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Sample records for gene expression cellular

  1. Molecular crowding shapes gene expression in synthetic cellular nanosystems.

    PubMed

    Tan, Cheemeng; Saurabh, Saumya; Bruchez, Marcel P; Schwartz, Russell; Leduc, Philip

    2013-08-01

    The integration of synthetic and cell-free biology has made tremendous strides towards creating artificial cellular nanosystems using concepts from solution-based chemistry, where only the concentrations of reacting species modulate gene expression rates. However, it is known that macromolecular crowding, a key feature in natural cells, can dramatically influence biochemical kinetics via volume exclusion effects, which reduce diffusion rates and enhance binding rates of macromolecules. Here, we demonstrate that macromolecular crowding can increase the robustness of gene expression by integrating synthetic cellular components of biological circuits and artificial cellular nanosystems. Furthermore, we reveal how ubiquitous cellular modules, including genetic components, a negative feedback loop and the size of the crowding molecules can fine-tune gene circuit response to molecular crowding. By bridging a key gap between artificial and living cells, our work has implications for efficient and robust control of both synthetic and natural cellular circuits.

  2. Molecular crowding shapes gene expression in synthetic cellular nanosystems

    PubMed Central

    Tan, Cheemeng; Saurabh, Saumya; Bruchez, Marcel; Schwartz, Russell; LeDuc, Philip

    2013-01-01

    Summary The integration of synthetic and cell-free biology has made tremendous strides towards creating artificial cellular nanosystems using concepts from solution-based chemistry: only the concentrations of reacting species modulate gene expression rates. However, it is known that macromolecular crowding, a key feature of natural cells, can dramatically influence biochemical kinetics by volume exclusion effects that reduce diffusion rates and enhance binding rates of macromolecules. Here, we demonstrate that macromolecular crowding can increase the robustness of gene expression through integrating synthetic cellular components of biological circuits and artificial cellular nanosystems. In addition, we reveal how ubiquitous cellular modules, including genetic components, a negative feedback loop, and the size of crowding molecules, can fine tune gene circuit response to molecular crowding. By bridging a key gap between artificial and living cells, our work has implications for efficient and robust control of both synthetic and natural cellular circuits. PMID:23851358

  3. Stably Expressed Genes Involved in Basic Cellular Functions

    PubMed Central

    Wang, Kejian; Fuscoe, James C.

    2017-01-01

    Stably Expressed Genes (SEGs) whose expression varies within a narrow range may be involved in core cellular processes necessary for basic functions. To identify such genes, we re-analyzed existing RNA-Seq gene expression profiles across 11 organs at 4 developmental stages (from immature to old age) in both sexes of F344 rats (n = 4/group; 320 samples). Expression changes (calculated as the maximum expression / minimum expression for each gene) of >19000 genes across organs, ages, and sexes ranged from 2.35 to >109-fold, with a median of 165-fold. The expression of 278 SEGs was found to vary ≤4-fold and these genes were significantly involved in protein catabolism (proteasome and ubiquitination), RNA transport, protein processing, and the spliceosome. Such stability of expression was further validated in human samples where the expression variability of the homologous human SEGs was significantly lower than that of other genes in the human genome. It was also found that the homologous human SEGs were generally less subject to non-synonymous mutation than other genes, as would be expected of stably expressed genes. We also found that knockout of SEG homologs in mouse models was more likely to cause complete preweaning lethality than non-SEG homologs, corroborating the fundamental roles played by SEGs in biological development. Such stably expressed genes and pathways across life-stages suggest that tight control of these processes is important in basic cellular functions and that perturbation by endogenous (e.g., genetics) or exogenous agents (e.g., drugs, environmental factors) may cause serious adverse effects. PMID:28125669

  4. Genetic determinants and cellular constraints in noisy gene expression

    PubMed Central

    Sanchez, Alvaro; Golding, Ido

    2014-01-01

    In individual cells, transcription is a random process obeying single-molecule kinetics. Often, it occurs in a bursty, intermittent manner. The frequency and size of these bursts affect the magnitude of temporal fluctuations in mRNA and protein content within a cell, creating variation or “noise” in gene expression. It is still unclear to what degree transcriptional kinetics are specific to each gene and determined by its promoter sequence. Alternative scenarios have been proposed, where the kinetics of transcription are governed by cellular constraints and follow universal rules across the genome. Evidence from genome-wide noise studies and from systematic perturbations of promoter sequences suggest that both scenarios—namely gene-specific versus genome-wide regulation of transcription kinetics— may be present to different degrees in bacteria, yeast and animal cells. PMID:24311680

  5. The Effect of Gravity Fields on Cellular Gene Expression

    NASA Technical Reports Server (NTRS)

    Hughes-Fulford, Millie

    1999-01-01

    Early theoretical analysis predicted that microgravity effects on the isolated cell would be minuscule at the subcellular level; however, these speculations have not proven true in the real world. Astronauts experience a significant bone and muscle loss in as little as 2 weeks of spaceflight and changes are seen at the cellular level soon after exposure to microgravity. Changes in biological systems may be primarily due to the lack of gravity and the resulting loss of mechanical stress on tissues and cells. Recent ground and flight studies examining the effects of gravity or mechanical stress on cells demonstrate marked changes in gene expression when relatively small changes in mechanical forces or gravity fields were made. Several immediate early genes (IEG) like c-fos and c-myc are induced by mechanical stimulation within minutes. In contrast, several investigators report that the absence of mechanical forces during space flight result in decreased sera response element (SRE) activity and attenuation of expression of IEGs such as c-fos, c-jun and cox-2 mRNAs. Clearly, these early changes in gene expression may have long term consequences on mechanically sensitive cells. In our early studies on STS-56, we reported four major changes in the osteoblast; 1) prostaglandin synthesis in flight, 2) changes in cellular morphology, 3) altered actin cytoskeleton and 4) reduced osteoblast growth after four days exposure to microgravity. Initially, it was believed that changes in fibronectin (FN) RNA, FN protein synthesis or subsequent FN matrix formation might account for the changes in cytoskeleton and/ or reduction of growth. However our recent studies on Biorack (STS-76, STS-81 and STS-84), using ground and in-flight 1-G controls, demonstrated that fibronectin synthesis and matrix formation were normal in microgravity. In addition, in our most recent Biorack paper, our laboratory has documented that relative protein synthesis and mRNA synthesis are not changed after 24

  6. 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.

  7. Gene Expression Profiling Reveals Early Cellular Responses to Intracellular Magnetic Labeling with Superparamagnetic Iron Oxide Nanoparticles

    PubMed Central

    Kedziorek, Dorota A.; Muja, Naser; Walczak, Piotr; Ruiz-Cabello, Jesus; Gilad, Assaf A.; Jie, Chunfa C.; Bulte, Jeff W. M.

    2010-01-01

    With MRI (stem) cell tracking having entered the clinic, studies on the cellular genomic response toward labeling are warranted. Gene expression profiling was applied to C17.2 neural stem cells following superparamagnetic iron oxide/PLL (poly-L-lysine) labeling over the course of 1 week. Relative to unlabeled cells, less than 1% of genes (49 total) exhibited greater than 2-fold difference in expression in response to superparamagnetic iron oxide/PLL labeling. In particular, transferrin receptor 1 (Tfrc) and heme oxygenase 1 (Hmox1) expression was downregulated early, whereas genes involved in lysosomal function (Sulf1) and detoxification (Clu, Cp, Gstm2, Mgst1) were upregulated at later time points. Relative to cells treated with PLL only, cells labeled with superparamagnetic iron oxide/PLL complexes exhibited differential expression of 1399 genes. Though these differentially expressed genes exhibited altered expression over time, the overall extent was limited. Gene ontology analysis of differentially expressed genes showed that genes encoding zinc-binding proteins are enriched after superparamagnetic iron oxide/PLL labeling relative to PLL only treatment, whereas members of the apoptosis/ programmed cell death pathway did not display increased expression. Overexpression of the differentially expressed genes Rnf138 and Abcc4 were confirmed by quantitative real-time polymerase chain reaction. These results demonstrate that, although early reactions responsible for iron homeostasis are induced, overall neural stem cell gene expression remains largely unaltered following superparamagnetic iron oxide/PLL labeling. PMID:20373404

  8. Introns and gene expression: Cellular constraints, transcriptional regulation, and evolutionary consequences

    PubMed Central

    Heyn, Patricia; Kalinka, Alex T; Tomancak, Pavel; Neugebauer, Karla M

    2015-01-01

    A gene's “expression profile” denotes the number of transcripts present relative to all other transcripts. The overall rate of transcript production is determined by transcription and RNA processing rates. While the speed of elongating RNA polymerase II has been characterized for many different genes and organisms, gene-architectural features – primarily the number and length of exons and introns – have recently emerged as important regulatory players. Several new studies indicate that rapidly cycling cells constrain gene-architecture toward short genes with a few introns, allowing efficient expression during short cell cycles. In contrast, longer genes with long introns exhibit delayed expression, which can serve as timing mechanisms for patterning processes. These findings indicate that cell cycle constraints drive the evolution of gene-architecture and shape the transcriptome of a given cell type. Furthermore, a tendency for short genes to be evolutionarily young hints at links between cellular constraints and the evolution of animal ontogeny. PMID:25400101

  9. A single-cell bioluminescence imaging system for monitoring cellular gene expression in a plant body.

    PubMed

    Muranaka, Tomoaki; Kubota, Saya; Oyama, Tokitaka

    2013-12-01

    Gene expression is a fundamental cellular process and expression dynamics are of great interest in life science. We succeeded in monitoring cellular gene expression in a duckweed plant, Lemna gibba, using bioluminescent reporters. Using particle bombardment, epidermal and mesophyll cells were transfected with the luciferase gene (luc+) under the control of a constitutive [Cauliflower mosaic virus 35S (CaMV35S)] and a rhythmic [Arabidopsis thaliana CIRCADIAN CLOCK ASSOCIATED 1 (AtCCA1)] promoter. Bioluminescence images were captured using an EM-CCD (electron multiply charged couple device) camera. Luminescent spots of the transfected cells in the plant body were quantitatively measured at the single-cell level. Luminescence intensities varied over a 1,000-fold range among CaMV35S::luc+-transfected cells in the same plant body and showed a log-normal-like frequency distribution. We monitored cellular gene expression under light-dark conditions by capturing bioluminescence images every hour. Luminescence traces of ≥50 individual cells in a frond were successfully obtained in each monitoring procedure. Rhythmic and constitutive luminescence behaviors were observed in cells transfected with AtCCA1::luc+ and CaMV35S::luc+, respectively. Diurnal rhythms were observed in every AtCCA1::luc+-introduced cell with traceable luminescence, and slight differences were detected in their rhythmic waveforms. Thus the single-cell bioluminescence monitoring system was useful for the characterization of cellular gene expression in a plant body.

  10. Cellular expansion and gene expression in the developing grape (Vitis vinifera L.).

    PubMed

    Schlosser, J; Olsson, N; Weis, M; Reid, K; Peng, F; Lund, S; Bowen, P

    2008-01-01

    Expression profiles of genes involved in cell wall metabolism and water transport were compared with changes in grape (Vitis vinifera L.) berry growth, basic chemical composition, and the shape, size, and wall thickness of cells within tissues of the berry pericarp. Expression of cell wall-modifying and aquaporin genes in berry pericarp tissues generally followed a bimodal expression profile with high levels of expression coinciding with the two periods of rapid berry growth, stages I and III, and low levels of expression corresponding to the slow-growth period, stage II. Cellular expansion was observed throughout all tissues during stage I, and only mesocarp cellular expansion was observed during stage III. Expansion of only exocarp cells was evident during transition between stages II and III. Cell wall-modifying and aquaporin gene expression profiles followed similar trends in exocarp and mesocarp tissues throughout berry development, with the exception of the up-regulation of pectin methylesterase, pectate lyase, two aquaporin genes (AQ1 and AQ2), and two expansin genes (EXP3 and EXPL) during stage II, which was delayed in the exocarp tissue compared with mesocarp tissue. Exocarp endo-(1-->3)-beta-glucanase and expansin-like gene expression was concurrent with increases in epidermal and hypodermal cell wall thickness. These results indicate a potential role of the grape berry skin in modulating grape berry growth.

  11. Tat is a multifunctional viral protein that modulates cellular gene expression and functions.

    PubMed

    Clark, Evan; Nava, Brenda; Caputi, Massimo

    2017-02-07

    The human immunodeficiency virus type I (HIV-1) has developed several strategies to condition the host environment to promote viral replication and spread. Viral proteins have evolved to perform multiple functions, aiding in the replication of the viral genome and modulating the cellular response to the infection. Tat is a small, versatile, viral protein that controls transcription of the HIV genome, regulates cellular gene expression and generates a permissive environment for viral replication by altering the immune response and facilitating viral spread to multiple tissues. Studies carried out utilizing biochemical, cellular, and genomic approaches show that the expression and activity of hundreds of genes and multiple molecular networks are modulated by Tat via multiple mechanisms.

  12. Posttranscriptional regulation of cellular gene expression by the c-myc oncogene

    SciTech Connect

    Prendergast, G.C.; Cole, M.D. . Dept. of Biology)

    1989-01-01

    The c-myc oncogene has been implicated in the development of many different cancers, yet the mechanism by which the c-myc protein alters cellular growth control has proven elusive. The authors used a cDNA hybridization difference assay to isolate two genes, mr1 and mr2, that were constitutively expressed (i.e., deregulated) in rodent fibroblast cell lines immortalized by transfection of a viral promoter-linked c-myc gene. Both cDNAs were serum inducible in quiescent G/sub o/ fibroblasts, suggesting that they are functionally related to cellular proliferative processes. Although there were significant differences in cytoplasmic mRNA levels between myc-immortalized and control cells, the rates of transcription and mRNA turnover of both genes were similar, suggesting that c-myc regulates mr1 and mr2 expression by some nuclear posttranscriptional mechanism. Their results provide evidence that c-myc can rapidly modulate cellular gene expression and suggest that c-myc may function in gene regulation at the level of RNA export, splicing, or nuclear RNA turnover.

  13. Building quantitative, three dimensional atlases of gene expression and morphology at cellular resolution

    PubMed Central

    Knowles, David W.; Biggin, Mark D.

    2013-01-01

    Animals comprise dynamic three-dimensional arrays of cells that express gene products in intricate spatial and temporal patterns that determine cellular differentiation and morphogenesis. A rigorous understanding of these developmental processes requires automated methods that quantitatively record and analyze complex morphologies and their associated patterns of gene expression at cellular resolution. Here we summarize light microscopy based approaches to establish permanent, quantitative datasets—atlases—that record this information. We focus on experiments that capture data for whole embryos or large areas of tissue in three dimensions, often at multiple time points. We compare and contrast the advantages and limitations of different methods and highlight some of the discoveries made. We emphasize the need for interdisciplinary collaborations and integrated experimental pipelines that link sample preparation, image acquisition, image analysis, database design, visualization and quantitative analysis. PMID:24123936

  14. Zoledronic acid and geranylgeraniol regulate cellular behaviour and angiogenic gene expression in human gingival fibroblasts.

    PubMed

    Zafar, S; Coates, D E; Cullinan, M P; Drummond, B K; Milne, T; Seymour, G J

    2014-10-01

    The mevalonate pathway (MVP) and the anti-angiogenic effect of bisphosphonates have been shown to play a role in the pathogenesis of bisphosphonate-related osteonecrosis of the jaw (BRONJ). This study determined the effect of the bisphosphonate, zoledronic acid and the replenishment of the MVP by geranylgeraniol on human gingival fibroblasts. Cell viability, apoptosis, morphological analysis using transmission electron microscopy, and gene expression for vascular endothelial growth factor A, bone morphogenic protein 2, ras homologue gene family member B, epiregulin and interferon-alpha were conducted. Results showed cellular viability was decreased in the presence of zoledronic acid and the co-addition of zoledronic acid with geranylgeraniol restored cell viability to control levels. Caspase 3/7 was detected in zoledronic-acid-treated cells indicating apoptosis. Transmission electron microscopy revealed dilation of the rough endoplasmic reticulum with zoledronic acid and the appearance of multiple lipid-like vesicles following the addition of geranylgeraniol. Zoledronic acid significantly (P < 0.05, FR > ± 2) up-regulated vascular endothelial growth factor A, bone morphogenic protein 2, ras homologue gene family member B and epiregulin at one or more time points but not interferon-alpha. Addition of geranylgeraniol resulted in a reduction in the expression of all five genes compared with zoledronic-acid-treated human gingival fibroblasts. The study concluded geranylgeraniol partially reversed the effects of zoledronic acid in human gingival fibroblasts both at the cellular and genetic levels, suggesting the regulation of these genes is mediated via the mevalonate pathway.

  15. Cellular Stress Response Gene Expression During Upper and Lower Body High Intensity Exercises

    PubMed Central

    Kochanowicz, Andrzej; Sawczyn, Stanisław; Niespodziński, Bartłomiej; Mieszkowski, Jan; Kochanowicz, Kazimierz

    2017-01-01

    Objectives The aim was to compare the effect of upper and lower body high-intensity exercise on chosen genes expression in athletes and non-athletes. Method Fourteen elite male artistic gymnasts (EAG) aged 20.6 ± 3.3 years and 14 physically active men (PAM) aged 19.9 ± 1.0 years performed lower and upper body 30 s Wingate Tests. Blood samples were collected before, 5 and 30 minutes after each effort to assess gene expression via PCR. Results Significantly higher mechanical parameters after lower body exercise was observed in both groups, for relative power (8.7 ± 1.2 W/kg in gymnasts, 7.2 ± 1.2 W/kg in controls, p = 0.01) and mean power (6.7 ± 0.7 W/kg in gymnasts, 5.4 ± 0.8 W/kg in controls, p = 0.01). No differences in lower versus upper body gene expression were detected for all tested genes as well as between gymnasts and physical active man. For IL-6 m-RNA time-dependent effect was observed. Conclusions Because of no significant differences in expression of genes associated with cellular stress response the similar adaptive effect to exercise may be obtained so by lower and upper body exercise. PMID:28141870

  16. Quality Controls in Cellular Immunotherapies: Rapid Assessment of Clinical Grade Dendritic Cells by Gene Expression Profiling

    PubMed Central

    Castiello, Luciano; Sabatino, Marianna; Zhao, Yingdong; Tumaini, Barbara; Ren, Jiaqiang; Ping, Jin; Wang, Ena; Wood, Lauren V; Marincola, Francesco M; Puri, Raj K; Stroncek, David F

    2013-01-01

    Cell-based immunotherapies are among the most promising approaches for developing effective and targeted immune response. However, their clinical usefulness and the evaluation of their efficacy rely heavily on complex quality control assessment. Therefore, rapid systematic methods are urgently needed for the in-depth characterization of relevant factors affecting newly developed cell product consistency and the identification of reliable markers for quality control. Using dendritic cells (DCs) as a model, we present a strategy to comprehensively characterize manufactured cellular products in order to define factors affecting their variability, quality and function. After generating clinical grade human monocyte-derived mature DCs (mDCs), we tested by gene expression profiling the degrees of product consistency related to the manufacturing process and variability due to intra- and interdonor factors, and how each factor affects single gene variation. Then, by calculating for each gene an index of variation we selected candidate markers for identity testing, and defined a set of genes that may be useful comparability and potency markers. Subsequently, we confirmed the observed gene index of variation in a larger clinical data set. In conclusion, using high-throughput technology we developed a method for the characterization of cellular therapies and the discovery of novel candidate quality assurance markers. PMID:23147403

  17. Vitamin D Metabolites Inhibit Hepatitis C Virus and Modulate Cellular Gene Expression

    PubMed Central

    Gutierrez, Julio A.; Jones, Krysten A.; Flores, Roxana; Singhania, Akul; Woelk, Christopher H.; Schooley, Robert T.; Wyles, David L.

    2015-01-01

    Background and Aims Previous studies suggest that low serum 25-hydroxyvitamin D [25(OH) D] levels are associated with reduced responsiveness to interferon and ribavirin therapy. We investigated the impact of vitamin D metabolites on HCV and cellular gene expression in cultured hepatoma cells. Methods HCV Replicon cell lines stably expressing luciferase reporter constructs (genotype 1b and 2a replicon) or JC1-Luc2a were incubated in the presence of vitamin D2, vitamin D3 or 1,25-dihydroxyvitamin D3 (1,25(OH)2D3). Presence of HCV was quantified by a luciferase reporter assay and immunoblot of the Core protein. Synergy of interferon-alpha A/D (IFN-α) and 1,25(OH)2D3 was evaluated using the Chou-Talalay method. Cellular gene expression by microarray analysis using Illumina Bead Chips and real-time quantitative PCR. Results Vitamin D2, D3 and 1,25(OH)2D3 each demonstrated anti-HCV activity at low micro molar concentrations. In vitro conversion from D3 to 25(OH)D3 was shown by LC/MS/MS. Combination indices of 1,25(OH)2D3 and IFN-α demonstrated a synergistic effect (0.23-0.46) and significantly reduced core expression by immunoblot. Differentially expressed genes were identified between Huh7.5.1 cells in the presence and absence of 1,25(OH)2D3 and HCV. Genes involved with classical effects of vitamin D metabolism and excretion were activated, along with genes linked to autophagy such as G-protein coupled receptor 37 (GPR37) and Hypoxia-inducible factor 1-alpha (HIF1a). Additionally, additive effects of 1,25(OH)2D3 and IFN-α were seen on mRNA expression of chemokine motif ligand 20 (CCL20). Conclusions This study shows that vitamin D reduces HCV protein production in cell culture synergistically with IFN-α. Vitamin D also activates gene expression independently and additively with IFN-α and this may explain its ability to aid in the clearance of HCV in vivo. PMID:26594646

  18. Disruption of a cystine transporter downregulates expression of genes involved in sulfur regulation and cellular respiration

    PubMed Central

    Simpkins, Jessica A.; Rickel, Kirby E.; Madeo, Marianna; Ahlers, Bethany A.; Carlisle, Gabriel B.; Nelson, Heidi J.; Cardillo, Andrew L.; Weber, Emily A.; Vitiello, Peter F.; Pearce, David A.

    2016-01-01

    ABSTRACT Cystine and cysteine are important molecules for pathways such as redox signaling and regulation, and thus identifying cellular deficits upon deletion of the Saccharomyces cerevisiae cystine transporter Ers1p allows for a further understanding of cystine homeostasis. Previous complementation studies using the human ortholog suggest yeast Ers1p is a cystine transporter. Human CTNS encodes the protein Cystinosin, a cystine transporter that is embedded in the lysosomal membrane and facilitates the export of cystine from the lysosome. When CTNS is mutated, cystine transport is disrupted, leading to cystine accumulation, the diagnostic hallmark of the lysosomal storage disorder cystinosis. Here, we provide biochemical evidence for Ers1p-dependent cystine transport. However, the accumulation of intracellular cystine is not observed when the ERS1 gene is deleted from ers1-Δ yeast, supporting the existence of modifier genes that provide a mechanism in ers1-Δ yeast that prevents or corrects cystine accumulation. Upon comparison of the transcriptomes of isogenic ERS1+ and ers1-Δ strains of S. cerevisiae by DNA microarray followed by targeted qPCR, sixteen genes were identified as being differentially expressed between the two genotypes. Genes that encode proteins functioning in sulfur regulation, cellular respiration, and general transport were enriched in our screen, demonstrating pleiotropic effects of ers1-Δ. These results give insight into yeast cystine regulation and the multiple, seemingly distal, pathways that involve proper cystine recycling. PMID:27142334

  19. JC virus induces altered patterns of cellular gene expression: Interferon-inducible genes as major transcriptional targets

    SciTech Connect

    Verma, Saguna; Ziegler, Katja; Ananthula, Praveen; Co, Juliene K.G.; Frisque, Richard J.; Yanagihara, Richard; Nerurkar, Vivek R. . E-mail: nerurkar@pbrc.hawaii.edu

    2006-02-20

    Human polyomavirus JC (JCV) infects 80% of the population worldwide. Primary infection, typically occurring during childhood, is asymptomatic in immunocompetent individuals and results in lifelong latency and persistent infection. However, among the severely immunocompromised, JCV may cause a fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). Virus-host interactions influencing persistence and pathogenicity are not well understood, although significant regulation of JCV activity is thought to occur at the level of transcription. Regulation of the JCV early and late promoters during the lytic cycle is a complex event that requires participation of both viral and cellular factors. We have used cDNA microarray technology to analyze global alterations in gene expression in JCV-permissive primary human fetal glial cells (PHFG). Expression of more than 400 cellular genes was altered, including many that influence cell proliferation, cell communication and interferon (IFN)-mediated host defense responses. Genes in the latter category included signal transducer and activator of transcription 1 (STAT1), interferon stimulating gene 56 (ISG56), myxovirus resistance 1 (MxA), 2'5'-oligoadenylate synthetase (OAS), and cig5. The expression of these genes was further confirmed in JCV-infected PHFG cells and the human glioblastoma cell line U87MG to ensure the specificity of JCV in inducing this strong antiviral response. Results obtained by real-time RT-PCR and Western blot analyses supported the microarray data and provide temporal information related to virus-induced changes in the IFN response pathway. Our data indicate that the induction of an antiviral response may be one of the cellular factors regulating/controlling JCV replication in immunocompetent hosts and therefore constraining the development of PML.

  20. BRCA1 haploinsufficiency leads to altered expression of genes involved in cellular proliferation and development.

    PubMed

    Feilotter, Harriet E; Michel, Claire; Uy, Paolo; Bathurst, Lauren; Davey, Scott

    2014-01-01

    The assessment of BRCA1 and BRCA2 coding sequences to identify pathogenic mutations associated with inherited breast/ovarian cancer syndrome has provided a method to identify high-risk individuals, allowing them to seek preventative treatments and strategies. However, the current test is expensive, and cannot differentiate between pathogenic variants and those that may be benign. Focusing only on one of the two BRCA partners, we have developed a biological assay for haploinsufficiency of BRCA1. Using a series of EBV-transformed cell lines, we explored gene expression patterns in cells that were BRCA1 wildtype compared to those that carried (heterozygous) BRCA1 pathogenic mutations. We identified a subset of 43 genes whose combined expression pattern is a sensitive predictor of BRCA1 status. The gene set was disproportionately made up of genes involved in cellular differentiation, lending credence to the hypothesis that single copy loss of BRCA1 function may impact differentiation, rendering cells more susceptible to undergoing malignant processes.

  1. Cellular Defense System Gene Expression Profiling of Human Whole Blood: Opportunities to Predict Health Benefits in Response to Diet12

    PubMed Central

    Drew, Janice E.

    2012-01-01

    Diet is a critical factor in the maintenance of human cellular defense systems, immunity, inflammation, redox regulation, metabolism, and DNA repair that ensure optimal health and reduce disease risk. Assessment of dietary modulation of cellular defense systems in humans has been limited due to difficulties in accessing target tissues. Notably, peripheral blood gene expression profiles associated with nonhematologic disease are detectable. Coupled with recent innovations in gene expression technologies, gene expression profiling of human blood to determine predictive markers associated with health status and dietary modulation is now a feasible prospect for nutrition scientists. This review focuses on cellular defense system gene expression profiling of human whole blood and the opportunities this presents, using recent technological advances, to predict health status and benefits conferred by diet. PMID:22797985

  2. Adult Drosophila melanogaster evolved for antibacterial defense invest in infection-induced expression of both humoral and cellular immunity genes

    PubMed Central

    2011-01-01

    Background While the transcription of innate immunity genes in response to bacterial infection has been well-characterised in the Drosophila model, we recently demonstrated the capacity for such transcription to evolve in flies selected for improved antibacterial defense. Here we use this experimental system to examine how insects invest in constitutive versus infection-induced transcription of immunity genes. These two strategies carry with them different consequences with respect to energetic and pleiotropic costs and may be more or less effective in improving defense depending on whether the genes contribute to humoral or cellular aspects of immunity. Findings Contrary to expectation we show that selection preferentially increased the infection-induced expression of both cellular and humoral immunity genes. Given their functional roles, infection induced increases in expression were expected for the humoral genes, while increases in constitutive expression were expected for the cellular genes. We also report a restricted ability to improve transcription of immunity genes that is on the order of 2-3 fold regardless of total transcription level of the gene. Conclusions The evolved increases in infection-induced expression of the cellular genes may result from specific cross talk with humoral pathways or from generalised strategies for enhancing immunity gene transcription. A failure to see improvements in constitutive expression of the cellular genes suggests either that increases might come at too great a cost or that patterns of expression in adults are decoupled from the larval phase where increases would be most effective. The similarity in fold change increase across all immunity genes may suggest a shared mechanism for the evolution of increased transcription in small, discrete units such as duplication of cis-regulatory elements. PMID:21859495

  3. Regulation of metallothionein gene expression and cellular zinc accumulation in a rat small intestinal cell line

    SciTech Connect

    Carlson, J.M.; Cousins, R.J. )

    1991-03-15

    The effects of extracellular zinc concentration on metallothionein gene expression and cellular zinc accumulation were studied in IRD-98 cells. This is a non-transformed clonal line established by Negrel, et al. from fetal rat small intestine which possess characteristics of small bowel epithelial cells. Cells were maintained in DMEM and grown to confluent monolayers. The response to media zinc concentrations over the range of 5-150 {mu}mol/L was assessed. After 24 h in culture, cell zinc and metallothionein protein concentrations were significantly increased in cells provided higher levels of media zinc. Subsequent time course experiments showed that cells exposed to higher zinc levels had significant elevations in both metallothionein mRNA, peaking at 24 h, and metallothionein protein increasing through 48 h. Furthermore, cell zinc concentrations were significantly increased. At 48 h of culture, greater than 50% of the additional cellular zinc accumulated could be attributed to elevated metallothionein protein levels. These cells represent a zinc-responsive model to examine the mechanism of zinc uptake and transcellular transport by intestinal cells and the regulatory factors involved.

  4. Three-dimensional morphology and gene expression in the Drosophila blastoderm at cellular resolution I: data acquisition pipeline

    PubMed Central

    Luengo Hendriks, Cris L; Keränen, Soile VE; Fowlkes, Charless C; Simirenko, Lisa; Weber, Gunther H; DePace, Angela H; Henriquez, Clara; Kaszuba, David W; Hamann, Bernd; Eisen, Michael B; Malik, Jitendra; Sudar, Damir; Biggin, Mark D; Knowles, David W

    2006-01-01

    Background To model and thoroughly understand animal transcription networks, it is essential to derive accurate spatial and temporal descriptions of developing gene expression patterns with cellular resolution. Results Here we describe a suite of methods that provide the first quantitative three-dimensional description of gene expression and morphology at cellular resolution in whole embryos. A database containing information derived from 1,282 embryos is released that describes the mRNA expression of 22 genes at multiple time points in the Drosophila blastoderm. We demonstrate that our methods are sufficiently accurate to detect previously undescribed features of morphology and gene expression. The cellular blastoderm is shown to have an intricate morphology of nuclear density patterns and apical/basal displacements that correlate with later well-known morphological features. Pair rule gene expression stripes, generally considered to specify patterning only along the anterior/posterior body axis, are shown to have complex changes in stripe location, stripe curvature, and expression level along the dorsal/ventral axis. Pair rule genes are also found to not always maintain the same register to each other. Conclusion The application of these quantitative methods to other developmental systems will likely reveal many other previously unknown features and provide a more rigorous understanding of developmental regulatory networks. PMID:17184546

  5. Myocardial Gene Transfer: Routes and Devices for Regulation of Transgene Expression by Modulation of Cellular Permeability

    PubMed Central

    Katz, Michael G.; Bridges, Charles R.

    2013-01-01

    Abstract Heart diseases are major causes of morbidity and mortality in Western society. Gene therapy approaches are becoming promising therapeutic modalities to improve underlying molecular processes affecting failing cardiomyocytes. Numerous cardiac clinical gene therapy trials have yet to demonstrate strong positive results and advantages over current pharmacotherapy. The success of gene therapy depends largely on the creation of a reliable and efficient delivery method. The establishment of such a system is determined by its ability to overcome the existing biological barriers, including cellular uptake and intracellular trafficking as well as modulation of cellular permeability. In this article, we describe a variety of physical and mechanical methods, based on the transient disruption of the cell membrane, which are applied in nonviral gene transfer. In addition, we focus on the use of different physiological techniques and devices and pharmacological agents to enhance endothelial permeability. Development of these methods will undoubtedly help solve major problems facing gene therapy. PMID:23427834

  6. Epstein–Barr virus transcription factor Zta acts through distal regulatory elements to directly control cellular gene expression

    PubMed Central

    Ramasubramanyan, Sharada; Osborn, Kay; Al-Mohammad, Rajaei; Naranjo Perez-Fernandez, Ijiel B.; Zuo, Jianmin; Balan, Nicolae; Godfrey, Anja; Patel, Harshil; Peters, Gordon; Rowe, Martin; Jenner, Richard G.; Sinclair, Alison J.

    2015-01-01

    Lytic replication of the human gamma herpes virus Epstein-Barr virus (EBV) is an essential prerequisite for the spread of the virus. Differential regulation of a limited number of cellular genes has been reported in B-cells during the viral lytic replication cycle. We asked whether a viral bZIP transcription factor, Zta (BZLF1, ZEBRA, EB1), drives some of these changes. Using genome-wide chromatin immunoprecipitation coupled to next-generation DNA sequencing (ChIP-seq) we established a map of Zta interactions across the human genome. Using sensitive transcriptome analyses we identified 2263 cellular genes whose expression is significantly changed during the EBV lytic replication cycle. Zta binds 278 of the regulated genes and the distribution of binding sites shows that Zta binds mostly to sites that are distal to transcription start sites. This differs from the prevailing view that Zta activates viral genes by binding exclusively at promoter elements. We show that a synthetic Zta binding element confers Zta regulation at a distance and that distal Zta binding sites from cellular genes can confer Zta-mediated regulation on a heterologous promoter. This leads us to propose that Zta directly reprograms the expression of cellular genes through distal elements. PMID:25779048

  7. Acute changes in cellular zinc alters zinc uptake rates prior to zinc transporter gene expression in Jurkat cells.

    PubMed

    Holland, Tai C; Killilea, David W; Shenvi, Swapna V; King, Janet C

    2015-12-01

    A coordinated network of zinc transporters and binding proteins tightly regulate cellular zinc levels. Canonical responses to zinc availability are thought to be mediated by changes in gene expression of key zinc transporters. We investigated the temporal relationships of actual zinc uptake with patterns of gene expression in membrane-bound zinc transporters in the human immortalized T lymphocyte Jurkat cell line. Cellular zinc levels were elevated or reduced with exogenous zinc sulfate or N,N,N',N-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN), respectively. Excess zinc resulted in a rapid 44 % decrease in the rate of zinc uptake within 10 min. After 120 min, the expression of metallothionein (positive control) increased, as well as the zinc exporter, ZnT1; however, the expression of zinc importers did not change during this time period. Zinc chelation with TPEN resulted in a rapid twofold increase in the rate of zinc uptake within 10 min. After 120 min, the expression of ZnT1 decreased, while again the expression of zinc importers did not change. Overall, zinc transporter gene expression kinetics did not match actual changes in cellular zinc uptake with exogenous zinc or TPEN treatments. This suggests zinc transporter regulation may be the initial response to changes in zinc within Jurkat cells.

  8. A Single-Cell Gene-Expression Profile Reveals Inter-Cellular Heterogeneity within Human Monocyte Subsets

    PubMed Central

    Gren, Susanne T.; Rasmussen, Thomas B.; Janciauskiene, Sabina; Håkansson, Katarina; Gerwien, Jens G.; Grip, Olof

    2015-01-01

    Human monocytes are a heterogeneous cell population classified into three different subsets: Classical CD14++CD16-, intermediate CD14++CD16+, and non-classical CD14+CD16++ monocytes. These subsets are distinguished by their differential expression of CD14 and CD16, and unique gene expression profile. So far, the variation in inter-cellular gene expression within the monocyte subsets is largely unknown. In this study, the cellular variation within each human monocyte subset from a single healthy donor was described by using a novel single-cell PCR gene-expression analysis tool. We investigated 86 different genes mainly encoding cell surface markers, and proteins involved in immune regulation. Within the three human monocyte subsets, our descriptive findings show multimodal expression of key immune response genes, such as CD40, NFⱪB1, RELA, TLR4, TLR8 and TLR9. Furthermore, we discovered one subgroup of cells within the classical monocytes, which showed alterations of 22 genes e.g. IRF8, CD40, CSF1R, NFⱪB1, RELA and TNF. Additionally one subgroup within the intermediate and non-classical monocytes also displayed distinct gene signatures by altered expression of 8 and 6 genes, respectively. Hence the three monocyte subsets can be further subdivided according to activation status and differentiation, independently of the traditional classification based on cell surface markers. Demonstrating the use and the ability to discover cell heterogeneity within defined populations of human monocytes is of great importance, and can be useful in unravelling inter-cellular variation in leukocyte populations, identifying subpopulations involved in disease pathogenesis and help tailor new therapies. PMID:26650546

  9. Functional characterization of calliphorid cell death genes and cellularization gene promoters for controlling gene expression and cell viability in early embryos.

    PubMed

    Edman, R M; Linger, R J; Belikoff, E J; Li, F; Sze, S-H; Tarone, A M; Scott, M J

    2015-02-01

    The New World screwworm fly, Cochliomyia hominivorax, and the Australian sheep blow fly, Lucilia cuprina, are major pests of livestock. The sterile insect technique was used to eradicate C. hominivorax from North and Central America. This involved area-wide releases of male and female flies that had been sterilized by radiation. Genetic systems have been developed for making 'male-only' strains that would improve the efficiency of genetic control of insect pests. One system involves induction of female lethality in embryos through activation of a pro-apoptotic gene by the tetracycline-dependent transactivator. Sex-specific expression is achieved using an intron from the transformer gene, which we previously isolated from several calliphorids. In the present study, we report the isolation of the promoters from the C. hominivorax slam and Lucilia sericata bnk cellularization genes and show that these promoters can drive expression of a GFP reporter gene in early embryos of transgenic L. cuprina. Additionally, we report the isolation of the L. sericata pro-apoptotic hid and rpr genes, identify conserved motifs in the encoded proteins and determine the relative expression of these genes at different stages of development. We show that widespread expression of the L. sericata pro-apoptotic genes was lethal in Drosophila melanogaster. The isolated gene promoters and pro-apoptotic genes could potentially be used to build transgenic embryonic sexing strains of calliphorid livestock pests.

  10. Pancreatic α- and β-cellular clocks have distinct molecular properties and impact on islet hormone secretion and gene expression.

    PubMed

    Petrenko, Volodymyr; Saini, Camille; Giovannoni, Laurianne; Gobet, Cedric; Sage, Daniel; Unser, Michael; Heddad Masson, Mounia; Gu, Guoqiang; Bosco, Domenico; Gachon, Frédéric; Philippe, Jacques; Dibner, Charna

    2017-02-15

    A critical role of circadian oscillators in orchestrating insulin secretion and islet gene transcription has been demonstrated recently. However, these studies focused on whole islets and did not explore the interplay between α-cell and β-cell clocks. We performed a parallel analysis of the molecular properties of α-cell and β-cell oscillators using a mouse model expressing three reporter genes: one labeling α cells, one specific for β cells, and a third monitoring circadian gene expression. Thus, phase entrainment properties, gene expression, and functional outputs of the α-cell and β-cell clockworks could be assessed in vivo and in vitro at the population and single-cell level. These experiments showed that α-cellular and β-cellular clocks are oscillating with distinct phases in vivo and in vitro. Diurnal transcriptome analysis in separated α and β cells revealed that a high number of genes with key roles in islet physiology, including regulators of glucose sensing and hormone secretion, are differentially expressed in these cell types. Moreover, temporal insulin and glucagon secretion exhibited distinct oscillatory profiles both in vivo and in vitro. Altogether, our data indicate that differential entrainment characteristics of circadian α-cell and β-cell clocks are an important feature in the temporal coordination of endocrine function and gene expression.

  11. Fat depot-specific differences in pref-1 gene expression and adipocyte cellularity between Wagyu and Holstein cattle.

    PubMed

    Yamada, Tomoya; Higuchi, Mikito; Nakanishi, Naoto

    2014-03-07

    Preadipocyte factor-1 (pref-1) is specifically expressed in preadipocytes and acts as a gatekeeper of adipogenesis by maintaining the preadipocyte state and preventing adipocyte differentiation. We hypothesized that the breed differences of adipogenic capacity in cattle could be explained by the expression level of pref-1. In this experiment, we studied the expression level of the pref-1 gene and adipocyte cellularity in subcutaneous and mesenteric adipose tissues of Japanese Black (Wagyu) and Holstein fattening cattle. In subcutaneous adipose tissue, there were no significant differences in the pref-1 gene expression levels and adipocyte sizes between the breeds. In contrast, the expression level of the pref-1 gene in mesenteric adipose tissue of Holsteins was significantly higher than that of Wagyu. In addition, the size of mesenteric adipocytes in Holsteins was significantly smaller than that of Wagyu. These results indicate that the breed differences of fattening cattle affect the expression pattern of the pref-1 gene and adipocyte cellularity in a fat depot-specific manner.

  12. Single-cell gene expression analyses of cellular reprogramming reveal a stochastic early and hierarchic late phase

    PubMed Central

    Buganim, Yosef; Faddah, Dina A.; Cheng, Albert W.; Itskovich, Elena; Markoulaki, Styliani; Ganz, Kibibi; Klemm, Sandy L.; van Oudenaarden, Alexander; Jaenisch, Rudolf

    2012-01-01

    During cellular reprogramming only a small fraction of cells become induced pluripotent stem cells (iPSCs). Previous analyses of gene expression during reprogramming were based on populations of cells, impeding single-cell level identification of reprogramming events. We utilized two gene expression technologies to profile 48 genes in single cells at various stages during the reprogramming process. Analysis of early stages revealed considerable variation in gene expression between cells in contrast to late stages. Expression of Esrrb, Utf1, Lin28, and Dppa2 is a better predictor for cells to progress into iPSCs than expression of Fbxo15, Fgf4, and Oct4 previously suggested to be reprogramming markers. Stochastic gene expression early in reprogramming is followed by a late hierarchical phase with Sox2 being the upstream factor in a gene expression hierarchy. Finally, downstream factors derived from the late phase, which do not include Oct4, Sox2, Klf4, c-Myc and Nanog, can activate the pluripotency circuitry. PMID:22980981

  13. PROX1 Gene is Differentially Expressed in Oral Cancer and Reduces Cellular Proliferation

    PubMed Central

    Rodrigues, Maria F.S.D.; de Oliveira Rodini, Camila; de Aquino Xavier, Flávia C.; Paiva, Katiúcia B.; Severino, Patrícia; Moyses, Raquel A.; López, Rossana M.; DeCicco, Rafael; Rocha, Lília A.; Carvalho, Marcos B.; Tajara, Eloiza H.; Nunes, Fabio D.

    2014-01-01

    Abstract Homeobox genes are a family of transcription factors that play a pivotal role in embryogenesis. Prospero homeobox 1 (PROX1) has been shown to function as a tumor suppressor gene or oncogene in various types of cancer, including oral squamous cell carcinoma (OSCC). We have previously identified PROX1 as a downregulated gene in OSCC. The aim of this study is to clarify the underlying mechanism by which PROX1 regulates tumorigenicity of OSCC cells. PROX1 mRNA and protein expression levels were first investigated in 40 samples of OSCC and in nontumor margins. Methylation and amplification analysis was also performed to assess the epigenetic and genetic mechanisms involved in controlling PROX1 expression. OSCC cell line SCC9 was also transfected to stably express the PROX1 gene. Next, SCC9-PROX1-overexpressing cells and controls were subjected to proliferation, differentiation, apoptosis, migration, and invasion assays in vitro. OSCC samples showed reduced PROX1 expression levels compared with nontumor margins. PROX1 amplification was associated with better overall survival. PROX1 overexpression reduces cell proliferation and downregulates cyclin D1. PROX1-overexpressing cells also exhibited reduced CK18 and CK19 expression and transcriptionally altered the expression of WISP3, GATA3, NOTCH1, and E2F1. Our results suggest that PROX1 functions as a tumor suppressor gene in oral carcinogenesis. PMID:25526434

  14. Cellular RNA homologous to the Abelson murine leukemia virus transforming gene: expression and relationship to the viral sequence.

    PubMed Central

    Wang, J Y; Baltimore, D

    1983-01-01

    To examine the expression of the cellular homolog of the Abelson murine leukemia virus transforming gene (the v-abl sequence), a DNA probe representing the v-abl sequence was prepared. The probe detected two cytoplasmic polyadenylic acid-containing c-abl RNAs of about 6.5 and 5.5 kilobases in a variety of rodent cells, and slightly larger RNAs were detected in human cells. These two RNA species were found in all normal tissues or cell lines examined, but at differing concentrations: liver cells had the least, fibroblastic cell lines had the most. By using a probe able to detect the cellular but not the viral gene, the two RNAs were shown to be present in Abelson murine leukemia virus-transformed cells at levels found either in their untransformed counterparts or in similar cell types transformed by other means. The target cells of the virus have a somewhat elevated level of the two RNAs although expression of the c-abl gene is not restricted to these cells. The v-abl sequence lacks 0.35 and 0.85 kilobases of the c-abl RNA on the 5' and 3' ends, respectively. Thus, the Abelson murine leukemia virus transforming gene is an internal fragment of the transcript of a normal cellular gene. Images PMID:6306446

  15. A Stable HeLa Cell Line That Inducibly Expresses Poliovirus 2Apro: Effects on Cellular and Viral Gene Expression

    PubMed Central

    Barco, Angel; Feduchi, Elena; Carrasco, Luis

    2000-01-01

    A HeLa cell clone (2A7d) that inducibly expresses the gene for poliovirus protease 2A (2Apro) under the control of tetracycline has been obtained. Synthesis of 2Apro induces severe morphological changes in 2A7d cells. One day after tetracycline removal, cells round up and a few hours later die. Poliovirus 2Apro cleaves both forms of initiation factor eIF4G, causing extensive inhibition of capped-mRNA translation a few hours after protease induction. Methoxysuccinyl-Ala-Ala-Pro-Val-chloromethylketone, a selective inhibitor of 2Apro, prevents both eIF4G cleavage and inhibition of translation but not cellular death. Expression of 2Apro still allows both the replication of poliovirus and the translation of mRNAs containing a picornavirus leader sequence, while vaccinia virus replication is drastically inhibited. Translation of transfected capped mRNA is blocked in 2A7d-On cells, while luciferase synthesis from a mRNA bearing a picornavirus internal ribosome entry site (IRES) sequence is enhanced by the presence of 2Apro. Moreover, synthesis of 2Apro in 2A7d cells complements the translational defect of a poliovirus 2Apro-defective variant. These results show that poliovirus 2Apro expression mimics some phenotypical characteristics of poliovirus-infected cells, such as cell rounding, inhibition of protein synthesis and enhancement of IRES-driven translation. This cell line constitutes a useful tool to further analyze 2Apro functions, to complement poliovirus 2Apro mutants, and to test antiviral compounds. PMID:10666269

  16. Identification of driving network of cellular differentiation from single sample time course gene expression data

    NASA Astrophysics Data System (ADS)

    Chen, Ye; Wolanyk, Nathaniel; Ilker, Tunc; Gao, Shouguo; Wang, Xujing

    Methods developed based on bifurcation theory have demonstrated their potential in driving network identification for complex human diseases, including the work by Chen, et al. Recently bifurcation theory has been successfully applied to model cellular differentiation. However, there one often faces a technical challenge in driving network prediction: time course cellular differentiation study often only contains one sample at each time point, while driving network prediction typically require multiple samples at each time point to infer the variation and interaction structures of candidate genes for the driving network. In this study, we investigate several methods to identify both the critical time point and the driving network through examination of how each time point affects the autocorrelation and phase locking. We apply these methods to a high-throughput sequencing (RNA-Seq) dataset of 42 subsets of thymocytes and mature peripheral T cells at multiple time points during their differentiation (GSE48138 from GEO). We compare the predicted driving genes with known transcription regulators of cellular differentiation. We will discuss the advantages and limitations of our proposed methods, as well as potential further improvements of our methods.

  17. Specificity of cellular expression of C. variopedatus polychaete innexin in the developing embryo: evolutionary aspects of innexins' heterogeneous gene structures.

    PubMed

    Potenza, Nicoletta; del Gaudio, Rosanna; Chiusano, Maria Luisa; Russo, Giuseppina Maria Rosaria; Geraci, Giuseppe

    2003-01-01

    Innexins are a family of membrane proteins involved in the formation of gap junctions in invertebrates. They have been found to participate in several aspects of cell differentiation and in embryonic patterning through the formation of specific intercellular communication channels. We present here data showing that the recently identified innexin of the marine worm Chaetopterus variopedatus is expressed only in particular cells of the early stage, demonstrating cell specificity of innexin expression also in polychaete annelids. Phylogenetic analysis of all known innexins results in a phylogenetic tree clearly distinguishing insect, nematode, and other invertebrate innexins. Comparative analysis of proteins and known related genes shows that the apparent similarity of protein composition, overall structural organization, and specificity of cellular expression, typical of innexins of all studied organisms, correspond to highly heterogeneous gene structures even for genes that are in close contiguity on the same chromosome. A possible evolutionary motive producing this situation is discussed.

  18. Expression of Arabidopsis FCS-Like Zinc finger genes is differentially regulated by sugars, cellular energy level, and abiotic stress.

    PubMed

    Jamsheer K, Muhammed; Laxmi, Ashverya

    2015-01-01

    Cellular energy status is an important regulator of plant growth, development, and stress mitigation. Environmental stresses ultimately lead to energy deficit in the cell which activates the SNF1-RELATED KINASE 1 (SnRK1) signaling cascade which eventually triggering a massive reprogramming of transcription to enable the plant to survive under low-energy conditions. The role of Arabidopsis thaliana FCS-Like Zinc finger (FLZ) gene family in energy and stress signaling is recently come to highlight after their interaction with kinase subunits of SnRK1 were identified. In a detailed expression analysis in different sugars, energy starvation, and replenishment series, we identified that the expression of most of the FLZ genes is differentially modulated by cellular energy level. It was found that FLZ gene family contains genes which are both positively and negatively regulated by energy deficit as well as energy-rich conditions. Genetic and pharmacological studies identified the role of HEXOKINASE 1- dependent and energy signaling pathways in the sugar-induced expression of FLZ genes. Further, these genes were also found to be highly responsive to different stresses as well as abscisic acid. In over-expression of kinase subunit of SnRK1, FLZ genes were found to be differentially regulated in accordance with their response toward energy fluctuation suggesting that these genes may work downstream to the established SnRK1 signaling under low-energy stress. Taken together, the present study provides a conceptual framework for further studies related to SnRK1-FLZ interaction in relation to sugar and energy signaling and stress response.

  19. Modulation of Estrogen Response Element-Driven Gene Expressions and Cellular Proliferation with Polar Directions by Designer Transcription Regulators

    PubMed Central

    Muyan, Mesut; Güpür, Gizem; Yaşar, Pelin; Ayaz, Gamze; User, Sırma Damla; Kazan, Hasan Hüseyin; Huang, Yanfang

    2015-01-01

    Estrogen receptor α (ERα), as a ligand-dependent transcription factor, mediates 17β-estradiol (E2) effects. ERα is a modular protein containing a DNA binding domain (DBD) and transcription activation domains (AD) located at the amino- and carboxyl-termini. The interaction of the E2-activated ERα dimer with estrogen response elements (EREs) of genes constitutes the initial step in the ERE-dependent signaling pathway necessary for alterations of cellular features. We previously constructed monomeric transcription activators, or monotransactivators, assembled from an engineered ERE-binding module (EBM) using the ERα-DBD and constitutively active ADs from other transcription factors. Monotransactivators modulated cell proliferation by activating and repressing ERE-driven gene expressions that simulate responses observed with E2-ERα. We reasoned here that integration of potent heterologous repression domains (RDs) into EBM could generate monotransrepressors that alter ERE-bearing gene expressions and cellular proliferation in directions opposite to those observed with E2-ERα or monotransactivators. Consistent with this, monotransrepressors suppressed reporter gene expressions that emulate the ERE-dependent signaling pathway. Moreover, a model monotransrepressor regulated DNA synthesis, cell cycle progression and proliferation of recombinant adenovirus infected ER-negative cells through decreasing as well as increasing gene expressions with polar directions compared with E2-ERα or monotransactivator. Our results indicate that an ‘activator’ or a ‘repressor’ possesses both transcription activating/enhancing and repressing/decreasing abilities within a chromatin context. Offering a protein engineering platform to alter signal pathway-specific gene expressions and cell growth, our approach could also be used for the development of tools for epigenetic modifications and for clinical interventions wherein multigenic de-regulations are an issue. PMID:26295471

  20. Microarray analysis of changes in cellular gene expression induced by productive infection of primary human astrocytes: implications for HAD.

    PubMed

    Kim, Seon-Young; Li, Jinliang; Bentsman, Galina; Brooks, Andrew I; Volsky, David J

    2004-12-01

    The role of astrocytes in HIV-1 associated dementia (HAD) is not well understood. HIV-1 binds efficiently to astrocytes but infects only a small fraction of the cells in vitro and in vivo. To gain insight into the biology of HIV-1-expressing astrocytes, we productively infected human fetal astrocytes with pseudotyped HIV-1 and employed Affymetrix oligonucleotide microarrays to determine global changes in cellular gene expression at the peak of virus production. With a twofold change as a cutoff, HIV-1 increased transcription of 266 genes in astrocytes and suppressed expression of 468. The functions of highly expressed genes included interferon-mediated antiviral responses (OAS1, IFIT1), intercellular contacts (SH3, glia-derived nexin), cell homing/adhesion (matrix metalloproteinases), and cell-cell signaling (neuropilin 1 and 2). Surprisingly, genes involved in innate immune responses of astrocytes were largely unaffected. The single most significant effect of HIV-1, however, was down-modulation of at least 55 genes involved in control of cell cycle, DNA replication, and cell proliferation, which were overrepresented in these categories with probability scores of 10(-10)-10(-26). Our data suggest that HIV-1 expression in astrocytes profoundly alters host cell biology, with potential consequences for the physiological function of astrocytes during HIV-1 infection in the brain.

  1. Effect of passage number on cellular response DNA-damaging agents: cell survival and gene expression

    SciTech Connect

    Chang-Liu, Chin-Mei; Wolschak, G.E.

    1996-03-01

    The effect of different passage numbers on plating efficiency, doubling time, cell growth, and radiation sensitivity was assessed in Syrian hamster embryo (SHE) cells. Changes in gene expression after UV or {gamma}-ray irradiation at different passage numbers were also examined. The SHE cells were maintained in culture medium for up to 64 passages. Cells were exposed to {sup 60}Co {gamma} rays or 254-m UV radiation. Differential display of cDNAs and Northern blots were used for the study of gene expression. With increasing passage number, SHE cells demonstrated decreased doubling time, increased plating efficiency, and a decreased yield in the number of cells per plate. Between passages 41 and 48 a ``crisis`` period was evident during which time cell growth in high serum (20%) was no longer optimal, and serum concentrations were reduced (to 10%) to maintain cell growth. Sensitivity to ionizing radiation was no different between early- and intermediate-passage cells. However, after UV exposure at low passages (passage 3), confluent cells were more sensitive to the killing effects of UV than were log-phase cells. At intermediate passages (passages 43, 48), confluent cells were slightly more radioresistant- than were log-phase cells. By passage 64, however, both confluent and log-phase cells showed similar patterns of UV sensitivity. Expression of {gamma}-actin, PCNA, and p53 transcripts did not change following UV exposure. p53 mRNA was induced following {gamma}-ray exposure of the intermediate (passage 45) epithelial cells. Differential display, however, revealed changes in expression of several transcripts following exposure to ionizing and ultraviolet radiations. The observed differences in radiation sensitivity associated with increasing passage number may be influenced by radiation-induced gene expression. We are conducting experiments to identify these genes.

  2. Derepression of hTERT gene expression promotes escape from oncogene-induced cellular senescence

    PubMed Central

    Patel, Priyanka L.; Suram, Anitha; Mirani, Neena; Bischof, Oliver; Herbig, Utz

    2016-01-01

    Oncogene-induced senescence (OIS) is a critical tumor-suppressing mechanism that restrains cancer progression at premalignant stages, in part by causing telomere dysfunction. Currently it is unknown whether this proliferative arrest presents a stable and therefore irreversible barrier to cancer progression. Here we demonstrate that cells frequently escape OIS induced by oncogenic H-Ras and B-Raf, after a prolonged period in the senescence arrested state. Cells that had escaped senescence displayed high oncogene expression levels, retained functional DNA damage responses, and acquired chromatin changes that promoted c-Myc–dependent expression of the human telomerase reverse transcriptase gene (hTERT). Telomerase was able to resolve existing telomeric DNA damage response foci and suppressed formation of new ones that were generated as a consequence of DNA replication stress and oncogenic signals. Inhibition of MAP kinase signaling, suppressing c-Myc expression, or inhibiting telomerase activity, caused telomere dysfunction and proliferative defects in cells that had escaped senescence, whereas ectopic expression of hTERT facilitated OIS escape. In human early neoplastic skin and breast tissue, hTERT expression was detected in cells that displayed features of senescence, suggesting that reactivation of telomerase expression in senescent cells is an early event during cancer progression in humans. Together, our data demonstrate that cells arrested in OIS retain the potential to escape senescence by mechanisms that involve derepression of hTERT expression. PMID:27503890

  3. Cellular Localization and Regulation of Expression of the PLET1 Gene in Porcine Placenta.

    PubMed

    Teng, Liu; Hong, Linjun; Liu, Ruize; Chen, Ran; Li, Xinyun; Yu, Mei

    2016-12-07

    The placenta expressed transcript 1 (PLET1) gene, which is expressed in placentas of pigs and mice, has been found to have a potential role in trophoblast cell fate decision in mice. Results of this study showed that the porcine PLET1 mRNA and protein were expressed exclusively in trophoblast cells on Days 15, 26, 50, and 95 of gestation (gestation length in the pig is 114 days), indicating that the PLET1 could be a useful marker for porcine trophoblast cells. Additionally, PLET1 protein was found to be redistributed from cytoplasm to the apical side of trophoblast cells as gestation progresses, which suggests a role of PLET1 in the establishment of a stable trophoblast and endometrial epithelial layers. In addition, two transcripts that differ in the 3' UTR length but encode identical protein were identified to be generated by the alternative cleavage and polyadenylation (APA), and the expression of PLET1-L transcript was significantly upregulated in porcine placentas as gestation progresses. Furthermore, we demonstrated the interaction between the miR-365-3p and PLET1 gene using luciferase assay system. Our findings imply an important role of PLET1 in the placental development in pigs.

  4. Cellular Localization and Regulation of Expression of the PLET1 Gene in Porcine Placenta

    PubMed Central

    Teng, Liu; Hong, Linjun; Liu, Ruize; Chen, Ran; Li, Xinyun; Yu, Mei

    2016-01-01

    The placenta expressed transcript 1 (PLET1) gene, which is expressed in placentas of pigs and mice, has been found to have a potential role in trophoblast cell fate decision in mice. Results of this study showed that the porcine PLET1 mRNA and protein were expressed exclusively in trophoblast cells on Days 15, 26, 50, and 95 of gestation (gestation length in the pig is 114 days), indicating that the PLET1 could be a useful marker for porcine trophoblast cells. Additionally, PLET1 protein was found to be redistributed from cytoplasm to the apical side of trophoblast cells as gestation progresses, which suggests a role of PLET1 in the establishment of a stable trophoblast and endometrial epithelial layers. In addition, two transcripts that differ in the 3′ UTR length but encode identical protein were identified to be generated by the alternative cleavage and polyadenylation (APA), and the expression of PLET1-L transcript was significantly upregulated in porcine placentas as gestation progresses. Furthermore, we demonstrated the interaction between the miR-365-3p and PLET1 gene using luciferase assay system. Our findings imply an important role of PLET1 in the placental development in pigs. PMID:27941613

  5. Alternative splicing, a new target to block cellular gene expression by poliovirus 2A protease

    SciTech Connect

    Alvarez, Enrique; Castello, Alfredo; Carrasco, Luis; Izquierdo, Jose M.

    2011-10-14

    Highlights: {yields} Novel role for poliovirus 2A protease as splicing modulator. {yields} Poliovirus 2A protease inhibits the alternative splicing of pre-mRNAs. {yields} Poliovirus 2A protease blocks the second catalytic step of splicing. -- Abstract: Viruses have developed multiple strategies to interfere with the gene expression of host cells at different stages to ensure their own survival. Here we report a new role for poliovirus 2A{sup pro} modulating the alternative splicing of pre-mRNAs. Expression of 2A{sup pro} potently inhibits splicing of reporter genes in HeLa cells. Low amounts of 2A{sup pro} abrogate Fas exon 6 skipping, whereas higher levels of protease fully abolish Fas and FGFR2 splicing. In vitro splicing of MINX mRNA using nuclear extracts is also strongly inhibited by 2A{sup pro}, leading to accumulation of the first exon and the lariat product containing the unspliced second exon. These findings reveal that the mechanism of action of 2A{sup pro} on splicing is to selectively block the second catalytic step.

  6. First cellular approach of the effects of global warming on groundwater organisms: a study of the HSP70 gene expression.

    PubMed

    Colson-Proch, Céline; Morales, Anne; Hervant, Frédéric; Konecny, Lara; Moulin, Colette; Douady, Christophe J

    2010-05-01

    Whereas the consequences of global warming at population or community levels are well documented, studies at the cellular level are still scarce. The study of the physiological or metabolic effects of such small increases in temperature (between +2 degrees C and +6 degrees C) is difficult because they are below the amplitude of the daily or seasonal thermal variations occurring in most environments. In contrast, subterranean biotopes are highly thermally buffered (+/-1 degrees C within a year), and underground water organisms could thus be particularly well suited to characterise cellular responses of global warming. To this purpose, we studied genes encoding chaperone proteins of the HSP70 family in amphipod crustaceans belonging to the ubiquitous subterranean genus Niphargus. An HSP70 sequence was identified in eight populations of two complexes of species of the Niphargus genus (Niphargus rhenorhodanensis and Niphargus virei complexes). Expression profiles were determined for one of these by reverse transcription and quantitative polymerase chain reaction, confirming the inducible nature of this gene. An increase in temperature of 2 degrees C seemed to be without effect on N. rhenorhodanensis physiology, whereas a heat shock of +6 degrees C represented an important thermal stress for these individuals. Thus, this study shows that although Niphargus individuals do not undergo any daily or seasonal thermal variations in underground water, they display an inducible HSP70 heat shock response. This controlled laboratory-based physiological experiment constitutes a first step towards field investigations of the cellular consequences of global warming on subterranean organisms.

  7. Primate Lentiviruses Modulate NF-κB Activity by Multiple Mechanisms to Fine-Tune Viral and Cellular Gene Expression

    PubMed Central

    Heusinger, Elena; Kirchhoff, Frank

    2017-01-01

    The transcription factor nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) plays a complex role during the replication of primate lentiviruses. On the one hand, NF-κB is essential for induction of efficient proviral gene expression. On the other hand, this transcription factor contributes to the innate immune response and induces expression of numerous cellular antiviral genes. Recent data suggest that primate lentiviruses cope with this challenge by boosting NF-κB activity early during the replication cycle to initiate Tat-driven viral transcription and suppressing it at later stages to minimize antiviral gene expression. Human and simian immunodeficiency viruses (HIV and SIV, respectively) initially exploit their accessory Nef protein to increase the responsiveness of infected CD4+ T cells to stimulation. Increased NF-κB activity initiates Tat expression and productive replication. These events happen quickly after infection since Nef is rapidly expressed at high levels. Later during infection, Nef proteins of HIV-2 and most SIVs exert a very different effect: by down-modulating the CD3 receptor, an essential factor for T cell receptor (TCR) signaling, they prevent stimulation of CD4+ T cells via antigen-presenting cells and hence suppress further induction of NF-κB and an effective antiviral immune response. Efficient LTR-driven viral transcription is maintained because it is largely independent of NF-κB in the presence of Tat. In contrast, human immunodeficiency virus type 1 (HIV-1) and its simian precursors have lost the CD3 down-modulation function of Nef and use the late viral protein U (Vpu) to inhibit NF-κB activity by suppressing its nuclear translocation. In this review, we discuss how HIV-1 and other primate lentiviruses might balance viral and antiviral gene expression through a tight temporal regulation of NF-κB activity throughout their replication cycle. PMID:28261165

  8. Identification, expression pattern, cellular location and potential role of the caveolin-1 gene from Artemia sinica.

    PubMed

    Li, Xuejie; Yao, Feng; Zhang, Wei; Cheng, Cheng; Chu, Bing; Liu, Yan; Mei, Yanli; Wu, Yang; Zou, Xiangyang; Hou, Lin

    2014-05-01

    Caveolins are integral membrane proteins that serve as scaffolds to recruit numerous signaling molecules. Caveolins play an important role in membrane trafficking, signal transduction, substrate transport and endocytosis in differentiated cells. In this study, a caveolin-1 gene from Artemia sinica (As-cav-1) was successfully cloned for the first time. The full-length cDNA of As-cav-1 comprises 974 bp, with a 675 bp open reading frame (ORF) that encodes a polypeptide of 224 amino acids with a caveolin scaffolding domain (CSD) and two transmembrane domains. Multiple sequence alignment revealed that the putative As-CAV-1 protein sequence was relatively conserved across species, especially in the CSD domain. Real-time PCR revealed high levels of the As-cav-1 transcript at 0h of embryo development. Furthermore, As-cav-1 transcripts were highly upregulated under high salinity (200‰) and low temperature stresses (15°C). To further characterize As-cav-1, recombinant pET30a-cav-1 protein was expressed using a prokaryotic expression system. The recombinant protein comprised 290 amino acids with a theoretical molecular weight of 32kDa, and a predicted isoelectric point of 5.6. Western blotting of the expression levels of As-CAV-1 during different embryo development stages revealed that As-CAV-1 levels decreased gradually during development stages from 0 h to 40 h, and increased at 3d. Furthermore, western blotting showed that As-CAV-1 was upregulated to its highest expression level by low temperature stress (15°C) and high salinity. Confocal laser microscopy analysis, using antibodies generated against the recombinant As-CAV-1 protein, showed that As-CAV-1 was mostly located in the cell membrane. Our results suggested that As-cav-1 plays a vital role in protecting embryos from high salt damage and low temperature stress, especially during post-diapause embryonic development.

  9. Reprogramming the genetic code: the emerging role of ribosomal frameshifting in regulating cellular gene expression

    PubMed Central

    Advani, Vivek M.; Dinman, Jonathan D.

    2016-01-01

    Reading frame maintenance is a critical property of ribosomes. However, a number of genetic elements have been described that can induce ribosomes to shift on mRNAs, the most well understood of which are a class that directs ribosomal slippage by one base in 5′ (-1) direction. This is referred to as programmed -1 ribosomal frameshifting (-1 PRF). Recently, a new -1 PRF promoting element was serendipitously discovered in a study examining the effects of stretches of adenosines in the coding sequences of mRNAs. Here, we discuss this finding, recent studies describing how -1 PRF is used to control gene expression in eukaryotes, and how -1 PRF is itself regulated. The implications of dysregulation of -1 PRF on human health are examined, as are possible new areas in which novel -1 PRF promoting elements might be discovered. PMID:26661048

  10. P-TEFb Kinase Complex Phosphorylates Histone H1 to Regulate Expression of Cellular and HIV-1 Genes*

    PubMed Central

    O'Brien, Siobhan K.; Cao, Hong; Nathans, Robin; Ali, Akbar; Rana, Tariq M.

    2010-01-01

    Transcription of HIV-1 genes depends on the RNA polymerase II kinase and elongation factor positive transcription elongation factor b (P-TEFb), the complex of cyclin T1 and CDK9. Recent evidence suggests that regulation of transcription by P-TEFb involves chromatin binding and modifying factors. To determine how P-TEFb may connect chromatin remodeling to transcription, we investigated the relationship between P-TEFb and histone H1. We identify histone H1 as a substrate for P-TEFb involved in cellular and HIV-1 transcription. We show that P-TEFb interacts with H1 and that P-TEFb inhibition by RNAi, flavopiridol, or dominant negative CDK9 expression correlates with loss of phosphorylation and mobility of H1 in vivo. Importantly, P-TEFb directs H1 phosphorylation in response to wild-type HIV-1 infection, but not Tat-mutant HIV-1 infection. Our results show that P-TEFb phosphorylates histone H1 at a specific C-terminal phosphorylation site. Expression of a mutant H1.1 that cannot be phosphorylated by P-TEFb also disrupts Tat transactivation in an HIV reporter cell line as well as transcription of the c-fos and hsp70 genes in HeLa cells. We identify histone H1 as a novel P-TEFb substrate, and our results suggest new roles for P-TEFb in both cellular and HIV-1 transcription. PMID:20551309

  11. Effects of polyamines on cellular innate immune response and the expression of immune-relevant genes in gilthead seabream leucocytes.

    PubMed

    Reyes-Becerril, Martha; Ascencio-Valle, Felipe; Tovar-Ramírez, Dariel; Meseguer, José; Esteban, María Ángeles

    2011-01-01

    It is well known that the polyamines spermidine and spermine, along with the diamine putrescine, are involved in many cellular processes and they are known to play an important role in the control of the innate immune response in higher vertebrates. However, to the best of our knowledge, no studies have focused on their immunological implications in other vertebrates, such as fish. For this reason, the effects of polyamines on the cellular innate immune response and immune-related gene expression were evaluated in vitro, using seabream head-kidney leucocytes (HKL). For this study, head-kidney leucocytes were incubated with the polyamines putrescine, spermine or spermidine (0.005 and 0.0025%) for 0.50, 1, 2 or 4 h. No significant effect was observed on either leucocyte viability or the innate cellular immune responses (peroxidase content and phagocytic and respiratory burst activities). The polyamines produced an increase in respiratory burst and phagocytic ability when leucocytes were incubated principally with putrescine (0.005 and 0.0025%) after 2 and 4 h of the experiment. Finally, the expression levels of immune-associated genes (IgM, MHCIα, MHCIIα, C3, IL-1β, CD8, Hep, NCCRP-1, CSF-1 and TLR) were quantified by real-time PCR and some of them (C3, MHCI, CD8, IgM and Hep) were up-regulated by the higher polyamine concentration. Further studies are needed to ascertain how polyamines control the immune system of seabream as well as which mechanisms are involved.

