NF-kB, Akt, Erk and NFAT. The propagated CAR T cells retained a diverse TCR repertoire and cellular transformation was not observed. The CARs with a constitutive growth phenotype displayed inferior antitumor effects and engraftment in vivo. Therefore the design of CARs that have a non-constitutive growth phenotype may be a strategy to improve efficacy and engraftment of CAR T cells. The identification of CARs that confer constitutive or non-constitutive growth patterns may explain observations that CAR T cells have differential survival patterns in clinical trials. PMID:25600436

  12. Ortho-aminoazotoluene activates mouse constitutive androstane receptor (mCAR) and increases expression of mCAR target genes

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Smetanina, Mariya A., E-mail: maria.smetanina@gmail.com; Laboratory of Gene Expression Control, Institute of Cytology and Genetics of the Siberian Branch of the Russian Academy of Sciences, prospekt Lavrentyeva 10, Novosibirsk 630090; Group of Pharmacogenomics, Institute of Chemical Biology and Fundamental Medicine of the Siberian Branch of the Russian Academy of Sciences, prospekt Lavrentyeva 8, Novosibirsk 630090

    2'-3-dimethyl-4-aminoazobenzene (ortho-aminoazotoluene, OAT) is an azo dye and a rodent carcinogen that has been evaluated by the International Agency for Research on Cancer (IARC) as a possible (class 2B) human carcinogen. Its mechanism of action remains unclear. We examined the role of the xenobiotic receptor Constitutive Androstane Receptor (CAR, NR1I3) as a mediator of the effects of OAT. We found that OAT increases mouse CAR (mCAR) transactivation in a dose-dependent manner. This effect is specific because another closely related azo dye, 3'-methyl-4-dimethyl-aminoazobenzene (3'MeDAB), did not activate mCAR. Real-time Q-PCR analysis in wild-type C57BL/6 mice revealed that OAT induces the hepaticmore » mRNA expression of the following CAR target genes: Cyp2b10, Cyp2c29, Cyp3a11, Ugt1a1, Mrp4, Mrp2 and c-Myc. CAR-null (Car{sup -/-}) mice showed no increased expression of these genes following OAT treatment, demonstrating that CAR is required for their OAT dependent induction. The OAT-induced CAR-dependent increase of Cyp2b10 and c-Myc expression was confirmed by Western blotting. Immunohistochemistry analysis of wild-type and Car{sup -/-} livers showed that OAT did not acutely induce hepatocyte proliferation, but at much later time points showed an unexpected CAR-dependent proliferative response. These studies demonstrate that mCAR is an OAT xenosensor, and indicate that at least some of the biological effects of this compound are mediated by this nuclear receptor. - Highlights: > The azo dye and mouse carcinogen OAT is a very effective mCAR activator. > OAT increases mCAR transactivation in a dose-dependent manner. > OAT CAR-dependently increases the expression of a specific subset of CAR target genes. > OAT induces an unexpectedly deferred, but CAR-dependent hepatocyte proliferation.« less

  13. Counteracting wetland overgrowth increases breeding and staging bird abundances

    NASA Astrophysics Data System (ADS)

    Lehikoinen, Petteri; Lehikoinen, Aleksi; Mikkola-Roos, Markku; Jaatinen, Kim

    2017-01-01

    Human actions have led to loss and degradation of wetlands, impairing their suitability as habitat especially for waterbirds. Such negative effects may be mitigated through habitat management. To date scientific evidence regarding the impacts of these actions remains scarce. We studied guild specific abundances of breeding and staging birds in response to habitat management on 15 Finnish wetlands. In this study management actions comprised several means of vegetation removal to thwart overgrowth. Management cost efficiency was assessed by examining the association between site-specific costs and bird abundances. Several bird guilds exhibited positive connections with both habitat management as well as with invested funds. Most importantly, however, red-listed species and species with special conservation concern as outlined by the EU showed positive correlations with management actions, underlining the conservation value of wetland management. The results suggest that grazing was especially efficient in restoring overgrown wetlands. As a whole this study makes it clear that wetland habitat management constitutes a feasible conservation tool. The marked association between invested funds and bird abundance may prove to be a valuable tool for decision makers when balancing costs and impact of conservation measures against one another.

  14. Constitutively Expressed IFITM3 Protein in Human Endothelial Cells Poses an Early Infection Block to Human Influenza Viruses

    PubMed Central

    Sun, Xiangjie; Zeng, Hui; Kumar, Amrita; Belser, Jessica A.; Maines, Taronna R.

    2016-01-01

    ABSTRACT A role for pulmonary endothelial cells in the orchestration of cytokine production and leukocyte recruitment during influenza virus infection, leading to severe lung damage, has been recently identified. As the mechanistic pathway for this ability is not fully known, we extended previous studies on influenza virus tropism in cultured human pulmonary endothelial cells. We found that a subset of avian influenza viruses, including potentially pandemic H5N1, H7N9, and H9N2 viruses, could infect human pulmonary endothelial cells (HULEC) with high efficiency compared to human H1N1 or H3N2 viruses. In HULEC, human influenza viruses were capable of binding to host cellular receptors, becoming internalized and initiating hemifusion but failing to uncoat the viral nucleocapsid and to replicate in host nuclei. Unlike numerous cell types, including epithelial cells, we found that pulmonary endothelial cells constitutively express a high level of the restriction protein IFITM3 in endosomal compartments. IFITM3 knockdown by small interfering RNA (siRNA) could partially rescue H1N1 virus infection in HULEC, suggesting IFITM3 proteins were involved in blocking human influenza virus infection in endothelial cells. In contrast, selected avian influenza viruses were able to escape IFITM3 restriction in endothelial cells, possibly by fusing in early endosomes at higher pH or by other, unknown mechanisms. Collectively, our study demonstrates that the human pulmonary endothelium possesses intrinsic immunity to human influenza viruses, in part due to the constitutive expression of IFITM3 proteins. Notably, certain avian influenza viruses have evolved to escape this restriction, possibly contributing to virus-induced pneumonia and severe lung disease in humans. IMPORTANCE Avian influenza viruses, including H5N1 and H7N9, have been associated with severe respiratory disease and fatal outcomes in humans. Although acute respiratory distress syndrome (ARDS) and progressive pulmonary

  15. Constitutive expression of CaPLA1 conferred enhanced growth and grain yield in transgenic rice plants.

    PubMed

    Park, Ki Youl; Kim, Eun Yu; Seo, Young Sam; Kim, Woo Taek

    2016-03-01

    Phospholipids are not only important components of cell membranes, but participate in diverse processes in higher plants. In this study, we generated Capsicum annuum phospholipiase A1 (CaPLA1) overexpressing transgenic rice (Oryza sativa L.) plants under the control of the maize ubiquitin promoter. The T4 CaPLA1-overexpressing rice plants (Ubi:CaPLA1) had a higher root:shoot mass ratio than the wild-type plants in the vegetative stage. Leaf epidermal cells from transgenic plants had more cells than wild-type plants. Genes that code for cyclin and lipid metabolic enzymes were up-regulated in the transgenic lines. When grown under typical paddy field conditions, the transgenic plants produced more tillers, longer panicles and more branches per panicle than the wild-type plants, all of which resulted in greater grain yield. Microarray analysis suggests that gene expressions that are related with cell proliferation, lipid metabolism, and redox state were widely altered in CaPLA1-overexpressing transgenic rice plants. Ubi:CaPLA1 plants had a reduced membrane peroxidation state, as determined by malondialdehyde and conjugated diene levels and higher peroxidase activity than wild-type rice plants. Furthermore, three isoprenoid synthetic genes encoding terpenoid synthase, hydroxysteroid dehydrogenase and 3-hydroxy-3-methyl-glutaryl-CoA reductase were up-regulated in CaPLA1-overexpressing plants. We suggest that constitutive expression of CaPLA1 conferred increased grain yield with enhanced growth in transgenic rice plants by alteration of gene activities related with cell proliferation, lipid metabolism, membrane peroxidation state and isoprenoid biosynthesis.

  16. RNA-Seq profiling of single bovine oocyte transcript abundance and its modulation by cytoplasmic polyadenylation.

    PubMed

    Reyes, Juan M; Chitwood, James L; Ross, Pablo J

    2015-02-01

    Molecular changes occurring during mammalian oocyte maturation are partly regulated by cytoplasmic polyadenylation (CP) and affect oocyte quality, yet the extent of CP activity during oocyte maturation remains unknown. Single bovine oocyte RNA sequencing (RNA-Seq) was performed to examine changes in transcript abundance during in vitro oocyte maturation in cattle. Polyadenylated RNA from individual germinal-vesicle and metaphase-II oocytes was amplified and processed for Illumina sequencing, producing approximately 30 million reads per replicate for each sample type. A total of 10,494 genes were found to be expressed, of which 2,455 were differentially expressed (adjusted P < 0.05 and fold change >2) between stages, with 503 and 1,952 genes respectively increasing and decreasing in abundance. Differentially expressed genes with complete 3'-untranslated-region sequence (279 increasing and 918 decreasing in polyadenylated transcript abundance) were examined for the presence, position, and distribution of motifs mediating CP, revealing enrichment (85%) and lack thereof (18%) in up- and down-regulated genes, respectively. Examination of total and polyadenylated RNA abundance by quantitative PCR validated these RNA-Seq findings. The observed increases in polyadenylated transcript abundance within the RNA-Seq data are likely due to CP, providing novel insight into targeted transcripts and resultant differential gene expression profiles that contribute to oocyte maturation. © 2015 Wiley Periodicals, Inc.

  17. Cell stress and translational inhibitors transiently increase the abundance of mammalian SINE transcripts.

    PubMed Central

    Liu, W M; Chu, W M; Choudary, P V; Schmid, C W

    1995-01-01

    The abundance of Alu RNA is transiently increased by heat shock in human cell lines. This effect is specific to Alu repeats among Pol III transcribed genes, since the abundance of 7SL, 7SK, 5S and U6 RNAs is essentially unaffected by heat shock. The rapid induction of Alu expression precedes the heat shock induction of mRNAs for the ubiquitin and HSP 70 heat shock genes. Heat shock mimetics also transiently induce Alu expression indicating that increased Alu expression is a general cell-stress response. Cycloheximide treatment rapidly and transiently increases the abundance of Alu RNA. Again, compared with other genes transcribed by Pol III, this increase is specific to Alu. However, as distinguished from the cell stress response, cycloheximide does not induce expression of HSP 70 and ubiquitin mRNAs. Puromycin also increases Alu expression, suggesting that this response is generally caused by translational inhibition. The response of mammalian SINEs to cell stress and translational inhibition is not limited to SINEs which are Alu homologues. Heat shock and cycloheximide each transiently induce Pol III directed expression of B1 and B2 RNAs in mouse cells and C-element RNA in rabbit cells. Together, these three species exemplify the known SINE composition of placental mammals, suggesting that mammalian SINEs are similarly regulated and may serve a common function. Images PMID:7784180

  18. MicroRNAs show a wide diversity of expression profiles in the developing and mature central nervous system

    PubMed Central

    Kapsimali, Marika; Kloosterman, Wigard P; de Bruijn, Ewart; Rosa, Frederic; Plasterk, Ronald HA; Wilson, Stephen W

    2007-01-01

    Background MicroRNA (miRNA) encoding genes are abundant in vertebrate genomes but very few have been studied in any detail. Bioinformatic tools allow prediction of miRNA targets and this information coupled with knowledge of miRNA expression profiles facilitates formulation of hypotheses of miRNA function. Although the central nervous system (CNS) is a prominent site of miRNA expression, virtually nothing is known about the spatial and temporal expression profiles of miRNAs in the brain. To provide an overview of the breadth of miRNA expression in the CNS, we performed a comprehensive analysis of the neuroanatomical expression profiles of 38 abundant conserved miRNAs in developing and adult zebrafish brain. Results Our results show miRNAs have a wide variety of different expression profiles in neural cells, including: expression in neuronal precursors and stem cells (for example, miR-92b); expression associated with transition from proliferation to differentiation (for example, miR-124); constitutive expression in mature neurons (miR-124 again); expression in both proliferative cells and their differentiated progeny (for example, miR-9); regionally restricted expression (for example, miR-222 in telencephalon); and cell-type specific expression (for example, miR-218a in motor neurons). Conclusion The data we present facilitate prediction of likely modes of miRNA function in the CNS and many miRNA expression profiles are consistent with the mutual exclusion mode of function in which there is spatial or temporal exclusion of miRNAs and their targets. However, some miRNAs, such as those with cell-type specific expression, are more likely to be co-expressed with their targets. Our data provide an important resource for future functional studies of miRNAs in the CNS. PMID:17711588

  19. The Constitutional Amendment Process

    ERIC Educational Resources Information Center

    Chism, Kahlil

    2005-01-01

    This article discusses the constitutional amendment process. Although the process is not described in great detail, Article V of the United States Constitution allows for and provides instruction on amending the Constitution. While the amendment process currently consists of six steps, the Constitution is nevertheless quite difficult to change.…

  20. Constitutional MLH1 methylation presenting with colonic polyposis syndrome and not Lynch syndrome.

    PubMed

    Kidambi, Trilokesh D; Blanco, Amie; Van Ziffle, Jessica; Terdiman, Jonathan P

    2016-04-01

    At least one-third of patients meeting clinical criteria for Lynch syndrome will have no germline mutation and constitutional epimutations leading to promoter methylation of MLH1 have been identified in a subset of these patients. We report the first case of constitutional MLH1 promoter methylation associated with a colonic polyposis syndrome in a 39 year-old man with a family history of colorectal cancer (CRC) and a personal history of 21 polyps identified over 8 years as well as the development of two synchronous CRCs over 16 months who was evaluated for a hereditary cancer syndrome. Immunohistochemistry (IHC) of multiple tumors showed absent MLH1 and PMS2 expression, though germline testing with Sanger sequencing and multiplex ligation-dependent probe amplification of these mismatch repair genes (MMR) genes was negative. A next generation sequencing panel of 29 genes also failed to identify a pathogenic mutation. Hypermethylation was identified in MLH1 intron 1 in tumor specimens along with buccal cells and peripheral white blood cells, confirming the diagnosis of constitutional MLH1 promoter methylation. This case highlights that constitutional MLH1 methylation should be considered in the differential diagnosis for a polyposis syndrome if IHC staining shows absent MMR gene expression.

  1. Detection of receptor ligands by monitoring selective stabilization of a Renilla luciferase-tagged, constitutively active mutant, G-protein-coupled receptor

    PubMed Central

    Ramsay, Douglas; Bevan, Nicola; Rees, Stephen; Milligan, Graeme

    2001-01-01

    The wild-type β2-adrenoceptor and a constitutively active mutant of this receptor were C-terminally tagged with luciferase from the sea pansy Renilla reniformis. C-terminal addition of Renilla luciferase did not substantially alter the levels of expression of either form of the receptor, the elevated constitutive activity of the mutant β2-adrenoceptor nor the capacity of isoprenaline to elevate cyclic AMP levels in intact cells expressing these constructs. Treatment of cells expressing constitutively active mutant β2-adrenoceptor-Renilla luciferase with antagonist/inverse agonist ligands resulted in upregulation of levels of this polypeptide which could be monitored by the elevated luciferase activity. The pEC50 for ligand-induced luciferase upregulation and ligand affinity to bind the receptor were highly correlated. Similar upregulation could be observed following sustained treatment with agonist ligands. These effects were only observed at a constitutively active mutant of the β2-adrenoceptor. Co-expression of the wild-type β2-adrenoceptor C-terminally tagged with the luciferase from Photinus pyralis did not result in ligand-induced upregulation of the levels of activity of this luciferase. Co-expression of the constitutively active mutant β2-adrenoceptor-Renilla luciferase and an equivalent mutant of the α1b-adrenoceptor C-terminally tagged with green fluorescent protein allowed pharmacological selectivity of adrenoceptor antagonists to be demonstrated. This approach offers a sensitive and convenient means, which is amenable to high throughput analysis, to monitor ligand binding to a constitutively active mutant receptor. As no prior knowledge of receptor ligands is required this approach may be suitable to identify ligands at orphan G protein-coupled receptors. PMID:11350868

  2. Constitutively expressing cell lines that secrete a truncated bovine herpes virus-1 glycoprotein (gpI) stimulate T-lymphocyte responsiveness.

    PubMed

    Leary, T P; Gao, Y; Splitter, G A

    1992-07-01

    The desire to obtain authentically glycosylated viral protein products in sufficient quantity for immunological study has led to the use of eucaryotic expression vectors for protein production. An additional advantage is that these protein products can be studied individually in the absence of their native viral environment. We have cloned a complementary DNA (cDNA) encoding bovine herpes virus-1 (BHV-1) glycoprotein 1 (gpI) into the eucaryotic expression vector, pZipNeo SVX1. Since this protein is normally embedded within the membrane of BHV-1 infected cells, we removed sequences encoding the transmembrane domain of the native protein. After transfection of the plasmid construct into the canine osteosarcoma cell line, D17, or Madin-Darby bovine kidney (MDBK) cells, a truncated BHV-1 (gpI) was secreted into the culture medium as demonstrated by radioimmunoprecipitation and SDS-PAGE. Both a CD4+ T-lymphocyte line specific for BHV-1 and freshly isolated T lymphocytes could recognize and respond to the secreted recombinant gpI. Further, recombinant gpI could elicit both antibody and cellular responses in cattle when used as an immunogen. Having established constitutively glycoprotein producing cell lines, future studies in vaccine evaluation of gpI will be facilitated.

  3. Recruitment of CREB1 and Histone Deacetylase 2 (HDAC2) to the Mouse Ltbp-1 Promoter Regulates its Constitutive Expression in a Dioxin Receptor-dependent Manner

    PubMed Central

    Gomez-Duran, Aurea; Ballestar, Esteban; Carvajal-Gonzalez, Jose M.; Marlowe, Jennifer L.; Puga, Alvaro; Esteller, Manel; Fernandez-Salguero, Pedro M.

    2010-01-01

    Latent TGFβ-binding protein 1 (LTBP-1) is a key regulator of TGFβ targeting and activation in the extracellular matrix. LTBP-1 is recognized as a major docking molecule to localize, and possibly to activate, TGFβ in the extracellular matrix. Despite this relevant function, the molecular mechanisms regulating Ltbp-1 transcription remain largely unknown. Previous results from our laboratory revealed that mouse embryonic fibroblasts (MEF) lacking dioxin receptor (AhR) had increased Ltbp-1 mRNA expression and elevated TGFβ activity, suggesting that AhR repressed Ltbp-1 transcription. Here, we have cloned the mouse Ltbp-1 gene promoter and analysed its mechanism of transcriptional repression by AhR. Reporter gene assays, AhR over-expression and site-directed mutagenesis showed that basal Ltbp-1 transcription is AhR-dependent. Chromatin immunoprecipitation (ChIP) and RNA interference (RNAi) revealed that AhR regulates Ltbp-1 transcription by a mechanism involving recruitment of co-activators such as CREB1 and co-repressors such as HDAC2 to the Ltbp-1 promoter. In AhR-expressing (AhR+/+) MEF cells, the recruitment of HDAC1, 2 and 4 correlated with decreased K8H4 acetylation and impaired binding of pCREBSer133 to the Ltbp-1 promoter, likely maintaining a constitutive repressed state. AhR−/− MEF cells had the opposite pattern of HDACs and pCREB1Ser133 binding to Ltbp-1 promoter, and therefore, over-expressed Ltbp-1 mRNA. In agreement, siRNA for HDAC2 increased Ltbp-1 expression and K8H4 acetylation in AhR+/+ but not in AhR−/− MEF cells. We suggest that HDAC2 binding keeps Ltbp-1 promoter repressed in AhR+/+ MEF cells, whereas in AhR-null MEF cells the absence of HDAC2 and the binding of pCREBSer133 allow Ltbp-1 transcription. Thus, epigenetics can contribute to constitutive Ltbp-1 repression by a mechanism requiring AhR activity. PMID:18508077

  4. Heterologous production of kasugamycin, an aminoglycoside antibiotic from Streptomyces kasugaensis, in Streptomyces lividans and Rhodococcus erythropolis L-88 by constitutive expression of the biosynthetic gene cluster.

    PubMed

    Kasuga, Kano; Sasaki, Akira; Matsuo, Takashi; Yamamoto, Chika; Minato, Yuiko; Kuwahara, Naoya; Fujii, Chikako; Kobayashi, Masayuki; Agematu, Hitosi; Tamura, Tomohiro; Komatsu, Mamoru; Ishikawa, Jun; Ikeda, Haruo; Kojima, Ikuo

    2017-05-01

    Kasugamycin (KSM), an aminoglycoside antibiotic isolated from Streptomyces kasugaensis cultures, has been used against rice blast disease for more than 50 years. We cloned the KSM biosynthetic gene (KBG) cluster from S. kasugaensis MB273-C4 and constructed three KBG cassettes (i.e., cassettes I-III) to enable heterologous production of KSM in many actinomycetes by constitutive expression of KBGs. Cassette I comprised all putative transcriptional units in the cluster, but it was placed under the control of the P neo promoter from Tn5. It was not maintained stably in Streptomyces lividans and did not transform Rhodococcus erythropolis. Cassette II retained the original arrangement of KBGs, except that the promoter of kasT, the specific activator gene for KBG, was replaced with P rpsJ , the constitutive promoter of rpsJ from Streptomyces avermitilis. To enhance the intracellular concentration of myo-inositol, an expression cassette of ino1 encoding the inositol-1-phosphate synthase from S. avermitilis was inserted into cassette II to generate cassette III. These two cassettes showed stable maintenance in S. lividans and R. erythropolis to produce KSM. Particularly, the transformants of S. lividans induced KSM production up to the same levels as those produced by S. kasugaensis. Furthermore, cassette III induced more KSM accumulation than cassette II in R. erythropolis, suggesting an exogenous supply of myo-inositol by the ino1 expression in the host. Cassettes II and III appear to be useful for heterologous KSM production in actinomycetes. Rhodococcus exhibiting a spherical form in liquid cultivation is also a promising heterologous host for antibiotic fermentation.

  5. Constitutive Expression of a Transcription Termination Factor by a Repressed Prophage: Promoters for Transcribing the Phage HK022 nun Gene

    PubMed Central

    King, Rodney A.; Madsen, Peter L.; Weisberg, Robert A.

    2000-01-01

    Lysogens of phage HK022 are resistant to infection by phage λ. Lambda resistance is caused by the action of the HK022 Nun protein, which prematurely terminates early λ transcripts. We report here that transcription of the nun gene initiates at a constitutive prophage promoter, PNun, located just upstream of the protein coding sequence. The 5′ end of the transcript was determined by primer extension analysis of RNA isolated from HK022 lysogens or RNA made in vitro by transcribing a template containing the promoter with purified Escherichia coli RNA polymerase. Inactivation of PNun by mutation greatly reduced Nun activity and Nun antigen in an HK022 lysogen. However, a low level of residual activity was detected, suggesting that a secondary promoter also contributes to nun expression. We found one possible secondary promoter, PNun′, just upstream of PNun. Neither promoter is likely to increase the expression of other phage genes in a lysogen because their transcripts should be terminated downstream of nun. We estimate that HK022 lysogens in stationary phase contain several hundred molecules of Nun per cell and that cells in exponential phase probably contain fewer. PMID:10629193

  6. Constitutively Expressed IFITM3 Protein in Human Endothelial Cells Poses an Early Infection Block to Human Influenza Viruses.

    PubMed

    Sun, Xiangjie; Zeng, Hui; Kumar, Amrita; Belser, Jessica A; Maines, Taronna R; Tumpey, Terrence M

    2016-12-15

    A role for pulmonary endothelial cells in the orchestration of cytokine production and leukocyte recruitment during influenza virus infection, leading to severe lung damage, has been recently identified. As the mechanistic pathway for this ability is not fully known, we extended previous studies on influenza virus tropism in cultured human pulmonary endothelial cells. We found that a subset of avian influenza viruses, including potentially pandemic H5N1, H7N9, and H9N2 viruses, could infect human pulmonary endothelial cells (HULEC) with high efficiency compared to human H1N1 or H3N2 viruses. In HULEC, human influenza viruses were capable of binding to host cellular receptors, becoming internalized and initiating hemifusion but failing to uncoat the viral nucleocapsid and to replicate in host nuclei. Unlike numerous cell types, including epithelial cells, we found that pulmonary endothelial cells constitutively express a high level of the restriction protein IFITM3 in endosomal compartments. IFITM3 knockdown by small interfering RNA (siRNA) could partially rescue H1N1 virus infection in HULEC, suggesting IFITM3 proteins were involved in blocking human influenza virus infection in endothelial cells. In contrast, selected avian influenza viruses were able to escape IFITM3 restriction in endothelial cells, possibly by fusing in early endosomes at higher pH or by other, unknown mechanisms. Collectively, our study demonstrates that the human pulmonary endothelium possesses intrinsic immunity to human influenza viruses, in part due to the constitutive expression of IFITM3 proteins. Notably, certain avian influenza viruses have evolved to escape this restriction, possibly contributing to virus-induced pneumonia and severe lung disease in humans. Avian influenza viruses, including H5N1 and H7N9, have been associated with severe respiratory disease and fatal outcomes in humans. Although acute respiratory distress syndrome (ARDS) and progressive pulmonary endothelial damage

  7. Cell-type specific expression of constitutively-active Rheb promotes regeneration of bulbospinal respiratory axons following cervical SCI.

    PubMed

    Urban, Mark W; Ghosh, Biswarup; Strojny, Laura R; Block, Cole G; Blazejewski, Sara M; Wright, Megan C; Smith, George M; Lepore, Angelo C

    2018-05-01

    Damage to respiratory neural circuitry and consequent loss of diaphragm function is a major cause of morbidity and mortality in individuals suffering from traumatic cervical spinal cord injury (SCI). Repair of CNS axons after SCI remains a therapeutic challenge, despite current efforts. SCI disrupts inspiratory signals originating in the rostral ventral respiratory group (rVRG) of the medulla from their phrenic motor neuron (PhMN) targets, resulting in loss of diaphragm function. Using a rat model of cervical hemisection SCI, we aimed to restore rVRG-PhMN-diaphragm circuitry by stimulating regeneration of injured rVRG axons via targeted induction of Rheb (ras homolog enriched in brain), a signaling molecule that regulates neuronal-intrinsic axon growth potential. Following C2 hemisection, we performed intra-rVRG injection of an adeno-associated virus serotype-2 (AAV2) vector that drives expression of a constitutively-active form of Rheb (cRheb). rVRG neuron-specific cRheb expression robustly increased mTOR pathway activity within the transduced rVRG neuron population ipsilateral to the hemisection, as assessed by levels of phosphorylated ribosomal S6 kinase. By co-injecting our novel AAV2-mCherry/WGA anterograde/trans-synaptic axonal tracer into rVRG, we found that cRheb expression promoted regeneration of injured rVRG axons into the lesion site, while we observed no rVRG axon regrowth with AAV2-GFP control. AAV2-cRheb also significantly reduced rVRG axonal dieback within the intact spinal cord rostral to the lesion. However, cRheb expression did not promote any recovery of ipsilateral hemi-diaphragm function, as assessed by inspiratory electromyography (EMG) burst amplitudes. This lack of functional recovery was likely because regrowing rVRG fibers did not extend back into the caudal spinal cord to synaptically reinnervate PhMNs that we retrogradely-labeled with cholera toxin B from the ipsilateral hemi-diaphragm. Our findings demonstrate that enhancing neuronal

  8. Co-regulation of the atrial natriuretic factor and cardiac myosin light chain-2 genes during alpha-adrenergic stimulation of neonatal rat ventricular cells. Identification of cis sequences within an embryonic and a constitutive contractile protein gene which mediate inducible expression.

    PubMed

    Knowlton, K U; Baracchini, E; Ross, R S; Harris, A N; Henderson, S A; Evans, S M; Glembotski, C C; Chien, K R

    1991-04-25

    To study the mechanisms which mediate the transcriptional activation of cardiac genes during alpha adrenergic stimulation, the present study examined the regulated expression of three cardiac genes, a ventricular embryonic gene (atrial natriuretic factor, ANF), a constitutively expressed contractile protein gene (cardiac MLC-2), and a cardiac sodium channel gene. alpha 1-Adrenergic stimulation activates the expression and release of ANF from neonatal ventricular cells. As assessed by RNase protection analyses, treatment with alpha-adrenergic agonists increases the steady-state levels of ANF mRNA by greater than 15-fold. However, a rat cardiac sodium channel gene mRNA is not induced, indicating that alpha-adrenergic stimulation does not lead to an increase in the expression of all cardiac genes. Studies employing a series of rat ANF luciferase and rat MLC-2 luciferase fusion genes identify 315- and 92-base pair cis regulatory sequences within an embryonic gene (ANF) and a constitutively expressed contractile protein gene (MLC-2), respectively, which mediate alpha-adrenergic-inducible gene expression. Transfection of various ANF luciferase reporters into neonatal rat ventricular cells demonstrated that upstream sequences which mediate tissue-specific expression (-3003 to -638) can be segregated from those responsible for inducibility. The lack of inducibility of a cardiac Na+ channel gene, and the segregation of ANF gene sequences which mediate cardiac specific from those which mediate inducible expression, provides further insight into the relationship between muscle-specific and inducible expression during cardiac myocyte hypertrophy. Based on these results, a testable model is proposed for the induction of embryonic cardiac genes and constitutively expressed contractile protein genes and the noninducibility of a subset of cardiac genes during alpha-adrenergic stimulation of neonatal rat ventricular cells.

  9. Community composition, abundance and biomass of zooplankton in Zhangzi Island waters, Northern Yellow Sea

    NASA Astrophysics Data System (ADS)

    Yin, Jiehui; Zhang, Guangtao; Li, Chaolun; Wang, Shiwei; Zhao, Zengxia; Wan, Aiyong

    2017-09-01

    Samples were collected monthly from the sea area around Zhangzi Island, northern Yellow Sea, from July 2009 to June 2010. Vertical net towing was used to examine spatial and temporal variability in zooplankton abundance and biomass. Overall, Calanus sinicus and Saggita crassa were the dominant species found during the study period, while the amphipod Themisto gracilipes was dominant in winter and spring. Vast numbers of the ctenophore species of the genus Beroe were found in October and November. It was not possible to count them, but they constituted a large portion of the total zooplankton biomass. Zooplankton species diversity was highest in October, and species evenness was highest in April. Zooplankton abundance (non-jellyfish) and biomass were highest in June and lowest in August, with annual averages of 131.3 ind./m³ and 217.5 mg/m³, respectively. Water temperature may be responsible for the variations in zooplankton abundance and biomass. Beroe biomass was negatively correlated with other zooplankton abundance. Longterm investigations will be carried out to learn more about the influence of the environment on zooplankton assemblages.

  10. Constitutively active transforming growth factor β receptor 1 in the mouse ovary promotes tumorigenesis

    PubMed Central

    Gao, Yang; Vincent, David F.; Davis, Anna Jane; Sansom, Owen J.; Bartholin, Laurent; Li, Qinglei

    2016-01-01

    Despite the well-established tumor suppressive role of TGFβ proteins, depletion of key TGFβ signaling components in the mouse ovary does not induce a growth advantage. To define the role of TGFβ signaling in ovarian tumorigenesis, we created a mouse model expressing a constitutively active TGFβ receptor 1 (TGFBR1) in ovarian somatic cells using conditional gain-of-function approach. Remarkably, these mice developed ovarian sex cord-stromal tumors with complete penetrance, leading to reproductive failure and mortality. The tumors expressed multiple granulosa cell markers and caused elevated serum inhibin and estradiol levels, reminiscent of granulosa cell tumors. Consistent with the tumorigenic effect, overactivation of TGFBR1 altered tumor microenvironment by promoting angiogenesis and enhanced ovarian cell proliferation, accompanied by impaired cell differentiation and dysregulated expression of critical genes in ovarian function. By further exploiting complementary genetic models, we substantiated our finding that constitutively active TGFBR1 is a potent oncogenic switch in mouse granulosa cells. In summary, overactivation of TGFBR1 drives gonadal tumor development. The TGFBR1 constitutively active mouse model phenocopies a number of morphological, hormonal, and molecular features of human granulosa cell tumors and are potentially valuable for preclinical testing of targeted therapies to treat granulosa cell tumors, a class of poorly defined ovarian malignancies. PMID:27344183

  11. Constitutional Issues--Watergate and the Constitution. Teaching with Documents.

    ERIC Educational Resources Information Center

    National Archives and Records Administration, Washington, DC.

    When U.S. President Richard Nixon resigned in 1974 in the wake of the Watergate scandal, it was only the second time that impeachment of a president had been considered. Although the U.S. Constitution has provisions for a person removed from office to be indicted, there are no guidelines in the Constitution about a President who has resigned. The…

  12. Long-term aldosterone administration increases renal Na+-Cl- cotransporter abundance in late distal convoluted tubule.

    PubMed

    Poulsen, Søren Brandt; Christensen, Birgitte Mønster

    2017-09-01

    Renal Na + -Cl - cotransporter (NCC) is expressed in early distal convoluted tubule (DCT) 1 and late DCT (DCT2). NCC activity can be stimulated by aldosterone administration, and the mechanism is assumed to depend on the enzyme 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2), which inactivates glucocorticoids that would otherwise occupy aldosterone receptors. Because 11β-HSD2 in rat may only be abundantly expressed in DCT2 cells and not in DCT1 cells, it has been speculated that aldosterone specifically stimulates NCC activity in DCT2 cells. In mice, however, it is debated if 11β-HSD2 is expressed in DCT2 cells. The present study examined whether aldosterone administration in mice stimulates NCC abundance and phosphorylation in DCT2 cells but not in DCT1 cells. B6/C57 male mice were administered 100 µg aldosterone·kg body weight -1 ·24 h -1 for 6 days and euthanized during isoflurane inhalation. Western blotting of whole kidney homogenate showed that aldosterone administration stimulated NCC and pT58-NCC abundances ( P < 0.001). In DCT1 cells, confocal microscopy detected no effect of the aldosterone administration on NCC and pT58-NCC abundances. By contrast, NCC and pT58-NCC abundances were stimulated by aldosterone administration in the middle of DCT2 ( P < 0.001 and <0.01, respectively) and at the junction between DCT2 and CNT ( P < 0.001 and <0.05, respectively). In contrast to rat, immunohistochemistry in mouse showed no/very weak 11β-HSD2 expression in DCT2 cells. Collectively, long-term aldosterone administration stimulates mouse NCC and pT58-NCC abundances in DCT2 cells and presumably not in DCT1 cells. Copyright © 2017 the American Physiological Society.

  13. Constitutively active RAS signaling reduces 1,25 dihydroxyvitamin D-mediated gene transcription in intestinal epithelial cells by reducing vitamin D receptor expression.

    PubMed

    DeSmet, Marsha L; Fleet, James C

    2017-10-01

    High vitamin D status is associated with reduced colon cancer risk but these studies ignore the diversity in the molecular etiology of colon cancer. RAS activating mutations are common in colon cancer and they activate pro-proliferative signaling pathways. We examined the impact of RAS activating mutations on 1,25 dihydroxyvitamin D (1,25(OH) 2 D)-mediated gene expression in cultured colon and intestinal cell lines. Transient transfection of Caco-2 cells with a constitutively active mutant K-RAS (G12 V) significantly reduced 1,25(OH) 2 D-induced activity of both a human 25-hydroxyvitamin D, 24 hydroxyase (CYP24A1) promoter-luciferase and an artificial 3X vitamin D response element (VDRE) promoter-luciferase reporter gene. Young Adult Mouse Colon (YAMC) and Rat Intestinal Epithelial (RIE) cell lines with stable expression of mutant H-RAS had suppressed 1,25(OH) 2 D-mediated induction of CYP24A1 mRNA. The RAS effects were associated with lower Vitamin D receptor (VDR) mRNA and protein levels in YAMC and RIE cells and they could be partially reversed by VDR overexpression. RAS-mediated suppression of VDR levels was not due to either reduced VDR mRNA stability or increased VDR gene methylation. However, chromatin accessibility to the VDR gene at the proximal promoter (-300bp), an enhancer region at -6kb, and an enhancer region located in exon 3 was significantly reduced in RAS transformed YAMC cells (YAMC-RAS). These data show that constitutively active RAS signaling suppresses 1,25(OH) 2 D-mediated gene transcription in colon epithelial cells by reducing VDR gene transcription but the mechanism for this suppression is not yet known. These data suggest that cancers with RAS-activating mutations may be less responsive to vitamin D mediated treatment or chemoprevention. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Essential Medicines in National Constitutions

    PubMed Central

    Toebes, Brigit; Hogerzeil, Hans

    2016-01-01

    Abstract A constitutional guarantee of access to essential medicines has been identified as an important indicator of government commitment to the progressive realization of the right to the highest attainable standard of health. The objective of this study was to evaluate provisions on access to essential medicines in national constitutions, to identify comprehensive examples of constitutional text on medicines that can be used as a model for other countries, and to evaluate the evolution of constitutional medicines-related rights since 2008. Relevant articles were selected from an inventory of constitutional texts from WHO member states. References to states’ legal obligations under international human rights law were evaluated. Twenty-two constitutions worldwide now oblige governments to protect and/or to fulfill accessibility of, availability of, and/or quality of medicines. Since 2008, state responsibilities to fulfill access to essential medicines have expanded in five constitutions, been maintained in four constitutions, and have regressed in one constitution. Government commitments to essential medicines are an important foundation of health system equity and are included increasingly in state constitutions. PMID:27781006

  15. Temporal changes in the expression of avian β-defensins in the chicken vagina during sexual maturation and Salmonella infection.

    PubMed

    Anastasiadou, Maria; Avdi, Melpomeni; Theodoridis, Alexandros; Michailidis, Georgios

    2013-06-01

    Avian β-defensins (AvβDs) constitute a family of antimicrobial peptides that are critical to innate immunity in chickens, providing protection against microbial pathogens including Salmonella Enteritidis (SE). As apart from the digestive tract another main route of SE colonization in birds is via infection of the oviduct and specifically of the vagina, the aim of this study was to investigate the expression of the complete family of AvβDs, in the chicken vagina in vivo, to determine whether sexual maturation affects their mRNA abundance and to investigate whether SE infection alters the vaginal AvβDs expression. Expression analysis revealed that 11 members of the AvβD family were expressed in the chicken vagina. Quantitative real-time PCR analysis revealed that the mRNA abundance of five AvβDs was up regulated and of one AvβD was down regulated with respect to sexual maturation. In addition SE infection resulted in a significant induction of AvβD5, 7, 10, 11, 12 and 14 in the vagina of sexually mature birds, and in a significant induction of AvβD5 and 11 in the vagina of aged birds. These findings provide strong evidence to suggest that an AvβD-mediated immune response mechanism exists in the chicken vagina providing protection against bacterial pathogens including Salmonella species.

  16. Making the Constitution Meaningful.

    ERIC Educational Resources Information Center

    Pelow, Randall A.

    1989-01-01

    Describes learning activities based on the U.S. Constitution that enhance higher level thinking skills in elementary students. One activity proposes a hypothetical constitutional amendment banning Saturday cartoons; a second taxes children's earnings; and other activities focus on dramatizing events surrounding the Constitutional Convention. (LS)

  17. Oceanic protists with different forms of acquired phototrophy display contrasting biogeographies and abundance.

    PubMed

    Leles, S G; Mitra, A; Flynn, K J; Stoecker, D K; Hansen, P J; Calbet, A; McManus, G B; Sanders, R W; Caron, D A; Not, F; Hallegraeff, G M; Pitta, P; Raven, J A; Johnson, M D; Glibert, P M; Våge, S

    2017-08-16

    This first comprehensive analysis of the global biogeography of marine protistan plankton with acquired phototrophy shows these mixotrophic organisms to be ubiquitous and abundant; however, their biogeography differs markedly between different functional groups. These mixotrophs, lacking a constitutive capacity for photosynthesis (i.e. non-constitutive mixotrophs, NCMs), acquire their phototrophic potential through either integration of prey-plastids or through endosymbiotic associations with photosynthetic microbes. Analysis of field data reveals that 40-60% of plankton traditionally labelled as (non-phototrophic) microzooplankton are actually NCMs, employing acquired phototrophy in addition to phagotrophy. Specialist NCMs acquire chloroplasts or endosymbionts from specific prey, while generalist NCMs obtain chloroplasts from a variety of prey. These contrasting functional types of NCMs exhibit distinct seasonal and spatial global distribution patterns. Mixotrophs reliant on 'stolen' chloroplasts, controlled by prey diversity and abundance, dominate in high-biomass areas. Mixotrophs harbouring intact symbionts are present in all waters and dominate particularly in oligotrophic open ocean systems. The contrasting temporal and spatial patterns of distribution of different mixotroph functional types across the oceanic provinces, as revealed in this study, challenges traditional interpretations of marine food web structures. Mixotrophs with acquired phototrophy (NCMs) warrant greater recognition in marine research. © 2017 The Author(s).

  18. Distribution and abundance of hematophagous flies (Glossinidae, Stomoxys, and Tabanidae) in two national parks of Gabon

    PubMed Central

    Bitome Essono, Paul Yannick; Dechaume-Moncharmont, François-Xavier; Mavoungou, Jacques; Obiang Mba, Régis; Duvallet, Gérard; Bretagnolle, François

    2015-01-01

    In order to minimize risks of pathogen transmission with the development of ecotourism in Gabon, a seasonal inventory has been performed in five contrasted biotopes in Ivindo (INP) and Moukalaba-Doudou (MDNP) National Parks. A total of 10,033 hematophagous flies were captured. The Glossinidae, with six different species identified, was the most abundant group and constitutes about 60% of the captured flies compared to the Stomoxys (6 species also identified) and Tabanidae with 28% and 12%, respectively. The Glossinidae showed a higher rate of capture in primary forest and in research camps. In INP, the Stomoxys showed a higher rate of capture in secondary forest and at village borders, whereas in MDNP the Stomoxys were captured more in the savannah area. Thus, each fly group seemed to reach maximum abundance in different habitats. The Glossinidae were more abundant in primary forest and near research camps while Stomoxys were more abundant in secondary forest and savannah. The Tabanidae did not show a clear habitat preference. PMID:26187781

  19. The Arabidopsis mutant cev1 has constitutively active jasmonate and ethylene signal pathways and enhanced resistance to pathogens.

    PubMed

    Ellis, C; Turner, J G

    2001-05-01

    Jasmonates (JAs) inhibit plant growth and induce plant defense responses. To define genes in the Arabidopsis JA signal pathway, we screened for mutants with constitutive expression of a luciferase reporter for the JA-responsive promoter from the vegetative storage protein gene VSP1. One mutant, named constitutive expression of VSP1 (cev1), produced plants that were smaller than wild type, had stunted roots with long root hairs, accumulated anthocyanin, had constitutive expression of the defense-related genes VSP1, VSP2, Thi2.1, PDF1.2, and CHI-B, and had enhanced resistance to powdery mildew diseases. Genetic evidence indicated that the cev1 phenotype required both COI1, an essential component of the JA signal pathway, and ETR1, which encodes the ethylene receptor. We conclude that cev1 stimulates both the JA and the ethylene signal pathways and that CEV1 regulates an early step in an Arabidopsis defense pathway.

  20. The Arabidopsis Mutant cev1 Has Constitutively Active Jasmonate and Ethylene Signal Pathways and Enhanced Resistance to Pathogens

    PubMed Central

    Ellis, Christine; Turner, John G.

    2001-01-01

    Jasmonates (JAs) inhibit plant growth and induce plant defense responses. To define genes in the Arabidopsis JA signal pathway, we screened for mutants with constitutive expression of a luciferase reporter for the JA-responsive promoter from the vegetative storage protein gene VSP1. One mutant, named constitutive expression of VSP1 (cev1), produced plants that were smaller than wild type, had stunted roots with long root hairs, accumulated anthocyanin, had constitutive expression of the defense-related genes VSP1, VSP2, Thi2.1, PDF1.2, and CHI-B, and had enhanced resistance to powdery mildew diseases. Genetic evidence indicated that the cev1 phenotype required both COI1, an essential component of the JA signal pathway, and ETR1, which encodes the ethylene receptor. We conclude that cev1 stimulates both the JA and the ethylene signal pathways and that CEV1 regulates an early step in an Arabidopsis defense pathway. PMID:11340179

  1. Argonautes promote male fertility and provide a paternal memory of germline gene expression in C. elegans

    PubMed Central

    Conine, Colin C.; Moresco, James J.; Gu, Weifeng; Shirayama, Masaki; Conte, Darryl; Yates, John R.; Mello, Craig C.

    2014-01-01

    SUMMARY During each life cycle germ cells preserve and pass on both genetic and epigenetic information. In C. elegans, the ALG-3/4 Argonaute proteins are expressed during male gametogenesis and promote male fertility. Here we show that the CSR-1 Argonaute functions with ALG-3/4 to positively regulate target genes required for spermiogenesis. Our findings suggest that ALG-3/4 functions during spermatogenesis to amplify a small-RNA signal that represents an epigenetic memory of male-specific gene expression. CSR-1, which is abundant in mature sperm, appears to transmit this memory to offspring. Surprisingly, in addition to small RNAs targeting male-specific genes, we show that males also harbor an extensive repertoire of CSR-1 small RNAs targeting oogenesis-specific mRNAs. Together these findings suggest that C. elegans sperm transmit not only the genome but also epigenetic binary signals in the form of Argonaute/small-RNA complexes that constitute a memory of gene expression in preceding generations. PMID:24360276

  2. [The Constitutional Court and new scenarios of biomedicine. (Constitutional reflections on the decision of the Constitutional Tribunal 116/1999, June 17)].

    PubMed

    Arruego Rodríguez, G; Chueca Rodríguez, R

    2000-01-01

    The recent ruling by the Constitutional Court (116/1999, 17 June) ended the process which had been initiated by the challenge filed by 63 conservative MPs against Law 35/1988, 22 November, on the Law on Human Assisted Reproduction Techniques. In our opinion, this ruling helps define the scope of the constitutional provisions used by the appellants in their challenge, provisions which had already been interpreted in lower rulings. The considerations given in this article are designed to establish the terms and framework which lawmakers and law experts should bear in mind when they prepare future, as will necessarily be the case. In view of some of the arguments used in the ruling, we believe it is appropriate to draw attention to some of the most salient constitutional aspects, such as the scope of the Constitutional Court's role as the ultimate judge of constitutionality, and the exact nature of the constitutional notion of fundamental right which, although complicated at times, is nonetheless a precise and accurate legal concept.

  3. Constitutive Modelling and Deformation Band Angle Predictions for High Porosity Sandstones

    NASA Astrophysics Data System (ADS)

    Richards, M. C.; Issen, K. A.; Ingraham, M. D.

    2017-12-01

    The development of a field-scale deformation model requires a constitutive framework that is capable of representing known material behavior and able to be calibrated using available mechanical response data. This work employs the principle of hyperplasticity (e.g., Houlsby and Puzrin, 2006) to develop such a constitutive framework for high porosity sandstone. Adapting the works of Zimmerman et al. (1986) and Collins and Houlsby (1997), the mechanical data set of Ingraham et al. (2013 a, b) was used to develop a specific constitutive framework for Castlegate sandstone, a high porosity fluvial-deposited reservoir analog rock. Using the mechanical data set of Ingraham et al. (2013 a, b), explicit expressions and material parameters of the elastic moduli and strain tensors were obtained. With these expressions, analytical and numerical techniques were then employed to partition the total mechanical strain into elastic, coupled, and plastic strain components. With the partitioned strain data, yield surfaces in true-stress space, coefficients of internal friction, dilatancy factors, along with the theorectical predictions of the deformation band angles were obtained. These results were also evaluated against band angle values obtained from a) measurements on specimen jackets (Ingraham et al., 2013a), b) plane fits through located acoustic emissions (AE) events (Ingraham et al. 2013b), and c) X-ray micro-computed tomography (micro-CT) calculations.

  4. Identification and expression profiles of the WRKY transcription factor family in Ricinus communis.

    PubMed

    Li, Hui-Liang; Zhang, Liang-Bo; Guo, Dong; Li, Chang-Zhu; Peng, Shi-Qing

    2012-07-25

    In plants, WRKY proteins constitute a large family of transcription factors. They are involved in many biological processes, such as plant development, metabolism, and responses to biotic and abiotic stresses. A large number of WRKY transcription factors have been reported from Arabidopsis, rice, and other higher plants. The recent publication of the draft genome sequence of castor bean (Ricinus communis) has allowed a genome-wide search for R. communis WRKY (RcWRKY) transcription factors and the comparison of these positively identified proteins with their homologs in model plants. A total of 47 WRKY genes were identified in the castor bean genome. According to the structural features of the WRKY domain, the RcWRKY are classified into seven main phylogenetic groups. Furthermore, putative orthologs of RcWRKY proteins in Arabidopsis and rice could now be assigned. An analysis of expression profiles of RcWRKY genes indicates that 47 WRKY genes display differential expressions either in their transcript abundance or expression patterns under normal growth conditions. Copyright © 2012 Elsevier B.V. All rights reserved.

  5. MLH1 constitutional and somatic methylation in patients with MLH1 negative tumors fulfilling the revised Bethesda criteria.

    PubMed

    Crucianelli, Francesca; Tricarico, Rossella; Turchetti, Daniela; Gorelli, Greta; Gensini, Francesca; Sestini, Roberta; Giunti, Laura; Pedroni, Monica; Ponz de Leon, Maurizio; Civitelli, Serenella; Genuardi, Maurizio

    2014-10-01

    Lynch syndrome (LS) is a tumor predisposing condition caused by constitutional defects in genes coding for components of the mismatch repair (MMR) apparatus. While hypermethylation of the promoter of the MMR gene MLH1 occurs in about 15% of colorectal cancer samples, it has also been observed as a constitutional alteration, in the absence of DNA sequence mutations, in a small number of LS patients. In order to obtain further insights on the phenotypic characteristics of MLH1 epimutation carriers, we investigated the somatic and constitutional MLH1 methylation status of 14 unrelated subjects with a suspicion of LS who were negative for MMR gene constitutional mutations and whose tumors did not express the MLH1 protein. A novel case of constitutional MLH1 epimutation was identified. This patient was affected with multiple primary tumors, including breast cancer, diagnosed starting from the age of 55 y. Investigation of her offspring by allele specific expression revealed that the epimutation was not stable across generations. We also found MLH1 hypermethylation in cancer samples from 4 additional patients who did not have evidence of constitutional defects. These patients had some characteristics of LS, namely early age at onset and/or positive family history, raising the possibility of genetic influences in the establishment of somatic MLH1 methylation.

  6. Effects of nutrient availabiiity on biomass allocation as well as constitutive and rapid induced herbivore resistance in poplar

    Treesearch

    Carolyn Glynn; Daniel A. Herms; Marie Egawa; Robert Hansen; William J. Mattson

    2003-01-01

    Many studies have examined effects of nutrient availability on constitutive herbivore resistance of plants, but few have addressed effects on expression of rapid induced resistance (RIR). We quantified effects of two levels of nutrient availability on growth, biomass allocation, photosynthesis, and constitutive secondary metabolism of black poplar (>i>Populus...

  7. Selection for low or high primary dormancy in Lolium rigidum Gaud seeds results in constitutive differences in stress protein expression and peroxidase activity

    PubMed Central

    Goggin, Danica E.; Powles, Stephen B.; Steadman, Kathryn J.

    2011-01-01

    Seed dormancy in wild Lolium rigidum Gaud (annual ryegrass) populations is highly variable and not well characterized at the biochemical level. To identify some of the determinants of dormancy level in these seeds, the proteomes of subpopulations selected for low and high levels of primary dormancy were compared by two-dimensional polyacrylamide gel electrophoresis of extracts from mature, dry seeds. High-dormancy seeds showed higher expression of small heat shock proteins, enolase, and glyoxalase I than the low-dormancy seeds. The functional relevance of these differences in protein expression was confirmed by the fact that high-dormancy seeds were more tolerant to high temperatures imposed at imbibition and had consistently higher glyoxalase I activity over 0–42 d dark stratification. Higher expression of a putative glutathione peroxidase in low-dormancy seeds was not accompanied by higher activity, but these seeds had a slightly more oxidized glutathione pool and higher total peroxidase activity. Overall, these biochemical and physiological differences suggest that L. rigidum seeds selected for low dormancy are more prepared for rapid germination via peroxidase-mediated cell wall weakening, whilst seeds selected for high dormancy are constitutively prepared to survive environmental stresses, even in the absence of stress during seed development. PMID:20974739

  8. Yeast Fex1p Is a Constitutively Expressed Fluoride Channel with Functional Asymmetry of Its Two Homologous Domains*

    PubMed Central

    Smith, Kathryn D.; Gordon, Patricia B.; Rivetta, Alberto; Allen, Kenneth E.; Berbasova, Tetyana; Slayman, Clifford; Strobel, Scott A.

    2015-01-01

    Fluoride is a ubiquitous environmental toxin with which all biological species must cope. A recently discovered family of fluoride export (FEX) proteins protects organisms from fluoride toxicity by removing it from the cell. We show here that FEX proteins in Saccharomyces cerevisiae function as ion channels that are selective for fluoride over chloride and that these proteins are constitutively expressed at the yeast plasma membrane. Continuous expression is in contrast to many other toxin exporters in yeast, and this, along with the fact that two nearly duplicate proteins are encoded in the yeast genome, suggests that the threat posed by fluoride ions is frequent and detrimental. Structurally, eukaryotic FEX proteins consist of two homologous four-transmembrane helix domains folded into an antiparallel dimer, where the orientation of the two domains is fixed by a single transmembrane linker helix. Using phylogenetic sequence conservation as a guide, we have identified several functionally important residues. There is substantial functional asymmetry in the effect of mutation at corresponding sites in the two domains. Specifically, mutations to residues in the C-terminal domain proved significantly more detrimental to function than did similar mutations in the N-terminal domain. Our data suggest particular residues that may be important to anion specificity, most notably the necessity of a positive charge near the end of TMH1 in the C-terminal domain. It is possible that a cationic charge at this location may create an electrostatic well for fluoride ions entering the channel from the cytoplasm. PMID:26055717

  9. Transcriptome profiling of a Saccharomyces cerevisiae mutant with a constitutively activated Ras/cAMP pathway.

    PubMed

    Jones, D L; Petty, J; Hoyle, D C; Hayes, A; Ragni, E; Popolo, L; Oliver, S G; Stateva, L I

    2003-12-16

    Often changes in gene expression levels have been considered significant only when above/below some arbitrarily chosen threshold. We investigated the effect of applying a purely statistical approach to microarray analysis and demonstrated that small changes in gene expression have biological significance. Whole genome microarray analysis of a pde2Delta mutant, constructed in the Saccharomyces cerevisiae reference strain FY23, revealed altered expression of approximately 11% of protein encoding genes. The mutant, characterized by constitutive activation of the Ras/cAMP pathway, has increased sensitivity to stress, reduced ability to assimilate nonfermentable carbon sources, and some cell wall integrity defects. Applying the Munich Information Centre for Protein Sequences (MIPS) functional categories revealed increased expression of genes related to ribosome biogenesis and downregulation of genes in the cell rescue, defense, cell death and aging category, suggesting a decreased response to stress conditions. A reduced level of gene expression in the unfolded protein response pathway (UPR) was observed. Cell wall genes whose expression was affected by this mutation were also identified. Several of the cAMP-responsive orphan genes, upon further investigation, revealed cell wall functions; others had previously unidentified phenotypes assigned to them. This investigation provides a statistical global transcriptome analysis of the cellular response to constitutive activation of the Ras/cAMP pathway.

  10. Analysing teachers' operations when teaching students: what constitutes scientific theories?

    NASA Astrophysics Data System (ADS)

    Holmqvist, Mona O.; Olander, Clas

    2017-05-01

    The aim of the study is to analyse teachers' efforts to develop secondary school students' knowledge and argumentation skills of what constitutes scientific theories. The analysis is based on Leontiev's three-level structure of activity (activity, action, and operation), as these levels correspond to the questions why, what, and how content is taught. The unit of analysis was a school development project in science education, where design-based interventions were conducted. Data comprised notes and minutes from eight meetings, plans, and video recordings of the lessons, and a written teacher evaluation. The teachers' (n = 7) learning actions were analysed to identify (a) concept formation in science education, (b) expressions of agency, (c) discursive manifestations of contradictions, and (d) patterns of interaction during the science interventions. Three lessons on what constitutes scientific theories were implemented in three different student groups (n = 24, 23, 24), framed by planning and evaluation meetings for each lesson. The results describe (1) the ways in which teachers became more skilled at ensuring instruction met their students' needs and (2) the ways in which teachers' operations during instruction changed as a result of their developed knowledge of how to express the content based on theoretical assumptions.

