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Sample records for 4-hydroxylase gene impact

  1. Complex regulation of prolyl-4-hydroxylases impacts root hair expansion.

    PubMed

    Velasquez, Silvia M; Ricardi, Martiniano M; Poulsen, Christian Peter; Oikawa, Ai; Dilokpimol, Adiphol; Halim, Adnan; Mangano, Silvina; Denita Juarez, Silvina Paola; Marzol, Eliana; Salgado Salter, Juan D; Dorosz, Javier Gloazzo; Borassi, Cecilia; Möller, Svenning Rune; Buono, Rafael; Ohsawa, Yukiko; Matsuoka, Ken; Otegui, Marisa S; Scheller, Henrik V; Geshi, Naomi; Petersen, Bent Larsen; Iusem, Norberto D; Estevez, José M

    2015-05-01

    Root hairs are single cells that develop by tip growth, a process shared with pollen tubes, axons, and fungal hyphae. However, structural plant cell walls impose constraints to accomplish tip growth. In addition to polysaccharides, plant cell walls are composed of hydroxyproline-rich glycoproteins (HRGPs), which include several groups of O-glycoproteins, including extensins (EXTs). Proline hydroxylation, an early post-translational modification (PTM) of HRGPs catalyzed by prolyl 4-hydroxylases (P4Hs), defines their subsequent O-glycosylation sites. In this work, our genetic analyses prove that P4H5, and to a lesser extent P4H2 and P4H13, are pivotal for root hair tip growth. Second, we demonstrate that P4H5 has in vitro preferred specificity for EXT substrates rather than for other HRGPs. Third, by P4H promoter and protein swapping approaches, we show that P4H2 and P4H13 have interchangeable functions but cannot replace P4H5. These three P4Hs are shown to be targeted to the secretory pathway, where P4H5 forms dimers with P4H2 and P4H13. Finally, we explore the impact of deficient proline hydroxylation on the cell wall architecture. Taken together, our results support a model in which correct peptidyl-proline hydroxylation on EXTs, and possibly in other HRGPs, is required for proper cell wall self-assembly and hence root hair elongation in Arabidopsis thaliana.

  2. Overexpression of cinnamate 4-hydroxylase gene enhances biosynthesis of decursinol angelate in Angelica gigas hairy roots.

    PubMed

    Park, Nam Il; Park, Jee Hee; Park, Sang Un

    2012-02-01

    Angelica gigas is a medicinal plant that produces pyranocoumarins, including decursin (D) and decursinol angelate (DA), which have neuroprotective, anticancer, and antiandrogenic effects. In this study, the coumarin biosynthetic pathway was engineered to increase the production of DA. Specifically, a vector was constructed which contained the A. gigas phenylalanine ammonia-lyase (AgPAL) and cinnamate 4-hydroxylase (AgC4H) genes that were driven by the cauliflower mosaic virus (CaMV) 35S promoter. Transgenic hairy roots that overexpressed AgPAL or AgC4H genes were obtained by using an Agrobacterium rhizogenes-mediated transformation system. Among them, only AgC4H-transgenic hairy root lines produced more DA than control transgenic hairy root lines. The enhanced gene expression corresponded to elevated C4H activities. This study showed the importance of C4H in the production of DA in A. gigas hairy root culture.

  3. Cinnamate 4-Hydroxylase (C4H) genes from Leucaena leucocephala: a pulp yielding leguminous tree.

    PubMed

    Kumar, Santosh; Omer, Sumita; Patel, Krunal; Khan, Bashir M

    2013-02-01

    Leucaena leucocephala is a leguminous tree species accounting for one-fourth of raw material supplied to paper and pulp industry in India. Cinnamate 4-Hydroxylase (C4H, EC 1.14.13.11) is the second gene of phenylpropanoid pathway and a member of cytochrome P450 family. There is currently intense interest to alter or modify lignin content of L. leucocephala. Three highly similar C4H alleles of LlC4H1 gene were isolated and characterized. The alleles shared more than 98 % sequence identity at amino acid level to each other. Binding of partial promoter of another C4H gene LlC4H2, to varying amounts of crude nuclear proteins isolated from leaf and stem tissues of L. leucocephala formed two loose and one strong complex, respectively, suggesting that the abundance of proteins that bind with the partial C4H promoter is higher in stem tissue than in leaf tissue. Quantitative Real Time PCR study suggested that among tissues of same age, root tissues had highest level of C4H transcripts. Maximum transcript level was observed in 30 day old root tissue. Among the tissues investigated, C4H activity was highest in 60 day old root tissues. Tissue specific quantitative comparison of lignin from developing seedling stage to 1 year old tree stage indicated that Klason lignin increased in tissues with age.

  4. Cloning and characterization of the rat HIF-1 alpha prolyl-4-hydroxylase-1 gene.

    PubMed

    Cobb, Ronald R; McClary, John; Manzana, Warren; Finster, Silke; Larsen, Brent; Blasko, Eric; Pearson, Jennifer; Biancalana, Sara; Kauser, Katalin; Bringmann, Peter; Light, David R; Schirm, Sabine

    2005-08-01

    Prolyl-4-hydroxylase domain-containing enzymes (PHDs) mediate the oxygen-dependent regulation of the heterodimeric transcription factor hypoxia-inducible factor-1 (HIF-1). Under normoxic conditions, one of the subunits of HIF-1, HIF-1alpha, is hydroxylated on specific proline residues to target HIF-1alpha for degradation by the ubiquitin-proteasome pathway. Under hypoxic conditions, the hydroxylation by the PHDs is attenuated by lack of the oxygen substrate, allowing HIF-1 to accumulate, translocate to the nucleus, and mediate HIF-mediated gene transcription. In several mammalian species including humans, three PHDs have been identified. We report here the cloning of a full-length rat cDNA that is highly homologous to the human and murine PHD-1 enzymes and encodes a protein that is 416 amino acids long. Both cDNA and protein are widely expressed in rat tissues and cell types. We demonstrate that purified and crude baculovirus-expressed rat PHD-1 exhibits HIF-1alpha specific prolyl hydroxylase activity with similar substrate affinities and is comparable to human PHD-1 protein.

  5. Cinnamic acid 4-hydroxylase of sorghum [Sorghum biocolor (L.) Moench] gene SbC4H1 restricts lignin synthesis in Arabidopsis

    USDA-ARS?s Scientific Manuscript database

    Cinnamic acid 4-hydroxylase (C4H) is the first hydroxylase enzyme of the phenylpropanoid pathway, and its content and activity affects the lignin synthesis. In this study, we isolated a C4H gene SbC4H1 from the suppression subtractive hybridization library of brown midrib (bmr) mutants of Sorghum b...

  6. Virus induced gene silencing of three putative prolyl 4-hydroxylases enhances plant growth in tomato (Solanum lycopersicum).

    PubMed

    Fragkostefanakis, Sotirios; Sedeek, Khalid E M; Raad, Maya; Zaki, Marwa Samir; Kalaitzis, Panagiotis

    2014-07-01

    Proline hydroxylation is a major posttranslational modification of hydroxyproline-rich glycoproteins (HRGPs) that is catalyzed by prolyl 4-hydroxylases (P4Hs). HRGPs such as arabinogalactan proteins (AGPs) and extensios play significant roles on cell wall structure and function and their implication in cell division and expansion has been reported. We used tobacco rattle virus (TRV)-based virus induced gene silencing to investigate the role of three tomato P4Hs, out of ten present in the tomato genome, in growth and development. Eight-days old tomato seedlings were infected with the appropriate TRV vectors and plants were allowed to grow under standard conditions for 6 weeks. Lower P4H mRNA levels were associated with lower hydroxyproline content in root and shoot tissues indicating successful gene silencing. P4H-silenced plants had longer roots and shoots and larger leaves. The increased leaf area can be attributed to increased cell division as indicated by the higher leaf epidermal cell number in SlP4H1- and SlP4H9-silenced plants. In contrast, SlP4H7-silenced plants had larger leaves due to enhanced cell expansion. Western blot analysis revealed that silencing of SlP4H7 and SlP4H9 was associated with reduced levels of JIM8-bound AGP and JIM11-bound extensin epitopes, while silencing of SlP4H1 reduced only the levels of AGP proteins. Collectively these results show that P4Hs have significant and distinct roles in cell division and expansion of tomato leaves.

  7. No association between schizophrenia and polymorphisms within the genes for debrisoquine 4-hydroxylase (CYP2D6) and the dopamine transporter (DAT)

    SciTech Connect

    Daniels, J.; Williams, J.; Asherson, P.; McGuffin, P.; Owen, M.

    1995-02-27

    It has been suggested that the cytochrome P450 mono-oxygenase, debrisoquine 4-hydroxylase, is involved in the catabolism and processing of neurotransmitters subsequent to their reuptake into target cells. It is also thought to be related to the dopamine transporter that acts to take released dopamine back up into presynaptic terminals. The present study used the association approach to test the hypothesis that mutations in the genes for debrisoquine 4-hydroxylase (CYP2D6) and the dopamine transporter (DAT) confer susceptibility to schizophrenia. There were no differences in allele or genotype frequencies between patients and controls in the mutations causing the poor metaboliser phenotype in CYP2D6. In addition there was no association found between schizophrenia and a 48 bp repeat within the 3{prime} untranslated region of DAT. 18 refs., 2 tabs.

  8. Prolyl 4-hydroxylase

    PubMed Central

    Gorres, Kelly L.; Raines, Ronald T.

    2010-01-01

    Posttranslational modifications can cause profound changes in protein function. Typically, these modifications are reversible, and thus provide a biochemical on–off switch. In contrast, proline residues are the substrates for an irreversible reaction that is the most common posttranslational modification in humans. This reaction, which is catalyzed by prolyl 4-hydroxylase (P4H), yields (2S,4R)-4-hydroxyproline (Hyp). The protein substrates for P4Hs are diverse. Likewise, the biological consequences of prolyl hydroxylation vary widely, and include altering protein conformation and protein–protein interactions, and enabling further modification. The best known role for Hyp is in stabilizing the collagen triple helix. Hyp is also found in proteins with collagen-like domains, as well as elastin, conotoxins, and argonaute 2. A prolyl hydroxylase domain protein acts on the hypoxia inducible factor α, which plays a key role in sensing molecular oxygen, and could act on inhibitory κB kinase and RNA polymerase II. P4Hs are not unique to animals, being found in plants and microbes as well. Here, we review the enzymic catalysts of prolyl hydroxylation, along with the chemical and biochemical consequences of this subtle but abundant posttranslational modification. PMID:20199358

  9. Sphingolipid base modifying enzymes in sunflower (Helianthus annuus): cloning and characterization of a C4-hydroxylase gene and a new paralogous Δ8-desaturase gene.

    PubMed

    Moreno-Pérez, Antonio J; Martínez-Force, Enrique; Garcés, Rafael; Salas, Joaquín J

    2011-05-15

    Sphingolipids are components of plant cell membranes that participate in the regulation of important physiological processes. Unlike their animal counterparts, plant sphingolipids are characterized by high levels of base C4-hydroxylation. Moreover, desaturation at the Δ8 position predominates over the Δ4 desaturation typically found in animal sphingolipids. These modifications are due to the action of C4-hydroxylases and Δ8-long chain base desaturases, and they are important for complex sphingolipids finally becoming functional. The long chain bases of sunflower sphingolipids have high levels of hydroxylated and unsaturated moieties. Here, a C4-long chain base hydroxylase was functionally characterized in sunflower plant, an enzyme that could complement the sur2Δ mutation when heterologously expressed in this yeast mutant deficient in hydroxylation. This hydroxylase was ubiquitously expressed in sunflower, with the highest levels found in the developing cotyledons. In addition, we identified a new Δ8-long base chain desaturase gene that displays strong homology to a previously reported desaturase gene. This desaturase was also expressed in yeast and was able to change the long chain base composition of the transformed host. We studied the expression of this desaturase and compared it with that of the other isoform described in sunflower. The desaturase form studied in this paper displayed higher expression levels in developing seeds.

  10. Developmental role of phenylalanine-ammonia-lyase (PAL) and cinnamate 4-hydroxylase (C4H) genes during adventitious rooting of Juglans regia L. microshoots.

    PubMed

    Cheniany, Monireh; Ganjeali, Ali

    2016-12-01

    Phenylalanine-ammonia-lyase and cinnamate-4-hydroxylase play important role in the phenylpropanoid pathway, which produces many biologically important secondary metabolites participating in normal plant development. Flavonol quercetin is the main representant of these compounds that has been identified in numerous Juglans spp. In this survey, the developmental expression patterns of PAL and C4H genes during in vitro rooting of two walnut cultivars 'Sunland' and 'Howard' was examined by RT-PCR. To understand the potential role in rooting, the changing pattern of endogenous content of quercetin was also analyzed by HPLC. The 'Sunland' with better capacity to root had more quercetin content during the "inductive phase" of rooting than 'Howard'. In each cultivar, the level of PAL transcripts showed the same behavior with the changing patterns of quercetin during root formation of microshoots. The positive correlation between the changes of quercetin and PAL-mRNA indicated that PAL gene may have an immediate effect on flavonoid pathway metabolites including quercetin. Although the behavioral change of C4H expression was similar in both cultivars during root formation (with significantly more level for 'Howard'), it was not coincide with the changes of quercerin concentrations. Our results showed that C4H function is important for the normal development, but its transcriptional regulation does not correlate with quercetin as an efficient phenolic compound for walnut rhizogenesis.

  11. Proline with or without hydroxyproline influences collagen concentration and regulates prolyl 4-hydroxylase α (I) gene expression in juvenile turbo ( Scophthalmus maximus L.)

    NASA Astrophysics Data System (ADS)

    Zhang, Kaikai; Mai, Kangsen; Xu, Wei; Zhou, Huihui; Liufu, Zhiguo; Zhang, Yanjiao; Peng, Mo; Ai, Qinghui

    2015-06-01

    This study was conducted to investigate the effect of dietary proline (Pro), and Pro and hydroxyproline (Hyp) in combination on the growth performance, total Hyp and collagen concentrations of tissues, and prolyl 4-hydroxylase α(I) (P4H α(I)) gene expression in juvenile turbot feeding high plant protein diets. A diet containing 50% crude protein and 12% crude lipid was formulated as the basal and control, on which other two protein and lipid contents identical experimental diets were formulated by supplementing the basal with either 0.75% Pro (Pro-0.75) or 0.75% Pro and 0.75% Hyp (Pro+Hyp). Four groups of fish in indoor seawater recirculating systems, 35 individuals each, were fed twice a day to apparent satiation for 10 weeks. The results showed that dietary Pro and Hyp supplementation had no significant effect on growth performance and feed utilization of juvenile turbot (P > 0.05). Total Hyp and collagen concentrations in muscle were significantly increased when dietary Pro and Hyp increased (P <0.05), and fish fed diet Pro+Hyp showed significantly higher free Hyp content in plasma than those fed other diets (P <0.05). The expression of P4H a(I) gene in liver and muscle was significantly up regulated in fish fed diet Pro-0.75 in comparison with control (P <0.05); however the gene was significantly down regulated in fish fed diet Pro+Hyp in muscle in comparison with fish fed diet Pro-0.75 (P <0.05). It can be concluded that supplement of crystal L-Pro and L-Hyp to high plant protein diets did not show positive effects on growth performance of juvenile turbot, but enhanced total collagen concentrations in muscle.

  12. Fermentation process for tetrameric human collagen prolyl 4-hydroxylase in Escherichia coli: improvement by gene optimisation of the PDI/beta subunit and repeated addition of the inducer anhydrotetracycline.

    PubMed

    Neubauer, Antje; Soini, Jaakko; Bollok, Monika; Zenker, Minette; Sandqvist, Janne; Myllyharju, Johanna; Neubauer, Peter

    2007-02-01

    The collagen prolyl 4-hydroxylases (C-P4Hs) that reside within the lumen of the endoplasmic reticulum (ER) are the key enzymes in the biosynthesis of collagens. The vertebrate enzymes are alpha(2)beta(2) tetramers consisting of two catalytic alpha subunits and two beta subunits that are identical to protein disulfide isomerase (PDI). Cytoplasmic production of an active human C-P4H has recently been described in the Origami (trxB gor) mutant Escherichia coli using a bicistronic vector with independent control of the alpha and PDI/beta subunit expression by the tetA and T5-lac promoters, respectively, enabling sequential induction (Neubauer, A., Neubauer, P., Myllyharju, J., 2005. High-level production of human collagen prolyl 4-hydroxylase in Escherichia coli. Matrix Biol. 24, 59-68). We show here that the yield of active C-P4H in shake flasks is increased 50-fold by improving the expression level of the PDI/beta subunit through gene optimisation. We also found that stable expression of the alpha subunit mRNA in a fed-batch fermentation process requires repeated additions of anhydrotetracycline. This finding may be of a wider general importance for the use of the tetA promoter in fed-batch cultivations, especially if recombinant proteins are expressed during long production phases. We also show that growth of the E. coli Origami strain to high cell density on a complex medium with consecutive sequential induction is difficult to achieve and that optimisation of similarly complicated systems can greatly benefit from the use of quantitative mRNA analysis for the evaluation of transcriptional bottlenecks. The optimisation approach resulted in a fermentation yield of 143 mg L(-1) of active C-P4H, corresponding to approximately 7.5% of the total soluble cell protein.

  13. Transmembrane prolyl 4-hydroxylase is a fourth prolyl 4-hydroxylase regulating EPO production and erythropoiesis.

    PubMed

    Laitala, Anu; Aro, Ellinoora; Walkinshaw, Gail; Mäki, Joni M; Rossi, Maarit; Heikkilä, Minna; Savolainen, Eeva-Riitta; Arend, Michael; Kivirikko, Kari I; Koivunen, Peppi; Myllyharju, Johanna

    2012-10-18

    An endoplasmic reticulum transmembrane prolyl 4-hydroxylase (P4H-TM) is able to hydroxylate the α subunit of the hypoxia-inducible factor (HIF) in vitro and in cultured cells, but nothing is known about its roles in mammalian erythropoiesis. We studied such roles here by administering a HIF-P4H inhibitor, FG-4497, to P4h-tm(-/-) mice. This caused larger increases in serum Epo concentration and kidney but not liver Hif-1α and Hif-2α protein and Epo mRNA levels than in wild-type mice, while the liver Hepcidin mRNA level was lower in the P4h-tm(-/-) mice than in the wild-type. Similar, but not identical, differences were also seen between FG-4497-treated Hif-p4h-2 hypomorphic (Hif-p4h-2(gt/gt)) and Hif-p4h-3(-/-) mice versus wild-type mice. FG-4497 administration increased hemoglobin and hematocrit values similarly in the P4h-tm(-/-) and wild-type mice, but caused higher increases in both values in the Hif-p4h-2(gt/gt) mice and in hematocrit value in the Hif-p4h-3(-/-) mice than in the wild-type. Hif-p4h-2(gt/gt)/P4h-tm(-/-) double gene-modified mice nevertheless had increased hemoglobin and hematocrit values without any FG-4497 administration, although no such abnormalities were seen in the Hif-p4h-2(gt/gt) or P4h-tm(-/-) mice. Our data thus indicate that P4H-TM plays a role in the regulation of EPO production, hepcidin expression, and erythropoiesis.

  14. Characterization of two carnation petal prolyl 4 hydroxylases.

    PubMed

    Vlad, Florina; Tiainen, Päivi; Owen, Carolyn; Spano, Thodhoraq; Daher, Firas Bou; Oualid, Fatiha; Senol, Namik Ozer; Vlad, Daniela; Myllyharju, Johanna; Kalaitzis, Panagiotis

    2010-10-01

    Prolyl 4-hydroxylases (P4Hs) catalyze the proline hydroxylation, a major post-translational modification, of hydroxyproline-rich glycoproteins. Two carnation petal P4H cDNAs, (Dianthus caryophyllus prolyl 4-hydroxylase) DcP4H1 and DcP4H2, were identified and characterized at the gene expression and biochemical level in order to investigate their role in flower senescence. Both mRNAs showed similar patterns of expression with stable transcript abundance during senescence progression and differential tissue-specific expression with DcP4H1 and DcP4H2 strongly expressed in ovaries and stems, respectively. Recombinant DcP4H1 and DcP4H2 proteins were produced and their catalytic properties were determined. Pyridine 2,4-dicarboxylate (PDCA) was identified as a potent inhibitor of the in vitro enzyme activity of both P4Hs and used to determine whether inhibition of proline hydroxylation in petals is involved in senescence progression of cut carnation flowers. PDCA suppressed the climacteric ethylene production indicating a strong correlation between the inhibition of DcP4H1 and DcP4H2 activity in vitro by PDCA and the suppression of climacteric ethylene production in cut carnation flowers.

  15. Regulation and Functional Expression of Cinnamate 4-Hydroxylase from Parsley

    PubMed Central

    Koopmann, Edda; Logemann, Elke; Hahlbrock, Klaus

    1999-01-01

    A previously isolated parsley (Petroselinum crispum) cDNA with high sequence similarity to cinnamate 4-hydroxylase (C4H) cDNAs from several plant sources was expressed in yeast (Saccharomyces cerevisiae) containing a plant NADPH:cytochrome P450 oxidoreductase and verified as encoding a functional C4H (CYP73A10). Low genomic complexity and the occurrence of a single type of cDNA suggest the existence of only one C4H gene in parsley. The encoded mRNA and protein, in contrast to those of a functionally related NADPH:cytochrome P450 oxidoreductase, were strictly coregulated with phenylalanine ammonia-lyase mRNA and protein, respectively, as demonstrated by coinduction under various conditions and colocalization in situ in cross-sections from several different parsley tissues. These results support the hypothesis that the genes encoding the core reactions of phenylpropanoid metabolism form a tight regulatory unit. PMID:9880345

  16. Expression analysis of kenaf cinnamate 4-hydroxylase (C4H) ortholog during developmental and stress responses

    USDA-ARS?s Scientific Manuscript database

    This study was conducted to clone and analyze the expression pattern of a C4H gene encoding cinnamate 4-hydroxylase from kenaf (Hibiscus cannabinus L.). A full-length C4H ortholog was cloned using degenerate primers and the RACE (rapid amplification of cDNA ends) method. The full-length C4H ortholog...

  17. Down-regulation of p-coumaroyl quinate/shikimate 3'-hydroxylase (C3'H) and cinnamate 4-hydroxylase (C4H) genes in the lignin biosynthetic pathway of Eucalyptus urophylla x E. grandis leads to improved sugar release

    DOE PAGES

    Sykes, Robert W.; Gjersing, Erica L.; Foutz, Kirk; ...

    2015-08-27

    In this study, lignocellulosic materials provide an attractive replacement for food-based crops used to produce ethanol. Understanding the interactions within the cell wall is vital to overcome the highly recalcitrant nature of biomass. One factor imparting plant cell wall recalcitrance is lignin, which can be manipulated by making changes in the lignin biosynthetic pathway. In this study, eucalyptus down-regulated in expression of cinnamate 4-hydroxylase (C4H, EC 1.14.13.11) or p-coumaroyl quinate/shikimate 3'-hydroxylase (C3'H, EC 1.14.13.36) were evaluated for cell wall composition and reduced recalcitrance.

  18. Hypoxia-inducible Factor Prolyl 4-Hydroxylase Inhibition A TARGET FOR NEUROPROTECTION IN THE CENTRAL NERVOUS SYSTEM*

    PubMed Central

    Siddiq, Ambreena; Ayoub, Issam A.; Chavez, Juan C.; Aminova, Leila; Shah, Sapan; LaManna, Joseph C.; Patton, Stephanie M.; Connor, James R.; Cherny, Robert A.; Volitakis, Irene; Bush, Ashley I.; Langsetmo, Ingrid; Seeley, Todd; Gunzler, Volkmar; Ratan, Rajiv R.

    2008-01-01

    Hypoxia-inducible factor (HIF) prolyl 4-hydroxylases are a family of iron- and 2-oxoglutarate-dependent dioxygenases that negatively regulate the stability of several proteins that have established roles in adaptation to hypoxic or oxidative stress. These proteins include the transcriptional activators HIF-1α and HIF-2α. The ability of the inhibitors of HIF prolyl 4-hydroxylases to stabilize proteins involved in adaptation in neurons and to prevent neuronal injury remains unclear. We reported that structurally diverse low molecular weight or peptide inhibitors of the HIF prolyl 4-hydroxylases stabilize HIF-1α and up-regulate HIF-dependent target genes (e.g. enolase, p21waf1/cip1, vascular endothelial growth factor, or erythropoietin) in embryonic cortical neurons in vitro or in adult rat brains in vivo. We also showed that structurally diverse HIF prolyl 4-hydroxylase inhibitors prevent oxidative death in vitro and ischemic injury in vivo. Taken together these findings identified low molecular weight and peptide HIF prolyl 4-hydroxylase inhibitors as novel neurological therapeutics for stroke as well as other diseases associated with oxidative stress. PMID:16227210

  19. RNAi down-regulation of cinnamate-4-hydroxylase increases artemisinin biosynthesis in Artemisia annua

    PubMed Central

    Kumar, Ritesh; Vashisth, Divya; Misra, Amita; Akhtar, Md Qussen; Jalil, Syed Uzma; Shanker, Karuna; Gupta, Madan Mohan; Rout, Prashant Kumar; Gupta, Anil Kumar; Shasany, Ajit Kumar

    2016-01-01

    Cinnamate-4-hydroxylase (C4H) converts trans-cinnamic acid (CA) to p-coumaric acid (COA) in the phenylpropanoid/lignin biosynthesis pathway. Earlier we reported increased expression of AaCYP71AV1 (an important gene of artemisinin biosynthesis pathway) caused by CA treatment in Artemisia annua. Hence, AaC4H gene was identified, cloned, characterized and silenced in A. annua with the assumption that the elevated internal CA due to knock down may increase the artemisinin yield. Accumulation of trans-cinnamic acid in the plant due to AaC4H knockdown was accompanied with the reduction of p-coumaric acid, total phenolics, anthocyanin, cinnamate-4-hydroxylase (C4H) and phenylalanine ammonia lyase (PAL) activities but increase in salicylic acid (SA) and artemisinin. Interestingly, feeding trans-cinnamic acid to the RNAi line increased the level of artemisinin along with benzoic (BA) and SA with no effect on the downstream metabolites p-coumaric acid, coniferylaldehyde and sinapaldehyde, whereas p-coumaric acid feeding increased the content of downstream coniferylaldehyde and sinapaldehyde with no effect on BA, SA, trans-cinnamic acid or artemisinin. SA is reported earlier to be inducing the artemisinin yield. This report demonstrates the link between the phenylpropanoid/lignin pathway with artemisinin pathway through SA, triggered by accumulation of trans-cinnamic acid because of the blockage at C4H. PMID:27220407

  20. Bioavailable affinity label for collagen prolyl 4-hydroxylase

    PubMed Central

    Vasta, James D.; Higgin, Joshua J.; Kersteen, Elizabeth A.

    2013-01-01

    Collagen is the most abundant protein in animals. Its prevalent 4-hydroxyproline residues contribute greatly to its conformational stability. The hydroxyl groups arise from a post-translational modification catalyzed by the non-heme iron-dependent enzyme, collagen prolyl 4-hydroxylase (P4H). Here, we report that 4-oxo-5,6-epoxyhexanoate, a mimic of the α-ketoglutarate co-substrate, inactivates human P4H. The inactivation installs a ketone functionality in P4H, providing a handle for proteomic experiments. Caenorhabditis elegans exposed to the esterified epoxy ketone displays the phenotype of a worm lacking P4H. Thus, this affinity label can be used to mediate collagen stability in an animal, as is desirable in the treatment of a variety of fibrotic diseases. PMID:23702396

  1. Down-regulation of p-coumaroyl quinate/shikimate 3'-hydroxylase (C3'H) and cinnamate 4-hydroxylase (C4H) genes in the lignin biosynthetic pathway of Eucalyptus urophylla × E. grandis leads to improved sugar release.

    PubMed

    Sykes, Robert W; Gjersing, Erica L; Foutz, Kirk; Rottmann, William H; Kuhn, Sean A; Foster, Cliff E; Ziebell, Angela; Turner, Geoffrey B; Decker, Stephen R; Hinchee, Maud A W; Davis, Mark F

    2015-01-01

    Lignocellulosic materials provide an attractive replacement for food-based crops used to produce ethanol. Understanding the interactions within the cell wall is vital to overcome the highly recalcitrant nature of biomass. One factor imparting plant cell wall recalcitrance is lignin, which can be manipulated by making changes in the lignin biosynthetic pathway. In this study, eucalyptus down-regulated in expression of cinnamate 4-hydroxylase (C4H, EC 1.14.13.11) or p-coumaroyl quinate/shikimate 3'-hydroxylase (C3'H, EC 1.14.13.36) were evaluated for cell wall composition and reduced recalcitrance. Eucalyptus trees with down-regulated C4H or C3'H expression displayed lowered overall lignin content. The control samples had an average of 29.6 %, the C3'H reduced lines had an average of 21.7 %, and the C4H reduced lines had an average of 18.9 % lignin from wet chemical analysis. The C3'H and C4H down-regulated lines had different lignin compositions with average S/G/H ratios of 48.5/33.2/18.3 for the C3'H reduced lines and 59.0/39.8/1.2 for the C4H reduced lines, compared to the control with 65.9/33.2/1.0. Both the C4H and C3'H down-regulated lines had reduced recalcitrance as indicated by increased sugar release as determined using enzymatic conversion assays utilizing both no pretreatment and a hot water pretreatment. Lowering lignin content rather than altering sinapyl alcohol/coniferyl alcohol/4-coumaryl alcohol ratios was found to have the largest impact on reducing recalcitrance of the transgenic eucalyptus variants. The development of lower recalcitrance trees opens up the possibility of using alternative pretreatment strategies in biomass conversion processes that can reduce processing costs.

  2. Inhibition of prolyl 4-hydroxylase decreases muscle fibrosis following chronic rotator cuff tear

    PubMed Central

    Gumucio, J. P.; Flood, M. D.; Bedi, A.; Kramer, H. F.; Russell, A. J.

    2017-01-01

    Objectives Rotator cuff tears are among the most frequent upper extremity injuries. Current treatment strategies do not address the poor quality of the muscle and tendon following chronic rotator cuff tears. Hypoxia-inducible factor-1 alpha (HIF-1α) is a transcription factor that activates many genes that are important in skeletal muscle regeneration. HIF-1α is inhibited under normal physiological conditions by the HIF prolyl 4-hydroxylases (PHDs). In this study, we used a pharmacological PHD inhibitor, GSK1120360A, to enhance the activity of HIF-1α following the repair of a chronic cuff tear, and measured muscle fibre contractility, fibrosis, gene expression, and enthesis mechanics. Methods Chronic supraspinatus tears were induced in adult rats, and repaired 28 days later. Rats received 0 mg/kg, 3 mg/kg, or 10 mg/kg GSK1120360A daily. Collagen content, contractility, fibre type distribution and size, the expression of genes involved in fibrosis, lipid accumulation, atrophy and inflammation, and the mechanical properties of the enthesis were then assessed two weeks following surgical repair. Results At two weeks following repair, treatment groups showed increased muscle mass but there was a 15% decrease in force production in the 10 mg/kg group from controls, and no difference between the 0 mg/kg and the 3 mg/kg groups. There was a decrease in the expression of several gene transcripts related to matrix accumulation and fibrosis, and a 50% decrease in collagen content in both treated groups compared with controls. Additionally, the expression of inflammatory genes was reduced in the treated groups compared with controls. Finally, PHD inhibition improved the maximum stress and displacement to failure in repaired tendons. Conclusions GSK1120360A resulted in improved enthesis mechanics with variable effects on muscle function. PHD inhibition may be beneficial for connective tissue injuries in which muscle atrophy has not occurred. Cite this article: J. P. Gumucio

  3. Inhibition of prolyl 4-hydroxylase decreases muscle fibrosis following chronic rotator cuff tear.

    PubMed

    Gumucio, J P; Flood, M D; Bedi, A; Kramer, H F; Russell, A J; Mendias, C L

    2017-01-01

    Rotator cuff tears are among the most frequent upper extremity injuries. Current treatment strategies do not address the poor quality of the muscle and tendon following chronic rotator cuff tears. Hypoxia-inducible factor-1 alpha (HIF-1α) is a transcription factor that activates many genes that are important in skeletal muscle regeneration. HIF-1α is inhibited under normal physiological conditions by the HIF prolyl 4-hydroxylases (PHDs). In this study, we used a pharmacological PHD inhibitor, GSK1120360A, to enhance the activity of HIF-1α following the repair of a chronic cuff tear, and measured muscle fibre contractility, fibrosis, gene expression, and enthesis mechanics. Chronic supraspinatus tears were induced in adult rats, and repaired 28 days later. Rats received 0 mg/kg, 3 mg/kg, or 10 mg/kg GSK1120360A daily. Collagen content, contractility, fibre type distribution and size, the expression of genes involved in fibrosis, lipid accumulation, atrophy and inflammation, and the mechanical properties of the enthesis were then assessed two weeks following surgical repair. At two weeks following repair, treatment groups showed increased muscle mass but there was a 15% decrease in force production in the 10 mg/kg group from controls, and no difference between the 0 mg/kg and the 3 mg/kg groups. There was a decrease in the expression of several gene transcripts related to matrix accumulation and fibrosis, and a 50% decrease in collagen content in both treated groups compared with controls. Additionally, the expression of inflammatory genes was reduced in the treated groups compared with controls. Finally, PHD inhibition improved the maximum stress and displacement to failure in repaired tendons. GSK1120360A resulted in improved enthesis mechanics with variable effects on muscle function. PHD inhibition may be beneficial for connective tissue injuries in which muscle atrophy has not occurred.Cite this article: J. P. Gumucio, M. D. Flood, A. Bedi, H. F. Kramer, A. J

  4. Hydroxylation of Human Type III Collagen Alpha Chain by Recombinant Coexpression with a Viral Prolyl 4-Hydroxylase in Escherichia coli.

    PubMed

    Shi, Jingjing; Ma, Xiaoxuan; Gao, Yuan; Fan, Daidi; Zhu, Chenhui; Mi, Yu; Xue, Wenjiao

    2017-08-01

    High-level expression of recombinant collagen by genetic engineering is urgently required. Recombinant collagen is different from natural collagen in its hydroxyproline (Hyp) content and thermal stability. To obtain hydroxylated collagen for applications in biomedicine and biomaterials, the human collagen α1(III) chain was co-expressed with the viral prolyl 4-hydroxylase A085R in Escherichia coli. Unlike previous reports using human prolyl 4-hydroxylase, this study examined the hydroxylation of full-length human collagen α1(III) chain (COL3A1) by viral prolyl 4-hydroxylase. The genes encoding these two proteins were controlled by different promoters, Ptac and PRPL, on a recombinant pKK223-3 plasmid. The sequencing results verified that the target genes were successfully inserted into the recombinant vector. Based on quantitative PCR, SDS-PAGE, and western blotting, successful expression by E. coli BL21(DE3) was detected at the mRNA and protein levels for both loci. Liquid chromatography-mass spectrometry (LC-MS/MS) results suggested that the highest Hyp yield was obtained when the two proteins were induced with 0.5 mM IPTG and heat-shock treatment at 50 °C, corresponding to high enzyme expression and low human collagen α1(III) chain expression levels. A biological activity analysis indicated that the recombinant collagen with the highest hydroxylation level supported the growth of baby hamster kidney cells, similar to observations for native collagen. The production of hydroxylated collagen in this study establishes a new method for collagen hydroxylation and provides a basis for the application of recombinant collagen expressed in E. coli.

  5. Down-regulation of p-coumaroyl quinate/shikimate 3'-hydroxylase (C3'H) and cinnamate 4-hydroxylase (C4H) genes in the lignin biosynthetic pathway of Eucalyptus urophylla x E. grandis leads to improved sugar release

    SciTech Connect

    Sykes, Robert W.; Gjersing, Erica L.; Foutz, Kirk; Rottmann, William H.; Kuhn, Sean A.; Foster, Cliff E.; Ziebell, Angela; Turner, Geoffrey B.; Decker, Stephen R.; Hinchee, Maud A. W.; Davis, Mark F.

    2015-08-27

    In this study, lignocellulosic materials provide an attractive replacement for food-based crops used to produce ethanol. Understanding the interactions within the cell wall is vital to overcome the highly recalcitrant nature of biomass. One factor imparting plant cell wall recalcitrance is lignin, which can be manipulated by making changes in the lignin biosynthetic pathway. In this study, eucalyptus down-regulated in expression of cinnamate 4-hydroxylase (C4H, EC 1.14.13.11) or p-coumaroyl quinate/shikimate 3'-hydroxylase (C3'H, EC 1.14.13.36) were evaluated for cell wall composition and reduced recalcitrance.

  6. Prolyl-4-hydroxylase α subunit 2 promotes breast cancer progression and metastasis by regulating collagen deposition.

    PubMed

    Xiong, Gaofeng; Deng, Lei; Zhu, Jieqing; Rychahou, Piotr G; Xu, Ren

    2014-01-02

    Increased collagen deposition provides physical and biochemical signals to support tumor growth and invasion during breast cancer development. Therefore, inhibition of collagen synthesis and deposition has been considered a strategy to suppress breast cancer progression. Collagen prolyl-4-hydroxylase α subunit 2 (P4HA2), an enzyme hydroxylating proline residues in -X-Pro-Gly- sequences, is a potential therapeutic target for the disorders associated with increased collagen deposition. However, expression and function of P4HA2 in breast cancer progression are not well investigated. Gene co-expression analysis was performed in the published microarray datasets to identify potential regulators of collagen I, III, and IV in human breast cancer tissue. Expression of P4HA2 was silenced by shRNAs, and its activity was inhibited by 1, 4-DPCA, a prolyl-4-hydroxylase inhibitor. Three-dimensional culture assay was used to analyze roles of P4HA2 in regulating malignant phenotypes of breast cancer cells. Reduced deposition of collagen I and IV was detected by Western blotting and immunofluorescence. Control and P4HA2-silenced breast cancer cells were injected into fat pad and tail vein of SCID mice to examine effect of P4HA2 on tumor growth and lung metastasis. Using gene co-expression analysis, we showed that P4HA2 was associated with expression of Col1A1, Col3A1, and Col4A1 during breast cancer development and progression. P4HA2 mRNA levels were significantly upregulated in breast cancer compared to normal mammary tissue. Increased mRNA levels of P4HA2 correlated with poor clinical outcome in breast cancer patients, which is independent of estrogen receptor status. Silencing P4HA2 expression or treatment with the P4HA inhibitor significantly inhibited cell proliferation and suppressed aggressive phenotypes of breast cancer cells in 3D culture, accompanied by reduced deposition of collagen I and IV. We also found that knockdown of P4HA2 inhibited mammary tumor growth and

  7. Cinnamate-4-hydroxylase expression in Arabidopsis. Regulation in response to development and the environment.

    PubMed Central

    Bell-Lelong, D A; Cusumano, J C; Meyer, K; Chapple, C

    1997-01-01

    Cinnamate-4-hydroxylase (C4H) is the first Cyt P450-dependent monooxygenase of the phenylpropanoid pathway. To study the expression of this gene in Arabidopsis thaliana, a C4H cDNA clone from the Arabidopsis expressed sequence tag database was identified and used to isolate its corresponding genomic clone. The entire C4H coding sequence plus 2.9 kb of its promoter were isolated on a 5.4-kb HindIII fragment of this cosmid. Inspection of the promoter sequence revealed the presence of a number of putative regulatory motifs previously identified in the promoters of other phenylpropanoid pathway genes. The expression of C4H was analyzed by RNA blot hybridization analysis and in transgenic Arabidopsis carrying a C4H-beta-glucuronidase transcriptional fusion. C4H message accumulation was light-dependent, but was detectable even in dark-grown seedlings. Consistent with these data, C4H mRNA was accumulated to light-grown levels in etiolated det1-1 mutant seedlings. C4H is widely expressed in various Arabidopsis tissues, particularly in roots and cells undergoing lignification. The C4H-driven beta-glucuronidase expression accurately reflected the tissue-specificity and wound-inducibility of the C4H promoter indicated by RNA blot hybridization analysis. A modest increase in C4H expression was observed in the tt8 mutant of Arabidopsis. PMID:9085570

  8. miR-190 Enhances HIF-Dependent Responses to Hypoxia in Drosophila by Inhibiting the Prolyl-4-hydroxylase Fatiga

    PubMed Central

    De Lella Ezcurra, Ana Laura; Bertolin, Agustina Paola; Kim, Kevin; Gándara, Lautaro; Luschnig, Stefan; Perrimon, Norbert; Melani, Mariana; Wappner, Pablo

    2016-01-01

    Cellular and systemic responses to low oxygen levels are principally mediated by Hypoxia Inducible Factors (HIFs), a family of evolutionary conserved heterodimeric transcription factors, whose alpha- and beta-subunits belong to the bHLH-PAS family. In normoxia, HIFα is hydroxylated by specific prolyl-4-hydroxylases, targeting it for proteasomal degradation, while in hypoxia the activity of these hydroxylases decreases due to low oxygen availability, leading to HIFα accumulation and expression of HIF target genes. To identify microRNAs required for maximal HIF activity, we conducted an overexpression screen in Drosophila melanogaster, evaluating the induction of a HIF transcriptional reporter. miR-190 overexpression enhanced HIF-dependent biological responses, including terminal sprouting of the tracheal system, while in miR-190 loss of function embryos the hypoxic response was impaired. In hypoxic conditions, miR-190 expression was upregulated and required for induction of HIF target genes by directly inhibiting the HIF prolyl-4-hydroxylase Fatiga. Thus, miR-190 is a novel regulator of the hypoxia response that represses the oxygen sensor Fatiga, leading to HIFα stabilization and enhancement of hypoxic responses. PMID:27223464

  9. miR-190 Enhances HIF-Dependent Responses to Hypoxia in Drosophila by Inhibiting the Prolyl-4-hydroxylase Fatiga.

    PubMed

    De Lella Ezcurra, Ana Laura; Bertolin, Agustina Paola; Kim, Kevin; Katz, Maximiliano Javier; Gándara, Lautaro; Misra, Tvisha; Luschnig, Stefan; Perrimon, Norbert; Melani, Mariana; Wappner, Pablo

    2016-05-01

    Cellular and systemic responses to low oxygen levels are principally mediated by Hypoxia Inducible Factors (HIFs), a family of evolutionary conserved heterodimeric transcription factors, whose alpha- and beta-subunits belong to the bHLH-PAS family. In normoxia, HIFα is hydroxylated by specific prolyl-4-hydroxylases, targeting it for proteasomal degradation, while in hypoxia the activity of these hydroxylases decreases due to low oxygen availability, leading to HIFα accumulation and expression of HIF target genes. To identify microRNAs required for maximal HIF activity, we conducted an overexpression screen in Drosophila melanogaster, evaluating the induction of a HIF transcriptional reporter. miR-190 overexpression enhanced HIF-dependent biological responses, including terminal sprouting of the tracheal system, while in miR-190 loss of function embryos the hypoxic response was impaired. In hypoxic conditions, miR-190 expression was upregulated and required for induction of HIF target genes by directly inhibiting the HIF prolyl-4-hydroxylase Fatiga. Thus, miR-190 is a novel regulator of the hypoxia response that represses the oxygen sensor Fatiga, leading to HIFα stabilization and enhancement of hypoxic responses.

  10. Role of NonO-histone interaction in TNFalpha-suppressed prolyl-4-hydroxylase alpha1.

    PubMed

    Zhang, Cheng; Zhang, Ming-Xiang; Shen, Ying H; Burks, Jared K; Li, Xiao-Nan; LeMaire, Scott A; Yoshimura, Koichi; Aoki, Hiroki; Matsuzaki, Masunori; An, Feng-Shuang; Engler, David A; Matsunami, Risë K; Coselli, Joseph S; Zhang, Yun; Wang, Xing Li

    2008-08-01

    Inflammation is a key process in cardiovascular diseases. The extracellular matrix (ECM) of the vasculature is a major target of inflammatory cytokines, and TNFalpha regulates ECM metabolism by affecting collagen production. In this study, we have examined the pathways mediating TNFalpha-induced suppression of prolyl-4 hydroxylase alpha1 (P4Halpha1), the rate-limiting isoform of P4H responsible for procollagen hydroxylation, maturation, and organization. Using human aortic smooth muscle cells, we found that TNFalpha activated the MKK4-JNK1 pathway, which induced histone (H) 4 lysine 12 acetylation within the TNFalpha response element in the P4Halpha1 promoter. The acetylated-H4 then recruited a transcription factor, NonO, which, in turn, recruited HDACs and induced H3 lysine 9 deacetylation, thereby inhibiting transcription of the P4Halpha1 promoter. Furthermore, we found that TNFalpha oxidized DJ-1, which may be essential for the NonO-P4Halpha1 interaction because treatment with gene specific siRNA to knockout DJ-1 eliminated the TNFalpha-induced NonO-P4Halpha1 interaction and its suppression. Our findings may be relevant to aortic aneurysm and dissection and the stability of the fibrous cap of atherosclerotic plaque in which collagen metabolism is important in arterial remodeling. Defining this cytokine-mediated regulatory pathway may provide novel molecular targets for therapeutic intervention in preventing plaque rupture and acute coronary occlusion.

  11. Notch Downregulation and Extramedullary Erythrocytosis in Hypoxia-Inducible Factor Prolyl 4-Hydroxylase 2-Deficient Mice.

    PubMed

    Myllymäki, Mikko N M; Määttä, Jenni; Dimova, Elitsa Y; Izzi, Valerio; Väisänen, Timo; Myllyharju, Johanna; Koivunen, Peppi; Serpi, Raisa

    2017-01-15

    Erythrocytosis is driven mainly by erythropoietin, which is regulated by hypoxia-inducible factor (HIF). Mutations in HIF prolyl 4-hydroxylase 2 (HIF-P4H-2) (PHD2/EGLN1), the major downregulator of HIFα subunits, are found in familiar erythrocytosis, and large-spectrum conditional inactivation of HIF-P4H-2 in mice leads to severe erythrocytosis. Although bone marrow is the primary site for erythropoiesis, spleen remains capable of extramedullary erythropoiesis. We studied HIF-P4H-2-deficient (Hif-p4h-2(gt/gt)) mice, which show slightly induced erythropoiesis upon aging despite nonincreased erythropoietin levels, and identified spleen as the site of extramedullary erythropoiesis. Splenic hematopoietic stem cells (HSCs) of these mice exhibited increased erythroid burst-forming unit (BFU-E) growth, and the mice were protected against anemia. HIF-1α and HIF-2α were stabilized in the spleens, while the Notch ligand genes Jag1, Jag2, and Dll1 and target Hes1 became downregulated upon aging HIF-2α dependently. Inhibition of Notch signaling in wild-type spleen HSCs phenocopied the increased BFU-E growth. HIFα stabilization can thus mediate non-erythropoietin-driven splenic erythropoiesis via altered Notch signaling.

  12. A steady-state kinetic analysis of the prolyl-4-hydroxylase mechanism.

    PubMed

    Soskel, N T; Kuby, S A

    1981-01-01

    Published kinetic data by Kivirikko, et al. on the prolyl-4-hydroxylase reaction have been re-evaluated using the overall steady-state velocity equation in the forward and reverse directions for an ordered ter ter kinetic mechanism. Qualitatively, the published data for prolyl-4-hydroxylase appear to fit the predicted patterns for this kinetic mechanism. More kinetic data are needed to confirm these results and to quantitate the kinetic parameters but, tentatively, the order of substrate addition would appear to be alpha-ketoglutarate, oxygen, and peptide; and the order of product release would be hydroxylated peptide (or collagen), carbon dioxide, and succinate.

  13. Functional Characterization and Subcellular Localization of Poplar (Populus trichocarpa × Populus deltoides) Cinnamate 4-Hydroxylase1

    PubMed Central

    Ro, Dae Kyun; Mah, Nancy; Ellis, Brian E.; Douglas, Carl J.

    2001-01-01

    Cinnamic acid 4-hydroxylase (C4H), a member of the cytochrome P450 monooxygenase superfamily, plays a central role in phenylpropanoid metabolism and lignin biosynthesis and possibly anchors a phenylpropanoid enzyme complex to the endoplasmic reticulum (ER). A full-length cDNA encoding C4H was isolated from a hybrid poplar (Populus trichocarpa × P. deltoides) young leaf cDNA library. RNA-blot analysis detected C4H transcripts in all organs tested, but the gene was most highly expressed in developing xylem. C4H expression was also strongly induced by elicitor-treatment in poplar cell cultures. To verify the catalytic activity of the putative C4H cDNA, two constructs, C4H and C4H fused to the FLAG epitope (C4H::FLAG), were expressed in yeast. Immunoblot analysis showed that C4H was present in the microsomal fraction and microsomal preparations from strains expressing both enzymes efficiently converted cinnamic acid to p-coumaric acid with high specific activities. To investigate the subcellular localization of C4H in vivo, a chimeric C4H-green fluorescent protein (GFP) gene was engineered and stably expressed in Arabidopsis. Confocal laser microscopy analysis clearly showed that in Arabidopsis the C4H::GFP chimeric enzyme was localized to the ER. When expressed in yeast, the C4H::GFP fusion enzyme was also active but displayed significantly lower specific activity than either C4H or C4H::FLAG in in vitro and in vivo enzyme assays. These data definitively show that C4H is localized to the ER in planta. PMID:11351095

  14. Overexpression of cinnamate 4-hydroxylase and 4-coumaroyl CoA ligase prompted flavone accumulation in Scutellaria baicalensis hairy roots.

    PubMed

    Kim, Young Seon; Kim, Yeon Bok; Kim, YeJi; Lee, Mi Young; Park, Sang Un

    2014-06-01

    Scutellaria baicalensis Georgi, a species of the Lamiaceae family, is considered as one of the 50 fundamental herbs used in traditional Chinese medicine. In order to enhance flavone (baicalein, baicalin, and wogonin) content in S. baicalensis roots, we overexpressed a single gene of cinnamate 4-hydroxylase (C4H) and 4-coumaroyl coenzyme A ligase (4CL) using an Agrobacterium rhizogenes-mediated system. SbC4H- and Sb4CL-overexpressed hairy root lines enhanced the transcript levels of SbC4H and Sb4CL compared with those in the control and also increased flavones contents by approximately 3- and 2.5-fold, respectively. We successfully engineered the flavone biosynthesis pathway for the production of beneficial flavones in S baicalensis hairy roots. The importance of upstream gene C4H and 4CL in flavone biosynthesis and the efficiency of metabolic engineering in promoting flavone biosynthesis in S. baicalensis hairy roots have been indicated in this study.

  15. Chemical Genetics Uncovers Novel Inhibitors of Lignification, Including p-Iodobenzoic Acid Targeting CINNAMATE-4-HYDROXYLASE1[OPEN

    PubMed Central

    Van de Wouwer, Dorien; Decou, Raphaël; Audenaert, Dominique; Nguyen, Long

    2016-01-01

    Plant secondary-thickened cell walls are characterized by the presence of lignin, a recalcitrant and hydrophobic polymer that provides mechanical strength and ensures long-distance water transport. Exactly the recalcitrance and hydrophobicity of lignin put a burden on the industrial processing efficiency of lignocellulosic biomass. Both forward and reverse genetic strategies have been used intensively to unravel the molecular mechanism of lignin deposition. As an alternative strategy, we introduce here a forward chemical genetic approach to find candidate inhibitors of lignification. A high-throughput assay to assess lignification in Arabidopsis (Arabidopsis thaliana) seedlings was developed and used to screen a 10-k library of structurally diverse, synthetic molecules. Of the 73 compounds that reduced lignin deposition, 39 that had a major impact were retained and classified into five clusters based on the shift they induced in the phenolic profile of Arabidopsis seedlings. One representative compound of each cluster was selected for further lignin-specific assays, leading to the identification of an aromatic compound that is processed in the plant into two fragments, both having inhibitory activity against lignification. One fragment, p-iodobenzoic acid, was further characterized as a new inhibitor of CINNAMATE 4-HYDROXYLASE, a key enzyme of the phenylpropanoid pathway synthesizing the building blocks of the lignin polymer. As such, we provide proof of concept of this chemical biology approach to screen for inhibitors of lignification and present a broad array of putative inhibitors of lignin deposition for further characterization. PMID:27485881

  16. Assay of prolyl 4-hydroxylase by the chromatographic determination of [14C]succinic acid on ion-exchange minicolumns.

    PubMed Central

    Cunliffe, C J; Franklin, T J; Gaskell, R M

    1986-01-01

    An assay for prolyl 4-hydroxylase (EC 1.14.11.2) is described which measures succinic acid produced during the decarboxylation of 2-oxoglutaric acid in the presence of poly(L-Pro-Gly-L-Pro). [1-14C]Succinic acid was separated from its precursor 2-oxo[5-14C]glutaric acid by using ion-exchange minicolumns. The contamination of succinic acid by 2-oxoglutaric acid was approx. 1%, and the recovery of succinic acid was 100%. Kinetic parameters of prolyl 4-hydroxylase measured by the assay showed good agreement with published values. Our experience indicates that the measurement of prolyl 4-hydroxylase by the production of succinic acid is especially suited to investigations involving large numbers of assays. PMID:3028379

  17. [Characterization of recombinant L-isoleucine-4-hydroxylase from Bacillus thuringiensis and its application in 4hydroxyisoleucine biosynthesis].

    PubMed

    Zhang, Chenglin; Liu, Yuan; Xue, Ning; Wang, Xinxin; Xie, Xixian; Xu, Qingyang; Chen, Ning

    2014-08-04

    L-isoleucine-4-hydroxylase (IDO) encoding gene ido from Bacillus thuringiensis TCCC 11826 was cloned and expressed, followed by enzyme characterization. In addition, recombinant strain was tested for its 4-Hydroxyisoleucine (4-HIL) biotransformation. Ido gene was amplified from B. thuringiensis TCCC 11826 genomic DNA and expressed in BL-IDO. Recombinant IDO was extracted, purified and characterized. Recombinant strain used for biotransformation of 4-HIL was constructed. Composed of 723 nucleotides encoding 240 amino acids (sharing 97.47% and 97.91% identities with that of B. thuringiensis 2-e-2), ido gene was cloned from B. thuringiensis TCCC 11826. The recombinant IDO contained a His1-X-Asp/Glu-Xn-His2 motif that is specific for Fe(II)/alpha-ketoglutarate-dependent hydroxylases and catalyzed L-isoleucine to 4-HIL. Normal hyperbolic kinetics was observed with L-Ile in the reaction by recombinant IDO. Lineweaver-Burk treatment of the data yielded apparent Km and the Vmax was 0.18 mmol/L and 2.10 micromol/min/mg, respectively. The optimum temperature and pH for the recombinant IDO was 35 degrees C and 7.0 respectively; moreover, the relative activity of the enzyme remain 85.10% after 5 h incubation at 35 degrees C. In all, recombinant strain harboring ido transformed 89.28% of L-isoleucine to 4-HIL. In this study, an ido (Accession No. KC884243) with novel sequence was isolated and enzymatic characteristics of recombinant IDO was systematically analyzed. In addition, we successfully achieved the biotransformation of 4-HIL from L-isoleucine. This work will lay theoretical foundation and practical basis on the microbial manufacture technology of 4-HIL and other amino acid derivatives. This work will lay theoretical foundation and practical basis on the microbial manufacture technology of 4-HIL and other amino acid derivatives.

  18. Molecular Cloning, Characterization, and Expression Analysis of a Prolyl 4-Hydroxylase from the Marine Sponge Chondrosia reniformis.

    PubMed

    Pozzolini, Marina; Scarfì, Sonia; Mussino, Francesca; Ferrando, Sara; Gallus, Lorenzo; Giovine, Marco

    2015-08-01

    Prolyl 4-hydroxylase (P4H) catalyzes the hydroxylation of proline residues in collagen. P4H has two functional subunits, α and β. Here, we report the cDNA cloning, characterization, and expression analysis of the α and β subunits of the P4H derived from the marine sponge Chondrosia reniformis. The amino acid sequence of the α subunit is 533 residues long with an M r of 59.14 kDa, while the β subunit counts 526 residues with an M r of 58.75 kDa. Phylogenetic analyses showed that αP4H and βP4H are more related to the mammalian sequences than to known invertebrate P4Hs. Western blot analysis of sponge lysate protein cross-linking revealed a band of 240 kDa corresponding to an α2β2 tetramer structure. This result suggests that P4H from marine sponges shares the same quaternary structure with vertebrate homologous enzymes. Gene expression analyses showed that αP4H transcript is higher in the choanosome than in the ectosome, while the study of factors affecting its expression in sponge fragmorphs revealed that soluble silicates had no effect on the αP4H levels, whereas ascorbic acid strongly upregulated the αP4H mRNA. Finally, treatment with two different tumor necrosis factor (TNF)-alpha inhibitors determined a significant downregulation of αP4H gene expression in fragmorphs demonstrating, for the first time in Porifera, a positive involvement of TNF in sponge matrix biosynthesis. The molecular characterization of P4H genes involved in collagen hydroxylation, including the mechanisms that regulate their expression, is a key step for future recombinant sponge collagen production and may be pivotal to understand pathological mechanisms related to extracellular matrix deposition in higher organisms.

  19. Acetanilide 4-hydroxylase and acetanilide 2-hydroxylase activity in hepatic microsomes from induced mice.

    PubMed

    Lewandowski, M; Chui, Y C; Levi, P; Hodgson, E

    1991-02-01

    A simple and sensitive method for the separation of 14C-labelled acetanilide, 4-hydroxyacetanilide, 3-hydroxyacetanilide and 2-hydroxyacetanilide was developed using thin-layer chromatography. This separation is the basis for the assay of acetanilide 4-hydroxylase and acetanilide 2-hydroxylase activity in liver microsomes from DBA2/N male mice that had been treated with phenobarbital, 3-methylcholanthrene, isosafrole or n-butylbenzodioxole. Microsomes were incubated with [14C]acetanilide and extracted with benzene and ethyl acetate. The extract was applied to silica gel plates and developed with a hexane/isopropanol/ammonium hydroxide/water solvent system. The radiolabelled phenolic metabolites and the parent compound were detected using a Berthold Automatic TLC Linear Analyzer. Although the 4-hydroxylated metabolite was the primary product detected, this method can be used to detect other phenolic metabolites.

  20. Prolyl-4-Hydroxylases Inhibitor Stabilizes HIF-1α and Increases Mitophagy to Reduce Cell Death After Experimental Retinal Detachment.

    PubMed

    Liu, Haiyang; Zhu, Hong; Li, Tong; Zhang, Pengfei; Wang, Ning; Sun, Xiaodong

    2016-04-01

    This study investigated the neuroprotective effect against photoreceptor cell death using prolyl-4-hydroxylases inhibitor (PHI), an HIF-1α stabilizer, in experimental retinal detachment (RD). RD was created in Brown Norway rats by subretinal injection of 1% sodium hyaluronate. FG-4592 (a PHI, 25 mg/kg) or a vehicle was administered every 2 days with retro-orbital injection. Photoreceptor death was evaluated by TdT-dUTP terminal nick-end labeling (TUNEL) assay 3 days after RD and by the thickness of the outer nuclear layer 7 days after RD. The mitophagy-related markers Hypoxia Inducible Factor 1α (HIF-1α), BCL2/adenovirus E1B 19 kDa protein-interacting protein 3 (BNIP3), autophagy-related gene 5 (Atg5), microtubule-associated protein 1 light chain 3 beta (LC3B), and FUN14 domain containing 1 (FUNDC1) were detected by Western blot and immunofluorescence. Transmission electron microscopy was used to observe ultramicro-morphological changes. Mitochondrial damage was evaluated by the measurement of reactive oxygen species (ROS) by in situ ROS detection with dihydroethidium. The accumulation of HIF-1α and BNIP3 significantly increased after PHI treatment (P < 0.05), the pattern of Atg5 and LC3 changed, and FUNDC1 and LC3 were colocated. More autophagic vacuoles engulfing mitochondria were observed in transmission electron microscopy sections after PHI treatment when compared with the control. ROS significantly decreased in the PHI-treatment group (P < 0.05). This resulted in reduced TUNEL-positive photoreceptors 3 days after RD and an increased thickness of the outer nuclear layer 7 days after RD (P < 0.05). HIF-1α stabilization as a result of PHI treatment, along with the enhancement of mitophagy, could provide protection against photoreceptor injury following RD, which might be mediated by excessive ROS generation.

  1. Biphenyl 4-Hydroxylases Involved in Aucuparin Biosynthesis in Rowan and Apple Are Cytochrome P450 736A Proteins1[OPEN

    PubMed Central

    Kaufholdt, David; Broggini, Giovanni A.L.; Flachowsky, Henryk; Hänsch, Robert

    2015-01-01

    Upon pathogen attack, fruit trees such as apple (Malus spp.) and pear (Pyrus spp.) accumulate biphenyl and dibenzofuran phytoalexins, with aucuparin as a major biphenyl compound. 4-Hydroxylation of the biphenyl scaffold, formed by biphenyl synthase (BIS), is catalyzed by a cytochrome P450 (CYP). The biphenyl 4-hydroxylase (B4H) coding sequence of rowan (Sorbus aucuparia) was isolated and functionally expressed in yeast (Saccharomyces cerevisiae). SaB4H was named CYP736A107. No catalytic function of CYP736 was known previously. SaB4H exhibited absolute specificity for 3-hydroxy-5-methoxybiphenyl. In rowan cell cultures treated with elicitor from the scab fungus, transient increases in the SaB4H, SaBIS, and phenylalanine ammonia lyase transcript levels preceded phytoalexin accumulation. Transient expression of a carboxyl-terminal reporter gene construct directed SaB4H to the endoplasmic reticulum. A construct lacking the amino-terminal leader and transmembrane domain caused cytoplasmic localization. Functional B4H coding sequences were also isolated from two apple (Malus × domestica) cultivars. The MdB4Hs were named CYP736A163. When stems of cv Golden Delicious were infected with the fire blight bacterium, highest MdB4H transcript levels were observed in the transition zone. In a phylogenetic tree, the three B4Hs were closest to coniferaldehyde 5-hydroxylases involved in lignin biosynthesis, suggesting a common ancestor. Coniferaldehyde and related compounds were not converted by SaB4H. PMID:25862456

  2. Detection of four oxidation sites in viral prolyl-4-hydroxylase by top-down mass spectrometry

    PubMed Central

    Ge, Ying; Lawhorn, Brian G.; ElNaggar, Mariam; Sze, Siu Kwan; Begley, Tadhg P.; McLafferty, Fred W.

    2003-01-01

    Oxidative inactivation is a common problem for enzymatic reactions that proceed via iron oxo intermediates. In an investigation of the inactivation of a viral prolyl-4-hydroxylase (26 kD), electrospray mass spectrometry (MS) directly shows the degree of oxidation under varying experimental conditions, but indicates the addition at most of three oxygen atoms per molecule. Thus, molecular ion masses (M + nO) of one sample indicate the oxygen atom adducts n = 0, 1, 2, 3, and 4 of 35, 41, 19, 5 ± 3, and <2%, respectively; “top-down” MS/MS of these ions show oxidation at the sites R28–V31, E95–F107, and K216 of 22%, 28%, and 34%, respectively, but with a possible (~4%) fourth site at V125–D150. However, for the doubly oxidized molecular ions (increasing the precursor oxygen content from 0.94 to 2), MS/MS showed an easily observable ~13% oxygen at the V125–D150 site. For the “bottom-up” approach, detection of the ~4% oxidation at the V125–D150 site by MS analysis of a proteolysis mixture would have been very difficult. The unmodified peptide containing this site would represent a few percent of the proteolysis mixture; the oxidized peptide not only would be just ~4% of this, but the uniqueness of its mass value (~1–2 kD) would be far less than the 11,933 Dalton value used here. Using different molecular ion precursors for top-down MS/MS also provides kinetic data from a single sample, that is, from molecular ions with 0.94 and 2 oxygens. Little oxidation occurs at V125–D150 until K216 is oxidized, suggesting that these are competitively catalyzed by the iron center; among several prolyl-4-hydroxylases the K216, H137, and D139 are conserved residues. PMID:14500890

  3. Engineering bacterial phenylalanine 4-hydroxylase for microbial synthesis of human neurotransmitter precursor 5-hydroxytryptophan.

    PubMed

    Lin, Yuheng; Sun, Xinxiao; Yuan, Qipeng; Yan, Yajun

    2014-07-18

    5-Hydroxytryptophan (5-HTP) is a drug that is clinically effective against depression, insomnia, obesity, chronic headaches, etc. It is only commercially produced by the extraction from the seeds of Griffonia simplicifolia because of a lack of synthetic methods. Here, we report the efficient microbial production of 5-HTP via combinatorial protein and metabolic engineering approaches. First, we reconstituted and screened prokaryotic phenylalanine 4-hydroxylase activity in Escherichia coli. Then, sequence- and structure-based protein engineering dramatically shifted its substrate preference, allowing for efficient conversion of tryptophan to 5-HTP. Importantly, E. coli endogenous tetrahydromonapterin (MH4) could be utilized as the coenzyme, when a foreign MH4 recycling mechanism was introduced. Whole-cell bioconversion allowed the high-level production of 5-HTP (1.1-1.2 g/L) from tryptophan in shake flasks. On this basis, metabolic engineering efforts were further made to achieve the de novo 5-HTP biosynthesis from glucose. This work not only holds great scale-up potential but also demonstrates a strategy for expanding the native metabolism of microorganisms.

  4. Immunolocalization of phenylalanine ammonia-lyase and cinnamate-4-hydroxylase in differentiating xylem of poplar.

    PubMed

    Sato, Takahiko; Takabe, Keiji; Fujita, Minoru

    2004-01-01

    Phenylalanine ammonia-lyase (PAL; EC 4.3.1.5) and cinnamate-4-hydroxylase (C4H; EC 1.14.13.11) are pivotal enzymes involved in lignification. We synthesized peptides as the epitopes according to the amino acid sequences of these enzymes, coupled them with hemocyanin, and injected them into mice. The antiserums against peptides of PAL and C4H specifically detected PAL and C4H in the crude enzymes extracted from differentiating xylem of poplar, respectively. PAL and C4H were localized in differentiating xylem of poplar. PAL labeling was mainly localized in the cytosol, and somewhat localized on the rough-endoplasmic reticulum (r-ER) and the Golgi apparatus. In contrast, C4H was mainly observed on r-ER and the Golgi apparatus. These findings suggest that conversion of phenylalanine to cinnamic acid occurs in the cytosol and the following reaction occurs near the membrane of r-ER and the Golgi apparatus. The possibility of coordinated localization of PAL and C4H is discussed.

  5. The effects of Urtica dioica L. leaf extract on aniline 4-hydroxylase in mice.

    PubMed

    Ozen, Tevfik; Korkmaz, Halil

    2009-01-01

    The effects of hydroalcoholic (80% ethanol-20% water) extract of Urtica dioica L. on microsomal aniline 4-hydroxylase (A4H) were investigated in the liver of Swiss albino mice (8- 10-weeks-old) treated with two doses (50 and 100 mg/kg body weight, given orally for 14 days ). The activities of A4H showed a significant increase in the liver at both dose levels of extract treatment. The hydroalcoholic extract of Urtica dioica induced the activities of A4H that had been increased by treatment of metal ions (Mg2+ and Ca2+) and the mixture of cofactors (NADH and NADPH). At saturated concentration of cofactor, microsomal A4H exhibited significantly even higher activities in the presence of the mixture of cofactors than NADPH and NADH. Mg2+ and Ca2+ ions acted as stimulants in vitro. The present results suggest that the hydroalcoholic extract of Urtica dioica may have modalatory effect on aniline hydroxylase at least in part and enhance the activity of A4H adding metals ions and cofactors.

  6. Bacillus anthracis Prolyl 4-Hydroxylase Modifies Collagen-like Substrates in Asymmetric Patterns*

    PubMed Central

    Schnicker, Nicholas J.; Dey, Mishtu

    2016-01-01

    Proline hydroxylation is the most prevalent post-translational modification in collagen. The resulting product trans-4-hydroxyproline (Hyp) is of critical importance for the stability and thus function of collagen, with defects leading to several diseases. Prolyl 4-hydroxylases (P4Hs) are mononuclear non-heme iron α-ketoglutarate (αKG)-dependent dioxygenases that catalyze Hyp formation. Although animal and plant P4Hs target peptidyl proline, prokaryotes have been known to use free l-proline as a precursor to form Hyp. The P4H from Bacillus anthracis (BaP4H) has been postulated to act on peptidyl proline in collagen peptides, making it unusual within the bacterial clade, but its true physiological substrate remains enigmatic. Here we use mass spectrometry, fluorescence binding, x-ray crystallography, and docking experiments to confirm that BaP4H recognizes and acts on peptidyl substrates but not free l-proline, using elements characteristic of an Fe(II)/αKG-dependent dioxygenases. We further show that BaP4H can hydroxylate unique peptidyl proline sites in collagen-derived peptides with asymmetric hydroxylation patterns. The cofactor-bound crystal structures of BaP4H reveal active site conformational changes that define open and closed forms and mimic “ready” and “product-released” states of the enzyme in the catalytic cycle. These results help to clarify the role of BaP4H as well as provide broader insights into human collagen P4H and proteins with poly-l-proline type II helices. PMID:27129244

  7. Bacillus anthracis Prolyl 4-Hydroxylase Modifies Collagen-like Substrates in Asymmetric Patterns.

    PubMed

    Schnicker, Nicholas J; Dey, Mishtu

    2016-06-17

    Proline hydroxylation is the most prevalent post-translational modification in collagen. The resulting product trans-4-hydroxyproline (Hyp) is of critical importance for the stability and thus function of collagen, with defects leading to several diseases. Prolyl 4-hydroxylases (P4Hs) are mononuclear non-heme iron α-ketoglutarate (αKG)-dependent dioxygenases that catalyze Hyp formation. Although animal and plant P4Hs target peptidyl proline, prokaryotes have been known to use free l-proline as a precursor to form Hyp. The P4H from Bacillus anthracis (BaP4H) has been postulated to act on peptidyl proline in collagen peptides, making it unusual within the bacterial clade, but its true physiological substrate remains enigmatic. Here we use mass spectrometry, fluorescence binding, x-ray crystallography, and docking experiments to confirm that BaP4H recognizes and acts on peptidyl substrates but not free l-proline, using elements characteristic of an Fe(II)/αKG-dependent dioxygenases. We further show that BaP4H can hydroxylate unique peptidyl proline sites in collagen-derived peptides with asymmetric hydroxylation patterns. The cofactor-bound crystal structures of BaP4H reveal active site conformational changes that define open and closed forms and mimic "ready" and "product-released" states of the enzyme in the catalytic cycle. These results help to clarify the role of BaP4H as well as provide broader insights into human collagen P4H and proteins with poly-l-proline type II helices. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  8. Prolyl-4-Hydroxylase 3 (PHD3) Expression Is Downregulated during Epithelial-to-Mesenchymal Transition

    PubMed Central

    Place, Trenton L.; Nauseef, Jones T.; Peterson, Maina K.; Henry, Michael D.; Mezhir, James J.; Domann, Frederick E.

    2013-01-01

    Prolyl-4-hydroxylation by the intracellular prolyl-4-hydroxylase enzymes (PHD1-3) serves as a master regulator of environmental oxygen sensing. The activity of these enzymes is tightly tied to tumorigenesis, as they regulate cell metabolism and angiogenesis through their control of hypoxia-inducible factor (HIF) stability. PHD3 specifically, is gaining attention for its broad function and rapidly accumulating array of non-HIF target proteins. Data from several recent studies suggest a role for PHD3 in the regulation of cell morphology and cell migration. In this study, we aimed to investigate this role by closely examining the relationship between PHD3 expression and epithelial-to-mesenchymal transition (EMT); a transcriptional program that plays a major role in controlling cell morphology and migratory capacity. Using human pancreatic ductal adenocarcinoma (PDA) cell lines and Madin-Darby Canine Kidney (MDCK) cells, we examined the correlation between several markers of EMT and PHD3 expression. We demonstrated that loss of PHD3 expression in PDA cell lines is highly correlated with a mesenchymal-like morphology and an increase in cell migratory capacity. We also found that induction of EMT in MDCK cells resulted in the specific downregulation of PHD3, whereas the expression of the other HIF-PHD enzymes was not affected. The results of this study clearly support a model by which the basal expression and hypoxic induction of PHD3 is suppressed by the EMT transcriptional program. This may be a novel mechanism by which migratory or metastasizing cells alter signaling through specific pathways that are sensitive to regulation by O2. The identification of downstream pathways that are affected by the suppression of PHD3 expression during EMT may provide important insight into the crosstalk between O2 and the migratory and metastatic potential of tumor cells. PMID:24367580

  9. Regulation of the Cinnamate 4-Hydroxylase (CYP73A1) in Jerusalem Artichoke Tubers in Response to Wounding and Chemical Treatments.

    PubMed Central

    Batard, Y.; Schalk, M.; Pierrel, M. A.; Zimmerlin, A.; Durst, F.; Werck-Reichhart, D.

    1997-01-01

    trans-Cinnamate 4-hydroxylase (C4H) is a plant-specific cytochrome (P450) that is encoded by the gene CYP73A and catalyzes the second step of the multibranched phenylpropanoid pathway. Increases in C4H activity in response to physical and chemical stresses have been well documented, but the mechanism of these increases has never been studied in detail. This paper reports on the regulatory mechanism controlling C4H activity in Jerusalem artichoke (Helianthus tuberosus) tubers in response to wounding and chemical treatments. We compared induction of C4H and other P450-catalyzed activities. C4H was moderately induced by chemicals relative to other P450s. Increases in enzyme activity, C4H protein, and transcripts were quantified and compared in tuber tissue 48 h after wounding and chemical treatments. Our data suggest that induction of the enzyme activity results primarily from gene activation. Time-course experiments were performed after wounding and aminopyrine treatment. Compared with wounded tissues, aminopyrine triggered an additional and delayed peak of transcript accumulation. The timing of the induced changes in activity, protein, and transcripts confirms that C4H induction results primarily from an increase in CYP73A1 mRNA, in both wounded and aminopyrine-treated tissues. However, posttranscriptional mechanisms might also contribute to the regulation of C4H activity. PMID:12223655

  10. Antisense and sense expression of cDNA coding for CYP73A15, a class II cinnamate 4-hydroxylase, leads to a delayed and reduced production of lignin in tobacco

    NASA Technical Reports Server (NTRS)

    Blee, K.; Choi, J. W.; O'Connell, A. P.; Jupe, S. C.; Schuch, W.; Lewis, N. G.; Bolwell, G. P.

    2001-01-01

    A number of plant species contain the class II of genes encoding the cytochrome P450, CYP73, the cognate protein of which cinnamic acid 4-hydroxylase, is the second enzyme of the phenylpropanoid pathway. In order to begin to determine possible functionality, tobacco has been transformed with a truncated French bean class II cinnamate hydroxylase (CYP73A15) in the sense and antisense orientations. Signals for C4H protein could be detected in vascular tissue from wild-type plants using heterologous probes. The transformed plants showed a normal phenotype, even though detectable C4H protein was much reduced in tissue prints. Young propagated transformants displayed a range of reduced C4H activities, as well as either reduced or no phloroglucinol-stainable lignin. However, all mature tobacco plants showed the accumulation of lignin, even though its deposition was apparently delayed. This was not due to induction of tyrosine ammonia-lyase activity, which was not detected, but instead it is presumed due to sufficient C4H residual activity. Analysis of the lignin content of the plants showed reductions of up to 30% with a slightly reduced syringyl to guaiacyl ratio as compared to wild type. This reduction level was favourable in comparison with some other targets in the lignification pathway that have been manipulated including that of class I cinnamate 4-hydroxylase. It is proposed that the class II cinnamate 4-hydroxylase might also function in lignification in a number of species including French bean and tobacco, based on these data.

  11. Antisense and sense expression of cDNA coding for CYP73A15, a class II cinnamate 4-hydroxylase, leads to a delayed and reduced production of lignin in tobacco

    NASA Technical Reports Server (NTRS)

    Blee, K.; Choi, J. W.; O'Connell, A. P.; Jupe, S. C.; Schuch, W.; Lewis, N. G.; Bolwell, G. P.

    2001-01-01

    A number of plant species contain the class II of genes encoding the cytochrome P450, CYP73, the cognate protein of which cinnamic acid 4-hydroxylase, is the second enzyme of the phenylpropanoid pathway. In order to begin to determine possible functionality, tobacco has been transformed with a truncated French bean class II cinnamate hydroxylase (CYP73A15) in the sense and antisense orientations. Signals for C4H protein could be detected in vascular tissue from wild-type plants using heterologous probes. The transformed plants showed a normal phenotype, even though detectable C4H protein was much reduced in tissue prints. Young propagated transformants displayed a range of reduced C4H activities, as well as either reduced or no phloroglucinol-stainable lignin. However, all mature tobacco plants showed the accumulation of lignin, even though its deposition was apparently delayed. This was not due to induction of tyrosine ammonia-lyase activity, which was not detected, but instead it is presumed due to sufficient C4H residual activity. Analysis of the lignin content of the plants showed reductions of up to 30% with a slightly reduced syringyl to guaiacyl ratio as compared to wild type. This reduction level was favourable in comparison with some other targets in the lignification pathway that have been manipulated including that of class I cinnamate 4-hydroxylase. It is proposed that the class II cinnamate 4-hydroxylase might also function in lignification in a number of species including French bean and tobacco, based on these data.

  12. Enzymatic changes in phenylalanine ammonia-lyase, cinnamic-4-hydroxylase, capsaicin synthase, and peroxidase activities in capsicum under drought stress.

    PubMed

    Phimchan, Paongpetch; Chanthai, Saksit; Bosland, Paul W; Techawongstien, Suchila

    2014-07-23

    Penylalanine ammonia-lyase (PAL), cinnamic-4-hydroxylase (C4H), capsaicin synthase (CS), and peroxidase (POD) are involved in the capsaicinoid biosynthesis pathway and may be altered in cultivars with different pungency levels. This study clarified the action of these enzymes under drought stress for hot Capsicum cultivars with low, medium,and high pungency levels. At the flowering stage, control plants were watered at field capacity, whereas drought-induced plants were subjected to gradual drought stress. Under drought stress, PAL, C4H, CS, and POD enzyme activities increased as compared to the non-drought-stressed plants. A novel discovery was that PAL was the critical enzyme in capsaicinoid biosynthesis under drought stress because its activities and capsaicinoid increased across the different pungency levels of hot pepper cultivars examined.

  13. Expression, Purification, Crystallization And Preliminary X-Ray Studies of a Prolyl-4-Hydroxylase Protein From Bacillus Anthracis

    SciTech Connect

    Miller, M.A.; Scott, E.E.; Limburg, J.

    2009-05-26

    Collagen prolyl-4-hydroxylase (C-P4H) catalyzes the hydroxylation of specific proline residues in procollagen, which is an essential step in collagen biosynthesis. A new form of P4H from Bacillus anthracis (anthrax-P4H) that shares many characteristics with the type I C-P4H from human has recently been characterized. The structure of anthrax-P4H could provide important insight into the chemistry of C-P4Hs and into the function of this unique homodimeric P4H. X-ray diffraction data of selenomethionine-labeled anthrax-P4H recombinantly expressed in Escherichia coli have been collected to 1.4 {angstrom} resolution.

  14. Isolation and characterization of isochorismate synthase and cinnamate 4-hydroxylase during salinity stress, wounding, and salicylic acid treatment in Carthamus tinctorius.

    PubMed

    Sadeghi, Mahnaz; Dehghan, Sara; Fischer, Rainer; Wenzel, Uwe; Vilcinskas, Andreas; Kavousi, Hamid Reza; Rahnamaeian, Mohammad

    2013-11-01

    Salicylic acid (SA) is a prominent signaling molecule during biotic and abiotic stresses in plants biosynthesized via cinnamate and isochorismate pathways. Cinnamate 4-hydroxylase (C4H) and isochorismate synthase (ICS) are the main enzymes in phenylpropanoid and isochorismate pathways, respectively. To investigate the actual roles of these genes in resistance mechanism to environmental stresses, here, the coding sequences of these enzymes in safflower (Carthamus tinctorius), as an oilseed industrial medicinal plant, were partially isolated and their expression profiles during salinity stress, wounding, and salicylic acid treatment were monitored. As a result, safflower ICS (CtICS) and C4H (CtC4H) were induced in early time points after wounding (3-6 h). Upon salinity stress, CtICS and CtC4H were highly expressed for the periods of 6-24 h and 3-6 h after treatment, respectively. It seems evident that ICS expression level is SA concentration dependent as if safflower treatment with 1 mM SA could induce ICS much stronger than that with 0.1 mM, while C4H is less likely to be so. Based on phylogenetic analysis, safflower ICS has maximum similarity to its ortholog in Vitis vinifera up to 69%, while C4H shows the highest similarity to its ortholog in Echinacea angustifolia up to 96%. Overall, the isolated genes of CtICS and CtC4H in safflower could be considered in plant breeding programs for salinity tolerance as well as for pathogen resistance.