  12. Effects of Selected Egyptian Honeys on the Cellular Ultrastructure and the Gene Expression Profile of Escherichia coli

    PubMed Central

    Elkhatib, Walid F.

    2016-01-01

    The purpose of this study was to: (i) evaluate the antibacterial activities of three Egyptian honeys collected from different floral sources (namely, citrus, clover, and marjoram) against Escherichia coli; (ii) investigate the effects of these honeys on bacterial ultrastructure; and (iii) assess the anti-virulence potential of these honeys, by examining their impacts on the expression of eight selected genes (involved in biofilm formation, quorum sensing, and stress survival) in the test organism. The minimum inhibitory concentration (MIC) of the honey samples against E. coli ATCC 8739 were assessed by the broth microdilution assay in the presence and absence of catalase enzyme. Impacts of the honeys on the cellular ultrastructure and the expression profiles of the selected genes of E. coli were examined using transmission electron microscopy (TEM) and quantitative real-time polymerase chain reaction (qPCR) analysis, respectively. The susceptibility tests showed promising antibacterial activities of all the tested honeys against E. coli. This was supported by the TEM observations, which revealed “ghost” cells lacking DNA, in addition to cells with increased vacuoles, and/or with irregular shrunken cytoplasm. Among the tested honeys, marjoram exhibited the highest total antibacterial activity and the highest levels of peroxide-dependent activity. The qPCR analysis showed that all honey-treated cells share a similar overall pattern of gene expression, with a trend toward reduced expression of the virulence genes of interest. Our results indicate that some varieties of the Egyptian honey have the potential to be effective inhibitor and virulence modulator of E. coli via multiple molecular targets. PMID:26954570

  13. Molecular cloning, expression analysis and cellular localization of an LFRFamide gene in the cuttlefish Sepiella japonica.

    PubMed

    Cao, Zi-Hao; Sun, Lian-Lian; Chi, Chang-Feng; Liu, Hui-Hui; Zhou, Li-Qing; Lv, Zhen-Ming; Wu, Chang-Wen

    2016-06-01

    Neuropeptides are important regulators of physiological processes in metazoans, such as feeding, reproduction, and heart activities. In this study, an LFRFamide gene was identified from the cuttlefish Sepiella japonica (designated as SjLFRFamide). The full-length sequence of SjLFRFamide cDNA has 841bp, and the open reading frame contains 567bp encoding 188 amino acids, which shared high similarity with precursor SOFaRP2 from Sepia officinalis. The deduced SjLFRFamdie precursor protein contains a signal peptide and four different FLPs (FMRFamide-like peptides): one pentapeptide (TIFRFamide), two hexapeptides (NSLFRFamide and GNLFRFamide) and one heptapeptide (PHTPFRFamide). Multiple sequence alignment showed that SjLFRFamide contains rather conserved mature peptides, which all ended in FRF. The phylogenetic analysis suggests that SjLFRFamide belongs to the LFRFamide subfamily. The tissue distribution analysis through quantitative real-time PCR method showed that SjLFRFamide mRNA is significantly expressed in the brain, and slight trace are detected in female nidamental gland and accessory nidamental gland. In situ hybridization assay of the brain indicated that SjLFRFamide is transcribed in several different functional lobes, suggesting SjLFRFamide might associate with multiple physiological regulations, such as feeding, chromatophore regulation and reproduction. This is the first study describing LFRFamide in S. japonica, which might have great importance for cuttlefish artificial breeding.

  14. Dietary Restriction Mitigates Cocaine-Induced Alterations of Olfactory Bulb Cellular Plasticity and Gene Expression, and Behavior

    PubMed Central

    Xu, Xiangru; Mughal, Mohamed R.; Hall, F. Scott; Perona, Maria T.G.; Pistell, Paul J.; Lathia, Justin D; Chigurupati, Srinivasulu; Becker, Kevin G; Ladenheim, Bruce; Niklason, Laura E; Uhl, George R.; Cadet, Jean Lud; Mattson, Mark P.

    2010-01-01

    Because the olfactory system plays a major role in food consumption, and because “food addiction” and associated morbidities have reached epidemic proportions, we tested the hypothesis that dietary energy restriction can modify adverse effects of cocaine on behavior and olfactory cellular and molecular plasticity. Mice maintained on an alternate day fasting (ADF) diet exhibited increased baseline locomotion and increased cocaine-sensitized locomotion during cocaine conditioning, despite no change in cocaine conditioned place preference, compared to mice fed ad libitum. Levels of dopamine and its metabolites in the olfactory bulb (OB) were suppressed in mice on the ADF diet compared to mice on the control diet, independent of acute or chronic cocaine treatment. The expression of several enzymes involved in dopamine metabolism including tyrosine hydroxylase, monoamine oxidases A and B (MAOA), and catechol-O-methyltransferase were significantly reduced in OBs of mice on the ADF diet. Both acute and chronic administration of cocaine suppressed the production of new OB cells, and this effect of cocaine was attenuated in mice on the ADF diet. Cocaine administration to mice on the control diet resulted in up-regulation of OB genes involved in mitochondrial energy metabolism, synaptic plasticity, cellular stress responses, and calcium- and cyclic AMP-mediated signaling, whereas multiple olfactory receptor genes were down-regulated by cocaine treatment. ADF abolished many of the effects of cocaine on OB gene expression. Our findings reveal that dietary energy intake modifies the neural substrates underlying some of the behavioral and physiological responses to repeated cocaine treatment, and also suggest novel roles for the olfactory system in addiction. The data further suggest that modification of dietary energy intake could provide a novel potential approach to addiction treatments. PMID:20456017

  15. Adipose depots differ in cellularity, adipokines produced, gene expression, and cell systems

    PubMed Central

    Dodson, Michael V; Du, Min; Wang, Songbo; Bergen, Werner G; Fernyhough-Culver, Melinda; Basu, Urmila; Poulos, Sylvia P; Hausman, Gary J

    2014-01-01

    The race to manage the health concerns related to excess fat deposition has spawned a proliferation of clinical and basic research efforts to understand variables including dietary uptake, metabolism, and lipid deposition by adipocytes. A full appreciation of these variables must also include a depot-specific understanding of content and location in order to elucidate mechanisms governing cellular development and regulation of fat deposition. Because adipose tissue depots contain various cell types, differences in the cellularity among and within adipose depots are presently being documented to ascertain functional differences. This has led to the possibility of there being, within any one adipose depot, cellular distinctions that essentially result in adipose depots within depots. The papers comprising this issue will underscore numerous differences in cellularity (development, histogenesis, growth, metabolic function, regulation) of different adipose depots. Such information is useful in deciphering adipose depot involvement both in normal physiology and in pathology. Obesity, diabetes, metabolic syndrome, carcass composition of meat animals, performance of elite athletes, physiology/pathophysiology of aging, and numerous other diseases might be altered with a greater understanding of adipose depots and the cells that comprise them—including stem cells—during initial development and subsequent periods of normal/abnormal growth into senescence. Once thought to be dormant and innocuous, the adipocyte is emerging as a dynamic and influential cell and research will continue to identify complex physiologic regulation of processes involved in adipose depot physiology. PMID:26317047

  16. Adrenoceptors in Brain: Cellular Gene Expression and Effects on Astrocytic Metabolism and [Ca2+]i

    PubMed Central

    Hertz, Leif; Lovatt, Ditte; Goldman, Steven A.; Nedergaard, Maiken

    2010-01-01

    Recent in vivo studies have established astrocytes as a major target for locus coeruleus activation (Bekar et al., Cereb. Cortex 18, 2789–2795), renewing interest in cell culture studies on noradrenergic effects on astrocytes in primary cultures and calling for additional information about the expression of adrenoceptor subtypes on different types of brain cells. In the present communication, mRNA expression of α1-, α2- and β-adrenergic receptors and their subtypes was determined in freshly-isolated, cell marker-defined populations of astrocytes, NG2-positive cells, microglia, endothelial cells, and Thy1-positive neurons (mainly glutamatergic projection neurons) in murine cerebral cortex. Immediately after dissection of frontal, parietal and occipital cortex of 10–12-week-old transgenic mice, which combined each cell-type marker with a specific fluorescent signal, the tissue was digested, triturated and centrifuged, yielding a solution of dissociated cells of all types, which were separated by fluorescence-activated cell sorting (FACS). mRNA expression in each cell fraction was determined by microarray analysis. α1A-Receptors were unequivocally expressed in astrocytes and NG2-positive cells, but absent in other cell types, and α1B-receptors were not expressed in any cell population. Among α2-receptors only α2A-receptors were expressed, unequivocally in astrocytes and NG-positive cells, tentatively in microglia and questionably in Thy1-positive neurons and endothelial cells. β1-Receptors were unequivocally expressed in astrocytes, tentatively in microglia, and questionably in neurons and endothelial cells, whereas β2-adrenergic receptors showed tentative expression in neurons and astrocytes and unequivocal expression in other cell types. This distribution was supported by immunochemical data and its relevance established by previous studies in well-differentiated primary cultures of mouse astrocytes, showing that stimulation of α2-adrenoceptors

  17. Relating Chemical Structure to Cellular Response: An Integrative Analysis of Gene Expression, Bioactivity, and Structural Data Across 11,000 Compounds.

    PubMed

    Chen, B; Greenside, P; Paik, H; Sirota, M; Hadley, D; Butte, A J

    2015-10-01

    A central premise in systems pharmacology is that structurally similar compounds have similar cellular responses; however, this principle often does not hold. One of the most widely used measures of cellular response is gene expression. By integrating gene expression data from Library of Integrated Network-based Cellular Signatures (LINCS) with chemical structure and bioactivity data from PubChem, we performed a large-scale correlation analysis of chemical structures and gene expression profiles of over 11,000 compounds taking into account confounding factors such as biological conditions (e.g., cell line, dose) and bioactivities. We found that structurally similar compounds do indeed yield similar gene expression profiles. There is an ∼20% chance that two structurally similar compounds (Tanimoto Coefficient ≥ 0.85) share significantly similar gene expression profiles. Regardless of structural similarity, two compounds tend to share similar gene expression profiles in a cell line when they are administrated at a higher dose or when the cell line is sensitive to both compounds.

  18. Kinetics of the cellular intake of a gene expression inducer at high concentrations.

    PubMed

    Tran, Huy; Oliveira, Samuel M D; Goncalves, Nadia; Ribeiro, Andre S

    2015-09-01

    From in vivo single-event measurements of the transient and steady-state transcription activity of a single-copy lac-ara-1 promoter in Escherichia coli, we characterize the intake kinetics of its inducer (IPTG) from the media. We show that the empirical data are well-fit by a model of intake assuming a bilayer membrane, with the passage through the second layer being rate-limiting, coupled to a stochastic, sub-Poissonian, multi-step transcription process. Using this model, we show that for a wide range of extracellular inducer levels (up to 1.25 mM) the intake process is diffusive-like, suggesting unsaturated membrane permeability. Inducer molecules travel from the periplasm to the cytoplasm in, on average, 31.7 minutes, strongly affecting cells' response time. The novel methodology followed here should aid the study of cellular intake mechanisms at the single-event level.

  19. Myocardial Gene Expression Profiling to Predict and Identify Cardiac Allograft Acute Cellular Rejection: The GET-Study

    PubMed Central

    Bodez, Diane; Hocini, Hakim; Tchitchek, Nicolas; Tisserand, Pascaline; Benhaiem, Nicole; Barau, Caroline; Kharoubi, Mounira; Guellich, Aziz; Guendouz, Soulef; Radu, Costin; Couetil, Jean-Paul; Ghaleh, Bijan; Dubois-Randé, Jean-Luc; Teiger, Emmanuel; Hittinger, Luc

    2016-01-01

    Aims Serial invasive endomyocardial biopsies (EMB) remain the gold standard for acute cellular rejection (ACR) diagnosis. However histological grading has several limitations. We aimed to explore the value of myocardial Gene Expression Profiling (GEP) for diagnosing and identifying predictive biomarkers of ACR. Methods A case-control study nested within a retrospective heart transplant patients cohort included 126 patients with median (IQR) age 50 (41–57) years and 111 (88%) males. Among 1157 EMB performed, 467 were eligible (i.e, corresponding to either ISHLT grade 0 or ≥3A), among which 36 were selected for GEP according to the grading: 0 (CISHLT, n = 13); rejection ≥3A (RISHLT, n = 13); 0 one month before ACR (BRISHLT, n = 10). Results We found 294 genes differentially expressed between CISHLT and RISHLT, mainly involved in immune activation, and inflammation. Hierarchical clustering showed a clear segregation of CISHLT and RISHLT groups and heterogeneity of GEP within RISHLT. All EMB presented immune activation, but some RISHLT EMB were strongly subject to inflammation, whereas others, closer to CISHLT, were characterized by structural modifications with lower inflammation level. We identified 15 probes significantly different between BRISHLT and CISHLT, including the gene of the muscular protein TTN. This result suggests that structural alterations precede inflammation in ACR. Linear Discriminant Analysis based on these 15 probes was able to identify the histological status of every 36 samples. Conclusion Myocardial GEP is a helpful method to accurately diagnose ACR, and predicts rejection one month before its histological occurrence. These results should be considered in cardiac allograft recipients’ care. PMID:27898719

  20. Minireview: Long Noncoding RNAs: New “Links” Between Gene Expression and Cellular Outcomes in Endocrinology

    PubMed Central

    Sun, Miao

    2013-01-01

    Recent advances in sequencing technologies have revealed that the genome is extensively transcribed, yielding a large repertoire of noncoding RNAs. These include long noncoding RNAs (lncRNAs), mRNA-like molecules that do not code for proteins, which are emerging as a new class of RNAs that play important roles in a variety of cellular processes. Ongoing studies are revealing new insights about lncRNAs, including their physiological functions, disease relationships, and molecular mechanisms of action. Characterized lncRNAs have been shown to interact with and modulate the activity of other RNAs and protein partners, leading to alterations in transcriptional and posttranscriptional regulatory processes. In this review, we summarize the key features of lncRNAs, their molecular mechanisms of action, biological functions, and therapeutic implications, particularly as they apply to the field of molecular endocrinology. In addition, we provide a brief overview of how molecular biologists are beginning to probe the identity, mechanisms, and functions of this emerging class of RNA molecules. PMID:23885095

  1. Analysis of Epidermal Growth Factor Receptor Related Gene Expression Changes in a Cellular and Animal Model of Parkinson’s Disease

    PubMed Central

    Kim, In-Su; Koppula, Sushruta; Park, Shin-Young; Choi, Dong-Kug

    2017-01-01

    We employed transcriptome analysis of epidermal growth factor receptor related gene expression changes in cellular and animal models of Parkinson’s disease (PD). We used a well-known Parkinsonian toxin 1-methyl-4-phenylpyridine (MPP+) to induce neuronal apoptosis in the human neuroblastoma SH-SY5Y cell line. The MPP+-treatment of SH-SY5Y cells was capable of inducing neuro-apoptosis, but it remains unclear what kinds of transcriptional genes are affected by MPP+ toxicity. Therefore the pathways that were significantly perturbed in MPP+ treated human neuroblastoma SH-SY5Y cells were identified based on genome-wide gene expression data at two time points (24 and 48 h). We found that the Epidermal Growth Factor Receptor (EGFR) pathway-related genes showed significantly differential expression at all time points. The EGFR pathway has been linked to diverse cellular events such as proliferation, differentiation, and apoptosis. Further, to evaluate the functional significance of the altered EGFR related gene expression observed in MPP+-treated SH-SY5Y cells, the EGFR related GJB2 (Cx26) gene expression was analyzed in an MPP+-intoxicated animal PD model. Our findings identify that the EGFR signaling pathway and its related genes, such as Cx26, might play a significant role in dopaminergic (DAergic) neuronal cell death during the process of neuro-apoptosis and therefore can be focused on as potential targets for therapeutic intervention. PMID:28212331

  2. Temporal regulation of global gene expression and cellular morphology in Xenopus kidney cells in response to clinorotation

    NASA Astrophysics Data System (ADS)

    Kitamoto, Junko; Fukui, Akimasa; Asashima, Makoto

    Here, we report changes gene expression and morphology of the renal epithelial cell line, A6, which was derived from Xenopus laevis adult kidney that had been induced by long-term culturing with a three-dimensional clinostat. An oligo microarray analysis on the A6 cells showed that mRNA levels for 52 out of 8091 genes were significantly altered in response to clinorotation. On day 5, there was no dramatic change in expression level, but by day 8 and day 10, either upregulation or downregulation of gene expression became evident. By day 15, the expression levels of 18 out of 52 genes had returned to the original levels, while the remaining 34 genes maintained the altered levels of expression. Quantitative analyses of gene expression by real-time PCR confirmed that changes in the mRNA levels of selected genes were found only under clinorotation and not under hypergravity (7 g) or ground control. Morphological changes including loss of dome-like structures and disorganization of both E-cadherin adherence junctions and cortical actin were also observed after 10 days of culturing with clinorotation. These results revealed that the expression of selected genes was altered specifically in A6 cells cultured under clinorotation.

  3. Murine Hyperglycemic Vasculopathy and Cardiomyopathy: Whole-Genome Gene Expression Analysis Predicts Cellular Targets and Regulatory Networks Influenced by Mannose Binding Lectin

    PubMed Central

    Zou, Chenhui; La Bonte, Laura R.; Pavlov, Vasile I.; Stahl, Gregory L.

    2012-01-01

    Hyperglycemia, in the absence of type 1 or 2 diabetes, is an independent risk factor for cardiovascular disease. We have previously demonstrated a central role for mannose binding lectin (MBL)-mediated cardiac dysfunction in acute hyperglycemic mice. In this study, we applied whole-genome microarray data analysis to investigate MBL’s role in systematic gene expression changes. The data predict possible intracellular events taking place in multiple cellular compartments such as enhanced insulin signaling pathway sensitivity, promoted mitochondrial respiratory function, improved cellular energy expenditure and protein quality control, improved cytoskeleton structure, and facilitated intracellular trafficking, all of which may contribute to the organismal health of MBL null mice against acute hyperglycemia. Our data show a tight association between gene expression profile and tissue function which might be a very useful tool in predicting cellular targets and regulatory networks connected with in vivo observations, providing clues for further mechanistic studies. PMID:22375142

  4. Cellular responses and gene expression profile changes due to bleomycin-induced DNA damage in human fibroblasts in space

    PubMed Central

    Kidane, Yared; Feiveson, Alan; Stodieck, Louis; Karouia, Fathi; Ramesh, Govindarajan; Rohde, Larry; Wu, Honglu

    2017-01-01

    Living organisms in space are constantly exposed to radiation, toxic chemicals or reactive oxygen species generated due to increased levels of environmental and psychological stresses. Understanding the impact of spaceflight factors, microgravity in particular, on cellular responses to DNA damage is essential for assessing the radiation risk for astronauts and the mutation rate in microorganisms. In a study conducted on the International Space Station, confluent human fibroblasts in culture were treated with bleomycin for three hours in the true microgravity environment. The degree of DNA damage was quantified by immunofluorescence staining for γ-H2AX, which is manifested in three types of staining patterns. Although similar percentages of these types of patterns were found between flight and ground cells, there was a slight shift in the distribution of foci counts in the flown cells with countable numbers of γ-H2AX foci. Comparison of the cells in confluent and in exponential growth conditions indicated that the proliferation rate between flight and the ground may be responsible for such a shift. We also performed a microarray analysis of gene expressions in response to bleomycin treatment. A qualitative comparison of the responsive pathways between the flown and ground cells showed similar responses with the p53 network being the top upstream regulator. The microarray data was confirmed with a PCR array analysis containing a set of genes involved in DNA damage signaling; with BBC3, CDKN1A, PCNA and PPM1D being significantly upregulated in both flight and ground cells after bleomycin treatment. Our results suggest that whether microgravity affects DNA damage response in space can be dependent on the cell type and cell growth condition. PMID:28248986

  5. Tetracapsuloides bryosalmonae infection affects the expression of genes involved in cellular signal transduction and iron metabolism in the kidney of the brown trout Salmo trutta.

    PubMed

    Kumar, Gokhlesh; Sarker, Subhodeep; Menanteau-Ledouble, Simon; El-Matbouli, Mansour

    2015-06-01

    Tetracapsuloides bryosalmonae is an enigmatic endoparasite which causes proliferative kidney disease in various species of salmonids in Europe and North America. The life cycle of the European strain of T. bryosalmonae generally completes in an invertebrate host freshwater bryozoan and vertebrate host brown trout (Salmo trutta) Linnaeus, 1758. Little is known about the gene expression in the kidney of brown trout during the developmental stages of T. bryosalmonae. In the present study, quantitative real-time PCR was applied to quantify the target genes of interest in the kidney of brown trout at different time points of T. bryosalmonae development. PCR primers specific for target genes were designed and optimized, and their gene expression levels were quantified in the cDNA kidney samples using SYBR Green Supermix. Expression of Rab GDP dissociation inhibitor beta, integral membrane protein 2B, NADH dehydrogenase 1 beta subcomplex subunit 6, and 26S protease regulatory subunit S10B were upregulated significantly in infected brown trout, while the expression of the ferritin M middle subunit was downregulated significantly. These results suggest that host genes involved in cellular signal transduction, proteasomal activities, including membrane transporters and cellular iron storage, are differentially upregulated or downregulated in the kidney of brown trout during parasite development. The gene expression pattern of infected renal tissue may support the development of intraluminal sporogonic stages of T. bryosalmonae in the renal tubular lumen of brown trout which may facilitate the release of viable parasite spores to transmit to the invertebrate host bryozoan.

  6. Loss of cone cyclic nucleotide-gated channel leads to alterations in light response modulating system and cellular stress response pathways: a gene expression profiling study

    PubMed Central

    Ma, Hongwei; Thapa, Arjun; Morris, Lynsie M.; Michalakis, Stylianos; Biel, Martin; Frank, Mark Barton; Bebak, Melissa; Ding, Xi-Qin

    2013-01-01

    The cone photoreceptor cyclic nucleotide-gated (CNG) channel is essential for central and color vision and visual acuity. Mutations in the channel subunits CNGA3 and CNGB3 are associated with achromatopsia and cone dystrophy. We investigated the gene expression profiles in mouse retina with CNG channel deficiency using whole genome expression microarrays. As cones comprise only 2 to 3% of the total photoreceptor population in the wild-type mouse retina, the mouse lines with CNG channel deficiency on a cone-dominant background, i.e. Cnga3−/−/Nrl−/− and Cngb3−/−/Nrl−/− mice, were used in our study. Comparative data analysis revealed a total of 105 genes altered in Cnga3−/−/Nrl−/− and 92 in Cngb3−/−/Nrl−/− retinas, relative to Nrl−/− retinas, with 27 genes changed in both genotypes. The differentially expressed genes primarily encode proteins associated with cell signaling, cellular function maintenance and gene expression. Ingenuity pathway analysis (IPA) identified 26 and 9 canonical pathways in Cnga3−/−/Nrl−/− and Cngb3−/−/Nrl−/− retinas, respectively, with 6 pathways being shared. The shared pathways include phototransduction, cAMP/PKA-mediated signaling, endothelin signaling, and EIF2/endoplasmic reticulum (ER) stress, whereas the IL-1, CREB, and purine metabolism signaling were found to specifically associate with Cnga3 deficiency. Thus, CNG channel deficiency differentially regulates genes that affect cell processes such as phototransduction, cellular survival and gene expression, and such regulations play a crucial role(s) in the retinal adaptation to impaired cone phototransduction. Though lack of Cnga3 and Cngb3 shares many common pathways, deficiency of Cnga3 causes more significant alterations in gene expression. This work provides insights into how cones respond to impaired phototransduction at the gene expression levels. PMID:23740940

  7. In vivo monitoring of transfected DNA, gene expression kinetics, and cellular immune responses in mice immunized with a human NIS gene-expressing plasmid.

    PubMed

    Son, Hye-Youn; Jeon, Yong-Hyun; Chung, June-Key; Kim, Chul-Woo

    2016-12-01

    In assessing the effectiveness of DNA vaccines, it is important to monitor: (1) the kinetics of target gene expression in vivo; and (2) the movement of cells that become transfected with the plasmid DNA used in the immunization of a subject. In this study, we used, as a visual imaging marker, expression of the transfected human sodium/iodide symporter (hNIS) gene, which enhances intracellular radio-pertechnetate (TcO4-) accumulation. After intradermal (i.d.) and systemic injection of mice with pcDNA-hNIS and radioactive Technetium-99m (Tc-99m), respectively, whole-body images were obtained by nuclear scintigraphy. The migration of mice cells transfected with the hNIS gene was monitored over a 2-week period by gamma-radioactivity counting of isolated cell populations and was demonstrated in peripheral lymphoid tissues, especially in the draining lymph nodes (dLNs). Beginning at 24 h after DNA inoculation and continuing for the 2-week monitoring period, hNIS-expressing cells were observed specifically in the T-cell-rich zones of the paracortical area of the dLNs. Over the same time period, high levels of INF-γ-secreting CD8 T-cells were found in the dLNs of the pcDNA-hNIS immunized mice. Tumor growth was also significantly retarded in the mice that received hNIS DNA immunization followed by inoculation with CT26 colorectal adenocarcinoma cells that had been transfected with the rat NIS gene (rNIS), which is 93% homologous to the hNIS gene. In conclusion, mouse cells transfected with hNIS DNA after i.d. immunization were found to traffic to the dLNs, and hNIS gene expression in these cells continued for at least 2 weeks post immunization. Furthermore, sequential presentation of NIS DNA to T-cells by migratory antigen presenting cells could induce NIS DNA-specific Th1 immune responses and thus retard the growth of NIS-expressing tumors.

  8. Cancer gene therapy targeting cellular apoptosis machinery.

    PubMed

    Jia, Lin-Tao; Chen, Si-Yi; Yang, An-Gang

    2012-11-01

    The unraveling of cellular apoptosis machinery provides novel targets for cancer treatment, and gene therapy targeting this suicidal system has been corroborated to cause inflammation-free autonomous elimination of neoplastic cells. The apoptotic machinery can be targeted by introduction of a gene encoding an inducer, mediator or executioner of apoptotic cell death or by inhibition of anti-apoptotic gene expression. Strategies targeting cancer cells, which are achieved by selective gene delivery, specific gene expression or secretion of target proteins via genetic modification of autologous cells, dictate the outcome of apoptosis-based cancer gene therapy. Despite so far limited clinical success, gene therapy targeting the apoptotic machinery has great potential to benefit patients with threatening malignancies provided the availability of efficient and specific gene delivery and administration systems.

  9. No Effect of the Transforming Growth Factor {beta}1 Promoter Polymorphism C-509T on TGFB1 Gene Expression, Protein Secretion, or Cellular Radiosensitivity

    SciTech Connect

    Reuther, Sebastian; Metzke, Elisabeth; Bonin, Michael; Petersen, Cordula; Dikomey, Ekkehard; Raabe, Annette

    2013-02-01

    Purpose: To study whether the promoter polymorphism (C-509T) affects transforming growth factor {beta}1 gene (TGFB1) expression, protein secretion, and/or cellular radiosensitivity for both human lymphocytes and fibroblasts. Methods and Materials: Experiments were performed with lymphocytes taken either from 124 breast cancer patients or 59 pairs of normal monozygotic twins. We used 15 normal human primary fibroblast strains as controls. The C-509T genotype was determined by polymerase chain reaction-restriction fragment length polymorphism or TaqMan single nucleotide polymorphism (SNP) genotyping assay. The cellular radiosensitivity of lymphocytes was measured by G0/1 assay and that of fibroblasts by colony assay. The amount of extracellular TGFB1 protein was determined by enzyme-linked immunosorbent assay, and TGFB1 expression was assessed via microarray analysis or reverse transcription-polymerase chain reaction. Results: The C-509T genotype was found not to be associated with cellular radiosensitivity, neither for lymphocytes (breast cancer patients, P=.811; healthy donors, P=.181) nor for fibroblasts (P=.589). Both TGFB1 expression and TGFB1 protein secretion showed considerable variation, which, however, did not depend on the C-509T genotype (protein secretion: P=.879; gene expression: lymphocytes, P=.134, fibroblasts, P=.605). There was also no general correlation between TGFB1 expression and cellular radiosensitivity (lymphocytes, P=.632; fibroblasts, P=.573). Conclusion: Our data indicate that any association between the SNP C-509T of TGFB1 and risk of normal tissue toxicity cannot be ascribed to a functional consequence of this SNP, either on the level of gene expression, protein secretion, or cellular radiosensitivity.

  10. Age-related changes in cellular protection, purification, and inflammation-related gene expression: role of dietary phytonutrients.

    PubMed

    Mastaloudis, Angela; Wood, Steven M

    2012-07-01

    Oxidative injury and inflammation are intimately involved in the aging process and the development of age-related diseases. To date, most nutritional antiaging strategies have focused solely on the delivery of exogenous antioxidants to combat the negative effects of aging. A promising new strategy is to identify nutrients and phytochemicals that can directly target intrinsic cytoprotective mechanisms, including modulation of the expression of (1) genes involved in the detoxification of xenobiotics, (2) genes involved in the synthesis and regulation of intrinsic antioxidants and antioxidant enzymes, (3) genes involved in the regulation of inflammation, and (4) vitagenes. The purpose of this review is to provide an overview of the age-related changes in gene expression related to oxidative stress, detoxification, and inflammatory processes, and to discuss natural compounds with the potential to oppose age-related changes in gene expression related to these processes, which therefore may be suitable for use in human antiaging research.

  11. Genome-wide analysis of phylogeny, expression profile and sub-cellular localization of SKP1-Like genes in wild tomato.

    PubMed

    Zhang, YueQin; Wang, CuiPing; Lin, QingFang; Gao, FengHua; Ma, Yan; Zhang, Min; Lin, YueHui; Ma, QingHu; Hua, XueJun

    2015-09-01

    SKP1 is a core component of SCF complex, a major type of E3 ubiquitin ligase catalyzing the last step in ubiquitin-mediated protein degradation pathway. In present study, SKP1 gene family in Solanum pimpinellifolium (SSK), a wild species of tomato, was investigated. A total of 19 SSK genes were identified through homologous search. Their chromosomal locations, gene structures, phylogeny, expression profiles, sub-cellular localizations and protein-protein interaction patterns with putative F-box proteins were analyzed in detail. The high homology and similar expression patterns among clustered SSK genes in chromosome suggested that they may have evolved from duplication events and are functionally redundant. Sub-cellular localization indicated that most of the SSK proteins are distributed in both cytosol and nucleus, except for SSK8, which is detected in cytosol only. Tissue-specific expression patterns suggested that many SSK genes may be involved in tomato fruit development. Furthermore, several SSK genes were found to be responsive to heat stress and salicylic acid treatment. Based on phylogenetic analysis, expression profiles and protein interaction property, we proposed that tomato SSK1 and SSK2 might have similar function to ASK1 and ASK2 in Arabidopsis.

  12. Inflammatory chemokines and their receptors in human visceral leishmaniasis: Gene expression profile in peripheral blood, splenic cellular sources and their impact on trafficking of inflammatory cells.

    PubMed

    Singh, Neetu; Sundar, Shyam

    2017-02-18

    Chemokines play an important role in determining cellular composition at inflammatory sites, and as such, influence disease outcome. In this study, we investigated the expression profile and splenic cellular source of various inflammatory chemokines and their receptors in human visceral leishmaniasis (VL). The expression of chemokines or their receptors was measured at the gene and protein level by employing real time qPCR and a cytometric bead array assay, respectively. In addition, the cellular source of chemokines and their receptors in the spleen was identified employing gene expression analyses in sequentially selected cell subsets. We identified elevated expression of CXCL10, CXCL9, CXCL8, and decreased CCL2 from VL patients. Further, we found reduced expression of the chemokine receptors CXCR1, CXCR2, CXCR3 and CCR2, but increased expression of CCR7 on VL PBMC, compared to endemic healthy controls. Additionally, splenic monocytes were found to be the major source of CXCL10, CXCL9 and CCR2, whereas T cells were the main source of CXCR3 and CCR7. We also report a strong association between plasma IFN-γ and CXCL-10, CXCL-9 levels. Enhanced parasite burden positively correlates with increased expression of CXCL10, CXCL9, IFN-γ and IL-10. Overall our result indicates that VL patients have an elevated inflammatory chemokine milieu which correlated with disease severity. However, expression of their chemokine receptors was significantly impaired, which may have contributed to reduced frequencies of blood monocytes and neutrophils in peripheral blood. In contrast, enhanced expression of CCR7 was associated with increased numbers of activated T cells in circulation. These findings highlight the importance of chemokines for recruitment of various cell populations in VL, and the knowledge gained may help in global understandings of the complex interaction between chemokines and pathological processes, and therefore will contribute towards the design of novel

  13. Analysis of Epstein-Barr virus and cellular gene expression during the early phases of Epstein-Barr virus lytic induction.

    PubMed

    Auburn, Helen; Zuckerman, Mark; Smith, Melvyn

    2016-11-01

    In order to develop novel host/pathogen real-time PCR assays for routine diagnostic use, early gene expression patterns from both Epstein-Barr virus (EBV) and Raji cells were examined after inducing the lytic life cycle using 12-O-tetradecanoyl-13-phorbol ester and sodium butyrate. Real-time PCR identified several highly induced (>90-fold) EBV lytic genes over a 48 h time course during the lytic induction phase. Latent genes were induced at low levels during this phase. The cellular response to lytic viral replication is poorly understood. Whole human genome microarray analysis identified 113 cellular genes regulated twofold or more by EBV, including 63 upregulated and 46 downregulated genes, over a 24 h time course post-induction. The most upregulated gene was CHI3L1, a chitinase-3-like 1 protein (18.1-fold; P<0.0084), and the most downregulated gene was TYMS, a thymidylate synthetase (-7.6-fold). Gene Ontology enrichment analysis using MetaCore software revealed cell cycle (core), cell cycle (role of anaphase-promoting complex) in cell cycle regulation) and lymphatic diseases as the most significantly represented biological network processes, canonical pathways and disease biomarkers, respectively. Chemotaxis, DNA damage and inflammation (IL-4 signalling) together with lymphoproliferative disorders and non-Hodgkin's lymphoma were significantly represented biological processes and disease biomarkers.

  14. Expression of Caveolin-1 reduces cellular responses to TGF-{beta}1 through down-regulating the expression of TGF-{beta} type II receptor gene in NIH3T3 fibroblast cells

    SciTech Connect

    Lee, Eun Kyung; Lee, Youn Sook; Han, In-Oc; Park, Seok Hee . E-mail: parks@skku.edu

    2007-07-27

    Transcriptional repression of Transforming Growth Factor-{beta} type II receptor (T{beta}RII) gene has been proposed to be one of the major mechanisms leading to TGF-{beta} resistance. In this study, we demonstrate that expression of Caveolin-1 (Cav-1) gene in NIH3T3 fibroblast cells down-regulates the expression of T{beta}RII gene in the transcriptional level, eventually resulting in the decreased responses to TGF-{beta}. The reduced expression of T{beta}RII gene by Cav-1 appeared to be due to the changes of the sequence-specific DNA binding proteins to either Positive Regulatory Element 1 (PRE1) or PRE2 of the T{beta}RII promoter. In addition, Cav-1 expression inhibited TGF-{beta}-mediated cellular proliferation and Plasminogen Activator Inhibitor (PAI)-1 gene expression as well as TGF-{beta}-induced luciferase activity. Furthermore, the inhibition of endogeneous Cav-1 by small interfering RNA increased the expression of T{beta}RII gene. These findings strongly suggest that expression of Cav-1 leads to the decreased cellular responsiveness to TGF-{beta} through down-regulating T{beta}RII gene expression.

  15. Gene expression profiling of bovine ovarian follicular and luteal cells provides insight into cellular identities and functions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    After ovulation, somatic cells of the ovarian follicle (theca and granulosa cells) become the small and large luteal cells of the corpus luteum. Aside from known cell type-specific receptors and steroidogenic enzymes, little is known about the differences in the gene expression profiles of these fou...

  16. A comparison of ovarian follicular and luteal cell gene expression profiles provides insight into cellular identities and functions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    After ovulation, somatic cells of the ovarian follicle (theca and granulosa cells) become the small and large luteal cells of the corpus luteum. Aside from known cell type-specific receptors and steroidogenic enzymes, little is known about the differences in the gene expression profiles of these fou...

  17. A stable HeLa cell line that inducibly expresses poliovirus 2A(pro): effects on cellular and viral gene expression.

    PubMed

    Barco, A; Feduchi, E; Carrasco, L

    2000-03-01

    A HeLa cell clone (2A7d) that inducibly expresses the gene for poliovirus protease 2A (2A(pro)) under the control of tetracycline has been obtained. Synthesis of 2A(pro) induces severe morphological changes in 2A7d cells. One day after tetracycline removal, cells round up and a few hours later die. Poliovirus 2A(pro) cleaves both forms of initiation factor eIF4G, causing extensive inhibition of capped-mRNA translation a few hours after protease induction. Methoxysuccinyl-Ala-Ala-Pro-Val-chloromethylketone, a selective inhibitor of 2A(pro), prevents both eIF4G cleavage and inhibition of translation but not cellular death. Expression of 2A(pro) still allows both the replication of poliovirus and the translation of mRNAs containing a picornavirus leader sequence, while vaccinia virus replication is drastically inhibited. Translation of transfected capped mRNA is blocked in 2A7d-On cells, while luciferase synthesis from a mRNA bearing a picornavirus internal ribosome entry site (IRES) sequence is enhanced by the presence of 2A(pro). Moreover, synthesis of 2A(pro) in 2A7d cells complements the translational defect of a poliovirus 2A(pro)-defective variant. These results show that poliovirus 2A(pro) expression mimics some phenotypical characteristics of poliovirus-infected cells, such as cell rounding, inhibition of protein synthesis and enhancement of IRES-driven translation. This cell line constitutes a useful tool to further analyze 2A(pro) functions, to complement poliovirus 2A(pro) mutants, and to test antiviral compounds.

  18. Ursolic Acid Inhibits Na+/K+-ATPase Activity and Prevents TNF-α-Induced Gene Expression by Blocking Amino Acid Transport and Cellular Protein Synthesis

    PubMed Central

    Yokomichi, Tomonobu; Morimoto, Kyoko; Oshima, Nana; Yamada, Yuriko; Fu, Liwei; Taketani, Shigeru; Ando, Masayoshi; Kataoka, Takao

    2011-01-01

    Pro-inflammatory cytokines, such as tumor necrosis factor (TNF)-α, induce the expression of a wide variety of genes, including intercellular adhesion molecule-1 (ICAM-1). Ursolic acid (3β-hydroxy-urs-12-en-28-oic acid) was identified to inhibit the cell-surface ICAM-1 expression induced by pro-inflammatory cytokines in human lung carcinoma A549 cells. Ursolic acid was found to inhibit the TNF-α-induced ICAM-1 protein expression almost completely, whereas the TNF-α-induced ICAM-1 mRNA expression and NF-κB signaling pathway were decreased only partially by ursolic acid. In line with these findings, ursolic acid prevented cellular protein synthesis as well as amino acid uptake, but did not obviously affect nucleoside uptake and the subsequent DNA/RNA syntheses. This inhibitory profile of ursolic acid was similar to that of the Na+/K+-ATPase inhibitor, ouabain, but not the translation inhibitor, cycloheximide. Consistent with this notion, ursolic acid was found to inhibit the catalytic activity of Na+/K+-ATPase. Thus, our present study reveals a novel molecular mechanism in which ursolic acid inhibits Na+/K+-ATPase activity and prevents the TNF-α-induced gene expression by blocking amino acid transport and cellular protein synthesis. PMID:24970122

  19. Effect of passage number on cellular response to DNA-damaging agents: Cell survival and gene expression

    SciTech Connect

    Chang-Liu, C.M.; Woloschak, G.E.

    1997-08-01

    The effect of different passage numbers on plating efficiency, doubling time, cell growth, and radiation sensitivity was assessed in Syrian hamster embryo (SHE) cells. Changes in gene expression after UV or {gamma}-ray irradiation at different passage numbers were also examined. The SHE cells were maintained in culture medium for up to 64 passages. Cells were exposed to {sup 60}Co {gamma} rays or 254-nm UV radiation. Differential display of cDNAs and northern blots were used for the study of gene expression. With increasing passage number, SHE cells demonstrated decreased doubling time, increased plating efficiency, and a decreased yield in the number of cells per plate. Between passages 41 and 48 a crisis period was evident during which time cell growth in high serum was no longer optimal, and serum concentrations were reduced to maintain cell growth. Sensitivity to ionizing radiation was no different between early- and intermediate-passage cells. However, after UV exposure at low passages (passage 3), confluent cells were more sensitive to the killing effects of UV than were log-phase cells. At intermediate passages (passages 43, 48), confluent cells were slightly more radioresistant than were log-phase cells. By passage 64, however, both confluent and log-phase cells showed similar patterns of UV sensitivity. Expression of {gamma}-actin, PCNA, and p53 transcripts did not change following UV exposure. p53 mRNA was induced following {gamma}-ray exposure of the intermediate (passage 45) epithelial cells. The observed differences in radiation sensitivity associated with increasing passage number may be influenced by radiation-induced gene expression. The authors are conducted experiments to identify these genes.

  20. Impaired antioxidant gene expression by pesticide residues and its relation with other cellular biomarkers in Nile Tilapia (Oreochromis niloticus) from Lake Burullus.

    PubMed

    El Nahas, Abeer F; Abdel-Razek, Mohamed A S; Helmy, Nashwa M; Mahmoud, Shawky; Ghazy, Haneen A

    2017-03-01

    Organochlorines and Organophosphorus are the most commonly used pesticides. These pesticides constitute a considerable contaminating threat due to their excessive agricultural usage which in turn contaminates the aquatic system through agricultural drainage. The aim of this study was to evaluate water and tissue residues of both pesticides in O. niloticus obtained from three different sections in Lake Burullus, Egypt. Assessment of relative change in mRNA levels of GST and Vtg (oxidative stress indicator) was done and its relation with other cellular biomarkers including apoptosis, which is assessed by Cellular apoptosis susceptibility transcript level (CAS), comet assay and micronucleus assays (genotoxicity indicator). Pesticide residue levels in water are fluctuating. In fish tissues, most residues were higher than those found in water and were associated with down regulation of hepatic GST gene and Vtg expression. CAS gene involved in apoptosis, its transcript is down regulated in middle and western sections of the lake with higher pesticide residues. Different degrees of DNA damages in O. niloticus' liver cells were demonstrated by comet assay. Significant increase in the micronucleated cells in the three sections of the lake was observed; the western section fish showed the highest number. Persistent exposures of fish to pesticide caused impairment of antioxidant gene expression. This negatively affects apoptosis associated with damaging DNA and chromosome fragments.

  1. Low-temperature atmospheric plasma increases the expression of anti-aging genes of skin cells without causing cellular damages.

    PubMed

    Choi, Jeong-Hae; Lee, Hyun-Wook; Lee, Jae-Koo; Hong, Jin-woo; Kim, Gyoo-cheon

    2013-03-01

    Efforts to employ various types of plasma in the field of skin care have increased consistently because it can regulate many biochemical reactions that are normally unaffected by light-based therapy. One method for skin rejuvenation adopted a high-temperature plasma generator to remove skin epithelial cells. In this case, the catalyzing effects of the plasma were rarely used due to the high temperature. Hence, the benefits of the plasma were not magnified. Recently, many types of low-temperature plasma devices have been developed for medical applications but their detailed functions and working mechanisms are unclear. The present study examined the effect of low-temperature microwave plasma on skin cells. Treatment with low-temperature plasma increased the expression of anti-aging genes in skin cells, including collagen, fibronectin and vascular endothelial growth factor. Furthermore, the plasma treatment did not cause cell death, but only induced slight cell growth arrest at the G2 phase. Although the cells treated with low-temperature plasma showed moderate growth arrest, there were no signs of thermal or genetic damage of skin cells. Overall, this low-temperature microwave plasma device induces the expressions of some anti-aging-related genes in skin cells without causing damage.

  2. Mitigating effects of L-selenomethionine on low-dose iron ion radiation-induced changes in gene expression associated with cellular stress.

    PubMed

    Nuth, Manunya; Kennedy, Ann R

    2013-07-01

    Ionizing radiation associated with highly energetic and charged heavy (HZE) particles poses a danger to astronauts during space travel. The aim of the present study was to evaluate the patterns of gene expression associated with cellular exposure to low-dose iron ion irradiation, in the presence and absence of L-selenomethionine (SeM). Human thyroid epithelial cells (HTori-3) were exposed to low-dose iron ion (1 GeV/n) irradiation at 10 or 20 cGy with or without SeM pretreatment. The cells were harvested 6 and 16 h post-irradiation and analyzed by the Affymetrix U133Av2 gene chip arrays. Genes exhibiting a 1.5-fold expression cut-off and 5% false discovery rate (FDR) were considered statistically significant and subsequently analyzed using the Database for Annotation, Visualization and Integrated Discovery (DAVID) for pathway analysis. Representative genes were further validated by real-time RT-PCR. Even at low doses of radiation from iron ions, global genome profiling of the irradiated cells revealed the upregulation of genes associated with the activation of stress-related signaling pathways (ubiquitin-mediated proteolysis, p53 signaling, cell cycle and apoptosis), which occurred in a dose-dependent manner. A 24-h pretreatment with SeM was shown to reduce the radiation effects by mitigating stress-related signaling pathways and downregulating certain genes associated with cell adhesion. The mechanism by which SeM prevents radiation-induced transformation in vitro may involve the suppression of the expression of genes associated with stress-related signaling and certain cell adhesion events.

  3. Complementation of non-tumorigenicity of HPV18-positive cervical carcinoma cells involves differential mRNA expression of cellular genes including potential tumor suppressor genes on chromosome 11q13.

    PubMed

    Kehrmann, Angela; Truong, Ha; Repenning, Antje; Boger, Regina; Klein-Hitpass, Ludger; Pascheberg, Ulrich; Beckmann, Alf; Opalka, Bertram; Kleine-Lowinski, Kerstin

    2013-01-01

    The fusion between human tumorigenic cells and normal human diploid fibroblasts results in non-tumorigenic hybrid cells, suggesting a dominant role for tumor suppressor genes in the generated hybrid cells. After long-term cultivation in vitro, tumorigenic segregants may arise. The loss of tumor suppressor genes on chromosome 11q13 has been postulated to be involved in the induction of the tumorigenic phenotype of human papillomavirus (HPV)18-positive cervical carcinoma cells and their derived tumorigenic hybrid cells after subcutaneous injection in immunocompromised mice. The aim of this study was the identification of novel cellular genes that may contribute to the suppression of the tumorigenic phenotype of non-tumorigenic hybrid cells in vivo. We used cDNA microarray technology to identify differentially expressed cellular genes in tumorigenic HPV18-positive hybrid and parental HeLa cells compared to non-tumorigenic HPV18-positive hybrid cells. We detected several as yet unknown cellular genes that play a role in cell differentiation, cell cycle progression, cell-cell communication, metastasis formation, angiogenesis, antigen presentation, and immune response. Apart from the known differentially expressed genes on 11q13 (e.g., phosphofurin acidic cluster sorting protein 1 (PACS1) and FOS ligand 1 (FOSL1 or Fra-1)), we detected novel differentially expressed cellular genes located within the tumor suppressor gene region (e.g., EGF-containing fibulin-like extracellular matrix protein 2 (EFEMP2) and leucine rich repeat containing 32 (LRRC32) (also known as glycoprotein-A repetitions predominant (GARP)) that may have potential tumor suppressor functions in this model system of non-tumorigenic and tumorigenic HeLa x fibroblast hybrid cells.

  4. Exercise Decreases Lipogenic Gene Expression in Adipose Tissue and Alters Adipocyte Cellularity during Weight Regain After Weight Loss

    PubMed Central

    Giles, Erin D.; Steig, Amy J.; Jackman, Matthew R.; Higgins, Janine A.; Johnson, Ginger C.; Lindstrom, Rachel C.; MacLean, Paul S.

    2016-01-01

    Exercise is a potent strategy to facilitate long-term weight maintenance. In addition to increasing energy expenditure and reducing appetite, exercise also favors the oxidation of dietary fat, which likely helps prevent weight re-gain. It is unclear whether this exercise-induced metabolic shift is due to changes in energy balance, or whether exercise imparts additional adaptations in the periphery that limit the storage and favor the oxidation of dietary fat. To answer this question, adipose tissue lipid metabolism and related gene expression were studied in obese rats following weight loss and during the first day of relapse to obesity. Mature, obese rats were weight-reduced for 2 weeks with or without daily treadmill exercise (EX). Rats were weight maintained for 6 weeks, followed by relapse on: (a) ad libitum low fat diet (LFD), (b) ad libitum LFD plus EX, or (c) a provision of LFD to match the positive energy imbalance of exercised, relapsing animals. 24 h retention of dietary- and de novo-derived fat were assessed directly using 14C palmitate/oleate and 3H20, respectively. Exercise decreased the size, but increased the number of adipocytes in both retroperitoneal (RP) and subcutaneous (SC) adipose depots, and prevented the relapse-induced increase in adipocyte size. Further, exercise decreased the expression of genes involved in lipid uptake (CD36 and LPL), de novo lipogenesis (FAS, ACC1), and triacylglycerol synthesis (MGAT and DGAT) in RP adipose during relapse following weight loss. This was consistent with the metabolic data, whereby exercise reduced retention of de novo-derived fat even when controlling for the positive energy imbalance. The decreased trafficking of dietary fat to adipose tissue with exercise was explained by reduced energy intake which attenuated energy imbalance during refeeding. Despite having decreased expression of lipogenic genes, the net retention of de novo-derived lipid was higher in both the RP and SC adipose of exercising

  5. Differential cellular gene expression in duck trachea infected with a highly or low pathogenic H5N1 avian influenza virus

    PubMed Central

    2013-01-01

    Background Avian influenza A (AI) viruses of subtypes H5 can cause serious disease outbreaks in poultry including panzootic due to H5N1 highly pathogenic (HP) viruses. These viruses are a threat not only for animal health but also public health due to their zoonotic potential. The domestic duck plays a major role in the epidemiological cycle of influenza virus subtypes H5 but little is known concerning host/pathogen interactions during influenza infection in duck species. In this study, a subtracted library from duck trachea (a primary site of influenza virus infection) was constructed to analyse and compare the host response after a highly or low pathogenic (LP) H5N1-infection. Results Here, we show that more than 200 different genes were differentially expressed in infected duck trachea to a significant degree. In addition, significant differentially expressed genes between LPAI- and HPAI-infected tracheas were observed. Gene ontology annotation was used and specific signalling pathways were identified. These pathways were different for LPAI and HPAI-infected tracheas, except for the CXCR4 signalling pathway which is implicated in immune response. A different modulation of genes in the CXCR4 signalling pathway and TRIM33 was induced in duck tracheas infected with a HPAI- or a LPAI-H5N1. Conclusion First, this study indicates that Suppressive Subtractive Hybridization (SSH) is an alternative approach to gain insights into the pathogenesis of influenza infection in ducks. Secondly, the results indicate that cellular gene expression in the duck trachea was differently modulated after infection with a LPAI-H5N1 or after infection with a HPAI-H5N1 virus. Such difference found in infected trachea, a primary infection site, could precede continuation of infection and could explain appearance of respiratory symptoms or not. PMID:24015922

  6. Identification of cellular genes induced in human cells after activation of the OAS/RNaseL pathway by vaccinia virus recombinants expressing these antiviral enzymes.

    PubMed

    Domingo-Gil, Elena; González, José Manuel; Esteban, Mariano

    2010-03-01

    Interferon (IFN) type I induces the expression of antiviral proteins such as 2',5'-oligoadenylate synthetases (OAS). The enzyme OAS is activated by dsRNA to produce 5'-phosphorylated, 2-5-linked oligoadenylates (2-5A) that activate RNaseL which, in turn, triggers RNA breakdown, leading to multiple biological functions. Although RNaseL is required for IFN antiviral function, there are many aspects of the molecular mechanisms that remain obscure. Here, we have used microarray analyses from human HeLa cells infected with vaccinia virus (VACV) recombinants expressing OAS-RNaseL enzymes (referred as 2-5A system) with the aim to identify host genes that are up- or down-regulated in the course of infection by the activation of this antiviral pathway. We found that activation of the 2-5A system from VACV recombinants produces a remarkable stimulation of transcription for genes that regulate many cellular processes, like those that promote cell growth arrest, GADD45B and KCTD11, apoptosis as CUL2, PDCD6, and TNFAIP8L2, IFN-stimulated genes as IFI6, and related to tumor suppression as PLA2G2A. The 2-5A system activation produces down-regulation of transcription of some genes that promote cell growth as RUNX2 and ESR2 and of genes in charge to maintain mitochondria homeostasis as MIPEP and COX5A. These results reveal new genes induced in response to the activation of the 2-5A system with roles in apoptosis, translational control, cell growth arrest, and tumor suppression.

  7. Evaluating long-term cellular effects of the arsenic species thio-DMA(V): qPCR-based gene expression as screening tool.

    PubMed

    Ebert, Franziska; Thomann, Marlies; Witt, Barbara; Müller, Sandra M; Meyer, Sören; Weber, Till; Christmann, Markus; Schwerdtle, Tanja

    2016-09-01

    Thio-dimethylarsinic acid (thio-DMA(V)) is a human urinary metabolite of the class 1 human carcinogen inorganic arsenic as well as of arsenosugars. Thio-DMA(V) exerts strong cellular toxicity, whereas its toxic modes of action are not fully understood. For the first time, this study characterises the impact of a long-term (21days) in vitro incubation of thio-DMA(V) on the expression of selected genes related to cell death, stress response, epigenetics and DNA repair. The observed upregulation of DNMT1 might be a cellular compensation to counterregulate the in a very recent study observed massive global DNA hypomethylation after chronic thio-DMA(V) incubation. Moreover, our data suggest that chronic exposure towards subcytotoxic, pico- to nanomolar concentrations of thio-DMA(V) causes a stress response in human urothelial cells. The upregulation of genes encoding for proteins of DNA repair (Apex1, Lig1, XRCC1, DDB2, XPG, ATR) as well as damage response (GADD45A, GADD45G, Trp53) indicate a potential genotoxic risk emanating from thio-DMA(V) after long-term incubation.

  8. Cellular Effect of High Doses of Silica-Coated Quantum Dot Profiled with High Throughput Gene Expression Analysis and High Content Cellomics Measurements

    PubMed Central

    Zhang, Tingting; Stilwell, Jackie L.; Gerion, Daniele; Ding, Lianghao; Elboudwarej, Omeed; Cooke, Patrick A.; Gray, Joe W.; Alivisatos, A. Paul; Chen, Fanqing Frank

    2009-01-01

    Quantum dots (Qdots) are now used extensively for labeling in biomedical research, and this use is predicted to grow because of their many advantages over alternative labeling methods. Uncoated Qdots made of core/shell CdSe/ZnS are toxic to cells because of the release of Cd2+ ions into the cellular environment. This problem has been partially overcome by coating Qdots with polymers, poly(ethylene glycol) (PEG), or other inert molecules. The most promising coating to date, for reducing toxicity, appears to be PEG. When PEG-coated silanized Qdots (PEG-silane-Qdots) are used to treat cells, toxicity is not observed, even at dosages above 10–20 nM, a concentration inducing death when cells are treated with polymer or mercaptoacid coated Qdots. Because of the importance of Qdots in current and future biomedical and clinical applications, we believe it is essential to more completely understand and verify this negative global response from cells treated with PEG-silane-Qdots. Consequently, we examined the molecular and cellular response of cells treated with two different dosages of PEG-silane-Qdots. Human fibroblasts were exposed to 8 and 80 nM of these Qdots, and both phenotypic as well as whole genome expression measurements were made. PEG-silane-Qdots did not induce any statistically significant cell cycle changes and minimal apoptosis/necrosis in lung fibroblasts (IMR-90) as measured by high content image analysis, regardless of the treatment dosage. A slight increase in apoptosis/necrosis was observed in treated human skin fibroblasts (HSF-42) at both the low and the high dosages. We performed genome-wide expression array analysis of HSF-42 exposed to doses 8 and 80 nM to link the global cell response to a molecular and genetic phenotype. We used a gene array containing ~22,000 total probe sets, containing 18,400 probe sets from known genes. Only ~50 genes (~0.2% of all the genes tested) exhibited a statistically significant change in expression level of greater

  9. In vitro mimicking of estrous cycle stages in porcine oviduct epithelium cells: estradiol and progesterone regulate differentiation, gene expression, and cellular function.

    PubMed

    Chen, Shuai; Einspanier, Ralf; Schoen, Jennifer

    2013-09-01

    Throughout the estrous cycle the oviduct epithelium undergoes dramatic morphological and functional changes. To elucidate cyclic cellular events and associated regulation mechanisms of 17beta estradiol (E2) and progesterone (P4), we mimicked estrous cycle stages in vitro using a culture system of primary porcine oviduct epithelium cells (POEC). Cells were polarized in an air/liquid interface and then treated with E2 and P4 for physiological time periods: In experiment 1, high concentration of P4 with low concentration of E2 for 10 days resembled diestrus; in experiment 2, following the previous diestrus, sequential high E2 with low P4 for 2.5 days represented estrus. Histomorphometry and electron microscopy showed cyclic changes in cellular height, cell population, and cilia density under the influence of hormone stimulation. Transepithelial electrical resistance was high in simulated diestrus but reduced in estrus. Thus, E2 and P4 affect cellular polarity, transformation of ciliated and secretory cells, as well as electrical conductivity of oviduct epithelium. Simulation of diestrus led to significant decrease in expression of hormone receptors (PGR and ESR1) and other epithelial markers (MUC16, OVGP1, and HSP90B1), while sequential simulated estrus caused an increase in these markers. The hormonal regulation of some marker genes was clearly time-dependent. Furthermore, POEC showed increased sperm-binding capacity in simulated estrus. In this study, we also present a novel approach based on the AndroVision software, which can be routinely utilized as a parameter for ciliary activity, and for the first time, we showed fluid movement patterns along the epithelium lining in vitro.

  10. Metabolomic changes during cellular transformation monitored by metabolite-metabolite correlation analysis and correlated with gene expression.

    PubMed

    Madhu, Basetti; Narita, Masako; Jauhiainen, Alexandra; Menon, Suraj; Stubbs, Marion; Tavaré, Simon; Narita, Masashi; Griffiths, John R

    To investigate metabolic changes during cellular transformation, we used a (1)H NMR based metabolite-metabolite correlation analysis (MMCA) method, which permits analysis of homeostatic mechanisms in cells at the steady state, in an inducible cell transformation model. Transcriptomic data were used to further explain the results. Transformed cells showed many more metabolite-metabolite correlations than control cells. Some had intuitively plausible explanations: a shift from glycolysis to amino acid oxidation after transformation was accompanied by a strongly positive correlation between glucose and glutamine and a strongly negative one between lactate and glutamate; there were also many correlations between the branched chain amino acids and the aromatic amino acids. Others remain puzzling: after transformation strong positive correlations developed between choline and a group of five amino acids, whereas the same amino acids showed negative correlations with phosphocholine, a membrane phospholipid precursor. MMCA in conjunction with transcriptome analysis has opened a new window into the metabolome.

  11. Inhibition of MDR1 gene expression and enhancing cellular uptake for effective colon cancer treatment using dual-surface–functionalized nanoparticles

    PubMed Central

    Xiao, Bo; Zhang, Mingzhen; Viennois, Emilie; Zhang, Yuchen; Wei, Na; Baker, Mark T.; Jung, Yunjin; Merlin, Didier

    2015-01-01

    Nanomedicine options for colon cancer therapy have been limited by the lack of suitable carriers capable of delivering sufficient drug into tumors to cause lethal toxicity. To circumvent this limitation, we fabricated a camptothecin (CPT)-loaded poly(lactic-co-glycolic acid) nanoparticle (NP) with dual-surface functionalization—Pluronic F127 and chitosan—for inhibiting multi-drug resistant gene 1 (MDR1) expression and enhancing tumor uptake. The resultant spherical NPs-P/C had a desirable particle size (~268 nm), slightly positive zeta-potential, and the ability to efficiently down-regulate the expression of MDR1. In vitro cytotoxicity tests revealed that the 24 and 48 h IC50 values of NPs-P/C1 were 2.03 and 0.67 µM, respectively, which were much lower than those for free CPT and other NPs. Interestingly, NPs-P/C1 showed the highest cellular uptake efficiency (approximately 85.5%) among the different drug formulations. Most importantly, treatment of colon tumor-bearing mice with various drug formulations confirmed that the introduction of Pluronic F127 and chitosan to the NP surface significantly enhanced the therapeutic efficacy of CPT, induced tumor cell apoptosis, and reduced systemic toxicity. Collectively, these findings suggest that our one-step–fabricated, dual-surface–functionalized NPs may hold promise as a readily scalable and effective drug carrier with clinical potential in colon cancer therapy. PMID:25701040

  12. The dopamine D2 receptor gene in lamprey, its expression in the striatum and cellular effects of D2 receptor activation.

    PubMed

    Robertson, Brita; Huerta-Ocampo, Icnelia; Ericsson, Jesper; Stephenson-Jones, Marcus; Pérez-Fernández, Juan; Bolam, J Paul; Diaz-Heijtz, Rochellys; Grillner, Sten

    2012-01-01

    All basal ganglia subnuclei have recently been identified in lampreys, the phylogenetically oldest group of vertebrates. Furthermore, the interconnectivity of these nuclei is similar to mammals and tyrosine hydroxylase-positive (dopaminergic) fibers have been detected within the input layer, the striatum. Striatal processing is critically dependent on the interplay with the dopamine system, and we explore here whether D2 receptors are expressed in the lamprey striatum and their potential role. We have identified a cDNA encoding the dopamine D2 receptor from the lamprey brain and the deduced protein sequence showed close phylogenetic relationship with other vertebrate D2 receptors, and an almost 100% identity within the transmembrane domains containing the amino acids essential for dopamine binding. There was a strong and distinct expression of D2 receptor mRNA in a subpopulation of striatal neurons, and in the same region tyrosine hydroxylase-immunoreactive synaptic terminals were identified at the ultrastructural level. The synaptic incidence of tyrosine hydroxylase-immunoreactive boutons was highest in a region ventrolateral to the compact layer of striatal neurons, a region where most striatal dendrites arborise. Application of a D2 receptor agonist modulates striatal neurons by causing a reduced spike discharge and a diminished post-inhibitory rebound. We conclude that the D2 receptor gene had already evolved in the earliest group of vertebrates, cyclostomes, when they diverged from the main vertebrate line of evolution (560 mya), and that it is expressed in striatum where it exerts similar cellular effects to that in other vertebrates. These results together with our previous published data (Stephenson-Jones et al. 2011, 2012) further emphasize the high degree of conservation of the basal ganglia, also with regard to the indirect loop, and its role as a basic mechanism for action selection in all vertebrates.

  13. Expression of Cellular Oncogenes in Human Malignancies

    NASA Astrophysics Data System (ADS)

    Slamon, Dennis J.; Dekernion, Jean B.; Verma, Inder M.; Cline, Martin J.

    1984-04-01

    Cellular oncogenes have been implicated in the induction of malignant transformation in some model systems in vitro and may be related to malignancies in vivo in some vertebrate species. This article describes a study of the expression of 15 cellular oncogenes in fresh human tumors from 54 patients, representing 20 different tumor types. More than one cellular oncogene was transcriptionally active in all of the tumors examined. In 14 patients it was possible to study normal and malignant tissue from the same organ. In many of these patients, the transcriptional activity of certain oncogenes was greater in the malignant than the normal tissue. The cellular fes (feline sarcoma) oncogene, not previously known to be transcribed in mammalian tissue, was found to be active in lung and hematopoietic malignancies.

  14. Gene Expression Reaction Norms Unravel the Molecular and Cellular Processes Underpinning the Plastic Phenotypes of Alternanthera Philoxeroides in Contrasting Hydrological Conditions

    PubMed Central

    Gao, Lexuan; Geng, Yupeng; Yang, Hongxing; Hu, Yonghong; Yang, Ji

    2015-01-01

    Alternanthera philoxeroides is an amphibious invasive weed that can colonize both aquatic and terrestrial habitats. Individuals growing in different habitats exhibit extensive phenotypic variation but little genetic differentiation. Little is known about the molecular basis underlying environment-induced phenotypic changes. Variation in transcript abundance in A. philoxeroides was characterized throughout the time-courses of pond and upland treatments using RNA-Sequencing. Seven thousand eight hundred and five genes demonstrated variable expression in response to different treatments, forming 11 transcriptionally coordinated gene groups. Functional enrichment analysis of plastically expressed genes revealed pathway changes in hormone-mediated signaling, osmotic adjustment, cell wall remodeling, and programmed cell death, providing a mechanistic understanding of the biological processes underlying the phenotypic changes in A. philoxeroides. Both transcriptional modulation of environmentally sensitive loci and environmentally dependent control of regulatory loci influenced the plastic responses to the environment. Phenotypic responses and gene expression patterns to contrasting hydrological conditions were compared between A. philoxeroides and its alien congener Alternanthera pungens. The terricolous A. pungens displayed limited phenotypic plasticity to different treatments. It was postulated based on gene expression comparison that the interspecific variation in plasticity between A. philoxeroides and A. pungens was not due to environmentally-mediated changes in hormone levels but to variations in the type and relative abundance of different signal transducers and receptors expressed in the target tissue. PMID:26617628

  15. Mechanoregulation of gene expression in fibroblasts

    PubMed Central

    Wang, James H.-C.; Thampatty, Bhavani P.; Lin, Jeen-Shang; Im, Hee-Jeong

    2010-01-01

    Mechanical loads placed on connective tissues alter gene expression in fibroblasts through mechanotransduction mechanisms by which cells convert mechanical signals into cellular biological events, such as gene expression of extracellular matrix components (e.g., collagen). This mechanical regulation of ECM gene expression affords maintenance of connective tissue homeostasis. However, mechanical loads can also interfere with homeostatic cellular gene expression and consequently cause the pathogenesis of connective tissue diseases such as tendinopathy and osteoarthritis. Therefore, the regulation of gene expression by mechanical loads is closely related to connective tissue physiology and pathology. This article reviews the effects of various mechanical loading conditions on gene regulation in fibroblasts and discusses several mechanotransduction mechanisms. Future research directions in mechanoregulation of gene expression are also suggested. PMID:17331678

  16. Different N-terminal isoforms of Oct-1 control expression of distinct sets of genes and their high levels in Namalwa Burkitt's lymphoma cells affect a wide range of cellular processes

    PubMed Central

    Pankratova, Elizaveta V.; Stepchenko, Alexander G.; Portseva, Tatiana; Mogila, Vladic A.; Georgieva, Sofia G.

    2016-01-01

    Oct-1 transcription factor has various functions in gene regulation. Its expression level is increased in several types of cancer and is associated with poor survival prognosis. Here we identified distinct Oct-1 protein isoforms in human cells and compared gene expression patterns and functions for Oct-1A, Oct-1L, and Oct-1X isoforms that differ by their N-terminal sequences. The longest isoform, Oct-1A, is abundantly expressed and is the main Oct-1 isoform in most of human tissues. The Oct-1L and the weakly expressed Oct-1X regulate the majority of Oct-1A targets as well as additional sets of genes. Oct-1X controls genes involved in DNA replication, DNA repair, RNA processing, and cellular response to stress. The high level of Oct-1 isoforms upregulates genes related to cell cycle progression and activates proliferation both in Namalwa Burkitt's lymphoma cells and primary human fibroblasts. It downregulates expression of genes related to antigen processing and presentation, cytokine-cytokine receptor interaction, oxidative metabolism, and cell adhesion, thus facilitating pro-oncogenic processes. PMID:27407111

  17. Short-term administration of rhGH increases markers of cellular proliferation, but not milk protein gene expression in normal lactating women.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Growth hormone is one of few pharmacologic agents known to augment milk production in humans. We hypothesized that recombinant human GH (rhGH) increases the expression of cell proliferation and milk protein synthesis genes. Sequential milk and blood samples collected over four days were obtained fro...

  18. Direct interaction of cellular hnRNP-F and NS1 of influenza A virus accelerates viral replication by modulation of viral transcriptional activity and host gene expression

    SciTech Connect

    Lee, Jun Han; Kim, Sung-Hak; Pascua, Philippe Noriel Q.; Song, Min-Suk; Baek, Yun Hee; Jin, Xun; Choi, Joong-Kook; Kim, Chul-Joong; Kim, Hyunggee; Choi, Young Ki

    2010-02-05

    To investigate novel NS1-interacting proteins, we conducted a yeast two-hybrid analysis, followed by co-immunoprecipitation assays. We identified heterogeneous nuclear ribonucleoprotein F (hnRNP-F) as a cellular protein interacting with NS1 during influenza A virus infection. Co-precipitation assays suggest that interaction between hnRNP-F and NS1 is a common and direct event among human or avian influenza viruses. NS1 and hnRNP-F co-localize in the nucleus of host cells, and the RNA-binding domain of NS1 directly interacts with the GY-rich region of hnRNP-F determined by GST pull-down assays with truncated proteins. Importantly, hnRNP-F expression levels in host cells indicate regulatory role on virus replication. hnRNP-F depletion by small interfering RNA (siRNA) shows 10- to 100-fold increases in virus titers corresponding to enhanced viral RNA polymerase activity. Our results delineate novel mechanism of action by which NS1 accelerates influenza virus replication by modulating normal cellular mRNA processes through direct interaction with cellular hnRNP-F protein.

  19. Study on connexin gene and protein expression and cellular distribution in relation to real-time proliferation of porcine granulosa cells.

