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Sample records for gene expression cellular

  1. Molecular crowding shapes gene expression in synthetic cellular nanosystems

    NASA Astrophysics Data System (ADS)

    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. Modeling gene expression using chromatin features in various cellular contexts

    PubMed Central

    2012-01-01

    Background Previous work has demonstrated that chromatin feature levels correlate with gene expression. The ENCODE project enables us to further explore this relationship using an unprecedented volume of data. Expression levels from more than 100,000 promoters were measured using a variety of high-throughput techniques applied to RNA extracted by different protocols from different cellular compartments of several human cell lines. ENCODE also generated the genome-wide mapping of eleven histone marks, one histone variant, and DNase I hypersensitivity sites in seven cell lines. Results We built a novel quantitative model to study the relationship between chromatin features and expression levels. Our study not only confirms that the general relationships found in previous studies hold across various cell lines, but also makes new suggestions about the relationship between chromatin features and gene expression levels. We found that expression status and expression levels can be predicted by different groups of chromatin features, both with high accuracy. We also found that expression levels measured by CAGE are better predicted than by RNA-PET or RNA-Seq, and different categories of chromatin features are the most predictive of expression for different RNA measurement methods. Additionally, PolyA+ RNA is overall more predictable than PolyA- RNA among different cell compartments, and PolyA+ cytosolic RNA measured with RNA-Seq is more predictable than PolyA+ nuclear RNA, while the opposite is true for PolyA- RNA. Conclusions Our study provides new insights into transcriptional regulation by analyzing chromatin features in different cellular contexts. PMID:22950368

  3. 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

  4. Control of Rta expression critically determines transcription of viral and cellular genes following gammaherpesvirus infection.

    PubMed

    Hair, James R; Lyons, Paul A; Smith, Kenneth G C; Efstathiou, Stacey

    2007-06-01

    The replication and transcriptional activator (Rta), encoded by ORF50 of gammaherpesviruses, initiates the lytic cycle of gene expression; therefore understanding the impact of Rta on viral and cellular gene expression is key to elucidating the transcriptional events governing productive infection and reactivation from latency. To this end, the impact of altering Rta transcription on viral and cellular gene expression was studied in the context of a whole virus infection. Recombinant murine gammaherpesvirus (MHV)-68 engineered to overexpress Rta greatly accelerated expression of specific lytic cycle ORFs, but repressed transcription of the major latency gene, ORF73. Increased expression of Rta accelerated the dysregulation in transcription of specific cellular genes when compared with cells infected with wild-type and revertant viruses. A subset of cellular genes was dysregulated only in cells infected with Rta-overexpressing virus, and never in those infected with non-overexpressing viruses. These data highlight the critical role of Rta abundance in governing viral and cellular gene transcription, and demonstrate the importance of understanding how the relative expression of ORF50 during the virus life cycle impacts on these processes. PMID:17485528

  5. Control of Rta expression critically determines transcription of viral and cellular genes following gammaherpesvirus infection

    PubMed Central

    Hair, James R.; Lyons, Paul A.; Smith, Kenneth G. C.; Efstathiou, Stacey

    2007-01-01

    The replication and transcriptional activator (Rta), encoded by ORF50 of gammaherpesviruses, initiates the lytic cycle of gene expression; therefore understanding the impact of Rta on viral and cellular gene expression is key to elucidating the transcriptional events governing productive infection and reactivation from latency. To this end, the impact of altering Rta transcription on viral and cellular gene expression was studied in the context of a whole virus infection. Recombinant murine gammaherpesvirus (MHV)-68 engineered to overexpress Rta greatly accelerated expression of specific lytic cycle ORFs, but repressed transcription of the major latency gene, ORF73. Increased expression of Rta accelerated the dysregulation in transcription of specific cellular genes when compared with cells infected with wild-type and revertant viruses. A subset of cellular genes was dysregulated only in cells infected with Rta-overexpressing virus, and never in those infected with non-overexpressing viruses. These data highlight the critical role of Rta abundance in governing viral and cellular gene transcription, and demonstrate the importance of understanding how the relative expression of ORF50 during the virus life cycle impacts on these processes. PMID:17485528

  6. Automated analysis of embryonic gene expression with cellular resolution in C. elegans

    PubMed Central

    Murray, John Isaac; Bao, Zhirong; Boyle, Thomas J.; Boeck, Max E.; Mericle, Barbara L.; Nicholas, Thomas J.; Zhao, Zhongying; Sandel, Matthew J.; Waterston, Robert H.

    2008-01-01

    We describe a system that permits the automated analysis of reporter gene expression in Caenorhabditis elegans with cellular resolution continuously during embryogenesis and demonstrate its utility by defining the expression patterns of reporters for several embryonically expressed transcription factors. The invariant cell lineage permits the automated alignment of multiple expression profiles, allowing the direct comparison of the expression of different genes' reporters. We have also used the system to monitor perturbations to normal development involving changes both in cell division timing and in cell fate. Systematic application could reveal the gene activity of each cell throughout development. PMID:18587405

  7. Differential cellular gene expression induced by hepatitis B and C viruses.

    PubMed

    Otsuka, Motoyuki; Aizaki, Hideki; Kato, Naoya; Suzuki, Tetsuro; Miyamura, Tatsuo; Omata, Masao; Seki, Naohiko

    2003-01-10

    Hepatitis B virus (HBV) is a hepatotropic virus that causes acute and chronic hepatocellular injury and hepatocellular carcinoma. To clarify how HBV proteins regulate host cellular gene expression, we used our in-house cDNA microarray and HepG2.2.15 cells, which are derived from HepG2 cells and produce all HBV proteins. Of 2304 genes investigated, several genes were differentially expressed in HepG2.2.15 cells compared with HepG2 cells. These genes included insulin-like growth factor II and alpha-fetoprotein, consistent with previous reports. Furthermore, we previously performed similar microarray analyses to clarify the effects of hepatitis C virus (HCV) proteins on host cells, using a HepG2-derivative cell line, which produces all HCV proteins. Using these two microarray results, we compared the differences in cellular gene expression induced by HBV and HCV proteins. The expression of the majority of genes investigated differed only slightly between HBV and HCV protein-producing cells. However, HBV and HCV proteins clearly regulated several genes in a reciprocal manner. Combined, these microarray results shed new light on the effects of HBV proteins on cellular gene expression and on the differences in the pathogenic activities of these two hepatitis viruses. PMID:12504104

  8. Cellular gene expression induced by parasite antigens and allergens in neonates from parasite-infected mothers.

    PubMed

    Soboslay, Peter T; Orlikowsky, Thorsten; Huang, Xiangsheng; Gille, Christian; Spring, Bärbel; Kocherscheidt, Lars; Agossou, Abram; Banla, Meba; Bonin, Michael; Köhler, Carsten

    2016-05-01

    Prenatal exposure to parasite antigens or allergens will influence the profile and strength of postnatal immune responses, such contact may tolerize and increase susceptibility to future infections or sensitize to environmental allergens. Exposure in utero to parasite antigens will distinctly alter cellular gene expression in newborns. Gene microarrays were applied to study gene expression in umbilical cord blood cell (UCBC) from parasite-exposed (Para-POS) and non-exposed (Para-NEG) neonates. UCBC were activated with antigens of helminth (Onchocerca volvulus), amoeba (Entamoeba histolytica) or allergens of mite (Dermatophagoides farinae). When UCBC from Para-POS and Para-NEG newborns were exposed to helminth antigens or allergens consistent differences occurred in the expression of genes encoding for MHC class I and II alleles, signal transducers of activation and transcription (STATs), cytokines, chemokines, immunoglobulin heavy and light chains, and molecules associated with immune regulation (SOCS, TLR, TGF), inflammation (TNF, CCR) and apoptosis (CASP). Expression of genes associated with innate immune responses were enhanced in Para-NEG, while in Para-POS, the expression of MHC class II and STAT genes was reduced. Within functional gene networks for cellular growth, proliferation and immune responses, Para-NEG neonates presented with significantly higher expression values than Para-POS. In Para-NEG newborns, the gene cluster and pathway analyses suggested that gene expression profiles may predispose for the development of immunological, hematological and dermatological disorders upon postnatal helminth parasite infection or allergen exposure. Thus, prenatal parasite contact will sensitize without generating aberrant inflammatory immune responses, and increased pro-inflammatory but decreased regulatory gene expression profiles will be present in those neonates lacking prenatal parasite antigen encounter. PMID:27062712

  9. Glucose Oxidase Induces Cellular Senescence in Immortal Renal Cells through ILK by Downregulating Klotho Gene Expression.

    PubMed

    Troyano-Suárez, Nuria; del Nogal-Avila, María; Mora, Inés; Sosa, Patricia; López-Ongil, Susana; Rodriguez-Puyol, Diego; Olmos, Gemma; Ruíz-Torres, María Piedad

    2015-01-01

    Cellular senescence can be prematurely induced by oxidative stress involved in aging. In this work, we were searching for novel intermediaries in oxidative stress-induced senescence, focusing our interest on integrin-linked kinase (ILK), a scaffold protein at cell-extracellular matrix (ECM) adhesion sites, and on the Klotho gene. Cultured renal cells were treated with glucose oxidase (GOx) for long time periods. GOx induced senescence, increasing senescence associated β-galactosidase activity and the expression of p16. In parallel, GOx increased ILK protein expression and activity. Ectopic overexpression of ILK in cells increased p16 expression, even in the absence of GOx, whereas downregulation of ILK inhibited the increase in p16 due to oxidative stress. Additionally, GOx reduced Klotho gene expression and cells overexpressing Klotho protein did not undergo senescence after GOx addition. We demonstrated a direct link between ILK and Klotho since silencing ILK expression in cells and mice increases Klotho expression and reduces p53 and p16 expression in renal cortex. In conclusion, oxidative stress induces cellular senescence in kidney cells by increasing ILK protein expression and activity, which in turn reduces Klotho expression. We hereby present ILK as a novel downregulator of Klotho gene expression. PMID:26583057

  10. Glucose Oxidase Induces Cellular Senescence in Immortal Renal Cells through ILK by Downregulating Klotho Gene Expression

    PubMed Central

    Troyano-Suárez, Nuria; del Nogal-Avila, María; Mora, Inés; Sosa, Patricia; López-Ongil, Susana; Rodriguez-Puyol, Diego; Olmos, Gemma; Ruíz-Torres, María Piedad

    2015-01-01

    Cellular senescence can be prematurely induced by oxidative stress involved in aging. In this work, we were searching for novel intermediaries in oxidative stress-induced senescence, focusing our interest on integrin-linked kinase (ILK), a scaffold protein at cell-extracellular matrix (ECM) adhesion sites, and on the Klotho gene. Cultured renal cells were treated with glucose oxidase (GOx) for long time periods. GOx induced senescence, increasing senescence associated β-galactosidase activity and the expression of p16. In parallel, GOx increased ILK protein expression and activity. Ectopic overexpression of ILK in cells increased p16 expression, even in the absence of GOx, whereas downregulation of ILK inhibited the increase in p16 due to oxidative stress. Additionally, GOx reduced Klotho gene expression and cells overexpressing Klotho protein did not undergo senescence after GOx addition. We demonstrated a direct link between ILK and Klotho since silencing ILK expression in cells and mice increases Klotho expression and reduces p53 and p16 expression in renal cortex. In conclusion, oxidative stress induces cellular senescence in kidney cells by increasing ILK protein expression and activity, which in turn reduces Klotho expression. We hereby present ILK as a novel downregulator of Klotho gene expression. PMID:26583057

  11. Control of adenovirus early gene expression: Posttranscriptional control mediated by both viral and cellular gene products

    SciTech Connect

    Katze, M.G.; Persson, H.; Philipson, L.

    1981-09-01

    An adenovirus type 5 host range mutant (hr-1) located in region E1A and phenotypically defective in expressing viral messenger ribonucleic acid (RNA) from other early regions was analyzed for accumulation of viral RNA in the presence of protein synthesis inhibitors. Nuclear RNA was transcribed from all early regions at the same rate, regardless of whether the drug was present or absent. As expected, low or undetectable levels of RNA were found in the cytoplasm of hr-1-infected cells compared with the wild-type adenovirus type 5 in the absence of drug. When anisomycin was added 30 min before hr-1 infection, cytoplasmic RNA was abundant from early regions E3 and E4 when assayed by filter hybridization. In accordance, early regions E3 and E4 viral messenger RNA species were detected by the S1 endonuclease mapping technique only in hr-1-infected cells that were treated with the drug. Similar results were obtained by in vitro translation studies. Together, these results suggest that this adenovirus type 5 mutant lacks a viral gene product necessary for accumulation of viral messenger RNA, but not for transcription. It is proposed that a cellular gene product serves as a negative regulator of viral messenger RNA accumulation at the posttranscriptional level.

  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. Three-dimensional morphology and gene expression in the Drosophila blastoderm at cellular resolution II: dynamics

    PubMed Central

    Keränen, Soile VE; Fowlkes, Charless C; Luengo Hendriks, Cris L; Sudar, Damir; Knowles, David W; Malik, Jitendra; Biggin, Mark D

    2006-01-01

    Background To accurately describe gene expression and computationally model animal transcriptional networks, it is essential to determine the changing locations of cells in developing embryos. Results Using automated image analysis methods, we provide the first quantitative description of temporal changes in morphology and gene expression at cellular resolution in whole embryos, using the Drosophila blastoderm as a model. Analyses based on both fixed and live embryos reveal complex, previously undetected three-dimensional changes in nuclear density patterns caused by nuclear movements prior to gastrulation. Gene expression patterns move, in part, with these changes in morphology, but additional spatial shifts in expression patterns are also seen, supporting a previously proposed model of pattern dynamics based on the induction and inhibition of gene expression. We show that mutations that disrupt either the anterior/posterior (a/p) or the dorsal/ventral (d/v) transcriptional cascades alter morphology and gene expression along both the a/p and d/v axes in a way suggesting that these two patterning systems interact via both transcriptional and morphological mechanisms. Conclusion Our work establishes a new strategy for measuring temporal changes in the locations of cells and gene expression patterns that uses fixed cell material and computational modeling. It also provides a coordinate framework for the blastoderm embryo that will allow increasingly accurate spatio-temporal modeling of both the transcriptional control network and morphogenesis. PMID:17184547

  14. 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

  15. FLI1 expression is correlated with breast cancer cellular growth, migration, and invasion and altered gene expression.

    PubMed

    Scheiber, Melissa N; Watson, Patricia M; Rumboldt, Tihana; Stanley, Connor; Wilson, Robert C; Findlay, Victoria J; Anderson, Paul E; Watson, Dennis K

    2014-10-01

    ETS factors have been shown to be dysregulated in breast cancer. ETS factors control the expression of genes involved in many biological processes, such as cellular proliferation, differentiation, and apoptosis. FLI1 is an ETS protein aberrantly expressed in retrovirus-induced hematological tumors, but limited attention has been directed towards elucidating the role of FLI1 in epithelial-derived cancers. Using data mining, we show that loss of FLI1 expression is associated with shorter survival and more aggressive phenotypes of breast cancer. Gain and loss of function cellular studies indicate the inhibitory effect of FLI1 expression on cellular growth, migration, and invasion. Using Fli1 mutant mice and both a transgenic murine breast cancer model and an orthotopic injection of syngeneic tumor cells indicates that reduced Fli1 contributes to accelerated tumor growth. Global expression analysis and RNA-Seq data from an invasive human breast cancer cell line with over expression of either FLI1 and another ETS gene, PDEF, shows changes in several cellular pathways associated with cancer, such as the cytokine-cytokine receptor interaction and PI3K-Akt signaling pathways. This study demonstrates a novel role for FLI1 in epithelial cells. In addition, these results reveal that FLI1 down-regulation in breast cancer may promote tumor progression. PMID:25379017

  16. FLI1 Expression is Correlated with Breast Cancer Cellular Growth, Migration, and Invasion and Altered Gene Expression

    PubMed Central

    Scheiber, Melissa N.; Watson, Patricia M.; Rumboldt, Tihana; Stanley, Connor; Wilson, Robert C.; Findlay, Victoria J.; Anderson, Paul E.; Watson, Dennis K.

    2014-01-01

    ETS factors have been shown to be dysregulated in breast cancer. ETS factors control the expression of genes involved in many biological processes, such as cellular proliferation, differentiation, and apoptosis. FLI1 is an ETS protein aberrantly expressed in retrovirus-induced hematological tumors, but limited attention has been directed towards elucidating the role of FLI1 in epithelial-derived cancers. Using data mining, we show that loss of FLI1 expression is associated with shorter survival and more aggressive phenotypes of breast cancer. Gain and loss of function cellular studies indicate the inhibitory effect of FLI1 expression on cellular growth, migration, and invasion. Using Fli1 mutant mice and both a transgenic murine breast cancer model and an orthotopic injection of syngeneic tumor cells indicates that reduced Fli1 contributes to accelerated tumor growth. Global expression analysis and RNA-Seq data from an invasive human breast cancer cell line with over expression of either FLI1 and another ETS gene, PDEF, shows changes in several cellular pathways associated with cancer, such as the cytokine-cytokine receptor interaction and PI3K-Akt signaling pathways. This study demonstrates a novel role for FLI1 in epithelial cells. In addition, these results reveal that FLI1 down-regulation in breast cancer may promote tumor progression. PMID:25379017

  17. Cellular Expression of Smarca4 (Brg1)-regulated Genes in Zebrafish Retinas

    PubMed Central

    2011-01-01

    Background In a recent genomic study, Leung et al. used a factorial microarray analysis to identify Smarca4 (Brg1)-regulated genes in micro-dissected zebrafish retinas. Two hundred and fifty nine genes were grouped in three-way ANOVA models which carried the most specific retinal change. To validate the microarray results and to elucidate cellular expression patterns of the significant genes for further characterization, 32 known genes were randomly selected from this group. In situ hybridization of these genes was performed on the same types of samples (wild-type (WT) and smarca4a50/a50 (yng) mutant) at the same stages (36 and 52 hours post-fertilization (hpf)) as in the microarray study. Results Thirty out of 32 riboprobes showed a positive in situ staining signal. Twenty seven out of these 30 genes were originally further classified as Smarca4-regulated retinal genes, while the remaining three as retinal-specific expression independent of Smarca4 regulation. It was found that 90.32% of the significant microarray comparisons that were used to identify Smarca4-regulated retinal genes had a corresponding qualitative expression change in the in situ hybridization comparisons. This is highly concordant with the theoretical true discovery rate of 95%. Hierarchical clustering was used to investigate the similarity of the cellular expression patterns of 25 out of the 27 Smarca4-regulated retinal genes that had a sufficiently high expression signal for an unambiguous identification of retinal expression domains. Three broad groups of expression pattern were identified; including 1) photoreceptor layer/outer nuclear layer specific expression at 52 hpf, 2) ganglion cell layer (GCL) and/or inner nuclear layer (INL) specific expression at both 36 & 52 hpf, and 3) GCL and/or INL specific expression at 52 hpf only. Some of these genes have recently been demonstrated to play key roles in retinal cell-type specification, differentiation and lamination. For the remaining three

  18. 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

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

    PubMed

    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; Vitiello, Seasson P

    2016-01-01

    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

  20. 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

  1. Regulation of viral and cellular gene expression by Kaposi's sarcoma-associated herpesvirus polyadenylated nuclear RNA.

    PubMed

    Rossetto, Cyprian C; Tarrant-Elorza, Margaret; Verma, Subhash; Purushothaman, Pravinkumar; Pari, Gregory S

    2013-05-01

    Kaposi's sarcoma-associated herpesvirus (KSHV) is the cause of Kaposi's sarcoma and body cavity lymphoma. In cell culture, KSHV results in a latent infection, and lytic reactivation is usually induced with the expression of K-Rta or by treatment with phorbol 12-myristate 13-acetate (TPA) and/or n-butyrate. Lytic infection is marked by the activation of the entire viral genomic transcription cascade and the production of infectious virus. KSHV-infected cells express a highly abundant, long, noncoding transcript referred to as polyadenylated nuclear RNA (PAN RNA). PAN RNA interacts with specific demethylases and physically binds to the KSHV genome to mediate activation of viral gene expression. A recombinant BACmid lacking the PAN RNA locus fails to express K-Rta and does not produce virus. We now show that the lack of PAN RNA expression results in the failure of the initiation of the entire KSHV transcription program. In addition to previous findings of an interaction with demethylases, we show that PAN RNA binds to protein components of Polycomb repression complex 2 (PRC2). RNA-Seq analysis using cell lines that express PAN RNA shows that transcription involving the expression of proteins involved in cell cycle, immune response, and inflammation is dysregulated. Expression of PAN RNA in various cell types results in an enhanced growth phenotype, higher cell densities, and increased survival compared to control cells. Also, PAN RNA expression mediates a decrease in the production of inflammatory cytokines. These data support a role for PAN RNA as a major global regulator of viral and cellular gene expression. PMID:23468496

  2. Ebola virion attachment and entry into human macrophages profoundly effects early cellular gene expression.

    PubMed

    Wahl-Jensen, Victoria; Kurz, Sabine; Feldmann, Friedericke; Buehler, Lukas K; Kindrachuk, Jason; DeFilippis, Victor; da Silva Correia, Jean; Früh, Klaus; Kuhn, Jens H; Burton, Dennis R; Feldmann, Heinz

    2011-10-01

    Zaire ebolavirus (ZEBOV) infections are associated with high lethality in primates. ZEBOV primarily targets mononuclear phagocytes, which are activated upon infection and secrete mediators believed to trigger initial stages of pathogenesis. The characterization of the responses of target cells to ZEBOV infection may therefore not only further understanding of pathogenesis but also suggest possible points of therapeutic intervention. Gene expression profiles of primary human macrophages exposed to ZEBOV were determined using DNA microarrays and quantitative PCR to gain insight into the cellular response immediately after cell entry. Significant changes in mRNA concentrations encoding for 88 cellular proteins were observed. Most of these proteins have not yet been implicated in ZEBOV infection. Some, however, are inflammatory mediators known to be elevated during the acute phase of disease in the blood of ZEBOV-infected humans. Interestingly, the cellular response occurred within the first hour of Ebola virion exposure, i.e. prior to virus gene expression. This observation supports the hypothesis that virion binding or entry mediated by the spike glycoprotein (GP(1,2)) is the primary stimulus for an initial response. Indeed, ZEBOV virions, LPS, and virus-like particles consisting of only the ZEBOV matrix protein VP40 and GP(1,2) (VLP(VP40-GP)) triggered comparable responses in macrophages, including pro-inflammatory and pro-apoptotic signals. In contrast, VLP(VP40) (particles lacking GP(1,2)) caused an aberrant response. This suggests that GP(1,2) binding to macrophages plays an important role in the immediate cellular response. PMID:22028943

  3. 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.

  4. BRCA1 Haploinsufficiency Leads to Altered Expression of Genes Involved in Cellular Proliferation and Development

    PubMed Central

    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. PMID:24950059

  5. Cellular defense system gene expression profiling of human whole blood: opportunities to predict health benefits in response to diet.

    PubMed

    Drew, Janice E

    2012-07-01

    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

  6. Modulation of Cellular and Viral Gene Expression by the Latency-Associated Nuclear Antigen of Kaposi's Sarcoma-Associated Herpesvirus

    PubMed Central

    Renne, Rolf; Barry, Chris; Dittmer, Dirk; Compitello, Nicole; Brown, Patrick O.; Ganem, Don

    2001-01-01

    Kaposi's sarcoma-associated herpesvirus (KSHV), also called human herpesvirus 8 (HHV-8), is the likely etiological agent of Kaposi's sarcoma and primary effusion lymphoma. Common to these malignancies is that tumor cells are latently infected with KSHV. Viral gene expression is limited to a few genes, one of which is the latency-associated nuclear antigen (LANA), the product of ORF73. Examination of the primary sequence of LANA reveals some structural features reminiscent of transcription factors, leading us to hypothesize that LANA may regulate viral and cellular transcription during latency. In reporter gene-based transient transfection assays, we found that LANA can have either positive or negative effects on gene expression. While expression of a reporter gene from several synthetic promoters was increased in the presence of LANA, expression from the human immunodeficiency virus (HIV) long terminal repeat (LTR)—and from NF-κB-dependent reporter genes—was reduced by LANA expression. In addition, the promoter of KSHV ORF73 itself is activated up to 5.5-fold by LANA. This autoregulation may be important in tumorigenesis, because two other genes (v-cyclin and v-FLIP) with likely roles in cell growth and survival are also controlled by this element. To identify cellular genes influenced by LANA, we employed cDNA array-based expression profiling. Six known genes (and nine expressed sequence tags) were found to be upregulated in LANA-expressing cell lines. One of these, Staf-50, is known to inhibit expression from the HIV LTR; most of the other known genes are interferon inducible, although the interferon genes themselves were not induced by LANA. These data demonstrate that LANA expression has effects on cellular and viral gene expression. We suggest that, whether direct or indirect in origin, these effects may play important roles in the pathobiology of KSHV infection. PMID:11119614

  7. Cellular Adhesion Gene SELP Is Associated with Rheumatoid Arthritis and Displays Differential Allelic Expression

    PubMed Central

    Petit-Teixeira, Elisabeth; Hugo Teixeira, Vitor; Steiner, Anke; Quente, Elfi; Wolfram, Grit; Scholz, Markus; Pierlot, Céline; Migliorini, Paola; Bombardieri, Stefano; Balsa, Alejandro; Westhovens, René; Barrera, Pilar; Radstake, Timothy R. D. J.; Alves, Helena; Bardin, Thomas; Prum, Bernard; Emmrich, Frank; Cornelis, François

    2014-01-01

    In rheumatoid arthritis (RA), a key event is infiltration of inflammatory immune cells into the synovial lining, possibly aggravated by dysregulation of cellular adhesion molecules. Therefore, single nucleotide polymorphisms of 14 genes involved in cellular adhesion processes (CAST, ITGA4, ITGB1, ITGB2, PECAM1, PTEN, PTPN11, PTPRC, PXN, SELE, SELP, SRC, TYK2, and VCAM1) were analyzed for association with RA. Association analysis was performed consecutively in three European RA family sample groups (Nfamilies = 407). Additionally, we investigated differential allelic expression, a possible functional consequence of genetic variants. SELP (selectin P, CD62P) SNP-allele rs6136-T was associated with risk for RA in two RA family sample groups as well as in global analysis of all three groups (ptotal = 0.003). This allele was also expressed preferentially (p<10−6) with a two- fold average increase in regulated samples. Differential expression is supported by data from Genevar MuTHER (p1 = 0.004; p2 = 0.0177). Evidence for influence of rs6136 on transcription factor binding was also found in silico and in public datasets reporting in vitro data. In summary, we found SELP rs6136-T to be associated with RA and with increased expression of SELP mRNA. SELP is located on the surface of endothelial cells and crucial for recruitment, adhesion, and migration of inflammatory cells into the joint. Genetically determined increased SELP expression levels might thus be a novel additional risk factor for RA. PMID:25147926

  8. Mechanisms of Cardiovascular Homeostasis and Pathophysiology--From Gene Expression, Signal Transduction to Cellular Communication.