  11. Expression of conventional and novel glucose transporters, GLUT1, -9, -10, and -12, in vascular smooth muscle cells

    PubMed Central

    Pyla, Rajkumar; Poulose, Ninu; Jun, John Y.

    2013-01-01

    Intimal hyperplasia is characterized by exaggerated proliferation of vascular smooth muscle cells (VSMCs). Enhanced VSMC growth is dependent on increased glucose uptake and metabolism. Facilitative glucose transporters (GLUTs) are comprised of conventional GLUT isoforms (GLUT1–5) and novel GLUT isoforms (GLUT6–14). Previous studies demonstrate that GLUT1 overexpression or GLUT10 downregulation contribute to phenotypic changes in VSMCs. To date, the expression profile of all 14 GLUT isoforms has not been fully examined in VSMCs. Using the proliferative and differentiated phenotypes of human aortic VSMCs, the present study has determined the relative abundance of GLUT1–14 mRNAs by quantitative real-time PCR analysis. Twelve GLUT mRNAs excluding GLUT7 and GLUT14 were detectable in VSMCs. In the proliferative phenotype, the relative abundance of key GLUT mRNAs was GLUT1 (∼43%) > GLUT10 (∼26%) > GLUT9 (∼13%) > GLUT12 (∼4%), whereas in the differentiated phenotype the relative abundance was GLUT10 (∼28%) > GLUT1 (∼25%) > GLUT12 (∼20%) > GLUT9 (∼14%), together constituting 86–87% of total GLUT transcripts. To confirm the expression of key GLUT proteins, immunoblot and immunocytochemical analyses were performed using GLUT isoform-specific primary antibodies. The protein bands characteristic of GLUT1, -9, -10, and -12 were detected in VSMCs in parallel with respective positive controls. In particular, GLUT1 protein expression showed different molecular forms representative of altered glycosylation. While GLUT1 protein displayed a predominant distribution in the plasma membrane, GLUT9, -10, and -12 proteins were mostly distributed in the intracellular compartments. The present study provides the first direct evidence for GLUT9 and GLUT12 expression in VSMCs in conjunction with the previously identified GLUT1 and GLUT10. PMID:23302780

  12. This Constitution: A Bicentennial Chronicle.

    ERIC Educational Resources Information Center

    This Constitution, 1984

    1984-01-01

    A series of articles focus on enduring Constitutional issues and methods for teaching about these in the classroom. Article 1 explores the nature of the judiciary as it has developed under the Constitution. Article 2, by discussing the uses of the Constitution in plays of the Federal Theater Project of the 1930's, examines the Constitution as a…

  13. Constitutive NF-κB activation and tumor-growth promotion by Romo1-mediated reactive oxygen species production

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chung, Jin Sil; Lee, Sora; Yoo, Young Do, E-mail: ydy1130@korea.ac.kr

    2014-08-08

    Highlights: • Romo1 expression is required for constitutive nuclear DNA-binding activity of NF-κB. • Romo1 depletion suppresses tumor growth in vivo. • Romo1 presents a potential therapeutic target for diseases. - Abstract: Deregulation of nuclear factor-κB (NF-κB) and related pathways contribute to tumor cell proliferation and invasion. Mechanisms for constitutive NF-κB activation are not fully explained; however, the underlying defects appear to generate and maintain pro-oxidative conditions. In hepatocellular carcinoma (HCC) tissues, up-regulation of reactive oxygen species modulator 1 (Romo1) correlates positively with tumor size. In the present study, we showed that Romo1 expression is required to maintain constitutive nuclearmore » DNA-binding activity of NF-κB and transcriptional activity through constitutive IκBα phosphorylation. Overexpression of Romo1 promoted p65 nuclear translocation and DNA-binding activity. We also show that Romo1 depletion suppressed anchorage-independent colony formation by HCC cells and suppressed tumor growth in vivo. Based on these findings, Romo1 may be a principal regulatory factor in the maintenance of constitutive NF-κB activation in tumor cells. In the interest of anti-proliferative treatments for cancer, Romo1 may also present a productive target for drug development.« less

  14. Nuclear receptor and target gene mRNA abundance in duodenum and colon of dogs with chronic enteropathies.

    PubMed

    Greger, D L; Gropp, F; Morel, C; Sauter, S; Blum, J W

    2006-11-01

    Nuclear receptors (NR), such as constitutive androstane receptor (CAR), pregnane X receptor (PXR) and peroxisome proliferator-associated receptors alpha and gamma (PPARalpha, PPARgamma) are mediators of inflammation and may be involved in inflammatory bowel disease (IBD) and food responsive diarrhea (FRD) of dogs. The present study compared mRNA abundance of NR and NR target genes [multi drug-resistance gene-1 (MDR1), multiple drug-resistance-associated proteins (MRD2, MRD3), cytochrome P450 (CYP3A12), phenol-sulfating phenol sulfotransferase (SULT1A1) and glutathione-S-transferase (GST A3-3)] in biopsies obtained from duodenum and colon of dogs with IBD and FRD and healthy control dogs (CON; n=7 per group). Upon first presentation of dogs, mRNA levels of PPARalpha, PPARgamma, CAR, PXR and RXRalpha in duodenum as well as PPARgamma, CAR, PXR and RXRalpha in colon were not different among groups (P>0.10). Although mRNA abundance of PPARalpha in colon of dogs with FRD was similar in both IBD and CON (P>0.10), PPARalpha mRNA abundance was higher in IBD than CON (P<0.05). Levels of mRNA of MDR1 in duodenum were higher in FRD than IBD (P<0.05) or CON (P<0.001). Compared with CON, abundances of mRNA for MRP2, CYP3A12 and SULT1A1 were higher in both FRD and IBD than CON (P<0.05). Differences in mRNA levels of PPARalpha and MRP2 in colon and MDR1, MRP2, CYP3A12 and SULT1A1 in duodenum may be indicative for enteropathy in FRD and (or) IBD dogs relative to healthy dogs. More importantly, increased expression of MDR1 in FRD relative to IBD in duodenum may be a useful diagnostic marker to distinguish dogs with FRD from dogs with IBD.

  15. Constitutive cyclooxygenase-2 is involved in central nociceptive processes in humans

    PubMed Central

    Martin, Frédéric; Fletcher, Dominique; Chauvin, Marcel; Bouhassira, Didier

    2007-01-01

    Background Prostaglandins play a major role in inflammation and pain. They are synthesised by the two cyclooxygenase (COX) isoforms: COX-1, which is expressed constitutively in many cell types and COX-2, which is induced at the site of inflammation. However, unlike peripheral tissues, COX-2 is expressed constitutively in the central nervous system and may play a role in nociceptive processes. The present study aimed to investigate the role of constitutive COX-2 in the spinal transmission of nociceptive signals in humans. Methods We used 12 healthy volunteers to compare the effects of the specific COX-2 inhibitor sodium parecoxib (1 mg/kg) or placebo, administered intravenously in a double-blind and cross-over fashion, on the electrophysiological recordings of the nociceptive flexion (RIII) reflex. The RIII reflex is an objective psychophysiological index of the spinal transmission of nociceptive signals and was recorded from the biceps femoris after electrical stimulation of the sural nerve. Two experiments, seven days apart, were carried out on each volunteer. On each experimental day, the effects of parecoxib or placebo were tested on: 1) the RIII reflex threshold, 2) the stimulus-response curves of the reflex up to the tolerance threshold (frequency of stimulation: 0.1 Hz); 3) the progressive increase of the reflex and pain sensations (i.e. “wind-up” phenomenon) induced by a series of 15 stimulations at a frequency of 1 Hz (intensity 20% above RIII threshold). Results Parecoxib, but not placebo, significantly reduced the slope of the stimulus-response curve, suggesting a reduction in the gain of the spinal transmision of nociceptive signals. By contrast, the “wind-up” phenomenon was not significantly altered after administration of parecoxib or placebo. Conclusions Our study shows that constitutive COX-2 modulates spinal nociceptive processes and that the anti-inflammatory and antinociceptive actions of COX-2 inhibitors are not necessarily related. PMID

  16. Environmental drivers of Ixodes ricinus abundance in forest fragments of rural European landscapes.

    PubMed

    Ehrmann, Steffen; Liira, Jaan; Gärtner, Stefanie; Hansen, Karin; Brunet, Jörg; Cousins, Sara A O; Deconchat, Marc; Decocq, Guillaume; De Frenne, Pieter; De Smedt, Pallieter; Diekmann, Martin; Gallet-Moron, Emilie; Kolb, Annette; Lenoir, Jonathan; Lindgren, Jessica; Naaf, Tobias; Paal, Taavi; Valdés, Alicia; Verheyen, Kris; Wulf, Monika; Scherer-Lorenzen, Michael

    2017-09-06

    The castor bean tick (Ixodes ricinus) transmits infectious diseases such as Lyme borreliosis, which constitutes an important ecosystem disservice. Despite many local studies, a comprehensive understanding of the key drivers of tick abundance at the continental scale is still lacking. We analyze a large set of environmental factors as potential drivers of I. ricinus abundance. Our multi-scale study was carried out in deciduous forest fragments dispersed within two contrasting rural landscapes of eight regions, along a macroclimatic gradient stretching from southern France to central Sweden and Estonia. We surveyed the abundance of I. ricinus, plant community composition, forest structure and soil properties and compiled data on landscape structure, macroclimate and habitat properties. We used linear mixed models to analyze patterns and derived the relative importance of the significant drivers. Many drivers had, on their own, either a moderate or small explanatory value for the abundance of I. ricinus, but combined they explained a substantial part of variation. This emphasizes the complex ecology of I. ricinus and the relevance of environmental factors for tick abundance. Macroclimate only explained a small fraction of variation, while properties of macro- and microhabitat, which buffer macroclimate, had a considerable impact on tick abundance. The amount of forest and the composition of the surrounding rural landscape were additionally important drivers of tick abundance. Functional (dispersules) and structural (density of tree and shrub layers) properties of the habitat patch played an important role. Various diversity metrics had only a small relative importance. Ontogenetic tick stages showed pronounced differences in their response. The abundance of nymphs and adults is explained by the preceding stage with a positive relationship, indicating a cumulative effect of drivers. Our findings suggest that the ecosystem disservices of tick-borne diseases, via the

  17. Abundance in proteins expressed after functional electrical stimulation cycling or arm cycling ergometry training in persons with chronic spinal cord injury.

    PubMed

    Gorgey, Ashraf S; Graham, Zachary A; Bauman, William A; Cardozo, Christopher; Gater, David R

    2017-07-01

    Longitudinal design. The study determined the effects of two forms of exercise training on the abundance of two proteins, (glucose transporter-4 [GLUT-4], adenosine monophosphate kinase [AMPK]) involved in glucose utilization and the transcriptional coactivator that regulates the genes involved in energy metabolism and mitochondrial biogenesis (peroxisome proliferator-activated receptor (PPAR) coactivator 1 alpha [PGC-1α]), in muscles in men with chronic motor-complete spinal cord injury (SCI). Clinical trial at a Medical Center. Nine men with chronic motor-complete SCI participated in functional electrical stimulation lower extremity cycling (FES-LEC; n = 4) or arm cycling ergometer (arm-cycling ergometer [ACE]; n = 5) 5 days/week for 16 weeks. Whole body composition was measured by dual energy X-ray absorptiometry. An intravenous glucose tolerance test was performed to measure glucose effectiveness (Sg) and insulin sensitivity (Si). Muscle biopsies of the right vastus lateralis (VL) and triceps muscles were collected one week prior to and post the exercise training intervention. Neither training intervention altered body composition or carbohydrate metabolism. GLUT-4 increased by 3.8 fold in the VL after FES training and increased 0.6 fold in the triceps after ACE training. PGC-1α increased by 2.3 fold in the VL after FES training and 3.8 fold in the triceps after ACE training. AMPK increased by 3.4 fold in the VL after FES training and in the triceps after ACE training. FES-LEC and ACE training were associated with greater protein expressions in the trained muscles by effectively influencing the abundance of GLUT-4, AMPK and PGC-1α. Thus, FES-LEC training of paralyzed muscle can modulate protein expression similar to that of trained and innervated muscle.

  18. Characterization of constitutive and putative differentially expressed mRNAs by means of expressed sequence tags, differential display reverse transcriptase-PCR and randomly amplified polymorphic DNA-PCR from the sand fly vector Lutzomyia longipalpis.

    PubMed

    Ramalho-Ortigão, J M; Temporal, P; de Oliveira , S M; Barbosa, A F; Vilela, M L; Rangel, E F; Brazil, R P; Traub-Cseko, Y M

    2001-01-01

    Molecular studies of insect disease vectors are of paramount importance for understanding parasite-vector relationship. Advances in this area have led to important findings regarding changes in vectors' physiology upon blood feeding and parasite infection. Mechanisms for interfering with the vectorial capacity of insects responsible for the transmission of diseases such as malaria, Chagas disease and dengue fever are being devised with the ultimate goal of developing transgenic insects. A primary necessity for this goal is information on gene expression and control in the target insect. Our group is investigating molecular aspects of the interaction between Leishmania parasites and Lutzomyia sand flies. As an initial step in our studies we have used random sequencing of cDNA clones from two expression libraries made from head/thorax and abdomen of sugar fed L. longipalpis for the identification of expressed sequence tags (EST). We applied differential display reverse transcriptase-PCR and randomly amplified polymorphic DNA-PCR to characterize differentially expressed mRNA from sugar and blood fed insects, and, in one case, from a L. (V.) braziliensis-infected L. longipalpis. We identified 37 cDNAs that have shown homology to known sequences from GeneBank. Of these, 32 cDNAs code for constitutive proteins such as zinc finger protein, glutamine synthetase, G binding protein, ubiquitin conjugating enzyme. Three are putative differentially expressed cDNAs from blood fed and Leishmania-infected midgut, a chitinase, a V-ATPase and a MAP kinase. Finally, two sequences are homologous to Drosophila melanogaster gene products recently discovered through the Drosophila genome initiative.

  19. Contesting the Constitution: The Constitutional Dialogues.

    ERIC Educational Resources Information Center

    Hilenski, Ferdinand Alexi

    This historical dramatization, prepared for presentation at the 1985 Wyoming Chatauqua, contains three dialogues, set during the administration of President Thomas Jefferson and presenting the issues surrounding the drafting and ratification of the U.S. Constitution. The dialogues are designed to be presented in three segments to permit discussion…

  20. Molecular Characterization of Aquaporin 1 and Aquaporin 3 from the Gills of the African Lungfish, Protopterus annectens, and Changes in Their Branchial mRNA Expression Levels and Protein Abundance during Three Phases of Aestivation.

    PubMed

    Chng, You R; Ong, Jasmine L Y; Ching, Biyun; Chen, Xiu L; Hiong, Kum C; Wong, Wai P; Chew, Shit F; Lam, Siew H; Ip, Yuen K

    2016-01-01

    African lungfishes can undergo long periods of aestivation on land during drought. During aestivation, lungfishes are confronted with desiccation and dehydration, and their gills become non-functional and covered with a thick layer of dried mucus. Aquaporins (Aqps) are a superfamily of integral membrane proteins which generally facilitate the permeation of water through plasma membranes. This study aimed to obtain the complete cDNA coding sequences of aqp1 and aqp3 from the gills of Protopterus annectens , and to determine their branchial mRNA and protein expression levels during the induction, maintenance and arousal phases of aestivation. Dendrogramic analyses of the deduced Aqp1 and Aqp3 amino acid sequences of P. annectens revealed their close relationships with those of Latimeria chalumnae and tetrapods. During the induction phase, there were significant decreases in the transcript levels of aqp1 and aqp3 in the gills of P. annectens , but the branchial Aqp1 and Aqp3 protein abundance remained unchanged. As changes in transcription might precede changes in translation, this could be regarded as an adaptive response to decrease the protein abundance of Aqp1 and Aqp3 in the subsequent maintenance phase of aestivation. As expected, the branchial transcript levels and protein abundance of aqp1 /Aqp1 and aqp3 /Aqp3 were significantly down-regulated during the maintenance phase, probably attributable to the shutdown of branchial functions and the cessation of volume regulation of branchial epithelial cells. Additionally, these changes could reduce the loss of water through branchial epithelial surfaces, supplementing the anti-desiccating property of the dried mucus. Upon arousal, it was essential for the lungfish to restore branchial functions. Indeed, the protein abundance of Aqp1 recovered partially, with complete recovery of mRNA expression level and protein abundance of Aqp3, in the gills of P. annectens after 3 days of arousal. These results provide insights

  1. Abundance of introduced species at home predicts abundance away in herbaceous communities

    USGS Publications Warehouse

    Firn, Jennifer; Moore, Joslin L.; MacDougall, Andrew S.; Borer, Elizabeth T.; Seabloom, Eric W.; HilleRisLambers, Janneke; Harpole, W. Stanley; Cleland, Elsa E.; Brown, Cynthia S.; Knops, Johannes M.H.; Prober, Suzanne M.; Pyke, David A.; Farrell, Kelly A.; Bakker, John D.; O'Halloran, Lydia R.; Adler, Peter B.; Collins, Scott L.; D'Antonio, Carla M.; Crawley, Michael J.; Wolkovich, Elizabeth M.; La Pierre, Kimberly J.; Melbourne, Brett A.; Hautier, Yann; Morgan, John W.; Leakey, Andrew D.B.; Kay, Adam; McCulley, Rebecca; Davies, Kendi F.; Stevens, Carly J.; Chu, Cheng-Jin; Holl, Karen D.; Klein, Julia A.; Fay, Phillip A.; Hagenah, Nicole; Kirkman, Kevin P.; Buckley, Yvonne M.

    2011-01-01

    Many ecosystems worldwide are dominated by introduced plant species, leading to loss of biodiversity and ecosystem function. A common but rarely tested assumption is that these plants are more abundant in introduced vs. native communities, because ecological or evolutionary-based shifts in populations underlie invasion success. Here, data for 26 herbaceous species at 39 sites, within eight countries, revealed that species abundances were similar at native (home) and introduced (away) sites - grass species were generally abundant home and away, while forbs were low in abundance, but more abundant at home. Sites with six or more of these species had similar community abundance hierarchies, suggesting that suites of introduced species are assembling similarly on different continents. Overall, we found that substantial changes to populations are not necessarily a pre-condition for invasion success and that increases in species abundance are unusual. Instead, abundance at home predicts abundance away, a potentially useful additional criterion for biosecurity programmes.

  2. MLH1 constitutional and somatic methylation in patients with MLH1 negative tumors fulfilling the revised Bethesda criteria

    PubMed Central

    Crucianelli, Francesca; Tricarico, Rossella; Turchetti, Daniela; Gorelli, Greta; Gensini, Francesca; Sestini, Roberta; Giunti, Laura; Pedroni, Monica; Ponz de Leon, Maurizio; Civitelli, Serenella; Genuardi, Maurizio

    2014-01-01

    Lynch syndrome (LS) is a tumor predisposing condition caused by constitutional defects in genes coding for components of the mismatch repair (MMR) apparatus. While hypermethylation of the promoter of the MMR gene MLH1 occurs in about 15% of colorectal cancer samples, it has also been observed as a constitutional alteration, in the absence of DNA sequence mutations, in a small number of LS patients. In order to obtain further insights on the phenotypic characteristics of MLH1 epimutation carriers, we investigated the somatic and constitutional MLH1 methylation status of 14 unrelated subjects with a suspicion of LS who were negative for MMR gene constitutional mutations and whose tumors did not express the MLH1 protein. A novel case of constitutional MLH1 epimutation was identified. This patient was affected with multiple primary tumors, including breast cancer, diagnosed starting from the age of 55 y. Investigation of her offspring by allele specific expression revealed that the epimutation was not stable across generations. We also found MLH1 hypermethylation in cancer samples from 4 additional patients who did not have evidence of constitutional defects. These patients had some characteristics of LS, namely early age at onset and/or positive family history, raising the possibility of genetic influences in the establishment of somatic MLH1 methylation. PMID:25437057

  3. Abscisic Acid-Dependent and -Independent Expression of the Carrot Late-Embryogenesis-Abundant-Class Gene Dc3 in Transgenic Tobacco Seedlings1

    PubMed Central

    Siddiqui, Najeeb U.; Chung, Hwa-Jee; Thomas, Terry L.; Drew, Malcolm C.

    1998-01-01

    We studied the expression of three promoter 5′ deletion constructs (−218, −599, and −1312) of the LEA (late embryogenesis abundant)-class gene Dc3 fused to β-glucuronidase (GUS), where each construct value refers to the number of base pairs upstream of the transcription start site at which the deletion occurred. The Dc3 gene is noted for its induction by abscisic acid (ABA), but its response to other plant hormones and various environmental stresses has not been reported previously for vegetative cells. Fourteen-day-old transgenic tobacco (Nicotiana tabacum L.) seedlings were exposed to dehydration, hypoxia, salinity, exogenous ethylene, or exogenous methyl jasmonate (MeJa). GUS activity was quantified fluorimetrically and expression was observed by histochemical staining of the seedlings. An increase in GUS activity was observed in plants with constructs −599 and −1312 in response to dehydration and salinity within 6 h of stress, and at 12 h in response to hypoxia. No increase in endogenous ABA was found in any of the three lines, even after 72 h of hypoxia. An ABA-independent increase in GUS activity was observed when endogenous ABA biosynthesis was blocked by fluridone and plants were exposed to 5 μL L−1 ethylene in air or 100 μm MeJa. Virtually no expression was observed in construct −218 in response to dehydration, salinity, or MeJa, but there was a moderate response to ethylene and hypoxia. This suggests that the region between −218 and −599 is necessary for ABA (dehydration and salinity)- and MeJa-dependent expression, whereas ethylene-mediated expression does not require this region of the promoter. PMID:9847092

  4. Distribution and abundance of hematophagous flies (Glossinidae, Stomoxys, and Tabanidae) in two national parks of Gabon.

    PubMed

    Bitome Essono, Paul Yannick; Dechaume-Moncharmont, François-Xavier; Mavoungou, Jacques; Obiang Mba, Régis; Duvallet, Gérard; Bretagnolle, François

    2015-01-01

    In order to minimize risks of pathogen transmission with the development of ecotourism in Gabon, a seasonal inventory has been performed in five contrasted biotopes in Ivindo (INP) and Moukalaba-Doudou (MDNP) National Parks. A total of 10,033 hematophagous flies were captured. The Glossinidae, with six different species identified, was the most abundant group and constitutes about 60% of the captured flies compared to the Stomoxys (6 species also identified) and Tabanidae with 28% and 12%, respectively. The Glossinidae showed a higher rate of capture in primary forest and in research camps. In INP, the Stomoxys showed a higher rate of capture in secondary forest and at village borders, whereas in MDNP the Stomoxys were captured more in the savannah area. Thus, each fly group seemed to reach maximum abundance in different habitats. The Glossinidae were more abundant in primary forest and near research camps while Stomoxys were more abundant in secondary forest and savannah. The Tabanidae did not show a clear habitat preference. © P.Y. Bitome Essono et al., published by EDP Sciences, 2015.

  5. Proteoglycans in Leiomyoma and Normal Myometrium: Abundance, Steroid Hormone Control, and Implications for Pathophysiology.

    PubMed

    Barker, Nichole M; Carrino, David A; Caplan, Arnold I; Hurd, William W; Liu, James H; Tan, Huiqing; Mesiano, Sam

    2016-03-01

    Uterine leiomyoma are a common benign pelvic tumors composed of modified smooth muscle cells and a large amount of extracellular matrix (ECM). The proteoglycan composition of the leiomyoma ECM is thought to affect pathophysiology of the disease. To test this hypothesis, we examined the abundance (by immunoblotting) and expression (by quantitative real-time polymerase chain reaction) of the proteoglycans biglycan, decorin, and versican in leiomyoma and normal myometrium and determined whether expression is affected by steroid hormones and menstrual phase. Leiomyoma and normal myometrium were collected from women (n = 17) undergoing hysterectomy or myomectomy. In vitro studies were performed on immortalized leiomyoma (UtLM) and normal myometrial (hTERT-HM) cells with and without exposure to estradiol and progesterone. In leiomyoma tissue, abundance of decorin messenger RNA (mRNA) and protein were 2.6-fold and 1.4-fold lower, respectively, compared with normal myometrium. Abundance of versican mRNA was not different between matched samples, whereas versican protein was increased 1.8-fold in leiomyoma compared with myometrium. Decorin mRNA was 2.4-fold lower in secretory phase leiomyoma compared with proliferative phase tissue. In UtLM cells, progesterone decreased the abundance of decorin mRNA by 1.3-fold. Lower decorin expression in leiomyoma compared with myometrium may contribute to disease growth and progression. As decorin inhibits the activity of specific growth factors, its reduced level in the leiomyoma cell microenvironment may promote cell proliferation and ECM deposition. Our data suggest that decorin expression in leiomyoma is inhibited by progesterone, which may be a mechanism by which the ovarian steroids affect leiomyoma growth and disease progression. © The Author(s) 2015.

  6. [Formation of individual somatotype parameters and features of constitutional organization in 7-12-year-old boys (longitudinal)].

    PubMed

    Kornienko, I A; Panasiuk, T V; Tambovtseva, R V

    1997-01-01

    Individual somatotype parameters and peculiarities of constitution in 7-12 years boys were evaluated in the present investigation. The age range studied was shown to be divided on 3 stages. Regular growth processes with the prevalence of "infantal" proportions occur at the age of 7-9. Signs of definite constitutional type are expressed yet insufficiently. The age of 10-11 is transitional which shows in delay of muscle growth. At the age of 11-12 prepubescent sets in, during which features of constitution types and appropriate somatotype parameters are distinctly manifested.

  7. Expression Atlas: gene and protein expression across multiple studies and organisms

    PubMed Central

    Tang, Y Amy; Bazant, Wojciech; Burke, Melissa; Fuentes, Alfonso Muñoz-Pomer; George, Nancy; Koskinen, Satu; Mohammed, Suhaib; Geniza, Matthew; Preece, Justin; Jarnuczak, Andrew F; Huber, Wolfgang; Stegle, Oliver; Brazma, Alvis; Petryszak, Robert

    2018-01-01

    Abstract Expression Atlas (http://www.ebi.ac.uk/gxa) is an added value database that provides information about gene and protein expression in different species and contexts, such as tissue, developmental stage, disease or cell type. The available public and controlled access data sets from different sources are curated and re-analysed using standardized, open source pipelines and made available for queries, download and visualization. As of August 2017, Expression Atlas holds data from 3,126 studies across 33 different species, including 731 from plants. Data from large-scale RNA sequencing studies including Blueprint, PCAWG, ENCODE, GTEx and HipSci can be visualized next to each other. In Expression Atlas, users can query genes or gene-sets of interest and explore their expression across or within species, tissues, developmental stages in a constitutive or differential context, representing the effects of diseases, conditions or experimental interventions. All processed data matrices are available for direct download in tab-delimited format or as R-data. In addition to the web interface, data sets can now be searched and downloaded through the Expression Atlas R package. Novel features and visualizations include the on-the-fly analysis of gene set overlaps and the option to view gene co-expression in experiments investigating constitutive gene expression across tissues or other conditions. PMID:29165655

  8. Circular RNA Expression: Its Potential Regulation and Function.

    PubMed

    Salzman, Julia

    2016-05-01

    In 2012, a new feature of eukaryotic gene expression emerged: ubiquitous expression of circular RNA (circRNA) from genes traditionally thought to express messenger or linear noncoding (nc)RNA only. CircRNAs are covalently closed, circular RNA molecules that typically comprise exonic sequences and are spliced at canonical splice sites. This feature of gene expression was first recognized in humans and mouse, but it quickly emerged that it was common across essentially all eukaryotes studied by molecular biologists. CircRNA abundance, and even which alternatively spliced circRNA isoforms are expressed, varies by cell type and can exceed the abundance of the traditional linear mRNA or ncRNA transcript. CircRNAs are enriched in the brain and increase in abundance during fetal development. Together, these features raise fundamental questions regarding the regulation of circRNA in cis and in trans, and its function. Copyright © 2016. Published by Elsevier Ltd.

  9. Constitutive mRNA expression and protein activity levels of nine ABC efflux transporters in seven permanent cell lines derived from different tissues of rainbow trout (Oncorhynchus mykiss).

    PubMed

    Fischer, Stephan; Loncar, Jovica; Zaja, Roko; Schnell, Sabine; Schirmer, Kristin; Smital, Tvrtko; Luckenbach, Till

    2011-01-25

    Permanent fish cell lines have become common model systems for determining ecotoxicological effects of pollutants. For these cell lines little is known on the cellular active transport mechanisms that control the amount of a compound entering the cell, such as the MXR (multixenobiotic resistance) system mediated by ATP binding cassette (ABC) transport proteins. Therefore, for toxic evaluation of chemicals with those cells information on MXR is important. We here present data on constitutive mRNA expression and protein activity levels of a series of ABC efflux transporters in seven permanent cell lines derived from liver (RTL-W1; R1) and liver hepatoma (RTH-149), gill (RTgill-W1), gonad (RTG-2), gut (RTgutGC) and brain (RTbrain) of rainbow trout (Oncorhynchus mykiss). In addition to known transporters abcb1 (designated here abcb1a), abcb11, abcc1-3, abcc5 and abcg2, we quantified expression levels of a newly identified abcb1 isoform (abcb1b) and abcc4, previously unknown in trout. Quantitative real time PCR (qPCR) indicated that mRNA of the examined ABC transporters was constitutively expressed in all cell lines. Transporter mRNA expression patterns were similar in all cell lines, with expression levels of abcc transporters being 80 to over 1000 fold higher than for abcg2, abcb1a/b and abcb11 (abcc1-5>abcg2>abcb1a/b, 11). Transporter activity in the cell lines was determined by measuring uptake of transporter type specific fluorescent substrates in the presence of activity inhibitors. The combination of the ABCB1 and ABCC transporter substrate calcein-AM with inhibitors cyclosporine A, PSC833 and MK571 resulted in a concentration-dependent fluorescence increase of up to 3-fold, whereas reversin 205 caused a slight, but not concentration-dependent fluorescence increase. Accumulation of the dyes Hoechst 33342 and 2',7'-dichlorodihydrofluorescein diacetate was basically unchanged in the presence of Ko134 and taurocholate, respectively, indicating low Abcg2 and Abcb11

  10. LentiPro26: novel stable cell lines for constitutive lentiviral vector production.

    PubMed

    Tomás, H A; Rodrigues, A F; Carrondo, M J T; Coroadinha, A S

    2018-03-27

    Lentiviral vectors (LVs) are excellent tools to promote gene transfer and stable gene expression. Their potential has been already demonstrated in gene therapy clinical trials for the treatment of diverse disorders. For large scale LV production, a stable producer system is desirable since it allows scalable and cost-effective viral productions, with increased reproducibility and safety. However, the development of stable systems has been challenging and time-consuming, being the selection of cells presenting high expression levels of Gag-Pro-Pol polyprotein and the cytotoxicity associated with some viral components, the main limitations. Hereby is described the establishment of a new LV producer cell line using a mutated less active viral protease to overcome potential cytotoxic limitations. The stable transfection of bicistronic expression cassettes with re-initiation of the translation mechanism enabled the generation of LentiPro26 packaging populations supporting high titers. Additionally, by skipping intermediate clone screening steps and performing only one final clone screening, it was possible to save time and generate LentiPro26-A59 cell line, that constitutively produces titers above 10 6 TU.mL -1 .day -1 , in less than six months. This work constitutes a step forward towards the development of improved LV producer cell lines, aiming to efficiently supply the clinical expanding gene therapy applications.

  11. A cherry protein and its gene, abundantly expressed in ripening fruit, have been identified as thaumatin-like.

    PubMed

    Fils-Lycaon, B R; Wiersma, P A; Eastwell, K C; Sautiere, P

    1996-05-01

    A 29-kD polypeptide is the most abundant soluble protein in ripe cherry fruit (Prunus avium L); accumulation begins at the onset of ripening as the fruit turns from yellow to red. This protein was extracted from ripe cherries and purified by size-exclusion and ion-exchange chromatography. Antibodies to the purified protein were used to screen a cDNA library from ripe cherries. Numerous recombinant plaques reacted positively with the antibodies; the DNA sequence of representative clones encoded a polypeptide of 245 amino acid residues. A signal peptide was indicated, and the predicted mature protein corresponded to the purified protein in size (23.3 kD, by mass spectrometry) and isoelectric point (4.2). A search of known protein sequences revealed a strong similarity between this polypeptide and the thaumatin family of pathogenesis-related proteins. The cherry thaumatin-like protein does not have a sweet taste, and no antifungal activity was seen in preliminary assays. Expression of the protein appears to be regulated at the gene level, with mRNA levels at their highest in the ripe fruit.

  12. A cherry protein and its gene, abundantly expressed in ripening fruit, have been identified as thaumatin-like.

    PubMed Central

    Fils-Lycaon, B R; Wiersma, P A; Eastwell, K C; Sautiere, P

    1996-01-01

    A 29-kD polypeptide is the most abundant soluble protein in ripe cherry fruit (Prunus avium L); accumulation begins at the onset of ripening as the fruit turns from yellow to red. This protein was extracted from ripe cherries and purified by size-exclusion and ion-exchange chromatography. Antibodies to the purified protein were used to screen a cDNA library from ripe cherries. Numerous recombinant plaques reacted positively with the antibodies; the DNA sequence of representative clones encoded a polypeptide of 245 amino acid residues. A signal peptide was indicated, and the predicted mature protein corresponded to the purified protein in size (23.3 kD, by mass spectrometry) and isoelectric point (4.2). A search of known protein sequences revealed a strong similarity between this polypeptide and the thaumatin family of pathogenesis-related proteins. The cherry thaumatin-like protein does not have a sweet taste, and no antifungal activity was seen in preliminary assays. Expression of the protein appears to be regulated at the gene level, with mRNA levels at their highest in the ripe fruit. PMID:8685266

  13. Estimating abundance

    USGS Publications Warehouse

    Sutherland, Chris; Royle, Andy

    2016-01-01

    This chapter provides a non-technical overview of ‘closed population capture–recapture’ models, a class of well-established models that are widely applied in ecology, such as removal sampling, covariate models, and distance sampling. These methods are regularly adopted for studies of reptiles, in order to estimate abundance from counts of marked individuals while accounting for imperfect detection. Thus, the chapter describes some classic closed population models for estimating abundance, with considerations for some recent extensions that provide a spatial context for the estimation of abundance, and therefore density. Finally, the chapter suggests some software for use in data analysis, such as the Windows-based program MARK, and provides an example of estimating abundance and density of reptiles using an artificial cover object survey of Slow Worms (Anguis fragilis).

  14. GDF-15 is abundantly expressed in plexiform lesions in patients with pulmonary arterial hypertension and affects proliferation and apoptosis of pulmonary endothelial cells

    PubMed Central

    2011-01-01

    Background Growth-differentiation factor-15 (GDF-15) is a stress-responsive, transforming growth factor-β-related cytokine, which has recently been reported to be elevated in serum of patients with idiopathic pulmonary arterial hypertension (IPAH). The aim of the study was to examine the expression and biological roles of GDF-15 in the lung of patients with pulmonary arterial hypertension (PAH). Methods GDF-15 expression in normal lungs and lung specimens of PAH patients were studied by real-time RT-PCR and immunohistochemistry. Using laser-assisted micro-dissection, GDF-15 expression was further analyzed within vascular compartments of PAH lungs. To elucidate the role of GDF-15 on endothelial cells, human pulmonary microvascular endothelial cells (HPMEC) were exposed to hypoxia and laminar shear stress. The effects of GDF-15 on the proliferation and cell death of HPMEC were studied using recombinant GDF-15 protein. Results GDF-15 expression was found to be increased in lung specimens from PAH patients, com-pared to normal lungs. GDF-15 was abundantly expressed in pulmonary vascular endothelial cells with a strong signal in the core of plexiform lesions. HPMEC responded with marked upregulation of GDF-15 to hypoxia and laminar shear stress. Apoptotic cell death of HPMEC was diminished, whereas HPMEC proliferation was either increased or decreased depending of the concentration of recombinant GDF-15 protein. Conclusions GDF-15 expression is increased in PAH lungs and appears predominantly located in vascular endothelial cells. The expression pattern as well as the observed effects on proliferation and apoptosis of pulmonary endothelial cells suggest a role of GDF-15 in the homeostasis of endothelial cells in PAH patients. PMID:21548946

  15. Natural Genetic Variation Influences Protein Abundances in C. elegans Developmental Signalling Pathways

    PubMed Central

    Singh, Kapil Dev; Roschitzki, Bernd; Snoek, L. Basten; Grossmann, Jonas; Zheng, Xue; Elvin, Mark; Kamkina, Polina; Schrimpf, Sabine P.; Poulin, Gino B.; Kammenga, Jan E.; Hengartner, Michael O.

    2016-01-01

    Complex traits, including common disease-related traits, are affected by many different genes that function in multiple pathways and networks. The apoptosis, MAPK, Notch, and Wnt signalling pathways play important roles in development and disease progression. At the moment we have a poor understanding of how allelic variation affects gene expression in these pathways at the level of translation. Here we report the effect of natural genetic variation on transcript and protein abundance involved in developmental signalling pathways in Caenorhabditis elegans. We used selected reaction monitoring to analyse proteins from the abovementioned four pathways in a set of recombinant inbred lines (RILs) generated from the wild-type strains N2 (Bristol) and CB4856 (Hawaii) to enable quantitative trait locus (QTL) mapping. About half of the cases from the 44 genes tested showed a statistically significant change in protein abundance between various strains, most of these were however very weak (below 1.3-fold change). We detected a distant QTL on the left arm of chromosome II that affected protein abundance of the phosphatidylserine receptor protein PSR-1, and two separate QTLs that influenced embryonic and ionizing radiation-induced apoptosis on chromosome IV. Our results demonstrate that natural variation in C. elegans is sufficient to cause significant changes in signalling pathways both at the gene expression (transcript and protein abundance) and phenotypic levels. PMID:26985669

  16. Pendrin protein abundance in the kidney is regulated by nitric oxide and cAMP.

    PubMed

    Thumova, Monika; Pech, Vladimir; Froehlich, Otto; Agazatian, Diana; Wang, Xiaonan; Verlander, Jill W; Kim, Young Hee; Wall, Susan M

    2012-09-15

    Pendrin is a Cl(-)/HCO(3)(-) exchanger, expressed in the apical regions of some intercalated cell subtypes, and is critical in the pressor response to angiotensin II. Since angiotensin type 1 receptor inhibitors reduce renal pendrin protein abundance in mice in vivo through a mechanism that is dependent on nitric oxide (NO), we asked if NO modulates renal pendrin expression in vitro and explored the mechanism by which it occurs. Thus we quantified pendrin protein abundance by confocal fluorescent microscopy in cultured mouse cortical collecting ducts (CCDs) and connecting tubules (CNTs). After overnight culture, CCDs maintain their tubular structure and maintain a solute gradient when perfused in vitro. Pendrin protein abundance increased 67% in CNT and 53% in CCD when NO synthase was inhibited (N(G)-nitro-L-arginine methyl ester, 100 μM), while NO donor (DETA NONOate, 200 μM) application reduced pendrin protein by ∼33% in the CCD and CNT. When CNTs were cultured in the presence of the guanylyl cyclase inhibitor 1H-[1,2,4] oxadiazolo[4,3-a]quinoxalin-1-one (10 μM), NO donors did not alter pendrin abundance. Conversely, pendrin protein abundance rose when cAMP content was increased by the application of an adenylyl cyclase agonist (forskolin, 10 μM), a cAMP analog (8-bromo-cAMP, 1 mM), or a phosphodiesterase inhibitor (BAY60-7550, 50 μM). Since NO reduces cellular cAMP in the CNT, we asked if NO reduces pendrin abundance by reducing cAMP. With blockade of cGMP-stimulated phosphodiesterase II, NO did not alter pendrin protein abundance. We conclude that NO acts through cAMP to reduce pendrin total protein abundance by enhancing cAMP degradation.

  17. Pendrin protein abundance in the kidney is regulated by nitric oxide and cAMP

    PubMed Central

    Thumova, Monika; Pech, Vladimir; Froehlich, Otto; Agazatian, Diana; Wang, Xiaonan; Verlander, Jill W.; Kim, Young Hee

    2012-01-01

    Pendrin is a Cl−/HCO3− exchanger, expressed in the apical regions of some intercalated cell subtypes, and is critical in the pressor response to angiotensin II. Since angiotensin type 1 receptor inhibitors reduce renal pendrin protein abundance in mice in vivo through a mechanism that is dependent on nitric oxide (NO), we asked if NO modulates renal pendrin expression in vitro and explored the mechanism by which it occurs. Thus we quantified pendrin protein abundance by confocal fluorescent microscopy in cultured mouse cortical collecting ducts (CCDs) and connecting tubules (CNTs). After overnight culture, CCDs maintain their tubular structure and maintain a solute gradient when perfused in vitro. Pendrin protein abundance increased 67% in CNT and 53% in CCD when NO synthase was inhibited (NG-nitro-l-arginine methyl ester, 100 μM), while NO donor (DETA NONOate, 200 μM) application reduced pendrin protein by ∼33% in the CCD and CNT. When CNTs were cultured in the presence of the guanylyl cyclase inhibitor 1H-[1,2,4] oxadiazolo[4,3-a]quinoxalin-1-one (10 μM), NO donors did not alter pendrin abundance. Conversely, pendrin protein abundance rose when cAMP content was increased by the application of an adenylyl cyclase agonist (forskolin, 10 μM), a cAMP analog (8-bromo-cAMP, 1 mM), or a phosphodiesterase inhibitor (BAY60-7550, 50 μM). Since NO reduces cellular cAMP in the CNT, we asked if NO reduces pendrin abundance by reducing cAMP. With blockade of cGMP-stimulated phosphodiesterase II, NO did not alter pendrin protein abundance. We conclude that NO acts through cAMP to reduce pendrin total protein abundance by enhancing cAMP degradation. PMID:22811483

  18. An Abundant Evolutionarily Conserved CSB-PiggyBac Fusion Protein Expressed in Cockayne Syndrome

    PubMed Central

    Newman, John C.; Bailey, Arnold D.; Fan, Hua-Ying; Pavelitz, Thomas; Weiner, Alan M.

    2008-01-01

    Cockayne syndrome (CS) is a devastating progeria most often caused by mutations in the CSB gene encoding a SWI/SNF family chromatin remodeling protein. Although all CSB mutations that cause CS are recessive, the complete absence of CSB protein does not cause CS. In addition, most CSB mutations are located beyond exon 5 and are thought to generate only C-terminally truncated protein fragments. We now show that a domesticated PiggyBac-like transposon PGBD3, residing within intron 5 of the CSB gene, functions as an alternative 3′ terminal exon. The alternatively spliced mRNA encodes a novel chimeric protein in which CSB exons 1–5 are joined in frame to the PiggyBac transposase. The resulting CSB-transposase fusion protein is as abundant as CSB protein itself in a variety of human cell lines, and continues to be expressed by primary CS cells in which functional CSB is lost due to mutations beyond exon 5. The CSB-transposase fusion protein has been highly conserved for at least 43 Myr since the divergence of humans and marmoset, and appears to be subject to selective pressure. The human genome contains over 600 nonautonomous PGBD3-related MER85 elements that were dispersed when the PGBD3 transposase was last active at least 37 Mya. Many of these MER85 elements are associated with genes which are involved in neuronal development, and are known to be regulated by CSB. We speculate that the CSB-transposase fusion protein has been conserved for host antitransposon defense, or to modulate gene regulation by MER85 elements, but may cause CS in the absence of functional CSB protein. PMID:18369450

  19. Identification of Differentially Abundant Proteins of Edwardsiella ictaluri during Iron Restriction

    PubMed Central

    Dumpala, Pradeep R.; Peterson, Brian C.; Lawrence, Mark L.; Karsi, Attila

    2015-01-01

    Edwardsiella ictaluri is a Gram-negative facultative anaerobe intracellular bacterium that causes enteric septicemia in channel catfish. Iron is an essential inorganic nutrient of bacteria and is crucial for bacterial invasion. Reduced availability of iron by the host may cause significant stress for bacterial pathogens and is considered a signal that leads to significant alteration in virulence gene expression. However, the precise effect of iron-restriction on E. ictaluri protein abundance is unknown. The purpose of this study was to identify differentially abundant proteins of E. ictaluri during in vitro iron-restricted conditions. We applied two-dimensional difference in gel electrophoresis (2D-DIGE) for determining differentially abundant proteins and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF/TOF MS) for protein identification. Gene ontology and pathway-based functional modeling of differentially abundant proteins was also conducted. A total of 50 unique differentially abundant proteins at a minimum of 2-fold (p ≤ 0.05) difference in abundance due to iron-restriction were detected. The numbers of up- and down-regulated proteins were 37 and 13, respectively. We noted several proteins, including EsrB, LamB, MalM, MalE, FdaA, and TonB-dependent heme/hemoglobin receptor family proteins responded to iron restriction in E. ictaluri. PMID:26168192

  20. Constitutively Active Akt Induces Ectodermal Defects and Impaired Bone Morphogenetic Protein Signaling

    PubMed Central

    Segrelles, Carmen; Moral, Marta; Lorz, Corina; Santos, Mirentxu; Lu, Jerry; Cascallana, José Luis; Lara, M. Fernanda; Carbajal, Steve; Martínez-Cruz, Ana Belén; García-Escudero, Ramón; Beltran, Linda; Segovia, José C.; Bravo, Ana

    2008-01-01

    Aberrant activation of the Akt pathway has been implicated in several human pathologies including cancer. However, current knowledge on the involvement of Akt signaling in development is limited. Previous data have suggested that Akt-mediated signaling may be an essential mediator of epidermal homeostasis through cell autonomous and noncell autonomous mechanisms. Here we report the developmental consequences of deregulated Akt activity in the basal layer of stratified epithelia, mediated by the expression of a constitutively active Akt1 (myrAkt) in transgenic mice. Contrary to mice overexpressing wild-type Akt1 (Aktwt), these myrAkt mice display, in a dose-dependent manner, altered development of ectodermally derived organs such as hair, teeth, nails, and epidermal glands. To identify the possible molecular mechanisms underlying these alterations, gene profiling approaches were used. We demonstrate that constitutive Akt activity disturbs the bone morphogenetic protein-dependent signaling pathway. In addition, these mice also display alterations in adult epidermal stem cells. Collectively, we show that epithelial tissue development and homeostasis is dependent on proper regulation of Akt expression and activity. PMID:17959825

  1. A constitutive relation for the viscous flow of an oriented fiber assembly

    NASA Technical Reports Server (NTRS)

    Pipes, R. B.; Hearle, J. W. S.; Beaussart, A. J.; Sastry, A. M.; Okine, R. K.

    1991-01-01

    A constitutive relation for an equivalent, homogeneous fluid is developed for the anisotropic viscous flow of an oriented assembly of discontinuous fibers suspended in a viscous fluid. The anisotropic viscous compliance matrix can be expressed in terms of three constants by assuming the equivalent fluid to be incompressible and the microstructure to have axial symmetry (transversely isotropic). By means of a micromechanics analysis, the three terms of the constitutive relation are expressed in terms of the viscosity of the matrix fluid, the fiber aspect ratio, and the fiber volume fraction. A comparison of the viscosity terms reveals that the elongational viscosity in the fiber direction varies as the square of the fiber aspect ratio and a complex function of the fiber volume fraction. Furthermore, the ratio of the axial elongational viscosity to the transverse elongational viscosity and both axial and transverse shear viscosities was shown to be 10 exp 4 - 10 exp 6 for fiber aspect ratio of 100-1000, except at extreme values of the fiber volume fraction.

  2. Activation of the Constitutive Androstane Receptor induces hepatic lipogenesis and regulates Pnpla3 gene expression in a LXR-independent way

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Marmugi, Alice; Lukowicz, Céline; Lasserre, Freder

    The Constitutive Androstane Receptor (CAR, NR1I3) has been newly described as a regulator of energy metabolism. A relevant number of studies using animal models of obesity suggest that CAR activation could be beneficial on the metabolic balance. However, this remains controversial and the underlying mechanisms are still unknown. This work aimed to investigate the effect of CAR activation on hepatic energy metabolism during physiological conditions, i.e. in mouse models not subjected to metabolic/nutritional stress. Gene expression profiling in the liver of CAR knockout and control mice on chow diet and treated with a CAR agonist highlighted CAR-mediated up-regulations of lipogenicmore » genes, concomitant with neutral lipid accumulation. A strong CAR-mediated up-regulation of the patatin-like phospholipase domain-containing protein 3 (Pnpla3) was demonstrated. Pnpla3 is a gene whose polymorphism is associated with the pathogenesis of nonalcoholic fatty liver disease (NAFLD) development. This observation was confirmed in human hepatocytes treated with the antiepileptic drug and CAR activator, phenobarbital and in immortalized human hepatocytes treated with CITCO. Studying the molecular mechanisms controlling Pnpla3 gene expression, we demonstrated that CAR does not act by a direct regulation of Pnpla3 transcription or via the Liver X Receptor but may rather involve the transcription factor Carbohydrate Responsive Element-binding protein. These data provide new insights into the regulation by CAR of glycolytic and lipogenic genes and on pathogenesis of steatosis. This also raises the question concerning the impact of drugs and environmental contaminants in lipid-associated metabolic diseases. - Highlights: • Induction of hepatic glycolytic and lipogenic genes upon CAR activation by TCPOBOP. • These effects are not mediated by the nuclear receptor LXR. • CAR activation resulted in hepatic lipid accumulation. • Pnpla3 expression is regulated by CAR in mouse

  3. Dynamic hybrid materials for constitutional self-instructed membranes.

    PubMed

    Cazacu, Adinela; Legrand, Yves-Marie; Pasc, Andreea; Nasr, Gihane; Van der Lee, Arie; Mahon, Eugene; Barboiu, Mihail

    2009-05-19

    Constitutional self-instructed membranes were developed and used for mimicking the adaptive structural functionality of natural ion-channel systems. These membranes are based on dynamic hybrid materials in which the functional self-organized macrocycles are reversibly connected with the inorganic silica through hydrophobic noncovalent interactions. Supramolecular columnar ion-channel architectures can be generated by reversible confinement within scaffolding hydrophobic silica mesopores. They can be structurally determined by using X-ray diffraction and morphologically tuned by alkali-salts templating. From the conceptual point of view, these membranes express a synergistic adaptive behavior: the simultaneous binding of the fittest cation and its anion would be a case of "homotropic allosteric interactions," because in time it increases the transport efficiency of the pore-contained superstructures by a selective evolving process toward the fittest ion channel. The hybrid membranes presented here represent dynamic constitutional systems evolving over time to form the fittest ion channels from a library of molecular and supramolecular components, or selecting the fittest ion pairs from a mixture of salts demonstrating flexible adaptation.

  4. Dynamic hybrid materials for constitutional self-instructed membranes

    PubMed Central

    Cazacu, Adinela; Legrand, Yves-Marie; Pasc, Andreea; Nasr, Gihane; Van der Lee, Arie; Mahon, Eugene; Barboiu, Mihail

    2009-01-01

    Constitutional self-instructed membranes were developed and used for mimicking the adaptive structural functionality of natural ion-channel systems. These membranes are based on dynamic hybrid materials in which the functional self-organized macrocycles are reversibly connected with the inorganic silica through hydrophobic noncovalent interactions. Supramolecular columnar ion-channel architectures can be generated by reversible confinement within scaffolding hydrophobic silica mesopores. They can be structurally determined by using X-ray diffraction and morphologically tuned by alkali-salts templating. From the conceptual point of view, these membranes express a synergistic adaptive behavior: the simultaneous binding of the fittest cation and its anion would be a case of “homotropic allosteric interactions,” because in time it increases the transport efficiency of the pore-contained superstructures by a selective evolving process toward the fittest ion channel. The hybrid membranes presented here represent dynamic constitutional systems evolving over time to form the fittest ion channels from a library of molecular and supramolecular components, or selecting the fittest ion pairs from a mixture of salts demonstrating flexible adaptation. PMID:19416909

  5. A novel hsp110-related gene, apg-1, that is abundantly expressed in the testis responds to a low temperature heat shock rather than the traditional elevated temperatures.

    PubMed

    Kaneko, Y; Nishiyama, H; Nonoguchi, K; Higashitsuji, H; Kishishita, M; Fujita, J

    1997-01-31

    We isolated a novel hsp110-related gene, apg-1, from a testis cDNA library. The apg-1 transcripts were constitutively expressed in the testicular germ cells and, in some degree, most tissues examined. In a mouse TAMA26 Sertoli cell line, apg-1 transcripts were induced in 2 h by a temperature shift from 32 to 39 degrees C, but not by a shift from 37 to 42 degrees C, the traditional heat stress, or a shift from 32 to 42 degrees C. The heat response pattern of hsp110 expression was similar to that of apg-1. Although induction of a hsp70 transcript was observed in 2 h by a shift from 32 to 39 degrees C, the induction was more apparent by a shift from 37 to 42 degrees C or from 32 to 42 degrees C. Essentially similar differential response patterns were observed among these genes in NIH/3T3 fibroblasts as well. The nuclear run-on assay and the native gel mobility shift assay demonstrated that, by the 32 to 39 degrees C temperature shift, the apg-1 gene was transcriptionally activated, and heat shock factor 1 bound to the heat shock elements in the 5'-flanking region of the apg-1 gene. These results demonstrated that expressions of apg-1, hsp110, and hsp70 could be heat-induced at a temperature lower than the traditional elevated temperatures in somatic cells of both testis and nontestis origin and suggest that the mechanisms regulating the transcript levels of apg-1 and hsp110 are different from those of hsp70. Furthermore, the constitutive expression in germ cells suggests that APG-1 plays a specific role in spermatogenesis as well as in stress response.

  6. One Percent Determination of the Primordial Deuterium Abundance

    NASA Astrophysics Data System (ADS)

    Cooke, Ryan J.; Pettini, Max; Steidel, Charles C.

    2018-03-01

    We report a reanalysis of a near-pristine absorption system, located at a redshift {z}abs}=2.52564 toward the quasar Q1243+307, based on the combination of archival and new data obtained with the HIRES echelle spectrograph on the Keck telescope. This absorption system, which has an oxygen abundance [O/H] = ‑2.769 ± 0.028 (≃1/600 of the solar abundance), is among the lowest metallicity systems currently known where a precise measurement of the deuterium abundance is afforded. Our detailed analysis of this system concludes, on the basis of eight D I absorption lines, that the deuterium abundance of this gas cloud is {log}}10({{D}}/{{H}})=-4.622+/- 0.015, which is in very good agreement with the results previously reported by Kirkman et al., but with an improvement on the precision of this single measurement by a factor of ∼3.5. Combining this new estimate with our previous sample of six high precision and homogeneously analyzed D/H measurements, we deduce that the primordial deuterium abundance is {log}}10{({{D}}/{{H}})}{{P}}=-4.5974+/- 0.0052 or, expressed as a linear quantity, {10}5{({{D}}/{{H}})}{{P}}=2.527+/- 0.030; this value corresponds to a one percent determination of the primordial deuterium abundance. Combining our result with a big bang nucleosynthesis (BBN) calculation that uses the latest nuclear physics input, we find that the baryon density derived from BBN agrees to within 2σ of the latest results from the Planck cosmic microwave background data. Based on observations collected at the W.M. Keck Observatory which is operated as a scientific partnership among the California Institute of Technology, the University of California, and the National Aeronautics and Space Administration. The Observatory was made possible by the generous financial support of the W.M. Keck Foundation.

  7. The constitutive lipid droplet protein PLIN2 regulates autophagy in liver.

    PubMed

    Tsai, Tsung-Huang; Chen, Elaine; Li, Lan; Saha, Pradip; Lee, Hsiao-Ju; Huang, Li-Shin; Shelness, Gregory S; Chan, Lawrence; Chang, Benny Hung-Junn

    2017-07-03

    Excess triglyceride (TG) accumulation in the liver underlies fatty liver disease, a highly prevalent ailment. TG occurs in the liver sequestered in lipid droplets, the major lipid storage organelle. Lipid droplets are home to the lipid droplet proteins, the most abundant of which are the perilipins (PLINs), encoded by 5 different genes, Plin1 to Plin5. Of the corresponding gene products, PLIN2 is the only constitutive and ubiquitously expressed lipid droplet protein that has been used as a protein marker for lipid droplets. We and others reported that plin2 -/- mice have an ∼60% reduction in TG content, and are protected against fatty liver disease. Here we show that PLIN2 overexpression protects lipid droplets against macroautophagy/autophagy, whereas PLIN2 deficiency enhances autophagy and depletes hepatic TG. The enhanced autophagy in plin2 -/- mice protects against severe ER stress-induced hepatosteatosis and hepatocyte apoptosis. In contrast, hepatic TG depletion resulting from other genetic and pharmacological manipulations has no effect on autophagy. Importantly, PLIN2 deficiency lowers cellular TG content in wild-type mouse embryonic fibroblasts (MEFs) via enhanced autophagy, but does not affect cellular TG content in atg7 -/- MEFs that are devoid of autophagic function. Conversely, adenovirus-shAtg7-mediated hepatic Atg7 knockdown per se does not alter the hepatic TG level, suggesting a more complex regulation in vivo. In sum, PLIN2 guards its own house, the lipid droplet. PLIN2 overexpression protects against autophagy, and its downregulation stimulates TG catabolism via autophagy.

