Sample records for upregulated cellular genes

  1. Expression of a GALACTINOL SYNTHASE Gene in Tomato Seeds Is Up-Regulated before Maturation

    E-print Network

    Bradford, Kent

    Expression of a GALACTINOL SYNTHASE Gene in Tomato Seeds Is Up-Regulated before Maturation of environmental stresses on plants. In seeds, proposed roles for RFOs include protecting cellular integrity during) seeds, and its expression was characterized in tomato seeds and seedlings. GOLS (LeGOLS-1) m

  2. Genes Upregulated in Human Fetal by Infection or Labor

    E-print Network

    Bryant-Greenwood, Gillian D.

    by The American College of Obstetricians and Gynecologists.) Preterm birth is a multifactorial disease, and itsGenes Upregulated in Human Fetal by Infection or Labor Membranes LILY S. TASHIMA, PhD, LYNNAE K. MILLAR, MD, AND GILLIAN D. BRYANT-GREENWOOD, PhD Objective: To determine whether suppression subtractive

  3. Progesterone Upregulates Gene Expression in Normal Human Thyroid Follicular Cells

    PubMed Central

    Bertoni, Ana Paula Santin; Brum, Ilma Simoni; Hillebrand, Ana Caroline; Furlanetto, Tania Weber

    2015-01-01

    Thyroid cancer and thyroid nodules are more prevalent in women than men, so female sex hormones may have an etiological role in these conditions. There are no data about direct effects of progesterone on thyroid cells, so the aim of the present study was to evaluate progesterone effects in the sodium-iodide symporter NIS, thyroglobulin TG, thyroperoxidase TPO, and KI-67 genes expression, in normal thyroid follicular cells, derived from human tissue. NIS, TG, TPO, and KI-67 mRNA expression increased significantly after TSH 20??UI/mL, respectively: 2.08 times, P < 0.0001; 2.39 times, P = 0.01; 1.58 times, P = 0.0003; and 1.87 times, P < 0.0001. In thyroid cells treated with 20??UI/mL TSH plus 10?nM progesterone, RNA expression of NIS, TG, and KI-67 genes increased, respectively: 1.78 times, P < 0.0001; 1.75 times, P = 0.037; and 1.95 times, P < 0.0001, and TPO mRNA expression also increased, though not significantly (1.77 times, P = 0.069). These effects were abolished by mifepristone, an antagonist of progesterone receptor, suggesting that genes involved in thyroid cell function and proliferation are upregulated by progesterone. This work provides evidence that progesterone has a direct effect on thyroid cells, upregulating genes involved in thyroid function and growth. PMID:26089899

  4. Progesterone Upregulates Gene Expression in Normal Human Thyroid Follicular Cells.

    PubMed

    Bertoni, Ana Paula Santin; Brum, Ilma Simoni; Hillebrand, Ana Caroline; Furlanetto, Tania Weber

    2015-01-01

    Thyroid cancer and thyroid nodules are more prevalent in women than men, so female sex hormones may have an etiological role in these conditions. There are no data about direct effects of progesterone on thyroid cells, so the aim of the present study was to evaluate progesterone effects in the sodium-iodide symporter NIS, thyroglobulin TG, thyroperoxidase TPO, and KI-67 genes expression, in normal thyroid follicular cells, derived from human tissue. NIS, TG, TPO, and KI-67 mRNA expression increased significantly after TSH 20??UI/mL, respectively: 2.08 times, P < 0.0001; 2.39 times, P = 0.01; 1.58 times, P = 0.0003; and 1.87 times, P < 0.0001. In thyroid cells treated with 20??UI/mL TSH plus 10?nM progesterone, RNA expression of NIS, TG, and KI-67 genes increased, respectively: 1.78 times, P < 0.0001; 1.75 times, P = 0.037; and 1.95 times, P < 0.0001, and TPO mRNA expression also increased, though not significantly (1.77 times, P = 0.069). These effects were abolished by mifepristone, an antagonist of progesterone receptor, suggesting that genes involved in thyroid cell function and proliferation are upregulated by progesterone. This work provides evidence that progesterone has a direct effect on thyroid cells, upregulating genes involved in thyroid function and growth. PMID:26089899

  5. Murine cytomegalovirus homologues of cellular immunomodulatory genes.

    PubMed

    Davis-Poynter, N J; Degli-Esposti, M; Farrell, H E

    1999-01-01

    The study of 'molecular mimicry' or 'genetic piracy', with respect to the utilisation of cellular genes captured and modified during the course of virus evolution, has been an area of increasing research with the expansion in virus genome sequencing. Examples of cellular immunomodulatory genes which have been captured from hosts have been identified in a number of viruses. This review concentrates upon studies of murine cytomegalovirus (MCMV), investigating the functions of viral genes homologous to G protein-coupled receptors, MHC class I and chemokines. The study of recombinant MCMV engineered with specific disruptions of these genes has revealed their significance during virus replication and dissemination within the host. In the case of the latter two classes of genes, evidence suggests they interfere with cellular immune responses, although the detailed mechanisms underlying this interference have yet to be delineated. PMID:10702715

  6. DNA stabilization by the upregulation of estrogen signaling in BRCA gene mutation carriers

    PubMed Central

    Suba, Zsuzsanna

    2015-01-01

    Currently available scientific evidence erroneously suggests that mutagenic weakness or loss of the BRCA1/2 genes may liberate the proliferative effects of estrogen signaling, which provokes DNA damage and genomic instability. Conversely, BRCA mutation seems to be an imbalanced defect, crudely inhibiting the upregulation of estrogen receptor expression and liganded transcriptional activity, whereas estrogen receptor-repressor functions become predominant. In BRCA-proficient cases, estrogen signaling orchestrates the activity of cell proliferation and differentiation with high safety, while upregulating the expression and DNA-stabilizing impact of BRCA genes. In turn, BRCA proteins promote estrogen signaling by proper estrogen synthesis via CYP19 gene regulation and by induction of the appropriate expression and transcriptional activity of estrogen receptors. In this exquisitely organized regulatory system, the dysfunction of each player may jeopardize genome stability and lead to severe chronic diseases, such as cancer development. Female organs, such as breast, endometrium, and ovary, exhibiting regular cyclic proliferative activity are particularly vulnerable in case of disturbances in either estrogen signaling or BRCA-mediated DNA repair. BRCA mutation carrier women may apparently be healthy or exhibit clinical signs of deficient estrogen signaling in spite of hyperestrogenism. Even women who enjoy sufficient compensatory DNA-defending activities are at risk of tumor development because many endogenous and environmental factors may jeopardize the mechanisms of extreme compensatory processes. Natural estrogens have numerous benefits in tumor prevention and therapy even in BRCA mutation carriers. There are no toxic effects even in sky-high doses and all physiologic cellular functions are strongly upregulated, while malignant tumor cells are recognized and killed in a Janus-faced manner. PMID:26028963

  7. Ginseng Berry Extract Prevents Atherogenesis via Anti-Inflammatory Action by Upregulating Phase II Gene Expression

    PubMed Central

    Kim, Chun-Ki; Cho, Dong Hui; Lee, Kyu-Sun; Lee, Dong-Keon; Park, Chan-Woong; Kim, Wan Gi; Lee, Sang Jun; Ha, Kwon-Soo; Goo Taeg, Oh; Kwon, Young-Guen; Kim, Young-Myeong

    2012-01-01

    Ginseng berry possesses higher ginsenoside content than its root, which has been traditionally used in herbal medicine for many human diseases, including atherosclerosis. We here examined the antiatherogenic effects of the Korean ginseng berry extract (KGBE) and investigated its underlying mechanism of action in vitro and in vivo. Administration of KGBE decreased atherosclerotic lesions, which was inversely correlated with the expression levels of phase II genes to include heme oxygenase-1 (HO-1) and glutamine-cysteine ligase (GCL). Furthermore, KGBE administration suppressed NF-?B-mediated expression of atherogenic inflammatory genes (TNF-?, IL-1?, iNOS, COX-2, ICAM-1, and VCAM-1), without altering serum cholesterol levels, in ApoE?/? mice fed a high fat-diet. Treatment with KGBE increased phase II gene expression and suppressed lipopolysaccharide-induced reactive oxygen species production, NF-?B activation, and inflammatory gene expression in primary macrophages. Importantly, these cellular events were blocked by selective inhibitors of HO-1 and GCL. In addition, these inhibitors reversed the suppressive effect of KGBE on TNF-?-mediated induction of ICAM-1 and VCAM-1, resulting in decreased interaction between endothelial cells and monocytes. These results suggest that KGBE ameliorates atherosclerosis by inhibiting NF-?B-mediated expression of atherogenic genes via upregulation of phase II enzymes and thus has therapeutic or preventive potential for atherosclerosis. PMID:23243449

  8. Zebrafish heat shock protein a4 genes in the intestinal epithelium are up-regulated during inflammation.

    PubMed

    Crawford, Katie C; Vega Flores, Maria; Oehlers, Stefan H; Hall, Christopher J; Crosier, Kathryn E; Crosier, Philip S

    2011-12-01

    A number of heat shock proteins (HSPs), including Hsp70 and Hsp110, function as molecular chaperones within intestinal epithelial cells that line the mammalian digestive system. HSPs confer cellular protection against environmental stress induced by chemical toxins or pathogens. There is interest in how members of this protein family might influence the progression of inflammatory bowel disease. Using the zebrafish model system, we report the expression of the duplicated hspa4 genes within the intestinal epithelium. The hspa4 genes belong to the Hsp110 family. We show that under inflammatory stress conditions within the gut, expression of these genes is up-regulated in a similar manner to that previously observed for mammalian Hsp70. Because of the amenability of the zebrafish to whole-animal screening protocols, the hspa4 genes could be used as effective read-outs for genetic, chemical and environmental factors that might influence intestinal inflammation. PMID:21557452

  9. Cellular microRNAs up-regulate transcription via interaction with promoter TATA-box motifs.

    PubMed

    Zhang, Yijun; Fan, Miaomiao; Zhang, Xue; Huang, Feng; Wu, Kang; Zhang, Junsong; Liu, Jun; Huang, Zhuoqiong; Luo, Haihua; Tao, Liang; Zhang, Hui

    2014-12-01

    The TATA box represents one of the most prevalent core promoters where the pre-initiation complexes (PICs) for gene transcription are assembled. This assembly is crucial for transcription initiation and well regulated. Here we show that some cellular microRNAs (miRNAs) are associated with RNA polymerase II (Pol II) and TATA box-binding protein (TBP) in human peripheral blood mononuclear cells (PBMCs). Among them, let-7i sequence specifically binds to the TATA-box motif of interleukin-2 (IL-2) gene and elevates IL-2 mRNA and protein production in CD4(+) T-lymphocytes in vitro and in vivo. Through direct interaction with the TATA-box motif, let-7i facilitates the PIC assembly and transcription initiation of IL-2 promoter. Several other cellular miRNAs, such as mir-138, mir-92a or mir-181d, also enhance the promoter activities via binding to the TATA-box motifs of insulin, calcitonin or c-myc, respectively. In agreement with the finding that an HIV-1-encoded miRNA could enhance viral replication through targeting the viral promoter TATA-box motif, our data demonstrate that the interaction with core transcription machinery is a novel mechanism for miRNAs to regulate gene expression. PMID:25336585

  10. Upregulation of the Coagulation Factor VII Gene during Glucose Deprivation Is Mediated by Activating Transcription Factor 4

    PubMed Central

    Cronin, Katherine R.; Mangan, Thomas P.; Carew, Josephine A.

    2012-01-01

    Background Constitutive production of blood coagulation proteins by hepatocytes is necessary for hemostasis. Stressful conditions trigger adaptive cellular responses and delay processing of most proteins, potentially affecting plasma levels of proteins secreted exclusively by hepatocytes. We examined the effect of glucose deprivation on expression of coagulation proteins by the human hepatoma cell line, HepG2. Methodology/Principal Findings Expression of coagulation factor VII, which is required for initiation of blood coagulation, was elevated by glucose deprivation, while expression of other coagulation proteins decreased. Realtime PCR and ELISA demonstrated that the relative percentage expression +/? SD of steady-state F7 mRNA and secreted factor VII antigen were significantly increased (from 100+/?15% to 188+/?27% and 100+/?8.8% to 176.3+/?17.3% respectively, p<0.001) at 24 hr of treatment. The integrated stress response was induced, as indicated by upregulation of transcription factor ATF4 and of additional stress-responsive genes. Small interfering RNAs directed against ATF4 potently reduced basal F7 expression, and prevented F7 upregulation by glucose deprivation. The response of the endogenous F7 gene was replicated in reporter gene assays, which further indicated that ATF4 effects were mediated via interaction with an amino acid response element in the F7 promoter. Conclusions/Significance Our data indicated that glucose deprivation enhanced F7 expression in a mechanism reliant on prior ATF4 upregulation primarily due to increased transcription from the ATF4 gene. Of five coagulation protein genes examined, only F7 was upregulated, suggesting that its functions may be important in a systemic response to glucose deprivation stress. PMID:22848420

  11. Transcriptional upregulation of the human MRP2 gene expression by serine/threonine protein kinase inhibitors.

    PubMed

    Pu?aski, L; Szemraj, J; Uchiumi, T; Kuwano, M; Bartosz, G

    2005-01-01

    Transcriptional regulation by cellular signalling pathways of multidrug resistance proteins that pump anticancer drugs out of cells is one of key issues in the development of the multidrug resistance phenotype. In our study, we have used the reporter gene approach as well as determination of mRNA levels in two cancer cell lines of human origin, MCF-7 and A549, to study the regulation of multidrug resistance proteins 2 and 3 (MRP2 AND MRP3) by serine/threonine protein kinases. Since a prototypic PKC inducer, PMA, caused a marked upregulation of transcription from both human MRP2 and MRP3 promoters, a role for PKC isoforms in positive control of expression of these proteins could be postulated. Interestingly, broad-spectrum serine-threonine protein kinase inhibitors which also inhibit PKC, staurosporine and H-7, stimulated expression from the MRP2 promoter instead of inhibiting it. This effect was not seen for MRP3. MRP2 induction by staurosporine and H-7 was shown to have phenotypic consequences in whole cells, rendering them more resistant to etoposide and increasing their ability to export calcein through the plasma membrane. These results point to the involvement of serine/threonine protein kinases in negative regulation of the human MRP2 gene and to the necessity of testing novel anti-cancer drugs acting as protein kinase inhibitors with regard to their potential ability to induce multidrug resistance. PMID:16602625

  12. Identification of a conserved set of upregulated genes in mouse skeletal muscle hypertrophy and regrowth.

    PubMed

    Chaillou, Thomas; Jackson, Janna R; England, Jonathan H; Kirby, Tyler J; Richards-White, Jena; Esser, Karyn A; Dupont-Versteegden, Esther E; McCarthy, John J

    2015-01-01

    The purpose of this study was to compare the gene expression profile of mouse skeletal muscle undergoing two forms of growth (hypertrophy and regrowth) with the goal of identifying a conserved set of differentially expressed genes. Expression profiling by microarray was performed on the plantaris muscle subjected to 1, 3, 5, 7, 10, and 14 days of hypertrophy or regrowth following 2 wk of hind-limb suspension. We identified 97 differentially expressed genes (?2-fold increase or ?50% decrease compared with control muscle) that were conserved during the two forms of muscle growth. The vast majority (?90%) of the differentially expressed genes was upregulated and occurred at a single time point (64 out of 86 genes), which most often was on the first day of the time course. Microarray analysis from the conserved upregulated genes showed a set of genes related to contractile apparatus and stress response at day 1, including three genes involved in mechanotransduction and four genes encoding heat shock proteins. Our analysis further identified three cell cycle-related genes at day and several genes associated with extracellular matrix (ECM) at both days 3 and 10. In conclusion, we have identified a core set of genes commonly upregulated in two forms of muscle growth that could play a role in the maintenance of sarcomere stability, ECM remodeling, cell proliferation, fast-to-slow fiber type transition, and the regulation of skeletal muscle growth. These findings suggest conserved regulatory mechanisms involved in the adaptation of skeletal muscle to increased mechanical loading. PMID:25554798

  13. Expression of the Abca-Subfamily of Genes in Abcc6-/- Mice – Upregulation of Abca4

    PubMed Central

    Li, Qiaoli; Uitto, Jouni

    2011-01-01

    Pseudoxanthoma elasticum (PXE), a heritable multi-system disorder, is caused by mutations in the ABCC6 gene primarily expressed in the liver. Recent analysis of cultured fibroblasts from patients with PXE has suggested compensatory alterations in the expression of the ABCA-subfamily of genes. We have now determined by quantitative RT-PCR the level of expression of Abca-family of genes in a mouse model of PXE developed by targeted ablation of Abcc6. The results indicated variable levels of mRNA for different Abca genes in the liver, however, only one of them, Abca4, was significantly, ~6.5-fold, upregulated in the Abcc6-/- mice in comparison to wild-type mice. In the same mice, Abca4 was not upregulated in the eyes or the kidney, suggesting that the upregulation of Abca4 in the liver is a tissue-specific compensatory consequence of the “knock-out” of Abcc6. PMID:21435020

  14. The glutathione peroxidase homologous gene from Chlamydomonas reinhardtii is transcriptionally up-regulated by singlet oxygen

    Microsoft Academic Search

    Urs Leisinger; Karin Rüfenacht; Beat Fischer; Manuel Pesaro; Arik Spengler; Alexander J. B. Zehnder; Rik I. L. Eggen

    2001-01-01

    The glutathione peroxidase homologous gene (Gpxh gene) in Chlamydomonas reinhardtii is up-regulated under oxidative stress conditions. The Gpxh gene showed a remarkably strong and fast induction by the singlet oxygen-generating photosensitizers neutral red, methylene blue and rose Bengal. The Gpxh mRNA levels strongly increased, albeit much more slowly, upon exposure to the organic hydroperoxides tert-butyl hydroperoxide (t-BOOH) and cumene hydroperoxide.

  15. 20-hydroxyecdysone upregulates Atg genes to induce autophagy in the Bombyx fat body

    PubMed Central

    Tian, Ling; Ma, Li; Guo, Enen; Deng, Xiaojuan; Ma, Sanyuan; Xia, Qingyou; Cao, Yang; Li, Sheng

    2013-01-01

    Autophagy is finely regulated at multiple levels and plays crucial roles in development and disease. In the fat body of the silkworm, Bombyx mori, autophagy occurs and Atg gene expression peaks during the nonfeeding molting and pupation stages when the steroid hormone (20-hydroxyecdysone; 20E) is high. Injection of 20E into the feeding larvae upregulated Atg genes and reduced TORC1 activity resulting in autophagy induction in the fat body. Conversely, RNAi knockdown of the 20E receptor partner (USP) or targeted overexpression of a dominant negative mutant of the 20E receptor (EcRDN) in the larval fat body reduced autophagy and downregulated the Atg genes, confirming the importance of 20E-induction of Atg gene expression during pupation. Moreover, in vitro treatments of the larval fat body with 20E upregulated the Atg genes. Five Atg genes were potentially 20E primary-responsive, and a 20E response element was identified in the Atg1 (ortholog of human ULK1) promoter region. Furthermore, RNAi knockdown of 4 key genes (namely Br-C, E74, HR3 and ?ftz-F1) in the 20E-triggered transcriptional cascade reduced autophagy and downregulated Atg genes to different levels. Taken together, we conclude that in addition to blocking TORC1 activity for autophagosome initiation, 20E upregulates Atg genes to induce autophagy in the Bombyx fat body. PMID:23674061

  16. Designer gene networks: Towards fundamental cellular control

    NASA Astrophysics Data System (ADS)

    Hasty, Jeff; Isaacs, Farren; Dolnik, Milos; McMillen, David; Collins, J. J.

    2001-03-01

    The engineered control of cellular function through the design of synthetic genetic networks is becoming plausible. Here we show how a naturally occurring network can be used as a parts list for artificial network design, and how model formulation leads to computational and analytical approaches relevant to nonlinear dynamics and statistical physics. We first review the relevant work on synthetic gene networks, highlighting the important experimental findings with regard to genetic switches and oscillators. We then present the derivation of a deterministic model describing the temporal evolution of the concentration of protein in a single-gene network. Bistability in the steady-state protein concentration arises naturally as a consequence of autoregulatory feedback, and we focus on the hysteretic properties of the protein concentration as a function of the degradation rate. We then formulate the effect of an external noise source which interacts with the protein degradation rate. We demonstrate the utility of such a formulation by constructing a protein switch, whereby external noise pulses are used to switch the protein concentration between two values. Following the lead of earlier work, we show how the addition of a second network component can be used to construct a relaxation oscillator, whereby the system is driven around the hysteresis loop. We highlight the frequency dependence on the tunable parameter values, and discuss design plausibility. We emphasize how the model equations can be used to develop design criteria for robust oscillations, and illustrate this point with parameter plots illuminating the oscillatory regions for given parameter values. We then turn to the utilization of an intrinsic cellular process as a means of controlling the oscillations. We consider a network design which exhibits self-sustained oscillations, and discuss the driving of the oscillator in the context of synchronization. Then, as a second design, we consider a synthetic network with parameter values near, but outside, the oscillatory boundary. In this case, we show how resonance can lead to the induction of oscillations and amplification of a cellular signal. Finally, we construct a toggle switch from positive regulatory elements, and compare the switching properties for this network with those of a network constructed using negative regulation. Our results demonstrate the utility of model analysis in the construction of synthetic gene regulatory networks.

  17. Identification of up-regulated genes in amphioxus neurula and the expression of AmphiFABP.

    PubMed

    Liu, Xiaohui; Sun, Yi; Zhang, Juyong; Li, Guang; Wang, Yiquan

    2011-01-01

    Amphioxus is a good model organism for understanding the origin and developmental mechanism of vertebrates owing to its important evolutionary position. During the developmental process of amphioxus embryo, the neurula is a crucial stage because of neural tube and notochord formation as well as somite emergence at this stage. In order to isolate genes up-regulated at the neurula stage, we constructed an 11-hour neurula subtracted cDNA library of amphioxus Branchiostoma belcheri and sequenced 204 ESTs representing 82 contigs. Comparative analysis revealed that 55% of those contigs were homologous to various known genes while 45% of them had no significant similarity to any known genes. Those observations imply that the un-identified ESTs might contain some new genes which are involved in the development of amphioxus neurula. Real-time quantitative PCR (RTqPCR) indicated that the expression levels of 14 genes are up-regulated after gastrulation among 20 assayed genes. Of those up-regulated genes, we further cloned and sequenced the full-length of fatty acid binding protein gene (AmphiFABP). The deduced protein sequence was similar to that of vertebrate brain FABP and heart FABP, and in situ hybridization displayed that AmphiFABP, similar to their vertebrate cognates, was expressed not only in nervous system but also in embryonic somite and gut, hinting a multifunctional property of AmphiFABP in amphioxus. PMID:21498921

  18. Gene up-regulation in response to predator kairomones in the water flea, Daphnia pulex

    PubMed Central

    2010-01-01

    Background Numerous cases of predator-induced polyphenisms, in which alternate phenotypes are produced in response to extrinsic stimuli, have been reported in aquatic taxa to date. The genus Daphnia (Branchiopoda, Cladocera) provides a model experimental system for the study of the developmental mechanisms and evolutionary processes associated with predator-induced polyphenisms. In D. pulex, juveniles form neckteeth in response to predatory kairomones released by Chaoborus larvae (Insecta, Diptera). Results Previous studies suggest that the timing of the sensitivity to kairomones in D. pulex can generally be divided into the embryonic and postembryonic developmental periods. We therefore examined which of the genes in the embryonic and first-instar juvenile stages exhibit different expression levels in the presence or absence of predator kairomones. Employing a candidate gene approach and identifying differentially-expressed genes revealed that the morphogenetic factors, Hox3, extradenticle and escargot, were up-regulated by kairomones in the postembryonic stage and may potentially be responsible for defense morph formation. In addition, the juvenile hormone pathway genes, JHAMT and Met, and the insulin signaling pathway genes, InR and IRS-1, were up-regulated in the first-instar stage. It is well known that these hormonal pathways are involved in physiological regulation following morphogenesis in many insect species. During the embryonic stage when morphotypes were determined, one of the novel genes identified by differential display was up-regulated, suggesting that this gene may be related to morphotype determination. Biological functions of the up-regulated genes are discussed in the context of defense morph formation. Conclusions It is suggested that, following the reception of kairomone signals, the identified genes are involved in a series of defensive phenotypic alterations and the production of a defensive phenotype. PMID:20433737

  19. Upregulated Genes In Sporadic, Idiopathic Pulmonary Arterial Hypertension

    Microsoft Academic Search

    Alasdair J Edgar; Matilde R Chacón; Anne E Bishop; Magdi H Yacoub; Julia M Polak

    2006-01-01

    BACKGROUND: To elucidate further the pathogenesis of sporadic, idiopathic pulmonary arterial hypertension (IPAH) and identify potential therapeutic avenues, differential gene expression in IPAH was examined by suppression subtractive hybridisation (SSH). METHODS: Peripheral lung samples were obtained immediately after removal from patients undergoing lung transplant for IPAH without familial disease, and control tissues consisted of similarly sampled pieces of donor lungs

  20. Biology Contribution Radiation-Induced Upregulation of Gene Expression From

    E-print Network

    Hemminki, Akseli

    ) was studied with and without radiation in three cell lines: breast cancer M4A4- LM3, prostate cancer PC-3MM2, Ph.D.,x Laura Ahtiainen, Ph.D.,*,y and Akseli Hemminki, M.D., Ph.D.*,y *Cancer Gene Therapy Group, Molecular Cancer Biology Program, Transplantation Laboratory, Haartman Institute, and Finnish Institute

  1. Expression Analysis of Up-Regulated Genes Responding to Plumbagin in Escherichia coli

    Microsoft Academic Search

    Jenn-Wei Chen; Chang-Ming Sun; Wei-Lun Sheng; Yu-Chen Wang; Wan-Jr Syu

    2006-01-01

    Plumbagin is found in many medicinal plants and has been reported to have antimicrobial activities. We examined the molecular responses of Escherichia coli to plumbagin by using a proteomic approach to search for bacterial genes up-regulated by the drug. The protein profile obtained was compared with that of E. coli without the plumbagin treatment. Subsequent analyses of the induced proteins

  2. Cellular senescence bypass screen identifies new putative tumor suppressor genes

    Microsoft Academic Search

    J F M Leal; J Fominaya; A Cascón; M V Guijarro; C Blanco-Aparicio; M Lleonart; M E Castro; S Ramon y Cajal; M Robledo; D H Beach; A Carnero

    2008-01-01

    Senescence is a mechanism that limits cellular lifespan and constitutes a barrier against cellular immortalization. To identify new senescence regulatory genes that might play a role in tumorigenesis, we have designed and performed a large-scale antisense-based genetic screen in primary mouse embryo fibroblasts (MEFs). Out of this screen, we have identified five different genes through which loss of function partially

  3. Mechanisms of Hypoxic Up-Regulation of Versican Gene Expression in Macrophages

    PubMed Central

    Sotoodehnejadnematalahi, Fattah; Staples, Karl J.; Chrysanthou, Elvina; Pearson, Helen; Ziegler-Heitbrock, Loems; Burke, Bernard

    2015-01-01

    Hypoxia is a hallmark of many pathological tissues. Macrophages accumulate in hypoxic sites and up-regulate a range of hypoxia-inducible genes. The matrix proteoglycan versican has been identified as one such gene, but the mechanisms responsible for hypoxic induction are not fully characterised. Here we investigate the up-regulation of versican by hypoxia in primary human monocyte-derived macrophages (HMDM), and, intriguingly, show that versican mRNA is up-regulated much more highly (>600 fold) by long term hypoxia (5 days) than by 1 day of hypoxia (48 fold). We report that versican mRNA decay rates are not affected by hypoxia, demonstrating that hypoxic induction of versican mRNA is mediated by increased transcription. Deletion analysis of the promoter identified two regions required for high level promoter activity of luciferase reporter constructs in human macrophages. The hypoxia-inducible transcription factor HIF-1 has previously been implicated as a key potential regulator of versican expression in hypoxia, however our data suggest that HIF-1 up-regulation is unlikely to be principally responsible for the high levels of induction observed in HMDM. Treatment of HMDM with two distinct specific inhibitors of Phosphoinositide 3-kinase (PI3K), LY290042 and wortmannin, significantly reduced induction of versican mRNA by hypoxia and provides evidence of a role for PI3K in hypoxic up-regulation of versican expression. PMID:26057378

  4. 75 FR 65640 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-26

    ...FDA-2010-N-0001] Cellular, Tissue and Gene Therapies Advisory Committee; Notice of...Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee. General...Branch, Office of Cellular, Tissue and Gene Therapies, Center for Biologics...

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

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-25

    ...Assessment of Investigational Cellular and Gene Therapy Products; Availability AGENCY...Assessment of Investigational Cellular and Gene Therapy Products'' dated November 2013...by the Office of Cellular, Tissue and Gene Therapies (OCTGT). The product...

  6. 78 FR 44133 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-23

    ...FDA-2013-N-0001] Cellular, Tissue and Gene Therapies Advisory Committee; Notice of...Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee. General...from the Office of Cellular, Tissue and Gene Therapies, Center for Biologics...

  7. 76 FR 22405 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-21

    ...FDA-2011-N-0002] Cellular, Tissue and Gene Therapies Advisory Committee; Notice of...Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee. General...the committee will discuss cellular and gene therapy products for the treatment of...

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

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-31

    ...FDA-2013-N-0001] Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of...of Committee: Cellular, Tissue, and Gene Therapies Advisory Committee. General...from the Office of Cellular, Tissue, and Gene Therapies, Center for Biologics...

  9. 76 FR 81513 - Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-28

    ...FDA-2011-N-0002] Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of...of Committee: Cellular, Tissue, and Gene Therapies Advisory Committee. General...Branch, Office of Cellular, Tissue and Gene Therapies, Center for Biologics...

  10. 76 FR 9028 - Guidance for Industry: Potency Tests for Cellular and Gene Therapy Products; Availability

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-02-16

    ...Industry: Potency Tests for Cellular and Gene Therapy Products; Availability AGENCY...Industry: Potency Tests for Cellular and Gene Therapy Products'' dated January 2011...provides manufacturers of cellular and gene therapy (CGT) products with...

  11. 77 FR 65693 - Cellular, Tissue and Gene Therapies Advisory Committee; Amendment of Notice

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-30

    ...FDA-2012-N-0001] Cellular, Tissue and Gene Therapies Advisory Committee; Amendment...a meeting of the Cellular, Tissue and Gene Therapies Advisory Committee. This meeting...a meeting of the Cellular, Tissue and Gene Therapies Advisory Committee would be...

  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

    ...Assessment of Investigational Cellular and Gene Therapy Products; Availability AGENCY...Assessment of Investigational Cellular and Gene Therapy Products,'' dated November 2012...CBER), Office of Cellular, Tissue, and Gene Therapies (OCTGT). The product...

  13. Peripheral challenge with a viral mimic upregulates expression of the complement genes in the hippocampus.

    PubMed

    Michalovicz, Lindsay T; Lally, Brent; Konat, Gregory W

    2015-08-15

    Peripheral challenge with a viral mimetic, polyinosinic-polycytidylic acid (PIC) induces hippocampal hyperexcitability in mice. Here, we characterized this hippocampal response through a whole genome transcriptome analysis. Intraperitoneal injection of PIC resulted in temporal dysregulation of 625 genes in the hippocampus, indicating an extensive genetic reprogramming. The bioinformatics analysis of these genes revealed the complement pathway to be the most significantly activated. The gene encoding complement factor B (CfB) exhibited the highest response, and its upregulation was commensurate with the development of hyperexcitability. Collectively, these results suggest that the induction of hippocampal hyperexcitability may be mediated by the alternative complement cascades. PMID:26198930

  14. Molecular cloning and functional analysis of a novel oncogene, cancer-upregulated gene 2 (CUG2)

    SciTech Connect

    Lee, Soojin [Department of Microbiology, Chungnam National University, Daejeon 305-764 (Korea, Republic of)]. E-mail: leesoojin@cnu.ac.kr; Gang, Jingu [Department of Internal Medicine, Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, Seoul (Korea, Republic of); Jeon, Sun Bok [LG Life Sciences, Ltd./R and D Park, Daejeon (Korea, Republic of); Choo, Seung Ho [LG Life Sciences, Ltd./R and D Park, Daejeon (Korea, Republic of); Lee, Bogman [LG Life Sciences, Ltd./R and D Park, Daejeon (Korea, Republic of); Kim, Young-Gun [LG Life Sciences, Ltd./R and D Park, Daejeon (Korea, Republic of); Lee, Yang Soon [Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-333 (Korea, Republic of); Jung, Jinyoung [Department of Microbiology, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Song, Si Young [Department of Internal Medicine, Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, Seoul (Korea, Republic of); Koh, Sang Seok [Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-333 (Korea, Republic of)]. E-mail: sskoh@kribb.re.kr

    2007-08-31

    We examined genome-wide differences in gene expression between tumor biopsies and normal tissues in order to identify differentially regulated genes in tumors. Cancer-upregulated gene 2 (CUG2) was identified as an expressed sequence tag (EST) that exhibits significant differential expression in multiple human cancer types. CUG2 showed weak sequence homology with the down-regulator of transcription 1 (DR1) gene, a human transcription repressor. We found that EGFP-CUG2 fusion proteins were predominantly localized in the nucleus, suggesting their putative role in gene regulation. In addition, CUG2-overexpressing mouse fibroblast cells exhibited distinct cancer-specific phenotypes in vitro and developed into tumors in nude mice. Taken together, these findings suggest that CUG2 is a novel tumor-associated gene that is commonly activated in various human cancers and exhibits high transforming activities; it possibly belongs to a transcription regulator family that is involved in tumor biogenesis.

  15. Social stress up-regulates inflammatory gene expression in the leukocyte transcriptome via ?-adrenergic induction of myelopoiesis

    PubMed Central

    Powell, Nicole D.; Sloan, Erica K.; Bailey, Michael T.; Arevalo, Jesusa M. G.; Miller, Gregory E.; Chen, Edith; Kobor, Michael S.; Reader, Brenda F.; Sheridan, John F.; Cole, Steven W.

    2013-01-01

    Across a variety of adverse life circumstances, such as social isolation and low socioeconomic status, mammalian immune cells have been found to show a conserved transcriptional response to adversity (CTRA) involving increased expression of proinflammatory genes. The present study examines whether such effects might stem in part from the selective up-regulation of a subpopulation of immature proinflammatory monocytes (Ly-6chigh in mice, CD16? in humans) within the circulating leukocyte pool. Transcriptome representation analyses showed relative expansion of the immature proinflammatory monocyte transcriptome in peripheral blood mononuclear cells from people subject to chronic social stress (low socioeconomic status) and mice subject to repeated social defeat. Cellular dissection of the mouse peripheral blood mononuclear cell transcriptome confirmed these results, and promoter-based bioinformatic analyses indicated increased activity of transcription factors involved in early myeloid lineage differentiation and proinflammatory effector function (PU.1, NF-?B, EGR1, MZF1, NRF2). Analysis of bone marrow hematopoiesis confirmed increased myelopoietic output of Ly-6chigh monocytes and Ly-6cintermediate granulocytes in mice subject to repeated social defeat, and these effects were blocked by pharmacologic antagonists of ?-adrenoreceptors and the myelopoietic growth factor GM-CSF. These results suggest that sympathetic nervous system-induced up-regulation of myelopoiesis mediates the proinflammatory component of the leukocyte CTRA dynamic and may contribute to the increased risk of inflammation-related disease associated with adverse social conditions. PMID:24062448

  16. Social stress up-regulates inflammatory gene expression in the leukocyte transcriptome via ?-adrenergic induction of myelopoiesis.

    PubMed

    Powell, Nicole D; Sloan, Erica K; Bailey, Michael T; Arevalo, Jesusa M G; Miller, Gregory E; Chen, Edith; Kobor, Michael S; Reader, Brenda F; Sheridan, John F; Cole, Steven W

    2013-10-01

    Across a variety of adverse life circumstances, such as social isolation and low socioeconomic status, mammalian immune cells have been found to show a conserved transcriptional response to adversity (CTRA) involving increased expression of proinflammatory genes. The present study examines whether such effects might stem in part from the selective up-regulation of a subpopulation of immature proinflammatory monocytes (Ly-6c(high) in mice, CD16(-) in humans) within the circulating leukocyte pool. Transcriptome representation analyses showed relative expansion of the immature proinflammatory monocyte transcriptome in peripheral blood mononuclear cells from people subject to chronic social stress (low socioeconomic status) and mice subject to repeated social defeat. Cellular dissection of the mouse peripheral blood mononuclear cell transcriptome confirmed these results, and promoter-based bioinformatic analyses indicated increased activity of transcription factors involved in early myeloid lineage differentiation and proinflammatory effector function (PU.1, NF-?B, EGR1, MZF1, NRF2). Analysis of bone marrow hematopoiesis confirmed increased myelopoietic output of Ly-6c(high) monocytes and Ly-6c(intermediate) granulocytes in mice subject to repeated social defeat, and these effects were blocked by pharmacologic antagonists of ?-adrenoreceptors and the myelopoietic growth factor GM-CSF. These results suggest that sympathetic nervous system-induced up-regulation of myelopoiesis mediates the proinflammatory component of the leukocyte CTRA dynamic and may contribute to the increased risk of inflammation-related disease associated with adverse social conditions. PMID:24062448

  17. Molecular crowding shapes gene expression in synthetic cellular nanosystems

    PubMed Central

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

    2013-01-01

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

  18. The longitudinal transcriptomic response of the substantia nigra to intrastriatal 6-hydroxydopamine reveals significant upregulation of regeneration-associated genes.

    PubMed

    Kanaan, Nicholas M; Collier, Timothy J; Cole-Strauss, Allyson; Grabinski, Tessa; Mattingly, Zachary R; Winn, Mary E; Steece-Collier, Kathy; Sortwell, Caryl E; Manfredsson, Fredric P; Lipton, Jack W

    2015-01-01

    We hypothesized that the study of gene expression at 1, 2, 4, 6 and 16 weeks in the substantia nigra (SN) after intrastriatal 6-OHDA in the Sprague-Dawley rat (rattus norvegicus) would identify cellular responses during the degenerative process that could be axoprotective. Specifically, we hypothesized that genes expressed within the SN that followed a profile of being highly upregulated early after the lesion (during active axonal degeneration) and then progressively declined to baseline over 16 weeks as DA neurons died are indicative of potential protective responses to the striatal 6-OHDA insult. Utilizing a ?-means cluster analysis strategy, we demonstrated that one such cluster followed this hypothesized expression pattern over time, and that this cluster contained several interrelated transcripts that are classified as regeneration-associated genes (RAGs) including Atf3, Sprr1a, Ecel1, Gadd45a, Gpnmb, Sox11, Mmp19, Srgap1, Rab15,Lifr, Trib3, Tgfb1, and Sema3c. All exemplar transcripts tested from this cluster (Sprr1a, Ecel1, Gadd45a, Atf3 and Sox11) were validated by qPCR and a smaller subset (Sprr1a, Gadd45a and Sox11) were shown to be exclusively localized to SN DA neurons using a dual label approach with RNAScope in situ hybridization and immunohistochemistry. Upregulation of RAGs is typically associated with the response to axonal injury in the peripheral nerves and was not previously reported as part of the axodegenerative process for DA neurons of the SN. Interestingly, as part of this cluster, other transcripts were identified based on their expression pattern but without a RAG provenance in the literature. These "RAG-like" transcripts need further characterization to determine if they possess similar functions to or interact with known RAG transcripts. Ultimately, it is hoped that some of the newly identified axodegeneration-reactive transcripts could be exploited as axoprotective therapies in PD and other neurodegenerative diseases. PMID:25992874

  19. Decoupling cellular memory from other gene expression characteristics

    NASA Astrophysics Data System (ADS)

    Balazsi, Gabor; Adams, Rhys; Nevozhay, Dmitry

    2010-03-01

    Non-conventional population level gene expression characteristics (such as the noise, cellular memory, skewness, modality, etc.) can have phenotypic impact and can affect cell population fitness independently of the gene expression mean. To study the phenotypic impact of gene expression characteristics other than the mean, they must be decoupled from the mean, and possibly from each other, i.e., two cell populations have to be established with similar means, but different non canonical gene expression characteristics. We study by experiment and mathematical modeling how positive feedback regulation can be used to decouple and adjust the cellular memory independently of the noise and the mean. We describe a state of ``population dynamic bistability'' where the cell population has bistable expression while individual cell lineages do not. Our results have implications for modeling gene expression bimodality and controlling cellular memory in cell populations.

  20. Naringenin confers protection against oxidative stress through upregulation of Nrf2 target genes in cardiomyoblast cells.

    PubMed

    Ramprasath, Tharmarajan; Senthamizharasi, Manivasagam; Vasudevan, Varadaraj; Sasikumar, Sundaresan; Yuvaraj, Subramani; Selvam, Govindan Sadasivam

    2014-06-01

    Cardiovascular diseases are the major health concern and the leading cause of death. Numerous studies have shown that oxidative stress stimuli have been incriminated in the pathogenesis of both acute and chronic heart disease. Though it is well known that bioflavonoids protect cells against reactive oxygen species (ROS)-induced damage, the molecular mechanisms involved are uncertain. Understanding the possible intracellular signaling pathways triggered by flavonoids will help to overcome the cardiac diseases resulting from oxidative stress. In the present study, we investigated whether naringenin (NGN) supplementation would improve the antioxidant defence under oxidative stress through the activation of Nrf2 signaling in cultured cardiomyoblast. NGN pretreatment significantly reduced stress-mediated apoptotic cell death and lipid peroxidation and showed increased level of reduced glutathione in H2O2-treated cardiomyoblast. In addition, NGN inhibited the production of NO and trigged the synthesis of antioxidant marker enzymes. Gene expression studies revealed that NGN upregulated the transcription of Akt and downregulated NF-?B and Caspase 3 genes. Notably, transcription of Nrf2 and its target genes was also upregulated. Taken together, the present study revealed that NGN elicits potent cytoprotective effect against oxidative stress by regulating Nrf2 and its target genes. In conclusion, the present work suggests that improving Nrf2 signaling by NGN supplementation would be a rational approach to facilitate ROS detoxification by augmenting both expression and activity of phase II detoxification enzymes for the alleviation of cardiac complications. PMID:24526395

  1. HMGB1 and HMGB2 proteins up-regulate cellular expression of human topoisomerase II

    Microsoft Academic Search

    Michal Stros; Eva Polanska ´; Sarka Pospisilova

    2009-01-01

    Topoisomerase IIa (topo IIa) is a nuclear enzyme involved in several critical processes, including chromosome replication, segregation and recom- bination. Previously we have shown that chromo- somal protein HMGB1 interacts with topo IIa, and stimulates its catalytic activity. Here we show the effect of HMGB1 on the activity of the human topo IIa gene promoter in different cell lines. We

  2. Rapid systemic up-regulation of genes after heat-wounding and electrical stimulation

    NASA Technical Reports Server (NTRS)

    Davies, E.; Vian, A.; Vian, C.; Stankovic, B.

    1997-01-01

    When one leaf of a tomato plant is electrically-stimulated or heat-wounded, proteinase inhibitor genes are rapidly up-regulated in distant leaves. The identity of the systemic wound signal(s) is not yet known, but major candidates include hormones transmitted via the phloem or the xylem, the electrically-stimulated self-propagating electrical signal in the phloem (the action potential, AP), or the heat-wound-induced surge in hydraulic pressure in the xylem evoking a local change in membrane potential in adjacent living cells (the variation potential, VP). In order to discriminate between these signals we have adopted two approaches. The first approach involves applying stimuli that evoke known signals and determining whether these signals have similar effects on the "model" transcripts for proteinase inhibitors (pin) and calmodulin (cal). Here we show that a heat wound almost invariably evokes a VP, while an electrical stimulation occasionally evokes an AP, and both of these signals induce accumulation of transcripts encoding proteinase inhibitors. The second approach involves identifying the array of genes turned on by heat-wounding. To this end, we have constructed a subtractive library for heat-wounded tissue, isolated over 800 putatively up-regulated clones, and shown that all but two of the fifty that we have analyzed by Northern hybridization are, indeed, up-regulated. Here we show the early kinetics of up-regulation of three of these transcripts in the terminal (4th) leaf in response to heat-wounding the 3rd leaf, about 5 cm away. Even though these transcripts show somewhat different time courses of induction, with one peaking at 30 min, another at 15 min, and another at 5 min after flaming of a distant leaf, they all exhibit a similar pattern, i.e., a transient period of transcript accumulation preceding a period of transcript decrease, followed by a second period of transcript accumulation.

  3. Expression of Autographa californica Multiple Nucleopolyhedrovirus Genes in Mammalian Cells and Upregulation of the Host ?-Actin Gene

    PubMed Central

    Fujita, Ryosuke; Matsuyama, Takahiro; Yamagishi, Junya; Sahara, Ken; Asano, Shinichiro; Bando, Hisanori

    2006-01-01

    The gene expression of Autographa californica multiple nucleopolyhedrovirus (AcMNPV) was examined in two types of mammalian cells, human HeLa14 and hamster BHK cells. DNA microarray analysis followed by reverse transcription-PCR identified at least 12 viral genes transcribed in both HeLa14 cells and BHK cells inoculated with AcMNPV. 5? rapid amplification of cDNA ends was carried out to examine the transcriptional fidelity of these genes in HeLa14 cells. The transcription of ie-1, ie-0 and gp64 was initiated at a baculovirus early gene motif, CAGT, accompanied by a TATA motif. In addition, the same splicing observed for ie-0 mRNA in Sf9 cells occurred in HeLa14 cells. While the transcription initiation sites for pe38 and p6.9 were not located in the CAGT motif, most of them were in a typical eukaryotic RNA polymerase II promoter structure (a conventional TATA motif and/or an initiator). Interestingly, the expression of ?-actin was upregulated in the mammalian cells inoculated with AcMNPV. Subsequent experiments using UV-inactivated virus confirmed the upregulation, suggesting that de novo synthesis of viral products is not required for the event. These results indicated that the AcMNPV genome acts as a template for transcription in mammalian cells through the usual infection pathway, though there is no evidence for the functional expression of viral genes at present. PMID:16474145

  4. Insecticide-Mediated Up-Regulation of Cytochrome P450 Genes in the Red Flour Beetle (Tribolium castaneum)

    PubMed Central

    Liang, Xiao; Xiao, Da; He, Yanping; Yao, Jianxiu; Zhu, Guonian; Zhu, Kun Yan

    2015-01-01

    Some cytochrome P450 (CYP) genes are known for their rapid up-regulation in response to insecticide exposures in insects. To date, however, limited information is available with respect to the relationships among the insecticide type, insecticide concentration, exposure duration and the up-regulated CYP genes. In this study, we examined the transcriptional response of eight selected CYP genes, including CYP4G7, CYP4Q4, CYP4BR3, CYP12H1, CYP6BK11, CYP9D4, CYP9Z5 and CYP345A1, to each of four insecticides in the red flour beetle, Tribolium castaneum. Reverse transcription quantitative PCR (RT-qPCR) revealed that CYP4G7 and CYP345A1 can be significantly up-regulated by cypermethrin (1.97- and 2.06-fold, respectively), permethrin (2.00- and 2.03-fold) and lambda-cyhalothrin (1.73- and 1.81-fold), whereas CYP4BR3 and CYP345A1 can be significantly up-regulated by imidacloprid (1.99- and 1.83-fold) when 20-day larvae were exposed to each of these insecticides at the concentration of LC20 for 24 h. Our studies also showed that similar levels of up-regulation can be achieved for CYP4G7, CYP4BR3 and CYP345A1 by cypermethrin, permethrin, lambda-cyhalothrin or imidacloprid with approximately one fourth of LC20 in 6 h. Our study demonstrated that up-regulation of these CYP genes was rapid and only required low concentrations of insecticides, and the up-regulation not only depended on the CYP genes but also the type of insecticides. Our results along with those from previous studies also indicated that there were no specific patterns for predicting the up-regulation of specific CYP gene families based on the insecticide classification. PMID:25607733

  5. Insecticide-mediated up-regulation of cytochrome P450 genes in the red flour beetle (Tribolium castaneum).

    PubMed

    Liang, Xiao; Xiao, Da; He, Yanping; Yao, Jianxiu; Zhu, Guonian; Zhu, Kun Yan

    2015-01-01

    Some cytochrome P450 (CYP) genes are known for their rapid up-regulation in response to insecticide exposures in insects. To date, however, limited information is available with respect to the relationships among the insecticide type, insecticide concentration, exposure duration and the up-regulated CYP genes. In this study, we examined the transcriptional response of eight selected CYP genes, including CYP4G7, CYP4Q4, CYP4BR3, CYP12H1, CYP6BK11, CYP9D4, CYP9Z5 and CYP345A1, to each of four insecticides in the red flour beetle, Tribolium castaneum. Reverse transcription quantitative PCR (RT-qPCR) revealed that CYP4G7 and CYP345A1 can be significantly up-regulated by cypermethrin (1.97- and 2.06-fold, respectively), permethrin (2.00- and 2.03-fold) and lambda-cyhalothrin (1.73- and 1.81-fold), whereas CYP4BR3 and CYP345A1 can be significantly up-regulated by imidacloprid (1.99- and 1.83-fold) when 20-day larvae were exposed to each of these insecticides at the concentration of LC20 for 24 h. Our studies also showed that similar levels of up-regulation can be achieved for CYP4G7, CYP4BR3 and CYP345A1 by cypermethrin, permethrin, lambda-cyhalothrin or imidacloprid with approximately one fourth of LC20 in 6 h. Our study demonstrated that up-regulation of these CYP genes was rapid and only required low concentrations of insecticides, and the up-regulation not only depended on the CYP genes but also the type of insecticides. Our results along with those from previous studies also indicated that there were no specific patterns for predicting the up-regulation of specific CYP gene families based on the insecticide classification. PMID:25607733

  6. 77 FR 63840 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-17

    ...FDA-2012-N-0001] Cellular, Tissue and Gene Therapies Advisory Committee; Notice of...Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee. General...hear updates of research programs in the Gene Transfer and Immunogenicity Branch,...

  7. 75 FR 66381 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-28

    ...FDA-2010-N-0001] Cellular, Tissue and Gene Therapies Advisory Committee; Notice of...Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee. General...Retroviral and Lentiviral Vector Based Gene Therapy Products. FDA intends to...

  8. The differentiation-related gene 1, Drg1, is markedly upregulated by androgens in LNCaP prostatic adenocarcinoma cells

    Microsoft Academic Search

    William Ulrix; Johannes V. Swinnen; Walter Heyns; Guido Verhoeven

    1999-01-01

    A differential display technique was used to identify androgen-regulated genes in LNCaP prostatic adenocarcinoma cells. One of the genes markedly upregulated by androgens proved to be identical to differentiation-related gene 1 (Drg1; also described as RTP, Cap43 and rit42), a gene whose expression has recently been shown to be diminished in colon, breast and prostate tumors. We show that Drg1

  9. Molecular cloning and functional characterization of a mouse gene upregulated by lipopolysaccharide treatment reveals alternative splicing

    SciTech Connect

    Du, Kejun; Chen, Yaoming; Dai, Zongming; Bi, Yuan; Cai, Tongjian [Department of Occupational and Environmental Health, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China)] [Department of Occupational and Environmental Health, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China); Hou, Lichao [Department of Anesthesiology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China)] [Department of Anesthesiology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China); Chai, Yubo; Song, Qinghe; Chen, Sumin [Department of Biochemistry and Molecular Biology, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China)] [Department of Biochemistry and Molecular Biology, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China); Luo, Wenjing, E-mail: luowenj@fmmu.edu.cn [Department of Occupational and Environmental Health, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China)] [Department of Occupational and Environmental Health, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China); Chen, Jingyuan, E-mail: jy_chen@fmmu.edu.cn [Department of Occupational and Environmental Health, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China)] [Department of Occupational and Environmental Health, Fourth Military Medical University, Xi'an 710032, Shaanxi Province (China)

    2010-01-01

    Treatment of mouse cells with lipopolysaccharide (LPS) potently initiates an inflammatory response, but the underlying mechanisms are unclear. We therefore sought to characterize cDNA sequences of a new mouse LPS-responsive gene, and to evaluate the effects of MLrg. Full-length cDNAs were obtained from LPS-treated NIH3T3 cells. We report that the MLrg gene produces two alternative splice products (GenBank Accession Nos. (DQ316984) and (DQ320011)), respectively, encoding MLrgW and MLrgS polypeptides. Both proteins contain zinc finger and leucine zipper domains and are thus potential regulators of transcription. Expression of MLrgW and MLrgS were robustly upregulated following LPS treatment, and the proteins were localized predominantly in the nuclear membrane and cytoplasm. In stable transfectants over-expressing MLrgW the proportion of cells in G1 phase was significantly reduced, while in cells over-expressing MLrgS the proportion of cells in G2 was significantly increased; both proteins are thus potential regulators of cell cycle progression. Upregulation of MLrgW and MLrgS may be an important component of the LPS inflammatory pathway and of the host response to infection with GNB.

  10. Egr-1, a master switch coordinating upregulation of divergent gene families underlying ischemic stress.

    PubMed

    Yan, S F; Fujita, T; Lu, J; Okada, K; Shan Zou, Y; Mackman, N; Pinsky, D J; Stern, D M

    2000-12-01

    Activation of the zinc-finger transcription factor early growth response (Egr)-1, initially linked to developmental processes, is shown here to function as a master switch activated by ischemia to trigger expression of pivotal regulators of inflammation, coagulation and vascular hyperpermeability. Chemokine, adhesion receptor, procoagulant and permeability-related genes are coordinately upregulated by rapid ischemia-mediated activation of Egr-1. Deletion of the gene encoding Egr-1 strikingly diminished expression of these mediators of vascular injury in a murine model of lung ischemia/reperfusion, and enhanced animal survival and organ function. Rapid activation of Egr-1 in response to oxygen deprivation primes the vasculature for dysfunction manifest during reperfusion. These studies define a central and unifying role for Egr-1 activation in the pathogenesis of ischemic tissue damage. PMID:11100120

  11. Gene expression profiling of endometrium versus bone marrow-derived mesenchymal stem cells: upregulation of cytokine genes.

    PubMed

    Gaafar, Taghrid; Osman, Omneya; Osman, Amira; Attia, Wael; Hamza, Hala; El Hawary, Rabab

    2014-10-01

    Postulated Stem/progenitor cells involved in endometrium regeneration are epithelial, mesenchymal, and endothelial. Bone marrow (BM) has been implicated in endometrial stem cells. We aimed at studying gene expression profiling of endometrial mesenchymal stem cells compared to BM MSCS to better understand their nature and functional phenotype. Endometrial tissues were obtained from premenopausal hysterectomies (n = 3), minced and enzymatically digested as well as Normal BM aspirates (n=3). Immunophenotyping, differentiation to mesoderm, and proliferation were studied. The expression profile of 84 genes relevant to mesenchymal stem cells was performed. Fold change calculations were determined with SA Biosciences data analysis software. VEGF, G-CSF, and GM-CSF in cultures supernatants of MSCs were assayed by Luminex immunoassay. Endo MSCs possess properties similar to BM MSCs. Cumulative population doubling was significantly higher in Endo MSCs compared to BM MSCs (p < 0.001). 52 core genes were shared between both generated MSCs including stemness, self-renewal, members of the Notch, TGFB, FGF, and WNT.16 downregulated genes (VCAM, IGF1)and 16 upregulated in Endo MSCs compared to BM (p < 0.05 ? fourfolds). They included mostly cytokine and growth factor genes G-CSF, GM-CSF, VWF, IL1b, GDF15, and KDR. VEGF and G-CSF levels were higher in Endo MSCs supernatants (p < 0.0001). Cells sharing MSC and endothelial cell characteristics could be isolated from the human endometrium. Endo MSCs share a core genetic profile with BM MSCs including stemness. They show upregulation of genes involved in vasculogenesis, angiogenesis, cell adhesion, growth proliferation, migration, and differentiation of endothelial cells, all contributing to endometrial function. PMID:24880484

  12. Transcription profiling during the cell cycle shows that a subset of Polycomb-targeted genes is upregulated during DNA replication

    PubMed Central

    Peña-Diaz, Javier; Hegre, Siv A.; Anderssen, Endre; Aas, Per A.; Mjelle, Robin; Gilfillan, Gregor D.; Lyle, Robert; Drabløs, Finn; Krokan, Hans E.; Sætrom, Pål

    2013-01-01

    Genome-wide gene expression analyses of the human somatic cell cycle have indicated that the set of cycling genes differ between primary and cancer cells. By identifying genes that have cell cycle dependent expression in HaCaT human keratinocytes and comparing these with previously identified cell cycle genes, we have identified three distinct groups of cell cycle genes. First, housekeeping genes enriched for known cell cycle functions; second, cell type-specific genes enriched for HaCaT-specific functions; and third, Polycomb-regulated genes. These Polycomb-regulated genes are specifically upregulated during DNA replication, and consistent with being epigenetically silenced in other cell cycle phases, these genes have lower expression than other cell cycle genes. We also find similar patterns in foreskin fibroblasts, indicating that replication-dependent expression of Polycomb-silenced genes is a prevalent but unrecognized regulatory mechanism. PMID:23325852

  13. Downregulation of protein tyrosine phosphatase PTPL1 alters cell cycle and upregulates invasion-related genes in prostate cancer cells.

    PubMed

    Castilla, Carolina; Flores, M Luz; Conde, José M; Medina, Rafael; Torrubia, Francisco J; Japón, Miguel A; Sáez, Carmen

    2012-04-01

    PTPL1, a non-receptor type protein tyrosine phosphatase, has been involved in the regulation of apoptosis and invasiveness of various tumour cell types, but its role in prostate cancer remained to be investigated. We report here that downregulation of PTPL1 by small interfering RNA in PC3 cells decreases cell proliferation and concomitantly reduces the expression of cell cycle-related proteins such as cyclins E and B1, PCNA, PTTG1 and phospho-histone H3. PTPL1 downregulation also increases the invasion ability of PC3 cells through Matrigel coated membranes. cDNA array of PTPL1-silenced PC3 cells versus control cells showed an upregulation of invasion-related genes such as uPA, uPAR, tPA, PAI-1, integrin ?6 and osteopontin. This increased expression was also confirmed in PTPL1-silenced DU145 prostate cancer cells by quantitative real time PCR and western blot. These findings suggest that PTPL1 is an important mediator of central cellular processes such as proliferation and invasion. PMID:22274591

  14. Histone acetylation associated up-regulation of the cell wall related genes is involved in salt stress induced maize root swelling

    PubMed Central

    2014-01-01

    Background Salt stress usually causes crop growth inhibition and yield decrease. Epigenetic regulation is involved in plant responses to environmental stimuli. The epigenetic regulation of the cell wall related genes associated with the salt-induced cellular response is still little known. This study aimed to analyze cell morphological alterations in maize roots as a consequence of excess salinity in relation to the transcriptional and epigenetic regulation of the cell wall related protein genes. Results In this study, maize seedling roots got shorter and displayed swelling after exposure to 200 mM NaCl for 48 h and 96 h. Cytological observation showed that the growth inhibition of maize roots was due to the reduction in meristematic zone cell division activity and elongation zone cell production. The enlargement of the stele tissue and cortex cells contributed to root swelling in the elongation zone. The cell wall is thought to be the major control point for cell enlargement. Cell wall related proteins include xyloglucan endotransglucosylase (XET), expansins (EXP), and the plasma membrane proton pump (MHA). RT-PCR results displayed an up-regulation of cell wall related ZmEXPA1, ZmEXPA3, ZmEXPA5, ZmEXPB1, ZmEXPB2 and ZmXET1 genes and the down-regulation of cell wall related ZmEXPB4 and ZmMHA genes as the duration of exposure was increased. Histone acetylation is regulated by HATs, which are often correlated with gene activation. The expression of histone acetyltransferase genes ZmHATB and ZmGCN5 was increased after 200 mM NaCl treatment, accompanied by an increase in the global acetylation levels of histones H3K9 and H4K5. ChIP experiment showed that the up-regulation of the ZmEXPB2 and ZmXET1 genes was associated with the elevated H3K9 acetylation levels on the promoter regions and coding regions of these two genes. Conclusions These data suggested that the up-regulation of some cell wall related genes mediated cell enlargement to possibly mitigate the salinity-induced ionic toxicity, and different genes had specific function in response to salt stress. Histone modification as a mediator may contribute to rapid regulation of cell wall related gene expression, which reduces the damage of excess salinity to plants. PMID:24758373

  15. Depletion of the xynB2 gene upregulates ?-xylosidase expression in C. crescentus.

    PubMed

    Corrêa, Juliana Moço; Mingori, Moara Rodrigues; Gandra, Rinaldo Ferreira; Loth, Eduardo Alexandre; Seixas, Flávio Augusto Vicente; Simão, Rita de Cássia Garcia

    2014-01-01

    Caulobacter crescentus is able to express several enzymes involved in the utilization of lignocellulosic biomasses. Five genes, xynB1-5, that encode ?-xylosidases are present in the genome of this bacterium. In this study, the xynB2 gene, which encodes ?-xylosidase II (CCNA_02442), was cloned under the control of the PxylX promoter to generate the O-xynB2 strain, which overexpresses the enzyme in the presence of xylose. In addition, a null mutant strain, ?-xynB2, was created by two homologous recombination events where the chromosomal xynB2 gene was replaced by a copy that was disrupted by the spectinomycin-resistant cassette. We demonstrated that C. crescentus cells lacking ?-xylosidase II upregulates the xynB genes inducing ?-xylosidase activity. Transcriptional analysis revealed that xynB1 (RT-PCR analysis) and xynB2 (lacZ transcription fusion) gene expression was induced in the ?-xynB2 cells, and high ?-xylosidase activity was observed in the presence of different agro-industrial residues in the null mutant strain, a characteristic that can be explored and applied in biotechnological processes. In contrast, overexpression of the xynB2 gene caused downregulation of the expression and activity of the ?-xylosidase. For example, the ?-xylosidase activity that was obtained in the presence of sugarcane bagasse was 7-fold and 16-fold higher than the activity measured in the C. crescentus parental and O-xynB2 cells, respectively. Our results suggest that ?-xylosidase II may have a role in controlling the expression of the xynB1 and xynB2 genes in C. crescentus. PMID:24142353

  16. Interleukin-6 upregulates paraoxonase 1 gene expression via an AKT/NF-?B-dependent pathway

    SciTech Connect

    Cheng, Chi-Chih [Department of Research, Taichung Veterans General Hospital, Taichung, Taiwan (China)] [Department of Research, Taichung Veterans General Hospital, Taichung, Taiwan (China); Hsueh, Chi-Mei [Department of Life Sciences, National Chung Hsing University, Taichung, Taiwan (China)] [Department of Life Sciences, National Chung Hsing University, Taichung, Taiwan (China); Chen, Chiu-Yuan [Graduate Institute of Natural Healing Sciences, Nanhua University, Chiayi, Taiwan (China)] [Graduate Institute of Natural Healing Sciences, Nanhua University, Chiayi, Taiwan (China); Chen, Tzu-Hsiu, E-mail: hsiu@mail.chna.edu.tw [Department of Health and Nutrition, Chia Nan University of Pharmacy and Science, Tainan, Taiwan (China)] [Department of Health and Nutrition, Chia Nan University of Pharmacy and Science, Tainan, Taiwan (China); Hsu, Shih-Lan, E-mail: h2326@vghtc.gov.tw [Department of Research, Taichung Veterans General Hospital, Taichung, Taiwan (China) [Department of Research, Taichung Veterans General Hospital, Taichung, Taiwan (China); Department of Applied Chemistry, National Chi Nan University, Puli, Nantou, Taiwan (China)

    2013-07-19

    Highlights: •IL-6 could induce PON1 gene expression. •IL-6 increased NF-?B protein expression and NF-?B-p50 and -p65 subunits nuclear translocation. •IL-6-induced PON1 up-regulation was through an AKT/NF-?B pathway. -- Abstract: The aim of this study is to investigate the relationship between paraoxonase 1 (PON1) and atherosclerosis-related inflammation. In this study, human hepatoma HepG2 cell line was used as a hepatocyte model to examine the effects of the pro-inflammatory cytokines on PON1 expression. The results showed that IL-6, but not TNF-? and IL-1?, significantly increased both the function and protein level of PON1; data from real-time RT-PCR analysis revealed that the IL-6-induced PON1 expression occurred at the transcriptional level. Increase of I?B kinase activity and I?B phosphorylation, and reduction of I?B protein level were also observed in IL-6-treated HepG2 cells compared with untreated culture. This event was accompanied by increase of NF-?B-p50 and -p65 nuclear translocation. Moreover, treatment with IL-6 augmented the DNA binding activity of NF-?B. Furthermore, pharmacological inhibition of NF-?B activation by PDTC and BAY 11-7082, markedly suppressed the IL-6-mediated PON1 expression. In addition, IL-6 increased the levels of phosphorylated protein kinase B (PKB, AKT). An AKT inhibitor LY294002 effectively suppressed IKK/I?B/NF-?B signaling and PON1 gene expression induced by IL-6. Our findings demonstrate that IL-6 upregulates PON1 gene expression through an AKT/NF-?B signaling axis in human hepatocyte-derived HepG2 cell line.

  17. Identification of putative immunologic targets for colon cancer prevention based on conserved gene upregulation from pre-invasive to malignant lesions

    PubMed Central

    Broussard, Elizabeth K.; Kim, Rachel; Wiley, Jesse C.; Marquez, Juan Pablo; Annis, James E.; Pritchard, David; Disis, Mary L.

    2013-01-01

    The length of time required for pre-invasive adenoma (AD) to progress to carcinoma, the immunogenicity of colorectal cancer (CRC), and the identification of high risk populations make development and testing of a prophylactic vaccine for the prevention of CRC possible. We hypothesized that genes upregulated in AD relative to normal tissue, which maintained increased expression in CRC, would encode proteins suitable as putative targets for immunoprevention. We evaluated existing AD and CRC microarray datasets and identified 160 genes that were 2-fold up-regulated in both AD and CRC relative to normal colon tissue. We further identified 23 genes that demonstrated protein over-expression in colon AD and CRC based on literature review. Silencing the most highly up-regulated genes, CDH3, CLDN1, KRT23, and MMP7, in AD and CRC cell lines resulted in a significant decrease in viability (p<0.0001) and proliferation (p<0.0001) as compared to controls and an increase in cellular apoptosis (p<0.05 for CDH3, KRT23). Results were duplicated across cell lines representing microsatellite instability (MSI), CpG island methylator (CIMP) and chromosomal instability (CIN) phenotypes suggesting immunologic elimination of cells expressing these proteins could impact the progression of all CRC phenotypes. To determine whether these proteins were immunogens, we interrogated sera from early stage CRC patients and controls and found significantly elevated CDH3 (p=0.006), KRT23 (p=0.0007), and MMP7 (p<0.0001) serum IgG in cases as compared to controls. These data demonstrate a high throughput approach to the identification of biologically relevant putative immunologic targets for CRC and identified 3 candidates suitable for vaccine development. PMID:23682078

  18. Isolation and characterization of a novel gene sfig in rat skeletal muscle up-regulated by spaceflight (STS-90).

    PubMed

    Kano, Mihoko; Kitano, Takako; Ikemoto, Madoka; Hirasaka, Katsuya; Asanoma, Yuki; Ogawa, Takayuki; Takeda, Shinichi; Nonaka, Ikuya; Adams, Gregory R; Baldwin, Kenneth M; Oarada, Motoko; Kishi, Kyoichi; Nikawa, Takeshi

    2003-02-01

    We obtained the skeletal muscle of rats exposed to weightless conditions during a 16-day-spaceflight (STS-90). By using a differential display technique, we identified 6 up-regulated and 3 down-regulated genes in the gastrocnemius muscle of the spaceflight rats, as compared to the ground control. The up-regulated genes included those coding Casitas B-lineage lymphoma-b, insulin growth factor binding protein-1, titin and mitochondrial gene 16 S rRNA and two novel genes (function unknown). The down-regulated genes included those encoding RNA polymerase II elongation factor-like protein, NADH dehydrogenase and one novel gene (function unknown). In the present study, we isolated and characterized one of two novel muscle genes that were remarkably up-regulated by spaceflight. The deduced amino acid sequence of the spaceflight-induced gene (sfig) comprises 86 amino acid residues and is well conserved from Drosophila to Homo sapiens. A putative leucine-zipper structure located at the N-terminal region of sfig suggests that this gene may encode a transcription factor. The up-regulated expression of this gene, confirmed by Northern blot analysis, was observed not only in the muscles of spaceflight rats but also in the muscles of tail-suspended rats, especially in the early stage of tail-suspension when gastrocnemius muscle atrophy initiated. The gene was predominantly expressed in the kidney, liver, small intestine and heart. When rat myoblastic L6 cells were grown to 100% confluence in the cell culture system, the expression of sfig was detected regardless of the cell differentiation state. These results suggest that spaceflight has many genetic effects on rat skeletal muscle. PMID:12630567

  19. 77 FR 73472 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-12-10

    ...Administration [Docket No. FDA-2012-N-0001] Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting AGENCY: Food...closed to the public. Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee. General Function of the...

  20. 76 FR 64951 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-10-19

    ...Administration [Docket No. FDA-2011-N-0002] Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting AGENCY: Food...open to the public. Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee. General Function of the...

  1. 78 FR 15726 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-12

    ...Administration [Docket No. FDA-2013-N-0001] Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting AGENCY: Food...closed to the public. Name of Committee: Cellular, Tissue and Gene Therapies Advisory Committee. General Function of the...

  2. 76 FR 18768 - Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-05

    ...Docket No. FDA-2011-N-0002] Cellular, Tissue, and Gene Therapies Advisory Committee; Notice of Meeting AGENCY: Food...closed to the public. Name of Committee: Cellular, Tissue, and Gene Therapies Advisory Committee. General Function of the...

  3. 76 FR 49774 - Cellular, Tissue and Gene Therapies Advisory Committee; Notice of Meeting

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-11

    ...Cellular, Tissue and Gene Therapies Advisory Committee...Cellular, Tissue and Gene Therapies Advisory Committee...thalassemia, Hurler syndrome, Krabbe disease, and...default.htm. Scroll down to the appropriate...

  4. Upregulation of TRAG3 gene in urothelial carcinoma of the bladder

    PubMed Central

    Karam, Jose A.; Huang, Sandra; Fan, Jinhai; Stanfield, Jennifer; Schultz, Roger A.; Pong, Rey-Chen; Sun, Xiankai; Mason, Ralph P.; Xie, Xian-Jin; Niu, Gang; Chen, Xiaoyuan; Frenkel, Eugene P.; Sagalowsky, Arthur I.; Hsieh, Jer-Tsong

    2010-01-01

    Conventional chemotherapy is commonly used for advanced stages of bladder cancer with modest success and high morbidity. Identifying markers of resistance will allow clinicians to tailor treatment to a specific patient population. T24-tumorigenic cell line was grown orthotopically in nude mice and monitored using bioluminescence imaging and microcomputed tomography until they developed metastases. Stable sublines were then developed from primary bladder (T24-P), lung (T24-L) and bone (T24-B) tissues. Chromosomal analysis and DNA microarray were used to characterize these sublines. qRT-PCR and immunohistochemistry (IHC) were used for validation. Epigenetic modifiers were used to study gene regulation. The cell viability was quantified with MTT assay. Chromosomal analysis revealed multiple alterations in metastatic cell lines compared to T24-P. DNA microarray analysis showed that Taxol-Resistance-Associated-Gene-3 (TRAG3) gene was the most upregulated gene. From qRT-PCR and IHC, TRAG3 was significantly higher in T24-L and T24-B than T24-P. TRAG3 gene expression is likely controlled by DNA methylation, but not histone acetylation. Interestingly, T24-B and T24-L cells were more resistant than T24-P to treatment with anti-microtubule agents such as docetaxel, paclitaxel and vinblastine. TRAG3 mRNA expression was higher in 20% of patients with ?pT2 (n=10) and 60% of patients with ?pT3 (n=20) compared to normal adjacent tissue (p=0.05). In addition, the median TRAG3 expression was 6.7-fold higher in ?pT3 tumors compared to ?pT2 tumors. Knowing the status of TRAG3 expression could help clinicians tailor treatment to a particular patient population that could benefit from treatment, while allocating patients with resistant tumors to new experimental therapies. PMID:20734393

  5. Increasing cancer-specific gene expression by targeting overexpressed ?5?1 integrin and upregulated transcriptional activity of NF-?B.

    PubMed

    Adil, Maroof M; Levine, Rachel M; Kokkoli, Efrosini

    2014-03-01

    We developed a modular multifunctional nonviral gene delivery system by targeting the overexpressed cancer surface receptor ?5?1 integrin and the upregulated transcriptional activity of the cancer resistance mediating transcription factor NF-?B, thereby introducing a new form of transcriptional targeting. NF-?B regulated therapy can improve specificity of gene expression in cancer tissue and also may offset NF-?B mediated cancer resistance. We delivered a luciferase gene under the control of an NF-?B responsive element (pNF-?B-Luc) encapsulated in a PR_b peptide functionalized stealth liposome that specifically targets the ?5?1 integrin and achieved increased gene expression in DLD-1 colorectal cancer cells compared to BJ-fibroblast healthy cells in vitro. The multitargeted system was also able to differentiate between cancer cells and healthy cells better than either of the individually targeted systems. In addition, we constructed a novel cancer therapeutic plasmid by cloning a highly potent diphtheria toxin fragment A (DTA) expressing gene under the control of an NF-?B responsive element (pNF-?B-DTA). A dose-dependent reduction of cellular protein expression and increased cytotoxicity in cancer cells was seen when transfected with PR_b functionalized stealth liposomes encapsulating the condensed pNF-?B-DTA plasmid. Our therapeutic delivery system specifically eradicated close to 70% of a variety of cancer cells while minimally affecting healthy cells in vitro. Furthermore, the modular nature of the nonviral design allows targeting novel pairs of extracellular receptors and upregulated transcription factors for applications beyond cancer gene therapy. PMID:24483950

  6. De-repressing LncRNA-Targeted Genes to Upregulate Gene Expression: Focus on Small Molecule Therapeutics

    PubMed Central

    Fatemi, Roya Pedram; Velmeshev, Dmitry; Faghihi, Mohammad Ali

    2014-01-01

    Non-protein coding RNAs (ncRNAs) make up the overwhelming majority of transcripts in the genome and have recently gained attention for their complex regulatory role in cells, including the regulation of protein-coding genes. Furthermore, ncRNAs play an important role in normal development and their expression levels are dysregulated in several diseases. Recently, several long noncoding RNAs (lncRNAs) have been shown to alter the epigenetic status of genomic loci and suppress the expression of target genes. This review will present examples of such a mechanism and focus on the potential to target lncRNAs for achieving therapeutic gene upregulation by de-repressing genes that are epigenetically silenced in various diseases. Finally, the potential to target lncRNAs, through their interactions with epigenetic enzymes, using various tools, such as small molecules, viral vectors and antisense oligonucleotides, will be discussed. We suggest that small molecule modulators of a novel class of drug targets, lncRNA-protein interactions, have great potential to treat some cancers, cardiovascular disease, and neurological disorders. PMID:25405465

  7. Microarray Analysis of Gene Expression Reveals that Cyclo-oxygenase-2 Gene Therapy Up-regulates Hematopoiesis and Down-regulates Inflammation During Endochondral Bone Fracture Healing

    PubMed Central

    Lau, K.-H. William; Popa, Nicoleta L.