    PubMed

    Akazawa, Hiroshi

    2015-01-01

    During embryogenesis, progenitor cells are specified and differentiated into mature cardiomyocytes. Soon after birth, the ability of cardiomyocytes to proliferate is strongly restrained, and thereafter, they grow in size without cell division. Under pathological conditions, cardiomyocytes show adaptive and maladaptive responses through complex intracellular signaling pathways and cross-talking networks of intercellular and inter-tissue communications, but ultimately, they become dysfunctional and undergo cell death or degeneration. Cardiovascular diseases remain the most prevalent, costly, disabling, and deadly medical conditions. To develop novel therapies for them, it is important to elucidate the underlying mechanisms that govern gene expression, signal transduction to cellular communication. In this review article for the 2014 SATO Memorial Award, an approach to uncover molecular and cellular pathophysiology is summarized, focusing on homeobox transcription factor Nkx2-5 in the transcriptional regulation of the cardiac gene program, 3-phosphoinositide-dependent kinase-1, in the regulation of postnatal cardiomyocyte growth, survival, and function, angiotensin II type 1 receptor in the development of pathological hypertrophy and remodeling, and mast cell infiltration in the pathogenesis of atrial remodeling and fibrillation. PMID:26538467

  9. 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

  10. 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

  11. 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. PMID:25225046

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

    PubMed

    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

    2015-04-20

    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

  13. 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. PMID:26420239

  14. 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

  15. Cellular Genes in the Mouse Regulate IN TRANS the Expression of Endogenous Mouse Mammary Tumor Viruses

    PubMed Central

    Traina-Dorge, Vicki L.; Carr, Jean K.; Bailey-Wilson, Joan E.; Elston, Robert C.; Taylor, Benjamin A.; Cohen, J. Craig

    1985-01-01

    The transcriptional activities of the eleven mouse mammary tumor virus (MMTV) proviruses endogenous to two sets of recombinant inbred (RI) mouse strains, BXD and BXH, were characterized. Comparison of the levels of virus-specific RNA quantitated in each strain showed no direct relationship between the presence of a particular endogenous provirus or with increasing numbers of proviruses. Association of specific genetic markers with the level of MMTV-specific RNA was examined by using multiple regression analysis. Several cellular loci as well as proviral loci were identified that were significantly associated with viral expression. Importantly, these cellular loci associated with MMTV expression segregated independently of viral sequences. PMID:2996982

  16. Human papillomavirus type 16 E7 perturbs DREAM to promote cellular proliferation and mitotic gene expression

    PubMed Central

    DeCaprio, James A.

    2014-01-01

    Study of the small DNA tumor viruses continues to provide valuable new insights into oncogenesis and fundamental biological processes. While much has already been revealed about how the human papillomaviruses (HPVs) can transform cells and contribute to cervical and oropharyngeal cancer, there clearly is much more to learn. In this issue of Oncogene, Pang et al. demonstrate that the high-risk HPV16 E7 oncogene can promote cellular proliferation by interacting with the DREAM (DP, RB-like, E2F and MuvB) complex at two distinct phases of the cell cycle (1). Consistent with earlier work, HPV16 E7 can bind to the retinoblastoma tumor suppressor (RB) family member p130 (RBL2) protein and promote its proteasome-mediated destruction thereby disrupting the DREAM complex and prevent exit from the cell cycle into quiescence. In addition, they demonstrate that HPV16 E7 can bind to MuvB core complex in association with BMYB and FOXM1 and activate gene expression during the G2 and M phase of the cell cycle. Thus, HPV16 E7 acts to prevent exit from the cell cycle entry and promotes mitotic proliferation and may account for the high levels of FOXM1 often observed in poor risk cervical cancers. PMID:24166507

  17. Human papillomavirus type 16 E7 perturbs DREAM to promote cellular proliferation and mitotic gene expression.

    PubMed

    DeCaprio, J A

    2014-07-31

    The study of the small DNA tumor viruses continues to provide valuable new insights into oncogenesis and fundamental biological processes. Although much has already been revealed about how the human papillomaviruses (HPVs) can transform cells and contribute to cervical and oropharyngeal cancer, there clearly is much more to learn. In this issue of Oncogene, Pang et al., doi:10.1038/onc.2013.426, demonstrate that the high-risk HPV16 E7 oncogene can promote cellular proliferation by interacting with the DREAM (DP, RB-like, E2F and MuvB) complex at two distinct phases of the cell cycle. Consistent with earlier work, HPV16 E7 can bind to the retinoblastoma tumor suppressor (RB) family member p130 (RBL2) protein and promote its proteasome-mediated destruction thereby disrupting the DREAM complex and can prevent exit from the cell cycle into quiescence. In addition, they demonstrate that HPV16 E7 can bind to MuvB core complex in association with BMYB and FOXM1 and activate gene expression during the G2 and M phase of the cell cycle. Thus, HPV16 E7 acts to prevent exit from the cell cycle entry and promotes mitotic proliferation and may account for the high levels of FOXM1 often observed in poor-risk cervical cancers. PMID:24166507

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

    PubMed

    Yamada, Tomoya; Higuchi, Mikito; Nakanishi, Naoto

    2014-03-01

    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. PMID:24525120

  19. 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.

  20. Computational evaluation of cellular metabolic costs successfully predicts genes whose expression is deleterious

    PubMed Central

    Wagner, Allon; Zarecki, Raphy; Reshef, Leah; Gochev, Camelia; Sorek, Rotem; Gophna, Uri; Ruppin, Eytan

    2013-01-01

    Gene suppression and overexpression are both fundamental tools in linking genotype to phenotype in model organisms. Computational methods have proven invaluable in studying and predicting the deleterious effects of gene deletions, and yet parallel computational methods for overexpression are still lacking. Here, we present Expression-Dependent Gene Effects (EDGE), an in silico method that can predict the deleterious effects resulting from overexpression of either native or foreign metabolic genes. We first test and validate EDGE’s predictive power in bacteria through a combination of small-scale growth experiments that we performed and analysis of extant large-scale datasets. Second, a broad cross-species analysis, ranging from microorganisms to multiple plant and human tissues, shows that genes that EDGE predicts to be deleterious when overexpressed are indeed typically down-regulated. This reflects a universal selection force keeping the expression of potentially deleterious genes in check. Third, EDGE-based analysis shows that cancer genetic reprogramming specifically suppresses genes whose overexpression impedes proliferation. The magnitude of this suppression is large enough to enable an almost perfect distinction between normal and cancerous tissues based solely on EDGE results. We expect EDGE to advance our understanding of human pathologies associated with up-regulation of particular transcripts and to facilitate the utilization of gene overexpression in metabolic engineering. PMID:24198337

  1. A Digital Framework to Build, Visualize and Analyze a Gene Expression Atlas with Cellular Resolution in Zebrafish Early Embryogenesis

    PubMed Central

    Castro-González, Carlos; Luengo-Oroz, Miguel A.; Duloquin, Louise; Savy, Thierry; Rizzi, Barbara; Desnoulez, Sophie; Doursat, René; Kergosien, Yannick L.; Ledesma-Carbayo, María J.; Bourgine, Paul

    2014-01-01

    A gene expression atlas is an essential resource to quantify and understand the multiscale processes of embryogenesis in time and space. The automated reconstruction of a prototypic 4D atlas for vertebrate early embryos, using multicolor fluorescence in situ hybridization with nuclear counterstain, requires dedicated computational strategies. To this goal, we designed an original methodological framework implemented in a software tool called Match-IT. With only minimal human supervision, our system is able to gather gene expression patterns observed in different analyzed embryos with phenotypic variability and map them onto a series of common 3D templates over time, creating a 4D atlas. This framework was used to construct an atlas composed of 6 gene expression templates from a cohort of zebrafish early embryos spanning 6 developmental stages from 4 to 6.3 hpf (hours post fertilization). They included 53 specimens, 181,415 detected cell nuclei and the segmentation of 98 gene expression patterns observed in 3D for 9 different genes. In addition, an interactive visualization software, Atlas-IT, was developed to inspect, supervise and analyze the atlas. Match-IT and Atlas-IT, including user manuals, representative datasets and video tutorials, are publicly and freely available online. We also propose computational methods and tools for the quantitative assessment of the gene expression templates at the cellular scale, with the identification, visualization and analysis of coexpression patterns, synexpression groups and their dynamics through developmental stages. PMID:24945246

  2. 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

  3. Identification of Novel Cellular Targets in Biliary Tract Cancers Using Global Gene Expression Technology

    PubMed Central

    Hansel, Donna E.; Rahman, Ayman; Hidalgo, Manuel; Thuluvath, Paul J.; Lillemoe, Keith D.; Shulick, Richard; Ku, Ja-Lok; Park, Jae-Gahb; Miyazaki, Kohje; Ashfaq, Raheela; Wistuba, Ignacio I.; Varma, Ram; Hawthorne, Lesleyann; Geradts, Joseph; Argani, Pedram; Maitra, Anirban

    2003-01-01

    Biliary tract carcinoma carries a poor prognosis, and difficulties with clinical management in patients with advanced disease are often due to frequent late-stage diagnosis, lack of serum markers, and limited information regarding biliary tumor pathogenesis. RNA-based global analyses of gene expression have led to the identification of a large number of up-regulated genes in several cancer types. We have used the recently developed Affymetrix U133A gene expression microarrays containing nearly 22,000 unique transcripts to obtain global gene expression profiles from normal biliary epithelial scrapings (n = 5), surgically resected biliary carcinomas (n = 11), and biliary cancer cell lines (n = 9). Microarray hybridization data were normalized using dCHIP (http://www.dCHIP.org) to identify differentially up-regulated genes in primary biliary cancers and biliary cancer cell lines and their expression profiles was compared to that of normal epithelial scrapings using the dCHIP software as well as Significance Analysis of Microarrays or SAM (http://www-stat.stanford.edu/∼tibs/SAM/). Comparison of the dCHIP and SAM datasets revealed an overlapping list of 282 genes expressed at greater than threefold levels in the cancers compared to normal epithelium (t-test P <0.1 in dCHIP, and median false discovery rate <10 in SAM). Several pathways integral to tumorigenesis were up-regulated in the biliary cancers, including proliferation and cell cycle antigens (eg, cyclins D2 and E2, cdc2/p34, and geminin), transcription factors (eg, homeobox B7 and islet-1), growth factors and growth factor receptors (eg, hepatocyte growth factor, amphiregulin, and insulin-like growth factor 1 receptor), and enzymes modulating sensitivity to chemotherapeutic agents (eg, cystathionine β synthase, dCMP deaminase, and CTP synthase). In addition, we identified several “pathway” genes that are rapidly emerging as novel therapeutic targets in cancer (eg, cytosolic phospholipase A2, an upstream

  4. Computational deconvolution of gene expression by individual host cellular subsets from microarray analyses of complex, parasite-infected whole tissues.

    PubMed

    Banskota, Nirad; Odegaard, Justin I; Rinaldi, Gabriel; Hsieh, Michael H

    2016-06-01

    Analyses of whole organs from parasite-infected animals can reveal the entirety of the host tissue transcriptome, but conventional approaches make it difficult to dissect out the contributions of individual cellular subsets to observed gene expression. Computational deconvolution of gene expression data may be one solution to this problem. We tested this potential solution by deconvoluting whole bladder gene expression microarray data derived from a model of experimental urogenital schistosomiasis. A supervised technique was used to group B-cell and T-cell related genes based on their cell types, with a semi-supervised technique to calculate the proportions of urothelial cells. We demonstrate that the deconvolution technique was able to group genes into their correct cell types with good accuracy. A clustering-based methodology was also used to improve prediction. However, incorrectly predicted genes could not be discriminated using this methodology. The incorrect predictions were primarily IgH- and IgK-related genes. To our knowledge, this is the first application of computational deconvolution to complex, parasite-infected whole tissues. Other computational techniques such as neural networks may need to be used to improve prediction. PMID:27025770

  5. 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.

  6. 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

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

    PubMed

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

    2016-08-23

    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

  8. Epstein-Barr virus induces cellular transcription factors to allow active expression of EBER genes by RNA polymerase III.

    PubMed

    Felton-Edkins, Zoë A; Kondrashov, Alexander; Karali, Dimitra; Fairley, Jennifer A; Dawson, Christopher W; Arrand, John R; Young, Lawrence S; White, Robert J

    2006-11-10

    The EBER genes of Epstein-Barr virus (EBV) are transcribed by RNA polymerase (pol) III to produce untranslated RNAs that are implicated in oncogenesis. These EBER transcripts are the most highly expressed viral gene products in EBV-transformed cells. We have identified changes to the cellular transcription machinery that may contribute to the high levels of EBER RNA. These include phosphorylation of ATF2, which interacts with EBER promoters. A second is induction of TFIIIC, a pol III-specific factor that activates EBER genes; all five subunits of TFIIIC are overexpressed in EBV-positive cells. In addition, EBV induces BDP1, a subunit of the pol III-specific factor TFIIIB. Although BDP1 is the only TFIIIB subunit induced by EBV, its induction is sufficient to stimulate EBER expression in vivo, implying a limiting function. The elevated levels of BDP1 and TFIIIC in EBV-positive cells stimulate production of tRNA, 7SL, and 5S rRNA. Abnormally high expression of these cellular pol III products may contribute to the ability of EBV to enhance growth potential. PMID:16956891

  9. Tertiary-Amine Functionalized Polyplexes Enhanced Cellular Uptake and Prolonged Gene Expression

    PubMed Central

    Lo, Chia-Wen; Chang, Yung; Lee, Jyun-Lin; Tsai, Wei-Bor; Chen, Wen-Shiang

    2014-01-01

    Ultrasound (US) has been found to facilitate the transport of DNA across cell membranes. However, the transfection efficiency is generally low, and the expression duration of the transfected gene is brief. In this study, a tertiary polycation, Poly(2-(dimethylamino) ethyl methacrylate) (PDMAEMA), was used as a carrier for US-mediated gene transfection. Its in-vitro and in-vivo effects on the transfection efficiency and the expression duration were evaluated. A mixture of pCI-neo-luc and PDMAEMA was transfected to cultured cells or mouse muscle by exposure to 1-MHz pulse US. A strong expression of luciferase was found 10 days after the transfection in vitro regardless of US exposure. However, effective transfection only occurred in the US groups in vivo. The transfection ability depended on the weight ratio of PDMAEMA to DNA, and was different for the in-vitro and in-vivo conditions. Lower weight ratios, e.g., 0.25, exhibited better in-vivo expression for at least 45 days. PMID:24827929

  10. Expression of cellular homologues of retroviral onc genes in human hematopoietic cells.

    PubMed Central

    Westin, E H; Wong-Staal, F; Gelmann, E P; Dalla-Favera, R; Papas, T S; Lautenberger, J A; Eva, A; Reddy, E P; Tronick, S R; Aaronson, S A; Gallo, R C

    1982-01-01

    Total cellular poly(A)-enriched RNA from a variety of fresh human leukemic blood cells and hematopoietic cell lines was analyzed for homology with molecularly cloned DNA probes containing the onc sequence of Abelson murine leukemia virus (Ab-MuLV), Harvey murine sarcoma virus (Ha-MuSV), simian sarcoma virus (SSV), and avian myelocytomatosis virus strain MC29. Results with the fresh blood cells paralleled those obtained with the cell lines. With Ab-MuLV and Ha-MuSV, multiple RNA bands were visualized in all cell types examined without significant variation in the relative intensities of the bands. When SSV was used as the probe, expression of related onc sequences was absent in all of the hematopoietic cell types examined except for one neoplastic T-cell line (HUT 102), which produces the human T-cell leukemia (lymphoma) retrovirus HTLV. In this cell line, a single band (4.2 kilobases) was observed. With MC29 as the probe, a single band of 2.7 kilobases was visualized in all cell types examined with only a 10 to 2-fold variation in intensity of hybridization. An exception was the promyelocytic cell line, HL60, which expressed approximately 10-fold more MC29-related onc sequences. With induction of differentiation of HL60 with either dimethyl sulfoxide or retinoic acid, a marked diminution in the amount of the MC29-related, but not the Ab-MuLV-related, onc message was observed. Images PMID:6283530

  11. Genomic mapping and cellular expression of human CPG2 transcripts in the SYNE1 gene.

    PubMed

    Loebrich, Sven; Rathje, Mette; Hager, Emily; Ataman, Bulent; Harmin, David A; Greenberg, Michael E; Nedivi, Elly

    2016-03-01

    Bipolar disorder (BD) is a prevalent and severe mood disorder characterized by recurrent episodes of mania and depression. Both genetic and environmental factors have been implicated in BD etiology, but the biological underpinnings remain elusive. Recent genome-wide association studies (GWAS) for identifying genes conferring risk for schizophrenia, BD, and major depression, identified an association between single-nucleotide polymorphisms (SNPs) in the SYNE1 gene and increased risk of BD. SYNE1 has also been identified as a risk locus for multiple other neurological or neuromuscular genetic disorders. The BD associated SNPs map within the gene region homologous to part of rat Syne1 encompassing the brain specific transcripts encoding CPG2, a postsynaptic neuronal protein localized to excitatory synapses and an important regulator of glutamate receptor internalization. Here, we use RNA-seq, ChIP-seq and RACE to map the human SYNE1 transcriptome, focusing on the CPG2 locus. We validate several CPG2 transcripts, including ones not previously annotated in public databases, and identify and clone a full-length CPG2 cDNA expressed in human neocortex, hippocampus and striatum. Using lenti-viral gene knock down/replacement and surface receptor internalization assays, we demonstrate that human CPG2 protein localizes to dendritic spines in rat hippocampal neurons and is functionally equivalent to rat CPG2 in regulating glutamate receptor internalization. This study provides a valuable gene-mapping framework for relating multiple genetic disease loci in SYNE1 with their transcripts, and for evaluating the effects of missense SNPs identified by patient genome sequencing on neuronal function. PMID:26704904

  12. 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.

  13. Differential expression of calcium-related genes in gastric cancer cells transfected with cellular prion protein.

    PubMed

    Liang, Jie; Luo, Guanhong; Ning, Xiaoxuan; Shi, Yongquan; Zhai, Huihong; Sun, Shiren; Jin, Haifeng; Liu, Zhenxiong; Zhang, Faming; Lu, Yuanyuan; Zhao, Yunping; Chen, Xiong; Zhang, Hongbo; Guo, Xuegang; Wu, Kaichun; Fan, Daiming

    2007-06-01

    The prion protein (PrPC) has a primary role in the pathogenesis of transmissible spongiform encephalopathies, which causes prion disorders partially due to Ca2+ dysregulation. In our previous work, we found that overexpressed PrPC in gastric cancer was involved in apoptosis, cell proliferation, and metastasis of gastric cancer. To better understand how PrPC acts in gastric cancer, a human microarray was performed to select differentially regulated genes that correlate with the biological function of PrPC. The microarray data were analyzed and revealed 3798 genes whose expression increased at least 2-fold in gastric cancer cells transfected with PrPC. These genes encode proteins involved in several aspects of cell biology, among which, we specially detected molecules related to calcium, especially the S100 calcium-binding proteins, and found that PrPC upregulates S100A1, S100A6, S100B, and S100P but downregulates CacyBP in gastric cancer cells. We also found that intracellular Ca2+ levels in cells transfected with PrPC increased, whereas these levels decreased in knockdowns of these cells. Taken together, PrPC might increase intracellular Ca2+, partially through calcium-binding proteins, or PrPC might upregulate the expression of S100 proteins, partially through stimulating the intracellular calcium level in gastric cancer. Though the underlying mechanisms need further exploration, this study provides a new insight into the role of PrPC in gastric cancer and enriches our knowledge of prion protein. PMID:17612632

  14. Centromere protein B of African green monkey cells: gene structure, cellular expression, and centromeric localization.

    PubMed Central

    Yoda, K; Nakamura, T; Masumoto, H; Suzuki, N; Kitagawa, K; Nakano, M; Shinjo, A; Okazaki, T

    1996-01-01

    Centromere protein B (CENP-B) is a centromeric DNA-binding protein which recognizes a 17-bp sequence (CENP-B box) in human and mouse centromeric satellite DNA. The African green monkey (AGM) is phylogenetically closer to humans than mice and is known to contain large amounts of alpha-satellite DNA, but there has been no report of CENP-B boxes or CENP-B in the centromere domains of its chromosomes. To elucidate the AGM CENP-B-CENP-B box interaction, we have analyzed the gene structure, expression, biochemical properties, and centromeric localization of its CENP-B. The amino acid sequence deduced from the cloned AGM CENP-B gene was established to be highly homologous to that of human and mouse CENP-B. In particular, the DNA binding and homodimer formation domains demonstrated 100% identity to their human and mouse counterparts. Immunoblotting and DNA mobility shift analyses revealed CENP-B to be expressed in AGM cell lines. As predicted from the gene structure, the AGM CENP-B in the cell extracts exhibited the same DNA binding specificity and homodimer forming activity as human CENP-B. By indirect immunofluorescent staining of AGM mitotic cells with anti-CENP-B antibodies, a centromere-specific localization of AGM CENP-B could be demonstrated. We also isolated AGM alpha-satellite DNA with a CENP-B box-like sequence with CENP-B affinity. These results not only prove that CENP-B functionally persists in AGM cells but also suggest that the AGM genome contains the recognition sequences for CENP-B (CENP-B boxes with the core recognition sequence or CENP-B box variants) in centromeric satellite DNA. PMID:8756674

  15. Limited but durable changes to cellular gene expression in a model of latent adenovirus infection are reflected in childhood leukemic cell lines

    PubMed Central

    Ornelles, D.A.; Gooding, L.R.; Dickherber, M.L.; Policard, M.; Garnett-Benson, C.

    2016-01-01

    Mucosal lymphocytes support latent infections of species C adenoviruses. Because infected lymphocytes resist re-infection with adenovirus, we sought to identify changes in cellular gene expression that could inhibit the infectious process. The expression of over 30,000 genes was evaluated by microarray in persistently infected B-and T-lymphocytic cells. BBS9, BNIP3, BTG3, CXADR, SLFN11 and SPARCL1 were the only genes differentially expressed between mock and infected B cells. Most of these genes are associated with oncogenesis or cancer progression. Histone deacetylase and DNA methyltransferase inhibitors released the repression of some of these genes. Cellular and viral gene expression was compared among leukemic cell lines following adenovirus infection. Childhood leukemic B-cell lines resist adenovirus infection and also show reduced expression of CXADR and SPARCL. Thus adenovirus induces limited changes to infected B-cell lines that are similar to changes observed in childhood leukemic cell lines. PMID:27085068

  16. Limited but durable changes to cellular gene expression in a model of latent adenovirus infection are reflected in childhood leukemic cell lines.

    PubMed

    Ornelles, D A; Gooding, L R; Dickherber, M L; Policard, M; Garnett-Benson, C

    2016-07-01

    Mucosal lymphocytes support latent infections of species C adenoviruses. Because infected lymphocytes resist re-infection with adenovirus, we sought to identify changes in cellular gene expression that could inhibit the infectious process. The expression of over 30,000 genes was evaluated by microarray in persistently infected B-and T-lymphocytic cells. BBS9, BNIP3, BTG3, CXADR, SLFN11 and SPARCL1 were the only genes differentially expressed between mock and infected B cells. Most of these genes are associated with oncogenesis or cancer progression. Histone deacetylase and DNA methyltransferase inhibitors released the repression of some of these genes. Cellular and viral gene expression was compared among leukemic cell lines following adenovirus infection. Childhood leukemic B-cell lines resist adenovirus infection and also show reduced expression of CXADR and SPARCL. Thus adenovirus induces limited changes to infected B-cell lines that are similar to changes observed in childhood leukemic cell lines. PMID:27085068

  17. Functional expression and cellular distribution of diastrophic dysplasia sulfate transporter (DTDST) gene mutations in HEK cells.