    PubMed

    Kempisty, B; Ziółkowska, A; Ciesiółka, S; Piotrowska, H; Antosik, P; Bukowska, D; Nowicki, M; Brüssow, K P; Zabel, M

    2014-01-01

    Granulosa cells (GCs) play an important role during follicle growth and development in preovulatory stage. Moreover, the proteins such as connexins are responsible for formation of protein channel between follicular-cumulus cells and oocyte. This study was aimed to investigate the role of connexin expression in porcine GCs in relation to their cellular distribution and real-time cell proliferation. In the present study, porcine GCs were isolated from the follicles of puberal gilts and then cultured in a real-time cellular analyzer (RTCA) system for 168 h. The expression levels of connexins (Cxs) Cx36, Cx37, Cx40 and Cx43 mRNA were measured by RQ-PCR analysis, and differences in the expression and distribution of Cx30, Cx31, Cx37, Cx43 and Cx45 proteins were analyzed by confocal microscopic visualization. We found higher level of Cx36, Cx37, and Cx43 mRNA expression in GCs at recovery (at 0 h of in vitro culture, IVC) compared to all analyzed time periods of IVC (24, 48, 72, 96, 120, 144 and 168 h; P<0.001). On the other hand, the expression level of Cx40 transcripts was higher after 24 h of IVC compared to 0 h and the other times of IVC (P<0.001). Similarly to mRNAs, the expression levels of Cx31, Cx37 and Cx45 proteins were higher before (0 h) compared to after 168 h of IVC. The expression of Cx30 and Cx43, however, did not vary between the groups. In all, the proteins were distributed throughout the cell membrane rather than in the cytoplasm both before and after IVC. After 24 h of IVC, we observed a significant increase in the proliferation of GCs (log phase). We found differences in the proliferation index between 72-96 and 96- 140 h within the same population of GCs. In conclusion, the decrease in the expression of Cx mRNAs and proteins following IVC could be associated with a breakdown in gap-junction connections (GJCs), and leads to the decreased of their activity, which may be a reason of non-functional existence of connexon in follicular granulosa cells

  20. Bacterial Cellular Engineering by Genome Editing and Gene Silencing

    PubMed Central

    Nakashima, Nobutaka; Miyazaki, Kentaro

    2014-01-01

    Genome editing is an important technology for bacterial cellular engineering, which is commonly conducted by homologous recombination-based procedures, including gene knockout (disruption), knock-in (insertion), and allelic exchange. In addition, some new recombination-independent approaches have emerged that utilize catalytic RNAs, artificial nucleases, nucleic acid analogs, and peptide nucleic acids. Apart from these methods, which directly modify the genomic structure, an alternative approach is to conditionally modify the gene expression profile at the posttranscriptional level without altering the genomes. This is performed by expressing antisense RNAs to knock down (silence) target mRNAs in vivo. This review describes the features and recent advances on methods used in genomic engineering and silencing technologies that are advantageously used for bacterial cellular engineering. PMID:24552876

  1. Cytotoxicity and expression of genes involved in the cellular stress response and apoptosis in mammalian fibroblast exposed to cotton cellulose nanofibers

    NASA Astrophysics Data System (ADS)

    Pereira, M. M.; Raposo, N. R. B.; Brayner, R.; Teixeira, E. M.; Oliveira, V.; Quintão, C. C. R.; Camargo, L. S. A.; Mattoso, L. H. C.; Brandão, H. M.

    2013-02-01

    Cellulose nanofibers (CNF) have mechanical properties that make them very attractive for applications in the construction of polymeric matrices, drug delivery and tissue engineering. However, little is known about their impact on mammalian cells. The objective of this study was to evaluate the cytotoxicity of CNF and their effect on gene expression of fibroblasts cultured in vitro. The morphology of CNF was analyzed by transmission electron microscopy and the surface charge by Zeta potential. Cell viability was analyzed by flow cytometry assay and gene expression of biomarkers focused on cell stress response such as Heat shock protein 70.1 (HSP70.1) and Peroxiredoxin 1 (PRDX1) and apoptosis as B-cell leukemia (BCL-2) and BCL-2 associated X protein (BAX) by RT-PCR assay. Low concentrations of CNF (0.02-100 μg ml-1) did not cause cell death; however, at concentrations above 200 μg ml-1, the nanofibers significantly decreased cell viability (86.41 ± 5.37%). The exposure to high concentrations of CNF (2000 and 5000 μg ml-1) resulted in increased HSP70.1, PRDX1 and BAX gene expression. The current study concludes that, under the conditions tested, high concentrations (2000 and 5000 μg ml-1) of CNF cause decreased cell viability and affect the expression of stress- and apoptosis-associated molecular markers.

  2. Hepatitis C virus dynamics and cellular gene expression in uPA-SCID chimeric mice with humanized livers during intravenous silibinin monotherapy

    SciTech Connect

    DebRoy, Swati; Hiraga, Nobuhiko; Imamura, Michio; Hayes, C. Nelson; Akamatsu, Sakura; Canini, Laetitia; Perelson, Alan S.; Pohl, Ralf T.; Persiani, Stefano; Uprichard, Susan L.; Tateno, Chise; Dahari, Harel; Chayama, Kazuaki

    2016-06-08

    Legalon SIL (SIL) is a chemically hydrophilized version of silibinin, an extract of milk thistle (Silybum marianum) seeds that has exhibited hepatoprotective and antiviral effectiveness against hepatitis C virus (HCV) in patients leading to viral clearance in combination with ribavirin. In this paper, to elucidate the incompletely understood mode of action of SIL against HCV, mathematical modelling of HCV kinetics and human hepatocyte gene expression studies were performed in uPA-SCID-chimeric mice with humanized livers. Chronically HCV-infected mice (n = 15) were treated for 14 days with daily intravenous SIL at 469, 265 or 61.5 mg/kg. Serum HCV and human albumin (hAlb) were measured frequently, and liver HCV RNA was analysed at days 3 and 14. Microarray analysis of human hepatocyte gene expression was performed at days 0, 3 and 14 of treatment. While hAlb remained constant, a biphasic viral decline in serum was observed consisting of a rapid 1st phase followed by a second slower phase (or plateau with the two lower SIL dosings). SIL effectiveness in blocking viral production was similar among dosing groups (median ε = 77%). However, the rate of HCV-infected hepatocyte decline, δ, was dose-dependent. Intracellular HCV RNA levels correlated (r = 0.66, P = 0.01) with serum HCV RNA. Pathway analysis revealed increased anti-inflammatory and antiproliferative gene expression in human hepatocytes in SIL-treated mice. Finally, the results suggest that SIL could lead to a continuous second-phase viral decline, that is potentially viral clearance, in the absence of adaptive immune response along with increased anti-inflammatory and antiproliferative gene expression in human hepatocytes.

  3. Hepatitis C virus dynamics and cellular gene expression in uPA-SCID chimeric mice with humanized livers during intravenous silibinin monotherapy

    DOE PAGES

    DebRoy, Swati; Hiraga, Nobuhiko; Imamura, Michio; ...

    2016-06-08

    Legalon SIL (SIL) is a chemically hydrophilized version of silibinin, an extract of milk thistle (Silybum marianum) seeds that has exhibited hepatoprotective and antiviral effectiveness against hepatitis C virus (HCV) in patients leading to viral clearance in combination with ribavirin. In this paper, to elucidate the incompletely understood mode of action of SIL against HCV, mathematical modelling of HCV kinetics and human hepatocyte gene expression studies were performed in uPA-SCID-chimeric mice with humanized livers. Chronically HCV-infected mice (n = 15) were treated for 14 days with daily intravenous SIL at 469, 265 or 61.5 mg/kg. Serum HCV and human albuminmore » (hAlb) were measured frequently, and liver HCV RNA was analysed at days 3 and 14. Microarray analysis of human hepatocyte gene expression was performed at days 0, 3 and 14 of treatment. While hAlb remained constant, a biphasic viral decline in serum was observed consisting of a rapid 1st phase followed by a second slower phase (or plateau with the two lower SIL dosings). SIL effectiveness in blocking viral production was similar among dosing groups (median ε = 77%). However, the rate of HCV-infected hepatocyte decline, δ, was dose-dependent. Intracellular HCV RNA levels correlated (r = 0.66, P = 0.01) with serum HCV RNA. Pathway analysis revealed increased anti-inflammatory and antiproliferative gene expression in human hepatocytes in SIL-treated mice. Finally, the results suggest that SIL could lead to a continuous second-phase viral decline, that is potentially viral clearance, in the absence of adaptive immune response along with increased anti-inflammatory and antiproliferative gene expression in human hepatocytes.« less

  4. Transition from somatic embryo to friable embryogenic callus in cassava: dynamic changes in cellular structure, physiological status, and gene expression profiles

    PubMed Central

    Ma, Qiuxiang; Zhou, Wenzhi; Zhang, Peng

    2015-01-01

    Friable embryogenic callus (FEC) is considered as the most suitable material for efficient genetic transformation of cassava. Heavy genotype dependence of FEC induction and amenability to somaclonal variation limits the production and maintenance of reliable FEC. Identifying key elements involved in biological processes from somatic embryos (SEs) to FEC at different stages provides critical insights for FEC improvement. Cytological observation showed a dramatic change of subcellular structures among SEs, fresh FEC (FFEC), and old FEC (OFEC). Decrease of sucrose and increase of fructose and glucose were detected in OFEC. A total of 6871 differentially expressed genes (DEGs) were identified from SEs, FFEC, and OFEC by RNA-seq. Analysis of the DEGs showed that FEC induction was accompanied by the process of dedifferentiation, whereas the epigenetics modification occurred during the continuous subculturing process. The cell structure was reconstructed, mainly including the GO terms of “cell periphery” and “external encapsulating structure”; in parallel, the internal mechanisms changed correspondingly, including the biological process of glycolysis and metabolisms of alanine, aspartate, and glutamate. The significant reduction of genomic DNA methylation in OFEC indicated altered gene expression via chromatin modification. These results indicate that the induction and long-term subculture of FEC is a complicated biological process involving changes of genome modification, gene expression, and subcellular reconstruction. The findings will be useful for improving FEC induction and maintenance from farmer-preferred cassava cultivars recalcitrant to genetic transformation, hence improving cassava through genetic engineering. PMID:26500668

  5. Characterization of Epstein-Barr virus (EBV) BZLF1 gene promoter variants and comparison of cellular gene expression profiles in Japanese patients with infectious mononucleosis, chronic active EBV infection, and EBV-associated hemophagocytic lymphohistiocytosis.

    PubMed

    Imajoh, Masayuki; Hashida, Yumiko; Murakami, Masanao; Maeda, Akihiko; Sato, Tetsuya; Fujieda, Mikiya; Wakiguchi, Hiroshi; Daibata, Masanori

    2012-06-01

    Epstein-Barr virus (EBV) genotypes can be distinguished based on gene sequence differences in EBV nuclear antigens 2, 3A, 3B, and 3C, and the BZLF1 promoter zone (Zp). EBV subtypes and BZLF1 Zp variants were examined in Japanese patients with infectious mononucleosis, chronic active EBV infection, and EBV-associated hemophagocytic lymphohistiocytosis. The results of EBV typing showed that samples of infectious mononucleosis, chronic active EBV infection, and EBV-associated hemophagocytic lymphohistiocytosis all belonged to EBV type 1. However, sequencing analysis of BZLF1 Zp found three polymorphic Zp variants in the same samples. The Zp-P prototype and the Zp-V3 variant were both detected in infectious mononucleosis and chronic active EBV infection. Furthermore, a novel variant previously identified in Chinese children with infectious mononucleosis, Zp-V1, was also found in 3 of 18 samples of infectious mononucleosis, where it coexisted with the Zp-P prototype. This is the first evidence that the EBV variant distribution in Japanese patients resembles that found in other Asian patients. The expression levels of 29 chronic active EBV infection-associated cellular genes were also compared in the three EBV-related disorders, using quantitative real-time reverse transcription polymerase chain reaction analysis. Two upregulated genes, RIPK2 and CDH9, were identified as common specific markers for chronic active EBV infection in both in vitro and in vivo studies. RIPK2 activates apoptosis and autophagy, and could be responsible for the pathogenesis of chronic active EBV infection.

  6. Metal-specific control of gene expression mediated by Bradyrhizobium japonicum Mur and Escherichia coli Fur is determined by the cellular context.

    PubMed

    Hohle, Thomas H; O'Brian, Mark R

    2016-07-01

    Bradyrhizobium japonicum Mur and Escherichia coli Fur are manganese- and iron-responsive transcriptional regulators, respectively, that belong to the same protein family. Here, we show that neither Mur nor Fur discriminate between Fe(2+) and Mn(2+) in vitro nor is there a metal preference for conferral of DNA-binding activity on the purified proteins. When expressed in E. coli, B. japonicum Mur responded to iron, but not manganese, as determined by in vivo promoter occupancy and transcriptional repression activity. Moreover, E. coli Fur activity was manganese-dependent in B. japonicum. Total and chelatable iron levels were higher in E. coli than in B. japonicum under identical growth conditions, and Mur responded to iron in a B. japonicum iron export mutant that accumulated high levels of the metal. However, elevated manganese content in E. coli did not confer activity on Fur or Mur, suggesting a regulatory pool of manganese in B. japonicum that is absent in E. coli. We conclude that the metal selectivity of Mur and Fur depends on the cellular context in which they function, not on intrinsic properties of the proteins. Also, the novel iron sensing mechanism found in the rhizobia may be an evolutionary adaptation to the cellular manganese status.

  7. Cellular Mechanism for Impaired Hepatitis C Virus Clearance by Interferon Associated with IFNL3 Gene Polymorphisms Relates to Intrahepatic Interferon-λ Expression.

    PubMed

    Ferraris, Pauline; Chandra, Partha K; Panigrahi, Rajesh; Aboulnasr, Fatma; Chava, Srinivas; Kurt, Ramazan; Pawlotsky, Jean-Michel; Wilkens, Ludwig; Osterlund, Pamela; Hartmann, Rune; Balart, Luis A; Wu, Tong; Dash, Srikanta

    2016-04-01

    The single nucleotide polymorphism located within the IFNL3 (also known as IL28B) promoter is one of the host factors associated with hepatitis C virus (HCV) clearance by interferon (IFN)-α therapy; however the mechanism remains unknown. We investigated how IL28B gene polymorphism influences HCV clearance with infected primary human hepatocytes, liver biopsies, and hepatoma cell lines. Our study confirms that the rs12979860-T/T genotype has a strong correlation with ss469415590-ΔG/ΔG single nucleotide polymorphism that produces IFN-λ4 protein. We found that IFN-α and IFN-λ1 antiviral activity against HCV was impaired in IL28B T/T infected hepatocytes compared with C/C genotype. Western blot analysis showed that IL28B TT genotype hepatocytes expressed higher levels of IFN-λ proteins (IL28B, IL-29), preactivated IFN-stimulated gene (ISG) expression, and impaired Stat phosphorylation when stimulated with either IFN-α or IFN-λ1. Furthermore, we showed that silencing IFN-λ1 in T/T cell line reduced basal ISG expression and improved antiviral activity. Likewise, overexpression of IFN-λ (1 to 4) in C/C cells induced basal ISG expression and prevented IFN-α antiviral activity. We showed that IFN-λ4, produced at low level only in T/T cells induced expression of IL28B and IL-29 and prevented IFN-α antiviral activity in HCV cell culture. Our results suggest that IFN-λ4 protein expression associated with the IL28B-T/T variant preactivates the Janus kinase-Stat signaling, leading to impaired HCV clearance by both IFN-α and IFN-λ.

  8. Regulatory Interaction between the Cellular Restriction Factor IFI16 and Viral pp65 (pUL83) Modulates Viral Gene Expression and IFI16 Protein Stability

    PubMed Central

    Pautasso, Sara; von Einem, Jens; Marschall, Manfred; Plachter, Bodo

    2016-01-01

    ABSTRACT A key player in the intrinsic resistance against human cytomegalovirus (HCMV) is the interferon-γ-inducible protein 16 (IFI16), which behaves as a viral DNA sensor in the first hours postinfection and as a repressor of viral gene transcription in the later stages. Previous studies on HCMV replication demonstrated that IFI16 binds to the viral protein kinase pUL97, undergoes phosphorylation, and relocalizes to the cytoplasm of infected cells. In this study, we demonstrate that the tegument protein pp65 (pUL83) recruits IFI16 to the promoter of the UL54 gene and downregulates viral replication, as shown by use of the HCMV mutant v65Stop, which lacks pp65 expression. Interestingly, at late time points of HCMV infection, IFI16 is stabilized by its interaction with pp65, which stood in contrast to IFI16 degradation, observed in herpes simplex virus 1 (HSV-1)-infected cells. Moreover, we found that its translocation to the cytoplasm, in addition to pUL97, strictly depends on pp65, as demonstrated with the HCMV mutant RV-VM1, which expresses a form of pp65 unable to translocate into the cytoplasm. Thus, these data reveal a dual role for pp65: during early infection, it modulates IFI16 activity at the promoter of immediate-early and early genes; subsequently, it delocalizes IFI16 from the nucleus into the cytoplasm, thereby stabilizing and protecting it from degradation. Overall, these data identify a novel activity of the pp65/IFI16 interactome involved in the regulation of UL54 gene expression and IFI16 stability during early and late phases of HCMV replication. IMPORTANCE The DNA sensor IFI16, a member of the PYHIN proteins, restricts HCMV replication by impairing viral DNA synthesis. Using a mutant virus lacking the tegument protein pp65 (v65Stop), we demonstrate that pp65 recruits IFI16 to the early UL54 gene promoter. As a putative counteraction to its restriction activity, pp65 supports the nucleocytoplasmic export of IFI16, which was demonstrated with the

  9. Responses of the European flounder Platichthys flesus to the chemical stress in estuaries: load of contaminants, gene expression, cellular impact and growth rate.

    PubMed

    Evrard, Estérine; Devaux, Alain; Bony, Sylvie; Burgeot, Thierry; Riso, Ricardo; Budzinski, Hélène; Le Du, Marie; Quiniou, Louis; Laroche, Jean

    2010-03-01

    European flounder responses to the chemical stress were assessed by a comparative approach on four estuaries displaying contrasted patterns of contamination. The contamination typology of the estuaries was investigated by individual measurements of contaminants in fish. Molecular and physiological responses were studied by gene expression, genotoxicity, neurotoxicity and growth rate. Fishes in contaminated estuaries were characterized by high levels of bioaccumulated contaminants, slow energetic metabolism and reduced growth rate, in contrast to the fish responses in the reference site. A seasonal effect was highlighted for contaminated flounder populations, with high PCB levels, high genotoxicity and elevated detoxification rate in summer compared with winter.

  10. Activation of cryptic 3' splice sites within introns of cellular genes following gene entrapment.

    PubMed

    Osipovich, Anna B; White-Grindley, Erica K; Hicks, Geoffrey G; Roshon, Michael J; Shaffer, Christian; Moore, Jason H; Ruley, H Earl

    2004-01-01

    Gene trap vectors developed for genome-wide mutagenesis can be used to study factors governing the expression of exons inserted throughout the genome. For example, entrapment vectors consisting of a partial 3'-terminal exon [i.e. a neomycin resistance gene (Neo), a poly(A) site, but no 3' splice site] were typically expressed following insertion into introns, from cellular transcripts that spliced to cryptic 3' splice sites present either within the targeting vector or in the adjacent intron. A vector (U3NeoSV1) containing the wild-type Neo sequence preferentially disrupted genes that spliced in-frame to a cryptic 3' splice site in the Neo coding sequence and expressed functional neomycin phosphotransferase fusion proteins. Removal of the cryptic Neo 3' splice site did not reduce the proportion of clones with inserts in introns; rather, the fusion transcripts utilized cryptic 3' splice sites present in the adjacent intron or generated by virus integration. However, gene entrapment with U3NeoSV2 was considerably more random than with U3NeoSV1, consistent with the widespread occurrence of potential 3' splice site sequences in the introns of cellular genes. These results clarify the mechanisms of gene entrapment by U3 gene trap vectors and illustrate features of exon definition required for 3' processing and polyadenylation of cellular transcripts.

  11. Activation of cryptic 3′ splice sites within introns of cellular genes following gene entrapment

    PubMed Central

    Osipovich, Anna B.; White-Grindley, Erica K.; Hicks, Geoffrey G.; Roshon, Michael J.; Shaffer, Christian; Moore, Jason H.; Ruley, H. Earl

    2004-01-01

    Gene trap vectors developed for genome-wide mutagenesis can be used to study factors governing the expression of exons inserted throughout the genome. For example, entrapment vectors consisting of a partial 3′-terminal exon [i.e. a neomycin resistance gene (Neo), a poly(A) site, but no 3′ splice site] were typically expressed following insertion into introns, from cellular transcripts that spliced to cryptic 3′ splice sites present either within the targeting vector or in the adjacent intron. A vector (U3NeoSV1) containing the wild-type Neo sequence preferentially disrupted genes that spliced in-frame to a cryptic 3′ splice site in the Neo coding sequence and expressed functional neomycin phosphotransferase fusion proteins. Removal of the cryptic Neo 3′ splice site did not reduce the proportion of clones with inserts in introns; rather, the fusion transcripts utilized cryptic 3′ splice sites present in the adjacent intron or generated by virus integration. However, gene entrapment with U3NeoSV2 was considerably more random than with U3NeoSV1, consistent with the widespread occurrence of potential 3′ splice site sequences in the introns of cellular genes. These results clarify the mechanisms of gene entrapment by U3 gene trap vectors and illustrate features of exon definition required for 3′ processing and polyadenylation of cellular transcripts. PMID:15155860

  12. Cellular functions of genetically imprinted genes in human and mouse as annotated in the gene ontology.

    PubMed

    Hamed, Mohamed; Ismael, Siba; Paulsen, Martina; Helms, Volkhard

    2012-01-01

    By analyzing the cellular functions of genetically imprinted genes as annotated in the Gene Ontology for human and mouse, we found that imprinted genes are often involved in developmental, transport and regulatory processes. In the human, paternally expressed genes are enriched in GO terms related to the development of organs and of anatomical structures. In the mouse, maternally expressed genes regulate cation transport as well as G-protein signaling processes. Furthermore, we investigated if imprinted genes are regulated by common transcription factors. We identified 25 TF families that showed an enrichment of binding sites in the set of imprinted genes in human and 40 TF families in mouse. In general, maternally and paternally expressed genes are not regulated by different transcription factors. The genes Nnat, Klf14, Blcap, Gnas and Ube3a contribute most to the enrichment of TF families. In the mouse, genes that are maternally expressed in placenta are enriched for AP1 binding sites. In the human, we found that these genes possessed binding sites for both, AP1 and SP1.

  13. Cellular Functions and Gene and Protein Expression Profiles in Endothelial Cells Derived from Moyamoya Disease-Specific iPS Cells

    PubMed Central

    Hamauchi, Shuji; Shichinohe, Hideo; Uchino, Haruto; Yamaguchi, Shigeru; Nakayama, Naoki; Kazumata, Ken; Osanai, Toshiya; Abumiya, Takeo; Houkin, Kiyohiro; Era, Takumi

    2016-01-01

    Background and purpose Moyamoya disease (MMD) is a slow, progressive steno-occlusive disease, arising in the terminal portions of the cerebral internal carotid artery. However, the functions and characteristics of the endothelial cells (ECs) in MMD are unknown. We analyzed these features using induced pluripotent stem cell (iPSC)-derived ECs. Methods iPSC lines were established from the peripheral blood of three patients with MMD carrying the variant RNF213 R4810K, and three healthy persons used as controls. After the endothelial differentiation of iPSCs, CD31+CD144+ cells were purified as ECs using a cell sorter. We analyzed their proliferation, angiogenesis, and responses to some angiogenic factors, namely VEGF, bFGF, TGF-β, and BMP4. The ECs were also analyzed using DNA microarray and proteomics to perform comprehensive gene and protein expression analysis. Results Angiogenesis was significantly impaired in MMD regardless of the presence of any angiogenic factor. On the contrary, endothelial proliferation was not significant between control- and MMD-derived cells. Regarding DNA microarray, pathway analysis illustrated that extracellular matrix (ECM) receptor-related genes, including integrin β3, were significantly downregulated in MMD. Proteomic analysis revealed that cytoskeleton-related proteins were downregulated and splicing regulation-related proteins were upregulated in MMD. Conclusions Downregulation of ECM receptor-related genes may be associated with impaired angiogenic activity in ECs derived from iPSCs from patients with MMD. Upregulation of splicing regulation-related proteins implied differences in splicing patterns between control and MMD ECs. PMID:27662211

  14. Engineering the cellular protein secretory pathway for enhancement of recombinant tissue plasminogen activator expression in Chinese hamster ovary cells: effects of CERT and XBP1s genes.

    PubMed

    Rahimpour, Azam; Vaziri, Behrouz; Moazzami, Reza; Nematollahi, Leila; Barkhordari, Farzaneh; Kokabee, Leila; Adeli, Ahmad; Mahboudi, Fereidoun

    2013-08-01

    Cell line development is the most critical and also the most time-consuming step in the production of recombinant therapeutic proteins. In this regard, a variety of vector and cell engineering strategies have been developed for generating high-producing mammalian cells; however, the cell line engineering approach seems to show various results on different recombinant protein producer cells. In order to improve the secretory capacity of a recombinant tissue plasminogen activator (t-PA)-producing Chinese hamster ovary (CHO) cell line, we developed cell line engineering approaches based on the ceramide transfer protein (CERT) and X-box binding protein 1 (XBP1) genes. For this purpose, CERT S132A, a mutant form of CERT that is resistant to phosphorylation, and XBP1s were overexpressed in a recombinant t-PA-producing CHO cell line. Overexpression of CERT S132A increased the specific productivity of t-PA-producing CHO cells up to 35%. In contrast, the heterologous expression of XBP1s did not affect the t-PA expression rate. Our results suggest that CERTS132A- based secretion engineering could be an effective strategy for enhancing recombinant t- PA production in CHO cells.

  15. Does FACS perturb gene expression?

    PubMed

    Richardson, Graham M; Lannigan, Joanne; Macara, Ian G

    2015-02-01

    Fluorescence activated cell sorting is the technique most commonly used to separate primary mammary epithelial sub-populations. Many studies incorporate this technique before analyzing gene expression within specific cellular lineages. However, to our knowledge, no one has examined the effects of fluorescence activated cell sorting (FACS) separation on short-term transcriptional profiles. In this study, we isolated a heterogeneous mixture of cells from the mouse mammary gland. To determine the effects of the isolation and separation process on gene expression, we harvested RNA from the cells before enzymatic digestion, following enzymatic digestion, and following a mock FACS sort where the entire cohort of cells was retained. A strict protocol was followed to minimize disruption to the cells, and to ensure that no subpopulations were enriched or lost. Microarray analysis demonstrated that FACS causes minimal disruptions to gene expression patterns, but prior steps in the mammary cell isolation process are followed by upregulation of 18 miRNA's and rapid decreases in their predicted target transcripts. © 2015 International Society for Advancement of Cytometry.

  16. Epstein-Barr virus growth/latency III program alters cellular microRNA expression

    SciTech Connect

    Cameron, Jennifer E. Fewell, Claire Yin, Qinyan McBride, Jane Wang Xia Lin Zhen

    2008-12-20

    The Epstein-Barr virus (EBV) is associated with lymphoid and epithelial cancers. Initial EBV infection alters lymphocyte gene expression, inducing cellular proliferation and differentiation as the virus transitions through consecutive latency transcription programs. Cellular microRNAs (miRNAs) are important regulators of signaling pathways and are implicated in carcinogenesis. The extent to which EBV exploits cellular miRNAs is unknown. Using micro-array analysis and quantitative PCR, we demonstrate differential expression of cellular miRNAs in type III versus type I EBV latency including elevated expression of miR-21, miR-23a, miR-24, miR-27a, miR-34a, miR-146a and b, and miR-155. In contrast, miR-28 expression was found to be lower in type III latency. The EBV-mediated regulation of cellular miRNAs may contribute to EBV signaling and associated cancers.

  17. Combined Effects of High-Dose Bisphenol A and Oxidizing Agent (KBrO3) on Cellular Microenvironment, Gene Expression, and Chromatin Structure of Ku70-deficient Mouse Embryonic Fibroblasts

    PubMed Central

    Gassman, Natalie R.; Coskun, Erdem; Jaruga, Pawel; Dizdaroglu, Miral; Wilson, Samuel H.

    2016-01-01

    Background: Exposure to bisphenol A (BPA) has been reported to alter global gene expression, induce epigenetic modifications, and interfere with complex regulatory networks of cells. In addition to these reprogramming events, we have demonstrated that BPA exposure generates reactive oxygen species and promotes cellular survival when co-exposed with the oxidizing agent potassium bromate (KBrO3). Objectives: We determined the cellular microenvironment changes induced by co-exposure of BPA and KBrO3 versus either agent alone. Methods: Ku70-deficient cells were exposed to 150 μM BPA, 20 mM KBrO3, or co-exposed to both agents. Four and 24 hr post-damage initiation by KBrO3, with BPA-only samples timed to coincide with these designated time points, we performed whole-genome microarray analysis and evaluated chromatin structure, DNA lesion load, glutathione content, and intracellular pH. Results: We found that 4 hr post-damage initiation, BPA exposure and co-exposure transiently condensed chromatin compared with untreated and KBrO3-only treated cells; the transcription of DNA repair proteins was also reduced. At this time point, BPA exposure and co-exposure also reduced the change in intracellular pH observed after treatment with KBrO3 alone. Twenty-four hours post-damage initiation, BPA-exposed cells showed less condensed chromatin than cells treated with KBrO3 alone; the intracellular pH of the co-exposed cells was significantly reduced compared with untreated and KBrO3-treated cells; and significant up-regulation of DNA repair proteins was observed after co-exposure. Conclusion: These results support the induction of an adaptive response by BPA co-exposure that alters the microcellular environment and modulates DNA repair. Further work is required to determine whether BPA induces similar DNA lesions in vivo at environmentally relevant doses; however, in the Ku70-deficient mouse embryonic fibroblasts, exposure to a high dose of BPA was associated with changes in the

  18. Mechanically enhanced microcapsules for cellular gene therapy.

    PubMed

    Shen, F; Mazumder, M A J; Burke, N A D; Stöver, H D H; Potter, M A

    2009-07-01

    Microcapsules bearing a covalently cross-linked coating have been developed for cellular gene therapy as an improvement on alginate-poly(L-lysine)-alginate (APA) microcapsules that only have ionic cross-linking. In this study, two mutually reactive polyelectrolytes, a polycation (designated C70), poly([2-(methacryloyloxy)ethyl]trimethylammonium chloride-co-2-aminoethyl methacrylate hydrochloride) and a polyanion (designated A70), poly(sodium methacrylate-co-2-(methacryloyloxy)ethyl acetoacetate), were used during the microcapsule fabrication. Ca-alginate beads were sequentially laminated with C70, A70, poly(L-lysine) (PLL), and alginate. The A70 reacts with both C70 and PLL to form a approximately 30 microm thick covalently cross-linked interpenetrating polymer network on the surface of the capsules. Confocal images confirmed the location of the C70/A70/PLL network and the stability of the network after 4 weeks implantation in mice. The mechanical and chemical resistance of the capsules was tested with a "stress test" where microcapsules were gently shaken in 0.003% EDTA for 15 min. APA capsules disappeared during this treatment, whereas the modified capsules, even those that had been retrieved from mice after 4-weeks implantation, remained intact. Analysis of solutions passing through model flat membranes showed that the molecular weight cut-off of alginate-C70-A70-PLL-alginate is similar to that of alginate-PLL-alginate. Recombinant cells encapsulated in APA and modified capsules were able to secrete luciferase into culture media. The modified capsules were found to capture some components of regular culture media used during preparation, causing an immune reaction in implanted mice, but use of UltraCulture serum-free medium was found to prevent this immune reaction. In vivo biocompatibility of the new capsules was similar to the APA capsules, with no sign of clinical toxicity on complete blood counts and liver function tests. The increased stability of the

  19. Differential expression of the ras gene family in mice.

    PubMed Central

    Leon, J; Guerrero, I; Pellicer, A

    1987-01-01

    We compared the expression of the ras gene family (H-ras, K-ras, and N-ras) in adult mouse tissues and during development. We found substantial variations in expression among different organs and in the amounts of the different transcripts originating from each gene, especially for the N-ras gene. The expression patterns were consistent with the reported preferential tissue activation of ras genes and suggested different cellular functions for each of the ras genes. Images PMID:3600635

  20. Inhibition of p38 MAPK during cellular activation modulate gene expression of head kidney leukocytes isolated from Atlantic salmon (Salmo salar) fed soy bean oil or fish oil based diets.

    PubMed

    Holen, E; Winterthun, S; Du, Z-Y; Krøvel, A V

    2011-01-01

    Head kidney leukocytes isolated from Atlantic salmon fed either a diet based on fish oil (FO) or soy bean oil (VO) were used in order to evaluate if different lipid sources could contribute to cellular activation of the salmon innate immune system. A specific inhibitor of p38 MAPK, SB202190, was used to investigate the effect of lipopolysaccharide (LPS) signalling in the head kidney leukocytes. The results show that LPS up regulate IL-1β, TNF-α, Cox2 expression in leukocytes isolated from fish fed either diet. The p38 MAPK inhibitor, SB202190, reduced the LPS induced expression of these genes in both dietary groups. In LPS stimulated leukocytes isolated from VO fed fish, SB202190 showed a clear dose dependent inhibitory effect on IL-1β, TNF-α and Cox2 expression. This effect was also observed for Cox2 in leukocytes isolated from FO fed fish. Furthermore, there was a stronger mean induction of Cox2 in LPS stimulated leucocytes isolated from the VO-group compared to LPS stimulated leukocytes isolated from the FO-group. In both dietary groups, LPS stimulation of salmon head kidney leukocytes increased the induction of CD83, a dendrite cell marker, while the inhibitor reduced CD83 expression in the VO fed fish only. The inhibitor also clearly reduced hsp27 expression in VO fed fish. Indicating a p38 MAPK feedback loop, LPS significantly inhibited the expression of p38MAPK itself in both diets, while SB202190 increased p38MAPK expression especially in the VO diet group. hsp70 expression was not affected by any treatment or feed composition. There were also differences in p38MAPK protein phosphorylation comparing treatment groups but no obvious difference comparing the two dietary groups. The results indicate that dietary fatty acids have the ability to modify signalling through p38 MAPK which may have consequences for the fish's ability to handle infections and stress. Signalling through p38MAPK is ligand dependent and affects gene and protein expression differently.

  1. Combined clustering models for the analysis of gene expression

    SciTech Connect

    Angelova, M. Ellman, J.

    2010-02-15

    Clustering has become one of the fundamental tools for analyzing gene expression and producing gene classifications. Clustering models enable finding patterns of similarity in order to understand gene function, gene regulation, cellular processes and sub-types of cells. The clustering results however have to be combined with sequence data or knowledge about gene functionality in order to make biologically meaningful conclusions. In this work, we explore a new model that integrates gene expression with sequence or text information.

  2. Gene expression profiles in irradiated cancer cells

    NASA Astrophysics Data System (ADS)

    Minafra, L.; Bravatà, V.; Russo, G.; Ripamonti, M.; Gilardi, M. C.

    2013-07-01

    Knowledge of the molecular and genetic mechanisms underlying cellular response to radiation may provide new avenues to develop innovative predictive tests of radiosensitivity of tumours and normal tissues and to improve individual therapy. Nowadays very few studies describe molecular changes induced by hadrontherapy treatments, therefore this field has to be explored and clarified. High-throughput methodologies, such as DNA microarray, allow us to analyse mRNA expression of thousands of genes simultaneously in order to discover new genes and pathways as targets of response to hadrontherapy. Our aim is to elucidate the molecular networks involved in the sensitivity/resistance of cancer cell lines subjected to hadrontherapy treatments with a genomewide approach by using cDNA microarray technology to identify gene expression profiles and candidate genes responsible of differential cellular responses.

  3. Gene expression profiles in irradiated cancer cells

    SciTech Connect

    Minafra, L.; Bravatà, V.; Russo, G.; Ripamonti, M.; Gilardi, M. C.

    2013-07-26

    Knowledge of the molecular and genetic mechanisms underlying cellular response to radiation may provide new avenues to develop innovative predictive tests of radiosensitivity of tumours and normal tissues and to improve individual therapy. Nowadays very few studies describe molecular changes induced by hadrontherapy treatments, therefore this field has to be explored and clarified. High-throughput methodologies, such as DNA microarray, allow us to analyse mRNA expression of thousands of genes simultaneously in order to discover new genes and pathways as targets of response to hadrontherapy. Our aim is to elucidate the molecular networks involved in the sensitivity/resistance of cancer cell lines subjected to hadrontherapy treatments with a genomewide approach by using cDNA microarray technology to identify gene expression profiles and candidate genes responsible of differential cellular responses.

  4. Protein splicing: selfish genes invade cellular proteins.

    PubMed

    Neff, N F

    1993-12-01

    Protein splicing is a series of enzymatic events involving intramolecular protein breakage, rejoining and intron homing, in which introns are able to promote the recombinative transposition of their own coding sequences. Eukaryotic and prokaryotic spliced proteins have conserved similar gene structure, but little amino acid identity. The genes coding for these spliced proteins contain internal in-frame introns that encode polypeptides that apparently self-excise from the resulting host protein sequences. Excision of the 'protein intron' is coupled with joining of the two flanking protein regions encoded by exons of the host gene. Some introns of this type encode DNA endonucleases, related to Group I RNA intron gene products, that stimulate gene conversion and self-transmission.

  5. 78 FR 70307 - Guidance for Industry: Preclinical Assessment of Investigational Cellular and Gene Therapy...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-25

    ... Investigational Cellular and Gene Therapy Products; Availability AGENCY: Food and Drug Administration, HHS. ACTION... entitled ``Guidance for Industry: Preclinical Assessment of Investigational Cellular and Gene Therapy... and Gene Therapies (OCTGT). The product areas covered by this guidance are cellular therapy,...

  6. Expression pattern, ethanol-metabolizing activities, and cellular localization of alcohol and aldehyde dehydrogenases in human large bowel: association of the functional polymorphisms of ADH and ALDH genes with hemorrhoids and colorectal cancer.

    PubMed

    Chiang, Chien-Ping; Jao, Shu-Wen; Lee, Shiao-Pieng; Chen, Pei-Chi; Chung, Chia-Chi; Lee, Shou-Lun; Nieh, Shin; Yin, Shih-Jiun

    2012-02-01

    Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) are principal enzymes responsible for metabolism of ethanol. Functional polymorphisms of ADH1B, ADH1C, and ALDH2 genes occur among racial populations. The goal of this study was to systematically determine the functional expressions and cellular localization of ADHs and ALDHs in human rectal mucosa, the lesions of adenocarcinoma and hemorrhoid, and the genetic association of allelic variations of ADH and ALDH with large bowel disorders. Twenty-one surgical specimens of rectal adenocarcinoma and the adjacent normal mucosa, including 16 paired tissues of rectal tumor, normal mucosae of rectum and sigmoid colon from the same individuals, and 18 surgical mixed hemorrhoid specimens and leukocyte DNA samples from 103 colorectal cancer patients, 67 hemorrhoid patients, and 545 control subjects recruited in previous study, were investigated. The isozyme/allozyme expression patterns of ADH and ALDH were identified by isoelectric focusing and the activities were assayed spectrophotometrically. The protein contents of ADH/ALDH isozymes were determined by immunoblotting using the corresponding purified class-specific antibodies; the cellular activity and protein localizations were detected by immunohistochemistry and histochemistry, respectively. Genotypes of ADH1B, ADH1C, and ALDH2 were determined by polymerase chain reaction-restriction fragment length polymorphisms. At 33mM ethanol, pH 7.5, the activity of ADH1C*1/1 phenotypes exhibited 87% higher than that of the ADH1C*1/*2 phenotypes in normal rectal mucosa. The activity of ALDH2-active phenotypes of rectal mucosa was 33% greater than ALDH2-inactive phenotypes at 200μM acetaldehyde. The protein contents in normal rectal mucosa were in the following order: ADH1>ALDH2>ADH3≈ALDH1A1, whereas those of ADH2, ADH4, and ALDH3A1 were fairly low. Both activity and content of ADH1 were significantly decreased in rectal tumors, whereas the ALDH activity remained

  7. Gene expression and cellular localization of ROMKs in the gills and kidney of Mozambique tilapia acclimated to fresh water with high potassium concentration.

    PubMed

    Furukawa, Fumiya; Watanabe, Soichi; Kakumura, Keigo; Hiroi, Junya; Kaneko, Toyoji

    2014-12-01

    Regulation of plasma K(+) levels in narrow ranges is vital to vertebrate animals. Since seawater (SW) teleosts are loaded with excess K(+), they constantly excrete K(+) from the gills. However, the K(+) regulatory mechanisms in freshwater (FW)-acclimated teleosts are still unclear. We aimed to identify the possible K(+) regulatory mechanisms in the gills and kidney, the two major osmoregulatory organs, of FW-acclimated Mozambique tilapia (Oreochromis mossambicus). As a potential molecular candidate for renal K(+) handling, a putative renal outer medullary K(+) channel (ROMK) was cloned from the tilapia kidney and tentatively named "ROMKb"; another ROMK previously cloned from the tilapia gills was thus renamed "ROMKa". The fish were acclimated to control FW or to high-K(+) (H-K) FW for 1 wk, and we assessed physiological responses of tilapia to H-K treatment. As a result, urinary K(+) levels were slightly higher in H-K fish, implying a role of the kidney in K(+) excretion. However, the mRNA expression levels of both ROMKa and ROMKb were very low in the kidney, while that of K(+)/Cl(-) cotransporter 1 (KCC1) was robust. In the gills, ROMKa mRNA was markedly upregulated in H-K fish. Immunofluorescence staining showed that branchial ROMKa was expressed at the apical membrane of type I and type III ionocytes, and the ROMKa immunosignals were more intense in H-K fish than in control fish. The present study suggests that branchial ROMKa takes a central role for K(+) regulation in FW conditions and that K(+) excretion via the gills is activated irrespective of environmental salinity.

  8. Modeling gene expression in time and space.

    PubMed

    Rué, Pau; Garcia-Ojalvo, Jordi

    2013-01-01

    Cell populations rarely exhibit gene-expression profiles that are homogeneous in time and space. In the temporal domain, dynamical behaviors such as oscillations and pulses of protein production pervade cell biology, underlying phenomena as diverse as circadian rhythmicity, cell cycle control, stress and damage responses, and stem-cell pluripotency. In multicellular populations, spatial heterogeneities are crucial for decision making and development, among many other functions. Cells need to exquisitely coordinate this temporal and spatial variation to survive. Although the spatiotemporal character of gene expression is challenging to quantify experimentally at the level of individual cells, it is beneficial from the modeling viewpoint, because it provides strong constraints that can be probed by theoretically analyzing mathematical models of candidate gene and protein circuits. Here, we review recent examples of temporal dynamics and spatial patterning in gene expression to show how modeling such phenomenology can help us unravel the molecular mechanisms of cellular function.

  9. An extensive network of coupling among gene expression machines.

    PubMed

    Maniatis, Tom; Reed, Robin

    2002-04-04

    Gene expression in eukaryotes requires several multi-component cellular machines. Each machine carries out a separate step in the gene expression pathway, which includes transcription, several pre-messenger RNA processing steps and the export of mature mRNA to the cytoplasm. Recent studies lead to the view that, in contrast to a simple linear assembly line, a complex and extensively coupled network has evolved to coordinate the activities of the gene expression machines. The extensive coupling is consistent with a model in which the machines are tethered to each other to form 'gene expression factories' that maximize the efficiency and specificity of each step in gene expression.

  10. Modulation at a cellular level of the thyroid hormone receptor-mediated gene expression by 1,2,5,6,9,10-hexabromocyclododecane (HBCD), 4,4'-diiodobiphenyl (DIB), and nitrofen (NIP).

    PubMed

    Yamada-Okabe, Toshiko; Sakai, Haruya; Kashima, Yuji; Yamada-Okabe, Hisafumi

    2005-01-15

    Previously, we demonstrated that some endocrine disrupting chemicals affected thyroid hormone receptor (TR)-mediated gene expression in HeLaTR cells that stably expressed the human TRalpha1. To examine whether widely used brominated flame retardants and pesticides affect TR-mediated gene expression, those with organohalogen, which is also present in T3, were screened. To monitor the TR-mediated gene expression, HeLaTR cells were transfected with a luciferase gene that was linked to the thyroid hormone responsive element. Thus, transcription of the luciferase gene in HeLaTR cells is driven by TR. By screening 38 chemical agents, it was found that 4,4'-diiodobiphenyl (DIB), markedly, and 1,2,5,6,9,10-hexabromocyclododecane (HBCD) and nitrofen (NIP), to a much lesser extent but significantly, enhanced the expression of the luciferase gene at concentrations that did not affect the growth of HeLaTR cells. DIB also augmented the E2-induced expression of the luciferase gene that was linked to the estrogen responsive element in MCF7 cells, whereas HBCD and NIP did not. These results indicate that DIB augments TR- and ER-mediated gene expression, but HBCD and NIP affect only TR-mediated gene expression. Thus, there is a potential risk that HBCD, DIB, and NIP act as endocrine disrupters in animals and human beings.

  11. Dynamics of single-cell gene expression

    PubMed Central

    Longo, Diane; Hasty, Jeff

    2006-01-01

    Cellular behavior has traditionally been investigated by utilizing bulk-scale methods that measure average values for a population of cells. Such population-wide studies mask the behavior of individual cells and are often insufficient for characterizing biological processes in which cellular heterogeneity plays a key role. A unifying theme of many recent studies has been a focus on the development and utilization of single-cell experimental techniques that are capable of probing key biological phenomena in individual living cells. Recently, novel information about gene expression dynamics has been obtained from single-cell experiments that draw upon the unique capabilities of fluorescent reporter proteins. PMID:17130866

  12. Carcinogen-induced trans activation of gene expression.

    PubMed Central

    Kleinberger, T; Flint, Y B; Blank, M; Etkin, S; Lavi, S

    1988-01-01

    We report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later. Images PMID:2835673

  13. Carcinogen-induced trans activation of gene expression

    SciTech Connect

    Kleinberger, T.; Flint, Y.B.; Blank, M.; Etkin, S.; Lavi, S.

    1988-03-01

    The authors report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later.

  14. Gene expression during normal and malignant differentiation

    SciTech Connect

    Andersson, L.C.; Gahmberg, C.G.; Ekblom, P.

    1985-01-01

    This book contains 18 selections. Some of the titles are: Exploring Carcinogenesis with Retroviral and Cellular Oncogenes; Retroviruses, Oncogenes and Evolution; HTLV and Human Neoplasi; Modes of Activation of cMyc Oncogene in B and T Lymphoid Tumors; The Structure and Function of the Epidermal Growth Factor Receptor: Its Relationship to the Protein Product of the V-ERB-B Oncogene; and Expression of Human Retrovirus Genes in Normal and Neoplastic Epithelial Cells.

  15. HIF-1α expression correlates with cellular apoptosis, angiogenesis and clinical prognosis in rectal carcinoma.

    PubMed

    Feng, Liu; Tao, Lin; Dawei, He; Xuliang, Li; Xiaodong, Luo

    2014-07-01

    Regional hypoxia caused by accelerated cell proliferation and overgrowth is an important characteristic of neoplasm. Hypoxia can cause a series of changes in gene transcription and protein expression, thereby not only inducing tumor cell resistance to radiotherapy and chemotherapy but also promoting tumor invasion and metastasis. This study aimed to investigate the relationship between HIF-1α expression and cellular apoptosis, angiogenesis and clinical prognosis in rectal carcinoma. In 113 rectal carcinoma cases, cellular apoptosis was analyzed by the in situ terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) assay, whereas the levels of HIF-1α expression, VEGF expression, microvessel density (MVD) and lymphatic vessel density(LVD) were examined by immunohistochemical staining. HIF-1 expression was detected in 67 of 113 rectal carcinoma cases (59.3 %). A positive correlation was found among HIF-1α expression, cellular apoptosis and angiogenesis. The 5-year survival rate in the HIF-1α-negative group was significantly higher than that in the HIF-1α-positive group (81.34 % versus 50 %, P < 0.05). According to the Cox regression analysis, HIF-1α expression, VEGF expression and cellular apoptosis index were independent risk factors for clinical prognosis in rectal carcinoma. Aberrant HIF-1α expression correlates with apoptosis inhibition, angiogenesis and poor prognosis in rectal carcinoma.

  16. Cell Type-Specific Expression Analysis to Identify Putative Cellular Mechanisms for Neurogenetic Disorders

    PubMed Central

    Xu, Xiaoxiao; Wells, Alan B.; O'Brien, David R.; Nehorai, Arye

    2014-01-01

    Recent advances have substantially increased the number of genes that are statistically associated with complex genetic disorders of the CNS such as autism and schizophrenia. It is now clear that there will likely be hundreds of distinct loci contributing to these disorders, underscoring a remarkable genetic heterogeneity. It is unclear whether this genetic heterogeneity indicates an equal heterogeneity of cellular mechanisms for these diseases. The commonality of symptoms across patients suggests there could be a functional convergence downstream of these loci upon a limited number of cell types or circuits that mediate the affected behaviors. One possible mechanism for this convergence would be the selective expression of at least a subset of these genes in the cell types that comprise these circuits. Using profiling data from mice and humans, we have developed and validated an approach, cell type-specific expression analysis, for identifying candidate cell populations likely to be disrupted across sets of patients with distinct genetic lesions. Using human genetics data and postmortem gene expression data, our approach can correctly identify the cell types for disorders of known cellular etiology, including narcolepsy and retinopathies. Applying this approach to autism, a disease where the cellular mechanism is unclear, indicates there may be multiple cellular routes to this disorder. Our approach may be useful for identifying common cellular mechanisms arising from distinct genetic lesions. PMID:24453331

  17. CIRCADIAN CLOCK AND CELL CYCLE GENE EXPRESSION

    PubMed Central

    Metz, Richard P.; Qu, Xiaoyu; Laffin, Brian; Earnest, David; Porter, Weston W.

    2009-01-01

    Mouse mammary epithelial cells (HC-11) and mammary tissues were analyzed for developmental changes in circadian clock, cellular proliferation and differentiation marker genes. Expression of the clock genes, Per1 and Bmal1, were elevated in differentiated HC-11 cells whereas Per2 mRNA levels were higher in undifferentiated cells. This differentiation-dependent profile of clock gene expression was consistent with that observed in mouse mammary glands as Per1 and Bmal1 mRNA levels were elevated in late pregnant and lactating mammary tissues, while Per2 expression was higher in proliferating virgin and early pregnant glands. In both HC-11 cells and mammary glands, elevated Per2 expression was positively correlated with c-Myc and Cyclin D1 mRNA levels while Per1 and Bmal1 expression changed in conjunction with ß-casein mRNA levels. Interestingly, developmental stage had differential effects on rhythms of clock gene expression in the mammary gland. These data suggest that circadian clock genes may play a role in mouse mammary gland development and differentiation. PMID:16261617

  18. The Role of Nuclear Bodies in Gene Expression and Disease

    PubMed Central

    Morimoto, Marie; Boerkoel, Cornelius F.

    2013-01-01

    This review summarizes the current understanding of the role of nuclear bodies in regulating gene expression. The compartmentalization of cellular processes, such as ribosome biogenesis, RNA processing, cellular response to stress, transcription, modification and assembly of spliceosomal snRNPs, histone gene synthesis and nuclear RNA retention, has significant implications for gene regulation. These functional nuclear domains include the nucleolus, nuclear speckle, nuclear stress body, transcription factory, Cajal body, Gemini of Cajal body, histone locus body and paraspeckle. We herein review the roles of nuclear bodies in regulating gene expression and their relation to human health and disease. PMID:24040563

  19. Method of controlling gene expression

    DOEpatents

    Peters, Norman K.; Frost, John W.; Long, Sharon R.

    1991-12-03

    A method of controlling expression of a DNA segment under the control of a nod gene promoter which comprises administering to a host containing a nod gene promoter an amount sufficient to control expression of the DNA segment of a compound of the formula: ##STR1## in which each R is independently H or OH, is described.

  20. The flow of gene expression.

    PubMed

    Misteli, Tom

    2004-03-01

    Gene expression is a highly interconnected multistep process. A recent meeting in Iguazu Falls, Argentina, highlighted the need to uncover both the molecular details of each single step as well as the mechanisms of coordination among processes in order to fully understand the expression of genes.

  1. Discovering modulators of gene expression

    PubMed Central

    Babur, Özgün; Demir, Emek; Gönen, Mithat; Sander, Chris; Dogrusoz, Ugur

    2010-01-01

    Proteins that modulate the activity of transcription factors, often called modulators, play a critical role in creating tissue- and context-specific gene expression responses to the signals cells receive. GEM (Gene Expression Modulation) is a probabilistic framework that predicts modulators, their affected targets and mode of action by combining gene expression profiles, protein–protein interactions and transcription factor–target relationships. Using GEM, we correctly predicted a significant number of androgen receptor modulators and observed that most modulators can both act as co-activators and co-repressors for different target genes. PMID:20466809

  2. Immediate–Early (IE) gene regulation of cytomegalovirus: IE1- and pp71-mediated viral strategies against cellular defenses

    PubMed Central

    Torres, Lilith; Tang, Qiyi

    2015-01-01

    Three crucial hurdles hinder studies on human cytomegalovirus (HCMV): strict species specificity, differences between in vivo and in vitro infection, and the complexity of gene regulation. Ever since the sequencing of the whole genome was first accomplished, functional studies on individual genes have been the mainstream in the CMV field. Gene regulation has therefore been elucidated in a more detailed fashion. However, viral gene regulation is largely controlled by both cellular and viral components. In other words, viral gene expression is determined by the virus–host interaction. Generally, cells respond to viral infection in a defensive pattern; at the same time, viruses try to counteract the cellular defense or else hide in the host (latency). Viruses evolve effective strategies against cellular defense in order to achieve replicative success. Whether or not they are successful, cellular defenses remain in the whole viral replication cycle: entry, immediate–early (IE) gene expression, early gene expression, DNA replication, late gene expression, and viral egress. Many viral strategies against cellular defense, and which occur in the immediate–early time of viral infection, have been documented. In this review, we will summarize the documented biological functions of IE1 and pp71 proteins, especially with regard to how they counteract cellular intrinsic defenses. PMID:25501994

  3. Studying the complex expression dependences between sets of coexpressed genes.

    PubMed

    Huerta, Mario; Casanova, Oriol; Barchino, Roberto; Flores, Jose; Querol, Enrique; Cedano, Juan

    2014-01-01

    Organisms simplify the orchestration of gene expression by coregulating genes whose products function together in the cell. The use of clustering methods to obtain sets of coexpressed genes from expression arrays is very common; nevertheless there are no appropriate tools to study the expression networks among these sets of coexpressed genes. The aim of the developed tools is to allow studying the complex expression dependences that exist between sets of coexpressed genes. For this purpose, we start detecting the nonlinear expression relationships between pairs of genes, plus the coexpressed genes. Next, we form networks among sets of coexpressed genes that maintain nonlinear expression dependences between all of them. The expression relationship between the sets of coexpressed genes is defined by the expression relationship between the skeletons of these sets, where this skeleton represents the coexpressed genes with a well-defined nonlinear expression relationship with the skeleton of the other sets. As a result, we can study the nonlinear expression relationships between a target gene and other sets of coexpressed genes, or start the study from the skeleton of the sets, to study the complex relationships of activation and deactivation between the sets of coexpressed genes that carry out the different cellular processes present in the expression experiments.

  4. Spinophilin expression determines cellular growth, cancer stemness and 5-flourouracil resistance in colorectal cancer

    PubMed Central

    Schwarzenbacher, Daniela; Deutsch, Alexander; Perakis, Samantha; Ling, Hui; Ivan, Cristina; Calin, George Adrian; Rinner, Beate; Gerger, Armin; Pichler, Martin

    2014-01-01

    The putative tumor suppressor gene spinophilin has been involved in cancer progression in several types of cancer. In this study, we explored the prognostic value of spinophilin expression in 162 colon adenocarcinoma patients. In addition, we generated stably expressing spinophilin-directed shRNA CRC cell lines and studied the influence of spinophilin expression on cellular phenotypes and molecular interactions. We independently confirmed that low spinophilin expression levels are associated with poor prognosis in CRC patients (p = 0.038). A reduction of spinophilin levels in p53 wild-type HCT116 and p53-mutated Caco-2 cells led to increased cellular growth rates and anchorage-independent growth (p<0.05). At molecular level, reduced spinophilin levels increased the expression of the transcription factor E2F-1. In addition, we observed an increased formation of tumor spheres, increased number of CD133 positive cells and an increased resistance to 5-flourouracil (p<0.05). Finally, treatment with the de-methylating agent 5-aza-dC increased spinophilin expression in CRC cells (p<0.05), corroborated by a correlation of spinophilin expression and extent of methylated CpG sites in the gene promoter region (p<0.001). In conclusion, gain of aggressive biological properties of CRC cells including cellular growth, cancer stem cell features and 5-flourouracil resistance partly explains the role of spinophilin in CRC. PMID:25261368

  5. A cellular memory module conveys epigenetic inheritance of hedgehog expression during Drosophila wing imaginal disc development.

    PubMed

    Maurange, Cédric; Paro, Renato

    2002-10-15

    In Drosophila, the Trithorax-group (trxG) and Polycomb-group (PcG) proteins interact with chromosomal elements, termed Cellular Memory Modules (CMMs). By modifying chromatin, this ensures a stable heritable maintenance of the transcriptional state of developmental regulators, like the homeotic genes, that is defined embryonically. We asked whether such CMMs could also control expression of genes involved in patterning imaginal discs during larval development. Our results demonstrate that expression of the hedgehog gene, once activated, is maintained by a CMM. In addition, our experiments indicate that the switching of such CMMs to an active state during larval stages, in contrast to embryonic stages, may require specific trans-activators. Our results suggest that the patterning of cells in particular developmental fields in the imaginal discs does not only rely on external cues from morphogens, but also depends on the previous history of the cells, as the control by CMMs ensures a preformatted gene expression pattern.

  6. Gene markers of cellular aging in human multipotent stromal cells in culture

    PubMed Central

    2014-01-01

    Introduction Human multipotent stromal cells (MSCs) isolated from bone marrow or other tissue sources have great potential to treat a wide range of injuries and disorders in the field of regenerative medicine and tissue engineering. In particular, MSCs have inherent characteristics to suppress the immune system and are being studied in clinical studies to prevent graft-versus-host disease. MSCs can be expanded in vitro and have potential for differentiation into multiple cell lineages. However, the impact of cell passaging on gene expression and function of the cells has not been determined. Methods Commercially available human MSCs derived from bone marrow from six different donors, grown under identical culture conditions and harvested at cell passages 3, 5, and 7, were analyzed with gene-expression profiling by using microarray technology. Results The phenotype of these cells did not change as reported previously; however, a statistical analysis revealed a set of 78 significant genes that were distinguishable in expression between passages 3 and 7. None of these significant genes corresponded to the markers established by the International Society for Cellular Therapy (ISCT) for MSC identification. When the significant gene lists were analyzed through pathway analysis, these genes were involved in the top-scoring networks of cellular growth and proliferation and cellular development. A meta-analysis of the literature for significant genes revealed that the MSCs seem to be undergoing differentiation into a senescent cell type when cultured extensively. Consistent with the differences in gene expression at passage 3 and 7, MSCs exhibited a significantly greater potential for cell division at passage 3 in comparison to passage 7. Conclusions Our results identified specific gene markers that distinguish aging MSCs grown in cell culture. Confirmatory studies are needed to correlate these molecular markers with biologic attributes that may facilitate the development

  7. Direct Introduction of Genes into Rats and Expression of the Genes

    NASA Astrophysics Data System (ADS)

    Benvenisty, Nissim; Reshef, Lea

    1986-12-01

    A method of introducing actively expressed genes into intact mammals is described. DNA precipitated with calcium phosphate has been injected intraperitoneally into newborn rats. The injected genes have been taken up and expressed by the animal tissues. To examine the generality of the method we have injected newborn rats with the chloramphenicol acetyltransferase prokaryotic gene fused with various viral and cellular gene promoters and the gene for hepatitis B surface antigen, and we observed appearance of chloramphenicol acetyltransferase activity and hepatitis B surface antigen in liver and spleen. In addition, administration of genes coding for hormones (insulin or growth hormone) resulted in their expression.

  8. Human Lacrimal Gland Gene Expression

    PubMed Central

    Aakalu, Vinay Kumar; Parameswaran, Sowmya; Maienschein-Cline, Mark; Bahroos, Neil; Shah, Dhara; Ali, Marwan; Krishnakumar, Subramanian

    2017-01-01

    Background The study of human lacrimal gland biology and development is limited. Lacrimal gland tissue is damaged or poorly functional in a number of disease states including dry eye disease. Development of cell based therapies for lacrimal gland diseases requires a better understanding of the gene expression and signaling pathways in lacrimal gland. Differential gene expression analysis between lacrimal gland and other embryologically similar tissues may be helpful in furthering our understanding of lacrimal gland development. Methods We performed global gene expression analysis of human lacrimal gland tissue using Affymetrix ® gene expression arrays. Primary data from our laboratory was compared with datasets available in the NLM GEO database for other surface ectodermal tissues including salivary gland, skin, conjunctiva and corneal epithelium. Results The analysis revealed statistically significant difference in the gene expression of lacrimal gland tissue compared to other ectodermal tissues. The lacrimal gland specific, cell surface secretory protein encoding genes and critical signaling pathways which distinguish lacrimal gland from other ectodermal tissues are described. Conclusions Differential gene expression in human lacrimal gland compared with other ectodermal tissue types revealed interesting patterns which may serve as the basis for future studies in directed differentiation among other areas. PMID:28081151

  9. Identification of human HK genes and gene expression regulation study in cancer from transcriptomics data analysis.

    PubMed

    Chen, Meili; Xiao, Jingfa; Zhang, Zhang; Liu, Jingxing; Wu, Jiayan; Yu, Jun

    2013-01-01

    The regulation of gene expression is essential for eukaryotes, as it drives the processes of cellular differentiation and morphogenesis, leading to the creation of different cell types in multicellular organisms. RNA-Sequencing (RNA-Seq) provides researchers with a powerful toolbox for characterization and quantification of transcriptome. Many different human tissue/cell transcriptome datasets coming from RNA-Seq technology are available on public data resource. The fundamental issue here is how to develop an effective analysis method to estimate expression pattern similarities between different tumor tissues and their corresponding normal tissues. We define the gene expression pattern from three directions: 1) expression breadth, which reflects gene expression on/off status, and mainly concerns ubiquitously expressed genes; 2) low/high or constant/variable expression genes, based on gene expression level and variation; and 3) the regulation of gene expression at the gene structure level. The cluster analysis indicates that gene expression pattern is higher related to physiological condition rather than tissue spatial distance. Two sets of human housekeeping (HK) genes are defined according to cell/tissue types, respectively. To characterize the gene expression pattern in gene expression level and variation, we firstly apply improved K-means algorithm and a gene expression variance model. We find that cancer-associated HK genes (a HK gene is specific in cancer group, while not in normal group) are expressed higher and more variable in cancer condition than in normal condition. Cancer-associated HK genes prefer to AT-rich genes, and they are enriched in cell cycle regulation related functions and constitute some cancer signatures. The expression of large genes is also avoided in cancer group. These studies will help us understand which cell type-specific patterns of gene expression differ among different cell types, and particularly for cancer.

  10. Differential gene expression in anatomical compartments of the human eye

    PubMed Central

    Diehn, Jennifer J; Diehn, Maximilian; Marmor, Michael F; Brown, Patrick O

    2005-01-01

    Background The human eye is composed of multiple compartments, diverse in form, function, and embryologic origin, that work in concert to provide us with our sense of sight. We set out to systematically characterize the global gene expression patterns that specify the distinctive characteristics of the various eye compartments. Results We used DNA microarrays representing approximately 30,000 human genes to analyze gene expression in the cornea, lens, iris, ciliary body, retina, and optic nerve. The distinctive patterns of expression in each compartment could be interpreted in relation to the physiology and cellular composition of each tissue. Notably, the sets of genes selectively expressed in the retina and in the lens were particularly large and diverse. Genes with roles in immune defense, particularly complement components, were expressed at especially high levels in the anterior segment tissues. We also found consistent differences between the gene expression patterns of the macula and peripheral retina, paralleling the differences in cell layer densities between these regions. Based on the hypothesis that genes responsible for diseases that affect a particular eye compartment are likely to be selectively expressed in that compartment, we compared our gene expression signatures with genetic mapping studies to identify candidate genes for diseases affecting the cornea, lens, and retina. Conclusion Through genome-scale gene expression profiling, we were able to discover distinct gene expression 'signatures' for each eye compartment and identified candidate disease genes that can serve as a reference database for investigating the physiology and pathophysiology of the eye. PMID:16168081

  11. Applying Attractor Dynamics to Infer Gene Regulatory Interactions Involved in Cellular Differentiation.

    PubMed

    Ghaffarizadeh, Ahmadreza; Podgorski, Gregory J; Flann, Nicholas S

    2017-02-27

    The dynamics of gene regulatory networks (GRNs) guide cellular differentiation. Determining the ways regulatory genes control expression of their targets is essential to understand and control cellular differentiation. The way a regulatory gene controls its target can be expressed as a gene regulatory function. Manual derivation of these regulatory functions is slow, error-prone and difficult to update as new information arises. Automating this process is a significant challenge and the subject of intensive effort. This work presents a novel approach to discovering biologically plausible gene regulatory interactions that control cellular differentiation. This method integrates known cell type expression data, genetic interactions, and knowledge of the effects of gene knockouts to determine likely GRN regulatory functions. We employ a genetic algorithm to search for candidate GRNs that use a set of transcription factors that control differentiation within a lineage. Nested canalyzing functions are used to constrain the search space to biologically plausible networks. The method identifies an ensemble of GRNs whose dynamics reproduce the gene expression pattern for each cell type within a particular lineage. The method's effectiveness was tested by inferring consensus GRNs for myeloid and pancreatic cell differentiation and comparing the predicted gene regulatory interactions to manually derived interactions. We identified many regulatory interactions reported in the literature and also found differences from published reports. These discrepancies suggest areas for biological studies of myeloid and pancreatic differentiation. We also performed a study that used defined synthetic networks to evaluate the accuracy of the automated search method and found that the search algorithm was able to discover the regulatory interactions in these defined networks with high accuracy. We suggest that the GRN functions derived from the methods described here can be used to fill

  12. Mesenchymal stem cells as cellular vehicles for prodrug gene therapy against tumors.

    PubMed

    Amara, Ikrame; Touati, Walid; Beaune, Philippe; de Waziers, Isabelle

    2014-10-01

    Gene-directed enzyme prodrug therapy (GDEPT) consists of targeted delivery to tumor cells of a suicide gene responsible for the in situ conversion of a prodrug into cytotoxic metabolites. One of the major impediments of GDEPT is to target specifically the tumor cells with the suicide gene. Among gene delivery methods, mesenchymal stem cells (MSCs) have emerged recently as potential cellular vehicles for gene delivery. MSCs are particularly suited for gene transduction. They exhibit remarkable migratory property towards tumors and their metastases and they are weakly immunogenic. This review will summarize the current knowledge about MSCs engineered to express different suicide genes (cytosine deaminase, thymidine kinase, carboxylesterase, cytochrome P450) to elicit a significant antitumor response against brain tumors, ovarian, hepatocellular, pancreatic, renal or medullary thyroid carcinomas, breast or prostate cancer and pulmonary metastases. The potential side effects of these MSC-based tumor therapies will also be considered to highlight certain aspects that need to be improved prior to clinical use.

  13. Monoallelic Gene Expression in Mammals.

    PubMed

    Chess, Andrew

    2016-11-23

    Monoallelic expression not due to cis-regulatory sequence polymorphism poses an intriguing problem in epigenetics because it requires the unequal treatment of two segments of DNA that are present in the same nucleus and that can indeed have absolutely identical sequences. Here, I focus on a few recent developments in the field of monoallelic expression that are of particular interest and raise interesting questions for future work. One development is regarding analyses of imprinted genes, in which recent work suggests the possibility that intriguing networks of imprinted genes exist and are important for genetic and physiological studies. Another issue that has been raised in recent years by a number of publications is the question of how skewed allelic expression should be for it to be designated as monoallelic expression and, further, what methods are appropriate or inappropriate for analyzing genomic data to examine allele-specific expression. Perhaps the most exciting recent development in mammalian monoallelic expression is a clever and carefully executed analysis of genetic diversity of autosomal genes subject to random monoallelic expression (RMAE), which provides compelling evidence for distinct evolutionary forces acting on random monoallelically expressed genes.

  14. Tuning noise in gene expression.

    PubMed

    Tyagi, Sanjay

    2015-05-05

    The relative contribution of promoter architecture and the associated chromatin environment in regulating gene expression noise has remained elusive. In their recent work, Arkin, Schaffer and colleagues (Dey et al, 2015) show that mean expression and noise for a given promoter at different genomic loci are uncorrelated and influenced by the local chromatin environment.