    2014-01-01

    Background Cyclo-oxygenase-2 (Cox-2) is an inflammatory mediator that is necessary for the tissue repair, including bone fracture healing. Although the application of Cox-2 gene therapy to a murine closed femoral fracture has accelerated bony union, but the beneficial effect was not observed until the endochondral stage of bone repair that is well after the inflammatory stage normally subsides. Methods To identify the molecular pathways through which Cox-2 regulates fracture healing, we examined gene expression profile in fracture tissues in response to Cox-2 gene therapy during the endochondral bone repair phase. Cox-2 gene therapy was applied to the closed murine femur fracture model. Microarray analysis was performed at 10 days post-fracture to examine global gene expression profile in the fracture tissues during the endochondral bone repair phase. The entire repertoire of significantly expressed genes was examined by gene set enrichment analysis, and the most up-regulated individual genes were evaluated further. Results The genes that normally promote inflammation were under-represented in the microarray analysis, and the expression of several inflammatory chemokines was significantly down-regulated. There was an up-regulation of two key transcription factor genes that regulate hematopoiesis and erythropoiesis. More surprisingly, there was no significant up-regulation in the genes that are normally involved in angiogenesis or bone formation. However, the expression of two tissue remodeling genes was up-regulated. Conclusions The down-regulation of the inflammatory genes in response to Cox-2 gene therapy was unexpected, given the pro-inflammatory role of prostaglandins. Cox-2 gene therapy could promote bony union through hematopoietic precursor proliferation during endochondral bone repair and thereby enhances subsequently fracture callus remodeling that leads to bony union of the fracture gap. PMID:25247155

  8. Identification of Potentially Neuroprotective Genes Upregulated by Neurotrophin Treatment of CA3 Neurons in the Injured Brain

    PubMed Central

    Malik, Saafan Z.; Motamedi, Shahab; Royo, Nicolas C.; LeBold, David

    2011-01-01

    Abstract Specific neurotrophic factors mediate histological and/or functional improvement in animal models of traumatic brain injury (TBI). In previous work, several lines of evidence indicated that the mammalian neurotrophin NT-4/5 is neuroprotective for hippocampal CA3 pyramidal neurons after experimental TBI. We hypothesized that NT-4/5 neuroprotection is mediated by changes in the expression of specific sets of genes, and that NT-4/5-regulated genes are potential therapeutic targets for blocking delayed neuronal death after TBI. In this study, we performed transcription profiling analysis of CA3 neurons to identify genes regulated by lateral fluid percussion injury, or by treatment with the trkB ligands NT-4/5 or brain-derived neurotrophic factor (BDNF). The results indicate extensive overlap between genes upregulated by neurotrophins and genes upregulated by injury, suggesting that the mechanism behind neurotrophin neuroprotection may mimic the brain's endogenous protective response. A subset of genes selected for further study in vitro exhibited neuroprotection against glutamate excitotoxicity. The neuroprotective genes identified in this study were upregulated at 30?h post-injury, and are thus expected to act during a clinically useful time frame of hours to days after injury. Modulation of these factors and pathways by genetic manipulation or small molecules may confer hippocampal neuroprotection in vivo in preclinical models of TBI. PMID:21083427

  9. Sampling-Dependent Up-regulation of Gene Expression in Sequential Samples of Human Airway Epithelial Cells

    PubMed Central

    Heguy, Adriana; Harvey, Ben-Gary; O’Connor, Timothy P; Hackett, Neil R; Crystal, Ronald G

    2003-01-01

    As part of a study of in vivo gene expression levels in the human airway epithelium in response to chronic cigarette smoking, we have identified a number of genes whose expression levels are altered in a time-dependent fashion resulting from the procedure used to sample epithelial cells. Fiberoptic bronchoscopy and airway epithelium brushing were used to obtain independent samples from a single individual, 1st from the right lung, followed by sampling of the left lung. We observed that a specific subset of early response genes encoding proteins involved in transcription, signal transduction, cell cycle/growth, and apoptosis were significantly up-regulated in the left lung samples (the 2nd region to be sampled) compared with the right lung samples (the 1st region to be sampled). This response was due to the temporal nature of the sampling procedure and not to inherent gene expression differences between airway epithelium of the right and left lungs. When the order of sampling was reversed, with the left airway epithelium sampled 1st, the same subset of genes were up-regulated in the samples obtained from the right airway epithelium. The time-dependent up-regulation of these genes was likely in response to the stress of the procedure and/or the anesthesia used. Sampling-dependent uncertainty of gene expression is likely a general phenomenon relevant to the procedures used for obtaining biological samples, particularly in humans where the sampling procedures are dependent on ensuring comfort and safety. PMID:15208741

  10. 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

  11. Upregulation of TRAG3 gene in urothelial carcinoma of the bladder.

    PubMed

    Karam, Jose A; Huang, Sandra; Fan, Jinhai; Stanfield, Jennifer; Schultz, Roger A; Pong, Rey-Chen; Sun, Xiankai; Mason, Ralph P; Xie, Xian-Jin; Niu, Gang; Chen, Xiaoyuan; Frenkel, Eugene P; Sagalowsky, Arthur I; Hsieh, Jer-Tsong

    2011-06-15

    Conventional chemotherapy is commonly used for advanced stages of bladder cancer with modest success and high morbidity. Identifying markers of resistance will allow clinicians to tailor treatment to a specific patient population. T24-tumorigenic cell line was grown orthotopically in nude mice and monitored using bioluminescence imaging and microcomputed tomography until they developed metastases. Stable sublines were then developed from primary bladder (T24-P), lung (T24-L) and bone (T24-B) tissues. Chromosomal analysis and DNA microarray were used to characterize these sublines. Real-time quantitative polymerase chain reaction and immunohistochemistry were used for validation. Epigenetic modifiers were used to study gene regulation. The cell viability was quantified with MTT assay. Chromosomal analysis revealed multiple alterations in metastatic cell lines compared to T24-P. DNA microarray analysis showed that taxol resistance-associated gene (TRAG) 3 was the most upregulated gene. From real-time quantitative polymerase chain reaction and immunohistochemistry, TRAG3 was significantly higher in T24-L and T24-B than T24-P. TRAG3 gene expression is likely controlled by DNA methylation but not histone acetylation. Interestingly, T24-B and T24-L cells were more resistant than T24-P to treatment with antimicrotubule agents such as docetaxel, paclitaxel and vinblastine. TRAG3 mRNA expression was higher in 20% of patients with ? pT2 (n = 10) and 60% of patients with ? pT3 (n = 20) compared to normal adjacent tissue (p = 0.05). In addition, the median TRAG3 expression was 6.7-fold higher in ? pT3 tumors compared to ? pT2 tumors. Knowing the status of TRAG3 expression could help clinicians tailor treatment to a particular patient population that could benefit from treatment, while allocating patients with resistant tumors to new experimental therapies. PMID:20734393

  12. Increased expression of peripheral blood leukocyte genes implicate CD14+ tissue macrophages in cellular intestine allograft rejection.

    PubMed

    Ashokkumar, Chethan; Ningappa, Mylarappa; Ranganathan, Sarangarajan; Higgs, Brandon W; Sun, Qing; Schmitt, Lori; Snyder, Sara; Dobberstein, Jennifer; Branca, Maria; Jaffe, Ronald; Zeevi, Adriana; Squires, Robert; Alissa, Feras; Shneider, Benjamin; Soltys, Kyle; Bond, Geoffrey; Abu-Elmagd, Kareem; Humar, Abhinav; Mazariegos, George; Hakonarson, Hakon; Sindhi, Rakesh

    2011-10-01

    Recurrent rejection shortens graft survival after intestinal transplantation (ITx) in children, most of whom also experience early acute cellular rejection (rejectors). To elucidate mechanisms common to early and recurrent rejection, we used a test cohort of 20 recipients to test the hypothesis that candidate peripheral blood leukocyte genes that trigger rejection episodes would be evident late after ITx during quiescent periods in genome-wide gene expression analysis and would achieve quantitative real-time PCR replication pre-ITx (another quiescent period) and in the early post-ITx period during first rejection episodes. Eight genes were significantly up-regulated among rejectors in the late post-ITx and pre-ITx periods, compared with nonrejectors: TBX21, CCL5, GNLY, SLAMF7, TGFBR3, NKG7, SYNE1, and GK5. Only CCL5 was also up-regulated in the early post-ITx period. Among resting peripheral blood leukocyte subsets in randomly sampled nonrejectors, CD14(+) monocytes expressed the CCL5 protein maximally. Compared with nonrejectors, rejectors demonstrated higher counts of both circulating CCL5(+)CD14(+) monocytes and intragraft CD14(+) monocyte-derived macrophages in immunohistochemistry of postperfusion and early post-ITx biopsies from the test and an independent replication cohort. Donor-specific alloreactivity measured with CD154(+) T-cytotoxic memory cells correlated with the CCL5 gene and intragraft CD14(+) monocyte-derived macrophages at graft reperfusion and early post-ITx. CCL5 gene up-regulation and CD14(+) macrophages likely prime cellular ITx rejection. Infiltration of reperfused intestine allografts with CD14(+) macrophages may predict rejection events. PMID:21854741

  13. Ribosomal RNA gene repeats, their stability and cellular senescence

    PubMed Central

    KOBAYASHI, Takehiko

    2014-01-01

    The ribosomal RNA gene (rDNA) repeats form a historically well-researched region in the chromosome. Their highly repetitive structure can be identified easily which has enabled studies on DNA replication, recombination, and transcription. The region is one of the most unstable regions in the genome because of deleterious recombination among the repeats. The ribosomal RNA gene repeats use a unique gene amplification system to restore the copy number after this has been reduced due to recombination. It has been shown that unstable features in the genome can accelerate cellular senescence that restricts the lifespan of a cell. Here, I will introduce a study by our group that shows how the stability of rDNA is maintained and affects lifespan. I propose that the ribosomal RNA gene repeats constitute a center from which the stability of the whole genome is regulated and the lifespan of the cell is controlled. PMID:24727936

  14. Upregulation of human PINK1 gene expression by NF?B signalling

    PubMed Central

    2014-01-01

    Parkinson’s disease (PD) is one of the major neurodegenerative disorders. Mitochondrial malfunction is implicated in PD pathogenesis. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN)-induced putative kinase 1 (PINK1), a serine/threonine kinase, plays an important role in the quality control of mitochondria and more than 70 PINK1 mutations have been identified to cause early-onset PD. However, the regulation of PINK1 gene expression remains elusive. In the present study, we identified the transcription start site (TSS) of the human PINK1 gene using switching mechanism at 5’end of RNA transcription (SMART RACE) assay. The TSS is located at 91 bp upstream of the translation start site ATG. The region with 104 bp was identified as the minimal promoter region by deletion analysis followed by dual luciferase assay. Four functional cis-acting nuclear factor kappa-light-chain-enhancer of activated B cells (NF?B)-binding sites within the PINK1 promoter were identified. NF?B overexpression led to the up-regulation of PINK1 expression in both HEK293 cells and SH-SY5Y cells. Consistently, lipopolysaccharide (LPS), a strong activator of NF?B, significantly increased PINK1 expression in SH-SY5Y cells. Taken together, our results clearly suggested that PINK1 expression is tightly regulated at its transcription level and NF?B is a positive regulator for PINK1 expression. PMID:25108683

  15. Efficacy of QCDCR formulated CpG ODN 2007 in Nile tilapia against Streptococcus iniae and identification of upregulated genes.

    PubMed

    Pridgeon, Julia W; Klesius, Phillip H; Mu, Xingjiang; Yancey, Robert J; Kievit, Michele S; Dominowski, Paul J

    2012-01-15

    The potential of using a QCDCR (quilA:cholesterol:dimethyl dioctadecyl ammonium bromide:carbopol:R1005 glycolipid) formulated CpG oligodeoxynucleotide (ODN), ODN 2007, to confer protection in Nile tilapia against Streptococcus iniae infection was evaluated in this study. At two days post treatment, QCDCR formulated ODN 2007 elicited significant (P<0.05) protection to Nile tilapia, with relative percent survival of 63% compared to fish treated by QCDCR alone. To understand the molecular mechanisms involved in the protective immunity elicited by ODN 2007, suppression subtractive cDNA hybridization technique was used to identify upregulated genes induced by ODN 2007. A total of 69 expressed sequence tags (ESTs) were identified from the subtractive cDNA library. Quantitative PCR revealed that 44 ESTs were significantly (P<0.05) upregulated by ODN 2007, including 29 highly (>10-fold) and 15 moderately (<10-fold) upregulated ESTs. Of all ESTs, putative peroxisomal sarcosine oxidase was upregulated the highest. The 69 ESTs only included six genes that had putative functions related to immunity, of which only two (putative glutaredoxin-1 and carboxypeptidase N catalytic chain) were confirmed to be significantly upregulated. Our results suggest that the protection elicited by ODN 2007 is mainly through innate immune responses directly or indirectly related to immunity. PMID:22129787

  16. Loss of E-cadherin promotes prostate cancer metastasis via upregulation of metastasis-associated gene 1 expression.

    PubMed

    Fan, Liangsheng; Wang, Hongyan; Xia, Xi; Rao, Yumei; Ma, Xiangyi; Ma, Ding; Wu, Peng; Chen, Gang

    2012-12-01

    E-cadherin is a key cell-to-cell adhesion molecule associated with the invasion and metastasis of tumor cells; however, the molecular mechanisms are not entirely understood. In this study, we investigated whether downregulation of E-cadherin by E-cadherin-specific small intefering RNA (siRNA) was able to promote malignant phenotypes of prostate cancer cells through upregulating the metastasis-associated gene 1 (MTA1) in vitro. The expression levels of E-cadherin in human paired prostate adenocarcinoma cell lines, PC-3M-2B4 (2B4) and PC-3M-1E8 (1E8), were investigated using western blot analysis. The alteration of malignant phenotypes associated with decreasing E-cadherin expression were assessed in 2B4 cells using wound-healing assays, solid-phase adhesion assays, invasion assays and cytoskeletal staining. The expression of E-cadherin and MTA1 in normal, localized and metastatic prostate cancer cells was analyzed using immunohistochemistry. Downregulation of E-cadherin using an RNA interference approach led to the upregulation of MTA1 expression, decreased tumor cell adhesion ability as well as enhanced cell mobility, invasion and cellular polarity compared with the controls (P<0.05). E-cadherin regulated MTA1 in a time-dependent manner. The correlation between E-cadherin and MTA1 was inversed in the prostate cancer group (P<0.05; r(s)=-0.434). The data suggest that E-cadherin plays an important role in prostate cancer metastasis, which is likely to be due to the regulation of MTA1 expression. E-cadherin may combine with MTA1 and alter the malignant phenotype of prostate cancer cells. A combined testing strategy for detecting MTA1 and E-cadherin may be sufficient for selecting high-risk patients with metastasis. Therefore, E-cadherin and MTA1 may be potential powerful factors for the treatment of various types of cancer. PMID:23205121

  17. Methylene blue upregulates Nrf2/ARE genes and prevents tau-related neurotoxicity.

    PubMed

    Stack, Cliona; Jainuddin, Shari; Elipenahli, Ceyhan; Gerges, Meri; Starkova, Natalia; Starkov, Anatoly A; Jové, Mariona; Portero-Otin, Manuel; Launay, Nathalie; Pujol, Aurora; Kaidery, Navneet Ammal; Thomas, Bobby; Tampellini, Davide; Beal, M Flint; Dumont, Magali

    2014-07-15

    Methylene blue (MB, methylthioninium chloride) is a phenothiazine that crosses the blood brain barrier and acts as a redox cycler. Among its beneficial properties are its abilities to act as an antioxidant, to reduce tau protein aggregation and to improve energy metabolism. These actions are of particular interest for the treatment of neurodegenerative diseases with tau protein aggregates known as tauopathies. The present study examined the effects of MB in the P301S mouse model of tauopathy. Both 4 mg/kg MB (low dose) and 40 mg/kg MB (high dose) were administered in the diet ad libitum from 1 to 10 months of age. We assessed behavior, tau pathology, oxidative damage, inflammation and numbers of mitochondria. MB improved the behavioral abnormalities and reduced tau pathology, inflammation and oxidative damage in the P301S mice. These beneficial effects were associated with increased expression of genes regulated by NF-E2-related factor 2 (Nrf2)/antioxidant response element (ARE), which play an important role in antioxidant defenses, preventing protein aggregation, and reducing inflammation. The activation of Nrf2/ARE genes is neuroprotective in other transgenic mouse models of neurodegenerative diseases and it appears to be an important mediator of the neuroprotective effects of MB in P301S mice. Moreover, we used Nrf2 knock out fibroblasts to show that the upregulation of Nrf2/ARE genes by MB is Nrf2 dependent and not due to secondary effects of the compound. These findings provide further evidence that MB has important neuroprotective effects that may be beneficial in the treatment of human neurodegenerative diseases with tau pathology. PMID:24556215

  18. Meta-analysis of heat- and chemically upregulated chaperone genes in plant and human cells

    PubMed Central

    Finka, Andrija; Mattoo, Rayees U. H.

    2010-01-01

    Molecular chaperones are central to cellular protein homeostasis. In mammals, protein misfolding diseases and aging cause inflammation and progressive tissue loss, in correlation with the accumulation of toxic protein aggregates and the defective expression of chaperone genes. Bacteria and non-diseased, non-aged eukaryotic cells effectively respond to heat shock by inducing the accumulation of heat-shock proteins (HSPs), many of which molecular chaperones involved in protein homeostasis, in reducing stress damages and promoting cellular recovery and thermotolerance. We performed a meta-analysis of published microarray data and compared expression profiles of HSP genes from mammalian and plant cells in response to heat or isothermal treatments with drugs. The differences and overlaps between HSP and chaperone genes were analyzed, and expression patterns were clustered and organized in a network. HSPs and chaperones only partly overlapped. Heat-shock induced a subset of chaperones primarily targeted to the cytoplasm and organelles but not to the endoplasmic reticulum, which organized into a network with a central core of Hsp90s, Hsp70s, and sHSPs. Heat was best mimicked by isothermal treatments with Hsp90 inhibitors, whereas less toxic drugs, some of which non-steroidal anti-inflammatory drugs, weakly expressed different subsets of Hsp chaperones. This type of analysis may uncover new HSP-inducing drugs to improve protein homeostasis in misfolding and aging diseases. Electronic supplementary material The online version of this article (doi:10.1007/s12192-010-0216-8) contains supplementary material, which is available to authorized users. PMID:20694844

  19. Non-Thermal Plasma Treatment Diminishes Fungal Viability and Up-Regulates Resistance Genes in a Plant Host

    PubMed Central

    Panngom, Kamonporn; Lee, Sang Hark; Park, Dae Hoon; Sim, Geon Bo; Kim, Yong Hee; Uhm, Han Sup; Park, Gyungsoon; Choi, Eun Ha

    2014-01-01

    Reactive oxygen and nitrogen species can have either harmful or beneficial effects on biological systems depending on the dose administered and the species of organism exposed, suggesting that application of reactive species can possibly produce contradictory effects in disease control, pathogen inactivation and activation of host resistance. A novel technology known as atmospheric-pressure non-thermal plasma represents a means of generating various reactive species that adversely affect pathogens (inactivation) while simultaneously up-regulating host defense genes. The anti-microbial efficacy of this technology was tested on the plant fungal pathogen Fusarium oxysporum f.sp. lycopersici and its susceptible host plant species Solanum lycopercicum. Germination of fungal spores suspended in saline was decreased over time after exposed to argon (Ar) plasma for 10 min. Although the majority of treated spores exhibited necrotic death, apoptosis was also observed along with the up-regulation of apoptosis related genes. Increases in the levels of peroxynitrite and nitrite in saline following plasma treatment may have been responsible for the observed spore death. In addition, increased transcription of pathogenesis related (PR) genes was observed in the roots of the susceptible tomato cultivar (S. lycopercicum) after exposure to the same Ar plasma dose used in fungal inactivation. These data suggest that atmospheric-pressure non-thermal plasma can be efficiently used to control plant fungal diseases by inactivating fungal pathogens and up-regulating mechanisms of host resistance. PMID:24911947

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

    Microsoft Academic Search

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

    2004-01-01

    Gene trap vectors developed for genome-wide muta- genesis can be used to study factors governing the expression of exons inserted throughout the genome. For example, entrapment vectors consist- ing of a partial 3¢-terminal exon (i.e. a neomycin resistance gene (Neo), a poly(A) site, but no 3¢ splice site) were typically expressed following insertion into introns, from cellular transcripts that spliced

  1. 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

  2. Genes involved in carnitine synthesis and carnitine uptake are up-regulated in the liver of sows during lactation

    PubMed Central

    2013-01-01

    Background Convincing evidence exist that carnitine synthesis and uptake of carnitine into cells is regulated by peroxisome proliferator-activated receptor ? (PPARA), a transcription factor which is physiologically activated during fasting or energy deprivation. Sows are typically in a negative energy balance during peak lactation. We investigated the hypothesis that genes involved in carnitine synthesis and uptake in the liver of sows are up-regulated during peak lactation. Findings Transcript levels of several PPAR? target genes involved in fatty acid uptake (FABP4, SLC25A20), fatty acid oxidation (ACOX1, CYP4A24) and ketogenesis (HMGCS2, FGF21) were elevated in the liver of lactating compared to non-lactating sows (P < 0.05). In addition, transcript levels of genes involved in carnitine synthesis (ALDH9A1, TMLHE, BBOX1) and carnitine uptake (SLC22A5) in the liver were greater in lactating than in non-lactating sows (P < 0.05). Carnitine concentrations in liver and plasma were about 20% and 50%, respectively, lower in lactating than in non-lactating sows (P < 0.05), which is likely due to an increased loss of carnitine via the milk. Conclusions The results of the present study show that PPAR? is activated in the liver of sows during lactation which leads to an up-regulation of genes involved in carnitine synthesis and carnitine uptake. The PPAR? mediated up-regulation of genes involved in carnitine synthesis and uptake in the liver of lactating sows may be regarded as an adaptive mechanism to maintain hepatic carnitine levels at a level sufficient to transport excessive amounts of fatty acids into the mitochondrion. PMID:23497718

  3. Cellular zinc sensors: MTF-1 regulation of gene expression.

    PubMed

    Andrews, G K

    2001-01-01

    Zinc metabolism in higher eukaryotes is complex, being controlled by uptake, efflux, and storage in individual cells, as well as in peripheral tissues and organs. Recently there have been advances in the understanding of the genes involved in these processes and their regulation. Metal-response element-binding transcription factor-1 (MTF-1) functions as a cellular zinc sensor which coordinates the expression of genes involved in zinc homeostasis, as well as protection against metal toxicity and oxidative stresses. In mice, these are known to include the metallothionein (MT), the zinc-transporter-1 (ZnT1) and the gamma-glutamylcysteine synthetase heavy chain (gammaGCShc) genes. The cysteine-rich MTs function as an intracellular metal-chelators that bind zinc with high affinity, whereas the transmembrane protein ZnT1 exports zinc from the cell. Gamma-glutamylcysteine synthetase controls the rate limiting step in glutathione (GSH) biosynthesis. GSH, which is present in mM concentrations in cells, effectively chelates large amounts of zinc in vitro. Both MT and GSH also function as antioxidants. The current model suggests that the zinc-finger domain of MTF-1 directly (and reversibly) binds to zinc. This metalloregulatory protein then adopts a DNA-binding conformation and translocates to the nucleus, where it binds to metal-response elements in these gene promoters leading to increased transcription. The six zinc-finger domain of this factor is highly conserved from insects to mammals, and biochemical studies confirm that the zinc-fingers are heterogeneous in function and in zinc-binding. Furthermore, the mouse MTF-1 gene is essential for development of the embryo, thus underscoring the importance of this transcription factor. PMID:11831458

  4. Berberine inhibits Wilms' tumor cell progression through upregulation of Wilms' tumor gene on the X chromosome.

    PubMed

    Liu, Yan; Liu, Sheng

    2013-11-01

    Wilms' tumor is a type of kidney cancer that affects young children. Although a number of Wilms' tumor samples have been collected through international trials, the mechanisms underlying its progression remain challenging to determine. Extensive studies have identified somatic mutations at several loci in Wilms' tumorigenesis, including WT1, catenin, Wilms' tumor gene on the X chromosome (WTX) and TP53. Berberine is a benzylisoquinoline alkaloid extracted from numerous types of medicinal plants and has been extensively used as a Chinese traditional medicine. Recently, berberine has been demonstrated to possess antitumoral activities. AMP-activated protein kinase (AMPK) is suggested to be one of the various cellular targets of berberine, which regulates tumor progression and metastasis. However, the specific involvement of berberine?induced AMPK activation and its effects on the proliferation potential of Wilms' tumor cells remains unknown. The present study investigated the berberine?induced activation of AMPK and its effects on G401 Wilms' tumor cell proliferation. The results demonstrated that berberine inhibited growth and decreased the expression of cell?cycle regulators in these cells. At the molecular level, berberine treatment led to a significant increase of WTX expression and G401 cells were protected against berberine?induced growth inhibition by small interfering RNA against WTX. In conclusion, these results suggest a novel mechanism that may contribute to the antineoplastic effects of berberine which was also demonstrated by recent population studies; however, further studies are required to investigate the potential therapeutic use of berberine in patients with Wilms' tumor. PMID:24002362

  5. Rapid acclimation of juvenile corals to CO2 -mediated acidification by upregulation of heat shock protein and Bcl-2 genes.

    PubMed

    Moya, A; Huisman, L; Forêt, S; Gattuso, J-P; Hayward, D C; Ball, E E; Miller, D J

    2015-01-01

    Corals play a key role in ocean ecosystems and carbonate balance, but their molecular response to ocean acidification remains unclear. The only previous whole-transcriptome study (Moya et al. Molecular Ecology, 2012; 21, 2440) documented extensive disruption of gene expression, particularly of genes encoding skeletal organic matrix proteins, in juvenile corals (Acropora millepora) after short-term (3 d) exposure to elevated pCO2 . In this study, whole-transcriptome analysis was used to compare the effects of such 'acute' (3 d) exposure to elevated pCO2 with a longer ('prolonged'; 9 d) period of exposure beginning immediately post-fertilization. Far fewer genes were differentially expressed under the 9-d treatment, and although the transcriptome data implied wholesale disruption of metabolism and calcification genes in the acute treatment experiment, expression of most genes was at control levels after prolonged treatment. There was little overlap between the genes responding to the acute and prolonged treatments, but heat shock proteins (HSPs) and heat shock factors (HSFs) were over-represented amongst the genes responding to both treatments. Amongst these was an HSP70 gene previously shown to be involved in acclimation to thermal stress in a field population of another acroporid coral. The most obvious feature of the molecular response in the 9-d treatment experiment was the upregulation of five distinct Bcl-2 family members, the majority predicted to be anti-apoptotic. This suggests that an important component of the longer term response to elevated CO2 is suppression of apoptosis. It therefore appears that juvenile A. millepora have the capacity to rapidly acclimate to elevated pCO2 , a process mediated by upregulation of specific HSPs and a suite of Bcl-2 family members. PMID:25444080

  6. Upregulation of two actin genes and redistribution of actin during diapause and cold stress in the northern house mosquito, Culex pipiens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Two actin genes cloned from Culex pipiens L. are upregulated during adult diapause. Though actins 1 and 2 were expressed throughout diapause, both genes were most highly expressed early in diapause. These changes in gene expression were accompanied by a conspicuous redistribution of polymerized acti...

  7. Parallel up-regulation of the profilin gene family following independent domestication of diploid and allopolyploid cotton (Gossypium)

    PubMed Central

    Bao, Ying; Hu, Guanjing; Flagel, Lex E.; Salmon, Armel; Bezanilla, Magdalena; Paterson, Andrew H.; Wang, Zining; Wendel, Jonathan F.

    2011-01-01

    Cotton is remarkable among our major crops in that four species were independently domesticated, two allopolyploids and two diploids. In each case thousands of years of human selection transformed sparsely flowering, perennial shrubs into highly productive crops with seeds bearing the vastly elongated and abundant single-celled hairs that comprise modern cotton fiber. The genetic underpinnings of these transformations are largely unknown, but comparative gene expression profiling experiments have demonstrated up-regulation of profilin accompanying domestication in all three species for which wild forms are known. Profilins are actin monomer binding proteins that are important in cytoskeletal dynamics and in cotton fiber elongation. We show that Gossypium diploids contain six profilin genes (GPRF1–GPRF6), located on four different chromosomes (eight chromosomes in the allopolyploid). All but one profilin (GPRF6) are expressed during cotton fiber development, and both homeologs of GPRF1–GPRF5 are expressed in fibers of the allopolyploids. Remarkably, quantitative RT-PCR and RNAseq data demonstrate that GPRF1–GPRF5 are all up-regulated, in parallel, in the three independently domesticated cottons in comparison with their wild counterparts. This result was additionally supported by iTRAQ proteomic data. In the allopolyploids, there This usage of novel should be fine, since it refers to a novel evolutionary process, not a novel discovery has been novel recruitment of the sixth profilin gene (GPRF6) as a result of domestication. This parallel up-regulation of an entire gene family in multiple species in response to strong directional selection is without precedent and suggests unwitting selection on one or more upstream transcription factors or other proteins that coordinately exercise control over profilin expression. PMID:22160709

  8. 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 hours exposure to microgravity. We did, however, find significant changes in osteoblast gene expression of IEGs, c-fos and cox-2 in microgravity exposure as compared to ground and in-flight 1-G controls. Subsequent ground studies suggest that the molecular mechanism underlying these changes may involve prostaglandin c-AMP receptors (EPs) and/or subsequent alteration of intracellular signaling in the absence of gravity.

  9. 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

  10. Upregulation of ERG Genes in Candida Species by Azoles and Other Sterol Biosynthesis Inhibitors

    Microsoft Academic Search

    KARL W. HENRY; JOSEPH T. NICKELS; THOMAS D. EDLIND

    2000-01-01

    Infections due to Candida albicans are usually treated with azole antifungals such as fluconazole, but treat- ment failure is not uncommon especially in immunocompromised individuals. Relatedly, in vitro studies dem- onstrate that azoles are nonfungicidal, with continued growth at strain-dependent rates even at high azole con- centrations. We hypothesized that upregulation of ERG11, which encodes the azole target enzyme lanosterol

  11. Structural Protein Interactions Predict Kinase-Inhibitor Interactions in Upregulated Pancreas Tumour Genes Expression Data

    Microsoft Academic Search

    Gihan Dawelbait; Christian Pilarsky; Yanju Zhang; Robert Grützmann; Michael Schroeder

    2005-01-01

    Micro-arrays can identify co-expressed genes at large scale. The gene expression analysis does however not show functional relation- ships between co-expressed genes. To address this problem, we link gene expression data to protein interaction data. For the gene products of co- expressed genes, we identify structural domains by sequence alignment and threading. Next, we use the protein structure interaction PSIMAP

  12. Detecting cellular reprogramming determinants by differential stability analysis of gene regulatory networks

    PubMed Central

    2013-01-01

    Background Cellular differentiation and reprogramming are processes that are carefully orchestrated by the activation and repression of specific sets of genes. An increasing amount of experimental results show that despite the large number of genes participating in transcriptional programs of cellular phenotypes, only few key genes, which are coined here as reprogramming determinants, are required to be directly perturbed in order to induce cellular reprogramming. However, identification of reprogramming determinants still remains a combinatorial problem, and the state-of-art methods addressing this issue rests on exhaustive experimentation or prior knowledge to narrow down the list of candidates. Results Here we present a computational method, without any preliminary selection of candidate genes, to identify reduced subsets of genes, which when perturbed can induce transitions between cellular phenotypes. The method relies on the expression profiles of two stable cellular phenotypes along with a topological analysis stability elements in the gene regulatory network that are necessary to cause this multi-stability. Since stable cellular phenotypes can be considered as attractors of gene regulatory networks, cell fate and cellular reprogramming involves transition between these attractors, and therefore current method searches for combinations of genes that are able to destabilize a specific initial attractor and stabilize the final one in response to the appropriate perturbations. Conclusions The method presented here represents a useful framework to assist researchers in the field of cellular reprogramming to design experimental strategies with potential applications in the regenerative medicine and disease modelling. PMID:24350678

  13. Heteroconium chaetospira Induces Resistance to Clubroot via Upregulation of Host Genes Involved in Jasmonic Acid, Ethylene, and Auxin Biosynthesis

    PubMed Central

    Lahlali, Rachid; McGregor, Linda; Song, Tao; Gossen, Bruce D.; Narisawa, Kazuhiko; Peng, Gary

    2014-01-01

    An endophytic fungus, Heteroconium chaetospira isolate BC2HB1 (Hc), suppressed clubroot (Plasmodiophora brassicae -Pb) on canola in growth-cabinet trials. Confocal microscopy demonstrated that Hc penetrated canola roots and colonized cortical tissues. Based on qPCR analysis, the amount of Hc DNA found in canola roots at 14 days after treatment was negatively correlated (r?=?0.92, P<0.001) with the severity of clubroot at 5 weeks after treatment at a low (2×105 spores pot?1) but not high (2×105 spores pot?1) dose of pathogen inoculum. Transcript levels of nine B. napus (Bn) genes in roots treated with Hc plus Pb, Pb alone and a nontreated control were analyzed using qPCR supplemented with biochemical analysis for the activity of phenylalanine ammonia lyases (PAL). These genes encode enzymes involved in several biosynthetic pathways related potentially to plant defence. Hc plus Pb increased the activity of PAL but not that of the other two genes (BnCCR and BnOPCL) involved also in phenylpropanoid biosynthesis, relative to Pb inoculation alone. In contrast, expression of several genes involved in the jasmonic acid (BnOPR2), ethylene (BnACO), auxin (BnAAO1), and PR-2 protein (BnPR-2) biosynthesis were upregulated by 63, 48, 3, and 3 fold, respectively, by Hc plus Pb over Pb alone. This indicates that these genes may be involved in inducing resistance in canola by Hc against clubroot. The upregulation of BnAAO1 appears to be related to both pathogenesis of clubroot and induced defence mechanisms in canola roots. This is the first report on regulation of specific host genes involved in induced plant resistance by a non-mycorrhizal endophyte. PMID:24714177

  14. Identification of Dmrt genes and their up-regulation during gonad transformation in the swamp eel (Monopterus albus).

    PubMed

    Sheng, Yue; Chen, Bo; Zhang, Liao; Luo, Majing; Cheng, Hanhua; Zhou, Rongjia

    2014-03-01

    The swamp eel is a teleost fish with a characteristic of natural sex reversal and an ideal model for vertebrate sexual development. However, underlying molecular mechanisms are poorly understood. We report the identification of five DM (doublesex and mab-3) domain genes in the swamp eel that include Dmrt2, Dmrt2b, Dmrt3, Dmrt4 and Dmrt5, which encode putative proteins of 527, 373, 471, 420 and 448 amino acids, respectively. Phylogenetic tree showed that these genes are clustered into corresponding branches of the DM genes in vertebrates. Southern blot analysis indicated that the Dmrt1-Dmrt3-Dmrt2 genes are tightly linked in a conserved gene cluster. Notably, these Dmrt genes are up-regulated during gonad transformation. Furthermore, mRNA in situ hybridisation showed that Dmrt2, Dmrt3, Dmrt4 and Dmrt5 are expressed in developing germ cells. These results are evidence that the DM genes are involved in sexual differentiation in the swamp eel. PMID:24390316

  15. Upregulation of Androgen-Responsive Genes and Transforming Growth Factor-?1 Cascade Genes in a Rat Model of Non-bacterial Prostatic Inflammation

    PubMed Central

    Funahashi, Yasuhito; O’Malley, Katherine J.; Kawamorita, Naoki; Tyagi, Pradeep; DeFranco, Donald B.; Takahashi, Ryosuke; Gotoh, Momokazu; Wang, Zhou; Yoshimura, Naoki

    2013-01-01

    Background Prostatic inflammation is associated with the development of prostatic hyperplasia. We investigated the effects of prostatic inflammation on expression levels of androgen-responsive genes and growth factors in the prostate. Methods Prostatic inflammation was induced by formalin injection into bilateral ventral lobes of the prostate of male SD rats. After 28 days, the prostate was harvested for analyses of proinflammatory cytokines, androgen-responsive genes in the epithelium and TGF-?1 cascade genes in the stroma. Some rats were given a COX-2 inhibitor (celecoxib; 10 mg/kg/day) by oral gavage for 28 days. Results The formalin-injected prostate exhibited widespread low-grade inflammation (< 50 leukocytes/10,000 ?m2) along with focal high-grade inflammation (> 100 leukocytes/10,000 ?m2) in limited areas. Compared to control, formalin-injected prostate exhibited a 2.5–6 fold increased protein expression of IL-1?, IL-1?, and IL-6. In the low-grade inflammatory regions, 3–9 fold and 2–3 fold upregulations of mRNA levels of androgen receptors/androgen-responsive genes and TGF-?1 cascade genes were respectively observed in the epithelium and stroma obtained by laser-capture microdissection. Positive staining for androgen receptors in the epithelial nuclei, and TGF-?1, IL-6 and COX-2 in the stroma was increased in the low-grade inflammation area. COX-2 inhibitor treatment suppressed these upregulations of cytokines, androgen-responsive and TGF-?1 cascade genes. Conclusions Prostatic inflammation induced increased expression of androgen-responsive genes in the epithelium and TGF-?1 cascade genes in the stroma, which were suppressed by COX-2 inhibitors, suggesting that activation of these genes in the low-grade inflammatory region might be involved in the development of symptomatic BPH. PMID:24446128

  16. Manganese Upregulates Cellular Prion Protein and Contributes to Altered Stabilization and Proteolysis: Relevance to Role of Metals in Pathogenesis of Prion Disease

    PubMed Central

    Choi, Christopher J.; Anantharam, Vellareddy; Martin, Dustin P.; Nicholson, Eric M.; Richt, Jürgen A.; Kanthasamy, Arthi; Kanthasamy, Anumantha G.

    2010-01-01

    Prion diseases are fatal neurodegenerative diseases resulting from misfolding of normal cellular prion (PrPC) into an abnormal form of scrapie prion (PrPSc). The cellular mechanisms underlying the misfolding of PrPC are not well understood. Since cellular prion proteins harbor divalent metal-binding sites in the N-terminal region, we examined the effect of manganese on PrPC processing in in vitro models of prion disease. Exposure to manganese significantly increased PrPC levels both in cytosolic and in membrane-rich fractions in a time-dependent manner. Manganese-induced PrPC upregulation was independent of messenger RNA transcription or stability. Additionally, manganese treatment did not alter the PrPC degradation by either proteasomal or lysosomal pathways. Interestingly, pulse-chase analysis showed that the PrPC turnover rate was significantly altered with manganese treatment, indicating increased stability of PrPC with the metal exposure. Limited proteolysis studies with proteinase-K further supported that manganese increases the stability of PrPC. Incubation of mouse brain slice cultures with manganese also resulted in increased prion protein levels and higher intracellular manganese accumulation. Furthermore, exposure of manganese to an infectious prion cell model, mouse Rocky Mountain Laboratory–infected CAD5 cells, significantly increased prion protein levels. Collectively, our results demonstrate for the first time that divalent metal manganese can alter the stability of prion proteins and suggest that manganese-induced stabilization of prion protein may play a role in prion protein misfolding and prion disease pathogenesis. PMID:20176619

  17. Genes Selectively Up-Regulated by Pheromone in White Cells Are Involved in Biofilm Formation in Candida albicans

    PubMed Central

    Sahni, Nidhi; Yi, Song; Daniels, Karla J.; Srikantha, Thyagarajan; Pujol, Claude; Soll, David R.

    2009-01-01

    To mate, MTL-homozygous strains of the yeast pathogen Candida albicans must switch from the white to opaque phase. Mating-competent opaque cells then release pheromone that induces polarization, a G1 block and conjugation tube formation in opaque cells of opposite mating type. Pheromone also induces mating-incompetent white cells to become adhesive and cohesive, and form thicker biofilms that facilitate mating. The pheromone response pathway of white cells shares the upstream components of that of opaque cells, but targets a different transcription factor. Here we demonstrate that the genes up-regulated by the pheromone in white cells are activated through a common cis-acting sequence, WPRE, which is distinct from the cis-acting sequence, OPRE, responsible for up-regulation in opaque cells. Furthermore, we find that these white-specific genes play roles in white cell biofilm formation, and are essential for biofilm formation in the absence of an added source of pheromone, suggesting either an autocrine or pheromone-independent mechanism. These results suggest an intimate, complex and unique relationship between switching, mating and MTL-homozygous white cell biofilm formation, the latter a presumed virulence factor in C. albicans. PMID:19798425

  18. Gamma tocotrienol, a potent radioprotector, preferentially upregulates expression of anti-apoptotic genes to promote intestinal cell survival.

    PubMed

    Suman, Shubhankar; Datta, Kamal; Chakraborty, Kushal; Kulkarni, Shilpa S; Doiron, Kathryn; Fornace, Albert J; Sree Kumar, K; Hauer-Jensen, Martin; Ghosh, Sanchita P

    2013-10-01

    Gamma tocotrienol (GT3) has been reported as a potent ameliorator of radiation-induced gastrointestinal (GI) toxicity when administered prophylactically. This study aimed to evaluate the role of GT3 mediated pro- and anti-apoptotic gene regulation in protecting mice from radiation-induced GI damage. Male 10- to 12-weeks-old CD2F1 mice were administered with a single dose of 200 mg/kg of GT3 or equal volume of vehicle (5% Tween-80) 24 h before exposure to 11 Gy of whole-body ?-radiation. Mouse jejunum was surgically removed 4 and 24h after radiation exposure, and was used for PCR array, histology, immunohistochemistry, and immunoblot analysis. Results were compared among vehicle pre-treated no radiation, vehicle pre-treated irradiated, and GT3 pre-treated irradiated groups. GT3 pretreated irradiated groups, both 4h and 24h after radiation, showed greater upregulation of anti-apoptotic gene expression than vehicle pretreated irradiated groups. TUNEL staining and intestinal crypt analysis showed protection of jejunum after GT3 pre-treatment and immunoblot results were supportive of PCR data. Our study demonstrated that GT3-mediated protection of intestinal cells from a GI-toxic dose of radiation occurred via upregulation of antiapoptotic and downregulation of pro-apoptotic factors, both at the transcript as well as at the protein levels. PMID:23941772

  19. Upregulation of Plasmid Genes during Stationary Phase in Synechocystis sp. Strain PCC 6803, a Cyanobacterium

    PubMed Central

    Berla, Bertram M.

    2012-01-01

    We analyzed DNA microarrays to identify highly expressed genes during stationary-phase growth of Synechocystis sp. PCC 6803. Many identified genes are on endogenous plasmids, with copy numbers between 0.4 and 7 per chromosome. The promoters of such genes will be useful for synthetic biology applications with this phototrophic host. PMID:22636001

  20. A Novel Gene Amplification Causes Upregulation of the PatAB ABC Transporter and Fluoroquinolone Resistance in Streptococcus pneumoniae

    PubMed Central

    Baylay, Alison J.; Ivens, Alasdair

    2015-01-01

    Overexpression of the ABC transporter genes patA and patB confers efflux-mediated fluoroquinolone resistance in Streptococcus pneumoniae and is also linked to pneumococcal stress responses. Although upregulation of patAB has been observed in many laboratory mutants and clinical isolates, the regulatory mechanisms controlling expression of these genes are unknown. In this study, we aimed to identify the cause of high-level constitutive overexpression of patAB in M184, a multidrug-resistant mutant of S. pneumoniae R6. Using a whole-genome transformation and sequencing approach, we identified a novel duplication of a 9.2-kb region of the M184 genome which included the patAB genes. This duplication did not affect growth and was semistable with a low segregation rate. The expression levels of patAB in M184 were much higher than those that could be fully explained by doubling of the gene dosage alone, and inactivation of the first copy of patA had no effect on multidrug resistance. Using a green fluorescent protein reporter system, increased patAB expression was ascribed to transcriptional read-through from a tRNA gene upstream of the second copy of patAB. This is the first report of a large genomic duplication causing antibiotic resistance in S. pneumoniae and also of a genomic duplication causing antibiotic resistance by a promoter switching mechanism. PMID:25779578

  1. Association between up-regulation of stress-responsive genes and hypomethylation of genomic DNA in tobacco plants

    Microsoft Academic Search

    Y. Wada; K. Miyamoto; T. Kusano; H. Sano

    2004-01-01

    Transcripts that specifically accumulate in transgenic tobacco plants expressing an anti-sense construct for a tobacco type I DNA methyltransferase, NtMET1, were screened by the differential display method. Of the 31 genes identified, 16 encoded proteins with known functions; ten of these were related to biotic and abiotic stress responses, and the other six to cellular functions. In order to examine

  2. Computational evaluation of cellular metabolic costs successfully predicts genes whose expression

    E-print Network

    Ruppin, Eytan

    Computational evaluation of cellular metabolic costs successfully predicts genes whose expression resulting from overexpression of either native or foreign metabolic genes. We first test and validate EDGE the expression of potentially deleterious genes in check. Third, EDGE- based analysis shows that cancer genetic

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

    PubMed

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

    2015-01-01

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

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

    PubMed Central

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

    2015-01-01

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

  5. Lipopolysaccharide pretreatment inhibits LPS-induced human umbilical cord mesenchymal stem cell apoptosis via upregulating the expression of cellular FLICE-inhibitory protein.

    PubMed

    Hou, Yu Sen; Liu, Ling Ying; Chai, Jia Ke; Yu, Yong Hui; Duan, Hong Jie; Hu, Quan; Yin, Hui Nan; Wang, Yi He; Zhuang, Shu Bo; Fan, Jun; Chu, Wan Li; Ma, Li

    2015-08-01

    Mesenchymal stem cell (MSC)-based regenerative therapy is currently regarded as a novel approach with which to repair damaged tissues. However, the efficiency of MSC transplantation is limited due to the low survival rate of engrafted MSCs. Lipopolysaccharide (LPS) production is increased in numerous diseases and serves an essential function in the regulation of apoptosis in a variety of cell types. Previous studies have indicated that low?dose LPS pretreatment contributes to cytoprotection. In the current study, LPS was demonstrated to induce apoptosis in human umbilical cord mesenchymal stem cells (hUCMSCs) via the activation of caspase, in a dose?dependent manner. Low?dose LPS pretreatment may protect hUCMSCs against apoptosis induced by high?dose LPS, by upregulating the expression of cellular FADD?like IL?1??converting enzyme?inhibitory protein (c?FLIP). The results of the present study indicate that pretreatment with an appropriate concentration of LPS may alleviate high-dose LPS-induced apoptosis. PMID:25955291

  6. The Yeast Homolog of Heme Oxygenase-1 Affords Cellular Antioxidant Protection via the Transcriptional Regulation of Known Antioxidant Genes*

    PubMed Central

    Collinson, Emma J.; Wimmer-Kleikamp, Sabine; Gerega, Sebastien K.; Yang, Yee Hwa; Parish, Christopher R.; Dawes, Ian W.; Stocker, Roland

    2011-01-01

    Heme oxygenase-1 (HO-1) degrades heme and protects cells from oxidative challenge. This antioxidant activity is thought to result from the HO-1 enzymatic activity, manifested by a decrease in the concentration of the pro-oxidant substrate heme, and an increase in the antioxidant product bilirubin. Using a global transcriptional approach, and yeast as a model, we show that HO-1 affords cellular protection via up-regulation of transcripts encoding enzymes involved in cellular antioxidant defense, rather than via its oxygenase activity. Like mammalian cells, yeast responds to oxidative stress by expressing its HO-1 homolog and, compared with the wild type, heme oxygenase-null mutant cells have increased sensitivity toward oxidants that is rescued by overexpression of human HO-1 or its yeast homolog. Increased oxidant sensitivity of heme oxygenase-null mutant cells is explained by a decrease in the expression of the genes encoding ?-glutamylcysteine synthetase, glutathione peroxidase, catalase, and methionine sulfoxide reductase, because overexpression of any of these genes affords partial, and overexpression of all four genes provides complete, protection to the null mutant. Genes encoding antioxidant enzymes represent only a small portion of the 480 differentially expressed transcripts in heme oxygenase-null mutants. Transcriptional regulation may be explained by the nuclear localization of heme oxygenase observed in oxidant-challenged cells. Our results challenge the notion that HO-1 functions simply as a catabolic and antioxidant enzyme. They indicate much broader functions for HO-1, the unraveling of which may help explain the multiple biological responses reported in animals as a result of altered HO-1 expression. PMID:21081499

  7. FUS1 acts as a tumor-suppressor gene by upregulating miR-197 in human glioblastoma.

    PubMed

    Xin, Jun; Zhang, Xue-Kui; Xin, De-You; Li, Xian-Feng; Sun, De-Ke; Ma, Yue-Ye; Tian, Li-Qiang

    2015-08-01

    Glioblastoma is the most common primary malignancy of the adult central nervous system (CNS) and is associated with an exceptionally poor prognosis. Elucidation of the pathogenesis and molecular changes will help us to further understand the pathogenesis and progression of the disease and offer new therapeutic targets. FUS1 (TUSC2, tumor suppressor candidate 2) is a tumor-suppressor gene located on human chromosome 3p21. Restoration of FUS1 function in human non-small cell lung cancer (NSCLC) cells was found to significantly inhibit tumor cell growth and modulate the chemosensitivity of lung cancer cells. Yet, its role in human glioblastoma has rarely been addressed. In the present study, we demonstrated that low expression of FUS1 was detected in high-grade human glioma, implying that FUS1 expression is negatively associated with progression of the disease. Subsequent studies confirmed that FUS1 overexpression inhibited the proliferation, migration and invasion of human glioblastoma cells. In addition, we found that FUS1 overexpression significantly upregulated miR-197 expression in the glioblastoma cells. We also revealed that miR-197 suppressed the proliferation, migration and invasion of the cells as well as the silencing of miR-197 attenuated the biological functions of FUS1. Using human glioblastoma tissue samples, we demonstrated that miR-197 is negatively associated with metastasis. All the results demonstrated that FUS1 acts as a tumor-suppressor gene by upregulating miR-197 in human glioblastoma and implied that restoration of FUS1 and miR-197 could be new therapeutic strategies for glioblastoma. PMID:26081814

  8. Swine PPAR-?2 expression upregulated in skeletal muscle of transgenic mice via the swine Myozenin-1 gene promoter.

    PubMed

    Ma, Juanjuan; Chai, Jin; Shang, Yangyang; Li, Yujiao; Chen, Ran; Jia, Jia; Jiang, Siwen; Peng, Jian

    2015-06-01

    Myozenin-1 (Myoz1) gene-encoded calsarcin-2 protein was expressed exclusively in fast-twitch muscles. Peroxisome proliferator-activated receptor ?2 (PPAR-?2) is a key regulator of adipocyte differentiation, fatty acid uptake and storage in mammals. In this study, transgenic (TG) mice were generated by injecting linearized DNA that contained mouse creatine kinase M-type enhancer, Myoz1 core promoter, swine PPAR-?2 (sPPAR-?2) and SV40 polyadenylation sequences into pronuclei of fertilized FVB/NJ mouse embryos using microinjection technology. Then, the TG mice were used to identify whether swine Myoz1 (sMyoz1) promoter could upregulate sPPAR-?2 expression in skeletal muscle in a TG mouse model. The results showed that the sMyoz1 promoter indeed upregulated sPPAR-?2 expression on both the RNA and protein levels. The target genes of PPAR-? in fat formation pathways, such as fatty acid-binding protein 4 (FABP4) and lipoprotein lipase (LPL), were also overexpressed on the RNA level. Meanwhile, the level of skeletal muscle triacylglycerol in TG mice was increased (P < 0.05), and the result of Oil Red-O staining in the skeletal muscle sections also showed that the number of lipid droplets was significantly increased in TG mice compared to wild-type mice, which might improve the intramuscular fat (IMF) content. For pork, the quality was mostly influenced by the IMF; the identification of swine muscle-specific promoter, sMyoz1, could further serve to develop transgenic pigs with higher intramuscular fat contents and improve pork quality. PMID:25421932

  9. Beta-lactam antibiotics modulate T-cell functions and gene expression via covalent binding to cellular albumin

    PubMed Central

    Mor, Felix; Cohen, Irun R.

    2013-01-01

    Recent work has suggested that beta-lactam antibiotics might directly affect eukaryotic cellular functions. Here, we studied the effects of commonly used beta-lactam antibiotics on rodent and human T cells in vitro and in vivo on T-cell–mediated experimental autoimmune diseases. We now report that experimental autoimmune encephalomyelitis and adjuvant arthritis were significantly more severe in rats treated with cefuroxime and other beta-lactams. T cells appeared to mediate the effect: an anti-myelin basic protein T-cell line treated with cefuroxime or penicillin was more encephalitogenic in adoptive transfer experiments. The beta-lactam ampicillin, in contrast to cefuroxime and penicillin, did not enhance encephalomyelitis, but did inhibit the autoimmune diabetes developing spontaneously in nonobese diabetic mice. Gene expression analysis of human peripheral blood T cells showed that numerous genes associated with T helper 2 (Th2) and T regulatory (Treg) differentiation were down-regulated in T cells stimulated in the presence of cefuroxime; these genes were up-regulated in the presence of ampicillin. The T-cell protein that covalently bound beta-lactam antibiotics was found to be albumin. Human and rodent T cells expressed albumin mRNA and protein, and penicillin-modified albumin was taken up by rat T cells, leading to enhanced encephalitogenicity. Thus, beta-lactam antibiotics in wide clinical use have marked effects on T-cell behavior; beta-lactam antibiotics can function as immunomodulators, apparently through covalent binding to albumin. PMID:23382225

  10. Upregulation of URI/RMP gene expression in cervical cancer by high-throughput tissue microarray analysis

    PubMed Central

    Gu, Junxia; Li, Xiaoyun; Liang, Yuting; Qiao, Longwei; Ran, Deyuan; Lu, Yaojuan; Li, Xingang; Wei, Wenxiang; Zheng, Qiping

    2013-01-01

    URI, or RMP, is a RNA polymerase II subunit RPB5-associated protein known to play essential roles in ubiquitination and transcription. Recently, we and others have shown that URI/RMP is also important for progression of hepatocellular carcinoma, ovarian, and prostate cancers. To identify the mechanistic basis of URI/RMP during multiple cellular processes, we investigated URI/RMP expression in a tissue microarray (TMA) containing multiple normal human tissues. The results showed that URI/RMP is ubiquitously but differentially expressed in these human tissues which partially explains its multiple cellular functions. To elucidate the role of URI/RMP during oncogenesis of multiple malignancies, especially the tumors of reproductive system, we analyzed URI/RMP expression in a TMA containing multiple reproductive system tumors. We did not observe significant difference of URI/RMP expression between cancerous and adjacent tissues of the prostate, breast, ovarian, and endometrial cancers. However, increased URI/RMP expression was observed in two of the three cases of cervical SCC (squamous cell carcinoma) cells compared to their adjacent epithelial cells. Moreover, we detected significantly upregulated URI/RMP expression not only in cervical cancers but also in pre-cancerous CINs (cervical intra-epithelial neoplasias) in a TMA that covers the whole spectrum of normal cervix, CINs, and cervical cancers. No difference of URI/RMP expression was observed between CINs and cervical cancers. Given the high risk of CINs (especially CIN3) turning into cervical cancer if left untreated, the increased URI/RMP expression in CINs as well as in cervical cancers suggest a clinical relevance of URI/RMP upon cervical cancer tumorigenesis and worth further investigation. PMID:23573313

  11. Identification of Upregulated Genes under Cold Stress in Cold-Tolerant Chickpea Using the cDNA-AFLP Approach

    PubMed Central

    Dinari, Ali; Niazi, Ali; Afsharifar, Ali Reza; Ramezani, Amin

    2013-01-01

    Low temperature injury is one of the most significant causes of crop damage worldwide. Cold acclimatization processes improve the freezing tolerance of plants. To identify genes of potential importance for acclimatzation to the cold and to elucidate the pathways that regulate this process, global transcriptome expression of the chickpea (Cicer arietinum L), a species of legume, was analyzed using the cDNA-AFLP technique. In total, we generated 4800 transcript-derived fragments (TDFs) using cDNA-AFLP in conjunction with 256 primer combinations. We only considered those cDNA fragments that seemed to be up-regulated during cold acclimatization. Of these, 102 TDFs with differential expression patterns were excised from gels and re-amplified by PCR. Fifty-four fragments were then cloned and sequenced. BLAST search of the GenBank non-redundant (nr) sequence database demonstrated that 77 percent of the TDFs belonged to known sequences with putative functions related to metabolism (31), transport (10), signal transduction pathways (15) and transcription factors (21). The last group of expressed transcripts showed homology to genes of unknown function (22). To further analyze and validate our cDNA-AFLP experiments, the expression of 9 TDFs during cold acclimatzatiion was confirmed using real time RT-PCR. The results of this research show that cDNA-AFLP is a powerful technique for investigating the expression pattern of chickpea genes under low-temperature stress. Moreover, our findings will help both to elucidate the molecular basis of low-temperature effects on the chickpea genome and to identify those genes that could increase the cold tolerance of the chickpea plant. PMID:23341906

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

    PubMed Central

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

    2010-01-01

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

  13. Inducible nitric oxide synthase expression in chronic viral hepatitis. Evidence for a virus-induced gene upregulation.

    PubMed Central

    Majano, P L; García-Monzón, C; López-Cabrera, M; Lara-Pezzi, E; Fernández-Ruiz, E; García-Iglesias, C; Borque, M J; Moreno-Otero, R

    1998-01-01

    Increased nitric oxide (NO) production may contribute to the pathological changes featuring in some inflammatory diseases, but the role of NO in chronic viral hepatitis is still unknown. We compared the inducible NO synthase (NOS2) expression in the liver of patients with chronic viral hepatitis with that of both nonviral liver disease and histologically normal liver. NOS2 expression was assessed by immunohistochemical and in situ hybridization studies of liver biopsy sections. An intense hepatocellular NOS2 reactivity was detected in chronic viral hepatitis, whereas it was weakly or not observed in nonviral liver disease or normal liver, respectively. In addition, we determined whether the hepatitis B virus (HBV) might regulate the synthesis of this enzyme. NOS2 mRNA and protein levels as well as enzyme activity were assessed in cytokine-stimulated HBV-transfected and untransfected hepatoma cells. Transfection with either HBV genome or HBV X gene resulted in induction of NOS2 mRNA expression, and the maximal induction of this transcript and NO production was observed in cytokine-stimulated HBV-transfected cells. These results indicate that hepatotropic viral infections are able to upregulate the NOS2 gene expression in human hepatocytes, suggesting that NO may mediate important pathogenic events in the course of chronic viral hepatitis. PMID:9525976

  14. Upregulation of Atrial Natriuretic Peptide Gene Expression in Remnant Kidney of Rats with Reduced Renal Mass

    Microsoft Academic Search

    KAZUHITO TOTSUNE; HARALD S. MACKENZIE; HIROKO TOTSUNE; JULIA L. TROY; JONATHAN LYTTON; BARRY M. BRENNER

    1998-01-01

    Atrial natriuretic peptide (ANP) is synthesized in the kidney but its physiologic significance there is unclear. To determine whether renal expression of the ANP gene is regu- lated, renal ANP mRNA expression was assessed in remnant kidneys after 5\\/6 nephrectomy in Munich-Wistar rats. In nor- mal sodium intake groups, ANP mRNA expression in the remnant kidney was significantly increased by

  15. Novel genes upregulated when NOTCH signalling is disrupted during hypothalamic development

    PubMed Central

    2013-01-01

    Background The generation of diverse neuronal types and subtypes from multipotent progenitors during development is crucial for assembling functional neural circuits in the adult central nervous system. It is well known that the Notch signalling pathway through the inhibition of proneural genes is a key regulator of neurogenesis in the vertebrate central nervous system. However, the role of Notch during hypothalamus formation along with its downstream effectors remains poorly defined. Results Here, we have transiently blocked Notch activity in chick embryos and used global gene expression analysis to provide evidence that Notch signalling modulates the generation of neurons in the early developing hypothalamus by lateral inhibition. Most importantly, we have taken advantage of this model to identify novel targets of Notch signalling, such as Tagln3 and Chga, which were expressed in hypothalamic neuronal nuclei. Conclusions These data give essential advances into the early generation of neurons in the hypothalamus. We demonstrate that inhibition of Notch signalling during early development of the hypothalamus enhances expression of several new markers. These genes must be considered as important new targets of the Notch/proneural network. PMID:24360028

  16. Target Genes of Neuron-Restrictive Silencer Factor Are Abnormally Up-Regulated in Human Myotilinopathy

    PubMed Central

    Barrachina, Marta; Moreno, Jesús; Juvés, Salvador; Moreno, Dolores; Olivé, Montse; Ferrer, Isidre

    2007-01-01

    Myotilinopathy is a subgroup of myofibrillar myopathies caused by mutations in the myotilin gene in which there is aggregation of abnormal cytoskeletal proteins and ubiquitin. We report here on the accumulation of neuron-related proteins such as ubiquitin carboxy-terminal hydrolase L1 (UCHL1), synaptosomal-associated protein 25, synaptophysin, and ?-internexin in aberrant protein aggregates in myotilinopathy. We have determined that the neuron-restrictive silencer factor (NRSF)/RE1 silencing transcription factor (REST), a transcription factor expressed in non-neuronal tissues repressing the expression of several neuronal genes, is reduced in myotilinopathies. Moreover, NRSF transfection reduces UCHL1, synaptosomal-associated protein 25, synaptophysin, and ?-internexin mRNA levels in DMS53 cells, whereas short interferring NRSF transfection increases UCHL1 and synaptophysin mRNA levels in U87-MG cells. Chromatin immunoprecipitation assays have shown that NRSF interacts with the UCHL1 promoter in U87-MG and HeLa cells. In silico analysis of the UCHL1 gene promoter sequence using the MatInspector software has predicted three potential neuron-restrictive silencer elements (NRSEs): NRSE1 located in the complementary DNA chain and NRSE2 and NRSE3 in intron 1, in the coding and complementary chains, respectively. Together, these findings show, for the first time, abnormal regulation of NRSF/REST as a mechanism associated with the aberrant expression of selected neuron-related proteins, which in turn accumulate in abnormal protein aggregates, in myotilinopathy. PMID:17823282

  17. Upregulated ex vivo expression of stress-responsive inflammatory pathway genes by LPS-challenged CD14+ monocytes in frail older adults

    PubMed Central

    Qu, Tao; Walston, Jeremy D.; Yang, Huanle; Fedarko, Neal S.; Xue, Qian-Li; Beamer, Brock A.; Ferrucci, Luigi; Rose, Noel R.; Leng, Sean X.

    2009-01-01

    Frailty has been increasingly recognized as an important clinical syndrome in old age. The frailty syndrome is characterized by chronic inflammation, decreased functional and physiologic reserve, and increased vulnerability to stressors, leading to disability and mortality. However, molecular mechanisms that contribute to inflammation activation and regulation in frail older adults have not been investigated. To begin to address this, we conducted a pathway-specific gene array analysis of 367 inflammatory pathway genes by lipopolysaccharide (LPS)-challenged CD14+ monocytes from 32 community-dwelling frail and age-, race-, and sex-paired nonfrail older adults (mean age 83 years, range 72–94). The results showed that ex vivo LPS-challenge induced average 2.0-fold or higher upregulated expression of 116 genes in frail participants and 85 genes in paired nonfrail controls. In addition, frail participants had 2-fold or higher upregulation in LPS-induced expression of 7 stress-responsive genes than nonfrail controls with validation by quantitative real time RT-PCR. These findings suggest upregulated expression of specific stress-responsive genes in monocyte-mediated inflammatory pathway in the syndrome of frailty with potential mechanistic and interventional implications. PMID:19027777

  18. COUP-TF Upregulates NGFI-A Gene Expression through an Sp1 Binding Site

    PubMed Central

    Pipaón, Carlos; Tsai, Sophia Y.; Tsai, Ming-Jer

    1999-01-01

    The formation of various tissues requires close communication between two groups of cells, epithelial and mesenchymal cells. COUP-TFs are transcription factors which have been shown to have functions in embryonic development. COUP-TFI is expressed mainly in the nervous system, and its targeted deletion leads to defects in the central and peripheral nervous systems. COUP-TFII is highly expressed in the mesenchymal component of the developing organs. A null mutation of COUP-TFII results in the malformation of the heart and blood vessels. From their expression pattern, we proposed that COUP-TFs regulate paracrine signals important for mesenchymal cell-epithelial cell interactions. In order to identify genes regulated by COUP-TF in this process, a rat urogenital mesenchymal cell line was stably transfected with a COUP-TFI expression vector. We found that NGFI-A, a gene with important functions in brain, organ, and vasculature development, has elevated mRNA and protein levels upon overexpression of COUP-TFI in these cells. A study of the promoter region of this gene identified a COUP-TF-responsive element between positions ?64 and ?46. Surprisingly, this region includes binding sites for members of the Sp1 family of transcription factors but no COUP-TF binding site. Mutations that abolish the Sp1 binding activity also impair the transactivation of the NGFI-A promoter by COUP-TF. Two regions of the COUP-TF molecule are shown to be important for NGFI-A activation: the DNA binding domain and the extreme C terminus of the putative ligand binding domain. The C-terminal region is likely to be important for interaction with coactivators. In fact, the coactivators p300 and steroid receptor activator 1 can enhance the transactivation of the NGFI-A promoter induced by COUP-TFI. Finally, we demonstrated that COUP-TF can directly interact with Sp1. Taken together, these results suggest that NGFI-A is a target gene for COUP-TFs and that the Sp1 family of transcription factors mediates its regulation by COUP-TFs. PMID:10082539

  19. Fibrinogen depletion attenuates Staphyloccocus aureus infection by preventing density-dependent virulence gene up-regulation.

    PubMed

    Rothfork, Jacob M; Dessus-Babus, Sophie; Van Wamel, Willem J B; Cheung, Ambrose L; Gresham, Hattie D

    2003-11-15

    Staphylococcus aureus undergoes a density-dependent conversion in phenotype from tissue-adhering to tissue-damaging and phagocyte-evading that is mediated in part by the quorum-sensing operon, agr, and its effector, RNAIII. Contributions of host factors to this mechanism for regulating virulence have not been studied. We hypothesized that fibrinogen, as a component of the inflammatory response, could create spatially constrained microenvironments around bacteria that increase density independently of bacterial numbers and thus potentiate quorum-sensing-dependent virulence gene expression. Here we show that transient fibrinogen depletion significantly reduces the bacterial burden and the consequential morbidity and mortality during experimental infection with wild-type S. aureus, but not with bacteria that lack expression of the quorum-sensing operon, agr. In addition, it inhibits in vivo activation of the promoter for the agr effector, RNAIII, and downstream targets of RNAIII, including alpha hemolysin and capsule production. Moreover, both in vitro and in vivo, the mechanism for promoting this phenotypic switch in virulence involves clumping of the bacteria, demonstrating that S. aureus responds to fibrinogen-mediated bacterial clumping by enhancing density-dependent virulence gene expression. These data demonstrate that down-modulation of specific inflammatory components of the host that augment bacterial quorum sensing can be a strategy for enhancing host defense against infection. PMID:14607942

  20. Review: Gene amplification--a cellular response to genotoxic stress

    SciTech Connect

    Lueke-Huhle, C. (Kernforschungszentrum Karlsruhe, Institut fuer Genetik und Toxikologie (West Germany))

    1989-10-01

    Recent years of cancer research have defined the role of key regulatory genes in oncogenesis. Oncogenes and suppressor genes are affected in the process of carcinogenesis either by mutations within the coding region, promoter mutations, or gene amplification. This review describes the authors studies on gene amplification in mammalian cells, with emphasis on the initiating events induced by carcinogenic chemicals and various types of radiation. The influence of genomic instability, cell dedifferentiation, and the malignant potential of a cell on their capacity to amplify genes is demonstrated by molecular biologic and cytogenetic studies on human and rodent cells. Cells that contain amplified DNA are at risk for chromosomal aberrations, sister chromatid exchanges, and rearrangements. Surviving cells show such cancer-prone genetic consequences. 52 references.

  1. MMTV accessory factor Naf affects cellular gene expression

    Microsoft Academic Search

    Christoph Metzner; Brian Salmons; Walter H. Gunzburg; Manfred Gemeiner; Ingrid Miller; Bernd Gesslbauer; Andreas Kungl; John A. Dangerfield

    2006-01-01

    Mouse mammary tumour virus (MMTV) encodes a viral superantigen (Sag) and a negative acting factor (Naf) which share parts of their coding sequence. Using 2-dimensional gel electrophoresis (2D-DIGE), we could show that at least 10 different cellular proteins were differentially expressed in Naf positive cells. Also, luciferase reporter expression was down-regulated in Naf expressing cells independent of the promoter used

  2. 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

  3. Up-regulation of transferrin receptor gene expression by granulocyte colony-stimulating factor in human myeloid leukemia cells.

    PubMed

    Morishita, Y; Kataoka, T; Towatari, M; Ito, T; Inoue, H; Ogura, M; Morishima, Y; Saito, H

    1990-12-15

    Granulocyte colony-stimulating factor (G-CSF) enhanced surface transferrin receptor (TfR) expression in two human myeloid leukemia cell lines, NKM-1 and NOMO-1, which possess G-CSF receptors. Radioligand-binding assay revealed that 10 ng/ml G-CSF significantly increased TfR to 186 +/- 20 and 276 +/- 38% of control for NKM-1 cells and NOMO-1 cells, respectively, in a 24-h culture. Scatchard analysis showed the increase of transferrin (Tf)-binding sites but no change in the receptor affinity. The enhanced TfR expression was not mediated either by the kinetic change of receptor cycling or by cellular iron content. Immunoprecipitation with anti-TfR antibody was used, and the increased biosynthesis of the receptor was demonstrated in G-CSF-stimulated cells. Northern blot analysis showed a 2- to 3-fold increase of TfR mRNA of NKM-1 cells cultured in medium containing Tf and G-CSF, whereas the mRNA declined without G-CSF. The effect of G-CSF on the TfR mRNA was observed within 2 h, which preceded the increase of surface TfR and the transition to the S phase of the cell cycle. G-CSF also potentiated TfR expression in freshly obtained myeloid leukemia cells. The present study shows up-regulation of TfR expression by G-CSF in myeloid leukemia cells and provides evidence that the regulation is mediated by controlling the steady-state level of the mRNA. PMID:1701357

  4. Beneficial effects of pioglitazone and metformin in murine model of polycystic ovaries via improvement of chemerin gene up-regulation

    PubMed Central

    2014-01-01

    Background Polycystic ovary syndrome (PCO) is recognized as the most common endocrinopathy in female. Chemerin is a novel adipocytokine that is expressed in ovary and upregulated in adipose tissue of obese, PCO patients. To date there is no report about the regulation of ovarian chemerin gene expression after PCO induction and treatment by insulin sensitizing drugs including pioglitazone and metformin. Thirty female rats were divided into six experimental groups with five rats in each group including control group, PCO group (i.m injection of 4 mg estradiol benzoate for 40 days), metformin treated (200 mg/kg/day for 21 days), pioglitazone treated (20 mg/kg/day, for 21 days), PCO?+?metformin and PCO?+?pioglitazone. PCO was detected by microscopic observation of vaginal smear and treatment by metformin and pioglitazone was initiated one week after that. Ovarian chemerin expression was analyzed by real time PCR and western blotting. Results Our results demonstrated that PCO induction resulted in elevation of chemerin mRNA and protein levels in ovary in concomitant with incidence of insulin resistance and increasing androgen and progesterone production. We observed that metformin and pioglitazone attenuated ovarian chemerin expression and improved insulin resistance and abnormal steroid production in PCO rats. Conclusion Based on data presented here we concluded that alteration of ovarian chemerin expression may has important role in PCO development and manipulation of chemerin expression or signaling by pioglitazone or metformin can be a novel therapeutic mechanism in the treatment of PCO patients by these drugs. PMID:24762064

  5. iNOS gene upregulation is associated with the early proliferative response of human lung fibroblasts to cytokine stimulation.

    PubMed

    Romanska, Hanna M; Polak, Julia M; Coleman, Robert A; James, Rowena S; Harmer, Daniel W; Allen, Jennifer C; Bishop, Anne E

    2002-07-01

    Increased release of oxidants has been implicated in the pathogenesis of pulmonary fibrosis. Previous work in the rat showed that formation of the early fibrotic lesion is associated with increased expression of inducible nitric oxide synthase (iNOS) in pulmonary fibroblasts. The aim of this study was to test the hypothesis that NO is involved in the activation of pulmonary fibroblasts. The effects of endogenous and exogenous NO on proliferation of human pulmonary fibroblasts were investigated by administration of cytomix or SNAP, respectively. At low concentrations, both treatments increased cell numbers, an effect attenuated by iNOS inhibitor or NO scavenger. Induction of iNOS was confirmed by measurement of nitrate/nitrite production and by immunodetection. Quantitative RT-PCR showed an increase in iNOS mRNA as early as 3 h after stimulation. These results support the hypothesis and show that upregulation of the iNOS gene is an early event in the proliferative response of human lung fibroblasts to inflammatory stimuli. PMID:12115884

  6. Thyroid Hormone Upregulates Hypothalamic kiss2 Gene in the Male Nile Tilapia, Oreochromis niloticus

    PubMed Central

    Ogawa, Satoshi; Ng, Kai We; Xue, Xiaoyu; Ramadasan, Priveena Nair; Sivalingam, Mageswary; Li, Shuisheng; Levavi-Sivan, Berta; Lin, Haoran; Liu, Xiaochun; Parhar, Ishwar S.

    2013-01-01

    Kisspeptin has recently been recognized as a critical regulator of reproductive function in vertebrates. During the sexual development, kisspeptin neurons receive sex steroids feedback to trigger gonadotropin-releasing hormone (GnRH) neurons. In teleosts, a positive correlation has been found between the thyroid status and the reproductive status. However, the role of thyroid hormone in the regulation of kisspeptin system remains unknown. We cloned and characterized a gene encoding kisspeptin (kiss2) in a cichlid fish, the Nile tilapia (Oreochromis niloticus). Expression of kiss2 mRNA in the brain was analyzed by in situ hybridization. The effect of thyroid hormone (triiodothyronine, T3) and hypothyroidism with methimazole (MMI) on kiss2 and the three GnRH types (gnrh1, gnrh2, and gnrh3) mRNA expression was analyzed by real-time PCR. Expression of thyroid hormone receptor mRNAs were analyzed in laser-captured kisspeptin and GnRH neurons by RT-PCR. The kiss2 mRNA expressing cells were seen in the nucleus of the lateral recess in the hypothalamus. Intraperitoneal administration of T3 (5??g/g body weight) to sexually mature male tilapia significantly increased kiss2 and gnrh1 mRNA levels at 24?h post injection (P?

  7. Activation of cellular genes by avian RNA tumor viruses.

    PubMed Central

    Groudine, M; Weintraub, H

    1980-01-01

    We demonstrated previously that chicken embryo fibroblasts accumulate approximately 100 copies of embryonic globin RNA after transformation by Rous sarcoma virus. Here we demonstrate that the globin gene in chicken embryo fibroblasts is activated by infection with two other oncogenic retroviruses, avian erythroblastosis virus and strain MC-29 of avian myeloblastosis virus, which contain transforming genes unrelated in nucleotide sequence content to each other or to the Rous sarcoma virus src gene. In addition, we have measured the genetic complexity of transformation by using established techniques for determining the number of different RNA sequences in specific populations of cells. Our results indicate that transformation of chicken embryo fibroblasts by Rous sarcoma virus results in the accumulation of RNA from approximately 1000 average-sized new transcription units. PMID:6254077

  8. Genes for tumor markers are clustered with cellular proto-oncogenes on human chromosomes.

    PubMed

    Hozier, J C; Mass, M J; Siegfried, J M

    1987-09-01

    We have analyzed the relative mapping positions of genes for polypeptides expressed abnormally in tumors (tumor markers) and cellular proto-oncogenes and find a remarkable degree of co-mapping of tumor marker genes with oncogenes in the human karyotype. We propose that aberrant expression of marker genes in tumors may be related to their proximity in the human genome to oncogenes expressed during the development of malignancy, and we suggest ways to test this hypothesis of concerted abnormal gene expression in mammalian tumor cells. PMID:3308073

  9. Epigenetic regulations in the IFN? signalling pathway: IFN?-mediated MHC class I upregulation on tumour cells is associated with DNA demethylation of antigen-presenting machinery genes

    PubMed Central

    Vlková, Veronika; Št?pánek, Ivan; Hrušková, Veronika; Šenigl, Filip; Mayerová, Veronika; Šrámek, Martin; Šímová, Jana; Bieblová, Jana; Indrová, Marie; Hejhal, Tomáš; Dérian, Nicolas; Klatzmann, David; Six, Adrien; Reiniš, Milan

    2014-01-01

    Downregulation of MHC class I expression on tumour cells, a common mechanism by which tumour cells can escape from specific immune responses, can be associated with coordinated silencing of antigen-presenting machinery genes. The expression of these genes can be restored by IFN?. In this study we documented association of DNA demethylation of selected antigen-presenting machinery genes located in the MHC genomic locus (TAP-1, TAP-2, LMP-2, LMP-7) upon IFN? treatment with MHC class I upregulation on tumour cells in several MHC class I-deficient murine tumour cell lines (TC-1/A9, TRAMP-C2, MK16 and MC15). Our data also documented higher methylation levels in these genes in TC-1/A9 cells, as compared to their parental MHC class I-positive TC-1 cells. IFN?-mediated DNA demethylation was relatively fast in comparison with demethylation induced by DNA methyltransferase inhibitor 5-azacytidine, and associated with increased histone H3 acetylation in the promoter regions of APM genes. Comparative transcriptome analysis in distinct MHC class I-deficient cell lines upon their treatment with either IFN? or epigenetic agents revealed that a set of genes, significantly enriched for the antigen presentation pathway, was regulated in the same manner. Our data demonstrate that IFN? acts as an epigenetic modifier when upregulating the expression of antigen-presenting machinery genes. PMID:25071011

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

    PubMed

    Muranaka, Tomoaki; Kubota, Saya; Oyama, Tokitaka

    2013-12-01

    Gene expression is a fundamental cellular process and expression dynamics are of great interest in life science. We succeeded in monitoring cellular gene expression in a duckweed plant, Lemna gibba, using bioluminescent reporters. Using particle bombardment, epidermal and mesophyll cells were transfected with the luciferase gene (luc+) under the control of a constitutive [Cauliflower mosaic virus 35S (CaMV35S)] and a rhythmic [Arabidopsis thaliana CIRCADIAN CLOCK ASSOCIATED 1 (AtCCA1)] promoter. Bioluminescence images were captured using an EM-CCD (electron multiply charged couple device) camera. Luminescent spots of the transfected cells in the plant body were quantitatively measured at the single-cell level. Luminescence intensities varied over a 1,000-fold range among CaMV35S::luc+-transfected cells in the same plant body and showed a log-normal-like frequency distribution. We monitored cellular gene expression under light-dark conditions by capturing bioluminescence images every hour. Luminescence traces of ?50 individual cells in a frond were successfully obtained in each monitoring procedure. Rhythmic and constitutive luminescence behaviors were observed in cells transfected with AtCCA1::luc+ and CaMV35S::luc+, respectively. Diurnal rhythms were observed in every AtCCA1::luc+-introduced cell with traceable luminescence, and slight differences were detected in their rhythmic waveforms. Thus the single-cell bioluminescence monitoring system was useful for the characterization of cellular gene expression in a plant body. PMID:24058151

  11. Cellular transformation by E1 genes of enteric adenoviruses.

    PubMed

    Cousin, C; Winter, N; Gomes, S A; D'Halluin, J C

    1991-03-01

    The ability of Ad40 and Ad41 E1A plus E1B genes to transform BRK cells was considerably lower than that of Ad5 and Ad12 corresponding genes. However, as for Ad5, the E1A genes of enteric adenoviruses could cooperate with an activated ras oncogene for full cell transformation and the Ad41 E1B could be complemented by E1A gene of Ad5 or Ad12 for cell transformation. Complementation studies suggested that the conserved region 1 of Ad41 E1A was responsible for this inefficient transformation. The Ad40- and Ad41-transformed cell lines exhibited a low level of major histocompatibility complex (MHC) class I antigens correlated to the low level of Ad12-transformed cells. Class I MHC antigen amounts expressed at the surface of the cells transformed by the weakly oncogenic Ad3 were between the high level of Ad5- and the low level of Ad12-transformed cells. PMID:1825253

  12. Manganese Upregulates Cellular Prion Protein and Contributes to Altered Stabilization and Proteolysis: Relevance to Role of Metals in Pathogenesis of Prion Disease

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Prion diseases are fatal neurodegenerative diseases resulting from misfolding of normal cellular prion (PrP**C) into an abnormal form of scrapie prion (PrP**Sc). The cellular mechanisms underlying the misfolding of PrP**C are not well understood. Since cellular prion proteins harbor divalent metal b...