  15. Isolation and characterization of isochorismate synthase and cinnamate 4-hydroxylase during salinity stress, wounding, and salicylic acid treatment in Carthamus tinctorius

    PubMed Central

    Sadeghi, Mahnaz; Dehghan, Sara; Fischer, Rainer; Wenzel, Uwe; Vilcinskas, Andreas; Kavousi, Hamid Reza; Rahnamaeian, Mohammad

    2013-01-01

    Salicylic acid (SA) is a prominent signaling molecule during biotic and abiotic stresses in plants biosynthesized via cinnamate and isochorismate pathways. Cinnamate 4-hydroxylase (C4H) and isochorismate synthase (ICS) are the main enzymes in phenylpropanoid and isochorismate pathways, respectively. To investigate the actual roles of these genes in resistance mechanism to environmental stresses, here, the coding sequences of these enzymes in safflower (Carthamus tinctorius), as an oilseed industrial medicinal plant, were partially isolated and their expression profiles during salinity stress, wounding, and salicylic acid treatment were monitored. As a result, safflower ICS (CtICS) and C4H (CtC4H) were induced in early time points after wounding (3–6 h). Upon salinity stress, CtICS and CtC4H were highly expressed for the periods of 6–24 h and 3–6 h after treatment, respectively. It seems evident that ICS expression level is SA concentration dependent as if safflower treatment with 1 mM SA could induce ICS much stronger than that with 0.1 mM, while C4H is less likely to be so. Based on phylogenetic analysis, safflower ICS has maximum similarity to its ortholog in Vitis vinifera up to 69%, while C4H shows the highest similarity to its ortholog in Echinacea angustifolia up to 96%. Overall, the isolated genes of CtICS and CtC4H in safflower could be considered in plant breeding programs for salinity tolerance as well as for pathogen resistance. PMID:24309561

  16. Cinnamate-4-hydroxylase expression in arabidopsis. Regulation in response to development and the environment

    SciTech Connect

    Bell-Lelong, D.A.; Cusumano, J.C.; Meyer, K.; Chapple, C.

    1997-03-01

    Cinnamate-r-hydroxylase (C4H) is the first Cyt P450-dependent monooxygenase of the phenylpropanoid pathway. To study the expression of this gene in Arabidopsis thaliana, a C4H cDNA clone from the Arabidopsis expressed sequence tag database was identified and used to isolate its corresponding genomic clone. The entire C4H coding sequence plus 2.9 kb of its promoter were isolated on a 5.4-kb HindIII fragment of this cosmid. Inspection of the promoter sequence revealed the presence of a number of putative regulatory motifs previously identified in the promoters of other phenylpropanoid pathway genes. The expression of C4H was analyzed by RNA blot hybridization analysis and in transgenic Arabidopsis carrying a C4H-{beta}-glucuronidase transcriptional fusion. C4H message accumulation was light-dependent, but was detectable even in dark-grown seedlings. Consistent with these data, C4H mRNA was accumulated to light-grown levels in etiolated det1-1 mutant seedlings. C4H is widely expressed in various Arabidopsis tissues, particularly in roots and cells undergoing lignification. The C4H-driven {beta}-glucuronidase expression accurately reflected the tissue-specificity and wound-inducibility of the C4H promoter indicated by RNA blot hybridization analysis. A modest increase in C4H expression was observed in the tt8 mutant of Arabidopsis. 77 refs., 5 figs.

  17. Comparison of aryl hydrocarbon hydroxylase and acetanilide 4-hydroxylase induction by polycyclic aromatic compounds in human and mouse cell lines.

    PubMed

    Jaiswal, A K; Nebert, D W; Eisen, H W

    1985-08-01

    The human MCF-7 and the mouse Hepa-1 cell culture lines were compared for aryl hydrocarbon hydroxylase and acetanilide 4-hydroxylase inducibility by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and benzo[a]anthracene (BA) and TCDD- and BA-specific binding in the cytosol and nucleus. The effective concentration of BA in the growth medium required to induce either enzyme to 50% of its maximally inducible activity (EC50) was the same (5-11 microM) in both MCF-7 and Hepa-1 cells. On the other hand, the EC50 for TCDD in MCF-7 cells (5-25 nM) was more than 40-fold greater than that in Hepa-1 cells (0.4 to 0.6 nM). P1-450- and P3-450-specific mouse cDNA probes were used to quantitate mRNA induction in the Hepa-1 cell line. P1-450 mRNA was induced markedly by TCDD and benzo[a] anthracene, whereas P3-450 mRNA was induced negligibly. A P1-450-specific human cDNA probe was used to quantitate P1-450 mRNA induction in the MCF-7 cell line. Aryl hydrocarbon hydroxylase inducibility by TCDD or BA always paralleled P1-450 mRNA inducibility in either the mouse or human line. Although the cytosolic Ah receptor in Hepa-1 cells was easily detected by sucrose density gradient centrifugation, gel permeation chromatography, and anion-exchange high-performance liquid chromatography, the cytosolic receptor cannot be detected in MCF-7 cells. Following in vivo exposure of cultures to radiolabeled TCDD, the intranuclear concentration of inducer-receptor complex was at least fifty times greater in Hepa-1 than MCF-7 cultures. The complete lack of measurable cytosolic receptor and almost totally absent inducer-receptor complex in the nucleus of MCF-7 cells was, therefore, out of proportion to its capacity for aryl hydrocarbon hydroxylase and acetanilide 4-hydroxylase inducibility. This MCF-7 line should provide an interesting model for a better understanding of the mechanisms of drug-metabolizing enzyme induction by polycyclic aromatic compounds, including the Ah receptor-mediated mechanism.

  18. Characterization of cDNAs, mRNAs, and proteins related to human liver microsomal cytochrome P-450 (S)-mephenytoin 4'-hydroxylase.

    PubMed

    Ged, C; Umbenhauer, D R; Bellew, T M; Bork, R W; Srivastava, P K; Shinriki, N; Lloyd, R S; Guengerich, F P

    1988-09-06

    mephenytoin 4'-hydroxylase activity. These results indicate that several closely related P-450 genes are all expressed in individual human livers. The MP-4/MP-8 gene products are proposed to be the ones most likely involved in mephenytoin 4'-hydroxylation, and much of the variation in catalytic activity among individuals is not a result of differences in levels of P-450MP-1 or mRNA but may be due to base differences in the structural gene(s).

  19. The Endothelial Prolyl-4-Hydroxylase Domain 2/Hypoxia-Inducible Factor 2 Axis Regulates Pulmonary Artery Pressure in Mice

    PubMed Central

    Rajendran, Ganeshkumar; Astleford, Lindsay; Michael, Mark; Schonfeld, Michael P.; Fields, Timothy; Shay, Sheila; French, Jaketa L.; West, James; Haase, Volker H.

    2016-01-01

    Hypoxia-inducible factors 1 and 2 (HIF-1 and -2) control oxygen supply to tissues by regulating erythropoiesis, angiogenesis and vascular homeostasis. HIFs are regulated in response to oxygen availability by prolyl-4-hydroxylase domain (PHD) proteins, with PHD2 being the main oxygen sensor that controls HIF activity under normoxia. In this study, we used a genetic approach to investigate the endothelial PHD2/HIF axis in the regulation of vascular function. We found that inactivation of Phd2 in endothelial cells specifically resulted in severe pulmonary hypertension (∼118% increase in right ventricular systolic pressure) but not polycythemia and was associated with abnormal muscularization of peripheral pulmonary arteries and right ventricular hypertrophy. Concurrent inactivation of either Hif1a or Hif2a in endothelial cell-specific Phd2 mutants demonstrated that the development of pulmonary hypertension was dependent on HIF-2α but not HIF-1α. Furthermore, endothelial HIF-2α was required for the development of increased pulmonary artery pressures in a model of pulmonary hypertension induced by chronic hypoxia. We propose that these HIF-2-dependent effects are partially due to increased expression of vasoconstrictor molecule endothelin 1 and a concomitant decrease in vasodilatory apelin receptor signaling. Taken together, our data identify endothelial HIF-2 as a key transcription factor in the pathogenesis of pulmonary hypertension. PMID:26976644

  20. Prolyl 4-Hydroxylase: Substrate Isosteres in Which an (E)- or (Z)-Alkene Replaces the Prolyl Peptide Bond.

    PubMed

    Vasta, James D; Choudhary, Amit; Jensen, Katrina H; McGrath, Nicholas A; Raines, Ronald T

    2017-01-10

    Collagen prolyl 4-hydroxylases (CP4Hs) catalyze a prevalent posttranslational modification, the hydroxylation of (2S)-proline residues in protocollagen strands. The ensuing (2S,4R)-4-hydroxyproline residues are necessary for the conformational stability of the collagen triple helix. Prolyl peptide bonds isomerize between cis and trans isomers, and the preference of the enzyme is unknown. We synthesized alkene isosteres of the cis and trans isomers to probe the conformational preferences of human CP4H1. We discovered that the presence of a prolyl peptide bond is necessary for catalysis. The cis isostere is, however, an inhibitor with a potency greater than that of the trans isostere, suggesting that the cis conformation of a prolyl peptide bond is recognized preferentially. Comparative studies with a Chlamydomonas reinhardtii P4H, which has a similar catalytic domain but lacks an N-terminal substrate-binding domain, showed a similar preference for the cis isostere. These findings support the hypothesis that the catalytic domain of CP4Hs recognizes the cis conformation of the prolyl peptide bond and inform the use of alkenes as isosteres for peptide bonds.

  1. Colocalization of L-phenylalanine ammonia-lyase and cinnamate 4-hydroxylase for metabolic channeling in phenylpropanoid biosynthesis.

    PubMed

    Achnine, Lahoucine; Blancaflor, Elison B; Rasmussen, Susanne; Dixon, Richard A

    2004-11-01

    Metabolic channeling has been proposed to occur at the entry point into plant phenylpropanoid biosynthesis. To determine whether isoforms of L-Phe ammonia-lyase (PAL), the first enzyme in the pathway, can associate with the next enzyme, the endomembrane-bound cinnamate 4-hydroxylase (C4H), to facilitate channeling, we generated transgenic tobacco (Nicotiana tabacum) plants independently expressing epitope-tagged versions of two PAL isoforms (PAL1 and PAL2) and C4H. Subcellular fractionation and protein gel blot analysis using epitope- and PAL isoform-specific antibodies indicated both microsomal and cytosolic locations of PAL1 but only cytosolic localization of PAL2. However, both PAL isoforms were microsomally localized in plants overexpressing C4H. These results, which suggest that C4H itself may organize the complex for membrane association of PAL, were confirmed using PAL-green fluorescent protein (GFP) fusions with localization by confocal microscopy. Coexpression of unlabeled PAL1 with PAL2-GFP resulted in a shift of fluorescence localization from endomembranes to cytosol in C4H overexpressing plants, whereas coexpression of unlabeled PAL2 with PAL1-GFP did not affect PAL1-GFP localization, indicating that PAL1 has a higher affinity for its membrane localization site than does PAL2. Dual-labeling immunofluorescence and fluorescence energy resonance transfer (FRET) studies confirmed colocalization of PAL and C4H. However, FRET analysis with acceptor photobleaching suggested that the colocalization was not tight.

  2. MiR-30e suppresses proliferation of hepatoma cells via targeting prolyl 4-hydroxylase subunit alpha-1 (P4HA1) mRNA

    SciTech Connect

    Feng, Guoxing; Shi, Hui; Li, Jiong; Yang, Zhe; Fang, Runping; Ye, Lihong; Zhang, Weiying; Zhang, Xiaodong

    2016-04-08

    Aberrant microRNA expression has been shown to be characteristic of many cancers. It has been reported that the expression levels of miR-30e are decreased in liver cancer tissues. However, the role of miR-30e in hepatocellular carcinoma remains poorly understood. In the present study, we investigated the significance of miR-30e in hepatocarcinogenesis. Bioinformatics analysis reveals a putative target site of miR-30e in the 3′-untranslated region (3′UTR) of prolyl 4-hydroxylase subunit alpha-1 (P4HA1) mRNA. Moreover, luciferase reporter gene assays verified that miR-30e directly targeted 3′UTR of P4HA1 mRNA. Then, we demonstrated that miR-30e was able to reduce the expression of P4HA1 at the levels of mRNA and protein using reverse transcription-polymerase chain reaction and Western blot analysis. Enforced expression of miR-30e suppressed proliferation of HepG2 cells by 5-ethynyl-2-deoxyuridine (EdU) assay and reduced colony formation of these cells by colony formation analysis. Conversely, anti-miR-30e enhanced the proliferation of hepatoma cells in vitro. Interestingly, the ectopic expression of P4HA1 could efficiently rescue the inhibition of cell proliferation mediated by miR-30e in HepG2 cells. Meanwhile, silencing of P4HA1 abolished the anti-miR-30e-induced proliferation of cells. Clinically, quantitative real-time PCR showed that miR-30e was down-regulated in liver tumor tissues relative to their peritumor tissues. The expression levels of miR-30e were negatively correlated to those of P4HA1 mRNA in clinical liver tumor tissues. Thus, we conclude that miR-30e suppresses proliferation of hepatoma cells through targeting P4HA1 mRNA. Our finding provides new insights into the mechanism of hepatocarcinogenesis. - Highlights: • P4HA1 is a novel target gene of miR-30e. • P4HA1 is increased in clinical HCC tissues. • MiR-30e is negatively correlated with P4HA1 in clinical HCC tissues. • MiR-30e suppresses the proliferation of HCC cells through

  3. TNF-α Suppresses Prolyl-4-Hydroxylase α1 Expression via the ASK1–JNK–NonO Pathway

    PubMed Central

    Zhang, Cheng; Zhang, Ming-Xiang; Shen, Ying H.; Burks, Jared K.; Zhang, Yun; Wang, Jian; LeMaire, Scott A.; Yoshimura, Koichi; Aoki, Hiroki; Coselli, Joseph S.; Wang, Xing Li

    2008-01-01

    Background Inflammation is known to contribute to the pathogenesis of vascular diseases in which arterial wall extracellular matrix (ECM) homeostasis is disrupted. Tumor necrosis factor-α (TNF-α), a pivotal cytokine that regulates ECM metabolism by increasing degradation and decreasing production of arterial collagens, is associated with vulnerable plaques and aortic aneurysms. Methods and Results In the current study, we showed that, when administered in doses of 1 to 100 ng/mL, TNF-α dose-dependently downregulated the expression of prolyl-4-hydroxylase αI [P4Hα (I)]—the rate-limiting subunit for the P4H enzyme essential for procollagen hydroxylation, secretion, and deposition in primary human aortic smooth muscle cells (HASMCs). Using a progressive deletion cloning approach, we characterized the TNF-α–responsive element (TaRE) in the human P4Hα (I) promoter and found that a negative regulatory region at the position of −32 to +18bp is responsible for ≈80% of TNF-α–mediated suppression. Using oligonucleotide-based transcription factor pull-down method in which proteins were resolved in 1-D gel electrophoresis and identified using LC-MS/MS, we identified the NonO protein binds this region. When NonO expression silenced with specific siRNA, we found that 70% of the TNF-α–mediated P4Hα suppression was abolished, which appeared to be mediated by the ASK1-JNK pathway. Conclusions Our findings define a novel molecular pathway for inflammation associated extracellular matrix dysregulation, which may account for atherosclerotic plaque rupture and aortic aneurysm formation. Further understanding of this pathway may facilitate development of novel therapeutics for vascular diseases. PMID:17478756

  4. Proline Availability Regulates Proline-4-Hydroxylase Synthesis and Substrate Uptake in Proline-Hydroxylating Recombinant Escherichia coli

    PubMed Central

    Falcioni, Francesco; Blank, Lars M.; Frick, Oliver; Karau, Andreas; Schmid, Andreas

    2013-01-01

    Microbial physiology plays a crucial role in whole-cell biotransformation, especially for redox reactions that depend on carbon and energy metabolism. In this study, regio- and enantio-selective proline hydroxylation with recombinant Escherichia coli expressing proline-4-hydroxylase (P4H) was investigated with respect to its interconnectivity to microbial physiology and metabolism. P4H production was found to depend on extracellular proline availability and on codon usage. Medium supplementation with proline did not alter p4h mRNA levels, indicating that P4H production depends on the availability of charged prolyl-tRNAs. Increasing the intracellular levels of soluble P4H did not result in an increase in resting cell activities above a certain threshold (depending on growth and assay temperature). Activities up to 5-fold higher were reached with permeabilized cells, confirming that host physiology and not the intracellular level of active P4H determines the achievable whole-cell proline hydroxylation activity. Metabolic flux analysis revealed that tricarboxylic acid cycle fluxes in growing biocatalytically active cells were significantly higher than proline hydroxylation rates. Remarkably, a catalysis-induced reduction of substrate uptake was observed, which correlated with reduced transcription of putA and putP, encoding proline dehydrogenase and the major proline transporter, respectively. These results provide evidence for a strong interference of catalytic activity with the regulation of proline uptake and metabolism. In terms of whole-cell biocatalyst efficiency, proline uptake and competition of P4H with proline catabolism are considered the most critical factors. PMID:23455348

  5. Monospecific polyclonal antibodies directed against purified cinnamate 4-hydroxylase from Helianthus tuberosus. Immunopurification, immunoquantitation, and interspecies cross-reactivity.

    PubMed Central

    Werck-Reichhart, D; Batard, Y; Kochs, G; Lesot, A; Durst, F

    1993-01-01

    We recently reported the purification of cinnamic acid 4-hydroxylase (CA4H), a cytochrome P-450 catalyzing the second reaction of the general phenylpropanoid pathway, from Jerusalem artichoke (Helianthus tuberosus L.) (B. Gabriac, D. Werck-Reichhart, H. Teutsch, F. Durst [1991] Arch Biochem Biophys 288: 302-309). Rabbit polyclonal antibodies were raised against the native and denaturated nitrocellulose-bound enzyme. Only the immunoglobulins G (IgGs) elicited upon immunization with native enzyme produced strong inhibition of catalytic activity and good cross-reactivity on western blots. In microsomes from H. tuberosus tissues induced by wounding and various chemicals, a positive correlation between catalytic activity and amounts of immunoreactive protein on western blots was observed. When coupled to cyanogen bromide-activated Sepharose, purified IgGs selectively retained CA4H activity from solubilized plant microsomes. Acid elution from the immunoaffinity matrix provided a rapid procedure for high-yield purification of the CA4H protein. The same IgGs immunoprecipitated a single protein from the in vitro translation products of mRNA isolated from wounded tissues. The apparent molecular weight (57,000) of this polypeptide was identical to that of CA4H purified from tuber microsomes. Immunochemical relatedness between CA4H from different plant species was demonstrated by strong inhibition of catalytic activity and immunopurification of several orthologous enzymes, using IgGs directed against CA4H from H. tuberosus. However, only limited interspecies cross-reactivity was observed on western blots. A careful immunochemical analysis indicates that CA4H immunoreactivity significantly differs from plant to plant. Results are discussed in terms of antibody specificity, enzyme glycosylation, and CA4H regulation. PMID:8278549

  6. Substrate specificity of human liver cytochrome P-450 debrisoquine 4-hydroxylase probed using immunochemical inhibition and chemical modeling.

    PubMed

    Wolff, T; Distlerath, L M; Worthington, M T; Groopman, J D; Hammons, G J; Kadlubar, F F; Prough, R A; Martin, M V; Guengerich, F P

    1985-05-01

    A significant population of humans (5 to 10%) are phenotypic poor metabolizers of debrisoquine. We have isolated the cytochrome P-450 isozyme from rat liver responsible for this activity and have shown that antibodies raised against the protein are able to inhibit this catalytic activity in human liver microsomes (Distlerath, L. M., and Guengerich, F. P., Proc. Natl. Acad. Sci. USA, 81: 7348-7352, 1984). These antibodies were utilized to determine which metabolic transformations are linked to debrisoquine 4-hydroxylation in human liver microsomes using techniques of immunochemical inhibition. The antibodies almost completely inhibited debrisoquine 4-hydroxylation and bufuralol 1'-hydroxylation in microsomes prepared from several different human livers. The oxidation of the pyrrolizidine alkaloids lasiocarpine and monocrotaline were inhibited by roughly one-third. The antibodies did not inhibit N,N-dimethylnitrosamine N-demethylation, oxidation of vinylidene chloride to 2,2-chloroacetaldehyde, oxidation of trichloroethylene to chloral, N-oxidation of azoprocarbazine, morphine N-demethylation, diazepam N-demethylation, oxidation of benzo(a)pyrene to alkali-soluble metabolites, oxidation of benzo(a)pyrene 7,8-dihydrodiol to products covalently bound to DNA, the N- and ring-oxidation of 1- and 2-naphthylamine and 2-aminofluorene, or the conversion of aflatoxin B1 to DNA adducts or aflatoxin Q1. Studies with space-filling models of the drugs the metabolism of which is associated with debrisoquine 4-hydroxylase in the literature indicated that all can be fitted to a general structure in which a basic nitrogen is about 5 A away from the site of carbon hydroxylation and a hydrophobic domain is near the site of hydroxylation. These results may be useful in predicting which chemicals may or may not be metabolized in an atypical manner by a segment of the human population.

  7. Enhancement of L-tryptophan 5-hydroxylation activity by structure-based modification of L-phenylalanine 4-hydroxylase from Chromobacterium violaceum.

    PubMed

    Kino, Kuniki; Hara, Ryotaro; Nozawa, Ai

    2009-09-01

    The objective of this study was to enhance l-tryptophan hydroxylation activity of l-phenylalanine 4-hydroxylase. It had been known that l-phenylalanine 4-hydroxylase from Chromobacterium violaceum could convert l-tryptophan to 5-hydroxy-l-tryptophan and l-phenylalanine to l-tyrosine; however, the activity for l-tryptophan was extremely low compared to l-phenylalanine activity levels. We used the information on the crystal structures of aromatic amino acid hydroxylases to generate C. violaceuml-phenylalanine 4-hydroxylase with high l-tryptophan hydroxylating activity. In silico structural modeling analysis suggested that hydrophobic and/or stacking interactions with the substrate and cofactor at L101 and W180 in C. violaceuml-phenylalanine 4-hydroxylase would increase hydroxylation activity. Based on this hypothesis, we introduced a saturation mutagenesis towards these sites followed by the evaluation of 5-hydroxy-l-tryptophan productivity using a modified Gibbs assay. Three and nine positive mutants were obtained from the L101 and W180 mutant libraries, respectively. Among the mutants, L101Y and W180F showed the highest l-tryptophan hydroxylation activity at the respective residues. Steady-state kinetic analysis revealed that k(cat) values for l-tryptophan hydroxylation were increased from 0.40 (wild-type) to 1.02 (L101Y) and 0.51 s(-1) (W180F). In addition, the double mutant (L101Y-W180F) displayed higher l-tryptophan hydroxylation activity than the wild-type and the W180F and L101Y mutants. The k(cat) value of L101Y-W180F increased to 2.08 s(-1), showing a 5.2-fold increase compared to wild-type enzyme levels.

  8. Hypoxia-inducible factor-1 (HIF-1) but not HIF-2 is essential for hypoxic induction of collagen prolyl 4-hydroxylases in primary newborn mouse epiphyseal growth plate chondrocytes.

    PubMed

    Aro, Ellinoora; Khatri, Richa; Gerard-O'Riley, Rita; Mangiavini, Laura; Myllyharju, Johanna; Schipani, Ernestina

    2012-10-26

    Hypoxia-inducible factors (HIFs) are the master regulators of hypoxia-responsive genes. They play a critical role in the survival, development, and differentiation of chondrocytes in the avascular hypoxic fetal growth plate, which is rich in extracellular matrix (ECM) and in its main component, collagens. Several genes involved in the synthesis, maintenance, and degradation of ECM are regulated by HIFs. Collagen prolyl 4-hydroxylases (C-P4Hs) are key enzymes in collagen synthesis because the resulting 4-hydroxyprolines are necessary for the stability of all collagen molecules. The vertebrate C-P4Hs are α(2)β(2) tetramers with three isoforms of the catalytic α subunit, yielding C-P4Hs of types I-III. C-P4H-I is the main form in most cells, but C-P4H-II is the major form in chondrocytes. We postulated here that post-translational modification of collagens, particularly 4-hydroxylation of proline residues, could be one of the modalities by which HIF regulates the adaptive responses of chondrocytes in fetal growth plates. To address this hypothesis, we used primary epiphyseal growth plate chondrocytes isolated from newborn mice with conditionally inactivated genes for HIF-1α, HIF-2α, or the von Hippel-Lindau protein. The data obtained showed that C-P4H α(I) and α(II) mRNA levels were increased in hypoxic chondrocytes in a manner dependent on HIF-1 but not on HIF-2. Furthermore, the increases in the C-P4H mRNA levels were associated with both increased amounts of the C-P4H tetramers and augmented C-P4H activity in hypoxia. The hypoxia inducibility of the C-P4H isoenzymes is thus likely to ensure sufficient C-P4H activity for collagen synthesis occurring in chondrocytes in a hypoxic environment.

  9. Novel characteristics and regulation of a divergent cinnamate 4-hydroxylase (CYP73A15) from French bean: engineering expression in yeast.

    PubMed

    Nedelkina, S; Jupe, S C; Blee, K A; Schalk, M; Werck-Reichhart, D; Bolwell, G P

    1999-04-01

    cDNAs showing high sequence similarity (>70%) over large stretches to plant CYP73A orthologues from other species were isolated from a cDNA library derived from mRNAs expressed in elicitor-treated suspension-cultured cells. These clones appear to code for a full-length 1554 bp open reading frame with a 78 bp 5'-untranslated region and a 140 bp 3'-untranslated region. The open reading frame, determined by sequence similarity, codes for a protein with a predicted Mr of 59229 and a pI of 8.8. It contains the conserved cysteine haem-binding site found in all cytochrome P450s. The protein encoded by this cDNA diverges however from other CYP73As in its N- and C-terminus and in four domains internally, so that overall sequence similarity is in the range 58-66%. Many clones contained an identical intron, which may be associated with a novel regulatory mechanism. Sequence similarity is sufficient for it to be classified as CYP73A15, although it is the least similar member of this family classified so far. The cDNA was expressed in yeast. Successful expression of cinnamate 4-hydroxylase activity required removal of the intron. High-level expression also required modification of the N-terminus to that of CYP73A1. Yeast did not process the intron at all and the leader sequence for A15 was not as compatible as that of A1. The mRNA for CYP73A15 was shown to be rapidly induced by elicitor treatment of suspension-cultured cells of French bean but induction was more transient than that of phenylalanine ammonia-lyase (PAL). In contrast, induction in cells undergoing xylogenesis was much more coordinate with PAL. The cloned cDNA may represent a cinnamate 4-hydroxylase isoform, whose expression is more related to differentiation than the responses to stress in which the majority of CYP73As cloned so far are involved.

  10. Cellular oxygen sensing: Importins and exportins are mediators of intracellular localisation of prolyl-4-hydroxylases PHD1 and PHD2

    SciTech Connect

    Steinhoff, Amrei; Pientka, Friederike Katharina; Moeckel, Sylvia; Kettelhake, Antje; Hartmann, Enno; Koehler, Matthias; Depping, Reinhard

    2009-10-02

    Hypoxia-inducible factors are crucial in the regulatory process of oxygen homeostasis of vertebrate cells. Inhibition of prolyl hydroxylation of HIF-{alpha} subunits by prolyl-hydroxylases (PHD1, PHD2 and PHD3) leads to transcription of a greater number of hypoxia responsive genes. We have investigated the subcellular distribution and the molecular mechanisms regulating the intracellular allocation of PHD1 and PHD2. As reported earlier we find PHD1 located exclusively in the nucleus. We demonstrate that nuclear import of PHD1 occurs importin {alpha}/{beta} dependently and relies on a nuclear localisation signal (NLS). By contrast PHD2 is cycling between nucleus and cytoplasm, and nuclear import seems to be independent of 'classical' importin {alpha}/{beta} receptors. Furthermore, we reveal that the exit of PHD2 from the nucleus requires CRM1 and the N-terminal 100 amino acids of the protein. Our findings provide new insights into the mechanisms of the regulation of the oxygen sensor cascade of PHDs in different cellular compartments.

  11. Overexpression of Prolyl-4-Hydroxylase-α1 Stabilizes but Increases Shear Stress-Induced Atherosclerotic Plaque in Apolipoprotein E-Deficient Mice

    PubMed Central

    Liu, Xin-xin; Li, Meng-meng; Zhang, Yu; Chen, Liang; Wang, Lin; Di, Ming-xue

    2016-01-01

    The rupture and erosion of atherosclerotic plaque can induce coronary thrombosis. Prolyl-4-hydroxylase (P4H) plays a central role in the synthesis of all known types of collagens, which are the most abundant constituent of the extracellular matrix in atherosclerotic plaque. The pathogenesis of atherosclerosis is thought to be in part caused by shear stress. In this study, we aimed to investigate a relationship between P4Hα1 and shear stress-induced atherosclerotic plaque. Carotid arteries of ApoE−/− mice were exposed to low and oscillatory shear stress conditions by the placement of a shear stress cast for 2 weeks; we divided 60 male ApoE−/− mice into three groups for treatments with saline (mock) (n = 20), empty lentivirus (lenti-EGFP) (n = 20), and lentivirus-P4Hα1 (lenti-P4Hα1) (n = 20). Our results reveal that after 2 weeks of lenti-P4Hα1 treatment both low and oscillatory shear stress-induced plaques increased collagen and the thickness of fibrous cap and decreased macrophage accumulation but no change in lipid accumulation. We also observed that overexpression of P4Ha1 increased plaque size. Our study suggests that P4Hα1 overexpression might be a potential therapeutic target in stabilizing vulnerable plaques. PMID:27818566

  12. Pichia pastoris production of a prolyl 4-hydroxylase derived from Chondrosia reniformis sponge: A new biotechnological tool for the recombinant production of marine collagen.

    PubMed

    Pozzolini, Marina; Scarfì, Sonia; Mussino, Francesca; Salis, Annalisa; Damonte, Gianluca; Benatti, Umberto; Giovine, Marco

    2015-08-20

    Prolyl 4-hydroxylase (P4H) is a α2β2 tetramer catalyzing the post-translational hydroxylation of prolines in collagen. Its recombinant production is mainly pursued to realize biotechnological tools able to generate animal contaminant-free hydroxylated collagen. One promising candidate for biomedical applications is the collagen extracted from the marine sponge Chondrosia reniformis, because of its biocompatibility and because is devoid of the health risks associated with bovine and porcine collagens. Here we report on the production and selection, by enzymatic and biomolecular analyses, of a triple transformed Pichia pastoris strain expressing a stable P4H tetramer derived from C. reniformis sponge and a hydroxylated non fibrillar procollagen polypeptide from the same animal. The percentage of recombinant procollagen hydroxylated prolines inside the transformed yeast was of 36.3% analyzed by mass spectrometry indicating that the recombinant enzyme is active on its natural substrate inside the yeast cell host. Furthermore, the recombinant sponge P4H has the ability to hydroxylate its natural substrate in both X and Y positions in the Xaa-Yaa-Gly collagenous triplets. In conclusion this Pichia system seems ideal for high-level production of hydroxylated sponge- or marine-derived collagen polypeptides as well as of conotoxins or other marine proteins of high pharmacological interest needing this particular post-translational modification.

  13. Microbial production of N-acetyl cis-4-hydroxy-L-proline by coexpression of the Rhizobium L-proline cis-4-hydroxylase and the yeast N-acetyltransferase Mpr1.

    PubMed

    Bach, Thi Mai Hoa; Hara, Ryotaro; Kino, Kuniki; Ohtsu, Iwao; Yoshida, Nobuyuki; Takagi, Hiroshi

    2013-01-01

    The proline analogue cis-4-hydroxy-L-proline (CHOP), which inhibits the biosynthesis of collagen, has been clinically evaluated as an anticancer drug, but its water solubility and low molecular weight limits its therapeutic potential since it is rapidly excreted. In addition, CHOP is too toxic to be practical as an anticancer drug, due primarily to its systematic effects on noncollagen proteins. To promote CHOP's retention in blood and/or to decrease its toxicity, N-acetylation of CHOP might be a novel approach as a prodrug. The present study was designed to achieve the microbial production of N-acetyl CHOP from L-proline by coexpression of L-proline cis-4-hydroxylases converting L-proline into CHOP (SmP4H) from the Rhizobium Sinorhizobium meliloti and N-acetyltransferase converting CHOP into N-acetyl CHOP (Mpr1) from the yeast Saccharomyces cerevisiae. We constructed a coexpression plasmid harboring both the SmP4H and Mpr1 genes and introduced it into Escherichia coli BL21(DE3) or its L-proline oxidase gene-disrupted (ΔputA) strain. M9 medium containing L-proline produced more N-acetyl CHOP than LB medium containing L-proline. E. coli ΔputA cells accumulated L-proline (by approximately 2-fold) compared to that in wild-type cells, but there was no significant difference in CHOP production between wild-type and ΔputA cells. The addition of NaCl and L-ascorbate resulted in a 2-fold increase in N-acetyl CHOP production in the L-proline-containing M9 medium. The highest yield of N-acetyl CHOP was achieved at 42 h cultivation in the optimized medium. Five unknown compounds were detected in the total protein reaction, probably due to the degradation of N-acetyl CHOP. Our results suggest that weakening of the degradation or deacetylation pathway improves the productivity of N-acetyl CHOP.

  14. Isolation and sequence of a cDNA encoding the Jerusalem artichoke cinnamate 4-hydroxylase, a major plant cytochrome P450 involved in the general phenylpropanoid pathway.

    PubMed Central

    Teutsch, H G; Hasenfratz, M P; Lesot, A; Stoltz, C; Garnier, J M; Jeltsch, J M; Durst, F; Werck-Reichhart, D

    1993-01-01

    Cinnamate 4-hydroxylase [CA4H; trans-cinnamate,NADPH:oxygen oxidoreductase (4-hydroxylating), EC 1.14.13.11] is a cytochrome P450 that catalyzes the first oxygenation step of the general phenylpropanoid metabolism in higher plants. The compounds formed are essential for lignification and defense against predators and pathogens. We recently reported the purification of this enzyme from Mn(2+)-induced Jerusalem artichoke (Helianthus tuberosus L.) tuber tissues. Highly selective polyclonal antibodies raised against the purified protein were used to screen a lambda gt11 cDNA expression library from wound-induced Jerusalem artichoke, allowing isolation of a 1130-base-pair insert. Typical P450 domains were identified in this incomplete sequence, which was used as a probe for the isolation of a 1.7-kilobase clone in a lambda gt10 library. A full-length open reading frame of 1515 base pairs, encoding a P450 protein of 505 residues (M(r) = 57,927), was sequenced. The N terminus, essentially composed of hydrophobic residues, matches perfectly the microsequenced N terminus of the purified protein. The calculated pI is 9.78, in agreement with the chromatographic behavior and two-dimensional electrophoretic analysis of CA4H. Synthesis of the corresponding mRNA is induced in wounded plant tissues, in correlation with CA4H enzymatic activity. This P450 protein exhibits the most similarity (28% amino acid identity) with avocado CYP71, but also good similarity with CYP17 and CYP21, or with CYP1 and CYP2 families. According to current criteria, it qualifies as a member of a new P450 family. Images Fig. 4 PMID:8097885

  15. Prolyl-4-hydroxylase Domain Protein 2 Controls NF-κB/p65 Transactivation and Enhances the Catabolic Effects of Inflammatory Cytokines on Cells of the Nucleus Pulposus*

    PubMed Central

    Li, Jun; Yuan, Wen; Jiang, Shuai; Ye, Wei; Yang, Hao; Shapiro, Irving M.; Risbud, Makarand V.

    2015-01-01

    Prolyl-4-hydroxylase (PHD) proteins are key in sensing tissue hypoxia. In nucleus pulposus (NP) cells, our previous work demonstrated that PHD isoforms have a differential contribution in controlling hypoxia-inducible factor (HIF)-α degradation and activity. Recently we have shown that a regulatory relationship exists between PHD3 and inflammatory cytokines in NP cells. With respect to PHD2, the most abundant PHD isoform in NP cells, very little is known concerning its function and regulation under inflammatory conditions that characterize intervertebral disc degeneration. Here, we show that PHD2 is a potent regulator of the catabolic activities of TNF-α; silencing of PHD2 significantly decreased TNF-α-induced expression of catabolic markers including SDC4, MMP-3, MMP-13, and ADAMTS5, as well as several inflammatory cytokines and chemokines, while partially restoring aggrecan and collagen II expression. Use of NF-κB reporters with ShPHD2, SiHIF-1α, as well as p65−/−, PHD2−/−, and PHD3−/− cells, shows that PHD2 serves as a co-activator of NF-κB/p65 signaling in HIF-1-independent fashion. Immunoprecipitation of endogenous and exogenously expressed tagged proteins, as well as fluorescence microscopy, indicates that following TNF-α treatment, PHD2 interacts and co-localizes with p65. Conversely, loss of function experiments using lentivirally delivered Sh-p65, Sh-IKKβ, and NF-κB inhibitor confirmed that cytokine-dependent PHD2 expression in NP cells requires NF-κB signaling. These findings clearly demonstrate that PHD2 forms a regulatory circuit with TNF-α via NF-κB and thereby plays an important role in enhancing activity of this cytokine. We propose that during disc degeneration PHD2 may offer a therapeutic target to mitigate the deleterious actions of TNF-α, a key proinflammatory cytokine. PMID:25635047

  16. Cell-free synthesis and assembly of prolyl 4-hydroxylase: the role of the beta-subunit (PDI) in preventing misfolding and aggregation of the alpha-subunit.

    PubMed Central

    John, D C; Grant, M E; Bulleid, N J

    1993-01-01

    Prolyl 4-hydroxylase (P4-H) catalyses a vital post-translational modification in the biosynthesis of collagen. The enzyme consists of two distinct polypeptides forming an alpha 2 beta 2 tetramer (alpha = 64 kDa, beta = 60 kDa), the beta-subunit being identical to the multifunctional enzyme protein disulfide isomerase (PDI). By studying the cell-free synthesis of the rat alpha-subunit of P4-H we have shown that the alpha-subunit can be translocated, glycosylated and the signal peptide cleaved by dog pancreatic microsomal membranes to yield both singly and doubly glycosylated forms. When translations were carried out under conditions which prevent disulfide bond formation, the product synthesized formed aggregates which were associated with the immunoglobulin heavy chain binding protein (BiP). Translations carried out under conditions that promote disulfide bond formation yielded a product that was not associated with BiP but formed a complex with the endogenous beta-subunit (PDI). Complex formation was detected by co-precipitation of the newly synthesized alpha-subunit with antibodies raised against PDI, by sucrose gradient centrifugation and by chemical cross-linking. When microsomal vesicles were depleted of PDI, BiP and other soluble endoplasmic reticulum proteins, no complex formation was observed and the alpha-subunit aggregated even under conditions that promote disulfide bond formation. We have therefore demonstrated that the enzyme P4-H can be assembled at synthesis in a cell-free system and that the solubility of the alpha-subunit is dependent upon its association with PDI. Images PMID:8385607

  17. Severe Extracellular Matrix Abnormalities and Chondrodysplasia in Mice Lacking Collagen Prolyl 4-Hydroxylase Isoenzyme II in Combination with a Reduced Amount of Isoenzyme I.

    PubMed

    Aro, Ellinoora; Salo, Antti M; Khatri, Richa; Finnilä, Mikko; Miinalainen, Ilkka; Sormunen, Raija; Pakkanen, Outi; Holster, Tiina; Soininen, Raija; Prein, Carina; Clausen-Schaumann, Hauke; Aszódi, Attila; Tuukkanen, Juha; Kivirikko, Kari I; Schipani, Ernestina; Myllyharju, Johanna

    2015-07-03

    Collagen prolyl 4-hydroxylases (C-P4H-I, C-P4H-II, and C-P4H-III) catalyze formation of 4-hydroxyproline residues required to form triple-helical collagen molecules. Vertebrate C-P4Hs are α2β2 tetramers differing in their catalytic α subunits. C-P4H-I is the major isoenzyme in most cells, and inactivation of its catalytic subunit (P4ha1(-/-)) leads to embryonic lethality in mouse, whereas P4ha1(+/-) mice have no abnormalities. To study the role of C-P4H-II, which predominates in chondrocytes, we generated P4ha2(-/-) mice. Surprisingly, they had no apparent phenotypic abnormalities. To assess possible functional complementarity, we established P4ha1(+/-);P4ha2(-/-) mice. They were smaller than their littermates, had moderate chondrodysplasia, and developed kyphosis. A transient inner cell death phenotype was detected in their developing growth plates. The columnar arrangement of proliferative chondrocytes was impaired, the amount of 4-hydroxyproline and the Tm of collagen II were reduced, and the extracellular matrix was softer in the growth plates of newborn P4ha1(+/-);P4ha2(-/-) mice. No signs of uncompensated ER stress were detected in the mutant growth plate chondrocytes. Some of these defects were also found in P4ha2(-/-) mice, although in a much milder form. Our data show that C-P4H-I can to a large extent compensate for the lack of C-P4H-II in proper endochondral bone development, but their combined partial and complete inactivation, respectively, leads to biomechanically impaired extracellular matrix, moderate chondrodysplasia, and kyphosis. Our mouse data suggest that inactivating mutations in human P4HA2 are not likely to lead to skeletal disorders, and a simultaneous decrease in P4HA1 function would most probably be required to generate such a disease phenotype. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. The prolyl 4-hydroxylase inhibitor GSK360A decreases post-stroke brain injury and sensory, motor, and cognitive behavioral deficits.

    PubMed

    Zhou, Jin; Li, Jie; Rosenbaum, Daniel M; Zhuang, Jian; Poon, Carrie; Qin, Pu; Rivera, Katrina; Lepore, John; Willette, Robert N; Hu, Erding; Barone, Frank C

    2017-01-01

    There is interest in pharmacologic preconditioning for end-organ protection by targeting the HIF system. This can be accomplished by inhibition of prolyl 4-hydroxylase (PHD). GSK360A is an orally active PHD inhibitor that has been previously shown to protect the failing heart. We hypothesized that PHD inhibition can also protect the brain from injuries and resulting behavioral deficits that can occur as a result of surgery. Thus, our goal was to investigate the effect of pre-stroke surgery brain protection using a verified GSK360A PHD inhibition paradigm on post-stroke surgery outcomes. Vehicle or an established protective dose (30 mg/kg, p.o.) of GSK360A was administered to male Sprague-Dawley rats. Initially, GSK360A pharmacokinetics and organ distribution were determined, and then PHD-HIF pharmacodynamic markers were measured (i.e., to validate the pharmacological effects of the GSK360A administration regimen). Results obtained using this validated PHD dose-regimen indicated significant improvement by GSK360A (30mg/kg); administered at 18 and 5 hours prior to transient middle cerebral artery occlusion (stroke). GSK360A exposure and plasma, kidney and brain HIF-PHD pharmacodynamics endpoints (e.g., erythropoietin; EPO and Vascular Endothelial Growth Factor; VEGF) were measured. GSK360A provided rapid exposure in plasma (7734 ng/ml), kidney (45-52% of plasma level) and brain (1-4% of plasma level), and increased kidney EPO mRNA (80-fold) and brain VEGF mRNA (2-fold). We also observed that GSK360A increased plasma EPO (300-fold) and VEGF (2-fold). Further assessments indicated that GSK360A reduced post-stroke surgery neurological deficits (47-64%), cognitive dysfunction (60-75%) and brain infarction (30%) 4 weeks later. Thus, PHD inhibition using GSK360A pretreatment produced long-term post-stroke brain protection and improved behavioral functioning. These data support PHD inhibition, specifically by GSK360A, as a potential strategy for pre-surgical use to reduce

  19. Impact of homeobox genes in gastrointestinal cancer

    PubMed Central

    Joo, Moon Kyung; Park, Jong-Jae; Chun, Hoon Jai

    2016-01-01

    Homeobox genes, including HOX and non-HOX genes, have been identified to be expressed aberrantly in solid tumors. In gastrointestinal (GI) cancers, most studies have focused on the function of non-HOX genes including caudal-related homeobox transcription factor 1 (CDX1) and CDX2. CDX2 is a crucial factor in the development of pre-cancerous lesions such as Barrett’s esophagus or intestinal metaplasia in the stomach, and its tumor suppressive role has been investigated in colorectal cancers. Recently, several HOX genes were reported to have specific roles in GI cancers; for example, HOXA13 in esophageal squamous cell cancer and HOXB7 in stomach and colorectal cancers. HOXD10 is upregulated in colorectal cancer while it is silenced epigenetically in gastric cancer. Thus, it is essential to examine the differential expression pattern of various homeobox genes in specific tumor types or cell lineages, and understand their underlying mechanisms. In this review, we summarize the available research on homeobox genes and present their potential value for the prediction of prognosis in GI cancers. PMID:27729732

  20. Catechol estrogen formation by brain tissue: characterization of a direct product isolation assay for estrogen-2- and 4-hydroxylase activity and its application to studies of 2- and 4-hydroxyestradiol formation by rabbit hypothalamus

    SciTech Connect

    Hersey, R.M.; Williams, K.I.; Weisz, J.

    1981-12-01

    A direct product isolation assay for quantifying the formation of 2- and 4-hydroxyestradiol (2-OHE2 and 4-OHE2) from (6,7-3H)estradiol by rabbit hypothalami in vitro was developed, and the assay was used to characterize some properties of estrogen-2- and 4-hydroxylase activity in this tissue. The reaction was carried out under conditions that minimized further metabolism of enzymatically formed catechol estrogens. A simple two-step separation procedure, involving the use of a neutral alumina column, followed by thin layer chromatography, was developed to isolate the enzymatically formed catechol estrogens in a radiochemically homogeneous form. The detergent, Tween-80, was found to activate the enzyme and was used routinely at a concentration of 0.1% in the assay. The formation of 2-OHE2 was linear up to 10 min and with increasing protein concentrations up to 150 micrograms/incubation. Similar values were obtained for 4-OHE2. Maximum velocities (Vmax) for the formation of 2- and 4-OHE2 were 190 and 270 pmol/mg protein . 10 min, respectively. The apparent Km values with respect to estradiol for 2-OHE2 and 4-OHE2 were 125 and 150 microM, respectively. The highest specific activity for the enzyme was present in the 100,000 X g supernatant (S3), while the activity in the microsomal fraction (P3) was less than that in the original homogenate. Enzyme activity depended on the presence of NADPH and oxygen and was inhibited by CO as well as by high concentrations of SKF-525A. Estrogen-2- and 4-hydroxylase activity in rabbit hypothalamus differed from that in rat liver in two respects. In the liver, enzyme activity was localized in the microsomal fraction and was virtually abolished by Tween-80. In contrast, enzyme activity in rabbit hypothalamus was maximal in the soluble fraction (100,000 X g supernatant)and was stimulated by the detergent.

  1. Human debrisoquine 4-hydroxylase (P450IID1): cDNA and deduced amino acid sequence and assignment of the CYP2D locus to chromosome 22.

    PubMed

    Gonzalez, F J; Vilbois, F; Hardwick, J P; McBride, O W; Nebert, D W; Gelboin, H V; Meyer, U A

    1988-02-01

    The enzyme P450db1 (db1) is responsible for the common human defect in drug oxidation known as the "debrisoquine/sparteine polymorphism." Polyclonal antibody against the rat db1 protein was used to screen a human liver lambda gt11 library for the db1 cDNA clone. A cDNA containing the full protein coding sequence was isolated; the deduced NH2-terminal sequence of this cDNA was identical to that derived from direct sequencing of the purified human db1 protein. Comparison of the human db1 with rat db1 revealed 71 and 73% similarities of nucleotides and amino acids, respectively. By use of human-rodent somatic cell hybrids the db1 gene was localized to human chromosome 22 (CYP2D locus).

  2. Molecular cloning and characterization of desacetoxyvindoline-4-hydroxylase, a 2-oxoglutarate dependent-dioxygenase involved in the biosynthesis of vindoline in Catharanthus roseus (L.) G. Don.

    PubMed

    Vazquez-Flota, F; De Carolis, E; Alarco, A M; De Luca, V

    1997-08-01

    A 2-oxoglutarate-dependent dioxygenase (EC 1.14.11.11) which catalyzes the 4-hydroxylation of desacetoxyvindoline was purified to homogeneity. Three oligopeptides isolated from a tryptic digest of the purified protein were microsequenced and one oligopeptide showed significant homology to hyoscyamine 6 beta-hydroxylase from Hyoscyamus niger. A 36-mer degenerate oligonucleotide based on this peptide sequence was used to screen a Catharanthus roseus cDNA library and three clones, cD4H-1 to -3, were isolated. Although none of the three clones were full-length, the open reading frame on each clone encoded a putative protein containing the sequence of all three peptides. Primer extension analysis suggested that cD4H-3, the longest cDNA clone, was missing 156 bp at the 5' end of the clone and sequencing of the genomic clone, gD4H-8, confirmed these results. Southern blot analysis suggested that d4h is present as a single-copy gene in C. roseus which is a diploid plant, and the significant differences in the sequence of the 3'-UTR between cD4H-1 and -3 suggest that they represent dimorphic alleles of the same hydroxylase. The identity of the clone was further confirmed when extracts of transformed Escherichia coli expressed D4H enzyme activity. The D4H clone encoded a putative protein of 401 amino acids with a calculated molecular mass of 45.5 kDa and the amino acid sequence showed a high degree of similarity with those of a growing family of 2-oxoglutarate-dependent dioxygenases of plant and fungal origin. The similarity was not restricted to the dioxygenase protein sequences but was also extended to the gene structure and organization since the 205 and 1720 bp introns of d4h were inserted around the same highly conserved amino acid consensus sequences as those for e8 protein, hyoscyamine-6 beta-hydroxylase and ethylene-forming enzyme. These results provide further support that a common ancestral gene is responsible for the appearance of this family of dioxygenases

  3. Impact of RNA degradation on gene expression profiling

    PubMed Central

    2010-01-01

    Background Gene expression profiling is a highly sensitive technique which is used for profiling tumor samples for medical prognosis. RNA quality and degradation influence the analysis results of gene expression profiles. The impact of this influence on the profiles and its medical impact is not fully understood. As patient samples are very valuable for clinical studies, it is necessary to establish criteria for the RNA quality to be able to use these samples in later analysis. Methods To investigate the effects of RNA integrity on gene expression profiling, whole genome expression arrays were used. We used tumor biopsies from patients diagnosed with locally advanced rectal cancer. To simulate degradation, the isolated total RNA of all patients was subjected to heat-induced degradation in a time-dependent manner. Expression profiling was then performed and data were analyzed bioinformatically to assess the differences. Results The differences introduced by RNA degradation were largely outweighed by the biological differences between the patients. Only a relatively small number of probes (275 out of 41,000) show a significant effect due to degradation. The genes that show the strongest effect due to RNA degradation were, especially, those with short mRNAs and probe positions near the 5' end. Conclusions Degraded RNA from tumor samples (RIN > 5) can still be used to perform gene expression analysis. A much higher biological variance between patients is observed compared to the effect that is imposed by degradation of RNA. Nevertheless there are genes, very short ones and those with the probe binding side close to the 5' end that should be excluded from gene expression analysis when working with degraded RNA. These results are limited to the Agilent 44 k microarray platform and should be carefully interpreted when transferring to other settings. PMID:20696062

  4. Impact of Statins on Gene Expression in Human Lung Tissues

    PubMed Central

    Lane, Jérôme; van Eeden, Stephan F.; Obeidat, Ma’en; Sin, Don D.; Tebbutt, Scott J.; Timens, Wim; Postma, Dirkje S.; Laviolette, Michel; Paré, Peter D.; Bossé, Yohan

    2015-01-01

    Statins are 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors that alter the synthesis of cholesterol. Some studies have shown a significant association of statins with improved respiratory health outcomes of patients with asthma, chronic obstructive pulmonary disease and lung cancer. Here we hypothesize that statins impact gene expression in human lungs and may reveal the pleiotropic effects of statins that are taking place directly in lung tissues. Human lung tissues were obtained from patients who underwent lung resection or transplantation. Gene expression was measured on a custom Affymetrix array in a discovery cohort (n = 408) and two replication sets (n = 341 and 282). Gene expression was evaluated by linear regression between statin users and non-users, adjusting for age, gender, smoking status, and other covariables. The results of each cohort were combined in a meta-analysis and biological pathways were studied using Gene Set Enrichment Analysis. The discovery set included 141 statin users. The lung mRNA expression levels of eighteen and three genes were up-regulated and down-regulated in statin users (FDR < 0.05), respectively. Twelve of the up-regulated genes were replicated in the first replication set, but none in the second (p-value < 0.05). Combining the discovery and replication sets into a meta-analysis improved the significance of the 12 up-regulated genes, which includes genes encoding enzymes and membrane proteins involved in cholesterol biosynthesis. Canonical biological pathways altered by statins in the lung include cholesterol, steroid, and terpenoid backbone biosynthesis. No genes encoding inflammatory, proteases, pro-fibrotic or growth factors were altered by statins, suggesting that the direct effect of statin in the lung do not go beyond its antilipidemic action. Although more studies are needed with specific lung cell types and different classes and doses of statins, the improved health outcomes and survival observed in statin

  5. Cinnamic acid 4-hydroxylase mechanism-based inactivation by psoralen derivatives: cloning and characterization of a C4H from a psoralen producing plant-Ruta graveolens-exhibiting low sensitivity to psoralen inactivation.

    PubMed

    Gravot, Antoine; Larbat, Romain; Hehn, Alain; Lièvre, Karine; Gontier, Eric; Goergen, Jean Louis; Bourgaud, Frédéric

    2004-02-01

    Cinnamate 4-hydroxylase (C4H, EC 1.14.13.11) complete cDNA was cloned from the leaves of Ruta graveolens, a psoralen producing plant. The recombinant enzyme (classified CYP73A32) was expressed in Saccharomyces cerevisiae. Mechanism-based inactivation was investigated using various psoralen derivatives. Only psoralen and 8-methoxypsoralen were found to inactivate C4H. The inactivation was dependent on the presence of NADPH, time of pre-incubation, and inhibitor concentration. Inactivation stoichiometry was 0.9 (+/-0.2) for CYP73A1 and 1.1 (+/-0.2) for CYP73A32. SDS-PAGE analysis demonstrated that [3H]psoralen was irreversibly bound to the C4H apoprotein. K(i) and k(inact) for psoralen and 8-methoxypsoralen inactivation on the two C4H revealed a lower sensitivity for CYP73A32 compared to CYP73A1. Inactivation kinetics were also determined for CYP73A10, a C4H from another furocoumarin-producing plant, Petroselinum crispum. This enzyme was found to behave like CYP73A32, with a weak sensitivity to psoralen and 8-MOP inactivation. Cinnamic acid hydroxylation is a key step in the biosynthesis of phenylpropanoid compounds, psoralen derivatives included. Our results suggest a possible evolution of R. graveolens and P. crispum C4H that might tolerate substantial levels of psoralen derivatives in the cytoplasmic compartment without a depletive effect on C4H and the general phenylpropanoid metabolism.

  6. The Impact of Gene Silencing on Horizontal Gene Transfer and Bacterial Evolution.

    PubMed

    Navarre, W W

    2016-01-01

    The H-NS family of DNA-binding proteins is the subject of intense study due to its important roles in the regulation of horizontally acquired genes critical for virulence, antibiotic resistance, and metabolism. Xenogeneic silencing proteins, typified by the H-NS protein of Escherichia coli, specifically target and downregulate expression from AT-rich genes by selectively recognizing specific structural features unique to the AT-rich minor groove. In doing so, these proteins facilitate bacterial evolution; enabling these cells to engage in horizontal gene transfer while buffering potential any detrimental fitness consequences that may result from it. Xenogeneic silencing and counter-silencing explain how bacterial cells can evolve effective gene regulatory strategies in the face of rampant gene gain and loss and it has extended our understanding of bacterial gene regulation beyond the classic operon model. Here we review the structures and mechanisms of xenogeneic silencers as well as their impact on bacterial evolution. Several H-NS-like proteins appear to play a role in facilitating gene transfer by other mechanisms including by regulating transposition, conjugation, and participating in the activation of virulence loci like the locus of enterocyte effacement pathogenicity island of pathogenic strains of E. coli. Evidence suggests that the critical determinants that dictate whether an H-NS-like protein will be a silencer or will perform a different function do not lie in the DNA-binding domain but, rather, in the domains that control oligomerization. This suggests that H-NS-like proteins are transcription factors that both recognize and alter the shape of DNA to exert specific effects that include but are not limited to gene silencing. © 2016 Elsevier Ltd All rights reserved.

  7. Impact of the cell division cycle on gene circuits

    NASA Astrophysics Data System (ADS)

    Bierbaum, Veronika; Klumpp, Stefan

    2015-12-01

    In growing cells, protein synthesis and cell growth are typically not synchronous, and, thus, protein concentrations vary over the cell division cycle. We have developed a theoretical description of genetic regulatory systems in bacteria that explicitly considers the cell division cycle to investigate its impact on gene expression. We calculate the cell-to-cell variations arising from cells being at different stages in the division cycle for unregulated genes and for basic regulatory mechanisms. These variations contribute to the extrinsic noise observed in single-cell experiments, and are most significant for proteins with short lifetimes. Negative autoregulation buffers against variation of protein concentration over the division cycle, but the effect is found to be relatively weak. Stronger buffering is achieved by an increased protein lifetime. Positive autoregulation can strongly amplify such variation if the parameters are set to values that lead to resonance-like behaviour. For cooperative positive autoregulation, the concentration variation over the division cycle diminishes the parameter region of bistability and modulates the switching times between the two stable states. The same effects are seen for a two-gene mutual-repression toggle switch. By contrast, an oscillatory circuit, the repressilator, is only weakly affected by the division cycle.

  8. Impact of gene stacking on gene flow: the case of maize.

    PubMed

    Paul, Lénaïc; Angevin, Frédérique; Collonnier, Cécile; Messéan, Antoine

    2012-04-01

    To respect the European labelling threshold for the adventitious presence of genetically modified organisms (GMOs) in food and feed, stakeholders mainly rely on real-time PCR analysis, which provides a measurement expressed as a percentage of GM-DNA. However, this measurement veils the complexity of gene flow, especially in the case of gene stacking. We have investigated the impact of gene stacking on adventitious GM presence due to pollen flow and seed admixture as well as its translation in terms of the percentage of GM-DNA in a non-GM maize harvest. In the case of varieties bearing one to four stacked events, we established a set of relationships between the percentage of GM kernels and the percentage of GM-DNA in a non-GM harvest as well as a set of relationships between the rate of seed admixture and the percentages of GM material in a non-GM harvest. Thanks to these relationships, and based on simulations with a gene flow model, we have been able to demonstrate that the number of events and the stacking structure of the emitting fields impact the ability of a non-GM maize producer to comply with given GM kernel or GM-DNA thresholds. We also show that a great variability in the rates of GM kernels, embryos and DNA results from seed admixture. Finally, the choice of a unit of measurement for a GM threshold in seed lots can have opposite effects on the ability of farmers to comply with a given threshold depending on whether they are crop or seed producers.

  9. Impact of gene editing on the study of cystic fibrosis.

    PubMed

    Harrison, Patrick T; Sanz, David J; Hollywood, Jennifer A

    2016-09-01

    Cystic fibrosis (CF) is a chronic and progressive autosomal recessive disorder of secretory epithelial cells, which causes obstructions in the lung airways and pancreatic ducts of 70,000 people worldwide (for recent review see Cutting Nat Rev Genet 16(1):45-56, 2015). The finding that mutations in the CFTR gene cause CF (Kerem et al. Science 245(4922):1073-1080, 1989; Riordan et al. Science 245(4922):1066-1073, 1989; Rommens et al. Science 245(4922):1059-1065, 1989), was hailed as the very happy middle of a story whose end is a cure for a fatal disease (Koshland Science 245(4922):1029, 1989). However, despite two licensed drugs (Ramsey et al. N Engl J Med 365(18):1663-1672, 2011; Wainwright et al. N Engl J Med 373(3):220-231, 2015), and a formal demonstration that repeated administration of CFTR cDNA to patients is safe and effects a modest but significant stabilisation of disease (Alton et al. Lancet Respir Med 3(9):684-691, 2015), we are still a long way from a cure, with many patients taking over 100 tablets per day, and a mean age at death of 28 years. The aim of this review is to discuss the impact on the study of CF of gene-editing techniques as they have developed over the last 30 years, up to and including the possibility of editing as a therapeutic approach.

  10. Changes in chondrocyte gene expression following in vitro impaction of porcine articular cartilage in an impact injury model.

    PubMed

    Ashwell, Melissa S; Gonda, Michael G; Gray, Kent; Maltecca, Christian; O'Nan, Audrey T; Cassady, Joseph P; Mente, Peter L

    2013-03-01

    Our objective was to monitor chondrocyte gene expression at 0, 3, 7, and 14 days following in vitro impaction to the articular surface of porcine patellae. Patellar facets were either axially impacted with a cylindrical impactor (25 mm/s loading rate) to a load level of 2,000 N or not impacted to serve as controls. After being placed in organ culture for 0, 3, 7, or 14 days, total RNA was isolated from full thickness cartilage slices and gene expression measured for 17 genes by quantitative real-time RT-PCR. Targeted genes included those encoding proteins involved with biological stress, inflammation, or anabolism and catabolism of cartilage extracellular matrix. Some gene expression changes were detected on the day of impaction, but most significant changes occurred at 14 days in culture. At 14 days in culture, 10 of the 17 genes were differentially expressed with col1a1 most significantly up-regulated in the impacted samples, suggesting impacted chondrocytes may have reverted to a fibroblast-like phenotype.

  11. Impact of recurrent gene duplication on adaptation of plant genomes

    PubMed Central

    2014-01-01

    Background Recurrent gene duplication and retention played an important role in angiosperm genome evolution. It has been hypothesized that these processes contribute significantly to plant adaptation but so far this hypothesis has not been tested at the genome scale. Results We studied available sequenced angiosperm genomes to assess the frequency of positive selection footprints in lineage specific expanded (LSE) gene families compared to single-copy genes using a dN/dS-based test in a phylogenetic framework. We found 5.38% of alignments in LSE genes with codons under positive selection. In contrast, we found no evidence for codons under positive selection in the single-copy reference set. An analysis at the branch level shows that purifying selection acted more strongly on single-copy genes than on LSE gene clusters. Moreover we detect significantly more branches indicating evolution under positive selection and/or relaxed constraint in LSE genes than in single-copy genes. Conclusions In this – to our knowledge –first genome-scale study we provide strong empirical support for the hypothesis that LSE genes fuel adaptation in angiosperms. Our conservative approach for detecting selection footprints as well as our results can be of interest for further studies on (plant) gene family evolution. PMID:24884640

  12. Potential impact of environmental bacteriophages in spreading antibiotic resistance genes.

    PubMed

    Muniesa, Maite; Colomer-Lluch, Marta; Jofre, Juan

    2013-06-01

    The idea that bacteriophage transduction plays a role in the horizontal transfer of antibiotic resistance genes is gaining momentum. Such transduction might be vital in horizontal transfer from environmental to human body-associated biomes and here we review many lines of evidence supporting this notion. It is well accepted that bacteriophages are the most abundant entities in most environments, where they have been shown to be quite persistent. This fact, together with the ability of many phages to infect bacteria belonging to different taxa, makes them suitable vehicles for gene transfer. Metagenomic studies confirm that substantial percentages of the bacteriophage particles present in most environments contain bacterial genes, including mobile genetic elements and antibiotic resistance genes. When specific genes of resistance to antibiotics are detected by real-time PCR in the bacteriophage populations of different environments, only tenfold lower numbers of these genes are observed, compared with those found in the corresponding bacterial populations. In addition, the antibiotic resistance genes from these bacteriophages are functional and generate resistance to the bacteria when these genes are transfected. Finally, reports about the transduction of antibiotic resistance genes are on the increase.

  13. Identification and Validation of Reference Genes and Their Impact on Normalized Gene Expression Studies across Cultivated and Wild Cicer Species.

    PubMed

    Reddy, Dumbala Srinivas; Bhatnagar-Mathur, Pooja; Reddy, Palakolanu Sudhakar; Sri Cindhuri, Katamreddy; Sivaji Ganesh, Adusumalli; Sharma, Kiran Kumar

    2016-01-01

    Quantitative Real-Time PCR (qPCR) is a preferred and reliable method for accurate quantification of gene expression to understand precise gene functions. A total of 25 candidate reference genes including traditional and new generation reference genes were selected and evaluated in a diverse set of chickpea samples. The samples used in this study included nine chickpea genotypes (Cicer spp.) comprising of cultivated and wild species, six abiotic stress treatments (drought, salinity, high vapor pressure deficit, abscisic acid, cold and heat shock), and five diverse tissues (leaf, root, flower, seedlings and seed). The geNorm, NormFinder and RefFinder algorithms used to identify stably expressed genes in four sample sets revealed stable expression of UCP and G6PD genes across genotypes, while TIP41 and CAC were highly stable under abiotic stress conditions. While PP2A and ABCT genes were ranked as best for different tissues, ABCT, UCP and CAC were most stable across all samples. This study demonstrated the usefulness of new generation reference genes for more accurate qPCR based gene expression quantification in cultivated as well as wild chickpea species. Validation of the best reference genes was carried out by studying their impact on normalization of aquaporin genes PIP1;4 and TIP3;1, in three contrasting chickpea genotypes under high vapor pressure deficit (VPD) treatment. The chickpea TIP3;1 gene got significantly up regulated under high VPD conditions with higher relative expression in the drought susceptible genotype, confirming the suitability of the selected reference genes for expression analysis. This is the first comprehensive study on the stability of the new generation reference genes for qPCR studies in chickpea across species, different tissues and abiotic stresses.

  14. Identification and Validation of Reference Genes and Their Impact on Normalized Gene Expression Studies across Cultivated and Wild Cicer Species

    PubMed Central

    Reddy, Palakolanu Sudhakar; Sri Cindhuri, Katamreddy; Sivaji Ganesh, Adusumalli; Sharma, Kiran Kumar

    2016-01-01

    Quantitative Real-Time PCR (qPCR) is a preferred and reliable method for accurate quantification of gene expression to understand precise gene functions. A total of 25 candidate reference genes including traditional and new generation reference genes were selected and evaluated in a diverse set of chickpea samples. The samples used in this study included nine chickpea genotypes (Cicer spp.) comprising of cultivated and wild species, six abiotic stress treatments (drought, salinity, high vapor pressure deficit, abscisic acid, cold and heat shock), and five diverse tissues (leaf, root, flower, seedlings and seed). The geNorm, NormFinder and RefFinder algorithms used to identify stably expressed genes in four sample sets revealed stable expression of UCP and G6PD genes across genotypes, while TIP41 and CAC were highly stable under abiotic stress conditions. While PP2A and ABCT genes were ranked as best for different tissues, ABCT, UCP and CAC were most stable across all samples. This study demonstrated the usefulness of new generation reference genes for more accurate qPCR based gene expression quantification in cultivated as well as wild chickpea species. Validation of the best reference genes was carried out by studying their impact on normalization of aquaporin genes PIP1;4 and TIP3;1, in three contrasting chickpea genotypes under high vapor pressure deficit (VPD) treatment. The chickpea TIP3;1 gene got significantly up regulated under high VPD conditions with higher relative expression in the drought susceptible genotype, confirming the suitability of the selected reference genes for expression analysis. This is the first comprehensive study on the stability of the new generation reference genes for qPCR studies in chickpea across species, different tissues and abiotic stresses. PMID:26863232

  15. Adaptive impact of the chimeric gene Quetzalcoatl in Drosophila melanogaster.

    PubMed

    Rogers, Rebekah L; Bedford, Trevor; Lyons, Ana M; Hartl, Daniel L

    2010-06-15

    Chimeric genes, which form through the genomic fusion of two protein-coding genes, are a significant source of evolutionary novelty in Drosophila melanogaster. However, the propensity of chimeric genes to produce adaptive phenotypic changes is not fully understood. Here, we describe the chimeric gene Quetzalcoatl (Qtzl; CG31864), which formed in the recent past and swept to fixation in D. melanogaster. Qtzl arose through a duplication on chromosome 2L that united a portion of the mitochondrially targeted peptide CG12264 with a segment of the polycomb gene escl. The 3' segment of the gene, which is derived from escl, is inherited out of frame, producing a unique peptide sequence. Nucleotide diversity is drastically reduced and site frequency spectra are significantly skewed surrounding the duplicated region, a finding consistent with a selective sweep on the duplicate region containing Qtzl. Qtzl has an expression profile that largely resembles that of escl, with expression in early pupae, adult females, and male testes. However, expression patterns appear to have been decoupled from both parental genes during later embryonic development and in head tissues of adult males, indicating that Qtzl has developed a distinct regulatory profile through the rearrangement of different 5' and 3' regulatory domains. Furthermore, misexpression of Qtzl suppresses defects in the formation of the neuromuscular junction in larvae, demonstrating that Qtzl can produce phenotypic effects in cells. Together, these results show that chimeric genes can produce structural and regulatory changes in a single mutational step and may be a major factor in adaptive evolution.

  16. Gene expression allelic imbalance in ovine brown adipose tissue impacts energy homeostasis.

    PubMed

    Ghazanfar, Shila; Vuocolo, Tony; Morrison, Janna L; Nicholas, Lisa M; McMillen, Isabella C; Yang, Jean Y H; Buckley, Michael J; Tellam, Ross L

    2017-01-01

    Heritable trait variation within a population of organisms is largely governed by DNA variations that impact gene transcription and protein function. Identifying genetic variants that affect complex functional traits is a primary aim of population genetics studies, especially in the context of human disease and agricultural production traits. The identification of alleles directly altering mRNA expression and thereby biological function is challenging due to difficulty in isolating direct effects of cis-acting genetic variations from indirect trans-acting genetic effects. Allele specific gene expression or allelic imbalance in gene expression (AI) occurring at heterozygous loci provides an opportunity to identify genes directly impacted by cis-acting genetic variants as indirect trans-acting effects equally impact the expression of both alleles. However, the identification of genes showing AI in the context of the expression of all genes remains a challenge due to a variety of technical and statistical issues. The current study focuses on the discovery of genes showing AI using single nucleotide polymorphisms as allelic reporters. By developing a computational and statistical process that addressed multiple analytical challenges, we ranked 5,809 genes for evidence of AI using RNA-Seq data derived from brown adipose tissue samples from a cohort of late gestation fetal lambs and then identified a conservative subgroup of 1,293 genes. Thus, AI was extensive, representing approximately 25% of the tested genes. Genes associated with AI were enriched for multiple Gene Ontology (GO) terms relating to lipid metabolism, mitochondrial function and the extracellular matrix. These functions suggest that cis-acting genetic variations causing AI in the population are preferentially impacting genes involved in energy homeostasis and tissue remodelling. These functions may contribute to production traits likely to be under genetic selection in the population.

  17. Gene expression allelic imbalance in ovine brown adipose tissue impacts energy homeostasis

    PubMed Central

    Ghazanfar, Shila; Vuocolo, Tony; Morrison, Janna L.; Nicholas, Lisa M.; McMillen, Isabella C.; Yang, Jean Y. H.; Buckley, Michael J.

    2017-01-01

    Heritable trait variation within a population of organisms is largely governed by DNA variations that impact gene transcription and protein function. Identifying genetic variants that affect complex functional traits is a primary aim of population genetics studies, especially in the context of human disease and agricultural production traits. The identification of alleles directly altering mRNA expression and thereby biological function is challenging due to difficulty in isolating direct effects of cis-acting genetic variations from indirect trans-acting genetic effects. Allele specific gene expression or allelic imbalance in gene expression (AI) occurring at heterozygous loci provides an opportunity to identify genes directly impacted by cis-acting genetic variants as indirect trans-acting effects equally impact the expression of both alleles. However, the identification of genes showing AI in the context of the expression of all genes remains a challenge due to a variety of technical and statistical issues. The current study focuses on the discovery of genes showing AI using single nucleotide polymorphisms as allelic reporters. By developing a computational and statistical process that addressed multiple analytical challenges, we ranked 5,809 genes for evidence of AI using RNA-Seq data derived from brown adipose tissue samples from a cohort of late gestation fetal lambs and then identified a conservative subgroup of 1,293 genes. Thus, AI was extensive, representing approximately 25% of the tested genes. Genes associated with AI were enriched for multiple Gene Ontology (GO) terms relating to lipid metabolism, mitochondrial function and the extracellular matrix. These functions suggest that cis-acting genetic variations causing AI in the population are preferentially impacting genes involved in energy homeostasis and tissue remodelling. These functions may contribute to production traits likely to be under genetic selection in the population. PMID:28665992

  18. Adaptive impact of the chimeric gene Quetzalcoatl in Drosophila melanogaster

    PubMed Central

    Rogers, Rebekah L.; Bedford, Trevor; Lyons, Ana M.; Hartl, Daniel L.

    2010-01-01

    Chimeric genes, which form through the genomic fusion of two protein-coding genes, are a significant source of evolutionary novelty in Drosophila melanogaster. However, the propensity of chimeric genes to produce adaptive phenotypic changes is not fully understood. Here, we describe the chimeric gene Quetzalcoatl (Qtzl; CG31864), which formed in the recent past and swept to fixation in D. melanogaster. Qtzl arose through a duplication on chromosome 2L that united a portion of the mitochondrially targeted peptide CG12264 with a segment of the polycomb gene escl. The 3′ segment of the gene, which is derived from escl, is inherited out of frame, producing a unique peptide sequence. Nucleotide diversity is drastically reduced and site frequency spectra are significantly skewed surrounding the duplicated region, a finding consistent with a selective sweep on the duplicate region containing Qtzl. Qtzl has an expression profile that largely resembles that of escl, with expression in early pupae, adult females, and male testes. However, expression patterns appear to have been decoupled from both parental genes during later embryonic development and in head tissues of adult males, indicating that Qtzl has developed a distinct regulatory profile through the rearrangement of different 5′ and 3′ regulatory domains. Furthermore, misexpression of Qtzl suppresses defects in the formation of the neuromuscular junction in larvae, demonstrating that Qtzl can produce phenotypic effects in cells. Together, these results show that chimeric genes can produce structural and regulatory changes in a single mutational step and may be a major factor in adaptive evolution. PMID:20534482

  19. Dietary intervention impact on gut microbial gene richness.

    PubMed

    Cotillard, Aurélie; Kennedy, Sean P; Kong, Ling Chun; Prifti, Edi; Pons, Nicolas; Le Chatelier, Emmanuelle; Almeida, Mathieu; Quinquis, Benoit; Levenez, Florence; Galleron, Nathalie; Gougis, Sophie; Rizkalla, Salwa; Batto, Jean-Michel; Renault, Pierre; Doré, Joel; Zucker, Jean-Daniel; Clément, Karine; Ehrlich, Stanislav Dusko

    2013-08-29

    Complex gene-environment interactions are considered important in the development of obesity. The composition of the gut microbiota can determine the efficacy of energy harvest from food and changes in dietary composition have been associated with changes in the composition of gut microbial populations. The capacity to explore microbiota composition was markedly improved by the development of metagenomic approaches, which have already allowed production of the first human gut microbial gene catalogue and stratifying individuals by their gut genomic profile into different enterotypes, but the analyses were carried out mainly in non-intervention settings. To investigate the temporal relationships between food intake, gut microbiota and metabolic and inflammatory phenotypes, we conducted diet-induced weight-loss and weight-stabilization interventions in a study sample of 38 obese and 11 overweight individuals. Here we report that individuals with reduced microbial gene richness (40%) present more pronounced dys-metabolism and low-grade inflammation, as observed concomitantly in the accompanying paper. Dietary intervention improves low gene richness and clinical phenotypes, but seems to be less efficient for inflammation variables in individuals with lower gene richness. Low gene richness may therefore have predictive potential for the efficacy of intervention.

  20. High-resolution genome-wide scan of genes, gene-networks and cellular systems impacting the yeast ionome

    USDA-ARS?s Scientific Manuscript database

    To balance the demand for uptake of essential elements with their potential toxicity living cells have complex regulatory mechanisms. Here, we describe a genome-wide screen to identify genes that impact the elemental composition (‘ionome’) of yeast Saccharomyces cerevisiae. Using inductively coupled...

  1. On the origins of Mendelian disease genes in man: the impact of gene duplication.

    PubMed

    Dickerson, Jonathan E; Robertson, David L

    2012-01-01

    Over 3,000 human diseases are known to be linked to heritable genetic variation, mapping to over 1,700 unique genes. Dating of the evolutionary age of these disease-associated genes has suggested that they have a tendency to be ancient, specifically coming into existence with early metazoa. The approach taken by past studies, however, assumes that the age of a disease is the same as the age of its common ancestor, ignoring the fundamental contribution of duplication events in the evolution of new genes and function. Here, we date both the common ancestor and the duplication history of known human disease-associated genes. We find that the majority of disease genes (80%) are genes that have been duplicated in their evolutionary history. Periods for which there are more disease-associated genes, for example, at the origins of bony vertebrates, are explained by the emergence of more genes at that time, and the majority of these are duplicates inferred to have arisen by whole-genome duplication. These relationships are similar for different disease types and the disease-associated gene's cellular function. This indicates that the emergence of duplication-associated diseases has been ongoing and approximately constant (relative to the retention of duplicate genes) throughout the evolution of life. This continued until approximately 390 Ma from which time relatively fewer novel genes came into existence on the human lineage, let alone disease genes. For single-copy genes associated with disease, we find that the numbers of disease genes decreases with recency. For the majority of duplicates, the disease-associated mutation is associated with just one of the duplicate copies. A universal explanation for heritable disease is, thus, that it is merely a by-product of the evolutionary process; the evolution of new genes (de novo or by duplication) results in the potential for new diseases to emerge.

  2. Gene Expression Elucidates Functional Impact of Polygenic Risk for Schizophrenia

    PubMed Central

    Fromer, Menachem; Roussos, Panos; Sieberts, Solveig K; Johnson, Jessica S; Kavanagh, David H; Perumal, Thanneer M; Ruderfer, Douglas M; Oh, Edwin C; Topol, Aaron; Shah, Hardik R; Klei, Lambertus L; Kramer, Robin; Pinto, Dalila; Gümüş, Zeynep H; Cicek, A. Ercument; Dang, Kristen K; Browne, Andrew; Lu, Cong; Xie, Lu; Readhead, Ben; Stahl, Eli A; Parvizi, Mahsa; Hamamsy, Tymor; Fullard, John F; Wang, Ying-Chih; Mahajan, Milind C; Derry, Jonathan M J; Dudley, Joel; Hemby, Scott E; Logsdon, Benjamin A; Talbot, Konrad; Raj, Towfique; Bennett, David A; De Jager, Philip L; Zhu, Jun; Zhang, Bin; Sullivan, Patrick F; Chess, Andrew; Purcell, Shaun M; Shinobu, Leslie A; Mangravite, Lara M; Toyoshiba, Hiroyoshi; Gur, Raquel E; Hahn, Chang-Gyu; Lewis, David A; Haroutunian, Vahram; Peters, Mette A; Lipska, Barbara K; Buxbaum, Joseph D; Schadt, Eric E; Hirai, Keisuke; Roeder, Kathryn; Brennand, Kristen J; Katsanis, Nicholas; Domenici, Enrico; Devlin, Bernie; Sklar, Pamela

    2016-01-01

    Over 100 genetic loci harbor schizophrenia associated variants, yet how these variants confer liability is uncertain. The CommonMind Consortium sequenced RNA from dorsolateral prefrontal cortex of schizophrenia cases (N = 258) and control subjects (N = 279), creating a resource of gene expression and its genetic regulation. Using this resource, ~20% of schizophrenia loci have variants that could contribute to altered gene expression and liability. In five loci, only a single gene was involved: FURIN, TSNARE1, CNTN4, CLCN3, or SNAP91. Altering expression of FURIN, TSNARE1, or CNTN4 changes neurodevelopment in zebrafish; knockdown of FURIN in human neural progenitor cells yields abnormal migration. Of 693 genes showing significant case/control differential expression, their fold changes are ≤ 1.33, and an independent cohort yields similar results. Gene co-expression implicates a network relevant for schizophrenia. Our findings show schizophrenia is polygenic and highlight the utility of this resource for mechanistic interpretations of genetic liability for brain diseases. PMID:27668389

  3. Impact of Solar Radiation on Gene Expression in Bacteria

    PubMed Central

    Matallana-Surget, Sabine; Wattiez, Ruddy

    2013-01-01

    Microorganisms often regulate their gene expression at the level of transcription and/or translation in response to solar radiation. In this review, we present the use of both transcriptomics and proteomics to advance knowledge in the field of bacterial response to damaging radiation. Those studies pertain to diverse application areas such as fundamental microbiology, water treatment, microbial ecology and astrobiology. Even though it has been demonstrated that mRNA abundance is not always consistent with the protein regulation, we present here an exhaustive review on how bacteria regulate their gene expression at both transcription and translation levels to enable biomarkers identification and comparison of gene regulation from one bacterial species to another. PMID:28250399

  4. The Impact of Human Gene Patents on Genetic Testing in the UK

    PubMed Central

    Hawkins, Naomi

    2011-01-01

    This paper reports the results of an empirical study examining the impact of human gene patents on the development and delivery of genetic tests in the public sector in the UK. The study found that, despite the potential for gene patents to have significant negative consequences for genetic testing, in fact, human gene patents have little or no impact on practice for those developing genetic tests in the public sector in the UK. This is not because patents are managed optimally; rather, gene patents are essentially ignored. This paper reports the factors that motivate this behavior. At least insofar as there seems to be no apparent problem of lack of patient access, there is no significant public health problem. However, there is divergence between the legal and the practical situation. Complacency about the lack of impact of patents on access to diagnostics is risky, and concerns about patents should be addressed proactively, rather than reactively. PMID:21150786

  5. Impact of physical activity and doping on epigenetic gene regulation.

    PubMed

    Schwarzenbach, Heidi

    2011-10-01

    To achieve success in sports, many athletes consume doping substances, such as anabolic androgenic steroids and growth hormones, and ignore the negative influence of these drugs on their health. Apart from the unethical aspect of doping in sports, it is essential to consider the tremendous risk it represents to their physical condition. The abuse of pharmaceuticals which improve athletic performance may alter the expression of specific genes involved in muscle and bone metabolism by epigenetic mechanisms, such as DNA methylation and histone modifications. Moreover, excessive and relentless training to increase the muscle mass, may also have an influence on the health of the athletes. This stress releases neurotransmitters and growth factors, and may affect the expression of endogenous genes by DNA methylation, too. This paper focuses on the relationship between epigenetic mechanisms and sports, highlights the potential consequences of abuse of doping drugs on gene expression, and describes methods to molecularly detect epigenetic changes of gene markers reflecting the physiological or metabolic effects of doping agents.