  15. Repression of gene expression by oxidative stress.

    PubMed Central

    Morel, Y; Barouki, R

    1999-01-01

    Gene expression is modulated by both physiological signals (hormones, cytokines, etc.) and environmental stimuli (physical parameters, xenobiotics, etc.). Oxidative stress appears to be a key pleiotropic modulator which may be involved in either pathway. Indeed, reactive oxygen species (ROS) have been described as second messengers for several growth factors and cytokines, but have also been shown to rise following cellular insults such as xenobiotic metabolism or enzymic deficiency. Extensive studies on the induction of stress-response genes by oxidative stress have been reported. In contrast, owing to the historical focus on gene induction, less attention has been paid to gene repression by ROS. However, a growing number of studies have shown that moderate (i.e. non-cytotoxic) oxidative stress specifically down-regulates the expression of various genes. In this review, we describe the alteration of several physiological functions resulting from oxidative-stress-mediated inhibition of gene transcription. We will then focus on the repressive oxidative modulation of various transcription factors elicited by ROS. PMID:10477257

  16. Gene Essentiality Is a Quantitative Property Linked to Cellular Evolvability.

    PubMed

    Liu, Gaowen; Yong, Mei Yun Jacy; Yurieva, Marina; Srinivasan, Kandhadayar Gopalan; Liu, Jaron; Lim, John Soon Yew; Poidinger, Michael; Wright, Graham Daniel; Zolezzi, Francesca; Choi, Hyungwon; Pavelka, Norman; Rancati, Giulia

    2015-12-03

    Gene essentiality is typically determined by assessing the viability of the corresponding mutant cells, but this definition fails to account for the ability of cells to adaptively evolve to genetic perturbations. Here, we performed a stringent screen to assess the degree to which Saccharomyces cerevisiae cells can survive the deletion of ~1,000 individual "essential" genes and found that ~9% of these genetic perturbations could in fact be overcome by adaptive evolution. Our analyses uncovered a genome-wide gradient of gene essentiality, with certain essential cellular functions being more "evolvable" than others. Ploidy changes were prevalent among the evolved mutant strains, and aneuploidy of a specific chromosome was adaptive for a class of evolvable nucleoporin mutants. These data justify a quantitative redefinition of gene essentiality that incorporates both viability and evolvability of the corresponding mutant cells and will enable selection of therapeutic targets associated with lower risk of emergence of drug resistance.

  17. Hepatic expression and cellular distribution of the glucose transporter family

    PubMed Central

    Karim, Sumera; Adams, David H; Lalor, Patricia F

    2012-01-01

    Glucose and other carbohydrates are transported into cells using members of a family of integral membrane glucose transporter (GLUT) molecules. To date 14 members of this family, also called the solute carrier 2A proteins have been identified which are divided on the basis of transport characteristics and sequence similarities into several families (Classes 1 to 3). The expression of these different receptor subtypes varies between different species, tissues and cellular subtypes and each has differential sensitivities to stimuli such as insulin. The liver is a contributor to metabolic carbohydrate homeostasis and is a major site for synthesis, storage and redistribution of carbohydrates. Situations in which the balance of glucose homeostasis is upset such as diabetes or the metabolic syndrome can lead metabolic disturbances that drive chronic organ damage and failure, confirming the importance of understanding the molecular regulation of hepatic glucose homeostasis. There is a considerable literature describing the expression and function of receptors that regulate glucose uptake and release by hepatocytes, the most import cells in glucose regulation and glycogen storage. However there is less appreciation of the roles of GLUTs expressed by non parenchymal cell types within the liver, all of which require carbohydrate to function. A better understanding of the detailed cellular distribution of GLUTs in human liver tissue may shed light on mechanisms underlying disease pathogenesis. This review summarises the available literature on hepatocellular expression of GLUTs in health and disease and highlights areas where further investigation is required. PMID:23239915

  18. Gene expression profiling in male genital lichen sclerosus

    PubMed Central

    Edmonds, Emma; Barton, Geraint; Buisson, Sandrine; Francis, Nick; Gotch, Frances; Game, Laurence; Haddad, Munther; Dinneen, Michael; Bunker, Chris

    2011-01-01

    Male genital lichen sclerosus (MGLSc) has a bimodal distribution in boys and men. It is associated with squamous cell carcinoma (SCC). The pathogenesis of MGLSc is unknown. HPV and autoimmune mechanisms have been mooted. Anti extracellular matrix protein (ECM)1 antibodies have been identified in women with GLSc. The gene expression pattern of LSc is unknown. Using DNA microarrays we studied differences in gene expression in healthy and diseased prepuces obtained at circumcision in adult males with MGLSc (n = 4), paediatric LSc (n = 2) and normal healthy paediatric foreskin (n = 4). In adult samples 51 genes with significantly increased expression and 87 genes with significantly reduced expression were identified; paediatric samples revealed 190 genes with significantly increased expression and 148 genes with significantly reduced expression. Concordance of expression profiles between adult and paediatric samples indicates the same disease process. Functional analysis revealed increased expression in the adult and child MGSLc samples in the immune response/cellular defence gene ontology (GO) category and reduced expression in other categories including genes related to squamous cancer. No specific HPV, autoimmune or squamous carcinogenesis-associated gene expression patterns were found. ECM1 and CABLES1 expression were significantly reduced in paediatric and adult samples respectively. PMID:21718371

  19. Tensor decomposition for multi-tissue gene expression experiments

    PubMed Central

    Hore, Victoria; Viñuela, Ana; Buil, Alfonso; Knight, Julian; McCarthy, Mark I; Small, Kerrin; Marchini, Jonathan

    2016-01-01

    Genome wide association studies of gene expression traits and other cellular phenotypes have been successful in revealing links between genetic variation and biological processes. The majority of discoveries have uncovered cis eQTL effects via mass univariate testing of SNPs against gene expression in single tissues. We present a Bayesian method for multi-tissue experiments focusing on uncovering gene networks linked to genetic variation. Our method decomposes the 3D array (or tensor) of gene expression measurements into a set of latent components. We identify sparse gene networks, which can then be tested for association against genetic variation genome-wide. We apply our method to a dataset of 845 individuals from the TwinsUK cohort with gene expression measured via RNA sequencing in adipose, LCLs and skin. We uncover several gene networks with a genetic basis and clear biological and statistical significance. Extensions of this approach will allow integration of multi-omic, environmental and phenotypic datasets. PMID:27479908

  20. A signature microRNA expression profile for the cellular response to thermal stress

    NASA Astrophysics Data System (ADS)

    Wilmink, Gerald J.; Roth, Caleb C.; Ketchum, Norma; Ibey, Bennett L.; Waterworth, Angela; Suarez, Maria; Roach, William P.

    2009-02-01

    Recently, an extensive layer of intra-cellular signals was discovered that was previously undetected by genetic radar. It is now known that this layer consists primarily of a class of short noncoding RNA species that are referred to as microRNAs (miRNAs). MiRNAs regulate protein synthesis at the post-transcriptional level, and studies have shown that they are involved in many fundamental cellular processes. In this study, we hypothesized that miRNAs may be involved in cellular stress response mechanisms, and that cells exposed to thermal stress may exhibit a signature miRNA expression profile indicative of their functional involvement in such mechanisms. To test our hypothesis, human dermal fibroblasts were exposed to an established hyperthermic protocol, and the ensuing miRNA expression levels were evaluated 4 hr post-exposure using microRNA microarray gene chips. The microarray data shows that 123 miRNAs were differentially expressed in cells exposed to thermal stress. We collectively refer to these miRNAs as thermalregulated microRNAs (TRMs). Since miRNA research is in its infancy, it is interesting to note that only 27 of the 123 TRMs are currently annotated in the Sanger miRNA registry. Prior to publication, we plan to submit the remaining novel 96 miRNA gene sequences for proper naming. Computational and thermodynamic modeling algorithms were employed to identify putative mRNA targets for the TRMs, and these studies predict that TRMs regulate the mRNA expression of various proteins that are involved in the cellular stress response. Future empirical studies will be conducted to validate these theoretical predictions, and to further examine the specific role that TRMs play in the cellular stress response.

  1. Temporal Changes in Gene Expression Profile during Mature Adipocyte Dedifferentiation

    PubMed Central

    Côté, Julie Anne; Guénard, Frédéric; Lessard, Julie; Lapointe, Marc; Biron, Simon

    2017-01-01

    Objective. To characterize changes in gene expression profile during human mature adipocyte dedifferentiation in ceiling culture. Methods. Subcutaneous (SC) and omental (OM) adipose tissue samples were obtained from 4 participants paired for age and BMI. Isolated adipocytes were dedifferentiated in ceiling culture. Gene expression analysis at days 0, 4, 7, and 12 of the cultures was performed using Affymetrix Human Gene 2.0 STvi arrays. Hierarchical clustering according to similarity of expression changes was used to identify overrepresented functions. Results. Four clusters gathered genes with similar expression between day 4 to day 7 but decreasing expression from day 7 to day 12. Most of these genes coded for proteins involved in adipocyte functions (LIPE, PLIN1, DGAT2, PNPLA2, ADIPOQ, CEBPA, LPL, FABP4, SCD, INSR, and LEP). Expression of several genes coding for proteins implicated in cellular proliferation and growth or cell cycle increased significantly from day 7 to day 12 (WNT5A, KITLG, and FGF5). Genes coding for extracellular matrix proteins were differentially expressed between days 0, 4, 7, and 12 (COL1A1, COL1A2, and COL6A3, MMP1, and TGFB1). Conclusion. Dedifferentiation is associated with downregulation of transcripts encoding proteins involved in mature adipocyte functions and upregulation of genes involved in matrix remodeling, cellular development, and cell cycle.

  2. Generally detected genes in comparative transcriptomics in bivalves: toward the identification of molecular markers of cellular stress response.

    PubMed

    Miao, Jingjing; Chi, Luping; Pan, Luqing; Song, Ying

    2015-01-01

    The specificity and representativeness of protein-coding genes identified by transcriptomics as biomarkers for environmental toxicological stress is crucial. We extracted the differential gene expression profile data from 49 published comparative transcriptomic studies of bivalves from January 2004 till November 2014 performed in 15 different bivalve species. Among the studies, 77 protein-coding genes were frequently detected when we use threefold of the average detection frequency as cut-off. Cellular organization and communication, protein and energy metabolism, stress response are the main functional classes of these proteins. We consider if these protein-coding genes represent common cellular stress responses of bivalves.

  3. Differential Gene Expression in Glaucoma

    PubMed Central

    Jakobs, Tatjana C.

    2014-01-01

    In glaucoma, regardless of its etiology, retinal ganglion cells degenerate and eventually die. Although age and elevated intraocular pressure (IOP) are the main risk factors, there are still many mysteries in the pathogenesis of glaucoma. The advent of genome-wide microarray expression screening together with the availability of animal models of the disease has allowed analysis of differential gene expression in all parts of the eye in glaucoma. This review will outline the findings of recent genome-wide expression studies and discuss their commonalities and differences. A common finding was the differential regulation of genes involved in inflammation and immunity, including the complement system and the cytokines transforming growth factor β (TGFβ) and tumor necrosis factor α (TNFα). Other genes of interest have roles in the extracellular matrix, cell–matrix interactions and adhesion, the cell cycle, and the endothelin system. PMID:24985133

  4. Differential gene expression in glaucoma.

    PubMed

    Jakobs, Tatjana C

    2014-07-01

    In glaucoma, regardless of its etiology, retinal ganglion cells degenerate and eventually die. Although age and elevated intraocular pressure (IOP) are the main risk factors, there are still many mysteries in the pathogenesis of glaucoma. The advent of genome-wide microarray expression screening together with the availability of animal models of the disease has allowed analysis of differential gene expression in all parts of the eye in glaucoma. This review will outline the findings of recent genome-wide expression studies and discuss their commonalities and differences. A common finding was the differential regulation of genes involved in inflammation and immunity, including the complement system and the cytokines transforming growth factor β (TGFβ) and tumor necrosis factor α (TNFα). Other genes of interest have roles in the extracellular matrix, cell-matrix interactions and adhesion, the cell cycle, and the endothelin system.

  5. Distribution of cellular HSV-1 receptor expression in human brain.

    PubMed

    Lathe, Richard; Haas, Juergen G

    2016-12-15

    Herpes simplex virus type 1 (HSV-1) is a neurotropic virus linked to a range of acute and chronic neurological disorders affecting distinct regions of the brain. Unusually, HSV-1 entry into cells requires the interaction of viral proteins glycoprotein D (gD) and glycoprotein B (gB) with distinct cellular receptor proteins. Several different gD and gB receptors have been identified, including TNFRSF14/HVEM and PVRL1/nectin 1 as gD receptors and PILRA, MAG, and MYH9 as gB receptors. We investigated the expression of these receptor molecules in different areas of the adult and developing human brain using online transcriptome databases. Whereas all HSV-1 receptors showed distinct expression patterns in different brain areas, the Allan Brain Atlas (ABA) reported increased expression of both gD and gB receptors in the hippocampus. Specifically, for PVRL1, TNFRFS14, and MYH9, the differential z scores for hippocampal expression, a measure of relative levels of increased expression, rose to 2.9, 2.9, and 2.5, respectively, comparable to the z score for the archetypical hippocampus-enriched mineralocorticoid receptor (NR3C2, z = 3.1). These data were confirmed at the Human Brain Transcriptome (HBT) database, but HBT data indicate that MAG expression is also enriched in hippocampus. The HBT database allowed the developmental pattern of expression to be investigated; we report that all HSV1 receptors markedly increase in expression levels between gestation and the postnatal/adult periods. These results suggest that differential receptor expression levels of several HSV-1 gD and gB receptors in the adult hippocampus are likely to underlie the susceptibility of this brain region to HSV-1 infection.

  6. Cellular dissection of the spinal cord motor column by BAC transgenesis and gene trapping in zebrafish.

    PubMed

    Asakawa, Kazuhide; Abe, Gembu; Kawakami, Koichi

    2013-01-01

    Bacterial artificial chromosome (BAC) transgenesis and gene/enhancer trapping are effective approaches for identification of genetically defined neuronal populations in the central nervous system (CNS). Here, we applied these techniques to zebrafish (Danio rerio) in order to obtain insights into the cellular architecture of the axial motor column in vertebrates. First, by using the BAC for the Mnx class homeodomain protein gene mnr2b/mnx2b, we established the mnGFF7 transgenic line expressing the Gal4FF transcriptional activator in a large part of the motor column. Single cell labeling of Gal4FF-expressing cells in the mnGFF7 line enabled a detailed investigation of the morphological characteristics of individual spinal motoneurons, as well as the overall organization of the motor column in a spinal segment. Secondly, from a large-scale gene trap screen, we identified transgenic lines that marked discrete subpopulations of spinal motoneurons with Gal4FF. Molecular characterization of these lines led to the identification of the ADAMTS3 gene, which encodes an evolutionarily conserved ADAMTS family of peptidases and is dynamically expressed in the ventral spinal cord. The transgenic fish established here, along with the identified gene, should facilitate an understanding of the cellular and molecular architecture of the spinal cord motor column and its connection to muscles in vertebrates.

  7. Transgenic Arabidopsis Gene Expression System

    NASA Technical Reports Server (NTRS)

    Ferl, Robert; Paul, Anna-Lisa

    2009-01-01

    The Transgenic Arabidopsis Gene Expression System (TAGES) investigation is one in a pair of investigations that use the Advanced Biological Research System (ABRS) facility. TAGES uses Arabidopsis thaliana, thale cress, with sensor promoter-reporter gene constructs that render the plants as biomonitors (an organism used to determine the quality of the surrounding environment) of their environment using real-time nondestructive Green Fluorescent Protein (GFP) imagery and traditional postflight analyses.

  8. Posttranscriptional Control of Gene Expression in Yeast

    PubMed Central

    McCarthy, John E. G.

    1998-01-01

    Studies of the budding yeast Saccharomyces cerevisiae have greatly advanced our understanding of the posttranscriptional steps of eukaryotic gene expression. Given the wide range of experimental tools applicable to S. cerevisiae and the recent determination of its complete genomic sequence, many of the key challenges of the posttranscriptional control field can be tackled particularly effectively by using this organism. This article reviews the current knowledge of the cellular components and mechanisms related to translation and mRNA decay, with the emphasis on the molecular basis for rate control and gene regulation. Recent progress in characterizing translation factors and their protein-protein and RNA-protein interactions has been rapid. Against the background of a growing body of structural information, the review discusses the thermodynamic and kinetic principles that govern the translation process. As in prokaryotic systems, translational initiation is a key point of control. Modulation of the activities of translational initiation factors imposes global regulation in the cell, while structural features of particular 5′ untranslated regions, such as upstream open reading frames and effector binding sites, allow for gene-specific regulation. Recent data have revealed many new details of the molecular mechanisms involved while providing insight into the functional overlaps and molecular networking that are apparently a key feature of evolving cellular systems. An overall picture of the mechanisms governing mRNA decay has only very recently begun to develop. The latest work has revealed new information about the mRNA decay pathways, the components of the mRNA degradation machinery, and the way in which these might relate to the translation apparatus. Overall, major challenges still to be addressed include the task of relating principles of posttranscriptional control to cellular compartmentalization and polysome structure and the role of molecular channelling

  9. Zipf's Law in Gene Expression

    NASA Astrophysics Data System (ADS)

    Furusawa, Chikara; Kaneko, Kunihiko

    2003-02-01

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

  10. 77 FR 65693 - Cellular, Tissue and Gene Therapies Advisory Committee; Amendment of Notice

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-30

    ... HUMAN SERVICES Food and Drug Administration Cellular, Tissue and Gene Therapies Advisory Committee... Register of October 17, 2012, FDA announced that a meeting of the Cellular, Tissue and Gene Therapies..., Office of Cellular, Tissue and Gene Therapies, Center for Biologics Evaluation and Research, FDA. On...

  11. 78 FR 44133 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-23

    ... HUMAN SERVICES Food and Drug Administration Cellular, Tissue and Gene Therapies Advisory Committee... be open to the public. Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee... on guidance documents issued from the Office of Cellular, Tissue and Gene Therapies, Center...

  12. 76 FR 9028 - Guidance for Industry: Potency Tests for Cellular and Gene Therapy Products; Availability

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-02-16

    ... Industry: Potency Tests for Cellular and Gene Therapy Products'' dated January 2011. The guidance document provides manufacturers of cellular and gene therapy (CGT) products with recommendations for developing... document entitled ``Guidance for Industry: Potency Tests for Cellular and Gene Therapy Products''...

  13. 77 FR 71194 - Draft Guidance for Industry: Preclinical Assessment of Investigational Cellular and Gene Therapy...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-11-29

    ... Investigational Cellular and Gene Therapy Products; Availability AGENCY: Food and Drug Administration, HHS. ACTION... for Biologics Research and Evaluation (CBER), Office of Cellular, Tissue, and Gene Therapies (OCTGT). The product areas covered by this guidance are cellular therapy, gene therapy, therapeutic...

  14. Neighboring Genes Show Correlated Evolution in Gene Expression.

    PubMed

    Ghanbarian, Avazeh T; Hurst, Laurence D

    2015-07-01

    When considering the evolution of a gene's expression profile, we commonly assume that this is unaffected by its genomic neighborhood. This is, however, in contrast to what we know about the lack of autonomy between neighboring genes in gene expression profiles in extant taxa. Indeed, in all eukaryotic genomes genes of similar expression-profile tend to cluster, reflecting chromatin level dynamics. Does it follow that if a gene increases expression in a particular lineage then the genomic neighbors will also increase in their expression or is gene expression evolution autonomous? To address this here we consider evolution of human gene expression since the human-chimp common ancestor, allowing for both variation in estimation of current expression level and error in Bayesian estimation of the ancestral state. We find that in all tissues and both sexes, the change in gene expression of a focal gene on average predicts the change in gene expression of neighbors. The effect is highly pronounced in the immediate vicinity (<100 kb) but extends much further. Sex-specific expression change is also genomically clustered. As genes increasing their expression in humans tend to avoid nuclear lamina domains and be enriched for the gene activator 5-hydroxymethylcytosine, we conclude that, most probably owing to chromatin level control of gene expression, a change in gene expression of one gene likely affects the expression evolution of neighbors, what we term expression piggybacking, an analog of hitchhiking.

  15. Hyperbaric oxygen treatment induces antioxidant gene expression.

    PubMed

    Godman, Cassandra A; Joshi, Rashmi; Giardina, Charles; Perdrizet, George; Hightower, Lawrence E

    2010-06-01

    Although the underlying molecular causes of aging are not entirely clear, hormetic agents like exercise, heat, and calorie restriction may generate a mild pro-oxidant stress that induces cell protective responses to promote healthy aging. As an individual ages, many cellular and physiological processes decline, including wound healing and reparative angiogenesis. This is particularly critical in patients with chronic non-healing wounds who tend to be older. We are interested in the potential beneficial effects of hyperbaric oxygen as a mild hormetic stress on human microvascular endothelial cells. We analyzed global gene expression changes in human endothelial cells following a hyperbaric exposure comparable to a clinical treatment. Our analysis revealed an upregulation of antioxidant, cytoprotective, and immediate early genes. This increase coincided with an increased resistance to a lethal oxidative stress. Our data indicate that hyperbaric oxygen can induce protection against oxidative insults in endothelial cells and may provide an easily administered hormetic treatment to help promote healthy aging.

  16. Expression of virus-encoded proteinases: functional and structural similarities with cellular enzymes.

    PubMed Central

    Dougherty, W G; Semler, B L

    1993-01-01

    Many viruses express their genome, or part of their genome, initially as a polyprotein precursor that undergoes proteolytic processing. Molecular genetic analyses of viral gene expression have revealed that many of these processing events are mediated by virus-encoded proteinases. Biochemical activity studies and structural analyses of these viral enzymes reveal that they have remarkable similarities to cellular proteinases. However, the viral proteinases have evolved unique features that permit them to function in a cellular environment. In this article, the current status of plant and animal virus proteinases is described along with their role in the viral replication cycle. The reactions catalyzed by viral proteinases are not simple enzyme-substrate interactions; rather, the processing steps are highly regulated, are coordinated with other viral processes, and frequently involve the participation of other factors. Images PMID:8302216

  17. HTLV-1 p30II: selective repressor of gene expression.

    PubMed

    Green, Patrick L

    2004-11-24

    Human T-lymphotropic virus type-1 (HTLV-1) is a complex retrovirus that causes adult T-cell leukemia/lymphoma (ATL) and is implicated in a variety of lymphocyte-mediated disorders. HTLV-1 pX ORF II encodes two proteins, p13II and p30II whose roles are beginning to be defined in the virus life cycle. Previous studies indicate the importance of these viral proteins in the ability of the virus to maintain viral loads and persist in an animal model of HTLV-1 infection. Intriguing new studies indicate that p30II is a multifunctional regulator that differentially modulates CREB and Tax-responsive element-mediated transcription through its interaction with CREB-binding protein (CBP)/p300 and specifically binds and represses tax/rex mRNA nuclear export. A new study characterized the role of p30II in regulation of cellular gene expression using comprehensive human gene arrays. Interestingly, p30II is an overall repressor of cellular gene expression, while selectively favoring the expression of regulatory gene pathways important to T lymphocytes. These new findings suggest that HTLV-1, which is associated with lymphoproliferative diseases, uses p30II to selectively repress cellular and viral gene expression to favor the survival of cellular targets ultimately resulting in leukemogenesis.

  18. Genome Wide Expression Profiling during Spinal Cord Regeneration Identifies Comprehensive Cellular Responses in Zebrafish

    PubMed Central

    Hui, Subhra Prakash; Sengupta, Dhriti; Lee, Serene Gek Ping; Sen, Triparna; Kundu, Sudip; Mathavan, Sinnakaruppan; Ghosh, Sukla

    2014-01-01

    Background Among the vertebrates, teleost and urodele amphibians are capable of regenerating their central nervous system. We have used zebrafish as a model to study spinal cord injury and regeneration. Relatively little is known about the molecular mechanisms underlying spinal cord regeneration and information based on high density oligonucleotide microarray was not available. We have used a high density microarray to profile the temporal transcriptome dynamics during the entire phenomenon. Results A total of 3842 genes expressed differentially with significant fold changes during spinal cord regeneration. Cluster analysis revealed event specific dynamic expression of genes related to inflammation, cell death, cell migration, cell proliferation, neurogenesis, neural patterning and axonal regrowth. Spatio-temporal analysis of stat3 expression suggested its possible function in controlling inflammation and cell proliferation. Genes involved in neurogenesis and their dorso-ventral patterning (sox2 and dbx2) are differentially expressed. Injury induced cell proliferation is controlled by many cell cycle regulators and some are commonly expressed in regenerating fin, heart and retina. Expression pattern of certain pathway genes are identified for the first time during regeneration of spinal cord. Several genes involved in PNS regeneration in mammals like stat3, socs3, atf3, mmp9 and sox11 are upregulated in zebrafish SCI thus creating PNS like environment after injury. Conclusion Our study provides a comprehensive genetic blue print of diverse cellular response(s) during regeneration of zebrafish spinal cord. The data highlights the importance of different event specific gene expression that could be better understood and manipulated further to induce successful regeneration in mammals. PMID:24465396

  19. Targeted cellular process profiling approach for uterine leiomyoma using cDNA microarray, proteomics and gene ontology analysis

    PubMed Central

    Ahn, Woong Shick; Kim, Ko-Woon; Bae, Su Mi; Yoon, Joo Hee; Lee, Joon Mo; Namkoong, Sung Eun; Kim, Jin Hong; Kim, Chong Kook; Lee, Young Joo; Kim, Yong-Wan

    2003-01-01

    This study utilized both cDNA microarray and two-dimensional protein gel electrophoresis technology to investigate the multiple interactions of genes and proteins involved in uterine leiomyoma pathophysiology. Also, the gene ontology analysis was used to systematically characterize the global expression profiles at cellular process levels. We profiled differentially expressed transcriptome and proteome in six-paired leiomyoma and normal myometrium. Screening up to 17 000 genes identified 21 upregulated and 50 downregulated genes. The gene-expression profiles were classified into mutually dependent 420 functional sets, resulting in 611 cellular processes according to the gene ontology. Also, protein analysis using two-dimensional gel electrophoresis identified 33 proteins (17 upregulated and 16 downregulated) of more than 500 total spots, which was classified into 302 cellular processes. Of these functional profilings, downregulations of transcriptomes and proteoms were shown in cell adhesion, cell motility, organogenesis, enzyme regulator, structural molecule activity and response to external stimulus functional activities that are supposed to play important roles in pathophysiology. In contrast, the upregulation was only shown in nucleic acid-binding activity. Taken together, potentially significant pathogenetic cellular processes were identified and showed that the downregulated functional profiling has a significant impact on the discovery of pathogenic pathway in leiomyoma. Also, the gene ontology analysis can overcome the complexity of expression profiles of cDNA microarray and two-dimensional protein analysis via its cellular process-level approach. Therefore, a valuable prognostic candidate gene with relevance to disease-specific pathogenesis can be found at cellular process levels. PMID:14748746

  20. Regulation of ABO gene expression.

    PubMed

    Kominato, Yoshihiko; Hata, Yukiko; Matsui, Kazuhiro; Takizawa, Hisao

    2005-07-01

    The ABO blood group system is important in blood transfusions and in identifying individuals during criminal investigations. Two carbohydrate antigens, the A and B antigens, and their antibodies constitute this system. Although biochemical and molecular genetic studies have demonstrated the molecular basis of the histo-blood group ABO system, some aspects remain to be elucidated. To explain the molecular basis of how the ABO genes are controlled in cell type-specific expression, during normal cell differentiation, and in cancer cells with invasive and metastatic potential that lack A/B antigens, it is essential to understand the regulatory mechanism of ABO gene transcription. We review the transcriptional regulation of the ABO gene, including positive and negative elements in the upstream region of the gene, and draw some inferences that help to explain the phenomena described above.

  1. Gene expression profile of pulpitis

    PubMed Central

    Galicia, Johnah C.; Henson, Brett R.; Parker, Joel S.; Khan, Asma A.

    2016-01-01

    The cost, prevalence and pain associated with endodontic disease necessitate an understanding of the fundamental molecular aspects of its pathogenesis. This study was aimed to identify the genetic contributors to pulpal pain and inflammation. Inflamed pulps were collected from patients diagnosed with irreversible pulpitis (n=20). Normal pulps from teeth extracted for various reasons served as controls (n=20). Pain level was assessed using a visual analog scale (VAS). Genome-wide microarray analysis was performed using Affymetrix GeneTitan Multichannel Instrument. The difference in gene expression levels were determined by the Significance Analysis of Microarray program using a false discovery rate (q-value) of 5%. Genes involved in immune response, cytokine-cytokine receptor interaction and signaling, integrin cell surface interactions, and others were expressed at relatively higher levels in the in the pulpitis group. Moreover, several genes known to modulate pain and inflammation showed differential expression in asymptomatic and mild pain patients (≥30mm on VAS) compared to those with moderate to severe pain. This exploratory study provides a molecular basis for the clinical diagnosis of pulpitis. With an enhanced understanding of pulpal inflammation, future studies on treatment and management of pulpitis and on pain associated with it can have a biological reference to bridge treatment strategies with pulpal biology. PMID:27052691

  2. Gene expression profile of pulpitis.

    PubMed

    Galicia, J C; Henson, B R; Parker, J S; Khan, A A

    2016-06-01

    The cost, prevalence and pain associated with endodontic disease necessitate an understanding of the fundamental molecular aspects of its pathogenesis. This study was aimed to identify the genetic contributors to pulpal pain and inflammation. Inflamed pulps were collected from patients diagnosed with irreversible pulpitis (n=20). Normal pulps from teeth extracted for various reasons served as controls (n=20). Pain level was assessed using a visual analog scale (VAS). Genome-wide microarray analysis was performed using Affymetrix GeneTitan Multichannel Instrument. The difference in gene expression levels were determined by the significance analysis of microarray program using a false discovery rate (q-value) of 5%. Genes involved in immune response, cytokine-cytokine receptor interaction and signaling, integrin cell surface interactions, and others were expressed at relatively higher levels in the pulpitis group. Moreover, several genes known to modulate pain and inflammation showed differential expression in asymptomatic and mild pain patients (⩾30 mm on VAS) compared with those with moderate to severe pain. This exploratory study provides a molecular basis for the clinical diagnosis of pulpitis. With an enhanced understanding of pulpal inflammation, future studies on treatment and management of pulpitis and on pain associated with it can have a biological reference to bridge treatment strategies with pulpal biology.

  3. Genetic screening of new genes responsible for cellular adaptation to hypoxia using a genome-wide shRNA library.

    PubMed

    Yoshino, Seiko; Hara, Toshiro; Weng, Jane S; Takahashi, Yuka; Seiki, Motoharu; Sakamoto, Takeharu

    2012-01-01

    Oxygen is a vital requirement for multi-cellular organisms to generate energy and cells have developed multiple compensatory mechanisms to adapt to stressful hypoxic conditions. Such adaptive mechanisms are intricately interconnected with other signaling pathways that regulate cellular functions such as cell growth. However, our understanding of the overall system governing the cellular response to the availability of oxygen remains limited. To identify new genes involved in the response to hypoxic stress, we have performed a genome-wide gene knockdown analysis in human lung carcinoma PC8 cells using an shRNA library carried by a lentiviral vector. The knockdown analysis was performed under both normoxic and hypoxic conditions to identify shRNA sequences enriched or lost in the resulting selected cell populations. Consequently, we identified 56 candidate genes that might contribute to the cellular response to hypoxia. Subsequent individual knockdown of each gene demonstrated that 13 of these have a significant effect upon oxygen-sensitive cell growth. The identification of BCL2L1, which encodes a Bcl-2 family protein that plays a role in cell survival by preventing apoptosis, validates the successful design of our screen. The other selected genes have not previously been directly implicated in the cellular response to hypoxia. Interestingly, hypoxia did not directly enhance the expression of any of the identified genes, suggesting that we have identified a new class of genes that have been missed by conventional gene expression analyses to identify hypoxia response genes. Thus, our genetic screening method using a genome-wide shRNA library and the newly-identified genes represent useful tools to analyze the cellular systems that respond to hypoxic stress.

  4. Identification of Cellular Genes Targeted by KSHV-Encoded MicroRNAs

    PubMed Central

    Samols, Mark A; Skalsky, Rebecca L; Maldonado, Ann M; Riva, Alberto; Lopez, M. Cecilia; Baker, Henry V; Renne, Rolf

    2007-01-01

    MicroRNAs (miRNAs) are 19 to 23 nucleotide–long RNAs that post-transcriptionally regulate gene expression. Human cells express several hundred miRNAs which regulate important biological pathways such as development, proliferation, and apoptosis. Recently, 12 miRNA genes have been identified within the genome of Kaposi sarcoma–associated herpesvirus; however, their functions are still unknown. To identify host cellular genes that may be targeted by these novel viral regulators, we performed gene expression profiling in cells stably expressing KSHV-encoded miRNAs. Data analysis revealed a set of 81 genes whose expression was significantly changed in the presence of miRNAs. While the majority of changes were below 2-fold, eight genes were down-regulated between 4- and 20-fold. We confirmed miRNA-dependent regulation for three of these genes and found that protein levels of thrombospondin 1 (THBS1) were decreased >10-fold. THBS1 has previously been reported to be down-regulated in Kaposi sarcoma lesions and has known activity as a strong tumor suppressor and anti-angiogenic factor, exerting its anti-angiogenic effect in part by activating the latent form of TGF-β. We show that reduced THBS1 expression in the presence of viral miRNAs translates into decreased TGF-β activity. These data suggest that KSHV-encoded miRNAs may contribute directly to pathogenesis by down-regulation of THBS1, a major regulator of cell adhesion, migration, and angiogenesis. PMID:17500590

  5. Expressing genes do not forget their LINEs: transposable elements and gene expression.

    PubMed

    Kines, Kristine J; Belancio, Victoria P

    2012-01-01

    Historically the accumulated mass of mammalian transposable elements (TEs), particularly those located within gene boundaries, was viewed as a genetic burden potentially detrimental to the genomic landscape. This notion has been strengthened by the discovery that transposable sequences can alter the architecture of the transcriptome, not only through insertion, but also long after the integration process is completed. Insertions previously considered harmless are now known to impact the expression of host genes via modification of the transcript quality or quantity, transcriptional interference, or by the control of pathways that affect the mRNA life-cycle. Conversely, several examples of the evolutionary advantageous impact of TEs on the host gene structure that diversified the cellular transcriptome are reported. TE-induced changes in gene expression can be tissue- or disease-specific, raising the possibility that the impact of TE sequences may vary during development, among normal cell types, and between normal and disease-affected tissues. The understanding of the rules and abundance of TE-interference with gene expression is in its infancy, and its contribution to human disease and/or evolution remains largely unexplored.

  6. Systems biology of lupus: mapping the impact of genomic and environmental factors on gene expression signatures, cellular signaling, metabolic pathways, hormonal and cytokine imbalance, and selecting targets for treatment.

    PubMed

    Perl, Andras

    2010-02-01

    Systemic lupus erythematosus (SLE) is characterized by the dysfunction of T cells, B cells, and dendritic cells, the release of pro-inflammatory nuclear materials from necrotic cells, and the formation of antinuclear antibodies (ANA) and immune complexes of ANA with DNA, RNA, and nuclear proteins. Activation of the mammalian target of rapamycin (mTOR) has recently emerged as a key factor in abnormal activation of T and B cells in SLE. In T cells, increased production of nitric oxide and mitochondrial hyperpolarization (MHP) were identified as metabolic checkpoints upstream of mTOR activation. mTOR controls the expression T-cell receptor-associated signaling proteins CD4 and CD3zeta through increased expression of the endosome recycling regulator Rab5 and HRES-1/Rab4 genes, enhances Ca2+ fluxing and skews the expression of tyrosine kinases both in T and B cells, and blocks the expression of Foxp3 and the generation of regulatory T cells. MHP, increased activity of mTOR, Rab GTPases, and Syk kinases, and enhanced Ca2+ flux have emerged as common T and B cell biomarkers and targets for treatment in SLE.

  7. Gene Expressions for Signal Transduction under Acidic Conditions

    PubMed Central

    Fukamachi, Toshihiko; Ikeda, Syunsuke; Wang, Xin; Saito, Hiromi; Tagawa, Masatoshi; Kobayashi, Hiroshi

    2013-01-01

    Although it is now well known that some diseased areas, such as cancer nests, inflammation loci, and infarction areas, are acidified, little is known about cellular signal transduction, gene expression, and cellular functions under acidic conditions. Our group showed that different signal proteins were activated under acidic conditions compared with those observed in a typical medium of around pH 7.4 that has been used until now. Investigations of gene expression under acidic conditions may be crucial to our understanding of signal transduction in acidic diseased areas. In this study, we investigated gene expression in mesothelioma cells cultured at an acidic pH using a DNA microarray technique. After 24 h culture at pH 6.7, expressions of 379 genes were increased more than twofold compared with those in cells cultured at pH 7.5. Genes encoding receptors, signal proteins including transcription factors, and cytokines including growth factors numbered 35, 32, and 17 among the 379 genes, respectively. Since the functions of 78 genes are unknown, it can be argued that cells may have other genes for signaling under acidic conditions. The expressions of 37 of the 379 genes were observed to increase after as little as 2 h. After 24 h culture at pH 6.7, expressions of 412 genes were repressed more than twofold compared with those in cells cultured at pH 7.5, and the 412 genes contained 35, 76, and 7 genes encoding receptors, signal proteins including transcription factors, and cytokines including growth factors, respectively. These results suggest that the signal pathways in acidic diseased areas are different, at least in part, from those examined with cells cultured at a pH of around 7.4. PMID:24705103

  8. The gsdf gene locus harbors evolutionary conserved and clustered genes preferentially expressed in fish previtellogenic oocytes.

    PubMed

    Gautier, Aude; Le Gac, Florence; Lareyre, Jean-Jacques

    2011-02-01

    The gonadal soma-derived factor (GSDF) belongs to the transforming growth factor-β superfamily and is conserved in teleostean fish species. Gsdf is specifically expressed in the gonads, and gene expression is restricted to the granulosa and Sertoli cells in trout and medaka. The gsdf gene expression is correlated to early testis differentiation in medaka and was shown to stimulate primordial germ cell and spermatogonia proliferation in trout. In the present study, we show that the gsdf gene localizes to a syntenic chromosomal fragment conserved among vertebrates although no gsdf-related gene is detected on the corresponding genomic region in tetrapods. We demonstrate using quantitative RT-PCR that most of the genes localized in the synteny are specifically expressed in medaka gonads. Gsdf is the only gene of the synteny with a much higher expression in the testis compared to the ovary. In contrast, gene expression pattern analysis of the gsdf surrounding genes (nup54, aff1, klhl8, sdad1, and ptpn13) indicates that these genes are preferentially expressed in the female gonads. The tissue distribution of these genes is highly similar in medaka and zebrafish, two teleostean species that have diverged more than 110 million years ago. The cellular localization of these genes was determined in medaka gonads using the whole-mount in situ hybridization technique. We confirm that gsdf gene expression is restricted to Sertoli and granulosa cells in contact with the premeiotic and meiotic cells. The nup54 gene is expressed in spermatocytes and previtellogenic oocytes. Transcripts corresponding to the ovary-specific genes (aff1, klhl8, and sdad1) are detected only in previtellogenic oocytes. No expression was detected in the gonocytes in 10 dpf embryos. In conclusion, we show that the gsdf gene localizes to a syntenic chromosomal fragment harboring evolutionary conserved genes in vertebrates. These genes are preferentially expressed in previtelloogenic oocytes, and thus, they

  9. Gene expression throughout a vertebrate's embryogenesis

    PubMed Central

    2011-01-01

    Background Describing the patterns of gene expression during embryonic development has broadened our understanding of the processes and patterns that define morphogenesis. Yet gene expression patterns have not been described throughout vertebrate embryogenesis. This study presents statistical analyses of gene expression during all 40 developmental stages in the teleost Fundulus heteroclitus using four biological replicates per stage. Results Patterns of gene expression for 7,000 genes appear to be important as they recapitulate developmental timing. Among the 45% of genes with significant expression differences between pairs of temporally adjacent stages, significant differences in gene expression vary from as few as five to more than 660. Five adjacent stages have disproportionately more significant changes in gene expression (> 200 genes) relative to other stages: four to eight and eight to sixteen cell stages, onset of circulation, pre and post-hatch, and during complete yolk absorption. The fewest differences among adjacent stages occur during gastrulation. Yet, at stage 16, (pre-mid-gastrulation) the largest number of genes has peak expression. This stage has an over representation of genes in oxidative respiration and protein expression (ribosomes, translational genes and proteases). Unexpectedly, among all ribosomal genes, both strong positive and negative correlations occur. Similar correlated patterns of expression occur among all significant genes. Conclusions These data provide statistical support for the temporal dynamics of developmental gene expression during all stages of vertebrate development. PMID:21356103

  10. Unstable Expression of Commonly Used Reference Genes in Rat Pancreatic Islets Early after Isolation Affects Results of Gene Expression Studies.

    PubMed

    Kosinová, Lucie; Cahová, Monika; Fábryová, Eva; Týcová, Irena; Koblas, Tomáš; Leontovyč, Ivan; Saudek, František; Kříž, Jan

    2016-01-01

    The use of RT-qPCR provides a powerful tool for gene expression studies; however, the proper interpretation of the obtained data is crucially dependent on accurate normalization based on stable reference genes. Recently, strong evidence has been shown indicating that the expression of many commonly used reference genes may vary significantly due to diverse experimental conditions. The isolation of pancreatic islets is a complicated procedure which creates severe mechanical and metabolic stress leading possibly to cellular damage and alteration of gene expression. Despite of this, freshly isolated islets frequently serve as a control in various gene expression and intervention studies. The aim of our study was to determine expression of 16 candidate reference genes and one gene of interest (F3) in isolated rat pancreatic islets during short-term cultivation in order to find a suitable endogenous control for gene expression studies. We compared the expression stability of the most commonly used reference genes and evaluated the reliability of relative and absolute quantification using RT-qPCR during 0-120 hrs after isolation. In freshly isolated islets, the expression of all tested genes was markedly depressed and it increased several times throughout the first 48 hrs of cultivation. We observed significant variability among samples at 0 and 24 hrs but substantial stabilization from 48 hrs onwards. During the first 48 hrs, relative quantification failed to reflect the real changes in respective mRNA concentrations while in the interval 48-120 hrs, the relative expression generally paralleled the results determined by absolute quantification. Thus, our data call into question the suitability of relative quantification for gene expression analysis in pancreatic islets during the first 48 hrs of cultivation, as the results may be significantly affected by unstable expression of reference genes. However, this method could provide reliable information from 48 hrs onwards.

  11. The Gene Expression Omnibus Database.

    PubMed

    Clough, Emily; Barrett, Tanya

    2016-01-01

    The Gene Expression Omnibus (GEO) database is an international public repository that archives and freely distributes high-throughput gene expression and other functional genomics data sets. Created in 2000 as a worldwide resource for gene expression studies, GEO has evolved with rapidly changing technologies and now accepts high-throughput data for many other data applications, including those that examine genome methylation, chromatin structure, and genome-protein interactions. GEO supports community-derived reporting standards that specify provision of several critical study elements including raw data, processed data, and descriptive metadata. The database not only provides access to data for tens of thousands of studies, but also offers various Web-based tools and strategies that enable users to locate data relevant to their specific interests, as well as to visualize and analyze the data. This chapter includes detailed descriptions of methods to query and download GEO data and use the analysis and visualization tools. The GEO homepage is at http://www.ncbi.nlm.nih.gov/geo/.

  12. Classification of genes based on gene expression analysis

    SciTech Connect

    Angelova, M. Myers, C. Faith, J.

    2008-05-15

    Systems biology and bioinformatics are now major fields for productive research. DNA microarrays and other array technologies and genome sequencing have advanced to the point that it is now possible to monitor gene expression on a genomic scale. Gene expression analysis is discussed and some important clustering techniques are considered. The patterns identified in the data suggest similarities in the gene behavior, which provides useful information for the gene functionalities. We discuss measures for investigating the homogeneity of gene expression data in order to optimize the clustering process. We contribute to the knowledge of functional roles and regulation of E. coli genes by proposing a classification of these genes based on consistently correlated genes in expression data and similarities of gene expression patterns. A new visualization tool for targeted projection pursuit and dimensionality reduction of gene expression data is demonstrated.

  13. Cellular retinol binding protein 1 could be a tumor suppressor gene in cervical cancer

    PubMed Central

    Mendoza-Rodriguez, Mónica; Arreola, Hugo; Valdivia, Alejandra; Peralta, Raúl; Serna, Humberto; Villegas, Vanessa; Romero, Pablo; Alvarado-Hernández, Beatriz; Paniagua, Lucero; Marrero-Rodríguez, Daniel; Meraz, Marco A; Salcedo, Mauricio

    2013-01-01

    Aims: Cervical Cancer (CC) is one of the most important health problems in women. It frequently presents genetic changes at chromosome region 3q21. This region contains the Cellular Retinol Binding Protein 1 gene (CRBP1) which has been implicated as an important element in the development of other types of cancer. The main goal of the present work was to determine the molecular alterations of CRBP1 and its relationship to CC. Methods: To determine the molecular alterations of CRBP1 gene in CC; twenty-six CC and twenty-six healthy cervix samples were evaluated for: 1) Copy number gain by real-time PCR analysis, 2) expression levels by an immunohistochemistry assay on tissue microarray, and 3) the methylation status of the CRBP1 promoter region. Results: The increase in CRBP1 copy number was observed in 10 out of the 26 CC samples analyzed, while healthy cervices samples showed no changes in the copy number. In addition, there was a lack of expression of the CRBP1 gene in an important number of the CC samples (17/26), and the CRBP1 gene promoter was methylated in 15/26 of the CC samples. Interestingly, there was a significant association between the lack of expression of the CRBP1 gene and its methylation status. Conclusions: The data indicates that, both activating and inactivating changes in the CRBP1 gene could be significant events in the development and progression of CC, and the lack of expression of the CRBP1 protein could be related with to the development of CC. We believe that there is enough evidence to consider to CRBP1 gene as a tumor suppressor gene for CC. PMID:24040446

  14. High expression of cellular retinol binding protein-1 in lung adenocarcinoma is associated with poor prognosis

    PubMed Central

    Doldo, Elena; Costanza, Gaetana; Ferlosio, Amedeo; Pompeo, Eugenio; Agostinelli, Sara; Bellezza, Guido; Mazzaglia, Donatella; Giunta, Alessandro; Sidoni, Angelo; Orlandi, Augusto

    2015-01-01

    Purpose Adenocarcinoma, the most common non-small cell lung cancer is a leading cause of death worldwide, with a low overall survival (OS) despite increasing attempts to achieve an early diagnosis and accomplish surgical and multimodality treatment strategies. Cellular retinol binding protein-1 (CRBP-1) regulates retinol bioavailability and cell differentiation, but its role in lung cancerogenesis remains uncertain. Experimental design CRBP-1 expression, clinical outcome and other prognostic factors were investigated in 167 lung adenocarcinoma patients. CRBP-1 expression was evaluated by immunohistochemistry of tissue microarray sections, gene copy number analysis and tumor methylation specific PCR. Effects of CRBP-1 expression on proliferation/apoptosis gene array, protein and transcripts were investigated in transfected A549 lung adenocarcinoma cells. Results CRBP-1High expression was observed in 62.3% of adenocarcinomas and correlated with increased tumor grade and reduced OS as an independent prognostic factor. CRBP-1 gene copy gain also associated with tumor CRBP-1High status and dedifferentiation. CRBP-1-transfected (CRBP-1+) A549 grew more than CRBP-1− A549 cells. At >1μM concentrations, all trans-retinoic acid and retinol reduced viability more in CRBP-1+ than in CRBP-1− A549 cells. CRBP-1+ A549 cells showed up-regulated RARα/ RXRα and proliferative and transcriptional genes including pAkt, pEGFR, pErk1/2, creb1 and c-jun, whereas RARβ and p53 were strongly down-regulated; pAkt/pErk/ pEGFR inhibitors counteracted proliferative advantage and increased RARα/RXRα, c-jun and CD44 expression in CRBP-1+ A549 cells. Conclusion CRBP-1High expression in lung adenocarcinoma correlated with increased tumor grade and reduced OS, likely through increased Akt/Erk/EGFR-mediated cell proliferation and differentiation. CRBP-1High expression can be considered an additional marker of poor prognosis in lung adenocarcinoma patients. PMID:26807202

  15. Clustering Algorithms: Their Application to Gene Expression Data

    PubMed Central

    Oyelade, Jelili; Isewon, Itunuoluwa; Oladipupo, Funke; Aromolaran, Olufemi; Uwoghiren, Efosa; Ameh, Faridah; Achas, Moses; Adebiyi, Ezekiel

    2016-01-01

    Gene expression data hide vital information required to understand the biological process that takes place in a particular organism in relation to its environment. Deciphering the hidden patterns in gene expression data proffers a prodigious preference to strengthen the understanding of functional genomics. The complexity of biological networks and the volume of genes present increase the challenges of comprehending and interpretation of the resulting mass of data, which consists of millions of measurements; these data also inhibit vagueness, imprecision, and noise. Therefore, the use of clustering techniques is a first step toward addressing these challenges, which is essential in the data mining process to reveal natural structures and identify interesting patterns in the underlying data. The clustering of gene expression data has been proven to be useful in making known the natural structure inherent in gene expression data, understanding gene functions, cellular processes, and subtypes of cells, mining useful information from noisy data, and understanding gene regulation. The other benefit of clustering gene expression data is the identification of homology, which is very important in vaccine design. This review examines the various clustering algorithms applicable to the gene expression data in order to discover and provide useful knowledge of the appropriate clustering technique that will guarantee stability and high degree of accuracy in its analysis procedure. PMID:27932867

  16. Expression profiles of subtracted mRNAs during cellular senescence in human mesenchymal stem cells derived from bone marrow.

    PubMed

    Yoo, Jung Ki; Choi, Seong-jun; Kim, Jin Kyeoung

    2013-05-01

    Cellular senescence is an irreversible cell cycle arrest that limits the replicative lifespan of cells. Senescence suppresses development of tumors by regulating aging factors, such as cyclin dependent kinase inhibitor (CKI) and telomerase. Suppression subtractive hybridization (SSH) was used to identify genes that were differentially expressed between young human mesenchymal stem cells (Y-hMSCs) and senescent human mesenchymal stem cells (S-hMSCs). We selected positive clones that were functionally characterized by referring to public databases using NCBI BLAST tool. This search revealed that 19 genes were downregulated, and 43 genes were upregulated in S-hMSCs relative to Y-hMSCs. Among subtracted clones in Y-hMSCs, most of genes markedly were related to metabolic functions. These genes, PDIA3, WDR1, FSTL1, COPG1, LMAN1, and PDIA6, significantly downregulated. Conversely, genes for subtracted clones in S-hMSCs were mostly associated with cell adhesion. In particular, the expression levels of 9 genes, HSP90B1, EID1, ATP2B4, DDAH1, PRNP, RAB1A, PGS5, TM4SF1 and SSR3, gradually increased during senescence. These genes have not previously been identified as being related to cellular senescence, but they seemed to be potentially affected during cellular senescence.

  17. The Role of Multiple Transcription Factors In Archaeal Gene Expression

    SciTech Connect

    Charles J. Daniels

    2008-09-23

    Since the inception of this research program, the project has focused on two central questions: What is the relationship between the 'eukaryal-like' transcription machinery of archaeal cells and its counterparts in eukaryal cells? And, how does the archaeal cell control gene expression using its mosaic of eukaryal core transcription machinery and its bacterial-like transcription regulatory proteins? During the grant period we have addressed these questions using a variety of in vivo approaches and have sought to specifically define the roles of the multiple TATA binding protein (TBP) and TFIIB-like (TFB) proteins in controlling gene expression in Haloferax volcanii. H. volcanii was initially chosen as a model for the Archaea based on the availability of suitable genetic tools; however, later studies showed that all haloarchaea possessed multiple tbp and tfb genes, which led to the proposal that multiple TBP and TFB proteins may function in a manner similar to alternative sigma factors in bacterial cells. In vivo transcription and promoter analysis established a clear relationship between the promoter requirements of haloarchaeal genes and those of the eukaryal RNA polymerase II promoter. Studies on heat shock gene promoters, and the demonstration that specific tfb genes were induced by heat shock, provided the first indication that TFB proteins may direct expression of specific gene families. The construction of strains lacking tbp or tfb genes, coupled with the finding that many of these genes are differentially expressed under varying growth conditions, provided further support for this model. Genetic tools were also developed that led to the construction of insertion and deletion mutants, and a novel gene expression scheme was designed that allowed the controlled expression of these genes in vivo. More recent studies have used a whole genome array to examine the expression of these genes and we have established a linkage between the expression of specific tfb

  18. Cellular unfolded protein response against viruses used in gene therapy

    PubMed Central

    Sen, Dwaipayan; Balakrishnan, Balaji; Jayandharan, Giridhara R.

    2014-01-01

    Viruses are excellent vehicles for gene therapy due to their natural ability to infect and deliver the cargo to specific tissues with high efficiency. Although such vectors are usually “gutted” and are replication defective, they are subjected to clearance by the host cells by immune recognition and destruction. Unfolded protein response (UPR) is a naturally evolved cyto-protective signaling pathway which is triggered due to endoplasmic reticulum (ER) stress caused by accumulation of unfolded/misfolded proteins in its lumen. The UPR signaling consists of three signaling pathways, namely PKR-like ER kinase, activating transcription factor 6, and inositol-requiring protein-1. Once activated, UPR triggers the production of ER molecular chaperones and stress response proteins to help reduce the protein load within the ER. This occurs by degradation of the misfolded proteins and ensues in the arrest of protein translation machinery. If the burden of protein load in ER is beyond its processing capacity, UPR can activate pro-apoptotic pathways or autophagy leading to cell death. Viruses are naturally evolved in hijacking the host cellular translation machinery to generate a large amount of proteins. This phenomenon disrupts ER homeostasis and leads to ER stress. Alternatively, in the case of gutted vectors used in gene therapy, the excess load of recombinant vectors administered and encountered by the cell can trigger UPR. Thus, in the context of gene therapy, UPR becomes a major roadblock that can potentially trigger inflammatory responses against the vectors and reduce the efficiency of gene transfer. PMID:24904562

  19. Understanding the mechanisms of ATPase beta family genes for cellular thermotolerance in crossbred bulls

    NASA Astrophysics Data System (ADS)

    Deb, Rajib; Sajjanar, Basavaraj; Singh, Umesh; Alex, Rani; Raja, T. V.; Alyethodi, Rafeeque R.; Kumar, Sushil; Sengar, Gyanendra; Sharma, Sheetal; Singh, Rani; Prakash, B.

    2015-12-01

    Na+/K+-ATPase is an integral membrane protein composed of a large catalytic subunit (alpha), a smaller glycoprotein subunit (beta), and gamma subunit. The beta subunit is essential for ion recognition as well as maintenance of the membrane integrity. Present study was aimed to analyze the expression pattern of ATPase beta subunit genes (ATPase B1, ATPase B2, and ATPase B3) among the crossbred bulls under different ambient temperatures (20-44 °C). The present study was also aimed to look into the relationship of HSP70 with the ATPase beta family genes. Our results demonstrated that among beta family genes, transcript abundance of ATPase B1 and ATPase B2 is significantly ( P < 0.05) higher during the thermal stress. Pearson correlation coefficient analysis revealed that the expression of ATPase Β1, ATPase B2, and ATPase B3 is highly correlated ( P < 0.01) with HSP70, representing that the change in the expression pattern of these genes is positive and synergistic. These may provide a foundation for understanding the mechanisms of ATPase beta family genes for cellular thermotolerance in cattle.

  20. Harnessing gene expression networks to prioritize candidate epileptic encephalopathy genes.

    PubMed

    Oliver, Karen L; Lukic, Vesna; Thorne, Natalie P; Berkovic, Samuel F; Scheffer, Ingrid E; Bahlo, Melanie

    2014-01-01

    We apply a novel gene expression network analysis to a cohort of 182 recently reported candidate Epileptic Encephalopathy genes to identify those most likely to be true Epileptic Encephalopathy genes. These candidate genes were identified as having single variants of likely pathogenic significance discovered in a large-scale massively parallel sequencing study. Candidate Epileptic Encephalopathy genes were prioritized according to their co-expression with 29 known Epileptic Encephalopathy genes. We utilized developing brain and adult brain gene expression data from the Allen Human Brain Atlas (AHBA) and compared this to data from Celsius: a large, heterogeneous gene expression data warehouse. We show replicable prioritization results using these three independent gene expression resources, two of which are brain-specific, with small sample size, and the third derived from a heterogeneous collection of tissues with large sample size. Of the nineteen genes that we predicted with the highest likelihood to be true Epileptic Encephalopathy genes, two (GNAO1 and GRIN2B) have recently been independently reported and confirmed. We compare our results to those produced by an established in silico prioritization approach called Endeavour, and finally present gene expression networks for the known and candidate Epileptic Encephalopathy genes. This highlights sub-networks of gene expression, particularly in the network derived from the adult AHBA gene expression dataset. These networks give clues to the likely biological interactions between Epileptic Encephalopathy genes, potentially highlighting underlying mechanisms and avenues for therapeutic targets.

  1. Cell Cycle Programs of Gene Expression Control Morphogenetic Protein Localization

    PubMed Central

    Lord, Matthew; Yang, Melody C.; Mischke, Michelle; Chant, John

    2000-01-01

    Genomic studies in yeast have revealed that one eighth of genes are cell cycle regulated in their expression. Almost without exception, the significance of cell cycle periodic gene expression has not been tested. Given that many such genes are critical to cellular morphogenesis, we wanted to examine the importance of periodic gene expression to this process. The expression profiles of two genes required for the axial pattern of cell division, BUD3 and BUD10/AXL2/SRO4, are strongly cell cycle regulated. BUD3 is expressed close to the onset of mitosis. BUD10 is expressed in late G1. Through promotor-swap experiments, the expression profile of each gene was altered and the consequences examined. We found that an S/G2 pulse of BUD3 expression controls the timing of Bud3p localization, but that this timing is not critical to Bud3p function. In contrast, a G1 pulse of BUD10 expression plays a direct role in Bud10p localization and function. Bud10p, a membrane protein, relies on the polarized secretory machinery specific to G1 to be delivered to its proper location. Such a secretion-based targeting mechanism for membrane proteins provides cells with flexibility in remodeling their architecture or evolving new forms. PMID:11134078

  2. The nociception genes painless and Piezo are required for the cellular immune response of Drosophila larvae to wasp parasitization.

    PubMed

    Tokusumi, Yumiko; Tokusumi, Tsuyoshi; Schulz, Robert A

    2017-03-22

    In vertebrates, interaction between the nervous system and immune system is important to protect a challenged host from stress inputs from external sources. In this study, we demonstrate that sensory neurons are involved in the cellular immune response elicited by wasp infestation of Drosophila larvae. Multidendritic class IV neurons sense contacts from external stimuli and induce avoidance behaviors for host defense. Our findings show that inactivation of these sensory neurons impairs the cellular response against wasp parasitization. We also demonstrate that the nociception genes encoding the mechanosensory receptors Painless and Piezo, both expressed in class IV neurons, are essential for the normal cellular immune response to parasite challenge.

  3. Implication of p53-dependent cellular senescence related gene, TARSH in tumor suppression

    SciTech Connect

    Wakoh, Takeshi; Uekawa, Natsuko; Terauchi, Kunihiko; Sugimoto, Masataka; Ishigami, Akihito; Shimada, Jun-ichi; Maruyama, Mitsuo

    2009-03-20

    A novel target of NESH-SH3 (TARSH) was identified as a cellular senescence related gene in mouse embryonic fibroblasts (MEFs) replicative senescence, the expression of which has been suppressed in primary clinical lung cancer specimens. However, the molecular mechanism underlying the regulation of TARSH involved in pulmonary tumorigenesis remains unclear. Here we demonstrate that the reduction of TARSH gene expression by short hairpin RNA (shRNA) system robustly inhibited the MEFs proliferation with increase in senescence-associated {beta}-galactosidase (SA-{beta}-gal) activity. Using p53{sup -/-} MEFs, we further suggest that this growth arrest by loss of TARSH is evoked by p53-dependent p21{sup Cip1} accumulation. Moreover, we also reveal that TARSH reduction induces multicentrosome in MEFs, which is linked in chromosome instability and tumor development. These results suggest that TARSH plays an important role in proliferation of replicative senescence and may serve as a trigger of tumor development.

  4. Gene Expression Profiling of Flagellar Disassembly in Chlamydomonas reinhardtii

    PubMed Central

    Chamberlain, Kara L.; Miller, Steven H.; Keller, Laura R.

    2008-01-01

    Flagella are sensory organelles that interact with the environment through signal transduction and gene expression networks. We used microarray profiling to examine gene regulation associated with flagellar length change in the green alga Chlamydomonas reinhardtii. Microarrays were probed with fluorescently labeled cDNAs synthesized from RNA extracted from cells before and during flagellar assembly or disassembly. Evaluation of the gene expression profiles identified >100 clones showing at least a twofold change in expression during flagellar length changes. Products of these genes are associated not only with flagellar structure and motility but also with other cellular responses, including signal transduction and metabolism. Expression of specific genes from each category was further characterized at higher resolution by using quantitative real-time PCR (qRT–PCR). Analysis and comparison of the gene expression profiles coupled to flagellar assembly and disassembly revealed that each process involves a new and uncharacterized whole-cell response to flagellar length changes. This analysis lays the groundwork for a more comprehensive understanding of the cellular and molecular networks regulating flagellar length changes. PMID:18493036

  5. Pulmonary Gene Expression Profiling of Inhaled Ricin

    DTIC Science & Technology

    2007-11-02

    in which 34 genes had statistically significant changes in gene expression. Transcripts identified by the assay included those that facilitate...gene expression. Transcripts identified by the assay included those that facilitate tissue healing (early growth response gene (egr)-1), regulate...impingement to determine aerosol concentration. Ricin concentrations from impinger samples were measured by protein assay (Pierce, MicroBCA, Rockford

  6. 76 FR 22405 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-21

    ... HUMAN SERVICES Food and Drug Administration Cellular, Tissue and Gene Therapies Advisory Committee... be open to the public. Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee... gene therapy products for the treatment of retinal disorders. Topics to be considered include...

  7. 77 FR 63840 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-17

    ... HUMAN SERVICES Food and Drug Administration Cellular, Tissue and Gene Therapies Advisory Committee..., Tissue and Gene Therapies Advisory Committee. General Function of the Committee: To provide advice and..., Office of Cellular, Tissue and Gene Therapies, Center for Biologics Evaluation and Research, and...

  8. 78 FR 79699 - Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-31

    ... No. FDA-2013-N-0001] Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of Meeting... the public. Name of Committee: Cellular, Tissue, and Gene Therapies Advisory Committee. General..., Tissue, and Gene Therapies, Center for Biologics Evaluation and Research (CBER), FDA. On February...

  9. 75 FR 66381 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-28

    ... HUMAN SERVICES Food and Drug Administration Cellular, Tissue and Gene Therapies Advisory Committee... be open to the public. Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee... Lentiviral Vector Based Gene Therapy Products. FDA intends to make background material available to...

  10. Emerging cellular and gene therapies for congenital anemias.

    PubMed

    Ludwig, Leif S; Khajuria, Rajiv K; Sankaran, Vijay G

    2016-12-01

    Congenital anemias comprise a group of blood disorders characterized by a reduction in the number of peripherally circulating erythrocytes. Various genetic etiologies have been identified that affect diverse aspects of erythroid physiology and broadly fall into two main categories: impaired production or increased destruction of mature erythrocytes. Current therapies are largely focused on symptomatic treatment and are often based on transfusion of donor-derived erythrocytes and management of complications. Hematopoietic stem cell transplantation represents the only curative option currently available for the majority of congenital anemias. Recent advances in gene therapy and genome editing hold promise for the development of additional curative strategies for these blood disorders. The relative ease of access to the hematopoietic stem cell compartment, as well as the possibility of genetic manipulation ex vivo and subsequent transplantation in an autologous manner, make blood disorders among the most amenable to cellular therapies. Here we review cell-based and gene therapy approaches, and discuss the limitations and prospects of emerging avenues, including genome editing tools and the use of pluripotent stem cells, for the treatment of congenital forms of anemia. © 2016 Wiley Periodicals, Inc.

  11. Does inbreeding affect gene expression in birds?

    PubMed

    Hansson, Bengt; Naurin, Sara; Hasselquist, Dennis

    2014-09-01

    Inbreeding increases homozygosity, exposes genome-wide recessive deleterious alleles and often reduces fitness. The physiological and reproductive consequences of inbreeding may be manifested already during gene regulation, but the degree to which inbreeding influences gene expression is unknown in most organisms, including in birds. To evaluate the pattern of inbreeding-affected gene expression over the genome and in relation to sex, we performed a transcriptome-wide gene expression (10 695 genes) study of brain tissue of 10-day-old inbred and outbred, male and female zebra finches. We found significantly lower gene expression in females compared with males at Z-linked genes, confirming that dosage compensation is incomplete in female birds. However, inbreeding did not affect gene expression at autosomal or sex-linked genes, neither in males nor in females. Analyses of single genes again found a clear sex-biased expression at Z-linked genes, whereas only a single gene was significantly affected by inbreeding. The weak effect of inbreeding on gene expression in zebra finches contrasts to the situation, for example, in Drosophila where inbreeding has been found to influence gene expression more generally and at stress-related genes in particular.

  12. Pathway network inference from gene expression data

    PubMed Central

    2014-01-01

    Background The development of high-throughput omics technologies enabled genome-wide measurements of the activity of cellular elements and provides the analytical resources for the progress of the Systems Biology discipline. Analysis and interpretation of gene expression data has evolved from the gene to the pathway and interaction level, i.e. from the detection of differentially expressed genes, to the establishment of gene interaction networks and the identification of enriched functional categories. Still, the understanding of biological systems requires a further level of analysis that addresses the characterization of the interaction between functional modules. Results We present a novel computational methodology to study the functional interconnections among the molecular elements of a biological system. The PANA approach uses high-throughput genomics measurements and a functional annotation scheme to extract an activity profile from each functional block -or pathway- followed by machine-learning methods to infer the relationships between these functional profiles. The result is a global, interconnected network of pathways that represents the functional cross-talk within the molecular system. We have applied this approach to describe the functional transcriptional connections during the yeast cell cycle and to identify pathways that change their connectivity in a disease condition using an Alzheimer example. Conclusions PANA is a useful tool to deepen in our understanding of the functional interdependences that operate within complex biological systems. We show the approach is algorithmically consistent and the inferred network is well supported by the available functional data. The method allows the dissection of the molecular basis of the functional connections and we describe the different regulatory mechanisms that explain the network's topology obtained for the yeast cell cycle data. PMID:25032889

  13. [Neuronal plasticity and gene expression].

    PubMed

    Sokolova, O O; Shtark, M B; Lisachev, P D

    2010-01-01

    Neuronal plasticity--a fundamental feature of brain--provides adequate interactions with dynamic environment. One of the most deeply investigated forms of the neuronal plasticity is a long-term potentiation (LTP)--a phenomenon underlying learning and memory. Signal paths activated during LTP converge into the nuclear of the neuron, giving rise to launch of the molecular-genetic programs, which mediate structural and functional remodeling of synapses. In the review data concerning involvement of multilevel gene expression into plastic change under neuronal activation are summarized.

  14. Novel redox nanomedicine improves gene expression of polyion complex vector

    NASA Astrophysics Data System (ADS)

    Toh, Kazuko; Yoshitomi, Toru; Ikeda, Yutaka; Nagasaki, Yukio

    2011-12-01

    Gene therapy has generated worldwide attention as a new medical technology. While non-viral gene vectors are promising candidates as gene carriers, they have several issues such as toxicity and low transfection efficiency. We have hypothesized that the generation of reactive oxygen species (ROS) affects gene expression in polyplex supported gene delivery systems. The effect of ROS on the gene expression of polyplex was evaluated using a nitroxide radical-containing nanoparticle (RNP) as an ROS scavenger. When polyethyleneimine (PEI)/pGL3 or PEI alone was added to the HeLa cells, ROS levels increased significantly. In contrast, when (PEI)/pGL3 or PEI was added with RNP, the ROS levels were suppressed. The luciferase expression was increased by the treatment with RNP in a dose-dependent manner and the cellular uptake of pDNA was also increased. Inflammatory cytokines play an important role in ROS generation in vivo. In particular, tumor necrosis factor (TNF)-α caused intracellular ROS generation in HeLa cells and decreased gene expression. RNP treatment suppressed ROS production even in the presence of TNF-α and increased gene expression. This anti-inflammatory property of RNP suggests that it may be used as an effective adjuvant for non-viral gene delivery systems.

  15. Inhibition of luciferase expression by synthetic hammerhead ribozymes and their cellular uptake.

    PubMed Central

    Bramlage, B; Alefelder, S; Marschall, P; Eckstein, F

    1999-01-01

    Two synthetic hammerhead ribozymes, one unmodified and the other with 2"-modifications and four phosphorothioate groups, targeting a single GUA site in the luciferase mRNA, were compared for their inhibition of gene expression in cell cultureand their cellular uptake was also analysed. A HeLa X1/5 cell line stably expressing luciferase, under an inducible promoter, was treated with these ribozymes by liposome-mediated transfection to determine their activity.Luciferase expression in cells was inhibited to approximately 50% with little difference between the unmodified and the 2"-modified ribozyme. A similar degree of inhibition was observed with two catalytically inactive ribozymes, indicating that inhibition was mainly due to an antisense effect. A ribozyme carrying a cholesterol moiety, applied to the cells without carrier, showed no inhibition. Northern blotting indicated a similar amount of cellular uptake of all ribozymes. The unmodified ribozyme was essentially evenly distributed between cytoplasm and nucleus, whereas a higher proportion of the phosphorothioate-containing ribozyme was observed in the nucleus. Fluorescence microscopy, including confocal microscopy using 5"-fluorescein-labelled ribozymes, showed that the unmodified and 2"-modified ribozymes were present in the cytoplasm and in the nucleus to a similar extent, whereas the fluorescence of the phosphorothioate-containing ribozyme was much stronger in the nucleus. Both ribozymes inhibited luciferase expression to a comparable degree, suggesting that the ribozyme in the nucleus did not contribute significantly to the inhibition. Ribozymes with a cholesterol moiety were predominantly trapped in the cell membrane, explaining their inability to interfere with gene expression. PMID:10454613

  16. Gene expression and dental enamel structure in developing mouse incisor.

    PubMed

    Sehic, Amer; Risnes, Steinar; Khan, Qalb-E-Saleem; Khuu, Cuong; Osmundsen, Harald

    2010-04-01

    At the mouse incisor tip the initially differentiated ameloblasts produce a thin, prism-free enamel, while further apically, in the immediate adjacent segment, the enamel thickness increases and the four-layered enamel of mouse incisor is formed. Comparative gene-expression profiling was carried out on RNA isolated from these two segments of incisor tooth germs at embryonic day (E)17.5 and at postnatal days (P)0, 1, 2, and 10 using microarrays to measure messenger RNA (mRNA) and microRNA (miRNA) species present in the segments. Validation of expression data was achieved using real-time reverse transcription-polymerase chain reaction (RT-PCR) and western blotting. Bioinformatic data suggested enhanced cellular apoptosis in the incisal tip segment, which, together with diminished expression of the Amelx and Enam genes, may contribute to the production of the thin enamel seen in this tooth segment. For genes exhibiting higher levels of expression in the adjacent segment where complex enamel is being formed, bioinformatic analysis suggested significant associations with cellular functions involving the actin cytoskeleton, cellular development, morphology, and movement. This is suggested to reflect that ameloblasts with Tomes' process are being organized in transverse rows, facilitating the transverse movement that results in prism decussation in the inner enamel of the adjacent segment. Bioinformatic analysis of miRNA expression data lends support to these suggestions.