  13. Significant up-regulation of a novel gene, CLCP1, in a highly metastatic lung cancer subline as well as in lung cancers in vivo

    Microsoft Academic Search

    Katsumi Koshikawa; Hirotaka Osada; Ken-ichi Kozaki; Hiroyuki Konishi; Akira Masuda; Yoshio Tatematsu; Tetsuya Mitsudomi; Akimasa Nakao; Takashi Takahashi

    2002-01-01

    Most lung cancer patients are unfortunately uncurable and die because of widespread metastases, thus indicating the importance of identification of molecules with a crucial role in this process. Our previous expression profiling analysis of a highly metastatic lung cancer cell line, NCI-H460-LNM35, and its parental low metastatic line, NCI-H460-N15, revealed significant up-regulation of both known and unknown genes in LNM35.

  14. The HD-Zip gene ATHB6 in Arabidopsis is expressed in developing leaves, roots and carpels and up-regulated by water deficit conditions

    Microsoft Academic Search

    Eva Söderman; Mattias Hjellström; Jan Fahleson; Peter Engström

    1999-01-01

    Homeodomain-leucine zipper (HD-Zip) proteins are transcription factors as yet found only in plants. We have characterized one HD-Zip gene, ATHB6, from Arabidopsis thaliana. ATHB6 was expressed constitutively in seedlings, but significantly up-regulated in seedlings subjected to water deficit, osmotic stress or exogenous treatment with abscisic acid (ABA), an induction being detectable within 30 min. The ATHB6 induction was impaired in

  15. B-cell Translocation Gene 2 (BTG2) Stimulates Cellular Antioxidant Defenses through the Antioxidant Transcription Factor NFE2L2 in Human Mammary Epithelial Cells*

    PubMed Central

    Karve, Tejaswita M.; Rosen, Eliot M.

    2012-01-01

    The B-cell translocation gene 2, BTG2, a member of the BTG/TOB (B-cell translocation gene/transducers of ErbB2) gene family, has been implicated in cell cycle regulation, normal development, and possibly tumor suppression. Previously, it was shown that BTG2 expression is lost or down-regulated in human breast cancers. We now report that BTG2 protects human mammary epithelial cells from oxidative stress due to hydrogen peroxide and other oxidants. BTG2 protection against oxidative stress is BRCA1-independent but requires the antioxidant transcription factor NFE2L2 and is associated with up-regulation of the expression of antioxidant enzymes, including catalase and superoxide dismutases 1 and 2. BTG2 stimulation of antioxidant gene expression is also NFE2L2-dependent. We further demonstrate that BTG2 is a binding partner for NFE2L2 and increases its transcriptional activity. In addition, BTG2 is detectable at the antioxidant response element (ARE) of several NFE2L2-responsive genes. Finally, we show that the ability of BTG2 to associate with NFE2L2, to protect cells against oxidative stress, and to stimulate antioxidant gene expression requires box B, a short highly conserved amino acid motif characteristic of BTG2/TOB family proteins, but does not require boxes A or C. These findings suggest a novel role for BTG2 as a co-activator for NFE2L2 in up-regulating cellular antioxidant defenses. PMID:22493435

  16. CEP290 gene transfer rescues Leber Congenital Amaurosis cellular phenotype

    PubMed Central

    Burnight, E.R.; Wiley, L.A.; Drack, A.V.; Braun, T.A.; Anfinson, K.R.; Kaalberg, E.E.; Halder, J.A.; Affatigato, L.M.; Mullins, R.F.; Stone, E.M.; Tucker, B.A.

    2014-01-01

    Mutations in CEP290 are the most common cause of Leber congenital amaurosis (LCA), a severe inherited retinal degenerative disease for which there is currently no cure. Autosomal recessive CEP290-associated LCA is a good candidate for gene-replacement therapy, and cells derived from affected individuals give researchers the ability to study human disease and therapeutic gene correction in vitro. Here we report the development of lentiviral vectors carrying full-length CEP290 for the purpose of correcting the CEP290 disease-specific phenotype in human cells. A lentiviral vector containing CMV-driven human full-length CEP290 was constructed. Following transduction of patient-specific, iPSC-derived, photoreceptor precursor cells, rt-PCR analysis and western blotting revealed vector-derived expression. Because CEP290 is important in ciliogenesis, the ability of fibroblast cultures from CEP290-associated LCA patients to form cilia was investigated. In cultures derived from these patients, fewer cells formed cilia compared to unaffected controls. Cilia that were formed were shorter in patient derived cells than in cells from unaffected individuals. Importantly, lentiviral delivery of CEP290 rescued the ciliogenesis defect. The successful construction and viral transfer of full-length CEP290 brings us closer to the goal of providing gene- and cell- based therapies for patients affected with this common form of LCA. PMID:24807808

  17. The low-density lipoprotein receptor gene family: a cellular Swiss army knife?

    Microsoft Academic Search

    Anders Nykjaer; Thomas E. Willnow

    2002-01-01

    The low-density lipoprotein receptor gene family is an evolutionarily conserved group of cell-surface receptors produced by mammals and other organisms. Initially thought to be endocytic receptors that mediate the uptake of lipoproteins, recent findings have shown that these receptors have other roles in a range of cellular processes. Among other activities, members of this family act as signal transducers in

  18. Upregulation of Autophagy-Related Gene-5 (ATG-5) Is Associated with Chemoresistance in Human Gastric Cancer

    PubMed Central

    Ge, Jie; Chen, Zihua; Huang, Jin; Chen, Jinxiang; Yuan, Weijie; Deng, Zhenghao; Chen, Zhikang

    2014-01-01

    Autophagy-related gene-5 (ATG-5) is one of the key regulators of autophagic cell death. It has been widely regarded as a protective molecular mechanism for tumor cells during the course of chemotherapy. In the present study, we investigated the expression pattern of ATG-5 and multidrug resistance-associated protein-1 (MRP-1) in 135 gastric cancers (GC) patients who were treated with epirubicin, cisplatin and 5-FU adjuvant chemotherapy (ECF) following surgical resection and explored their potential clinical significance. We found that both ATG-5 (77.78%) and MRP-1 (79.26%) were highly expressed in GC patients. ATG-5 expression was significantly associated with depth of wall invasion, TNM stages and distant metastasis of GC (P<0.05), whereas MRP-1 expression was significantly linked with tumor size, depth of wall invasion, lymph node metastasis, TNM stages and differentiation status (P<0.05). ATG-5 expression was positively correlated with MRP-1 (rp?=?0.616, P<0.01). Increased expression of ATG-5 and MPR-1 was significantly correlated with poor overall survival (OS; P<0.01) and disease free survival (DFS; P<0.01) of our GC cohort. Furthermore, we demonstrated that ATG-5 was involved in drug resistant of GC cells, which was mainly through regulating autophagy. Our data suggest that upregulated expression of ATG-5, an important molecular feature of protective autophagy, is associated with chemoresistance in GC. Expression of ATG-5 and MRP-1 may be independent prognostic markers for GC treatment. PMID:25329677

  19. IL-4 Up-Regulates MiR-21 and the MiRNAs Hosted in the CLCN5 Gene in Chronic Lymphocytic Leukemia.

    PubMed

    Ruiz-Lafuente, Natalia; Alcaraz-García, María-José; Sebastián-Ruiz, Silvia; García-Serna, Azahara-María; Gómez-Espuch, Joaquín; Moraleda, José-María; Minguela, Alfredo; García-Alonso, Ana-María; Parrado, Antonio

    2015-01-01

    Interleukin 4 (IL-4) induces B-cell differentiation and survival of chronic lymphocytic leukemia (CLL) cells. MicroRNAs (miRNAs) regulate mRNA and protein expression, and several miRNAs, deregulated in CLL, might play roles as oncogenes or tumor suppressors. We have studied the miRNA profile of CLL, and its response to IL-4, by oligonucleotide microarrays, resulting in the detection of a set of 129 mature miRNAs consistently expressed in CLL, which included 41 differentially expressed compared to normal B cells (NBC), and 6 significantly underexpressed in ZAP-70 positive patients. IL-4 stimulation brought about up-regulation of the 5p and 3p mature variants of the miR-21 gene, which maps immediately downstream to the VMP1 gene, and of the mature forms generated from the miR-362 (3p and 5p), miR-500a (3p), miR-502 (3p), and miR-532 (3p and 5p) genes, which map within the third intron of the CLCN5 gene. Both genes are in turn regulated by IL-4, suggesting that these miRNAs were regulated by IL-4 as passengers from their carrier genes. Their levels of up-regulation by IL-4 significantly correlated with cytoprotection. MiR-21 has been reported to be leukemogenic, associated to bad prognosis in CLL, and the miRNA more frequently overexpressed in human cancer. Up-regulation by IL-4 of miR-21 and the miRNAs hosted in the CLCN5 locus may contribute to evasion of apoptosis of CLL cells. These findings indicate that the IL-4 pathway and the miRNAs induced by IL-4 are promising targets for the development of novel therapies in CLL. PMID:25909590

  20. IL-4 Up-Regulates MiR-21 and the MiRNAs Hosted in the CLCN5 Gene in Chronic Lymphocytic Leukemia

    PubMed Central

    Sebastián-Ruiz, Silvia; García-Serna, Azahara-María; Gómez-Espuch, Joaquín; Moraleda, José-María; Minguela, Alfredo; García-Alonso, Ana-María; Parrado, Antonio

    2015-01-01

    Interleukin 4 (IL-4) induces B-cell differentiation and survival of chronic lymphocytic leukemia (CLL) cells. MicroRNAs (miRNAs) regulate mRNA and protein expression, and several miRNAs, deregulated in CLL, might play roles as oncogenes or tumor suppressors. We have studied the miRNA profile of CLL, and its response to IL-4, by oligonucleotide microarrays, resulting in the detection of a set of 129 mature miRNAs consistently expressed in CLL, which included 41 differentially expressed compared to normal B cells (NBC), and 6 significantly underexpressed in ZAP-70 positive patients. IL-4 stimulation brought about up-regulation of the 5p and 3p mature variants of the miR-21 gene, which maps immediately downstream to the VMP1 gene, and of the mature forms generated from the miR-362 (3p and 5p), miR-500a (3p), miR-502 (3p), and miR-532 (3p and 5p) genes, which map within the third intron of the CLCN5 gene. Both genes are in turn regulated by IL-4, suggesting that these miRNAs were regulated by IL-4 as passengers from their carrier genes. Their levels of up-regulation by IL-4 significantly correlated with cytoprotection. MiR-21 has been reported to be leukemogenic, associated to bad prognosis in CLL, and the miRNA more frequently overexpressed in human cancer. Up-regulation by IL-4 of miR-21 and the miRNAs hosted in the CLCN5 locus may contribute to evasion of apoptosis of CLL cells. These findings indicate that the IL-4 pathway and the miRNAs induced by IL-4 are promising targets for the development of novel therapies in CLL. PMID:25909590

  1. An efficient method for in vitro gene delivery via regulation of cellular endocytosis pathway

    PubMed Central

    Luo, Jing; Li, Caixia; Chen, Jianlin; Wang, Gang; Gao, Rong; Gu, Zhongwei

    2015-01-01

    Transfection efficiency was the primary goal for in vitro gene delivery mediated by nonviral gene carriers. Here, we report a modified gene transfection method that could greatly increase the efficiency of, and accelerate the process mediated by, 25 kDa branched polyethyleneimine and Lipofectamine™ 2000 in a broad range of cell strains, including tumor, normal, primary, and embryonic stem cells. In this method, the combination of transfection procedure with optimized complexation volume had a determinant effect on gene delivery result. The superiorities of the method were found to be related to the change of cellular endocytosis pathway and decrease of particle size. The efficient and simple method established in this study can be widely used for in vitro gene delivery into cultured cells. We think it may also be applicable for many more nonviral gene delivery materials than polyethyleneimine and liposome. PMID:25767387

  2. Regulated expression of CXCR4 constitutive active mutants revealed the up-modulated chemotaxis and up-regulation of genes crucial for CXCR4 mediated homing and engraftment of hematopoietic stem/progenitor cells

    PubMed Central

    Sharma, M; Afrin, F; Tripathi, RP; Gangenahalli, G

    2013-01-01

    SDF-1/CXCR4 axis plays a principle role in the homing and engraftment of hematopoietic stem/progenitor cells (HSPCs), a process that defines cells ability to reach and seed recipient bone marrow niche following their intravenous infusion. However, the proper functioning of CXCR4 downstream signaling depends upon consistent optimal expression of both SDF-1 ligand and its receptor CXCR4, which in turn is variable and regulated by several factors. The constitutive active mutants of CXCR4 (N119A and N119S) being able to induce autonomous downstream signaling, overcome the limitation of ligand-receptor interaction for induction of CXCR4 signaling. Therefore, we intended to explore their potential in Chemotaxis; a key cellular process which crucially regulates cells homing to bone marrow. In present study, Tet-on inducible gene expression vector system was used for doxycycline inducible regulated transgene expression of CXCR4 active mutants in hematopoietic stem progenitor cell line K-562. Both of these mutants revealed significantly enhanced Chemotaxis to SDF-1 gradient as compared to wild type. Furthermore, gene expression profiling of these genetically engineered cells as assessed by microarray analysis revealed the up-regulation of group of genes that are known to play a crucial role in CXCR4 mediated cells homing and engraftment. Hence, this study suggest the potential prospects of CXCR4 active mutants in research and development aimed to improve the efficiency of cells in the mechanism of homing and engraftment process. PMID:24693205

  3. 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 of assays to test the quality of MSCs before clinical use. PMID:24780490

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

    SciTech Connect

    Prendergast, G.C.; Cole, M.D. (Princeton Univ., NJ (USA). 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.

  5. Genes Upregulated in Winter Wheat (Triticum aestivum L.) during Mild Freezing and Subsequent Thawing Suggest Sequential Activation of Multiple Response Mechanisms

    PubMed Central

    Skinner, Daniel Z.

    2015-01-01

    Exposing fully cold-acclimated wheat plants to a mild freeze-thaw cycle of ?3°C for 24h followed by +3°C for 24 or 48h results in dramatically improved tolerance of subsequent exposure to sub-freezing temperatures. Gene enrichment analysis of crown tissue from plants collected before or after the ?3°C freeze or after thawing at +3°C for 24 or 48h revealed that many biological processes and molecular functions were activated during the freeze-thaw cycle in an increasing cascade of responses such that over 150 processes or functions were significantly enhanced by the end of the 48 h, post-freeze thaw. Nearly 2,000 individual genes were upregulated more than 2-fold over the 72 h course of freezing and thawing, but more than 70% of these genes were upregulated during only one of the time periods examined, suggesting a series of genes and gene functions were involved in activation of the processes that led to enhanced freezing tolerance. This series of functions appeared to include extensive cell signaling, activation of stress response mechanisms and the phenylpropanoid biosynthetic pathway, extensive modification of secondary metabolites, and physical restructuring of cell membranes. By identifying plant lines that are especially able to activate these multiple mechanisms it may be possible to develop lines with enhanced winterhardiness. PMID:26173115

  6. Stochastic Fluctuations and Distributed Control of Gene Expression Impact Cellular Memory

    PubMed Central

    Corre, Guillaume; Stockholm, Daniel; Arnaud, Ophélie; Kaneko, Gaël; Viñuelas, José; Yamagata, Yoshiaki; Neildez-Nguyen, Thi My Anh; Kupiec, Jean-Jacques; Beslon, Guillaume; Gandrillon, Olivier; Paldi, András

    2014-01-01

    Despite the stochastic noise that characterizes all cellular processes the cells are able to maintain and transmit to their daughter cells the stable level of gene expression. In order to better understand this phenomenon, we investigated the temporal dynamics of gene expression variation using a double reporter gene model. We compared cell clones with transgenes coding for highly stable mRNA and fluorescent proteins with clones expressing destabilized mRNA-s and proteins. Both types of clones displayed strong heterogeneity of reporter gene expression levels. However, cells expressing stable gene products produced daughter cells with similar level of reporter proteins, while in cell clones with short mRNA and protein half-lives the epigenetic memory of the gene expression level was completely suppressed. Computer simulations also confirmed the role of mRNA and protein stability in the conservation of constant gene expression levels over several cell generations. These data indicate that the conservation of a stable phenotype in a cellular lineage may largely depend on the slow turnover of mRNA-s and proteins. PMID:25531401

  7. Identification of genes up-regulated by retinoic-acid-induced differentiation of the human neuronal precursor cell line NTERA-2 cl.D1.

    PubMed

    Leypoldt, F; Lewerenz, J; Methner, A

    2001-02-01

    The human teratocarcinoma cell line NTERA-2 cl.D1 (NT2 cells) can be induced with retinoic acid and cell aggregation to yield postmitotic neurones. This seems to model the in vivo situation, as high concentrations of retinoic acid, retinoic acid binding proteins, and receptors have been detected in the embryonic CNS and the developing spinal cord suggesting a role for retinoic acid in neurogenesis. Suppression subtractive hybridization was used to detect genes up-regulated by this paradigm of neuronal differentiation. Microfibril-associated glycoprotein 2 was found to be drastically up-regulated and has not been implicated in neuronal differentiation before. Suppression subtractive hybridization also identified DYRK4, a homologue of the Drosophila gene minibrain. Minibrain mutations result in specific defects in the development of the fly central nervous system. In adult rats, DYRK4 is only expressed in testis, but our results suggest an additional role for DYRK4 in neuronal differentiation. We have shown that suppression subtractive hybridization in conjunction with an efficient screening procedure is a valuable tool to produce a repertoire of differentially expressed genes and propose a new physiological role for several identified genes and expressed sequence tags. PMID:11158252

  8. Dietary fermentable fiber upregulated immune related genes expression, increased innate immune response and resistance of rainbow trout (Oncorhynchus mykiss) against Aeromonas hydrophila.

    PubMed

    Yarahmadi, Peyman; Kolangi Miandare, Hamed; Farahmand, Hamid; Mirvaghefi, Alireza; Hoseinifar, Seyed Hossein

    2014-12-01

    This trial was carried out to investigate the effects of dietary administration of Vitacel(®), a commercial fermentable fiber, on immune related genes (Lysozyme, TNF? and HSP70) expression, innate immune response and resistance of rainbow trout against Aeromonas hydrophila. 120 healthy rainbow trout (81.65 ± 1.49 g) were distributed in six fiberglass tanks assigned to two treatments. The treatments were feeding rainbow trout with diets supplemented with 0 (control) or 10 g kg(-1) Vitacel(®) for 45 days. The results revealed that administration of fermentable fiber significantly (P < 0.05) upregulated lysozyme and TNF? gene expression. HSP70 gene expression was significantly lower in Vitacel(®) fed fish at the end of trial (P < 0.05). Furthermore dietary administrations of Vitacel(®) remarkably elevated rainbow trout innate immune parameters include serum lysozyme, ACH50, bactericidal activity and agglutination antibody titer (P < 0.05). Administration of 10 g kg(-1) Vitacel(®) significantly increased rainbow trout resistance against A. hydrophila (P < 0.05). The results of present study revealed that dietary Vitacel(®) can upregulates immune related genes expression and elevates innate immune response and disease resistance of rainbow trout. PMID:25218276

  9. YPEL3, a p53-regulated gene that induces cellular senescence

    PubMed Central

    Kelley, Kevin; Miller, Kelly R.; Todd, Amber; Kelley, Amy; Tuttle, Rebecca

    2010-01-01

    Cellular senescence, the limited ability of cultured normal cells to divide, can result from cellular damage triggered through oncogene activation (premature senescence) or the loss of telomeres following successive rounds of DNA replication (replicative senescence). While both processes require a functional p53 signaling pathway, relevant downstream p53 targets have been difficult to identify. Discovery of senescence activators is important because induction of tumor cell senescence may represent a therapeutic approach for the treatment of cancer. In microarray studies where p53 was reactivated in MCF7 cells, we discovered that YPEL3 (Yippee-like-3), a member of a recently discovered family of putative zinc finger motif coding genes consisting of YPEL1-5, is a p53-regulated gene. YPEL3 expression induced by DNA damage leads to p53 recruitment to a cis-acting DNA response element located near the human YPEL3 promoter. Physiological induction of YPEL3 results in a substantial decrease in cell viability associated with an increase in cellular senescence. Through the use of RNAi and H-ras induction of cellular senescence, we demonstrate that YPEL3 activates cellular senescence downstream of p53. Consistent with its growth suppressive activity, YPEL3 gene expression is repressed in ovarian tumor samples. One mechanism of YPEL3 downregulation in ovarian tumor cell lines appears to be hypermethylation of a CpG island upstream of the YPEL3 promoter. We believe these findings point to YPEL3 being a novel tumor suppressor, which upon induction triggers a permanent growth arrest in human tumor and normal cells. PMID:20388804

  10. MicroRNA-31 controls phenotypic modulation of human vascular smooth muscle cells by regulating its target gene cellular repressor of E1A-stimulated genes

    SciTech Connect

    Wang, Jie, E-mail: wj2170@qq.com [Xijing Hospital, Fourth Military Medical University, Xi'an 710032 (China); Cardiovascular Research Institute and Key Laboratory of Cardiology, Shenyang Northern Hospital, Shenyang 110840 (China); Yan, Cheng-Hui, E-mail: yanch1029@163.com [Cardiovascular Research Institute and Key Laboratory of Cardiology, Shenyang Northern Hospital, Shenyang 110840 (China); Li, Yang, E-mail: liyang19830925@126.com [Cardiovascular Research Institute and Key Laboratory of Cardiology, Shenyang Northern Hospital, Shenyang 110840 (China); Xu, Kai, E-mail: xukai2001@gmail.com [Cardiovascular Research Institute and Key Laboratory of Cardiology, Shenyang Northern Hospital, Shenyang 110840 (China); Tian, Xiao-Xiang, E-mail: tian_xx@163.com [Cardiovascular Research Institute and Key Laboratory of Cardiology, Shenyang Northern Hospital, Shenyang 110840 (China); Peng, Cheng-Fei, E-mail: pengchengfei2000@126.com [Cardiovascular Research Institute and Key Laboratory of Cardiology, Shenyang Northern Hospital, Shenyang 110840 (China); Tao, Jie, E-mail: taojie1976@163.com [Cardiovascular Research Institute and Key Laboratory of Cardiology, Shenyang Northern Hospital, Shenyang 110840 (China); Sun, Ming-Yu, E-mail: sunmingyu1976@126.com [Cardiovascular Research Institute and Key Laboratory of Cardiology, Shenyang Northern Hospital, Shenyang 110840 (China); Han, Ya-Ling, E-mail: yalinghan@gmail.com [Cardiovascular Research Institute and Key Laboratory of Cardiology, Shenyang Northern Hospital, Shenyang 110840 (China)

    2013-05-01

    Phenotypic modulation of vascular smooth muscle cells (VSMCs) plays a critical role in the pathogenesis of a variety of proliferative vascular diseases. The cellular repressor of E1A-stimulated genes (CREG) has been shown to play an important role in phenotypic modulation of VSMCs. However, the mechanism regulating CREG upstream signaling remains unclear. MicroRNAs (miRNAs) have recently been found to play a critical role in cell differentiation via target-gene regulation. This study aimed to identify a miRNA that binds directly to CREG, and may thus be involved in CREG-mediated VSMC phenotypic modulation. Computational analysis indicated that miR-31 bound to the CREG mRNA 3? untranslated region (3?-UTR). miR-31 was upregulated in quiescent differentiated VSMCs and downregulated in proliferative cells stimulated by platelet-derived growth factor and serum starvation, demonstrating a negative relationship with the VSMC differentiation marker genes, smooth muscle ?-actin, calponin and CREG. Using gain-of-function and loss-of-function approaches, CREG and VSMC differentiation marker gene expression levels were shown to be suppressed by a miR-31 mimic, but increased by a miR-31 inhibitor at both protein and mRNA levels. Notably, miR-31 overexpression or inhibition affected luciferase expression driven by the CREG 3?-UTR containing the miR-31 binding site. Furthermore, miR-31-mediated VSMC phenotypic modulation was inhibited in CREG-knockdown human VSMCs. We also determined miR-31 levels in the serum of patients with coronary artery disease (CAD), with or without in stent restenosis and in healthy controls. miR-31 levels were higher in the serum of CAD patients with restenosis compared to CAD patients without restenosis and in healthy controls. In summary, these data demonstrate that miR-31 not only directly binds to its target gene CREG and modulates the VSMC phenotype through this interaction, but also can be an important biomarker in diseases involving VSMC phenotypic modulation. These novel findings may have extensive implications for the diagnosis and therapy of a variety of proliferative vascular diseases. - Highlights: ? MiR-31 modulates CREG expression by binding directly to the human CREG mRNA 3?-UTR. ? MiR-31 mediates the human VSMC phenotypic modulation by regulating the expression of human CREG. ? Serum miR-31 may act as an important biomarker in diseases involving in stent restenosis after PCI.

  11. Long terminal repeat regions from exogenous but not endogenous feline leukemia viruses transactivate cellular gene expression.

    PubMed

    Ghosh, S K; Roy-Burman, P; Faller, D V

    2000-10-01

    We have previously reported that the long terminal repeat (LTR) region of feline leukemia viruses (FeLVs) can enhance expression of certain cellular genes such as the collagenase IV gene and MCP-1 in trans (S. K. Ghosh and D. V. Faller, J. Virol. 73:4931-4940, 1999). Genomic DNA of all healthy feline species also contains LTR-like sequences that are related to exogenous FeLV LTRs. In this study, we evaluated the cellular gene transactivational potential of these endogenous FeLV LTR sequences. Unlike their exogenous FeLV counterparts, neither nearly full-length endogenous FeLV molecular clones (CFE-6 and CFE-16) nor their isolated LTRs were able to activate collagenase IV gene or MCP-1 expression in transient transfection assays. We had also demonstrated previously that production of an RNA transcript from exogenous FeLV LTRs correlates with their transactivational activity. In the present study, we demonstrate that the endogenous FeLV LTRs do not generate LTR-specific RNA transcripts in the feline embryo fibroblast cell line AH927. Furthermore, infection of AH927 cells by an exogenous FeLV subgroup A virus did not induce production of such LTR-specific transcripts from the endogenous proviral genomes, although the LTR-specific transcripts from the exogenous virus were readily detected. Finally, LTR-specific transcripts were not generated in BALB/3T3 cells transiently transfected with isolated CFE-6 LTR, in contrast to transfections with LTRs from exogenous viruses. Our data thus suggest that the inability of endogenous FeLV LTRs in gene transactivation is not due to cell line specificity or presence of any upstream inhibitory cis-acting element. Endogenous, nonleukemogenic FeLV LTRs, therefore, do not transactivate cellular gene expression, and this property appears to be specific to exogenous, leukemogenic FeLVs. PMID:11000248

  12. Long Terminal Repeat Regions from Exogenous but Not Endogenous Feline Leukemia Viruses Transactivate Cellular Gene Expression

    PubMed Central

    Ghosh, Sajal K.; Roy-Burman, Pradip; Faller, Douglas V.

    2000-01-01

    We have previously reported that the long terminal repeat (LTR) region of feline leukemia viruses (FeLVs) can enhance expression of certain cellular genes such as the collagenase IV gene and MCP-1 in trans (S. K. Ghosh and D. V. Faller, J. Virol. 73:4931–4940, 1999). Genomic DNA of all healthy feline species also contains LTR-like sequences that are related to exogenous FeLV LTRs. In this study, we evaluated the cellular gene transactivational potential of these endogenous FeLV LTR sequences. Unlike their exogenous FeLV counterparts, neither nearly full-length endogenous FeLV molecular clones (CFE-6 and CFE-16) nor their isolated LTRs were able to activate collagenase IV gene or MCP-1 expression in transient transfection assays. We had also demonstrated previously that production of an RNA transcript from exogenous FeLV LTRs correlates with their transactivational activity. In the present study, we demonstrate that the endogenous FeLV LTRs do not generate LTR-specific RNA transcripts in the feline embryo fibroblast cell line AH927. Furthermore, infection of AH927 cells by an exogenous FeLV subgroup A virus did not induce production of such LTR-specific transcripts from the endogenous proviral genomes, although the LTR-specific transcripts from the exogenous virus were readily detected. Finally, LTR-specific transcripts were not generated in BALB/3T3 cells transiently transfected with isolated CFE-6 LTR, in contrast to transfections with LTRs from exogenous viruses. Our data thus suggest that the inability of endogenous FeLV LTRs in gene transactivation is not due to cell line specificity or presence of any upstream inhibitory cis-acting element. Endogenous, nonleukemogenic FeLV LTRs, therefore, do not transactivate cellular gene expression, and this property appears to be specific to exogenous, leukemogenic FeLVs. PMID:11000248

  13. Signatures of gene expression noise in cellular systems Julia Rausenberger a,b,*, Christian Fleck b,c

    E-print Network

    Timmer, Jens

    Review Signatures of gene expression noise in cellular systems Julia Rausenberger a,b,*, Christian t i c l e i n f o Article history: Available online 11 June 2009 Keywords: Gene expression noise Stochastic modeling Transcription factor Population distribution a b s t r a c t Noise in gene expression

  14. Accelerated methylation of ribosomal RNA genes during the cellular senescence of Werner syndrome fibroblasts

    Microsoft Academic Search

    AMRITA MACHWE; DAVID K. ORREN; VILHELM A. BOHR

    2000-01-01

    Ribosomal DNA (rDNA) metabolism has been implicated in cellular and organismal aging. The role of rDNA in premature and normal human aging was investigated by measuring rDNA gene copy number, the level of rDNA methylation, and rRNA expression during the in vitro senescence of primary fibroblasts from normal (young and old) donors and from Werner syndrome (WS) patients. In compari-

  15. Upregulation of heat shock protein genes by envenomation of ectoparasitoid Bracon hebetor in larval host of Indian meal moth Plodia interpunctella.

    PubMed

    Shim, Jae-Kyoung; Ha, Dae-Myung; Nho, Si-Kab; Song, Kyung-Sik; Lee, Kyeong-Yeoll

    2008-03-01

    Effect of envenomation of ectoparasitoid Bracon hebetor was determined on the heart rate and the expression of shsp, hsc70 and hsp90 of the lepidopteran host Plodia interpunctella. Envenomated host larvae were promptly immobilized but heart rate was not changed until 4 days after envenomation. Northern hybridization showed that each hsp gene was differentially influenced by envenomation: continued high induction of shsp, gradual strong induction of hsc70, but no induction of hsp90. Our results suggest that upregulation of both shsp and hsc70 may produce potent factors that have important roles in the mechanism of host-parasitoid relationship. PMID:17981295

  16. Cellular Gene Expression Survey of Vaccinia Virus Infection of Human HeLa Cells

    PubMed Central

    Guerra, Susana; López-Fernández, Luis A.; Pascual-Montano, Alberto; Muñoz, Manuel; Harshman, Keith; Esteban, Mariano

    2003-01-01

    Vaccinia virus (VV) is a cytocidal virus that causes major changes in host cell machinery shortly after infecting cells. To define the consequences of virus infection on host gene expression, we used microarrays of approximately 15,000 human cDNAs to examine expression levels of mRNAs isolated at 2, 6, and 16 h postinfection from cultures of infected HeLa cells. The majority of profiling changes during VV infection corresponded to downregulation of genes at 16 h postinfection. Differentially expressed genes were clustered into seven groups to identify common regulatory pathways, with most of them (90%) belonging to clusters 6 and 7, which represent genes whose expression was repressed after infection. Cluster 1, however, contained 37 transcripts (2.81%) showing a robust pattern of induction that was maintained during the course of infection. Genes in cluster 1 included those for Wiskott-Aldrich syndrome protein (WASP) family member WASF1, thymosine, adenosine A2a receptor, glutamate decarboxylase 2, CD-80 antigen, KIAA0888 protein, selenophosphate synthetase, pericentrin, and attractin as well as several expressed sequence tags. We analyzed in more detail the fate of WASP protein in VV-infected cells, because a related family member, N-WASP, is involved in viral motility. WASP protein accumulated in the course of infection; its increase required viral DNA replication and de novo protein synthesis, and it localized in cytoplasmic structures distinct from uninfected cells. This study is the first quantitative analysis of host gene expression following VV infection of cultured human cells, demonstrating global changes in the expression profile, and identifies upregulated genes with potential roles in the virus replication cycle. PMID:12743306

  17. Upregulation of human telomerase reverse transcriptase mRNA expression by in vitro transfection of hepatitis B virus X gene into human hepatocarcinoma and cholangiocarcinoma cells

    PubMed Central

    Qu, Zhen-Liang; Zou, Sheng-Quan; Cui, Nai-Qiang; Wu, Xian-Zhong; Qin, Ming-Fang; Kong, Di; Zhou, Zhen-Li

    2005-01-01

    AIM: To study the changes of human telomerase reverse transcriptase (hTERT) mRNA expression in human hepatocarcinoma cell lines (HepG2) and cholangiocarcinoma cell lines (QBC939) after HBx gene transfection and to illustrate the significance of transcriptional regulation of hTERT gene by HBx gene in the carcinogenesis. METHODS: HepG2 and QBC939 cell lines were cultured and co-transfected with eukaryotic expression vector containing the HBx coding region and cloning vector containing enhanced green fluorescent protein (EGFP) coding sequence using lipid-mediated gene transduction technique. Thirty-six hours after transfection, EGFP expression in cells was used as the indicator of successful transfection. Flow cytometry was performed to determine the transfection efficiency. Cells were harvested and total RNA was extracted using TRIzol® reagent. The expression of hTERT mRNA in HepG2 and QBC939 cell lines was assayed by reverse transcription-polymerase chain reaction. The expression of HBx protein in both cell lines was detected by immunocytochemical staining and Western blotting. RESULTS: Flow cytometry showed that the transfection efficiency was 46.4% in HepG2 cells and 29.6% in QBC939 cells for both HBx gene expression vector and blank vector. The expression of hTERT mRNA was meaningfully increased in HepG2 and QBC939 cell lines when transfected with HBx gene expression vector compared to those transfected with OPTI-MEM medium and blank vector. Immunocytochemical staining and Western blotting revealed HBx protein expression in HepG2 and QBC939 cells only when transfected with HBx gene. CONCLUSION: HBx gene transfection can upregulate the transcriptional expression of hTERT mRNA. The transactiv-ation of hTERT gene by HBx gene is a newfound mechanism for pathogenesis of hepatocarcinomas and cholangioca-rcinomas after HBV infection. PMID:16237755

  18. Coordinate up-regulation of low-density lipoprotein receptor and cyclo-oxygenase-2 gene expression in human colorectal cells and in colorectal adenocarcinoma biopsies

    NASA Technical Reports Server (NTRS)

    Lum, D. F.; McQuaid, K. R.; Gilbertson, V. L.; Hughes-Fulford, M.

    1999-01-01

    Many colorectal cancers have high levels of cyclo-oxygenase 2 (COX-2), an enzyme that metabolizes the essential fatty acids into prostaglandins. Since the low-density lipoprotein receptor (LDLr) is involved in the uptake of essential fatty acids, we studied the effect of LDL on growth and gene regulation in colorectal cancer cells. DiFi cells grown in lipoprotein-deficient sera (LPDS) grew more slowly than cells with LDL. LDLr antibody caused significant inhibition of tumor cell growth but did not affect controls. In addition, LDL uptake did not change in the presence of excess LDL, suggesting that ldlr mRNA lacks normal feedback regulation in some colorectal cancers. Analysis of the ldlr mRNA showed that excess LDL in the medium did not cause down-regulation of the message even after 24 hr. The second portion of the study examined the mRNA expression of ldlr and its co-regulation with cox-2 in normal and tumor specimens from patients with colorectal adenocarcinomas. The ratio of tumor:paired normal mucosa of mRNA expression of ldlr and of cox-2 was measured in specimens taken during colonoscopy. ldlr and cox-2 transcripts were apparent in 11 of 11 carcinomas. There was significant coordinate up-regulation both of ldlr and of cox-2 in 6 of 11 (55%) tumors compared with normal colonic mucosa. There was no up-regulation of cox-2 without concomitant up-regulation of ldlr. These data suggest that the LDLr is abnormally regulated in some colorectal tumors and may play a role in the up-regulation of cox-2. Copyright 1999 Wiley-Liss, Inc.

  19. Transcriptional up-regulation of antioxidant genes by PPAR{delta} inhibits angiotensin II-induced premature senescence in vascular smooth muscle cells

    SciTech Connect

    Kim, Hyo Jung; Ham, Sun Ah [Department of Pharmacology, Gyeongsang Institute of Health Science, Gyeongsang National University School of Medicine, Jinju (Korea, Republic of)] [Department of Pharmacology, Gyeongsang Institute of Health Science, Gyeongsang National University School of Medicine, Jinju (Korea, Republic of); Paek, Kyung Shin [Department of Nursing, Semyung University, Jechon (Korea, Republic of)] [Department of Nursing, Semyung University, Jechon (Korea, Republic of); Hwang, Jung Seok; Jung, Si Young; Kim, Min Young; Jin, Hanna; Kang, Eun Sil; Woo, Im Sun; Kim, Hye Jung; Lee, Jae Heun; Chang, Ki Churl [Department of Pharmacology, Gyeongsang Institute of Health Science, Gyeongsang National University School of Medicine, Jinju (Korea, Republic of)] [Department of Pharmacology, Gyeongsang Institute of Health Science, Gyeongsang National University School of Medicine, Jinju (Korea, Republic of); Han, Chang Woo [Department of Internal Medicine, Pusan National University School of Korean Medicine, Yangsan (Korea, Republic of)] [Department of Internal Medicine, Pusan National University School of Korean Medicine, Yangsan (Korea, Republic of); Seo, Han Geuk, E-mail: hgseo@gnu.ac.kr [Department of Pharmacology, Gyeongsang Institute of Health Science, Gyeongsang National University School of Medicine, Jinju (Korea, Republic of)

    2011-03-25

    Research highlights: {yields} Activation of PPAR{delta} by GW501516 significantly inhibited Ang II-induced premature senescence in hVSMCs. {yields} Agonist-activated PPAR{delta} suppressed generation of Ang II-triggered ROS with a concomitant reduction in DNA damage. {yields} GW501516 up-regulated expression of antioxidant genes, such as GPx1, Trx1, Mn-SOD and HO-1. {yields} Knock-down of these antioxidant genes abolished the effects of GW501516 on ROS production and premature senescence. -- Abstract: This study evaluated peroxisome proliferator-activated receptor (PPAR) {delta} as a potential target for therapeutic intervention in Ang II-induced senescence in human vascular smooth muscle cells (hVSMCs). Activation of PPAR{delta} by GW501516, a specific agonist of PPAR{delta}, significantly inhibited the Ang II-induced premature senescence of hVSMCs. Agonist-activated PPAR{delta} suppressed the generation of Ang II-triggered reactive oxygen species (ROS) with a concomitant reduction in DNA damage. Notably, GW501516 up-regulated the expression of antioxidant genes, such as glutathione peroxidase 1, thioredoxin 1, manganese superoxide dismutase and heme oxygenase 1. siRNA-mediated down-regulation of these antioxidant genes almost completely abolished the effects of GW501516 on ROS production and premature senescence in hVSMCs treated with Ang II. Taken together, the enhanced transcription of antioxidant genes is responsible for the PPAR{delta}-mediated inhibition of premature senescence through sequestration of ROS in hVSMCs treated with Ang II.

  20. Iron deficiency upregulates Egr1 expression.

    PubMed

    Lee, Seung-Min; Lee, Sun Bok; Prywes, Ron; Vulpe, Christopher D

    2015-07-01

    Iron-deficient anemia is a prevalent disease among humans. We searched for genes regulated by iron deficiency and its regulated mechanism. cDNA microarrays were performed using Hepa1c1c7 cells treated with 100 ?M desferrioxamine (DFO), an iron chelator. Early growth response 1 (Egr1) was upregulated with at least 20-fold increase within 4 h and lasted for 24 h, which was confirmed by qRT-PCR. This activation was not seen by ferric ammonium citrate (FAC). DFO increased the transcriptional activity of Egr1-luc (-604 to +160) and serum response element (SRE)-luc reporters by 2.7-folds. In addition, cycloheximide lowered DFO-induced Egr1 mRNA levels. The upregulation of Egr1 by DFO was accompanied by sustained ERK signals along with phosphorylation of Elk-1. The ERK inhibitor (PD98059) prevented the DFO-induced Egr1 mRNAs. Overexpression of Elk-1 mutant (pElk-1S383A) decreased Egr1 reporter activity. DFO lowered reactive oxygen species (ROS) production and increased caspase 3/7 activity and cell death. DFO-induced iron deficiency upregulates Egr1 in part through transcriptional activation via ERK and Elk-1 signals, which may be important in the regulation of cell death in hepatoma cells. Our study demonstrated that iron depletion controlled the expression of Egr1, which might contribute to decisions about cellular fate in response to iron deficiency. PMID:25981695

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

    PubMed

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

    2014-10-01

    Gene-directed enzyme prodrug therapy (GDEPT) consists of targeted delivery to tumor cells of a suicide gene responsible for the in situ conversion of a prodrug into cytotoxic metabolites. One of the major impediments of GDEPT is to target specifically the tumor cells with the suicide gene. Among gene delivery methods, mesenchymal stem cells (MSCs) have emerged recently as potential cellular vehicles for gene delivery. MSCs are particularly suited for gene transduction. They exhibit remarkable migratory property towards tumors and their metastases and they are weakly immunogenic. This review will summarize the current knowledge about MSCs engineered to express different suicide genes (cytosine deaminase, thymidine kinase, carboxylesterase, cytochrome P450) to elicit a significant antitumor response against brain tumors, ovarian, hepatocellular, pancreatic, renal or medullary thyroid carcinomas, breast or prostate cancer and pulmonary metastases. The potential side effects of these MSC-based tumor therapies will also be considered to highlight certain aspects that need to be improved prior to clinical use. PMID:24977933

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

    PubMed

    Nuth, Manunya; Kennedy, Ann R

    2013-07-01

    Ionizing radiation associated with highly energetic and charged heavy (HZE) particles poses a danger to astronauts during space travel. The aim of the present study was to evaluate the patterns of gene expression associated with cellular exposure to low-dose iron ion irradiation, in the presence and absence of L-selenomethionine (SeM). Human thyroid epithelial cells (HTori-3) were exposed to low-dose iron ion (1 GeV/n) irradiation at 10 or 20 cGy with or without SeM pretreatment. The cells were harvested 6 and 16 h post-irradiation and analyzed by the Affymetrix U133Av2 gene chip arrays. Genes exhibiting a 1.5-fold expression cut-off and 5% false discovery rate (FDR) were considered statistically significant and subsequently analyzed using the Database for Annotation, Visualization and Integrated Discovery (DAVID) for pathway analysis. Representative genes were further validated by real-time RT-PCR. Even at low doses of radiation from iron ions, global genome profiling of the irradiated cells revealed the upregulation of genes associated with the activation of stress-related signaling pathways (ubiquitin-mediated proteolysis, p53 signaling, cell cycle and apoptosis), which occurred in a dose-dependent manner. A 24-h pretreatment with SeM was shown to reduce the radiation effects by mitigating stress-related signaling pathways and downregulating certain genes associated with cell adhesion. The mechanism by which SeM prevents radiation-induced transformation in vitro may involve the suppression of the expression of genes associated with stress-related signaling and certain cell adhesion events. PMID:23946774

  3. Developing the concept of adoptive cellular gene therapy of rheumatoid arthritis.

    PubMed

    Tarner, Ingo H; Neumann, Elena; Gay, Steffen; Fathman, C Garrison; Müller-Ladner, Ulf

    2006-02-01

    Progressive destruction of articular cartilage and bone is the pivotal problem of rheumatoid arthritis (RA). Joint destruction is the cause of severe disability and determines the long-term outcome of disease. Conventional therapy does not control this destructive process sufficiently and the anti-rheumatic drugs available today can cause severe systemic adverse effects. Local application of chondroprotective and osteoprotective agents by means of gene therapy would be an attractive alternative to conventional therapy of RA and could provide long-term expression of the therapeutic agents and minimize systemic adverse effects. For this purpose, we have developed the concept of adoptive cellular gene therapy. This treatment strategy is based on using genetically engineered cells that home specifically to sites of autoimmune inflammation and thus allow local delivery of therapeutic gene products. Ex vivo transduction of these cells avoids systemic exposure of the host to the transgene-encoding vector and thus adds to the safety of this approach. In this article of the CIS Spring School in Autoimmune Diseases 2005 proceedings, we review our work on developing the strategy of adoptive cellular gene therapy and summarize recent advances in the evaluation of therapeutic effects and the identification of novel therapeutic targets. PMID:16431349

  4. Real-time Transcriptional Profiling of Cellular and Viral Gene Expression during Lytic Cytomegalovirus Infection

    PubMed Central

    Marcinowski, Lisa; Lidschreiber, Michael; Windhager, Lukas; Rieder, Martina; Bosse, Jens B.; Rädle, Bernd; Bonfert, Thomas; Györy, Ildiko; de Graaf, Miranda; da Costa, Olivia Prazeres; Rosenstiel, Philip; Friedel, Caroline C.; Zimmer, Ralf; Ruzsics, Zsolt; Dölken, Lars

    2012-01-01

    During viral infections cellular gene expression is subject to rapid alterations induced by both viral and antiviral mechanisms. In this study, we applied metabolic labeling of newly transcribed RNA with 4-thiouridine (4sU-tagging) to dissect the real-time kinetics of cellular and viral transcriptional activity during lytic murine cytomegalovirus (MCMV) infection. Microarray profiling on newly transcribed RNA obtained at different times during the first six hours of MCMV infection revealed discrete functional clusters of cellular genes regulated with distinct kinetics at surprising temporal resolution. Immediately upon virus entry, a cluster of NF-?B- and interferon-regulated genes was induced. Rapid viral counter-regulation of this coincided with a very transient DNA-damage response, followed by a delayed ER-stress response. Rapid counter-regulation of all three clusters indicated the involvement of novel viral regulators targeting these pathways. In addition, down-regulation of two clusters involved in cell-differentiation (rapid repression) and cell-cycle (delayed repression) was observed. Promoter analysis revealed all five clusters to be associated with distinct transcription factors, of which NF-?B and c-Myc were validated to precisely match the respective transcriptional changes observed in newly transcribed RNA. 4sU-tagging also allowed us to study the real-time kinetics of viral gene expression in the absence of any interfering virion-associated-RNA. Both qRT-PCR and next-generation sequencing demonstrated a sharp peak of viral gene expression during the first two hours of infection including transcription of immediate-early, early and even well characterized late genes. Interestingly, this was subject to rapid gene silencing by 5–6 hours post infection. Despite the rapid increase in viral DNA load during viral DNA replication, transcriptional activity of some viral genes remained remarkably constant until late-stage infection, or was subject to further continuous decline. In summary, this study pioneers real-time transcriptional analysis during a lytic herpesvirus infection and highlights numerous novel regulatory aspects of virus-host-cell interaction. PMID:22969428

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

    PubMed Central

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

    2015-01-01

    Background Defective cellular transport processes can lead to aberrant accumulation of trace elements, iron, small molecules and hormones in the cell, which in turn may promote the formation of reactive oxygen species, promoting DNA damage and aberrant expression of key regulatory cancer genes. As DNA damage and uncontrolled proliferation are hallmarks of cancer, including epithelial ovarian cancer (EOC), we hypothesized that inherited variation in the cellular transport genes contributes to EOC risk. Methods In total, DNA samples were obtained from 14,525 case subjects with invasive EOC and from 23,447 controls from 43 sites in the Ovarian Cancer Association Consortium (OCAC). Two hundred seventy nine SNPs, representing 131 genes, were genotyped using an Illumina Infinium iSelect BeadChip as part of the Collaborative Oncological Gene-environment Study (COGS). SNP analyses were conducted using unconditional logistic regression under a log-additive model, and the FDR q<0.2 was applied to adjust for multiple comparisons. Results The most significant evidence of an association for all invasive cancers combined and for the serous subtype was observed for SNP rs17216603 in the iron transporter gene HEPH (invasive: OR = 0.85, P = 0.00026; serous: OR = 0.81, P = 0.00020); this SNP was also associated with the borderline/low malignant potential (LMP) tumors (P = 0.021). Other genes significantly associated with EOC histological subtypes (p<0.05) included the UGT1A (endometrioid), SLC25A45 (mucinous), SLC39A11 (low malignant potential), and SERPINA7 (clear cell carcinoma). In addition, 1785 SNPs in six genes (HEPH, MGST1, SERPINA, SLC25A45, SLC39A11 and UGT1A) were imputed from the 1000 Genomes Project and examined for association with INV EOC in white-European subjects. The most significant imputed SNP was rs117729793 in SLC39A11 (per allele, OR = 2.55, 95% CI = 1.5-4.35, p = 5.66x10-4). Conclusion These results, generated on a large cohort of women, revealed associations between inherited cellular transport gene variants and risk of EOC histologic subtypes. PMID:26091520

  6. Upregulation of Mouse Genes in HSV-1 Latent TG after Butyrate Treatment Implicates the Multiple Roles of the LAT-ICP0 Locus

    PubMed Central

    Clement, Christian; Bhattacharjee, Partha S.; Kumar, Manish; Foster, Timothy P.; Thompson, Hilary W.

    2011-01-01

    Purpose. To determine host response by gene expression in HSV-1 latent trigeminal ganglia (TG) after sodium butyrate (NaBu) treatment. Methods. Corneas of 6-week-old female BALB/c mice were scarified and inoculated with HSV-1 17Syn+ (high phenotypic reactivator) or its mutant 17?Pst(LAT?) (low phenotypic reactivator) at 104 plaque-forming units/eye. NaBu-induced viral reactivation was by intraperitoneal (IP) administration at postinfection (PI) day 28, followed by euthanasia after 1 hour. NaBu-treated, uninfected mice served as the control. The resultant labeled cRNA from TG isolated total RNA was hybridized to gene microarray chips containing 14,000 mouse genes. Quantitative real-time PCR was performed to confirm gene expression. Results. Differential induction of gene expression between 17Syn+ and its mutant 17?Pst(LAT?) was designated as NaBu-induced gene expression and yielded significant upregulation of 2- to 16-fold of 0.4% (56/14,000) host genes probed, comprising mainly nucleosome assembly and binding, central nervous system structural activity, hormonal activity, and signaling activity. Approximately 0.2% (24/14,000) of the host genes, mainly of the same functional categories were downregulated 3- to 11-fold. Immune activity was minor in comparison to our reports on gene expression during latency and heat stress induction. Euchromatin analysis revealed that the LAT-ICP0 locus is amenable to the effects of NaBu. Histone activity was detected by early transcription of histone cluster 2 H2be (Hist2h2be). Conclusions. NaBu-induced reactivation of HSV-1 is twofold: drug action involving significant moderation of specific host epigenetic changes and failure to elicit or suppress immune activity at the early time point of 1 hour. PMID:20881297

  7. Identification of up-regulated genes in flag leaves during rice grain filling and characterization of OsNAC5, a new ABA-dependent transcription factor.

    PubMed

    Sperotto, Raul A; Ricachenevsky, Felipe K; Duarte, Guilherme L; Boff, Tatiana; Lopes, Karina L; Sperb, Edilena R; Grusak, Michael A; Fett, Janette Palma

    2009-10-01

    Rice is a poor source of micronutrients such as iron and zinc. To help clarify the molecular mechanisms that regulate metal mobilization from leaves to developing seeds, we conducted suppression subtractive hybridization analysis in flag leaves of two rice cultivars. Flag leaves are the major source of remobilized metals for developing seeds. We isolated 78 sequences up-regulated in flag leaves at the grain filling stage relative to the panicle exertion stage. Differential expression of selected genes (encoding 7 transport proteins, the OsNAS3 enzyme and the OsNAC5 transcription factor) was confirmed by quantitative RT-PCR. We show that OsNAC5 expression is up-regulated by natural (aging) and induced senescence processes (dark, ABA application, high salinity, cold and Fe-deficiency) and its expression is not affected in the presence of 6-benzylaminopurine (a senescence inhibitor) under dark-induced senescence. Salt induction of OsNAC5 expression is abolished by nicotinamide, an inhibitor of ABA effects. This result and the presence of cis-acting elements in the promoter region of the OsNAC5 gene suggest an ABA-dependent regulation. Using four different rice cultivars, we show that OsNAC5 up-regulation is higher and earlier in flag leaves and panicles of IR75862 plants, which have higher seed concentrations of Fe, Zn and protein. We suggest that OsNAC5 is a novel senescence-associated ABA-dependent NAC transcription factor and its function could be related to Fe, Zn and amino acids remobilization from green tissues to seeds. PMID:19697058

  8. Lack of GCN5 remarkably enhances the resistance against prolonged endoplasmic reticulum stress-induced apoptosis through up-regulation of Bcl-2 gene expression.

    PubMed

    Kikuchi, Hidehiko; Kuribayashi, Futoshi; Mimuro, Hitomi; Imajoh-Ohmi, Shinobu; Nakayama, Masami; Takami, Yasunari; Nishitoh, Hideki; Nakayama, Tatsuo

    2015-08-01

    The endoplasmic reticulum (ER), a complex membrane structure, has important roles in all eukaryotic cells. Catastrophe of its functions would lead to ER stress that causes various diseases such as cancer, neurodegenerative diseases, diabetes and so on. Prolonged ER stress could trigger apoptosis via activation of various signal transduction pathways. To investigate physiological roles of histone acetyltransferase GCN5 in regulation of ER stress, we analyzed responses of homozygous GCN5-deficient DT40 mutants, ?GCN5, against ER stress. GCN5-deficiency in DT40 caused drastic resistance against apoptosis induced by pharmacological ER stress agents (thapsigargin and tunicamycin). Pharmaceutical analysis using specific Bcl-2 inhibitors showed that the drastic resistance against prolonged ER stress-induced apoptosis is, in part, due to up-regulation of Bcl-2 gene expression in ?GCN5. These data revealed that GCN5 is involved in regulation of prolonged ER stress-induced apoptosis through controlling Bcl-2 gene expression. PMID:26086109

  9. Large-scale cellular-resolution gene profiling in human neocortex reveals species-specific molecular signatures

    PubMed Central

    Zeng, Hongkui; Shen, Elaine H.; Hohmann, John G.; Oh, Wook Seung; Bernard, Amy; Royall, Joshua J.; Glattfelder, Katie J.; Sunkin, Susan M.; Morris, John A.; Guillozet-Bongaarts, Angela L.; Smith, Kimberly A.; Ebbert, Amanda J.; Swanson, Beryl; Kuan, Leonard; Page, Damon T.; Overly, Caroline C.; Lein, Ed S.; Hawrylycz, Michael J.; Hof, Patrick R.; Hyde, Thomas M.; Kleinman, Joel E.; Jones, Allan R.

    2012-01-01

    Summary Although there have been major advances in elucidating the functional biology of the human brain, relatively little is known of its cellular and molecular organization. Here we report a large-scale characterization of the expression of ~1,000 genes important for neural functions, by in situ hybridization with cellular resolution in visual and temporal cortices of adult human brains. These data reveal diverse gene expression patterns and remarkable conservation of each individual gene’s expression among individuals (95%), cortical areas (84%), and between human and mouse (79%). A small but substantial number of genes (21%) exhibited species-differential expression. Distinct molecular signatures, comprised of genes both common between species and unique to each, were identified for each major cortical cell type. The data suggest that gene expression profile changes may contribute to differential cortical function across species, in particular, a shift from corticosubcortical to more predominant corticocortical communications in the human brain. PMID:22500809

  10. p53 protein-mediated up-regulation of MAP kinase phosphatase 3 (MKP-3) contributes to the establishment of the cellular senescent phenotype through dephosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2).

    PubMed

    Zhang, Hui; Chi, Yuan; Gao, Kun; Zhang, Xiling; Yao, Jian

    2015-01-01

    Growth arrest is one of the essential features of cellular senescence. At present, the precise mechanisms responsible for the establishment of the senescence-associated arrested phenotype are still incompletely understood. Given that ERK1/2 is one of the major kinases controlling cell growth and proliferation, we examined the possible implication of ERK1/2. Exposure of normal rat epithelial cells to etoposide caused cellular senescence, as manifested by enlarged cell size, a flattened cell body, reduced cell proliferation, enhanced ?-galactosidase activity, and elevated p53 and p21. Senescent cells displayed a blunted response to growth factor-induced cell proliferation, which was preceded by impaired ERK1/2 activation. Further analysis revealed that senescent cells expressed a significantly higher level of mitogen-activated protein phosphatase 3 (MKP-3, a cytosolic ERK1/2-targeted phosphatase), which was suppressed by blocking the transcriptional activity of the tumor suppressor p53 with pifithrin-?. Inhibition of MKP-3 activity with a specific inhibitor or siRNA enhanced basal ERK1/2 phosphorylation and promoted cell proliferation. Apart from its role in growth arrest, impairment of ERK1/2 also contributed to the resistance of senescent cells to oxidant-elicited cell injury. These results therefore indicate that p53-mediated up-regulation of MKP-3 contributes to the establishment of the senescent cellular phenotype through dephosphorylating ERK1/2. Impairment of ERK1/2 activation could be an important mechanism by which p53 controls cellular senescence. PMID:25414256

  11. Modifications of chromatin structure and gene expression following induced alterations of cellular shape.

    PubMed

    Vergani, Laura; Grattarola, Myriam; Nicolini, Claudio

    2004-08-01

    In higher eukaryotes cellular shape is a dynamic element which can be altered by external and internal factors (i.e. surface interactions, temperature, ionic strength). Our question was: might modifications of cell shape reflect on nuclear morphology and architecture and hence on chromatin function, in order to represent a mechanism of cell regulation? We altered the shape of cultured fibroblasts by coating the growth substratum with synthetic polymers, which alternatively increased and decreased the adhesiveness. By means of Fluorescence microscopy we analysed the modifications of cell and nucleus architecture induced by the different substrata. Then we used differential scanning calorimetry to investigate if a remodelling of chromatin structure was associated with the induced morphological changes. Finally, we evaluated if the observed modifications of chromatin condensation affect the transcriptional profile. At this stage of the work we focused on just four genes (c-myc, c-fos, c-jun and collagen) and we analysed their expression by dot blot hybridization and RT-PCR. The results confirm that mechanical factors external to the cell, such as the physico-chemical features of the substratum, are able to modulate gene transcription through a remodelling of chromatin structure. Therefore the work supports our starting hypothesis of a regulatory pathway connecting in sequence cellular morphomety/nuclear architecture/chromatin structure/gene expression. PMID:15147724

  12. Kaposi's Sarcoma-Associated Herpesvirus Viral Interferon Regulatory Factor 4 (vIRF4) Targets Expression of Cellular IRF4 and the Myc Gene To Facilitate Lytic Replication

    PubMed Central

    Do?anay, Sultan; Chung, Brian; Toth, Zsolt; Brulois, Kevin; Lee, Stacy; Kanketayeva, Zhansaya; Feng, Pinghui; Ha, Taekjip

    2014-01-01

    Besides an essential transcriptional factor for B cell development and function, cellular interferon regulatory factor 4 (c-IRF4) directly regulates expression of the c-Myc gene, which is not only associated with various B cell lymphomas but also required for herpesvirus latency and pathogenesis. Kaposi's sarcoma-associated herpesvirus (KSHV), the etiological agent of Kaposi's sarcoma and primary effusion lymphoma, has developed a unique mechanism to deregulate host antiviral innate immunity and growth control by incorporating four viral homologs (vIRF1 to -4) of cellular IRFs into its genome. Previous studies have shown that several KSHV latent proteins, including vIRF3, vFLIP, and LANA, target the expression, function, and stability of c-Myc to establish and maintain viral latency. Here we report that the KSHV vIRF4 lytic protein robustly suppresses expression of c-IRF4 and c-Myc, reshaping host gene expression profiles to facilitate viral lytic replication. Genomewide gene expression analysis revealed that KSHV vIRF4 grossly affects host gene expression by upregulating and downregulating 118 genes and 166 genes, respectively, by at least 2-fold. Remarkably, vIRF4 suppressed c-Myc expression by 11-fold, which was directed primarily by the deregulation of c-IRF4 expression. Real-time quantitative PCR (RT-qPCR), single-molecule in situ hybridization, and chromatin immunoprecipitation assays showed that vIRF4 not only reduces c-IRF4 expression but also competes with c-IRF4 for binding to the specific promoter region of the c-Myc gene, resulting in drastic suppression of c-Myc expression. Consequently, the loss of vIRF4 function in the suppression of c-IRF4 and c-Myc expression ultimately led to a reduction of KSHV lytic replication capacity. These results indicate that the KSHV vIRF4 lytic protein comprehensively targets the expression and function of c-IRF4 to downregulate c-Myc expression, generating a favorable environment for viral lytic replication. Finally, this study further reinforces the important role of the c-Myc gene in KSHV lytic replication and latency. PMID:24335298

  13. Kaposi's sarcoma-associated herpesvirus viral interferon regulatory factor 4 (vIRF4) targets expression of cellular IRF4 and the Myc gene to facilitate lytic replication.

    PubMed

    Lee, Hye-Ra; Do?anay, Sultan; Chung, Brian; Toth, Zsolt; Brulois, Kevin; Lee, Stacy; Kanketayeva, Zhansaya; Feng, Pinghui; Ha, Taekjip; Jung, Jae U

    2014-02-01

    Besides an essential transcriptional factor for B cell development and function, cellular interferon regulatory factor 4 (c-IRF4) directly regulates expression of the c-Myc gene, which is not only associated with various B cell lymphomas but also required for herpesvirus latency and pathogenesis. Kaposi's sarcoma-associated herpesvirus (KSHV), the etiological agent of Kaposi's sarcoma and primary effusion lymphoma, has developed a unique mechanism to deregulate host antiviral innate immunity and growth control by incorporating four viral homologs (vIRF1 to -4) of cellular IRFs into its genome. Previous studies have shown that several KSHV latent proteins, including vIRF3, vFLIP, and LANA, target the expression, function, and stability of c-Myc to establish and maintain viral latency. Here we report that the KSHV vIRF4 lytic protein robustly suppresses expression of c-IRF4 and c-Myc, reshaping host gene expression profiles to facilitate viral lytic replication. Genomewide gene expression analysis revealed that KSHV vIRF4 grossly affects host gene expression by upregulating and downregulating 118 genes and 166 genes, respectively, by at least 2-fold. Remarkably, vIRF4 suppressed c-Myc expression by 11-fold, which was directed primarily by the deregulation of c-IRF4 expression. Real-time quantitative PCR (RT-qPCR), single-molecule in situ hybridization, and chromatin immunoprecipitation assays showed that vIRF4 not only reduces c-IRF4 expression but also competes with c-IRF4 for binding to the specific promoter region of the c-Myc gene, resulting in drastic suppression of c-Myc expression. Consequently, the loss of vIRF4 function in the suppression of c-IRF4 and c-Myc expression ultimately led to a reduction of KSHV lytic replication capacity. These results indicate that the KSHV vIRF4 lytic protein comprehensively targets the expression and function of c-IRF4 to downregulate c-Myc expression, generating a favorable environment for viral lytic replication. Finally, this study further reinforces the important role of the c-Myc gene in KSHV lytic replication and latency. PMID:24335298

  14. 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

  15. Oxidized frying oil up-regulates hepatic acyl-CoA oxidase and cytochrome P450 4 A1 genes in rats and activates PPARalpha.

    PubMed

    Chao, P M; Chao, C Y; Lin, F J; Huang, C

    2001-12-01

    Oxidized LDL (oxLDL) and its component hydroxy fatty acids were shown to activate peroxisome proliferator-activating receptor alpha (PPARalpha) and gamma (PPARgamma). To test the hypothesis that lipid oxidation products in oxidized frying oil (OFO) can activate PPARalpha and up-regulate its target genes, a feeding experiment and a transactivation experiment were conducted. Based on a 2 x 2 factorial design, four groups of Sprague-Dawley male weanling rats were fed diets containing either high (20 g/100 g, HO and HF) or low (5 g/100 g, LO and LF) levels of oxidized frying soybean oil (HO and LO) or fresh soybean oil (HF and LF) for 6 wk. The OFO sample was prepared by frying wheat dough sheets in soybean oil at 205 +/- 5 degrees C for 24 h. OFO dose dependently and significantly increased (P < 0.05) mRNA of acyl-CoA oxidase (ACO) and cytochrome P(450) 4A1(CYP4A1) in liver of rats. Dietary OFO also dose dependently increased liver microsomal CYP4A protein (P < 0.05). The activity of hepatic ACO of the HO group was sixfold that of the HF group (P < 0.05). Plasma total lipids, liver triglycerides, cholesterol and total lipids were reduced in rats fed the LO and HO diets (P < 0.05). Through the ligand binding domain of PPARalpha, the hydrolyzed OFO enhanced the expression of alkaline phosphatase (ALP) reporter gene to a significantly greater extent (P < 0.05) than the hydrolyzed fresh soybean oil in a transactivation assay using a clone of CHO K1 cells stably expressing Gal4-PPARalpha chimeric receptor and UAS4-ALP reporter. The results support our hypothesis that dietary OFO, by activating PPARalpha, up-regulates the expression of PPARalpha downstream genes and alters lipid metabolism in rats. PMID:11739861

  16. The human immunodeficiency virus type-1 Tat protein upregulates Bcl-2 gene expression in Jurkat T-cell lines and primary peripheral blood mononuclear cells.

    PubMed

    Zauli, G; Gibellini, D; Caputo, A; Bassini, A; Negrini, M; Monne, M; Mazzoni, M; Capitani, S

    1995-11-15

    The regulatory Tat protein of human immunodeficiency virus type-1 (HIV-1) exerts a pleyotropic activity on the survival and proliferation of different cell types in culture. In this report, we investigated the effect of either endogenous or exogenous Tat on Bcl-2 proto-oncogene expression and cell survival in Jurkat T-cell lines and primary peripheral blood mononuclear cells. Stable and transient transfections of Jurkat cells with the cDNA of tat and a plasmid containing Bcl-2 promoter in front of CAT (Bcl-2 Pr/CAT) stimulated CAT activity and showed an increase of Bcl-2 mRNA and protein expression. This effect was specifically related to tat, because Jurkat cells transfected with the cDNA of tat in antisense orientation, tat carrying a mutation in the amino acid cys22-gly22, or the control vector alone (pRPneo-SL3) did not show any significant difference in Bcl-2 promoter activity with respect to parental Jurkat cells. We also observed a specific correlation between tat-induced Bcl-2 gene expression and inhibition of apoptosis induced by serum withdrawal. Our results suggest that the structural integrity of the activation domain of Tat was required for the promotion of the Bcl-2 promoter and Jurkat cell survival, because a single mutation in the aminoacid cys22 was sufficient to completely block the upregulation of Bcl-2 and inhibition of apoptosis. Moreover, picomolar concentrations of native or recombinant Tat were able to upregulate Bcl-2 expression both in Jurkat and primary peripheral blood mononuclear cells, suggesting that extracellular Tat, actively released by infected cells, may also play a significant role in suppressing apoptosis. An aberrant cell survival of lymphoid cells consequent to the upregulation of Bcl-2 may represent an additional pathogenetic mechanism that could help explain both the dysregulated immune response and the frequent occurrence of hyperplastic/neoplastic disorders in HIV-1-seropositive individuals. PMID:7579350

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

    PubMed Central

    Drew, Janice E.

    2012-01-01

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

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

    SciTech Connect

    Wakoh, Takeshi; Uekawa, Natsuko [Department of Mechanism of Aging, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, 36-3, Gengo, Morioka-Cho, Obu-City, Aichi 474-8522 (Japan); Terauchi, Kunihiko [Department of Cardiovascular and Thoracic Surgery, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, 465 Kajii-cho, Kawaramachi-Hirokoji, Kamigyo-ku, Kyoto 602-8566 (Japan); Sugimoto, Masataka [Department of Mechanism of Aging, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, 36-3, Gengo, Morioka-Cho, Obu-City, Aichi 474-8522 (Japan); Ishigami, Akihito [Department of Biochemistry, Faculty of Pharmaceutical Sciences, Toho University, Miyama 2-2-1, Funabashi, Chiba 274-8510 (Japan); Shimada, Jun-ichi [Department of Cardiovascular and Thoracic Surgery, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, 465 Kajii-cho, Kawaramachi-Hirokoji, Kamigyo-ku, Kyoto 602-8566 (Japan); Maruyama, Mitsuo [Department of Mechanism of Aging, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, 36-3, Gengo, Morioka-Cho, Obu-City, Aichi 474-8522 (Japan)], E-mail: michan@nils.go.jp

    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.

  19. UP-REGULATION OF A NOVEL WHEAT LECTIN-LIKE GENE IS ASSOCIATED WITH RESISTANCE TO HESSIAN FLY LARVAE

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The incompatible interaction of wheat (Triticum aestivum) plants with Hessian fly (Mayetiola destructor) results in larval death when avirulent larvae attempt to establish a feeding site at the base of wheat seedlings. The resistance response is governed by a gene-for-gene recognition event, which o...

  20. Neuronal excitation upregulates Tbr1, a high-confidence risk gene of autism, mediating Grin2b expression in the adult brain

    PubMed Central

    Chuang, Hsiu-Chun; Huang, Tzyy-Nan; Hsueh, Yi-Ping

    2014-01-01

    The activity-regulated gene expression of transcription factors is required for neural plasticity and function in response to neuronal stimulation. T-brain-1 (TBR1), a critical neuron-specific transcription factor for forebrain development, has been recognized as a high-confidence risk gene for autism spectrum disorders. Here, we show that in addition to its role in brain development, Tbr1 responds to neuronal activation and further modulates the Grin2b expression in adult brains and mature neurons. The expression levels of Tbr1 were investigated using both immunostaining and quantitative reverse transcription polymerase chain reaction (RT-PCR) analyses. We found that the mRNA and protein expression levels of Tbr1 are induced by excitatory synaptic transmission driven by bicuculline or glutamate treatment in cultured mature neurons. The upregulation of Tbr1 expression requires the activation of both ?-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) and N-methyl-D-aspartate (NMDA) receptors. Furthermore, behavioral training triggers Tbr1 induction in the adult mouse brain. The elevation of Tbr1 expression is associated with Grin2b upregulation in both mature neurons and adult brains. Using Tbr1-deficient neurons, we further demonstrated that TBR1 is required for the induction of Grin2b upon neuronal activation. Taken together with the previous studies showing that TBR1 binds the Grin2b promoter and controls expression of luciferase reporter driven by Grin2b promoter, the evidence suggests that TBR1 directly controls Grin2b expression in mature neurons. We also found that the addition of the calcium/calmodulin-dependent protein kinase II (CaMKII) antagonist KN-93, but not the calcium-dependent phosphatase calcineurin antagonist cyclosporin A, to cultured mature neurons noticeably inhibited Tbr1 induction, indicating that neuronal activation upregulates Tbr1 expression in a CaMKII-dependent manner. In conclusion, our study suggests that Tbr1 plays an important role in adult mouse brains in response to neuronal activation to modulate the activity-regulated gene transcription required for neural plasticity. PMID:25309323

  1. A functional screen for copper homeostasis genes identifies a pharmacologically tractable cellular system

    PubMed Central

    2014-01-01

    Background Copper is essential for the survival of aerobic organisms. If copper is not properly regulated in the body however, it can be extremely cytotoxic and genetic mutations that compromise copper homeostasis result in severe clinical phenotypes. Understanding how cells maintain optimal copper levels is therefore highly relevant to human health. Results We found that addition of copper (Cu) to culture medium leads to increased respiratory growth of yeast, a phenotype which we then systematically and quantitatively measured in 5050 homozygous diploid deletion strains. Cu’s positive effect on respiratory growth was quantitatively reduced in deletion strains representing 73 different genes, the function of which identify increased iron uptake as a cause of the increase in growth rate. Conversely, these effects were enhanced in strains representing 93 genes. Many of these strains exhibited respiratory defects that were specifically rescued by supplementing the growth medium with Cu. Among the genes identified are known and direct regulators of copper homeostasis, genes required to maintain low vacuolar pH, and genes where evidence supporting a functional link with Cu has been heretofore lacking. Roughly half of the genes are conserved in man, and several of these are associated with Mendelian disorders, including the Cu-imbalance syndromes Menkes and Wilson’s disease. We additionally demonstrate that pharmacological agents, including the approved drug disulfiram, can rescue Cu-deficiencies of both environmental and genetic origin. Conclusions A functional screen in yeast has expanded the list of genes required for Cu-dependent fitness, revealing a complex cellular system with implications for human health. Respiratory fitness defects arising from perturbations in this system can be corrected with pharmacological agents that increase intracellular copper concentrations. PMID:24708151

  2. env Gene of chicken RNA tumor viruses: extent of conservation in cellular and viral genomes.

    PubMed

    Fujita, D J; Tal, J; Varmus, H E; Bishop, J M

    1978-09-01

    The env gene of avian sarcoma-leukosis viruses codes for envelope glycoproteins that determine viral host range, antigenic specificity, and interference patterns. We used molecular hybridization to analyze the natural distribution and possible origins of the nucleotide sequences that encode env; our work exploited the availability of radioactive DNA (cDNA(gp)) complementary to most or all of env. env sequences were detectable in the DNAs of chickens which synthesized an env gene product (chick helper factor positive) encoded by an endogenous viral gene and also in the DNAs of chickens which synthesized little or no env gene product (chick helper factor negative). env sequences were not detectable in DNAs from Japanese quail, ring-necked pheasant, golden pheasant, duck, squab, salmon sperm, or calf thymus. The detection of sequences closely related to viral env only in chicken DNA contrasts sharply with the demonstration that the transforming gene (src) of avian sarcoma viruses has readily detectable homologues in the DNAs of all avian species tested [D. Stehelin, H. E. Varmus, J. M. Bishop, and P. K. Vogt, Nature (London) 260: 170-173, 1976] and in the DNAs of other vertebrates (D. Spector, personal communication). Thermal denaturation studies on duplexes formed between cDNA(gp) and chicken DNA and also between cDNA(gp) and RNAs of subgroup A to E viruses derived from chickens indicated that these duplexes were well matched. In contrast, cDNA(gp) did not form stable hybrids with RNAs of viruses which were isolated from ring-necked and golden pheasants. We conclude that substantial portions of nucleotide sequences within the env genes of viruses of subgroups A to E are closely related and that these genes probably have a common, perhaps cellular, evolutionary origin. PMID:212576

  3. Genome-wide transcriptomic analysis of cotton under drought stress reveal significant down-regulation of genes and pathways involved in fibre elongation and up-regulation of defense responsive genes.

    PubMed

    Padmalatha, Kethireddy Venkata; Dhandapani, Gurusamy; Kanakachari, Mogilicherla; Kumar, Saravanan; Dass, Abhishek; Patil, Deepak Prabhakar; Rajamani, Vijayalakshmi; Kumar, Krishan; Pathak, Ranjana; Rawat, Bhupendra; Leelavathi, Sadhu; Reddy, Palakolanu Sudhakar; Jain, Neha; Powar, Kasu N; Hiremath, Vamadevaiah; Katageri, Ishwarappa S; Reddy, Malireddy K; Solanke, Amolkumar U; Reddy, Vanga Siva; Kumar, Polumetla Ananda

    2012-02-01

    Cotton is an important source of natural fibre used in the textile industry and the productivity of the crop is adversely affected by drought stress. High throughput transcriptomic analyses were used to identify genes involved in fibre development. However, not much information is available on cotton genome response in developing fibres under drought stress. In the present study a genome wide transcriptome analysis was carried out to identify differentially expressed genes at various stages of fibre growth under drought stress. Our study identified a number of genes differentially expressed during fibre elongation as compared to other stages. High level up-regulation of genes encoding for enzymes involved in pectin modification and cytoskeleton proteins was observed at fibre initiation stage. While a large number of genes encoding transcription factors (AP2-EREBP, WRKY, NAC and C2H2), osmoprotectants, ion transporters and heat shock proteins and pathways involved in hormone (ABA, ethylene and JA) biosynthesis and signal transduction were up-regulated and genes involved in phenylpropanoid and flavonoid biosynthesis, pentose and glucuronate interconversions and starch and sucrose metabolism pathways were down-regulated during fibre elongation. This study showed that drought has relatively less impact on fibre initiation but has profound effect on fibre elongation by down-regulating important genes involved in cell wall loosening and expansion process. The comprehensive transcriptome analysis under drought stress has provided valuable information on differentially expressed genes and pathways during fibre development that will be useful in developing drought tolerant cotton cultivars without compromising fibre quality. PMID:22143977

  4. Clock gene mutation modulates the cellular sensitivity to genotoxic stress through altering the expression of N-methylpurine DNA glycosylase gene

    Microsoft Academic Search

    Jahye Kim; Naoya Matsunaga; Satoru Koyanagi; Shigehiro Ohdo

    2009-01-01

    Although Clock gene product, a component of the circadian pacemaker, has been suggested to participate in the regulation of cellular sensitivity to genotoxic stress, the underlying mechanism remains to be fully understood. In this study, we showed that Clock gene mutation modulates the sensitivity of hepatocytes to alkylating agent-induced genotoxic stress through altering the expression of N-methylpurine DNA glycosylase (MPG),

  5. Modified pectin-based carrier for gene delivery: Cellular barriers in gene delivery course

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The use of biodegradable and biocompatible polysaccharides as DNA carriers has high potential for gene therapy applications. Pectin is a structural plant polysaccharide heterogeneous with respect to its chemical structure. It contains branches rich in galactose residues which serve as potential liga...