  6. The impact of rare variation on gene expression across tissues.

    PubMed

    Li, Xin; Kim, Yungil; Tsang, Emily K; Davis, Joe R; Damani, Farhan N; Chiang, Colby; Hess, Gaelen T; Zappala, Zachary; Strober, Benjamin J; Scott, Alexandra J; Li, Amy; Ganna, Andrea; Bassik, Michael C; Merker, Jason D; Hall, Ira M; Battle, Alexis; Montgomery, Stephen B

    2017-10-11

    Rare genetic variants are abundant in humans and are expected to contribute to individual disease risk. While genetic association studies have successfully identified common genetic variants associated with susceptibility, these studies are not practical for identifying rare variants. Efforts to distinguish pathogenic variants from benign rare variants have leveraged the genetic code to identify deleterious protein-coding alleles, but no analogous code exists for non-coding variants. Therefore, ascertaining which rare variants have phenotypic effects remains a major challenge. Rare non-coding variants have been associated with extreme gene expression in studies using single tissues, but their effects across tissues are unknown. Here we identify gene expression outliers, or individuals showing extreme expression levels for a particular gene, across 44 human tissues by using combined analyses of whole genomes and multi-tissue RNA-sequencing data from the Genotype-Tissue Expression (GTEx) project v6p release. We find that 58% of underexpression and 28% of overexpression outliers have nearby conserved rare variants compared to 8% of non-outliers. Additionally, we developed RIVER (RNA-informed variant effect on regulation), a Bayesian statistical model that incorporates expression data to predict a regulatory effect for rare variants with higher accuracy than models using genomic annotations alone. Overall, we demonstrate that rare variants contribute to large gene expression changes across tissues and provide an integrative method for interpretation of rare variants in individual genomes.

  7. Impact of ACTH Signaling on Transcriptional Regulation of Steroidogenic Genes

    PubMed Central

    Ruggiero, Carmen; Lalli, Enzo

    2016-01-01

    The trophic peptide hormone adrenocorticotropic (ACTH) stimulates steroid hormone biosynthesis evoking both a rapid, acute response and a long-term, chronic response, via the activation of cAMP/protein kinase A (PKA) signaling. The acute response is initiated by the mobilization of cholesterol from lipid stores and its delivery to the inner mitochondrial membrane, a process that is mediated by the steroidogenic acute regulatory protein. The chronic response results in the increased coordinated transcription of genes encoding steroidogenic enzymes. ACTH binding to its cognate receptor, melanocortin 2 receptor (MC2R), stimulates adenylyl cyclase, thus inducing cAMP production, PKA activation, and phosphorylation of specific nuclear factors, which bind to target promoters and facilitate coactivator protein recruitment to direct steroidogenic gene transcription. This review provides a general view of the transcriptional control exerted by the ACTH/cAMP system on the expression of genes encoding for steroidogenic enzymes in the adrenal cortex. Special emphasis will be given to the transcription factors required to mediate ACTH-dependent transcription of steroidogenic genes. PMID:27065945

  8. Estrogen Signaling Multiple Pathways to Impact Gene Transcription

    PubMed Central

    Marino, Maria; Galluzzo, Paola; Ascenzi, Paolo

    2006-01-01

    Steroid hormones exert profound effects on cell growth, development, differentiation, and homeostasis. Their effects are mediated through specific intracellular steroid receptors that act via multiple mechanisms. Among others, the action mechanism starting upon 17β-estradiol (E2) binds to its receptors (ER) is considered a paradigmatic example of how steroid hormones function. Ligand-activated ER dimerizes and translocates in the nucleus where it recognizes specific hormone response elements located in or near promoter DNA regions of target genes. Behind the classical genomic mechanism shared with other steroid hormones, E2 also modulates gene expression by a second indirect mechanism that involves the interaction of ER with other transcription factors which, in turn, bind their cognate DNA elements. In this case, ER modulates the activities of transcription factors such as the activator protein (AP)-1, nuclear factor-κB (NF-κB) and stimulating protein-1 (Sp-1), by stabilizing DNA-protein complexes and/or recruiting co-activators. In addition, E2 binding to ER may also exert rapid actions that start with the activation of a variety of signal transduction pathways (e.g. ERK/MAPK, p38/MAPK, PI3K/AKT, PLC/PKC). The debate about the contribution of different ER-mediated signaling pathways to coordinate the expression of specific sets of genes is still open. This review will focus on the recent knowledge about the mechanism by which ERs regulate the expression of target genes and the emerging field of integration of membrane and nuclear receptor signaling, giving examples of the ways by which the genomic and non-genomic actions of ERs on target genes converge. PMID:18369406

  9. Impact of obesity-related genes in Spanish population

    PubMed Central

    2013-01-01

    Background The objective was to investigate the association between BMI and single nucleotide polymorphisms previously identified of obesity-related genes in two Spanish populations. Forty SNPs in 23 obesity-related genes were evaluated in a rural population characterized by a high prevalence of obesity (869 subjects, mean age 46 yr, 62% women, 36% obese) and in an urban population (1425 subjects, mean age 54 yr, 50% women, 19% obese). Genotyping was assessed by using SNPlex and PLINK for the association analysis. Results Polymorphisms of the FTO were significantly associated with BMI, in the rural population (beta 0.87, p-value <0.001). None of the other SNPs showed significant association after Bonferroni correction in the two populations or in the pooled analysis. A weighted genetic risk score (wGRS) was constructed using the risk alleles of the Tag-SNPs with a positive Beta parameter in both populations. From the first to the fifth quintile of the score, the BMI increased 0.45 kg/m2 in Hortega and 2.0 kg/m2 in Pizarra. Overall, the obesity predictive value was low (less than 1%). Conclusion The risk associated with polymorphisms is low and the overall effect on BMI or obesity prediction is minimal. A weighted genetic risk score based on genes mainly acting through central nervous system mechanisms was associated with BMI but it yields minimal clinical prediction for the obesity risk in the general population. PMID:24267414

  10. Impact of experience-dependent and -independent factors on gene expression in songbird brain

    PubMed Central

    Drnevich, Jenny; Replogle, Kirstin L.; Lovell, Peter; Hahn, Thomas P.; Johnson, Frank; Mast, Thomas G.; Nordeen, Ernest; Nordeen, Kathy; Strand, Christy; London, Sarah E.; Mukai, Motoko; Wingfield, John C.; Arnold, Arthur P.; Ball, Gregory F.; Brenowitz, Eliot A.; Wade, Juli; Mello, Claudio V.; Clayton, David F.

    2012-01-01

    Songbirds provide rich natural models for studying the relationships between brain anatomy, behavior, environmental signals, and gene expression. Under the Songbird Neurogenomics Initiative, investigators from 11 laboratories collected brain samples from six species of songbird under a range of experimental conditions, and 488 of these samples were analyzed systematically for gene expression by microarray. ANOVA was used to test 32 planned contrasts in the data, revealing the relative impact of different factors. The brain region from which tissue was taken had the greatest influence on gene expression profile, affecting the majority of signals measured by 18,848 cDNA spots on the microarray. Social and environmental manipulations had a highly variable impact, interpreted here as a manifestation of paradoxical “constitutive plasticity” (fewer inducible genes) during periods of enhanced behavioral responsiveness. Several specific genes were identified that may be important in the evolution of linkages between environmental signals and behavior. The data were also analyzed using weighted gene coexpression network analysis, followed by gene ontology analysis. This revealed modules of coexpressed genes that are also enriched for specific functional annotations, such as “ribosome” (expressed more highly in juvenile brain) and “dopamine metabolic process” (expressed more highly in striatal song control nucleus area X). These results underscore the complexity of influences on neural gene expression and provide a resource for studying how these influences are integrated during natural experience. PMID:23045667

  11. Clinical impact of recurrently mutated genes on lymphoma diagnostics: state-of-the-art and beyond.

    PubMed

    Rosenquist, Richard; Rosenwald, Andreas; Du, Ming-Qing; Gaidano, Gianluca; Groenen, Patricia; Wotherspoon, Andrew; Ghia, Paolo; Gaulard, Philippe; Campo, Elias; Stamatopoulos, Kostas

    2016-09-01

    Similar to the inherent clinical heterogeneity of most, if not all, lymphoma entities, the genetic landscape of these tumors is markedly complex in the majority of cases, with a rapidly growing list of recurrently mutated genes discovered in recent years by next-generation sequencing technology. Whilst a few genes have been implied to have diagnostic, prognostic and even predictive impact, most gene mutations still require rigorous validation in larger, preferably prospective patient series, to scrutinize their potential role in lymphoma diagnostics and patient management. In selected entities, a predominantly mutated gene is identified in almost all cases (e.g. Waldenström's macroglobulinemia/lymphoplasmacytic lymphoma and hairy-cell leukemia), while for the vast majority of lymphomas a quite diverse mutation pattern is observed, with a limited number of frequently mutated genes followed by a seemingly endless tail of genes with mutations at a low frequency. Herein, the European Expert Group on NGS-based Diagnostics in Lymphomas (EGNL) summarizes the current status of this ever-evolving field, and, based on the present evidence level, segregates mutations into the following categories: i) immediate impact on treatment decisions, ii) diagnostic impact, iii) prognostic impact, iv) potential clinical impact in the near future, or v) should only be considered for research purposes. In the coming years, coordinated efforts aiming to apply targeted next-generation sequencing in large patient series will be needed in order to elucidate if a particular gene mutation will have an immediate impact on the lymphoma classification, and ultimately aid clinical decision making. Copyright© Ferrata Storti Foundation.

  12. Gene flow, invasiveness, and ecological impact of genetically modified crops.

    PubMed

    Warwick, Suzanne I; Beckie, Hugh J; Hall, Linda M

    2009-06-01

    The main environmental concerns about genetically modified (GM) crops are the potential weediness or invasiveness in the crop itself or in its wild or weedy relatives as a result of transgene movement. Here we briefly review evidence for pollen- and seed-mediated gene flow from GM crops to non-GM or other GM crops and to wild relatives. The report focuses on the effect of abiotic and biotic stress-tolerance traits on plant fitness and their potential to increase weedy or invasive tendencies. An evaluation of weediness and invasive traits that contribute to the success of agricultural weeds and invasive plants was of limited value in predicting the effect of biotic and abiotic stress-tolerance GM traits, suggesting context-specific evaluation rather than generalizations. Fitness data on herbicide, insect, and disease resistance, as well as cold-, drought-, and salinity-tolerance traits, are reviewed. We describe useful ecological models predicting the effects of gene flow and altered fitness in GM crops and wild/weedy relatives, as well as suitable mitigation measures. A better understanding of factors controlling population size, dynamics, and range limits in weedy volunteer GM crop and related host or target weed populations is necessary before the effect of biotic and abiotic stress-tolerance GM traits can be fully assessed.

  13. Impact of gene patents on the development of molecular diagnostics.

    PubMed

    Toneguzzo, Frances

    2011-07-01

    There is a widely held view that gene patents in particular and patents in general, because of the exclusionary rights that they provide, are inhibiting the development of and access to critical molecular diagnostic testing. This is a highly relevant issue for healthcare delivery as we move towards personalized medicine, which relies heavily on genetic testing to tailor treatments that are specific for individual characteristics. Critics of the patent system hope to void or diminish the exclusionary aspect of patents by removing genes from the definition of what is patentable, by increasing the number of activities that fall within the research use exemption, or by compelling patent holders to license their rights non-exclusively. Although a re-examination of what constitutes patentable subject matter is an important undertaking, narrowing the definition of patentable subject matter is at best only a partial solution. Erosion of the patent system through compulsory licensing or expansion of the research use exemption runs the risk of destroying important incentives without also fully addressing the problem. To promote solutions that truly address the issues, this article distinguishes documented facts from perceptions and suggests alternative approaches to explore. The author believes that efforts to undermine the patent system are simply counterproductive and that time would be better spent addressing the real issues that lie within molecular diagnostic development.

  14. Knowing your genes: does this impact behaviour change?

    PubMed

    O'Donovan, Clare B; Walsh, Marianne C; Gibney, Michael J; Brennan, Lorraine; Gibney, Eileen R

    2017-08-01

    It is postulated that knowledge of genotype may be more powerful than other types of personalised information in terms of motivating behaviour change. However, there is also a danger that disclosure of genetic risk may promote a fatalistic attitude and demotivate individuals. The original concept of personalised nutrition (PN) focused on genotype-based tailored dietary advice; however, PN can also be delivered based on assessment of dietary intake and phenotypic measures. Whilst dietitians currently provide PN advice based on diet and phenotype, genotype-based PN advice is not so readily available. The aim of this review is to examine the evidence for genotype-based personalised information on motivating behaviour change, and factors which may affect the impact of genotype-based personalised advice. Recent findings in PN will also be discussed, with respect to a large European study, Food4Me, which investigated the impact of varying levels of PN advice on motivating behaviour change. The researchers reported that PN advice resulted in greater dietary changes compared with general healthy eating advice, but no additional benefit was observed for PN advice based on phenotype and genotype information. Within Food4Me, work from our group revealed that knowledge of MTHFR genotype did not significantly improve intakes of dietary folate. In general, evidence is weak with regard to genotype-based PN advice. For future work, studies should test the impact of PN advice developed on a strong nutrigenetic evidence base, ensure an appropriate study design for the research question asked, and incorporate behaviour change techniques into the intervention.

  15. Impact of lux gene insertion on bacterial surface properties and transport.

    PubMed

    Chen, Gang; Srinivasa Ranga, Vijay Penagonda; Mao, Yongjun; Chen, Kevin; Qiao, Hanzi

    2008-03-01

    Genetic markers have been in popular use for tracing microbial movement in the environment. However, the impact of genetic marker insertion on microbial surface properties and consequent transport is often ignored. For this research, we investigated the impact of luminescence-based genetic marker insertion on bacterial surface properties and transport. Typical Gram-positive bacterial strains of Lactobacillus casei, Streptococcus mitis and Micrococcus luteus were used as model bacterial strains in this research. We manipulated gene transfer to observe the impact of lux gene insertion on bacterial surface properties based on contact angle measurements, and we conducted column experiments to evaluate the impact of lux gene insertion on bacterial transport. After lux gene insertion, bacterial interactions with the porous media increased, demonstrating stronger deposition potential in the porous media. Accordingly, retention of the daughter strains increased. Lux gene insertion also resulted in an increase in bacterial dispersion and equilibrium adsorption in the porous media. The bacterial deposition coefficient was found to correlate with the free energy of interactions between bacteria and the porous media.

  16. Aberrant DNA methylation impacts gene expression and prognosis in breast cancer subtypes.

    PubMed

    Győrffy, Balázs; Bottai, Giulia; Fleischer, Thomas; Munkácsy, Gyöngyi; Budczies, Jan; Paladini, Laura; Børresen-Dale, Anne-Lise; Kristensen, Vessela N; Santarpia, Libero

    2016-01-01

    DNA methylation has a substantial impact on gene expression, affecting the prognosis of breast cancer (BC) patients dependent on molecular subtypes. In this study, we investigated the prognostic relevance of the expression of genes reported as aberrantly methylated, and the link between gene expression and DNA methylation in BC subtypes. The prognostic value of the expression of 144 aberrantly methylated genes was evaluated in ER+/HER2-, HER2+, and ER-/HER2- molecular BC subtypes, in a meta-analysis of two large transcriptomic cohorts of BC patients (n = 1,938 and n = 1,640). The correlation between gene expression and DNA methylation in distinct gene regions was also investigated in an independent dataset of 104 BCs. Survival and Pearson correlation analyses were computed for each gene separately. The expression of 48 genes was significantly associated with BC prognosis (p < 0.05), and 32 of these prognostic genes exhibited a direct expression-methylation correlation. The expression of several immune-related genes, including CD3D and HLA-A, was associated with both relapse-free survival (HR = 0.42, p = 3.5E-06; HR = 0.35, p = 1.7E-08) and overall survival (HR = 0.50, p = 5.5E-04; HR = 0.68, p = 4.5E-02) in ER-/HER2- BCs. On the overall, the distribution of both positive and negative expression-methylation correlation in distinct gene regions have different effects on gene expression and prognosis in BC subtypes. This large-scale meta-analysis allowed the identification of several genes consistently associated with prognosis, whose DNA methylation could represent a promising biomarker for prognostication and clinical stratification of patients with distinct BC subtypes.

  17. Impact of Panel Gene Testing for Hereditary Breast and Ovarian Cancer on Patients.

    PubMed

    Lumish, Heidi S; Steinfeld, Hallie; Koval, Carrie; Russo, Donna; Levinson, Elana; Wynn, Julia; Duong, James; Chung, Wendy K

    2017-03-29

    Recent advances in next generation sequencing have enabled panel gene testing, or simultaneous testing for mutations in multiple genes for a clinical condition. With more extensive and widespread genetic testing, there will be increased detection of genes with moderate penetrance without established clinical guidelines and of variants of uncertain significance (VUS), or genetic variants unknown to either be disease-causing or benign. This study surveyed 232 patients who underwent genetic counseling for hereditary breast and ovarian cancer to examine the impact of panel gene testing on psychological outcomes, patient understanding, and utilization of genetic information. The survey used standardized instruments including the Impact of Event Scale (IES), Multidimensional Impact of Cancer Risk Assessment (MICRA), Satisfaction with Decision Instrument (SWD), Ambiguity Tolerance Scale (AT-20), genetics knowledge, and utilization of genetic test results. Study results suggested that unaffected individuals with a family history of breast or ovarian cancer who received positive results were most significantly impacted by intrusive thoughts, avoidance, and distress. However, scores were also modestly elevated among unaffected patients with a family history of breast and ovarian cancer who received VUS, highlighting the impact of ambiguous results that are frequent among patients undergoing genetic testing with large panels of genes. Potential risk factors for increased genetic testing-specific distress in this study included younger age, black or African American race, Hispanic origin, lower education level, and lower genetic knowledge and highlight the need for developing strategies to provide effective counseling and education to these communities, particularly when genetic testing utilizes gene panels that more commonly return VUS. More detailed pre-test education and counseling may help patients appreciate the probability of various types of test results and how results

  18. Current impact of gene technology on healthcare. A map of economic assessments.

    PubMed

    Rogowski, Wolf

    2007-02-01

    It has been claimed that gene technology will induce revolutionary changes in healthcare. This paper investigates how and to what extent these changes have been economically assessed. A generic framework was developed to distinguish between methodologically similar evaluations of healthcare technology. Methodological issues and the current state of economic evidence concerning human DNA technology were extracted from publications within these groups of evaluations. Economic evaluations of "healthcare consisting of gene technology" were identified primarily for in vitro diagnostics for hereditary disease and others for pharmacogenetics and molecular pathology. "Healthcare enabled by gene technology" is far more encompassing and includes, e.g., biotechnology drugs for which various health economic evaluations can be found. Yet here, the impact of gene technology intertwines with the impact of other technologies and is therefore hardly susceptible to evaluation. The fields of evaluation may be classified best according to the two dimensions "purpose" and "stage of development". Current evaluations cover screening, diagnostic and treatment technologies in investigational, new and established stages. Apart from prenatal screening, healthcare consisting of gene technology was cost saving only for genotype tests replacing continuous phenotype tests and for one pharmacogenetic test. Conclusive evidence of favourable cost-effectiveness ratios is available only for few conditions. Hypotheses about the impact of gene technology on healthcare must be explicit about the definition of "genetic" medicine. A general statement regarding healthcare enabled by gene technology is not possible. Based on current evidence, an era of healthcare consisting of gene technology built on widespread predictive testing is not desirable from a health economic viewpoint.

  19. The 'Fat Mass and Obesity Related' (FTO) gene: Mechanisms of Impact on Obesity and Energy Balance.

    PubMed

    Speakman, John R

    2015-03-01

    A cluster of single nucleotide polymorphisms (SNPs) in the first intron of the fat mass and obesity related (FTO) gene were the first common variants discovered to be associated with body mass index and body fatness. This review summarises what has been later discovered about the biology of FTO drawing together information from both human and animal studies. Subsequent work showed that the 'at risk' alleles of these SNPs are associated with greater food intake and increased hunger/lowered satiety, but are not associated with altered resting energy expenditure or low physical activity in humans. FTO is an FE (II) and 2-oxoglutarate dependent DNA/RNA methylase. Contrasting the impact of the SNPs on energy balance in humans, knocking out or reducing activity of the Fto gene in the mouse resulted in lowered adiposity, elevated energy expenditure with no impact on food intake (but the impact on expenditure is disputed). In contrast, overexpression of the gene in mice led to elevated food intake and adiposity, with no impact on expenditure. In rodents, the Fto gene is widely expressed in the brain including hypothalamic nuclei linked to food intake regulation. Since its activity is 2-oxoglutarate dependent it could potentially act as a sensor of citrate acid cycle flux, but this function has been dismissed, and instead it has been suggested to be much more likely to act as an amino acid sensor, linking circulating AAs to the mammalian target of rapamycin complex 1. This may be fundamental to its role in development but the link to obesity is less clear. It has been recently suggested that although the obesity related SNPs reside in the first intron of FTO, they may not only impact FTO but mediate their obesity effects via nearby genes (notably RPGRIP1L and IRX3).

  20. Impact of IL-27 on hepatocyte antiviral gene expression and function

    PubMed Central

    2016-01-01

    Background: Interleukin (IL)-27 is a member of the IL-6/IL-12 family of cytokines. It is a potent cytokine, with potential antiviral impact, and has been shown to play a role in modulating functions of diverse cell types, including Th1, Th2, and NK and B cells, demonstrating both pro- and anti-inflammatory roles.  In hepatocytes, it is capable of inducing signal transducer and activator of transcription (STAT)1, STAT3 and interferon-stimulated genes. Methods: To address its role in viral hepatitis, the antiviral activity of IL-27 against hepatitis C virus (HCV) and hepatitis B virus (HBV) was tested in vitro using cell-culture-derived infectious HCV (HCVcc) cell culture system and the HepaRG HBV cell culture model. To further investigate the impact of IL-27 on hepatocytes, Huh7.5 cells were treated with IL-27 to analyse the differentially expressed genes by microarray analysis. Furthermore, by quantitative PCR, we analyzed the up-regulation of chemokine (CXCL)-10 in response to IL-27. Results: In both HCV and HBV infection models, we observed only a modest direct antiviral effect. Microarray analysis showed that the up-regulated genes mostly belonged to antigen presentation and DNA replication pathways, and involved strong up-regulation of CXCL-10, a gene associated with liver inflammation. Overall, gene set enrichment analysis showed a striking correlation of these genes with those up-regulated in response to related cytokines in diverse cell populations. Conclusion: Our data indicate that IL-27 can have a significant pro-inflammatory impact in vitro, although the direct antiviral effect is modest. It may have a potential impact on hepatocyte function, especially chemokine expression and antigen presentation. PMID:28058287

  1. Tandem repeat variation in human and great ape populations and its impact on gene expression divergence

    PubMed Central

    Bilgin Sonay, Tugce; Carvalho, Tiago; Robinson, Mark D.; Greminger, Maja P.; Krützen, Michael; Comas, David; Highnam, Gareth; Mittelman, David; Sharp, Andrew; Marques-Bonet, Tomàs; Wagner, Andreas

    2015-01-01

    Tandem repeats (TRs) are stretches of DNA that are highly variable in length and mutate rapidly. They are thus an important source of genetic variation. This variation is highly informative for population and conservation genetics. It has also been associated with several pathological conditions and with gene expression regulation. However, genome-wide surveys of TR variation in humans and closely related species have been scarce due to technical difficulties derived from short-read technology. Here we explored the genome-wide diversity of TRs in a panel of 83 human and nonhuman great ape genomes, in a total of six different species, and studied their impact on gene expression evolution. We found that population diversity patterns can be efficiently captured with short TRs (repeat unit length, 1–5 bp). We examined the potential evolutionary role of TRs in gene expression differences between humans and primates by using 30,275 larger TRs (repeat unit length, 2–50 bp). Genes that contained TRs in the promoters, in their 3′ untranslated region, in introns, and in exons had higher expression divergence than genes without repeats in the regions. Polymorphic small repeats (1–5 bp) had also higher expression divergence compared with genes with fixed or no TRs in the gene promoters. Our findings highlight the potential contribution of TRs to human evolution through gene regulation. PMID:26290536

  2. Epigenetics meets genetics in acute myeloid leukemia: clinical impact of a novel seven-gene score.

    PubMed

    Marcucci, Guido; Yan, Pearlly; Maharry, Kati; Frankhouser, David; Nicolet, Deedra; Metzeler, Klaus H; Kohlschmidt, Jessica; Mrózek, Krzysztof; Wu, Yue-Zhong; Bucci, Donna; Curfman, John P; Whitman, Susan P; Eisfeld, Ann-Kathrin; Mendler, Jason H; Schwind, Sebastian; Becker, Heiko; Bär, Constance; Carroll, Andrew J; Baer, Maria R; Wetzler, Meir; Carter, Thomas H; Powell, Bayard L; Kolitz, Jonathan E; Byrd, John C; Plass, Christoph; Garzon, Ramiro; Caligiuri, Michael A; Stone, Richard M; Volinia, Stefano; Bundschuh, Ralf; Bloomfield, Clara D

    2014-02-20

    Molecular risk stratification of acute myeloid leukemia (AML) is largely based on genetic markers. However, epigenetic changes, including DNA methylation, deregulate gene expression and may also have prognostic impact. We evaluated the clinical relevance of integrating DNA methylation and genetic information in AML. Next-generation sequencing analysis of methylated DNA identified differentially methylated regions (DMRs) associated with prognostic mutations in older (≥ 60 years) cytogenetically normal (CN) patients with AML (n = 134). Genes with promoter DMRs and expression levels significantly associated with outcome were used to compute a prognostic gene expression weighted summary score that was tested and validated in four independent patient sets (n = 355). In the training set, we identified seven genes (CD34, RHOC, SCRN1, F2RL1, FAM92A1, MIR155HG, and VWA8) with promoter DMRs and expression associated with overall survival (OS; P ≤ .001). Each gene had high DMR methylation and lower expression, which were associated with better outcome. A weighted summary expression score of the seven gene expression levels was computed. A low score was associated with a higher complete remission (CR) rate and longer disease-free survival and OS (P < .001 for all end points). This was validated in multivariable models and in two younger (< 60 years) and two older independent sets of patients with CN-AML. Considering the seven genes individually, the fewer the genes with high expression, the better the outcome. Younger and older patients with no genes or one gene with high expression had the best outcomes (CR rate, 94% and 87%, respectively; 3-year OS, 80% and 42%, respectively). A seven-gene score encompassing epigenetic and genetic prognostic information identifies novel AML subsets that are meaningful for treatment guidance.

  3. Epigenetics Meets Genetics in Acute Myeloid Leukemia: Clinical Impact of a Novel Seven-Gene Score

    PubMed Central

    Marcucci, Guido; Yan, Pearlly; Maharry, Kati; Frankhouser, David; Nicolet, Deedra; Metzeler, Klaus H.; Kohlschmidt, Jessica; Mrózek, Krzysztof; Wu, Yue-Zhong; Bucci, Donna; Curfman, John P.; Whitman, Susan P.; Eisfeld, Ann-Kathrin; Mendler, Jason H.; Schwind, Sebastian; Becker, Heiko; Bär, Constance; Carroll, Andrew J.; Baer, Maria R.; Wetzler, Meir; Carter, Thomas H.; Powell, Bayard L.; Kolitz, Jonathan E.; Byrd, John C.; Plass, Christoph; Garzon, Ramiro; Caligiuri, Michael A.; Stone, Richard M.; Volinia, Stefano; Bundschuh, Ralf; Bloomfield, Clara D.

    2014-01-01

    Purpose Molecular risk stratification of acute myeloid leukemia (AML) is largely based on genetic markers. However, epigenetic changes, including DNA methylation, deregulate gene expression and may also have prognostic impact. We evaluated the clinical relevance of integrating DNA methylation and genetic information in AML. Methods Next-generation sequencing analysis of methylated DNA identified differentially methylated regions (DMRs) associated with prognostic mutations in older (≥ 60 years) cytogenetically normal (CN) patients with AML (n = 134). Genes with promoter DMRs and expression levels significantly associated with outcome were used to compute a prognostic gene expression weighted summary score that was tested and validated in four independent patient sets (n = 355). Results In the training set, we identified seven genes (CD34, RHOC, SCRN1, F2RL1, FAM92A1, MIR155HG, and VWA8) with promoter DMRs and expression associated with overall survival (OS; P ≤ .001). Each gene had high DMR methylation and lower expression, which were associated with better outcome. A weighted summary expression score of the seven gene expression levels was computed. A low score was associated with a higher complete remission (CR) rate and longer disease-free survival and OS (P < .001 for all end points). This was validated in multivariable models and in two younger (< 60 years) and two older independent sets of patients with CN-AML. Considering the seven genes individually, the fewer the genes with high expression, the better the outcome. Younger and older patients with no genes or one gene with high expression had the best outcomes (CR rate, 94% and 87%, respectively; 3-year OS, 80% and 42%, respectively). Conclusion A seven-gene score encompassing epigenetic and genetic prognostic information identifies novel AML subsets that are meaningful for treatment guidance. PMID:24378410

  4. Impact of estrogen receptor α gene and oxytocin receptor gene polymorphisms on female sexuality

    PubMed Central

    Armeni, Anastasia K; Assimakopoulos, Konstantinos; Marioli, Dimitra; Koika, Vassiliki; Michaelidou, Euthychia; Mourtzi, Niki; Iconomou, Gregoris

    2017-01-01

    Over the past decades, research attention has increasingly been paid to the neurobiological component of sexual behavior. The aim of the present study was to investigate the correlation of estrogen receptor α (ERA) gene polymorphism (rs2234693-PvuII) (T→C substitution) and oxytocin receptor gene polymorphism (rs53576) (G→A substitution) with sexuality parameters of young, healthy women. One hundred thirty-three Greek heterosexual women, students in higher education institutions, 20–25 years of age, sexually active, with normal menstrual cycles (28–35 days), were recruited in the study. Exclusion criteria were chronic and/or major psychiatric diseases, use of oral contraceptive pills (OCs), polycystic ovary syndrome (PCOS), thyroid diseases as well as drugs that are implicated in hypothalamus–pituitary–gonadal axis. T allele (wildtype) of rs2234693 (PvuII) polymorphism of ERA gene was correlated with increased levels of arousal and lubrication, whereas A allele (polymorphic) of rs53576 (OXTR) polymorphism was correlated with increased arousal levels. The simultaneous presence of both T allele of rs2234693 (PvuII) and A allele of rs53576 (OXTR) polymorphisms (T + A group) was correlated with increased arousal, orgasm levels as well as female sexual function index full score. To our knowledge, this is the first study to investigate the interaction between ERA and OXTR with regard to sexual function in women. Female sexuality is a complex behavioral trait that encompasses both biological and psychological components. It seems that variability in female sexual response stems from genetic variability that characterizes endocrine, neurotransmitter and central nervous system influences. PMID:28069897

  5. Impact of estrogen receptor α gene and oxytocin receptor gene polymorphisms on female sexuality.

    PubMed

    Armeni, Anastasia K; Assimakopoulos, Konstantinos; Marioli, Dimitra; Koika, Vassiliki; Michaelidou, Euthychia; Mourtzi, Niki; Iconomou, Gregoris; Georgopoulos, Neoklis A

    2017-01-01

    Over the past decades, research attention has increasingly been paid to the neurobiological component of sexual behavior. The aim of the present study was to investigate the correlation of estrogen receptor α (ERA) gene polymorphism (rs2234693-PvuII) (T→C substitution) and oxytocin receptor gene polymorphism (rs53576) (G→A substitution) with sexuality parameters of young, healthy women. One hundred thirty-three Greek heterosexual women, students in higher education institutions, 20-25 years of age, sexually active, with normal menstrual cycles (28-35 days), were recruited in the study. Exclusion criteria were chronic and/or major psychiatric diseases, use of oral contraceptive pills (OCs), polycystic ovary syndrome (PCOS), thyroid diseases as well as drugs that are implicated in hypothalamus-pituitary-gonadal axis. T allele (wildtype) of rs2234693 (PvuII) polymorphism of ERA gene was correlated with increased levels of arousal and lubrication, whereas A allele (polymorphic) of rs53576 (OXTR) polymorphism was correlated with increased arousal levels. The simultaneous presence of both T allele of rs2234693 (PvuII) and A allele of rs53576 (OXTR) polymorphisms (T + A group) was correlated with increased arousal, orgasm levels as well as female sexual function index full score. To our knowledge, this is the first study to investigate the interaction between ERA and OXTR with regard to sexual function in women. Female sexuality is a complex behavioral trait that encompasses both biological and psychological components. It seems that variability in female sexual response stems from genetic variability that characterizes endocrine, neurotransmitter and central nervous system influences.

  6. Genetic variants in microRNA genes: impact on microRNA expression, function, and disease

    PubMed Central

    Cammaerts, Sophia; Strazisar, Mojca; De Rijk, Peter; Del Favero, Jurgen

    2015-01-01

    MicroRNAs (miRNAs) are important regulators of gene expression and like any other gene, their coding sequences are subject to genetic variation. Variants in miRNA genes can have profound effects on miRNA functionality at all levels, including miRNA transcription, maturation, and target specificity, and as such they can also contribute to disease. The impact of variants in miRNA genes is the focus of the present review. To put these effects into context, we first discuss the requirements of miRNA transcripts for maturation. In the last part an overview of available databases and tools and experimental approaches to investigate miRNA variants related to human disease is presented. PMID:26052338

  7. Ancestry dynamics in a South American population: The impact of gene flow and preferential mating.

    PubMed

    Hedrick, Philip W

    2017-07-01

    European ancestry in many populations in Latin America at autosomal loci is often higher than that from X-linked loci indicating more European male ancestry and more Amerindian female ancestry. Generally, this has been attributed to more European male gene flow but could also result from an advantage to European mating or reproductive success. Population genetic models were developed to investigate the dynamics of gene flow and mating or reproductive success. Using estimates of autosomal and X-chromosome European ancestry, the amount of male gene flow or mating or reproductive advantage for Europeans, or those with European ancestry, was estimated. In a population from Antioquia, Colombia with an estimated 79% European autosomal ancestry and an estimated 69% European X-chromosome ancestry, about 15% male gene flow from Europe or about 20% mating or reproductive advantage of Europeans over Amerindians resulted in these levels of European ancestry in the contemporary population. Combinations of gene flow and mating advantage were nearly additive in their impact. Gene flow, mating advantage, or a combination of both factors, are consistent with observed levels of European ancestry in a Latin American population. This approach provides a general methodology to determine the levels of gene flow and mating differences that can explain the observed contemporary differences in ancestry from autosomes and X-chromosomes. © 2017 Wiley Periodicals, Inc.

  8. Impact of gene patents on diagnostic testing: a new patent landscaping method applied to spinocerebellar ataxia.

    PubMed

    Berthels, Nele; Matthijs, Gert; Van Overwalle, Geertrui

    2011-11-01

    Recent reports in Europe and the United States raise concern about the potential negative impact of gene patents on the freedom to operate of diagnosticians and on the access of patients to genetic diagnostic services. Patents, historically seen as legal instruments to trigger innovation, could cause undesired side effects in the public health domain. Clear empirical evidence on the alleged hindering effect of gene patents is still scarce. We therefore developed a patent categorization method to determine which gene patents could indeed be problematic. The method is applied to patents relevant for genetic testing of spinocerebellar ataxia (SCA). The SCA test is probably the most widely used DNA test in (adult) neurology, as well as one of the most challenging due to the heterogeneity of the disease. Typically tested as a gene panel covering the five common SCA subtypes, we show that the patenting of SCA genes and testing methods and the associated licensing conditions could have far-reaching consequences on legitimate access to this gene panel. Moreover, with genetic testing being increasingly standardized, simply ignoring patents is unlikely to hold out indefinitely. This paper aims to differentiate among so-called 'gene patents' by lifting out the truly problematic ones. In doing so, awareness is raised among all stakeholders in the genetic diagnostics field who are not necessarily familiar with the ins and outs of patenting and licensing.

  9. Impact of gene patents on diagnostic testing: a new patent landscaping method applied to spinocerebellar ataxia

    PubMed Central

    Berthels, Nele; Matthijs, Gert; Van Overwalle, Geertrui

    2011-01-01

    Recent reports in Europe and the United States raise concern about the potential negative impact of gene patents on the freedom to operate of diagnosticians and on the access of patients to genetic diagnostic services. Patents, historically seen as legal instruments to trigger innovation, could cause undesired side effects in the public health domain. Clear empirical evidence on the alleged hindering effect of gene patents is still scarce. We therefore developed a patent categorization method to determine which gene patents could indeed be problematic. The method is applied to patents relevant for genetic testing of spinocerebellar ataxia (SCA). The SCA test is probably the most widely used DNA test in (adult) neurology, as well as one of the most challenging due to the heterogeneity of the disease. Typically tested as a gene panel covering the five common SCA subtypes, we show that the patenting of SCA genes and testing methods and the associated licensing conditions could have far-reaching consequences on legitimate access to this gene panel. Moreover, with genetic testing being increasingly standardized, simply ignoring patents is unlikely to hold out indefinitely. This paper aims to differentiate among so-called ‘gene patents' by lifting out the truly problematic ones. In doing so, awareness is raised among all stakeholders in the genetic diagnostics field who are not necessarily familiar with the ins and outs of patenting and licensing. PMID:21811306

  10. Gene mutations differently impact the prognosis of the myelodysplastic and myeloproliferative classes of chronic myelomonocytic leukemia.

    PubMed

    Cervera, Nathalie; Itzykson, Raphael; Coppin, Emilie; Prebet, Thomas; Murati, Anne; Legall, Stevan; Vey, Norbert; Solary, Eric; Birnbaum, Daniel; Gelsi-Boyer, Véronique

    2014-06-01

    Initially classified in the myelodysplastic syndromes (MDSs), chronic myelomonocytic leukemia (CMML) is currently considered as a MDS/myeloproliferative neoplasm. Two classes-myelodysplastic and myeloproliferative-have been distinguished upon the level of the white blood cell count (threshold 13 G/L). We analyzed mutations in 19 genes reported in CMML to determine if and how these mutations impacted the respective prognosis of the two classes. We defined four major mutated pathways (DNA methylation, ASXL1, splicing, and signaling) and determined their prognostic impact. The number of mutated pathways impacted overall survival in the myelodysplastic class but not in the myeloproliferative class. The myeloproliferative class had a worse prognosis than the myelodysplastic class and was impacted by RUNX1 mutations only. Our results argue for a reclassification of CMML based on the myelodysplastic/myeloproliferative status. © 2014 Wiley Periodicals, Inc.

  11. Impact of variation in the BDNF gene on social stress sensitivity and the buffering impact of positive emotions: replication and extension of a gene-environment interaction.

    PubMed

    van Winkel, Mark; Peeters, Frenk; van Winkel, Ruud; Kenis, Gunter; Collip, Dina; Geschwind, Nicole; Jacobs, Nele; Derom, Catherine; Thiery, Evert; van Os, Jim; Myin-Germeys, Inez; Wichers, Marieke

    2014-06-01

    A previous study reported that social stress sensitivity is moderated by the brain-derived-neurotrophic-factor(Val66Met) (BDNF rs6265) genotype. Additionally, positive emotions partially neutralize this moderating effect. The current study aimed to: (i) replicate in a new independent sample of subjects with residual depressive symptoms the moderating effect of BDNF(Val66Met) genotype on social stress sensitivity, (ii) replicate the neutralizing impact of positive emotions, (iii) extend these analyses to other variations in the BDNF gene in the new independent sample and the original sample of non-depressed individuals. Previous findings were replicated in an experience sampling method (ESM) study. Negative Affect (NA) responses to social stress were stronger in "Val/Met" carriers of BDNF(Val66Met) compared to "Val/Val" carriers. Positive emotions neutralized the moderating effect of BDNF(Val66Met) genotype on social stress sensitivity in a dose-response fashion. Finally, two of four additional BDNF SNPs (rs11030101, rs2049046) showed similar moderating effects on social stress-sensitivity across both samples. The neutralizing effect of positive emotions on the moderating effects of these two additional SNPs was found in one sample. In conclusion, ESM has important advantages in gene-environment (GxE) research and may attribute to more consistent findings in future GxE research. This study shows how the impact of BDNF genetic variation on depressive symptoms may be explained by its impact on subtle daily life responses to social stress. Further, it shows that the generation of positive affect (PA) can buffer social stress sensitivity and partially undo the genetic susceptibility.

  12. Bacteria and Genes Involved in Arsenic Speciation in Sediment Impacted by Long-Term Gold Mining

    PubMed Central

    Costa, Patrícia S.; Scholte, Larissa L. S.; Reis, Mariana P.; Chaves, Anderson V.; Oliveira, Pollyanna L.; Itabayana, Luiza B.; Suhadolnik, Maria Luiza S.; Barbosa, Francisco A. R.; Chartone-Souza, Edmar; Nascimento, Andréa M. A.

    2014-01-01

    The bacterial community and genes involved in geobiocycling of arsenic (As) from sediment impacted by long-term gold mining were characterized through culture-based analysis of As-transforming bacteria and metagenomic studies of the arsC, arrA, and aioA genes. Sediment was collected from the historically gold mining impacted Mina stream, located in one of the world’s largest mining regions known as the “Iron Quadrangle”. A total of 123 As-resistant bacteria were recovered from the enrichment cultures, which were phenotypically and genotypically characterized for As-transformation. A diverse As-resistant bacteria community was found through phylogenetic analyses of the 16S rRNA gene. Bacterial isolates were affiliated with Proteobacteria, Firmicutes, and Actinobacteria and were represented by 20 genera. Most were AsV-reducing (72%), whereas AsIII-oxidizing accounted for 20%. Bacteria harboring the arsC gene predominated (85%), followed by aioA (20%) and arrA (7%). Additionally, we identified two novel As-transforming genera, Thermomonas and Pannonibacter. Metagenomic analysis of arsC, aioA, and arrA sequences confirmed the presence of these genes, with arrA sequences being more closely related to uncultured organisms. Evolutionary analyses revealed high genetic similarity between some arsC and aioA sequences obtained from isolates and clone libraries, suggesting that those isolates may represent environmentally important bacteria acting in As speciation. In addition, our findings show that the diversity of arrA genes is wider than earlier described, once none arrA-OTUs were affiliated with known reference strains. Therefore, the molecular diversity of arrA genes is far from being fully explored deserving further attention. PMID:24755825

  13. Impacts of Neanderthal-Introgressed Sequences on the Landscape of Human Gene Expression.

    PubMed

    McCoy, Rajiv C; Wakefield, Jon; Akey, Joshua M

    2017-02-23

    Regulatory variation influencing gene expression is a key contributor to phenotypic diversity, both within and between species. Unfortunately, RNA degrades too rapidly to be recovered from fossil remains, limiting functional genomic insights about our extinct hominin relatives. Many Neanderthal sequences survive in modern humans due to ancient hybridization, providing an opportunity to assess their contributions to transcriptional variation and to test hypotheses about regulatory evolution. We developed a flexible Bayesian statistical approach to quantify allele-specific expression (ASE) in complex RNA-seq datasets. We identified widespread expression differences between Neanderthal and modern human alleles, indicating pervasive cis-regulatory impacts of introgression. Brain regions and testes exhibited significant downregulation of Neanderthal alleles relative to other tissues, consistent with natural selection influencing the tissue-specific regulatory landscape. Our study demonstrates that Neanderthal-inherited sequences are not silent remnants of ancient interbreeding but have measurable impacts on gene expression that contribute to variation in modern human phenotypes.

  14. Growth-rate regulated genes have profound impact on interpretation of transcriptome profiling in Saccharomyces cerevisiae

    PubMed Central

    Regenberg, Birgitte; Grotkjær, Thomas; Winther, Ole; Fausbøll, Anders; Åkesson, Mats; Bro, Christoffer; Hansen, Lars Kai; Brunak, Søren; Nielsen, Jens

    2006-01-01

    Background Growth rate is central to the development of cells in all organisms. However, little is known about the impact of changing growth rates. We used continuous cultures to control growth rate and studied the transcriptional program of the model eukaryote Saccharomyces cerevisiae, with generation times varying between 2 and 35 hours. Results A total of 5930 transcripts were identified at the different growth rates studied. Consensus clustering of these revealed that half of all yeast genes are affected by the specific growth rate, and that the changes are similar to those found when cells are exposed to different types of stress (>80% overlap). Genes with decreased transcript levels in response to faster growth are largely of unknown function (>50%) whereas genes with increased transcript levels are involved in macromolecular biosynthesis such as those that encode ribosomal proteins. This group also covers most targets of the transcriptional activator RAP1, which is also known to be involved in replication. A positive correlation between the location of replication origins and the location of growth-regulated genes suggests a role for replication in growth rate regulation. Conclusion Our data show that the cellular growth rate has great influence on transcriptional regulation. This, in turn, implies that one should be cautious when comparing mutants with different growth rates. Our findings also indicate that much of the regulation is coordinated via the chromosomal location of the affected genes, which may be valuable information for the control of heterologous gene expression in metabolic engineering. PMID:17105650

  15. Impact of karyotype organization on interlocus recombination between T cell receptor genes in Equidae.

    PubMed

    Drbalova, Jitka; Musilova, Petra; Kubickova, Svatava; Sebestova, Hana; Vahala, Jiri; Rubes, Jiri

    2014-01-01

    The T cell receptor (TCR) genes (TRA, TRB, TRD and TRG) reside in 3 different chromosomal regions. During the maturation of T lymphocytes, the TCR genes are rearranged by site-specific recombination, a process that also predisposes T cells to aberrant rearrangements. Illegitimate recombination between the TCR genes occurs at a low level in healthy individuals, but this frequency may correlate with the risk of lymphoma. The aim of this work was to investigate interlocus recombination in equids. Illegitimate rearrangements were studied in peripheral blood lymphocytes by FISH with painting and BAC probes and by sequencing of PCR products, and the frequencies of recombination were assessed in horses and 4 other equids. The presence of several trans-rearrangement products between the TRA and TRG genes was verified by PCR in all investigated equids. Frequencies of trans-rearrangements in horses are higher than in humans, and colocalization of the TCR genes on the same chromosome increases the incidence of trans-rearrangements between them. The orientation of the TCR genes does not impact interlocus recombination itself but does affect the viability of cells carrying its products and consequently the number of trans-rearrangements observed in lymphocytes.

  16. Impact of Neutron Exposure on Global Gene Expression in a Human Peripheral Blood Model.

    PubMed

    Broustas, Constantinos G; Xu, Yanping; Harken, Andrew D; Chowdhury, Mashkura; Garty, Guy; Amundson, Sally A

    2017-04-01

    The detonation of an improvised nuclear device would produce prompt radiation consisting of both photons (gamma rays) and neutrons. While much effort in recent years has gone into the development of radiation biodosimetry methods suitable for mass triage, the possible effect of neutrons on the endpoints studied has remained largely uninvestigated. We have used a novel neutron irradiator with an energy spectrum based on that 1-1.5 km from the epicenter of the Hiroshima blast to begin examining the effect of neutrons on global gene expression, and the impact this may have on the development of gene expression signatures for radiation biodosimetry. We have exposed peripheral blood from healthy human donors to 0.1, 0.3, 0.5 or 1 Gy of neutrons ex vivo using our neutron irradiator, and compared the transcriptomic response 24 h later to that resulting from sham exposure or exposure to 0.1, 0.3, 0.5, 1, 2 or 4 Gy of photons (X rays). We identified 125 genes that responded significantly to both radiation qualities as a function of dose, with the magnitude of response to neutrons generally being greater than that seen after X-ray exposure. Gene ontology analysis suggested broad involvement of the p53 signaling pathway and general DNA damage response functions across all doses of both radiation qualities. Regulation of immune response and chromatin-related functions were implicated only following the highest doses of neutrons, suggesting a physiological impact of greater DNA damage. We also identified several genes that seem to respond primarily as a function of dose, with less effect of radiation quality. We confirmed this pattern of response by quantitative real-time RT-PCR for BAX, TNFRSF10B, ITLN2 and AEN and suggest that gene expression may provide a means to differentiate between total dose and a neutron component.

  17. Impact of Neutron Exposure on Global Gene Expression in a Human Peripheral Blood Model

    PubMed Central

    Broustas, Constantinos G.; Xu, Yanping; Harken, Andrew D.; Chowdhury, Mashkura; Garty, Guy; Amundson, Sally A.

    2017-01-01

    The detonation of an improvised nuclear device would produce prompt radiation consisting of both photons (gamma rays) and neutrons. While much effort in recent years has gone into the development of radiation biodosimetry methods suitable for mass triage, the possible effect of neutrons on the endpoints studied has remained largely uninvestigated. We have used a novel neutron irradiator with an energy spectrum based on that 1–1.5 km from the epicenter of the Hiroshima blast to begin examining the effect of neutrons on global gene expression, and the impact this may have on the development of gene expression signatures for radiation biodosimetry. We have exposed peripheral blood from healthy human donors to 0.1, 0.3, 0.5 or 1 Gy of neutrons ex vivo using our neutron irradiator, and compared the transcriptomic response 24 h later to that resulting from sham exposure or exposure to 0.1, 0.3, 0.5, 1, 2 or 4 Gy of photons (X rays). We identified 125 genes that responded significantly to both radiation qualities as a function of dose, with the magnitude of response to neutrons generally being greater than that seen after X-ray exposure. Gene ontology analysis suggested broad involvement of the p53 signaling pathway and general DNA damage response functions across all doses of both radiation qualities. Regulation of immune response and chromatin-related functions were implicated only following the highest doses of neutrons, suggesting a physiological impact of greater DNA damage. We also identified several genes that seem to respond primarily as a function of dose, with less effect of radiation quality. We confirmed this pattern of response by quantitative real-time RT-PCR for BAX, TNFRSF10B, ITLN2 and AEN and suggest that gene expression may provide a means to differentiate between total dose and a neutron component. PMID:28140791

  18. Microevolution of Duplications and Deletions and Their Impact on Gene Expression in the Nematode Pristionchus pacificus

    PubMed Central

    2015-01-01

    The evolution of diversity across the animal kingdom has been accompanied by tremendous gene loss and gain. While comparative genomics has been fruitful to characterize differences in gene content across highly diverged species, little is known about the microevolution of structural variations that cause these differences in the first place. In order to investigate the genomic impact of structural variations, we made use of genomic and transcriptomic data from the nematode Pristionchus pacificus, which has been established as a satellite model to Caenorhabditis elegans for comparative biology. We exploit the fact that P. pacificus is a highly diverse species for which various genomic data including the draft genome of a sister species P. exspectatus is available. Based on resequencing coverage data for two natural isolates we identified large (> 2kb) deletions and duplications relative to the reference strain. By restriction to completely syntenic regions between P. pacificus and P. exspectatus, we were able to polarize the comparison and to assess the impact of structural variations on expression levels. We found that while loss of genes correlates with lack of expression, duplication of genes has virtually no effect on gene expression. Further investigating expression of individual copies at sites that segregate between the duplicates, we found in the majority of cases only one of the copies to be expressed. Nevertheless, we still find that certain gene classes are strongly depleted in deletions as well as duplications, suggesting evolutionary constraint acting on synteny. In summary, our results are consistent with a model, where most structural variations are either deleterious or neutral and provide first insights into the microevolution of structural variations in the P. pacificus genome. PMID:26125626

  19. Impact of gene gains, losses and duplication modes on the origin and diversification of vertebrates.

    PubMed

    Cañestro, Cristian; Albalat, Ricard; Irimia, Manuel; Garcia-Fernàndez, Jordi

    2013-02-01

    The study of the evolutionary origin of vertebrates has been linked to the study of genome duplications since Susumo Ohno suggested that the successful diversification of vertebrate innovations was facilitated by two rounds of whole-genome duplication (2R-WGD) in the stem vertebrate. Since then, studies on the functional evolution of many genes duplicated in the vertebrate lineage have provided the grounds to support experimentally this link. This article reviews cases of gene duplications derived either from the 2R-WGD or from local gene duplication events in vertebrates, analyzing their impact on the evolution of developmental innovations. We analyze how gene regulatory networks can be rewired by the activity of transposable elements after genome duplications, discuss how different mechanisms of duplication might affect the fate of duplicated genes, and how the loss of gene duplicates might influence the fate of surviving paralogs. We also discuss the evolutionary relationships between gene duplication and alternative splicing, in particular in the vertebrate lineage. Finally, we discuss the role that the 2R-WGD might have played in the evolution of vertebrate developmental gene networks, paying special attention to those related to vertebrate key features such as neural crest cells, placodes, and the complex tripartite brain. In this context, we argue that current evidences points that the 2R-WGD may not be linked to the origin of vertebrate innovations, but to their subsequent diversification in a broad variety of complex structures and functions that facilitated the successful transition from peaceful filter-feeding non-vertebrate ancestors to voracious vertebrate predators. Copyright © 2013 Elsevier Ltd. All rights reserved.

  20. Multiple Gene-Environment Interactions on the Angiogenesis Gene-Pathway Impact Rectal Cancer Risk and Survival.

    PubMed

    Sharafeldin, Noha; Slattery, Martha L; Liu, Qi; Franco-Villalobos, Conrado; Caan, Bette J; Potter, John D; Yasui, Yutaka

    2017-09-28

    Characterization of gene-environment interactions (GEIs) in cancer is limited. We aimed at identifying GEIs in rectal cancer focusing on a relevant biologic process involving the angiogenesis pathway and relevant environmental exposures: cigarette smoking, alcohol consumption, and animal protein intake. We analyzed data from 747 rectal cancer cases and 956 controls from the Diet, Activity and Lifestyle as a Risk Factor for Rectal Cancer study. We applied a 3-step analysis approach: first, we searched for interactions among single nucleotide polymorphisms on the pathway genes; second, we searched for interactions among the genes, both steps using Logic regression; third, we examined the GEIs significant at the 5% level using logistic regression for cancer risk and Cox proportional hazards models for survival. Permutation-based test was used for multiple testing adjustment. We identified 8 significant GEIs associated with risk among 6 genes adjusting for multiple testing: TNF (OR = 1.85, 95% CI: 1.10, 3.11), TLR4 (OR = 2.34, 95% CI: 1.38, 3.98), and EGR2 (OR = 2.23, 95% CI: 1.04, 4.78) with smoking; IGF1R (OR = 1.69, 95% CI: 1.04, 2.72), TLR4 (OR = 2.10, 95% CI: 1.22, 3.60) and EGR2 (OR = 2.12, 95% CI: 1.01, 4.46) with alcohol; and PDGFB (OR = 1.75, 95% CI: 1.04, 2.92) and MMP1 (OR = 2.44, 95% CI: 1.24, 4.81) with protein. Five GEIs were associated with survival at the 5% significance level but not after multiple testing adjustment: CXCR1 (HR = 2.06, 95% CI: 1.13, 3.75) with smoking; and KDR (HR = 4.36, 95% CI: 1.62, 11.73), TLR2 (HR = 9.06, 95% CI: 1.14, 72.11), EGR2 (HR = 2.45, 95% CI: 1.42, 4.22), and EGFR (HR = 6.33, 95% CI: 1.95, 20.54) with protein. GEIs between angiogenesis genes and smoking, alcohol, and animal protein impact rectal cancer risk. Our results support the importance of considering the biologic hypothesis to characterize GEIs associated with cancer outcomes.

  1. Psychological resilience and the gene regulatory impact of posttraumatic stress in Nepali child soldiers

    PubMed Central

    Worthman, Carol M.; Adhikari, Ramesh P.; Luitel, Nagendra P.; Arevalo, Jesusa M. G.; Ma, Jeffrey; McCreath, Heather; Seeman, Teresa E.; Crimmins, Eileen M.; Cole, Steven W.

    2016-01-01

    Adverse social conditions in early life have been linked to increased expression of proinflammatory genes and reduced expression of antiviral genes in circulating immune cells—the conserved transcriptional response to adversity (CTRA). However, it remains unclear whether such effects are specific to the Western, educated, industrialized, rich, and democratic (WEIRD) cultural environments in which previous research has been conducted. To assess the roles of early adversity and individual psychological resilience in immune system gene regulation within a non-WEIRD population, we evaluated CTRA gene-expression profiles in 254 former child soldiers and matched noncombatant civilians 5 y after the People’s War in Nepal. CTRA gene expression was up-regulated in former child soldiers. These effects were linked to the degree of experienced trauma and associated distress—that is, posttraumatic stress disorder (PTSD) severity—more than to child soldier status per se. Self-perceived psychological resilience was associated with marked buffering of CTRA activation such that PTSD-affected former child soldiers with high levels of personal resilience showed molecular profiles comparable to those of PTSD-free civilians. These results suggest that CTRA responses to early life adversity are not restricted to WEIRD cultural contexts and they underscore the key role of resilience in determining the molecular impact of adverse environments. PMID:27402736

  2. Kindness in the blood: A randomized controlled trial of the gene regulatory impact of prosocial behavior.

    PubMed

    Nelson-Coffey, S Katherine; Fritz, Megan M; Lyubomirsky, Sonja; Cole, Steve W

    2017-07-01

    Prosocial behavior is linked to longevity, but few studies have experimentally manipulated prosocial behavior to identify the causal mechanisms underlying this association. One possible mediating pathway involves changes in gene expression that may subsequently influence disease development or resistance. In the current study, we examined changes in a leukocyte gene expression profile known as the Conserved Transcriptional Response to Adversity (CTRA) in 159 adults who were randomly assigned for 4 weeks to engage in prosocial behavior directed towards specific others, prosocial behavior directed towards the world in general, self-focused kindness, or a neutral control task. Those randomized to prosocial behavior towards specific others demonstrated improvements (i.e., reductions) in leukocyte expression of CTRA indicator genes. No significant changes in CTRA gene expression were observed in the other 3 conditions. These findings suggest that prosocial behavior can causally impact leukocyte gene expression profiles in ways that might potentially help explain the previously observed health advantages associated with social ties. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Impact of blue LED irradiation on proliferation and gene expression of cultured human keratinocytes

    NASA Astrophysics Data System (ADS)

    Becker, Anja; Sticht, Carsten; Dweep, Harsh; van Abeelen, Frank A.; Gretz, Norbert; Oversluizen, Gerrit

    2015-03-01

    Blue light is known for its anti-microbial, anti-proliferative and anti-inflammatory effects. Furthermore, it is already used for the treatment of neonatal jaundice and acne. However, little is known about the exact mechanisms of action on gene expression level. The aim of this study was to assess the impact of blue LED irradiation on the proliferation and gene expression in immortalized human keratinocytes (HaCaT) in vitro. Furthermore its safety was assessed. XTT-tests revealed a decrease in cell proliferation in blue light irradiated cells depending on the duration of light irradiation. Moreover, gene expression analysis demonstrated deregulated genes already 3 hours after blue light irradiation. 24 hours after blue light irradiation the effects seemed to be even more pronounced. The oxidative stress response was significantly increased, pointing to increased ROS production due to blue light, as well as steroid hormone biosynthesis. Downregulated pathways or biological processes were connected to anti-inflammatory response. Interestingly, also the melanoma pathway contained significantly downregulated genes 24 hours after blue light irradiation, which stands in accordance to literature that blue light can also inhibit proliferation in cancer cells. First tests with melanoma cells revealed a decrease in cell proliferation after blue light irradiation. In conclusion, blue light irradiation might open avenues to new therapeutic regimens; at least blue light seems to have no effect that induces cancer growth or formation.

  4. Psychological resilience and the gene regulatory impact of posttraumatic stress in Nepali child soldiers.

    PubMed

    Kohrt, Brandon A; Worthman, Carol M; Adhikari, Ramesh P; Luitel, Nagendra P; Arevalo, Jesusa M G; Ma, Jeffrey; McCreath, Heather; Seeman, Teresa E; Crimmins, Eileen M; Cole, Steven W

    2016-07-19

    Adverse social conditions in early life have been linked to increased expression of proinflammatory genes and reduced expression of antiviral genes in circulating immune cells-the conserved transcriptional response to adversity (CTRA). However, it remains unclear whether such effects are specific to the Western, educated, industrialized, rich, and democratic (WEIRD) cultural environments in which previous research has been conducted. To assess the roles of early adversity and individual psychological resilience in immune system gene regulation within a non-WEIRD population, we evaluated CTRA gene-expression profiles in 254 former child soldiers and matched noncombatant civilians 5 y after the People's War in Nepal. CTRA gene expression was up-regulated in former child soldiers. These effects were linked to the degree of experienced trauma and associated distress-that is, posttraumatic stress disorder (PTSD) severity-more than to child soldier status per se. Self-perceived psychological resilience was associated with marked buffering of CTRA activation such that PTSD-affected former child soldiers with high levels of personal resilience showed molecular profiles comparable to those of PTSD-free civilians. These results suggest that CTRA responses to early life adversity are not restricted to WEIRD cultural contexts and they underscore the key role of resilience in determining the molecular impact of adverse environments.

  5. Tetracycline residues and tetracycline resistance genes in groundwater impacted by swine production facilities

    USGS Publications Warehouse

    Mackie, R.I.; Koike, S.; Krapac, I.; Chee-Sanford, J.; Maxwell, Susan; Aminov, R.I.

    2006-01-01

    Antibiotics are used at therapeutic levels to treat disease; at slightly lower levels as prophylactics; and at low, subtherapeutic levels for growth promotion and improvement of feed efficiency. Over 88% of swine producers in the United States gave antimicrobials to grower/finisher pigs in feed as a growth promoter in 2000. It is estimated that ca. 75% of antibiotics are not absorbed by animals and are excreted in urine and feces. The extensive use of antibiotics in swine production has resulted in antibiotic resistance in many intestinal bacteria, which are also excreted in swine feces, resulting in dissemination of resistance genes into the environment.To assess the impact of manure management on groundwater quality, groundwater samples have been collected near two swine confinement facilities that use lagoons for manure storage and treatment. Several key contaminant indicators-including inorganic ions, antibiotics, and antibiotic resistance genes-were analyzed in groundwater collected from the monitoring wells. Chloride, ammonium, potassium, and sodium were predominant inorganic constituents in the manure samples and served as indicators of groundwater contamination. Based on these analyses, shallow groundwater has been impacted by lagoon seepage at both sites. Liquid chromatography-mass spectroscopy (LC-MS) was used to measure the dissolved concentrations of tetracycline, chlortetracycline, and oxytetracycline in groundwater and manure. Although tetracyclines were regularly used at both facilities, they were infrequently detected in manure samples and then at relatively trace concentrations. Concentrations of all tetracyclines and their breakdown products in the groundwater sampled were generally less than 0.5 ??g/L.Bacterial tetracycline resistance genes served as distinct genotypic markers to indicate the dissemination and mobility of antibiotic resistance genes that originated from the lagoons. Applying PCR to genomic DNA extracted from the lagoon and

  6. Impact of virulence genes on sepsis severity and survival in Escherichia coli bacteremia

    PubMed Central

    Mora-Rillo, Marta; Fernández-Romero, Natalia; Francisco, Carolina Navarro-San; Díez-Sebastián, Jesús; Romero-Gómez, Maria Pilar; Fernández, Francisco Arnalich; López, Jose Ramon Arribas; Mingorance, Jesús

    2015-01-01

    Extraintestinal pathogenic Escherichia coli (ExPEC) are a frequent cause of bacteremia and sepsis, but the role of ExPEC genetic virulence factors (VFs) in sepsis development and outcome is ill-defined. Prospective study including 120 adult patients with E. coli bacteremia to investigate the impact of bacterial and host factors on sepsis severity and mortality. Patients' clinical and demographic data were registered. Phylogenetic background of E. coli isolates was analyzed by SNP pyrosequencing and VFs by PCR. The E. coli isolates presented an epidemic population structure with 6 dominant clones making up to half of the isolates. VF gene profiles were highly diverse. Multivariate analysis for sepsis severity showed that the presence of cnf and blaTEM genes increased the risk of severe illness by 6.75 (95% confidence interval [CI] 1.79–24.71) and 2.59 (95% CI 1.04–6.43) times respectively, while each point in the Pitt score increased the risk by 1.34 (95% CI 1.02–1.76) times. Multivariate analysis for mortality showed that active chemotherapy (OR 17.87, 95% CI 3.35–95.45), McCabe-Jackson Index (OR for rapidly fatal category 120.15, 95% CI 4.19–3446.23), Pitt index (OR 1.78, 95% CI 1.25–2.56) and presence of fyuA gene (OR 8.05, 95% CI 1.37–47.12) were associated to increased mortality while the presence of P fimbriae genes had a protective role (OR 0.094, 95%IC 0.018–0.494). Bacteremic E. coli had a high diversity of genetic backgrounds and VF gene profiles. Bacterial VFs and host determinants had an impact on disease evolution and mortality. PMID:25654604

  7. CARDIOPULMONARY GENE EXPRESSION PROFILES IN NORMO- AND SPONTANEOUSLY HYPERSENSITIVE (SH) RATS: IMPACT OF PARTICULATE MATTER (PM) EXPOSURE

    EPA Science Inventory

    CARDIOPULMONARY GENE EXPRESSION PROFILES IN NORMO- AND SPONTANEOUSLY HYPERTENSIVE (SH) RATS: IMPACT OF PARTICULATE MATTER (PM) EXPOSURE. SS Nadadur UP Kodavanti, Pulmonary Toxicology Branch, ETD, ORD, NHEERL, US Environmental Protection Agency, Research Triangle Park, NC 27711.

  8. CARDIOPULMONARY GENE EXPRESSION PROFILES IN NORMO- AND SPONTANEOUSLY HYPERSENSITIVE (SH) RATS: IMPACT OF PARTICULATE MATTER (PM) EXPOSURE

    EPA Science Inventory

    CARDIOPULMONARY GENE EXPRESSION PROFILES IN NORMO- AND SPONTANEOUSLY HYPERTENSIVE (SH) RATS: IMPACT OF PARTICULATE MATTER (PM) EXPOSURE. SS Nadadur UP Kodavanti, Pulmonary Toxicology Branch, ETD, ORD, NHEERL, US Environmental Protection Agency, Research Triangle Park, NC 27711.

  9. Middle age has a significant impact on gene expression during skin wound healing in male mice.

    PubMed

    Yanai, Hagai; Lumenta, David Benjamin; Vierlinger, Klemens; Hofner, Manuela; Kitzinger, Hugo-Benito; Kamolz, Lars-Peter; Nöhammer, Christa; Chilosi, Marco; Fraifeld, Vadim E

    2016-08-01

    The vast majority of research on the impact of age on skin wound healing (WH) compares old animals to young ones. The middle age is often ignored in biogerontological research despite the fact that many functions that decline in an age-dependent manner have starting points in mid-life. With this in mind, we examined gene expression patterns during skin WH in late middle-aged versus young adult male mice, using the head and back punch models. The rationale behind this study was that the impact of age would first be detectable at the transcriptional level. We pinpointed several pathways which were over-activated in the middle-aged mice, both in the intact skin and during WH. Among them were various metabolic, immune-inflammatory and growth-promoting pathways. These transcriptional changes were much more pronounced in the head than in the back. In summary, the middle age has a significant impact on gene expression in intact and healing skin. It seems that the head punch model is more sensitive to the effect of age than the back model, and we suggest that it should be more widely applied in aging research on wound healing.

  10. The impact of endurance exercise on global and AMPK gene-specific DNA methylation

    SciTech Connect

    King-Himmelreich, Tanya S.; Schramm, Stefanie; Wolters, Miriam C.; Schmetzer, Julia; Möser, Christine V.; Knothe, Claudia; Geisslinger, Gerd

    2016-05-27

    Alterations in gene expression as a consequence of physical exercise are frequently described. The mechanism of these regulations might depend on epigenetic changes in global or gene-specific DNA methylation levels. The AMP-activated protein kinase (AMPK) plays a key role in maintenance of energy homeostasis and is activated by increases in the AMP/ATP ratio as occurring in skeletal muscles after sporting activity. To analyze whether exercise has an impact on the methylation status of the AMPK promoter, we determined the AMPK methylation status in human blood samples from patients before and after sporting activity in the context of rehabilitation as well as in skeletal muscles of trained and untrained mice. Further, we examined long interspersed nuclear element 1 (LINE-1) as indicator of global DNA methylation changes. Our results revealed that light sporting activity in mice and humans does not alter global DNA methylation but has an effect on methylation of specific CpG sites in the AMPKα2 gene. These regulations were associated with a reduced AMPKα2 mRNA and protein expression in muscle tissue, pointing at a contribution of the methylation status to AMPK expression. Taken together, these results suggest that exercise influences AMPKα2 gene methylation in human blood and eminently in the skeletal muscle of mice and therefore might repress AMPKα2 gene expression. -- Highlights: •AMPK gene methylation increases after moderate endurance exercise in humans and mice. •AMPKα mRNA and protein decrease after moderate endurance exercise in mice. •Global DNA methylation is not affected under the same conditions.

  11. Clinical impact of a targeted next-generation sequencing gene panel for autoinflammation and vasculitis

    PubMed Central

    Standing, Ariane; Keylock, Annette; Price-Kuehne, Fiona; Melo Gomes, Sonia; Rowczenio, Dorota; Nanthapisal, Sira; Cullup, Thomas; Nyanhete, Rodney; Ashton, Emma; Murphy, Claire; Clarke, Megan; Ahlfors, Helena; Jenkins, Lucy; Gilmour, Kimberly; Eleftheriou, Despina; Lachmann, Helen J.; Hawkins, Philip N.; Klein, Nigel; Brogan, Paul A.

    2017-01-01

    Background Monogenic autoinflammatory diseases (AID) are a rapidly expanding group of genetically diverse but phenotypically overlapping systemic inflammatory disorders associated with dysregulated innate immunity. They cause significant morbidity, mortality and economic burden. Here, we aimed to develop and evaluate the clinical impact of a NGS targeted gene panel, the “Vasculitis and Inflammation Panel” (VIP) for AID and vasculitis. Methods The Agilent SureDesign tool was used to design 2 versions of VIP; VIP1 targeting 113 genes, and a later version, VIP2, targeting 166 genes. Captured and indexed libraries (QXT Target Enrichment System) prepared for 72 patients were sequenced as a multiplex of 16 samples on an Illumina MiSeq sequencer in 150bp paired-end mode. The cohort comprised 22 positive control DNA samples from patients with previously validated mutations in a variety of the genes; and 50 prospective samples from patients with suspected AID in whom previous Sanger based genetic screening had been non-diagnostic. Results VIP was sensitive and specific at detecting all the different types of known mutations in 22 positive controls, including gene deletion, small INDELS, and somatic mosaicism with allele fraction as low as 3%. Six/50 patients (12%) with unclassified AID had at least one class 5 (clearly pathogenic) variant; and 11/50 (22%) had at least one likely pathogenic variant (class 4). Overall, testing with VIP resulted in a firm or strongly suspected molecular diagnosis in 16/50 patients (32%). Conclusions The high diagnostic yield and accuracy of this comprehensive targeted gene panel validate the use of broad NGS-based testing for patients with suspected AID. PMID:28750028

  12. Impact of high predation risk on genome-wide hippocampal gene expression in snowshoe hares.

    PubMed

    Lavergne, Sophia G; McGowan, Patrick O; Krebs, Charles J; Boonstra, Rudy

    2014-11-01

    The population dynamics of snowshoe hares (Lepus americanus) are fundamental to the ecosystem dynamics of Canada's boreal forest. During the 8- to 11-year population cycle, hare densities can fluctuate up to 40-fold. Predators in this system (lynx, coyotes, great-horned owls) affect population numbers not only through direct mortality but also through sublethal effects. The chronic stress hypothesis posits that high predation risk during the decline severely stresses hares, leading to greater stress responses, heightened ability to mobilize cortisol and energy, and a poorer body condition. These effects may result in, or be mediated by, differential gene expression. We used an oligonucleotide microarray designed for a closely-related species, the European rabbit (Oryctolagus cuniculus), to characterize differences in genome-wide hippocampal RNA transcript abundance in wild hares from the Yukon during peak and decline phases of a single cycle. A total of 106 genes were differentially regulated between phases. Array results were validated with quantitative real-time PCR, and mammalian protein sequence similarity was used to infer gene function. In comparison to hares from the peak, decline phase hares showed increased expression of genes involved in metabolic processes and hormone response, and decreased expression of immune response and blood cell formation genes. We found evidence for predation risk effects on the expression of genes whose putative functions correspond with physiological impacts known to be induced by predation risk in snowshoe hares. This study shows, for the first time, a link between changes in demography and alterations in neural RNA transcript abundance in a natural population.

  13. The impact of founder effects, gene flow, and European admixture on native American genetic diversity.

    PubMed

    Hunley, Keith; Healy, Meghan

    2011-12-01

    Recent studies have concluded that the global pattern of neutral genetic diversity in humans reflects a series of founder effects and population movements associated with our recent expansion out of Africa. In contrast, regional studies tend to emphasize the significance of more complex patterns of colonization, gene flow, and secondary population movements in shaping patterns of diversity. Our objective in this study is to examine how founder effects, gene flow, and European admixture have molded patterns of neutral genetic diversity in the Americas. Our strategy is to test the fit of a serial founder effects process to the pattern of neutral autosomal genetic variation and to examine the contribution of gene flow and European admixture to departures from fit. The genetic data consist of 678 autosomal microsatellite loci assayed by Wang and colleagues in 530 individuals in 29 widely distributed Native American populations. We find that previous evidence for serial founder effects in the Americas may be driven in part by high levels of European admixture in northern North America, intermediate levels in Central America, and low levels in eastern South America. Geographically patterned admixture may also account for previously reported genetic differences between Andean and Amazonian groups. Though admixture has obscured the precise details of precontact evolutionary processes, we find that genetic diversity is still largely hierarchically structured and that gene flow between neighboring groups has had surprisingly little impact on macrogeographic patterns of genetic diversity in the Americas. 2011 Wiley Periodicals, Inc.

  14. Apolipoprotein A5: A newly identified gene impacting plasmatriglyceride levels in humans and mice

    SciTech Connect

    Pennacchio, Len A.; Rubin, Edward M.

    2002-09-15

    Apolipoprotein A5 (APOA5) is a newly described member of theapolipoprotein gene family whose initial discovery arose from comparativesequence analysis of the mammalian APOA1/C3/A4 gene cluster. Functionalstudies in mice indicated that alteration in the level of APOA5significantly impacted plasma triglyceride concentrations. Miceover-expressing human APOA5 displayed significantly reducedtriglycerides, while mice lacking apoA5 had a large increase in thislipid parameter. Studies in humans have also suggested an important rolefor APOA5 in determining plasma triglyceride concentrations. In theseexperiments, polymorphisms in the human gene were found to define severalcommon haplotypes that were associated with significant changes intriglyceride concentrations in multiple populations. Several separateclinical studies have provided consistent and strong support for theeffect with 24 percent of Caucasians, 35 percent of African-Americans and53 percent of Hispanics carrying APOA5 haplotypes associated withincreased plasma triglyceride levels. In summary, APOA5 represents anewly discovered gene involved in triglyceride metabolism in both humansand mice whose mechanism of action remains to be deciphered.

  15. The impact of non-electrical factors on electrical gene transfer

    PubMed Central

    Hu, Jiemiao; Cutrera, Jeffry; Li, Shulin

    2014-01-01

    Electrical pulses directly and effectively boost both in vitro and in vivo gene transfer, but this process is greatly affected by non-electrical factors that exist during electroporation. These factors include, but are not limited to, the types of cells or tissues used, the property of DNA, DNA formulation, and the expressed protein. In this mini-review, we only describe and discuss a summary of DNA properties and selected DNA formulations on gene transfer via electroporation. The properties of DNA were selected for review because a substantial amount of remarkable work has been performed during the past few years but has received less notice than other work, although DNA properties appear to be critical for boosting electroporation delivery. The selected formulations will be covered in this mini-review because we are only interested in the simple formulations that could be used for cell or gene therapy via electroporation. Plus, there was an extensive review of DNA formulations in the first edition of this book. The formulations discussed in this mini-review represent novel developments in recent years and may impact electroporation significantly. These advancements in DNA formulations could prove to be important for gene delivery and disease treatment. PMID:24510810

  16. The Impact of RNA-seq Alignment Pipeline on Detection of Differentially Expressed Genes.

    PubMed

    Yang, Cheng; Wu, Po-Yen; Phan, John H; Wang, May D

    2014-12-01

    RNA-seq data analysis pipelines are generally composed of sequence alignment, expression quantification, expression normalization, and differentially expressed gene (DEG) detection. Each step has numerous specific tools or algorithms, so we cannot explore all combinatorial pipelines and provide a comprehensive comparison of pipeline performance. To understand the mechanism of RNA-seq data analysis pipelines and provide some useful information for pipeline selection, we believe it is necessary to analyze the interactions among pipeline components. In this paper, by combining different alignment algorithms with the same quantification, normalization, and DEG detection tools, we construct nine RNA-seq pipelines to analyze the impact of RNA-seq alignment on downstream applications of gene expression estimates. Specifically, we find moderate linear correlation between the number of DEGs detected and the percentage of reads aligned with zero mismatch.

  17. Potential impact of human mitochondrial replacement on global policy regarding germline gene modification.

    PubMed

    Ishii, Tetsuya

    2014-08-01

    Previous discussions regarding human germline gene modification led to a global consensus that no germline should undergo genetic modification. However, the UK Human Fertilisation and Embryology Authority, having conducted at the UK Government's request a scientific review and a wide public consultation, provided advice to the Government on the pros and cons of Parliament's lifting a ban on altering mitochondrial DNA content of human oocytes and embryos, so as to permit the prevention of maternal transmission of mitochondrial diseases. In this commentary, relevant ethical and biomedical issues are examined and requirements for proceeding with this novel procedure are suggested. Additionally, potentially significant impacts of the UK legalization on global policy concerning germline gene modification are discussed in the context of recent advances in genome-editing technology. It is concluded that international harmonization is needed, as well as further ethical and practical consideration, prior to the legalization of human mitochondrial replacement.

  18. Analysis of the multi-copied genes and the impact of the redundant protein coding sequences on gene annotation in prokaryotic genomes.