  8. RNA protein interactions governing expression of the most abundant protein in human body, type I collagen.

    PubMed

    Stefanovic, Branko

    2013-01-01

    Type I collagen is the most abundant protein in human body. The protein turns over slowly and its replacement synthesis is low. However, in wound healing or in pathological fibrosis the cells can increase production of type I collagen several hundred fold. This increase is predominantly due to posttranscriptional regulation, including increased half-life of collagen messenger RNAs (mRNAs) and their increased translatability. Type I collagen is composed of two α1 and one α2 polypeptides that fold into a triple helix. This stoichiometry is strictly regulated to prevent detrimental synthesis of α1 homotrimers. Collagen polypeptides are co-translationally modified and the rate of modifications is in dynamic equilibrium with the rate of folding, suggesting coordinated translation of collagen α1(I) and α2(I) polypeptides. Collagen α1(I) mRNA has in the 3' untranslated region (UTR) a C-rich sequence that binds protein αCP, this binding stabilizes the mRNA in collagen producing cells. In the 5' UTR both collagen mRNAs have a conserved stem-loop (5' SL) structure. The 5' SL is critical for high collagen expression, knock in mice with disruption of the 5' SL are resistant to liver fibrosis. the 5' SL binds protein LARP6 with strict sequence specificity and high affinity. LARP6 recruits RNA helicase A to facilitate translation initiation and associates collagen mRNAs with vimentin and nonmuscle myosin filaments. Binding to vimentin stabilizes collagen mRNAs, while nonmuscle myosin regulates coordinated translation of α1(I) and α2(I) mRNAs. When nonmuscle myosin filaments are disrupted the cells secrete only α1 homotrimers. Thus, the mechanism governing high collagen expression involves two RNA binding proteins and development of cytoskeletal filaments. Copyright © 2013 John Wiley & Sons, Ltd.

  9. This Constitution: A Bicentennial Chronicle.

    ERIC Educational Resources Information Center

    This Constitution, 1986

    1986-01-01

    Providing a link between constitutional scholars and the planners of school and public programs observing the Bicentennial of the United States Constitution, this series of the Bicentennial Chronicles features articles that provide a link between scholars of the Constitution and the people who will be planning programs for the public and for the…

  10. Preubiquitinated chimeric ErbB2 is constitutively endocytosed and subsequently degraded in lysosomes

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Vuong, Tram Thu; Berger, Christian; Bertelsen, Vibeke

    The oncoprotein ErbB2 is endocytosis-deficient, probably due to its interaction with Heat shock protein 90. We previously demonstrated that clathrin-dependent endocytosis of ErbB2 is induced upon incubation of cells with Ansamycin derivatives, such as geldanamycin and its derivative 17-AAG. Furthermore, we have previously demonstrated that a preubiquitinated chimeric EGFR (EGFR-Ub{sub 4}) is constitutively endocytosed in a clathrin-dependent manner. We now demonstrate that also an ErbB2-Ub{sub 4} chimera is endocytosed constitutively and clathrin-dependently. Upon expression, the ErbB2-Ub{sub 4} was further ubiquitinated, and by Western blotting, we demonstrated the formation of both Lys48-linked and Lys63-linked polyubiquitin chains. ErbB2-Ub{sub 4} was constitutively internalizedmore » and eventually sorted to late endosomes and lysosomes where the fusion protein was degraded. ErbB2-Ub{sub 4} was not cleaved prior to internalization. Interestingly, over-expression of Ubiquitin Interaction Motif-containing dominant negative fragments of the clathrin adaptor proteins epsin1 and Eps15 negatively affected endocytosis of ErbB2. Altogether, this argues that ubiquitination is sufficient to induce clathrin-mediated endocytosis and lysosomal degradation of the otherwise plasma membrane localized ErbB2. Also, it appears that C-terminal cleavage is not required for endocytosis. -- Highlights: ► A chimera containing ErbB2 and a tetra-Ubiquitin chain internalizes constitutively. ► Receptor fragmentation is not required for endocytosis of ErbB2. ► Ubiquitination is sufficient to induce endocytosis and degradation of ErbB2. ► ErbB2-Ub4 is internalized clathrin-dependently.« less

  11. Heat-shock protein 70 (Hsp70) expression in four limpets of the genus Lottia: interspecific variation in constitutive and inducible synthesis correlates with in situ exposure to heat stress.

    PubMed

    Dong, Yunwei; Miller, Luke P; Sanders, Jon G; Somero, George N

    2008-10-01

    Limpets of the genus Lottia occupy a broad vertical distribution on wave-exposed rocky shores, a range that encompasses gradients in the frequency and severity of thermal and desiccation stress brought on by aerial emersion. Using western blot analysis of levels of heat-shock protein 70 (Hsp70), we examined the heat-shock responses of four Lottia congeners: Lottia scabra and L. austrodigitalis, which occur in the high-intertidal zone, and L. pelta and L. scutum, which are restricted to the low- and mid-intertidal zones. Our results suggest distinct strategies of Hsp70 expression in limpets occupying different heights and orientations in the rocky intertidal zone. In freshly field-collected animals and in specimens acclimated at ambient temperature ( approximately 14 degrees C) for 14 days, the two high-intertidal species had higher constitutive levels of Hsp70 than the low- and mid-intertidal species. During aerial exposure to high temperatures, the two low-shore species and L. austrodigitalis exhibited an onset of Hsp70 expression at 28 degrees C; no induction of Hsp70 occurred in L. scabra. Our findings suggest that high-intertidal congeners of Lottia employ a "preparative defense" strategy involving maintenance of high constitutive levels of Hsp70 in their cells as a mechanism for protection against periods of extreme and unpredictable heat stress.

  12. Protein Kinase A Regulates Constitutive Expression of Small Heat-Shock Genes in an Msn2/4p-Independent and Hsf1p-Dependent Manner in Saccharomyces cerevisiae

    PubMed Central

    Ferguson, Scott B.; Anderson, Erik S.; Harshaw, Robyn B.; Thate, Tim; Craig, Nancy L.; Nelson, Hillary C. M.

    2005-01-01

    Hsf1p, the heat-shock transcription factor from Saccharomyces cerevisiae, has a low level of constitutive transcriptional activity and is kept in this state through negative regulation. In an effort to understand this negative regulation, we developed a novel genetic selection that detects altered expression from the HSP26 promoter. Using this reporter strain, we identified mutations and dosage compensators in the Ras/cAMP signaling pathway that decrease cAMP levels and increase expression from the HSP26 promoter. In yeast, low cAMP levels reduce the catalytic activity of the cAMP-dependent kinase PKA. Previous studies had proposed that the stress response transcription factors Msn2p/4p, but not Hsf1p, are repressed by PKA. However, we found that reduction or elimination of PKA activity strongly derepresses transcription of the small heat-shock genes HSP26 and HSP12, even in the absence of MSN2/4. In a strain deleted for MSN2/4 and the PKA catalytic subunits, expression of HSP12 and HSP26 depends on HSF1 expression. Our findings indicate that Hsf1p functions downstream of PKA and suggest that PKA might be involved in negative regulation of Hsf1p activity. These results represent a major change in our understanding of how PKA signaling influences the heat-shock response and heat-shock protein expression. PMID:15545649

  13. Distribution and sequence homogeneity of an abundant satellite DNA in the beetle, Tenebrio molitor.

    PubMed Central

    Davis, C A; Wyatt, G R

    1989-01-01

    The mealworm beetle, Tenebrio molitor, contains an unusually abundant and homogeneous satellite DNA which constitutes up to 60% of its genome. The satellite DNA is shown to be present in all of the chromosomes by in situ hybridization. 18 dimers of the repeat unit were cloned and sequenced. The consensus sequence is 142 nt long and lacks any internal repeat structure. Monomers of the sequence are very similar, showing on average a 2% divergence from the calculated consensus. Variant nucleotides are scattered randomly throughout the sequence although some variants are more common than others. Neighboring repeat units are no more alike than randomly chosen ones. The results suggest that some mechanism, perhaps gene conversion, is acting to maintain the homogeneity of the satellite DNA despite its abundance and distribution on all of the chromosomes. Images PMID:2762148

  14. Megakaryocytic differentiation induced by constitutive activation of mitogen-activated protein kinase kinase.

    PubMed Central

    Whalen, A M; Galasinski, S C; Shapiro, P S; Nahreini, T S; Ahn, N G

    1997-01-01

    The K562 erythroleukemia cell line was used to study the molecular mechanisms regulating lineage commitment of hematopoietic stem cells. Phorbol esters, which initiate megakaryocyte differentiation in this cell line, caused a rapid increase in extracellular-signal-regulated kinase (ERK), which remained elevated for 2 h and returned to near-basal levels by 24 h. In the absence of extracellular stimuli, ERK could be activated by expression of constitutively active mutants of mitogen-activated protein (MAP) kinase kinase (MKK), resulting in cell adhesion and spreading, increased cell size, inhibition of cell growth, and induction of the platelet-specific integrin alphaIIb beta3, all hallmarks of megakaryocytic differentiation. In contrast, expression of wild-type MKK had little effect. In addition, constitutively active MKK suppressed the expression of an erythroid marker, alpha-globin, indicating the ability to suppress cellular responses necessary for alternative cell lineages. The MKK inhibitor PD98059 blocked MKK/ERK activation and cellular responses to phorbol ester, demonstrating that activation of MKK is necessary and sufficient to induce a differentiation program along the megakaryocyte lineage. Thus, the MAP kinase cascade, which promotes cell growth and proliferation in many cell types, instead inhibits cell proliferation and initiates lineage-specific differentiation in K562 cells, establishing a model system to investigate the mechanisms by which this signal transduction pathway specifies cell fate and developmental processes. PMID:9121442

  15. Megakaryocytic differentiation induced by constitutive activation of mitogen-activated protein kinase kinase.

    PubMed

    Whalen, A M; Galasinski, S C; Shapiro, P S; Nahreini, T S; Ahn, N G

    1997-04-01

    The K562 erythroleukemia cell line was used to study the molecular mechanisms regulating lineage commitment of hematopoietic stem cells. Phorbol esters, which initiate megakaryocyte differentiation in this cell line, caused a rapid increase in extracellular-signal-regulated kinase (ERK), which remained elevated for 2 h and returned to near-basal levels by 24 h. In the absence of extracellular stimuli, ERK could be activated by expression of constitutively active mutants of mitogen-activated protein (MAP) kinase kinase (MKK), resulting in cell adhesion and spreading, increased cell size, inhibition of cell growth, and induction of the platelet-specific integrin alphaIIb beta3, all hallmarks of megakaryocytic differentiation. In contrast, expression of wild-type MKK had little effect. In addition, constitutively active MKK suppressed the expression of an erythroid marker, alpha-globin, indicating the ability to suppress cellular responses necessary for alternative cell lineages. The MKK inhibitor PD98059 blocked MKK/ERK activation and cellular responses to phorbol ester, demonstrating that activation of MKK is necessary and sufficient to induce a differentiation program along the megakaryocyte lineage. Thus, the MAP kinase cascade, which promotes cell growth and proliferation in many cell types, instead inhibits cell proliferation and initiates lineage-specific differentiation in K562 cells, establishing a model system to investigate the mechanisms by which this signal transduction pathway specifies cell fate and developmental processes.

  16. Constitutive expression of a grapevine polygalacturonase-inhibiting protein affects gene expression and cell wall properties in uninfected tobacco

    PubMed Central

    2011-01-01

    Background Polygalacturonase-inhibiting proteins (PGIPs) directly limit the effective ingress of fungal pathogens by inhibiting cell wall-degrading endopolygalacturonases (ePGs). Transgenic tobacco plants over-expressing grapevine (Vitis vinifera) Vvpgip1 have previously been shown to be resistant to Botrytis infection. In this study we characterized two of these PGIP over-expressing lines with known resistance phenotypes by gene expression and hormone profiling in the absence of pathogen infection. Results Global gene expression was performed by a cross-species microarray approach using a potato cDNA microarray. The degree of potential cross-hybridization between probes was modeled by a novel computational workflow designed in-house. Probe annotations were updated by predicting probe-to-transcript hybridizations and combining information derived from other plant species. Comparing uninfected Vvpgip1-overexpressing lines to wild-type (WT), 318 probes showed significant change in expression. Functional groups of genes involved in metabolism and associated to the cell wall were identified and consequent cell wall analysis revealed increased lignin-levels in the transgenic lines, but no major differences in cell wall-derived polysaccharides. GO enrichment analysis also identified genes responsive to auxin, which was supported by elevated indole-acetic acid (IAA) levels in the transgenic lines. Finally, a down-regulation of xyloglucan endotransglycosylase/hydrolases (XTHs), which are important in cell wall remodeling, was linked to a decrease in total XTH activity. Conclusions This evaluation of PGIP over-expressing plants performed under pathogen-free conditions to exclude the classical PGIP-ePG inhibition interaction indicates additional roles for PGIPs beyond the inhibition of ePGs. PMID:22078230

  17. Molecular cloning, characterization and expression profiles of thioredoxin 1 and thioredoxin 2 genes in Mytilus galloprovincialis

    NASA Astrophysics Data System (ADS)

    Wang, Qing; Ning, Xuanxuan; Pei, Dong; Zhao, Jianmin; You, Liping; Wang, Chunyan; Wu, Huifeng

    2013-05-01

    Thioredoxin (Trx) proteins are involved in many biological processes especially the regulation of cellular redox homeostasis. In this study, two Trx cDNAs were cloned from the mussel Mytilus galloprovincialis using rapid amplifi cation of cDNA ends-polymerase chain reaction (RACE-PCR). The two cDNAs were named MgTrx1 and MgTrx2, respectively. The open reading frames of MgTrx1 and MgTrx2 were 318 and 507 base pairs (bp) and they encoded proteins of 105 and 168 amino acids with estimated molecular masses of 11.45 and 18.93 kDa, respectively. Sequence analysis revealed that both proteins possessed the conserved active site dithiol motif Cys-Gly-Pro-Cys. In addition, MgTrx2 also possessed a putative mitochondrial targeting signal suggesting that it is located in the mitochondria. Quantitative real-time polymerase chain reaction (qPCR) revealed that both MgTrx1 and MgTrx2 were constitutively expressed in all tissues examined. The MgTrx1 transcript was most abundant in hemocytes and gills, whereas the MgTrx2 transcript was most abundant in gonad, hepatopancreas, gill and hemocytes. Following Vibrio anguillarum challenge, the expression of MgTrx1 was up-regulated and reached its peak, at a value 10-fold the initial value, at 24 h. Subsequently, expression returned back to the original level. In contrast, the expression level of MgTrx2 was down-regulated following bacterial stimulation, with one fi fth of the control level evident at 12 h post challenge. These results suggest that MgTrx1 and MgTrx2 may play important roles in the response of M. galloprovincialis to bacterial challenge.

  18. Prevalence of MLH1 constitutional epimutations as a cause of Lynch syndrome in unselected versus selected consecutive series of patients with colorectal cancer.

    PubMed

    Castillejo, Adela; Hernández-Illán, Eva; Rodriguez-Soler, María; Pérez-Carbonell, Lucía; Egoavil, Cecilia; Barberá, Victor M; Castillejo, María-Isabel; Guarinos, Carla; Martínez-de-Dueñas, Eduardo; Juan, María-Jose; Sánchez-Heras, Ana-Beatriz; García-Casado, Zaida; Ruiz-Ponte, Clara; Brea-Fernández, Alejandro; Juárez, Miriam; Bujanda, Luis; Clofent, Juan; Llor, Xavier; Andreu, Montserrat; Castells, Antoni; Carracedo, Angel; Alenda, Cristina; Payá, Artemio; Jover, Rodrigo; Soto, José-Luis

    2015-07-01

    The prevalence of MLH1 constitutional epimutations in the general population is unknown. We sought to analyse the prevalence of MLH1 constitutional epimutations in unselected and selected series of patients with colorectal cancer (CRC). Patients with diagnoses of CRC (n=2123) were included in the unselected group. For comparison, a group of 847 selected patients with CRC who fulfilled the revised Bethesda guidelines (rBG) were also included. Somatic and constitutional MLH1 methylation was assayed via methylation-specific multiplex ligation-dependent probe amplification of cases lacking MLH1 expression. Germline alterations in mismatch-repair (MMR) genes were assessed via Sanger sequencing and methylation-specific multiplex ligation-dependent probe amplification. Loss of MLH1 expression occurred in 5.5% of the unselected series and 12.5% of the selected series (p<0.0001). No constitutional epimutations in MLH1 were detected in the unselected population (0/62); five cases from the selected series were positive for MLH1 epimutations (15.6%, 5/32; p=0.004). Our results suggest a negligible prevalence of MLH1 constitutional epimutations in unselected cases of CRC. Therefore, MLH1 constitutional epimutation analysis should be conducted only for patients who fulfil the rBG and who lack MLH1 expression with methylated MLH1. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

  19. Space-based measurements of elemental abundances and their relation to solar abundances

    NASA Technical Reports Server (NTRS)

    Coplan, M. A.; Ogilvie, K. W.; Bochsler, P.; Geiss, J.

    1990-01-01

    The Ion Composition Instrument (ICI) aboard the ISEE-3/ICE spacecraft was in the solar wind continuously from August 1978 to December 1982. The results made it possible to establish long-term average solar wind abundance values for helium, oxygen, neon, silicon, and iron. The Charge-Energy-Mass instrument aboard the CCE spacecraft of the AMPTE mission has measured the abundance of these elements in the magnetosheath and has also added carbon, nitrogen, magnesium, and sulfur to the list. There is strong evidence that these magnetosheath abundances are representative of the solar wind. Other sources of solar wind abundances are Solar Energetic Particle experiments and Apollo lunar foils. When comparing the abundances from all of these sources with photospheric abundances, it is clear that helium is depleted in the solar wind while silicon and iron are enhanced. Solar wind abundances for carbon, nitrogen, oxygen, and neon correlate well with the photospheric values. The incorporation of minor ions into the solar wind appears to depend upon both the ionization times for the elements and the Coulomb drag exerted by the outflowing proton flux.

  20. A geometric rationale for invariance, covariance and constitutive relations

    NASA Astrophysics Data System (ADS)

    Romano, Giovanni; Barretta, Raffaele; Diaco, Marina

    2018-01-01

    There are, in each branch of science, statements which, expressed in ambiguous or even incorrect but seemingly friendly manner, were repeated for a long time and eventually became diffusely accepted. Objectivity of physical fields and of their time rates and frame indifference of constitutive relations are among such notions. A geometric reflection on the description of frame changes as spacetime automorphisms, on induced push-pull transformations and on proper physico-mathematical definitions of material, spatial and spacetime tensor fields and of their time-derivatives along the motion, is here carried out with the aim of pointing out essential notions and of unveiling false claims. Theoretical and computational aspects of nonlinear continuum mechanics, and especially those pertaining to constitutive relations, involving material fields and their time rates, gain decisive conceptual and operative improvement from a proper geometric treatment. Outcomes of the geometric analysis are frame covariance of spacetime velocity, material stretching and material spin. A univocal and frame-covariant tool for evaluation of time rates of material fields is provided by the Lie derivative along the motion. The postulate of frame covariance of material fields is assessed to be a natural physical requirement which cannot interfere with the formulation of constitutive laws, with claims of the contrary stemming from an improper imposition of equality in place of equivalence.

  1. Constitutive expression of fluorescent protein by Aspergillus var. niger and Aspergillus carbonarius to monitor fungal colonization in maize plants.

    PubMed

    Palencia, Edwin Rene; Glenn, Anthony Elbie; Hinton, Dorothy Mae; Bacon, Charles Wilson

    2013-09-01

    Aspergillus niger and Aspergillus carbonarius are two species in the Aspergillus section Nigri (black-spored aspergilli) frequently associated with peanut (Arachis hypogea), maize (Zea mays), and other plants as pathogens. These infections are symptomless and as such are major concerns since some black aspergilli produce important mycotoxins, ochratoxins A, and the fumonisins. To facilitate the study of the black aspergilli-maize interactions with maize during the early stages of infections, we developed a method that used the enhanced yellow fluorescent protein (eYFP) and the monomeric red fluorescent protein (mRFP1) to transform A. niger and A. carbonarius, respectively. The results were constitutive expressions of the fluorescent genes that were stable in the cytoplasms of hyphae and conidia under natural environmental conditions. The hyphal in planta distribution in 21-day-old seedlings of maize were similar wild type and transformants of A. niger and A. carbonarius. The in planta studies indicated that both wild type and transformants internally colonized leaf, stem and root tissues of maize seedlings, without any visible disease symptoms. Yellow and red fluorescent strains were capable of invading epidermal cells of maize roots intercellularly within the first 3 days after inoculation, but intracellular hyphal growth was more evident after 7 days of inoculation. We also tested the capacity of fluorescent transformants to produce ochratoxin A and the results with A. carbonarius showed that this transgenic strain produced similar concentrations of this secondary metabolite. This is the first report on the in planta expression of fluorescent proteins that should be useful to study the internal plant colonization patterns of two ochratoxigenic species in the Aspergillus section Nigri. © 2013.

  2. Global Survey of National Constitutions: Mapping Constitutional Commitments to Sexual and Reproductive Health and Rights.

    PubMed

    Pizzarossa, Lucía Berro; Perehudoff, Katrina

    2017-12-01

    General Comment No. 22, issued in 2016 by the Committee on Economic, Social and Cultural Rights (CESCR), clarifies states' legal duties to respect, protect, and fulfill the right to sexual and reproductive health (SRH). Our study analyzes domestic constitutions around the world to investigate whether and to what extent the right to sexual and reproductive health is respected, protected, and fulfilled; to what extent these provisions are inclusive and non-discriminatory; and to what degree the interlinkages between this and other human rights are acknowledged. Of the 195 constitutions accessed, 27 enshrine sexual and/or reproductive health, and seven adopt restrictive approaches to this right. In the 27 constitutions, provisions most frequently enshrine respect of one's sexual health and family planning decisions, the protection of sexual health, and the provision of reproductive health care and family planning services (fulfillment). Most of the 27 constitutions fail to adequately respect reproductive health rights; to protect reproductive health, family planning, and abortion services from third-party interference; and to fulfill all dimensions of sexual health and access to abortion. Three of the 27 constitutions enshrine a universal right to SRH, and additional constitutions protect specific vulnerable groups (such as women, children) and/or restrict the scope of rights holders to couples. Among the 27 constitutions, nine explicitly link the right to sexual and reproductive health to the rights to education, science, and/or to make autonomous decisions about sexuality and reproduction. Our results can serve as a baseline measure to track constitutional reforms in pursuit of the realization of sexual and reproductive health and rights, and as building blocks for future lawmakers committed to realizing these rights through domestic legal reform.

  3. APOGEE Chemical Abundances of the Sagittarius Dwarf Galaxy

    NASA Astrophysics Data System (ADS)

    Hasselquist, Sten; Shetrone, Matthew; Smith, Verne; Holtzman, Jon; McWilliam, Andrew; Fernández-Trincado, J. G.; Beers, Timothy C.; Majewski, Steven R.; Nidever, David L.; Tang, Baitian; Tissera, Patricia B.; Fernández Alvar, Emma; Allende Prieto, Carlos; Almeida, Andres; Anguiano, Borja; Battaglia, Giuseppina; Carigi, Leticia; Delgado Inglada, Gloria; Frinchaboy, Peter; García-Hernández, D. A.; Geisler, Doug; Minniti, Dante; Placco, Vinicius M.; Schultheis, Mathias; Sobeck, Jennifer; Villanova, Sandro

    2017-08-01

    The Apache Point Observatory Galactic Evolution Experiment provides the opportunity of measuring elemental abundances for C, N, O, Na, Mg, Al, Si, P, K, Ca, V, Cr, Mn, Fe, Co, and Ni in vast numbers of stars. We analyze thechemical-abundance patterns of these elements for 158 red giant stars belonging to the Sagittarius dwarf galaxy (Sgr). This is the largest sample of Sgr stars with detailed chemical abundances, and it is the first time that C, N, P, K, V, Cr, Co, and Ni have been studied at high resolution in this galaxy. We find that the Sgr stars with [Fe/H] ≳ -0.8 are deficient in all elemental abundance ratios (expressed as [X/Fe]) relative to the Milky Way, suggesting that the Sgr stars observed today were formed from gas that was less enriched by Type II SNe than stars formed in the Milky Way. By examining the relative deficiencies of the hydrostatic (O, Na, Mg, and Al) and explosive (Si, P, K, and Mn) elements, our analysis supports the argument that previous generations of Sgr stars were formed with a top-light initial mass function, one lacking the most massive stars that would normally pollute the interstellar medium with the hydrostatic elements. We use a simple chemical-evolution model, flexCE, to further support our claim and conclude that recent stellar generations of Fornax and the Large Magellanic Cloud could also have formed according to a top-light initial mass function.

  4. Global Survey of National Constitutions

    PubMed Central

    Perehudoff, Katrina

    2017-01-01

    Abstract General Comment No. 22, issued in 2016 by the Committee on Economic, Social and Cultural Rights (CESCR), clarifies states’ legal duties to respect, protect, and fulfill the right to sexual and reproductive health (SRH). Our study analyzes domestic constitutions around the world to investigate whether and to what extent the right to sexual and reproductive health is respected, protected, and fulfilled; to what extent these provisions are inclusive and non-discriminatory; and to what degree the interlinkages between this and other human rights are acknowledged. Of the 195 constitutions accessed, 27 enshrine sexual and/or reproductive health, and seven adopt restrictive approaches to this right. In the 27 constitutions, provisions most frequently enshrine respect of one’s sexual health and family planning decisions, the protection of sexual health, and the provision of reproductive health care and family planning services (fulfillment). Most of the 27 constitutions fail to adequately respect reproductive health rights; to protect reproductive health, family planning, and abortion services from third-party interference; and to fulfill all dimensions of sexual health and access to abortion. Three of the 27 constitutions enshrine a universal right to SRH, and additional constitutions protect specific vulnerable groups (such as women, children) and/or restrict the scope of rights holders to couples. Among the 27 constitutions, nine explicitly link the right to sexual and reproductive health to the rights to education, science, and/or to make autonomous decisions about sexuality and reproduction. Our results can serve as a baseline measure to track constitutional reforms in pursuit of the realization of sexual and reproductive health and rights, and as building blocks for future lawmakers committed to realizing these rights through domestic legal reform. PMID:29302182

  5. Teaching About the Constitution.

    ERIC Educational Resources Information Center

    White, Charles S.

    1988-01-01

    Reviews "The U.S. Constitution Then and Now," a two-unit program using the integrated database and word processing capabilities of AppleWorks. For grades 7-12, the units simulate the constitutional convention and the principles of free speech and privacy. Concludes that with adequate time, the program can provide a potentially powerful…

  6. The U.S. Constitution in Today's World.

    ERIC Educational Resources Information Center

    Patrick, John J.

    A comparative study of constitutions and governments in world history is a key to deeper understanding of the U.S. Constitution. While many countries have constitutions, the United States is among a minority of nations in today's world that has a constitutional government. Many nations' constitutions truly guarantee few protections of individual…

  7. CFTR expression and organ damage in cystic fibrosis

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tizzano, E.; Chitayat, D.; Buchwald, M.

    1994-09-01

    To assist our understanding of the origin of organ damage caused by cystic fibrosis (CF) disease, we have analyzed the pattern of expression of the CF gene (CFTR). mRNA in situ hybridization analysis was carried out in human fetal, newborn, infant and adult tissues and the abundance of the mRNA was correlated with the known pathology at the various stages of human development. Analysis of the pattern of expression indicates a constitutive level of mRNA in gastrointestinal tissues starting during early development and maintained throughout life. Prenatal respiratory tissues show qualitative and quantitative major differences in comparison to postnatal lungmore » samples. Male reproductive tissues show high levels of expression in the head of the epididymis compared with the rest of the male ducts. Female reproductive tissues show a variable pattern of expression at different stages during fetal development and during puberty probably due to changes in hormonal levels. Gastrointestinal and male reproductive tissues have a consistent pathology at birth, whereas no lung abnormalities have been described in newborns affected by CF. Our results show that there is no exact correlations between organ damage present at birth and the degree of CFTR expression. To explain these observations, we hypothesize that the pathogenesis of organ damage in CF depend on at least three factors: the rate of CFTR-mediated fluid secretion, differences in genotype and environmental factors, such as the amount of macromolecules in the lumen of the ducts. This last element predicts that damage will occur in tissues with high protein loads and low flow rates (e.g. pancreas, epididymis), where the absence of CFTR function leads to obstruction and pathology. Organs that express CFTR but with no significant damage (e.g. prenatal lung, female reproductive tissues), will have a low protein load and a high flow rates.« less

  8. Light differentially regulates cell division and the mRNA abundance of pea nucleolin during de-etiolation

    NASA Technical Reports Server (NTRS)

    Reichler, S. A.; Balk, J.; Brown, M. E.; Woodruff, K.; Clark, G. B.; Roux, S. J.

    2001-01-01

    The abundance of plant nucleolin mRNA is regulated during de-etiolation by phytochrome. A close correlation between the mRNA abundance of nucleolin and mitosis has also been previously reported. These results raised the question of whether the effects of light on nucleolin mRNA expression were a consequence of light effects on mitosis. To test this we compared the kinetics of light-mediated increases in cell proliferation with that of light-mediated changes in the abundance of nucleolin mRNA using plumules of dark-grown pea (Pisum sativum) seedlings. These experiments show that S-phase increases 9 h after a red light pulse, followed by M-phase increases in the plumule leaves at 12 h post-irradiation, a time course consistent with separately measured kinetics of red light-induced increases in the expression of cell cycle-regulated genes. These increases in cell cycle-regulated genes are photoreversible, implying that the light-induced increases in cell proliferation are, like nucleolin mRNA expression, regulated via phytochrome. Red light stimulates increases in the mRNA for nucleolin at 6 h post-irradiation, prior to any cell proliferation changes and concurrent with the reported timing of phytochrome-mediated increases of rRNA abundance. After a green light pulse, nucleolin mRNA levels increase without increasing S-phase or M-phase. Studies in animals and yeast indicate that nucleolin plays a significant role in ribosome biosynthesis. Consistent with this function, pea nucleolin can rescue nucleolin deletion mutants of yeast that are defective in rRNA synthesis. Our data show that during de-etiolation, the increased expression of nucleolin mRNA is more directly regulated by light than by mitosis.

  9. Expression of the Arabidopsis thaliana BBX32 gene in soybean increases grain yield.

    PubMed

    Preuss, Sasha B; Meister, Robert; Xu, Qingzhang; Urwin, Carl P; Tripodi, Federico A; Screen, Steven E; Anil, Veena S; Zhu, Shuquan; Morrell, James A; Liu, Grace; Ratcliffe, Oliver J; Reuber, T Lynne; Khanna, Rajnish; Goldman, Barry S; Bell, Erin; Ziegler, Todd E; McClerren, Amanda L; Ruff, Thomas G; Petracek, Marie E

    2012-01-01

    Crop yield is a highly complex quantitative trait. Historically, successful breeding for improved grain yield has led to crop plants with improved source capacity, altered plant architecture, and increased resistance to abiotic and biotic stresses. To date, transgenic approaches towards improving crop grain yield have primarily focused on protecting plants from herbicide, insects, or disease. In contrast, we have focused on identifying genes that, when expressed in soybean, improve the intrinsic ability of the plant to yield more. Through the large scale screening of candidate genes in transgenic soybean, we identified an Arabidopsis thaliana B-box domain gene (AtBBX32) that significantly increases soybean grain yield year after year in multiple transgenic events in multi-location field trials. In order to understand the underlying physiological changes that are associated with increased yield in transgenic soybean, we examined phenotypic differences in two AtBBX32-expressing lines and found increases in plant height and node, flower, pod, and seed number. We propose that these phenotypic changes are likely the result of changes in the timing of reproductive development in transgenic soybean that lead to the increased duration of the pod and seed development period. Consistent with the role of BBX32 in A. thaliana in regulating light signaling, we show that the constitutive expression of AtBBX32 in soybean alters the abundance of a subset of gene transcripts in the early morning hours. In particular, AtBBX32 alters transcript levels of the soybean clock genes GmTOC1 and LHY-CCA1-like2 (GmLCL2). We propose that through the expression of AtBBX32 and modulation of the abundance of circadian clock genes during the transition from dark to light, the timing of critical phases of reproductive development are altered. These findings demonstrate a specific role for AtBBX32 in modulating soybean development, and demonstrate the validity of expressing single genes in crops to

  10. Immunohistochemical localization of constitutive and inducible cyclo-oxygenases in rat uterus during the oestrous cycle and pregnancy.

    PubMed

    Fang, L; Chatterjee, S; Dong, Y L; Gangula, P R; Yallampalli, C

    1998-06-01

    The uterus is a rich source of eicosanoids synthesized from arachidonic acid metabolism through the cyclo-oxygenase pathway. Two isoforms of cyclo-oxygenase, constitutive (COX-I) and inducible (COX-II) enzyme, have been reported. In the present study, we have immunohistochemically mapped the distribution of both COX-I and COX-II during various physiological states of the rat uterus. Uterine tissue was collected from female rats (a) during different stages of the oestrous cycle, (b) on days 1, 4, 8 and 18 of gestation, (c) after spontaneous delivery and (d) post partum, and fixed in Bouin's fixative. After paraffin wax embedding, 5-microm-thick sections were immunohistochemically stained by the ABC technique. Observation of the stained sections under the light microscope revealed that, in non-pregnant rat uterus, both COX-I and COX-II were abundantly expressed in the endometrium, with minimal staining observed in the myometrium. Staining was more prominent in epithelial cells than in stromal cells. The intensity of staining in epithelial cells was highest at pro-oestrus and oestrus and lowest at dioestrus. In pregnant rats, although the expression of both COX-I and COX-II was localized primarily to the endometrium with very little staining in the myometrium on day 1 of gestation, both of these enzymes were also apparent in myometrial cells by day 4 of gestation. The staining intensity of endometrial and myometrial cells increased further with the progression of gestation, being maximal at the time of spontaneous delivery. During the post-partum period, however, the staining intensity for both of the enzymes in endometrium and myometrium was decreased. Thus, our studies show that the expression of cyclo-oxygenases in various uterine cells vary with the oestrous cycle and with pregnancy. Furthermore, prominent increases in the expression of cyclo-oxygenases in the myometrium during pregnancy and parturition imply that the cyclo-oxygenase system in the myometrium may

  11. Biaxial experiments supporting the development of constitutive theories for advanced high-temperature materials

    NASA Technical Reports Server (NTRS)

    Ellis, J. R.

    1988-01-01

    Complex states of stress and strain are introduced into components during service in engineering applications. It follows that analysis of such components requires material descriptions, or constitutive theories, which reflect the tensorial nature of stress and strain. For applications involving stress levels above yield, the situation is more complex in that material response is both nonlinear and history dependent. This has led to the development of viscoplastic constitutive theories which introduce time by expressing the flow and evolutionary equation in the form of time derivatives. Models were developed here which can be used to analyze high temperature components manufactured from advanced composite materials. In parallel with these studies, effort was directed at developing multiaxial testing techniques to verify the various theories. Recent progress in the development of constitutive theories from both the theoretical and experimental viewpoints are outlined. One important aspect is that material descriptions for advanced composite materials which can be implemented in general purpose finite element codes and used for practical design are verified.

  12. Identification of chimeric antigen receptors that mediate constitutive or inducible proliferation of T cells.

    PubMed

    Frigault, Matthew J; Lee, Jihyun; Basil, Maria Ciocca; Carpenito, Carmine; Motohashi, Shinichiro; Scholler, John; Kawalekar, Omkar U; Guedan, Sonia; McGettigan, Shannon E; Posey, Avery D; Ang, Sonny; Cooper, Laurence J N; Platt, Jesse M; Johnson, F Brad; Paulos, Chrystal M; Zhao, Yangbing; Kalos, Michael; Milone, Michael C; June, Carl H

    2015-04-01

    This study compared second-generation chimeric antigen receptors (CAR) encoding signaling domains composed of CD28, ICOS, and 4-1BB (TNFRSF9). Here, we report that certain CARs endow T cells with the ability to undergo long-term autonomous proliferation. Transduction of primary human T cells with lentiviral vectors encoding some of the CARs resulted in sustained proliferation for up to 3 months following a single stimulation through the T-cell receptor (TCR). Sustained numeric expansion was independent of cognate antigen and did not require the addition of exogenous cytokines or feeder cells after a single stimulation of the TCR and CD28. Results from gene array and functional assays linked sustained cytokine secretion and expression of T-bet (TBX21), EOMES, and GATA-3 to the effect. Sustained expression of the endogenous IL2 locus has not been reported in primary T cells. Sustained proliferation was dependent on CAR structure and high expression, the latter of which was necessary but not sufficient. The mechanism involves constitutive signaling through NF-κB, AKT, ERK, and NFAT. The propagated CAR T cells retained a diverse TCR repertoire, and cellular transformation was not observed. The CARs with a constitutive growth phenotype displayed inferior antitumor effects and engraftment in vivo. Therefore, the design of CARs that have a nonconstitutive growth phenotype may be a strategy to improve efficacy and engraftment of CAR T cells. The identification of CARs that confer constitutive or nonconstitutive growth patterns may explain observations that CAR T cells have differential survival patterns in clinical trials. ©2015 American Association for Cancer Research.

  13. Activation of the Constitutive Androstane Receptor induces hepatic lipogenesis and regulates Pnpla3 gene expression in a LXR-independent way.

    PubMed

    Marmugi, Alice; Lukowicz, Céline; Lasserre, Frederic; Montagner, Alexandra; Polizzi, Arnaud; Ducheix, Simon; Goron, Adeline; Gamet-Payrastre, Laurence; Gerbal-Chaloin, Sabine; Pascussi, Jean Marc; Moldes, Marthe; Pineau, Thierry; Guillou, Hervé; Mselli-Lakhal, Laila

    2016-07-15

    The Constitutive Androstane Receptor (CAR, NR1I3) has been newly described as a regulator of energy metabolism. A relevant number of studies using animal models of obesity suggest that CAR activation could be beneficial on the metabolic balance. However, this remains controversial and the underlying mechanisms are still unknown. This work aimed to investigate the effect of CAR activation on hepatic energy metabolism during physiological conditions, i.e. in mouse models not subjected to metabolic/nutritional stress. Gene expression profiling in the liver of CAR knockout and control mice on chow diet and treated with a CAR agonist highlighted CAR-mediated up-regulations of lipogenic genes, concomitant with neutral lipid accumulation. A strong CAR-mediated up-regulation of the patatin-like phospholipase domain-containing protein 3 (Pnpla3) was demonstrated. Pnpla3 is a gene whose polymorphism is associated with the pathogenesis of nonalcoholic fatty liver disease (NAFLD) development. This observation was confirmed in human hepatocytes treated with the antiepileptic drug and CAR activator, phenobarbital and in immortalized human hepatocytes treated with CITCO. Studying the molecular mechanisms controlling Pnpla3 gene expression, we demonstrated that CAR does not act by a direct regulation of Pnpla3 transcription or via the Liver X Receptor but may rather involve the transcription factor Carbohydrate Responsive Element-binding protein. These data provide new insights into the regulation by CAR of glycolytic and lipogenic genes and on pathogenesis of steatosis. This also raises the question concerning the impact of drugs and environmental contaminants in lipid-associated metabolic diseases. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Computational Analysis and Low-Scale Constitutive Expression of Laccases Synthetic Genes GlLCC1 from Ganoderma lucidum and POXA 1B from Pleurotus ostreatus in Pichia pastoris

    PubMed Central

    Reyes-Guzmán, Edwin Alfredo; Poutou-Piñales, Raúl A.; Reyes-Montaño, Edgar Antonio; Pedroza-Rodríguez, Aura Marina; Rodríguez-Vázquez, Refugio; Cardozo-Bernal, Ángela M.

    2015-01-01

    Lacasses are multicopper oxidases that can catalyze aromatic and non-aromatic compounds concomitantly with reduction of molecular oxygen to water. Fungal laccases have generated a growing interest due to their biotechnological potential applications, such as lignocellulosic material delignification, biopulping and biobleaching, wastewater treatment, and transformation of toxic organic pollutants. In this work we selected fungal genes encoding for laccase enzymes GlLCC1 in Ganoderma lucidum and POXA 1B in Pleurotus ostreatus. These genes were optimized for codon use, GC content, and regions generating secondary structures. Laccase proposed computational models, and their interaction with ABTS [2, 2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)] substrate was evaluated by molecular docking. Synthetic genes were cloned under the control of Pichia pastoris glyceraldehyde-3-phosphate dehydrogenase (GAP) constitutive promoter. P. pastoris X-33 was transformed with pGAPZαA-LaccGluc-Stop and pGAPZαA-LaccPost-Stop constructs. Optimization reduced GC content by 47 and 49% for LaccGluc-Stop and LaccPost-Stop genes, respectively. A codon adaptation index of 0.84 was obtained for both genes. 3D structure analysis using SuperPose revealed LaccGluc-Stop is similar to the laccase crystallographic structure 1GYC of Trametes versicolor. Interaction analysis of the 3D models validated through ABTS, demonstrated higher substrate affinity for LaccPost-Stop, in agreement with our experimental results with enzymatic activities of 451.08 ± 6.46 UL-1 compared to activities of 0.13 ± 0.028 UL-1 for LaccGluc-Stop. This study demonstrated that G. lucidum GlLCC1 and P. ostreatus POXA 1B gene optimization resulted in constitutive gene expression under GAP promoter and α-factor leader in P. pastoris. These are important findings in light of recombinant enzyme expression system utility for environmentally friendly designed expression systems, because of the wide range of substrates

  15. Computational analysis and low-scale constitutive expression of laccases synthetic genes GlLCC1 from Ganoderma lucidum and POXA 1B from Pleurotus ostreatus in Pichia pastoris.

    PubMed

    Rivera-Hoyos, Claudia M; Morales-Álvarez, Edwin David; Poveda-Cuevas, Sergio Alejandro; Reyes-Guzmán, Edwin Alfredo; Poutou-Piñales, Raúl A; Reyes-Montaño, Edgar Antonio; Pedroza-Rodríguez, Aura Marina; Rodríguez-Vázquez, Refugio; Cardozo-Bernal, Ángela M

    2015-01-01

    Lacasses are multicopper oxidases that can catalyze aromatic and non-aromatic compounds concomitantly with reduction of molecular oxygen to water. Fungal laccases have generated a growing interest due to their biotechnological potential applications, such as lignocellulosic material delignification, biopulping and biobleaching, wastewater treatment, and transformation of toxic organic pollutants. In this work we selected fungal genes encoding for laccase enzymes GlLCC1 in Ganoderma lucidum and POXA 1B in Pleurotus ostreatus. These genes were optimized for codon use, GC content, and regions generating secondary structures. Laccase proposed computational models, and their interaction with ABTS [2, 2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)] substrate was evaluated by molecular docking. Synthetic genes were cloned under the control of Pichia pastoris glyceraldehyde-3-phosphate dehydrogenase (GAP) constitutive promoter. P. pastoris X-33 was transformed with pGAPZαA-LaccGluc-Stop and pGAPZαA-LaccPost-Stop constructs. Optimization reduced GC content by 47 and 49% for LaccGluc-Stop and LaccPost-Stop genes, respectively. A codon adaptation index of 0.84 was obtained for both genes. 3D structure analysis using SuperPose revealed LaccGluc-Stop is similar to the laccase crystallographic structure 1GYC of Trametes versicolor. Interaction analysis of the 3D models validated through ABTS, demonstrated higher substrate affinity for LaccPost-Stop, in agreement with our experimental results with enzymatic activities of 451.08 ± 6.46 UL-1 compared to activities of 0.13 ± 0.028 UL-1 for LaccGluc-Stop. This study demonstrated that G. lucidum GlLCC1 and P. ostreatus POXA 1B gene optimization resulted in constitutive gene expression under GAP promoter and α-factor leader in P. pastoris. These are important findings in light of recombinant enzyme expression system utility for environmentally friendly designed expression systems, because of the wide range of substrates

  16. Aldosterone modulates thiazide-sensitive sodium chloride cotransporter abundance via DUSP6-mediated ERK1/2 signaling pathway.

    PubMed

    Feng, Xiuyan; Zhang, Yiqian; Shao, Ningjun; Wang, Yanhui; Zhuang, Zhizhi; Wu, Ping; Lee, Matthew J; Liu, Yingli; Wang, Xiaonan; Zhuang, Jieqiu; Delpire, Eric; Gu, Dingying; Cai, Hui

    2015-05-15

    Thiazide-sensitive sodium chloride cotransporter (NCC) plays an important role in maintaining blood pressure. Aldosterone is known to modulate NCC abundance. Previous studies reported that dietary salts modulated NCC abundance through either WNK4 [with no lysine (k) kinase 4]-SPAK (Ste20-related proline alanine-rich kinase) or WNK4-extracellular signal-regulated kinase-1 and -2 (ERK1/2) signaling pathways. To exclude the influence of SPAK signaling pathway on the role of the aldosterone-mediated ERK1/2 pathway in NCC regulation, we investigated the effects of dietary salt changes and aldosterone on NCC abundance in SPAK knockout (KO) mice. We found that in SPAK KO mice low-salt diet significantly increased total NCC abundance while reducing ERK1/2 phosphorylation, whereas high-salt diet decreased total NCC while increasing ERK1/2 phosphorylation. Importantly, exogenous aldosterone administration increased total NCC abundance in SPAK KO mice while increasing DUSP6 expression, an ERK1/2-specific phosphatase, and led to decreasing ERK1/2 phosphorylation without changing the ratio of phospho-T53-NCC/total NCC. In mouse distal convoluted tubule (mDCT) cells, aldosterone increased DUSP6 expression while reducing ERK1/2 phosphorylation. DUSP6 Knockdown increased ERK1/2 phosphorylation while reducing total NCC expression. Inhibition of DUSP6 by (E)-2-benzylidene-3-(cyclohexylamino)-2,3-dihydro-1H-inden-1-one increased ERK1/2 phosphorylation and reversed the aldosterone-mediated increments of NCC partly by increasing NCC ubiquitination. Therefore, these data suggest that aldosterone modulates NCC abundance via altering NCC ubiquitination through a DUSP6-dependent ERK1/2 signal pathway in SPAK KO mice and part of the effects of dietary salt changes may be mediated by aldosterone in the DCTs.

  17. Aldosterone modulates thiazide-sensitive sodium chloride cotransporter abundance via DUSP6-mediated ERK1/2 signaling pathway

    PubMed Central

    Feng, Xiuyan; Zhang, Yiqian; Shao, Ningjun; Wang, Yanhui; Zhuang, Zhizhi; Wu, Ping; Lee, Matthew J.; Liu, Yingli; Wang, Xiaonan; Zhuang, Jieqiu; Delpire, Eric; Gu, Dingying

    2015-01-01

    Thiazide-sensitive sodium chloride cotransporter (NCC) plays an important role in maintaining blood pressure. Aldosterone is known to modulate NCC abundance. Previous studies reported that dietary salts modulated NCC abundance through either WNK4 [with no lysine (k) kinase 4]-SPAK (Ste20-related proline alanine-rich kinase) or WNK4-extracellular signal-regulated kinase-1 and -2 (ERK1/2) signaling pathways. To exclude the influence of SPAK signaling pathway on the role of the aldosterone-mediated ERK1/2 pathway in NCC regulation, we investigated the effects of dietary salt changes and aldosterone on NCC abundance in SPAK knockout (KO) mice. We found that in SPAK KO mice low-salt diet significantly increased total NCC abundance while reducing ERK1/2 phosphorylation, whereas high-salt diet decreased total NCC while increasing ERK1/2 phosphorylation. Importantly, exogenous aldosterone administration increased total NCC abundance in SPAK KO mice while increasing DUSP6 expression, an ERK1/2-specific phosphatase, and led to decreasing ERK1/2 phosphorylation without changing the ratio of phospho-T53-NCC/total NCC. In mouse distal convoluted tubule (mDCT) cells, aldosterone increased DUSP6 expression while reducing ERK1/2 phosphorylation. DUSP6 Knockdown increased ERK1/2 phosphorylation while reducing total NCC expression. Inhibition of DUSP6 by (E)-2-benzylidene-3-(cyclohexylamino)-2,3-dihydro-1H-inden-1-one increased ERK1/2 phosphorylation and reversed the aldosterone-mediated increments of NCC partly by increasing NCC ubiquitination. Therefore, these data suggest that aldosterone modulates NCC abundance via altering NCC ubiquitination through a DUSP6-dependent ERK1/2 signal pathway in SPAK KO mice and part of the effects of dietary salt changes may be mediated by aldosterone in the DCTs. PMID:25761881

  18. Caffeine Bitterness is Related to Daily Caffeine Intake and Bitter Receptor mRNA Abundance in Human Taste Tissue

    PubMed Central

    Lipchock, Sarah V.; Spielman, Andrew I.; Mennella, Julie A.; Mansfield, Corrine J.; Hwang, Liang-Dar; Douglas, Jennifer E.; Reed, Danielle R.

    2018-01-01

    We investigated whether the abundance of bitter receptor mRNA expression from human taste papillae is related to an individual’s perceptual ratings of bitter intensity and habitual intake of bitter drinks. Ratings of the bitterness of caffeine and quinine and three other bitter stimuli (urea, propylthiouracil, and denatonium benzoate) were compared with relative taste papilla mRNA abundance of bitter receptors that respond to the corresponding bitter stimuli in cell-based assays (TAS2R4, TAS2R10, TAS2R38, TAS2R43, and TAS2R46). We calculated caffeine and quinine intake from a food frequency questionnaire. The bitterness of caffeine was related to the abundance of the combined mRNA expression of these known receptors, r = 0.47, p = .05, and self-reported daily caffeine intake, t(18) = 2.78, p = .012. The results of linear modeling indicated that 47% of the variance among subjects in the rating of caffeine bitterness was accounted for by these two factors (habitual caffeine intake and taste receptor mRNA abundance). We observed no such relationships for quinine but consumption of its primary dietary form (tonic water) was uncommon. Overall, diet and TAS2R gene expression in taste papillae are related to individual differences in caffeine perception. PMID:28118781

  19. We the People: Indiana and the United States Constitution. Lectures in Observance of the Bicentennial of the Constitution.

    ERIC Educational Resources Information Center

    Furlong, Partick J.; And Others

    These lectures, presented in observance of the U.S. Constitution bicentennial celebration, consider selected constitutionally significant law cases that occurred in Indiana. These cases are representative of U.S. constitutional development and of the relationship of Indiana to the U.S. Constitution. Patrick Furlong, in "The South Bend…

  20. The Constitution by Cell

    ERIC Educational Resources Information Center

    Greenhut, Stephanie; Jones, Megan

    2010-01-01

    On their visit to the National Archives Experience in Washington, D.C., students in Jenni Ashley and Gay Brock's U.S. history classes at the Potomac School in McLean, Virginia, participated in a pilot program called "The Constitution by Cell." Armed with their cell phones, a basic understanding of the Constitution, and a willingness to…

  1. Reduced expression of IL-12 p35 by SJL/J macrophages responding to Theiler's virus infection is associated with constitutive activation of IRF-3

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Dahlberg, Angela; Auble, Mark R.; Petro, Thomas M.

    2006-09-30

    Macrophages responding to viral infections may contribute to autoimmune demyelinating diseases (ADD). Macrophages from ADD-susceptible SJL/J mice responding to Theiler's Virus (TMEV) infection, the TLR7 agonist loxoribine, or the TLR4 agonist-LPS expressed less IL-12 p35 but more IL-12/23 p40 and IFN-{beta} than macrophages from ADD-resistant B10.S mice. While expression of IRF-1 and -7 was similar between B10.S and SJL/J TMEV-infected macrophages, SJL/J but not B10.S macrophages exhibited constitutively active IRF-3. In contrast to overexpressed IRF-1, IRF-5, and IRF-7, which stimulated p35 promoter reporter activity, overexpressed IRF-3 repressed p35 promoter activity in response to TMEV infection, loxoribine, IFN-{gamma}/LPS, but not IFN-{gamma}more » alone. IRF-3 lessened but did not eliminate IRF-1-stimulated p35 promoter activity. Repression by IRF-3 required bp -172 to -122 of the p35 promoter. The data suggest that pre-activated IRF-3 is a major factor in the differences in IL-12 production between B10.S and SJL/J macrophages responding to TMEV.« less

  2. A landscape of circular RNA expression in the human heart.

    PubMed

    Tan, Wilson L W; Lim, Benson T S; Anene-Nzelu, Chukwuemeka G O; Ackers-Johnson, Matthew; Dashi, Albert; See, Kelvin; Tiang, Zenia; Lee, Dominic Paul; Chua, Wee Woon; Luu, Tuan D A; Li, Peter Y Q; Richards, Arthur Mark; Foo, Roger S Y

    2017-03-01

    Circular RNA (circRNA) is a newly validated class of single-stranded RNA, ubiquitously expressed in mammalian tissues and possessing key functions including acting as microRNA sponges and as transcriptional regulators by binding to RNA-binding proteins. While independent studies confirm the expression of circRNA in various tissue types, genome-wide circRNA expression in the heart has yet to be described in detail. We performed deep RNA-sequencing on ribosomal-depleted RNA isolated from 12 human hearts, 25 mouse hearts and across a 28-day differentiation time-course of human embryonic stem cell-derived cardiomyocytes. Using purpose-designed bioinformatics tools, we uncovered a total of 15 318 and 3017 cardiac circRNA within human and mouse, respectively. Their abundance generally correlates with the abundance of their cognate linear RNA, but selected circRNAs exist at disproportionately higher abundance. Top highly expressed circRNA corresponded to key cardiac genes including Titin (TTN), RYR2, and DMD. The most abundant cardiac-expressed circRNA is a cytoplasmic localized single-exon circSLC8A1-1. The longest human transcript TTN alone generates up to 415 different exonic circRNA isoforms, the majority (83%) of which originates from the I-band domain. Finally, we confirmed the expression of selected cardiac circRNA by RT-PCR, Sanger sequencing and single molecule RNA-fluorescence in situ hybridization. Our data provide a detailed circRNA expression landscape in hearts. There is a high-abundance of specific cardiac-expressed circRNA. These findings open up a new avenue for future investigation into this emerging class of RNA. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2016. For Permissions, please email: journals.permissions@oup.com.

  3. Constitutional Reform for Conflict Management

    DTIC Science & Technology

    2014-04-01

    conflict. Empirical studies reveal that both accommodative and integrative constitutional design can produce political stability , if properly...reform, and the ability of various constitutional designs to promote democracy and political stability .

  4. The low-abundance transcriptome reveals novel biomarkers, specific intracellular pathways and targetable genes associated with advanced gastric cancer.

    PubMed

    Bizama, Carolina; Benavente, Felipe; Salvatierra, Edgardo; Gutiérrez-Moraga, Ana; Espinoza, Jaime A; Fernández, Elmer A; Roa, Iván; Mazzolini, Guillermo; Sagredo, Eduardo A; Gidekel, Manuel; Podhajcer, Osvaldo L

    2014-02-15

    Studies on the low-abundance transcriptome are of paramount importance for identifying the intimate mechanisms of tumor progression that can lead to novel therapies. The aim of the present study was to identify novel markers and targetable genes and pathways in advanced human gastric cancer through analyses of the low-abundance transcriptome. The procedure involved an initial subtractive hybridization step, followed by global gene expression analysis using microarrays. We observed profound differences, both at the single gene and gene ontology levels, between the low-abundance transcriptome and the whole transcriptome. Analysis of the low-abundance transcriptome led to the identification and validation by tissue microarrays of novel biomarkers, such as LAMA3 and TTN; moreover, we identified cancer type-specific intracellular pathways and targetable genes, such as IRS2, IL17, IFNγ, VEGF-C, WISP1, FZD5 and CTBP1 that were not detectable by whole transcriptome analyses. We also demonstrated that knocking down the expression of CTBP1 sensitized gastric cancer cells to mainstay chemotherapeutic drugs. We conclude that the analysis of the low-abundance transcriptome provides useful insights into the molecular basis and treatment of cancer. © 2013 UICC.

  5. Cell surface expression of bacterial esterase A by Saccharomyces cerevisiae and its enhancement by constitutive activation of the cellular unfolded protein response.