  6. Echinacea purpurea up-regulates CYP1A2, CYP3A4 and MDR1 gene expression by activation of pregnane X receptor pathway.

    PubMed

    Awortwe, Charles; Manda, Vamshi K; Avonto, Cristina; Khan, Shabana I; Khan, Ikhlas A; Walker, Larry A; Bouic, Patrick J; Rosenkranz, Bernd

    2015-03-01

    1.This study investigated the mechanism underlying Echinacea-mediated induction of CYP1A2, CYP3A4 and MDR1 in terms of human pregnane X receptor (PXR) activation. 2.Crude extracts and fractions of Echinacea purpurea were tested for PXR activation in HepG2 cells by a reporter gene assay. Quantitative real-time PCR was carried out to determine their effects on CYP1A2 and CYP3A4 mRNA expressions. Capsules and fractions were risk ranked as high, intermediate and remote risk of drug-metabolizing enzymes induction based on EC50 values determined for respective CYPs. 3.?Fractions F1, F2 and capsule (2660) strongly activated PXR with 5-, 4- and 3.5-fold increase in activity, respectively. Echinacea preparations potentiated up-regulation of CYP1A2, CYP3A4 and MDR1 via PXR activation. 4.Thus E. purpurea preparations cause herb-drug interaction by up-regulating CYP1A2, CYP3A4 and P-gp via PXR activation. PMID:25377539

  7. Helicobacter pylori cytotoxin-associated gene A protein upregulates ?-enolase expression via Src/MEK/ERK pathway: implication for progression of gastric cancer.

    PubMed

    Chen, Shuaiyin; Duan, Guangcai; Zhang, Rongguang; Fan, Qingtang

    2014-08-01

    Persistent infection with Helicobacter pylori confers an increased risk for the development of gastric cancer. In our previous investigations, we found that ENO1 was overexpression in cagA-positive H. pylori-infected gastric epithelial AGS cells by proteomic method, in contrast to the isogenic cagA knock out mutant H. pylori-infected cells. ENO1 is a newly identified oncoprotein overexpressed in some cancer. However, the relationship between H. pylori infection and ENO1 expression still remains undefined. The AGS gastric cancer cells were transfected with WT-cagA plasmid and PR-cagA plasmids. Expression of ENO1 mRNA and protein were measured by real-time quantitative PCR and western blot analysis. Signal protein inhibitor treatment was used to investigate the signal pathways. It was found that the ENO1 mRNA and protein overexpression levels were dependent on cagA gene expression and CagA protein phosphorylation. Further analysis revealed that the Src, MEK and ERK pathway was involved in this upregulation effect. Our data suggest that ENO1 was upregulated by CagA protein through activating the Src and MEK/ERK signal pathways, thereby providing a novel mechanism underlying H. pylori-mediated gastric diseases. PMID:24841372

  8. Gene Expression Analysis of the 26S Proteasome Subunit PSMB4 Reveals Significant Upregulation, Different Expression and Association with Proliferation in Human Pulmonary Neuroendocrine Tumours

    PubMed Central

    Mairinger, Fabian Dominik; Walter, Robert Fred Henry; Theegarten, Dirk; Hager, Thomas; Vollbrecht, Claudia; Christoph, Daniel Christian; Worm, Karl; Ting, Saskia; Werner, Robert; Stamatis, Georgios; Mairinger, Thomas; Baba, Hideo; Zarogoulidis, Konstantinos; Huang, Haidong; Li, Qiang; Tsakiridis, Kosmas; Zarogoulidis, Paul; Schmid, Kurt Werner; Wohlschlaeger, Jeremias

    2014-01-01

    Background: Proteasomal subunit PSMB4 was suggested to be a survival gene in an animal model of hepatocellular carcinoma and in glioblastoma cell lines. In pulmonary adenocarcinoma, a high expression of these genes was found to be associated with poor differentiation and survival. This study investigates the gene expression levels of 26S proteasome subunits in human pulmonary neuroendocrine tumours including typical (TC) and atypical (AC) carcinoid tumours as well as small cell (SCLC) and large cell (LCNEC) neuroendocrine carcinomas. Material and methods: Gene expression levels of proteasomal subunits (PSMA1, PSMA5, PSMB4, PSMB5 and PSMD1) were investigated in 80 neuroendocrine pulmonary tumours (each 20 TC, AC, LCNLC and SCLC) and compared to controls. mRNA levels were determined by using TaqMan assays. Immunohistochemistry on tissue microarrays (TMA) was performed to determine the expression of ki67, cleaved caspase 3 and PSMB4. Results: All proteasomal subunit gene expressions were significantly upregulated in TC, AC, SCLC and LCNEC compared to controls. PSMB4 mRNA is differently expressed between all neuroendocrine tumour subtypes demonstrating the highest expression and greatest range in LCNEC (p=0.043), and is significantly associated with proliferative activity (p=0.039). Conclusion: In line with other 26S proteasomal subunits PSMB4 is significantly increased, but differently expressed between pulmonary neuroendocrine tumours and is associated with the proliferative activity. Unlike in pulmonary adenocarcinomas, no association with biological behaviour was observed, suggesting that increased proteasomal subunit gene expression is a common and probably early event in the tumorigenesis of pulmonary neuroendocrine tumours regardless of their differentiation. PMID:25157275

  9. Selective up-regulation of LXR-regulated genes ABCA1, ABCG1, and APOE in macrophages through increased endogenous synthesis of 24(S),25-epoxycholesterol.

    PubMed

    Beyea, Michael M; Heslop, Claire L; Sawyez, Cynthia G; Edwards, Jane Y; Markle, Janet G; Hegele, Robert A; Huff, Murray W

    2007-02-23

    Liver X receptor (LXR) activation represents a mechanism to prevent macrophage foam cell formation. Previously, we demonstrated that partial inhibition of oxidosqualene:lanosterol cyclase (OSC) stimulated synthesis of the LXR agonist 24(S),25-epoxycholesterol (24(S),25-epoxy) and enhanced ABCA1-mediated cholesterol efflux. In contrast to a synthetic, nonsteroidal LXR activator, TO-901317, triglyceride accumulation was not observed. In the present study, we determined whether endogenous 24(S),25-epoxy synthesis selectively enhanced expression of macrophage LXR-regulated cholesterol efflux genes but not genes that regulate fatty acid metabolism. THP-1 human macrophages incubated with the OSC inhibitor (OSCi) RO0714565 (15 nM) significantly reduced cholesterol synthesis and maximized synthesis of 24(S),25-epoxy. Endogenous 24(S),25-epoxy increased ABCA1, ABCG1, and APOE mRNA abundance and consequently increased cholesterol efflux to apoAI. In contrast, OSCi had no effect on LXR-regulated genes LPL (lipoprotein lipase) and FAS (fatty acid synthase). TO-901317 (>or=10 nM) significantly enhanced expression of all genes examined. OSCi and TO-901317 increased the mRNA and precursor form of SREBP-1c, a major regulator of fatty acid and triglyceride synthesis. However, conversion of the precursor to the active form (nSREBP-1c) was blocked by OSCi-induced 24(S),25-epoxy but not by TO-901317 (>or=10 nm), which instead markedly increased nSREBP-1c. Disruption of nSREBP-1c formation by 24(S),25-epoxy accounted for diminished FAS and LPL expression. In summary, endogenous synthesis of 24(S),25-epoxy selectively up-regulates expression of macrophage LXR-regulated cholesterol efflux genes without stimulating genes linked to fatty acid and triglyceride synthesis. PMID:17186944

  10. A cellular genetics approach identifies gene-drug interactions and pinpoints drug toxicity pathway nodes.

    PubMed

    Suzuki, Oscar T; Frick, Amber; Parks, Bethany B; Trask, O Joseph; Butz, Natasha; Steffy, Brian; Chan, Emmanuel; Scoville, David K; Healy, Eric; Benton, Cristina; McQuaid, Patricia E; Thomas, Russell S; Wiltshire, Tim

    2014-01-01

    New approaches to toxicity testing have incorporated high-throughput screening across a broad-range of in vitro assays to identify potential key events in response to chemical or drug treatment. To date, these approaches have primarily utilized repurposed drug discovery assays. In this study, we describe an approach that combines in vitro screening with genetic approaches for the experimental identification of genes and pathways involved in chemical or drug toxicity. Primary embryonic fibroblasts isolated from 32 genetically-characterized inbred mouse strains were treated in concentration-response format with 65 compounds, including pharmaceutical drugs, environmental chemicals, and compounds with known modes-of-action. Integrated cellular responses were measured at 24 and 72 h using high-content imaging and included cell loss, membrane permeability, mitochondrial function, and apoptosis. Genetic association analysis of cross-strain differences in the cellular responses resulted in a collection of candidate loci potentially underlying the variable strain response to each chemical. As a demonstration of the approach, one candidate gene involved in rotenone sensitivity, Cybb, was experimentally validated in vitro and in vivo. Pathway analysis on the combined list of candidate loci across all chemicals identified a number of over-connected nodes that may serve as core regulatory points in toxicity pathways. PMID:25221565

  11. PPAR{alpha} gene expression is up-regulated by LXR and PXR activators in the small intestine

    SciTech Connect

    Inoue, Jun [Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo, Tokyo 113-8657 (Japan); Basic Research Activities for Innovative Biosciences (BRAIN) (Japan); Satoh, Shin-ichi; Kita, Mariko [Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo, Tokyo 113-8657 (Japan); Nakahara, Mayuko [Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo, Tokyo 113-8657 (Japan); Basic Research Activities for Innovative Biosciences (BRAIN) (Japan); Hachimura, Satoshi [Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo, Tokyo 113-8657 (Japan); Miyata, Masaaki [Division of Drug Metabolism and Molecular Toxicology, Graduate School of Pharmaceutical Sciences, Tohoku University (Japan); Nishimaki-Mogami, Tomoko [Division of Cell Signaling, National Institute of Health Sciences (Japan); Sato, Ryuichiro [Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo, Tokyo 113-8657 (Japan); Basic Research Activities for Innovative Biosciences (BRAIN) (Japan)], E-mail: aroysato@mail.ecc.u-tokyo.ac.jp

    2008-07-11

    LXR, PXR, and PPAR{alpha} are members of a nuclear receptor family which regulate the expression of genes involved in lipid metabolism. Here, we show the administration of T0901317 stimulates PPAR{alpha} gene expression in the small intestine but not in the liver of both normal and FXR-null mice. The administration of LXR specific ligand GW3965, or PXR specific ligand PCN has the same effect, indicating that ligand-dependent activation of LXR and PXR, but not FXR, is responsible for the increased gene expression of PPAR{alpha} in the mouse small intestine.

  12. Gene expression is dynamically regulated in retinal progenitor cells prior to and during overt cellular differentiation.

    PubMed

    Dixit, Rajiv; Tachibana, Nobuhiko; Touahri, Yacine; Zinyk, Dawn; Logan, Cairine; Schuurmans, Carol

    2014-01-01

    The retina is comprised of one glial and six neuronal populations that are generated from a multipotent pool of retinal progenitor cells (RPCs) during development. To give rise to these different cell types, RPCs undergo temporal identity transitions, displaying distinct gene expression profiles at different stages of differentiation. Little, however, is known about temporal differences in RPC identities prior to the onset of overt cellular differentiation, during the period when a retinal identity is gradually acquired. Here we examined the sequential onset of expression of regional markers (i.e., homeodomain transcription factors) and cell fate determinants (i.e., basic-helix-loop-helix transcription factors and neurogenic genes) in RPCs from the earliest appearance of a morphologically-distinct retina. By performing a comparative analysis of the expression of a panel of 27 homeodomain, basic-helix-loop-helix and Notch pathway genes between embryonic day (E) 8.75 and postnatal day (P) 9, we identified six distinct RPC molecular profiles. At E8.75, the earliest stage assayed, murine RPCs expressed five homeodomain genes and a single neurogenic gene (Pax6, Six3, Six6, Rx, Otx2, Hes1). This early gene expression profile was remarkably similar to that of 'early' RPCs in the amphibian ciliary marginal zone (CMZ), where RPCs are compartmentalised according to developmental stage, and homologs of Pax6, Six3 and Rx are expressed in the 'early' stem cell zone. As development proceeds, expression of additional homeodomain, bHLH and neurogenic genes was gradually initiated in murine RPCs, allowing distinct genetic profiles to also be defined at E9.5, E10.5, E12.5, E15.5 and P0. In addition, RPCs in the postnatal ciliary margin, where retinal stem cells are retained throughout life, displayed a unique molecular signature, expressing all of the early-onset genes as well as several late-onset markers, indicative of a 'mixed' temporal identity. Taken together, the identification of temporal differences in gene expression in mammalian RPCs during pre-neurogenic developmental stages leads to new insights into how regional identities are progressively acquired during development, while comparisons at later stages highlight the dynamic nature of gene expression in temporally distinct RPC pools. PMID:24148613

  13. Evaluating thermodynamic models of enhancer activity on cellular resolution gene expression data.

    PubMed

    Samee, Abul Hassan; Sinha, Saurabh

    2013-07-15

    With the advent of high throughput sequencing and high resolution transcriptomic technologies, there exists today an unprecedented opportunity to understand gene regulation at a quantitative level. State of the art models of the relationship between regulatory sequence and gene expression have shown great promise, but also suffer from some major shortcomings. In this paper, we identify and address methodological challenges pertaining to quantitative modeling of gene expression from sequence, and test our models on the anterior-posterior patterning system in the Drosophila embryo. We first develop a framework to process cellular resolution three-dimensional gene expression data from the Drosophila embryo and create data sets on which quantitative models can be trained. Next we propose a new score, called 'weighted pattern generating potential' (w-PGP), to evaluate model predictions, and show its advantages over the two most common scoring schemes in use today. The model building exercise uses w-PGP as the evaluation score and adopts a systematic strategy to increase a model's complexity while guarding against over-fitting. Our model identifies three transcription factors--ZELDA, SLOPPY-PAIRED, and NUBBIN--that have not been previously incorporated in quantitative models of this system, as having significant regulatory influence. Finally, we show how fitting quantitative models on data sets comprising a handful of enhancers, as reported in earlier work, may lead to unreliable models. PMID:23624421

  14. Anti-HIV agent MAP30 modulates the expression profile of viral and cellular genes for proliferation and apoptosis in AIDS-related lymphoma cells infected with Kaposi's sarcoma-associated virus.

    PubMed

    Sun, Y; Huang, P L; Li, J J; Huang, Y Q; Zhang, L; Huang, P L; Lee-Huang, S

    2001-10-01

    The anti-HIV agent MAP30 (Momordica anti-HIV protein, 30 kDa) inhibits the proliferation of BC-2, an AIDS-related primary effusion lymphoma (PEL) cell line derived from an AIDS patient. BC-2 cells are latently infected with Kaposi's sarcoma-associated herpes virus (KSHV), also known as human herpes virus 8 (HHV8). We examined the effect of MAP30 on the expression of viral and cellular genes in BC-2 during latent and lytic states of the viral life cycle. By Northern analysis and RT-PCR, we found that MAP30 downregulates the expression of viral cyclin D (vCD), viral interleukin-6 (vIL-6), and viral FLIP (vFLIP), genes involved in cell cycle regulation, viral pathogenesis, and apoptosis. By pathway-specific cDNA microarray analysis, we found that BC-2 cells express high levels of egr-1, ATF-2, hsp27, hsp90, IkappaB, mdm2, skp1, and IL-2, cellular genes involved in mitogenesis, tumorigenesis, and inhibition of apoptosis in NFkappaB and p53 signaling pathways. These results define for the first time the specific cellular pathways involved in AIDS-related tumorigenesis and suggest specific novel targets for the treatment. Furthermore, we found that MAP30 downregulates the expression of egr-1, ATF-2, hsp27, hsp90, IkappaB, mdm2, and Skp1, while it upregulates the pro-apoptotic-related genes Bax, CRADD, and caspase-3. Thus, MAP30 modulates the expression of both viral and cellular genes involved in KS pathogenesis. These results provide valuable insight into the molecular mechanisms of MAP30 anti-KS action and suggest its utility as a therapeutic agent against AIDS-related tumors. PMID:11573962

  15. Solenopsis invicta transferrin: cDNA cloning, gene architecture, and up-regulation in response to Beauveria bassiana infection.

    PubMed

    Valles, Steven M; Pereira, Roberto M

    2005-09-26

    Transferrin genes from several insects have been shown to be induced in response to bacterial or fungal infection. We were interested to know whether transferrin genes in the red imported fire ant, Solenopsis invicta, are similarly induced by microbial challenge. Hence, the cDNA and structure of a gene exhibiting significant homology to insect transferrins were elucidated for S. invicta. The cDNA was comprised of 2417 nucleotides, excluding the poly(A) tail, with a large open reading frame of 2106 nucleotides. The predicted translation product of the S. invicta tranferrin (SiTf) gene was a 702 amino acid polypeptide with an estimated molecular mass of 77.3 kDa and a pI value of 5.66, characteristics consistent with transferrin proteins. Comparative analysis of genomic and cDNA sequences revealed that the SiTf gene was comprised of 8 exons. Quantitative real-time PCR was used to examine the expression of SiTf. Expression of SiTf was induced in worker ants exposed to Beauveria bassiana conidia. Autoclave-killed conidia did not elicit a SiTf induction response from worker ants. Genes, like SiTf, responding to microbe attack or infection may provide a unique approach to assist in the discovery of microbial control organisms for the target insect pest. PMID:16039806

  16. Quercetin glycosides induced neuroprotection by changes in the gene expression in a cellular model of Parkinson's disease.

    PubMed

    Magalingam, Kasthuri Bai; Radhakrishnan, Ammu; Ramdas, Premdass; Haleagrahara, Nagaraja

    2015-03-01

    Quercetin glycosides, rutin and isoquercitrin, are potent antioxidants that have been found to possess neuroprotective effect in diseases like Parkinson's and Alzheimer's disease. In the present study, we have examined the gene expression changes with rutin and isoquercitrin pretreatment on 6-hydroxydopamine (6-OHDA)-treated toxicity in rat pheochromocytoma (PC12) cells. PC12 cells were pretreated with rutin or isoquercitrin and subsequently exposed to 6-OHDA. Rutin-pretreated PC12 attenuated the Park2, Park5, Park7, Casp3, and Casp7 genes which were expressed significantly in the 6-OHDA-treated PC12 cells. Rutin upregulated the TH gene which is important in dopamine biosynthesis, but isoquercitrin pretreatment did not affect the expression of this gene. Both rutin and isoquercitrin pretreatments upregulated the ion transport and antiapoptotic genes (NSF and Opa1). The qPCR array data were further validated by qRT-PCR using four primers, Park5, Park7, Casp3, and TH. This finding suggests that changes in the expression levels of transcripts encoded by genes that participate in ubiquitin pathway and dopamine biosynthesis may be involved in Parkinson's disease. PMID:25129099

  17. Upregulation of CRMP4, a new prostate cancer metastasis suppressor gene, inhibits tumor growth in a nude mouse intratibial injection model.

    PubMed

    Zhou, Wei; Xie, Peigen; Pang, Mao; Yang, Bu; Fang, Youqiang; Shu, Tao; Liu, Chang; Wang, Xuan; Zhang, Liangming; Li, Shangfu; Rong, Limin

    2015-01-01

    Prostate cancer, the most commonly diagnosed male cancer in North America, has a high incidence of bone metastasis. Our previous study showed collapsin response mediator protein 4 (CRMP4) gene inhibited prostate cancer migration and invasion. In this study, we investigated whether overexpression of CRMP4 gene in prostate cancer cells inhibit tumor bone metastasis. The stable prostate cancer cells overexpressing the CRMP4 gene were constructed using lentivirus infection. Prostate cancer bone metastasis nude mouse model was built though orthotopic prostate implantation, intracardiac injection and intratibial injection with CRMP4 overexpress and control cancer cells. Small animal PET/CT scanning results showed no difference of bone metastatic capacity in orthotopic and intracardiac injection models between CRMP4 overexpression and control group, while CRMP4 overexpression inhibited tumor growth in the intratibial injection model. Moreover, our in vitro study showed CRMP4 overexpression downregulates the Neuropilin1 (NRP1) expression and upregulate the Noggin expression. Immunohistochemical staining of the hind limbs of intratibial injection model was confirmed with cytological experiments. Taken together, our research indicated CRMP4 inhibits prostate cancer cells growth in the nude mouse bone microenvironment and this effect may relate with regulation of NRP1 and Noggin expression. PMID:25338524

  18. Enhanced human bone marrow stromal cell affinity for modified poly(L-lactide) surfaces by the upregulation of adhesion molecular genes.

    PubMed

    Mao, Xueli; Peng, Hui; Ling, Junqi; Friis, Thor; Whittaker, Andrew K; Crawford, Ross; Xiao, Yin

    2009-12-01

    To enhance and regulate cell affinity for poly (L-lactic acid) (PLLA) based materials, two hydrophilic ligands, poly (ethylene glycol) (PEG) and poly (L-lysine) (PLL), were used to develop triblock copolymers: methoxy-terminated poly (ethylene glycol)-block-poly (L-lactide)-block-poly (L-lysine) (MPEG-b-PLLA-b-PLL) in order to regulate protein absorption and cell adhesion. Bone marrow stromal cells (BMSCs) were cultured on different composition of MPEG-b-PLLA-b-PLL copolymer films to determine the effect of modified polymer surfaces on BMSC attachment. To understand the molecular mechanism governing the initial cell adhesion on difference polymer surfaces, the mRNA expression of 84 human extracellular matrix (ECM) and adhesion molecules was analysed using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). It was found that down regulation of adhesion molecules was responsible for the impaired BMSC attachment on PLLA surface. MPEG-b-PLLA-b-PLL copolymer films improved significantly the cell adhesion and cytoskeleton expression by upregulation of relevant molecule genes significantly. Six adhesion genes (CDH1, ITGL, NCAM1, SGCE, COL16A1, and LAMA3) were most significantly influenced by the modified PLLA surfaces. In summary, polymer surfaces altered adhesion molecule gene expression of BMSCs, which consequently regulated cell initial attachment on modified PLLA surfaces. PMID:19796804

  19. Characterization of a Gene Encoding Clathrin Heavy Chain in Maize Up-Regulated by Salicylic Acid, Abscisic Acid and High Boron Supply

    PubMed Central

    Zeng, Mu-Heng; Liu, Sheng-Hong; Yang, Miao-Xian; Zhang, Ya-Jun; Liang, Jia-Yong; Wan, Xiao-Rong; Liang, Hong

    2013-01-01

    Clathrin, a three-legged triskelion composed of three clathrin heavy chains (CHCs) and three light chains (CLCs), plays a critical role in clathrin-mediated endocytosis (CME) in eukaryotic cells. In this study, the genes ZmCHC1 and ZmCHC2 encoding clathrin heavy chain in maize were cloned and characterized for the first time in monocots. ZmCHC1 encodes a 1693-amino acid-protein including 29 exons and 28 introns, and ZmCHC2 encodes a 1746-amino acid-protein including 28 exons and 27 introns. The high similarities of gene structure, protein sequences and 3D models among ZmCHC1, and Arabidopsis AtCHC1 and AtCHC2 suggest their similar functions in CME. ZmCHC1 gene is predominantly expressed in maize roots instead of ubiquitous expression of ZmCHC2. Consistent with a typical predicted salicylic acid (SA)-responsive element and four predicted ABA-responsive elements (ABREs) in the promoter sequence of ZmCHC1, the expression of ZmCHC1 instead of ZmCHC2 in maize roots is significantly up-regulated by SA or ABA, suggesting that ZmCHC1 gene may be involved in the SA signaling pathway in maize defense responses. The expressions of ZmCHC1 and ZmCHC2 genes in maize are down-regulated by azide or cold treatment, further revealing the energy requirement of CME and suggesting that CME in plants is sensitive to low temperatures. PMID:23880865

  20. Early Life Manganese Exposure Upregulates Tumor-Associated Genes in the Hypothalamus of Female Rats: Relationship to Manganese-Induced Precocious Puberty

    PubMed Central

    Dees, William L.

    2013-01-01

    Prepubertal exposure to low, but elevated levels of manganese (Mn) can induce increased secretions of puberty-related hormones resulting in precocious pubertal development in female rats. These events are due to an action of the element within the hypothalamus to induce the secretion of gonadotropin-releasing hormone (GnRH). Because of these prepubertal effects of Mn and because precocious puberty is a serious neuroendocrine disorder, we have assessed whether early life exposure to this environmental element is capable of precociously upregulating the expression of a select group of genes previously associated with tumor growth or suppression, and that have more recently been shown to increase at the normal time of puberty. Female rat pups received a daily dose of either 10mg/kg manganese(II) chloride or an equal volume of saline by gastric gavage from postnatal day 12 through day 22 or 29. At this time, blood was collected for estradiol analysis and hypothalamic brain tissue frozen on dry ice until assessed for gene expressions. Rats exposed to the elevated levels of Mn showed a precocious increase in GnRH gene expression in the preoptic area and rostral hypothalamus on day 29, an action associated with precociously increased expressions of specific tumor-associated, puberty-related genes. These results demonstrate for the first time that prepubertal Mn exposure is capable of activating specific upstream genes regulating hypothalamic GnRH and suggest that these actions are involved in the mechanism by which this element can induce precocious puberty. PMID:23997110

  1. Insulin and insulin-like growth factor I up-regulate GLUT4 gene expression in fetal brown adipocytes, in a phosphoinositide 3-kinase-dependent manner.

    PubMed Central

    Valverde, A M; Navarro, P; Teruel, T; Conejo, R; Benito, M; Lorenzo, M

    1999-01-01

    Fetal brown adipocytes cultured in a serum-free medium, containing 5 mM glucose, expressed both GLUT4 and GLUT1 glucose transporters at the mRNA and protein level. Treatment with either insulin or insulin-like growth factor (IGF)-I at physiological concentrations up-regulates the expression of the GLUT4 gene, producing a time-dependent mRNA accumulation (7-fold increase at 24 h) and a 2.5-fold increase in the amount of protein in the total membrane fraction. However, insulin treatment down-regulates GLUT1 mRNA and protein expression. Moreover, either insulin or IGF-I transactivates a full-promoter GLUT4-chloramphenicol acetyltransferase gene (CAT) construct transiently transfected to the cells, without affecting GLUT1-CAT activity. In consequence, insulin treatment for 24 h increased by 3-fold the basal glucose uptake. Inhibition of phosphoinositide (PI) 3-kinase activity with chemical agents such as wortmannin or LY294002 partially blocked insulin-induced GLUT4 mRNA accumulation, insulin-induced GLUT4 protein content, GLUT4-CAT transactivation and glucose uptake. Furthermore, co-transfection of brown adipocytes with a dominant-negative form of PI 3-kinase precluded the transactivation of the GLUT4 promoter by insulin. However, inhibition of p70S6 kinase (p70(s6k)) with rapamycin or of mitogen-activated protein kinase (MAPK) with PD098059 does not preclude insulin effects on GLUT4 gene expression or glucose uptake. Our results show for the first time a positive effect of insulin on GLUT4 gene expression in fetal brown adipocytes, suggesting the existence of insulin response element(s) in its promoter. Moreover, PI 3-kinase, but not p70(s6k) or MAPK, is an essential requirement for insulin regulation of GLUT4 gene expression. PMID:9895282

  2. Housekeeping genes in prostate tumorigenesis.

    PubMed

    Byun, Jinyoung; Logothetis, Christopher J; Gorlov, Ivan P

    2009-12-01

    Housekeeping (HK) genes are involved in basic cellular functions and tend to be constitutively expressed across various tissues and conditions. A number of studies have analyzed the value of HK genes as an internal standard for assessing gene expression, but the role of HK genes in cancer development has never been specifically addressed. In this study, we sought to evaluate the expression of HK genes during prostate tumorigenesis. We performed a meta-analysis of gene expression during the transition from normal prostate (NP) to localized prostate cancer (LPC) (i.e., NP > LPC) and from localized to metastatic prostate cancer (MPC) (i.e., LPC > MPC). We found that HK genes are more likely to be differentially expressed during prostate tumorigenesis than is the average gene in the human genome, suggesting that prostate tumorigenesis is driven by modulation of the expression of HK genes. Cell-cycle genes and proliferation markers were up-regulated in both NP > LPC and LPC > MPC transitions. We also found that the genes encoding ribosomal proteins were up-regulated in the NP > LPC and down-regulated in the LPC > MPC transition. The expression of heat shock proteins was up-regulated during the LPC > MPC transition, suggesting that in its advanced stages, prostate tumor is under cellular stress. The results of these analyses suggest that during prostate tumorigenesis, there is a period when the tumor is under cellular stress and, therefore, may be the most vulnerable and responsive to treatment. PMID:19551858

  3. A Sexual Shift Induced by Silencing of a Single Insulin-Like Gene in Crayfish: Ovarian Upregulation and Testicular Degeneration

    PubMed Central

    Rosen, Ohad; Manor, Rivka; Weil, Simy; Gafni, Ohad; Linial, Assaf; Aflalo, Eliahu D.; Ventura, Tomer; Sagi, Amir

    2010-01-01

    In sequential hermaphrodites, intersexuality occurs naturally, usually as a transition state during sexual re-differentiation processes. In crustaceans, male sexual differentiation is controlled by the male-specific androgenic gland (AG). An AG-specific insulin-like gene, previously identified in the red-claw crayfish Cherax quadricarinatus (designated Cq-IAG), was found in this study to be the prominent transcript in an AG cDNA subtractive library. In C. quadricarinatus, sexual plasticity is exhibited by intersex individuals in the form of an active male reproductive system and male secondary sex characters, along with a constantly arrested ovary. This intersexuality was exploited to follow changes caused by single gene silencing, accomplished via dsRNA injection. Cq-IAG silencing induced dramatic sex-related alterations, including male feature feminization, a reduction in sperm production, extensive testicular degeneration, expression of the vitellogenin gene, and accumulation of yolk proteins in the developing oocytes. Upon silencing of the gene, AG cells hypertrophied, possibly to compensate for low hormone levels, as reflected in the poor production of the insulin-like hormone (and revealed by immunohistochemistry). These results demonstrate both the functionality of Cq-IAG as an androgenic hormone-encoding gene and the dependence of male gonad viability on the Cq-IAG product. This study is the first to provide evidence that silencing an insulin-like gene in intersex C. quadricarinatus feminizes male-related phenotypes. These findings, moreover, contribute to the understanding of the regulation of sexual shifts, whether naturally occurring in sequential hermaphrodites or abnormally induced by endocrine disruptors found in the environment, and offer insight into an unusual gender-related link to the evolution of insulins. PMID:21151555

  4. RT-qPCR reveals opsin gene upregulation associated with age and sex in guppies (Poecilia reticulata) - a species with color-based sexual selection and 11 visual-opsin genes

    PubMed Central

    2011-01-01

    Background PCR-based surveys have shown that guppies (Poecilia reticulata) have an unusually large visual-opsin gene repertoire. This has led to speculation that opsin duplication and divergence has enhanced the evolution of elaborate male coloration because it improves spectral sensitivity and/or discrimination in females. However, this conjecture on evolutionary connections between opsin repertoire, vision, mate choice, and male coloration was generated with little data on gene expression. Here, we used RT-qPCR to survey visual-opsin gene expression in the eyes of males, females, and juveniles in order to further understand color-based sexual selection from the perspective of the visual system. Results Juvenile and adult (male and female) guppies express 10 visual opsins at varying levels in the eye. Two opsin genes in juveniles, SWS2B and RH2-2, accounted for >85% of all visual-opsin transcripts in the eye, excluding RH1. This relative abundance (RA) value dropped to about 65% in adults, as LWS-A180 expression increased from approximately 3% to 20% RA. The juvenile-to-female transition also showed LWS-S180 upregulation from about 1.5% to 7% RA. Finally, we found that expression in guppies' SWS2-LWS gene cluster is negatively correlated with distance from a candidate locus control region (LCR). Conclusions Selective pressures influencing visual-opsin gene expression appear to differ among age and sex. LWS upregulation in females is implicated in augmenting spectral discrimination of male coloration and courtship displays. In males, enhanced discrimination of carotenoid-rich food and possibly rival males are strong candidate selective pressures driving LWS upregulation. These developmental changes in expression suggest that adults possess better wavelength discrimination than juveniles. Opsin expression within the SWS2-LWS gene cluster appears to be regulated, in part, by a common LCR. Finally, by comparing our RT-qPCR data to MSP data, we were able to propose the first opsin-to-?max assignments for all photoreceptor types in the cone mosaic. PMID:21447186

  5. Intracranial self stimulation upregulates the expression of synaptic plasticity related genes and Arc protein expression in rat hippocampus.

    PubMed

    Kádár, E; Huguet, G; Aldavert-Vera, L; Morgado-Bernal, I; Segura-Torres, P

    2013-11-01

    Post-training lateral hypothalamus (LH) intracranial self stimulation (ICSS) has a reliable enhancing effect on explicit memory formation evaluated in hippocampus-dependent tasks such as the Morris water maze. In this study, the effects of ICSS on gene expression in the hippocampus are examined 4.5 h post treatment by using oligonucleotide microarray and real-time PCR, and by measuring Arc protein levels in the different layers of hippocampal subfields through immunofluorescence. The microarray data analysis resulted in 65 significantly regulated genes in rat ICSS hippocampi compared to sham, including cAMP-mediated signaling as one of the most significantly enriched Database for Annotation, Visualization and Integrated Discovery (DAVID) functional categories. In particular, expression of CREB-dependent synaptic plasticity related genes (c-Fos, Arc, Bdnf, Ptgs-2 and Crem and Icer) was regulated in a time-dependent manner following treatment administration. Immunofluorescence results showed that ICSS treatment induced a significant increase in Arc protein expression in CA1 and DG hippocampal subfields. This empirical evidence supports our hypothesis that the effect of ICSS on improved or restored memory functions might be mediated by increased hippocampal expression of activity-dependent synaptic plasticity related genes, including Arc protein expression, as neural mechanisms related to memory consolidation. PMID:23898803

  6. Exposure to cell phone radiation up-regulates apoptosis genes in primary cultures of neurons and astrocytes

    Microsoft Academic Search

    Tian-Yong Zhao; Shi-Ping Zou; Pamela E. Knapp

    2007-01-01

    The health effects of cell phone radiation exposure are a growing public concern. This study investigated whether expression of genes related to cell death pathways are dysregulated in primary cultured neurons and astrocytes by exposure to a working Global System for Mobile Communication (GSM) cell phone rated at a frequency of 1900MHz. Primary cultures were exposed to cell phone emissions

  7. Genes Up-Regulated in Tolerant Cavendish Banana Roots in Response to Fusarium oxysporum f. sp. cubense Infection1

    E-print Network

    Programme, Scottish Crop Research Institute, Invergowrie, Dundee, DD2 5DA, UK Keywords: catalase, defence-associated genes (catalase 2, pectin acetyl esterase (PAE), PR-1 and PR-3) were selected for expression profile, xylanase inhibitor, peroxidase, catalase 2, metallothionein, response regulator 6 and tripsin inhibitor

  8. Increased leptin expression in endometriosis cells is associated with endometrial stromal cell proliferation and leptin gene up-regulation

    Microsoft Academic Search

    Meng-Hsing Wu; Pei-Ching Chuang; Hsiu-Mei Chen; Chen-Chung Lin; Shaw-Jenq Tsai

    2002-01-01

    Endometriosis is a polygenic disease with complex, multifactorial aetiologies affecting ~10% of women of reproductive age. Leptin is the product of the ob gene, which is related to reproductive function and immunological alteration. The angiogenic and mitogenic action of leptin may influence the formation of endometriosis. This study was aimed at determining whether leptin and leptin receptor expression differs in

  9. Sustained ?-catenin activity in dermal fibroblasts promotes fibrosis by up-regulating expression of extracellular matrix protein-coding genes.

    PubMed

    Hamburg-Shields, Emily; DiNuoscio, Gregg J; Mullin, Nathaniel K; Lafayatis, Robert; Atit, Radhika P

    2015-04-01

    Fibrosis is an end-stage response to tissue injury that is associated with loss of organ function as a result of excess extracellular matrix (ECM) production by fibroblasts. In skin, pathological fibrosis is evident during keloid scar formation, systemic sclerosis (SSc) and morphea. Dermal fibroblasts in these fibrotic diseases exhibit increased Wnt/?-catenin signalling, a pathway that is sufficient to cause fibrosis in mice. However, in the context of this complex pathology, the precise pro-fibrotic consequences of Wnt/?-catenin signalling are not known. We found that expression of stabilized ?-catenin in mouse dermal fibroblasts resulted in spontaneous, progressive skin fibrosis with thickened collagen fibres and altered collagen fibril morphology. The fibrotic phenotype was predominated by resident dermal fibroblasts. Genome-wide profiling of the fibrotic mouse dermis revealed elevated expression of matrix-encoding genes, and the promoter regions of these genes were enriched for Tcf/Lef family transcription factor binding sites. Additionally, we identified 32 ?-catenin-responsive genes in our mouse model that are also over-expressed in human fibrotic tissues and poised for regulation by Tcf/Lef family transcription factors. Therefore, we have uncovered a matrix-regulatory role for stabilized ?-catenin in fibroblasts in vivo and have defined a set of ?-catenin-responsive genes with relevance to fibrotic disease. PMID:25385294

  10. ARACNE: An Algorithm for the Reconstruction of Gene Regulatory Networks in a Mammalian Cellular Context

    PubMed Central

    Margolin, Adam A; Nemenman, Ilya; Basso, Katia; Wiggins, Chris; Stolovitzky, Gustavo; Favera, Riccardo Dalla; Califano, Andrea

    2006-01-01

    Background Elucidating gene regulatory networks is crucial for understanding normal cell physiology and complex pathologic phenotypes. Existing computational methods for the genome-wide "reverse engineering" of such networks have been successful only for lower eukaryotes with simple genomes. Here we present ARACNE, a novel algorithm, using microarray expression profiles, specifically designed to scale up to the complexity of regulatory networks in mammalian cells, yet general enough to address a wider range of network deconvolution problems. This method uses an information theoretic approach to eliminate the majority of indirect interactions inferred by co-expression methods. Results We prove that ARACNE reconstructs the network exactly (asymptotically) if the effect of loops in the network topology is negligible, and we show that the algorithm works well in practice, even in the presence of numerous loops and complex topologies. We assess ARACNE's ability to reconstruct transcriptional regulatory networks using both a realistic synthetic dataset and a microarray dataset from human B cells. On synthetic datasets ARACNE achieves very low error rates and outperforms established methods, such as Relevance Networks and Bayesian Networks. Application to the deconvolution of genetic networks in human B cells demonstrates ARACNE's ability to infer validated transcriptional targets of the cMYC proto-oncogene. We also study the effects of misestimation of mutual information on network reconstruction, and show that algorithms based on mutual information ranking are more resilient to estimation errors. Conclusion ARACNE shows promise in identifying direct transcriptional interactions in mammalian cellular networks, a problem that has challenged existing reverse engineering algorithms. This approach should enhance our ability to use microarray data to elucidate functional mechanisms that underlie cellular processes and to identify molecular targets of pharmacological compounds in mammalian cellular networks. PMID:16723010

  11. Complex cellular functions of the von Hippel–Lindau tumor suppressor gene: insights from model organisms

    PubMed Central

    Hsu, T

    2012-01-01

    The von Hippel–Lindau tumor suppressor gene (VHL) has attracted intensive interest not only because its mutations predispose carriers to devastating tumors, but also because it is involved in oxygen sensing under physiological conditions. VHL loss-of-function mutations result in organ-specific tumors, such as hemangioblastoma of the central nervous system and renal cell carcinoma, both untreatable with conventional chemotherapies. The VHL protein is best known as an E3 ubiquitin ligase that targets hypoxia-inducible factor-? (HIF-?), but many diverse, non-canonical cellular functions have also been assigned to VHL, mainly based on studies in cell culture systems. As such, although the HIF-dependent role of VHL is critical, the full spectrum of pathophysiological functions of VHL is still unresolved. Such understanding requires careful cross-referencing with physiologically relevant experimental models. Studies in model systems, such as Caenorhabditis elegans, Drosophila, zebrafish and mouse have provided critical in vivo confirmation of the VHL–HIF pathway, and verification of potentially important cellular functions including microtubule stabilization and epithelial morphogenesis. More recently, animal models have also suggested systemic roles of VHL in hematopoiesis, metabolic homeostasis and inflammation. In this review, the studies performed in model organisms will be summarized and placed in context with existing clinical and in vitro data. PMID:21996733

  12. Salmonella enterica Serovar Typhimurium Colonizing the Lumen of the Chicken Intestine Grows Slowly and Upregulates a Unique Set of Virulence and Metabolism Genes?

    PubMed Central

    Harvey, P. C.; Watson, M.; Hulme, S.; Jones, M. A.; Lovell, M.; Berchieri, A.; Young, J.; Bumstead, N.; Barrow, P.

    2011-01-01

    The pattern of global gene expression in Salmonella enterica serovar Typhimurium bacteria harvested from the chicken intestinal lumen (cecum) was compared with that of a late-log-phase LB broth culture using a whole-genome microarray. Levels of transcription, translation, and cell division in vivo were lower than those in vitro. S. Typhimurium appeared to be using carbon sources, such as propionate, 1,2-propanediol, and ethanolamine, in addition to melibiose and ascorbate, the latter possibly transformed to d-xylulose. Amino acid starvation appeared to be a factor during colonization. Bacteria in the lumen were non- or weakly motile and nonchemotactic but showed upregulation of a number of fimbrial and Salmonella pathogenicity island 3 (SPI-3) and 5 genes, suggesting a close physical association with the host during colonization. S. Typhimurium bacteria harvested from the cecal mucosa showed an expression profile similar to that of bacteria from the intestinal lumen, except that levels of transcription, translation, and cell division were higher and glucose may also have been used as a carbon source. PMID:21768276

  13. IL-17A enhances the expression of profibrotic genes through upregulation of the TGF-? receptor on hepatic stellate cells in a JNK-dependent manner.

    PubMed

    Fabre, Thomas; Kared, Hassen; Friedman, Scott L; Shoukry, Naglaa H

    2014-10-15

    Activation of hepatic stellate cells (HSCs) is a key event in the initiation of liver fibrosis, characterized by enhanced extracellular matrix production and altered degradation. Activation of HSCs can be modulated by cytokines produced by immune cells. Recent reports have implicated the proinflammatory cytokine IL-17A in liver fibrosis progression. We hypothesized that IL-17A may enhance activation of HSCs and induction of the fibrogenic signals in these cells. The human HSC line LX2 and primary human HSCs were stimulated with increasing doses of IL-17A and compared with TGF-?- and PBS-treated cells as positive and negative controls, respectively. IL-17A alone did not induce activation of HSCs. However, IL-17A sensitized HSCs to the action of suboptimal doses of TGF-? as confirmed by strong induction of ?-smooth muscle actin, collagen type I (COL1A1), and tissue inhibitor of matrix metalloproteinase I gene expression and protein production. IL-17A specifically upregulated the cell surface expression of TGF-?RII following stimulation. Pretreatment of HSCs with IL-17A enhanced signaling through TGF-?RII as observed by increased phosphorylation of SMAD2/3 in response to stimulation with suboptimal doses of TGF-?. This enhanced TGF-? response of HSCs induced by IL-17A was JNK-dependent. Our results suggest a novel profibrotic function for IL-17A by enhancing the response of HSCs to TGF-? through activation of the JNK pathway. IL-17A acts through upregulation and stabilization of TGF-?RII, leading to increased SMAD2/3 signaling. These findings represent a novel example of cooperative signaling between an immune cytokine and a fibrogenic receptor. PMID:25210118

  14. Glutathione S-Transferase (GST) Gene Diversity in the Crustacean Calanus finmarchicus - Contributors to Cellular Detoxification.

    PubMed

    Roncalli, Vittoria; Cieslak, Matthew C; Passamaneck, Yale; Christie, Andrew E; Lenz, Petra H

    2015-01-01

    Detoxification is a fundamental cellular stress defense mechanism, which allows an organism to survive or even thrive in the presence of environmental toxins and/or pollutants. The glutathione S-transferase (GST) superfamily is a set of enzymes involved in the detoxification process. This highly diverse protein superfamily is characterized by multiple gene duplications, with over 40 GST genes reported in some insects. However, less is known about the GST superfamily in marine organisms, including crustaceans. The availability of two de novo transcriptomes for the copepod, Calanus finmarchicus, provided an opportunity for an in depth study of the GST superfamily in a marine crustacean. The transcriptomes were searched for putative GST-encoding transcripts using known GST proteins from three arthropods as queries. The identified transcripts were then translated into proteins, analyzed for structural domains, and annotated using reciprocal BLAST analysis. Mining the two transcriptomes yielded a total of 41 predicted GST proteins belonging to the cytosolic, mitochondrial or microsomal classes. Phylogenetic analysis of the cytosolic GSTs validated their annotation into six different subclasses. The predicted proteins are likely to represent the products of distinct genes, suggesting that the diversity of GSTs in C. finmarchicus exceeds or rivals that described for insects. Analysis of relative gene expression in different developmental stages indicated low levels of GST expression in embryos, and relatively high expression in late copepodites and adult females for several cytosolic GSTs. A diverse diet and complex life history are factors that might be driving the multiplicity of GSTs in C. finmarchicus, as this copepod is commonly exposed to a variety of natural toxins. Hence, diversity in detoxification pathway proteins may well be key to their survival. PMID:25945801

  15. MPI on Blue Gene\\/L : Designing an Efficient General Purpose Messaging Solution for a Large Cellular System

    Microsoft Academic Search

    George Almasi; Charles Archer; Jose G. Castanos; Manish Gupta; Xavier Martorell; Jose E. Moreira; William Gropp; Silvius Rus; Brian Toonen

    The Blue Gene\\/L supercomputer uses system-on-a-chip integration and a highly scalable 65,536 node cellular architecture to deliver 360 Teraflops of peak computing power. Efficient operation of the machine requires a fast, scalable and standards compliant MPI library. Researchers at IBM and Argonne National Labs are porting the MPICH2 library to Blue Gene\\/L. We present the current state of the design

  16. Microarray analysis of differentially expressed genes in placental tissue of pre-eclampsia: up-regulation of obesity-related genes

    Microsoft Academic Search

    T. Reimer; D. Koczan; B. Gerber; D. Richter; H. J. Thiesen; K. Friese

    2002-01-01

    Susceptibility genes present in both mother and fetus most likely contribute to the risk of pre-eclampsia. Placental biopsies were therefore investigated by high-density DNA microarray analysis to determine genes differentially regulated within chorionic villous tissue in pre-eclampsia and normal pregnancy. The pooled RNAs of pre-eclamptic and normotensive subjects were hybridized to the HuGeneFL array representing sequences from ~5600 full-length human

  17. Against All Odds: Trehalose-6-Phosphate Synthase and Trehalase Genes in the Bdelloid Rotifer Adineta vaga Were Acquired by Horizontal Gene Transfer and Are Upregulated during Desiccation

    PubMed Central

    Hespeels, Boris; Li, Xiang; Flot, Jean-François; Pigneur, Lise-Marie; Malaisse, Jeremy; Da Silva, Corinne; Van Doninck, Karine

    2015-01-01

    The disaccharide sugar trehalose is essential for desiccation resistance in most metazoans that survive dryness; however, neither trehalose nor the enzymes involved in its metabolism have ever been detected in bdelloid rotifers despite their extreme resistance to desiccation. Here we screened the genome of the bdelloid rotifer Adineta vaga for genes involved in trehalose metabolism. We discovered a total of four putative trehalose-6-phosphate synthase (TPS) and seven putative trehalase (TRE) gene copies in the genome of this ameiotic organism; however, no trehalose-6-phosphate phosphatase (TPP) gene or domain was detected. The four TPS copies of A. vaga appear more closely related to plant and fungi proteins, as well as to some protists, whereas the seven TRE copies fall in bacterial clades. Therefore, A. vaga likely acquired its trehalose biosynthesis and hydrolysis genes by horizontal gene transfers. Nearly all residues important for substrate binding in the predicted TPS domains are highly conserved, supporting the hypothesis that several copies of the genes might be functional. Besides, RNAseq library screening showed that trehalase genes were highly expressed compared to TPS genes, explaining probably why trehalose had not been detected in previous studies of bdelloids. A strong overexpression of their TPS genes was observed when bdelloids enter desiccation, suggesting a possible signaling role of trehalose-6-phosphate or trehalose in this process. PMID:26161530

  18. The Majority of Viral-Cellular Fusion Transcripts in Cervical Carcinomas Cotranscribe Cellular Sequences of Known or Predicted Genes

    Microsoft Academic Search

    Irene Kraus; Corina Driesch; Svetlana Vinokurova; Eivind Hovig; Magnus von Knebel Doeberitz; Matthias Durst

    2008-01-01

    Integration of human papillomavirus (HPV) DNA into the host genome is a frequent event in cervical carcinogenesis and is reported to occur at randomly selected chromosomal sites. However,as the databases are being up-dated continuously, the knowledge based on sequenced viral integration sites also expands. In this study,viral-cellular fusion transcripts of a preselected group of 74 cervical carcinoma or cervical intraepithelial

  19. Panax ginseng extract modulates oxidative stress, DNA fragmentation and up-regulate gene expression in rats sub chronically treated with aflatoxin B1 and fumonisin B 1.

    PubMed

    Hassan, Aziza M; Abdel-Aziem, Sekena H; El-Nekeety, Aziza A; Abdel-Wahhab, Mosaad A

    2014-04-19

    Aflatoxins and fumonisins are important food-borne mycotoxins implicated in human health and have cytotoxic effects. The aims of the current study were to evaluate the protective role of Panax ginseng extract (PGE) against the synergistic effect of subchronic administration of aflatoxin B1 (AFB1) and fumonisin B1 (FB1) on DNA and gene expression in rat. Female Sprague-Dawley rats were divided into eight groups (ten rats/group) and treated for 12 weeks including the control group, the group having received AFB1 (80 µg/kg bw), the group having received FB1 (100 µg/kg bw), the group having received AFB1 plus FB1 and the groups having received PGE (20 mg/kg bw) alone or with AFB1 and/or FB1. At the end of experiment, liver and kidney were collected for the determination of DNA fragmentation, lipid peroxidation (LP), glutathione (GSH) contents and alterations in gene expression. The results indicated that these mycotoxins increased DNA fragmentation, LP and decreased GSH content in liver and kidney and down-regulated gene expression of antioxidants enzymes. The combined treatments with AFB1 and/or FB1 plus PGE suppressed DNA fragmentation only in the liver, normalized LP and increased GSH in the liver and kidney as well as up-regulated the expression of GPx, SOD1 and CAT mRNA. It could be concluded that AFB1 and FB1 have synergistic genotoxic effects. PGE induced protective effects against their oxidative stress and genotoxicity through its antioxidant properties. PMID:24748134

  20. Genistein, a soy isoflavone, up-regulates expression of antioxidant genes: involvement of estrogen receptors, ERK1/2, and NFkappaB.

    PubMed

    Borrás, Consuelo; Gambini, Juan; Gómez-Cabrera, M Carmen; Sastre, Juan; Pallardó, Federico V; Mann, Giovanni E; Viña, Jose

    2006-10-01

    We have previously reported that estrogens up-regulate longevity-associated genes. As recent evidence has shown that estrogen replacement therapy is associated with an increased risk of cardiovascular disease, we have studied the effects of genistein, a soy isoflavone with a similar structure to estradiol, on the expression of antioxidant, longevity-related genes. MCF-7 cells (human mammary gland tumor cell line) were incubated for 48 h with 0.5 microM genistein, a concentration found in the plasma of populations consuming diets rich in soy protein. Peroxide levels were determined by fluorimetry, activation of extracellular-signal regulated kinase (ERK1/2), and nuclear factor kappaB (NFkappaB)-signaling pathways by Western blot analysis and ELISA, respectively, and mRNA expression of antioxidant genes by real-time reverse transcriptase-polymerase chain reaction (RT-PCR). Inhibition of basal peroxide levels in MCF-7 cells by genistein was prevented by pretreatment of cells with the estrogen receptor antagonist tamoxifen. Phosphorylation of extracellular regulated kinase (ERK)1/2 led to an activation of NFkappaB, as indicated by increased p50 subunit expression in nuclear extracts, and increased mRNA levels of the antioxidant enzyme manganese-superoxide dismutase (MnSOD). Inhibition of ERK1/2 abrogated genistein-mediated NFkappaB activation and elevated expression of MnSOD. Our molecular studies may provide a basis to determine the effects of genistein and other soy protein-derived products on longevity in both animals and the human population. PMID:16966488

  1. Overfeeding energy upregulates peroxisome proliferator-activated receptor (PPAR)?-controlled adipogenic and lipolytic gene networks but does not affect proinflammatory markers in visceral and subcutaneous adipose depots of Holstein cows.

    PubMed

    Ji, P; Drackley, J K; Khan, M J; Loor, J J

    2014-06-01

    Our objective was to determine the effects of overfeeding energy on gene expression in mesenteric (MAT), omental (OAT), and subcutaneous (SAT) adipose tissue (AT) from nonpregnant and nonlactating Holstein cows. Eighteen cows were randomly assigned to either a low energy [LE, net energy for lactation (NE(L)) = 1.35 Mcal/kg of dry matter (DM)] or high energy (HE, NE(L) = 1.62 Mcal/kg of DM) diets for 8 wk. Cows were then euthanized and subsamples of MAT, OAT, and SAT were harvested for transcript profiling via quantitative PCR of 34 genes involved in lipogenesis, triacylglycerol (TAG) synthesis, lipolysis, lactate signaling, transcription regulation, and inflammation. The interaction of dietary energy and AT depot was only significant for LPL, which indicated a consistent response among the 3 sites. The expression of key genes related to de novo fatty acid synthesis (FASN) and desaturation (SCD) was upregulated by HE compared with LE. Other genes associated with those processes, such as ACLY, ACACA, ELOVL6, FABP4, GPAM, and LPIN1, were numerically upregulated by HE. The expression of lipolytic (PNPLA2 and ABHD5) genes was upregulated and the antilypolytic lactate receptor HCAR1 was downregulated with HE compared with LE. The putative transcription regulator THRSP was upregulated and the transcription regulator PPARG tended to be upregulated by HE, whereas SREBF1 was downregulated. Among adipocytokines, HE tended to upregulate the expression of CCL2, whereas IL6R was downregulated. Overall, results indicated that overfeeding energy may increase AT mass at least in part by stimulating transcription of the network encompassing key genes associated with de novo synthesis. In response to energy overfeeding, the expression of PPARG rather than SREBF1 was closely associated with most adipogenic or lipogenic genes. However, the transcriptional activity of these regulators needs to be verified to confirm their role in the regulation of adipogenesis or lipogenesis in bovine AT. Overfeeding energy also may predispose cows to greater lipolytic potential by stimulating expression of TAG hydrolysis genes while inhibiting signaling via hydroxycarboxylic acid receptor (HCAR1), which is a novel antilipolytic regulator. Our results do not support an overt inflammatory response in adipose tissues in response to an 8-wk energy overfeeding. PMID:24704238

  2. Urotensin II upregulates migration and cytokine gene expression in leukocytes of the African clawed frog, Xenopus laevis.

    PubMed

    Tomiyama, Shiori; Nakamachi, Tomoya; Uchiyama, Minoru; Matsuda, Kouhei; Konno, Norifumi

    2015-05-15

    Urotensin II (UII) exhibits diverse physiological actions including vasoconstriction, locomotor activity, osmoregulation, and immune response via the UII receptor (UTR) in mammals. However, in amphibians the function of the UII-UTR system remains unknown. In the present study, we investigated the potential immune function of UII using leukocytes isolated from the African clawed frog, Xenopus laevis. Stimulation of male frogs with lipopolysaccharide increased mRNA expression of UII and UTR in leukocytes, suggesting that inflammatory stimuli induce activation of the UII-UTR system. Migration assays showed that both UII and UII-related peptide enhanced migration of leukocytes in a dose-dependent manner, and that UII effect was inhibited by the UTR antagonist urantide. Inhibition of Rho kinase with Y-27632 abolished UII-induced migration, suggesting that it depends on the activation of RhoA/Rho kinase. Treatment of isolated leukocytes with UII increased the expression of several cytokine genes including tumor necrosis factor-?, interleukin-1?, and macrophage migration inhibitory factor, and the effects were abolished by urantide. These results suggest that in amphibian leukocytes the UII-UTR system is involved in the activation of leukocyte migration and cytokine gene expression in response to inflammatory stimuli. PMID:25907658

  3. Toxoplasma gondii Lysine Acetyltransferase GCN5-A Functions in the Cellular Response to Alkaline Stress and Expression of Cyst Genes

    PubMed Central

    Naguleswaran, Arunasalam; Elias, Eliana V.; McClintick, Jeanette; Edenberg, Howard J.; Sullivan, William J.

    2010-01-01

    Parasitic protozoa such as the apicomplexan Toxoplasma gondii progress through their life cycle in response to stimuli in the environment or host organism. Very little is known about how proliferating tachyzoites reprogram their expressed genome in response to stresses that prompt development into latent bradyzoite cysts. We have previously linked histone acetylation with the expression of stage-specific genes, but the factors involved remain to be determined. We sought to determine if GCN5, which operates as a transcriptional co-activator by virtue of its histone acetyltransferase (HAT) activity, contributed to stress-induced changes in gene expression in Toxoplasma. In contrast to other lower eukaryotes, Toxoplasma has duplicated its GCN5 lysine acetyltransferase (KAT). Disruption of the gene encoding for TgGCN5-A in type I RH strain did not produce a severe phenotype under normal culture conditions, but here we show that the TgGCN5-A null mutant is deficient in responding to alkaline pH, a common stress used to induce bradyzoite differentiation in vitro. We performed a genome-wide analysis of the Toxoplasma transcriptional response to alkaline pH stress, finding that parasites deleted for TgGCN5-A fail to up-regulate 74% of the stress response genes that are induced 2-fold or more in wild-type. Using chromatin immunoprecipitation, we verify an enrichment of TgGCN5-A at the upstream regions of genes activated by alkaline pH exposure. The TgGCN5-A knockout is also incapable of up-regulating key marker genes expressed during development of the latent cyst form, and is impaired in its ability to recover from alkaline stress. Complementation of the TgGCN5-A knockout restores the expression of these stress-induced genes and reverses the stress recovery defect. These results establish TgGCN5-A as a major contributor to the alkaline stress response in RH strain Toxoplasma. PMID:21179246

  4. Structure-guided mutational analysis of gene regulation by the Bacillus subtilis pbuE adenine-responsive riboswitch in a cellular context.

    PubMed

    Marcano-Velázquez, Joan G; Batey, Robert T

    2015-02-13

    Riboswitches are a broadly distributed form of RNA-based gene regulation in Bacteria and, more rarely, Archaea and Eukarya. Most often found in the 5'-leader sequence of bacterial mRNAs, they are generally composed of two functional domains: a receptor (aptamer) domain that binds an effector molecule and a regulatory domain (or expression platform) that instructs the expression machinery. One of the most studied riboswitches is the Bacillus subtilis adenine-responsive pbuE riboswitch, which regulates gene expression at the transcriptional level, up-regulating expression in response to increased intracellular effector concentrations. In this work, we analyzed sequence and structural elements that contribute to efficient ligand-dependent regulatory activity in a co-transcriptional and cellular context. Unexpectedly, we found that the P1 helix, which acts as the antitermination element of the switch in this RNA, supported ligand-dependent activation of a reporter gene over a broad spectrum of lengths from 3 to 10 bp. This same trend was also observed using a minimal in vitro single-turnover transcription assay, revealing that this behavior is intrinsic to the RNA sequence. We also found that the sequences at the distal tip of the terminator not directly involved in alternative secondary structure formation are highly important for efficient regulation. These data strongly support a model in which the switch is highly localized to the P1 helix adjacent to the ligand-binding pocket that likely presents a local kinetic block to invasion of the aptamer by the terminator. PMID:25550163

  5. Mechanistic links between cellular trade-offs, gene expression, and growth.

    PubMed

    Weiße, Andrea Y; Oyarzún, Diego A; Danos, Vincent; Swain, Peter S

    2015-03-01

    Intracellular processes rarely work in isolation but continually interact with the rest of the cell. In microbes, for example, we now know that gene expression across the whole genome typically changes with growth rate. The mechanisms driving such global regulation, however, are not well understood. Here we consider three trade-offs that, because of limitations in levels of cellular energy, free ribosomes, and proteins, are faced by all living cells and we construct a mechanistic model that comprises these trade-offs. Our model couples gene expression with growth rate and growth rate with a growing population of cells. We show that the model recovers Monod's law for the growth of microbes and two other empirical relationships connecting growth rate to the mass fraction of ribosomes. Further, we can explain growth-related effects in dosage compensation by paralogs and predict host-circuit interactions in synthetic biology. Simulating competitions between strains, we find that the regulation of metabolic pathways may have evolved not to match expression of enzymes to levels of extracellular substrates in changing environments but rather to balance a trade-off between exploiting one type of nutrient over another. Although coarse-grained, the trade-offs that the model embodies are fundamental, and, as such, our modeling framework has potentially wide application, including in both biotechnology and medicine. PMID:25695966

  6. Maximizing gene delivery efficiencies of cationic helical polypeptides via balanced membrane penetration and cellular targeting

    PubMed Central

    Zheng, Nan; Yin, Lichen; Song, Ziyuan; Ma, Liang; Tang, Haoyu; Gabrielson, Nathan P.; Lu, Hua; Cheng, Jianjun

    2014-01-01

    The application of non-viral gene delivery vectors is often accompanied with the poor correlation between transfection efficiency and the safety profiles of vectors: vectors with high transfection efficiencies often suffer from high toxicities, making it unlikely to improve their efficiencies by increasing the DNA dosage. In the current study, we developed a ternary complex system which consisted of a highly membrane-active cationic helical polypeptide (PVBLG-8), a low-toxic, membrane-inactive cationic helical polypeptide (PVBLG-7) capable of mediating mannose receptor targeting, and DNA. The PVBLG-7 moiety notably enhanced the cellular uptake and transfection efficiency of PVBLG-8 in a variety of mannose receptor-expressing cell types (HeLa, COS-7, and Raw 264.7), while it did not compromise the membrane permeability of PVBLG-8 or bring additional cytotoxicities. Because of the simplicity and adjustability of the self-assembly approach, optimal formulations of the ternary complexes with a proper balance between membrane activity and targeting capability were easily identified in each specific cell type. The optimal ternary complexes displayed desired cell tolerability and markedly outperformed the PVBLG-8/DNA binary complexes as well as commercial reagent Lipofectamine™ 2000 in terms of transfection efficiency. This study therefore provides an effective and facile strategy to overcome the efficiency-toxicity poor correlation of non-viral vectors, which contributes insights into the design strategy of effective and safe non-viral gene delivery vectors. PMID:24211080

  7. Lack of poly(ADP-ribose) polymerase-1 gene product enhances cellular sensitivity to arsenite.

    PubMed

    Poonepalli, Anuradha; Balakrishnan, Lakshmidevi; Khaw, Aik Kia; Low, Grace Kah Mun; Jayapal, Manikandan; Bhattacharjee, Rabindra N; Akira, Shizuo; Balajee, Adayabalam S; Hande, M Prakash

    2005-12-01

    Arsenite (As3+) has long been known to induce cancer and other degenerative diseases. Arsenite exerts its toxicity in part by generating reactive oxygen species. Identification of genetic factors that contribute to arsenic mutagenicity and carcinogenicity is critical for the treatment and prevention of arsenic exposure in human population. As poly(ADP-ribose) polymerase (PARP) is critical for genomic DNA stability, role of PARP-1 was evaluated in arsenic-induced cytotoxic and genotoxic effects. Our study revealed that telomere attrition, probably owing to arsenite-induced oxidative stress, was much more pronounced in PARP-1-/- mouse embryonic fibroblasts (MEF; 40%) compared with PARP-1+/+ MEFs (10-20%). Correlation observed between telomere reduction and apoptotic death in PARP-1 null cells strongly indicates that the telomere attrition might be a trigger for enhanced apoptotic death after arsenite treatment. Elevated DNA damage detected by alkaline comet assay points to an impaired repair ability of arsenite-induced DNA lesions in PARP-1-/- MEFs. Consistent with elevated DNA damage, increased micronuclei induction reflecting gross genomic instability was also observed in arsenite-treated PARP-1-/- MEFs. Microarray analysis has revealed that arsenite treatment altered the expression of about 311 genes majority of which have known functions in cellular responses to stress/external stimulus and cell growth and/or maintenance. Our results suggest an important role for PARP-1 gene product in the maintenance of chromosome-genome stability in response to arsenite-induced DNA damage. PMID:16322246

  8. Polymorphisms in Cytokine and Cellular Adhesion Molecule Genes and Susceptibility to Hematotoxicity among Workers Exposed to Benzene

    Microsoft Academic Search

    Qing Lan; Luoping Zhang; Min Shen; Martyn T. Smith; Guilan Li; Roel Vermeulen; Stephen M. Rappaport; Matthew S. Forrest; Richard B. Hayes; Martha Linet; Mustafa Dosemeci; Rona S. Weinberg; Songnian Yin; Meredith Yeager; Robert Welch; Suramya Waidyanatha; Sungkyoon Kim; Stephen Chanock; Nathaniel Rothman

    2005-01-01

    Benzene is a recognized hematotoxin and leukemogen but its mechanism of action and the role of genetic susceptibility are still unclear. Cytokines, chemokines, and cellular adhesion molecules are soluble proteins that play an important regulatory role in hematopoiesis. We therefore hypothesized that variation in these genes could influence benzene-induced hematotoxicity. We analyzed common, well-studied single- nucleotide polymorphisms (SNPs) in 20

  9. Cellular and molecular mechanisms that mediate basal and tumour necrosis factor-alpha-induced regulation of myosin light chain kinase gene activity.

    PubMed

    Ye, Dongmei; Ma, Thomas Y

    2008-08-01

    The patients with Crohn's disease (CD) have a 'leaky gut' manifested by an increase in intestinal epithelial tight junction (TJ) permeability. Tumour necrosis factor-alpha (TNF-alpha) is a proto-typical pro-inflammatory cytokine that plays a central role in intestinal inflammation of CD. An important pro-inflammatory action of TNF-alpha is to cause a functional opening of intestinal TJ barrier. Previous studies have shown that TNF-alpha increase in TJ permeability was regulated by an increase in myosin light chain kinase (MLCK) gene activity and protein expression. The major aim of this study was to elucidate the cellular and molecular mechanisms that mediate basal and TNF-alpha-induced increase in MLCK gene activity. By progressive 5' deletion, minimal MLCK promoter was localized between -313 to +118 on MLCK promoter. A p53 binding site located within minimal promoter region was identified as an essential determinant for basal promoter activity. A 4 bp start site and a 5 bp downstream promoter element were required for MLCK gene activity. TNF-alpha-induced increase in MLCK promoter activity was mediated by NF-kappaB activation. There were eight kappaB binding sites on MLCK promoter. The NF-kappaB1 site at +48 to +57 mediated TNF-alpha-induced increase in MLCK promoter activity. The NF-kappaB2 site at -325 to -316 had a repressive role on promoter activity. The opposite effects on promoter activity were due to differences in the NF-kappaB dimer type binding to the kappaB sites. p50/p65 dimer preferentially binds to the NF-kappaB1 site and up-regulates promoter activity; while p50/p50 dimer preferentially binds to the NF-kappaB2 site and down-regulates promoter activity. In conclusion, we have identified the minimal MLCK promoter region, essential molecular determinants and molecular mechanisms that mediate basal and TNF-alpha-induced modulation of MLCK promoter activity in Caco-2 intestinal epithelial cells. These studies provide novel insight into the cellular and molecular mechanisms that regulate basal and TNF-alpha-induced modulation of MLCK gene activity. PMID:18363837

  10. c-Jun/c-Fos heterodimers regulate cellular genes via a newly identified class of methylated DNA sequence motifs

    PubMed Central

    Gustems, Montse; Woellmer, Anne; Rothbauer, Ulrich; Eck, Sebastian H.; Wieland, Thomas; Lutter, Dominik; Hammerschmidt, Wolfgang

    2014-01-01

    CpG methylation in mammalian DNA is known to interfere with gene expression by inhibiting the binding of transactivators to their cognate sequence motifs or recruiting proteins involved in gene repression. An Epstein–Barr virus-encoded transcription factor, Zta, was the first example of a sequence-specific transcription factor that preferentially recognizes and selectively binds DNA sequence motifs with methylated CpG residues, reverses epigenetic silencing and activates gene transcription. The DNA binding domain of Zta is homologous to c-Fos, a member of the cellular AP-1 (activator protein 1) transcription factor family, which regulates cell proliferation and survival, apoptosis, transformation and oncogenesis. We have identified a novel AP-1 binding site termed meAP-1, which contains a CpG dinucleotide. If methylated, meAP-1 sites are preferentially bound by the AP-1 heterodimer c-Jun/c-Fos in vitro and in cellular chromatin in vivo. In activated human primary B cells, c-Jun/c-Fos locates to these methylated elements in promoter regions of transcriptionally activated genes. Reminiscent of the viral Zta protein, c-Jun/c-Fos is the first identified cellular member of the AP-1 family of transactivators that can induce expression of genes with methylated, hence repressed promoters, reversing epigenetic silencing. PMID:24371273

  11. Inhibin beta E is upregulated by drug-induced endoplasmic reticulum stress as a transcriptional target gene of ATF4

    SciTech Connect

    Brüning, Ansgar, E-mail: ansgar.bruening@med.uni-muenchen.de; Matsingou, Christina; Brem, German Johannes; Rahmeh, Martina; Mylonas, Ioannis

    2012-10-15

    Inhibins and activins are gonadal peptide hormones of the transforming growth factor-? super family with important functions in the reproductive system. By contrast, the recently identified inhibin ?E subunit, primarily expressed in liver cells, appears to exert functions unrelated to the reproductive system. Previously shown downregulation of inhibin ?E in hepatoma cells and anti-proliferative effects of ectopic inhibin ?E overexpression indicated growth-regulatory effects of inhibin ?E. We observed a selective re-expression of the inhibin ?E subunit in HepG2 hepatoblastoma cells, MCF7 breast cancer cells, and HeLa cervical cancer cells under endoplasmic reticulum stress conditions induced by tunicamycin, thapsigargin, and nelfinavir. Analysis of XPB1 splicing and ATF4 activation revealed that inhibin ?E re-expression was associated with induction of the endoplasmic reticulum stress reaction by these drugs. Transfection of an ATF4 expression plasmid specifically induced inhibin ?E expression in HeLa cells and indicates inhibin ?E as a hitherto unidentified target gene of ATF4, a key transcription factor of the endoplasmic reticulum stress response. Therefore, the inhibin ?E subunit defines not only a new player but also a possible new marker for drug-induced endoplasmic reticulum stress. -- Highlights: ? Endoplasmic reticulum stress induces inhibin beta E expression. ? Inhibin beta E is regulated by the transcription factor ATF4. ? Inhibin beta E expression can be used as a marker for drug-induced ER stress.

  12. Apoptosis Induction of Human Bladder Cancer Cells by Sanguinarine through Reactive Oxygen Species-Mediated Up-Regulation of Early Growth Response Gene-1

    PubMed Central

    Han, Min Ho; Park, Cheol; Jin, Cheng-Yun; Kim, Gi-Young; Chang, Young-Chae; Moon, Sung-Kwon; Kim, Wun-Jae; Choi, Yung Hyun

    2013-01-01

    Although the effects of sanguinarine, a benzophenanthridine alkaloid, on the inhibition of some kinds of cancer cell growth have been established, the underlying mechanisms are not completely understood. This study investigated possible mechanisms by which sanguinarine exerts its anticancer action in cultured human bladder cancer cell lines (T24, EJ, and 5637). Sanguinarine treatment resulted in concentration-response growth inhibition of the bladder cancer cells by inducing apoptosis. Sanguinarine-induced apoptosis was correlated with the up-regulation of Bax, the down-regulation of Bid and XIAP, the activation of caspases (-3, -8, and -9), and the generation of increased reactive oxygen species (ROS). The ROS scavenger N-acetyl cysteine (NAC) completely reversed the sanguinarine-triggered apoptotic events. In addition, sanguinarine effectively increased the activation of the c-Jun N-terminal kinase (JNK) and the expression of the early growth response gene-1 (Egr-1), which was recovered by pretreatment with NAC. Furthermore, knockdown of Egr-1 expression by small interfering RNA attenuated sanguinarine-induced apoptosis, but not the JNK inhibitor, indicating that the interception of ROS generation blocked the sanguinarine-induced apoptotic effects via deregulation of the expression of Egr-1 proteins. Taken together, the data provide evidence that sanguinarine is a potent anticancer agent, which inhibits the growth of bladder cancer cells and induces their apoptosis through the generation of free radicals. PMID:23717422

  13. Interferon-Stimulated Gene 15 Upregulation Precedes the Development of Blood-Brain Barrier Disruption and Cerebral Edema after Traumatic Brain Injury in Young Mice.

    PubMed

    Rossi, Janet L; Todd, Tracey; Daniels, Zachary; Bazan, Nicolas G; Belayev, Ludmila

    2015-07-15

    Recent studies show that myosin light chain kinase (MLCK) plays a pivotal role in development of cerebral edema, a known complication following traumatic brain injury (TBI) in children and a contributing factor to worsened neurologic recovery. Interferon-stimulated gene 15 (ISG15) is upregulated after cerebral ischemia and is neuroprotective. The significant role of ISG15 after TBI has not been studied. Postnatal Day (PND) 21 and PND24 mice were subjected to lateral closed-skull injury with impact depth of 2.0 or 2.25?mm. Behavior was examined at 7?d using two-object novel recognition and Wire Hang tests. Mice were sacrificed at 6?h, 12?h, 24?h, 48?h, 72?h, and 7?d. ISG15 and MLCK were analyzed by Western blot and immunohistochemistry, blood-brain barrier (BBB) disruption with Evans Blue (EB), and cerebral edema with wet/dry weights. EB extravasation and edema peaked at 72?h in both ages. PND21 mice had more severe neurological deficits, compared with PND24 mice. PND24 mice showed peak ISG15 expression at 6?h, and PND21 mice at 72?h. MLCK peaked in both age groups at 12?h and co-localized with ISG15 on immunohistochemistry and co-immunoprecipitation. These studies provide evidence, ISG15 is elevated following TBI in mice, preceding MLCK elevation, development of BBB disruption, and cerebral edema. PMID:25669448

  14. Antiaging gene Klotho enhances glucose-induced insulin secretion by up-regulating plasma membrane levels of TRPV2 in MIN6 ?-cells.

    PubMed

    Lin, Yi; Sun, Zhongjie

    2012-07-01

    Klotho is a recently discovered antiaging gene. Klotho is expressed in mouse pancreatic islets and in insulinoma ?-cells (MIN6 ?-cells). The purpose of this study was to investigate whether Klotho plays a role in the regulation of insulin secretion in MIN6 ?-cells by overexpression and silencing of Klotho. It is interesting that overexpression of Klotho increased glucose-induced insulin secretion in MIN6 ?-cells. Overexpression of mouse Klotho protein also significantly increased plasma membrane levels of transient receptor potential V2 (TRPV2), calcium entry, and the glucose-induced increase in intracellular calcium. On the other hand, knockdown of Klotho by siRNA significantly decreased plasma membrane levels of TRPV2 and attenuated glucose-induced calcium entry and insulin secretion. Tranilast, a selective inhibitor of TRPV2, abolished the promoting effects of overexpression of Klotho on glucose-induced calcium entry and insulin secretion in MIN6 cells. Thus, TRPV2 lies in the downstream of Klotho in the regulation of glucose-induced insulin secretion. This study demonstrated, for the first time, that Klotho may enhance glucose-induced insulin secretion by up-regulating plasma membrane levels of TRPV2 and thus glucose-induced calcium responses. These findings reveal a previously unidentified role of Klotho in the regulation of glucose-induced insulin secretion in MIN6 ?-cells. PMID:22597535

  15. Increasing of temperature induces pathogenicity of Streptococcus agalactiae and the up-regulation of inflammatory related genes in infected Nile tilapia (Oreochromis niloticus).

    PubMed

    Kayansamruaj, Pattanapon; Pirarat, Nopadon; Hirono, Ikuo; Rodkhum, Channarong

    2014-08-01

    Temperature strongly affects the health of aquatic poikilotherms. In Nile tilapia (Oreochromis niloticus), elevated water temperatures increase the severity of streptococcosis. Here we investigated the effects of temperature on the vulnerability and inflammatory response of Nile tilapia to Streptococcus agalactiae (Group B streptococci; GBS). At 35 and 28 °C, GBS took 4 and 7h, respectively to reach the log-phase and, when incubated with tilapia whole blood, experienced survival rates of 97% and 2%, respectively. The hemolysis activity of GBS grown at 35 °C was five times higher than that of GBS grown at 28 °C. GBS expressed cylE (?-hemolysin/cytolysin), cfb (CAMP factor) and PI-2b (pili-backbone) much more strongly at 35 °C than at 28 °C. Challenging Nile tilapia reared at 35 and 28 °C with GBS resulted in accumulated mortalities of about 85% and 45%, respectively. At 35 °C, infected tilapia exhibited tremendous inflammatory responses due to a dramatic up-regulation (30-40-fold) of inflammatory-related genes (cyclooxygenase-2, IL-1? and TNF-?) between 6 and 96 h-post infection. These results suggest that the increase of GBS pathogenicity to Nile tilapia induced by elevated temperature is associated with massive inflammatory responses, which may lead to acute mortality. PMID:24856132

  16. Characterization and expression analysis of the transferrin gene in Nile tilapia (Oreochromis niloticus) and its upregulation in response to Streptococcus agalactiae infection.