    PubMed

    Karniski, Lawrence P

    2004-10-01

    Defects in sulfate transport in chondrocytes lead to undersulfation of the cartilage extracellular matrix proteoglycans. Mutations in the diastrophic dysplasia sulfate transporter (DTDST) gene have been linked to four chondrodysplasias of varying severity. To characterize disease-causing mutations of DTDST, we expressed DTDST-mediated sulfate transport in mammalian HEK-293 cells and determined that the wild-type protein is glycosylated and localized to the cell plasma membrane. Four mutations, A715V, C653S, Q454P and R279W, stimulated sulfate transport at rates only 39-62% of wild-type DTDST. These four mutations were expressed on the plasma membrane of the cell, but the amount of expressed protein was reduced when compared with wild-type DTDST. The Q454P mutant is unique in that it is not properly glycosylated in HEK cells. There was no difference in sulfate transport activity between cells transfected with either the DeltaV340 or the G678V mutations and control HEK cells. Furthermore, the G678V mutation is not expressed along the plasma membrane, but is trapped within the cytoplasm. When comparing the sulfate transport capacity of each DTDST mutation with the chondrodysplasia in which it has been identified, we find that individuals with severe achondrogenesis 1B phenotype have null mutations on both DTDST alleles. Heterozygotes for both a null mutation and a partial-function mutation result in either atelosteogenesis type 2 or DTD, whereas the milder, recessive multiple epiphyseal dysplasia phenotype is homozygous for partial-function mutations. In contrast to previous studies in Xenopus laevis oocytes, we find a strong correlation between the severity of the phenotype and the level of residual transport function in mammalian cells. PMID:15294877

  18. Expression of a cellular gene cloned in herpes simplex virus: rabbit beta-globin is regulated as an early viral gene in infected fibroblasts.

    PubMed Central

    Smiley, J R; Smibert, C; Everett, R D

    1987-01-01

    We constructed nondefective herpes simplex virus type 1 recombinants bearing the intact rabbit beta-globin gene inserted into the viral gene for thymidine kinase to study the expression of a cellular gene when it is present in the viral genome during lytic viral infections. The globin promoter was activated to high levels during productive infection of Vero cells, giving rise to properly spliced and processed cytoplasmic globin transcripts. Expression of globin RNA occurred with early kinetics, was not affected by blocking viral DNA replication, and was strongly inhibited by preventing viral immediate-early protein synthesis with cycloheximide. These results support the hypothesis that temporal control of herpes simplex virus early gene expression is accomplished by mechanisms that are not restricted to viral promoters. In addition, these data show that a cellular transcript can be correctly processed and can accumulate to high levels during viral infection; this indicates that the mechanisms of virally induced shutoff of host RNA accumulation and degradation of host mRNAs do not depend on sequence-specific differentiation between host and viral RNAs. These findings also suggest that herpesviruses have considerable potential as high-capacity gene transfer vectors for a variety of applications. Images PMID:3037101

  19. Stage specific gene expression and cellular localization of two isoforms of the serine hydroxymethyltransferase in the protozoan parasite Leishmania.

    PubMed

    Gagnon, Dominic; Foucher, Aude; Girard, Isabelle; Ouellette, Marc

    2006-11-01

    Serine hydroxymethyltransferase (SHMT) catalyses the reversible conversion of serine and tetrahydrofolate to glycine and methylene-tetrahydrofolate. The recent completion of the genome sequence of Leishmania major revealed the presence of two genes coding for two isoforms of this protein. In silico analysis showed that one isoform had an extension at its N-terminus and was predicted to localize to the mitochondrion. The situation is different in other kinetoplastid parasites with only one SHMT encoding gene in Trypanosoma cruzi and no SHMT encoding gene in Trypanosoma brucei. The two L. major SHMT genes were cloned in frame with the green fluorescent protein and the resulting fusion proteins showed differential localization: the short form (SHMT-S) was found in the cytosol while the long one (SHMT-L) was found in an organelle that has hallmarks of the parasite mitochondrion. Indeed, SHMT-L had a similar cellular fractionation pattern as the mitochondrial HSP60 as determined by digitonin fractionation. Both SHMT-S and SHMT-L genes were expressed preferentially in the amastigote stage of the parasite and the RNA levels of SHMT-L could be modulated by glycine, serine, and folate. Overexpression of SHMT-S increased resistance to the antifolate methotrexate and to a lower level to the inhibitor thiosemicarbazide in a rich folate containing medium. These findings suggest that folate metabolism is compartmentalised in Leishmania and that SHMT RNA levels are responsive to environmental conditions. PMID:16876889

  20. 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

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

    PubMed

    Wasfi, Reham; Elkhatib, Walid F; Khairalla, Ahmed S

    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

  2. Transcriptional and post-transcriptional regulation of HIV-1 gene expression: role of cellular factors for Tat and Rev.

    PubMed

    Nekhai, Sergei; Jeang, Kuan-Teh

    2006-12-01

    The emergence of drug-resistant HIV-1 strains presents a challenge for the design of new therapy. Targeting host cell factors that regulate HIV-1 replication might be one way to overcome the propensity for HIV-1 to mutate in order to develop resistance to antivirals. This article reviews the interplay between viral proteins Tat and Rev and their cellular cofactors in the transcriptional and post-transcriptional regulation of HIV-1 gene expression. HIV-1 Tat regulates viral transcription by recruiting cellular factors to the HIV promoter. Tat interacts with protein kinase complexes Cdk9/cyclin T1 and Cdk2/cyclin E; acetyltransferases p300/CBP, p300/CBP-associated factor and hGCN5; protein phosphatases and other factors. HIV-1 Rev regulates post-transcriptional processing of viral mRNAs. Rev primarily functions to export unspliced and partially spliced viral RNAs from the nucleus into the cytoplasm. For this activity, Rev cooperates with cellular transport protein CRM1 and RNA helicases DDX1 and DDX3, amongst others. PMID:17661632

  3. Human T lymphotropic virus type-1 p30II alters cellular gene expression to selectively enhance signaling pathways that activate T lymphocytes

    PubMed Central

    Michael, Bindhu; Nair, Amrithraj M; Hiraragi, Hajime; Shen, Lei; Feuer, Gerold; Boris-Lawrie, Kathleen; Lairmore, Michael D

    2004-01-01

    Background Human T-lymphotropic virus type-1 (HTLV-1) is a deltaretrovirus that causes adult T-cell leukemia/lymphoma and is implicated in a variety of lymphocyte-mediated disorders. HTLV-1 contains both regulatory and accessory genes in four pX open reading frames. pX ORF-II encodes two proteins, p13II and p30II, which are incompletely defined in the virus life cycle or HTLV-1 pathogenesis. Proviral clones of the virus with pX ORF-II mutations diminish the ability of the virus to maintain viral loads in vivo. Exogenous expression of p30II differentially modulates CREB and Tax-responsive element-mediated transcription through its interaction with CREB-binding protein/p300 and represses tax/rex RNA nuclear export. Results Herein, we further characterized the role of p30II in regulation of cellular gene expression, using stable p30II expression system employing lentiviral vectors to test cellular gene expression with Affymetrix U133A arrays, representing ~33,000 human genes. Reporter assays in Jurkat T cells and RT-PCR in Jurkat and primary CD4+ T-lymphocytes were used to confirm selected gene expression patterns. Our data reveals alterations of interrelated pathways of cell proliferation, T-cell signaling, apoptosis and cell cycle in p30II expressing Jurkat T cells. In all categories, p30II appeared to be an overall repressor of cellular gene expression, while selectively increasing the expression of certain key regulatory genes. Conclusions We are the first to demonstrate that p30II, while repressing the expression of many genes, selectively activates key gene pathways involved in T-cell signaling/activation. Collectively, our data suggests that this complex retrovirus, associated with lymphoproliferative diseases, relies upon accessory gene products to modify cellular environment to promote clonal expansion of the virus genome and thus maintain proviral loads in vivo. PMID:15560845

  4. 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. PMID:26494614

  5. Development and Characterization of Novel Empty Adenovirus Capsids and Their Impact on Cellular Gene Expression

    PubMed Central

    Stilwell, Jackie L.; McCarty, Douglas M.; Negishi, Atsuko; Superfine, Richard; Samulski, R. Jude

    2003-01-01

    Adenovirus (Ad) has been extensively studied as a eukaryotic viral vector. As these vectors have evolved from first-generation vectors to vectors that contain either very few or no viral genes (“gutless” Ad), significant reductions in the host innate immune response upon infection have been observed. Regardless of these vector improvements an unknown amount of toxicity has been associated with the virion structural proteins. Here we demonstrate the ability to generate high particle numbers (1011 to 1012) of Ad empty virions based on a modification of Cre/lox gutless Ad vectors. Using a battery of analyses (electron microscopy, atomic force microscopy, confocal images, and competition assays) we characterized this reagent and determined that it (i) makes intact virion particles, (ii) competes for receptor binding with wild-type Ad, and (iii) enters the cell proficiently, demonstrating an ability to carry out essential steps of viral entry. To further study the biological impact of these Ad empty virions on infected cells, we carried out DNA microarray analysis. Compared to that for recombinant Ad, the number of mRNAs modulated upon infection was significantly reduced but the expression signatures were similar. This reagent provides a valuable tool for studies of Ad in that researchers can examine the effect of infection in the presence of the virion capsid alone. PMID:14610209

  6. Dietary restriction mitigates cocaine-induced alterations of olfactory bulb cellular plasticity and gene expression, and behavior.

    PubMed

    Xu, Xiangru; Mughal, Mohamed R; Scott Hall, F; 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

    2010-07-01

    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 with mice fed ad libitum. Levels of dopamine and its metabolites in the olfactory bulb (OB) were suppressed in mice on the ADF diet compared with 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, 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 cAMP-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

  7. Quantitative in situ hybridization for the study of gene expression at the regional and cellular levels.

    PubMed

    Le Moine, Catherine

    2003-08-01

    Quantitative in situ hybridization allows measurement of mRNA level modifications in a variety of experimental conditions. This analysis may be performed both at the regional anatomical and cellular levels by densitometry, neuronal counting and silver grain measurements. PMID:18428577

  8. 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

  9. Interactions between small viral RNAs of vesicular stomatitis virus and components of cellular gene expression.

    PubMed

    Keene, J D

    1985-05-01

    Recent interest in the details of virus-host interactions has come to focus on molecular contacts between cell factors and components of viruses. These generally concern protein-protein and protein-nucleic acid interactions. In this review, protein-nucleic acid interactions involving viral transcription products and cell proteins are considered. Also examined here will be the hypothesis that such interactions have evolved because viruses have adopted cellular processes to favour their own replication and that the consequences of this co-evolution can be the disruption of the macromolecular functions of the cell, and eventual cytopathology. For its own survival in the long term, the host may evolve more refined mechanisms to evade the damage levied by the intruding virus. PMID:2856377

  10. 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. PMID:26223179

  11. Hormone-regulated v-rel estrogen receptor fusion protein: reversible induction of cell transformation and cellular gene expression.

    PubMed

    Boehmelt, G; Walker, A; Kabrun, N; Mellitzer, G; Beug, H; Zenke, M; Enrietto, P J

    1992-12-01

    We describe the construction of a v-rel estrogen receptor fusion protein (v-relER) which allows the regulation of v-rel oncoprotein activity by hormone. In the presence of estrogen, v-relER readily transformed primary chicken fibroblasts and bone marrow cells in vitro. In both cell types, v-rel-specific transformation was critically dependent on the presence of estrogen or the estrogen agonist 4-hydroxytamoxifen (OHT). Withdrawal of estrogen or application of an estrogen antagonist, ICI164,384 (ICI) caused a reversal of the transformed phenotype. We also demonstrate that the v-relER protein binds to NF-kappa B sites in an estrogen-dependent manner, thereby showing that sequence-specific DNA binding of v-relER is critical for the activation of its transforming capacity. In transient transfection experiments, we failed to demonstrate a clear repressor or activator function of the v-rel moiety in v-relER. However, in v-relER-transformed bone marrow cells, estrogen and OHT induced elevated mRNA levels of two cellular genes whose expression is constitutive and high in v-rel-transformed cells. These results suggest that v-rel might exert part of its activity as an activator of rel-responsive genes. PMID:1425595

  12. Effect of human immunodeficiency virus type 1 protein R (vpr) gene expression on basic cellular function of fission yeast Schizosaccharomyces pombe.

    PubMed Central

    Zhao, Y; Cao, J; O'Gorman, M R; Yu, M; Yogev, R

    1996-01-01

    The human immunodeficiency virus type 1 (HIV-1) Vpr protein affects cell morphology and prevents proliferation of human cells by induction of cell cycle G2 arrest. In this study, we used the fission yeast Schizosaccharomyces pombe as a model system to investigate the cellular effects of HIV-1 vpr gene expression. The vpr gene was cloned into an inducible fission yeast gene expression vector and expressed in wild-type S. pombe cells, and using these cells, we were able to demonstrate the specific Vpr-induced effects by induction and suppression of vpr gene expression. Induction of HIV-1 vpr gene expression affected S. pombe at the colonial, cellular, and molecular levels. Specifically, Vpr induced small-colony formation, polymorphic cells, growth delay, and cell cycle G2 arrest. Additionally, Vpr-induced G2 arrest appeared to be independent of cell size and morphological changes. The cell cycle G2 arrest correlated with increased phosphorylation of p34cdc2, suggesting negative regulation of mitosis by HIV-1 Vpr. Treatment of Vpr-induced cell with a protein phosphatase inhibitor, okadaic acid, transiently suppressed cell cycle arrest and morphological changes. This observation implicates possible involvement of protein phosphatase(s) in the effects of Vpr. Together, these data showed that the HIV-1 Vpr-induced cellular changes in S. pombe are similar to those observed in human cells. Therefore, the S. pombe system is suited for further investigation of the HIV-1 vpr gene functions. PMID:8709199

  13. Temporal sequence and cellular origin of interleukin-2 stimulated cytokine gene expression.

    PubMed Central

    Saraya, K. A.; Balkwill, F. R.

    1993-01-01

    A study of activation of the cytokine network by interleukin 2, IL-2, may provide a rationale for devising cytokine combination and cytokine antagonist treatments with increased anti-tumour efficacy and decreased toxicity. We have investigated the expression of mRNA for 13 cytokines and three transcription factors during in vitro culture of peripheral blood mononuclear cells, PBMC, with IL-2. A consistent pattern of induction was seen in nine individuals, with early (2-24 h) induction of IL-1 beta, IL-6, tumour necrosis factor, TNF, lymphotoxin, LT, and gro. TNF and LT mRNA was expressed continually throughout culture, but levels of mRNA for IL-1 beta, IL-6, and gro declined by 24-48 h. After 48 h, PBMC began to express mRNA for IFN-gamma, IL-5, GM-CSF, and M-CSF. At 15 min to 1 h post IL-2 mRNA for c-fos, c-jun, and c-myc, and TNF was induced in three individuals studied. IL-4, IFN-alpha, and IL-1 alpha mRNA was not detected. Only a minority of cells expressed mRNA for TNF, IL-1 beta, IL-6 and IFN-gamma, and monocytes were the main source. Levels of cytokine protein in culture supernatants mirrored the pattern of mRNA induction. This in vitro model shows clear parallels with the reported in vivo production of cytokines during IL-2 therapy, and may prove useful in designing new therapeutic strategies. Images Figure 1 Figure 2 Figure 3 PMID:8439502

  14. Gene Expression Profile Changes and Cellular Responses to Bleomycin-Induced DNA Damage in Human Fibroblast Cells in Space

    NASA Technical Reports Server (NTRS)

    Lu, Tao; Zhang, Ye; Kidane, Yared; Feiveson, Alan; Stodieck, Louis; Karouia, Fathi; Rohde, Larry; Wu, Honglu

    2016-01-01

    Living organisms are constantly exposed to space radiation that consists of energetic protons and other heavier charged particles. In addition, DNA in space can be damaged by 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 affects the accuracy of the radiation risk assessment for astronauts and the mutation rate in microorganisms. Although possible synergistic effects of space radiation and microgravity have been investigated since the early days of the human space program, the published results were mostly conflicting and inconsistent. To investigate the effects of spaceflight on cellular responses to DNA damage, confluent human fibroblast cells (AG1522) flown on the International Space Station (ISS) were treated with bleomycin for three hours in the true microgravity environment, which induced DNA damages including double-strand breaks (DSB). Damages in the DNA were quantified by immunofluorescence staining for ?-H2AX, which showed similar percentages of different types of stained cells between flight and ground. However, there was a slight shift in the distribution of the ?-H2AX foci number in the flown cells with countable 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. A microarray analysis of gene expressions in response to bleomycin treatment was also performed. Comparison of the responsive pathways between the flown and ground cells showed similar responses with the p53 network being the top upstream regulator. Similar responses at the RNA level between different gravity conditions were also observed with a PCR array analysis containing a set of genes involved in DNA damage signaling; with BBC3, CDKN1A, PCNA and PPM1D being significantly

  15. 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.

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

    PubMed

    Maningat, Patricia D; Sen, Partha; Rijnkels, Monique; Hadsell, Darryl L; Bray, Molly S; Haymond, Morey W

    2011-04-27

    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 from five normal lactating women. Following 24 h of baseline milk and blood sampling, rhGH (0.1 mg/kg/day) was administered subcutaneously once daily for 3 days. Gene expression changes were determined by microarray studies utilizing milk fat globule RNA isolated from each milk sample. Following rhGH administration, DNA synthesis and cell cycle genes were induced, while no significant changes were observed in the expression of milk synthesis genes. Expression of glycolysis and citric acid cycle genes were increased by day 4 compared with day 1, while lipid synthesis genes displayed a circadian-like pattern. Cell cycle gene upregulation occurred after a lag of ∼2 days, likely explaining the failure to increase milk production after only 3 days of rhGH treatment. We conclude that rhGH induces expression of cellular proliferation and metabolism genes but does not induce milk protein gene expression, as potential mechanisms for increasing milk production and could account for the known effect of rhGH to increase milk production following 7-10 days. PMID:21205870

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

    PubMed Central

    Maningat, Patricia D.; Sen, Partha; Rijnkels, Monique; Hadsell, Darryl L.; Bray, Molly S.

    2011-01-01

    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 from five normal lactating women. Following 24 h of baseline milk and blood sampling, rhGH (0.1 mg/kg/day) was administered subcutaneously once daily for 3 days. Gene expression changes were determined by microarray studies utilizing milk fat globule RNA isolated from each milk sample. Following rhGH administration, DNA synthesis and cell cycle genes were induced, while no significant changes were observed in the expression of milk synthesis genes. Expression of glycolysis and citric acid cycle genes were increased by day 4 compared with day 1, while lipid synthesis genes displayed a circadian-like pattern. Cell cycle gene upregulation occurred after a lag of ∼2 days, likely explaining the failure to increase milk production after only 3 days of rhGH treatment. We conclude that rhGH induces expression of cellular proliferation and metabolism genes but does not induce milk protein gene expression, as potential mechanisms for increasing milk production and could account for the known effect of rhGH to increase milk production following 7–10 days. PMID:21205870

  18. GENE EXPRESSION NETWORKS

    EPA Science Inventory

    "Gene expression network" is the term used to describe the interplay, simple or complex, between two or more gene products in performing a specific cellular function. Although the delineation of such networks is complicated by the existence of multiple and subtle types of intera...

  19. Role of Accessory Proteins of HTLV-1 in Viral Replication, T Cell Activation, and Cellular Gene Expression

    PubMed Central

    Michael, Bindhu; Nair, Amithraj; Lairmore, Michael D.

    2010-01-01

    Human T-cell lymphotropic virus type 1 (HTLV-1), causes adult T cell leukemia/lymphoma (ATLL), and initiates a variety of immune mediated disorders. The viral genome encodes common structural and enzymatic proteins characteristic of all retroviruses and utilizes alternative splicing and alternate codon usage to make several regulatory and accessory proteins encoded in the pX region (pX ORF I to IV). Recent studies indicate that the accessory proteins p12I, p27I, p13II, and p30II, encoded by pX ORF I and II, contribute to viral replication and the ability of the virus to maintain typical in vivo expression levels. Proviral clones that are mutated in either pX ORF I or II, while fully competent in cell culture, are severely limited in their replicative capacity in a rabbit model. These HTLV-1 accessory proteins are critical for establishment of viral infectivity, enhance T- lymphocyte activation and potentially alter gene transcription and mitochondrial function. HTLV-1 pX ORF I expression is critical to the viral infectivity in resting primary lymphocytes suggesting a role for the calcineurin-binding protein p12I in lymphocyte activation. The endoplasmic reticulum and cis-Golgi localizing p12I activates NFAT, a key T cell transcription factor, through calcium-mediated signaling pathways and may lower the threshold of lymphocyte activation via the JAK/STAT pathway. In contrast p30II localizes to the nucleus and represses viral promoter activity, but may regulate cellular gene expression through p300/CBP or related co-activators of transcription. The mitochondrial localizing p13II induces morphologic changes in the organelle and may influence energy metabolism infected cells. Future studies of the molecular details HTLV-1 “accessory” proteins interactions will provide important new directions for investigations of HTLV-1 and related viruses associated with lymphoproliferative diseases. Thus, the accessory proteins of HTLV-1, once thought to be dispensable for

  20. 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. PMID:25786607

  1. 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

  2. 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

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

    2013-10-01

    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

  3. Reconstructing and analysing cellular states, space and time from gene expression profiles of many cells and single cells.

    PubMed

    Francesconi, Mirko; Lehner, Ben

    2015-10-01

    Genome-wide gene expression profiling is a fast, cheap and standardised analysis that provides a high dimensional measurement of the state of a biological sample. In this review we describe computational methods that can be applied to identify and interpret sources of variance in gene expression in whole organisms, organs, tissues or single cells. This allows the identification of constituent cell types and states in complex mixtures, the reconstruction of temporal trajectories of development, differentiation and progression, and the reconstruction of spatial patterning. When applied to genetically variable samples, these methods allow the efficient investigation of how genetic variation influences gene expression and biological processes in space and time. PMID:26273952

  4. 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.

  5. Inhibition of cellular proliferation by the Wilms' tumor suppressor WT1 is associated with suppression of insulin-like growth factor I receptor gene expression.

    PubMed Central

    Werner, H; Shen-Orr, Z; Rauscher, F J; Morris, J F; Roberts, C T; LeRoith, D

    1995-01-01

    We have investigated the regulation of the insulin-like growth factor I receptor (IGF-I-R) gene promoter by the Wilms' tumor suppressor WT1 in intact cells. The levels of endogenous IGF-I-R mRNA and the activity of IGF-I-R gene promoter fragments in luciferase reporter constructs were found to be significantly higher in G401 cells (a Wilms' tumor-derived cell line lacking detectable WT1 mRNA) than in 293 cells (a human embryonic kidney cell line which expresses significant levels of WT1 mRNA). To study whether WT1 could suppress the expression of the endogenous IGF-I-R gene, WT1-negative G401 cells were stably transfected with a WT1 expression vector. Expression of WT1 mRNA in G401 cells resulted in a significant decrease in the rate of cellular proliferation, which was associated with a reduction in the levels of IGF-I-R mRNA, promoter activity, and ligand binding and with a reduction in IGF-I-stimulated cellular proliferation, thymidine incorporation, and anchorage-independent growth. These data suggest that a major aspect of the action of the WT1 tumor suppressor is the repression of IGF-I-R gene expression. PMID:7791758

  6. Blood cell gene expression associated with cellular stress defense is modulated by antioxidant-rich food in a randomised controlled clinical trial of male smokers

    PubMed Central

    2010-01-01

    Background Plant-based diets rich in fruit and vegetables can prevent development of several chronic age-related diseases. However, the mechanisms behind this protective effect are not elucidated. We have tested the hypothesis that intake of antioxidant-rich foods can affect groups of genes associated with cellular stress defence in human blood cells. Trial registration number: NCT00520819 http://clinicaltrials.gov. Methods In an 8-week dietary intervention study, 102 healthy male smokers were randomised to either a diet rich in various antioxidant-rich foods, a kiwifruit diet (three kiwifruits/d added to the regular diet) or a control group. Blood cell gene expression profiles were obtained from 10 randomly selected individuals of each group. Diet-induced changes on gene expression were compared to controls using a novel application of the gene set enrichment analysis (GSEA) on transcription profiles obtained using Affymetrix HG-U133-Plus 2.0 whole genome arrays. Results Changes were observed in the blood cell gene expression profiles in both intervention groups when compared to the control group. Groups of genes involved in regulation of cellular stress defence, such as DNA repair, apoptosis and hypoxia, were significantly upregulated (GSEA, FDR q-values < 5%) by both diets compared to the control group. Genes with common regulatory motifs for aryl hydrocarbon receptor (AhR) and AhR nuclear translocator (AhR/ARNT) were upregulated by both interventions (FDR q-values < 5%). Plasma antioxidant biomarkers (polyphenols/carotenoids) increased in both groups. Conclusions The observed changes in the blood cell gene expression profiles suggest that the beneficial effects of a plant-based diet on human health may be mediated through optimization of defence processes. PMID:20846424

  7. 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.