    PubMed

    Breinig, Frank; Diehl, Björn; Rau, Sabrina; Zimmer, Christian; Schwab, Helmut; Schmitt, Manfred J

    2006-11-01

    Yeast cell surface display is a powerful tool for expression and immobilization of biocatalytically active proteins on a unicellular eukaryote. Here bacterial carboxylesterase EstA from Burkholderia gladioli was covalently anchored into the cell wall of Saccharomyces cerevisiae by in-frame fusion to the endogenous yeast proteins Kre1p, Cwp2p, and Flo1p. When p-nitrophenyl acetate was used as a substrate, the esterase specific activities of yeast expressing the protein fusions were 103 mU mg(-1) protein for Kre1/EstA/Cwp2p and 72 mU mg(-1) protein for Kre1/EstA/Flo1p. In vivo cell wall targeting was confirmed by esterase solubilization after laminarinase treatment and immunofluorescence microscopy. EstA expression resulted in cell wall-associated esterase activities of 2.72 U mg(-1) protein for Kre1/EstA/Cwp2p and 1.27 U mg(-1) protein for Kre1/EstA/Flo1p. Furthermore, esterase display on the yeast cell surface enabled the cells to effectively grow on the esterase-dependent carbon source glycerol triacetate (Triacetin). In the case of Kre1/EstA/Flo1p, in vivo maturation within the yeast secretory pathway and final incorporation into the wall were further enhanced when there was constitutive activation of the unfolded protein response pathway. Our results demonstrate that esterase cell surface display in yeast, which, as shown here, is remarkably more effective than EstA surface display in Escherichia coli, can be further optimized by activating the protein folding machinery in the eukaryotic secretion pathway.

  6. Constitutional multicenter bank linked to Sasang constitutional phenotypic data.

    PubMed

    Jin, Hee-Jeong; Baek, Younghwa; Kim, Ho-Seok; Ryu, Jonghyang; Lee, Siwoo

    2015-03-10

    Biobanks are more important in medical area because they can give researchers data for demonstrating and validating their research. In this study, we developed a biobank called the Korea Constitutional Multicenter Bank (KCMB) based on Sasang Constitutional Medicine (SCM). The aim of the KCMB was a foundation to providing the scientific basis of SCM. The KCMB has been constructed since 2006 in 24 Korean medical clinics with collection of questionnaire data, physical measurements and biological information comprised the results from blood test and DNA analyses. All participants were prescribed Sasang Constitution (SC)-specific herbal remedies for the treatment, and showed improvement of original symptoms as confirmed by Korean medicine doctor. Collected data went through de-identification process using the electronic case report form system. For calculation of several SC type specific tendencies, we used the direct standardization and Chi-square tests. The KCMB collected clinical information from 3,711 study participants (1,353 men and 2,358 women) aged more than 10 years. The mean age (± standard deviation) was 47.1 (±16.6) and 47.7 (±15.8) years for men and women respectively. After applying the direct standardization, the estimated constitutional distributions for the SC types were as follows: 39.2% for Tae-eumin(TE), 27.1% for Soeumin(SE), 33.7% for Soyangyin(SY), and non-zero but below 0.1% for Taeyangyin(TY). The estimated distribution of TE was about 10% less, while that of SY and SE were slightly more than the distribution reported by Jema Lee established the SCM. Based on the participants' medical history within the KCMB, each SC type had notably different frequencies for some diseases such as hypertension, diabetes, hyperlipidemia, stroke, and obesity (P < 0.001). The KCMB may serve to verify and validate SCM theories and practices. It may also provide new insights into SCM mechanisms. The results from many studies using the KCMB data are of great

  7. Ceruloplasmin expression by human peripheral blood lymphocytes: a new link between immunity and iron metabolism.

    PubMed

    Banha, João; Marques, Liliana; Oliveira, Rita; Martins, Maria de Fátima; Paixão, Eleonora; Pereira, Dina; Malhó, Rui; Penque, Deborah; Costa, Luciana

    2008-02-01

    Ceruloplasmin (CP) is a multicopper oxidase involved in the acute phase reaction to stress. Although the physiological role of CP is uncertain, its role in iron (Fe) homeostasis and protection against free radical-initiated cell injury has been widely documented. Previous studies showed the existence of two molecular isoforms of CP: secreted CP (sCP) and a membrane glycosylphosphatidylinositol (GPI)-anchored form of CP (GPI-CP). sCP is produced mainly by the liver and is abundant in human serum whereas GPI-CP is expressed in mammalian astrocytes, rat leptomeningeal cells, and Sertolli cells. Herein, we show using RT-PCR that human peripheral blood lymphocytes (huPBL) constitutively express the transcripts for both CP molecular isoforms previously reported. Also, expression of CP in huPBL is demonstrated by immunofluorescence with confocal microscopy and flow cytometry analysis using cells isolated from healthy blood donors with normal Fe status. Importantly, the results obtained show that natural killer cells have a significantly higher CP expression compared to all other major lymphocyte subsets. In this context, the involvement of lymphocyte-derived CP on host defense processes via its anti/prooxidant properties is proposed, giving further support for a close functional interaction between the immune system and the Fe metabolism.

  8. [Constitutional-somatotopic factors in the clinical picture of alcoholic psychoses and obvious signs of paranoid schizophrenia].

    PubMed

    Kornetov, N A; Gubernik, V Ia

    1980-01-01

    The clinical and anthropometrical methods were used to study the constitutional traits of 240 patients with psychotic forms of alcoholism and 300 patients with paranoid schizophrenia. The somatotypes were assessed by the Rees-Eysenck method. The prepsychotic parameters of alcoholism and the structure of alcoholic delirium were studied in 78 patients, as well as the manifest symptoms of paranoid schizophrenia. The study demonstrated a syndromal modified expressivity in exo- and endogenous processes, depending upon the somatotypical constitution. Simple syndromes were more frequently found in patients with a pyknic somatotype, and complicated in normosthenical and asthenic types of body built. It is suggested that the constitutional and somatotypical factors be included into the multifactorial analysis of the syndrome structure and prognosis in the development of the psychosis.

  9. Insulin-regulated aminopeptidase immunoreactivity is abundantly present in human hypothalamus and posterior pituitary gland, with reduced expression in paraventricular and suprachiasmatic neurons in chronic schizophrenia.

    PubMed

    Bernstein, Hans-Gert; Müller, Susan; Dobrowolny, Hendrik; Wolke, Carmen; Lendeckel, Uwe; Bukowska, Alicja; Keilhoff, Gerburg; Becker, Axel; Trübner, Kurt; Steiner, Johann; Bogerts, Bernhard

    2017-08-01

    The vasopressin- and oxytocin-degrading enzyme insulin-regulated aminopeptidase (IRAP) is expressed in various organs including the brain. However, knowledge about its presence in human hypothalamus is fragmentary. Functionally, for a number of reasons (genetic linkage, hydrolysis of oxytocin and vasopressin, its role as angiotensin IV receptor in learning and memory and others) IRAP might play a role in schizophrenia. We studied the regional and cellular localization of IRAP in normal human brain with special emphasis on the hypothalamus and determined numerical densities of IRAP-expressing cells in the paraventricular, supraoptic and suprachiasmatic nuclei in schizophrenia patients and controls. By using immunohistochemistry and Western blot analysis, IRAP was immunolocalized in postmortem human brains. Cell countings were performed to estimate numbers and numerical densities of IRAP immunoreactive hypothalamic neurons in schizophrenia patients and control cases. Shape, size and regional distribution of IRAP-expressing cells, as well the lack of co-localization with the glia marker glutamine synthetase, show that IRAP is expressed in neurons. IRAP immunoreactive cells were observed in the hippocampal formation, cerebral cortex, thalamus, amygdala and, abundantly, hypothalamus. Double labeling experiments (IRAP and oxytocin/neurophysin 1, IRAP with vasopressin/neurophysin 2) revealed that IRAP is present in oxytocinergic and in vasopressinergic neurons. In schizophrenia patients, the numerical density of IRAP-expressing neurons in the paraventricular and the suprachiasmatic nuclei is significantly reduced, which might be associated with the reduction in neurophysin-containing neurons in these nuclei in schizophrenia. The pathophysiological role of lowered hypothalamic IRAP expression in schizophrenia remains to be established.

  10. Metalloprotease cleavage of the N terminus of the orphan G protein-coupled receptor GPR37L1 reduces its constitutive activity.

    PubMed

    Coleman, James L J; Ngo, Tony; Schmidt, Johannes; Mrad, Nadine; Liew, Chu Kong; Jones, Nicole M; Graham, Robert M; Smith, Nicola J

    2016-04-12

    Little is known about the pharmacology or physiology of GPR37L1, a G protein (heterotrimeric guanine nucleotide-binding protein)-coupled receptor that is abundant in the cerebellum. Mice deficient in this receptor exhibit precocious cerebellar development and hypertension. We showed that GPR37L1 coupled to the G protein Gα(s) when heterologously expressed in cultured cells in the absence of any added ligand, whereas a mutant receptor that lacked the amino terminus was inactive. Conversely, inhibition of ADAMs (a disintegrin and metalloproteases) enhanced receptor activity, indicating that the presence of the amino terminus is necessary for GPR37L1 signaling. Metalloprotease-dependent processing of GPR37L1 was evident in rodent cerebellum, where we detected predominantly the cleaved, inactive form. However, comparison of the accumulation of cAMP (adenosine 3',5'-monophosphate) in response to phosphodiesterase inhibition in cerebellar slice preparations from wild-type and GPR37L1-null mice showed that some constitutive signaling remained in the wild-type mice. In reporter assays of Gα(s) or Gα(i) signaling, the synthetic, prosaposin-derived peptide prosaptide (TX14A) did not increase GPR37L1 activity. Our data indicate that GPR37L1 may be a constitutively active receptor, or perhaps its ligand is present under the conditions that we used for analysis, and that the activity of this receptor is instead controlled by signals that regulate metalloprotease activity in the tissue. Copyright © 2016, American Association for the Advancement of Science.

  11. Zipf's Law in Gene Expression

    NASA Astrophysics Data System (ADS)

    Furusawa, Chikara; Kaneko, Kunihiko

    2003-02-01

    Using data from gene expression databases on various organisms and tissues, including yeast, nematodes, human normal and cancer tissues, and embryonic stem cells, we found that the abundances of expressed genes exhibit a power-law distribution with an exponent close to -1; i.e., they obey Zipf’s law. Furthermore, by simulations of a simple model with an intracellular reaction network, we found that Zipf’s law of chemical abundance is a universal feature of cells where such a network optimizes the efficiency and faithfulness of self-reproduction. These findings provide novel insights into the nature of the organization of reaction dynamics in living cells.

  12. Constitutional aneuploidy and cancer predisposition.

    PubMed

    Ganmore, Ithamar; Smooha, Gil; Izraeli, Shai

    2009-04-15

    Constitutional aneuploidies are rare syndromes associated with multiple developmental abnormalities and the alterations in the risk for specific cancers. Acquired somatic chromosomal aneuploidies are the most common genetic aberrations in sporadic cancers. Thus studies of these rare constitutional aneuploidy syndromes are important not only for patient counseling and clinical management, but also for deciphering the mechanisms by which chromosomal aneuploidy affect cancer initiation and progression. Here we review the major constitutional aneuploidy syndromes and suggest some general mechanisms for the associated cancer predisposition.

  13. Chromosome engineering of Escherichia coli for constitutive production of salvianic acid A.

    PubMed

    Zhou, Liang; Ding, Qi; Jiang, Guo-Zhen; Liu, Zhen-Ning; Wang, Hai-Yan; Zhao, Guang-Rong

    2017-05-16

    Salvianic acid A (SAA), a valuable natural product from herbal plant Salvia miltiorrhiza, exhibits excellent antioxidant activities on food industries and efficacious therapeutic potential on cardiovascular diseases. Recently, production of SAA in engineered Escherichia coli was established via the artificial biosynthetic pathway of SAA on the multiple plasmids in our previous work. However, the plasmid-mediated system required to supplement expensive inducers and antibiotics during the fermentation process, restricting scale-up production of SAA. Microbial cell factory would be an attractive approach for constitutive production of SAA by chromosome engineering. The limited enzymatic reactions in SAA biosynthetic pathway from glucose were grouped into three modules, which were sequentially integrated into chromosome of engineered E. coli by λ Red homologous recombination method. With starting strain E. coli BAK5, in which the ptsG, pykF, pykA, pheA and tyrR genes were previously deleted, chassis strain BAK11 was constructed for constitutive production of precursor L-tyrosine by replacing the 17.7-kb mao-paa cluster with module 1 (P lacUV5 -aroG fbr -tyrA fbr -aroE) and the lacI gene with module 2 (P trc -glk-tktA-ppsA). The synthetic 5tacs promoter demonstrated the optimal strength to drive the expression of hpaBC-d-ldh Y52A in module 3, which then was inserted at the position between nupG and speC on the chromosome of strain BAK11. The final strain BKD13 produced 5.6 g/L of SAA by fed-batch fermentation in 60 h from glucose without any antibiotics and inducers supplemented. The plasmid-free and inducer-free strain for SAA production was developed by targeted integration of the constitutive expression of SAA biosynthetic genes into E. coli chromosome. Our work provides the industrial potential for constitutive production of SAA by the indel microbial cell factory and also sets an example of further producing other valuable natural and unnatural products.

  14. Ligand stimulation of ErbB4 and a constitutively-active ErbB4 mutant result in different biological responses in human pancreatic tumor cell lines

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Mill, Christopher P.; Auburn University Harrison School of Pharmacy, Auburn, AL 36849-5501; Gettinger, Kathleen L.

    2011-02-15

    Pancreatic cancer is the fourth leading cause of cancer death in the United States. Indeed, it has been estimated that 37,000 Americans will die from this disease in 2010. Late diagnosis, chemoresistance, and radioresistance of these tumors are major reasons for poor patient outcome, spurring the search for pancreatic cancer early diagnostic and therapeutic targets. ErbB4 (HER4) is a member of the ErbB family of receptor tyrosine kinases (RTKs), a family that also includes the Epidermal Growth Factor Receptor (EGFR/ErbB1/HER1), Neu/ErbB2/HER2, and ErbB3/HER3. These RTKs play central roles in many human malignancies by regulating cell proliferation, survival, differentiation, invasiveness, motility,more » and apoptosis. In this report we demonstrate that human pancreatic tumor cell lines exhibit minimal ErbB4 expression; in contrast, these cell lines exhibit varied and in some cases abundant expression and basal tyrosine phosphorylation of EGFR, ErbB2, and ErbB3. Expression of a constitutively-dimerized and -active ErbB4 mutant inhibits clonogenic proliferation of CaPan-1, HPAC, MIA PaCa-2, and PANC-1 pancreatic tumor cell lines. In contrast, expression of wild-type ErbB4 in pancreatic tumor cell lines potentiates stimulation of anchorage-independent colony formation by the ErbB4 ligand Neuregulin 1{beta}. These results illustrate the multiple roles that ErbB4 may be playing in pancreatic tumorigenesis and tumor progression.« less

  15. Constituting children's bodily integrity.

    PubMed

    Hill, B Jessie

    2015-04-01

    Children have a constitutional right to bodily integrity. Courts do not hesitate to vindicate that right when children are abused by state actors. Moreover, in at least some cases, a child's right to bodily integrity applies within the family, giving the child the right to avoid unwanted physical intrusions regardless of the parents' wishes. Nonetheless, the scope of this right vis-à-vis the parents is unclear; the extent to which it applies beyond the narrow context of abortion and contraception has been almost entirely unexplored and untheorized. This Article is the first in the legal literature to analyze the constitutional right of minors to bodily integrity within the family by spanning traditionally disparate doctrinal categories such as abortion rights; corporal punishment; medical decisionmaking; and nontherapeutic physical interventions such as tattooing, piercing, and circumcision. However, the constitutional right of minors to bodily integrity raises complex philosophical questions concerning the proper relationship between family and state, as well as difficult doctrinal and theoretical issues concerning the ever-murky idea of state action. This Article canvasses those issues with the ultimate goal of delineating a constitutional right of bodily security and autonomy for children.

  16. Retroelements (LINEs and SINEs) in vole genomes: differential distribution in the constitutive heterochromatin.

    PubMed

    Acosta, M J; Marchal, J A; Fernández-Espartero, C H; Bullejos, M; Sánchez, A

    2008-01-01

    The chromosomal distribution of mobile genetic elements is scarcely known in Arvicolinae species, but could be of relevance to understand the origin and complex evolution of the sex chromosome heterochromatin. In this work we cloned two retrotransposon sequences, L1 and SINE-B1, from the genome of Chionomys nivalis and investigated their chromosomal distribution on several arvicoline species. Our results demonstrate first that both retroelements are the most abundant repeated DNA sequences in the genome of these species. L1 elements, in most species, are highly accumulated in the sex chromosomes compared to the autosomes. This favoured L1 insertion could have played an important role in the origin of the enlarged heterochromatic blocks existing in the sex chromosomes of some Microtus species. Also, we propose that L1 accumulation on the X heterochromatin could have been the consequence of different, independent and rapid amplification processes acting in each species. SINE elements, however, were completely lacking from the constitutive heterochromatin, either in autosomes or in the heterochromatic blocks of sex chromosomes. These data could indicate that some SINE elements are incompatible with the formation of heterochromatic complexes and hence are necessarily missing from the constitutive heterochromatin.

  17. Regulation of infection efficiency in a globally abundant marine Bacteriodetes virus

    DOE PAGES

    Howard-Varona, Cristina; Roux, Simon; Dore, Hugo; ...

    2016-05-17

    Microbes impact human health and disease, industrial processes and natural ecosystems, but do so under the influence of viruses. Problematically, knowledge of viral infection efficiencies and outcomes (e.g. lysis, lysogeny) derives from few model systems that over-represent efficient, lytic infections and under-represent virus-host natural diversity. Here we sought to understand how infection efficiency is regulated in an environmental Bacteroidetes virus that represents a globally abundant viral group and has drastically different infection efficiencies when infecting two nearly identical bacterial strains. To this end, we quantified bacterial virus (phage) and host DNA, transcripts and phage particles throughout the infection of bothmore » bacterial hosts. While the phage transcriptome was similar during both infections, host transcriptional differences appeared to have altered infection efficiency. Specifically, host transcriptomes suggested that the phage failed to repress early host expression in the inefficient nfection, thereby allowing the host to respond against infection by delaying phage DNA replication and protein translation. Further measurements showed that phage DNA and particle production were delayed (by >30 minutes) and reduced (by >50%) in the inefficient versus efficient infection as the host over-expressed DNA degradation genes and under-expressed translation genes, respectively. Together these results suggest that multiple levels of regulation can impact infection efficiencies as failure to repress host transcription allowed the host to defend against both phage DNA and protein production. Given that this phage type is ubiquitous and abundant in the global oceans and that variably efficient viral infections are likely common in any ecosystem with varying phage-host abundances and physiological states, these data provide a critically needed foundation for understanding and modeling viral infection efficiency in nature.« less

  18. Regulation of infection efficiency in a globally abundant marine Bacteriodetes virus

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Howard-Varona, Cristina; Roux, Simon; Dore, Hugo

    Microbes impact human health and disease, industrial processes and natural ecosystems, but do so under the influence of viruses. Problematically, knowledge of viral infection efficiencies and outcomes (e.g. lysis, lysogeny) derives from few model systems that over-represent efficient, lytic infections and under-represent virus-host natural diversity. Here we sought to understand how infection efficiency is regulated in an environmental Bacteroidetes virus that represents a globally abundant viral group and has drastically different infection efficiencies when infecting two nearly identical bacterial strains. To this end, we quantified bacterial virus (phage) and host DNA, transcripts and phage particles throughout the infection of bothmore » bacterial hosts. While the phage transcriptome was similar during both infections, host transcriptional differences appeared to have altered infection efficiency. Specifically, host transcriptomes suggested that the phage failed to repress early host expression in the inefficient nfection, thereby allowing the host to respond against infection by delaying phage DNA replication and protein translation. Further measurements showed that phage DNA and particle production were delayed (by >30 minutes) and reduced (by >50%) in the inefficient versus efficient infection as the host over-expressed DNA degradation genes and under-expressed translation genes, respectively. Together these results suggest that multiple levels of regulation can impact infection efficiencies as failure to repress host transcription allowed the host to defend against both phage DNA and protein production. Given that this phage type is ubiquitous and abundant in the global oceans and that variably efficient viral infections are likely common in any ecosystem with varying phage-host abundances and physiological states, these data provide a critically needed foundation for understanding and modeling viral infection efficiency in nature.« less

  19. [Comparative research into the process of forming the theory of constitution in ancient western medicine and that of four trigrams constitution in Korean medicine and contents of two theories of constitution].

    PubMed

    Park, Joo-Hong

    2009-06-01

    After conducting comparative research into the process of forming the Theory of Constitution in Ancient Western Medicine and that of Four Trigrams Constitution(Sasang Constitution) in Korean Medicine and contents of two Theories of Constitution in terms of medical history, both theories were found to be formed by an interaction between philosophy and medicine, followed by a combination of the two, on a philosophical basis. The Theory of Constitution in Ancient Western Medicine began with the Theory of Four Elements presented by Empedocles, followed by the Theory of Four Humors presented by Hippocrates and the Theory of Four Temperaments by Galenos, forming and developing the Theory of Constitution. After the Middle Ages, there was no significant advance in the Theory of Constitution by modern times ; however, it developed into the theory of constitution type of Kretschmer and others after the 19th century and into the scientific theory of constitution based on genetics presented by Garrod and others early in the 20th century. The Theory of Four Trigrams Constitution began with the Theory of Constitution in Huangdi Neijing, followed by developments and influences of existing medicine called beginning, restoration, and revival periods and DongeuisoosebowonSaSangChoBonGwon based on the original philosophy of Four Trigrams presented by Lee Je-ma, which is found in GyeokChiGo, DongMuYuGo and so on, ultimately forming and developing into the Theory of Four Trigrams Constitution in Dongeuisoosebowon. Recently, a lot of research is being conducted into making it objective in order to achieve reproducibility in diagnosis and so forth of Four Trigrams Constitution.

  20. Constitutional aneuploidy and cancer predisposition†

    PubMed Central

    Ganmore, Ithamar; Smooha, Gil; Izraeli, Shai

    2009-01-01

    Constitutional aneuploidies are rare syndromes associated with multiple developmental abnormalities and the alterations in the risk for specific cancers. Acquired somatic chromosomal aneuploidies are the most common genetic aberrations in sporadic cancers. Thus studies of these rare constitutional aneuploidy syndromes are important not only for patient counseling and clinical management, but also for deciphering the mechanisms by which chromosomal aneuploidy affect cancer initiation and progression. Here we review the major constitutional aneuploidy syndromes and suggest some general mechanisms for the associated cancer predisposition. PMID:19297405

  1. Can Occupancy–Abundance Models Be Used to Monitor Wolf Abundance?

    PubMed Central

    Latham, M. Cecilia; Latham, A. David M.; Webb, Nathan F.; Mccutchen, Nicole A.; Boutin, Stan

    2014-01-01

    Estimating the abundance of wild carnivores is of foremost importance for conservation and management. However, given their elusive habits, direct observations of these animals are difficult to obtain, so abundance is more commonly estimated from sign surveys or radio-marked individuals. These methods can be costly and difficult, particularly in large areas with heavy forest cover. As an alternative, recent research has suggested that wolf abundance can be estimated from occupancy–abundance curves derived from “virtual” surveys of simulated wolf track networks. Although potentially more cost-effective, the utility of this approach hinges on its robustness to violations of its assumptions. We assessed the sensitivity of the occupancy–abundance approach to four assumptions: variation in wolf movement rates, changes in pack cohesion, presence of lone wolves, and size of survey units. Our simulations showed that occupancy rates and wolf pack abundances were biased high if track surveys were conducted when wolves made long compared to short movements, wolf packs were moving as multiple hunting units as opposed to a cohesive pack, and lone wolves were moving throughout the surveyed landscape. We also found that larger survey units (400 and 576 km2) were more robust to changes in these factors than smaller survey units (36 and 144 km2). However, occupancy rates derived from large survey units rapidly reached an asymptote at 100% occupancy, suggesting that these large units are inappropriate for areas with moderate to high wolf densities (>15 wolves/1,000 km2). Virtually-derived occupancy–abundance relationships can be a useful method for monitoring wolves and other elusive wildlife if applied within certain constraints, in particular biological knowledge of the surveyed species needs to be incorporated into the design of the occupancy surveys. Further, we suggest that the applicability of this method could be extended by directly incorporating some of its

  2. Trade-offs between induced and constitutive resistance in two pine species: secondary chemistry, effective antiherbivore-resistance, and effect of nutrient availability

    Treesearch

    Luis Sampedro; Xoaquín Moreira; Rafael Zas

    2012-01-01

    Constitutive chemical defenses, always expressed in the plants, and plastic defensive responses, those mobilized in response to plant injury or other cues or herbivory risk, differ in their benefits in terms of fitness for long-lived plants. Induced defenses are considered to be less expensive than constitutive preformed defenses since the cost is realized only when...

  3. APOGEE Chemical Abundances of the Sagittarius Dwarf Galaxy

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hasselquist, Sten; Holtzman, Jon; Shetrone, Matthew

    The Apache Point Observatory Galactic Evolution Experiment provides the opportunity of measuring elemental abundances for C, N, O, Na, Mg, Al, Si, P, K, Ca, V, Cr, Mn, Fe, Co, and Ni in vast numbers of stars. We analyze thechemical-abundance patterns of these elements for 158 red giant stars belonging to the Sagittarius dwarf galaxy (Sgr). This is the largest sample of Sgr stars with detailed chemical abundances, and it is the first time that C, N, P, K, V, Cr, Co, and Ni have been studied at high resolution in this galaxy. We find that the Sgr stars withmore » [Fe/H] ≳ −0.8 are deficient in all elemental abundance ratios (expressed as [X/Fe]) relative to the Milky Way, suggesting that the Sgr stars observed today were formed from gas that was less enriched by Type II SNe than stars formed in the Milky Way. By examining the relative deficiencies of the hydrostatic (O, Na, Mg, and Al) and explosive (Si, P, K, and Mn) elements, our analysis supports the argument that previous generations of Sgr stars were formed with a top-light initial mass function, one lacking the most massive stars that would normally pollute the interstellar medium with the hydrostatic elements. We use a simple chemical-evolution model, flexCE, to further support our claim and conclude that recent stellar generations of Fornax and the Large Magellanic Cloud could also have formed according to a top-light initial mass function.« less

  4. Increased Production of Fatty Acids and Triglycerides in Aspergillus oryzae by Enhancing Expressions of Fatty Acid Synthesis-Related Genes

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tamano, Koichi; Bruno, Kenneth S.; Karagiosis, Sue A.

    2013-01-01

    Microbial production of fats and oils is being developedas a means of converting biomass to biofuels. Here we investigate enhancing expression of enzymes involved in the production of fatty acids and triglycerides as a means to increase production of these compounds in Aspergillusoryzae. Examination of the A.oryzaegenome demonstrates that it contains twofatty acid synthases and several other genes that are predicted to be part of this biosynthetic pathway. We enhancedthe expressionof fatty acid synthesis-related genes by replacing their promoters with thepromoter fromthe constitutively highly expressedgene tef1. We demonstrate that by simply increasing the expression of the fatty acid synthasegenes wemore » successfullyincreasedtheproduction of fatty acids and triglyceridesby more than two fold. Enhancement of expression of the fatty acid pathway genes ATP-citrate lyase and palmitoyl-ACP thioesteraseincreasedproductivity to a lesser extent.Increasing expression ofacetyl-CoA carboxylase caused no detectable change in fatty acid levels. Increases in message level for each gene were monitored usingquantitative real-time RT-PCR. Our data demonstrates that a simple increase in the abundance of fatty acid synthase genes can increase the detectable amount of fatty acids.« less

  5. 29 CFR 452.18 - Constitutional officers.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... officer refers to a person holding a position identified as an officer by the constitution and bylaws of... constitution identifies the holder of such a position as an officer. On the other hand, legislative representatives who are required to be elected by the constitution and bylaws of a labor organization are not...

  6. 7 CFR 718.201 - Farm constitution.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 7 2010-01-01 2010-01-01 false Farm constitution. 718.201 Section 718.201 Agriculture... Reconstitution of Farms, Allotments, Quotas, and Bases § 718.201 Farm constitution. (a) In order to implement... this section. The constitution and identification of land as a farm for the first time and the...

  7. 29 CFR 452.18 - Constitutional officers.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... officer refers to a person holding a position identified as an officer by the constitution and bylaws of... constitution identifies the holder of such a position as an officer. On the other hand, legislative representatives who are required to be elected by the constitution and bylaws of a labor organization are not...

  8. 7 CFR 718.201 - Farm constitution.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 7 2012-01-01 2012-01-01 false Farm constitution. 718.201 Section 718.201 Agriculture... Reconstitution of Farms, Allotments, Quotas, and Bases § 718.201 Farm constitution. (a) In order to implement... this section. The constitution and identification of land as a farm for the first time and the...

  9. 7 CFR 718.201 - Farm constitution.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 7 2013-01-01 2013-01-01 false Farm constitution. 718.201 Section 718.201 Agriculture... Reconstitution of Farms, Allotments, Quotas, and Bases § 718.201 Farm constitution. (a) In order to implement... this section. The constitution and identification of land as a farm for the first time and the...

  10. 29 CFR 452.18 - Constitutional officers.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... officer refers to a person holding a position identified as an officer by the constitution and bylaws of... constitution identifies the holder of such a position as an officer. On the other hand, legislative representatives who are required to be elected by the constitution and bylaws of a labor organization are not...

  11. 7 CFR 718.201 - Farm constitution.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 7 2011-01-01 2011-01-01 false Farm constitution. 718.201 Section 718.201 Agriculture... Reconstitution of Farms, Allotments, Quotas, and Bases § 718.201 Farm constitution. (a) In order to implement... this section. The constitution and identification of land as a farm for the first time and the...

  12. 7 CFR 718.201 - Farm constitution.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 7 2014-01-01 2014-01-01 false Farm constitution. 718.201 Section 718.201 Agriculture... Reconstitution of Farms, Allotments, Quotas, and Bases § 718.201 Farm constitution. (a) In order to implement... this section. The constitution and identification of land as a farm for the first time and the...

  13. 29 CFR 452.18 - Constitutional officers.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... officer refers to a person holding a position identified as an officer by the constitution and bylaws of... constitution identifies the holder of such a position as an officer. On the other hand, legislative representatives who are required to be elected by the constitution and bylaws of a labor organization are not...

  14. Digital gene expression for non-model organisms

    PubMed Central

    Hong, Lewis Z.; Li, Jun; Schmidt-Küntzel, Anne; Warren, Wesley C.; Barsh, Gregory S.

    2011-01-01

    Next-generation sequencing technologies offer new approaches for global measurements of gene expression but are mostly limited to organisms for which a high-quality assembled reference genome sequence is available. We present a method for gene expression profiling called EDGE, or EcoP15I-tagged Digital Gene Expression, based on ultra-high-throughput sequencing of 27-bp cDNA fragments that uniquely tag the corresponding gene, thereby allowing direct quantification of transcript abundance. We show that EDGE is capable of assaying for expression in >99% of genes in the genome and achieves saturation after 6–8 million reads. EDGE exhibits very little technical noise, reveals a large (106) dynamic range of gene expression, and is particularly suited for quantification of transcript abundance in non-model organisms where a high-quality annotated genome is not available. In a direct comparison with RNA-seq, both methods provide similar assessments of relative transcript abundance, but EDGE does better at detecting gene expression differences for poorly expressed genes and does not exhibit transcript length bias. Applying EDGE to laboratory mice, we show that a loss-of-function mutation in the melanocortin 1 receptor (Mc1r), recognized as a Mendelian determinant of yellow hair color in many different mammals, also causes reduced expression of genes involved in the interferon response. To illustrate the application of EDGE to a non-model organism, we examine skin biopsy samples from a cheetah (Acinonyx jubatus) and identify genes likely to control differences in the color of spotted versus non-spotted regions. PMID:21844123

  15. Expression Plasmids for Use in Candida glabrata

    PubMed Central

    Zordan, Rebecca E.; Ren, Yuxia; Pan, Shih-Jung; Rotondo, Giuseppe; Peñas, Alejandro De Las; Iluore, Joseph; Cormack, Brendan P.

    2013-01-01

    We describe a series of CEN/ARS episomal plasmids containing different Candida glabrata promoters, allowing for a range of constitutive or regulated expression of proteins in C. glabrata. The set of promoters includes three constitutive promoters (EGD2pr, HHT2pr, PDC1pr), two macrophage/phagocytosis-induced promoters (ACO2pr, LYS21pr), and one nutritionally regulated promoter (MET3pr). Each promoter was cloned into two plasmid backbones that differ in their selectable marker, URA3, or the dominant-selectable NAT1 gene, which confers resistance to the drug nourseothricin. Expression from the 12 resulting plasmids was assessed using GFP as a reporter and flow cytometry or quantitative reverse-transcription polymerase chain reaction to assess expression levels. Together this set of plasmids expands the toolkit of expression vectors available for use with C. glabrata. PMID:23934995

  16. Phenol emulsion-enhanced DNA-driven subtractive cDNA cloning: isolation of low-abundance monkey cortex-specific mRNAs.

    PubMed Central

    Travis, G H; Sutcliffe, J G

    1988-01-01

    To isolate cDNA clones of low-abundance mRNAs expressed in monkey cerebral cortex but absent from cerebellum, we developed an improved subtractive cDNA cloning procedure that requires only modest quantities of mRNA. Plasmid DNA from a monkey cerebellum cDNA library was hybridized in large excess to radiolabeled monkey cortex cDNA in a phenol emulsion-enhanced reaction. The unhybridized cortex cDNA was isolated by chromatography on hydroxyapatite and used to probe colonies from a monkey cortex cDNA library. Of 60,000 colonies screened, 163 clones were isolated and confirmed by colony hybridization or RNA blotting to represent mRNAs, ranging from 0.001% to 0.1% abundance, specific to or highly enriched in cerebral cortex relative to cerebellum. Clones of one medium-abundance mRNA were recovered almost quantitatively. Two of the lower-abundance mRNAs were expressed at levels reduced by a factor of 10 in Alzheimer disease relative to normal human cortex. One of these was identified as the monkey preprosomatostatin I mRNA. Images PMID:2894033

  17. Childhood tuberculosis is associated with decreased abundance of T cell gene transcripts and impaired T cell function.

    PubMed

    Hemingway, Cheryl; Berk, Maurice; Anderson, Suzanne T; Wright, Victoria J; Hamilton, Shea; Eleftherohorinou, Hariklia; Kaforou, Myrsini; Goldgof, Greg M; Hickman, Katy; Kampmann, Beate; Schoeman, Johan; Eley, Brian; Beatty, David; Pienaar, Sandra; Nicol, Mark P; Griffiths, Michael J; Waddell, Simon J; Newton, Sandra M; Coin, Lachlan J; Relman, David A; Montana, Giovanni; Levin, Michael

    2017-01-01

    The WHO estimates around a million children contract tuberculosis (TB) annually with over 80 000 deaths from dissemination of infection outside of the lungs. The insidious onset and association with skin test anergy suggests failure of the immune system to both recognise and respond to infection. To understand the immune mechanisms, we studied genome-wide whole blood RNA expression in children with TB meningitis (TBM). Findings were validated in a second cohort of children with TBM and pulmonary TB (PTB), and functional T-cell responses studied in a third cohort of children with TBM, other extrapulmonary TB (EPTB) and PTB. The predominant RNA transcriptional response in children with TBM was decreased abundance of multiple genes, with 140/204 (68%) of all differentially regulated genes showing reduced abundance compared to healthy controls. Findings were validated in a second cohort with concordance of the direction of differential expression in both TBM (r2 = 0.78 p = 2x10-16) and PTB patients (r2 = 0.71 p = 2x10-16) when compared to a second group of healthy controls. Although the direction of expression of these significant genes was similar in the PTB patients, the magnitude of differential transcript abundance was less in PTB than in TBM. The majority of genes were involved in activation of leucocytes (p = 2.67E-11) and T-cell receptor signalling (p = 6.56E-07). Less abundant gene expression in immune cells was associated with a functional defect in T-cell proliferation that recovered after full TB treatment (p<0.0003). Multiple genes involved in T-cell activation show decreased abundance in children with acute TB, who also have impaired functional T-cell responses. Our data suggest that childhood TB is associated with an acquired immune defect, potentially resulting in failure to contain the pathogen. Elucidation of the mechanism causing the immune paresis may identify new treatment and prevention strategies.

  18. APOGEE Chemical Abundances of the Sagittarius Dwarf Galaxy System

    NASA Astrophysics Data System (ADS)

    Hasselquist, Sten; Shetrone, Matthew D.; Smith, Verne V.; Holtzman, Jon A.; McWilliam, Andrew; APOGEE Team

    2018-06-01

    The Apache Point Observatory Galactic Evolution Experiment provides the opportunity of measuring elemental abundances for C, N, O, Na, Mg, Al, Si, P, K, Ca, V, Cr, Mn, Fe, Co, and Ni in vast numbers of stars. We analyze the chemical-abundance patterns of these elements for 158 red giant stars belonging to the Sagittarius dwarf galaxy (Sgr). This is the largest sample of Sgr stars with detailed chemical abundances, and it is the first time that C, N, P, K, V, Cr, Co, and Ni have been studied at high resolution in this galaxy. We find that the Sgr stars with [Fe/H] > -0.8 are deficient in all elemental abundance ratios (expressed as [X/Fe]) relative to the Milky Way, suggesting that the Sgr stars observed today were formed from gas that was less enriched by Type II SNe than stars formed in the Milky Way. By examining the relative deficiencies of the hydrostatic (O, Na, Mg, and Al) and explosive (Si, P, K, and Mn) elements, our analysis supports the argument that previous generations of Sgr stars were formed with a top-light initial mass function, one lacking the most massive stars that would normally pollute the interstellar medium with the hydrostatic elements. We use a simple chemical-evolution model, flexCE, to further support our claim and conclude that recent stellar generations of Fornax and the Large Magellanic Cloud could also have formed according to a top-light initial mass function. We then exploit the unique chemical abundance patters of the Sgr core to trace stars belonging to the Sgr tidal streams elsewhere in the Milky Way.

  19. Listeria-vectored vaccine expressing the Mycobacterium tuberculosis 30 kDa major secretory protein via the constitutively active prfA* regulon boosts BCG efficacy against tuberculosis.

    PubMed

    Jia, Qingmei; Dillon, Barbara Jane; Masleša-Galić, Saša; Horwitz, Marcus A

    2017-06-19

    A potent vaccine against tuberculosis, one of the world's deadliest diseases, is needed to enhance the immunity of people worldwide, most of whom have been vaccinated with the partially effective BCG vaccine. Here we investigate novel live attenuated recombinant Listeria monocytogenes (rLm) vaccines expressing the Mycobacterium tuberculosis (Mtb) 30 kDa major secretory protein (r30/Ag85B) (rLm30) as heterologous booster vaccines in animals primed with BCG. Using three attenuated Lm vectors, rLm Δ actA (LmI), rLm Δ actA Δ inlB (LmII), and rLm Δ actA Δ inlB prfA * (LmIII), we constructed five rLm30 vaccine candidates expressing the r30 linked in-frame to the Lm Listeriolycin O signal sequence and driven by the hly promoter (h30) or linked in-frame to the ActA N-terminus and driven by the actA promoter (a30). All five rLm30 vaccines secreted r30 in broth and macrophages; while rLm expressing r30 via a constitutively active prfA * regulon (rLmIII/a30) expressed the greatest amount of r30 in broth culture, all five rLm vaccines expressed equivalent amounts of r30 in infected macrophages. In comparative studies, boosting BCG-immunized mice with rLmIII/a30 induced the strongest antigen-specific T-cell responses, including splenic and lung polyfunctional CD4+ T-cells expressing the three cytokines of interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α), and interleukin-2 (IL-2) ( P < 0.001) and splenic and lung CD8+ T-cells expressing IFN-γ ( P < 0.0001). In mice and guinea pigs, rLmIII/a30 and rLmI/h30 vaccines were generally more potent booster vaccines than r30 in adjuvant and a recombinant adenovirus vaccine expressing r30. In a setting in which BCG alone was highly immunoprotective, boosting mice with rLmIII/a30, the most potent of the vaccines, significantly enhanced protection against aerosolized Mtb ( P <0.01). Copyright © 2017 American Society for Microbiology.

  20. Distribution, abundance and habitat use of deep diving cetaceans in the North-East Atlantic

    NASA Astrophysics Data System (ADS)

    Rogan, Emer; Cañadas, Ana; Macleod, Kelly; Santos, M. Begoña; Mikkelsen, Bjarni; Uriarte, Ainhize; Van Canneyt, Olivier; Vázquez, José Antonio; Hammond, Philip S.

    2017-07-01

    In spite of their oceanic habitat, deep diving cetacean species have been found to be affected by anthropogenic activities, with potential population impacts of high intensity sounds generated by naval research and oil prospecting receiving the most attention. Improving the knowledge of the distribution and abundance of this poorly known group is an essential prerequisite to inform mitigation strategies seeking to minimize their spatial and temporal overlap with human activities. We provide for the first time abundance estimates for five deep diving cetacean species (sperm whale, long-finned pilot whale, northern bottlenose whale, Cuvier's beaked whale and Sowerby's beaked whale) using data from three dedicated cetacean sighting surveys that covered the oceanic and shelf waters of the North-East Atlantic. Density surface modelling was used to obtain model-based estimates of abundance and to explore the physical and biological characteristics of the habitat used by these species. Distribution of all species was found to be significantly related to depth, distance from the 2000m depth contour, the contour index (a measure of variability in the seabed) and sea surface temperature. Predicted distribution maps also suggest that there is little spatial overlap between these species. Our results represent the best abundance estimates for deep-diving whales in the North-East Atlantic, predict areas of high density during summer and constitute important baseline information to guide future risk assessments of human activities on these species, evaluate potential spatial and temporal trends and inform EU Directives and future conservation efforts.

  1. Evidence Supporting a Role for Constitutive Ghrelin Receptor Signaling in Fasting-Induced Hyperphagia in Male Mice.

    PubMed

    Fernandez, Gimena; Cabral, Agustina; Andreoli, María F; Labarthe, Alexandra; M'Kadmi, Céline; Ramos, Jorge G; Marie, Jacky; Fehrentz, Jean-Alain; Epelbaum, Jacques; Tolle, Virginie; Perello, Mario

    2018-02-01

    Ghrelin is a potent orexigenic peptide hormone that acts through the growth hormone secretagogue receptor (GHSR), a G protein-coupled receptor highly expressed in the hypothalamus. In vitro studies have shown that GHSR displays a high constitutive activity, whose physiological relevance is uncertain. As GHSR gene expression in the hypothalamus is known to increase in fasting conditions, we tested the hypothesis that constitutive GHSR activity at the hypothalamic level drives the fasting-induced hyperphagia. We found that refed wild-type (WT) mice displayed a robust hyperphagia that continued for 5 days after refeeding and changed their food intake daily pattern. Fasted WT mice showed an increase in plasma ghrelin levels, as well as in GHSR expression levels and ghrelin binding sites in the hypothalamic arcuate nucleus. When fasting-refeeding responses were evaluated in ghrelin- or GHSR-deficient mice, only the latter displayed an ∼15% smaller hyperphagia, compared with WT mice. Finally, fasting-induced hyperphagia of WT mice was significantly smaller in mice centrally treated with the GHSR inverse agonist K-(D-1-Nal)-FwLL-NH2, compared with mice treated with vehicle, whereas it was unaffected in mice centrally treated with the GHSR antagonists D-Lys3-growth hormone-releasing peptide 6 or JMV2959. Taken together, genetic models and pharmacological results support the notion that constitutive GHSR activity modulates the magnitude of the compensatory hyperphagia triggered by fasting. Thus, the hypothalamic GHSR signaling system could affect the set point of daily food intake, independently of plasma ghrelin levels, in situations of negative energy balance. Copyright © 2018 Endocrine Society.

  2. Cleanup liability and the Constitution

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Friedland, D.M.; Hagen, P.E.

    It was observed in the July 1992 issue of this Journal that a plain reading of the Constitution's prohibition on [open quotes][ital ex post facto][close quotes] suggests that some environmental statutes such as the Comprehensive Environmental Response, Compensation, and Liability Act of 1980 (CERCLA or Superfund) and the Resource Conservation and Recovery Act of 1976 (RCRA) conflict with Constitutional principles. Like many Constitutional principles, however, the Supreme Court's interpretation of the Constitution's bar on [ital ex post facto] laws has a long history. The Court has consistently interpreted this clause as limited to criminal or penal statutes. This article discussesmore » the history of the [ital ex post facto] clause, the retroactive application of CERCLA and RCRA, the decision that retroactive application of CERCLA and RCRA does not violate the [ital ex post facto] clause, and laws, regulations, and guidance. 27 refs.« less

  3. Housing system influences abundance of Pax3 and Pax7 in postnatal chicken skeletal muscles.

    PubMed

    Yin, H D; Li, D Y; Zhang, L; Yang, M Y; Zhao, X L; Wang, Y; Liu, Y P; Zhu, Q

    2014-06-01

    Paired box (Pax) proteins 3 and 7 are associated with activation of muscle satellite cells and play a major role in hyperplastic and hypertrophic growth in postnatal skeletal muscle fibers. The objective of this study was to evaluate the effect of housing system on abundance of Pax3 and Pax7 in postnatal chicken skeletal muscles. At 42 d, 1,200 chickens with similar BW were randomly assigned to cage, pen, and free-range group. The mRNA abundance was measured in pectoralis major and thigh muscle at d 56, 70, and 84, and the protein expression was quantified at d 84. Increases in mRNA abundance of PAX3 and PAX7 with age were less pronounced in caged system chickens than in pen and free-range chickens from d 56 to 84, and free-range chickens showed a more pronounced increase in gene expression with age compared with penned chickens. At d 84, quantities of PAX3 and PAX7 mRNA and protein were highest in both pectoralis major and thigh muscle of chickens raised in the free-range group, lowest in penned chickens, and intermediate in caged chickens (P < 0.05). These data indicate that housing system may influence muscle fiber muscle accretion by coordinating the expression of Pax3 and Pax7 in adult chicken skeletal muscles. Poultry Science Association Inc.

  4. Lack of in vitro constitutive activity for four previously reported TSH receptor mutations identified in patients with nonautoimmune hyperthyroidism and hot thyroid carcinomas.

    PubMed

    Jaeschke, Holger; Mueller, Sandra; Eszlinger, Markus; Paschke, Ralf

    2010-12-01

    Constitutively activating mutations (CAMs) of the TSHR are the major cause for nonautoimmune hyperthyroidism. Re-examination of constitutive activity previously determined in CHO cell lines recently demonstrated the caveats for the in vitro determination of constitutive TSHR activity, which leads to false positive conclusions regarding the molecular origin of hyperthyroidism or hot thyroid carcinomas. Mutations L677V and T620I identified in hot thyroid carcinomas were previously characterized in CHO and in 3T3-Vill cell lines, respectively, stably expressing the mutant without determination of TSHR expression. F666L identified in a patient with hot thyroid nodules, I691F in a family with nonautoimmune hyperthyroidism and F631I identified in a hot thyroid carcinoma were not characterized for their in vitro function. Therefore, we decided to (re)evaluate the in vitro function of these five TSHR variants by determination of cell surface expression, and intracellular cAMP and inositol phosphate levels and performed additionally linear regression analyses to determine basal activity independently from the mutant's cell surface expression in COS-7 and HEK(GT) cells. Only one (F631I) of the five investigated TSHR variants displayed constitutive activity for G(α) s signalling and showed correlation with the clinical phenotype. The previous false classification of T620I and L677V as CAMs is most likely related to the fact that both mutations were characterized in cell lines stably expressing the mutated receptor construct without assessing the respective receptor number per cell. Other molecular aetiologies for the nonautoimmune hyperthyroidism and/or hot thyroid carcinomas in these three patients and one family should be elucidated. © 2010 Blackwell Publishing Ltd.

  5. The impacts of land use change on malaria vector abundance in a water-limited, highland region of Ethiopia.

    PubMed

    Stryker, Jody J; Bomblies, Arne

    2012-12-01

    Changes in land use and climate are expected to alter the risk of malaria transmission in areas where rainfall limits vector abundance. We use a coupled hydrology-entomology model to investigate the effects of land use change on hydrological processes impacting mosquito abundance in a highland village of Ethiopia. Land use affects partitioning of rainfall into infiltration and runoff that reaches small-scale topographic depressions, which constitute the primary breeding habitat of Anopheles arabiensis mosquitoes. A physically based hydrology model isolates hydrological mechanisms by which land use impacts pool formation and persistence, and an agent-based entomology model evaluates the response of mosquito populations. This approach reproduced observed interannual variability in mosquito abundance between the 2009 and 2010 wet seasons. Several scenarios of land cover were then evaluated using the calibrated, field-validated model. Model results show variation in pool persistence and depth, as well as in mosquito abundance, due to land use changes alone. The model showed particular sensitivity to surface roughness, but also to root zone uptake. Scenarios in which land use was modified from agriculture to forest generally resulted in lowest mosquito abundance predictions; classification of the entire domain as rainforest produced a 34% decrease in abundance compared to 2010 results. This study also showed that in addition to vegetation type, spatial proximity of land use change to habitat locations has an impact on mosquito abundance. This modeling approach can be applied to assess impacts of climate and land use conditions that fall outside of the range of previously observed variability.

  6. The Impacts of Land Use Change on Malaria Vector Abundance in a Water-Limited Highland Region of Ethiopia

    NASA Astrophysics Data System (ADS)

    Stryker, J.; Bomblies, A.

    2012-12-01

    Changes in land use and climate are expected to alter risk of malaria transmission in areas where rainfall limits vector abundance. We use a coupled hydrology-entomology model to investigate the effects of land use change on hydrological processes impacting mosquito abundance in a highland village of Ethiopia. Land use affects partitioning of rainfall into infiltration and runoff that reaches small-scale topographic depressions, which constitute the primary breeding habitat of Anopheles arabiensis mosquitoes. A physically-based hydrology model isolates hydrological mechanisms by which land use impacts pool formation and persistence, and an agent-based entomology model evaluates the response of mosquito populations. This approach reproduced observed interannual variability in mosquito abundance between the 2009 and 2010 wet seasons. Several scenarios of land cover were then evaluated using the calibrated, field-validated model. Model results show variation in pool persistence and depth, as well as in mosquito abundance, due to land use changes alone. The model showed particular sensitivity to surface roughness, but also to root zone uptake. Scenarios in which land use was modified from agriculture to forest generally resulted in lowest mosquito abundance predictions; classification of the entire domain as rainforest produced a 34% decrease in abundance compared to 2010 results. This study also showed that in addition to vegetation type, spatial proximity of land use change to habitat locations has an impact on mosquito abundance. This modeling approach can be applied to assess impacts of climate and land use conditions that fall outside of the range of previously observed variability.

  7. Snake venoms are integrated systems, but abundant venom proteins evolve more rapidly.

    PubMed

    Aird, Steven D; Aggarwal, Shikha; Villar-Briones, Alejandro; Tin, Mandy Man-Ying; Terada, Kouki; Mikheyev, Alexander S

    2015-08-28

    While many studies have shown that extracellular proteins evolve rapidly, how selection acts on them remains poorly understood. We used snake venoms to understand the interaction between ecology, expression level, and evolutionary rate in secreted protein systems. Venomous snakes employ well-integrated systems of proteins and organic constituents to immobilize prey. Venoms are generally optimized to subdue preferred prey more effectively than non-prey, and many venom protein families manifest positive selection and rapid gene family diversification. Although previous studies have illuminated how individual venom protein families evolve, how selection acts on venoms as integrated systems, is unknown. Using next-generation transcriptome sequencing and mass spectrometry, we examined microevolution in two pitvipers, allopatrically separated for at least 1.6 million years, and their hybrids. Transcriptomes of parental species had generally similar compositions in regard to protein families, but for a given protein family, the homologs present and concentrations thereof sometimes differed dramatically. For instance, a phospholipase A2 transcript comprising 73.4 % of the Protobothrops elegans transcriptome, was barely present in the P. flavoviridis transcriptome (<0.05 %). Hybrids produced most proteins found in both parental venoms. Protein evolutionary rates were positively correlated with transcriptomic and proteomic abundances, and the most abundant proteins showed positive selection. This pattern holds with the addition of four other published crotaline transcriptomes, from two more genera, and also for the recently published king cobra genome, suggesting that rapid evolution of abundant proteins may be generally true for snake venoms. Looking more broadly at Protobothrops, we show that rapid evolution of the most abundant components is due to positive selection, suggesting an interplay between abundance and adaptation. Given log-scale differences in toxin

  8. AQP2 Abundance is Regulated by the E3-Ligase CHIP Via HSP70.

    PubMed

    Centrone, Mariangela; Ranieri, Marianna; Di Mise, Annarita; Berlingerio, Sante Princiero; Russo, Annamaria; Deen, Peter M T; Staub, Olivier; Valenti, Giovanna; Tamma, Grazia

    2017-01-01

    AQP2 expression is mainly controlled by vasopressin-dependent changes in protein abundance which is in turn regulated by AQP2 ubiquitylation and degradation, however the proteins involved in these processes are largely unknown. Here, we investigated the potential role of the CHIP E3 ligase in AQP2 regulation. MCD4 cells and kidney slices were used to study the involvement of the E3 ligase CHIP on AQP2 protein abundance by cell homogenization and immunoprecipitation followed by immunoblotting. We found that AQP2 complexes with CHIP in renal tissue. Expression of CHIP increased proteasomal degradation of AQP2 and HSP70 abundance, a molecular signature of HSP90 inhibition. Increased HSP70 level, secondary to CHIP expression, promoted ERK signaling resulting in increased AQP2 phosphorylation at S261. Phosphorylation of AQP2 at S256 and T269 were instead downregulated. Next, we investigated HSP70 interaction with AQP2, which is important for endocytosis. Compared with AQP2-wt, HSP70 binding decreased in AQP2-S256D and AQP2-S256D-S261D, while increased in AQP2-S256D-S261A. Surprisingly, expression of CHIP-delUbox, displaying a loss of E3 ligase activity, still induced AQP2 degradation, indicating that CHIP does not ubiquitylate and degrade AQP2 itself. Conversely, the AQP2 half-life was increased upon the expression of CHIP-delTPR a domain which binds Hsc70/HSP70 and HSP90. HSP70 has been reported to bind other E3 ligases such as MDM2. Notably, we found that co-expression of CHIP and MDM2 increased AQP2 degradation, whereas co-expression of CHIP with MDM2-delRING, an inactive form of MDM2, impaired AQP2 degradation. Our findings indicate CHIP as a master regulator of AQP2 degradation via HSP70 that has dual functions: (1) as chaperone for AQP2 and (2) as an anchoring protein for MDM2 E3 ligase, which is likely to be involved in AQP2 degradation. © 2017 The Author(s). Published by S. Karger AG, Basel.

  9. Expressing Constitutively Active Rheb in Adult Neurons after a Complete Spinal Cord Injury Enhances Axonal Regeneration beyond a Chondroitinase-Treated Glial Scar

    PubMed Central

    Wu, Di; Klaw, Michelle C.; Connors, Theresa; Kholodilov, Nikolai; Burke, Robert E.

    2015-01-01

    After a spinal cord injury (SCI), CNS axons fail to regenerate, resulting in permanent deficits. This is due to: (1) the presence of inhibitory molecules, e.g., chondroitin sulfate proteoglycans (CSPG), in the glial scar at the lesion; and (2) the diminished growth capacity of adult neurons. We sought to determine whether expressing a constitutively active form of the GTPase Rheb (caRheb) in adult neurons after a complete SCI in rats improves intrinsic growth potential to result in axon regeneration out of a growth-supportive peripheral nerve grafted (PNG) into the SCI cavity. We also hypothesized that treating the glial scar with chondroitinase ABC (ChABC), which digests CSPG, would further allow caRheb-transduced neurons to extend axons across the distal graft interface. We found that targeting this pathway at a clinically relevant post-SCI time point improves both sprouting and regeneration of axons. CaRheb increased the number of axons, but not the number of neurons, that projected into the PNG, indicative of augmented sprouting. We also saw that caRheb enhanced sprouting far rostral to the injury. CaRheb not only increased growth rostral and into the graft, it also resulted in significantly more regrowth of axons across a ChABC-treated scar into caudal spinal cord. CaRheb+ neurons had higher levels of growth-associated-43, suggestive of a newly identified mechanism for mTOR-mediated enhancement of regeneration. Thus, we demonstrate for the first time that simultaneously addressing intrinsic and scar-associated, extrinsic impediments to regeneration results in significant regrowth beyond an extremely challenging, complete SCI site. SIGNIFICANCE STATEMENT After spinal cord injury (SCI), CNS axons fail to regenerate, resulting in permanent deficits. This is due to the diminished growth capacity of adult neurons and the presence of inhibitory molecules in the scar at the lesion. We sought to simultaneously counter both of these obstacles to achieve more robust

  10. Expressing Constitutively Active Rheb in Adult Neurons after a Complete Spinal Cord Injury Enhances Axonal Regeneration beyond a Chondroitinase-Treated Glial Scar.