  17. Gene expression analysis of endometrium reveals progesterone resistance and candidate susceptibility genes in women with endometriosis.

    PubMed

    Burney, Richard O; Talbi, Said; Hamilton, Amy E; Vo, Kim Chi; Nyegaard, Mette; Nezhat, Camran R; Lessey, Bruce A; Giudice, Linda C

    2007-08-01

    The identification of molecular differences in the endometrium of women with endometriosis is an important step toward understanding the pathogenesis of this condition and toward developing novel strategies for the treatment of associated infertility and pain. In this study, we conducted global gene expression analysis of endometrium from women with and without moderate/severe stage endometriosis and compared the gene expression signatures across various phases of the menstrual cycle. The transcriptome analysis revealed molecular dysregulation of the proliferative-to-secretory transition in endometrium of women with endometriosis. Paralleled gene expression analysis of endometrial specimens obtained during the early secretory phase demonstrated a signature of enhanced cellular survival and persistent expression of genes involved in DNA synthesis and cellular mitosis in the setting of endometriosis. Comparative gene expression analysis of progesterone-regulated genes in secretory phase endometrium confirmed the observation of attenuated progesterone response. Additionally, interesting candidate susceptibility genes were identified that may be associated with this disorder, including FOXO1A, MIG6, and CYP26A1. Collectively these findings provide a framework for further investigations on causality and mechanisms underlying attenuated progesterone response in endometrium of women with endometriosis.

  18. Noncytopathic Sindbis virus RNA vectors for heterologous gene expression

    PubMed Central

    Agapov, Eugene V.; Frolov, Ilya; Lindenbach, Brett D.; Prágai, Béla M.; Schlesinger, Sondra; Rice, Charles M.

    1998-01-01

    Infection of vertebrate cells with alphaviruses normally leads to prodigious expression of virus-encoded genes and a dramatic inhibition of host protein synthesis. Recombinant Sindbis viruses and replicons have been useful as vectors for high level foreign gene expression, but the cytopathic effects of viral replication have limited their use to transient studies. We recently selected Sindbis replicons capable of persistent, noncytopathic growth in BHK cells and describe here a new generation of Sindbis vectors useful for long-term foreign gene expression based on such replicons. Foreign genes of interest as well as the dominant selectable marker puromycin N-acteyltransferase, which confers resistance to the drug puromycin, were expressed as subgenomic transcripts of noncytopathic replicons or defective-interfering genomes complemented in trans by a replicon. Based on these strategies, we developed vectors that can be initiated via either RNA or DNA transfection and analyzed them for their level and stability of foreign gene expression. Noncytopathic Sindbis vectors express reasonably high levels of protein in nearly every cell. These vectors should prove to be flexible tools for the rapid expression of heterologous genes under conditions in which cellular metabolism is not perturbed, and we illustrate their utility with a number of foreign proteins. PMID:9789028

  19. Roles of neural precursor cell expressed, developmentally downregulated 9 in tumor-associated cellular processes (Review).

    PubMed

    Zhang, Sisen; Wu, Lihua

    2015-11-01

    Neural precursor cell expressed, developmentally downregulated 9 (NEDD9), a gene exclusively expressed in the brain during embryonic stages but not in brains of adult mice, is an important cytoskeletal protein and regarded as a 'router/hub' in cellular signal transduction processes connecting external stimulation signals with downstream target proteins that can directly promote tumor metastasis. Numerous studies showed that NEDD9 has an essential role in cell proliferation, apoptosis, adhesion, migration and invasion. The roles of NEDD9, including the underlying mechanisms of its regulation of cell migration, its distinctive functions in various tumor stages and its association with other diseases, are required to be elucidated at large. Future studies of NEDD9 may provide a more profound understanding of the development of tumor invasiveness and NEDD9 may serve as a potential novel target for tumor therapy. The present review examined the significant roles of NEDD9 in the abovementioned processes.

  20. Cell cycle, apoptosis, cellular uptake and whole-transcriptome microarray gene expression analysis of HeLa cells treated with a ruthenium(II)-arene complex with an isoquinoline-3-carboxylic acid ligand.

    PubMed

    Jovanović, Katarina K; Tanić, Miljana; Ivanović, Ivanka; Gligorijević, Nevenka; Dojčinović, Biljana P; Radulović, Siniša

    2016-10-01

    Ruthenium(II)-arene complexes are promising drug candidates for the therapy of solid tumors. In previous work, seven new compounds of the general formula [Ru(η(6)-p-cymene)(L(1-7))Cl] were synthesized and characterized, of which the complex with L=isoquinoline-3-carboxylic acid (RuT7) was two times as active on HeLa cells compared to normal cell line MRC-5, as indicated by IC50 values determined after 48h of incubation (45.4±3.0 vs. 84.2±5.7μM, respectively). In the present study, cell cycle analysis of HeLa cells treated with RuT7 showed S phase arrest and an increase in sub-G1 population. The apoptotic potential of the title compound was confirmed with the Annexin V-FITC/PI assay together with a morphological evaluation of cells using fluorescent microscopy. Analysis of the intracellular accumulation of ruthenium showed 8.9ng Ru/10(6) cells after 6h of incubation. To gain further insight in the molecular mechanism of action of RuT7 on HeLa cells, a whole-transcriptome microarray gene expression analysis was performed. Analysis of functional categories and signaling and biochemical pathways associated with the response of HeLa cells to treatment with RuT7 showed that it leads the cells through the intrinsic (mitochondrial) apoptotic pathway, via indirect DNA damage due to the action of reactive oxygen species, and through direct DNA binding of RuT7. Statistical analysis for enrichment of gene sets associated with known drug-induced toxicities identified fewer associated toxicity profiles in RuT7-treated cells compared to cisplatin treatment. Altogether these results provide the basis for further development of RuT7 in animal and pre-clinical studies as a potential drug candidate.

  1. Gene Expression Profiling in the Type 1 Diabetes Rat Diaphragm

    PubMed Central

    van Lunteren, Erik; Moyer, Michelle

    2009-01-01

    Background Respiratory muscle contractile performance is impaired by diabetes, mechanisms of which included altered carbohydrate and lipid metabolism, oxidative stress and changes in membrane electrophysiology. The present study examined to what extent these cellular perturbations involve changes in gene expression. Methodology/Principal Findings Diaphragm muscle from streptozotocin-diabetic rats was analyzed with Affymetrix gene expression arrays. Diaphragm from diabetic rats had 105 genes with at least ±2-fold significantly changed expression (55 increased, 50 decreased), and these were assigned to gene ontology groups based on over-representation analysis using DAVID software. There was increased expression of genes involved in palmitoyl-CoA hydrolase activity (a component of lipid metabolism) (P = 0.037, n = 2 genes, fold change 4.2 to 27.5) and reduced expression of genes related to carbohydrate metabolism (P = 0.000061, n = 8 genes, fold change −2.0 to −8.5). Other gene ontology groups among upregulated genes were protein ubiquitination (P = 0.0053, n = 4, fold change 2.2 to 3.4), oxidoreductase activity (P = 0.024, n = 8, fold change 2.1 to 6.0), and morphogenesis (P = 0.012, n = 10, fold change 2.1 to 4.3). Other downregulated gene groups were extracellular region (including extracellular matrix and collagen) (P = 0.00032, n = 13, fold change −2.2 to −3.7) and organogenesis (P = 0.032, n = 7, fold change −2.1 to −3.7). Real-time PCR confirmed the directionality of changes in gene expression for 30 of 31 genes tested. Conclusions/Significance These data indicate that in diaphragm muscle type 1 diabetes increases expression of genes involved in lipid energetics, oxidative stress and protein ubiquitination, decreases expression of genes involved in carbohydrate metabolism, and has little effect on expression of ion channel genes. Reciprocal changes in expression of genes involved in

  2. Analysis of global gene expression profiles to identify differentially expressed genes critical for embryo development in Brassica rapa.

    PubMed

    Zhang, Yu; Peng, Lifang; Wu, Ya; Shen, Yanyue; Wu, Xiaoming; Wang, Jianbo

    2014-11-01

    Embryo development represents a crucial developmental period in the life cycle of flowering plants. To gain insights into the genetic programs that control embryo development in Brassica rapa L., RNA sequencing technology was used to perform transcriptome profiling analysis of B. rapa developing embryos. The results generated 42,906,229 sequence reads aligned with 32,941 genes. In total, 27,760, 28,871, 28,384, and 25,653 genes were identified from embryos at globular, heart, early cotyledon, and mature developmental stages, respectively, and analysis between stages revealed a subset of stage-specific genes. We next investigated 9,884 differentially expressed genes with more than fivefold changes in expression and false discovery rate ≤ 0.001 from three adjacent-stage comparisons; 1,514, 3,831, and 6,633 genes were detected between globular and heart stage embryo libraries, heart stage and early cotyledon stage, and early cotyledon and mature stage, respectively. Large numbers of genes related to cellular process, metabolism process, response to stimulus, and biological process were expressed during the early and middle stages of embryo development. Fatty acid biosynthesis, biosynthesis of secondary metabolites, and photosynthesis-related genes were expressed predominantly in embryos at the middle stage. Genes for lipid metabolism and storage proteins were highly expressed in the middle and late stages of embryo development. We also identified 911 transcription factor genes that show differential expression across embryo developmental stages. These results increase our understanding of the complex molecular and cellular events during embryo development in B. rapa and provide a foundation for future studies on other oilseed crops.

  3. 77 FR 73472 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-12-10

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF HEALTH AND HUMAN SERVICES Food and Drug Administration Cellular, Tissue and Gene Therapies Advisory Committee... and Gene Therapies Advisory Committee. General Function of the Committee: To provide advice...

  4. Transient expression and cellular localization of recombinant proteins in cultured insect cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Heterologous protein expression systems are used for production of recombinant proteins, interpretation of cellular trafficking/localization, and for the determination of biochemical function of proteins at the sub-organismal level. Although baculovirus expression systems are increasingly used for ...

  5. Development and application of a rat ovarian gene expression database.

    PubMed

    Jo, Misung; Gieske, Mary C; Payne, Charles E; Wheeler-Price, Sarah E; Gieske, Joseph B; Ignatius, Ignatius V; Curry, Thomas E; Ko, Chemyong

    2004-11-01

    The pituitary gonadotropins play a key role in follicular development and ovulation through the induction of specific genes. To identify these genes, we have constructed a genome-wide rat ovarian gene expression database (rOGED). The database was constructed from total RNA isolated from intact ovaries, granulosa cells, or residual ovarian tissues collected from immature pregnant mare serum gonadotropin (PMSG)/human chorionic gonadotropin-treated rats at 0 h (no PMSG), 12 h, and 48 h post PMSG, as well as 6 and 12 h post human chorionic gonadotropin. The total RNA was used for DNA microarray analysis using Affymetrix Rat Expression Arrays 230A and 230B (Affymetrix, Santa Clara, CA). The microarray data were compiled and used for display of individual gene expression profiles through specially developed software. The final rOGED provides immediate analysis of temporal gene expression profiles for over 28,000 genes in intact ovaries, granulosa cells, and residual ovarian tissue during follicular growth and the preovulatory period. The accuracy of the rOGED was validated against the gene profiles for over 20 known genes. The utility of the rOGED was demonstrated by identifying six genes that have not been described in the rat periovulatory ovary. The mRNA expression patterns and cellular localization for each of these six genes (estrogen sulfotransferase, synaptosomal-associated protein 25 kDa, runt-related transcription factor, calgranulin B, alpha1-macroglobulin, and MAPK phosphotase-3) were confirmed by Northern blot analyses and in situ hybridization, respectively. The current findings demonstrate that the rOGED can be used as an instant reference for ovarian gene expression profiles, as well as a reliable resource for identifying important yet, to date, unknown ovarian genes.

  6. Differential Gene Expression in Human Cerebrovascular Malformations

    PubMed Central

    Shenkar, Robert; Elliott, J. Paul; Diener, Katrina; Gault, Judith; Hu, Ling-Jia; Cohrs, Randall J.; Phang, Tzulip; Hunter, Lawrence; Breeze, Robert E.; Awad, Issam A.

    2009-01-01

    OBJECTIVE We sought to identify genes with differential expression in cerebral cavernous malformations (CCMs), arteriovenous malformations (AVMs), and control superficial temporal arteries (STAs) and to confirm differential expression of genes previously implicated in the pathobiology of these lesions. METHODS Total ribonucleic acid was isolated from four CCM, four AVM, and three STA surgical specimens and used to quantify lesion-specific messenger ribonucleic acid expression levels on human gene arrays. Data were analyzed with the use of two separate methodologies: gene discovery and confirmation analysis. RESULTS The gene discovery method identified 42 genes that were significantly up-regulated and 36 genes that were significantly down-regulated in CCMs as compared with AVMs and STAs (P = 0.006). Similarly, 48 genes were significantly up-regulated and 59 genes were significantly down-regulated in AVMs as compared with CCMs and STAs (P = 0.006). The confirmation analysis showed significant differential expression (P < 0.05) in 11 of 15 genes (angiogenesis factors, receptors, and structural proteins) that previously had been reported to be expressed differentially in CCMs and AVMs in immunohistochemical analysis. CONCLUSION We identify numerous genes that are differentially expressed in CCMs and AVMs and correlate expression with the immunohistochemistry of genes implicated in cerebrovascular malformations. In future efforts, we will aim to confirm candidate genes specifically related to the pathobiology of cerebrovascular malformations and determine their biological systems and mechanistic relevance. PMID:12535382

  7. Cellular localization of transforming growth factor-beta expression in bleomycin-induced pulmonary fibrosis.

    PubMed Central

    Zhang, K.; Flanders, K. C.; Phan, S. H.

    1995-01-01

    Bleomycin-induced pulmonary fibrosis is associated with increased lung transforming growth factor-beta (TGF-beta) gene expression, but cellular localization of the source of this expression has not been unequivocally established. In this study, lung fibrosis was induced in rats by endotracheal bleomycin injection on day 0 and, on selected days afterwards, lungs were harvested for in situ hybridization, immunohistochemical and histochemical analyses for TGF-beta 1 mRNA and protein expression, and cell identification. The results show that control lungs express essentially no detectable TGF-beta 1 mRNA or protein in the parenchyma. Before day 3 after bleomycin treatment, scattered bronchiolar epithelial cells, mononuclear cells, and eosinophils expressed elevated levels of TGF-beta 1. Between days 3 and 14, there was a major increase in the number of eosinophils, myofibroblasts, and fibroblasts strongly expressing TGF-beta 1 mRNA and protein. TGF-beta 1-producing cells were predominantly localized within areas of injury and active fibrosis. After day 14, the intensity and number of TGF-beta 1-expressing cells significantly declined and were predominantly found in fibroblasts in fibrotic areas. The expression of TGF-beta 1 protein was generally coincident with that for mRNA with the exception of bronchiolar epithelial cells in which strong protein expression was unaccompanied by a commensurate increase in mRNA. The study demonstrates that myofibroblasts, fibroblasts, and eosinophils represent the major sources of increased lung TGF-beta 1 expression in this model of pulmonary fibrosis. Images Figure 2 Figure 3 Figure 4 PMID:7543734

  8. Membrane channel gene expression in human costal and articular chondrocytes

    PubMed Central

    Asmar, A.; Barrett-Jolley, R.; Werner, A.; Kelly, R.; Stacey, M.

    2016-01-01

    ABSTRACT Chondrocytes are the uniquely resident cells found in all types of cartilage and key to their function is the ability to respond to mechanical loads with changes of metabolic activity. This mechanotransduction property is, in part, mediated through the activity of a range of expressed transmembrane channels; ion channels, gap junction proteins, and porins. Appropriate expression of ion channels has been shown essential for production of extracellular matrix and differential expression of transmembrane channels is correlated to musculoskeletal diseases such as osteoarthritis and Albers-Schönberg. In this study we analyzed the consistency of gene expression between channelomes of chondrocytes from human articular and costal (teenage and fetal origin) cartilages. Notably, we found 14 ion channel genes commonly expressed between articular and both types of costal cartilage chondrocytes. There were several other ion channel genes expressed only in articular (6 genes) or costal chondrocytes (5 genes). Significant differences in expression of BEST1 and KCNJ2 (Kir2.1) were observed between fetal and teenage costal cartilage. Interestingly, the large Ca2+ activated potassium channel (BKα, or KCNMA1) was very highly expressed in all chondrocytes examined. Expression of the gap junction genes for Panx1, GJA1 (Cx43) and GJC1 (Cx45) was also observed in chondrocytes from all cartilage samples. Together, this data highlights similarities between chondrocyte membrane channel gene expressions in cells derived from different anatomical sites, and may imply that common electrophysiological signaling pathways underlie cellular control. The high expression of a range of mechanically and metabolically sensitive membrane channels suggest that chondrocyte mechanotransduction may be more complex than previously thought. PMID:27116676

  9. Macrophages in gene therapy: cellular delivery vehicles and in vivo targets.

    PubMed

    Burke, B; Sumner, S; Maitland, N; Lewis, C E

    2002-09-01

    The appearance and activation of macrophages are thought to be rapid events in the development of many pathological lesions, including malignant tumors, atherosclerotic plaques, and arthritic joints. This has prompted recent attempts to use macrophages as novel cellular vehicles for gene therapy, in which macrophages are genetically modified ex vivo and then reintroduced into the body with the hope that a proportion will then home to the diseased site. Here, we critically review the efficacy of various gene transfer methods (viral, bacterial, protozoan, and various chemical and physical methods) in transfecting macrophages in vitro, and the results obtained when transfected macrophages are used as gene delivery vehicles. Finally, we discuss the use of various viral and nonviral methods to transfer genes to macrophages in vivo. As will be seen, definitive evidence for the use of macrophages as gene transfer vehicles has yet to be provided and awaits detailed trafficking studies in vivo. Moreover, although methods for transfecting macrophages have improved considerably in efficiency in recent years, targeting of gene transfer specifically to macrophages in vivo remains a problem. However, possible solutions to this include placing transgenes under the control of macrophage-specific promoters to limit expression to macrophages or stably transfecting CD34(+) precursors of monocytes/macrophages and then differentiating these cells into monocytes/macrophages ex vivo. The latter approach could conceivably lead to the bone marrow precursor cells of patients with inherited genetic disorders being permanently fortified or even replaced with genetically modified cells.

  10. Norovirus gene expression and replication.

    PubMed

    Thorne, Lucy G; Goodfellow, Ian G

    2014-02-01

    Noroviruses are small, positive-sense RNA viruses within the family Caliciviridae, and are now accepted widely as a major cause of acute gastroenteritis in both developed and developing countries. Despite their impact, our understanding of the life cycle of noroviruses has lagged behind that of other RNA viruses due to the inability to culture human noroviruses (HuNVs). Our knowledge of norovirus biology has improved significantly over the past decade as a result of numerous technological advances. The use of a HuNV replicon, improved biochemical and cell-based assays, combined with the discovery of a murine norovirus capable of replication in cell culture, has improved greatly our understanding of the molecular mechanisms of norovirus genome translation and replication, as well as the interaction with host cell processes. In this review, the current state of knowledge of the intracellular life of noroviruses is discussed with particular emphasis on the mechanisms of viral gene expression and viral genome replication.

  11. 78 FR 15726 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-12

    ... HUMAN SERVICES Food and Drug Administration Cellular, Tissue and Gene Therapies Advisory Committee... portion of the meeting will be closed to the public. Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee. General Function of the Committee: To provide advice and recommendations...

  12. 76 FR 18768 - Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-05

    ... HUMAN SERVICES Food and Drug Administration Cellular, Tissue, and Gene Therapies Advisory Committee... portion of the meeting will be closed to the public. Name of Committee: Cellular, Tissue, and Gene Therapies Advisory Committee. General Function of the Committee: To provide advice and recommendations...

  13. 76 FR 64951 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-10-19

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF HEALTH AND HUMAN SERVICES Food and Drug Administration Cellular, Tissue and Gene Therapies Advisory Committee... be open to the public. Name of Committee: Cellular, Tissue and Gene Therapies Advisory...

  14. GIM3E: Condition-specific Models of Cellular Metabolism Developed from Metabolomics and Expression Data

    SciTech Connect

    Schmidt, Brian; Ebrahim, Ali; Metz, Thomas O.; Adkins, Joshua N.; Palsson, Bernard O.; Hyduke, Daniel R.

    2013-11-15

    Motivation: Genome-scale metabolic models have been used extensively to investigate alterations in cellular metabolism. The accuracy of these models to represent cellular metabolism in specific conditions has been improved by constraining the model with omics data sources. However, few practical methods for integrating metabolomics data with other omics data sources into genome-scale models of metabolism have been reported. Results: GIMMME (Gene Inactivation Moderated by Metabolism, Metabolomics, and Expression) is an algorithm that enables the development of condition-specific models based on an objective function, transcriptomics, and intracellular metabolomics data. GIMMME establishes metabolite utilization requirements with metabolomics data, uses model-paired transcriptomics data to find experimentally supported solutions, and also provides calculations of the turnover (production / consumption) flux of metabolites. GIMMME was employed to investigate the effects of integrating additional omics datasets to create increasingly constrained solution spaces of Salmonella Typhimurium metabolism during growth in both rich and virulence media. This integration proved to be informative and resulted in a requirement of additional active reactions (12 in each case) or metabolites (26 or 29, respectively). The addition of constraints from transcriptomics also impacted the allowed solution space, and the cellular metabolites with turnover fluxes that were necessarily altered by the change in conditions increased from 118 to 271 of 1397. Availability: GIMMME has been implemented in Python and requires a COBRApy 0.2.x. The algorithm and sample data described here are freely available at: http://opencobra.sourceforge.net/

  15. Analytical workflow profiling gene expression in murine macrophages

    PubMed Central

    Nixon, Scott E.; González-Peña, Dianelys; Lawson, Marcus A.; McCusker, Robert H.; Hernandez, Alvaro G.; O’Connor, Jason C.; Dantzer, Robert; Kelley, Keith W.

    2015-01-01

    Comprehensive and simultaneous analysis of all genes in a biological sample is a capability of RNA-Seq technology. Analysis of the entire transcriptome benefits from summarization of genes at the functional level. As a cellular response of interest not previously explored with RNA-Seq, peritoneal macrophages from mice under two conditions (control and immunologically challenged) were analyzed for gene expression differences. Quantification of individual transcripts modeled RNA-Seq read distribution and uncertainty (using a Beta Negative Binomial distribution), then tested for differential transcript expression (False Discovery Rate-adjusted p-value < 0.05). Enrichment of functional categories utilized the list of differentially expressed genes. A total of 2079 differentially expressed transcripts representing 1884 genes were detected. Enrichment of 92 categories from Gene Ontology Biological Processes and Molecular Functions, and KEGG pathways were grouped into 6 clusters. Clusters included defense and inflammatory response (Enrichment Score = 11.24) and ribosomal activity (Enrichment Score = 17.89). Our work provides a context to the fine detail of individual gene expression differences in murine peritoneal macrophages during immunological challenge with high throughput RNA-Seq. PMID:25708305

  16. Gene expression profiling in peanut using high density oligonucleotide microarrays

    PubMed Central

    Payton, Paxton; Kottapalli, Kameswara Rao; Rowland, Diane; Faircloth, Wilson; Guo, Baozhu; Burow, Mark; Puppala, Naveen; Gallo, Maria

    2009-01-01

    Background Transcriptome expression analysis in peanut to date has been limited to a relatively small set of genes and only recently has a significant number of ESTs been released into the public domain. Utilization of these ESTs for oligonucleotide microarrays provides a means to investigate large-scale transcript responses to a variety of developmental and environmental signals, ultimately improving our understanding of plant biology. Results We have developed a high-density oligonucleotide microarray for peanut using 49,205 publicly available ESTs and tested the utility of this array for expression profiling in a variety of peanut tissues. To identify putatively tissue-specific genes and demonstrate the utility of this array for expression profiling in a variety of peanut tissues, we compared transcript levels in pod, peg, leaf, stem, and root tissues. Results from this experiment showed 108 putatively pod-specific/abundant genes, as well as transcripts whose expression was low or undetected in pod compared to peg, leaf, stem, or root. The transcripts significantly over-represented in pod include genes responsible for seed storage proteins and desiccation (e.g., late-embryogenesis abundant proteins, aquaporins, legumin B), oil production, and cellular defense. Additionally, almost half of the pod-abundant genes represent unknown genes allowing for the possibility of associating putative function to these previously uncharacterized genes. Conclusion The peanut oligonucleotide array represents the majority of publicly available peanut ESTs and can be used as a tool for expression profiling studies in diverse tissues. PMID:19523230

  17. Common Genetic Variation In Cellular Transport Genes and Epithelial Ovarian Cancer (EOC) Risk

    PubMed Central

    Chornokur, Ganna; Lin, Hui-Yi; Tyrer, Jonathan P.; Lawrenson, Kate; Dennis, Joe; Amankwah, Ernest K.; Qu, Xiaotao; Tsai, Ya-Yu; Jim, Heather S. L.; Chen, Zhihua; Chen, Ann Y.; Permuth-Wey, Jennifer; Aben, Katja KH.; Anton-Culver, Hoda; Antonenkova, Natalia; Bruinsma, Fiona; Bandera, Elisa V.; Bean, Yukie T.; Beckmann, Matthias W.; Bisogna, Maria; Bjorge, Line; Bogdanova, Natalia; Brinton, Louise A.; Brooks-Wilson, Angela; Bunker, Clareann H.; Butzow, Ralf; Campbell, Ian G.; Carty, Karen; Chang-Claude, Jenny; Cook, Linda S.; Cramer, Daniel W.; Cunningham, Julie M.; Cybulski, Cezary; Dansonka-Mieszkowska, Agnieszka; du Bois, Andreas; Despierre, Evelyn; Dicks, Ed; Doherty, Jennifer A.; Dörk, Thilo; Dürst, Matthias; Easton, Douglas F.; Eccles, Diana M.; Edwards, Robert P.; Ekici, Arif B.; Fasching, Peter A.; Fridley, Brooke L.; Gao, Yu-Tang; Gentry-Maharaj, Aleksandra; Giles, Graham G.; Glasspool, Rosalind; Goodman, Marc T.; Gronwald, Jacek; Harrington, Patricia; Harter, Philipp; Hein, Alexander; Heitz, Florian; Hildebrandt, Michelle A. T.; Hillemanns, Peter; Hogdall, Claus K.; Hogdall, Estrid; Hosono, Satoyo; Jakubowska, Anna; Jensen, Allan; Ji, Bu-Tian; Karlan, Beth Y.; Kelemen, Linda E.; Kellar, Mellissa; Kiemeney, Lambertus A.; Krakstad, Camilla; Kjaer, Susanne K.; Kupryjanczyk, Jolanta; Lambrechts, Diether; Lambrechts, Sandrina; Le, Nhu D.; Lee, Alice W.; Lele, Shashi; Leminen, Arto; Lester, Jenny; Levine, Douglas A.; Liang, Dong; Lim, Boon Kiong; Lissowska, Jolanta; Lu, Karen; Lubinski, Jan; Lundvall, Lene; Massuger, Leon F. A. G.; Matsuo, Keitaro; McGuire, Valerie; McLaughlin, John R.; McNeish, Iain; Menon, Usha; Milne, Roger L.; Modugno, Francesmary; Moysich, Kirsten B.; Ness, Roberta B.; Nevanlinna, Heli; Eilber, Ursula; Odunsi, Kunle; Olson, Sara H.; Orlow, Irene; Orsulic, Sandra; Weber, Rachel Palmieri; Paul, James; Pearce, Celeste L.; Pejovic, Tanja; Pelttari, Liisa M.; Pike, Malcolm C.; Poole, Elizabeth M.; Risch, Harvey A.; Rosen, Barry; Rossing, Mary Anne; Rothstein, Joseph H.; Rudolph, Anja; Runnebaum, Ingo B.; Rzepecka, Iwona K.; Salvesen, Helga B.; Schernhammer, Eva; Schwaab, Ira; Shu, Xiao-Ou; Shvetsov, Yurii B.; Siddiqui, Nadeem; Sieh, Weiva; Song, Honglin; Southey, Melissa C.; Spiewankiewicz, Beata; Sucheston, Lara; Teo, Soo-Hwang; Terry, Kathryn L.; Thompson, Pamela J.; Thomsen, Lotte; Tangen, Ingvild L.; Tworoger, Shelley S.; van Altena, Anne M.; Vierkant, Robert A.; Vergote, Ignace; Walsh, Christine S.; Wang-Gohrke, Shan; Wentzensen, Nicolas; Whittemore, Alice S.; Wicklund, Kristine G.; Wilkens, Lynne R.; Wu, Anna H.; Wu, Xifeng; Woo, Yin-Ling; Yang, Hannah; Zheng, Wei; Ziogas, Argyrios; Hasmad, Hanis N.; Berchuck, Andrew; Iversen, Edwin S.; Schildkraut, Joellen M.; Ramus, Susan J.; Goode, Ellen L.; Monteiro, Alvaro N. A.; Gayther, Simon A.; Narod, Steven A.; Pharoah, Paul D. P.; Sellers, Thomas A.; Phelan, Catherine M.

    2015-01-01

    Background Defective cellular transport processes can lead to aberrant accumulation of trace elements, iron, small molecules and hormones in the cell, which in turn may promote the formation of reactive oxygen species, promoting DNA damage and aberrant expression of key regulatory cancer genes. As DNA damage and uncontrolled proliferation are hallmarks of cancer, including epithelial ovarian cancer (EOC), we hypothesized that inherited variation in the cellular transport genes contributes to EOC risk. Methods In total, DNA samples were obtained from 14,525 case subjects with invasive EOC and from 23,447 controls from 43 sites in the Ovarian Cancer Association Consortium (OCAC). Two hundred seventy nine SNPs, representing 131 genes, were genotyped using an Illumina Infinium iSelect BeadChip as part of the Collaborative Oncological Gene-environment Study (COGS). SNP analyses were conducted using unconditional logistic regression under a log-additive model, and the FDR q<0.2 was applied to adjust for multiple comparisons. Results The most significant evidence of an association for all invasive cancers combined and for the serous subtype was observed for SNP rs17216603 in the iron transporter gene HEPH (invasive: OR = 0.85, P = 0.00026; serous: OR = 0.81, P = 0.00020); this SNP was also associated with the borderline/low malignant potential (LMP) tumors (P = 0.021). Other genes significantly associated with EOC histological subtypes (p<0.05) included the UGT1A (endometrioid), SLC25A45 (mucinous), SLC39A11 (low malignant potential), and SERPINA7 (clear cell carcinoma). In addition, 1785 SNPs in six genes (HEPH, MGST1, SERPINA, SLC25A45, SLC39A11 and UGT1A) were imputed from the 1000 Genomes Project and examined for association with INV EOC in white-European subjects. The most significant imputed SNP was rs117729793 in SLC39A11 (per allele, OR = 2.55, 95% CI = 1.5-4.35, p = 5.66x10-4). Conclusion These results, generated on a large cohort of women, revealed associations

  18. Human immunodeficiency virus type 1 Tat increases the expression of cleavage and polyadenylation specificity factor 73-kilodalton subunit modulating cellular and viral expression.

    PubMed

    Calzado, Marco A; Sancho, Rocío; Muñoz, Eduardo

    2004-07-01

    The human immunodeficiency virus type 1 (HIV-1) Tat protein, which is essential for HIV gene expression and viral replication, is known to mediate pleiotropic effects on various cell functions. For instance, Tat protein is able to regulate the rate of transcription of host cellular genes and to interact with the signaling machinery, leading to cellular dysfunction. To study the effect that HIV-1 Tat exerts on the host cell, we identified several genes that were up- or down-regulated in tat-expressing cell lines by using the differential display method. HIV-1 Tat specifically increases the expression of the cleavage and polyadenylation specificity factor (CPSF) 73-kDa subunit (CPSF3) without affecting the expression of the 160- and 100-kDa subunits of the CPSF complex. This complex comprises four subunits and has a key function in the 3'-end processing of pre-mRNAs by a coordinated interaction with other factors. CPSF3 overexpression experiments and knockdown of the endogenous CPSF3 by mRNA interference have shown that this subunit of the complex is an important regulatory protein for both viral and cellular gene expression. In addition to the known CPSF3 function in RNA polyadenylation, we also present evidence that this protein exerts transcriptional activities by repressing the mdm2 gene promoter. Thus, HIV-1-Tat up-regulation of CPSF3 could represent a novel mechanism by which this virus increases mRNA processing, causing an increase in both cell and viral gene expression.

  19. Familial aggregation analysis of gene expressions

    PubMed Central

    Rao, Shao-Qi; Xu, Liang-De; Zhang, Guang-Mei; Li, Xia; Li, Lin; Shen, Gong-Qing; Jiang, Yang; Yang, Yue-Ying; Gong, Bin-Sheng; Jiang, Wei; Zhang, Fan; Xiao, Yun; Wang, Qing K

    2007-01-01

    Traditional studies of familial aggregation are aimed at defining the genetic (and non-genetic) causes of a disease from physiological or clinical traits. However, there has been little attempt to use genome-wide gene expressions, the direct phenotypic measures of genes, as the traits to investigate several extended issues regarding the distributions of familially aggregated genes on chromosomes or in functions. In this study we conducted a genome-wide familial aggregation analysis by using the in vitro cell gene expressions of 3300 human autosome genes (Problem 1 data provided to Genetic Analysis Workshop 15) in order to answer three basic genetics questions. First, we investigated how gene expressions aggregate among different types (degrees) of relative pairs. Second, we conducted a bioinformatics analysis of highly familially aggregated genes to see how they are distributed on chromosomes. Third, we performed a gene ontology enrichment test of familially aggregated genes to find evidence to support their functional consensus. The results indicated that 1) gene expressions did aggregate in families, especially between sibs. Of 3300 human genes analyzed, there were a total of 1105 genes with one or more significant (empirical p < 0.05) familial correlation; 2) there were several genomic hot spots where highly familially aggregated genes (e.g., the chromosome 6 HLA genes cluster) were clustered; 3) as we expected, gene ontology enrichment tests revealed that the 1105 genes were aggregating not only in families but also in functional categories. PMID:18466548

  20. Correlating chemical sensitivity and basal gene expression reveals mechanism of action | Office of Cancer Genomics

    Cancer.gov

    Changes in cellular gene expression in response to small-molecule or genetic perturbations have yielded signatures that can connect unknown mechanisms of action (MoA) to ones previously established. We hypothesized that differential basal gene expression could be correlated with patterns of small-molecule sensitivity across many cell lines to illuminate the actions of compounds whose MoA are unknown.

  1. Differentially expressed genes in giant cell tumor of bone.

    PubMed

    Babeto, Erica; Conceição, André Luis Giacometti; Valsechi, Marina Curado; Peitl Junior, Paulo; de Campos Zuccari, Débora Aparecida Pires; de Lima, Luiz Guilherme Cernaglia Aureliano; Bonilha, Jane Lopes; de Freitas Calmon, Marília; Cordeiro, José Antônio; Rahal, Paula

    2011-04-01

    Giant cells tumors of bone (GCTB) are benign in nature but cause osteolytic destruction with a number of particular characteristics. These tumors can have uncertain biological behavior often contain a significant proportion of highly multinucleated cells, and may show aggressive behavior. We have studied differential gene expression in GCTB that may give a better understanding of their physiopathology, and might be helpful in prognosis and treatment. Rapid subtractive hybridization (RaSH) was used to identify and measure novel genes that appear to be differentially expressed, including KTN1, NEB, ROCK1, and ZAK using quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry in the samples of GCTBs compared to normal bone tissue. Normal bone was used in the methodology RaSH for comparison with the GCTB in identification of differentially expressed genes. Functional annotation indicated that these genes are involved in cellular processes related to their tumor phenotype. The differential expression of KTN1, ROCK1, and ZAK was independently confirmed by qRT-PCR and immunohistochemistry. The expression of the KTN1 and ROCK1 genes were increased in samples by qRT-PCR and immunohistochemistry, and ZAK had reduced expression. Since ZAK have CpG islands in their promoter region and low expression in tumor tissue, their methylation pattern was analyzed by MSP-PCR. The genes identified KTN1, ROCK1, and ZAK may be responsible for loss of cellular homeostasis in GCTB since they are responsible for various functions related to tumorigenesis such as cell migration, cytoskeletal organization, apoptosis, and cell cycle control and thus may contribute at some stage in the process of formation and development of GCTB.

  2. The light gene of Drosophila melanogaster encodes a homologue of VPS41, a yeast gene involved in cellular-protein trafficking.

    PubMed

    Warner, T S; Sinclair, D A; Fitzpatrick, K A; Singh, M; Devlin, R H; Honda, B M

    1998-04-01

    Mutations in a number of genes affect eye colour in Drosophila melanogaster; some of these "eye-colour" genes have been shown to be involved in various aspects of cellular transport processes. In addition, combinations of viable mutant alleles of some of these genes, such as carnation (car) combined with either light (lt) or deep-orange (dor) mutants, show lethal interactions. Recently, dor was shown to be homologous to the yeast gene PEP3 (VPS18), which is known to be involved in intracellular trafficking. We have undertaken to extend our earlier work on the lt gene, in order to examine in more detail its expression pattern and to characterize its gene product via sequencing of a cloned cDNA. The gene appears to be expressed at relatively high levels in all stages and tissues examined, and shows strong homology to VPS41, a gene involved in cellular-protein trafficking in yeast and higher eukaryotes. Further genetic experiments also point to a role for lt in transport processes: we describe lethal interactions between viable alleles of lt and dor, as well as phenotypic interactions (reductions in eye pigment) between allels of lt and another eye-colour gene, garnet (g), whose gene product has close homology to a subunit of the human adaptor complex, AP-3.

  3. Methods for monitoring multiple gene expression

    SciTech Connect

    Berka, Randy; Bachkirova, Elena; Rey, Michael

    2012-05-01

    The present invention relates to methods for monitoring differential expression of a plurality of genes in a first filamentous fungal cell relative to expression of the same genes in one or more second filamentous fungal cells using microarrays containing Trichoderma reesei ESTs or SSH clones, or a combination thereof. The present invention also relates to computer readable media and substrates containing such array features for monitoring expression of a plurality of genes in filamentous fungal cells.

  4. Methods for monitoring multiple gene expression

    SciTech Connect

    Berka, Randy; Bachkirova, Elena; Rey, Michael

    2013-10-01

    The present invention relates to methods for monitoring differential expression of a plurality of genes in a first filamentous fungal cell relative to expression of the same genes in one or more second filamentous fungal cells using microarrays containing Trichoderma reesei ESTs or SSH clones, or a combination thereof. The present invention also relates to computer readable media and substrates containing such array features for monitoring expression of a plurality of genes in filamentous fungal cells.

  5. Methods for monitoring multiple gene expression

    DOEpatents

    Berka, Randy; Bachkirova, Elena; Rey, Michael

    2008-06-01

    The present invention relates to methods for monitoring differential expression of a plurality of genes in a first filamentous fungal cell relative to expression of the same genes in one or more second filamentous fungal cells using microarrays containing Trichoderma reesei ESTs or SSH clones, or a combination thereof. The present invention also relates to computer readable media and substrates containing such array features for monitoring expression of a plurality of genes in filamentous fungal cells.

  6. The role of TREX in gene expression and disease

    PubMed Central

    Heath, Catherine G.; Viphakone, Nicolas; Wilson, Stuart A.

    2016-01-01

    TRanscription and EXport (TREX) is a conserved multisubunit complex essential for embryogenesis, organogenesis and cellular differentiation throughout life. By linking transcription, mRNA processing and export together, it exerts a physiologically vital role in the gene expression pathway. In addition, this complex prevents DNA damage and regulates the cell cycle by ensuring optimal gene expression. As the extent of TREX activity in viral infections, amyotrophic lateral sclerosis and cancer emerges, the need for a greater understanding of TREX function becomes evident. A complete elucidation of the composition, function and interactions of the complex will provide the framework for understanding the molecular basis for a variety of diseases. This review details the known composition of TREX, how it is regulated and its cellular functions with an emphasis on mammalian systems. PMID:27679854

  7. Ribozymes, riboswitches and beyond: regulation of gene expression without proteins

    PubMed Central

    Serganov, Alexander; Patel, Dinshaw J.

    2015-01-01

    Although various functions of RNA are carried out in conjunction with proteins, some catalytic RNAs, or ribozymes, which contribute to a range of cellular processes, require little or no assistance from proteins. Furthermore, the discovery of metabolite-sensing riboswitches and other types of RNA sensors has revealed RNA-based mechanisms that cells use to regulate gene expression in response to internal and external changes. Structural studies have shown how these RNAs can carry out a range of functions. In addition, the contribution of ribozymes and riboswitches to gene expression is being revealed as far more widespread than was previously appreciated. These findings have implications for understanding how cellular functions might have evolved from RNA-based origins. PMID:17846637

  8. Identification of Haemophilus ducreyi genes expressed during human infection.

    PubMed

    Bauer, Margaret E; Fortney, Kate R; Harrison, Alistair; Janowicz, Diane M; Munson, Robert S; Spinola, Stanley M

    2008-04-01

    To identify Haemophilus ducreyi transcripts that are expressed during human infection, we used selective capture of transcribed sequences (SCOTS) with RNA isolated from pustules obtained from three volunteers infected with H. ducreyi, and with RNA isolated from broth-grown bacteria used to infect volunteers. With SCOTS, competitive hybridization of tissue-derived and broth-derived sequences identifies genes that may be preferentially expressed in vivo. Among the three tissue specimens, we identified 531 genes expressed in vivo. Southern blot analysis of 60 genes from each tissue showed that 87 % of the identified genes hybridized better with cDNA derived from tissue specimens than with cDNA derived from broth-grown bacteria. RT-PCR on nine additional pustules confirmed in vivo expression of 10 of 11 selected genes in other volunteers. Of the 531 genes, 139 were identified in at least two volunteers. These 139 genes fell into several functional categories, including biosynthesis and metabolism, regulation, and cellular processes, such as transcription, translation, cell division, DNA replication and repair, and transport. Detection of genes involved in anaerobic and aerobic respiration indicated that H. ducreyi likely encounters both microenvironments within the pustule. Other genes detected suggest an increase in DNA damage and stress in vivo. Genes involved in virulence in other bacterial pathogens and 32 genes encoding hypothetical proteins were identified, and may represent novel virulence factors. We identified three genes, lspA1, lspA2 and tadA, known to be required for virulence in humans. This is the first study to broadly define transcripts expressed by H. ducreyi in humans.

  9. Leucine modulates the effect of Walker factor, a proteolysis-inducing factor-like protein from Walker tumours, on gene expression and cellular activity in C2C12 myotubes.

    PubMed

    Gonçalves, Estela Maria; Salomão, Emilianne Miguel; Gomes-Marcondes, Maria Cristina Cintra

    2013-10-01

    Cancer-cachexia causes severe weight loss, particularly from the wasting of skeletal muscle, which occurs due to increased protein catabolism and/or decreased protein synthesis. The muscle protein degradation observed in cancer patients is mediated by a specific cytokine, proteolysis-inducing factor (PIF), which is produced by the tumour. This protein increases the ubiquitin-proteasome pathway activity, and the synthesis of muscle protein in these patients can be affected by several factors, including nutrient-related signalling. Some nutrients, such as leucine, can decrease the ubiquitin-proteasome pathway activity and increase the skeletal muscle protein content in cachectic animals. In this study, we investigated the effects of leucine on cell viability, morphology, functional proteasome activity, enzymatic activity, and protein synthesis and degradation in C2C12 myotubes exposed to the proteolysis-inducing factor (PIF)-like protein purified from Walker tumour-bearing rats. Walker factor (WF) had no cytotoxic effects on myotube cells and morphological characteristics were not altered in the presence of WF and/or leucine. However, increased alkaline phosphatase activity was observed. At higher WF concentrations, chymotrypsin-like activity, cathepsin B activity and 20S proteasome gene expression increased. Treating myotubes with leucine before exposure to WF causes leads to a decrease in proteasome activity as well as the activity of chymotrypsin and cathepsin enzymes. Total protein synthesis decreased in WF-treated cells concomitantly as protein degradation increased. After leucine exposure, the observed effects of WF were minimal or even reverted in some cases. Taken together, these results suggest an important modulatory effect for leucine on the effects of WF in C2C12 myotube cells.

  10. Producing a Mammalian GFP Expression Vector Containing Neomycin Resistance Gene.

    PubMed

    Izadi, Manizheh; Abiri, Maryam; Keramatipour, Mohammad

    2009-04-01

    The green fluorescent protein (GFP) was originally isolated from the Jellyfish Aequorea Victoria that fluoresces green when exposed to blue light. GFP protein is composed of 238 amino acids with the molecular mass of 26.9 kD. The GFP gene is frequently used in cellular and molecular biology as a reporter gene. To date, many bacterial, yeast, fungal, plants, fly and mammalian cells, including human, have been created which express GFP. Martin Chalfie, Osamu Shimomura, and Roger Tsien were awarded the 2008 noble prize in chemistry for their discovery and development of GFP. In many studies on mammalian cells, GFP gene is introduced into cells using vector-based systems or a recombinant virus to track the location of a target protein or to study the expression level of the gene of interest, but in these studies there is no selection marker to normalize transfection. According to the importance of neomycin gene as a selection marker in mammalian cells, we aimed to produce a GFP expression vector that contains neomycin gene. GFP gene was separated from pEGFP-N1 vector and was inserted in the back-bone of pCDNA3.1/His/LacZ vector that contained the neomycin gene. The resulted vector contained GFP beside neomycin gene.

  11. VESPUCCI: Exploring Patterns of Gene Expression in Grapevine

    PubMed Central

    Moretto, Marco; Sonego, Paolo; Pilati, Stefania; Malacarne, Giulia; Costantini, Laura; Grzeskowiak, Lukasz; Bagagli, Giorgia; Grando, Maria Stella; Moser, Claudio; Engelen, Kristof

    2016-01-01

    Large-scale transcriptional studies aim to decipher the dynamic cellular responses to a stimulus, like different environmental conditions. In the era of high-throughput omics biology, the most used technologies for these purposes are microarray and RNA-Seq, whose data are usually required to be deposited in public repositories upon publication. Such repositories have the enormous potential to provide a comprehensive view of how different experimental conditions lead to expression changes, by comparing gene expression across all possible measured conditions. Unfortunately, this task is greatly impaired by differences among experimental platforms that make direct comparisons difficult. In this paper, we present the Vitis Expression Studies Platform Using COLOMBOS Compendia Instances (VESPUCCI), a gene expression compendium for grapevine which was built by adapting an approach originally developed for bacteria, and show how it can be used to investigate complex gene expression patterns. We integrated nearly all publicly available microarray and RNA-Seq expression data: 1608 gene expression samples from 10 different technological platforms. Each sample has been manually annotated using a controlled vocabulary developed ad hoc to ensure both human readability and computational tractability. Expression data in the compendium can be visually explored using several tools provided by the web interface or can be programmatically accessed using the REST interface. VESPUCCI is freely accessible at http://vespucci.colombos.fmach.it. PMID:27242836

  12. Heterosis and differential gene expression in hybrids and parents in Bombyx mori by digital gene expression profiling.

    PubMed

    Wang, Hua; Fang, Yan; Wang, Lipeng; Zhu, Wenjuan; Ji, Haipeng; Wang, Haiying; Xu, Shiqing; Sima, Yanghu

    2015-03-04

    Heterosis is a concern to all breeders, but the mechanism of heterosis remains unknown. In F1 organisms, genetic material is inherited from the two parents and theoretically, heterosis might be caused by differences in gene expression or modification. Differential gene expression was analyzed in hybrids and parents in Bombyx mori. The results showed that there were significant changes in gene expression in the fat body involving biological regulation, cellular and metabolic processes. Consistent trends in expression patterns covering different hybrid combinations were seen in 74 genes. Moreover, these differential gene expression patterns included overdominance, dominance, and additive effects. By correlating these patterns with economic traits, a potential relationship was found. Differential gene expression was seen in different cross combinations and in different sexes. In addition, a regulatory mechanism involving metabolism and ErbB signaling pathways was also found, suggesting that such a network might also be related to heterosis in Bombyx mori. Together, our data provide a comprehensive overview and useful resource for transcriptional analysis of heterosis of Bombyx mori.

  13. Heterosis and differential gene expression in hybrids and parents in Bombyx mori by digital gene expression profiling

    PubMed Central

    Wang, Hua; Fang, Yan; Wang, Lipeng; Zhu, Wenjuan; Ji, Haipeng; Wang, Haiying; Xu, Shiqing; Sima, Yanghu

    2015-01-01

    Heterosis is a concern to all breeders, but the mechanism of heterosis remains unknown. In F1 organisms, genetic material is inherited from the two parents and theoretically, heterosis might be caused by differences in gene expression or modification. Differential gene expression was analyzed in hybrids and parents in Bombyx mori. The results showed that there were significant changes in gene expression in the fat body involving biological regulation, cellular and metabolic processes. Consistent trends in expression patterns covering different hybrid combinations were seen in 74 genes. Moreover, these differential gene expression patterns included overdominance, dominance, and additive effects. By correlating these patterns with economic traits, a potential relationship was found. Differential gene expression was seen in different cross combinations and in different sexes. In addition, a regulatory mechanism involving metabolism and ErbB signaling pathways was also found, suggesting that such a network might also be related to heterosis in Bombyx mori. Together, our data provide a comprehensive overview and useful resource for transcriptional analysis of heterosis of Bombyx mori. PMID:25736158

  14. Gene expression control by Bacillus anthracis purine riboswitches.

    PubMed

    Kirchner, Marion; Schneider, Sabine

    2017-02-16

    In all kingdoms of life, cellular replication relies on the presence of nucleosides and nucleotides, the building blocks of nucleic acids and the main source of energy. In bacteria, the availability of metabolites sometimes directly regulates the expression of enzymes and proteins involved in purine salvage, biosynthesis and uptake through riboswitches. Riboswitches are located in bacterial mRNAs and can control gene expression by conformational changes in response to ligand binding. We have established an inverse reporter gene system in Bacillus subtilis that allows us to monitor riboswitch-controlled gene expression. We used it to investigate the activity of five potential purine riboswitches from B. anthracis in response to different purines and pyrimidines. Furthermore, in vitro studies on the aptamer domains of the riboswitches reveal their variation in guanine binding affinity ranging from nM to µM. These data do not only provide insight into metabolite sensing but can also aid to engineer artificial cell regulatory systems.

  15. Baicalin increases developmental competence of mouse embryos in vitro by inhibiting cellular apoptosis and modulating HSP70 and DNMT expression

    PubMed Central

    QI, Xiaonan; LI, Huatao; CONG, Xia; WANG, Xin; JIANG, Zhongling; CAO, Rongfeng; TIAN, Wenru

    2016-01-01

    Scutellaria baicalensis has been effectively used in Chinese traditional medicine to prevent miscarriages. However, little information is available on its mechanism of action. This study is designed specifically to reveal how baicalin, the main effective ingredient of S. baicalensis, improves developmental competence of embryos in vitro, using the mouse as a model. Mouse pronuclear embryos were cultured in KSOM medium supplemented with (0, 2, 4 and 8 μg/ml) baicalin. The results demonstrated that in vitro culture conditions significantly decreased the blastocyst developmental rate and blastocyst quality, possibly due to increased cellular stress and apoptosis. Baicalin (4 µg/ml) significantly increased 2- and 4-cell cleavage rates, morula developmental rate, and blastocyst developmental rate and cell number of in vitro-cultured mouse embryos. Moreover, baicalin increased the expression of Gja1, Cdh1, Bcl-2, and Dnmt3a genes, decreased the expression of Dnmt1 gene, and decreased cellular stress and apoptosis as it decreased the expression of HSP70, CASP3, and BAX and increased BCL-2 expression in blastocysts cultured in vitro. In conclusion, baicalin improves developmental competence of in vitro-cultured mouse embryos through inhibition of cellular apoptosis and HSP70 expression, and improvement of DNA methylation. PMID:27478062

  16. Lentiviral Gene Therapy Using Cellular Promoters Cures Type 1 Gaucher Disease in Mice

    PubMed Central

    Dahl, Maria; Doyle, Alexander; Olsson, Karin; Månsson, Jan-Eric; Marques, André R A; Mirzaian, Mina; Aerts, Johannes M; Ehinger, Mats; Rothe, Michael; Modlich, Ute; Schambach, Axel; Karlsson, Stefan

    2015-01-01

    Gaucher disease is caused by an inherited deficiency of the enzyme glucosylceramidase. Due to the lack of a fully functional enzyme, there is progressive build-up of the lipid component glucosylceramide. Insufficient glucosylceramidase activity results in hepatosplenomegaly, cytopenias, and bone disease in patients. Gene therapy represents a future therapeutic option for patients unresponsive to enzyme replacement therapy and lacking a suitable bone marrow donor. By proof-of-principle experiments, we have previously demonstrated a reversal of symptoms in a murine disease model of type 1 Gaucher disease, using gammaretroviral vectors harboring strong viral promoters to drive glucosidase β-acid (GBA) gene expression. To investigate whether safer vectors can correct the enzyme deficiency, we utilized self-inactivating lentiviral vectors (SIN LVs) with the GBA gene under the control of human phosphoglycerate kinase (PGK) and CD68 promoter, respectively. Here, we report prevention of, as well as reversal of, manifest disease symptoms after lentiviral gene transfer. Glucosylceramidase activity above levels required for clearance of glucosylceramide from tissues resulted in reversal of splenomegaly, reduced Gaucher cell infiltration and a restoration of hematological parameters. These findings support the use of SIN-LVs with cellular promoters in future clinical gene therapy protocols for type 1 Gaucher disease. PMID:25655314

  17. Lentiviral gene therapy using cellular promoters cures type 1 Gaucher disease in mice.

    PubMed

    Dahl, Maria; Doyle, Alexander; Olsson, Karin; Månsson, Jan-Eric; Marques, André R A; Mirzaian, Mina; Aerts, Johannes M; Ehinger, Mats; Rothe, Michael; Modlich, Ute; Schambach, Axel; Karlsson, Stefan

    2015-05-01

    Gaucher disease is caused by an inherited deficiency of the enzyme glucosylceramidase. Due to the lack of a fully functional enzyme, there is progressive build-up of the lipid component glucosylceramide. Insufficient glucosylceramidase activity results in hepatosplenomegaly, cytopenias, and bone disease in patients. Gene therapy represents a future therapeutic option for patients unresponsive to enzyme replacement therapy and lacking a suitable bone marrow donor. By proof-of-principle experiments, we have previously demonstrated a reversal of symptoms in a murine disease model of type 1 Gaucher disease, using gammaretroviral vectors harboring strong viral promoters to drive glucosidase β-acid (GBA) gene expression. To investigate whether safer vectors can correct the enzyme deficiency, we utilized self-inactivating lentiviral vectors (SIN LVs) with the GBA gene under the control of human phosphoglycerate kinase (PGK) and CD68 promoter, respectively. Here, we report prevention of, as well as reversal of, manifest disease symptoms after lentiviral gene transfer. Glucosylceramidase activity above levels required for clearance of glucosylceramide from tissues resulted in reversal of splenomegaly, reduced Gaucher cell infiltration and a restoration of hematological parameters. These findings support the use of SIN-LVs with cellular promoters in future clinical gene therapy protocols for type 1 Gaucher disease.

  18. Estimation and Testing of Gene Expression Heterosis

    PubMed Central

    Liu, Peng; Nettleton, Dan

    2014-01-01

    Heterosis, also known as the hybrid vigor, occurs when the mean phenotype of hybrid off-spring is superior to that of its two inbred parents. The heterosis phenomenon is extensively utilized in agriculture though the molecular basis is still unknown. In an effort to understand phenotypic heterosis at the molecular level, researchers have begun to compare expression levels of thousands of genes between parental inbred lines and their hybrid offspring to search for evidence of gene expression heterosis. Standard statistical approaches for separately analyzing expression data for each gene can produce biased and highly variable estimates and unreliable tests of heterosis. To address these shortcomings, we develop a hierarchical model to borrow information across genes. Using our modeling framework, we derive empirical Bayes estimators and an inference strategy to identify gene expression heterosis. Simulation results show that our proposed method outperforms the more traditional strategy used to detect gene expression heterosis. This article has supplementary material online. PMID:25435758

  19. Estimation and Testing of Gene Expression Heterosis.

    PubMed

    Ji, Tieming; Liu, Peng; Nettleton, Dan

    2014-09-01

    Heterosis, also known as the hybrid vigor, occurs when the mean phenotype of hybrid off-spring is superior to that of its two inbred parents. The heterosis phenomenon is extensively utilized in agriculture though the molecular basis is still unknown. In an effort to understand phenotypic heterosis at the molecular level, researchers have begun to compare expression levels of thousands of genes between parental inbred lines and their hybrid offspring to search for evidence of gene expression heterosis. Standard statistical approaches for separately analyzing expression data for each gene can produce biased and highly variable estimates and unreliable tests of heterosis. To address these shortcomings, we develop a hierarchical model to borrow information across genes. Using our modeling framework, we derive empirical Bayes estimators and an inference strategy to identify gene expression heterosis. Simulation results show that our proposed method outperforms the more traditional strategy used to detect gene expression heterosis. This article has supplementary material online.

  20. cDNA macroarray analysis of gene expression in synoviocytes stimulated with TNFalpha.

    PubMed

    Sugiyama, Tomoyasu; Ishii, Shizuko; Yamamoto, Jun ichi; Irie, Ryotaro; Saito, Kaoru; Otuki, Tetsuji; Wakamatsu, Ai; Suzuki, Yuzuru; Hio, Yuri; Ota, Toshio; Nishikawa, Tetsuo; Sugano, Sumio; Masuho, Yasuhiko; Isogai, Takao

    2002-04-24

    Gene expression of synoviocytes stimulated with tumor necrosis factor-alpha (TNFalpha) was studied by macroarray analysis to elucidate the cellular response and identify new biological functions of known and unknown genes. 10035 cDNA clones were used to make cDNA macroarrays of representative genes. Synoviocytes expressed large amounts of fibronectin and collagen mRNA. Statistical analysis of the macroarray data revealed 26 genes, including six new genes, which underwent significant alteration of gene expression in response to TNFalpha stimulation. These findings suggest that the synoviocyte response to TNFalpha stimulation forms the basis of development of various aspects of the pathophysiology of rheumatoid arthritis.

  1. Gene expression profiles of bronchoalveolar cells in Pulmonary TB

    PubMed Central

    Raju, Bindu; Hoshino, Yoshihiko; Belitskaya-Lévy, Ilana; Dawson, Rod; Ress, Stanley; Gold, Jeffrey A.; Condos, Rany; Pine, Richard; Brown, Stuart; Nolan, Anna; Rom, William N.; Weiden, Michael D.

    2008-01-01

    The host response to Mycobacterium tuberculosis includes macrophage activation, inflammation with increased immune effector cells, tissue necrosis and cavity formation, and fibrosis, distortion, and bronchiectasis. To evaluate the molecular basis of the immune response in the lungs of patients with active pulmonary tuberculosis (TB), we used bronchoalveolar lavage to obtain cells at the site of infection. Affymetrix Genechip micro-arrays and cDNA nylon filter microarrays interrogated gene expression in BAL cells from 11 healthy controls and 17 patients with active pulmonary TB. We found altered gene expression for 69 genes in TB versus normal controls that included cell surface markers, cytokines, chemokines, receptors, transcription factors, and complement components. In addition, TB BAL cell gene expression patternssegregated into 2 groups: one suggestive of a T helper type 1 (Th1) cellular immune response with increased STAT-4, IFN-γ receptor, and MIG expression with increased IFN-γ protein levels in BAL fluid; the other group displayed characteristics of Th2 immunity with increased STAT-6, CD81, and IL-10 receptor expression. We were able to demonstrate that a Th2 presentation could change to a Th1 pattern after anti-tuberculous treatment in one TB patient studied serially. These gene expression data support the conclusion that pulmonary TB produces a global change in the BAL cell transcriptome with manifestations of either Th1 or Th2 immunity. PMID:17921069

  2. Identification of Differentially Expressed Genes Between Osteoblasts and Osteocytes

    PubMed Central

    Paic, Frane; Igwe, John C.; Ravi, Nori; Kronenberg, Mark S.; Franceschetti, Tiziana; Harrington, Patrick; Kuo, Lynn; Shin, Don-Guk; Rowe, David W.; Harris, Stephen E.; Kalajzic, Ivo

    2009-01-01

    Osteocytes represent the most abundant cellular component of mammalian bones with important functions in bone mass maintenance and remodeling. To elucidate the differential gene expression between osteoblasts and osteocytes we completed a comprehensive analysis of their gene profiles. Selective identification of these two mature populations was achieved by utilization of visual markers of bone lineage cells. We have utilized dual GFP reporter mice in which osteocytes are expressing GFP (topaz) directed by the DMP1 promoter, while osteoblasts are identified by expression of GFP (cyan) driven by 2.3kb of the Col1a1 promoter. Histological analysis of 7-day-old neonatal calvaria confirmed the expression pattern of DMP1GFP in osteocytes and Col2.3 in osteoblasts and osteocytes. To isolate distinct populations of cells we utilized fluorescent activated cell sorting (FACS). Cells suspensions were subjected to RNA extraction, in vitro transcription and labeling of cDNA and gene expression was analyzed using the Illumina WG-6v1 BeadChip. Following normalization of raw data from four biological replicates, 3444 genes were called present in all three sorted cell populations: GFP negative, Col2.3cyan+ (osteoblasts), and DMP1topaz+(preosteocytes and osteocytes). We present the genes that showed in excess of a 2-fold change for gene expression between DMP1topaz+ and Col2.3cyan+ cells. The selected genes were classified and grouped according to their associated gene ontology terms. Genes clustered to osteogenesis and skeletal development such as Bmp4, Bmp8a, Dmp1, Enpp1, Phex and Ank were highly expressed in DMP1topaz+cells. Most of the genes encoding extracellular matrix components and secreted proteins had lower expression in DMP1topaz+ cells, while most of the genes encoding plasma membrane proteins were increased. Interestingly a large number of genes associated with muscle development and function and with neuronal phenotype were increased in DMP1topaz+ cells, indicating

  3. Comparison between the viral transforming gene (src) of recovered avian sarcoma virus and its cellular homolog.

    PubMed Central

    Takeya, T; Hanafusa, H; Junghans, R P; Ju, G; Skalka, A M

    1981-01-01

    Recovered avian sarcoma viruses are recombinants between transformation-defective mutants of Rous sarcoma virus and the chicken cellular gene homologous to the src gene of Rous sarcoma virus. We have constructed and analyzed molecular clones of viral deoxyribonucleic acid from recovered avian sarcoma virus and its transformation-competent progenitor, the Schmidt-Ruppin A strain of Rous sarcoma virus. A 2.0-megadalton EcoRI fragment containing the entire src gene from each of these clones was subcloned and characterized. These fragments were also used as probes to isolate recombinant phage clones containing the cellular counterpart of the viral src gene, termed cellular src, from a lambda library of chicken deoxyribonucleic acid. The structure of cellular src was analyzed by restriction endonuclease mapping and electron microscopy. Restriction endonuclease mapping revealed extensive similarity between the src regions of Rous sarcoma virus and recovered avian sarcoma virus, but striking differences between the viral src's and cellular src. Electron microscopic analysis of heteroduplexes between recovered virus src and cellular src revealed a 1.8-kilobase region of homology. In the cellular gene, the homologous region was interrupted by seven nonhomologous regions which we interpret to be intervening sequences. We estimate the minimum length of cellular src to be about 7.2 kilobases. These findings have implications concerning the mechanism of formation of recovered virus src and possibly other cell-derived retrovirus transforming genes. Images PMID:6287213

  4. Evolution of viruses by acquisition of cellular RNA or DNA nucleotide sequences and genes: an introduction.

    PubMed

    Becker, Y

    2000-01-01

    The origins of virus evolution may be traced to Archeabacteria since Inouye and Inouye (6) discovered a retroelement with a gene for reverse transcriptase in the bacterial genome and in the satellite, multiple copy single stranded DNA (msDNA) in the soil bacterium Myxococcus xanthus. It was possible (8) to define the evolution of retroelements in eukaryotic cells of plants, insects (gypsy retrovirus) and vertebrates. The replication of RNA viruses in eukaryotic cells allowed for the viral RNA genome to integrate a cellular ubiquitin mRNA, as reported for BVDV (24). Another example is the integration of 28S ribosomal RNA into the hemagglutinin gene of an influenza virus. This change in the hemagglutinin gene led to an increased pathogenicity of the influenza virus (25). In contrast to RNA viruses, DNA viruses had evolved by inserting cDNA molecules derived from mRNA transcripts of cellular genes or foreign viral RNA. It is of interest that the virus acquired cellular genes in the genomes of DNA viruses represent genes that code for proteins that inhibit cellular molecular processes related to HLA class I and II molecules. The other acquired genes are cellular genes that code for cytokines that are capable of inhibiting antigen presentation to T cells by antigen presenting cells (APC) by dendritic Langerhans cells. The acquisition of cellular genes by DNA viruses enhances their pathogenicity by inhibiting the hosts' defense systems.

  5. HMGA2 expression in white adipose tissue linking cellular senescence with diabetes.

    PubMed

    Markowski, Dominique Nadine; Thies, Helge Wilhelm; Gottlieb, Andrea; Wenk, Heiner; Wischnewsky, Manfred; Bullerdiek, Jörn

    2013-09-01

    There is a clear link between overweight, gain of white adipose tissue, and diabetes type 2 (T2D). The molecular mechanism of the gain of adipose tissue is linked with the expression of high mobility group protein AT-hook 2 (HMGA2), and recent studies revealed an association with a SNP near HMGA2. In this study, we investigated the gene expression of HMGA2, p14 (Arf) , CDKN1A, and BAX in human abdominal subcutaneous white adipose tissue from 157 patients. We found a significant higher HMGA2 expression in obese individuals than in non-obese patients. Furthermore, the HMGA2 expression in white adipose tissue in patient with type 2 diabetes was significantly higher than in nondiabetic patients. There is an association between the DNA-binding nonhistone protein HMGA2 and the risk of developing T2D that remains mechanistically unexplained so far. Likewise, p14(Arf), an inducer of cellular senescence, has been associated with the occurrence of T2D. The data of the present study provide evidence that both proteins act within the same network to drive proliferation of adipose tissue stem and precursor cells, senescence, and increased risk of T2D, respectively.

  6. Independent component analysis of Alzheimer's DNA microarray gene expression data

    PubMed Central

    Kong, Wei; Mou, Xiaoyang; Liu, Qingzhong; Chen, Zhongxue; Vanderburg, Charles R; Rogers, Jack T; Huang, Xudong

    2009-01-01

    Background Gene microarray technology is an effective tool to investigate the simultaneous activity of multiple cellular pathways from hundreds to thousands of genes. However, because data in the colossal amounts generated by DNA microarray technology are usually complex, noisy, high-dimensional, and often hindered by low statistical power, their exploitation is difficult. To overcome these problems, two kinds of unsupervised analysis methods for microarray data: principal component analysis (PCA) and independent component analysis (ICA) have been developed to accomplish the task. PCA projects the data into a new space spanned by the principal components that are mutually orthonormal to each other. The constraint of mutual orthogonality and second-order statistics technique within PCA algorithms, however, may not be applied to the biological systems studied. Extracting and characterizing the most informative features of the biological signals, however, require higher-order statistics. Results ICA is one of the unsupervised algorithms that can extract higher-order statistical structures from data and has been applied to DNA microarray gene expression data analysis. We performed FastICA method on DNA microarray gene expression data from Alzheimer's disease (AD) hippocampal tissue samples and consequential gene clustering. Experimental results showed that the ICA method can improve the clustering results of AD samples and identify significant genes. More than 50 significant genes with high expression levels in severe AD were extracted, representing immunity-related protein, metal-related protein, membrane protein, lipoprotein, neuropeptide, cytoskeleton protein, cellular binding protein, and ribosomal protein. Within the aforementioned categories, our method also found 37 significant genes with low expression levels. Moreover, it is worth noting that some oncogenes and phosphorylation-related proteins are expressed in low levels. In comparison to the PCA and support

  7. Gene Expression Patterns in Ovarian Carcinomas

    PubMed Central

    Schaner, Marci E.; Ross, Douglas T.; Ciaravino, Giuseppe; Sørlie, Therese; Troyanskaya, Olga; Diehn, Maximilian; Wang, Yan C.; Duran, George E.; Sikic, Thomas L.; Caldeira, Sandra; Skomedal, Hanne; Tu, I-Ping; Hernandez-Boussard, Tina; Johnson, Steven W.; O'Dwyer, Peter J.; Fero, Michael J.; Kristensen, Gunnar B.; Børresen-Dale, Anne-Lise; Hastie, Trevor; Tibshirani, Robert; van de Rijn, Matt; Teng, Nelson N.; Longacre, Teri A.; Botstein, David; Brown, Patrick O.; Sikic, Branimir I.