    PubMed

    Poochai, Watsida; Choowongkomon, Kiattawee; Srisapoome, Prapansak; Unajak, Sasimanas; Areechon, Nontawith

    2014-10-01

    In this study, full-length tilapia transferrin (OnTF) isolated from liver cDNA of Nile tilapia (Oreochromis niloticus) was found to have an open reading frame of 2,091-bp encoding 696 amino acid residues. Two additional amino acids: Gly(369) and Gly(370) were observed compared with the reported Nile tilapia transferrin protein sequence. Pre-mature protein has a predicted molecular weight of 78.2 kDa, while mature protein is 73.28 kDa in size. Comparative sequence analysis with transferrin from other species revealed two major putative iron-binding domains designated as the N-lobe and the C-lobe in accordance with the transferrin protein characteristics. The predicted tertiary structure of tilapia transferrin confirmed the presence of iron and anion-binding sites on both lobes that are conserved among transferrins from other species. Quantitative real-time PCR analysis showed significantly higher expression of tilapia transferrin gene in liver than in other tissues (p < 0.05). Transferrin expression in tilapia experimentally infected with 10(6) and 10(8) colony-forming units mL(-1) of Streptococcus agalactiae was significantly upregulated at 24 and 12 h post-infection (hpi), respectively, and decreased afterward. Iron-deficiency in serum of bacterially infected fish was detected at 48 and 24 hpi, respectively. The expression pattern of the transferrin gene and the iron levels of infected tilapia in this study were consistent with the function of transferrin in innate immunity. PMID:24770882

  17. Aspirin influences megakaryocytic gene expression leading to up-regulation of multidrug resistance protein-4 in human platelets

    PubMed Central

    Massimi, Isabella; Guerriero, Raffaella; Lotti, Lavinia Vittoria; Lulli, Valentina; Borgognone, Alessandra; Romani, Federico; Barillà, Francesco; Gaudio, Carlo; Gabbianelli, Marco; Frati, Luigi; Pulcinelli, Fabio M

    2014-01-01

    Aim The aim of the study was to investigate whether human megakaryocytic cells have an adaptive response to aspirin treatment, leading to an enhancement of multidrug resistance protein-4 (MRP4) expression in circulating platelets responsible for a reduced aspirin action. We recently found that platelet MRP4 overexpression has a role in reducing aspirin action in patients after by-pass surgery. Aspirin enhances MRP4-mRNA levels in rat liver and drug administration transcriptionally regulates MRP4 gene expression through peroxisome proliferator-activated receptor-? (PPAR?). Methods The effects induced by aspirin or PPAR? agonist (WY14643) on MRP4 modulation were evaluated in vitro in a human megakaryoblastic DAMI cell line, in megakaryocytes (MKs) and in platelets obtained from human haematopoietic progenitor cell (HPC) cultures, and in vivo platelets obtained from aspirin treated healthy volunteers (HV). Results In DAMI cells, aspirin and WY14643 treatment induced a significant increase in MRP4 and PPAR? expression. In human MKs grown in the presence of either aspirin or WY14643, MRP4 and PPAR?-mRNA were higher than in control cultures and derived platelets showed an enhancement in MRP4 protein expression. The ability of aspirin to modulate MRP4 expression in MKs and to transfer it to platelets was also confirmed in vivo. In fact, we found the highest MRP4 mRNA and protein expression in platelets obtained from HV after 15 days' aspirin treatment. Conclusions The present study provides evidence, for the first time, that aspirin treatment affects the platelet protein pattern through MK genomic modulation. This work represents an innovative and attractive approach, useful both to identify patients less sensitive to aspirin and to improve pharmacological treatment in cardiovascular high-risk patients. PMID:24902864

  18. Dnmt3a1 Upregulates Transcription of Distinct Genes and Targets Chromosomal Gene Clusters for Epigenetic Silencing in Mouse Embryonic Stem Cells ?

    PubMed Central

    Kotini, Andriana G.; Mpakali, Anastasia; Agalioti, Theodora

    2011-01-01

    Dnmt3a1 and Dnmt3a2 are two de novo DNA methyltransferases expressed in mouse embryonic stem cells (mESCs). They differ in that a 219-amino-acid (aa) amino (N)-terminal noncatalytic domain is present only in Dnmt3a1. Here, we examined the unique functions of Dnmt3a1 in mESCs by targeting the coding sequence of the Dnmt3a1 N-terminal domain tagged with enhanced green fluorescent protein (GFP) for insertion into the mouse Rosa26 locus. Using these targeted cells (GFP-3a1Nter), we showed that Dnmt3a1 was efficiently recruited to the silenced Oct3/4 and activated Vtn (vitronectin) gene promoters via its unique N-terminal domain. This recruitment affected the two genes in contrasting ways, compromising Oct3/4 gene promoter DNA methylation to prevent consolidation of the silent state while significantly reducing Vtn transcription. We used this negative effect of the Dnmt3a1 N-terminal domain to investigate the extent of transcriptional regulation by Dnmt3a1 in mESCs by using microarrays. A small group of all-trans retinoic acid (tRA)-inducible genes had lower transcript levels in GFP-3a1Nter cells than in wild-type mESCs. Intriguingly, this group included genes that are important for fetal nutrition, placenta development, and metabolic functions and is enriched for a distinct set of imprinted genes. We also identified a larger group of genes that showed higher transcript levels in the GFP-3a1Nter-expressing cells than in wild-type mESCs, including pluripotency factors and key regulators of primordial germ cell differentiation. Thus, Dnmt3a1 in mESCs functions primarily as a negative and to a lesser extent as a positive regulator of transcription. Our findings suggest that Dnmt3a1 positively affects transcription of specific genes at the promoter level and targets chromosomal domains to epigenetically silence gene clusters in mESCs. PMID:21262766

  19. Massive bowel resection upregulates the intestinal mRNA expression levels of cellular retinol-binding protein II and apolipoprotein A-IV and alters the intestinal vitamin A status in rats.

    PubMed

    Hebiguchi, Taku; Mezaki, Yoshihiro; Morii, Mayako; Watanabe, Ryo; Yoshikawa, Kiwamu; Miura, Mitsutaka; Imai, Katsuyuki; Senoo, Haruki; Yoshino, Hiroaki

    2015-03-01

    Short bowel (SB) syndrome causes the malabsorption of various nutrients. Among these, vitamin A is important for a number of physiological activities. Vitamin A is absorbed by epithelial cells of the small intestine and is discharged into the lymphatic vessels as a component of chylomicrons and is delivered to the liver. In the present study, we used a rat model of SB syndrome in order to assess its effects on the expression of genes associated with the absorption, transport and metabolism of vitamin A. In the rats with SB, the intestinal mRNA expression levels of cellular retinol-binding protein II (CRBP II, gene symbol Rbp2) and apolipoprotein A-IV (gene symbol Apoa4) were higher than those in the sham-operated rats, as shown by RT-qPCR. Immunohistochemical analysis revealed that absorptive epithelial cells stained positive for both CRBP II and lecithin retinol acyltransferase, which are both required for the effective esterification of vitamin A. In the rats with SB, the retinol content in the ileum and the retinyl ester content in the jejunum were lower than those in the sham-operated rats, as shown by quantitative analysis of retinol and retinyl esters by high performance liquid chromatography. These results suggest that the elevated mRNA expression levels of Rbp2 and Apoa4 in the rats with SB contribute to the effective esterification and transport of vitamin A. PMID:25585692

  20. L-Arginine ameliorates cardiac left ventricular oxidative stress by upregulating eNOS and Nrf2 target genes in alloxan-induced hyperglycemic rats

    SciTech Connect

    Ramprasath, Tharmarajan; Hamenth Kumar, Palani; Syed Mohamed Puhari, Shanavas; Senthil Murugan, Ponniah; Vasudevan, Varadaraj [Molecular Cardiology Unit, Department of Biochemistry, Center for Excellence in Genomic Sciences, School of Biological Sciences, Madurai Kamaraj University, Madurai 625 021, Tamilnadu (India)] [Molecular Cardiology Unit, Department of Biochemistry, Center for Excellence in Genomic Sciences, School of Biological Sciences, Madurai Kamaraj University, Madurai 625 021, Tamilnadu (India); Selvam, Govindan Sadasivam, E-mail: drselvamgsbiochem@rediffmail.com [Molecular Cardiology Unit, Department of Biochemistry, Center for Excellence in Genomic Sciences, School of Biological Sciences, Madurai Kamaraj University, Madurai 625 021, Tamilnadu (India)

    2012-11-23

    Highlights: Black-Right-Pointing-Pointer L-Arginine treatment reduced the metabolic disturbances in diabetic animals. Black-Right-Pointing-Pointer Antioxidant marker proteins were found high in myocardium by L-arginine treatment. Black-Right-Pointing-Pointer Elevated antioxidant status, mediates the reduced TBA-reactivity in left ventricle. Black-Right-Pointing-Pointer L-Arginine treatment enhanced the Nrf2 and eNOS signaling in left ventricle. Black-Right-Pointing-Pointer Improved cell survival signaling by arginine, offers a novel tactic for targeting. -- Abstract: Hyperglycemia is independently related with excessive morbidity and mortality in cardiovascular disorders. L-Arginine-nitric oxide (NO) pathway and the involvement of NO in modulating nuclear factor-E2-related factor-2 (Nrf2) signaling were well established. In the present study we investigated, whether L-arginine supplementation would improve the myocardial antioxidant defense under hyperglycemia through activation of Nrf2 signaling. Diabetes was induced by alloxan monohydrate (90 mg kg{sup -1} body weight) in rats. Both non-diabetic and diabetic group of rats were divided into three subgroups and they were administered either with L-arginine (2.25%) or L-NAME (0.01%) in drinking water for 12 days. Results showed that L-arginine treatment reduced the metabolic disturbances in diabetic rats. Antioxidant enzymes and glutathione levels were found to be increased in heart left ventricles, thereby reduction of lipid peroxidation by L-arginine treatment. Heart histopathological analysis further validates the reversal of typical diabetic characteristics consisting of alterations in myofibers and myofibrillary degeneration. qRT-PCR studies revealed that L-arginine treatment upregulated the transcription of Akt and downregulated NF-{kappa}B. Notably, transcription of eNOS and Nrf2 target genes was also upregulated, which were accompanied by enhanced expression of Nrf2 in left ventricular tissue from diabetic and control rats. Under these findings, we suggest that targeting of eNOS and Nrf2 signaling by L-arginine supplementation could be used as a potential treatment method to alleviate the late diabetic complications.

  1. Spatial association of apoptosis-related gene expression and cellular death in clinical neuroblastoma.

    PubMed Central

    Hoehner, J. C.; Gestblom, C.; Olsen, L.; PÃ¥hlman, S.

    1997-01-01

    Several unique features of neuroblastoma (NB), including the capacity for spontaneous regression and maturation to benign pathology, suggest that genes that regulate cellular proliferation, survival and differentiation may be involved in directing clinical tumour aggressiveness. The in situ expression of Bcl-2, Rb, p21, p53 and Bax proteins, as well as the proliferation marker proliferating cell nuclear antigen (PCNA) were examined immunocytochemically in a selection of 38 stage- and outcome-identified NB tumours. Apoptotic cells were identified morphologically and by a DNA fragmentation labelling technique (TUNEL). Although the tumour cell density of Bcl-2, p53, Bax, PCNA and TUNEL positivity correlated with patient survival, a spatially organized expression pattern was further recognized in stroma-poor differentiating tumours. Immature tumour cells adjacent to thin fibrovascular stroma are proliferating, as evidenced by PCNA positivity, and often express Bcl-2. At increasing distance from this fibrovascular stroma, intermediately differentiated tumour cells express Rb, while with more advanced differentiation, proliferation ceases and Bcl-2 immunoreactivity is lost. The most differentiated tumour cells, which often express p53, and occasionally p21 and Bax, lie adjacent to TUNEL-positive, morphologically apoptotic cells. This spatial organization in favourable outcome NB tumours suggests that physiological regulation of differentiation and apoptosis may be involved in tumour regression. Images Figure 2 PMID:9099968

  2. Upregulation of genes for C-reactive protein and related pentraxin/complement proteins in photodynamic therapy-treated human tumor cells: enrolment of PI3K/Akt and AP-1.

    PubMed

    Merchant, Soroush; Korbelik, Mladen

    2013-06-01

    Treatment of mouse tumors by photodynamic therapy (PDT) was reported to trigger the production of serum amyloid P component (SAP), a prototypic acute phase reactant in the mouse, that occurs in the targeted tumor as well as distant sites dominated by host's liver. It was also shown that the SAP gene becomes upregulated and protein produced in mouse tumor cells treated by PDT in vitro. Present study revealed that, in addition to SAP, increased expression of genes encoding related pentraxin and complement proteins, including PTX3, C1q and ficolin B, can be found in mouse LLC tumor cells treated by PDT. Since in humans C-reactive protein (CRP) is more important acute phase reactant than SAP, the expression of gene encoding this pentraxin protein was examined in human lung tumor A549 cells treated by PDT. The results demonstrated a PDT dose-dependent upregulation of CRP gene, as well as of PTX3 and ficolin 1 genes in these cells. Investigation into the signal transduction process underlying PDT-induced human CRP gene upregulation using specific inhibitors of critical signaling elements revealed critical role played by PI3K/Akt pathway. Downstream DNA transcription factor largely responsible for this increased CRP gene expression is AP-1 with possible cooperation of HIF-1. It was suggested that cells sensing to have sustained a mortal injury from PDT can turn on molecular programs ensuring that the disposal of their corpses (facilitated by CRP and related pentraxin and complement components) is swift and efficient. PMID:23182717

  3. Identification of SNPs in Cellular Retinol Binding Protein 1 and Cellular Retinol Binding Protein 3 Genes and Their Associations with Laying Performance Traits in Erlang Mountainous Chicken

    PubMed Central

    Wang, Yan; Xiao, Li-Hua; Zhao, Xiao-Ling; Liu, Yi-Ping; Zhu, Qing

    2014-01-01

    CRBP1 (cellular retinol binding protein 1) and CRBP3 (cellular retinol binding protein 3), are important components of the retinoid signaling pathway and take part in vitamin A absorption, transport and metabolism. Based on the role of vitamin A in chicken laying performance, we investigated the polymorphism of CRBP1 and CRBP3 genes in 349 chickens using single strand conformation polymorphism and DNA sequencing methods. Only one polymorphism was identified in the third intron of CRBP1, two polymorphisms were detected in CRBP3; they were located in the second intron and the third intron respectively. The association studies between these three SNPs and laying performance traits were performed in Erlang mountainous chicken. Notably, the SNP g.14604G>T of CRBP1 was shown to be significantly associated with body weight at first egg (BWFE), age at first egg (AFE), weight at first egg (WFE) and total number of eggs with 300 age (EN). The CRBP3 polymorphism g.934C>G was associated with AFE, and the g.1324A>G was associated with AFE and BWFE, but none of these polymorphisms were associated with egg quality traits. Haplotype combinations constructed on these two SNPs of CRBP3 gene were associated with BWFE and AFE. In particular, diplotype H2H2 had positive effect on AFE, BWFE, EN, and average egg-laying interval. We herein describe for the first time basic research on the polymorphism of chicken CRBP1 and CRBP3 genes that is predictive of genetic potential for laying performance in chicken. PMID:25083100

  4. HMOX1 and NQO1 Genes are Upregulated in Response to Contact Sensitizers in Dendritic Cells and THP1 Cell Line: Role of the Keap1\\/Nrf2 Pathway

    Microsoft Academic Search

    Nadege Ade; Fanny Leon; Marc Pallardy; Jean-Luc Peiffer; Saadia Kerdine-Romer; Marie-Helene Tissier; Pierre-Antoine Bonnet; Isabelle Fabre; Jean-Claude Ourlin

    2009-01-01

    Electrophilicity is one of the most common features of skin contact sensitizers and is necessary for protein haptenation. The Keap1 (Kelch-like ECH-associated protein 1)\\/Nrf2 -signaling pathway is dedicated to the detection of electrophilic stress in cells leading to the upregulation of genes involved in protection or neutralization of chemical reactive species. Signals provided by chemical stress could play an important

  5. CD84 is markedly up-regulated in Kawasaki disease arteriopathy

    PubMed Central

    Reindel, R; Bischof, J; Kim, K-Y A; Orenstein, J M; Soares, M B; Baker, S C; Shulman, S T; Perlman, E J; Lingen, M W; Pink, A J; Trevenen, C; Rowley, A H

    2014-01-01

    The major goals of Kawasaki disease (KD) therapy are to reduce inflammation and prevent thrombosis in the coronary arteries (CA), but some children do not respond to currently available non-specific therapies. New treatments have been difficult to develop because the molecular pathogenesis is unknown. In order to identify dysregulated gene expression in KD CA, we performed high-throughput RNA sequencing on KD and control CA, validated potentially dysregulated genes by real-time reverse transcription–polymerase chain reaction (RT–PCR) and localized protein expression by immunohistochemistry. Signalling lymphocyte activation molecule CD84 was up-regulated 16-fold (P < 0·01) in acute KD CA (within 2 months of onset) and 32-fold (P < 0·01) in chronic CA (5 months to years after onset). CD84 was localized to inflammatory cells in KD tissues. Genes associated with cellular proliferation, motility and survival were also up-regulated in KD CA, and immune activation molecules MX2 and SP140 were up-regulated in chronic KD. CD84, which facilitates immune responses and stabilizes platelet aggregates, is markedly up-regulated in KD CA in patients with acute and chronic arterial disease. We provide the first molecular evidence of dysregulated inflammatory responses persisting for months to years in CA significantly damaged by KD. PMID:24635044

  6. The hepatitis B virus X protein increases the cellular level of TATA-binding protein, which mediates transactivation of RNA polymerase III genes

    SciTech Connect

    Wang, Horng-Dar; Johnson, D.L. [Univ. of Southern California, Los Angeles, CA (United States); Yuh, Chio-Hwa [California Institute of Technology, Pasadena, CA (United States)] [and others

    1995-12-01

    This report decribes the mechanism by which the hepatitis B virus X gene product induces RNA polymerase III genes. The RNA pol III transcription system serves as model for understanding the mechanism of X in the transactivation of cellular genes in both Drosophila and rat cell lines. 53 refs., 7 figs., 1 tab.

  7. Up-regulation of type II collagen gene by 17?-estradiol in articular chondrocytes involves Sp1/3, Sox-9, and estrogen receptor ?

    PubMed

    Maneix, Laure; Servent, Aurélie; Porée, Benoît; Ollitrault, David; Branly, Thomas; Bigot, Nicolas; Boujrad, Noureddine; Flouriot, Gilles; Demoor, Magali; Boumediene, Karim; Moslemi, Safa; Galéra, Philippe

    2014-08-01

    The existence of a link between estrogen deprivation and osteoarthritis (OA) in postmenopausal women suggests that 17?-estradiol (17?-E2) may be a modulator of cartilage homeostasis. Here, we demonstrate that 17?-E2 stimulates, via its receptor human estrogen receptor ? 66 (hER?66), type II collagen expression in differentiated and dedifferentiated (reflecting the OA phenotype) articular chondrocytes. Transactivation of type II collagen gene (COL2A1) by ligand-independent transactivation domain (AF-1) of hER?66 was mediated by "GC" binding sites of the -266/-63-bp promoter, through physical interactions between ER?, Sp1/Sp3, Sox9, and p300, as demonstrated in chromatin immunoprecipitation (ChIP) and Re-Chromatin Immuno-Precipitation (Re-ChIP) assays in primary and dedifferentiated cells. 17?-E2 and hER?66 increased the DNA-binding activities of Sp1/Sp3 and Sox-9 to both COL2A1 promoter and enhancer regions. Besides, Sp1, Sp3, and Sox-9 small interfering RNAs (siRNAs) prevented hER?66-induced transactivation of COL2A1, suggesting that these factors and their respective cis-regions are required for hER?66-mediated COL2A1 up-regulation. Our results highlight the genomic pathway by which 17?-E2 and hER?66 modulate Sp1/Sp3 heteromer binding activity and simultaneously participate in the recruitment of the essential factors Sox-9 and p300 involved respectively in the chondrocyte-differentiated status and COL2A1 transcriptional activation. These novel findings could therefore be attractive for tissue engineering of cartilage in OA, by the fact that 17?-E2 could promote chondrocyte redifferentiation. PMID:25081415

  8. Quantum Dots in an Amphiphilic Polyethyleneimine Derivative Platform for Cellular Labeling, Targeting, Gene Delivery, and Ratiometric Oxygen Sensing.

    PubMed

    Park, Joonhyuck; Lee, Junhwa; Kwag, Jungheon; Baek, Yeonggyeong; Kim, Bumju; Yoon, Calvin Jinse; Bok, Seoyeon; Cho, So-Hye; Kim, Ki Hean; Ahn, G-One; Kim, Sungjee

    2015-06-23

    Amphiphilic polyethyleneimine derivatives (amPEIs) were synthesized and used to encapsulate dozens of quantum dots (QDs). The QD-amPEI composite was ?100 nm in hydrodynamic diameter and had the slightly positive outer surface that suited well for cellular internalization. The QD-amPEI showed very efficient QD cellular labeling with the labeled cell fluorescence intensity more than 10 times higher than conventional techniques such as Lipofectamine-assisted QD delivery. QD-amPEI was optimal for maximal intracellular QD delivery by the large QD payload and the rapid endocytosis kinetics. QD-amPEI platform technology was demonstrated for gene delivery, cell-specific labeling, and ratiometric oxygen sensing. Our QD-amPEI platform has two partitions: positive outer surface and hydrophobic inside pocket. The outer positive surface was further exploited for gene delivery and targeting. Co-delivery of QDs and GFP silencing RNAs was successfully demonstrated by assembling siRNAs to the outer surfaces, which showed the transfection efficiency an order of magnitude higher than conventional gene transfections. Hyaluronic acids were tethered onto the QD-amPEI for cell-specific targeted labeling which showed the specific-to-nonspecific signal ratio over 100. The inside hydrophobic compartment was further applied for cohosting oxygen sensing phosphorescence Ru dyes along with QDs. The QD-Ru-amPEI oxygen probe showed accurate and reversible oxygen sensing capability by the ratiometric photoluminescence signals, which was successfully applied to cellular and spheroid models. PMID:26057729

  9. B?cell translocation 1 gene inhibits cellular metastasis?associated behavior in breast cancer.

    PubMed

    Li, Wei; Zou, Shi-Tao; Zhu, Ran; Wan, Jian-Mei; Xu, Yan; Wu, Hao-Rong

    2014-06-01

    B-cell translocation gene 1 (BTG1) is a member of the BTG/transducer of ERBB2 family, which regulates cell cycle progression in a variety of cell types and may have a role in inhibiting proliferation, promoting apoptosis and stimulating cellular differentiation in numerous cell types. However, the role of BTG1 in cancer metastasis is yet to be elucidated. In the present study, analysis of clinical specimens revealed that BTG1 mRNA levels were lower in lymph node metastases than those in benign breast tumors and normal human breast tissue. The effect of BTG1 on the metastatic behavior of breast cancer cells following stable transfection with a BTG1 expression vector was also investigated. The overexpression of BTG1 was observed to inhibit cell adhesion, migration and invasion. Furthermore, the overexpression of BTG1 was found to be involved in the inhibition of the metastasis-related proteins matrix metalloproteinase-2 and -9, as well as the promotion of the cell-cell adhesion-associated protein E-cadherin. In syngeneic nude mice breast tumor models, hepatic metastasis and angiogenesis were observed in the mice injected with the control cells, but not in those injected with pcDNA3-BTG1 cells. Immunohistochemistry revealed that overexpression of BTG1 decreased vascular endothelial growth factor expression in tumors. To the best of our knowledge, this is the first study to show that BTG1 overexpression decreases migration and invasion of breast cancer cells and thereby inhibits distant metastasis in mice breast tumor models. PMID:24714932

  10. Pteromalus puparum venom impairs host cellular immune responses by decreasing expression of its scavenger receptor gene.

    PubMed

    Fang, Qi; Wang, Lei; Zhu, Yangkeng; Stanley, David W; Chen, Xuexin; Hu, Cui; Ye, Gongyin

    2011-11-01

    Insect host/parasitoid interactions are co-evolved systems in which host defenses are balanced by parasitoid mechanisms to disable or hide from host immune effectors. Although there is a rich literature on these systems, parasitoid immune-disabling mechanisms have not been fully elucidated. Here we report on a newly discovered immune-disabling mechanism in the Pieris rapae/Pteromalus puparum host/parasitoid system. Because venom injections and parasitization suppresses host phagocytosis, we turned attention to the P. rapae scavenger receptor (Pr-SR), posing the hypothesis that P. puparum venom suppresses expression of the host Pr-SR gene. To test our hypothesis, we cloned a full-length cDNA of the Pr-SR. Multiple sequences alignment showed the deduced amino acid sequence of Pr-SR is similar to scavenger receptors of other lepidopterans. Bacterial and bead injections induced Pr-SR mRNA and protein expression, which peaked at 4h post-bead injection. Venom injection inhibited Pr-SR expression. Pr-SR was specifically expressed in granulocytes compared to plasmatocytes. We localized the Pr-SR protein in cytoplasm and cellular membrane, with no evidence of secretion into host plasma. Double-strand RNA designed to Pr-SR mRNA silenced expression of Pr-SR and significantly impaired host phagocytosis and encapsulation reactions. Venom injections similarly silenced Pr-SR expression during the first 8h post-treatment, after which the silencing effects gradually abated. We infer from these findings that one mechanism of impairing P. rapae hemocytic immune reactions is by silencing expression of Pr-SR. PMID:21802512

  11. Modulation of Enhancer Looping and Differential Gene Targeting by Epstein-Barr Virus Transcription Factors Directs Cellular Reprogramming

    PubMed Central

    McClellan, Michael J.; Wood, C. David; Ojeniyi, Opeoluwa; Cooper, Tim J.; Kanhere, Aditi; Arvey, Aaron; Webb, Helen M.; Palermo, Richard D.; Harth-Hertle, Marie L.; Kempkes, Bettina; Jenner, Richard G.; West, Michelle J.

    2013-01-01

    Epstein-Barr virus (EBV) epigenetically reprogrammes B-lymphocytes to drive immortalization and facilitate viral persistence. Host-cell transcription is perturbed principally through the actions of EBV EBNA 2, 3A, 3B and 3C, with cellular genes deregulated by specific combinations of these EBNAs through unknown mechanisms. Comparing human genome binding by these viral transcription factors, we discovered that 25% of binding sites were shared by EBNA 2 and the EBNA 3s and were located predominantly in enhancers. Moreover, 80% of potential EBNA 3A, 3B or 3C target genes were also targeted by EBNA 2, implicating extensive interplay between EBNA 2 and 3 proteins in cellular reprogramming. Investigating shared enhancer sites neighbouring two new targets (WEE1 and CTBP2) we discovered that EBNA 3 proteins repress transcription by modulating enhancer-promoter loop formation to establish repressive chromatin hubs or prevent assembly of active hubs. Re-ChIP analysis revealed that EBNA 2 and 3 proteins do not bind simultaneously at shared sites but compete for binding thereby modulating enhancer-promoter interactions. At an EBNA 3-only intergenic enhancer site between ADAM28 and ADAMDEC1 EBNA 3C was also able to independently direct epigenetic repression of both genes through enhancer-promoter looping. Significantly, studying shared or unique EBNA 3 binding sites at WEE1, CTBP2, ITGAL (LFA-1 alpha chain), BCL2L11 (Bim) and the ADAMs, we also discovered that different sets of EBNA 3 proteins bind regulatory elements in a gene and cell-type specific manner. Binding profiles correlated with the effects of individual EBNA 3 proteins on the expression of these genes, providing a molecular basis for the targeting of different sets of cellular genes by the EBNA 3s. Our results therefore highlight the influence of the genomic and cellular context in determining the specificity of gene deregulation by EBV and provide a paradigm for host-cell reprogramming through modulation of enhancer-promoter interactions by viral transcription factors. PMID:24068937

  12. PpCBF3 from Cold-Tolerant Kentucky Bluegrass Involved in Freezing Tolerance Associated with Up-Regulation of Cold-Related Genes in Transgenic Arabidopsis thaliana

    PubMed Central

    Chen, Yu; Xu, Bin; Yang, Zhimin; Huang, Bingru

    2015-01-01

    Dehydration-Responsive Element Binding proteins (DREB)/C-repeat (CRT) Binding Factors (CBF) have been identified as transcriptional activators during plant responses to cold stress. The objective of this study was to determine the physiological roles of a CBF gene isolated from a cold-tolerant perennial grass species, Kentucky bluegrass (Poa pratensis L.), which designated as PpCBF3, in regulating plant tolerance to freezing stress. Transient transformation of Arabidopsis thaliana mesophyll protoplast with PpCBF3-eGFP fused protein showed that PpCBF3 was localized to the nucleus. RT-PCR analysis showed that PpCBF3 was specifically induced by cold stress (4°C) but not by drought stress [induced by 20% polyethylene glycol 6000 solution (PEG-6000)] or salt stress (150 mM NaCl). Transgenic Arabidopsis overexpressing PpCBF3 showed significant improvement in freezing (-20°C) tolerance demonstrated by a lower percentage of chlorotic leaves, lower cellular electrolyte leakage (EL) and H2O2 and O2.- content, and higher chlorophyll content and photochemical efficiency compared to the wild type. Relative mRNA expression level analysis by qRT-PCR indicated that the improved freezing tolerance of transgenic Arabidopsis plants overexpressing PpCBF3 was conferred by sustained activation of downstream cold responsive (COR) genes. Other interesting phenotypic changes in the PpCBF3-transgenic Arabidopsis plants included late flowering and slow growth or ‘dwarfism’, both of which are desirable phenotypic traits for perennial turfgrasses. Therefore, PpCBF3 has potential to be used in genetic engineering for improvement of turfgrass freezing tolerance and other desirable traits. PMID:26177510

  13. Expression of lipogenic genes is upregulated in the heart with exercise training-induced but not pressure overload-induced left ventricular hypertrophy.

    PubMed

    Dobrzyn, Pawel; Pyrkowska, Aleksandra; Duda, Monika K; Bednarski, Tomasz; Maczewski, Michal; Langfort, Jozef; Dobrzyn, Agnieszka

    2013-06-15

    Cardiac hypertrophy is accompanied by molecular remodeling that affects different cellular pathways, including fatty acid (FA) utilization. In the present study, we show that cardiac lipid metabolism is differentially regulated in response to physiological (endurance training) and pathological [abdominal aortic banding (AAB)] hypertrophic stimuli. Physiological hypertrophy was accompanied by an increased expression of lipogenic genes and the activation of sterol regulatory element-binding protein-1c and Akt signaling. Additionally, FA oxidation pathways regulated by AMP-activated protein kinase (AMPK) and peroxisome proliferator activated receptor-? (PPAR?) were induced in trained hearts. Cardiac lipid content was not changed by physiological stimulation, underlining balanced lipid utilization in the trained heart. Moreover, pathological hypertrophy induced the AMPK-regulated oxidative pathway, whereas PPAR? and expression of its downstream targets, i.e., acyl-CoA oxidase and carnitine palmitoyltransferase I, were not affected by AAB. In contrast, pathological hypertrophy leads to cardiac triglyceride (TG) and diacylglycerol (DAG) accumulation, although the expression of lipogenic genes and the levels of FA transport proteins (CD36 and FATP) were not changed or reduced compared with the sham group. A possible explanation for this phenomenon is a decrease in lipolysis, as evidenced by the increased content of adipose triglyceride lipase inhibitor G0S2, the increased phosphorylation of hormone-sensitive lipase at Ser(565), and the decreased protein levels of DAG lipase that attenuate TG and DAG contents. The increased TG and DAG accumulation observed in AAB-induced hypertrophy might have lipotoxic effects, thereby predisposing to cardiomyopathy and heart failure in the future. PMID:23632628

  14. HMOX1 and NQO1 genes are upregulated in response to contact sensitizers in dendritic cells and THP-1 cell line: role of the Keap1/Nrf2 pathway.

    PubMed

    Ade, Nadège; Leon, Fanny; Pallardy, Marc; Peiffer, Jean-Luc; Kerdine-Romer, Saadia; Tissier, Marie-Hélène; Bonnet, Pierre-Antoine; Fabre, Isabelle; Ourlin, Jean-Claude

    2009-02-01

    Electrophilicity is one of the most common features of skin contact sensitizers and is necessary for protein haptenation. The Keap1 (Kelch-like ECH-associated protein 1)/Nrf2 -signaling pathway is dedicated to the detection of electrophilic stress in cells leading to the upregulation of genes involved in protection or neutralization of chemical reactive species. Signals provided by chemical stress could play an important role in dendritic cell activation and the aim of this work was to test whether contact sensitizers were specific activators of the Keap1/Nrf2 pathway. CD34-derived dendritic cells (CD34-DC) and the THP-1 myeloid cell line were treated by a panel of sensitizers (Ni, 1-chloro 2,4-dinitrobenzene, cinnamaldehyde, 7-hydroxycitronellal, 1,4-dihydroquinone, alpha-methyl-trans-cinnamaldehyde, 2-4-tert-(butylbenzyl)propionaldehyde or Lilial, and 1,4-phenylenediamine), irritants (sodium dodecyl sulfate, benzalkonium chloride), and a nonsensitizer molecule (chlorobenzene). Three well-known Nrf2 activators (tert-butylhydroquinone, lipoic acid, sulforaphane) were also tested. Expression of hmox1 and nqo1 was measured using real-time PCR and cellular accumulation of Nrf2 was assessed by Western blot. Our results showed an increased expression at early time points of hmox1 and nqo1 mRNAs in response to sensitizers but not to irritants. Accumulation of the Nrf2 protein was also observed only with chemical sensitizers. A significant inhibition of the expression of hmox1 and nqo1 mRNAs and CD86 expression was found in 1-chloro 2,4-dinitrobenzene-treated THP-1 cells preincubated with N-acetyl cysteine, a glutathione precursor. Altogether, these data suggested that the Keap1/Nrf2-signaling pathway was activated by electrophilic molecules including sensitizers in dendritic cells and in the THP-1 cell line. Monitoring of this pathway may provide new biomarkers (e.g., Nrf2, hmox1) for the detection of the sensitization potential of chemicals. PMID:19033392

  15. Delineation and interpretation of gene networks towards their effect in cellular physiology- A reverse engineering approach for the identification of critical molecular players, through the use of ontologies

    Microsoft Academic Search

    K. Moutselos; I. Maglogiannis; A. Chatziioannou

    2010-01-01

    Exploiting ontologies, provides clues regarding the involvement of certain molecular processes in the cellular phenotypic manifestation. However, identifying individual molecular actors (genes, proteins, etc.) for targeted biological validation in a generic, prioritized, fashion, based in objective measures of their effects in the cellular physiology, remains a challenge. In this work, a new meta-analysis algorithm is proposed for the holistic interpretation

  16. Bacterial diversity among small-subunit rRNA gene clones and cellular isolates from the same seawater sample.

    PubMed Central

    Suzuki, M T; Rappé, M S; Haimberger, Z W; Winfield, H; Adair, N; Ströbel, J; Giovannoni, S J

    1997-01-01

    Numerous investigations applying the cloning and sequencing of rRNA genes (rDNAs) to the study of marine bacterioplankton diversity have shown that the sequences of genes cloned directly from environmental DNA do not correspond to the genes of cultured marine taxa. These results have been interpreted as support for the hypothesis that the most abundant heterotrophic marine bacterioplankton species are not readily culturable by commonly used methods. However, an alternative explanation is that marine bacterioplankton can be easily cultured but are not well represented in sequence databases. To further examine this question, we compared the small-subunit (SSU) rDNAs of 127 cellular clones isolated from a water sample collected off the Oregon coast to 58 bacterial SSU rDNAs cloned from environmental DNAs from the same water sample. The results revealed little overlap between partial SSU rDNA sequences from the cellular clones and the environmental clone library. An exception was the SSU rDNA sequence recovered from a cellular clone belonging to the Pseudomonas subgroup of the gamma subclass of the class Proteobacteria, which was related to a single gene cloned directly from the same water sample (OCS181) (similarity, 94.6%). In addition, partial SSU rDNA sequences from three of the cultured strains matched a novel rDNA clone related to the gamma subclass of the Proteobacteria found previously in an environmental clone library from marine aggregates (AGG53) (similarity, 94.3 to 99.6%). Our results support the hypothesis that many of the most abundant bacterioplankton species are not readily culturable by standard methods but also show that heterotrophic bacterioplankton that are culturable on media with high organic contents include many strains for which SSU rDNA sequences are not available in sequence databases. PMID:9055415

  17. Expression of Novel Gene Products Upregulated by Disuse is Normalized by an Osteogenic Mechanical Stimulus: Evidence for the Molecular Basis of a Low Level Biomechanical Countermeasure for Osteoporosis?

    NASA Technical Reports Server (NTRS)

    Rubin, C.; Zhi, J.; Xu, G.; Cute, M.; McLeod, K.; Hadjiargyrou, M.

    1999-01-01

    The National Research Council's report entitled: A Strategy for Space Biology and Medical Science, highlighted several areas of fundamental scientific investigation which must be addressed to make long-term space exploration not only feasible, but safe. This "Goldberg Strategy," as well as several subsequent reports published by the NRC's Space Studies Board (e.g., Assessment of Programs in Space Biology and Medicine, Smith et. al., 1991), suggests that the principal hurdle to man's extended presence in space is the osteopenia which parallels reduced gravity. Ironically, the most significant risk to the skeleton may only be realized on return to normal gravitational fields, and full recovery of bone mass may never occur. Effective counter-measures to this microgravity induced bone loss are thus essential. Considering the similarities of space and aging induced osteopenia, an indisputable benefit of such a prophylaxis would be its potential as a treatment for the bone loss which plagues over 25 million people in the U.S. The osteogenic potential of mechanical strain is strongly frequency dependent, with sensitivity increasing up through at least 60 Hz (cycles per second). One hundred seconds per day of a 1 Hz cyclic loading will inhibit disuse osteopenia only if sufficient in magnitude to engender 1000 microstrain (mu(epsilon)) in the tissue. When loading is applied at 30 Hz, however, mechanical strains on the order of 5O mu(epsilon) (approx. 1% of the peak strains which occur in bone during vigorous functional activity), can stimulate bone formation in a duration dependent manner. In longer term animal studies, strains of less than 10 mu(epsilon), induced non-invasively via a whole body vibration, will stimulate bone formation on the surfaces of trabeculae, increase bone density, and improve strength. Finally, preliminary results from a double blind prospective clinical trial shows promise in inhibiting the bone loss which parallels the menopause. Based on these observations, we propose that these high frequency, low magnitude, mechanical strains effectively serve as a "surrogate" for musculoskeletal ground reaction forces, and thus represent an ideal countermeasure to the osteopenia which parallels microgravity conditions. The specific goal of this NASA funded work is to identify genes in bone upregulated by disuse, and to determine the efficacy of an osteogenic mechanical stimulus to downregulate their expression.

  18. Cellular Gene Expression Profiles in Rhesus Macaques Challenged Mucosally with a Pathogenic R5 Tropic Simian Human Immunodeficiency Virus Isolate

    PubMed Central

    Pise-Masison, Cynthia A.; Radonovich, Michael F.; Brady, John; Lee, Jae K.; Cheon, Soo-Young; Markham, Phillip; Cristillo, Anthony; Pal, Ranajit

    2008-01-01

    Abstract Insights into the host factors that contribute to an effective antiviral immune response may be obtained by examining global gene expression in simian human immunodeficiency virus (SHIV)-infected nonhuman primates that exhibit different virological outcomes. Immune responses and gene expression profiles in peripheral blood mononuclear cells (PBMCs) were compared between animals that controlled or did not control viremia after infection. Rectal inoculation of eight rhesus macaques with R5-tropic SHIVSF162P3 resulted in a high level of plasma viremia during the acute phase of infection. The viremia was controlled to below levels of detection in six of these animals at the set point (controllers), whereas two animals had persistent viremia throughout the 140 wk that the animals were monitored (non-controllers). CD4+ T-cell counts declined slightly in both controllers and non-controllers in the acute phase of infection, but CD4+ T-cell counts continued to decline only in the non-controllers. Neutralizing antibodies to the challenge virus were variable and could not account for the control of viremia. However, analysis of the cellular gene expression profiles in the PBMCs from both groups of animals revealed distinctive gene expression patterns between controllers and non-controllers. Using the paired LPE test, 59 genes with p values <0.01 were identified and specific differences in the gene expression profiles in PBMCs from controllers versus non-controllers were detected. PMID:19115930

  19. A digital framework to build, visualize and analyze a gene expression atlas with cellular resolution in zebrafish early embryogenesis.

    PubMed

    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; Peyriéras, Nadine; Santos, Andrés

    2014-06-01

    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

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

    PubMed Central

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

    2012-01-01

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

  1. A Targeted UAS-RNAi Screen in Drosophila Larvae Identifies Wound Closure Genes Regulating Distinct Cellular Processes

    PubMed Central

    Lesch, Christine; Jo, Juyeon; Wu, Yujane; Fish, Greg S.; Galko, Michael J.

    2010-01-01

    Robust mechanisms for tissue repair are critical for survival of multicellular organisms. Efficient cutaneous wound repair requires the migration of cells at the wound edge and farther back within the epidermal sheet, but the genes that control and coordinate these migrations remain obscure. This is in part because a systematic screening approach for in vivo identification and classification of postembryonic wound closure genes has yet to be developed. Here, we performed a proof-of-principle reporter-based in vivo RNAi screen in the Drosophila melanogaster larval epidermis to identify genes required for normal wound closure. Among the candidate genes tested were kinases and transcriptional mediators of the Jun N-terminal kinase (JNK) signaling pathway shown to be required for epithelial sheet migration during development. Also targeted were genes involved in actin cytoskeletal remodeling. Importantly, RNAi knockdown of both canonical and noncanonical members of the JNK pathway caused open wounds, as did several genes involved in actin cytoskeletal remodeling. Our analysis of JNK pathway components reveals redundancy among the upstream activating kinases and distinct roles for the downstream transcription factors DJun and DFos. Quantitative and qualitative morphological classification of the open wound phenotypes and evaluation of JNK activation suggest that multiple cellular processes are required in the migrating epidermal cells, including functions specific to cells at the wound edge and others specific to cells farther back within the epidermal sheet. Together, our results identify a new set of conserved wound closure genes, determine putative functional roles for these genes within the migrating epidermal sheet, and provide a template for a broader in vivo RNAi screen to discover the full complement of genes required for wound closure during larval epidermal wound healing. PMID:20813879

  2. Maximizing gene delivery efficiencies of cationic helical polypeptides via balanced membrane penetration and cellular targeting

    E-print Network

    Cheng, Jianjun

    Maximizing gene delivery efficiencies of cationic helical polypeptides via balanced membrane: Received 26 July 2013 Accepted 24 September 2013 Available online 7 November 2013 Keywords: Non-viral gene a c t The application of non-viral gene delivery vectors is often accompanied with the poor

  3. A Novel Vitamin D-Regulated Immediate-Early Gene, IEX-1, Alters Cellular Growth and Apoptosis

    PubMed Central

    Pittelkow, Mark R.; Salisbury, Jeffrey L.; Grande, Joseph R; Im, Hee-Jeong; Feldmann, Kathrin A.; Schilling, David

    2010-01-01

    1?,25-Dihydroxyvitamin D3 (1?,25(OH)2D3) inhibits the expression of an immediate-early gene, IEX-1, which is involved in the regulation of cellular growth and apoptosis in a variety of cells. 1?,25(OH)2D3 alters the subcellular localization of IEX-1 by causing an efflux of IEX-1 from the nucleus, and the sterol decreases the expression of IEX-1 messenger RNA in cells via a novel DR3 repeat-type DNA response element. PMID:12899517

  4. Exposure of a 23F Serotype Strain of Streptococcus pneumoniae to Cigarette Smoke Condensate Is Associated with Selective Upregulation of Genes Encoding the Two-Component Regulatory System 11 (TCS11)

    PubMed Central

    Herbert, Jenny A.; Mitchell, Timothy J.; Dix-Peek, Thérèse; Dickens, Caroline; Anderson, Ronald; Feldman, Charles

    2014-01-01

    Alterations in whole genome expression profiles following exposure of the pneumococcus (strain 172, serotype 23F) to cigarette smoke condensate (160??g/mL) for 15 and 60?min have been determined using the TIGR4 DNA microarray chip. Exposure to CSC resulted in the significant (P < 0.014–0.0006) upregulation of the genes encoding the two-component regulatory system 11 (TCS11), consisting of the sensor kinase, hk11, and its cognate response regulator, rr11, in the setting of increased biofilm formation. These effects of cigarette smoke on the pneumococcus may contribute to colonization of the airways by this microbial pathogen. PMID:25013815

  5. Cloning, expression and cellular localization of the Doublesex gene in the water flea, Daphnia carinata, during different developmental stages.

    PubMed

    Zhang, Mingqing; Li, Haixia; Liu, Ajing; Wu, Donglei; Wang, Danli; Zhao, Yunlong

    2014-10-25

    In this study, one of Doublesex genes from the common freshwater cladoceran Daphnia carinata, designated DapcaDsx1, was cloned using primers based on homologous sequences and rapid amplification of cDNA ends (RACE). qPCR was employed to quantify differences in DapcaDsx1 expression between the different sexual phases, with expression levels being higher in sexual females. The role of DapcaDsx1 in the reproductive transformation was further investigated in parthenogenetic-phase females and sexual-phase females using whole-mount in situ hybridization. This cellular localization study showed specific expression of DapcaDsx1 in the thoracic segments, second antenna and part of the ventral carapace. Higher expression levels were exhibited in sexual females compared to parthenogenetic females. This suggests that the DapcaDsx1 gene plays significant roles in switching modes of reproduction and during sexual differentiation. PMID:25130908

  6. MicroRNAs Regulate Cellular ATP Levels by Targeting Mitochondrial Energy Metabolism Genes during C2C12 Myoblast Differentiation.

    PubMed

    Siengdee, Puntita; Trakooljul, Nares; Murani, Eduard; Schwerin, Manfred; Wimmers, Klaus; Ponsuksili, Siriluck

    2015-01-01

    In our previous study, we identified an miRNA regulatory network involved in energy metabolism in porcine muscle. To better understand the involvement of miRNAs in cellular ATP production and energy metabolism, here we used C2C12 myoblasts, in which ATP levels increase during differentiation, to identify miRNAs modulating these processes. ATP level, miRNA and mRNA microarray expression profiles during C2C12 differentiation into myotubes were assessed. The results suggest 14 miRNAs (miR-423-3p, miR-17, miR-130b, miR-301a/b, miR-345, miR-15a, miR-16a, miR-128, miR-615, miR-1968, miR-1a/b, and miR-194) as cellular ATP regulators targeting genes involved in mitochondrial energy metabolism (Cox4i2, Cox6a2, Ndufb7, Ndufs4, Ndufs5, and Ndufv1) during C2C12 differentiation. Among these, miR-423-3p showed a high inverse correlation with increasing ATP levels. Besides having implications in promoting cell growth and cell cycle progression, its function in cellular ATP regulation is yet unknown. Therefore, miR-423-3p was selected and validated for the function together with its potential target, Cox6a2. Overexpression of miR-423-3p in C2C12 myogenic differentiation lead to decreased cellular ATP level and decreased expression of Cox6a2 compared to the negative control. These results suggest miR-423-3p as a novel regulator of ATP/energy metabolism by targeting Cox6a2. PMID:26010876

  7. MicroRNAs Regulate Cellular ATP Levels by Targeting Mitochondrial Energy Metabolism Genes during C2C12 Myoblast Differentiation

    PubMed Central

    Siengdee, Puntita; Trakooljul, Nares; Murani, Eduard; Schwerin, Manfred; Wimmers, Klaus; Ponsuksili, Siriluck

    2015-01-01

    In our previous study, we identified an miRNA regulatory network involved in energy metabolism in porcine muscle. To better understand the involvement of miRNAs in cellular ATP production and energy metabolism, here we used C2C12 myoblasts, in which ATP levels increase during differentiation, to identify miRNAs modulating these processes. ATP level, miRNA and mRNA microarray expression profiles during C2C12 differentiation into myotubes were assessed. The results suggest 14 miRNAs (miR-423-3p, miR-17, miR-130b, miR-301a/b, miR-345, miR-15a, miR-16a, miR-128, miR-615, miR-1968, miR-1a/b, and miR-194) as cellular ATP regulators targeting genes involved in mitochondrial energy metabolism (Cox4i2, Cox6a2, Ndufb7, Ndufs4, Ndufs5, and Ndufv1) during C2C12 differentiation. Among these, miR-423-3p showed a high inverse correlation with increasing ATP levels. Besides having implications in promoting cell growth and cell cycle progression, its function in cellular ATP regulation is yet unknown. Therefore, miR-423-3p was selected and validated for the function together with its potential target, Cox6a2. Overexpression of miR-423-3p in C2C12 myogenic differentiation lead to decreased cellular ATP level and decreased expression of Cox6a2 compared to the negative control. These results suggest miR-423-3p as a novel regulator of ATP/energy metabolism by targeting Cox6a2. PMID:26010876

  8. TNF-{alpha} upregulates the A{sub 2B} adenosine receptor gene: The role of NAD(P)H oxidase 4

    SciTech Connect

    St Hilaire, Cynthia; Koupenova, Milka; Carroll, Shannon H.; Smith, Barbara D. [Department of Biochemistry, Whitaker Cardiovascular Institute, Boston University School of Medicine, 715 Albany Street, Boston, MA 02118 (United States); Ravid, Katya [Department of Biochemistry, Whitaker Cardiovascular Institute, Boston University School of Medicine, 715 Albany Street, Boston, MA 02118 (United States)], E-mail: ravid@biochem.bumc.bu.edu

    2008-10-24

    Proliferation of vascular smooth muscle cells (VSMC), oxidative stress, and elevated inflammatory cytokines are some of the components that contribute to plaque formation in the vasculature. The cytokine tumor necrosis factor-alpha (TNF-{alpha}) is released during vascular injury, and contributes to lesion formation also by affecting VSMC proliferation. Recently, an A{sub 2B} adenosine receptor (A{sub 2B}AR) knockout mouse illustrated that this receptor is a tissue protector, in that it inhibits VSMC proliferation and attenuates the inflammatory response following injury, including the release of TNF-{alpha}. Here, we show a regulatory loop by which TNF-{alpha} upregulates the A{sub 2B}AR in VSMC in vitro and in vivo. The effect of this cytokine is mimicked by its known downstream target, NAD(P)H oxidase 4 (Nox4). Nox4 upregulates the A{sub 2B}AR, and Nox inhibitors dampen the effect of TNF-{alpha}. Hence, our study is the first to show that signaling associated with Nox4 is also able to upregulate the tissue protecting A{sub 2B}AR.

  9. PIKfyve upregulates CFTR activity.

    PubMed

    Gehring, Eva-Maria; Lam, Rebecca S; Siraskar, Gulab; Koutsouki, Evgenia; Seebohm, Guiscard; Ureche, Oana N; Ureche, Liviu; Baltaev, Ravshan; Tavare, Jeremy M; Lang, Florian

    2009-12-18

    The cystic fibrosis transmembrane conductance regulator (CFTR) is a cAMP-activated Cl(-) channel critically important in Cl(-) secreting epithelia. Mutations in the CFTR gene, such as (DeltaF508)CFTR leads to cystic fibrosis, a severe disease with defective Cl(-) secretion. CFTR is stimulated by the serum and glucocorticoid-inducible kinase SGK1. The SGK1 dependent regulation of several carriers and channels involves the phosphatidylinositol-3-phosphate-5-kinase PIKfyve, which similarly mediates the regulation of glucose carriers by PKB/Akt. The present study was thus performed to elucidate whether PKB/Akt and PIKfyve are regulators of CFTR. To this end CFTR or (DeltaF508)CFTR were expressed in Xenopus oocytes alone or together with PKB, PIKfyve or the SGK1/PKB resistant mutant (S318A)PIKfyve, and the current generated by cAMP upregulation with 10muM forskolin+1mM IBMX determined utilizing dual electrode voltage clamp. As a result, forskolin/IBMX treatment triggered a current (I(cAMP)) in CFTR-expressing Xenopus oocytes, but not in oocytes expressing (DeltaF508)CFTR. Coexpression of PKB/Akt and PIKfyve, but not of (S318A)PIKfyve, stimulated I(cAMP) in CFTR-expressing ( approximately 2- to 3-fold) but not in (DeltaF508)CFTR-expressing or water injected Xenopus oocytes. Immunohistochemistry revealed that the coexpression of PIKfyve, but not of (S318A)PIKfyve, enhanced the CFTR protein abundance but not the (DeltaF508)CFTR protein abundance in CFTR or (DeltaF508)CFTR-expressing oocytes. The present observations reveal a novel powerful regulator of intact but not of defective CFTR. PMID:19852935

  10. Upregulation of genes related to bone formation by ?-amino butyric acid and ?-oryzanol in germinated brown rice is via the activation of GABAB-receptors and reduction of serum IL-6 in rats

    PubMed Central

    Muhammad, Sani Ismaila; Maznah, Ismail; Mahmud, Rozi; Zuki, Abu Bakar Zakaria; Imam, Mustapha Umar

    2013-01-01

    Background Osteoporosis and other bone degenerative diseases are among the most challenging non-communicable diseases to treat. Previous works relate bone loss due to osteoporosis with oxidative stress generated by free radicals and inflammatory cytokines. Alternative therapy to hormone replacement has been an area of interest to researchers for almost three decades due to hormone therapy-associated side effects. Methods In this study, we investigated the effects of gamma-amino butyric acid (GABA), gamma-oryzanol (ORZ), acylated steryl glucosides (ASG), and phenolic extracts from germinated brown rice (GBR) on the expression of genes related to bone metabolism, such as bone morphogenic protein-2 (BMP-2), secreted protein acidic and rich in cysteine (SPARC), runt-related transcription factor 2 (RUNX-2), osteoblast-specific transcription factor osterix (Osx), periostin, osteoblast specific factor (Postn), collagen 1&2 (Col1&2), calcitonin receptor gene (CGRP); body weight measurement and also serum interleukin-6 (IL-6) and osteocalcin, in serum and bone. Rats were treated with GBR, ORZ, GABA, and ASG at (100 and 200 mg/kg); estrogen (0.2 mg/kg), or remifemin (10 and 20 mg/kg), compared to ovariectomized non-treated group as well as non-ovariectomized non-treated (sham) group. Enzyme-linked immunosorbent assay was used to measure the IL-6 and osteocalcin levels at week 2, 4, and 8, while the gene expression in the bone tissue was determined using the Genetic Analysis System (Beckman Coulter Inc., Brea, CA, USA). Results The results indicate that groups treated with GABA (100 and 200 mg/kg) showed significant upregulation of SPARC, calcitonin receptor, and BMP-2 genes (P < 0.05), while the ORZ-treated group (100 and 200 mg/kg) revealed significant (P < 0.05) upregulation of Osx, Postn, RUNX-2, and Col1&2. Similarly, IL-6 concentration decreased, while osteocalcin levels increased significantly (P < 0.05) in the treated groups as compared to ovariectomized non-treated groups. Conclusion GABA and ORZ from GBR stimulates osteoblastogenesis by upregulation of bone formation genes, possibly via the activation of GABAB receptors and by inhibiting the activity of inflammatory cytokines and reactive oxygen species. Therefore, it could be used effectively in the management of osteoporosis. PMID:24098073

  11. Upregulation of Opioid Receptors

    Microsoft Academic Search

    Ellen M. Unterwald; Richard D. Howells

    \\u000a It is well established that chronic exposure to opioid receptor antagonists can result in opioid receptor upregulation. The\\u000a phenomenon of antagonist-induced receptor upregulation is not unique to the opioid system but is common to many receptor systems\\u000a including adenergic, cholinergic, serotinergic, and dopaminergic receptors. Chronic administration of naloxone or naltrexone\\u000a reliably produces increases in binding to opioid receptors both in

  12. Contribution of Viral Mimics of Cellular Genes to KSHV Infection and Disease

    PubMed Central

    Sakakibara, Shuhei; Tosato, Giovanna

    2014-01-01

    Kaposi’s sarcoma-associated herpesvirus (KSHV, also named Human herpesvirus 8 HHV-8) is the cause of Kaposi sarcoma (KS), the most common malignancy in HIV-infected individuals worldwide, primary effusion lymphoma (PEL) and multicentric Castleman disease (MCD). KSHV is a double-stranded DNA virus that encodes several homologues of cellular proteins. The structural similarity between viral and host proteins explains why some viral homologues function as their host counterparts, but sometimes at unusual anatomical sites and inappropriate times. In other cases, structural modification in the viral proteins can suppress or override the function of the host homologue, contributing to KSHV-related diseases. For example, viral IL-6 (vIL-6) is sufficiently different from human IL-6 to activate gp130 signaling independent of the ? subunit. As a consequence, vIL-6 can activate many cell types that are unresponsive to cellular IL-6, contributing to MCD disease manifestations. Here, we discuss the molecular biology of KSHV homologues of cellular products as conduits of virus/host interaction with a focus on identifying new strategies for therapy of KS and other KSHV-related diseases. PMID:25243371

  13. Integrated Cellular and Gene Interaction Model for Cell Migration in Embryonic Development

    E-print Network

    Song, Joe

    of biological phenomena, including animal coats [12], human brain development [1], and gene regulatory expressions behave regarding each other as an integrated system is a profound question in biology. Intuitively

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

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

  15. BAC-recombineering for studying plant gene regulation: developmental control and cellular localization of SnRK1 kinase subunits.

    PubMed

    Bitrián, Marta; Roodbarkelari, Farshad; Horváth, Mihály; Koncz, Csaba

    2011-03-01

    Recombineering, permitting precise modification of genes within bacterial artificial chromosomes (BACs) through homologous recombination mediated by lambda phage-encoded Red proteins, is a widely used powerful tool in mouse, Caenorhabditis and Drosophila genetics. As Agrobacterium-mediated transfer of large DNA inserts from binary BACs and TACs into plants occurs at low frequency, recombineering is so far seldom exploited in the analysis of plant gene functions. We have constructed binary plant transformation vectors, which are suitable for gap-repair cloning of genes from BACs using recombineering methods previously developed for other organisms. Here we show that recombineering facilitates PCR-based generation of precise translational fusions between coding sequences of fluorescent reporter and plant proteins using galK-based exchange recombination. The modified target genes alone or as part of a larger gene cluster can be transferred by high-frequency gap-repair into plant transformation vectors, stably maintained in Agrobacterium and transformed without alteration into plants. Versatile application of plant BAC-recombineering is illustrated by the analysis of developmental regulation and cellular localization of interacting AKIN10 catalytic and SNF4 activating subunits of Arabidopsis Snf1-related (SnRK1) protein kinase using in vivo imaging. To validate full functionality and in vivo interaction of tagged SnRK1 subunits, it is demonstrated that immunoprecipitated SNF4-YFP is bound to a kinase that phosphorylates SnRK1 candidate substrates, and that the GFP- and YFP-tagged kinase subunits co-immunoprecipitate with endogenous wild type AKIN10 and SNF4. PMID:21235649

  16. Rapid Up-Regulation of HKT1, a High-Affinity Potassium Transporter Gene, in Roots of Barley and Wheat following Withdrawal of Potassium1

    PubMed Central

    Wang, Tie-Bang; Gassmann, Walter; Rubio, Francisco; Schroeder, Julian I.; Glass, Anthony D.M.

    1998-01-01

    High-affinity K+ uptake in plant roots is rapidly up-regulated when K+ is withheld and down-regulated when K+ is resupplied. These processes make important contributions to plant K+ homeostasis. A cDNA coding for a high-affinity K+ transporter, HKT1, was earlier cloned from wheat (Triticum aestivum L.) roots and functionally characterized. We demonstrate here that in both barley (Hordeum vulgare L.) and wheat roots, a rapid and large up-regulation of HKT1 mRNA levels resulted when K+ was withdrawn from growth media. This effect was specific for K+; withholding N caused a modest reduction of HKT1 mRNA levels. Up-regulation of HKT1 transcript levels in barley roots occurred within 4 h of removing K+, which corresponds to the documented increase of high-affinity K+ uptake in roots following removal of K+. Increased expression of HKT1 mRNA was evident before a decline in total root K+ concentration could be detected. Resupply of 1 mm K+ was sufficient to strongly reduce HKT1 transcript levels. In wheat root cortical cells, both membrane depolarizations in response to 100 ?m K+, Cs+, and Rb+, and high-affinity K+ uptake were enhanced by K+ deprivation. Thus, in both plant systems the observed physiological changes associated with manipulating external K+ supply were correlated with levels of HKT1 mRNA expression. Implications of these findings for K+ sensing and regulation of the HKT1 mRNA levels in plant roots are discussed. PMID:9765551

  17. A method for cellular localization of gene expression via quantitative in situ hybridization in plants.

    PubMed

    Küpper, Hendrik; Seib, Laura Ort; Sivaguru, Mayandi; Hoekenga, Owen A; Kochian, Leon V

    2007-04-01

    A quantitative in situ hybridization technique (quantitative whole-mount in situ hybridization, QISH) for plants is described. It employs direct hybridization of fluorescently labelled gene-specific oligonucleotides in large tissue pieces combined with optical sectioning. It dramatically increases the throughput compared with conventional antibody- and microtome-based in situ mRNA hybridization methods, while simultaneously eliminating artefact-prone preparation steps that prevent reliable quantification in conventional methods. The key feature of this technique is the quantification of gene expression using housekeeping genes (cytosolic GAPDH and 18S RNA) as internal standards. This feature enables a correction of varying cytoplasm/vacuole ratios in different cell types, as well as tissue optical effects and non-specific signals. The quantitative nature of the technique allows for analysis of gene expression in response to different environmental conditions, as well as tissue- and age-dependent differences in gene expression patterns. In addition to testing tissue permeabilization, structural preservation, specificity, linearity and tissue optical effects, we verified the reliability of the technique with three Arabidopsis thaliana genes of known function and distribution. These were the rbcL gene for ribulose 1,5-bisphosphate carboxylase, the developmentally related gene SCARECROW (AtSCR) and PHOT-1, a photoreceptor kinase. As expected, rbcL mRNA was found in all photosynthetic cells, while SCR mRNA was detected mainly in bundle sheath cells and PHOT-1 was found predominantly in epidermal and cortical cells of the apical hook of light-grown seedlings. As an application example, QISH was used to measure transcript abundance for a zinc transporter from the ZIP family of transporters in the Zn/Cd hyperaccumulator model plant, Thlaspi caerulescens, and the related non-accumulator Thlaspi arvense. This showed that QISH can be used to compare differences in mRNA levels between cell types, plant growth conditions and plant species. Messenger RNA for the zinc transporter gene ZNT1 was abundant in photosynthetic cells, but not in the epidermal storage cells where metal hyperaccumulation in T. caerulescens occurs. This indicates that ZNT1 does not directly participate in metal hyperaccumulation within the leaf. Growing T. caerulescens with high zinc levels strongly reduced ZNT1 transcript abundance in the spongy mesophyll cells, but less in the other cell types. In T. arvense, ZNT1 mRNA levels were generally much lower, and were furthermore drastically reduced by growth at increased zinc levels, confirming earlier reports regarding ZNT1 regulation in these two Thlaspi species. PMID:17397510

  18. Upregulation of the immune protein gene hemolin in the epidermis during the wandering larval stage of the Indian meal moth, Plodia interpunctella.

    PubMed

    Aye, Tin Tin; Shim, Jae-Kyoung; Rhee, In-Koo; Lee, Kyeong-Yeoll

    2008-08-01

    Expression of hemolin, which generates an immune protein, was up-regulated in wandering fifth instar larval stage of Plodia interpunctella. The mRNA level peaked in the middle of the wandering stage. Major expression was in the epidermis, rather than in the fat body or gut. To test a possible ecdysteroid effect on hemolin induction we treated with RH-5992, an ecdysteroid agonist, and KK-42, which inhibits ecdysteroid biosynthesis in both feeding and wandering fifth instar larvae. When feeding larvae were treated with RH-5992 the hemolin mRNA level was increased. When wandering larvae were treated with KK-42 its level was reduced. In addition, when KK-42-treated larvae were subsequently treated with RH-5992 the hemolin mRNA level was recovered. These results strongly suggest that ecdysteroid up-regulates the expression of hemolin mRNA. Hormonal and bacterial effects on hemolin induction were further analyzed at the tissue level. Major induction of hemolin mRNA was detected following both RH-5992 treatment and bacterial injection in the epidermis of both feeding and wandering larvae. Minor induction of hemolin was detected in the fat body following a bacterial injection, but not RH-5992 treatment. We infer that in P. interpunctella larvae, the epidermis is the major tissue for hemolin induction in naïve insects and in insects manipulated with bacterial and hormonal treatments. PMID:18675821

  19. Blue light-promoted rice leaf bending and unrolling are due to up-regulated brassinosteroid biosynthesis genes accompanied by accumulation of castasterone.

    PubMed

    Asahina, Masashi; Tamaki, Yuji; Sakamoto, Tomoaki; Shibata, Kyomi; Nomura, Takahito; Yokota, Takao

    2014-08-01

    In this study the relationship between blue light- and brassinosteroid-enhanced leaf lamina bending and unrolling in rice was investigated. Twenty-four hours (h) irradiation with white or blue light increased endogenous brassinosteroid levels, especially those of typhasterol and castasterone, in aerial tissues of rice seedlings. There was an accompanying up-regulation of transcript levels of CYP85A1/OsDWARF, encoding an enzyme catalyzing C-6 oxidation, after 6h under either white or blue light. These effects were not observed in seedlings placed under far-red or red light regimes. It was concluded that blue light up-regulates the levels of several cytochrome P450 enzymes including CYP85A1, thereby promoting the synthesis of castasterone, a biologically active brassinosteroid in rice. Based on these findings, it is considered that blue light-mediated rice leaf bending and unrolling are consequences of the enhanced biosynthesis of endogenous castasterone. In contrast to aerial tissues, brassinosteroid synthesis in roots appeared to be negatively regulated by white, blue and red light but positively controlled by far-red light. PMID:24856112

  20. Upregulation of human telomerase reverse transcriptase mRNA expression by in vitro transfection of hepatitis B virus X gene into human hepatocarcinoma and cholangiocarcinoma cells

    Microsoft Academic Search

    Zhen-Liang Qu; Sheng-Quan Zou; Nai-Qiang Cui; Xian-Zhong Wu; Ming-Fang Qin; Zhen-Li Zhou

    2005-01-01

    Abstract Abstract Abstract Abstract AIM: To study the changes of human telomerase reverse transcriptase (hTERT) mRNA expression in human hepatocarcinoma cell lines (HepG2) and cholangiocarcinoma cell lines (QBC939) after HBx gene transfection and to illustrate the significance of transcriptional regulation of hTERT gene by HBx gene in the carcinogenesis. METHODS: HepG2 and QBC939 cell lines were cultured and co-transfected with

  1. Kinetochore genes are coordinately up-regulated in human tumors as part of a FoxM1-related cell division program

    E-print Network

    Thiru, Prathapan

    The key player in directing proper chromosome segregation is the macromolecular kinetochore complex, which mediates DNA–microtubule interactions. Previous studies testing individual kinetochore genes documented examples ...

  2. Upregulated, 7q21-22 amplicon candidate gene SHFM1 confers oncogenic advantage by suppressing p53 function in gastric cancer.

    PubMed

    Tamilzhalagan, Sembulingam; Muthuswami, Muthulakshmi; Periasamy, Jayaprakash; Lee, Ming Hui; Rha, Sun Young; Tan, Patrick; Ganesan, Kumaresan

    2015-06-01

    Chromosomal aberrations are hallmarks of cancers and the locus of frequent genomic amplifications often harbors key cancer driver genes. Many genomic amplicons remain larger with hundreds of genes and the key drivers remain to be identified by an amplification-wide systematic analysis. The 7q21.12-q22.3 genomic amplification is frequent in gastric cancers which occur in ~10% of the patients and multiple cell lines. This 7q21.12-q22.3 amplicon has not yet been completely analyzed towards identifying the driver genes and their functional contribution in oncogenesis. The amplitude and prevalence indicate the important role conferred by this amplicon in gastric cancers. Among the 159 genes of this amplicon, 12 genes are found over-expressed in primary gastric tumors and cell lines. Many of the over-expressed genes show negative association with p53 transcriptional activity. RNAi based functional screening of the genes reveal, SHFM1 as key gastric cancer driver gene. SHFM1 confers cell cycle progression and resistance to p53 stabilizing drugs in gastric cancer cells. SHFM1 also activates Src, MAPK/ERK and PI3K/Akt signaling pathways. This is the first integrative genomic investigation of 7q21.12-q22.3 amplicon revealing the potential oncogenic candidacy of 12 genes. The oncogenic contribution of SHFM1, mediated by the p53 suppressive feature has been demonstrated in gastric cancer cells. PMID:25697906

  3. Regulation of Gene Expression and Cellular Localization of Cloned Klebsiella aerogenes (K. pneumoniae) Urease

    Microsoft Academic Search

    SCOTT B. MULROONEY; H. S. PANKRATZ; ROBERT P. HAUSINGER

    1989-01-01

    The genes for Klebsiella aerogenes (K. pneumoniae) urease were cloned and the protein was overexpressed (up to 18% of total protein consisted of this enzyme) in several hosts. The small size of the DNA encoding urease (3.5 kb), the restriction map, and the regulation of enzyme expression directed by the recombinant plasmid are distinct from other cloned ureases. Nickel concentration

  4. Cellular Pharmacology and Molecular Biology of the Trabecular Meshwork Inducible glucocorticoid Response Gene Product

    Microsoft Academic Search

    Jon R. Polansky; Don J. Fauss; Pu Chen; Hua Chen; Elke Lütjen-Drecoll; Douglas Johnson; Ron M. Kurtz; Zhi-Dong Ma; Ernest Bloom; Thai D. Nguyen

    1997-01-01

    Studies of the effects of glucocorticoid (GC) and oxidative stress stimuli in differentiated cultures of human trabecular meshwork (HTM) cells have provided the rationale for our studies of a major new gene termed TIGR (trabecular meshwork inducible GC response). The TIGR clone was isolated by differential library screening using selection criteria based on the induction pattern of a new protein\\/glycoprotein

  5. Retrovolution: HIV–Driven Evolution of Cellular Genes and Improvement of Anticancer Drug Activation

    PubMed Central

    Rossolillo, Paola; Winter, Flore; Simon-Loriere, Etienne; Gallois-Montbrun, Sarah; Negroni, Matteo

    2012-01-01

    In evolution strategies aimed at isolating molecules with new functions, screening for the desired phenotype is generally performed in vitro or in bacteria. When the final goal of the strategy is the modification of the human cell, the mutants selected with these preliminary screenings may fail to confer the desired phenotype, due to the complex networks that regulate gene expression in higher eukaryotes. We developed a system where, by mimicking successive infection cycles with HIV-1 derived vectors containing the gene target of the evolution in their genome, libraries of gene mutants are generated in the human cell, where they can be directly screened. As a proof of concept we created a library of mutants of the human deoxycytidine kinase (dCK) gene, involved in the activation of nucleoside analogues used in cancer treatment, with the aim of isolating a variant sensitizing cancer cells to the chemotherapy compound Gemcitabine, to be used in gene therapy for anti-cancer approaches or as a poorly immunogenic negative selection marker for cell transplantation approaches. We describe the isolation of a dCK mutant, G12, inducing a 300-fold sensitization to Gemcitabine in cells originally resistant to the prodrug (Messa 10K), an effect 60 times stronger than the one induced by the wt enzyme. The phenotype is observed in different tumour cell lines irrespective of the insertion site of the transgene and is due to a change in specificity of the mutated kinase in favour of the nucleoside analogue. The mutations characterizing G12 are distant from the active site of the enzyme and are unpredictable on a rational basis, fully validating the pragmatic approach followed. Besides the potential interest of the G12 dCK variant for therapeutic purposes, the methodology developed is of interest for a large panel of applications in biotechnology and basic research. PMID:22927829

  6. Association of Angiotensin-Converting Enzyme (ACE) Gene Polymorphism with Inflammation and Cellular Cytotoxicity in Vitiligo Patients

    PubMed Central

    Hasan, Nermeen; Zahra, Amr; Fayez, Salwa

    2015-01-01

    Background Vitiligo is a disorder with profound heterogeneity in its aetio-pathophysiology. Angiotensin converting enzyme (ACE) plays an important role in the physiology of the vasculature, blood pressure and inflammation. An insertion/deletion (I/D) polymorphism of the ACE gene was reported be associated with the development of vitiligo. Objective Our aim was to evaluate the ACE I/D polymorphism in vitiligo patients and controls. Our second aim was to find a possible association between ACE gene polymorphism and inflammatory mediators (as interleukin (IL)-6) and/or cellular cytotoxicity induced by serum nitrite (as a breakdown product of the cytotoxic nitric oxide) in vitiligo patients. Methods This case-control study included 74 vitiligo patients and 75 apparently healthy controls. The distribution of ACE gene I/D genotype was investigated using PCR. Serum ACE, IL-6 and nitrite were measured by colorimetric method, ELISA and Griess assay respectively. Results The ACE allele frequency was significantly different between vitiligo patients and healthy controls (P = 0.026). However there was no significant difference between the ACE genotyping frequency in both groups (P = 0.115). There were statistically significant higher VIDA score (P = 0.007), and serum IL-6 (P < 0.001) in patients with the DD genotype when compared to other genotypes. Serum nitrite in patients with the DD genotype was significantly higher (P = 0.007) when compared to patients with II genotype. Serum levels of ACE, IL-6 and nitrite in vitiligo patients were statistically significantly higher than those in controls. Conclusion As a conclusion, ACE gene polymorphism might grant susceptibility to develop vitiligo. Serum IL-6 and nitrite levels might have an important role in the pathogenesis of vitiligo. Targeting these two factors might have an implication in the treatment of some resistant cases. PMID:26177100

  7. The Tumor Suppressor Gene Brca1 Is Required for Embryonic Cellular Proliferation in the Mouse

    Microsoft Academic Search

    Razqallah Hakem; José Luis de la Pompa; Christian Sirard; Rong Mo; Minna Woo; Anne Hakem; Andrew Wakeham; Julia Potter; Armin Reitmair; Filio Billia; Eduardo Firpo; Chi Chung Hui; Jim Roberts; Janet Rossant; Tak W Mak

    1996-01-01

    Mutations of the BRCA1 gene in humans are associated with predisposition to breast and ovarian cancers. We show here that Brca1+\\/? mice are normal and fertile and lack tumors by age eleven months. Homozygous Brca15-6 mutant mice die before day 7.5 of embryogenesis. Mutant embryos are poorly developed, with no evidence of mesoderm formation. The extraembryonic region is abnormal, but

  8. Scrapie and cellular PrP isoforms are encoded by the same chromosomal gene

    Microsoft Academic Search

    K BASLER; M Scott; M Wälchli; M P McKinley

    1986-01-01

    Abstract PrP 27-30 is the major protein in purified preparations of scrapie agent. An almost complete PrP cDNA was used to select PrP-related genomic,clones from normal hamster DNA. The gene contains a noncoding exon of 56 to 82 bp and a 2 kb coding exon, separated by a 10 kb intron. Transcription initiates at the same multiple sites in vivo

  9. Roles of the CDC24 gene product in cellular morphogenesis during the Saccharomyces cerevisiae cell cycle.

    PubMed

    Sloat, B F; Adams, A; Pringle, J R

    1981-06-01

    Temperature-sensitive yeast mutants defective in gene CDC24 continued to grow (i.e., increase in cell mass and cell volume) at restrictive temperature (36 degrees C) but were unable to form buds. Staining with the fluorescent dye Calcofluor showed that the mutants were also unable to form normal bud scars (the discrete chitin rings formed in the cell wall at budding sites) at 36 degrees C; instead, large amounts of chitin were deposited randomly over the surfaces of the growing unbudded cells. Labeling of cell-wall mannan with fluorescein isothiocyanate-conjugated concanavalin A suggested that mannan incorporation was also delocalized in mutant cells grown at 36 degrees C. Although the mutants have well-defined execution points just before bud emergence, inactivation of the CDC24 gene product in budded cells led both to selective growth of mother cells rather than of buds and to delocalized chitin deposition, indicating that the CDC24 gene product functions in the normal localization of growth in budded as well as in unbudded cells. Growth of the mutant strains at temperatures less than 36 degrees C revealed allele-specific differences in behavior. Two strains produced buds of abnormal shape during growth at 33 degrees C. Moreover, these same strains displayed abnormal localization of budding sites when growth at 24 degrees C (the normal permissive temperature for the mutants); in each case, the abnormal pattern of budding sites segregated with the temperature sensitivity in crosses. Thus, the CDC24 gene product seems to be involved in selection of the budding site, formation of the chitin ring at that site, the subsequent localization of new cell wall growth to the budding site and the growing bud, and the balance between tip growth and uniform growth of the bud that leads to the normal cell shape. PMID:7019215

  10. Gene expression patterns in Euglena gracilis: Insights into the cellular response to environmental stress

    Microsoft Academic Search

    Verónica dos Santos Ferreira; Iara Rocchetta; Visitación Conforti; Shellie Bench; Robert Feldman; Mariano J. Levin

    2007-01-01

    To better understand Euglena gracilis gene expression under different stress conditions (Chromium, Streptomycin or darkness), we undertook a survey of the E. gracilis transcriptome by cDNA sequencing and microarray analysis. First, we constructed a non-normalized cDNA library from the E. gracilis UTEX strain and sequenced a total of 1000 cDNAs. Six hundred and ten of these ESTs were similar to

  11. Betaine aldehyde dehydrogenase genes from Arabidopsis with different sub-cellular localization affect stress responses

    Microsoft Academic Search

    Tagnon D. Missihoun; Jessica Schmitz; Rebecca Klug; Hans-Hubert Kirch; Dorothea Bartels

    2011-01-01

    Arabidopsis thaliana belongs to those plants that do not naturally accumulate glycine betaine (GB), although its genome contains two genes, ALDH10A8 and ALDH10A9 that code for betaine aldehyde dehydrogenases (BADHs). BADHs were initially known to catalyze the last step of the biosynthesis\\u000a of GB in plants. But they can also oxidize metabolism-derived aminoaldehydes to their corresponding amino acids in some

  12. Convergence of genes and cellular pathways dysregulated in autism spectrum disorders.

    PubMed

    Pinto, Dalila; Delaby, Elsa; Merico, Daniele; Barbosa, Mafalda; Merikangas, Alison; Klei, Lambertus; Thiruvahindrapuram, Bhooma; Xu, Xiao; Ziman, Robert; Wang, Zhuozhi; Vorstman, Jacob A S; Thompson, Ann; Regan, Regina; Pilorge, Marion; Pellecchia, Giovanna; Pagnamenta, Alistair T; Oliveira, Bárbara; Marshall, Christian R; Magalhaes, Tiago R; Lowe, Jennifer K; Howe, Jennifer L; Griswold, Anthony J; Gilbert, John; Duketis, Eftichia; Dombroski, Beth A; De Jonge, Maretha V; Cuccaro, Michael; Crawford, Emily L; Correia, Catarina T; Conroy, Judith; Conceição, Inês C; Chiocchetti, Andreas G; Casey, Jillian P; Cai, Guiqing; Cabrol, Christelle; Bolshakova, Nadia; Bacchelli, Elena; Anney, Richard; Gallinger, Steven; Cotterchio, Michelle; Casey, Graham; Zwaigenbaum, Lonnie; Wittemeyer, Kerstin; Wing, Kirsty; Wallace, Simon; van Engeland, Herman; Tryfon, Ana; Thomson, Susanne; Soorya, Latha; Rogé, Bernadette; Roberts, Wendy; Poustka, Fritz; Mouga, Susana; Minshew, Nancy; McInnes, L Alison; McGrew, Susan G; Lord, Catherine; Leboyer, Marion; Le Couteur, Ann S; Kolevzon, Alexander; Jiménez González, Patricia; Jacob, Suma; Holt, Richard; Guter, Stephen; Green, Jonathan; Green, Andrew; Gillberg, Christopher; Fernandez, Bridget A; Duque, Frederico; Delorme, Richard; Dawson, Geraldine; Chaste, Pauline; Café, Cátia; Brennan, Sean; Bourgeron, Thomas; Bolton, Patrick F; Bölte, Sven; Bernier, Raphael; Baird, Gillian; Bailey, Anthony J; Anagnostou, Evdokia; Almeida, Joana; Wijsman, Ellen M; Vieland, Veronica J; Vicente, Astrid M; Schellenberg, Gerard D; Pericak-Vance, Margaret; Paterson, Andrew D; Parr, Jeremy R; Oliveira, Guiomar; Nurnberger, John I; Monaco, Anthony P; Maestrini, Elena; Klauck, Sabine M; Hakonarson, Hakon; Haines, Jonathan L; Geschwind, Daniel H; Freitag, Christine M; Folstein, Susan E; Ennis, Sean; Coon, Hilary; Battaglia, Agatino; Szatmari, Peter; Sutcliffe, James S; Hallmayer, Joachim; Gill, Michael; Cook, Edwin H; Buxbaum, Joseph D; Devlin, Bernie; Gallagher, Louise; Betancur, Catalina; Scherer, Stephen W

    2014-05-01

    Rare copy-number variation (CNV) is an important source of risk for autism spectrum disorders (ASDs). We analyzed 2,446 ASD-affected families and confirmed an excess of genic deletions and duplications in affected versus control groups (1.41-fold, p = 1.0 × 10(-5)) and an increase in affected subjects carrying exonic pathogenic CNVs overlapping known loci associated with dominant or X-linked ASD and intellectual disability (odds ratio = 12.62, p = 2.7 × 10(-15), ?3% of ASD subjects). Pathogenic CNVs, often showing variable expressivity, included rare de novo and inherited events at 36 loci, implicating ASD-associated genes (CHD2, HDAC4, and GDI1) previously linked to other neurodevelopmental disorders, as well as other genes such as SETD5, MIR137, and HDAC9. Consistent with hypothesized gender-specific modulators, females with ASD were more likely to have highly penetrant CNVs (p = 0.017) and were also overrepresented among subjects with fragile X syndrome protein targets (p = 0.02). Genes affected by de novo CNVs and/or loss-of-function single-nucleotide variants converged on networks related to neuronal signaling and development, synapse function, and chromatin regulation. PMID:24768552

  13. Convergence of Genes and Cellular Pathways Dysregulated in Autism Spectrum Disorders

    PubMed Central

    Pinto, Dalila; Delaby, Elsa; Merico, Daniele; Barbosa, Mafalda; Merikangas, Alison; Klei, Lambertus; Thiruvahindrapuram, Bhooma; Xu, Xiao; Ziman, Robert; Wang, Zhuozhi; Vorstman, Jacob A.S.; Thompson, Ann; Regan, Regina; Pilorge, Marion; Pellecchia, Giovanna; Pagnamenta, Alistair T.; Oliveira, Bárbara; Marshall, Christian R.; Magalhaes, Tiago R.; Lowe, Jennifer K.; Howe, Jennifer L.; Griswold, Anthony J.; Gilbert, John; Duketis, Eftichia; Dombroski, Beth A.; De Jonge, Maretha V.; Cuccaro, Michael; Crawford, Emily L.; Correia, Catarina T.; Conroy, Judith; Conceição, Inês C.; Chiocchetti, Andreas G.; Casey, Jillian P.; Cai, Guiqing; Cabrol, Christelle; Bolshakova, Nadia; Bacchelli, Elena; Anney, Richard; Gallinger, Steven; Cotterchio, Michelle; Casey, Graham; Zwaigenbaum, Lonnie; Wittemeyer, Kerstin; Wing, Kirsty; Wallace, Simon; van Engeland, Herman; Tryfon, Ana; Thomson, Susanne; Soorya, Latha; Rogé, Bernadette; Roberts, Wendy; Poustka, Fritz; Mouga, Susana; Minshew, Nancy; McInnes, L. Alison; McGrew, Susan G.; Lord, Catherine; Leboyer, Marion; Le Couteur, Ann S.; Kolevzon, Alexander; Jiménez González, Patricia; Jacob, Suma; Holt, Richard; Guter, Stephen; Green, Jonathan; Green, Andrew; Gillberg, Christopher; Fernandez, Bridget A.; Duque, Frederico; Delorme, Richard; Dawson, Geraldine; Chaste, Pauline; Café, Cátia; Brennan, Sean; Bourgeron, Thomas; Bolton, Patrick F.; Bölte, Sven; Bernier, Raphael; Baird, Gillian; Bailey, Anthony J.; Anagnostou, Evdokia; Almeida, Joana; Wijsman, Ellen M.; Vieland, Veronica J.; Vicente, Astrid M.; Schellenberg, Gerard D.; Pericak-Vance, Margaret; Paterson, Andrew D.; Parr, Jeremy R.; Oliveira, Guiomar; Nurnberger, John I.; Monaco, Anthony P.; Maestrini, Elena; Klauck, Sabine M.; Hakonarson, Hakon; Haines, Jonathan L.; Geschwind, Daniel H.; Freitag, Christine M.; Folstein, Susan E.; Ennis, Sean; Coon, Hilary; Battaglia, Agatino; Szatmari, Peter; Sutcliffe, James S.; Hallmayer, Joachim; Gill, Michael; Cook, Edwin H.; Buxbaum, Joseph D.; Devlin, Bernie; Gallagher, Louise; Betancur, Catalina; Scherer, Stephen W.