  8. High-resolution gene expression analysis of the developing mouse kidney defines novel cellular compartments within the nephron progenitor population.

    PubMed

    Mugford, Joshua W; Yu, Jing; Kobayashi, Akio; McMahon, Andrew P

    2009-09-15

    The functional unit of the kidney is the nephron. During its organogenesis, the mammalian metanephric kidney generates thousands of nephrons over a protracted period of fetal life. All nephrons are derived from a population of self-renewing multi-potent progenitor cells, termed the cap mesenchyme. However, our understanding of the molecular and cellular mechanisms underlying nephron development is at an early stage. In order to identify factors involved in nephrogenesis, we performed a high-resolution, spatial profiling of a number of transcriptional regulators expressed within the cap mesenchyme and early developing nephron. Our results demonstrate novel, stereotypic, spatially defined cellular sub-domains within the cap mesenchyme, which may, in part, reflect induction of nephron precursors. These results suggest a hitherto unappreciated complexity of cell states that accompany the assembly of the metanephric kidney, likely reflecting diverse regulatory actions such as the maintenance and induction of nephron progenitors. PMID:19591821

  9. High-resolution gene expression analysis of the developing mouse kidney defines novel cellular compartments within the nephron progenitor population

    PubMed Central

    Mugford, Joshua W.; Yu, Jing; Kobayashi, Akio; McMahon, Andrew P.

    2009-01-01

    The functional unit of the kidney is the nephron. During its organogenesis, the mammalian metanephric kidney generates thousands of nephrons over a protracted period of fetal life. All nephrons are derived from a population of self-renewing multi-potent progenitor cells, termed the cap mesenchyme. However, our understanding of the molecular and cellular mechanisms underlying nephron development is at an early stage. In order to identify factors involved in nephrogenesis, we performed a high-resolution, spatial profiling of a number of transcriptional regulators expressed within the cap mesenchyme and early developing nephron. Our results demonstrate novel, stereotypic, spatially defined cellular sub-domains within the cap mesenchyme, which may, in part, reflect induction of nephron precursors. These results suggest a hitherto unappreciated complexity of cell states that accompany the assembly of the metanephric kidney, likely reflecting diverse regulatory actions such as the maintenance and induction of nephron progenitors. PMID:19591821

  10. Evaluation of cellular uptake and intracellular trafficking as determining factors of gene expression for amino acid-substituted gemini surfactant-based DNA nanoparticles

    PubMed Central

    2012-01-01

    Background Gene transfer using non-viral vectors offers a non-immunogenic and safe method of gene delivery. Cellular uptake and intracellular trafficking of the nanoparticles can impact on the transfection efficiency of these vectors. Therefore, understanding the physicochemical properties that may influence the cellular uptake and the intracellular trafficking can aid the design of more efficient non-viral gene delivery systems. Recently, we developed novel amino acid-substituted gemini surfactants that showed higher transfection efficiency than their parent compound. In this study, we evaluated the mechanism of cellular uptake of the plasmid/gemini surfactant/helper lipid nanoparticles and their effect on the transfection efficiency. Results Nanoparticles were incubated with Sf 1 Ep cells in the presence of different endocytic inhibitors and gene expression (interferon-γ) was measured using ELISA. Clathrin-mediated and caveolae-mediated uptake were found to be equally contributing to cellular internalization of both P/12-7NH-12/L (parent gemini surfactant) and P/12-7NGK-12/L (amino acid-substituted gemini surfactant) nanoparticles. The plasmid and the helper lipid were fluorescently tagged to track the nanoparticles inside the cells, using confocal laser scanning microscopy. Transmission electron microscopy images showed that the P/12-7NGK-12/L particles were cylindrical while the P/12-7NH-12/L particles were spherical which may influence the cellular uptake behaviour of these particles. Dye exclusion assay and pH-titration of the nanoparticles suggested that high buffering capacity, pH-dependent increase in particle size and balanced DNA binding properties may be contributing to a more efficient endosomal escape of P/12-7NGK-12/L compared to the P/12-7NH-12/L nanoparticles, leading to higher gene expression. Conclusion Amino-acid substitution in the spacer of gemini surfactant did not alter the cellular uptake pathway, showing similar pattern to the

  11. 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...

  12. De-regulation of gene expression and alternative splicing affects distinct cellular pathways in the aging hippocampus

    PubMed Central

    Stilling, Roman M.; Benito, Eva; Gertig, Michael; Barth, Jonas; Capece, Vincenzo; Burkhardt, Susanne; Bonn, Stefan; Fischer, Andre

    2014-01-01

    Aging is accompanied by gradually increasing impairment of cognitive abilities and constitutes the main risk factor of neurodegenerative conditions like Alzheimer's disease (AD). The underlying mechanisms are however not well understood. Here we analyze the hippocampal transcriptome of young adult mice and two groups of mice at advanced age using RNA sequencing. This approach enabled us to test differential expression of coding and non-coding transcripts, as well as differential splicing and RNA editing. We report a specific age-associated gene expression signature that is associated with major genetic risk factors for late-onset AD (LOAD). This signature is dominated by neuroinflammatory processes, specifically activation of the complement system at the level of increased gene expression, while de-regulation of neuronal plasticity appears to be mediated by compromised RNA splicing. PMID:25431548

  13. 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.

  14. Molecular and cellular effects of cis-9, trans-11-conjugated linoleic acid in enterocytes: effects on proliferation, differentiation, and gene expression.

    PubMed

    Lampen, A; Leifheit, M; Voss, J; Nau, H

    2005-06-15

    It has been hypothesized that dietary conjugated linoleic acids (CLA) may inhibit colon tumorigenesis. The aim of our study was to investigate the cellular and molecular effects of cis-9 (9Z), trans-11 (11E)-CLA on the proliferation, differentiation, interaction with peroxisome proliferator-activated receptors (PPARs), and expression of genes relevant in the APC-beta-catenin-TCF4 signalling pathway in human HT-29 and Caco-2 colon cells. We found that 9Z,11E-CLA inhibited the proliferation of HT-29 and Caco-2 cells. Trans-vaccenic acid (VA) showed no antiproliferative effects at all. We determined that 9Z,11E-CLA induced cell differentiation as measured by intestinal alkaline phosphatase (IAP) enzyme activity in Caco-2 cells, mRNA expression of IAP, and activation of a 5' flanking region of IAP. The 9Z,11E-CLA activated human PPARdelta as measured in a reporter gene assay. Treatment of HT29 cells in the poliferation phase with 9Z,11E-CLA repressed mRNA-expression of proliferation genes such as c-myc, cyclin D1 and c-jun in a concentration dependent manner. The promoter activities of c-myc and AP1 were also inhibited after incubation with 9Z,11E-CLA. beta-Catenin mRNA and protein expression was also repressed by the treatment with 9Z,11E-CLA. In addition, the mRNA expression of PPARdelta was repressed by treatment of the HT-29 cells with 9Z,11E-CLA. We conclude that 9Z,11E-CLA has an antiproliferative effect at the cellular and molecular levels in human colon cells. The results indicate that the preventive effects of CLA in the development of colon cancer may be due to their downregulation of some target genes of the APC-beta-catenin-TCF-4- and PPARdelta signalling pathway. PMID:15935729

  15. Host cellular microRNA involvement in the control of hepatitis B virus gene expression and replication

    PubMed Central

    Mizuguchi, Yoshiaki; Takizawa, Toshihiro; Uchida, Eiji

    2015-01-01

    A large number of studies have demonstrated that the synergistic collaboration of a number of microRNAs (miRNAs), their growth factors and their downstream agents is required for the initiation and completion of pathogenesis in the liver. miRNAs are thought to exert a profound effect on almost every aspect of liver biology and pathology. Accumulating evidence indicates that several miRNAs are involved in the hepatitis B virus (HBV) life cycle and infectivity, in addition to HBV-associated liver diseases including fibrosis, cirrhosis and hepatocellular carcinoma (HCC). In turn, HBV can modulate the expression of several cellular miRNAs, thus promoting a favorable environment for its replication and survival. In this review, we focused on the involvement of host cellular miRNAs that are directly and indirectly associated with HBV RNA or HBV associated transcription factors. Exploring different facets of the interactions among miRNA, HBV and HCV infections, and the carcinogenesis and progress of HCC, could facilitate the development of novel and effective treatment approaches for liver disease. PMID:25866606

  16. Cooperative activation of human papillomavirus type 8 gene expression by the E2 protein and the cellular coactivator p300.

    PubMed

    Müller, Andreas; Ritzkowsky, Andreas; Steger, Gertrud

    2002-11-01

    The E2 proteins of papillomaviruses (PV) bind to the coactivator CBP/p300 as do many other transcription factors, but the precise role of CBP/p300 in E2-specific functions is not yet understood. We show that the E2 protein of human PV type 8 (HPV8) directly binds to p300. Activation of HPV8 gene expression by low amounts of HPV8 E2 was stimulated up to sevenfold by coexpression of p300. The interaction between E2 and p300 may play a role in differentiation-dependent activation of PV gene expression, since we can show that the expression level of p300 increases during keratinocyte differentiation. Surprisingly, sequence-specific binding of E2 to its recognition sites within the regulatory region of HPV8 is not necessary for this cooperation, indicating that E2 can be recruited to the promoter via protein-protein interaction. HPV8 E2 binds via its N-terminal activation domain (AD), its C-terminal DNA binding domain (DBD), and its internal hinge region to p300 in vitro. Transient-transfection assays revealed that the AD is necessary and sufficient for cooperative activation with p300. However, we provide evidence that the interaction of the hinge and the DBD of HPV8 E2 with p300 may contribute. Our data suggest an important role of p300 in regulation of HPV8 gene expression and reveal a new mechanism by which E2 may be recruited to a promoter to activate transcription without sequence specific DNA binding. PMID:12368347

  17. 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. PMID:22773133

  18. 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. PMID:23946774

  19. 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

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

    PubMed

    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 (14)C palmitate/oleate and (3)H20, 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

  1. Alpha2-macroglobulin from an Atlantic shrimp: biochemical characterization, sub-cellular localization and gene expression upon fungal challenge.

    PubMed

    Perazzolo, Luciane Maria; Bachère, Evelyne; Rosa, Rafael Diego; Goncalves, Priscila; Andreatta, Edemar Roberto; Daffre, Sirlei; Barracco, Margherita Anna

    2011-12-01

    In this study, we report on the isolation and characterization of an alpha2-macroglobulin (α2M) from the plasma of the pink shrimp Farfantepenaeus paulensis, its sub-cellular localization and transcriptional changes after infection by fungi. The molecular mass of the α2M was estimated at 389 kDa by gel filtration and 197 kDa by SDS-PAGE, under reducing conditions, suggesting that α2M from F. paulensis consists of two identical sub-units, covalently linked by disulphide bonds. The N-terminal amino acid sequence of the α2M from F. paulensis was very similar to those of other penaeid shrimps, crayfish and lobster (70-90% identity) and to a less extent with that of freshwater prawn (40% identity). A monoclonal antibody raised against the Marsupenaeus japonicus α2M made it possible to demonstrate that α2M of F. paulensis is stored in the vesicles of the shrimp granular hemocytes (through immunogold assay). Quantitative real-time PCR (qPCR) analysis showed that α2M mRNA transcripts significantly increased 24 h after an experimental infection with the shrimp pathogen Fusarium solani and it returned to the basal levels at 48 h post-injection. This is the first report on a α2M characterization in an Atlantic penaeid species and its expression profile upon a fungal infection. PMID:21888978

  2. Cellular and molecular changes associated with competence acquisition during passion fruit somatic embryogenesis: ultrastructural characterization and analysis of SERK gene expression.

    PubMed

    Rocha, Diego Ismael; Pinto, Daniela Lopes Paim; Vieira, Lorena Melo; Tanaka, Francisco André Ossamu; Dornelas, Marcelo Carnier; Otoni, Wagner Campos

    2016-03-01

    The integration of cellular and molecular data is essential for understanding the mechanisms involved in the acquisition of competence by plant somatic cells and the cytological changes that underlie this process. In the present study, we investigated the dynamics and fate of Passiflora edulis Sims cotyledon explants that were committed to somatic embryogenesis by characterizing the associated ultrastructural events and analysing the expression of a putative P. edulis ortholog of the Somatic Embryogenesis Receptor-like Kinase (SERK) gene. Embryogenic calli were obtained from zygotic embryo explants cultured on Murashige and Skoog medium supplemented with 2,4-dichlorophenoxyacetic acid and 6-benzyladenine. Callus formation was initiated by the division of cells derived from the protodermal and subprotodermal cells on the abaxial side of the cotyledons. The isodiametric protodermal cells of the cotyledon explants adopted a columnar shape and became meristematic at the onset of PeSERK expression, which was not initially detected in explant cells. Therefore, we propose that these changes represent the first observable steps towards the acquisition of a competent state within this regeneration system. PeSERK expression was limited to the early stages of somatic embryogenesis; the expression of this gene was confined to proembryogenic zones and was absent in the embryos after the globular stage. Our data also demonstrated that the dynamics of the mobilization of reserve compounds correlated with the differentiation of the embryogenic callus. PMID:26008651

  3. 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. PMID:27320638

  4. 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

  5. A genetically engineered live-attenuated simian-human immunodeficiency virus that co-expresses the RANTES gene improves the magnitude of cellular immunity in rhesus macaques

    SciTech Connect

    Shimizu, Yuya; Inaba, Katsuhisa; Kaneyasu, Kentaro; Ibuki, Kentaro; Himeno, Ai; Okoba, Masashi; Goto, Yoshitaka; Hayami, Masanori; Miura, Tomoyuki; Haga, Takeshi . E-mail: a0d518u@cc.miyazaki-u.ac.jp

    2007-04-25

    Regulated-on-activation-normal-T-cell-expressed-and-secreted (RANTES), a CC-chemokine, enhances antigen-specific T helper (Th) type-1 responses against HIV-1. To evaluate the adjuvant effects of RANTES against HIV vaccine candidate in SHIV-macaque models, we genetically engineered a live-attenuated SHIV to express the RANTES gene (SHIV-RANTES) and characterized the virus's properties in vivo. After the vaccination, the plasma viral loads were same in the SHIV-RANTES-inoculated monkeys and the parental nef-deleted SHIV (SHIV-NI)-inoculated monkeys. SHIV-RANTES provided some immunity in monkeys by remarkably increasing the antigen-specific CD4{sup +} Th cell-proliferative response and by inducing an antigen-specific IFN-{gamma} ELISpot response. The magnitude of the immunity in SHIV-RANTES-immunized animals, however, failed to afford greater protection against a heterologous pathogenic SHIV (SHIV-C2/1) challenge compared to control SHIV-NI-immunized animals. SHIV-RANTES immunized monkeys, elicited robust cellular CD4{sup +} Th responses and IFN-{gamma} ELISpot responses after SHIV-C2/1 challenge. These findings suggest that the chemokine RANTES can augment vaccine-elicited, HIV-specific CD4{sup +} T cell responses.

  6. 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

  7. Modulation of p53β and p53γ expression by regulating the alternative splicing of TP53 gene modifies cellular response.

    PubMed

    Marcel, V; Fernandes, K; Terrier, O; Lane, D P; Bourdon, J-C

    2014-09-01

    In addition to the tumor suppressor p53 protein, also termed p53α, the TP53 gene produces p53β and p53γ through alternative splicing of exons 9β and 9γ located within TP53 intron 9. Here we report that both TG003, a specific inhibitor of Cdc2-like kinases (Clk) that regulates the alternative splicing pre-mRNA pathway, and knockdown of SFRS1 increase expression of endogenous p53β and p53γ at mRNA and protein levels. Development of a TP53 intron 9 minigene shows that TG003 treatment and knockdown of SFRS1 promote inclusion of TP53 exons 9β/9γ. In a series of 85 primary breast tumors, a significant association was observed between expression of SFRS1 and α variant, supporting our experimental data. Using siRNA specifically targeting exons 9β/9γ, we demonstrate that cell growth can be driven by modulating p53β and p53γ expression in an opposite manner, depending on the cellular context. In MCF7 cells, p53β and p53γ promote apoptosis, thus inhibiting cell growth. By transient transfection, we show that p53β enhanced p53α transcriptional activity on the p21 and Bax promoters, while p53γ increased p53α transcriptional activity on the Bax promoter only. Moreover, p53β and p53γ co-immunoprecipitate with p53α only in the presence of p53-responsive promoter. Interestingly, although p53β and p53γ promote apoptosis in MCF7 cells, p53β and p53γ maintain cell growth in response to TG003 in a p53α-dependent manner. The dual activities of p53β and p53γ isoforms observed in non-treated and TG003-treated cells may result from the impact of TG003 on both expression and activities of p53 isoforms. Overall, our data suggest that p53β and p53γ regulate cellular response to modulation of alternative splicing pre-mRNA pathway by a small drug inhibitor. The development of novel drugs targeting alternative splicing process could be used as a novel therapeutic approach in human cancers. PMID:24926616

  8. High-Content Imaging and Gene Expression Approaches To Unravel the Effect of Surface Functionality on Cellular Interactions of Silver Nanoparticles.

    PubMed

    Manshian, Bella B; Pfeiffer, Christian; Pelaz, Beatriz; Heimerl, Thomas; Gallego, Marta; Möller, Marco; del Pino, Pablo; Himmelreich, Uwe; Parak, Wolfgang J; Soenen, Stefaan J

    2015-10-27

    The toxic effects of Ag nanoparticles (NPs) remain an issue of debate, where the respective contribution of the NPs themselves and of free Ag(+) ions present in the NP stock suspensions and after intracellular NP corrosion are not fully understood. Here, we employ a recently set up methodology based on high-content (HC) imaging combined with high-content gene expression studies to examine the interaction of three types of Ag NPs with identical core sizes, but coated with either mercaptoundecanoic acid (MUA), dodecylamine-modified poly(isobutylene-alt-maleic anhydride) (PMA), or poly(ethylene glycol) (PEG)-conjugated PMA with two types of cultured cells (primary human umbilical vein endothelial cells (HUVEC) and murine C17.2 neural progenitor cells). As a control, cells were also exposed to free Ag(+) ions at the same concentration as present in the respective Ag NP stock suspensions. The data reveal clear effects of the NP surface properties on cellular interactions. PEGylation of the NPs significantly reduces their cellular uptake efficiency, whereas MUA-NPs are more prone to agglomeration in complex tissue culture media. PEG-NPs display the lowest levels of toxicity, which is in line with their reduced cell uptake. MUA-NPs display the highest levels of toxicity, caused by autophagy, cell membrane damage, mitochondrial damage, and cytoskeletal deformations. At similar intracellular NP levels, PEG-NPs induce the highest levels of reactive oxygen species (ROS), but do not affect the cell cytoskeleton, in contrast to MUA- and PMA-NPs. Gene expression studies support the findings above, defining autophagy and cell membrane damage-related necrosis as main toxicity pathways. Additionally, immunotoxicity, DNA damage responses, and hypoxia-like toxicity were observed for PMA- and especially MUA-NPs. Together, these data reveal that Ag(+) ions do contribute to Ag NP-associated toxicity, particularly upon intracellular degradation. The different surface properties of the

  9. Regulation of gene expression by NFAT transcription factors in hibernating ground squirrels is dependent on the cellular environment.

    PubMed

    Zhang, Yichi; Storey, Kenneth B

    2016-09-01

    Calcineurin is a calmodulin-stimulated phosphatase that regulates the nuclear translocation of nuclear factor of activated T cell (NFAT) c1-4 through dephosphorylation. We believe that this mechanism plays various roles in the remodeling and maintenance of Ictidomys tridecemlineatus skeletal muscle. During hibernation, bouts of torpor and arousal take place, and squirrels do not lose muscle mass despite being inactive. Protein expression of Ca(2+) signaling proteins were studied using immunoblotting. A DNA-protein interaction ELISA technique was created to test the binding of NFATs in the nucleus to DNA probes containing the NFAT response element under environmental conditions reflective of those during hibernation. Calcineurin protein levels increased by 3.08-fold during torpor (compared to euthermic control), whereas calpain1 levels also rose by 3.66-fold during torpor. Calmodulin levels were elevated upon entering torpor. NFATc4 binding to DNA showed a 1.4-fold increase during torpor, and we found that this binding was further enhanced when 600 nM of Ca(2+) was supplemented. We also found that decreasing the temperature of ELISAs resulted in progressive decreases in the binding of NFATs c1, c3, and c4 to DNA. In summary, calmodulin and calpain1 appear to activate calcineurin and NFATc4 during torpor. NFAT binding to target promoters is affected by intranuclear [Ca(2+)] and environmental temperatures. Therefore, Ca(2+) signaling and temperature changes play key roles in regulation of the NFAT-calcineurin pathway in skeletal muscle of hibernating 13-lined ground squirrels over the torpor-arousal cycle, and they may contribute to the avoidance of disuse-induced muscle atrophy that occurs naturally in these animals. PMID:27344571

  10. Aberrant Gene Expression in Humans

    PubMed Central

    Yang, Ence; Ji, Guoli; Brinkmeyer-Langford, Candice L.; Cai, James J.

    2015-01-01

    Gene expression as an intermediate molecular phenotype has been a focus of research interest. In particular, studies of expression quantitative trait loci (eQTL) have offered promise for understanding gene regulation through the discovery of genetic variants that explain variation in gene expression levels. Existing eQTL methods are designed for assessing the effects of common variants, but not rare variants. Here, we address the problem by establishing a novel analytical framework for evaluating the effects of rare or private variants on gene expression. Our method starts from the identification of outlier individuals that show markedly different gene expression from the majority of a population, and then reveals the contributions of private SNPs to the aberrant gene expression in these outliers. Using population-scale mRNA sequencing data, we identify outlier individuals using a multivariate approach. We find that outlier individuals are more readily detected with respect to gene sets that include genes involved in cellular regulation and signal transduction, and less likely to be detected with respect to the gene sets with genes involved in metabolic pathways and other fundamental molecular functions. Analysis of polymorphic data suggests that private SNPs of outlier individuals are enriched in the enhancer and promoter regions of corresponding aberrantly-expressed genes, suggesting a specific regulatory role of private SNPs, while the commonly-occurring regulatory genetic variants (i.e., eQTL SNPs) show little evidence of involvement. Additional data suggest that non-genetic factors may also underlie aberrant gene expression. Taken together, our findings advance a novel viewpoint relevant to situations wherein common eQTLs fail to predict gene expression when heritable, rare inter-individual variation exists. The analytical framework we describe, taking into consideration the reality of differential phenotypic robustness, may be valuable for investigating

  11. Identification of cellular senescence-specific genes by comparative transcriptomics.

    PubMed

    Nagano, Taiki; Nakano, Masayuki; Nakashima, Akio; Onishi, Kengo; Yamao, Shunsuke; Enari, Masato; Kikkawa, Ushio; Kamada, Shinji

    2016-01-01

    Cellular senescence is defined as permanent cell cycle arrest induced by various stresses. Although the p53 transcriptional activity is essential for senescence induction, the downstream genes that are crucial for senescence remain unsolved. Here, by using a developed experimental system in which cellular senescence or apoptosis is induced preferentially by altering concentration of etoposide, a DNA-damaging drug, we compared gene expression profiles of senescent and apoptotic cells by microarray analysis. Subtraction of the expression profile of apoptotic cells identified 20 genes upregulated specifically in senescent cells. Furthermore, 6 out of 20 genes showed p53-dependent upregulation by comparing gene expression between p53-proficient and -deficient cells. These 6 genes were also upregulated during replicative senescence of normal human diploid fibroblasts, suggesting that upregulation of these genes is a general phenomenon in senescence. Among these genes, 2 genes (PRODH and DAO) were found to be directly regulated by p53, and ectopic expression of 4 genes (PRODH, DAO, EPN3, and GPR172B) affected senescence phenotypes induced by etoposide treatment. Collectively, our results identified several proteins as novel downstream effectors of p53-mediated senescence and provided new clues for further research on the complex signalling networks underlying the induction and maintenance of senescence. PMID:27545311

  12. Identification of cellular senescence-specific genes by comparative transcriptomics

    PubMed Central

    Nagano, Taiki; Nakano, Masayuki; Nakashima, Akio; Onishi, Kengo; Yamao, Shunsuke; Enari, Masato; Kikkawa, Ushio; Kamada, Shinji

    2016-01-01

    Cellular senescence is defined as permanent cell cycle arrest induced by various stresses. Although the p53 transcriptional activity is essential for senescence induction, the downstream genes that are crucial for senescence remain unsolved. Here, by using a developed experimental system in which cellular senescence or apoptosis is induced preferentially by altering concentration of etoposide, a DNA-damaging drug, we compared gene expression profiles of senescent and apoptotic cells by microarray analysis. Subtraction of the expression profile of apoptotic cells identified 20 genes upregulated specifically in senescent cells. Furthermore, 6 out of 20 genes showed p53-dependent upregulation by comparing gene expression between p53-proficient and -deficient cells. These 6 genes were also upregulated during replicative senescence of normal human diploid fibroblasts, suggesting that upregulation of these genes is a general phenomenon in senescence. Among these genes, 2 genes (PRODH and DAO) were found to be directly regulated by p53, and ectopic expression of 4 genes (PRODH, DAO, EPN3, and GPR172B) affected senescence phenotypes induced by etoposide treatment. Collectively, our results identified several proteins as novel downstream effectors of p53-mediated senescence and provided new clues for further research on the complex signalling networks underlying the induction and maintenance of senescence. PMID:27545311

  13. Changes in gene expression and cellular localization of insulin-like growth factors 1 and 2 in the ovaries during ovary development of the yellowtail, Seriola quinqueradiata.