    2003-01-01

    We used DNA microarrays to characterize the global gene expression patterns in surface epithelial cancers of the ovary. We identified groups of genes that distinguished the clear cell subtype from other ovarian carcinomas, grade I and II from grade III serous papillary carcinomas, and ovarian from breast carcinomas. Six clear cell carcinomas were distinguished from 36 other ovarian carcinomas (predominantly serous papillary) based on their gene expression patterns. The differences may yield insights into the worse prognosis and therapeutic resistance associated with clear cell carcinomas. A comparison of the gene expression patterns in the ovarian cancers to published data of gene expression in breast cancers revealed a large number of differentially expressed genes. We identified a group of 62 genes that correctly classified all 125 breast and ovarian cancer specimens. Among the best discriminators more highly expressed in the ovarian carcinomas were PAX8 (paired box gene 8), mesothelin, and ephrin-B1 (EFNB1). Although estrogen receptor was expressed in both the ovarian and breast cancers, genes that are coregulated with the estrogen receptor in breast cancers, including GATA-3, LIV-1, and X-box binding protein 1, did not show a similar pattern of coexpression in the ovarian cancers. PMID:12960427

  8. Arabidopsis gene expression patterns during spaceflight

    NASA Astrophysics Data System (ADS)

    Paul, A.-L.; Ferl, R. J.

    The exposure of Arabidopsis thaliana (Arabidopsis) plants to spaceflight environments resulted in the differential expression of hundreds of genes. A 5 day mission on orbiter Columbia in 1999 (STS-93) carried transgenic Arabidopsis plants engineered with a transgene composed of the alcohol dehydrogenase (Adh) gene promoter linked to the β -Glucuronidase (GUS) reporter gene. The plants were used to evaluate the effects of spaceflight on two fronts. First, expression patterns visualized with the Adh/GUS transgene were used to address specifically the possibility that spaceflight induces a hypoxic stress response, and to assess whether any spaceflight response was similar to control terrestrial hypoxia-induced gene expression patterns. (Paul et al., Plant Physiol. 2001, 126:613). Second, genome-wide patterns of native gene expression were evaluated utilizing the Affymetrix ATH1 GeneChip? array of 8,000 Arabidopsis genes. As a control for the veracity of the array analyses, a selection of genes identified with the arrays was further characterized with quantitative Real-Time RT PCR (ABI - TaqmanTM). Comparison of the patterns of expression for arrays of hybridized with RNA isolated from plants exposed to spaceflight compared to the control arrays revealed hundreds of genes that were differentially expressed in response to spaceflight, yet most genes that are hallmarks of hypoxic stress were unaffected. These results will be discussed in light of current models for plant responses to the spaceflight environment, and with regard to potential future flight opportunities.

  9. Effect of adenovirus infection on expression of human histone genes.

    PubMed Central

    Flint, S J; Plumb, M A; Yang, U C; Stein, G S; Stein, J L

    1984-01-01

    The influence of adenovirus type 2 infection of HeLa cells upon expression of human histone genes was examined as a function of the period of infection. Histone RNA synthesis was assayed after run-off transcription in nuclei isolated from mock-infected cells and after various periods of adenovirus infection. Histone protein synthesis was measured by [3H]leucine labeling of intact cells and fluorography of electrophoretically fractionated nuclear and cytoplasmic proteins. The cellular representation of RNA species complementary to more than 13 different human histone genes was determined by RNA blot analysis of total cellular, nuclear or cytoplasmic RNA by using a series of 32P-labeled cloned human histone genes as hybridization probes and also by analysis of 3H-labeled histone mRNA species synthesized in intact cells. By 18 h after infection, HeLa cell DNA synthesis and all parameters of histone gene expression, including transcription and the nuclear and cytoplasmic concentrations of core and H1 mRNA species, were reduced to less than 5 to 10% of the control values. By contrast, transcription and processing of other cellular mRNA sequences have been shown to continue throughout this period of infection. The early period of adenovirus infection was marked by an inhibition of transcription of histone genes that accompanied the reduction in rate of HeLa cell DNA synthesis. These results suggest that the adenovirus-induced inhibition of histone gene expression is mediated in part at the transcriptional level. However, the persistence of histone mRNA species at concentrations comparable to those of mock-infected control cells during the early phase of the infection, despite a reduction in histone gene transcription and histone protein synthesis, implies that histone gene expression is also regulated post-transcriptionally in adenovirus-infected cells. These results suggest that the tight coupling between histone mRNA concentrations and the rate of cellular DNA

  10. Stratified gene expression analysis identifies major amyotrophic lateral sclerosis genes.

    PubMed

    Jones, Ashley R; Troakes, Claire; King, Andrew; Sahni, Vibhu; De Jong, Simone; Bossers, Koen; Papouli, Efterpi; Mirza, Muddassar; Al-Sarraj, Safa; Shaw, Christopher E; Shaw, Pamela J; Kirby, Janine; Veldink, Jan H; Macklis, Jeffrey D; Powell, John F; Al-Chalabi, Ammar

    2015-05-01

    Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease of motor neurons resulting in progressive paralysis. Gene expression studies of ALS only rarely identify the same gene pathways as gene association studies. We hypothesized that analyzing tissues by matching on degree of disease severity would identify different patterns of gene expression from a traditional case-control comparison. We analyzed gene expression changes in 4 postmortem central nervous system regions, stratified by severity of motor neuron loss. An overall comparison of cases (n = 6) and controls (n = 3) identified known ALS gene, SOX5, as showing differential expression (log2 fold change = 0.09, p = 5.5 × 10(-5)). Analyses stratified by disease severity identified expression changes in C9orf72 (p = 2.77 × 10(-3)), MATR3 (p = 3.46 × 10(-3)), and VEGFA (p = 8.21 × 10(-4)), all implicated in ALS through genetic studies, and changes in other genes in pathways involving RNA processing and immune response. These findings suggest that analysis of gene expression stratified by disease severity can identify major ALS genes and may be more efficient than traditional case-control comparison.

  11. A Genetic Approach to Promoter Recognition during Trans Induction of Viral Gene Expression

    NASA Astrophysics Data System (ADS)

    Coen, Donald M.; Weinheimer, Steven P.; McKnight, Steven L.

    1986-10-01

    Viral infection of mammalian cells entails the regulated induction of viral gene expression. The induction of many viral genes, including the herpes simplex virus gene encoding thymidine kinase (tk), depends on viral regulatory proteins that act in trans. Because recognition of the tk promoter by cellular transcription factors is well understood, its trans induction by viral regulatory proteins may serve as a useful model for the regulation of eukaryotic gene expression. A comprehensive set of mutations was therefore introduced into the chromosome of herpes simplex virus at the tk promoter to directly analyze the effects of promoter mutations on tk transcription. The promoter domains required for efficient tk expression under conditions of trans induction corresponded to those important for recognition by cellular transcription factors. Thus, trans induction of tk expression may be catalyzed initially by the interaction of viral regulatory proteins with cellular transcription factors.

  12. Evolution and Expression of Paxillin Genes in Teleost Fish

    PubMed Central

    Jacob, Andrew E.; Turner, Christopher E.; Amack, Jeffrey D.

    2016-01-01

    Background Paxillin family proteins regulate intracellular signaling downstream of extracellular matrix adhesion. Tissue expression patterns and cellular functions of Paxillin proteins during embryo development remain poorly understood. Additionally, the evolution of this gene family has not been thoroughly investigated. Results This report characterizes the evolution and expression of a novel Paxillin gene, called Paxillin-b, in Teleosts. Alignments indicate that Teleost Paxillin-a and Paxillin-b proteins are highly homologous to each other and to human Paxillin. Phylogenetic and synteny analyses suggest that these genes originated from the duplication of an ancestral Paxillin gene that was in a common ancestor of Teleosts and Tetrapods. Analysis of the spatiotemporal expression profiles of Paxillin-a and Paxillin-b using zebrafish revealed both overlapping and distinct domains for Paxillin-a and Paxillin-b during embryo development. Localization of zebrafish Paxillin orthologs expressed in mammalian cells demonstrated that both proteins localize to focal adhesions, similar to mammalian Paxillin. This suggests these proteins regulate adhesion-dependent processes in their endogenous tissues. Conclusion Paxillin-a and Paxillin-b were generated by duplication in Teleosts. These genes likely play similar roles as Paxillin genes in other organisms. This work provides a framework for functional investigation of Paxillin family members during development using the zebrafish as an in vivo model system. PMID:27806088

  13. GOAL: automated Gene Ontology analysis of expression profiles.

    PubMed

    Volinia, Stefano; Evangelisti, Rita; Francioso, Francesca; Arcelli, Diego; Carella, Massimo; Gasparini, Paolo

    2004-07-01

    One of the most common problems encountered while deciphering results from expression profiling experiments is in relating differential expression of genes to molecular functions and cellular processes. A second important problem is that of comparing experiments performed by different labs using different microarray platforms, or even unrelated techniques. Gene Ontology (GO) is now used to describe biological features, since GO terms are associated with genes, to overcome the apparent distance between expression profiles and biological comprehension. Here we describe the development, implementation and use of GOAL (Gene Ontology Automated Lexicon), a web-based application for the identification of functions and processes regulated in microarray and SAGE (serial analysis of gene expression) experiments. We applied GOAL to a range of experimental datasets related to different biological problems, including cancer and the cell cycle. By using GOAL, reported and novel relevant processes were identified in a number of experiments by our collaborators and by us. Different datasets could also be compared with each other to define conserved functional modules. GOAL allows a seamless and high-level analysis of expression profiles and is implemented as a free WWW resource (http://microarrays.unife.it).

  14. Monitoring Cellular Phosphorylation Signaling Pathways into Chromatin and Down to the Gene Level*

    PubMed Central

    Han, Yumiao; Yuan, Zuo-Fei; Molden, Rosalynn C.; Garcia, Benjamin A.

    2016-01-01

    Protein phosphorylation, one of the most common and important modifications of acute and reversible regulation of protein function, plays a dominant role in almost all cellular processes. These signaling events regulate cellular responses, including proliferation, differentiation, metabolism, survival, and apoptosis. Several studies have been successfully used to identify phosphorylated proteins and dynamic changes in phosphorylation status after stimulation. Nevertheless, it is still rather difficult to elucidate precise complex phosphorylation signaling pathways. In particular, how signal transduction pathways directly communicate from the outer cell surface through cytoplasmic space and then directly into chromatin networks to change the transcriptional and epigenetic landscape remains poorly understood. Here, we describe the optimization and comparison of methods based on thiophosphorylation affinity enrichment, which can be utilized to monitor phosphorylation signaling into chromatin by isolation of phosphoprotein containing nucleosomes, a method we term phosphorylation-specific chromatin affinity purification (PS-ChAP). We utilized this PS-ChAP1 approach in combination with quantitative proteomics to identify changes in the phosphorylation status of chromatin-bound proteins on nucleosomes following perturbation of transcriptional processes. We also demonstrate that this method can be employed to map phosphoprotein signaling into chromatin containing nucleosomes through identifying the genes those phosphorylated proteins are found on via thiophosphate PS-ChAP-qPCR. Thus, our results showed that PS-ChAP offers a new strategy for studying cellular signaling and chromatin biology, allowing us to directly and comprehensively investigate phosphorylation signaling into chromatin to investigate if these pathways are involved in altering gene expression. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium with the data set

  15. 76 FR 81513 - Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-28

    ... HUMAN SERVICES Food and Drug Administration Cellular, Tissue, and Gene Therapies Advisory Committee..., Tissue, and Gene Therapies Advisory Committee. General Function of the Committee: To provide advice and... Gene Therapies, Center for Biologics Evaluation and Research, FDA. FDA intends to make...

  16. A Weakened Transcriptional Enhancer Yields Variegated Gene Expression

    PubMed Central

    Collins, Cathy; Azmi, Peter; Berru, Maribel; Zhu, Xiaofu; Shulman, Marc J.

    2006-01-01

    Identical genes in the same cellular environment are sometimes expressed differently. In some cases, including the immunoglobulin heavy chain (IgH) locus, this type of differential gene expression has been related to the absence of a transcriptional enhancer. To gain additional information on the role of the IgH enhancer, we examined expression driven by enhancers that were merely weakened, rather than fully deleted, using both mutations and insulators to impair enhancer activity. For this purpose we used a LoxP/Cre system to place a reporter gene at the same genomic site of a stable cell line. Whereas expression of the reporter gene was uniformly high in the presence of the normal, uninsulated enhancer and undetectable in its absence, weakened enhancers yielded variegated expression of the reporter gene; i.e., the average level of expression of the same gene differed in different clones, and expression varied significantly among cells within individual clones. These results indicate that the weakened enhancer allows the reporter gene to exist in at least two states. Subtle aspects of the variegation suggest that the IgH enhancer decreases the average duration (half-life) of the silent state. This analysis has also tested the conventional wisdom that enhancer activity is independent of distance and orientation. Thus, our analysis of mutant (truncated) forms of the IgH enhancer revealed that the 250 bp core enhancer was active in its normal position, ∼1.4 kb 3′ of the promoter, but inactive ∼6 kb 3′, indicating that the activity of the core enhancer was distance-dependent. A longer segment – the core enhancer plus ∼1 kb of 3′ flanking material, including the 3′ matrix attachment region – was active, and the activity of this longer segment was orientation-dependent. Our data suggest that this 3′ flank includes binding sites for at least two activators. PMID:17183661

  17. Gene Expression Noise, Fitness Landscapes, and Evolution

    NASA Astrophysics Data System (ADS)

    Charlebois, Daniel

    The stochastic (or noisy) process of gene expression can have fitness consequences for living organisms. For example, gene expression noise facilitates the development of drug resistance by increasing the time scale at which beneficial phenotypic states can be maintained. The present work investigates the relationship between gene expression noise and the fitness landscape. By incorporating the costs and benefits of gene expression, we track how the fluctuation magnitude and timescale of expression noise evolve in simulations of cell populations under stress. We find that properties of expression noise evolve to maximize fitness on the fitness landscape, and that low levels of expression noise emerge when the fitness benefits of gene expression exceed the fitness costs (and that high levels of noise emerge when the costs of expression exceed the benefits). The findings from our theoretical/computational work offer new hypotheses on the development of drug resistance, some of which are now being investigated in evolution experiments in our laboratory using well-characterized synthetic gene regulatory networks in budding yeast. Nserc Postdoctoral Fellowship (Grant No. PDF-453977-2014).

  18. Gene Expression Dynamics Accompanying the Sponge Thermal Stress Response

    PubMed Central

    Guzman, Christine; Conaco, Cecilia

    2016-01-01

    Marine sponges are important members of coral reef ecosystems. Thus, their responses to changes in ocean chemistry and environmental conditions, particularly to higher seawater temperatures, will have potential impacts on the future of these reefs. To better understand the sponge thermal stress response, we investigated gene expression dynamics in the shallow water sponge, Haliclona tubifera (order Haplosclerida, class Demospongiae), subjected to elevated temperature. Using high-throughput transcriptome sequencing, we show that these conditions result in the activation of various processes that interact to maintain cellular homeostasis. Short-term thermal stress resulted in the induction of heat shock proteins, antioxidants, and genes involved in signal transduction and innate immunity pathways. Prolonged exposure to thermal stress affected the expression of genes involved in cellular damage repair, apoptosis, signaling and transcription. Interestingly, exposure to sublethal temperatures may improve the ability of the sponge to mitigate cellular damage under more extreme stress conditions. These insights into the potential mechanisms of adaptation and resilience of sponges contribute to a better understanding of sponge conservation status and the prediction of ecosystem trajectories under future climate conditions. PMID:27788197

  19. Gene Expression Measurement Module (GEMM) - a fully automated, miniaturized instrument for measuring gene expression in space

    NASA Astrophysics Data System (ADS)

    Karouia, Fathi; Ricco, Antonio; Pohorille, Andrew; Peyvan, Kianoosh

    2012-07-01

    The capability to measure gene expression on board spacecrafts opens the doors to a large number of experiments on the influence of space environment on biological systems that will profoundly impact our ability to conduct safe and effective space travel, and might also shed light on terrestrial physiology or biological function and human disease and aging processes. Measurements of gene expression will help us to understand adaptation of terrestrial life to conditions beyond the planet of origin, identify deleterious effects of the space environment on a wide range of organisms from microbes to humans, develop effective countermeasures against these effects, determine metabolic basis of microbial pathogenicity and drug resistance, test our ability to sustain and grow in space organisms that can be used for life support and in situ resource utilization during long-duration space exploration, and monitor both the spacecraft environment and crew health. These and other applications hold significant potential for discoveries in space biology, biotechnology and medicine. Accordingly, supported by funding from the NASA Astrobiology Science and Technology Instrument Development Program, we are developing a fully automated, miniaturized, integrated fluidic system for small spacecraft capable of in-situ measuring microbial expression of thousands of genes from multiple samples. The instrument will be capable of (1) lysing bacterial cell walls, (2) extracting and purifying RNA released from cells, (3) hybridizing it on a microarray and (4) providing electrochemical readout, all in a microfluidics cartridge. The prototype under development is suitable for deployment on nanosatellite platforms developed by the NASA Small Spacecraft Office. The first target application is to cultivate and measure gene expression of the photosynthetic bacterium Synechococcus elongatus, i.e. a cyanobacterium known to exhibit remarkable metabolic diversity and resilience to adverse conditions

  20. Sequential Logic Model Deciphers Dynamic Transcriptional Control of Gene Expressions

    PubMed Central

    Yeo, Zhen Xuan; Wong, Sum Thai; Arjunan, Satya Nanda Vel; Piras, Vincent; Tomita, Masaru; Selvarajoo, Kumar; Giuliani, Alessandro; Tsuchiya, Masa

    2007-01-01

    Background Cellular signaling involves a sequence of events from ligand binding to membrane receptors through transcription factors activation and the induction of mRNA expression. The transcriptional-regulatory system plays a pivotal role in the control of gene expression. A novel computational approach to the study of gene regulation circuits is presented here. Methodology Based on the concept of finite state machine, which provides a discrete view of gene regulation, a novel sequential logic model (SLM) is developed to decipher control mechanisms of dynamic transcriptional regulation of gene expressions. The SLM technique is also used to systematically analyze the dynamic function of transcriptional inputs, the dependency and cooperativity, such as synergy effect, among the binding sites with respect to when, how much and how fast the gene of interest is expressed. Principal Findings SLM is verified by a set of well studied expression data on endo16 of Strongylocentrotus purpuratus (sea urchin) during the embryonic midgut development. A dynamic regulatory mechanism for endo16 expression controlled by three binding sites, UI, R and Otx is identified and demonstrated to be consistent with experimental findings. Furthermore, we show that during transition from specification to differentiation in wild type endo16 expression profile, SLM reveals three binary activities are not sufficient to explain the transcriptional regulation of endo16 expression and additional activities of binding sites are required. Further analyses suggest detailed mechanism of R switch activity where indirect dependency occurs in between UI activity and R switch during specification to differentiation stage. Conclusions/Significance The sequential logic formalism allows for a simplification of regulation network dynamics going from a continuous to a discrete representation of gene activation in time. In effect our SLM is non-parametric and model-independent, yet providing rich biological

  1. Epigenetic determinants of cardiovascular gene expression: vascular endothelium.

    PubMed

    Man, Hon-Sum Jeffrey; Yan, Matthew S; Lee, John Jy; Marsden, Philip A

    2016-07-01

    The modern landscape of gene regulation involves interacting factors that ultimately lead to gene activation or repression. Epigenetic mechanisms provide a perspective of cellular phenotype as dynamically regulated and responsive to input. This perspective is supported by the generation of induced pluripotent stem cells from fully differentiated cell types. In vascular endothelial cells, evidence suggests that epigenetic mechanisms play a major role in the expression of endothelial cell-specific genes such as the endothelial nitric oxide synthase (NOS3/eNOS). These mechanisms are also important for eNOS expression in response to environmental stimuli such as hypoxia and shear stress. A newer paradigm in epigenetics, long noncoding RNAs offer a link between genetic variation, epigenetic regulation and disease. While the understanding of epigenetic mechanisms is early in its course, it is becoming clear that approaches to understanding the interaction of these factors and their inputs will be necessary to improve outcomes in cardiovascular disease.

  2. Interferon-Stimulated Gene 15 in the Control of Cellular Responses to Genotoxic Stress

    PubMed Central

    Jeon, Young Joo; Park, Jong Ho; Chung, Chin Ha

    2017-01-01

    Error-free replication and repair of DNA are pivotal to organisms for faithful transmission of their genetic information. Cells orchestrate complex signaling networks that sense and resolve DNA damage. Post-translational protein modifications by ubiquitin and ubiquitin-like proteins, including SUMO and NEDD8, are critically involved in DNA damage response (DDR) and DNA damage tolerance (DDT). The expression of interferon-stimulated gene 15 (ISG15), the first identified ubiquitin-like protein, has recently been shown to be induced under various DNA damage conditions, such as exposure to UV, camptothecin, and doxorubicin. Here we overview the recent findings on the role of ISG15 and its conjugation to target proteins (e.g., p53, ΔNp63α, and PCNA) in the control of cellular responses to genotoxic stress, such as the inhibition of cell growth and tumorigenesis. PMID:28241406

  3. Quantitative analysis of cell-type specific gene expression in the green alga Volvox carteri

    PubMed Central

    Nematollahi, Ghazaleh; Kianianmomeni, Arash; Hallmann, Armin

    2006-01-01

    Background The multicellular alga Volvox carteri possesses only two cell types: mortal, motile somatic cells and potentially immortal, immotile reproductive cells. It is therefore an attractive model system for studying how cell-autonomous cytodifferentiation is programmed within a genome. Moreover, there are ongoing genome projects both in Volvox carteri and in the closely related unicellular alga Chlamydomonas reinhardtii. However, gene sequencing is only the beginning. To identify cell-type specific expression and to determine relative expression rates, we evaluate the potential of real-time RT-PCR for quantifying gene transcript levels. Results Here we analyze a diversified pool of 39 target genes by real-time RT-PCR for each cell type. This gene pool contains previously known genes with unknown localization of cellular expression, 28 novel genes which are described in this study for the first time, and a few known, cell-type specific genes as a control. The respective gene products are, for instance, part of photosynthesis, cellular regulation, stress response, or transport processes. We provide expression data for all these genes. Conclusion The results show that quantitative real-time RT-PCR is a favorable approach to analyze cell-type specific gene expression in Volvox, which can be extended to a much larger number of genes or to developmental or metabolic mutants. Our expression data also provide a basis for a detailed analysis of individual, previously unknown, cell-type specifically expressed genes. PMID:17184518

  4. Expression profiling in spondyloarthropathy synovial biopsies highlights changes in expression of inflammatory genes in conjunction with tissue remodelling genes

    PubMed Central

    2013-01-01

    Background In the spondyloarthropathies, the underlying molecular and cellular pathways driving disease are poorly understood. By undertaking a study in knee synovial biopsies from spondyloarthropathy (SpA) and ankylosing spondylitis (AS) patients we aimed to elucidate dysregulated genes and pathways. Methods RNA was extracted from six SpA, two AS, three osteoarthritis (OA) and four normal control knee synovial biopsies. Whole genome expression profiling was undertaken using the Illumina DASL system, which assays 24000 cDNA probes. Differentially expressed candidate genes were then validated using quantitative PCR and immunohistochemistry. Results Four hundred and sixteen differentially expressed genes were identified that clearly delineated between AS/SpA and control groups. Pathway analysis showed altered gene-expression in oxidoreductase activity, B-cell associated, matrix catabolic, and metabolic pathways. Altered "myogene" profiling was also identified. The inflammatory mediator, MMP3, was strongly upregulated (5-fold) in AS/SpA samples and the Wnt pathway inhibitors DKK3 (2.7-fold) and Kremen1 (1.5-fold) were downregulated. Conclusions Altered expression profiling in SpA and AS samples demonstrates that disease pathogenesis is associated with both systemic inflammation as well as local tissue alterations that may underlie tissue damaging modelling and remodelling outcomes. This supports the hypothesis that initial systemic inflammation in spondyloarthropathies transfers to and persists in the local joint environment, and might subsequently mediate changes in genes directly involved in the destructive tissue remodelling. PMID:24330574

  5. Gene-expression signatures of Atlantic salmon's plastic life cycle

    USGS Publications Warehouse

    Aubin-Horth, N.; Letcher, B.H.; Hofmann, H.A.

    2009-01-01

    How genomic expression differs as a function of life history variation is largely unknown. Atlantic salmon exhibits extreme alternative life histories. We defined the gene-expression signatures of wild-caught salmon at two different life stages by comparing the brain expression profiles of mature sneaker males and immature males, and early migrants and late migrants. In addition to life-stage-specific signatures, we discovered a surprisingly large gene set that was differentially regulated-at similar magnitudes, yet in opposite direction-in both life history transitions. We suggest that this co-variation is not a consequence of many independent cellular and molecular switches in the same direction but rather represents the molecular equivalent of a physiological shift orchestrated by one or very few master regulators. ?? 2009 Elsevier Inc. All rights reserved.

  6. Gene expression in the etiology of schizophrenia.

    PubMed

    Bray, Nicholas J

    2008-05-01

    Gene expression represents a fundamental interface between genes and environment in the development and ongoing plasticity of the human brain. Individual differences in gene expression are likely to underpin much of human diversity, including psychiatric illness. In the past decade, the development of microarray and proteomic technology has enabled global description of gene expression in schizophrenia. However, it is difficult on the basis of gene expression assays alone to distinguish between those changes that constitute primary etiology and those that reflect secondary pathology, compensatory mechanisms, or confounding influences. In this respect, tests of genetic association with schizophrenia will be instructive because changes in gene expression that result from gene variants that are associated with the disorder are likely to be of primary etiological significance. However, regulatory polymorphism is extremely difficult to recognize on the basis of sequence interrogation alone. Functional assays at the messenger RNA and/or protein level will be essential in elucidating the molecular mechanisms underlying genetic association with schizophrenia and are likely to become increasingly important in the identification of regulatory variants with which to test for association with the disorder and related traits. Once established, etiologically relevant changes in gene expression can be recapitulated in model systems in order to elucidate the molecular and physiological pathways that may ultimately give rise to the condition.

  7. Variation-preserving normalization unveils blind spots in gene expression profiling

    PubMed Central

    Roca, Carlos P.; Gomes, Susana I. L.; Amorim, Mónica J. B.; Scott-Fordsmand, Janeck J.

    2017-01-01

    RNA-Seq and gene expression microarrays provide comprehensive profiles of gene activity, but lack of reproducibility has hindered their application. A key challenge in the data analysis is the normalization of gene expression levels, which is currently performed following the implicit assumption that most genes are not differentially expressed. Here, we present a mathematical approach to normalization that makes no assumption of this sort. We have found that variation in gene expression is much larger than currently believed, and that it can be measured with available assays. Our results also explain, at least partially, the reproducibility problems encountered in transcriptomics studies. We expect that this improvement in detection will help efforts to realize the full potential of gene expression profiling, especially in analyses of cellular processes involving complex modulations of gene expression. PMID:28276435

  8. Noise minimisation in gene expression switches.

    PubMed

    Monteoliva, Diana; McCarthy, Christina B; Diambra, Luis

    2013-01-01

    Gene expression is subject to stochastic variation which leads to fluctuations in the rate of protein production. Recently, a study in yeast at a genomic scale showed that, in some cases, gene expression variability alters phenotypes while, in other cases, these remain unchanged despite fluctuations in the expression of other genes. These studies suggested that noise in gene expression is a physiologically relevant trait and, to prevent harmful stochastic variation in the expression levels of some genes, it can be subject to minimisation. However, the mechanisms for noise minimisation are still unclear. In the present work, we analysed how noise expression depends on the architecture of the cis-regulatory system, in particular on the number of regulatory binding sites. Using analytical calculations and stochastic simulations, we found that the fluctuation level in noise expression decreased with the number of regulatory sites when regulatory transcription factors interacted with only one other bound transcription factor. In contrast, we observed that there was an optimal number of binding sites when transcription factors interacted with many bound transcription factors. This finding suggested a new mechanism for preventing large fluctuations in the expression of genes which are sensitive to the concentration of regulators.

  9. Noise Minimisation in Gene Expression Switches

    PubMed Central

    Monteoliva, Diana; McCarthy, Christina B.; Diambra, Luis

    2013-01-01

    Gene expression is subject to stochastic variation which leads to fluctuations in the rate of protein production. Recently, a study in yeast at a genomic scale showed that, in some cases, gene expression variability alters phenotypes while, in other cases, these remain unchanged despite fluctuations in the expression of other genes. These studies suggested that noise in gene expression is a physiologically relevant trait and, to prevent harmful stochastic variation in the expression levels of some genes, it can be subject to minimisation. However, the mechanisms for noise minimisation are still unclear. In the present work, we analysed how noise expression depends on the architecture of the cis-regulatory system, in particular on the number of regulatory binding sites. Using analytical calculations and stochastic simulations, we found that the fluctuation level in noise expression decreased with the number of regulatory sites when regulatory transcription factors interacted with only one other bound transcription factor. In contrast, we observed that there was an optimal number of binding sites when transcription factors interacted with many bound transcription factors. This finding suggested a new mechanism for preventing large fluctuations in the expression of genes which are sensitive to the concentration of regulators. PMID:24376783

  10. Tenomodulin Expression in the Periodontal Ligament Enhances Cellular Adhesion

    PubMed Central

    Komiyama, Yuske; Ohba, Shinsuke; Shimohata, Nobuyuki; Nakajima, Keiji; Hojo, Hironori; Yano, Fumiko; Takato, Tsuyoshi; Docheva, Denitsa; Shukunami, Chisa; Hiraki, Yuji; Chung, Ung-il

    2013-01-01

    Tenomodulin (Tnmd) is a type II transmembrane protein characteristically expressed in dense connective tissues such as tendons and ligaments. Its expression in the periodontal ligament (PDL) has also been demonstrated, though the timing and function remain unclear. We investigated the expression of Tnmd during murine tooth eruption and explored its biological functions in vitro. Tnmd expression was related to the time of eruption when occlusal force was transferred to the teeth and surrounding tissues. Tnmd overexpression enhanced cell adhesion in NIH3T3 and human PDL cells. In addition, Tnmd-knockout fibroblasts showed decreased cell adhesion. In the extracellular portions of Tnmd, the BRICHOS domain or CS region was found to be responsible for Tnmd-mediated enhancement of cell adhesion. These results suggest that Tnmd acts on the maturation or maintenance of the PDL by positively regulating cell adhesion via its BRICHOS domain. PMID:23593173

  11. The relative cellular levels of CP2a and CP2b potentiates erythroid cell-specific expression of the {alpha}-globin gene by regulating the nuclear localization of CP2c

    SciTech Connect

    Chae, Ji Hyung; Kang, Ho Chul; Kim, Chul Geun

    2009-03-20

    CP2b activates {alpha}-globin expression in an erythroid cell-specific manner, through interaction with CP2c and PIAS1. Although CP2a is identical to CP2b except for lacking an exon encoding additional 36 amino acids and has the intrinsic DNA binding and transactivation properties, it does not exert any role in {alpha}-globin expression. Investigation of subcellular localization of exogenous CP2 proteins revealed that CP2a and CP2b were exclusively localized in the cytosol and nucleus, respectively. The CP2b-specific exon was in charge of the nuclear localization of CP2b. Interestingly, subcellular localization of CP2c was either in the nucleus or cytosol depending on the relative level of CP2a and CP2b although CP2c intrinsically localized in the cytosol in the absence of CP2a/CP2b. Finally, dramatic increment of hemoglobin expression was correlated with nuclear translocation of CP2c during MEL cell differentiation. Our data suggest that CP2b potentiate erythroid cell-specific {alpha}-globin expression by recruiting CP2c into the nucleus.

  12. Dynamic Post-Transcriptional Regulation of HIV-1 Gene Expression

    PubMed Central

    Kula, Anna; Marcello, Alessandro

    2012-01-01

    Gene expression of the human immunodeficiency virus type 1 (HIV-1) is a highly regulated process. Basal transcription of the integrated provirus generates early transcripts that encode for the viral products Tat and Rev. Tat promotes the elongation of RNA polymerase while Rev mediates the nuclear export of viral RNAs that contain the Rev-responsive RNA element (RRE). These RNAs are exported from the nucleus to allow expression of Gag-Pol and Env proteins and for the production of full-length genomic RNAs. A balance exists between completely processed mRNAs and RRE-containing RNAs. Rev functions as an adaptor that recruits cellular factors to re-direct singly spliced and unspliced viral RNAs to nuclear export. The aim of this review is to address the dynamic regulation of this post-transcriptional pathway in light of recent findings that implicate several novel cellular cofactors of Rev function. PMID:24832221

  13. Nucleosome repositioning underlies dynamic gene expression.

    PubMed

    Nocetti, Nicolas; Whitehouse, Iestyn

    2016-03-15

    Nucleosome repositioning at gene promoters is a fundamental aspect of the regulation of gene expression. However, the extent to which nucleosome repositioning is used within eukaryotic genomes is poorly understood. Here we report a comprehensive analysis of nucleosome positions as budding yeast transit through an ultradian cycle in which expression of >50% of all genes is highly synchronized. We present evidence of extensive nucleosome repositioning at thousands of gene promoters as genes are activated and repressed. During activation, nucleosomes are relocated to allow sites of general transcription factor binding and transcription initiation to become accessible. The extent of nucleosome shifting is closely related to the dynamic range of gene transcription and generally related to DNA sequence properties and use of the coactivators TFIID or SAGA. However, dynamic gene expression is not limited to SAGA-regulated promoters and is an inherent feature of most genes. While nucleosome repositioning occurs pervasively, we found that a class of genes required for growth experience acute nucleosome shifting as cells enter the cell cycle. Significantly, our data identify that the ATP-dependent chromatin-remodeling enzyme Snf2 plays a fundamental role in nucleosome repositioning and the expression of growth genes. We also reveal that nucleosome organization changes extensively in concert with phases of the cell cycle, with large, regularly spaced nucleosome arrays being established in mitosis. Collectively, our data and analysis provide a framework for understanding nucleosome dynamics in relation to fundamental DNA-dependent transactions.

  14. Caenorhabditis elegans metabolic gene regulatory networks govern the cellular economy.

    PubMed

    Watson, Emma; Walhout, Albertha J M

    2014-10-01

    Diet greatly impacts metabolism in health and disease. In response to the presence or absence of specific nutrients, metabolic gene regulatory networks sense the metabolic state of the cell and regulate metabolic flux accordingly, for instance by the transcriptional control of metabolic enzymes. Here, we discuss recent insights regarding metazoan metabolic regulatory networks using the nematode Caenorhabditis elegans as a model, including the modular organization of metabolic gene regulatory networks, the prominent impact of diet on the transcriptome and metabolome, specialized roles of nuclear hormone receptors (NHRs) in responding to dietary conditions, regulation of metabolic genes and metabolic regulators by miRNAs, and feedback between metabolic genes and their regulators.

  15. Gene Expression Patterns in Human Liver Cancers

    PubMed Central

    Chen, Xin; Cheung, Siu Tim; So, Samuel; Fan, Sheung Tat; Barry, Christopher; Higgins, John; Lai, Kin-Man; Ji, Jiafu; Dudoit, Sandrine; Ng, Irene O.L.; van de Rijn, Matt; Botstein, David; Brown, Patrick O.

    2002-01-01

    Hepatocellular carcinoma (HCC) is a leading cause of death worldwide. Using cDNA microarrays to characterize patterns of gene expression in HCC, we found consistent differences between the expression patterns in HCC compared with those seen in nontumor liver tissues. The expression patterns in HCC were also readily distinguished from those associated with tumors metastatic to liver. The global gene expression patterns intrinsic to each tumor were sufficiently distinctive that multiple tumor nodules from the same patient could usually be recognized and distinguished from all the others in the large sample set on the basis of their gene expression patterns alone. The distinctive gene expression patterns are characteristic of the tumors and not the patient; the expression programs seen in clonally independent tumor nodules in the same patient were no more similar than those in tumors from different patients. Moreover, clonally related tumor masses that showed distinct expression profiles were also distinguished by genotypic differences. Some features of the gene expression patterns were associated with specific phenotypic and genotypic characteristics of the tumors, including growth rate, vascular invasion, and p53 overexpression. PMID:12058060

  16. Gene Expression Signature-Based Screening Identifies New Broadly Effective Influenza A Antivirals

    PubMed Central

    Josset, Laurence; Textoris, Julien; Loriod, Béatrice; Ferraris, Olivier; Moules, Vincent; Lina, Bruno; N'Guyen, Catherine; Diaz, Jean-Jacques; Rosa-Calatrava, Manuel

    2010-01-01

    Classical antiviral therapies target viral proteins and are consequently subject to resistance. To counteract this limitation, alternative strategies have been developed that target cellular factors. We hypothesized that such an approach could also be useful to identify broad-spectrum antivirals. The influenza A virus was used as a model for its viral diversity and because of the need to develop therapies against unpredictable viruses as recently underlined by the H1N1 pandemic. We proposed to identify a gene-expression signature associated with infection by different influenza A virus subtypes which would allow the identification of potential antiviral drugs with a broad anti-influenza spectrum of activity. We analyzed the cellular gene expression response to infection with five different human and avian influenza A virus strains and identified 300 genes as differentially expressed between infected and non-infected samples. The most 20 dysregulated genes were used to screen the connectivity map, a database of drug-associated gene expression profiles. Candidate antivirals were then identified by their inverse correlation to the query signature. We hypothesized that such molecules would induce an unfavorable cellular environment for influenza virus replication. Eight potential antivirals including ribavirin were identified and their effects were tested in vitro on five influenza A strains. Six of the molecules inhibited influenza viral growth. The new pandemic H1N1 virus, which was not used to define the gene expression signature of infection, was inhibited by five out of the eight identified molecules, demonstrating that this strategy could contribute to identifying new broad anti-influenza agents acting on cellular gene expression. The identified infection signature genes, the expression of which are modified upon infection, could encode cellular proteins involved in the viral life cycle. This is the first study showing that gene expression-based screening can be

  17. Ezrin Inhibition Up-regulates Stress Response Gene Expression.

    PubMed

    Çelik, Haydar; Bulut, Gülay; Han, Jenny; Graham, Garrett T; Minas, Tsion Z; Conn, Erin J; Hong, Sung-Hyeok; Pauly, Gary T; Hayran, Mutlu; Li, Xin; Özdemirli, Metin; Ayhan, Ayşe; Rudek, Michelle A; Toretsky, Jeffrey A; Üren, Aykut

    2016-06-17

    Ezrin is a member of the ERM (ezrin/radixin/moesin) family of proteins that links cortical cytoskeleton to the plasma membrane. High expression of ezrin correlates with poor prognosis and metastasis in osteosarcoma. In this study, to uncover specific cellular responses evoked by ezrin inhibition that can be used as a specific pharmacodynamic marker(s), we profiled global gene expression in osteosarcoma cells after treatment with small molecule ezrin inhibitors, NSC305787 and NSC668394. We identified and validated several up-regulated integrated stress response genes including PTGS2, ATF3, DDIT3, DDIT4, TRIB3, and ATF4 as novel ezrin-regulated transcripts. Analysis of transcriptional response in skin and peripheral blood mononuclear cells from NSC305787-treated mice compared with a control group revealed that, among those genes, the stress gene DDIT4/REDD1 may be used as a surrogate pharmacodynamic marker of ezrin inhibitor compound activity. In addition, we validated the anti-metastatic effects of NSC305787 in reducing the incidence of lung metastasis in a genetically engineered mouse model of osteosarcoma and evaluated the pharmacokinetics of NSC305787 and NSC668394 in mice. In conclusion, our findings suggest that cytoplasmic ezrin, previously considered a dormant and inactive protein, has important functions in regulating gene expression that may result in down-regulation of stress response genes.

  18. Digital gene expression signatures for maize development.

    PubMed

    Eveland, Andrea L; Satoh-Nagasawa, Namiko; Goldshmidt, Alexander; Meyer, Sandra; Beatty, Mary; Sakai, Hajime; Ware, Doreen; Jackson, David

    2010-11-01

    Genome-wide expression signatures detect specific perturbations in developmental programs and contribute to functional resolution of key regulatory networks. In maize (Zea mays) inflorescences, mutations in the RAMOSA (RA) genes affect the determinacy of axillary meristems and thus alter branching patterns, an important agronomic trait. In this work, we developed and tested a framework for analysis of tag-based, digital gene expression profiles using Illumina's high-throughput sequencing technology and the newly assembled B73 maize reference genome. We also used a mutation in the RA3 gene to identify putative expression signatures specific to stem cell fate in axillary meristem determinacy. The RA3 gene encodes a trehalose-6-phosphate phosphatase and may act at the interface between developmental and metabolic processes. Deep sequencing of digital gene expression libraries, representing three biological replicate ear samples from wild-type and ra3 plants, generated 27 million 20- to 21-nucleotide reads with frequencies spanning 4 orders of magnitude. Unique sequence tags were anchored to 3'-ends of individual transcripts by DpnII and NlaIII digests, which were multiplexed during sequencing. We mapped 86% of nonredundant signature tags to the maize genome, which associated with 37,117 gene models and unannotated regions of expression. In total, 66% of genes were detected by at least nine reads in immature maize ears. We used comparative genomics to leverage existing information from Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa) in functional analyses of differentially expressed maize genes. Results from this study provide a basis for the analysis of short-read expression data in maize and resolved specific expression signatures that will help define mechanisms of action for the RA3 gene.

  19. A gene expression atlas of the domestic pig

    PubMed Central

    2012-01-01

    Background This work describes the first genome-wide analysis of the transcriptional landscape of the pig. A new porcine Affymetrix expression array was designed in order to provide comprehensive coverage of the known pig transcriptome. The new array was used to generate a genome-wide expression atlas of pig tissues derived from 62 tissue/cell types. These data were subjected to network correlation analysis and clustering. Results The analysis presented here provides a detailed functional clustering of the pig transcriptome where transcripts are grouped according to their expression pattern, so one can infer the function of an uncharacterized gene from the company it keeps and the locations in which it is expressed. We describe the overall transcriptional signatures present in the tissue atlas, where possible assigning those signatures to specific cell populations or pathways. In particular, we discuss the expression signatures associated with the gastrointestinal tract, an organ that was sampled at 15 sites along its length and whose biology in the pig is similar to human. We identify sets of genes that define specialized cellular compartments and region-specific digestive functions. Finally, we performed a network analysis of the transcription factors expressed in the gastrointestinal tract and demonstrate how they sub-divide into functional groups that may control cellular gastrointestinal development. Conclusions As an important livestock animal with a physiology that is more similar than mouse to man, we provide a major new resource for understanding gene expression with respect to the known physiology of mammalian tissues and cells. The data and analyses are available on the websites http://biogps.org and http://www.macrophages.com/pig-atlas. PMID:23153189

  20. Gene expression homeostasis and chromosome architecture

    PubMed Central

    Seshasayee, Aswin Sai Narain

    2014-01-01

    In rapidly growing populations of bacterial cells, including those of the model organism Escherichia coli, genes essential for growth - such as those involved in protein synthesis - are expressed at high levels; this is in contrast to many horizontally-acquired genes, which are maintained at low transcriptional levels.1 This balance in gene expression states between 2 distinct classes of genes is established by a galaxy of transcriptional regulators, including the so-called nucleoid associated proteins (NAP) that contribute to shaping the chromosome.2 Besides these active players in gene regulation, it is not too far-fetched to anticipate that genome organization in terms of how genes are arranged on the chromosome,3 which is the result of long-drawn transactions among genome rearrangement processes and selection, and the manner in which it is structured inside the cell, plays a role in establishing this balance. A recent study from our group has contributed to the literature investigating the interplay between global transcriptional regulators and genome organization in establishing gene expression homeostasis.4 In particular, we address a triangle of functional interactions among genome organization, gene expression homeostasis and horizontal gene transfer. PMID:25997086

  1. Unmasking ultradian rhythms in gene expression

    PubMed Central

    van der Veen, Daan R.; Gerkema, Menno P.

    2017-01-01

    Biological oscillations with an ultradian time scale of 1 to several hours include cycles in behavioral arousal, episodic glucocorticoid release, and gene expression. Ultradian rhythms are thought to have an extrinsic origin because of a perceived absence of ultradian rhythmicity in vitro and a lack of known molecular ultradian oscillators. We designed a novel, non–spectral-analysis method of separating ultradian from circadian components and applied it to a published gene expression dataset with an ultradian sampling resolution. Ultradian rhythms in mouse hepatocytes in vivo have been published, and we validated our approach using this control by confirming 175 of 323 ultradian genes identified in a prior study and found 862 additional ultradian genes. For the first time, we now report ultradian expression of >900 genes in vitro. Sixty genes exhibited ultradian transcriptional rhythmicity, both in vivo and in vitro, including 5 genes involved in the cell cycle. Within these 60 genes, we identified significant enrichment of specific DNA motifs in the 1000 bp proximal promotor, some of which associate with known transcriptional factors. These findings are in strong support of instrinsically driven ultradian rhythms and expose potential molecular mechanisms and functions underlying ultradian rhythms that remain unknown.—Van der Veen, D. R., Gerkema, M. P. Unmasking ultradian rhythms in gene expression. PMID:27871062

  2. Expression of polarity genes in human cancer.

    PubMed

    Lin, Wan-Hsin; Asmann, Yan W; Anastasiadis, Panos Z

    2015-01-01

    Polarity protein complexes are crucial for epithelial apical-basal polarity and directed cell migration. Since alterations of these processes are common in cancer, polarity proteins have been proposed to function as tumor suppressors or oncogenic promoters. Here, we review the current understanding of polarity protein functions in epithelial homeostasis, as well as tumor formation and progression. As most previous studies focused on the function of single polarity proteins in simplified model systems, we used a genomics approach to systematically examine and identify the expression profiles of polarity genes in human cancer. The expression profiles of polarity genes were distinct in different human tissues and classified cancer types. Additionally, polarity expression profiles correlated with disease progression and aggressiveness, as well as with identified cancer types, where specific polarity genes were commonly altered. In the case of Scribble, gene expression analysis indicated its common amplification and upregulation in human cancer, suggesting a tumor promoting function.

  3. Regulation of Gene Expression in Protozoa Parasites

    PubMed Central

    Gomez, Consuelo; Esther Ramirez, M.; Calixto-Galvez, Mercedes; Medel, Olivia; Rodríguez, Mario A.

    2010-01-01

    Infections with protozoa parasites are associated with high burdens of morbidity and mortality across the developing world. Despite extensive efforts to control the transmission of these parasites, the spread of populations resistant to drugs and the lack of effective vaccines against them contribute to their persistence as major public health problems. Parasites should perform a strict control on the expression of genes involved in their pathogenicity, differentiation, immune evasion, or drug resistance, and the comprehension of the mechanisms implicated in that control could help to develop novel therapeutic strategies. However, until now these mechanisms are poorly understood in protozoa. Recent investigations into gene expression in protozoa parasites suggest that they possess many of the canonical machineries employed by higher eukaryotes for the control of gene expression at transcriptional, posttranscriptional, and epigenetic levels, but they also contain exclusive mechanisms. Here, we review the current understanding about the regulation of gene expression in Plasmodium sp., Trypanosomatids, Entamoeba histolytica and Trichomonas vaginalis. PMID:20204171

  4. Dynamic modeling of gene expression data

    NASA Technical Reports Server (NTRS)

    Holter, N. S.; Maritan, A.; Cieplak, M.; Fedoroff, N. V.; Banavar, J. R.

    2001-01-01

    We describe the time evolution of gene expression levels by using a time translational matrix to predict future expression levels of genes based on their expression levels at some initial time. We deduce the time translational matrix for previously published DNA microarray gene expression data sets by modeling them within a linear framework by using the characteristic modes obtained by singular value decomposition. The resulting time translation matrix provides a measure of the relationships among the modes and governs their time evolution. We show that a truncated matrix linking just a few modes is a good approximation of the full time translation matrix. This finding suggests that the number of essential connections among the genes is small.

  5. Distal chromatin structure influences local nucleosome positions and gene expression.

    PubMed

    Jansen, An; van der Zande, Elisa; Meert, Wim; Fink, Gerald R; Verstrepen, Kevin J

    2012-05-01

    The positions of nucleosomes across the genome influence several cellular processes, including gene transcription. However, our understanding of the factors dictating where nucleosomes are located and how this affects gene regulation is still limited. Here, we perform an extensive in vivo study to investigate the influence of the neighboring chromatin structure on local nucleosome positioning and gene expression. Using truncated versions of the Saccharomyces cerevisiae URA3 gene, we show that nucleosome positions in the URA3 promoter are at least partly determined by the local DNA sequence, with so-called 'anti-nucleosomal elements' like poly(dA:dT) tracts being key determinants of nucleosome positions. In addition, we show that changes in the nucleosome positions in the URA3 promoter strongly affect the promoter activity. Most interestingly, in addition to demonstrating the effect of the local DNA sequence, our study provides novel in vivo evidence that nucleosome positions are also affected by the position of neighboring nucleosomes. Nucleosome structure may therefore be an important selective force for conservation of gene order on a chromosome, because relocating a gene to another genomic position (where the positions of neighboring nucleosomes are different from the original locus) can have dramatic consequences for the gene's nucleosome structure and thus its expression.

  6. Gene expression analysis of precision-cut human liver slices indicates stable expression of ADME-Tox related genes

    SciTech Connect

    Elferink, M.G.L.; Olinga, P.; van Leeuwen, E.M.; Bauerschmidt, S.; Polman, J.; Schoonen, W.G.; Heisterkamp, S.H.; Groothuis, G.M.M.

    2011-05-15

    In the process of drug development it is of high importance to test the safety of new drugs with predictive value for human toxicity. A promising approach of toxicity testing is based on shifts in gene expression profiling of the liver. Toxicity screening based on animal liver cells cannot be directly extrapolated to humans due to species differences. The aim of this study was to evaluate precision-cut human liver slices as in vitro method for the prediction of human specific toxicity by toxicogenomics. The liver slices contain all cell types of the liver in their natural architecture. This is important since drug-induced toxicity often is a multi-cellular process. Previously we showed that toxicogenomic analysis of rat liver slices is highly predictive for rat in vivo toxicity. In this study we investigated the levels of gene expression during incubation up to 24 h with Affymetrix microarray technology. The analysis was focused on a broad spectrum of genes related to stress and toxicity, and on genes encoding for phase-I, -II and -III metabolizing enzymes and transporters. Observed changes in gene expression were associated with cytoskeleton remodeling, extracellular matrix and cell adhesion, but for the ADME-Tox related genes only minor changes were observed. PCA analysis showed that changes in gene expression were not associated with age, sex or source of the human livers. Slices treated with acetaminophen showed patterns of gene expression related to its toxicity. These results indicate that precision-cut human liver slices are relatively stable during 24 h of incubation and represent a valuable model for human in vitro hepatotoxicity testing despite the human inter-individual variability.

  7. Mining Gene Expression Data of Multiple Sclerosis

    PubMed Central

    Zhu, Zhenli; Huang, Zhengliang; Li, Ke

    2014-01-01

    Objectives Microarray produces a large amount of gene expression data, containing various biological implications. The challenge is to detect a panel of discriminative genes associated with disease. This study proposed a robust classification model for gene selection using gene expression data, and performed an analysis to identify disease-related genes using multiple sclerosis as an example. Materials and methods Gene expression profiles based on the transcriptome of peripheral blood mononuclear cells from a total of 44 samples from 26 multiple sclerosis patients and 18 individuals with other neurological diseases (control) were analyzed. Feature selection algorithms including Support Vector Machine based on Recursive Feature Elimination, Receiver Operating Characteristic Curve, and Boruta algorithms were jointly performed to select candidate genes associating with multiple sclerosis. Multiple classification models categorized samples into two different groups based on the identified genes. Models’ performance was evaluated using cross-validation methods, and an optimal classifier for gene selection was determined. Results An overlapping feature set was identified consisting of 8 genes that were differentially expressed between the two phenotype groups. The genes were significantly associated with the pathways of apoptosis and cytokine-cytokine receptor interaction. TNFSF10 was significantly associated with multiple sclerosis. A Support Vector Machine model was established based on the featured genes and gave a practical accuracy of ∼86%. This binary classification model also outperformed the other models in terms of Sensitivity, Specificity and F1 score. Conclusions The combined analytical framework integrating feature ranking algorithms and Support Vector Machine model could be used for selecting genes for other diseases. PMID:24932510

  8. Noise-based switches and amplifiers for gene expression

    PubMed Central

    Hasty, Jeff; Pradines, Joel; Dolnik, Milos; Collins, J. J.

    2000-01-01

    The regulation of cellular function is often controlled at the level of gene transcription. Such genetic regulation usually consists of interacting networks, whereby gene products from a single network can act to control their own expression or the production of protein in another network. Engineered control of cellular function through the design and manipulation of such networks lies within the constraints of current technology. Here we develop a model describing the regulation of gene expression and elucidate the effects of noise on the formulation. We consider a single network derived from bacteriophage λ and construct a two-parameter deterministic model describing the temporal evolution of the concentration of λ repressor protein. Bistability in the steady-state protein concentration arises naturally, and we show how the bistable regime is enhanced with the addition of the first operator site in the promotor region. We then show how additive and multiplicative external noise can be used to regulate expression. In the additive case, we demonstrate the utility of such control through the construction of a protein switch, whereby protein production is turned “on” and “off” by using short noise pulses. In the multiplicative case, we show that small deviations in the transcription rate can lead to large fluctuations in the production of protein, and we describe how these fluctuations can be used to amplify protein production significantly. These results suggest that an external noise source could be used as a switch and/or amplifier for gene expression. Such a development could have important implications for gene therapy. PMID:10681449

  9. Constructing gene co-expression networks and predicting functions of unknown genes by random matrix theory

    PubMed Central

    Luo, Feng; Yang, Yunfeng; Zhong, Jianxin; Gao, Haichun; Khan, Latifur; Thompson, Dorothea K; Zhou, Jizhong

    2007-01-01

    Background Large-scale sequencing of entire genomes has ushered in a new age in biology. One of the next grand challenges is to dissect the cellular networks consisting of many individual functional modules. Defining co-expression networks without ambiguity based on genome-wide microarray data is difficult and current methods are not robust and consistent with different data sets. This is particularly problematic for little understood organisms since not much existing biological knowledge can be exploited for determining the threshold to differentiate true correlation from random noise. Random matrix theory (RMT), which has been widely and successfully used in physics, is a powerful approach to distinguish system-specific, non-random properties embedded in complex systems from random noise. Here, we have hypothesized that the universal predictions of RMT are also applicable to biological systems and the correlation threshold can be determined by characterizing the correlation matrix of microarray profiles using random matrix theory. Results Application of random matrix theory to microarray data of S. oneidensis, E. coli, yeast, A. thaliana, Drosophila, mouse and human indicates that there is a sharp transition of nearest neighbour spacing distribution (NNSD) of correlation matrix after gradually removing certain elements insider the matrix. Testing on an in silico modular model has demonstrated that this transition can be used to determine the correlation threshold for revealing modular co-expression networks. The co-expression network derived from yeast cell cycling microarray data is supported by gene annotation. The topological properties of the resulting co-expression network agree well with the general properties of biological networks. Computational evaluations have showed that RMT approach is sensitive and robust. Furthermore, evaluation on sampled expression data of an in silico modular gene system has showed that under-sampled expressions do not affect the

  10. Amino acid regulation of gene expression.

    PubMed Central

    Fafournoux, P; Bruhat, A; Jousse, C

    2000-01-01

    The impact of nutrients on gene expression in mammals has become an important area of research. Nevertheless, the current understanding of the amino acid-dependent control of gene expression is limited. Because amino acids have multiple and important functions, their homoeostasis has to be finely maintained. However, amino-acidaemia can be affected by certain nutritional conditions or various forms of stress. It follows that mammals have to adjust several of their physiological functions involved in the adaptation to amino acid availability by regulating the expression of numerous genes. The aim of the present review is to examine the role of amino acids in regulating mammalian gene expression and protein turnover. It has been reported that some genes involved in the control of growth or amino acid metabolism are regulated by amino acid availability. For instance, limitation of several amino acids greatly increases the expression of the genes encoding insulin-like growth factor binding protein-1, CHOP (C/EBP homologous protein, where C/EBP is CCAAT/enhancer binding protein) and asparagine synthetase. Elevated mRNA levels result from both an increase in the rate of transcription and an increase in mRNA stability. Several observations suggest that the amino acid regulation of gene expression observed in mammalian cells and the general control process described in yeast share common features. Moreover, amino acid response elements have been characterized in the promoters of the CHOP and asparagine synthetase genes. Taken together, the results discussed in the present review demonstrate that amino acids, by themselves, can, in concert with hormones, play an important role in the control of gene expression. PMID:10998343

  11. Prion search and cellular prion protein expression in stranded dolphins.

    PubMed

    Di Guardo, G; Cocumelli, C; Meoli, R; Barbaro, K; Terracciano, G; Di Francesco, C E; Mazzariol, S; Eleni, C

    2012-01-01

    The recent description of a prion disease (PD) case in a free-ranging bottlenose dolphin (Tursiops truncatus) prompted us to carry out an extensive search for the disease-associated isoform (PrPSc) of the cellular prion protein (PrPC) in the brain and in a range of lymphoid tissues from 23 striped dolphins (Stenella coeruleoalba), 5 bottlenose dolphins and 2 Risso s dolphins (Grampus griseus) found stranded between 2007 and 2012 along the Italian coastline. Three striped dolphins and one bottlenose dolphin showed microscopic lesions of encephalitis, with no evidence of spongiform brain lesions being detected in any of the 30 free-ranging cetaceans investigated herein. Nevertheless, we could still observe a prominent PrPC immunoreactivity in the brain as well as in lymphoid tissues from these dolphins. Although immunohistochemical and Western blot investigations yielded negative results for PrPSc deposition in all tissues from the dolphins under study, the reported occurrence of a spontaneous PD case in a wild dolphin is an intriguing issue and a matter of concern for both prion biology and intra/inter-species transmissibility, as well as for cetacean conservation medicine.

  12. Imputing gene expression to maximize platform compatibility.

    PubMed

    Zhou, Weizhuang; Han, Lichy; Altman, Russ B

    2017-02-15

    Microarray measurements of gene expression constitute a large fraction of publicly shared biological data, and are available in the Gene Expression Omnibus (GEO). Many studies use GEO data to shape hypotheses and improve statistical power. Within GEO, the Affymetrix HG-U133A and HG-U133 Plus 2.0 are the two most commonly used microarray platforms for human samples; the HG-U133 Plus 2.0 platform contains 54 220 probes and the HG-U133A array contains a proper subset (21 722 probes). When different platforms are involved, the subset of common genes is most easily compared. This approach results in the exclusion of substantial measured data and can limit downstream analysis. To predict the expression values for the genes unique to the HG-U133 Plus 2.0 platform, we constructed a series of gene expression inference models based on genes common to both platforms. Our model predicts gene expression values that are within the variability observed in controlled replicate studies and are highly correlated with measured data. Using six previously published studies, we also demonstrate the improved performance of the enlarged feature space generated by our model in downstream analysis.

  13. Optimal Reference Genes for Gene Expression Normalization in Trichomonas vaginalis.

    PubMed

    dos Santos, Odelta; de Vargas Rigo, Graziela; Frasson, Amanda Piccoli; Macedo, Alexandre José; Tasca, Tiana

    2015-01-01

    Trichomonas vaginalis is the etiologic agent of trichomonosis, the most common non-viral sexually transmitted disease worldwide. This infection is associated with several health consequences, including cervical and prostate cancers and HIV acquisition. Gene expression analysis has been facilitated because of available genome sequences and large-scale transcriptomes in T. vaginalis, particularly using quantitative real-time polymerase chain reaction (qRT-PCR), one of the most used methods for molecular studies. Reference genes for normalization are crucial to ensure the accuracy of this method. However, to the best of our knowledge, a systematic validation of reference genes has not been performed for T. vaginalis. In this study, the transcripts of nine candidate reference genes were quantified using qRT-PCR under different cultivation conditions, and the stability of these genes was compared using the geNorm and NormFinder algorithms. The most stable reference genes were α-tubulin, actin and DNATopII, and, conversely, the widely used T. vaginalis reference genes GAPDH and β-tubulin were less stable. The PFOR gene was used to validate the reliability of the use of these candidate reference genes. As expected, the PFOR gene was upregulated when the trophozoites were cultivated with ferrous ammonium sulfate when the DNATopII, α-tubulin and actin genes were used as normalizing gene. By contrast, the PFOR gene was downregulated when the GAPDH gene was used as an internal control, leading to misinterpretation of the data. These results provide an important starting point for reference gene selection and gene expression analysis with qRT-PCR studies of T. vaginalis.

  14. Glycosyltransferase Gene Expression Profiles Classify Cancer Types and Propose Prognostic Subtypes

    NASA Astrophysics Data System (ADS)

    Ashkani, Jahanshah; Naidoo, Kevin J.

    2016-05-01

    Aberrant glycosylation in tumours stem from altered glycosyltransferase (GT) gene expression but can the expression profiles of these signature genes be used to classify cancer types and lead to cancer subtype discovery? The differential structural changes to cellular glycan structures are predominantly regulated by the expression patterns of GT genes and are a hallmark of neoplastic cell metamorphoses. We found that the expression of 210 GT genes taken from 1893 cancer patient samples in The Cancer Genome Atlas (TCGA) microarray data are able to classify six cancers; breast, ovarian, glioblastoma, kidney, colon and lung. The GT gene expression profiles are used to develop cancer classifiers and propose subtypes. The subclassification of breast cancer solid tumour samples illustrates the discovery of subgroups from GT genes that match well against basal-like and HER2-enriched subtypes and correlates to clinical, mutation and survival data. This cancer type glycosyltransferase gene signature finding provides foundational evidence for the centrality of glycosylation in cancer.

  15. A primer on molecular biology for imagers: II. Transcription and gene expression.

    PubMed

    Pandit, Sunil D; Li, King C P

    2004-03-01

    The process of gene expression is complex and highly regulated to ensure that the right gene is expressed at the right place, at the right time, and in regulated amounts. The cell has multiple levels at which it controls the expression of a transcript including gene expression, alternate splicing, and stability of the transcript. Alternate splicing to generate different RNA species from a given gene and DNA rearrangements where genes are rearranged during cellular differentiation (eg, immunoglobulin genes) are additional mechanisms used to generate diversity in complex organisms. Epigenetic mechanisms such as methylation where CpG-rich islands in the promoter region depending on their methylation status can also modulate gene expression. The reader is requested to refer to the books, review articles, and web sites for additional information.

  16. Glycosyltransferase Gene Expression Profiles Classify Cancer Types and Propose Prognostic Subtypes

    PubMed Central

    Ashkani, Jahanshah; Naidoo, Kevin J.

    2016-01-01

    Aberrant glycosylation in tumours stem from altered glycosyltransferase (GT) gene expression but can the expression profiles of these signature genes be used to classify cancer types and lead to cancer subtype discovery? The differential structural changes to cellular glycan structures are predominantly regulated by the expression patterns of GT genes and are a hallmark of neoplastic cell metamorphoses. We found that the expression of 210 GT genes taken from 1893 cancer patient samples in The Cancer Genome Atlas (TCGA) microarray data are able to classify six cancers; breast, ovarian, glioblastoma, kidney, colon and lung. The GT gene expression profiles are used to develop cancer classifiers and propose subtypes. The subclassification of breast cancer solid tumour samples illustrates the discovery of subgroups from GT genes that match well against basal-like and HER2-enriched subtypes and correlates to clinical, mutation and survival data. This cancer type glycosyltransferase gene signature finding provides foundational evidence for the centrality of glycosylation in cancer. PMID:27198045

  17. Perspectives: Gene Expression in Fisheries Management

    USGS Publications Warehouse

    Nielsen, Jennifer L.; Pavey, Scott A.

    2010-01-01

    Functional genes and gene expression have been connected to physiological traits linked to effective production and broodstock selection in aquaculture, selective implications of commercial fish harvest, and adaptive changes reflected in non-commercial fish populations subject to human disturbance and climate change. Gene mapping using single nucleotide polymorphisms (SNPs) to identify functional genes, gene expression (analogue microarrays and real-time PCR), and digital sequencing technologies looking at RNA transcripts present new concepts and opportunities in support of effective and sustainable fisheries. Genomic tools have been rapidly growing in aquaculture research addressing aspects of fish health, toxicology, and early development. Genomic technologies linking effects in functional genes involved in growth, maturation and life history development have been tied to selection resulting from harvest practices. Incorporating new and ever-increasing knowledge of fish genomes is opening a different perspective on local adaptation that will prove invaluable in wild fish conservation and management. Conservation of fish stocks is rapidly incorporating research on critical adaptive responses directed at the effects of human disturbance and climate change through gene expression studies. Genomic studies of fish populations can be generally grouped into three broad categories: 1) evolutionary genomics and biodiversity; 2) adaptive physiological responses to a changing environment; and 3) adaptive behavioral genomics and life history diversity. We review current genomic research in fisheries focusing on those that use microarrays to explore differences in gene expression among phenotypes and within or across populations, information that is critically important to the conservation of fish and their relationship to humans.

  18. Applications of queueing theory to stochastic models of gene expression

    NASA Astrophysics Data System (ADS)

    Kulkarni, Rahul

    2012-02-01

    The intrinsic stochasticity of cellular processes implies that analysis of fluctuations (`noise') is often essential for quantitative modeling of gene expression. Recent single-cell experiments have carried out such analysis to characterize moments and entire probability distributions for quantities of interest, e.g. mRNA and protein levels across a population of cells. Correspondingly, there is a need to develop general analytical tools for modeling and interpretation of data obtained from such single-cell experiments. One such approach involves the mapping between models of stochastic gene expression and systems analyzed in queueing theory. The talk will provide an overview of this approach and discuss how theorems from queueing theory (e.g. Little's Law) can be used to derive exact results for general stochastic models of gene expression. In the limit that gene expression occurs in bursts, analytical results can be obtained which provide insight into the effects of different regulatory mechanisms on the noise in protein steady-state distributions. In particular, the approach can be used to analyze the effect of post-transcriptional regulation by non-coding RNAs leading to new insights and experimentally testable predictions.

  19. Control of gene expression in trypanosomes.

    PubMed Central

    Vanhamme, L; Pays, E

    1995-01-01

    Trypanosomes are protozoan agents of major parasitic diseases such as Chagas' disease in South America and sleeping sickness of humans and nagana disease of cattle in Africa. They are transmitted to mammalian hosts by specific insect vectors. Their life cycle consists of a succession of differentiation and growth phases requiring regulated gene expression to adapt to the changing extracellular environment. Typical of such stage-specific expression is that of the major surface antigens of Trypanosoma brucei, procyclin in the procyclic (insect) form and the variant surface glycoprotein (VSG) in the bloodstream (mammalian) form. In trypanosomes, the regulation of gene expression is effected mainly at posttranscriptional levels, since primary transcription of most of the genes occurs in long polycistronic units and is constitutive. The transcripts are processed by transsplicing and polyadenylation under the influence of intergenic polypyrimidine tracts. These events show some developmental regulation. Untranslated sequences of the mRNAs seem to play a prominent role in the stage-specific control of individual gene expression, through a modulation of mRNA abundance. The VSG and procyclin transcription units exhibit particular features that are probably related to the need for a high level of expression. The promoters and RNA polymerase driving the expression of these units resemble those of the ribosomal genes. Their mutually exclusive expression is ensured by controls operating at several levels, including RNA elongation. Antigenic variation in the bloodstream is achieved through DNA rearrangements or alternative activation of the telomeric VSG gene expression sites. Recent discoveries, such as the existence of a novel nucleotide in telomeric DNA and the generation of point mutations in VSG genes, have shed new light on the mechanisms and consequences of antigenic variation. PMID:7603410

  20. Resource Sharing Controls Gene Expression Bursting.

    PubMed

    Caveney, Patrick M; Norred, S Elizabeth; Chin, Charles W; Boreyko, Jonathan B; Razooky, Brandon S; Retterer, Scott T; Collier, C Patrick; Simpson, Michael L

    2017-02-17

    Episodic gene expression, with periods of high expression separated by periods of no expression, is a pervasive biological phenomenon. This bursty pattern of expression draws from a finite reservoir of expression machinery in a highly time variant way, i.e., requiring no resources most of the time but drawing heavily on them during short intense bursts, that intimately links expression bursting and resource sharing. Yet, most recent investigations have focused on specific molecular mechanisms intrinsic to the bursty behavior of individual genes, while little is known about the interplay between resource sharing and global expression bursting behavior. Here, we confine Escherichia coli cell extract in both cell-sized microfluidic chambers and lipid-based vesicles to explore how resource sharing influences expression bursting. Interestingly, expression burst size, but not burst frequency, is highly sensitive to the size of the shared transcription and translation resource pools. The intriguing implication of these results is that expression bursts are more readily amplified than initiated, suggesting that burst formation occurs through positive feedback or cooperativity. When extrapolated to prokaryotic cells, these results suggest that large translational bursts may be correlated with large transcriptional bursts. This correlation is supported by recently reported transcription and translation bursting studies in E. coli. The results reported here demonstrate a strong intimate link between global expression burst patterns and resource sharing, and they suggest that bursting plays an important role in optimizing the use of limited, shared expression resources.

  1. Application of multidisciplinary analysis to gene expression.

    SciTech Connect

    Wang, Xuefel; Kang, Huining; Fields, Chris; Cowie, Jim R.; Davidson, George S.; Haaland, David Michael; Sibirtsev, Valeriy; Mosquera-Caro, Monica P.; Xu, Yuexian; Martin, Shawn Bryan; Helman, Paul; Andries, Erik; Ar, Kerem; Potter, Jeffrey; Willman, Cheryl L.; Murphy, Maurice H.