    2014-01-01

    Rare copy-number variation (CNV) is an important source of risk for autism spectrum disorders (ASDs). We analyzed 2,446 ASD-affected families and confirmed an excess of genic deletions and duplications in affected versus control groups (1.41-fold, p = 1.0 × 10?5) and an increase in affected subjects carrying exonic pathogenic CNVs overlapping known loci associated with dominant or X-linked ASD and intellectual disability (odds ratio = 12.62, p = 2.7 × 10?15, ?3% of ASD subjects). Pathogenic CNVs, often showing variable expressivity, included rare de novo and inherited events at 36 loci, implicating ASD-associated genes (CHD2, HDAC4, and GDI1) previously linked to other neurodevelopmental disorders, as well as other genes such as SETD5, MIR137, and HDAC9. Consistent with hypothesized gender-specific modulators, females with ASD were more likely to have highly penetrant CNVs (p = 0.017) and were also overrepresented among subjects with fragile X syndrome protein targets (p = 0.02). Genes affected by de novo CNVs and/or loss-of-function single-nucleotide variants converged on networks related to neuronal signaling and development, synapse function, and chromatin regulation. PMID:24768552

  14. Coupling of Cellular Processes and Their Coordinated Oscillations under Continuous Light in Cyanothece sp. ATCC 51142, a Diazotrophic Unicellular Cyanobacterium.

    PubMed

    Krishnakumar, S; Gaudana, Sandeep B; Vinh, Nguyen X; Viswanathan, Ganesh A; Chetty, Madhu; Wangikar, Pramod P

    2015-01-01

    Unicellular diazotrophic cyanobacteria such as Cyanothece sp. ATCC 51142 (henceforth Cyanothece), temporally separate the oxygen sensitive nitrogen fixation from oxygen evolving photosynthesis not only under diurnal cycles (LD) but also in continuous light (LL). However, recent reports demonstrate that the oscillations in LL occur with a shorter cycle time of ~11 h. We find that indeed, majority of the genes oscillate in LL with this cycle time. Genes that are upregulated at a particular time of day under diurnal cycle also get upregulated at an equivalent metabolic phase under LL suggesting tight coupling of various cellular events with each other and with the cell's metabolic status. A number of metabolic processes get upregulated in a coordinated fashion during the respiratory phase under LL including glycogen degradation, glycolysis, oxidative pentose phosphate pathway, and tricarboxylic acid cycle. These precede nitrogen fixation apparently to ensure sufficient energy and anoxic environment needed for the nitrogenase enzyme. Photosynthetic phase sees upregulation of photosystem II, carbonate transport, carbon concentrating mechanism, RuBisCO, glycogen synthesis and light harvesting antenna pigment biosynthesis. In Synechococcus elongates PCC 7942, a non-nitrogen fixing cyanobacteria, expression of a relatively smaller fraction of genes oscillates under LL condition with the major periodicity being 24 h. In contrast, the entire cellular machinery of Cyanothece orchestrates coordinated oscillation in anticipation of the ensuing metabolic phase in both LD and LL. These results may have important implications in understanding the timing of various cellular events and in engineering cyanobacteria for biofuel production. PMID:25973856

  15. Correlation of rare coding variants in the gene encoding human glucokinase regulatory protein with phenotypic, cellular, and kinetic outcomes.

    PubMed

    Rees, Matthew G; Ng, David; Ruppert, Sarah; Turner, Clesson; Beer, Nicola L; Swift, Amy J; Morken, Mario A; Below, Jennifer E; Blech, Ilana; Mullikin, James C; McCarthy, Mark I; Biesecker, Leslie G; Gloyn, Anna L; Collins, Francis S

    2012-01-01

    Defining the genetic contribution of rare variants to common diseases is a major basic and clinical science challenge that could offer new insights into disease etiology and provide potential for directed gene- and pathway-based prevention and treatment. Common and rare nonsynonymous variants in the GCKR gene are associated with alterations in metabolic traits, most notably serum triglyceride levels. GCKR encodes glucokinase regulatory protein (GKRP), a predominantly nuclear protein that inhibits hepatic glucokinase (GCK) and plays a critical role in glucose homeostasis. The mode of action of rare GCKR variants remains unexplored. We identified 19 nonsynonymous GCKR variants among 800 individuals from the ClinSeq medical sequencing project. Excluding the previously described common missense variant p.Pro446Leu, all variants were rare in the cohort. Accordingly, we functionally characterized all variants to evaluate their potential phenotypic effects. Defects were observed for the majority of the rare variants after assessment of cellular localization, ability to interact with GCK, and kinetic activity of the encoded proteins. Comparing the individuals with functional rare variants to those without such variants showed associations with lipid phenotypes. Our findings suggest that, while nonsynonymous GCKR variants, excluding p.Pro446Leu, are rare in individuals of mixed European descent, the majority do affect protein function. In sum, this study utilizes computational, cell biological, and biochemical methods to present a model for interpreting the clinical significance of rare genetic variants in common disease. PMID:22182842

  16. DaTrypsin, a novel clip-domain serine proteinase gene up-regulated during winter and summer diapauses of the onion maggot, Delia antiqua

    E-print Network

    Monteiro, Antónia

    diapauses of the onion maggot, Delia antiqua Bin Chena,b,c,*, Takumi Kayukawaa , Haobo Jiangd , Anto of winter (WD) and summer diapauses (SD), we screened for diapause-specific genes in the onion maggot, Delia

  17. Human ImmunodeficiencyVirus Type 1 tat Gene Up-regulates Interleukin 4 Receptors on a Human B-Lymphoblastoid Cell Line

    Microsoft Academic Search

    Raj K. Puri; Bharat B. Aggarwal

    1992-01-01

    The human immunodeficiency virus type I (HIV-1) regulatory gene, tat III, is a powerful fra\\/is-activator of gene expression from the viral long terminal repeat and is essential for HIV replication. In addition, tat III protein has been shown to be immunosuppressive as indicated by the inhibition of antigen mediated T-cell proliferation. To further test whether tat III might play a

  18. hEST2, the Putative Human Telomerase Catalytic Subunit Gene, Is Up-Regulated in Tumor Cells and during Immortalization

    Microsoft Academic Search

    Matthew Meyerson; Christopher M Counter; Elinor Ng Eaton; Leif W Ellisen; Philipp Steiner; Stephanie Dickinson Caddle; Liuda Ziaugra; Roderick L Beijersbergen; Michael J Davidoff; Qingyun Liu; Silvia Bacchetti; Daniel A Haber; Robert A Weinberg

    1997-01-01

    Telomerase, the ribonucleoprotein enzyme that elongates telomeres, is repressed in normal human somatic cells but is reactivated during tumor progression. We report the cloning of a human gene, hEST2, that shares significant sequence similarity with the telomerase catalytic subunit genes of lower eukaryotes. hEST2 is expressed at high levels in primary tumors, cancer cell lines, and telomerase-positive tissues but is

  19. RNA pol II transcript abundance controls condensin accumulation at mitotically up-regulated and heat-shock-inducible genes in fission yeast.

    PubMed

    Nakazawa, Norihiko; Sajiki, Kenichi; Xu, Xingya; Villar-Briones, Alejandro; Arakawa, Orie; Yanagida, Mitsuhiro

    2015-06-01

    Condensin plays fundamental roles in chromosome dynamics. In this study, we determined the binding sites of condensin on fission yeast (Schizosaccharomyces pombe) chromosomes at the level of nucleotide sequences using chromatin immunoprecipitation (ChIP) and ChIP sequencing (ChIP-seq). We found that condensin binds to RNA polymerase I-, II- and III-transcribed genes during both mitosis and interphase, and we focused on pol II constitutive and inducible genes. Accumulation sites for condensin are distinct from those of cohesin and DNA topoisomerase II. Using cell cycle stage and heat-shock-inducible genes, we show that pol II-mediated transcripts cause condensin accumulation. First, condensin's enrichment on mitotically activated genes was abolished by deleting the sep1(+) gene that encodes an M-phase-specific forkhead transcription factor. Second, by raising the temperature, condensin accumulation was rapidly induced at heat-shock protein genes in interphase and even during mid-mitosis. In interphase, condensin accumulates preferentially during the postreplicative phase. Pol II-mediated transcription was neither repressed nor activated by condensin, as levels of transcripts per se did not change when mutant condensin failed to associate with chromosomal DNA. However, massive chromosome missegregation occurred, suggesting that abundant pol II transcription may require active condensin before proper chromosome segregation. PMID:25847133

  20. RNA pol II transcript abundance controls condensin accumulation at mitotically up-regulated and heat-shock-inducible genes in fission yeast

    PubMed Central

    Nakazawa, Norihiko; Sajiki, Kenichi; Xu, Xingya; Villar-Briones, Alejandro; Arakawa, Orie; Yanagida, Mitsuhiro

    2015-01-01

    Condensin plays fundamental roles in chromosome dynamics. In this study, we determined the binding sites of condensin on fission yeast (Schizosaccharomyces pombe) chromosomes at the level of nucleotide sequences using chromatin immunoprecipitation (ChIP) and ChIP sequencing (ChIP-seq). We found that condensin binds to RNA polymerase I-, II- and III-transcribed genes during both mitosis and interphase, and we focused on pol II constitutive and inducible genes. Accumulation sites for condensin are distinct from those of cohesin and DNA topoisomerase II. Using cell cycle stage and heat-shock-inducible genes, we show that pol II-mediated transcripts cause condensin accumulation. First, condensin's enrichment on mitotically activated genes was abolished by deleting the sep1+ gene that encodes an M-phase-specific forkhead transcription factor. Second, by raising the temperature, condensin accumulation was rapidly induced at heat-shock protein genes in interphase and even during mid-mitosis. In interphase, condensin accumulates preferentially during the postreplicative phase. Pol II-mediated transcription was neither repressed nor activated by condensin, as levels of transcripts per se did not change when mutant condensin failed to associate with chromosomal DNA. However, massive chromosome missegregation occurred, suggesting that abundant pol II transcription may require active condensin before proper chromosome segregation. PMID:25847133

  1. Up-Regulation of mRNA Ventricular PRNP Prion Protein Gene Expression in Air Pollution Highly Exposed Young Urbanites: Endoplasmic Reticulum Stress, Glucose Regulated Protein 78, and Nanosized Particles

    PubMed Central

    Villarreal-Calderon, Rodolfo; Franco-Lira, Maricela; González-Maciel, Angélica; Reynoso-Robles, Rafael; Harritt, Lou; Pérez-Guillé, Beatriz; Ferreira-Azevedo, Lara; Drecktrah, Dan; Zhu, Hongtu; Sun, Qiang; Torres-Jardón, Ricardo; Aragón-Flores, Mariana; Calderón-Garcidueñas, Ana; Diaz, Philippe; Calderón-Garcidueñas, Lilian

    2013-01-01

    Mexico City Metropolitan Area children and young adults exposed to high concentrations of air pollutants including fine and ultrafine particulate matter (PM) vs. clean air controls, exhibit myocardial inflammation and inflammasome activation with a differential right and left ventricular expression of key inflammatory genes and inflammasomes. We investigated the mRNA expression levels of the prion protein gene PRNP, which plays an important role in the protection against oxidative stress and metal toxicity, and the glucose regulated protein 78, a key protein in endoplasmic reticulum (ER) stress signaling, in ventricular autopsy samples from 30 children and young adults age 19.97 ± 6.8 years with a lifetime of low (n:4) vs. high (n:26) air pollution exposures. Light microscopy and transmission electron microscopy studies were carried out in human ventricles, and electron microscopy studies were also done in 5 young, highly exposed Mexico City dogs. There was significant left ventricular PRNP and bi-ventricular GRP78 mRNA up-regulation in Mexico City young urbanites vs. controls. PRNP up-regulation in the left ventricle was significantly different from the right, p < 0.0001, and there was a strong left ventricular PRNP and GRP78 correlation (p = 0.0005). Marked abnormalities in capillary endothelial cells, numerous nanosized particles in myocardial ER and in abnormal mitochondria characterized the highly exposed ventricles. Early and sustained cardiac ER stress could result in detrimental irreversible consequences in urban children, and while highly complex systems maintain myocardial homeostasis, failure to compensate for chronic myocardial inflammation, oxidative and ER stress, and particles damaging myocardial organelles may prime the development of pathophysiological cardiovascular states in young urbanites. Nanosized PM could play a key cardiac myocyte toxicity role. PMID:24287918

  2. Sox9/Sox6 and Sp1 are involved in the insulin-like growth factor-I-mediated upregulation of human type II collagen gene expression in articular chondrocytes.

    PubMed

    Renard, Emmanuelle; Porée, Benoît; Chadjichristos, Christos; Kypriotou, Magdalini; Maneix, Laure; Bigot, Nicolas; Legendre, Florence; Ollitrault, David; De Crombrugghe, Benoît; Malléin-Gérin, Frédéric; Moslemi, Safa; Demoor, Magali; Boumediene, Karim; Galéra, Philippe

    2012-06-01

    Type II collagen is a marker of articular cartilage encoded by the COL2A1 gene. The nature of the trans factors involved in the upregulation of this gene by insulin-like growth factor-I (IGF-I) remains unclear. We found that IGF-I increased type II collagen synthesis by a transcriptional control mechanism involving a 715-bp region within the COL2A1 first-intron specific enhancer. The overproduction of L-Sox5/Sox6/Sox9 and Sp1 and decoy experiments targeting these factors demonstrated their action in concert in IGF-I trans-activation. These results were supported by the data obtained in knockdown experiments in which siRNA against Sox9/Sox6 and Sp1 prevented the IGF-I-induced increase in collagen II production. Indeed, each of these trans-activators increased the expression of others. IGF-I increased the binding of Sox9 and Sp1/Sp3 to their cis elements in the enhancer, and we provide the first evidence of Sox9 interaction with the promoter by chromatin immunoprecipitation. Interactions with COL2A1 were also observed for Sp1, p300/CBP, and Tip60. Finally, a physical interaction between Sox9, p300, Sp3, and Sp1 was detected. These data demonstrate the role of Sox9, Sp1/Sp3, and euchromatin-associated factors (p300, Tip60) in the IGF-I-induced upregulation of COL2A1, indicating possible use of this growth factor in articular cartilage engineering applications to promote repair in patients with degenerative diseases, such as osteoarthritis. PMID:22215151

  3. Analysis of the human guanylin gene and the processing and cellular localization of the peptide.

    PubMed

    Hill, O; Kuhn, M; Zucht, H D; Cetin, Y; Kulaksiz, H; Adermann, K; Klock, G; Rechkemmer, G; Forssmann, W G; Mägert, H J

    1995-03-14

    The complete cell biological analysis of human guanylin, a recently discovered regulatory peptide, is offered in this investigation: (i) the nucleotide sequence of the gene, (ii) the isolation and characterization of its circulating molecular form, and (iii) its localization in enterochromaffin cells of the gut. As determined by molecular cloning, DNA sequencing, and comparison with the known cDNA sequence, the approximately 2.6-kbp large gene consists of three exons interrupted by two introns. The putative promoter region contains a TTTAAAA sequence motif and several potential binding sites for transcription factors such as AP-1, AP-2, Sp 1, and glucocorticoid receptors. The isolated hormonal form of guanylin is a 94-amino acid peptide with a molecular mass of 10.3 kDa. Western blot analysis of RP-HPLC fractions from blood plasma confirms this molecular form. Thus, guanylin is synthesized by gut enterochromaffin cells as a prohormone of 115 amino acids and is processed to the molecular form of 94 amino acids circulating in the blood. PMID:7892222

  4. Cellular immune response against firefly luciferase after sleeping beauty-mediated gene transfer in vivo.

    PubMed

    Podetz-Pedersen, Kelly M; Vezys, Vaiva; Somia, Nikunj V; Russell, Stephen J; McIvor, R Scott

    2014-11-01

    The Sleeping Beauty (SB) transposon system has been shown to mediate new gene sequence integration resulting in long-term expression. Here the effectiveness of hyperactive SB100X transposase was tested, and we found that hydrodynamic co-delivery of a firefly luciferase transposon (pT2/CaL) along with SB100X transposase (pCMV-SB100X) resulted in remarkably sustained, high levels of luciferase expression. However, after 4 weeks there was a rapid, animal-by-animal loss of luciferase expression that was not observed in immunodeficient mice. We hypothesized that this sustained, high-level luciferase expression achieved using the SB100X transposase elicits an immune response in pT2/CaL co-administered mice, which was supported by the rapid loss of luciferase expression upon challenge of previously treated animals and in naive animals adoptively transferred with splenocytes from previously treated animals. Specificity of the immune response to luciferase was demonstrated by increased cytokine expression in splenocytes after exposure to luciferase peptide in parallel with MHC I-luciferase peptide tetramer binding. This anti-luciferase immune response observed following continuous, high-level luciferase expression in vivo clearly impacts its use as an in vivo reporter. As both an immunogen and an extremely sensitive reporter, luciferase is also a useful model system for the study of immune responses following in vivo gene transfer and expression. PMID:25093708

  5. Identification of Cellular Genes Affecting the Infectivity of Foot-and-Mouth Disease Virus?

    PubMed Central

    Piccone, Maria E.; Feng, Yanan; Chang, Annie C. Y.; Mosseri, Ronen; Lu, Quan; Kutish, Gerald F.; Lu, Zhiqiang; Burrage, Thomas G.; Gooch, Christina; Rock, Daniel L.; Cohen, Stanley N.

    2009-01-01

    Foot-and-mouth disease virus (FMDV) produces one of the most infectious of all livestock diseases, causing extensive economic loss in areas of breakout. Like other viral pathogens, FMDV recruits proteins encoded by host cell genes to accomplish the entry, replication, and release of infectious viral particles. To identify such host-encoded proteins, we employed an antisense RNA strategy and a lentivirus-based library containing approximately 40,000 human expressed sequence tags (ESTs) to randomly inactivate chromosomal genes in a bovine kidney cell line (LF-BK) that is highly susceptible to FMDV infection and then isolated clones that survived multiple rounds of exposure to the virus. Here, we report the identification of ESTs whose expression in antisense orientation limited host cell killing by FMDV and restricted viral propagation. The role of one such EST, that of ectonucleoside triphosphate diphosphohydrolase 6 (NTPDase6; also known as CD39L2), a membrane-associated ectonucleoside triphosphate diphosphohydrolase that previously was not suspected of involvement in the propagation of viral pathogens and which we now show is required for normal synthesis of FMDV RNA and proteins, is described in this report. PMID:19369337

  6. Cellular and functional analysis of four mutations located in the mitochondrial ATPase6 gene.

    PubMed

    Vazquez-Memije, Martha Elisa; Rizza, Teresa; Meschini, Maria Chiara; Nesti, Claudia; Santorelli, Filippo Maria; Carrozzo, Rosalba

    2009-04-01

    The smallest rotary motor of living cells, F0F1-ATP synthase, couples proton flow-generated by the OXPHOS system-from the intermembrane space back to the matrix with the conversion of ADP to ATP. While all mutations affecting the multisubunit complexes of the OXPHOS system probably impact on the cell's output of ATP, only mutations in complex V can be considered to affect this output directly. So far, most of the F0F1-ATP synthase variations have been detected in the mitochondrial ATPase6 gene. In this study, the four most frequent mutations in the ATPase6 gene, namely L156R, L217R, L156P, and L217P, are studied for the first time together, both in primary cells and in cybrid clones. Arginine ("R") mutations were associated with a much more severe phenotype than Proline ("P") mutations, in terms of both biochemical activity and growth capacity. Also, a threshold effect in both "R" mutations appeared at 50% mutation load. Different mechanisms seemed to emerge for the two "R" mutations: the F1 seemed loosely bound to the membrane in the L156R mutant, whereas the L217R mutant induced low activity of complex V, possibly the result of a reduced rate of proton flow through the A6 channel. PMID:19160410

  7. Cellular imaging demonstrates genetic mosaicism in heterozygous carriers of an X-linked ciliopathy gene.

    PubMed

    Pyo Park, Sung; Hwan Hong, In; Tsang, Stephen H; Chang, Stanley

    2013-11-01

    X-linked retinitis pigmentosa (XLRP) is the least common genetic type of retinitis pigmentosa; however, it has extremely devastating consequences to patients' activities of daily living. RPGR and RP2 genes expressed in the photoreceptor sensory cilia are predominantly implicated in XLRP; however, the interpretation of genetic mutations and their correlation with clinical phenotypes remain unknown, and the role of these genes in photoreceptor cilia function is not completely elucidated. Therefore, we evaluated structural characteristics in five female obligate carriers of XLRP by using state-of-the-art non-invasive imaging methods, including adaptive optics (AO) scanning laser ophthalmoscopy (SLO). In all five carriers examined, qualitative and quantitative analyses by AO SLO imaging revealed a mosaic pattern of cone disruption, even in the absence of visual symptoms, normal visual acuity and normal macular thickness, on optical coherence tomography and mildly subnormal full-field cone electroretinographic findings. As the technique is sensitive to the level of a single cone, the ability to visualize the cone cells in vivo should be especially useful in other retinal diseases. In addition, further investigation of XLRP carriers may yield insight into how cone structures change over time and ultimately enable understanding of the role of RPGR and RP2 in cone cell survival. PMID:23443027

  8. Epigenetic transcriptional repression of cellular genes by a viral SET protein

    PubMed Central

    Mujtaba, Shiraz; Manzur, Karishma L.; Gurnon, James R.; Kang, Ming; Van Etten, James L.; Zhou, Ming-Ming

    2010-01-01

    Viruses recruit host proteins to secure viral genome maintenance and replication. However, whether they modify host histones directly to interfere with chromatin-based transcription is unknown. Here we report that Paramecium bursaria chlorella virus 1 (PBCV-1) encodes a functional SET domain histone Lys methyltransferase (HKMTase) termed vSET, which is linked to rapid inhibition of host transcription after viral infection. We show that vSET is packaged in the PBCV-1 virion, and that it contains a nuclear localization signal and probably represses host transcription by methylating histone H3 at Lys 27 (H3K27), a modification known to trigger gene silencing in eukaryotes. We also show that vSET induces cell accumulation at the G2/M phase by recruiting the Polycomb repressive complex CBX8 to the methylated H3K27 site in a heterologous system. vSET-like proteins that have H3K27 methylation activity are conserved in chlorella viruses. Our findings suggest a viral mechanism to repress gene transcription by direct modification of chromatin by PBCV-1 vSET. PMID:19160493

  9. Allyl isothiocyanate depletes glutathione and upregulates expression of glutathione S-transferases in Arabidopsis thaliana.

    PubMed

    Øverby, Anders; Stokland, Ragni A; Åsberg, Signe E; Sporsheim, Bjørnar; Bones, Atle M

    2015-01-01

    Allyl isothiocyanate (AITC) is a phytochemical associated with plant defense in plants from the Brassicaceae family. AITC has long been recognized as a countermeasure against external threats, but recent reports suggest that AITC is also involved in the onset of defense-related mechanisms such as the regulation of stomatal aperture. However, the underlying cellular modes of action in plants remain scarcely investigated. Here we report evidence of an AITC-induced depletion of glutathione (GSH) and the effect on gene expression of the detoxification enzyme family glutathione S-transferases (GSTs) in Arabidopsis thaliana. Treatment of A. thaliana wild-type with AITC resulted in a time- and dose-dependent depletion of cellular GSH. AITC-exposure of mutant lines vtc1 and pad2-1 with elevated and reduced GSH-levels, displayed enhanced and decreased AITC-tolerance, respectively. AITC-exposure also led to increased ROS-levels in the roots and loss of chlorophyll which are symptoms of oxidative stress. Following exposure to AITC, we found that GSH rapidly recovered to the same level as in the control plant, suggesting an effective route for replenishment of GSH or a rapid detoxification of AITC. Transcriptional analysis of genes encoding GSTs showed an upregulation in response to AITC. These findings demonstrate cellular effects by AITC involving a reversible depletion of the GSH-pool, induced oxidative stress, and elevated expression of GST-encoding genes. PMID:25954298

  10. Respiratory Viral Infections and Subversion of Cellular Antioxidant Defenses

    PubMed Central

    Komaravelli, Narayana; Casola, Antonella

    2014-01-01

    Reactive oxygen species (ROS) formation is part of normal cellular aerobic metabolism, due to respiration and oxidation of nutrients in order to generate energy. Low levels of ROS are involved in cellular signaling and are well controlled by the cellular antioxidant defense system. Elevated levels of ROS generation due to pollutants, toxins and radiation exposure, as well as infections, are associated with oxidative stress causing cellular damage. Several respiratory viruses, including respiratory syncytial virus (RSV), human metapneumovirus (hMPV) and influenza, induce increased ROS formation, both intracellularly and as a result of increased inflammatory cell recruitment at the site of infection. They also reduce antioxidant enzyme (AOE) levels and/or activity, leading to unbalanced oxidative-antioxidant status and subsequent oxidative cell damage. Expression of several AOE is controlled by the activation of the nuclear transcription factor NF-E2-related factor 2 (Nrf2), through binding to the antioxidant responsive element (ARE) present in the AOE gene promoters. While exposure to several pro-oxidant stimuli usually leads to Nrf2 activation and upregulation of AOE expression, respiratory viral infections are associated with inhibition of AOE expression/activity, which in the case of RSV and hMPV is associated with reduced Nrf2 nuclear localization, decreased cellular levels and reduced ARE-dependent gene transcription. Therefore, administration of antioxidant mimetics or Nrf2 inducers represents potential viable therapeutic approaches to viral-induced diseases, such as respiratory infections and other infections associated with decreased cellular antioxidant capacity. PMID:25584194

  11. Identification and Upregulation of Biosynthetic Genes Required for Accumulation of Mycosporine-2-Glycine under Salt Stress Conditions in the Halotolerant Cyanobacterium Aphanothece halophytica

    PubMed Central

    Waditee-Sirisattha, Rungaroon; Kageyama, Hakuto; Sopun, Warangkana; Tanaka, Yoshito

    2014-01-01

    Mycosporine-like amino acids (MAAs) are valuable molecules that are the basis for important photoprotective constituents. Here we report molecular analysis of mycosporine-like amino acid biosynthetic genes from the halotolerant cyanobacterium Aphanothece halophytica, which can survive at high salinity and alkaline pH. This extremophile was found to have a unique MAA core (4-deoxygadusol)-synthesizing gene separated from three other genes. In vivo analysis showed accumulation of the mycosporine-2-glycine but not shinorine or mycosporine-glycine. Mycosporine-2-glycine accumulation was stimulated more under the stress condition of high salinity than UV-B radiation. The Aphanothece MAA biosynthetic genes also manifested a strong transcript level response to salt stress. Furthermore, the transformed Escherichia coli and Synechococcus strains expressing four putative Aphanothece MAA genes under the control of a native promoter were found to be capable of synthesizing mycosporine-2-glycine. The accumulation level of mycosporine-2-glycine was again higher under the high-salinity condition. In the transformed E. coli cells, its level was approximately 85.2 ± 0.7 ?mol/g (dry weight). Successful production of a large amount of mycosporine in these cells provides a new opportunity in the search for an alternative natural sunscreen compound source. PMID:24375141

  12. High hydrostatic pressure extract of garlic increases the HDL cholesterol level via up-regulation of apolipoprotein A-I gene expression in rats fed a high-fat diet

    PubMed Central

    2012-01-01

    Background Cardiovascular disease (CVD) is the number one cause of mortality worldwide and a low high-density lipoprotein cholesterol (HDL-C) level is an important marker of CVD risk. Garlic (Allium sativum) has been widely used in the clinic for treatment of CVD and regulation of lipid metabolism. This study investigated the effects of a high hydrostatic pressure extract of garlic (HEG) on HDL-C level and regulation of hepatic apolipoprotein A-I (apoA-I) gene expression. Methods Male Sprague–Dawley rats were divided into two groups and maintained on a high-fat control diet (CON) or high-fat control diet supplemented with high hydrostatic pressure extract of garlic (HEG) for 5 weeks. Changes in the expression of genes related to HDL-C metabolism were analyzed in liver, together with biometric and blood parameters. Results In the HEG group, the plasma triglyceride (TG) and low-density lipoprotein cholesterol (LDL-C) levels were significantly decreased in comparison with the CON group (P?gene expression of ATP-binding cassette transporter A1 (ABCA1) and lecithin:cholesterol acyltransferase (LCAT), importantly involved in the biogenesis in HDL, were also up-regulated by dietary HEG. Conclusions These results suggest that HEG ameliorates plasma lipid profiles and attenuates hepatic lipid accumulation in the high-fat fed rats. Our findings provides that the effects of HEG on the increase of the plasma HDL-C level was at least partially mediated by up-regulation of hepatic genes expression such as apoA-I, ABCA1, and LCAT in rats fed a high-fat diet. PMID:22713542

  13. Activation of PPAR{delta} up-regulates fatty acid oxidation and energy uncoupling genes of mitochondria and reduces palmitate-induced apoptosis in pancreatic {beta}-cells

    SciTech Connect

    Wan, Jun; Jiang, Li; Lue, Qingguo; Ke, Linqiu [Department of Endocrinology, West China Hospital of Sichuan University, 37 Guoxue Lane, Chengdu, Sichuan 610041 (China)] [Department of Endocrinology, West China Hospital of Sichuan University, 37 Guoxue Lane, Chengdu, Sichuan 610041 (China); Li, Xiaoyu [State Key Laboratory of Oral Diseases, Sichuan University, No. 14, 3rd Section, Renmin South Road, Chengdu, Sichuan 610041 (China)] [State Key Laboratory of Oral Diseases, Sichuan University, No. 14, 3rd Section, Renmin South Road, Chengdu, Sichuan 610041 (China); Tong, Nanwei, E-mail: buddyjun@hotmail.com [Department of Endocrinology, West China Hospital of Sichuan University, 37 Guoxue Lane, Chengdu, Sichuan 610041 (China)] [Department of Endocrinology, West China Hospital of Sichuan University, 37 Guoxue Lane, Chengdu, Sichuan 610041 (China)

    2010-01-15

    Recent evidence indicates that decreased oxidative capacity, lipotoxicity, and mitochondrial aberrations contribute to the development of insulin resistance and type 2 diabetes. The goal of this study was to investigate the effects of peroxisome proliferator-activated receptor {delta} (PPAR{delta}) activation on lipid oxidation, mitochondrial function, and insulin secretion in pancreatic {beta}-cells. After HIT-T15 cells (a {beta}-cell line) were exposed to high concentrations of palmitate and GW501516 (GW; a selective agonist of PPAR{delta}), we found that administration of GW increased the expression of PPAR{delta} mRNA. GW-induced activation of PPAR{delta} up-regulated carnitine palmitoyltransferase 1 (CPT1), long-chain acyl-CoA dehydrogenase (LCAD), pyruvate dehydrogenase kinase 4 (PDK4), and uncoupling protein 2 (UCP2); alleviated mitochondrial swelling; attenuated apoptosis; and reduced basal insulin secretion induced by increased palmitate in HIT cells. These results suggest that activation of PPAR{delta} plays an important role in protecting pancreatic {beta}-cells against aberrations caused by lipotoxicity in metabolic syndrome and diabetes.

  14. Werner syndrome protein limits MYC-induced cellular senescence

    PubMed Central

    Grandori, Carla; Wu, Kou-Juey; Fernandez, Paula; Ngouenet, Celine; Grim, Jonathan; Clurman, Bruce E.; Moser, Michael J.; Oshima, Junko; Russell, David W.; Swisshelm, Karen; Frank, Scott; Amati, Bruno; Dalla-Favera, Riccardo; Monnat, Raymond J.

    2003-01-01

    The MYC oncoprotein is a transcription factor that coordinates cell growth and division. MYC overexpression exacerbates genomic instability and sensitizes cells to apoptotic stimuli. Here we demonstrate that MYC directly stimulates transcription of the human Werner syndrome gene, WRN, which encodes a conserved RecQ helicase. Loss-of-function mutations in WRN lead to genomic instability, an elevated cancer risk, and premature cellular senescence. The overexpression of MYC in WRN syndrome fibroblasts or after WRN depletion from control fibroblasts led to rapid cellular senescence that could not be suppressed by hTERT expression. We propose that WRN up-regulation by MYC may promote MYC-driven tumorigenesis by preventing cellular senescence. PMID:12842909

  15. NF-?Bp65 and Expression of Its Pro-Inflammatory Target Genes Are Upregulated in the Subcutaneous Adipose Tissue of Cachectic Cancer Patients

    PubMed Central

    Gonzalez Camargo, Rodolfo; Mendes dos Reis Riccardi, Daniela; Quintas Teixeira Ribeiro, Henrique; Carlos Carnevali, Luiz; Marques de Matos-Neto, Emidio; Enjiu, Lucas; Xavier Neves, Rodrigo; Darck Carola Correia Lima, Joanna; Galvão Figuerêdo, Raquel; Sérgio Martins de Alcântara, Paulo; Maximiano, Linda; Otoch, José; Batista, Miguel Luiz; Püschel, Gerhard; Seelaender, Marilia

    2015-01-01

    Cancer cachexia, of which the most notable symptom is severe and rapid weight loss, is present in the majority of patients with advanced cancer. Inflammatory mediators play an important role in the development of cachexia, envisaged as a chronic inflammatory syndrome. The white adipose tissue (WAT) is one of the first compartments affected in cancer cachexia and suffers a high rate of lipolysis. It secretes several cytokines capable of directly regulating intermediate metabolism. A common pathway in the regulation of the expression of pro-inflammatory cytokines in WAT is the activation of the nuclear transcription factor kappa-B (NF-?B). We have examined the gene expression of the subunits NF-?Bp65 and NF-?Bp50, as well as NF-?Bp65 and NF-?Bp50 binding, the gene expression of pro-inflammatory mediators under NF-?B control (IL-1?, IL-6, INF-?, TNF-?, MCP-1), and its inhibitory protein, nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (I?B-?). The observational study involved 35 patients (control group, n = 12 and cancer group, n = 23, further divided into cachectic and non-cachectic). NF-?Bp65 and its target genes expression (TNF-?, IL-1?, MCP-1 and I?B-?) were significantly higher in cachectic cancer patients. Moreover, NF-?Bp65 gene expression correlated positively with the expression of its target genes. The results strongly suggest that the NF-?B pathway plays a role in the promotion of WAT inflammation during cachexia. PMID:26053616

  16. NF-?Bp65 and Expression of Its Pro-Inflammatory Target Genes Are Upregulated in the Subcutaneous Adipose Tissue of Cachectic Cancer Patients.

    PubMed

    Camargo, Rodolfo Gonzalez; Riccardi, Daniela Mendes Dos Reis; Ribeiro, Henrique Quintas Teixeira; Carnevali, Luiz Carlos; de Matos-Neto, Emidio Marques; Enjiu, Lucas; Neves, Rodrigo Xavier; Lima, Joanna Darck Carola Correia; Figuerêdo, Raquel Galvão; de Alcântara, Paulo Sérgio Martins; Maximiano, Linda; Otoch, José; Batista, Miguel; Püschel, Gerhard; Seelaender, Marilia

    2015-01-01

    Cancer cachexia, of which the most notable symptom is severe and rapid weight loss, is present in the majority of patients with advanced cancer. Inflammatory mediators play an important role in the development of cachexia, envisaged as a chronic inflammatory syndrome. The white adipose tissue (WAT) is one of the first compartments affected in cancer cachexia and suffers a high rate of lipolysis. It secretes several cytokines capable of directly regulating intermediate metabolism. A common pathway in the regulation of the expression of pro-inflammatory cytokines in WAT is the activation of the nuclear transcription factor kappa-B (NF-?B). We have examined the gene expression of the subunits NF-?Bp65 and NF-?Bp50, as well as NF-?Bp65 and NF-?Bp50 binding, the gene expression of pro-inflammatory mediators under NF-?B control (IL-1?, IL-6, INF-?, TNF-?, MCP-1), and its inhibitory protein, nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (I?B-?). The observational study involved 35 patients (control group, n = 12 and cancer group, n = 23, further divided into cachectic and non-cachectic). NF-?Bp65 and its target genes expression (TNF-?, IL-1?, MCP-1 and I?B-?) were significantly higher in cachectic cancer patients. Moreover, NF-?Bp65 gene expression correlated positively with the expression of its target genes. The results strongly suggest that the NF-?B pathway plays a role in the promotion of WAT inflammation during cachexia. PMID:26053616

  17. Cellular uptake, stability, visualization by 'Naturstoff reagent A', and multidrug resistance protein 1 gene-regulatory activity of cyanidin in human keratinocytes.

    PubMed

    Ernst, I M A; Wagner, A E; Lipinski, S; Skrbek, S; Ruefer, C E; Desel, C; Rimbach, G

    2010-03-01

    There is increasing interest in the role of anthocyanidins as potential skin protective phytochemicals. However, little is known if and to what extent anthocyanidins are taken up by the human skin. In the present study cellular uptake (as determined by HPLC), stability, and gene-regulatory activity of cyanidin were determined in human HaCaT keratinocytes in culture. Using the fluorescent dye Naturstoff reagent A cyanidin was visualized in order to determine its cellular accumulation via flow cytometry and fluorescence microscopy. Cyanidin was rapidly taken up by HaCaT cells at relatively low concentrations. Following incubation, cellular cyanidin levels decreased time-dependently most likely due to degradation into protocatechuic acid and phloroglucinol aldehyde. Confocal laser scanning microscopy data demonstrated that cyanidin was mainly present in the cytoplasm. Cellular uptake of cyanidin was accompanied by an inhibition of multidrug resistance protein 1 (involved in cellular efflux of flavonoids) mRNA-levels indicating its gene-regulatory activity. Naturstoff reagent A seems to be a promising fluorescent dye to visualize cyanidin in keratinocytes. PMID:19897037

  18. Birt-Hogg-Dubé renal tumors are genetically distinct from other renal neoplasias and are associated with up-regulation of mitochondrial gene expression

    PubMed Central

    2010-01-01

    Background Germline mutations in the folliculin (FLCN) gene are associated with the development of Birt-Hogg-Dubé syndrome (BHDS), a disease characterized by papular skin lesions, a high occurrence of spontaneous pneumothorax, and the development of renal neoplasias. The majority of renal tumors that arise in BHDS-affected individuals are histologically similar to sporadic chromophobe renal cell carcinoma (RCC) and sporadic renal oncocytoma. However, most sporadic tumors lack FLCN mutations and the extent to which the BHDS-derived renal tumors share genetic defects associated with the sporadic tumors has not been well studied. Methods BHDS individuals were identified symptomatically and FLCN mutations were confirmed by DNA sequencing. Comparative gene expression profiling analyses were carried out on renal tumors isolated from individuals afflicted with BHDS and a panel of sporadic renal tumors of different subtypes using discriminate and clustering approaches. qRT-PCR was used to confirm selected results of the gene expression analyses. We further analyzed differentially expressed genes using gene set enrichment analysis and pathway analysis approaches. Pathway analysis results were confirmed by generation of independent pathway signatures and application to additional datasets. Results Renal tumors isolated from individuals with BHDS showed distinct gene expression and cytogenetic characteristics from sporadic renal oncocytoma and chromophobe RCC. The most prominent molecular feature of BHDS-derived kidney tumors was high expression of mitochondria-and oxidative phosphorylation (OXPHOS)-associated genes. This mitochondria expression phenotype was associated with deregulation of the PGC-1?-TFAM signaling axis. Loss of FLCN expression across various tumor types is also associated with increased nuclear mitochondrial gene expression. Conclusions Our results support a genetic distinction between BHDS-associated tumors and other renal neoplasias. In addition, deregulation of the PGC-1?-TFAM signaling axis is most pronounced in renal tumors that harbor FLCN mutations and in tumors from other organs that have relatively low expression of FLCN. These results are consistent with the recently discovered interaction between FLCN and AMPK and support a model in which FLCN is a regulator of mitochondrial function. PMID:21162720

  19. KSHV encoded LANA upregulates Pim-1 and is a substrate for its kinase activity

    SciTech Connect

    Bajaj, Bharat G. [Department of Microbiology and the Tumor Virology Program, Abramson Comprehensive Cancer Center, University of Pennsylvania Medical School, 201E Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA 19104 (United States); Verma, Subhash C. [Department of Microbiology and the Tumor Virology Program, Abramson Comprehensive Cancer Center, University of Pennsylvania Medical School, 201E Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA 19104 (United States); Lan, Ke [Department of Microbiology and the Tumor Virology Program, Abramson Comprehensive Cancer Center, University of Pennsylvania Medical School, 201E Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA 19104 (United States); Cotter, Murray A. [Department of Microbiology and the Tumor Virology Program, Abramson Comprehensive Cancer Center, University of Pennsylvania Medical School, 201E Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA 19104 (United States); Woodman, Zenda L. [Department of Microbiology and the Tumor Virology Program, Abramson Comprehensive Cancer Center, University of Pennsylvania Medical School, 201E Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA 19104 (United States); Robertson, Erle S. [Department of Microbiology and the Tumor Virology Program, Abramson Comprehensive Cancer Center, University of Pennsylvania Medical School, 201E Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA 19104 (United States)]. E-mail: erle@mail.med.upenn.edu

    2006-07-20

    Pim kinases are proto-oncogenes that are upregulated in a number of B cell cancers, including Epstein-Barr Virus (EBV) associated Burkitt's lymphoma. They have also been shown to be upregulated in Kaposi sarcoma-associated herpes virus (KSHV) infected primary B cells. Most cells in KSHV-associated tumors are latently infected and express only a small subset of viral genes, with KSHV latency associated nuclear antigen (LANA) being constitutively expressed. LANA regulates the transcription of a large number of cellular and viral genes. Here, we show that LANA upregulates transcription from the Pim-1 promoter (pPim-1) and map this activation to a region in the promoter located within the sequence (-681 to +37). We show that LANA expressing cells can proliferate faster and are better protected from drug induced apoptosis. Since transition through cell cycle check points and anti-apoptosis are functions associated with Pim-1, it is likely that higher Pim-1 expression in cells expressing LANA is responsible, at least in part, for this effect. A Pim-1 phosphorylation site was also identified within the amino-terminal domain of LANA. Using in vitro kinase assays, we confirmed that LANA was indeed a Pim-1 substrate, and the failure of Pim-1 to phosphorylate LANA mutated at SS205/6RR identified this site as the specific serine residues phosphorylated by Pim-1. This report provides valuable insight into yet another cellular signaling pathway subverted by KSHV LANA and suggests a contribution to KSHV related oncogenesis.

  20. OVER-ACCUMULATION OF HIGHER POLYAMINES IN RIPENING TRANSGENIC TOMATO FRUIT REVIVES METABOLIC MEMORY, UPREGULATES ANABOLISM-RELATED GENES, AND POSITIVELY IMPACTS NUTRITIONAL QUALITY

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Modern science is making strides into our understanding of the interrelationships between diet and disease. Thus, functional genomics research that not only deciphers the functional roles of genes but also to understand how a particular diet or a component thereof influences a particular disease has...

  1. Mechanism of enhanced responses after combination photodynamic therapy (cPDT) in carcinoma cells involves C/EBP-mediated transcriptional upregulation of the coproporphyrinogen oxidase (CPO) gene

    NASA Astrophysics Data System (ADS)

    Anand, Sanjay; Hasan, Tayyaba; Maytin, Edward V.

    2013-03-01

    Photodynamic therapy (PDT) with aminolevulinate (ALA) is widely accepted as an effective treatment for superficial carcinomas and pre-cancers. However, PDT is still suboptimal for deeper tumors, mainly due to inadequate ALA penetration and subsequent conversion to PpIX. We are interested in improving the effectiveness of photodynamic therapy (PDT) for deep tumors, using a combination approach (cPDT) in which target protoporphyrin (PpIX) levels are significantly enhanced by differentiation caused by giving Vitamin D or methotrexate (MTX) for 3 days prior to ALAPDT. In LNCaP and MEL cells, a strong correlation between inducible differentiation and expression of C/EBP transcription factors, as well as between differentiation and mRNA levels of CPO (a key heme-synthetic enzyme), indicates the possibility of CPO transcriptional regulation by the C/EBPs. Sequence analysis of the first 1300 base pairs of the murine CPO upstream region revealed 15 consensus C/EBP binding sites. Electrophoretic Mobility Shift Assays (EMSA) proved that these sites form specific complexes that have strong, moderate or weak affinities for C/EBPs. However, in the context of the full-length CPO promoter, inactivation of any type of site (strong or weak) reduced CPO promoter activity (luciferase assay) to nearly the same extent, suggesting cooperative interactions. A comparative analysis of murine and human CPO promoters revealed possible protein-protein interactions between C/EBPs and several neighboring transcription factors such as NFkB, Sp1, AP-1, CBP/p300 and CREB (an enhanceosome complex). Overall, these results confirm that C/EBP's are important for CPO expression via complex mechanisms which upregulate PpIX and enhance the outcome of cPDT.

  2. The Gene Ontology (GO) Cellular Component Ontology: integration with SAO (Subcellular Anatomy Ontology) and other recent developments

    PubMed Central

    2013-01-01

    Background The Gene Ontology (GO) (http://www.geneontology.org/) contains a set of terms for describing the activity and actions of gene products across all kingdoms of life. Each of these activities is executed in a location within a cell or in the vicinity of a cell. In order to capture this context, the GO includes a sub-ontology called the Cellular Component (CC) ontology (GO-CCO). The primary use of this ontology is for GO annotation, but it has also been used for phenotype annotation, and for the annotation of images. Another ontology with similar scope to the GO-CCO is the Subcellular Anatomy Ontology (SAO), part of the Neuroscience Information Framework Standard (NIFSTD) suite of ontologies. The SAO also covers cell components, but in the domain of neuroscience. Description Recently, the GO-CCO was enriched in content and links to the Biological Process and Molecular Function branches of GO as well as to other ontologies. This was achieved in several ways. We carried out an amalgamation of SAO terms with GO-CCO ones; as a result, nearly 100 new neuroscience-related terms were added to the GO. The GO-CCO also contains relationships to GO Biological Process and Molecular Function terms, as well as connecting to external ontologies such as the Cell Ontology (CL). Terms representing protein complexes in the Protein Ontology (PRO) reference GO-CCO terms for their species-generic counterparts. GO-CCO terms can also be used to search a variety of databases. Conclusions In this publication we provide an overview of the GO-CCO, its overall design, and some recent extensions that make use of additional spatial information. One of the most recent developments of the GO-CCO was the merging in of the SAO, resulting in a single unified ontology designed to serve the needs of GO annotators as well as the specific needs of the neuroscience community. PMID:24093723

  3. Immortalization by c-myc, H-ras, and Ela oncogenes induces differential cellular gene expression and growth factor responses

    SciTech Connect

    Kelekar, A.; Cole, M.D.

    1987-11-01

    Early-passage rat kidney cells were immortalized or rescued from senescence with three different oncogenes: viral promoter-driven c-myc, H-ras (Val-12), and adenovirus type 5 E1a. The normal c-myc and H-ras (Gly-12) were unable to immortalize cells under similar conditions. Quantitation of RNA in the ras-immortalized lines demonstrated that the H-ras oncogene was expressed at a level equivalent to that of the normal H-ras gene in established human or rat cell lines. Cell lines immortalized by different oncogenes were found to have distinct growth responses to individual growth factors in a short-term assay. E1a-immortalized cells were largely independent of serum growth factors, whereas c-myc-immortalized cells responded to serum better than to epidermal growth factor and insulin. H-ras-immortalized cells responded significantly to insulin alone and gave a maximal response to epidermal growth factor and insulin. Several cellular genes associated with platelet-derived growth factor stimulation, including c-myc, were expressed at high levels in the H-ras-immortalized cells, and c-myc expression was deregulated, suggesting that the H-ras oncogene has provided a ''competence'' function. H-ras-immortalized cells could not be morphologically transformed by secondary transfection with a long terminal repeat-c-myc oncogene, but secondary transfection of the same cells with H-ras (Val-12) produced morphologically transformed colonies that had 20- to 40-fold higher levels of H-ras oncogene expression. Thus transformation in this system is dependent on high levels of H-ras oncogene expression rather than on the presence of activated H-ras and c-myc oncogenes in the same cell.

  4. Restoration of cellular ubiquitin reverses impairments in neuronal development caused by disruption of the polyubiquitin gene Ubb.

    PubMed

    Ryu, Han-Wook; Park, Chul-Woo; Ryu, Kwon-Yul

    2014-10-24

    Disruption of the polyubiquitin gene Ubb leads to hypothalamic neurodegeneration and metabolic disorders, including obesity and sleep abnormalities, in mice. However, it has yet to be determined whether or not these neural phenotypes in Ubb(-/-) mice are directly caused by cell autonomous defects in maintaining proper levels of ubiquitin (Ub). To directly demonstrate that reduced levels of Ub are sufficient to cause neuronal abnormalities, we investigated the characteristics of cultured neurons isolated from Ubb(-/-) mouse embryonic brains. We found that neuronal morphology, neurite outgrowth, and synaptic development were significantly impaired in Ubb(-/-) neurons. Furthermore, we observed the growth of astrocytes in Ubb(-/-) cell cultures despite the fact that cells were cultured under conditions promoting neuronal growth. When the reduced levels of free Ub, but not Ub conjugates, in Ubb(-/-) cells were restored to those of wild-type cells by providing exogenous Ub via lentivirus-mediated delivery, the increased apoptosis observed in Ubb(-/-) cells was almost completely abolished. Ectopic expression of Ub also improved neuronal and glial phenotypes observed in Ubb(-/-) cells. Therefore, our study suggests that Ub homeostasis, or the maintenance of cellular free Ub above certain threshold levels, is essential for proper neuronal development and survival. PMID:25280998

  5. Transposition of a reconstructed Harbinger element in human cells and functional homology with two transposon-derived cellular genes

    PubMed Central

    Sinzelle, Ludivine; Kapitonov, Vladimir V.; Grzela, Dawid P.; Jursch, Tobias; Jurka, Jerzy; Izsvák, Zsuzsanna; Ivics, Zoltán

    2008-01-01

    Ancient, inactive copies of transposable elements of the PIF/Harbinger superfamily have been described in vertebrates. We reconstructed components of the Harbinger3_DR transposon in zebrafish, including a transposase and a second, transposon-encoded protein that has a Myb-like trihelix domain. The reconstructed Harbinger transposon shows efficient cut-and-paste transposition in human cells and preferentially inserts into a 15-bp consensus target sequence. The Myb-like protein is required for transposition and physically interacts with the N-terminal region of the transposase via its C-terminal domain. The Myb-like protein enables transposition in part by promoting nuclear import of the transposase, by directly binding to subterminal regions of the transposon, and by recruiting the transposase to the transposon ends. We investigated the functions of two transposon-derived human proteins: HARBI1, a domesticated transposase-derived protein, and NAIF1, which contains a trihelix motif similar to that described in the Myb-like protein. Physical interaction, subcellular localization, and DNA-binding activities of HARBI1 and NAIF1 suggest strong functional homologies between the Harbinger3_DR system and their related, host-encoded counterparts. The Harbinger transposon will serve as a useful experimental system for transposon biology and for investigating the enzymatic functions of domesticated, transposon-derived cellular genes. PMID:18339812

  6. Targeted mutation of the gene for cellular glutathione peroxidase (Gpx1) increases noise-induced hearing loss in mice.

    PubMed

    Ohlemiller, K K; McFadden, S L; Ding, D L; Lear, P M; Ho, Y S

    2000-11-01

    Reactive oxygen species (ROS) and oxidative stress have been implicated in cochlear injury following loud noise and ototoxins. Genetic mutations that impair antioxidant defenses would be expected to increase cochlear injury following acute insults and to contribute to cumulative injury that presents as age-related hearing loss. We examined whether genetically based deficiency of cellular glutathione peroxidase, a major antioxidant enzyme, increases noise-induced hearing loss in mice. Two-month-old "knockout" mice with a targeted inactivating mutation of the gene coding for glutathione peroxidase (Gpx1) and wild type controls were exposed to broadband noise for one hour at 110 dB SPL. Auditory brainstem response (ABR) thresholds at test frequencies ranging from 5 to 40 kHz were obtained two and four weeks after exposure to determine the stable permanent component of the hearing loss. Depending on test frequency, (compared with controls) Gpx1 knockout mice showed up to 16 dB higher ABR thresholds prior to noise exposure, and up to 15 dB greater noise-induced hearing loss, compared with normal control. Within the cochlear base, there was also a significant contribution of the knockout to inner and outer hair cell loss, as well as nerve fiber loss. Our results support a link between genetic impairment of antioxidant defenses, vulnerability of the cochlea injury, and cochlear degeneration. Such impairment produces characteristics expected of some mutations associated with age-related hearing loss and offers one possible mechanism for their action. PMID:11545230

  7. Host-pathogen interactions: Host resistance factor Nramp1 up-regulates the expression of Salmonella pathogenicity island-2 virulence genes

    Microsoft Academic Search

    Michelle L. Zaharik; Bruce A. Vallance; José L. Puente; Philippe Gros; B. Brett Finlay

    2002-01-01

    Nramp1 (Natural resistance-associated macrophage protein-1; also known as Slc11a1) is a host resistance gene that provides protection against several intracellular pathogens, including Salmonella enterica serovar Typhimurium. Little is known about the dynamic interplay that occurs between mammalian host resistance determinants such as Nramp1 and pathogens during infection. To explore these interactions, we examined the effect of Nramp1 on expression of

  8. The expression of MaEXP1, a Melilotus alba expansin gene, is upregulated during the sweetclover-Sinorhizobium meliloti interaction.

    PubMed

    Giordano, Walter; Hirsch, Ann M

    2004-06-01

    Expansins are a highly conserved group of cell wall-localized proteins that appear to mediate changes in cell wall plasticity during cell expansion or differentiation. The accumulation of expansin protein or the mRNA for specific expansin gene family members has been correlated with the growth of various plant organs. Because expansin proteins are closely associated with plant cell wall expansion, and as part of a larger study to determine the role of different gene products in the legume-Rhizobium spp. symbiosis, we investigated whether a Melilotus alba (white sweetclover) expansin gene is expressed during nodule development. A cDNA fragment encoding an expansin gene (EXP) was isolated from Sinorhizobium meliloti-inoculated sweetclover root RNA by reverse-transcriptase polymerase chain reaction using degenerate primers, and a full-length sweetclover expansin sequence (MaEXP1) was obtained using 5' and 3' rapid amplification of cDNA end cloning. The predicted amino acid of the sweetclover expansin is highly conserved with the various alpha-expansins in the GenBank database. MaEXP1 contains a series of eight cysteines and four tryptophans that are conserved in the alpha-expansin protein family. Northern analysis and whole-mount in situ hybridization analyses indicate that MaEXP1 mRNA expression is enhanced in roots within hours after inoculation with S. meliloti and in nodules. Western and immunolocalization studies using a cucumber expansin antibody demonstrated that a cross-reacting protein accumulated in the expanding cells of the nodule. PMID:15195944

  9. Targeting Neurons of Rat Nucleus Tractus Solitarii with the Gene Transfer Vector Adeno-Associated Virus Type 2 to Up-Regulate Neuronal Nitric Oxide Synthase

    Microsoft Academic Search

    Li-Hsien LinDeidre; Deidre Nitschke Dragon; Jingwen Jin; William T. Talman

    2011-01-01

    Adeno-associated virus (AAV) has distinct advantages over other viral vectors in delivering genes of interest to the brain.\\u000a AAV mainly transfects neurons, produces no toxicity or inflammatory responses, and yields long-term transgene expression.\\u000a In this study, we first tested the hypothesis that AAV serotype 2 (AAV2) selectively transfects neurons but not glial cells\\u000a in the nucleus tractus solitarii (NTS) by

  10. Obesity, Independent of p53 Gene Dosage, Promotes Mammary Tumor Progression and Upregulates the p53 Regulator MicroRNA-504

    PubMed Central

    Ford, Nikki A.; Dunlap, Sarah M.; Wheatley, Karrie E.; Hursting, Stephen D.

    2013-01-01

    Obesity, prevalent in >35% of US women, is an established risk and progression factor for postmenopausal breast cancer, and strategies to break the obesity-breast cancer link are urgently needed. Approximately 30% of breast cancers carry p53 tumor suppressor gene alterations; however, the effects of obesity on breast cancer progression in relation to p53 gene dosage are unclear. Using murine models of postmenopausal breast cancer, we characterized the interactive effects of diet-induced obesity (DIO) and p53 gene dosage on mammary tumor growth and associated p53-related regulatory mechanisms. Ovariectomized C57BL/6 mice were randomly assigned to receive a DIO or control diet, and (at 10 weeks) orthotopic injection of MMTV-Wnt-1 p53+/? or MMTV-Wnt-1 p53+/+ mammary tumor cells (n?=?20 mice per diet and genotype group). DIO and control diets produced distinct phenotypes (mean percent body fat at 10 weeks: 57% and 39%, respectively, P<0.001). Regardless of phenotype, time to first palpable tumor was 57% less for Wnt-1 p53+/? than Wnt-1 p53+/+ tumors. Regardless of tumoral p53 genotype, DIO (relative to control) increased tumor burden, tumor cell proliferation (Ki-67), severity of tumor pathology, local tissue invasion, epithelial-to-mesenchymal transition (EMT) programming, and tumoral microRNA-504 (a negative regulator of p53) expression; and suppressed p53, p21, and estrogen receptor-alpha protein expression. These findings in murine models of postmenopausal breast cancer suggest that obesity may augment procancer effects related to p53 gene alterations. Furthermore, microRNA-504, an obesity-responsive negative regulator of p53 and putative EMT regulator, may represent a novel molecular target for breaking the obesity-breast cancer link. PMID:23840816

  11. Expression of human kinase suppressor of Ras 2 (hKSR-2) gene in HL60 leukemia cells is directly upregulated by 1,25-dihydroxyvitamn D3 and is required for optimal cell differentiation

    PubMed Central

    Wang, Xuening; Wang, Tian-Tian; White, John H.; Studzinski, George P.

    2012-01-01

    Induction of terminal differentiation of neoplastic cells offers potential for a novel approach to cancer therapy. One of the agents being investigated for this purpose in preclinical studies is 1,25-dihydroxyvitamin D3 (1,25D), which can convert myeloid leukemia cells into normal monocyte-like cells, but the molecular mechanisms underlying this process are not fully understood. Here, we report that 1,25D upregulates the expression of hKSR-2, a new member of a small family of proteins that exhibit evolutionarily conserved function of potentiating ras signaling. The upregulation of hKSR-2 is direct, as it occurs in the presence of cycloheximide, and occurs primarily at the transcriptional level, via activation of vitamin D receptor, which acts as a ligand-activated transcription factor. Two VDRE-type motifs identified in the hKSR-2 gene bind VDR-RXR alpha heterodimers present in nuclear extracts of 1,25D-treated HL60 cells, and chromatin immunoprecipitation assays show that these VDRE motifs bind VDR in 1,25D-dependent manner in intact cells, coincident with the recruitment of RNA polymerase II to these motifs. Treatment of the cells with siRNA to hKSR-2 reduced the proportion of the most highly differentiated cells in 1,25D-treated cultures. These results demonstrate that hKSR-2 is a direct target of 1,25D in HL60 cells, and is required for optimal monocytic differentiation. PMID:17599832

  12. Expression of human kinase suppressor of Ras 2 (hKSR-2) gene in HL60 leukemia cells is directly upregulated by 1,25-dihydroxyvitamin D(3) and is required for optimal cell differentiation.

    PubMed

    Wang, Xuening; Wang, Tian-Tian; White, John H; Studzinski, George P

    2007-08-15

    Induction of terminal differentiation of neoplastic cells offers potential for a novel approach to cancer therapy. One of the agents being investigated for this purpose in preclinical studies is 1,25-dihydroxyvitamin D(3) (1,25D), which can convert myeloid leukemia cells into normal monocyte-like cells, but the molecular mechanisms underlying this process are not fully understood. Here, we report that 1,25D upregulates the expression of hKSR-2, a new member of a small family of proteins that exhibit evolutionarily conserved function of potentiating ras signaling. The upregulation of hKSR-2 is direct, as it occurs in the presence of cycloheximide, and occurs primarily at the transcriptional level, via activation of vitamin D receptor, which acts as a ligand-activated transcription factor. Two VDRE-type motifs identified in the hKSR-2 gene bind VDR-RXR alpha heterodimers present in nuclear extracts of 1,25D-treated HL60 cells, and chromatin immunoprecipitation assays show that these VDRE motifs bind VDR in 1,25D-dependent manner in intact cells, coincident with the recruitment of RNA polymerase II to these motifs. Treatment of the cells with siRNA to hKSR-2 reduced the proportion of the most highly differentiated cells in 1,25D-treated cultures. These results demonstrate that hKSR-2 is a direct target of 1,25D in HL60 cells, and is required for optimal monocytic differentiation. PMID:17599832

  13. Expression of human kinase suppressor of Ras 2 (hKSR-2) gene in HL60 leukemia cells is directly upregulated by 1,25-dihydroxyvitamin D{sub 3} and is required for optimal cell differentiation

    SciTech Connect

    Wang Xuening [Department of Pathology and Laboratory Medicine, UMDNJ-New Jersey Medical School, 185 South Orange Avenue, C-543, Newark, NJ 07103 (United States); Wang, T.-T. [Department of Physiology, McGill University, Montreal, Quebec (Canada); White, John H. [Department of Physiology, McGill University, Montreal, Quebec (Canada); Studzinski, George P. [Department of Pathology and Laboratory Medicine, UMDNJ-New Jersey Medical School, 185 South Orange Avenue, C-543, Newark, NJ 07103 (United States)]. E-mail: studzins@umdnj.edu

    2007-08-15

    Induction of terminal differentiation of neoplastic cells offers potential for a novel approach to cancer therapy. One of the agents being investigated for this purpose in preclinical studies is 1,25-dihydroxyvitamin D{sub 3} (1,25D), which can convert myeloid leukemia cells into normal monocyte-like cells, but the molecular mechanisms underlying this process are not fully understood. Here, we report that 1,25D upregulates the expression of hKSR-2, a new member of a small family of proteins that exhibit evolutionarily conserved function of potentiating ras signaling. The upregulation of hKSR-2 is direct, as it occurs in the presence of cycloheximide, and occurs primarily at the transcriptional level, via activation of vitamin D receptor, which acts as a ligand-activated transcription factor. Two VDRE-type motifs identified in the hKSR-2 gene bind VDR-RXR alpha heterodimers present in nuclear extracts of 1,25D-treated HL60 cells, and chromatin immunoprecipitation assays show that these VDRE motifs bind VDR in 1,25D-dependent manner in intact cells, coincident with the recruitment of RNA polymerase II to these motifs. Treatment of the cells with siRNA to hKSR-2 reduced the proportion of the most highly differentiated cells in 1,25D-treated cultures. These results demonstrate that hKSR-2 is a direct target of 1,25D in HL60 cells, and is required for optimal monocytic differentiation.

  14. Age-associated reduction of cellular spreading/mechanical force up-regulates matrix metalloproteinase-1 expression and collagen fibril fragmentation via c-Jun/AP-1 in human dermal fibroblasts

    PubMed Central

    Qin, Zhaoping; Voorhees, John J; Fisher, Gary J; Quan, Taihao

    2014-01-01

    The dermal compartment of human skin is largely composed of dense collagen-rich fibrils, which provide structural and mechanical support. Skin dermal fibroblasts, the major collagen-producing cells, are interact with collagen fibrils to maintain cell spreading and mechanical force for function. A characteristic feature of aged human skin is fragmentation of collagen fibrils, which is initiated by matrix metalloproteinase 1 (MMP-1). Fragmentation impairs fibroblast attachment and thereby reduces spreading. Here, we investigated the relationship among fibroblast spreading, mechanical force, MMP-1 expression, and collagen fibril fragmentation. Reduced fibroblast spreading due to cytoskeletal disruption was associated with reduced cellular mechanical force, as determined by atomic force microscopy. These reductions substantially induced MMP-1 expression, which led to collagen fibril fragmentation and disorganization in three-dimensional collagen lattices. Constraining fibroblast size by culturing on slides coated with collagen micropatterns also significantly induced MMP-1 expression. Reduced spreading/mechanical force induced transcription factor c-Jun and its binding to a canonical AP-1 binding site in the MMP-1 proximal promoter. Blocking c-Jun function with dominant negative mutant c-Jun significantly reduced induction of MMP-1 expression in response to reduced spreading/mechanical force. Furthermore, restoration of fibroblast spreading/mechanical force led to decline of c-Jun and MMP-1 levels and eliminated collagen fibril fragmentation and disorganization. These data reveal a novel mechanism by which alteration of fibroblast shape/mechanical force regulates c-Jun/AP-1-dependent expression of MMP-1 and consequent collagen fibril fragmentation. This mechanism provides a foundation for understanding the cellular and molecular basis of age-related collagen fragmentation in human skin. PMID:25201474

  15. The TLR4 D299G and T399I SNPs are constitutively active to up-regulate expression of Trif-dependent genes.

    PubMed

    Hold, Georgina L; Berry, Susan; Saunders, Karin A; Drew, Janice; Mayer, Claus; Brookes, Heather; Gay, Nick J; El-Omar, Emad M; Bryant, Clare E

    2014-01-01

    Dysregulated Toll-Like Receptor (TLR) signalling and genetic polymorphisms in these proteins are linked to many human diseases. We investigated TLR4 functional variants D299G and T399I to assess the impact on LPS-induced responsiveness in comparison to wild-type TLR4. The mechanism by which this occurs in unclear as these SNPs do not lie within the lipid A binding domain or dimerisation sites of the LPS-TLR4/MD2 receptor complexes. Transfection of TLR4D299G, TLR4T399I or TLR4D299G. T399I into HEK cells resulted in constitutive activation of an NF-?B reporter gene and a blunting of the LPS-induced reporter activation compared to WT-TLR4. Unstimulated human monocyte/macrophages, from patients with the D299G and T399I SNPs demonstrated a downregulation of many genes, particularly Tram/Trif signalling pathway constitutents compared to the TLR4 wild-type subjects supporting the concept of basal receptor activity. Monocyte/macrophages from carriers of the TLR4 D299G and T399I polymorphisms stimulated with LPS showed >6 fold lower levels of NF-?B and ?12 fold higher IFN-? gene expression levels compared to wild-type subjects (P<0.05; MWU test) and dramatically altered resultant cytokine profiles. We conclude that these TLR4 SNPs affect constitutive receptor activity which impacts on the hosts ability to respond to LPS challenge leading to a dysregulated sub-optimal immune response to infection. PMID:25365308

  16. The TLR4 D299G and T399I SNPs Are Constitutively Active to Up-Regulate Expression of Trif-Dependent Genes

    PubMed Central

    Hold, Georgina L.; Berry, Susan; Saunders, Karin A.; Drew, Janice; Mayer, Claus; Brookes, Heather; Gay, Nick J.; El-Omar, Emad M.; Bryant, Clare E.

    2014-01-01

    Dysregulated Toll-Like Receptor (TLR) signalling and genetic polymorphisms in these proteins are linked to many human diseases. We investigated TLR4 functional variants D299G and T399I to assess the impact on LPS-induced responsiveness in comparison to wild-type TLR4. The mechanism by which this occurs in unclear as these SNPs do not lie within the lipid A binding domain or dimerisation sites of the LPS-TLR4/MD2 receptor complexes. Transfection of TLR4D299G, TLR4T399I or TLR4D299G. T399I into HEK cells resulted in constitutive activation of an NF-?B reporter gene and a blunting of the LPS-induced reporter activation compared to WT-TLR4. Unstimulated human monocyte/macrophages, from patients with the D299G and T399I SNPs demonstrated a downregulation of many genes, particularly Tram/Trif signalling pathway constitutents compared to the TLR4 wild-type subjects supporting the concept of basal receptor activity. Monocyte/macrophages from carriers of the TLR4 D299G and T399I polymorphisms stimulated with LPS showed >6 fold lower levels of NF-?B and ?12 fold higher IFN-? gene expression levels compared to wild-type subjects (P<0.05; MWU test) and dramatically altered resultant cytokine profiles. We conclude that these TLR4 SNPs affect constitutive receptor activity which impacts on the hosts ability to respond to LPS challenge leading to a dysregulated sub-optimal immune response to infection. PMID:25365308

  17. 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 [Laboratory of Radiobiology and Experimental Radiooncology, University Hospital Hamburg-Eppendorf, Hamburg (Germany)] [Laboratory of Radiobiology and Experimental Radiooncology, University Hospital Hamburg-Eppendorf, Hamburg (Germany); Bonin, Michael [Department of Medical Genetics, University of Tuebingen (Germany)] [Department of Medical Genetics, University of Tuebingen (Germany); Petersen, Cordula [Clinic of Radiotherapy and Radiooncology, University Hospital Hamburg-Eppendorf, Hamburg (Germany)] [Clinic of Radiotherapy and Radiooncology, University Hospital Hamburg-Eppendorf, Hamburg (Germany); Dikomey, Ekkehard, E-mail: dikomey@uke.de [Laboratory of Radiobiology and Experimental Radiooncology, University Hospital Hamburg-Eppendorf, Hamburg (Germany)] [Laboratory of Radiobiology and Experimental Radiooncology, University Hospital Hamburg-Eppendorf, Hamburg (Germany); Raabe, Annette [Laboratory of Radiobiology and Experimental Radiooncology, University Hospital Hamburg-Eppendorf, Hamburg (Germany)] [Laboratory of Radiobiology and Experimental Radiooncology, University Hospital Hamburg-Eppendorf, Hamburg (Germany)

    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.

  18. Rhodnius prolixus: identification of immune-related genes up-regulated in response to pathogens and parasites using suppressive subtractive hybridization.