    PubMed

    Higuchi, Kentaro; Gen, Koichiro; Izumida, Daisuke; Kazeto, Yukinori; Hotta, Takuro; Takashi, Toshinori; Aono, Hideaki; Soyano, Kiyoshi

    2016-06-01

    A method of controlling the somatic growth and reproduction of yellowtail fish (Seriola quinqueradiata) is needed in order to establish methods for the efficient aquaculture production of the species. However, little information about the hormonal interactions between somatic growth and reproduction is available for marine teleosts. There is accumulating evidence that insulin-like growth factor (IGF), a major hormone related somatic growth, plays an important role in fish reproduction. As the first step toward understanding the physiological role of IGF in the development of yellowtail ovaries, we characterized the expression and cellular localization of IGF-1 and IGF-2 in the ovary during development. We histologically classified the maturity of two-year-old females with ovaries at various developmental stages into the perinucleolar (Pn), yolk vesicle (Yv), primary yolk (Py), secondary yolk and tertiary yolk (Ty) stages, according to the most advanced type of oocyte present. The IGF-1 gene expression showed constitutively high levels at the different developmental stages, although IGF-1 mRNA levels tended to increase from the Py to the Ty stage with vitellogenesis, reaching maximum levels during the Ty stage. The IGF-2 mRNA levels increased as ovarian development advanced. Using immunohistochemistry methods, immunoreactive IGF-1 was mainly detected in the theca cells of ovarian follicles during late secondary oocyte growth, and in part of the granulosa cells of Ty stage oocytes. IGF-2 immunoreactivity was observed in all granulosa cells in layer in Ty stage oocytes. These results indicate that follicular IGFs may be involved in yellowtail reproduction via autocrine/paracrine mechanisms. PMID:26764214

  14. Impaired NFKBIE gene function decreases cellular uptake of methotrexate by down-regulating SLC19A1 expression in a human rheumatoid arthritis cell line

    PubMed Central

    Imamura, Hitoshi; Yoshina, Sawako; Ikari, Katsunori; Miyazawa, Keiji; Momohara, Shigeki; Mitani, Shohei

    2016-01-01

    Abstract Objective: A non-synonymous single nucleotide polymorphism (nsSNP, rs2233434, Val194Ala) in the NFKBIE (nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, epsilon) gene is known to be a rheumatoid arthritis (RA) susceptibility polymorphism in the Japanese RA population and could be closely associated with nuclear factor kappaB (NF-κB) activity. Inflammation caused by RA is sometimes associated with changes in expression levels of MTX (methotrexate) pathway-related genes. It is of interest to examine whether the NFKBIE gene had any influences on the mode of MTX action. Methods: Both knockdown of NFKBIE gene expression and overexpression of wild-type NFKBIE and Val194Ala mutation were performed. A transfected human RA synovial cell line was cultured and then gene expressions in the MTX pathway were measured. In addition, we measured the uptake and efflux of MTX derivatives under the NFKBIE knockdown condition. Results: Knockdown of NFKBIE reduced the mRNA for SLC19A1, a main MTX membrane transporter, and the intracellular accumulations of MTX derivatives. Moreover, our experiments also confirmed that overexpression of Val194Ala mutant NFKBIE decreased the SLC19A1 mRNA when compared to that of wild-type NFKBIE. Conclusions: We suggest that the impairment of NFKBIE gene function can reduce the uptake of MTX into cells, suggesting that the gene is an important factor for the RA outcome. PMID:26587663

  15. 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

  16. Genome-Wide Analysis Using Exon Arrays Demonstrates an Important Role for Expression of Extra-Cellular Matrix, Fibrotic Control and Tissue Remodelling Genes in Dupuytren's Disease

    PubMed Central

    Ham, Seungmin; de Kretser, David; Southwick, Graeme; Sprung, Carl N.

    2013-01-01

    Dupuytren's disease (DD) is a classic example of pathological fibrosis which results in a debilitating disorder affecting a large sector of the human population. It is characterized by excessive local proliferation of fibroblasts and over-production of collagen and other components of extracellular matrix (ECM) in the palmar fascia. The fibrosis progressively results in contracture of elements between the palmar fascia and skin causing flexion deformity or clawing of the fingers and a severe reduction in hand function. While much is known about the pathogenesis and surgical treatment of DD, little is known about the factors that cause its onset and progression, despite many years of research. Gene expression patterns in DD patients now offers the potential to identify genes that direct the pathogenesis of DD. In this study we used primary cultures of fibroblasts derived from excisional biopsies of fibrotic tissue from DD patients to compare the gene expression profiles on a genome-wide basis with normal control fibroblasts. Our investigations have identified genes that may be involved with DD pathogenesis including some which are directly relevant to fibrosis. In particular, these include significantly reduced expression levels of three matrix metallopeptidases (MMP1, MMP3, MMP16), follistatin, and STAT1, and significantly increased expression levels of fibroblast growth factors (FGF9, FGF11), a number of collagen genes and other ECM genes in DD patient samples. Many of these gene products are known to be involved in fibrosis, tumour formation and in the normal processes of tissue remodelling. In addition, alternative splicing was identified in some DD associated genes. These highly sensitive genomic investigations provide new insight into the molecular mechanisms that may underpin the development and progression of DD. PMID:23554969

  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. Cellular Distribution and Gene Expression Pattern of Metastasin (S100A4), Calgranulin A (S100A8), and Calgranulin B (S100A9) in Oral Lesions as Markers for Molecular Pathology.

    PubMed

    Reckenbeil, Jan; Kraus, Dominik; Probstmeier, Rainer; Allam, Jean-Pierre; Novak, Natalija; Frentzen, Matthias; Martini, Markus; Wenghoefer, Matthias; Winter, Jochen

    2016-07-01

    The objective of this study was to analyze cellular localization and expression levels of oncologic relevant members of the S100 family in common oral lesions.Biopsies of various oral lesions were analyzed. S100A4 showed a higher expression rate in leukoplakias and oral squamous cell carcinomas. Transcript levels of S100A8 and S100A9 were significantly decreased in malignant OSCCs. A correlation could be drawn between the expression levels of these genes and the pathological characteristics of the investigated lesions. S100A4, A8, and A9 proteins represent promising marker genes to evaluate the risk potential of suspicious oral lesions in molecular pathology. PMID:27294692

  20. 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

  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 load and expression of a unique subset of cellular genes in circulating lymphoid cells differentiate non-responders from responders to pegylated interferon alpha-ribavirin treatment.

    PubMed

    Pham, Tram N Q; Lin, Dolly M H; Mulrooney-Cousins, Patricia M; Churchill, Norma D; Kowala-Piaskowska, Arleta; Mozer-Lisewska, Iwona; Machaj, Anna; Pazgan-Simon, Monika; Zalewska, Malgorzata; Simon, Krzysztof; King, Dawn; Reddy, S Bharati; Michalak, Tomasz I

    2013-03-01

    Based on investigations of liver biopsy material, certain cellular genes have been implicated as correlates of success or failure to interferon alpha-ribavirin (IFN/RBV) therapy against hepatitis C. The current study aimed at determining whether expression of host genes thought to be relevant to HCV replication in the liver would be correlated with HCV infection status in peripheral blood mononuclear cells (PBMCs) and also with patient responsiveness to IFN/RBV treatment. Therefore, PBMCs from patients with chronic hepatitis C responding (n = 35) or not (n = 49) to IFN/RBV and from healthy controls (n = 15) were evaluated for HCV RNA load and cellular gene expression. Non-responders had 3- to 10-fold higher basal levels of interleukin (IL)-8, IFN-stimulated gene 15 (ISG15), 2',5'-oligoadenylate synthetase (OAS), and Toll-like receptors (TLR)-4, -5, and -7 compared to responders. Non-responders with similar post-treatment follow-ups as responders persistently expressed 6- to 20-fold greater levels of IL-8, ISG15, and OAS after therapy. Higher expression of IFN-α, IFN-γ, and IFN-λ was found in PBMCs of individuals achieving sustained virological response, either before or after therapy. Pre-treatment HCV RNA loads in PBMCs of non-responders were significantly higher (P = 0.016) than those of responders. In conclusion, the data indicate that immune cells of responders and non-responders to IFN/RBV therapy exhibited significantly different virological and host gene expression profiles. Elevated baseline HCV loads and TLR-4, -5, and -7 levels, and persistently high levels of IL-8, ISG15, and OAS were correlated with IFN non-responsiveness. The results warrant further investigations on the utilization of PBMCs for predicting success or failure to IFN-based therapies. PMID:23280583

  3. 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

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

    PubMed

    DebRoy, S; Hiraga, N; Imamura, M; Hayes, C N; Akamatsu, S; Canini, L; Perelson, A S; Pohl, R T; Persiani, S; Uprichard, S L; Tateno, C; Dahari, H; Chayama, K

    2016-09-01

    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. 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. 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. PMID:27272497

  5. 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

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

    PubMed

    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

  7. Cellular MYCro Economics: Balancing MYC Function with MYC Expression

    PubMed Central

    Levens, David

    2013-01-01

    The expression levels of the MYC oncoprotein have long been recognized to be associated with the outputs of major cellular processes including proliferation, cell growth, apoptosis, differentiation, and metabolism. Therefore, to understand how MYC operates, it is important to define quantitatively the relationship between MYC input and expression output for its targets as well as the higher-order relationships between the expression levels of subnetwork components and the flow of information and materials through those networks. Two different views of MYC are considered, first as a molecular microeconomic manager orchestrating specific positive and negative responses at individual promoters in collaboration with other transcription and chromatin components, and second, as a macroeconomic czar imposing an overarching rule onto all active genes. In either case, c-myc promoter output requires multiple inputs and exploits diverse mechanisms to tune expression to the appropriate levels relative to the thresholds of expression that separate health and disease. PMID:24186489

  8. 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. PMID:26896692

  9. Analysis of Expression, Cellular Localization, and Function of Three Inhibitors of Apoptosis (IAPs) from Litopenaeus vannamei during WSSV Infection and in Regulation of Antimicrobial Peptide Genes (AMPs)

    PubMed Central

    Wang, Pei-Hui; Wan, Ding-Hui; Gu, Zhi-Hua; Qiu, Wei; Chen, Yong-Gui; Weng, Shao-Ping; Yu, Xiao-Qiang; He, Jian-Guo

    2013-01-01

    Inhibitors of apoptosis (IAPs) play important roles in apoptosis and NF-κB activation. In this study, we cloned and characterized three IAPs (LvIAP1-3) from the Pacific white shrimp, Litopenaeusvannamei. LvIAP1-3 proteins shared signature domains and exhibited significant similarities with other IAP family proteins. The tissue distributions of LvIAP1-3 were studied. The expression of LvIAP1-3 was induced in the muscle after white spot syndrome virus (WSSV) infection. LvIAP1 expression in the gill, hemocytes, hepatopancreas, and intestine was responsive to WSSV and Vibrioalginolyticus infections. LvIAP2 expression in the gill, hemocytes, and hepatopancreas was also responsive to WSSV infection. The expression of LvIAP3 in the gill, hemocytes, and intestine was reduced after V. alginolyticus infection. When overexpressed in Drosophila S2 cells, GFP labeled-LvIAP2 was distributed in the cytoplasm and appeared as speck-like aggregates in the nucleus. Both LvIAP1 and LvIAP3 were widely distributed throughout the cytoplasm and nucleus. The expression of LvIAP1, LvIAP2, and LvIAP3 was significantly knocked down by dsRNA-mediated gene silencing. In the gill of LvIAP1- or LvIAP3-silenced shrimp, the expression of WSSV VP28 was significantly higher than that of the dsGFP control group, suggesting that LvIAP1 and LvIAP3 may play protective roles in host defense against WSSV infection. Intriguingly, the LvIAP2-silenced shrimp all died within 48 hours after dsLvIAP2 injection. In the hemocytes of LvIAP2-silenced shrimps, the expression of antimicrobial peptide genes (AMPs), including Penaeidins, lysozyme, crustins, Vibriopenaeicidae-induced cysteine and proline-rich peptides (VICPs), was significantly downregulated, while the expression of anti-lipopolysaccharide factors (ALFs) was upregulated. Moreover, LvIAP2 activated the promoters of the NF-κB pathway-controlled AMPs, such as shrimp Penaeidins and Drosophila drosomycin and attacin A, in Drosophila S2 cells. Taken together

  10. Gene expression technology

    SciTech Connect

    Goeddel, D.V. )

    1990-01-01

    The articles in this volume were assemble to enable the reader to design effective strategies for the expression of cloned genes and cDNAs. More than a compilation of papers describing the multitude of techniques now available for expressing cloned genes, this volume provides a manual that should prove useful for solving the majority of expression problems one likely to encounter. The four major expression systems commonly available to most investigators are stressed: Escherichia coli, Bacillus subtilis, yeast, and mammalian cells. Each of these system has its advantages and disadvantages, details of which are found in Chapter 1 and the strategic overviews for the four major sections of the volume. The papers in each of these sections provide many suggestions on how to proceed if initial expression levels are not sufficient.

  11. Gene expression networks.

    PubMed

    Thomas, Reuben; Portier, Christopher J

    2013-01-01

    With the advent of microarrays and next-generation biotechnologies, the use of gene expression data has become ubiquitous in biological research. One potential drawback of these data is that they are very rich in features or genes though cost considerations allow for the use of only relatively small sample sizes. A useful way of getting at biologically meaningful interpretations of the environmental or toxicological condition of interest would be to make inferences at the level of a priori defined biochemical pathways or networks of interacting genes or proteins that are known to perform certain biological functions. This chapter describes approaches taken in the literature to make such inferences at the biochemical pathway level. In addition this chapter describes approaches to create hypotheses on genes playing important roles in response to a treatment, using organism level gene coexpression or protein-protein interaction networks. Also, approaches to reverse engineer gene networks or methods that seek to identify novel interactions between genes are described. Given the relatively small sample numbers typically available, these reverse engineering approaches are generally useful in inferring interactions only among a relatively small or an order 10 number of genes. Finally, given the vast amounts of publicly available gene expression data from different sources, this chapter summarizes the important sources of these data and characteristics of these sources or databases. In line with the overall aims of this book of providing practical knowledge to a researcher interested in analyzing gene expression data from a network perspective, the chapter provides convenient publicly accessible tools for performing analyses described, and in addition describe three motivating examples taken from the published literature that illustrate some of the relevant analyses. PMID:23086841

  12. Antisense RNA inactivation of gene expression of a cell-cell adhesion protein (gp64) in the cellular slime mold Polysphondylium pallidum.

    PubMed

    Funamoto, S; Ochiai, H

    1996-05-01

    The gp64 protein of Polysphondylium pallidum has been shown to mediate EDTA-stable cell-cell adhesion. To explore the functional role of gp64, we made an antisense RNA expression construct designed to prevent the gene expression of gp64; the construct was introduced into P. pallidum cells and the transformants were characterised. The antisense RNA-expressing clone L3mc2 which had just been harvested at the growth phase tended to re-form in aggregates smaller in size than did the parental cells in either the presence or absence of 10 mM EDTA. In contrast, 6.5-hour starved L3mc2 cells remained considerably dissociated from each other after 5 minutes gyrating, although aggregation gradually increased by 50% during a further 55 minutes gyrating in the presence of 10 mM EDTA. Correspondingly, L3mc2 lacked specifically the cell-cell adhesion protein, gp64. We therefore conclude that the gp64 protein is involved in forming the EDTA-resistant cell-cell contact. In spite of the absence of gp64, L3mc2 exhibited normal developmental processes, a fact which demonstrates that another cell-cell adhesion system exists in the development of Polysphondylium. This is the first report in which an antisense RNA technique was successfully applied to Polysphondylium. PMID:8743948

  13. 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. PMID:23226257

  14. Seasonal Effects on Gene Expression

    PubMed Central

    Goldinger, Anita; Shakhbazov, Konstantin; Henders, Anjali K.; McRae, Allan F.; Montgomery, Grant W.; Powell, Joseph E.

    2015-01-01

    Many health conditions, ranging from psychiatric disorders to cardiovascular disease, display notable seasonal variation in severity and onset. In order to understand the molecular processes underlying this phenomenon, we have examined seasonal variation in the transcriptome of 606 healthy individuals. We show that 74 transcripts associated with a 12-month seasonal cycle were enriched for processes involved in DNA repair and binding. An additional 94 transcripts demonstrated significant seasonal variability that was largely influenced by blood cell count levels. These transcripts were enriched for immune function, protein production, and specific cellular markers for lymphocytes. Accordingly, cell counts for erythrocytes, platelets, neutrophils, monocytes, and CD19 cells demonstrated significant association with a 12-month seasonal cycle. These results demonstrate that seasonal variation is an important environmental regulator of gene expression and blood cell composition. Notable changes in leukocyte counts and genes involved in immune function indicate that immune cell physiology varies throughout the year in healthy individuals. PMID:26023781

  15. The OsCYP19-4 Gene Is Expressed as Multiple Alternatively Spliced Transcripts Encoding Isoforms with Distinct Cellular Localizations and PPIase Activities under Cold Stress.

    PubMed

    Lee, Areum; Lee, Sang Sook; Jung, Won Yong; Park, Hyun Ji; Lim, Bo Ra; Kim, Hyun-Soon; Ahn, Jun Cheul; Cho, Hye Sun

    2016-01-01

    Alternative splicing (AS) is an important molecular mechanism by which single genes can generate multiple mRNA isoforms. We reported previously that, in Oryza sativa, the cyclophilin 19-4 (OsCYP19-4.1) transcript was significantly upregulated in response to cold stress, and that transgenic plants were cold tolerant. Here we show that, under cold stress, OsCYP19-4 produces eight transcript variants by intron retention and exon skipping, resulting in production of four distinct protein isoforms. The OsCYP19-4 AS isoforms exhibited different cellular localizations in the epidermal cells: in contrast to OsCYP19-4.1, the OsCYP19-4.2 and OsCYP19-4.3 proteins were primarily targeted to guard and subsidiary cells, whereas OsCYP19-4.5, which consists largely of an endoplasmic reticulum (ER) targeting signal, was co-localized with the RFP-BiP marker in the ER. In OsCYP19-4.2, the key residues of the PPIase domain are altered; consistent with this, recombinant OsCYP19-4.2 had significantly lower PPIase activity than OsCYP19-4.1 in vitro. Specific protein-protein interactions between OsCYP19-4.2/3 and AtRCN1 were verified in yeast two-hybrid (Y2H) and bimolecular fluoresence complementation (BiFC assays), although the OsCYP19-4 isoforms could not bind each other. Based on these results, we propose that two OsCYP19-4 AS isoforms, OsCYP19-4.2 and OsCYP19-4.3, play roles linking auxin transport and cold stress via interactions with RCN1. PMID:27447607

  16. The OsCYP19-4 Gene Is Expressed as Multiple Alternatively Spliced Transcripts Encoding Isoforms with Distinct Cellular Localizations and PPIase Activities under Cold Stress

    PubMed Central

    Lee, Areum; Lee, Sang Sook; Jung, Won Yong; Park, Hyun Ji; Lim, Bo Ra; Kim, Hyun-Soon; Ahn, Jun Cheul; Cho, Hye Sun

    2016-01-01

    Alternative splicing (AS) is an important molecular mechanism by which single genes can generate multiple mRNA isoforms. We reported previously that, in Oryza sativa, the cyclophilin 19-4 (OsCYP19-4.1) transcript was significantly upregulated in response to cold stress, and that transgenic plants were cold tolerant. Here we show that, under cold stress, OsCYP19-4 produces eight transcript variants by intron retention and exon skipping, resulting in production of four distinct protein isoforms. The OsCYP19-4 AS isoforms exhibited different cellular localizations in the epidermal cells: in contrast to OsCYP19-4.1, the OsCYP19-4.2 and OsCYP19-4.3 proteins were primarily targeted to guard and subsidiary cells, whereas OsCYP19-4.5, which consists largely of an endoplasmic reticulum (ER) targeting signal, was co-localized with the RFP-BiP marker in the ER. In OsCYP19-4.2, the key residues of the PPIase domain are altered; consistent with this, recombinant OsCYP19-4.2 had significantly lower PPIase activity than OsCYP19-4.1 in vitro. Specific protein-protein interactions between OsCYP19-4.2/3 and AtRCN1 were verified in yeast two-hybrid (Y2H) and bimolecular fluoresence complementation (BiFC assays), although the OsCYP19-4 isoforms could not bind each other. Based on these results, we propose that two OsCYP19-4 AS isoforms, OsCYP19-4.2 and OsCYP19-4.3, play roles linking auxin transport and cold stress via interactions with RCN1. PMID:27447607

  17. Induction of cellular hsp70 expression by human cytomegalovirus.

    PubMed Central

    Santomenna, L D; Colberg-Poley, A M

    1990-01-01

    Expression of the cellular heat shock protein 70 gene (hsp70) is transiently induced by human cytomegalovirus (HCMV) infection of permissive human diploid fibroblasts. Induction of the cellular heat shock response during critical times of infection had previously been reported to alter the growth of HCMV in vitro. Thus, a potential interaction between heat shock proteins and HCMV expression was indicated. HCMV dramatically increased expression of hsp70 RNA within 8 h of infection. hsp70 RNA remained elevated at 24 and 48 h postinfection and decreased to low levels of 72 h postinfection. Induction of HSP70 protein occurred more slowly; inducible HSP70 protein encoded by this RNA increased within 16 h postinfection and continued to increase throughout infection until 72 h postinfection, when the highest abundance of inducible HSP70 protein was observed. Cells that received both heat (43 degrees C for 70 min) treatment and HCMV infection expressed hsp70 RNA to levels above the sum of levels present in cells given either treatment alone. Furthermore, hsp70 RNA induction occurred earlier and remained elevated longer than in cells infected with HCMV alone or in cells treated with heat alone, respectively. Nevertheless, the pattern of HCMV immediate-early transcript expression at 2, 4, and 6 h postinfection appeared to be unchanged by this prior heat treatment. Our results suggest that heat shock treatment and HCMV infection can act additively in stimulating hsp70 RNA expression. The previously reported stimulation of HCMV growth in vitro following the heat shock response apparently does not result from alterations in the steady-state expression of HCMV immediate-early transcripts. Images PMID:2157870

  18. Induction of plasminogen activator inhibitor 1 gene expression in murine liver by lipopolysaccharide. Cellular localization and role of endogenous tumor necrosis factor-alpha.

    PubMed Central

    Fearns, C.; Loskutoff, D. J.

    1997-01-01

    We previously demonstrated that lipopolysaccharide (LPS) induces plasminogen activator inhibitor 1 (PAI-1) gene expression primarily in endothelial cells in most organs of the mouse, with maximal induction by 3 hours. Here we show that induction in the liver occurs in a distinctly different pattern. For example, the increase in PAI-1 mRNA in liver was biphasic with an initial peak at 1 to 2 hours and a second peak at 6 to 8 hours. Moreover, in situ hybridization experiments revealed that PAI-1 mRNA was induced in both endothelial cells and hepatocytes. The endothelial cell response was monophasic and maximal between 1 and 4 hours, whereas the hepatocyte response was biphasic, peaking at 2 hours and again at 6 to 8 hours. To determine possible mechanisms involved in the induction of PAI-1 by LPS, we analyzed the tissues for changes in tumor necrosis factor (TNF)-alpha LPS caused a rapid induction of TNF-alpha mRNA in Kupffer cells, detectable within 15 minutes. Pretreatment of mice with anti-TNF antiserum before challenge with LPS reduced the subsequent increase in plasma levels of PAI-1 by 50 to 70% and significantly reduced the level of induction of PAI-1 mRNA in the liver at both early and late times. Pretreatment appeared to inhibit induction primarily within hepatocytes. These results suggest that LPS may induce PAI-1 in endothelial cells and hepatocytes by different mechanisms. Images Figure 3 Figure 4 Figure 7 PMID:9033272

  19. 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.

  20. 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.

  1. 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.