    PubMed

    Yu, Jia-Feng; Chen, Qing-Li; Ren, Jing; Yang, Yan-Ling; Wang, Ji-Hua; Sun, Xiao

    2015-07-07

    The important roles of duplicated genes in evolutional process have been recognized in bacteria, archaebacteria and eukaryotes, while there is very little study on the multi-copied protein coding genes that share sequence identity of 100%. In this paper, the multi-copied protein coding genes in a number of prokaryotic genomes are comprehensively analyzed firstly. The results show that 0-15.93% of the protein coding genes in each genome are multi-copied genes and 0-16.49% of the protein coding genes in each genome are highly similar with the sequence identity ≥ 80%. Function and COG (Clusters of Orthologous Groups of proteins) analysis shows that 64.64% of multi-copied genes concentrate on the function of transposase and 86.28% of the COG assigned multi-copied genes concentrate on the COG code of 'L'. Furthermore, the impact of redundant protein coding sequences on the gene prediction results is studied. The results show that the problem of protein coding sequence redundancies cannot be ignored and the consistency of the gene annotation results before and after excluding the redundant sequences is negatively related with the sequences redundancy degree of the protein coding sequences in the training set.

  19. Burden Analysis of Rare Microdeletions Suggests a Strong Impact of Neurodevelopmental Genes in Genetic Generalised Epilepsies

    PubMed Central

    Trucks, Holger; Schulz, Herbert; de Kovel, Carolien G.; Kasteleijn-Nolst Trenité, Dorothée; Sonsma, Anja C. M.; Koeleman, Bobby P.; Lindhout, Dick; Weber, Yvonne G.; Lerche, Holger; Kapser, Claudia; Schankin, Christoph J.; Kunz, Wolfram S.; Surges, Rainer; Elger, Christian E.; Gaus, Verena; Schmitz, Bettina; Helbig, Ingo; Muhle, Hiltrud; Stephani, Ulrich; Klein, Karl M.; Rosenow, Felix; Neubauer, Bernd A.; Reinthaler, Eva M.; Zimprich, Fritz; Feucht, Martha; Møller, Rikke S.; Hjalgrim, Helle; De Jonghe, Peter; Suls, Arvid; Lieb, Wolfgang; Franke, Andre; Strauch, Konstantin; Gieger, Christian; Schurmann, Claudia; Schminke, Ulf; Nürnberg, Peter; Sander, Thomas

    2015-01-01

    Genetic generalised epilepsy (GGE) is the most common form of genetic epilepsy, accounting for 20% of all epilepsies. Genomic copy number variations (CNVs) constitute important genetic risk factors of common GGE syndromes. In our present genome-wide burden analysis, large (≥ 400 kb) and rare (< 1%) autosomal microdeletions with high calling confidence (≥ 200 markers) were assessed by the Affymetrix SNP 6.0 array in European case-control cohorts of 1,366 GGE patients and 5,234 ancestry-matched controls. We aimed to: 1) assess the microdeletion burden in common GGE syndromes, 2) estimate the relative contribution of recurrent microdeletions at genomic rearrangement hotspots and non-recurrent microdeletions, and 3) identify potential candidate genes for GGE. We found a significant excess of microdeletions in 7.3% of GGE patients compared to 4.0% in controls (P = 1.8 x 10-7; OR = 1.9). Recurrent microdeletions at seven known genomic hotspots accounted for 36.9% of all microdeletions identified in the GGE cohort and showed a 7.5-fold increased burden (P = 2.6 x 10-17) relative to controls. Microdeletions affecting either a gene previously implicated in neurodevelopmental disorders (P = 8.0 x 10-18, OR = 4.6) or an evolutionarily conserved brain-expressed gene related to autism spectrum disorder (P = 1.3 x 10-12, OR = 4.1) were significantly enriched in the GGE patients. Microdeletions found only in GGE patients harboured a high proportion of genes previously associated with epilepsy and neuropsychiatric disorders (NRXN1, RBFOX1, PCDH7, KCNA2, EPM2A, RORB, PLCB1). Our results demonstrate that the significantly increased burden of large and rare microdeletions in GGE patients is largely confined to recurrent hotspot microdeletions and microdeletions affecting neurodevelopmental genes, suggesting a strong impact of fundamental neurodevelopmental processes in the pathogenesis of common GGE syndromes. PMID:25950944

  20. Impact of hot and cold exposure on human skeletal muscle gene expression.

    PubMed

    Zak, Roksana B; Shute, Robert J; Heesch, Matthew W S; La Salle, D Taylor; Bubak, Matthew P; Dinan, Nicholas E; Laursen, Terence L; Slivka, Dustin R

    2017-03-01

    Many human diseases lead to a loss of skeletal muscle metabolic function and mass. Local and environmental temperature can modulate the exercise-stimulated response of several genes involved in mitochondrial biogenesis and skeletal muscle function in a human model. However, the impact of environmental temperature, independent of exercise, has not been addressed in a human model. Thus, the purpose of this study was to compare the effects of exposure to hot, cold, and room temperature conditions on skeletal muscle gene expression related to mitochondrial biogenesis and muscle mass. Recreationally trained male subjects (n = 12) had muscle biopsies taken from the vastus lateralis before and after 3 h of exposure to hot (33 °C), cold (7 °C), or room temperature (20 °C) conditions. Temperature had no effect on most of the genes related to mitochondrial biogenesis, myogenesis, or proteolysis (p > 0.05). Core temperature was significantly higher in hot and cold environments compared with room temperature (37.2 ± 0.1 °C, p = 0.001; 37.1 ± 0.1 °C, p = 0.013; 36.9 ± 0.1 °C, respectively). Whole-body oxygen consumption was also significantly higher in hot and cold compared with room temperature (0.38 ± 0.01 L·min(-1), p < 0.001; 0.52 ± 0.03 L·min(-1), p < 0.001; 0.35 ± 0.01 L·min(-1), respectively). In conclusion, these data show that acute temperature exposure alone does not elicit significant changes in skeletal muscle gene expression. When considered in conjunction with previous research, exercise appears to be a necessary component to observe gene expression alterations between different environmental temperatures in humans.

  1. Burden analysis of rare microdeletions suggests a strong impact of neurodevelopmental genes in genetic generalised epilepsies.

    PubMed

    Lal, Dennis; Ruppert, Ann-Kathrin; Trucks, Holger; Schulz, Herbert; de Kovel, Carolien G; Kasteleijn-Nolst Trenité, Dorothée; Sonsma, Anja C M; Koeleman, Bobby P; Lindhout, Dick; Weber, Yvonne G; Lerche, Holger; Kapser, Claudia; Schankin, Christoph J; Kunz, Wolfram S; Surges, Rainer; Elger, Christian E; Gaus, Verena; Schmitz, Bettina; Helbig, Ingo; Muhle, Hiltrud; Stephani, Ulrich; Klein, Karl M; Rosenow, Felix; Neubauer, Bernd A; Reinthaler, Eva M; Zimprich, Fritz; Feucht, Martha; Møller, Rikke S; Hjalgrim, Helle; De Jonghe, Peter; Suls, Arvid; Lieb, Wolfgang; Franke, Andre; Strauch, Konstantin; Gieger, Christian; Schurmann, Claudia; Schminke, Ulf; Nürnberg, Peter; Sander, Thomas

    2015-05-01

    Genetic generalised epilepsy (GGE) is the most common form of genetic epilepsy, accounting for 20% of all epilepsies. Genomic copy number variations (CNVs) constitute important genetic risk factors of common GGE syndromes. In our present genome-wide burden analysis, large (≥ 400 kb) and rare (< 1%) autosomal microdeletions with high calling confidence (≥ 200 markers) were assessed by the Affymetrix SNP 6.0 array in European case-control cohorts of 1,366 GGE patients and 5,234 ancestry-matched controls. We aimed to: 1) assess the microdeletion burden in common GGE syndromes, 2) estimate the relative contribution of recurrent microdeletions at genomic rearrangement hotspots and non-recurrent microdeletions, and 3) identify potential candidate genes for GGE. We found a significant excess of microdeletions in 7.3% of GGE patients compared to 4.0% in controls (P = 1.8 x 10-7; OR = 1.9). Recurrent microdeletions at seven known genomic hotspots accounted for 36.9% of all microdeletions identified in the GGE cohort and showed a 7.5-fold increased burden (P = 2.6 x 10-17) relative to controls. Microdeletions affecting either a gene previously implicated in neurodevelopmental disorders (P = 8.0 x 10-18, OR = 4.6) or an evolutionarily conserved brain-expressed gene related to autism spectrum disorder (P = 1.3 x 10-12, OR = 4.1) were significantly enriched in the GGE patients. Microdeletions found only in GGE patients harboured a high proportion of genes previously associated with epilepsy and neuropsychiatric disorders (NRXN1, RBFOX1, PCDH7, KCNA2, EPM2A, RORB, PLCB1). Our results demonstrate that the significantly increased burden of large and rare microdeletions in GGE patients is largely confined to recurrent hotspot microdeletions and microdeletions affecting neurodevelopmental genes, suggesting a strong impact of fundamental neurodevelopmental processes in the pathogenesis of common GGE syndromes.

  2. Characterisation of porin genes from Mycobacterium fortuitum and their impact on growth.

    PubMed

    Sharbati, Soroush; Schramm, Kira; Rempel, Sonja; Wang, Hwa; Andrich, Ronny; Tykiel, Verena; Kunisch, Ralph; Lewin, Astrid

    2009-02-09

    Highly pathogenic mycobacteria like Mycobacterium tuberculosis are characterised by their slow growth and their ability to reside and multiply in the very hostile phagosomal environment and a correlation between the growth rate of mycobacteria and their pathogenicity has been hypothesised. Here, porin genes from M. fortuitum were cloned and characterised to address their impact on the growth rate of fast-growing and pathogenic mycobacteria. Two genes encoding porins orthologous to MspA from M. smegmatis, porM1 and porM2, were cloned from M. fortuitum strains, which were originally isolated from human patients. Both porin genes were at least partially able to complement the mutations of a M. smegmatis mutant strain lacking the genes mspA and mspC with respect to the growth rate. PorM1 and porM2 were present in different strains of M. fortuitum including the type strain. Comparative expression analysis of porM genes revealed divergent porin expression among analysed M. fortuitum strains. Repression of the expression of porins by antisense technique decreased the growth rates of different M. fortuitum. The effects of over-expression of porM1 as well as porM2 varied depending on the strain and the concentration of antibiotic added to the medium and indicated that PorM1 and PorM2 enhance the growth of M. fortuitum strains, but also the diffusion of the antibiotic kanamycin into the cells. This study demonstrates the important role of porin expression in growth as well as antibiotic susceptibility of the opportunistic bacterium M. fortuitum.

  3. Widespread Impact of Chromosomal Inversions on Gene Expression Uncovers Robustness via Phenotypic Buffering

    PubMed Central

    Naseeb, Samina; Carter, Zorana; Minnis, David; Donaldson, Ian; Zeef, Leo; Delneri, Daniela

    2016-01-01

    The nonrandom gene organization in eukaryotes plays a significant role in genome evolution and function. Chromosomal structural changes impact meiotic fitness and, in several organisms, are associated with speciation and rapid adaptation to different environments. Small sized chromosomal inversions, encompassing few genes, are pervasive in Saccharomyces “sensu stricto” species, while larger inversions are less common in yeasts compared with higher eukaryotes. To explore the effect of gene order on phenotype, reproductive isolation, and gene expression, we engineered 16 Saccharomyces cerevisiae strains carrying all possible paracentric and pericentric inversions between Ty1 elements, a natural substrate for rearrangements. We found that 4 inversions were lethal, while the other 12 did not show any fitness advantage or disadvantage in rich and minimal media. At meiosis, only a weak negative correlation with fitness was seen with the size of the inverted region. However, significantly lower fertility was seen in heterozygote invertant strains carrying recombination hotspots within the breakpoints. Altered transcription was observed throughout the genome rather than being overrepresented within the inversions. In spite of the large difference in gene expression in the inverted strains, mitotic fitness was not impaired in the majority of the 94 conditions tested, indicating that the robustness of the expression network buffers the deleterious effects of structural changes in several environments. Overall, our results support the notion that transcriptional changes may compensate for Ty-mediated rearrangements resulting in the maintenance of a constant phenotype, and suggest that large inversions in yeast are unlikely to be a selectable trait during vegetative growth. PMID:26929245

  4. Distribution patterns and impact of transposable elements in genes of green algae.

    PubMed

    Philippsen, Gisele S; Avaca-Crusca, Juliana S; Araujo, Ana P U; DeMarco, Ricardo

    2016-12-05

    Transposable elements (TEs) are DNA sequences able to transpose in the host genome, a remarkable feature that enables them to influence evolutive trajectories of species. An investigation about the TE distribution and TE impact in different gene regions of the green algae species Chlamydomonas reinhardtii and Volvox carteri was performed. Our results indicate that TEs are very scarce near introns boundaries, suggesting that insertions in this region are negatively selected. This contrasts with previous results showing enrichment of tandem repeats in introns boundaries and suggests that different evolutionary forces are acting in these different classes of repeats. Despite the relatively low abundance of TEs in the genome of green algae when compared to mammals, the proportion of poly(A) sites derived from TEs found in C. reinhardtii was similar to that described in human and mice. This fact, associated with the enrichment of TEs in gene 5' and 3' flanks of C. reinhardtii, opens up the possibility that TEs may have considerably contributed for gene regulatory sequences evolution in this species. Moreover, it was possible identify several instances of TE exonization for C. reinhardtii, with a particularly interesting case from a gene coding for Condensin II, a protein involved in the maintenance of chromosomal structure, where the addition of a transposomal PHD finger may contribute to binding specificity of this protein. Taken together, our results suggest that the low abundance of TEs in green algae genomes is correlated with a strict negative selection process, combined with the retention of copies that contribute positively with gene structures. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Impact of Hot and Cold Exposure on Human Skeletal Muscle Gene Expression

    PubMed Central

    Zak, Roksana B.; Shute, Robert J.; Heesch, Matthew W.S.; La Salle, D. Taylor; Bubak, Matthew P.; Dinan, Nicholas E.; Laursen, Terence L.; Slivka, Dustin R.

    2017-01-01

    Many human diseases lead to a loss of skeletal muscle metabolic function and mass. Local and environmental temperature can modulate the exercise-stimulated response of several genes involved in mitochondrial biogenesis and skeletal muscle function in a human model. However, the impact of environmental temperature, independent of exercise, has not been addressed in a human model. Thus, the purpose of this study was to compare the effects of exposure to hot, cold, and room temperature conditions on skeletal muscle gene expression related to mitochondrial biogenesis and muscle mass. METHODS Recreationally trained male subjects (n=12) had muscle biopsies taken from the vastus lateralis before and after 3 h exposure to hot (33 °C), cold (7 °C), or room temperature (20 °C) conditions. RESULTS Temperature had no effect on most of the genes related to mitochondrial biogenesis, myogenesis, or proteolysis (p > 0.05). Core temperature was significantly higher in hot and cold environments compared to room temperature (37.2 ± 0.1 °C, p = 0.001; 37.1 ± 0.1 °C, p = 0.013; 36.9 ± 0.1 °C, respectively). Whole body oxygen consumption was also significantly higher in hot and cold compared to room temperature (0.38 ± 0.01 L·min−1, p < 0.001; 0.52 ± 0.03 L·min−1, p < 0.001; 0.35 ± 0.01 L·min−1, respectively). CONCLUSIONS These data show that acute temperature exposure alone does not elicit significant changes in skeletal muscle gene expression. When considered in conjunction with previous research, exercise appears to be a necessary component to observe gene expression alterations between different environmental temperatures in humans. PMID:28177744

  6. Characterisation of porin genes from Mycobacterium fortuitum and their impact on growth

    PubMed Central

    2009-01-01

    Background Highly pathogenic mycobacteria like Mycobacterium tuberculosis are characterised by their slow growth and their ability to reside and multiply in the very hostile phagosomal environment and a correlation between the growth rate of mycobacteria and their pathogenicity has been hypothesised. Here, porin genes from M. fortuitum were cloned and characterised to address their impact on the growth rate of fast-growing and pathogenic mycobacteria. Results Two genes encoding porins orthologous to MspA from M. smegmatis, porM1 and porM2, were cloned from M. fortuitum strains, which were originally isolated from human patients. Both porin genes were at least partially able to complement the mutations of a M. smegmatis mutant strain lacking the genes mspA and mspC with respect to the growth rate. PorM1 and porM2 were present in different strains of M. fortuitum including the type strain. Comparative expression analysis of porM genes revealed divergent porin expression among analysed M. fortuitum strains. Repression of the expression of porins by antisense technique decreased the growth rates of different M. fortuitum. The effects of over-expression of porM1 as well as porM2 varied depending on the strain and the concentration of antibiotic added to the medium and indicated that PorM1 and PorM2 enhance the growth of M. fortuitum strains, but also the diffusion of the antibiotic kanamycin into the cells. Conclusion This study demonstrates the important role of porin expression in growth as well as antibiotic susceptibility of the opportunistic bacterium M. fortuitum. PMID:19203364

  7. Diversity and Impact of Rare Variants in Genes Encoding the Platelet G Protein-Coupled Receptors

    PubMed Central

    Jones, Matthew L.; Norman, Jane E.; Morgan, Neil V.; Mundell, Stuart J.; Lordkipanidzé, Marie; Lowe, Gillian C.; Daly, Martina E.; Simpson, Michael A.; Drake, Sian; Watson, Steve P.

    2015-01-01

    Summary Platelet responses to activating agonists are influenced by common population variants within or near G protein-coupled receptor (GPCR) genes that affect receptor activity. However, the impact of rare GPCR gene variants is unknown. We describe the rare single nucleotide variants (SNVs) in the coding and splice regions of 18 GPCR genes in 7,595 exomes from the 1,000-genomes and Exome Sequencing Project databases and in 31 cases with inherited platelet function disorders (IPFDs). In the population databases, the GPCR gene target regions contained 740 SNVs (318 synonymous, 410 missense, 7 stop gain and 6 splice region) of which 70% had global minor allele frequency (MAF) < 0.05%. Functional annotation using six computational algorithms, experimental evidence and structural data identified 156/740 (21%) SNVs as potentially damaging to GPCR function, most commonly in regions encoding the transmembrane and C-terminal intracellular receptor domains. In 31 index cases with IPFDs (Gi-pathway defect n=15; secretion defect n=11; thromboxane pathway defect n=3 and complex defect n=2) there were 256 SNVs in the target regions of 15 stimulatory platelet GPCRs (34 unique; 12 with MAF<1% and 22 with MAF ≥ 1%). These included rare variants predicting R122H, P258T and V207A substitutions in the P2Y12 receptor that were annotated as potentially damaging, but only partially explained the platelet function defects in each case. Our data highlight that potentially damaging variants in platelet GPCR genes have low individual frequencies, but are collectively abundant in the population. Potentially damaging variants are also present in pedigrees with IPFDs and may contribute to complex laboratory phenotypes. PMID:25567036

  8. Diversity and impact of rare variants in genes encoding the platelet G protein-coupled receptors.

    PubMed

    Jones, Matthew L; Norman, Jane E; Morgan, Neil V; Mundell, Stuart J; Lordkipanidzé, Marie; Lowe, Gillian C; Daly, Martina E; Simpson, Michael A; Drake, Sian; Watson, Steve P; Mumford, Andrew D

    2015-04-01

    Platelet responses to activating agonists are influenced by common population variants within or near G protein-coupled receptor (GPCR) genes that affect receptor activity. However, the impact of rare GPCR gene variants is unknown. We describe the rare single nucleotide variants (SNVs) in the coding and splice regions of 18 GPCR genes in 7,595 exomes from the 1,000-genomes and Exome Sequencing Project databases and in 31 cases with inherited platelet function disorders (IPFDs). In the population databases, the GPCR gene target regions contained 740 SNVs (318 synonymous, 410 missense, 7 stop gain and 6 splice region) of which 70 % had global minor allele frequency (MAF) < 0.05 %. Functional annotation using six computational algorithms, experimental evidence and structural data identified 156/740 (21 %) SNVs as potentially damaging to GPCR function, most commonly in regions encoding the transmembrane and C-terminal intracellular receptor domains. In 31 index cases with IPFDs (Gi-pathway defect n=15; secretion defect n=11; thromboxane pathway defect n=3 and complex defect n=2) there were 256 SNVs in the target regions of 15 stimulatory platelet GPCRs (34 unique; 12 with MAF< 1 % and 22 with MAF≥ 1 %). These included rare variants predicting R122H, P258T and V207A substitutions in the P2Y12 receptor that were annotated as potentially damaging, but only partially explained the platelet function defects in each case. Our data highlight that potentially damaging variants in platelet GPCR genes have low individual frequencies, but are collectively abundant in the population. Potentially damaging variants are also present in pedigrees with IPFDs and may contribute to complex laboratory phenotypes.

  9. Chemical dispersant potentiates crude oil impacts on growth, reproduction, and gene expression in Caenorhabditis elegans.

    PubMed

    Zhang, Yanqiong; Chen, Dongliang; Ennis, Adrien C; Polli, Joseph R; Xiao, Peng; Zhang, Baohong; Stellwag, Edmund J; Overton, Anthony; Pan, Xiaoping

    2013-02-01

    The economic, environmental, and human health impacts of the deepwater horizon (DWH) oil spill have been of significant concern in the general public and among scientists. This study employs parallel experiments to test the effects of crude oil from the DWH oil well, chemical dispersant Corexit 9500A, and dispersant-oil mixture on growth and reproduction in the model organism Caenorhabditis elegans. Both the crude oil and the dispersant significantly inhibited the reproduction of C. elegans. Dose-dependent inhibitions of hatched larvae production were observed in worms exposed to both crude oil and dispersant. Importantly, the chemical dispersant Corexit 9500A potentiated crude oil effects; dispersant-oil mixture induced more significant effects than oil or dispersant-alone exposures. While oil-alone exposure and dispersant-alone exposure have none to moderate inhibitory effects on hatched larvae production, respectively, the mixture of dispersant and oil induced much more significant inhibition of offspring production. The production of hatched larvae was almost completely inhibited by several high concentrations of the dispersant-oil mixture. This suggests a sensitive bioassay for future investigation of oil/dispersant impacts on organisms. We also investigated the effects of crude oil/dispersant exposure at the molecular level by measuring the expressions of 31 functional genes. Results showed that the dispersant and the dispersant-oil mixture induced aberrant expressions of 12 protein-coding genes (cat-4, trxr-2, sdhb-1, lev-8, lin-39, unc-115, prdx-3, sod-1, acr-16, ric-3, unc-68, and acr-8). These 12 genes are associated with a variety of biological processes, including egg-laying, oxidative stress, muscle contraction, and neurological functions. In summary, the toxicity potentiating effect of chemical dispersant must be taken into consideration in future crude oil cleanup applications.

  10. Temperature and Development Impacts on Housekeeping Gene Expression in Cowpea Aphid, Aphis craccivora (Hemiptera: Aphidiae)

    PubMed Central

    Liu, Yong; Zhou, Xuguo

    2015-01-01

    Quantitative real-time PCR (qRT-PCR) is a powerful technique to quantify gene expression. To standardize gene expression studies and obtain more accurate qRT-PCR analysis, normalization relative to consistently expressed housekeeping genes (HKGs) is required. In this study, ten candidate HKGs including elongation factor 1 α (EF1A), ribosomal protein L11 (RPL11), ribosomal protein L14 (RPL14), ribosomal protein S8 (RPS8), ribosomal protein S23 (RPS23), NADH-ubiquinone oxidoreductase (NADH), vacuolar-type H+-ATPase (ATPase), heat shock protein 70 (HSP70), 18S ribosomal RNA (18S), and 12S ribosomal RNA (12S) from the cowpea aphid, Aphis craccivora Koch were selected. Four algorithms, geNorm, Normfinder, BestKeeper, and the ΔCt method were employed to evaluate the expression profiles of these HKGs as endogenous controls across different developmental stages and temperature regimes. Based on RefFinder, which integrates all four analytical algorithms to compare and rank the candidate HKGs, RPS8, RPL14, and RPL11 were the three most stable HKGs across different developmental stages and temperature conditions. This study is the first step to establish a standardized qRT-PCR analysis in A. craccivora following the MIQE guideline. Results from this study lay a foundation for the genomics and functional genomics research in this sap-sucking insect pest with substantial economic impact. PMID:26090683

  11. Interactions between DNA and gemini surfactant: impact on gene therapy: part II.

    PubMed

    Ahmed, Taksim; Kamel, Amany O; Wettig, Shawn D

    2016-02-01

    Nonviral gene delivery, provides distinct treatment modalities for the inherited and acquired diseases, relies upon the encapsulation of a gene of interest, which is then ideally delivered to the target cells. Variations in the chemical structure of gemini surfactants and subsequent physicochemical characteristics of the gemini-based lipoplexes and their impact on efficient gene transfection were assessed in part I, which was published in first March 2016 issue of Nanomedicine (1103). In order to design an efficient vector using gemini surfactants, the interaction of the surfactant with DNA and other components of the delivery system must be characterized, and more critically, well understood. Such studies will help to understand how nonviral transfection complexes, in general, overcome various cellular barriers. The Langmuir-Blodgett monolayer studies, atomic force microscopy, differential scanning calorimetry, isothermal titration calorimetry, small-angle x-ray scattering, are extensively used to evaluate the interaction behavior of gemini surfactants with DNA and other vector components. Part II of this review focuses on the use of these unique techniques to understand their interaction with DNA.

  12. Phevalin (aureusimine B) production by Staphylococcus aureus biofilm and impacts on human keratinocyte gene expression.

    PubMed

    Secor, Patrick R; Jennings, Laura K; James, Garth A; Kirker, Kelly R; Pulcini, Elinor Delancey; McInnerney, Kate; Gerlach, Robin; Livinghouse, Tom; Hilmer, Jonathan K; Bothner, Brian; Fleckman, Philip; Olerud, John E; Stewart, Philip S

    2012-01-01

    Staphylococcus aureus biofilms are associated with chronic skin infections and are orders of magnitude more resistant to antimicrobials and host responses. S. aureus contains conserved nonribosomal peptide synthetases that produce the cyclic dipeptides tyrvalin and phevalin (aureusimine A and B, respectively). The biological function of these compounds has been speculated to be involved in virulence factor gene expression in S. aureus, protease inhibition in eukaryotic cells, and interspecies bacterial communication. However, the exact biological role of these compounds is unknown. Here, we report that S. aureus biofilms produce greater amounts of phevalin than their planktonic counterparts. Phevalin had no obvious impact on the extracellular metabolome of S. aureus as measured by high-performance liquid chromatography-mass spectrometry and nuclear magnetic resonance. When administered to human keratinocytes, phevalin had a modest effect on gene expression. However, conditioned medium from S. aureus spiked with phevalin amplified differences in keratinocyte gene expression compared to conditioned medium alone. Phevalin may be exploited as potential biomarker and/or therapeutic target for chronic, S. aureus biofilm-based infections.

  13. Using Blood Informative Transcripts in Geographical Genomics: Impact of Lifestyle on Gene Expression in Fijians

    PubMed Central

    Nath, Artika Praveeta; Arafat, Dalia; Gibson, Greg

    2012-01-01

    In previous geographical genomics studies of the impact of lifestyle on gene expression inferred from microarray analysis of peripheral blood samples, we described the complex influences of culture, ethnicity, and gender in Morocco, and of pregnancy in Brisbane. Here we describe the use of nanofluidic Fluidigm quantitative RT-PCR arrays targeted at a set of 96 transcripts that are broadly informative of the major axes of immune gene expression, to explore the population structure of transcription in Fiji. As in Morocco, major differences are seen between the peripheral blood transcriptomes of rural villagers and residents of the capital city, Suva. The effect is much greater in Indian villages than in Melanesian highlanders and appears to be similar with respect to the nature of at least two axes of variation. Gender differences are much smaller than ethnicity or lifestyle effects. Body mass index is shown to associate with one of the axes as it does in Atlanta and Brisbane, establishing a link between the epidemiological transition of human metabolic disease, and gene expression profiles. PMID:23162571

  14. Growth of Yersinia pseudotuberculosis in human plasma: impacts on virulence and metabolic gene expression

    PubMed Central

    Rosso, Marie-Laure; Chauvaux, Sylvie; Dessein, Rodrigue; Laurans, Caroline; Frangeul, Lionel; Lacroix, Céline; Schiavo, Angèle; Dillies, Marie-Agnès; Foulon, Jeannine; Coppée, Jean-Yves; Médigue, Claudine; Carniel, Elisabeth; Simonet, Michel; Marceau, Michaël

    2008-01-01

    Background In man, infection by the Gram-negative enteropathogen Yersinia pseudotuberculosis is usually limited to the terminal ileum. However, in immunocompromised patients, the microorganism may disseminate from the digestive tract and thus cause a systemic infection with septicemia. Results To gain insight into the metabolic pathways and virulence factors expressed by the bacterium at the blood stage of pseudotuberculosis, we compared the overall gene transcription patterns (the transcriptome) of bacterial cells cultured in either human plasma or Luria-Bertani medium. The most marked plasma-triggered metabolic consequence in Y. pseudotuberculosis was the switch to high glucose consumption, which is reminiscent of the acetogenic pathway (known as "glucose overflow") in Escherichia coli. However, upregulation of the glyoxylate shunt enzymes suggests that (in contrast to E. coli) acetate may be further metabolized in Y. pseudotuberculosis. Our data also indicate that the bloodstream environment can regulate major virulence genes (positively or negatively); the yadA adhesin gene and most of the transcriptional units of the pYV-encoded type III secretion apparatus were found to be upregulated, whereas transcription of the pH6 antigen locus was strongly repressed. Conclusion Our results suggest that plasma growth of Y. pseudotuberculosis is responsible for major transcriptional regulatory events and prompts key metabolic reorientations within the bacterium, which may in turn have an impact on virulence. PMID:19055764

  15. The Differential Impact of Oxytocin Receptor Gene in Violence-Exposed Boys and Girls.

    PubMed

    Merrill, Livia C; Jones, Christopher W; Drury, Stacy S; Theall, Katherine P

    2017-03-21

    Childhood violence exposure is a prevalent public health problem. Understanding the lasting impact of violence requires an enhanced appreciation for the complex effects of violence across behavioral, physiologic, and molecular outcomes. This subject matched, cross-sectional study of 80 children explored the impact of violence exposure across behavioral, physiologic, and cellular outcomes. Externalizing behavior, diurnal cortisol rhythm, and telomere length (TL) were examined in a community recruited cohort of Black youth. Given evidence that genetic variation contributes to individual differences in response to the environment, we further tested whether a polymorphism in the oxytocin receptor gene (OXTR rs53576) moderated associations between violence and youth outcomes. Exposure to violence was directly associated with increased externalizing behavior, but no direct association of violence was found with cortisol or TL. Oxytocin genotype, however, moderated the association between violence and both cortisol and TL, suggesting that pathways linked to oxytocin may contribute to individual differences in the physiologic and molecular consequences of violence exposure. Sex differences with OXTR in cortisol and TL outcomes were also detected. Taken together, these findings suggest that there are complex pathways through which violence exposure impacts children, and that these pathways differ by both genetic variation and the sex of the child.

  16. Rat Models of Cardiovascular Disease Demonstrate Distinctive Pulmonary Gene Expressions for Vascular Response Genes: Impact of Ozone Exposure

    EPA Science Inventory

    Comparative gene expression profiling of multiple tissues from rat strains with genetic predisposition to diverse cardiovascular diseases (CVD) can help decode the transcriptional program that governs organ-specific functions. We examined expressions of CVD genes in the lungs of ...

  17. Rat Models of Cardiovascular Disease Demonstrate Distinctive Pulmonary Gene Expressions for Vascular Response Genes: Impact of Ozone Exposure

    EPA Science Inventory

    Comparative gene expression profiling of multiple tissues from rat strains with genetic predisposition to diverse cardiovascular diseases (CVD) can help decode the transcriptional program that governs organ-specific functions. We examined expressions of CVD genes in the lungs of ...

  18. The autism risk genes MET and PLAUR differentially impact cortical development

    PubMed Central

    Eagleson, Kathie L.; Campbell, Daniel B.; Thompson, Barbara L.; Bergman, Mica Y.; Levitt, Pat

    2013-01-01

    Met signaling in cortical interneuron development in vitro and a reduction of these neurons in uPAR (mouse ortholog of PLAUR) null mice, suggesting that disruption of either gene impacts cortical development similarly. Here, we modify this conclusion, reporting that interneuron numbers are unchanged in the neocortex of Metfx/fx/ Dlx5/6cre mice, in which Met is ablated from cells arising from the ventral telencephalon (VTel). Consistent with this, Met transcript is not detected in the VTel during interneuron genesis and migration; furthermore, during the postnatal period of interneuron maturation, Met is co-expressed in glutamatergic projection neurons, but not interneurons. Low levels of Met protein are expressed in the VTel at E12.5 and E14.5, likely reflecting the arrival of Met-containing corticofugal axons. Met expression, however, is induced in E12.5 VTel cells after 2 days in vitro, perhaps underlying discrepancies between observations in vitro and in Metfx/fx/ Dlx5/6cre mice. We suggest that, in vivo, Met impacts the development of cortical projection neurons, whereas uPAR influences interneuron maturation. An altered balance between excitation and inhibition has been postulated as a biological mechanism for ASD; this imbalance could arise from different risk genes differentially affecting either or both elements. PMID:21328570

  19. Memory acquisition and retrieval impact different epigenetic processes that regulate gene expression

    PubMed Central

    2015-01-01

    Background A fundamental question in neuroscience is how memories are stored and retrieved in the brain. Long-term memory formation requires transcription, translation and epigenetic processes that control gene expression. Thus, characterizing genome-wide the transcriptional changes that occur after memory acquisition and retrieval is of broad interest and importance. Genome-wide technologies are commonly used to interrogate transcriptional changes in discovery-based approaches. Their ability to increase scientific insight beyond traditional candidate gene approaches, however, is usually hindered by batch effects and other sources of unwanted variation, which are particularly hard to control in the study of brain and behavior. Results We examined genome-wide gene expression after contextual conditioning in the mouse hippocampus, a brain region essential for learning and memory, at all the time-points in which inhibiting transcription has been shown to impair memory formation. We show that most of the variance in gene expression is not due to conditioning and that by removing unwanted variance through additional normalization we are able provide novel biological insights. In particular, we show that genes downregulated by memory acquisition and retrieval impact different functions: chromatin assembly and RNA processing, respectively. Levels of histone 2A variant H2AB are reduced only following acquisition, a finding we confirmed using quantitative proteomics. On the other hand, splicing factor Rbfox1 and NMDA receptor-dependent microRNA miR-219 are only downregulated after retrieval, accompanied by an increase in protein levels of miR-219 target CAMKIIγ. Conclusions We provide a thorough characterization of coding and non-coding gene expression during long-term memory formation. We demonstrate that unwanted variance dominates the signal in transcriptional studies of learning and memory and introduce the removal of unwanted variance through normalization as a

  20. Impact of host gene polymorphisms on susceptibility to chronic hepatitis B virus infection.

    PubMed

    Moudi, Bita; Heidari, Zahra; Mahmoudzadeh-Sagheb, Hamidreza

    2016-10-01

    Hepatitis B virus (HBV) infection can result in a number of different clinical conditions, including asymptomatic HBV carriers to chronic hepatitis and primary hepatocellular carcinoma. Variations in cytokine genes have been discussed to affect the natural history of HBV infection. These cytokines may involve in the viral binding to the cells, modulating the host immune response to infection and pathological changes in the liver, and affecting the antiviral therapies. Various studies reveal that SNPs play an important role in pathogenesis of HBV. On the other hand, various outcomes of infection cannot be completely shown by genetic factors because these studies have inconsistent results with regard to the possible impacts of host genetic polymorphisms on susceptibility to infection. Therefore, to identify the real effects of host genetic factors in HBV susceptibility and natural history of the disease, studies with large sample size will be needed. In addition, due to the complex interactions of genetic factors it is better to identify synergies of several SNPs. Such studies can provide better insights into the novel methods of diagnosis and treatment. Current review will discuss significant genetic variations in cytokine genes that may affect the susceptibility to the chronic HBV infection. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. The Impact of Gene Polymorphisms on the Success of Anticholinergic Treatment in Children with Overactive Bladder

    PubMed Central

    Konac, Ece; Ure, Iyimser; Senol, Cem; Onen, Ilke Hacer; Sozen, Sinan; Menevse, Adnan

    2015-01-01

    Aim. To determine the impact of gene polymorphisms on detrusor contraction-relaxation harmony in children with lower urinary tract symptoms (LUTS). Materials and Methods. Toilet trained children older than 5 years of age with LUTS and normal neurological examination underwent videourodynamic study. The control group was composed of age matched children with no voiding complaints. The study group who filled out the voiding dysfunction symptom score before and after the treatment received standard oxybutynin treatment and was reevaluated 1 year after treatment. Genomic DNA was isolated from all patients and subjected to PCR for amplification. Genotyping of ARGHEF10, ROCK2, ADRB3, and CYP3A4 was carried out with Polymerase Chain Reaction- Restriction Fragment Length Polymorphism (PCR-RFLP) method. Results. 34 (45%) and 42 (55%) patients were enrolled in the study and control group, respectively. ARGEF10 GG, ADRB3 TC, and CYP3A4 AG genotype patients displayed insignificant difference between pre- and posttreatment voiding dysfunction symptom score and bladder volumes. Conclusions. The polymorphism of genes in the cholinergic pathway did not significantly differ clinical parameters. On the other hand, polymorphic patients in the adrenergic pathway seemed to suffer from clinical disappointment. For this reason, we think that the neglected adrenergic pathway could be a new therapeutic target for the treatment of anticholinergic resistant LUTS in children. PMID:26166934

  2. Low-level copy number changes of MYC genes have a prognostic impact in medulloblastoma.

    PubMed

    Zitterbart, Karel; Filkova, Hana; Tomasikova, Lenka; Necesalova, Eva; Zambo, Iva; Kantorova, Dagmar; Slamova, Iva; Vranova, Vladimira; Zezulkova, Dita; Pesakova, Martina; Pavelka, Zdenek; Veselska, Renata; Kuglik, Petr; Sterba, Jaroslav

    2011-03-01

    High-level amplifications of MYC genes are associated with poor outcomes in childhood medulloblastoma (MB). However, the occurrence of MYCN and MYCC copy number increases below the intense amplification pattern is rarely reported, and its clinical impact has not yet been determined. Here, we describe this phenomenon and its prognostic significance in a cohort of 29 MB patients. Using interphase fluorescence in situ hybridization (I-FISH), low-level copy number alterations, i.e. gain of MYCN, were shown in 5/27 (19%) samples, whereas amplification was revealed in only 1/27 (4%) samples. MYCC gain was revealed in 6/29 (21%) MB, while amplification was disclosed in only 2/29 (7%). Hyperploidy and co-incidence of gains in both MYC loci were frequently observed in samples with copy number aberrations. Survival analysis has clearly shown that MYC copy number increases are associated with lowered event-free survival and overall survival in MB. In the case of MYCN, this negative correlation was statistically significant. We conclude that limited numerical alterations in loci 2p24 (MYCN) and 8q24 (MYCC), as assessed by I-FISH, are present in MB with a higher frequency than high-level amplifications. Poor prognoses were observed in patients with copy number increases in MYC genes. Our data illustrate the importance of further investigations in multicenter trials to better refine the emerging genomic-based prognostic stratification in MB.

  3. Plasmapheresis eliminates the negative impact of AAV antibodies on microdystrophin gene expression following vascular delivery.

    PubMed

    Chicoine, L G; Montgomery, C L; Bremer, W G; Shontz, K M; Griffin, D A; Heller, K N; Lewis, S; Malik, V; Grose, W E; Shilling, C J; Campbell, K J; Preston, T J; Coley, B D; Martin, P T; Walker, C M; Clark, K R; Sahenk, Z; Mendell, J R; Rodino-Klapac, L R

    2014-02-01

    Duchenne muscular dystrophy is a monogenic disease potentially treatable by gene replacement. Use of recombinant adeno-associated virus (AAV) will ultimately require a vascular approach to broadly transduce muscle cells. We tested the impact of preexisting AAV antibodies on microdystrophin expression following vascular delivery to nonhuman primates. Rhesus macaques were treated by isolated limb perfusion using a fluoroscopically guided catheter. In addition to serostatus stratification, the animals were placed into one of the three immune suppression groups: no immune suppression, prednisone, and triple immune suppression (prednisone, tacrolimus, and mycophenolate mofetil). The animals were analyzed for transgene expression at 3 or 6 months. Microdystrophin expression was visualized in AAV, rhesus serotype 74 sero-negative animals (mean: 48.0 ± 20.8%) that was attenuated in sero-positive animals (19.6 ± 18.7%). Immunosuppression did not affect transgene expression. Importantly, removal of AAV binding antibodies by plasmapheresis in AAV sero-positive animals resulted in high-level transduction (60.8 ± 18.0%), which is comparable with that of AAV sero-negative animals (53.7 ± 7.6%), whereas non-pheresed sero-positive animals demonstrated significantly lower transduction levels (10.1 ± 6.0%). These data support the hypothesis that removal of AAV binding antibodies by plasmapheresis permits successful and sustained gene transfer in the presence of preexisting immunity (natural infection) to AAV.

  4. Impact of Enriched Environment on Murine T Cell Differentiation and Gene Expression Profile

    PubMed Central

    Rattazzi, Lorenza; Piras, Giuseppa; Brod, Samuel; Smith, Koval; Ono, Masahiro; D’Acquisto, Fulvio

    2016-01-01

    T cells are known to be plastic and to change their phenotype according to the cellular and biochemical milieu they are embedded in. In this study, we transposed this concept at a macroscopic level assessing whether changes in the environmental housing conditions of C57/BL6 mice would influence the phenotype and function of T cells. Our study shows that exposure to 2 weeks in an enriched environment (EE) does not impact the T cell repertoire in vivo and causes no changes in the early TCR-driven activation events of these cells. Surprisingly, however, T cells from enriched mice showed a unique T helper effector cell phenotype upon differentiation in vitro. This was featured by a significant reduction in their ability to produce IFN-γ and by an increased release of IL-10 and IL-17. Microarray analysis of these cells also revealed a unique gene fingerprint with key signaling pathways involved in autoimmunity being modulated. Together, our results provide first evidence for a specific effect of EE on T cell differentiation and its associated changes in gene expression profile. In addition, our study sheds new light on the possible mechanisms by which changes in environmental factors can significantly influence the immune response of the host and favor the resolution of the inflammatory response. PMID:27746779

  5. Deletion of Lipoteichoic Acid Synthase Impacts Expression of Genes Encoding Cell Surface Proteins in Lactobacillus acidophilus.

    PubMed

    Selle, Kurt; Goh, Yong J; Johnson, Brant R; O'Flaherty, Sarah; Andersen, Joakim M; Barrangou, Rodolphe; Klaenhammer, Todd R

    2017-01-01

    Lactobacillus acidophilus NCFM is a well-characterized probiotic microorganism, supported by a decade of genomic and functional phenotypic investigations. L. acidophilus deficient in lipoteichoic acid (LTA), a major immunostimulant in Gram-positive bacteria, has been shown to shift immune system responses in animal disease models. However, the pleiotropic effects of removing LTA from the cell surface in lactobacilli are unknown. In this study, we surveyed the global transcriptional and extracellular protein profiles of two strains of L. acidophilus deficient in LTA. Twenty-four differentially expressed genes specific to the LTA-deficient strains were identified, including a predicted heavy metal resistance operon and several putative peptidoglycan hydrolases. Cell morphology and manganese sensitivity phenotypes were assessed in relation to the putative functions of differentially expressed genes. LTA-deficient L. acidophilus exhibited elongated cellular morphology and their growth was severely inhibited by elevated manganese concentrations. Exoproteomic surveys revealed distinct changes in the composition and relative abundances of several extracellular proteins and showed a bias of intracellular proteins in LTA-deficient strains of L. acidophilus. Taken together, these results elucidate the impact of ltaS deletion on the transcriptome and extracellular proteins of L. acidophilus, suggesting roles of LTA in cell morphology and ion homeostasis as a structural component of the Gram positive cell wall.

  6. Deletion of Lipoteichoic Acid Synthase Impacts Expression of Genes Encoding Cell Surface Proteins in Lactobacillus acidophilus

    PubMed Central

    Selle, Kurt; Goh, Yong J.; Johnson, Brant R.; O’Flaherty, Sarah; Andersen, Joakim M.; Barrangou, Rodolphe; Klaenhammer, Todd R.

    2017-01-01

    Lactobacillus acidophilus NCFM is a well-characterized probiotic microorganism, supported by a decade of genomic and functional phenotypic investigations. L. acidophilus deficient in lipoteichoic acid (LTA), a major immunostimulant in Gram-positive bacteria, has been shown to shift immune system responses in animal disease models. However, the pleiotropic effects of removing LTA from the cell surface in lactobacilli are unknown. In this study, we surveyed the global transcriptional and extracellular protein profiles of two strains of L. acidophilus deficient in LTA. Twenty-four differentially expressed genes specific to the LTA-deficient strains were identified, including a predicted heavy metal resistance operon and several putative peptidoglycan hydrolases. Cell morphology and manganese sensitivity phenotypes were assessed in relation to the putative functions of differentially expressed genes. LTA-deficient L. acidophilus exhibited elongated cellular morphology and their growth was severely inhibited by elevated manganese concentrations. Exoproteomic surveys revealed distinct changes in the composition and relative abundances of several extracellular proteins and showed a bias of intracellular proteins in LTA-deficient strains of L. acidophilus. Taken together, these results elucidate the impact of ltaS deletion on the transcriptome and extracellular proteins of L. acidophilus, suggesting roles of LTA in cell morphology and ion homeostasis as a structural component of the Gram positive cell wall. PMID:28443071

  7. The impact of H63D HFE gene carriage on hemoglobin and iron status in children.

    PubMed

    Barbara, Kaczorowska-Hac; Marcin, Luszczyk; Jedrzej, Antosiewicz; Wieslaw, Ziolkowski; Elzbieta, Adamkiewicz-Drozynska; Malgorzata, Mysliwiec; Ewa, Milosz; Jacek, Kaczor Jan

    2016-12-01

    The molecular mechanism that regulates iron homeostasis is based on a network of signals, which reflect on the iron requirements of the body. Hereditary hemochromatosis is a heterogenic metabolic syndrome which is due to unchecked transfer of iron into the bloodstream and its toxic effects on parenchymatous organs. It is caused by the mutation of genes that encode proteins that help hepcidin to monitor serum iron. These proteins include the human hemochromatosis protein -HFE, transferrin-receptor 2, hemojuvelin in rare instances, and ferroportin. HFE-related hemochromatosis is the most frequent form of the disease. Interestingly, the low penetrance of polymorphic HFE genes results in rare clinical presentation of the disease, predominantly in middle-aged males. Taking into account the wide dispersion of HFE mutation in our population and also its unknown role in heterozygotes, we analyzed the impact of H63D HFE carriage in the developmental age, with respect to gender, on the iron status and hemoglobin concentration of carriers in comparison to those of wild-type HFE gene (12.7 ± 3.07 years, 42 boys and 41 girls). H63D carriers presented higher blood iron, transferrin saturation, and ferritin concentration than wild-type probands (p < 0.05.) Interestingly, male H63D carriers showed higher hemoglobin concentration than the unburdened children. Moreover, in the H63D carrier group, a positive correlation between iron and hemoglobin was noted. In conclusion, this study demonstrates that changes in iron metabolism occur at a young age in HFE heterozygotes.

  8. Impact of gene polymorphisms of gonadotropins and their receptors on human reproductive success.

    PubMed

    Casarini, Livio; Santi, Daniele; Marino, Marco

    2015-12-01

    Gonadotropins and their receptors' genes carry several single-nucleotide polymorphisms resulting in endocrine genotypes modulating reproductive parameters, diseases, and lifespan leading to important implications for reproductive success and potential relevance during human evolution. Here we illustrate common genotypes of the gonadotropins and gonadotropin receptors' genes and their clinical implications in phenotypes relevant for reproduction such as ovarian cycle length, age of menopause, testosterone levels, polycystic ovary syndrome, and cancer. We then discuss their possible role in human reproduction and adaptation to the environment. Gonadotropins and their receptors' variants are differently distributed among human populations. Some hints suggest that they may be the result of natural selection that occurred in ancient times, increasing the individual chance of successful mating, pregnancy, and effective post-natal parental cares. The gender-related differences in the regulation of the reproductive endocrine systems imply that many of these genotypes may lead to sex-dependent effects, increasing the chance of mating and reproductive success in one sex at the expenses of the other sex. Also, we suggest that sexual conflicts within the FSH and LH-choriogonadotropin receptor genes contributed to maintain genotypes linked to subfertility among humans. Because the distribution of polymorphic markers results in a defined geographical pattern due to human migrations rather than natural selection, these polymorphisms may have had only a weak impact on reproductive success. On the contrary, such genotypes could acquire relevant consequences in the modern, developed societies in which parenthood attempts often occur at a later age, during a short, suboptimal reproductive window, making clinical fertility treatments necessary.

  9. Impact of dairy manure pre-application treatment on manure composition, soil dynamics of antibiotic resistance genes, and abundance of antibiotic-resistance genes on vegetables at harvest.

    PubMed

    Tien, Yuan-Ching; Li, Bing; Zhang, Tong; Scott, Andrew; Murray, Roger; Sabourin, Lyne; Marti, Romain; Topp, Edward

    2017-03-01

    Manuring ground used for crop production is an important agricultural practice. Should antibiotic-resistant enteric bacteria carried in the manure be transferred to crops that are consumed raw, their consumption by humans or animals will represent a route of exposure to antibiotic resistance genes. Treatment of manures prior to land application is a potential management option to reduce the abundance of antibiotic resistance genes entrained with manure application. In this study, dairy manure that was untreated, anaerobically digested, mechanically dewatered or composted was applied to field plots that were then cropped to lettuce, carrots and radishes. The impact of treatment on manure composition, persistence of antibiotic resistance gene targets in soil following application, and distribution of antibiotic resistance genes and bacteria on vegetables at harvest was determined. Composted manure had the lowest abundance of antibiotic resistance gene targets compared to the other manures. There was no significant difference in the persistence characteristics of antibiotic resistance genes following land application of the various manures. Compared to unmanured soil, antibiotic resistance genes were detected more frequently in soil receiving raw or digested manure, whereas they were not in soil receiving composted manure. The present study suggests that vegetables grown in ground receiving raw or digested manure are at risk of contamination with manure-borne antibiotic resistant bacteria, whereas vegetables grown in ground receiving composted manure are less so.

  10. Metagenomic profiling of historic Colorado Front Range flood impact on distribution of riverine antibiotic resistance genes

    NASA Astrophysics Data System (ADS)

    Garner, Emily; Wallace, Joshua S.; Argoty, Gustavo Arango; Wilkinson, Caitlin; Fahrenfeld, Nicole; Heath, Lenwood S.; Zhang, Liqing; Arabi, Mazdak; Aga, Diana S.; Pruden, Amy

    2016-12-01

    Record-breaking floods in September 2013 caused massive damage to homes and infrastructure across the Colorado Front Range and heavily impacted the Cache La Poudre River watershed. Given the unique nature of this watershed as a test-bed for tracking environmental pathways of antibiotic resistance gene (ARG) dissemination, we sought to determine the impact of extreme flooding on ARG reservoirs in river water and sediment. We utilized high-throughput DNA sequencing to obtain metagenomic profiles of ARGs before and after flooding, and investigated 23 antibiotics and 14 metals as putative selective agents during post-flood recovery. With 277 ARG subtypes identified across samples, total bulk water ARGs decreased following the flood but recovered to near pre-flood abundances by ten months post-flood at both a pristine site and at a site historically heavily influenced by wastewater treatment plants and animal feeding operations. Network analysis of de novo assembled sequencing reads into 52,556 scaffolds identified ARGs likely located on mobile genetic elements, with up to 11 ARGs per plasmid-associated scaffold. Bulk water bacterial phylogeny correlated with ARG profiles while sediment phylogeny varied along the river’s anthropogenic gradient. This rare flood afforded the opportunity to gain deeper insight into factors influencing the spread of ARGs in watersheds.

  11. Metagenomic profiling of historic Colorado Front Range flood impact on distribution of riverine antibiotic resistance genes

    PubMed Central

    Garner, Emily; Wallace, Joshua S.; Argoty, Gustavo Arango; Wilkinson, Caitlin; Fahrenfeld, Nicole; Heath, Lenwood S.; Zhang, Liqing; Arabi, Mazdak; Aga, Diana S.; Pruden, Amy

    2016-01-01

    Record-breaking floods in September 2013 caused massive damage to homes and infrastructure across the Colorado Front Range and heavily impacted the Cache La Poudre River watershed. Given the unique nature of this watershed as a test-bed for tracking environmental pathways of antibiotic resistance gene (ARG) dissemination, we sought to determine the impact of extreme flooding on ARG reservoirs in river water and sediment. We utilized high-throughput DNA sequencing to obtain metagenomic profiles of ARGs before and after flooding, and investigated 23 antibiotics and 14 metals as putative selective agents during post-flood recovery. With 277 ARG subtypes identified across samples, total bulk water ARGs decreased following the flood but recovered to near pre-flood abundances by ten months post-flood at both a pristine site and at a site historically heavily influenced by wastewater treatment plants and animal feeding operations. Network analysis of de novo assembled sequencing reads into 52,556 scaffolds identified ARGs likely located on mobile genetic elements, with up to 11 ARGs per plasmid-associated scaffold. Bulk water bacterial phylogeny correlated with ARG profiles while sediment phylogeny varied along the river’s anthropogenic gradient. This rare flood afforded the opportunity to gain deeper insight into factors influencing the spread of ARGs in watersheds. PMID:27917931

  12. Linking gene regulation and the exo-metabolome: A comparative transcriptomics approach to identify genes that impact on the production of volatile aroma compounds in yeast

    PubMed Central

    Rossouw, Debra; Næs, Tormod; Bauer, Florian F

    2008-01-01

    Background 'Omics' tools provide novel opportunities for system-wide analysis of complex cellular functions. Secondary metabolism is an example of a complex network of biochemical pathways, which, although well mapped from a biochemical point of view, is not well understood with regards to its physiological roles and genetic and biochemical regulation. Many of the metabolites produced by this network such as higher alcohols and esters are significant aroma impact compounds in fermentation products, and different yeast strains are known to produce highly divergent aroma profiles. Here, we investigated whether we can predict the impact of specific genes of known or unknown function on this metabolic network by combining whole transcriptome and partial exo-metabolome analysis. Results For this purpose, the gene expression levels of five different industrial wine yeast strains that produce divergent aroma profiles were established at three different time points of alcoholic fermentation in synthetic wine must. A matrix of gene expression data was generated and integrated with the concentrations of volatile aroma compounds measured at the same time points. This relatively unbiased approach to the study of volatile aroma compounds enabled us to identify candidate genes for aroma profile modification. Five of these genes, namely YMR210W, BAT1, AAD10, AAD14 and ACS1 were selected for overexpression in commercial wine yeast, VIN13. Analysis of the data show a statistically significant correlation between the changes in the exo-metabome of the overexpressing strains and the changes that were predicted based on the unbiased alignment of transcriptomic and exo-metabolomic data. Conclusion The data suggest that a comparative transcriptomics and metabolomics approach can be used to identify the metabolic impacts of the expression of individual genes in complex systems, and the amenability of transcriptomic data to direct applications of biotechnological relevance. PMID:18990252

  13. Impact of obesity and nitric oxide synthase gene G894T polymorphism on essential hypertension.

    PubMed

    Wrzosek, M; Sokal, M; Sawicka, A; Wlodarczyk, M; Glowala, M; Wrzosek, M; Kosior, M; Talalaj, M; Biecek, P; Nowicka, G

    2015-10-01

    Hypertension is a multifactorial disease caused by environmental, metabolic and genetic factors, but little is currently known on the complex interplay between these factors and blood pressure. The aim of the present study was to assess the potential impact of obesity, and angiotensin-converting enzyme (ACE) I/D polymorphism and endothelial nitric oxide synthase gene (NOS3) 4a/4b, G894T and -T786C variants on the essential hypertension. The study group consisted of 1,027 Caucasian adults of Polish nationality (45.5 ± 13.6 years old), of which 401 met the criteria for hypertension. Body weight, height and blood pressure were measured and data on self-reported smoking status were collected. Fasting blood glucose, total cholesterol, LDL-cholesterol, HDL-cholesterol, triglycerides were determined by standard procedures. The ACE I/D polymorphism and three polymorphisms in NOS3 gene (4a/4b, G894T, -T786C) were detected by the PCR method. Multivariable logistic regression demonstrated that age above 45 years, diabetes, dyslipidemia, smoking and male sex are important risk factors for hypertension and no significant influence of variants in ACE and NOS3 genes on this risk was recognized. Obese subjects had a 3.27-times higher risk (OR = 3.27, 95% CI: 2.37 - 4.52) of hypertension than non-obese, and in obese the NOS3 894T allele was associated with 1.37 fold higher risk of hypertension (P = 0.031). The distribution of NOS3 G894T genotypes supported the co-dominant (OR = 1.35, P = 0.034, Pfit = 0.435) or recessive (OR = 2.00, P = 0.046, Pfit = 0.286), but not dominant model of inheritance (P = 0.100). The study indicates that in obese NOS3 G894T polymorphism may enhance hypertension risk. However, in the presence of such strong risk factors as age, diabetes and smoking, the impact of this genetic variant seems to be attenuated. Further studies are needed to reveal the usefulness of G894T polymorphism in hypertension risk assessment in obese.

  14. The impact of gene expression variation on the robustness and evolvability of a developmental gene regulatory network.

    PubMed

    Garfield, David A; Runcie, Daniel E; Babbitt, Courtney C; Haygood, Ralph; Nielsen, William J; Wray, Gregory A

    2013-10-01

    Regulatory interactions buffer development against genetic and environmental perturbations, but adaptation requires phenotypes to change. We investigated the relationship between robustness and evolvability within the gene regulatory network underlying development of the larval skeleton in the sea urchin Strongylocentrotus purpuratus. We find extensive variation in gene expression in this network throughout development in a natural population, some of which has a heritable genetic basis. Switch-like regulatory interactions predominate during early development, buffer expression variation, and may promote the accumulation of cryptic genetic variation affecting early stages. Regulatory interactions during later development are typically more sensitive (linear), allowing variation in expression to affect downstream target genes. Variation in skeletal morphology is associated primarily with expression variation of a few, primarily structural, genes at terminal positions within the network. These results indicate that the position and properties of gene interactions within a network can have important evolutionary consequences independent of their immediate regulatory role.

  15. The Impact of Gene Expression Variation on the Robustness and Evolvability of a Developmental Gene Regulatory Network

    PubMed Central

    Garfield, David A.; Runcie, Daniel E.; Babbitt, Courtney C.; Haygood, Ralph; Nielsen, William J.; Wray, Gregory A.

    2013-01-01

    Regulatory interactions buffer development against genetic and environmental perturbations, but adaptation requires phenotypes to change. We investigated the relationship between robustness and evolvability within the gene regulatory network underlying development of the larval skeleton in the sea urchin Strongylocentrotus purpuratus. We find extensive variation in gene expression in this network throughout development in a natural population, some of which has a heritable genetic basis. Switch-like regulatory interactions predominate during early development, buffer expression variation, and may promote the accumulation of cryptic genetic variation affecting early stages. Regulatory interactions during later development are typically more sensitive (linear), allowing variation in expression to affect downstream target genes. Variation in skeletal morphology is associated primarily with expression variation of a few, primarily structural, genes at terminal positions within the network. These results indicate that the position and properties of gene interactions within a network can have important evolutionary consequences independent of their immediate regulatory role. PMID:24204211

  16. Antimicrobial-resistant bacterial populations and antimicrobial resistance genes obtained from environments impacted by livestock and municipal waste

    USDA-ARS?s Scientific Manuscript database

    This study compared the populations of antimicrobial-resistant bacteria and the repertoire of antimicrobial resistance genes in four environments: effluent of three municipal waste water treatment facilities, three cattle feedlot runoff catchment ponds, three swine waste lagoons, and two "low impact...

  17. Impact of SCP-2/SCP-x gene ablation and dietary cholesterol on hepatic lipid accumulation

    PubMed Central

    Klipsic, Devon; Landrock, Danilo; Martin, Gregory G.; McIntosh, Avery L.; Landrock, Kerstin K.; Mackie, John T.; Schroeder, Friedhelm

    2015-01-01

    While a high-cholesterol diet induces hepatic steatosis, the role of intracellular sterol carrier protein-2/sterol carrier protein-x (SCP-2/SCP-x) proteins is unknown. We hypothesized that ablating SCP-2/SCP-x [double knockout (DKO)] would impact hepatic lipids (cholesterol and cholesteryl ester), especially in high-cholesterol-fed mice. DKO did not alter food consumption, and body weight (BW) gain decreased especially in females, concomitant with hepatic steatosis in females and less so in males. DKO-induced steatosis in control-fed wild-type (WT) mice was associated with 1) loss of SCP-2; 2) upregulation of liver fatty acid binding protein (L-FABP); 3) increased mRNA and/or protein levels of sterol regulatory element binding proteins (SREBP1 and SREBP2) as well as increased expression of target genes of cholesterol synthesis (Hmgcs1 and Hmgcr) and fatty acid synthesis (Acc1 and Fas); and 4) cholesteryl ester accumulation was also associated with increased acyl-CoA cholesterol acyltransferase-2 (ACAT2) in males. DKO exacerbated the high-cholesterol diet-induced hepatic cholesterol and glyceride accumulation, without further increasing SREBP1, SREBP2, or target genes. This exacerbation was associated both with loss of SCP-2 and concomitant downregulation of Ceh/Hsl, apolipoprotein B (ApoB), MTP, and/or L-FABP protein expression. DKO diminished the ability to secrete excess cholesterol into bile and oxidize cholesterol to bile acid for biliary excretion, especially in females. This suggested that SCP-2/SCP-x affects cholesterol transport to particular intracellular compartments, with ablation resulting in less to the endoplasmic reticulum for SREBP regulation, making more available for cholesteryl ester synthesis, for cholesteryl-ester storage in lipid droplets, and for bile salt synthesis and/or secretion. These alterations are significant findings, since they affect key processes in regulation of sterol metabolism. PMID:26113298

  18. Impact of the GeneXpert MTB/RIF Technology on Tuberculosis Control.

    PubMed

    Stevens, Wendy Susan; Scott, Lesley; Noble, Lara; Gous, Natasha; Dheda, Keertan

    2017-01-01

    Molecular technology revolutionized the diagnosis of tuberculosis (TB) with a paradigm shift to faster, more sensitive, clinically relevant patient care. The most recent molecular leader is the GeneXpert MTB/RIF assay (Xpert) (Cepheid, Sunnyvale, CA), which was endorsed by the World Health Organization with unprecedented speed in December 2010 as the initial diagnostic for detection of HIV-associated TB and for where high rates of drug resistance are suspected. South Africa elected to take an aggressive smear replacement approach to facilitate earlier diagnosis and treatment through the decision to implement the Xpert assay nationally in March 2011, against the backdrop of approximately 6.3 million HIV-infected individuals, one of highest global TB and HIV coinfection rates, no available implementation models, uncertainties around field performance and program costs, and lack of guidance on how to operationalize the assay into existing complex clinical algorithms. South Africa's national implementation was conducted as a phased, forecasted, and managed approach (March 2011 to September 2013), through political will and both treasury-funded and donor-funded support. Today there are 314 GeneXperts across 207 microscopy centers; over 8 million assays have been conducted, and South Africa accounts for over half the global test cartridge usage. As with any implementation of new technology, challenges were encountered, both predicted and unexpected. This chapter discusses the challenges and consequences of such large-scale implementation efforts, the opportunities for new innovations, and the need to strengthen health systems, as well as the impact of the Xpert assay on rifampin-sensitive and multidrug-resistant TB patient care that translated into global TB control as we move toward the sustainable development goals.

  19. Impact of gene molecular evolution on phylogenetic reconstruction: a case study in the rosids (Superorder Rosanae, Angiosperms).

    PubMed

    Hilu, Khidir W; Black, Chelsea M; Oza, Dipan

    2014-01-01

    Rate of substitution of genomic regions is among the most debated intrinsic features that impact phylogenetic informativeness. However, this variable is also coupled with rates of nonsynonymous substitutions that underscore the nature and degree of selection on the selected genes. To empirically address these variables, we constructed four completely overlapping data sets of plastid matK, atpB, rbcL, and mitochondrial matR genes and used the rosid lineage (angiosperms) as a working platform. The genes differ in combinations of overall rates of nucleotide and amino acid substitutions. Tree robustness, homoplasy, accuracy in contrast to a reference tree, and phylogenetic informativeness are evaluated. The rapidly evolving/unconstrained matK faired best, whereas remaining genes varied in degrees of contribution to rosid phylogenetics across the lineage's 108 million years evolutionary history. Phylogenetic accuracy was low with the slowly evolving/unconstrained matR despite least amount of homoplasy. Third codon positions contributed the highest amount of parsimony informative sites, resolution and informativeness, but magnitude varied with gene mode of evolution. These findings are in clear contrast with the views that rapidly evolving regions and the 3rd codon position have inevitable negative impact on phylogenetic reconstruction at deep historic level due to accumulation of multiple hits and subsequent elevation in homoplasy and saturation. Relaxed evolutionary constraint in rapidly evolving genes distributes substitutions across codon positions, an evolutionary mode expected to reduce the frequency of multiple hits. These findings should be tested at deeper evolutionary histories.

  20. Impact of Gene Molecular Evolution on Phylogenetic Reconstruction: A Case Study in the Rosids (Superorder Rosanae, Angiosperms)

    PubMed Central

    Hilu, Khidir W.; Black, Chelsea M.; Oza, Dipan

    2014-01-01

    Rate of substitution of genomic regions is among the most debated intrinsic features that impact phylogenetic informativeness. However, this variable is also coupled with rates of nonsynonymous substitutions that underscore the nature and degree of selection on the selected genes. To empirically address these variables, we constructed four completely overlapping data sets of plastid matK, atpB, rbcL, and mitochondrial matR genes and used the rosid lineage (angiosperms) as a working platform. The genes differ in combinations of overall rates of nucleotide and amino acid substitutions. Tree robustness, homoplasy, accuracy in contrast to a reference tree, and phylogenetic informativeness are evaluated. The rapidly evolving/unconstrained matK faired best, whereas remaining genes varied in degrees of contribution to rosid phylogenetics across the lineage's 108 million years evolutionary history. Phylogenetic accuracy was low with the slowly evolving/unconstrained matR despite least amount of homoplasy. Third codon positions contributed the highest amount of parsimony informative sites, resolution and informativeness, but magnitude varied with gene mode of evolution. These findings are in clear contrast with the views that rapidly evolving regions and the 3rd codon position have inevitable negative impact on phylogenetic reconstruction at deep historic level due to accumulation of multiple hits and subsequent elevation in homoplasy and saturation. Relaxed evolutionary constraint in rapidly evolving genes distributes substitutions across codon positions, an evolutionary mode expected to reduce the frequency of multiple hits. These findings should be tested at deeper evolutionary histories. PMID:24932884

  1. Palliating the impact of fixation of a major gene on the genetic variation of artificially selected polygenes.

    PubMed

    Sánchez, Leopoldo; Caballero, Armando; Santiago, Enrique

    2006-10-01

    Selective sweeps of variation caused by fixation of major genes may have a dramatic impact on the genetic gain from background polygenic variation, particularly in the genome regions closely linked to the major gene. The response to selection can be restrained because of the reduced selection intensity and the reduced effective population size caused by the increase in frequency of the major gene. In the context of a selected population where fixation of a known major gene is desired, the question arises as to which is the optimal path of increase in frequency of the gene so that the selective sweep of variation resulting from its fixation is minimized. Using basic theoretical arguments we propose a frequency path that maximizes simultaneously the effective population size applicable to the selected background and the selection intensity on the polygenic variation by minimizing the average squared selection intensity on the major gene over generations up to a given fixation time. We also propose the use of mating between carriers and non-carriers of the major gene, in order to promote the effective recombination between the major gene and its linked polygenic background. Using a locus-based computer simulation assuming different degrees of linkage, we show that the path proposed is more effective than a similar path recently published, and that the combination of the selection and mating methods provides an efficient way to palliate the negative effects of a selective sweep.

  2. The Impact of Selection, Gene Conversion, and Biased Sampling on the Assessment of Microbial Demography.

    PubMed

    Lapierre, Marguerite; Blin, Camille; Lambert, Amaury; Achaz, Guillaume; Rocha, Eduardo P C

    2016-07-01

    Recent studies have linked demographic changes and epidemiological patterns in bacterial populations using coalescent-based approaches. We identified 26 studies using skyline plots and found that 21 inferred overall population expansion. This surprising result led us to analyze the impact of natural selection, recombination (gene conversion), and sampling biases on demographic inference using skyline plots and site frequency spectra (SFS). Forward simulations based on biologically relevant parameters from Escherichia coli populations showed that theoretical arguments on the detrimental impact of recombination and especially natural selection on the reconstructed genealogies cannot be ignored in practice. In fact, both processes systematically lead to spurious interpretations of population expansion in skyline plots (and in SFS for selection). Weak purifying selection, and especially positive selection, had important effects on skyline plots, showing patterns akin to those of population expansions. State-of-the-art techniques to remove recombination further amplified these biases. We simulated three common sampling biases in microbiological research: uniform, clustered, and mixed sampling. Alone, or together with recombination and selection, they further mislead demographic inferences producing almost any possible skyline shape or SFS. Interestingly, sampling sub-populations also affected skyline plots and SFS, because the coalescent rates of populations and their sub-populations had different distributions. This study suggests that extreme caution is needed to infer demographic changes solely based on reconstructed genealogies. We suggest that the development of novel sampling strategies and the joint analyzes of diverse population genetic methods are strictly necessary to estimate demographic changes in populations where selection, recombination, and biased sampling are present. © The Author 2016. Published by Oxford University Press on behalf of the Society for

  3. Elevated atmospheric CO2 impacts abundance and diversity of nitrogen cycling functional genes in soil.

    PubMed

    Kelly, John J; Peterson, Emily; Winkelman, Jonathan; Walter, Teagan J; Rier, Steven T; Tuchman, Nancy C

    2013-02-01

    The concentration of CO(2) in the Earth's atmosphere has increased over the last century. Although this increase is unlikely to have direct effects on soil microbial communities, increased atmospheric CO(2) may impact soil ecosystems indirectly through plant responses. This study tested the hypothesis that exposure of plants to elevated CO(2) would impact soil microorganisms responsible for key nitrogen cycling processes, specifically denitrification and nitrification. We grew trembling aspen (Populus tremuloides) trees in outdoor chambers under ambient (360 ppm) or elevated (720 ppm) levels of CO(2) for 5 years and analyzed the microbial communities in the soils below the trees using quantitative polymerase chain reaction and clone library sequencing targeting the nitrite reductase (nirK) and ammonia monooxygenase (amoA) genes. We observed a more than twofold increase in copy numbers of nirK and a decrease in nirK diversity with CO(2) enrichment, with an increased predominance of Bradyrhizobia-like nirK sequences. We suggest that this dramatic increase in nirK-containing bacteria may have contributed to the significant loss of soil N in the CO(2)-treated chambers. Elevated CO(2) also resulted in a significant decrease in copy numbers of bacterial amoA, but no change in archaeal amoA copy numbers. The decrease in abundance of bacterial amoA was likely a result of the loss of soil N in the CO(2)-treated chambers, while the lack of response for archaeal amoA supports the hypothesis that physiological differences in these two groups of ammonia oxidizers may enable them to occupy distinct ecological niches and respond differently to environmental change.

  4. The Impact of Selection, Gene Conversion, and Biased Sampling on the Assessment of Microbial Demography

    PubMed Central

    Lapierre, Marguerite; Blin, Camille; Lambert, Amaury; Achaz, Guillaume; Rocha, Eduardo P. C.

    2016-01-01

    Recent studies have linked demographic changes and epidemiological patterns in bacterial populations using coalescent-based approaches. We identified 26 studies using skyline plots and found that 21 inferred overall population expansion. This surprising result led us to analyze the impact of natural selection, recombination (gene conversion), and sampling biases on demographic inference using skyline plots and site frequency spectra (SFS). Forward simulations based on biologically relevant parameters from Escherichia coli populations showed that theoretical arguments on the detrimental impact of recombination and especially natural selection on the reconstructed genealogies cannot be ignored in practice. In fact, both processes systematically lead to spurious interpretations of population expansion in skyline plots (and in SFS for selection). Weak purifying selection, and especially positive selection, had important effects on skyline plots, showing patterns akin to those of population expansions. State-of-the-art techniques to remove recombination further amplified these biases. We simulated three common sampling biases in microbiological research: uniform, clustered, and mixed sampling. Alone, or together with recombination and selection, they further mislead demographic inferences producing almost any possible skyline shape or SFS. Interestingly, sampling sub-populations also affected skyline plots and SFS, because the coalescent rates of populations and their sub-populations had different distributions. This study suggests that extreme caution is needed to infer demographic changes solely based on reconstructed genealogies. We suggest that the development of novel sampling strategies and the joint analyzes of diverse population genetic methods are strictly necessary to estimate demographic changes in populations where selection, recombination, and biased sampling are present. PMID:26931140

  5. The combined impact of metabolic gene polymorphisms on elite endurance athlete status and related phenotypes.

    PubMed

    Ahmetov, Ildus I; Williams, Alun G; Popov, Daniil V; Lyubaeva, Ekaterina V; Hakimullina, Albina M; Fedotovskaya, Olga N; Mozhayskaya, Irina A; Vinogradova, Olga L; Astratenkova, Irina V; Montgomery, Hugh E; Rogozkin, Viktor A

    2009-12-01

    Endurance performance is a complex phenotype subject to the influence of both environmental and genetic factors. Although the last decade has seen a variety of specific genetic factors proposed, many in metabolic pathways, each is likely to make a limited contribution to an 'elite' phenotype: it seems more likely that such status depends on the simultaneous presence of multiple such variants. The aim of the study was to investigate individually and in combination the association of common metabolic gene polymorphisms with endurance athlete status, the proportion of slow-twitch muscle fibers and maximal oxygen consumption. A total of 1,423 Russian athletes and 1,132 controls were genotyped for 15 gene polymorphisms, of which most were previously reported to be associated with athlete status or related intermediate phenotypes. Muscle fiber composition of m. vastus lateralis in 45 healthy men was determined by immunohistochemistry. Maximal oxygen consumption of 50 male rowers of national competitive standard was determined during an incremental test to exhaustion on a rowing ergometer. Ten 'endurance alleles' (NFATC4 Gly160, PPARA rs4253778 G, PPARD rs2016520 C, PPARGC1A Gly482, PPARGC1B 203Pro, PPP3R1 promoter 5I, TFAM 12Thr, UCP2 55Val, UCP3 rs1800849 T and VEGFA rs2010963 C) were first identified showing discrete associations with elite endurance athlete status. Next, to assess the combined impact of all 10 gene polymorphisms, all athletes were classified according to the number of 'endurance' alleles they possessed. The proportion of subjects with a high (≥9) number of 'endurance' alleles was greater in the best endurance athletes compared with controls (85.7 vs. 37.8%, P = 7.6 × 10(-6)). The number of 'endurance' alleles was shown to be positively correlated (r = 0.50; P = 4.0 × 10(-4)) with the proportion of fatigue-resistant slow-twitch fibers, and with maximal oxygen consumption (r = 0.46; P = 7.0 × 10(-4)). These data suggest that the likelihood of

  6. Evidence of current impact of climate change on life: a walk from genes to the biosphere.

    PubMed

    Peñuelas, Josep; Sardans, Jordi; Estiarte, Marc; Ogaya, Romà; Carnicer, Jofre; Coll, Marta; Barbeta, Adria; Rivas-Ubach, Albert; Llusià, Joan; Garbulsky, Martin; Filella, Iolanda; Jump, Alistair S

    2013-08-01

    We review the evidence of how organisms and populations are currently responding to climate change through phenotypic plasticity, genotypic evolution, changes in distribution and, in some cases, local extinction. Organisms alter their gene expression and metabolism to increase the concentrations of several antistress compounds and to change their physiology, phenology, growth and reproduction in response to climate change. Rapid adaptation and microevolution occur at the population level. Together with these phenotypic and genotypic adaptations, the movement of organisms and the turnover of populations can lead to migration toward habitats with better conditions unless hindered by barriers. Both migration and local extinction of populations have occurred. However, many unknowns for all these processes remain. The roles of phenotypic plasticity and genotypic evolution and their possible trade-offs and links with population structure warrant further research. The application of omic techniques to ecological studies will greatly favor this research. It remains poorly understood how climate change will result in asymmetrical responses of species and how it will interact with other increasing global impacts, such as N eutrophication, changes in environmental N : P ratios and species invasion, among many others. The biogeochemical and biophysical feedbacks on climate of all these changes in vegetation are also poorly understood. We here review the evidence of responses to climate change and discuss the perspectives for increasing our knowledge of the interactions between climate change and life. © 2013 John Wiley & Sons Ltd.

  7. Impact of mild temperature hardening on thermotolerance, fecundity, and Hsp gene expression in Liriomyza huidobrensis.

    PubMed

    Huang, Li-Hua; Chen, Bing; Kang, Le

    2007-12-01

    The pea leafminer, Liriomyza huidobrensis, is one of the most important economic insect pests around the world. Its population fluctuates greatly with seasonal change in China, and temperature was thought to be one of the important reasons. In attempt to further explore the impact of disadvantageous temperature on L. huidobrensis, 1-day-old adults were shocked at various temperatures (10, 25, 32, and 35 degrees C, respectively) for 4h, and the effects on thermotolerance, feeding, and fecundity were studied. Meanwhile the expression of five heat shock genes (hsp90, 70, 60, 40, and 20) was examined by real-time quantitative PCR. Our results showed that both 32 and 35 degrees C hardenings remarkably increased adult heat resistance, whereas cold tolerance was not improved accordingly. No cross resistance in response to cold and heat stresses was observed. Both adult feeding and fecundity were dramatically reduced, but no effect was observed on egg hatching, larval survival, pupal eclosion, or sex ratio. The results indicate that the deleterious effect on fecundity is the result of direct cessation of oviposition during the period of stress. Simultaneously, the mRNA levels of hsp70 and hsp20 significantly increased upon thermal hardening. Taken together, our results suggest that mild heat hardening improves thermotolerance of L. huidobrensis at the cost of impairment on fecundity, and the induced expression of hsp70 and hsp20 may play an important role in balancing the functional tradeoff.

  8. [Histamine H₁ receptor gene as an allergic diseases-sensitive gene and its impact on therapeutics for allergic diseases].

    PubMed

    Mizuguchi, Hiroyuki; Kitamura, Yoshiaki; Kondo, Yuto; Kuroda, Wakana; Yoshida, Haruka; Miyamoto, Yuko; Hattori, Masashi; Takeda, Noriaki; Fukui, Hiroyuki

    2011-02-01

    Therapeutics targeting disease-sensitive genes are required for the therapy of multifactorial diseases. There is no clinical report on therapeutics for allergic disease-sensitive genes. We are focusing on the histamine H₁ receptor (H1R) as a sensitive gene. H1R mediates allergy histamine signals. H1R is a rate-limiting molecule of the H1R signal because the signal is increased with elevated receptor expression level. We discovered that the stimulation of H1R induced H1R gene expression through PKCδ activation, resulting in receptor upregulation. The mechanism of H1R gene expression was revealed to play a key role in the receptor expression level in studies using cultured HeLa cells and allergic rhinitis model rats. Preseasonal prophylactic treatment with antihistamines is recommended for the therapy of pollinosis. However, the mechanism of the therapy remains to be elucidated. We demonstrated that repeated pretreatment treatment with antihistamines in the allergic rhinitis model rats resulted not only in improvement of symptoms but also in suppressed elevation of H1R mRNA levels in the nasal mucosa. A clinical trial was then initiated. When symptoms and H1R mRNA levels in the nasal mucosa of pollinosis patients with or without preseasonal prophylactic treatment with antihistamines were examined, both symptoms and high levels of H1R mRNA were significantly improved in treated compared with untreated patients. These results strongly suggest that H1R is an allergic disease-sensitive gene.