    PubMed

    Ursic-Bedoya, Raul J; Lowenberger, Carl A

    2007-01-01

    We report the identification of immune-related molecules from the fat body, and intestine of Rhodnius prolixus, an important vector of Chagas disease. Insects were challenged by introducing pathogens or Trypanosoma cruzi, the parasite that causes Chagas disease, into the hemocoel. RNA from intestines, or fat body were isolated 24h after stimulation. We used suppressive subtractive hybridization to identify immune-related genes, generated three subtracted libraries, sequenced the clones and assembled the sequences. The functional annotation revealed expressed sequence tags (ESTs) generated in response to various stimuli in all tissues, and included pathogen recognition molecules, regulatory molecules, and effector molecules. PMID:16824597

  19. KRIT1 loss of function causes a ROS-dependent upregulation of c-Jun

    PubMed Central

    Goitre, Luca; De Luca, Elisa; Braggion, Stefano; Trapani, Eliana; Guglielmotto, Michela; Biasi, Fiorella; Forni, Marco; Moglia, Andrea; Trabalzini, Lorenza; Retta, Saverio Francesco

    2014-01-01

    Loss-of-function mutations in the KRIT1 gene (CCM1) have been associated with the pathogenesis of cerebral cavernous malformations (CCM), a major cerebrovascular disease. However, KRIT1 functions and CCM pathogenetic mechanisms remain incompletely understood. Indeed, recent experiments in animal models have clearly demonstrated that the homozygous loss of KRIT1 is not sufficient to induce CCM lesions, suggesting that additional factors are necessary to cause CCM disease. Previously, we found that KRIT1 is involved in the maintenance of the intracellular reactive oxygen species (ROS) homeostasis to prevent ROS-induced cellular dysfunctions, including a reduced ability to maintain a quiescent state. Here, we show that KRIT1 loss of function leads to enhanced expression and phosphorylation of the redox-sensitive transcription factor c-Jun, as well as induction of its downstream target COX-2, in both cellular models and human CCM tissues. Furthermore, we demonstrate that c-Jun upregulation can be reversed by either KRIT1 re-expression or ROS scavenging, whereas KRIT1 overexpression prevents forced upregulation of c-Jun induced by oxidative stimuli. Taken together with the reported role of c-Jun in vascular dysfunctions triggered by oxidative stress, our findings shed new light on the molecular mechanisms underlying KRIT1 function and CCM pathogenesis. PMID:24291398

  20. Gibberellin regulates Arabidopsis seed germination via RGL2, a GAI/RGA-like gene whose expression is up-regulated following imbibition

    PubMed Central

    Lee, Sorcheng; Cheng, Hui; King, Kathryn E.; Wang, Weefuen; He, Yawen; Hussain, Alamgir; Lo, Jane; Harberd, Nicholas P.; Peng, Jinrong

    2002-01-01

    The germination of Arabidopsis seeds is promoted by gibberellin (GA). Arabidopsis GAI, and RGA are genes encoding key GA signal-transduction components (GAI and RGA) that mediate GA regulation of stem elongation. The Arabidopsis genome contains two further genes, RGL1 and RGL2, that encode proteins (RGL1 and RGL2) that are closely related to GAI and RGA. Here, we show that RGL2 regulates seed germination in response to GA, and that RGL1, GAI, and RGA do not. In addition, we show that RGL2 transcript levels rise rapidly following seed imbibition, and then decline rapidly as germination proceeds. In situ GUS staining revealed that RGL2 expression in imbibed seeds is restricted to elongating regions of pre-emergent and recently emerged radicles. These observations indicate that RGL2 is a negative regulator of GA responses that acts specifically to control seed germination rather than stem elongation. Furthermore, as RGL2 expression is imbibition inducible, RGL2 may function as an integrator of environmental and endogenous cues to control seed germination. PMID:11877383

  1. DNA immunization with a plasmid carrying the gene of hepatitis C virus protein 5A (NS5A) induces an effective cellular immune response

    Microsoft Academic Search

    O. V. Masalova; E. I. Lesnova; V. V. Grabovetskii; O. A. Smirnova; T. I. Ulanova; A. N. Burkov; A. V. Ivanov; A. D. Zaberezhnyi; R. I. Ataullakhanov; A. A. Kushch

    2010-01-01

    In spite of extensive research, no effective vaccine against hepatitis C virus (HCV) has been developed so far. DNA immunization\\u000a is a potent technique of vaccine design strongly promoting the cellular arm of immune response. The genes encoding nonstructural\\u000a HCV proteins (NS2-NS5B) are promising candidates for vaccine development. NS5A is a protein involved in viral pathogenesis,\\u000a in the induction of

  2. Hepatitis C virus ARFP\\/F protein interacts with cellular MM1 protein and enhances the gene trans-activation activity of c-Myc

    Microsoft Academic Search

    Hsin-Chieh Ma; Ta-Wei Lin; Huichun Li; Sanae M. M. Iguchi-Ariga; Hiroyoshi Ariga; Yu-Li Chuang; Jing-Hsiung Ou; Shih-Yen Lo

    2008-01-01

    The ARFP\\/F protein is synthesized from the +1 reading frame of the hepatitis C virus (HCV) core protein gene. The function\\u000a of this protein remains unknown. To study the function of the HCV ARFP\\/F protein, we have conducted the yeast two-hybrid screening\\u000a experiment to identify cellular proteins that may interact with the ARFP\\/F protein. MM-1, a c-Myc interacting protein, was

  3. Transmissible Gastroenteritis Virus (TGEV) Infection Alters the Expression of Cellular MicroRNA Species That Affect Transcription of TGEV Gene 7

    PubMed Central

    Song, Xiangjun; Zhao, Xiaomin; Huang, Yong; Xiang, Hailing; Zhang, Wenlong; Tong, Dewen

    2015-01-01

    Transmissible gastroenteritis virus (TGEV) is a member of Coronaviridae family. TGEV infection has emerged as a major cause of severe gastroenteritis and leads to alterations of many cellular processes. Meanwhile, the pathogenic mechanism of TGEV is still unclear. microRNAs (miRNAs) are a novel class of small non-coding RNAs which are involved in the regulation of numerous biological processes such as viral infection and cell apoptosis. Accumulating data show that miRNAs are involved in the process of coronavirus infection such as replication of severe acute respiratory syndrome coronavirus (SARS-CoV). However, the link between miRNAs and TGEV infection is unknown. In this study, we performed microRNA microarray assay and predicted targets of altered miRNAs. The results showed TGEV infection caused the change of miRNAs profile. Then we selected miR-4331 for further analysis and subsequently identified cell division cycle-associated protein 7 (CDCA7) as the target of miR-4331. Moreover, miR-4331 showed the ability to inhibit transcription of TGEV gene 7 (a non-structure gene) via directly targeting CDCA7. In conclusion, differentially expressed miR-4331 that is caused by TGEV infection can suppress transcription of TGEV gene 7 via targeting cellular CDCA7. Our key finding is that TGEV selectively manipulates the expression of some cellular miRNAs to regulate its subgenomic transcription.

  4. miR-24-2 regulates genes in survival pathway and demonstrates potential in reducing cellular viability in combination with docetaxel.

    PubMed

    Manvati, Siddharth; Mangalhara, Kailash Chandra; Kalaiarasan, P; Srivastava, Niloo; Bamezai, R N K

    2015-08-10

    MicroRNAs the small (18-22 in length) noncoding RNA molecules are negative regulators of gene expression, modulating biological processes of cell differentiation, survival and death. The latter two phenomena are critical in tumour biology. We provide here the results of human genome wide target prediction of one such microRNA, hsa-miR-24-2, shown to target genes essential for initiating cellular stability and cell survival. The protein-protein interaction study showed important nodes which could affect cell cycle progression and differential oncogenesis. An analysis of hsa-miR-24-2 in sporadic breast tumours showed a negative correlation with metastasis and increasing nodes. The conclusion drawn of hsa-miR-24-2 targeting the genes of cell survival correlated with the methylation profile and resultant transcription factor binding site gain or loss in support of absence of cell survival. In order to accentuate the potential of hsa-miR-24-2 to reduce cellular viability under experimental conditions, in vitro studies in the presence and absence of anti-cancer drugs, such as docetaxel resulted in a significant decrease in cellular viability even at a 200-fold reduced dose of the drug in combination with hsa-miR-24-2. PMID:25943634

  5. NMDA-stimulated ERK1/2 Signaling and the Transcriptional Up-regulation of Plasticity-related Genes are Developmentally Regulated following in vitro Neuronal Maturation

    PubMed Central

    Zhou, Xianju; Moon, Changjong; Zheng, Fei; Luo, Yongneng; Soellner, Deborah; Nuñez, Joseph L.; Wang, Hongbing

    2010-01-01

    The general features of neuroplasticity are developmentally regulated. Although it has been hypothesized that the loss of plasticity in mature neurons may be due to synaptic saturation and functional reduction of NMDA receptors (NMDAR), the molecular mechanisms remain largely unknown. We examined the effects of NMDAR activation and KCl-mediated membrane depolarization on ERK1/2 signaling following in vitro maturation of cultured cortical neurons. Although NMDA stimulated robust increase of intracellular calcium at both DIV (day in vitro) 3 and 14, the activation of ERK1/2 and CREB was impaired at DIV 14. Specifically, the phosphorylation of ERK1/2 was stimulated by both NMDA and KCl at DIV 3. However, at DIV 14, NMDA-, but not KCl-stimulated ERK1/2 and CREB phosphorylation was significantly diminished. Consistently, the NMDA-induced transcription of ERK/CREB-regulated genes Bdnf exon 4, Arc and zif268 was significantly attenuated at DIV 14. Moreover, compared to DIV 3 neurons, the basal level of phosphorylated-ERK1/2 in DIV 14 neurons increased tremendously following maturation, and was more susceptible to dephosphorylation. Blocking calcium channels by nifedipine or NMDAR by APV caused more dramatic ERK dephosphorylation in DIV 14 neurons. We further demonstrate that the loss of plasticity-related signaling is unrelated to NMDA-induced cell death of the DIV 14 neurons. Taken together, these results suggest that the attenuation of certain aspects of neuroplasticity following maturation may be due to the reduction of NMDAR-mediated gene transcription and a saturation of ERK1/2 activity. PMID:19396876

  6. Abscisic acid prevents the coalescence of protein storage vacuoles by upregulating expression of a tonoplast intrinsic protein gene in barley aleurone.

    PubMed

    Lee, Sung-eun; Yim, Hui-kyung; Lim, Mi-na; Yoon, In sun; Kim, Jeong hoe; Hwang, Yong-sic

    2015-03-01

    Tonoplast intrinsic proteins (TIPs) are integral membrane proteins that are known to function in plants as aquaporins. Here, we propose another role for TIPs during the fusion of protein storage vacuoles (PSVs) in aleurone cells, a process that is promoted by gibberellic acid (GA) and prevented by abscisic acid (ABA). Studies of the expression of barley (Hordeum vulgare) TIP genes (HvTIP) showed that GA specifically decreased the abundance of HvTIP1;2 and HvTIP3;1 transcripts, while ABA strongly increased expression of HvTIP3;1. Increased or decreased expression of HvTIP3;1 interfered with the hormonal effects on vacuolation in aleurone protoplasts. HvTIP3;1 gain-of-function experiments delayed GA-induced vacuolation, whereas HvTIP3;1 loss-of-function experiments promoted vacuolation in ABA-treated aleurone cells. These results indicate that TIP plays a key role in preventing the coalescence of small PSVs in aleurone cells. Hormonal regulation of the HvTIP3;1 promoter is similar to the regulation of the endogenous gene, indicating that induction of the transcription of HvTIP3;1 by ABA is a critical factor in the prevention of PSV coalescence in response to ABA. Promoter analysis using deletions and site-directed mutagenesis of sequences identified three cis-acting elements that are responsible for ABA responsiveness in the HvTIP3;1 promoter. Promoter analysis also showed that ABA responsiveness of the HvTIP3;1 promoter is likely to occur via a unique regulatory system distinct from that involving the ABA-response promoter complexes. PMID:25477530

  7. 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 viral replication, have proven to be directly involved in viral spread in vivo and represent potential targets for therapeutic intervention against HTLV-1 infection and disease. PMID:15358581

  8. Decreased cellular cholesterol efflux is a common cause of familial hypoalphalipoproteinemia: role of the ABCA1 gene mutations

    Microsoft Academic Search

    Stephanie Mott; Lu Yu; Michel Marcil; Betsie Boucher; Colette Rondeau; Jacques Genest Jr

    2000-01-01

    Background. High density lipoproteins (HDL) are complex lipoprotein particles involved in reverse cholesterol (C) transport and are negatively associated with the risk for coronary artery disease (CAD). We have described a disorder of familial HDL deficiency (FHD) due to abnormal cellular cholesterol efflux. In the present study, we investigated cellular cholesterol efflux on skin fibroblast from 15 probands with moderate

  9. The Charcot-Marie-Tooth type 2A gene product, Mfn2, up-regulates fuel oxidation through expression of OXPHOS system.

    PubMed

    Pich, Sara; Bach, Daniel; Briones, Paz; Liesa, Marc; Camps, Marta; Testar, Xavier; Palacín, Manuel; Zorzano, Antonio

    2005-06-01

    Mitofusin-2 (Mfn2) is a mitochondrial membrane protein that participates in mitochondrial fusion in mammalian cells and mutations in the Mfn2 gene cause Charcot-Marie-Tooth neuropathy type 2A. Here, we show that Mfn2 loss-of-function inhibits pyruvate, glucose and fatty acid oxidation and reduces mitochondrial membrane potential, whereas Mfn2 gain-of-function increases glucose oxidation and mitochondrial membrane potential. As to the mechanisms involved, we have found that Mfn2 loss-of-function represses nuclear-encoded subunits of OXPHOS complexes I, II, III and V, whereas Mfn2 overexpression induced the subunits of complexes I, IV and V. Obesity-induced Mfn2 deficiency in rat skeletal muscle was also associated with a decrease in the subunits of complexes I, II, III and V. In addition, the effect of Mfn2 overexpression on mitochondrial metabolism was mimicked by a truncated Mfn2 mutant that is inactive as a mitochondrial fusion protein. Our results indicate that Mfn2 triggers mitochondrial energization, at least in part, by regulating OXPHOS expression through signals that are independent of its role as a mitochondrial fusion protein. PMID:15829499

  10. New 2-(aryloxy)-3-phenylpropanoic acids as peroxisome proliferator-activated receptor ?/? dual agonists able to upregulate mitochondrial carnitine shuttle system gene expression.

    PubMed

    Laghezza, A; Pochetti, G; Lavecchia, A; Fracchiolla, G; Faliti, S; Piemontese, L; Di Giovanni, C; Iacobazzi, V; Infantino, V; Montanari, R; Capelli, D; Tortorella, P; Loiodice, F

    2013-01-10

    The preparation of a series of 2-(aryloxy)-3-phenylpropanoic acids, resulting from the introduction of different substituents into the biphenyl system of the previously reported peroxisome proliferator-activated receptor ?/? (PPAR?/?) dual agonist 1, allowed the identification of new ligands with higher potency on PPAR? and fine-tuned moderate PPAR? activity. For the most promising stereoisomer (S)-16, X-ray and calorimetric studies in PPAR? revealed, at high ligand concentration, the presence of two molecules simultaneously bound to the receptor. On the basis of these results and docking experiments in both receptor subtypes, a molecular explanation was provided for its different behavior as a full and partial agonist of PPAR? and PPAR?, respectively. The effects of (S)-16 on mitochondrial acylcarnitine carrier and carnitine-palmitoyl-transferase 1 gene expression, two key components of the carnitine shuttle system, were also investigated, allowing the hypothesis of a more beneficial pharmacological profile of this compound compared to the less potent PPAR? agonist fibrates currently used in therapy. PMID:23171045

  11. miR-148a is upregulated by Twist1 and T-bet and promotes Th1-cell survival by regulating the proapoptotic gene Bim

    PubMed Central

    Haftmann, Claudia; Stittrich, Anna-Barbara; Zimmermann, Jakob; Fang, Zhuo; Hradilkova, Kristyna; Bardua, Markus; Westendorf, Kerstin; Heinz, Gitta A; Riedel, René; Siede, Julia; Lehmann, Katrin; Weinberger, Esther E; Zimmel, David; Lauer, Uta; Häupl, Thomas; Sieper, Joachim; Backhaus, Marina; Neumann, Christian; Hoffmann, Ute; Porstner, Martina; Chen, Wei; Grün, Joachim R; Baumgrass, Ria; Matz, Mareen; Löhning, Max; Scheffold, Alexander; Wittmann, Jürgen; Chang, Hyun-Dong; Rajewsky, Nikolaus; Jäck, Hans-Martin; Radbruch, Andreas; Mashreghi, Mir-Farzin

    2015-01-01

    Repeatedly activated T helper 1 (Th1) cells present during chronic inflammation can efficiently adapt to the inflammatory milieu, for example, by expressing the transcription factor Twist1, which limits the immunopathology caused by Th1 cells. Here, we show that in repeatedly activated murine Th1 cells, Twist1 and T-bet induce expression of microRNA-148a (miR-148a). miR-148a regulates expression of the proapoptotic gene Bim, resulting in a decreased Bim/Bcl2 ratio. Inhibition of miR-148a by antagomirs in repeatedly activated Th1 cells increases the expression of Bim, leading to enhanced apoptosis. Knockdown of Bim expression by siRNA in miR-148a antagomir-treated cells restores viability of the Th1 cells, demonstrating that miR-148a controls survival by regulating Bim expression. Thus, Twist1 and T-bet not only control the differentiation and function of Th1 cells, but also their persistence in chronic inflammation. PMID:25486906

  12. Hyperphosphatemia-induced nanocrystals upregulate the expression of bone morphogenetic protein-2 and osteopontin genes in mouse smooth muscle cells in vitro

    PubMed Central

    Sage, Andrew P.; Lu, Jinxiu; Tintut, Yin; Demer, Linda L.

    2011-01-01

    Vascular calcification, which contributes to cardiovascular disease in patients with uremic hyperphosphatemia, is associated with vascular cell expression of osteogenic genes, including bone morphogenetic protein (BMP)-2 and osteopontin (OPN). High inorganic phosphate levels in vitro stimulate the osteogenic conversion of smooth muscle cells; however, the mechanism governing this is not clear. We found that high-phosphate medium increased the expression of BMP-2 and OPN in mouse smooth muscle cells in culture. However, this effect was lost in the presence of the mineralization inhibitor, pyrophosphate, suggesting a contribution of calcium phosphate crystals. Addition of 1–2 mmol/l phosphate alone to growth medium was sufficient to induce nanosized crystals after 1 day at 37 °C. Isolated crystals were about 160 nm in diameter and had a calcium to phosphate ratio of 1.35, consistent with the hydroxyapatite precursor octacalcium phosphate. Nanocrystal formation increased fourfold in the absence of serum, was blocked by fetuin-A, and was dependent on time and on the concentrations of phosphate and calcium. Purified synthetic hydroxyapatite nanocrystals and isolated high-phosphate-induced nanocrystals, but not nanocrystal-free high-phosphate medium, also induced BMP-2 and OPN. Thus, our results suggest that BMP-2 and OPN are induced by calcium phosphate nanocrystals, rather than soluble phosphate. This mechanism may contribute, in part, to hyperphosphatemia-related vascular cell differentiation and calcification. PMID:20944546

  13. Up-regulation of genes involved in N-acetylglucosamine uptake and metabolism suggests a recycling mode of chitin in intraradical mycelium of arbuscular mycorrhizal fungi.

    PubMed

    Kobae, Yoshihiro; Kawachi, Miki; Saito, Katsuharu; Kikuchi, Yusuke; Ezawa, Tatsuhiro; Maeshima, Masayoshi; Hata, Shingo; Fujiwara, Toru

    2015-07-01

    Arbuscular mycorrhizal (AM) fungi colonize roots and form two kinds of mycelium, intraradical mycelium (IRM) and extraradical mycelium (ERM). Arbuscules are characteristic IRM structures that highly branch within host cells in order to mediate resource exchange between the symbionts. They are ephemeral structures and at the end of their life span, arbuscular branches collapse from the tip, fungal cytoplasm withdraws, and the whole arbuscule shrinks into fungal clumps. The exoskeleton of an arbuscule contains structured chitin, which is a polymer of N-acetylglucosamine (GlcNAc), whereas a collapsed arbuscule does not. The molecular mechanisms underlying the turnover of chitin in AM fungi remain unknown. Here, a GlcNAc transporter, RiNGT, was identified from the AM fungus Rhizophagus irregularis. Yeast mutants defective in endogenous GlcNAc uptake and expressing RiNGT took up (14)C-GlcNAc, and the optimum uptake was at acidic pH values (pH 4.0-4.5). The transcript levels of RiNGT in IRM in mycorrhizal Lotus japonicus roots were over 1000 times higher than those in ERM. GlcNAc-6-phosphate deacetylase (DAC1) and glucosamine-6-phosphate isomerase (NAG1) genes, which are related to the GlcNAc catabolism pathway, were also induced in IRM. Altogether, data suggest the existence of an enhanced recycling mode of GlcNAc in IRM of AM fungi. PMID:25564438

  14. Up-regulation of the embryonic self-renewal network through reversible polyploidy in irradiated p53-mutant tumour cells

    SciTech Connect

    Salmina, Kristine, E-mail: kristine@biomed.lu.lv [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia)] [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia); Jankevics, Eriks, E-mail: eriks@biomed.lu.lv [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia)] [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia); Huna, Anda, E-mail: anima-l@inbox.lv [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia)] [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia); Perminov, Dmitry, E-mail: perminov@biomed.lu.lv [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia)] [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia); Radovica, Ilze, E-mail: ilze@biomed.lu.lv [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia)] [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia); Klymenko, Tetyana, E-mail: TKlymenko@picr.man.ac.uk [Paterson Institute of Cancer Research, Manchester University, M20 4BX (United Kingdom)] [Paterson Institute of Cancer Research, Manchester University, M20 4BX (United Kingdom); Ivanov, Andrey, E-mail: a.ivanov@beatson.gla.ac.uk [Beatson Institute, Glasgow Centre for Cancer Research, Glasgow University, G61 4LG (United Kingdom)] [Beatson Institute, Glasgow Centre for Cancer Research, Glasgow University, G61 4LG (United Kingdom); Jascenko, Elina, E-mail: ejascenko@gmail.com [Latvian Institute of Organic Synthesis, Riga, LV-1006 (Latvia)] [Latvian Institute of Organic Synthesis, Riga, LV-1006 (Latvia); Scherthan, Harry, E-mail: harryscherthan@bundeswehr.org [Inst. fuer Radiobiologie der Bundeswehr in Verbindung mit der Univ. Ulm, Munich (Germany)] [Inst. fuer Radiobiologie der Bundeswehr in Verbindung mit der Univ. Ulm, Munich (Germany); Cragg, Mark, E-mail: m.s.cragg@soton.ac.uk [Cancer Sciences Division, Southampton University School of Medicine, General Hospital, Southampton SO16 6YD (United Kingdom)] [Cancer Sciences Division, Southampton University School of Medicine, General Hospital, Southampton SO16 6YD (United Kingdom); Erenpreisa, Jekaterina, E-mail: katrina@biomed.lu.lv [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia)] [Latvian Biomedical Research and Study Centre, Riga, LV-1067 (Latvia)

    2010-08-01

    We have previously documented that transient polyploidy is a potential cell survival strategy underlying the clonogenic re-growth of tumour cells after genotoxic treatment. In an attempt to better define this mechanism, we recently documented the key role of meiotic genes in regulating the DNA repair and return of the endopolyploid tumour cells (ETC) to diploidy through reduction divisions after irradiation. Here, we studied the role of the pluripotency and self-renewal stem cell genes NANOG, OCT4 and SOX2 in this polyploidy-dependent survival mechanism. In irradiation-resistant p53-mutated lymphoma cell-lines (Namalwa and WI-L2-NS) but not sensitive p53 wild-type counterparts (TK6), low background expression of OCT4 and NANOG was up-regulated by ionising radiation with protein accumulation evident in ETC as detected by OCT4/DNA flow cytometry and immunofluorescence (IF). IF analysis also showed that the ETC generate PML bodies that appear to concentrate OCT4, NANOG and SOX2 proteins, which extend into complex nuclear networks. These polyploid tumour cells resist apoptosis, overcome cellular senescence and undergo bi- and multi-polar divisions transmitting the up-regulated OCT4, NANOG and SOX2 self-renewal cassette to their descendents. Altogether, our observations indicate that irradiation-induced ETC up-regulate key components of germ-line cells, which potentially facilitate survival and propagation of the tumour cell population.

  15. Upregulation of TRIM5? gene expression after live-attenuated simian immunodeficiency virus vaccination in Mauritian cynomolgus macaques, but TRIM5? genotype has no impact on virus acquisition or vaccination outcome.

    PubMed

    Mattiuzzo, Giada; Rose, Nicola J; Almond, Neil; Towers, Greg J; Berry, Neil

    2013-03-01

    Polymorphism in the TRIM5?/TRIMcyp gene, which interacts with the lentiviral capsid, has been shown to impact on simian immunodeficiency virus (SIV) replication in certain macaque species. Here, in the context of a live-attenuated SIV vaccine study conducted in Mauritian-origin cynomolgus macaques (MCM), we demonstrate upregulation of TRIM5? expression in multiple lymphoid tissues immediately following vaccination. Despite this, the restricted range of TRIM5? genotypes and lack of TRIMcyp variants had no or only limited impact on the replication kinetics in vivo of either the SIVmac viral vaccine or wild-type SIVsmE660 challenge. Additionally, there appeared to be no impact of TRIM5? genotype on the outcome of homologous or heterologous vaccination/challenge studies. The limited spectrum of TRIM5? polymorphism in MCM appears to minimize host bias to provide consistency of replication for SIVmac/SIVsm viruses in vivo, and therefore on vaccination and pathogenesis studies conducted in this species. PMID:23152371

  16. Brain-specific noncoding RNAs are likely to originate in repeats and may play a role in up-regulating genes in cis.

    PubMed

    Francescatto, Margherita; Vitezic, Morana; Heutink, Peter; Saxena, Alka

    2014-09-01

    The mouse and human brain express a large number of noncoding RNAs (ncRNAs). Some of these are known to participate in neural progenitor cell fate determination, cell differentiation, neuronal and synaptic plasticity and transposable elements derived ncRNAs contribute to somatic variation. Dysregulation of specific long ncRNAs (lncRNAs) has been shown in neuro-developmental and neuro-degenerative diseases thus highlighting the importance of lncRNAs in brain function. Even though it is known that lncRNAs are expressed in cells at low levels in a tissue-specific manner, bioinformatics analyses of brain-specific ncRNAs has not been performed. We analyzed previously published custom microarray ncRNA expression data generated from twelve human tissues to identify tissue-specific ncRNAs. We find that among the 12 tissues studied, brain has the largest number of ncRNAs. Our analyses show that genes in the vicinity of brain-specific ncRNAs are significantly up regulated in the brain. Investigations of repeat representation show that brain-specific ncRNAs are significantly more likely to originate in repeat regions especially DNA/TcMar-Tigger compared with non-tissue-specific ncRNAs. We find SINE/Alus depleted from brain-specific dataset when compared with non-tissue-specific ncRNAs. Our data provide a bioinformatics comparison between brain-specific and non tissue-specific ncRNAs. This article is part of a Directed Issue entitled: The Non-coding RNA Revolution. PMID:24993078

  17. Calcium Upregulation by Percutaneous Administration of Gene Therapy in Cardiac Disease (CUPID Trial), a First-in-Human Phase 1/2 Clinical Trial

    PubMed Central

    JASKI, BRIAN E.; JESSUP, MARIELL L.; MANCINI, DONNA M.; CAPPOLA, THOMAS P.; PAULY, DANIEL F.; GREENBERG, BARRY; BOROW, KENNETH; DITTRICH, HOWARD; ZSEBO, KRISZTINA M.; HAJJAR, ROGER J.

    2009-01-01

    Background SERCA2a deficiency is commonly seen in advanced heart failure (HF). This study is designed to investigate safety and biological effects of enzyme replacement using gene transfer in patients with advanced HF. Methods and Results A total of 9 patients with advanced HF (New York Heart Association [NYHA] Class III/IV, ejection fraction [EF] ?30%, maximal oxygen uptake [VO2 max] <16 mL·kg·min, with maximal pharmacological and device therapy) received a single intracoronary infusion of AAV1/SER-CA2a in the open-label portion of this ongoing study. Doses administered ranged from 1.4 × 1011 to 3 × 1012 DNase resistant particles per patient. We present 6- to 12-month follow-up data for these patients. AAV1/SERCA2a demonstrated an acceptable safety profile in this advanced HF population. Of the 9 patients treated, several demonstrated improvements from baseline to month 6 across a number of parameters important in HF, including symptomatic (NYHA and Minnesota Living with Heart Failure Questionnaire, 5 patients), functional (6-minute walk test and VO2 max, 4 patients), biomarker (NT-ProBNP, 2 patients), and LV function/remodeling (EF and end-systolic volume, 5 patients). Of note, 2 patients who failed to improve had preexisting anti-AAV1 neutralizing antibodies. Conclusions Quantitative evidence of biological activity across a number of parameters important for assessing HF status could be detected in several patients without preexisting neutralizing antibodies in this open-label study, although the number of patients in each cohort is too small to conduct statistical analyses. These findings support the initiation of the Phase 2 double-blind, placebo-controlled portion of this study. PMID:19327618

  18. Up-regulation of calcitonin gene-related peptide in trigeminal ganglion following chronic exposure to paracetamol in a CSD migraine animal model.

    PubMed

    Yisarakun, Waranurin; Chantong, Chattraporn; Supornsilpchai, Weera; Thongtan, Thananya; Srikiatkhachorn, Anan; Reuangwechvorachai, Preecha; Maneesri-le Grand, Supang

    2015-06-01

    Previously, our group has demonstrated that chronic paracetamol (APAP) treatment induces alterations to the trigeminovascular nociceptive system in the cortical spreading depression (CSD) migraine animal model. The calcitonin gene related peptide (CGRP) is a key neuropeptide involved in the activation of the trigeminovascular nociceptive system. Therefore, this study examined the expression levels of CGRP in the trigeminal ganglion (TG) after chronic APAP exposure (0, 15, and 30 days) using a CSD model. Rats were divided into control, CSD only, APAP only and APAP treatment with CSD groups. A single injection (i.p.) of APAP (200?mg/kg body weight) was given to the 0-day APAP-treated groups, while the other APAP-treated groups received daily injections for 15 and 30 days. CSD was induced by the topical application of KCl to the parietal cortex. The protein expression of CGRP in the TG was evaluated by immunohistochemistry, and the CGRP mRNA level was investigated by real-time quantitative reverse transcription polymerase chain reaction. The results revealed that the induction of CSD significantly increased the level of CGRP protein but had no effect on CGRP mRNA level. Pretreatment with APAP 1 hour before CSD activation significantly reduced CGRP expression induced by CSD. In contrast, chronic treatment with APAP (15 and 30 days) significantly enhanced CGRP expression in both protein and mRNA levels when compared with the control groups. In combination with CSD, the expression of CGRP further increased in the animal with 30 day treatment. These findings indicate that chronic treatment with APAP induces an increase of CGRP expression in the TG. This alteration may be associated with the increased trigeminovascular nociception observed in our previous studies. PMID:25998753

  19. Abnormal Apoptosis of Trophoblastic Cells Is Related to the Up-Regulation of CYP11A Gene in Placenta of Preeclampsia Patients

    PubMed Central

    Chen, Yan; Liu, Xinghui; Xi, Mingrong

    2013-01-01

    Abnormal placenta trophoblast proliferation and apoptosis is related to the pathogenesis of preeclampsia. Emerging evidence has also indicated that key pregnancy-associated hormones, such as hCG, progesterone, are found in high concentration at the maternal-fetal interface. The purpose of this study was to investigate the expression of CYP11A, a key enzyme in steroid hormone synthesis and metabolism, in normal pregnancy and severe preeclampsia placenta and to explore the underlying mechanism of the relationship between the altered CYP11A expression and onset of preeclampsia. Immunohistochemistry method was used to study the localization of CYP11A-encoded protein P450scc in the placenta; reverse transcription polymerase chain reaction (RT-PCR) and Western blotting were used to examine CYP11A expression at mRNA and protein levels in patients with severe preeclampsia and normal placental tissue. CYP11A overexpression in trophoblastic cells was used to evaluate the effect on viability. TUNEL staining was used to determine whether overexpression of CYP11A could affect trophoblastic cell apoptosis. The results showed that CYP11A was selectively expressed in the cytoplasm of the placental trophoblastic cells. CYP11A expression were significantly increased in severe preeclampsia compared with normal pregnancy in both mRNA and protein levels. Multiple regression analysis indicated that CYP11A gene expression was positively correlated to ALT level and Plt, while negatively correlated to INR. Overexpression of CYP11A reduced trophoblastic cell proliferation and induced HTR8/SVneo cells apoptosis through activation of activated caspase-3 expression. These results suggest that abnormally high expression of CYP11A inhibits trophoblastic proliferation and increases apoptosis and therefore could be involved in the pathogenesis of preeclampsia. PMID:23555723

  20. Single rol Genes from the Agrobacterium rhizogenes T(L)-DNA Alter Some of the Cellular Responses to Auxin in Nicotiana tabacum.

    PubMed

    Maurel, C; Barbier-Brygoo, H; Spena, A; Tempé, J; Guern, J

    1991-09-01

    Two kinds of cellular responses to auxin, the hyperpolarization of protoplasts and the division of protoplast-derived cells, were compared in Nicotiana tabacum plants transformed by different T-DNA fragments of Agrobacterium rhizogenes strain A4. Using transmembrane potential difference measurements to characterize hormonal sensitivity of mesophyll protoplasts, we found a 30-fold increase in sensitivity to auxin in protoplasts transformed by the whole Ri A4 T-DNA. Furthermore, the rol genes of the Ri A4 T(L)-DNA, together or as single genes, were able to increase the sensitivity to auxin by factors up to 10(4). The different effects of the single rol genes on the sensitivity of mesophyll protoplasts to auxin, rolB being the most powerful, were consistent with their respective rhizogenic effects on leaf fragments (A Spena, T Schmülling, C Koncz, J Schell [1987] EMBO J 6: 3891-3899). No difference was seen concerning the effects of auxin on division of cells derived from normal or transformed protoplasts. These results suggest that only some cellular responses to auxin could be selectively altered by rol genes. They also show that rol-transformed tobaccos can be a model system to study auxin action in plants. PMID:16668373

  1. H3K4 demethylation by Jarid1a and Jarid1b contributes to retinoblastoma-mediated gene silencing during cellular senescence.

    PubMed

    Chicas, Agustin; Kapoor, Avnish; Wang, Xiaowo; Aksoy, Ozlem; Evertts, Adam G; Zhang, Michael Q; Garcia, Benjamin A; Bernstein, Emily; Lowe, Scott W

    2012-06-01

    Cellular senescence is a tumor-suppressive program that involves chromatin reorganization and specific changes in gene expression that trigger an irreversible cell-cycle arrest. Here we combine quantitative mass spectrometry, ChIP deep-sequencing, and functional studies to determine the role of histone modifications on chromatin structure and gene-expression alterations associated with senescence in primary human cells. We uncover distinct senescence-associated changes in histone-modification patterns consistent with a repressive chromatin environment and link the establishment of one of these patterns--loss of H3K4 methylation--to the retinoblastoma tumor suppressor and the H3K4 demethylases Jarid1a and Jarid1b. Our results show that Jarid1a/b-mediated H3K4 demethylation contributes to silencing of retinoblastoma target genes in senescent cells, suggesting a mechanism by which retinoblastoma triggers gene silencing. Therefore, we link the Jarid1a and Jarid1b demethylases to a tumor-suppressor network controlling cellular senescence. PMID:22615382

  2. Single rol Genes from the Agrobacterium rhizogenes TL-DNA Alter Some of the Cellular Responses to Auxin in Nicotiana tabacum 1

    PubMed Central

    Maurel, Christophe; Barbier-Brygoo, Hélène; Spena, Angelo; Tempé, Jacques; Guern, Jean

    1991-01-01

    Two kinds of cellular responses to auxin, the hyperpolarization of protoplasts and the division of protoplast-derived cells, were compared in Nicotiana tabacum plants transformed by different T-DNA fragments of Agrobacterium rhizogenes strain A4. Using transmembrane potential difference measurements to characterize hormonal sensitivity of mesophyll protoplasts, we found a 30-fold increase in sensitivity to auxin in protoplasts transformed by the whole Ri A4 T-DNA. Furthermore, the rol genes of the Ri A4 TL-DNA, together or as single genes, were able to increase the sensitivity to auxin by factors up to 104. The different effects of the single rol genes on the sensitivity of mesophyll protoplasts to auxin, rolB being the most powerful, were consistent with their respective rhizogenic effects on leaf fragments (A Spena, T Schmülling, C Koncz, J Schell [1987] EMBO J 6: 3891-3899). No difference was seen concerning the effects of auxin on division of cells derived from normal or transformed protoplasts. These results suggest that only some cellular responses to auxin could be selectively altered by rol genes. They also show that rol-transformed tobaccos can be a model system to study auxin action in plants. PMID:16668373

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

    PubMed

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

    1995-07-01

    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

  4. Utrophin upregulation in Duchenne muscular dystrophy.

    PubMed

    Hirst, R C; McCullagh, K J A; Davies, K E

    2005-12-01

    Duchenne Muscular Dystrophy (DMD) is a devastating, progressive muscle wasting disease for which there is currently no effective treatment. DMD is caused by mutations in the dystrophin gene many of which result in the absence of the large cytoskeletal protein dystrophin at the sarcolemma. Over-expression of utrophin, the autosomal paralogue of dystrophin, as a transgene in the mdx mouse (the mouse model of DMD) has demonstrated that utrophin can prevent the muscle pathology. Thus, up-regulation of utrophin in DMD muscle is a potential therapy for DMD. In this review we discuss recent advances in our understanding of the regulatory pathways controlling utrophin expression and the various approaches that have been applied to increasing the level of utrophin in the mdx mouse. These results are very encouraging and suggest that pharmacological up-regulation of utrophin may well be a feasible approach to therapy for DMD. PMID:16629055

  5. Overexpression of the Gene Encoding GTP:Mannose-1-Phosphate Guanyltransferase, mpg1, Increases Cellular GDP-Mannose Levels and Protein Mannosylation in Trichoderma reesei

    PubMed Central

    Zakrzewska, Anna; Palamarczyk, Grazyna; Krotkiewski, Hubert; Zdebska, Ewa; Saloheimo, Markku; Penttilä, Merja; Kruszewska, Joanna S.

    2003-01-01

    To elucidate the regulation and limiting factors in the glycosylation of secreted proteins, the mpg1 and dpm1 genes from Trichoderma reesei (Hypocrea jecorina) encoding GTP:?-d-mannose-1-phosphate guanyltransferase and dolichyl phosphate mannose synthase (DPMS), respectively, were overexpressed in T. reesei. No significant increases were observed in DPMS activity or protein secretion in dpm1-overexpressing transformants, whereas overexpression of mpg1 led to a twofold increase in GDP-mannose (GDPMan) levels. GDPMan was effectively utilized by mannnosyltransferases and resulted in hypermannosylation of secreted proteins in both N and O glycosylation. Overexpression of the mpg1 gene also increased the transcription of the dpm1 gene and DPMS activity. Our data indicate that the level of cellular GDPMan can play a major regulatory role in protein glycosylation in T. reesei. ? PMID:12902219

  6. Interplay between TAp73 Protein and Selected Activator Protein-1 (AP-1) Family Members Promotes AP-1 Target Gene Activation and Cellular Growth.

    PubMed

    Subramanian, Deepa; Bunjobpol, Wilawan; Sabapathy, Kanaga

    2015-07-24

    Unlike p53, which is mutated at a high rate in human cancers, its homologue p73 is not mutated but is often overexpressed, suggesting a possible context-dependent role in growth promotion. Previously, we have shown that co-expression of TAp73 with the proto-oncogene c-Jun can augment cellular growth and potentiate transactivation of activator protein (AP)-1 target genes such as cyclin D1. Here, we provide further mechanistic insights into the cooperative activity between these two transcription factors. Our data show that TAp73-mediated AP-1 target gene transactivation relies on c-Jun dimerization and requires the canonical AP-1 sites on target gene promoters. Interestingly, only selected members of the Fos family of proteins such as c-Fos and Fra1 were found to cooperate with TAp73 in a c-Jun-dependent manner to transactivate AP-1 target promoters. Inducible expression of TAp73 led to the recruitment of these Fos family members to the AP-1 target promoters on which TAp73 was found to be bound near the AP-1 site. Consistent with the binding of TAp73 and AP-1 members on the target promoters in a c-Jun-dependent manner, TAp73 was observed to physically interact with c-Jun specifically at the chromatin via its carboxyl-terminal region. Furthermore, co-expression of c-Fos or Fra1 was able to cooperate with TAp73 in potentiating cellular growth, similarly to c-Jun. These data together suggest that TAp73 plays a vital role in activation of AP-1 target genes via direct binding to c-Jun at the target promoters, leading to enhanced loading of other AP-1 family members, thereby leading to cellular growth. PMID:26018080

  7. Kallikrein 4 (KLK4), a new member of the human kallikrein gene family is up-regulated by estrogen and progesterone in the human endometrial cancer cell line, KLE.

    PubMed

    Myers, S A; Clements, J A

    2001-05-01

    Endometrial cancer is the fourth most common female malignancy in women in developed countries. Estrogen, and to a lesser degree, progesterone, regulate specific target genes that are involved in endometrial tumorigenesis. A family of proteases involved in cellular proliferation, extracellular matrix degradation and thus, implicated in tumorigenesis, and regulated by estrogen and progesterone in a number of systems, are the tissue kallikreins (KLKs). KLK4, a new member of the KLK gene family, was found to be expressed to varying levels in a number of endometrial cancer cell lines- HEC1A, HEC1B, Ishikawa, RL95-2 and KLE- at both the mRNA and protein level. On the addition of 10 nmol/L estradiol, progesterone, or a combination of both over a 48 h period, an increase in the intracellular protein levels of K4 were observed when compared to the control (untreated) cells. We have also identified a novel KLK4 transcript with a complete exon 4 deletion. The significance of this alternative transcript, which would give rise to a truncated protein without a serine residue (which is essential for catalytic activity), is yet to be established. These cell lines now provide a model system to study the role of KLK4 and the molecular mechanisms of KLK4 regulation by estrogen and progesterone, in endometrial tumorigenesis. PMID:11344246

  8. Mechanisms of Nrf2/Keap1-Dependent Phase II Cytoprotective and Detoxifying Gene Expression and Potential Cellular Targets of Chemopreventive Isothiocyanates

    PubMed Central

    Das, Biswa Nath; Kim, Young-Woo

    2013-01-01

    Isothiocyanates (ITCs) are abundantly found in cruciferous vegetables. Epidemiological studies suggest that chronic consumption of cruciferous vegetables can lower the overall risk of cancer. Natural ITCs are key chemopreventive ingredients of cruciferous vegetables, and one of the prime chemopreventive mechanisms of natural isothiocyanates is the induction of Nrf2/ARE-dependent gene expression that plays a critical role in cellular defense against electrophiles and reactive oxygen species. In the present review, we first discuss the underlying mechanisms how natural ITCs affect the intracellular signaling kinase cascades to regulate the Keap1/Nrf2 activities, thereby inducing phase II cytoprotective and detoxifying enzymes. We also discuss the potential cellular protein targets to which natural ITCs are directly conjugated and how these events aid in the chemopreventive effects of natural ITCs. Finally, we discuss the posttranslational modifications of Keap1 and nucleocytoplasmic trafficking of Nrf2 in response to electrophiles and oxidants. PMID:23781297

  9. Multiple-Integrations of HPV16 Genome and Altered Transcription of Viral Oncogenes and Cellular Genes Are Associated with the Development of Cervical Cancer

    PubMed Central

    Lin, Mao; Duan, Ping; Ye, Lulu; Chen, Jun; Chen, Xiangmin; Zhang, Lifang; Xue, Xiangyang

    2014-01-01

    The constitutive expression of the high-risk HPV E6 and E7 viral oncogenes is the major cause of cervical cancer. To comprehensively explore the composition of HPV16 early transcripts and their genomic annotation, cervical squamous epithelial tissues from 40 HPV16-infected patients were collected for analysis of papillomavirus oncogene transcripts (APOT). We observed different transcription patterns of HPV16 oncogenes in progression of cervical lesions to cervical cancer and identified one novel transcript. Multiple-integration events in the tissues of cervical carcinoma (CxCa) are significantly more often than those of low-grade squamous intraepithelial lesions (LSIL) and high-grade squamous intraepithelial lesions (HSIL). Moreover, most cellular genes within or near these integration sites are cancer-associated genes. Taken together, this study suggests that the multiple-integrations of HPV genome during persistent viral infection, which thereby alters the expression patterns of viral oncogenes and integration-related cellular genes, play a crucial role in progression of cervical lesions to cervix cancer. PMID:24992025

  10. The Aspergillus nidulans sepA gene encodes an FH1/2 protein involved in cytokinesis and the maintenance of cellular polarity.

    PubMed

    Harris, S D; Hamer, L; Sharpless, K E; Hamer, J E

    1997-06-16

    Cytokinesis (septation) in the fungus Aspergillus nidulans occurs through the formation of a transient actin ring at the incipient division site. Temperature-sensitive mutations in the sepA gene prevent septation and cause defects in the maintenance of cellular polarity, without affecting growth and nuclear division. The sepA gene encodes a member of the growing family of FH1/2 proteins, which appear to have roles in morphogenesis and cytokinesis in organisms such as yeast and Drosophila. Results from temperature shift and immunofluorescence microscopy experiments strongly suggest that sepA function requires a preceding mitosis and that sepA acts prior to actin ring formation. Deletion mutants of sepA exhibit temperature-sensitive growth and severe delays in septation at the permissive temperature, indicating that expression of another gene may compensate for the loss of sepA. Conidiophores formed by sepA mutants exhibit abnormal branching of the stalk and vesicle. These results suggest that sepA interacts with the actin cytoskeleton to promote formation of the actin ring during cytokinesis and that sepA is also required for maintenance of cellular polarity during hyphal growth and asexual morphogenesis. PMID:9218790

  11. Genome-wide transcriptional profiling analysis reveals annexin A6 as a novel EZH2 target gene involving gastric cellular proliferation.

    PubMed

    Qi, Ying; Zhang, Xiaoli; Kang, Yani; Wu, Jun; Chen, Jian; Li, Hua; Guo, Yan; Liu, Bingya; Shao, Zhifeng; Zhao, Xiaodong

    2015-06-16

    A histone methyltransferase enhancer of zeste homologue 2 (EZH2) catalyzes trimethylation at histone H3 lysine27 (H3K27me3) and is frequently dysregulated in a wide range of human cancers. EZH2-mediated gene silencing contributes to carcinogenesis and regulates stem cell maintenance and differentiation; however, the underlining mechanisms remain to be completely understood. Here, we found that downregulation of EZH2 by RNA interference (RNAi) in gastric cancer cells suppresses cell growth, migration, invasion, and induces cell cycle arrest. Transcriptome analysis identified 1223 EZH2 responsive genes upon EZH2 knockdown. These genes are involved in the biological processes of cell cycle, proliferation and metastasis. Particularly, we found that annexin A6 (ANXA6) is a new target of EZH2 and is repressed in gastric cancer cells. Restoration of ANXA6 expression inhibits gastric cellular proliferation. We further demonstrated that EZH2-mediated H3K27me3, rather than promoter DNA methylation, is primarily responsible for ANXA6 inhibition. Taken together, our results provide a framework for understanding EZH2 biology and reveal ANXA6 as a new EZH2 target involving gastric cellular proliferation. PMID:25947258

  12. Upregulation of Parkin in Endophilin Mutant Mice

    PubMed Central

    Cao, Mian; Milosevic, Ira; Giovedi, Silvia

    2014-01-01

    Several proteins encoded by PD genes are implicated in synaptic vesicle traffic. Endophilin, a key factor in the endocytosis of synaptic vesicles, was shown to bind to, and be ubiquitinated by, the PD-linked E3 ubiquitin ligase Parkin. Here we report that Parkin's level is specifically upregulated in brain and fibroblasts of endophilin mutant mice due to increased transcriptional regulation. Studies of transfected HEK293T cells show that Parkin ubiquitinates not only endophilin, but also its major binding partners, dynamin and synaptojanin 1. These results converge with the recently reported functional relationship of endophilin to the PD gene LRRK2 and with the identification of a PD-linked synaptojanin 1 mutation, in providing evidence for a link between PD and endocytosis genes. PMID:25471590

  13. Bacterial diversity among small-subunit rRNA gene clones and cellular isolates from the same seawater sample

    Microsoft Academic Search

    MARCELINO T. SUZUKI; MICHAEL S. RAPPE; ZARA W. HAIMBERGER

    1997-01-01

    Numerous investigations applying the cloning and sequencing of rRNA genes (rDNAs) to the study of marine bacterioplankton diversity have shown that the sequences of genes cloned directly from environmental DNA do not correspond to the genes of cultured marine taxa. These results have been interpreted as support for the hypothesis that the most abundant heterotrophic marine bacterioplankton species are not

  14. Abnormal gene expression and gene fusion in lung adenocarcinoma with high-throughput RNA sequencing.

    PubMed

    Yang, Z-H; Zheng, R; Gao, Y; Zhang, Q; Zhang, H

    2014-02-01

    To explore the universal law of the abnormal gene expression and the structural variation of genes related to lung adenocarcinoma, the gene expression profile of GSE37765 were downloaded from Gene Expression Omnibus database. The differentially expressed genes (DEGs) were analyzed with t-test and NOISeq tool, and the core DEGs were screened out by combining with another RNA-seq data containing totally 77 pairs of samples in 77 patients with lung adenocarcinoma. Moreover, the functional annotation of the core DEGs was performed by using the Database for Annotation Visualization and Integrated Discovery following selection of oncogene and tumor suppressor by combining with tumor suppressor genes and Cancer Genes database, and motif-finding of core DEGs was performed with motif-finding algorithm Seqpos. We also used Tophat-fusion tool to further explore the fusion genes. In total, 850 downregulated DEGs and 206 upregulated DEGs were screened out in lung adenocarcinoma tissues. Next, we selected 543 core DEGs, including 401 downregulated and 142 upregulated genes, and vasculature development (P=1.89E-06) was significantly enriched among downregulated core genes, as well as mitosis (P=6.26E-04) enriched among upregulated core genes. On the basis of the cellular localization analysis of core genes, wnt-1-induced secreted protein 1 (WISP1) and receptor (G protein-coupled) activity modifying protein 1 (RAMP1) identified mainly located in extracellular region and extracellular space. We also screened one oncogene, v-myb avian myeloblastosis viral oncogene homolog-like 2 (MYBL2). Moreover, transcription factor GATA2 was mined by motif-finding analysis. Finally, four fusion genes belonged to the human leukocyte antigen (HLA) family. WISP1, RAMP1, MYBL2 and GATA2 could be potential targets of treatment for lung adenocarcinoma and the fusion of HLA family genes might have important roles in lung adenocarcinoma. PMID:24503571

  15. Up-regulation of nuclear factor E2-related factor 2 upon SVCV infection.

    PubMed

    Yang, Yi; Huang, Jian; Li, Lijuan; Lin, Li; Zhai, Yanhua; Chen, Xiaoxuan; Liu, Xueqin; Wu, Zhixin; Yuan, Junfa

    2014-09-01

    Nuclear factor E2 - related factor 2 (Nrf2) is a crucial transcription factor that regulates the basal and inducible expression of many antioxidant response element (ARE)-dependent genes, including heme oxygenase-1 (HO-1) and superoxide dismutase 1 (SOD1). The Nrf2/ARE pathway has been regarded as a critical switch in the initiation of cellular defence systems for surviving oxidative insults and viral infection. In this study, the Nrf2 gene of EPC cells, which is originally derived from Pimephales promelas, was cloned, and an investigation on the interactions between Nrf2 and spring viraemia of carp virus (SVCV) was performed. These results demonstrated that the virus facilitated the nuclear accumulation of Nrf2 and up-regulated its transcriptional and protein profiles in EPC cells. In addition, exogenous activation of Nrf2 conferred EPC cells with a higher cellular total antioxidant capacity via an increase in the expression of HO-1 and SOD1, but did not suppress the replication of SVCV. PMID:25038284

  16. Gain-of-function mutations in IFIH1 cause a spectrum of human disease phenotypes associated with upregulated type I interferon signaling

    PubMed Central

    Jenkinson, Emma M; Forte, Gabriella MA; Anderson, Beverley H; Ariaudo, Giada; Bader-Meunier, Brigitte; Baildam, Eileen M; Battini, Roberta; Beresford, Michael W; Casarano, Manuela; Chouchane, Mondher; Cimaz, Rolando; Collins, Abigail E; Cordeiro, Nuno JV; Dale, Russell C; Davidson, Joyce E; De Waele, Liesbeth; Desguerre, Isabelle; Faivre, Laurence; Fazzi, Elisa; Isidor, Bertrand; Lagae, Lieven; Latchman, Andrew R; Lebon, Pierre; Li, Chumei; Livingston, John H; Lourenço, Charles M; Mancardi, Maria Margherita; Masurel-Paulet, Alice; McInnes, Iain B; Menezes, Manoj P; Mignot, Cyril; O’Sullivan, James; Orcesi, Simona; Picco, Paolo P; Riva, Enrica; Robinson, Robert A; Rodriguez, Diana; Salvatici, Elisabetta; Scott, Christiaan; Szybowska, Marta; Tolmie, John L; Vanderver, Adeline; Vanhulle, Catherine; Vieira, Jose Pedro; Webb, Kate; Whitney, Robyn N; Williams, Simon G; Wolfe, Lynne A; Zuberi, Sameer M; Hur, Sun; Crow, Yanick J

    2014-01-01

    The type I interferon system is integral to human antiviral immunity. However, inappropriate stimulation or defective negative regulation of this system can lead to inflammatory disease. We sought to determine the molecular basis of genetically uncharacterized cases of the type I interferonopathy Aicardi-Goutières syndrome, and of other patients with undefined neurological and immunological phenotypes also demonstrating an upregulated type I interferon response. We found that heterozygous mutations in the cytosolic double-stranded RNA receptor gene IFIH1 (MDA5) cause a spectrum of neuro-immunological features consistently associated with an enhanced interferon state. Cellular and biochemical assays indicate that these mutations confer a gain-of-function - so that mutant IFIH1 binds RNA more avidly, leading to increased baseline and ligand-induced interferon signaling. Our results demonstrate that aberrant sensing of nucleic acids can cause immune upregulation. PMID:24686847

  17. Expression of Cellular myc and mos Genes in Undifferentiated B Cell Lymphomas of Burkitt and Non-Burkitt Types

    Microsoft Academic Search

    Robert T. Maguire; Terry S. Robins; Snorri S. Thorgeirsson; Carole A. Heilman

    1983-01-01

    Burkitt lymphomas contain reciprocal translocations between chromosome 8 and one of the chromosomes containing the immunoglobulin gene loci, prompting speculation that consequent activation of a crucial gene(s) on chromosome 8 might be involved in the generation of these tumors. Recently the human counterparts of the retroviral oncogenes v-myc and v-mos have been mapped to chromosome 8. We have, therefore, analyzed

  18. Cellular repressor of E1A-stimulated genes is a bona fide lysosomal protein which undergoes proteolytic maturation during its biosynthesis

    SciTech Connect

    Schaehs, Philipp; Weidinger, Petra; Probst, Olivia C.; Svoboda, Barbara [Institut fuer Angewandte Genetik und Zellbiologie, Universitaet fuer Bodenkultur Wien, Muthgasse 18, A-1190 Vienna (Austria); Stadlmann, Johannes [Department fuer Chemie, Universitaet fuer Bodenkultur Wien, Muthgasse 18, A-1190 Vienna (Austria); Beug, Hartmut; Waerner, Thomas [Research Institute of Molecular Pathology, Dr. Bohr-Gasse 7, A-1030 Vienna (Austria); Mach, Lukas [Institut fuer Angewandte Genetik und Zellbiologie, Universitaet fuer Bodenkultur Wien, Muthgasse 18, A-1190 Vienna (Austria)], E-mail: lukas.mach@boku.ac.at

    2008-10-01

    Cellular repressor of E1A-stimulated genes (CREG) has been reported to be a secretory glycoprotein implicated in cellular growth and differentiation. We now show that CREG is predominantly localized within intracellular compartments. Intracellular CREG was found to lack an N-terminal peptide present in the secreted form of the protein. In contrast to normal cells, CREG is largely secreted by fibroblasts missing both mannose 6-phosphate receptors. This is not observed in cells lacking only one of them. Mass spectrometric analysis of recombinant CREG revealed that the protein contains phosphorylated oligosaccharides at either of its two N-glycosylation sites. Cellular CREG was found to cosediment with lysosomal markers upon subcellular fractionation by density-gradient centrifugation. In fibroblasts expressing a CREG-GFP fusion construct, the heterologous protein was detected in compartments containing lysosomal proteins. Immunolocalization of endogenous CREG confirmed that intracellular CREG is localized in lysosomes. Proteolytic processing of intracellular CREG involves the action of lysosomal cysteine proteinases. These results establish that CREG is a lysosomal protein that undergoes proteolytic maturation in the course of its biosynthesis, carries the mannose 6-phosphate recognition marker and depends on the interaction with mannose 6-phosphate receptors for efficient delivery to lysosomes.

  19. Comparative gene expression profiling of adult mouse ovary-derived oogonial stem cells supports a distinct cellular identity

    PubMed Central

    Imudia, Anthony N.; Wang, Ning; Tanaka, Yoshihiro; White, Yvonne A.R.; Woods, Dori C.; Tilly, Jonathan L.

    2013-01-01

    Objective Perform gene expression profiling of adult mouse ovary-derived oogonial stem cells (OSCs). Design Experimental animal study. Setting Research laboratory. Animal(s) Adult C57BL/6 female mice. Intervention(s) None. Main outcome measure(s) Gene expression profiles were compared between freshly isolated and cultured OSCs, as well as between OSCs and embryonic stem cells (ESCs), fetal primordial germ cells (PGCs) and spermatogonial stem cells (SSCs); OSC yield from ovaries versus meiotic gene activation during the estrous cycle was determined. Result(s) Freshly isolated OSCs, PGCs and SSCs exhibited distinct gene expression profiles. Cultured OSCs maintained their germline gene expression pattern, but gained expression of pluripotency markers found in PGCs and ESCs. Cultured OSCs also expressed the meiotic marker, stimulated by retinoic acid gene 8 (Stra8). In vivo, OSC yield was higher from luteal versus follicular phase ovaries and this was inversely related to Stra8 expression. Conclusion(s) Freshly isolated OSCs exhibit a germline gene expression profile that overlaps with, but is distinct from, that of PGCs and SSCs. After in vitro expansion, OSCs activate expression of pluripotency genes found in freshly isolated PGCs. In vivo, OSC numbers in the ovaries fluctuate during the estrous cycle, with the highest numbers noted during the luteal phase. This is followed by activation of Stra8 expression during the follicular phase, which may signify a wave of neo-oogenesis to partially offset follicular loss through atresia and ovulation in the prior cycle. PMID:23876535

  20. Molecular and Cellular Endocrinology 283 (2008) 3848 Auto-regulation of estrogen receptor subtypes and gene expression profiling

    E-print Network

    Xia, Xuhua

    2008-01-01

    induction of liver VTG was observed. In the testes (7 d) and telencephalon (7 d), E2 induced ER conditions. Although aromatase B levels increased in the brain, the majority of candidate genes identified. © 2007 Elsevier Ireland Ltd. All rights reserved. Keywords: Estrogen receptors; Fish; Aromatase; Gene

  1. RT-qPCR reveals opsin gene upregulation associated with age and sex in guppies ( Poecilia reticulata ) - a species with color-based sexual selection and 11 visual-opsin genes

    Microsoft Academic Search

    Christopher RJ Laver; John S Taylor

    2011-01-01

    Background  PCR-based surveys have shown that guppies (Poecilia reticulata) have an unusually large visual-opsin gene repertoire. This has led to speculation that opsin duplication and divergence\\u000a has enhanced the evolution of elaborate male coloration because it improves spectral sensitivity and\\/or discrimination in\\u000a females. However, this conjecture on evolutionary connections between opsin repertoire, vision, mate choice, and male coloration\\u000a was generated with

  2. 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

  3. Sex as a response to oxidative stress: a twofold increase in cellular reactive oxygen species activates sex genes

    Microsoft Academic Search

    Aurora M. Nedelcu; Oana Marcu; Richard E. Michod

    2004-01-01

    Organisms are constantly subjected to factors that can alter the cellular redox balance and result in the formation of a series of highly reactive molecules known as reactive oxygen species (ROS). As ROS can be damaging to biological structures, cells evolved a series of mechanisms (e.g. cell-cycle arrest, pro- grammed cell death) to respond to high levels of ROS (i.e.

  4. Aryl hydrocarbon receptor activation in hematopoietic stem/progenitor cells alters cell function and pathway-specific gene modulation reflecting changes in cellular trafficking and migration.

    PubMed

    Casado, Fanny L; Singh, Kameshwar P; Gasiewicz, Thomas A

    2011-10-01

    The aryl hydrocarbon receptor (AhR) is a transcription factor belonging to the Per-ARNT-Sim family of proteins. These proteins sense molecules and stimuli from the cellular/tissue environment and initiate signaling cascades to elicit appropriate cellular responses. Recent literature reports suggest an important function of AhR in hematopoietic stem cell (HSC) biology. However, the molecular mechanisms by which AhR signaling regulates HSC functions are unknown. In previous studies, we and others reported that treatment of mice with the AhR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) compromises the competitive reconstitution of bone marrow (BM) cells into irradiated host animals. Additional studies indicated a requirement for AhR in hematopoietic cells and not marrow microenvironment cells. In this study, we tested the hypothesis that TCDD-mediated phenotypic and functional changes of HSCs are a result of changes in gene expression that disrupt stem cell numbers and/or their migration. TCDD treatment to mice increased the numbers of phenotypically defined HSCs in BM. These cells showed compromised migration to the BM in vivo and to the chemokine CXCL12 in vitro, as well as increased expression of the leukemia-associated receptors CD184 (CXCR4) and CD44. Gene expression profiles at 6 and 12 h after exposure were consistent with the phenotypic and functional changes observed. The expressions of Scin, Nqo1, Flnb, Mmp8, Ilf9, and Slamf7 were consistently altered. TCDD also disrupted expression of other genes involved in hematological system development and function including Fos, JunB, Egr1, Ptgs2 (Cox2), and Cxcl2. These data support a molecular mechanism for an AhR ligand to disrupt the homeostatic cell signaling of HSCs that may promote altered HSC function. PMID:21791576

  5. A Novel Role for Keratin 17 in Coordinating Oncogenic Transformation and Cellular Adhesion in Ewing Sarcoma

    PubMed Central

    Sankar, Savita; Tanner, Jason M.; Bell, Russell; Chaturvedi, Aashi; Randall, R. Lor; Beckerle, Mary C.

    2013-01-01

    Oncogenic transformation in Ewing sarcoma is caused by EWS/FLI, an aberrant transcription factor fusion oncogene. Glioma-associated oncogene homolog 1 (GLI1) is a critical target gene activated by EWS/FLI, but the mechanism by which GLI1 contributes to the transformed phenotype of Ewing sarcoma was unknown. In this work, we identify keratin 17 (KRT17) as a direct downstream target gene upregulated by GLI1. We demonstrate that KRT17 regulates cellular adhesion by activating AKT/PKB (protein kinase B) signaling. In addition, KRT17 is necessary for oncogenic transformation in Ewing sarcoma and accounts for much of the GLI1-mediated transformation function but via a mechanism independent of AKT signaling. Taken together, our data reveal previously unknown molecular functions for a cytoplasmic intermediate filament protein, KRT17, in coordinating EWS/FLI- and GLI1-mediated oncogenic transformation and cellular adhesion in Ewing sarcoma. PMID:24043308

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

    E-print Network

    Luengo-Oroz, Miguel A.

    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, ...

  7. Eicosapentaenoic acid increases lipolysis through up-regulation of the lipolytic gene expression and down-regulation of the adipogenic gene expression in 3T3-L1 adipocytes

    PubMed Central

    Lee, Mak-Soon; Kwun, In-Sook

    2007-01-01

    In this study, we investigated the lipolytic effects of eicosapentaenoic acid (EPA) in 3T3-L1 adipocytes. The differentiated 3T3-L1 adipocytes were treated in a serum-free medium with 300 ?M of EPA for 3, 6, 12, and 24 h. In comparison with the control, intracellular lipid accumulation was significantly decreased by 24% at 24 h following the addition of EPA (P < 0.05). Under the same experimental conditions, there was an increase of glycerol and free fatty acids (FFAs). The mRNA level of carnitine palmitoyltransferase I-a, a component of the fatty-acid shuttle system involved in the mitochondrial oxidation of long-chain fatty acids, was also significantly elevated by EPA (P < 0.05). However, the expression of peroxisome proliferator-activated receptor-? and acetyl-CoA carboxylase (ACC), which are involved in adipogenesis, was significantly down-regulated by EPA (P < 0.05). These results suggest that EPA may modulate lipid metabolism by stimulation of lipolysis, which was associated with induction of lipolytic gene expression and suppression of adipogenic gene expression in 3T3-L1 adipocytes. PMID:18850226

  8. Homologous DNA sequences and cellular factors are implicated in the control of glucagon and insulin gene expression.

    PubMed Central

    Cordier-Bussat, M; Morel, C; Philippe, J

    1995-01-01

    The glucagon gene is specifically expressed in the alpha cells of pancreatic islets. The promoter of the glucagon gene is responsible for this specificity. Within the promoter, the upstream promoter element G1 is critical to restrict expression to the alpha cells. We define here a composite DNA control element, G4, localized upstream of G1 between nucleotides -100 and -140 which functions as an islet-specific activator in both glucagon- and insulin-producing cells but not in nonislet cells. G4 contains at least three protein binding sites. The most proximal site, E2, is highly homologous to the E1, SMS-UE, and B elements of the rat insulin I, somastatin, and elastase I genes, respectively, and interacts with a pancreas-specific complex; the distal site, E3, represents an E box which is identical to the E boxes of the rat insulin I and II genes and binds to a complex similar or identical to IEF1 which has been implicated in the tissue-specific control of insulin gene expression. These two sites necessitate a third element, the intervening sequence, to activate transcription. We conclude that the first 140 bp of the glucagon gene promoter contains at least two DNA control elements responsible for pancreatic alpha-cell-specific expression: G4, an islet cell-specific element sharing common binding sites with the insulin gene, and G1, which restricts glucagon gene expression to the alpha cells. This double control of specificity might have relevance during islet cell differentiation. PMID:7791796

  9. Induction of Interferon-Stimulated Genes on the IL-4 Response Axis by Epstein-Barr Virus Infected Human B Cells; Relevance to Cellular Transformation

    PubMed Central

    Wei, Wenbin; Vockerodt, Martina; Murray, Paul G.; Woodman, Ciaran B.; Rowe, Martin

    2013-01-01

    Epstein-Barr virus (EBV) is an oncogenic virus that is associated with the pathogenesis of several human lymphoid malignancies, including Hodgkin's lymphoma. Infection of normal resting B cells with EBV results in activation to lymphoblasts that are phenotypically similar to those generated by physiological stimulation with CD40L plus IL-4. One important difference is that infection leads to the establishment of permanently growing lymphoblastoid cell lines, whereas CD40L/IL-4 blasts have finite proliferation lifespans. To identify early events which might later determine why EBV infected blasts go on to establish transformed cell lines, we performed global transcriptome analyses on resting B cells and on EBV and CD40L/IL-4 blasts after 7 days culture. As anticipated there was considerable overlap in the transcriptomes of the two types of lymphoblasts when compared to the original resting B cells, reflecting common changes associated with lymphocyte activation and proliferation. Of interest to us was a subset of 255 genes that were differentially expressed between EBV and CD40L/IL-4 blasts. Genes which were more highly expressed in EBV blasts were substantially and significantly enriched for a set of interferon-stimulated genes which on further in silico analyses were found to be repressed by IL-4 in other cell contexts and to be up-regulated in micro-dissected malignant cells from Hodgkin's lymphoma biopsies when compared to their normal germinal center cell counterparts. We hypothesized that EBV and IL-4 were targeting and discordantly regulating a common set of genes. This was supported experimentally in our B cell model where IL-4 stimulation partially reversed transcriptional changes which follow EBV infection and it impaired the efficiency of EBV-induced B cell transformation. Taken together, these data suggest that the discordant regulation of interferon and IL-4 pathway genes by EBV that occurs early following infection of B cells has relevance to the development or maintenance of an EBV-associated malignancy. PMID:23724103

  10. Cellular homeoproteins, SATB1 and CDP, bind to the unique region between the human cytomegalovirus UL127 and major immediate-early genes

    SciTech Connect

    Lee Jialing [Expression Engineering Group, Bioprocessing Technology Institute, 20 Biopolis Way, 06-01 Centros, Singapore 138668 (Singapore); Klase, Zachary [Department of Biochemistry and Molecular Biology, George Washington University School of Medicine, Washington, DC 20037 (United States); Gao Xiaoqi; Caldwell, Jeremy S. [Genomics Institute of the Novartis Research Foundation, 10675 John Jay Hopkins Drive, San Diego, CA 92121 (United States); Stinski, Mark F. [Department of Microbiology, Carver College of Medicine, University of Iowa, Iowa City, IA 52242 (United States); Kashanchi, Fatah [Department of Biochemistry and Molecular Biology, George Washington University School of Medicine, Washington, DC 20037 (United States); Chao, S.-H. [Expression Engineering Group, Bioprocessing Technology Institute, 20 Biopolis Way, 06-01 Centros, Singapore 138668 (Singapore)], E-mail: jimmy_chao@bti.a-star.edu.sg

    2007-09-15

    An AT-rich region of the human cytomegalovirus (CMV) genome between the UL127 open reading frame and the major immediate-early (MIE) enhancer is referred to as the unique region (UR). It has been shown that the UR represses activation of transcription from the UL127 promoter and functions as a boundary between the divergent UL127 and MIE genes during human CMV infection [Angulo, A., Kerry, D., Huang, H., Borst, E.M., Razinsky, A., Wu, J., Hobom, U., Messerle, M., Ghazal, P., 2000. Identification of a boundary domain adjacent to the potent human cytomegalovirus enhancer that represses transcription of the divergent UL127 promoter. J. Virol. 74 (6), 2826-2839; Lundquist, C.A., Meier, J.L., Stinski, M.F., 1999. A strong negative transcriptional regulatory region between the human cytomegalovirus UL127 gene and the major immediate-early enhancer. J. Virol. 73 (11), 9039-9052]. A putative forkhead box-like (FOX-like) site, AAATCAATATT, was identified in the UR and found to play a key role in repression of the UL127 promoter in recombinant virus-infected cells [Lashmit, P.E., Lundquist, C.A., Meier, J.L., Stinski, M.F., 2004. Cellular repressor inhibits human cytomegalovirus transcription from the UL127 promoter. J. Virol. 78 (10), 5113-5123]. However, the cellular factors which associate with the UR and FOX-like region remain to be determined. We reported previously that pancreatic-duodenal homeobox factor-1 (PDX1) bound to a 45-bp element located within the UR [Chao, S.H., Harada, J.N., Hyndman, F., Gao, X., Nelson, C.G., Chanda, S.K., Caldwell, J.S., 2004. PDX1, a Cellular Homeoprotein, Binds to and Regulates the Activity of Human Cytomegalovirus Immediate Early Promoter. J. Biol. Chem. 279 (16), 16111-16120]. Here we demonstrate that two additional cellular homeoproteins, special AT-rich sequence binding protein 1 (SATB1) and CCAAT displacement protein (CDP), bind to the human CMV UR in vitro and in vivo. Furthermore, CDP is identified as a FOX-like binding protein and a repressor of the UL127 promoter, while SATB1 has no effect on UL127 expression. Since CDP is known as a transcription repressor and a nuclear matrix-associated region binding protein, CDP may have a role in the regulation of human CMV transcription.

  11. Cellular homeoproteins, SATB1 and CDP, bind to the unique region between the human cytomegalovirus UL127 and major immediate-early genes.

    PubMed

    Lee, Jialing; Klase, Zachary; Gao, Xiaoqi; Caldwell, Jeremy S; Stinski, Mark F; Kashanchi, Fatah; Chao, Sheng-Hao

    2007-09-15

    An AT-rich region of the human cytomegalovirus (CMV) genome between the UL127 open reading frame and the major immediate-early (MIE) enhancer is referred to as the unique region (UR). It has been shown that the UR represses activation of transcription from the UL127 promoter and functions as a boundary between the divergent UL127 and MIE genes during human CMV infection [Angulo, A., Kerry, D., Huang, H., Borst, E.M., Razinsky, A., Wu, J., Hobom, U., Messerle, M., Ghazal, P., 2000. Identification of a boundary domain adjacent to the potent human cytomegalovirus enhancer that represses transcription of the divergent UL127 promoter. J. Virol. 74 (6), 2826-2839; Lundquist, C.A., Meier, J.L., Stinski, M.F., 1999. A strong negative transcriptional regulatory region between the human cytomegalovirus UL127 gene and the major immediate-early enhancer. J. Virol. 73 (11), 9039-9052]. A putative forkhead box-like (FOX-like) site, AAATCAATATT, was identified in the UR and found to play a key role in repression of the UL127 promoter in recombinant virus-infected cells [Lashmit, P.E., Lundquist, C.A., Meier, J.L., Stinski, M.F., 2004. Cellular repressor inhibits human cytomegalovirus transcription from the UL127 promoter. J. Virol. 78 (10), 5113-5123]. However, the cellular factors which associate with the UR and FOX-like region remain to be determined. We reported previously that pancreatic-duodenal homeobox factor-1 (PDX1) bound to a 45-bp element located within the UR [Chao, S.H., Harada, J.N., Hyndman, F., Gao, X., Nelson, C.G., Chanda, S.K., Caldwell, J.S., 2004. PDX1, a Cellular Homeoprotein, Binds to and Regulates the Activity of Human Cytomegalovirus Immediate Early Promoter. J. Biol. Chem. 279 (16), 16111-16120]. Here we demonstrate that two additional cellular homeoproteins, special AT-rich sequence binding protein 1 (SATB1) and CCAAT displacement protein (CDP), bind to the human CMV UR in vitro and in vivo. Furthermore, CDP is identified as a FOX-like binding protein and a repressor of the UL127 promoter, while SATB1 has no effect on UL127 expression. Since CDP is known as a transcription repressor and a nuclear matrix-associated region binding protein, CDP may have a role in the regulation of human CMV transcription. PMID:17512569

  12. 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 than 2-fold. Genes activated in treated cells included those involved in carbohydrate binding, intracellular vesicle formation, and cellular response to stress. Conversely, PEG-silane-Qdots induce a down-regulation of genes involved in controlling the M-phase progression of mitosis, spindle formation, and cytokinesis. Promoter analysis of these results reveals that expression changes may be attributed to the down-regulation of FOXM and BHLB2 transcription factors. Remarkably, PEG-silane-Qdots, unlike carbon nanotubes, do not activate genes indicative of a strong immune and inflammatory response or heavy-metal-related toxicity. The experimental evidence shows that CdSe/ZnS Qdots, if appropriately protected, induce negligible toxicity to the model cell system studied here, even when exposed to high dosages. This study indicates that PEG-coated silanized Qdots pose minimal impact to cells and are a very promising alternative to uncoated Qdots. PMID:16608287

  13. Upregulation of Heat Shock Proteins is Essential for Cold Survival during Insect Diapause

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Diapause, the dormancy common to overwintering insects, evokes a unique pattern of gene expression. In the flesh fly most, but not all, of the fly’s heat shock proteins (Hsps) are upregulated. The diapause upregulated Hsps include two members of the Hsp70 family, one member of the Hsp60 family (TC...

  14. To Investigate the Necessity of STRA6 Upregulation in T Cells during T Cell Immune Responses

    PubMed Central

    Charpentier, Tania; Lamarre, Alain; Zhong, Ming; Sun, Hui; Mao, Jianning; Qi, Shijie; Luo, Hongyu; Wu, Jiangping

    2013-01-01

    Our earlier study revealed that STRA6 (stimulated by retinoic acid gene 6) was up-regulated within 3 h of TCR stimulation. STRA6 is the high-affinity receptor for plasma retinol-binding protein (RBP) and mediates cellular vitamin A uptake. We generated STRA6 knockout (KO) mice to assess whether such up-regulation was critical for T-cell activation, differentiation and function. STRA6 KO mice under vitamin A sufficient conditions were fertile without apparent anomalies upon visual inspection. The size, cellularity and lymphocyte subpopulations of STRA6 KO thymus and spleen were comparable to those of their wild type (WT) controls. KO and WT T cells were similar in terms of TCR-stimulated proliferation in vitro and homeostatic expansion in vivo. Naive KO CD4 cells differentiated in vitro into Th1, Th2, Th17 as well as regulatory T cells in an analogous manner as their WT counterparts. In vivo experiments revealed that anti-viral immune responses to lymphocytic choriomeningitis virus in KO mice were comparable to those of WT controls. We also demonstrated that STRA6 KO and WT mice had similar glucose tolerance. Total vitamin A levels are dramatically lower in the eyes of KO mice as compared to those of WT mice, but the levels in other organs were not significantly affected after STRA6 deletion under vitamin A sufficient conditions, indicating that the eye is the mouse organ most sensitive to the loss of STRA6. Our results demonstrate that 1) in vitamin A sufficiency, the deletion of STRA6 in T cells does no affect the T-cell immune responses so-far tested, including those depend on STAT5 signaling; 2) STRA6-independent vitamin A uptake compensated the lack of STRA6 in lymphoid organs under vitamin A sufficient conditions in mice; 3) STRA6 is critical for vitamin A uptake in the eyes even in vitamin A sufficiency. PMID:24391722

  15. Effects of AC/DC magnetic fields, frequency, and nanoparticle aspect ratio on cellular transfection of gene vectors

    NASA Astrophysics Data System (ADS)

    Ford, Kris; Mair, Lamar; Fisher, Mike; Rowshon Alam, Md.; Juliano, Rudolph; Superfine, Richard

    2008-10-01

    In order to make non-viral gene delivery a useful tool in the study and treatment of genetic disorders, it is imperative that these methodologies be further refined to yield optimal results. Transfection of magnetic nanoparticles and nanorods are used as non-viral gene vectors to transfect HeLa EGFP-654 cells that stably express a mutated enhanced green fluorescent protein (EGFP) gene. We deliver antisense oligonucleotides to these cells designed to correct the aberrant splicing caused by the mutation in the EGFP gene. We also transfect human bronchial endothelial cells and immortalized WI-38 lung cells with pEGFP-N1 vectors. To achieve this we bind the genes to magnetic nanoparticles and nanorods and introduce magnetic fields to effect transfection. We wish to examine the effects of magnetic fields on the transfection of these particles and the benefits of using alternating (AC) magnetic fields in improving transfection rates over direct (DC) magnetic fields. We specifically look at the frequency dependence of the AC field and particle aspect ratio as it pertains to influencing transfection rate. We posit that the increase in angular momentum brought about by the AC field and the high aspect ratio of the nanorod particles, is vital to generating the force needed to move the particle through the cell membrane.