    PubMed

    Wu, Di; Klaw, Michelle C; Connors, Theresa; Kholodilov, Nikolai; Burke, Robert E; Tom, Veronica J

    2015-08-05

    After a spinal cord injury (SCI), CNS axons fail to regenerate, resulting in permanent deficits. This is due to: (1) the presence of inhibitory molecules, e.g., chondroitin sulfate proteoglycans (CSPG), in the glial scar at the lesion; and (2) the diminished growth capacity of adult neurons. We sought to determine whether expressing a constitutively active form of the GTPase Rheb (caRheb) in adult neurons after a complete SCI in rats improves intrinsic growth potential to result in axon regeneration out of a growth-supportive peripheral nerve grafted (PNG) into the SCI cavity. We also hypothesized that treating the glial scar with chondroitinase ABC (ChABC), which digests CSPG, would further allow caRheb-transduced neurons to extend axons across the distal graft interface. We found that targeting this pathway at a clinically relevant post-SCI time point improves both sprouting and regeneration of axons. CaRheb increased the number of axons, but not the number of neurons, that projected into the PNG, indicative of augmented sprouting. We also saw that caRheb enhanced sprouting far rostral to the injury. CaRheb not only increased growth rostral and into the graft, it also resulted in significantly more regrowth of axons across a ChABC-treated scar into caudal spinal cord. CaRheb(+) neurons had higher levels of growth-associated-43, suggestive of a newly identified mechanism for mTOR-mediated enhancement of regeneration. Thus, we demonstrate for the first time that simultaneously addressing intrinsic and scar-associated, extrinsic impediments to regeneration results in significant regrowth beyond an extremely challenging, complete SCI site. After spinal cord injury (SCI), CNS axons fail to regenerate, resulting in permanent deficits. This is due to the diminished growth capacity of adult neurons and the presence of inhibitory molecules in the scar at the lesion. We sought to simultaneously counter both of these obstacles to achieve more robust regeneration after

  11. Omega-3 fatty acids attenuate constitutive and insulin-induced CD36 expression through a suppression of PPAR α/γ activity in microvascular endothelial cells.

    PubMed

    Madonna, Rosalinda; Salerni, Sara; Schiavone, Deborah; Glatz, Jan F; Geng, Yong-Jian; De Caterina, Raffaele

    2011-09-01

    Microvascular dysfunction occurs in insulin resistance and/or hyperinsulinaemia. Enhanced uptake of free fatty acids (FFA) and oxidised low-density lipoproteins (oxLDL) may lead to oxidative stress and microvascular dysfunction interacting with CD36, a PPARα/γ-regulated scavenger receptor and long-chain FFA transporter. We investigated CD36 expression and CD36-mediated oxLDL uptake before and after insulin treatment in human dermal microvascular endothelial cells (HMVECs), ± different types of fatty acids (FA), including palmitic, oleic, linoleic, arachidonic, eicosapentaenoic (EPA), and docosahexaenoic (DHA) acids. Insulin (10(-8) and 10(-7) M) time-dependently increased DiI-oxLDL uptake and CD36 surface expression (by 30 ± 13%, p<0.05 vs. untreated control after 24 hours incubation), as assessed by ELISA and flow cytometry, an effect that was potentiated by the PI3-kinase inhibitor wortmannin and reverted by the ERK1/2 inhibitor PD98059 and the PPARα/γ antagonist GW9662. A ≥ 24 hour exposure to 50 μM DHA or EPA, but not other FA, blunted both the constitutive (by 23 ± 3% and 29 ± 2%, respectively, p<0.05 for both) and insulin-induced CD36 expressions (by 45 ± 27 % and 12 ± 3 %, respectively, p<0.05 for both), along with insulin-induced uptake of DiI-oxLDL and the downregulation of phosphorylated endothelial nitric oxide synthase (P-eNOS). At gel shift assays, DHA reverted insulin-induced basal and oxLDL-stimulated transactivation of PPRE and DNA binding of PPARα/γ and NF-κB. In conclusion, omega-3 fatty acids blunt the increased CD36 expression and activity promoted by high concentrations of insulin. Such mechanisms may be the basis for the use of omega-3 fatty acids in diabetic microvasculopathy.

  12. The solar photospheric abundance of hafnium and thorium. Results from CO5BOLD 3D hydrodynamic model atmospheres

    NASA Astrophysics Data System (ADS)

    Caffau, E.; Sbordone, L.; Ludwig, H.-G.; Bonifacio, P.; Steffen, M.; Behara, N. T.

    2008-05-01

    Context: The stable element hafnium (Hf) and the radioactive element thorium (Th) were recently suggested as a suitable pair for radioactive dating of stars. The applicability of this elemental pair needs to be established for stellar spectroscopy. Aims: We aim at a spectroscopic determination of the abundance of Hf and Th in the solar photosphere based on a CO5BOLD 3D hydrodynamical model atmosphere. We put this into a wider context by investigating 3D abundance corrections for a set of G- and F-type dwarfs. Methods: High-resolution, high signal-to-noise solar spectra were compared to line synthesis calculations performed on a solar CO5BOLD model. For the other atmospheres, we compared synthetic spectra of CO5BOLD 3D and associated 1D models. Results: For Hf we find a photospheric abundance A(Hf) = 0.87 ± 0.04, in good agreement with a previous analysis, based on 1D model atmospheres. The weak Th II 401.9 nm line constitutes the only Th abundance indicator available in the solar spectrum. It lies in the red wing of a Ni-Fe blend exhibiting a non-negligible convective asymmetry. Accounting for the asymmetry-related additional absorption, we obtain A(Th) = 0.08 ± 0.03, consistent with the meteoritic abundance, and about 0.1 dex lower than obtained in previous photospheric abundance determinations. Conclusions: Only for the second time, to our knowledge, has a non-negligible effect of convective line asymmetries on an abundance derivation been highlighted. Three-dimensional hydrodynamical simulations should be employed to measure Th abundances in dwarfs if similar blending is present, as in the solar case. In contrast, 3D effects on Hf abundances are small in G- to mid F-type dwarfs and sub-giants, and 1D model atmospheres can be conveniently used.

  13. NEON AND OXYGEN ABUNDANCES AND ABUNDANCE RATIO IN THE SOLAR CORONA

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Landi, E.; Testa, P., E-mail: elandi@umich.edu

    2015-02-20

    In this work we determine the Ne/O abundance ratio from Solar and Heliospheric Observatory (SOHO)/Solar Ultraviolet Measurement of Emitted Radiation (SUMER) off-disk observations of quiescent streamers over the 1996-2008 period. We find that the Ne/O ratio is approximately constant over solar cycle 23 from 1996 to 2005, at a value of 0.099 ± 0.017; this value is lower than the transition region determinations from the quiet Sun used to infer the neon photospheric abundance from the oxygen photospheric abundance. Also, the Ne/O ratio we determined from SUMER is in excellent agreement with in situ determinations from ACE/SWICS. In 2005-2008, the Ne/O abundancemore » ratio increased with time and reached 0.25 ± 0.05, following the same trend found in the slowest wind analyzed by ACE/SWICS. Further, we measure the absolute abundance in the corona for both oxygen and neon from the data set of 1996 November 22, obtaining A {sub o} = 8.99 ± 0.04 and A {sub Ne} = 7.92 ± 0.03, and we find that both elements are affected by the first ionization potential (FIP) effect, with oxygen being enhanced by a factor of 1.4-2.1 over its photospheric abundance, and neon being changed by a factor of 0.75-1.20. We conclude that the Ne/O ratio is not constant in the solar atmosphere, both in time and at different heights, and that it cannot be reliably used to infer the neon abundance in the photosphere. Also, we argue that the FIP effect was less effective during the minimum of solar cycle 24, and that the Ne/O = 0.25 ± 0.05 value measured at that time is closer to the true photospheric value, leading to a neon photospheric abundance larger than assumed by ≈40%. We discuss the implications of these results for the solar abundance problem, for the FIP effect, and for the identification of the source regions of the solar wind.« less

  14. Induced miR-99a expression represses Mtor cooperatively with miR-150 to promote regulatory T-cell differentiation

    PubMed Central

    Warth, Sebastian C; Hoefig, Kai P; Hiekel, Anian; Schallenberg, Sonja; Jovanovic, Ksenija; Klein, Ludger; Kretschmer, Karsten; Ansel, K Mark; Heissmeyer, Vigo

    2015-01-01

    Peripheral induction of regulatory T (Treg) cells provides essential protection from inappropriate immune responses. CD4+ T cells that lack endogenous miRNAs are impaired to differentiate into Treg cells, but the relevant miRNAs are unknown. We performed an overexpression screen with T-cell-expressed miRNAs in naive mouse CD4+ T cells undergoing Treg differentiation. Among 130 candidates, the screen identified 29 miRNAs with a negative and 10 miRNAs with a positive effect. Testing reciprocal Th17 differentiation revealed specific functions for miR-100, miR-99a and miR-10b, since all of these promoted the Treg and inhibited the Th17 program without impacting on viability, proliferation and activation. miR-99a cooperated with miR-150 to repress the expression of the Th17-promoting factor mTOR. The comparably low expression of miR-99a was strongly increased by the Treg cell inducer “retinoic acid”, and the abundantly expressed miR-150 could only repress Mtor in the presence of miR-99a. Our data suggest that induction of Treg cell differentiation is regulated by a miRNA network, which involves cooperation of constitutively expressed as well as inducible miRNAs. PMID:25712478

  15. Isolation, characterization, and evaluation of three Citrus sinensis-derived constitutive gene promoters.

    PubMed

    Erpen, L; Tavano, E C R; Harakava, R; Dutt, M; Grosser, J W; Piedade, S M S; Mendes, B M J; Mourão Filho, F A A

    2018-05-23

    Regulatory sequences from the citrus constitutive genes cyclophilin (CsCYP), glyceraldehyde-3-phosphate dehydrogenase C2 (CsGAPC2), and elongation factor 1-alpha (CsEF1) were isolated, fused to the uidA gene, and qualitatively and quantitatively evaluated in transgenic sweet orange plants. The 5' upstream region of a gene (the promoter) is the most important component for the initiation and regulation of gene transcription of both native genes and transgenes in plants. The isolation and characterization of gene regulatory sequences are essential to the development of intragenic or cisgenic genetic manipulation strategies, which imply the use of genetic material from the same species or from closely related species. We describe herein the isolation and evaluation of the promoter sequence from three constitutively expressed citrus genes: cyclophilin (CsCYP), glyceraldehyde-3-phosphate dehydrogenase C2 (CsGAPC2), and elongation factor 1-alpha (CsEF1). The functionality of the promoters was confirmed by a histochemical GUS assay in leaves, stems, and roots of stably transformed citrus plants expressing the promoter-uidA construct. Lower uidA mRNA levels were detected when the transgene was under the control of citrus promoters as compared to the expression under the control of the CaMV35S promoter. The association of the uidA gene with the citrus-derived promoters resulted in mRNA levels of up to 60-41.8% of the value obtained with the construct containing CaMV35S driving the uidA gene. Moreover, a lower inter-individual variability in transgene expression was observed amongst the different transgenic lines, where gene constructs containing citrus-derived promoters were used. In silico analysis of the citrus-derived promoter sequences revealed that their activity may be controlled by several putative cis-regulatory elements. These citrus promoters will expand the availability of regulatory sequences for driving gene expression in citrus gene-modification programs.

  16. Nonequilibrium electromagnetics: Local and macroscopic fields and constitutive relationships

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Baker-Jarvis, James; Kabos, Pavel; Holloway, Christopher L.

    We study the electrodynamics of materials using a Liouville-Hamiltonian-based statistical-mechanical theory. Our goal is to develop electrodynamics from an ensemble-average viewpoint that is valid for microscopic and nonequilibrium systems at molecular to submolecular scales. This approach is not based on a Taylor series expansion of the charge density to obtain the multipoles. Instead, expressions of the molecular multipoles are used in an inverse problem to obtain the averaging statistical-density function that is used to obtain the macroscopic fields. The advantages of this method are that the averaging function is constructed in a self-consistent manner and the molecules can either bemore » treated as point multipoles or contain more microstructure. Expressions for the local and macroscopic fields are obtained, and evolution equations for the constitutive parameters are developed. We derive equations for the local field as functions of the applied, polarization, magnetization, strain density, and macroscopic fields.« less

  17. Mining microarrays for metabolic meaning: nutritional regulation of hypothalamic gene expression.

    PubMed

    Mobbs, Charles V; Yen, Kelvin; Mastaitis, Jason; Nguyen, Ha; Watson, Elizabeth; Wurmbach, Elisa; Sealfon, Stuart C; Brooks, Andrew; Salton, Stephen R J

    2004-06-01

    DNA microarray analysis has been used to investigate relative changes in the level of gene expression in the CNS, including changes that are associated with disease, injury, psychiatric disorders, drug exposure or withdrawal, and memory formation. We have used oligonucleotide microarrays to identify hypothalamic genes that respond to nutritional manipulation. In addition to commonly used microarray analysis based on criteria such as fold-regulation, we have also found that simply carrying out multiple t tests then sorting by P value constitutes a highly reliable method to detect true regulation, as assessed by real-time polymerase chain reaction (PCR), even for relatively low abundance genes or relatively low magnitude of regulation. Such analyses directly suggested novel mechanisms that mediate effects of nutritional state on neuroendocrine function and are being used to identify regulated gene products that may elucidate the metabolic pathology of obese ob/ob, lean Vgf-/Vgf-, and other models with profound metabolic impairments.

  18. Cell Surface Expression of Bacterial Esterase A by Saccharomyces cerevisiae and Its Enhancement by Constitutive Activation of the Cellular Unfolded Protein Response▿ †

    PubMed Central

    Breinig, Frank; Diehl, Björn; Rau, Sabrina; Zimmer, Christian; Schwab, Helmut; Schmitt, Manfred J.

    2006-01-01

    Yeast cell surface display is a powerful tool for expression and immobilization of biocatalytically active proteins on a unicellular eukaryote. Here bacterial carboxylesterase EstA from Burkholderia gladioli was covalently anchored into the cell wall of Saccharomyces cerevisiae by in-frame fusion to the endogenous yeast proteins Kre1p, Cwp2p, and Flo1p. When p-nitrophenyl acetate was used as a substrate, the esterase specific activities of yeast expressing the protein fusions were 103 mU mg−1 protein for Kre1/EstA/Cwp2p and 72 mU mg−1 protein for Kre1/EstA/Flo1p. In vivo cell wall targeting was confirmed by esterase solubilization after laminarinase treatment and immunofluorescence microscopy. EstA expression resulted in cell wall-associated esterase activities of 2.72 U mg−1 protein for Kre1/EstA/Cwp2p and 1.27 U mg−1 protein for Kre1/EstA/Flo1p. Furthermore, esterase display on the yeast cell surface enabled the cells to effectively grow on the esterase-dependent carbon source glycerol triacetate (Triacetin). In the case of Kre1/EstA/Flo1p, in vivo maturation within the yeast secretory pathway and final incorporation into the wall were further enhanced when there was constitutive activation of the unfolded protein response pathway. Our results demonstrate that esterase cell surface display in yeast, which, as shown here, is remarkably more effective than EstA surface display in Escherichia coli, can be further optimized by activating the protein folding machinery in the eukaryotic secretion pathway. PMID:16980424

  19. Actinide abundances in ordinary chondrites

    NASA Technical Reports Server (NTRS)

    Hagee, B.; Bernatowicz, T. J.; Podosek, F. A.; Johnson, M. L.; Burnett, D. S.

    1990-01-01

    Measurements of actinide and light REE (LREE) abundances and of phosphate abundances in equilibrated ordinary chondrites were obtained and were used to define the Pu abundance in the solar system and to determine the degree of variation of actinide and LREE abundances. The results were also used to compare directly the Pu/U ratio with the earlier obtained ratio determined indirectly, as (Pu/Nd)x(Nd/U), assuming that Pu behaves chemically as a LREE. The data, combined with high-accuracy isotope-dilution data from the literature, show that the degree of gram-scale variability of the Th, U, and LREE abundances for equilibrated ordinary chondrites is a factor of 2-3 for absolute abundances and up to 50 percent for relative abundances. The observed variations are interpreted as reflecting the differences in the compositions and/or proportions of solar nebula components accreted to ordinary chondrite parent bodies.

  20. Essential Medicines in National Constitutions: Progress Since 2008.

    PubMed

    Katrina Perehudoff, S; Toebes, Brigit; Hogerzeil, Hans

    2016-06-01

    A constitutional guarantee of access to essential medicines has been identified as an important indicator of government commitment to the progressive realization of the right to the highest attainable standard of health. The objective of this study was to evaluate provisions on access to essential medicines in national constitutions, to identify comprehensive examples of constitutional text on medicines that can be used as a model for other countries, and to evaluate the evolution of constitutional medicines-related rights since 2008. Relevant articles were selected from an inventory of constitutional texts from WHO member states. References to states' legal obligations under international human rights law were evaluated. Twenty-two constitutions worldwide now oblige governments to protect and/or to fulfill accessibility of, availability of, and/or quality of medicines. Since 2008, state responsibilities to fulfill access to essential medicines have expanded in five constitutions, been maintained in four constitutions, and have regressed in one constitution. Government commitments to essential medicines are an important foundation of health system equity and are included increasingly in state constitutions.

  1. Remarks on turbulent constitutive relations

    NASA Technical Reports Server (NTRS)

    Shih, Tsan-Hsing; Lumley, John L.

    1993-01-01

    The paper demonstrates that the concept of turbulent constitutive relations can be used to construct general models for various turbulent correlations. Some of the Generalized Cayley-Hamilton formulas for relating tensor products of higher extension to tensor products of lower extension are introduced. The combination of dimensional analysis and invariant theory can lead to 'turbulent constitutive relations' (or general turbulence models) for, in principle, any turbulent correlations. As examples, the constitutive relations for Reynolds stresses and scalar fluxes are derived. The results are consistent with ones from Renormalization Group (RNG) theory and two-scale Direct-Interaction Approximation (DIA) method, but with a more general form.

  2. How Capitalistic Is the Constitution?

    ERIC Educational Resources Information Center

    Goldwin, Robert A., Ed.; Schambra, William A., Ed.

    Second in a three-part series designed to help prepare the nation for a thoughtful observance of the Constitutional bicentennial, this publication contains seven essays on the topic of capitalism and the Constitution. "American Democracy and the Acquisitive Spirit" (Marc F. Plattner) supports the argument that the framers of the…

  3. Oxygen abundances in halo stars

    NASA Astrophysics Data System (ADS)

    Bessell, Michael S.; Sutherland, Ralph S.; Ruan, Kui

    1991-12-01

    The present study determines the oxygen abundance for a sample of metal-poor G dwarfs by analysis of OH lines between 3080 and 3200 A and the permitted high-excitation far-red O I triple. The oxygen abundances determined from the low-excitation OH lines are up to 0.55 dex lower than those measured from the high-excitation O I lines. The abundances for the far-red O I triplet lines agree with those rederived from Abia and Rebolo (1989), and the abundances from the OH lines in dwarfs and giants are in agreement with the rederived O abundances of Barbuy (1988) and others from the forbidden resonance O I line. Because the chi = 0.1.7 eV OH lines are formed in the same layers as the majority of Fe, Ti, and other neutral metal lines used for abundance analyses, it is argued that the OH lines and the forbidden O I line yield the true oxygen abundances relative to the metals.

  4. Enhanced constitutive invasion activity in human nontumorigenic keratinocytes exposed to a low level of barium for a long time.

    PubMed

    Thang, Nguyen D; Yajima, Ichiro; Ohnuma, Shoko; Ohgami, Nobutaka; Kumasaka, Mayuko Y; Ichihara, Gaku; Kato, Masashi

    2015-02-01

    We have recently demonstrated that exposure to barium for a short time (≤4 days) and at a low level (5 µM = 687 µg/L) promotes invasion of human nontumorigenic HaCaT cells, which have characteristics similar to those of normal keratinocytes, suggesting that exposure to barium for a short time enhances malignant characteristics. Here we examined the effect of exposure to low level of barium for a long time, a condition mimicking the exposure to barium through well water, on malignant characteristics of HaCaT keratinocytes. Constitutive invasion activity, focal adhesion kinase (FAK) protein expression and activity, and matrix metalloproteinase 14 (MMP14) protein expression in primary cultured normal human epidermal keratinocytes, HaCaT keratinocytes, and HSC5 and A431 human squamous cell carcinoma cells were augmented following an increase in malignancy grade of the cells. Constitutive invasion activity, FAK phosphorylation, and MMP14 expression levels of HaCaT keratinocytes after treatment with 5 µM barium for 4 months were significantly higher than those of control untreated HaCaT keratinocytes. Taken together, our results suggest that exposure to a low level of barium for a long time enhances constitutive malignant characteristics of HaCaT keratinocytes via regulatory molecules (FAK and MMP14) for invasion. © 2013 Wiley Periodicals, Inc.

  5. The Constitution and Its Critics

    ERIC Educational Resources Information Center

    Main, Thomas J.

    2011-01-01

    In planning a freshman undergraduate curriculum with colleagues recently, the question arose as to what type of understanding educators wanted to impart to their students about the Constitution. The alleged defects of the Constitution that these books point to are wide-ranging and can be classified into various categories. Some problems--such as…

  6. The fish fauna of Anambra river basin, Nigeria: species abundance and morphometry.

    PubMed

    Odo, Gregory Ejikeme; Didigwu, Nwani Christopher; Eyo, Joseph Effiong

    2009-01-01

    The fish yields of most Nigeria inland waters are generally on the decline for causes that may range from inadequate management of the fisheries to degradation of the water bodies. Sustainable exploitation requires knowledge of the ichthyofaunal composition in the water bodies. We did a survey of fish species in Anambra river basin for 22 months. Fish samples were collected using four different gears -hook and line of size 13, caste nets, gill nets, and cages of mesh sizes of 50 mm, 75 mm, and 100 mm each. We recorded 52 fish species belonging to 17 families: 171, 236, and 169 individuals at Ogurugu, Otuocha, and Nsugbe stations respectively. Two families, Characidae, 19.5%, and Mochokidae, 11.8%, constituted the dominant fish families in the river. The dominant fish species were Citherinus citherius, 9.02%, and Alestes nurse, 7.1%. Other fish species with significant abundance were Synodontis clarias 6.9%, Macrolepidotus curvier 5.7%, Labeo coubie 5.4%, Distichodus rostrtus 4.9%, and Schilbe mystus 4.5%. The meristic features of the two most abundant fish species caught are as follows: Citharinus citharius dorsal fins 20, anal fins 30, caudal fins 21, pectoral fins, 9 and 8 ventral fins, and Alestes nurse 10 dorsal fins, 14 anal fins, 31 caudal fins, 7 pectoral fins and 6 ventral fins. The morphometric features of the two most abundant fish species are Citharinus citharius total length 300 mm, standard length 231 mm, head length 69 mm, body length 101 mm, body girth 176 mm, body weight 900 mg. Alestes nurse total length 200, standard length 140 mm, head length 60 mm, body length 80 mm, body girth 120 mm, body weight 400 mg. The most abundant animal utilizing the basin was Ardea cinerea (D3) with 22.2% occurrence (D4) and this was followed by Caprini with 13.51%, and Varanus niloticus, 10.04%. The least abundant animals utilizing basin were Chephalophus rufilatus, and Erythrocebus patas, with 0.58% each of occurrence.

  7. Quantitative Differential Expression Analysis Reveals Mir-7 As Major Islet MicroRNA

    PubMed Central

    Bravo-Egana, Valia; Rosero, Samuel; Molano, R. Damaris; Pileggi, Antonello; Ricordi, Camillo; Domínguez-Bendala, Juan; Pastori, Ricardo L.

    2008-01-01

    MicroRNAs (miRNAs) are non-coding gene products that regulate gene expression through specific binding to target mRNAs. Cell-specific patterns of miRNAs are associated with the acquisition and maintenance of a given phenotype, such as endocrine pancreas (islets). We hypothesized that a subset of miRNAs could be differentially expressed in the islets. Using miRNA microarray technology and quantitative RT-PCR we identified a subset of miRNAs that are the most differentially expressed islet miRNAs (ratio islet/acinar >150-fold), mir-7 being the most abundant. A similarly high ratio for mir-7 was observed in human islets. The ratio islet/acinar for mir-375, a previously described islet miRNA, was <10, and is 2.5X more abundant in the islets than mir-7. Therefore, we conclude that mir-7 is the most abundant endocrine miRNA in islets while mir-375 is the most abundant intra-islet miRNA. Our results may offer new insights into regulatory pathways of islet gene expression. PMID:18086561

  8. Expression and Functional Activities of Selected Sulfotransferase Isoforms in BeWo Cells and Primary Cytotrophoblast Cells

    PubMed Central

    Mitra, Pallabi; Audus, Kenneth L.

    2009-01-01

    Several cytosolic sulfotransferase enzyme isoforms are functional in placenta but there is limited information available on the utility of cultured trophoblast cells for studying sulfation. The trophoblast cell layer constitutes the rate-determining barrier for trans-placental transfer. The objective of this work was to examine the mRNA expression and enzyme activities of four sulfotransferase isoforms reported to be functional in human placenta (SULT1A1, SULT1A3, SULT1E1, and SULT2A1) in primary cytotrophoblast cells and the trophoblast-like BeWo cell line. Reverse transcription polymerase chain reaction (RT-PCR) was performed to determine mRNA expression. Enzyme activities were assessed using the following substrates: 4-nitrophenol for SULT1A1, dopamine for SULT1A3, 17β-estradiol for SULT1E1, and dehydroepiandrosterone for SULT2A1. For 4-nitrophenol and dopamine sulfation, apparent Km values, response to inhibitors (2,6-dichloro-4-nitrophenol and sodium chloride), and thermal stability profiles indicated that 4-nitrophenol and dopamine sulfation in BeWo cells were being mediated by SULT1A1 and SULT1A3, respectively. SULT1A1 and SULT1A3 were also functional in the cytotrophoblast cells. Both at the protein and at the mRNA levels, SULT1A1 was more abundant in BeWo cells in comparison to the primary cytotrophoblast cells. SULT1E1 and SULT2A1 mRNA were not detected in the cytotrophoblasts. SULT1E1 mRNA was weakly expressed in BeWo but there was negligible functional activity. Although SULT2A1 mRNA was abundantly expressed in BeWo, Western blot and enzyme activities revealed that the protein is not expressed in BeWo cells. The results suggest that the BeWo cells and the cytotrophoblast cells can be used to examine the roles of SULT1A1 and SULT1A3 in placental metabolism. PMID:19646966

  9. The Constitution and Citizenship Education.

    ERIC Educational Resources Information Center

    Shoemaker, Rebecca S.

    The paper takes the position that the study and understanding of the United States Constitution should be a critical part of citizenship education, especially as its Bicentennial approaches. Several factors suggest that the Constitution has become the most durable document of its kind in history, and that its teaching should be centered in both…

  10. Differential display of abundantly expressed genes of Trichoderma harzianum during colonization of tomato-germinating seeds and roots.

    PubMed

    Mehrabi-Koushki, Mehdi; Rouhani, Hamid; Mahdikhani-Moghaddam, Esmat

    2012-11-01

    The identification of Trichoderma genes whose expression is altered during early stages of interaction with developing roots of germinated seeds is an important step toward understanding the rhizosphere competency of Trichoderma spp. The potential of 13 Trichoderma strains to colonize tomato root and promote plant growth has been evaluated. All used strains successfully propagated in spermosphere and continued their growth in rhizoplane simultaneously root enlargement while the strains T6 and T7 were the most abundant in the apical segment of roots. Root colonization in most strains associated with promoting the roots and shoots growth while they significantly increased up to 43 and 40 % roots and shoots dry weights, respectively. Differential display reverse transcriptase-PCR (DDRT-PCR) has been developed to detect differentially expressed genes in the previously selected strain, Trichoderma harzianum T7, during colonization stages of tomato-germinating seeds and roots. Amplified DDRT-PCR products were analyzed on gel agarose and 62 differential bands excised, purified, cloned, and sequenced. Obtained ESTs were submit-queried to NCBI database by BLASTx search and gene ontology hierarchy. Most of transcripts (29 EST) corresponds to known and hypothetical proteins such as secretion-related small GTPase, 40S ribosomal protein S3a, 3-hydroxybutyryl-CoA dehydrogenase, DNA repair protein rad50, lipid phosphate phosphatase-related protein type 3, nuclear essential protein, phospholipase A2, fatty acid desaturase, nuclear pore complex subunit Nup133, ubiquitin-activating enzyme, and 60S ribosomal protein L40. Also, 13 of these sequences showed no homology (E > 0.05) with public databases and considered as novel genes. Some of these ESTs corresponded to genes encodes enzymes potentially involved in nutritional support of microorganisms which have obvious importance in the establishment of Trichoderma in spermosphere and rhizosphere, via potentially functioning in

  11. Stellar Oxygen Abundances

    NASA Astrophysics Data System (ADS)

    King, Jeremy

    1994-04-01

    This dissertation addresses several issues concerning stellar oxygen abundances. The 7774 {\\AA} O I triplet equivalent widths of Abia & Rebolo [1989, AJ, 347, 186] for metal-poor dwarfs are found to be systematically too high. I also argue that current effective temperatures used in halo star abundance studies may be ~150 K too low. New color-Teff relations are derived for metal-poor stars. Using the revised Teff values and improved equivalent widths for the 7774A O I triplet, the mean [O/Fe] ratio for a handful of halo stars is found to be +0.52 with no dependence on Teff or [Fe/H]. Possible cosmological implications of the hotter Teff scale are discussed along with additional evidence supporting the need for a higher temperature scale for metal-poor stars. Our Teff scale leads to a Spite Li plateau value of N(Li)=2.28 +/- 0.09. A conservative minimal primordial value of N(Li)=2.35 is inferred. If errors in the observations and models are considered, consistency with standard models of Big Bang nucleosynthesis is still achieved with this larger Li abundance. The revised Teff scale raises the observed B/Be ratio of HD 140283 from 10 to 12, making its value more comfortably consistent with the production of the observed B and Be by ordinary spallation. Our Teff values are found to be in good agreement with values predicted from both the Victoria and Yale isochrone color-Teff relations. Thus, it appears likely that no changes in globular cluster ages would result. Next, we examine the location of the break in the [O/Fe] versus [Fe/H] plane in a quantitative fashion. Analysis of a relatively homogeneous data set does not favor any unique break point in the range -1.7 /= -3), in agreement with the new results for halo dwarfs. We find that the gap in the observed [O/H] distribution, noted by Wheeler et al

  12. State Constitutional Commitment to Health and Health Care and Population Health Outcomes: Evidence From Historical US Data

    PubMed Central

    2015-01-01

    Objectives. I investigated whether the introduction of health and health care provisions in US state constitutions can make health systems more equitable and improve health outcomes by urging state policymakers and administrative agencies to uphold their human rights obligations at state level. Methods. I constructed a panel of infant mortality rates from 50 US states over the period 1929 through 2000 to examine their association with the timing and details of introducing a constitutional right to health and health care provisions. Results. The introduction of a stronger constitutional commitment that obligates state legislature to provide health care was associated with a subsequent reduction in the infant mortality rate of approximately 7.8%. The introduction of provisions explicitly targeting the poor was also associated with a reduction in the infant mortality rate of 6.5%. These health benefits are primarily evident in non-White populations. Conclusions. This empirical result supports Elizabeth Leonard’s view that although state constitutional rights have been poorly enforced through the judiciary, a constitutional expression of health care duties has fueled the political and social process, ultimately allowing states to identify the best way to address citizens’ health inequality concerns. PMID:25905857

  13. State constitutional commitment to health and health care and population health outcomes: evidence from historical US data.

    PubMed

    Matsuura, Hiroaki

    2015-07-01

    I investigated whether the introduction of health and health care provisions in US state constitutions can make health systems more equitable and improve health outcomes by urging state policymakers and administrative agencies to uphold their human rights obligations at state level. I constructed a panel of infant mortality rates from 50 US states over the period 1929 through 2000 to examine their association with the timing and details of introducing a constitutional right to health and health care provisions. The introduction of a stronger constitutional commitment that obligates state legislature to provide health care was associated with a subsequent reduction in the infant mortality rate of approximately 7.8%. The introduction of provisions explicitly targeting the poor was also associated with a reduction in the infant mortality rate of 6.5%. These health benefits are primarily evident in non-White populations. This empirical result supports Elizabeth Leonard's view that although state constitutional rights have been poorly enforced through the judiciary, a constitutional expression of health care duties has fueled the political and social process, ultimately allowing states to identify the best way to address citizens' health inequality concerns.

  14. Constitutional Conservatism

    ERIC Educational Resources Information Center

    Berkowitz, Peter

    2009-01-01

    After their dismal performance in election 2008, conservatives are taking stock. As they examine the causes that have driven them into the political wilderness and as they explore paths out, they should also take heart. After all, election 2008 shows that America's constitutional order is working as designed. Indeed, while sorting out their errors…

  15. Discrete False-Discovery Rate Improves Identification of Differentially Abundant Microbes.

    PubMed

    Jiang, Lingjing; Amir, Amnon; Morton, James T; Heller, Ruth; Arias-Castro, Ery; Knight, Rob

    2017-01-01

    Differential abundance testing is a critical task in microbiome studies that is complicated by the sparsity of data matrices. Here we adapt for microbiome studies a solution from the field of gene expression analysis to produce a new method, discrete false-discovery rate (DS-FDR), that greatly improves the power to detect differential taxa by exploiting the discreteness of the data. Additionally, DS-FDR is relatively robust to the number of noninformative features, and thus removes the problem of filtering taxonomy tables by an arbitrary abundance threshold. We show by using a combination of simulations and reanalysis of nine real-world microbiome data sets that this new method outperforms existing methods at the differential abundance testing task, producing a false-discovery rate that is up to threefold more accurate, and halves the number of samples required to find a given difference (thus increasing the efficiency of microbiome experiments considerably). We therefore expect DS-FDR to be widely applied in microbiome studies. IMPORTANCE DS-FDR can achieve higher statistical power to detect significant findings in sparse and noisy microbiome data compared to the commonly used Benjamini-Hochberg procedure and other FDR-controlling procedures.

  16. Closing in on the constitution of consciousness

    PubMed Central

    Miller, Steven M.

    2014-01-01

    The science of consciousness is a nascent and thriving field of research that is founded on identifying the minimally sufficient neural correlates of consciousness. However, I have argued that it is the neural constitution of consciousness that science seeks to understand and that there are no evident strategies for distinguishing the correlates and constitution of (phenomenal) consciousness. Here I review this correlation/constitution distinction problem and challenge the existing foundations of consciousness science. I present the main analyses from a longer paper in press on this issue, focusing on recording, inhibition, stimulation, and combined inhibition/stimulation strategies, including proposal of the Jenga analogy to illustrate why identifying the minimally sufficient neural correlates of consciousness should not be considered the ultimate target of consciousness science. Thereafter I suggest that while combined inhibition and stimulation strategies might identify some constitutive neural activities—indeed minimally sufficient constitutive neural activities—such strategies fail to identify the whole neural constitution of consciousness and thus the correlation/constitution distinction problem is not fully solved. Various clarifications, potential objections and related scientific and philosophical issues are also discussed and I conclude by proposing new foundational claims for consciousness science. PMID:25452738

  17. Abundance estimation and conservation biology

    USGS Publications Warehouse

    Nichols, J.D.; MacKenzie, D.I.

    2004-01-01

    Abundance is the state variable of interest in most population–level ecological research and in most programs involving management and conservation of animal populations. Abundance is the single parameter of interest in capture–recapture models for closed populations (e.g., Darroch, 1958; Otis et al., 1978; Chao, 2001). The initial capture–recapture models developed for partially (Darroch, 1959) and completely (Jolly, 1965; Seber, 1965) open populations represented efforts to relax the restrictive assumption of population closure for the purpose of estimating abundance. Subsequent emphases in capture–recapture work were on survival rate estimation in the 1970’s and 1980’s (e.g., Burnham et al., 1987; Lebreton et al.,1992), and on movement estimation in the 1990’s (Brownie et al., 1993; Schwarz et al., 1993). However, from the mid–1990’s until the present time, capture–recapture investigators have expressed a renewed interest in abundance and related parameters (Pradel, 1996; Schwarz & Arnason, 1996; Schwarz, 2001). The focus of this session was abundance, and presentations covered topics ranging from estimation of abundance and rate of change in abundance, to inferences about the demographic processes underlying changes in abundance, to occupancy as a surrogate of abundance. The plenary paper by Link & Barker (2004) is provocative and very interesting, and it contains a number of important messages and suggestions. Link & Barker (2004) emphasize that the increasing complexity of capture–recapture models has resulted in large numbers of parameters and that a challenge to ecologists is to extract ecological signals from this complexity. They offer hierarchical models as a natural approach to inference in which traditional parameters are viewed as realizations of stochastic processes. These processes are governed by hyperparameters, and the inferential approach focuses on these hyperparameters. Link & Barker (2004) also suggest that our attention

  18. Increased skin barrier disruption by sodium lauryl sulfate in mice expressing a constitutively active STAT6 in T cells.

    PubMed

    DaSilva, Sonia C; Sahu, Ravi P; Konger, Raymond L; Perkins, Susan M; Kaplan, Mark H; Travers, Jeffrey B

    2012-01-01

    Atopic dermatitis (AD) is a pruritic, chronic inflammatory skin disease that affects 10-20% of children and 1-3% of adults worldwide. Recent studies have indicated that the ability of Th2 cytokines, such as interleukin-4 (IL-4) to regulate skin barrier function may be a predisposing factor for AD development. The present studies examined the ability of increased Th2 activity to affect cutaneous barrier function in vivo and epidermal thickening. Mice that express a constitutively active Signal Transducer and Activator of Transcription 6 (STAT6VT) have increased Th2 cells and a predisposition to allergic inflammation were used in these studies, they demonstrate that topical treatment with the irritant sodium lauryl sulfate (SLS) caused increased transepidermal water loss and epidermal thickening in STAT6VT mice over similarly treated wild-type mice. The proliferation marker Ki-67 was increased in the epidermis of STAT6VT compared to the wild-type mice. However, these differences do not appear to be linked to the addition of an irritant as control-treated STAT6VT skin also exhibited elevated Ki-67 levels, suggesting that the increased epidermal thickness in SLS-treated STAT6VT mice is primarily driven by epidermal cell hypertrophy rather than an increase in cellular proliferation. Our results suggest that an environment with increased Th2 cytokines results in abnormal responses to topical irritants.

  19. Constitutive expression of nitrate reductase allows normal growth and development of Nicotiana plumbaginifolia plants.

    PubMed Central

    Vincentz, M; Caboche, M

    1991-01-01

    A nitrate reductase (NR) deficient mutant of Nicotiana plumbaginifolia totally impaired in the production of NR transcript and protein was restored for NR activity by transformation with a chimaeric NR gene. This gene was composed of a full-length tobacco NR cDNA fused to the CaMV 35S promoter and to termination signals from the tobacco NR gene. The transgenic plants we obtained were viable and fertile and expressed from one-fifth to three times the wild-type NR activity in their leaves. The analysis of chimeric NR gene expression in these plants showed, by comparison with wild-type plants, that the regulation of NR gene expression by light, nitrate and circadian rhythm takes place at the transcriptional level. However, unlike nitrate, light was required for the accumulation of NR protein in transgenic plants, suggesting that NR expression is also controlled at the translational and/or post-translational level. Images PMID:2022181

  20. Altered synaptic marker abundance in the hippocampal stratum oriens of Ts65Dn mice is associated with exuberant expression of versican

    PubMed Central

    Howell, Matthew D; Gottschall, Paul E

    2012-01-01

    DS (Down syndrome), resulting from trisomy of chromosome 21, is the most common cause of genetic mental retardation; however, the molecular mechanisms underlying the cognitive deficits are poorly understood. Growing data indicate that changes in abundance or type of CSPGs (chondroitin sulfate proteoglycans) in the ECM (extracellular matrix) can influence synaptic structure and plasticity. The purpose of this study was to identify changes in synaptic structure in the hippocampus in a model of DS, the Ts65Dn mouse, and to determine the relationship to proteoglycan abundance and/or cleavage and cognitive disability. We measured synaptic proteins by ELISA and changes in lectican expression and processing in the hippocampus of young and old Ts65Dn mice and LMCs (littermate controls). In young (5 months old) Ts65Dn hippocampal extracts, we found a significant increase in the postsynaptic protein PSD-95 (postsynaptic density 95) compared with LMCs. In aged (20 months old) Ts65Dn hippocampus, this increase was localized to hippocampal stratum oriens extracts compared with LMCs. Aged Ts65Dn mice exhibited impaired hippocampal-dependent spatial learning and memory in the RAWM (radial-arm water maze) and a marked increase in levels of the lectican versican V2 in stratum oriens that correlated with the number of errors made in the final RAWM block. Ts65Dn stratum oriens PNNs (perineuronal nets), an extension of the ECM enveloping mostly inhibitory interneurons, were dispersed over a larger area compared with LMC mice. Taken together, these data suggest a possible association with alterations in the ECM and inhibitory neurotransmission in the Ts65Dn hippocampus which could contribute to cognitive deficits. PMID:22225533

  1. IAOseq: inferring abundance of overlapping genes using RNA-seq data.

    PubMed

    Sun, Hong; Yang, Shuang; Tun, Liangliang; Li, Yixue

    2015-01-01

    Overlapping transcription constitutes a common mechanism for regulating gene expression. A major limitation of the overlapping transcription assays is the lack of high throughput expression data. We developed a new tool (IAOseq) that is based on reads distributions along the transcribed regions to identify the expression levels of overlapping genes from standard RNA-seq data. Compared with five commonly used quantification methods, IAOseq showed better performance in the estimation accuracy of overlapping transcription levels. For the same strand overlapping transcription, currently existing high-throughput methods are rarely available to distinguish which strand was present in the original mRNA template. The IAOseq results showed that the commonly used methods gave an average of 1.6 fold overestimation of the expression levels of same strand overlapping genes. This work provides a useful tool for mining overlapping transcription levels from standard RNA-seq libraries. IAOseq could be used to help us understand the complex regulatory mechanism mediated by overlapping transcripts. IAOseq is freely available at http://lifecenter.sgst.cn/main/en/IAO_seq.jsp.

  2. The Late Embryogenesis Abundant Protein Family in Cassava (Manihot esculenta Crantz): Genome-Wide Characterization and Expression during Abiotic Stress.

    PubMed

    Wu, Chunlai; Hu, Wei; Yan, Yan; Tie, Weiwei; Ding, Zehong; Guo, Jianchun; He, Guangyuan

    2018-05-17

    Late embryogenesis abundant (LEA) proteins, as a highly diverse group of polypeptides, play an important role in plant adaptation to abiotic stress; however, LEAs from cassava have not been studied in cassava. In this study, 26 LEA members were genome-wide identified from cassava, which were clustered into seven subfamily according to evolutionary relationship, protein motif, and gene structure analyses. Chromosomal location and duplication event analyses suggested that 26 MeLEAs distributed in 10 chromosomes and 11 MeLEA paralogues were subjected to purifying selection. Transcriptomic analysis showed the expression profiles of MeLEAs in different tissues of stem, leaves, and storage roots of three accessions. Comparative transcriptomic analysis revealed that the function of MeLEAs in response to drought may be differentiated in different accessions. Compared with the wild subspecies W14, more MeLEA genes were activated in cultivated varieties Arg7 and SC124 after drought treatment. Several MeLEA genes showed induction under various stresses and related signaling treatments. Taken together, this study demonstrates the transcriptional control of MeLEAs in tissue development and the responses to abiotic stress in cassava and identifies candidate genes for improving crop resistance to abiotic stress.

  3. Theoretical Issues of the Constitutional Regulation Mechanism

    ERIC Educational Resources Information Center

    Zhussupova, Guldaray B.; Zhailyaubayev, Rassul T.; Ukin, Symbat K.; Shunayeva, Sylu M.; Nurmagambetov, Rachit G.

    2016-01-01

    The purpose of this research is to define the concept of "constitutional regulation mechanism." The definition of the concept of "constitutional regulation mechanism" will give jurists and legislators a theoretical framework for developing legal sciences, such as the constitutional law and the theory of state and law. The…

  4. Constitutive expression of a salinity-induced wheat WRKY transcription factor enhances salinity and ionic stress tolerance in transgenic Arabidopsis thaliana.

    PubMed

    Qin, Yuxiang; Tian, Yanchen; Han, Lu; Yang, Xinchao

    2013-10-25

    The isolation and characterization of TaWRKY79, a wheat class II WRKY transcription factor, is described. Its 1297 bp coding region includes a 987 bp long open reading frame. TaWRKY79 was induced by stressing seedlings with either NaCl or abscisic acid (ABA). When a fusion between an 843 bp segment upstream of the TaWRKY79 coding sequence and GUS was introduced into Arabidopsis thaliana, GUS staining indicated that this upstream segment captured the sequence(s) required to respond to ABA or NaCl treatment. When TaWRKY79 was constitutively expressed as a transgene in A. thaliana, the transgenic plants showed an improved capacity to extend their primary root in the presence of either 100 mM NaCl, 10 mM LiCl or 2 μM ABA. The inference was that TaWRKY79 enhanced the level of tolerance to both salinity and ionic stress, while reducing the level of sensitivity to ABA. The ABA-related genes ABA1, ABA2 ABI1 and ABI5 were all up-regulated in the TaWRKY79 transgenic plants, suggesting that the transcription factor operates in an ABA-dependent pathway. Copyright © 2013. Published by Elsevier Inc.

  5. 75 FR 57835 - Constitution Day and Citizenship Day, Constitution Week, 2010

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-09-22

    ... vision of freedom and justice for all. In remembrance of the signing of the Constitution and in... leaders of civic, social, and educational organizations, to conduct ceremonies and programs that recognize...

  6. Targeting chromatin binding regulation of constitutively active AR variants to overcome prostate cancer resistance to endocrine-based therapies

    PubMed Central

    Chan, Siu Chiu; Selth, Luke A.; Li, Yingming; Nyquist, Michael D.; Miao, Lu; Bradner, James E.; Raj, Ganesh V.; Tilley, Wayne D.; Dehm, Scott M.

    2015-01-01

    Androgen receptor (AR) variants (AR-Vs) expressed in prostate cancer (PCa) lack the AR ligand binding domain (LBD) and function as constitutively active transcription factors. AR-V expression in patient tissues or circulating tumor cells is associated with resistance to AR-targeting endocrine therapies and poor outcomes. Here, we investigated the mechanisms governing chromatin binding of AR-Vs with the goal of identifying therapeutic vulnerabilities. By chromatin immunoprecipitation and sequencing (ChIP-seq) and complementary biochemical experiments, we show that AR-Vs display a binding preference for the same canonical high-affinity androgen response elements (AREs) that are preferentially engaged by AR, albeit with lower affinity. Dimerization was an absolute requirement for constitutive AR-V DNA binding and transcriptional activation. Treatment with the bromodomain and extraterminal (BET) inhibitor JQ1 resulted in inhibition of AR-V chromatin binding and impaired AR-V driven PCa cell growth in vitro and in vivo. Importantly, this was associated with a novel JQ1 action of down-regulating AR-V transcript and protein expression. Overall, this study demonstrates that AR-Vs broadly restore AR chromatin binding events that are otherwise suppressed during endocrine therapy, and provides pre-clinical rationale for BET inhibition as a strategy for inhibiting expression and chromatin binding of AR-Vs in PCa. PMID:25908785

  7. OXYGEN ABUNDANCES IN CEPHEIDS

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Luck, R. E.; Andrievsky, S. M.; Korotin, S. N.

    2013-07-01

    Oxygen abundances in later-type stars, and intermediate-mass stars in particular, are usually determined from the [O I] line at 630.0 nm, and to a lesser extent, from the O I triplet at 615.7 nm. The near-IR triplets at 777.4 nm and 844.6 nm are strong in these stars and generally do not suffer from severe blending with other species. However, these latter two triplets suffer from strong non-local thermodynamic equilibrium (NLTE) effects and thus see limited use in abundance analyses. In this paper, we derive oxygen abundances in a large sample of Cepheids using the near-IR triplets from an NLTEmore » analysis, and compare those abundances to values derived from a local thermodynamic equilibrium (LTE) analysis of the [O I] 630.0 nm line and the O I 615.7 nm triplet as well as LTE abundances for the 777.4 nm triplet. All of these lines suffer from line strength problems making them sensitive to either measurement complications (weak lines) or to line saturation difficulties (strong lines). Upon this realization, the LTE results for the [O I] lines and the O I 615.7 nm triplet are in adequate agreement with the abundance from the NLTE analysis of the near-IR triplets.« less

  8. [Body composition and constitution: a constitutional syndrome (1st of 2 parts)].

    PubMed

    Terán Díaz, E

    1999-04-01

    Constitutional syndrome alters body constitution modifying (usually decreasing) two of its dimensions--weight and perimeters--by changing the composition of one, several or every body levels. Apart of the cause, the basic physiopathological process that characterizes this new syndrome is the amino acid mobilization from the muscle (proteolysis). As soon as fat loss has no consequence to the organism, proteolysis reduces the muscle mass and life is in danger. Actually, there is no effective treatment to improve the nitrogen balance by medication or hormones in speed catabolic states but it can also approach us to more proper therapeutics for these so frequent processes in clinic.

  9. Rumex acetosa Y chromosomes: constitutive or facultative heterochromatin?

    PubMed

    Mosiołek, Magdalena; Pasierbek, Paweł; Malarz, Janusz; Moś, Maria; Joachimiak, Andrzej J

    2005-01-01

    Condensed Y chromosomes in Rumex acetosa L. root-tip nuclei were studied using 5-azaC treatment and immunohistochemical detection of methylated histones. Although Y chromosomes were decondensed within root meristem in vivo, they became condensed and heteropycnotic in roots cultured in vitro. 5-azacytidine (5-azaC) treatment of cultured roots caused transitional dispersion of their Y chromosome bodies, but 7 days after removal of the drug from the culture medium, Y heterochromatin recondensed and again became visible. The response of Rumex sex chromatin to 5-azaC was compared with that of condensed segments of pericentromeric heterochromatin in Rhoeo spathacea (Sw.) Steam roots. It was shown that Rhoeo chromocentres, composed of AT-rich constitutive heterochromatin, did not undergo decondensation after 5-azaC treatment. The Y-bodies observed within male nuclei of R. acetosa were globally enriched with H3 histone, demethylated at lysine 4 and methylated at lysine 9. This is the first report of histone tail-modification in condensed sex chromatin in plants. Our results suggest that the interphase condensation of Y chromosomes in Rumex is facultative rather than constitutive. Furthermore, the observed response of Y-bodies to 5-azaC may result indirectly from demethylation and the subsequent altered expression of unknown genes controlling tissue-specific Y-inactivation as opposed to the global demethylation of Y-chromosome DNA.

  10. Power: Constitutional Update. Bar/School Partnership Programs Series.

    ERIC Educational Resources Information Center

    American Bar Association, Chicago, IL. Special Committee on Youth Education for Citizenship.

    The fourth in a special series of handbooks dealing with constitutional themes, this document looks at power in the context of the U.S. Constitution. "The Constitution's Prescription for Freedom" (L. Peach) examines the separation of powers provided for in the Constitution. "The Concept of Power" (C. Roach) is a series of…

  11. Ordered Rate Constitutive Theories: Development of Rate Constitutive Equations for Solids, Liquids, and Gases

    DTIC Science & Technology

    2010-08-18

    the author(s) and should not contrued as an official Department of the Army position, policy or decision, unless so designated by other documentation...Daniel S. Nunez 0.50 Yong-Ting Ma 0.50 Tristan Moody 0.50 1.50FTE Equivalent: 3Total Number: Names of Post Doctorates PERCENT_SUPPORTEDNAME FTE...demonstrated that the constitutive theory for ordered thermofluids of all orders is indeed rate constitutive theory. The reseach work presented in this chapter

  12. American Focus on World Constitutions. Teacher's Guide.

    ERIC Educational Resources Information Center

    Holmes, Stanley T., III

    This curriculum project was designed to familiarize high school students with their own constitutional roots while gaining a better understanding of governmental systems developed by other nations. The project uses the U.S. Constitution as a baseline for analyzing the constitutions of other nations, and is intended to supplement courses in such…

  13. Estimating abundance: Chapter 27

    USGS Publications Warehouse

    Royle, J. Andrew

    2016-01-01

    This chapter provides a non-technical overview of ‘closed population capture–recapture’ models, a class of well-established models that are widely applied in ecology, such as removal sampling, covariate models, and distance sampling. These methods are regularly adopted for studies of reptiles, in order to estimate abundance from counts of marked individuals while accounting for imperfect detection. Thus, the chapter describes some classic closed population models for estimating abundance, with considerations for some recent extensions that provide a spatial context for the estimation of abundance, and therefore density. Finally, the chapter suggests some software for use in data analysis, such as the Windows-based program MARK, and provides an example of estimating abundance and density of reptiles using an artificial cover object survey of Slow Worms (Anguis fragilis).

  14. Pseudo-constitutivity of nitrate-responsive genes in nitrate reductase mutants

    PubMed Central

    Schinko, Thorsten; Gallmetzer, Andreas; Amillis, Sotiris; Strauss, Joseph

    2013-01-01

    In fungi, transcriptional activation of genes involved in NO3- assimilation requires the presence of an inducer (nitrate or nitrite) and low intracellular concentrations of the pathway products ammonium or glutamine. In Aspergillus nidulans, the two transcription factors NirA and AreA act synergistically to mediate nitrate/nitrite induction and nitrogen metabolite derepression, respectively. In all studied fungi and in plants, mutants lacking nitrate reductase (NR) activity express nitrate-metabolizing enzymes constitutively without the addition of inducer molecules. Based on their work in A. nidulans, Cove and Pateman proposed an “autoregulation control” model for the synthesis of nitrate metabolizing enzymes in which the functional nitrate reductase molecule would act as co-repressor in the absence and as co-inducer in the presence of nitrate. However, NR mutants could simply show “pseudo-constitutivity” due to induction by nitrate which accumulates over time in NR-deficient strains. Here we examined this possibility using strains which lack flavohemoglobins (fhbs), and are thus unable to generate nitrate internally, in combination with nitrate transporter mutations (nrtA, nrtB) and a GFP-labeled NirA protein. Using different combinations of genotypes we demonstrate that nitrate transporters are functional also in NR null mutants and show that the constitutive phenotype of NR mutants is not due to nitrate accumulation from intracellular sources but depends on the activity of nitrate transporters. However, these transporters are not required for nitrate signaling because addition of external nitrate (10 mM) leads to standard induction of nitrate assimilatory genes in the nitrate transporter double mutants. We finally show that NR does not regulate NirA localization and activity, and thus the autoregulation model, in which NR would act as a co-repressor of NirA in the absence of nitrate, is unlikely to be correct. Results from this study instead suggest

  15. Expression of endoglucanases in Pichia pastoris under control of the GAP promoter

    PubMed Central

    2014-01-01

    Background Plant-derived biomass is a potential alternative to fossil feedstocks for a greener economy. Enzymatic saccharification of biomass has been studied extensively and endoglucanases have been found to be a prerequisite for quick initial liquefaction of biomass under industrial conditions. Pichia pastoris, widely used for heterologous protein expression, can be utilized for fungal endoglucanase production. The recently marketed PichiaPink™ expression system allows for rapid clone selection, and employs the methanol inducible AOX1 promoter to ensure high protein expression levels. However, methanol is toxic and poses a fire hazard, issues which become more significant at an industrial scale. It is possible to eliminate these risks and still maintain high productivity by switching to the constitutive GAP promoter. Results In the present study, a plasmid carrying the constitutive GAP promoter was created for PichiaPink™. We then studied expression of two endoglucanases, AfCel12A from Aspergillus fumigatus and TaCel5A from Thermoascus aurantiacus, regulated by either the AOX1 promoter or the GAP promoter. Initial experiments in tubes and small bioreactors showed that the levels of AfCel12A obtained with the constitutive promoter were similar or higher, compared to the AOX1 promoter, whereas the levels of TaCel5A were somewhat lower. After optimization of cultivation conditions using a 15-l bioreactor, the recombinant P. pastoris strains utilizing the GAP promoter produced ca. 3–5 g/l of total secreted protein, with CMCase activity equivalent to 1200 nkat/ml AfCel12A and 170 nkat/ml TaCel5A. Conclusions We present a strategy for constitutive recombinant protein expression in the novel PichiaPink™ system. Both AfCel12A and TaCel5A were successfully expressed constitutively in P. pastoris under the GAP promoter. Reasonable protein levels were reached after optimizing cultivation conditions. PMID:24742273

  16. Expression of endoglucanases in Pichia pastoris under control of the GAP promoter.

    PubMed

    Várnai, Anikó; Tang, Campbell; Bengtsson, Oskar; Atterton, Andrew; Mathiesen, Geir; Eijsink, Vincent G H

    2014-04-18

    Plant-derived biomass is a potential alternative to fossil feedstocks for a greener economy. Enzymatic saccharification of biomass has been studied extensively and endoglucanases have been found to be a prerequisite for quick initial liquefaction of biomass under industrial conditions. Pichia pastoris, widely used for heterologous protein expression, can be utilized for fungal endoglucanase production. The recently marketed PichiaPink™ expression system allows for rapid clone selection, and employs the methanol inducible AOX1 promoter to ensure high protein expression levels. However, methanol is toxic and poses a fire hazard, issues which become more significant at an industrial scale. It is possible to eliminate these risks and still maintain high productivity by switching to the constitutive GAP promoter. In the present study, a plasmid carrying the constitutive GAP promoter was created for PichiaPink™. We then studied expression of two endoglucanases, AfCel12A from Aspergillus fumigatus and TaCel5A from Thermoascus aurantiacus, regulated by either the AOX1 promoter or the GAP promoter. Initial experiments in tubes and small bioreactors showed that the levels of AfCel12A obtained with the constitutive promoter were similar or higher, compared to the AOX1 promoter, whereas the levels of TaCel5A were somewhat lower. After optimization of cultivation conditions using a 15-l bioreactor, the recombinant P. pastoris strains utilizing the GAP promoter produced ca. 3-5 g/l of total secreted protein, with CMCase activity equivalent to 1200 nkat/ml AfCel12A and 170 nkat/ml TaCel5A. We present a strategy for constitutive recombinant protein expression in the novel PichiaPink™ system. Both AfCel12A and TaCel5A were successfully expressed constitutively in P. pastoris under the GAP promoter. Reasonable protein levels were reached after optimizing cultivation conditions.