  2. 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

  3. Methodological Limitations in Determining Astrocytic Gene Expression

    PubMed Central

    Peng, Liang; Guo, Chuang; Wang, Tao; Li, Baoman; Gu, Li; Wang, Zhanyou

    2013-01-01

    Traditionally, astrocytic mRNA and protein expression are studied by in situ hybridization (ISH) and immunohistochemically. This led to the concept that astrocytes lack aralar, a component of the malate-aspartate-shuttle. At least similar aralar mRNA and protein expression in astrocytes and neurons isolated by fluorescence-assisted cell sorting (FACS) reversed this opinion. Demonstration of expression of other astrocytic genes may also be erroneous. Literature data based on morphological methods were therefore compared with mRNA expression in cells obtained by recently developed methods for determination of cell-specific gene expression. All Na,K-ATPase-α subunits were demonstrated by immunohistochemistry (IHC), but there are problems with the cotransporter NKCC1. Glutamate and GABA transporter gene expression was well determined immunohistochemically. The same applies to expression of many genes of glucose metabolism, whereas a single study based on findings in bacterial artificial chromosome (BAC) transgenic animals showed very low astrocytic expression of hexokinase. Gene expression of the equilibrative nucleoside transporters ENT1 and ENT2 was recognized by ISH, but ENT3 was not. The same applies to the concentrative transporters CNT2 and CNT3. All were clearly expressed in FACS-isolated cells, followed by biochemical analysis. ENT3 was enriched in astrocytes. Expression of many nucleoside transporter genes were shown by microarray analysis, whereas other important genes were not. Results in cultured astrocytes resembled those obtained by FACS. These findings call for reappraisal of cellular nucleoside transporter expression. FACS cell yield is small. Further development of cell separation methods to render methods more easily available and less animal and cost consuming and parallel studies of astrocytic mRNA and protein expression by ISH/IHC and other methods are necessary, but new methods also need to be thoroughly checked. PMID:24324456

  4. GENES FOR TUMOR MARKERS ARE CLUSTERED WITH CELLULAR PROTO-ONCOGENES ON HUMAN CHROMOSOMES

    EPA Science Inventory

    The relative mapping positions of genes for polypeptides expressed abnormally in tumors (tumor markers) and cellular proto-oncogenes were analyzed and a remarkable degree of co-mapping of tumor marker genes with oncogenes in the human karyotype were found. It is proposed that abe...

  5. Gene Express Inc.

    PubMed

    Saccomanno, Colette F

    2006-07-01

    Gene Express, Inc. is a technology-licensing company and provider of Standardized Reverse Transcription Polymerase Chain Reaction (StaRT-PCR) services. Designed by and for clinical researchers involved in pharmaceutical, biomarker and molecular diagnostic product development, StaRT-PCR is a unique quantitative and standardized multigene expression measurement platform. StaRT-PCR meets all of the performance characteristics defined by the US FDA as required to support regulatory submissions [101,102] , and by the Clinical Laboratory Improvement Act of 1988 (CLIA) as necessary to support diagnostic testing [1] . A standardized mixture of internal standards (SMIS), manufactured in bulk, provides integrated quality control wherein each native template target gene is measured relative to a competitive template internal standard. Bulk production enables the compilation of a comprehensive standardized database from across multiple experiments, across collaborating laboratories and across the entire clinical development lifecycle of a given compound or diagnostic product. For the first time, all these data are able to be directly compared. Access to such a database can dramatically shorten the time from investigational new drug (IND) to new drug application (NDA), or save time and money by hastening a substantiated 'no-go' decision. High-throughput StaRT-PCR is conducted at the company's automated Standardized Expression Measurement (SEM) Center. Currently optimized for detection on a microcapillary electrophoretic platform, StaRT-PCR products also may be analyzed on microarray, high-performance liquid chromatography (HPLC), or matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) platforms. SEM Center services deliver standardized genomic data--data that will accelerate the application of pharmacogenomic technology to new drug and diagnostic test development and facilitate personalized medicine. PMID:16886903

  6. Association between the expression of LHR, FSHR and CYP19 genes, cellular distribution of encoded proteins and proliferation of porcine granulosa cells in real-time.

    PubMed

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

    2014-01-01

    The process of granulosa cell luteinization is part of the main process determining growth, differentiation and proliferation of these cells. Although the mechanisms underlying the regulation of luteinizing hormone receptor (LHR), follicle stimulating hormone receptor (FSHR) and cytochrome P450 aromatase expression in mammalian granulosa cells is well understood, still little is known about the expression of mRNA and encoded proteins in relation to cell proliferation and luteinization in vitro. Porcine granulosa cells were observed in vitro at a168-h period while undergoing real-time proliferation using an RTCA system. Furthermore, LHR, FSHR and CYP19 mRNA expression were detected using RQ-PCR after 168 h of in vitro culture (IVC) at 24-h intervals, and LHR, FSHR and P450arom were examined by confocal microscopic observation at 0 h, 24 h, 48 h, 96 h, and 168 h of IVC. We found increased expression of LHR and CYP19 mRNA at 24 h and 48 h of IVC compared to the other stages (P less than 0.01, P less than 0.001), whereas FSHR mRNA was higher only at 0 h (P less than 0.001). In contrast, LHR, FSHR and P450arom protein expression was significantly higher at the end of the 168-h IVC period compared to 0 h, 24 h, 48 h and 96 h (P less than 0.001). LHR, FSHR and P450arom were distributed in the cytoplasm of porcine GCs at each time point of IVC. When analyzing cell proliferation, differences in cell index were observed (at least P less than 0.05) between the first (0-24 h) and the last period (144-168 h) of IVC; however, soon after 24 h of IVC a logarithmic increase in proliferation was also seen. We assume that the expression of LHR, FSHR and CYP19 mRNAs depends on the period of in vitro cultivation and may be linked with the luteinization process of porcine GCs. Furthermore, the patterns of mRNA and protein expression suggest a post-transcriptional regulation of LHR, FSHR and P450arom. In summary, it can be presumed that mRNA and protein expression and in vitro

  7. Evolution of gene expression after gene amplification.

    PubMed

    Garcia, Nelson; Zhang, Wei; Wu, Yongrui; Messing, Joachim

    2015-05-01

    We took a rather unique approach to investigate the conservation of gene expression of prolamin storage protein genes across two different subfamilies of the Poaceae. We took advantage of oat plants carrying single maize chromosomes in different cultivars, called oat-maize addition (OMA) lines, which permitted us to determine whether regulation of gene expression was conserved between the two species. We found that γ-zeins are expressed in OMA7.06, which carries maize chromosome 7 even in the absence of the trans-acting maize prolamin-box-binding factor (PBF), which regulates their expression. This is likely because oat PBF can substitute for the function of maize PBF as shown in our transient expression data, using a γ-zein promoter fused to green fluorescent protein (GFP). Despite this conservation, the younger, recently amplified prolamin genes in maize, absent in oat, are not expressed in the corresponding OMAs. However, maize can express the oldest prolamin gene, the wheat high-molecular weight glutenin Dx5 gene, even when maize Pbf is knocked down (through PbfRNAi), and/or another maize transcription factor, Opaque-2 (O2) is knocked out (in maize o2 mutant). Therefore, older genes are conserved in their regulation, whereas younger ones diverged during evolution and eventually acquired a new repertoire of suitable transcriptional activators. PMID:25912045

  8. Evolution of Gene Expression after Gene Amplification

    PubMed Central

    Garcia, Nelson; Zhang, Wei; Wu, Yongrui; Messing, Joachim

    2015-01-01

    We took a rather unique approach to investigate the conservation of gene expression of prolamin storage protein genes across two different subfamilies of the Poaceae. We took advantage of oat plants carrying single maize chromosomes in different cultivars, called oat–maize addition (OMA) lines, which permitted us to determine whether regulation of gene expression was conserved between the two species. We found that γ-zeins are expressed in OMA7.06, which carries maize chromosome 7 even in the absence of the trans-acting maize prolamin-box-binding factor (PBF), which regulates their expression. This is likely because oat PBF can substitute for the function of maize PBF as shown in our transient expression data, using a γ-zein promoter fused to green fluorescent protein (GFP). Despite this conservation, the younger, recently amplified prolamin genes in maize, absent in oat, are not expressed in the corresponding OMAs. However, maize can express the oldest prolamin gene, the wheat high-molecular weight glutenin Dx5 gene, even when maize Pbf is knocked down (through PbfRNAi), and/or another maize transcription factor, Opaque-2 (O2) is knocked out (in maize o2 mutant). Therefore, older genes are conserved in their regulation, whereas younger ones diverged during evolution and eventually acquired a new repertoire of suitable transcriptional activators. PMID:25912045

  9. High expression hampers horizontal gene transfer.

    PubMed

    Park, Chungoo; Zhang, Jianzhi

    2012-01-01

    Horizontal gene transfer (HGT), the movement of genetic material from one species to another, is a common phenomenon in prokaryotic evolution. Although the rate of HGT is known to vary among genes, our understanding of the cause of this variation, currently summarized by two rules, is far from complete. The first rule states that informational genes, which are involved in DNA replication, transcription, and translation, have lower transferabilities than operational genes. The second rule asserts that protein interactivity negatively impacts gene transferability. Here, we hypothesize that high expression hampers HGT, because the fitness cost of an HGT to the recipient, arising from the 1) energy expenditure in transcription and translation, 2) cytotoxic protein misfolding, 3) reduction in cellular translational efficiency, 4) detrimental protein misinteraction, and 5) disturbance of the optimal protein concentration or cell physiology, increases with the expression level of the transferred gene. To test this hypothesis, we examined laboratory and natural HGTs to Escherichia coli. We observed lower transferabilities of more highly expressed genes, even after controlling the confounding factors from the two established rules and the genic GC content. Furthermore, expression level predicts gene transferability better than all other factors examined. We also confirmed the significant negative impact of gene expression on the rate of HGTs to 127 of 133 genomes of eubacteria and archaebacteria. Together, these findings establish the gene expression level as a major determinant of horizontal gene transferability. They also suggest that most successful HGTs are initially slightly deleterious, fixed because of their negligibly low costs rather than high benefits to the recipient. PMID:22436996

  10. Control of RANKL Gene Expression

    PubMed Central

    O'Brien, Charles A.

    2009-01-01

    Osteoclasts are highly specialized cells capable of degrading mineralized tissue and form at different regions of bone to meet different physiological needs, such as mobilization of calcium, modeling of bone structure, and remodeling of bone matrix. Osteoclast production is elevated in a number of pathological conditions, many of which lead to loss of bone mass. Whether normal or pathological, osteoclastogenesis strictly depends upon support from accessory cells which supply cytokines required for osteoclast differentiation. Only one of these cytokines, receptor activator of NFκB ligand (RANKL), is absolutely essential for osteoclast formation throughout life and is thus expressed by all cell types that support osteoclast differentiation. The central role of RANKL in bone resorption is highlighted by the fact that it is the basis for a new therapy to inhibit bone loss. This review will discuss mechanisms that control RANKL gene expression in different osteoclast-support cells and how the study of such mechanisms may lead to a better understanding of the cellular interactions that drive normal and pathological bone resorption. PMID:19716455

  11. Serial analysis of gene expression.

    PubMed

    Velculescu, V E; Zhang, L; Vogelstein, B; Kinzler, K W

    1995-10-20

    The characteristics of an organism are determined by the genes expressed within it. A method was developed, called serial analysis of gene expression (SAGE), that allows the quantitative and simultaneous analysis of a large number of transcripts. To demonstrate this strategy, short diagnostic sequence tags were isolated from pancreas, concatenated, and cloned. Manual sequencing of 1000 tags revealed a gene expression pattern characteristic of pancreatic function. New pancreatic transcripts corresponding to novel tags were identified. SAGE should provide a broadly applicable means for the quantitative cataloging and comparison of expressed genes in a variety of normal, developmental, and disease states. PMID:7570003

  12. 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... document entitled ``Guidance for Industry: Preclinical Assessment of Investigational Cellular and Gene... for Biologics Research and Evaluation (CBER), Office of Cellular, Tissue, and Gene Therapies...

  13. 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. PMID:25298512

  14. Global analysis of patterns of gene expression during Drosophila embryogenesis

    PubMed Central

    Tomancak, Pavel; Berman, Benjamin P; Beaton, Amy; Weiszmann, Richard; Kwan, Elaine; Hartenstein, Volker; Celniker, Susan E; Rubin, Gerald M

    2007-01-01

    Background Cell and tissue specific gene expression is a defining feature of embryonic development in multi-cellular organisms. However, the range of gene expression patterns, the extent of the correlation of expression with function, and the classes of genes whose spatial expression are tightly regulated have been unclear due to the lack of an unbiased, genome-wide survey of gene expression patterns. Results We determined and documented embryonic expression patterns for 6,003 (44%) of the 13,659 protein-coding genes identified in the Drosophila melanogaster genome with over 70,000 images and controlled vocabulary annotations. Individual expression patterns are extraordinarily diverse, but by supplementing qualitative in situ hybridization data with quantitative microarray time-course data using a hybrid clustering strategy, we identify groups of genes with similar expression. Of 4,496 genes with detectable expression in the embryo, 2,549 (57%) fall into 10 clusters representing broad expression patterns. The remaining 1,947 (43%) genes fall into 29 clusters representing restricted expression, 20% patterned as early as blastoderm, with the majority restricted to differentiated cell types, such as epithelia, nervous system, or muscle. We investigate the relationship between expression clusters and known molecular and cellular-physiological functions. Conclusion Nearly 60% of the genes with detectable expression exhibit broad patterns reflecting quantitative rather than qualitative differences between tissues. The other 40% show tissue-restricted expression; the expression patterns of over 1,500 of these genes are documented here for the first time. Within each of these categories, we identified clusters of genes associated with particular cellular and developmental functions. PMID:17645804

  15. 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.

  16. 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

  17. Gene Expression in Oligodendroglial Tumors

    PubMed Central

    Shaw, Elisabeth J.; Haylock, Brian; Husband, David; du Plessis, Daniel; Sibson, D. Ross; Warnke, Peter C.; Walker, Carol

    2010-01-01

    Background: Oligodendroglial tumors with 1p/19q loss are more likely to be chemosensitive and have longer survival than those with intact 1p/19q, but not all respond to chemotherapy, warranting investigation of the biological basis of chemosensitivity. Methods: Gene expression profiling was performed using amplified antisense RNA from 28 oligodendroglial tumors treated with chemotherapy (26 serial stereotactic biopsy, 2 resection). Expression of differentially expressed genes was validated by real-time PCR. Results: Unsupervised hierarchical clustering showed clustering of multiple samples from the same case in 14/17 cases and identified subgroups associated with tumor grade and 1p/19q status. 176 genes were differentially expressed, 164 being associated with 1p/19q loss (86% not on 1p or 19q). 94 genes differed between responders and non-responders to chemotherapy; 12 were not associated with 1p/19q loss. Significant differential expression was confirmed in 11/13 selected genes. Novel genes associated with response to therapy included SSBP2, GFRA1, FAP and RASD1. IQGAP1, INA, TGIF1, NR2F2 and MYCBP were differentially expressed in oligodendroglial tumors with 1p/19q loss. Conclusion: Gene expression profiling using serial stereotactic biopsies indicated greater homogeneity within tumors than between tumors. Genes associated with 1p/19q status or response were identified warranting further elucidation of their role in oligodendroglial tumors. PMID:20966545

  18. 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.

  19. 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

  20. Epigenetics, cellular memory and gene regulation.

    PubMed

    Henikoff, Steven; Greally, John M

    2016-07-25

    The field described as 'epigenetics' has captured the imagination of scientists and the lay public. Advances in our understanding of chromatin and gene regulatory mechanisms have had impact on drug development, fueling excitement in the lay public about the prospects of applying this knowledge to address health issues. However, when describing these scientific advances as 'epigenetic', we encounter the problem that this term means different things to different people, starting within the scientific community and amplified in the popular press. To help researchers understand some of the misconceptions in the field and to communicate the science accurately to each other and the lay audience, here we review the basis for many of the assumptions made about what are currently referred to as epigenetic processes. PMID:27458904

  1. Identification and sequence analysis of a new member of the mouse HSP70 gene family and characterization of its unique cellular and developmental pattern of expression in the male germ line.

    PubMed Central

    Zakeri, Z F; Wolgemuth, D J; Hunt, C R

    1988-01-01

    A unique member of the mouse HSP70 gene family has been isolated and characterized with respect to its DNA sequence organization and expression. The gene contains extensive similarity to a heat shock-inducible HSP70 gene within the coding region but diverges in both 3' and 5' nontranslated regions. The gene does not yield transcripts in response to heat shock in mouse L cells. Rather, the gene appears to be activated uniquely in the male germ line. Analysis of RNA from different developmental stages and from enriched populations of spermatogenic cells revealed that this gene is expressed during the prophase stage of meiosis. A transcript different in size from the major heat-inducible mouse transcripts is most abundant in meiotic prophase spermatocytes and decreases in abundance in postmeiotic stages of spermatogenesis. This pattern of expression is distinct from that observed for another member of this gene family, which was previously shown to be expressed abundantly in postmeiotic germ cells. These observations suggest that specific HSP70 gene family members play distinct roles in the differentiation of the germ cell lineage in mammals. Images PMID:3405224

  2. Quantitative imaging of gene expression in Drosophila embryos.

    PubMed

    Surkova, Svetlana; Myasnikova, Ekaterina; Kozlov, Konstantin N; Pisarev, Andrei; Reinitz, John; Samsonova, Maria

    2013-06-01

    Quantitative measurements derived using sophisticated microscopy techniques are essential for understanding the basic principles that control the behavior of biological systems. Here we describe a data pipeline developed to extract quantitative data on segmentation gene expression from confocal images of gene expression patterns in Drosophila. The pipeline consists of image segmentation, background removal, temporal characterization of an embryo, data registration, and data averaging. This pipeline has been successfully applied to obtain quantitative gene expression data at cellular resolution in space and at 6.5-min resolution in time. It has also enabled the construction of a spatiotemporal atlas of segmentation gene expression. We describe the software used to construct a workflow for extracting quantitative data on segmentation gene expression and the BREReA package, which implements the methods for background removal and registration of segmentation gene expression patterns. PMID:23734022

  3. Single-cell PCR profiling of gene expression in hematopoiesis.

    PubMed

    Teles, José; Enver, Tariq; Pina, Cristina

    2014-01-01

    Single-cell analysis of gene expression offers the possibility of exploring cellular and molecular heterogeneity in stem and developmental cell systems, including cancer, to infer routes of cellular specification and their respective gene regulatory modules. PCR-based technologies, although limited to the analysis of a predefined set of genes, afford a cost-effective balance of throughput and biological information and have become a method of choice in stem cell laboratories. Here we describe an experimental and analytical protocol based on the Fluidigm microfluidics platform for the simultaneous expression analysis of 48 or 96 genes in multiples of 48 or 96 cells. We detail wet laboratory procedures and describe clustering, principal component analysis, correlation, and classification tools for the inference of cellular pathways and gene networks. PMID:25062620

  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. Population-level control of gene expression

    NASA Astrophysics Data System (ADS)

    Nevozhay, Dmitry; Adams, Rhys; van Itallie, Elizabeth; Bennett, Matthew; Balazsi, Gabor

    2011-03-01

    Gene expression is the process that translates genetic information into proteins, that determine the way cells live, function and even die. It was demonstrated that cells with identical genomes exposed to the same environment can differ in their protein composition and therefore phenotypes. Protein levels can vary between cells due to the stochastic nature of intracellular biochemical events, indicating that the genotype-phenotype connection is not deterministic at the cellular level. We asked whether genomes could encode isogenic cell populations more reliably than single cells. To address this question, we built two gene circuits to control three cell population-level characteristics: gene expression mean, coefficient of variation and non-genetic memory of previous expression states. Indeed, we found that these population-level characteristics were more predictable than the gene expression of single cells in a well-controlled environment. This research was supported by the NIH Director's New Innovator Award 1DP2 OD006481-01 and Welch Foundation Grant C-1729.

  6. 75 FR 65640 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-26

    ... 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 and... Tumor Vaccines and Biotechnology Branch, Office of Cellular, Tissue and Gene Therapies, Center...

  7. 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

  8. Dual transcriptional-translational cascade permits cellular level tuneable expression control

    PubMed Central

    Morra, Rosa; Shankar, Jayendra; Robinson, Christopher J.; Halliwell, Samantha; Butler, Lisa; Upton, Mathew; Hay, Sam; Micklefield, Jason; Dixon, Neil

    2016-01-01

    The ability to induce gene expression in a small molecule dependent manner has led to many applications in target discovery, functional elucidation and bio-production. To date these applications have relied on a limited set of protein-based control mechanisms operating at the level of transcription initiation. The discovery, design and reengineering of riboswitches offer an alternative means by which to control gene expression. Here we report the development and characterization of a novel tunable recombinant expression system, termed RiboTite, which operates at both the transcriptional and translational level. Using standard inducible promoters and orthogonal riboswitches, a multi-layered modular genetic control circuit was developed to control the expression of both bacteriophage T7 RNA polymerase and recombinant gene(s) of interest. The system was benchmarked against a number of commonly used E. coli expression systems, and shows tight basal control, precise analogue tunability of gene expression at the cellular level, dose-dependent regulation of protein production rates over extended growth periods and enhanced cell viability. This novel system expands the number of E. coli expression systems for use in recombinant protein production and represents a major performance enhancement over and above the most widely used expression systems. PMID:26405200

  9. Gene expression profiling in male genital lichen sclerosus.

    PubMed

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

    2011-10-01

    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

  10. Nuclear Neighborhoods and Gene Expression

    PubMed Central

    Zhao, Rui; Bodnar, Megan S.; Spector, David L.

    2009-01-01

    Summary The eukaryotic nucleus is a highly compartmentalized and dynamic environment. Chromosome territories are arranged non-randomly within the nucleus and numerous studies have indicated that a gene’s position in the nucleus can impact its transcriptional activity. Here, we focus on recent advances in our understanding of the influence of specific nuclear neighborhoods on gene expression or repression. Nuclear neighborhoods associated with transcriptional repression include the inner nuclear membrane/nuclear lamina and peri-nucleolar chromatin, whereas neighborhoods surrounding the nuclear pore complex, PML nuclear bodies, and nuclear speckles seem to be transcriptionally permissive. While nuclear position appears to play an important role in gene expression, it is likely to be only one piece of a flexible puzzle that incorporates numerous parameters. We are still at a very early, yet exciting stage in our journey toward deciphering the mechanism(s) that govern the permissiveness of gene expression/repression within different nuclear neighborhoods. PMID:19339170

  11. 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

  12. 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.

  13. From gene expressions to genetic networks

    NASA Astrophysics Data System (ADS)

    Cieplak, Marek

    2009-03-01

    A method based on the principle of entropy maximization is used to identify the gene interaction network with the highest probability of giving rise to experimentally observed transcript profiles [1]. In its simplest form, the method yields the pairwise gene interaction network, but it can also be extended to deduce higher order correlations. Analysis of microarray data from genes in Saccharomyces cerevisiae chemostat cultures exhibiting energy metabollic oscillations identifies a gene interaction network that reflects the intracellular communication pathways. These pathways adjust cellular metabolic activity and cell division to the limiting nutrient conditions that trigger metabolic oscillations. The success of the present approach in extracting meaningful genetic connections suggests that the maximum entropy principle is a useful concept for understanding living systems, as it is for other complex, nonequilibrium systems. The time-dependent behavior of the genetic network is found to involve only a few fundamental modes [2,3]. [4pt] REFERENCES:[0pt] [1] T. R. Lezon, J. R. Banavar, M. Cieplak, A. Maritan, and N. Fedoroff, Using the principle of entropy maximization to infer genetic interaction networks from gene expression patterns, Proc. Natl. Acad. Sci. (USA) 103, 19033-19038 (2006) [0pt] [2] N. S. Holter, M. Mitra, A. Maritan, M. Cieplak, J. R. Banavar, and N. V. Fedoroff, Fundamental patterns underlying gene expression profiles: simplicity from complexity, Proc. Natl. Acad. Sci. USA 97, 8409-8414 (2000) [0pt] [3] N. S. Holter, A. Maritan, M. Cieplak, N. V. Fedoroff, and J. R. Banavar, Dynamic modeling of gene expression data, Proc. Natl. Acad. Sci. USA 98, 1693-1698 (2001)

  14. 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

  15. Neighboring Genes Show Correlated Evolution in Gene Expression

    PubMed Central

    Ghanbarian, Avazeh T.; Hurst, Laurence D.

    2015-01-01

    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. PMID:25743543

  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

    2015-12-01

    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. PMID:26627736

  17. Nuclear structure, gene expression and development.

    PubMed

    Brown, K

    1999-01-01

    This article considers the extent to which features of nuclear structure are involved in the regulation of genome function. The recent renaissance in imaging technology has inspired a new determination to assign specific functions to nuclear domains or structures, many of which have been described as "factories" to express the idea that they coordinate nuclear processes in an efficient way. Visual data have been combined with genetic and biochemical information to support the idea that nuclear organization has functional significance. Particular DNA sequences or chromatin structures may nucleate domains that are permissive or restrictive of transcription, to which active or inactive loci could be recruited. Associations within the nucleus, as well as many nuclear structures, are transient and change dynamically during cell cycle progression and development. Despite this complexity, elucidation of the possible structural basis of epigenetic phenomena, such as the inheritance of a "cellular memory" of gene expression status, is an important goal for cell biology. Topics for discussion include the regulatory effect of chromatin structure on gene expression, putative "nuclear addresses" for genes and proteins, the functional significance of nuclear bodies, and the role of the nuclear matrix in nuclear compartmentalization. PMID:10651237

  18. Transition Metals in Control of Gene Expression

    NASA Astrophysics Data System (ADS)

    O'Halloran, Thomas V.