  16. The Shh Signaling Pathway Is Upregulated in Multiple Cell Types in Cortical Ischemia and Influences the Outcome of Stroke in an Animal Model

    PubMed Central

    Jin, Yongmin; Raviv, Nataly; Barnett, Austin; Bambakidis, Nicholas C.; Filichia, Emily; Luo, Yu

    2015-01-01

    Recently the sonic hedgehog (shh) signaling pathway has been shown to play an important role in regulating repair and regenerative responses after brain injury, including ischemia. However, the precise cellular components that express and upregulate the shh gene and the cellular components that respond to shh signaling remain to be identified. In this study, using a distal MCA occlusion model, our data show that the shh signal is upregulated both at the cortical area near the injury site and in the adjacent striatum. Multiple cell types upregulate shh signaling in ischemic brain, including neurons, reactive astrocytes and nestin-expressing cells. The shh signaling pathway genes are also expressed in the neural stem cells (NSCs) niche in the subventricular zone (SVZ). Conditional deletion of the shh gene in nestin-expressing cells both at the SVZ niche and at the ischemic site lead to significantly more severe behavioral deficits in these shh iKO mice after cortical stroke, measured using an automated open field locomotion apparatus (Student’s t-test, p<0.05). In contrast, animals given post-stroke treatment with the shh signaling agonist (SAG) demonstrated less deficits in behavioral function, compared to vehicle-treated mice. At 7 days after stroke, SAG-treated mice showed higher values in multiple horizontal movement parameters compared to vehicle treated mice (Student’s t-test, p<0.05) whereas there were no differences in pre-stroke measurements, (Student’s t-test, p>0.05). In summary, our data demonstrate that shh signaling plays critical and ongoing roles in response to ischemic injury and modulation of shh signaling in vivo alters the functional outcome after cortical ischemic injury. PMID:25927436

  17. Endogenous Morphine/Nitric Oxide-Coupled Regulation of Cellular Physiology and Gene Expression: Implications for Cancer Biology

    PubMed Central

    Stefano, George B.; Kream, Richard M.; Mantione, Kirk J.; Sheehan, Melinda; Cadet, Patrick; Zhu, Wei; Bilfinger, Thomas V.; Esch, Tobias

    2008-01-01

    Cancer is a simplistic, yet complicated, process that promotes uncontrolled growth. In this regard, this unconstrained proliferation may represent primitive phenomena whereby cellular regulation is suspended or compromised. Given the new empirical evidence for a morphinergic presence and its profound modulatory actions on several cellular processes it is not an overstatement to hypothesize that morphine may represent a key chemical messenger in the process of modulating proliferation of diverse cells. This has been recently demonstrated by the finding of a novel opiate-alkaloid selective receptor subtype in human multilineage progenitor cells (MLPC). Adding to the significance of morphinergic signaling are the findings of its presence in plant, invertebrate and vertebrate cells, which also have been shown to synthesize this messenger as well. Interestingly, we and others have shown that some cancerous tissues contain morphine. Furthermore, in medullary histolytic reticulosis, which is exemplified by cells having hyperactivity, the mu3 (?3) opiate select receptor was not present. Thus, it would appear that morphinergic signaling has inserted itself in many processes taking a long time to evolve, including those regulating the proliferation of cells across diverse phyla. PMID:18203618

  18. Novel senescence associated gene, YPEL3, is repressed by estrogen in ER+ mammary tumor cells and required for tamoxifen-induced cellular senescence

    PubMed Central

    Tuttle, Rebecca; Miller, Kelly R.; Maiorano, J. Nicholas; Termuhlen, Paula M.; Gao, Yongping; Berberich, Steven J.

    2011-01-01

    Estrogen signaling plays an important role in breast carcinogenesis. An increased understanding of estrogen gene targets and their effects will allow for more directed and effective therapies for individuals with breast cancer, particularly those with estrogen receptor positive tumors resistant to tamoxifen therapy. Here, we identify YPEL3 as a growth suppressive protein down-regulated by estrogen in estrogen receptor positive breast cancer cell lines. Estrogen repression of YPEL3 expression was found to be independent of p53 but dependent on estrogen receptor alpha expression. Importantly, YPEL3 expression, which is induced by the removal of estrogen or treatment with tamoxifen triggers cellular senescence in MCF-7 cells while loss of YPEL3 increases the growth rate of MCF-7 cells. Taken together these findings suggest that YPEL3 may represent a potential target for directed hormonal therapy for estrogen receptor positive breast cancer patients. PMID:21671470

  19. Meta-analysis of retrograde signaling in Arabidopsis thaliana reveals a core module of genes embedded in complex cellular signaling networks.

    PubMed

    Gläßer, Christine; Haberer, Georg; Finkemeier, Iris; Pfannschmidt, Thomas; Kleine, Tatjana; Leister, Dario; Dietz, Karl-Josef; Häusler, Rainer Erich; Grimm, Bernhard; Mayer, Klaus Franz Xaver

    2014-07-01

    Plastid-to-nucleus signaling is essential for the coordination and adjustment of cellular metabolism in response to environmental and developmental cues of plant cells. A variety of operational retrograde signaling pathways have been described that are thought to be triggered by reactive oxygen species, photosynthesis redox imbalance, tetrapyrrole intermediates, and other metabolic traits. Here we report a meta-analysis based on transcriptome and protein interaction data. Comparing the output of these pathways reveals the commonalities and peculiarities stimulated by six different sources impinging on operational retrograde signaling. Our study provides novel insights into the interplay of these pathways, supporting the existence of an as-yet unknown core response module of genes being regulated under all conditions tested. Our analysis further highlights affiliated regulatory cis-elements and classifies abscisic acid and auxin-based signaling as secondary components involved in the response cascades following a plastidial signal. Our study provides a global analysis of structure and interfaces of different pathways involved in plastid-to-nucleus signaling and a new view on this complex cellular communication network. PMID:24719466

  20. [Antiapoptotic gene bcl-2 prevents cellular senescence program reactivation induced by histone deacetylase inhibitor sodium butyrate in E1A and cHa-ras transformed rat fibroblasts].

    PubMed

    Gordeev, S A; Bykova, T V; Zubova, S G; Aksenov, N D; Pospelova, T V

    2015-01-01

    We have investigated the role of apoptosis resistance gene bcl-2 in the activation of cellular senescence program induced by histone deacetylase inhibitor (HDACi) sodium butyrate (NaBut) in transformed rat fibroblasts. This study was conducted in a resistant to apoptosis induction cell line of rat embryo fibroblasts transfor- med by oncogenes E1A, cHa-ras and bcl-2 (ERasBcl). The parent cell line transformed with only EJA and cHa-ras (ERas) was used as a control. It has been found that NaBut reduces proliferation rate of ERasBcl cells significantly weaker than of ERas transformed cells, despite the fact that the G1 cell cycle arrest was observed in both cell lines. After NaBut treatment, hypertrophy of the apoptosis resistant transformants ERasBcl also was reduced compared to parent cell line ERas, due to less activation of mTORC1, which is known to control the synthesis of protein and ribosome biogenesis. The degree of mTORC1 activation was as.sessed by its target proteins phosphorylation: the ribosomal S6 protein and 4E-BP1--inhibitor of translation initiation factor eIF4E. Since cell senescence process may be associated with changes in autophagy regulation, we analyzed the dynamics of one of the main autophagosome formation markers--protein LC3. The accumulation of lipid-bound form LC3-II changes significantly in ERasBcl cells after NaBut treatment and has transient nature. The set of analyzed cellular senescence markers suggests that a high level of apoptosis resistance gene bcl-2 expression prevents the realization of tumor-suppressor senescence program induced by HDACi sodium butyrate treatment. PMID:26035971

  1. Cellular engineering

    Microsoft Academic Search

    Robert M. Nerem

    1991-01-01

    Cellular engineering applies the principles and methods of engineering to the problems of cell and molecular biology of both\\u000a a basic and applied nature. As biomedical engineering has shifted from the organ and tissue level to the cellular and sub-cellular\\u000a level, cellular engineering has emerged as a new area. A cornerstone of much of this activity is cell culture technology,

  2. Up-regulation of a thermogenesis-related gene (UCP1) and down-regulation of PPARgamma and aP2 genes in adipose tissue: possible features of the antiobesity effects of a beta3-adrenergic agonist.

    PubMed

    Margareto, J; Larrarte, E; Marti, A; Martinez, J A

    2001-06-15

    A number of experiments have demonstrated the antiobesity effects of beta(3)-adrenergic receptor stimulation by promoting thermogenesis and/or lipolysis. While many studies have been performed in order to develop beta(3)-adrenergic agonists as a novel strategy in the management of obesity, more information is needed about the mechanisms involved in thermogenesis and the actions of these drugs on adipocyte differentiation. To address this, the possible thermogenic and antiadipogenic properties of Tertatolol, a beta(3)-adrenergic agonist, in a diet-induced obesity model has been tested. Animals fed on a high-fat diet gained more weight and fat mass as compared with control and high-fat fed animals treated with Tertatolol. A RT-PCR was carried out in white adipose tissue specific genes involved in thermogenesis such as uncoupling proteins (UCPs) and adipogenesis such as peroxisome proliferator-activated receptor (PPARgamma2), retinoid receptors (RXRalpha/RARalpha), and fatty acid binding protein (aP2). Levels of UCP1 mRNA were augmented in the Tertatolol-treated group as compared to non-treated high-fat fed animals, while the beta(3)-adrenergic agonist treatment significantly decreased the expression levels of aP2 and transcription factors such as PPARgamma2 and the ratio RXRalpha/RARalpha as compared to obese rats. Altogether these data suggest that the antiobesity effects of beta(3)-adrenergic agonists are not limited to the promotion of thermogenesis and/or lipolysis and support the implication that these beta(3)-adrenergic agonists also affect fat deposition by impairing adipogenesis in white adipose tissue (WAT). PMID:11377376

  3. Use of zinc-finger nucleases to knock out the WAS gene in K562 cells: a human cellular model for Wiskott-Aldrich syndrome.

    PubMed

    Toscano, Miguel G; Anderson, Per; Muñoz, Pilar; Lucena, Gema; Cobo, Marién; Benabdellah, Karim; Gregory, Philip D; Holmes, Michael C; Martin, Francisco

    2013-03-01

    Mutations in the WAS gene cause Wiskott-Aldrich syndrome (WAS), which is characterized by eczema, immunodeficiency and microthrombocytopenia. Although the role of WASP in lymphocytes and myeloid cells is well characterized, its role on megakaryocyte (MK) development is poorly understood. In order to develop a human cellular model that mimics the megakaryocytic-derived defects observed in WAS patients we used K562 cells, a well-known model for study of megakaryocytic development. We knocked out the WAS gene in K562 cells using a zinc-finger nuclease (ZFN) pair targeting the WAS intron 1 and a homologous donor DNA that disrupted WASP expression. Knockout of WASP on K562 cells (K562WASKO cells) resulted in several megakaryocytic-related defects such as morphological alterations, lower expression of CD41, lower increments in F-actin polymerization upon stimulation, reduced CD43 expression and increased phosphatidylserine exposure. All these defects have been previously described either in WAS-knockout mice or in WAS patients, validating K562WASKO as a cell model for WAS. However, K562WASPKO cells showed also increased basal F-actin and adhesion, increased expression of CD61 and reduced expression of TGF? and Factor VIII, defects that have never been described before for WAS-deficient cells. Interestingly, these phenotypic alterations correlate with different roles for WASP in megakaryocytic differentiation. All phenotypic alterations observed in K562WASKO cells were alleviated upon expression of WAS following lentiviral transduction, confirming the role of WASP in these phenotypes. In summary, in this work we have validated a human cellular model, K562WASPKO, that mimics the megakaryocytic-related defects found in WAS-knockout mice and have found evidences for a role of WASP as regulator of megakaryocytic differentiation. We propose the use of K562WASPKO cells as a tool to study the molecular mechanisms involved in the megakaryocytic-related defects observed in WAS patients and as a cellular model to study new therapeutic strategies. PMID:23324327

  4. Low-dose cisplatin and 5-fluorouracil in combination can repress increased gene expression of cellular resistance determinants to themselves.

    PubMed

    Nishiyama, M; Yamamoto, W; Park, J S; Okamoto, R; Hanaoka, H; Takano, H; Saito, N; Matsukawa, M; Shirasaka, T; Kurihara, M

    1999-09-01

    The synergistic mechanism of cisplatin (CDDP) and 5-fluorouracil (5-FU) in combination remains unclear, despite its substantial antitumor activity, which has been demonstrated clinically. To clarify the mechanism(s), we determined the sensitivity or resistance factors to either drug in seven gastrointestinal cancer cell lines and then analyzed the altered gene expression after different exposures to CDDP and 5-FU. At the basal gene expression level, glutathione S-transferase pi (GSTpi) expression correlated with the observed resistance to CDDP, whereas dihydropyrimidine dehydrogenase (DPD) and multidrug resistance-associated protein (MRP) expression was related to 5-FU resistance. GSTpi, DPD, and MRP expression increased in response to the respective drug, but they also increased in response to the other drug as well. Additionally, 5-FU revealed a drastically increased thymidylate synthase (TS) gene expression in 5-FU-resistant cells. However, the increasing actions of CDDP and 5-FU on GSTpi, DPD, MRP, and TS expression varied according to the exposure time, concentration, and schedule. A low concentration of CDDP (1 microg/ml, 30 min) followed by 5-FU (0.5 microg/ml, 72 h) was found to cause a less increased expression of DPD, MRP, GSTpi, and TS than either drug alone, thus resulting in synergistic cytotoxicity in 5-FU-resistant COLO201 and CDDP-resistant HCC-48 cells. The sequential combination of CDDP and 5-FU inhibited the growth of human normal renal proximal tubule cells by less than 20%. Low concentrations of CDDP followed by continuous exposure to 5-FU can repress increased gene expression related to both drug resistances, thereby being synergistically cytotoxic in human gastrointestinal cancer cells. PMID:10499641

  5. Involvement of upregulation of DEPDC1 (DEP domain containing 1) in bladder carcinogenesis

    Microsoft Academic Search

    M Kanehira; Y Harada; R Takata; T Shuin; T Miki; T Fujioka; Y Nakamura; T Katagiri

    2007-01-01

    In an attempt to disclose mechanisms of bladder carcinogenesis and discover novel target molecules for development of treatment, we applied a cDNA microarray to screen genes that were significantly transactivated in bladder cancer cells. Among the upregulated genes, we here focused on a novel gene, (DEPDC1) DEP domain containing 1, whose overexpression was confirmed by northern blot and immunohistochemical analyses.

  6. Effect of hydrophobic scaffold on the cellular uptake and gene transfection activities of DNA-encapsulating liposomal nanoparticles via intracerebroventricular administration.

    PubMed

    Akita, Hidetaka; Nakatani, Taichi; Kuroki, Kimiko; Maenaka, Katsumi; Tange, Kota; Nakai, Yuta; Harashima, Hideyoshi

    2015-07-25

    Efficient DNA carriers are needed as a gene medication for curing brain disorders. In the present study, the function of a neutral lipid envelope-type nanoparticle (LNP) encapsulating pDNA was evaluated after intracerebroventricular administration. The lipid envelope was composed of a series of SS-cleavable and pH-activated lipid like materials (ssPalm) including myristic acid, vitamin A and vitamin E in the hydrophobic scaffold (LNPssPalmM, LNPssPalmA, LNPssPalmE, respectively). The LNPssPalmA and LNPssPalmE were extensively distributed in the corpus callosum, and then gene expression occurred mainly astrocytes in this region, while not in LNPssPalmM. The recombinant human ApoE3-dependent enhancement of the uptake into an astrocyte-derived cell line (KT-5) was observed in LNPssPalmA and LNPssPalmE. Thus, ApoE in the brain plays a key role in the cellular uptake of these particles by astrocytes, and this uptake is dependent on the structure of the hydrophobic scaffold. PMID:26003418

  7. Cellular effects and gene expression after exposure to amorphous silica nanoparticles in vitro Rasmus Foldbjerg1, Christiane Beer1, Jing Wang2, Duncan S. Sutherland2, Herman Autrup1

    E-print Network

    Schierup, Mikkel Heide

    Cellular effects and gene expression after exposure to amorphous silica nanoparticles in vitro, Denmark Much of the concerns regarding engineered nanoparticle (NP) toxicity are based on knowledge from previous studies on ambient and environmental particles. E.g., the effects of exposure to silica dust

  8. Sleeping Beauty transposon-based system for cellular reprogramming and targeted gene insertion in induced pluripotent stem cells

    PubMed Central

    Grabundzija, Ivana; Wang, Jichang; Sebe, Attila; Erdei, Zsuzsanna; Kajdi, Robert; Devaraj, Anantharam; Steinemann, Doris; Szuhai, Károly; Stein, Ulrike; Cantz, Tobias; Schambach, Axel; Baum, Christopher; Izsvák, Zsuzsanna; Sarkadi, Balázs; Ivics, Zoltán

    2013-01-01

    The discovery of direct cell reprogramming and induced pluripotent stem (iPS) cell technology opened up new avenues for the application of non-viral, transposon-based gene delivery systems. The Sleeping Beauty (SB) transposon is highly advanced for versatile genetic manipulations in mammalian cells. We established iPS cell reprogramming of mouse embryonic fibroblasts and human foreskin fibroblasts by transposition of OSKM (Oct4, Sox2, Klf4 and c-Myc) and OSKML (OSKM + Lin28) expression cassettes mobilized by the SB100X hyperactive transposase. The efficiency of iPS cell derivation with SB transposon system was in the range of that obtained with retroviral vectors. Co-expression of the miRNA302/367 cluster together with OSKM significantly improved reprogramming efficiency and accelerated the temporal kinetics of reprogramming. The iPS cells displayed a stable karyotype, and hallmarks of pluripotency including expression of stem cell markers and the ability to differentiate into embryoid bodies in vitro. We demonstrate Cre recombinase-mediated exchange allowing simultaneous removal of the reprogramming cassette and targeted knock-in of an expression cassette of interest into the transposon-tagged locus in mouse iPS cells. This strategy would allow correction of a genetic defect by site-specific insertion of a therapeutic gene construct into ‘safe harbor’ sites in the genomes of autologous, patient-derived iPS cells. PMID:23275558

  9. Long-Term Channel Block Is Required to Inhibit Cellular Transformation by Human Ether-à-Go-Go–Related Gene (hERG1) Potassium Channels

    PubMed Central

    Pier, David M.; Shehatou, George S. G.; Giblett, Susan; Pullar, Christine E.; Trezise, Derek J.; Pritchard, Catrin A.; Challiss, R. A. John

    2014-01-01

    Both human ether-à-go-go–related gene (hERG1) and the closely related human ether-à-go-go (hEAG1) channel are aberrantly expressed in a large proportion of human cancers. In the present study, we demonstrate that transfection of hERG1 into mouse fibroblasts is sufficient to induce many features characteristic of malignant transformation. An important finding of this work is that this transformation could be reversed by chronic incubation (for 2–3 weeks) with the hERG channel blocker dofetilide (100 nM), whereas more acute applications (for 1–2 days) were ineffective. The hERG1 expression resulted in a profound loss of cell contact inhibition, multiple layers of overgrowing cells, and high saturation densities. Cells also changed from fibroblast-like to a more spindle-shaped morphology, which was associated with a smaller cell size, a dramatic increase in cell polarization, a reduction in the number of actin stress fibers, and less punctate labeling of focal adhesions. Analysis of single-cell migration and scratch-wound closure clearly demonstrated that hERG1-expressing cells migrated more rapidly than vector-transfected control cells. In contrast to previous studies on hEAG1, there were no increases in rates of proliferation, or loss of growth factor dependency; however, hERG1-expressing cells were capable of substrate-independent growth. Allogeneic transplantation of hERG1-expressing cells into nude mice resulted in an increased incidence of tumors. In contrast to hEAG1, the mechanism of cellular transformation is dependent on ion conduction. Trafficking-deficient and conduction-deficient hERG1 mutants also prevented cellular transformation. These results provide evidence that hERG1 expression is sufficient to induce cellular transformation by a mechanism distinct from hEAG1. The most important conclusion of this study is that selective hERG1 channel blockers have therapeutic potential in the treatment of hERG1-expressing cancers. PMID:24830940

  10. 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-?B 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-?B pathway-controlled AMPs, such as shrimp Penaeidins and Drosophila drosomycin and attacin A, in Drosophila S2 cells. Taken together, these results reveal that LvIAP1 and LvIAP3 might participate in the host defense against WSSV infection, and LvIAP2 might be involved in the regulation of shrimp AMPs. PMID:23967321

  11. EBV BART MicroRNAs Target Multiple Pro-apoptotic Cellular Genes to Promote Epithelial Cell Survival

    PubMed Central

    Kang, Dong; Skalsky, Rebecca L.; Cullen, Bryan R.

    2015-01-01

    Epstein-Barr virus (EBV) is a ubiquitous human ?-herpesvirus that can give rise to cancers of both B-cell and epithelial cell origin. In EBV-induced cancers of epithelial origin, including nasopharyngeal carcinomas (NPCs) and gastric carcinomas, the latent EBV genome expresses very high levels of a cluster of 22 viral pre-miRNAs, called the miR-BARTs, and these have previously been shown to confer a degree of resistance to pro-apoptotic drugs. Here, we present an analysis of the ability of individual miR-BART pre-miRNAs to confer an anti-apoptotic phenotype and report that five of the 22 miR-BARTs demonstrate this ability. We next used photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) to globally identify the mRNA targets bound by these miR-BARTs in latently infected epithelial cells. This led to the identification of ten mRNAs encoding pro-apoptotic mRNA targets, all of which could be confirmed as valid targets for the five anti-apoptotic miR-BARTs by indicator assays and by demonstrating that ectopic expression of physiological levels of the relevant miR-BART in the epithelial cell line AGS resulted in a significant repression of the target mRNA as well as the encoded protein product. Using RNA interference, we further demonstrated that knockdown of at least seven of these cellular miR-BART target transcripts phenocopies the anti-apoptotic activity seen upon expression of the relevant EBV miR-BART miRNA. Together, these observations validate previously published reports arguing that the miR-BARTs can exert an anti-apoptotic effect in EBV-infected epithelial cells and provide a mechanistic explanation for this activity. Moreover, these results identify and validate a substantial number of novel mRNA targets for the anti-apoptotic miR-BARTs. PMID:26070070

  12. EBV BART MicroRNAs Target Multiple Pro-apoptotic Cellular Genes to Promote Epithelial Cell Survival.

    PubMed

    Kang, Dong; Skalsky, Rebecca L; Cullen, Bryan R

    2015-06-01

    Epstein-Barr virus (EBV) is a ubiquitous human ?-herpesvirus that can give rise to cancers of both B-cell and epithelial cell origin. In EBV-induced cancers of epithelial origin, including nasopharyngeal carcinomas (NPCs) and gastric carcinomas, the latent EBV genome expresses very high levels of a cluster of 22 viral pre-miRNAs, called the miR-BARTs, and these have previously been shown to confer a degree of resistance to pro-apoptotic drugs. Here, we present an analysis of the ability of individual miR-BART pre-miRNAs to confer an anti-apoptotic phenotype and report that five of the 22 miR-BARTs demonstrate this ability. We next used photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) to globally identify the mRNA targets bound by these miR-BARTs in latently infected epithelial cells. This led to the identification of ten mRNAs encoding pro-apoptotic mRNA targets, all of which could be confirmed as valid targets for the five anti-apoptotic miR-BARTs by indicator assays and by demonstrating that ectopic expression of physiological levels of the relevant miR-BART in the epithelial cell line AGS resulted in a significant repression of the target mRNA as well as the encoded protein product. Using RNA interference, we further demonstrated that knockdown of at least seven of these cellular miR-BART target transcripts phenocopies the anti-apoptotic activity seen upon expression of the relevant EBV miR-BART miRNA. Together, these observations validate previously published reports arguing that the miR-BARTs can exert an anti-apoptotic effect in EBV-infected epithelial cells and provide a mechanistic explanation for this activity. Moreover, these results identify and validate a substantial number of novel mRNA targets for the anti-apoptotic miR-BARTs. PMID:26070070

  13. A novel embryological theory of autism causation involving endogenous biochemicals capable of initiating cellular gene transcription: a possible link between twelve autism risk factors and the autism 'epidemic'.

    PubMed

    King, Chiara R

    2011-05-01

    Human alpha-fetoprotein is a pregnancy-associated protein with an undetermined physiological role. As human alpha-fetoprotein binds retinoids and inhibits estrogen-dependent cancer cell proliferation, and because retinoic acid (a retinol metabolite) and estradiol (an estrogen) can both initiate cellular gene transcription, it is hypothesized here that alpha-fetoprotein functions during critical gestational periods to prevent retinoic acid and maternal estradiol from inappropriately stimulating gene expression in developing brain regions which are sensitive to these chemicals. Prenatal/maternal factors linked to increased autism risk include valproic acid, thalidomide, alcohol, rubella, cytomegalovirus, depression, schizophrenia, obsessive-compulsive disorder, autoimmune disease, stress, allergic reaction, and hypothyroidism. It will be shown how each of these risk factors may initiate expression of genes which are sensitive to retinoic acid and/or estradiol - whether by direct promotion or by reducing production of alpha-fetoprotein. It is thus hypothesized here that autism is not a genetic disorder, but is rather an epigenetic disruption in brain development caused by gestational exposure to chemicals and/or conditions which either inhibit alpha-fetoprotein production or directly promote retinoic acid-sensitive or estradiol-sensitive gene expression. This causation model leads to potential chemical explanations for autistic brain morphology, the distinct symptomatology of Asperger's syndrome, and the differences between high-functioning and low-functioning autism with regard to mental retardation, physical malformation, and sex ratio. It will be discussed how folic acid may cause autism under the retinoic acid/estradiol model, and the history of prenatal folic acid supplementation will be shown to coincide with the history of what is popularly known as the autism epidemic. It is thus hypothesized here that prenatal folic acid supplementation has contributed to the post-1980 increase in US autism diagnoses. In addition to explaining the epidemic within the wider retinoic acid/estradiol model of causation, this theory leads to potential explanations for certain genetic findings in autism, autistic regression, and changing trends in autism symptomatology with regard to mental retardation, wheat allergy, and gastrointestinal problems. PMID:21388746

  14. Expression of the prion protein gene (PRNP) and cellular prion protein (PrPc) in cattle and sheep fetuses and maternal tissues during pregnancy.

    PubMed

    Thumdee, Patama; Ponsuksili, Siriluck; Murani, Eduard; Nganvongpanit, Korakot; Gehrig, Bernhard; Tesfaye, Dawit; Gilles, Markus; Hoelker, Michael; Jennen, Danyel; Griese, Josef; Schellander, Karl; Wimmers, Klaus

    2007-01-01

    We investigated the expression of prion protein gene both on mRNA and protein levels in bovine and ovine female reproductive organs during gestation and various tissues of their fetuses. The fetal tissues of both species included brain, cotyledon, heart, intestine, kidney, liver, lung, and muscle. In cattle, prion protein gene (PRNP) transcripts were detected by semiquantitative RT-PCR in reproductive tissues such as ovary, oviduct, endometrium, myometrium, follicles, and granulosa cells. In various tissues of 2-month-old fetuses, higher expression levels were found in brain and cotyledon compared to the other tissues. To detect the expression of the gene transcript in in vivo preimplantation embryos and 1-month-old fetuses, real-time PCR was performed showing that the level of gene expression in zygote stage was significantly higher (p < or = 0.05) than that of the other stages. Sheep were categorized as resistant (RI) or high susceptible (R5) to scrapie according to their PRNP genotype. In both genotype groups, the PRNP mRNA was detectable in all tissues studied including ovary, oviduct, endometrium, myometrium, and caruncle of ewes and all tissues of 2-month-old fetuses of both groups. Comparison between reproductive organs demonstrates the highest expression level in caruncle tissue of R1 ewes, whereas the level was high in brain and low in liver of both R1 and R5 fetuses. In addition, real-time RT-PCR was performed in immature oocytes, mature oocytes, in vivo embryos at morula stage, and 1-month-old fetuses. The results showed that the relative expression levels of the ovine PRNP mRNA in mature oocytes and morula stage embryos were significantly lower than those in immature oocytes and 1-month-old fetuses (p < or = 0.05). Western blot analyses revealed the immunoreactive bands corresponding to the cellular prion protein (PrPc) in all maternal and fetal tissues examined of both cattle and sheep. Moreover, immunohistochemical staining implicated localization of the PrPc in ovarian cortex and ovarian medulla of both species. However, PrPc was not detected in oocyte, granulosa cells, theca cells, and corpus luteum in this study. PMID:17605301

  15. The transcription elongation factor ELL2 is specifically upregulated in HTLV-1-infected T-cells and is dependent on the viral oncoprotein Tax.

    PubMed

    Mann, Melanie C; Strobel, Sarah; Fleckenstein, Bernhard; Kress, Andrea K

    2014-09-01

    The oncoprotein Tax of human T-cell leukemia virus type 1 (HTLV-1) is a potent transactivator of viral and cellular transcription. Here, we identified ELL2 as the sole transcription elongation factor to be specifically upregulated in HTLV-1-/Tax-transformed T-cells. Tax contributes to regulation of ELL2, since transient transfection of Tax increases ELL2 mRNA, Tax transactivates the ELL2 promoter, and repression of Tax results in decrease of ELL2 in transformed T-lymphocytes. However, we also measured upregulation of ELL2 in HTLV-1-transformed cells exhibiting undetectable amounts of Tax, suggesting that ELL2 can still be maintained independent of continuous Tax expression. We further show that Tax and ELL2 synergistically activate the HTLV-1 promoter, indicating that ELL2 cooperates with Tax in viral transactivation. This is supported by our findings that Tax and ELL2 accumulate in nuclear fractions and that they co-precipitate upon co-expression in transiently-transfected cells. Thus, upregulation of ELL2 could contribute to HTLV-1 gene regulation. PMID:25058508

  16. Cellular and molecular mechanisms of accelerated fracture healing by COX2 gene therapy: studies in a mouse model of multiple fractures.

    PubMed

    Lau, K-H William; Kothari, Vishal; Das, Amitava; Zhang, Xiao-Bing; Baylink, David J

    2013-04-01

    This study sought to determine the cellular and molecular mechanisms of cyclooxygenase-2 (COX2) gene therapy to accelerate fracture repair in a mouse multiple tibial fractures model. The lenti-COX2 (or lenti-gfp control vector) was injected into fractures on day 1 post-fracture. At days 3-7, the COX2 treatment increased Sdf1-, Cxcr4-, Nes-, and Podxl-expressing mesenchymal stem cells (MSCs) within fracture calluses, suggesting an enhanced MSC recruitment or expansion. The COX2-treated mice formed smaller cartilaginous calluses that had less cartilage tissues than control mice. The expression of Sox9 mRNA was 7-fold less in COX2-treated than in control calluses at day 14, implying that COX2 reduces chondrocytic differentiation of MSCs. The therapy also enhanced angiogenesis as reflected by increased immunostaining of CD31, vWF, and ?-SMA over controls in the cartilaginous callus at day 14-21. At which time, the COX2 gene therapy promoted bony remodeling of the cartilaginous callus to bridge the fracture gap that was accompanied by 2-fold increase in osteoclasts along the surface of the woven bone and an onset of osteogenesis. Blocking angiogenesis with daily injection of endostatin from day 4 to day 10 into fracture sites blocked the COX2-mediated reduction of callus size that was associated with an increase in hypertrophic chondrocytes and concomitant reduction in osteoclasts. In conclusion, COX2 accelerates fracture healing in part through three biological actions: 1) increased recruitment/expansion of MSCs; 2) decreased cartilaginous callus formation; and 3) increased angiogenesis-dependent cartilage remodeling. These effects were associated with an earlier onset of bony bridging of the fracture gap. PMID:23314071

  17. Calcitonin gene-related peptide promotes cellular changes in trigeminal neurons and glia implicated in peripheral and central sensitization

    PubMed Central

    2011-01-01

    Background Calcitonin gene-related peptide (CGRP), a neuropeptide released from trigeminal nerves, is implicated in the underlying pathology of temporomandibular joint disorder (TMD). Elevated levels of CGRP in the joint capsule correlate with inflammation and pain. CGRP mediates neurogenic inflammation in peripheral tissues by increasing blood flow, recruiting immune cells, and activating sensory neurons. The goal of this study was to investigate the capability of CGRP to promote peripheral and central sensitization in a model of TMD. Results Temporal changes in protein expression in trigeminal ganglia and spinal trigeminal nucleus were determined by immunohistochemistry following injection of CGRP in the temporomandibular joint (TMJ) capsule of male Sprague-Dawley rats. CGRP stimulated expression of the active forms of the MAP kinases p38 and ERK, and PKA in trigeminal ganglia at 2 and 24 hours. CGRP also caused a sustained increase in the expression of c-Fos neurons in the spinal trigeminal nucleus. In contrast, levels of P2X3 in spinal neurons were only significantly elevated at 2 hours in response to CGRP. In addition, CGRP stimulated expression of GFAP in astrocytes and OX-42 in microglia at 2 and 24 hours post injection. Conclusions Our results demonstrate that an elevated level of CGRP in the joint, which is associated with TMD, stimulate neuronal and glial expression of proteins implicated in the development of peripheral and central sensitization. Based on our findings, we propose that inhibition of CGRP-mediated activation of trigeminal neurons and glial cells with selective non-peptide CGRP receptor antagonists would be beneficial in the treatment of TMD. PMID:22145886

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

    PubMed

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

    2015-04-01

    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 ?m, 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

  19. DNA Microarray Analysis of Cyanobacterial Gene Expression during Acclimation to High Light

    PubMed Central

    Hihara, Yukako; Kamei, Ayako; Kanehisa, Minoru; Kaplan, Aaron; Ikeuchi, Masahiko

    2001-01-01

    DNA microarrays bearing nearly all of the genes of the unicellular cyanobacterium Synechocystis sp PCC 6803 were used to examine the temporal program of gene expression during acclimation from low to high light intensity. A complete pattern is provided of gene expression during acclimation of a photosynthetic organism to changing light intensity. More than 160 responsive genes were identified and classified into distinct sets. Genes involved in light absorption and photochemical reactions were downregulated within 15 min of exposure to high light intensity, whereas those associated with CO2 fixation and protection from photoinhibition were upregulated. Changes in the expression of genes involved in replication, transcription, and translation, which were induced to support cellular proliferation, occurred later. Several unidentified open reading frames were induced or repressed. The possible involvement of these genes in the acclimation to high light conditions is discussed. PMID:11283337

  20. Induction of interferon-stimulated genes by Simian virus 40 T antigens

    SciTech Connect

    Rathi, Abhilasha V.; Cantalupo, Paul G. [Department of Biological Sciences, University of Pittsburgh, 559 Crawford Hall, Pittsburgh, PA 15260 (United States); Sarkar, Saumendra N. [University of Pittsburgh Cancer Institute, Hillman Cancer Research Pavilion, 5117 Centre Avenue, Pittsburgh, PA 15213 (United States); Pipas, James M., E-mail: pipas@pitt.ed [Department of Biological Sciences, University of Pittsburgh, 559 Crawford Hall, Pittsburgh, PA 15260 (United States)

    2010-10-25

    Simian virus 40 (SV40) large T antigen (TAg) is a multifunctional oncoprotein essential for productive viral infection and for cellular transformation. We have used microarray analysis to examine the global changes in cellular gene expression induced by wild-type T antigen (TAg{sup wt}) and TAg-mutants in mouse embryo fibroblasts (MEFs). The expression profile of approximately 800 cellular genes was altered by TAg{sup wt} and a truncated TAg (TAg{sup N136}), including many genes that influence cell cycle, DNA-replication, transcription, chromatin structure and DNA repair. Unexpectedly, we found a significant number of immune response genes upregulated by TAg{sup wt} including many interferon-stimulated genes (ISGs) such as ISG56, OAS, Rsad2, Ifi27 and Mx1. Additionally, we also observed activation of STAT1 by TAg{sup wt}. Our genetic studies using several TAg-mutants reveal an unexplored function of TAg and indicate that the LXCXE motif and p53 binding are required for the upregulation of ISGs.

  1. Alternation in the Glycolipid Transfer Protein Expression Causes Changes in the Cellular Lipidome

    PubMed Central

    Kjellberg, Matti A.; Backman, Anders P. E.; Ohvo-Rekilä, Henna; Mattjus, Peter

    2014-01-01

    The glycolipid transfer protein (GLTP) catalyzes the binding and transport of glycolipids, but not phospholipids or neutral lipids. With its all-alpha helical fold, it is the founding member for a new superfamily, however its biological role still remains unclear. We have analyzed changes in the HeLa cell lipidome in response to down- and up-regulation of GLTP expression. We used metabolic labeling and thin layer chromatography analysis, complemented with a lipidomics mass spectroscopic approach. HeLa cells were treated with GLTP siRNA or were transiently overexpressing the GLTP gene. We identified eight different lipid classes that changed as a result of the GLTP down- or up-regulation treatments; glucosylceramide, lactosylceramide, globotriaosylceramide, ceramide, sphingomyelin, cholesterol-esters, diacylglycerol and phosphatidylserine. We discovered that the amount of globotriaosylceramide (Gb3) was extensively lowered after down-regulation of GLTP. Further, an up-regulation of GLTP caused a substantial increase in both the Gb3 and glucosylceramide levels compared to the controls. Total galactosylceramide levels remained unchanged. Both lactosylceramide and ceramide showed small changes, an increase with increasing GLTP and a decrease in the HeLa cell GLTP knockdowns. The cholesterol-esters and diacylglycerol masses increased in cells that had upregulated GLTP protein levels, wheras down-regulation did not affect their amounts. For the glycerophospholipids, phosphatidylserine was the only species that was lower in GLTP overexpressing cells. Phosphatidylethanolamine, phosphatidylglyerol and phosphatidylinositol remained unaltered. A total of 142 lipid species were profiled and quantified using shotgun lipidomics analyses. This work provides for the first time insights into how alternations in the levels of a protein that binds and transfers glycolipids affects the cellular lipid metabolism. We discuss the observed changes in the lipidome and how these relate to GLTP. We suggest, that GLTP not only could be a significant player in cellular sphingolipid metabolism, but also could have a much broader role in the overall lipid metabolism. PMID:24824606

  2. Bexarotene induces cellular senescence in MMTV-Neu mouse model of mammary carcinogenesis.

    PubMed

    Shilkaitis, Anne; Bratescu, Laura; Green, Albert; Yamada, Tohru; Christov, Konstantin

    2013-04-01

    Previous studies have shown that retinoids and rexinoids can prevent breast cancer in animal models and in women with increased risk of developing the disease. The cellular effects of these vitamin A analogues have been primarily associated with induction of differentiation and inhibition of proliferation. In this study, we tested the hypothesis that bexarotene (LGD1069, Targretin), a rexinoid, can not only inhibit cell proliferation but also induce cellular senescence in mammary epithelial cells, premalignant lesions, and tumors of the MMTV-Neu model of mammary carcinogenesis, which develops estrogen receptor-negative tumors. Mice with palpable mammary tumors were treated for 4 weeks with bexarotene at 80 or 40 mg/kg body weight, and senescent cells were determined by SA-?-Gal assay. Bexarotene decreased in a dose-dependent manner the multiplicity of premalignant lesions and tumors, and this was associated with inhibition of cell proliferation and induction of cellular senescence and apoptosis. By double labeling of senescent cells, first by SA-?-Gal and then by antibodies against genes related to cellular senescence, we found that p21, p16, and RAR?, but not p53, were upregulated by bexarotene in mammary tumors and in breast cancer cell lines, suggesting involvement of multiple signaling pathways in mediating the senescence program of rexinoids. These findings indicate that, in addition to cell proliferation and apoptosis, cellular senescence could be used as a potential biomarker of response in breast cancer prevention and therapy studies with rexinoids and possibly with other antitumor agents. PMID:23430755

  3. Genome-wide differential gene expression in immortalized DF-1 chicken embryo fibroblast cell line

    PubMed Central

    2011-01-01

    Background When compared to primary chicken embryo fibroblast (CEF) cells, the immortal DF-1 CEF line exhibits enhanced growth rates and susceptibility to oxidative stress. Although genes responsible for cell cycle regulation and antioxidant functions have been identified, the genome-wide transcription profile of immortal DF-1 CEF cells has not been previously reported. Global gene expression in primary CEF and DF-1 cells was performed using a 4X44K chicken oligo microarray. Results A total of 3876 differentially expressed genes were identified with a 2 fold level cutoff that included 1706 up-regulated and 2170 down-regulated genes in DF-1 cells. Network and functional analyses using Ingenuity Pathways Analysis (IPA, Ingenuity® Systems, http://www.ingenuity.com) revealed that 902 of 3876 differentially expressed genes were classified into a number of functional groups including cellular growth and proliferation, cell cycle, cellular movement, cancer, genetic disorders, and cell death. Also, the top 5 gene networks with intermolecular connections were identified. Bioinformatic analyses suggested that DF-1 cells were characterized by enhanced molecular mechanisms for cell cycle progression and proliferation, suppressing cell death pathways, altered cellular morphogenesis, and accelerated capacity for molecule transport. Key molecules for these functions include E2F1, BRCA1, SRC, CASP3, and the peroxidases. Conclusions The global gene expression profiles provide insight into the cellular mechanisms that regulate the unique characteristics observed in immortal DF-1 CEF cells. PMID:22111699

  4. Identification of cellular target genes of the Epstein-Barr virus transactivator Zta: activation of transforming growth factor beta igh3 (TGF-beta igh3) and TGF-beta 1.

    PubMed Central

    Cayrol, C; Flemington, E K

    1995-01-01

    The lytic switch transactivator Zta initiates the ordered cascade of Epstein-Barr virus gene expression that culminates in virus production. Zta is a sequence-specific DNA-binding protein that transactivates early viral promotes via cis-acting sequences. Activation of some of these genes is mediated through binding to consensus AP-1 promoter elements. This observation suggests that Zta may also regulate the expression of cellular genes. While many targets of Zta have been identified in the Epstein-Barr virus genome, putative host cell targets remain largely unknown. To address this issue, a tetracycline-regulated Zta expression system was generated, and differential hybridization screening was used to isolate Zta-responsive cellular genes. The major target identified by this analysis is a gene encoding a fasciclin-like secreted factor, transforming growth factor beta igh3 (TGF-beta igh3), that was originally identified as a gene that is responsive to the potent immunosuppressor TGF-beta 1. Northern (RNA) blot analysis demonstrated that induction of Zta expression results in a 10-fold increase in TGF-beta igh3 mRNA levels. Zta was also found to increase TGF-beta 1 mRNA levels as well as the amount of active TGF-beta 1 secreted into the medium. Interestingly, alpha 1-collagen IV, which has been shown to potentiate the effects of TGF-beta 1, is also a cellular target of Zta. These results suggest that Zta could play a role in modulating the host cell environment through activating the expression of secreted factors. PMID:7769680

  5. The Effects of Dietary n-3 Polyunsaturated Fatty Acids Delivered in Chylomicron Remnants on the Transcription of Genes Regulating Synthesis and Secretion of Very-Low-Density Lipoprotein by the Liver: Modulation by Cellular Oxidative State

    Microsoft Academic Search

    KATHLEEN M. BOTHAM; X IAOZHONG ZHENG; MARIAROSARIA NAPOLITANO; MICHAEL AVELLA; CLAUDIO CAVALLARI; ROBERTO RIVABENE; ELENA BRAVO

    The influence of chylomicron remnants enriched in n-3 or n-6 polyunsaturated fatty acids (PUFA) (derived from fish or corn oil, respectively) on the expression of mRNA for four genes involved in the regulation of the synthesis, assembly, and se- cretion of very-low-density lipoprotein (VLDL) in the liver was investigated in normal rat hepatocytes and after manipulation of the cellular oxidative

  6. Overexpressing cellular repressor of E1A-stimulated genes protects mesenchymal stem cells against hypoxia- and serum deprivation-induced apoptosis by activation of PI3K\\/Akt

    Microsoft Academic Search

    Jie Deng; Yaling Han; Chenhui Yan; XIaoxiang Tian; Jie Tao; Jian Kang; Shaohua Li

    2010-01-01

    Bone marrow-derived mesenchymal stem cells (MSCs) have great potential for repair after myocardial infarction. However, poor\\u000a viability of transplanted MSCs in the ischemic heart has limited their therapeutic potential. Cellular repressor of E1A-stimulated\\u000a genes (CREG) has been identified as a potent inhibitor of apoptosis. The aim of this study was to investigate the anti-apoptotic\\u000a effects of CREG on MSCs under

  7. Human Cytomegalovirus Upregulates Expression of the Lectin Galectin 9 via Induction of Beta Interferon

    PubMed Central

    McSharry, Brian P.; Forbes, Simone K.; Cao, John Z.; Avdic, Selmir; Machala, Emily A.; Gottlieb, David J.; Abendroth, Allison

    2014-01-01

    Regulation of the lectin galectin 9 (Gal-9) was investigated for the first time during human cytomegalovirus (HCMV) infection. Gal-9 transcription was significantly upregulated in transplant recipients with reactivated HCMV in vivo. In vitro, Gal-9 was potently upregulated by HCMV independently of viral gene expression, with interferon beta (IFN-?) identified as the mediator of this effect. This study defines an immunoregulatory protein potently increased by HCMV infection and a novel mechanism to control Gal-9 through IFN-? induction. PMID:25008927

  8. Molecular and cellular mechanisms of cardiotoxicity.

    PubMed Central

    Kang, Y J

    2001-01-01

    Cardiotoxicity resulting from detrimental environmental insults has been recognized for a long time. However, extensive studies of the mechanisms involved had not been undertaken until recent years. Advances in molecular biology provide powerful tools and make such studies possible. We are gathering information about cellular events, signaling pathways, and molecular mechanisms of myocardial toxicologic responses to environmental toxicants and pollutants. Severe acute toxic insults cause cardiac cell death instantly. In the early response to mild environmental stimuli, biochemical changes such as alterations in calcium homeostasis occur. These may lead to cardiac arrhythmia, which most often is reversible. Prolonged stimuli activate transcription factors such as activator protein-1 through elevation of intracellular calcium and the subsequent activation of calcineurin. Upregulation by activated transcription factors of hypertrophic genes results in heart hypertrophy, which is a short-term adaptive response to detrimental factors. However, further development of hypertrophy will lead to severe and irreversible cardiomyopathy, and eventually heart failure. From cardiac hypertrophy to heart failure, myocardial cells undergo extensive biochemical and molecular changes. Cardiac hypertrophy causes tissue hypoperfusion, which activates compensatory mechanisms such as production of angiotensin II and norepinephrine. Both further stimulate cardiac hypertrophy and, importantly, activate counterregulatory mechanisms including overexpression of atrial natriuretic peptide and b-type natriuretic peptide, and production of cytokines such as tumor necrosis factor-alpha. This counterregulation leads to myocardial remodeling as well as cell death through apoptosis and necrosis. Cell death through activation of mitochondrial factors and other pathways constitutes an important cellular mechanism of heart failure. Our current knowledge of cardiotoxicity is limited. Further extensive studies are warranted for a comprehensive understanding of this field. PMID:11250803

  9. Antibody-Dependent Cellular Cytotoxicity-Mediating Antibodies from an HIV-1 Vaccine Efficacy Trial Target Multiple Epitopes and Preferentially Use the VH1 Gene Family

    PubMed Central

    Pollara, Justin; Moody, M. Anthony; Alpert, Michael D.; Chen, Xi; Hwang, Kwan-Ki; Gilbert, Peter B.; Huang, Ying; Gurley, Thaddeus C.; Kozink, Daniel M.; Marshall, Dawn J.; Whitesides, John F.; Tsao, Chun-Yen; Kaewkungwal, Jaranit; Nitayaphan, Sorachai; Pitisuttithum, Punnee; Rerks-Ngarm, Supachai; Kim, Jerome H.; Michael, Nelson L.; Tomaras, Georgia D.; Montefiori, David C.; Lewis, George K.; DeVico, Anthony; Evans, David T.; Ferrari, Guido; Liao, Hua-Xin; Haynes, Barton F.

    2012-01-01

    The ALVAC-HIV/AIDSVAX-B/E RV144 vaccine trial showed an estimated efficacy of 31%. RV144 secondary immune correlate analysis demonstrated that the combination of low plasma anti-HIV-1 Env IgA antibodies and high levels of antibody-dependent cellular cytotoxicity (ADCC) inversely correlate with infection risk. One hypothesis is that the observed protection in RV144 is partially due to ADCC-mediating antibodies. We found that the majority (73 to 90%) of a representative group of vaccinees displayed plasma ADCC activity, usually (96.2%) blocked by competition with the C1 region-specific A32 Fab fragment. Using memory B-cell cultures and antigen-specific B-cell sorting, we isolated 23 ADCC-mediating nonclonally related antibodies from 6 vaccine recipients. These antibodies targeted A32-blockable conformational epitopes (n = 19), a non-A32-blockable conformational epitope (n = 1), and the gp120 Env variable loops (n = 3). Fourteen antibodies mediated cross-clade target cell killing. ADCC-mediating antibodies displayed modest levels of V-heavy (VH) chain somatic mutation (0.5 to 1.5%) and also displayed a disproportionate usage of VH1 family genes (74%), a phenomenon recently described for CD4-binding site broadly neutralizing antibodies (bNAbs). Maximal ADCC activity of VH1 antibodies correlated with mutation frequency. The polyclonality and low mutation frequency of these VH1 antibodies reveal fundamental differences in the regulation and maturation of these ADCC-mediating responses compared to VH1 bNAbs. PMID:22896626

  10. Molecular and cellular changes in the lumbar spinal cord following thoracic injury: regulation by treadmill locomotor training.

    PubMed

    Shin, Hae Young; Kim, Hyosil; Kwon, Min Jung; Hwang, Dong Hoon; Lee, KiYoung; Kim, Byung Gon

    2014-01-01

    Traumatic spinal cord injury (SCI) often leads to debilitating loss of locomotor function. Neuroplasticity of spinal circuitry underlies some functional recovery and therefore represents a therapeutic target to improve locomotor function following SCI. However, the cellular and molecular mechanisms mediating neuroplasticity below the lesion level are not fully understood. The present study performed a gene expression profiling in the rat lumbar spinal cord at 1 and 3 weeks after contusive SCI at T9. Another group of rats received treadmill locomotor training (TMT) until 3 weeks, and gene expression profiles were compared between animals with and without TMT. Microarray analysis showed that many inflammation-related genes were robustly upregulated in the lumbar spinal cord at both 1 and 3 weeks after thoracic injury. Notably, several components involved in an early complement activation pathway were concurrently upregulated. In line with the microarray finding, the number of microglia substantially increased not only in the white matter but also in the gray matter. C3 and complement receptor 3 were intensely expressed in the ventral horn after injury. Furthermore, synaptic puncta near ventral motor neurons were frequently colocalized with microglia after injury, implicating complement activation and microglial cells in synaptic remodeling in the lumbar locomotor circuitry after SCI. Interestingly, TMT did not influence the injury-induced upregulation of inflammation-related genes. Instead, TMT restored pre-injury expression patterns of several genes that were downregulated by injury. Notably, TMT increased the expression of genes involved in neuroplasticity (Arc, Nrcam) and angiogenesis (Adam8, Tie1), suggesting that TMT may improve locomotor function in part by promoting neurovascular remodeling in the lumbar motor circuitry. PMID:24520355

  11. Iron upregulates melanogenesis in cultured retinal pigment epithelial cells.

    PubMed

    Wolkow, Natalie; Li, Yafeng; Maminishkis, Arvydas; Song, Ying; Alekseev, Oleg; Iacovelli, Jared; Song, Delu; Lee, Jennifer C; Dunaief, Joshua L

    2014-11-01

    The purpose of our studies was to examine the relationship between iron and melanogenesis in retinal pigment epithelial cells, as prior observations had suggested that iron may promote melanogenesis. This relationship has potential clinical importance, as both iron overload and hyperpigmentation are associated with age-related macular degeneration (AMD). Human fetal retinal pigment epithelial cells and ARPE-19 cells were treated with iron in the form of ferric ammonium citrate, after which quantitative RT-PCR and electron microscopy were performed. Melanogenesis genes tyrosinase, tyrosinase-related protein 1, Hermansky-Pudlak Syndrome 3, premelanosome protein and dopachrome tautomerase were upregulated, as was the melanogenesis-controlling transcription factor, microphthalmia-associated transcription factor (MITF). Iron-treated cells had increased pigmentation and melanosome number. Multiple transcription factors upstream of MITF were upregulated by iron. PMID:25277027

  12. Oxidized low-density lipoproteins upregulate proline oxidase to initiate ROS-dependent autophagy

    PubMed Central

    Zabirnyk, Olga; Liu, Wei; Khalil, Shadi; Sharma, Anit; Phang, James M.

    2010-01-01

    Epidemiological studies showed that high levels of oxidized low-density lipoproteins (oxLDLs) are associated with increased cancer risk. We examined the direct effect of physiologic concentrations oxLDL on cancer cells. OxLDLs were cytotoxic and activate both apoptosis and autophagy. OxLDLs have ligands for peroxisome proliferator-activated receptor gamma and upregulated proline oxidase (POX) through this nuclear receptor. We identified 7-ketocholesterol (7KC) as a main component responsible for the latter. To elucidate the role of POX in oxLDL-mediated cytotoxicity, we knocked down POX via small interfering RNA and found that this (i) further reduced viability of cancer cells treated with oxLDL; (ii) decreased oxLDL-associated reactive oxygen species generation; (iii) decreased autophagy measured via beclin-1 protein level and light-chain 3 protein (LC3)-I into LC3-II conversion. Using POX-expressing cell model, we established that single POX overexpression was sufficient to activate autophagy. Thus, it led to autophagosomes accumulation and increased conversion of LC3-I into LC3-II. Moreover, beclin-1 gene expression was directly dependent on POX catalytic activity, namely the generation of POX-dependent superoxide. We conclude that POX is critical in the cellular response to the noxious effects of oxLDL by activating protective autophagy. PMID:19942609

  13. IL-6 Trans-signaling-STAT3 Pathway Mediates ECM and Cellular Proliferation in Fibroblasts from Hypertrophic Scar

    PubMed Central

    Ray, Sutapa; Ju, Xiaoxi; Sun, Hong; Finnerty, Celeste C; Herndon, David N; Brasier, Allan R

    2012-01-01

    The molecular mechanisms behind the pathogenesis of post-burn hypertrophic scar (HS) remain unclear. Here, we investigate the role of interleukin-6 (IL-6) trans-signaling-STAT3 pathway in HS fibroblasts (HSF) derived from burned-induced HS skin. HSF showed increased Tyr 705 STAT3 phosphorylation over normal fibroblast (NF) after IL-6•IL-6R? stimulation by immunoassays. The endogenous STAT3 target gene, SOCS3, was upregulated in HSF and showed increased STAT3 binding on its promoter relative to NF in Chromatin Immunoprecipitation assay. We observed that the cell surface signaling transducer glycoprotein 130 is upregulated in HSF using Q-RT-PCR and flow cytometry. The production of excessive extracellular matrix (ECM), including the expression of alpha2 (1) procollagen (Col1A2) and fibronectin 1 (FN) were seen in HSFs. A STAT3 peptide inhibitor abrogated FN and Col1A2 gene expression in HSF indicating involvement of STAT3 in ECM production. The cellular proliferation markers Cyclin D1, Bcl-Xl and c-Myc were also upregulated in HSF and knockdown of STAT3 by siRNA attenuated c-Myc expression indicating the essential role of STAT3 in fibroblast proliferation. Taken together, our results suggest that the IL-6-trans-signaling-STAT3 pathway may play an integral role in HS pathogenesis and disruption of this pathway could be a potential therapeutic strategy for the treatment of burn-induced HS. PMID:23303450

  14. Modification of erucic acid content in Indian mustard (Brassica juncea) by up-regulation and down-regulation of the Brassica juncea FAT TY ACID ELONGATION1 (BjFAE1) gene.

    PubMed

    Kanrar, S; Venkateswari, J; Dureja, P; Kirti, P B; Chopra, V L

    2006-03-01

    In Brassicas, the Fatty Acid Elongation1 (FAE1) gene product, a 3-ketoacyl-CoA synthase, is the first in a 4-enzyme complex involved in the synthesis of erucic acid from oleic acid. The FAE1 homologue from Brassica juncea cv. Pusa Bold was cloned in a binary vector both in sense and antisense orientations under the control of the CaMV35S promoter. The recombinant binary vectors were used to transform B. juncea cv. RLM 198 via Agrobacterium tumefaciens. The presence of the transgene was confirmed by polymerase chain reaction and Southern hybridization. Northern and western analyses showed the expression of the gene and protein, respectively, in the transgenic plants. Analyses of the fatty acid profile of the seed oil from homozygous T4 generation seeds revealed that over-expression of the FAE1 gene caused a 36% increase in the percent of erucic acid (37-49% compared to 36% in untransformed control). The down-regulation of FAE1 caused an 86% decrease in the percent of erucic acid to as low as 5% in the seed oil of transgenic plants. Thus, it is clearly possible to alter erucic acid content of mustard by altering the expression level of the FAE 1 gene. PMID:16322995

  15. Responses of plant seedlings to hypergravity: cellular and molecular aspects

    NASA Astrophysics Data System (ADS)

    Hoson, T.; Yoshioka, R.; Soga, K.; Wakabayashi, K.; Takeba, G.

    Hypergravity produced by centrifugation has been used to analyze the responses of plant seedlings to gravity stimulus. Elongation growth of stem organs is suppressed by hypergravity, which can be recognized as a way for plants to resist gravitational force. The mechanisms inducing growth suppression under hypergravity conditions were analyzed at cellular and molecular levels. When growth was suppressed by hypergravity, a decrease in the cell wall extensibility was brought about in various plants. Hypergravity also induced a cell wall thickening and an increase in the molecular mass of the certain hemicellulosic polysaccharides. Both a decrease in the activities hydrolyzing such polysaccharides and an increase in the apoplast pH were involved in such changes in the cell wall constituents. Thus, the cell wall metabolism is greatly modified under hypergravity conditions, which causes a decrease in the cell wall extensibility, thereby inhibiting elongation growth in stem organs. On the other hand, to identify genes involved in hypergravity-induced growth suppression, changes in gene expression by hypergravity treatment were analyzed in Arabidopsis hypocotyls by differential display method. Sixty-two genes were expressed differentially: expression levels of 39 genes increased, whereas those of 23 genes decreased under hypergravity conditions. The expression of these genes was further analyzed using RT-PCR. One of genes upregulated by hypergravity encoded hydroxymethylglutaryl-CoA reductase (HMGR), which catalyzes a reaction producing mevalonic acid, a key precursor of hormones such as gibberellic acid and abscisic acid. The expression of HMGR gene increased within several hours after hypergravity treatment. Also, compactin, an inhibitor of HMGR activity, prevented hypergravity-induced growth suppression, suggesting that HMGR is involved in suppression of Arabidopsis hypocotyl growth by hypergravity. In addition, hypergravity increased the expression levels of CCR1 and ERD15, which were shown to take part in the signaling pathway of environmental stimuli such as temperature and water. These cellular and molecular changes appear to be involved in a series of events leading to growth suppression of stem organs under hypergravity conditions.

  16. Global Gene Expression Profile for Swarming Bacillus cereus Bacteria?†

    PubMed Central

    Salvetti, Sara; Faegri, Karoline; Ghelardi, Emilia; Kolstø, Anne-Brit; Senesi, Sonia

    2011-01-01

    Bacillus cereus can use swarming to move over and colonize solid surfaces in different environments. This kind of motility is a collective behavior accompanied by the production of long and hyperflagellate swarm cells. In this study, the genome-wide transcriptional response of B. cereus ATCC 14579 during swarming was analyzed. Swarming was shown to trigger the differential expression (>2-fold change) of 118 genes. Downregulated genes included those required for basic cellular metabolism. In accordance with the hyperflagellate phenotype of the swarm cell, genes encoding flagellin were overexpressed. Some genes associated with K+ transport, phBC6A51 phage genes, and the binding component of the enterotoxin hemolysin BL (HBL) were also induced. Quantitative reverse transcription-PCR (qRT-PCR) experiments indicated an almost 2-fold upregulation of the entire hbl operon during swarming. Finally, BC1435 and BC1436, orthologs of liaI-liaH that are known to be involved in the resistance of Bacillus subtilis to daptomycin, were upregulated under swarming conditions. Accordingly, phenotypic assays showed reduced susceptibility of swarming B. cereus cells to daptomycin, and Pspac-induced hyper-expression of these genes in liquid medium highlighted the role of BC1435 and BC1436 in the response of B. cereus to daptomycin. PMID:21642396

  17. Sparse feature selection methods identify unexpected global cellular response to strontium-containing materials.

    PubMed

    Autefage, Hélène; Gentleman, Eileen; Littmann, Elena; Hedegaard, Martin A B; Von Erlach, Thomas; O'Donnell, Matthew; Burden, Frank R; Winkler, David A; Stevens, Molly M

    2015-04-01

    Despite the increasing sophistication of biomaterials design and functional characterization studies, little is known regarding cells' global response to biomaterials. Here, we combined nontargeted holistic biological and physical science techniques to evaluate how simple strontium ion incorporation within the well-described biomaterial 45S5 bioactive glass (BG) influences the global response of human mesenchymal stem cells. Our objective analyses of whole gene-expression profiles, confirmed by standard molecular biology techniques, revealed that strontium-substituted BG up-regulated the isoprenoid pathway, suggesting an influence on both sterol metabolite synthesis and protein prenylation processes. This up-regulation was accompanied by increases in cellular and membrane cholesterol and lipid raft contents as determined by Raman spectroscopy mapping and total internal reflection fluorescence microscopy analyses and by an increase in cellular content of phosphorylated myosin II light chain. Our unexpected findings of this strong metabolic pathway regulation as a response to biomaterial composition highlight the benefits of discovery-driven nonreductionist approaches to gain a deeper understanding of global cell-material interactions and suggest alternative research routes for evaluating biomaterials to improve their design. PMID:25831522

  18. Phosphorylation Drives an Apoptotic Protein to Activate Antiapoptotic Genes

    PubMed Central

    Halder, Umesh Chandra; Bhowmick, Rahul; Roy Mukherjee, Tapasi; Nayak, Mukti Kant; Chawla-Sarkar, Mamta

    2013-01-01

    During infection, viral proteins target cellular pathways that regulate cellular innate immune responses and cell death. We demonstrate that influenza A virus matrix 1 protein (M1), an established proapoptotic protein, activates nuclear factor-?B member RelB-mediated survival genes (cIAP1, cIAP2, and cFLIP), a function that is linked with its nuclear translocation during early infection. Death domain-associated protein 6 (Daxx) is a transcription co-repressor of the RelB-responsive gene promoters. During influenza virus infection M1 binds to and stabilizes Daxx protein by preventing its ubiquitination and proteasomal degradation. Binding of M1 with Daxx through its Daxx binding motif prevents binding of RelB and Daxx, resulting in up-regulation of survival genes. This interaction also prevents promoter recruitment of DNA methyltransferases (Dnmt1 and Dnmt3a) and lowers CpG methylation of the survival gene promoters, leading to the activation of these genes. Thus, M1 prevents repressional function of Daxx during infection, thereby exerting a survival role. In addition to its nuclear localization signal, translocation of M1 to the nucleus depends on cellular kinase-mediated phosphorylation as the protein kinase C inhibitor calphostin C effectively down-regulates virus replication. The study reconciles the ambiguity of dual antagonistic function of viral protein and potentiates a possible target to limit virus infection. PMID:23548901

  19. Gene Dose Influences Cellular and Calcium Channel Dysregulation in Heterozygous and Homozygous T4826I-RYR1 Malignant Hyperthermia-susceptible Muscle*

    PubMed Central

    Barrientos, Genaro C.; Feng, Wei; Truong, Kim; Matthaei, Klaus I.; Yang, Tianzhong; Allen, Paul D.; Lopez, José R.; Pessah, Isaac N.

    2012-01-01

    Malignant hyperthermia susceptibility (MHS) is primarily conferred by mutations within ryanodine receptor type 1 (RYR1). Here we address how the MHS mutation T4826I within the S4-S5 linker influences excitation-contraction coupling and resting myoplasmic Ca2+ concentration ([Ca2+]rest) in flexor digitorum brevis (FDB) and vastus lateralis prepared from heterozygous (Het) and homozygous (Hom) T4826I-RYR1 knock-in mice (Yuen, B. T., Boncompagni, S., Feng, W., Yang, T., Lopez, J. R., Matthaei, K. I., Goth, S. R., Protasi, F., Franzini-Armstrong, C., Allen, P. D., and Pessah, I. N. (2011) FASEB J. doi:22131268). FDB responses to electrical stimuli and acute halothane (0.1%, v/v) exposure showed a rank order of Hom ? Het ? WT. Release of Ca2+ from the sarcoplasmic reticulum and Ca2+ entry contributed to halothane-triggered increases in [Ca2+]rest in Hom FDBs and elicited pronounced Ca2+ oscillations in ?30% of FDBs tested. Genotype contributed significantly elevated [Ca2+]rest (Hom > Het > WT) measured in vivo using ion-selective microelectrodes. Het and Hom oxygen consumption rates measured in intact myotubes using the Seahorse Bioscience (Billerica, MA) flux analyzer and mitochondrial content measured with MitoTracker were lower than WT, whereas total cellular calpain activity was higher than WT. Muscle membranes did not differ in RYR1 expression nor in Ser2844 phosphorylation among the genotypes. Single channel analysis showed highly divergent gating behavior with Hom and WT favoring open and closed states, respectively, whereas Het exhibited heterogeneous gating behaviors. [3H]Ryanodine binding analysis revealed a gene dose influence on binding density and regulation by Ca2+, Mg2+, and temperature. Pronounced abnormalities inherent in T4826I-RYR1 channels confer MHS and promote basal disturbances of excitation-contraction coupling, [Ca2+]rest, and oxygen consumption rates. Considering that both Het and Hom T4826I-RYR1 mice are viable, the remarkable isolated single channel dysfunction mediated through this mutation in S4-S5 cytoplasmic linker must be highly regulated in vivo. PMID:22139840

  20. Overexpression of cellular repressor of E1A-stimulated genes inhibits TNF-{alpha}-induced apoptosis via NF-{kappa}B in mesenchymal stem cells

    SciTech Connect

    Peng, Cheng-Fei [Xijing Hospital, Fourth Military Medical University, Xi'an (China) [Xijing Hospital, Fourth Military Medical University, Xi'an (China); Cardiovascular Research Institute and Department of Cardiology, Shenyang Northern Hospital, Shenyang (China); Han, Ya-Ling, E-mail: hanyaling53@gmail.com [Cardiovascular Research Institute and Department of Cardiology, Shenyang Northern Hospital, Shenyang (China)] [Cardiovascular Research Institute and Department of Cardiology, Shenyang Northern Hospital, Shenyang (China); Jie-Deng,; Yan, Cheng-Hui; Jian-Kang,; Bo-Luan,; Jie-Li [Cardiovascular Research Institute and Department of Cardiology, Shenyang Northern Hospital, Shenyang (China)] [Cardiovascular Research Institute and Department of Cardiology, Shenyang Northern Hospital, Shenyang (China)

    2011-03-25

    Research highlights: {yields} CREG protected MSCs from tumor necrosis factor-{alpha} (TNF-{alpha}) induced apoptosis. {yields} CREG inhibits the phosphorylation of I{kappa}B{alpha} and prevents the activation of NF-{kappa}B. {yields} CREG inhibits NF-{kappa}B nuclear translocation and pro-apoptosis protein transcription. {yields} CREG anti-apoptotic effect involves inhibition of the death receptor pathway. {yields} p53 is downregulated by CREG via NF-{kappa}B pathway under TNF-{alpha} stimulation. -- Abstract: Bone marrow-derived mesenchymal stem cells (MSCs) show great potential for therapeutic repair after myocardial infarction. However, poor viability of transplanted MSCs in the ischemic heart has limited their use. Cellular repressor of E1A-stimulated genes (CREG) has been identified as a potent inhibitor of apoptosis. This study therefore aimed to determine if rat bone marrow MSCs transfected with CREG-were able to effectively resist apoptosis induced by inflammatory mediators, and to demonstrate the mechanism of CREG action. Apoptosis was determined by flow cytometric and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling assays. The pathways mediating these apoptotic effects were investigated by Western blotting. Overexpression of CREG markedly protected MSCs from tumor necrosis factor-{alpha} (TNF-{alpha}) induced apoptosis by 50% after 10 h, through inhibition of the death-receptor-mediated apoptotic pathway, leading to attenuation of caspase-8 and caspase-3. Moreover, CREG resisted the serine phosphorylation of I{kappa}B{alpha} and prevented the nuclear translocation of the transcription factor nuclear factor-{kappa}B (NF-{kappa}B) under TNF-{alpha} stimulation. Treatment of cells with the NF-{kappa}B inhibitor pyrrolidine dithiocarbamate (PDTC) significantly increased the transcription of pro-apoptosis proteins (p53 and Fas) by NF-{kappa}B, and attenuated the anti-apoptotic effects of CREG on MSCs. The results of this study indicate that CREG acts as a novel and potent survival factor in MSCs, and may therefore be a useful therapeutic adjunct for transplanting MSCs into the damaged heart after myocardial infarction.

  1. Bitter Gourd (Momordica charantia) Extract Activates Peroxisome Proliferator-Activated Receptors and Upregulates the Expression of the Acyl CoA Oxidase Gene in H4IIEC3 Hepatoma Cells

    Microsoft Academic Search

    Che-Yi Chao; Ching-jang Huang

    2003-01-01

    Peroxisome proliferator-activated receptor ? (PPAR?) is a ligand-dependent transcription factor that regulates the expression of genes involved in lipid metabolism and transport. Ligands\\/activators of PPAR?, like fibrate-type drugs, may have hypolipidemic effects. To identify food that contains activators of PPAR?, a transactivation assay employing a clone of CHO-K1 cells stably transfected with a (UAS)4-tk-alkaline phosphatase reporter and a chimeric receptor

  2. Comparison of gene expression signatures of diamide, H2O2 and menadione exposed Aspergillus nidulans cultures – linking genome-wide transcriptional changes to cellular physiology

    Microsoft Academic Search

    István Pócsi; Márton Miskei; Zsolt Karányi; Tamás Emri; Patricia Ayoubi; Tünde Pusztahelyi; György Balla; Rolf A Prade

    2005-01-01

    BACKGROUND: In addition to their cytotoxic nature, reactive oxygen species (ROS) are also signal molecules in diverse cellular processes in eukaryotic organisms. Linking genome-wide transcriptional changes to cellular physiology in oxidative stress-exposed Aspergillus nidulans cultures provides the opportunity to estimate the sizes of peroxide (O22-), superoxide (O2•-) and glutathione\\/glutathione disulphide (GSH\\/GSSG) redox imbalance responses. RESULTS: Genome-wide transcriptional changes triggered by

  3. E2F7, a novel target, is up-regulated by p53 and mediates DNA damage-dependent transcriptional repression

    PubMed Central

    Carvajal, Luis A.; Hamard, Pierre-Jacques; Tonnessen, Crystal; Manfredi, James J.

    2012-01-01

    The p53 tumor suppressor protein is a transcription factor that exerts its effects on the cell cycle via regulation of gene expression. Although the mechanism of p53-dependent transcriptional activation has been well-studied, the molecular basis for p53-mediated repression has been elusive. The E2F family of transcription factors has been implicated in regulation of cell cycle-related genes, with E2F6, E2F7, and E2F8 playing key roles in repression. In response to cellular DNA damage, E2F7, but not E2F6 or E2F8, is up-regulated in a p53-dependent manner, with p53 being sufficient to increase expression of E2F7. Indeed, p53 occupies the promoter of the E2F7 gene after genotoxic stress, consistent with E2F7 being a novel p53 target. Ablation of E2F7 expression abrogates p53-dependent repression of a subset of its targets, including E2F1 and DHFR, in response to DNA damage. Furthermore, E2F7 occupancy of the E2F1 and DHFR promoters is detected, and expression of E2F7 is sufficient to inhibit cell proliferation. Taken together, these results show that p53-dependent transcriptional up-regulation of its target, E2F7, leads to repression of relevant gene expression. In turn, this E2F7-dependent mechanism contributes to p53-dependent cell cycle arrest in response to DNA damage. PMID:22802528

  4. Mining the bladder cancer-associated genes by an integrated strategy for the construction and analysis of differential co-expression networks

    PubMed Central

    2015-01-01

    Background Bladder cancer is the most common malignant tumor of the urinary system and it is a heterogeneous disease with both superficial and invasive growth. However, its aetiological agent is still unclear. And it is indispensable to find key genes or modules causing the bladder cancer. Based on gene expression microarray datasets, constructing differential co-expression networks (DCNs) is an important method to investigate diseases and there have been some relevant good tools such as R package 'WGCNA', 'DCGL'. Results Employing an integrated strategy, 36 up-regulated differentially expressed genes (DEGs) and 356 down-regulated DEGs were selected and main functions of those DEGs are cellular physiological precess(24 up-regulated DEGs; 167 down-regulated DEGs) and cellular metabolism (19 up-regulated DEGs; 104 down-regulated DEGs). The up-regulated DEGs are mainly involved in the the pathways related to "metabolism". By comparing two DCNs between the normal and cancer states, we found some great changes in hub genes and topological structure, which suggest that the modules of two different DCNs change a lot. Especially, we screened some hub genes of a differential subnetwork between the normal and the cancer states and then do bioinformatics analysis for them. Conclusions Through constructing and analyzing two differential co-expression networks at different states using the screened DEGs, we found some hub genes associated with the bladder cancer. The results of the bioinformatics analysis for those hub genes will support the biological experiments and the further treatment of the bladder cancer. PMID:25707808

  5. Antioxidant pathways are up-regulated during biological nitrogen fixation to prevent ROS-induced nitrogenase inhibition in Gluconacetobacter diazotrophicus.

    PubMed

    Alquéres, Sylvia M C; Oliveira, Jose Henrique M; Nogueira, Eduardo M; Guedes, Helma V; Oliveira, Pedro L; Câmara, Fernando; Baldani, Jose I; Martins, Orlando B

    2010-10-01

    Gluconacetobacter diazotrophicus, an endophyte isolated from sugarcane, is a strict aerobe that fixates N(2). This process is catalyzed by nitrogenase and requires copious amounts of ATP. Nitrogenase activity is extremely sensitive to inhibition by oxygen and reactive oxygen species (ROS). However, the elevated oxidative metabolic rates required to sustain biological nitrogen fixation (BNF) may favor an increased production of ROS. Here, we explored this paradox and observed that ROS levels are, in fact, decreased in nitrogen-fixing cells due to the up-regulation of transcript levels of six ROS-detoxifying genes. A cluster analyses based on common expression patterns revealed the existence of a stable cluster with 99.8% similarity made up of the genes encoding the ?-subunit of nitrogenase Mo-Fe protein (nifD), superoxide dismutase (sodA) and catalase type E (katE). Finally, nitrogenase activity was inhibited in a dose-dependent manner by paraquat, a redox cycler that increases cellular ROS levels. Our data revealed that ROS can strongly inhibit nitrogenase activity, and G. diazotrophicus alters its redox metabolism during BNF by increasing antioxidant transcript levels resulting in a lower ROS generation. We suggest that careful controlled ROS production during this critical phase is an adaptive mechanism to allow nitrogen fixation. PMID:20697694

  6. Expression of osteoblast and osteoclast regulatory genes in the bone marrow microenvironment in multiple myeloma: only up-regulation of Wnt inhibitors SFRP3 and DKK1 is associated with lytic bone disease.

    PubMed

    Kristensen, Ida B; Christensen, Jacob Haaber; Lyng, Maria B; Møller, Michael B; Pedersen, Lise; Rasmussen, Lars M; Ditzel, Henrik J; Abildgaard, Niels

    2014-04-01

    Multiple myeloma (MM) lytic bone disease (LBD) is caused by osteoclast activation and osteoblast inhibition. RANK/RANKL/OPG play central roles in osteoclast activation and Wnt inhibitor DKK1 in osteoblast inhibition. The role of other Wnt inhibitors is less clear. We evaluated gene expression of osteoclast regulators (RANK, RANKL, OPG, TRAIL, MIP1A), Wnt inhibitors (DKK1, SFRP2, SFRP3, sclerostin, WIF1) and osteoblast transcription factors (RUNX2, osterix) by quantitative reverse transcriptase polymerase chain reaction (RT-PCR) in the bone marrow (BM) microenvironment using snap-frozen BM biopsies, thereby achieving minimal post-sampling manipulation, and gene expression profiling (GEP) data, reflecting the in vivo situation. We analyzed 110 biopsies from newly diagnosed patients with MM and monoclonal gammopathy of unknown significance (MGUS) and healthy volunteers. LBD was evaluated using standard radiographs and the bone resorption marker CTX-1. Protein levels were evaluated by enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry. Among Wnt inhibitors, only SFRP3 and DKK1 were significantly overexpressed in advanced LBD, correlating with protein levels. SFRP3 correlated with CTX-1. Our findings support osteoblast inhibition as the driving force behind MM LBD. PMID:23915193