  9. Global analysis of somatic structural genomic alterations and their impact on gene expression in diverse human cancers

    PubMed Central

    Alaei-Mahabadi, Babak; Karlsson, Joakim W.; Nilsson, Jonas A.; Larsson, Erik

    2016-01-01

    Tumor genomes are mosaics of somatic structural variants (SVs) that may contribute to the activation of oncogenes or inactivation of tumor suppressors, for example, by altering gene copy number amplitude. However, there are multiple other ways in which SVs can modulate transcription, but the general impact of such events on tumor transcriptional output has not been systematically determined. Here we use whole-genome sequencing data to map SVs across 600 tumors and 18 cancers, and investigate the relationship between SVs, copy number alterations (CNAs), and mRNA expression. We find that 34% of CNA breakpoints can be clarified structurally and that most amplifications are due to tandem duplications. We observe frequent swapping of strong and weak promoters in the context of gene fusions, and find that this has a measurable global impact on mRNA levels. Interestingly, several long noncoding RNAs were strongly activated by this mechanism. Additionally, SVs were confirmed in telomere reverse transcriptase (TERT) upstream regions in several cancers, associated with elevated TERT mRNA levels. We also highlight high-confidence gene fusions supported by both genomic and transcriptomic evidence, including a previously undescribed paired box 8 (PAX8)–nuclear factor, erythroid 2 like 2 (NFE2L2) fusion in thyroid carcinoma. In summary, we combine SV, CNA, and expression data to provide insights into the structural basis of CNAs as well as the impact of SVs on gene expression in tumors. PMID:27856756

  10. The impact of human and mouse differences in NOS2 gene expression on the brain's redox and immune environment.

    PubMed

    Hoos, Michael D; Vitek, Michael P; Ridnour, Lisa A; Wilson, Joan; Jansen, Marilyn; Everhart, Angela; Wink, David A; Colton, Carol A

    2014-11-17

    Mouse models are used in the study of human disease. Despite well-known homologies, the difference in immune response between mice and humans impacts the application of data derived from mice to human disease outcomes. Nitric oxide synthase-2 (NOS2) is a key gene that displays species-specific outcomes via altered regulation of the gene promoter and via post-transcriptional mechanisms in humans that are not found in mice. The resulting levels of NO produced by activation of human NOS2 are different from the levels of NO produced by mouse Nos2. Since both tissue redox environment and immune responsiveness are regulated by the level of NO and its interactions, we investigated the significance of mouse and human differences on brain oxidative stress and on immune activation in HuNOS2tg/mNos2-/- mice that express the entire human NOS2 gene and that lack a functional mNos2 compared to wild type (WT) mice that express normal mNos2. Similarly to human, brain tissue from HuNOS2tg/mNos2-/- mice showed the presence of a NOS2 gene 3'UTR binding site. We also identified miRNA-939, the binding partner for this site, in mouse brain lysates and further demonstrated reduced levels of nitric oxide (NO) typical of the human immune response on injection with lipopolysaccharide (LPS). HuNOS2tg/mNos2-/- brain samples were probed for characteristic differences in redox and immune gene profiles compared to WT mice using gene arrays. Selected genes were also compared against mNos2-/- brain lysates. Reconstitution of the human NOS2 gene significantly altered genes that encode multiple anti-oxidant proteins, oxidases, DNA repair, mitochondrial proteins and redox regulated immune proteins. Expression levels of typical pro-inflammatory, anti-inflammatory and chemokine genes were not significantly different with the exception of increased TNFα and Ccr1 mRNA expression in the HuNOS2tg/mNos2-/- mice compared to WT or mNos2-/- mice. NO is a principle factor in establishing the tissue redox

  11. The impact of microRNAs on transcriptional heterogeneity and gene co-expression across single embryonic stem cells

    PubMed Central

    Gambardella, Gennaro; Carissimo, Annamaria; Chen, Amy; Cutillo, Luisa; Nowakowski, Tomasz J.; di Bernardo, Diego; Blelloch, Robert

    2017-01-01

    MicroRNAs act posttranscriptionally to suppress multiple target genes within a cell population. To what extent this multi-target suppression occurs in individual cells and how it impacts transcriptional heterogeneity and gene co-expression remains unknown. Here we used single-cell sequencing combined with introduction of individual microRNAs. miR-294 and let-7c were introduced into otherwise microRNA-deficient Dgcr8 knockout mouse embryonic stem cells. Both microRNAs induce suppression and correlated expression of their respective gene targets. The two microRNAs had opposing effects on transcriptional heterogeneity within the cell population, with let-7c increasing and miR-294 decreasing the heterogeneity between cells. Furthermore, let-7c promotes, whereas miR-294 suppresses, the phasing of cell cycle genes. These results show at the individual cell level how a microRNA simultaneously has impacts on its many targets and how that in turn can influence a population of cells. The findings have important implications in the understanding of how microRNAs influence the co-expression of genes and pathways, and thus ultimately cell fate. PMID:28102192

  12. Gene patents, patenting life and the impact of court rulings on US stem cell patents and research.

    PubMed

    Matthews, Kirstin R W; Cuchiara, Maude L

    2014-03-01

    In June 2013, the US Supreme Court ruled that naturally occurring genes were unpatentable in the case Association for Molecular Pathology v. Myriad Genetics. Up until this decision, Myriad Genetics was the only company in the USA that could legally conduct diagnostic testing for BRCA1 and 2, genes that are linked to familial breast and ovarian cancer. The court case and rulings garnered discussion in public about patenting biological materials. This paper will describe the progression of the Myriad Genetics case, similar US rulings and biological intellectual property policies. In addition, it will discuss the impact of the case on biological patents - specifically those for human embryonic stem cells.

  13. Does human activity impact the natural antibiotic resistance background? Abundance of antibiotic resistance genes in 21 Swiss lakes.

    PubMed

    Czekalski, Nadine; Sigdel, Radhika; Birtel, Julia; Matthews, Blake; Bürgmann, Helmut

    2015-08-01

    Antibiotic resistance genes (ARGs) are emerging environmental contaminants, known to be continuously discharged into the aquatic environment via human and animal waste. Freshwater aquatic environments represent potential reservoirs for ARG and potentially allow sewage-derived ARG to persist and spread in the environment. This may create increased opportunities for an eventual contact with, and gene transfer to, human and animal pathogens via the food chain or drinking water. However, assessment of this risk requires a better understanding of the level and variability of the natural resistance background and the extent of the human impact. We have analyzed water samples from 21 Swiss lakes, taken at sampling points that were not under the direct influence of local contamination sources and analyzed the relative abundance of ARG using quantitative real-time PCR. Copy numbers of genes mediating resistance to three different broad-spectrum antibiotic classes (sulfonamides: sul1, sul2, tetracyclines: tet(B), tet(M), tet(W) and fluoroquinolones: qnrA) were normalized to copy numbers of bacterial 16S rRNA genes. We used multiple linear regression to assess if ARG abundance is related to human activities in the catchment, microbial community composition and the eutrophication status of the lakes. Sul genes were detected in all sampled lakes, whereas only four lakes contained quantifiable numbers of tet genes, and qnrA remained below detection in all lakes. Our data indicate higher abundance of sul1 in lakes with increasing number and capacity of wastewater treatment plants (WWTPs) in the catchment. sul2 abundance was rather related to long water residence times and eutrophication status. Our study demonstrates the potential of freshwater lakes to preserve antibiotic resistance genes, and provides a reference for ARG abundance from lake systems with low human impact as a baseline for assessing ARG contamination in lake water.

  14. Impact of pe_pgrs33 Gene Polymorphisms on Mycobacterium tuberculosis Infection and Pathogenesis.

    PubMed

    Camassa, Serena; Palucci, Ivana; Iantomasi, Raffaella; Cubeddu, Tiziana; Minerva, Mariachiara; De Maio, Flavio; Jouny, Samuel; Petruccioli, Elisa; Goletti, Delia; Ria, Francesco; Sali, Michela; Sanguinetti, Maurizio; Manganelli, Riccardo; Rocca, Stefano; Brodin, Priscille; Delogu, Giovanni

    2017-01-01

    PE_PGRS33 is a surface-exposed protein of Mycobacterium tuberculosis (Mtb) which exerts its role in macrophages entry and immunomodulation. In this study, we aimed to investigate the polymorphisms in the pe_pgrs33 gene of Mtb clinical isolates and evaluate their impact on protein functions. We sequenced pe_pgrs33 in a collection of 135 clinical strains, genotyped by 15-loci MIRU-VNTR and spoligotyping and belonging to the Mtb complex (MTBC). Overall, an association between pe_pgrs33 alleles and MTBC genotypes was observed and a dN/dS ratio of 0.64 was obtained, suggesting that a purifying selective pressure is acting on pe_pgrs33 against deleterious SNPs. Among a total of 19 pe_pgrs33 alleles identified in this study, 5 were cloned and used to complement the pe_pgrs33 knock-out mutant strain of Mtb H37Rv (MtbΔ33) to assess the functional impact of the respective polymorphisms in in vitro infections of primary macrophages. In human monocyte-derived macrophages (MDMs) infection, large in-frame and frameshift mutations were unable to restore the phenotype of Mtb H37Rv, impairing the cell entry capacity of Mtb, but neither its intracellular replication rate nor its immunomodulatory properties. In vivo studies performed in the murine model of tuberculosis (TB) demonstrated that the MtbΔ33 mutant strain was not impaired in the ability to infect and replicate in the lung tissue compared to the parental strain. Interestingly, MtbΔ33 showed an enhanced virulence during the chronic steps of infection compared to Mtb H37Rv. Similarly, the complementation of MtbΔ33 with a frameshift allele also resulted in a Mtb strain capable of causing a surprisingly enhanced tissue damage in murine lungs, during the chronic steps of infection. Together, these results further support the role of PE_PGRS33 in the pathogenesis and virulence of Mtb.

  15. Impact of mosquito gene drive on malaria elimination in a computational model with explicit spatial and temporal dynamics.

    PubMed

    Eckhoff, Philip A; Wenger, Edward A; Godfray, H Charles J; Burt, Austin

    2017-01-10

    The renewed effort to eliminate malaria and permanently remove its tremendous burden highlights questions of what combination of tools would be sufficient in various settings and what new tools need to be developed. Gene drive mosquitoes constitute a promising set of tools, with multiple different possible approaches including population replacement with introduced genes limiting malaria transmission, driving-Y chromosomes to collapse a mosquito population, and gene drive disrupting a fertility gene and thereby achieving population suppression or collapse. Each of these approaches has had recent success and advances under laboratory conditions, raising the urgency for understanding how each could be deployed in the real world and the potential impacts of each. New analyses are needed as existing models of gene drive primarily focus on nonseasonal or nonspatial dynamics. We use a mechanistic, spatially explicit, stochastic, individual-based mathematical model to simulate each gene drive approach in a variety of sub-Saharan African settings. Each approach exhibits a broad region of gene construct parameter space with successful elimination of malaria transmission due to the targeted vector species. The introduction of realistic seasonality in vector population dynamics facilitates gene drive success compared with nonseasonal analyses. Spatial simulations illustrate constraints on release timing, frequency, and spatial density in the most challenging settings for construct success. Within its parameter space for success, each gene drive approach provides a tool for malaria elimination unlike anything presently available. Provided potential barriers to success are surmounted, each achieves high efficacy at reducing transmission potential and lower delivery requirements in logistically challenged settings.

  16. Impact of mosquito gene drive on malaria elimination in a computational model with explicit spatial and temporal dynamics

    PubMed Central

    Eckhoff, Philip A.; Wenger, Edward A.; Godfray, H. Charles J.; Burt, Austin

    2017-01-01

    The renewed effort to eliminate malaria and permanently remove its tremendous burden highlights questions of what combination of tools would be sufficient in various settings and what new tools need to be developed. Gene drive mosquitoes constitute a promising set of tools, with multiple different possible approaches including population replacement with introduced genes limiting malaria transmission, driving-Y chromosomes to collapse a mosquito population, and gene drive disrupting a fertility gene and thereby achieving population suppression or collapse. Each of these approaches has had recent success and advances under laboratory conditions, raising the urgency for understanding how each could be deployed in the real world and the potential impacts of each. New analyses are needed as existing models of gene drive primarily focus on nonseasonal or nonspatial dynamics. We use a mechanistic, spatially explicit, stochastic, individual-based mathematical model to simulate each gene drive approach in a variety of sub-Saharan African settings. Each approach exhibits a broad region of gene construct parameter space with successful elimination of malaria transmission due to the targeted vector species. The introduction of realistic seasonality in vector population dynamics facilitates gene drive success compared with nonseasonal analyses. Spatial simulations illustrate constraints on release timing, frequency, and spatial density in the most challenging settings for construct success. Within its parameter space for success, each gene drive approach provides a tool for malaria elimination unlike anything presently available. Provided potential barriers to success are surmounted, each achieves high efficacy at reducing transmission potential and lower delivery requirements in logistically challenged settings. PMID:28028208

  17. Determinants of Human Adipose Tissue Gene Expression: Impact of Diet, Sex, Metabolic Status, and Cis Genetic Regulation

    PubMed Central

    Viguerie, Nathalie; Montastier, Emilie; Maoret, Jean-José; Roussel, Balbine; Combes, Marion; Valle, Carine; Villa-Vialaneix, Nathalie; Iacovoni, Jason S.; Martinez, J. Alfredo; Holst, Claus; Astrup, Arne; Vidal, Hubert; Clément, Karine; Hager, Jorg; Saris, Wim H. M.; Langin, Dominique

    2012-01-01

    Weight control diets favorably affect parameters of the metabolic syndrome and delay the onset of diabetic complications. The adaptations occurring in adipose tissue (AT) are likely to have a profound impact on the whole body response as AT is a key target of dietary intervention. Identification of environmental and individual factors controlling AT adaptation is therefore essential. Here, expression of 271 transcripts, selected for regulation according to obesity and weight changes, was determined in 515 individuals before, after 8-week low-calorie diet-induced weight loss, and after 26-week ad libitum weight maintenance diets. For 175 genes, opposite regulation was observed during calorie restriction and weight maintenance phases, independently of variations in body weight. Metabolism and immunity genes showed inverse profiles. During the dietary intervention, network-based analyses revealed strong interconnection between expression of genes involved in de novo lipogenesis and components of the metabolic syndrome. Sex had a marked influence on AT expression of 88 transcripts, which persisted during the entire dietary intervention and after control for fat mass. In women, the influence of body mass index on expression of a subset of genes persisted during the dietary intervention. Twenty-two genes revealed a metabolic syndrome signature common to men and women. Genetic control of AT gene expression by cis signals was observed for 46 genes. Dietary intervention, sex, and cis genetic variants independently controlled AT gene expression. These analyses help understanding the relative importance of environmental and individual factors that control the expression of human AT genes and therefore may foster strategies aimed at improving AT function in metabolic diseases. PMID:23028366

  18. Impact of the PDE4D gene polymorphism and additional SNP-SNP and gene-smoking interaction on ischemic stroke risk in Chinese Han population.

    PubMed

    Wang, Xianxiang; Sun, Zhongwu; Zhang, Yiquan; Tian, Xuefeng; Li, Qingxin; Luo, Jing

    2017-04-01

    To investigate the association between phosphodiesterase 4D gene (PDE4D) gene single nucleotide polymorphisms (SNPs) and ischemic stroke (IS) risk, and impact of additional SNP- SNP and gene- smoking interaction on IS risk in Chinese population. A total of 1228 subjects (666 males, 562 females) were selected, including 610 IS patients and 618 control subjects. Logistic regression model was used to examine the association between SNPs in PDE4D gene and IS risk. Generalized multifactor dimensionality reduction (GMDR) was employed to analyze the SNP- SNP and gene- smoking interaction. IS risks were significantly higher in carriers of A allele of rs12188950 polymorphism than those with GG genotype (GA + AA vs. GG), adjusted OR (95%CI) = 1.61 (1.26-2.19), and also significantly higher in carriers of T allele of rs966221 polymorphism than those with CC (CT + TT vs. CC), adjusted OR (95%CI) = 1.82 (1.39-2.23). We found that there was a significant SNP- SNP interaction between rs966221 and rs12188950. Subjects with CT or TT of rs966221 and GA or AA of rs12188950 genotype have the highest IS risk, compared to subjects with CC of rs966221 and GG of rs12188950 genotype, OR (95%CI) was 3.52 (2.68-4.69). We also found a significant gene-environment interaction between rs966221 and smoking. Smokers with CT or TT of rs966221 genotype have the highest IS risk, compared to never smokers with CC of rs966221 genotype, OR (95%CI) was 3.97 (2.25-5.71). Our results support an important association of rs966221 and rs12188950 minor allele and its interaction with increased risk of IS risk, and additional interaction between rs966221 and smoking.

  19. Impact of deletion of envelope-related genes of recombinant Sendai viruses on immune responses following pulmonary gene transfer of neonatal mice.

    PubMed

    Tanaka, S; Yonemitsu, Y; Yoshida, K; Okano, S; Kondo, H; Inoue, M; Hasegawa, M; Masumoto, K; Suita, S; Taguchi, T; Sueishi, K

    2007-07-01

    We demonstrated previously that the additive-type recombinant Sendai virus (rSeV) is highly efficient for use in pulmonary gene transfer; however, rSeV exhibits inflammatory responses. To overcome this problem, we tested newly developed non-transmissible constructs, namely, temperature-sensitive F-deleted vector, rSeV/dF (ts-rSeV/dF) and a rSeV with all the envelope-related genes deleted (rSeV/dFdMdHN), for pulmonary gene transfer in neonatal mice, by assessing their toxicity and immune responses. The gene expression in the lungs of neonatal ICR mice peaked on day 2, then gradually decreased until almost disappearing at 14 days after infection in all constructs. Loss of body weight and mortality rate, however, were dramatically improved in mice treated with SeV/dFdMdHN (mortality=0%, n=41) and ts-rSeV/dF (24.2%, n=33) compared with additive rSeV (70.7%, n=58). Although the deletion of envelope-related genes of SeV had a small impact on the production of antibody and cytotoxic T-lymphocyte activity in both adults and neonates, a dramatic reduction was found in the events related to innate responses, including the production of proinflammatory cytokines, particularly in the case of neonates. These results indicate that pulmonary gene transfer using SeV/dFdMdHN warrants further investigation for its possible use in developing safer therapeutics for neonatal lung diseases, including cystic fibrosis.

  20. Evolutionary analysis of multidrug resistance genes in fungi - impact of gene duplication and family conservation.

    PubMed

    Gossani, Cristiani; Bellieny-Rabelo, Daniel; Venancio, Thiago M

    2014-11-01

    Although the emergence of bacterial drug resistance is of great concern to the scientific community, few studies have evaluated this phenomenon systematically in fungi by using genome-wide datasets. In the present study, we assembled a large compendium of Saccharomyces cerevisiae chemical genetic data to study the evolution of multidrug resistance genes (MDRs) in the fungal lineage. We found that MDRs typically emerge in widely conserved families, most of which containing homologs from pathogenic fungi, such as Candida albicans and Coccidioides immitis, which could favor the evolution of drug resistance in those species. By integrating data from chemical genetics with protein family conservation, genetic and protein interactions, we found that gene families rarely have more than one MDR, indicating that paralogs evolve asymmetrically with regard to multidrug resistance roles. Furthermore, MDRs have more genetic and protein interaction partners than non-MDRs, supporting their participation in complex biochemical systems underlying the tolerance to multiple bioactive molecules. MDRs share more chemical genetic interactions with other MDRs than with non-MDRs, regardless of their evolutionary affinity. These results suggest the existence of an intricate system involved in the global drug tolerance phenotypes. Finally, MDRs are more likely to be hit repeatedly by mutations in laboratory evolution experiments, indicating that they have great adaptive potential. The results presented here not only reveal the main genomic features underlying the evolution of MDRs, but also shed light on the gene families from which drug resistance is more likely to emerge in fungi.

  1. The Gene Messenger Impact Project: An Innovative Genetics Continuing Education Strategy for Primary Care Providers.

    PubMed

    Carroll, June C; Grad, Roland; Allanson, Judith E; Pluye, Pierre; Permaul, Joanne A; Pimlott, Nicholas; Wilson, Brenda J

    2016-01-01

    Primary care providers (PCP) will need to be integrally involved in the delivery of genomic medicine. The GenetiKit trial demonstrated effectiveness of a knowledge translation intervention on family physicians' (FP) genetics referral decision-making. Most wanted to continue receiving Gene Messengers (GM), evidence-based summaries of new genetic tests with primary care recommendations. Our objective was to determine the value of GMs as a continuing education (CE) strategy in genomic medicine for FPs. Using a "push" model, we invited 19,060 members of the College of Family Physicians of Canada to participate. Participants read GMs online, receiving 12 emailed topics over 6 months. Participants completed an online Information Assessment Method questionnaire evaluating GMs on four constructs: cognitive impact, relevance, intended use of information for a patient, and expected health benefits. One thousand four hundred two FPs participated, 55% rated at least one GM. Most (73%) indicated their practice would be improved after reading GMs, with referral to genetics ranked highly. Of those who rated a GM relevant, 94% would apply it to at least one patient and 79% would expect health benefits. This method of CE was found useful for genetics by 88% and 94% wanted to continue receiving GMs. FPs found this novel CE strategy, brief individual reflective e-learning, to be valuable for learning about genetics. This method of information delivery may be an especially effective method for CE in genomic medicine where discoveries occur at a rapid pace and lack of knowledge is a barrier to integration of genetic services.

  2. Widespread impact of horizontal gene transfer on plant colonization of land

    PubMed Central

    Yue, Jipei; Hu, Xiangyang; Sun, Hang; Yang, Yongping; Huang, Jinling

    2012-01-01

    In complex multicellular eukaryotes such as animals and plants, horizontal gene transfer is commonly considered rare with very limited evolutionary significance. Here we show that horizontal gene transfer is a dynamic process occurring frequently in the early evolution of land plants. Our genome analyses of the moss Physcomitrella patens identified 57 families of nuclear genes that were acquired from prokaryotes, fungi or viruses. Many of these gene families were transferred to the ancestors of green or land plants. Available experimental evidence shows that these anciently acquired genes are involved in some essential or plant-specific activities such as xylem formation, plant defence, nitrogen recycling as well as the biosynthesis of starch, polyamines, hormones and glutathione. These findings suggest that horizontal gene transfer had a critical role in the transition of plants from aquatic to terrestrial environments. On the basis of these findings, we propose a model of horizontal gene transfer mechanism in nonvascular and seedless vascular plants. PMID:23093189

  3. Widespread impact of horizontal gene transfer on plant colonization of land.

    PubMed

    Yue, Jipei; Hu, Xiangyang; Sun, Hang; Yang, Yongping; Huang, Jinling

    2012-01-01

    In complex multicellular eukaryotes such as animals and plants, horizontal gene transfer is commonly considered rare with very limited evolutionary significance. Here we show that horizontal gene transfer is a dynamic process occurring frequently in the early evolution of land plants. Our genome analyses of the moss Physcomitrella patens identified 57 families of nuclear genes that were acquired from prokaryotes, fungi or viruses. Many of these gene families were transferred to the ancestors of green or land plants. Available experimental evidence shows that these anciently acquired genes are involved in some essential or plant-specific activities such as xylem formation, plant defence, nitrogen recycling as well as the biosynthesis of starch, polyamines, hormones and glutathione. These findings suggest that horizontal gene transfer had a critical role in the transition of plants from aquatic to terrestrial environments. On the basis of these findings, we propose a model of horizontal gene transfer mechanism in nonvascular and seedless vascular plants.

  4. Cancer predisposition genes: molecular mechanisms and clinical impact on personalized cancer care: examples of Lynch and HBOC syndromes.

    PubMed

    Wang, Qing

    2016-02-01

    Up to 10% of cancers occur through the inherited mutation of a group of genes called cancer predisposition genes. Individuals who carry a mutant allele of these genes have an increased susceptibility to cancer. A growing number of cancer susceptibility genes are being identified, and the physiopathology of germline mutation-based cancer development is also being elucidated with accumulating clinical and molecular data. More importantly, the identification of familial mutations has become routine practice, which is a perfect example of bench-to-bed translational medicine. Recently, other clinical applications of predisposition genes have been exploited, especially as efficient biomarkers predicting prognosis or response to treatment. Thus, it appears interesting to give an overview of the advances and impacts of predisposition genes in personalized cancer care by taking representative and common cancer syndromes as examples: Lynch syndrome for the first example, which is related to cancer susceptibility, and breast and ovary cancer syndrome for the second example, which involves BRCA deficiency-related targeted therapy.

  5. Classification of human chromosome 21 gene-expression variations in Down syndrome: impact on disease phenotypes.

    PubMed

    Aït Yahya-Graison, E; Aubert, J; Dauphinot, L; Rivals, I; Prieur, M; Golfier, G; Rossier, J; Personnaz, L; Creau, N; Bléhaut, H; Robin, S; Delabar, J M; Potier, M-C

    2007-09-01

    Down syndrome caused by chromosome 21 trisomy is the most common genetic cause of mental retardation in humans. Disruption of the phenotype is thought to be the result of gene-dosage imbalance. Variations in chromosome 21 gene expression in Down syndrome were analyzed in lymphoblastoid cells derived from patients and control individuals. Of the 359 genes and predictions displayed on a specifically designed high-content chromosome 21 microarray, one-third were expressed in lymphoblastoid cells. We performed a mixed-model analysis of variance to find genes that are differentially expressed in Down syndrome independent of sex and interindividual variations. In addition, we identified genes with variations between Down syndrome and control samples that were significantly different from the gene-dosage effect (1.5). Microarray data were validated by quantitative polymerase chain reaction. We found that 29% of the expressed chromosome 21 transcripts are overexpressed in Down syndrome and correspond to either genes or open reading frames. Among these, 22% are increased proportional to the gene-dosage effect, and 7% are amplified. The other 71% of expressed sequences are either compensated (56%, with a large proportion of predicted genes and antisense transcripts) or highly variable among individuals (15%). Thus, most of the chromosome 21 transcripts are compensated for the gene-dosage effect. Overexpressed genes are likely to be involved in the Down syndrome phenotype, in contrast to the compensated genes. Highly variable genes could account for phenotypic variations observed in patients. Finally, we show that alternative transcripts belonging to the same gene are similarly regulated in Down syndrome but sense and antisense transcripts are not.

  6. Classification of Human Chromosome 21 Gene-Expression Variations in Down Syndrome: Impact on Disease Phenotypes

    PubMed Central

    Aït Yahya-Graison, E. ; Aubert, J. ; Dauphinot, L. ; Rivals, I. ; Prieur, M. ; Golfier, G. ; Rossier, J. ; Personnaz, L. ; Créau, N. ; Bléhaut, H. ; Robin, S. ; Delabar, J. M. ; Potier, M.-C. 

    2007-01-01

    Down syndrome caused by chromosome 21 trisomy is the most common genetic cause of mental retardation in humans. Disruption of the phenotype is thought to be the result of gene-dosage imbalance. Variations in chromosome 21 gene expression in Down syndrome were analyzed in lymphoblastoid cells derived from patients and control individuals. Of the 359 genes and predictions displayed on a specifically designed high-content chromosome 21 microarray, one-third were expressed in lymphoblastoid cells. We performed a mixed-model analysis of variance to find genes that are differentially expressed in Down syndrome independent of sex and interindividual variations. In addition, we identified genes with variations between Down syndrome and control samples that were significantly different from the gene-dosage effect (1.5). Microarray data were validated by quantitative polymerase chain reaction. We found that 29% of the expressed chromosome 21 transcripts are overexpressed in Down syndrome and correspond to either genes or open reading frames. Among these, 22% are increased proportional to the gene-dosage effect, and 7% are amplified. The other 71% of expressed sequences are either compensated (56%, with a large proportion of predicted genes and antisense transcripts) or highly variable among individuals (15%). Thus, most of the chromosome 21 transcripts are compensated for the gene-dosage effect. Overexpressed genes are likely to be involved in the Down syndrome phenotype, in contrast to the compensated genes. Highly variable genes could account for phenotypic variations observed in patients. Finally, we show that alternative transcripts belonging to the same gene are similarly regulated in Down syndrome but sense and antisense transcripts are not. PMID:17701894

  7. Phylogenetic diversity of archaea and the archaeal ammonia monooxygenase gene in uranium mining-impacted locations in Bulgaria.

    PubMed

    Radeva, Galina; Kenarova, Anelia; Bachvarova, Velina; Flemming, Katrin; Popov, Ivan; Vassilev, Dimitar; Selenska-Pobell, Sonja

    2014-01-01

    Uranium mining and milling activities adversely affect the microbial populations of impacted sites. The negative effects of uranium on soil bacteria and fungi are well studied, but little is known about the effects of radionuclides and heavy metals on archaea. The composition and diversity of archaeal communities inhabiting the waste pile of the Sliven uranium mine and the soil of the Buhovo uranium mine were investigated using 16S rRNA gene retrieval. A total of 355 archaeal clones were selected, and their 16S rDNA inserts were analysed by restriction fragment length polymorphism (RFLP) discriminating 14 different RFLP types. All evaluated archaeal 16S rRNA gene sequences belong to the 1.1b/Nitrososphaera cluster of Crenarchaeota. The composition of the archaeal community is distinct for each site of interest and dependent on environmental characteristics, including pollution levels. Since the members of 1.1b/Nitrososphaera cluster have been implicated in the nitrogen cycle, the archaeal communities from these sites were probed for the presence of the ammonia monooxygenase gene (amoA). Our data indicate that amoA gene sequences are distributed in a similar manner as in Crenarchaeota, suggesting that archaeal nitrification processes in uranium mining-impacted locations are under the control of the same key factors controlling archaeal diversity.

  8. Phylogenetic Diversity of Archaea and the Archaeal Ammonia Monooxygenase Gene in Uranium Mining-Impacted Locations in Bulgaria

    PubMed Central

    Radeva, Galina; Kenarova, Anelia; Bachvarova, Velina; Popov, Ivan; Selenska-Pobell, Sonja

    2014-01-01

    Uranium mining and milling activities adversely affect the microbial populations of impacted sites. The negative effects of uranium on soil bacteria and fungi are well studied, but little is known about the effects of radionuclides and heavy metals on archaea. The composition and diversity of archaeal communities inhabiting the waste pile of the Sliven uranium mine and the soil of the Buhovo uranium mine were investigated using 16S rRNA gene retrieval. A total of 355 archaeal clones were selected, and their 16S rDNA inserts were analysed by restriction fragment length polymorphism (RFLP) discriminating 14 different RFLP types. All evaluated archaeal 16S rRNA gene sequences belong to the 1.1b/Nitrososphaera cluster of Crenarchaeota. The composition of the archaeal community is distinct for each site of interest and dependent on environmental characteristics, including pollution levels. Since the members of 1.1b/Nitrososphaera cluster have been implicated in the nitrogen cycle, the archaeal communities from these sites were probed for the presence of the ammonia monooxygenase gene (amoA). Our data indicate that amoA gene sequences are distributed in a similar manner as in Crenarchaeota, suggesting that archaeal nitrification processes in uranium mining-impacted locations are under the control of the same key factors controlling archaeal diversity. PMID:24711725

  9. [Impact of SIRT1 gene expression on the development and treatment of the metabolic syndrome in oncological patients].

    PubMed

    Wawryka, Joanna; Barg, Ewa

    2016-01-01

    Sirtuins - products of gene SIRT expression have been divided into 7 classes, according to the amino acid composition and location of the cell. Those factors, called longevities proteins, are a group of histone deacetylases, depend on nicotinamide adenine dinucleotide (NAD). Particularly noteworthy is the protein sirtuin 1, which further deacetylates numerous transcription factors, receptors and enzymes. Through its action reduces the activity of glucocorticoid receptors in the body. Products of gene SIRT1 expression is responsible for apoptosis, differentiation, senescence cells, also affect the regulation of carbohydrate and lipid metabolism. Cardioprotective and hypotensive impact is also very important. SIRT1 reduces the accumulation of fat and decreases the risk of visceral obesity. Low gene expression of SIRT1 therefore predispose to the development of metabolic syndrome. Homeostasis sirtuin 1 disorders can also be observed in certain neoplastic diseases, primarily hormone-dependent breast, ovarian and prostate cancer, as well as it can cause leukemias and lymphomas. Components, activating expression of gen SIRT1 or a molecule with biological properties sirtuin 1, may have promising impact for treatment of diabetes mellitus type 2, obesity, hypertension, dyslipidemia. Analyzing, the pleiotropic effect of sirtuin 1 and numerous metabolic pathways, appear to be particularly beneficial effect of supplementation molecules increasing the level of expression gene SIRT1, in treatment of acute lymphoblastic leukemia with using high-dosing glicocorticosteroid therapy. Which would reduce the number of early and late complications of oncological treatment and increase patient survival. Compound requires further study.

  10. Experimental evidence of genome-wide impact of ecological selection during early stages of speciation-with-gene-flow.

    PubMed

    Egan, Scott P; Ragland, Gregory J; Assour, Lauren; Powell, Thomas H Q; Hood, Glen R; Emrich, Scott; Nosil, Patrik; Feder, Jeffrey L

    2015-08-01

    Theory predicts that speciation-with-gene-flow is more likely when the consequences of selection for population divergence transitions from mainly direct effects of selection acting on individual genes to a collective property of all selected genes in the genome. Thus, understanding the direct impacts of ecologically based selection, as well as the indirect effects due to correlations among loci, is critical to understanding speciation. Here, we measure the genome-wide impacts of host-associated selection between hawthorn and apple host races of Rhagoletis pomonella (Diptera: Tephritidae), a model for contemporary speciation-with-gene-flow. Allele frequency shifts of 32 455 SNPs induced in a selection experiment based on host phenology were genome wide and highly concordant with genetic divergence between co-occurring apple and hawthorn flies in nature. This striking genome-wide similarity between experimental and natural populations of R. pomonella underscores the importance of ecological selection at early stages of divergence and calls for further integration of studies of eco-evolutionary dynamics and genome divergence. © 2015 The Authors Ecology Letters published by John Wiley & Sons Ltd and CNRS.

  11. Genomic Data Quality Impacts Automated Detection of Lateral Gene Transfer in Fungi

    PubMed Central

    Dupont, Pierre-Yves; Cox, Murray P.

    2017-01-01

    Lateral gene transfer (LGT, also known as horizontal gene transfer), an atypical mechanism of transferring genes between species, has almost become the default explanation for genes that display an unexpected composition or phylogeny. Numerous methods of detecting LGT events all rely on two fundamental strategies: primary structure composition or gene tree/species tree comparisons. Discouragingly, the results of these different approaches rarely coincide. With the wealth of genome data now available, detection of laterally transferred genes is increasingly being attempted in large uncurated eukaryotic datasets. However, detection methods depend greatly on the quality of the underlying genomic data, which are typically complex for eukaryotes. Furthermore, given the automated nature of genomic data collection, it is typically impractical to manually verify all protein or gene models, orthology predictions, and multiple sequence alignments, requiring researchers to accept a substantial margin of error in their datasets. Using a test case comprising plant-associated genomes across the fungal kingdom, this study reveals that composition- and phylogeny-based methods have little statistical power to detect laterally transferred genes. In particular, phylogenetic methods reveal extreme levels of topological variation in fungal gene trees, the vast majority of which show departures from the canonical species tree. Therefore, it is inherently challenging to detect LGT events in typical eukaryotic genomes. This finding is in striking contrast to the large number of claims for laterally transferred genes in eukaryotic species that routinely appear in the literature, and questions how many of these proposed examples are statistically well supported. PMID:28235827

  12. Antimicrobial-Resistant Bacterial Populations and Antimicrobial Resistance Genes Obtained from Environments Impacted by Livestock and Municipal Waste.

    PubMed

    Agga, Getahun E; Arthur, Terrance M; Durso, Lisa M; Harhay, Dayna M; Schmidt, John W

    2015-01-01

    This study compared the populations of antimicrobial-resistant bacteria and the repertoire of antimicrobial resistance genes in four environments: effluent of three municipal wastewater treatment facilities, three cattle feedlot runoff catchment ponds, three swine waste lagoons, and two "low impact" environments (an urban lake and a relict prairie). Multiple liquid and solid samples were collected from each environment. The prevalences and concentrations of antimicrobial-resistant (AMR) Gram-negative (Escherichia coli and Salmonella enterica) and Gram-positive (enterococci) bacteria were determined from individual samples (n = 174). The prevalences of 84 antimicrobial resistance genes in metagenomic DNA isolated from samples pooled (n = 44) by collection date, location, and sample type were determined. The prevalences and concentrations of AMR E. coli and Salmonella were similar among the livestock and municipal sample sources. The levels of erythromycin-resistant enterococci were significantly higher in liquid samples from cattle catchment ponds and swine waste lagoons than in liquid samples from municipal wastewater treatment facilities, but solid samples from these environments did not differ significantly. Similarly, trimethoprim/sulfamethoxazole-resistant E. coli concentrations were significantly higher in swine liquid than in municipal liquid samples, but there was no difference in solid samples. Multivariate analysis of the distribution of antimicrobial resistance genes using principal coordinate analysis showed distinct clustering of samples with livestock (cattle and swine), low impact environment and municipal samples forming three separate clusters. The numbers of class A beta-lactamase, class C beta-lactamase, and fluoroquinolone resistance genes detected were significantly higher (P < 0.05) in municipal samples than in cattle runoff or swine lagoon samples. In conclusion, we report that AMR is a very widespread phenomenon and that similar prevalences

  13. B chromosome contains active genes and impacts the transcription of A chromosomes in maize (Zea mays L.).

    PubMed

    Huang, Wei; Du, Yan; Zhao, Xin; Jin, Weiwei

    2016-04-16

    The dispensable maize (Zea mays L.) B chromosome is highly heterochromatic and widely believed to be devoid of functional genes. Although low-copy B chromosome causes no obvious phenotype variation, its existence might influence A genome gene expression. Previous studies suggested that B chromosomes are evolved from standard chromosomes; therefore, they might contain genic regions showing homology with A chromosome sequences. Our data suggested that maize B chromosome influences the A-genome transcription with stronger effect associated with an increase in copy number of B chromosome. In total 130 differently expressed genes were detected in comparison between with and without B chromosome lines. These differentially expressed genes are mainly involved in cell metabolism and nucleotide binding. Using Starter + B, we amplified ten B chromosome loci with high sequence similarity to A-genome genes. Fluorescence in situ hybridization (FISH) confirmed that at least four ~5 kb-sized genes are located on the B chromosome. In addition, through de novo assembly of the reads not unmapped to maize B73 reference genome together with PCR validation, we found three B-located LTR; in particular, one of them, the 3.2 kb comp75688, is expressed in a B-dosage dependent manner. We found that in the presence of maize B chromosome, the transcription of A genome genes was altered, with more impact by the increase of the B chromosome number. The B-located transcriptionally active genes showed high similarity to their A-genome homologues, and retrotransposons on B chromosome also have partial homologous to A genome sequences. Our data shed more lights on the genome structure and evolution of the maize B chromosome.

  14. Kinase impact assessment in the landscape of fusion genes that retain kinase domains: a pan-cancer study.

    PubMed

    Kim, Pora; Jia, Peilin; Zhao, Zhongming

    2016-12-24

    Assessing the impact of kinase in gene fusion is essential for both identifying driver fusion genes (FGs) and developing molecular targeted therapies. Kinase domain retention is a crucial factor in kinase fusion genes (KFGs), but such a systematic investigation has not been done yet. To this end, we analyzed kinase domain retention (KDR) status in chimeric protein sequences of 914 KFGs covering 312 kinases across 13 major cancer types. Based on 171 kinase domain-retained KFGs including 101 kinases, we studied their recurrence, kinase groups, fusion partners, exon-based expression depth, short DNA motifs around the break points and networks. Our results, such as more KDR than 5'-kinase fusion genes, combinatorial effects between 3'-KDR kinases and their 5'-partners and a signal transduction-specific DNA sequence motif in the break point intronic sequences, supported positive selection on 3'-kinase fusion genes in cancer. We introduced a degree-of-frequency (DoF) score to measure the possible number of KFGs of a kinase. Interestingly, kinases with high DoF scores tended to undergo strong gene expression alteration at the break points. Furthermore, our KDR gene fusion network analysis revealed six of the seven kinases with the highest DoF scores (ALK, BRAF, MET, NTRK1, NTRK3 and RET) were all observed in thyroid carcinoma. Finally, we summarized common features of 'effective' (highly recurrent) kinases in gene fusions such as expression alteration at break point, redundant usage in multiple cancer types and 3'-location tendency. Collectively, our findings are useful for prioritizing driver kinases and FGs and provided insights into KFGs' clinical implications. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  15. Impact of DNA vector topology on non-viral gene therapeutic safety and efficacy.

    PubMed

    Sum, Chi H; Wettig, Shawn; Slavcev, Roderick A

    2014-01-01

    Gene therapy continues to grow as an emerging treatment strategy toward numerous diseases. However, such prospects are hindered by the use of viral vectors prompting significant safety concerns along with limitations concerning repeat administrations, size of delivered gene construct, scale-up, high production costs, contamination during production, and lack of desired tissue selectivity. Non-viral gene delivery demonstrates the potential to address the abovementioned limitations, but itself generally suffers from low efficacy. Continuing efforts have been made to develop innovative delivery systems, synthetic gene carriers, and DNA vectors in a concerted attempt to enhance gene delivery suitable for clinical applications. In this review, we focus on the advances in the design of novel DNA vectors catered to enhance transfection and transgene expression and their influences on the efficacy and safety of existing and emerging delivery systems and synthetic vectors for non viral gene delivery.

  16. Does your gene need a background check? How genetic background impacts the analysis of mutations, genes, and evolution.

    PubMed

    Chandler, Christopher H; Chari, Sudarshan; Dworkin, Ian

    2013-06-01

    The premise of genetic analysis is that a causal link exists between phenotypic and allelic variation. However, it has long been documented that mutant phenotypes are not a simple result of a single DNA lesion, but are instead due to interactions of the focal allele with other genes and the environment. Although an experimentally rigorous approach focused on individual mutations and isogenic control strains has facilitated amazing progress within genetics and related fields, a glimpse back suggests that a vast complexity has been omitted from our current understanding of allelic effects. Armed with traditional genetic analyses and the foundational knowledge they have provided, we argue that the time and tools are ripe to return to the underexplored aspects of gene function and embrace the context-dependent nature of genetic effects. We assert that a broad understanding of genetic effects and the evolutionary dynamics of alleles requires identifying how mutational outcomes depend upon the 'wild type' genetic background. Furthermore, we discuss how best to exploit genetic background effects to broaden genetic research programs.

  17. Gene Model Annotations for Drosophila melanogaster: Impact of High-Throughput Data

    PubMed Central

    Matthews, Beverley B.; dos Santos, Gilberto; Crosby, Madeline A.; Emmert, David B.; St. Pierre, Susan E.; Gramates, L. Sian; Zhou, Pinglei; Schroeder, Andrew J.; Falls, Kathleen; Strelets, Victor; Russo, Susan M.; Gelbart, William M.

    2015-01-01

    We report the current status of the FlyBase annotated gene set for Drosophila melanogaster and highlight improvements based on high-throughput data. The FlyBase annotated gene set consists entirely of manually annotated gene models, with the exception of some classes of small non-coding RNAs. All gene models have been reviewed using evidence from high-throughput datasets, primarily from the modENCODE project. These datasets include RNA-Seq coverage data, RNA-Seq junction data, transcription start site profiles, and translation stop-codon read-through predictions. New annotation guidelines were developed to take into account the use of the high-throughput data. We describe how this flood of new data was incorporated into thousands of new and revised annotations. FlyBase has adopted a philosophy of excluding low-confidence and low-frequency data from gene model annotations; we also do not attempt to represent all possible permutations for complex and modularly organized genes. This has allowed us to produce a high-confidence, manageable gene annotation dataset that is available at FlyBase (http://flybase.org). Interesting aspects of new annotations include new genes (coding, non-coding, and antisense), many genes with alternative transcripts with very long 3′ UTRs (up to 15–18 kb), and a stunning mismatch in the number of male-specific genes (approximately 13% of all annotated gene models) vs. female-specific genes (less than 1%). The number of identified pseudogenes and mutations in the sequenced strain also increased significantly. We discuss remaining challenges, for instance, identification of functional small polypeptides and detection of alternative translation starts. PMID:26109357

  18. Gene Model Annotations for Drosophila melanogaster: Impact of High-Throughput Data.

    PubMed

    Matthews, Beverley B; Dos Santos, Gilberto; Crosby, Madeline A; Emmert, David B; St Pierre, Susan E; Gramates, L Sian; Zhou, Pinglei; Schroeder, Andrew J; Falls, Kathleen; Strelets, Victor; Russo, Susan M; Gelbart, William M

    2015-06-24

    We report the current status of the FlyBase annotated gene set for Drosophila melanogaster and highlight improvements based on high-throughput data. The FlyBase annotated gene set consists entirely of manually annotated gene models, with the exception of some classes of small non-coding RNAs. All gene models have been reviewed using evidence from high-throughput datasets, primarily from the modENCODE project. These datasets include RNA-Seq coverage data, RNA-Seq junction data, transcription start site profiles, and translation stop-codon read-through predictions. New annotation guidelines were developed to take into account the use of the high-throughput data. We describe how this flood of new data was incorporated into thousands of new and revised annotations. FlyBase has adopted a philosophy of excluding low-confidence and low-frequency data from gene model annotations; we also do not attempt to represent all possible permutations for complex and modularly organized genes. This has allowed us to produce a high-confidence, manageable gene annotation dataset that is available at FlyBase (http://flybase.org). Interesting aspects of new annotations include new genes (coding, non-coding, and antisense), many genes with alternative transcripts with very long 3' UTRs (up to 15-18 kb), and a stunning mismatch in the number of male-specific genes (approximately 13% of all annotated gene models) vs. female-specific genes (less than 1%). The number of identified pseudogenes and mutations in the sequenced strain also increased significantly. We discuss remaining challenges, for instance, identification of functional small polypeptides and detection of alternative translation starts.

  19. Host gene constraints and genomic context impact the expression and evolution of human microRNAs

    PubMed Central

    França, Gustavo S.; Vibranovski, Maria D.; Galante, Pedro A. F.

    2016-01-01

    Increasing evidence has shown that recent miRNAs tend to emerge within coding genes. Here we conjecture that human miRNA evolution is tightly influenced by the genomic context, especially by host genes. Our findings show a preferential emergence of intragenic miRNAs within old genes. We found that miRNAs within old host genes are significantly more broadly expressed than those within young ones. Young miRNAs within old genes are more broadly expressed than their intergenic counterparts, suggesting that young miRNAs have an initial advantage by residing in old genes, and benefit from their hosts' expression control and from the exposure to diverse cellular contexts and target genes. Our results demonstrate that host genes may provide stronger expression constraints to intragenic miRNAs in the long run. We also report associated functional implications, highlighting the genomic context and host genes as driving factors for the expression and evolution of human miRNAs. PMID:27109497

  20. Copy Number Deletion Has Little Impact on Gene Expression Levels in Racehorses

    PubMed Central

    Park, Kyung-Do; Kim, Hyeongmin; Hwang, Jae Yeon; Lee, Chang-Kyu; Do, Kyoung-Tag; Kim, Heui-Soo; Yang, Young-Mok; Kwon, Young-jun; Kim, Jaemin; Kim, Hyeon Jeong; Song, Ki-Duk; Oh, Jae-Don; Kim, Heebal; Cho, Byung-Wook; Cho, Seoae; Lee, Hak-Kyo

    2014-01-01

    Copy number variations (CNVs), important genetic factors for study of human diseases, may have as large of an effect on phenotype as do single nucleotide polymorphisms. Indeed, it is widely accepted that CNVs are associated with differential disease susceptibility. However, the relationships between CNVs and gene expression have not been characterized in the horse. In this study, we investigated the effects of copy number deletion in the blood and muscle transcriptomes of Thoroughbred racing horses. We identified a total of 1,246 CNVs of deletion polymorphisms using DNA re-sequencing data from 18 Thoroughbred racing horses. To discover the tendencies between CNV status and gene expression levels, we extracted CNVs of four Thoroughbred racing horses of which RNA sequencing was available. We found that 252 pairs of CNVs and genes were associated in the four horse samples. We did not observe a clear and consistent relationship between the deletion status of CNVs and gene expression levels before and after exercise in blood and muscle. However, we found some pairs of CNVs and associated genes that indicated relationships with gene expression levels: a positive relationship with genes responsible for membrane structure or cytoskeleton and a negative relationship with genes involved in disease. This study will lead to conceptual advances in understanding the relationship between CNVs and global gene expression in the horse. PMID:25178379

  1. Genomic Instability of Osteosarcoma Cell Lines in Culture: Impact on the Prediction of Metastasis Relevant Genes

    PubMed Central

    Muff, Roman; Rath, Prisni; Ram Kumar, Ram Mohan; Husmann, Knut; Born, Walter; Baudis, Michael; Fuchs, Bruno

    2015-01-01

    Background Osteosarcoma is a rare but highly malignant cancer of the bone. As a consequence, the number of established cell lines used for experimental in vitro and in vivo osteosarcoma research is limited and the value of these cell lines relies on their stability during culture. Here we investigated the stability in gene expression by microarray analysis and array genomic hybridization of three low metastatic cell lines and derivatives thereof with increased metastatic potential using cells of different passages. Principal Findings The osteosarcoma cell lines showed altered gene expression during in vitro culture, and it was more pronounced in two metastatic cell lines compared to the respective parental cells. Chromosomal instability contributed in part to the altered gene expression in SAOS and LM5 cells with low and high metastatic potential. To identify metastasis-relevant genes in a background of passage-dependent altered gene expression, genes involved in "Pathways in cancer" that were consistently regulated under all passage comparisons were evaluated. Genes belonging to "Hedgehog signaling pathway" and "Wnt signaling pathway" were significantly up-regulated, and IHH, WNT10B and TCF7 were found up-regulated in all three metastatic compared to the parental cell lines. Conclusions Considerable instability during culture in terms of gene expression and chromosomal aberrations was observed in osteosarcoma cell lines. The use of cells from different passages and a search for genes consistently regulated in early and late passages allows the analysis of metastasis-relevant genes despite the observed instability in gene expression in osteosarcoma cell lines during culture. PMID:25992885

  2. Impact of methionine synthase gene and methylenetetrahydrofolate reductase gene polymorphisms on the risk of sudden sensorineural hearing loss.

    PubMed

    Gross, Menachem; Friedman, Gideon; Eliashar, Ron; Koren-Morag, Nira; Goldschmidt, Neta; Atta, Iman Abou; Ben-Yehuda, Arie

    2006-01-01

    Idiopathic sudden sensorineural hearing loss (SSNHL) represents a frequently encountered otological disease of unknown etiology. In recent years, several inherited risk factors have been found in the pathogenesis of vascular diseases. In the present study, we determined whether specific polymorphism or the combination of polymorphisms in folate-dependent homocysteine metabolism genes can act as predisposing inherited vascular risk factors in the development of SSNHL. We conducted a prospective case-control study using DNA samples extracted from 81 patients diagnosed as suffering from SSNHL and 264 healthy control subjects. Three functional polymorphisms were analyzed by polymerase chain reaction amplification, restriction enzyme digestion, and DNA fragment separation by electrophoresis: methylenetetrahydrofolate reductase (MTHFR) C677T, MTHFR A1298C, and methionine synthase (MTR) A2756G polymorphisms. The prevalence of the homozygous genotype of MTR 2756GG in the SSNHL patients (9%) was significantly higher than in the control group (4%) (p = 0.011). The allelic frequency of the G allele of the MTR A2756G polymorphism among SSNHL patients (12.5%) was also significantly higher than in the control group (5%) (p = 0.033). The prevalence of patients possessing two polymorphisms (31%) and three polymorphisms (17%) in the SSNHL group was significantly higher than in the control group (23 and 9%, respectively; p = 0.019). The frequency of patients with a very high rank risk (double homozygous) was significantly higher in the SSNHL group, MTHFR 677TT/MTR 2675GG--7%, than the frequency of patients in the control group, MTHFR 677TT/MTR 2675GG--3% (p = 0.030). Certain polymorphisms in genes encoding enzymes in the folate-dependent homocysteine metabolism are associated with SSNHL. In our case-control study, a significant association between MTR 2756GG genotype and SSNHL was found which may represent an inherited vascular risk factor in the pathogenesis of SSNHL.

  3. Aerosol from Tobacco Heating System 2.2 has reduced impact on mouse heart gene expression compared with cigarette smoke.

    PubMed

    Szostak, Justyna; Boué, Stéphanie; Talikka, Marja; Guedj, Emmanuel; Martin, Florian; Phillips, Blaine; Ivanov, Nikolai V; Peitsch, Manuel C; Hoeng, Julia

    2017-03-01

    Experimental studies clearly demonstrate a causal effect of cigarette smoking on cardiovascular disease. To reduce the individual risk and population harm caused by smoking, alternative products to cigarettes are being developed. We recently reported on an apolipoprotein E-deficient (Apoe(-/-)) mouse inhalation study that compared the effects of exposure to aerosol from a candidate modified risk tobacco product, Tobacco Heating System 2.2 (THS2.2), and smoke from the reference cigarette (3R4F) on pulmonary and vascular biology. Here, we applied a transcriptomics approach to evaluate the impact of the exposure to 3R4F smoke and THS2.2 aerosol on heart tissues from the same cohort of mice. The systems response profiles demonstrated that 3R4F smoke exposure led to time-dependent transcriptomics changes (False Discovery Rate (FDR) < 0.05; 44 differentially expressed genes at 3-months; 491 at 8-months). Analysis of differentially expressed genes in the heart tissue indicated that 3R4F exposure induced the downregulation of genes involved in cytoskeleton organization and the contractile function of the heart, notably genes that encode beta actin (Actb), actinin alpha 4 (Actn4), and filamin C (Flnc). This was accompanied by the downregulation of genes related to the inflammatory response. None of these effects were observed in the group exposed to THS2.2 aerosol.

  4. Impacts of anthropogenic activity on the ecology of class 1 integrons and integron-associated genes in the environment

    PubMed Central

    Gaze, William H; Zhang, Lihong; Abdouslam, Nouradin A; Hawkey, Peter M; Calvo-Bado, Leo; Royle, Jeremy; Brown, Helen; Davis, Susan; Kay, Paul; Boxall, Alistair B A; Wellington, Elizabeth M H

    2011-01-01

    The impact of human activity on the selection for antibiotic resistance in the environment is largely unknown, although considerable amounts of antibiotics are introduced through domestic wastewater and farm animal waste. Selection for resistance may occur by exposure to antibiotic residues or by co-selection for mobile genetic elements (MGEs) which carry genes of varying activity. Class 1 integrons are genetic elements that carry antibiotic and quaternary ammonium compound (QAC) resistance genes that confer resistance to detergents and biocides. This study aimed to investigate the prevalence and diversity of class 1 integron and integron-associated QAC resistance genes in bacteria associated with industrial waste, sewage sludge and pig slurry. We show that prevalence of class 1 integrons is higher in bacteria exposed to detergents and/or antibiotic residues, specifically in sewage sludge and pig slurry compared with agricultural soils to which these waste products are amended. We also show that QAC resistance genes are more prevalent in the presence of detergents. Studies of class 1 integron prevalence in sewage sludge amended soil showed measurable differences compared with controls. Insertion sequence elements were discovered in integrons from QAC contaminated sediment, acting as powerful promoters likely to upregulate cassette gene expression. On the basis of this data, >1 × 1019 bacteria carrying class 1 integrons enter the United Kingdom environment by disposal of sewage sludge each year. PMID:21368907

  5. Prenatal exposure to arsenic and cadmium impacts infectious disease-related genes within the glucocorticoid receptor signal transduction pathway.

    PubMed

    Rager, Julia E; Yosim, Andrew; Fry, Rebecca C

    2014-12-03

    There is increasing evidence that environmental agents mediate susceptibility to infectious disease. Studies support the impact of prenatal/early life exposure to the environmental metals inorganic arsenic (iAs) and cadmium (Cd) on increased risk for susceptibility to infection. The specific biological mechanisms that underlie such exposure-mediated effects remain understudied. This research aimed to identify key genes/signal transduction pathways that associate prenatal exposure to these toxic metals with changes in infectious disease susceptibility using a Comparative Genomic Enrichment Method (CGEM). Using CGEM an infectious disease gene (IDG) database was developed comprising 1085 genes with known roles in viral, bacterial, and parasitic disease pathways. Subsequently, datasets collected from human pregnancy cohorts exposed to iAs or Cd were examined in relationship to the IDGs, specifically focusing on data representing epigenetic modifications (5-methyl cytosine), genomic perturbations (mRNA expression), and proteomic shifts (protein expression). A set of 82 infection and exposure-related genes was identified and found to be enriched for their role in the glucocorticoid receptor signal transduction pathway. Given their common identification across numerous human cohorts and their known toxicological role in disease, the identified genes within the glucocorticoid signal transduction pathway may underlie altered infectious disease susceptibility associated with prenatal exposures to the toxic metals iAs and Cd in humans.

  6. Prenatal Exposure to Arsenic and Cadmium Impacts Infectious Disease-Related Genes within the Glucocorticoid Receptor Signal Transduction Pathway

    PubMed Central

    Rager, Julia E.; Yosim, Andrew; Fry, Rebecca C.

    2014-01-01

    There is increasing evidence that environmental agents mediate susceptibility to infectious disease. Studies support the impact of prenatal/early life exposure to the environmental metals inorganic arsenic (iAs) and cadmium (Cd) on increased risk for susceptibility to infection. The specific biological mechanisms that underlie such exposure-mediated effects remain understudied. This research aimed to identify key genes/signal transduction pathways that associate prenatal exposure to these toxic metals with changes in infectious disease susceptibility using a Comparative Genomic Enrichment Method (CGEM). Using CGEM an infectious disease gene (IDG) database was developed comprising 1085 genes with known roles in viral, bacterial, and parasitic disease pathways. Subsequently, datasets collected from human pregnancy cohorts exposed to iAs or Cd were examined in relationship to the IDGs, specifically focusing on data representing epigenetic modifications (5-methyl cytosine), genomic perturbations (mRNA expression), and proteomic shifts (protein expression). A set of 82 infection and exposure-related genes was identified and found to be enriched for their role in the glucocorticoid receptor signal transduction pathway. Given their common identification across numerous human cohorts and their known toxicological role in disease, the identified genes within the glucocorticoid signal transduction pathway may underlie altered infectious disease susceptibility associated with prenatal exposures to the toxic metals iAs and Cd in humans. PMID:25479081

  7. geneLAB: Expanding the Impact of NASA's Biological Research in Space

    NASA Technical Reports Server (NTRS)

    Rayl, Nicole; Smith, Jeffrey D.

    2014-01-01

    The geneLAB project is designed to leverage the value of large 'omics' datasets from molecular biology projects conducted on the ISS by making these datasets available, citable, discoverable, interpretable, reusable, and reproducible. geneLAB will create a collaboration space with an integrated set of tools for depositing, accessing, analyzing, and modeling these diverse datasets from spaceflight and related terrestrial studies.

  8. Transposable elements: insertion pattern and impact on gene expression evolution in hominids.

    PubMed

    Warnefors, Maria; Pereira, Vini; Eyre-Walker, Adam

    2010-08-01

    Transposable elements (TEs) can affect the regulation of nearby genes through several mechanisms. Here, we examine to what extent recent TE insertions have contributed to the evolution of gene expression in hominids. We compare expression levels of human and chimpanzee orthologs and detect a weak increase in expression divergence (ED) for genes with species-specific TE insertions compared with unaffected genes. However, we show that genes with TE insertions predating the human-chimpanzee split also exhibit a similar increase in ED and therefore conclude that the increase is not due to the transcriptional influence of the TEs. These results are further confirmed by lineage-specific analysis of ED, using rhesus macaque as an outgroup: Human-chimpanzee ortholog pairs, where one ortholog has suffered TE insertion but not the other, do not show increased ED along the lineage where the insertion occurred, relative to the other lineage. We also show that genes with recent TE insertions tend to produce more alternative transcripts but find no evidence that the TEs themselves promote transcript diversity. Finally, we observe that TEs are enriched upstream relative to downstream of genes and show that this is due to insertional bias, rather than selection, because this bias is only observed in genes expressed in the germ line. This provides an alternative neutral explanation for the accumulation of TEs in upstream sequences.

  9. Impact of Cigarette Smoke on the Human and Mouse Lungs: A Gene-Expression Comparison Study

    PubMed Central

    Morissette, Mathieu C.; Lamontagne, Maxime; Bérubé, Jean-Christophe; Gaschler, Gordon; Williams, Andrew; Yauk, Carole; Couture, Christian; Laviolette, Michel; Hogg, James C.; Timens, Wim; Halappanavar, Sabina; Stampfli, Martin R.; Bossé, Yohan

    2014-01-01

    Cigarette smoke is well known for its adverse effects on human health, especially on the lungs. Basic research is essential to identify the mechanisms involved in the development of cigarette smoke-related diseases, but translation of new findings from pre-clinical models to the clinic remains difficult. In the present study, we aimed at comparing the gene expression signature between the lungs of human smokers and mice exposed to cigarette smoke to identify the similarities and differences. Using human and mouse whole-genome gene expression arrays, changes in gene expression, signaling pathways and biological functions were assessed. We found that genes significantly modulated by cigarette smoke in humans were enriched for genes modulated by cigarette smoke in mice, suggesting a similar response of both species. Sixteen smoking-induced genes were in common between humans and mice including six newly reported to be modulated by cigarette smoke. In addition, we identified a new conserved pulmonary response to cigarette smoke in the induction of phospholipid metabolism/degradation pathways. Finally, the majority of biological functions modulated by cigarette smoke in humans were also affected in mice. Altogether, the present study provides information on similarities and differences in lung gene expression response to cigarette smoke that exist between human and mouse. Our results foster the idea that animal models should be used to study the involvement of pathways rather than single genes in human diseases. PMID:24663285

  10. UPDATE ON SLA GENES AND THEIR IMPACT ON IMMUNE AND DISEASE INTERACTIONS

    USDA-ARS?s Scientific Manuscript database

    The swine major histocompatibility complex (MHC) or swine leukocyte antigen (SLA) complex is one of the most gene-dense regions in the swine genome. It consists of three major gene clusters, the SLA class I, class III and class II regions, that span ~1.1, 0.7 and 0.5 Mb, respectively, making the swi...

  11. Impact of C-Myc gene-related aberrations in newly diagnosed myeloma with bortezomib/dexamethasone therapy.

    PubMed

    Sekiguchi, Naohiro; Ootsubo, Kaori; Wagatsuma, Miyuki; Midorikawa, Kiyoe; Nagata, Akihisa; Noto, Satoshi; Yamada, Kazuaki; Takezako, Naoki

    2014-03-01

    Recent studies have suggested that c-Myc over-expression may be a factor indicating poor prognosis in multiple myeloma (MM), although c-Myc gene-related abnormalities, including translocation and gene amplification, have not been fully investigated in the novel agent era. Additional chromosome 8 may be considered as aggressive disease in the 1990s. To clarify the impact of these aberrations, we retrospectively analyzed newly diagnosed MM (NDMM) and relapsed/refractory MM (RRMM) with bortezomib and dexamethasone induction therapy. In the present study, the high-risk group was defined as having at least one of the following present: non-hyperdiploidy, IgH/FGFR3, and del p53. Forty NDMM cases were analyzed. At the median follow-up duration of 14.1 months, 14 RRMM were recognized. The proportions of patients in the high-risk, c-Myc gene-related aberrations, and additional chromosome 8 groups at diagnosis were 45.5, 22.5, and 10 %, respectively. The proportions of patients who developed RRMM in the high-risk, c-Myc gene-related aberrations, and additional chromosome 8 groups were 41.7, 77.7, and 50 %, respectively. Furthermore, patients with c-Myc gene-related abnormalities tended to exhibit inferior progression-free survival (PFS), and those with c-Myc gene-related abnormalities and/or additional chromosome 8 showed statistically shorter PFS. Therefore, c-Myc gene-related abnormalities and additional chromosome 8 may be related to a poorer prognosis.

  12. Gene-diet interactions and their impact on colorectal cancer risk

    PubMed Central

    Kantor, Elizabeth D.; Giovannucci, Edward L.

    2015-01-01

    A number of studies have evaluated the role of gene-diet interaction in the etiology of colorectal cancer (CRC). Historically, these studies focused on established dietary risk factors and genes involved in their metabolism. However, results from these candidate gene studies were inconsistent, possibly due to multiple testing and publication bias. In recent years, genome-wide association studies have identified a number of CRC susceptibility loci, and subsequent meta-analyses have observed limited evidence that diet may modify the risk associated with these susceptibility loci. Statistical techniques have been recently developed to evaluate the presence of interaction across the entire genome; results from these genome-wide studies have demonstrated limited evidence of interaction and have failed to replicate results from candidate gene studies and those using established susceptibility loci. However, larger sample sizes are likely needed to elucidate modest or weak interaction in genome-wide studies of gene-diet interaction. PMID:25844273

  13. Impact of Adenovirus E4-ORF3 Oligomerization and Protein Localization on Cellular Gene Expression.

    PubMed

    Vink, Elizabeth I; Zheng, Yueting; Yeasmin, Rukhsana; Stamminger, Thomas; Krug, Laurie T; Hearing, Patrick

    2015-05-13

    The Adenovirus E4-ORF3 protein facilitates virus replication through the relocalization of cellular proteins into nuclear inclusions termed tracks. This sequestration event disrupts antiviral properties associated with target proteins. Relocalization of Mre11-Rad50-Nbs1 proteins prevents the DNA damage response from inhibiting Ad replication. Relocalization of PML and Daxx impedes the interferon-mediated antiviral response. Several E4-ORF3 targets regulate gene expression, linking E4-ORF3 to transcriptional control. Furthermore, E4-ORF3 was shown to promote the formation of heterochromatin, down-regulating p53-dependent gene expression. Here, we characterize how E4-ORF3 alters cellular gene expression. Using an inducible, E4-ORF3-expressing cell line, we performed microarray experiments to highlight cellular gene expression changes influenced by E4-ORF3 expression, identifying over four hundred target genes. Enrichment analysis of these genes suggests that E4-ORF3 influences factors involved in signal transduction and cellular defense, among others. The expression of mutant E4-ORF3 proteins revealed that nuclear track formation is necessary to induce these expression changes. Through the generation of knockdown cells, we demonstrate that the observed expression changes may be independent of Daxx and TRIM33 suggesting that an additional factor(s) may be responsible. The ability of E4-ORF3 to manipulate cellular gene expression through the sequestration of cellular proteins implicates a novel role for E4-ORF3 in transcriptional regulation.

  14. Impact of microRNA regulation on variation in human gene expression

    PubMed Central

    Lu, Jian; Clark, Andrew G.

    2012-01-01

    MicroRNAs (miRNAs) are endogenously expressed small RNAs that regulate expression of mRNAs at the post-transcriptional level. The consequence of miRNA regulation is hypothesized to reduce the expression variation of target genes. However, it is possible that mutations in miRNAs and target sites cause rewiring of the miRNA regulatory networks resulting in increased variation in gene expression. By examining variation in gene expression patterns in human populations and between human and other primate species, we find that miRNAs have stabilized expression of a small number of target genes during primate evolution. Compared with genes not regulated by miRNAs, however, genes regulated by miRNAs overall have higher expression variation at the population level, and they display greater variation in expression among human ethnic groups or between human and other primate species. By integrating expression data with genotypes determined in the HapMap 3 and the 1000 Genomes Projects, we found that expression variation in miRNAs, genetic variants in miRNA loci, and mutations in miRNA target sites are important sources of elevated expression variation of miRNA target genes. A reasonable case can be made that natural selection is driving this pattern of variation. PMID:22456605

  15. Impact of Adenovirus E4-ORF3 Oligomerization and Protein Localization on Cellular Gene Expression

    PubMed Central

    Vink, Elizabeth I.; Zheng, Yueting; Yeasmin, Rukhsana; Stamminger, Thomas; Krug, Laurie T.; Hearing, Patrick

    2015-01-01

    The Adenovirus E4-ORF3 protein facilitates virus replication through the relocalization of cellular proteins into nuclear inclusions termed tracks. This sequestration event disrupts antiviral properties associated with target proteins. Relocalization of Mre11-Rad50-Nbs1 proteins prevents the DNA damage response from inhibiting Ad replication. Relocalization of PML and Daxx impedes the interferon-mediated antiviral response. Several E4-ORF3 targets regulate gene expression, linking E4-ORF3 to transcriptional control. Furthermore, E4-ORF3 was shown to promote the formation of heterochromatin, down-regulating p53-dependent gene expression. Here, we characterize how E4-ORF3 alters cellular gene expression. Using an inducible, E4-ORF3-expressing cell line, we performed microarray experiments to highlight cellular gene expression changes influenced by E4-ORF3 expression, identifying over four hundred target genes. Enrichment analysis of these genes suggests that E4-ORF3 influences factors involved in signal transduction and cellular defense, among others. The expression of mutant E4-ORF3 proteins revealed that nuclear track formation is necessary to induce these expression changes. Through the generation of knockdown cells, we demonstrate that the observed expression changes may be independent of Daxx and TRIM33 suggesting that an additional factor(s) may be responsible. The ability of E4-ORF3 to manipulate cellular gene expression through the sequestration of cellular proteins implicates a novel role for E4-ORF3 in transcriptional regulation. PMID:25984715

  16. Gene x environment interactions impact endometrial function and the menstrual cycle: PROGINS, life history, anthropometry, and physical activity.

    PubMed

    Rowe, Elizabeth J; Eisenstein, Toby K; Meissler, Joseph; Rockwell, L Christie

    2013-01-01

    We assessed the impact of a high frequency, functionally significant allelic variant of the progesterone receptor gene (PROGINS) on endometrial function and menstrual cycle characteristics. Further we asked whether PROGINS moderates the impact of life history characteristics, anthropometric measures, and physical activity on endometrial function. Fifty-two women were genotyped for the PROGINS variant, provided life history information, and had anthropometric measurements made. Women monitored their menstrual bleeding for three cycles, performed mid-cycle urinary ovulation tests, and recorded physical activity. A subset of women provided daily saliva samples and had mid-luteal endometrial thickness measurements taken during the third menstrual cycle. Salivary progesterone was assayed using ELISAs. The direct impact of PROGINS on endometrial and menstrual cycle characteristics was determined via independent t-tests with Bonferroni correction. Interactions between PROGINS and covariates were assessed by moderated regression. PROGINS did not directly impact any indicator of endometrial function. However, PROGINS caused an increase in menstrual cycle length with increasing mid-luteal progesterone levels; the opposite relationship was present in noncarriers (P < 0.05). Additionally, PROGINS interacted with four of six anthropometric measures (BMI, waist circumference, height, and waist-hip ratio) to impact endometrial function, however, interactions between PROGINS and life history variables, or physical activity was limited. The gene x environment interactions we report suggest that PROGINS alters endometrial sensitivity to maternal energetic condition. Thus, the possibility of genetically-based variation in sensitivity to energetic stress should be considered in future adaptive models of women's reproduction. Copyright © 2013 Wiley Periodicals, Inc.

  17. Targeted gene correction minimally impacts whole-genome mutational load in human-disease-specific induced pluripotent stem cell clones.

    PubMed

    Suzuki, Keiichiro; Yu, Chang; Qu, Jing; Li, Mo; Yao, Xiaotian; Yuan, Tingting; Goebl, April; Tang, Senwei; Ren, Ruotong; Aizawa, Emi; Zhang, Fan; Xu, Xiuling; Soligalla, Rupa Devi; Chen, Feng; Kim, Jessica; Kim, Na Young; Liao, Hsin-Kai; Benner, Chris; Esteban, Concepcion Rodriguez; Jin, Yabin; Liu, Guang-Hui; Li, Yingrui; Izpisua Belmonte, Juan Carlos

    2014-07-03

    The utility of genome editing technologies for disease modeling and developing cellular therapies has been extensively documented, but the impact of these technologies on mutational load at the whole-genome level remains unclear. We performed whole-genome sequencing to evaluate the mutational load at single-base resolution in individual gene-corrected human induced pluripotent stem cell (hiPSC) clones in three different disease models. In single-cell clones, gene correction by helper-dependent adenoviral vector (HDAdV) or Transcription Activator-Like Effector Nuclease (TALEN) exhibited few off-target effects and a low level of sequence variation, comparable to that accumulated in routine hiPSC culture. The sequence variants were randomly distributed and unique to individual clones. We also combined both technologies and developed a TALEN-HDAdV hybrid vector, which significantly increased gene-correction efficiency in hiPSCs. Therefore, with careful monitoring via whole-genome sequencing it is possible to apply genome editing to human pluripotent cells with minimal impact on genomic mutational load.

  18. Impacts of ocean acidification on gene expression and biomineralisation in the Pacific oyster Crassostrea gigas Thunberg, 1793

    NASA Astrophysics Data System (ADS)

    Bagusche, F.; Pouvreau, S.; Trueman, C.; Long, S.; Hauton, C.

    2012-04-01

    The published evidence of impacts of ocean acidification and on marine calcifiers has emphasized the need to understand the molecular mechanisms of biomineralisation. Crassostrea gigas is an ideal organism to examine these processes as: 1) the hatchery rearing of larval stages is well constrained, 2) studies have established an ontogenetic switch in deposition of carbonate polymorphs from aragonite in larval shells to calcite in adults and 3) it is a globally-important commercial species. Research summarized in this presentation will identify some of the molecular mechanisms involved in calcification processes during ontogeny of Crassostrea gigas, as well as possible impacts of changes in environmental conditions such as temperature and pH. Data will be presented from a quantitative real-time PCR study of the changes in gene expression during development in different environments. Additionally scanning electron microscopy and infrared spectroscopy analyses of shell microstructures and composition will be summarised to correlate changes in gene expression with end-point differences in shell structure. Preliminary results suggest that changes in the environmental conditions lead to differences in expression patterns of genes involved in biomineralisation processes. The combined effects of ambient seawater temperature and low pH show the greatest negative effect on larval shell development, identified as malformations, eroded shell surfaces and a significant decrease in shell size. However, the effect of higher seawater temperature seems to amend the effects of ocean acidification on larval shell development.

  19. Impact of pre-existing immunity on gene transfer to nonhuman primate liver with adeno-associated virus 8 vectors.

    PubMed

    Wang, Lili; Calcedo, Roberto; Bell, Peter; Lin, Jianping; Grant, Rebecca L; Siegel, Don L; Wilson, James M

    2011-11-01

    Vectors based on the primate-derived adeno-associated virus serotype 8 (AAV8) are being evaluated in preclinical and clinical models. Natural infections with related AAVs activate memory B cells that produce antibodies capable of modulating the efficacy and safety of the vector. We have evaluated the biology of AAV8 gene transfer in macaque liver, with a focus on assessing the impact of pre-existing humoral immunity. Twenty-one macaques with various levels of AAV neutralizing antibody (NAb) were injected intravenously with AAV8 vector expressing green fluorescent protein. Pre-existing antibody titers in excess of 1:10 substantially diminished hepatocyte transduction that, in the absence of NAbs, was highly efficient. Vector-specific NAb diminished liver deposition of genomes and unexpectedly increased genome distribution to the spleen. The majority of animals showed high-level and stable sequestration of vector capsid protein by follicular dendritic cells of splenic germinal centers. These studies illustrate how natural immunity to a virus that is related to a vector can impact the efficacy and potential safety of in vivo gene therapy. We propose to use the in vitro transduction inhibition assay to evaluate research subjects before gene therapy and to preclude from systemic AAV8 trials those that have titers in excess of 1:10.

  20. Upon accounting for the impact of isoenzyme loss, gene deletion costs anticorrelate with their evolutionary rates

    DOE PAGES

    Jacobs, Christopher; Lambourne, Luke; Xia, Yu; ...

    2017-01-20

    Here, system-level metabolic network models enable the computation of growth and metabolic phenotypes from an organism's genome. In particular, flux balance approaches have been used to estimate the contribution of individual metabolic genes to organismal fitness, offering the opportunity to test whether such contributions carry information about the evolutionary pressure on the corresponding genes. Previous failure to identify the expected negative correlation between such computed gene-loss cost and sequence-derived evolutionary rates in Saccharomyces cerevisiae has been ascribed to a real biological gap between a gene's fitness contribution to an organism "here and now"º and the same gene's historical importance asmore » evidenced by its accumulated mutations over millions of years of evolution. Here we show that this negative correlation does exist, and can be exposed by revisiting a broadly employed assumption of flux balance models. In particular, we introduce a new metric that we call "function-loss cost", which estimates the cost of a gene loss event as the total potential functional impairment caused by that loss. This new metric displays significant negative correlation with evolutionary rate, across several thousand minimal environments. We demonstrate that the improvement gained using function-loss cost over gene-loss cost is explained by replacing the base assumption that isoenzymes provide unlimited capacity for backup with the assumption that isoenzymes are completely non-redundant. We further show that this change of the assumption regarding isoenzymes increases the recall of epistatic interactions predicted by the flux balance model at the cost of a reduction in the precision of the predictions. In addition to suggesting that the gene-to-reaction mapping in genome-scale flux balance models should be used with caution, our analysis provides new evidence that evolutionary gene importance captures much more than strict essentiality.« less

  1. Upon Accounting for the Impact of Isoenzyme Loss, Gene Deletion Costs Anticorrelate with Their Evolutionary Rates.

    PubMed

    Jacobs, Christopher; Lambourne, Luke; Xia, Yu; Segrè, Daniel

    2017-01-01

    System-level metabolic network models enable the computation of growth and metabolic phenotypes from an organism's genome. In particular, flux balance approaches have been used to estimate the contribution of individual metabolic genes to organismal fitness, offering the opportunity to test whether such contributions carry information about the evolutionary pressure on the corresponding genes. Previous failure to identify the expected negative correlation between such computed gene-loss cost and sequence-derived evolutionary rates in Saccharomyces cerevisiae has been ascribed to a real biological gap between a gene's fitness contribution to an organism "here and now" and the same gene's historical importance as evidenced by its accumulated mutations over millions of years of evolution. Here we show that this negative correlation does exist, and can be exposed by revisiting a broadly employed assumption of flux balance models. In particular, we introduce a new metric that we call "function-loss cost", which estimates the cost of a gene loss event as the total potential functional impairment caused by that loss. This new metric displays significant negative correlation with evolutionary rate, across several thousand minimal environments. We demonstrate that the improvement gained using function-loss cost over gene-loss cost is explained by replacing the base assumption that isoenzymes provide unlimited capacity for backup with the assumption that isoenzymes are completely non-redundant. We further show that this change of the assumption regarding isoenzymes increases the recall of epistatic interactions predicted by the flux balance model at the cost of a reduction in the precision of the predictions. In addition to suggesting that the gene-to-reaction mapping in genome-scale flux balance models should be used with caution, our analysis provides new evidence that evolutionary gene importance captures much more than strict essentiality.

  2. The impact of PPARα activation on whole genome gene expression in human precision cut liver slices.

    PubMed

    Janssen, Aafke W F; Betzel, Bark; Stoopen, Geert; Berends, Frits J; Janssen, Ignace M; Peijnenburg, Ad A; Kersten, Sander

    2015-10-08

    Studies in mice have shown that PPARα is an important regulator of lipid metabolism in liver and key transcription factor involved in the adaptive response to fasting. However, much less is known about the role of PPARα in human liver. Here we set out to study the function of PPARα in human liver via analysis of whole genome gene regulation in human liver slices treated with the PPARα agonist Wy14643. Quantitative PCR indicated that PPARα is well expressed in human liver and human liver slices and that the classical PPARα targets PLIN2, VLDLR, ANGPTL4, CPT1A and PDK4 are robustly induced by PPARα activation. Transcriptomics analysis indicated that 617 genes were upregulated and 665 genes were downregulated by PPARα activation (q value < 0.05). Many genes induced by PPARα activation were involved in lipid metabolism (ACSL5, AGPAT9, FADS1, SLC27A4), xenobiotic metabolism (POR, ABCC2, CYP3A5) or the unfolded protein response, whereas most of the downregulated genes were involved in immune-related pathways. Among the most highly repressed genes upon PPARα activation were several chemokines (e.g. CXCL9-11, CCL8, CX3CL1, CXCL6), interferon γ-induced genes (e.g. IFITM1, IFIT1, IFIT2, IFIT3) and numerous other immune-related genes (e.g. TLR3, NOS2, and LCN2). Comparative analysis of gene regulation by Wy14643 between human liver slices and primary human hepatocytes showed that down-regulation of gene expression by PPARα is much better captured by liver slices as compared to primary hepatocytes. In particular, PPARα activation markedly suppressed immunity/inflammation-related genes in human liver slices but not in primary hepatocytes. Finally, several putative new target genes of PPARα were identified that were commonly induced by PPARα activation in the two human liver model systems, including TSKU, RHOF, CA12 and VSIG10L. Our paper demonstrates the suitability and superiority of human liver slices over primary hepatocytes for studying the functional

  3. Annuity payments can increase patient access to innovative cell and gene therapies under England's net budget impact test.