    1993-08-01

    Metalloproteins play structural and catalytic roles in gene expression. The metalloregulatory proteins are a subclass that exerts metal-responsive control of genes involved in respiration, metabolism, and metal-specific homeostasis or stress-response systems, such as iron uptake and storage, copper efflux, and mercury detoxification. Two allosteric mechanisms for control of gene expression were first discovered in metalloregulatory systems: an iron-responsive translational control mechanism for ferritin production and a mercury-responsive DNA-distortion mechanism for transcriptional control of detoxification genes. These otherwise unrelated mechanisms give rise to a rapid physiological response when metal ion concentrations exceed a dangerous threshold. Molecular recognition in these allosteric metal ion receptors is achieved through atypical coordination geometries, cluster formation, or complexes with prosthetic groups, such as sulfide and heme. Thus, many of the inorganic assemblies that otherwise buttress the structure of biopolymers or catalyze substrate transformation in active sites of enzymes have also been adapted to serve sensor functions in the metalloregulatory proteins. Mechanistic studies of these metal-sensor protein interactions are providing new insights into fundamental aspects of inorganic chemistry, molecular biology, and cellular physiology.

  19. Gene expression during memory formation.

    PubMed

    Igaz, Lionel Muller; Bekinschtein, Pedro; Vianna, Monica M R; Izquierdo, Ivan; Medina, Jorge H

    2004-01-01

    For several decades, neuroscientists have provided many clues that point out the involvement of de novo gene expression during the formation of long-lasting forms of memory. However, information regarding the transcriptional response networks involved in memory formation has been scarce and fragmented. With the advent of genome-based technologies, combined with more classical approaches (i.e., pharmacology and biochemistry), it is now feasible to address those relevant questions--which gene products are modulated, and when that processes are necessary for the proper storage of memories--with unprecedented resolution and scale. Using one-trial inhibitory (passive) avoidance training of rats, one of the most studied tasks so far, we found two time windows of sensitivity to transcriptional and translational inhibitors infused into the hippocampus: around the time of training and 3-6 h after training. Remarkably, these periods perfectly overlap with the involvement of hippocampal cAMP/PKA (protein kinase A) signaling pathways in memory consolidation. Given the complexity of transcriptional responses in the brain, particularly those related to processing of behavioral information, it was clearly necessary to address this issue with a multi-variable, parallel-oriented approach. We used cDNA arrays to screen for candidate inhibitory avoidance learning-related genes and analyze the dynamic pattern of gene expression that emerges during memory consolidation. These include genes involved in intracellular kinase networks, synaptic function, DNA-binding and chromatin modification, transcriptional activation and repression, translation, membrane receptors, and oncogenes, among others. Our findings suggest that differential and orchestrated hippocampal gene expression is necessary in both early and late periods of long-term memory consolidation. Additionally, this kind of studies may lead to the identification and characterization of genes that are relevant for the pathogenesis

  20. Whirlin increases TRPV1 channel expression and cellular stability.

    PubMed

    Ciardo, Maria Grazia; Andrés-Bordería, Amparo; Cuesta, Natalia; Valente, Pierluigi; Camprubí-Robles, María; Yang, Jun; Planells-Cases, Rosa; Ferrer-Montiel, Antonio

    2016-01-01

    The expression and function of TRPV1 are influenced by its interaction with cellular proteins. Here, we identify Whirlin, a cytoskeletal PDZ-scaffold protein implicated in hearing, vision and mechanosensory transduction, as an interacting partner of TRPV1. Whirlin associates with TRPV1 in cell lines and in primary cultures of rat nociceptors. Whirlin is expressed in 55% of mouse sensory C-fibers, including peptidergic and non-peptidergic nociceptors, and co-localizes with TRPV1 in 70% of them. Heterologous expression of Whirlin increased TRPV1 protein expression and trafficking to the plasma membrane, and promoted receptor clustering. Silencing Whirlin expression with siRNA or blocking protein translation resulted in a concomitant degradation of TRPV1 that could be prevented by inhibiting the proteasome. The degradation kinetics of TRPV1 upon arresting protein translation mirrored that of Whirlin in cells co-expressing both proteins, suggesting a parallel degradation mechanism. Noteworthy, Whirlin expression significantly reduced TRPV1 degradation induced by prolonged exposure to capsaicin. Thus, our findings indicate that Whirlin and TRPV1 are associated in a subset of nociceptors and that TRPV1 protein stability is increased through the interaction with the cytoskeletal scaffold protein. Our results suggest that the Whirlin–TRPV1 complex may represent a novel molecular target and its pharmacological disruption might be a therapeutic strategy for the treatment of peripheral TRPV1-mediated disorders. PMID:26516054

  1. 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.

  2. Using gene expression profiling to evaluate cellular responses in mouse lungs exposed to V2O5 and a group of other mouse lung tumorigens and non-tumorigens.

    PubMed

    Black, Michael B; Dodd, Darol E; McMullen, Patrick D; Pendse, Salil; MacGregor, Judith A; Gollapudi, B Bhaskar; Andersen, Melvin E

    2015-10-01

    Many compounds test positive for lung tumors in two-year NTP carcinogenicity bioassays in B6C3F1 mice. V2O5 was identified as a lung carcinogen in this assay, leading to its IARC (International Agency for Research on Cancer) classification as group 2b or a "possible" human carcinogen. To assess potential tumorigenic mode of action of V2O5, we compared gene expression and gene ontology enrichment in lung tissue of female B6C3F1 mice exposed for 13 weeks to a V2O5 particulate aerosol at a tumorigenic level (2.0 mg/m(3)). Relative to 12 other compounds also tested for carcinogenicity in 2-year bioassays in mice, there were 1026 differentially expressed genes with V2O5, of which 483 were unique to V2O5. Ontology analysis of the 1026 V2O5 differentially expressed genes showed enrichment for hyaluronan and sphingolipid metabolism, adenylate cyclase functions, c-AMP signaling and PKA activation/signaling. Enrichment of lipids/lipoprotein metabolism and inflammatory pathways were consistent with previously reported clinical findings. Enrichment of c-AMP and PKA signaling pathways may arise due to inhibition of phosphatases, a known biological action of vanadate. We saw no enrichment for DNA-damage, oxidative stress, cell cycle, or apoptosis pathway signaling in mouse lungs exposed to V2O5 which is in contrast with past studies evaluating in vivo gene expression in target tissues of other carcinogens (arsenic, formaldehyde, naphthalene and chloroprene). PMID:26210822

  3. 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. PMID:27052691

  4. Systems Biophysics of Gene Expression

    PubMed Central

    Vilar, Jose M.G.; Saiz, Leonor

    2013-01-01

    Gene expression is a process central to any form of life. It involves multiple temporal and functional scales that extend from specific protein-DNA interactions to the coordinated regulation of multiple genes in response to intracellular and extracellular changes. This diversity in scales poses fundamental challenges to the use of traditional approaches to fully understand even the simplest gene expression systems. Recent advances in computational systems biophysics have provided promising avenues to reliably integrate the molecular detail of biophysical process into the system behavior. Here, we review recent advances in the description of gene regulation as a system of biophysical processes that extend from specific protein-DNA interactions to the combinatorial assembly of nucleoprotein complexes. There is now basic mechanistic understanding on how promoters controlled by multiple, local and distal, DNA binding sites for transcription factors can actively control transcriptional noise, cell-to-cell variability, and other properties of gene regulation, including precision and flexibility of the transcriptional responses. PMID:23790365

  5. Control of Renin Gene Expression

    PubMed Central

    Glenn, Sean T.; Jones, Craig A.; Gross, Kenneth W.; Pan, Li

    2015-01-01

    Renin, as part of the renin-angiotensin system, plays a critical role in the regulation of blood pressure, electrolyte homeostasis, mammalian renal development and progression of fibrotic/hypertrophic diseases. Renin gene transcription is subject to complex developmental and tissue-specific regulation. Initial studies using the mouse As4.1 cell line, which has many characteristics of the renin-expressing juxtaglomerular cells of the kidney, have identified a proximal promoter region (−197 to −50 bp) and an enhancer (−2866 to −2625 bp) upstream of the Ren-1c gene, which are critical for renin gene expression. The proximal promoter region contains several transcription factor-binding sites including a binding site for the products of the developmental control genes Hox. The enhancer consists of at least 11 transcription factor-binding sites and is responsive to various signal transduction pathways including cAMP, retinoic acid, endothelin-1, and cytokines, all of which are known to alter renin mRNA levels. Furthermore, in vivo models have validated several of these key components found within the proximal promoter region and the enhancer as well as other key sites necessary for renin gene transcription. PMID:22576577

  6. Global Gene Expression in Staphylococcus aureus Biofilms

    PubMed Central

    Beenken, Karen E.; Dunman, Paul M.; McAleese, Fionnuala; Macapagal, Daphne; Murphy, Ellen; Projan, Steven J.; Blevins, Jon S.; Smeltzer, Mark S.

    2004-01-01

    We previously demonstrated that mutation of the staphylococcal accessory regulator (sarA) in a clinical isolate of Staphylococcus aureus (UAMS-1) results in an impaired capacity to form a biofilm in vitro (K. E. Beenken, J. S. Blevins, and M. S. Smeltzer, Infect. Immun. 71:4206-4211, 2003). In this report, we used a murine model of catheter-based biofilm formation to demonstrate that a UAMS-1 sarA mutant also has a reduced capacity to form a biofilm in vivo. Surprisingly, mutation of the UAMS-1 ica locus had little impact on biofilm formation in vitro or in vivo. In an effort to identify additional loci that might be relevant to biofilm formation and/or the adaptive response required for persistence of S. aureus within a biofilm, we isolated total cellular RNA from UAMS-1 harvested from a biofilm grown in a flow cell and compared the transcriptional profile of this RNA to RNA isolated from both exponential- and stationary-phase planktonic cultures. Comparisons were done using a custom-made Affymetrix GeneChip representing the genomic complement of six strains of S. aureus (COL, N315, Mu50, NCTC 8325, EMRSA-16 [strain 252], and MSSA-476). The results confirm that the sessile lifestyle associated with persistence within a biofilm is distinct by comparison to the lifestyles of both the exponential and postexponential phases of planktonic culture. Indeed, we identified 48 genes in which expression was induced at least twofold in biofilms over expression under both planktonic conditions. Similarly, we identified 84 genes in which expression was repressed by a factor of at least 2 compared to expression under both planktonic conditions. A primary theme that emerged from the analysis of these genes is that persistence within a biofilm requires an adaptive response that limits the deleterious effects of the reduced pH associated with anaerobic growth conditions. PMID:15231800

  7. 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...

  8. Multidimensional regulation of gene expression in the C. elegans embryo

    PubMed Central

    Murray, John Isaac; Boyle, Thomas J.; Preston, Elicia; Vafeados, Dionne; Mericle, Barbara; Weisdepp, Peter; Zhao, Zhongying; Bao, Zhirong; Boeck, Max; Waterston, Robert H.

    2012-01-01

    How cells adopt different expression patterns is a fundamental question of developmental biology. We quantitatively measured reporter expression of 127 genes, primarily transcription factors, in every cell and with high temporal resolution in C. elegans embryos. Embryonic cells are highly distinct in their gene expression; expression of the 127 genes studied here can distinguish nearly all pairs of cells, even between cells of the same tissue type. We observed recurrent lineage-regulated expression patterns for many genes in diverse contexts. These patterns are regulated in part by the TCF-LEF transcription factor POP-1. Other genes' reporters exhibited patterns correlated with tissue, position, and left–right asymmetry. Sequential patterns both within tissues and series of sublineages suggest regulatory pathways. Expression patterns often differ between embryonic and larval stages for the same genes, emphasizing the importance of profiling expression in different stages. This work greatly expands the number of genes in each of these categories and provides the first large-scale, digitally based, cellular resolution compendium of gene expression dynamics in live animals. The resulting data sets will be a useful resource for future research. PMID:22508763

  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. Congruence of tissue expression profiles from Gene Expression Atlas, SAGEmap and TissueInfo databases

    PubMed Central

    Huminiecki, Lukasz; Lloyd, Andrew T; Wolfe, Kenneth H

    2003-01-01

    Background Extracting biological knowledge from large amounts of gene expression information deposited in public databases is a major challenge of the postgenomic era. Additional insights may be derived by data integration and cross-platform comparisons of expression profiles. However, database meta-analysis is complicated by differences in experimental technologies, data post-processing, database formats, and inconsistent gene and sample annotation. Results We have analysed expression profiles from three public databases: Gene Expression Atlas, SAGEmap and TissueInfo. These are repositories of oligonucleotide microarray, Serial Analysis of Gene Expression and Expressed Sequence Tag human gene expression data respectively. We devised a method, Preferential Expression Measure, to identify genes that are significantly over- or under-expressed in any given tissue. We examined intra- and inter-database consistency of Preferential Expression Measures. There was good correlation between replicate experiments of oligonucleotide microarray data, but there was less coherence in expression profiles as measured by Serial Analysis of Gene Expression and Expressed Sequence Tag counts. We investigated inter-database correlations for six tissue categories, for which data were present in the three databases. Significant positive correlations were found for brain, prostate and vascular endothelium but not for ovary, kidney, and pancreas. Conclusion We show that data from Gene Expression Atlas, SAGEmap and TissueInfo can be integrated using the UniGene gene index, and that expression profiles correlate relatively well when large numbers of tags are available or when tissue cellular composition is simple. Finally, in the case of brain, we demonstrate that when PEM values show good correlation, predictions of tissue-specific expression based on integrated data are very accurate. PMID:12885301

  11. Gene Expression Studies in Mosquitoes

    PubMed Central

    Chen, Xlao-Guang; Mathur, Geetika; James, Anthony A.

    2009-01-01

    Research on gene expression in mosquitoes is motivated by both basic and applied interests. Studies of genes involved in hematophagy, reproduction, olfaction, and immune responses reveal an exquisite confluence of biological adaptations that result in these highly-successful life forms. The requirement of female mosquitoes for a bloodmeal for propagation has been exploited by a wide diversity of viral, protozoan and metazoan pathogens as part of their life cycles. Identifying genes involved in host-seeking, blood feeding and digestion, reproduction, insecticide resistance and susceptibility/refractoriness to pathogen development is expected to provide the bases for the development of novel methods to control mosquito-borne diseases. Advances in mosquito transgenesis technologies, the availability of whole genome sequence information, mass sequencing and analyses of transcriptomes and RNAi techniques will assist development of these tools as well as deepen the understanding of the underlying genetic components for biological phenomena characteristic of these insect species. PMID:19161831

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

    PubMed Central

    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

  13. The Gene Expression Omnibus database

    PubMed Central

    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/. PMID:27008011

  14. 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/. PMID:27008011

  15. Classification of genes based on gene expression analysis

    NASA Astrophysics Data System (ADS)

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

    2008-05-01

    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.

  16. 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.

  17. 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

  18. Gene expression in the brain during reovirus encephalitis

    PubMed Central

    Tyler, Kenneth L; Leser, J Smith; Phang, Tzu L; Clarke, Penny

    2010-01-01

    Viral encephalitis remains a significant cause of morbidity and mortality throughout the world. We performed microarray analysis to identify genes and pathways that are differentially regulated during reovirus encephalitis and that may provide novel therapeutic targets for virus-induced diseases of the central nervous system (CNS). An increase in the expression of 130 cellular genes was found in the brains of reovirus-infected mice at early times post infection, compared to mock-infected controls. The up-regulation of these genes was consistent with activation of innate immune responses, particularly interferon signaling. At later times post infection, when significant CNS injury is present and mice exhibit signs of severe neurologic disease, many more (1374) genes were up-regulated, indicating that increased gene expression correlates with disease pathology. Virus-induced gene expression at late times post infection was again consistent with the activation of innate immune responses. However, additional significant pathways included those associated with cytokine signaling and apoptosis, both of which can contribute to CNS injury. This is the first report comparing virus-induced cellular gene and pathway regulation at early and late times following virus infection of the brain. The shift of virus-induced gene expression from innate immune responses at early times post infection to cytokine signaling and apoptosis at later times suggests a potential therapeutic strategy that preserves early protective responses whilst inhibiting later responses that contribute to pathogenesis. PMID:20158406

  19. 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

  20. Identification of four soybean reference genes for gene expression normalization

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Gene expression analysis requires the use of reference genes stably expressed independently of specific tissues or environmental conditions. Housekeeping genes (e.g., actin, tubulin, ribosomal, polyubiquitin and elongation factor 1-alpha) are commonly used as reference genes with the assumption tha...

  1. Mitochondrial RNA granules: Compartmentalizing mitochondrial gene expression.

    PubMed

    Jourdain, Alexis A; Boehm, Erik; Maundrell, Kinsey; Martinou, Jean-Claude

    2016-03-14

    In mitochondria, DNA replication, gene expression, and RNA degradation machineries coexist within a common nondelimited space, raising the question of how functional compartmentalization of gene expression is achieved. Here, we discuss the recently characterized "mitochondrial RNA granules," mitochondrial subdomains with an emerging role in the regulation of gene expression. PMID:26953349

  2. Spatial reconstruction of single-cell gene expression

    PubMed Central

    Satija, Rahul; Farrell, Jeffrey A.; Gennert, David; Schier, Alexander F.; Regev, Aviv

    2015-01-01

    Spatial localization is a key determinant of cellular fate and behavior, but spatial RNA assays traditionally rely on staining for a limited number of RNA species. In contrast, single-cell RNA-seq allows for deep profiling of cellular gene expression, but established methods separate cells from their native spatial context. Here we present Seurat, a computational strategy to infer cellular localization by integrating single-cell RNA-seq data with in situ RNA patterns. We applied Seurat to spatially map 851 single cells from dissociated zebrafish (Danio rerio) embryos, inferring a transcriptome-wide map of spatial patterning. We confirmed Seurat’s accuracy using several experimental approaches, and used it to identify a set of archetypal expression patterns and spatial markers. Additionally, Seurat correctly localizes rare subpopulations, accurately mapping both spatially restricted and scattered groups. Seurat will be applicable to mapping cellular localization within complex patterned tissues in diverse systems. PMID:25867923

  3. Mammalian Mss51 is a skeletal muscle-specific gene modulating cellular metabolism

    PubMed Central

    Moyer, Adam L.; Wagner, Kathryn R.

    2015-01-01

    Background The transforming growth factor β (TGF-β) signaling pathways modulate skeletal muscle growth, regeneration, and cellular metabolism. Several recent gene expression studies have shown that inhibition of myostatin and TGF-β1 signaling consistently leads to a significant reduction in expression of Mss51, also named Zmynd17. The function of mammalian Mss51 is unknown although a putative homolog in yeast is a mitochondrial translational activator. Objective The objective of this work was to characterize mammalian Mss51. Methods Quantitative RT-PCR and immunoblot of subcellular fractionation were used to determine expression patterns and localization of Mss51. The CRISPR/Cas9 system was used to reduce expression of Mss51 in C2C12 myoblasts and the function of Mss51 was evaluated in assays of proliferation, differentiation and cellular metabolism. Results Mss51 was predominantly expressed in skeletal muscle and in those muscles dominated by fast-twitch fibers. In vitro, its expression was upregulated upon differentiation of C2C12 myoblasts into myotubes. Expression of Mss51 was modulated in response to altered TGF-β family signaling. In human muscle, Mss51 localized to the mitochondria. Its genetic disruption resulted in increased levels of cellular ATP, β-oxidation, glycolysis, and oxidative phosphorylation. Conclusions Mss51 is a novel, skeletal muscle-specific gene and a key target of myostatin and TGF-β1 signaling. Unlike myostatin, TGF-β1 and IGF-1, Mss51 does not regulate myoblast proliferation or differentiation. Rather, Mss51 appears to be one of the effectors of these growth factors on metabolic processes including fatty acid oxidation, glycolysis and oxidative phosphorylation. PMID:26634192

  4. The transcriptional regulation of regucalcin gene expression.

    PubMed

    Yamaguchi, Masayoshi

    2011-01-01

    Regucalcin, which is discovered as a calcium-binding protein in 1978, has been shown to play a multifunctional role in many tissues and cell types; regucalcin has been proposed to play a pivotal role in keeping cell homeostasis and function for cell response. Regucalcin and its gene are identified in over 15 species consisting of regucalcin family. Comparison of the nucleotide sequences of regucalcin from vertebrate species is highly conserved in their coding region with throughout evolution. The regucalcin gene is localized on the chromosome X in rat and human. The organization of rat regucalcin gene consists of seven exons and six introns and several consensus regulatory elements exist upstream of the 5'-flanking region. AP-1, NF1-A1, RGPR-p117, β-catenin, and other factors have been found to be a transcription factor in the enhancement of regucalcin gene promoter activity. The transcription activity of regucalcin gene is enhanced through intracellular signaling factors that are mediated through the phosphorylation and dephosphorylation of nuclear protein in vitro. Regucalcin mRNA and its protein are markedly expressed in the liver and kidney cortex of rats. The expression of regucalcin mRNA in the liver and kidney cortex has been shown to stimulate by hormonal factors (including calcium, calcitonin, parathyroid hormone, insulin, estrogen, and dexamethasone) in vivo. Regucalcin mRNA expression is enhanced in the regenerating liver after partial hepatectomy of rats in vivo. The expression of regucalcin mRNA in the liver and kidney with pathophysiological state has been shown to suppress, suggesting an involvement of regucalcin in disease. Liver regucalcin expression is down-regulated in tumor cells, suggesting a suppressive role in the development of carcinogenesis. Liver regucalcin is markedly released into the serum of rats with chemically induced liver injury in vivo. Serum regucalcin has a potential sensitivity as a specific biochemical marker of chronic

  5. Does inbreeding affect gene expression in birds?

    PubMed Central

    Hansson, Bengt; Naurin, Sara; Hasselquist, Dennis

    2014-01-01

    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. PMID:25232028

  6. MRI of Transgene Expression: Correlation to Therapeutic Gene Expression

    PubMed Central

    Ichikawa, Tomotsugu; Högemanny, Dagmar; Saeki, Yoshinaga; Tyminski, Edyta; Terada, Kinya; Weissleder, Ralph; Chiocca, E Antonio; Basilion, James P

    2002-01-01

    Abstract Magnetic resonance imaging (MRI) can provide highresolution 3D maps of structural and functional information, yet its use of mapping in vivo gene expression has only recently been explored. A potential application for this technology is to noninvasively image transgene expression. The current study explores the latter using a nonregulatable internalizing engineered transferrin receptor (ETR) whose expression can be probed for with a superparamagnetic Tf-CLIO probe. Using an HSV-based amplicon vector system for transgene delivery, we demonstrate that: 1) ETR is a sensitive MR marker gene; 2) several transgenes can be efficiently expressed from a single amplicon; 3) expression of each transgene results in functional gene product; and 4) ETR gene expression correlates with expression of therapeutic genes when the latter are contained within the same amplicon. These data, taken together, suggest that MRI of ETR expression can serve as a surrogate for measuring therapeutic transgene expression. PMID:12407446

  7. Adult mouse brain gene expression patterns bear an embryologic imprint

    PubMed Central

    Zapala, Matthew A.; Hovatta, Iiris; Ellison, Julie A.; Wodicka, Lisa; Del Rio, Jo A.; Tennant, Richard; Tynan, Wendy; Broide, Ron S.; Helton, Rob; Stoveken, Barbara S.; Winrow, Christopher; Lockhart, Daniel J.; Reilly, John F.; Young, Warren G.; Bloom, Floyd E.; Lockhart, David J.; Barlow, Carrolee

    2005-01-01

    The current model to explain the organization of the mammalian nervous system is based on studies of anatomy, embryology, and evolution. To further investigate the molecular organization of the adult mammalian brain, we have built a gene expression-based brain map. We measured gene expression patterns for 24 neural tissues covering the mouse central nervous system and found, surprisingly, that the adult brain bears a transcriptional “imprint” consistent with both embryological origins and classic evolutionary relationships. Embryonic cellular position along the anterior–posterior axis of the neural tube was shown to be closely associated with, and possibly a determinant of, the gene expression patterns in adult structures. We also observed a significant number of embryonic patterning and homeobox genes with region-specific expression in the adult nervous system. The relationships between global expression patterns for different anatomical regions and the nature of the observed region-specific genes suggest that the adult brain retains a degree of overall gene expression established during embryogenesis that is important for regional specificity and the functional relationships between regions in the adult. The complete collection of extensively annotated gene expression data along with data mining and visualization tools have been made available on a publicly accessible web site (www.barlow-lockhart-brainmapnimhgrant.org). PMID:16002470

  8. 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

  9. 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.