  17. Stable CoT-1 repeat RNA is abundant and associated with euchromatic interphase chromosomes

    PubMed Central

    Hall, Lisa L.; Carone, Dawn M.; Gomez, Alvin; Kolpa, Heather J.; Byron, Meg; Mehta, Nitish; Fackelmayer, Frank O.; Lawrence, Jeanne B.

    2014-01-01

    SUMMARY Recent studies recognize a vast diversity of non-coding RNAs with largely unknown functions, but few have examined interspersed repeat sequences, which constitute almost half our genome. RNA hybridization in situ using CoT-1 (highly repeated) DNA probes detects surprisingly abundant euchromatin-associated RNA comprised predominantly of repeat sequences (“CoT-1 RNA”), including LINE-1. CoT-1-hybridizing RNA strictly localizes to the interphase chromosome territory in cis, and remains stably associated with the chromosome territory following prolonged transcriptional inhibition. The CoT-1 RNA territory resists mechanical disruption and fractionates with the non-chromatin scaffold, but can be experimentally released. Loss of repeat-rich, stable nuclear RNAs from euchromatin corresponds to aberrant chromatin distribution and condensation. CoT-1 RNA has several properties similar to XIST chromosomal RNA, but is excluded from chromatin condensed by XIST. These findings impact two “black boxes” of genome science: the poorly understood diversity of non-coding RNA and the unexplained abundance of repetitive elements. PMID:24581492

  18. South African court rejects country's new constitution.

    PubMed

    1996-09-20

    Fundamental principles designed to ensure that South Africa's new constitution upholds a wide range of individual rights and freedoms and establishes a responsive government with a balanced separation of powers, including recognition of the role of traditional tribal leadership, were adopted into the current interim constitution shortly before the 1994 free elections which brought Nelson Mandela and the African National Congress to power. In a judgement issued on September 6, 1996, South Africa's Constitutional Court rejected the country's new draft constitution, arguing that it failed to meet the standards of nine of the 34 principles established at the Kempton Park negotiations. The Constitutional Assembly is comprised of a joint meeting of the National Assembly and Senate. One of the court's major objections to the constitution concerned the proposed structure of rule, which was seen to give inadequate power to South Africa's nine provinces as compared with the national government. However, the bill of rights was almost entirely upheld. The bill would create a favorable environment for legalized abortion and guarantee a universal right of access to health care, including reproductive health services

  19. hebp3, a novel member of the heme-binding protein gene family, is expressed in the medaka meninges with higher abundance in females due to a direct stimulating action of ovarian estrogens.

    PubMed

    Nakasone, Kiyoshi; Nagahama, Yoshitaka; Okubo, Kataaki

    2013-02-01

    The brains of teleost fish exhibit remarkable sexual plasticity throughout their life span. To dissect the molecular basis for the development and reversal of sex differences in the teleost brain, we screened for genes differentially expressed between sexes in the brain of medaka (Oryzias latipes). One of the genes identified in the screen as being preferentially expressed in females was found to be a new member of the heme-binding protein gene family that includes hebp1 and hebp2 and was designated here as hebp3. The medaka hebp3 is expressed in the meninges with higher abundance in females, whereas there is no expression within the brain parenchyma. This female-biased expression of hebp3 is not attributable to the direct action of sex chromosome genes but results from the transient and reversible action of estrogens derived from the ovary. Moreover, estrogens directly activate the transcription of hebp3 via a palindromic estrogen-responsive element in the hebp3 promoter. Taken together, our findings demonstrate that hebp3 is a novel transcriptional target of estrogens, with female-biased expression in the meninges. The definite but reversible sexual dimorphism of the meningeal hebp3 expression may contribute to the development and reversal of sex differences in the teleost brain.

  20. [The right to health. Constitutional dimensions].

    PubMed

    Pestalozza, C

    2007-09-01

    A fundamental "right to health" is expressly guaranteed by the constitutions of several Bundesländer, but unknown to the German Federal Constitution. Instead the Federal Constitutional Court has - especially on the basis of related human rights - developed the obligation of the State to protect everybody's life and physical integrity, which in some respects comes near to a "right to health". The State's autonomy in financial matters, the scarcity of its financial resources and the individual's natural responsibility for his own health though advise against exaggerated hopes set in a "right to health".

  1. Tissue-specific mRNA expression profiling in grape berry tissues

    PubMed Central

    Grimplet, Jerome; Deluc, Laurent G; Tillett, Richard L; Wheatley, Matthew D; Schlauch, Karen A; Cramer, Grant R; Cushman, John C

    2007-01-01

    Background Berries of grape (Vitis vinifera) contain three major tissue types (skin, pulp and seed) all of which contribute to the aroma, color, and flavor characters of wine. The pericarp, which is composed of the exocarp (skin) and mesocarp (pulp), not only functions to protect and feed the developing seed, but also to assist in the dispersal of the mature seed by avian and mammalian vectors. The skin provides volatile and nonvolatile aroma and color compounds, the pulp contributes organic acids and sugars, and the seeds provide condensed tannins, all of which are important to the formation of organoleptic characteristics of wine. In order to understand the transcriptional network responsible for controlling tissue-specific mRNA expression patterns, mRNA expression profiling was conducted on each tissue of mature berries of V. vinifera Cabernet Sauvignon using the Affymetrix GeneChip® Vitis oligonucleotide microarray ver. 1.0. In order to monitor the influence of water-deficit stress on tissue-specific expression patterns, mRNA expression profiles were also compared from mature berries harvested from vines subjected to well-watered or water-deficit conditions. Results Overall, berry tissues were found to express approximately 76% of genes represented on the Vitis microarray. Approximately 60% of these genes exhibited significant differential expression in one or more of the three major tissue types with more than 28% of genes showing pronounced (2-fold or greater) differences in mRNA expression. The largest difference in tissue-specific expression was observed between the seed and pulp/skin. Exocarp tissue, which is involved in pathogen defense and pigment production, showed higher mRNA abundance relative to other berry tissues for genes involved with flavonoid biosynthesis, pathogen resistance, and cell wall modification. Mesocarp tissue, which is considered a nutritive tissue, exhibited a higher mRNA abundance of genes involved in cell wall function and

  2. Late Embryogenesis Abundant (LEA) Constitutes a Large and Diverse Family of Proteins Involved in Development and Abiotic Stress Responses in Sweet Orange (Citrus sinensis L. Osb.)

    PubMed Central

    Pedrosa, Andresa Muniz; Martins, Cristina de Paula Santos; Gonçalves, Luana Pereira; Costa, Marcio Gilberto Cardoso

    2015-01-01

    Late Embryogenesis Abundant (LEA) proteins are an ubiquitous group of polypeptides that were first described to accumulate during plant seed dehydration, at the later stages of embryogenesis. Since then they have also been recorded in vegetative plant tissues experiencing water limitation and in anhydrobiotic bacteria and invertebrates and, thereby, correlated with the acquisition of desiccation tolerance. This study provides the first comprehensive study about the LEA gene family in sweet orange (Citrus sinensis L. Osb.), the most important and widely grown fruit crop around the world. A surprisingly high number (72) of genes encoding C. sinensis LEAs (CsLEAs) were identified and classified into seven groups (LEA_1, LEA_2, LEA_3 and LEA_4, LEA_5, DEHYDRIN and SMP) based on their predicted amino acid sequences and also on their phylogenetic relationships with the complete set of Arabidopsis thaliana LEA proteins (AtLEAs). Approximately 60% of the CsLEAs identified in this study belongs to the unusual LEA_2 group of more hydrophobic LEA proteins, while the other LEA groups contained a relatively small number of members typically hydrophilic. A correlation between gene structure and motif composition was observed within each LEA group. Investigation of their chromosomal localizations revealed that the CsLEAs were non-randomly distributed across all nine chromosomes and that 33% of all CsLEAs are segmentally or tandemly duplicated genes. Analysis of the upstream sequences required for transcription revealed the presence of various stress-responsive cis-acting regulatory elements in the promoter regions of CsLEAs, including ABRE, DRE/CRT, MYBS and LTRE. Expression analysis using both RNA-seq data and quantitative real-time RT-PCR (qPCR) revealed that the CsLEA genes are widely expressed in various tissues, and that many genes containing the ABRE promoter sequence are induced by drought, salt and PEG. These results provide a useful reference for further exploration of

  3. Late Embryogenesis Abundant (LEA) Constitutes a Large and Diverse Family of Proteins Involved in Development and Abiotic Stress Responses in Sweet Orange (Citrus sinensis L. Osb.).

    PubMed

    Pedrosa, Andresa Muniz; Martins, Cristina de Paula Santos; Gonçalves, Luana Pereira; Costa, Marcio Gilberto Cardoso

    2015-01-01

    Late Embryogenesis Abundant (LEA) proteins are an ubiquitous group of polypeptides that were first described to accumulate during plant seed dehydration, at the later stages of embryogenesis. Since then they have also been recorded in vegetative plant tissues experiencing water limitation and in anhydrobiotic bacteria and invertebrates and, thereby, correlated with the acquisition of desiccation tolerance. This study provides the first comprehensive study about the LEA gene family in sweet orange (Citrus sinensis L. Osb.), the most important and widely grown fruit crop around the world. A surprisingly high number (72) of genes encoding C. sinensis LEAs (CsLEAs) were identified and classified into seven groups (LEA_1, LEA_2, LEA_3 and LEA_4, LEA_5, DEHYDRIN and SMP) based on their predicted amino acid sequences and also on their phylogenetic relationships with the complete set of Arabidopsis thaliana LEA proteins (AtLEAs). Approximately 60% of the CsLEAs identified in this study belongs to the unusual LEA_2 group of more hydrophobic LEA proteins, while the other LEA groups contained a relatively small number of members typically hydrophilic. A correlation between gene structure and motif composition was observed within each LEA group. Investigation of their chromosomal localizations revealed that the CsLEAs were non-randomly distributed across all nine chromosomes and that 33% of all CsLEAs are segmentally or tandemly duplicated genes. Analysis of the upstream sequences required for transcription revealed the presence of various stress-responsive cis-acting regulatory elements in the promoter regions of CsLEAs, including ABRE, DRE/CRT, MYBS and LTRE. Expression analysis using both RNA-seq data and quantitative real-time RT-PCR (qPCR) revealed that the CsLEA genes are widely expressed in various tissues, and that many genes containing the ABRE promoter sequence are induced by drought, salt and PEG. These results provide a useful reference for further exploration of

  4. A Method for the Construction of Hereditary Constitutive Equations of Laminates Bases on a Hereditary Constitutive Equation for a Layer

    NASA Astrophysics Data System (ADS)

    Dumansky, Alexander M.; Tairova, Lyudmila P.

    2008-09-01

    A method for the construction of hereditary constitutive equation is proposed for the laminate on the basis of hereditary constitutive equations of a layer. The method is developed from the assumption that in the directions of axes of orthotropy the layer follows elastic behavior, and obeys hereditary constitutive equations under shear. The constitutive equations of the laminate are constructed on the basis of classical laminate theory and algebra of resolvent operators. Effective matrix algorithm and relationships of operator algebra are used to derive visco-elastic stiffness and compliance of the laminate. The example of construction of hereditary constitutive equations of cross-ply carbon fiber-reinforced plastic is presented.

  5. The transketolase gene family of the resurrection plant Craterostigma plantagineum: differential expression during the rehydration phase.

    PubMed Central

    Bernacchia, G; Schwall, G; Lottspeich, F; Salamini, F; Bartels, D

    1995-01-01

    Transketolases, key enzymes of the reductive and oxidative pentose phosphate pathways, are responsible for the synthesis of sugar phosphate intermediates. Here we report the first molecular analysis of transketolase genes from plants. Three distinct classes of transketolase-encoding cDNA clones were isolated from the desiccation-tolerant resurrection plant Craterostigma plantagineum. One class represented by the transcript tkt3 is constitutively expressed in leaves and roots under all physiological conditions tested. By biochemical analysis and protein sequencing of purified transketolase, it was shown that tkt3 is expressed in three enzymatically active isoforms. An intriguing discovery was that accumulation of the two other transketolase transcripts, tkt7 and tkt10, is preferentially associated with the rehydration process of the desiccated plant; whereas tkt10 is only expressed in leaves, tkt7 was detected in leaves and roots. This observation suggests a possible role for these transketolases in the conversion of sugars, which are a major phenomenon in the rehydration process. Despite an abundant level of tkt7 and tkt10 transcripts in rehydrating leaves, proteins could not be isolated. This is due in part to a translational control mechanism acting on the loading of mRNAs to polysomes. Images PMID:7859749

  6. Expression and characterization of constitutive heat shock protein 70.1 (HSPA-1A) gene in in vitro produced and in vivo-derived buffalo (Bubalus bubalis) embryos.

    PubMed

    Sharma, G T; Nath, A; Prasad, S; Singhal, S; Singh, N; Gade, N E; Dubey, P K; Saikumar, G

    2012-12-01

    Cells are blessed with a group of stress protector molecules known as heat shock proteins (HSPs), amongst them HSP70, encoded by HSPA-1A gene, is most abundant and highly conserved protein. Variety of stresses hampers the developmental competence of embryos under in vivo and in vitro conditions. Present work was designed to study the quantitative expression of HSPA-1A mRNA in immature oocytes (IMO), matured oocytes (MO), in vitro produced (IVP) and in vivo-derived (IVD) buffalo embryos to assess the level of stress to which embryos are exposed under in vivo and in vitro culture conditions. Further, HSPA-1A gene sequence was analysed to determine its homology with other mammalian sequences. The mRNA expression analysis was carried out on 72 oocytes (40 IMO; 32 MO), 76 IVP and 55 IVD buffalo embryos. Expression of HSPA-1A was found in oocytes and throughout the developmental stages of embryos examined irrespective of the embryo source; however, higher (p < 0.05) expression was observed in 8-16 cell, morula and blastocyst stages of IVP embryos as compared to IVD embryos. Phylogenetic analysis of bubaline HSPA-1A revealed that it shares 91-98% identity with other mammalian sequences. It can be concluded that higher level of HSPA-1A mRNA in IVP embryos in comparison with in vivo-derived embryos is an indicator of cellular stress in IVP system. This study suggests need for further optimization of in vitro culture system in which HSPA-1A gene could be used as a stress biomarker during pre-implantation development. © 2012 Blackwell Verlag GmbH.

  7. Differential protein abundance of a basolateral MCT1 transporter in the human gastrointestinal tract.

    PubMed

    Al-Mosauwi, Hashemeya; Ryan, Elizabeth; McGrane, Alison; Riveros-Beltran, Stefanie; Walpole, Caragh; Dempsey, Eugene; Courtney, Danielle; Fearon, Naomi; Winter, Desmond; Baird, Alan; Stewart, Gavin

    2016-12-01

    Bacterially derived short chain fatty acids (SCFAs), such as butyrate, are vital in maintaining the symbiotic relationship that exists between humans and their gastrointestinal microbial populations. A key step in this process is the transport of SCFAs across colonic epithelial cells via MCT1 transporters. This study investigated MCT1 protein abundance in various human intestinal tissues. Initial RT-PCR analysis confirmed the expected MCT1 RNA expression pattern of colon > small intestine > stomach. Using surgical resection samples, immunoblot analysis detected higher abundance of a 45 kDa MCT1 protein in colonic tissue compared to ileum tissue (P < 0.001, N = 4, unpaired t-test). Importantly, MCT1 abundance was found to be significantly lower in sigmoid colon compared to ascending colon (P < 0.01, N = 8-11, ANOVA). Finally, immunolocalization studies confirmed MCT1 to be abundant in the basolateral membranes of surface epithelial cells of the ascending, transverse, and descending colon, but significantly less prevalent in the sigmoid colon (P < 0.05, N = 5-21, ANOVA). In conclusion, these data confirm that basolateral MCT1 protein abundance is correlated to levels of bacterially derived SCFAs along the human gastrointestinal tract. These findings highlight the importance of precise tissue location in studies comparing colonic MCT1 abundance between normal and diseased states. © 2016 International Federation for Cell Biology.

  8. Transgenic expression of medicago truncatula PR10 and PR5 promoters in alfalfa shows pathogen-induced up-regulation of transgene expression

    USDA-ARS?s Scientific Manuscript database

    Genetic modification of alfalfa to introduce novel traits requires promoters for controlling gene expression. Promoters that are constitutively activated for expression of genes that enhance disease resistance pose a great energy load on the plant and exert a strong selective pressure on the pathoge...

  9. 78 FR 57777 - Constitution Day and Citizenship Day, Constitution Week, 2013

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-09-19

    ... that in the face of impossible odds, those who love their country can change it. In remembrance of the signing of the Constitution and in recognition of the Americans who strive to uphold the duties and...

  10. RNA-Seq Alignment to Individualized Genomes Improves Transcript Abundance Estimates in Multiparent Populations

    PubMed Central

    Munger, Steven C.; Raghupathy, Narayanan; Choi, Kwangbom; Simons, Allen K.; Gatti, Daniel M.; Hinerfeld, Douglas A.; Svenson, Karen L.; Keller, Mark P.; Attie, Alan D.; Hibbs, Matthew A.; Graber, Joel H.; Chesler, Elissa J.; Churchill, Gary A.

    2014-01-01

    Massively parallel RNA sequencing (RNA-seq) has yielded a wealth of new insights into transcriptional regulation. A first step in the analysis of RNA-seq data is the alignment of short sequence reads to a common reference genome or transcriptome. Genetic variants that distinguish individual genomes from the reference sequence can cause reads to be misaligned, resulting in biased estimates of transcript abundance. Fine-tuning of read alignment algorithms does not correct this problem. We have developed Seqnature software to construct individualized diploid genomes and transcriptomes for multiparent populations and have implemented a complete analysis pipeline that incorporates other existing software tools. We demonstrate in simulated and real data sets that alignment to individualized transcriptomes increases read mapping accuracy, improves estimation of transcript abundance, and enables the direct estimation of allele-specific expression. Moreover, when applied to expression QTL mapping we find that our individualized alignment strategy corrects false-positive linkage signals and unmasks hidden associations. We recommend the use of individualized diploid genomes over reference sequence alignment for all applications of high-throughput sequencing technology in genetically diverse populations. PMID:25236449

  11. Cognition research and constitutional classification in Chinese medicine.

    PubMed

    Wang, Ji; Li, Yingshuai; Ni, Cheng; Zhang, Huimin; Li, Lingru; Wang, Qi

    2011-01-01

    In the Western medicine system, scholars have explained individual differences in terms of behaviour and thinking, leading to the emergence of various classification theories on individual differences. Traditional Chinese medicine has long observed human constitutions. Modern Chinese medicine studies have also involved study of human constitutions; however, differences exist in the ways traditional and modern Chinese medicine explore individual constitutions. In the late 1970s, the constitutional theory of Chinese medicine was proposed. This theory takes a global and dynamic view of human differences (e.g., the shape of the human body, function, psychology, and other characteristics) based on arguments from traditional Chinese medicine. The establishment of a standard for classifying constitutions into nine modules was critical for clinical application of this theory. In this review, we describe the history and recent research progress of this theory, and compare it with related studies in the western medicine system. Several research methods, including philology, informatics, epidemiology, and molecular biology, in classifying constitutions used in the constitutional theory of Chinese medicine were discussed. In summary, this constitutional theory of Chinese medicine can be used in clinical practice and would contribute to health control of patients.

  12. Temporal transcriptome analysis of the white-rot fungus Obba rivulosa shows expression of a constitutive set of plant cell wall degradation targeted genes during growth on solid spruce wood.

    PubMed

    Marinović, Mila; Aguilar-Pontes, Maria Victoria; Zhou, Miaomiao; Miettinen, Otto; de Vries, Ronald P; Mäkelä, Miia R; Hildén, Kristiina

    2018-03-01

    The basidiomycete white-rot fungus Obba rivulosa, a close relative of Gelatoporia (Ceriporiopsis) subvermispora, is an efficient degrader of softwood. The dikaryotic O. rivulosa strain T241i (FBCC949) has been shown to selectively remove lignin from spruce wood prior to depolymerization of plant cell wall polysaccharides, thus possessing potential in biotechnological applications such as pretreatment of wood in pulp and paper industry. In this work, we studied the time-course of the conversion of spruce by the genome-sequenced monokaryotic O. rivulosa strain 3A-2, which is derived from the dikaryon T241i, to get insight into transcriptome level changes during prolonged solid state cultivation. During 8-week cultivation, O. rivulosa expressed a constitutive set of genes encoding putative plant cell wall degrading enzymes. High level of expression of the genes targeted towards all plant cell wall polymers was detected at 2-week time point, after which majority of the genes showed reduced expression. This implicated non-selective degradation of lignin by the O. rivulosa monokaryon and suggests high variation between mono- and dikaryotic strains of the white-rot fungi with respect to their abilities to convert plant cell wall polymers. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. A constitutive rheological model for agglomerating blood derived from nonequilibrium thermodynamics

    NASA Astrophysics Data System (ADS)

    Tsimouri, Ioanna Ch.; Stephanou, Pavlos S.; Mavrantzas, Vlasis G.

    2018-03-01

    Red blood cells tend to aggregate in the presence of plasma proteins, forming structures known as rouleaux. Here, we derive a constitutive rheological model for human blood which accounts for the formation and dissociation of rouleaux using the generalized bracket formulation of nonequilibrium thermodynamics. Similar to the model derived by Owens and co-workers ["A non-homogeneous constitutive model for human blood. Part 1. Model derivation and steady flow," J. Fluid Mech. 617, 327-354 (2008)] through polymer network theory, each rouleau in our model is represented as a dumbbell; the corresponding structural variable is the conformation tensor of the dumbbell. The kinetics of rouleau formation and dissociation is treated as in the work of Germann et al. ["Nonequilibrium thermodynamic modeling of the structure and rheology of concentrated wormlike micellar solutions," J. Non-Newton. Fluid Mech. 196, 51-57 (2013)] by assuming a set of reversible reactions, each characterized by a forward and a reverse rate constant. The final set of evolution equations for the microstructure of each rouleau and the expression for the stress tensor turn out to be very similar to those of Owens and co-workers. However, by explicitly considering a mechanism for the formation and breakage of rouleaux, our model further provides expressions for the aggregation and disaggregation rates appearing in the final transport equations, which in the kinetic theory-based network model of Owens were absent and had to be specified separately. Despite this, the two models are found to provide similar descriptions of experimental data on the size distribution of rouleaux.

  14. Increased skin barrier disruption by sodium lauryl sulfate in mice expressing a constitutively active STAT6 in T cells

    PubMed Central

    DaSilva, Sonia C.; Sahu, Ravi P.; Konger, Raymond L.; Perkins, Susan M.; Kaplan, Mark H.; Travers, Jeffrey B.

    2011-01-01

    Atopic dermatitis (AD) is a pruritic, chronic inflammatory skin disease that affects 10–20% of children and 1–3% of adults worldwide. Recent studies have indicated that the ability of Th2 cytokines such as interleukin-4 (IL-4) to regulate skin barrier function may be a predisposing factor for AD development. The present studies examined the ability of increased Th2 activity to affect cutaneous barrier function in vivo and epidermal thickening. Mice that express a constitutively active Signal Transducer and Activator of Transcription 6 (STAT6VT) have increased Th2 cells and a predisposition to allergic inflammation were used in these studies; they demonstrate that topical treatment with the irritant sodium lauryl sulfate (SLS) caused increased transepidermal water loss and epidermal thickening in STAT6VT mice over similarly treated wild-type mice. The proliferation marker Ki-67 was increased in the epidermis of STAT6VT compared to wild-type mice. However, these differences do not appear to be linked to the addition of an irritant as control-treated STAT6VT skin also exhibited elevated Ki-67 levels, suggesting that the increased epidermal thickness in SLS-treated STAT6VT mice is primarily driven by epidermal cell hypertrophy rather than an increase in cellular proliferation. Our results suggest that an environment with increased Th2 cytokines results in abnormal responses to topical irritants. PMID:21959772

  15. Conditional and constitutive expression of a Tbx1-GFP fusion protein in mice

    PubMed Central

    2013-01-01

    Background Velo-cardio-facial syndrome/DiGeorge syndrome (VCFS/DGS) is caused by a 1.5-3 Mb microdeletion of chromosome 22q11.2, frequently referred to as 22q11.2 deletion syndrome (22q11DS). This region includes TBX1, a T-box transcription factor gene that contributes to the etiology of 22q11DS. The requirement for TBX1 in mammalian development is dosage-sensitive, such that loss-of-function (LOF) and gain-of-function (GOF) of TBX1 in both mice and humans results in disease relevant congenital malformations. Results To further gain insight into the role of Tbx1 in development, we have targeted the Rosa26 locus to generate a new GOF mouse model in which a Tbx1-GFP fusion protein is expressed conditionally using the Cre/LoxP system. Tbx1-GFP expression is driven by the endogenous Rosa26 promoter resulting in ectopic and persistent expression. Tbx1 is pivotal for proper ear and heart development; ectopic activation of Tbx1-GFP in the otic vesicle by Pax2-Cre and Foxg1-Cre represses neurogenesis and produces morphological defects of the inner ear. Overexpression of a single copy of Tbx1-GFP using Tbx1Cre/+ was viable, while overexpression of both copies resulted in neonatal lethality with cardiac outflow tract defects. We have partially rescued inner ear and heart anomalies in Tbx1Cre/- null embryos by expression of Tbx1-GFP. Conclusions We have generated a new mouse model to conditionally overexpress a GFP-tagged Tbx1 protein in vivo. This provides a useful tool to investigate in vivo direct downstream targets and protein binding partners of Tbx1. PMID:23971992

  16. Solar abundance of silicon

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Holweger, H.

    1973-07-01

    An analysis of 19 photospheric Si I lines whose oscillator strengths have recently been detertmined by Garz (1973) leads to a solar abundance of silicon, log epsilon /sub Si/ = 7.65 plus or minus 0.07, on the scale where log epsilon /sub H/ = 12. Together with the sodium abundance determained earlier by the same method, a solar abundance ratio /sup epsilon /Na//sup epsilon /Si = 0.045 ( plus or minus 10%) results. Within the error limits this a grees wtth the meteoritic ratio found in carbonaceous chondrites. Results concerning line-broadening by hydrogen are discussed. (auth)

  17. Genetic Approach to Elucidation of Sasang Constitutional Medicine

    PubMed Central

    Kim, Bu-Yeo; Cha, Seongwon; Jin, Hee-Jeong

    2009-01-01

    Sasang Constitutional Medicine (SCM) offers a medical principle that classifies humans into four constitution groups and guides their treatment with constitution-matched medical assistance. The principle of this traditional medicine, although requires significant scientific support, appears to suggest a genetic influence on constitution type. The relative frequency of constitution types in a population, for instance, has remained relatively constant since Jema Lee first described them from his observations. In addition, the body compartment concept of SCM appears to be related to the anterio–posterior patterning of the embryonic gut and associated internal organs. This study describes the attributes of the constitution concept of SCM that can be interpreted in the language of genetics and current approaches to identity the genetic factors that make up the constitution. These efforts should make it possible to interpret the principle of this traditional medicine scientifically. Considering the recent trend in medicine that pursues individualized or tailored medical offerings, once SCM is proven to be explainable with scientific evidence, it will be able to contribute to and take a place in the rapidly evolving medicine environment. PMID:19745011

  18. Fructooligosaccharides production by Schedonorus arundinaceus sucrose:sucrose 1-fructosyltransferase constitutively expressed to high levels in Pichia pastoris.

    PubMed

    Hernández, Lázaro; Menéndez, Carmen; Pérez, Enrique R; Martínez, Duniesky; Alfonso, Dubiel; Trujillo, Luis E; Ramírez, Ricardo; Sobrino, Alina; Mazola, Yuliet; Musacchio, Alexis; Pimentel, Eulogio

    2018-01-20

    The non-saccharolytic yeast Pichia pastoris was engineered to express constitutively the mature region of sucrose:sucrose 1-fructosyltransferase (1-SST, EC 2.4.1.99) from Tall fescue (Schedonorus arundinaceus). The increase of the transgene dosage from one to nine copies enhanced 7.9-fold the recombinant enzyme (Sa1-SSTrec) yield without causing cell toxicity. Secretion driven by the Saccharomyces cerevisiae α-factor signal peptide resulted in periplasmic retention (38%) and extracellular release (62%) of Sa1-SSTrec to an overall activity of 102.1 U/ml when biomass reached (106 g/l, dry weight) in fed-batch fermentation using cane sugar for cell growth. The volumetric productivity of the nine-copy clone PGFT6x-308 at the end of fermentation (72 h) was 1422.2 U/l/h. Sa1-SSTrec purified from the culture supernatant was a monomeric glycoprotein optimally active at pH 5.0-6.0 and 45-50 °C. The removal of N-linked oligosaccharides by Endo Hf treatment decreased the enzyme stability but had no effect on the substrate and product specificities. Sa1-SSTrec converted sucrose (600 g/l) into 1-kestose (GF 2 ) and nystose (GF 3 ) in a ratio 9:1 with their sum representing 55-60% (w/w) of the total carbohydrates in the reaction mixture. Variations in the sucrose (100-800 g/l) or enzyme (1.5-15 units per gram of substrate) concentrations kept unaltered the product profile. Sa1-SSTrec is an attractive candidate enzyme for the industrial production of short-chain fructooligosaccharides, most particularly 1-kestose. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Expression of a Dianthus flavonoid glucosyltransferase in Saccharomyces cerevisiae for whole-cell biocatalysis.

    PubMed

    Werner, Sean R; Morgan, John A

    2009-07-15

    Glycosyltransferases are promising biocatalysts for the synthesis of small molecule glycosides. In this study, Saccharomyces cerevisiae expressing a flavonoid glucosyltransferase (GT) from Dianthus caryophyllus (carnation) was investigated as a whole-cell biocatalyst. Two yeast expression systems were compared using the flavonoid naringenin as a model substrate. Under in vitro conditions, naringenin-7-O-glucoside was formed and a higher specific glucosyl transfer activity was found using a galactose inducible expression system compared to a constitutive expression system. However, S. cerevisiae expressing the GT constitutively was significantly more productive than the galactose inducible system under in vivo conditions. Interestingly, the glycosides were recovered directly from the culture broth and did not accumulate intracellularly. A previously uncharacterized naringenin glycoside formed using the D. caryophyllus GT was identified as naringenin-4'-O-glucoside. It was found that S. cerevisiae cells hydrolyze naringenin-7-O-glucoside during whole-cell biocatalysis, resulting in a low final glycoside titer. When phloretin was added as a substrate to the yeast strain expressing the GT constitutively, the natural product phlorizin was formed. This study demonstrates S. cerevisiae is a promising whole-cell biocatalyst host for the production of valuable glycosides.

  20. Correlation Between Expression of Recombinant Proteins and Abundance of H3K4Me3 on the Enhancer of Human Cytomegalovirus Major Immediate-Early Promoter.

    PubMed

    Soo, Benjamin P C; Tay, Julian; Ng, Shirelle; Ho, Steven C L; Yang, Yuansheng; Chao, Sheng-Hao

    2017-08-01

    Role of epigenetic regulation in the control of gene expression is well established. The impact of several epigenetic mechanisms, such as DNA methylation and histone acetylation, on recombinant protein production in mammalian cells has been investigated recently. Here we investigate the correlation between the selected epigenetic markers and five trastuzumab biosimilar-producing Chinese hamster ovary (CHO) cell lines in which the expression of trastuzumab is driven by human cytomegalovirus (HCMV) major immediate-early (MIE) promoter. We chose the producing clones in which transcription was the determinative step for the production of recombinant trastuzumab. We found that the abundance of trimethylation of histone 3 at lysine 4 (H3K4Me3) on the enhancer of HCMV MIE promoter correlated well with the relative titers of recombinant trastuzumab among the clones. Such close correlation was not observed between the recombinant protein and other epigenetic markers examined in our study. Our results demonstrate that the HCMV MIE enhancer-bound H3K4Me3 epigenetic marker may be used as the epigenetic indicator to predict the relative production of recombinant proteins between the producing CHO cell lines.

  1. Micromechanics of Composite Materials Governed by Vector Constitutive Laws

    NASA Technical Reports Server (NTRS)

    Bednarcyk, Brett A.; Aboudi, Jacob; Arnold, Steven M.

    2017-01-01

    The high-fidelity generalized method of cells micromechanics theory has been extended for the prediction of the effective property tensor and the corresponding local field distributions for composites whose constituents are governed by vector constitutive laws. As shown, the shear analogy, which can predict effective transverse properties, is not valid in the general three-dimensional case. Consequently, a general derivation is presented that is applicable to both continuously and discontinuously reinforced composites with arbitrary vector constitutive laws and periodic microstructures. Results are given for thermal and electric problems, effective properties and local field distributions, ordered and random microstructures, as well as complex geometries including woven composites. Comparisons of the theory's predictions are made to test data, numerical analysis, and classical expressions from the literature. Further, classical methods cannot provide the local field distributions in the composite, and it is demonstrated that, as the percolation threshold is approached, their predictions are increasingly unreliable. XXXX It has been observed that the bonding between the fibers and matrix in composite materials can be imperfect. In the context of thermal conductivity, such imperfect interfaces have been investigated in micromechanical models by Dunn and Taya (1993), Duan and Karihaloo (2007), Nan et al. (1997) and Hashin (2001). The present HFGMC micromechanical method, derived for perfectly bonded composite materials governed by vector constitutive laws, can be easily generalized to include the effects of weak bonding between the constituents. Such generalizations, in the context of the mechanical micromechanics problem, involve introduction of a traction-separation law at the fiber/matrix interface and have been presented by Aboudi (1987), Bednarcyk and Arnold (2002), Bednarcyk et al. (2004) and Aboudi et al. (2013) and will be addressed in the future.

  2. Antioxidant supplement inhibits skeletal muscle constitutive autophagy rather than fasting-induced autophagy in mice.

    PubMed

    Qi, Zhengtang; He, Qiang; Ji, Liu; Ding, Shuzhe

    2014-01-01

    In this study, we tested the hypothesis that NAC administration leads to reduced oxidative stress and thus to decreased expression of autophagy markers in young mice. Our results reveal that NAC administration results in reduced muscle mRNA levels of several autophagy markers, including Beclin-1, Atg7, LC3, Atg9, and LAMP2. However, NAC supplement fails to block the activation of skeletal muscle autophagy in response to fasting, because fasting significantly increases the mRNA level of several autophagy markers and LC3 lipidation. We further examined the effects of NAC administration on mitochondrial antioxidant capacity in fed and 24-hour fasted mice. Our results clearly show that NAC administration depresses the expression of manganese superoxide dismutase (MnSOD) and TP53-induced glycolysis and apoptosis regulator (TIGAR), both of which play a predominant antioxidant role in mitochondria by reducing ROS level. In addition, we found no beneficial effect of NAC supplement on muscle mass but it can protect from muscle loss in response to fasting. Collectively, our findings indicate that ROS is required for skeletal muscle constitutive autophagy, rather than starvation-induced autophagy, and that antioxidant NAC inhibits constitutive autophagy by the regulation of mitochondrial ROS production and antioxidant capacity.

  3. Antioxidant Supplement Inhibits Skeletal Muscle Constitutive Autophagy rather than Fasting-Induced Autophagy in Mice

    PubMed Central

    Qi, Zhengtang; He, Qiang; Ji, Liu; Ding, Shuzhe

    2014-01-01

    In this study, we tested the hypothesis that NAC administration leads to reduced oxidative stress and thus to decreased expression of autophagy markers in young mice. Our results reveal that NAC administration results in reduced muscle mRNA levels of several autophagy markers, including Beclin-1, Atg7, LC3, Atg9, and LAMP2. However, NAC supplement fails to block the activation of skeletal muscle autophagy in response to fasting, because fasting significantly increases the mRNA level of several autophagy markers and LC3 lipidation. We further examined the effects of NAC administration on mitochondrial antioxidant capacity in fed and 24-hour fasted mice. Our results clearly show that NAC administration depresses the expression of manganese superoxide dismutase (MnSOD) and TP53-induced glycolysis and apoptosis regulator (TIGAR), both of which play a predominant antioxidant role in mitochondria by reducing ROS level. In addition, we found no beneficial effect of NAC supplement on muscle mass but it can protect from muscle loss in response to fasting. Collectively, our findings indicate that ROS is required for skeletal muscle constitutive autophagy, rather than starvation-induced autophagy, and that antioxidant NAC inhibits constitutive autophagy by the regulation of mitochondrial ROS production and antioxidant capacity. PMID:25028602

  4. Histone methylation mediates plasticity of human FOXP3(+) regulatory T cells by modulating signature gene expressions.

    PubMed

    He, Haiqi; Ni, Bing; Tian, Yi; Tian, Zhiqiang; Chen, Yanke; Liu, Zhengwen; Yang, Xiaomei; Lv, Yi; Zhang, Yong

    2014-03-01

    CD4(+) FOXP3(+) regulatory T (Treg) cells constitute a heterogeneous and plastic T-cell lineage that plays a pivotal role in maintaining immune homeostasis and immune tolerance. However, the fate of human Treg cells after loss of FOXP3 expression and the epigenetic mechanisms contributing to such a phenotype switch remain to be fully elucidated. In the current study, we demonstrate that human CD4(+) CD25(high) CD127(low/-) Treg cells convert to two subpopulations with distinctive FOXP3(+) and FOXP3(-) phenotypes following in vitro culture with anti-CD3/CD28 and interleukin-2. Digital gene expression analysis showed that upon in vitro expansion, human Treg cells down-regulated Treg cell signature genes, such as FOXP3, CTLA4, ICOS, IKZF2 and LRRC32, but up-regulated a set of T helper lineage-associated genes, especially T helper type 2 (Th2)-associated, such as GATA3, GFI1 and IL13. Subsequent chromatin immunoprecipitation-sequencing of these subpopulations yielded genome-wide maps of their H3K4me3 and H3K27me3 profiles. Surprisingly, reprogramming of Treg cells was associated with differential histone modifications, as evidenced by decreased abundance of permissive H3K4me3 within the down-regulated Treg cell signature genes, such as FOXP3, CTLA4 and LRRC32 loci, and increased abundance of H3K4me3 within the Th2-associated genes, such as IL4 and IL5; however, the H3K27me3 modification profile was not significantly different between the two subpopulations. In conclusion, this study revealed that loss of FOXP3 expression from human Treg cells during in vitro expansion can induce reprogramming to a T helper cell phenotype with a gene expression signature dominated by Th2 lineage-associated genes, and that this cell type conversion may be mediated by histone methylation events. © 2013 John Wiley & Sons Ltd.

  5. Histone methylation mediates plasticity of human FOXP3+ regulatory T cells by modulating signature gene expressions

    PubMed Central

    He, Haiqi; Ni, Bing; Tian, Yi; Tian, Zhiqiang; Chen, Yanke; Liu, Zhengwen; Yang, Xiaomei; Lv, Yi; Zhang, Yong

    2014-01-01

    CD4+ FOXP3+ regulatory T (Treg) cells constitute a heterogeneous and plastic T-cell lineage that plays a pivotal role in maintaining immune homeostasis and immune tolerance. However, the fate of human Treg cells after loss of FOXP3 expression and the epigenetic mechanisms contributing to such a phenotype switch remain to be fully elucidated. In the current study, we demonstrate that human CD4+ CD25high CD127low/− Treg cells convert to two subpopulations with distinctive FOXP3+ and FOXP3− phenotypes following in vitro culture with anti-CD3/CD28 and interleukin-2. Digital gene expression analysis showed that upon in vitro expansion, human Treg cells down-regulated Treg cell signature genes, such as FOXP3, CTLA4, ICOS, IKZF2 and LRRC32, but up-regulated a set of T helper lineage-associated genes, especially T helper type 2 (Th2)-associated, such as GATA3, GFI1 and IL13. Subsequent chromatin immunoprecipitation-sequencing of these subpopulations yielded genome-wide maps of their H3K4me3 and H3K27me3 profiles. Surprisingly, reprogramming of Treg cells was associated with differential histone modifications, as evidenced by decreased abundance of permissive H3K4me3 within the down-regulated Treg cell signature genes, such as FOXP3, CTLA4 and LRRC32 loci, and increased abundance of H3K4me3 within the Th2-associated genes, such as IL4 and IL5; however, the H3K27me3 modification profile was not significantly different between the two subpopulations. In conclusion, this study revealed that loss of FOXP3 expression from human Treg cells during in vitro expansion can induce reprogramming to a T helper cell phenotype with a gene expression signature dominated by Th2 lineage-associated genes, and that this cell type conversion may be mediated by histone methylation events. PMID:24152290

  6. The Dynamics of Transcript Abundance during Cellularization of Developing Barley Endosperm1[OPEN

    PubMed Central

    Zhang, Runxuan; Burton, Rachel A; Shirley, Neil J.; Little, Alan; Morris, Jenny; Milne, Linda

    2016-01-01

    Within the cereal grain, the endosperm and its nutrient reserves are critical for successful germination and in the context of grain utilization. The identification of molecular determinants of early endosperm development, particularly regulators of cell division and cell wall deposition, would help predict end-use properties such as yield, quality, and nutritional value. Custom microarray data have been generated using RNA isolated from developing barley grain endosperm 3 d to 8 d after pollination (DAP). Comparisons of transcript abundance over time revealed 47 gene expression modules that can be clustered into 10 broad groups. Superimposing these modules upon cytological data allowed patterns of transcript abundance to be linked with key stages of early grain development. Here, attention was focused on how the datasets could be mined to explore and define the processes of cell wall biosynthesis, remodeling, and degradation. Using a combination of spatial molecular network and gene ontology enrichment analyses, it is shown that genes involved in cell wall metabolism are found in multiple modules, but cluster into two main groups that exhibit peak expression at 3 DAP to 4 DAP and 5 DAP to 8 DAP. The presence of transcription factor genes in these modules allowed candidate genes for the control of wall metabolism during early barley grain development to be identified. The data are publicly available through a dedicated web interface (https://ics.hutton.ac.uk/barseed/), where they can be used to interrogate co- and differential expression for any other genes, groups of genes, or transcription factors expressed during early endosperm development. PMID:26754666

  7. Frederick Douglass Changed My Mind about the Constitution

    ERIC Educational Resources Information Center

    Oakes, James

    2008-01-01

    Frederick Douglass dramatically and publicly changed his own mind about the Constitution. Like Frederick Douglass, the author had originally viewed the Constitution as pro-slavery. Yet a close look at Douglass's writings revealed a Constitution that empowered the federal government to abolish slavery.

  8. Changes in transcript abundance relating to colony collapse disorder in honey bees (Apis mellifera).

    PubMed

    Johnson, Reed M; Evans, Jay D; Robinson, Gene E; Berenbaum, May R

    2009-09-01

    Colony collapse disorder (CCD) is a mysterious disappearance of honey bees that has beset beekeepers in the United States since late 2006. Pathogens and other environmental stresses, including pesticides, have been linked to CCD, but a causal relationship has not yet been demonstrated. Because the gut acts as a primary interface between the honey bee and its environment as a site of entry for pathogens and toxins, we used whole-genome microarrays to compare gene expression between guts of bees from CCD colonies originating on both the east and west coasts of the United States and guts of bees from healthy colonies sampled before the emergence of CCD. Considerable variation in gene expression was associated with the geographical origin of bees, but a consensus list of 65 transcripts was identified as potential markers for CCD status. Overall, elevated expression of pesticide response genes was not observed. Genes involved in immune response showed no clear trend in expression pattern despite the increased prevalence of viruses and other pathogens in CCD colonies. Microarray analysis revealed unusual ribosomal RNA fragments that were conspicuously more abundant in the guts of CCD bees. The presence of these fragments may be a possible consequence of picorna-like viral infection, including deformed wing virus and Israeli acute paralysis virus, and may be related to arrested translation. Ribosomal fragment abundance and presence of multiple viruses may prove to be useful diagnostic markers for colonies afflicted with CCD.

  9. Changes in transcript abundance relating to colony collapse disorder in honey bees (Apis mellifera)

    PubMed Central

    Johnson, Reed M.; Evans, Jay D.; Robinson, Gene E.; Berenbaum, May R.

    2009-01-01

    Colony collapse disorder (CCD) is a mysterious disappearance of honey bees that has beset beekeepers in the United States since late 2006. Pathogens and other environmental stresses, including pesticides, have been linked to CCD, but a causal relationship has not yet been demonstrated. Because the gut acts as a primary interface between the honey bee and its environment as a site of entry for pathogens and toxins, we used whole-genome microarrays to compare gene expression between guts of bees from CCD colonies originating on both the east and west coasts of the United States and guts of bees from healthy colonies sampled before the emergence of CCD. Considerable variation in gene expression was associated with the geographical origin of bees, but a consensus list of 65 transcripts was identified as potential markers for CCD status. Overall, elevated expression of pesticide response genes was not observed. Genes involved in immune response showed no clear trend in expression pattern despite the increased prevalence of viruses and other pathogens in CCD colonies. Microarray analysis revealed unusual ribosomal RNA fragments that were conspicuously more abundant in the guts of CCD bees. The presence of these fragments may be a possible consequence of picorna-like viral infection, including deformed wing virus and Israeli acute paralysis virus, and may be related to arrested translation. Ribosomal fragment abundance and presence of multiple viruses may prove to be useful diagnostic markers for colonies afflicted with CCD. PMID:19706391

  10. Presidential Signing Statements: Constitutional and Institutional Implications

    DTIC Science & Technology

    2007-04-13

    Association (ABA) recently publishing a report declaring that these instruments are “contrary to the rule of law and our constitutional separation of powers ” when...Force on Presidential Signing Statements and the Separation of Powers Doctrine at p. 5 (August 2006). Presidential Signing Statements: Constitutional...constitutional separation of powers ” when they “claim the authority or state the intention to disregard or decline to enforce all or part of a law...or to

  11. Explicit robust schemes for implementation of general principal value-based constitutive models

    NASA Technical Reports Server (NTRS)

    Arnold, S. M.; Saleeb, A. F.; Tan, H. Q.; Zhang, Y.

    1993-01-01

    The issue of developing effective and robust schemes to implement general hyperelastic constitutive models is addressed. To this end, special purpose functions are used to symbolically derive, evaluate, and automatically generate the associated FORTRAN code for the explicit forms of the corresponding stress function and material tangent stiffness tensors. These explicit forms are valid for the entire deformation range. The analytical form of these explicit expressions is given here for the case in which the strain-energy potential is taken as a nonseparable polynomial function of the principle stretches.

  12. Stability-driven nonnegative matrix factorization to interpret spatial gene expression and build local gene networks.

    PubMed

    Wu, Siqi; Joseph, Antony; Hammonds, Ann S; Celniker, Susan E; Yu, Bin; Frise, Erwin

    2016-04-19

    Spatial gene expression patterns enable the detection of local covariability and are extremely useful for identifying local gene interactions during normal development. The abundance of spatial expression data in recent years has led to the modeling and analysis of regulatory networks. The inherent complexity of such data makes it a challenge to extract biological information. We developed staNMF, a method that combines a scalable implementation of nonnegative matrix factorization (NMF) with a new stability-driven model selection criterion. When applied to a set ofDrosophilaearly embryonic spatial gene expression images, one of the largest datasets of its kind, staNMF identified 21 principal patterns (PP). Providing a compact yet biologically interpretable representation ofDrosophilaexpression patterns, PP are comparable to a fate map generated experimentally by laser ablation and show exceptional promise as a data-driven alternative to manual annotations. Our analysis mapped genes to cell-fate programs and assigned putative biological roles to uncharacterized genes. Finally, we used the PP to generate local transcription factor regulatory networks. Spatially local correlation networks were constructed for six PP that span along the embryonic anterior-posterior axis. Using a two-tail 5% cutoff on correlation, we reproduced 10 of the 11 links in the well-studied gap gene network. The performance of PP with theDrosophiladata suggests that staNMF provides informative decompositions and constitutes a useful computational lens through which to extract biological insight from complex and often noisy gene expression data.

  13. Identification of a Polyomavirus microRNA Highly Expressed in Tumors

    PubMed Central

    Chen, Chun Jung; Cox, Jennifer E.; Azarm, Kristopher; Wylie, Karen N.; Woolard, Kevin D.; Pesavento, Patricia A.; Sullivan, Christopher S.

    2014-01-01

    Polyomaviruses (PyVs) are associated with tumors including Merkel cell carcinoma (MCC). Several PyVs encode microRNAs (miRNAs) but to date no abundant PyV miRNAs have been reported in tumors. To better understand the function of the Merkel cell PyV (MCPyV) miRNA, we examined phylogenetically-related viruses for miRNA expression. We show that two primate PyVs and the more distantly-related raccoon PyV (RacPyV) encode miRNAs that share genomic position and partial sequence identity with MCPyV miRNAs. Unlike MCPyV miRNA in MCC, RacPyV miRNA is highly abundant in raccoon tumors. RacPyV miRNA negatively regulates reporters of early viral (T antigen) transcripts, yet robust viral miRNA expression is tolerated in tumors. We also identify raccoon miRNAs expressed in RacPyV-associated neuroglial brain tumors, including several likely oncogenic miRNAs (oncomiRs). This work describes the first PyV miRNA abundantly expressed in tumors and is consistent with a possible role for both host and viral miRNAs in RacPyV-associated tumors. PMID:25514573

  14. The symbolic constitution of addiction: language, alienation, ambivalence.

    PubMed

    Kemp, Ryan

    2012-07-01

    The author offers an articulation of addiction, via existential-phenomenology and Lacanian psychoanalysis, where it is argued that the addicted subject is constituted via a symbolic structuring evolving from societal practices, laws and the effects of language. Language carries a heritage, which bears on the knowledge and practices of designated subjects and practitioners of that discourse. Addiction, as one particular form of embodied existence and knowledgeable practice, finds expression through the speech and habits of the addict. Addiction, it is argued, is symbolically saturated with ambivalence and alienation. Also the addict is described as the complete modern technocratic subject, consumed by the ideology of consumption. The clinical implications are briefly explored where it is noted that two major approaches to addiction, namely 12-step fellowships and motivational interviewing, both attend to language as a critical component of their treatment approach.

  15. Stability-driven nonnegative matrix factorization to interpret spatial gene expression and build local gene networks

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wu, Siqi; Joseph, Antony; Hammonds, Ann S.

    Spatial gene expression patterns enable the detection of local covariability and are extremely useful for identifying local gene interactions during normal development. The abundance of spatial expression data in recent years has led to the modeling and analysis of regulatory networks. The inherent complexity of such data makes it a challenge to extract biological information. We developed staNMF, a method that combines a scalable implementation of nonnegative matrix factorization (NMF) with a new stability-driven model selection criterion. When applied to a set of Drosophila early embryonic spatial gene expression images, one of the largest datasets of its kind, staNMF identifiedmore » 21 principal patterns (PP). Providing a compact yet biologically interpretable representation of Drosophila expression patterns, PP are comparable to a fate map generated experimentally by laser ablation and show exceptional promise as a data-driven alternative to manual annotations. Our analysis mapped genes to cell-fate programs and assigned putative biological roles to uncharacterized genes. Finally, we used the PP to generate local transcription factor regulatory networks. Spatially local correlation networks were constructed for six PP that span along the embryonic anterior-posterior axis. Using a two-tail 5% cutoff on correlation, we reproduced 10 of the 11 links in the well-studied gap gene network. In conclusion, the performance of PP with the Drosophila data suggests that staNMF provides informative decompositions and constitutes a useful computational lens through which to extract biological insight from complex and often noisy gene expression data.« less

  16. Stability-driven nonnegative matrix factorization to interpret spatial gene expression and build local gene networks

    DOE PAGES

    Wu, Siqi; Joseph, Antony; Hammonds, Ann S.; ...

    2016-04-06

    Spatial gene expression patterns enable the detection of local covariability and are extremely useful for identifying local gene interactions during normal development. The abundance of spatial expression data in recent years has led to the modeling and analysis of regulatory networks. The inherent complexity of such data makes it a challenge to extract biological information. We developed staNMF, a method that combines a scalable implementation of nonnegative matrix factorization (NMF) with a new stability-driven model selection criterion. When applied to a set of Drosophila early embryonic spatial gene expression images, one of the largest datasets of its kind, staNMF identifiedmore » 21 principal patterns (PP). Providing a compact yet biologically interpretable representation of Drosophila expression patterns, PP are comparable to a fate map generated experimentally by laser ablation and show exceptional promise as a data-driven alternative to manual annotations. Our analysis mapped genes to cell-fate programs and assigned putative biological roles to uncharacterized genes. Finally, we used the PP to generate local transcription factor regulatory networks. Spatially local correlation networks were constructed for six PP that span along the embryonic anterior-posterior axis. Using a two-tail 5% cutoff on correlation, we reproduced 10 of the 11 links in the well-studied gap gene network. In conclusion, the performance of PP with the Drosophila data suggests that staNMF provides informative decompositions and constitutes a useful computational lens through which to extract biological insight from complex and often noisy gene expression data.« less

  17. [Etiologic spectrum of solitary constitutional syndrome].

    PubMed

    Hernández Hernández, J L; Matorras Galán, P; Riancho Moral, J A; González-Macías, J

    2002-07-01

    To know the spectrum of diseases responsible for the solitary constitutional syndrome in our setting. This syndrome was defined as a clinical picture characterized by the presence of asthenia, anorexia, and weight loss of at least 5% of body weight in the last six months, not associated with any other symptom or sign suggesting the diagnosis of an organ or system disease. All patients diagnosed of the solitary constitutional syndrome (328) in a tertiary-care level teaching hospital between January 1991 and December 1996. Fifty-two (170) percent of patients with solitary constitutional syndrome were males and 48% (158) females. The mean age was 65.4%, ranging from 15 to 97 years. The average of the monthly estimated weight loss was 3 to 4 kilograms. A total of 115 (35%) malignant neoplasms and 5 (1.5%) benign tumors were diagnosed. The most common malignant tumors corresponded to the digestive tract (51.3% of the total malignant tumors). The second cause in frequency of the solitary constitutional syndrome corresponded to psychiatric diseases, with a total of 80 patients (24.3%). A total of 116 non-neoplastic organic diseases were detected, with digestive tract diseases --mainly peptic disease-- being the most common cause in this group. After follow-up, only in twenty cases were we unable to detect the underlying disease responsible for the syndrome. In nine of these, the solitary constitutional syndrome was self-limited. Forty-four percent of patients had at least another concomitant disease and in 24% of patients more than one associated condition was found. The most common diseases responsible for the solitary constitutional syndrome were, by decreasing frequency, malignant tumors, psychiatric disorders, and non-malignant organic diseases located in the digestive tract. A better knowledge of the etiological spectrum of this syndrome might be useful for a more efficient management of these patients.