    PubMed

    Jørgensen, Jesper; Kefalas, Panos

    2017-01-01

    Background: Cell and gene therapies have the potential to provide therapeutic breakthroughs, but the high costs of researching, developing, manufacturing and delivering them translate into prices that may challenge healthcare budgets. Various measures exist that aim to address the affordability challenge, including reducing price, limiting patient numbers and/or linking remuneration to product performance. Objective: To explore how the net budget impact test recently introduced in England can affect patient access to high-value, one-off cell and gene therapies, and how managed entry agreements can improve access. Methods: We use a hypothetical example where a new high-value, one-off therapy launches in an indication where it displaces a relatively low cost chronic treatment. We calculate the number of patients that can be treated without exceeding the £20 million net budget impact threshold, and compare results for scenarios where a full upfront payment is used, and where annuity-based payments are used. Results: Charging a full upfront payment at the time of treatment can lead to suboptimal patient access. Conclusion: Annuity-based payments in combination with an outcomes-based remuneration scheme reduce consequences of decision uncertainty and can increase patient access, without exceeding the net budget impact test.

  4. Annuity payments can increase patient access to innovative cell and gene therapies under England’s net budget impact test

    PubMed Central

    Jørgensen, Jesper; Kefalas, Panos

    2017-01-01

    ABSTRACT Background: Cell and gene therapies have the potential to provide therapeutic breakthroughs, but the high costs of researching, developing, manufacturing and delivering them translate into prices that may challenge healthcare budgets. Various measures exist that aim to address the affordability challenge, including reducing price, limiting patient numbers and/or linking remuneration to product performance. Objective: To explore how the net budget impact test recently introduced in England can affect patient access to high-value, one-off cell and gene therapies, and how managed entry agreements can improve access. Methods: We use a hypothetical example where a new high-value, one-off therapy launches in an indication where it displaces a relatively low cost chronic treatment. We calculate the number of patients that can be treated without exceeding the £20 million net budget impact threshold, and compare results for scenarios where a full upfront payment is used, and where annuity-based payments are used. Results: Charging a full upfront payment at the time of treatment can lead to suboptimal patient access. Conclusion: Annuity-based payments in combination with an outcomes-based remuneration scheme reduce consequences of decision uncertainty and can increase patient access, without exceeding the net budget impact test. PMID:28839525

  5. Genes

    MedlinePlus

    ... Search Search MedlinePlus GO GO About MedlinePlus Site Map FAQs Customer Support Health Topics Drugs & Supplements Videos & Tools Español You Are Here: Home → Medical Encyclopedia → Genes URL of this page: //medlineplus.gov/ency/article/ ...

  6. Aggregation of AcMNPV LEF-10 and Its Impact on Viral Late Gene Expression

    PubMed Central

    Xu, Xiaodong; Zhou, Xinyu; Nan, Hao; Zhao, Yu; Bai, Yu; Ou, Yanmei; Chen, Hongying

    2016-01-01

    The Autographa californica multiple nucleopolyhedrovirus (AcMNPV) late expression factor gene lef-10 has been identified to be required for viral late gene expression by transient expression assay. Our previous work has shown that the gene product LEF-10 can form very stable high-molecular-weight complexes, but the structure and function of the protein remain unknown. In this study, we demonstrated that LEF-10 was essential for the replication of AcMNPV, and its truncated fragment containing amino acid residues 1 to 48 were sufficient to support the virus survival. Our data also suggested that the LEF-10 could spontaneously aggregate to form punctate spots in virus infected Sf9 cells at low frequency, and the aggregation of the protein could be induced by LEF-10 over-expression. When the protein aggregated to form punctate spots, soluble LEF-10 proteins were depleted and this could result in the down-regulation of viral late gene expression. PMID:27152613

  7. The impact of gene-environment interaction on alcohol use disorders.

    PubMed

    Dick, Danielle M; Kendler, Kenneth S

    2012-01-01

    This article describes three types of gene-environment interactions and the challenges inherent in interpreting these interactions. It also reports on what is known about gene-environment interactions in the field of alcohol use disorders (AUDs). Twin studies of the interaction of genetic and environmental influences on AUDs have resulted in relatively consistent findings and have suggested general mechanisms for interaction effects. These studies generally find that environments that exert more social control (e.g., higher parental monitoring, less migratory neighborhoods, etc.) tend to reduce genetic influences, whereas other environments allow greater opportunity to express genetic predispositions, such as those characterized by more deviant peers and greater alcohol availability. Conversely, the gene-environment literature that has been developed surrounding specific genes has focused largely on the role of stress as a moderator of genetic effects.

  8. Splicing factor gene mutations in the myelodysplastic syndromes: impact on disease phenotype and therapeutic applications.

    PubMed

    Pellagatti, Andrea; Boultwood, Jacqueline

    2017-01-01

    Splicing factor gene mutations are the most frequent mutations found in patients with the myeloid malignancy myelodysplastic syndrome (MDS), suggesting that spliceosomal dysfunction plays a major role in disease pathogenesis. The aberrantly spliced target genes and deregulated cellular pathways associated with the commonly mutated splicing factor genes in MDS (SF3B1, SRSF2 and U2AF1) are being identified, illuminating the molecular mechanisms underlying MDS. Emerging data from mouse modeling studies indicate that the presence of splicing factor gene mutations can lead to bone marrow hematopoietic stem/myeloid progenitor cell expansion, impaired hematopoiesis and dysplastic differentiation that are hallmarks of MDS. Importantly, recent evidence suggests that spliceosome inhibitors and splicing modulators may have therapeutic value in the treatment of splicing factor mutant myeloid malignancies.

  9. Complex MHC class I gene transcription profiles and their functional impact in orangutans

    PubMed Central

    de Groot, Natasja G.; Heijmans, Corrine M.C.; van der Wiel, Marit K.H.; Blokhuis, Jeroen H.; Mulder, Arend; Guethlein, Lisbeth A.; Doxiadis, Gaby G.M.; Claas, Frans H.J.; Parham, Peter; Bontrop, Ronald E.

    2015-01-01

    MHC haplotypes of humans and the African great ape species have one copy of the MHC-A, -B, and -C genes. In contrast, MHC haplotypes of orangutans, the Asian great ape species, exhibit variation in the number of gene copies. An in-depth analysis of the MHC class I gene repertoire in the two orangutan species, Pongo abelii and Pongo pygmaeus, is presented here. This analysis involved Sanger and next-generation sequencing methodologies, revealing diverse and complicated transcription profiles for orangutan MHC-A, -B, and -C. Thirty-five previously unreported MHC class I alleles are described. The data demonstrate that each orangutan MHC haplotype has one copy of the MHC-A gene, and that the MHC-B region has been subject to duplication, giving rise to at least three MHC-B genes. The MHC-B*03 and -B*08 lineages of alleles each account for a separate MHC-B gene. All MHC-B*08 allotypes have the C1-epitope motif recognized by KIR. At least one other MHC-B gene is present, pointing to MHC-B alleles that are not B*03 or B*08. The MHC-C gene is present only on some haplotypes, and each MHC-C allotype has the C1-epitope. The transcription profiles demonstrate that MHC-A alleles are highly transcribed, whereas MHC-C alleles, when present, are transcribed at very low levels. The MHC-B alleles are transcribed to a variable extent and over a wide range. For those orangutan MHC class I allotypes that are detected by human monoclonal anti-HLA class I antibodies, the level of cell-surface expression of proteins correlates with the level of transcription of the allele. PMID:26685209

  10. Impact of smoking cessation on global gene expression in the bronchial epithelium of chronic smokers

    PubMed Central

    Zhang, Li; Lee, Jack; Tang, Hongli; Fan, You-Hong; Xiao, Lianchun; Ren, Hening; Kurie, Jonathan; Morice, Rodolfo C; Hong, Waun Ki; Mao, Li

    2014-01-01

    Cigarette smoke is the major cause of lung cancer and can interact in complex ways with drugs for lung cancer prevention or therapy. Molecular genetic research promises to elucidate the biologic mechanisms underlying divergent drug effects in smokers versus non-smokers and to help in developing new approaches for controlling lung cancer. The present study compared global gene expression profiles (determined via Affymetrix microarray measurements in bronchial epithelial cells) between chronic smokers, former smokers, and never smokers. Smoking effects on global gene expression were determined from a combined analysis of three independent datasets. Differential expression between current and never smokers occurred in 591 of the 13,902 genes measured on the microarrays (P < 0.01 and >2 fold change; pooled data)—a profound effect. In contrast, differential expression between current and former smokers occurred in only 145 of the measured genes (P < 0.01 and >2 fold change; pooled data). Nine of these 145 genes showed consistent and significant changes in each of the three datasets (P < 0.01 and >2 fold change), with 8 being down-regulated in former smokers. Seven of the 8 down-regulated genes, including CYP1B1 and 3 AKR genes, influence the metabolism of carcinogens and/or therapeutic/chemopreventive agents. Our data comparing former and current smokers allowed us to pinpoint the genes involved in smoking–drug interactions in lung cancer prevention and therapy. These findings have important implications for developing new targeted and dosing approaches for prevention and therapy in the lung and other sites, highlighting the importance of monitoring smoking status in patients receiving oncologic drug interventions. PMID:19138944

  11. The Genomic Impact of DNA CpG Methylation on Gene Expression; Relationships in Prostate Cancer.

    PubMed

    Long, Mark D; Smiraglia, Dominic J; Campbell, Moray J

    2017-02-14

    The process of DNA CpG methylation has been extensively investigated for over 50 years and revealed associations between changing methylation status of CpG islands and gene expression. As a result, DNA CpG methylation is implicated in the control of gene expression in developmental and homeostasis processes, as well as being a cancer-driver mechanism. The development of genome-wide technologies and sophisticated statistical analytical approaches has ushered in an era of widespread analyses, for example in the cancer arena, of the relationships between altered DNA CpG methylation, gene expression, and tumor status. The remarkable increase in the volume of such genomic data, for example, through investigators from the Cancer Genome Atlas (TCGA), has allowed dissection of the relationships between DNA CpG methylation density and distribution, gene expression, and tumor outcome. In this manner, it is now possible to test that the genome-wide correlations are measurable between changes in DNA CpG methylation and gene expression. Perhaps surprisingly is that these associations can only be detected for hundreds, but not thousands, of genes, and the direction of the correlations are both positive and negative. This, perhaps, suggests that CpG methylation events in cancer systems can act as disease drivers but the effects are possibly more restricted than suspected. Additionally, the positive and negative correlations suggest direct and indirect events and an incomplete understanding. Within the prostate cancer TCGA cohort, we examined the relationships between expression of genes that control DNA methylation, known targets of DNA methylation and tumor status. This revealed that genes that control the synthesis of S-adenosyl-l-methionine (SAM) associate with altered expression of DNA methylation targets in a subset of aggressive tumors.

  12. Impact of thawing on reference gene expression stability in renal cell carcinoma samples.

    PubMed

    Ma, Yi; Dai, HuiLi; Kong, XianMing; Wang, LiMin

    2012-09-01

    More and more samples are obtained from biobanks for biomedical research; however, some of these samples may undergo thawing before processing. We aim to evaluate the reference gene expression stability in thawed renal cell carcinoma samples. Sixteen matched malignant and nonmalignant renal tissue samples were obtained and each sample was divided into 4 aliquots before being snap frozen and stored at -80°C. By quantitative real-time polymerase chain reaction, a time-course study was conducted on the thawed tissue to evaluate the expression stability of a panel of the 10 most frequently used reference genes in renal cell carcinom samples: ACTB, ALAS1, B2M, GAPDH, HMBS, HPRT, PPIA, RPLP0,TBP, and TUBB. As shown by geNorm M values, PPIA was the most stable gene at the 0-, 15-, and 30-minute time points (M=0.82, 0.85, and 0.76, respectively), whereas GAPDH was ranked last at the 5-, 15-, and 30-minute time points (M=1.38, 1.44, and 1.39, respectively). A positive correlation was found by linear regression between the thawing time and 2 to the power of crossing point values of all candidate reference genes (P<0.05). The mean coefficient of variance of all reference genes increased significantly at time points 5, 15, and 30 minutes compared with 0 minutes (P<0.01). In conclusion, using the geNorm algorithm, PPIA was identified as the most stably expressed gene between malignant and nonmalignant renal tissue samples that were thawed for similar time periods. All the reference genes showed high variations along with the thawing time; it should be recommended to use a combination of several candidate reference genes when comparing samples thawed for different time periods.

  13. Impact of five SNPs in dopamine-related genes on executive function.

    PubMed

    Mitaki, S; Isomura, M; Maniwa, K; Yamasaki, M; Nagai, A; Nabika, T; Yamaguchi, S

    2013-01-01

    Dopamine neurotransmission is a critical factor for executive function, which is controlled by the prefrontal cortex in humans. Although the contribution of genetic factors to the regulation of brain dopaminergic activity is widely acknowledged, identification of a genotype-phenotype association has not yet been clearly established. In this study, we therefore evaluated the effects of five functional single-nucleotide polymorphisms (SNPs) in specific genes related to dopamine neurotransmission on executive function in a general population. Participants of the health examination at the Shimane Institute of Health Science were recruited for this study (n = 964). To evaluate executive function, the Frontal Assessment Battery (FAB) was administered. SNPs were genotyped using the TaqMan method. A significant association was found between an SNP in the catechol-O-methyltransferase (COMT) gene (rs4680) encoding the low-activity Met allele and FAB score (P = 0.003). Of note, the flexibility subset of the FAB was associated with the SNP in COMT (P = 0.003) after adjustment for confounding factors. The generalized multifactor dimensionality reduction method identified that the combination of two SNPs in the COMT gene (rs4680) and the dopamine D4 receptor gene (rs1800955) had a significant effect on FAB score. Our study indicates a contribution of rs4680 in the COMT gene to the variability in executive function, as assessed by the FAB. In addition, we have indicated that a complex gene-gene interaction between SNPs in the genes related to dopamine neurotransmission may influence executive function in a general population. © 2012 John Wiley & Sons A/S.

  14. Antibiotic resistance genes in surface water of eutrophic urban lakes are related to heavy metals, antibiotics, lake morphology and anthropic impact.

    PubMed

    Yang, Yuyi; Xu, Chen; Cao, Xinhua; Lin, Hui; Wang, Jun

    2017-08-01

    Urban lakes are impacted by heavy human activities and represent potential reservoirs for antibiotic resistance genes. In this study, six urban lakes in Wuhan, central China were selected to analyze the distribution of sulfonamide resistance (sul) genes, tetracycline resistance (tet) genes and quinolone resistance (qnr) genes and their relationship with heavy metals, antibiotics, lake morphology and anthropic impact. sul1 and sul2 were detected in all six lakes and dominated the types of antibiotic resistance genes, which accounted for 86.28-97.79% of the total antibiotic resistance gene abundance. For eight tested tet genes, antibiotic efflux pumps (tetA, tetB, tetC, and tetG) genes were all observed in six lakes and had higher relative abundance than ribosomal protection protein genes (tetM and tetQ). For 4 plasmid mediated quinolone resistance genes, only qnrD is found in all six lakes. The class I integron (intI1) is also found to be a very important media for antibiotic resistance gene propagation in urban lakes. The results of redundancy analysis and variation partitioning analysis showed that antibiotic and co-selection with heavy metals were the major factors driving the propagation of antibiotic resistance genes in six urban lakes. The heavily eutrophic Nanhu Lake and Shahu Lake which located in a high density building area with heavy human activities had the higher relative abundance of total antibiotic resistance genes. Our study could provide a useful reference for antibiotic resistance gene abundance in urban lakes with high anthropic impact.

  15. Antimicrobial-Resistant Bacterial Populations and Antimicrobial Resistance Genes Obtained from Environments Impacted by Livestock and Municipal Waste

    PubMed Central

    Durso, Lisa M.; Harhay, Dayna M.; Schmidt, John W.

    2015-01-01

    This study compared the populations of antimicrobial-resistant bacteria and the repertoire of antimicrobial resistance genes in four environments: effluent of three municipal wastewater treatment facilities, three cattle feedlot runoff catchment ponds, three swine waste lagoons, and two “low impact” environments (an urban lake and a relict prairie). Multiple liquid and solid samples were collected from each environment. The prevalences and concentrations of antimicrobial-resistant (AMR) Gram-negative (Escherichia coli and Salmonella enterica) and Gram-positive (enterococci) bacteria were determined from individual samples (n = 174). The prevalences of 84 antimicrobial resistance genes in metagenomic DNA isolated from samples pooled (n = 44) by collection date, location, and sample type were determined. The prevalences and concentrations of AMR E. coli and Salmonella were similar among the livestock and municipal sample sources. The levels of erythromycin-resistant enterococci were significantly higher in liquid samples from cattle catchment ponds and swine waste lagoons than in liquid samples from municipal wastewater treatment facilities, but solid samples from these environments did not differ significantly. Similarly, trimethoprim/sulfamethoxazole-resistant E. coli concentrations were significantly higher in swine liquid than in municipal liquid samples, but there was no difference in solid samples. Multivariate analysis of the distribution of antimicrobial resistance genes using principal coordinate analysis showed distinct clustering of samples with livestock (cattle and swine), low impact environment and municipal samples forming three separate clusters. The numbers of class A beta-lactamase, class C beta-lactamase, and fluoroquinolone resistance genes detected were significantly higher (P < 0.05) in municipal samples than in cattle runoff or swine lagoon samples. In conclusion, we report that AMR is a very widespread phenomenon and that similar

  16. An integrative study on the impact of highly differentially methylated genes on expression and cancer etiology

    PubMed Central

    2017-01-01

    DNA methylation is an important epigenetic phenomenon that plays a key role in the regulation of expression. Most of the studies on the topic of methylation’s role in cancer mechanisms include analyses based on differential methylation, with the integration of expression information as supporting evidence. In the present study, we sought to identify methylation-driven patterns by also integrating protein-protein interaction information. We performed integrative analyses of DNA methylation, expression, SNP and copy number data on paired samples from six different cancer types. As a result, we found that genes that show a methylation change larger than 32.2% may influence cancer-related genes via fewer interaction steps and with much higher percentages compared with genes showing a methylation change less than 32.2%. Additionally, we investigated whether there were shared cancer mechanisms among different cancer types. Specifically, five cancer types shared a change in AGTR1 and IGF1 genes, which implies that there may be similar underlying disease mechanisms among these cancers. Additionally, when the focus was placed on distinctly altered genes within each cancer type, we identified various cancer-specific genes that are also supported in the literature and may play crucial roles as therapeutic targets. Overall, our novel graph-based approach for identifying methylation-driven patterns will improve our understanding of the effects of methylation on cancer progression and lead to improved knowledge of cancer etiology. PMID:28178311

  17. Impact of genes highly correlated with MMSET myeloma on survival of Non-MMSET myeloma patients

    PubMed Central

    Wu, S. Peter; Pfeiffer, Ruth M.; Ahn, Inhye E.; Mailankody, Sham; Sonneveld, Pieter; van Duin, Mark; Munshi, Nikhil; Walker, Brian; Morgan, Gareth; Landgren, Ola

    2017-01-01

    Purpose The poor prognosis of multiple myeloma with t(4;14) is driven by the fusion of genes encoding MMSET and immunoglobulin heavy chain. Specific genes affected by MMSET and their clinical implications in Non-MMSET myeloma remain undetermined. Experimental design We obtained gene-expression profiles of 1,032 newly diagnosed myeloma patients enrolled in Total Therapy 2, Total Therapy 3, Myeloma IX, and HOVON65-GMMGHD4 trials, and 156 patients from Multiple Myeloma Resource Collection. Probes most correlated with MMSET myeloma were selected based on a multivariable linear regression and Bonferroni correction, and refined based on the strength of association with survival in Non-MMSET patients. Results Ten MMSET-like probes were associated with poor survival in Non-MMSET myeloma. Non-MMSET myeloma patients in the highest quartile of the 10-gene signature (MMSET-like myeloma) had 5-year overall survival similar to that of MMSET myeloma (highest quartile vs. lowest quartile hazard ratio [HR]=2.0, 95% confidence interval [CI] 1.5-2.8 in MMSET-like myeloma; HR=2.3, 95% CI: 1.6-3.3 in MMSET myeloma). Analyses of MMSET-like gene signature suggested the involvement of p53 and MYC pathways. Conclusion MMSET-like gene signature captures a subset of high-risk myeloma patients under-represented by conventional risk stratification platforms, and defines a distinct biological subtype. PMID:26847058

  18. Impact of gene patents and licensing practices on access to genetic testing for hearing loss.

    PubMed

    Chandrasekharan, Subhashini; Fiffer, Melissa

    2010-04-01

    Genetic testing for heritable hearing loss involves a mix of patented and unpatented genes, mutations and testing methods. More than half of all hearing loss is linked to inherited mutations, and five genes are most commonly tested for in the United States. There are no patents on three of these genes, but Athena Diagnostics holds exclusive licenses to test for a common mutation in the GJB2 gene associated with about 50% of all cases as well as mutations in the MTRNR1 gene. This fragmented intellectual property landscape made hearing loss a useful case study to assess whether patent rights in genetic testing can proliferate or overlap, and whether it is possible to gather the rights necessary to perform testing. Testing for hearing loss is widely available, primarily from academic medical centers. Based on literature reviews and interviews with researchers, research on the genetics of hearing loss has generally not been impeded by patents. There is no consistent evidence of a premium in testing prices attributable to patent status. Athena Diagnostics has, however, used its intellectual property to discourage other providers from offering some tests. There is no definitive answer about the suitability of current patenting and licensing of commonly tested genes because of continuing legal uncertainty about the extent of enforcement of patent rights. Clinicians have also expressed concerns that multiplex tests will be difficult to develop because of overlapping intellectual property and conflict with Athena's sole provider business model.

  19. Using ALoFT to determine the impact of putative loss-of-function variants in protein-coding genes.

    PubMed

    Balasubramanian, Suganthi; Fu, Yao; Pawashe, Mayur; McGillivray, Patrick; Jin, Mike; Liu, Jeremy; Karczewski, Konrad J; MacArthur, Daniel G; Gerstein, Mark

    2017-08-29

    Variants predicted to result in the loss of function of human genes have attracted interest because of their clinical impact and surprising prevalence in healthy individuals. Here, we present ALoFT (annotation of loss-of-function transcripts), a method to annotate and predict the disease-causing potential of loss-of-function variants. Using data from Mendelian disease-gene discovery projects, we show that ALoFT can distinguish between loss-of-function variants that are deleterious as heterozygotes and those causing disease only in the homozygous state. Investigation of variants discovered in healthy populations suggests that each individual carries at least two heterozygous premature stop alleles that could potentially lead to disease if present as homozygotes. When applied to de novo putative loss-of-function variants in autism-affected families, ALoFT distinguishes between deleterious variants in patients and benign variants in unaffected siblings. Finally, analysis of somatic variants in >6500 cancer exomes shows that putative loss-of-function variants predicted to be deleterious by ALoFT are enriched in known driver genes.Variants causing loss of function (LoF) of human genes have clinical implications. Here, the authors present a method to predict disease-causing potential of LoF variants, ALoFT (annotation of Loss-of-Function Transcripts) and show its application to interpreting LoF variants in different contexts.

  20. Stocking impacts the expression of candidate genes and physiological condition in introgressed brook charr (Salvelinus fontinalis) populations

    PubMed Central

    Lamaze, Fabien C; Garant, Dany; Bernatchez, Louis

    2013-01-01

    Translocation of plants and animal populations between environments is one of the major forms of anthropogenic perturbation experienced by pristine populations, and consequently, human-mediated hybridization by stocking practices between wild and exogenous conspecifics is of increasing concern. In this study, we compared the expression of seven candidate genes involved in multifactorial traits and regulatory pathways for growth as a function of level of introgressive hybridization between wild and domestic brook charr to test the null hypothesis of no effect of introgression on wild fish. Our analyses revealed that the expression of two of the genes tested, cytochrome c oxidase VIIa and the growth hormone receptor isoform I, was positively correlated with the level of introgression. We also observed a positive relationship between the extent of introgression and physiological status quantified by the Fulton's condition index. The expression of other genes was influenced by other variables, including year of sampling (reflecting different thermal conditions), sampling method and lake of origin. This is the first demonstration in nature that introgression from stocked populations has an impact on the expression of genes playing a role in important biological functions that may be related with fitness in wild introgressed populations. PMID:23467764

  1. Transcriptome-wide effects of inverted SINEs on gene expression and their impact on RNA polymerase II activity.

    PubMed

    Tajaddod, Mansoureh; Tanzer, Andrea; Licht, Konstantin; Wolfinger, Michael T; Badelt, Stefan; Huber, Florian; Pusch, Oliver; Schopoff, Sandy; Janisiw, Michael; Hofacker, Ivo; Jantsch, Michael F

    2016-10-25

    Short interspersed elements (SINEs) represent the most abundant group of non-long-terminal repeat transposable elements in mammalian genomes. In primates, Alu elements are the most prominent and homogenous representatives of SINEs. Due to their frequent insertion within or close to coding regions, SINEs have been suggested to play a crucial role during genome evolution. Moreover, Alu elements within mRNAs have also been reported to control gene expression at different levels. Here, we undertake a genome-wide analysis of insertion patterns of human Alus within transcribed portions of the genome. Multiple, nearby insertions of SINEs within one transcript are more abundant in tandem orientation than in inverted orientation. Indeed, analysis of transcriptome-wide expression levels of 15 ENCODE cell lines suggests a cis-repressive effect of inverted Alu elements on gene expression. Using reporter assays, we show that the negative effect of inverted SINEs on gene expression is independent of known sensors of double-stranded RNAs. Instead, transcriptional elongation seems impaired, leading to reduced mRNA levels. Our study suggests that there is a bias against multiple SINE insertions that can promote intramolecular base pairing within a transcript. Moreover, at a genome-wide level, mRNAs harboring inverted SINEs are less expressed than mRNAs harboring single or tandemly arranged SINEs. Finally, we demonstrate a novel mechanism by which inverted SINEs can impact on gene expression by interfering with RNA polymerase II.

  2. Identification of Nucleotide-Level Changes Impacting Gene Content and Genome Evolution in Orthopoxviruses

    PubMed Central

    Hatcher, Eneida L.; Hendrickson, Robert Curtis

    2014-01-01

    ABSTRACT Poxviruses are composed of large double-stranded DNA (dsDNA) genomes coding for several hundred genes whose variation has supported virus adaptation to a wide variety of hosts over their long evolutionary history. Comparative genomics has suggested that the Orthopoxvirus genus in particular has undergone reductive evolution, with the most recent common ancestor likely possessing a gene complement consisting of all genes present in any existing modern-day orthopoxvirus species, similar to the current Cowpox virus species. As orthopoxviruses adapt to new environments, the selection pressure on individual genes may be altered, driving sequence divergence and possible loss of function. This is evidenced by accumulation of mutations and loss of protein-coding open reading frames (ORFs) that progress from individual missense mutations to gene truncation through the introduction of early stop mutations (ESMs), gene fragmentation, and in some cases, a total loss of the ORF. In this study, we have constructed a whole-genome alignment for representative isolates from each Orthopoxvirus species and used it to identify the nucleotide-level changes that have led to gene content variation. By identifying the changes that have led to ESMs, we were able to determine that short indels were the major cause of gene truncations and that the genome length is inversely proportional to the number of ESMs present. We also identified the number and types of protein functional motifs still present in truncated genes to assess their functional significance. IMPORTANCE This work contributes to our understanding of reductive evolution in poxviruses by identifying genomic remnants such as single nucleotide polymorphisms (SNPs) and indels left behind by evolutionary processes. Our comprehensive analysis of the genomic changes leading to gene truncation and fragmentation was able to detect some of the remnants of these evolutionary processes still present in orthopoxvirus genomes and

  3. Upon Accounting for the Impact of Isoenzyme Loss, Gene Deletion Costs Anticorrelate with Their Evolutionary Rates

    PubMed Central

    Xia, Yu; Segrè, Daniel

    2017-01-01

    System-level metabolic network models enable the computation of growth and metabolic phenotypes from an organism’s genome. In particular, flux balance approaches have been used to estimate the contribution of individual metabolic genes to organismal fitness, offering the opportunity to test whether such contributions carry information about the evolutionary pressure on the corresponding genes. Previous failure to identify the expected negative correlation between such computed gene-loss cost and sequence-derived evolutionary rates in Saccharomyces cerevisiae has been ascribed to a real biological gap between a gene’s fitness contribution to an organism “here and now” and the same gene’s historical importance as evidenced by its accumulated mutations over millions of years of evolution. Here we show that this negative correlation does exist, and can be exposed by revisiting a broadly employed assumption of flux balance models. In particular, we introduce a new metric that we call “function-loss cost”, which estimates the cost of a gene loss event as the total potential functional impairment caused by that loss. This new metric displays significant negative correlation with evolutionary rate, across several thousand minimal environments. We demonstrate that the improvement gained using function-loss cost over gene-loss cost is explained by replacing the base assumption that isoenzymes provide unlimited capacity for backup with the assumption that isoenzymes are completely non-redundant. We further show that this change of the assumption regarding isoenzymes increases the recall of epistatic interactions predicted by the flux balance model at the cost of a reduction in the precision of the predictions. In addition to suggesting that the gene-to-reaction mapping in genome-scale flux balance models should be used with caution, our analysis provides new evidence that evolutionary gene importance captures much more than strict essentiality. PMID:28107392

  4. Impact of the IL-10 Promoter Gene Polymorphisms in the Severity of Chronic Hepatitis B Infection

    PubMed Central

    Ghaleh Baghi, Sahand; Alavian, Seyed Moayed; Mehrnoush, Leila; Salimi, Shima

    2015-01-01

    Background: Interleukin-10 (IL-10) is an important anti-inflammatory cytokine. The polymorphisms of its promoter gene have been considered to be related with the chronicity of hepatitis B infection. Objectives: The aim of this study was to evaluate the polymorphisms at different positions in the IL-10 promoter gene in patients with chronic hepatitis B. Patients and Methods: Totally, 166 patients with chronic hepatitis B infection were enrolled. Genotypes at different positions (i.e. -819, - 592, and - 1082) in the IL-10 gene promoter were determined. Results: The C/A genotype at position -592, C/T genotype at position -819, and GCC/ATA haplotype of the IL-10 gene promoter were significantly more common in the patients with cirrhosis. The genotypes were significantly different between the hepatitis B e antigen (HBeAg)-negative and HBeAg-positive patients at position -592 (C/A and C/C), position -819 (C/C and C/T), and position -1082 (A/A and G/A). Conclusions: Some IL-10 promoter gene polymorphisms predisposed the infected hepatitis B virus cases to cirrhosis in our study population. PMID:26300930

  5. Impact of gene patents and licensing practices on access to genetic testing for Alzheimer disease.

    PubMed

    Skeehan, Katie; Heaney, Christopher; Cook-Deegan, Robert

    2010-04-01

    Genetic testing for Alzheimer disease includes genotyping for apolipoprotein E, for late-onset Alzheimer disease, and three rare autosomal dominant, early-onset forms of Alzheimer disease associated with different genes (APP, PSEN1, and PSEN2). According to researchers, patents have not impeded research in the field, nor were patents an important consideration in the quest for the genetic risk factors. Athena Diagnostics holds exclusive licenses from Duke University for three "method" patents covering apolipoprotein E genetic testing. Athena offers tests for apolipoprotein E and genes associated with early-onset, autosomal-dominant Alzheimer disease. One of those presenilin genes is patented and exclusively licensed to Athena; the other presenilin gene was patented but the patent was allowed to lapse; and one (amyloid precursor protein) is patented as a research tool. Direct-to-consumer testing is available for some Alzheimer disease-related genes, apparently without a license. Athena Diagnostics consolidated its position in the market for Alzheimer disease genetic testing by collecting exclusive rights to patents arising from university research. Duke University also used its licenses to Athena to enforce adherence to clinical guidelines, including elimination of the service from Smart Genetics, which was offering direct-to-consumer risk assessment based on apolipoprotein E genotyping.

  6. Impact of azithromycin treatment on macrophage gene expression in subjects with cystic fibrosis.

    PubMed

    Cory, Theodore J; Birket, Susan E; Murphy, Brian S; Hayes, Don; Anstead, Michael I; Kanga, Jamshed F; Kuhn, Robert J; Bush, Heather M; Feola, David J

    2014-03-01

    Azithromycin treatment improves clinical parameters in patients with CF, and alters macrophage activation from a pro-inflammatory (M1) phenotype to a pro-fibrotic, alternatively activated (M2) phenotype. The transcriptional profile of cells from patients receiving azithromycin is unknown. Gene expression in association with macrophage polarization, inflammation, and tissue remodeling was assessed from sputum samples collected from patients with CF. Transcriptional profiles and clinical characteristics, including azithromycin therapy, were compared. Expression of NOS2 and TNFα was decreased in subjects receiving azithromycin, whereas expression of M2-associated genes was unaffected. Principal component analysis revealed gene expression profiles consistent with M1- (MMP9, NOS2, and TLR4) or M2-polarization (CCL18, fibronectin, and MR1) in select subject groups. These expression signatures did not significantly correlate with clinical characteristics. Pro-inflammatory gene expression was low in subjects receiving AZM. Genes were stratified into groupings characteristic of M1- or M2-polarization, suggesting that overall polarization status is distinct among patient groups. © 2013.

  7. Interactions between DNA and Gemini surfactant: impact on gene therapy: part I.

    PubMed

    Ahmed, Taksim; Kamel, Amany O; Wettig, Shawn D

    2016-02-01

    Nonviral gene therapy using gemini surfactants is a unique approach to medicine that can be adapted toward the treatment of various diseases. Recently, gemini surfactants have been utilized as candidates for the formation of nonviral vectors. The chemical structure of the surfactant (variations in the alkyl tail length and spacer/head group) and the resulting physicochemical properties of the lipoplexes are critical parameters for efficient gene transfection. Moreover, studying the interaction of the surfactant with DNA can help in designing an efficient vector and understanding how transfection complexes overcome various cellular barriers. Part I of this review provides an overview of various types of gemini surfactants designed for gene therapy and their transfection efficiency; and Part II will focus on different novel methods utilized to understand the interactions between the gemini and DNA in a lipoplex.

  8. Interaction between smoking history and gene expression levels impacts survival of breast cancer patients.

    PubMed

    Andres, Sarah A; Bickett, Katie E; Alatoum, Mohammad A; Kalbfleisch, Theodore S; Brock, Guy N; Wittliff, James L

    2015-08-01

    In contrast to studies focused on cigarette smoking and risk of breast cancer occurrence, this study explored the influence of smoking on breast cancer recurrence and progression. The goal was to evaluate the interaction between smoking history and gene expression levels on recurrence and overall survival of breast cancer patients. Multivariable Cox proportional hazards models were fitted for 48 cigarette smokers, 50 non-smokers, and the total population separately to determine which gene expressions and gene expression/cigarette usage interaction terms were significant in predicting overall and disease-free survival in breast cancer patients. Using methods similar to Andres et al. (BMC Cancer 13:326, 2013a; Horm Cancer 4:208-221, 2013b), multivariable analyses revealed CENPN, CETN1, CYP1A1, IRF2, LECT2, and NCOA1 to be important predictors for both breast carcinoma recurrence and mortality among smokers. Additionally, COMT was important for recurrence, and NAT1 and RIPK1 were important for mortality. In contrast, only IRF2, CETN1, and CYP1A1 were significant for disease recurrence and mortality among non-smokers, with NAT2 additionally significant for survival. Analysis of interaction between smoking status and gene expression values using the combined samples revealed significant interactions between smoking status and CYP1A1, LECT2, and CETN1. Signatures consisting of 7-8 genes were highly predictive for breast cancer recurrence and overall survival among smokers, with median C-index values of 0.8 and 0.73 for overall survival and recurrence, respectively. In contrast, median C-index values for non-smokers was only 0.59. Hence, significant interactions between gene expression and smoking status can play a key role in predicting breast cancer patient outcomes.

  9. Frequency of undetected CYP2D6 hybrid genes in clinical samples: impact on phenotype prediction.

    PubMed

    Black, John Logan; Walker, Denise L; O'Kane, Dennis J; Harmandayan, Maria

    2012-01-01

    Cytochrome P450 2D6 (CYP2D6) is highly polymorphic. CYP2D6-2D7 hybrid genes can be present in samples containing CYP2D6*4 and CYP2D6*10 alleles. CYP2D7-2D6 hybrid genes can be present in samples with duplication signals and in samples with homozygous genotyping results. The frequency of hybrid genes in clinical samples is unknown. We evaluated 1390 samples for undetected hybrid genes by polymerase chain reaction (PCR) amplification, PCR fragment analysis, TaqMan copy number assays, DNA sequencing, and allele-specific primer extension assay. Of 508 CYP2D6*4-containing samples, 109 (21.5%) harbored CYP2D6*68 + *4-like, whereas 9 (1.8%) harbored CYP2D6*4N + *4-like. Of 209 CYP2D6*10-containing samples, 44 (21.1%) were found to have CYP2D6*36 + *10. Of 332 homozygous samples, 4 (1.2%) harbored a single CYP2D7-2D6 hybrid, and of 341 samples with duplication signals, 25 (7.3%) harbored an undetected CYP2D7-2D6 hybrid. Phenotype before and after accurate genotyping was predicted using a method in clinical use. The presence of hybrid genes had no effect on the phenotype prediction of CYP2D6*4- and CYP2D6*10-containing samples. Four of four (100%) homozygous samples containing a CYP2D7-2D6 gene had a change in predicted phenotype, and 23 of 25 (92%) samples with a duplication signal and a CYP2D7-2D6 gene had a change in predicted phenotype. Four novel genes were identified (CYP2D6*13A1 variants 1 and 2, CYP2D6*13G1, and CYP2D6*13G2), and two novel hybrid tandem structures consisting of CYP2D6*13B + *68×2 + *4-like and CYP2D6*13A1 variant 2 + *1×N were observed.

  10. Impact of Natural IgM Concentration on Gene Therapy with Adenovirus Type 5 Vectors

    PubMed Central

    Qiu, Qi; Xu, Zhili; Tian, Jie; Moitra, Rituparna; Gunti, Sreenivasulu; Notkins, Abner L.

    2014-01-01

    Natural IgM inhibits gene transfer by adenovirus type 5 (Ad5) vectors. We show that polyreactive natural IgM antibodies bind to Ad5 and that inhibition of liver transduction by IgM depends on Kupffer cells. By manipulating IgM concentration in vivo, we demonstrate that IgM inhibits liver transduction in a concentration-dependent manner. We further show that differences in natural IgM between BALB/c and C57BL/6 mice contribute to lower efficiency of Ad5 gene transfer in BALB/c mice. PMID:25552715

  11. The impact of low molecular weight heparin on obstetric outcomes among unexplained recurrent miscarriages complicated with methylenetetrahydrofolate reductase gene polymorphism.

    PubMed

    Cetin, Orkun; Karaman, Erbil; Cim, Numan; Dirik, Deniz; Sahin, Hanim Guler; Kara, Erdal; Esen, Ramazan

    2017-01-01

    The association between methylenetetrahydrofolate reductase gene polymorphisms and unexplained recurrent miscarriage is elusive. The recommendations for improving pregnancy outcomes in these patients keep changing based on the available evidence. The aim of this study is to analyze the impact of low molecular weight heparin on obstetric outcomes of recurrent miscarriage patients complicated with methylenetetrahydrofolate reductase gene polymorphism. We reviewed medical records of 121 patients with a history of recurrent miscarriage complicated by methylenetetrahydrofolate reductase gene polymorphisms, retrospectively. From among them, 68 patients were treated only with folic acid and iron. The remaining 53 patients were treated with folic acid, iron and prophylactic doses of low molecular weight heparin. The subsequent pregnancy outcomes of these patients were noted. The live birth rate was higher in patients with anticoagulant therapy than in patients without anticoagulant therapy (48.5% vs. 69.8%, respectively, p: 0.015) and the congenital anomaly rate was lower in anticoagulant therapy group (17.6% vs. 3.8%, respectively, p: 0.022). The other obstetric outcomes were found to be similar between the two groups. The current study demonstrated that low molecular weight heparin improved the live birth rates among unex-plained recurrent miscarriage patients complicated with methylenetetrahydrofolate reductase gene polymorphisms. How-ever, the routine use of low molecular weight heparin did not improve the late pregnancy complications in these selected patients in the eastern region of our country. Further studies are needed to discriminate the effect of anticoagulation on the live birth rate of each of methylenetetrahydrofolate reductase gene polymorphism type.

  12. Impact of I/D polymorphism of ACE gene on risk of development and course of chronic obstructive pulmonary disease

    PubMed Central

    Homa-Mlak, Iwona; Powrózek, Tomasz; Mackiewicz, Barbara; Michnar, Marek; Krawczyk, Paweł; Dziedzic, Marcin; Rubinsztajn, Renata; Chazan, Ryszarda; Milanowski, Janusz; Małecka-Massalska, Teresa

    2016-01-01

    Introduction Chronic obstructive pulmonary disease (COPD) affects more than 10% of the world's population over 40 years of age. The main exogenous risk factor is cigarette smoking; however, only 20% of smokers develop COPD, indicating that some other factors, e.g. genetic, may play an important role in the disease pathogenesis. Recent research indicates that ACE (angiotensin-converting enzyme) may be a susceptibility gene for asthma or COPD. The aim of our study was to determine the influence of I/D (insertion/deletion) polymorphism of the ACE gene (AluYa5, rs4646994) on the risk and course of COPD. Material and methods We investigated ACE I/D polymorphism in 206 COPD and 165 healthy Caucasian subjects. Results In the generalized linear model (GLZ) analysis of the influence of selected factors on presence of COPD we found a significant independent effect for male sex (repeatedly increases the risk of COPD, OR = 7.7, p = 0.049), as well as smoking or lower body mass index, but only in combination with older age (OR = 0.96, p = 0.003 and OR = 1.005, p = 0.04 respectively). Interestingly, analysis of factors which may influence the risk of a higher number of exacerbations demonstrated that occurrence of DD genotype, but only in men, is associated with a lower risk (OR = 0.7, p = 0.03) of this complication. Conclusions We suggest that ACE may not be a susceptibility gene for the origin of COPD but a disease-modifying gene. Since the impact of I/D polymorphism of the ACE gene on COPD risk is moderate or negligible, other molecular changes, that will help predict the development of this disease, should still be sought. PMID:27186170

  13. Impact of treatment strategies on cephalosporin and tetracycline resistance gene quantities in the bovine fecal metagenome.

    PubMed

    Kanwar, Neena; Scott, H Morgan; Norby, Bo; Loneragan, Guy H; Vinasco, Javier; Cottell, Jennifer L; Chalmers, Gabhan; Chengappa, Muckatira M; Bai, Jianfa; Boerlin, Patrick

    2014-05-29

    The study objective was to determine the effects of two treatment regimens on quantities of ceftiofur and tetracycline resistance genes in feedlot cattle. The two regimens were ceftiofur crystalline-free acid (CCFA) administered to either one or all steers within a pen and subsequent feeding/not feeding of therapeutic doses of chlortetracycline. A 26-day randomized controlled field trial was conducted on 176 steers. Real-time PCR was used to quantify bla(CMY-2), bla(CTX-M), tet(A), tet(B), and 16S rRNA gene copies/gram of feces from community DNA. A significant increase in ceftiofur resistance and a decrease in tetracycline resistance elements were observed among the treatment groups in which all steers received CCFA treatment, expressed as gene copies/gram of feces. Subsequent chlortetracycline administration led to rapid expansion of both ceftiofur and tetracycline resistance gene copies/gram of feces. Our data suggest that chlortetracycline is contraindicated when attempting to avoid expansion of resistance to critically important third-generation cephalosporins.

  14. Gene-environment interactions and the impact on obesity and lipid profile phenotypes

    USDA-ARS?s Scientific Manuscript database

    Sequencing the human genome provided the data, human intellectual capital and technology, particularly in terms of infrastructure and methodologies, to begin discovering genes involved in a wide range of human diseases and afflictions. This has led to a resurgence in genetics with the advent of geno...

  15. Impact of treatment strategies on cephalosporin and tetracycline resistance gene quantities in the bovine fecal metagenome

    PubMed Central

    Kanwar, Neena; Scott, H. Morgan; Norby, Bo; Loneragan, Guy H.; Vinasco, Javier; Cottell, Jennifer L.; Chalmers, Gabhan; Chengappa, Muckatira M.; Bai, Jianfa; Boerlin, Patrick

    2014-01-01

    The study objective was to determine the effects of two treatment regimens on quantities of ceftiofur and tetracycline resistance genes in feedlot cattle. The two regimens were ceftiofur crystalline-free acid (CCFA) administered to either one or all steers within a pen and subsequent feeding/not feeding of therapeutic doses of chlortetracycline. A 26-day randomized controlled field trial was conducted on 176 steers. Real-time PCR was used to quantify blaCMY-2, blaCTX-M, tet(A), tet(B), and 16S rRNA gene copies/gram of feces from community DNA. A significant increase in ceftiofur resistance and a decrease in tetracycline resistance elements were observed among the treatment groups in which all steers received CCFA treatment, expressed as gene copies/gram of feces. Subsequent chlortetracycline administration led to rapid expansion of both ceftiofur and tetracycline resistance gene copies/gram of feces. Our data suggest that chlortetracycline is contraindicated when attempting to avoid expansion of resistance to critically important third-generation cephalosporins. PMID:24872333

  16. Population diversity and adaptive evolution in keratinization genes: impact of environment in shaping skin phenotypes.

    PubMed

    Gautam, Pramod; Chaurasia, Amit; Bhattacharya, Aniket; Grover, Ritika; Mukerji, Mitali; Natarajan, Vivek T

    2015-03-01

    Several studies have demonstrated the role of climatic factors in shaping skin phenotypes, particularly pigmentation. Keratinization is another well-designed feature of human skin, which is involved in modulating transepidermal water loss (TEWL). Although this physiological process is closely linked to climate, presently it is not clear whether genetic diversity is observed in keratinization and whether this process also responds to the environmental pressure. To address this, we adopted a multipronged approach, which involved analysis of 1) copy number variations in diverse Indian and HapMap populations from varied geographical regions; 2) genetic association with geoclimatic parameters in 61 populations of dbCLINE database in a set of 549 genes from four processes namely keratinization, pigmentation, epidermal differentiation, and housekeeping functions; 3) sequence divergence in 4,316 orthologous promoters and corresponding exonic regions of human and chimpanzee with macaque as outgroup, and 4) protein sequence divergence (Ka/Ks) across nine vertebrate classes, which differ in their extent of TEWL. Our analyses demonstrate that keratinization and epidermal differentiation genes are under accelerated evolution in the human lineage, relative to pigmentation and housekeeping genes. We show that this entire pathway may have been driven by environmental selection pressure through concordant functional polymorphisms across several genes involved in skin keratinization. Remarkably, this underappreciated function of skin may be a crucial determinant of adaptation to diverse environmental pressures across world populations.

  17. Impact of Hfq on Global Gene Expression and Intracellular Survival in Brucella melitensis

    PubMed Central

    Du, Xinying; Yuan, Xitong; Wang, Zhoujia; Gong, Chunli; Zhuang, Yubin; Lei, Shuangshuang; Su, Xiao; Wang, Xuesong; Huang, Liuyu; Zhong, Zhijun; Peng, Guangneng; Yuan, Jing; Chen, Zeliang; Wang, Yufei

    2013-01-01

    Brucella melitensis is a facultative intracellular bacterium that replicates within macrophages. The ability of brucellae to survive and multiply in the hostile environment of host macrophages is essential to its virulence. The RNA-binding protein Hfq is a global regulator that is involved in stress resistance and pathogenicity. Here we demonstrate that Hfq is essential for stress adaptation and intracellular survival in B. melitensis. A B. melitensis hfq deletion mutant exhibits reduced survival under environmental stresses and is attenuated in cultured macrophages and mice. Microarray-based transcriptome analyses revealed that 359 genes involved in numerous cellular processes were dysregulated in the hfq mutant. From these same samples the proteins were also prepared for proteomic analysis to directly identify Hfq-regulated proteins. Fifty-five proteins with significantly affected expression were identified in the hfq mutant. Our results demonstrate that Hfq regulates many genes and/or proteins involved in metabolism, virulence, and stress responses, including those potentially involved in the adaptation of Brucella to the oxidative, acid, heat stress, and antibacterial peptides encountered within the host. The dysregulation of such genes and/or proteins could contribute to the attenuated hfq mutant phenotype. These findings highlight the involvement of Hfq as a key regulator of Brucella gene expression and facilitate our understanding of the role of Hfq in environmental stress adaptation and intracellular survival of B. melitensis. PMID:23977181

  18. Impact of enhancin genes on potency of LdNPV in gypsy moth

    Treesearch

    Kelli Hoover; Jim McNeil; Alyssa Gendron; James. Slavicek

    2011-01-01

    Lymantria dispar nucleopolyhedrovirus (LdNPV) contains two enhancin genes (E1 and E2) encoding proteases that degrade key peritrophic matrix (PM) proteins, thereby promoting infection and mortality by the virus. In a previous study, gypsy moth larvae inoculated with LdNPV in which both E1 and E2 were deleted (double deletion virus) resulted in a non-...

  19. Differential Impact of the "FMR1" Gene on Visual Processing in Fragile X Syndrome

    ERIC Educational Resources Information Center

    Kogan, Cary S.; Boutet, Isabelle; Cornish, Kim; Zangenehpour, Shahin; Mullen, Kathy T.; Holden, Jeanette J. A.; Kaloustian, Vazken M. Der; Andermann, Eva; Chaudhuri, Avi

    2004-01-01

    Fragile X syndrome (FXS) is the most common form of heritable mental retardation, affecting (~ around) 1 in 4000 males. The syndrome arises from expansion of a trinucleotide repeat in the 5'-untranslated region of the fragile X mental retardation 1 ("FMR1") gene, leading to methylation of the promoter sequence and lack of the fragile X mental…

  20. The Protein Kinase KIS Impacts Gene Expression during Development and Fear Conditioning in Adult Mice

    PubMed Central

    Manceau, Valérie; Kremmer, Elisabeth; Nabel, Elizabeth G.; Maucuer, Alexandre

    2012-01-01

    The brain-enriched protein kinase KIS (product of the gene UHMK1) has been shown to phosphorylate the human splicing factor SF1 in vitro. This phosphorylation in turn favors the formation of a U2AF65-SF1-RNA complex which occurs at the 3′ end of introns at an early stage of spliceosome assembly. Here, we analyzed the effects of KIS knockout on mouse SF1 phosphorylation, physiology, adult behavior, and gene expression in the neonate brain. We found SF1 isoforms are differently expressed in KIS-ko mouse brains and fibroblasts. Re-expression of KIS in fibroblasts restores a wild type distribution of SF1 isoforms, confirming the link between KIS and SF1. Microarray analysis of transcripts in the neonate brain revealed a subtle down-regulation of brain specific genes including cys-loop ligand-gated ion channels and metabolic enzymes. Q-PCR analyses confirmed these defects and point to an increase of pre-mRNA over mRNA ratios, likely due to changes in splicing efficiency. While performing similarly in prepulse inhibition and most other behavioral tests, KIS-ko mice differ in spontaneous activity and contextual fear conditioning. This difference suggests that disregulation of gene expression due to KIS inactivation affects specific brain functions. PMID:22937132

  1. Meiotic drive impacts expression and evolution of x-linked genes in stalk-eyed flies.

    PubMed

    Reinhardt, Josephine A; Brand, Cara L; Paczolt, Kimberly A; Johns, Philip M; Baker, Richard H; Wilkinson, Gerald S

    2014-01-01

    Although sex chromosome meiotic drive has been observed in a variety of species for over 50 years, the genes causing drive are only known in a few cases, and none of these cases cause distorted sex-ratios in nature. In stalk-eyed flies (Teleopsis dalmanni), driving X chromosomes are commonly found at frequencies approaching 30% in the wild, but the genetic basis of drive has remained elusive due to reduced recombination between driving and non-driving X chromosomes. Here, we used RNAseq to identify transcripts that are differentially expressed between males carrying either a driving X (XSR) or a standard X chromosome (XST), and found hundreds of these, the majority of which are X-linked. Drive-associated transcripts show increased levels of sequence divergence (dN/dS) compared to a control set, and are predominantly expressed either in testes or in the gonads of both sexes. Finally, we confirmed that XSR and XST are highly divergent by estimating sequence differentiation between the RNAseq pools. We found that X-linked transcripts were often strongly differentiated (whereas most autosomal transcripts were not), supporting the presence of a relatively large region of recombination suppression on XSR presumably caused by one or more inversions. We have identified a group of genes that are good candidates for further study into the causes and consequences of sex-chromosome drive, and demonstrated that meiotic drive has had a profound effect on sequence evolution and gene expression of X-linked genes in this species.

  2. Essentiality of mmpL3 and impact of its silencing on Mycobacterium tuberculosis gene expression

    PubMed Central

    Degiacomi, Giulia; Benjak, Andrej; Madacki, Jan; Boldrin, Francesca; Provvedi, Roberta; Palù, Giorgio; Kordulakova, Jana; Cole, Stewart T.; Manganelli, Riccardo

    2017-01-01

    MmpL3 is an inner membrane transporter of Mycobacterium tuberculosis responsible for the export of trehalose momomycolate, a precursor of the mycobacterial outer membrane component trehalose dimycolate (TDM), as well as mycolic acids bound to arabinogalactan. MmpL3 represents an emerging target for tuberculosis therapy. In this paper, we describe the construction and characterization of an mmpL3 knockdown strain of M. tuberculosis. Downregulation of mmpL3 led to a stop in bacterial division and rapid cell death, preceded by the accumulation of TDM precursors. MmpL3 was also shown to be essential for growth in monocyte-derived human macrophages. Using RNA-seq we also found that MmpL3 depletion caused up-regulation of 47 genes and down-regulation of 23 genes (at least 3-fold change and false discovery rate ≤1%). Several genes related to osmoprotection and metal homeostasis were induced, while several genes related to energy production and mycolic acids biosynthesis were repressed suggesting that inability to synthesize a correct outer membrane leads to changes in cellular permeability and a metabolic downshift. PMID:28240248

  3. Differential Impact of the "FMR1" Gene on Visual Processing in Fragile X Syndrome

    ERIC Educational Resources Information Center

    Kogan, Cary S.; Boutet, Isabelle; Cornish, Kim; Zangenehpour, Shahin; Mullen, Kathy T.; Holden, Jeanette J. A.; Kaloustian, Vazken M. Der; Andermann, Eva; Chaudhuri, Avi

    2004-01-01

    Fragile X syndrome (FXS) is the most common form of heritable mental retardation, affecting (~ around) 1 in 4000 males. The syndrome arises from expansion of a trinucleotide repeat in the 5'-untranslated region of the fragile X mental retardation 1 ("FMR1") gene, leading to methylation of the promoter sequence and lack of the fragile X mental…

  4. Serratia entomophila bet gene induction and the impact of glycine betaine accumulation on desiccation tolerance.

    PubMed

    Sheen, T R; O'Callaghan, M; Smalley, D J; Ronson, C W; Hurst, M R H

    2013-02-01

    The genes involved in choline transport and oxidation to glycine betaine in the biopesticidal bacterium Serratia entomophila were characterized, and the potential of osmoprotectants, coupled with increased NaCl concentrations, to improve the desiccation tolerance of this species was investigated. Serratia entomophila carries sequences similar to the Escherichia coli betTIBA genes encoding a choline transporter and dehydrogenase, a betaine aldehyde dehydrogenase and a regulatory protein. Disruption of betA abolished the ability of Ser. entomophila to utilize choline as a carbon source. Quantitative reverse-transcriptase PCR analysis revealed that betA transcription was reduced compared to that of the upstream genes in the operon, and that NaCl and choline induced bet gene expression. Glycine betaine and choline increased the NaCl tolerance of Ser. entomophila, and osmotically preconditioned cultures survived better than control cultures following desiccation and immediately after application to agricultural soil. Addition of glycine betaine and NaCl to growth medium can greatly enhance the desiccation survival of Ser. entomophila, and its initial survival in soil. Serratia entomophila is sensitive to desiccation and does not persist under low soil moisture conditions. Techniques described here for enhancing the desiccation survival of Ser. entomophila can be used to improve formulations of this bacterium, and allow its application under a wider range of environmental conditions. © 2012 AgResearch.

  5. Potential impact of a single nucleotide polymorphism in the hyaluronan synthase 1 gene in Waldenstrom's macroglobulinemia.

    PubMed

    Adamia, Sophia; Treon, Steven P; Reiman, Tony; Tournilhac, Olivier; McQuarrie, Carrie; Mant, Michael J; Belch, Andrew R; Pilarski, Linda M

    2005-03-01

    The hyaluronan synthase 1 (HAS1) gene encodes a plasma membrane protein that synthesizes hyaluronan, an extracellular matrix molecule. Previously, in patients with Waldenstrom's macroglobulinemia (WM), we detected upregulation of HAS1 transcripts and identified aberrant splice variants of this gene. Aberrant splicing of HAS1 results from activation of cryptic splice sites. In turn, activation of cryptic donor and acceptor splice sites can be promoted by mutations occurring upstream of these sites and/or at the branch point of slicing. We measured the frequency of the HAS1 833A/G polymorphism (ie, single-nucleotide polymorphism; SNP) in patients with WM and healthy donors. Additionally, HAS1 gene expression was evaluated in the same group of patients. Our observations so far suggest that HAS1 833A/G SNPs contribute to aberrant splicing of this gene; this idea is supported by the fact that 833A/G SNP is located on an exonic splicing enhancer motif. Based on the results obtained thus far, we speculate that individuals with HAS1 833G/G genotype are predisposed toward aberrant HAS1 splicing and expression of HAS1 variants, resulting in an enhanced risk of developing WM. Study of a larger group of patients and healthy donors is needed to confirm these speculations and to evaluate the prognostic significance of these findings.

  6. Genetic variation of six desaturase genes in flax and their impact on fatty acid composition.

    PubMed

    Thambugala, Dinushika; Duguid, Scott; Loewen, Evelyn; Rowland, Gordon; Booker, Helen; You, Frank M; Cloutier, Sylvie

    2013-10-01

    Flax (Linum usitatissimum L.) is one of the richest plant sources of omega-3 fatty acids praised for their health benefits. In this study, the extent of the genetic variability of genes encoding stearoyl-ACP desaturase (SAD), and fatty acid desaturase 2 (FAD2) and 3 (FAD3) was determined by sequencing the six paralogous genes from 120 flax accessions representing a broad range of germplasm including some EMS mutant lines. A total of 6 alleles for sad1 and sad2, 21 for fad2a, 5 for fad2b, 15 for fad3a and 18 for fad3b were identified. Deduced amino acid sequences of the alleles predicted 4, 2, 3, 4, 6 and 7 isoforms, respectively. Allele frequencies varied greatly across genes. Fad3a, with 110 SNPs and 19 indels, and fad3b, with 50 SNPs and 5 indels, showed the highest levels of genetic variations. While most of the SNPs and all the indels were silent mutations, both genes carried nonsense SNP mutations resulting in premature stop codons, a feature not observed in sad and fad2 genes. Some alleles and isoforms discovered in induced mutant lines were absent in the natural germplasm. Correlation of these genotypic data with fatty acid composition data of 120 flax accessions phenotyped in six field experiments revealed statistically significant effects of some of the SAD and FAD isoforms on fatty acid composition, oil content and iodine value. The novel allelic variants and isoforms identified for the six desaturases will be a resource for the development of oilseed flax with unique and useful fatty acid profiles.

  7. Oxytocin Pathway Genes: Evolutionary Ancient System Impacting on Human Affiliation, Sociality, and Psychopathology.

    PubMed

    Feldman, Ruth; Monakhov, Mikhail; Pratt, Maayan; Ebstein, Richard P

    2016-02-01

    Oxytocin (OT), a nonapeptide signaling molecule originating from an ancestral peptide, appears in different variants across all vertebrate and several invertebrate species. Throughout animal evolution, neuropeptidergic signaling has been adapted by organisms for regulating response to rapidly changing environments. The family of OT-like molecules affects both peripheral tissues implicated in reproduction, homeostasis, and energy balance, as well as neuromodulation of social behavior, stress regulation, and associative learning in species ranging from nematodes to humans. After describing the OT-signaling pathway, we review research on the three genes most extensively studied in humans: the OT receptor (OXTR), the structural gene for OT (OXT/neurophysin-I), and CD38. Consistent with the notion that sociality should be studied from the perspective of social life at the species level, we address human social functions in relation to OT-pathway genes, including parenting, empathy, and using social relationships to manage stress. We then describe associations between OT-pathway genes with psychopathologies involving social dysfunctions such as autism, depression, or schizophrenia. Human research particularly underscored the involvement of two OXTR single nucleotide polymorphisms (rs53576, rs2254298) with fewer studies focusing on other OXTR (rs7632287, rs1042778, rs2268494, rs2268490), OXT (rs2740210, rs4813627, rs4813625), and CD38 (rs3796863, rs6449197) single nucleotide polymorphisms. Overall, studies provide evidence for the involvement of OT-pathway genes in human social functions but also suggest that factors such as gender, culture, and early environment often confound attempts to replicate first findings. We conclude by discussing epigenetics, conceptual implications within an evolutionary perspective, and future directions, especially the need to refine phenotypes, carefully characterize early environments, and integrate observations of social behavior across

  8. Impacts of Nonsynonymous Single Nucleotide Polymorphisms of Adiponectin Receptor 1 Gene on Corresponding Protein Stability: A Computational Approach

    PubMed Central

    Saleh, Md. Abu; Solayman, Md.; Paul, Sudip; Saha, Moumoni; Khalil, Md. Ibrahim; Gan, Siew Hua

    2016-01-01

    Despite the reported association of adiponectin receptor 1 (ADIPOR1) gene mutations with vulnerability to several human metabolic diseases, there is lack of computational analysis on the functional and structural impacts of single nucleotide polymorphisms (SNPs) of the human ADIPOR1 at protein level. Therefore, sequence- and structure-based computational tools were employed in this study to functionally and structurally characterize the coding nsSNPs of ADIPOR1 gene listed in the dbSNP database. Our in silico analysis by SIFT, nsSNPAnalyzer, PolyPhen-2, Fathmm, I-Mutant 2.0, SNPs&GO, PhD-SNP, PANTHER, and SNPeffect tools identified the nsSNPs with distorting functional impacts, namely, rs765425383 (A348G), rs752071352 (H341Y), rs759555652 (R324L), rs200326086 (L224F), and rs766267373 (L143P) from 74 nsSNPs of ADIPOR1 gene. Finally the aforementioned five deleterious nsSNPs were introduced using Swiss-PDB Viewer package within the X-ray crystal structure of ADIPOR1 protein, and changes in free energy for these mutations were computed. Although increased free energy was observed for all the mutants, the nsSNP H341Y caused the highest energy increase amongst all. RMSD and TM scores predicted that mutants were structurally similar to wild type protein. Our analyses suggested that the aforementioned variants especially H341Y could directly or indirectly destabilize the amino acid interactions and hydrogen bonding networks of ADIPOR1. PMID:27294143

  9. Impact of life history traits on gene flow: A multispecies systematic review across oceanographic barriers in the Mediterranean Sea

    PubMed Central

    Rives, Borja; Schunter, Celia; Macpherson, Enrique

    2017-01-01

    Background Marine species can demonstrate strong genetic differentiation and population structure despite the hypothesis of open seas and high connectivity. Some suggested drivers causing the genetic breaks are oceanographic barriers and the species’ biology. We assessed the relevance of seven major oceanographic fronts on species connectivity while considering their dispersal capacity and life strategy. Methods We systematically reviewed the scientific articles reporting population genetic differentiation along the Mediterranean Sea and across the Atlantic-Mediterranean transition. We retained those considering at least one sampling locality at each side of an oceanographic front, and at least two localities with no-front between them to correctly assess the effect of the front. To estimate the impact of life history characteristics affecting connectivity we considered the planktonic larval duration (PLD) and adult life strategy. Results Oceanographic barriers in the Mediterranean Sea seem to reduce gene flow globally; however, this effect is not homogeneous considering the life history traits of the species. The effect of the oceanographic fronts reduces gene flow in highly mobile species with PLD larger than 2–4 weeks. Benthic sessile species and/or with short PLD (< 2 weeks) have more significant genetic breaks between localities than species with higher motility; however, genetic differentiation occurs independently of the presence of a front. Conclusion Genetic connectivity is important for populations to recover from anthropogenic or natural impacts. We show that species with low mobility, mostly habitat-formers, have high genetic differentiation but low gene flow reduction mediated by the front, therefore, considering the importance of these species, we emphasize the vulnerability of the Mediterranean ecosystems and the necessity of protection strategies based on the whole ecosystem. PMID:28489878

  10. Impact of life history traits on gene flow: A multispecies systematic review across oceanographic barriers in the Mediterranean Sea.

    PubMed

    Pascual, Marta; Rives, Borja; Schunter, Celia; Macpherson, Enrique

    2017-01-01

    Marine species can demonstrate strong genetic differentiation and population structure despite the hypothesis of open seas and high connectivity. Some suggested drivers causing the genetic breaks are oceanographic barriers and the species' biology. We assessed the relevance of seven major oceanographic fronts on species connectivity while considering their dispersal capacity and life strategy. We systematically reviewed the scientific articles reporting population genetic differentiation along the Mediterranean Sea and across the Atlantic-Mediterranean transition. We retained those considering at least one sampling locality at each side of an oceanographic front, and at least two localities with no-front between them to correctly assess the effect of the front. To estimate the impact of life history characteristics affecting connectivity we considered the planktonic larval duration (PLD) and adult life strategy. Oceanographic barriers in the Mediterranean Sea seem to reduce gene flow globally; however, this effect is not homogeneous considering the life history traits of the species. The effect of the oceanographic fronts reduces gene flow in highly mobile species with PLD larger than 2-4 weeks. Benthic sessile species and/or with short PLD (< 2 weeks) have more significant genetic breaks between localities than species with higher motility; however, genetic differentiation occurs independently of the presence of a front. Genetic connectivity is important for populations to recover from anthropogenic or natural impacts. We show that species with low mobility, mostly habitat-formers, have high genetic differentiation but low gene flow reduction mediated by the front, therefore, considering the importance of these species, we emphasize the vulnerability of the Mediterranean ecosystems and the necessity of protection strategies based on the whole ecosystem.

  11. Impact of diurnal temperature variation on grape berry development, proanthocyanidin accumulation, and the expression of flavonoid pathway genes

    PubMed Central

    Cohen, Seth D.; Tarara, Julie M.; Gambetta, Greg A.; Matthews, Mark A.; Kennedy, James A.

    2012-01-01

    Little is known about the impact of temperature on proanthocyanidin (PA) accumulation in grape skins, despite its significance in berry composition and wine quality. Field-grown grapes (cv. Merlot) were cooled during the day or heated at night by +/–8 °C, from fruit set to véraison in three seasons, to determine the effect of temperature on PA accumulation. Total PA content per berry varied only in one year, when PA content was highest in heated berries (1.46 mg berry−1) and lowest in cooled berries (0.97 mg berry−1). In two years, cooling berries resulted in a significant increase in the proportion of (–)-epigallocatechin as an extension subunit. In the third year, rates of berry development, PA accumulation, and the expression levels of several genes involved in flavonoid biosynthesis were assessed. Heating and cooling berries altered the initial rates of PA accumulation, which was correlated strongly with the expression of core genes in the flavonoid pathway. Both heating and cooling altered the rate of berry growth and coloration, and the expression of several structural genes within the flavonoid pathway. PMID:22268158

  12. The impact of ICAM1 and VCAM1 gene polymorphisms on chronic allograft nephropathy and transplanted kidney function.

    PubMed

    Kłoda, K; Domański, L; Pawlik, A; Wiśniewska, M; Safranow, K; Ciechanowski, K

    2013-01-01

    ICAM-1 and VCAM-1 adhesion molecules play important roles in the immune response and emergence of chronic allograft nephropathy (CAN). The several polymorphisms of ICAM1 and VCAM1 genes are associated with changes in molecular expression therefore affecting allograft function and immune responses after kidney transplantation. The aim of this study was to examine the impact of polymorphisms in ICAM1 and VCAM1 genes on biopsy-proven CAN and renal allograft function. The 270 Caucasian renal transplant recipients (166 men and 104 women) were genotyped for the rs5498 ICAM1 and rs1041163 and rs3170794 VCAM1 gene polymorphisms using real-time polymerase chain reaction. There was no correlation between polymorphisms and CAN. Creatinine concentrations in the first month after transplantation differed between the rs5498 ICAM1 genotypes (P = .095), being higher for GG carriers (AA + AG vs GG, P =.07) albeit not with statistical significance. Creatinine concentrations at 12, 24, and 36 months after transplantation differed significantly among rs5498 ICAM1 genotypes (P = .0046, P =.016, and P = .02) and were higher among GG carriers (AA + AG vs GG, P = .001, P = .004, and P = .006). Rs5498 ICAM1 GG genotype and receipient male gender were independent factors associated with higher creatinine concentrations. These results suggest that the rs5498 ICAM1 GG genotype may be associated with long-term allograft function. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Impact of murine intestinal apolipoprotein A-IV expression on regional lipid absorption, gene expression, and growth

    PubMed Central

    Simon, Trang; Cook, Victoria R.; Rao, Anuradha; Weinberg, Richard B.

    2011-01-01

    Apolipoprotein A-IV (apoA-IV) is synthesized by intestinal enterocytes during lipid absorption and secreted into lymph on the surface of nascent chylomicrons. A compelling body of evidence supports a central role of apoA-IV in facilitating intestinal lipid absorption and in regulating satiety, yet a longstanding conundrum is that no abnormalities in fat absorption, feeding behavior, or weight gain were observed in chow-fed apoA-IV knockout (A4KO) mice. Herein we reevaluated the impact of apoA-IV expression in C57BL6 and A4KO mice fed a high-fat diet. Fat balance and lymph cannulation studies found no effect of intestinal apoA-IV gene expression on the efficiency of fatty acid absorption, but gut sac transport studies revealed that apoA-IV differentially modulates lipid transport and the number and size of secreted triglyceride-rich lipoproteins in different anatomic regions of the small bowel. ApoA-IV gene deletion increased expression of other genes involved in chylomicron assembly, impaired the ability of A4KO mice to gain weight and increase adipose tissue mass, and increased the distal gut hormone response to a high-fat diet. Together these findings suggest that apoA-IV may play a unique role in integrating feeding behavior, intestinal lipid absorption, and energy storage. PMID:21840868

  14. Nutritional impacts on gene expression in the surface mucosa of blue catfish (Ictalurus furcatus)

    USDA-ARS?s Scientific Manuscript database

    Short-term feed deprivation is a common occurrence in both wild and farmed fish species, due to reproductive processes, seasonal variations in temperature, or in response to a disease outbreak. Fasting can have dramatic physiological and biological onsequences for fish, including impacts on mucosal ...

  15. Structure and transcriptional impact of divergent repetitive elements inserted within Phanerochaete chrysosporium strain RP-78 genes

    Treesearch

    Luis F. Larrondo; Paulo Canessa; Rafael Vicuna; Philip Stewart; Amber Vanden Wymelenberg; Dan Cullen

    2007-01-01

    We describe the structure, organization, and transcriptional impact of repetitive elements within the lignin-degrading basidiomycete, Phanerochaete chrysosporium. Searches of the P. chrysosporium genome revealed five copies of pce1, a 1,750-nt non-autonomous, class II element. Alleles encoding a putative glucosyltransferase and a cytochrome P450 harbor pce insertions...

  16. Impact of duplicate gene copies on phylogenetic analysis and divergence time estimates in butterflies

    PubMed Central

    Pohl, Nélida; Sison-Mangus, Marilou P; Yee, Emily N; Liswi, Saif W; Briscoe, Adriana D

    2009-01-01

    Background The increase in availability of genomic sequences for a wide range of organisms has revealed gene duplication to be a relatively common event. Encounters with duplicate gene copies have consequently become almost inevitable in the context of collecting gene sequences for inferring species trees. Here we examine the effect of incorporating duplicate gene copies evolving at different rates on tree reconstruction and time estimation of recent and deep divergences in butterflies. Results Sequences from ultraviolet-sensitive (UVRh), blue-sensitive (BRh), and long-wavelength sensitive (LWRh) opsins,EF-1α and COI were obtained from 27 taxa representing the five major butterfly families (5535 bp total). Both BRh and LWRh are present in multiple copies in some butterfly lineages and the different copies evolve at different rates. Regardless of the phylogenetic reconstruction method used, we found that analyses of combined data sets using either slower or faster evolving copies of duplicate genes resulted in a single topology in agreement with our current understanding of butterfly family relationships based on morphology and molecules. Interestingly, individual analyses of BRh and LWRh sequences also recovered these family-level relationships. Two different relaxed clock methods resulted in similar divergence time estimates at the shallower nodes in the tree, regardless of whether faster or slower evolving copies were used, with larger discrepancies observed at deeper nodes in the phylogeny. The time of divergence between the monarch butterfly Danaus plexippus and the queen D. gilippus (15.3–35.6 Mya) was found to be much older than the time of divergence between monarch co-mimic Limenitis archippus and red-spotted purple L. arthemis (4.7–13.6 Mya), and overlapping with the time of divergence of the co-mimetic passionflower butterflies Heliconius erato and H. melpomene (13.5–26.1 Mya). Our family-level results are congruent with recent estimates found in

  17. Impact of duplicate gene copies on phylogenetic analysis and divergence time estimates in butterflies.

    PubMed

    Pohl, Nélida; Sison-Mangus, Marilou P; Yee, Emily N; Liswi, Saif W; Briscoe, Adriana D

    2009-05-13

    The increase in availability of genomic sequences for a wide range of organisms has revealed gene duplication to be a relatively common event. Encounters with duplicate gene copies have consequently become almost inevitable in the context of collecting gene sequences for inferring species trees. Here we examine the effect of incorporating duplicate gene copies evolving at different rates on tree reconstruction and time estimation of recent and deep divergences in butterflies. Sequences from ultraviolet-sensitive (UVRh), blue-sensitive (BRh), and long-wavelength sensitive (LWRh) opsins,EF-1 and COI were obtained from 27 taxa representing the five major butterfly families (5535 bp total). Both BRh and LWRh are present in multiple copies in some butterfly lineages and the different copies evolve at different rates. Regardless of the phylogenetic reconstruction method used, we found that analyses of combined data sets using either slower or faster evolving copies of duplicate genes resulted in a single topology in agreement with our current understanding of butterfly family relationships based on morphology and molecules. Interestingly, individual analyses of BRh and LWRh sequences also recovered these family-level relationships. Two different relaxed clock methods resulted in similar divergence time estimates at the shallower nodes in the tree, regardless of whether faster or slower evolving copies were used, with larger discrepancies observed at deeper nodes in the phylogeny. The time of divergence between the monarch butterfly Danaus plexippus and the queen D. gilippus (15.3-35.6 Mya) was found to be much older than the time of divergence between monarch co-mimic Limenitis archippus and red-spotted purple L. arthemis (4.7-13.6 Mya), and overlapping with the time of divergence of the co-mimetic passionflower butterflies Heliconius erato and H. melpomene (13.5-26.1 Mya). Our family-level results are congruent with recent estimates found in the literature and indicate

  18. Impact of XPD gene polymorphism on risk of prostate cancer on north Indian population.

    PubMed

    Sobti, Ranbir Chander; Berhane, Nega; Melese, Shiferaw; Mahdi, Salih Abdul; Gupta, Libsy; Thakur, Hitender; Singh, Neha

    2012-03-01

    Prostate cancer is the second most diagnosed cancer in men next to skin cancer in the developed world. Risk of disease varies most prominently with age, ethnicity, family history, and diet. Genetic polymorphism of some genes has been implicated in increasing the risk. The XPD (Xeroderma pigmentosum group D) gene codes for a DNA helicase involved in transcription and nucleotide excision repair. The aim of this study is to evaluate the effect of XPD 751 Lys/Gln polymorphism on risk of prostate cancer on north Indian patients. Blood sample from 150 prostate cancer patients, 150 from Prostate Hyper Plasia and equal number of samples from healthy control groups was collected from North India. The polymerase chain reaction and restrictive fragment length polymorphism techniques were implemented. Statistically non-significant increase risk of prostate cancer was observed with patients having Gln/Gln genotype (OR 1.62, 95% CI).

  19. Impact of diethylhexyl phthalate on gene expression and development of mammary glands of pregnant mouse.

    PubMed

    Li, Lan; Liu, Jing-Cai; Zhao, Yong; Lai, Fang-Nong; Yang, Fan; Ge, Wei; Dou, Cheng-Li; Shen, Wei; Zhang, Xi-Feng; Chen, Hong

    2015-10-01

    The widely used diethylhexyl phthalate (DEHP) is a known endocrine disruptor that causes persistent alterations in the structure and function of female reproductive system, including ovaries, uterus and oviducts. To explore the molecular mechanism of the effect of DEHP on the development of mammary glands, we investigated the cell cycle, growth, proliferation and gene expression of mammary gland cells of pregnant mice exposed to DEHP. It was demonstrated, for the first time, that the mammary gland cells of pregnant mice treated with DEHP for 0.5-3.5 days post-coitum had increased proliferation, growth rate and number of cells in the G2/S phase. The expression of cell proliferation-related genes was significantly altered after short time and low-dose DEHP treatment of mammary gland cells in vivo and in vitro. These findings showed adverse effects of DEHP on mammary gland cells in pregnant mice.

  20. Heme Signaling Impacts Global Gene Expression, Immunity and Dengue Virus Infectivity in Aedes aegypti

    PubMed Central

    Bottino-Rojas, Vanessa; Talyuli, Octávio A. C.; Jupatanakul, Natapong; Sim, Shuzhen; Dimopoulos, George; Venancio, Thiago M.; Bahia, Ana C.; Sorgine, Marcos H.; Oliveira, Pedro L.; Paiva-Silva, Gabriela O.

    2015-01-01

    Blood-feeding mosquitoes are exposed to high levels of heme, the product of hemoglobin degradation. Heme is a pro-oxidant that influences a variety of cellular processes. We performed a global analysis of heme-regulated Aedes aegypti (yellow fever mosquito) transcriptional changes to better understand influence on mosquito physiology at the molecular level. We observed an iron- and reactive oxygen species (ROS)-independent signaling induced by heme that comprised genes related to redox metabolism. By modulating the abundance of these transcripts, heme possibly acts as a danger signaling molecule. Furthermore, heme triggered critical changes in the expression of energy metabolism and immune response genes, altering the susceptibility towards bacteria and dengue virus. These findings seem to have implications on the adaptation of mosquitoes to hematophagy and consequently on their ability to transmit diseases. Altogether, these results may also contribute to the understanding of heme cell biology in eukaryotic cells. PMID:26275150

  1. The Impact of Hypergravity and Vibration on Gene and Protein Expression of Thyroid Cells

    NASA Astrophysics Data System (ADS)

    Wehland, Markus; Warnke, Elisabeth; Frett, Timo; Hemmersbach, Ruth; Hauslage, Jens; Ma, Xiao; Aleshcheva, Ganna; Pietsch, Jessica; Bauer, Johann; Grimm, Daniela

    2016-06-01

    Experiments in space either on orbital missions on-board the ISS, or in suborbital missions using sounding rockets, like TEXUS as well as parabolic flight campaigns are still the gold standard to achieve real microgravity conditions in the field of gravitational biology and medicine. However, during launch, and in flight, hypergravity and vibrations occur which might interfere with the effects of microgravity. It is therefore important to know these effects and discriminate them from the microgravity effects. This can be achieved by ground-based facilities like centrifuges or vibration platforms. Recently, we have conducted several experiments with different thyroid cancer cell lines. This study, as part of the ESA-CORA-GBF 2010-203 project, focused on the influence of vibration and hypergravity on benign human thyroid follicular epithelial cells (Nthy-ori 3-1 cell line). Gene and in part protein expression regulation under both conditions were analyzed for VCAN, ITGA10, ITGB1, OPN, ADAM19, ANXA1, TNFA, ABL2, ACTB, PFN2, TLN1, EZR, RDX, MSN, CTGF, PRKCA, and PRKAA1 using quantitative real-time PCR and Western Blot. We found that hypergravity and vibration affected genes and proteins involved in the extracellular matrix, the cytoskeleton, apoptosis, cell growth and signaling. Vibration always led to a down-regulation, whereas hypergravity resulted in a more heterogeneous expression pattern. Overall we conclude that both conditions can influence gene regulation and production of various genes and proteins. As a consequence, it is important to perform control experiments on hypergravity and vibration facilities in parallel to flight experiments.

  2. Interactions of early adversity with stress-related gene polymorphisms impact regional brain structure in females

    PubMed Central

    Gupta, Arpana; Labus, Jennifer; Kilpatrick, Lisa A.; Bonyadi, Mariam; Ashe-McNalley, Cody; Heendeniya, Nuwanthi; Bradesi, Sylvie; Chang, Lin; Mayer, Emeran A.