  10. 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

  11. Noise in gene expression is coupled to growth rate.

    PubMed

    Keren, Leeat; van Dijk, David; Weingarten-Gabbay, Shira; Davidi, Dan; Jona, Ghil; Weinberger, Adina; Milo, Ron; Segal, Eran

    2015-12-01

    Genetically identical cells exposed to the same environment display variability in gene expression (noise), with important consequences for the fidelity of cellular regulation and biological function. Although population average gene expression is tightly coupled to growth rate, the effects of changes in environmental conditions on expression variability are not known. Here, we measure the single-cell expression distributions of approximately 900 Saccharomyces cerevisiae promoters across four environmental conditions using flow cytometry, and find that gene expression noise is tightly coupled to the environment and is generally higher at lower growth rates. Nutrient-poor conditions, which support lower growth rates, display elevated levels of noise for most promoters, regardless of their specific expression values. We present a simple model of noise in expression that results from having an asynchronous population, with cells at different cell-cycle stages, and with different partitioning of the cells between the stages at different growth rates. This model predicts non-monotonic global changes in noise at different growth rates as well as overall higher variability in expression for cell-cycle-regulated genes in all conditions. The consistency between this model and our data, as well as with noise measurements of cells growing in a chemostat at well-defined growth rates, suggests that cell-cycle heterogeneity is a major contributor to gene expression noise. Finally, we identify gene and promoter features that play a role in gene expression noise across conditions. Our results show the existence of growth-related global changes in gene expression noise and suggest their potential phenotypic implications. PMID:26355006

  12. Noise in gene expression is coupled to growth rate

    PubMed Central

    Keren, Leeat; van Dijk, David; Weingarten-Gabbay, Shira; Davidi, Dan; Jona, Ghil; Weinberger, Adina; Milo, Ron; Segal, Eran

    2015-01-01

    Genetically identical cells exposed to the same environment display variability in gene expression (noise), with important consequences for the fidelity of cellular regulation and biological function. Although population average gene expression is tightly coupled to growth rate, the effects of changes in environmental conditions on expression variability are not known. Here, we measure the single-cell expression distributions of approximately 900 Saccharomyces cerevisiae promoters across four environmental conditions using flow cytometry, and find that gene expression noise is tightly coupled to the environment and is generally higher at lower growth rates. Nutrient-poor conditions, which support lower growth rates, display elevated levels of noise for most promoters, regardless of their specific expression values. We present a simple model of noise in expression that results from having an asynchronous population, with cells at different cell-cycle stages, and with different partitioning of the cells between the stages at different growth rates. This model predicts non-monotonic global changes in noise at different growth rates as well as overall higher variability in expression for cell-cycle–regulated genes in all conditions. The consistency between this model and our data, as well as with noise measurements of cells growing in a chemostat at well-defined growth rates, suggests that cell-cycle heterogeneity is a major contributor to gene expression noise. Finally, we identify gene and promoter features that play a role in gene expression noise across conditions. Our results show the existence of growth-related global changes in gene expression noise and suggest their potential phenotypic implications. PMID:26355006

  13. 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

  14. 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. PMID:23466301

  15. Concise review: Nanoparticles and cellular carriers-allies in cancer imaging and cellular gene therapy?

    PubMed Central

    Tang, Catherine; Russell, Pamela J; Martiniello-Wilks, Rosetta; J Rasko, John E; Khatri, Aparajita

    2010-01-01

    Ineffective treatment and poor patient management continue to plague the arena of clinical oncology. The crucial issues include inadequate treatment efficacy due to ineffective targeting of cancer deposits, systemic toxicities, suboptimal cancer detection and disease monitoring. This has led to the quest for clinically relevant, innovative multifaceted solutions such as development of targeted and traceable therapies. Mesenchymal stem cells (MSCs) have the intrinsic ability to “home” to growing tumors and are hypoimmunogenic. Therefore, these can be used as (a) “Trojan Horses” to deliver gene therapy directly into the tumors and (b) carriers of nanoparticles to allow cell tracking and simultaneous cancer detection. The camouflage of MSC carriers can potentially tackle the issues of safety, vector, and/or transgene immunogenicity as well as nanoparticle clearance and toxicity. The versatility of the nanotechnology platform could allow cellular tracking using single or multimodal imaging modalities. Toward that end, noninvasive magnetic resonance imaging (MRI) is fast becoming a clinical favorite, though there is scope for improvement in its accuracy and sensitivity. In that, use of superparamagnetic iron-oxide nanoparticles (SPION) as MRI contrast enhancers may be the best option for tracking therapeutic MSC. The prospects and consequences of synergistic approaches using MSC carriers, gene therapy, and SPION in developing cancer diagnostics and therapeutics are discussed. STEM CELLS 2010; 28:1686–1702. PMID:20629172

  16. Influence of mitochondria on gene expression in a citrus cybrid.

    PubMed

    Bassene, Jean-Baptiste; Froelicher, Yann; Navarro, Luis; Ollitrault, Patrick; Ancillo, Gema

    2011-06-01

    The production of cybrids, combining nucleus of a species with alien cytoplasmic organelles, is a valuable method used for improvement of various crops. Several citrus cybrids have been created by somatic hybridization. These genotypes are interesting models to analyze the impact of cytoplasmic genome change on nuclear genome expression. Herein, we report genome-wide gene expression analysis in leaves of a citrus cybrid between C. reticulata cv 'Willowleaf mandarin' and C. limon cv 'Eureka lemon' compared with its lemon parent, using a Citrus 20K cDNA microarray. Molecular analysis showed that this cybrid possesses nuclear and chloroplast genomes of Eureka lemon plus mitochondria from Willowleaf mandarin and, therefore, can be considered as a lemon bearing foreign mitochondria. Mandarin mitochondria influenced the expression of a large set of lemon nuclear genes causing an over-expression of 480 of them and repression of 39 genes. Quantitative real-time RT-PCR further confirmed the credibility of microarray data. Genes over-expressed in cybrid leaves are predominantly attributed to the functional category "cellular protein metabolism" whereas in the down-regulated none functional category was enriched. Overall, mitochondria replacement affected different nuclear genes including particularly genes predicted to be involved in mitochondrial retrograde signaling. Mitochondria regulate all cell structures even chloroplast status. These results suggest that nuclear gene expression is modulated with respect to new information received from the foreign organelle, with the final objective to suit specific needs to ensure better cell physiological balance. PMID:21308470

  17. 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.

  18. Regulation of gene expression in the nervous system

    SciTech Connect

    Giuffrida Stella, A.M.; Perez-Polo, J.R.; deVellis, J.

    1990-01-01

    This book offers an up-to-date account of the latest research findings concerned with the regulatory mechanisms of gene expression in neuronal and glial cells under different conditions. The book explores the cellular and neurobiological aspects of important phenomena of the nervous system and its role in health, disease and injury. Contributions form prominent scientists in the field address a variety of specific topics concerned with gene expression in the nervous system - from growth, hormonal and trophic factors to neural tissue reactions in injury or aging.

  19. Macrophage Expression of Inflammatory Genes in Response to EMCV Infection

    PubMed Central

    Shaheen, Zachary R.; Corbett, John A.

    2015-01-01

    The expression and production of type 1 interferon is the classic cellular response to virus infection. In addition to this antiviral response, virus infection also stimulates the production of proinflammatory mediators. In this review, the pathways controlling the induction of inflammatory genes and the roles that these inflammatory mediators contribute to host defense against viral pathogens will be discussed. Specific focus will be on the role of the chemokine receptor CCR5, as a signaling receptor controlling the activation of pathways leading to virus-induced inflammatory gene expression. PMID:26295266

  20. 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. PMID:25214014

  1. Regulatory Nexus of Synthesis and Degradation Deciphers Cellular Nrf2 Expression Levels

    PubMed Central

    Suzuki, Takafumi; Shibata, Tatsuhiro; Takaya, Kai; Shiraishi, Kouya; Kohno, Takashi; Kunitoh, Hideo; Tsuta, Koji; Furuta, Koh; Goto, Koichi; Hosoda, Fumie; Sakamoto, Hiromi; Motohashi, Hozumi

    2013-01-01

    Transcription factor Nrf2 (NF-E2-related factor 2) is essential for oxidative and electrophilic stress responses. While it has been well characterized that Nrf2 activity is tightly regulated at the protein level through proteasomal degradation via Keap1 (Kelch-like ECH-associated protein 1)-mediated ubiquitination, not much attention has been paid to the supply side of Nrf2, especially regulation of Nrf2 gene transcription. Here we report that manipulation of Nrf2 transcription is effective in changing the final Nrf2 protein level and activity of cellular defense against oxidative stress even in the presence of Keap1 and under efficient Nrf2 degradation, determined using genetically engineered mouse models. In excellent agreement with this finding, we found that minor A/A homozygotes of a single nucleotide polymorphism (SNP) in the human NRF2 upstream promoter region (rs6721961) exhibited significantly diminished NRF2 gene expression and, consequently, an increased risk of lung cancer, especially those who had ever smoked. Our results support the notion that in addition to control over proteasomal degradation and derepression from degradation/repression, the transcriptional level of the Nrf2 gene acts as another important regulatory point to define cellular Nrf2 levels. These results thus verify the critical importance of human SNPs that influence the levels of transcription of the NRF2 gene for future personalized medicine. PMID:23572560

  2. Regulatory nexus of synthesis and degradation deciphers cellular Nrf2 expression levels.

    PubMed

    Suzuki, Takafumi; Shibata, Tatsuhiro; Takaya, Kai; Shiraishi, Kouya; Kohno, Takashi; Kunitoh, Hideo; Tsuta, Koji; Furuta, Koh; Goto, Koichi; Hosoda, Fumie; Sakamoto, Hiromi; Motohashi, Hozumi; Yamamoto, Masayuki

    2013-06-01

    Transcription factor Nrf2 (NF-E2-related factor 2) is essential for oxidative and electrophilic stress responses. While it has been well characterized that Nrf2 activity is tightly regulated at the protein level through proteasomal degradation via Keap1 (Kelch-like ECH-associated protein 1)-mediated ubiquitination, not much attention has been paid to the supply side of Nrf2, especially regulation of Nrf2 gene transcription. Here we report that manipulation of Nrf2 transcription is effective in changing the final Nrf2 protein level and activity of cellular defense against oxidative stress even in the presence of Keap1 and under efficient Nrf2 degradation, determined using genetically engineered mouse models. In excellent agreement with this finding, we found that minor A/A homozygotes of a single nucleotide polymorphism (SNP) in the human NRF2 upstream promoter region (rs6721961) exhibited significantly diminished NRF2 gene expression and, consequently, an increased risk of lung cancer, especially those who had ever smoked. Our results support the notion that in addition to control over proteasomal degradation and derepression from degradation/repression, the transcriptional level of the Nrf2 gene acts as another important regulatory point to define cellular Nrf2 levels. These results thus verify the critical importance of human SNPs that influence the levels of transcription of the NRF2 gene for future personalized medicine. PMID:23572560

  3. Dopamine receptor-mediated regulation of neuronal "clock" gene expression.

    PubMed

    Imbesi, M; Yildiz, S; Dirim Arslan, A; Sharma, R; Manev, H; Uz, T

    2009-01-23

    Using a transgenic mice model (i.e. "clock" knockouts), clock transcription factors have been suggested as critical regulators of dopaminergic behaviors induced by drugs of abuse. Moreover, it has been shown that systemic administration of psychostimulants, such as cocaine and methamphetamine regulates the striatal expression of clock genes. However, it is not known whether dopamine receptors mediate these regulatory effects of psychostimulants at the cellular level. Primary striatal neurons in culture express dopamine receptors as well as clock genes and have been successfully used in studying dopamine receptor functioning. Therefore, we investigated the role of dopamine receptors on neuronal clock gene expression in this model using specific receptor agonists. We found an inhibitory effect on the expression of mClock and mPer1 genes with the D2-class (i.e. D2/D3) receptor agonist quinpirole. We also found a generalized stimulatory effect on the expression of clock genes mPer1, mClock, mNPAS2 (neuronal PAS domain protein 2), and mBmal1 with the D1-class (i.e. D1) receptor agonist SKF38393. Further, we tested whether systemic administration of dopamine receptor agonists causes similar changes in striatal clock gene expression in vivo. We found quinpirole-induced alterations in mPER1 protein levels in the mouse striatum (i.e. rhythm shift). Collectively, our results indicate that the dopamine receptor system may mediate psychostimulant-induced changes in clock gene expression. Using striatal neurons in culture as a model, further research is needed to better understand how dopamine signaling modulates the expression dynamics of clock genes (i.e. intracellular signaling pathways) and thereby influences neuronal gene expression, neuronal transmission, and brain functioning. PMID:19017537

  4. 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.

  5. 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

  6. Gene Expression: Sizing it all up

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genomic architecture appears to be a largely unexplored component of gene expression. Although surely not the end of the story, we are learning that when it comes to gene expression, size is important. We have been surprised to find that certain patterns of expression, tissue-specific versus constit...

  7. 78 FR 79699 - Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-31

    ... 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..., Tissue, and Gene Therapies, Center for Biologics Evaluation and Research (CBER), FDA. On February...

  8. 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...

  9. 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...

  10. 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

  11. Serial Expression Analysis: a web tool for the analysis of serial gene expression data

    PubMed Central

    Nueda, Maria José; Carbonell, José; Medina, Ignacio; Dopazo, Joaquín; Conesa, Ana

    2010-01-01

    Serial transcriptomics experiments investigate the dynamics of gene expression changes associated with a quantitative variable such as time or dosage. The statistical analysis of these data implies the study of global and gene-specific expression trends, the identification of significant serial changes, the comparison of expression profiles and the assessment of transcriptional changes in terms of cellular processes. We have created the SEA (Serial Expression Analysis) suite to provide a complete web-based resource for the analysis of serial transcriptomics data. SEA offers five different algorithms based on univariate, multivariate and functional profiling strategies framed within a user-friendly interface and a project-oriented architecture to facilitate the analysis of serial gene expression data sets from different perspectives. SEA is available at sea.bioinfo.cipf.es. PMID:20525784

  12. 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

  13. 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.

  14. Methods for monitoring multiple gene expression

    DOEpatents

    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.

  15. Methods for monitoring multiple gene expression

    DOEpatents

    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.

  16. 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.

  17. 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. PMID:23407141

  18. 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

  19. Functional clustering of time series gene expression data by Granger causality

    PubMed Central

    2012-01-01

    Background A common approach for time series gene expression data analysis includes the clustering of genes with similar expression patterns throughout time. Clustered gene expression profiles point to the joint contribution of groups of genes to a particular cellular process. However, since genes belong to intricate networks, other features, besides comparable expression patterns, should provide additional information for the identification of functionally similar genes. Results In this study we perform gene clustering through the identification of Granger causality between and within sets of time series gene expression data. Granger causality is based on the idea that the cause of an event cannot come after its consequence. Conclusions This kind of analysis can be used as a complementary approach for functional clustering, wherein genes would be clustered not solely based on their expression similarity but on their topological proximity built according to the intensity of Granger causality among them. PMID:23107425

  20. Gene expression in major depressive disorder.

    PubMed

    Jansen, R; Penninx, B W J H; Madar, V; Xia, K; Milaneschi, Y; Hottenga, J J; Hammerschlag, A R; Beekman, A; van der Wee, N; Smit, J H; Brooks, A I; Tischfield, J; Posthuma, D; Schoevers, R; van Grootheest, G; Willemsen, G; de Geus, E J; Boomsma, D I; Wright, F A; Zou, F; Sun, W; Sullivan, P F

    2016-03-01

    The search for genetic variants underlying major depressive disorder (MDD) has not yet provided firm leads to its underlying molecular biology. A complementary approach is to study gene expression in relation to MDD. We measured gene expression in peripheral blood from 1848 subjects from The Netherlands Study of Depression and Anxiety. Subjects were divided into current MDD (N=882), remitted MDD (N=635) and control (N=331) groups. MDD status and gene expression were measured again 2 years later in 414 subjects. The strongest gene expression differences were between the current MDD and control groups (129 genes at false-discovery rate, FDR<0.1). Gene expression differences across MDD status were largely unrelated to antidepressant use, inflammatory status and blood cell counts. Genes associated with MDD were enriched for interleukin-6 (IL-6)-signaling and natural killer (NK) cell pathways. We identified 13 gene expression clusters with specific clusters enriched for genes involved in NK cell activation (downregulated in current MDD, FDR=5.8 × 10(-5)) and IL-6 pathways (upregulated in current MDD, FDR=3.2 × 10(-3)). Longitudinal analyses largely confirmed results observed in the cross-sectional data. Comparisons of gene expression results to the Psychiatric Genomics Consortium (PGC) MDD genome-wide association study results revealed overlap with DVL3. In conclusion, multiple gene expression associations with MDD were identified and suggest a measurable impact of current MDD state on gene expression. Identified genes and gene clusters are enriched with immune pathways previously associated with the etiology of MDD, in line with the immune suppression and immune activation hypothesis of MDD. PMID:26008736

  1. 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

  2. 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

  3. Analysis of Gene Expression Patterns Using Biclustering.

    PubMed

    Roy, Swarup; Bhattacharyya, Dhruba K; Kalita, Jugal K

    2016-01-01

    Mining microarray data to unearth interesting expression profile patterns for discovery of in silico biological knowledge is an emerging area of research in computational biology. A group of functionally related genes may have similar expression patterns under a set of conditions or at some time points. Biclustering is an important data mining tool that has been successfully used to analyze gene expression data for biologically significant cluster discovery. The purpose of this chapter is to introduce interesting patterns that may be observed in expression data and discuss the role of biclustering techniques in detecting interesting functional gene groups with similar expression patterns. PMID:26350227

  4. Xenbase: gene expression and improved integration.

    PubMed

    Bowes, Jeff B; Snyder, Kevin A; Segerdell, Erik; Jarabek, Chris J; Azam, Kenan; Zorn, Aaron M; Vize, Peter D

    2010-01-01

    Xenbase (www.xenbase.org), the model organism database for Xenopus laevis and X. (Silurana) tropicalis, is the principal centralized resource of genomic, development data and community information for Xenopus research. Recent improvements include the addition of the literature and interaction tabs to gene catalog pages. New content has been added including a section on gene expression patterns that incorporates image data from the literature, large scale screens and community submissions. Gene expression data are integrated into the gene catalog via an expression tab and is also searchable by multiple criteria using an expression search interface. The gene catalog has grown to contain over 15,000 genes. Collaboration with the European Xenopus Research Center (EXRC) has resulted in a stock center section with data on frog lines supplied by the EXRC. Numerous improvements have also been made to search and navigation. Xenbase is also the source of the Xenopus Anatomical Ontology and the clearinghouse for Xenopus gene nomenclature. PMID:19884130

  5. Hyperglycosylated HCG expression in pregnancy: cellular origin and clinical applications.

    PubMed

    Kovalevskaya, G; Kakuma, T; Schlatterer, J; O'Connor, J F

    2007-01-01

    Employing a monoclonal antibody (B152) specific for a carbohydrate epitope found on a choriocarcinoma derived hCG, it was discovered that a similar hCG isoform is expressed during early pregnancy. This form differs from later pregnancy hCG in carbohydrate moieties. Profiling of these two hCG isoforms throughout pregnancy utilized two IRMA's: B152-B207 ("hyperglycosylated hCG"-specific assay) and B109-B108 (an IRMA for standard intact hCG isoforms in the WHO hCG reference preparation). The WHO hCG standard was used in both assays. Values were presented as a ratio of hCG isoform concentrations (B152/B109 ratio). In early pregnancy urine concentrations of B152 hCG were significantly higher in normal pregnancy (NP) compared to early pregnancy loss (EPL). Matched serum-urine samples from the first and third trimesters revealed that the B152 hCG form is predominant in both serum and urine in the first trimester compared with the third trimester. The proportion of the B152 hCG (HhCG) form is higher in urine than in matched serum. There was a significant difference in the B152/B109 ratio between days 5 and 20 from time of embryo transfer in normally developing pregnancy versus EPL in the urine of IVF patients. In spontaneous abortion (SA) the level of B109 hCG remained higher in NP compared with SA. However, the B152/B109 ratio declined with gestational age faster in SA than in NP suggesting perhaps a different loss mechanism in SA versus EPL. The cellular origin of the different hCG glycoforms was identified by assay of cell media from cytotrophoblasts (CTBs) and syncytiotrophoblasts (STBs). Isolated CTBs expressed predominantly HhCG. The level of expression was the highest in the first trimester. STBs were the source of the less glycosylated B109 hCG isoform. Analysis of hCG glycoforms during early pregnancy can distinguish pregnancies that will fail from those that will proceed normally. Since the B152 assay does not effectively discriminate between intact HhCG and free

  6. 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. PMID:26324022

  7. HOXB homeobox gene expression in cervical carcinoma.

    PubMed

    López, R; Garrido, E; Piña, P; Hidalgo, A; Lazos, M; Ochoa, R; Salcedo, M

    2006-01-01

    The homeobox (HOX) genes are a family of transcription factors that bind to specific DNA sequences in target genes regulating gene expression. Thirty-nine HOX genes have been mapped in four conserved clusters: A, B, C, and D; they act as master genes regulating the identity of body segments along the anteroposterior axis of the embryo. The role played by HOX genes in adult cell differentiation is unclear to date, but growing evidence suggests that they may play an important role in the development of cancer. To study the role played by HOX genes in cervical cancer, in the present work, we analyzed the expression of HOXB genes and the localization of their transcripts in human cervical tissues. Reverse transcription-polymerase chain reaction analysis and nonradioactive RNA in situ hybridization were used to detect HOXB expression in 11 normal cervical tissues and 17 cervical carcinomas. It was determined that HOXB1, B3, B5, B6, B7, B8, and B9 genes are expressed in normal adult cervical epithelium and squamous cervical carcinomas. Interestingly, HOXB2, HOXB4, and HOXB13 gene expression was found only in tumor tissues. Our findings suggest that the new expression of HOXB2, HOXB4, and B13 genes is involved in cervical cancer. PMID:16445654

  8. Gene Expression Profiling of Gastric Cancer

    PubMed Central

    Marimuthu, Arivusudar; Jacob, Harrys K.C.; Jakharia, Aniruddha; Subbannayya, Yashwanth; Keerthikumar, Shivakumar; Kashyap, Manoj Kumar; Goel, Renu; Balakrishnan, Lavanya; Dwivedi, Sutopa; Pathare, Swapnali; Dikshit, Jyoti Bajpai; Maharudraiah, Jagadeesha; Singh, Sujay; Sameer Kumar, Ghantasala S; Vijayakumar, M.; Veerendra Kumar, Kariyanakatte Veeraiah; Premalatha, Chennagiri Shrinivasamurthy; Tata, Pramila; Hariharan, Ramesh; Roa, Juan Carlos; Prasad, T.S.K; Chaerkady, Raghothama; Kumar, Rekha Vijay; Pandey, Akhilesh

    2015-01-01

    Gastric cancer is the second leading cause of cancer death worldwide, both in men and women. A genomewide gene expression analysis was carried out to identify differentially expressed genes in gastric adenocarcinoma tissues as compared to adjacent normal tissues. We used Agilent’s whole human genome oligonucleotide microarray platform representing ~41,000 genes to carry out gene expression analysis. Two-color microarray analysis was employed to directly compare the expression of genes between tumor and normal tissues. Through this approach, we identified several previously known candidate genes along with a number of novel candidate genes in gastric cancer. Testican-1 (SPOCK1) was one of the novel molecules that was 10-fold upregulated in tumors. Using tissue microarrays, we validated the expression of testican-1 by immunohistochemical staining. It was overexpressed in 56% (160/282) of the cases tested. Pathway analysis led to the identification of several networks in which SPOCK1 was among the topmost networks of interacting genes. By gene enrichment analysis, we identified several genes involved in cell adhesion and cell proliferation to be significantly upregulated while those corresponding to metabolic pathways were significantly downregulated. The differentially expressed genes identified in this study are candidate biomarkers for gastric adenoacarcinoma. PMID:27030788

  9. Gene expression profiling in developing human hippocampus.

    PubMed

    Zhang, Yan; Mei, Pinchao; Lou, Rong; Zhang, Michael Q; Wu, Guanyun; Qiang, Boqin; Zhang, Zhengguo; Shen, Yan

    2002-10-15

    The gene expression profile of developing human hippocampus is of particular interest and importance to neurobiologists devoted to development of the human brain and related diseases. To gain further molecular insight into the developmental and functional characteristics, we analyzed the expression profile of active genes in developing human hippocampus. Expressed sequence tags (ESTs) were selected by sequencing randomly selected clones from an original 3'-directed cDNA library of 150-day human fetal hippocampus, and a digital expression profile of 946 known genes that could be divided into 16 categories was generated. We also used for comparison 14 other expression profiles of related human neural cells/tissues, including human adult hippocampus. To yield more confidence regarding differential expression, a method was applied to attach normalized expression data to genes with a low false-positive rate (<0.05). Finally, hierarchical cluster analysis was used to exhibit related gene expression patterns. Our results are in accordance with anatomical and physiological observations made during the developmental process of the human hippocampus. Furthermore, some novel findings appeared to be unique to our results. The abundant expression of genes for cell surface components and disease-related genes drew our attention. Twenty-four genes are significantly different from adult, and 13 genes might be developing hippocampus-specific candidate genes, including wnt2b and some Alzheimer's disease-related genes. Our results could provide useful information on the ontogeny, development, and function of cells in the human hippocampus at the molecular level and underscore the utility of large-scale, parallel gene expression analyses in the study of complex biological phenomena. PMID:12271469

  10. 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...