  18. From Confederation to Constitution: 1781-1789.

    ERIC Educational Resources Information Center

    Urofsky, Melvin I.; Cox, Nancy

    Students should achieve a higher level of understanding and appreciation of the evolving nature of the U.S. Constitution and its relevance to contemporary societal issues by studying historical documents from the period of time between the Articles of Confederation and the Constitution. This document begins with a history of that period and of the…

  19. Identification of small RNAs abundant in Burkholderia cenocepacia biofilms reveal putative regulators with a potential role in carbon and iron metabolism.

    PubMed

    Sass, Andrea; Kiekens, Sanne; Coenye, Tom

    2017-11-15

    Small RNAs play a regulatory role in many central metabolic processes of bacteria, as well as in developmental processes such as biofilm formation. Small RNAs of Burkholderia cenocepacia, an opportunistic pathogenic beta-proteobacterium, are to date not well characterised. To address that, we performed genome-wide transcriptome structure analysis of biofilm grown B. cenocepacia J2315. 41 unannotated short transcripts were identified in intergenic regions of the B. cenocepacia genome. 15 of these short transcripts, highly abundant in biofilms, widely conserved in Burkholderia sp. and without known function, were selected for in-depth analysis. Expression profiling showed that most of these sRNAs are more abundant in biofilms than in planktonic cultures. Many are also highly abundant in cells grown in minimal media, suggesting they are involved in adaptation to nutrient limitation and growth arrest. Their computationally predicted targets include a high proportion of genes involved in carbon metabolism. Expression and target genes of one sRNA suggest a potential role in regulating iron homoeostasis. The strategy used for this study to detect sRNAs expressed in B. cenocepacia biofilms has successfully identified sRNAs with a regulatory function.

  20. The rcsA Promoter of Pantoea stewartii subsp. stewartii Features a Low-Level Constitutive Promoter and an EsaR Quorum-Sensing-Regulated Promoter

    PubMed Central

    Carlier, Aurelien L.; von Bodman, S. B.

    2006-01-01

    The upstream region of the Pantoea stewartii rcsA gene features two promoters, one for constitutive basal-level expression and a second autoregulated promoter for induced expression. The EsaR quorum-sensing repressor binds to a site centered between the two promoters, blocking transcription elongation from the regulated promoter under noninducing conditions. PMID:16740966

  1. 29 CFR 402.1 - Labor organization constitution and bylaws.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 29 Labor 2 2014-07-01 2014-07-01 false Labor organization constitution and bylaws. 402.1 Section... constitution and bylaws. Every labor organization shall adopt a constitution and bylaws consistent with the... a constitution and bylaws which it has previously adopted and under which it is operating when the...

  2. 29 CFR 402.1 - Labor organization constitution and bylaws.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 29 Labor 2 2012-07-01 2012-07-01 false Labor organization constitution and bylaws. 402.1 Section... constitution and bylaws. Every labor organization shall adopt a constitution and bylaws consistent with the... a constitution and bylaws which it has previously adopted and under which it is operating when the...

  3. 29 CFR 402.1 - Labor organization constitution and bylaws.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 29 Labor 2 2010-07-01 2010-07-01 false Labor organization constitution and bylaws. 402.1 Section... constitution and bylaws. Every labor organization shall adopt a constitution and bylaws consistent with the... a constitution and bylaws which it has previously adopted and under which it is operating when the...

  4. 29 CFR 402.1 - Labor organization constitution and bylaws.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 29 Labor 2 2011-07-01 2011-07-01 false Labor organization constitution and bylaws. 402.1 Section... constitution and bylaws. Every labor organization shall adopt a constitution and bylaws consistent with the... a constitution and bylaws which it has previously adopted and under which it is operating when the...

  5. 29 CFR 402.1 - Labor organization constitution and bylaws.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 29 Labor 2 2013-07-01 2013-07-01 false Labor organization constitution and bylaws. 402.1 Section... constitution and bylaws. Every labor organization shall adopt a constitution and bylaws consistent with the... a constitution and bylaws which it has previously adopted and under which it is operating when the...

  6. Nonlinear Constitutive Relations for High Temperature Application, 1984

    NASA Technical Reports Server (NTRS)

    1985-01-01

    Nonlinear constitutive relations for high temperature applications were discussed. The state of the art in nonlinear constitutive modeling of high temperature materials was reviewed and the need for future research and development efforts in this area was identified. Considerable research efforts are urgently needed in the development of nonlinear constitutive relations for high temperature applications prompted by recent advances in high temperature materials technology and new demands on material and component performance. Topics discussed include: constitutive modeling, numerical methods, material testing, and structural applications.

  7. Novel Gal3 proteins showing altered Gal80p binding cause constitutive transcription of Gal4p-activated genes in Saccharomyces cerevisiae.

    PubMed Central

    Blank, T E; Woods, M P; Lebo, C M; Xin, P; Hopper, J E

    1997-01-01

    Gal4p-mediated activation of galactose gene expression in Saccharomyces cerevisiae normally requires both galactose and the activity of Gal3p. Recent evidence suggests that in cells exposed to galactose, Gal3p binds to and inhibits Ga180p, an inhibitor of the transcriptional activator Gal4p. Here, we report on the isolation and characterization of novel mutant forms of Gal3p that can induce Gal4p activity independently of galactose. Five mutant GAL3(c) alleles were isolated by using a selection demanding constitutive expression of a GAL1 promoter-driven HIS3 gene. This constitutive effect is not due to overproduction of Gal3p. The level of constitutive GAL gene expression in cells bearing different GAL3(c) alleles varies over more than a fourfold range and increases in response to galactose. Utilizing glutathione S-transferase-Gal3p fusions, we determined that the mutant Gal3p proteins show altered Gal80p-binding characteristics. The Gal3p mutant proteins differ in their requirements for galactose and ATP for their Gal80p-binding ability. The behavior of the novel Gal3p proteins provides strong support for a model wherein galactose causes an alteration in Gal3p that increases either its ability to bind to Gal80p or its access to Gal80p. With the Gal3p-Gal80p interaction being a critical step in the induction process, the Gal3p proteins constitute an important new reagent for studying the induction mechanism through both in vivo and in vitro methods. PMID:9111326

  8. 77 FR 57981 - Constitution Day and Citizenship Day, Constitution Week, 2012

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-09-18

    ... than two centuries, our country has drawn enterprising men and women from around the world--individuals... on the basic rights and responsibilities of citizenship, the founding documents from which they were... Constitution, and reaffirm our commitment to the rights and obligations of citizenship in this great Nation...

  9. 29 CFR 452.109 - Constitution of labor organization.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 29 Labor 2 2013-07-01 2013-07-01 false Constitution of labor organization. 452.109 Section 452.109... AND DISCLOSURE ACT OF 1959 Election Procedures; Rights of Members § 452.109 Constitution of labor organization. Elections must be conducted in accordance with the constitution and bylaws of the organization...

  10. 29 CFR 452.109 - Constitution of labor organization.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 29 Labor 2 2010-07-01 2010-07-01 false Constitution of labor organization. 452.109 Section 452.109... AND DISCLOSURE ACT OF 1959 Election Procedures; Rights of Members § 452.109 Constitution of labor organization. Elections must be conducted in accordance with the constitution and bylaws of the organization...

  11. 29 CFR 452.109 - Constitution of labor organization.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 29 Labor 2 2014-07-01 2014-07-01 false Constitution of labor organization. 452.109 Section 452.109... AND DISCLOSURE ACT OF 1959 Election Procedures; Rights of Members § 452.109 Constitution of labor organization. Elections must be conducted in accordance with the constitution and bylaws of the organization...

  12. 29 CFR 452.109 - Constitution of labor organization.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 29 Labor 2 2012-07-01 2012-07-01 false Constitution of labor organization. 452.109 Section 452.109... AND DISCLOSURE ACT OF 1959 Election Procedures; Rights of Members § 452.109 Constitution of labor organization. Elections must be conducted in accordance with the constitution and bylaws of the organization...

  13. 29 CFR 452.109 - Constitution of labor organization.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 29 Labor 2 2011-07-01 2011-07-01 false Constitution of labor organization. 452.109 Section 452.109... AND DISCLOSURE ACT OF 1959 Election Procedures; Rights of Members § 452.109 Constitution of labor organization. Elections must be conducted in accordance with the constitution and bylaws of the organization...

  14. James Madison's "Public" As Interpreter of the Constitution.

    ERIC Educational Resources Information Center

    Dewey, Donald O.

    James Madison's thoughts on various interpretations of the Constitution maintain that public opinion is the ultimate method of legitimizing the document. The Constitution must prevail against mere public opinion, but public opinion may be used to establish the meaning of the Constitution when conflicting interpretations exist. The public good and…

  15. Ectopic ERK Expression Induces Phenotypic Conversion of C10 Cells and Alters DNA Methyltransferase Expression

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Sontag, Ryan L.; Weber, Thomas J.

    2012-05-04

    In some model systems constitutive extracellular signal regulated kinase (ERK) activation is sufficient to promote an oncogenic phenotype. Here we investigate whether constitutive ERK expression influences phenotypic conversion in murine C10 type II alveolar epithelial cells. C10 cells were stably transduced with an ERK1-green fluorescent protein (ERK1-GFP) chimera or empty vector and ectopic ERK expression was associated with the acquisition of soft agar focus-forming potential in late passage, but not early passage cells. Late passage ERK1-GFP cells exhibited a significant increase in the expression of DNA methyl transferases (DNMT1 and 3b) and a marked increase in sensitivity to 5-azacytidine (5-azaC)-mediatedmore » toxicity, relative to early passage ERK1-GFP cells and vector controls. The expression of xeroderma pigmentosum complementation group A (XPA) and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) were significantly increased in late passage cells, suggesting enhanced DNA damage recognition and repair activity which we interpret as a reflection of genomic instability. Phospho-ERK levels were dramatically decreased in late passage ERK1-GFP cells, relative to early passage and vector controls, and phospho-ERK levels were restored by treatment with sodium orthovanadate, indicating a role for phosphatase activity in this response. Collectively these observations suggest that ectopic ERK expression promotes phenotypic conversion of C10 cells that is associated with latent effects on epigenetic programming and phosphatase activities.« less

  16. The Constitutional Heritage.

    ERIC Educational Resources Information Center

    Baxter, Maurice

    Changing political, social, economic, and intellectual conditions over the past two hundred years have demanded innovation and adjustment of legal doctrine, thus giving the United States Constitution a character which the framers of the document could not have predicted. Historically, one must not only understand developments since 1787 but also…

  17. Health care law versus constitutional law.

    PubMed

    Hall, Mark A

    2013-04-01

    National Federation of Independent Business v. Sebelius, the Supreme Court's ruling on the Patient Protection and Affordable Care Act, is a landmark decision - both for constitutional law and for health care law and policy. Others will study its implications for constitutional limits on a range of federal powers beyond health care. This article considers to what extent the decision is also about health care law, properly conceived. Under one view, health care law is the subdiscipline that inquires how courts and government actors take account of the special features of medicine that make legal or policy issues especially problematic - rather than regarding health care delivery and finance more generically, like most any other economic or social enterprise. Viewed this way, the opinions from the Court's conservative justices are mainly about general constitutional law principles. In contrast, Justice Ruth Bader Ginsburg's dissenting opinion for the four more liberal justices is just as much about health care law as it is about constitutional law. Her opinion gives detailed attention to the unique features of health care finance and delivery in order to inform her analysis of constitutional precedents and principles. Thus, the Court's multiple opinions give a vivid depiction of the compelling contrasts between communal versus individualistic conceptions of caring for those in need, and between health care and health insurance as ordinary commodities versus ones that merit special economic, social, and legal status.

  18. Constitution Day: Start the School Year with Civics

    ERIC Educational Resources Information Center

    Lesser, Eli J.

    2006-01-01

    The celebration and recognition of Constitution Day on September 17th is now required by federal law. The new law, known colloquially as the Byrd Amendment, requires all schools receiving any federal assistance, from kindergarten to higher education, to teach students about the Constitution on Constitution Day. The concept of a school's civic…

  19. Constitutional Provisions on the Press: A World View.

    ERIC Educational Resources Information Center

    Paraschos, Manny

    A study examined the legal treatment of the press in constitutions or other basic legal institutional documents from around the world. Sixty-three constitutions or basic documents from the Western World, the Communist Bloc, the Middle East, Africa, Southeast Asia, and Latin America were analyzed. Analysis revealed that most constitutions open with…

  20. 42 CFR 23.28 - What events constitute default?

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 42 Public Health 1 2012-10-01 2012-10-01 false What events constitute default? 23.28 Section 23.28... SERVICE CORPS Private Practice Special Loans for Former Corps Members § 23.28 What events constitute default? The following events will constitute defaults of the loan agreement: (a) Failure to make full...

  1. 42 CFR 23.28 - What events constitute default?

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 42 Public Health 1 2013-10-01 2013-10-01 false What events constitute default? 23.28 Section 23.28... SERVICE CORPS Private Practice Special Loans for Former Corps Members § 23.28 What events constitute default? The following events will constitute defaults of the loan agreement: (a) Failure to make full...

  2. 42 CFR 23.28 - What events constitute default?

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 42 Public Health 1 2010-10-01 2010-10-01 false What events constitute default? 23.28 Section 23.28... SERVICE CORPS Private Practice Special Loans for Former Corps Members § 23.28 What events constitute default? The following events will constitute defaults of the loan agreement: (a) Failure to make full...

  3. 42 CFR 23.28 - What events constitute default?

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 42 Public Health 1 2014-10-01 2014-10-01 false What events constitute default? 23.28 Section 23.28... SERVICE CORPS Private Practice Special Loans for Former Corps Members § 23.28 What events constitute default? The following events will constitute defaults of the loan agreement: (a) Failure to make full...

  4. Constitutional and acquired autosomal aneuploidy.

    PubMed

    Jackson-Cook, Colleen

    2011-12-01

    Chromosomal imbalances can result from numerical or structural anomalies. Numerical chromosomal abnormalities are often referred to as aneuploid conditions. This article focuses on the occurrence of constitutional and acquired autosomal aneuploidy in humans. Topics covered include frequency, mosaicism, phenotypic findings, and etiology. The article concludes with a consideration of anticipated advances that might allow for the development of screening tests and/or lead to improvements in our understanding and management of the role that aneuploidy plays in the aging process and acquisition of age-related and constitutional conditions.

  5. Altered Expression of Porcine Piwi Genes and piRNA during Development

    PubMed Central

    Kowalczykiewicz, Dorota; Pawlak, Piotr; Lechniak, Dorota; Wrzesinski, Jan

    2012-01-01

    Three Sus scrofa Piwi genes (Piwil1, Piwil2 and Piwil4) encoding proteins of 861, 985 and 853 aminoacids, respectively, were cloned and sequenced. Alignment of the Piwi proteins showed the high identity between Sus scrofa and Homo sapiens. Relative transcript abundance of porcine Piwil1, Piwil2 and Piwil4 genes in testes, ovaries and oocytes derived from sexually immature and mature animals was examined using Real-Time PCR. Expression of the three Piwi mRNAs was proved to be tissue specific and restricted exclusively to the gonads. In testes of adult pigs the highest relative transcript abundance was observed for the Sus scrofa Piwil1 gene. On the other hand, in testes of neonatal pigs the Piwil1 transcript level was over 2–fold reduced while the level of Piwil2 transcript was higher. As regards the expression of the Piwil4 transcript, its level was 34-fold elevated in testes of neonatal piglet when compared to adult male. In ovaries of prepubertal and pubertal female pigs transcript abundance of the three Piwi genes was significantly reduced in comparison with testes. However, similarly to testes, in ovaries of neonatal pigs the Piwil2 gene was characterized by the highest relative transcript abundance among the three Piwi genes analysed. In prepubertal and pubertal oocytes Piwil1 transcript was the most abundant whereas the expression of Piwil4 was undetectable. We also demonstrated that expression of piRNA occurs preferentially in the gonads of adult male and female pigs. Moreover, a piRNA subset isolated from ovaries was 2–3 nucleotides longer than the piRNA from testes. PMID:22952772

  6. A constitutive theory of reacting electrolyte mixtures

    NASA Astrophysics Data System (ADS)

    Costa Reis, Martina; Wang, Yongqi; Bono Maurizio Sacchi Bassi, Adalberto

    2013-11-01

    A constitutive theory of reacting electrolyte mixtures is formulated. The intermolecular interactions among the constituents of the mixture are accounted for through additional freedom degrees to each constituent of the mixture. Balance equations for polar reacting continuum mixtures are accordingly formulated and a proper set of constitutive equations is derived with basis in the Müller-Liu formulation of the second law of thermodynamics. Moreover, the non-equilibrium and equilibrium responses of the reacting mixture are investigated in detail by emphasizing the inner and reactive structures of the medium. From the balance laws and constitutive relations, the effects of molecular structure of constituents upon the fluid flow are studied. It is also demonstrated that the local thermodynamic equilibrium state can be reached without imposing that the set of independent constitutive variables is time independent, neither spatially homogeneous nor null. The resulting constitutive relations presented throughout this work are of relevance to many practical applications, such as swelling of clays, developing of bio and polymeric membranes, and use of electrorheological fluids in industrial processes. The first author acknowledges financial support from National Counsel of Technological and Scientific Development (CNPq) and German Academic Exchange Service (DAAD).

  7. A constitutively expressed pair of rpoE2-chrR2 in Azospirillum brasilense Sp7 is required for survival under antibiotic and oxidative stress.

    PubMed

    Gupta, Namrata; Kumar, Santosh; Mishra, Mukti Nath; Tripathi, Anil Kumar

    2013-02-01

    Extracytoplasmic function (ECF) sigma factors (σ(E)) are known to bring about changes in gene expression to enable bacteria to adapt to different stresses. The Azospirillum brasilense Sp245 genome harbours nine genes encoding σ(E), of which two are adjacent to the genes encoding ChrR-type zinc-binding anti-sigma (ZAS) factors. We describe here the role and regulation of a new pair of rpoE-chrR, which was found in the genome of A. brasilense Sp7 in addition to the previously described rpoE-chrR pair (designated rpoE1-chrR1). The rpoE2-chrR2 pair is also cotranscribed, and their products show protein-protein interaction. The -10 and -35 promoter elements of rpoE2-chrR2 and rpoE1-chrR1 were similar but not identical. Unlike the promoter of rpoE1-chrR1, the rpoE2-chrR2 promoter was neither autoregulated nor induced by oxidative stress. Inactivation of chrR2 or overexpression of rpoE2 in A. brasilense Sp7 resulted in an overproduction of carotenoids. It also conferred resistance to oxidative stresses and antibiotics. By controlling the synthesis of carotenoids, initiation and elongation of translation, protein folding and purine biosynthesis, RpoE2 seems to play a crucial role in preventing and repairing the cellular damage caused by oxidative stress. Lack of autoregulation and constitutive expression of rpoE2-chrR2 suggest that RpoE2-ChrR2 may provide a rapid mechanism to cope with oxidative stress, wherein singlet oxygen ((1)O(2))-mediated dissociation of the RpoE2-ChrR2 complex might release RpoE2 to drive the expression of its target genes.

  8. Constitutive polymorphic cyanogenesis in the Australian rainforest tree, Ryparosa kurrangii (Achariaceae).

    PubMed

    Webber, Bruce L; Miller, Rebecca E; Woodrow, Ian E

    2007-08-01

    Cyanogenesis, the liberation of volatile hydrogen cyanide from endogenous cyanide-containing compounds, is a proven plant defence mechanism and the particular cyanogens involved have taxonomic utility. The cyclopentenoncyanhydrin glycoside gynocardin was the only cyanogen isolated from foliar tissue of the rare Australian rainforest tree, Ryparosa kurrangii (Achariaceae). Mechanical damage simulating foliar herbivory did not induce a significant increase in the expression of cyanogenesis over a 24h period, indicating cyanogenic herbivore defence in R. kurrangii is constitutive. The cyanogenic potential of mature leaves was quantitatively polymorphic between trees in a natural population, ranging from 0.54 to 4.77 mg CN g(-1) dry wt leaf tissue.

  9. Expression of the Maize Dof1 Transcription Factor in Wheat and Sorghum

    PubMed Central

    Peña, Pamela A.; Quach, Truyen; Sato, Shirley; Ge, Zhengxiang; Nersesian, Natalya; Changa, Taity; Dweikat, Ismail; Soundararajan, Madhavan; Clemente, Tom E.

    2017-01-01

    Nitrogen is essential for plant growth and development. Improving the ability of plants to acquire and assimilate nitrogen more efficiently is a key agronomic parameter that will augment sustainability in agriculture. A transcription factor approach was pursued to address improvement of nitrogen use efficiency in two major commodity crops. To this end, the Zea mays Dof1 (ZmDof1) transcription factor was expressed in both wheat (Triticum aestivum) and sorghum (Sorghum bicolor) either constitutively, UBI4 promoter from sugarcane, or in a tissue specific fashion via the maize rbcS1 promoter. The primary transcription activation target of ZmDof1, phosphoenolpyruvate carboxylase (PEPC), is observed in transgenic wheat events. Expression ZmDof1 under control of the rbcs1 promoter translates to increase in biomass and yield components in wheat. However, constitutive expression of ZmDof1 led to the down-regulation of genes involved in photosynthesis and the functional apparatus of chloroplasts, and an outcome that negatively impacts photosynthesis, height, and biomass in wheat. Similar patterns were also observed in sorghum transgenic events harboring the constitutive expression cassette of ZmDof1. These results indicate that transcription factor strategies to boost agronomic phenotypic outcomes in crops need to consider expression patterns of the genetic elements to be introduced. PMID:28424717

  10. NCBI GEO: mining millions of expression profiles--database and tools.

    PubMed

    Barrett, Tanya; Suzek, Tugba O; Troup, Dennis B; Wilhite, Stephen E; Ngau, Wing-Chi; Ledoux, Pierre; Rudnev, Dmitry; Lash, Alex E; Fujibuchi, Wataru; Edgar, Ron

    2005-01-01

    The Gene Expression Omnibus (GEO) at the National Center for Biotechnology Information (NCBI) is the largest fully public repository for high-throughput molecular abundance data, primarily gene expression data. The database has a flexible and open design that allows the submission, storage and retrieval of many data types. These data include microarray-based experiments measuring the abundance of mRNA, genomic DNA and protein molecules, as well as non-array-based technologies such as serial analysis of gene expression (SAGE) and mass spectrometry proteomic technology. GEO currently holds over 30,000 submissions representing approximately half a billion individual molecular abundance measurements, for over 100 organisms. Here, we describe recent database developments that facilitate effective mining and visualization of these data. Features are provided to examine data from both experiment- and gene-centric perspectives using user-friendly Web-based interfaces accessible to those without computational or microarray-related analytical expertise. The GEO database is publicly accessible through the World Wide Web at http://www.ncbi.nlm.nih.gov/geo.

  11. METHANE AND NITROGEN ABUNDANCES ON PLUTO AND ERIS

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tegler, S. C.; Cornelison, D. M.; Abernathy, M. R.

    We present spectra of Eris from the MMT 6.5 m Telescope and Red Channel Spectrograph (5700-9800 A, 5 A pixel{sup -1}) on Mt. Hopkins, AZ, and of Pluto from the Steward Observatory 2.3 m Telescope and Boller and Chivens Spectrograph (7100-9400 A, 2 A pixel{sup -1}) on Kitt Peak, AZ. In addition, we present laboratory transmission spectra of methane-nitrogen and methane-argon ice mixtures. By anchoring our analysis in methane and nitrogen solubilities in one another as expressed in the phase diagram of Prokhvatilov and Yantsevich, and comparing methane bands in our Eris and Pluto spectra and methane bands in ourmore » laboratory spectra of methane and nitrogen ice mixtures, we find Eris' bulk methane and nitrogen abundances are {approx}10% and {approx}90% and Pluto's bulk methane and nitrogen abundances are {approx}3% and {approx}97%. Such abundances for Pluto are consistent with values reported in the literature. It appears that the bulk volatile composition of Eris is similar to the bulk volatile composition of Pluto. Both objects appear to be dominated by nitrogen ice. Our analysis also suggests, unlike previous work reported in the literature, that the methane and nitrogen stoichiometry is constant with depth into the surface of Eris. Finally, we point out that our Eris spectrum is also consistent with a laboratory ice mixture consisting of 40% methane and 60% argon. Although we cannot rule out an argon-rich surface, it seems more likely that nitrogen is the dominant species on Eris because the nitrogen ice 2.15 {mu}m band is seen in spectra of Pluto and Triton.« less

  12. Conservation of protein abundance patterns reveals the regulatory architecture of the EGFR-MAPK pathway

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Shi, T.; Niepel, M.; McDermott, J. E.

    It is not known whether cancer cells generally show quantitative differences in the expression of signaling pathway proteins that could dysregulate signal transduction. To explore this issue, we first defined the primary components of the EGF-MAPK pathway in normal human mammary epithelial cells, identifying 16 core proteins and 10 feedback regulators. We then quantified their absolute abundance across a panel of normal and cancer cell lines. We found that core pathway proteins were expressed at very similar levels across all cell types. In contrast, the EGFR and transcriptionally controlled feedback regulators were expressed at highly variable levels. The absolute abundancemore » of most core pathway proteins was between 50,000- 70,000 copies per cell, but the adaptors SOS1, SOS2, and GAB1 were found at far lower levels (2,000-5,000 per cell). MAPK signaling showed saturation in all cells between 3,000-10,000 occupied EGFR, consistent with the idea that low adaptor levels limit signaling. Our results suggest that the core MAPK pathway is essentially invariant across different cell types, with cell- specific differences in signaling likely due to variable levels of feedback regulators. The low abundance of adaptors relative to the EGFR could be responsible for previous observation of saturable signaling, endocytosis, and high affinity EGFR.« less

  13. Identification and Characterization of 40 Isolated Rehmannia glutinosa MYB Family Genes and Their Expression Profiles in Response to Shading and Continuous Cropping

    PubMed Central

    Wang, Fengqing; Suo, Yanfei; Wei, He; Li, Mingjie; Xie, Caixia; Wang, Lina; Chen, Xinjian; Zhang, Zhongyi

    2015-01-01

    The v-myb avian myeloblastosis viral oncogene homolog (MYB) superfamily constitutes one of the most abundant groups of transcription factors (TFs) described in plants. To date, little is known about the MYB genes in Rehmannia glutinosa. Forty unique MYB genes with full-length cDNA sequences were isolated. These 40 genes were grouped into five categories, one R1R2R3-MYB, four TRFL MYBs, four SMH MYBs, 25 R2R3-MYBs, and six MYB-related members. The MYB DNA-binding domain (DBD) sequence composition was conserved among proteins of the same subgroup. As expected, most of the closely related members in the phylogenetic tree exhibited common motifs. Additionally, the gene structure and motifs of the R. glutinosa MYB genes were analyzed. MYB gene expression was analyzed in the leaf and the tuberous root under two abiotic stress conditions. Expression profiles showed that most R. glutinosa MYB genes were expressed in the leaf and the tuberous root, suggesting that MYB genes are involved in various physiological and developmental processes in R. glutinosa. Seven MYB genes were up-regulated in response to shading in at least one tissue. Two MYB genes showed increased expression and 13 MYB genes showed decreased expression in the tuberous root under continuous cropping. This investigation is the first comprehensive study of the MYB gene family in R. glutinosa. PMID:26147429

  14. Development of an IP-Free Biotechnology Platform for Constitutive Production of HPV16 L1 Capsid Protein Using the Pichia pastoris PGK1 Promoter.

    PubMed

    Mariz, F C; Coimbra, E C; Jesus, A L S; Nascimento, L M; Torres, F A G; Freitas, A C

    2015-01-01

    The human papillomavirus (HPV) L1 major capsid protein, which forms the basis of the currently available vaccines against cervical cancer, self-assembles into virus-like particles (VLPs) when expressed heterologously. We report the development of a biotechnology platform for HPV16 L1 protein expression based on the constitutive PGK1 promoter (PPGK1) from the methylotrophic yeast Pichia pastoris. The L1 gene was cloned under regulation of PPGK1 into pPGKΔ3 expression vector to achieve intracellular expression. In parallel, secretion of the L1 protein was obtained through the use of an alternative vector called pPGKΔ3α, in which a codon optimized α-factor signal sequence was inserted. We devised a work-flow based on the detection of the L1 protein by dot blot, colony blot, and western blot to classify the positive clones. Finally, intracellular HPV VLPs assembly was demonstrated for the first time in yeast cells. This study opens up perspectives for the establishment of an innovative platform for the production of HPV VLPs or other viral antigens for vaccination purposes, based on constitutive expression in P. pastoris.

  15. Relative abundance of total subgingival plaque-specific bacteria in salivary microbiota reflects the overall periodontal condition in patients with periodontitis.

    PubMed

    Kageyama, Shinya; Takeshita, Toru; Asakawa, Mikari; Shibata, Yukie; Takeuchi, Kenji; Yamanaka, Wataru; Yamashita, Yoshihisa

    2017-01-01

    Increasing attention is being focused on evaluating the salivary microbiota as a promising method for monitoring oral health; however, its bacterial composition greatly differs from that of dental plaque microbiota, which is a dominant etiologic factor of oral diseases. This study evaluated the relative abundance of subgingival plaque-specific bacteria in the salivary microbiota and examined a relationship between the abundance and severity of periodontal condition in patients with periodontitis. Four samples (subgingival and supragingival plaques, saliva, and tongue coating) per each subject were collected from 14 patients with a broad range of severity of periodontitis before periodontal therapy. The bacterial composition was analyzed by 16S rRNA gene amplicon sequencing using Ion PGM. Of the 66 species-level operational taxonomic units (OTUs) representing the mean relative abundance of ≥ 1% in any of the four niches, 12 OTUs corresponding to known periodontal pathogens, including Porphyromonas gingivalis, were characteristically predominant in the subgingival plaque and constituted 37.3 ± 22.9% of the microbiota. The total relative abundance of these OTUs occupied only 1.6 ± 1.2% of the salivary microbiota, but significantly correlated with the percentage of diseased sites (periodontal pocket depth ≥ 4 mm; r = 0.78, P < 0.001), in addition to the abundance of subgingival plaque microbiota (r = 0.61, P = 0.02). After periodontal therapy, the total relative abundance of these 12 OTUs was evaluated as well as before periodontal therapy and reductions of the abundance through periodontal therapy were strongly correlated in saliva and subgingival plaque (r = 0.81, P < 0.001). Based on these results, salivary microbiota might be a promising target for the evaluation of subgingival plaque-derived bacteria representing the present condition of periodontal health.

  16. Relative abundance of total subgingival plaque-specific bacteria in salivary microbiota reflects the overall periodontal condition in patients with periodontitis

    PubMed Central

    Kageyama, Shinya; Takeshita, Toru; Asakawa, Mikari; Shibata, Yukie; Takeuchi, Kenji; Yamanaka, Wataru

    2017-01-01

    Increasing attention is being focused on evaluating the salivary microbiota as a promising method for monitoring oral health; however, its bacterial composition greatly differs from that of dental plaque microbiota, which is a dominant etiologic factor of oral diseases. This study evaluated the relative abundance of subgingival plaque-specific bacteria in the salivary microbiota and examined a relationship between the abundance and severity of periodontal condition in patients with periodontitis. Four samples (subgingival and supragingival plaques, saliva, and tongue coating) per each subject were collected from 14 patients with a broad range of severity of periodontitis before periodontal therapy. The bacterial composition was analyzed by 16S rRNA gene amplicon sequencing using Ion PGM. Of the 66 species-level operational taxonomic units (OTUs) representing the mean relative abundance of ≥ 1% in any of the four niches, 12 OTUs corresponding to known periodontal pathogens, including Porphyromonas gingivalis, were characteristically predominant in the subgingival plaque and constituted 37.3 ± 22.9% of the microbiota. The total relative abundance of these OTUs occupied only 1.6 ± 1.2% of the salivary microbiota, but significantly correlated with the percentage of diseased sites (periodontal pocket depth ≥ 4 mm; r = 0.78, P < 0.001), in addition to the abundance of subgingival plaque microbiota (r = 0.61, P = 0.02). After periodontal therapy, the total relative abundance of these 12 OTUs was evaluated as well as before periodontal therapy and reductions of the abundance through periodontal therapy were strongly correlated in saliva and subgingival plaque (r = 0.81, P < 0.001). Based on these results, salivary microbiota might be a promising target for the evaluation of subgingival plaque-derived bacteria representing the present condition of periodontal health. PMID:28369125

  17. Constitutive expression of two apple (Malus x domestica Borkh.) homolog genes of LIKE HETEROCHROMATIN PROTEIN1 affects flowering time and whole-plant growth in transgenic Arabidopsis.

    PubMed

    Mimida, Naozumi; Kidou, Shin-Ichiro; Kotoda, Nobuhiro

    2007-09-01

    Fruit trees, such as apple (Malus x domestica Borkh.), are woody perennial plants with a long juvenile phase. The biological analysis for the regulation of flowering time provides insights into the reduction of juvenile phase and the acceleration of breeding in fruit trees. In Arabidopsis, LIKE HETEROCHROMATIN PROTEIN1 (LHP1) is involved in epigenetic silencing of the target genes such as flowering genes. We isolated and characterized twin apple LHP1 homolog genes, MdLHP1a and MdLHP1b. These genes may have been generated as a result of ancient genome duplication. Although the putative MdLHP1 proteins showed lower similarity to any other known plant LHP1 homologs, a chromo domain, a chromo shadow domain, and the nuclear localization signal motifs were highly conserved among them. RT-PCR analysis showed that MdLHP1a and MdLHP1b were expressed constantly in developing shoot apices of apple trees throughout the growing season. Constitutive expression of MdLHP1a or MdLHP1b could compensate for the pleiotropic phenotype of lhp1/tfl2 mutant, suggesting that apple LHP1 homolog genes are involved in the regulation of flowering time and whole-plant growth. Based on these results, LHP1 homolog genes might have rapidly evolved among plant species, but the protein functions were conserved, at least between Arabidopsis and apple.

  18. Bacterial 16S rRNA gene analysis revealed that bacteria related to Arcobacter spp. constitute an abundant and common component of the oyster microbiota (Tiostrea chilensis).

    PubMed

    Romero, J; García-Varela, M; Laclette, J P; Espejo, R T

    2002-11-01

    To explore the bacterial microbiota in Chilean oyster (Tiostrea chilensis), a molecular approach that permits detection of different bacteria, independently of their capacity to grow in culture media, was used. Bacterial diversity was assessed by analysis of both the 16S rDNA and the 16S-23S intergenic region, obtained by PCR amplifications of DNA extracted from depurated oysters. RFLP of the PCR amplified 16S rDNA showed a prevailing pattern in most of the individuals analyzed, indicating that a few bacterial species were relatively abundant and common in oysters. Cloning and sequencing of the 16S rDNA with the prevailing RFLP pattern indicated that this rRNA was most closely related to Arcobacter spp. However, analysis by the size of the amplified 16S-23S rRNA intergenic regions revealed not Arcobacter spp. but Staphylococcus spp. related bacteria as a major and common component in oyster. These different results may be caused by the absence of target for one of the primers employed for amplification of the intergenic region. Neither of the two bacteria species found in large abundance was recovered after culturing under aerobic, anaerobic, or microaerophilic conditions. This result, however, is expected because the number of bacteria recovered after cultivation was less than 0.01% of the total. All together, these observations suggest that Arcobacter-related strains are probably abundant and common in the Chilean oyster bacterial microbiota.

  19. Constitutive STAT5 Activation Correlates With Better Survival in Cervical Cancer Patients Treated With Radiation Therapy

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chen, Helen H.W.; Institute of Clinical Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan; Chou, Cheng-Yang

    2012-02-01

    Purpose: Constitutively activated signal transducers and activators of transcription (STAT) factors, in particular STAT1, STAT3, and STAT5, have been detected in a wide variety of human primary tumors and have been demonstrated to directly contribute to oncogenesis. However, the expression pattern of these STATs in cervical carcinoma is still unknown, as is whether or not they have prognostic significance. This study investigated the expression patterns of STAT1, STAT3, and STAT5 in cervical cancer and their associations with clinical outcomes in patients treated with radical radiation therapy. Methods and Materials: A total of 165 consecutive patients with International Federation of Gynecologymore » and Obstetrics (FIGO) Stages IB to IVA cervical cancer underwent radical radiation therapy, including external beam and/or high-dose-rate brachytherapy between 1989 and 2002. Immunohistochemical studies of their formalin-fixed, paraffin-embedded tissues were performed. Univariate and multivariate analyses were performed to identify and to evaluate the effects of these factors affecting patient survival. Results: Constitutive activations of STAT1, STAT3, and STAT5 were observed in 11%, 22%, and 61% of the participants, respectively. While STAT5 activation was associated with significantly better metastasis-free survival (p < 0.01) and overall survival (p = 0.04), STAT1 and STAT3 activation were not. Multivariate analyses showed that STAT5 activation, bulky tumor ({>=}4 cm), advanced stage (FIGO Stages III and IV), and brachytherapy (yes vs. no) were independent prognostic factors for cause-specific overall survival. None of the STATs was associated with local relapse. STAT5 activation (odds ratio = 0.29, 95% confidence interval = 0.13-0.63) and advanced stage (odds ratio = 2.54; 95% confidence interval = 1.03-6.26) were independent predictors of distant metastasis. Conclusions: This is the first report to provide the overall expression patterns and prognostic

  20. Linking species abundance distributions in numerical abundance and biomass through simple assumptions about community structure.

    PubMed

    Henderson, Peter A; Magurran, Anne E

    2010-05-22

    Species abundance distributions (SADs) are widely used as a tool for summarizing ecological communities but may have different shapes, depending on the currency used to measure species importance. We develop a simple plotting method that links SADs in the alternative currencies of numerical abundance and biomass and is underpinned by testable predictions about how organisms occupy physical space. When log numerical abundance is plotted against log biomass, the species lie within an approximately triangular region. Simple energetic and sampling constraints explain the triangular form. The dispersion of species within this triangle is the key to understanding why SADs of numerical abundance and biomass can differ. Given regular or random species dispersion, we can predict the shape of the SAD for both currencies under a variety of sampling regimes. We argue that this dispersion pattern will lie between regular and random for the following reasons. First, regular dispersion patterns will result if communities are comprised groups of organisms that use different components of the physical space (e.g. open water, the sea bed surface or rock crevices in a marine fish assemblage), and if the abundance of species in each of these spatial guilds is linked to the way individuals of varying size use the habitat. Second, temporal variation in abundance and sampling error will tend to randomize this regular pattern. Data from two intensively studied marine ecosystems offer empirical support for these predictions. Our approach also has application in environmental monitoring and the recognition of anthropogenic disturbance, which may change the shape of the triangular region by, for example, the loss of large body size top predators that occur at low abundance.

  1. Linking species abundance distributions in numerical abundance and biomass through simple assumptions about community structure

    PubMed Central

    Henderson, Peter A.; Magurran, Anne E.

    2010-01-01

    Species abundance distributions (SADs) are widely used as a tool for summarizing ecological communities but may have different shapes, depending on the currency used to measure species importance. We develop a simple plotting method that links SADs in the alternative currencies of numerical abundance and biomass and is underpinned by testable predictions about how organisms occupy physical space. When log numerical abundance is plotted against log biomass, the species lie within an approximately triangular region. Simple energetic and sampling constraints explain the triangular form. The dispersion of species within this triangle is the key to understanding why SADs of numerical abundance and biomass can differ. Given regular or random species dispersion, we can predict the shape of the SAD for both currencies under a variety of sampling regimes. We argue that this dispersion pattern will lie between regular and random for the following reasons. First, regular dispersion patterns will result if communities are comprised groups of organisms that use different components of the physical space (e.g. open water, the sea bed surface or rock crevices in a marine fish assemblage), and if the abundance of species in each of these spatial guilds is linked to the way individuals of varying size use the habitat. Second, temporal variation in abundance and sampling error will tend to randomize this regular pattern. Data from two intensively studied marine ecosystems offer empirical support for these predictions. Our approach also has application in environmental monitoring and the recognition of anthropogenic disturbance, which may change the shape of the triangular region by, for example, the loss of large body size top predators that occur at low abundance. PMID:20071388

  2. Expression of human placental lactogen and variant growth hormone genes in placentas.

    PubMed

    Martinez-Rodriguez, H G; Guerra-Rodriguez, N E; Iturbe-Cantu, M A; Martinez-Torres, A; Barrera-Saldaña, H A

    1997-01-01

    Previous studies comparing the expression levels of human placental lactogen (hPL) genes have shown varying results, due to, perhaps, the fact that in all of them only one placenta was being analyzed. Here, the expression of hPL and growth hormone variant (hGH-V) genes in fifteen term placentas was comparatively analyzed at the RNA level, using reverse transcription coupled to polymerase chain reaction (RT-PCR). The abundance of the combined RNA transcripts derived from these genes varied from one placenta to another. The authors found that hPL-4 transcripts were more abundant than those of hPL-3 in most samples (ratios from 1:1 to 6:1), transcripts from the putative hPL-1 pseudogene were more abundant at the unprocessed stage while those of the hGH-V gene were mostly processed. Again, the authors of this study observed wide variation from placenta to placenta in the abundance of both of these types of transcripts. The same was observed when a group of six placentas from abortuses and nine from pregnancies complicated by preclampsia, diabetes and hypertension was studied. The authors conclude that the disagreeing results reported in the literature which are not in agreement concerning the expression levels of hPL genes could be explained by normal variations of their expression levels among the different placentas analyzed.

  3. The wheat resistance gene Lr34 results in the constitutive induction of multiple defense pathways in transgenic barley.

    PubMed

    Chauhan, Harsh; Boni, Rainer; Bucher, Rahel; Kuhn, Benjamin; Buchmann, Gabriele; Sucher, Justine; Selter, Liselotte L; Hensel, Goetz; Kumlehn, Jochen; Bigler, Laurent; Glauser, Gaëtan; Wicker, Thomas; Krattinger, Simon G; Keller, Beat

    2015-10-01

    The wheat gene Lr34 encodes an ABCG-type transporter which provides durable resistance against multiple pathogens. Lr34 is functional as a transgene in barley, but its mode of action has remained largely unknown both in wheat and barley. Here we studied gene expression in uninfected barley lines transgenic for Lr34. Genes from multiple defense pathways contributing to basal and inducible disease resistance were constitutively active in seedlings and mature leaves. In addition, the hormones jasmonic acid and salicylic acid were induced to high levels, and increased levels of lignin as well as hordatines were observed. These results demonstrate a strong, constitutive re-programming of metabolism by Lr34. The resistant Lr34 allele (Lr34res) encodes a protein that differs by two amino acid polymorphisms from the susceptible Lr34sus allele. The deletion of a single phenylalanine residue in Lr34sus was sufficient to induce the characteristic Lr34-based responses. Combination of Lr34res and Lr34sus in the same plant resulted in a reduction of Lr34res expression by 8- to 20-fold when the low-expressing Lr34res line BG8 was used as a parent. Crosses with the high-expressing Lr34res line BG9 resulted in an increase of Lr34sus expression by 13- to 16-fold in progenies that inherited both alleles. These results indicate an interaction of the two Lr34 alleles on the transcriptional level. Reduction of Lr34res expression in BG8 crosses reduced the negative pleiotropic effects of Lr34res on barley growth and vigor without compromising disease resistance, suggesting that transgenic combination of Lr34res and Lr34sus can result in agronomically useful resistance. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

  4. Upholding Indigenous Freedoms of Religion and Medicine: Peyotists at the 1906-1908 Oklahoma Constitutional Convention and First Legislature

    ERIC Educational Resources Information Center

    Wiedman, Dennis

    2012-01-01

    In the five hundred years of European and American globalization of the world, seldom have Indigenous peoples been invited to a constitutional convention and first legislature to express their perspectives and concerns. Rarely in the five-hundred-year history of the European and American colonization of the world were the rights of the Indigenous…

  5. Improved stress tolerance and productivity in transgenic rice plants constitutively expressing the Oryza sativa glutathione synthetase OsGS under paddy field conditions.

    PubMed

    Park, Seong-Im; Kim, Young-Saeng; Kim, Jin-Ju; Mok, Ji-Eun; Kim, Yul-Ho; Park, Hyang-Mi; Kim, Il-Sup; Yoon, Ho-Sung

    2017-08-01

    Reactive oxygen species, which increase under various environmental stresses, have deleterious effects on plants. An important antioxidant, glutathione, is used to detoxify reactive oxygen species in plant cells and is mainly produced by two enzymes: gamma-glutamylcysteine synthetase (γ-ECS) and glutathione synthetase (GS). To evaluate the functional roles of the glutathione synthetase gene (OsGS) in rice, we generated four independent transgenic rice plants (TG1-TG4) that overexpressed OsGS under the control of the constitutively expressed OsCc1 promoter. When grown under natural paddy field conditions, the TG rice plants exhibited greater growth development, higher chlorophyll content, and higher GSH/GSSH ratios than control wild-type (WT) rice plants. Subsequently, the TG rice plants enhanced redox homeostasis by preventing hydroperoxide-mediated membrane damage, which improved their adaptation to environmental stresses. As a result, TG rice plants improved rice grain yield and total biomass following increases in panicle number and number of spikelets per panicle, despite differences in climate during the cultivation periods of 2014 and 2015. Overall, our results indicate that OsGS overexpression improved redox homeostasis by enhancing the glutathione pool, which resulted in greater tolerance to environmental stresses in the paddy fields. Copyright © 2017. Published by Elsevier GmbH.

  6. Antisense sequences of the nbl gene induce apoptosis in the human promyelocytic leukemia cell line HL-60.

    PubMed

    Naora, H; Nishida, T; Shindo, Y; Adachi, M; Naora, H

    1998-04-01

    Apoptosis is induced by the transcriptional inhibitor actinomycin D (Act D) in various cell types, particularly many leukemic cell lines such as HL-60. A common feature of these cell lines is their high constitutive expression level of the nbl gene, which was originally isolated by virtue of its abundance in a Namalwa Burkitt lymphoma cDNA library. In contrast, cell lines which constitutively express nbl at low levels appear not to undergo typical apoptotic death in response to Act D. Apoptotic induction by Act D in cells which normally express nbl at high levels was found in this study to be closely associated with a decline in nbl mRNA levels, raising the possibility that apoptosis could be induced by lowering nbl expression levels in such cells. Transient expression of nbl antisense sequences in HL-60 cells decreased cell viability, and induced typical apoptotic morphology such as cell shrinkage, chromatin condensation and nuclear fragmentation. Incubation with nbl antisense oligomers also induced similar features in HL-60 cells and in another high nb-expressing cell line, Jurkat, but had little effect in HepG2 cells which constitutively express nbl at low levels. These findings suggest that lowering constitutively high levels of nbl expression can induce apoptosis.

  7. Constitutive activation of CaMKKα signaling is sufficient but not necessary for mTORC1 activation and growth in mouse skeletal muscle.

    PubMed

    Ferey, Jeremie L A; Brault, Jeffrey J; Smith, Cheryl A S; Witczak, Carol A

    2014-10-15

    Skeletal muscle loading/overload stimulates the Ca²⁺-activated, serine/threonine kinase Ca²⁺/calmodulin-dependent protein kinase kinase-α (CaMKKα); yet to date, no studies have examined whether CaMKKα regulates muscle growth. The purpose of this study was to determine if constitutive activation of CaMKKα signaling could stimulate muscle growth and if so whether CaMKKα is essential for this process. CaMKKα signaling was selectively activated in mouse muscle via expression of a constitutively active form of CaMKKα using in vivo electroporation. After 2 wk, constitutively active CaMKKα expression increased muscle weight (~10%) and protein content (~10%), demonstrating that activation of CaMKKα signaling can stimulate muscle growth. To determine if active CaMKKα expression stimulated muscle growth via increased mammalian target of rapamycin complex 1 (mTORC1) signaling and protein synthesis, [³H]phenylalanine incorporation into proteins was assessed with or without the mTORC1 inhibitor rapamycin. Constitutively active CaMKKα increased protein synthesis ~60%, and this increase was prevented by rapamycin, demonstrating a critical role for mTORC1 in this process. To determine if CaMKKα is essential for growth, muscles from CaMKKα knockout mice were stimulated to hypertrophy via unilateral ablation of synergist muscles (overload). Surprisingly, compared with wild-type mice, muscles from CaMKKα knockout mice exhibited greater growth (~15%) and phosphorylation of the mTORC1 substrate 70-kDa ribosomal protein S6 kinase (Thr³⁸⁹; ~50%), demonstrating that CaMKKα is not essential for overload-induced mTORC1 activation or muscle growth. Collectively, these results demonstrate that activation of CaMKKα signaling is sufficient but not necessary for activation of mTORC1 signaling and growth in mouse skeletal muscle. Copyright © 2014 the American Physiological Society.

  8. Constitutive activation of CaMKKα signaling is sufficient but not necessary for mTORC1 activation and growth in mouse skeletal muscle

    PubMed Central

    Ferey, Jeremie L. A.; Brault, Jeffrey J.; Smith, Cheryl A. S.

    2014-01-01

    Skeletal muscle loading/overload stimulates the Ca2+-activated, serine/threonine kinase Ca2+/calmodulin-dependent protein kinase kinase-α (CaMKKα); yet to date, no studies have examined whether CaMKKα regulates muscle growth. The purpose of this study was to determine if constitutive activation of CaMKKα signaling could stimulate muscle growth and if so whether CaMKKα is essential for this process. CaMKKα signaling was selectively activated in mouse muscle via expression of a constitutively active form of CaMKKα using in vivo electroporation. After 2 wk, constitutively active CaMKKα expression increased muscle weight (∼10%) and protein content (∼10%), demonstrating that activation of CaMKKα signaling can stimulate muscle growth. To determine if active CaMKKα expression stimulated muscle growth via increased mammalian target of rapamycin complex 1 (mTORC1) signaling and protein synthesis, [3H]phenylalanine incorporation into proteins was assessed with or without the mTORC1 inhibitor rapamycin. Constitutively active CaMKKα increased protein synthesis ∼60%, and this increase was prevented by rapamycin, demonstrating a critical role for mTORC1 in this process. To determine if CaMKKα is essential for growth, muscles from CaMKKα knockout mice were stimulated to hypertrophy via unilateral ablation of synergist muscles (overload). Surprisingly, compared with wild-type mice, muscles from CaMKKα knockout mice exhibited greater growth (∼15%) and phosphorylation of the mTORC1 substrate 70-kDa ribosomal protein S6 kinase (Thr389; ∼50%), demonstrating that CaMKKα is not essential for overload-induced mTORC1 activation or muscle growth. Collectively, these results demonstrate that activation of CaMKKα signaling is sufficient but not necessary for activation of mTORC1 signaling and growth in mouse skeletal muscle. PMID:25159322

  9. Up-regulation of phosphoinositide metabolism in tobacco cells constitutively expressing the human type I inositol polyphosphate 5-phosphatase

    NASA Technical Reports Server (NTRS)

    Perera, Imara Y.; Love, John; Heilmann, Ingo; Thompson, William F.; Boss, Wendy F.; Brown, C. S. (Principal Investigator)

    2002-01-01

    To evaluate the impact of suppressing inositol 1,4,5-trisphosphate (InsP(3)) in plants, tobacco (Nicotiana tabacum) cells were transformed with the human type I inositol polyphosphate 5-phosphatase (InsP 5-ptase), an enzyme which specifically hydrolyzes InsP(3). The transgenic cell lines showed a 12- to 25-fold increase in InsP 5-ptase activity in vitro and a 60% to 80% reduction in basal InsP(3) compared with wild-type cells. Stimulation with Mas-7, a synthetic analog of the wasp venom peptide mastoparan, resulted in an approximately 2-fold increase in InsP(3) in both wild-type and transgenic cells. However, even with stimulation, InsP(3) levels in the transgenic cells did not reach wild-type basal values, suggesting that InsP(3) signaling is compromised. Analysis of whole-cell lipids indicated that phosphatidylinositol 4,5-bisphosphate (PtdInsP(2)), the lipid precursor of InsP(3), was greatly reduced in the transgenic cells. In vitro assays of enzymes involved in PtdInsP(2) metabolism showed that the activity of the PtdInsP(2)-hydrolyzing enzyme phospholipase C was not significantly altered in the transgenic cells. In contrast, the activity of the plasma membrane PtdInsP 5 kinase was increased by approximately 3-fold in the transgenic cells. In vivo labeling studies revealed a greater incorporation of (32)P into PtdInsP(2) in the transgenic cells compared with the wild type, indicating that the rate of PtdInsP(2) synthesis was increased. These studies show that the constitutive expression of the human type I InsP 5-ptase in tobacco cells leads to an up-regulation of the phosphoinositide pathway and highlight the importance of PtdInsP(2) synthesis as a regulatory step in this system.

  10. Analysis of the Plasma Proteome in COPD: Novel Low Abundance Proteins Reflect the Severity of Lung Remodeling

    PubMed Central

    Merali, Salim; Barrero, Carlos A.; Bowler, Russell P.; Chen, Diane Er; Criner, Gerard; Braverman, Alan; Litwin, Samuel; Yeung, Anthony; Kelsen, Steven G.

    2015-01-01

    The search for COPD biomarkers has largely employed a targeted approach that focuses on plasma proteins involved in the systemic inflammatory response and in lung injury and repair. This proof of concept study was designed to test the idea that an open, unbiased, in-depth proteomics approach could identify novel, low abundance plasma proteins i.e., ng/mL concentration, which could serve as potential biomarkers. Differentially expressed proteins were identified in a discovery group with severe COPD (FEV1 <45% predicted; n = 10). Subjects with normal lung function matched for age, sex, ethnicity and smoking history served as controls (n = 10). Pooled plasma from each group was exhaustively immunodepleted of abundant proteins, d separated by 1-D gel electrophoresis and extensively fractionated prior to LC-tandem mass spectroscopy (GeLC-MS). Thirty one differentially expressed proteins were identified in the discovery group including markers of lung defense against oxidant stress, alveolar macrophage activation, and lung tissue injury and repair. Four of the 31 proteins (i.e., GRP78, soluble CD163, IL1AP and MSPT9) were measured in a separate verification group of 80 subjects with varying COPD severity by immunoassay. All 4 were significantly altered in COPD and 2 (GRP78 and soluble CD163) correlated with both FEV1 and the extent of emphysema. In-depth, plasma proteomic analysis identified a group of novel, differentially expressed, low abundance proteins that reflect known pathogenic mechanisms and the severity of lung remodeling in COPD. These proteins may also prove useful as COPD biomarkers. PMID:24111704

  11. Analysis of the plasma proteome in COPD: Novel low abundance proteins reflect the severity of lung remodeling.

    PubMed

    Merali, Salim; Barrero, Carlos A; Bowler, Russell P; Chen, Diane Er; Criner, Gerard; Braverman, Alan; Litwin, Samuel; Yeung, Anthony; Kelsen, Steven G

    2014-04-01

    The search for COPD biomarkers has largely employed a targeted approach that focuses on plasma proteins involved in the systemic inflammatory response and in lung injury and repair. This proof of concept study was designed to test the idea that an open, unbiased, in-depth proteomics approach could identify novel, low abundance plasma proteins i.e., ng/mL concentration, which could serve as potential biomarkers. Differentially expressed proteins were identified in a discovery group with severe COPD (FEV1 <45% predicted; n = 10). Subjects with normal lung function matched for age, sex, ethnicity and smoking history served as controls (n = 10). Pooled plasma from each group was exhaustively immunodepleted of abundant proteins, d separated by 1-D gel electrophoresis and extensively fractionated prior to LC-tandem mass spectroscopy (GeLC-MS). Thirty one differentially expressed proteins were identified in the discovery group including markers of lung defense against oxidant stress, alveolar macrophage activation, and lung tissue injury and repair. Four of the 31 proteins (i.e., GRP78, soluble CD163, IL1AP and MSPT9) were measured in a separate verification group of 80 subjects with varying COPD severity by immunoassay. All 4 were significantly altered in COPD and 2 (GRP78 and soluble CD163) correlated with both FEV1 and the extent of emphysema. In-depth, plasma proteomic analysis identified a group of novel, differentially expressed, low abundance proteins that reflect known pathogenic mechanisms and the severity of lung remodeling in COPD. These proteins may also prove useful as COPD biomarkers.