    2004-01-01

    Molecular analysis of cancer, at the genomic level, could lead to individualized patient diagnostics and treatments. The developments to follow will signal a significant paradigm shift in the clinical management of human cancer. Despite our initial hopes, however, it seems that simple analysis of microarray data cannot elucidate clinically significant gene functions and mechanisms. Extracting biological information from microarray data requires a complicated path involving multidisciplinary teams of biomedical researchers, computer scientists, mathematicians, statisticians, and computational linguists. The integration of the diverse outputs of each team is the limiting factor in the progress to discover candidate genes and pathways associated with the molecular biology of cancer. Specifically, one must deal with sets of significant genes identified by each method and extract whatever useful information may be found by comparing these different gene lists. Here we present our experience with such comparisons, and share methods developed in the analysis of an infant leukemia cohort studied on Affymetrix HG-U95A arrays. In particular, spatial gene clustering, hyper-dimensional projections, and computational linguistics were used to compare different gene lists. In spatial gene clustering, different gene lists are grouped together and visualized on a three-dimensional expression map, where genes with similar expressions are co-located. In another approach, projections from gene expression space onto a sphere clarify how groups of genes can jointly have more predictive power than groups of individually selected genes. Finally, online literature is automatically rearranged to present information about genes common to multiple groups, or to contrast the differences between the lists. The combination of these methods has improved our understanding of infant leukemia. While the complicated reality of the biology dashed our initial, optimistic hopes for simple answers from

  2. Epigenetic balance of gene expression by Polycomb and COMPASS families.

    PubMed

    Piunti, Andrea; Shilatifard, Ali

    2016-06-03

    Epigenetic regulation of gene expression in metazoans is central for establishing cellular diversity, and its deregulation can result in pathological conditions. Although transcription factors are essential for implementing gene expression programs, they do not function in isolation and require the recruitment of various chromatin-modifying and -remodeling machineries. A classic example of developmental chromatin regulation is the balanced activities of the Polycomb group (PcG) proteins within the PRC1 and PRC2 complexes, and the Trithorax group (TrxG) proteins within the COMPASS family, which are highly mutated in a large number of human diseases. In this review, we will discuss the latest findings regarding the properties of the PcG and COMPASS families and the insight they provide into the epigenetic control of transcription under physiological and pathological settings.

  3. Hippocampal gene expression changes underlying stress sensitization and recovery.

    PubMed

    Gray, J D; Rubin, T G; Hunter, R G; McEwen, B S

    2014-11-01

    behavioral response to a novel stressor. These findings establish a baseline profile of normal recovery and adaptation to stress. Importantly, they will serve as a conceptual basis to facilitate the future study of the cellular and regional basis of gene expression changes that lead to impaired recovery from stress, such as those that occur in mood and anxiety disorders.

  4. Hippocampal gene expression changes underlying stress sensitization and recovery

    PubMed Central

    Gray, Jason D.; Rubin, Todd G.; Hunter, Richard G.; McEwen, Bruce S.

    2013-01-01

    recovery and adaptation to stress. Importantly, they will serve as a conceptual basis to facilitate the future study of the cellular and regional basis of gene expression changes as well as genetic risk factors and adverse early life experiences that lead to impaired recovery from stress such as occurs in mood and anxiety disorders. PMID:24342991

  5. A structured population modeling framework for quantifying and predicting gene expression noise in flow cytometry data.

    PubMed

    Flores, Kevin B

    2013-07-01

    We formulated a structured population model with distributed parameters to identify mechanisms that contribute to gene expression noise in time-dependent flow cytometry data. The model was validated using cell population-level gene expression data from two experiments with synthetically engineered eukaryotic cells. Our model captures the qualitative noise features of both experiments and accurately fit the data from the first experiment. Our results suggest that cellular switching between high and low expression states and transcriptional re-initiation are important factors needed to accurately describe gene expression noise with a structured population model.

  6. CXCL11 Expression by Keratinocytes Occurs Transiently Between Reaching Confluence and Cellular Compaction

    PubMed Central

    Huen, Arthur C.; Marathi, Archana; Nam, Peter K.; Wells, Alan

    2016-01-01

    Objective: To investigate whether differentiation or cellular confluence is responsible for CXCL11 expression patterns in re-epithelialization. Approach: In vitro model systems of re-epithelialization using the HaCaT keratinocyte cell line were utilized in monitoring expression of differentiation markers, including desmoplakin and various cytokeratins while evaluating for an association with chemokine CXCL11 expression. Results: CXCL11 expression was elevated in sparse culture with peak expression near the time of confluence. This somewhat followed the accumulation of desmoplakin in detergent-insoluble pool of proteins. However, in postconfluent, despite continued accumulation of desmoplakin within cells, CXCL11 expression decreased to baseline levels. This biphasic pattern was also seen in low calcium culture, an environment that inhibits keratinocyte differentiation and accumulation of desmosomal proteins. Highest CXCL11-expressing areas best correlated with newly confluent areas within culture expressing basal keratin 14, but also activated keratin 6. Innovation: Achievement of a threshold cellular density induces cell signaling cascade through CXCR3 that, in addition to other undiscovered pathways, can progress cutaneous wounds from the proliferative into the remodeling phases of cutaneous wound healing. Conclusion: These results suggest that the achievement of confluence with increased cellular density by migrating keratinocytes at the wound edge triggers expression of CXCL11. Since CXCR3 stimulation in endothelial cells results in apoptosis and causes neovascular pruning, whereas stimulation of CXCR3 in fibroblasts results decreased motility and cellular contraction, we speculate that CXCL11 expression by epidermal cells upon achieving cellular confluence could be the source of CXCR3 stimulation in the dermis ushering a transition from proliferative to remodeling phases of wound healing. PMID:28078185

  7. Chemically regulated gene expression in plants.

    PubMed

    Padidam, Malla

    2003-04-01

    Chemically inducible systems that activate or inactivate gene expression have many potential applications in the determination of gene function and in plant biotechnology. The precise timing and control of gene expression are important aspects of chemically inducible systems. Several systems have been developed and used to analyze gene function, marker-free plant transformation, site-specific DNA excision, activation tagging, conditional genetic complementation, and restoration of male fertility. Chemicals that are used to regulate transgene expression include the antibiotic tetracycline, the steroids dexamethasone and estradiol, copper, ethanol, the inducer of pathogen-related proteins benzothiadiazol, herbicide safeners, and the insecticide methoxyfenozide. Systems that are suitable for field application are particularly useful for experimental systems and have potential applications in biotechnology.

  8. Phylogenetic evidence for extensive lateral acquisition of cellular genes by Nucleocytoplasmic large DNA viruses

    PubMed Central

    2008-01-01

    Background Nucleo-Cytoplasmic Large DNA viruses (NCLDV), a diverse group that infects a wide range of eukaryotic hosts, exhibit a large heterogeneity in genome size (between 100 kb and 1.2 Mb) but have been suggested to form a monophyletic group on the basis of a small subset of approximately 30 conserved genes. NCLDV were proposed to have evolved by simplification from cellular organism although some of the giant NCLDV have clearly grown by gene accretion from a bacterial origin. Results We demonstrate here that many NCLDV lineages appear to have undergone frequent gene exchange in two different ways. Viruses which infect protists directly (Mimivirus) or algae which exist as intracellular protists symbionts (Phycodnaviruses) acquire genes from a bacterial source. Metazoan viruses such as the Poxviruses show a predominant acquisition of host genes. In both cases, the laterally acquired genes show a strong tendency to be positioned at the tip of the genome. Surprisingly, several core genes believed to be ancestral in the family appear to have undergone lateral gene transfers, suggesting that the NCLDV ancestor might have had a smaller genome than previously believed. Moreover, our data show that the larger the genome, the higher is the number of laterally acquired genes. This pattern is incompatible with a genome reduction from a cellular ancestor. Conclusion We propose that the NCLDV viruses have evolved by significant growth of a simple DNA virus by gene acquisition from cellular sources. PMID:19036122

  9. Symbiont modulates expression of specific gene categories in Angomonas deanei

    PubMed Central

    Penha, Luciana Loureiro; Hoffmann, Luísa; de Souza, Silvanna Sant’Anna; Martins, Allan Cézar de Azevedo; Bottaro, Thayane; Prosdocimi, Francisco; Faffe, Débora Souza; Motta, Maria Cristina Machado; Ürményi, Turán Péter; Silva, Rosane

    2016-01-01

    Trypanosomatids are parasites that cause disease in humans, animals, and plants. Most are non-pathogenic and some harbor a symbiotic bacterium. Endosymbiosis is part of the evolutionary process of vital cell functions such as respiration and photosynthesis. Angomonas deanei is an example of a symbiont-containing trypanosomatid. In this paper, we sought to investigate how symbionts influence host cells by characterising and comparing the transcriptomes of the symbiont-containing A. deanei (wild type) and the symbiont-free aposymbiotic strains. The comparison revealed that the presence of the symbiont modulates several differentially expressed genes. Empirical analysis of differential gene expression showed that 216 of the 7625 modulated genes were significantly changed. Finally, gene set enrichment analysis revealed that the largest categories of genes that downregulated in the absence of the symbiont were those involved in oxidation-reduction process, ATP hydrolysis coupled proton transport and glycolysis. In contrast, among the upregulated gene categories were those involved in proteolysis, microtubule-based movement, and cellular metabolic process. Our results provide valuable information for dissecting the mechanism of endosymbiosis in A. deanei. PMID:27706380

  10. Expression of Alzheimer's disease risk genes in ischemic brain degeneration.

    PubMed

    Ułamek-Kozioł, Marzena; Pluta, Ryszard; Januszewski, Sławomir; Kocki, Janusz; Bogucka-Kocka, Anna; Czuczwar, Stanisław J

    2016-12-01

    We review the Alzheimer-related expression of genes following brain ischemia as risk factors for late-onset of sporadic Alzheimer's disease and their role in Alzheimer's disease ischemia-reperfusion pathogenesis. More recent advances in understanding ischemic etiology of Alzheimer's disease have revealed dysregulation of Alzheimer-associated genes including amyloid protein precursor, β-secretase, presenilin 1 and 2, autophagy, mitophagy and apoptosis. We review the relationship between these genes dysregulated by brain ischemia and the cellular and neuropathological characteristics of Alzheimer's disease. Here we summarize the latest studies supporting the theory that Alzheimer-related genes play an important role in ischemic brain injury and that ischemia is a needful and leading supplier to the onset and progression of sporadic Alzheimer's disease. Although the exact molecular mechanisms of ischemic dependent neurodegenerative disease and neuronal susceptibility finally are unknown, a downregulated expression of neuronal defense genes like alfa-secretase in the ischemic brain makes the neurons less able to resist injury. The recent challenge is to find ways to raise the adaptive reserve of the brain to overcome such ischemic-associated deficits and support and/or promote neuronal survival. Understanding the mechanisms underlying the association of these genes with risk for Alzheimer's disease will provide the most meaningful targets for therapeutic development to date.

  11. A spatial and temporal map of C. elegans gene expression.

    PubMed

    Spencer, W Clay; Zeller, Georg; Watson, Joseph D; Henz, Stefan R; Watkins, Kathie L; McWhirter, Rebecca D; Petersen, Sarah; Sreedharan, Vipin T; Widmer, Christian; Jo, Jeanyoung; Reinke, Valerie; Petrella, Lisa; Strome, Susan; Von Stetina, Stephen E; Katz, Menachem; Shaham, Shai; Rätsch, Gunnar; Miller, David M

    2011-02-01

    The C. elegans genome has been completely sequenced, and the developmental anatomy of this model organism is described at single-cell resolution. Here we utilize strategies that exploit this precisely defined architecture to link gene expression to cell type. We obtained RNAs from specific cells and from each developmental stage using tissue-specific promoters to mark cells for isolation by FACS or for mRNA extraction by the mRNA-tagging method. We then generated gene expression profiles of more than 30 different cells and developmental stages using tiling arrays. Machine-learning-based analysis detected transcripts corresponding to established gene models and revealed novel transcriptionally active regions (TARs) in noncoding domains that comprise at least 10% of the total C. elegans genome. Our results show that about 75% of transcripts with detectable expression are differentially expressed among developmental stages and across cell types. Examination of known tissue- and cell-specific transcripts validates these data sets and suggests that newly identified TARs may exercise cell-specific functions. Additionally, we used self-organizing maps to define groups of coregulated transcripts and applied regulatory element analysis to identify known transcription factor- and miRNA-binding sites, as well as novel motifs that likely function to control subsets of these genes. By using cell-specific, whole-genome profiling strategies, we have detected a large number of novel transcripts and produced high-resolution gene expression maps that provide a basis for establishing the roles of individual genes in cellular differentiation.

  12. Changes in gene expression and signal transduction in microgravity

    NASA Technical Reports Server (NTRS)

    Hughes-Fulford, M.

    2001-01-01

    Studies from space flights over the past three decades have demonstrated that basic physiological changes occur in humans during space flight. These changes include cephalic fluid shifts, loss of fluid and electrolytes, loss of muscle mass, space motion sickness, anemia, reduced immune response, and loss of calcium and mineralized bone. The cause of most of these manifestations is not known and until recently, the general approach was to investigate general systemic changes, not basic cellular responses to microgravity. This laboratory has recently studied gene growth and activation of normal osteoblasts (MC3T3-El) during spaceflight. Osteoblast cells were grown on glass coverslips and loaded in the Biorack plunger boxes. The osteoblasts were launched in a serum deprived state, activated in microgravity and collected in microgravity. The osteoblasts were examined for changes in gene expression and signal transduction. Approximately one day after growth activation significant changes were observed in gene expression in 0-G flight samples. Immediate early growth genes/growth factors cox-2, c-myc, bcl2, TGF beta1, bFGF and PCNA showed a significant diminished mRNA induction in microgravity FCS activated cells when compared to ground and 1-G flight controls. Cox-1 was not detected in any of the samples. There were no significant differences in the expression of reference gene mRNA between the ground, 0-G and 1-G samples. The data suggest that quiescent osteoblasts are slower to enter the cell cycle in microgravity and that the lack of gravity itself may be a significant factor in bone loss in spaceflight. Preliminary data from our STS 76 flight experiment support our hypothesis that a basic biological response occurs at the tissue, cellular, and molecular level in 0-G. Here we examine ground-based and space flown data to help us understand the mechanism of bone loss in microgravity.

  13. Paternally expressed genes predominate in the placenta.

    PubMed

    Wang, Xu; Miller, Donald C; Harman, Rebecca; Antczak, Douglas F; Clark, Andrew G

    2013-06-25

    The discovery of genomic imprinting through studies of manipulated mouse embryos indicated that the paternal genome has a major influence on placental development. However, previous research has not demonstrated paternal bias in imprinted genes. We applied RNA sequencing to trophoblast tissue from reciprocal hybrids of horse and donkey, where genotypic differences allowed parent-of-origin identification of most expressed genes. Using this approach, we identified a core group of 15 ancient imprinted genes, of which 10 were paternally expressed. An additional 78 candidate imprinted genes identified by RNA sequencing also showed paternal bias. Pyrosequencing was used to confirm the imprinting status of six of the genes, including the insulin receptor (INSR), which may play a role in growth regulation with its reciprocally imprinted ligand, histone acetyltransferase-1 (HAT1), a gene involved in chromatin modification, and lymphocyte antigen 6 complex, locus G6C, a newly identified imprinted gene in the major histocompatibility complex. The 78 candidate imprinted genes displayed parent-of-origin expression bias in placenta but not fetus, and most showed less than 100% silencing of the imprinted allele. Some displayed variability in imprinting status among individuals. This variability results in a unique epigenetic signature for each placenta that contributes to variation in the intrauterine environment and thus presents the opportunity for natural selection to operate on parent-of-origin differential regulation. Taken together, these features highlight the plasticity of imprinting in mammals and the central importance of the placenta as a target tissue for genomic imprinting.

  14. Hepatic Xenobiotic Metabolizing Enzyme Gene Expression ...

    EPA Pesticide Factsheets

    BACKGROUND: Differences in responses to environmental chemicals and drugs between life stages are likely due in part to differences in the expression of xenobiotic metabolizing enzymes and transporters (XMETs). No comprehensive analysis of the mRNA expression of XMETs has been carried out through life stages in any species. RESULTS: Using full-genome arrays, the mRNA expression of all XMETs and their regulatory proteins was examined during fetal (gestation day (GD) 19), neonatal (postnatal day (PND) 7), prepubescent (PND32), middle age (12 months), and old age (18 and 24 months) in the C57BL/6J (C57) mouse liver and compared to adults. Fetal and neonatal life stages exhibited dramatic differences in XMET mRNA expression compared to the relatively minor effects of old age. The total number of XMET probe sets that differed from adults was 636, 500, 84, 5, 43, and 102 for GD19, PND7, PND32, 12 months, 18 months and 24 months, respectively. At all life stages except PND32, under-expressed genes outnumbered over-expressed genes. The altered XMETs included those in all of the major metabolic and transport phases including introduction of reactive or polar groups (Phase I), conjugation (Phase II) and excretion (Phase III). In the fetus and neonate, parallel increases in expression were noted in the dioxin receptor, Nrf2 components and their regulated genes while nuclear receptors and regulated genes were generally down-regulated. Suppression of male-specific XMETs w

  15. Three gene expression vector sets for concurrently expressing multiple genes in Saccharomyces cerevisiae.

    PubMed

    Ishii, Jun; Kondo, Takashi; Makino, Harumi; Ogura, Akira; Matsuda, Fumio; Kondo, Akihiko

    2014-05-01

    Yeast has the potential to be used in bulk-scale fermentative production of fuels and chemicals due to its tolerance for low pH and robustness for autolysis. However, expression of multiple external genes in one host yeast strain is considerably labor-intensive due to the lack of polycistronic transcription. To promote the metabolic engineering of yeast, we generated systematic and convenient genetic engineering tools to express multiple genes in Saccharomyces cerevisiae. We constructed a series of multi-copy and integration vector sets for concurrently expressing two or three genes in S. cerevisiae by embedding three classical promoters. The comparative expression capabilities of the constructed vectors were monitored with green fluorescent protein, and the concurrent expression of genes was monitored with three different fluorescent proteins. Our multiple gene expression tool will be helpful to the advanced construction of genetically engineered yeast strains in a variety of research fields other than metabolic engineering.

  16. Kinetics of Kaposi's sarcoma-associated herpesvirus gene expression.

    PubMed

    Sun, R; Lin, S F; Staskus, K; Gradoville, L; Grogan, E; Haase, A; Miller, G

    1999-03-01

    Herpesvirus gene expression can be classified into four distinct kinetic stages: latent, immediate early, early, and late. Here we characterize the kinetic class of a group of 16 Kaposi's sarcoma-associated herpesvirus (KSHV)/human herpesvirus 8 genes in a cultured primary effusion cell line and examine the expression of a subset of these genes in KS biopsies. Expression of two latent genes, LANA and vFLIP, was constitutive and was not induced by chemicals that induce the lytic cycle in primary effusion lymphoma (PEL) cell lines. An immediate-early gene, Rta (open reading frame 50 [ORF50]), was induced within 4 h of the addition of n-butyrate, and its 3.6-kb mRNA was resistant to inhibition by cycloheximide. Early genes, including K3 and K5 that are homologues of the "immediate-early" gene of bovine herpesvirus 4, K8 that is a positional homologue of Epstein-Barr virus BZLF1, vMIP II, vIL-6, and polyadenylated nuclear (PAN) RNA, appeared 8 to 13 h after chemical induction. A second group of early genes that were slightly delayed in their appearance included viral DHFR, thymidylate synthase, vMIP I, G protein-coupled receptor, K12, vBcl2, and a lytic transcript that overlapped LANA. The transcript of sVCA (ORF65), a late gene whose expression was abolished by Phosphonoacetic acid, an inhibitor of KSHV DNA replication, did not appear until 30 h after induction. Single-cell assays indicated that the induction of lytic cycle transcripts resulted from the recruitment of additional cells into the lytic cycle. In situ hybridization of KS biopsies showed that about 3% of spindle-shaped tumor cells expressed Rta, ORF K8, vIL-6, vMIP I, vBcl-2, PAN RNA, and sVCA. Our study shows that several KSHV-encoded homologues of cellular cytokines, chemokines, and antiapoptotic factors are expressed during the viral lytic cycle in PEL cell lines and in KS biopsies. The lytic cycle of KSHV, probably under the initial control of the KSHV/Rta gene, may directly contribute to tumor

  17. Differential effects of detergents on keratinocyte gene expression.

    PubMed

    van Ruissen, F; Le, M; Carroll, J M; van der Valk, P G; Schalkwijk, J

    1998-04-01

    We have studied the effect of various detergents on keratinocyte gene expression in vitro, using an anionic detergent (sodium dodecyl sulfate), a cationic detergent cetyltrimethylammoniumbromide (CTAB), and two nonionic detergents, Nonidet P-40 and Tween-20. We measured the effect of these detergents on direct cellular toxicity (lactate dehydrogenase release), on the expression of markers for normal differentiation (cytokeratin 1 and involucrin expression), and on disturbed keratinocyte differentiation (SKALP) by northern blot analysis. As reported in other studies, large differences were noted in direct cellular toxicity. In a culture model that mimics normal epidermal differentiation we found that low concentrations of sodium dodecyl sulfate could induce the expression of SKALP, a proteinase inhibitor that is not normally expressed in human epidermis but is found in hyperproliferative skin. Sodium dodecyl sulfate caused upregulation of involucrin and downregulation of cytokeratin 1 expression, which is associated with the hyperproliferative/inflammatory epidermal phenotype found in psoriasis, wound healing, and skin irritation. These changes were not induced after treatment of cultures with CTAB, Triton X-100, and Nonidet-P40. This effect appeared to be specific for the class of anionic detergents because sodium dodecyl benzene sulfonate and sodium laurate also induced SKALP expression. These in vitro findings showed only a partial correlation with the potential of different detergents to induce clinical, biophysical, and cell biologic changes in vivo in human skin. Both sodium dodecyl sulfate and CTAB were found to cause induction and upregulation of SKALP and involucrin at low doses following a 24 h patch test, whereas high concentrations of Triton X-100 did not. Sodium dodecyl sulfate induced higher rates of transepidermal water loss, whereas CTAB treated skin showed more signs of cellular toxicity. We conclude that the action of anionic detergents on

  18. Gene expression profile of HIV-1 Tat expressing cells: a close interplay between proliferative and differentiation signals

    PubMed Central

    de la Fuente, Cynthia; Santiago, Francisco; Deng, Longwen; Eadie, Carolyne; Zilberman, Irene; Kehn, Kylene; Maddukuri, Anil; Baylor, Shanese; Wu, Kaili; Lee, Chee Gun; Pumfery, Anne; Kashanchi, Fatah

    2002-01-01

    Background Expression profiling holds great promise for rapid host genome functional analysis. It is plausible that host expression profiling in an infection could serve as a universal phenotype in virally infected cells. Here, we describe the effect of one of the most critical viral activators, Tat, in HIV-1 infected and Tat expressing cells. We utilized microarray analysis from uninfected, latently HIV-1 infected cells, as well as cells that express Tat, to decipher some of the cellular changes associated with this viral activator. Results Utilizing uninfected, HIV-1 latently infected cells, and Tat expressing cells, we observed that most of the cellular host genes in Tat expressing cells were down-regulated. The down-regulation in Tat expressing cells is most apparent on cellular receptors that have intrinsic receptor tyrosine kinase (RTK) activity and signal transduction members that mediate RTK function, including Ras-Raf-MEK pathway. Co-activators of transcription, such as p300/CBP and SRC-1, which mediate gene expression related to hormone receptor genes, were also found to be down-regulated. Down-regulation of receptors may allow latent HIV-1 infected cells to either hide from the immune system or avoid extracellular differentiation signals. Some of the genes that were up-regulated included co-receptors for HIV-1 entry, translation machinery, and cell cycle regulatory proteins. Conclusions We have demonstrated, through a microarray approach, that HIV-1 Tat is able to regulate many cellular genes that are involved in cell signaling, translation and ultimately control the host proliferative and differentiation signals. PMID:12069692

  19. Tools and resources for analyzing gene expression changes in glaucomatous neurodegeneration.

    PubMed

    Nickells, Robert W; Pelzel, Heather R

    2015-12-01

    Evaluating gene expression changes presents one of the most powerful interrogative approaches to study the molecular, biochemical, and cellular pathways associated with glaucomatous disease pathology. Technologies to study gene expression profiles in glaucoma are wide ranging. Qualitative techniques provide the power of localizing expression changes to individual cells, but are not robust to evaluate differences in expression changes. Alternatively, quantitative changes provide a high level of stringency to quantify changes in gene expression. Additionally, advances in high throughput analysis and bioinformatics have dramatically improved the number of individual genes that can be evaluated in a single experiment, while dramatically reducing amounts of input tissue/starting material. Together, gene expression profiling and proteomics have yielded new insights on the roles of neuroinflammation, the complement cascade, and metabolic shutdown as important players in the pathology of the optic nerve head and retina in this disease.

  20. Tools and resources for analyzing gene expression changes in glaucomatous neurodegeneration

    PubMed Central

    Nickells, Robert W.; Pelzel, Heather R.

    2015-01-01

    Evaluating gene expression changes presents one of the most powerful interrogative approaches to study the molecular, biochemical, and cellular pathways associated with glaucomatous disease pathology. Technologies to study gene expression profiles in glaucoma are wide ranging. Qualitative techniques provide the power of localizing expression changes to individual cells, but are not robust to evaluate differences in expression changes. Alternatively, quantitative changes provide a high level of stringency to quantify changes in gene expression. Additionally, advances in high throughput analysis and bioinformatics have dramatically improved the number of individual genes that can be evaluated in a single experiment, while dramatically reducing amounts of input tissue/starting material. Together, gene expression profiling and proteomics have yielded new insights on the roles of neuroinflammation, the complement cascade, and metabolic shutdown as important players in the pathology of the optic nerve head and retina in this disease. PMID:25999234

  1. Gene expression profiles of FABP genes in protochordates, Ciona intestinalis and Branchiostoma belcheri.

    PubMed

    Orito, Wataru; Ohhira, Fuyuko; Ogasawara, Michio

    2015-11-01

    Fatty-acid-binding proteins (FABPs) are small intracellular proteins associated with the transportation of fatty acids. Members of the FABPs share similar amino acid sequences and tertiary structures and form, together with a member of the cellular retinol-binding proteins (CRBPs), the intracellular-lipid-binding protein (iLBP) family. In vertebrates, several types of FABP have been isolated and classified into three subfamilies: 2-4. In invertebrates, several FABP-related proteins have been reported in protostomes and amphioxus; however, little is known about the relationship between their phylogenetic positions and expression patterns. We have performed a genome-wide survey of FABP-related genes in protochordates: amphioxus Branchiostoma belcheri and the ascidian Ciona intestinalis. Comprehensive BLAST searches in NCBI and the Ciona Ghost Database by using amino acid sequences of all FABPs have revealed that the ascidian C. intestinalis and amphioxus B. belcheri contain six and seven FABP-related genes in their haploid genomes, respectively. Expression pattern analyses by whole-mount in situ hybridization in Ciona transparent juveniles and serial-section in situ hybridizations in adult amphioxus have revealed that all genes are mainly expressed in the postpharyngeal digestive tract. In particular, the expression of FABP-related genes of subfamily-2 (liver/ileum type) and subfamily-3 (intestinal type) in the ascidian pyloric gland and amphioxus hepatic cecum provides insight into the evolution of hepatic-related structures of chordates and FABP-related genes.

  2. Expression of myriapod pair rule gene orthologs

    PubMed Central

    2011-01-01

    Background Segmentation is a hallmark of the arthropods; most knowledge about the molecular basis of arthropod segmentation comes from work on the fly Drosophila melanogaster. In this species a hierarchic cascade of segmentation genes subdivides the blastoderm stepwise into single segment wide regions. However, segmentation in the fly is a derived feature since all segments form virtually simultaneously. Conversely, in the vast majority of arthropods the posterior segments form one at a time from a posterior pre-segmental zone. The pair rule genes (PRGs) comprise an important level of the Drosophila segmentation gene cascade and are indeed the first genes that are expressed in typical transverse stripes in the early embryo. Information on expression and function of PRGs outside the insects, however, is scarce. Results Here we present the expression of the pair rule gene orthologs in the pill millipede Glomeris marginata (Myriapoda: Diplopoda). We find evidence that these genes are involved in segmentation and that components of the hierarchic interaction of the gene network as found in insects may be conserved. We further provide evidence that segments are formed in a single-segment periodicity rather than in pairs of two like in another myriapod, the centipede Strigamia maritima. Finally we show that decoupling of dorsal and ventral segmentation in Glomeris appears already at the level of the PRGs. Conclusions Although the pair rule gene network is partially conserved among insects and myriapods, some aspects of PRG interaction are, as suggested by expression pattern analysis, convergent, even within the Myriapoda. Conserved expression patterns of PRGs in insects and myriapods, however, may represent ancestral features involved in segmenting the arthropod ancestor. PMID:21352542

  3. Human AZU-1 gene, variants thereof and expressed gene products

    DOEpatents

    Chen, Huei-Mei; Bissell, Mina

    2004-06-22

    A human AZU-1 gene, mutants, variants and fragments thereof. Protein products encoded by the AZU-1 gene and homologs encoded by the variants of AZU-1 gene acting as tumor suppressors or markers of malignancy progression and tumorigenicity reversion. Identification, isolation and characterization of AZU-1 and AZU-2 genes localized to a tumor suppressive locus at chromosome 10q26, highly expressed in nonmalignant and premalignant cells derived from a human breast tumor progression model. A recombinant full length protein sequences encoded by the AZU-1 gene and nucleotide sequences of AZU-1 and AZU-2 genes and variant and fragments thereof. Monoclonal or polyclonal antibodies specific to AZU-1, AZU-2 encoded protein and to AZU-1, or AZU-2 encoded protein homologs.

  4. Rubisco gene expression in C4 plants.

    PubMed

    Patel, Minesh; Berry, James O

    2008-01-01

    In leaves of most C(4) plants, ribulose 1,5 bisphosphate carboxylase (Rubisco) accumulates only in bundle sheath (bs) cells that surround the vascular centres, and not in mesophyll (mp) cells. It has been shown previously that in the C(4) dicots amaranth and Flaveria bidentis, post-transcriptional control of mRNA translation and stability mediate the C(4) expression patterns of genes encoding the large and small Rubisco subunits (chloroplast rbcL and nuclear RbcS, respectively). Translational control appears to regulate bs cell-specific Rubisco gene expression during early dicot leaf development, while control of mRNA stability appears to mediate bs-specific accumulation of RbcS and rbcL transcripts in mature leaves. Post-transcriptional control is also involved in the regulation of Rubisco gene expression by light, and in response to photosynthetic activity. Transgenic and transient expression studies in F. bidentis provide direct evidence for post-transcriptional control of bs cell-specific RbcS expression, which is mediated by the 5' and 3' untranslated regions (UTRs) of the mRNA. Comparisons of Rubisco gene expression in these dicots and in the monocot maize indicates possible commonalities in the regulation of RbcS and rbcL genes in these divergent C(4) species. Now that the role of post-transcriptional regulation in C(4) gene expression has been established, it is likely that future studies of mRNA-protein interactions will address long-standing questions about the establishment and maintenance of cell type-specificity in these plants. Some of these regulatory mechanisms may have ancestral origins in C(3) species, through modification of pre-existing factors, or by the acquisition of novel C(4) processes.

  5. BDNF and NT-3 expression by using glucocorticoid-induced bicistronic expression vector pGC-BDNF-IRES-NT3 protects apoptotic cells in a cellular injury model.

    PubMed

    Wang, Yongxiang; Gu, Jiaxiang; Wang, Jingcheng; Feng, Xingmin; Tao, Yuping; Jiang, Baichuan; He, Jinshan; Wang, Qiang; Yang, Jiandong; Zhang, Shenfei; Cai, Jun; Sun, Yu

    2012-04-11

    Spinal cord injury (SCI) is a severe traumatic disease in the central nervous system with high incidence and high morbidity. Recent study demonstrated that cell transplantation therapy may improve local microenvironment of the injury site and promote nerve regeneration to restore spinal cord functions. In this study, we constructed a glucocorticoid-induced bicistronic eukaryotic expression vector pGC-BDNF-IRES-NT3 by using molecular cloning techniques and examined the protective effect of neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) expressed by this vector in a rat spinal cord injury (SCI) model. We first connected glucocorticoid response element (GRE) to cytomegalovirus (CMV) promoter and then the GRE-CMV gene was inserted into pEGFP-1 vector to construct the eukaryotic expression vector pGC-EGFP. Western blot analysis was used to confirm the expression of EGFP by transfecting this vector in RN-DSC cells. The IRES was used to connect BDNF gene and NT-3 gene and replaced the EGFP gene in pGC-EGFP plasmid to form the bicistronic expression vector-pGC-BDNF-IRES-NT3. After RN-DSC cells were transfected with the plasmid and treated with glucocorticoid, BDNF and NT-3 expression in the culture medium were measured by ELISA method. Finally, we found that combination therapy with the transfection of this vector and glucocorticoid had an anti-apoptotic effect in a cellular SCI model of RN-DSC cells. Therefore, the co-expression of BDNF and NT-3 by using this vector rescued the injured cells. This provided useful information for the gene-modification cell transplantation combined with glucocorticoid for the treatment of SCI.

  6. Alternative-splicing-mediated gene expression

    NASA Astrophysics Data System (ADS)

    Wang, Qianliang; Zhou, Tianshou

    2014-01-01

    Alternative splicing (AS) is a fundamental process during gene expression and has been found to be ubiquitous in eukaryotes. However, how AS impacts gene expression levels both quantitatively and qualitatively remains to be fully explored. Here, we analyze two common models of gene expression, each incorporating a simple splice mechanism that a pre-mRNA is spliced into two mature mRNA isoforms in a probabilistic manner. In the constitutive expression case, we show that the steady-state molecular numbers of two mature mRNA isoforms follow mutually independent Poisson distributions. In the bursting expression case, we demonstrate that the tail decay of the steady-state distribution for both mature mRNA isoforms that in general are not mutually independent can be characterized by the product of mean burst size and splicing probability. In both cases, we find that AS can efficiently modulate both the variability (measured by variance) and the noise level of the total mature mRNA, and in particular, the latter is always lower than the noise level of the pre-mRNA, implying that AS always reduces the noise. These results altogether reveal that AS is a mechanism of efficiently controlling the gene expression noise.

  7. Molecular mechanisms of alpha-fetoprotein gene expression.

    PubMed

    Lazarevich, N L

    2000-01-01

    Alpha-fetoprotein (AFP) is the main component of mammalian fetal serum. It is synthesized by visceral endoderm of the yolk sac and by fetal liver. Immediately after birth AFP level in blood decreases dramatically. AFP synthesis is reactivated in liver tumors and germinogeneous teratoblastomas, in a lesser degree after chemical and mechanical liver injuries followed by regeneration (i.e., acute viral hepatitis). AFP blood level change is an important marker for liver tumors that is widely used in clinical practice. Therefore, the study of the molecular and cellular mechanisms participating in regulation of the oncoembryonal protein AFP is an important task. On various experimental models it has been shown that the expression is regulated mainly on the transcriptional level, the AFP gene having a 7 kb regulatory region upstream. Within this region a tissue-specific promoter, three independent enhancers, and a silencer that is at least partially responsible for AFP gene expression decrease in adult liver have been defined. Some ubiquitous and some tissue-specific transcription factors, including hepatocyte nuclear factors (HNFs), which mediate the transcription of most of the liver-specific genes, have been shown to bind to the promoter. However, the mechanisms determining drastic changes of AFP synthesis level in the course of ontogenesis and carcinogenesis remain incompletely clarified. Also, little is known about negative regulators of AFP gene expression in cells of non-hepatic origin and in adult liver.

  8. Gene activation properties of a mouse DNA sequence isolated by expression selection.

    PubMed Central

    von Hoyningen-Huene, V; Norbury, C; Griffiths, M; Fried, M

    1986-01-01

    The MES-1 element was previously isolated from restricted total mouse cellular DNA by "expression selection"--the ability to reactivate expression of a test gene devoid of its 5' enhancer sequences. Mes-1 has been tested in long-term transformation and short-term CAT expression assays. In both assays MES-1 is active independent of orientation and at a distance when placed 5' to the test gene. The element is active with heterologous promoters and functions efficiently in both rat and mouse cells. MES-1 activates expression by increasing transcription from the test gene's own start (cap) site. Thus the expression selection technique can be used for the isolation of DNA sequences with enhancer-like properties from total cellular DNA. Images PMID:3016657

  9. The Interrelationship between Promoter Strength, Gene Expression, and Growth Rate

    PubMed Central

    Klesmith, Justin R.; Detwiler, Emily E.; Tomek, Kyle J.; Whitehead, Timothy A.

    2014-01-01

    In exponentially growing bacteria, expression of heterologous protein impedes cellular growth rates. Quantitative understanding of the relationship between expression and growth rate will advance our ability to forward engineer bacteria, important for metabolic engineering and synthetic biology applications. Recently, a work described a scaling model based on optimal allocation of ribosomes for protein translation. This model quantitatively predicts a linear relationship between microbial growth rate and heterologous protein expression with no free parameters. With the aim of validating this model, we have rigorously quantified the fitness cost of gene expression by using a library of synthetic constitutive promoters to drive expression of two separate proteins (eGFP and amiE) in E. coli in different strains and growth media. In all cases, we demonstrate that the fitness cost is consistent with the previous findings. We expand upon the previous theory by introducing a simple promoter activity model to quantitatively predict how basal promoter strength relates to growth rate and protein expression. We then estimate the amount of protein expression needed to support high flux through a heterologous metabolic pathway and predict the sizable fitness cost associated with enzyme production. This work has broad implications across applied biological sciences because it allows for prediction of the interplay between promoter strength, protein expression, and the resulting cost to microbial growth rates. PMID:25286161

  10. Genomic modulators of gene expression in human neutrophils

    PubMed Central

    Naranbhai, Vivek; Fairfax, Benjamin P.; Makino, Seiko; Humburg, Peter; Wong, Daniel; Ng, Esther; Hill, Adrian V. S.; Knight, Julian C.

    2015-01-01

    Neutrophils form the most abundant leukocyte subset and are central to many disease processes. Technical challenges in transcriptomic profiling have prohibited genomic approaches to date. Here we map expression quantitative trait loci (eQTL) in peripheral blood CD16+ neutrophils from 101 healthy European adults. We identify cis-eQTL for 3281 neutrophil-expressed genes including many implicated in neutrophil function, with 450 of these not previously observed in myeloid or lymphoid cells. Paired comparison with monocyte eQTL demonstrates nuanced conditioning of genetic regulation of gene expression by cellular context, which relates to cell-type-specific DNA methylation and histone modifications. Neutrophil eQTL are markedly enriched for trait-associated variants particularly autoimmune, allergy and infectious disease. We further demonstrate how eQTL in PADI4 and NOD2 delineate risk variant function in rheumatoid arthritis, leprosy and Crohn's disease. Taken together, these data help advance understanding of the genetics of gene expression, neutrophil biology and immune-related diseases. PMID:26151758

  11. Variability of Gene Expression Identifies Transcriptional Regulators of Early Human Embryonic Development

    PubMed Central

    Hasegawa, Yu; Taylor, Deanne; Ovchinnikov, Dmitry A.; Wolvetang, Ernst J.; de Torrenté, Laurence; Mar, Jessica C.

    2015-01-01

    An analysis of gene expression variability can provide an insightful window into how regulatory control is distributed across the transcriptome. In a single cell analysis, the inter-cellular variability of gene expression measures the consistency of transcript copy numbers observed between cells in the same population. Application of these ideas to the study of early human embryonic development may reveal important insights into the transcriptional programs controlling this process, based on which components are most tightly regulated. Using a published single cell RNA-seq data set of human embryos collected at four-cell, eight-cell, morula and blastocyst stages, we identified genes with the most stable, invariant expression across all four developmental stages. Stably-expressed genes were found to be enriched for those sharing indispensable features, including essentiality, haploinsufficiency, and ubiquitous expression. The stable genes were less likely to be associated with loss-of-function variant genes or human recessive disease genes affected by a DNA copy number variant deletion, suggesting that stable genes have a functional impact on the regulation of some of the basic cellular processes. Genes with low expression variability at early stages of development are involved in regulation of DNA methylation, responses to hypoxia and telomerase activity, whereas by the blastocyst stage, low-variability genes are enriched for metabolic processes as well as telomerase signaling. Based on changes in expression variability, we identified a putative set of gene expression markers of morulae and blastocyst stages. Experimental validation of a blastocyst-expressed variability marker demonstrated that HDDC2 plays a role in the maintenance of pluripotency in human ES and iPS cells. Collectively our analyses identified new regulators involved in human embryonic development that would have otherwise been missed using methods that focus on assessment of the average expression

  12. [Expression and immunity of multi-HIV B'/C subype genes in replicating DNA vaccines].

    PubMed

    Gao, Ying-ying; Deng, Yao; Qi, Xiang-rong; Zhang, Xiang-min; Meng, Xin; Wang, Hui-juan; Tan, Wen-jie; Ruan, Li

    2010-05-01

    To understand the effect of various gene structures of HIV B'/C subtype on the gene expression and immunity in DNA vaccine, replicating DNA vector pSCK2 was used to construct seven DNA vaccines carrying one or more of HIV B'/C subtype genes: gagpol, gp160 and rtn (rev, tat and nef fusion gene). Immunofluorescence staining indicated that Gag, Gp160, Rev, Tat and Nef could be expressed from the seven DNA vaccines. Stronger expression was observed with the gene in single-gene expression plasmid or with the gene located at upper-IRES in double- or multi-gene expression plasmid. ELISA test showed that Gag induced higher antibody response, but the antibody titers stimulated by Gp160, Pol, or RTN were very low. Both Gag single-gene expression plasmid and Gag-RTN double-gene expression plasmid separately inoculating induced stronger antibody response against Gag than Gag-Gp160 double-gene expression plasmid and Gagpol-Gp160-RTN multi-gene expression plasmid or combined inoculation of Gag and Gp160 single-gene expression plasmids did. ELISPOT detection showed that all the seven DNA vaccines could stimulate cellular immune response against Gag, Pol, Gp160, Tat, and Nef, respectively. Gagpol or Gp160 single-gene expression plasmid separately inoculating stimulated the strongest cellular immune response. Tat and Nef expressed in all the plasmids induced similar immune response. These results indicated that HIV B'/C subtype genes gagpol, gp160 and rtn could be efficiently expressed in the replicating DNA vaccine vector, single-gene expression plasmid had the higher gene expression level and induced stronger immune response; combined immunization of Gagpol and Gp160 had dramatically lower immunity than Gagpol or Gp160 separated immunization did. Immunity of RTN had no difference between combined and separated immunizations. Therefore, in case of immunization with DNA vaccines containing different HIV genes, it is necessary to optimize the combined immunization procedure

  13. Visual exploration of three-dimensional gene expression using physical views and linked abstract views.

    PubMed

    Weber, Gunther H; Rübel, Oliver; Huang, Min-Yu; DePace, Angela H; Fowlkes, Charless C; Keränen, Soile V E; Luengo Hendriks, Cris L; Hagen, Hans; Knowles, David W; Malik, Jitendra; Biggin, Mark D; Hamann, Bernd

    2009-01-01

    During animal development, complex patterns of gene expression provide positional information within the embryo. To better understand the underlying gene regulatory networks, the Berkeley Drosophila Transcription Network Project (BDTNP) has developed methods that support quantitative computational analysis of three-dimensional (3D) gene expression in early Drosophila embryos at cellular resolution. We introduce PointCloudXplore (PCX), an interactive visualization tool that supports visual exploration of relationships between different genes' expression using a combination of established visualization techniques. Two aspects of gene expression are of particular interest: 1) gene expression patterns defined by the spatial locations of cells expressing a gene and 2) relationships between the expression levels of multiple genes. PCX provides users with two corresponding classes of data views: 1) Physical Views based on the spatial relationships of cells in the embryo and 2) Abstract Views that discard spatial information and plot expression levels of multiple genes with respect to each other. Cell Selectors highlight data associated with subsets of embryo cells within a View. Using linking, these selected cells can be viewed in multiple representations. We describe PCX as a 3D gene expression visualization tool and provide examples of how it has been used by BDTNP biologists to generate new hypotheses.

  14. Analysis of Host Gene Expression Profile in HIV-1 and HIV-2 Infected T-Cells.

    PubMed

    Devadas, Krishnakumar; Biswas, Santanu; Haleyurgirisetty, Mohan; Wood, Owen; Ragupathy, Viswanath; Lee, Sherwin; Hewlett, Indira

    2016-01-01

    HIV replication is closely regulated by a complex pathway of host factors, many of them being determinants of cell tropism and host susceptibility to HIV infection. These host factors are known to exert a positive or negative influence on the replication of the two major types of HIV, HIV-1 and HIV-2, thereby modulating virus infectivity, host response to infection and ultimately disease progression profiles characteristic of these two types. Understanding the differential regulation of host cellular factors in response to HIV-1 and HIV-2 infections will help us to understand the apparent differences in rates of disease progression and pathogenesis. This knowledge would aid in the discovery of new biomarkers that may serve as novel targets for therapy and diagnosis. The objective of this study was to determine the differential expression of host genes in response to HIV-1/HIV-2 infection. To achieve this, we analyzed the effects of HIV-1 (MN) and HIV-2 (ROD) infection on the expression of host factors in PBMC at the RNA level using the Agilent Whole Human Genome Oligo Microarray. Differentially expressed genes were identified and their biological functions determined. Host gene expression profiles were significantly changed. Gene expression profiling analysis identified a subset of differentially expressed genes in HIV-1 and HIV-2 infected cells. Genes involved in cellular metabolism, apoptosis, immune cell proliferation and activation, cytokines, chemokines, and transcription factors were differentially expressed in HIV-1 infected cells. Relatively few genes were differentially expressed in cells infected with HIV-2.

  15. Analysis of Host Gene Expression Profile in HIV-1 and HIV-2 Infected T-Cells

    PubMed Central

    Devadas, Krishnakumar; Biswas, Santanu; Haleyurgirisetty, Mohan; Wood, Owen; Ragupathy, Viswanath; Lee, Sherwin; Hewlett, Indira

    2016-01-01

    HIV replication is closely regulated by a complex pathway of host factors, many of them being determinants of cell tropism and host susceptibility to HIV infection. These host factors are known to exert a positive or negative influence on the replication of the two major types of HIV, HIV-1 and HIV-2, thereby modulating virus infectivity, host response to infection and ultimately disease progression profiles characteristic of these two types. Understanding the differential regulation of host cellular factors in response to HIV-1 and HIV-2 infections will help us to understand the apparent differences in rates of disease progression and pathogenesis. This knowledge would aid in the discovery of new biomarkers that may serve as novel targets for therapy and diagnosis. The objective of this study was to determine the differential expression of host genes in response to HIV-1/HIV-2 infection. To achieve this, we analyzed the effects of HIV-1 (MN) and HIV-2 (ROD) infection on the expression of host factors in PBMC at the RNA level using the Agilent Whole Human Genome Oligo Microarray. Differentially expressed genes were identified and their biological functions determined. Host gene expression profiles were significantly changed. Gene expression profiling analysis identified a subset of differentially expressed genes in HIV-1 and HIV-2 infected cells. Genes involved in cellular metabolism, apoptosis, immune cell proliferation and activation, cytokines, chemokines, and transcription factors were differentially expressed in HIV-1 infected cells. Relatively few genes were differentially expressed in cells infected with HIV-2. PMID:26821323

  16. The CD27+ memory B cells display changes in the gene expression pattern in elderly individuals

    PubMed Central

    Báez, Alicia; Álvarez-Laderas, Isabel; Piruat, José I; Caballero-Velázquez, Teresa; Barbado, María Victoria; Millán-Uclés, África; Medrano, Mayte; García-Guerrero, Estefanía; Sánchez-Abarca, Luis Ignacio; Pérez-Simón, José Antonio

    2015-01-01

    Memory B cells (MBCs) have a long lifespan compared with naive B cells (NBCs), remaining viable for years. This could predispose them to suffer misbalances in the gene expression pattern in the long term, which might be involved in the development of age-related B-cell disorders. In order to identify genes whose expression might change during life, we analysed the gene expression patterns of CD27− NBCs versus CD27+ MBCs in young and old subjects. Using microarray assays we observed that the expression pattern of CD27− NBCs versus CD27+ MBCs is significantly different. Furthermore, to evaluate the age effect, we compared the gene expression pattern of young versus aged subjects in both cell populations. Interestingly, we did not find significant differences in the CD27− NBC population between young and aged individuals, whereas we found 925 genes differentially expressed in CD27+ MBCs. Among these genes, 193 were also differentially expressed in CD27+ MBCs compared with CD27− NBCs, most of them involved in cell survival, cell growth and proliferation, cellular development and gene expression. We conclude that gene expression profiles of CD27− NBCs and CD27+ MBCs are different. Moreover, whereas the gene expression pattern of CD27+ MBCs varies with age, the same does not happen in CD27− NBCs. This suggests that MBCs undergo time-dependent changes, which could underlie a higher susceptibility to dysfunction with age. PMID:25196729

  17. Fasting-induced changes in ECL cell gene expression.

    PubMed

    Lambrecht, Nils W G; Yakubov, Iskandar; Sachs, George

    2007-10-22

    Gastric enterochromaffin-like (ECL) cells release histamine in response to food because of elevation of gastrin and neural release of pituitary adenylate cyclase-activating peptide (PACAP). Acid secretion is at a basal level in the absence of food but is rapidly stimulated with feeding. Rats fasted for 24 h showed a significant decrease of mucosal histamine despite steady-state expression of the histamine-synthesizing enzyme histidine decarboxylase (HDC). Comparative transcriptomal analysis using gene expression oligonucleotide microarrays of 95% pure ECL cells from fed and 24-h fasted rats, thereby eliminating mRNA contamination from other gastric mucosal cell types, identified significantly increased gene expression of the enzymes histidase and urocanase catabolizing the HDC substrate L-histidine but significantly decreased expression of the cellular L-histidine uptake transporter SN2 and of the vesicular monoamine transporter 2 (VMAT-2) responsible for histamine uptake into secretory vesicles. This was confirmed by reverse transcriptase-quantitative polymerase chain reaction of gastric fundic mucosal samples from fed and 24-h fasted rats. The decrease of VMAT-2 gene expression was also shown by a decrease in VMAT-2 protein content in protein extracts from fed and 24-h fasted rats compared with equal amounts of HDC protein and Na-K-ATPase alpha(1)-subunit protein content. These results indicate that rat gastric ECL cells regulate their histamine content during 24-h fasting not by a change in HDC gene or protein expression but by regulation of substrate concentration for HDC and a decreased histamine secretory pool.

  18. EvoCor: a platform for predicting functionally related genes using phylogenetic and expression profiles.

    PubMed

    Dittmar, W James; McIver, Lauren; Michalak, Pawel; Garner, Harold R; Valdez, Gregorio

    2014-07-01

    The wealth of publicly available gene expression and genomic data provides unique opportunities for computational inference to discover groups of genes that function to control specific cellular processes. Such genes are likely to have co-evolved and be expressed in the same tissues and cells. Unfortunately, the expertise and computational resources required to compare tens of genomes and gene expression data sets make this type of analysis difficult for the average end-user. Here, we describe the implementation of a web server that predicts genes involved in affecting specific cellular processes together with a gene of interest. We termed the server 'EvoCor', to denote that it detects functional relationships among genes through evolutionary analysis and gene expression correlation. This web server integrates profiles of sequence divergence derived by a Hidden Markov Model (HMM) and tissue-wide gene expression patterns to determine putative functional linkages between pairs of genes. This server is easy to use and freely available at http://pilot-hmm.vbi.vt.edu/.

  19. Visualizing Gene Expression In Situ

    SciTech Connect

    Burlage, R.S.

    1998-11-02

    Visualizing bacterial cells and describing their responses to the environment are difficult tasks. Their small size is the chief reason for the difficulty, which means that we must often use many millions of cells in a sample in order to determine what the average response of the bacteria is. However, an average response can sometimes mask important events in bacterial physiology, which means that our understanding of these organisms will suffer. We have used a variety of instruments to visualize bacterial cells, all of which tell us something different about the sample. We use a fluorescence activated cell sorter to sort cells based on the fluorescence provided by bioreporter genes, and these can be used to select for particular genetic mutations. Cells can be visualized by epifluorescent microscopy, and sensitive photodetectors can be added that allow us to find a single bacterial cell that is fluorescent or bioluminescent. We have also used standard photomultipliers to examine cell aggregates as field bioreporter microorganisms. Examples of each of these instruments show how our understanding of bacterial physiology has changed with the technology.

  20. Gene expression profile in pelvic organ prolapse†

    PubMed Central

    Brizzolara, S.S.; Killeen, J.; Urschitz, J.

    2009-01-01

    It was hypothesized that the processes contributing to pelvic organ prolapse (POP) may be identified by transcriptional profiling of pelvic connective tissue in conjunction with light microscopy. In order to test this, we performed a frequency-matched case–control study of women undergoing hysterectomy for POP and controls. Total RNA, extracted from uterosacral and round ligament samples used to generate labeled cRNA, was hybridized to microarrays and analyzed for the expression of 32 878 genes. Significance Analysis of Microarrays (Stanford University, CA, USA) identified differentially expressed genes used for ontoanalysis. Quantitative PCR (qPCR) confirmed results. Light microscopy confirmed the tissue type and assessed inflammatory infiltration. The analysis of 34 arrays revealed 249 differentially expressed genes with fold changes (FC) larger than 1.5 and false discovery rates ≤5.2%. Immunity and defense was the most significant biological process differentially expressed in POP. qPCR confirmed the elevated steady-state mRNA levels for four genes: interleukin-6 (FC 9.8), thrombospondin 1 (FC 3.5) and prostaglandin-endoperoxide synthase 2 (FC 2.4) and activating transcription factor 3 (FC 2.6). Light microscopy showed all the samples were composed of fibromuscular connective tissue with no inflammatory infiltrates. In conclusion, genes enriched for ‘immunity and defense’ contribute to POP independent of inflammatory infiltrates. PMID:19056808

  1. Clustering of High Throughput Gene Expression Data

    PubMed Central

    Pirim, Harun; Ekşioğlu, Burak; Perkins, Andy; Yüceer, Çetin

    2012-01-01

    High throughput biological data need to be processed, analyzed, and interpreted to address problems in life sciences. Bioinformatics, computational biology, and systems biology deal with biological problems using computational methods. Clustering is one of the methods used to gain insight into biological processes, particularly at the genomics level. Clearly, clustering can be used in many areas of biological data analysis. However, this paper presents a review of the current clustering algorithms designed especially for analyzing gene expression data. It is also intended to introduce one of the main problems in bioinformatics - clustering gene expression data - to the operations research community. PMID:23144527

  2. Facilitated diffusion buffers noise in gene expression.

    PubMed

    Schoech, Armin P; Zabet, Nicolae Radu

    2014-09-01

    Transcription factors perform facilitated diffusion [three-dimensional (3D) diffusion in the cytosol and 1D diffusion on the DNA] when binding to their target sites to regulate gene expression. Here, we investigated the influence of this binding mechanism on the noise in gene expression. Our results showed that, for biologically relevant parameters, the binding process can be represented by a two-state Markov model and that the accelerated target finding due to facilitated diffusion leads to a reduction in both the mRNA and the protein noise.

  3. Facilitated diffusion buffers noise in gene expression

    NASA Astrophysics Data System (ADS)

    Schoech, Armin P.; Zabet, Nicolae Radu

    2014-09-01

    Transcription factors perform facilitated diffusion [three-dimensional (3D) diffusion in the cytosol and 1D diffusion on the DNA] when binding to their target sites to regulate gene expression. Here, we investigated the influence of this binding mechanism on the noise in gene expression. Our results showed that, for biologically relevant parameters, the binding process can be represented by a two-state Markov model and that the accelerated target finding due to facilitated diffusion leads to a reduction in both the mRNA and the protein noise.

  4. Objective and subjective probability in gene expression.

    PubMed

    Velasco, Joel D

    2012-09-01

    In this paper I address the question of whether the probabilities that appear in models of stochastic gene expression are objective or subjective. I argue that while our best models of the phenomena in question are stochastic models, this fact should not lead us to automatically assume that the processes are inherently stochastic. After distinguishing between models and reality, I give a brief introduction to the philosophical problem of the interpretation of probability statements. I argue that the objective vs. subjective distinction is a false dichotomy and is an unhelpful distinction in this case. Instead, the probabilities in our models of gene expression exhibit standard features of both objectivity and subjectivity.

  5. Inferring biological functions and associated transcriptional regulators using gene set expression coherence analysis

    PubMed Central

    Kim, Tae-Min; Chung, Yeun-Jun; Rhyu, Mun-Gan; Ho Jung, Myeong

    2007-01-01

    Background Gene clustering has been widely used to group genes with similar expression pattern in microarray data analysis. Subsequent enrichment analysis using predefined gene sets can provide clues on which functional themes or regulatory sequence motifs are associated with individual gene clusters. In spite of the potential utility, gene clustering and enrichment analysis have been used in separate platforms, thus, the development of integrative algorithm linking both methods is highly challenging. Results In this study, we propose an algorithm for discovery of molecular functions and elucidation of transcriptional logics using two kinds of gene information, functional and regulatory motif gene sets. The algorithm, termed gene set expression coherence analysis first selects functional gene sets with significantly high expression coherences. Those candidate gene sets are further processed into a number of functionally related themes or functional clusters according to the expression similarities. Each functional cluster is then, investigated for the enrichment of transcriptional regulatory motifs using modified gene set enrichment analysis and regulatory motif gene sets. The method was tested for two publicly available expression profiles representing murine myogenesis and erythropoiesis. For respective profiles, our algorithm identified myocyte- and erythrocyte-related molecular functions, along with the putative transcriptional regulators for the corresponding molecular functions. Conclusion As an integrative and comprehensive method for the analysis of large-scaled gene expression profiles, our method is able to generate a set of testable hypotheses: the transcriptional regulator X regulates function Y under cellular condition Z. GSECA algorithm is implemented into freely available software package. PMID:18021416

  6. Genomic signatures of germline gene expression.

    PubMed

    McVicker, Graham; Green, Phil

    2010-11-01

    Transcribed regions in the human genome differ from adjacent intergenic regions in transposable element density, crossover rates, and asymmetric substitution and sequence composition patterns. We tested whether these differences reflect selection or are instead a byproduct of germline transcription, using publicly available gene expression data from a variety of germline and somatic tissues. Crossover rate shows a strong negative correlation with gene expression in meiotic tissues, suggesting that crossover is inhibited by transcription. Strand-biased composition (G+T content) and A → G versus T → C substitution asymmetry are both positively correlated with germline gene expression. We find no evidence for a strand bias in allele frequency data, implying that the substitution asymmetry reflects a mutation rather than a fixation bias. The density of transposable elements is positively correlated with germline expression, suggesting that such elements preferentially insert into regions that are actively transcribed. For each of the features examined, our analyses favor a nonselective explanation for the observed trends and point to the role of germline gene expression in shaping the mammalian genome.

  7. Comparison of microarray and sage techniques in gene expression analysis of human glioblastoma.

    PubMed

    Kavsan, V M; Dmitrenko, V V; Shostak, K O; Bukreieva, T V; Vitak, N Y; Simirenko, O E; Malisheva, T A; Shamayev, M I; Rozumenko, V D; Zozulya, Y A

    2007-01-01

    To enhance glioblastoma (GB) marker discovery we compared gene expression in GB with human normal brain (NB) by accessing SAGE Genie web site and compared obtained results with published data. Nine GB and five NB SAGE-libraries were analyzed using the Digital Gene Expression Displayer (DGED), the results of DGED were tested by Northern blot analysis and RT-PCR of arbitrary selected genes. Review of available data from the articles on gene expression profiling by microarray-based hybridization showed as few as 35 overlapped genes with increased expression in GB. Some of them were identified in four articles, but most genes in three or even in two investigations. There was found also some differences between SAGE results of GB analysis. Digital Gene Expression Displayer approach revealed 676 genes differentially expressed in GB vs. NB with cut-off ratio: twofold change and P < or = 0.05. Differential expression of selectedgenes obtained by DGED was confirmed by Northern analysis and RT-PCR. Altogether, only 105 of 955 genes presented in published investigations were among the genes obtained by DGED. Comparison of the results obtained by microarrays and SAGE is very complicated because authors present only the most prominent differentially expressed genes. However, even available data give quite poor overlapping of genes revealed by microarrays. Some differences between results obtained by SAGE in different investigations can be explained by high dependence on the statistical methods used. As for now, the best solution to search for molecular tumor markers is to compare all available results and to select only those genes, which significant expression in tumor combined with very low expression in normal tissues was reproduced in several articles. 105 differentially expressed genes, common to both methods, can be included in the list of candidates for the molecular typing of GBs. Some genes, encoded cell surface or extra-cellular proteins may be useful for targeting

  8. [Imprinting genes and it's expression in Arabidopsis].

    PubMed

    Zhang, Hong-Yu; Xu, Pei-Zhou; Yang, Hua; Wu, Xian-Jun

    2010-07-01

    Genomic imprinting refers to the phenomenon that the expression of a gene copy depends on its parent of origin. The Arabidopsis imprinted FIS (Fertilisation-independent seed) genes, mea, fis2, and fie, play essential roles in the repression of central cell and the regulation of early endosperm development. fis mutants display two phenotypes: autonomous diploid endosperm development when fertilization is absent and un-cellularised endosperm formation when fertilization occurs. The FIS Polycomb protein complex including the above three FIS proteins catalyzes histone H3 K27 tri-methylation on target loci. DME (DEMETER), a DNA glycosylase, and AtMET1 (Methyltransferase1), a DNA methyltransferase, are involved in the regulation of imprinted expression of both mea and fis2. This review summarizes the studies on the Arabidopsis imprinted FIS genes and other related genes. Recent works have shown that the insertion of transposons may affect nearby gene expression, which may be the main driving force behind the evolution of genomic imprinting. This summary covers the achievements on Arabidopsis imprinted genes will provide important information for studies on genomic imprinting in the important crops such as rice and maize.

  9. Dysregulated MicroRNA Expression Profiles and Potential Cellular, Circulating and Polymorphic Biomarkers in Non-Hodgkin Lymphoma

    PubMed Central

    Bradshaw, Gabrielle; Sutherland, Heidi G.; Haupt, Larisa M.; Griffiths, Lyn R.

    2016-01-01

    A large number of studies have focused on identifying molecular biomarkers, including microRNAs (miRNAs) to aid in the diagnosis and prognosis of the most common subtypes of non-Hodgkin lymphoma (NHL), Diffuse Large B-cell Lymphoma and Follicular Lymphoma. NHL is difficult to diagnose and treat with many cases becoming resistant to chemotherapy, hence the need to identify improved biomarkers to aid in both diagnosis and treatment modalities. This review summarises more recent research on the dysregulated miRNA expression profiles found in NHL, as well as the regulatory role and biomarker potential of cellular and circulating miRNAs found in tissue and serum, respectively. In addition, the emerging field of research focusing on miRNA single nucleotide polymorphisms (miRSNPs) in genes of the miRNA biogenesis pathway, in miRNA genes themselves, and in their target sites may provide new insights on gene expression changes in these genes. These miRSNPs may impact miRNA networks and have been shown to play a role in a host of different cancer types including haematological malignancies. With respect to NHL, a number of SNPs in miRNA-binding sites in target genes have been shown to be associated with overall survival. PMID:27999330

  10. Sequence and gene expression evolution of paralogous genes in willows

    PubMed Central

    Harikrishnan, Srilakshmy L.; Pucholt, Pascal; Berlin, Sofia

    2015-01-01

    Whole genome duplications (WGD) have had strong impacts on species diversification by triggering evolutionary novelties, however, relatively little is known about the balance between gene loss and forces involved in the retention of duplicated genes originating from a WGD. We analyzed putative Salicoid duplicates in willows, originating from the Salicoid WGD, which took place more than 45 Mya. Contigs were constructed by de novo assembly of RNA-seq data derived from leaves and roots from two genotypes. Among the 48,508 contigs, 3,778 pairs were, based on fourfold synonymous third-codon transversion rates and syntenic positions, predicted to be Salicoid duplicates. Both copies were in most cases expressed in both tissues and 74% were significantly differentially expressed. Mean Ka/Ks was 0.23, suggesting that the Salicoid duplicates are evolving by purifying selection. Gene Ontology enrichment analyses showed that functions related to DNA- and nucleic acid binding were over-represented among the non-differentially expressed Salicoid duplicates, while functions related to biosynthesis and metabolism were over-represented among the differentially expressed Salicoid duplicates. We propose that the differentially expressed Salicoid duplicates are regulatory neo- and/or subfunctionalized, while the non-differentially expressed are dose sensitive, hence, functionally conserved. Multiple evolutionary processes, thus drive the retention of Salicoid duplicates in willows. PMID:26689951

  11. Sequence and gene expression evolution of paralogous genes in willows.

    PubMed

    Harikrishnan, Srilakshmy L; Pucholt, Pascal; Berlin, Sofia

    2015-12-22

    Whole genome duplications (WGD) have had strong impacts on species diversification by triggering evolutionary novelties, however, relatively little is known about the balance between gene loss and forces involved in the retention of duplicated genes originating from a WGD. We analyzed putative Salicoid duplicates in willows, originating from the Salicoid WGD, which took place more than 45 Mya. Contigs were constructed by de novo assembly of RNA-seq data derived from leaves and roots from two genotypes. Among the 48,508 contigs, 3,778 pairs were, based on fourfold synonymous third-codon transversion rates and syntenic positions, predicted to be Salicoid duplicates. Both copies were in most cases expressed in both tissues and 74% were significantly differentially expressed. Mean Ka/Ks was 0.23, suggesting that the Salicoid duplicates are evolving by purifying selection. Gene Ontology enrichment analyses showed that functions related to DNA- and nucleic acid binding were over-represented among the non-differentially expressed Salicoid duplicates, while functions related to biosynthesis and metabolism were over-represented among the differentially expressed Salicoid duplicates. We propose that the differentially expressed Salicoid duplicates are regulatory neo- and/or subfunctionalized, while the non-differentially expressed are dose sensitive, hence, functionally conserved. Multiple evolutionary processes, thus drive the retention of Salicoid duplicates in willows.

  12. Neonatal cellular and gene therapies for mucopolysaccharidoses: the earlier the better?

    PubMed

    Tomatsu, Shunji; Azario, Isabella; Sawamoto, Kazuki; Pievani, Alice Silvia; Biondi, Andrea; Serafini, Marta

    2016-03-01

    Mucopolysaccharidoses (MPSs) are a group of lysosomal storage disorders (LSDs). The increasing interest in newborn screening procedures for LSDs underlines the need for alternative cellular and gene therapy approaches to be developed during the perinatal period, supporting the treatment of MPS patients before the onset of clinical signs and symptoms. The rationale for considering these early therapies results from the clinical experience in the treatment of MPSs and other genetic disorders. The normal or gene-corrected hematopoiesis transplanted in patients can produce the missing protein at levels sufficient to improve and/or halt the disease-related abnormalities. However, these current therapies are only partially successful, probably due to the limited efficacy of the protein provided through the hematopoiesis. An alternative explanation is that the time at which the cellular or gene therapy procedures are performed could be too late to prevent pre-existing or progressive organ damage. Considering these aspects, in the last several years, novel cellular and gene therapy approaches have been tested in different animal models at birth, a highly early stage, showing that precocious treatment is critical to prevent long-term pathological consequences. This review provides insights into the state-of-art accomplishments made with neonatal cellular and gene-based therapies and the major barriers that need to be overcome before they can be implemented in the medical community.

  13. The TRANSFAC system on gene expression regulation.

    PubMed

    Wingender, E; Chen, X; Fricke, E; Geffers, R; Hehl, R; Liebich, I; Krull, M; Matys, V; Michael, H; Ohnhäuser, R; Prüss, M; Schacherer, F; Thiele, S; Urbach, S

    2001-01-01

    The TRANSFAC database on transcription factors and their DNA-binding sites and profiles (http://www.gene-regulation.de/) has been quantitatively extended and supplemented by a number of modules. These modules give information about pathologically relevant mutations in regulatory regions and transcription factor genes (PathoDB), scaffold/matrix attached regions (S/MARt DB), signal transduction (TRANSPATH) and gene expression sources (CYTOMER). Altogether, these distinct database modules constitute the TRANSFAC system. They are accompanied by a number of program routines for identifying potential transcription factor binding sites or for localizing individual components in the regulatory network of a cell.

  14. Identification of differentially expressed genes involved in transient regeneration of the neonatal C57BL/6J mouse heart by digital gene expression profiling.

    PubMed

    Liu, Ming; Zhu, Jin-Gai; Yu, Zhang-Bin; Song, Gui-Xian; Shen, Ya-Hui; Liu, Yao-Qiu; Zhu, Chun; Qian, Ling-Mei

    2014-06-01

    Accumulating evidence has revealed that the mammalian heart possesses a measurable capacity for renewal. Neonatal mice retain a regenerative capacity over a short time-frame (≤6 days), but this capacity is lost by 7 days of age. In the present study, differential gene expression profiling of mouse cardiac tissue was performed to further elucidate the mechanisms underlying this process. The global gene expression patterns of the neonatal C57BL/6J mouse heart were examined at three key time-points (1, 6 and 7 days old) using digital gene expression analysis. In the distribution of total clean tags, high-expression tags (>100 copies) were found to be predominant, whereas low expression tags (<5 copies) occupied the majority of distinct tag distributions. In total, 306 differentially expressed genes (DEGs) were detected in cardiac tissue, with the expression levels of 115 genes upregulated and those of 191 genes downregulated in 7-day-old mice compared with expression levels in 1- and 6-day-old mice, respectively. The expression levels of five DEGs were confirmed using quantitative polymerase chain reaction. Gene ontology analysis revealed a large proportion of DEGs distributed throughout the cell, and these DEGs were associated with binding as well as catalytic, hydrolase, transferase and molecular transducer activities. Furthermore, these genes were involved in cellular, metabolic and developmental processes, as well as biological regulation and signaling pathways. Pathway analysis identified the oxidative phosphorylation pathway to be the process most significantly putatively affected by the differential expression of these genes. These data provide the basis for future analysis of the gene expression patterns that regulate the molecular mechanism of cardiac regeneration.