    2015-01-01

    Early adverse life events (EALs) have been associated with regional thinning of the subgenual cingulate cortex (sgACC), a brain region implicated in the development of disorders of mood and affect, and often comorbid functional pain disorders, such as irritable bowel syndrome (IBS). Regional neuroinflammation related to chronic stress system activation has been suggested as a possible mechanism underlying these neuroplastic changes. However, the interaction of genetic and environmental factors in these changes is poorly understood. The current study aimed to evaluate the interactions of EALs and candidate gene polymorphisms in influencing thickness of the sgACC. 210 female subjects (137 healthy controls; 73 IBS) were genotyped for stress and inflammation-related gene polymorphisms. Genetic variation with EALs, and diagnosis on sgACC thickness was examined, while controlling for race, age, and total brain volume. Compared to HCs, IBS had significantly reduced sgACC thickness (p = 0.03). Regardless of disease group (IBS vs. HC), thinning of the left sgACC was associated with a significant gene-gene environment interaction between the IL-1β genotype, the NR3C1 haplotype, and a history of EALs (p = 0.05). Reduced sgACC thickness in women with the minor IL-1β allele, was associated with EAL total scores regardless of NR3C1 haplotype status (p = 0.02). In subjects homozygous for the major IL-1β allele, reduced sgACC with increasing levels of EALs was seen only with the less common NR3C1 haplotype (p = 0.02). These findings support an interaction between polymorphisms related to stress and inflammation and early adverse life events in modulating a key region of the emotion arousal circuit. PMID:25630611

  3. Contrasting nitrogen fertilization treatments impact xylem gene expression and secondary cell wall lignification in Eucalyptus.

    PubMed

    Camargo, Eduardo Leal Oliveira; Nascimento, Leandro Costa; Soler, Marçal; Salazar, Marcela Mendes; Lepikson-Neto, Jorge; Marques, Wesley Leoricy; Alves, Ana; Teixeira, Paulo José Pereira Lima; Mieczkowski, Piotr; Carazzolle, Marcelo Falsarella; Martinez, Yves; Deckmann, Ana Carolina; Rodrigues, José Carlos; Grima-Pettenati, Jacqueline; Pereira, Gonçalo Amarante Guimarães

    2014-09-28

    Nitrogen (N) is a main nutrient required for tree growth and biomass accumulation. In this study, we analyzed the effects of contrasting nitrogen fertilization treatments on the phenotypes of fast growing Eucalyptus hybrids (E. urophylla x E. grandis) with a special focus on xylem secondary cell walls and global gene expression patterns. Histological observations of the xylem secondary cell walls further confirmed by chemical analyses showed that lignin was reduced by luxuriant fertilization, whereas a consistent lignin deposition was observed in trees grown in N-limiting conditions. Also, the syringyl/guaiacyl (S/G) ratio was significantly lower in luxuriant nitrogen samples. Deep sequencing RNAseq analyses allowed us to identify a high number of differentially expressed genes (1,469) between contrasting N treatments. This number is dramatically higher than those obtained in similar studies performed in poplar but using microarrays. Remarkably, all the genes involved the general phenylpropanoid metabolism and lignin pathway were found to be down-regulated in response to high N availability. These findings further confirmed by RT-qPCR are in agreement with the reduced amount of lignin in xylem secondary cell walls of these plants. This work enabled us to identify, at the whole genome level, xylem genes differentially regulated by N availability, some of which are involved in the environmental control of xylogenesis. It further illustrates that N fertilization can be used to alter the quantity and quality of lignocellulosic biomass in Eucalyptus, offering exciting prospects for the pulp and paper industry and for the use of short coppices plantations to produce second generation biofuels.

  4. Impact of Intramammary Treatment on Gene Expression Profiles in Bovine Escherichia coli Mastitis

    PubMed Central

    Sipka, Anja; Klaessig, Suzanne; Duhamel, Gerald E.; Swinkels, Jantijn; Rainard, Pascal; Schukken, Ynte

    2014-01-01

    Clinical mastitis caused by E. coli accounts for significant production losses and animal welfare concerns on dairy farms worldwide. The benefits of therapeutic intervention in mild to moderate cases are incompletely understood. We investigated the effect of intramammary treatment with cefapirin alone or in combination with prednisolone on gene expression profiles in experimentally-induced E. coli mastitis in six mid-lactating Holstein Friesian cows. Cows were challenged with E. coli in 3 quarters and received 4 doses of 300 mg cefapirin in one quarter and 4 doses of 300 mg cefapirin together with 20 mg prednisolone in another quarter. At 24 h (n = 3) or 48 h (n = 3) post-challenge, tissue samples from control and treated quarters were collected for microarray analysis. Gene expression analysis of challenged, un-treated quarters revealed an up-regulation of transcripts associated with immune response functions compared to un-challenged quarters. Both treatments resulted in down-regulation of these transcripts compared to challenged, un-treated quarters most prominently for genes representing Chemokine and TLR-signaling pathways. Gene expression of Lipopolysaccharide Binding Protein (LBP), CCL2 and CXCL2 were only significantly down-regulated in cefapirin-prednisolone-treated quarters compared to un-treated controls. Down-regulation of chemokines was further confirmed on the basis of protein levels in milk whey for CXCL1, CXCL2 and CXCL8 in both treatments with a greater decrease in cefapirin-prednisolone-treated quarters. The data reveal a significant effect of treatment on cell recruitment with a more pronounced effect in cefapirin-prednisolone treated quarters. Provided a rapid bacteriological clearance, combination therapy may prevent neutrophil-induced tissue damage and promote recovery of the gland. PMID:24454893

  5. Gene therapy of hepatocarcinoma: a long way from the concept to the therapeutical impact.

    PubMed

    Gérolami, René; Uch, Rathviro; Bréchot, Christian; Mannoni, Patrice; Bagnis, Claude

    2003-09-01

    Hepatocellular carcinoma (HCC), the most prevalent histological form of primary liver cancer is one of the most frequent cancer worldwide. This pathology still requires the development of new therapeutical approaches. Gene therapy strategies focusing on the genetic manipulation of accessory cells involved in the immune reaction against cancer cells, or on the direct transduction of tumor cells with transgenes able to "suicide" cancer cells have been largely developed for more than ten years.

  6. Meiotic Drive Impacts Expression and Evolution of X-Linked Genes in Stalk-Eyed Flies

    PubMed Central

    Reinhardt, Josephine A.; Brand, Cara L.; Paczolt, Kimberly A.; Johns, Philip M.; Baker, Richard H.; Wilkinson, Gerald S.

    2014-01-01

    Although sex chromosome meiotic drive has been observed in a variety of species for over 50 years, the genes causing drive are only known in a few cases, and none of these cases cause distorted sex-ratios in nature. In stalk-eyed flies (Teleopsis dalmanni), driving X chromosomes are commonly found at frequencies approaching 30% in the wild, but the genetic basis of drive has remained elusive due to reduced recombination between driving and non-driving X chromosomes. Here, we used RNAseq to identify transcripts that are differentially expressed between males carrying either a driving X (XSR) or a standard X chromosome (XST), and found hundreds of these, the majority of which are X-linked. Drive-associated transcripts show increased levels of sequence divergence (dN/dS) compared to a control set, and are predominantly expressed either in testes or in the gonads of both sexes. Finally, we confirmed that XSR and XST are highly divergent by estimating sequence differentiation between the RNAseq pools. We found that X-linked transcripts were often strongly differentiated (whereas most autosomal transcripts were not), supporting the presence of a relatively large region of recombination suppression on XSR presumably caused by one or more inversions. We have identified a group of genes that are good candidates for further study into the causes and consequences of sex-chromosome drive, and demonstrated that meiotic drive has had a profound effect on sequence evolution and gene expression of X-linked genes in this species. PMID:24832132

  7. High-Resolution Gene Flow Model for Assessing Environmental Impacts of Transgene Escape Based on Biological Parameters and Wind Speed.

    PubMed

    Wang, Lei; Haccou, Patsy; Lu, Bao-Rong

    2016-01-01

    Environmental impacts caused by transgene flow from genetically engineered (GE) crops to their wild relatives mediated by pollination are longstanding biosafety concerns worldwide. Mathematical modeling provides a useful tool for estimating frequencies of pollen-mediated gene flow (PMGF) that are critical for assessing such environmental impacts. However, most PMGF models are impractical for this purpose because their parameterization requires actual data from field experiments. In addition, most of these models are usually too general and ignored the important biological characteristics of concerned plant species; and therefore cannot provide accurate prediction for PMGF frequencies. It is necessary to develop more accurate PMGF models based on biological and climatic parameters that can be easily measured in situ. Here, we present a quasi-mechanistic PMGF model that only requires the input of biological and wind speed parameters without actual data from field experiments. Validation of the quasi-mechanistic model based on five sets of published data from field experiments showed significant correlations between the model-simulated and field experimental-generated PMGF frequencies. These results suggest accurate prediction for PMGF frequencies using this model, provided that the necessary biological parameters and wind speed data are available. This model can largely facilitate the assessment and management of environmental impacts caused by transgene flow, such as determining transgene flow frequencies at a particular spatial distance, and establishing spatial isolation between a GE crop and its coexisting non-GE counterparts and wild relatives.

  8. High-Resolution Gene Flow Model for Assessing Environmental Impacts of Transgene Escape Based on Biological Parameters and Wind Speed

    PubMed Central

    Wang, Lei; Haccou, Patsy; Lu, Bao-Rong

    2016-01-01

    Environmental impacts caused by transgene flow from genetically engineered (GE) crops to their wild relatives mediated by pollination are longstanding biosafety concerns worldwide. Mathematical modeling provides a useful tool for estimating frequencies of pollen-mediated gene flow (PMGF) that are critical for assessing such environmental impacts. However, most PMGF models are impractical for this purpose because their parameterization requires actual data from field experiments. In addition, most of these models are usually too general and ignored the important biological characteristics of concerned plant species; and therefore cannot provide accurate prediction for PMGF frequencies. It is necessary to develop more accurate PMGF models based on biological and climatic parameters that can be easily measured in situ. Here, we present a quasi-mechanistic PMGF model that only requires the input of biological and wind speed parameters without actual data from field experiments. Validation of the quasi-mechanistic model based on five sets of published data from field experiments showed significant correlations between the model-simulated and field experimental-generated PMGF frequencies. These results suggest accurate prediction for PMGF frequencies using this model, provided that the necessary biological parameters and wind speed data are available. This model can largely facilitate the assessment and management of environmental impacts caused by transgene flow, such as determining transgene flow frequencies at a particular spatial distance, and establishing spatial isolation between a GE crop and its coexisting non-GE counterparts and wild relatives. PMID:26959240

  9. Many amino acid substitution variants identified in DNA repair genes during human population screenings are predicted to impact protein function

    SciTech Connect

    Xi, T; Jones, I M; Mohrenweiser, H W

    2003-11-03

    Over 520 different amino acid substitution variants have been previously identified in the systematic screening of 91 human DNA repair genes for sequence variation. Two algorithms were employed to predict the impact of these amino acid substitutions on protein activity. Sorting Intolerant From Tolerant (SIFT) classified 226 of 508 variants (44%) as ''Intolerant''. Polymorphism Phenotyping (PolyPhen) classed 165 of 489 amino acid substitutions (34%) as ''Probably or Possibly Damaging''. Another 9-15% of the variants were classed as ''Potentially Intolerant or Damaging''. The results from the two algorithms are highly associated, with concordance in predicted impact observed for {approx}62% of the variants. Twenty one to thirty one percent of the variant proteins are predicted to exhibit reduced activity by both algorithms. These variants occur at slightly lower individual allele frequency than do the variants classified as ''Tolerant'' or ''Benign''. Both algorithms correctly predicted the impact of 26 functionally characterized amino acid substitutions in the APE1 protein on biochemical activity, with one exception. It is concluded that a substantial fraction of the missense variants observed in the general human population are functionally relevant. These variants are expected to be the molecular genetic and biochemical basis for the associations of reduced DNA repair capacity phenotypes with elevated cancer risk.

  10. Impact of Gene Patents and Licensing Practices on Access to Genetic Testing for Long QT Syndrome

    PubMed Central

    Angrist, Misha; Chandrasekharan, Subhashini; Heaney, Christopher; Cook-Deegan, Robert

    2010-01-01

    Genetic testing for Long QT syndrome (LQTS) exemplifies patenting and exclusive licensing with different outcomes at different times. Exclusive licensing from the University of Utah changed the business model from sole provider to two US providers of LQTS testing. LQTS is associated with mutations in many genes, ten of which are now tested by two competing firms in the United States, PGxHealth and GeneDx. Until 2009, PGxHealth was sole provider, based largely on exclusive rights to patents from the University of Utah and other academic institutions. University of Utah patents were initially licensed to DNA Sciences, whose patent rights were acquired by Gennaissance, and then by Clinical Data, Inc., which owns PGxHealth. In 2002, DNA Sciences “cleared the market” by sending cease and desist patent enforcement letters to university and reference laboratories offering LQTS genetic testing. There was no test on the market for a one- to two-year period. From 2005-2008, most LQTS-related patents were controlled by Clinical Data, Inc., and its subsidiary PGxHealth. BioReference Laboratories, Inc., secured countervailing exclusive patent rights starting in 2006, also from the University of Utah, and broke the PGxHealth monopoly in early 2009, creating a duopoly for genetic testing in the United States, and expanding the number of genes for which commercial testing is available from five to ten. PMID:20393304

  11. Predicting the impact of deleterious single point mutations in SMAD gene family using structural bioinformatics approach.

    PubMed

    George Priya Doss, C; Nagasundaram, N; Tanwar, Himani

    2012-06-01

    Functional alteration in SMAD proteins leads to dis-regulation of its mechanism results in possibilities of high risk diseases like fibrosis, cancer, juvenile polyposis etc. Studying single nucleotide polymorphism (SNP) in SMAD genes helps understand the malfunction of these proteins. In this study, we focused on deleterious effects of nsSNPs in both structural and functional level using publically available bioinformatics tools. We have mainly focused on identifying deleterious nsSNPs in both structural and functional level in SMAD genes by using SIFT, PolyPhen, SNPs&GO, I-Mutant 3.0, MUpro and PANTHER. Structure analysis was carried out with the major mutation that occurred in the native protein coded by SMAD genes and its amino acid positions (R358W, K306S, R310G, S433R and R361C). SRide was used to check the stability of the native and mutant modelled proteins. In addition, we used MAPPER to identify SNPs present in transcription factor binding sites. These findings demonstrate that the in silico approaches can be used efficiently to identify potential candidate SNPs in large scale analysis.

  12. Impact of chromatin structure and dynamics on PR signaling. The initial steps in hormonal gene regulation.

    PubMed

    Beato, Miguel; Vicent, Guillermo P

    2012-06-24

    Gene regulation requires access of transcription factors to DNA sequences of target genes, which is limited by the compaction of DNA in chromatin. Based on our studies on the Progesterone receptor (PR)-dependent hormonal induction of mouse mammary tumor virus (MMTV) promoter we found that remodeling of the various levels of chromatin organization is a complex and necessary prerequisite for regulation. Two consecutive cycles are essential for transcriptional activation, both involving the collaboration between activated protein kinases, histone modifying enzymes and ATP-dependent chromatin remodelers. The first cycle ends with the displacement of histone H1 and decompaction of higher order chromatin structure. The second cycle leads to the displacement of dimers of histones H2A and H2B resulting in opening of nucleosomes. In both cases the hormone receptor recruits an ATP-dependent chromatin remodeler, whose binding to chromatin is stabilized by distinct histone modifications. The final result is to facilitate full occupancy of the cis regulatory sites and access for the basal transcription machinery. Thus, activation of PR-target genes involves a very rapid coordination of enzymatic activities via crosstalk with various kinase-signaling pathways. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  13. Quercetin Impacts Expression of Metabolism- and Obesity-Associated Genes in SGBS Adipocytes.

    PubMed

    Leiherer, Andreas; Stoemmer, Kathrin; Muendlein, Axel; Saely, Christoph H; Kinz, Elena; Brandtner, Eva M; Fraunberger, Peter; Drexel, Heinz

    2016-05-12

    Obesity is characterized by the rapid expansion of visceral adipose tissue, resulting in a hypoxic environment in adipose tissue which leads to a profound change of gene expression in adipocytes. As a consequence, there is a dysregulation of metabolism and adipokine secretion in adipose tissue leading to the development of systemic inflammation and finally resulting in the onset of metabolic diseases. The flavonoid quercetin as well as other secondary plant metabolites also referred to as phytochemicals have anti-oxidant, anti-inflammatory, and anti-diabetic effects known to be protective in view of obesity-related-diseases. Nevertheless, its underlying molecular mechanism is still obscure and thus the focus of this study was to explore the influence of quercetin on human SGBS (Simpson Golabi Behmel Syndrome) adipocytes' gene expression. We revealed for the first time that quercetin significantly changed expression of adipokine (Angptl4, adipsin, irisin and PAI-1) and glycolysis-involved (ENO2, PFKP and PFKFB4) genes, and that this effect not only antagonized but in part even overcompensated the effect mediated by hypoxia in adipocytes. Thus, these results are explained by the recently proposed hypothesis that the protective effect of quercetin is not solely due to its free radical-scavenging activity but also to a direct effect on mitochondrial processes, and they demonstrate that quercetin might have the potential to counteract the development of obesity-associated complications.

  14. Basic concepts of epigenetics: impact of environmental signals on gene expression.

    PubMed

    Mazzio, Elizabeth A; Soliman, Karam F A

    2012-02-01

    Through epigenetic modifications, specific long-term phenotypic consequences can arise from environmental influence on slowly evolving genomic DNA. Heritable epigenetic information regulates nucleosomal arrangement around DNA and determines patterns of gene silencing or active transcription. One of the greatest challenges in the study of epigenetics as it relates to disease is the enormous diversity of proteins, histone modifications and DNA methylation patterns associated with each unique maladaptive phenotype. This is further complicated by a limitless combination of environmental cues that could alter the epigenome of specific cell types, tissues, organs and systems. In addition, complexities arise from the interpretation of studies describing analogous but not identical processes in flies, plants, worms, yeast, ciliated protozoans, tumor cells and mammals. This review integrates fundamental basic concepts of epigenetics with specific focus on how the epigenetic machinery interacts and operates in continuity to silence or activate gene expression. Topics covered include the connection between DNA methylation, methyl-CpG-binding proteins, transcriptional repression complexes, histone residues, histone modifications that mediate gene repression or relaxation, histone core variant stability, H1 histone linker flexibility, FACT complex, nucleosomal remodeling complexes, HP1 and nuclear lamins.

  15. Impact of single nucleotide polymorphisms in HBB gene causing haemoglobinopathies: in silico analysis.

    PubMed

    George Priya Doss, C; Rao, Sethumadhavan

    2009-04-01

    Single nucleotide polymorphisms (SNPs) are being intensively studied to understand the biological basis of complex traits and diseases. Deleterious mutations of the human beta-globin gene (HBB) are responsible for beta-thalassaemia and other haemoglobinopathies, which are the most common genetic diseases of blood. Single amino acid substitutions in the globin chain are the commonest forms of haemoglobinopathy. Although many haemoglobinopathies present similar structural abnormal points, their functions sometimes are different. Here, using computational methods, we analysed the genetic variations that can alter the expression and function of the HBB gene. We applied an evolutionary perspective to screen the SNPs using a sequence homology-based SIFT tool, which suggested that 210 (90%) non-synonymous (ns)SNPs were found to be deleterious. The structure-based approach PolyPhen server suggested that 134 (57%) nsSNPS may disrupt protein function and structure. The PupaSuite tool predicted the phenotypic effect of SNPs on the structure and function of the affected protein. Structure analysis was carried out with the major mutation that occurred in the native protein coded by the HBB gene in HbC, HbD, HbE and HbS. The amino acid residues in the native and mutant modelled protein were further analysed for solvent accessibility, and secondary structure to check the stability of the proteins. The functional analysis presented here may be a good model for further research.

  16. Quercetin Impacts Expression of Metabolism- and Obesity-Associated Genes in SGBS Adipocytes

    PubMed Central

    Leiherer, Andreas; Stoemmer, Kathrin; Muendlein, Axel; Saely, Christoph H.; Kinz, Elena; Brandtner, Eva M.; Fraunberger, Peter; Drexel, Heinz

    2016-01-01

    Obesity is characterized by the rapid expansion of visceral adipose tissue, resulting in a hypoxic environment in adipose tissue which leads to a profound change of gene expression in adipocytes. As a consequence, there is a dysregulation of metabolism and adipokine secretion in adipose tissue leading to the development of systemic inflammation and finally resulting in the onset of metabolic diseases. The flavonoid quercetin as well as other secondary plant metabolites also referred to as phytochemicals have anti-oxidant, anti-inflammatory, and anti-diabetic effects known to be protective in view of obesity-related-diseases. Nevertheless, its underlying molecular mechanism is still obscure and thus the focus of this study was to explore the influence of quercetin on human SGBS (Simpson Golabi Behmel Syndrome) adipocytes’ gene expression. We revealed for the first time that quercetin significantly changed expression of adipokine (Angptl4, adipsin, irisin and PAI-1) and glycolysis-involved (ENO2, PFKP and PFKFB4) genes, and that this effect not only antagonized but in part even overcompensated the effect mediated by hypoxia in adipocytes. Thus, these results are explained by the recently proposed hypothesis that the protective effect of quercetin is not solely due to its free radical-scavenging activity but also to a direct effect on mitochondrial processes, and they demonstrate that quercetin might have the potential to counteract the development of obesity-associated complications. PMID:27187453

  17. γ-Resorcylate Catabolic-Pathway Genes in the Soil Actinomycete Rhodococcus jostii RHA1

    PubMed Central

    Kasai, Daisuke; Araki, Naoto; Motoi, Kota; Yoshikawa, Shota; Iino, Toju; Imai, Shunsuke; Masai, Eiji

    2015-01-01

    The Rhodococcus jostii RHA1 gene cluster required for γ-resorcylate (GRA) catabolism was characterized. The cluster includes tsdA, tsdB, tsdC, tsdD, tsdR, tsdT, and tsdX, which encode GRA decarboxylase, resorcinol 4-hydroxylase, hydroxyquinol 1,2-dioxygenase, maleylacetate reductase, an IclR-type regulator, a major facilitator superfamily transporter, and a putative hydrolase, respectively. The tsdA gene conferred GRA decarboxylase activity on Escherichia coli. Purified TsdB oxidized NADH in the presence of resorcinol, suggesting that tsdB encodes a unique NADH-specific single-component resorcinol 4-hydroxylase. Mutations in either tsdA or tsdB resulted in growth deficiency on GRA. The tsdC and tsdD genes conferred hydroxyquinol 1,2-dioxygenase and maleylacetate reductase activities, respectively, on E. coli. Inactivation of tsdT significantly retarded the growth of RHA1 on GRA. The growth retardation was partially suppressed under acidic conditions, suggesting the involvement of tsdT in GRA uptake. Reverse transcription-PCR analysis revealed that the tsd genes constitute three transcriptional units, the tsdBADC and tsdTX operons and tsdR. Transcription of the tsdBADC and tsdTX operons was induced during growth on GRA. Inactivation of tsdR derepressed transcription of the tsdBADC and tsdTX operons in the absence of GRA, suggesting that tsd gene transcription is negatively regulated by the tsdR-encoded regulator. Binding of TsdR to the tsdR-tsdB and tsdT-tsdR intergenic regions was inhibited by the addition of GRA, indicating that GRA interacts with TsdR as an effector molecule. PMID:26319878

  18. DNA methylation in glioblastoma: impact on gene expression and clinical outcome.

    PubMed

    Etcheverry, Amandine; Aubry, Marc; de Tayrac, Marie; Vauleon, Elodie; Boniface, Rachel; Guenot, Frederique; Saikali, Stephan; Hamlat, Abderrahmane; Riffaud, Laurent; Menei, Philippe; Quillien, Veronique; Mosser, Jean

    2010-12-14

    Changes in promoter DNA methylation pattern of genes involved in key biological pathways have been reported in glioblastoma. Genome-wide assessments of DNA methylation levels are now required to decipher the epigenetic events involved in the aggressive phenotype of glioblastoma, and to guide new treatment strategies. We performed a whole-genome integrative analysis of methylation and gene expression profiles in 40 newly diagnosed glioblastoma patients. We also screened for associations between the level of methylation of CpG sites and overall survival in a cohort of 50 patients uniformly treated by surgery, radiotherapy and chemotherapy with concomitant and adjuvant temozolomide (STUPP protocol). The methylation analysis identified 616 CpG sites differentially methylated between glioblastoma and control brain, a quarter of which was differentially expressed in a concordant way. Thirteen of the genes with concordant CpG sites displayed an inverse correlation between promoter methylation and expression level in glioblastomas: B3GNT5, FABP7, ZNF217, BST2, OAS1, SLC13A5, GSTM5, ME1, UBXD3, TSPYL5, FAAH, C7orf13, and C3orf14. Survival analysis identified six CpG sites associated with overall survival. SOX10 promoter methylation status (two CpG sites) stratified patients similarly to MGMT status, but with a higher Area Under the Curve (0.78 vs. 0.71, p-value < 5e-04). The methylation status of the FNDC3B, TBX3, DGKI, and FSD1 promoters identified patients with MGMT-methylated tumors that did not respond to STUPP treatment (p-value < 1e-04). This study provides the first genome-wide integrative analysis of DNA methylation and gene expression profiles obtained from the same GBM cohort. We also present a methylome-based survival analysis for one of the largest uniformly treated GBM cohort ever studied, for more than 27,000 CpG sites. We have identified genes whose expression may be tightly regulated by epigenetic mechanisms and markers that may guide treatment decisions.

  19. The impact of ICAM1 and VCAM1 gene polymorphisms on long-term renal transplant function and recipient outcomes.

    PubMed

    Kłoda, Karolina; Domański, Leszek; Pawlik, Andrzej; Safranow, Krzysztof; Ciechanowski, Kazimierz

    2013-05-20

    Intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) are implicated in endothelial cell-leukocyte adhesion. Several functional polymorphisms that affect the expression of ICAM-1 and VCAM-1, and therefore post-transplant immunological response, have been found in genes of these molecules. The aim of this study was to examine the impact of ICAM1 and VCAM1 gene polymorphisms on long-term renal transplant function and recipient outcome during a 5-year follow-up period. The study enrolled 269 Caucasian renal transplant recipients (165 males, 104 females, mean age 47.58 ± 12.96 years) remaining under observation for 5 years. Genotyping of the c.1405A>G (Lys469Glu, rs5498) ICAM1, T(-1592)C (rs1041163), and T(-833)C (rs3170794) VCAM1 genes polymorphisms was performed using real-time PCR. The rs5498 ICAM1 GG genotype was an independent risk factor associated with increased creatinine concentration at 12, 24, 36, 48, and 60 months after transplantation (p=0.0004, p=0.02, p=0.01, p=0.01 and p=0.04, respectively). Creatinine concentrations 36 and 48 months after transplantation were higher among CC homozygotes of the rs1041163 VCAM1 (TT+CT vs. CC, p=0.049 and p=0.04, respectively). The C allele of the rs1041163 VCAM1 gene polymorphism was identified as a risk factor for dialysis after transplantation (HR=3.323, 95%CI=1.44-7.67, p=0.005). The C allele of the rs1041163 VCAM1 gene was at the border of statistical significance as a risk factor for death after transplantation (p=0.06). These results suggest that the rs5498 ICAM1 GG genotype is correlated with early and long-term allograft failure. The C allele of the rs1041163 VCAM1 gene may be associated with long-term allograft failure and graft loss, as well as increased morbidity after transplantation.

  20. Arsenic and phosphate rock impacted the abundance and diversity of bacterial arsenic oxidase and reductase genes in rhizosphere of As-hyperaccumulator Pteris vittata.

    PubMed

    Han, Yong-He; Fu, Jing-Wei; Xiang, Ping; Cao, Yue; Rathinasabapathi, Bala; Chen, Yanshan; Ma, Lena Q

    2017-01-05

    Microbially-mediated arsenic (As) transformation in soils affects As speciation and plant uptake. However, little is known about the impacts of As on bacterial communities and their functional genes in the rhizosphere of As-hyperaccumulator Pteris vittata. In this study, arsenite (AsIII) oxidase genes (aroA-like) and arsenate (AsV) reductase genes (arsC) were amplified from three soils, which were amended with 50mgkg(-1) As and/or 1.5% phosphate rock (PR) and grew P. vittata for 90 d. The aroA-like genes in the rhizosphere were 50 times more abundant than arsC genes, consistent with the dominance of AsV in soils. According to functional gene alignment, most bacteria belonged to α-, β- and γ-Proteobacteria. Moreover, aroA-like genes showed a higher biodiversity than arsC genes based on clone library analysis and could be grouped into nine clusters based on terminal restriction fragment length polymorphism (T-RFLP) analysis. Besides, AsV amendment elevated aroA-like gene diversity, but decreased arsC gene diversity. Redundancy analysis indicated that soil pH, available Ca and P, and AsV concentration were key factors driving diverse compositions in aroA-like gene community. This work identified new opportunities to screen for As-oxidizing and/or -reducing bacteria to aid phytoremediation of As-contaminated soils.

  1. Impact of APOE gene polymorphisms on the lipid profile in an Algerian population

    PubMed Central

    2013-01-01

    Background The importance of apolipoprotein E (APOE) in lipid and lipoprotein metabolism is well established. However, the impact of APOE polymorphisms has never been investigated in an Algerian population. This study assessed, for the fist time, the relationships between three APOE polymorphisms (epsilon, rs439401, rs4420638) and plasma lipid concentrations in a general population sample from Algeria. Methods The association analysis was performed in the ISOR study, a representative sample of the population living in Oran (787 subjects aged between 30 and 64). Polymorphisms were considered both individually and as haplotypes. Results In the ISOR sample, APOE ϵ4 allele carriers had higher plasma triglyceride (p=0.0002), total cholesterol (p=0.009) and LDL-cholesterol (p=0.003) levels than ϵ3 allele carriers. No significant associations were detected for the rs4420638 and rs439401 SNPs. Linkage disequilibrium and haplotype analyses confirmed the respectively deleterious and protective impacts of the ϵ4 and ϵ2 alleles on LDL-cholesterol levels and showed that the G allele of the rs4420638 polymorphism may exert a protective effect on LDL-cholesterol levels in subjects bearing the APOE epsilon 4 allele. Conclusion Our results showed that (i) the APOE epsilon polymorphism has the expected impact on the plasma lipid profile and (ii) the rs4420638 G allele may counterbalance the deleterious effect of the ϵ4 allele on LDL-cholesterol levels in an Algerian population. PMID:24160669

  2. Global Renal Gene Expression Profiling Analysis in B2-Kinin Receptor Null Mice: Impact of Diabetes

    PubMed Central

    Jaffa, Miran A.; Kobeissy, Firas; Al Hariri, Moustafa; Chalhoub, Hussein; Eid, Assaad; Ziyadeh, Fuad N.; Jaffa, Ayad A.

    2012-01-01

    Diabetic nephropathy (DN), the leading cause of end-stage renal failure, is clinically manifested by albuminuria and a progressive decline in glomerular filtration rate. The risk factors and mechanisms that contribute to the development and progression of DN are still incompletely defined. To address the involvement of bradykinin B2-receptors (B2R) in DN, we used a genome wide approach to study the effects of diabetes on differential renal gene expression profile in wild type and B2R knockout (B2R−/−) mice. Diabetes was induced with streptozotocin and plasma glucose levels and albumin excretion rate (AER) were measured at predetermined times throughout the 23 week study period. Longitudinal analysis of AER indicated that diabetic B2R−/−D null mice had a significantly decreased AER levels compared to wild type B2R+/+D mice (P = 0.0005). Results from the global microarray study comparing gene expression profiles among four groups of mice respectively: (B2R+/+C, B2R+/+D, B2R−/−C and B2R−/−D) highlighted the role of several altered pathological pathways in response to disruption of B2R and to the diabetic state that included: endothelial injury, oxidative stress, insulin and lipid metabolism and inflammatory process with a marked alteration in the pro-apoptotic genes. The findings of the present study provide a global genomics view of biomarkers that highlight the mechanisms and putative pathways involved in DN. PMID:23028588

  3. Impact of Pre-Analytical Variables on Cancer Targeted Gene Sequencing Efficiency

    PubMed Central

    Araujo, Luiz H.; Timmers, Cynthia; Shilo, Konstantin; Zhao, Weiqiang; Zhang, Jianying; Yu, Lianbo; Natarajan, Thanemozhi G.; Miller, Clinton J.; Yilmaz, Ayse Selen; Liu, Tom; Amann, Joseph; Lapa e Silva, José Roberto; Ferreira, Carlos Gil; Carbone, David P.

    2015-01-01

    Tumor specimens are often preserved as formalin-fixed paraffin-embedded (FFPE) tissue blocks, the most common clinical source for DNA sequencing. Herein, we evaluated the effect of pre-sequencing parameters to guide proper sample selection for targeted gene sequencing. Data from 113 FFPE lung tumor specimens were collected, and targeted gene sequencing was performed. Libraries were constructed using custom probes and were paired-end sequenced on a next generation sequencing platform. A PCR-based quality control (QC) assay was utilized to determine DNA quality, and a ratio was generated in comparison to control DNA. We observed that FFPE storage time, PCR/QC ratio, and DNA input in the library preparation were significantly correlated to most parameters of sequencing efficiency including depth of coverage, alignment rate, insert size, and read quality. A combined score using the three parameters was generated and proved highly accurate to predict sequencing metrics. We also showed wide read count variability within the genome, with worse coverage in regions of low GC content like in KRAS. Sample quality and GC content had independent effects on sequencing depth, and the worst results were observed in regions of low GC content in samples with poor quality. Our data confirm that FFPE samples are a reliable source for targeted gene sequencing in cancer, provided adequate sample quality controls are exercised. Tissue quality should be routinely assessed for pre-analytical factors, and sequencing depth may be limited in genomic regions of low GC content if suboptimal samples are utilized. PMID:26605948

  4. Impact of IL-22 gene polymorphism on human immunodeficiency virus infection in Han Chinese patients.

    PubMed

    Hu, Jun; Li, Yi; Chen, Lin; Yang, Zhengrong; Zhao, Guanglu; Wang, Yushu; Cheng, Jinquan; Zhao, Jin; Peng, Ying

    2016-12-01

    To analyze the polymorphism of the IL-22 gene in Han Chinese patients and to evaluate the influence of IL-22 polymorphism on human immunodeficiency virus (HIV) infection. IL-22 gene polymorphism was analyzed in 73 blood samples from healthy participants. The influence of the genotype and allele distribution of three single nucleotide polymorphisms (rs2227484, rs2227485, and rs2227513) of IL-22 on HIV infection was evaluated in 619 HIV seropositive patients and 619 healthy controls. To determine the association between the rs2227513 genotype and IL-22 levels in plasma, we randomly selected 29 HIV seropositive blood samples and 15 healthy blood samples and measured the levels of IL-22. Nine single nucleotide polymorphism loci of the IL-22 gene were found (rs2227484, rs2227485, rs2227491, rs2227508, rs2227513, rs1179249, rs1179250, rs1179251, and rs1182844). Stratified analysis (by sex) showed a higher association of HIV infection and the A/G genotype and G allele at rs2227513 in women, but not in men (A/G genotype odds ratio = 5.24, 95% confidence interval = 1.13-24.27; allele G odds ratio = 5.27, 95% confidence interval 1.15-24.23). The rs2227513 A/G genotype was also associated with significantly higher levels of plasma IL-22, regardless of whether the patient was HIV seropositive or seronegative. Our results suggest that IL-22 production in blood might act as a pathogenic factor in HIV infection. Copyright © 2014. Published by Elsevier B.V.

  5. The major histocompatibility complex genes impact pain response in DA and DA.1U rats.

    PubMed

    Guo, Yuan; Yao, Fan-Rong; Cao, Dong-Yuan; Li, Li; Wang, Hui-Sheng; Xie, Wen; Zhao, Yan

    2015-08-01

    Our recent studies have shown that the difference in basal pain sensitivity to mechanical and thermal stimulation between Dark-Agouti (DA) rats and a novel congenic DA.1U rats is major histocompatibility complex (MHC) genes dependent. In the present study, we further used DA and DA.1U rats to investigate the role of MHC genes in formalin-induced pain model by behavioral, electrophysiological and immunohistochemical methods. Behavioral results showed biphasic nociceptive behaviors increased significantly following the intraplantar injection of formalin in the hindpaw of DA and DA.1U rats. The main nociceptive behaviors were lifting and licking, especially in DA rats (P<0.001 and P<0.01). The composite pain scores (CPS) in DA rats were significantly higher than those in DA.1U rats in both phases of the formalin test (P<0.01). Electrophysiological results also showed the biphasic increase in discharge rates of C and Aδ fibers of L5 dorsal root in the two strains, and the net change of the discharge rate of DA rats was significantly higher than that of DA.1U rats (P<0.05). The mechanical thresholds decreased after formalin injection in both strains (P<0.01), and the net change in the mechanical threshold in DA was greater than that in DA.1U rats (P<0.05). The expression of RT1-B, representation of MHC class II molecule, in laminae I-II of L4/5 spinal cord in DA rats was significantly higher than that in DA.1U rats in the respective experimental group (P<0.05). These results suggested that both DA and DA.1U rats exhibited nociceptive responses in formalin-induced pain model and DA rats were more sensitive to noxious chemical stimulus than DA.1U rats, indicating that MHC genes might contribute to the difference in pain sensitivity.

  6. The impact of horizontal gene transfer on the biology of Clostridium difficile.

    PubMed

    Roberts, Adam P; Allan, Elaine; Mullany, Peter

    2014-01-01

    Clostridium difficile infection (CDI) is now recognised as the main cause of healthcare associated diarrhoea. Over the recent years there has been a change in the epidemiology of CDI with certain related strains dominating infection. These strains have been termed hyper-virulent and have successfully spread across the globe. Many C. difficile strains have had their genomes completely sequenced allowing researchers to build up a very detailed picture of the contribution of horizontal gene transfer to the adaptive potential, through the acquisition of mobile DNA, of this organism. Here, we review and discuss the contribution of mobile genetic elements to the biology of this clinically important pathogen.

  7. Prognostic Impact of 21-Gene Recurrence Score in Patients With Stage IV Breast Cancer: TBCRC 013.

    PubMed

    King, Tari A; Lyman, Jaclyn P; Gonen, Mithat; Voci, Amy; De Brot, Marina; Boafo, Camilla; Sing, Amy Pratt; Hwang, E Shelley; Alvarado, Michael D; Liu, Minetta C; Boughey, Judy C; McGuire, Kandace P; Van Poznak, Catherine H; Jacobs, Lisa K; Meszoely, Ingrid M; Krontiras, Helen; Babiera, Gildy V; Norton, Larry; Morrow, Monica; Hudis, Clifford A

    2016-07-10

    The objective of this study was to determine whether the 21-gene Recurrence Score (RS) provides clinically meaningful information in patients with de novo stage IV breast cancer enrolled in the Translational Breast Cancer Research Consortium (TBCRC) 013. TBCRC 013 was a multicenter prospective registry that evaluated the role of surgery of the primary tumor in patients with de novo stage IV breast cancer. From July 2009 to April 2012, 127 patients from 14 sites were enrolled; 109 (86%) patients had pretreatment primary tumor samples suitable for 21-gene RS analysis. Clinical variables, time to first progression (TTP), and 2-year overall survival (OS) were correlated with the 21-gene RS by using log-rank, Kaplan-Meier, and Cox regression. Median patient age was 52 years (21 to 79 years); the majority had hormone receptor-positive/human epidermal growth factor receptor 2 (HER2)-negative (72 [66%]) or hormone receptor-positive/HER2-positive (20 [18%]) breast cancer. At a median follow-up of 29 months, median TTP was 20 months (95% CI, 16 to 26 months), and median survival was 49 months (95% CI, 40 months to not reached). An RS was generated for 101 (93%) primary tumor samples: 22 (23%) low risk (< 18), 29 (28%) intermediate risk (18 to 30); and 50 (49%) high risk (≥ 31). For all patients, RS was associated with TTP (P = .01) and 2-year OS (P = .04). In multivariable Cox regression models among 69 patients with estrogen receptor (ER)-positive/HER2-negative cancer, RS was independently prognostic for TTP (hazard ratio, 1.40; 95% CI, 1.05 to 1.86; P = .02) and 2-year OS (hazard ratio, 1.83; 95% CI, 1.14 to 2.95; P = .013). The 21-gene RS is independently prognostic for both TTP and 2-year OS in ER-positive/HER2-negative de novo stage IV breast cancer. Prospective validation is needed to determine the potential role for this assay in the clinical management of this patient subset. © 2016 by American Society of Clinical Oncology.

  8. Impact of Mannose-Binding Protein Gene Polymorphisms in Omani Sickle Cell Disease Patients

    PubMed Central

    Zachariah, Mathew; Al Zadjali, Shoaib; Bashir, Wafa; Al Ambusaidi, Rahma; Misquith, Rhea; Wali, Yasser; Pathare, Anil

    2016-01-01

    Objectives Our aim was to study mannose-binding protein (MBP) polymorphisms in exonic and promoter region and correlate it with associated infections and vasoocculsive (VOC) episodes in sickle cell disease (SCD) patients since MBP plays an important role in innate immunity by activating the complement system. Methods We studied the genetic polymorphisms in the Exon 1 (alleles A/O) and promoter region (alleles Y/X; H/L, P/Q) of the MBL2 gene, in SCD patients as an increased incidence of infections is seen in these patients. A PCR-based, targeted genomic DNA sequencing of MBL2 was used to study 68 SCD Omani patients and 44 controls (healthy voluntary blood donors). Results In SCD patients, the frequency of the genotype related to the high production of MBL was 0.35 (YA/YA) and for intermediate/low production was 0.65 (YA/XA, XA/XA, YA/YO, XA/YO, YO/YO). The observed frequencies of MBL2 gene promoter polymorphism (-221, Y/X) were 44.4% and 20.5% for the heterozygous genotype Y/X and 3.2% and 2.2% for the homozygous (X/X) respectively between SCD patients and controls. MBL2 Exon1 gene mutations were 29.4% and 50% for the heterozygous genotype A/O and 5.9% and 6.8% respectively for the homozygous (O/O) genotype between SCD patients and controls. The distribution of variant MBL2 gene polymorphisms did not show any correlation in SCD patients with or without VOC attacks (p=0.16; OR −0.486; CI=0.177 −1.33), however, it was correlated with infections (p=0.0162; OR −3.55; CI 1.25–10.04). Conclusions Although the frequency of the genotypes and haplotypes of MBL2 in SCD patients did not differ from controls, overall in the SCD patient cohort the increased representation of variant alleles was significantly correlated with infections (p<0.05). However, these variant MBL2 polymorphisms did not seem to play a significant role in the VOC episodes in this SCD cohort. PMID:26977272

  9. Unraveling the estrogen receptor (er) genes in Atlantic salmon (Salmo salar) reveals expression differences between the two adult life stages but little impact from polychlorinated biphenyl (PCB) load.

    PubMed

    Nikoleris, Lina; Hansson, Maria C

    2015-01-15

    Estrogen receptors (ers) not only are activated by hormones but also interact with many human-derived environmental contaminants. Here, we present evidence for four expressed er genes in Atlantic salmon cDNA - two more ers (erα2 and erβ2) than previously published. To determine if er gene expression differs between two adult life-stages we sampled 20 adult salmon from the feeding phase in the Baltic Sea and during migration in the River Mörrum, Sweden. Results show that all four er genes are present in the investigated tissues, except for erα2 not appearing in the spleen. Overall, a profile analysis reveals the erα1 gene to be the most highly expressed er gene in both female and male Baltic Sea salmon tissues, and also in female River Mörrum salmon. In contrast, this gene has the lowest gene expression level of the four er genes in male salmon from the River Mörrum. The erα2 gene is expressed at the lowest levels in both female/male Baltic Sea salmon and in female River Mörrum salmon. Statistical analyses indicate a significant and complex interaction where both sex and adult life stage can impact er gene expression. Regression analyses did not demonstrate any significant relationship between polychlorinated biphenyl (PCB) body burden and er gene expression level, suggesting that accumulated pollutants from the Baltic Sea may be deactivated inside the salmon's lipid tissues and have limited impact on er activity. This study is the first comprehensive analysis of four er gene expression levels in two wild salmon populations from two different adult life stages where information about PCB load is also available.

  10. Parental divorce and adolescent delinquency: ruling out the impact of common genes.

    PubMed

    Burt, S Alexandra; Barnes, Ashlee R; McGue, Matt; Iacono, William G

    2008-11-01

    Although the well-documented association between parental divorce and adolescent delinquency is generally assumed to be environmental (i.e., causal) in origin, genetic mediation is also possible. Namely, the behavior problems often found in children of divorce could derive from similar pathology in the parents, pathology that is both heritable and increases the risk that the parent will experience divorce. To test these alternative hypotheses, the authors made use of a novel design that incorporated timing of divorce in a sample of 610 adoptive and biological families. They reasoned that if genes common to parent and child mediate this association, nonadopted youth should manifest increased delinquency in the presence of parental divorce even if the divorce preceded their birth (i.e., was from a prior parental relationship). However, should the association be environmental in origin, the authors reasoned that adolescents should manifest increased delinquency only in response to divorce exposure, and this association should not vary by adoption status. Results firmly supported the latter, suggesting that it is the experience of parental divorce, and not common genes, that drives the association between divorce and adolescent delinquency.

  11. Impact of Gene Patents and Licensing Practices on Access to Genetic Testing for Hereditary Hemochromatosis

    PubMed Central

    Chandrasekharan, Subhashini; Pitlick, Emily; Heaney, Christopher; Cook-Deegan, Robert

    2010-01-01

    Hereditary hemochromatosis (HH) is an iron metabolism disorder that leads to excess iron buildup, especially in the heart, liver, and pancreas. Mutations in the HFE gene are the single most common cause of HH, which can be treated effectively if diagnosed early. Patents cover the HFE gene, related proteins, screening methods, and testing kits. Most initial testing for HH is biochemical, but HFE DNA testing or genotyping is used to confirm a diagnosis of inherited hemochromatosis. Concerns over patents covering HFE testing emerged in 2002, when scholars argued that exclusive licensing and the patent-enabled sole provider model then in place led to high prices and limited access. Critics of the sole provider model noted that the test was available at multiple laboratories prior to the enforcement of patents. By 2007, however, Bio-Rad, Limited, acquired the key intellectual property and sub-licensed it widely. In part because of broad, non-exclusive licensing, there are now multiple providers and testing technologies, and research continues. This case study illustrates how both changes in intellectual property ownership and evolving clinical utility of HFE genetic testing in the last decade have effected the licensing of patents and availability of genetic testing. PMID:20393306

  12. Impact of Bi-Axial Shear on Atherogenic Gene Expression by Endothelial Cells.

    PubMed

    Chakraborty, Amlan; Chakraborty, Sutirtha; Jala, Venkatakrishna R; Thomas, Jonathan M; Sharp, M Keith; Berson, R Eric; Haribabu, Bodduluri

    2016-10-01

    This study demonstrated the effects of the directionality of oscillatory wall shear stress (WSS) on proliferation and proatherogenic gene expression (I-CAM, E-Selectin, and IL-6) in the presence of inflammatory mediators leukotriene B4 (LTB4) and bacterial lipopolysaccharide (LPS) from endothelial cells grown in an orbiting culture dish. Computational fluid dynamics (CFD) was applied to quantify the flow in the dish, while an analytical solution representing an extension of Stokes second problem was used for validation. Results indicated that WSS magnitude was relatively constant near the center of the dish and oscillated significantly (0-0.9 Pa) near the side walls. Experiments showed that LTB4 dominated the shear effects on cell proliferation and area. Addition of LPS didn't change proliferation, but significantly affected cell area. The expression of I-CAM1, E-Selectin and IL-6 were altered by directional oscillatory shear index (DOSI, a measure of the biaxiality of oscillatory shear), but not shear magnitude. The significance of DOSI was further reinforced by the strength of its interactions with other atherogenic factors. Hence, directionality of shear appears to be an important factor in regulating gene expression and provides a potential explanation of the propensity for increased vascular lesions in regions in the arteries with oscillating biaxial flow.

  13. The impact of osteopontin gene variations on multiple sclerosis development and progression.

    PubMed

    Comi, Cristoforo; Cappellano, Giuseppe; Chiocchetti, Annalisa; Orilieri, Elisabetta; Buttini, Sara; Ghezzi, Laura; Galimberti, Daniela; Guerini, Franca; Barizzone, Nadia; Perla, Franco; Leone, Maurizio; D'Alfonso, Sandra; Caputo, Domenico; Scarpini, Elio; Cantello, Roberto; Dianzani, Umberto

    2012-01-01

    Osteopontin is a proinflammatory molecule, modulating TH1 and TH17 responses. Several reports suggest its involvement in multiple sclerosis (MS) pathogenesis. We previously reported that OPN gene variations at the 3' end are a predisposing factor for MS development and evolution. In this paper, we extended our analysis to a gene variation at the 5' end on the -156G > GG single nucleotide polymorphism (SNP) and replicated our previous findings at the 3' end on the +1239A > C SNP. We found that only +1239A > C SNP displayed a statistically significant association with MS development, but both +1239A > C and -156G > GG had an influence on MS progression, since patients homozygous for both +1239A and -156GG alleles displayed slower progression of disability and slower switch to secondary progression than those carrying +1239C and/or -156G and those homozygous for +1239A only. Moreover, patients homozygous for +1239A also displayed a significantly lower relapse rate than those carrying +1239C, which is in line with the established role of OPN in MS relapses.

  14. The impact of FMR1 gene mutations on human reproduction and development: a systematic review.

    PubMed

    Noto, Vincenzo; Harrity, Conor; Walsh, David; Marron, Kevin

    2016-09-01

    This is a comprehensive review of the literature in this field attempting to put the FMR1 gene and its evaluation into context, both in general and for the reproductive health audience. Online database search of publications with systematic review of all papers relevant to ovarian reserve and assisted reproduction was done. Relevant papers were identified and assessed, and an attempt was made to understand, rationalize and explain the divergent views in this field of study. Seminal and original illustrations were employed. FMR1 is a highly conserved gene whose interpretation and effect on outcomes remains controversial in the reproductive health setting. Recent re-evaluations of the commonly accepted normal range have yielded interesting tools for possibly explaining unexpected outcomes in assisted reproduction. Fragile X investigations should perhaps become more routinely assessed in the reproductive health setting, particularly following a failed treatment cycle where oocyte quality is thought to be a contributing factor, or in the presence of a surprise finding of diminished ovarian reserve in a young patient.

  15. The Impact of Osteopontin Gene Variations on Multiple Sclerosis Development and Progression

    PubMed Central

    Comi, Cristoforo; Cappellano, Giuseppe; Chiocchetti, Annalisa; Orilieri, Elisabetta; Buttini, Sara; Ghezzi, Laura; Galimberti, Daniela; Guerini, Franca; Barizzone, Nadia; Perla, Franco; Leone, Maurizio; D'Alfonso, Sandra; Caputo, Domenico; Scarpini, Elio; Cantello, Roberto; Dianzani, Umberto

    2012-01-01

    Osteopontin is a proinflammatory molecule, modulating TH1 and TH17 responses. Several reports suggest its involvement in multiple sclerosis (MS) pathogenesis. We previously reported that OPN gene variations at the 3′ end are a predisposing factor for MS development and evolution. In this paper, we extended our analysis to a gene variation at the 5′ end on the −156G > GG single nucleotide polymorphism (SNP) and replicated our previous findings at the 3′ end on the +1239A > C SNP. We found that only +1239A > C SNP displayed a statistically significant association with MS development, but both +1239A > C and −156G > GG had an influence on MS progression, since patients homozygous for both +1239A and −156GG alleles displayed slower progression of disability and slower switch to secondary progression than those carrying +1239C and/or −156G and those homozygous for +1239A only. Moreover, patients homozygous for +1239A also displayed a significantly lower relapse rate than those carrying +1239C, which is in line with the established role of OPN in MS relapses. PMID:23008732

  16. Impact of Common Variation in Bone-Related Genes on Type 2 Diabetes and Related Traits

    PubMed Central

    Billings, Liana K.; Hsu, Yi-Hsiang; Ackerman, Rachel J.; Dupuis, Josée; Voight, Benjamin F.; Rasmussen-Torvik, Laura J.; Hercberg, Serge; Lathrop, Mark; Barnes, Daniel; Langenberg, Claudia; Hui, Jennie; Fu, Mao; Bouatia-Naji, Nabila; Lecoeur, Cecile; An, Ping; Magnusson, Patrik K.; Surakka, Ida; Ripatti, Samuli; Christiansen, Lene; Dalgård, Christine; Folkersen, Lasse; Grundberg, Elin; Eriksson, Per; Kaprio, Jaakko; Ohm Kyvik, Kirsten; Pedersen, Nancy L.; Borecki, Ingrid B.; Province, Michael A.; Balkau, Beverley; Froguel, Philippe; Shuldiner, Alan R.; Palmer, Lyle J.; Wareham, Nick; Meneton, Pierre; Johnson, Toby; Pankow, James S.; Karasik, David; Meigs, James B.; Kiel, Douglas P.; Florez, Jose C.

    2012-01-01

    Exploring genetic pleiotropy can provide clues to a mechanism underlying the observed epidemiological association between type 2 diabetes and heightened fracture risk. We examined genetic variants associated with bone mineral density (BMD) for association with type 2 diabetes and glycemic traits in large well-phenotyped and -genotyped consortia. We undertook follow-up analysis in ∼19,000 individuals and assessed gene expression. We queried single nucleotide polymorphisms (SNPs) associated with BMD at levels of genome-wide significance, variants in linkage disequilibrium (r2 > 0.5), and BMD candidate genes. SNP rs6867040, at the ITGA1 locus, was associated with a 0.0166 mmol/L (0.004) increase in fasting glucose per C allele in the combined analysis. Genetic variants in the ITGA1 locus were associated with its expression in the liver but not in adipose tissue. ITGA1 variants appeared among the top loci associated with type 2 diabetes, fasting insulin, β-cell function by homeostasis model assessment, and 2-h post–oral glucose tolerance test glucose and insulin levels. ITGA1 has demonstrated genetic pleiotropy in prior studies, and its suggested role in liver fibrosis, insulin secretion, and bone healing lends credence to its contribution to both osteoporosis and type 2 diabetes. These findings further underscore the link between skeletal and glucose metabolism and highlight a locus to direct future investigations. PMID:22698912

  17. Gastric Cancer and Helicobacter pylori: Impact of hopQII Gene.

    PubMed

    Kazemi, E; Kahrizi, D; Moradi, M T; Sohrabi, M; Yari, K

    2016-02-29

    The Helicobacter pylori is a Gram-negative, microaerophilic bacterium found usually in the stomach and use a number of mechanisms to survive in the stomach lumen. The presence of these bacteria in the stomach can lead to gastritis and reduction in stomach acid production. Acute inflammation can directly damage to the peripheral cells that are responsible for the secretion of acid. The risk of developing gastric carcinoma is associated to heterogeneity of Helicobacter pylori virulence factors. The HopQII is one of the outer membrane proteins involved in bacterial adherence to gastric mucosa and has been suggested to also play a role in the virulence of H. pylori. The purpose of the current study was to investigate the association between different H. pylori virulence hopQII allele and patients with gastroduodenal disorders. For this purpose 58 stomach biopsies of patients with gastric cancer and 100 saliva samples from healthy individuals were collected. Then genomic DNA was purified and PCR for was done for desired genes via specific primers. The H. pylori infections were diagnosed by PCR for GlmM gene. Then frequencies of hopQII+ and hopQII- genotypes was determined in H. pylori infected cases. Statistical analysis showed that there were not significant differences between healthy and diseased ones for genotype hopQII+.

  18. The impact of unprotected T cells in RNAi-based gene therapy for HIV-AIDS.

    PubMed

    Herrera-Carrillo, Elena; Liu, Ying Poi; Berkhout, Ben

    2014-03-01

    RNA interference (RNAi) is highly effective in inhibiting human immunodeficiency virus type 1 (HIV-1) replication by the expression of antiviral short hairpin RNA (shRNA) in stably transduced T-cell lines. For the development of a durable gene therapy that prevents viral escape, we proposed to combine multiple shRNAs against highly conserved regions of the HIV-1 RNA genome. The future in vivo application of such a gene therapy protocol will reach only a fraction of the T cells, such that HIV-1 replication will continue in the unmodified T cells, thereby possibly frustrating the therapy by generation of HIV-1 variants that escape from the inhibition imposed by the protected cells. We studied virus inhibition and evolution in pure cultures of shRNA-expressing cells versus mixed cell cultures of protected and unprotected T cells. The addition of the unprotected T cells indeed seems to accelerate HIV-1 evolution and escape from a single shRNA inhibitor. However, expression of three antiviral shRNAs from a single lentiviral vector prevents virus escape even in the presence of unprotected cells. These results support the idea to validate the therapeutic potential of this anti-HIV approach in appropriate in vivo models.

  19. Polymorphisms of the murine mitochondrial ND4, CYTB and COX3 genes impact hematopoiesis during aging

    PubMed Central

    Timmer, Katrin; Sekora, Anett; Knübel, Gudrun; Escobar, Hugo Murua; Fuellen, Georg; Ibrahim, Saleh M.; Tiedge, Markus; Baltrusch, Simone; Jaster, Robert; Köhling, Rüdiger; Junghanss, Christian

    2016-01-01

    During aging, mitochondrial DNA (mtDNA) can accumulate mutations leading to increasing levels of reactive oxygen species (ROS). Increased ROS were described to activate formerly quiescent hematopoietic stem cells (HSC). Mutations in mtDNA were shown to enhance the risk for myelodysplastic syndrome and leukemia. However, the complex relationship between mtDNA variations, ROS and aging of the hematopoietic system is not fully understood. Herein, three mouse strains with mtDNA polymorphisms in genes of respiratory chain complexes I (ND4), III (CYTB) and IV (COX3) were compared to a reference strain during aging. Analysis focused on ROS and ATP levels, bone marrow composition and blood counts. Additionally, hematopoietic restoration capacity following cytotoxic stress was tested. Mice with polymorphisms in ND4 and CYTB gene had significantly decreasing ROS levels in bone marrow cells during aging, without effecting ATP levels. In addition, the frequency of stem and progenitor cells increased during aging but the amount of lymphocytes in the peripheral blood decreased during aging. In summary, the presence of mtDNA polymorphisms affecting the respiratory chain complexes I, III and IV was associated with altered ROS levels as well as changes in BM and peripheral blood composition during aging. PMID:27626489

  20. Major intrinsic proteins (MIPs) in plants: a complex gene family with major impacts on plant phenotype.

    PubMed

    Forrest, Kerrie L; Bhave, Mrinal

    2007-10-01

    The ubiquitous cell membrane proteins called aquaporins are now firmly established as channel proteins that control the specific transport of water molecules across cell membranes in all living organisms. The aquaporins are thus likely to be of fundamental significance to all facets of plant growth and development affected by plant-water relations. A majority of plant aquaporins have been found to share essential structural features with the human aquaporin and exhibit water-transporting ability in various functional assays, and some have been shown experimentally to be of critical importance to plant survival. Furthermore, substantial evidence is now available from a number of plant species that shows differential gene expression of aquaporins in response to abiotic stresses such as salinity, drought, or cold and clearly establishes the aquaporins as major players in the response of plants to conditions that affect water availability. This review summarizes the function and regulation of these genes to develop a greater understanding of the response of plants to water insufficiency, and particularly, to identify tolerant genotypes of major crop species including wheat and rice and plants that are important in agroforestry.

  1. Reassessing domain architecture evolution of metazoan proteins: major impact of gene prediction errors.

    PubMed

    Nagy, Alinda; Szláma, György; Szarka, Eszter; Trexler, Mária; Bányai, László; Patthy, László

    2011-07-13

    In view of the fact that appearance of novel protein domain architectures (DA) is closely associated with biological innovations, there is a growing interest in the genome-scale reconstruction of the evolutionary history of the domain architectures of multidomain proteins. In such analyses, however, it is usually ignored that a significant proportion of Metazoan sequences analyzed is mispredicted and that this may seriously affect the validity of the conclusions. To estimate the contribution of errors in gene prediction to differences in DA of predicted proteins, we have used the high quality manually curated UniProtKB/Swiss-Prot database as a reference. For genome-scale analysis of domain architectures of predicted proteins we focused on RefSeq, EnsEMBL and NCBI's GNOMON predicted sequences of Metazoan species with completely sequenced genomes. Comparison of the DA of UniProtKB/Swiss-Prot sequences of worm, fly, zebrafish, frog, chick, mouse, rat and orangutan with those of human Swiss-Prot entries have identified relatively few cases where orthologs had different DA, although the percentage with different DA increased with evolutionary distance. In contrast with this, comparison of the DA of human, orangutan, rat, mouse, chicken, frog, zebrafish, worm and fly RefSeq, EnsEMBL and NCBI's GNOMON predicted protein sequences with those of the corresponding/orthologous human Swiss-Prot entries identified a significantly higher proportion of domain architecture differences than in the case of the comparison of Swiss-Prot entries. Analysis of RefSeq, EnsEMBL and NCBI's GNOMON predicted protein sequences with DAs different from those of their Swiss-Prot orthologs confirmed that the higher rate of domain architecture differences is due to errors in gene prediction, the majority of which could be corrected with our FixPred protocol. We have also demonstrated that contamination of databases with incomplete, abnormal or mispredicted sequences introduces a bias in DA

  2. Reassessing Domain Architecture Evolution of Metazoan Proteins: Major Impact of Gene Prediction Errors

    PubMed Central

    Nagy, Alinda; Szláma, György; Szarka, Eszter; Trexler, Mária; Bányai, László; Patthy, László

    2011-01-01

    In view of the fact that appearance of novel protein domain architectures (DA) is closely associated with biological innovations, there is a growing interest in the genome-scale reconstruction of the evolutionary history of the domain architectures of multidomain proteins. In such analyses, however, it is usually ignored that a significant proportion of Metazoan sequences analyzed is mispredicted and that this may seriously affect the validity of the conclusions. To estimate the contribution of errors in gene prediction to differences in DA of predicted proteins, we have used the high quality manually curated UniProtKB/Swiss-Prot database as a reference. For genome-scale analysis of domain architectures of predicted proteins we focused on RefSeq, EnsEMBL and NCBI's GNOMON predicted sequences of Metazoan species with completely sequenced genomes. Comparison of the DA of UniProtKB/Swiss-Prot sequences of worm, fly, zebrafish, frog, chick, mouse, rat and orangutan with those of human Swiss-Prot entries have identified relatively few cases where orthologs had different DA, although the percentage with different DA increased with evolutionary distance. In contrast with this, comparison of the DA of human, orangutan, rat, mouse, chicken, frog, zebrafish, worm and fly RefSeq, EnsEMBL and NCBI's GNOMON predicted protein sequences with those of the corresponding/orthologous human Swiss-Prot entries identified a significantly higher proportion of domain architecture differences than in the case of the comparison of Swiss-Prot entries. Analysis of RefSeq, EnsEMBL and NCBI's GNOMON predicted protein sequences with DAs different from those of their Swiss-Prot orthologs confirmed that the higher rate of domain architecture differences is due to errors in gene prediction, the majority of which could be corrected with our FixPred protocol. We have also demonstrated that contamination of databases with incomplete, abnormal or mispredicted sequences introduces a bias in DA

  3. OCD candidate gene SLC1A1/EAAT3 impacts basal ganglia-mediated activity and stereotypic behavior.

    PubMed

    Zike, Isaac D; Chohan, Muhammad O; Kopelman, Jared M; Krasnow, Emily N; Flicker, Daniel; Nautiyal, Katherine M; Bubser, Michael; Kellendonk, Christoph; Jones, Carrie K; Stanwood, Gregg; Tanaka, Kenji Fransis; Moore, Holly; Ahmari, Susanne E; Veenstra-VanderWeele, Jeremy

    2017-05-30

    Obsessive-compulsive disorder (OCD) is a chronic, disabling condition with inadequate treatment options that leave most patients with substantial residual symptoms. Structural, neurochemical, and behavioral findings point to a significant role for basal ganglia circuits and for the glutamate system in OCD. Genetic linkage and association studies in OCD point to SLC1A1, which encodes the neuronal glutamate/aspartate/cysteine transporter excitatory amino acid transporter 3 (EAAT3)/excitatory amino acid transporter 1 (EAAC1). However, no previous studies have investigated EAAT3 in basal ganglia circuits or in relation to OCD-related behavior. Here, we report a model of Slc1a1 loss based on an excisable STOP cassette that yields successful ablation of EAAT3 expression and function. Using amphetamine as a probe, we found that EAAT3 loss prevents expected increases in (i) locomotor activity, (ii) stereotypy, and (iii) immediate early gene induction in the dorsal striatum following amphetamine administration. Further, Slc1a1-STOP mice showed diminished grooming in an SKF-38393 challenge experiment, a pharmacologic model of OCD-like grooming behavior. This reduced grooming is accompanied by reduced dopamine D1 receptor binding in the dorsal striatum of Slc1a1-STOP mice. Slc1a1-STOP mice also exhibit reduced extracellular dopamine concentrations in the dorsal striatum both at baseline and following amphetamine challenge. Viral-mediated restoration of Slc1a1/EAAT3 expression in the midbrain but not in the striatum results in partial rescue of amphetamine-induced locomotion and stereotypy in Slc1a1-STOP mice, consistent with an impact of EAAT3 loss on presynaptic dopaminergic function. Collectively, these findings indicate that the most consistently associated OCD candidate gene impacts basal ganglia-dependent repetitive behaviors.

  4. CO2 induced seawater acidification impacts sea urchin larval development II: gene expression patterns in pluteus larvae.

    PubMed

    Stumpp, M; Dupont, S; Thorndyke, M C; Melzner, F

    2011-11-01

    Extensive use of fossil fuels is leading to increasing CO(2) concentrations in the atmosphere and causes changes in the carbonate chemistry of the oceans which represents a major sink for anthropogenic CO(2). As a result, the oceans' surface pH is expected to decrease by ca. 0.4 units by the year 2100, a major change with potentially negative consequences for some marine species. Because of their carbonate skeleton, sea urchins and their larval stages are regarded as likely to be one of the more sensitive taxa. In order to investigate sensitivity of pre-feeding (2 days post-fertilization) and feeding (4 and 7 days post-fertilization) pluteus larvae, we raised Strongylocentrotus purpuratus embryos in control (pH 8.1 and pCO(2) 41 Pa e.g. 399 μatm) and CO(2) acidified seawater with pH of 7.7 (pCO(2) 134 Pa e.g. 1318 μatm) and investigated growth, calcification and survival. At three time points (day 2, day 4 and day 7 post-fertilization), we measured the expression of 26 representative genes important for metabolism, calcification and ion regulation using RT-qPCR. After one week of development, we observed a significant difference in growth. Maximum differences in size were detected at day 4 (ca. 10% reduction in body length). A comparison of gene expression patterns using PCA and ANOSIM clearly distinguished between the different age groups (two-way ANOSIM: Global R=1) while acidification effects were less pronounced (Global R=0.518). Significant differences in gene expression patterns (ANOSIM R=0.938, SIMPER: 4.3% difference) were also detected at day 4 leading to the hypothesis that differences between CO(2) treatments could reflect patterns of expression seen in control experiments of a younger larva and thus a developmental artifact rather than a direct CO(2) effect. We found an up regulation of metabolic genes (between 10%and 20% in ATP-synthase, citrate synthase, pyruvate kinase and thiolase at day 4) and down regulation of calcification related genes

  5. A research program for the socioeconomic impacts of gene editing regulation.

    PubMed

    Whelan, Agustina I; Lema, Martin A

    2017-01-02

    Gene editing technologies are a group of recent innovations in plant breeding using molecular biology, which have in common the capability of introducing a site-directed mutation or deletion in the genome. The first cases of crops improved with these technologies are approaching the market; this has raised an international debate regarding if they should be regulated as genetically modified crops or just as another form of mutagenesis under conventional breeding. This dilemma for policymakers not only entails issues pertaining safety information and legal/regulatory definitions. It also demands borrowing tools developed in the field of social studies of science and technology, as an additional basis for sound decision making.

  6. Impact of mutations in Toll-like receptor pathway genes on esophageal carcinogenesis.

    PubMed

    Fels Elliott, Daffolyn Rachael; Perner, Juliane; Li, Xiaodun; Symmons, Martyn F; Verstak, Brett; Eldridge, Matthew; Bower, Lawrence; O'Donovan, Maria; Gay, Nick J; Fitzgerald, Rebecca C

    2017-05-01

    Esophageal adenocarcinoma (EAC) develops in an inflammatory microenvironment with reduced microbial diversity, but mechanisms for these influences remain poorly characterized. We hypothesized that mutations targeting the Toll-like receptor (TLR) pathway could disrupt innate immune signaling and promote a microenvironment that favors tumorigenesis. Through interrogating whole genome sequencing data from 171 EAC patients, we showed that non-synonymous mutations collectively affect the TLR pathway in 25/171 (14.6%, PathScan p = 8.7x10-5) tumors. TLR mutant cases were associated with more proximal tumors and metastatic disease, indicating possible clinical significance of these mutations. Only rare mutations were identified in adjacent Barrett's esophagus samples. We validated our findings in an external EAC dataset with non-synonymous TLR pathway mutations in 33/149 (22.1%, PathScan p = 0.05) tumors, and in other solid tumor types exposed to microbiomes in the COSMIC database (10,318 samples), including uterine endometrioid carcinoma (188/320, 58.8%), cutaneous melanoma (377/988, 38.2%), colorectal adenocarcinoma (402/1519, 26.5%), and stomach adenocarcinoma (151/579, 26.1%). TLR4 was the most frequently mutated gene with eleven mutations in 10/171 (5.8%) of EAC tumors. The TLR4 mutants E439G, S570I, F703C and R787H were confirmed to have impaired reactivity to bacterial lipopolysaccharide with marked reductions in signaling by luciferase reporter assays. Overall, our findings show that TLR pathway genes are recurrently mutated in EAC, and TLR4 mutations have decreased responsiveness to bacterial lipopolysaccharide and may play a role in disease pathogenesis in a subset of patients.

  7. MicroRNA-433 Dampens Glucocorticoid Receptor Signaling, Impacting Circadian Rhythm and Osteoblastic Gene Expression.

    PubMed

    Smith, Spenser S; Dole, Neha S; Franceschetti, Tiziana; Hrdlicka, Henry C; Delany, Anne M

    2016-10-07

    Serum glucocorticoids play a critical role in synchronizing circadian rhythm in peripheral tissues, and multiple mechanisms regulate tissue sensitivity to glucocorticoids. In the skeleton, circadian rhythm helps coordinate bone formation and resorption. Circadian rhythm is regulated through transcriptional and post-transcriptional feedback loops that include microRNAs. How microRNAs regulate circadian rhythm in bone is unexplored. We show that in mouse calvaria, miR-433 displays robust circadian rhythm, peaking just after dark. In C3H/10T1/2 cells synchronized with a pulse of dexamethasone, inhibition of miR-433 using a tough decoy altered the period and amplitude of Per2 gene expression, suggesting that miR-433 regulates rhythm. Although miR-433 does not directly target the Per2 3'-UTR, it does target two rhythmically expressed genes in calvaria, Igf1 and Hif1α. miR-433 can target the glucocorticoid receptor; however, glucocorticoid receptor protein abundance was unaffected in miR-433 decoy cells. Rather, miR-433 inhibition dramatically enhanced glucocorticoid signaling due to increased nuclear receptor translocation, activating glucocorticoid receptor transcriptional targets. Last, in calvaria of transgenic mice expressing a miR-433 decoy in osteoblastic cells (Col3.6 promoter), the amplitude of Per2 and Bmal1 mRNA rhythm was increased, confirming that miR-433 regulates circadian rhythm. miR-433 was previously shown to target Runx2, and mRNA for Runx2 and its downstream target, osteocalcin, were also increased in miR-433 decoy mouse calvaria. We hypothesize that miR-433 helps maintain circadian rhythm in osteoblasts by regulating sensitivity to glucocorticoid receptor signaling.

  8. Impact of mutations in Toll-like receptor pathway genes on esophageal carcinogenesis

    PubMed Central

    Fels Elliott, Daffolyn Rachael; Perner, Juliane; Li, Xiaodun; Symmons, Martyn F.; Verstak, Brett; Bower, Lawrence; O’Donovan, Maria; Fitzgerald, Rebecca C.

    2017-01-01

    Esophageal adenocarcinoma (EAC) develops in an inflammatory microenvironment with reduced microbial diversity, but mechanisms for these influences remain poorly characterized. We hypothesized that mutations targeting the Toll-like receptor (TLR) pathway could disrupt innate immune signaling and promote a microenvironment that favors tumorigenesis. Through interrogating whole genome sequencing data from 171 EAC patients, we showed that non-synonymous mutations collectively affect the TLR pathway in 25/171 (14.6%, PathScan p = 8.7x10-5) tumors. TLR mutant cases were associated with more proximal tumors and metastatic disease, indicating possible clinical significance of these mutations. Only rare mutations were identified in adjacent Barrett’s esophagus samples. We validated our findings in an external EAC dataset with non-synonymous TLR pathway mutations in 33/149 (22.1%, PathScan p = 0.05) tumors, and in other solid tumor types exposed to microbiomes in the COSMIC database (10,318 samples), including uterine endometrioid carcinoma (188/320, 58.8%), cutaneous melanoma (377/988, 38.2%), colorectal adenocarcinoma (402/1519, 26.5%), and stomach adenocarcinoma (151/579, 26.1%). TLR4 was the most frequently mutated gene with eleven mutations in 10/171 (5.8%) of EAC tumors. The TLR4 mutants E439G, S570I, F703C and R787H were confirmed to have impaired reactivity to bacterial lipopolysaccharide with marked reductions in signaling by luciferase reporter assays. Overall, our findings show that TLR pathway genes are recurrently mutated in EAC, and TLR4 mutations have decreased responsiveness to bacterial lipopolysaccharide and may play a role in disease pathogenesis in a subset of patients. PMID:28531216

  9. Interplay of Promoter Usage and Intragenic CpG Content: Impact on GFP Reporter Gene Expression.

    PubMed

    Krinner, Simone; Heitzer, Asli; Asbach, Benedikt; Wagner, Ralf

    2015-12-01

    Successful therapeutic protein production in vitro and in vivo requires efficient and long-term transgene expression supported by optimized vector and transgene cis-regulatory sequence elements. This study provides a comparative analysis of CpG-rich, highly expressed, versus CpG-depleted, poorly expressed green fluorescent protein (GFP) reporter transgenes, transcribed by various promoters in two different cell systems. Long-term GFP expression from a defined locus in stable Chinese hamster ovary cells was clearly influenced by the combination of transgene CpG content and promoter usage, as shown by differential silencing effects on selection pressure removal among the cytomegalovirus (CMV) promoter and elongation factor (EF)-1α promoter. Whereas a high intragenic CpG content promoted local DNA methylation, CpG depletion rather accelerated transgene loss and increased the local chromatin density. On lentiviral transfer of various expression modules into epigenetically sensitive P19 embryonic pluripotent carcinoma cells, CMV promoter usage led to rapid gene silencing irrespective of the intragenic CpG content. In contrast, EF-1α promoter-controlled constructs showed delayed silencing activity and high-level transgene expression, in particular when the CpG-rich GFP reporter was used. Notably, GFP silencing in P19 cells could be prevented completely by the bidirectional, dual divergently transcribed A2UCOE (ubiquitously acting chromatin-opening element derived from the human HNRPA2B1-CBX3 locus) promoter. Because the level of GFP expression by the A2UCOE promoter was entirely unaffected by the intragenic CpG level, we suggest that A2UCOE can overcome chromatin compaction resulting from intragenic CpG depletion due to its ascribed chromatin-opening abilities. Our analyses provide insights into the interplay of the intragenic CpG content with promoter sequences and regulatory sequence elements, thus contributing toward the design of therapeutic transgene expression

  10. Deletion of the APOBEC3B gene strongly impacts susceptibility to falciparum malaria.

    PubMed

    Jha, Pankaj; Sinha, Swapnil; Kanchan, Kanika; Qidwai, Tabish; Narang, Ankita; Singh, Prashant Kumar; Pati, Sudhanshu S; Mohanty, Sanjib; Mishra, Saroj K; Sharma, Surya K; Awasthi, Shally; Venkatesh, Vimala; Jain, Sanjeev; Basu, Analabha; Xu, Shuhua; Mukerji, Mitali; Habib, Saman

    2012-01-01

    APOBEC3B, a gene involved in innate response, exhibits insertion-deletion polymorphism across world populations. We observed the insertion allele to be nearly fixed in malaria endemic regions of sub-Saharan Africa as well as populations with high malaria incidence in the past. This prompted us to investigate the possible association of the polymorphism with falciparum malaria. We studied the distribution of APOBEC3B, in 25 diverse Indian populations comprising of 500 samples and 176 severe or non-severe Plasmodium falciparum patients and 174 ethnically-matched uninfected individuals from a P. falciparum endemic and a non-endemic region of India. The deletion frequencies ranged from 0% to 43% in the Indian populations. The frequency of the insertion allele strikingly correlated with the endemicity map of P. falciparum malaria in India. A strong association of the deletion allele with susceptibility to falciparum malaria in the endemic region (non-severe vs. control, Odds ratio=4.96, P value=9.5E(-06); severe vs. control, OR=4.36, P value=5.76E(-05)) was observed. Although the frequency of deletion allele was higher in the non-endemic region, there was a significant association of the homozygous deletion genotype with malaria (OR=3.17, 95% CI=1.10-10.32, P value=0.0177). Our study also presents a case for malaria as a positive selection force for the APOBEC3B insertion and suggests a major role for this gene in innate immunity against malaria.

  11. Sexual Polyploidization in Medicago sativa L.: Impact on the Phenotype, Gene Transcription, and Genome Methylation.

    PubMed

    Rosellini, Daniele; Ferradini, Nicoletta; Allegrucci, Stefano; Capomaccio, Stefano; Zago, Elisa Debora; Leonetti, Paola; Balech, Bachir; Aversano, Riccardo; Carputo, Domenico; Reale, Lara; Veronesi, Fabio

    2016-04-07

    Polyploidization as the consequence of 2n gamete formation is a prominent mechanism in plant evolution. Studying its effects on the genome, and on genome expression, has both basic and applied interest. We crossed two diploid (2n = 2x = 16) Medicago sativa plants, a subsp. falcata seed parent, and a coerulea × falcata pollen parent that form a mixture of n and 2n eggs and pollen, respectively. Such a cross produced full-sib diploid and tetraploid (2n = 4x = 32) hybrids, the latter being the result of bilateral sexual polyploidization (BSP). These unique materials allowed us to investigate the effects of BSP, and to separate the effect of intraspecific hybridization from those of polyploidization by comparing 2x with 4x full sib progeny plants. Simple sequence repeat marker segregation demonstrated tetrasomic inheritance for all chromosomes but one, demonstrating that these neotetraploids are true autotetraploids. BSP brought about increased biomass, earlier flowering, higher seed set and weight, and larger leaves with larger cells. Microarray analyses with M. truncatula gene chips showed that several hundred genes, related to diverse metabolic functions, changed their expression level as a consequence of polyploidization. In addition, cytosine methylation increased in 2x, but not in 4x, hybrids. Our results indicate that sexual polyploidization induces significant transcriptional novelty, possibly mediated in part by DNA methylation, and phenotypic novelty that could underpin improved adaptation and reproductive success of tetraploid M. sativa with respect to its diploid progenitor. These polyploidy-induced changes may have promoted the adoption of tetraploid alfalfa in agriculture. Copyright © 2016 Rosellini et al.