  12. An Insulin-Like Growth Factor 1 Receptor Inhibitor Induces CYP3A4 Expression through a Pregnane X Receptor-Independent, Noncanonical Constitutive Androstane Receptor-Related Mechanism

    PubMed Central

    Li, Linhao; Sinz, Michael W.; Zimmermann, Kurt

    2012-01-01

    Inhibition of insulin-like growth factor-1 receptor (IGF-1R) signaling represents an attractive therapeutic strategy for cancer treatment. A first-generation IGF-1R inhibitor (R)-4-(3-(3-chlorophenyl)-3-hydroxypropyl)-3-(4-methyl-6-morpholino-1H-benzo[d]imidazol-2-yl)pyridin-2(1H)-one (BMS-536924), however, was associated with potent CYP3A4 induction mediated by pregnane X receptor (PXR; NR1I2) transactivation. Structural activity-based modification led to the synthesis of 4-(1-(2-(4-((2-(4-chloro-1H-pyrazol-1-yl)ethyl)amino)-2-oxo-1,2-dihydropyridin-3-yl)-4-methyl-1H-benzo[d]imidazol-6-yl)piperidin-4-yl) piperazine-1-carboxylate (BMS-665351) with no PXR activity while maintaining its ability to inhibit IGF-1R. However, BMS-665351 significantly induces CYP3A4 expression in human primary hepatocytes (HPHs). Here, we report a novel nonclassical constitutive androstane receptor (CAR; NR1I3)-related pathway of BMS-665351-mediated CYP3A4 induction. BMS-665351 treatment resulted in the significant induction of CYP3A4 in HPHs and HepG2 cells, but failed to activate either PXR or CAR in cell-based reporter assays. Moreover, BMS-665351 at concentrations that induce CYP3A4 expression was unable to translocate human CAR from the cytoplasm to the nucleus of HPHs, which represents the initial step of CAR activation. Nevertheless, quantitative polymerase chain reaction analysis demonstrated that BMS-665351 significantly enhanced the expression of CYP3A4 in CAR- but not PXR-transfected HepG2 and Huh7 cells. It is noteworthy that BMS-665351 selectively induced the expression of CAR but not PXR in all tested hepatic cell systems. Synergistic induction of CYP3A4 was observed in HPHs cotreated with BMS-665351 and prototypical activators of CAR but not PXR. In summary, our results indicate that BMS-665351-mediated induction of CYP3A4 is CAR-dependent, but BMS-665351 itself is not a typical activator of either CAR or PXR, rather it functions as a selective inducer of CAR expression and

  13. A constitutive model for the forces of a magnetic bearing including eddy currents

    NASA Technical Reports Server (NTRS)

    Taylor, D. L.; Hebbale, K. V.

    1993-01-01

    A multiple magnet bearing can be developed from N individual electromagnets. The constitutive relationships for a single magnet in such a bearing is presented. Analytical expressions are developed for a magnet with poles arranged circumferencially. Maxwell's field equations are used so the model easily includes the effects of induced eddy currents due to the rotation of the journal. Eddy currents must be included in any dynamic model because they are the only speed dependent parameter and may lead to a critical speed for the bearing. The model is applicable to bearings using attraction or repulsion.

  14. Identification of constitutional MLH1 epimutations and promoter variants in colorectal cancer patients from the Colon Cancer Family Registry

    PubMed Central

    Ward, Robyn L.; Dobbins, Timothy; Lindor, Noralane M.; Rapkins, Robert W.; Hitchins, Megan P.

    2013-01-01

    Purpose: Constitutional MLH1 epimutations manifest as promoter methylation and silencing of the affected allele in normal tissues, predisposing to Lynch syndrome–associated cancers. This study investigated their frequency and inheritance. Methods: A total of 416 individuals with a colorectal cancer showing loss of MLH1 expression and without deleterious germline mutations in MLH1 were ascertained from the Colon Cancer Family Registry (C-CFR). Constitutive DNA samples were screened for MLH1 methylation in all 416 subjects and for promoter sequence changes in 357 individuals. Results: Constitutional MLH1 epimutations were identified in 16 subjects. Of these, seven (1.7%) had mono- or hemi-allelic methylation and eight had low-level methylation (2%). In one subject the epimutation was linked to the c.-27C>A promoter variant. Testing of 37 relatives from nine probands revealed paternal transmission of low-level methylation segregating with a c.+27G>A variant in one case. Five additional probands had a promoter variant without an MLH1 epimutation, with three showing diminished promoter activity in functional assays. Conclusion: Although rare, sequence changes in the regulatory region of MLH1 and aberrant methylation may alone or together predispose to the development of cancer. Screening for these changes is warranted in individuals who have a negative germline sequence screen of MLH1 and loss of MLH1 expression in their tumor. PMID:22878509

  15. Automatic Control of Gene Expression in Mammalian Cells.

    PubMed

    Fracassi, Chiara; Postiglione, Lorena; Fiore, Gianfranco; di Bernardo, Diego

    2016-04-15

    Automatic control of gene expression in living cells is paramount importance to characterize both endogenous gene regulatory networks and synthetic circuits. In addition, such a technology can be used to maintain the expression of synthetic circuit components in an optimal range in order to ensure reliable performance. Here we present a microfluidics-based method to automatically control gene expression from the tetracycline-inducible promoter in mammalian cells in real time. Our approach is based on the negative-feedback control engineering paradigm. We validated our method in a monoclonal population of cells constitutively expressing a fluorescent reporter protein (d2EYFP) downstream of a minimal CMV promoter with seven tet-responsive operator motifs (CMV-TET). These cells also constitutively express the tetracycline transactivator protein (tTA). In cells grown in standard growth medium, tTA is able to bind the CMV-TET promoter, causing d2EYFP to be maximally expressed. Upon addition of tetracycline to the culture medium, tTA detaches from the CMV-TET promoter, thus preventing d2EYFP expression. We tested two different model-independent control algorithms (relay and proportional-integral (PI)) to force a monoclonal population of cells to express an intermediate level of d2EYFP equal to 50% of its maximum expression level for up to 3500 min. The control input is either tetracycline-rich or standard growth medium. We demonstrated that both the relay and PI controllers can regulate gene expression at the desired level, despite oscillations (dampened in the case of the PI controller) around the chosen set point.

  16. Immune challenge differentially affects transcript abundance of three antimicrobial peptides in hemocytes from the moth Pseudoplusia includens.

    PubMed

    Lavine, M D; Chen, G; Strand, M R

    2005-12-01

    Inducible expression of antimicrobial peptides and other humoral immune factors by the insect fat body is well documented. Hemocytes comprise the second essential arm of the insect immune system but it is unclear whether antimicrobial peptide genes are expressed by all or only some types of hemocytes. Here we report the cloning of cecropin A (Pi-cecA), lebocin (Pi-leb) and lysozyme (Pi-lys) homologs from the moth Pseudoplusia includens. Relative-quantitative real-time PCR (rq-rtPCR) indicated that transcript abundance for each antimicrobial gene increased in fat body and hemocytes following immune challenge with the Gram-negative bacterium Escherichia coli. Relative transcript abundance of Pi-cecA was much higher in fat body than hemocytes. In contrast, transcript levels of Pi-leb were three-fold lower in hemocytes than fat body while transcript levels of Pi-lys were three-fold higher. Estimates for the overall contribution of the fat body and hemocytes to antimicrobial peptide expression suggested that hemocytes contribute significantly to Pi-lys transcript levels in larvae but produce much smaller amounts of Pi-cecA and Pi-leb compared to the fat body. Each antimicrobial peptide was also inducibly expressed in hemocytes following challenge with the Gram-positive bacterium Micrococcus luteus or when hemocytes formed capsules around chromatography beads. Analysis of hemocyte types indicated that granulocytes and plasmatocytes expressed all three antimicrobial peptides, whereas spherule cells and oenocytoids expressed only lysozyme. Transcriptional profiles of these antimicrobial genes were similar in granulocytes and plasmatocytes in vivo but were very different in vitro.

  17. Constitutive Behavior and Processing Map of T2 Pure Copper Deformed from 293 to 1073 K

    NASA Astrophysics Data System (ADS)

    Liu, Ying; Xiong, Wei; Yang, Qing; Zeng, Ji-Wei; Zhu, Wen; Sunkulp, Goel

    2018-02-01

    The deformation behavior of T2 pure copper compressed from 293 to 1073 K with strain rates from 0.01 to 10 s-1 was investigated. The constitutive equations were established by the Arrhenius constitutive model, which can be expressed as a piecewise function of temperature with two sections, in the ranges 293-723 K and 723-1073 K. The processing maps were established according to the dynamic material model for strains of 0.2, 0.4, 0.6, and 0.8, and the optimal processing parameters of T2 copper were determined accordingly. In order to obtain a better understanding of the deformation behavior, the microstructures of the compressed samples were studied by electron back-scattered diffraction. The grains tend to be more refined with decreases in temperature and increases in strain rate.

  18. The Abundance and Activity of Nitrate-Reducing Microbial Populations in Estuarine Sediments

    NASA Astrophysics Data System (ADS)

    Cardarelli, E.; Francis, C. A.

    2014-12-01

    Estuaries are productive ecosystems that ameliorate nutrient and metal contaminants from surficial water supplies. At the intersection of terrestrial and aquatic environments, estuarine sediments host major microbially-mediated geochemical transformations. These include denitrification (the conversion of nitrate to nitrous oxide and/or dinitrogen) and dissimilatory nitrate reduction to ammonium (DNRA). Denitrification has historically been seen as the predominant nitrate attenuation process and functions as an effective sink for nitrate. DNRA has previously been believed to be a minor nitrate reduction process and transforms nitrate within the ecosystem to ammonium, a more biologically available N species. Recent studies have compared the two processes in coastal environments and determined fluctuating environmental conditions may suppress denitrification, supporting an increased role for DNRA in the N cycle. Nitrate availability and salinity are factors thought to influence the membership of the microbial communities present, and the nitrate reduction process that predominates. The aim of this study is to investigate how nitrate concentration and salinity alter the transcript abundances of N cycling functional gene markers for denitrification (nirK, nirS) and DNRA (nrfA) in estuarine sediments at the mouth of the hypernutrified Old Salinas River, CA. Short-term whole core incubations amended with artificial freshwater/artificial seawater (2 psu, 35 psu) and with varying NO3- concentrations (200mM, 2000mM) were conducted to assess the activity as well as the abundance of the nitrate-reducing microbial populations present. Gene expression of nirK, nirS, and nrfA at the conclusion of the incubations was quantified using reverse transcription quantitative polymerase chain reaction (RT-qPCR). High abundances of nirK, nirS, and nrfA under particular conditions coupled with the resulting geochemical data ultimately provides insight onto how the aforementioned factors

  19. Bringing abundance into environmental politics: Constructing a Zionist network of water abundance, immigration, and colonization.

    PubMed

    Alatout, Samer

    2009-06-01

    For more than five decades, resource scarcity has been the lead story in debates over environmental politics. More importantly, and whenever environmental politics implies conflict, resource scarcity is constructed as the culprit. Abundance of resources, if at all visited in the literature, holds less importance. Resource abundance is seen, at best, as the other side of scarcity--maybe the successful conclusion of multiple interventions that may turn scarcity into abundance. This paper reinstates abundance as a politico-environmental category in its own right. Rather than relegating abundance to a second-order environmental actor that matters only on occasion, this paper foregrounds it as a crucial element in modern environmental politics. On the substantive level, and using insights from science and technology studies, especially a slightly modified actor-network framework, I describe the emergence and consolidation of a Zionist network of abundance, immigration, and colonization in Palestine between 1918 and 1948. The essential argument here is that water abundance was constructed as fact, and became a political rallying point around which a techno-political network emerged that included a great number of elements. To name just a few, the following were enrolled in the service of such a network: geologists, geophysicists, Zionist settlement experts, Zionist organizations, political and technical categories of all sorts, Palestinians as the negated others, Palestinian revolts in search of political rights, the British Mandate authorities, the hydrological system of Palestine, and the absorptive capacity of Palestine, among others. The point was to successfully articulate these disparate elements into a network that seeks opening Palestine for Jewish immigration, redefining Palestinian geography and history through Judeo-Christian Biblical narratives, and, in the process, de-legitimizing political Palestinian presence in historic Palestine.

  20. Multiple marker abundance profiling: combining selected reaction monitoring and data-dependent acquisition for rapid estimation of organelle abundance in subcellular samples

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hooper, Cornelia M.; Stevens, Tim J.; Saukkonen, Anna

    Measuring changes in protein or organelle abundance in the cell is an essential, but challenging aspect of cell biology. Frequently-used methods for determining organelle abundance typically rely on detection of a very few marker proteins, so are unsatisfactory. In silico estimates of protein abundances from publicly available protein spectra can provide useful standard abundance values but contain only data from tissue proteomes, and are not coupled to organelle localization data. A new protein abundance score, the normalized protein abundance scale (NPAS), expands on the number of scored proteins and the scoring accuracy of lower-abundance proteins in Arabidopsis. NPAS was combinedmore » with subcellular protein localization data, facilitating quantitative estimations of organelle abundance during routine experimental procedures. A suite of targeted proteomics markers for subcellular compartment markers was developed, enabling independent verification of in silico estimates for relative organelle abundance. Estimation of relative organelle abundance was found to be reproducible and consistent over a range of tissues and growth conditions. In silico abundance estimations and localization data have been combined into an online tool, multiple marker abundance profiling, available in the SUBA4 toolbox (http://suba.live).« less

  1. Multiple marker abundance profiling: combining selected reaction monitoring and data-dependent acquisition for rapid estimation of organelle abundance in subcellular samples

    DOE PAGES

    Hooper, Cornelia M.; Stevens, Tim J.; Saukkonen, Anna; ...

    2017-10-12

    Measuring changes in protein or organelle abundance in the cell is an essential, but challenging aspect of cell biology. Frequently-used methods for determining organelle abundance typically rely on detection of a very few marker proteins, so are unsatisfactory. In silico estimates of protein abundances from publicly available protein spectra can provide useful standard abundance values but contain only data from tissue proteomes, and are not coupled to organelle localization data. A new protein abundance score, the normalized protein abundance scale (NPAS), expands on the number of scored proteins and the scoring accuracy of lower-abundance proteins in Arabidopsis. NPAS was combinedmore » with subcellular protein localization data, facilitating quantitative estimations of organelle abundance during routine experimental procedures. A suite of targeted proteomics markers for subcellular compartment markers was developed, enabling independent verification of in silico estimates for relative organelle abundance. Estimation of relative organelle abundance was found to be reproducible and consistent over a range of tissues and growth conditions. In silico abundance estimations and localization data have been combined into an online tool, multiple marker abundance profiling, available in the SUBA4 toolbox (http://suba.live).« less

  2. FABP4-Cre mediated expression of constitutively active ChREBP protects against obesity

    USDA-ARS?s Scientific Manuscript database

    Carbohydrate response element binding protein (ChREBP) regulates cellular glucose and lipid homeostasis. Although ChREBP is highly expressed in many key metabolic tissues, the role of ChREBP in most of those tissues and consequent effects on whole-body glucose and lipid metabolism are not well under...

  3. The Constitution at 200: Celebration Amidst Controversy.

    ERIC Educational Resources Information Center

    Collins, Sheila D.

    1987-01-01

    Current debates about the Constitution fall into the three following categories: (1) reappraisals of consitutional origins; (2) disagreements on hermeneutical principles used in contemporary applications; and (3) discussions of contemporary events whose consequences for law and political stability could not have been foreseen by the Constitution's…

  4. Public and Constitutional Support for Character Education.

    ERIC Educational Resources Information Center

    Vessels, Gordon G.; Boyd, Stephen M.

    1996-01-01

    Character education thrives on an informed understanding of constitutional principles and an inclusive commitment-building process. U.S. Supreme Court opinions that clarify public school students' free speech rights have established values education as a constitutionally acceptable practice. Challenges might lie in possible violations of the First…

  5. A microRNA feedback loop regulates global microRNA abundance during aging.

    PubMed

    Inukai, Sachi; Pincus, Zachary; de Lencastre, Alexandre; Slack, Frank J

    2018-02-01

    Expression levels of many microRNAs (miRNAs) change during aging, notably declining globally in a number of organisms and tissues across taxa. However, little is known about the mechanisms or the biological relevance for this change. We investigated the network of genes that controls miRNA transcription and processing during C. elegans aging. We found that miRNA biogenesis genes are highly networked with transcription factors and aging-associated miRNAs. In particular, miR-71, known to influence life span and itself up-regulated during aging, represses alg-1 /Argonaute expression post-transcriptionally during aging. Increased ALG-1 abundance in mir-71 loss-of-function mutants led to globally increased miRNA expression. Interestingly, these mutants demonstrated widespread mRNA expression dysregulation and diminished levels of variability both in gene expression and in overall life span. Thus, the progressive molecular decline often thought to be the result of accumulated damage over an organism's life may be partially explained by a miRNA-directed mechanism of age-associated decline. © 2018 Inukai et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  6. Multiple factors affect diversity and abundance of ammonia-oxidizing microorganisms in iron mine soil.

    PubMed

    Xing, Yi; Si, Yan-Xiao; Hong, Chen; Li, Yang

    2015-07-01

    Ammonia oxidation by microorganisms is a critical process in the nitrogen cycle. In this study, four soil samples collected from a desert zone in an iron-exploration area and others from farmland and planted forest soil in an iron mine surrounding area. We analyzed the abundance and diversity of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in iron-mining area near the Miyun reservoir using ammonia monooxygenase. A subunit gene (amoA) as molecular biomarker. Quantitative polymerase chain reaction was applied to explore the relationships between the abundance of AOA and AOB and soil physicochemical parameters. The results showed that AOA were more abundant than AOB and may play a more dominant role in the ammonia-oxidizing process in the whole region. PCR-denaturing gradient gel electrophoresis was used to analyze the structural changes of AOA and AOB. The results showed that AOB were much more diverse than AOA. Nitrosospira cluster three constitute the majority of AOB, and AOA were dominated by group 1.1b in the soil. Redundancy analysis was performed to explore the physicochemical parameters potentially important to AOA and AOB. Soil characteristics (i.e. water, ammonia, organic carbon, total nitrogen, available phosphorus, and soil type) were proposed to potentially contribute to the distributions of AOB, whereas Cd was also closely correlated to the distributions of AOB. The community of AOA correlated with ammonium and water contents. These results highlight the importance of multiple drivers in microbial niche formation as well as their affect on ammonia oxidizer composition, both which have significant consequences for ecosystem nitrogen functioning.

  7. 78 FR 39721 - Constitution Pipeline Company, LLC; Notice of Application

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-02

    ...] Constitution Pipeline Company, LLC; Notice of Application Take notice that on June 13, 2013, Constitution Pipeline Company, LLC (Constitution), having its principal place of business at 2800 Post Oak Boulevard..., Chenango, Delaware, and Schoharie Counties, New York.\\1\\ Constitution further requests that the Commission...

  8. Monitoring Butterfly Abundance: Beyond Pollard Walks

    PubMed Central

    Pellet, Jérôme; Bried, Jason T.; Parietti, David; Gander, Antoine; Heer, Patrick O.; Cherix, Daniel; Arlettaz, Raphaël

    2012-01-01

    Most butterfly monitoring protocols rely on counts along transects (Pollard walks) to generate species abundance indices and track population trends. It is still too often ignored that a population count results from two processes: the biological process (true abundance) and the statistical process (our ability to properly quantify abundance). Because individual detectability tends to vary in space (e.g., among sites) and time (e.g., among years), it remains unclear whether index counts truly reflect population sizes and trends. This study compares capture-mark-recapture (absolute abundance) and count-index (relative abundance) monitoring methods in three species (Maculinea nausithous and Iolana iolas: Lycaenidae; Minois dryas: Satyridae) in contrasted habitat types. We demonstrate that intraspecific variability in individual detectability under standard monitoring conditions is probably the rule rather than the exception, which questions the reliability of count-based indices to estimate and compare specific population abundance. Our results suggest that the accuracy of count-based methods depends heavily on the ecology and behavior of the target species, as well as on the type of habitat in which surveys take place. Monitoring programs designed to assess the abundance and trends in butterfly populations should incorporate a measure of detectability. We discuss the relative advantages and inconveniences of current monitoring methods and analytical approaches with respect to the characteristics of the species under scrutiny and resources availability. PMID:22859980

  9. Variable directionality of gene expression changes across generations does not constitute negative evidence of epigenetic inheritance.

    PubMed

    Sharma, Abhay

    2015-01-01

    Transgenerational epigenetic inheritance in mammals has been controversial due to inherent difficulties in its experimental demonstration. A recent report has, however, opened a new front in the ongoing debate by claiming that endocrine disrupting chemicals, contrary to previous findings, do not cause effects across generations. This claim is based on the observation that gene expression changes induced by these chemicals in the exposed and unexposed generations are mainly in the opposite direction. This analysis shows that the pattern of gene expression reported in the two generations is not expected by chance and is suggestive of transmission across generations. A meta-analysis of diverse data sets related to endocrine disruptor-induced transgenerational gene expression alterations, including the data provided in the said report, further suggests that effects of endocrine disrupting chemicals persist in unexposed generations. Based on the prior evidence of phenotypic variability and gene expression alterations in opposite direction between generations, it is argued here that calling evidence of mismatched directionality in gene expression in experiments testing potential of environmental agents in inducing epigenetic inheritance of phenotypic traits as negative is untenable. This is expected to settle the newly raised doubts over epigenetic inheritance in mammals.

  10. The boron abundance of Procyon

    NASA Technical Reports Server (NTRS)

    Lemke, Michael; Lambert, David L.; Edvardsson, Bengt

    1993-01-01

    The B I 2496.8 A resonance line and HST/GHRS echelle spectra are used with model atmospheres and synthetic spectra to derive the B abundance of the F dwarfs Procyon (Alpha Canis Minoris), Theta Ursae Majoris, and Iota Pegasi. The B abundance of Theta UMa and Iota Peg is similar to that derived by Boesgaard and Heacox (1978) from the B II resonance line in spectra of A- and B-type stars. These two dwarfs show normal abundances of Li, Be, and B. Procyon, which is highly depleted in Li and Be, is depleted in B by a factor of at least 3. Comparison of the spectra of Procyon and the halo dwarf HD 140283 shows that the B abundance assigned by Duncan et al. (1992) to three halo dwarfs is not greatly overestimated as a result of contamination of the B I line by an unidentified line.

  11. Subcellular Localization of Large Yellow Croaker ( Larimichthys crocea) TLR21 and Expression Profiling of Its Gene in Immune Response

    NASA Astrophysics Data System (ADS)

    Sun, Qingxue; Fan, Zejun; Yao, Cuiluan

    2018-04-01

    Toll-like receptor 21 (TLR21) is a non-mammalian type TLR, and plays an important role in innate immune response in fish. In this paper, the full-length cDNA sequence of TLR21 gene was identified and characterized from large yellow croaker, Larimichthys crocea and was termed as LcTLR21. It consists of 3365 bp, including a 5'-terminal untranslated region (UTR) of 97 bp, a 3'-terminal UTR of 331 bp, and an open reading frame (ORF) of 2937 bp encoding a polypeptide of 978 amino acid residues. The deduced LcTLR21 contains a signal peptide domain at N-terminal, 12 leucine-rich repeats (LRRs) at the extracellular region, a transmembrane domain and a cytoplasmic toll-interleukin-1 receptor (TIR) domain at the C-terminal. Subcellular localization analysis revealed that the LcTLR21-GFP was constitutively expressed in cytoplasm. Tissue expression analysis indicated that LcTLR21 gene broadly expressed in most of the examined tissues, with the most predominant abundance in spleen, followed by head-kidney and liver, while the weakest expression was detected in brain. The expression level of LcTLR21 after LPS, poly I:C and Vibrio parahaemolyticus challenges was investigated in spleen, head-kidney and liver. LcTLR21 gene transcripts increased significantly in all examined tissues after the challenges, and the highest expression level was detected in liver at 24 h after poly I:C stimulation ( P < 0.05), suggesting that LcTLR21 might play a crucial role in fish resistance to viral and bacterial infections.

  12. [Women, gender, and the Constitution].

    PubMed

    1993-12-01

    Although all the constitutions of Latin America directly or indirectly acknowledge the juridical equality of the sexes, these patriarchal societies continue to maintain institutional power in male hands and to neutralize legal actions favoring women. International instruments such as the Convention on Elimination of All Forms of Discrimination Against Women, approved by the UN in 1979, have given a firmer basis to policies and actions to improve the status of women. Obstacles to full equality of Latin American women are rooted in economic and sociopolitical factors, but lack of true political will also plays a significant role. A number of new laws in the past several years as well as the new Constitution have improved the legal position of Colombian women. The new Constitution recognizes fundamental rights that may be claimed directly before a judge, and social, economic, and collective rights requiring legislative development. Article 43 of the new Constitution states that women will not be subjected to any form of discrimination. Another norm states that women will enjoy special assistance and protection before and after childbirth, in recognition of the social functions of maternity. Article 43 also states that women who are heads of households will receive special assistance, but the corresponding regulations have not yet been promulgated. The mechanism of tutelage has become an important recourse that has been used in several cases in which fundamental rights of women have been violated or threatened because of their sex. The order of tutelage has been used in cases of adolescents expelled from school for pregnancy and of abused wives, as well as to force recognition of the social and economic contributions of housework.

  13. Bioluminescence Resonance Energy Transfer Studies Reveal Constitutive Dimerization of the Human Lutropin Receptor and a Lack of Correlation between Receptor Activation and the Propensity for Dimerization*

    PubMed Central

    Guan, Rongbin; Feng, Xiuyan; Wu, Xueqing; Zhang, Meilin; Zhang, Xuesen; Hébert, Terence E.; Segaloff, Deborah L.

    2009-01-01

    Previous studies from our laboratory using co-immunoprecipitation techniques suggested that the human lutropin receptor (hLHR) constitutively self-associates into dimers/oligomers and that agonist treatment of cells either increased hLHR dimerization/oligomerization and/or stabilized hLHR dimers/oligomers to detergent solubilization (Tao, Y. X., Johnson, N. B., and Segaloff, D. L. (2004) J. Biol. Chem. 279, 5904–5914). In this study, bioluminescence resonance energy transfer (BRET2) analyses confirmed that the hLHR constitutively self-associates in living cells. After subcellular fractionation, hLHR dimers/oligomers were detected in both the plasma membrane and endoplasmic reticulum (ER). Further evidence supporting the constitutive formation of hLHR dimer/oligomers in the ER is provided by data showing homodimerization of misfolded hLHR mutants that are retained in the ER. These mutants, when co-expressed with wild-type receptor, are shown by BRET2 to heterodimerize, accounting for their dominant-negative effects on cell surface receptor expression. Hormone desorption assays using intact cells demonstrate allosterism between hLHR protomers, indicating functional cell surface hLHR dimers. However, quantitative BRET2 analyses in intact cells indicate a lack of effect of agonist on the propensity of the hLHR to dimerize. Using purified plasma membranes, human chorionic gonadotropin was similarly observed to have no effect on the BRET2 signal. An examination of the propensity for constitutively active and signaling inactive hLHR mutants to dimerize further showed no correlation between dimerization and the activation state of the hLHR. Taken altogether, our data suggest that hLHR dimers/oligomers are formed early in the biosynthetic pathway in the ER, are constitutively expressed on the plasma membrane, and are not affected by the activation state of the hLHR. PMID:19147490

  14. Clinical study on constitutional herbal tea for treating chronic fatigue.

    PubMed

    Park, Soo-Jung; Bae, Young-Chun; Choi, Na-Rae; Ryu, Seung-Yeob; Kwon, Young-Mi; Joo, Jong-Cheon

    2014-12-01

    This study was designed to evaluate the efficacy and the safety of constitutional herbal tea for treating chronic fatigue with no diagnosed cause, which is called Mibyeong in Korea. Males and females with ages between 40 and 59 years who had complained of fatigue for 1 month consistently or for 6 months intermittently without a definite cause were recruited. At the same time, a Chalder fatigue scale (CFS) score of 19 was essential for participation in this study. Sixty five subjects completed the entire process, including blood tests and tests with medical devices. Five assessments of health status were accomplished over 8 weeks by using the CFS and the visual analogue scale (VAS). To ensure that the constitutional herbal tea was being safely used, we conducted and analyzed renal function and liver function tests. For the diagnosis of the Sasang constitution, the Sasang Constitutional Analysis Tool (SCAT) was used, and a specialist in Sasang constitutional medicine made the final diagnosis based on the SCAT result. Constitutional herbal tea was served four weeks after the first visit. The subjects took the constitutional herbal tea twice a day for one month. The results are as follows: The CFS and the VAS scores were significantly improved for the subjects in the constitutional herbal tea. No abnormalities were found on the blood tests to evaluate safety after taking the constitutional herbal tea. The improvements in the CFS and the VAS scores due to the constitutional herbal tea had no significant differences according to the Sasang constitution. Constitutional herbal tea may be used to reduce fatigue and improve health and has no adverse effect on either the kidney or the liver.

  15. An Internet Guide to Teaching the Constitution

    ERIC Educational Resources Information Center

    Risinger, C. Frederick

    2005-01-01

    In 2004, Congress passed and President Bush signed legislation designating every September 17 as Constitution Day and suggesting that all schools teach about the Constitution on that day. There are innumerable websites, countless lesson plans and a multitude of other teaching resources on the topic. Teachers were inundated with brochures,…

  16. 27 CFR 71.128 - What constitutes record.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 2 2014-04-01 2014-04-01 false What constitutes record. 71.128 Section 71.128 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU... Miscellaneous Record § 71.128 What constitutes record. The transcript of testimony, pleadings and exhibits, all...

  17. 27 CFR 71.128 - What constitutes record.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 2 2010-04-01 2010-04-01 false What constitutes record. 71.128 Section 71.128 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU... Miscellaneous Record § 71.128 What constitutes record. The transcript of testimony, pleadings and exhibits, all...

  18. 29 CFR 452.3 - Interpretations of constitution and bylaws.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 29 Labor 2 2014-07-01 2014-07-01 false Interpretations of constitution and bylaws. 452.3 Section... REPORTING AND DISCLOSURE ACT OF 1959 General Considerations § 452.3 Interpretations of constitution and bylaws. The interpretation consistently placed on a union's constitution by the responsible union...

  19. 29 CFR 452.3 - Interpretations of constitution and bylaws.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 29 Labor 2 2010-07-01 2010-07-01 false Interpretations of constitution and bylaws. 452.3 Section... REPORTING AND DISCLOSURE ACT OF 1959 General Considerations § 452.3 Interpretations of constitution and bylaws. The interpretation consistently placed on a union's constitution by the responsible union...

  20. 29 CFR 452.3 - Interpretations of constitution and bylaws.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 29 Labor 2 2013-07-01 2013-07-01 false Interpretations of constitution and bylaws. 452.3 Section... REPORTING AND DISCLOSURE ACT OF 1959 General Considerations § 452.3 Interpretations of constitution and bylaws. The interpretation consistently placed on a union's constitution by the responsible union...

  1. 29 CFR 452.3 - Interpretations of constitution and bylaws.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 29 Labor 2 2011-07-01 2011-07-01 false Interpretations of constitution and bylaws. 452.3 Section... REPORTING AND DISCLOSURE ACT OF 1959 General Considerations § 452.3 Interpretations of constitution and bylaws. The interpretation consistently placed on a union's constitution by the responsible union...

  2. 29 CFR 452.3 - Interpretations of constitution and bylaws.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 29 Labor 2 2012-07-01 2012-07-01 false Interpretations of constitution and bylaws. 452.3 Section... REPORTING AND DISCLOSURE ACT OF 1959 General Considerations § 452.3 Interpretations of constitution and bylaws. The interpretation consistently placed on a union's constitution by the responsible union...

  3. Predicting Sasang Constitution Using Body-Shape Information

    PubMed Central

    Jang, Eunsu; Do, Jun-Hyeong; Jin, HeeJeong; Park, KiHyun; Ku, Boncho; Lee, Siwoo; Kim, Jong Yeol

    2012-01-01

    Objectives. Body measurement plays a pivotal role not only in the diagnosis of disease but also in the classification of typology. Sasang constitutional medicine, which is one of the forms of Traditional Korean Medicine, is considered to be strongly associated with body shape. We attempted to determine whether a Sasang constitutional analytic tool based on body shape information (SCAT-B) could predict Sasang constitution (SC). Methods. After surveying 23 Oriental medical clinics, 2,677 subjects were recruited and body shape information was collected. The SCAT-Bs for males and females were developed using multinomial logistic regression. Stepwise forward-variable selection was applied using the score statistic and Wald's test. Results. The predictive rates of the SCAT-B for Tae-eumin (TE), Soeumin (SE), and Soyangin (SY) types in males and females were 80.2%, 56.9%, and 37.7% (males) and 69.3%, 38.9%, and 50.0% (females) in the training set and were 74%, 70.1%, and 35% (males), and 67.4%, 66.3%, and 53.7% (females) in the test set, respectively. Higher constitutional probability scores showed a trend for association with higher predictability. Conclusions. This study shows that the Sasang constitutional analytic tool, which is based on body shape information, may be relatively highly predictive of TE type but may be less predictive when used for SY type. PMID:22792124

  4. Constitutive apical membrane recycling in Aplysia enterocytes.

    PubMed

    Keeton, Robert Aaron; Runge, Steven William; Moran, William Michael

    2004-11-01

    In Aplysia californica enterocytes, alanine-stimulated Na+ absorption increases both apical membrane exocytosis and fractional capacitance (fCa; a measure of relative apical membrane surface area). These increases are thought to reduce membrane tension during periods of nutrient absorption that cause the enterocytes to swell osmotically. In the absence of alanine, exocytosis and fCa are constant. These findings imply equal rates of constitutive endocytosis and exocytosis and constitutive recycling of the apical plasma membrane. Thus, the purpose of this study was to confirm and determine the relative extent of constitutive apical membrane recycling in Aplysia enterocytes. Biotinylated lectins are commonly used to label plasma membranes and to investigate plasma membrane recycling. Of fourteen biotinylated lectins tested, biotinylated wheat germ agglutinin (bWGA) bound preferentially to the enterocytes apical surface. Therefore, we used bWGA, avidin D (which binds tightly to biotin), and the UV fluorophore 7-amino-4-methylcoumarin-3-acetic acid (AMCA)-conjugated avidin D to assess the extent of constitutive apical membrane recycling. A temperature-dependent (20 vs. 4 degrees C) experimental protocol employed the use of two tissues from each of five snails and resulted in a approximately 60% difference in apical surface fluorescence intensity. Because the extent of membrane recycling is proportional to the difference in surface fluorescence intensity, this difference reveals a relatively high rate of constitutive apical membrane recycling in Aplysia enterocytes.

  5. Nonlinear Constitutive Modeling of Piezoelectric Ceramics

    NASA Astrophysics Data System (ADS)

    Xu, Jia; Li, Chao; Wang, Haibo; Zhu, Zhiwen

    2017-12-01

    Nonlinear constitutive modeling of piezoelectric ceramics is discussed in this paper. Van der Pol item is introduced to explain the simple hysteretic curve. Improved nonlinear difference items are used to interpret the hysteresis phenomena of piezoelectric ceramics. The fitting effect of the model on experimental data is proved by the partial least-square regression method. The results show that this method can describe the real curve well. The results of this paper are helpful to piezoelectric ceramics constitutive modeling.

  6. CHEMICAL ABUNDANCES IN THE EXTERNALLY POLLUTED WHITE DWARF GD 40: EVIDENCE OF A ROCKY EXTRASOLAR MINOR PLANET

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Klein, B.; Jura, M.; Zuckerman, B.

    2010-02-01

    We present Keck/High Resolution Echelle Spectrometer data with model atmosphere analysis of the helium-dominated polluted white dwarf GD 40, in which we measure atmospheric abundances relative to helium of nine elements: H, O, Mg, Si, Ca, Ti, Cr, Mn, and Fe. Apart from hydrogen, whose association with the other contaminants is uncertain, this material most likely accreted from GD 40's circumstellar dust disk whose existence is demonstrated by excess infrared emission. The data are best explained by accretion of rocky planetary material, in which heavy elements are largely contained within oxides, derived from a tidally disrupted minor planet at leastmore » the mass of Juno, and probably as massive as Vesta. The relatively low hydrogen abundance sets an upper limit of 10% water by mass in the inferred parent body, and the relatively high abundances of refractory elements, Ca and Ti, may indicate high-temperature processing. While the overall constitution of the parent body is similar to the bulk Earth being over 85% by mass composed of oxygen, magnesium, silicon, and iron, we find n(Si)/n(Mg) = 0.30 +- 0.11, significantly smaller than the ratio near unity for the bulk Earth, chondrites, the Sun, and nearby stars. This result suggests that differentiation occurred within the parent body.« less

  7. A monoallelic-to-biallelic T-cell transcriptional switch regulates GATA3 abundance

    PubMed Central

    Ku, Chia-Jui; Lim, Kim-Chew; Kalantry, Sundeep; Maillard, Ivan; Engel, James Douglas; Hosoya, Tomonori

    2015-01-01

    Protein abundance must be precisely regulated throughout life, and nowhere is the stringency of this requirement more evident than during T-cell development: A twofold increase in the abundance of transcription factor GATA3 results in thymic lymphoma, while reduced GATA3 leads to diminished T-cell production. GATA3 haploinsufficiency also causes human HDR (hypoparathyroidism, deafness, and renal dysplasia) syndrome, often accompanied by immunodeficiency. Here we show that loss of one Gata3 allele leads to diminished expansion (and compromised development) of immature T cells as well as aberrant induction of myeloid transcription factor PU.1. This effect is at least in part mediated transcriptionally: We discovered that Gata3 is monoallelically expressed in a parent of origin-independent manner in hematopoietic stem cells and early T-cell progenitors. Curiously, half of the developing cells switch to biallelic Gata3 transcription abruptly at midthymopoiesis. We show that the monoallelic-to-biallelic transcriptional switch is stably maintained and therefore is not a stochastic phenomenon. This unique mechanism, if adopted by other regulatory genes, may provide new biological insights into the rather prevalent phenomenon of monoallelic expression of autosomal genes as well as into the variably penetrant pathophysiological spectrum of phenotypes observed in many human syndromes that are due to haploinsufficiency of the affected gene. PMID:26385963

  8. ABUNDANT OR RARE? A HYBRID APPROACH FOR DETERMINING SPECIES RELATIVE ABUNDANCE AT AN ECOREGOIONAL SCALE

    EPA Science Inventory

    Everyone knows what abundant and rare species are, but quantifying the concept proves elusive. As part of an EPA/USGS project to assess near-coastal species vulnerability to climate change affects, we designed a hybrid approach to determine species relative abundance at an ecoreg...

  9. A search for stars of very low metal abundance. VI. Detailed abundances of 313 metal-poor stars

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Roederer, Ian U.; Preston, George W.; Thompson, Ian B.

    2014-06-01

    We present radial velocities, equivalent widths, model atmosphere parameters, and abundances or upper limits for 53 species of 48 elements derived from high resolution optical spectroscopy of 313 metal-poor stars. A majority of these stars were selected from the metal-poor candidates of the HK Survey of Beers, Preston, and Shectman. We derive detailed abundances for 61% of these stars for the first time. Spectra were obtained during a 10 yr observing campaign using the Magellan Inamori Kyocera Echelle spectrograph on the Magellan Telescopes at Las Campanas Observatory, the Robert G. Tull Coudé Spectrograph on the Harlan J. Smith Telescope atmore » McDonald Observatory, and the High Resolution Spectrograph on the Hobby-Eberly Telescope at McDonald Observatory. We perform a standard LTE abundance analysis using MARCS model atmospheres, and we apply line-by-line statistical corrections to minimize systematic abundance differences arising when different sets of lines are available for analysis. We identify several abundance correlations with effective temperature. A comparison with previous abundance analyses reveals significant differences in stellar parameters, which we investigate in detail. Our metallicities are, on average, lower by ≈0.25 dex for red giants and ≈0.04 dex for subgiants. Our sample contains 19 stars with [Fe/H] ≤–3.5, 84 stars with [Fe/H] ≤–3.0, and 210 stars with [Fe/H] ≤–2.5. Detailed abundances are presented here or elsewhere for 91% of the 209 stars with [Fe/H] ≤–2.5 as estimated from medium resolution spectroscopy by Beers, Preston, and Shectman. We will discuss the interpretation of these abundances in subsequent papers.« less

  10. Liberty, Power, and the American Constitutional Heritage.

    ERIC Educational Resources Information Center

    Madison, James H.

    The principles, values, and issues of our constitutional heritage that should be emphasized in citizenship education are based on the concepts of liberty and power. The Constitution is not a sacred icon formulated by immortals, but rather a changing and controversial framework guided by a diverse group of practical politicians, sensitive to their…

  11. Identification of genes specifically or preferentially expressed in maize silk reveals similarity and diversity in transcript abundance of different dry stigmas.

    PubMed

    Xu, Xiao Hui; Chen, Hao; Sang, Ya Lin; Wang, Fang; Ma, Jun Ping; Gao, Xin-Qi; Zhang, Xian Sheng

    2012-07-02

    In plants, pollination is a critical step in reproduction. During pollination, constant communication between male pollen and the female stigma is required for pollen adhesion, germination, and tube growth. The detailed mechanisms of stigma-mediated reproductive processes, however, remain largely unknown. Maize (Zea mays L.), one of the world's most important crops, has been extensively used as a model species to study molecular mechanisms of pollen and stigma interaction. A comprehensive analysis of maize silk transcriptome may provide valuable information for investigating stigma functionality. A comparative analysis of expression profiles between maize silk and dry stigmas of other species might reveal conserved and diverse mechanisms that underlie stigma-mediated reproductive processes in various plant species. Transcript abundance profiles of mature silk, mature pollen, mature ovary, and seedling were investigated using RNA-seq. By comparing the transcriptomes of these tissues, we identified 1,427 genes specifically or preferentially expressed in maize silk. Bioinformatic analyses of these genes revealed many genes with known functions in plant reproduction as well as novel candidate genes that encode amino acid transporters, peptide and oligopeptide transporters, and cysteine-rich receptor-like kinases. In addition, comparison of gene sets specifically or preferentially expressed in stigmas of maize, rice (Oryza sativa L.), and Arabidopsis (Arabidopsis thaliana [L.] Heynh.) identified a number of homologous genes involved either in pollen adhesion, hydration, and germination or in initial growth and penetration of pollen tubes into the stigma surface. The comparison also indicated that maize shares a more similar profile and larger number of conserved genes with rice than with Arabidopsis, and that amino acid and lipid transport-related genes are distinctively overrepresented in maize. Many of the novel genes uncovered in this study are potentially involved

  12. Identification of genes specifically or preferentially expressed in maize silk reveals similarity and diversity in transcript abundance of different dry stigmas

    PubMed Central

    2012-01-01

    Background In plants, pollination is a critical step in reproduction. During pollination, constant communication between male pollen and the female stigma is required for pollen adhesion, germination, and tube growth. The detailed mechanisms of stigma-mediated reproductive processes, however, remain largely unknown. Maize (Zea mays L.), one of the world’s most important crops, has been extensively used as a model species to study molecular mechanisms of pollen and stigma interaction. A comprehensive analysis of maize silk transcriptome may provide valuable information for investigating stigma functionality. A comparative analysis of expression profiles between maize silk and dry stigmas of other species might reveal conserved and diverse mechanisms that underlie stigma-mediated reproductive processes in various plant species. Results Transcript abundance profiles of mature silk, mature pollen, mature ovary, and seedling were investigated using RNA-seq. By comparing the transcriptomes of these tissues, we identified 1,427 genes specifically or preferentially expressed in maize silk. Bioinformatic analyses of these genes revealed many genes with known functions in plant reproduction as well as novel candidate genes that encode amino acid transporters, peptide and oligopeptide transporters, and cysteine-rich receptor-like kinases. In addition, comparison of gene sets specifically or preferentially expressed in stigmas of maize, rice (Oryza sativa L.), and Arabidopsis (Arabidopsis thaliana [L.] Heynh.) identified a number of homologous genes involved either in pollen adhesion, hydration, and germination or in initial growth and penetration of pollen tubes into the stigma surface. The comparison also indicated that maize shares a more similar profile and larger number of conserved genes with rice than with Arabidopsis, and that amino acid and lipid transport-related genes are distinctively overrepresented in maize. Conclusions Many of the novel genes uncovered in this

  13. Comparison of normalization methods for the analysis of metagenomic gene abundance data.

    PubMed

    Pereira, Mariana Buongermino; Wallroth, Mikael; Jonsson, Viktor; Kristiansson, Erik

    2018-04-20

    In shotgun metagenomics, microbial communities are studied through direct sequencing of DNA without any prior cultivation. By comparing gene abundances estimated from the generated sequencing reads, functional differences between the communities can be identified. However, gene abundance data is affected by high levels of systematic variability, which can greatly reduce the statistical power and introduce false positives. Normalization, which is the process where systematic variability is identified and removed, is therefore a vital part of the data analysis. A wide range of normalization methods for high-dimensional count data has been proposed but their performance on the analysis of shotgun metagenomic data has not been evaluated. Here, we present a systematic evaluation of nine normalization methods for gene abundance data. The methods were evaluated through resampling of three comprehensive datasets, creating a realistic setting that preserved the unique characteristics of metagenomic data. Performance was measured in terms of the methods ability to identify differentially abundant genes (DAGs), correctly calculate unbiased p-values and control the false discovery rate (FDR). Our results showed that the choice of normalization method has a large impact on the end results. When the DAGs were asymmetrically present between the experimental conditions, many normalization methods had a reduced true positive rate (TPR) and a high false positive rate (FPR). The methods trimmed mean of M-values (TMM) and relative log expression (RLE) had the overall highest performance and are therefore recommended for the analysis of gene abundance data. For larger sample sizes, CSS also showed satisfactory performance. This study emphasizes the importance of selecting a suitable normalization methods in the analysis of data from shotgun metagenomics. Our results also demonstrate that improper methods may result in unacceptably high levels of false positives, which in turn may lead

  14. Adjustable under-expression of yeast mating pathway proteins in Saccharomyces cerevisiae using a programmed ribosomal frameshift.

    PubMed

    Choi, Min-Yeon; Park, Sang-Hyun

    2016-06-01

    Experimental research in molecular biology frequently relies on the promotion or suppression of gene expression, an important tool in the study of its functions. Although yeast is among the most studied model systems with the ease of maintenance and manipulation, current experimental methods are mostly limited to gene deletion, suppression or overexpression of genes. Therefore, the ability to reduce protein expressions and then observing the effects would promote a better understanding of the exact functions and their interactions. Reducing protein expression is mainly limited by the difficulties associated with controlling the reduction level, and in some cases, the initial endogenous abundance is too low. For the under-expression to be useful as an experimental tool, repeatability and stability of reduced expression is important. We found that cis-elements in programmed -1 ribosomal frameshifting (-1RFS) of beet western yellow virus (BWYV) could be utilized to reduced protein expression in Saccharomyces cerevisiae. The two main advantages of using -1RFS are adjustable reduction rates and ease of use. To demonstrate the utility of this under-expression system, examples of reduced protein abundance were shown using yeast mating pathway components. The abundance of MAP kinase Fus3 was reduced to approximately 28-75 % of the wild-type value. Other MAP kinase mating pathway components, including Ste5, Ste11, and Ste7, were also under-expressed to verify that the -1RFS system works with different proteins. Furthermore, reduced Fus3 abundance altered the overall signal transduction outcome of the mating pathway, demonstrating the potential for further studies of signal transduction adjustment via under-expression.

  15. Lipoxin A4 Attenuates Constitutive and TGF-β1–Dependent Profibrotic Activity in Human Lung Myofibroblasts

    PubMed Central

    Feghali-Bostwick, Carol A.; Amrani, Yassine; Bradding, Peter

    2015-01-01

    Idiopathic pulmonary fibrosis (IPF) is a common, progressive, and invariably lethal interstitial lung disease with no effective therapy. The key cell driving the development of fibrosis is the myofibroblast. Lipoxin A4 (LXA4) is an anti-inflammatory lipid, important in the resolution of inflammation, and it has potential antifibrotic activity. However, the effects of LXA4 on primary human lung myofibroblasts (HLMFs) have not previously been investigated. Therefore, the aim of this study was to examine the effects of LXA4 on TGF-β1–dependent responses in IPF- and nonfibrotic control (NFC)–derived HLMFs. HLMFs were isolated from IPF and NFC patients and grown in vitro. The effects of LXA4 on HLMF proliferation, collagen secretion, α-smooth muscle actin (αSMA) expression, and Smad2/3 activation were examined constitutively and following TGF-β1 stimulation. The LXA4 receptor (ALXR) was expressed in both NFC- and IPF-derived HLMFs. LXA4 (10−10 and 10−8 mol) reduced constitutive αSMA expression, actin stress fiber formation, contraction, and nuclear Smad2/3, indicating regression from a myofibroblast to fibroblast phenotype. LXA4 also significantly inhibited FBS-dependent proliferation and TGF-β1–dependent collagen secretion, αSMA expression, and Smad2/3 nuclear translocation in IPF-derived HLMFs. LXA4 did not inhibit Smad2/3 phosphorylation. In summary, LXA4 attenuated profibrotic HLMF activity and promoted HLMF regression to a quiescent fibroblast phenotype. LXA4 or its stable analogs delivered by aerosol may offer a novel approach to the treatment of IPF. PMID:26276873

  16. Clinical Study on Constitutional Herbal Tea for Treating Chronic Fatigue

    PubMed Central

    Park, Soo-Jung; Bae, Young-Chun; Choi, Na-Rae; Ryu, Seung-Yeob; Kwon, Young-Mi; Joo, Jong-Cheon

    2014-01-01

    Objectives: This study was designed to evaluate the efficacy and the safety of constitutional herbal tea for treating chronic fatigue with no diagnosed cause, which is called Mibyeong in Korea. Methods: Males and females with ages between 40 and 59 years who had complained of fatigue for 1 month consistently or for 6 months intermittently without a definite cause were recruited. At the same time, a Chalder fatigue scale (CFS) score of 19 was essential for participation in this study. Sixty five subjects completed the entire process, including blood tests and tests with medical devices. Five assessments of health status were accomplished over 8 weeks by using the CFS and the visual analogue scale (VAS). To ensure that the constitutional herbal tea was being safely used, we conducted and analyzed renal function and liver function tests. For the diagnosis of the Sasang constitution, the Sasang Constitutional Analysis Tool (SCAT) was used, and a specialist in Sasang constitutional medicine made the final diagnosis based on the SCAT result. Constitutional herbal tea was served four weeks after the first visit. The subjects took the constitutional herbal tea twice a day for one month. Results: The results are as follows: The CFS and the VAS scores were significantly improved for the subjects in the constitutional herbal tea. No abnormalities were found on the blood tests to evaluate safety after taking the constitutional herbal tea. The improvements in the CFS and the VAS scores due to the constitutional herbal tea had no significant differences according to the Sasang constitution. Conclusion: Constitutional herbal tea may be used to reduce fatigue and improve health and has no adverse effect on either the kidney or the liver. PMID:25780720

  17. Climate and local abundance in freshwater fishes

    PubMed Central

    Knouft, Jason H.; Anthony, Melissa M.

    2016-01-01

    Identifying factors regulating variation in numbers of individuals among populations across a species' distribution is a fundamental goal in ecology. A common prediction, often referred to as the abundant-centre hypothesis, suggests that abundance is highest near the centre of a species' range. However, because of the primary focus on the geographical position of a population, this framework provides little insight into the environmental factors regulating local abundance. While range-wide variation in population abundance associated with environmental conditions has been investigated in terrestrial species, the relationship between climate and local abundance in freshwater taxa across species' distributions is not well understood. We used GIS-based temperature and precipitation data to determine the relationships between climatic conditions and range-wide variation in local abundance for 19 species of North American freshwater fishes. Climate predicted a portion of the variation in local abundance among populations for 18 species. In addition, the relationship between climatic conditions and local abundance varied among species, which is expected as lineages partition the environment across geographical space. The influence of local habitat quality on species persistence is well documented; however, our results also indicate the importance of climate in regulating population sizes across a species geographical range, even in aquatic taxa. PMID:27429769

  18. Constitutive activation of MEK1 in chondrocytes causes Stat1-independent achondroplasia-like dwarfism and rescues the Fgfr3-deficient mouse phenotype

    PubMed Central

    Murakami, Shunichi; Balmes, Gener; McKinney, Sandra; Zhang, Zhaoping; Givol, David; de Crombrugghe, Benoit

    2004-01-01

    We generated transgenic mice that express a constitutively active mutant of MEK1 in chondrocytes. These mice showed a dwarf phenotype similar to achondroplasia, the most common human dwarfism, caused by activating mutations in FGFR3. These mice displayed incomplete hypertrophy of chondrocytes in the growth plates and a general delay in endochondral ossification, whereas chondrocyte proliferation was unaffected. Immunohistochemical analysis of the cranial base in transgenic embryos showed reduced staining for collagen type X and persistent expression of Sox9 in chondrocytes. These observations indicate that the MAPK pathway inhibits hypertrophic differentiation of chondrocytes and negatively regulates bone growth without inhibiting chondrocyte proliferation. Expression of a constitutively active mutant of MEK1 in chondrocytes of Fgfr3-deficient mice inhibited skeletal overgrowth, strongly suggesting that regulation of bone growth by FGFR3 is mediated at least in part by the MAPK pathway. Although loss of Stat1 restored the reduced chondrocyte proliferation in mice expressing an achondroplasia mutant of Fgfr3, it did not rescue the reduced hypertrophic zone, the delay in formation of secondary ossification centers, and the achondroplasia-like phenotype. These observations suggest a model in which Fgfr3 signaling inhibits bone growth by inhibiting chondrocyte differentiation through the MAPK pathway and by inhibiting chondrocyte proliferation through Stat1. PMID:14871928

  19. The Environmental Estrogen, Nonylphenol, Activates the Constitutive Androstane Receptor

    PubMed Central

    Hernandez, Juan P.; Huang, Wendong; Chapman, Laura M.; Chua, Steven; Moore, David D.; Baldwin, William S.

    2007-01-01

    Nonylphenol (NP) and its parent compounds, the nonylphenol ethoxylates are some of the most prevalent chemicals found in U.S. waterways. NP is also resistant to biodegradation and is a known environmental estrogen, which makes NP a chemical of concern. Our data show that NP also activates the constitutive androstane receptor (CAR), an orphan nuclear receptor important in the induction of detoxification enzymes, including the P450s. Transactivation assays demonstrate that NP increases murine CAR (mCAR) transcriptional activity, and NP treatment can overcome the inhibitory effects of the inverse agonist, androstanol, on mCAR activation. Treatment of wild-type (CAR +/+) mice with NP at 50 or 75 mg/kg/day increases Cyp2b protein expression in a dose-dependent manner as demonstrated by Western blotting, and was confirmed by quantitative reverse transcription–PCR of Cyp2b10 transcript levels. CAR-null (CAR −/−) mice show no increased expression of Cyp2b following NP treatment, indicating that CAR is required for NP-mediated Cyp2b induction. In addition, NP increases the translocation of CAR into the nucleus, which is the key step in the commencement of CAR's transcriptional activity. NP also induced CYP2B6 in primary human hepatocytes, and increased Cyp2b10 messenger RNA and protein expression in humanized CAR mice, indicating that NP is an activator of human CAR as well. In conclusion, NP is a CAR activator, and this was demonstrated in vitro with transactivation assays and in vivo with transgenic CAR mouse models. PMID:17483497

  20. Acute and constitutive increases in central serotonin levels reduce social play behaviour in peri-adolescent rats

    PubMed Central

    Schiepers, Olga J. G.; Schoffelmeer, Anton N. M.; Cuppen, Edwin; Vanderschuren, Louk J. M. J.

    2007-01-01

    Rationale Serotonin is an important modulator of social behaviour. Individual differences in serotonergic signalling are considered to be a marker of personality that is stable throughout lifetime. While a large body of evidence indicates that central serotonin levels are inversely related to aggression and sexual behaviour in adult rats, the relationship between serotonin and social behaviour during peri-adolescence has hardly been explored. Objective To study the effect of acute and constitutive increases in serotonin neurotransmission on social behaviour in peri-adolescent rats. Materials and methods Social behaviour in peri-adolesent rats (28–35 days old) was studied after genetic ablation of the serotonin transporter, causing constitutively increased extra-neuronal serotonin levels, and after acute treatment with the serotonin reuptake inhibitor fluoxetine or the serotonin releasing agent 3,4-methylenedioxymethamphetamine (MDMA). A distinction was made between social play behaviour that mainly occurs during peri-adolescence, and non-playful social interactions that are abundant during the entire lifespan of rats. Results In serotonin transporter knockout rats, social play behaviour was markedly reduced, while non-playful aspects of social interaction were unaffected. Acute treatment with fluoxetine or MDMA dose-dependently inhibited social play behaviour. MDMA also suppressed non-playful social interaction but at higher doses than those required to reduce social play. Fluoxetine did not affect non-playful social interaction. Conclusions These data show that both acute and constitutive increases in serotonergic neurotransmission reduce social play behaviour in peri-adolescent rats. Together with our previous findings of reduced aggressive and sexual behaviour in adult serotonin transporter knockout rats, these data support the notion that serotonin modulates social behaviour in a trait-like manner. PMID:17661017