  15. Marker gene tethering by nucleoporins affects gene expression in plants.

    PubMed

    Smith, Sarah; Galinha, Carla; Desset, Sophie; Tolmie, Frances; Evans, David; Tatout, Christophe; Graumann, Katja

    2015-01-01

    In non-plant systems, chromatin association with the nuclear periphery affects gene expression, where interactions with nuclear envelope proteins can repress and interactions with nucleoporins can enhance transcription. In plants, both hetero- and euchromatin can localize at the nuclear periphery, but the effect of proximity to the nuclear periphery on gene expression remains largely unknown. This study explores the putative function of Seh1 and Nup50a nucleoporins on gene expression by using the Lac Operator / Lac Repressor (LacI-LacO) system adapted to Arabidopsis thaliana. We used LacO fused to the luciferase reporter gene (LacO:Luc) to investigate whether binding of the LacO:Luc transgene to nucleoporin:LacI protein fusions alters luciferase expression. Two separate nucleoporin-LacI-YFP fusions were introduced into single insert, homozygous LacO:Luc Arabidopsis plants. Homozygous plants carrying LacO:Luc and a single insert of either Seh1-LacI-YFP or Nup50a-LacI-YFP were tested for luciferase activity and compared to plants containing LacO:Luc only. Seh1-LacI-YFP increased, while Nup50a-LacI-YFP decreased luciferase activity. Seh1-LacI-YFP accumulated at the nuclear periphery as expected, while Nup50a-LacI-YFP was nucleoplasmic and was not selected for further study. Protein and RNA levels of luciferase were quantified by western blotting and RT-qPCR, respectively. Increased luciferase activity in LacO:Luc+Seh1-LacI-YFP plants was correlated with increased luciferase protein and RNA levels. This change of luciferase expression was abolished by disruption of LacI-LacO binding by treating with IPTG in young seedlings, rosette leaves and inflorescences. This study suggests that association with the nuclear periphery is involved in the regulation of gene expression in plants.

  16. Integrating constitutive gene expression and chemoactivity: mining the NCI60 anticancer screen.

    PubMed

    Covell, David G

    2012-01-01

    Studies into the genetic origins of tumor cell chemoactivity pose significant challenges to bioinformatic mining efforts. Connections between measures of gene expression and chemoactivity have the potential to identify clinical biomarkers of compound response, cellular pathways important to efficacy and potential toxicities; all vital to anticancer drug development. An investigation has been conducted that jointly explores tumor-cell constitutive NCI60 gene expression profiles and small-molecule NCI60 growth inhibition chemoactivity profiles, viewed from novel applications of self-organizing maps (SOMs) and pathway-centric analyses of gene expressions, to identify subsets of over- and under-expressed pathway genes that discriminate chemo-sensitive and chemo-insensitive tumor cell types. Linear Discriminant Analysis (LDA) is used to quantify the accuracy of discriminating genes to predict tumor cell chemoactivity. LDA results find 15% higher prediction accuracies, using ∼30% fewer genes, for pathway-derived discriminating genes when compared to genes derived using conventional gene expression-chemoactivity correlations. The proposed pathway-centric data mining procedure was used to derive discriminating genes for ten well-known compounds. Discriminating genes were further evaluated using gene set enrichment analysis (GSEA) to reveal a cellular genetic landscape, comprised of small numbers of key over and under expressed on- and off-target pathway genes, as important for a compound's tumor cell chemoactivity. Literature-based validations are provided as support for chemo-important pathways derived from this procedure. Qualitatively similar results are found when using gene expression measurements derived from different microarray platforms. The data used in this analysis is available at http://pubchem.ncbi.nlm.nih.gov/andhttp://www.ncbi.nlm.nih.gov/projects/geo (GPL96, GSE32474).

  17. Transgenic control of perforin gene expression

    SciTech Connect

    Lichtenheld, M.G.; Podack, E.R.; Levy, R.B.

    1995-03-01

    Perforin is a pore-forming effector molecule of CTL and NK cells. To characterize perforin gene expression and its transcriptional control mechanisms in vivo, expression of a cell surface tag, i.e., human CD4, was driven by 5.1 kb of the murin perforin 5{prime} flanking and promoter region in transgenic mice. Six out of seven transgenic lines expressed the perforin-tag hybrid gene at low to intermediate levels, depending on the integration site. Transgene expression occurred in all cells that physiologically are able to express perforin. At the whole organ level, significant amounts of transgenic mRNA and endogenous perforin mRNA were co-expressed in the lymphoid organs, as well as in the lung, the ileum, the oviduct/uterus, and the bone marrow. At the single cell level, the perforin tag was present on NK cells and on CD8{sup +}, as well as on CD4{sup +} cells. Also targeted were Thy-1.2{sup +} {gamma}{delta} T cells, but not Thy-1.2{sup -} {gamma}{delta} T cells, B cells, nor monocytes. During thymic T cell development, transgene expression occurred in double negative (CD4{sup -}CD8{sup -}) thymocytes and was detected at all subsequent stages, but exceeded the expression levels of the endogenous gene in the thymus. In conclusion, the analyzed perforin 5{prime} flanking and promoter region contains important cis-acting sequences that restrict perforin expression to T cells and NK cells, and therefore provides a unique tool for manipulating T cell and/or Nk cell-mediated immune responses in transgenic mice. On the other hand, the normal control of perforin gene expression involves at least one additional negative control mechanism that was not mediated by the transgenic promoter and upstream region. This control restricts perforin gene expression in thymically developing T cells and in most resting peripheral T cells, but can be released upon T cell activation. 43 refs., 7 figs., 1 tab.

  18. Organization and expression of hair follicle genes.

    PubMed

    Rogers, G E; Powell, B C

    1993-07-01

    Several families of proteins are expressed in the growth of hair and an estimated 50-100 proteins constitute the final hair fiber. The cumbersome nomenclature for naming these different proteins has led to a proposal to modify that which is currently used for epidermal keratins. Investigations of the organization of hair genes indicate that the members of each family are clustered in the genome and their expression could be under some general control. Interestingly, the protein called trichohyalin, markedly distinct from the hair proteins, is produced in the inner root sheath cells and the gene for it has been found to be located at the same human chromosome locus as the genes for profilaggrin, involucrin, and loricrin. A mainstream objective is to identify controls responsible for the production in the hair cortex of keratin intermediate filaments (IFs) and two large groups of keratin-associated proteins (KAPs) rich in the amino acids cysteine or glycine/tyrosine. A specific family of cysteine-rich proteins is expressed in the hair cuticle. Comparisons of promoter regions of IF genes and KAP genes, including a recently characterized gene for a glycine/tyrosine-rich protein, have revealed putative hair-specific motifs in addition to known elements that regulate gene expression. In the sheep, the patterns of expression in hair differentiation are particularly interesting insofar as there are distinct segments of para- and orthocortical type cells that have significantly different pathways of expression. The testing of candidate hair-specific regulatory sequences by mouse transgenesis has produced several interesting hair phenotypes. Transgenic sheep over-expressing keratin genes but showing no hair growth change have been obtained and compared with the equivalent transgenic hair-loss mice. Studies of the effects of amino acid supply on the rate of hair growth have demonstrated that with cysteine supplementation of sheep a perturbation occurs in which there is a

  19. Regulation of Calreticulin Gene Expression by Calcium

    PubMed Central

    Waser, Mathilde; Mesaeli, Nasrin; Spencer, Charlotte; Michalak, Marek

    1997-01-01

    We have isolated and characterized a 12-kb mouse genomic DNA fragment containing the entire calreticulin gene and 2.14 kb of the promoter region. The mouse calreticulin gene consists of nine exons and eight introns, and it spans 4.2 kb of genomic DNA. A 1.8-kb fragment of the calreticulin promoter was subcloned into a reporter gene plasmid containing chloramphenicol acetyltransferase. This construct was then used in transient and stable transfection of NIH/ 3T3 cells. Treatment of transfected cells either with the Ca2+ ionophore A23187, or with the ER Ca2+-ATPase inhibitor thapsigargin, resulted in a five- to sevenfold increase of the expression of chloramphenicol acetyltransferase protein. Transactivation of the calreticulin promoter was also increased by fourfold in NIH/3T3 cells treated with bradykinin, a hormone that induces Ca2+ release from the intracellular Ca2+ stores. Analysis of the promoter deletion constructs revealed that A23187- and thapsigargin-responsive regions are confined to two regions (−115 to −260 and −685 to −1,763) in the calreticulin promoter that contain the CCAAT nucleotide sequences. Northern blot analysis of cells treated with A23187, or with thapsigargin, revealed a fivefold increase in calreticulin mRNA levels. Thapsigargin also induced a fourfold increase in calreticulun protein levels. Importantly, we show by nuclear run-on transcription analysis that calreticulin gene transcription is increased in NIH/3T3 cells treated with A23187 and thapsigargin in vivo. This increase in gene expression required over 4 h of continuous incubation with the drugs and was also sensitive to treatment with cycloheximide, suggesting that it is dependent on protein synthesis. Changes in the concentration of extracellular and cytoplasmic Ca2+ did not affect the increased expression of the calreticulin gene. These studies suggest that stress response to the depletion of intracellular Ca2+ stores induces expression of the calreticulin gene in vitro

  20. ATR controls cellular adaptation to hypoxia through positive regulation of hypoxia-inducible factor 1 (HIF-1) expression.

    PubMed

    Fallone, F; Britton, S; Nieto, L; Salles, B; Muller, C

    2013-09-12

    Tumor cells adaptation to severe oxygen deprivation (hypoxia) plays a major role in tumor progression. The transcription factor HIF-1 (hypoxia-inducible factor 1), whose α-subunit is stabilized under hypoxic conditions is a key component of this process. Recent studies showed that two members of the phosphoinositide 3-kinase-related kinases (PIKKs) family, ATM (ataxia telangiectasia mutated) and DNA-PK (DNA-dependent protein kinase), regulate the hypoxic-dependent accumulation of HIF-1. These proteins initiate cellular stress responses when DNA damage occurs. In addition, it has been demonstrated that extreme hypoxia induces a replicative stress resulting in regions of single-stranded DNA at stalled replication forks and the activation of ATR (ataxia telangiectasia and Rad3 related protein), another member of the PIKKs family. Here, we show that even less severe hypoxia (0.1% O2) also induces activation of ATR through replicative stress. Importantly, in using either transiently silenced ATR cells, cells expressing an inactive form of ATR or cells exposed to an ATR inhibitor (CGK733), we demonstrate that hypoxic ATR activation positively regulates the key transcription factor HIF-1 independently of the checkpoint kinase Chk1. We show that ATR kinase activity regulates HIF-1α at the translational level and we find that the elements necessary for the regulation of HIF-1α translation are located within the coding region of HIF-1α mRNA. Finally, by using three independent cellular models, we clearly show that the loss of ATR expression and/or kinase activity results in the decrease of HIF-1 DNA binding under hypoxia and consequently affects protein expression levels of two HIF-1 target genes, GLUT-1 and CAIX. Taken together, our data show a new function for ATR in cellular adaptation to hypoxia through regulation of HIF-1α translation. Our work offers new prospect for cancer therapy using ATR inhibitors with the potential to decrease cellular adaptation in hypoxic

  1. Visual Exploration of Three-dimensional Gene Expression Using Physical Views and Linked Abstract Views

    PubMed Central

    Weber, Gunther H.; Rübel, Oliver; Huang, Min-Yu; DePace, Angela H.; Fowlkes, Charless C.; Keränen, Soile V. E.; Luengo Hendriks, Cris L.; Hagen, Hans; Knowles, David W.; Malik, Jitendra; Biggin, Mark D.; Hamann, Bernd

    2011-01-01

    During animal development, complex patterns of gene expression provide positional information within the embryo. To better understand the underlying gene regulatory networks, the Berkeley Drosophila Transcription Network Project (BDTNP) has developed methods that support quantitative computational analysis of three-dimensional (3D) gene expression in early Drosophila embryos at cellular resolution. We introduce PointCloudXplore, an interactive visualization tool that supports visual exploration of relationships between different genes’ expression using a combination of established visualization techniques. Two aspects of gene expression are of particular interest: (i) gene expression patterns defined by the spatial locations of cells expressing a gene, and (ii) relationships between the expression levels of multiple genes. PointCloudXplore provides users with two corresponding classes of data views: (i) Physical Views based on the spatial relationships of cells in the embryo, and (ii) Abstract Views that discard spatial information and plot expression levels of multiple genes with respect to each other. Cell Selectors highlight data associated with subsets of embryo cells within a View. Using linking, these selected cells can be viewed in multiple representations. We describe PCX as a 3D gene expression visualization tool and provide examples of how it has been used by BDTNP biologists to generate new hypotheses. PMID:19407353

  2. Neuron type-specific miRNA represses two broadly expressed genes to modulate an avoidance behavior in C. elegans

    PubMed Central

    Drexel, Tanja; Mahofsky, Katharina; Latham, Richard; Zimmer, Manuel

    2016-01-01

    Two broad gene classes are distinguished within multicellular organisms: cell type-specific genes, which confer particular cellular properties, and ubiquitous genes that support general cellular functions. However, certain so-called ubiquitous genes show functionally relevant cell type-specific repression. How such repression is achieved is poorly understood. MicroRNAs (miRNAs) are repressors, many of which are expressed with high cell type specificity. Here we show that mir-791, expressed exclusively in the CO2-sensing neurons in Caenorhabditis elegans, represses two otherwise broadly expressed genes. This repression is necessary for normal neuronal function and behavior of the animals toward CO2. miRNA-mediated repression of broadly transcribed genes is a previously unappreciated strategy for cellular specialization. PMID:27688400

  3. The frustrated gene: origins of eukaryotic gene expression

    PubMed Central

    Madhani, Hiten D.

    2014-01-01

    Eukarytotic gene expression is frustrated by a series of steps that are generally not observed in prokaryotes and are therefore not essential for the basic chemistry of transcription and translation. Their evolution may have been driven by the need to defend against parasitic nucleic acids. PMID:24209615

  4. Reconstructing differentially co-expressed gene modules and regulatory networks of soybean cells

    PubMed Central

    2012-01-01

    Background Current experimental evidence indicates that functionally related genes show coordinated expression in order to perform their cellular functions. In this way, the cell transcriptional machinery can respond optimally to internal or external stimuli. This provides a research opportunity to identify and study co-expressed gene modules whose transcription is controlled by shared gene regulatory networks. Results We developed and integrated a set of computational methods of differential gene expression analysis, gene clustering, gene network inference, gene function prediction, and DNA motif identification to automatically identify differentially co-expressed gene modules, reconstruct their regulatory networks, and validate their correctness. We tested the methods using microarray data derived from soybean cells grown under various stress conditions. Our methods were able to identify 42 coherent gene modules within which average gene expression correlation coefficients are greater than 0.8 and reconstruct their putative regulatory networks. A total of 32 modules and their regulatory networks were further validated by the coherence of predicted gene functions and the consistency of putative transcription factor binding motifs. Approximately half of the 32 modules were partially supported by the literature, which demonstrates that the bioinformatic methods used can help elucidate the molecular responses of soybean cells upon various environmental stresses. Conclusions The bioinformatics methods and genome-wide data sources for gene expression, clustering, regulation, and function analysis were integrated seamlessly into one modular protocol to systematically analyze and infer modules and networks from only differential expression genes in soybean cells grown under stress conditions. Our approach appears to effectively reduce the complexity of the problem, and is sufficiently robust and accurate to generate a rather complete and detailed view of putative soybean

  5. Trigger finger, tendinosis, and intratendinous gene expression.

    PubMed

    Lundin, A-C; Aspenberg, P; Eliasson, P

    2014-04-01

    The pathogenesis of trigger finger has generally been ascribed to primary changes in the first annular ligament. In contrast, we recently found histological changes in the tendons, similar to the findings in Achilles tendinosis or tendinopathy. We therefore hypothesized that trigger finger tendons would show differences in gene expression in comparison to normal tendons in a pattern similar to what is published for Achilles tendinosis. We performed quantitative real-time polymerase chain reaction on biopsies from finger flexor tendons, 13 trigger fingers and 13 apparently healthy control tendons, to assess the expression of 10 genes which have been described to be differently expressed in tendinosis (collagen type 1a1, collagen 3a1, MMP-2, MMP-3, ADAMTS-5, TIMP-3, aggrecan, biglycan, decorin, and versican). In trigger finger tendons, collagen types 1a1 and 3a1, aggrecan and biglycan were all up-regulated, and MMP-3and TIMP-3 were down-regulated. These changes were statistically significant and have been previously described for Achilles tendinosis. The remaining four genes were not significantly altered. The changes in gene expression support the hypothesis that trigger finger is a form of tendinosis. Because trigger finger is a common condition, often treated surgically, it could provide opportunities for clinical research on tendinosis.

  6. The low noise limit in gene expression

    SciTech Connect

    Dar, Roy D.; Weinberger, Leor S.; Cox, Chris D.; Simpson, Michael L.; Razooky, Brandon S.

    2015-10-21

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

  7. The Low Noise Limit in Gene Expression

    PubMed Central

    Dar, Roy D.; Razooky, Brandon S.; Weinberger, Leor S.; Cox, Chris D.; Simpson, Michael L.

    2015-01-01

    Protein noise measurements are increasingly used to elucidate biophysical parameters. Unfortunately noise analyses are often at odds with directly measured parameters. Here we show that these inconsistencies arise from two problematic analytical choices: (i) the assumption that protein translation rate is invariant for different proteins of different abundances, which has inadvertently led to (ii) the assumption that a large constitutive extrinsic noise sets the low noise limit in gene expression. While growing evidence suggests that transcriptional bursting may set the low noise limit, variability in translational bursting has been largely ignored. We show that genome-wide systematic variation in translational efficiency can–and in the case of E. coli does–control the low noise limit in gene expression. Therefore constitutive extrinsic noise is small and only plays a role in the absence of a systematic variation in translational efficiency. These results show the existence of two distinct expression noise patterns: (1) a global noise floor uniformly imposed on all genes by expression bursting; and (2) high noise distributed to only a select group of genes. PMID:26488303

  8. Digital gene expression signatures for maize development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genome-wide expression signatures detect specific perturbations in developmental programs and contribute to functional resolution of key regulatory networks. In maize (Zea mays) inflorescences, mutations in the RAMOSA (RA) genes affect determinacy of axillary meristems and thus alter branching patt...

  9. Analysis of baseline gene expression levels from ...

    EPA Pesticide Factsheets

    The use of gene expression profiling to predict chemical mode of action would be enhanced by better characterization of variance due to individual, environmental, and technical factors. Meta-analysis of microarray data from untreated or vehicle-treated animals within the control arm of toxicogenomics studies has yielded useful information on baseline fluctuations in gene expression. A dataset of control animal microarray expression data was assembled by a working group of the Health and Environmental Sciences Institute's Technical Committee on the Application of Genomics in Mechanism Based Risk Assessment in order to provide a public resource for assessments of variability in baseline gene expression. Data from over 500 Affymetrix microarrays from control rat liver and kidney were collected from 16 different institutions. Thirty-five biological and technical factors were obtained for each animal, describing a wide range of study characteristics, and a subset were evaluated in detail for their contribution to total variability using multivariate statistical and graphical techniques. The study factors that emerged as key sources of variability included gender, organ section, strain, and fasting state. These and other study factors were identified as key descriptors that should be included in the minimal information about a toxicogenomics study needed for interpretation of results by an independent source. Genes that are the most and least variable, gender-selectiv

  10. Multiple Stochastic Point Processes in Gene Expression

    NASA Astrophysics Data System (ADS)

    Murugan, Rajamanickam

    2008-04-01

    We generalize the idea of multiple-stochasticity in chemical reaction systems to gene expression. Using Chemical Langevin Equation approach we investigate how this multiple-stochasticity can influence the overall molecular number fluctuations. We show that the main sources of this multiple-stochasticity in gene expression could be the randomness in transcription and translation initiation times which in turn originates from the underlying bio-macromolecular recognition processes such as the site-specific DNA-protein interactions and therefore can be internally regulated by the supra-molecular structural factors such as the condensation/super-coiling of DNA. Our theory predicts that (1) in case of gene expression system, the variances ( φ) introduced by the randomness in transcription and translation initiation-times approximately scales with the degree of condensation ( s) of DNA or mRNA as φ ∝ s -6. From the theoretical analysis of the Fano factor as well as coefficient of variation associated with the protein number fluctuations we predict that (2) unlike the singly-stochastic case where the Fano factor has been shown to be a monotonous function of translation rate, in case of multiple-stochastic gene expression the Fano factor is a turn over function with a definite minimum. This in turn suggests that the multiple-stochastic processes can also be well tuned to behave like a singly-stochastic point processes by adjusting the rate parameters.

  11. The low noise limit in gene expression

    DOE PAGES

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

    2015-10-21

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

  12. CHO gene expression profiling in biopharmaceutical process analysis and design.

    PubMed

    Schaub, Jochen; Clemens, Christoph; Schorn, Peter; Hildebrandt, Tobias; Rust, Werner; Mennerich, Detlev; Kaufmann, Hitto; Schulz, Torsten W

    2010-02-01

    Increase in both productivity and product yields in biopharmaceutical process development with recombinant protein producing mammalian cells can be mainly attributed to the advancements in cell line development, media, and process optimization. Only recently, genome-scale technologies enable a system-level analysis to elucidate the complex biomolecular basis of protein production in mammalian cells promising an increased process understanding and the deduction of knowledge-based approaches for further process optimization. Here, the use of gene expression profiling for the analysis of a low titer (LT) and high titer (HT) fed batch process using the same IgG producing CHO cell line was investigated. We found that gene expression (i) significantly differed in HT versus LT process conditions due to differences in applied chemically defined, serum-free media, (ii) changed over the time course of the fed batch processes, and that (iii) both metabolic pathways and 14 biological functions such as cellular growth or cell death were affected. Furthermore, detailed analysis of metabolism in a standard process format revealed the potential use of transcriptomics for rational media design as is shown for the case of lipid metabolism where the product titer could be increased by about 20% based on a lipid modified basal medium. The results demonstrate that gene expression profiling can be an important tool for mammalian biopharmaceutical process analysis and optimization.

  13. Expression of mouse metallothionein genes in tobacco

    SciTech Connect

    Maiti, I.B.; Yeargan, R.; Wagner, G.J.; Hunt, A.G. )

    1990-05-01

    We have expressed a mouse metallothionein (NT) gene in tobacco under control of the cauliflower mosaic virus (CaMV) 35S promoter and a pea ribulose-1,5-bisphosphate carboxylase small subunit (rbcS) gene promoter. Seedlings in which MT gene expression is driven by the 35S promoter are resistant to toxic levels of cadmium. Mature plants carrying the 35S-MT gene accumulate less Cd in their leaves when exposed to low levels of Cd in laboratory growth conditions. Plants with the rbcS-MT construction express this gene in a light-regulated and tissue-specific manner, as expected. Moreover, the MT levels in leaves in these plants are about 20% of those seen in 35S-MT plants. These plants are currently being tested for Cd resistance. In addition, a small field evaluation of 35S-MT lines for Cd levels is being evaluated. These experiments will address the possibility of using MTs to alter Cd levels in crop species.

  14. Aging elevates metabolic gene expression in brain cholinergic neurons.

    PubMed

    Baskerville, Karen A; Kent, Caroline; Personett, David; Lai, Weil R; Park, Peter J; Coleman, Paul; McKinney, Michael

    2008-12-01

    The basal forebrain (BF) cholinergic system is selectively vulnerable in human brain diseases, while the cholinergic groups in the upper pons of the brainstem (BS) resist neurodegeneration. Cholinergic neurons (200 per region per animal) were laser-microdissected from five young (8 months) and five aged (24 months) F344 rats from the BF and the BS pontine lateral dorsal tegmental/pedunculopontine nuclei (LDTN/PPN) and their expression profiles were obtained. The bioinformatics program SigPathway was used to identify gene groups and pathways that were selectively affected by aging. In the BF cholinergic system, aging most significantly altered genes involved with a variety of metabolic functions. In contrast, BS cholinergic neuronal age effects included gene groupings related to neuronal plasticity and a broad range of normal cellular functions. Transcription factor GA-binding protein alpha (GABPalpha), which controls expression of nuclear genes encoding mitochondrial proteins, was more strongly upregulated in the BF cholinergic neurons (+107%) than in the BS cholinergic population (+40%). The results suggest that aging elicits elevates metabolic activity in cholinergic populations and that this occurs to a much greater degree in the BF group than in the BS group.

  15. Regulation of methane genes and genome expression

    SciTech Connect

    John N. Reeve

    2009-09-09

    At the start of this project, it was known that methanogens were Archaeabacteria (now Archaea) and were therefore predicted to have gene expression and regulatory systems different from Bacteria, but few of the molecular biology details were established. The goals were then to establish the structures and organizations of genes in methanogens, and to develop the genetic technologies needed to investigate and dissect methanogen gene expression and regulation in vivo. By cloning and sequencing, we established the gene and operon structures of all of the “methane” genes that encode the enzymes that catalyze methane biosynthesis from carbon dioxide and hydrogen. This work identified unique sequences in the methane gene that we designated mcrA, that encodes the largest subunit of methyl-coenzyme M reductase, that could be used to identify methanogen DNA and establish methanogen phylogenetic relationships. McrA sequences are now the accepted standard and used extensively as hybridization probes to identify and quantify methanogens in environmental research. With the methane genes in hand, we used northern blot and then later whole-genome microarray hybridization analyses to establish how growth phase and substrate availability regulated methane gene expression in Methanobacterium thermautotrophicus ΔH (now Methanothermobacter thermautotrophicus). Isoenzymes or pairs of functionally equivalent enzymes catalyze several steps in the hydrogen-dependent reduction of carbon dioxide to methane. We established that hydrogen availability determine which of these pairs of methane genes is expressed and therefore which of the alternative enzymes is employed to catalyze methane biosynthesis under different environmental conditions. As were unable to establish a reliable genetic system for M. thermautotrophicus, we developed in vitro transcription as an alternative system to investigate methanogen gene expression and regulation. This led to the discovery that an archaeal protein

  16. Fluid Mechanics, Arterial Disease, and Gene Expression

    PubMed Central

    Tarbell, John M.; Shi, Zhong-Dong; Dunn, Jessilyn; Jo, Hanjoong

    2014-01-01

    This review places modern research developments in vascular mechanobiology in the context of hemodynamic phenomena in the cardiovascular system and the discrete localization of vascular disease. The modern origins of this field are traced, beginning in the 1960s when associations between flow characteristics, particularly blood flow–induced wall shear stress, and the localization of atherosclerotic plaques were uncovered, and continuing to fluid shear stress effects on the vascular lining endothelial) cells (ECs), including their effects on EC morphology, biochemical production, and gene expression. The earliest single-gene studies and genome-wide analyses are considered. The final section moves from the ECs lining the vessel wall to the smooth muscle cells and fibroblasts within the wall that are fluid me chanically activated by interstitial flow that imposes shear stresses on their surfaces comparable with those of flowing blood on EC surfaces. Interstitial flow stimulates biochemical production and gene expression, much like blood flow on ECs. PMID:25360054

  17. Fluid Mechanics, Arterial Disease, and Gene Expression.

    PubMed

    Tarbell, John M; Shi, Zhong-Dong; Dunn, Jessilyn; Jo, Hanjoong

    2014-01-01

    This review places modern research developments in vascular mechanobiology in the context of hemodynamic phenomena in the cardiovascular system and the discrete localization of vascular disease. The modern origins of this field are traced, beginning in the 1960s when associations between flow characteristics, particularly blood flow-induced wall shear stress, and the localization of atherosclerotic plaques were uncovered, and continuing to fluid shear stress effects on the vascular lining endothelial) cells (ECs), including their effects on EC morphology, biochemical production, and gene expression. The earliest single-gene studies and genome-wide analyses are considered. The final section moves from the ECs lining the vessel wall to the smooth muscle cells and fibroblasts within the wall that are fluid me chanically activated by interstitial flow that imposes shear stresses on their surfaces comparable with those of flowing blood on EC surfaces. Interstitial flow stimulates biochemical production and gene expression, much like blood flow on ECs.

  18. Gene expression profiling of human ovarian tumours

    PubMed Central

    Biade, S; Marinucci, M; Schick, J; Roberts, D; Workman, G; Sage, E H; O'Dwyer, P J; LiVolsi, V A; Johnson, S W

    2006-01-01

    There is currently a lack of reliable diagnostic and prognostic markers for ovarian cancer. We established gene expression profiles for 120 human ovarian tumours to identify determinants of histologic subtype, grade and degree of malignancy. Unsupervised cluster analysis of the most variable set of expression data resulted in three major tumour groups. One consisted predominantly of benign tumours, one contained mostly malignant tumours, and one was comprised of a mixture of borderline and malignant tumours. Using two supervised approaches, we identified a set of genes that distinguished the benign, borderline and malignant phenotypes. These algorithms were unable to establish profiles for histologic subtype or grade. To validate these findings, the expression of 21 candidate genes selected from these analyses was measured by quantitative RT–PCR using an independent set of tumour samples. Hierarchical clustering of these data resulted in two major groups, one benign and one malignant, with the borderline tumours interspersed between the two groups. These results indicate that borderline ovarian tumours may be classified as either benign or malignant, and that this classifier could be useful for predicting the clinical course of borderline tumours. Immunohistochemical analysis also demonstrated increased expression of CD24 antigen in malignant versus benign tumour tissue. The data that we have generated will contribute to a growing body of expression data that more accurately define the biologic and clinical characteristics of ovarian cancers. PMID:16969345

  19. Gene expression profiling of human ovarian tumours.

    PubMed

    Biade, S; Marinucci, M; Schick, J; Roberts, D; Workman, G; Sage, E H; O'Dwyer, P J; Livolsi, V A; Johnson, S W

    2006-10-23

    There is currently a lack of reliable diagnostic and prognostic markers for ovarian cancer. We established gene expression profiles for 120 human ovarian tumours to identify determinants of histologic subtype, grade and degree of malignancy. Unsupervised cluster analysis of the most variable set of expression data resulted in three major tumour groups. One consisted predominantly of benign tumours, one contained mostly malignant tumours, and one was comprised of a mixture of borderline and malignant tumours. Using two supervised approaches, we identified a set of genes that distinguished the benign, borderline and malignant phenotypes. These algorithms were unable to establish profiles for histologic subtype or grade. To validate these findings, the expression of 21 candidate genes selected from these analyses was measured by quantitative RT-PCR using an independent set of tumour samples. Hierarchical clustering of these data resulted in two major groups, one benign and one malignant, with the borderline tumours interspersed between the two groups. These results indicate that borderline ovarian tumours may be classified as either benign or malignant, and that this classifier could be useful for predicting the clinical course of borderline tumours. Immunohistochemical analysis also demonstrated increased expression of CD24 antigen in malignant versus benign tumour tissue. The data that we have generated will contribute to a growing body of expression data that more accurately define the biologic and clinical characteristics of ovarian cancers.

  20. Dysregulation of host cellular genes targeted by human papillomavirus (HPV) integration contributes to HPV-related cervical carcinogenesis.

    PubMed

    Zhang, Ruiyang; Shen, Congle; Zhao, Lijun; Wang, Jianliu; McCrae, Malcolm; Chen, Xiangmei; Lu, Fengmin

    2016-03-01

    Integration of human papillomavirus (HPV) viral DNA into the human genome has been postulated as an important etiological event during cervical carcinogenesis. Several recent reports suggested a possible role for such integration-targeted cellular genes (ITGs) in cervical carcinogenesis. Therefore, a comprehensive analysis of HPV integration events was undertaken using data collected from 14 publications, with 499 integration loci on human chromosomes included. It revealed that HPV DNA preferred to integrate into intragenic regions and gene-dense regions of human chromosomes. Intriguingly, the host cellular genes nearby the integration sites were found to be more transcriptionally active compared with control. Furthermore, analysis of the integration sites in the human genome revealed that there were several integration hotspots although all chromosomes were represented. The ITGs identified were found to be enriched in tumor-related terms and pathways using gene ontology and KEGG analysis. In line with this, three of six ITGs tested were found aberrantly expressed in cervical cancer tissues. Among them, it was demonstrated for the first time that MPPED2 could induce HeLa cell and SiHa cell G1/S transition block and cell proliferation retardation. Moreover, "knocking out" the integrated HPV fragment in HeLa cell line decreased expression of MYC located ∼500 kb downstream of the integration site, which provided the first experimental evidence supporting the hypothesis that integrated HPV fragment influence MYC expression via long distance chromatin interaction. Overall, the results of this comprehensive analysis implicated that dysregulation of ITGs caused by viral integration as possibly having an etiological involvement in cervical carcinogenesis.

  1. Modulation of enhancer looping and differential gene targeting by Epstein-Barr virus transcription factors directs cellular reprogramming.

    PubMed

    McClellan, Michael J; Wood, C David; Ojeniyi, Opeoluwa; Cooper, Tim J; Kanhere, Aditi; Arvey, Aaron; Webb, Helen M; Palermo, Richard D; Harth-Hertle, Marie L; Kempkes, Bettina; Jenner, Richard G; West, Michelle J

    2013-09-01

    Epstein-Barr virus (EBV) epigenetically reprogrammes B-lymphocytes to drive immortalization and facilitate viral persistence. Host-cell transcription is perturbed principally through the actions of EBV EBNA 2, 3A, 3B and 3C, with cellular genes deregulated by specific combinations of these EBNAs through unknown mechanisms. Comparing human genome binding by these viral transcription factors, we discovered that 25% of binding sites were shared by EBNA 2 and the EBNA 3s and were located predominantly in enhancers. Moreover, 80% of potential EBNA 3A, 3B or 3C target genes were also targeted by EBNA 2, implicating extensive interplay between EBNA 2 and 3 proteins in cellular reprogramming. Investigating shared enhancer sites neighbouring two new targets (WEE1 and CTBP2) we discovered that EBNA 3 proteins repress transcription by modulating enhancer-promoter loop formation to establish repressive chromatin hubs or prevent assembly of active hubs. Re-ChIP analysis revealed that EBNA 2 and 3 proteins do not bind simultaneously at shared sites but compete for binding thereby modulating enhancer-promoter interactions. At an EBNA 3-only intergenic enhancer site between ADAM28 and ADAMDEC1 EBNA 3C was also able to independently direct epigenetic repression of both genes through enhancer-promoter looping. Significantly, studying shared or unique EBNA 3 binding sites at WEE1, CTBP2, ITGAL (LFA-1 alpha chain), BCL2L11 (Bim) and the ADAMs, we also discovered that different sets of EBNA 3 proteins bind regulatory elements in a gene and cell-type specific manner. Binding profiles correlated with the effects of individual EBNA 3 proteins on the expression of these genes, providing a molecular basis for the targeting of different sets of cellular genes by the EBNA 3s. Our results therefore highlight the influence of the genomic and cellular context in determining the specificity of gene deregulation by EBV and provide a paradigm for host-cell reprogramming through modulation of

  2. [Structure and expression of thyroglobulin gene].

    PubMed

    Vassart, G; Brocas, H; Christophe, D; de Martynoff, G; Leriche, A; Mercken, L; Pohl, V; Van Heuverswyn, B

    1982-01-01

    Thyroglobulin is composed of two 300000 dalton polypeptide chains, translated from an 8000 base mRNA. Preparation of a full length cDNA and its cloning in E. coli have lead to the demonstration that the polypeptides of thyroglobulin protomers were identical. Used as molecular probes, the cloned cDNA allowed the isolation of a fragment of thyroglobulin gene. Electron microscopic studies have demonstrated that this gene contains more than 90% intronic material separating small size exons (less than 200 bp). Sequencing of bovine thyroglobulin structural gene is in progress. Preliminary results show evidence for the existence of repetitive segments. Availability of cloned DNA complementary to bovine and human thyroglobulin mRNA allows the study of genetic defects of thyroglobulin gene expression in the human and in various animal models.

  3. Determining Physical Mechanisms of Gene Expression Regulation from Single Cell Gene Expression Data

    PubMed Central

    Moignard, Victoria; Göttgens, Berthold; Adryan, Boris

    2016-01-01

    Many genes are expressed in bursts, which can contribute to cell-to-cell heterogeneity. It is now possible to measure this heterogeneity with high throughput single cell gene expression assays (single cell qPCR and RNA-seq). These experimental approaches generate gene expression distributions which can be used to estimate the kinetic parameters of gene expression bursting, namely the rate that genes turn on, the rate that genes turn off, and the rate of transcription. We construct a complete pipeline for the analysis of single cell qPCR data that uses the mathematics behind bursty expression to develop more accurate and robust algorithms for analyzing the origin of heterogeneity in experimental samples, specifically an algorithm for clustering cells by their bursting behavior (Simulated Annealing for Bursty Expression Clustering, SABEC) and a statistical tool for comparing the kinetic parameters of bursty expression across populations of cells (Estimation of Parameter changes in Kinetics, EPiK). We applied these methods to hematopoiesis, including a new single cell dataset in which transcription factors (TFs) involved in the earliest branchpoint of blood differentiation were individually up- and down-regulated. We could identify two unique sub-populations within a seemingly homogenous group of hematopoietic stem cells. In addition, we could predict regulatory mechanisms controlling the expression levels of eighteen key hematopoietic transcription factors throughout differentiation. Detailed information about gene regulatory mechanisms can therefore be obtained simply from high throughput single cell gene expression data, which should be widely applicable given the rapid expansion of single cell genomics. PMID:27551778

  4. Coevolution of gene expression among interacting proteins

    SciTech Connect

    Fraser, Hunter B.; Hirsh, Aaron E.; Wall, Dennis P.; Eisen,Michael B.

    2004-03-01

    Physically interacting proteins or parts of proteins are expected to evolve in a coordinated manner that preserves proper interactions. Such coevolution at the amino acid-sequence level is well documented and has been used to predict interacting proteins, domains, and amino acids. Interacting proteins are also often precisely coexpressed with one another, presumably to maintain proper stoichiometry among interacting components. Here, we show that the expression levels of physically interacting proteins coevolve. We estimate average expression levels of genes from four closely related fungi of the genus Saccharomyces using the codon adaptation index and show that expression levels of interacting proteins exhibit coordinated changes in these different species. We find that this coevolution of expression is a more powerful predictor of physical interaction than is coevolution of amino acid sequence. These results demonstrate previously uncharacterized coevolution of gene expression, adding a different dimension to the study of the coevolution of interacting proteins and underscoring the importance of maintaining coexpression of interacting proteins over evolutionary time. Our results also suggest that expression coevolution can be used for computational prediction of protein protein interactions.

  5. Differential var gene expression in children with malaria and antidromic effects on host gene expression.

    PubMed

    Kalmbach, Yvonne; Rottmann, Matthias; Kombila, Maryvonne; Kremsner, Peter G; Beck, Hans-Peter; Kun, Jürgen F J

    2010-07-15

    Among 62 children with mild malaria, cerebral malaria, or severe malarial anemia, we analyzed the transcription of different var gene types. There was no difference in parasitemia level or body temperature between groups. However, a significantly different expression pattern was observed in children with cerebral malaria, compared with that in patients in the other 2 groups: children with cerebral malaria had lower expression of the upsA subtype but higher expression of the upsB and upsC subtypes. Furthermore, expression of human genes responsive to tumor necrosis factor and hypoxia correlated with distinct ups types.

  6. Age-related changes in the expression of schizophrenia susceptibility genes in the human prefrontal cortex.

    PubMed

    Colantuoni, Carlo; Hyde, Thomas M; Mitkus, Shruti; Joseph, Andrew; Sartorius, Leah; Aguirre, Claudia; Creswell, Johanna; Johnson, Elizabeth; Deep-Soboslay, Amy; Herman, Mary M; Lipska, Barbara K; Weinberger, Daniel R; Kleinman, Joel E

    2008-09-01

    The molecular basis of complex neuropsychiatric disorders most likely involves many genes. In recent years, specific genetic variations influencing risk for schizophrenia and other neuropsychiatric disorders have been reported. We have used custom DNA microarrays and qPCR to investigate the expression of putative schizophrenia susceptibility genes and related genes of interest in the normal human brain. Expression of 31 genes was measured in Brodmann's area 10 (BA10) in the prefrontal cortex of 72 postmortem brain samples spanning half a century of human aging (18-67 years), each without history of neuropsychiatric illness, neurological disease, or drug abuse. Examination of expression across age allowed the identification of genes whose expression patterns correlate with age, as well as genes that share common expression patterns and that possibly participate in common cellular mechanisms related to the emergence of schizophrenia in early adult life. The expression of GRM3 and RGS4 decreased across the entire age range surveyed, while that of PRODH and DARPP-32 was shown to increase with age. NRG1, ERBB3, and NGFR show expression changes during the years of greatest risk for the development of schizophrenia. Expression of FEZ1, GAD1, and RGS4 showed especially high correlation with one another, in addition to the strongest mean levels of absolute correlation with all other genes studied here. All microarray data are available at NCBI's Gene Expression Omnibus: GEO Series accession number GSE11546 (http://www.ncbi.nlm.nih.gov/geo) [corrected

  7. Transcriptome-Level Signatures in Gene Expression and Gene Expression Variability during Bacterial Adaptive Evolution

    PubMed Central

    Erickson, Keesha E.; Otoupal, Peter B.

    2017-01-01

    ABSTRACT Antibiotic-resistant bacteria are an increasingly serious public health concern, as strains emerge that demonstrate resistance to almost all available treatments. One factor that contributes to the crisis is the adaptive ability of bacteria, which exhibit remarkable phenotypic and gene expression heterogeneity in order to gain a survival advantage in damaging environments. This high degree of variability in gene expression across biological populations makes it a challenging task to identify key regulators of bacterial adaptation. Here, we research the regulation of adaptive resistance by investigating transcriptome profiles of Escherichia coli upon adaptation to disparate toxins, including antibiotics and biofuels. We locate potential target genes via conventional gene expression analysis as well as using a new analysis technique examining differential gene expression variability. By investigating trends across the diverse adaptation conditions, we identify a focused set of genes with conserved behavior, including those involved in cell motility, metabolism, membrane structure, and transport, and several genes of unknown function. To validate the biological relevance of the observed changes, we synthetically perturb gene expression using clustered regularly interspaced short palindromic repeat (CRISPR)-dCas9. Manipulation of select genes in combination with antibiotic treatment promotes adaptive resistance as demonstrated by an increased degree of antibiotic tolerance and heterogeneity in MICs. We study the mechanisms by which identified genes influence adaptation and find that select differentially variable genes have the potential to impact metabolic rates, mutation rates, and motility. Overall, this work provides evidence for a complex nongenetic response, encompassing shifts in gene expression and gene expression variability, which underlies adaptive resistance. IMPORTANCE Even initially sensitive bacteria can rapidly thwart antibiotic treatment

  8. Transcriptome-Level Signatures in Gene Expression and Gene Expression Variability during Bacterial Adaptive Evolution.

    PubMed

    Erickson, Keesha E; Otoupal, Peter B; Chatterjee, Anushree

    2017-01-01

    Antibiotic-resistant bacteria are an increasingly serious public health concern, as strains emerge that demonstrate resistance to almost all available treatments. One factor that contributes to the crisis is the adaptive ability of bacteria, which exhibit remarkable phenotypic and gene expression heterogeneity in order to gain a survival advantage in damaging environments. This high degree of variability in gene expression across biological populations makes it a challenging task to identify key regulators of bacterial adaptation. Here, we research the regulation of adaptive resistance by investigating transcriptome profiles of Escherichia coli upon adaptation to disparate toxins, including antibiotics and biofuels. We locate potential target genes via conventional gene expression analysis as well as using a new analysis technique examining differential gene expression variability. By investigating trends across the diverse adaptation conditions, we identify a focused set of genes with conserved behavior, including those involved in cell motility, metabolism, membrane structure, and transport, and several genes of unknown function. To validate the biological relevance of the observed changes, we synthetically perturb gene expression using clustered regularly interspaced short palindromic repeat (CRISPR)-dCas9. Manipulation of select genes in combination with antibiotic treatment promotes adaptive resistance as demonstrated by an increased degree of antibiotic tolerance and heterogeneity in MICs. We study the mechanisms by which identified genes influence adaptation and find that select differentially variable genes have the potential to impact metabolic rates, mutation rates, and motility. Overall, this work provides evidence for a complex nongenetic response, encompassing shifts in gene expression and gene expression variability, which underlies adaptive resistance. IMPORTANCE Even initially sensitive bacteria can rapidly thwart antibiotic treatment through stress

  9. Nutrient sensing and insulin signaling in neuropeptide-expressing immortalized, hypothalamic neurons: A cellular model of insulin resistance.

    PubMed

    Fick, Laura J; Belsham, Denise D

    2010-08-15

    Obesity and type 2 diabetes mellitus represent a significant global health crisis. These two interrelated diseases are typified by perturbed insulin signaling in the hypothalamus. Using novel hypothalamic cell lines, we have begun to elucidate the molecular and intracellular mechanisms involved in the hypothalamic control of energy homeostasis and insulin resistance. In this review, we present evidence of insulin and glucose signaling pathways that lead to changes in neuropeptide gene expression. We have identified some of the molecular mechanisms involved in the control of de novo hypothalamic insulin mRNA expression. And finally, we have defined key mechanisms involved in the etiology of cellular insulin resistance in hypothalamic neurons that may play a fundamental role in cases of high levels of insulin or saturated fatty acids, often linked to the exacerbation of obesity and diabetes.

  10. Gene expression regulation in roots under drought.

    PubMed

    Janiak, Agnieszka; Kwaśniewski, Mirosław; Szarejko, Iwona

    2016-02-01

    Stress signalling and regulatory networks controlling expression of target genes are the basis of plant response to drought. Roots are the first organs exposed to water deficiency in the soil and are the place of drought sensing. Signalling cascades transfer chemical signals toward the shoot and initiate molecular responses that lead to the biochemical and morphological changes that allow plants to be protected against water loss and to tolerate stress conditions. Here, we present an overview of signalling network and gene expression regulation pathways that are actively induced in roots under drought stress. In particular, the role of several transcription factor (TF) families, including DREB, AP2/ERF, NAC, bZIP, MYC, CAMTA, Alfin-like and Q-type ZFP, in the regulation of root response to drought are highlighted. The information provided includes available data on mutual interactions between these TFs together with their regulation by plant hormones and other signalling molecules. The most significant downstream target genes and molecular processes that are controlled by the regulatory factors are given. These data are also coupled with information about the influence of the described regulatory networks on root traits and root development which may translate to enhanced drought tolerance. This is the first literature survey demonstrating the gene expression regulatory machinery that is induced by drought stress, presented from the perspective of roots.

  11. Expression of bacterial genes in plant cells.

    PubMed Central

    Fraley, R T; Rogers, S G; Horsch, R B; Sanders, P R; Flick, J S; Adams, S P; Bittner, M L; Brand, L A; Fink, C L; Fry, J S; Galluppi, G R; Goldberg, S B; Hoffmann, N L; Woo, S C

    1983-01-01

    Chimeric bacterial genes conferring resistance to aminoglycoside antibiotics have been inserted into the Agrobacterium tumefaciens tumor-inducing (Ti) plasmid and introduced into plant cells by in vitro transformation techniques. The chimeric genes contain the nopaline synthase 5' and 3' regulatory regions joined to the genes for neomycin phosphotransferase type I or type II. The chimeric genes were cloned into an intermediate vector, pMON120, and inserted into pTiB6S3 by recombination and then introduced into petunia and tobacco cells by cocultivating A. tumefaciens cells with protoplast-derived cells. Southern hybridization was used to confirm the presence of the chimeric genes in the transformed plant tissues. Expression of the chimeric genes was determined by the ability of the transformed cells to proliferate on medium containing normally inhibitory levels of kanamycin (50 micrograms/ml) or other aminoglycoside antibiotics. Plant cells transformed by wild-type pTiB6S3 or derivatives carrying the bacterial neomycin phosphotransferase genes with their own promoters failed to grow under these conditions. The significance of these results for plant genetic engineering is discussed. Images PMID:6308651

  12. Gene Expression of Corals in Response to Macroalgal Competitors

    PubMed Central

    Shearer, Tonya L.; Snell, Terry W.; Hay, Mark E.

    2014-01-01

    As corals decline and macroalgae proliferate on coral reefs, coral-macroalgal competition becomes more frequent and ecologically important. Whether corals are damaged by these interactions depends on susceptibility of the coral and traits of macroalgal competitors. Investigating changes in gene expression of corals and their intracellular symbiotic algae, Symbiodinium, in response to contact with different macroalgae provides insight into the biological processes and cellular pathways affected by competition with macroalgae. We evaluated the gene expression profiles of coral and Symbiodinium genes from two confamilial corals, Acropora millepora and Montipora digitata, after 6 h and 48 h of contact with four common macroalgae that differ in their allelopathic potency to corals. Contacts with macroalgae affected different biological pathways in the more susceptible (A. millepora) versus the more resistant (M. digitata) coral. Genes of coral hosts and of their associated Symbiodinium also responded in species-specific and time-specific ways to each macroalga. Changes in number and expression intensity of affected genes were greater after 6 h compared to 48 h of contact and were greater following contact with Chlorodesmis fastigiata and Amphiroa crassa than following contact with Galaxaura filamentosa or Turbinaria conoides. We documented a divergence in transcriptional responses between two confamilial corals and their associated Symbiodinium, as well as a diversity of dynamic responses within each coral species with respect to the species of macroalgal competitor and the duration of exposure to that competitor. These responses included early initiation of immune processes by Montipora, which is more resistant to damage after long-term macroalgal contact. Activation of the immune response by corals that better resist algal competition is consistent with the hypothesis that some macroalgal effects on corals may be mediated by microbial pathogens. PMID:25500576

  13. Transient gene expression in electroporated Solanum protoplasts.

    PubMed

    Jones, H; Ooms, G; Jones, M G

    1989-11-01

    Electroporation was used to evaluate parameters important in transient gene expression in potato protoplasts. The protoplasts were from leaves of wild potato Solanum brevidens, and from leaves, tubers and suspension cells of cultivated Solanum tuberosum cv. Désirée. Reporter enzyme activity, chloramphenicol acetyl transferase (CAT) under the control of the cauliflower mosaic virus (CaMV) 35S promoter, depended on the field strength and the pulse duration used for electroporation. Using field pulses of 85 ms duration, the optimum field strengths for maximum CAT activity were: S. brevidens mesophyll protoplasts--250 V/cm; Désirée mesophyll protoplasts--225 V/cm; Désirée suspension culture protoplasts--225 V/cm; and Désirée tuber protoplasts--150 V/cm. The optimum field strengths correlated inversely with the size of the protoplasts electroporated; this is consistent with biophysical theory. In time courses, maximum CAT activity (in Désirée mesophyll protoplasts) occurred 36-48 h after electroporation. Examination at optimised conditions of a chimaeric gene consisting of a class II patatin promoter linked to the beta-glucuronidase (gus) gene, showed expression (at DNA concentrations between 0-10 pmol/ml) comparable to the CaMV 35S promoter in both tuber and mesophyll protoplasts. At higher DNA concentrations (20-30 pmol/ml) the patatin promoter directed 4-5 times higher levels of gus expression. Implications and potential contributions towards studying gene expression, in particular of homologous genes in potato, are discussed.

  14. Expression profiling of hypothetical genes in Desulfovibrio vulgaris leads to improved functional annotation

    PubMed Central

    Elias, Dwayne A.; Mukhopadhyay, Aindrila; Joachimiak, Marcin P.; Drury, Elliott C.; Redding, Alyssa M.; Yen, Huei-Che B.; Fields, Matthew W.; Hazen, Terry C.; Arkin, Adam P.; Keasling, Jay D.; Wall, Judy D.

    2009-01-01

    Hypothetical (HyP) and conserved HyP genes account for >30% of sequenced bacterial genomes. For the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough, 347 of the 3634 genes were annotated as conserved HyP (9.5%) along with 887 HyP genes (24.4%). Given the large fraction of the genome, it is plausible that some of these genes serve critical cellular roles. The study goals were to determine which genes were expressed and provide a more functionally based annotation. To accomplish this, expression profiles of 1234 HyP and conserved genes were used from transcriptomic datasets of 11 environmental stresses, complemented with shotgun LC–MS/MS and AMT tag proteomic data. Genes were divided into putatively polycistronic operons and those predicted to be monocistronic, then classified by basal expression levels and grouped according to changes in expression for one or multiple stresses. One thousand two hundred and twelve of these genes were transcribed with 786 producing detectable proteins. There was no evidence for expression of 17 predicted genes. Except for the latter, monocistronic gene annotation was expanded using the above criteria along with matching Clusters of Orthologous Groups. Polycistronic genes were annotated in the same manner with inferences from their proximity to more confidently annotated genes. Two targeted deletion mutants were used as test cases to determine the relevance of the inferred functional annotations. PMID:19293273

  15. Expression profiling of hypothetical genes in Desulfovibrio vulgaris leads to improved functional annotation

    SciTech Connect

    Elias, Dwayne A.; Mukhopadhyay, Aindrila; Joachimiak, Marcin P.; Drury, Elliott C.; Redding, Alyssa M.; Yen, Huei-Che B.; Fields, Matthew W.; Hazen, Terry C.; Arkin, Adam P.; Keasling, Jay D.; Wall, Judy D.

    2008-10-27

    Hypothetical and conserved hypothetical genes account for>30percent of sequenced bacterial genomes. For the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough, 347 of the 3634 genes were annotated as conserved hypothetical (9.5percent) along with 887 hypothetical genes (24.4percent). Given the large fraction of the genome, it is plausible that some of these genes serve critical cellular roles. The study goals were to determine which genes were expressed and provide a more functionally based annotation. To accomplish this, expression profiles of 1234 hypothetical and conserved genes were used from transcriptomic datasets of 11 environmental stresses, complemented with shotgun LC-MS/MS and AMT tag proteomic data. Genes were divided into putatively polycistronic operons and those predicted to be monocistronic, then classified by basal expression levels and grouped according to changes in expression for one or multiple stresses. 1212 of these genes were transcribed with 786 producing detectable proteins. There was no evidence for expression of 17 predicted genes. Except for the latter, monocistronic gene annotation was expanded using the above criteria along with matching Clusters of Orthologous Groups. Polycistronic genes were annotated in the same manner with inferences from their proximity to more confidently annotated genes. Two targeted deletion mutants were used as test cases to determine the relevance of the inferred functional annotations.

  16. Cellular and temporal expression of NADPH oxidase (NOX) isotypes after brain injury

    PubMed Central

    2013-01-01

    Background Brain injury results in an increase in the activity of the reactive oxygen species generating NADPH oxidase (NOX) enzymes. Preliminary studies have shown that NOX2, NOX3, and NOX4 are the most prominently expressed NOX isotypes in the brain. However, the cellular and temporal expression profile of these isotypes in the injured and non-injured brain is currently unclear. Methods Double immunofluorescence for NOX isotypes and brain cell types was performed at acute (24 hours), sub-acute (7 days), and chronic (28 days) time points after controlled cortical impact-induced brain injury or sham-injury in rats. Results NOX2, NOX3, and NOX4 isotypes were found to be expressed in neurons, astrocytes, and microglia, and this expression was dependent on both cellular source and post-injury time. NOX4 was found in all cell types assessed, while NOX3 was positively identified in neurons only, and NOX2 was identified in microglia and neurons. NOX2 was the most responsive to injury, increasing primarily in microglia in response to injury. Quantitation of this isotype showed a significant increase in NOX2 expression at 24 hours, with reduced expression at 7 days and 28 days post-injury, although expression remained above sham levels at later time points. Cellular confirmation using purified primary or cell line culture demonstrated similar patterns in microglia, astrocytes, and neurons. Further, inhibition of NOX, and more specifically NOX2, reduced pro-inflammatory activity in microglia, demonstrating that NOX is not only up-regulated after stimulation, but may also play a significant role in post-injury neuroinflammation. Conclusions This study illustrates the expression profiles of NOX isotypes in the brain after injury, and demonstrates that NOX2, and to a lesser extent, NOX4, may be responsible for the majority of oxidative stress observed acutely after traumatic brain injury. These data may provide insight into the design of future therapeutic approaches. PMID

  17. Contaminant loading in remote Arctic lakes affects cellular stress-related proteins expression in feral charr.

    USGS Publications Warehouse

    Wiseman, Steve; Jorgensen, Even H.; Maule, Alec G.; Vijayan, Mathilakath M.

    2011-01-01

    The remote Arctic lakes on Bjornoya Island, Norway, offer a unique opportunity to study possible affect of lifelong contaminant exposure in wild populations of landlocked Arctic charr (Salvelinus alpinus). This is because Lake Ellasjoen has persistent organic pollutant (POP) levels that are significantly greater than in the nearby Lake Oyangen. We examined whether this differential contaminant loading was reflected in the expression of protein markers of exposure and effect in the native fish. We assessed the expressions of cellular stress markers, including cytochrome P4501A (Cyp1A), heat shock protein 70 (hsp70), and glucocorticoid receptor (GR) in feral charr from the two lakes. The average polychlorinated biphenyl (PCB) load in the charr liver from Ellasjoen was approximately 25-fold higher than in individuals from Oyangen. Liver Cyp1A protein expression was significantly higher in individuals from Ellasjoen compared with Oyangen, confirming differential PCB exposure. There was no significant difference in hsp70 protein expression in charr liver between the two lakes. However, brain hsp70 protein expression was significantly elevated in charr from Ellasjoen compared with Oyangen. Also, liver GR protein expression was significantly higher in the Ellasjoen charr compared with Oyangen charr. Taken together, our results suggest changes to cellular stress-related protein expression as a possible adaptation to chronic-contaminant exposure in feral charr in the Norwegian high-Arctic.

  18. Androgens regulate gene expression in avian skeletal muscles.

    PubMed

    Fuxjager, Matthew J; Barske, Julia; Du, Sienmi; Day, Lainy B; Schlinger, Barney A

    2012-01-01

    Circulating androgens in adult reproductively active male vertebrates influence a diversity of organ systems and thus are considered costly. Recently, we obtained evidence that androgen receptors (AR) are expressed in several skeletal muscles of three passeriform birds, the golden-collared manakin (Manacus vitellinus), zebra finch (Taenopygia guttata), and ochre-bellied flycatcher (Mionectes oleagieus). Because skeletal muscles that control wing movement make up the bulk of a bird's body mass, evidence for widespread effects of androgen action on these muscles would greatly expand the functional impact of androgens beyond their well-characterized effects on relatively discrete targets throughout the avian body. To investigate this issue, we use quantitative PCR (qPCR) to determine if androgens alter gene mRNA expression patterns in wing musculature of wild golden-collared manakins and captive zebra finches. In manakins, the androgen testosterone (T) up-regulated expression of parvalbumin (PV) and insulin-like growth factor I (IGF-I), two genes whose products enhance cellular Ca(2+) cycling and hypertrophy of skeletal muscle fibers. In T-treated zebra finches, the anti-androgen flutamide blunted PV and IGF-I expression. These results suggest that certain transcriptional effects of androgen action via AR are conserved in passerine skeletal muscle tissue. When we examined wing muscles of manakins, zebra finches and ochre-bellied flycatchers, we found that expression of PV and IGF-I varied across species and in a manner consistent with a function for AR-dependent gene regulation. Together, these findings imply that androgens have the potential to act on avian muscle in a way that may enhance the physicality required for successful reproduction.

  19. Androgens Regulate Gene Expression in Avian Skeletal Muscles

    PubMed Central

    Fuxjager, Matthew J.; Barske, Julia; Du, Sienmi; Day, Lainy B.; Schlinger, Barney A.

    2012-01-01

    Circulating androgens in adult reproductively active male vertebrates influence a diversity of organ systems and thus are considered costly. Recently, we obtained evidence that androgen receptors (AR) are expressed in several skeletal muscles of three passeriform birds, the golden-collared manakin (Manacus vitellinus), zebra finch (Taenopygia guttata), and ochre-bellied flycatcher (Mionectes oleagieus). Because skeletal muscles that control wing movement make up the bulk of a bird’s body mass, evidence for widespread effects of androgen action on these muscles would greatly expand the functional impact of androgens beyond their well-characterized effects on relatively discrete targets throughout the avian body. To investigate this issue, we use quantitative PCR (qPCR) to determine if androgens alter gene mRNA expression patterns in wing musculature of wild golden-collared manakins and captive zebra finches. In manakins, the androgen testosterone (T) up-regulated expression of parvalbumin (PV) and insulin-like growth factor I (IGF-I), two genes whose products enhance cellular Ca2+ cycling and hypertrophy of skeletal muscle fibers. In T-treated zebra finches, the anti-androgen flutamide blunted PV and IGF-I expression. These results suggest that certain transcriptional effects of androgen action via AR are conserved in passerine skeletal muscle tissue. When we examined wing muscles of manakins, zebra finches and ochre-bellied flycatchers, we found that expression of PV and IGF-I varied across species and in a manner consistent with a function for AR-dependent gene regulation. Together, these findings imply that androgens have the potential to act on avian muscle in a way that may enhance the physicality required for successful reproduction. PMID:23284699

  20. Alpha1-antitrypsin gene therapy modulates cellular immunity and efficiently prevents type 1 diabetes in nonobese diabetic mice.

    PubMed

    Lu, Yuanqing; Tang, Mei; Wasserfall, Clive; Kou, Zhongchen; Campbell-Thompson, Martha; Gardemann, Thomas; Crawford, James; Atkinson, Mark; Song, Sihong

    2006-06-01

    An imbalance of the immune-regulatory pathways plays an important role in the development of type 1 diabetes. Therefore, immunoregulatory and antiinflammatory strategies hold great potential for the prevention of this autoimmune disease. Studies have demonstrated that two serine proteinase inhibitors, alpha1-antitrypsin (AAT) and elafin, act as potent antiinflammatory agents. In the present study, we sought to develop an efficient gene therapy approach to prevent type 1 diabetes. Cohorts of 4-week-old female nonobese diabetic (NOD) mice were injected intramuscularly with rAAV1-CB-hAAT, rAAV1-CB-hElafin, or saline. AAV1 vector mediated sustained high levels of transgene expression, sufficient to overcome a humoral immune response against hAAT. AAT gene therapy, contrary to elafin and saline, was remarkably effective in preventing type 1 diabetes. T cell receptor spectratyping indicated that AAT gene therapy altered T cell repertoire diversity in splenocytes from NOD mice. Adoptive transfer experiments demonstrated that AAT gene therapy attenuated cellular immunity associated with beta cell destruction. This study demonstrates that AAT gene therapy attenuates cell-mediated autoimmunity, alters the T cell receptor repertoire, and efficiently prevents type 1 diabetes in the NOD mouse model. These results strongly suggest that rAAV1-mediated AAT gene therapy may be useful as a novel approach to prevent type 1 diabetes.

  1. Trichloroethylene effects on gene expression during cardiac development

    SciTech Connect

    Collier, John Michael; Selmin, Ornella; Johnson, Paula D.; Runyan, Raymond B.

    2003-05-09

    Background: Halogenated hydrocarbon exposure is associated with changes in gene expression in adult and embryonic tissue. The present study was undertaken to identify differentially expressed mRNA transcripts in embryonic hearts from Sprague-Dawley rats exposed to trichloroethylene (TCE) or potential bio-transformation products of TCE, Dichloroethylene (DCE) and Trichloroacetic acid (TCAA). Methods: cDNA subtractive hybridization was used to selectively amplify expressed mRNA in either control or day 11 embryonic rat hearts exposed to one of these halogenated hydrocarbons from day 0 to 11. The doses used were 1100 and 110 ppm (8300 and 830 mu M) TCE, 110 and 11 ppm (1100 and 110 mu M) DCE, 27.3 and 2.75 mg/ml (100 and 10 mM) TCAA. Control animals were given distilled drinking water throughout the period of experiments. Results: Sequencing of over 100 clones derived from halogenated hydrocarbon exposed groups=resulted in identification of numerous differentially regulate gene sequences. Up-regulated transcripts identified include genes associated with stress response (Hsp 70) and homeostasis (several ribosomal proteins). Down-regulated transcripts include extracellular matrix components (GPI-p137 and vimentin) and Ca2 + responsive proteins (Serca-2 Ca2+-ATPase and beta-catenin). Two possible markers for fetal TCE exposure were identified: Serca-2 and GPI-p137, a GPI-linked protein of unknown function. Both markers show a dose-related decrease in mRNA transcript levels associated with fetal exposure to TCE. Differential regulation of expression of both markers by TCE was confirmed by dot blot analysis and semi-quantitative RT-PCR. Levels of exposure between 100 and 250 ppb (0.76 and 1.9 mu M) TCE are sufficient to decrease expression of both the Ca2+-AT Pase and GPI-p137. Conclusion: Sequences down-regulated with TCE exposure appear to be those associated with cellular=housekeeping, cell adhesion and developmental processes, while TCE=exposure up-regulates expression

  2. Toward stable gene expression in CHO cells

    PubMed Central

    Mariati; Koh, Esther YC; Yeo, Jessna HM; Ho, Steven CL; Yang, Yuansheng

    2014-01-01

    Maintaining high gene expression level during long-term culture is critical when producing therapeutic recombinant proteins using mammalian cells. Transcriptional silencing of promoters, most likely due to epigenetic events such as DNA methylation and histone modifications, is one of the major mechanisms causing production instability. Previous studies demonstrated that the core CpG island element (IE) from the hamster adenine phosphoribosyltransferase gene is effective to prevent DNA methylation. We generated one set of modified human cytomegalovirus (hCMV) promoters by insertion of one or two copies of IE in either forward or reverse orientations into different locations of the hCMV promoter. The modified hCMV with one copy of IE inserted between the hCMV enhancer and core promoter in reverse orientation (MR1) was most effective at enhancing expression stability in CHO cells without comprising expression level when compared with the wild type hCMV. We also found that insertion of IE into a chimeric murine CMV (mCMV) enhancer and human elongation factor-1α core (hEF) promoter in reverse orientation did not enhance expression stability, indicating that the effect of IE on expression stability is possibly promoter specific. PMID:25482237

  3. Engineering Genes for Predictable Protein Expression

    PubMed Central

    Gustafsson, Claes; Minshull, Jeremy; Govindarajan, Sridhar; Ness, Jon; Villalobos, Alan; Welch, Mark

    2013-01-01

    The DNA sequence used to encode a polypeptide can have dramatic effects on its expression. Lack of readily available tools has until recently inhibited meaningful experimental investigation of this phenomenon. Advances in synthetic biology and the application of modern engineering approaches now provide the tools for systematic analysis of the sequence variables affecting heterologous expression of recombinant proteins. We here discuss how these new tools are being applied and how they circumvent the constraints of previous approaches, highlighting some of the surprising and promising results emerging from the developing field of gene engineering. PMID:22425659

  4. Engineering genes for predictable protein expression.

    PubMed

    Gustafsson, Claes; Minshull, Jeremy; Govindarajan, Sridhar; Ness, Jon; Villalobos, Alan; Welch, Mark

    2012-05-01

    The DNA sequence used to encode a polypeptide can have dramatic effects on its expression. Lack of readily available tools has until recently inhibited meaningful experimental investigation of this phenomenon. Advances in synthetic biology and the application of modern engineering approaches now provide the tools for systematic analysis of the sequence variables affecting heterologous expression of recombinant proteins. We here discuss how these new tools are being applied and how they circumvent the constraints of previous approaches, highlighting some of the surprising and promising results emerging from the developing field of gene engineering.

  5. Circular RNA and gene expression profiles in gastric cancer based on microarray chip technology.

    PubMed

    Sui, Weiguo; Shi, Zhoufang; Xue, Wen; Ou, Minglin; Zhu, Ying; Chen, Jiejing; Lin, Hua; Liu, Fuhua; Dai, Yong

    2017-03-01

    The aim of the present study was to screen gastric cancer (GC) tissue and adjacent tissue for differences in mRNA and circular (circRNA) expression, to analyze the differences in circRNA and mRNA expression, and to investigate the circRNA expression in gastric carcinoma and its mechanism. circRNA and mRNA differential expression profiles generated using Agilent microarray technology were analyzed in the GC tissues and adjacent tissues. qRT-PCR was used to verify the differential expression of circRNAs and mRNAs according to the interactions between circRNAs and miRNAs as well as the possible existence of miRNA and mRNA interactions. We found that: i) the circRNA expression profile revealed 1,285 significant differences in circRNA expression, with circRNA expression downregulated in 594 samples and upregulated in 691 samples via interactions with miRNAs. The qRT-PCR validation experiments showed that hsa_circRNA_400071, hsa_circRNA_000543 and hsa_circRNA_001959 expression was consistent with the microarray analysis results. ii) 29,112 genes were found in the GC tissues and adjacent tissues, including 5,460 differentially expressed genes. Among them, 2,390 differentially expressed genes were upregulated and 3,070 genes were downregulated. Gene Ontology (GO) analysis of the differentially expressed genes revealed these genes involved in biological process classification, cellular component classification and molecular function classification. Pathway analysis of the differentially expressed genes identified 83 significantly enriched genes, including 28 upregulated genes and 55 downregulated genes. iii) 69 differentially expressed circRNAs were found that might adsorb specific miRNAs to regulate the expression of their target gene mRNAs. The conclusions are: i) differentially expressed circRNAs had corresponding miRNA binding sites. These circRNAs regulated the expression of target genes through interactions with miRNAs and might become new molecular biomarkers for GC

  6. Cancer outlier differential gene expression detection.

    PubMed

    Wu, Baolin

    2007-07-01

    We study statistical methods to detect cancer genes that are over- or down-expressed in some but not all samples in a disease group. This has proven useful in cancer studies where oncogenes are activated only in a small subset of samples. We propose the outlier robust t-statistic (ORT), which is intuitively motivated from the t-statistic, the most commonly used differential gene expression detection method. Using real and simulation studies, we compare the ORT to the recently proposed cancer outlier profile analysis (Tomlins and others, 2005) and the outlier sum statistic of Tibshirani and Hastie (2006). The proposed method often has more detection power and smaller false discovery rates. Supplementary information can be found at http://www.biostat.umn.edu/~baolin/research/ort.html.

  7. Programming gene expression with combinatorial promoters

    PubMed Central

    Cox, Robert Sidney; Surette, Michael G; Elowitz, Michael B

    2007-01-01

    Promoters control the expression of genes in response to one or more transcription factors (TFs). The architecture of a promoter is the arrangement and type of binding sites within it. To understand natural genetic circuits and to design promoters for synthetic biology, it is essential to understand the relationship between promoter function and architecture. We constructed a combinatorial library of random promoter architectures. We characterized 288 promoters in Escherichia coli, each containing up to three inputs from four different TFs. The library design allowed for multiple −10 and −35 boxes, and we observed varied promoter strength over five decades. To further analyze the functional repertoire, we defined a representation of promoter function in terms of regulatory range, logic type, and symmetry. Using these results, we identified heuristic rules for programming gene expression with combinatorial promoters. PMID:18004278