  12. Sexual Polyploidization in Medicago sativa L.: Impact on the Phenotype, Gene Transcription, and Genome Methylation

    PubMed Central

    Rosellini, Daniele; Ferradini, Nicoletta; Allegrucci, Stefano; Capomaccio, Stefano; Zago, Elisa Debora; Leonetti, Paola; Balech, Bachir; Aversano, Riccardo; Carputo, Domenico; Reale, Lara; Veronesi, Fabio

    2016-01-01

    Polyploidization as the consequence of 2n gamete formation is a prominent mechanism in plant evolution. Studying its effects on the genome, and on genome expression, has both basic and applied interest. We crossed two diploid (2n = 2x = 16) Medicago sativa plants, a subsp. falcata seed parent, and a coerulea × falcata pollen parent that form a mixture of n and 2n eggs and pollen, respectively. Such a cross produced full-sib diploid and tetraploid (2n = 4x = 32) hybrids, the latter being the result of bilateral sexual polyploidization (BSP). These unique materials allowed us to investigate the effects of BSP, and to separate the effect of intraspecific hybridization from those of polyploidization by comparing 2x with 4x full sib progeny plants. Simple sequence repeat marker segregation demonstrated tetrasomic inheritance for all chromosomes but one, demonstrating that these neotetraploids are true autotetraploids. BSP brought about increased biomass, earlier flowering, higher seed set and weight, and larger leaves with larger cells. Microarray analyses with M. truncatula gene chips showed that several hundred genes, related to diverse metabolic functions, changed their expression level as a consequence of polyploidization. In addition, cytosine methylation increased in 2x, but not in 4x, hybrids. Our results indicate that sexual polyploidization induces significant transcriptional novelty, possibly mediated in part by DNA methylation, and phenotypic novelty that could underpin improved adaptation and reproductive success of tetraploid M. sativa with respect to its diploid progenitor. These polyploidy-induced changes may have promoted the adoption of tetraploid alfalfa in agriculture. PMID:26858330

  13. Impact of Gene Patents and Licensing Practices on Access to Genetic Testing for Cystic Fibrosis

    PubMed Central

    Chandrasekharan, Subhashini; Heaney, Christopher; James, Tamara; Conover, Chris; Cook-Deegan, Robert

    2010-01-01

    Cystic fibrosis (CF) is one of the most commonly tested autosomal recessive disorders in the US. Clinical CF is associated with mutations in the CFTR gene, of which the most common mutation among Caucasians, ΔF508, was identified in 1989. The University of Michigan, Johns Hopkins University, and the Hospital for Sick Children, where much of the initial research occurred, hold key patents for CF genetic sequences, mutations and methods for detecting them. Several patents including the one that covers detection of the ΔF508 mutation are jointly held by the University of Michigan and the Hospital for Sick Children in Toronto, with Michigan administering patent licensing in the US. The University of Michigan broadly licenses the ΔF508 patent for genetic testing with over 60 providers of genetic testing to date. Genetic testing is now used in newborn screening, diagnosis, and reproductive decisions. Interviews with key researchers and intellectual property managers, a survey of laboratories’ prices for CF genetic testing, a review of literature on CF tests’ cost effectiveness, and a review of the developing market for CF testing provide no evidence that patents have significantly hindered access to genetic tests for CF or prevented financially cost-effective screening. Current licensing practices for cystic fibrosis (CF) genetic testing appear to facilitate both academic research and commercial testing. More than one thousand different CFTR mutations have been identified, and research continues to determine their clinical significance. Patents have been nonexclusively licensed for diagnostic use, and have been variably licensed for gene transfer and other therapeutic applications. The Cystic Fibrosis Foundation has been engaged in licensing decisions, making CF a model of collaborative and cooperative patenting and licensing practice. PMID:20393308

  14. Impact of Huntington Disease Gene-Positive Status on Pre-Symptomatic Young Adults and Recommendations for Genetic Counselors.

    PubMed

    Gong, Ping; Fanos, Joanna H; Korty, Lauren; Siskind, Carly E; Hanson-Kahn, Andrea K

    2016-12-01

    Huntington disease (HD) is an autosomal dominant, progressive neurodegenerative disorder for which there is no cure. Predictive testing for HD is available to asymptomatic at-risk individuals. Approximately half of the population undergoing predictive testing for HD consists of young adults (≤35 years old). Finishing one's education, starting a career, engaging in romantic relationships and becoming a parent are key milestones of young adulthood. We conducted a qualitative study to explore how testing gene-positive for HD influences young adults' attainment of these milestones, and to identify major challenges that pre-symptomatic young adults face to aid the development of targeted genetic counseling. Results of our study demonstrate that 1) knowing one's gene-positive status results in an urgency to reach milestones and positively changes young adults' approach to life; 2) testing positive influences young adults' education and career choices, romantic relationships, and family planning; 3) young adults desire flexible and tailored genetic counseling to address needs and concerns unique to this population. Findings of this study contribute to the understanding of the impact of predictive testing for HD on young adults, and highlight issues unique to this population that call for further research, intervention and advocacy.

  15. Null Mutants of Individual RABA Genes Impact the Proportion of Different Cell Wall Components in Stem Tissue of Arabidopsis thaliana

    PubMed Central

    Lunn, Daniel; Gaddipati, Sanyasi R.; Tucker, Gregory A.; Lycett, Grantley W.

    2013-01-01

    In Arabidopsis, and other plants, the RABA GTPases (orthologous to the Rab11a of mammals) have expanded in number and diversity and have been shown to belong to eight sub clades, some of which have been implicated in controlling vesicles that traffic cell wall polymers and enzymes that synthesise or modify them to the cell wall. In order to investigate this, we have investigated whether T-DNA insertion knockouts of individual RABA genes belonging to different sub clades, impact on the composition of the plant cell wall. Single gene knockouts of the RABA1, RABA2 and RABA4 sub clades primarily affected the percentage composition of pectin, cellulose and hemicellulose within the cell wall, respectively, despite having no obvious phenotype in the whole plant. We hypothesise that vesicles carrying specific types of cargoes from the Golgi to the cell surface may be regulated by particular sub types of RABA proteins, a finding that could have wider implications for how trafficking systems work and could be a useful tool in cell wall research and other fields of plant biology. PMID:24124508

  16. Sequence variations of the partially dominant DELLA gene Rht-B1c in wheat and their functional impacts.

    PubMed

    Wen, Wen; Deng, Qingyan; Jia, Haiyan; Wei, Lingzhu; Wei, Jingbo; Wan, Hongshen; Yang, Liming; Cao, Wenjin; Ma, Zhengqiang

    2013-08-01

    Rht-B1c, allelic to the DELLA protein-encoding gene Rht-B1a, is a natural mutation documented in common wheat (Triticum aestivum). It confers variation to a number of traits related to cell and plant morphology, seed dormancy, and photosynthesis. The present study was conducted to examine the sequence variations of Rht-B1c and their functional impacts. The results showed that Rht-B1c was partially dominant or co-dominant for plant height, and exhibited an increased dwarfing effect. At the sequence level, Rht-B1c differed from Rht-B1a by one 2kb Veju retrotransposon insertion, three coding region single nucleotide polymorphisms (SNPs), one 197bp insertion, and four SNPs in the 1kb upstream sequence. Haplotype investigations, association analyses, transient expression assays, and expression profiling showed that the Veju insertion was primarily responsible for the extreme dwarfing effect. It was found that the Veju insertion changed processing of the Rht-B1c transcripts and resulted in DELLA motif primary structure disruption. Expression assays showed that Rht-B1c caused reduction of total Rht-1 transcript levels, and up-regulation of GATA-like transcription factors and genes positively regulated by these factors, suggesting that one way in which Rht-1 proteins affect plant growth and development is through GATA-like transcription factor regulation.

  17. The Impact of CYP1A2 and CYP2E1 Genes Polymorphism on Theophylline Response.

    PubMed

    Sutrisna, Em

    2016-11-01

    Theophylline is a medicine with narrow therapeutic index. This implies that a small change in dosage would cause side effects. Theophylline is metabolized by CYP1A2 and CYP2E1. The aim of this review is to know the impact of CYP1A2 and CYP2E1 genes polymorphism on theophylline response. The review was done by searching literature in Pubmed and Science Direct databases with keywords 'polymorphism', 'pharmacogenetic', 'CYP1A2', 'CYP2E1' and 'theophylline'. There were 5 research articles from Pubmed and 65 articles (21 research articles, 23 review articles and 21 book chapters) from Science Direct. The exclusion criteria were - articles discussing about polymorphism but not CYP1A2 or CYP2E1, the ones with a mention of theophylline but not about its metabolism, articles on CYP1A2 and/or 2E1 polymorphism but not on the effect on theophylline. Thus, 33 articles were reviewed due to their suitability. The review discusses the influence of polymorphism of CYP1A2 and CYP2E1 genes on theophylline response.

  18. Impact of the HTR3A gene with early life trauma on emotional brain networks and depressed mood.

    PubMed

    Gatt, Justine M; Williams, Leanne M; Schofield, Peter R; Dobson-Stone, Carol; Paul, Robert H; Grieve, Stuart M; Clark, C Richard; Gordon, Evian; Nemeroff, Charles B

    2010-08-01

    The risk for mental illnesses such as depression is increasingly conceptualized as the product of gene-environment interactions and their impact on brain structure and function. The role of serotonin 3A receptor gene (HTR3A -42C>T polymorphism) and its interaction with early life stress (ELS) was investigated in view of the receptor's localization to brain regions central to emotion processing. Fronto-limbic grey matter (GM) loss was measured using magnetic resonance imaging and assessed using voxel-based morphometry analysis in 397 nonclinical individuals from the Brain Resource International Database. Negative mood symptoms were also assessed. The HTR3A CC genotype group, compared to the T carriers, demonstrated comparative loss to GM in hippocampal structures, which extended to the frontal cortices for those CC genotype individuals also exposed to ELS. Elevations in depressed mood were also evident. These findings suggest that the HTR3A CC genotype may be associated with alterations in brain structures central to emotion processing, particularly when exposed to stress, and further highlight the potential role of the serotonin system in the pathophysiology of affective disorders. In contrast, those individuals with the T allele, in particular the TT genotype, may be more protected from such alterations combined with minimal exposure to ELS events. 2010 Wiley-Liss, Inc.

  19. Impact of cluster thinning on transcriptional regulation of anthocyanin biosynthesis-related genes in 'Summer Black' grapes.

    PubMed

    Xi, Xiaojun; Zha, Qian; Jiang, Aili; Tian, Yihua

    2016-07-01

    Cluster thinning is an agronomic practice that strongly affects anthocyanin biosynthesis in the skin of grape berries. However, the impact of cluster thinning on anthocyanin biosynthesis has not been fully elucidated at the molecular level. Here, we investigated its effects on the berry quality, the biosynthesis of anthocyanins, and the expression levels of related genes from the onset of véraison to harvest in 'Summer Black' grapes. It was observed that the total soluble solid and anthocyanin content in berry skin significantly increased under cluster thinning, whereas the berry weight and titratable acidity showed no differences from the beginning of véraison to harvest. The expression level of most anthocyanin biosynthesis-related genes was significantly up-regulated by cluster thinning from the beginning of véraison and was higher at its end compared to the control. Up-regulation of flavonoid 3',5'-hydroxylase (F3'5'H) and O-methyltransferase (OMT) expression, and down-regulation of flavonoid 3'-hydroxylase (F3'H) expression were observed, which might be the cause of shift in the anthocyanin profile. These findings provide insights into the molecular basis of the relationship between cluster thinning and anthocyanin biosynthesis in the grape berry skin. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  20. Genome-Wide Nucleosome Occupancy and Positioning and Their Impact on Gene Expression and Evolution in Plants.

    PubMed

    Zhang, Tao; Zhang, Wenli; Jiang, Jiming

    2015-08-01

    The fundamental unit of chromatin is the nucleosome that consists of a protein octamer composed of the four core histones (Hs; H3, H4, H2A, and H2B) wrapped by 147 bp of DNA. Nucleosome occupancy and positioning have proven to be dynamic and have a critical impact on expression, regulation, and evolution of eukaryotic genes. We developed nucleosome occupancy and positioning data sets using leaf tissue of rice (Oryza sativa) and both leaf and flower tissues of Arabidopsis (Arabidopsis thaliana). We show that model plant and animal species share the fundamental characteristics associated with nucleosome dynamics. Only 12% and 16% of the Arabidopsis and rice genomes, respectively, were occupied by well-positioned nucleosomes. The cores of positioned nucleosomes were enriched with G/C dinucleotides and showed a lower C→T mutation rate than the linker sequences. We discovered that nucleosomes associated with heterochromatic regions were more spaced with longer linkers than those in euchromatic regions in both plant species. Surprisingly, different nucleosome densities were found to be associated with chromatin in leaf and flower tissues in Arabidopsis. We show that deep MNase-seq data sets can be used to map nucleosome occupancy of specific genomic loci and reveal gene expression patterns correlated with chromatin dynamics in plant genomes. © 2015 American Society of Plant Biologists. All Rights Reserved.

  1. Impacts of light and temperature on shoot branching gradient and expression of strigolactone synthesis and signalling genes in rose.

    PubMed

    Djennane, Samia; Hibrand-Saint Oyant, Laurence; Kawamura, Koji; Lalanne, David; Laffaire, Michel; Thouroude, Tatiana; Chalain, Séverine; Sakr, Soulaiman; Boumaza, Rachid; Foucher, Fabrice; Leduc, Nathalie

    2014-03-01

    Light and temperature are two environmental factors that deeply affect bud outgrowth. However, little is known about their impact on the bud burst gradient along a stem and their interactions with the molecular mechanisms of bud burst control. We investigated this question in two acrotonic rose cultivars. We demonstrated that the darkening of distal buds or exposure to cold (5 °C) prior to transfer to mild temperatures (20 °C) both repress acrotony, allowing the burst of quiescent medial and proximal buds. We sequenced the strigolactone pathway MAX-homologous genes in rose and studied their expression in buds and internodes along the stem. Only expressions of RwMAX1, RwMAX2 and RwMAX4 were detected. Darkening of the distal part of the shoot triggered a strong increase of RwMAX2 expression in darkened buds and bark-phloem samples, whereas it suppressed the acropetal gradient of the expression of RwMAX1 observed in stems fully exposed to light. Cold treatment induced an acropetal gradient of expression of RwMAX1 in internodes and of RwMAX2 in buds along the stem. Our results suggest that the bud burst gradient along the stem cannot be explained by a gradient of expression of RwMAX genes but rather by their local level of expression at each individual position.

  2. Staphylococcus aureus Biofilm and Planktonic cultures differentially impact gene expression, mapk phosphorylation, and cytokine production in human keratinocytes.

    PubMed

    Secor, Patrick R; James, Garth A; Fleckman, Philip; Olerud, John E; McInnerney, Kate; Stewart, Philip S

    2011-06-21

    Many chronic diseases, such as non-healing wounds are characterized by prolonged inflammation and respond poorly to conventional treatment. Bacterial biofilms are a major impediment to wound healing. Persistent infection of the skin allows the formation of complex bacterial communities termed biofilm. Bacteria living in biofilms are phenotypically distinct from their planktonic counterparts and are orders of magnitude more resistant to antibiotics, host immune response, and environmental stress. Staphylococcus aureus is prevalent in cutaneous infections such as chronic wounds and is an important human pathogen. The impact of S. aureus soluble products in biofilm-conditioned medium (BCM) or in planktonic-conditioned medium (PCM) on human keratinocytes was investigated. Proteomic analysis of BCM and PCM revealed differential protein compositions with PCM containing several enzymes involved in glycolysis. Global gene expression of keratinocytes exposed to biofilm and planktonic S. aureus was analyzed after four hours of exposure. Gene ontology terms associated with responses to bacteria, inflammation, apoptosis, chemotaxis, and signal transduction were enriched in BCM treated keratinocytes. Several transcripts encoding cytokines were also upregulated by BCM after four hours. ELISA analysis of cytokines confirmed microarray results at four hours and revealed that after 24 hours of exposure, S. aureus biofilm induced sustained low level cytokine production compared to near exponential increases of cytokines in planktonic treated keratinocytes. The reduction in cytokines produced by keratinocytes exposed to biofilm was accompanied by suppressed phosphorylation of MAPKs. Chemical inhibition of MAPKs did not drastically reduce cytokine production in BCM-treated keratinocytes suggesting that the majority of cytokine production is mediated through MAPK-independent mechanisms. Collectively the results indicate that S. aureus biofilms induce a distinct inflammatory response

  3. Impact of fusion gene status versus histology on risk-stratification for rhabdomyosarcoma: Retrospective analyses of patients on UK trials.

    PubMed

    Selfe, Joanna; Olmos, David; Al-Saadi, Reem; Thway, Khin; Chisholm, Julia; Kelsey, Anna; Shipley, Janet

    2017-07-01

    Long-term toxicities from current treatments are a major issue in paediatric cancer. Previous studies, including our own, have shown prognostic value for the presence of PAX3/7-FOXO1 fusion genes in rhabdomyosarcoma (RMS). It is proposed to introduce PAX3/7-FOXO1 positivity as a component of risk stratification, rather than alveolar histology, in future clinical trials. To assess the potential impact of this reclassification, we have determined the changes to risk category assignment of 210 histologically reviewed patients treated in the UK from previous malignant mesenchymal tumour clinical trials for non-metastatic RMS based on identification of PAX3/7-FOXO1 by fluorescence in situ hybridisation and/or reverse transcription PCR. Using fusion gene positivity in the current risk stratification would reassign 7% of patients to different European Paediatric Soft Tissue Sarcoma Study Group (EpSSG) risk groups. The next European trial would have 80% power to detect differences in event-free survival of 15% over 10 years and 20% over 5 years in reassigned patients. This would decrease treatment for over a quarter of patients with alveolar histology tumours that lack PAX3/7-FOXO1. Fusion gene status used in stratification may result in significant numbers of patients benefitting from lower treatment-associated toxicity. Prospective testing to show this reassignment maintains current survival rates is now required and is shown to be feasible based on estimated recruitment to a future EpSSG trial. Together with developing novel therapeutic strategies for patients identified as higher risk, this may ultimately improve the outcome and quality of life for patients with RMS. © 2016 Wiley Periodicals, Inc.

  4. Different impacts of manure and chemical fertilizers on bacterial community structure and antibiotic resistance genes in arable soils.

    PubMed

    Liu, Peng; Jia, Shuyu; He, Xiwei; Zhang, Xuxiang; Ye, Lin

    2017-09-04

    Both manure and chemical fertilizers are widely used in modern agriculture. However, the impacts of different fertilizers on bacterial community structure and antibiotic resistance genes (ARGs) in arable soils still remain unclear. In this study, high-throughput sequencing and quantitative PCR were employed to investigate the bacterial community structure, ARGs and mobile genetic elements (MGEs) influenced by the application of different fertilizers, including chemical fertilizers, piggery manure and straw ash. The results showed that the application of fertilizers could significantly change the soil bacterial community and the abundance of Gaiella under phylum Actinobacteria was significantly reduced from 12.9% in unfertilized soil to 4.1%-7.4% in fertilized soil (P < 0.05). It was also found that the application of manure could cause a transient effect on soil resistome composition and the relative abundance of ARGs increased from 7.37 ppm to 32.10 ppm. The abundance of aminoglycoside, sulfonamide and tetracycline resistance genes greatly increased after manure fertilization and then gradually returned to normal levels with the decay of some intestinal bacteria carrying ARGs. In contrast, the application of chemical fertilizers and straw ash significantly changed the bacterial community structure but exerted little effect on soil resistome. Overall, the results of this study illustrated the different effects of different fertilizers on the soil resistome and revealed that the changes of soil resistome induced by manure application mainly resulted from alteration of bacteria community rather than the horizontal gene transfer. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Epistasis × environment interactions among Arabidopsis thaliana glucosinolate genes impact complex traits and fitness in the field.

    PubMed

    Kerwin, Rachel E; Feusier, Julie; Muok, Alise; Lin, Catherine; Larson, Brandon; Copeland, Daniel; Corwin, Jason A; Rubin, Matthew J; Francisco, Marta; Li, Baohua; Joseph, Bindu; Weinig, Cynthia; Kliebenstein, Daniel J

    2017-08-01

    Despite the growing number of studies showing that genotype × environment and epistatic interactions control fitness, the influences of epistasis × environment interactions on adaptive trait evolution remain largely uncharacterized. Across three field trials, we quantified aliphatic glucosinolate (GSL) defense chemistry, leaf damage, and relative fitness using mutant lines of Arabidopsis thaliana varying at pairs of causal aliphatic GSL defense genes to test the impact of epistatic and epistasis × environment interactions on adaptive trait variation. We found that aliphatic GSL accumulation was primarily influenced by additive and epistatic genetic variation, leaf damage was primarily influenced by environmental variation and relative fitness was primarily influenced by epistasis and epistasis × environment interactions. Epistasis × environment interactions accounted for up to 48% of the relative fitness variation in the field. At a single field site, the impact of epistasis on relative fitness varied significantly over 2 yr, showing that epistasis × environment interactions within a location can be temporally dynamic. These results suggest that the environmental dependency of epistasis can profoundly influence the response to selection, shaping the adaptive trajectories of natural populations in complex ways, and deserves further consideration in future evolutionary studies. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  6. Maternal stress retards fetal development in mice with transcriptome-wide impact on gene expression profiles of the limb.

    PubMed

    Choe, Han Kyoung; Son, Gi Hoon; Chung, Sooyoung; Kim, Myungjin; Sun, Woong; Kim, Hyun; Geum, Dongho; Kim, Kyungjin

    2011-03-01

    The environment of a pregnant mother has a life-long impact on later life of offspring. Maternal stress is known to cause low birth weight and programs several physiological dysfunctions in offspring. However, the direct effects of maternal stress on the developing fetus remain largely unknown. The present study focused on the effect of chronic maternal stress on the developmental program and its molecular mechanisms. Pregnant mice were given 6-hour immobilization stress every day from 8.5 days post coitum. Fetal body weight was significantly decreased by maternal stress throughout development. Importantly, developmental events were retarded in the stressed fetuses. Around embryonic day 13.5 (E13.5), the developmental increment of somite numbers was delayed, although this difference recovered by E15.5. Limb bud formation and regression of interdigital webbing were also retarded by approximately 0.5 days. Subsequently, transcriptomes of developing limbs were analyzed by cDNA microarrays. Approximately, one-tenth of detected transcripts were significantly influenced by maternal stress. Q-PCR AQ analyses further demonstrated that the expression of a subset of limb development-associated genes, including Igf1, Aldh1a2, and Acta1, was changed in the stressed fetus. In conclusion, our findings suggest that maternal stress can retard limb and somite development in mice, with profound impacts on the developmental genetic program of limb.

  7. Impact of acetaminophen consumption and resistance exercise on extracellular matrix gene expression in human skeletal muscle.

    PubMed

    Patel, Shivam H; D'Lugos, Andrew C; Eldon, Erica R; Curtis, Donald; Dickinson, Jared M; Carroll, Chad C

    2017-07-01

    Acetaminophen (APAP) given during chronic exercise reduces skeletal muscle collagen and cross-linking in rats. We propose that the effect of APAP on muscle extracellular matrix (ECM) may, in part, be mediated by dysregulation of the balance between matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs). The purpose of this study was to evaluate the impact of APAP consumption during acute resistance exercise (RE) on several regulators of the ECM in human skeletal muscle. In a double-blinded, placebo-controlled, randomized crossover design, recreationally active men (n = 8, 25 ± 2 yr) performed two trials of knee extension. Placebo (PLA) or APAP (1,000 mg/6 h) was given for 24 h before and immediately following RE. Vastus lateralis biopsies were taken at baseline and 1 and 3 h post-RE. Quantitative RT-PCR was used to determine differences in mRNA expression. MMP-2, type I collagen, and type III collagen mRNA expression was not altered by exercise or APAP (P > 0.05). When compared with PLA, TIMP-1 expression was lower at 1 h post-RE during APAP conditions but greater than PLA at 3 h post-RE (P < 0.05). MMP-9 expression and protein levels were elevated at 3 h post-RE independent of treatment (P < 0.05). Lysyl oxidase expression was greater at 3 h post-RE during APAP consumption (P < 0.05) compared with PLA. MMP-2 and TIMP-1 protein was not altered by RE or APAP (P > 0.05). Phosphorylation of ERK1/2 and p38-MAPK increased (P < 0.05) with RE but was not influenced by APAP. Our findings do not support our hypothesis and suggest that short-term APAP consumption before RE has a small impact on the measured ECM molecules in human skeletal muscle following acute RE. Copyright © 2017 the American Physiological Society.

  8. Impact of gamma-glutamyl carboxylase gene polymorphisms on warfarin dose requirement: a systematic review and meta-analysis.

    PubMed

    Sun, Yifan; Wu, Zhitong; Li, Shan; Qin, Xue; Li, Taijie; Xie, Li; Deng, Yan; Chen, Junqing

    2015-04-01

    The Gamma-glutamyl carboxylase (GGCX) gene, as with Vitamin K Epoxide Reductase Complex Subunit 1 (VKORC1), CytochromeP450 Complex Subunit 14 F2 (CYP4F2) and CytochromeP450 Complex Subunit2C9 (CYP2C9), is a candidate predictor for appropriate maintenance warfarin dose. However, the association between GGCX gene polymorphisms and warfarin dose requirement is still controversial. To quantify the influence of GGCX polymorphisms on warfarin dose requirements, we performed a systematic review and meta-analysis. According to PRISRM statement (Preferred reporting items for systematic reviews and meta-analyses), a comprehensive literature search was undertaken through August 2014 looking for eligible studies in Embase, Pubmed,Web of Science and the Cochrane Library. The impact of GGCX polymorphisms on mean daily warfarin dose (MDWD) was counted by means of Z test. RevMan 5.2.7 software (developed by the Cochrane Collaboration) was applied to analyze the relationship between GGCX gene polymorphisms and warfarin dose requirements. Nineteen articles including 21 studies with a total of 6957 patients were included in the meta-analysis. Among three investigated single nucleotide polymorphisms (SNPs), rs11676382 showed higher CC genotype frequencies in Asian than those in Caucasian (97.7% vs. 86.9%); patients who were "G carriers" (that is, carried the GGCX rs11676382 CG or GG genotypes) required 27% lower warfarin dose than CC genotype [95%Confidence Interval (CI)=17%-37%, P=0.000, I(2)%=82.0 and PQ=0.000], moreover, stratified analysis by ethnicity showed similar results in Caucasian (23% lower, 95%CI=12%-33%), but not in Asian. With respect to genetic variation of rs699664 and rs121714145 SNPs, no significant impact on warfarin dose requirements were demonstrated. This meta-analysis suggested that GGCX rs11676382 polymorphism may be one of factors affecting the dose of warfarin requirement, and the effects are different in different ethnicities. Further studies about this

  9. Impacts of selective logging on inbreeding and gene flow in two Amazonian timber species with contrasting ecological and reproductive characteristics.

    PubMed

    Vinson, C C; Kanashiro, M; Harris, S A; Boshier, D H

    2015-01-01

    Selective logging in Brazil allows for the removal of up to 90% of trees above 50 cm diameter of a given timber species, independent of a species' life history characteristics or how quickly it will recover. The genetic and demographic effects of selective logging on two Amazonian timber species (Dipteryx odorata Leguminosae, Jacaranda copaia Bignoniaceae) with contrasting ecological and reproductive characteristics were assessed in the same forest. Genetic diversity and gene flow were characterized by genotyping adults and seed sampled before and after logging, using hypervariable microsatellite markers. Overall, there were no short-term genetic impacts on the J. copaia population, with commercial application of current Brazilian forest management regulations. In contrast, for D. Odorata, selective logging showed a range of genetic impacts, with a 10% loss of alleles, and reductions in siring by pollen from trees within the 546-ha study area (23-11%) and in the number of pollen donors per progeny array (2.8-1.6), illustrating the importance of the surrounding landscape. Asynchrony in flowering between D. odorata trees led to trees with no breeding partners, which could limit the species reproduction and regeneration under current regulations. The results are summarized with other published studies from the same site and the implications for forest management discussed. The different types and levels of impacts associated with each species support the idea that ecological and genetic information by species, ecological guild or reproductive group is essential in helping to derive sustainable logging guidelines for tropical forests.

  10. Antibiotic resistance genes in an urban river as impacted by bacterial community and physicochemical parameters.

    PubMed

    Zhou, Zhen-Chao; Zheng, Ji; Wei, Yuan-Yuan; Chen, Tao; Dahlgren, Randy A; Shang, Xu; Chen, Hong

    2017-09-01

    Antibiotic resistance genes (ARGs) in urban rivers are a serious public health concern in regions with poorly planned, rapid development. To gain insights into the predominant factors affecting the fate of ARGs in a highly polluted urban river in eastern China, a total of 285 ARGs, microbial communities, and 20 physicochemical parameters were analyzed for 17 sites. A total of 258 unique ARGs were detected using high-throughput qPCR, and the absolute abundance of total ARGs was positively correlated with total organic carbon and total dissolved nitrogen concentrations (P < 0.01). ARG abundance and diversity were greatly altered by microbial community structure. Variation partitioning analysis showed that the combined effects of multiple factors contributed to the profile and dissemination of ARGs, and variation of microbial communities was the major factor affecting the distribution of ARGs. The disparate distribution of some bacteria, including Bacteroides from mammalian gastrointestinal flora, Burkholderia from zoonotic infectious diseases, and Zoogloea from wastewater treatment, indicates that the urban river was strongly influenced by point-source pollution. Results imply that microbial community shifts caused by changes in water quality may lead to the spread of ARGs, and point-source pollution in urban rivers requires greater attention to control the transfer of ARGs between environmental bacteria and pathogens.

  11. Dopamine D2 gene expression interacts with environmental enrichment to impact lifespan and behavior

    PubMed Central

    Thanos, Panayotis K.; Hamilton, John; O'Rourke, Joseph R.; Napoli, Anthony; Febo, Marcelo; Volkow, Nora D.; Blum, Kenneth; Gold, Mark

    2016-01-01

    Aging produces cellular, molecular, and behavioral changes affecting many areas of the brain. The dopamine (DA) system is known to be vulnerable to the effects of aging, which regulate behavioral functions such as locomotor activity, body weight, and reward and cognition. In particular, age-related DA D2 receptor (D2R) changes have been of particular interest given its relationship with addiction and other rewarding behavioral properties. Male and female wild-type (Drd2 +/+), heterozygous (Drd2 +/−) and knockout (Drd2 −/−) mice were reared post-weaning in either an enriched environment (EE) or a deprived environment (DE). Over the course of their lifespan, body weight and locomotor activity was assessed. While an EE was generally found to be correlated with longer lifespan, these increases were only found in mice with normal or decreased expression of the D2 gene. Drd2 +/+ EE mice lived nearly 16% longer than their DE counterparts. Drd2 +/+ and Drd2 +/− EE mice lived 22% and 21% longer than Drd2 −/− EE mice, respectively. Moreover, both body weight and locomotor activity were moderated by environmental factors. In addition, EE mice show greater behavioral variability between genotypes compared to DE mice with respect to body weight and locomotor activity. PMID:26992232

  12. Reduced Impact of Pyrimethamine Drug Pressure on Plasmodium malariae Dihydrofolate Reductase Gene

    PubMed Central

    Khim, Nimol; Kim, Saorin; Bouchier, Christiane; Tichit, Magali; Ariey, Frédéric; Fandeur, Thierry; Chim, Pheaktra; Ke, Sopheakvatey; Sum, Sarorn; Man, Somnang; Ratsimbasoa, Arsène; Durand, Rémy

    2012-01-01

    Molecular investigations performed following the emergence of sulfadoxine-pyrimethamine (SP) resistance in Plasmodium falciparum have allowed the identification of the dihydrofolate reductase (DHFR) enzyme as the target of pyrimethamine. Although clinical cases of Plasmodium malariae are not usually treated with antifolate therapy, incorrect diagnosis and the high frequency of undetected mixed infections has probably exposed non-P. falciparum parasites to antifolate therapy in many areas. In this context, we aimed to assess the worldwide genetic diversity of the P. malariae dhfr gene in 123 samples collected in Africa and Asia, areas with different histories of SP use. Among the 10 polymorphic sites found, we have observed 7 new mutations (K55E, S58R, S59A, F168S, N194S, D207G, and T221A), which led us to describe 6 new DHFR proteins. All isolates from African countries were classified as wild type, while new mutations and haplotypes were recognized as exclusive to Madagascar (except for the double mutations at nucleotides 341 and 342 [S114N] found in one Cambodian isolate). Among these nonsynonymous mutations, two were likely related to pyrimethamine resistance: S58R (corresponding to C59R in P. falciparum and S58R in Plasmodium vivax; observed in one Malagasy sample) and S114N (corresponding to S108N in P. falciparum and S117N in P. vivax; observed in three Cambodian samples). PMID:22123682

  13. Reduced impact of pyrimethamine drug pressure on Plasmodium malariae dihydrofolate reductase gene.

    PubMed

    Khim, Nimol; Kim, Saorin; Bouchier, Christiane; Tichit, Magali; Ariey, Frédéric; Fandeur, Thierry; Chim, Pheaktra; Ke, Sopheakvatey; Sum, Sarorn; Man, Somnang; Ratsimbasoa, Arsène; Durand, Rémy; Ménard, Didier

    2012-02-01

    Molecular investigations performed following the emergence of sulfadoxine-pyrimethamine (SP) resistance in Plasmodium falciparum have allowed the identification of the dihydrofolate reductase (DHFR) enzyme as the target of pyrimethamine. Although clinical cases of Plasmodium malariae are not usually treated with antifolate therapy, incorrect diagnosis and the high frequency of undetected mixed infections has probably exposed non-P. falciparum parasites to antifolate therapy in many areas. In this context, we aimed to assess the worldwide genetic diversity of the P. malariae dhfr gene in 123 samples collected in Africa and Asia, areas with different histories of SP use. Among the 10 polymorphic sites found, we have observed 7 new mutations (K55E, S58R, S59A, F168S, N194S, D207G, and T221A), which led us to describe 6 new DHFR proteins. All isolates from African countries were classified as wild type, while new mutations and haplotypes were recognized as exclusive to Madagascar (except for the double mutations at nucleotides 341 and 342 [S114N] found in one Cambodian isolate). Among these nonsynonymous mutations, two were likely related to pyrimethamine resistance: S58R (corresponding to C59R in P. falciparum and S58R in Plasmodium vivax; observed in one Malagasy sample) and S114N (corresponding to S108N in P. falciparum and S117N in P. vivax; observed in three Cambodian samples).

  14. The impact of gene expression regulation on evolution of extracellular signaling pathways.

    PubMed

    Charoensawan, Varodom; Adryan, Boris; Martin, Stephen; Söllner, Christian; Thisse, Bernard; Thisse, Christine; Wright, Gavin J; Teichmann, Sarah A

    2010-12-01

    Extracellular protein interactions are crucial to the development of multicellular organisms because they initiate signaling pathways and enable cellular recognition cues. Despite their importance, extracellular protein interactions are often under-represented in large scale protein interaction data sets because most high throughput assays are not designed to detect low affinity extracellular interactions. Due to the lack of a comprehensive data set, the evolution of extracellular signaling pathways has remained largely a mystery. We investigated this question using a combined data set of physical pairwise interactions between zebrafish extracellular proteins, mainly from the immunoglobulin superfamily and leucine-rich repeat families, and their spatiotemporal expression profiles. We took advantage of known homology between proteins to estimate the relative rates of changes of four parameters after gene duplication, namely extracellular protein interaction, expression pattern, and the divergence of extracellular and intracellular protein sequences. We showed that change in expression profile is a major contributor to the evolution of signaling pathways followed by divergence in intracellular protein sequence, whereas extracellular sequence and interaction profiles were relatively more conserved. Rapidly evolving expression profiles will eventually drive other parameters to diverge more quickly because differentially expressed proteins get exposed to different environments and potential binding partners. This allows homologous extracellular receptors to attain specialized functions and become specific to tissues and/or developmental stages.

  15. Sox2 gene amplification significantly impacts overall survival in serous epithelial ovarian cancer.

    PubMed

    Belotte, Jimmy; Fletcher, Nicole M; Alexis, Mitchell; Morris, Robert T; Munkarah, Adnan R; Diamond, Michael P; Saed, Ghassan M

    2015-01-01

    Epithelial ovarian cancer (EOC) is the deadliest gynecologic cancer. Recently, the existence of ovarian cancer stem cells has been reported. Sox2, Nanog and Oct4 are key markers of "stemness". The objective of this study was to determine whether Sox2, Nanog, and Oct4 are associated with EOC and poor outcome. The expression of these markers was assessed by immunofluorescence staining and real-time RT-PCR in human EOC cell lines MDAH-2774 and SKOV-3, while the cancer genome atlas (TCGA) dataset was analyzed for associations with survival. Sox2, Nanog and Oct4 (POU5F1) were all detected by immunofluorescence staining and these results were confirmed by real-time RT-PCR. The TCGA dataset revealed a 26%, 9%, and 6% amplification of Sox2, Nanog and POU5F1, respectively. Additionally, K-M survival analyses showed a significant median overall survival difference (41 versus 48.3 months, P = .01) for Sox2 amplification, but not for Nanog (44.1 versus 36.2 months, P > .05) and POU5F1 (43.5 versus 45.0 months, P > .05). Our results suggest that Sox2 gene amplification significantly influences overall survival. © The Author(s) 2014.

  16. Obesity risk prediction among women of Upper Egypt: The impact of serum vaspin and vaspin rs2236242 gene polymorphism.

    PubMed

    Abdel Ghany, Soad M; Sayed, Ayat A; El-Deek, Sahar E M; ElBadre, Hala M; Dahpy, Marwa A; Saleh, Medhat A; Sharaf El-Deen, Hanan; Mustafa, Mohamed H

    2017-08-30

    Vaspin is an adipokine that is potentially linking obesity, insulin resistance, metabolic syndrome and type-2 diabetes. The present study aimed to investigate the impact of vaspin rs2236242 gene polymorphism on the risk of obesity, diabetes, their metabolic traits, and serum vaspin levels in a sample of Upper Egyptian women. A total of 224 subjects, 112 obese (62 non diabetics, 50 diabetics) and 112 controls were included in this case control study. Vaspin gene rs2236242 polymorphism was performed using tetra-amplification refractory mutation system-polymerase chain reaction (T-ARMS-PCR) and serum vaspin levels were estimated by ELISA. The minor (A) allele of vaspin rs2236242 gene polymorphism was significantly lower in obese (30.8%) than controls (43.7%) (P=0.005). The protective effect was evident in dominant and recessive inheritance models (TT vs TA+AA, P=0.004 and TT+TA vs AA, P=0.036). After adjusting genotypes for diabetes there were no significant association between vaspin rs2236242 gene polymorphism and obesity but significant association was maintained in the obese diabetics. Vaspin serum levels were found to be lower in minor protective (AA) genotype carriers than the other two genotypes (P<0.001). In the mean-time serum vaspin levels were significantly higher in obese diabetics and non-diabetics than controls (P<0.001 each).There were significant positive correlations between vaspin levels and hs-CRP, cholesterol, LDL-C, fasting glucose, HOMA-IR, insulin, and ALT values (P<0.05 each) and a negative correlation with HDL-C (P<0.01). The minor A allele of vaspin rs2236242 polymorphism plays a protective role against obesity and diabetes but this relation is largely ascribed to its effect on insulin resistance. The serum vaspin concentration was lower in minor protective allele carriers. To the best of our knowledge, this is the first study of vaspin SNP in Upper Egyptian women. The entire understanding of vaspin intimate mechanistic action might enable

  17. Impacts of environmental stress on growth, secondary metabolite biosynthetic gene clusters and metabolite production of xerotolerant/xerophilic fungi.

    PubMed

    Medina, Angel; Schmidt-Heydt, Markus; Rodríguez, Alicia; Parra, Roberto; Geisen, Rolf; Magan, Naresh

    2015-08-01

    This paper examines the impact that single and interacting environmental stress factors have on tolerance mechanisms, molecular ecology and the relationship with secondary metabolite production by a group of mycotoxigenic species of economic importance. Growth of these fungi (Aspergillus flavus, A.ochraceus, A.carbonarius, Penicillium nordicum and P. verrucosum) is influenced by water and temperature interactions and type of solute used to induce water stress. Such abiotic stresses are overcome by the synthesis of increased amounts of low molecular weight sugar alcohols, especially glycerol and erythritol, to enable them to remain active under abiotic stress. This is accompanied by increased expression of sugar transporter genes, e.g., in A. flavus, which provides the nutritional means of tolerating such stress. The optimum conditions of water activity (a w) × temperature stress for growth are often different from those for secondary metabolite production. The genes for toxin production are clustered together and their relative expression is influenced by abiotic interacting stress factors. For example., A. flavus synthesises aflatoxins under water stress in non-ionic solutes. In contrast, P. nordicum specifically occupies a high salt (0.87 a w = 22% NaCl) niche such as cured meats, and produces ochratoxin A (OTA). There is differential and temporal expression of the genes in the secondary metabolite clusters in response to a w × temperature stress. We have used a microarray and integrated data on growth, relative expression of key genes in the biosynthetic pathways for secondary metabolite production and toxin production using a mixed growth model. This was used to correlate these factors and predict the toxin levels produced under different abiotic stress conditions. This system approach to integrate these different data sets and model the relationships could be a powerful tool for predicting the relative toxin production under extreme stress conditions

  18. Peroxisome biogenesis in mammalian cells: The impact of genes and environment.

    PubMed

    Farr, Rebecca L; Lismont, Celien; Terlecky, Stanley R; Fransen, Marc

    2016-05-01

    The initiation and progression of many human diseases are mediated by a complex interplay of genetic, epigenetic, and environmental factors. As all diseases begin with an imbalance at the cellular level, it is essential to understand how various types of molecular aberrations, metabolic changes, and environmental stressors function as switching points in essential communication networks. In recent years, peroxisomes have emerged as important intracellular hubs for redox-, lipid-, inflammatory-, and nucleic acid-mediated signaling pathways. In this review, we focus on how nature and nurture modulate peroxisome biogenesis and function in mammalian cells. First, we review emerging evidence that changes in peroxisome activity can be linked to the epigenetic regulation of cell function. Next, we outline how defects in peroxisome biogenesis may directly impact cellular pathways involved in the development of disease. In addition, we discuss how changes in the cellular microenvironment can modulate peroxisome biogenesis and function. Finally, given the importance of peroxisome function in multiple aspects of health, disease, and aging, we highlight the need for more research in this still understudied field. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. The Psychosocial Impact of Carrying a Debated Variant in the GLA Gene.

    PubMed

    Macklin, Sarah; Laney, Dawn; Lisi, Emily; Atherton, Andrea; Smith, Elizabeth

    2017-08-10

    The clinical significance of the c.427G>A (p.A143T) variant in GLA is a topic of debate within the lysosomal storage disease community. A review of the literature and published case reports found the clinical impact of the variant to range from classic Fabry symptoms to healthy unaffected males with normal alpha- galactosidase enzyme levels, leaving clinicians unsure of how to manage these individuals. As the number of states testing for Fabry disease on their newborn screening panel has increased, more people with this variant are being identified. The goal of this project was to learn how the uncertainty surrounding the clinical significance of the p.A143T variant affects those with this change. A self-response questionnaire was developed to explore this topic. In addition to evaluating participant feelings, the questionnaire explored individuals' beliefs regarding the pathogenicity of the variant. Results suggest that people have diverse feelings regarding reclassification of the p.A143T variant. Around half of those surveyed reported feeling frustrated by the lack of clear information. Despite the ambiguity regarding the health consequences of this variant, many participants felt that knowing this result helps guide medical management.

  20. Converging evidence for an impact of a functional NOS gene variation on anxiety-related processes

    PubMed Central

    Haaker, Jan; Glotzbach-Schoon, Evelyn; Schümann, Dirk; Andreatta, Marta; Mechias, Marie-Luise; Raczka, Karolina; Gartmann, Nina; Büchel, Christian; Mühlberger, Andreas; Pauli, Paul; Reif, Andreas; Kalisch, Raffael; Lonsdorf, Tina B.

    2016-01-01

    Being a complex phenotype with substantial heritability, anxiety and related phenotypes are characterized by a complex polygenic basis. Thereby, one candidate pathway is neuronal nitric oxide (NO) signaling, and accordingly, rodent studies have identified NO synthase (NOS-I), encoded by NOS1, as a strong molecular candidate for modulating anxiety and hippocampus-dependent learning processes. Using a multi-dimensional and -methodological replication approach, we investigated the impact of a functional promoter polymorphism (NOS1-ex1f-VNTR) on human anxiety-related phenotypes in a total of 1019 healthy controls in five different studies. Homozygous carriers of the NOS1-ex1f short-allele displayed enhanced trait anxiety, worrying and depression scores. Furthermore, short-allele carriers were characterized by increased anxious apprehension during contextual fear conditioning. While autonomous measures (fear-potentiated startle) provided only suggestive evidence for a modulatory role of NOS1-ex1f-VNTR on (contextual) fear conditioning processes, neural activation at the amygdala/anterior hippocampus junction was significantly increased in short-allele carriers during context conditioning. Notably, this could not be attributed to morphological differences. In accordance with data from a plethora of rodent studies, we here provide converging evidence from behavioral, subjective, psychophysiological and neuroimaging studies in large human cohorts that NOS-I plays an important role in anxious apprehension but provide only limited evidence for a role in (contextual) fear conditioning. PMID:26746182

  1. Impact of REV-ERB alpha gene polymorphisms on obesity phenotypes in adult and adolescent samples.

    PubMed

    Goumidi, L; Grechez, A; Dumont, J; Cottel, D; Kafatos, A; Moreno, L A; Molnar, D; Moschonis, G; Gottrand, F; Huybrechts, I; Dallongeville, J; Amouyel, P; Delaunay, F; Meirhaeghe, A

    2013-05-01

    REV-ERBα has been shown to regulate adipogenesis and lipid metabolism as well as to link the circadian timing system to whole body metabolic homeostasis. We thus tested whether polymorphisms in REV-ERBα could be associated with metabolic phenotypes in human population samples. We analyzed the associations between 5 REV-ERBα polymorphisms and anthropometric (body weight, body mass index (BMI), waist and hip circumferences), biochemical (plasma lipid, glucose and insulin levels) and clinical (systolic and diastolic blood pressure) variables in three population-based studies (MONICA Lille n=1155 adults, MONA LISA Lille n=1170 adults and HELENA n=1155 adolescents). We assessed in vitro, the potential influence of one REV-ERBα polymorphism in transient transfection assays using two different cell lines. We observed significant and consistent associations between the T minor allele of the REV-ERBα rs2071427 polymorphism (located in intron 1) and higher BMI (mean allele effect=+0.33 kg m(-2)) in the MONICA Lille (P=0.02), MONA LISA (P=0.02) and HELENA (P=0.03) studies. The odds ratios for obesity associated with this allele were 1.67 (1.00-2.79) (P=0.05) in MONICA Lille, 1.29 (1.01-1.65) (P=0.04) in MONA LISA Lille and the odds ratio for overweight was 1.48 (1.08-2.03) (P=0.01) in HELENA. In transfection experiments in human hepatocyte-derived cell lines, the REV-ERBα intron 1 directed the transcription of a luciferase reporter gene independently of the rs2071427 polymorphism. Our results suggest that the REV-ERBα rs2071427 polymorphism modulates body fat mass in both adult and young people.

  2. Economic Impact of Gene Expression Profiling in Patients with Early-Stage Breast Cancer in France

    PubMed Central

    Katz, Gregory; Romano, Olivier; Foa, Cyril; Vataire, Anne-Lise; Chantelard, Jean-Victor; Hervé, Robert; Barletta, Hugues; Durieux, Axel; Martin, Jean-Pierre; Salmon, Rémy

    2015-01-01

    Background and Aims The heterogeneous nature of breast cancer can make decisions on adjuvant chemotherapy following surgical resection challenging. Oncotype DX is a validated gene expression profiling test that predicts the likelihood of adjuvant chemotherapy benefit in early-stage breast cancer. The aim of this study is to determine the costs of chemotherapy in private hospitals in France, and evaluate the cost-effectiveness of Oncotype DX from national insurance and societal perspectives. Methods A multicenter study was conducted in seven French private hospitals, capturing retrospective data from 106 patient files. Cost estimates were used in conjunction with a published Markov model to assess the cost-effectiveness of using Oncotype DX to inform chemotherapy decision making versus standard care. Sensitivity analyses were performed. Results The cost of adjuvant chemotherapy in private hospitals was estimated at EUR 8,218 per patient from a national insurance perspective and EUR 10,305 from a societal perspective. Cost-effectiveness analysis indicated that introducing Oncotype DX improved life expectancy (+0.18 years) and quality-adjusted life expectancy (+0.17 QALYs) versus standard care. Oncotype DX was found cost-effective from a national insurance perspective (EUR 2,134 per QALY gained) and cost saving from a societal perspective versus standard care. Inclusion of lost productivity costs in the modeling analysis meant that costs for eligible patients undergoing Oncotype DX testing were on average EUR 602 lower than costs for those receiving standard care. Conclusions As Oncotype DX was found both cost and life-saving from a societal perspective, the test was considered to be dominant to standard care. However, the delay in coverage has the potential to erode the quality of the French healthcare system, thus depriving patients of technologies that could improve clinical outcomes and allow healthcare professionals to better allocate hospital resources to

  3. Impact of obesity on IL-12 family gene expression in insulin responsive tissues.

    PubMed

    Nam, Heesun; Ferguson, Bradley S; Stephens, Jacqueline M; Morrison, Ron F

    2013-01-01

    Mounting evidence has established a role for chronic inflammation in the development of obesity-induced insulin resistance, as genetic ablation of pro-inflammatory cytokines and chemokines elevated in obesity improves insulin signaling in vitro and in vivo. Recent evidence further highlights interleukin (IL)-12 family cytokines as prospective inflammatory mediators linking obesity to insulin resistance. In this study, we present empirical evidence demonstrating that IL-12 family related genes are expressed and regulated in insulin-responsive tissues under conditions of obesity. First, we report that respective mRNAs for each of the known members of this cytokine family are expressed within detectable ranges in WAT, skeletal muscle, liver and heart. Second, we show that these cytokines and their cognate receptors are divergently regulated with genetic obesity in a tissue-specific manner. Third, we demonstrate that select IL-12 family cytokines are regulated in WAT in a manner that is dependent on the developmental stage of obesity as well as the inflammatory progression associated with obesity. Fourth, we report that respective mRNAs for IL-12 cytokines and receptors are also expressed and divergently regulated in cultured adipocytes under conditions of inflammatory stress. To our knowledge, this report is the first study to systemically evaluated mRNA expression of all IL-12 family cytokines and receptors in any tissue under conditions of obesity highlighting select family members as potential mediators linking excess nutrient intake to metabolic diseases such as insulin resistance, diabetes and heart disease. Copyright © 2012 Elsevier B.V. All rights reserved.

  4. The VEGF gene polymorphism impacts brain volume and arterial blood volume.

    PubMed

    Takeuchi, Hikaru; Tomita, Hiroaki; Taki, Yasuyuki; Kikuchi, Yoshie; Ono, Chiaki; Yu, Zhiqian; Sekiguchi, Atsushi; Nouchi, Rui; Kotozaki, Yuka; Nakagawa, Seishu; Miyauchi, Carlos Makoto; Iizuka, Kunio; Yokoyama, Ryoichi; Shinada, Takamitsu; Yamamoto, Yuki; Hanawa, Sugiko; Araki, Tsuyoshi; Kunitoki, Keiko; Sassa, Yuko; Kawashima, Ryuta

    2017-04-12

    Vascular endothelial growth factor (VEGF) plays a critical role in the angiogenesis and proliferation of various types of cells such as neurons, astroglia, and endothelial cells in the brain. A common polymorphism in the VEGF gene (-2578 C/A) is associated with circulating VEGF levels, cancers and Alzheimer's disease. Nonetheless, the effects of this polymorphism on normal human brain volume, arterial blood volume, and blood supply remain unclear. In this study, the effects of this polymorphism on the total gray matter volume (TGMV) and total white matter volume (TWMV) using T1-weighted structural images and the total arterial blood volume (TABV) and mean cerebral blood flow (mCBF) during rest using arterial spin labeling (ASL) in 765 young adult humans were investigated. Voxel-by-voxel whole-brain analyses of these measures were also performed. Multiple regression analyses with age and sex as covariates revealed that the VEGF genotype (number of C alleles) was significantly and positively correlated with TGMV, TWMV, and TABV as well as with regional gray and white matter volumes in widespread areas and regional arterial blood volume in some areas with high arterial blood volume. However, these regional associations were not seen when the corresponding global signal was included as a covariate in the multiple regression analyses, indicating that we failed to obtain evidence of region-specific associations between these brain measures and the genotype. The results suggest that the VEGF-2578C allele, is associated with changes in the vascular system that lead to increased blood volume and larger brain volume. Hum Brain Mapp, 2017. © 2017 Wiley Periodicals, Inc.

  5. The Impact and Cost of Scaling up GeneXpert MTB/RIF in South Africa

    PubMed Central

    Meyer-Rath, Gesine; Schnippel, Kathryn; Long, Lawrence; MacLeod, William; Sanne, Ian; Stevens, Wendy; Pillay, Sagie; Pillay, Yogan; Rosen, Sydney

    2012-01-01

    Objective We estimated the incremental cost and impact on diagnosis and treatment uptake of national rollout of Xpert MTB/RIF technology (Xpert) for the diagnosis of pulmonary TB above the cost of current guidelines for the years 2011 to 2016 in South Africa. Methods We parameterised a population-level decision model with data from national-level TB databases (n = 199,511) and implementation studies. The model follows cohorts of TB suspects from diagnosis to treatment under current diagnostic guidelines or an algorithm that includes Xpert. Assumptions include the number of TB suspects, symptom prevalence of 5.5%, annual suspect growth rate of 10%, and 2010 public-sector salaries and drug and service delivery costs. Xpert test costs are based on data from an in-country pilot evaluation and assumptions about when global volumes allowing cartridge discounts will be reached. Results At full scale, Xpert will increase the number of TB cases diagnosed per year by 30%–37% and the number of MDR-TB cases diagnosed by 69%–71%. It will diagnose 81% of patients after the first visit, compared to 46% currently. The cost of TB diagnosis per suspect will increase by 55% to USD 60–61 and the cost of diagnosis and treatment per TB case treated by 8% to USD 797–873. The incremental capital cost of the Xpert scale-up will be USD 22 million and the incremental recurrent cost USD 287–316 million over six years. Conclusion Xpert will increase both the number of TB cases diagnosed and treated and the cost of TB diagnosis. These results do not include savings due to reduced transmission of TB as a result of earlier diagnosis and treatment initiation. PMID:22693561

  6. Moderate nutrient restriction influences transcript abundance of genes impacting production efficiencies of beef cattle in fetal liver, muscle, and cerebrum by d 50 of gestation

    USDA-ARS?s Scientific Manuscript database

    We hypothesized that a moderate maternal nutrient restriction during the first 50 d of gestation in beef heifers would affect transcript abundance of genes impacting production efficiency phenotypes in fetal liver, muscle, and cerebrum. Fourteen Angus-cross heifers were estrus synchronized and assig...

  7. Impact of a single session of intermittent pneumatic leg compressions on skeletal muscle and isolated artery gene expression in rats

    PubMed Central

    Roseguini, Bruno T.; Arce-Esquivel, Arturo A.; Newcomer, Sean C.

    2011-01-01

    Intermittent pneumatic leg compressions (IPC) have proven to be an effective noninvasive approach for treatment of patients with claudication, but the mechanisms underlying the clinical benefits remain elusive. In the present study, a rodent model of claudication produced by bilateral ligation of the femoral artery was used to investigate the acute impact of a single session of IPC (150 min) on hemodynamics, skeletal muscle (tibialis anterior), and isolated collateral artery (perforating artery) expression of a subset of genes associated with inflammation and vascular remodeling. In addition, the effect of compression frequency (15 vs. 3 compressions/min) on the expression of these factors was studied. In ligated animals, IPC evoked an increase of monocyte chemoattractant protein-1 (MCP-1) and cytokine-induced neutrophil chemoattractant 1 (CXCL1) mRNA (P < 0.01) and immunostaining (P < 0.05), as well as a minor increase in VEGF immunostaining in the muscle endomysium 150 min postintervention. Further, collateral arteries from these animals showed an increased expression of MCP-1 (approximately twofold, P = 0.02). These effects were most evident in the group exposed to the high-frequency protocol (15 compressions/min). In contrast, IPC in sham-operated control animals evoked a modest initial upregulation of VEGF (P = 0.01), MCP-1 (P = 0.02), and CXCL1 (P = 0.03) mRNA in the muscle without concomitant changes in protein levels. No changes in gene expression were observed in arteries isolated from sham animals. In conclusion, IPC acutely up-regulates the expression of important factors involved in vascular remodeling in the compressed muscle and collateral arteries in a model of hindlimb ischemia. These effects appear to be dependent on the compression frequency, such that a high compression frequency (15 compressions/min) evokes more consistent and robust effects compared with the frequency commonly employed clinically to treat patients with claudication (3

  8. End-to-end gene fusions and their impact on the production of multifunctional biomass degrading enzymes

    SciTech Connect

    Rizk, Mazen; Antranikian, Garabed; Elleuche, Skander

    2012-11-09

    Highlights: Black-Right-Pointing-Pointer Multifunctional enzymes offer an interesting approach for biomass degradation. Black-Right-Pointing-Pointer Size and conformation of separate constructs play a role in the effectiveness of chimeras. Black-Right-Pointing-Pointer A connecting linker allows for maximal flexibility and increased thermostability. Black-Right-Pointing-Pointer Genes with functional similarities are the best choice for fusion candidates. -- Abstract: The reduction of fossil fuels, coupled with its increase in price, has made the search for alternative energy resources more plausible. One of the topics gaining fast interest is the utilization of lignocellulose, the main component of plants. Its primary constituents, cellulose and hemicellulose, can be degraded by a series of enzymes present in microorganisms, into simple sugars, later used for bioethanol production. Thermophilic bacteria have proven to be an interesting source of enzymes required for hydrolysis since they can withstand high and denaturing temperatures, which are usually required for processes involving biomass degradation. However, the cost associated with the whole enzymatic process is staggering. A solution for cost effective and highly active production is through the construction of multifunctional enzyme complexes harboring the function of more than one enzyme needed for the hydrolysis process. There are various strategies for the degradation of complex biomass ranging from the regulation of the enzymes involved, to cellulosomes, and proteins harboring more than one enzymatic activity. In this review, the construction of multifunctional biomass degrading enzymes through end-to-end gene fusions, and its impact on production and activity by choosing the enzymes and linkers is assessed.

  9. Prognostic impact of Wilms tumor gene mutations in Egyptian patients with acute myeloid leukemia with normal karyotype.

    PubMed

    Zidan, Magda Abdel Aziz; Kamal Shaaban, Howyda M; Elghannam, Doaa M

    2014-07-01

    The Wilms' tumor (WT1) gene mutations were detected in patients with most forms of acute leukemia. However, the biological significance and the prognostic impact of WT1 mutation in Egyptian patients with acute myeloid leukemia with normal karyotype (AML-NK) are still uncertain. We aimed to evaluate the incidence and clinical relevance of WT1 gene mutations in acute myeloid leukemia with normal karyotype (AML-NK). Exons 7 and 9 of WT1 were screened in samples from 216 adult NK-AML using polymerase chain reaction single-strand conformation polymorphism techniques. Twenty-three patients (10.6%) harbored WT1 mutations. Younger ages and higher marrow blasts were significantly associated with WT1 mutations (P = 0.006 and 0.003 respectively). Complete remission rates were significantly lower in patients with WT1 mutations than those with WT1 wild-type (P = 0.015). Resistance, relapse, and mortality rates were significantly higher in patients with WT1 mutations than those without (P = 0.041, 0.016, and 0.008 respectively). WT1 mutations were inversely associated with NPM1 mutations (P = 0.007). Patients with WT1 mutations had worse disease-free survival (P < 0.001) and overall survival (P < 0.001) than patients with WT1 wild-type. In multivariable analyses, WT1 mutations independently predicted worse DFS (P < 0.001; hazard ratio [HR] 0.036) and overall survival (P = 0.001; HR = 0.376) when controlling for age, total leukocytic count (TLC), and NPM1 mutational status. In conclusion, WT1 mutations are a negative prognostic indicator in intensively treated patients with AML-NK, may be a part of molecularly based risk assessment and risk-adapted treatment stratification of patients with AML-NK.

  10. Efficient removal of antibiotics in surface-flow constructed wetlands, with no observed impact on antibiotic resistance genes.

    PubMed

    Berglund, Björn; Khan, Ghazanfar Ali; Weisner, Stefan E B; Ehde, Per Magnus; Fick, Jerker; Lindgren, Per-Eric

    2014-04-01

    Recently, there have been growing concerns about pharmaceuticals including antibiotics as environmental contaminants. Antibiotics of concentrations commonly encountered in wastewater have been suggested to affect bacterial population dynamics and to promote dissemination of antibiotic resistance. Conventional wastewater treatment processes do not always adequately remove pharmaceuticals causing environmental dissemination of low levels of these compounds. Using constructed wetlands as an additional treatment step after sewage treatment plants have been proposed as a cheap alternative to increase reduction of wastewater contaminants, however this means that the natural microbial community of the wetlands becomes exposed to elevated levels of antibiotics. In this study, experimental surface-flow wetlands in Sweden were continuously exposed to antibiotics of concentrations commonly encountered in wastewater. The aim was to assess the antibiotic removal efficiency of constructed wetlands and to evaluate the impact of low levels of antibiotics on bacterial diversity, resistance development and expression in the wetland bacterial community. Antibiotic concentrations were measured using liquid chromatography-mass spectrometry and the effect on the bacterial diversity was assessed with 16S rRNA-based denaturing gradient gel electrophoresis. Real-time PCR was used to detect and quantify antibiotic resistance genes and integrons in the wetlands, during and after the exposure period. The results indicated that the antibiotic removal efficiency of constructed wetlands was comparable to conventional wastewater treatment schemes. Furthermore, short-term treatment of the constructed wetlands with environmentally relevant concentrations (i.e. 100-2000 ng×l(-1)) of antibiotics did not significantly affect resistance gene concentrations, suggesting that surface-flow constructed wetlands are well-suited for wastewater treatment purposes. Copyright © 2014 Elsevier B.V. All rights

  11. A gene to organism approach—assessing the impact of environmental pollution in eelpout (Zoarces viviparus) females and larvae

    PubMed Central

    Carney Almroth, Bethanie; Albertsson, Eva; Coltellaro, Mariateresa; Bignell, John Paul; Hanson, Niklas; Scarcelli, Vittoria; Fagerholm, Björn; Parkkonen, Jari; Wijkmark, Emma; Frenzilli, Giada; Förlin, Lars; Sturve, Joachim

    2015-01-01

    Abstract A broad biomarker approach was applied to study the effects of marine pollution along the Swedish west coast using the teleost eelpout (Zoarces viviparus) as the sentinel species. Measurements were performed on different biological levels, from the molecular to the organismal, including measurements of messenger RNA (mRNA), proteins, cellular and tissue changes, and reproductive success. Results revealed that eelpout captured in Stenungsund had significantly higher hepatic ethoxyresorufin O‐deethylase activity, high levels of both cytochrome P4501A and diablo homolog mRNA, and high prevalence of dead larvae and nuclear damage in erythrocytes. Eelpout collected in Göteborg harbor displayed extensive macrovesicular steatosis, whereby the majority of hepatocytes were affected throughout the liver, which could indicate an effect on lipid metabolism. Results also indicate that eelpouts collected at polluted sites might have an affected immune system, with lower mRNA expression of genes involved in the innate immune system and a higher number of lymphocytes. Biomarker assessment also was performed on livers dissected from unborn eelpout larvae collected from the ovary of the females. No significant differences were noted, which might indicate that the larvae to some extent are protected from effects of environmental pollutants. In conclusion, usage of the selected set of biological markers, covering responses from gene to organism, has demonstrated site‐specific biomarker patterns that provided a broad and comprehensive picture of the impact of environmental stressors. Environ Toxicol Chem 2015;34:1511–1523. © 2015 The Authors. Published by SETAC. PMID:25663503

  12. Impact of CUX2 on the Female Mouse Liver Transcriptome: Activation of Female-Biased Genes and Repression of Male-Biased Genes

    PubMed Central

    Conforto, Tara L.; Zhang, Yijing; Sherman, Jennifer

    2012-01-01

    The growth hormone-regulated transcription factors STAT5 and BCL6 coordinately regulate sex differences in mouse liver, primarily through effects in male liver, where male-biased genes are upregulated and many female-biased genes are actively repressed. Here we investigated whether CUX2, a highly female-specific liver transcription factor, contributes to an analogous regulatory network in female liver. Adenoviral overexpression of CUX2 in male liver induced 36% of female-biased genes and repressed 35% of male-biased genes. In female liver, CUX2 small interfering RNA (siRNA) preferentially induced genes repressed by adenovirus expressing CUX2 (adeno-CUX2) in male liver, and it preferentially repressed genes induced by adeno-CUX2 in male liver. CUX2 binding in female liver chromatin was enriched at sites of male-biased DNase hypersensitivity and at genomic regions showing male-enriched STAT5 binding. CUX2 binding was also enriched near genes repressed by adeno-CUX2 in male liver or induced by CUX2 siRNA in female liver but not at genes induced by adeno-CUX2, indicating that CUX2 binding is preferentially associated with gene repression. Nevertheless, direct CUX2 binding was seen at several highly female-specific genes that were positively regulated by CUX2, including A1bg, Cyp2b9, Cyp3a44, Tox, and Trim24. CUX2 expression and chromatin binding were high in immature male liver, where repression of adult male-biased genes and expression of adult female-biased genes are common, suggesting that the downregulation of CUX2 in male liver at puberty contributes to the developmental changes establishing adult patterns of sex-specific gene expression. PMID:22966202

  13. [Impact of Pax-8 gene interference on mitochondrial function and cardiomyocyte apoptosis].

    PubMed

    Dai, Xiao-chun; Zhou, Xi; Huang, Xiao-yan; Wang, Liang-guo; Lin, Su; Yang, De-ye

    2013-01-01

    To observe the effects of paired box gene 8 (Pax-8) silencing by RNA interference on mitochondrial function and cardiomyocytes apoptosis. The cultured H9C2 (2-1) myocytes were divided into 3 groups: short interference RNA targeting Pax-8 (Pax-8 siRNA) group, non-specific siRNA group as the negative control (NC siRNA), and blank control group (BC siRNA). Fluorescence spectrophotometry was used to detect the activity of caspase-3. RT-PCR was performed to detect mRNA expression of Bcl2 and Bax. The protein expression of Bcl2, Bax and cytoplasm of Cytochrome was examined by Western blot. Changes of ΔΨm were detected by flow cytometry.ΔΨm with JC-1 monomer/polymer ratio was calculated for measuring mitochondrial depolarization proportion. Compared to NC siRNA and BC siRNA group (0.075 ± 0.021, 0.072 ± 0.019), the activity of caspase-3 in Pax-8 siRNA group (0.167 ± 0.012) was significantly increased (P < 0.05); Bcl2 mRNA and protein expression in Pax-8 siRNA group (0.61 ± 0.06, 0.94 ± 0.11) were significantly downregulated compared with NC siRNA group (0.90 ± 0.070, 1.39 ± 0.15) and BC siRNA group (0.94 ± 0.087, 1.49 ± 0.20) (P < 0.05); Bax mRNA and protein expression in Pax-8 siRNA group (1.05 ± 0.10, 1.25 ± 0.12) were markedly upregulated compared with NC siRNA group (0.72 ± 0.03, 0.99 ± 0.12) and BC siRNA group (0.64 ± 0.03, 0.92 ± 0.06), P < 0.05; cytosolic cytochrome expression in Pax-8 siRNA group (0.75 ± 0.14) was significantly upregulated compared with NC siRNA group (0.51 ± 0.06) and BC siRNA group (0.48 ± 0.07) (P < 0.05); JC-1 monomer/polymer ratio in Pax-8 siRNA group (0.163 ± 0.011) was significantly increased compared with NC siRNA group (0.092 ± 0.015) and BC siRNA group (0.072 ± 0.025) (P < 0.05) indicating mitochondrial membrane potential was significantly reduced in Pax-8 siRNA group. Above parameters were similar between NC siRNA group and BC siRNA group (P > 0.05). Inhibiting Pax-8 results in enhanced cardiomyocytes apoptosis

  14. Impact of the 3D Microenvironment on Phenotype, Gene Expression, and EGFR Inhibition of Colorectal Cancer Cell Lines

    PubMed Central

    Deenen, René; Schmidt, Stephan; Messner, Isabelle; Schäfer, Karl-Ludwig; Baldus, Stephan E.; Huckenbeck, Wolfgang; Piekorz, Roland P.; Knoefel, Wolfram T.; Krieg, Andreas; Stoecklein, Nikolas H.

    2013-01-01

    Three-dimensional (3D) tumor cell cultures grown in laminin-rich-extracellular matrix (lrECM) are considered to reflect human tumors more realistic as compared to cells grown as monolayer on plastic. Here, we systematically investigated the impact of ECM on phenotype, gene expression, EGFR signaling pathway, and on EGFR inhibition in commonly used colorectal cancer (CRC) cell lines. LrECM on-top (3D) culture assays were performed with the CRC cell lines SW-480, HT-29, DLD-1, LOVO, CACO-2, COLO-205 and COLO-206F. Morphology of lrECM cultivated CRC cell lines was determined by phase contrast and confocal laser scanning fluorescence microscopy. Proliferation of cells was examined by MTT assay, invasive capacity of the cell lines was assayed using Matrigel-coated Boyden chambers, and migratory activity was determined employing the Fence assay. Differential gene expression was analyzed at the transcriptional level by the Agilent array platform. EGFR was inhibited by using the specific small molecule inhibitor AG1478. A specific spheroid growth pattern was observed for all investigated CRC cell lines. DLD-1, HT-29 and SW-480 and CACO-2 exhibited a clear solid tumor cell formation, while LOVO, COLO-205 and COLO-206F were characterized by forming grape-like structures. Although the occurrence of a spheroid morphology did not correlate with an altered migratory, invasive, or proliferative capacity of CRC cell lines, gene expression was clearly altered in cells grown on lrECM as compared to 2D cultures. Interestingly, in KRAS wild-type cell lines, inhibition of EGFR was less effective in lrECM (3D) cultures as compared to 2D cell cultures. Thus, comparing both 2D and 3D cell culture models, our data support the influence of the ECM on cancer growth. Compared to conventional 2D cell culture, the lrECM (3D) cell culture model offers the opportunity to investigate permanent CRC cell lines under more physiological conditions, i.e. in the context of molecular therapeutic targets

  15. Impact of estrogen receptor gene polymorphisms and mRNA levels on obesity and lipolysis – a cohort study

    PubMed Central

    Nilsson, Maria; Dahlman, Ingrid; Jiao, Hong; Gustafsson, Jan-Åke; Arner, Peter; Dahlman-Wright, Karin

    2007-01-01

    Background The estrogen receptors α and β (ESR1, ESR2) have been implicated in adiposity, lipid metabolism and feeding behaviour. In this report we analyse ESR1 and ESR2 gene single nucleotide polymorphisms (SNPs) for association with obesity. We also relate adipose tissue ESR1 mRNA levels and ESR1 SNPs to adipocyte lipolysis and lipogenesis phenotypes. Methods 23 ESR1 and 11 ESR2 tag-SNPs, covering most of the common haplotype variation in each gene according to HAPMAP data, were analysed by Chi2 for association with obesity in a cohort comprising 705 adults with severe obesity and 402 lean individuals. Results were replicated in a cohort comprising 837 obese and 613 lean subjects. About 80% of both cohorts comprised women and 20% men. Adipose tissue ESR1 mRNA was quantified in 122 women and related to lipolysis and lipogenesis by multiple regression. ESR1 SNPs were analysed for association with adipocyte lipolysis and lipogenesis phenotypes in 204 obese women by simple regression. Results No ESR1 SNP was associated with obesity. Five ESR2 SNPs displayed nominal significant allelic association with obesity in women and one in men. The two ESR2 SNPs associated with obesity with nominal P value < 0.01 were genotyped in a second cohort where no association with obesity was observed. There was an inverse correlation between ESR1 mRNA levels in abdominal subcutaneous (sc) adipose tissue and basal lipolysis, as well as responsiveness to adrenoceptor agonists independent of age and BMI (P value 0.009–0.045). ESR1 rs532010 was associated with lipolytic sensitivity to noradrenaline (nominal P value 0.012), and ESR1 rs1884051 with responsiveness to the non-selective beta-adrenoceptor agonist isoprenaline (nominal P value 0.05). These associations became non-significant after Bonferroni correction. Conclusion ESR1 gene alleles are unlikely to be a major cause of obesity in women. A minor importance of ESR2 on severe obesity cannot be excluded. The inverse correlation

  16. The impact of RNA standardization and heterogeneous gene expression on the results of cDNA array of human breast carcinoma.

    PubMed

    Khoshnoud, Reza; He, Qimin; Sylván, Maria; Khoshnoud, Aida; Ivarsson, Madleen; Fornander, Tommy; Bergh, Jonas; Frisell, Jan; Rutqvist, Lars-Erik; Skog, Sven

    2010-05-01

    cDNA microarray is an established technique. However, difficulties such as handling tissue samples under RNase-free conditions, the heterogeneous tumor composition, i.e. non-malignant versus malignant cells and different pathologic types of malignant cells, and lack of appropriate reference may limit the potentially benefit of this method in clinical use. In this study, we examined how standardization of gene expression to total mg RNA or mg tissue and tumor heterogeneity affect the final results. We found that the gene expression of human breast tumors was approximately 9 times higher in malignant tissue as compared to the non-malignant tissue when expressed per total mg RNA, but approximately 40 times higher when expressed per mg tissue. Genes that were expected to act as housekeeping genes (PUC18, RPL and beta-actin) varied between different parts of the tumor and also between non-malignant and malignant tissues, excluding them as reference genes. We also found that the gene expression differed in various parts of the breast tumor, probably due to a mixture of different types of cells, i.e. non-malignant and malignant cells. To find out if the variations in the gene expression were due to cell heterogeneity we used microdissection to collect malignant cells separately. We found that the gene expression was markedly different in the isolated malignant cells as compared to the gene expression of the bulk tumor tissue. Thus, to be able to evaluate results from cDNA array gene expression experiments it is, to our opinion, necessary to work with pure tumor cell populations, until solid information is available on the impact of stromal component. Housekeeping genes should be handling with care and mg tissue may be preferred instead of microg RNA for standardization.

  17. CYP1A2 and CYP2D6 Gene Polymorphisms in Schizophrenic Patients with Neuroleptic Drug-Induced Side Effects.

    PubMed

    Ivanova, S A; Filipenko, M L; Vyalova, N M; Voronina, E N; Pozhidaev, I V; Osmanova, D Z; Ivanov, M V; Fedorenko, O Yu; Semke, A V; Bokhan, N A

    2016-03-01

    Polymorphic variants of CYP1A2 and CYP2D6 genes of the cytochrome P450 system were studied in patients with schizophrenia with drug-induced motor disorders and hyperprolactinemia against the background of long-term neuroleptic therapy. We revealed an association of polymorphic variant C-163A CYP1A2*1F of CYP1A2 gene with tardive dyskinesia and association of polymorphic variant 1846G>A CY2D6*4 and genotype A/A of CYP2D6 gene (responsible for debrisoquin-4-hydroxylase synthesis) with limbotruncal tardive dyskinesia in patients with schizophrenia receiving neuroleptics for a long time.

  18. The acute impact of polyphenols from Hibiscus sabdariffa in metabolic homeostasis: an approach combining metabolomics and gene-expression analyses.

    PubMed

    Beltrán-Debón, Raúl; Rodríguez-Gallego, Esther; Fernández-Arroyo, Salvador; Senan-Campos, Oriol; Massucci, Francesco A; Hernández-Aguilera, Anna; Sales-Pardo, Marta; Guimerà, Roger; Camps, Jordi; Menendez, Javier A; Joven, Jorge

    2015-09-01

    We explored the acute multifunctional effects of polyphenols from Hibiscus sabdariffa in humans to assess possible consequences on the host's health. The expected dynamic response was studied using a combination of transcriptomics and metabolomics to integrate specific functional pathways through network-based methods and to generate hypotheses established by acute metabolic effects and/or modifications in the expression of relevant genes. Data were obtained from healthy male volunteers after 3 hours of ingestion of an aqueous Hibiscus sabdariffa extract. The data were compared with data obtained prior to the ingestion, and the overall findings suggest that these particular polyphenols had a simultaneous role in mitochondrial function, energy homeostasis and protection of the cardiovascular system. These findings suggest beneficial actions in inflammation, endothelial dysfunction, and oxidation, which are interrelated mechanisms. Among other effects, the activation of the heme oxygenase-biliverdin reductase axis, the systemic inhibition of the renin-angiotensin system, the inhibition of the angiotensin-converting enzyme, and several actions mirroring those of the peroxisome proliferator-activated receptor agonists further support this notion. We also found concordant findings in the serum of the participants, which include a decrease in cortisol levels and a significant increase in the active vasodilator metabolite of bradykinin (des-Arg(9)-bradykinin). Therefore, our data support the view that polyphenols from Hibiscus sabdariffa play a regulatory role in metabolic health and in the maintenance of blood pressure, thus implying a multi-faceted impact in metabolic and cardiovascular diseases.

  19. Epidermal growth factor receptor and AKT1 gene copy numbers by multi-gene fluorescence in situ hybridization impact on prognosis in breast cancer.

    PubMed

    Li, Jiao; Su, Wei; Zhang, Sheng; Hu, Yunhui; Liu, Jingjing; Zhang, Xiaobei; Bai, Jingchao; Yuan, Weiping; Hu, Linping; Cheng, Tao; Zetterberg, Anders; Lei, Zhenmin; Zhang, Jin

    2015-05-01

    The epidermal growth factor receptor (EGFR)/PI3K/AKT signaling pathway aberrations play significant roles in breast cancer occurrence and development. However, the status of EGFR and AKT1 gene copy numbers remains unclear. In this study, we showed that the rates of EGFR and AKT1 gene copy number alterations were associated with the prognosis of breast cancer. Among 205 patients, high EGFR and AKT1 gene copy numbers were observed in 34.6% and 27.8% of cases by multi-gene fluorescence in situ hybridization, respectively. Co-heightened EGFR/AKT1 gene copy numbers were identified in 11.7% cases. No changes were found in 49.3% of patients. Although changes in EGFR and AKT1 gene copy numbers had no correlation with patients' age, tumor stage, histological grade and the expression status of other molecular makers, high EGFR (P = 0.0002) but not AKT1 (P = 0.1177) gene copy numbers correlated with poor 5-year overall survival. The patients with co-heightened EGFR/AKT1 gene copy numbers displayed a poorer prognosis than those with tumors with only high EGFR gene copy numbers (P = 0.0383). Both Univariate (U) and COX multivariate (C) analyses revealed that high EGFR and AKT1 gene copy numbers (P = 0.000 [U], P = 0.0001 [C]), similar to histological grade (P = 0.001 [U], P = 0.012 [C]) and lymph node metastasis (P = 0.046 [U], P = 0.158 [C]), were independent prognostic indicators of 5-year overall survival. These results indicate that high EGFR and AKT1 gene copy numbers were relatively frequent in breast cancer. Co-heightened EGFR/AKT1 gene copy numbers had a worse outcome than those with only high EGFR gene copy numbers, suggesting that evaluation of these two genes together may be useful for selecting patients for anti-EGFR-targeted therapy or anti-EGFR/AKT1-targeted therapy and for predicting outcomes. © 2015 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

  20. Impacts of gene bioaugmentation with pJP4-harboring bacteria of 2,4-D-contaminated soil slurry on the indigenous microbial community.

    PubMed

    Inoue, Daisuke; Yamazaki, Yuji; Tsutsui, Hirofumi; Sei, Kazunari; Soda, Satoshi; Fujita, Masanori; Ike, Michihiko

    2012-04-01

    Gene bioaugmentation is a bioremediation strategy that enhances biodegradative potential via dissemination of degradative genes from introduced microorganisms to indigenous microorganisms. Bioremediation experiments using 2,4-dichlorophenoxyacetic acid (2,4-D)-contaminated soil slurry and strains of Pseudomonas putida or Escherichia coli harboring a self-transmissible 2,4-D degradative plasmid pJP4 were conducted in microcosms to assess possible effects of gene bioaugmentation on the overall microbial community structure and ecological functions (carbon source utilization and nitrogen transformation potentials). Although exogenous bacteria decreased rapidly, 2,4-D degradation was stimulated in bioaugmented microcosms, possibly because of the occurrence of transconjugants by the transfer of pJP4. Terminal restriction fragment length polymorphism analysis revealed that, although the bacterial community structure was disturbed immediately after introducing exogenous bacteria to the inoculated microcosms, it gradually approached that of the uninoculated microcosms. Biolog assay, nitrate reduction assay, and monitoring of the amoA gene of ammonia-oxidizing bacteria and nirK and nirS genes of denitrifying bacteria showed no irretrievable depressive effects of gene bioaugmentation on the carbon source utilization and nitrogen transformation potentials. These results may suggest that gene bioaugmentation with P. putida and E. coli strains harboring pJP4 is effective for the degradation of 2,4-D in soil without large impacts on the indigenous microbial community.